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Journal of Neuropathology & Experimental Neurology:
doi: 10.1097/01.jnen.0000446983.27392.a4
Abstracts

American Association of Neuropathologists, Inc., Abstracts of the 90th Annual Meeting, June 12–15, 2014, Portland, Oregon

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PLATFORM SESSION 1: TUMORS 1 - PREDICTIVE GLIOMA MARKERS Friday 8-10 AM, June 13, 2014 Fashion Ballroom

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1

Predicting the Likelihood of an IDH1 or IDH2 Mutation in Patients Diagnosed with Infiltrative Gliomas

Craig Horbinski1, Zoya Voronovich2, Kenneth Clark2, Isaac Hands1, Jonathan Mannas1, Meggen Walsh1, Marina Nikiforova2, Eric Durbin1, Heidi Weiss1, Li Chen1. 1University of Kentucky; 2University of Pittsburgh

Background: Several variables are associated with isocitrate dehydrogenase 1 or 2 (IDH1/2) mutation likelihood in gliomas, though no guidelines yet exist for when testing is warranted, especially when an R132H IDH1 immunostain is negative.

Methods: A cohort of 89 patients was used to build IDH1/2 mutation prediction models in WHO grade II-IV gliomas, and an external cohort of 100 patients was used for validation. Logistic regression and backward model selection with Akaike information criterion were used to develop prediction models. A third cohort from The Cancer Genome Atlas (TCGA) was also used for additional validation.

Results: A multivariate model, incorporating patient age, glioblastoma (GBM) diagnosis, and prior history of grade II or III glioma, was developed to predict IDH1/2 mutation probability. This model generated an area under the curve (AUC) of 0.934 (95% CI: 0.878, 0.978) in the external validation cohort and 0.941 (95% CI: 0.918, 0.962) in the TCGA cohort. When R132H IDH1 immunostain information was added, AUC increased to 0.986 (95% CI: 0.967, 0.998). This model had an AUC of 0.947 (95% CI: 0.891, 0.995) in predicting whether an R132H IDH1 immunonegative case harbored a less common IDH1 or IDH2 mutation. Its ability to predict IDH1/2 mutation status in grades II-IV gliomas from the Cancer Genome Atlas (94%) was superior to the 84% accuracy of a board-certified neuropathologist who evaluated the same cohort. An interactive web-based application for calculating the probability of an IDH1/2 mutation is now available using these models ( http://www.kcr.uky.edu/webapps/IDH/app.html).

Conclusions: We have integrated multiple variables to generate a probability of an IDH1/2 mutation. The associated web-based application can help triage diffuse gliomas that would benefit from mutation testing in both clinical and research settings.

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2

Glutamine Based PET Imaging Facilitates Enhanced Metabolic Detection of Gliomas in Vivo

Sriram Venneti1, Mark Dunphy1, Hanwen Zhang1, Kenneth Pitter1, Carl Campos1, Sean Carlin1, Serge Lyashchenko1, Karl Plöessl2, Daniel Rohle1, Antonio Omuro1, Justin Cross1, Cameron Brennan1, Wolfgang Weber1, Eric Holland3, Ingo Mellinghoff1, Hank Kung2, Jason Lewis1, Craig Thompson1. 1Memorial Sloan-Kettering Cancer Center; 2University of Pennsylvania; 3University of Washington

Glutamine is the most abundant plasma amino acid and many cancers show altered glutamine metabolism to support their growth and proliferation. We evaluated glutamine metabolism in gliomas using biochemical and in vivo PET imaging approaches. Our results demonstrate that glutamine is a key TCA cycle anaplerotic substrate in gliomas and that glutamine is metabolized to generate 2-HG in IDH1 mutant gliomas. To begin to understand glutamine metabolism in vivo we used PET imaging with18F-labeled glutamine (18F-FGln), which showed high uptake in gliomas but low background uptake in the surrounding brain in RCAS-PDGF/PTEN null and IDH1 mutant glioma animal models. This facilitated clear tumor delineation in contrast to that seen with 18F-FDG. We did not observe 18F-FGln uptake in animals with neuroinflammation with a disrupted BBB. Further, 18F-FGln uptake was specifically reduced on chemo/radiation therapy corresponding with a histologic decrease in tumor volume. Finally, 18F-FGln showed high avidity in a human glioma patient with low uptake in the surrounding brain. These data suggest that 18F-FGln is specifically taken up by gliomas, can be used to detect the metabolic state of gliomas in vivo and may serve as a valuable tool in the clinical management of gliomas.

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3

Simplified Grading System for WHO Grade II-III Gliomas based on 1p/19q Status, IDH 1/2 Mutation Status, and Nestin Expression

Kimmo Hatanpaa1, Vamsidhara Vemireddy1, Tianshen Hu1, Jack Raisanen1, Chan Foong1, Bruce Mickey1, Samuel Barnett1, Dwight Oliver1, Paul Yell1, Edward Pan1, Dennis Burns1, Charles White1, Elizabeth Maher1, Robert Bachoo1. 1UT Southwestern Medical Center

Infiltrating astrocytomas and oligoastrocytomas of low to anaplastic grade (WHO grades II-III), in spite of being associated with a wide range of clinical outcomes, are often difficult to subclassify and grade. Unlike oligodendrogliomas and anaplastic oligodendrogliomas, which can be identified by the presence of 1p/19q LOH, and glioblastomas, which are diagnosed based on objective features on routine hematoxylin and eosin sections, the WHO criteria for the subclassification of grade II-III astrocytomas and oligoastrocytomas (A+OA II-III) are not as clearly defined. The current criteria depend on features such as nuclear roundness and significant mitotic activity, leaving much room for subjective interpretation in individual cases.

We recently identified nestin, a protein expressed by stem cells, as a strong prognostic marker in human A+OA II-III tumors (p=0.0004; n=50) (Hatanpaa et al., J Neurooncol 117:183-9, 2014). Both nestin protein and mRNA level correlated with total survival. In multivariate survival analysis, nestin and IDH 1/2 mutation status remained highly significant, whereas histological covariates failed to reach significance (WHO grade II vs. III, presence of oligodendroglial component, MIB-1 index). All tumors were negative for 1p/19q LOH by PCR and EGFR amplification by CISH.

Here, we show that by combining information on nestin positivity, evaluated by immunohistochemistry, and IDH 1/2 mutation status, evaluated by immunohistochemistry for the IDH1 R132H mutation and followed by IDH 1/2 sequencing of the cases that are negative, A+OA II-III tumors can be divided into three groups with markedly different clinical outcomes (p<0.0001): (1) nestin negative and IDH mutated (both favorable), (2) nestin negative and IDHwt or nestin positive and IDH mutated (mixed favorable/unfavorable), and (3) nestin positive and IDHwt (both unfavorable). The total survival was favorable and similar to 1p/19q codeleted oligodendrogliomas for group #1, intermediate for group #2, and dismal and similar to glioblastomas (median survival, 1.5 years) for group #3.

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4

Loss of CDKN2A/p16 is Associated with Shortened Overall Survival in Grade II and III Gliomas

Gerald Reis1, Melike Pekmezci1, Helen Hansen1, Roxanne Marshall1, Terri Rice1, Joanna Phillips1, John Wiencke1, Margaret Wrensch1, Kyle Walsh1, Arie Perry1. 1University of California San Francisco

Background: Lower WHO grade (II and III) gliomas (LGG) vary widely in clinical behavior and are classified as astrocytoma (A2), oligodendroglioma (O2), and oligoastrocytoma (OA2) or anaplastic astrocytoma (A3), oligodendroglioma (O3), and oligoastrocytoma (OA3). Anaplasia depends greatly on mitotic activity with CDKN2A loss considered the most common mechanism for cell cycle dysregulation. Thus, we investigated whether loss of the CDKN2A gene or its product, p16, is associated with patient survival across pathologically and genetically defined subtypes.

Methods: UCSF Adult Glioma Study cases (95 males and 70 females; average age: 43.2 and 41.1 years, respectively) consisted of 57 O2, 34 A2, 29 OA2, 20 O3, 16 A3, 9 OA3. Molecular parameters included isocitrate dehydrogenase (IDH1 or IDH2) (97%) and 1p19q status (88% of oligodendroglial tumors). CDKN2A deletion was assessed by FISH, while p16 and MIB1 (Ki-67) labeling indices (LI) were determined immunohistochemically. Cox proportional hazards modeled overall survival, stratifying by histology and adjusting for sex, age, grade, and molecular subtype. 66 events (deaths) were observed with median follow-up time of 6.4 years.

Results: Reduced p16 LI was associated with decreased overall survival across all grades (P=0.05) and adjusted for age, histology, and IDH status (P=0.02). CDKN2A deletion was strongly associated with worse survival in astrocytomas (P=0.002; HR=4.4; 95% CI=1.73-11.0) but not oligodendrogliomas or oligoastrocytomas (P=0.60 and 0.96, respectively). IDH mutation and CDKN2A deletion were inversely associated across all subtypes (p=0.03). CDKN2A FISH and p16 IHC results were only weakly associated (P=0.10), while no clear associations between CDKN2A and MIB1 (P=0.21) or p16 and MIB1 (P=0.21) were found.

Conclusions: Loss of p16 expression is associated with shortened survival in all LGG, whereas CDKN2A deletions decrease survival in astrocytomas, most being IDH wild type. CDKN2A/p16 studies may provide further clinical aid in LGG sub-stratification beyond IDH and 1p19q studies.

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5

Biopsy Site has Important Prognostic Implications in Gliomas

Rocio Guevara de Bonis1, Marta Brell1, Javier Ibanez1, Carmen Vidal1, Francisca Nebot1, Marta Couce2. 1Hospital Universitari Son Espases, Palma, Spain; 2University Hospitals Case Medical Center

Predictive biomarkers are part of the diagnostic tests battery in gliomas. Some patients, in spite of bearing predictors of poor behavior will have longer survivals; whereas others, with good prognostic markers, will have shorter life expectancies. Recently, the generally accepted good predictive value of O6-methylguanine-DNA methyltransferase promoter methylation (MGMTpm), was questioned. In addition, one study reported that only those patients with wild type isocitrate dehydrogenase 1 (IDH1wt) would benefit from that prediction. Biochemical and molecular characterization of distant regions of gliomas could be key to understand these different clinical responses.

44 patients undergoing gross total resection of gliomas were consented for the study (11 low-grade gliomas (LG), and 33 high-grade gliomas (HG)) and followed from 36 to 85 months. Central and peripheral tumoral regions were examined. IDH1 mutation (IDH1mut) (by immunohistochemistry and sequencing analysis), IDH1 mRNA expression and MGMTpm were evaluated.

1) IDH1 mutation was present in 72.7 % LG and 18.2 % HG, and showed no regional differences. 2) IDH1 mutation was associated with longer survival, particularly in HG (p<0.05). 3) HG, had significantly higher expression of IDH1mRNA as compared with LG (p<0.05). 4) A positive correlation between IDH1 mutation and MGMTpm was identified (p<0.01). 5) MGMT was more frequently methylated in central regions in both types of gliomas (29.4% HG and 42.9 % LG were methylated in central and not in peripheral regions). 6) Chemotherapy was beneficial for those patients with concomitant MGMTpm, and IDH1mut.

We conclude that the combination of IDH1mut and MGMTpm predict the best possible outcome for these patients. MGMTpm alone does not increase the survival in IDH1wt gliomas in our study. In addition, we found that biopsies from central regions in gliomas are more informative in order to accurately characterize the MGMTpm in these tumors.

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6

Is hTERT Immunohistochemistry Predictive of 1p/19q Co-deletion in Grade II-III Diffuse Gliomas?

Christina Appin1, Daniel Brat1. 1Emory University School of Medicine

Background: One mechanism for maintaining telomere length in diffuse gliomas is by activating somatic mutation of the human telomerase reverse transcriptase (hTERT) promoter. hTERT promoter mutations are associated with increased gene expression and are frequently present in IDH-mutated, 1p/19q co-deleted and in IDH-wildtype diffuse gliomas, but rarely in those that are IDH-mutated, 1p/19q intact. We investigated whether the immunohistochemical (IHC) expression of hTERT was predictive of 1p/19q co-deletion among IDH-mutated diffuse gliomas.

Methods: IHC for hTERT and IDH1 mutant protein (R132H) was performed on 43 grade II-III diffuse gliomas. Fluorescence in situ hybridization (FISH) was performed to assess 1p and 19q status. Diffuse gliomas immunostained for hTERT included 20 IDH1-mutated, 1p/19q co-deleted tumors; 15 IDH1-mutated, 1p/19q intact tumors and 8 IDH1-wildtype, 1p/19q intact tumors. Statistical analysis of correlations between alterations was performed using Chi-square analysis.

Results: IHC expression of hTERT was detected in 15 of 20 (75%) IDH1-mutated, 1p/19q co-deleted gliomas; 6 of 8 (75%) IDH1-wildtype gliomas; and 6 of 15 (40%) IDH1-mutated, 1p/19q intact gliomas. Among the IDH1-mutated gliomas, we found a significant correlation between hTERT expression and 1p/19q co-deletion (p = 0.0364). However, IHC expression of hTERT was only 75% sensitive and 60% specific for 1p/19q co-deletion within IDH1-mutated gliomas. The positive predictive value of hTERT IHC for 1p/19q co-deletion was 71% and the negative predictive value was 64%.

Conclusions: hTERT IHC expression was noted in 75% of IDH1-mutated, 1p/19q co-deleted tumors and 75% of IDH1 wildtype, 1p/19q intact tumors. However, the frequency of hTERT IHC expression in IDH1-mutated, 1p/19q intact gliomas (40%) suggests that this test is not highly specific or predictive for 1p/19q co-deletion.

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7

TERT Promoter Mutation is Associated with Older Age at Diagnosis, Independent of Glioma Grade, Histology and IDH1/2 Status

Melike Pekmezci1, Gerald Reis1, Helen Hansen1, Terri Rice1, Shichun Zheng1, John Wiencke1, Arie Perry1, Margaret Wrensch1, Kyle Walsh1. 1University of California San Francisco

Background: Young age at diagnosis is a favorable prognostic factor and impacts clinical management of patients with infiltrating glioma. The literature suggests that grade II tumors and oligodendroglial tumors are diagnosed at younger ages than anaplastic and astrocytic tumors, respectively. With the exception of isocitrate dehydrogenase (IDH) mutations, molecular features driving these age differences remain largely unknown. Heritable variation in the telomerase reverse transcriptase (TERT) gene is associated with later age at diagnosis in glioma patients. We sought to evaluate whether somatic TERT mutations are also associated with later age at diagnosis in glioma patients, within strata of grade, histology and IDH1/2 status.

Methods: Adults (age ≥18 years) with histopathologically confirmed infiltrating glioma were selected from the UCSF Adult Glioma Study (n=351). The TERT promoter region was analyzed by Sanger sequencing and tumors were classified as TERT-wild type and TERT-mutated. Associations between TERT status and age at diagnosis were assessed by t-tests.

Results: TERT promoter mutation was associated with an approximately 8 year older age at diagnosis across all glioma grades (gradeIV p=0.0003, GradeIII p=0.014, GradeII p=0.0001) and histologies (Glioblastoma p=0.0003, Astrocytoma p=0.007, Oligodedroglioma p=0.16, Mixed p=0.005). Among patients with IDH wild-type grade II-III gliomas, TERT promoter mutation was associated with a 10 year older age at diagnosis (p=0.0065). Among patients with IDH-mutated grade II-III gliomas, TERT promoter mutation was associated with a 7 year older age at diagnosis (p=0.0002). Overall, patients with IDH mutation who lack TERT promoter mutation have the youngest age at diagnosis (Average=36.5). This group consists primarily of astrocytic neoplasms.

Conclusions: Somatic mutations in IDH are associated with younger age at diagnosis, while somatic mutations in TERT promoter are associated with older age at diagnosis. Among patients with IDH mutated tumors, those with TERT mutation (primarily oligodendroglioma) are diagnosed at later ages than those without TERT mutation (primarily astrocytoma).

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8

Efficacy of Mono and Dual PI3K and MAPK Inhibition in Glioblastoma and Triple-Negative Breast Cancer Brain Metastasis Models

Robert McNeill1, Amanda Van Swearingen1, Ryan Bash1, Salma Azam1, Demitra Canoutas1, Brian Constance1, Ralf Schmid1, Carey Anders1, C. Miller1. 1University of North Carolina Chapel Hill

Glioblastoma (GBM) and triple-negative breast cancer brain metastases (BCBM) are biologically aggressive with limited therapeutic options. The MAPK and PI3K arms of receptor tyrosine kinase (RTK) signaling are attractive therapeutic targets because they are mutationally activated in both diseases. We defined the in vitro and in vivo efficacy of brain penetrant pan-PI3K (BKM120) and MEK1/2 (AZD6244) inhibitors in GBM and BCBM models. GBM was modeled using G1/S-defective murine (TRP) and human (NHA-RAS-AKT) astrocytes with genetically activated PI3K and MAPK signaling. BCBM was modeled using SUM149, an established, basal-like, BRCA1-mutant human cell line. BKM and AZD inhibited in vitro growth of both TRP and NHA-RAS-AKT astrocytes by >95 and ∼75% at 1-100 μM. BKM inhibited downstream PI3K signaling (decreased p-Akt and p-S6), while AZD6244 inhibited MAPK signaling (decreased p-Erk). Compensatory activation of the non-targeted pathway was evident in TRP astrocytes treated with either drug alone. Dual inhibition was required to ablate compensatory signaling and was synergistic at ∼4-110 μM in vitro. Daily treatment of established (10 d) orthotopic TRP GBM allografts in syngeneic mice with BKM (30 mpk), AZD (37 mpk), or both (25 and 18 mpk) did not delay growth by bioluminescence imaging nor provide a significant survival benefit (96, 108, and 84% of untreated animals). In contrast, while BKM and AZD mono-therapy inhibited SUM149 cell growth similar to GBM astrocytes, drug synergism was evident at ∼0.08-10 μM and the same treatment regimens delayed growth of established (20 d) intracranial SUM149 xenografts and prolonged median survival (116, 160, and >178% of untreated animals). Thus, despite similar in vitro efficacy in both model systems, dual BKM and AZD therapy was less synergistic and efficacious in the GBM vs. BCBM models tested here. These results suggest that GBM and BCBM may have unique, intrinsic resistance mechanisms to PI3K and MAPK inhibitors.

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PLATFORM SESSION 2: NEURODEGENERATIVE 1 - AD, CTE, PRIONS Friday 8-10 AM, June 13, 2014 Culture Ballroom

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9

Pyroglutamylated Amyloid-β correlates with Hyperphosphorylated Tau and Severity of Alzheimer’s Disease

Johannes Attems1, Lauren Walker2, Radmila Santic3, Ajeet Rijal Upadhaya4, Sean Colloby1, Dietmar Thal4, Alan Thomas1, Achim Schneeberger3, Markus Mandler3. 1Institute for Ageing and Health, Newcastle University; 2Newcastle University; 3Affiris AG; 4Laboratory of Neuropathology, University of Ulm

Pyroglutamylated amyloid-β has been suggested to play a major role in Alzheimer’s disease pathogenesis as amyloid-β oligomers containing pyroglutamylated amyloid-β might initiate tau dependent cytotoxicity. We aimed to further elucidate the associations between pyroglutamylated amyloid-β, amyloid-β and hyperphosphorylated tau in human brain tissue. We examined 41 post mortem brains (mean age 79.6 years, SE: ±1.5) of both Alzheimer’s disease (n=18) and controls. Adjacent sections from each frontal and entorhinal cortex, were stained with pyroglutamylated amyloid-β, hyperphosphorylated tau and amyloid-β specific antibodies. We used image analysis to quantitatively assess loads for pyroglutamylated amyloid-β, hyperphosphorylated tau and non-pyroglutamylated amyloid-β. All loads were significantly higher in Alzheimer’s disease as compared to controls (p≤0.01). However, regression analysis using amyloid-β loads as independent variables revealed that only frontal pyroglutamylated amyloid-β load independently predicted the presence of Alzheimer’s disease (p=0.01). In frontal and entorhinal cortices pyroglutamylated amyloid-β load independently predicted hyperphosphorylated tau load (P<0.001), while non-pyroglutamylated amyloid-β failed to do so. All loads correlated with neurofibrillary tangle Braak stages and Thal amyloid-β phases (P<0.01). However, partial correlation analysis revealed respective correlations in the frontal cortex only for pyroglutamylated amyloid-β load while in the entorhinal cortex respective correlations were seen for both hyperphosphorylated tau and non-pyroglutamylated amyloid-β loads. Mini Mental State Examination scores were predicted by entorhinal hyperphosphorylated tau load independent of other entorhinal loads (P<0.001) and by frontal pyroglutamylated amyloid-β load independent of frontal non-pyroglutamylated amyloid-β load (P=0.01). Here, we report an association between pyroglutamylated amyloid-β and hyperphosphorylated tau in human brain tissue and an influence of frontal pyroglutamylated amyloid-β on both the severity of Alzheimer’s disease neuropathology and clinical dementia. Our findings further support the notion that pyroglutamylated amyloid-β may represent an important link between amyloid-β and hyperphosphorylated tau and investigations into its role as diagnostic and therapeutic target in Alzheimer’s disease are warranted.

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10

TDP-43 Influences Cognition, Memory Loss and Hippocampal Atrophy in Alzheimer’s Disease

Keith Josephs1, Jennifer Whitwell2, Stephen Weigand3, Melissa Murray4, Nirubol Tosakulwong3, Amanda Liesinger4, Leonard Petrucelli5, David Knopman1, Bradley Boeve1, Robert Ivnik6, Glenn Smith6, Clifford Jack2, Joseph Parisi7, Ronald Petersen1, Dennis Dickson4. 1Mayo Clinic, Department of Neurology; 2Mayo Clinic, Department of Radiology; 3Mayo Clinic, Department of Health Science Research (Biostatistics); 4Mayo Clinic, Department of Neuroscience (Neuropathology); 5Mayo Clinic, Department of Neuroscience (Molecular Neuroscience); 6Mayo Clinic, Department of Psychiatry (Neuropsychology); 7Mayo Clinic, Department of Laboratory Medicine and Pathology

The aim of this study was to determine whether the TAR DNA-binding protein of 43kDa(TDP-43) has any independent effect on the clinical and neuroimaging features typically ascribed to Alzheimer’s disease (AD) pathology, and whether TDP-43 pathology could help to shed light on the phenomenon of resilient cognition that has been observed in AD. Three hundred and forty-two subjects pathologically diagnosed with intermediate-high probability AD were screened for the presence and distribution of TDP-43. In addition, we quantitatively measured the burden of TDP-43 in the dentate gyrus of the hippocampus. All subjects had been classified as cognitively impaired or cognitively normal, prior to death. Atlas-based parcellation and voxel-based morphometry were used to assess regional atrophy on MRI. Regression models controlling for age at death, apolipoprotein ε4, Braak stage, CERAD, Lewy bodies and vascular pathologies were utilized to explore effects between TDP-43 and cognition or brain atrophy, stratified by Braak stage. Additionally, we determined whether the effects of TDP-43 on outcome variables were mediated by hippocampal sclerosis. One-hundred and ninety-five (57%) cases were TDP-positive. After accounting for age at death, apolipoprotein ε4, and the other AD related pathologies, TDP-43 had a strong effect on cognition, memory loss, and medial temporal atrophy in AD. These effects were not mediated by hippocampal sclerosis. The TDP-positive subjects were 10X more likely to be cognitively impaired at death compared to the TDP-negative subjects. Greater cognitive impairment and medial temporal atrophy were associated with greater TDP-43 burden and more extensive TDP-43 distribution. TDP-43 is an important factor in the manifestation of the clinico-imaging features of AD and appears to be able to overpower what has been termed resilient brain aging. TDP-43 therefore should be considered a potential therapeutic target for the treatment of AD.

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11

Novel Fluid Biomarkers for Brain Amyloid in Presymptomatic Alzheimer Disease

Richard Perrin1, Hua Weng2, Kelley Coalier1, Anne Fagan1, Chengjie Xiong1, David Holtzman1. 1Washington University School of Medicine; 2Division of Biostatistics, Washington University School of Medicine

Background: Clinical trials for Alzheimer disease (AD) are now targeting Aβ production or clearance to combat amyloid deposition. In this circumstance, low CSF Aβ42 may serve as an indicator of target engagement, but will lose its utility as an independent surrogate marker for brain amyloid. PET ligands Pittsburgh compound B (PIB) and florbetapir (Amyvid) may still be effective for detecting plaques, but PIB is inconvenient to use at distance from a cyclotron, florbetapir may have limited sensitivity for small changes in plaque burden, and both are expensive. In this study, we sought to identify novel fluid biomarkers for very early amyloid deposition.

Methods: Paired CSF and plasma samples, collected from 142 Knight ADRC participants with normal cognition (Clinical Dementia Rating [CDR] 0) within 2.5 years of PET-PIB imaging, were analyzed using the 190MAP biomarker discovery panel at Myriad/RBM (Luminex platform). Mean cortical PIB binding potential [MCBP] above/below 0.18 was treated as a categorical variable for receiver operating characteristic (ROC) analyses. Logistic regression models were used to evaluate markers in combination. Age, gender, education, and APOE-ɛ4 status were treated as covariates.

Results: In multivariate ROC analyses, 22 plasma and 17 novel CSF proteins yielded areas under the curve (AUC) > 0.6 (max 0.662 & 0.687) for predicting MCBP >0.18. CSF tau/Aβ42 ratio combined with p-tau181 was the best predictor, with AUC = 0.963. As an alternative (e.g. for trials targeting Aβ42), a combination of CSF tau, α-1-AT and adiponectin yielded an AUC = 0.797. In plasma, a combination of apoA2, apoE, and PAI-1 yielded an AUC of 0.748.

Conclusions/Discussion: Many novel fluid proteins have potential as early brain amyloid biomarkers. These may facilitate clinical trials that directly target Aβ, aid screening for preclinical AD, and provide insights into the pathophysiology surrounding very early plaque deposition.

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12

Beclin1/BECN1 Interacts with Surface APP and Facilitates Its Internalization and Sorting for Autophagosomal Degradation

Edward Plowey1, Wan Zhu1, Gayathri Swaminathan1. 1Stanford University School of Medicine

Macroautophagy, hereafter designated autophagy, is a vesicular trafficking pathway for autolysosomal degradation that is thought to protect neurons from diverse stressors. Recent studies suggest that deficiencies in the BECN1-PIK3C3 complex, an essential autophagy regulatory protein complex, may be an important factor in age-related neurodegenerative diseases including Alzheimer disease (AD). Studies in BECN1+/- mice and cell line models have highlighted an important role for BECN1 in the regulation of amyloid precursor protein (APP) metabolism and in reducing the secretion of its toxic metabolite amyloid-beta. A better understanding of the cellular and molecular mechanisms underlying BECN1- mediated APP degradation is crucial for the design of novel therapeutic strategies to reduce amyloid-beta secretion and toxicity in AD. We hypothesized that BECN1 interacts with APP and promotes its trafficking for autolysosomal degradation in lieu of amyloidogenic metabolism in the endosomal pathway. Using co-immunoprecipitations, we identified novel interactions of APP with the autophagy regulatory proteins BECN1, PIK3C3, UVRAG and ATG16L1. We further found that APP associates with BECN1 at the plasma membrane by co-immunoprecipitation from cell-surface fractions and that BECN1 promotes surface APP internalization and trafficking to autophagosomes. Mapping studies using BECN1 and APP mutants indicated that the association between APP and BECN1 is primarily mediated by the evolutionary conserved domain of Beclin1 and by the C-terminus of APP . Furthermore, BECN1 binding to APP was regulated by presence of ubiquitination sites, spanning lysines 649-651, in APP C-terminus while mutation of APP endocytic motif (YENPTY) had no effect. In summary, our studies reveal a novel functional interaction between surface APP and BECN1 and its significance in regulation of APP sorting from the cell surface for degradation.

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13

Beta-amyloid Accumulation in Chronic Traumatic Encephalopathy

Thor Stein1, Victor Alvarez2, John Crary3, Gaoyuan Meng1, Ann McKee1. 1Boston VA Medical Center; 2Boston University; 3Columbia University Medical Center

Chronic traumatic encephalopathy (CTE) is a neurodegenerative disease induced by repeated mild traumatic brain injuries (TBIs). It is a tauopathy characterized by neurofibrillary tangles and tau-positive processes that preferentially involve the cortical sulci, medial temporal lobe, diencephalon, and brainstem. In CTE the tau pathology is predominantly subpial, perivascular, and within the sulcal depths. This tau pathology may be due to axonal injury, disruptions of the blood brain barrier, and vascular damage—alterations that also have been linked to Alzheimer disease (AD). In fact, trauma is a suspected risk factor for AD, and multiple mild TBIs may play a causative role in the development of AD as well as CTE. Thus, we examined our cohort of athletes and military veterans with CTE (n=92) for beta-amyloid deposition with standard histology, immunohistochemistry, ELISA, and APOE genotyping. We find that Aβ neuritic plaques occur in 37% of all cases of CTE and are significantly associated with age at death and the presence of APOE ε4 allele, but not with the duration of trauma exposure, disease duration, or CTE stage adjusted for age. We also examined sulcal versus gyral levels of Aβ and found that Aβ1-40, but not Aβ1-42, is significantly increased within the sulcus in both AD and CTE with AD subjects. The percentage of leptomeningeal vessels with cerebral amyloid angiopathy is significantly elevated within the sulcus of subjects with CTE and Aβ, but not in AD subjects alone. Overall, this suggests that parenchymal beta-amyloid accumulation occurs independently of mild traumatic brain injury and CTE, but that cerebral amyloid angiopathy is exacerbated by trauma at the sulcal depths.

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14

Single Episode of Severe Axonal Injury in Humans Can Lead to Tau Pathology Resembling Chronic Traumatic Encephalopathy

Sarah Edgerton1, Sharon Shively1, Elliott Mufson2, Dushyant Purohit3, Daniel Perl4. 1Henry M. Jackson Foundation for the Advancement of Military Medicine; 2Rush University School of Medicine; 3Icahn School of Medicine at Mount Sinai; 4Uniformed Service University, F. Edward Hebert School of Medicine

Chronic traumatic encephalopathy (CTE) is a neurodegenerative disorder associated with repetitive mild traumatic brain injury (TBI). In CTE, tau proteins aggregate forming neurofibrillary tangles (NFTs) and astrocytic tangles in a stereotyped distribution, favoring subpial sulcal depths, perivascular regions and superficial neocortical layers. Some have suggested that these tau aggregates develop following axonal damage. Therefore, we tested the hypothesis that axonal damage in the human brain can give rise to neurofibrillary and astrocytic tangle formation by analyzing postmortem brain tissues from six schizophrenic patients who underwent prefrontal leucotomy. Because leucotomy involves severing axons leading to and from the prefrontal cortex, this procedure represents a single, severe axonal injury without external cortical impact in patients who survived at least another 40 years. We examined cortical tissues at the lesion sites, prefrontal cortices rostral and frontal cortices caudal to the lesion sites, temporal cortex and hippocampus with immunohistochemistry for abnormal tau. Temporal cortex and hippocampus revealed scant NFTs, consistent with age. In all 6 cases, significant abnormal tau was detected in the gray matter adjacent to the lesion sites, with primarily astrocytic tangles in depths of sulci and NFTs in superficial neocortical layers, but not in the prefrontal and frontal cortices distant to the lesion sites. We conclude that axonal damage from leucotomy leads to accumulations of abnormal tau in gray matter adjacent to the lesion with features resembling CTE. Since significant tau pathology was not detected in prefrontal cortices, the data do not support the hypothesis that chronic neuronal deafferentation alone leads to abnormal tau accumulation. Moreover, since leucotomy lacks external cortical impact, the data suggest that selective accumulation of tau at depths of sulci may be related to underlying axonal damage rather than mechanical stresses during TBI.

(Supported by the US Army Military Research and Materiel Command and the CNRM.)

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15

Glycans Modulate the Transmissibility of PrPSc and the sCJDMM2 and sFI Phenotypes

Laura Cracco1, Ignazio Cali1,2, Anya Hurley1, Liuting Qing1, Qingzhong Kong1, Pierluigi Gambetti1. 1National Prion Disease Surveillance Center, Case Western Reserve University; 2Second University of Naples

Sporadic fatal insomnia (sFI) and the subtype of sporadic Creutzfeldt-Jakob disease (sCJD) identified as sCJDMM2 share methionine homozygosity at codon 129 (129MM) of the prion protein (PrP) gene and scrapie PrP (PrPSc) type 2 although they are associated with different phenotypes. Our data suggest that the sFI and sCJDMM2 phenotypic heterogeneity is in part related to the diversity of the PrPSc glycans in these two diseases.

To further investigate the role of glycans in phenotypic determination, we transmitted sCJDMM2 and sFI to i) transgenic (Tg) mice expressing normally glycosylated human (Hu) PrPC [Tg(HuPrPC)] and ii) Tg mice expressing human PrP free of glycans [Tg(HuPrPglycKO)]; incubation periods, histopathologies and PrPSc characteristics were compared.

Tg(HuPrPC) mice challenged with sCJDMM2 and sFI became symptomatic 631±93 and 554±60 days post inoculation (dpi), respectively. sCJDMM2-inoculated mice showed large-vacuole spongiform degeneration (SD) and a PrP immunostaining pattern similar to that of sCJDMM2. Minimal small-vacuole SD was observed in half of the sFI-inoculated mice and was associated with sFI-like PrP immunostaining. Proteinase K-resistant PrPSc (resPrPSc) type 2 was detected in both groups of mice. In contrast, sCJDMM2- and sFI-inoculated Tg(HuPrPglycKO) became symptomatic 278±36 and 354±155 dpi, respectively. Both displayed similar histopathologies characterized by widespread SD and PrPSc deposition in focal plaque-like and large granules. In both, resPrPSc formed only one band of ∼19 kDa matching the electrophoretic mobility of unglycosylated resPrPSc type 2.

In conclusion: 1) sCJDMM2 and sFI can be transmitted to Tg(HuPrPC) mice, producing different histopathologies; 2) transmission to Tg mice expressing human glycan-free PrP appears to abrogate the histopathological differences of these two diseases, supporting a role of glycans in phenotype determination; 3) in our model, PrP glycan ablation increases the transmission efficiency of both sCJDMM2 and sFI.

(Supported by P01AG 14359, CDC UR8/CCU515004, NS052319 and Charles S. Britton Fund).

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16

Transmissibility and propagation of co-existing prions of sporadic Creutzfeldt-Jakob disease into humanized transgenic mice

Ignazio Cali1,2, Wenquan Zou1, Laura Cracco1, Anya Hurley1, Tetsuyuky Kitamoto3, Qingzhong Kong1, Pierluigi Gambetti1. 1National Prion Disease Surveillance Center, Case Western Reserve University; 2Second University of Naples; 3Graduate School of Medicine, Tohoku University, Sendai, Japan

About 40% of sporadic Creutzfeldt-Jakob disease patients with methionine (M) homozygosity at PrP gene codon 129 show co-existence of the scrapie prion protein (PrPSc) types 1 and 2 (sCJDMM1-2). To investigate the mechanism of mixed PrPSc 1-2 types formation, we examined co-occurrence, transmissibility, disease phenotype prevalence and brain propagation of PrPSc using brain homogenates from a) sCJDMM1-2 harboring PK-resistant PrPSc (resPrPSc) types 1 and 2, b) in vitro mixed types 1 and 2 individually harvested from separate regions of sCJDMM1-2 and c) in vitro mixed PrPSc from sCJDMM1 and sCJDMM2, following intracerebral inoculation into transgenic mice expressing human PrPC-129M. The ratios of type 1/type 2 were 50%/50%, 35%/65% and 10%/90% in a), b) and c), respectively. Mice were sacrificed after 95, 131, 187 days post inoculation (dpi). All mice showed only resPrPSc type 1. Mice inoculated with PrPSc type 1-2 had incubation time, lesion profiles and resPrPSc features identical to those of mice inoculated with sCJDMM1 (183±22 dpi) and different from those of sCJDMM2-challenged mice (609±139 dpi). At 95 dpi, mice showed tiny amounts of resPrPSc type 1 in the central brain with spongiosis and PrP immunostaining in the thalamus. At 131 dpi, resPrPSc type 1 spread to the cerebral cortex and brain stem. The lesion profile reflected the resPrPSc distribution. At 187 dpi, resPrPSc type 1 affected the whole brain. We conclude that: 1) Mice reproduce PrPSc type 1 and sCJDMM1 phenotype regardless of PrPSc types 1 and 2 ratios in the inoculum. 2) Replication rate of type 2 is much slower than that of type 1 and is not accelerated by the presence of type 1. 3) In sCJDMM1-2, type 1 maintains the same infectivity, propagation and phenotype characteristics of type 1 of sCJDMM1.

4) Topography and timing of PrPSc propagation are highly reproducible.

(Supported by P01AG14359, CDCUR8/CCU515004, NIHNS062787, NS052319 and Charles S. Britton).

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PLATFORM SESSION 3: INFLAMMATORY, OPHTHALMIC, OTHER Friday 2-4 PM, June 13, 2014 Fashion Ballroom

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17

Transient Receptor Potential Melastatin 4 Expression in Human Cerebral Infarcts

Rupal Mehta1, Svetlana Ivanova1, Cigdem Tosun1, Rudy Castellani1, Volodymyr Gerzanich1, J Simard1. 1University of Maryland

Transient receptor potential melastatin 4 (TRPM4), a mammalian cation channel, is transcriptionally upregulated in neural and vascular cells following acute subarachnoid hemorrhage in humans and animal models. Prior studies document a leading role of this molecule in cytotoxic edema formation and necrotic cell death following diverse acute central nervous system injuries. The expression of TRPM4 in human cerebral infarcts, however, has not previously been systematically analyzed. In this study, we evaluated expression of TRPM4 in postmortem specimens obtained from 15 patients within the first 31 days after clinical onset of focal cerebral ischemia. Significant elevation of TRPM4 protein was found in neurons, astrocytes and endothelial cells in infarcted cerebrum, relative to contralateral and control tissues. Upregulation of TRPM4 protein was corroborated using in situ hybridization for TRPM4 mRNA. Furthermore, co-association of TRPM4 and sulfonylurea 1 (SUR1) was detected using co-immunoprecipitation and Förster resonance energy transfer (FRET) experiments. These findings document upregulation of SUR1-TRPM4 ion channels during focal ischemic stroke in humans. These new data indicate that SUR1-TRPM4 channel upregulation and activation are likely key molecular events leading to cell death during acute stroke, suggesting that SUR1 may be a promising novel treatment target for patients with acute cerebral infarction.

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18

Lymphocytic Hypophysitis: A Single Centre Experience of 11 Cases

Marc Del Bigio1, Angela Miller2, Jayantha Herath3. 1Pathology - University of Manitoba; 2University of Manitoba; 3Ontario Forensic Pathology Service

In the province of Manitoba Canada (population 1.4 million) all neurosurgical procedures and almost all autopsies are done at a single center. We did a retrospective review spanning the years 1996-2012, searching records for documentation of inflammation in the pituitary (2228 glands from adult autopsies, 1267 from pediatric autopsies, 339 surgical biopsies). Proven or likely infectious causes and cases with predominantly neutrophilic inflammation associated with hemorrhage or infarction were excluded. This left 11 cases with non-granulomatous predominantly T lymphocytic inflammation (7 autopsy, 4 surgical). The autopsy cases ranged from 11-64 years and the surgical cases 27-82 years (total 5 females including 1 pregnant). Among the autopsy cases, 3/7 had a clearly unrelated cause of death and no history of relevant symptoms; the pituitary inflammation was considered incidental. One had a history of seizures; there was no anatomical cause of death. In the 7 autopsy cases, other endocrine organs were available for analysis; 4/4 adrenal glands, 4/5 thyroid glands, and 2/5 pancreases had T lymphocytic inflammation within the parenchyma. In four cases the decedent had exhibited pre-mortem clinical syndrome associated with that organ (38 year female hypothyroidism; 49 year female hyperprolactinemia; 44 year male Addison disease and hypothyroidism; 49 year female diabetes mellitus); in the first three, pituitary dysfunction was considered contributory to the deaths. Immunohistochemical staining showed reduced immunoreactivity for some or all of the pituitary hormones in all of the cases. Although the prevalence of autoimmune lymphocytic hypophysitis is low, we conclude that examination of the pituitary gland is an important part of the autopsy in individuals who die unexpectedly, particularly if they have a history of endocrine disease or recent neurological symptoms.

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19

Myelin Loss in Adult-onset Leukoencephalopathy/Leukodystrophy with Axonal Spheroids is Secondary to Axonal Loss

Murad Alturkustani1,2, Julia Keith3, Lili-Naz Hazrati4, Lee-Cyn Ang1. 1London Health Sciences Centre, University of Western, London, ON; 2Department of Pathology, King Abdulaziz University and Hospital, Jeddah, Kingdom of Saudi Arabia, 3Sunnybrook Health Sciences Centre, University of Toronto, Toronto, ON; 4Tanz Centre for Research in Neurodegenerative Diseases, Toronto, ON

Background: Whether the myelin loss in adult-onset leukoencephalopathy/leukodystrophy with axonal spheroids (ALAS) is a primary demyelinating process or secondary to axonal degeneration has not been clearly established. The confluent myelin loss with U-fibre sparing gives an impression of a primary demyelinating/dysmyelinating disease. Although, the prevalence of axonal spheroids would favor a primary axonal degeneration with secondary demyelination, occasional axonal spheroids have been described in chronic primary demyelination as in multiple sclerosis (MS). In spite of the recent discovery of CSF1R mutation in ALAS, it is uncertain how this mutation correlates with the observed pathology.

Methods: We examined neuropathological features in 5 ALAS cases (3 males: 2 females; age 39-61 years) and 3 chronic MS cases (1 male: 2 females; age 50-73 years). Special histochemical stains, immunohistochemical stains and ultrastructural studies were used to study the axonal and myelin pathology.

Results: The white matter pathology in ALAS cases can be characterized as 3 evolving phases: 1) white matter with numerous spheroids in a background of well-myelinated fibres or slight myelin pallor; 2) white matter showing mixture of well-preserved myelinated fibres and degenerating fibres with sparse to moderate number of spheroids; and 3) leukodystrophy-like pattern of confluent myelin loss with relative preservation of the subcortical U-fibres, few spheroids and myelinated fibres. No convincing areas with demyelinated but relatively intact axons have been identified in our ALAS cases. In contrast, the chronic MS plaques show complete myelin loss with relatively preserved axons. Axonal spheroids are occasionally present in these plaques but not in the areas with preserved myelin.

Conclusions: Our study supports the notion that the myelin loss in ALAS is secondary to axonal degeneration.

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20

Characteristic of neural tissue in ovarian teratomas associated to Anti-NMDA receptor encephalitis (ANMDARE)

David Munoz1, Gregory Day2, Simin Laiq2, David Tang-Wai22. 1St. Michael’s Hospital, University of Toronto; 2University of Toronto

Objective & Methods: ANMDARE is the most common form of paraneoplastic encephalitis that preferentially affects women of reproductive age; over half have ovarian teratomas (OT). Most women with OT, however, do not develop ANMDARE. We investigated the histological differences between OT in 5 women with ANMDARE (mean age 28.2±8.6 years) and all OT resected at a single hospital during 2013 (39 OT from 38 women, mean age 34.8±10.0 years).

Results: Neural tissue was identified in 80% (4/5) of ANMDARE-associated OT and 51.3% (20/39) of control OT. Dysplastic neurons, often showing binucleation or multinucleation, were present in all cases with neural tissue but not in any controls (p=0.0001). If found in the brain, one case would have been diagnosed as a ganglioneuroblastoma, as dysplastic neurons occurred together with mitotically active neuroblasts and maturing neurons; the other 3 OT contained mature neurons and glial cells and would have been called gangliogliomas. In one OT the area showing this change was tiny, less than 300 um in diameter. Neural tissue was intimately associated with inflammatory cell infiltrates in all four cases but not in any controls (p=0.0001). The neural tissue often formed a rim around lymphoid follicles, suggesting that the centrally-located inflammatory cells may have focally obliterated underlying dysplastic neurons. Central nervous system tissue in control OT often showed ferruginated neurons and/or Rosenthal fibers, construed as degenerative/reactive changes.

Conclusion: We conclude that ANMDARE-associated OT differs from ordinary OT in the presence of dysplastic neurons in neural tissue and topographical relation of inflammatory infiltrates. The very small size of the areas involved in some cases suggests that dysplastic neurons may be easily missed. We hypothesize that the development of dysplastic neurons in OT triggers the inflammatory response that results in the production of ANMDARE and the partial obliteration of areas with dysplastic change.

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21

Astrocytic Regulation of Synaptic NMDA Receptors

Junghyun Hahn1, Xianhong Wang1, Marta Margeta1. 1University of California San Francisco

A growing body of evidence indicates that astrocytes regulate the formation and function of excitatory synapses, but little is known about the effect of glia on glutamate NMDA receptors (NMDARs). To investigate glial modulation of neuronal NMDARs, we performed electrophysiological, cell biological, and biochemical experiments on hippocampal neurons cultured either alone, with a mixed population of glia, or with pure astrocytes. While there was no significant change in EC50 of the NMDA-induced current between various culture conditions, the maximal current density was twice as large in mixed compared to neuronal cultures; a similar increase in NMDAR current density was seen when neurons were cultured in the presence of pure astrocytes, either directly or indirectly. The amplitude of NMDAR-mediated (but not AMPA receptor-mediated) miniature excitatory postsynaptic currents was significantly greater in mixed relative to neuronal cultures, resulting in a significantly lower synaptic AMPA/NMDA current ratio when neurons were cultured with glia; however, there was no difference in plasma membrane expression of major (NR1, NR2A and NR2B) NMDAR subunits between mixed and neuronal cultures. Astrocyte-induced potentiation of NMDAR currents was limited to the synaptic pool of NMDARs; furthermore, the proportion of NR2B subunit-dependent NMDAR currents, but not NR2A-dependent NMDAR currents, was greater in mixed than in neuronal cultures. Taken together, these data indicate that astrocyte-mediated increase in the neuronal NMDAR current (1) is postsynaptic; (2) is not due to an increase in the total number of synapses, (3) is induced by one or more astrocyte-secreted soluble factors, and (4) reflects posttranslational regulation of NR2B-containing NMDAR channels already present at postsynaptic sites, presumably on a rapid time scale. Given the role of NR2B NMDARs in synaptic plasticity and neuronal survival, our findings suggest that NMDAR modulation is an important mechanism by which astrocytes can impact neuronal circuits in both normal and disease states.

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22

A Novel Telomere Phenotype In Human Retinal Photoreceptors

William Bell1, Christopher Heaphy1, Ian Rosenthal1, Alan Meeker1, Charles Eberhart1. 1The Johns Hopkins Medical Institutions

Many neoplasms maintain telomeres via a telomerase-independent mechanism termed alternative lengthening of telomeres (ALT), but normal human tissues are not known to utilize such pathways. We previously surveyed over 6,000 different malignancies with telomere-specific fluorescence in situ hybridization (FISH) and found that the ALT phenotype is particularly common among neural neoplasms, but was never identified in the 541 benign neoplasms and 264 diverse non-neoplastic tissue samples examined. We recently expanded our analysis to include ocular samples, and unexpectedly identified ALT-like telomere DNA foci in normal retina. Using rod and cone specific antibodies, it was determined that the phenotype was largely restricted to rods. In total, 124 human retinas were examined, including 8 fetal, 22 infant, 10 childhood and 84 adult eyes. The ALT-like phenotype was fairly homogenous within each retina, but some variation was noted from eye to eye within each age group. The ALT-like telomere foci showed an age-dependent onset. They were never seen in fetal eyes, and were very rare in infants and children. Our adult eye cohort included 38 cases with pathological conditions such as diabetic retinopathy, macular degeneration, glaucoma and retinitis pigmentosa, and the ALT-like phenotype was very prominent in these. Interestingly, we also analyzed the eyes of two patients with an inherited short telomere syndrome caused by reduced telomerase function. These both showed the ALT-like phenotype in photoreceptors, raising the possibility that, like cancer-associated ALT, this retinal phenotype is independent of telomerase activity. We analyzed eyes from several rodents of varying ages, but these did not show the ALT-like telomere phenotype. In summary, we identify ALT-like telomere DNA foci in human retinal cells not present in any other normal tissue examined to date. The process is not detected in children, and its function is not yet clear.

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23

Intraocular Medulloepitheliomas in Children and Adults show Markers of Retinal Development and Glioneuronal Differentiation

Matthew Rose1, Alia Rashid2, Frederick Jakobiec2. 1Dept. of Pathology, Brigham and Women’s Hospital, Harvard; 2Dept. of Ophthalmology, Massachusetts Eye and Ear Infirmary, Harvard

Intraocular medulloepithelioma is a rare tumor arising most often near the ciliary body of the eye. It is frequently characterized by tubular and/or papillary growth of pseudostratified neuroepithelial ribbons, sometimes mixed with differentiating regions. While they generally exhibit a distinct morphology, these tumors can display regions of heterologous differentiation. We seek to further characterize the patterns of gene expression in medulloepithelioma to determine whether these tumors exhibit immunohistochemical markers reflecting their histogenetic origin. We report a series of five cases of medulloepithelioma including three pediatric (ages 1, 6, and 8 months) and two adult cases (ages 30 and 45 years), with four arising near the ciliary body and one arising from a retinal coloboma. All tumors expressed S100. In addition, both of the classic pediatric cases arising near the ciliary body showed a remarkably similar immunohistochemical expression pattern with CRX labeling primitive neuroectodermal units and GFAP and NeuN labeling areas of glial and neuronal differentiation, respectively. One of these tumors was very small and involved primarily the non-pigmented ciliary epithelium from the ora serrata to the iris, offering a glimpse into the early stages of tumor growth. One adult case likewise showed CRX and synaptophysin expression, while the other showed only CRX expression without markers of glioneuronal differentiation. In contrast, the last pediatric case appeared to arise from a coloboma and showed areas of myogenic rather than glioneuronal differentiation, although it focally showed primitive neuroectodermal morphology. With follow-up intervals ranging from eight months to eleven years, all five patients are alive with no reported recurrence of tumor following enucleation. Overall, these findings highlight immunohistochemical markers of retinal development and glioneuronal differentiation which are consistent with the origin of medulloepitheliomas in the non-pigmented ciliary epithelium of the pars plicata or pars plana interfacing with the peripheral retinal ora serrata.

