Nitrogen Permease Regulator Like-2 (NPRL2) truncating mutation causes Ohtahara syndrome with incomplete penetrance: expanding the genotype-phenotype correlations : Clinical Dysmorphology

Journal Logo

Short Case Reports

Nitrogen Permease Regulator Like-2 (NPRL2) truncating mutation causes Ohtahara syndrome with incomplete penetrance: expanding the genotype-phenotype correlations

Zhou, Xinga,*; Chen, Feng-Yingb,*; Ye, Xing-Guanga; Liu, Zhi-Ganga,c

Author Information
doi: 10.1097/MCD.0000000000000428
  • Open

List of key features


Mild downslanting of palpebral fissures

High forehead

Prominent eyebrows

Deep-set eyes

Full lips

Saddle nose

High arched palate


Ohtahara syndrome is one of the most severe and earliest forms of infant epilepsy. It is characterized by tonic spasms, intractable seizures, suppression-burst pattern on the electroencephalogram (EEG), and severe psychomotor retardation (Kato et al., 2007). Previous studies have shown that genetic factors play an important role in the cause of Ohtahara syndrome (Gan et al., 2019; Kang et al., 2019). The established causative genes include PRRT2, KCNQ2, SCN1A, SCN2A, STXBP1, CDKL5, and ARX. In addition, structural anomalies including neuronal migration disorder or dysgenesis are often associated (Song et al., 2021). Mutations in Nitrogen Permease Regulator Like-2 (NPRL2) are being recognized increasingly as a major player in producing brain structural anomalies (Sun et al., 2021). Here, we first report on a familial case with Ohtahara syndrome with a heterozygous nonsense mutation of NPRL2 (NM_006545.5: c.27C>A, p.Cys9Ter) (Fig. 1a and b). Identification of this mutation expands the genotype spectrum of NPRL2.

Fig. 1:
Pedigree of the family, and the electroencephalography (EEG) and imaging data of the patients with NPRL2 mutation. (a) The proband affected with Ohtahara syndrome is indicated by red arrow. Individuals with mutation c.27C>A (p.Cys9Ter) are indicated by m/+, and individuals negative for the mutation are indicated by +/+. (b) Sanger sequencing of the proband and her parents showed c.27C>A mutation in the NPRL2 gene, which was inherited from the father. (c) EEG showed a suppression-burst pattern and recorded epileptic spasms (at the age of 9 days). (d) Brain MRI showed left perisylvian polymicrogyria with an almost complete absence of gyrification and thick cortex (at the age of 10 days). (e) EEG showed hemispheric irregular poly-spikes, poly-sharp waves, and slow waves predominantly in the left central-temporal regions (at the age of 4 months).

