Familial Epilepsy Associated with Concurrent CHRNB2 Mutation and RBFOX1 Exon Deletion

doi:10.21203/rs.3.rs-2493257/v1

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Familial Epilepsy Associated with Concurrent CHRNB2 Mutation and RBFOX1 Exon Deletion

https://doi.org/10.21203/rs.3.rs-2493257/v1

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Background

Understanding the genetic basis of epilepsy may lead to an improved understanding of the etiology, more precise medical management, and ultimately improved outcomes. It is imperative for patients with epilepsy to obtain a molecular diagnosis, especially when a strong familial epilepsy is discovered.

Case presentation

The proband was a 19 years old female who experienced focal onset seizures, with presenting symptoms of feeling dizzy, disorientation, and passing out. Her EEG studies revealed interictally focal slow and sharp waves in either the left or right hemisphere. EEG monitoring data showed that the seizures arose from the left fronto-temporal regions. Her brain MRI was normal. The proband sister also suffered from focal onset seizures. Her EEG showed focal epileptiform discharge in the right temporal regions, and a subsequent brain MRI was unrevealing. Two genetic tests were conducted for the proband: 1) array CGH revealed 16p13.3 deletion but no 22q deletion; and 2) NGS Epilepsy Panel revealed a few variants of uncertain significance (VUS) including in CHRNB2 (c.1423A >G, p.Ile475Val) and RBFOX1 (exon1-2 deletion). The proband’s sister also carries both the CHRNB2 variant and RBFOX1 deletion. The proband’s father carries the CHRNB2 variant her brother and her mother carry the deletion of RBFOX1.

Conclusions

In this family, the co-expression of the CHRNB2 variant and RBFOX1 deletion may cause the clinical seizures seen in the proband and her sister. It is also possible that the RBFOX1 deletion is associated with an increased risk of seizure disorder with variable expressivity.

focal onset seizure

epilepsy

variants of uncertain significance

CHRNB2

RBFOX1

Epilepsy, a common neurological disorder affecting over 50 million people worldwide, exhibits high heterogeneity by clinical presentation, causative etiologies, triggering factors, etc. Finding a treatable cause of epilepsy is the foundation of precision medicine, which considers individual patient characteristics and offers customized treatment for patients with epilepsy [1, 2]. With advances in genetic sequencing and variant/mutation detecting technology, more and more epilepsy cases were studied. It was estimated that, in 70% of epilepsy cases, genetic factors are responsible either as a single genetic variant in rare epilepsies (epilepsy syndromes) or in multiple genetic variants acting along with different environmental factors as in common epilepsies [3, 4, 5]. Understanding the genetic basis of epilepsy may lead to an improved understanding of the etiology of epilepsy, precision medical management, and ultimately improved outcomes [6]. Such an approach has long been advocated. It is imperative for patients with epilepsy to obtain a molecular diagnosis as early as possible, especially when a strong familial history is discovered [6].

To date, different approaches have been applied to explore the genetic factors in epileptogenesis and to discover new treatment targets [4]. A series of the causative gene(s) for certain epilepsy syndromes have been identified as mutations in some genes and may be the core cause of epilepsy (SCN1A mutations associated with febrile seizures plus syndrome), while other genes may be associated with brain structure lesions and epilepsy (TSC genes cause tuberous sclerosis, the KIRT1 and the CCM2 gene causes cavernous malformations) [4]. Due to the heterogenous phenotypic features of epilepsy involved in changes in the sequence and expression of multiple genes (instead of a single gene), as well as the modification and regulation of genes by multiple factors, gene co-expression network analysis may help to reveal the molecular regulatory mechanisms during epileptogenesis [7]. The utility of genetic testing in peripheral blood samples from patients with epilepsy remains less explored, especially the discovery of variants of uncertain significance (VUS), which often leaves challenges for epileptologists’ clinical practice [8].