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24

Opthalmological Abnormalities in CTE

Victor Alvarez1, John Crary2, Nathalie Slick1, Brian Fry1, Thor Stein3, Ann McKee3. 1Boston University School of Medicine; 2Columbia Presbyterian School of Medicine; 3VA Boston Healthcare System, Boston University School of Medicine

Repetitive concussive injury is associated with the development of chronic traumatic encephalopathy (CTE), a progressive brain degeneration characterized by hyperphosphorylated tau (p-tau) neurofibrillary tangles in the brain (McKee et al., 2009, Goldstein et al., 2012, McKee et al 2013). CTE produces symptoms of irritability, impulsivity, depression, memory loss, and suicidality that may progress to dementia and parkinsonism in late stages. The neuropathological changes of CTE are distinctive and easily distinguished from other neurodegenerative diseases, including Alzheimer’s disease. Key neuropathological features of CTE are p-tau neurofibrillary tangles around small blood vessels, neuroinflammation, and deposition of abnormal aggregates of TDP-43, a protein associated with ALS, alpha B crystallin, a small heat shock protein, and p62, an autophagosome marker. We have diagnosed CTE at post-mortem examination in over 100 former football players, hockey players, boxers and military veterans of the Iraq and Afghanistan conflicts exposed to blast injury (McKee et al 2013). Currently, there is no method of detecting CTE during life and the diagnosis can only be made at autopsy. Nothing is presently known about the eye pathology of CTE but if a distinctive profile of ocular abnormalities were identified, it would raise the possibility that CTE could be diagnosed during life by eye examination. Our preliminary studies demonstrate that p-tau inclusions, neuroinflammation and abnormal aggregates of the proteins TDP-43, alpha B cystallin, and p62 are found post-mortem in the inner plexiform layer of the retina in individuals with advanced CTE that were not found in age-matched controls. These findings suggest that the ocular pathology of CTE is unique and that methods of in vivo detection might be developed to diagnose CTE opthalmologically during life.

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PLATFORM SESSION 4: DEVELOPMENTAL/PEDIATRIC Friday 2-4 PM, June 13, 2014 Culture Ballroom

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25

Development of the Hippocampal Formation in Human: Part I-Pyramidal Cell Layer

Sara Cipriani1, Catherine Verney1, Jeannette Nardelli1, Anne-Lise Delezoide1, Pierre Gressens1, Homa Adle-Biassette2. 1INSERM UMR 1141; 2Department of Pathology, Lariboisiere Hospital, APHP & INSERM UMR 1141

This is the first detailed study describing the development of hippocampal formation in human embryos and fetuses from 8 weeks of gestation (GW) to midgestation. Multiple immunohistochemical labelings were performed to study the progenitors using Ki67 associated with nestin, SOX2, PAX6 and TBR2, and post mitotic neuronal markers using NeuN, CUX1, SATB2, CTIP2, TBR1, NeuroD1, doublecortin, CaBP, and Calretinin. Our results showed the presence of a gradient of neurogenesis starting first in the entorhinal cortex and extending toward the ammonic plate. The proliferation of Pax6+ or Sox2+ progenitors was already observed at 9 GW in the ventricular zone. Intermediate-stage progenitors labeled by Tbr2 were present at 11 GW in the subventricular zone. The density of both types of progenitors progressively decreased around 20GW. Postmitotic neurons appeared during the embryonic stage. In the ammonic plate Tbr1, CTIP2 and Cux1 labeled numerous cells, some of which were colabeled with Sox2 and Pax6. They progressively migrated to the deep and superficial layers of the ammonic plate until midgestation. The density of SATB2-labeled neurons in the ammonic plate was lower compared to the other layer markers and to the subicular / entorhinal cortical plates.

In conclusion, neurogenesis in the hippocampal formation starts during the embryonal period. The pattern of expression of transcription factors suggests that postmitotic neurons are produced directly from funding progenitors and from intermediate-stage progenitors. The lamination of the pyramidal layer follows an in an inside out gradient, producing a three-layered ammonic plate compared to the entorhinal cortex.

The research leading to these results was performed in the frame of DEVELAGE project “Pathways common to brain development and ageing: defining strategies for preventive therapy and diagnostics” (HEALTH-F2-2011-278486) and has received funding from the European Community’s 7th Framework Programme (FP7/2007-2013).

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26

Fetal Nucleus/fasciculus Solitarius: Synaptophysin Maturation

Harvey Sarnat1, Laura Flores-Sarnat1. 1University of Calgary and Alberta Children’s Hospital

Background: The nucleus/fasciculus solitarius is the respiratory centre of the brain, also known as the pneumotaxic centre. It consists of paired longitudinal columns in the tegmentum of the medulla oblongata, extending to the cervical spinal cord. A white matter core of longitudinal axons (fasciculus) is surrounded by neurons (nucleus), a primitive arrangement. Afferent fibres to the nucleus solitarius arise mainly from chemoreceptors and stretch receptors in respiratory muscles and lung. Efferent axons descend the fasciculus to innervate spinal motor neurons to the diaphragm and other respiratory muscles. This nucleus is essential for fetal rhythmical respiratory movements observed by real-time ultrasound and postnatal respiration.

Methods: Sections of medulla oblongata of 20 normal fetuses of 9-40 weeks gestation were studied at autopsy by immunoreactivity to synaptophysin and other markers of neuronal maturation: NeuN, calretinin, NSE. Three fetal chromosomopathies, two preterm neonates with apnoea of prematurity and 2 cases of Möbius syndrome also were examined.

Results: Synaptogenesis in the nucleus solitarius begins early in the human fetus, at 10-11 weeks gestation, and it is advanced by 15 weeks. Delayed synaptophysin expression was found in 2 premature infants with apnoea but not in infants with chromosomopathies; maturation of other neuronal markers was not altered. The nucleus was bilaterally infarcted in 2 infants with Möbius syndrome.

Conclusions: Hypoxia, metabolic disturbances and other adverse events affecting late 1st and 2nd trimester fetuses may impair the rate of formation of synapses in the nucleus solitarius and explain some cases of apnoea of prematurity and SIDS. Symmetrical watershed tegmental infarcts may involve the nucleus solitarius to account for central respiratory insufficiency in neonates with Möbius syndrome.

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27

Radial Glia Defects and the Pathogenesis of Germinal Matrix Hemorrhage.

Jennifer Cotter1, Mianzhi Tang1, Eric Huang1. 1University of California San Francisco, Department of Pathology

Germinal matrix hemorrhage (GMH) is a life-threatening condition that affects over 12000 infants per year in the US. Its pathogenesis has been attributed to alterations in hemodynamics and/or vascular fragility, but the mechanism of its development remains poorly understood. We evaluated human autopsy tissues from the Pediatric Neuropathology Research Lab to study potential developmental alterations in the setting of GMH. Brain sections from preterm infants with GMH (n=5) and without (n=13), ranging from gestational ages of 16 to 36 weeks were evaluated. Using stereology, we found no differences in vascular branching or vascular density within the germinal matrix (GM) between controls and GMH cases. Furthermore, in situ hybridization for vasculogenic factors, VEGF, TGFbeta and integrin beta8, showed no detectable differences in the expression of these genes in the GM. To characterize how altered neurogenesis within the GM may contribute to the pathogenesis of GMH, we performed immunohistochemistry for Ki-67, Nkx2.1, Sox2, Nestin, and NeuN. Consistent with previously published data, human GMH cases showed reduced Ki-67 and Sox2 labeling indices within the GM when compared to controls. Intriguingly, similar to mouse ganglionic eminence (GE), human GM could be further sub-divided into a medial compartment (MGE) with high Nkx2.1 expression and a lateral compartment (LGE) with low Nkx2.1. Using Nestin as a marker for radial glia, we found that in control cases, the MGE showed higher radial glia density than the LGE. In contrast, GMH cases over 28 weeks gestational age demonstrated decreased Nestin-positive radial glia not only adjacent to hemorrhage, but also in the contralateral GM. The finding of reduced radial glia density in bilateral GM of preterm infants with GMH suggests that the developmental differences in radial glia between controls and the brains of preterm infants with GMH most likely contribute to the pathogenesis of GMH.

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28

A Century of Brain Hemorrhages in Autopsied Premature Infants at a Tertiary Pediatric Hospital

Marco Hefti1, Robin Haynes2, Rebecca Folkerth3, Felicia Trachtenberg4, Joseph Volpe2, Hannah Kinney2. 1Department of Pathology, Beth Israel Deaconess/Harvard Medical School; 2Boston Children’s Hospital; 3Brigham and Women’s Hospital; 4New England Research Institutes

The care of premature infants in the 20th century is remarkable for technical advances that have dramatically improved survival, but little is known about changes in the neuropathology of the premature infant over this time frame. We hypothesize that neuropathologic entities have changed over the last century, disappearing because of improvements in care, or arising anew due to unforeseen complications of new treatments and/or increased length of survival. To begin to test this hypothesis, we examined germinal matrix hemorrhage with intraventricular hemorrhage (GMH-IVH) and intraparenchymal cerebellar hemorrhage (CH) in 351 premature infants (gestational age 24-37 weeks) autopsied at Boston Children’s Hospital from 1924 to 2010. There was a median of 39 cases/decade (range 16-66). Over the course of the study, gestational age decreased from 31.6 (1924-1930) to 29.0 (2001-2010) weeks (p<0.001), and survival increased from 1.5 to 14.5 days (p<0.001). The incidence of GMH-IVH increased from 2.7% before 1950 to 37.7% from 1970-1980, and then decreased to 15.4% after 1990 (p<0.001), as adjusted for infant gestational age and length of survival time. The incidence of CH showed a significant linear trend, increasing from 2.7% prior to 1950 to 20.5% after 1990 (p=0.005). The incidence of GM-IVH increased >3-fold around the time of the introduction of positive pressure ventilation into premature intensive care in the mid-1960s. The increased incidence of GM-IVH in the 1970-1980s likely reflects respiratory and hemodynamic imbalances complicating mechanical ventilation; the subsequent decreased incidence likely reflects stabilization of respiratory function with the use of surfactant beginning in the 1980s. In contrast, the incidence of CH gradually increased into the 1990’s, suggesting that factors other than those related to mechanical ventilation are involved and require elucidation.

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29

Hippocampal Anomalies in Sudden Unexplained Death in Young Children: An Extended Series.

Marco Hefti1, Jane Cryan1, Elisabeth Haas2, Marjorie Grafe3, Laura Crandall4, David Patterson1, Lisa Teot1, Felicia Trachtenberg5, Hannah Kinney1, Henry Krous2. 1Department of Pathology, Boston Children’s Hospital; 2Department of Pathology, Rady Children’s Hospital San Diego; 3Department of Pathology, Oregon Health and Sciences University; 4SUDC Program, Hackensack, NJ; 5New England Research Institute, Watertown, MA

Sudden unexplained death in childhood (SUDC) is the sudden death of a child older than 1 year that remains unexplained after a review of the clinical history, circumstances of death, autopsy, and ancillary testing; its incidence in children 1-3 years is 1.3/100,000. Previously, we reported a putative new entity of SUDC associated with hippocampal/temporal lobe (TL) maldevelopment with or without a personal and/or family history of febrile seizures (FS) in a subset of children aged 1-6 (Kinney HC et al. Ped Dev Pathol 2019; 12: 455-463). Sixty-four SUDC cases were accrued from 2002-2007 in an autopsy study of sudden unexpected death (77% SUDC, 23% explained); of the 49 SUDC cases, 46% with hippocampal sections (12/26) had substantial microdysgenesis. From 2007-2011, we accrued 88 additional cases (SUDC, 88%). We tested the hypothesis that a larger sample size confirms the observation of hippocampal anomalies associated with SUDC. We report combined findings in the cohort of 151 cases (1-6 years) based upon review of clinical records, autopsy and scene reports, microscopic slides, and family surveys. The demise categories were: SUDC, 80% (n=121); explained/known causes, 13% (n=19) (e.g., infection, accidents); undetermined, 5% (n=7), and epilepsy, 3% (n=4). There was no difference in age at death, male gender, or preterm birth among the groups. In the SUDC group, 49% (59/121) had a personal and/or family history of FS compared to 11% (2/19) of known causes (p=0.003). Of the SUDC cases, 97% were discovered after a sleep period, and 86% in the prone position. Hippocampal/TL review revealed 43% (36/83) of SUDC cases had gross and/or microscopic anomalies compared to 13% (2/16) of known causes (p=0.02). Of these SUDC cases, 31% had a personal FS history, and 17%, a family FS history only. Future research is needed towards identifying living young children at risk for sudden death.

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30

Construction of a Single Adenoviral Vector Carrying Both ZFN and Donor for the Treatment of Lysosomal Storage Diseases

Qinwen Mao1, Weifeng Zhang2, Esther Bit-Ivan1, Eileen Bigio1, Haibin Xia2. 1Northwestern University Feinberg School of Medicine; 2Shaanxi Normal University, China

Many lysosomal storage diseases (LSDs) have devastating consequences including neurodegenerative diseases. Although enzyme replacement therapy is a potential treatment option, this approach needs repeated injection over the lifetime of the patients. Gene-addition strategy using viral vectors has the potential for long-term expression of the therapeutic protein, however, serious adverse effects, e.g., immune response to in vivo administered viral vectors, and insertional activation of proto-oncogenes, can occur. An alternative to gene addition that can minimize the oncogenic risk of gene therapy is targeted gene correction via homologous recombination. Zinc-finger nucleases (ZFNs) show promise as reagents that can mediate high-frequency homologous recombination in the presence of a donor. This technique has shown success in ex vivo correction of a disease-causing mutation by using induced pluripotent stem cells, which can be useful for the treatment of diseases affecting cells that can be removed and returned to the patient. However, many LSDs might need in vivo gene correction in affected organs, which requires the efficient introduction of gene-targeting components (i.e., ZFNs and donor fragment) in vivo. We utilized adenovirus, a high-efficiency and safe vector with sufficient size to fit both ZFN and donor to mediate gene correction in vivo. We have overcome the challenges of packing both ZFN and donor in a single adenovirus, and produced ZFN expressing adenovirus with high titer. We also show that this novel system efficiently mediates targeted genome editing. In summary, adenovirus may be a promising vector for in vivo gene correction in disease-affected organs in lysosomal storage diseases.

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31

The KCa3.1 blocker TRAM-34 Reduces Activated Microglia and ASD Like Behavior in a Rat Model of Neonatal HI Brain Injury

Mirna Lechpammer1, Yen Tran2, Philip Huebner2, Yi-Je Chen3, Robert Berman4, Lee-Way Jin2, Heike Wulff3. 1University of California Davis School of Medicine; 2Department of Pathology & Laboratory Medicine, UC Davis; 3Department of Pharmacology, UC Davis; 4Department of Neurosurgery, UC Davis

Neonatal hypoxia/ischemia (HI) causes Encephalopathy of Prematurity (EOP), which is associated with neurobehavioral deficits and autism spectrum disorders (ASD) in later life. We assessed the role of the intermediate-conductance calcium-activated K+ channel KCa3.1 in microglia activation after neonatal HI brain injury. We also evaluated the neuroprotective efficacy of the selective small molecule KCa3.1 inhibitor TRAM-34, developed by our group, at a molecular level and by behavioral assessments in a rat model of EOP.

We investigated brain tissue from male Long Evans rats after selective white matter (WM) injury produced in P6 by unilateral carotid artery ligation (UCL) followed by severe hypoxia (UCL/HI). The immunocytochemical analysis has shown an increased expression of KCa3.1 in activated microglia (Iba1+) in rat WM in P10 following UCL/HI at P6. TRAM-34 (40 mg/kg i.p. every 12 hours for 3 days) initiated 12 hours after UCL/HI inhibited microglial response at P10 as demonstrated by the reduced number of activated microglia in WM compared to vehicle treated controls. Likewise, a significant reduction (p=0.04) in WM injury (MBP staining) was observed at P14 in TRAM-34 treated rats (40 mg/kg i.p. every 12 hours for 7 days) compared to controls. When their behavior was assessed by the three-chambered sociability test, young rats (P31-33; n=15) exposed to UCL/HI at P6 exhibited resistance to change and impaired preference for social novelty. Importantly, TRAM-34 has prevented this ASD-like behavior and restored normal preference for social novelty as observed in healthy controls.

Presented results indicate increased immunoreactivity of KCa3.1 in activated microglia following HI pro-inflammatory stimuli in neonatal rat brain. Our data also demonstrate feasibility of prevention/reversal of cognitive consequences of EOP by in vivo pharmacological blockade of KCa3.1, which in turn may have translational clinical potential for this as yet untreatable cause of neurocognitive deficits, including autism and cerebral palsy.

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32

Developmental Synaptic Plasticity Defects in a Mouse Model of Down Syndrome

Christopher William1, Lubna Saqran2, Matthew Stern2, Matthew Frosch1, Bradley Hyman3. 1Neuropathology Service, Massachusetts General Hospital, Boston, MA; 2Department of Neurology, Massachusetts General Hospital; 3MassGeneral Institute for Neurodegenerative Disease, MGH, Boston, MA

Understanding the molecular basis for cognitive delay in Down syndrome (DS) is a challenge that would be aided by assays of circuit function in models of the disease. Synaptic plasticity is critical for circuitry refinement; to explore whether defects in synaptic plasticity may contribute to developmental impairment in DS, we have assessed visual system plasticity in a partial duplication mouse model of DS, the Ts65Dn line.

Ocular dominance plasticity is the process by which loss of vision in one eye, monocular deprivation (MD), causes changes in connectivity that result in an increase in the area of primary visual cortex responsive to the non-deprived eye and a strengthening of the relative magnitude of responses to stimulation of the non-deprived eye compared to stimulation of the deprived eye. Trisomic and non-trisomic litter mate mice underwent MD via lid suture over six days between P27 and P35. Expression of the immediate early gene Arc was used to determine the width of the cortical domain responsive to the non-deprived eye ipsilateral to that eye. To assess the magnitude of cortical responses to stimulation of each eye, optical imaging of intrinsic signals was performed on awake, head-fixed mice, before and after MD. Following MD, trisomic mice fail to demonstrate an expansion of the domain of visual cortex responsive to the non-deprived eye (trisomic width, 944 ± 77.89 microns, n=5; non-trisomic width, 1087 ± 87.44 microns, n=13; p=0.0028, T-test). Non-trisomic mice demonstrate strengthening of responses to non-deprived eye stimulation (pre-MD dF/F, 0.37%, n=6; post-MD, 0.52%; p=0.01, T-test), however, trisomic mice fail to demonstrate an increase in response strength following MD (pre-MD, 0.5%, post-MD, 0.49%, n=9; p=0.21, T-test). These data suggest that Ts65Dn mice demonstrate plasticity defects that will be used in future studies to explore the molecular basis for developmental and cognitive impairment in DS.

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POSTERS: DAY 1 10-10:30 AM, 4-4:30 PM, June 13, 2014 Foyer

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33

An Interesting Case Of Sarcoid Myopathy

Luis Gonzalez-Cuyar1, Desiree Marshall1, Nina Bozinov1, Zackary Hoffer1, Michael Weiss1, B. Distad1, C. Keene1. 1University of Washington

We report the case of a 69-year-old man with clinical history of sarcoidosis initially diagnosed in his 20’s with lifelong weakness that has progressively worsened over the last 2-3 years. Physical exam was notable for diffuse weakness and wasting (proximal worse than distal) as well as finger flexor and extensor weakness. MRI showed a nonspecific myopathy that involved the pelvis as well as the bilateral thighs. Electromyography revealed a mild predominantly chronic myopathy and laboratory studies revealed creatine kinase of 53 U/L. The patient underwent a left vastus biopsy which was characterized by myonecrosis with patchy lymphohistiocytic and granulomatous inflammation, HLA-I upregulation and neurogenic atrophy in the setting of endomysial fibrosis and moderate to focally severe fiber size variation. Rimmed vacuoles, TDP-43 positive inclusions and ultrastructural evidence of sarcoplasmic filamentous inclusions were also identified. Chronic sarcoid myopathy syndrome is characterized by symmetrical muscle weakness and wasting in the limb girdle and proximal limbs with often normal serum CK and granulomatous inflammation on muscle biopsy. Although rimmed vacuoles are a relatively non-specific myopathic feature, examples of sarcoid myopathy in concomitance with features of inclusion body myositis (IBM) have rarely been reported. Coexisting neurogenic features, as in this case, are often present.

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34

Neuromuscular Pathology of Bannayan-Riley-Ruvalcaba Syndrome: A Rare Case Report and Review of Literature

Keng-Chih Su1, Negar Khanlou1, Anthony Verity1, Jennifer Yi1. 1Neuropathology Department, David Geffen School of Medicine at UCLA

Bannayan–Riley–Ruvalcaba syndrome (BRRS), now a category of PTEN hamartoma-tumor syndromes (PHTS), is an autosomal dominant genodermatosis resulting from mutations in PTEN. A neuromuscular clinical presentation is also seen in affected young children. However, reports on associated neuromuscular pathology are rare in the literature. We report a 5-year-old with infantile cerebral palsy and clinical consideration of BRSS, presenting with acute leg pain on exertion, left leg hypertrophy and fatigability. Motor weakness was noted in the affected limb at examination. MRI of the left leg showed focal edema and enhancement without mass lesion. A left rectus femoris muscle biopsy was obtained. The fascicular architecture was overall preserved while the epimysium was site of angiolipoid hamartomas associated with extensive nodular lymphohistiocytic inflammation. Intra-tumoral vascular changes including perivascular inflammation and intimal hyperplasia were frequently noted. Myofiber study was abnormal including dysmaturation neuromyopathy, an unusual subsarcolemmal pattern of intra-fiber lipid distribution without hyperlipidation, neurogenic change and type II fiber smallness. Conclusion: To our knowledge skeletal myofiber pathology observed in this biopsy was scarcely reported in patients with BRRS. A direct association of dysmaturation phenomenon with PTEN status cannot be ruled out and requires further investigation. This case also underlines the importance of skeletal muscle biopsy not only in patients with BRRS but also in young children with early neuromuscular symptoms. The syndrome is known to be underdiagnosed because of its phenotypic variability, its incomplete penetrance, and the frequency of its individual components in the general population. While macrocephaly, intestinal polyposis and sub-mucocutaneous tumors are common findings, the neuromuscular presentations are less recognized as heralding symptoms. In addition to access to tumor pathology, the muscle biopsy allows for the diagnosis and study of skeletal muscle damage in early stages of its progression and proactive clinical management of its complications.

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35

Novel Homozygous Cofilin-2 (CFL2) Mutation Causing a Congenital Form of Nemaline Myopathy with Filamentous Inclusions

Leslie Bruch1, Sheraden Mundy2, Natalie Hauser2, Joseph Shen2. 1University of Iowa; 2Children’s Hospital Central California

Nemaline myopathy most often presents as a congenital myopathy characterized by hypotonia and muscle weakness; it is pathologically distinguished by the presence of nemaline rods. Seven causative genes have thus far been identified with 6 of the 7 encoding proteins associated with sarcomeric thin filaments. The function of the seventh gene has not yet been identified. Cofilin-2 (CFL2) encodes an intracellular protein that is involved in the regulation of actin filament dynamics. Mutations in CFL2 are known to be responsible for nemaline myopathies in only two families, both with consanguinity. We describe a novel homozygous mutation in CFL2 presenting as congenital hypotonia with pathologic features showing overlap between those of nemaline myopathy and other protein aggregate myopathies. The proband initially presented at 3 weeks of age due to poor oral intake and respiratory distress, but feeding difficulty had been present since birth. He was born to consanguineous parents after an unremarkable pregnancy. His weakness progressed to ventilator dependency, and he received a tracheostomy and gastrostomy button placement. At 3.5 months of age he had minimal movement of his extremities. CPK was elevated up to 780 U/L. A left quadriceps muscle biopsy showed increased fiber size variation and scattered fibers with inclusions composed of finely granular red-purple material on trichrome as well as a few fibers with larger inclusions characteristic of nemaline rods. No necrosis, inflammation, fiber type disproportion or predominance was found. Electron microscopy demonstrated areas of sarcomeric disarray with filamentous inclusions, mini-cores and occasional nemaline bodies or rods. Sanger sequencing revealed a homozygous mutation in exon 4 (c.353C>A, p.Ala118Asp). This new case represents the third family described with mutations in CFL2 and further expands the clinical and pathologic phenotype associated with this genotype.

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36

Diagnostic Pitfall with Use of Alpha-dystroglycan Immunostain in Perimortem Muscle Biopsies

Amanda Kan1, Sophelia Chan2. 1Department of Clinical Pathology, Tuen Mun Hospital, Hong Kong; 2Department of Paediatrics and Adolescent Medicine, Queen Mary Hospital

Abnormal glycosylation of alpha-dystroglycan (aDG) occurs in certain forms of congenital muscular dystrophy. Immunostaining for aDG is a convenient and useful tool in workup for congenital muscular dystrophy in muscle biopsy. The degree of reduction of aDG immunolabelling often correlates with disease severity. However, we encounter two perimortem muscle biopsies with no clinical evidence of congenital muscular dystrophy but absence immunostaining for aDG. We propose the loss of staining or alteration protein expression affecting the group of dystrophin and dystrophin-associated complex at the terminal stage of life being physiological or artifactual.

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37

Unusual Pathologic Features in a Patient with Laing Early Onset Distal Myopathy

Marta Margeta1, Grant de la Motte2, Nigel Laing3. 1University of California San Francisco; 2Palo Alto Medical Foundation; 3University of Western Australia

Laing early onset distal myopathy (MPD1) is a rare form of distal myopathy caused by mutations in MHY7 myosin heavy gene. Clinical presentation of MPD1 patients is fairly stereotypical (development of complete bilateral foot drop in the early childhood, followed by a very slow clinical progression in subsequent years); in contrast, the reported biopsy findings are relatively nonspecific. Here, we present a patient with MPD1 whose muscle biopsy showed distinctive light microscopic and ultrastructural features. The patient, a 39 year old male, developed bilateral foot drop at age 5, but is otherwise asymptomatic and physically active. Physical exam showed bilateral weakness of ankle dorsiflexors (1/5), toe extensors (1/5), and finger extensors (4+/5); creatine kinase level was within the normal limits. The disease first developed in the patient’s paternal grandmother and is transmitted in autosomal dominant fashion, with all affected family members showing the same clinical phenotype. The quadriceps femoris biopsy showed moderate variation in muscle fiber diameters, mild endomysial fibrosis, scattered nuclear clumps, and type 1 fiber predominance (85%), but no degenerating/regenerating fibers or rimmed vacuoles. In addition, randomly distributed type 1 (but no type 2) fibers showed abnormal invaginations of sarcolemma that were most readily apparent on ATPase, NADH and Toluidine blue stains. On ultrastructural analysis, these abnormal membrane invaginations were reminiscent of myotendinous junctions, except for the degree of complexity and the lack of clustering in a single area of the specimen. Genetic analysis of the patient and three affected relatives showed heterozygous deletion of GAG triplet at position 4522-4524 of MHY7 gene, which results in a deletion of a glutamate residue at position 1508 of the beta myosin heavy chain protein [c.4522_4524delGAG (p.Glu1508del)]. This case suggests that the spectrum of histopathologic findings associated with MPD1 is wider than previously believed.

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38

Granulomatous Fungal Myositis Presenting as Isolated Unilateral Myalgia: A Case Report

Osama Elkadi1, Jiang Qian2. 1Albany Medical Center/College; 2Pathology, Albany Med Ctr Hosp/APS

Fungal myositis is a rare but serious form of myositis, with very few cases reported in the literature. It usually occurs in the setting of immunocompromise, especially in AIDS patients. The most common isolated fungal organisms are aspergillus and cryptococcus. Here we report a case of isolated fungal myositis without evidence of disseminated disease.

The patient is a 48 year-old African American female with history of cryptogenic cirrhosis, status post liver transplant on immunosuppression. She presented with left lower extremity pain, tenderness and mild swelling. MRI showed increased STIR signals most evident in the flexor compartment, suggesting myositis. After ruling out DVT and compartment syndrome, a left gastrocnemius muscle biopsy was obtained, which revealed necrotic / regenerative muscle fibers and a non-necrotizing, vaguely granulomatous inflammation with dense mononuclear infiltrates composed mainly of CD3 positive T lymphocytes (CD4-dominant) and CD68 positive histiocytes with occasional multinucleated giant cells. Among the inflammatory infiltrates were multiple small spherical forms which were pale and slightly refractile on H&E stain, negative on AFB, mucicarmine and alcian blue stains, but confirmed on GMS and PAS-D stains to be yeast-form fungi, most consistent with blastomyces species. The patient received systemic antifungal therapy, amphotericin, followed by oral fluconazole. CSF and blood cultures were negative, as was culture from the second muscle biopsy 23 days after the initial diagnosis. Two and half months later, she had low-grade fever and abdominal pain, but otherwise stabilized. Her HIV testing was negative. She had elevated anti-nuclear antibody titer in the past, but anti-mitochondria antibody, smooth muscle antibody and rheumatoid factor were all within normal.

This case illustrated that a high index of suspicion for infectious etiology should be raised in inflammatory myopathy work-up in immunocompromised patients, so that early accurate diagnosis can be achieved and appropriate treatment promptly administered.

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39

Regional Necrotic Myopathies: Border Zone Muscle Fiber Necrosis Associated with Dermatomyositis Syndromes and Neoplasms

Chunyu Cai1, Rati Choksi2, Ali Alshehri2, Alan Pestronk2. 1Department of Pathology and Immunology, WUSM; 2Department of Neurology, Washington University School of Medicine

Introduction: Necrosis and regeneration of scattered muscle fibers are common features of many active myopathies. We studied a series of patients with acquired myopathies whose myopathology included an unusual pattern of regional, rather than scattered, muscle fiber necrosis and regeneration.

Methods: Retrospective review of clinical, laboratory, myopathologic and ultrastructual features of seven patients with acquired myopathies having regional necrosis and regeneration of muscle fibers on muscle biopsy.

Results: Clinical features included proximal symmetric weakness with a subacute onset (100%), dysphagia (83%), myalgias (100%), and a skin rash (67%). Serum creatine kinase was often (83%) very high (>1,600 U/L). An associated malignancy was common (71%). Survival was less than one year in 43%. Myopathology included regions of muscle fiber necrosis or regeneration in border zone territories between intermediate-sized perimysial vessels. Muscle fibers and connective tissue in regions of necrosis show C5b-9 deposition and expression of the ischemia marker carbonic anhydrase IX. In areas between the regions of necrotic fibers and perimysial vessels, endomysial capillaries are enlarged and stain for alkaline phosphatase and muscle fibers have upregulated MHC Class I. Intermediate-sized perimysial veins had abnormal structure and increased cellularity, likely marginating macrophages and polymorphonuclear leukocytes, in the wall. There were no aggregates of mononuclear cells.

Conclusions: Regional necrotic myopathies (RNM) are a distinctive myopathologic, often paraneoplastic, subgroup of dermatomyositis. Muscle fiber damage may be due to ischemia in border zone regions between intermediate perimysial blood vessels.

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40

Unusual Presentation of Hereditary Sensory Autonomic Neuropathy

Armine Darbinyan1, Kenneth Hughes1, Mary Fowkes1. 1Pathology, Icahn School of Medicine, Mount Sinai Medical Center, NY

We describe an unusual presentation of hereditary sensory autonomic neuropathy in a young woman. The patient was diagnosed at age 9 with biopsy proven autoimmune hepatitis and was treated with azathioprine. She remained active and attended her 1st year of college. At age 18 she developed EBV mononucleosis and subsequently diffuse paresthesias with progressive weakness thought to be Guillann Barr Syndrome variant. She was treated with IVIG with mild improvement. She then developed pancreatitis with autonomic instability and became wheelchair bound with bladder incontinence. Neurologic examination revealed intact CN2-12, profound sensory and proprioceptive loss, weak deep tendon reflexes, 3-4/5 motor strength of upper and lower extremities, mild dysmetria, and inaccurate rapid alternating movements. A motor nerve conduction study revealed absent sensory potentials consistent with a diagnosis of predominantly sensory neuropathy or neuronopathy (ganglionopathy). She developed global encephalopathy in the context of hepatic decompensation, worsening ascites, rising ammonia, with persistent asterixis and expired two months following her initial EBV diagnosis.

Autopsy microscopic examination revealed moderate loss of dorsal root ganglion neurons and diffuse axonal loss in the dorsal columns of the spinal cord without associated inflammation. There was preservation of motor neurons and axons in the descending corticospinal tract of the spinal cord. A deltoid muscle section revealed scattered myocytes with moderate atrophy, no myocyte necrosis, no increase in internalized nuclei, no increase in endomysial or perimysial fibrosis, and no inflammation.

This histologic phenotype is most consistent with hereditary sensory autonomic neuropathy which can have an associated component of muscle weakness with atrophy as seen in this case. Acquired disorders would unlikely present with selective neuronal loss as seen in this case. The finding of preserved motor neurons and axons rules out hereditary motor and sensory neuropathy. However, the clinical picture of autoimmune hepatitis and pancreatitis has not been previously reported.

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41

Paraneoplastic Sensory Neuronopathy In A 63-year-old Man Without A Demonstratable Underlying Malignancy

Jenny Smith1, Jeremy Deisch1. 1Loma Linda University

Paraneoplastic disorders of the nervous system comprise a heterogeneous group of conditions that are united by autoimmune damage to elements of the central nervous system, mediated by antibodies directed against antigens that cross react with both neoplastic cells and nervous system constituents (i.e. onconeural antibodies). The prototypical example of paraneoplastic nervous system damage is sensory neuropathy with or without central nervous system damage. Many of these cases harbor anti-Hu antibodies. An underlying malignancy is identified in most cases, usually small cell carcinoma of lung origin. We present a case of paraneoplastic sensory neuronopathy with minor components of limbic encephalitis and Purkinje cell damage, without demonstrable malignancy by either thorough clinical investigation or autopsy study. The patient was a 63-year-old man with a 45 year smoking history that presented with a forty pound weight loss and three month history of weakness, numbness, and parasthesias. All extremities were weak, most prominent in the proximal legs. Deep tendon reflexes were decreased throughout. An EMG demonstrated a prominent sensory axonal polyneuropathy. Radiologic and endoscopic search for malignancy demonstrated FDG avid hilar and retroperitoneal lymphadenopathy, but was otherwise negative. Serum onconeural antibody testing revealed anti-Hu, as well as Purkinje cell fluourescence. Anti-Yo antibodies were not detected. A full autopsy was performed, and no malignancy was identified. Neuropathologic examination demonstrated prominent ganglionic depopulation of all dorsal root ganglia. Scattered small aggregates of lymphocytes where present within the dorsal nerve roots and ganglia. Aggregates of residual satellite cells (“nodules of Nageotte”) were frequent. The spinal cord showed prominent degeneration of the dorsal columns. Perivascular chronic inflammation limited to the basal ganglia was noted; the brain was otherwise unremarkable. The cerebellum showed slight Purkinje cell depopulation and Bergmann gliosis. This case demonstrates an example of paraneoplastic sensory neuronopathy and encephalomyelitis with clinical and pathologically occult malignancy.

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42

Expression of SERPING1 is Increased in Reactive Astrogliosis Associated with Neuroinflammation

Amber Nolan1, Nicole Croom1, Han Lee1, Marta Margeta1. 1University of California San Francisco

Reactive astrogliosis was once thought of as a uniform, stereotyped response to injury of the brain. However, recent gene expression profiling data suggests that astrocytic reaction is heterogeneous, with gene expression patterns that depend on the nature of the original insult and thus provide candidates for new reactive astrocyte markers that will be clinically useful in differentiation of equivocal pathologic states. SERPING1 (also known as C1 inhibitor), a serine protease inhibitor which primarily functions to inhibit the complement system, was recently shown to be preferentially expressed in a mouse model of inflammatory brain injury compared to a mouse model of brain ischemia. To asses if this association held true in human disease, we analyzed expression of SERPING1 using immunohistochemistry on paraffin-embedded tissue from surgical and autopsy cases of encephalitis, cerebral ischemic infarct, and pathologically unremarkable brain tissue. Our preliminary results replicate the existing mouse model data. Specifically, while control brain tissue exhibits patchy and weak SERPING1 staining in the cortical and hippocampal pyramidal neurons, brain tissue from patients with encephalitis demonstrates strong cytoplasmic expression of this protein in a population of reactive astrocytes (as confirmed by staining for glial fibrillary astrocytic protein) and to a lesser extent in a subpopulation of nearby neurons. In contrast, infarct tissue shows faint cytoplasmic staining for SERPING1 in reactive astrocytes and neighboring neurons. These results confirm that SERPING1 expression is upregulated to a greater extent in inflammatory compared to ischemic astrogliosis and suggest that SERPING1 may be a clinically useful marker of neuroinflammation in biopsies from patients with an unknown neuropathologic process. Given the recently established role of complement in synapse pruning, increased expression of SERPING1 by neurons and reactive astrocytes may be important for limiting the excessive synapse loss in neuroinflammatory conditions.

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43

Two Cases of Fatal Cerebral Edema with CNS Angiitis

Denise Ng1, Harry Vinters1. 1University of California, Los Angeles

We report two cases of fatal cerebral edema arising as a complication of two different types of central nervous system angiitis. CASE 1: A 27-year-old male presented with flu-like symptoms, fever, truncal rash and myalgia after treatment of cutaneous abscess. Six days after his initial presentation, he developed diarrhea, nausea, vomiting, altered mental status and a witnessed grand mal seizure. Despite initial CT head being unremarkable, he had another seizure during a lumbar puncture. His repeat imaging showed severe global cerebral edema with cerebellar tonsillar herniation. His other laboratory investigations showed mildly elevated eosinophil count and CSF protein of 406 mg/dL. His medical history includes previous episode of meningoencephalitis with rash of unknown etiology 10 years previously. Autopsy brain examination shows bilateral cerebellar tonsillar herniation with midbrain compression and multifocal subacute vasculitis, consistent with primary angiitis of the central nervous system. CASE 2: A 71-year-old woman with past medical history of quadriplegia for severe spinal stenosis, rheumatoid arthritis, osteoporosis, and hypertension presented with headache and altered mental status for approximately 3 days. Initial CT head showed right temporal lobe edema and right to let midline shift. She was treated with steroids for presumed acute disseminated encephalomyelitis. She became more obtunded and later hypothermic with unstable blood pressures. Repeat imaging showed worsening cerebral edema. At autopsy, the brain showed edema with Duret hemorrhages. Focally severe granulomatous angiitis in vessels immunoreactive for Abeta was diagnostic for Abeta-related granulomatous angiitis. CONCLUSION: Although both ABRA and PCNSV have distinct clinicopathological features, both cases show that vascular injury can cause impairment of mechanisms important in fluid regulation. Cerebral edema is an uncommon manifestation of vasculitis in the absence of hemorrhage, and may be a result of impaired blood-brain barrier function associated with the angiitis.

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44

Autoimmune Polyglandular Syndrome Type 1 with Calcification in the Cortical Gray and White Matter and the Basal Ganglia

Fahad Bafakih1, Paul Benson2, M. Beatriz Lopes1. 1University of Virginia; 2Office of the Chief Medical Examiner of Virginia - Western Office

Autoimmune polyglandular syndromes are rare endocrinopathies that are associated with abnormal endocrine functions and nonendocrine autoimmune dysfunctions. Autoimmune polyglandular syndrome (APS) type 1 presents in childhood and might show other manifestations of disease as mucocutaneous candidiasis and ectodermal dystrophy. The disease is caused by mutations in the autoimmune regulator gene (AIRE) coding for a putative transcription factor. Autoimmune polyglandular syndrome type 2 is seen in adults and is more common than type 1, presenting with autoimmune thyroid disease, adrenal insufficiency and diabetes mellitus type I, and is not associated with AIRE gene mutation. Autoimmune polyglandular syndrome type 3 presents with autoimmune thyroid disease and other autoimmune disease with the exception of adrenal involvement. There is no known treatment for these diseases and management consistent mainly of hormone replacement and supportive therapy.

Herein, we report an autopsy of a 27-year-old female with APS type 1 with extensive calcification in the cortical gray and white matter and basal ganglia. The clinical presentation during early childhood included hypoglycemia, hypoparathyroidism, and adrenal crisis. She received multiple hormonal replacement therapies. At age 17, she had an episode of mental status changes thought at the time to likely be multifactorial, a combination of hypoglycemia, HSV meningitis and, perhaps, adrenal crisis. Head CT scan at that time showed extensive calcification of the basal ganglia and scattered calcification of the cortical gray and white matter located in the fronto-parietal lobes that had been documented prior to this episode. The patient was lost in follow-up. At age 27, she was found unresponsive at home and a forensic autopsy was performed. The mechanism resulting the extensive calcification of the deep gray matter is believed to be due to hormonal imbalance specifically hypoparathyroidism and adrenal insufficiency.

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45

Comparative Analysis of Neuronal Vulnerability of the Anterior Cingulate Cortex in Hereditary and Sporadic Tauopathies

Adrian Oblak1, Jill Murrell1, Rose Richardson1, Francine Epperson1, Bernardino Ghetti1. 1Indiana University School of Medicine, Dept Path and Lab Med

In hereditary Frontotemporal Dementia and Parkinsonism linked to chromosome 17 associated with the Microtuble Associated Protein Tau (MAPT) mutations (FTDP-17MAPT), specific neuronal populations may be more vulnerable than in sporadic tauopathies. We have focused on the analysis of hereditary and sporadic tauopathies characterized by 4-repeat (4R) tau. Brains of subjects carrying the following MAPT mutations: IVS10+3, IVS10+16, and P301L were used as well as brains with a neuropathologic diagnosis of corticobasal degeneration (CBD) and progressive supranuclear palsy (PSP). Histologic methods included hematoxylin and eosin with Luxol fast blue and Thioflavin S. For immunohistochemistry, antibodies raised against tau protein (AT8, 4R tau and 3R tau) and glial fibrillary acidic protein were used. Grey level index acquisition was used to determine the relative density and distribution of tau-immunoreactive (tau-ir) neurons within cortical layers of the anterior cingulate cortex. Furthermore, neuronal loss and gliosis in the same area was assessed. Tau-ir neurons were more frequently observed in layer II than in layers I, III, V or VI for both the IVS10+3 and IVS10+16 mutation. In the P301L mutation, tau-ir neurons were most numerous in layers II-III and VI than in I and V. Overall, in cases with the IVS10+16 mutation, there were more tau-ir neurons in all layers as compared to the cases with the IVS10+3 and P301L mutation. In CBD and PSP, tau-ir neurons were distributed evenly throughout cortical layers. Overall fewer tau-ir neurons were observed than in hereditary cases. The present findings demonstrate that tau is differentially distributed throughout the anterior cingulate cortex in the cases of hereditary and sporadic tauopathies examined. Cases of FTDP-17MAPT are more likely to demonstrate susceptibility of cortical layers in the anterior cingulate cortex when compared to sporadic tauopathies.

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46

Brainstem Pathology in Frontotemporal Degeneration Associated with the MAPT P301L Mutation

Melissa Gener1, Jill Murrell2, Adrian Oblak2, Bernardino Ghetti2. 1Indiana University School of Medicine; 2Indiana University School of Medicine Dept of Pathology and Lab Med

Among mutations in the Microtubule Associated Protein Tau gene, P301L is one of the most frequent. We studied a case characterized by a long clinical course, extremely severe cortical and subcortical atrophy, and prominent involvement of the brainstem.

A 44-year-old woman presented with sudden-onset of delusions and hallucinations. Retrospectively, disease onset may have been insidious, beginning at age 40. She was first treated for a psychiatric disorder; however, initial symptoms were followed by a rapid cognitive decline. At age 46, imaging studies revealed hypometabolism and atrophy of the frontal and temporal lobes, bilaterally. She continued to deteriorate neurologically and was admitted to a long-term nursing facility, where she died at 53 years of age.

The brain and spinal cord were obtained at autopsy. The brain was extremely atrophic and weighed 652 grams. Atrophy was particularly severe in the frontal and temporal lobes, as well as in the brainstem. Histological examination revealed marked neuronal loss and gliosis in the frontal and temporal cortices. In the midbrain, only a few neurons had survived in the substantia nigra. There was a significant loss of myelinated axons in the hemispheric white matter and in descending tracts. Immunohistochemistry was carried out using a panel of antibodies to tau including AT8, 4 repeat, and 3 repeat. Neuronal and glial tau were immunolabeled with AT8 and 4 repeat tau antibodies. Tau deposits were most numerous in the cortex of the frontal and temporal lobes, amygdala, dorsal midbrain, locus coerleus, median raphe nucleus, red nucleus, and the dorsal motor nucleus of vagus nerve. DNA, obtained from autopsy brain tissue, revealed a C to T nucleotide substitution at codon 301 resulting in a leucine for proline change (P301L).

The significance of 4 repeat tau pathology, in the brainstem as it relates to autonomic functions may need further investigation.

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47

Effect Of Salvadora Persica Leaf Extract In Transgenic Drosophila Model Of Parkinson’s Disease

Yasir Siddique1, Tanveer Beg2, Falaq Naz1, Smita Jyoti1. 1Dept of Zoology, Aligarh Muslim University, Aligarh (UP), India; 2Biology Dept,Faculty of Science, Jazan University, Jazan, Saudi Arabia

Background: There are various genetic models of PD based on alpha-synuclein (αS) expression (mutant, or wild type) in mice as well as in flies. The over expression of either wild type or mutant form of αS in transgenic Drosophila leads to the formation of Lewy Bodies (LB) resulting in the loss of dopaminergic neurons and behavioral abnormality. Plants having medicinal properties have gained importance because of their beneficial effect on humans. In this context the effect of Salvadora persica leaf extract was studied on the transgenic Drosophila (Parkinson’s disease) model flies expressing normal human alpha synuclein (h-αS) in the neurons.

Methods: The leaf extract was prepared in acetone. The flies were cultured on standard Drosophila food at 25°C. Crosses were set up using six virgin females of UAS-Hsap/SNCA.F5B were mated to three males of GAL4elav. The progeny expressing the human α-synuclein (PD flies) were exposed to 0.1, 0.5, and 1.0 μl/ml of S. persica mixed in the diet for 24 days. Hsap/SNCA.F strains were taken as control. The effect of extract was studied on the climbing ability, behavioral pattern, oxidative stress and apoptosis in the brain of PD model flies.

Results: The exposure of flies to 0.1, 0.5 and 1.0 μl/ml showed a dose dependent significant delay in the loss of climbing ability and activity pattern, reduction in the oxidative stress and apoptosis in PD model flies.

Conclusion: S. persica leaf extract is potent in reducing PD symptoms.

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48

Effect Of Gingerol On The Oxidative Stress In The Brains Of Transgenic Drosophila Model Of Parkinson’s Disease

Tanveer Beg1, Yasir Siddique2, Smita Jyoti2, Falaq Naz2, Rahul Sachdev2. 1Biology Dept,Faculty of Science, Jazan University, Jazan, Saudi Arabia; 2Dept of Zoology, Aligarh Muslim University, Aligarh (UP), India

Background: Oxidative stress is thought to be mechanistically involved in PD pathogenesis. Gingerol is a component of ginger and may have anti-oxidant properties. We studied the effects of gingerol supplementation on oxidative stress in Parkinson’s disease (PD) model flies.

Methods: Drosophila expressing human alpha-synuclein (PD flies) were exposed to 50, 100 and 150μM of gingerol mixed in the diet for 24 days. To measure lipid peroxidation, brain homogenates (10 brains/group; five replicates/group) were prepared in Tris HCl (20 mM), mixed with 1-methyl-2-phenylindole, acetonitrile, methanol and HCl, and incubated at 45°C for 40 min prior to absorbance measurements at 586 nm. The protein carbonyl content was estimated according to the protocol described by Hawkins et al. (2009). Homogenates were mixed with 2, 4-dinitrophenyl hydrazine and incubated for 20 min followed by addition of trichloroacetic acid (TCA). Following incubation at -20°C for 15 min, mixtures were centrifuged and the pellet obtained was washed twice by ice cold ethanol:ethylacetate (1:1). Pellets were re-dissolved in guanidine hydrochloride and the absorbance was read at 370 nm.

Results: Gingerol supplementation resulted in a dose dependent significant decrease in lipid peroxidation ranging from a 30% decrease at 50 μM gingerol to a 59% decrease at 150 μM gingerol (p<0.05). Gingerol supplementation also resulted in a dose dependent significant decrease in protein carbonyl content ranging from a 17% decrease at 50 μM gingerol to a 28% decrease at 150 μM gingerol (p<0.05).

Conclusion: Gingerol is potent in reducing the oxidative stress in the brains of PD model flies. Gingerol should be validated in mammalian model systems before use as a potential therapy for PD in humans.

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49

Apigenin Extends Lifespan And Improves The Activity Pattern Of Parkinson’s Disease Model Flies

Yasir Siddique1, Tanveer Beg2, Falaq Naz1, Smita Jyoti1, Rahul Sachdev1. 1Dept of Zoology, Aligarh Muslim University, Aligarh (UP), India; 2Biology Dept, Faculty of Science, Jazan University, Jazan, Saudi Arabia

Background: The Drosophila model for Parkinson’s Disease (PD) based on wild-type alpha synuclein expression in flies were used in this study. A time dependent loss of dopaminergic neurons and the formation of intracellular aggregates of αS (Lewy bodies) have been reported in the PD model flies. The effect of apigenin, an antioxidant found in most vegetables and fruits, was studied on the life span and activity pattern of the PD model flies.

