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Next-Gen Genomics Hit Lymphoma Diagnosis, Therapy

Fuerst, Mark L.

doi: 10.1097/01.COT.0000470861.57351.c1


NEW YORK—The application of next-generation sequencing technologies has established the genomic landscape of lymphoid malignancies. Recurrent genomic alterations have a major impact on the clinical management of lymphoid malignancies by refining diagnosis and prognosis. As discussed here at the “Modern Radiation for Lymphoma: Updated Role and New Rules” meeting at Memorial Sloan-Kettering Cancer Center, this has led to predictions of therapy response and the development of therapeutic targets.

Genomic studies have had an impact on the diagnosis of lymphoma, said Ahmet Dogan, MD, PhD, Chief of the Hematopathology Service at MSKCC—and there have been some recent updates.

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Hot Issues in Lymphoma Pathology

Mediastinal grey zone lymphoma (MGZL) was recognized in 2008 by the World Health Organization, defined as having “poorly defined criteria, including cases that do not fulfill the morphological and/or phenotypic criteria for primary mediastinal B-cell lymphoma (PMBL) or classical Hodgkin lymphoma (HL) in the mediastinum, but that do exhibit transitional features between these two entities.”

The fairly new concept of grey zone lymphoma has recently been refined. It predominantly affects young adult males and shares many immunophenotypic and genetic features with PMBL and classical HL. “A distinct methylation profile suggests a distinct biology, with an aggressive course with clinical outcome worse that both classical HL and PMBL,” he said.

In addition, genetic changes in classical HL, MGZL, and PMBL have also all been noted. Three major biological processes are altered:

  • The JAK STAT pathway, including JAK2 and STAT6;
  • Immune regulatory genes, such as programmed death-ligand 1 (PD-L1) and PD-L2; and
  • The NFkB pathway.

“FISH studies note amplification of JAK2, and more recently, researchers have developed an elegant association by isolating Hodgkin and Reed/Sternberg cells,” Dogan said.

In a discussion of aggressive B-cell lymphomas, he highlighted Burkitt lymphoma. Classical disease includes MYC translocation and several mutations, while variants are MYC translocation-negative and have ID3 mutations, including Burkitt lymphoma-like lymphoma with 11q aberrations.

High-grade B-cell lymphomas, with MYC and BCL2 or BCL6 rearrangements (double hit), contain the morphology of diffuse large B-cell lymphoma (DLBCL) or B-cell lymphoma unclassifiable (BCL-U). This excludes other histologies with a “double hit,” he said. High-grade B-cell lymphoma NOS (not otherwise specified) have no double hit BCL-U or other aggressive cytologies.

For anaplastic large cell lymphoma (ALCL) ALK-positive (ALK+), new fusion partners have been identified, as well as morphological variants. ALCL, ALK+ disease generally appears in the second or third decades, and can be localized or systemic. Histology is sinusoidal or paracortical, with “hallmark cells,” classic, small cell, lymphohistiocytic, and Hodgkin-like variants, he said. The immunophenotype is CD30+, ALK+, loss of other T-cell markers, with a good prognosis.

Of the morphologic variants of ALCL ALK+, 60 percent of patients have the common type variant. All of the patients with this variant are CD30-positive (CD30+).

ALCL ALK-negative (ALK-) disease includes breast implant-associated ALCL and disease with new genetic risk predictors. Among patients with ALCL, ALK- disease, they are generally in their sixth or seventh decades and have systemic disease. “Histology is identical to the classical variant of ALCL-ALK+,” he said. The immunophenotype includes CD30+, ALK-, and loss of other T-cell markers. The prognosis is worse than ALCL, ALK+ patients, but better than those with peripheral T-cell lymphoma NOS.

He noted that there are four genetic subtypes of ALCL, ALK-, including ALK rearrangement, DUSP22 rearrangement, P63 rearrangement, and triple negative. DUSP22 is found in one-third of cases. “With FISH screening, we know that DUSP22-positive patients have better outcomes than DUSP-negative patients. We can use this as a predictor of outcome,” Dogan said.

Using genomic profiling, a set of mutations has emerged in T-cell lymphoma. Disrupted JAK/STAT cytokine signaling pathways in ALCL, ALK- leads to multiple JAK STAT mutations—in particular, JAK1 and STAT3. “These are potential therapeutic targets, with a number of therapies available,” he said.

Summing up, he said: “Next-generation sequencing technologies have redefined the genomic alterations in lymphoma. This new knowledge has led to refinement of diagnostic criteria, enhanced prognostic prediction, and identified molecular alterations that could be targeted for therapy. Targeted genomic profiling of tumor for recurrent mutations of clinical value should be considered as part of standard care.”