Clinical report

The proband is a 5-month-old girl, born to consanguineous parents. She was born at full term following a normal pregnancy. Her birth weight was 3100 gr (25th percentile), length was 47 cm (10th percentile), and head circumference was 32 cm (10th percentile). She started having multiple clusters of flexor epileptic spasms and focal seizures, lasting 20–40 s (occurred around 5–8 times per day) on the third day of life. EEG recordings showed a burst suppression pattern since the 9 days of life (Fig. 1c). Metabolic tests showed normal results. Brain MRI scans showed left perisylvian polymicrogyria with an almost complete absence of gyrification and thick cortex at 10 days (Fig. 1d). Malformation of cortical development (MCD) was diagnosed. Various trials with different medications including sodium valproate, topiramate, and lamotrigine failed to reduce seizures in the first months of life. Vigabatrin was added at 4 months of age at a dose of 40 mg/kg/day with seizure frequency decreased (occurred around 1–3 times per day). Follow-up EEG showed hemispheric irregular poly-spikes, poly-sharp waves, and slow waves predominantly in the left central-temporal regions (at the age of 4 months) (Fig. 1e). Throughout follow-up, the child exhibited severe hypotonia, quadriplegia, absence of spontaneous movements, no acquired developmental milestones, and no eye tracking. On examination, minor dysmorphic facial features were noted, including slight down slanting of palpebral fissures, high forehead, prominent eyebrows, deep-set eyes, saddle nose, full lips, and high arched palate. There were no neurocutaneous markers. Then, we performed whole-exome sequencing of the proband and the parents and found a novel nonsense mutation c.27C>A (p.Cys9Ter) in the NPRL2 gene (Fig. 1a). This mutation was inherited from her unaffected father and was validated by Sanger sequencing (Fig. 1b). It causes truncation of the NPRL2 protein at amino acid position 9 and has not been observed in any of the previously reported Ohtahara syndrome patients. The nonsense mutation c.27C>A of NPRL2 rating was described as predicted null variant in a gene where loss of function is a known mechanism of disease + absent in population databases and was classified as likely pathogenic according to the American College of Medical Genetics and Genomics criteria and guidelines (Richards et al., 2015). Notably, her father’s younger sister and grandfather’s sister also had a history of epilepsy, and her father’s younger sister also had brain structural anomalies, suggesting that they may also carry the same mutation. Unfortunately, their specific diagnosis and treatment were unknown, and we were unable to obtain their peripheral blood samples for sequencing. All the above evidences suggested that this gene is mutated in the paternal line; however, her father and grandfather had no asymptomatic, incomplete penetrance is potentially one of the explanations.


NPRL2 resides on chromosome 3q21.31 and encodes nitrogen permease regulator-like-2 protein (NPRL2). Together with the proteins NPRL3 and DEPDC5, NPRL2 is part of protein complex named GTPase-activating protein activity towards RAGs (GATOR) complex 1 (GATOR1) (Ricos et al., 2016). GATOR1 is a repressor of the mammalian target of rapamycin (mTOR) signaling pathway, and mTOR is a master regulator of cell growth, proliferation, metabolism, migration, protein synthesis, and transcription (Bar-Peled et al., 2013). Pathogenic NPRL2 mutations can cause loss of function of the GATOR1 complex, thereby cause deregulation of mTOR signaling. Deregulation of mTOR signaling has been implicated developmental brain malformations (Crino, 2016), explaining the Ohtahara syndrome associated with MCD caused by NPRL2 truncating mutations.

Previously, a total of 15 NPRL2 variants have been identified in patients featured by focal epilepsies (Sun et al., 2021). We reviewed the phenotypes of all 16 NPRL2 variants reported so far (in the literature and in this study) (Fig. 2). We found that at least 75% of cases have NPRL2 variants inherited from an unaffected parent or a parent with mild phenotype (Table 1), suggesting that incomplete penetrance is a prominent feature of NPRL2-related epilepsy. A total of 11 cases with the results of MRI can be obtained, which showed that three cases carrying nonsense mutations and one case carrying splicing mutation presented with MCD. Among the seven cases with normal brain MRI, two cases carried nonsense mutations and two cases carried splicing mutations. Therefore, the number of existing cases is insufficient to support the association between MCD and null mutations of NPRL2. On the other hand, one case with nonsense mutation, whose MRI findings were normal, had a PET-MRI evocative of right frontoinsular hypometabolism, and histopathologic examination of resected brain tissue was compatible with focal cortical dysplasia Ia, suggesting that PET-MRI may be helpful to detect focal structural abnormalities, especially in patients with null mutations. The onset age of patients with NPRL2-related epilepsy can occur from 3 days to 18 years, and six cases occurred within 1 year old. Oral antiepileptic drugs, including oxcarbazepine, sodium valproate, topiramate and vitamin B6, ketogenic diet, vagus nerve stimulation, and surgery were partly effective. No obvious correlation between the NPRL2 mutations and the mechanisms of action of antiseizure medications was found.