Previously variants in CHRNB2 (cholinergic receptor nicotinic beta 2 subunit) have been studied and linked to autosomal dominant nocturnal frontal lobe epilepsy [9]. Partial deletions of the RBFOX1(RNA binding fox-1 homolog 1) gene encoding the neuronal splicing regulator have been reported in a wide range of neurodevelopmental and neuropsychiatric disorders [10]. Our case report describes the discovery of VUS of the CHRNB2 and RBFOX1 genes in a multi-generation family with epilepsy.

A three-generation family was studied. The proband and her sister were studied with electroclinical and radiological phenotyping which incorporated EEG, brain imaging, and molecular genetic analysis. The medical information and genetic tests were obtained from the other family members.

Clinical Characteristics

The proband was a 19 years old female when she experienced her first episodes, she collapsed while dancing. Thereafter, she continued with episodes when she felt dizzy, disoriented, then passed out. The episodes lasted 30–60 seconds. She was amnestic of some her spells. A subsequent cardiac workup with event monitoring and a stress test were unrevealing. Her brain MRI did not reveal a lesional pathology. To further characterize her episodes, she underwent video EEG (vEEG) monitoring. Interictally, focal slow and sharp waves were seen in either left or right centro-parietal regions, or right fronto-central regions. One focal non-impaired seizure was reported and recorded. Clinically the patient felt dizzy, lightheaded, and experienced palpitations. She was briefly confused and not knowing where she was. Then she was able to recall what happened and described the event to her family and providers. Reviewing the video recording, during the episode, she laid back with her eyelids blinking. Then she sat up and rubbed her eyes when the ictal pattern ended. Electroencephalographically, or EEG showed the ictal pattern started as left fronto-temporal rhythmicity, best seen at T3. At the ictal onset, low voltage 4-5Hz rhythmicity was seen in the left temporal regions, followed by modest voltage amplitude 3-4Hz rhythmic delta activity with sharply contoured morphology in the same regions. The frequency then slowed to 3Hz with even higher voltage recordings. The ictal pattern ended with irregular delta activity in the left hemisphere followed by normal awake background recording. The seizure pattern lasted for 30 seconds. Ictally, the heart rate increased from a baseline of 54 bpm to 73 bpm transiently (Fig. 1).

Based on the vEEG morning data, it was confirmed that the proband suffers from focal onset seizures. With further exploration of her family history, the proband sister has childhood onset focal onset seizures, and the proband’s maternal aunt and maternal cousin also have childhood onset seizures with a mixture of focal and generalized seizures (Fig. 2).

The proband’s sister had her first event at 15 years old. Thereafter, she continued experiencing focal onset seizures with or without impaired awareness (often clustering seizures), despite treatments with different anti-seizure medications (ASMs). Her brain MRI was unrevealing. Her EEG showed focal epileptiform discharges in either the left or right temporal regions.

Genetic Testing

Due to a strong family history of epilepsy, we extended the work up with genetic tests. The following genetic test results were discovered in the proband; 1) array CGH (comparative genomic hybridization) revealed 16p13.3 deletion but no 22q deletion, and 2) NGS (next generation sequencing) Epilepsy Panel revealed a few VUS: CHRNB2 (c.1423A > G, p.Ile475Val), RBFOX1 (exon1-2 deletion), SYNJ1, PCLO, PEX10, QARS (Table 1). The discovered genetic variants have not been previously reported in the scientific literature or in the HGMD databases.

With help from medical genetics consultation, genetic tests were obtained from the proband’s immediate family members. The proband’s sister also carries both the CHRNB2 variant and the RBFOX1 deletion. Family studies confirmed the 16p13.3 deletion (including the RBFOX1 deletion) was maternally inherited and the CHRNB2 VUS was paternally inherited; the proband’s father carries CHRNB2 variant, proband’s mother and brother carry the deletion of RBFOX1. None of the carriers (proband father, mother, and brother) have seizures. Another maternal cousin and her son were discovered to have the 22q deletion syndrome with birth organ defects, such information was thought to be irrelevant to the proband and her sister’s epilepsy. The maternal aunt and cousin who have seizures were not tested, as additional family studies were unlikely to add very much information given the revealed inheriting pattern.