Methods: Flies were maintained on standard Drosophila food at 25°C. Crosses were set up using six virgin UAS-Hsap/SNCA.F5B females mated to three GAL4elav males. The progeny expressing human α-synuclein (PD flies) were exposed to 0.1, 0.5 and 1.0 μl/ml of apigenin mixed in the diet. Hsap/SNCA.F strains were used as controls. To determine lifespan, the newly enclosed male flies (control and PD) were placed in culture tubes (10 flies per tube) containing 0.1, 0.5, and 1.0 μl/ml of apigenin mixed in diet. Flies were transferred to new diet every 3 days and the numbers of dead flies were recorded at 3 days intervals until all flies had died. Motor activity was monitored by using Drosophila activity monitors (DAMs) starting on the 12th day. Activity was recorded every hour for a total of 280 hrs.

Results: The results obtained for lifespan determination showed that apigenin extended the average lifespan of the male PD flies. A dose dependent delay in the loss of activity pattern was observed in PD flies exposed to 0.1, 0.5, and 1.0 μl/ml of apigenin. No change was observed in the activity pattern of control flies.

Conclusion: Apigenin is potent in reducing PD symptoms in the Drosophila model for PD.

(The grant (No.F.30-1/2013 (SA-II)/RA-2012-14-GE-UTT-858) received as UGC Research Award (2012-14) from the University Grants Commission, New Delhi is thankfully acknowledged).

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50

Death of Neurons in the Substantia Nigra Prior to Incidental Lewy Body Disease

Eric Richfield1, Kavita Prasad1, John Hedreen2. 1Robert Wood Johnson Medical School/Rutgers Universtity; 2Mailman Research Center

We used the quantitative assessment of several measures from a cross section through the substantia nigra of different groups of individuals. Measures included counts of tyrosine hydroxylase (TH+) and neuromelanin (NM+) expressing and non-expressing neurons, TH+ and phosphorylated α-synuclein+ (pαSYN+) neurites, and three NM-related measures. The groups included age-matched elderly controls, patients with Incidental Lewy body disease (ILBD) and the Parkinson’s disease phenotype (PDP). We used one tissue section at the level of the 3rd nerve stained for TH and pαSYN. We divided all cases into seven distinct grades based on quantitative data. The grades included control, asymptomatic individuals (PDP0a, PDP0b, and PDP0c), and symptomatic PDP patients (PDP1, PDP2, and PDP3). The grades demonstrated a progression of disease severity. Some measures declined with progression/duration of disease (TH+/NM+ neurons), some measures increased (ratio of pαSYN+/TH+ neurites), and many measures were biphasic: increasing and then declining (NM-related). Various measures of NM (representing loss of pigmented neurons) demonstrated that cell death began at a grade of disease prior to the presence of Lewy bodies. Cell death was also present in ILBD. The pathology of the PDP measured in this poster demonstrates this is a dynamic and progressive disease. The interpretation of post mortem findings will require assessment of grade to be useful.

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51

Effect of Lewy Bodies on Substantia Nigra Dopaminergic Neuron Cell Health

Knarik Arkun1, Ann Rice2, Essie Komlada2, James Bennett2. 1Virginia Commonwealth University Medical Center; 2VCU Parkinson’s and Movement Center

Parkinson’s disease is the second most common neurodegenerative disease. Lewy bodies and Lewy neurites with premature neuronal loss are hallmarks of the disease. It is still unclear what triggers neuronal loss. In this study we would like to determine if Lewy bodies are beneficial or deleterious to neuronal health by measuring mtDNA copy numbers in cells with and without Lewy bodies in the substantia nigra. Initial studies used paraffin embedded H&E stained sections from the substantia nigra of four Parkinson’s patients. 15 neurons with and without Lewy bodies by laser capture microdissection (LCM) from each case in triplicate. DNA was extracted and qPCR was performed for 4 genes (ND2, ND4, COX3 and 12S) to measure the neuronal well-being on the mitochondrial level. We found no difference in the expression levels of all four genes between cells with and without Lewy bodies in each case. However, there were differences of mtDNA copy number expression between the cases and between the genes. In summary, the preliminary data shows no difference in mtDNA copy numbers in cells with and without Lewy bodies, which indicates that Lewy body inclusions are not deleterious to mitochondria. A subsequent study using immunohistochemistry for α-synuclein and fresh frozen tissue is being conducted to elucidate the influence of the presence of Lewy bodies on mtDNA copy number.

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52

Evaluation of Lewy Pathology in Enteric Neurons from Living Patients

Annie Hiniker1, Ryan Gill1, Chadwick Christine1, Robert Nussbaum1. 1University of California San Francisco

Diminished GI motility, resulting in constipation, is often one of the earliest clinical manifestations of Parkinson’s disease (PD) and can precede PD diagnosis by years. Additionally, autopsy series clearly demonstrate the presence of Lewy pathology in enteric neurons in almost all PD patients. We and others have hypothesized that enteric neurons may show Lewy pathology prior to the diagnosis of PD, making GI biopsies such as those obtained during colonoscopy a potential diagnostic tool. To examine this hypothesis, we are examining all GI surgical and biopsy specimens from patients with clinical diagnosis of PD at our institution. As proof of principle, we first examined colonic resection specimens from patients with an ICD-9 coded diagnosis of PD (332.0). Six resections from patients with this diagnosis who also fit our criteria for PD were compared to matched (age, gender, reason for resection) control resections. All resection specimens demonstrated submucosal and myenteric neurons. Immunohistochemical staining with anti-phosphorylated alpha-synuclein antibody revealed rare Lewy bodies in neurons of 5 of 6 PD cases (predominantly myenteric plexus neurons) and no Lewy bodies in the six matched control cases. We next searched for all colonic biopsy specimens from PD patients; this yielded twelve biopsies. However, only 4/12 biopsies demonstrated neurons. These samples are currently being evaluated for Lewy pathology. Finally, an additional search for surgical specimens from other GI locations known to demonstrate Lewy pathology at autopsy, including salivary gland, esophagus, stomach, and small intestine, was performed and yielded 27 additional cases, currently being evaluated for neuron content. Our initial data suggest that Lewy pathology is present in the GI tract of living patients but that random biopsies may be limited in their diagnostic utility due to the rarity of diagnostic pathology.

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53

Hippocampal sclerosis in Lewy body disease

Naoya Aoki1, Melissa Murray2, Kotaro Ogaki2, Shinsuke Fujioka2, Owen Ross2, Dennis Dickson2. 1Mayo Clinic Neuropathology Laboratory, Jacksonville, Florida; 2Mayo Clinic

Hippocampal sclerosis (HpScl) is a common neurodegenerative disorder in the elderly, which occurs alone or accompanied by other disorders, especially frontotemporal lobar degeneration with TDP-43 pathology (FTLD-TDP). It is increasingly clear that TDP-43 pathology is present in most cases of HpScl. TDP-43 pathology is also common in Alzheimer disease (AD) and has also been reported in Lewy body disease (LBD). So far, no study has focused on the frequency and characteristics of HpScl in LBD. Our goals were (1) to clarify the pathological characteristics of HpScl in LBD, and (2) to determine the importance of TDP-43 pathology in the diagnosis of HpScl. We examined 670 LBD cases (381 diffuse LBD and 289 transitional LBD). All cases were evaluated with thioflavin S fluorescent microscopy for AD pathology, as well as immunohistochemistry for α-synuclein and TDP-43. HpScl was detected in 34 cases (5%). Older age, higher Braak neurofibrillary tangle stage and Thal amyloid phase, as well as the presence of TDP-43 pathology were associated with HpScl. There was no difference in the frequency of HpScl between transitional and diffuse LBD. TDP-43 pathology was detected in all HpScl cases and 116/493 (24%) of cases without HpScl. All HpScl cases had a pattern of TDP-43 pathology consistent with FTLD-TDP Type A; however, the pathology were restricted to limbic and inferomedial temporal lobe in 85% of the cases. All HpScl cases had TDP-43 pathology in the parahippocampal gyrus and 88% had neuronal intranuclear inclusions. Fine neurites in CA1 sector were detected in all HpScl cases. We conclude that HpScl in LBD is a TDP-43 proteinopathy associated with older age and greater AD pathology, but not α-synuclein distribution. The results suggest that neuronal loss in HpScl is not merely due to AD pathology, but may potentially be due to the same underlying mechanism of FTLD-TDP.

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54

Flow Cytometry Analysis of Synaptosomes from Human Brain Reveals Changes Specific to Lewy Body and Alzheimer’s Disease

Nadia Postupna1, C Keene1, Caitlin Latimer1, Emily Sherfield1, Rachel Van Gelder1, Jeffrey Ojemann1, Thomas Montine1, Martin Darvas1. 1University of Washington

Synaptic dysfunction is thought to play an important role in the pathophysiology of neurodegenerative diseases, such as Alzheimer’s disease (AD) and Lewy body disease (LBD). To improve our understanding of synaptic alterations in health and disease, we investigated synaptosomes prepared from post-mortem human cerebral cortex, putamen, and two regions of the caudate nucleus, dorso-lateral (DL) and ventro-medial (VM), regions commonly affected in AD and LBD. We observed that the fraction of synaptosomal particles with reactivity for dopamine transporter (DAT) was significantly reduced in the putamen and VM caudate of patients with neuropathological diagnosis of LBD. As expected, these differences also were reflected in direct measurements of dopamine (DA) and its metabolite, 3,4-dihydroxyphenylacetic acid (DOPAC), in caudate and putamen of LBD patients. The fraction of synaptosomal particles positive for amyloid β (Aβ) was significantly increased in frontal cortical samples of patients with the neuropathological diagnosis of severe AD, and was positively correlated with disease progression. We also prepared synaptosomes from the striatum of mice with severe loss of DA neurons (Slc6a3-DTR mice) and wild-type littermate controls. We observed dramatically reduced levels of DAT-positive synaptosomes in Slc6a3-DTR mice following exposure to diphtheria toxin (DT). Striatal levels of DA and DOPAC in Slc6a3-DTR mice also were reduced significantly following DT exposure. We conclude that flow cytometric analysis of synaptosomes prepared from human or mouse brain provides an opportunity to study dopaminergic and glutamatergic synaptic content in tissue and detect pathological changes at the level of the synapse in LBD as well as AD.

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55

Coincident Alzheimer’s Disease Modifies Alpha-synuclein Pathology in Lewy body Disease

Pallavi Gopal1, Jon Toledo2, Kevin Raible2, Erin Abner3, David Irwin2, Johannes Brettschneider4, Steven Arnold5, Howard Hurtig5, Peter Nelson3, Charles Adler6, Thomas Beach7, John Trojanowski2. 1Hospital of the University of Pennsylvania; 2Center for Neurodegenerative Disease Research; 3Sanders-Brown Center on Aging, University of Kentucky; 4University of Ulm, Center for Neurodegenerative Disease Research; 5Department of Neurology, University of Pennsylvania; 6Parkinson’s Disease and Movement Disorders Center, Mayo Clinic; 7Banner Sun Health Research Institute

Alzheimer’s disease (AD) and Parkinson’s disease (PD) are the two most common, frequently co-occurring, neurodegenerative diseases. Dementia with Lewy Bodies (DLB) is defined by dementia with fluctuating cognitive symptoms and the appearance of a parkinsonian syndrome >1 year after dementia onset. Staging systems have been proposed for tau and Aβ deposits in AD as well as for alpha-synuclein pathology (ASP) in PD and DLB although no consensus has been reached.

We studied the distribution of ASP in DLB subjects with coincident AD (AD+DLB, n=313) and compared it to the distribution in PD cases with and without AD (PD+AD, n=71; PD, n=134) from the University of Pennsylvania (UPenn) and the Banner Sun Health Research Institute. Cases were classified by ASP distribution into amygdala predominant, brainstem predominant, limbic (with/ without brainstem involvement) and neocortical using a modified Unified Staging System for Lewy Body Disorders scheme. Dopamine transporter immunohistochemistry was performed on a subset of the UPenn cohort.

Amygdala predominant ASP category was present in AD+DLB cases and not PD cases. The limbic category presented a lower burden of brainstem, subcortical and frontal ASP in the DLB cases compared to the PD cases. In the neocortical category, PD only cases showed a lower burden of temporal and angular cortex ASP than the PD+AD and the DLB only cases. PD and PD+AD groups in the UPenn cohort showed a higher burden of ASP in the substantia nigra compared to the DLB only group. The nigro-striatal pathway, defined by dopamine transporter immunohistochemistry, was relatively preserved in the DLB+AD groups compared to the PD only and PD+AD groups.

Conclusions: Presence of AD pathology modifies the burden and distribution of ASP in the alpha-synucleinopathies studied. The different patterns of ASP spread may explain the heterogeneity observed when classifying the different patterns of distribution of Lewy body pathology.

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56

Clinicopathologic Correlations of FTLD-TDP Type B with High and Low Burden of TDP-43 Positive Inclusions in the Dentate Gyrus

Esther Bit-Ivan1, Anne Koronkiewicz1, Melanie Peterson1, Alfred Rademaker1, Mallory Ward1, Qinwen Mao1, Sandra Weintraub1, Nailah Siddique1, Teepu Siddque1, M. Marsel Mesulam1, Eileen Bigio1. 1Northwestern University Feinberg School of Medicine

The FTLD-TDP harmonized classification divides FTLD-TDP into four groups, Types A through D, based on the location, type, and density of TDP-43 positive inclusions, which correlate with clinical presentation and genetic findings. Type B, which generally presents with behavioral bvFTD +/- ALS, was observed to have few to numerous neuronal cytoplasmic inclusions in the dentate gyrus, with density inversely proportional to cortical TDP. To date, the significance of this difference in dentate gyrus (DG) TDP-43 inclusion burden has not been studied. The aim of the current study is determine whether DG TDP-43 inclusion burden correlates with specific clinical or pathologic factors. Thirty-two cases with the final diagnosis of FTLD-TDP Type B were selected and medical records were reviewed. All 32 cases also either had both clinical and pathologic ALS or ALS pathology without clinical ALS. H&E slides were assessed for neuronal loss and gliosis (NL&G) of the hippocampal formation, dentate molecular layer rarefaction, and degenerative changes in the upper and lower motor neuron pathways. Immunohistochemical stains for TDP-43 of the hippocampus and other regions were performed. Dentate gyrus inclusions were graded as 0=none, 1=rare, 2=mild, 3=moderate or 4=frequent. Nine of 32 cases had low TDP-43 burden (Group 1, grades 0 to 2), and 23 cases had high TDP-43 burden (Group 2, grade 3 and 4). p62 immunostains were performed to look for C9orf72-specific pathology. Results showed that Group 2 cases more commonly had dementia at onset of disease (p = 0.007) and greater subicular NL&G (p = 0.012) than did Group 1 cases. Other trends in Group 2 cases included lower brain weight and greater CA1 NL&G. There was no correlation with duration of disease, age, presence of memory problems, gender, cortical TDP pathology, or C9orf72 hexanucleotide repeat pathology. Investigation into the significance of these findings is warranted.

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57

Clinicopathologic report of ocular involvement in ALS patients with C9ORF72 Mutation

Eileen Bigio1, Amani Fawzi1, Esther Bit-Ivan1, Joseph Simonett1, Patryk Purta1, Heather Moss2, Nailah Siddique1, Nicholas Volpe1, Teepu Siddque1. 1Northwestern University Feinberg School of Medicine; 2University of Illinois at Chicago

Recently, a GGGGCC hexanucleotide repeat expansion in intron 1 of the C9orf72 gene was identified as the most common pathologic mutation in familial and sporadic ALS/FTD, familial ALS, and familial FTD. Multiple unique pathologic features have been identified in brains of c9orf72 mutation carriers (c9-ALS/FTD), including star and dot-shaped p62 positive, p-TDP-43 negative neuronal cytoplasmic and intranuclear inclusions in the cerebellum, hippocampus, and neocortex. These inclusions also contain dipeptide repeat proteins, likely formed from non-conventional, non-ATG initiated translation of the expanded hexanucleotide repeat. Neuro-ophthalmologic changes have been described in ALS patients, but histopathologic studies of ocular involvement have been limited to studies of oculomotor and trochlear nuclei. In our study of a patient with C9orf72 mutation, histologic examination of the brain revealed ALS, no FTLD-TDP, and p62 positive inclusions typical of c9 FTD-ALS. Examination of visual pathway related regions showed no histologic abnormality. p62 showed inclusions in the occipital cortex, as has been reported in c9 cases, but the remainder of the visual system had no p62 pathology and there were no TDP-43 positive inclusions. Immunofluorescent labeling studies of retinal sections demonstrated abundant dot and semi-lunar shaped p62-positive inclusions that were localized to the inner nuclear layer (INL) in the macula and peripheral retina. Most inclusions were found in a peri-nuclear location and were seen at all levels of the INL, with rare inclusions also seen in the inner plexiform and ganglion cell layer. No p62+ inclusions were seen in a control retina. Inclusions were also labeled with antibodies to poly-GA dipeptide repeat which co-localized with the p62 inclusions, and with ubiquitin, but not ubiquilin-2 or TDP-43. This is the first report that identifies disease-specific deposits of ALS/FTLD in the retinal tissue of a patient with C9orf72 mutation, in whom corresponding ocular functional deficits were identified prior to her passing.

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58

Incidental, Insidious and Late Onset Neuronal Intermediate Filament Inclusion Disease in an 85-Year-Old Male – a Case Report

Jason Chiang1, Lananh Nguyen1, Nigel Cairns2, Oscar Lopez3, Julia Kofler4. 1Univ Pittsburgh Medical Center Department of Pathology; 2Washington University in St. Louis, Department of Neurology; 3University of Pittsburgh Department of Neurology; 4Univ Pittsburgh Medical Center Department of Pathology

Neuronal intermediate filament inclusion disease (NIFID), a subgroup of frontotemporal lobar degeneration with FUS-positive inclusions (FTLD-FUS), usually occurs in individuals younger than 60 years with short disease duration (3-7 years). The inclusions are typically present abundantly in the neocortex, hippocampus and basal ganglia.

Here we report incidental findings of NIFID in an 85-year-old male with a 19-year history of progressive memory deficits and depression, but no significant motor symptoms except for mild action tremors. Neuropathologic evaluation using NIA-AA criteria revealed a high level of Alzheimer’s disease (AD) neuropathologic change (A3, B3, C3) and amygdala-predominant Lewy body disease. In addition, autopsy revealed widespread presence of round eosinophilic neuronal cytoplasmic inclusions, which were strongly positive for alpha-internexin and p62 but largely negative for tau and alpha-synuclein. The highest frequency of inclusions was found in amygdala and claustrum. Within cortical regions, deeper layers of the primary visual cortex were most densely affected, followed by pre- and postcentral gyri. Other cortical areas showed only sparse inclusions. Moderate numbers of inclusions were found in hippocampal CA3 and CA1 areas and substantia nigra. Rare inclusions were seen in the basal ganglia, thalamus, inferior olive, colliculi and medullary tegmentum. The inclusions exhibited variable immunoreactivity for FUS, ranging from negative to weakly positive.

As the patient’s symptoms are sufficiently explained by the AD and Lewy body pathologies, the NIFID changes appear to be an incidental finding of uncertain significance. Several features including older age, longer disease duration, absence of motor symptoms and limited cortical and basal ganglia involvement distinguish this case from typical NIFID. Similar weak FUS reactivity as in our case has been described for a few cases of NIFID in older patients. In conclusion, this case broadens the spectrum of NIFID and suggests that NIFID in older individuals may represent a biochemically and phenotypically distinct variant.

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59

Adult-Onset Alexander Disease in a Paraplegic African-American Male with a Rare D138N Mutation in the GFAP Gene

Ki-Eun Chang1, Nancy Edwards2, Bibhuti Mishra3, Mark Hallett2, Abhik Ray-Chaudhury4. 1Penn State College of Medicine/NIH; 2NINDS, NIH; 3Inova Fairfax Hospital; 4NIH

Contrary to the fatal form of infantile Alexander Disease, Adult-onset Alexander Disease (AOAD) is a unique variant, characterized by a protracted clinical course, dominant mutation of the GFAP gene, atrophy of the brainstem and upper cervical spinal cord, and the presence of Rosenthal fibers on histology. Although the overall incidence is rare, complications of AOAD often mimic common and/or devastating disorders, subjecting the patient and medical caregivers to undue anxiety and clinical conundrum. Also, unfamiliarity with the heritable pattern of AOAD may lead to improper follow-up of potentially affected family members. Here, we report a case of AOAD in a 52 year-old African-American man with relevant clinical history, MRI findings, histology, and a GFAP mutation (c.382 G>A 9p.Asp28Asn) that has only been reported once previously.

Our patient had an extensive medical history with complications attributable to bulbar and pyramidal pathology such as paraplegia, chronic aspiration pneumonitis, and recurrent urinary tract infections. MR imaging revealed severe atrophy of the medulla and upper cervical cord. Final diagnosis was confirmed by the presence of large number of Rosenthal fibers, scattered mostly in a perivascular pattern over multiple regions of the brain on post-mortem examination. Foci of white matter demyelination were also observed. We searched the current literature and mutation databases and found only a single account of the same mutation in a middle-aged man. Interestingly, both patients were older at symptom presentation and suffered mainly from pyramidal involvement. The ethnicity of the other patient was not explicitly stated, which is not an uncommon scenario in most reports describing AOAD. Not only does this make our case potentially unique, but further highlights the principle that genetic confirmation should be pursued if presented with characteristic MRI findings in the right clinical context and should not be deterred by the age or ethnicity of the patient.

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60

GABA-ergic and Glycinergic Synaptic Deficit in Friedreich Ataxia

Arnulf Koeppen1, R Ramirez1, Alyssa Becker1, Joseph Mazurkiewicz2. 1VA Medical Center; 2Albany Medical College

Progressive atrophy of large glutamatergic neurons of the dentate nucleus (DN) is a characteristic central nervous system lesion of Friedreich ataxia (FRDA), and loss of cerebellar output is the pathophysiological correlate of disabling ataxia. Most afferent fibers in the DN derive from Purkinje cells that effect inhibitory synaptic transmission by gamma-aminobutyric acid (GABA). In addition, DN neurons receive inhibitory glycinergic impulses from intrinsic DN nerve cells. Grumose degeneration in the DN of patients with FRDA consists of clusters of abnormal axon terminals that react with antibodies to synaptophysin and glutamic acid decarboxylase (GAD). Grumose degeneration is more prominent in cases with partial preservation of large neurons but may be entirely absent in cases of long disease duration. Enzymatic GAD activity is the rate-limiting step in the biosynthesis of GABA, and GAD reaction product identifies neuronal cell bodies, axons, and axon terminals as GABA-ergic. Small GABA-ergic nerve cells in the DN are the principal source of GABA-ergic afferents in the contralateral inferior olivary nucleus. Glycinergic neurons are recognizable by their reaction with an antibody to the glycine transporter 2. Positive-contrast immunohistochemistry and double-label immunofluorescence of paraffin-embedded archival specimens of FRDA show that grumose degeneration represents proliferation of GABA-ergic terminals and retraction from neuronal plasma membranes. The proposed mechanism is loss of GABA-A receptors due to failure of the postsynaptic protein gephyrin. Gephyrin positions GABA-A and glycine receptors to the plasma membrane and anchors them to the cellular cytoskeleton. Some gephyrin-positive small neurons remain even in advanced cases and display reaction product of GABA-A and glycine receptors. Persistence of small GABA-ergic neurons in the DN explains the lack of transsynaptic degeneration of the inferior olivary nuclei in FRDA.

(Supported by National Institutes of Health, R01NS069454; and Friedreich’s Ataxia Research Alliance).

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61

Neurpathologic Features of Spinocerebellar Ataxia 5 (Lincoln’s Ataxia.)

Margaret Flanagan1, Luis Gonzalez-Cuyar2, Jake Hemingway2, Zachary Hoffer2, Thomas Montine2, Thomas Bird2, C. Keene2. 1University of Washington Medical Center; 2University of Washington

Spinocerebellar Ataxia type 5 (SCA5) belongs to a heterogeneous group of autosomal dominant, progressive neurodegenerative disorders characterized by ataxia and cerebellar degeneration. SCA5 is caused by mutations in the Beta-III Spectrin gene (SPTBN2) located in chromosome 11. SCA5 progresses slower then other Spinocerebellar Ataxias and does not shorten lifespan. Herein we report an 87-year-old man with a clinical diagnosis of Spinocerebellar Ataxia type 5 (SCA5) diagnosed at age 48 who exhibited slowly progressive truncal and extremity ataxia (onset age 25), dysarthria, dysphagia, and intention tremor. His family history was significant for his father, five siblings, two children, and other relatives with similar ataxic symptoms. Postmortem examination revealed that the left half of the brain, brainstem and cerebellum weighed 580 grams with the left posterior fossa contents contributing 50 grams. Grossly the cerebellum was small with atrophy of the anterior vermis and the cerebellar peduncles. The inferior olivary nuclei appeared grossly unremarkable. Histologically there was diffuse loss of Purkinje cells with “empty baskets” in most areas of cerebellar cortex, a thinning of the molecular layer and mild granule cell loss. The inferior olivary nuclei were gliotic with mild-to-moderate neuronal loss. The basal pontine nuclei, red nuclei, cranial motor nuclei, posterior columns, Clarke’s columns and spinocerebellar tracts were unaffected. This is the second reported case of post mortem examination of SCA5 and, similar to the case reported by Clark and Ranum, our impression was that the pathology was that of a simple cerebellar cortical degeneration, principally targeted to Purkinje cells with other changes in granular neurons and inferior olives likely the result of long-term loss of Purkinje cells.

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62

Neuropathological Features of Early Onset Huntington’s Disease with Marked Cerebellar Atrophy

Caitlin Latimer1, Patrick Cimino2, Zachary Hoffer1, Louis Gonzalez-Cuyar1, Thomas Montine1, Thomas Bird1, C Keene1. 1University of Washington; 2Washington University

Early onset Huntington’s disease (EOHD), as its adult counterpart, is an autosomal dominant disorder with anticipation caused by trinucleotide (CAG) repeat expansion in the IT15 (Huntingtin) gene located in the short arm of chromosome four. It is defined as Huntington’s disease arising before age 21 and accounts for 5-10% of the total Huntington’s disease (HD) cases. Cases arising in patients younger than ten years of age have also been reported, and account for approximately 1% of the total cases. Clinically these patients differ from the adult patients in presenting with myoclonus, seizures, Parkinsonism and cognitive decline. Imaging but not neuropathological evaluations of EOHD cases with cerebellar atrophy are reported in the literature. Herein we report a case of a six-year-old boy with paternally inherited EOHD of 169 CAG trinucleotide repeats who presented at approximately age four with speech difficulties and developmental delay who progressed to medically intractable generalized tonic-clonic seizures. Gross neuropathological examination revealed mild frontal cortical atrophy, as well as atrophy of the caudate and putamen consistent with gross Vonsattel grade 3. There was cerebellar cortical atrophy mainly involving the medial folia (vermis). Microscopically, sections of the striatum showed gliotic gray matter nuclei with moderate neuron loss and gliosis corresponding to histologic Vonsattel grade 3 HD. There was substantial Purkinje cell loss and mild Bergmann gliosis. Huntingtin positive intranuclear inclusions were present in the striatum and frontal cortex. A review of the English literature shows that out of the reported cases of EOHD only four have been histopathologically characterized, all without cerebellar atrophy.

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63

Neurodegeneration in Bipolar Disorders

Ayako Shioya1, Kunimasa Arima2, Yukio Kakuta3, Takefumi Yuzuriha4, Akira Tamaoka5, Shigeo Murayama6, Yuko Saito7. 1Dept Neuropath, National Center Hosp. Neurol and Psyc; 2Dept of Psych, National Center Hospital of Neurology and Ps; 3Department of Pathology, Yokohama Rosai Hospital; 4Department of Psychiatry, Hizen Psychiatric Center; 5Dept of Neurol, Graduate School of Comprehensive Human Scienc; 6Brain Bank for Aging Research, Tokyo Metropolitan Institute of Geronto; 7National Center Hospital of Neurology and Psychiatry

Introduction: Neurodegeneration in mood disorder receives considerable attention. We examined patients with bipolar disorder (BPD), who died in late adulthood, in order to check any modification in age- related neurodegeneration as in chronic traumatic brain encephalopathy.

Materials and Methods: Eleven consecutive autopsy cases in our brain banks, who received definite clinical diagnosis of BPD, were employed for this study. Serial six- um- thick sections were obtained from archival paraffin blocks of representative areas. The sections were stained with H.E., K.B. and Gallyas- Braak silver method, as well as immunocytochemically with anti- phosphorylated tau, amyloid beta and alpha- synuclein andibodies. Age and gender- matched controls were selected for each case. Clinical information was retrospectively collected from medical charts.

Results: All cases were males and average age of death was 69 years of age. Average clinical course was 27 years. Four cases presented with cognitive impairment. Brain weight ranged from 1,116 to 1,325 g except for a case with hypoxic encephalopathy after hanging of suicidal attempt. Neuropathological diagnosis included dementia with grain (2), argyrophilic grain disease (2), corticobasal degenerarion (CBD) (1), Lewy body disease (LBD) (1), hypoxic encephalopathy (1), cerebral infarction (1) and unremarkable (3). All cases contained AGs with Stage 0.5 to III. Three cases died in the 50s and one fulfilled morphological criteria of dementia with Lewy body, limbic form. The other two cases, including one with suicidal attempt, showed AGs, abundant in the brain stem and amygdala. Two cases died in 60s and one showed AG preferentially in brain stem and amygdala and another in limbic predominance. The cases who died after 70s presented AG in limbic predominance like controls, except for one case with CBD.

Discussion & Conclusion: Our study showed that tauopathy or alpha- synucleionopathy is a pathological background of a certain group of BPD.

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64

Brain Biopsy in Dementia or Neurologic Decline of Unknown Etiology

Shino Magaki1, William Yong1, Negar Khanlou1, Harry Vinters1. 1Section of Neuropathology, UCLA Department of Pathology & Lab Medicine

Brain biopsies have an uncertain role in the diagnosis of patients with dementia or neurologic decline of unknown etiology. They are often performed as a last resort after an exhaustive panel of less invasive tests and procedures have failed to lead to a definitive diagnosis. The objective of this study was to evaluate the sensitivity of brain biopsies in this patient group through the retrospective analysis of 53 brain biopsies performed for neurologic disease of unknown etiology at a single tertiary care institution between December 2001 and December 2011. Patients with known nonlymphomatous neoplasms thought to be associated with the neurologic symptoms or with immunodeficiency were excluded from the study. The clinical presentation, imaging and laboratory tests were compared between diagnostic groups to identify factors more likely to yield a diagnosis. Sixty two percent of the biopsies were diagnostic (33 out of 53), with the most common histologic diagnosis of primary central nervous system lymphoma (PCNSL) in 14 of 53 patients (26% of total) followed by infarct in four cases (7.5%). A few of the patients were found to have rare unexpected diseases such as lymphomatosis cerebri, neurosarcoidosis, and neuroaxonal leukodystrophy. Complications from biopsy were uncommon and included infection with abscess formation at the biopsy site and hemorrhage. These results suggest that brain biopsies may be useful in difficult cases in which less invasive measures have been unable to reach a diagnosis.

(This study was supported in part by P50 AG16570).

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65

An Autopsy Case of Granulomatous Amoebic Meningoencephalomyelitis Caused by Balamuthia Mandrillaris in Japan

Hajime Miyata1, Kenju Hara2, Ken Saitoh3, Masae Ryufuku1, Haruka Ouchi2, Ken Shibano2, Kenji Yagita4, Hideaki Ishiguro2. 1Neuropathology, Research Institute for Brain and Blood Vessels - Akita; 2Neurology, Akita Red Cross Hospital; 3Pathology, Akita Red Cross Hospital; 4Parasitology, National Institute of Infectious Diseases

CNS infections caused by pathogenic free-living amoebae have mortality rate over 90%. These include primary amoebic meningoencephalitis caused by Naegleria fowleri and granulomatous amoebic meningoencephalitis caused by Balamuthia mandrillaris and Acanthamoeba species.

A 69-year-old Japanese man developed fever in mid-summer followed by transient unconsciousness on the next day. He has the clinical histories of membranous nephropathy since the age 35, hypertension and appendix cancer treated with the right hemicolectomy at the age 67. The possibility of basilar artery thromboembolism was considered based on the initial CT scan showing a hypodense area in the right mesial posterior temporal lobe. However, he gradually complained of headache and nausea. CSF examination and cytology 2 weeks after the onset revealed lymphoplasmacytosis, elevated protein (>200mg/dl) and decreased glucose (12mg/dl) contents, with negative culture for bacteria, negative PCR tests for Mycobacterium tuberculosis, and negative cryptococcal antigen. The possibility of metastatic cancer with carcinomatous meningitis was also considered based on the clinical course and laboratory examinations as well as the repeated CT scan showing enlargement of the lesion with partial contrast enhancement and obstructive hydrocephalus. The patient died 50 days after the onset. The autopsy brain and spinal cord showed multiple foci of fresh hemorrhagic and non-hemorrhagic softening, histologically consisting of extensive necrosis and granulomatous inflammatory changes with perivascular amoeba trophozoites and cysts immunohistochemically positive for Balamuthia mandrillaris. Trophozoites showed a single nucleus with occasional multiple nucleoli, and transmission electron microscopy demonstrated triple-walled cysts. No obvious foci of amoebic infection were detected in the visceral organs.

Eight of 10 reported cases with amoebic encephalitis in Japan since 1976 are caused by Balamuthia mandrillaris. Amoebic infection, particularly Balamuthia mandrillaris, should be included in the differential diagnosis of non-purulent meningoencephalitis showing lymphoplasmacytosis, elevated protein and decreased glucose contents in CSF, with a space-occupying CNS lesion on CT and MRI.

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66

A 29-year-old Pregnant Woman with Worsening Left Hemiparesis, Encephalopathy, and Hemodynamic Instability: A Case of SSPE

Gerald Reis1, Jana Ritter2, William Bellini2, Andrew Bollen1. 1University of California San Francisco; 2Centers for Disease Control and Prevention

A 29-year-old G3P2 woman was transferred to this hospital for progressively worsening encephalopathy, left hemiparesis, and hemodynamic instability. Her symptoms started approximately 6 weeks earlier when she developed weakness and gait problems. Examination at that time found left lower extremity rigidity, hyperreflexia, and a positive Babinski sign. MRI imaging of the brain and spine was unremarkable. Two weeks later she developed encephalopathy and worsening left-sided dysfunction involving both upper and lower extremities. Initial work-up for infectious and autoimmune causes at an outside hospital was negative, except that the cerebrospinal fluid (CSF) analysis showed an elevated IgG concentration of 25 mg/dL with ≥ 2 oligoclonal bands. She had an uncomplicated spontaneous delivery of a preterm healthy infant at 34 weeks and 2 days, but her cognitive function deteriorated further and she became hemodynamically unstable. A repeat brain MRI demonstrated generalized volume loss and multifocal cortical and subcortical T2 hyperintense lesions with evidence of corticospinal tract degeneration. The clinical differential diagnosis included inflammatory, vascular, infectious, neoplastic, paraneoplastic, and metabolic etiologies. She underwent a brain biopsy and additional CSF studies at our institution. The brain biopsy showed characteristic viral inclusions of the type seen in subacute sclerosing panencephalitis (SSPE), and this was confirmed by both immunohistochemical and electron microscopic evaluation. DNA sequence analysis at the Center for Disease Control demonstrated a close relationship to genotype D7. The patient received treatment with intrathecal interferon alpha 2B without improvement and died 2 months later. This case documents an approximately 6-month progression to death in a pregnant patient with SSPE.

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67

Balamuthia Amoebic Encephalitis, Diagnostic Challenges from Surgical Pathology to Autopsy: A Case Report

Keng-Chih Su1, Harry Vinters1, Negar Khanlou1, William Yong1, Annie Wu1, Wun-Ju Shieh2, Dianna Blau2, Atis Muehlenbachs2. 1Neuropathology Department, David Geffen School of Medicine at UCLA; 2Centers for Disease Control and Prevention (CDC), Atlanta, GA

Balamuthia mandrillaris (BM), with a case mortality rate of >98% nationally, is a fatal clinicopathologic mimic of treatable granulomatous encephalitides. We report a case of central nervous system (CNS) Balamuthia infection presenting initially with seizures. The decedent was a 63-year-old man California resident with an uncomplicated medical history and fully active until presentation. Post-ictal MRI of the brain with contrast showed multiple enhancing tumors involving predominantly the right cerebellar hemisphere, left gyrus, right temporal lobe, and left frontoparietal lobes. Initial brain biopsy was reported as extensively necrotic and therefore interpreted as non-diagnostic. A second biopsy targeting cerebellar lesion showed a necrotizing lymphohistiocytic lesion and abundant amoeba. Careful family interview revealed a history of several recent camping trips in central California including a visit to a bat cave. Patient deteriorated rapidly and died within two months of initial presentation. The autopsy showed multifocal hemorrhagic necrosis involving cerebral lobes and cerebellum. Post-mortem CNS samples were submitted to the CDC in Atlanta for further analysis. These were positive for Balamuthia mandrillaris by Real-Time PCR. This case illustrates the importance of detailed clinical history in identification of fatal causes of granulomatous encephalitis from those that are amenable to treatment. It also draws attention to identification of the amoeba by “standard morphology” since there are no special stains available to highlight the parasite in the tissue samples. California remains among areas endemic for the amoebic encephalitis. Surgical pathologists are to be aware of the possibility of such rare cases.

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68

Fatal Seronegative Rickettsia rickettsii Meningoencephalitis in an Infant

Hope Richard1, Christine Fuller1. 1Virginia Commonwealth University Health System

Rocky Mountain spotted fever is a tick-transmitted infection caused by Rickettsia rickettsii. Patients typically present with fever, headache, and rash; left untreated, complications include systemic microinfarcts secondary to microvasculitis, and (meningo)encephalitis. Rapid diagnosis and treatment are crucial to prevent permanent neurologic sequelae or death. Herein we present a 3 month old female infant who was up to date on her vaccinations and healthy until 2 days prior to admission when her mother noticed a fever for which she was treated with acetaminophen. The following day a targetoid rash developed on her palms and soles of the feet which subsequently spread to the chest, at which time she was admitted to the hospital and treated with antibiotics (ceftriaxone) with resolution of the rash. The fever remained but was managed with anti-pyretics, and blood cultures remained negative. On hospital day 4 the patient was noted to be pancytopenic and lumbar puncture showed elevated WBCs (12-18/mm3), lactate (6.5 mmol/L), and protein (207 mg/dL). Serology studies for numerous infectious agents including Rickettsia were negative. Antibiotics for viral and bacterial meningitis were started (Ceftriaxone, Vancomycin, and Doxycycline) and the patient was intubated. Her status continued to decline, and the family decided to withdraw care. At autopsy, the cerebrum appeared pale and edematous. The leptomeninges were grossly normal without exudate. On coronal sections, edema was noted but there were no discrete intraparenchymal lesions or hemorrhages. Microscopic examination revealed plump reactive-appearing vascular endothelium together with numerous microglial nodules present within the spinal cord, brain stem, cerebrum and cerebellum. Foci of chronic inflammation were noted throughout the meninges, and rare perivascular mononuclear aggregates and microinfarcts were seen. Petechia were absent. Routine bacterial, viral, and fungal stains were negative; however, immunohistochemical staining was positive for Rickettsial species, and confirmatory PCR studies yielded Rickettsia rickettsii as the etiologic agent.

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69

Amebic Meningoencephalitis with Acanthamoeba spp

Stewart Neill1, Christina Appin1, Daniel Brat1, Jeannette Guarner1. 1Department of Pathology and Laboratory Medicine, Emory University

Background: Intracranial amebic infections are rare events that are nearly universally fatal. They are caused by several species of free-living amoeba, with significant inter-species variation as to usual routes of infection, disease course, and inflammatory response. We present a case of Acanthamoeba CNS infection.

Case History: A 73 year-old male who was 4 months status post renal transplant for end-stage renal disease presented with fever, fatigue and gait instability and soon developed progressive neurologic decline. Head CT demonstrated a rim-enhancing hypodensity in the right frontal lobe. An infectious disease workup, including lumbar puncture, was non-diagnostic. Brain MRI revealed diffusion restriction and T2/FLAIR signal within the cortices and brainstem along with leptomeningeal enhancement. Brain biopsy demonstrated abundant necrosis, a mixed inflammatory infiltrate, and amoebic cysts and trophozoites. The patient expired 2 days following biopsy, 12 days after presentation. Review of cerebrospinal fluid (CSF) cytology revealed amoeba.

Autopsy Findings: A complete autopsy found no evidence of extracranial amebiasis. The brain showed leptomeningeal opacification with underlying parenchymal necrosis, particularly around the base of the brain and brainstem. Microscopic examination revealed amoebic trophozoite and cyst forms within the leptomeninges and extending perivascularly into the parenchyma. A moderate mixed granulomatous and necrotizing inflammatory infiltrate was present. Morphology and immunohistochemistry were indicative of Acanthamoeba spp.

Discussion: Acanthamoeba species usually trigger a focal granulomatous encephalitis secondary to hematogenous dissemination from an extracranial source. While meningeal involvement may occur, Acanthamoeba organisms tend to cluster around parenchymal vessels and rarely are found in CSF samples. Death typically occurs within a few weeks to a month. By contrast, Naegleria spp. infections usually seed the CSF, produce a primary meningitis, and quickly precipitate death. Our case shows that a primary meningeal infection with secondary spread to the cortex is part of the spectrum of disease of Acanthamoeba infections.

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70

Lack of Findings to Support a Role for HPV in a Cohort of Focal Cortical Dysplasia, Type IIB

Declan McGuone1, Kevin Shapiro1, Kevin Staley1, Anat Stemmer-Rachamimov1. 1Massachusetts General Hospital, Harvard University, Boston MA; 2UCSF School of Medicine, CA

Introduction: Focal cortical dysplasia (FCD) is increasingly recognized as an important cause of medically refractory epilepsy. Neuropathologic examination of surgical specimens in these patients reveals a combination of architectural and cytologic abnormalities. FCD type IIB (FCDIIB) is specifically defined by focal abnormal cortical architecture with dysmorphic neurons and balloon cells. Recently, two groups have independently reported high risk human papilloma virus (HPV) type 16 in balloon cells in a high proportion of human FCDIIB specimens, implying a potential mechanistic relationship between high risk HPV infection during development and focal cortical malformations (1-2). We sought to analyze an independent surgical cohort for the presence of HPV 16 DNA.

Methods: All Focal cortical dysplasia cases reported over a 19 year period were retrieved from the pathology files of the Massachusetts General Hospital. The cases were reviewed independently by two neuropathologists for histological confirmation and PCR for high risk HPV was performed on all cases meeting diagnostic criteria for FCDIIB. In situ hybridization for HPV was performed on a subset of these cases. All patient data were de-identified.

Results: Six patients with medically refractory epilepsy and biopsy proven FCDIIB (age range 4 -48 years) were identified. HPV PCR for high risk HPV failed to identify HPV nucleic acids in any of these cases. In situ hybridization studies performed on a subset of patients were also negative.

Conclusion: We have not been able to identify high risk HPV nucleic acids in any cases reviewed. These data imply mechanisms other than HPV infection are also likely to be important in the pathogenesis of a subset of FCDIIB cases.

References:

1: Chen J et al. Detection of human papillomavirus in human focal cortical dysplasia type IIB. Ann Neurol. 2012;72(6):881-92.

2: Liu S etal. Viral Infection and Focal Cortical Dysplasia. Ltr to the Editor Ann Neurol. 2013.

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71

A Tissue Microarray Approach to the Immunohistochemical Characterization of Pituitary Adenoma

William McDonald1, Nilanjana Banerji1, Joel Money3, Kelsey McDonald4. 1Allina Health; 2John Nasseff Neuroscience Institute; 3Hospital Pathology Associates; 4Inst for Med Informatics, Biometry and Epidemiology, Essen, Germany

Pituitary adenomas are common tumors of the pituitary gland, but institutions vary greatly in diagnostic strategy and no optimal diagnostic algorithm has been defined. Tissue microarrays (TMAs) are collections of tissue samples placed in an orderly array for the production of histological slides. While laborious to construct, they allow rapid, inexpensive comparison of multiple tumors under the same histological conditions. Our goal is to use a TMA containing pituitary macroadenomas to choose from immunohistochemical (IHC) stains of Pit-1, SF-1, anterior pituitary hormones, and alpha subunit of human chorionic gonadotropin to create a more efficient, cost-effective diagnostic algorithm. Thirty-eight pituitary macroadenomas and six control cases (representing various tumor types and normal pituitary) were selected for inclusion in the TMA. Correlation and cluster analyses were performed using the R statistical program, and show, for example, significant positive correlation between Pit-1 and prolactin, growth hormone, and TSH (Pearson’s r: 0.42-0.93) and significant negative correlation with SF-1 (Pearson’s r: -0.69), as expected. This suggests internal consistency and opportunities for a more efficient panel. A diagnostic algorithm using stepwise IHC staining to diagnose pituitary adenomas is discussed.

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72

CD1a Immunohistochemistry in Pituitary Lesions

David Pisapia1, Ehud Lavi1. 1Weill Cornell Medical College

Within the differential diagnosis of patients presenting with sellar or suprasellar lesions identified radiologically is Langerhans cell histiocytosis (LCH). CD1a staining is often used in the clinical setting to confirm the presence of an abnormal proliferation of Langerhans cells on histological sections and often contributes to the diagnosis of LCH. Here, we report that the MTB1 monoclonal antibody to CD1a reacts to adenohypophyseal epithelial elements and, particularly in inflammatory lesions, that this may introduce diagnostic uncertainty. We show that immunohistochemistry for CD1a shows positivity in a variety of settings including normal pituitary tissue resected at autopsy, surgically resected non-neoplastic pituitary tissue, and pituitary tissue that is found embedded within the context of inflammatory lesions and proliferative lesions other than LCH. Moreover, CD1a staining was found to be negative in null-cell pituitary adenomas suggesting cross-reactivity may be caused by the presence of hormonally derived antigens within adenohypophyseal cells. We propose that in the setting of pituitary lesions, a panel of stains should be used including CD1a, Langerin, BRAF-V600E, and synaptophysin in conjunction with morphological analysis before a diagnosis of Langerhans histiocytosis is rendered, and that CD1a positivity by itself should be interpreted with caution.

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73

Pituitary Adenomas: VUV (Very Unusual Variants)

Bette Kleinschmidt-DeMasters1, T Cummings2, M Lopes3. 1University of Colorado Anschutz Medical Campus; 2Duke University Medical Center; 3University of Virginia Health System

Background: Pituitary adenomas constitute more than 85% of sellar region masses, are usually easily diagnosed on routine hematoxylin and eosin, and show a homogeneous cell population. Interruption of this otherwise mundane appearance by excessive fibrosis, microcalcifications, amyloid deposition, architectural sinusoidal pattern, nuclear enlargement/pleomorphism, or interspersed cells with ganglionic metaplasia are well-known features of adenomas as well as many other endocrine tumors and do not engender surprise or misdiagnosis on the part of the pathologist. However, occasional pituitary adenomas show histological features that extend beyond these known variants and these are minimally illustrated in the literature.

Design: Review of the files of three neuropathologists practicing at centers with large numbers of sellar region masses to identify morphological variants they have encountered.

Results: Five cases of adenomas were identified that met study criteria: 3 gonadotroph adenomas with (one each) osseous metaplasia, CAM5.2 immunohistochemistry (IHC)+ fibrous bodies, and abundant eosinophilic intralysosomal proteinaceous deposits (confirmed by EM), one mixed prolactinoma-growth hormone-alpha subunit-IHC+ adenoma with overwhelming benign inflammatory infiltrates and lymphoid follicle formation, and one densely-granulated GH adenoma with numerous interspersed small nerve twigs (despite intraoperative correlation of no involvement of dura or cavernous sinus). The gonadotroph adenoma further showed extensive invasion on neuroimaging and loss of E-cadherin IHC.

Conclusion: Very unusual variants (VUVs) happen, even within otherwise very histologically-mundane pituitary adenomas. These can give pause during the routine examination of adenomas and thus are worth highlighting for neuropathologists.

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74

Stellate Amyloid Deposition in a Densely Granulated-Somatotroph Type Pituitary Adenoma

Nitin Agarwal1, Priyanka Singh2, Jean Eloy1, James Liu1, Ada Baisre1. 1Rutgers New Jersey Medical School; 2Wayne State University School of Medicine

The authors describe a unique case of stellate amyloid deposition in a densely granulated somatotroph type (growth-hormone producing) pituitary adenoma and review the literature on amyloid deposition in endocrine-active pituitary adenomas. A 54-year-old acromegalic man with hypertension and obstructive sleep apnea presented with a pituitary macroadenoma. Endocrinologic work-up revealed elevated serum growth hormone and insulin-like growth factor-1. The patient underwent complete removal of the tumor via an endoscopic endonasal transsphenoidal approach resulting in biochemical remission. Pathologic examination revealed a growth hormone producing pituitary adenoma with abundant stellate amyloid deposition. Amyloid deposition involving the pituitary gland usually occurs as part of a systemic disorder, but it can occasionally be seen within pituitary adenomas, either as nodular/spheroid or more rarely as stellate deposits. The presence of amyloid has been described more commonly in prolactinomas and somatotroph (growth hormone-producing) adenomas. Growth-hormone producing pituitary adenomas are commonly characterized by spheroid pattern of deposition; however, histological analysis of the case presented here reveals a growth hormone producing adenoma with abundant stellate amyloid deposits.