Table 1 - Epilepsy-related NPRL2 variants and their phenotypes
cDNA variant/Protein alteration Inherited Onset age Phenotypes AEDs use Outcome EEG MRI
c.27C>A/p.Cys9* Paternal 3 days 1 IS and FE, 1 Unaffected, 2 UE VPA, VGB, LTG, TPM, currently VPA, VGB, TPM Uncontrolled sz; Global developmental delay Interictal: suppression-burst MCD
c.68_69delTC/p.Ile23Asnfs*6 Paternal 3 years 1 FLE, 1 TLE, 1 ETCS, 2 Unaffected CBZ, VPA VGB, LTG, TPM,LEV, LCS, CLZ After right frontoorbital resection at 18 y: > 50% sz reduction Interictal: right frontoinsular and frontoorbital onset Normal. PET: right frontoinsularHypometabolism. Histology: FCD
c.100C>T/p.Arg34* Paternal 11 months 2 NFLE VPA, TPM, CBZ, LEV, LTG, PGB, CLB, LCS Drug resistant for 3y; Engel III at 7y after 2nd surgery, improved on VNS; Intellectual disability Interictal: discharges in right cingulate gyrus. FCD in right frontal lobe
c.257delG/p.G86Afs*24 Paternal 10 days 1 IS, 1 Unaffected ACTH,TPM Uncontrolled sz; Global developmental delay Hypsarrhythmia Normal
c.562C>T/p.Gln188* N/A 2 years 1 UE N/A N/A N/A Left superior frontal gyrus FCD
c.883C>T/p.Arg295* Maternal N/A 1 TLE, 1 Unaffected N/A N/A N/A N/A
339 + 2T>C/p.(?) Maternal 7 months 2 FE (IS;FS) vitamin B6 and TPM sz free; Moderate developmental delay Interictal: frequent discharges in the left frontal and central regions Normal
c.683 + 1G>C/p.(?) N/A <1 week 1 AS PB, pyridoxine, VGB, ACTH, BZD Drug resistant for 6 m; sz free after surgery; Intellectual disability Interictal: multifocal epileptiform abnormalities with left predominance FCD in left parieto-tempora
c.933-1G>A/p.(?) Paternal 3 years 11 months 1 TLE, 1 FE OXC sz free Right anterior temporal region/– Normal
c.232C>T/p.Arg78Cys Maternal 18 years 1 TLE,1 UE, 1 Unaffected LTG,VPA Uncontrolled sz Interictal: Lefttemporal slowing and spike-and-wave. Ictal: Lefttemporal onset Normal
c.289G>A/p.Ala97Thr Maternal 8 years 1 FLE, 1 Unaffected VPA sz free Interictal: Rightfrontal discharges Normal
c.314T>C/p.Leu105Pro Maternal N/A 2 NFLE,1 FE, 1 nocturnal TCS, 1 Unaffected N/A N/A N/A N/A
c.329C>G/p.Thr110Ser Maternal N/A 1 TLE, 1 Unaffected N/A N/A N/A N/A
c.640G>C/p.Asp214His N/A N/A 1 FLE N/A N/A N/A N/A
c.949G>A/p.Gly317Arg Paternal < 1y 2 UE N/A N/A N/A N/A
c.1134C>G/p.Cys378Trp N/A 3 years 8 months 1FLE CBZ, LEV, CLB, VPA, LCS, OXC, LTG, PHT, TPM, ketogenic diet Drug resistant for 14y (weekly nocturnal sz, improvement on ketogenic diet) Interictal: right frontal epileptiform activity. Ictal (subclinical): right fronto-central sharp waves, activated by sleep Normal
AEDs, antiepileptic drugs; AS, asymmetric spasms; ACTH, adrenocorticotrophic hormone; CBZ, carbamazepine; CLB, clobazam; CLZ, clonazepam; ETCS, epilepsy with tonic–clonic seizures; ECSWS, epileptic encephalopathy with continuous spike-and-wave in slow-wave sleep; ETX, ethosuximide; FEVF, focal epilepsy with variable foci; FLE, frontal lobe epilepsy; FS, febrile seizures: FE, focal epilepsy; FCD, focal cortical dysplasia; GBP, gabapentin; IS, infantile spasm; LCS, lacosamide; LEV, levetiracetam; LTG, lamotrigine; MCD, malformations of cortical development; N/A, not available; NFLE, nocturnal frontal lobe epilepsy; OXC, oxcarbazepine; PB, phenobarbital; PHT, phenytoin; PMP, perampanel; PGB, gregabalin; SUDEP, sudden unexplained death in epilepsy patients; SWC, spike-wave complexes; sz, seizures; TCS, tonic–clonic seizures; TLE, temporal lobe epilepsy; TPM, topiramate; UE, unclassified epilepsy; VGB, vigabatrin; VPA, valproic acid; VNS, vagus nerve stimulation; ZNS, zonisamide.