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Molecular Pathogenesis of B-cell Lymphoma

In a talk on the molecular pathogenesis of B-cell lymphoma, Ricardo Dall-Favera, MD, Professor & Director of the Institute for Cancer Genetics and Professor of Pathology at Columbia University, noted that diffuse large B-cell lymphoma, the most common and aggressive subtype of B-cell non-Hodgkin lymphoma (NHL) in adults, has marked heterogeneity in morphologic, molecular, and clinical features, and that several phenotypic subgroups have been recognized by gene-expression profile studies.

Such analysis has identified two distinct molecular subgroups of DLBCL; and copy number analysis and whole exome capture and next-generation sequencing have led to gene-expression profiling of about 100 DLBCL primary biopsies. Multiple recurrently mutated genes in DLBCL have been identified, with common and distinct pathways in DLBCL subtypes.

Follicular lymphoma (FL), the second most common type of B-cell NHL, is characterized by a pattern of relapses with decreasing sensitivity to therapy. Histological transformation to more aggressive malignancies occurs in approximately 20 to 60 percent of patients, with survival post-transformation of less than two years, Dall-Favera said.

An experimental approach has used copy number analysis and whole exome capture and next-generation sequencing to identify recurrently mutated genes to compare unselected FL cases with de novo DLBCL.

There are still open questions about FL transformation, he said:

  • What is the history of clonal evolution during transformation (mostly this is a divergent evolution pattern);
  • What are the molecular determinants of FL transformation (chromatin modification and apoptosis are involved, and in transformed FL, there is a cell cycle, proliferation, and DNA damage response); and
  • Is transformed FL a distinct entity separate from de novo DLBCL (it is more similar to germinal center B-cell DLBCL, but has a unique genetic profile, and most FL and transformed FL derive from a common mutated precursor cell through divergent clonal evolution)?


“In 10 years we will recognize more DLBCL,” Dall-Favera predicted. “We will fully understand how to target the disease once we understand its pathogenesis. The good news is we know that this cancer is a complex, multistep genetic disease with more than 110 potentially functioning mutations, and cellular pathways will help define why some of these lesions occur together.”

Some genetic lesions are acquired or selected at transformation, he continued. For example, beta-2 microglobulin (B2M) is an invariant subunit of class I human leukocyte antigen (HLA I). “HLA-I complex displays peptides of endogenous origin on the cell surface and plays an important role in immune recognition.”

The B2M gene is a target of genetic lesions in DLBCL and transformed FL, and defects in B2M expression are associated with the lack of cell surface HLA-I.

A new model shows the evolution of FL to high-grade DLBCL. Resistance to apoptosis and epigenetic reprogramming lead to cell cycle deregulation, DNA damage response, enhanced proliferation, and immune evasion, he said.

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Immunological Modulation

Tim Illidge, PhD, Professor of Targeted Therapy and Oncology at the Institute of Cancer Sciences of Manchester University in the UK, said: “Cancer immunotherapy has finally arrived, and we are exploiting immune checkpoint inhibitors.”

A key mechanism of immune evasion is direct inhibition of cytotoxic T cells, with T-cell activation a two-step process: antigen recognition, followed by the generation of an antigen-independent co-regulatory signal that determines whether the T cell will be switched on or off in response to the antigen, he said. This second step is overseen by the immune checkpoint pathways, which are either stimulatory or inhibitory.”

In describing the importance of the programmed death 1 (PD 1)/PD-L1 axis in Hodgkin lymphoma, he noted that preclinical studies suggest that Reed-Sternberg cells exploit the PD-1 pathway to evade immune detection. In classical HL, alterations in chromosome 9p24 1 increase the abundance of the PD-1 ligands.



“PD-L1 and PD-L2 promote their induction through JAK-STAT signaling. The rationale for therapy is that anti-PD-1-blocking antibody could inhibit tumor immune evasion in patients with relapsed or refractory HL,” Illidge said.

A handful of therapeutic agents that target the PD-1/PD-L pathway are now being tested. “PD-1 pathway blockade is promising for the future of HL therapy,” he said.

“The increasing success for immune checkpoint blockade strategies in anticancer therapies has led to a massive expansion of clinical trials. Monoclonal antibodies targeting PD-1/PD-L1 are likely to make an impact on hematologic malignancies, particularly HL. Well-designed clinical trials with associated translational immunological research are the key to improving outcome for patients.”

Copyright © 2015 Wolters Kluwer Health, Inc. All rights reserved.
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