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75

Sellar Atypical Teratoid Rhabdoid Tumor (AT/RT)–A Rare Tumor of Middle Aged Women

Derick Aranda1, Mark Jentoft1, Joseph Parisi1. 1Mayo Clinic

Atypical Teratoid Rhabdoid Tumor (AT/RT) is a rare tumor that typically occurs in the posterior fossa of young children, usually under 2 years of age. There have been infrequent reports of CNS AT/RT outside the posterior fossa and in adults; occurrence in the sella region is rare with 7 previously reported cases, curiously all in woman. We describe 2 additional cases of sellar AT/RT in 2 middle age (36 and 47-year-old) women that clinically and radiologically mimicked pituitary adenoma. Histologically, the neoplasms demonstrated high cellularity and malignant features with foci of hemorrhage and necrosis; identification of a subset of large pleomorphic cells with abundant eosinophilic cytoplasm suggestive of rhabdoid cells prompted consideration of AT/RT that was confirmed by immunoprofile and loss of INI1 expression. Follow-up is available in only 4 of the 7 previously reported cases, with death in 3 (3-30 months) and survival in 1 (6 months). With this limited data, definite conclusions regarding prognosis are difficult. Our cases both showed response to chemoradiation at 4 and 11 month followup. Although rare, when confronted with a sellar lesion with unusual histological features, the pathologist should consider the possibility of AT/RT. The addition of these two cases to the already existing literature of sellar AT/RT will aid in the understanding of this rare and unusual entity.

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76

Salivary Gland Rests in Rathke Cleft Cysts. A Review of the Literature

Dibson Gondim1, Gregory Bosh1, Atul Agarwal1, Daniel Fulkerson1, Jose Bonnin1. 1Indiana University School of Medicine

Salivary gland rests were found in approximately 3.4 per cent of pituitary glands examined in a large autopsy series, and both benign and malignant salivary-like tumors involving the sellar region have been reported. A 16-year-old female developed visual loss in the left eye over a 3-day period. Her visual acuity was 20/200 on the left eye and normal on the right. An MRI scan revealed a 1.7x 1.5 x 1.3 cm bilobed peripherally enhancing cystic mass in the sella turcica. The lesion extended to the suprasellar cistern and there was some mass effect on the optic chiasm. A subsequent CT scan showed a large aerated sphenoid sinus and bright punctate densities in the sellar region. In the differential diagnosis, craniopharygioma and Rathke cleft cyst were considered. Because of the rapid progression of her visual loss, a transsphenoidal resection of the lesion was performed. The cyst contained a greenish mucinous fluid. Histopathological examination revealed fragments of adenohypophysis and a cyst lined by cuboidal, focally ciliated, epithelium. The cyst wall contained multiple islands of serous acinar glands compatible with salivary gland rests. Two years later she was found to have bilateral pheochromocytomas, which were resected. An identical twin also had bilateral pheochromocytomas, resected at 6 years of age and later was found to have an asymptomatic sellar cyst. Although Rathke cleft cysts involving the sellar region are not uncommon and despite the relatively frequent occurrence of salivary gland rests in the pituitary gland, such rests are rarely observed in the cyst walls. Unlike some cases of craniopharyngioma, Rathke cleft cysts have not been linked to any genetic abnormalities and are not known to be part of one of the familial tumor syndromes.

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77

A Rare Craniopharyngioma Recurred 21 years Later in Lumbosacral Dura, with Multiple Local and Cerebral Ectopic Recurrences

Osama Elkadi1, Jiang Qian2. 1Pathology and Lab Med, Albany Med Coll; 2Pathology, Albany Med Ctr Hosp/APS

Craniopharyngioma is a histologically benign tumor in the suprasellar region arising from the epithelial remnants of craniopharyngeal duct. It is classified into adamantinomatous and papillary types. Recurrence is not uncommon, and most occur in the original surgery site. Here we report a rare craniopharyngioma case in a 28 years-old male with local ectopic and distant/metastatic recurrences.

At age 4y, this patient presented with a suprasellar mass and underwent subfrontal resection. The mass was diagnosed as adamantinomatous cranipharyngioma (ACP). 3 years later suprasellar recurrence occurred, treated with resection and radiation. At age 10y, the patient presented with a right temporal lobe cystic mass, and at age 14y, a left frontal lobe mass, both diagnosed as ACP. At age 25y, the patient complained of increasing lower back pain, and MRI study revealed an intradural heterogeneously enhancing mass in the thecal sac at the L5-S1 level. Resection specimen demonstrated classic features of ACP: ribbons and nests of columnar and squamoid epithelial cells with peripheral palisading, in a loose reticulate stroma, myxoid cysts, and wet keratin composed of “ghost” cells and calcifications. 3 years after the last surgery, he was tumor-free at the sellar region and recurrence sites, but developed seizures, sleep disorders, and endocrinopathy.

Ectopic local recurrence of craniopharyngioma is uncommon, and distant/metastatic spread is even rarer with very few cases reported. Most of these recurrences are hypothesized to be due to seeding along the surgical pathway, or along the cerebrospinal fluid to a distant location. Since such recurrences can occur many years after the initial diagnosis, and no clinical or pathological parameters can reliably predict their occurrence, long-term follow-up with multidisciplinary approach is warranted. For many tumors, metastasis is a sign of malignancy, but its biological significance in craniopharyngioma remains to be elucidated and study of more such cases is needed.

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78

Pineoblastomas in Adults: Outcomes in a Series of Twelve Patients

Melissa Gener1, Aaron Cohen-Gadol1, Jamie Van Gompel2, Jeremy Cardinal1, Fredric Meyer2, Mohammad Ariai2, Mark Jentoft2, Jose Bonnin1. 1Indiana University School of Medicine; 2Mayo Clinic

Pineoblastomas are uncommon primitive neurectodermal tumors and most occur in children. They are exceedingly rare in adults. Few published case reports or small series analyzed the various aspects of these tumors in adult and pediatric patients. We report a series of 12 pineoblastomas in adults from 2 institutions over a period of 24 years. The clinical, radiologic, and pathologic features, and the clinical outcomes were studied and compared with previously reported cases in children and adults. Our patients ranged from 24-81 years of age and all but 1 presented with symptoms of obstructive hydrocephalus. Records of the radiological findings were available in all patients, but the actual neuroimaging studies were available only in 3. Ten patients had pineal masses on initial imaging and 11 had evidence of hydrocephalus. One patient had dissemination along the CSF pathways on follow-up imaging. Three cases had gross total resection, while subtotal resection was performed in 3, and diagnostic biopsies were obtained in 6 cases. Pathologically, the tumors had the classical morphological and immunohistochemical features of pineoblastomas. Postoperatively, ten patients received radiotherapy and chemotherapy was also given to five of them. When compared with previously reported cases, several differences were noted in clinical outcomes. Of our 12 patients, only 5 died of disease (average length of survival 118 months). Five patients are alive with no evidence of disease (average length of follow up 92 months). One patient died of unrelated causes and one was lost to follow up. The general consensus is that patients with subtotal resections or diagnostic biopsies have a much worse prognosis. Of our 9 patients with biopsy or subtotal resection, 5 are alive, and 3 died of disease. Although our series is small, the data suggests that pineoblastomas in adults have a less aggressive clinical course than in the pediatric population.

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79

Lack of BRAF-V600E Mutation in Papillary Tumor of the Pineal Region (PTPR)

Patrick Cimino1, Joseph Corbo1, Arie Perry2, Sonika Dahiya1. 1Washington University School of Medicine; 2University of California San Francisco

Background: Papillary Tumor of the Pineal Region (PTPR) is an extremely rare central nervous system (CNS) tumor with a variably aggressive clinical behavior, corresponding to WHO grade II-III. Very little is known about the genetic mutations comprising PTPR. Recent studies have shown that other papillary tumors harbor BRAF-V600E mutations, namely papillary thyroid carcinoma and papillary craniopharyngioma, the latter of which is a midline CNS papillary tumor like PTPR. For this study, we hypothesized that PTPR may contain the BRAF-V600E mutation and that it may be found at high frequencies like other midline papillary tumors.

Design: A search of our institutional files from 2004-2014 showed a total of 12 PTPR cases occurring in 10 patients. Chart review was performed to obtain demographics and pertinent clinical information. Of these 10 patients, 7 had additional material to be used for immunohistochemistry (IHC). IHC was performed using an anti-BRAF-V600E antibody (Spring Bioscience) utilizing a Benchmark Ultra instrument (Ventana Medical Systems Inc.).

Results: All 7 patients presented with symptoms of obstructive hydrocephalus. The patient age range was 1-60 years (average 29.9 years). There was a 3:4 male to female ratio. Initial magnetic resonance imaging characteristics tended to include partially cystic masses with heterogeneous post-contrast enhancement. The tumor size ranged from 1.1-4.4 cm (average 2.5 cm). IHC for BRAF-V600E demonstrated negativity in all 7/7 (100%) of cases.

Conclusions: In our cohort, none of the 7 PTPR cases harbored the BRAF-V600E mutation. This rate is unlike that of other midline papillary tumors, and suggests that these tumors despite their papillary phenotype may have a distinctive molecular background.

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80

Bifocal Papillary Tumor of the Pineal Region (PTPR) with Unusual Cytogenetic Features

Murat Gokden1, Bret Mobley2, Warren Sanger3, Hilary Nickols2. 1University of Arkansas for Medical Sciences; 2Vanderbilt University School of Medicine; 3University of Nebraska Medical Center

PTPR is an uncommon neoplasm codified in the 2007 World Health Organization classification as grade II-III due to variable biologic behavior. Cytogenetic and molecular diagnostic studies are rare, yielding no definitive genetic signature. Here, we report a case of PTPR with a bifocal presentation and unusual cytogenetic features.

A 25-year-old man presented with headache, disconjugate gaze and confusion. A mass in the pineal region and a smaller mass in the suprasellar region, with identical imaging characteristics, were identified. The former extended through the aqueduct partially into the 4th ventricle, was subtotally resected due to extensive adhesions to ependymal surfaces and showed typical PTPR histology. Cytogenetics revealed losses of chromosomes 3, 7, 10 and 14, additional losses of chromosomes Y and 18, and gains of chromosomes 3, 8 and 9. He is stable, receiving radiation therapy 6 months post-surgery.

This case has 2 unusual features: 1. Two separate mass lesions at presentation, 2. Losses of chromosomes 3, 7, 10, 14, 18 and Y, and gains of chromosomes 3, 8 and 9. Although leptomeningeal and craniospinal spread have been rarely reported, to our knowledge, only one previous report of synchronous pineal and suprasellar region masses exists, showing subsequent leptomeningeal dissemination. Our case also presented as bifocal lesions in these locations. It is yet to be seen whether it represents early phase of dissemination. Losses of chromosomes 10 and 22, and gains of chromosomes 4, 5, 8 and 11 have been reported more commonly. Gains of chromosomes 8 and 9 with loss of 10 in our case are compatible with prior reports, supporting a primary role of these changes in tumor etiology. We also identified unique losses of chromosomes 3, 7, 14, 18 and Y, adding to the available data and underscoring the genetic variability associated with the biology of these unusual neoplasms.

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81

The Utility of OCT4, CD117, and PLAP in Diagnosing Germinoma with Crush Artifact in the Central Nervous System

Zhe Piao1, Daniel Won2, Eric Stiner2, Todd Goldenberg2. 1Kaiser Permanente at Fontana, CA; 2Department of Neurosurgery, Kaiser Permanente Medical Center, Fontana

Background: Germinoma is the most common intracranial germ cell neoplasm. It is extremely challenging to make a diagnosis, particularly in a small specimen with severe crush artifact. The goal of this study was to evaluate the diagnostic utility of OCT4, CD117, and PLAP for germinoma with severe crush artifact.

Design: A total of 5 germinomas, 2 from the 3rd ventricle, 2 from suprasellar region, and 1 from pineal region, were included in this study. Immunostains of OCT4, CD117, and PLAP were performed on formalin-fixed, paraffin-embedded tissue. The positivity of crushed tumor area was compared with non-crushed tumor area in each slide.

Results: Four of 5 germinomas showed severe crush artifact. A few tumor cells showed positivity for OCT4, CD117, and PLAP in the background of severe crush artifact. The germinoma tumor cells without crush artifact showed strong positivity for OCT4, CD117, and PLAP. This result indicates that the positive cells in the crush artifact area are the germinoma tumor cells. The results of tumor positivity are summarized in the table 1. Among 3 markers, OCT4 and PLAP were better than CD117 in terms of positivity.

Conclusion: By using a panel of immunostains for OCT4, CD117, and PLAP, one can make a diagnosis of germinoma with severe crush artifact.

TABLE 1
TABLE 1
Image Tools
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82

Aggressive Juvenile Xanthogranuloma of Posterior Cranial Fossa and Temporal Bone: A Case Report

Nishant Tiwari1, Guy Marshall1, Frank Gannon1. 1Baylor College of Medicine, Houston, TX

Background: Juvenile xanthogranuloma (JXG) is a histiocytic disease of the non-Langerhans cell type. The intracranial JXGs have been known to present without cutaneous or systemic lesions. We present a case of a young adult (oldest reported) with an extra axial posterior fossa and temporal bone JXG.

Case Presentation: 20 year-old male presented to ENT service with a 6 month history of gradual onset of headache, vertigo and progressive hearing loss with pulsatile tinnitus in his left ear. Multidirectional spontaneous nystagmus was also noted. Pre-op MRI of the brain showed a large T2 isointense and T1 hyperintense mass in the posterior cranial fossa with direct erosion of temporal bone and obliteration of the left transverse-sigmoid sinus and jugular bulb.

Pathology: The lesion consisted mainly of benign histiocytes with minor sub-population of lipidized forms; occasional multinucleated giant cells and pleomorphic cells. Admixed with the histiocytes were abundant spindled fibroblasts and think ropy collagen, Larger thick collagen strands give an overall nodular appearance. Lesion also shows minor but significant areas with hemorrhage and granulation tissue with patchy areas of dystrophic calcification. The lesion also shows entrapment of degenerating bone, skeletal muscle and adipose tissue suggesting the infiltrative growth of the lesion. No cellular atypia or mitosis was present.

Follow up and progression: Surgical intervention was pursued and led to a significant resolution of symptoms with small residual disease. Tumor has since gradually grown over 10 month at all residual foci.

Conclusions: The intracranial JXGs in adults behave differently from the pediatric counterparts which are known to involute with time. Surgical resection without gross-total resection is likely to be inadequate. Corticosteroids, chemotherapy and radiation have been administered in such cases to achieve resolution.

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83

Non-Psammomatous Melanotic Schwannoma in Meckel’s Cave

Zachary Hoffer1, Gordana Juric-Sekhar1, Caitlin Latimer1, Jing Zhang1, Lia Halasz1, Manuel Ferreira1, C. Dirk Keene1, Luis Gonzalez-Cuyar1. 1University of Washington

Melanotic schwannomas are rare neuroectodermally-derived tumors that are typically associated with spinal roots, generally at the cervical and thoracic levels. However, rarely, melanocytic schwannomas present as skull base masses, and less than a handful of cases have been reported in association with the trigeminal nerve. Given the rare occurrence of trigeminal melanotic schwannomas as well as the different biological behavior and treatment modalities required, it is important for the neuropathologist to distinguish between melanocytic schwannoma and malignant melanoma, which are nearly identical based on histology and immunohistochemistry. Here, we report the case of a 60-year-old man who presented with one year of right-sided facial numbness and sharp shooting pain typical of trigeminal neuralgia. Magnetic resonance imaging revealed a 2.5 cm heterogeneously enhancing lesion in Meckel’s cave. The patient underwent surgical resection and intraoperatively the mass was dark brown to black. Histopathologically, the mass was a pigmented epithelioid neoplasm with low mitotic activity that lacked psammoma bodies and adipocyte differentiation. Immunohistochemically, it was positive for S100, Melan-A and HMB-45. In addition, immunohistochemical staining for collagen type IV revealed a delicate basement membrane surrounding numerous individual cells which favored a diagnosis of melanotic schwannoma over malignant melanoma. The patient underwent postoperative radiation therapy, and has been followed for two years with surveillance imaging without evidence of recurrence. This case is one of only a handful of cases of melanotic schwannoma involving the skull base, and illustrates the diagnostic challenges faced when distinguishing it from the more common malignant melanoma.

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84

Salivary Duct Carcinoma Presenting as a CP Angle Mass

Stewart Neill1, Kelly Magliocca1, Patricia Hudgins2, Matthew Schniederjan1. 1Department of Pathology and Laboratory Medicine, Emory University; 2Department of Radiology, Emory University

Background: The cerebellopontine angle (CPA) is a unique location within the CNS with a well-defined differential diagnosis for masses. Schwannomas and meningiomas are most common, but other primary and metastatic lesions have been reported, including some salivary gland neoplasms. We present the case of a patient with a salivary duct carcinoma (SDC) presenting as a CPA mass.

Case History: The patient was a 75-year-old male with progressive left facial pain and weakness. MRI had shown a left enhancing CPA and internal auditory canal mass that was interpreted as a schwannoma at an outside facility. Radiation therapy was pursued, but the tumor progressed and the patient presented to our hospital for biopsy and tumor debulking. Review of previous MRI studies revealed an ipsilateral parotidectomy defect with no mass in the parotid bed. CT scans of the chest, abdomen, and pelvis did not show a likely primary mass.

Pathologic Findings: Microscopic examination revealed an epithelioid neoplasm organized in variably sized nests with focal comedo-type necrosis, reminiscent of breast carcinoma. Neoplastic cells showed granular eosinophilic cytoplasm and atypical nuclei with prominent nucleoli. Immunohistochemistry revealed the tumor to be positive for CK7, CK5, androgen receptor, and Her2, and focally positive for GCDFP. The tumor was negative for CK20, TTF-1, napsin, PSA, prostein, p40, CDX-2, villin, synaptophysin, and chromogranin. A FISH assay for Her2/neu amplification was negative.

Discussion: SDC is an aggressive salivary gland neoplasm most often found in the parotid gland of older men. Whereas other salivary gland neoplasms have been reported as metastatic or possible primary tumors at the CPA, SDC has not been reported per our literature review. The pathologic findings in this case indicate SDC, and the radiologic findings are suggestive of prior parotidectomy. This entity may be added among the extended differential diagnosis of lesions found at the CPA.

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85

Radiation-Induced Atypical Meningiomas Exhibit Higher Recurrence Rates Than Sporadic Meningiomas of the Same Grade

Sarah Martin1, Eyas Hattab1. 1Department of Pathology, Indiana University School of Medicine

It is widely known that radiation-induced meningiomas overall tend to behave more aggressively, are more likely to recur, and have higher histologic grades than sporadic ones. However, studies comparing radiation-induced and sporadic meningiomas, grade for grade, are lacking. The aim of our study was to compare the clinical behavior of radiation-induced atypical meningiomas to that of sporadic atypical meningiomas and answer the question of whether the “history of prior irradiation” has any added value. Our institution’s archives were searched from 1986 to 2008 for meningiomas that qualified as atypical/grade II by the 2007 WHO grading criteria. One hundred twenty-five such patients were identified. The clinical medical records for these patients were reviewed. Eight (6%) patients were found to have had prior cranial radiation within the field of meningioma for various other CNS malignancies, including astrocytoma, germinoma and medulloblastoma. The median latency time from radiation to development of meningioma was 23 years, ranging between 14 and 25 years. Recurrence data for all 125 patients was obtained. Seven of the eight (87.5%) patients with radiation-induced meningiomas had tumor recurrence, while only 29.1% of non-radiation-induced meningiomas recurred. Pearson chi-square analysis demonstrated a statistically significant difference between the two groups (p=.001). Three of the patients with radiation-induced meningiomas had more than one recurrence. Our results demonstrate striking differences in clinical behavior within a cohort of grade II meningiomas. The presence of prior radiation puts the patient at higher risk for recurrence compared with other patients with meningiomas of the same grade. Such information is helpful in the risk stratification of such a heterogeneous group of tumors. Finally, while histologic criteria have long been the primary means of predicting tumor behavior, perhaps clinical criteria such as the presence of prior irradiation should also be taken into consideration in the grading of these tumors.

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86

WHO Grade I Meningiomas with Atypical Features: Correlation of Histopathology with Clinical Outcome

Declan McGuone1, Ariel Marciscano2, Andrzej Niemierko1, William Curry1, Fred Barker II1, Robert Martuza1, Kevin Oh1, Jay Loeffler1, Helen Shih1, Anat Stemmer-Rachamimov1. 1Massachusetts General Hospital, Harvard University, Boston MA; 2The Johns Hopkins Hospital, Baltimore, MD

Introduction: WHO grade I meningiomas with <2 atypical features, not fulfilling 2007 WHO criteria for grade II meningioma, may behave more aggressively than WHO grade I meningiomas without atypical features. We evaluated prognostic significance of atypical features in WHO grade I meningiomas.

Methods: All cases of WHO grade I meningioma with <2 atypical features (increased cellularity, sheeting, high N/C ratio, necrosis, prominent nucleoli) were reviewed and compared to cases without atypical features. Clinical data were reviewed. Cox regression and Kaplan-Meier (K-M) survival analysis was performed.

Results: 148 cases were retrieved (n =77 with <2 atypical features, n=71 with no atypical features). Median patient follow up was 37.5 months. 30 patients had progression/recurrence (P/R) after initial treatment and 22 of 30 (73%) of these had atypical features. The 1-year and 5-year P/R rates were 9.6% vs 1.4% and 30.8% vs 13.8 % for tumors with and without atypical features. Foci of necrosis (hazard ratio [HR], 10.5; 95% CI, 2.9 to 37.8), prominent nucleoli (HR, 6.3; 95% CI, 1.8 to 21.7) and sheeting (HR, 3.8; 95% CI, 1.2 to 11.6) were associated with increased risk of P/R. The median MIB-1 index was higher (p<0.001) in cases with atypical features (4.2%) versus those without (1.9%). The % of patients with MIB-1 proliferative rates > 3% at diagnosis was 65% for patients with atypical features and 17% for those without (p<0.001). Patients without grade I Simpson resection had an increased P/R risk (p<0.001) compared to those with grade I resection.

Conclusion: WHO Grade I meningiomas with atypical features have an increased risk of P/R compared to WHO grade I meningiomas without atypical features. Most cases correlate with a >3% MIB1 proliferation index. These findings support documenting atypical features in WHO grade I meningiomas as these patients may benefit from additional surgery and radiation.

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87

Rhabdoid-Like Meningioma: A Case Report

Caterina Giannini1, Jonathan Fratkin2, Josephine Wyatt-Ashmead2, Patrice Abell Aleff3. 1Mayo Clinic College of Medicine; 2University of Mississippi Medical Center, Jackson, MS; 3Mayo Clinic

Rhabdoid meningiomas (RM) are highly aggressive tumors, classified as WHO grade III. They are composed of loosely cohesive cells with abundant eosinophilic cytoplasm, eccentric nuclei, and hyaline paranuclear inclusions. By electron microscopy (EM), these consist of intermediate filament whorls often entrapping lysosomes and other organelles. Cytoplasmic inclusions frequently stain with vimentin and, at times, with cytokeratin or GFAP. In absence of EM, RM is frequently an H&E diagnosis. Rhabdoid cytology may be focal, present at first diagnosis or at recurrence, and often becomes increasingly prominent with subsequent resections suggesting that rhabdoid differentiation represents a morphologic marker of malignant transformation. Clinically aggressive RM typically demonstrates independent histological features of anaplasia. The behavior of RM with only focal rhabdoid features and/or without other malignant features remains indeterminate. A 57-year-old man presented with a generalized tonic-clonic seizure. MRI demonstrated a right temporoparietal well-circumscribed homogeneously enhancing mass 2.7×2.7×2.8 cm, associated with a peripherally enhancing cyst 1.8×1.7×1.2 cm and marked surrounding vasogenic edema. The tumor on H&E showed patternless growth and presence of cytoplasmic eosinophilic inclusions consistent with rhabdoid differentiation. Mitotic activity was low, and the tumor did not show macronucleoli, necrosis, or small cell transformation. There was no evidence of parenchymal brain invasion. To our surprise, under EM, the cytoplasmic inclusions did not correspond to the whorls of intermediate filaments typical of RM but rather to complex cytoplasmic invaginations with scattered intercellular junctions. Our findings prompt caution in making a diagnosis of RM (WHO grade III) simply based on optic microscopy appearance in absence of EM confirmation of rhabdoid differentiation in a tumor which does not show features of anaplasia. Additional cases are under study to evaluate the frequency of these findings since, if not an isolated phenomenon, this observation could highly impact diagnosis, classification, and grading of RM at large.

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88

Leptomeningeal Melanomatosis With Signet-Ring Cell Features Mimicking Leptomeningeal Carcinomatosis: A Postmortem Examination

Sarah Martin1, Andrew Fabiano2, Robert Fenstermaker2, Richard Cheney1, Jingxin Qiu1. 1Roswell Park Cancer Institute Department of Pathology; 2Roswell Park Cancer Institute Department of Neurosurgery

We present the case of a 63-year-old man clinically diagnosed with leptomeningeal carcinomatosis, who upon postmortem examination was found instead to have leptomeningeal melanomatosis with signet-ring cell features. This patient presented with no known history of cancer and six months history of anorexia, weight loss and fatigue, followed by two weeks of confusion and difficulty with speaking and ambulating. Brain MRI showed multiple variably sized contrast-enhancing lesions and diffuse abnormal leptomeningeal contrast enhancement. Full body CT imaging revealed no detectable lesions elsewhere in the body. Antemortem CSF cytology was diagnosed as metastatic adenocarcinoma on multiple occasions based upon the signet ring cell morphology. Immunohistochemical studies were not performed. The patient expired despite treatment with Temodar, whole brain radiation, and intrathecal methotrexate. Postmortem examination of the brain revealed no gross abnormality. Microscopic examination showed a subdural collection of discohesive tumor cells with marked nuclear pleomorphism, hyperchromasia, and frequent signet-ring or rhabdoid morphology, as well as a striking, diffuse infiltration of tumor cells in the leptomeninges. Intraparenchymal deposits were also seen in many areas. General autopsy also identified tumor metastases in the larynx and testes. Tumor cells were immunoreactive for melanoma markers (S100, Melan-A, Tyrosinase and HMB-45) and negative for cytokeratins and mucin. A diagnosis of leptomeningeal melanomatosis with signet-ring cell features was rendered. The patient had no known history of melanoma and no primary lesion was discovered at autopsy. While most patients with metastatic melanoma do have a known history, in a small percentage no primary site is identified. We discuss possible etiologies for this phenomenon and emphasize the potential pitfall of signet-ring melanoma clinically and cytologically mimicking adenocarcinoma. Melanoma with signet-ring cell features should be included in the differential diagnosis for cases of undiagnosed suspected adenocarcinoma.

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89

Primary Malignant Melanoma of the Leptomeninges with GNA11 (Q209L) Mutation: Case Report and Literature Review

Michael Lynch1, G. Timothy Reiter1, Joseph Drabick1, Charles Specht1. 1Penn State - Hershey Medical Center

We report the case of a 64-year-old female without significant past medical history who presented with abdominal pain, weight loss, new onset back and bilateral leg pain, and trouble walking. Magnetic resonance imaging (MRI) revealed a well-defined, 1.6 cm enhancing intradural extramedullary mass at T11. Numerous smaller enhancing leptomeningeal nodules surrounded the brain, cervical spinal cord, and cauda equina. Surgery was recommended for the lesion at T11 to decompress the spinal cord and obtain a tissue diagnosis. Intraoperative findings included widespread deposition of melanin within the exposed arachnoid membrane and tumor. Histologically, the resected tumor is malignant melanoma. Immunohistochemically, the tumor is positive for Melan-A (Mart-1), HMB-45, and microphthalmia transcription factor (MITF). Next generation sequencing of this tumor shows a GNA11 (Q209L) mutation. Dermatologic, gynecologic, and ophthalmologic evaluation revealed no primary tumor outside the central nervous system (CNS). Activating mutations in the Gq alpha subunit of GNAQ or GNA11 can lead to constitutively activated mitogen-activated protein kinase (MAPK) and protein kinase C (PKC) pathways; this may occur without BRAF or RAS mutations. Previous studies have shown that mutations in either GNAQ or GNA11 are often found in uveal melanoma. Primary CNS melanomas are rare, but the few reported analyses have demonstrated frequent mutations in GNAQ or GNA11. These mutations are not common in cutaneous melanoma. Understanding this molecular distinction is important for proper diagnosis and has potential future therapeutic implications. As an example, recent studies have shown that PKC inhibitors in combination with MEK inhibition lead to sustained MAPK pathway inhibition and tumor shrinkage in vivo. Evaluation for GNAQ and GNA11 mutation status can thus provide biological information that is of significant utility for treatment planning when malignant melanoma is encountered in the meninges. We aim to highlight the molecular profile and histologic differential of these tumors.

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90

Analysis of Select Angiogenic Markers in Melanoma Brain Metastases

Dimitri Trembath1, Stergios Moschos2, Anna Snavely3, Evan Bradler4, Nana Nikolaishvilli-Feinberg3, Bentley Midkiff4, Jonette Werley5, Michal Krauze6, Ronald Hamilton5. 1Department of Pathology and Lab Medicine, UNC-Chapel Hill; 2Department of Medicine, UNC-Chapel Hill; 3Lineberger Comprehensive Cancer Center, UNC-Chapel Hill, NC; 4The University of North Carolina School of Medicine, Chapel Hill, NC; 5Department of Pathology, UPMC, Pittsburgh, PA; 6Department of Medicine, UPMC, Pittsburgh, PA

Background: We have previously demonstrated that the presence of intra-tumoral hemorrhage and lack of immune infiltrates in craniotomy specimens of melanoma brain metastases (MBM) are associated with shorter overall survival (OS), defined as time from craniotomy to death. We hypothesize that immune checkpoint protein(s), angiogenic proteins, cytokines, hypoxia, and the density of mature vs. immature blood vessels (MBV, IBV) are important prognostic factors.

Methods: Craniotomy resections of melanoma brain metastases were reviewed for the presence/absence of tumor, normal brain, immune infiltrate, and gliosis. These specimens were then stained for angiogenic/hypoxic factors (bFGF, VEGF, HIF1α), blood vessel density (CD31), PD-L1, and for immature (CD31+SMA-) or mature blood vessels (CD31+SMA+). Each marker was analyzed in the different compartments described above using an Aperio imaging system and Definiens Tissue Studio.

Results: An average of 44 cases was analyzed for each of the 7 stains. Treating each variable as continuous using Cox proportional hazards, none were associated with OS. When survival information was used to define the optimal cut-point for each variable between cases with long versus short OS, high tumor expression of PD-L1 and bFGF, and low tumor expression of HIF1α were associated with worse OS (unadjusted p < 0.05; hazard ratios 1.92, 2.2, and 2.1 respectively). PD-L1 showed significantly higher expression in tumor compared to normal brain and there was a significant inverse association between PD-L1 expression in tumor and immune infiltrate.

Conclusions: Our results suggest that neither blood vessel density nor maturity within MBM is a significant prognostic factor. Tumor response to hypoxia by up-regulation of HIF1α may paradoxically be a favorable prognostic factor. More importantly, high expression of PD-L1 in MBM is an adverse prognostic factor, and may be related to the tumor’s immunosuppressive effects in the brain microenvironment, raising the possibility of a new therapeutic target.

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91

Epithelioid Schwannoma of a Spinal Nerve Root

Rachael Vaubel1, Howard Chang2, Karen Fritchie1, Mark Jentoft1. 1Mayo Clinic, Rochester, MN; 2Sparrow Health System, Lansing, MI

Even though the diagnosis of schwannoma is usually straightforward in lesions with classic features, tumors with predominantly epithelioid morphology or marked myxoid change may present a diagnostic challenge. We present a case of a slow-growing, dumbbell-shaped mass present within a thoracic neural foramen. Histologic examination revealed a proliferation of epithelioid cells in an abundant myxoid background raising the possibility of chordoma or chondrosarcoma. However, immunohistochemical studies showed the tumor cells to exhibit strong and diffuse reactivity for S100 and collagen IV while cytokeratin and brachyury were negative. This immunophenotype, in conjunction with ultrastructural studies showing long-spaced collagen and enveloping cytoplasmic processes, confirmed its schwann cell origin. The diagnosis of malignant peripheral nerve sheath tumor was considered but the lack of mitoses, necrosis and low Ki-67 labeling index supported the diagnosis of epithelioid schwannoma. This case illustrates the need for awareness of the epithelioid variant of schwannoma at this location to avoid misclassification as a malignant mesenchymal neoplasm.

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92

Intracranial Angiolipoma in a 23 year Old Male with Sturge-Weber Syndrome

Christopher Jones1, Aaron Wagner1, Gary Pearl1. 1Orlando Health

Angiolipomas of the central nervous system (CNS) are rare neoplasms. There are approximately 100 reported cases in the medical literature. They most commonly occur in the extradural space of the thoracic spine. Less commonly, they may arise in the cavernous sinus and the orbit. Rarely (∼10 cases) they have been reported in the parasellar space.

The intracranial manifestations of Sturge-Weber Syndrome are typically characterized by diffuse leptomeningeal vascular malformations, historically but incorrectly termed angiomata. They can be identified by contrast enhanced magnetic resonance imaging (MRI) as a diffuse leptomeningeal enhancement, most commonly of the superior convexities, that may fill the sulci and cause atrophy of the underlying parenchyma. Large intracranial neoplasms are not associated with the syndrome. To our knowledge, the development of an intracranial angiolipoma in a patient with Sturge-Weber Syndrome has not previously been reported.

We present a case of a large 11 cm angiolipoma in a 23 year old male with Sturge-Weber syndrome. The mass is seen on MRI as a solid and bilobed enhancing mass with a suprasellar epicenter. The tumor fills the bilateral anterior temporal extra-axial space connecting in the midline across the perimesencephalic cistern and prepontine cistern. The tumor fills the perimesencephalic cistern and completely encases the Circle of Willis, optic chiasm and pituitary gland, compressing the brainstem.

Stereotactic core needle biopsy was performed which revealed mature fat and blood vessels. Immunohistochemistry confirmed the presence of CD31-positive vasculature and S100-positive adipocytes. The tissue did not demonstrate staining with anti-HMB45, mitigating against angiomyolipoma.

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93

Intracranial Osteosarcoma Arising in Fibrous Dysplasia

Christine James1, Darnell Josiah1, Patrick Bacaj1, Charles Rosen1, Kymberly Gyure1. 1West Virginia University

Fibrous dysplasia is a generally benign fibro-osseous proliferation occurring in bone. The skull and facial bones are commonly affected, particularly in the polyostotic form of fibrous dysplasia. Malignant transformation is extremely rare (<1% of cases) and is often associated with prior radiation therapy. We report an unusual case of spontaneous osteosarcoma arising in a patient with a prior history of pathologically confirmed fibrous dysplasia. A 42-year-old man presented to the emergency department with a five-day history of progressive vision loss in his left eye. He had a history of multiple previous surgeries for craniofacial fibrous dysplasia. Imaging studies revealed extensive involvement of the craniofacial bones with fibrous dysplasia, initially thought to be unchanged when compared to his prior imaging studies. He subsequently underwent a left frontotemporal craniotomy, posterior orbitotomy, and clinoidectomy to decompress the optic nerve. A soft tissue mass was discovered growing medial to the optic nerve and involving the medial orbit. The mass was debulked, and sections revealed a high-grade sarcoma with focal osteoid formation. Although rare, malignant transformation should be considered in the differential diagnosis of patients with a history of fibrous dysplasia, particularly in those with rapid progression of their symptoms.

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94

Epstein-Barr Virus-Positive Primary CNS Lymphomas Associated with Intracranial Mass Lesions in Immunocompetent patients

Yasuo Sugita1, Koichi Ohshima1, Jun Masuoka2, Yoshizo Kimura3, Koichi Higaki3, Susumu Nakashima4. 1Department of Pathology, Kurume University School of Medicine; 2Department of Neurosurgery, Saga University Faculty of Medicine; 3Department of Pathology, St Mary’s Hospital; 4Department of Neurosurgery, St. Mary’s Hospital

The Epstein-Barr virus (EBV) has been implicated in the pathogenesis of some primary CNS lymphmas (PCNSLs) in immunocompetent patients. To investigate the role of EBV in pathogenesis for PCNSLs associated with intracranial mass lesions, the present study assessed three unusual cases. Case 1 was a 66-year-old man with a history of gait disturbance. Neuroimagings (NIs) revealed a solid and a cystic lesion in the posterior fossa. Histologically, the solid tumor showed a proliferation of pleomorphic immunoblastic-like cells. The cystic tumor showed keratinous debris, which was consistent with an epidermoid cyst (EC). Case 2 was a 57-year-old woman with gait and hearing disturbances. NIs revealed a cystic mass in the cerebellopontine angle. Histologically, the tumor showed keratinous debris, which was consistent with an EC. In addition, atypical lymphocytes scattered adjacent an EC without forming a mass. Case 3 was a 77-year-old man with a history of head injury. NIs revealed a right frontal-temporal subdural lesion. Histological examinations showed atypical lymphocytes conglomerated in the thick outer membrane of the organized chronic subdural hematoma (CSH). Immunohistochemically, atypical lymphocytes of all these cases were positive for CD20, and negative for CD3. On in situ hybridization, atypical lymphocytes in all cases showed positive signals for EBV-encoded small RNAs. Based on these findings, the proliferations of atypical lymphocytes in all cases were diagnosed as EBV-positive PCNSLs. It is well known that ECs and CSHs cause severe inflammatory reactions in the surrounding tissues. Therefore, these results indicated that long-standing chronic inflammation caused by intracranial masses and EBV infections have contributed to the pathogenesis of PCNSLs. In addition, Cases 2 and 3 may be an earlier-stage of lymphoma when compared with Case 1. This underscores the need for careful attention to the potential presence of atypical lymphocytes during the pathologist’s review of the intracranial mass lesions.

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95

An Unusual Case of an EBV-positive CNS Lymphoma in an Immunocompetent Adult

Brian Bockelman1, Cristina Vincentelli1, Vathany Sriganeshan1, Amilcar Castellano-Sanchez2. 1A.M. Rywlin M.D. Department of Pathology Mount Sinai Medical Center; 2Herbert Wertheim College of Medicine Florida International University

CNS diffuse large B-cell lymphomas comprise approximately 2-3 percent of all brain tumors and less than 1 percent of all non-Hodgkin lymphomas. These rare neoplasms are usually not associated with Epstein-Barr virus in immunocompetent patients. We present a case of a 59 year old woman with a two week history of progressive left upper and lower extremity weakness while vacationing with her husband in the United States. An MRI revealed scattered ring-enhancing lesions in the right inferior frontal gyrus, right thalamus and lentiform nucleus extending through the corona radiata to the centrum semiovale, and left hypothalamus. There was effacement of the right lateral ventricle, third ventricle and ambient cistern with a 5 mm leftward midline shift. Additionally observed was extensive vasogenic edema around the lesions. The patient was taken to surgery and a biopsy was performed. Microscopic examination revealed brain tissue with reactive gliosis and areas with a dense angiocentric infiltrate with associated necrosis. The infiltrate was composed of a pleomorphic population of cells including small lymphocytes, histiocytes, medium to larger atypical cells with irregular nuclear contours, as well as scattered large cells with vesicular nuclei. The large atypical cells were positive for CD45, CD20 (weak), CD79a, PAX5, MUM1, and CD30. Additionally, EBER in situ hybridization was positive. The diagnosis of EBV-positive CNS diffuse large B-cell lymphoma was made. The fact that the patient is not immunocompromised along with histological findings of relatively scattered neoplastic cells in an abundant background of T lymphocytes and focal angioinvasion, which are not the characteristic features of this entity, made this a challenging case. The clinical presentation, imaging, and microscopic findings along with the differential diagnosis are discussed.

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96

Lymphomas in Intraoperative Consultations (IOC) in Neuropathology

Murat Gokden1, Melody Harrison1. 1University of Arkansas for Medical Sciences

Many times, IOC results in a change in surgical procedure and/or handling of the tissue. We have encountered several such instances over the years, especially with the IOC diagnosis of unsuspected lymphoma. Based on this observation, we reviewed our lymphoma diagnoses in IOC.

Neuropathology files were searched for cases with a final diagnosis of lymphoma in 2000-2014. Clinical findings, history, radiology reports and operative reports of these cases were reviewed. Cases with no mention of lymphoma as a possibility until neuropathology was involved were identified.

A total of 71 cases with a final diagnosis of lymphoma were identified. Nineteen of these had no mention of lymphoma in their diagnostic work-up before neuropathology was involved. IOC was performed in 15, with a diagnosis such as “atypical lymphoid infiltrate” or “favor lymphoma” that initiated additional work-up. In these 19 patients (age range: 22-78 years; mean age: 61 years; male:female=1:4), most common radiologic diagnoses were metastatic carcinoma, glioma, meningioma, or a combination of these. Most common location was cerebral hemispheres. Unusual locations were trigeminal nerve, cerebello-pontine angle, dura-based with infiltration into bone mimicking meningioma, and diffuse meningeal, mimicking meningioma en plaque. The final diagnosis in 17 cases was diffuse large B-cell lymphoma. One marginal zone lymphoma presented mimicking meningioma en plaque and one follicular lymphoma presented mimicking a meningioma.

These results emphasize the frequency of unexpected presentations in IOC and the importance of approaching cases with an open mind. The absence of a consistent presentation pattern may add to the potential problem. Even though other unexpected diagnoses are made in IOC, lymphomas can be particularly surprising to pathologists, radiologists and clinicians alike, emphasizing the direct immediate impact and the need for consistet use of IOC.

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97

Divergent Glioneuronal Differentiation in Metastatic Intracranial Neuroendocrine Carcinomas

Boleslaw Lach1, Suhita Joshi2, Naresh Murty3, Nasim Huq4. 1McMaster University, Hamilton Health Sciences; 2Greater Niagara General Hospital, Niagara Health System; 3Department of Neurosurgery, McMaster University; 4Clinical Surgery, McMaster University

Metastatic intracranial neuroendocrine carcinomas usually display a phenotype of primary tumours. We present three cases of metastatic carcinomas with different patterns as well as variable degree of divergent glioneuronal differentiation in intracranial lesions.

Patient #1: 77 yo. male with a history of multiple resections of squamous cell carcinomas and a Merkle cell carcinoma(MCC) of the head, was admitted for removal of a large "meningioma". The tumour displayed remarkable degree of pleomorphism, reminiscent of GBM, and immunoreactivity for synaptophysin(SYN),CD56,tubulin(TBL)and Tau. It was negative for GFAP, neurofilaments (NF),keratins(CK) and chromogranin (CHG). Review of the skin MCC revealed scattered NF+ ganglionic cells and immunoreactivity of small cells for many CK and neuronal markers (SYN,CHG,NF,CD56,Tau) as well as a small area of GFAP+ cells. Lymph node metastasis was SYN+,CK+ and CHG+, with no ganglionic or glial cells.

Patient #2: 71 yo. female with intracerebral occipital tumour. The resection specimen was a highly pleomorphic tumour displaying wide-spread reactivity for CK (AE1/3), SYN, NF, CHRG, GFAP, Tau, p53,and TTF-1. The tumour was negative for EMA and subgroup of CK(7,19,20). Carcinoma of lung was discovered post-operatively.

Patient #3: 77 yo. male with known small cell carcinoma(SCC) of lung. Resection specimen showed scattered neoplastic cells positive for GFAP. Otherwise immunohistochemistry revealed rectivity for SYN, CHG, Tau and CD56 as expected from SCC of lung.

These cases demonstrate the ability of poorly differentiated extracranial neuroendocrine tumours to differentiate to glial and ganglionic cells, occasionally mimicking primary tumours of the central nervous system.

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98

Experimental Photothrombotic White Matter Infarct with Marked Motor Deficit: A New Animal Model

Min-Cheol Lee1, Hyoung-Ihl Kim2, Kyung-Wha Lee1, Young Kim1, Hyung-Sun Kim2. 1Department of Pathology, Chonnam National University Medical School; 2Gwangju Institute of Science and Technology

We describe a new method for inducing a circumscribed subcortical capsular infarct (SCI), which results in persistent motor impairment in rats as a model to study post-stroke recovery. Photothrombotic destruction of the posterior limb of the internal capsule (PLIC) was performed using a low energy green laser and Rose Bengal in Sprague Dawley rats (male; n=38; 250-300g). Motor performance of all animals was assessed using forelimb placing, forelimb use asymmetry, and single pellet reaching test scores. Based on the degree of motor recovery, rats were dichotomized into either the poor recovery group (PRG) or the moderate recovery group (MRG). Imaging assessment of the impact of subcortical capsular infarct on brain metabolism was done using 18FDG microPET. Photothrombotic lesioning with low light energy selectively disrupted circumscribed capsular fibers. MRG demonstrated recovery of motor performance after one week but PRG showed a persistent motor impairment for more than 3 weeks. Damage to PLIC more effectively produced a severe motor deficit. Histological examination revealed that PRG had complete, full thickness destruction of the axon bundles in the anterior part of the PLIC. MRG show a partial lesion in the same area. Analysis of PET data revealed decreased regional glucose metabolism in the ipsilesional motor and bilateral sensory cortex and increased metabolism in the contralesional motor cortex and bilateral hippocampus during the early recovery period after SCI. These behavioral, histological and functional imaging findings support the usefulness of this novel SCI rat model for investigating motor recovery and the mechanism underlying SCI.

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99

Cerebral Biopsies in Posterior “Irreversible” Encephalopathy Syndrome (PIES)

Hannes Vogel1, Jennifer Ziskin1, Gregory Moes2. 1Stanford University; 2Kaiser Permanente

Posterior reversible encephalopathy syndrome (PRES) occurs when the posterior cerebral circulation fails to autoregulate in response to acute changes in blood pressure. Hyperperfusion causes disruption of the blood brain barrier and vasogenic edema, most commonly in parieto-occipital regions but without infarction. Pathological studies have reported fibrinoid vascular necrosis. Clinicians may neglect the possibility of PRES in cases in which hypertension is overshadowed by other considerations or atypical radiographic findings. We present three cases of acute parieto-occipital infarction which were biopsied for clinical considerations other than hypertension. Case 1 is a 49 yo male who presented with acute neurological deficits and a blood pressure of 190/112. MRI demonstrated a rim enhancing lesion in the occipital lobe, cerebellum and punctate lesions in deep gray and white matter and frontal lobes. Because of poor dentition, a cerebral abscess was suspected in the occipital lobe lesion. The biopsy showed recent infarction without infection. Case 2 was a 44 yo female with headache, left-sided weakness, and a generalized tonic-clonic seizure. Blood pressure was elevated to 205/121. MRI upon admission demonstrated enhancing parieto-occipital brain lesions with surrounding vasogenic edema. Concern for infection led to a stereotactic biopsy of the larger right parieto-occipital lobe lesion which demonstrated laminar necrosis without infection. Case 3 was a 20 year old female with recently documented hypertension, who developed sudden deficits and unresponsiveness, A CT showed left basal ganglion hemorrhage. Blood pressures to 220/173 were documented. She developed new lesions of both parietal lobes which were biopsied for possible vasculitis, which showed recent infarction without vasculitis. In all cases, the final clinical diagnosis was PRES with superimposed infarctions, thus we revise the term to reflect an irreversible condition. Neuropathologists should seek a history of severe hypertension when biopsies of contrast-enhancing or tumefactive lesions of parieto-occipital regions only show recent infarction.

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100

Histopathological Features of Intracranial Vascular Involvement in Neurofibromatosis Type I, Ehlers-Danlos Type IV, and FMD

Bette Kleinschmidt-DeMasters1, Seth Lummus1. 1University of Colorado Anschutz Medical Campus

Background: Recent studies suggest shared epidemiologic characteristics, common pathological weakening of arterial walls, and clinical outcomes amongst the nonatherosclerotic cerebrovascular arteriopathies. Few studies directly compare histological features of intracranial meningeal and parenchymal vasculature in major arteriopathies that affect CNS, i.e., fibromuscular dysplasia (FMD), Ehlers-Danlos type IV (ED-IV, vascular type), and neurofibromatosis type I (NF1).

Methods: Three adult autopsy cases of FMD in patients succumbing from basilar artery aneurysm rupture, vertebral artery dissection, and Moyamoya disease, all known patterns of FMD central nervous system (CNS) involvement, were studied with focus on intracranial rather than extracranial vascular changes. FMD was only confidently identified in autopsy specimens when multiple sections (optimally the entire Circle of Willis) were subjected to microscopic examination. A fourth FMD case of a massive, subcutaneous scalp cirsoid aneurysm showed classic "string-of-beads" gross appearance and microscopic medial hypertrophy. Direct comparison of FMD intracranial vascular changes was made with those in 1 autopsy patient with ED-IV and 4 autopsy patients with NF1.

Results: All three disorders involved cerebral vessels, albeit each with distinctive histological features and distribution, despite shared clinical and epidemiological characteristics. Disordered collagen within the muscularis was confined to FMD, best seen on picrosirius red histochemistry. Ehlers-Danlos type IV showed aneurysmal formation and eccentric intimal thickening of Circle of Willis vasculature but no obvious abnormalities of muscularis. Extensive leptomeningeal smaller-caliber arterial disease was confined to NF1 and particularly affected spinal cord. Fibrocellular smooth muscle intimal proliferation within parenchymal cerebral arteries was confirmed on smooth muscle actin immunohistochemistry in Moyamoya.

Conclusions: Cranial/intracranial artery involvement is not rare in these conditions, but requires extensive sampling to fully appreciate features.