Fig. 2:
Schematic diagram of NPRL2 domain, and the position of the 16 NPRL2 mutations and corresponding phenotypes. Malformation of cortical development (MCD) and normal MRI associated mutations were colored red and green, respectively.

In summary, this is the first report of Ohtahara syndrome caused by NPRL2 mutation, expanding the genotype spectrum of NPRL2. Infants with a diagnosis of Ohtahara syndrome, genetic testing for NPRL2 variants is recommended.


The authors are deeply grateful to the patients and clinicians who participated in this work. This work was supported by Foshan Science and Technology Bureau (Grant Nos. 2020001003419).

Conflicts of interest

There are no conflicts of interest.


Bar-Peled L, Chantranupong L, Cherniack AD, Chen WW, Ottina KA, Grabiner BC, et al. (2013). A tumor suppressor complex with GAP activity for the Rag GTPases that signal amino acid sufficiency to mTORC1. Science 340:1100–1106.
Crino PB (2016). The mTOR signalling cascade: paving new roads to cure neurological disease. Nat Rev Neurol 12:379–392.
Gan J, Cai Q, Galer P, Ma D, Chen X, Huang J, et al. (2019). Mapping the knowledge structure and trends of epilepsy genetics over the past decade: a co-word analysis based on medical subject headings terms. Medicine (Baltimore) 98:e16782.
Kang KW, Kim W, Cho YW, Lee SK, Jung KY, Shin W, et al. (2019). Genetic characteristics of non-familial epilepsy. PeerJ 7:e8278.
Kato M, Saitoh S, Kamei A, Shiraishi H, Ueda Y, Akasaka M, et al. (2007). A longer polyalanine expansion mutation in the ARX gene causes early infantile epileptic encephalopathy with suppression-burst pattern (Ohtahara syndrome). Am J Hum Genet 81:361–366.
Richards S, Aziz N, Bale S, Bick D, Das S, Gastier-Foster J, et al.; ACMG Laboratory Quality Assurance Committee (2015). Standards and guidelines for the interpretation of sequence variants: a joint consensus recommendation of the American college of medical genetics and genomics and the association for molecular pathology. Genet Med 17:405–424.
Ricos MG, Hodgson BL, Pippucci T, Saidin A, Ong YS, Heron SE, et al.; Epilepsy Electroclinical Study Group (2016). Mutations in the mammalian target of rapamycin pathway regulators NPRL2 and NPRL3 cause focal epilepsy. Ann Neurol 79:120–131.
Song TY, Deng J, Fang F, Chen CH, Wang XH, Wang X, et al. (2021). [The etiology of 340 infants with early-onset epilepsy]. Zhonghua Er Ke Za Zhi 59:387–392.
Sun Y, Wan L, Yan H, Li Z, Yang G (2021). Phenotypic and genotypic characterization of NPRL2-related epilepsy: two case reports and literature review. Front Neurol 12:780799.
Copyright © 2022 The Author(s). Published by Wolters Kluwer Health, Inc.