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PLATFORM SESSION 5: TUMORS 2 - MOLECULAR PHENOTYPES Saturday 8-10 AM, June 14, 2014 Fashion Ballroom

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101

Chromosome Band 7q34 Deletions Resulting in KIAA1549-BRAF and FAM131B-BRAF Fusions in Pediatric Low Grade Gliomas

Jacquelyn Roth1, Mariarita Santi1, Avrum Pollock1, Brian Harding1, Lucy Rorke-Adams1, Laura Tooke1, Jaclyn Biegel1. 1Children’s Hopsital of Philadelphia

The majority of low grade gliomas (LGGs) in children are characterized by constitutive activation of the MAPK pathway through a variety of mechanisms, including mutations in BRAF and KRAS, inactivation of NF1, and KIAA1549-BRAF, FAM131B-BRAF and SRGAP3-RAF1 gene fusions. The KIAA1549-BRAF fusion is typically the result of a 2.0 Mb tandem duplication in 7q34 and is most common in pilocytic astrocytoma (PA). In the present study, single nucleotide polymorphism (SNP)-based array analysis of three LGGs demonstrated deletions in 7q34 that resulted in a BRAF fusion. One low grade neuronal-glial tumor had three deletions in 7q33 and 7q34. The most proximal deletion encompassed the 3` end of KIAA1549 while the most distal deletion included exons 3-7 of BRAF. RT-PCR analysis demonstrated a fusion between exon 15 of KIAA1549 and exon 9 of BRAF. SNP array analysis of a dysembryoplastic neuroepithelial tumor (DNT)/ganglioglioma (GG) revealed a 2.6 Mb deletion which included the 5` end of BRAF and extended to the 3` end of FAM131B. A PA displayed two deletions, which were 15 kb and 450 kb in size, with the most proximal deletion encompassing exons 4-7 of BRAF. Subsequent SNP array analysis of a recurrent PA from this patient yielded two additional deletions in 7q34, the most distal of which included the 3` end of FAM131B. RT-PCR analyses confirmed a fusion between exon 2 of FAM131B and exon 9 of BRAF in both the DNT/GG and PA. The presence of fusion transcripts in these three LGGs highlights the utility of SNP array analysis to identify deletions that are suggestive of fusion proteins, and confirms that such fusions are not restricted to PA. BRAF fusions can result from multiple non-overlapping deletions suggesting variant complex mechanisms of formation.

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102

A Clinicopathologic Study of Hypothalamic Pediatric Low-Grade Gliomas with BRAF V600E Mutation

Cheng-Ying Ho1, Gary Mason2, Ashley Campbell1, Mahtab Tehrani3, Brent Orr4, Roger Packer2, Fausto Rodriguez5. 1Divison of Pathology, Children’s National Medical Center; 2Department of Neurology, Children’s National Medical Center; 3Department of Pathology, Inova Fairfax Hospital; 4Department of Pathology, St. Jude Children’s Research Hospital; 5Department of Pathology, Johns Hopkins Hospital

Background: BRAF V600E mutation is frequently associated with pleomorphic xanthoastrocytomas (PXAs) and gangliogliomas (GGs). It is also detected in ∼5% of other pediatric low-grade gliomas (PLGAs), including pilocytic astrocytomas (PAs), pilomyxoid astrocytomas (PMAs) and diffuse astrocytomas (DAs). The aim of this study was to evaluate the clinicopathologic features of non-PXA/GG PLGAs that harbor the BRAF V600E mutation.

Design: After excluding PXAs and GGs, we performed a retrospective analysis of 18 BRAF V600E PLGAs from five institutions. Approximately half of the BRAF V600E tumors were located in the hypothalamus/optic chiasm (9/18, 50%). To investigate the association between the mutation and clinicopathologic features of tumors at this anatomic site, 9 V600E and 8 non-V600E (fusion or wild-type) hypothalamic PLGAs were evaluated. Thirteen cases had available clinical information and follow-up for over a year. 14 (8 V600E and 6 non-V600E) specimens were available for microscopic review.

Results: The BRAF V600E hypothalamic PLGAs appeared to affect infants (under one year of age) predominantly (56%, 5/9). Our analysis revealed similar histologic features in all hypothalamic BRAFV600E tumors (100%, 9/9): a monophasic dense compact architecture without microcystic or pilomyxoid patterns. Rosenthal fibers and eosinophilic granular bodies were rare to absent. Because the classic features of PAs were absent, nearly all hypothalamic BRAF V600E tumors were initially classified as low-grade astrocytomas. All 13 cases in the study received a subtotal resection or biopsy of the tumor. Ten of 13 patients had post-operative chemotherapy. Tumor progression or recurrence was observed in 86% (6/7) of BRAFV600E PLGAs, 25% (1/4) non-V600E PAs and 100% (2/2) non-V600E PMAs.

Conclusions: BRAF V600E mutation is commonly present in the hypothalamic region of infants. Hypothalamic BRAF V600E PLGAs appear to be associated with a unique histologic pattern, and have a tendency for a more aggressive clinical course.

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103

Identification of Molecular and Pathologic Subsets of Low Grade Gliomas and Glioneuronal Tumors by microRNA profiling

Heather Ames1, María Vizcaíno-Villalobos2, Fausto Rodriguez1. 1Johns Hopkins Medical School, Department of Pathology; 2Universidad Nacional Autónoma de México, Department of Cell Biology

Low-grade (WHO I-II) gliomas are tumors of the central nervous system that often have good prognosis following total resection, however they can create many neurological complications due to mass effect, and may be difficult to resect depending on anatomic location. They may also contain neuronal components to variable extent. With these characteristics, low-grade gliomas and glioneuronal tumors represent a valuable pool from which to identify potential molecular targets for early stages of tumorigenesis in the brain. MicroRNAs have been identified as mechanisms of epigenetic gene regulation in cells – pathways that are inherently targetable and amplifiable due to their simple nucleic acid structure. Given the importance of microRNA expression to tumor development, NanoString technology was applied to formalin-fixed paraffin-embedded tissues (n=45) to quantify the global expression of over 800 microRNAs in nine types of low-grade neoplasms of the brain: subependymal giant cell astrocytoma (SEGA) (n=6), pilocytic astrocytoma (n=6), pleomorphic xanthoastrocytomas (n=7), gangliogliomas (n=6), dysembryoplastic neuroepithelial tumor (n=5), angiocentric glioma (n=3), pediatric oligodendroglioma (n=3), pediatric diffuse astrocytoma (n=3), rosette forming glioneuronal tumor (n=2) and non-neoplastic brain controls (n=4). Raw data was normalized using nSolver, filtered for minimum expression threshold, and log2 transformed prior to analysis. MeV software was used to generate hierarchical clusters following significance evaluation via significant analysis of microarrays (SAMs). Hierarchical clustering separated tumors from non-neoplastic brain. These differentially expressed microRNAs included many that have been previously identified as altered in tumors of neural origin and important in neural differentiation. When looking at individual tumors, SEGAs clustered sharply together, consistent with a unique microRNA expression signature in this tumor subtype in particular, compared to other low grade glial and glioneuronal tumors. In summary, low-grade gliomas show alterations in microRNA expression patterns that may provide insights into their biology and/or make these tumors molecularly targetable.

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104

Cytogenetic Progression of Oligodendroglioma Assessed Using Single Nucleotide Polymorphism Array

David Nauen1, Andrew Guajardo1, Fayez Jabboure1, Peter Burger1, Christopher Gocke1. 1Johns Hopkins

The prognosis of oligodendroglioma (ODG), a relatively common infiltrating glioma, differs markedly between the typical tumor (WHO Grade II) and its anaplastic counterpart (WHO grade III). ODG is characterized by alterations, generally loss, of chromosome arms 1p and 19q. While this hallmark abnormality has been a focal point of investigation, additional cytogenetic changes that may underlie the progression of the tumor to higher grade have been less studied. Single nucleotide polymorphism (SNP) array is widely used in diagnosis of glial tumors. Although routine clinical reporting is limited in this context to chromosomes 1 and 19, SNP array data provides information on the status of all chromosomes. To assess mechanisms of pathogenesis and progression of ODG we review SNP array data for all such tumors assayed at our institution since 2011, when microarray analysis entered routine use. We confirm that chromosomal abnormalities in grade II ODG are not limited to 1p and 19q. A number of other abnormalities are noted, with loss of heterozygosity (LOH) in chromosomes 4 and 9 frequent. We find a marked increase in the average number of chromosomal abnormalities for grade III v grade II ODG (average 3.5 v 0.9), and that additional abnormalities recur in anaplastic ODG at high frequency. A number of chromosomes show LOH to different extent among different chromosomes, which may reflect distinct subpopulations during tumor progression. These findings may have implications for the increased mortality associated with grade III oligodendroglioma and may ultimately inform the search for more effective therapies.

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105

Next Generation Sequencing and Immunohistochemistry for EGFR, p53 and Mutant IDH1 in High-Grade Gliomas

Lyndsey Emery1, William Motley2, Donald O’Rourke1, Steven Brem1, Robert Daber1, Maria Martinez-Lage1. 1Hospital of the University of Pennsylvania; 2Perlman School of Medicine (University of Pennsylvania)

Background: Key genomic alterations in high-grade gliomas include TP53, IDH1, and EGFR. We report the correlation of next generation sequencing (NGS) in detecting point mutations and amplifications compared with routine immunohistochemistry.

Methods: Eighty-three high-grade gliomas were analyzed over 12 months. Immunohistochemical staining for mutated IDH1 (R132H), p53 and EGFR was performed and rated by qualitative or semi-quantitative scoring (0-3+). DNA from formalin-fixed paraffin embedded tissue was evaluated using a 47-gene panel via Illumina Mi-Seq.

Results: 81 tumors had perfect correlation between IDH1 R132H-specific staining and mutational status by sequencing (71 negative, 10 positive). One case did not stain with R132H-specific antibody, but had an R132L mutation by sequencing. 31 samples had TP53 mutations by NGS, of which 25 had moderate-strong p53 staining, 5 had a subset of p53-positive cells and one sample showed no staining. Among the 50 samples without TP53 mutations, 12 had moderate-strong p53 staining, 34 had a subset of p53-positive cells and 4 samples were negative. 31 samples had disease-associated EGFR mutations and/or amplification and 2+ or 3+ membranous EGFR staining. Two cases with EGFRvIII variant mutations had 1+ staining. No cases with mutations/amplification of EGFR showed negative EGFR staining. Among 33 samples with no NGS-detected EGFR mutations/amplifications, staining was variable, ranging from 1+ to 3+. Interestingly, of the 23 samples with 2+ or 3+ EGFR staining within this subset, 1 had a mutational “variant of unknown significance” in EGFR; 9 had “variants of unknown significance” within other genes, 6 of which had mutations/amplifications in PDGFRA.

Conclusions: NGS offers powerful complementary information in high-grade gliomas, which is specific and, potentially, more sensitive than classic immunohistochemistry. Furthermore, NGS could identify novel mutation-based subgroups among tumors with or without p53/EGFR staining. Application of these findings to outcome data could provide additional prognostic information and identify new therapeutic targets.

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106

Evaluation of Targeted Next Generation Sequencing (NGS) for Assessment of Variations in Gliomas

Ada Baisre1, Nitin Agarwal1, Helen Fernandes2, Anjali Seth3. 1RUTGERS-New Jersey Medical School; 2Weill Cornell Medical College-New York-Presbyterian Hospital; 3University Hospital

Introduction: Molecular markers including 1p/19q co-deletion (LOH), MGMT promoter hypermethylation, 10q (PTEN) loss and mutations in IDH 1&2 are currently used for diagnostic, prognostic and therapeutic purposes in a variety of CNS gliomas. More recently, next generation sequencing (NGS) is being used and numerous genomic abnormalities are being identified in several types of cancers. In this study we have stratified glioma specimens using these molecular markers. In addition, we used targeted NGS to detect differences between GBMs and other gliomas.

Materials and Methods: Formalin fixed paraffin embedded tissue (FFPE) from patients (n= 82) diagnosed with gliomas have been included in this study. Oligodendrogliomas, oligoastrocytomas, astrocytomas (non-GBM tumor group) and glioblastomas (GBM) were assessed for allelic status of chromosomes 1p, 19q and 10q by capillary electrophoresis. IDH1 and IDH2 mutation was also assessed using real time PCR and melt curve analysis. MGMT promoter methylation was assessed by pyrosequencing. A subset of these glioma specimens (n=49) were subjected to NGS using the Ion Torrent Personal Genome Machine (PGM) platform. Genomic DNA libraries were prepared using the Life Technologies Cancer panel.

Results: A majority (79%) of the non-GBM tumor group harbored IDH mutations in comparison to GBMs (29%). Although 44% of the non-GBM specimens co-expressed molecular markers such as 1p/19q co-deletion, MGMT promoter methylation and IDH mutations, none of the GBMs co-expressed these markers collectively. Genomic variants using NGS were observed in all groups of gliomas in TP53, PIK3CA, IDH, MET and APC genes. Variants in KIT, SMAD4, FLT3, FLT3, STK11, FGFR3, PDGFRA, KDR, FBXW, PTPN1, BRAF and RBI were also observed at varying frequencies in GBMs and LOH-positive oligodendrogliomas but were absent in the remainder of the non-GBM group. 33% of LOH-positive oligodendrogliomas and 25% of non-GBM harbored alterations in the SMO gene that were entirely absent in the GBMs.

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107

Differential Expression of Genes in Primary and Recurrent GBM Suggests a Transition Toward More Aggressive Molecular Subtype

Gerald Reis1, Wood Matthew1, Jason Huse2, Joanna Phillips1. 1University of California San Francisco; 2Memorial Sloan-Kettering Cancer Center

Introduction: Glioblastoma (GBM) is the most common primary malignant brain tumor in adults and is molecularly heterogeneous. Despite advances in therapy, disease progression is often relentless and prognosis remains poor. It has been suggested that tumors may acquire a more aggressive and potentially more uniform molecular profile at recurrence. To investigate this question we analyzed the gene expression pattern of GBM at initial diagnosis and at recurrence.

Methods: The population consisted of 32 patients, 18 of which were male and 14 female (mean age: 59.8 and 54.4 years, respectively). There were 30 newly diagnosed (NEW) and 10 recurrent (RECUR) GBM, including 7-paired samples. The average time to recurrence was 494 days. RNA was extracted from formalin-fixed paraffin-embedded (FFPE) material, and integrity was verified with Agilent Bioanalyzer. Select genes implicated in GBM (n=173) were tested using the nCounter NanoString Analysis System. Pairwise and between-group comparisons were made using Wilcoxon and Mann Whitney tests.

Results: While the majority of the genes tested (154 genes; 89%) showed no significant change, OLIG2, CD44, and GPNMB were among the differentially expressed genes. OLIG2 was significantly lower in the recurrent group (NEW= 1937+/-349, RECUR=676+/-314; p=0.004) while CD44 was significantly higher (NEW=2410+/-313, RECUR=5183+/-1605; p=0.0282), suggesting a shift toward a more mesenchymal subtype. The expression of GPNMB was significantly higher in recurrent GBM (NEW=796+/-418, RECUR=1347+/-481, p=0.0085), raising the possibility of poorer prognosis in recurrent disease. OLIG2, CD44, and GPNMB showed no significant difference on pairwise comparison (P=0.38, 0.69, and 0.09, respectively), though the sample size is small.

Conclusion: The observed changes in the OLIG2, CD44, and GPNMB expression would suggest a transition towards a more aggressive molecular phenotype in recurrent GBM. In addition, they raise the possibility that primary GBM progression reflects a convergence of molecular subtypes.

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108

Reprogramming Cell Circuits in Glioblastoma

Mario Suvà1, Anoop Patel1, Bradley Bernstein1, David Louis1. 1Massachusetts General Hospital

Developmental fate decisions are dictated by master transcription factors (TFs) that interact with cis-regulatory elements to direct transcriptional programs. In glioblastoma (GBM), a subset of stem-like tumor-propagating cells (TPCs) appears to drive tumor progression and underlie therapeutic resistance, yet remain poorly understood. We identified a core set of neurodevelopmental TFs (POU3F2, SOX2, SALL2, OLIG2) essential for GBM propagation. These TFs coordinately bind and activate TPC-specific regulatory elements, and are sufficient to fully reprogram differentiated GBM cells to ‘induced’ TPCs, recapitulating the epigenetic landscape and phenotype of native TPCs. We reconstruct a network model that highlights critical interactions and identifies novel therapeutic targets for eliminating TPCs. Finally, using single-cell RNA-seq, we show that this network displays a gradient of activity in primary human GBM, thus supporting the relevance of our findings in vivo. Our study establishes the epigenetic basis of a developmental hierarchy in GBM, provides detailed insight into underlying gene regulatory programs, and suggests attendant therapeutic strategies.

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PLATFORM SESSION 6: MUSCLE, NERVE, INFECTIOUS Saturday 8-10 AM, June 14, 2014 Culture Ballroom

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109

Motor Nerve Biopsy

Arthur Hays1, Dale Lange2. 1Therapath; 2Hospital for Special Surgery, New York, NY

Background: Sural nerve biopsy is of limited utility, particularly when the disorder is predominantly or purely motor, such as in motor neuron disease (MND), multifocal motor neuropathy (MMN), chronic inflammatory demyelinating polyneuropathy (CIDP) and other motor neuropathies (MN). When confronting this differential, motor nerve biopsy can potentially help to distinguish motor neuropathies from MND (Riva N et al. Ann Neurol 2011;69:197-201).

Materials & Methods: Biopsy of a motor nerve and muscle were performed in 46 patients. Depending on the distribution of weakness, biopsy was performed on the gracilis muscle and the motor branch of the internal obturator nerve or the pronator teres muscle and its motor nerve supply. Nerve samples were evaluated by routine histology, semithin sections and teased myelinated fibers. Muscle biopsies were prepared by standard methods.

Results: The typical findings in MND included loss of myelinated fibers (sometimes patchy) and sparse myelin debris. Motor neuropathies were identified by nerve fiber loss and excessive regenerative clusters of myelinated fibers or segmental remyelination or both. The differential clinical diagnosis was MND vs MMN or MN in 17 patients because of a non-diagnostic work up. Of these, 11 biopsies yielded a diagnosis of MN rather than MND, 5 had probable MND and one suggested CIDP. Twelve patients met clinical criteria of MND, and the motor nerve was consistent with MND in 9, axonal MN in 2 and equivocal in one. Two were thought to have MMN, but the pathology suggested MND in one and equivocal in the other. Eight of the motor nerve biopsies had no abnormalities. Finally, samples of two other patients suggested CIDP, and one had an inflammatory myopathy.

Conclusions: Motor nerve biopsy is useful in confirming the clinical suspicion of MND and can provide evidence of a potentially treatable neuropathy such as MMN and CIDP.

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110

Treatment with ActRIIB-mFc Improves Lifespan, Behavior and Pathology in the Acta1 H40Y Murine Model of Nemaline Myopathy

Jennifer Tinklenberg1, Hui Meng1, Lin Yang2, Edna Hardeman3, Scott Pearsall4, Robert Fitts5, Michael Lawlor6. 1Medical College of Wisconsin, Milwaukee, WI, USA; 2University of Kentucky, Lexington, KY, USA; 3School of Medical Sciences, UNSW Australia, Sydney, Australia; 4Acceleron Pharma Inc., Boston, MA, USA; 5Marquette University, Milwaukee, WI, USA; 6Medical College of Wisconsin/Children’s Hospital of Wisconsin

Nemaline myopathy (NM) is a congenital muscle disease that can cause a range of clinical symptoms, including perinatal death due to respiratory insufficiency in severe cases. NM is one of the three main types of congenital myopathy, and it is associated with mutations in nine genes (NEB, ACTA1, TPM3, TPM2, TNNT1, CFL2, KBTBD13, KLHL40, and KLHL41). Despite the clinical and genetic heterogeneity seen in human NM patients, it is consistently diagnosed on muscle biopsy by the presence of cytoplasmic rod-like structures (nemaline rods) composed of cytoskeletal material. The interference with skeletal muscle contraction by nemaline rods provides one mechanism for muscle weakness, but there is little correlation between the number of nemaline rods observed and symptomatic severity. Myofiber smallness is also found in many cases of NM and may represent a cause of weakness that can be counteracted by treatment. As we have used ActRIIB-mFc (an inhibitor of myostatin signaling) to promote hypertrophy and increase strength in our prior murine work, we believe that this agent could dramatically improve the weakness seen in NM mice through myofiber hypertrophy. In this study, we tested the efficacy of ActRIIB-mFc treatment in the Acta1 H40Y mouse model of nemaline myopathy, which develops weakness and nemaline rod formation and in which half of male hemizygote mice die between 1-3 months of life. Treatment of Acta1 H40Y mice has thus far shown dramatic improvements in lifespan, animal mass, and open field activity and more modest trends toward increased forelimb grip strength in ActRIIB-mFc treated mice. Our early pathological studies also indicate increased muscle mass (across all major muscles) and increased myofiber size in ActRIIB-mFc treated Acta1 H40Y mice. These preliminary studies demonstrate the substantial potential of ActRIIB-mFc in the treatment of NM.

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111

Keap1/Nrf2 Stress Response Pathway in Autophagic Vacuolar Myopathies

Steve Duleh1, Xianhong Wang1, Marta Margeta1. 1University of California San Francisco

Nrf2, the transcriptional master regulator of antioxidant stress response, is regulated through interaction with its cytoplasmic inhibitor, Keap1. Under basal conditions, Keap1 targets Nrf2 for proteasomal degradation; stimuli that activate Nrf2 signaling lead to dissociation of Keap1/Nrf2 complex, Nrf2 translocation into the nucleus, and increase in the transcription of Nrf2-regulated genes. p62/SQSTM1 – an adapter protein that accumulates following autophagy inhibition and can serve as a diagnostic marker for human autophagic vacuolar myopathies (AVMs) – was recently shown to compete with Nrf2 for Keap1 binding, resulting in Nrf2 pathway activation. In this study, we examined whether Keap1 can serve as an alternative diagnostic marker for human AVMs and whether Nrf2 activity is increased in these disorders. Immunohistochemical staining was performed on formalin-fixed, paraffin-embedded tissue from 47 muscle biopsies divided into five groups: normal control muscle, chloroquine- or colchicine-treated non-AVM control muscle, chloroquine or colchicine AVM, polymyositis, and inclusion body myositis (IBM). In drug-induced AVMs and IBM, but not in control muscle groups or polymyositis, Keap1 antibody labeled the same sarcoplasmic structures as antibodies against p62 and LC3-II (an autophagosome marker that also binds p62); in fact, Keap1 can be used as an alternative diagnostic marker for both AVMs. The sequestration of Keap1 into p62- and LC3-II-positive protein aggregates was associated with increased protein level of Nrf2 target genes Nqo1 and Hmox1 in AVM compared to the control muscle. Similarly, chloroquine treatment of C2C12 muscle cells in vitro led to increase in protein levels of LC3-II and p62 and to nuclear translocation of Nrf2. Taken together, our findings indicate that autophagy impairment leads to upregulation of Nrf2 signaling in human AVMs; future work will show whether Nrf2 activation in this setting is beneficial or harmful, as recently shown for Nrf2-induced redutive stress in the αB-crystallin model of protein aggregation cardiomyopathy.

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112

A Novel MTM1 Mutation in a Case of X-linked Myotubular/Centronuclear Myopathy

Nitin Agarwal1, Helio Pedro2, Karen Valdez-Gonzalez2, Madhuri Hegde3, Ada Baisre1. 1Rutgers New Jersey Medical School; 2Hackensack University Medical Center; 3Emory University

We are reporting a case of X-linked myotubular/centronuclear myopathy [XLMTM] with a novel MTM1 mutation in a 5-week-old baby boy born with severe hypotonia. After extensive testing with normal results, including SMN1/2 copy number analysis, acid alpha-glucosidase enzyme assay, chromosomal microarray, Prader-Willi methylation analysis, very long chain fatty acid profile, pipecolic acid, urine organic acids, plasma amino acids, and complete myotonia evaluation, a muscle biopsy was performed. The biopsy was composed of over 50% Type 1 myofibers, which were slightly larger than the Type 2 fibers. Several fibers, mostly Type 1, either had a single centrally placed nucleus or an area seemingly bare on H&E and myofibrillar ATPases (pH 9.4, 4.6 & 4.3). However these areas showed accumulation of oxidative enzymes and mitochondria that were emphasized with NADH, PAS, acid phosphatase and Gomori-Trichrome. These classical features of myotubular/centronuclear myoapathy prompted molecular/genetic testing. Incidentally a few small foci of extramedullary hematopoiesis were noted in the muscle. When the biopsy revealed findings suggestive of XLMTM, the family recalled additional information in their family history. The patient’s maternal grandmother had had a stillborn son and one of her brothers had also been stillborn. This history seemed to be consistent with diagnosis of an X-linked condition. MTM1 gene testing was then ordered. Sequence analysis of the entire coding region of the MTM1 gene identified a c.142_143delGA mutation in exon 4. This is a two nucleotide deletion that is predicted to result in a shift of the open reading frame leading to the introduction of a premature stop codon. This is to our knowledge the first report of such mutation in XLMTM, and although novel, it is of the type predicted to cause disease.

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113

Congenital Myasthenic Syndrome with Vacuolar Changes and Tubular Aggregates Caused by Mutations in DPAGT1

Carolyn Rysgaard1, Duygu Selcen2, Katherine Mathews1, Andrew Engel2, Steven Moore1. 1The University of Iowa; 2Mayo Clinic, Rochester, MN

Congenital myasthenic syndromes (CMS) are genetically and phenotypically heterogeneous disorders that present a diagnostic challenge to both the neurologist and muscle pathologist. We present a CMS case with diagnostic clues observed in a muscle biopsy. The patient was first evaluated for hypotonia and delayed motor milestones at 12 months of age. She had variable but largely stable motor skills until age 12 years when she presented to clinic with worsened weakness and fatigability affecting her limb-girdle musculature and sparing ocular and bulbar muscles. At age 12 years, a quadriceps biopsy revealed marked fiber size variation, type II fiber predominance and vacuolar changes. Electron microscopy demonstrated small tubular aggregates (TA) and increased lysosomes, confirmed by LAMP2 immunofluorescence. The differential included Pompe disease; however, acid alpha-glucosidase (GAA) enzyme activity was normal. The diagnosis of CMS was reached after finding a decremental response to repetitive nerve stimulation and identifying compound heterozygous mutations in dolichyl-phosphate N-acetyl glucosamine phosphotransferase (DPAGT1) (Selcen et al. Neurology, 2014, in press). Involved in glycosylation and export of acetylcholine receptor (AChR) subunits, DPAGT1 regulates the expression of AChR at the motor endplate. As in this case, previously published patients with DPAGT1 mutations presented in childhood with limb-girdle weakness and minimal ocular or bulbar involvement. Limb-girdle pattern myopathy in patients with CMS has also been associated with mutations in DOK7 and GFPT1. Muscle biopsy TAs are a hallmark of at least two limb-girdle pattern CMS genotypes, DPAGT1 and GFPT1 (also linked to defective glycosylation and decreased AChRs). So while routine muscle biopsies in CMS are generally nondiagnostic, the presence of TAs in the appropriate clinical context can suggest the diagnosis of CMS and guide clinical management. Vacuolar changes with lysosomal proliferation appear to be a novel finding in this patient; their significance is undetermined.

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114

Identification of a Novel Polyomavirus in a Pancreatic Transplant Recipient with Retinal Blindness and Vasculitic Myopathy

Phyllis Faust1, Nischay Mishra2, Roy Rhodes3, Marcus Pereira4, James Pipas5, Komal Jain6, Amit Kapoor2, Thomas Briese8, W. Lipkin2. 1Columbia University, Department of Pathology and Cell Biology; 2Columbia University, Center for Infection and Immunity; 3Rutgers Robert Wood Johnson Medical School, Department of Pathology; 4Columbia University, College of Physicians and Surgeons; 5University of Pittsburgh, Department of Biological Sciences; 6Columbia University, Mailman School of Public Health; 7Columbia University, Epidemiology

A 33-year-old woman was admitted to hospital with 3-week history of fatigue, myalgias, decreasing visual acuity and progressive weakness. Her medical history was notable for Type 1 diabetes mellitus, peripheral neuropathy and retinopathy leading to pancreas transplantation 11 months earlier. She lived in New Jersey, and was evacuated through sewage-contaminated floodwaters in the aftermath of Superstorm Sandy in October of 2012. Her illness began two weeks thereafter. On hospital admission, her creatine kinase was initially normal but increased, peaking at over 8000 U/L. Serum assays by PCR for HIV, EBV, parvovirus and CMV, and for antibodies to influenza, adenovirus, mycoplasma, HTLV and HHV-6 were negative. Her vision deteriorated to detection of only bright light; retinal mapping showed acute macular ischemia due to thrombosis. She developed necrotic plaques on her face, scalp and hands. A muscle biopsy showed microvasculitis and microthromboses in endomysial capillaries, widespread endothelial cell atypia and viral particles in endothelial cell nuclei. Immunostains for cytomegalovirus, herpes simplex virus, adenovirus and BK/JC virus were negative. High-throughput sequencing of nucleic acid from the muscle biopsy revealed a novel polyomavirus with greatest homology to chimpanzee polyomaviruses. In situ hybridization demonstrated viral genome in endothelial cells in muscle and skin biopsies. RNA transcripts encoding large T, small T and a novel T antigen were detected in muscle. After steroid and plasmapheresis treatments, her muscle strength improved but blindness persisted. Blood collected 10 months after illness onset revealed viral DNA in serum, no viral mutations, and antibodies to VP1 and VP2 capsid proteins. New Jersey polyomavirus (NJPyV-2013) is a novel polyomavirus that appears to have tropism for vascular endothelial cells. Whether this infection represents reactivation related to immunosuppression or a newly acquired infection is unknown. Its prevalence and scope of tissue tropism in human infections will require application of diagnostic tests for NJPyV-2013.

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115

Neisseria Elongata Encephalitis Presenting as an Enhancing Brain Mass

Jennifer Cotter1, Daniel Kerrigan2, Atis Muehlenbachs3, Sherif Zaki3, Andrew Bollen1. 1University of California San Francisco, Department of Pathology; 2Pathology Consultants, Springfield, OR; 3National Center for Emerging and Zoonotic Infectious Diseases

Neisseria elongata is a commensal organism typically present in human oral flora, but has been associated with invasive disease, including endocarditis and osteomyelitis. A 25-year-old woman presented with a one-month history of a progressively severe headache. There were some associated visual changes, including “seeing black” when standing. CT imaging demonstrated a large attenuation in the left frontal lobe, with midline shift. MRI showed an enhancing left frontal lobe mass in the subcortical white matter, with surrounding edema. An initial biopsy obtained via left frontal craniotomy showed mild astrogliosis. A second biopsy performed three weeks later demonstrated mixed inflammation with prominent plasma cell and macrophage components, in a perivascular distribution. Vascular involvement by this inflammation was present, without fibrinoid necrosis. The surrounding parenchyma contained reactive gliosis, microglial activation, and microvascular proliferation. The leptomeninges were involved by a macrophage-predominant infiltrate. Overall, the pattern of chronic inflammation was concerning for an infectious etiology, but organisms were not identified by a spectrum of special and immunohistochemical stains (AFB, PAS, GMS, Steiner, Gram, Toxoplasma, HSV, and EBV). Cultures collected from brain tissue and CSF at the time of the second biopsy showed no growth over six weeks. Consultation was sought from the Centers for Disease Control and Prevention. Immunohistochemistry for Treponema spp, other spirochetes, free-living ameba, Mycobacterium spp, and Tropheryma whipplei was negative. No molecular evidence of Treponema pallidum was identified by PCR. Paneubacterial 16S rDNA PCR and sequencing detected DNA with 99% homology to Neisseria elongata. In reviewing the neuropathology literature, Neisseria elongata infection was the presumed cause of a brain abscess in a single case report of a patient with positive blood cultures. Herein we present the first neuropathologic evidence of CNS involvement by this organism. The apparent induction of microvascular proliferation may account for the MRI finding of gadolinium enhancement.

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116

Pathology of HTLV-1 Infection in the Brain

Jiancong Liang1, Jenny Libien1, Marcello DiStasio2, Yuetsu Tanaka3. 1Department of Pathology, SUNY Downstate Medical Center; 2SUNY Downstate Medical Center; 3Department of Immunology, University of the Ryukyus, Japan

Human T-lymphotropic virus type 1 (HTLV-1) infection has been shown to cause progressive spastic paraparesis (HAM/TSP), likely due to spinal cord damage initiated by the infiltrating HTLV-1-infected T cells. Although neurologic and radiologic abnormalities of the brain have been described in HTLV-1 patients, little is known about the underlying pathology in these individuals. We characterized the brains and spinal cords of three HTLV-1 patients who died of adult T-cell lymphoma/leukemia (ATLL) and compared them to three brains and spinal cords of age and gender matched patients with no evidence of neurologic disease. One of the three HTLV-1 patients had a clinical diagnosis of HAM/TSP and none had central nervous system (CNS) involvement with ATLL. On postmortem neuropathologic exam, perivascular cuffing by chronic inflammatory cells, a characteristic finding reported in HAM/TSP, was present in the spinal cords of two patients and in the brain of the one patient with HAM/TSP. We assessed microglial activation and microglial nodule formation using CD68 immunohistochemical stain with quantification by color deconvolution. Microglial nodules were seen in all three brains. Brainstem, cerebellar white matter/dentate nuclei, and spinal cord showed marked microglial activation, while basal ganglia and cerebral white matter exhibited mild to moderate microglial activation, and cerebral and cerebellar cortex and hippocampus did not have elevated microglial activity compared to control brains. By immunofluorescence, HTLV-1 antigens Tax and Rex were detected using Lt-4 and Rec-6 antibodies, respectively, in a few microglia and astrocytes of the brainstem, indicating HTLV-1 infection of non-neuronal cells in the CNS. These findings provide a neuropathologic basis for the clinical symptoms and radiologic abnormalities related to the brains of HTLV-1 patients, and suggest that direct viral infection of the cellular components of the CNS may contribute to the pathogenesis of neurologic disease in HTLV-1 infection.

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PLATFORM SESSION 7: TUMORS 3 – OTHER Saturday 2–4 AM, June 14, 2014 Fashion Ballroom

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117

Altered Receptor Tyrosine Kinase Regulation Promotes Growth Factor Adaptability in Nf2-mutant Schwann Cells

Christian Davidson1, Ching-Hui Liu1, Jeremie Vitte2, Marco Giovannini2, Andrea McClatchey1. 1Massachusetts General Hospital; 2University of California-Los Angeles

Schwannomas are a major source of morbidity in patients with neurofibromatosis type 2 (NF2). Moreover, the sheer number of schwannomas often precludes effective surgical management in these patients. This underscores the need for efficacious medical intervention driven by precise knowledge of the aberrant signaling pathways driving schwannoma formation. We have previously reported that Nf2 deficiency promotes the ability of Schwann cells to grow without their requisite growth factor, neuregulin. We now demonstrate that, even prior to neuregulin independence, neuregulin-dependent Nf2-/- Schwann cells have altered receptor tyrosine kinase (RTK) signaling, as revealed by Western analysis and RTK-targeted drugs. In addition, we also show differential expression of RTKs in murine and human schwannoma cells, and that this difference results in differential pharmacodynamic responses to targeted therapies. Furthermore, these altered RTK dynamics facilitate the ability of schwannomas to adapt to alternative growth factors to fuel their proliferation. Lastly, we present signaling analysis that provides insight into how growth factor adaptability facilitates neuregulin independent proliferation. Collectively, our data establishes a framework for future studies aimed at both biomarker identification and rational design of clinical trials in NF2 patients.

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118

Aberrantly Expressed ErbB4 Promotes Malignant Peripheral Nerve Sheath Tumor Pathogenesis

Stephanie Brosius1, Stephanie Byer1, Amber Moon1, Kevin Roth1, Steven Carroll2. 1University of Alabama at Birmingham; 2Medical University of South Carolina

Malignant peripheral nerve sheath tumors (MPNSTs) are the most common malignancy associated with neurofibromatosis type 1. We have presented evidence that dysregulated signaling by the growth factor neuregulin-1 (NRG1) promotes MPNST pathogenesis. Curiously, MPNSTs may express both erbB3, the only NRG1 receptor found in non-neoplastic Schwann cells, and erbB4, a second NRG1 receptor with distinct functional characteristics. Thus, we hypothesize that aberrant erbB4 expression is common in MPNSTs and that it promotes tumorigenesis via actions distinct from those regulated by erbB3. We found that erbB4 was expressed in 9/9 human MPNST lines and 27/30 (90%) surgically resected MPNSTs. ErbB4 was also uniformly expressed in MPNSTs arising in genetically engineered mice that develop these tumors (P0-GGFβ3; Trp53+/- mice). To establish the function of ErbB4, we generated P0-GGFβ3; Trp53+/-; Erbb4flox/flox mice and ablated Erbb4 in MPNSTs derived from these mice with Ad5CMV-Cre-eGFP. Compared to non-ablated controls, Erbb4 null MPNST cells demonstrated decreases in both proliferation and viability. Orthotopic allografts of Erbb4 null cells also had smaller volumes compared to control grafts. Erbb4 knockdown in human MPNST cells similarly inhibited DNA synthesis and cell viability. Interestingly, Erbb4 ablation did not disrupt Ras or MAPK activation. Antibody arrays instead indicated that erbB4 regulates the PI3K/Akt/mTOR signaling cascade and the phosphorylation of multiple STAT proteins and other molecules that promote oncogenesis. We conclude that aberrantly expressed erbB4 promotes the proliferation and the survival of MPNSTs via the activation of key non-Ras dependent signaling cascades. ErbB4 is thus a candidate for targeted therapy in MPNSTs.

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119

Succinate Dehydrogenase (SDH) Deficiency in Non-familial Pituitary Adenomas

Mark Curtis1, Stacey Mardekian2, Markku Miettinen3, Edward Andrews2. 1Thomas Jefferson University Hospital; 2Department of Pathology,Thomas Jefferson University Hospital; 3Laboratory of Pathology, National Cancer Institute, United States

Succinate dehydrogenase (SDH) deficiency is pro-oncogenic in a variety of tumors, including extra-adrenal paragangliomas, a subset of gastrointestinal stromal tumors, and renal cancers. Whereas SDH deficient tumors typically occur in the setting of germline mutations, SDH deficient tumors also appear sporadically. Several families with germline mutations in the genes encoding SDH have been associated with pituitary adenomas. SDH is an inner mitochondrial membrane-bound, multimeric enzyme that plays a critical role in the Kreb cycle and can transfer electrons directly to the ubiquinone pool. The SDH complex consists of four subunits: SDHA, SDHB, SDHC and SDHD. In order to determine whether SDH deficiency plays a role in sporadic, non-familial pituitary adenomas, we analyzed 37 pituitary adenomas for SDH deficiency from patients with no known familial tumor syndrome and without other known SDH deficiency associated neoplasms. SDH deficiency was determined by immunohistochemistry, since the loss of any of the SDH subunits results in the loss of SDHB expression, owing to destabilization of the entire SDH complex. Of the 37 pituitary adenomas analyzed, 15 patients were female and 22 male. The mean age of the patients was 53 years (range 27-85). Two pituitary adenomas showed complete loss of SDHB staining. Both of these were in males aged 56 years and 27 years. In the 56 year-old, the tumor also showed loss of SDHA staining, indicating the basis of the loss of SDHB. In the literature, SDHA deficiency correlates with an SDHA mutation. Our identification of SDH deficiency in 5.4% of the pituitary adenomas analyzed suggests SDH deficiency may be a relatively common factor contributing to the development of pituitary adenomas in patients not suspected of having a syndrome associated with pituitary adenomas.

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120

Immune Cell Infiltrates in Sparsely Granulated and Densely Granulated Growth Hormone Pituitary Adenomas

Jian-Qiang Lu1, Benjamin Adam1, Robert Broad2, Constance Chik3. 1Department of Laboratory Medicine and Pathology, University of Alberta; 2Department of Surgery/Neurosurgery, University of Alberta; 3Department of Medicine, University of Alberta

The tumor immune microenvironment can influence tumor development and outcome. Immune cell infiltrates in pituitary adenomas are rarely examined. Sparsely granulated growth hormone (SG-GH) pituitary adenomas are distinct from densely granulated GH (DG-GH) pituitary adenomas, as SG-GH adenomas have special properties such as the formation of intracytoplasmic fibrous bodies and different biologic behavior. The cause of these distinctions and specifically fibrous body formation remains enigmatic. This study was to examine immune cell infiltrates in SG-GH adenomas, compared with those in DG-GH adenomas. We examined consecutive cases of SG-GH (n =8) and DG-GH (n=6) adenomas diagnosed between 2009 and 2013; cases with concurrent inflammatory disorders, previous operations (including recurrences) or hemorrhage of the pituitary were excluded from this study. Quantitative analysis was performed by counting and averaging the numbers of immunoreactive cells in 10 ∼ 30 consecutive high-power fields (HPFs). The numbers of CD68-immunoreactive macrophages were found to be significantly higher in SG-GH (8.73 ± 3.73 per HPF) than DG-GH (4.00 ± 2.38 per HPF; p < 0.001, two-tailed t-test) adenomas. There were no statistically significant differences between SG-GH and DG-GH adenomas in infiltrates of CD4-immunoreactive T-cells (4.39 ± 3.79 per HPF and 2.74 ± 2.80 per HPF, respectively; p = 0.38, two-tailed t-test) and CD8-immunoreactive cytotoxic T-cells (4.42 ± 4.56 per HPF and 3.26 ± 2.02 per HPF, respectively; p = 0.53, two-tailed t-test). No difference was seen between two groups in low Ki-67 proliferation index or p53 nuclear immunoreactivity. Our results suggest that SG-GH adenomas possess more macrophage infiltrates, compared to DG-GH adenomas, which may contribute and/or be secondary to the formation of fibrous bodies and other properties of SG-GH adenomas. Since tumor-infiltrating macrophages have been found to be immunosuppressive and tumor supportive, our observation helps understand the more aggressive growth of SG-GH compared with DG-GH adenomas.

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121

Histological Predictors of Progression and Radiosensitivity in Cranial Atypical Meningioma after Total or Subtotal Resection

Chunyu Cai1, Sam Sun2, Richard Perrin1, Clifford Robinson3, Michael Chicoine4, Albert Kim4. 1Department of Pathology and Immunology, 2Washington University School of Medicine; 3Department of Radiation Oncology, Washington University School of Med; 4Department of Neurosurgery, Washington University School of Med

Introduction: Atypical meningiomas (AM), WHO grade II, are histologically heterogenous and exhibit a broad range of clinical aggressiveness. The necessity of adjuvant radiation following gross total resection (GTR) or subtotal resection (STR) remains controversial. The goal of the current study is to assess clinicopathological features associated with progression of cranial AM following resection and with the efficacy of adjuvant radiation therapy (ART).

Methods: Fifteen clinicopathological parameters were analyzed from a total of 210 AM patients who underwent either primary GTR (151 patients, 45 months mean follow-up), or primary STR (59 patients, 67 months mean follow-up) at Barnes Jewish Hospital. The impacts of ART on progression were assessed by Kaplan-Meier analysis.

Results: The risk factors for disease progression differed between patients who underwent GTR and STR. After GTR, 13/151 patients (8.6%) experienced recurrence (median 47.0 months). Multivariate analysis identified mitotic index (MI) ≥8HPF (p = 0.006) and brain invasion (p = 0.001) as predictors of recurrence. Kaplan- Meier analysis demonstrated no difference in progression-free survival (PFS) or overall survival (OS) after GTR vs. GTR/ART (p = 0.9, p = 0.9). After STR, 27/59 patients (46%) experienced recurrence (median 30 months). Compared to STR alone, ART post STR significantly improved PFS in AMs without spontaneous necrosis (p=0.001), but had no such effect in AMs with spontaneous necrosis (p=0.6).

Conclusions: AM recurrences post GTR are uncommon; after GTR, ART should be reserved for cases with brain invasion or MI ≥8/10HPF. Post STR, ART significantly improves PFS but only for tumors without spontaneous necrosis. The latter finding indicates that spontaneous necrosis may be used as a biomarker for “triaging” patients to receive ART post STR, and suggests that tumoral ischemia might confer radioresistance.

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122

Meningiomas That Meet Grade II by 3 Criteria Have an Increased Rate of Recurrence

Christina Appin1, Matthew Schniederjan1, Stewart Neill1, Robert Press1, Roshan Prabhu1, Ian Crocker1, Daniel Brat1. 1Emory University School of Medicine, Atlanta, GA

Background: Meningiomas are graded according to the World Health Organization (WHO) criteria as grade I, II or III. While grade I meningiomas are considered benign, grade II and III tumors recur more frequently and have a worse prognosis. Criteria for classification as grade II include 1) brain invasion; 2) specific histologic patterns (chordoid and clear cell); 3) increased mitoses; and 4) the presence of at least 3 of 5 atypical features (increased cellularity, sheet-like growth, prominent nucleoli, ’spontaneous’ or ’geographic’ necrosis and small cells with a high nuclear to cytoplasmic ratio). The purpose of this study was to determine whether meningiomas that meet more than one criterion for grade II are more aggressive than those that meet only one criterion.

Design: Seventy-nine grade II meningiomas resected at Emory University Hospitals from 1999-2010 were included. Slides, pathology reports, and patient charts from each case were reviewed. Meningiomas were grouped according to number of histopathologic criteria met to establish a grade II diagnosis and correlated with time to recurrence and survival data. Median follow-up time was 3.2 years. Statistical analysis was performed using Fisher’s exact test and SAS-JMP software.

Results: Meningiomas that met 3 criteria (n=8) for classification as a grade II neoplasm had a higher recurrence rate (87.5%) than those that only met one (n=43) or two (n=26) criteria (30.2%; p=0.00397 and 30.7%; p=0.0112, respectively). No significant difference in recurrence rates were noted for meningiomas that met one criterion compared to those that met two. Within the timeframe of follow-up, time to recurrence and patient survival did not differ between study groups.

Conclusion: Meningiomas that meet 3 criteria for grade II have a higher recurrence rate than those that meet 1 or 2 criteria.

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123

NHERF1/EBP50 is a Marker of Ependymoma

Maria-Magdalena Georgescu1, Paul Yell1, Ping Shang1, Kimmo Hatanpaa1, Lauren Langford2, Charles White 3rd1, Jack Raisanen1. 1Universit of Texas Southwestern; 2MD Anderson Cancer Center

NHERF1/EBP50 (Na+/H+ exchanger 3 regulating factor 1; ezrin-radixin-moesin (ERM) binding phosphoprotein 50), an adaptor molecule composed of two tandem PDZ domains and a carboxyl-terminal ERM-binding region, is predominantly localized at the apical plasma membrane of normal epithelia and has been recently implicated in the progression of various human malignancies. We generated NHERF1-deficient mice and described structural defects of the intestinal brush border membrane, as well as lactation deficits. We report here that non-obstructive hydrocephaly also develops with partial penetrance in NHERF1-deficient mice. Examination of mouse and human brain tissues revealed strong NHERF1 expression at the apical plasma membrane of ependymal cells, suggesting contribution of these cells to the development of hydrocephaly, perhaps through abnormal cilia motility. Based on these findings and on our previous functional and mechanistic studies showing a tumor suppressor role for NHERF1 by delocalization from the apical plasma membrane, we analyzed the expression and localization of NHERF1 in a series of tumors of ependymal origin. Remarkably, we found a robust staining for intracytoplasmic lumens (ICLs) in 100% of ependymomas (n=13), including the cases with negative EMA staining. A similar but focal staining pattern was also present in subependymomas (n=7) and in all but one of the anaplastic ependymoma cases examined (n=5). True rosettes were highlighted by NHERF1 staining in both ependymomas and anaplastic ependymomas. This staining was not present in other tumors, including meningioma, schwannoma, brainstem glioma and oligodendroglioma, and was only rarely seen in glioblastoma (2/10 cases). These preliminary data denoting NHERF1 as marker for ependymoma will be validated on a higher number of cases. Since we have also characterized and reported alterations of NHERF1 in glioblastoma and colorectal cancer, the differential diagnosis, functional implications and in vitro modeling for therapeutic testing will be discussed.

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124

A Novel Medulloblastoma Model Mimicking Human Disease

Guo Zhu1, Sherri Rankin2, Lionel Chow3, Chunxu Qu2, David Ellison2, Suzanne Baker2. 1St. Jude Children’s Res. Hosp. and The University of Tennessee HSC; 2St. Jude Children’s Research Hospital; 3Cincinnati Children’s Hospital Medical Center

PTEN and TP53 are recurrently mutated tumor suppressor genes in human medulloblastoma. Pten loss induces diverse outcomes in different developmental contexts. To investigate Pten function in early postnatal brain development and tumorigenesis, we conditionally inactivated Pten in neural stem/progenitor cells in neonatal brain using Nestin-CreER transgenic mouse. Pten inactivation created a perivascular arrangement of hyperplastic atypical undifferentiated cells in the cerebellum that did not progress to neoplasia during the lifespan of the mouse. Co-deletion of Pten and Trp53 cooperated to produce fully penetrant medulloblastomas originating from cells in these perivascular locations. Pten / Trp53 double knock-out medulloblastomas showed a neuronal immunophenotype and a vascular network with a hemangiopericytic morphology. The neuronal immunophenotype included immunoreactivity for GabaARα6, which was absent in tumors arising from Tp53 knockout alone.

EdU pulse-chase experiments demonstrated a perivascular cancer stem cell population in double knock-out medulloblastomas. The Pten and Trp53 double knock-out medulloblastomas showed recurrent somatic inactivating mutations in the tumor suppressor gene Ptch1. Gene expression profiles showed that this model recapitulated the subgroup of human medulloblastomas with de-regulated SHH signaling. The models presented here show that Pten loss is sufficient to drive the formation of a pre-neoplastic ‘perivascular niche’ that can be transformed into medulloblastoma by additional mutations. We conclude that Pten normally acts to prevent ongoing proliferation of a normally undetectable stem cell/progenitor niche in postnatal cerebellum capable of acting as the cell of origin for the SHH subgroup of medulloblastoma.

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PLATFORM SESSION 8: NEURODEGENERATIVE 2 - ALS, FTD, PD, OTHER Saturday 2–4 AM, June 14, 2014 Culture Ballroom

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125

Incidental Brain Pathologies in Prospectively Followed Normal Elderly Brain Bank Volunteers

Brittany Dugger1, Joseph Hentz2, Charles Adler2, Marwan Sabbagh1, Holly Shill1, Sandra Jacobson1, John Caviness2, Christine Belden1, Erika Driver-Dunckley2, Kathryn Davis1, Lucia Sue1, Thomas Beach1. 1Banner Sun Health Research Institute; 2Mayo Clinic, Scottsdale Arizona

We recently investigated the clinicopathological outcomes of initially normal elderly subjects who enrolled in our brain and body donation program (Dugger et al. J Neuropathol Exp Neurol. 2014). In addition to the defining histopathology for certain neurodegenerative disorders, other pathologies may exist within the brain, including cerebral white matter rarefaction (CWMR), argyrophilic grains (Arg), cerebral amyloid angiopathy (CAA), and glial tauopathies. Furthermore, deposits of proteins that define certain diseases, such as TAR DNA binding protein-43 (TDP-43), may exist but cases lack corresponding clinical features during life. We sought to determine the frequencies of these pathologies in a prospectively-assessed, community-based autopsy series. We utilized the same 119 cases from our previous study, all having normal cognitive and movement disorder assessments at study entry. Of these cases, 52% were female, median age at study entry was 83.5 years (range 67 to 99), and median duration from first visit until death was 4.3 years (range 0-10). All 119 cases contained at least one of the examined pathologies. With respect to the 87 subjects who were still clinically normal at death, 47 (54%) had CWMR, 22 (25%) Arg, 37 (43%) CAA, 8 (9%) glial tauopathies, and 41 (47%) TDP-43. Of the 30 who converted to a clinicopathologically defined neurodegenerative disease by the time of death, 27 cases (90%) had CWMR, 12 (40%) with Arg, 19 (63%) with CAA, 6 (20%) with glial tauopathies, and 15 (50%) with TDP-43. Pathology groups are not mutually exclusive; particular overlap was found. Although limited by a relatively small sample size, these other pathologies found within initially normal elderly subjects represent a rough estimate of the incidence that exist within brains of elderly individuals over a defined time period.

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126

Neuropathologic Analysis Of 59 Centenarian Brains

Masaki Takao1, Shigeo Murayama1, Hiroyuki Sumikura1, Akane Nogami1, Akiko Uchino1, Yuta Nakano1, Hiroyuki Hatsuta1, Maki Obata1, Nobuyoshi Hirose2. 1Tokyo Metropolitan Geriatric Hospital and Institute of Gerontology; 2Department of Geriatric Medicine, School of Medicine Keio University

Purpose: To describe neuropathologic background of the centenarian brains.

Materials and Methods: From 1972 to 2013, we obtained 59 centenarian brains out of 8538 autopsy cases in our hospital. We reviewed their medical records and analyzed neuropathology using standard protocols and staging methodologies.

Results: The mean age at death was 101.7 ± 2.1 y (range; 100-111). There were 11 men and 48 women. In 43 individuals that we could obtain CDR scores (0-0.5, 19 cases; 1-3, 24 cases), high likelihood AD cases were 0 in CDR 0-0.5 and 5 in CDR 1-3. The brain weight of men (1212.3 ± 28.4 g) was heavier than that of women (1066.6 ± 13.6 g). 53 cases (89.8%) belonged to Braak NFT stages I-II (n=14) and III-IV (n=39). Only 6 cases (10.1%) were stage V. There were no brains of stage VI. Senile plaques (SPs) grading of CERAD was as follows: none in 10, sparse in 21, moderate in 11 and frequent in 17 cases. According to NIA-Reagan criteria of Alzheimer disease (AD), low, intermediate and high likelihood ADs were 12 (20.3%), 9 (15.2%) and 6 (10.2%) cases, respectively. NFT predominant pathology (NFT stage = III-VI/CERAD SP grading = none or sparse) was seen in 19 cases (32.2%). There were NFT stage I-II/moderate SPs in 2 cases, and NFT stage III-IV/frequent SPs in 11 cases. Although Lewy body (LB) pathology was observed in 19 (32.2%) cases, 17 cases remained in incidental or pre-symptomatic condition. Tufted astrocytes were rarely seen (n=3, 5.1%). Argyrophilic grains were present in 34 cases (57.6%). 23 cases (39.0%) had cerebral infarcts.

Conclusions: In the centenarian brains, advanced AD pathology as well as LB pathology may be uncommon condition.

(Supported in part by CBSN, MHLW and Daiwa Securities Health Foundation.)

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127

Association Between Hippocampal Sclerosis of Aging (HS-Aging) Pathology and Sulfonylurea Drug Exposure in NACC

Peter Nelson1, Erin Abner1, Walter Kukull2, Sarah Monsell2, Eseosa Ighodaro1, David Fardo1. 1University of Kentucky; 2University of Washington

Hippocampal sclerosis of aging (HS-Aging) is a prevalent brain disease of the elderly with no known environmental risk factors. HS-Aging is currently defined by pathological manifestations: neuron loss, gliosis, and atrophy in the hippocampal formation, not associated with Alzheimer’s-type plaques and tangles. HS-Aging is among the most prevalent neuropathologic features of advanced old age. Substantial disease-specific cognitive impairment is directly associated with HS-Aging when comorbid pathologies are taken into consideration. We tested the hypothesis (based on a human genomic screen beyond the scope of this Abstract) that exposure to sulfonylurea drugs is associated with altered risk for HS-Aging pathology. Sulfonylureas are a popular class of orally administered anti-diabetic medication. Data were obtained from the National Alzheimer’s Coordinating Center (NACC) database with both detailed longitudinal drug history and postmortem data. The study sample was 624 patients who died after age 85, between 2010-2013, with longitudinal drug history and autopsy data (n=108 with HS-Aging pathology, n=517 without HS-Aging pathology). “Sulfonylurea use” indicated self-reported use of any sulfonylurea drug at any longitudinal visit. Crude OR for use of any sulfonylurea for individuals with HS-Aging pathology was 2.14 (95% CI: 1.06–4.68, p=0.03); age-adjusted OR=2.19 (95% CI: 1.04–4.63, p=0.04). The higher HS-Aging pathology in persons taking sulfonylurea medication was not apparently associated with the patients’ year of pathological diagnosis, nor did sulfonylurea users have higher burden of AD pathology. HS-Aging pathology does not appear to be associated directly with diabetes per se because history of diabetes has a null association with HS-Aging pathology – for self-reported diabetes (age-adjusted OR=1.08 (95% CI: 0.58-2.03). These data provide preliminary support for the hypothesis that among elderly human subjects, sulfonylurea use may cause or exacerbate HS-Aging pathology. This study also underscores the importance of pathology-based stratification of dementia etiology.

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128

The Role of the Unfolded Protein Response in Sporadic and Lrrk2-Driven Parkinson’s Disease

Annie Hiniker1, Scott Oakes1. 1University of California San Francisco

Delineating the mechanisms leading to dopaminergic neuronal death in Parkinson’s disease (PD) is an unsolved problem that is critical to the development of effective PD therapeutics. A large body of evidence points to a key role for protein misfolding and aggregation in PD; however, activation of the unfolded protein response (UPR), an essential network of intracellular signal transduction pathways that reacts to misfolded proteins and modulates the cell’s response, has not been fully assessed. We are exploring the role of the UPR in PD using a multifaceted approach. First, we are assessing the activation status of the UPR in a cohort of 20 PD patients and 20 controls (not significantly different in age, gender, brain weight, or postmortem interval) using immunohistochemical evaluation of a panel of proteins known to be upregulated or phosphorylated under UPR activation. Second, we are exploring the possibility that Lrrk2, the gene most commonly mutated in genetic PD, may be important in triggering the UPR through an interaction with the ER transmembrane kinase/RNase Ire1a. Ire1a is a master regulator of the UPR that allows cells to adapt to remediable stresses but signals apoptosis under extreme stress conditions. Our lab has built a series of powerful genetic tools to independently control Lrrk2 and Ire1a and test their effects on cell survival. Finally, we will apply our immunohistochemical panel for UPR activation to a small cohort of PD patients with known Lrrk2 mutations. Better small molecule inhibitors against components of the UPR, including Ire1a, have recently become available. Therefore, if we find that neurodegeneration in PD is related to activation of UPR-induced apoptosis, we are positioned to rapidly test whether modulating the UPR has therapeutic benefit in pre-clinical models of PD.

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129

The Basal Forebrain and Hypothalamus are Involved in a Subset of Patients with Amyotrophic Lateral Sclerosis (ALS)

Matthew Cykowski1, Hidehiro Takei1, Paul Schulz2, Suzanne Powell1. 1Houston Methodist Hospital; 2University of Texas Medical School at Houston

Background: Amyotrophic lateral sclerosis (ALS) is associated with frontotemporal cognitive impairment and autonomic nervous system (ANS) dysfunction. Given the weight loss, changes in metabolism, and autonomic dysfunction observed in ALS, and changes in cognition, we hypothesized that there would be neuropathologic involvement of basal forebrain and hypothalamus in ALS patients.

Materials and Methods: Autopsy materials and records from 29 ALS patients were studied with IRB approval. Anatomic regions evaluated for TAR DNA-binding protein (TDP-43)-positive inclusions and neurites included the lateral septal area (LSA), bed nucleus of the stria terminalis (BNST), ventral pallidum (VP), ventral striatum including Islands of Calleja (VS/IsC), substantia innominata (SI) including nucleus of the diagonal band (NDB) and nucleus basalis (NB), medial hypothalamic preoptic (HPO) and tuberal levels (HTL), including the paraventricular nucleus (HPV), posterior (PHA) and lateral (LHA) hypothalamic areas, mammillary body and adjacent tuberal nuclei (MB/TN), and supraoptic nucleus (SON).

Results: Patients (23 males, 6 females) had a mean age + standard deviation (SD) of 62.1 +/- 8.51 years and a mean brain weight + SD of 1344.5 +/- 145.6 grams. Five (of 24) cases had hypothalamic involvement, including LHA (4 cases), PHA (3), HTL (3), MB/TN (3), and PO/PV (1). Five (of 22) cases had involvement of small or medium-sized neurons in SI/NDB/NB; a single case had filamentous inclusions in the NDB. Seven (of 21) cases had involvement of VS/IsC. BNST and LSA were involved in six (of 17) and one (of 10) cases, respectively. VP was not involved (18 cases).

Conclusions: Hypothalamic and basal forebrain involvement occurred in three patients with only cord, brainstem, and/or agranular frontal cortex involvement, whereas in five other patients it was associated with more widespread TDP-43 deposition. This observation raises the possibility that hypothalamic/basal forebrain TDP-43 deposition may precede deposition in other well-known locations for ALS pathology.

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130

Methylation of RAN Translated Glycine-Arginine Dipeptide Repeats in C9ORF72 FTLD and ALS

Dennis Dickson1, Kevin Bieniek2. 1Mayo Clinic; 2Mayo Clinic Graduate School

We have previously described repeat-associated non-ATG (RAN) translation of the expanded GGGGCC hexanucleotide repeat in C9ORF72 frontotemporal dementia and amyotrophic lateral sclerosis (c9FTD/ALS). However, the cytotoxicity of the generated dipeptide repeats (DPR) remains to be determined. One potential mechanism is through arginine methylation of glycine arginine repeats. With endogenous RGG/RG motif-containing proteins facilitating cellular pathways such as DNA damage signaling, transcription, pre-mRNA splicing, translation, and apoptosis, it is conceivable that generation of the (GR)n DPR can disrupt equilibrium in these systems. We observed arginine methylation of neuronal cytoplasmic inclusions in c9FTD/ALS with immunohistochemistry. Immunoreactivity was observed for monomethylarginine as well as symmetric and asymmetric dimethylarginine. These lesions corresponded to anatomical regions of high DPR neuropathologic burden, including the cerebellum and hippocampus. In addition, methylarginine-immunopositive neuronal cytoplasmic inclusions were often “star-shaped” in morphology, similar to DPR and p62/sequestosome-1 in c9FTD/ALS. Immunohistochemistry of serial sections in the hippocampus with polyclonal antibodies specific to dimethylarginine and the (GR)n DPR revealed colocalization of these epitopes suggestive of direct arginine methylation of this DPR repeat. Arginine methylation of RAN translated (GR)n DPR appears to be a novel and prominent neuropathologic feature of c9FTD/ALS that may have important implications for disease mechanism and future treatment strategies.

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131

C9orf72 Hypermethylation Influences Repeat Expansion Associated Pathology in ALS/FTD

Elaine Liu1, Jenny Russ1, Kathryn Wu1, Donald Neal1, Eunran Suh1, Anna McNally1, David Irwin1, Vivianna Van Deerlin1, Edward B. Lee1. 1University of Pennsylvania

Hexanucleotide repeat expansions of C9orf72 are the most common genetic cause of amyotrophic lateral sclerosis and frontotemporal degeneration. The mutation consists of a large intronic GGGGCC repeat expansion which is associated with reduced C9orf72 expression, leading to the suggestion that haploinsufficiency contributes to disease. The repeat expansion is also associated with the accumulation of potentially toxic RNA and protein aggregates, supporting the hypothesis that the C9orf72 mutation may cause disease through a toxic gain of function. CpG methylation is known to protect the genome against unstable DNA elements and to stably silence inappropriate gene expression. C9orf72 promoter methylation has recently been associated with the repeat expansion mutation. We report here detailed characterization of C9orf72 promoter methylation from human peripheral blood and brain tissue of repeat expansion mutation carriers. Based on studies in lymphoblastoid cell cultures from repeat-expanded and non-expanded ALS patients, we found that C9orf72 methylation leads to transcriptional silencing which can be reversed by the DNA methyltransferase inhbitor, 5-aza-2’-deoxycytidine. C9orf72 promoter methylation also significantly reduces cellular vulnerability to oxidative and autophagic stressors, accumulation of RNA foci, and accumulation of dipeptide repeat protein aggregates. These results indicate that epigentic silencing of mutant C9orf72 allele may modulate disease pathogenesis in amyotrophic lateral sclerosis and frontotemporal degeneration linked to hexanucleotide repeat expansions.

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132

A Loss of Function Mutation in the C9ORF72 Mouse Ortholog Results in Motor System Degeneration

Daniel Mordes1, Naoki Suzuki2, Johnny Salameh3, Morag Stewart4, Asif Maroof2, Steve Han3, Robert Brown3, Kevin Eggan5. 1Massachusetts General Hospital; 2Harvard Stem Cell Institute, Harvard University; 3Department of Neurology, University of Massachusetts Medical School; 4Program in Cellular and Molecular Medicine, Boston Children’s Hospital; 5HHMI, Harvard Stem Cell Institute, Harvard University

A hexanucleotide repeat expansion at C9ORF72 has recently been found in a significant fraction of patients suffering from amyotrophic lateral sclerosis (ALS). However, it remains to be determined whether this mutation acts through a gain of function or loss of function mechanism. We have recently shown that the mouse C9ORF72 ortholog was most highly transcribed in the neuronal populations that are sensitive to degeneration in ALS and frontotemporal dementia. Here we show that heterozygous and homozygous mice harboring a loss of function mutation in the ortholog of C9ORF72 are viable, survive to adulthood and initially display motor system functionality similar to their wild type litter mates. However, as heterozygous animals aged, we found they displayed a significantly increased rate of mortality. Death in heterozygous animals was associated with declining motor function that was accompanied by muscle atrophy. Homozygous mutant animals displayed a similar but accelerated and quantitatively more severe phenotype. Immunohistochemical analysis of CNS tissue for ALS-associated pathology will be presented. Overall, these findings support the hypothesis that reduced C9ORF72 function and haploinsufficiency resulting from the repeat expansion that many patients harbor contributes directly to the development of ALS.

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POSTERS: DAY 2 Saturday 10–10:30 AM, 4–4:40 PM, June 14, 2014 Foyer

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133

Autopsy Neuropathology of a Case of Microcephaly-Thin Corpus Callosum Syndrome

Malak Abedalthagafi1, David Urion2, Elizabeth Engle2, Hannah Kinney2, Rebecca Folkerth1. 1Brigham and Women’s Hospital- Harvard University; 2Boston Children’s Hospital- Harvard University

A syndrome of microcephaly and thin corpus callosum in patients with spastic quadriparesis and global cognitive deficits has been described on the basis of neuroimaging in families of Arab heritage. Recently, one such disorder was mapped to 8q23.2-q24.12 in 4 members of 3 consanguineous Bedouin families (Halevy et al, 2012. Pediatr Neurol 46:363). The underlying neuropathology of the syndrome, however, has not been reported, and thus the neuroanatomic substrate is poorly understood. We performed autopsy evaluation of an 18-year-old woman with spastic quadriparesis and global delay with microcephaly-thin corpus callosum with the additional clinical feature of medically refractory epilepsy. She was of Pakistani origin, and had a sibling with a similar neurologic disorder by family report, and death at age 7. We found profound microcephaly (brain wt 350g, expected 1100g), with sparing of the cerebellum. The temporal lobes were also distportionately small. The corpus callosum was razor-thin throughout its length, and disproportionately smaller than the hypoplastic white matter of the parietal, frontal, and occipital lobes. A small white matter bundle resembling a Probst bundle was noted in the periventricular frontal region, suggesting that the thin corpus callosum was due to dysfunctional axonal guidance across the midline. Descending corticospinal tracts through the brainstem bilaterally were hypoplastic, suggesting defective axonal guidance of the pyramidal system. Cerebral cortical abnormalities included persistent vertical cortex, poor gray-white distinction with prominent interstitial white matter neuron population, and focal subarachnoid glioneuronal heterotopia. Basal nuclei were unremarkable. Death was due to respiratory infection complicating a neurologically debilitated state. The major pathology was developmental in origin and not acquired. We postulate that it is due to a mutation in a gene critically involved in both neuronal development (proliferation, migration) and axonal targeting. The gene defect is under investigation.

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134

Rhombencephalosynapsis in a Patient with Goldenhar Syndrome

Melissa Gener1, Stephanie Slemp1, Justin Richey1, Dean Hawley1, Jose Bonnin1. 1Indiana University School of Medicine

Goldenhar syndrome (facioauriculovertebral sequence, FAV sequence) is a rare genetic disorder characterized by anomalies in the eyes, ears, and vertebral column. Central nervous system anomalies have been reported in approximately 50% of cases. They range from mild hydrocephalus to severe malformations, including Arnold-Chiari malformation, with or without syringomyelia or hydromyelia, encephalocele, holoprosencephaly, arhinencephaly, lissencephaly and others. Cerebellar abnormalities have been reported in association with Goldenhar syndrome. We report a case of an 11-year-old female with this condition. Neuroimaging studies revealed ventriculomegaly and diffuse white matter loss, particularly in the peritrigonal region, atrophy of the corpus callosum, hypoplasia of the middle ear, fusion of the ossicles, hypoplasia of the right external auditory canal and absence of the right temporalis muscle. Fusion of the cerebellar hemispheres was also documented. A CT scan of the spine showed focal levoscoliosis in the lumbar region, centered at L3-L4 as well as multiple segmentation anomalies of the midthoracic vertebral bodies and S-shaped scoliosis of the thoracic spine. She had esophageal atresia with tracheoesophageal fistula as well as a history multiple gastrointestinal surgical procedures including a Nissen fundoplication and G-tube placement. She also had a horseshoe kidney with multiple episodes of urinary tract infections. The patient was admitted because of severe abdominal distention following two days of diarrhea. She deteriorated rapidly and went into cardiac arrest. Resuscitation efforts were unsuccessful. An autopsy confirmed the presence of multiple anomalies in the spine, absence of right pinna, bicuspid aortic valve, horseshoe kidney and hydrocephaly. Examination of the hindbrain revealed absence of the cerebellar vermis and fusion of the cerebellar hemispheres and dentate nuclei. Although cerebellar hypoplasia has been observed in association of Goldenhar syndrome, to our knowledge, rhombencephalosynapsis has not been previously reported in association with this syndrome.

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135

Characterization of Neuropathologic Findings in a 16 month-old Female with Pfeiffer Syndrome

Bret Evers1, Dennis Burns1, Charles Timmons2, Veena Rajaram2. 1UTSW; 2UTSW/CMC

Pfeiffer syndrome, a rare genetic disease caused by mutations in members of the fibroblast growth factor receptor (FGFR) family, is characterized by multiple developmental anomalies, namely craniosynostoses, mid-face hypoplasia, digit abnormalities, and mental retardation. Currently, the neuropathologic findings in Pfeiffer syndrome have been limited to those found in four fetal cases. Here, we describe the neuropathologic abnormalities observed in a 16 month-old female with a confirmed S351C mutation in FGFR2. Gross autopsy findings of the CNS included megalencephaly, hydrocephalus, temporal lobe enlargement with hippocampal dysmorphogenesis, atrophy of the corpus callosum, absence of the septum pellucidum, Chiari type I malformation, and caudal deviation of the brainstem with a small pons. Microscopically, foci of polymicrogyria and cerebral white matter neuronal heterotopias were observed, indicative of a neuronal migration defect; and sections through the hippocampi revealed moderate to severe disorganization of the hippocampal pyramidal cell layer and dentate gyrus. Additionally, brainstem sections showed generalized white matter atrophy of the cerebral peduncles, basal pons, and medullary pyramids. These various findings can be explained by the role of FGFR2 in the development and differentiation of the CNS. Our studies not only further characterize the neuropathology of Pfeiffer syndrome but also are the first to provide details on its postnatal course.

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136

Novel Nonsense Mutation of FLNA in a 26 year Old Female with Periventricular Nodular Heterotopias and Severe Emphysema

Patricia kirby1, Erica Savage2, Benjamin Darbro2. 1University of Iowa Hospitals and Clinics, Dept of Pathology; 2University of Iowa College of Medicine

Numerous mutations of the filamin A gene (FLNA) have been reported in the English literature with the most common association being periventricular nodular heterotopias (PVNH). As filamin A is integral in cell cytoskeleton formation, mutations also result in non-CNS abnormalities including cardiac, skeletal, platelets, pulmonary, gastrointestinal, soft tissue and urogenital pathology. PVNH together with pulmonary disease is uncommon, presenting almost exclusively in infants with a spectrum of tracheobronchomalacia, atelectasis, pulmonary cysts and congenital emphysema. Pulmonary hypertension and oxygen dependence are reported in rare survivors.

We report a 26 year old female with Scheuermann’s kyphosis in whom bilateral PVNH and severe emphysema were found at autopsy. She had presented for surgical correction of progressive kyphosis to ameliorate worsening dyspnea. Her past medical history was notable for moderately severe aortic and mitral incompetence, macrothrombocytopenia, joint laxity and prior inguinal and hiatus hernia repairs. She was of normal intelligence and had no dysmorphic facial features, extremity anomalies or history of seizures. Her post-operative course was complicated by an abrupt onset of pulmonary hypertension with congestive cardiac failure. A saddle embolus was suspected but not identified on emergent surgical exploration. She developed a refractory bleeding diathesis with subsequent multiorgan failure. Death occurred on post-operative day 14.

Autopsy revealed severe pulmonary emphysema and diffuse bilateral PVNH with glomeruloid blood vessels. Whole exome sequencing was performed on DNA extracted from peripheral blood. An average of 100x coverage was obtained across the targeted exonic regions. Focused analysis of the FLNA gene revealed a novel nonsense mutation in exon 11 that is similar to those known to cause PVNH (NM_001456.3 c.1675G>A, p.Q559*).

This patient’s numerous pathologies expand the phenotypic presentation of FLNA cases in the literature.

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137

Cortical and Leptomeningeal Angiomatosis Associated Epilepsy in a 68 Year Old Female: A Case Report

Nishant Tiwari1, Daniel Grimmer1, J Goodman1, Meenakshi Bhattacharjee2. 1Baylor College of Medicine, Houston, TX; 2University of Texas Medical School, Houston, TX

Sturge Weber Syndrome (SWS) can be classified clinically based upon the extent of disease and location of the angiomatosis. Sporadic type III SWS is the CNS selective entity, associated with isolated leptomeningeal and cortical involvement without facial port wine stain. These patients frequently present with refractory epilepsy and migraine headaches. Symptoms usually develop early, and more than 95% cases are symptomatic by 5 years. Very rarely have patients presented at older ages.

We present a 67 year old woman who developed epilepsy at age 52. She does not have migranous headaches, port wine stains or glaucoma, or significant family history. MRI showed linear enhancement, with cortical calcifications and atrophy in the left frontal lobe. Biopsy of the lesion in the non-eloquent cortex was performed.

Pathologic examination showed increased cortical parenchymal and meningeal vasculature, and associated gyral calcifications associated with the cortical vessels. Immunohistochemistry was performed to better define the pathology. GFAP highlighted reactive astrocyosis. Neu-N staining showed mild dysplastic features, and phosphorlylated neurofilament revealed rare stained neurons. The pathologic features were consistent with a diagnosis of SWS with associated cortical dysplasia.

To the best of out knowledge, this case appears to be the oldest presentation and diagnosis reported in the english literature. In SWS Type III, the diagnosis of purely CNS disease can be delayed due to lack of clinical suspicion. However, the recently described GNAQ somatic mutation (c.548G→A, p.Arg183Gln) seen in up to 88% of patients with SWS may help in recognition of a wider spectrum of the disease.

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138

Neuropathology of a Case of Joubert Syndrome with an Unusual Genetic Background

Douglas Miller1, Eric Destrampe2, Carl Stacy2. 1University of Missouri School of Medicine; 2Dept of Pathology & Anatomical Sciences, U Missouri School of Medicine

A 3 year old died unexpectedly. He carried a diagnosis of Joubert Syndrome (JS), with multicystic kidneys, retinal dystrophy, epilepsy, ptosis, hypospadias, and brain abnormalities. He had been unable to walk or talk, had severe developmental delay, and had episodic tachypnea (a feature of some cases of JS). He allegedly watched television but did not track a light or finger visually. He was hypotonic with normal deep tendon reflexes. Imaging had disclosed vermian aplasia and a “molar tooth” conformation of the midbrain and pons in axial images. Genetic testing revealed heterozygosity for two different mutations in the CEP290 gene and an unrelated deletion from 1p21.3.

Neuropathological examination confirmed vermian aplasia with dilatation of the aqueduct and fourth ventricle but no cisterna magna cyst. The midbrain was short rostro-caudally with thickening and elongation of the undecussated superior cerebellar peduncles (giving the “molar tooth” appearance). The corticospinal tracts did not decussate in the medulla, consequently there were no pyramids; the pontine pyramidal tracts were small and laterally displaced. The oculomotor nerve fascicles arose from dispersed large neurons not discrete nuclei, curved laterally around the lower cerebral peduncles and then medially, emerging into the subarachnoid space within heterotopic neuroglial tissue in which they were myelinated by oligodendrocytes, not Schwann cells. The basis pontis nuclei were hypoplastic and the inferior olivary and dentate nuclei were dysplastic. There was focal foliar dysplasia. In addition to posterior fossa abnormalities, there were multiple foci of cortical dysplasia, patchy abnormal myelination of the hemispheric white matter, and absence of the optic radiations and striae of Gennari.

There are few descriptions of the neuropathological abnormalities of JS, none from a child with this genetic background. Our case shares many features with published descriptions and clearly fits the syndrome but has additional abnormalities which may reflect the unusual genetics.

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139

BRAFV600E Mutation is Not Associated with Focal Cortical Dysplasia

Sonika Dahiya1, Devon Haydon1, Jeffrey Leonard1, David Gutmann1, Robert Schmidt1. 1Washington University School of Medicine

Introduction: BRAFv600e mutation is common in low-grade CNS tumors with nearly half of gangliogliomas (GG) harboring this point mutation. Nevertheless, although one-third of GGs may be associated with focal cortical dysplasia (FCD) and both of these are included in the mTORopathy spectrum, the role of this specific mutation in FCD has remained rather unexplored. Furthermore, while this mutation may be present in both the glial and neuronal components, it is more frequent in the latter. In the current study, we investigated the role of BRAFV600E mutation in FCD presenting either as isolated lesions and/or collision lesions which had concurrent GG.

Design: We performed immunohistochemistry for BRAFv600e on eleven cases of isolated FCD cases and five cases with collision lesions (FCD and ganglioglioma). Immunostaining was also done for phospho-S6 (p-S6; marker of mTOR activation).

Results: While all of FCD alone (0/11) cases were negative for this point mutation, two cases with collision lesions (2/5 cases) showed expression of the mutant protein limited to GG areas only. All the cases in our cohort, including both the components in collision lesions, were positive for p-S6 expression.

Conclusions: Expression of p-S6 in all our cases is consistent with the notion that both FCD and GG exhinit mTOR signaling activation. However, the lack of BRAFv600e may be a signature of neoplasia.

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140

Development of the Hippocampal Formation in Human: Part II- Dentate Gyrus

Sara Cipriani1, Catherine Verney1, Jeannette Nardelli1, Pierre Gressens1, Homa Adle-Biassette2. 1INSERM UMR 1141; 2Department of Pathology, Lariboisiere Hospital, APHP & INSERM UMR1141

This is the first detailed study describing the development of hippocampal formation in human embryos and fetuses from 8 weeks of gestation (GW) to midgestation. Multiple immunohistochemical labelings were performed to study the progenitors using Ki67 associated with nestin, SOX2, PAX6 and TBR2, and post mitotic neuronal markers using NeuN, CUX1, SATB2, CTIP2, TBR1, NeuroD1, doublecortin, CaBP, and Calretinin. Our results showed the presence of SOX2/Ki67 double-labeled cells in the ventricular zone of the hippocampal formation next to the hem from 9 GW. The secondary dentate matrix, a germinative matrix located adjacent to the VZ of the dentate neuroepithelium was present around 11 GW. It consisted of proliferative cells composed of Pax6+, Sox2+ and Tbr2+ progenitors and migratory cells. It extended tangentially towards the dentate gyrus following a subpial migration route and established the proliferative tertiary dentate matrix in the hilus. The dentate gyrus became progressively identifiable from 13 GW. Dentate granule cells were composed of two main postmitotic neurons, labelled by Cux1 and CTIP2 transcription factors markers. In conclusion, the histological stages of formation of the dentate gyrus in human are roughly similar to that described in rodents. The pattern of expression of the transcription factors is different from the pyramidal layer of the hippocampus and from the neocortex. Animal studies suggest that all granule cells appear to project to CA3, forming the mossy fibres. The present study suggests that the densely packed granule cells in human are composed of different neuronal subtypes during development. Neuronal subtypes and connections of the dentate gyrus have yet to be investigated.

The research leading to these results was performed in the frame of DEVELAGE project “Pathways common to brain development and ageing: defining strategies for preventive therapy and diagnostics” (HEALTH-F2-2011-278486) and has received funding from the European Community’s 7th Framework Programme (FP7/2007-2013).

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141

Aprosencephaly-Atelencephaly in a Neonate with Severe Facial features of Holoprosencephaly

Nitin Agarwal1, Ada Baisre1. 1Rutgers New Jersey Medical School

We report a case of the very rare atelencephaly/aprosencephaly spectrum, also termed aprosencephaly (XK) syndrome with severe craniofacial features of holoprosencephaly. The case is that of a baby girl born at 34 weeks of gestation to a 21-year-old mother without prenatal care and past medical history of a “previously abnormal fetus aborted during the first trimester”. On external examination, craniofacial abnormalities typically seen in holoprosencephaly were severe, including one fused anterior/posterior fontanelle, anopthalmic cyclopia, arhinia, and hypoplastic mouth. Internal examination of the cranial vault showed absence of the anterior cranial fossa, orbits and periorbital structures, with a hypoplastic middle cranial fossa with absent sella turcica and pituitary gland. The cerebral hemispheres were also absent and the cranium was filled with approximately 500 cc of translucent-yellow fluid. A poorly formed diencephalon and relatively preserved infratentorial structures and spinal cord were identified. The general autopsy also revealed hypoplasia of the adrenal glands and pulmonary hyaline membranes. There was a normal female karyotype [46XX].

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142

Shaken Baby – Review of Old Cases

Roland Auer1. 1Université de Montreal

Shaken baby syndrome continues to be diagnosed based on a triad of retinal hemorrhage, subdural hemorrhage and encephalopathy. As part of a move to close medicolegal cases that were still open, the author was asked to review a case from 1991 by the police and a defense attorney. Two questions were posed. The first was to determine whether the diagnosis could be sustained after consultative review. The second was whether the case would be interpreted differently now 23 years after the diagnosis of shaken baby had first been entertained based on subdural hemorrhage, retinal hemorrhage and encephalopathy. Clinical history revealed a 5 day duration of a low grade febrile illness with sudden deterioration, gasping for air and cardio respiratory arrest. Blood cultures were taken and antibiotics were administered in hospital but the child rapidly died. Review of the case revealed severe hemorrhagic bronchopneumonia in all 5 lobes of the lungs, with little aeration. Hemorrhage was also seen in the kidneys, adrenal glands, retina, leptomeninges of the optic nerve and subdural space. A severe global ischemic encephalopathy was found, with ischemic necrosis comprising acidophilic neurons in most brain regions examined. Because of the presence of the triad, the father was charged in the death of his 5-month-old daughter, as he was the last caregiver prior to bringing her to hospital. The science done over the past 20 years has undermined the biophysics of producing the triad by shaking. Retinal and subdural hemorrhage once thought to be pathognomonic of shaken baby is now known to have many causes such as infection and reperfusion hemorrhage after cardiac arrest, as in the present case. A review of 8 local cases shows that hemorrhages can occur from numerous causes other than trauma. These facts are important to avoid wrongful convictions of innocent parents.

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143

Toddler with Microcephaly and Basal Ganglia Calcifications:? Aicardi-Goutieres Syndrome

Veena Rajaram1, Esther Bit-Ivan2, Russell Fetzer3, Jason Wang1, Naseem Uddin1. 1UT Southwestern/ Children’s Medical Center, Dallas; 2Feinberg School of Medicine, Northwestern University; 3UT Southwestern Medical Center, Dallas, TX

Aicardi-Goutieres syndrome (AGS) is an inherited progressive encephalopathy characterized by basal ganglia calcifications with cerebrospinal fluid (CSF) lymphocytosis and increased interferon-α level relative to the serum levels. Clinically, the patients have microcephaly with developmental delay/regression, spasticity, dystonia, abnormal eye movements and epilepsy. Other clinical findings include feeding difficulties, recurrent mild fever, chilblains, and transient elevation in liver enzymes. Imaging and neuropathology studies report bilateral basal ganglia calcifications with calcifications in the white matter, thalami, dentate nuclei and small vessel calcifications. Mutations in TREX1, RNASEH2B, RNASEH2C, RNASEH2A and SAMHD1 genes are thought to account for over 90% of these cases. We report a case with clinical and neuropathological findings most consistent with AGS. The patient was a 20-month old boy with microcephaly, feeding difficulties and developmental lag noticed at 7 months of age. He had developmental delay and developed spastic quadriparesis with dystonia and alternating exotropia. Further neurological evaluation was pending. He had recurrent fever with bronchiolitis. He presented with a few days of fever, severe respiratory distress and unresponsiveness. CT scan at this time showed marked hydrocephalus with symmetric dystrophic calcifications in the basal ganglia, cerebral periventricular white matter and deep cerebellar nuclei. He had mildly elevated liver enzymes. Investigations for TORCH and other infections were negative. He continued to deteriorate and care was withdrawn. At autopsy, grossly, he had microcephaly with dilated lateral ventricles, atrophy and yellow discoloration of the basal ganglia. Histologically, there was neuronal loss with gliosis in the basal ganglia and thalami, white matter loss with gliosis in the cortex and cerebellum and diffuse parenchymal and small vessel calcifications. Though a CSF analysis was not performed, in the absence of an infectious etiology, the clinical and histological findings are most consistent with AGS. Genetic testing for the above genes is in process.

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144

Mesial Temporal Sclerosis with Dysplastic Dentate Fascia Neurons: A Case Report.

Matthew Wood1, Arie Perry2. 1University of California, San Francisco; 2University of California, San Francisco, Division of Neuropathology

Background: Hippocampal sclerosis (HS) is the most common finding in surgical specimens for medically refractory epilepsy. In some cases, a second seizure-associated lesion is identified (dual pathology). HS associated with focal cortical dysplasia (FCD IIIa) is one example, although in such cases dysplasia is typically restricted to neocortex. Rare cases of HS have been reported showing dentate fascia granular cell dispersion combined with CD34-positive balloon cells, with or without associated neocortical FCD. Here, we describe an additional example of dual HS/FCD with CD34-positive balloon cells and dysplastic ganglion cells in the dentate fascia.

Case Report: A 33 year-old right-handed woman presented with medically refractory seizures. MR imaging showed abnormal left hippocampal T2 hyperintensity, architectural distortion, and atrophy. Electroencephalographic monitoring during two seizures indicated a left hemispheric origin. The patient underwent surgical resection of the left hippocampus, temporal lobe tip, and amygdala. Sections showed marked neuronal loss and gliosis involving the CA1 and CA4 regions, along with granule cell dispersion, consistent with HS. Dysmorphic ganglion cells and balloon cells were noted within and immediately surrounding the dentate fascia. A subset of dysmorphic neurons were neurofilament-positive, and a subset of the balloon cells expressed GFAP. In addition, focal CD34-positive balloon cells with atypical arborizing processes were identified. The left temporal tip resection showed focally jumbled neurofilament-positive neurons with slightly enlarged cell bodies, but no CD34-positive population (FCD IIa).

Summary: We report a rare case of epilepsy-associated HS combined with neocortical FCD IIa, with dysmorphic ganglion-like cells and CD34-positive balloon cells in the dentate fascia. Whether this reflects a purely developmental process or involves secondary changes associated with HS remains unclear.

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145

Glycogen Storage Disease Type III: Neuropathologic Phenotype Associated with Mutations in the AGL Gene

Kathy Newell1, Robert Reynders2, Jill Murrell3. 1KU School of Medicine, Dept of Pathology and Laboratory Medicine; 2KU School of Medicine, Department of Neurology; 3IU School of Medicine, Department of Pathology and Laboratory Medicine

Glycogen storage disease type III (GSD III), caused by a deficiency of glycogen debranching enzyme, affects liver, skeletal muscle, and heart. Central nervous system (CNS) involvement has not been described to our knowledge. We present the neuropathologic findings in a 27 year-old woman diagnosed with GSD III at age 1-1/2 years during a metabolic disorder workup. AGL gene mutations were later identified. She developed end-stage cirrhosis, underwent orthotopic liver transplant at age 24, and died at age 27 with invasive aspergillosis and Pneumocystis pneumonia while on chronic immunosuppressive therapy. Neurologic examination in the last week of her life documented brisk biceps, brachioradialis, and patellar deep tendon reflexes. A full autopsy confirmed glycogen storage in the heart and skeletal muscle. Histological sections from the fixed brain were stained with hematoxylin and eosin with/without Luxol fast blue. Periodic acid Schiff (PAS) stains with/without diastase were performed on select sections. Multiple brain stem nuclei, including cranial nerve nuclei VI, VII, XII, dorsal motor nucleus of X, spinal trigeminal nucleus, and the lateral reticular nucleus, and spinal cord anterior horn cells contained numerous neurons with vacuolated, clear cytoplasm with fine eosinophilic deposits that were PAS-positive and diastase sensitive, compatible with glycogen. Ultrastructural evaluation of the deposits is underway. The cerebral cortex and deep nuclei did not show obvious neuronal storage. White matter myelin was preserved. Multifocal brain abscesses contained branching septate hyphae, confirmed as Aspergillus fumigatus from postmortem cultures. Frozen brain tissue was reserved for confirmation of the AGL gene mutation. Autopsy neuropathological evaluation remains an essential tool in the study and documentation of rare disorders, such as glycogen storage disease.

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146

Inadvertent Intrathecal Vincristine Administration with Widespread Axonal Injury Demonstrated by APP Immunohistochemistry

Jesse Kresak1, Marie Rivera-Zengotita1, Martha Burt1, Anthony Yachnis1. 1University of Florida

Vincristine is a Vinca alkaloid chemotherapeutic agent used to treat hematologic and some solid-organ neoplasms. As a microtubule inhibitor, an unfortunate side-effect of this drug is disruption of axonal transport. Thus, the drug is intended for intravenous use only. We report a 59-year old man with diffuse large B-cell lymphoma who inadvertently received intrathecal vincristine. The patient experienced progressive neurologic decline despite attempts at CSF lavage with fresh frozen plasma and died five days after the incident. Grossly, there was evidence of prior shunt placement, the brain was mildly swollen, and the white matter appeared congested. Histological study revealed pallor of the centrum semiovale, focal perivascular hemorrhages, and widespread axonal spheroids that were demonstrated by immunohistochemical study for amyloid precursor protein (APP). There was subependymal spongiosis and focal hydropic-like change of the ependyma. Proximal axon segments of anterior horn neurons showed many APP-reactive spheroids as they exited the spinal cord. Anterior spinal nerve roots also contained APP-positive swellings. No significant pathologic changes were identified in the gray matter. In particular, there was no Purkinje cell loss. No evidence of residual lymphoma was identified. This case illustrates diffuse axonal injury caused by toxic disruption of axonal transport and emphasizes the need for continued awareness of this preventable medical error.

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147

Leptomeningeal Transthyretin Amyloidosis: A Case Study

Jennifer Ziskin1, Anna Okumu2, Christopher Adams2, Edward Plowey1. 1Stanford University Medical Center, Department of Pathology; 2Stanford University Mass Spectrometry, Stanford, CA

Background: Transthyretin amyloidosis is usually a systemic disease, however, rare mutations are associated with leptomeningeal amyloidosis that may lead to dementia and ataxia. Case reports of leptomeningeal transthyretin amyloidosis associated with the Tyr69His variant of transthyretin (TTR) are rare and do not discuss the neuropathologic features of the disease. Characterization of the associated histopathologic features is important to elucidate the role of leptomeningeal amyloidosis in neurodegeneration and dementia.

Design: We report a case of leptomeningeal amyloidosis in a 72 year old man with no significant family history who suffered from dementia and ataxia. A complete autopsy, including brain examination, was performed. Mass spectroscopy (MS) was employed to analyze the leptomeningeal amyloid.

Results: Histologic sections revealed systemic amyloidosis affecting the heart, lungs and other visceral organs. Widespread vascular and extravascular amyloid was documented in the leptomeninges. The amyloid extended into superficial cortical penetrating vessels and was prominently deposited in subpial and subependymal distributions throughout the brain. Other conspicuous neuropathologic findings included neocortical molecular layer gliosis, gliosis of the alveus and fimbria and cerebellar cortical atrophy. In addition to immunoreactive hippocampal neurons, a phospho-tau immunostain (AT8) revealed neocortical threads. No significant amyloid-beta immunoreactivity was seen. TTR gene sequencing performed on the paraffin embedded tissue identified a point mutation in exon 3 encoding a Tyr69His mutation. MS performed on the amyloid in paraffin block cores demonstrated abundant TTR with an 8-fold higher concentration of variant Tyr69His TTR compared to wild-type TTR.

Conclusions: Mutant TTR is a rare cause of leptomeningeal amyloidosis. We demonstrate the utility of MS in identifying transthyretin variant Tyr69His in a patient with leptomeningeal amyloidosis. We report additional neuropathologic features including phospho-tau positive neocortical threads and injury to hippocampal outflow structures, factors which may contribute to the dementia phenotype.

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148

3R Predominant Tauopathy with Globular Glial Inclusions

Stewart Neill1, Jonathan Glass2, Allan Levey2, Monica Parker3, Deborah Cooper2, Marla Gearing1. 1Department of Pathology and Laboratory Medicine, Emory University; 2Department of Neurology, Emory University; 3Department of Medicine, Emory University

Background: Numerous sporadic and familial neurodegenerative disorders involving aggregations of the microtubule-associated protein tau have been reported over the past several decades. Most of these tauopathies have shown a predominance of 4-repeat (4R) tau isoforms, the major exception being the 3R predominant Pick disease. The globular glial tauopathies (GGT) are 4R tauopathies showing small round tau-positive inclusions in white matter glia. We report a case displaying similar inclusions with a predominance of 3R tau.

Case History: The patient was a Caucasian male who initially presented to our facility at age 82 with short-term memory loss that progressed to include behavioral disinhibition and agitation. No family history of dementia was noted. A clinical diagnosis of mixed Alzheimer’s and vascular dementia was made. The patient’s memory loss and behavioral problems progressed until his death at age 89.

Autopsy Findings: The brain showed marked cortical atrophy of the frontal, parietal, and temporal lobes; the hippocampus, amygdala, and entorhinal cortex were severely affected. Microscopic examination showed neuronal loss and gliosis in atrophic limbic areas; sections of atrophic cortex showed primarily superficial spongiosis. A lacunar infarct was noted along with moderate small-vessel thickening. Special and immunohistochemical stains documented findings sufficient for diagnoses of probable Alzheimer’s disease (CERAD criteria) and neocortical Lewy body disease. TDP-43 immunohistochemistry revealed deposition in numerous supratentorial locations. Tau immunostains revealed the most striking findings: neurofibrillary tangles in cortical, deep gray, brainstem, and spinal cord structures, along with prominent astrocytic and oligodendroglial inclusions in white matter. These inclusions were diffusely 3R tau-positive and 4R tau-negative by immunohistochemistry and displayed a distinct morphology with numerous small round inclusions clustered around glial nuclei. No Pick bodies were seen.

Discussion: The finding of widespread 3R globular inclusions in white matter glia appears novel. It may expand the range of 3R tauopathies, and GGTs in particular.

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149

Childhood Neurodegenerative Disease with Widespread Tauopathy

Dibson Gondim1, Jill Murrell1, Adrian Oblak1, Deborah Sokol1, Laurence Walsh1, Gregory Bosh1, Ruben Vidal1, Michel Goedert2, Bernardino Ghetti1, Jose Bonnin1. 1Indiana University School of Medicine; 2Medical Research Council Laboratory of Molecular Biology (UK)

Severe accumulation of pathologic tau protein in neurons throughout the cerebral cortex and subcortical structures occurs in several adult-onset sporadic and genetically determined neurodegenerative diseases. On the contrary tau is less known to accumulate in diseases of childhood. Tau deposits have been observed in a number of unrelated conditions, including Niemann-Pick disease type C, pantothenate-kinase-associated neurodegeneration, subacute sclerosing panencephalitis, postencephalitic parkinsonism, SLC9A6-related mental retardation, and chronic traumatic encephalopathy. We report the case of a 14-year-old male with a neurodegenerative disease characterized by severe tau deposition. The child had no significant developmental or neurological problems until age 2 when speech and cognition started to deteriorate and was diagnosed as having autistic spectrum disorder. Multiple MRIs did not reveal brain abnormalities until age 11, when generalized cortical atrophy, ventriculomegaly, thinning of the corpus callosum and atrophy of the brainstem were documented. Since then, he developed behavioral problems, gait abnormalities, dysphagia, constant drooling, mild protrusion of the tongue and slight ptosis. He also had some cogwheeling, marked rigidity in both lower extremities and tended to fall backward. Extensive work-up did not reveal any genetic and mitochondrial disorders, enzymatic abnormalities, or storage diseases. An EEG confirmed the presence of severe slowing in the background and bifrontal epileptiform discharges that became generalized at times. Later in his course, he developed severe spasticity and mutism. He died at age 14. At autopsy, the brain weighed 1040 grams. Cerebrum, cerebellum and brainstem were atrophic. By histology and immunohistochemistry, numerous neurofibrillary tangles, tau-positive neurons, astrocytic plaques and neuropil threads were seen throughout the brain. DNA was extracted from liver, postmortem. The MAPT gene was sequenced and no mutation was found. Further chromosomal and molecular genetic studies are in progress to better define the cause of this disorder and the pathogenesis of the severe tauopathy.

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150

Creutzfeldt-Jakob Disease: Review of the Past Twenty Three Year Experience

Francine Epperson1, Adrian Oblak1, Jill Murrell1, Rose Richardson1, Brenda Dupree1, Pedro Piccardo2, Pierluigi Gambetti3, Bernardino Ghetti1. 1IU School of Medicine, Dept of Pathology and Laboratory Medicine; 2Food and Drug Administration; 3Case Western Reserve University

The aim of this report is to review the work of the last 38 years devoted to identifying prion diseases (p.d.) in the neuropathology laboratory at Indiana University School of Medicine, Department of Pathology and Laboratory Medicine. This analysis might be useful for planning resources needed for the development of a prion-dedicated laboratory in an academic setting. We have summarized the experience related to Creutzfeldt-Jakob disease (CJD) between 1991-2014, dividing these 23 years in two periods, before (1991-1996) and after (1997-2014) the development of the National Prion Disease Pathology Surveillance Center (NPDPSC). Between 1991 and 2014, a total of 256 cases (243 autopsies and 13 brain biopsies) were obtained to rule out a diagnosis of CJD. There were 117 cases from Indiana, 129 from 22 other states and 10 from two foreign countries. Prionopathies were diagnosed in 195 cases, including 168 sCJD (155 autopsies, 13 biopsies), 24 fCJD, one GSS, one FFI, and one variably protease-sensitive prionopathy (VPSPr). Sixty one cases were negative for p.d., and included diagnoses of AD, DLB, FTD/MND, other neurodegenerative disease, and cancer. Prior to the development of the NPDPSC (1991-1996), 48 cases were studied (46 prionopathies and 2 other). Between 1997 and 2014, 208 cases were received, 64 were confirmed CJD from Indiana and 85 from other states. The cases from Indiana were received from the following counties: 9 each from Marion and Allen, 4 each from Delaware and Johnson, 2 each from nine counties, and 1 each from 20 counties. The recent discovery of VPSPr and its clinical similarity to other degenerative diseases highlights the urgency of recognizing that at least one laboratory for prion diseases should be available in every state to develop accurate epidemiological data related to prionopathies in the US.

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151

Gerstmann-Sträussler-Scheinker Disease PRNP A117V: Prion Protein Deposition in Neurosensory Retina

Kathy Newell1, Francine Epperson2, Masaki Takao2, Martin Farlow3, Frederick Unverzagt4, Bernardino Ghetti2. 1KU School of Medicine, Dept of Pathology and Laboratory Medicine; 2IU School of Medicine, Dept of Pathology and Laboratory Medicine; 3IU School of Medicine, Department of Neurology; 4IU School of Medicine, Department of Psychiatry

Gerstmann-Sträussler-Scheinker disease (GSS) is a neurodegenerative disease associated with mutations in the Prion Protein (PRNP) gene and characterized by deposition of prion protein (PrP) in the brain. In sporadic and variant Creutzfeldt-Jakob disease (sCJD, vCJD), deposition of PrP in the retina has been reported, but limited information is available about retinal involvement in hereditary prionopathies. We studied a 41-year-old woman carrying the A117V mutation in PRNP. The proband presented with amnesia and dysphasia, followed by mild fine tremor in the right hand. Eight months later, saccadic pursuit eye movements, mild lower extremity weakness, mild ataxia, and generalized hyperreflexia were noted. Neuropsychological testing revealed severe general cognitive dysfunction, and the mini-mental status exam score was 13/30. She died at age 42. The brain and eyes were obtained at autopsy. The brain was moderately atrophic, and mild spongiform change was observed in the cerebral cortex and basal ganglia. PrP deposits in the form of unicentric and multicentric plaques were widespread. The cytoarchitecture of the retina was well preserved, but PrP immunopositive deposits were numerous in the outer plexiform layer (OPL). The deposits were aligned in the retinal layer that corresponds to the synapses between axons of photoreceptor cells and processes of cells of the inner nuclear layer. No PrP-immunopositivity was seen in the corneoscleral coat, the uvea, or the optic nerve. A similar pattern of retinal involvement was observed in GSS associated with the F198S mutation. Deposition of PrP in the retina has also been reported in sCJD and vCJD in both the OPL and inner plexiform layer (IPL). In these cases, there was a florid deposition in the IPL, but this was absent in GSS patients. The clinical and biological significance of PrP deposition in the retina has not been fully investigated.

(Funding source: P30AG010133)

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152

Familial Creutzfeldt-Jakob Disease Associated with the PRNP E200K-129V Haplotype: Report of a New Kindred

Adrian Oblak1, Jill Murrell1, Rose Richardson1, Francine Epperson1, Bernardino Ghetti1, Wei Chen2, Daniel Bonnin1, Martin Farlow3, Pierluigi Gambetti2. 1Indiana University School of Medicine, Dept Path and Lab Med; 2Case Western Reserve University; 3Indiana University School of Medicine, Dept of Neurology

Familial prionopathies are neurodegenerative diseases caused by mutations of the Prion Protein (PRNP) gene. They include familial Creutzfeldt-Jakob disease (fCJD), Gerstmann-Sträussler-Scheinker disease and Fatal Familial Insomnia. Approximately 20 mutations have been reported to be associated with fCJD and they are transmitted in an autosomal dominant pattern. Clinical and neuropathologic features of fCJD are comparable to those of sporadic CJD. We report a family in which four members have the PRNP E200K-129V haplotype and died of the disease. All four subjects carried methionine in the normal PRNP allele. The oldest subject, a female, died at 85 years. Her mother, three of her sisters, and a brother were also reported to have died of the same disease. Three of her daughters died of fCJD at ages 56, 55 and 51 years of age. Neuropathologic examination revealed mild to moderate cerebral atrophy with preferential involvement of the frontal and temporal cortices and the hippocampus. There was extensive prion protein (PrP) immunopositivity with a synaptic pattern in the cerebral cortex, basal ganglia, amygdala, hippocampus and parahippocampal gyrus, cerebellum and midbrain. Intraneuronal PrP immunopositive inclusions were prominent in the cerebral cortex, basal ganglia, entorhinal cortex and dentate nucleus of the cerebellum. Neuronal loss, gliosis and spongiform changes were observed in the same regions. Alzheimer disease pathology was also present in two members of the family. The E200K-129M haplotype is the most common form of fCJD while there are only five reported cases of E200K-129V haplotype. The presence of intraneuronal PrP immunopositive inclusions appears to be a consistent finding in subjects with the E200K mutation; however, their presence needs to be further investigated in other prionopathies.

(Funding sources: P30AG010133; NIH P01 AG-14359, Charles S. Britton Fund and CDC UR8/CCU515004)

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153

Gerstmann-Sträussler-Scheinker Disease Associated with the P102L-129M Haplotype: Clinical and Pathologic Heterogeneity

Daniel Bonnin1, Jill Murrell1, Francine Epperson1, Matthew Frosch2, E Tessa Hedley-Whyte2, Bernardino Ghetti1. 1Indiana University School of Medicine - Department of Pathology; 2C.S. Kubik Laboratory for Neuropathology - Massachusetts General

Gerstmann-Sträussler-Scheinker disease (GSS) is an autosomal dominant prionopathy frequently presenting with cerebellar ataxia and subsequent cognitive decline. GSS associated with the P102L-129M haplotype in the Prion Protein (PRNP) gene has clinical and neuropathologic characteristics that may occur in various combinations and with differing degrees of severity. A 41-year-old female presented with a four to six week history of cognitive decline. The subject’s condition deteriorated rapidly with short-term memory deficit, difficulty processing information and dysphasia. She died five months after the onset of symptoms. An autopsy was carried out. The subject’s father had an initial presentation interpreted as amyotrophic lateral sclerosis and a clinical course of 2.5 years. He died at age 62. He was neuropathologically and genetically reported to have the P102L-129M haplotype of the PRNP gene associated with GSS. The subject and two of her three siblings carried the same haplotype. The subject’s brain revealed no significant cortical atrophy or white matter loss, weighing 1,385 grams. Histologically, there were marked spongiform changes in the frontal and temporal lobes, cingulate gyrus, basal ganglia and entorhinal cortex. Immunohistochemical stains for PrP revealed numerous unicentric and multicentric plaques in the cerebral cortex, deep grey matter nuclei, substantia innominata, hippocampus, entorhinal cortex, cerebellum, and midbrain. In the spinal cord, there was moderate loss of myelinated fibers in the posterior and lateral columns, as well as neuronal loss in the anterior horns. The spinal cord changes contrasted with those seen in the subject’s father, in which severe loss of anterior horn neurons and pyramidal tract degeneration was observed. In the two patients examined, the neuropathologic features in the brain were similar, while age of symptom onset, initial clinical manifestations and disease duration were significantly different.

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154

Neuronal Tau and Parenchymal PrP in Gerstmann-Sträussler-Scheinker Disease Associated with the PRNP Q217R Mutation

Adrian Oblak1, Jill Murrell1, Rose Richardson1, Francine Epperson1, Bernardino Ghetti1. 1Indiana University School of Medicine, Dept Path and Lab Med

Previous studies have shown that in Gerstmann-Sträussler-Scheinker disease (GSS) neurons may be severely affected by deposition of tau protein in many cortical and subcortical gray matter regions. Severe tau deposition has been shown in GSS associated with the F198S-129V haplotype and other prionopathies. We now report the results of neuropathologic studies in three individuals carrying the Q217R-129V haplotype and belonging to one family. A male, his son and daughter died at age 59, 50 and 64, respectively. Both siblings had been diagnosed as suffering from GSS. Histologic studies of the three brains were carried out using the following methods: hematoxylin and eosin with Luxol fast blue (H&E/LFB), Bodian, Woelke-Heidenhain and Thioflavin S. For immunohistochemistry, antibodies against tau, amyloid beta, α-synuclein, glial fibrillary acidic protein, and prion protein (PrP) were used. Microscopically, PrP immunostaining and Thioflavin S, revealed abundant, unicentric, multicentric and diffuse amyloid plaques in frontal, temporal, parietal and occipital cortical regions. Amyloid and diffuse PrP immunopositive plaques were also observed in basal ganglia, thalamus, hippocampus, brain stem, pons, medulla, cerebellum and dentate nucleus. Tau immunohistochemistry revealed neurofibrillary tangles, neuropil threads and neurites of the crown surrounding PrP immunopositive amyloid cores in cortex, hippocampus, striatum, thalamus, substantia nigra, and locus coeruleus. In the white matter, tau-immunoreactive axons were also present. Neuronal loss and gliosis were apparent in the regions affected by PrP and tau pathology. Neuronal loss was particularly severe in the substantia nigra. Lewy bodies and Lewy neurites were present in the substantia nigra of the father. The present study shows that there is a consistent coexistence of intraneuronal tau deposition in the presence of parenchymal PrP diffuse and amyloid plaques in Gerstmann-Sträussler-Scheinker disease associated with the Q217R mutation.

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155

The Presence of Total Tau in Peripheral Tissues of Alzheimer’s Disease

Brittany Dugger1, Charisse Whiteside1, Chera Maarouf1, Thomas Beach1, Travis Dunckley2, Bessie Meechoovet2, Alex Roher1. 1Banner Sun Health Research Institute; 2Tanslational Genomics Research Institute

Tau is one of the main protein aggregates within the brains of Alzheimer’s disease (AD) patients; however, it has not been extensively examined within human peripheral organs. The purpose of our study was to determine the levels of total Tau in areas amendable for biopsy as compared to the brain within AD cases. We examined human postmortem tissue from the abdominal skin, liver, scalp, sigmoid colon, and submandibular gland for total Tau in 18 AD cases and 2 non-demented control cases- NC. Tissues were subject to Western blots analyses and enzyme-linked immunosorbent assays. Frontal gray matter of NC and AD were used as a frame of reference for each respective group. The brain had the highest levels of total Tau, having an average of 4150ng/mg total protein for NC and 2482ng/mg total protein for AD, followed by the submandibular gland (136ng/mg NC, 117ng/mg AD), sigmoid colon (21ng/mg NC, 22ng/mg AD), liver (12ng/mg NC, 19ng/mg AD), scalp (19ng/mg NC, 14ng/mg AD), and abdominal skin (10ng/mg NC, 20ng/mg AD). These results demonstrate that Tau is present in measurable quantities within these peripheral tissues but not to the levels that are detected within the brain. Future studies investigating different phosphorylated species of Tau are planned.

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156

Neuropathologic Assessment of Alzheimer’s Disease Neuroimaging Initiative (ADNI) Participants

Nigel Cairns1, Richard Perrin1, Erin Householder1, Deborah Carter1, Benjamin Vincent1, John Morris1. 1Washington University School of Medicine

Introduction: The mission of the Alzheimer’s Disease Neuroimaging Initiative (ADNI) is to define the progression of Alzheimer’s Disease. A major goal of the ADNI study has been to collect and validate data such as MRI and PET images, cerebral spinal fluid, and blood biomarkers as predictors of the disease in autopsy-confirmed cases. The objectives of the ADNI Neuropathology Core are to facilitate autopsy consent, brain collection, and perform standardized neuropathologic assessments of all ADNI participants who come to autopsy at the 58 ADNI sites in the USA and Canada.

Methods: The ADNI-NPC maintains a central laboratory to provide uniform neuropathologic assessments using the operational criteria for the classification of AD and other pathologies defined by the National Alzheimer Coordinating Center (NACC). Results: Since the inception of the NPC in 2008 the autopsy rate is 36/54 (66.7%). AD was present in all cases and the second most frequent comorbidity was Lewy body disease. Additional pathologies included: hippocampal sclerosis, TDP-43 proteinopathy, argyrophilic grain disease, small vessel disease and infarcts. Multimodal correlations were observed in the clinical, biomarker, imaging, and neuropathologic data [1].

Conclusions: The Neuropathology Core has established procedures to coordinate collection of brains from all ADNI sites and undertake standardized neuropathologic assessments. The NPC will facilitate clinical and multimodal biomarker correlates of neuropathology in the ADNI cohort.

Reference: Toledo JB and Cairns NJ et al. Clinical and multimodal biomarker correlates of neuropathology. Acta Neuropathol. Commun. 2013;1:6.

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157

The Dominantly Inherited Alzheimer Network (DIAN): The Essential Role of the Neuropathology Core

Nigel Cairns1, Richard Perrin1, Erin Householder1, Deborah Carter1, Benjamin Vincent1, John Morris1. 1Washington University School of Medicine

Background: The Dominantly Inherited Alzheimer Network (DIAN) is a collaborative effort of international Alzheimer disease (AD) centers that are conducting a multifaceted prospective biomarker study in individuals at-risk for autosomal dominant AD (ADAD) [1]. Neuropathology is performed on participants who come to autopsy to determine the distribution and severity of lesions and the presence of co-morbidities which may influence clinical, cognitive, imaging and biochemical measures of disease progression.

Methods: The Neuropathology Core maintains a central laboratory to provide uniform neuropathologic assessments defined by the National Alzheimer Coordinating Center (NACC). The DIAN-NPC maintains a state-of-the-art brain bank of DIAN-derived brain tissue to promote biomarker and multi-disciplinary clinico-pathologic studies.

Results: Since the implementation of the DIAN Neuropathology Core (DIAN-NPC), there have been 10 deaths of DIAN participants or family members, 2 of whom have clinical, biomarker, and neuroimaging data. Of 10 autopsies, the neuropathologic diagnoses include: AD (100%) and dementia with Lewy bodies (40%). Although the overall numbers to date are small, these data demonstrate that the Neuropathology Core has established the administrative organization to harvest brains from DIAN participants who come to autopsy at the participating sites.

Conclusions: The Neuropathology Core has: (1) implemented a protocol to solicit permission for brain autopsy in DIAN participants at all 11 sites in the USA, Europe, and Australia, who die; (2) established procedures to send brain tissue to the Neuropathology Core for a standardized and uniform neuropathologic assessment; and (3) determined the presence of co-existent pathologies with AD in the autopsied cases. It is possible that these co-morbidities may contribute to the variance in DIAN data.

Reference: Bateman RJ, et al. Dominantly Inherited Alzheimer Network. Clinical and biomarker changes in dominantly inherited Alzheimer’s disease. New England Journal of Medicine 2012; 367: 795-804.

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158

Alzheimer Disease and Diffuse Lewy Body Disease Coexist in a PSEN1 N135S Mutation Carrier

Bernardino Ghetti1, Jill Murrell1, Martin Farlow2, Frederick Unverzagt3, Adrian Oblak1, Francine Epperson1, Rose Richardson1, Shannon Risacher4, Andrew Saykin4, John Morris5, Nigel Cairns5. 1Indiana University School of Medicine, Dept Pathology and Lab Medicine; 2Indiana University School of Medicine, Dept of Neurology; 3Indiana University School of Medicine, Dept of Psychiatry; 4Indiana University School of Medicine, Dept of Radiology; 5Washington University in St. Louis School of Medicine

The neuropathologic phenotype of Alzheimer disease (AD) associated with PSEN1 mutations is not uniform. To illustrate this characteristic, we describe the case of a male, who showed gait difficulties at age 39. When examined at age 41, he was repetitive and had word finding difficulties. Subsequently, he developed generalized seizures and a rapid decline, requiring full-time nursing care. On examination at 42, his movements were apraxic, but tremor was absent. Spasticity predominantly involved the lower extremities. His speech was strained and spastic; he had bilateral Babinski signs. His Clinical Dementia Rating was 3.0, consistent with severe dementia. At baseline and follow-up, [18F]FDG and [11C]PiB PET showed generalized hypometabolism and a positive amyloid signal, most evident in the striatum. He died at age 43. At autopsy, the brain weighed 1530 grams. For histology, hematoxylin and eosin with Luxol fast blue and Thioflavin S were used. For immunohistochemistry, antibodies against tau, amyloid beta (Aβ), glial fibrillary acidic protein, and α-synuclein were used. Aβ cored plaques and diffuse deposits were seen in the neocortex and hippocampus, while diffuse Aβ deposits were predominant in the striatum and cerebellum. Aβ angiopathy was severe. Tau-immunopositive neurons and neuropil threads were numerous in the cerebral cortex, dorsal pontine gray and raphe. α-synuclein immunopositive Lewy bodies and neurites were abundant in frontal and cingulate cortices and substantia nigra. DNA was extracted from brain tissue; the Presenilin 1 (PSEN1) gene was sequenced, revealing an A to G point mutation predicting an amino acid substitution of asparagine (N) to serine (S) at residue 135. The neuropathologic phenotype associated with the PSEN1 N135S mutation has been reported previously; however the present case is noteworthy for the coexistence of Diffuse Lewy Body Disease with severe AD.

(Funding sources: P30AG010133, U19AG032438)

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159

Familial Dementia Associated with the Novel PSEN1 F176V Mutation

Bernardino Ghetti1, Adrian Oblak1, Rose Richardson1, Francine Epperson1, Jill Murrell1. 1IU School of Medicine, Dept Pathology and Laboratory Medicine

The Presenilin 1 (PSEN1) gene is known to have over 180 mutations. We report a case in which a novel mutation has been identified. A history of presenile dementia was traced back three generations preceding that of the proband. The subject, a female, showed symptoms of memory impairment at the age of 59 years. Following a neurological examination, it was concluded that she was in the early stages of Alzheimer disease. Her short term memory progressively deteriorated in the ensuing years. At age 69, her mini mental status exam score was 20. At age 70, a computed tomography scan revealed mild diffuse cerebral atrophy and at age 71, a positron emission tomography scan with flurodeoxyglucose revealed hypometabolism in the posterior parietal region of the left cerebral hemisphere. Her cognitive status continued to deteriorate and she died at age 77. At autopsy, the brain weighed 838 grams. DNA was extracted from brain tissue and the PSEN1 gene was sequenced. The molecular genetic analysis revealed a TTT>GTT point mutation predicting an amino acid substitution of phenylalanine (F) to valine (V) in the PSEN1 gene at residue 176. The brain was severely atrophic. There was moderate to severe atrophy of the head of the caudate nucleus and severe atrophy of the hippocampus and parahippocampal cortex. For neuropathology, histological methods included hematoxylin and eosin with Luxol fast blue, Bielschowsky and Thioflavin S. For immunohistochemistry, antibodies against tau, amyloid beta (Aβ), and α-synuclein were used. Numerous β-amyloid immunopositive plaques and numerous tau immunopositive neurons and neuropil threads were seen throughout the cerebral cortex. The hemispheric white matter showed marked loss of myelinated fibers. Amyloid angiopathy and parenchymal Aβ deposition were severe in the cerebellum. Additional studies on this kindred are needed to clarify the clinical and pathologic phenotypes associated with the novel PSEN1 F176V mutation.

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160

Neuropathologic Phenotype of Dementia Associated with PSEN1 H163R Mutation

Bernardino Ghetti1, Jill Murrell1, Adrian Oblak1, Rose Richardson1, Jordan Grafman1, Alan Lerner3. 1IU School of Medicine, Dept Pathology and Laboratory Medicine; 2Rehabilitation Institute of Chicago; 3Case Western Reserve University

Mutations in Presenilin 1 (PSEN1) gene are numerous; however not for all of them has the associated neuropathology been investigated. We describe the neuropathologic phenotype in an individual affected by an early-onset. A 45-year-old man began experiencing depression and symptoms of cognitive impairment, following the death of a brother. At the age of 48 years, he was diagnosed as having frontotemporal dementia (FTD). At age 52, a neurological exam revealed severe aphasia. A positron emission tomography scan with flurodeoxyglucose showed generalized hypometabolism in the cerebral hemispheres with greater decrease in the left frontal, temporal and parietal lobes. A computed tomography revealed a volume loss greater in the left temporal lobe. At age 53, he was hospitalized and discharged with a diagnosis of FTD, associated with behavioral disturbance and agitation. The patient died at age 55. At autopsy, the brain weight was 1130 g. For histology, hematoxylin and eosin with Luxol fast blue, Bielschowsky and Thioflavin S were used. For immunohistochemistry, antibodies against tau, amyloid beta (Aβ), glial fibrillary acidic protein, and α-synuclein were used. Numerous Aβ immunopositive plaques, tau-immunopositive neurons, neuropil threads, and neurites surrounding plaque cores were seen in the cerebral cortex. Amyloid angiopathy and diffuse parenchymal Aβ deposits were severe in the cerebellum. Tau immunopositive neurons were numerous in the substantia nigra, in the dorsal pontine gray and in the raphe. α-synuclein immunopositive Lewy bodies and neurites were numerous in the amygdala, the adjacent temporal cortex and the basal forebrain. In these areas, the neuropil showed spongiform-like changes. DNA was extracted from brain tissue and PSEN1 was sequenced revealing a CAT>CGT point mutation predicting an amino acid substitution of histidine (H) to arginine (R) at residue 163. The PSEN1 H163R mutation has been previously reported; however the neuropathologic phenotype associated with this mutation had not been documented.

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161

Oxidative Damage is Correlated with Mitochondrial Abnormalities in Aging but not Alzheimer Disease

George Perry1, Rasha Shammas1, Xiongwei Zhu2, Xinglong Wang2, Hyoung-gon Lee2, Rudy Castellani3, Akihiko Nunomura4. 1The University of Texas at San Antonio, San Antonio, Texas; 2Case Western Reserve University, Cleveland, Ohio; 3University of Maryland, Baltimore, MD, USA; 4University of Yamanshi, Yamanshi, Japan

Alzheimer disease (AD) and aging are marked by oxidative damage and mitochondrial abnormalities. Since mitochondria can play a critical role in oxidative damage, we conducted this study to identify the relationship of oxidized RNA, 8-hydroxyguanosine (8OHG), and mitochondrial DNA (mtDNA) accumulation in AD and aging individuals. Abnormalities were examined by using densitometry of hippocampal pyramidal neurons: mtDNA accumulation as a marker of mitophagy and oxidative damage by 8OHG. Among aging individuals, oxidative damage and mtDNA were highly correlated (correlation coefficient = 0.86). While both 8OHG and mtDNA were at higher levels in AD individuals, they were uncorrelated (correlation coefficient = 0.06). In contrast, as we found before, oxidative damage was inversely correlated with amyloid-β; it was unrelated in normal aging individuals. These results suggest that oxidative damage is directly related to mitophagy in aging individuals. With the onset of AD, amyloid-β plays a strong antioxidant role. These findings indicate that the onset of AD is marked by a pleotrophic change in oxidative stress, one characterized by a change from mitochondria to amyloid-β dependency.

George Perry is supported by the Semmes Foundation, and by a grant from the National Institute on Minority Health and Health Disparities (G12MD007591) from the National Institutes of Health.

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162

Screening for Pathology Consistent with Chronic Traumatic Encephalopathy in Neurodegenerative Diseases

Kevin Bieniek1, Ann McKee2, Dennis Dickson3. 1Mayo Graduate School, Department of Neuroscience; 2Center for the Study of Traumatic Encephalopathy, Boston University; 3Department of Neuroscience, Mayo Clinic Florida

Chronic traumatic encephalopathy (CTE) is a progressive neurodegenerative disease linked to subconcussive repetitive traumatic brain injury. Neuropathologically, CTE is defined by deposition of hyperphosphorylated tau in neuritic threads and neurofibrillary tangles, with variable axonal pathology and transactive response DNA-binding protein 43 (TDP-43) pathology. In its earliest stages, CTE pathology is focal in convexity frontal lobe, at the depths of sulci and perivascular. CTE can occur in other neurodegenerative diseases. While CTE often lacks amyloid-β and α-synuclein pathology, Alzheimer’s disease (AD) and Lewy body disease (LBD) pathology have been reported with CTE pathology. Upper and lower motor neuron disease, amyotrophic lateral sclerosis (ALS), has also been reported in CTE. While current studies focus on further characterizing this disease in cohorts of athletes and military veterans, there are no studies that have screened for CTE pathology in neurodegenerative diseases. The Mayo Clinic Jacksonville Brain Bank currently holds over 6,500 brains from individuals with memory and/or motor disorders. We identified 350 men that were at risk for CTE (based upon gender, age, brain weight, and no primary tauopathy) and retrospectively reviewed available medical records for history of involvement in contact sports or documented head trauma. We screened these cases with tau immunohistochemistry and found 10 (3%) with pathologic features compatible with CTE. All cases had primary diagnoses of AD, LBD, ALS or frontotemporal lobar degeneration. CTE-related tau pathology was not detected in 56 disease and aged-matched women. We also screened 68 individuals with ALS, and found 5 additional men with pathology consistent with CTE. This study demonstrates the importance of screening for CTE in individuals with history of involvement in contact sports or documented head trauma even though they have other concomitant neurodegenerative pathologies.

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163

“APP and Aβ-immunoreactive Plaques” in Traumatic Brain Injury: An Underappreciated Neuropathological Finding?

Ross Reichard1, Cristiane Ida1. 1Mayo Clinic

Amyloid precursor protein (APP) is an integral membrane protein highly expressed in neuronal synapses. Although its function is not fully understood, it has been implicated as a regulator of synapse formation, neural plasticity and iron export. APP proteolysis generates beta amyloid (Aβ), including amyloid fibrillar isoforms (Aβ40 and Aβ42), which form the amyloid plaques seen in neurodegenerative diseases, especially in Alzheimer disease. Studies have demonstrated that, in addition to the characteristic diffuse traumatic axonal injury (dTAI) APP-immunoreactive axonal spheroids/bulbs, Aβ deposition may be seen within short time (hours) of the traumatic event . We report two insightful autopsy cases that illustrate “APP and Aβ -immunoreactive plaques” in a setting of fatal traumatic brain injury. Both individuals did not have significant past neurological history and were involved in road traffic accident: 1) 50-year-old woman who sustained multiple skull and cervical fractures and blunt force trauma of the head with diffuse subarachnoid hemorrhage (SAH) with intraventricular extension, and multiple cerebral acute contusions, and survived for 3 days; histologically, in addition to dTAI associated with global hypoxic-ischemic brain injury, numerous cortical tau-negative, variably Aβ-immunoreactive “APP-immunoreactive plaques” were identified; 2) 53 year-old man who suffered skull fractures and blunt force trauma to the head, with diffuse SAH, extensive cerebral contusions and survived for 6 hours; histologically, besides dTAI and Alzheimer disease neuropathologic changes distinct cortical tau-negative, variably Aβ-immunoreactive “APP-immunoreactive plaques” were present. These cases illustrate a plaque morphologically “invisible” on H&E stains, highlighted by immunostaining for APP, which may be underappreciated in severe traumatic brain injury. These findings also support the view that induction of APP is a response to neuronal stresses that, although possibly protective, may progress to a neurodegenerative disease process in susceptible individuals, and suggesting a pathophysiological association between Alzheimer disease and traumatic brain injury.

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164

Myopericytoma Involving The Orbit In A 78-year-old Female

Jenny Smith1, Jeremy Deisch1. 1Loma Linda University

Myopericytomas are uncommon tumors of presumed perivascular myoid origin. According to the World Health Organization (WHO), myopericytomas occur as part of a spectrum of tumors arising from perivascular contractile cells, including myofibroma/myofibromatosis, glomus tumors, and “true” hemangiopericytomas. Myopericytomas typically affect adults, but examples in pediatric and elderly patients have been reported. The subcutaneous tissues of distal extremities are most often affected, but nearly all superficial anatomic sites may be affected. In the central nervous system, one presumed cerebral hemispheric parenchymal tumor and one thoracic epidural mass have been reported. Otherwise, there are no reports of myopericytomas involving the nervous system. We report a case of a myopericytoma involving the orbit in a 78-year-old woman with a three-year-history of a slowly progressive right eye pain, bulging, and vision loss. An MRI demonstrated a 5.1 cm well circumscribed heterogeneously enhancing mass in the right orbit, encasing the optic nerve. Biopsy revealed a richly vascular mesenchymal neoplasm with numerous thin-walled, variably sized sinusoidal vessels and a bland proliferation of spindled monotonous cells with scant to moderate amounts of eosinophilic cytoplasm. Significant cytologic atypia was lacking, and mitotic figures were not increased. By immunohistochemistry, the tumor cells were strongly and diffusely positive for vimentin and patchy positive for smooth muscle actin (SMA). CD31/CD34 immunostains highlighted the lesional blood vessels. S-100, cytokeratin, EMA, neurofilament, and GFAP immunostains were negative. The tumor was treated with radiotherapy, resulting in decreased enhancement and a decrease in tumor size by 1.5 cm during ten months of follow-up. This case demonstrates the first reported example of a myopericytoma involving the orbit, expanding the already wide differential diagnosis of orbital region tumors. Neuropathologists should be aware of this entity, as orbital region tumors often fall under the purview of neuropathologists.

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165

An Unusual Presentation of Gliosarcoma: A Case of Primary Tumor Arising in the Optic Nerve.

Yevgeniy Sychev1, Luis Gonzalez-Cuyar1, Raghu Mudumbai1, C. Dirk Keene1. 1University of Washington

Gliosarcoma is a subtype accounting for a small percentage of glioblastoma multiforme, a highly malignant central nervous system tumor. While glioblastoma multiforme is the most common primary malignant tumor in the brain, it is encountered only rarely in the optic nerve. Gliosarcomas commonly arise in the temporal lobes and until now there have been no reports of this entity as a primary optic nerve tumor. Here we report a case of an optic nerve gliosarcoma. A 90 year old demented woman presented with one year history of progressive proptosis, neovascular glaucoma, blindness and ocular pain. She underwent enucleation following diagnosis of optic nerve tumor. Microscopically the tumor demonstrated effacement of the normal nerve, invasion of the retina and a biphasic architecture demonstrating foci of pleomorphic epithelioid and spindle cells. Electron microscopy demonstrated abundant intermediate filaments but no myofibers, sarcomeric differentiation, melanosomes or pre-melanosomes. The neoplasm was negative for melan-A, HMB45, tyrosinase, synaptophysin, alpha smooth muscle actin, epithelial membrane antigen. AE1/AE2 pan-cytokeratin cocktail showed focal intermediate positivity. S-100 and vimentin immunostaining was strongly positive in both spindle and glial populations, while GFAP immunostaining was primarily limited to glial portions. Multiple foci of mGFAP negative cells stained strongly positively for reticulin. These features are characteristic of gliosarcomas encountered in the brain.

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166

The Prognostic Value of EGFR Amplification in MGMT Methylated Glioblastomas

Seema Shroff1, Katharine McNeill2, Irina Mikolaenko1, David Zagzag1, Matija Snuderl1. 1NYU Langone Medical Center; 2Laura and Isaac Perlmutter Cancer Center, NYU

Background: Glioblastoma (GBM) is the most common malignant brain tumor of adults with a 5-year survival <5% from diagnosis. MGMT gene promoter methylation is the most frequently performed molecular test in neuropathology clinical practice. The use of EGFR gene amplification as a prognostic maker is not well established. We investigated the prognostic role of EGFR amplification in subsets of patients with glioblastoma. We specifically sought if there was additional prognostic value of EGFR status in patients with MGMT methylated GBM.

Methods: We performed a retrospective analysis of 87 adult patients with GBM. We analyzed MGMT, EGFR, IDH1 R132H, p53 and BRAF V600E status and correlated with clinical data.

Results: Our cohort consisted of 77 patients (51 males and 26 females) with median age of 63 (8-88). Complete clinical, pathologic and molecular data were available for 20 patients. Median survival for the cohort was 4 months (range 1-24). MGMT was methylated in 9/30 patients (30%) with median progression free survival (PFS) of 8 months (range 4-24), compared to MGMT non-methylated patients (70%)(median 4.5, range 1-12, p-value = 0.03). EGFR was amplified in 10/23 (43%) patients with median PFS of 4.5 months (range 3-24). EGFR non-amplified patients (57%) showed PFS of 4 months (range 1-10). PFS in patients with MGMT methylation who had EGFR amplification was 6.75 months (range 4-24) in comparison with EGFR non-amplified, PFS = 4 months (range 4, p-value = N.S.). PFS in patients with no MGMT methylation who had EGFR amplification was 4.5 months (range = 3-7) compared to EGFR non-amplified tumors (PFS= 3.25 months, range 1-10).

Conclusions: MGMT methylation status is a superior predictive marker in GBM. EGFR amplification in MGMT methylated GBMs does not have additional prognostic value.

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167

Improved Adenoviral Transduction of Glioblastoma Cells by Aptamer Modification

Qinwen Mao1, Hao Chen2, Esther Bit-Ivan1, Eileen Bigio1, Haibin Xia2. 1Northwestern University Feinberg School of Medicine; 2Shaanxi Normal University, China

Adenovirus type 5 (Ad5) is one of the most commonly used vector systems for gene and viral therapy for glioblastoma. However, targeting of adenovirus to human glioblastoma remains a challenge due to the low expression level of CAR (coxsackievirus and adenovirus receptor), the known adenovirus receptor, in glioma cells. To improve the targeting efficiency of Ad5 to glioblastoma, transductional retargeting strategies, including genetic capsid modification and adaptor-based strategy, have been used. The latter involves the use of a molecular bridge to retarget the Ad from its native primary receptor to a different cell surface receptor. Aptamers are single-stranded oligonucleotides that bind at high affinity to a target molecule, and are good candidates for targeted cancer therapy. In this study, to construct an aptamer-modified Ad5, we first genetically modified the hypervariable region 5 (HVR5) of Ad hexon with biotin acceptor peptide (BAP), which would be metabolically biotinylated during virus production in HEK293 cells. Then the biotin labeled aptamer was attached to the modified Ad through avidin-biotin binding. The aptamers used in this study include AS1411 and GBI-10. The former is a DNA aptamer that targets nucleolin, a nuclear matrix protein found on the surface of cancer cells. The latter is a DNA aptamer that can recognize the extracellular matrix protein tenascin-C on the surface of human glioblastoma cells. To examine if aptamer-modification of the hexon protein could improve the adenoviral transduction efficiency in gliomas, glioblastoma U251 cells were transduced with aptamer-modified Ads. Our results show that the transduction efficiency of AS1411- or GBI-10-modified Ad in U251 cells is approximately 4.1-fold or 5.2-fold higher than that of the control vectors. The data indicate that aptamer modified adenovirus would be a useful tool for glioblastoma gene and viral therapy.

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168

Overexpression of Eg5 Correlates with High Grade Astrocytic Neoplasm

Liqiong Liu1, Xichun Liu2, Marcus Mare2, Aaron Dumont2, Haitao Zhang2, Dong Yang2, Zhenggang Xiong2. 1Louisiana State University; 2Tulane University

Objective: To investigate the relationship between Eg5 expression and histopathological grade of astrocytoma for potential application of Eg5 as a diagnostic marker and a therapeutic target of astrocytomas.

Methods: Eg5 expression was evaluated by immunohistochemical examination on 92 specimens including 25 cases of glioblastoma (WHO grade IV), 22 cases of anaplastic astrocytoma (WHO grade III), 20 cases of diffuse astrocytoma (WHO grade II), and 21 cases of pilocytic astrocytoma (WHO grade I). The histopathological characteristics and Eg5 expression level of each tumor were assessed under microscope and by image analysis computer software. Correlation between Eg5 expression and histopathological grade of astrocytoma was statistically analyzed.

Results: Astrocytic tumors exhibited significant correlation of expression of Eg5 with higher WHO histopathological grades (p<0.001). Eg5 is expressed in 51%-98% (mean: 76.88%) of neoplastic cells in glioblastoma, 34%-57% (mean: 43.59%) of neoplastic cells in anaplastic astrocytoma, 6%-36% (mean: 18.60%) of neoplastic cells in diffuse astrocytoma, and 2%-28% (mean: 13.48%) of neoplastic cells in pilocytic astrocytoma.

Conclusions: Overexpression of Eg5 associates with high-grade astrocytic neoplasm, and it may represent an independent diagnostic and prognostic factor in grading astrocytic tumors and predicting prognosis of astrocytic tumor patients.

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169

Detection of CD133 Expression in U87 Glioblastoma Cell Line Using Novel Anti-CD133 Monoclonal Antibodies

Esther Bit-Ivan1, Haibin Xia2, Eileen Bigio1, Qinwen Mao1. 1Northwestern University Feinberg School of Medicine; 2Shaanxi Normal University, China

In glioblastoma, CD133 identifies a subpopulation of stem-like tumor cells, named CSCs, which are able to initiate tumor growth and are highly resistant to conventional chemo-radiotherapy. However, some studies show that some CD133-negative glioma cells are also able to self-renew and retain tumorigenic potential. It remains obscure whether CD133 is a reliable CSC marker in gliomas and whether a population of cancer stem cells that do not express CD133 at all even exists. These conflicts might be due to the detection limit of currently available anti-CD133 antibodies. In this study, we generated novel anti-human CD133 monoclonal antibodies (McAb) using two recombinant extracellular domains of human CD133, CD133 ectodomain 1 (amino acid 171 to 420) and CD133 ectodomain 2 (amino acid 507-716). Immunofluorescence microscopy and Western blot analysis revealed that at least two antibodies (C1E3 and C1E4) were directed against CD133 ectodomain 1, and four of them (including C2E1) were directed against ectodomain 2. Similar to the commercially available anti-CD133 antibodies, C1E3 only detected glycosylated full length CD133 band in Caco-2 cells (a human colorectal adenocarcinoma line) but failed to detect any CD133 expression in the human glioblastoma cell line, U87. Interestingly, C2E1 McAb, which revealed both glycosylated or non-glycosylated full length CD133 bands in Caco-2 cells, detected high levels of non-glycosylated full length CD133 protein in U87 cells, which previous studies had shown were devoid of CD133 expression. In addition, CD133 overexpression led to aberrant glycosylation of CD133 in U87 cells. Moreover, both C1E3 and C2E1 antibodies suppressed cell proliferation when incubated with Caco-2, or U87 cells overexpressing CD133. These novel antibodies might be of significant value for further exploration of the expression and function of CD133 in cancer stem/progenitor cells, and their usefulness in cancer therapeutics deserves further exploration.

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170

Recurrent Glioblastoma Is Associated With Increased Expression of Mesenchymal Markers

Matthew Wood1, Gerald Reis2, Joanna Phillips2. 1University of California, San Francisco, Department of Pathology; 2Department of Pathology, Division of Neuropathology

Background: Glioblastoma (GBM) is a molecularly heterogeneous disease with a uniformly poor prognosis due to invasiveness and resistance to therapy. A number of studies have focused on the heterogeneity of GBM at initial presentation, but few have characterized whether heterogeneity is retained at recurrence. To begin to address this question, we generated a tissue microarray (TMA) for analysis of paired primary and recurrent GBM.

Design: We identified recurrent GBM cases diagnosed between 1996 and 2013, and reviewed clinical data to obtain population characteristics and therapeutic histories. Up to 3 tissue cores from primary and recurrent tumors were assembled in tissue microarrays. Arrays were immunostained for a number of markers including Ki67, CD34, phosphorylated S6 kinase (pS6K), YKL40, phosphorylated STAT3 (pSTAT3), CD44, OLIG2, EGFR, and IDH1R132H.

Results: The study population included 14 male and 10 female patients (N=24), with mean age at diagnosis of 53 years (range 31 to 73). The mean time to recurrence was 531 days (range 75 to 2411). As expected, primary tumors exhibited a diversity of phenotypes, including mutant IDH1 (15%; 3/20), robust EGFR expression (45%; 9/20), a range of Ki67 positive cells (2-60%), and a mesenchymal phenotype (34%; 6/18), denoted by robust expression of at least two mesenchymal markers (pSTAT3, YKL40, or CD44). Fifteen primary and recurrent GBM cases had adequate tissue for analysis of paired samples. At recurrence there was an increase in tumors with a mesenchymal phenotype (27% vs. 80%), including one IDH1 mutant tumor. A single IDH1 mutant tumor showed reduced mesenchymal marker expression at recurrence.

Conclusions: We generated a tissue microarray of paired primary and recurrent GBM for analysis of molecular heterogeneity and mesenchymal phenotype characteristics. At recurrence a significant subset of tumors acquired expression of mesenchymal markers. Studies to determine whether this reflects a change in overall tumor heterogeneity are ongoing.

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171

Genomic Sequencing Reveals PI3KCA Mutations in Two Cases of Glioblastoma with Sarcomatous Differentiation

Lyndsey Emery1, Elizabeth Azzato1, Robert Daber1, Maria Martinez-Lage1. 1Hospital of the University of Pennsylvania

Background: Gliosarcomas are a rare subset of glioblastoma (GBM) characterized by a biphasic appearance with mesenchymal differentiation, accounting for approximately 1-5% of GBM diagnoses1. Phosphoinositide 3-kinase, catalytic, alpha polypeptide (PIK3CA) is a member of the PI3K family of enzymes, which function in numerous roles in the PI3K/AKT/mTOR signaling pathway, including cell growth, proliferation, differentiation, intracellular trafficking and survival. In turn, these enzymes are commonly identified as mutated in numerous cancers. PIK3CA mutations are present in approximately 11% of glioblastomas2. Here we present two cases of WHO grade IV malignant gliomas, a gliosarcoma and a glioblastoma with prominent sarcomatous features, with PIK3CA mutations identified by genomic sequencing.

Cases: Case 1 is from a 71-year-old male who underwent tumor resection with pathology demonstrating gliosarcoma, WHO grade IV. Next-generation sequencing using a panel of 47 genes identified a missense mutation in TP53 (p.R273P, c.818G>C) and an additional missense mutation in PIK3CA (p.H1047R, c.3140A>G). Case 2 is from an 81-year-old female who underwent tumor resection with pathology demonstrating glioblastoma, WHO grade IV, with focal sarcomatous features. Interval imaging showed tumor progression and she underwent a second resection, which demonstrated recurrent/residual glioblastoma. This specimen showed more prominent sarcomatous differentiation. Her original tumor specimen was submitted for genomic sequencing, which identified amplification of EGFR and an additional missense mutation PIK3CA (p.G1049S, c.3145G>A).

Conclusions: To our knowledge, PI3KCA missense mutations in similar amino acid regions have not been described in gliosarcomas or glioblastomas with prominent sarcomatous differentiation. Further investigation is needed to determine if PI3KCA mutations are common in sarcomatous malignant gliomas, which may provide insight into the pathogenesis of this rare subset of glioblastomas. Furthermore, the mutation identified in Case 1 has been shown to predict response to PI3K/AKT/mTOR inhibitors, opening possibilities for targeted therapy in patients with high grade gliomas with sarcomatous differentiation3.

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172

Glioblastoma with Melanoma-like Histology and Widespread Extracranial Metastases. A Case Report with EM Analysis.

Michael Presta1, Rami Al-Rohil1, Walter Jacobson2, Julie Pilitsis2, Jiang Qian3. 1Pathology, Albany Medical College/Center Hospital; 2Neurosurgery, Albany Medical College/Center Hospital; 3Pathology, Albany Med Ctr Hosp/APS

Glioblastoma (GBM) is an aggressive astrocytoma with poor prognosis. Classic GBM is readily recognizable pathologically, but occasional cases pose diagnostic challenges due to their unusual features. Local spread and recurrence occur commonly in GBM, but extracranial metastasis is quite rare. Here we report a case of GBM with multiple systemic metastases, with supporting electron microscopy (EM) study. The patient was a 55 year old male who presented with three weeks of bilateral frontal headaches, without vision changes, sensory or motor deficits, and an unremarkable physical examination. MRI identified a right frontal enhancing mass and 3 small enhancements in the left fronto-parietal region. CT of the internal organs found no pathology. He underwent a right frontal gross total resection with no residual enhancement on post-op imaging. Resection specimen revealed a high grade tumor composed of sheets of melanoma-like, round epithelioid cells arranged perivascularly, with mitotic activity, microvascular proliferation, and extensive necrosis. Tumor cells stained strongly for S100 and vimentin, with limited GFAP and p53 expression, and stained negative for epithelial and melanocytic markers. Further EM study supported a glial origin of the tumor, showing intermediate filaments without melanin pigment. GBM was diagnosed, and the patient was treated with Temozolomide. Follow-up MRI at 3 months identified 12 enhancing areas in both cerebral hemispheres and the right cerebellum. PET scan revealed multiple pulmonary, intra-peritoneal and soft tissue masses in the trunk. The largest lung mass was biopsied, confirmed to be metastatic GBM. The patient died shortly thereafter, less than 5 months following initial brain surgery. While the adverse impact of extracranial spread of GBM on overall survival is debatable, its presence should be recognized so that proper clinical management can be planned. For unusual GBM cases like ours, with overlapping features with melanoma, EM is a useful adjunct in aiding diagnosis.

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173

Prevalence, Pathology, Prognostic Factors, and Survival in Latino Americans with Glioblastoma

Maryam Shabihkhani1, Yalda Behbahanian1, Kourosh Naeini2, Diviya Gupta1, Bowen Wei1, Gregory Lucey1, Lauren Hanna1, Desiree Sanchez1, Sergey Mareninov1, Timothy Cloughesy3, William Yong1. 1Department of Pathology, University of California, Los Angeles; 2Department of Radiology, University of California,LosAngeles; 3Department of Neurology, University of California, Los Angeles

There is limited data regarding the prevalence, pathology, prognostic factors, and outcome in Hispanic/Latino Americans. Given that Latinos comprise the largest and fastest growing minority group in the US, understanding the characteristics of glioblastoma in this population is an important public health concern. A literature search was conducted using Google Scholar and PubMed. Survival statistics and clinical and demographic variables were also extracted from the Survival, Epidemiology and End Results (SEER) database for patients diagnosed with glioblastoma (GBM) from 1973 to 2010. Descriptive statistics were used to describe the population and survival was analyzed using Kaplan-Meier curves and proportional hazard models. This study included 45,187 GBM patients, of whom 83.6% were non-Latino white, 7.6% were Latino, 5% were African American, and 3.9% were Asian/Other. There was no statistical significance in median age, gender, surgery, and radiotherapy between groups. Improved prognosis in Latinos and non-Latinos is associated with age <60 years, use of radiation, and gross total resection. One year survival rate is 31.5% in non-Latino populations and 35.9% in Latinos (p value= 0.6). The SEER database showed no difference in the prevalence of either giant cell glioblastoma or gliosarcoma between Latino and non-Latino populations. We did not identify literature regarding the prevalence of MGMT methylation or IDH1 mutations in Latinos. While 7.6% of GBM patients in the SEER database are Latino, it has been reported that 1.9% of Cancer Therapy Evaluation Program (CTEP) clinical trial patients are Hispanic suggesting that they may be under-represented. More detailed evaluation and comparison of sub-groups within the Latino population and against other ethnic groups may be informative. Additional molecular data regarding GBMs in Latinos is desirable.

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174

Glioblastoma Presenting As Lung Metastasis: A Unique Case Report And Review Of The Literature

Denise Ng1, Annie Wu1, Yalda Behbahanian1, William Yong1, Neda Moatamed1, Negar Khanlou1, Phioanh Nghiemphu1, Linda Liau1, Harry Vinters1. 1University of California, Los Angeles

We report a case of glioblastoma (GBM) presenting as pleural effusions originally considered a mesothelioma. CASE REPORT: The patient was a 54-year-old woman with no significant prior medical or surgical history. She presented with acute shortness of breath after strenuous exercise, prompting an emergency room consultation. Chest x-ray revealed a large left pleural effusion with radiologic consideration of mesothelioma. Patient underwent repeated thoracenteses. Malignant cells of undetermined origin were noted on cytologic analysis of the pleural fluid. Patient was originally scheduled for PET scanning, however, she began to complain of dysgeusia, heralding a tonic clonic seizure with loss of consciousness. Head CT revealed a diffuse T2 hyperintense, non-enhancing 2.1 cm mass in the medial right temporal lobe of the brain. Concomitant thoracic PET scan showed hypermetabolic areas in the ribs, vertebrae and lungs. Patient underwent a bronchoscopy and biopsy of the lung lesion, which was interpreted as malignancy not otherwise specified. Meanwhile, she underwent resection of the newly discovered brain lesion. Histopathologic examination showed a GBM, WHO grade IV. In view of these findings, prior pleural fluid samples and lung biopsies were reevaluated for possibility of a metastatic glioma. All specimens showed GFAP immunopositive tumor cells. Additional MRI revealed a space-occupying lesion in the lower cervical and upper thoracic spinal canal with continuity with the left pleural space. CONCLUSION: This case is unique in its presentation with respiratory symptoms due to the lung metastases, but also reminds pathologists of the metastatic potential of primary brain tumors. Molecular profiling of glial tumors has been used as a therapeutic guideline in recent years. Detailed studies are required to determine their significance in identifying tumors with greater metastatic potential. These molecular characteristics, including those pertaining to this case, will be further discussed.

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175

VE1 Immunoreactivity Patterns in Epithelioid Glioblastomas Positive for BRAF V600E Mutation

Bette Kleinschmidt-DeMasters1, Nicholas Foreman2, Seth Lummus3. 1University of Colorado Anschutz Medical Campus; 2The Children’s Hospital Colorado;3University of Colorado Dept. of Pathology

Background: Epithelioid glioblastomas (E-GBMs) are one of several GBM types not currently mentioned in the WHO 2007 classification system. However, fifty percent of these manifest BRAF V600E mutation, compared to a small percentage of ordinary GBMs, suggesting that they may possess a unique genetic signature - better qualifying them as variants rather than a pattern. Further, a targeted therapy for tumors with BRAF V600E mutation, vemurafinib, may make testing BRAF status important for treatment. It is unclear whether BRAF VE1 immunohistochemistry (IHC) can substitute for Sanger sequencing in these tumors, or if the IHC pattern differs between E-GBMs and other mutated tumor types, gangliogliomas (GG) and anaplastic pleomorphic xanthoastrocytomas (a-PXA). Finally, proof of principle has yet to be reported that E-GBMs with this mutation might respond to vemurafinib.

Design: We conducted VE1 IHC on our previously studied cohorts of E-GBMs, GGs, and a-PXAs (all known to possess the mutation) and compared the patterns of immunostaining. We also report the results of two patients recently treated with vemurafinib - one pediatric a-PXA and one of the E-GBMs.

Results: All E-GBMs manifested strongly diffuse cytoplasmic immunoreactivity, with uniform intensity in 7/8 and diffuse positivity in the 8th, with scattered more-intensely immunoreactive tumor cells. GGs demonstrated far more intense immunoreactivity in the ganglion, than glial, tumor component. One E-GBM patient with initial gross total resection quickly recurred within 4 months, required a second resection, and then was placed on vemurafinib; she remains tumor-free 23 months later without neuroimaging of residual disease. One pediatric a-PXA recurred despite drug therapy and required stereotactic radiosurgery, but also survives 32 months later.

Conclusions: E-GBMs manifest the same diffuse strong IHC immunoreactivity pattern seen in melanoma and, although anecdotal, can respond to targeted therapy.

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176

“Gliomatosis encephali” as a Novel Category of Brain Tumors: The First Autopsy Case Report of Gliomatosis Cerebelli

Asa Nakahara1, Toshikazu Yoshida2, Masanobu Yazawa2, Takashi Ehara3, Jun Nakayama4, Akiyoshi Kakita5, Ryosuke Ogura5, Mika Asakawa6, Emi Suzuki-Kouyama6, Kiyomitsu Oyanagi6. 1Shinshu Univ. Sch. of Med.; 2Dept. of Med. (Neurol.), Fujimi-kogen Med. Center; 3Dept. of Pathol., Shinshu Univ. Sch. of Med.; 4Det. of Molecular Pathol., Shinshu Univ. Graduate Sch. of Med.; 5Dept of Pathol., Brain Res. Inst., Univ. of Niigata; 6Dept. of Brain Disease Res., Shinshu Univ. Sch. of Med.

Gliomatosis cerebri is a rare diffuse glioma that is neither mass-forming nor necrotic, and does not disrupt existing structures. Gliomatosis occurring in the cerebellum is known as gliomatosis cerebelli, and only three such cases examined by biopsy have been reported. Here we describe the first autopsy findings of a patient who was diagnosed as having gliomatosis in the cerebellum. Neuropathological examination identified the tumor cells as being positive for glial fibrillary acidic protein, vimentin and nestin, with atypical nuclei that were cashew-nut- or dishcloth-gourdshaped. These tumor cells were dense in the right cerebellum, but also spread broadly throughout the brain including the left cerebrum and optic nerve. Mitotic figures were frequently seen in the cerebellum, brain stem and cerebrum. Scherer’s secondary structures were evident not only in the cerebellum but also the cerebrum. No necrosis, microvascular proliferation or destruction of anatomical structures was detected in the whole brain. Differences in the origin of the tumors of the gliomatoses cerbri and cerebelli suggests these tumors are different types of brain tumors. Thus the findings support that the gliomatosis cerebelli is a novel type of brain tumor classification. Furthermore, by the similarities of the histological features among the tumors, it appears appropriate to establish a novel category of “gliomatosis encephali” which includes both gliomatosis cerebri and gliomatosis cerebelli.

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177

Astrocytomas as Secondary Tumors in Patients Treated for Medulloblastoma: A Case Series

Dilys Chen1, Marco Hefti1, Sandro Santagata2, Rolf Pfannl1. 1Department of Pathology, Beth Israel Deaconess/Harvard Medical School; 2Department of Pathology, Boston Children’s Hospital

The increasing number of medulloblastoma survivors after aggressive multi-modality therapy makes understanding long-term sequelae critically important. We report the largest series of patients with astrocytomas occurring after treatment for medulloblastoma to date (n=5), and the longest reported latency. Medulloblastoma was diagnosed at a mean age of 3.5 years (range 2 to 6) and the secondary astrocytoma a mean of 14 years later (6 to 30). Of our cases, three were glioblastomas (WHO Grade IV) and two anaplastic astrocytomas (Grade III). Four cases were in the posterior fossa, while one was in the right temporal lobe. Four patients died of their secondary tumor, with a mean survival of 10.8 months, while one of the two anaplastic astrocytoma patients was alive at 8 months post-diagnosis. The right temporal lobe tumor had chromosomal analysis performed by array-based comparative genomic hybridization and was found to have only an 8.4 MB single copy loss of 9p21.3-9p21.1 resulting in a loss of CDKN2a which is rare in medulloblastomas, but common in high grade astrocytomas. It is unlikely that any of these tumors were sporadic, as high grade gliomas, particularly in the posterior fossa, are rare in children and young adults. In addition three of our cases showed clear histological changes consistent with previous radiation effect adjacent to the tumor. It is highly unlikely that any of these tumors were derived from the original medulloblastoma cells, given the long latency and the absence of residual/recurrent medulloblastoma in the interim, although this cannot be completely excluded. Radiation-induced glioblastomas are uncommon but are known to occur in patients with prior therapy for medulloblastoma in childhood. Although these cases are rare, these examples demonstrate the need for adequate surveillance.

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178

ImmunoFISH is a Reliable Technique for the Assessment of 1p and 19q Status in Oligodendrogliomas.

Peter Gould1, Céline Duval2, Marie de Tayrac3, François Sanschagrin2, Karine Michaud1, Stéphan Saikali1. 1Hôpital de l’Enfant-Jésus du CHU de Québec; 2CHU de Québec; 3Université de Rennes

We developed an immunoFISH technique for the study of oligodendrogliomas by combining a classical immunohistochemistry technique using MIB-1 antibody with a standard FISH technique using commercial 1p36 and 19q13 chromosomal probes on the same section.

This technique permits an analysis of 1p and 19q status restricted to the actively proliferating tumour cell population, which may be advantageous for small tissue samples with a mixture of tumour and non-tumour cells. Validation was performed on a series of 36 pre-selected oligodendrogliomas (12 grade II, 21 grade III and 3 grade IV) and compared to the results previously obtained by FISH.

ImmunoFISH technique appears easy to perform and to analyse and is little more time-consuming than the FISH technique. Inter-observer reliability of immunoFISH is high when detecting deletions by the ratio method (1p, κ = 0.71; 19q, κ = 0.72) but higher when using the combination method (1p, κ = 0.86; 19q, κ = 0.95).Compared to FISH, immunoFISH exhibits a very high sensitivity (∼ 100 %) and a high specificity (∼ 90 %) for 1p and /or 19q deleted cases. The sensitivity decreases slightly for non-deleted cases (∼ 85 %) as does specificity (∼ 60 % for 1p and ∼ 85 % for 19q). For unbalanced cases the sensitivity remains high for both 1p (∼ 90 %) and 19q status (∼ 100 %) but specificity decreases (∼ 50 % for both 1p and 19q status). There was no significant difference between FISH and immunoFISH results calculated on 60, 40 or 20 cells, regardless of the method of calculation used (combination or ratio) or the type of chromosomal status studied (Chi-square test not significant). Our study demonstrates the reliability of the immunoFISH technique in the study of 1p and 19q status in oligodendrogliomas and emphasizes its advantage in poorly cellular tumour specimens.

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179

A Case of Bone-Only Extraneural Metastatic WHO Grade II Oligodendroglioma

Zachary Hoffer1, Desiree Marshall1, Luis Gonzalez-Cuyar1, C. Dirk Keene1, Theodore Burke1, Marc Chamberlain1, Donald Born2, Margaret Flanagan1. 1University of Washington; 2University of Washington, Stanford University

Metastases to the brain are the most common form of intracranial malignancy; however, extraneural metastases of primary intracranial neoplasms are very uncommon. Examples of primary intracranial tumors exhibiting extraneural spread are meningiomas and hemangiopericytomas in adults and medulloblastomas and ependymomas in pediatric patients. Gliomas rarely metastasize, but when they exhibit extraneural spread it is more commonly originating from glioblastomas and gliosarcomas. Here we report the case of a 48-year-old man who presented with headaches and a seizure prompting imaging which revealed a non-contrast enhancing left parenchymal frontal tumor. Approximately one year later the patient underwent resection of a WHO grade II oligodendroglioma. The patient did well after surgery and was expectantly managed for five years until he developed primary site tumor recurrence; review of the original pathology showed the tumor was co-deleted for chromosomes 1p and 19q. Standard dose temozolomide therapy was administered for one year, and the patient did well for an additional three years with stable radiographic disease until he developed subacute progressive lower back pain prompting imaging. Plain radiographs and thoracolumbar magnetic resonance imaging revealed osteosclerotic lesions distributed in multiple thoracic vertebrae and the sacrum. Computerized tomography of the chest, abdomen and pelvis confirmed multiple metastases but no peripheral primary site of disease. A biopsy of the sacral lesion showed an epithelioid tumor composed of neoplastic cells with prominent nucleoli and eosinophilic cytoplasm that were immunoreactive to GFAP and S100 and co-deleted for 1p and 19q. IDH-1 (R132H) testing by immunohistochemistry in the brain primary and sacral tumor was negative. Although the histology was slightly different, taken together the immunohistochemical and molecular features were consistent with metastatic oligodendroglioma. Recognizing the rarity of extraneural metastatic oligodendrogliomas, it is unclear if immunohistochemically wild type IDH-1 (R132H), such as our case, have an increased propensity to metastasize.

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180

Alpha-Internexin Immunoexpression in Oligodendrogliomas – Potential Diagnostic and Pathogenetic Relevance.

Suash Sharma1, Dustin Gertsch1. 1Medical College of Georgia, Georgia Regents University

With the exception of 1p19q codeletion in oligodendrogliomas, no other markers reliably differentiate prognostically distinct oligodendrogliomas from diffuse astrocytomas. We evaluated immunoexpression of alpha-internexin (INA), a neuron-specific gene that encodes a 66-kd neurofilament-interacting intermediate filament protein and is one of the most differentially expressed genes between mutually exclusive 1p19q-codeleted and EGFR-amplified high-grade gliomas, in oligodendrogliomas and associated mini-gemistocytes. Thirteen archival cases of oligodendrogliomas of all histologic grades were immunostained for alpha internexin, with autopsy brain tissue as positive control. The staining of tumor cells were interpreted as 1+ (<10% positive cells), 2+ (11-50%) and 3+ (>50%), while carefully excluding the known immunostaining of native neurons and axons. Extensive (3+) staining with INA was seen in 5/13 (38%) oligodendrogliomas. While some staining with INA was seen in 7/7 (100%) oligodendrogliomas with 1p19q codeletion and in 2/3 (66%) without the codeletion, however >10% cell staining was seen in only 4/7 (57%) oligodendrogliomas with codeletion (comparable to Buckley et al). Minigemistocytes were stained in 11/12 (82%) with INA. Extensive (3+) staining was identified in 3/6 (50%) high-grade oligodendrogliomas (grades 3 or 4), as compared to 2/7 (29%) grade-2 oligodendrogliomas. To conclude, our preliminary study showed extensive immunoexpression (>50% tumor cells) of alpha-internexin in 38% oligodendrogliomas. Although more selective than the literature, its positivity in oligodendroglial mini-gemistocytes, and higher proportion of high-grade oligodendrogliomas suggests that it may be of diagnostic value and further support evidence of neuronal differentiation.

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181

Ki-67 and GFAP Dual Stain Employed in Surgical Neuropathology

Paul Mckeever1, Sandra Camelo-Piragua1, Kristina Fields1, Jonathan Mchugh1. 1University of Michigan Department of Pathology

Background: Both glial fibrillary acidic protein (GFAP) and Ki-67 markers are used extensively in surgical neuropathology. Distinguishing neoplastic from other proliferating cells is important to determining proliferative activity of a tumor. Does staining one tissue section for both GFAP and Ki-67 help make such distinctions?

Design: Twenty gliomas and ten other lesions were used to compare the value of single GFAP and Ki-67 immunostaining versus dual staining (DS) for Ki-67/GFAP. Ki-67 stained the tissue first, and its hard reaction product retained the proliferation index during additional staining. From a specimen block, Ki-67/GFAP DS section and individual serial sections stained for either Ki-67 or GFAP were used to quantify Ki-67/GFAP +/+, +/-, -/+ , -/- cells, and to quantitatively compare these cell types. A comment was included if any stains enhanced interpretation or enabled diagnosis.

Results: Ki-67/GFAP DS outperformed serial sections of single stains in identifying individual GFAP+ proliferating and non-proliferating cells. Ki-67/GFAP DS provided better information about specimens containing well mixed populations of GFAP positive and GFAP negative cells: Mixed glial and neuronal tumors, oligoastrocytomas, and margins of astrocytomas. In mixtures of astrocytes and other cells, Ki-67/GFAP DS highlighted the population of proliferating GFAP+ astrocytes when present. Ki-67/GFAP DS increased accuracy of counting proliferation indices of astrocytomas and ependymomas by excluding proliferating GFAP negative cells associated with small vessels and leukocytes. Ki-67/GFAP DS has been used for half a year on various diagnostic problems. It has been particularly helpful with fibrous tumors invading brain versus gliosarcoma, anaplastic astrocytoma versus glioblastoma, and margin of diffuse astrocytoma versus reactive changes. Cases will be illustrated.

Conclusions: Ki-67 and GFAP dual staining is most useful in evaluating lesions containing mixed populations of GFAP positive and GFAP negative cells. In these situations, dual staining was superior to sections of single stains.

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182

Methylthioadenosine Phosphatase Expression in Pilocytic Astrocytomas and its Relationship with Oncogene-Induced Senescence

Aline Becker1, Cristovam Scapulatempo-Neto1, Weder Menezes1, Carlos Clara1, Ricardo Oliveira2, Helio Machado2, Marileila Varella-Garcia3, Luciano Neder4, Rui Reis5. 1Molecular Oncology Research Center, Barretos Cancer Hospital, Brasil; 2Ribeirão Preto School of Medicine, University of São Paulo/FMRP-USP; 3Division of Pathology National Institute of Cancer, Braga, Portugal; 4Ribeirão Preto Medical School - FMRP/USP; 5Molecular Oncol Res Center, Barretos, Brasil and ICVS, Univ. Minho, PT

Background: Pilocytic astrocytomas (PAs) are the foremost glioma in children and in general presents an indolent course. The main involved molecular mechanism is constitutive activation of MAPK pathway, mostly through KIAA-BRAF (K:B) fusion. This molecular alteration may trigger oncogene-induced senescence (OIS), a mechanism recently implicated in the benign behavior of PAs. In fact, an increased expression of methylthioadenosine phosphatase (MTAP) gene has been referred in astrocytes of dementia and ageing animal models, related to the phenomenon of senescence. In contrast, loss of MTAP expression is related to poor prognosis in leukemias and other malignancies, including glioblastomas. Thus far, the role of MTAP in PAs is unknown. Herein, we aimed to assess MTAP expression and its role in the biologic behavior of PAs.

Methods: A cohort of 69 patients (1.2:1 M/F, median 11.6 years-old), was evaluated to assess the expression of MTAP by immunohistochemistry in FFPE on two tissue microarrays (TMAs) platforms and correlated with clinicopathological and molecular data of K:B fusion by fluorescence in situ hybridization (FISH), with a customized dual-target dual-color probe.

Results: More than 95% of the PAs (66/69) showed overexpression of MTAP, contrasting to the faint reaction in the astrocytes of the adjacent non-neoplastic cerebellar cortex. The MTAP overexpression was positively correlated with cerebellar lesions (p=0.025) and the presence of K-B fusion (p=0.028). No other significant differences were noted.

Conclusions: To best of our knowledge, this is the first study that shows an overexpression of MTAP in PAs with a reduced expression in adjacent non-neoplastic tissue. This pattern of expression reinforces a positive relationship between constitutive activation of MAPK pathway through K-B fusion and the OIS phenomenon. This fact could be one of the mechanisms responsible for the indolent behavior of PAs, but further studies with larger cohorts are necessary to confirm this relationship. Support: PAIP

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183

Immunohistochemical Characteristics of Centyrin & DARPin Targets in Brainstem Gliomas and Correlation with Molecular Markers

Viktor Zherebitskiy1, Sakir Gultekin1, Cynthia Hawkins2, Christopher Corless3, Kellie Nazemi1, Charles Keller1. 1Oregon Health and Science University; 2The Hospital for Sick Children/ University of Toronto; 3Knight Diagnostic Laboratories/ Oregon Health and Science University

Brainstem gliomas are deadly due to their location. They occur mainly in the pediatric and adolescent population. They are not readily amenable to surgical treatment and respond poorly to combined radiation and chemotherapy. Centyrins and DARPins belong to scaffold proteins family. They have many pharmacologic advantages over monoclonal antibodies currently used for targeted therapies. It is important to confirm that their effector proteins have specific targets in the gliomas. We examined tissue from 17 DIPG cases (6 surgical biopsies, 2 autopsy cases and 9 in TMA archival material) studying distribution of 7 antigenic targets (IL4, IL4R, IL13, IL13RA1, IL13RA2, EGFR and MET/HGFR) using immunohistochemistry. Two neuropathologists (VZ & SHG) interpreted the results using semi-quantitative scoring system. The tumor material was also screened for variety of molecular targets including CDKN2A, EGFR, IGF1R, MET, MGMT, NF1, MYCN, PDGFRA, PTEN, RB1, P53, H3F3A, HIST1H3B and ATRX. We found that 86% of DIPGs were positive for IL4 and only 7% of them showed positive immunostaining for IL13. However, their receptors (IL4R, IL13RA1 and IL13RA2) were expressed in 75-85% cases. In about a half of the positive cases the expression was strong and global (>25% of positive tumor cells). The oncogenes (EGFR and MET) were expressed in about 40% of cases. There was no straightforward correlation between potential centyrin and DARPin targets studied by immunohistochemistry and molecular tumor markers. CONCLUSION: Our study showed fairly high and frequent expression of IL4 and IL13 receptors in the brainstem gliomas. These data can be used in designing specific centyrin/DARPin-based drugs for targeted therapies of DIPGs. Further studies are needed to find correlations between centyrin/DARPin targets and molecular makeup of the brainstem gliomas.

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184

Adult Brainstem Gliomas: A Case Series With Radiological, Pathological, And Clinical Correlation

Kimmo Hatanpaa1, Gregory Martin1, Jack Raisanen1, Chan Foong1, Bruce Mickey1, Charles White1, Robert Oberle1. 1UT Southwestern Medical Center

Brainstem gliomas are common in children and classifiable based on radiological and histopathological features into two clinically relevant groups: diffusely infiltrating pontine gliomas, which are associated with an unfavorable prognosis, and dorsal exophytic gliomas, which are associated with a favorable prognosis. In contrast, brainstem gliomas are rare in adults, and the clinical significance of radiological and histopathological features is less well established.

We reviewed the imaging findings, histopathological features, and clinical outcome in a series of 17 adult patients with histologically proven brainstem gliomas. The series consisted of 6 histologically classic pilocytic astrocytomas (WHO grade I), 1 diffuse astrocytoma (WHO grade II), 5 unclassifiable low-grade astrocytomas (WHO grade I or II), 1 anaplastic astrocytoma (WHO grade III), and 4 glioblastomas (WHO grade IV). The radiological features that were analyzed included the presence and pattern of contrast enhancement and whether the tumor was intrinsic or exophytic.

High-grade (WHO grade III-IV) gliomas were associated with a significantly shorter total survival compared with low-grade (WHO grade I-II) gliomas (p=0.0005). Specifically, all 6 patients with pilocytic astrocytomas were alive after a median follow-up of 7 years, whereas 4 of the 5 patients with high-grade tumors were dead after a median follow-up of 7.5 months. Four of the five patients with unclassifiable low-grade tumors were alive after a median follow-up of 3 years. None of the radiological variables was significantly associated with survival. Our data suggests that standard MRI techniques are poor predictors of clinical outcome and supports a low threshold for pursuing histological diagnosis in adult patients with suspected brainstem gliomas.

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185

Pediatric Brainstem Tumors with Features of Angiocentric Glioma: Report of Two Cases

Marie Rivera-Zengotita1, Jesse Kresak2, David Pincus3, Anthony Yachnis2. 1University of Florida; 2Department of Pathology, University of Florida; 3Department of Neurosurgery, University of Florida

Angiocentric glioma is a rare low grade neuroepithelial tumor with characteristic angiocentric growth pattern and ependymal differentiation. It typically affects children and young adults that present with chronic intractable seizures and a cerebral lesion. Such tumors arise most commonly in the frontoparietal or temporal regions as solid T2 hyperintense areas that occasionally show a "stalk-like" extension to the adjacent ventricle. We report the clinicopathologic features of two pediatric patients with low-grade brainstem tumors that showed histopathological and immunohistochemical features of angiocentric glioma. The first patient, a 5 year old girl, presented with double vision, headache, and nausea and was found to have papilledema on exam. Imaging studies revealed a T2 bright, non-enhancing lesion of the midbrain that caused aqueductal obstruction and hydrocephalus. The second patient was a 6 year old boy that had a non-enhancing diffuse pontine lesion that was bright on T2/FLAIR images and had a large exophytic component projecting into the fourth ventricle. Histologic examination of both cases revealed a neoplasm composed of monomorphous bipolar spindle cells, with nuclei containing granular ("crisp") chromatin, that were arranged in a distinctive angiocentric growth pattern. Tumor cells were immunoreactive for GFAP and showed a perinuclear dot-like pattern of reactivity for EMA. Only a rare mitosis was identified and the Ki-67 (MIB1) labeling index was low. The brainstem location and histologic features of these tumors are similar to a previously reported angiocentric glioma-like tumor that arose in the midbrain (Covington et al. Pediatr Neurosurg 2009;45:429-33). On follow-up, both patients reported here are alive with stable residual disease after 6 years (first patient) and 5 months follow-up. These findings suggest that brainstem tumors with angiocentric glioma-like features may represent a distinctive subtype of low grade glioma that should be included in the differential diagnosis of brainstem/posterior fossa lesions in the pediatric population.

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186

Composite Atypical Ganglioglioma (GG)-Pleomorphic Xanthoastrocytoma (PXA), Recurrent: A Case Report

Areli Cuevas-Ocampo1, Luis Moral1. 1Baystate Medical Center/Tufts University School of Medicine

Composite GG/PXA is a rare entity displaying histologically a GG adjacent to a PXA, with the second component recognized at presentation or recurrence. It shares clinicopathologic features of both individual tumors and when anaplasia is present it is usually associated with the PXA elements. We report a case of a composite GG/PXA presenting as atypical GG and recurring twice as PXA. The patient is a 24-year-old man who developed left-sided arm numbness and tingling. Imaging studies revealed a 5.4 × 3.6 × 3.1 cm solid/cystic enhancing right parietal mass. Gross total tumor resection was performed. Microscopically the hypercellular tumor had papillary formations, eosinophilic granular bodies, neoplastic cells with glial and neuronal phenotypes, no neurocytes, and up to 3 mitoses/10 HPFs. Microvascular proliferation or necrosis was not seen. Pleomorphic, xanthomatous cells and the classic reticulin network were absent. Frequent tumor cells were positive for GFAP and synaptophysin, some for p53 and few expressed CD34 and NF, with a Ki-67 index of 3%. The first and second recurrences occurred five and ten months later and exhibited the characteristic histopathologic findings of PXA with similar immunohistochemical results as above, except for only scattered synaptophysin positive tumor cells. Ganglion cells were not identified. Mitoses, necrosis or microvascular proliferations were not found in both recurrences. The original and recurrent tumors were positive for BRAF V600E mutation. We are reporting this case as a composite GG/PXA presenting as atypical GG and recurring with the classical histologic features of PXA. It suggests the possibility that mitoses, papillary formations and p53 immunoreactivity in some GG/PXAs, present in the GG component in this case, may be associated with aggressive clinical behavior. It also emphasizes the existing overlap between PXA and GG and supports the notion that PXA may correspond to a glioneuronal rather than a pure astrocytic neoplasm.

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187

Molecular Analysis of Pilomyxoid Astrocytoma and Variable Tendency Toward Maturation to Pilocytic Astrocytoma

Bette Kleinschmidt-DeMasters1, Andrew Donson2, Nicholas Foreman2. 1University of Colorado Anschutz Medical Campus; 2Children’s Hospital Colorado

Background: PMA was codified in WHO 2007 as a variant of PA, although it was given a different grade (WHO grade II) due to more aggressive behavior. Occasional reports have shown maturation of PMA to PA or intermediate forms of PMA-PA, validating the concept of keeping them within the PA spectrum. Additional information is needed regarding the clinical and biological relationships between PMA and PA.

Design: We reviewed our clinical and histological experience with pure PMAs (n=8), and analyzed gene expression using Affymetrix U113plus2 microarray chips. Findings were contrasted with PA gene expression profiles (n=10). Those >2-fold overexpressed were identified and subsequently analyzed using the web resource DAVID (Database for Annotation, Visualization and Integrated Discovery) and GSEA (Gene Set Enrichment Analysis).

Results: Clinical behavior varied considerably in our cohort; PMAs involving optic chiasm or cerebellum did significantly better and 4/4 patients survived, albeit with deficits. Diencephalic syndrome, seen in 4/4 PMAs centered in hypothalamus, was associated with demise in all cases. 1 of these 4 PMAs underwent autopsy and both maturation towards PA and transformation to GBM was identified within the cerebrospinal tumor deposits. In contrast, one surviving non-diencephalic PMA patient has undergone 5 subsequent surgical excisions, with documented progressive maturation towards PA over time. Many genes overexpressed in PMA versus PA were cell-cycle and mitosis-related. After ranking most differentially-expressed genes in PMAs, we identified upregulation of IMP3 (45-fold), (as reported, JNEN 2013:72:442), as well as key molecules in nervous system development, HMGA2 (16-fold) and EPHB2 (7-fold) (all p<0.00001).

Conclusions: PMAs have very variable behaviors and definite biological differences.

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188

BRAF Testing in Pleomorphic Xanthoastrocytoma: Insights from 3 Pediatric Cases

David Pisapia1, Ehud Lavi1. 1Weill Cornell Medical College

The presence of BRAF alterations is becoming increasingly recognized in a variety of pediatric central nervous system tumors. Moreover, the existence of small molecule inhibitors that target this mutated receptor may offer therapeutic options in more aggressive lesions in addition to surgery, conventional chemotherapy, and radiation. Certain entities such as pleomorphic xanthoastrocytoma (PXA), ganglioglioma (GG), and pilocytic astrocytomas (PCA) frequently show BRAF alterations and in histologically ambiguous cases, such molecular analysis may be a helpful adjunctive study in formulating a differential diagnosis. Here we highlight three informative cases that ultimately harbored BRAF alterations. In the first case the finding of a BRAF-V600E mutation was used to confirm the presence of a recurrent PXA with sarcomatous features where both the histology and location were distinct from the original tumor. In the second case, the presence of the BRAF-V600E mutation was used on limited biopsy tissue to support the notion that the astrocytic component of the lesion represented a tumor along the spectrum of PXA, GG, or PCA as opposed to the spectrum of diffusely infiltrating astrocytomas. Finally, a previously unreported CCDC6-BRAF fusion protein was found in the third case, which showed definitive histological features of the PXA/GG spectrum of tumors. This report highlights the diagnostic utility of BRAF mutations in addition to reporting a novel BRAF fusion protein that is postulated to act with functional similarity to that of BRAF-V600E. We find that targeted sequencing of this gene can be of great utility, particularly in diagnosing tumors that lie within the PXA/GG spectrum and in cases with histological ambiguity. Such testing is not only useful diagnostically, but also may serve as the basis for therapeutic decision making and the ability to refine prognosis.

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189

Cerebellar Pleomorphic Xanthoastrocytoma in a Patient with Neurofibromatosis Type 1: A Case Report.

Hidehiro Takei1, Meenakshi Bhattacharjee2. 1Houston Methodist Hospital; 2University of Texas Medical School at Houston

Pleomorphic xanthoastrocytoma (PXA) is an uncommon tumor of young adults, typically occurring supratentorially. It is generally considered to be a low grade, circumscribed tumor treated by surgical resection, with a relatively favorable outcome. Cases of cerebellar PXA have been rare, and cases associated with neurofibromatosis type 1 (NF1; PXA-NF1) are even rarer, with only 2 cases having been reported to date. We present here a third case of PXA-NF1 with unusual features.

The patient was a 33-year-old woman presenting with a history of headache for about 6 weeks. Her medical and family history was significant for NF1. Brain MRI revealed a 3.4 cm ill-defined lesion with a gyriform enhancing pattern in the left cerebellum, mimicking gangliocytoma. The patient had gross total resection of the lesion, and had an unremarkable postoperative course. Histologically, the lesion showed typical PXA (WHO grade II) features characterized by a cellular astrocytic proliferation with bizarre, pleomorphic tumor giant cells, some multinucleated, abundant eosinophilic granular bodies, and a typical intercellular reticulin network. Immunohistochemically, scattered synaptophysin positive tumor cells were identified. Notable however, the majority of tumor showed growth in the subpial compartment, with thickening of the cerebellar folia externally. P53-immunostaining was negative. Three months following surgery, local recurrence was detected, and radiation therapy was given. One year after the first surgery, she underwent surgical resection of the recurrent/residual tumor. Histologically, the recurrent tumor showed very similar features to the initially resected tumor, with additional radiation-related changes. All 3 reported cases of cerebellar PXA-NF1 including ours, showed recurrence. The reported cases of supratentorial PXA-NF1 have not been associated with recurrence, and thus, exhibit the same behavior as sporadic non-NF1-PXA. In contrast, cerebellar PXA-NF1 may not have such s favorable outcome.

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190

Anaplastic Pleomorphic Xanthoastrocytoma: Report of Two Diverse Cases

Suash Sharma1, Ravi Raghavan2. 1Medical College of Georgia, Georgia Regents University; 2Loma Linda University Medical Center

We profile two cases of anaplastic pleomorphic xanthoastrocytoma (APXA), a rare malignant variant for which diagnostic criteria are not well established in the WHO classification. Case 1: A 7-year old female presented with a large right fronto-parietal cystic mass with mural nodule. The tumor was mostly densely cellular and consisted of pleomorphic anaplastic astrocytic cells exhibiting frequent mitotic activity (7 per 10 hpf), palisaded necrosis, microvascular proliferation a Ki-67 labeling index of 20%, and negative IDH1. Differential diagnosis was between glioblastoma and APXA. Given the interspersed areas of lower cellularity, abundant eosinophilic granular bodies (EGBs), focally increased peri-/inter-cellular reticulin, xanthomatous and pleomorphic cells, and relative circumscription, the findings were most consistent with an APXA. Case 2: A 65 year-old male presented with a large left temporal, partially enhancing tumor with mass effect. Histopathology revealed a glial neoplasm with relatively sharp borders, and composed of GFAP-immunoreactive and focally CD34-positive markedly pleomorphic astrocytic cells with gemistocytic forms, bizarre giant cells, high mitotic activity, vascular endothelial proliferation, and necrosis with pseudopalisading. Interspersed were mildly sclerotic areas of lesser cellularity and mitotic activity, focal xanthomatous change, and reticulin staining. EGBs were scant or absent. An admixed neuronal/ganglionic component was not identifiable, although rare giant cells were focally NFP-positive. Losses on chromosome 10 (PTEN/DMBT1) or evidence of EGFR amplification characteristic of de-novo glioblastomas were absent, but gains on chromosome 7 were present. Overall impression was that of a high-grade glioma, indistinguishable from glioblastoma, that in view of imaging, histology and longer survival was best regarded an APXA. This patient survived for 5 years. To conclude, characteristic histology especially EGBs, with reticulin and CD34 expression, in the clinical context of slow-growing and circumscribed tumors can facilitate a diagnosis of APXA, with its implications of greater prognostic variability (including longer survival) as compared to conventional glioblastomas.

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191

Anaplastic Pilocytic Astrocytoma of the Cerebellum with a BRAF V600E Mutation

Ada Baisre1, Nitin Agarwal1, Anjali Seth2, James Liu1. 1Rutgers-New Jersey Medical School; 2University Hospital

We report a case of pilocytic astrocytoma with anaplastic features and a BRAF codon 600 mutation (V600E). Pilocytic astrocytoma (PA) is a relatively indolent, WHO Grade I neoplasm of children and young adults, arising commonly in the cerebellum, although it can arise from other locations as well. Anaplastic features are rarely seen in pilocytic astrocytomas and the criteria are not well established. Previous studies have shown that the biologic behavior of such tumors is similar to that of WHO Grade III astrocytomas, hence the survival rate is decreased compared to typical PAs. Fusion variants involving BRAF and KIAA1549 have been found in ∼80% of pilocytic astrocytomas. BRAF V600E mutations, although not common, can also be found in PAs, but have been seen more frequently in PAs located in the supratentorial compartment. Our case is that of a 30-year-old man who presented with sudden loss of consciousness and was found to have hydrocephalus due to a 2.4 × 2.6 × 4.2 cm, partially cystic, heterogeneously enhancing mass within the cerebellar vermis. A midline suboccipital craniectomy was performed and gross total resection of the lesion was achieved. Histopathological examination showed a tumor composed of highly cellular areas mixed with low cellular areas of classic pilocytic astrocytoma. The highly cellular areas were histologically variable, including spindle, epithelioid, and focally chordoid morphology. The mitotic activity in these areas was ∼1-2 × 10 HPF and a focus of necrosis was identified surrounded by microvascular proliferation (glomeruloid tufts). Both anaplastic features and the presence of the BRAF V600E mutation are uncommon findings in pilocytic astrocytomas. Therefore, it is not clear whether the BRAF V600E mutation in our case played a role in the development of these anaplastic features, and hence, a more aggressive behavior with a probable decreased survival.

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192

Low Grade Glioneuronal Neoplasm of the Third Ventricle With PIK3CA Mutation. A Case Report and Review of the Literature

Patrick Malafronte1, Laurence Davidson1, Brett Theeler1. 1Walter Reed National Military Medical Center

Low-grade glioneuronal neoplasms are a heterogeneous group of tumors. While their histologic and clinical features are often distinct, occasional cases present in unusual sites with overlapping histologic appearances. In such settings it can be difficult if not impossible to definitively classify lesions, leading to frustration on the part of treating clinicians. Several ancillary tests are available to assist in differentiating histologically similar neoplasms, but these have not been extensively studied in this group of CNS tumors. We present a case of a 35 year old man presenting with a one week history of headache and intermittent, slowly progressive horizontal diplopia. MRI demonstrated a non-enhancing T2 hyperintense lesion within the body and posterior third ventricle with involvement of bilateral thalami and associated hydrocephalus. Microscopic sections revealed a neoplasm composed of oligodendroglioma-like cells with focal fibrillar and perivascular pseudorosettes in a mucinous microcystic background. There were many areas of microvascular proliferation. The neoplastic cells stained focally with GFAP. Synaptophysin reactivity was largely restricted within and around rosettes. The MIB-1 index was <2%. IDH-1 and p53 IHC were negative. Molecular analysis demonstrated intact 1p19q and no BRAF mutation (V600E). PIK3CA mutation analysis detected a mutation in exon 9 (E545K, c.1633G>A). PIK3CA mutations have mostly been reported in high grade gliomas; however, they can also be seen in some rosette-forming glioneuronal tumors (RGNT). Other tumors in the differential for this case (DNET, pilocytic astrocytoma) have not been extensively examined for PIC3CA mutations, but given the potential therapeutic benefit of PI3K/AKT/mTOR inhibitors, such study seems indicated, especially for use in recurrent or unresectable tumors. In addition, the discovery of potential recurrent molecular abnormalities may help to better classify this group of understudied neoplasms.

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193

Multinodular and Vacuolating Neuronal Lesions: An Evolving Classification of Low-Grade Neoplasms of the Cerebrum

Lyndsey Emery1, Donald O’Rourke1, Maria Martinez-Lage1. 1Hospital of the University of Pennsylvania

Background: Low-grade glioneuronal tumors include gangliogliomas, dysembryoplastic neuroepithelial tumors and papillary glioneuronal tumors. However, current classification does not necessarily encompass the entire spectrum of low-grade lesions with neuronal differentiation. Recently, twelve cases were reported as “multinodular and vacuolating neuronal tumors” exhibiting similar presentations, radiographic findings, histopathologic features and outcomes1,2. Here, we present two additional cases, which are thought to belong to this novel entity.

Cases: The first case is from a 24-year-old female with long-standing seizures refractory to multiple medications, since age 5. MRI revealed a focus of T2/FLAIR cortical and subcortical hyperintense signal within the left occipital lobe, with mild gyral expansion and minimal mass effect, for which she elected surgical resection. The second case is from a 30-year-old female with no known seizures. She underwent MRI for intermittent episodes of vertigo, which revealed a cluster of well-delineated, variable sized T2/FLAIR hyperintense cysts in the left frontal subcortical white matter. Interval imaging was stable; however, she elected for surgical resection. Histopathologic examination of both specimens revealed areas of distinct to coalescent nodules within the subcortical white matter at the cortical-white matter junction and focally extending into the cortex. The nodules were composed of bland cells with round to ovoid nuclei, open chromatin, variably prominent nucleoli and moderate amounts of eosinophilic cytoplasm in a background of vacuolated neuropil. Immunohistochemical stains demonstrate neuronal origin. Case 1 was also notable for a robust CD34-positive nodular meshwork. The overlying cortex shows evidence of architectural disarray, without significant mitotic activity, necrosis or microvascular proliferation. Next-generation sequencing of 47 gene hot spots (including BRAF) was negative for somatic mutations in Case 1.

Conclusions: We have identified two cases morphologically and clinically consistent with the recently described “multinodular and vacuolating neuronal tumors”. We propose that these are low-grade neoplastic, or possibly hamartomatous, lesions warranting further characterization.

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194

Synaptophysin Positive Atypical Neuroepithelial Tumor With Diffuse GFAP Expression at Recurrence: A Case Report.

Areli Cuevas-Ocampo1, Luis Moral1. 1Baystate Medical Center/Tufts University School of Medicine

The overlap between oligodendroglioma and extraventricular neurocytoma is rare and has been reported seldom in the medical literature. The biological behavior is thought to be analogous to that of conventional oligodendroglioma. We report a neuroepithelial neoplasm with neurocytic/oligodendrocytic differentiation and atypical histologic features. The patient is a 30-year-old woman who presented with sudden elevation of intracranial pressure. Imaging studies revealed a 6.6 × 5.4 × 4.0 cm solid/cystic right parietal lobe mass which recurred two years later. The patient underwent gross total tumor resection on both occasions confirmed by imaging studies. Histologically, the initial and recurrent tumors showed similar features; a high cellular density with oval to spindle-shaped nuclei, mild- moderate pleomorphism, perinuclear spaces in some cells, thick-walled vessels, focal calcifications and microvascular proliferation. The first resection showed a focally well-demarcated brain/tumor interface, strong reactivity for synaptophysin and variable staining for GFAP, 3 mitoses/10 HPF, Ki-67 of 7.6% and no necrosis. The recurrence displayed focal necrosis, 6 mitoses/10 HPF, strong immunoreactivity for GFAP but no staining for synaptophysin. Ki-67 appeared to be higher than in the initial resection. Neither tumor was immunoreactive for IDH1 or neurofilament protein. 1p/19q analysis by FISH failed to demonstrate co-deletion in both lesions. Although this did not exclude oligodendroglioma, the lack of 1p/19q co-deletion, and negative IDH1, favored the initial diagnosis of extraventricular neurocytoma. Electron microscopy was attempted out of the paraffin blocks but the results were inconclusive. The exact classification of this tumor is somewhat unclear since they share similar morphologic features but a dissimilar immunophenotype. Whether this neoplasm represents an extraventricular neurocytoma with oligodendrocytic differentiation or an oligodendroglioma with neurocytic differentiation is open to discussion. Its atypical histologic features will likely determine its clinical behavior. This case demonstrates the intriguing relationship that exists between oligodendrocytic and neurocytic neoplasms.

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195

An Intracranial Secondary Malignant Neoplasm after Wilms Tumor in a Four- Year -Old Girl

Rong Li1, Dava Sue Cleveland1, Jeffrey Blount2, Alyssa Reddy3, James Post1, David Kelly1. 1Department of Pathology and Laboratory Medicine, Children’s of Alabama; 2Department of Neurosurgery, Children’s of Alabama; 3Department of Neurology, Children’s of Alabama

Wilms tumor (WT) is the most common renal tumor in children with overall survival rate of about 85%. However, the survivors of WT have substantial risk of developing secondary malignant neoplasms, the most common tumor type of which is carcinoma, followed by sarcoma and leukemia. Rare cases of medulloblastoma and primitive neuroectodermal tumor have also been documented in WT patients. However, to the best of our knowledge, ependymoma has not been reported previously in patients after WT. Herein, we present the case of a four-year-old female who initially presented with left-sided abdominal asymmetry in September 2012. Imaging studies revealed a large left-sided abdominal mass. A diagnosis of classic triphasic WT with favorable histology was made by biopsy and a left nephrectomy was performed after initial chemotherapy. The patient was treated with abdominal radiation and additional chemotherapy. She subsequently presented with chronic headache in November 2013. MRI scan demonstrated a heterogeneously enhancing fourth ventricular mass. Suboccipital craniotomy was performed and microscopic examination demonstrates an anaplastic ependymoma characterized by numerous perivascular pseudorosettes and scattered true rosettes formed by relatively uniform cells. Portions of this tumor displayed marked hypercellularity and increased nuclear hyperchromasia. Microvascular proliferation, occasional mitotic figures, and geographic tumor necrosis were also present. The tumor cells were diffusely positive for glial fibrillary acidic protein and displayed dot-like positivity for epithelial membrane antigen in scattered tumor cells. In summary, we describe a case of fourth ventricular anaplastic ependymoma in a child who has been treated successfully for WT. Clinicians, radiologists, and neuropathologists should be aware of this rare association between ependymoma and WT. Interestingly, the Wilms tumor gene (WT1) is expressed in numerous neuroepithelial tumors, including ependymoma, and is increased with the grade of malignancy. The important role of WT1 in tumorigenesis and progression of brain tumor is worthy of further investigation.

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196

Malignant Evolution of Desmoplastic Glioma with Anaplastic Features, Including Postmortem Examination.

Catherine Stefaniuk1, Mohamed El Hag1, Duncan Stearns1, Michael Coffey1, Mark Cohen1, Marta Couce1. 1Case Western Medical Center/University Hospitals

Desmoplastic infantile gliomas (DIG) are WHO grade I supratentorial neoplasms of voluminous size with divergent differentiation and intense desmoplasia which typically follow a benign course following resection, even when accompanied by anaplastic features. Here we report a girl diagnosed with an anaplastic DIG at 3 months of age who died of disease two years later. We describe the original histologic findings (Diagnostic Slide Session Case 2012-5) and compare them with two subsequent surgeries and post-mortem evaluation. MRI at presentation demonstrated a large hemispheric cystic mass with enhancing nodular components. Complete resection was accomplished in two stages. Microscopic evaluation of the original tumor demonstrated a meningeal-based desmoplastic astrocytic neoplasm with regionally high mitotic activity and palisading necrosis. Three months later, MRI demonstrated increased enhancement surrounding the resection cavity. A repeat resection showed similar pathologic findings. The patient was given six months of vincristine, cyclophosphamide, cisplatin and etoposide without radiation (POG protocol #8633). One month later she had a third recurrence, which showed recurrent/residual glioma, reactive astrocytosis with dystrophic calcifications, and focal cortical dysplasia. The patient went under hospice care and died two years after initial diagnosis. Autopsy showed a large necrotic mass affecting the entire right cerebral hemisphere and crossing the corpus callosum, with extensive subarachnoid and subependymal spread. Histological findings were similar to those seen in her previous resections. As demonstrated by this case and others, DIGs have the potential to behave aggressively. We suggest that ominous histologic findings such as sarcomatoid architecture, palisading necrosis, and increased mitotic figures be graded on their own merit and not be considered within the spectrum of a WHO grade I tumor.

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197

Disseminated Choroid Plexus Papillomas in Adults. A Review of the Literature

Marwah Abdulkader1, Gregory Bosh1, Edward Dropcho1, Aaron Cohen-Gadol1, Jose Bonnin1. 1Indiana University School of Medicine

Choroid plexus papillomas (CPPs) are uncommon, usually intraventricular, low-grade tumors accounting for less than 1% of all intracranial neoplasms and 1-3% of brain tumors in children. Dissemination of CPPs to multiple levels of the neuraxis has been observed. A 21-year-old female had a craniotomy with resection of a CPP involving the right lateral ventricle. She had multifocal disease and not all areas of the tumor were resected. No postoperative radiation or chemotherapy was given but no progression was observed over the ensuing years. Seventeen years later, she developed episodes of dizziness, some short-term memory impairment and episodes of confusion. At a visit to her primary care physician, because of a gingival abscess, a follow-up MRI was obtained. This revealed hydrocephalus and multiple intraventricular and subarachnoid lesions compatible with disseminated CPPs. The tumors masses involved both lateral ventricles as well as the third and fourth ventricles. The largest one was in the third ventricle, extended into the suprasellar compartment and compressed the optic chiasm. This lesion was resected via an endoscopic craniofacial approach. The surgical procedure has been reported elsewhere. Histopathologically, the tumor revealed the characteristic architectural and cytologic features of CPPs, with very low mitotic and Ki-67 labeling indices. Multifocal CPPs at the time of initial presentation is an extremely rare occurrence and only twenty-two cases of disseminated CPPs after the original surgical resection have been reported in the literature. With some exceptions, disseminated CPPs have been observed in adults and they involved multiple sites along the CSF pathways. Occasionally, intraparenchymal extension has been documented and secondary involvement in the suprasellar cistern was previously reported in only five cases. There is no consensus on the postoperative treatment of disseminated CPPs. Generally, the metastatic lesions maintain the low-grade features of the primary tumor.

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198

Recurrent Subependymoma of Fourth Ventricle with Unusual Histological Features: A Case Report

Nishant Tiwari1, Suzanne Powell2, Hidehiro Takei2. 1Baylor College of Medicine, Houston, TX; 2The Methodist Hospital, Houston, TX

Subependymoma is a rare variant of ependymal neoplasm with distinct histological features. It is a slow-growing and well-circumscribed tumor, corresponding to WHO grade I. Many cases are incidentally found at autopsy, while surgically resected cases are usually those arising from the 4th ventricle. Subependymoma is characterized histologically by clusters of ependymal tumor cells arranged in a gliofibrillary matrix with no high grade features. We present here a very unusual case of recurrent subependymoma with high grade histological features.

The patient was a 62-year-old female who previously underwent a gross total resection of a 4th ventricular subependymoma 6 years ago. No recurrence had been found by follow-up MRI until 4 years ago. Recently, she presented with symptoms related to increased intracranial pressure and was found to have an enhancing mass in the 4th ventricle by MRI. She subsequently underwent resection of the recurrent tumor.

Histologically, the resected tumor showed a characteristic histologic pattern of subependymoma with small clusters and nests of ependymal cells with an interspersed gliofibrillary matrix. No true ependymal rosettes or perivascular pseudorosettes were present. In addition, there were very unusual histological features, including focal areas of necrosis, microvascular proliferation, thrombosed blood vessels, and scattered mitotic figures. Immunohistochemically, scattered p53 positive tumor cells were noted, and a Ki-67 labeling index was approximately 5% (15% in the highest areas).

Subependymomas are known to be low-grade lesions and are usually cured if completely excised. The tumor presented here is very unique in that several high grade pathological features were found in an otherwise typical subependymoma. Although subependymomas can have higher grade tumor components (most commonly with ependymoma; hybrid tumor), high grade pathologic features are usually not seen. Our case may represent anaplastic transformation of subependymoma, although no such examples have been reported to date, and close clinical follow-up has been recommended.

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199

Ependymal Tumor of the Pituitary

Derick Aranda1, Joseph Parisi1, John Atkinson1, Mark Jentoft1. 1Mayo Clinic

Although a wide range of histologic entities may occur in the sella, only rare cases of ependymal lesions have been described in this region. We report a 60-year-old man who presented with a mass lesion within the sella turcica clinically thought to represent a pituitary adenoma that demonstrated histomorphologic, immunohistochemical, and ultrastructural features of an ependymoma. The lesion showed immunoreactivity to GFAP, EMA, and S100; but none to pituitary hormones or chromogranin. It is important to separate this entity from pituitary adenoma since the clinical behavior of sellar lesions with ependymal differentiation is not known. The tumor also stained positively for TTF 1, which is of interest since the origin of these lesions is not certain. A hypothesis is that these ependymal tumors of the pituitary represent a variant of pituicytoma, which originate from “ependymal pituicytes.” TTF 1 is expressed in the ventral forebrain and pituitary during development. Although TTF 1 positivity supports a variant of pituicytoma, it rarely has been reported in cases of 3rd ventricular ependymoma. Consequently, the origin of this lesion remains speculative, although recognition of this entity will stimulate future studies and follow-up.

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200

Combining Whole Slide Imaging with Telepathology for Neuropathology Intraoperative Consultation

Jo Elle Peterson1, Andreana Rivera1, J Goodman2, Michael Thrall1, Suzanne Powell1. 1Houston Methodist Hospital; 2Baylor College of Medicine

Whole slide imaging (WSI) is a technology employed at the Houston Methodist Hospital system to create digital reconstructions of pathology glass slides. This methodology facilitates rapid long-distance transmission of images for viewing by branch hospital pathologists and remote consultations for diagnostic problems. For a standard tissue section, WSI creates a two-dimensional image at 20× magnification. Cytology slides, where cellular detail is of critical importance, however, require three-dimensional evaluation. While it is possible to create a more three-dimensional image with WSI, it takes longer, requires more memory, and is slow to load and manipulate. For our routine neuropathology intraoperative consultation, both cytologic preparations and frozen tissue sections are utilized for diagnostic purposes. WSI for cytologic preparations in the intraoperative setting is not possible, given the requirement of a 20 minute turn-around-time. Intranet-connected microscope-mounted cameras (netcams) make remote evaluation of cytologic preparations possible by live streaming of images. We examined the combined use of these technologies in the remote diagnosis of intraoperative consultation neuropathology specimens. Board certified neuropathologists remotely evaluated twenty-five neuropathology cases in a simulated intraoperative consultation setting. All cases had two cytologic preparations. Twenty-three cases had a single frozen section slide, while two cases had two frozen section slides. The following times were recorded: time to perform WSI and make imaging available to the neuropathologist; time reviewing cytologic preparations via netcam; time reviewing WSI reconstruction(s); and total time to diagnosis (turn-around-time). A turn-around-time of less than 20 minutes was possible in all cases, including those containing multiple frozen section slides and those requiring rescanning due to WSI system malfunctions. The turn-around-times ranged from 5:10 (minutes:seconds) to 19:13 amongst the neuropathologists. We conclude that the combined use of netcams and WSI is feasible in the intraoperative setting for remote neuropathology specimens.

Copyright © 2014 by the American Association of Neuropathologists, Inc.

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