VUS Protein Structure Analysis

Since the patient's father is seizure-free, it is unclear if the CHRNB2 VUS is associated with a significant risk for seizures. We applied Alphafold AI (artificial intelligence) and deep learning to investigate if the discovered CHRNB2 gene mutation affects the protein structure. It revealed that the discovered CHRNB2 VUS is not likely to impact the protein structure (Fig. 3), which ratified that such VUS is unlikely to cause familial epilepsy.

CHRNB2 is responsible for the clinical phenotype in some Autosomal Dominant Nocturnal Frontal Lobe Epilepsy (ADNFLE) patients [9]. The electroclinical studies from the proband and her sister did not provide evidence that their seizures were frontal lobe epilepsy. Instead, both were likely suffering from temporal lobe epilepsy. As we predicted, the discovered CHRNB2 VUS is not likely to impact the protein structure – indicative of the fact that CHRNB2 VUS is unlikely pathogenic.

RBFOX1, a neuron-specific splicing factor that exerts both positive and negative regulatory effects on alternative splicing and has been studied and demonstrated associations in several neurodevelopmental and neuropsychiatric disorders including autism spectrum disorder (ASD), developmental delay, epilepsy, ADHD (attention-deficit/hyperactivity disorder), and psychiatric disorders [10, 11]. Within our reported family, it is interesting to note that both sisters and the carriers (proband’s mother and brother) with the discovered RBFOX1 deletion variant do not present with any neuropsychiatric clinical manifestation. Such observation raised the possibility that in this family, that the RBFOX1 deletion may be associated with an increased risk of seizure disorder with variable expressivity.

It is interesting to note that the proband and her sister are the only family members with reported seizures, and they carry both VUS in CHRNB2 and the RBFOX1 deletion. We further explored the possibility of co-expression of VUS in CHRNB2 and the RBFOX1 deletion affecting the phenotype in the proband and her sister.

A recent gene ontology analysis revealed that genes with decreased expression, affected by RBFOX1, are involved in neuronal development, synapse function, and cell differentiation (Table 2). Within the gene members identified, CHRNB2 was noted within the expression network affecting neurogenesis, neuronal development, and synaptic transmission [10]. It is likely that the co-expression of both the CHRNB2 variant and RBFOX1 deletion may have caused the clinical seizures seen in the proband and her sister.

Though the precise causative mechanism of co-expression between RBFOX1 deletion and the VUS in CHRNB2 remains to be further studied, our observation and analysis highlight the need for increasing awareness of and access to genetic testing for adult epilepsy patients with a strong epileptic family history. The VUS are often rare enough that there are no previous reports. Based on family studies and genome-wide association studies, a fair number of VUS might be reclassified [12].

Genetic findings will provide valuable insight into the natural history of epilepsy heterogeneity. Most recently, the awareness of the impact of genetic background on developing drug-resistant epilepsy has gradually increased, and new genetic techniques are proving very useful in clinical practice and may help clinicians to explain the clinical presentation of drug-resistant epilepsy [13]. In addition, this will likely help us understand the impact of personalized and precise pharmacological treatment on epilepsy [14].

EEG: electroencephalogram

VUS: variants of uncertain significance

CGH: comparative genomic hybridization

NGS: next generation sequencing

SCN1A: sodium channel protein type 1 subunit alpha

TSC: Tuberous Sclerosis Complex

KIRT1: KREV interaction trapped 1

CCM2: cerebral cavernous malformations

CHRNB2: cholinergic receptor nicotinic beta 2 subunit

RBFOX1: RNA binding fox-1 homolog 1

Availability of data and materials

The datasets during and/or analyzed during the current study available from the corresponding author on reasonable request.

Funding: The study was not supported by any funding.

Author information

Authors and Affiliations

Math Science Technology Center, Paul Laurence Dunbar High School, Lexington, KY 40513; Tony Xu

Department of Neurology, Kaiser Permanente Sacramento Medical Center, Sacramento, CA 95825;

North Valley Comprehensive Epilepsy Program, Kaiser Permanente Sacramento Medical Center, Sacramento, CA 95825;

Luong Dorris, Ning Zhong

Contributions

LD and NZ collected the clinical information. TX performed protein structure analysis. TX and NZ drafted the manuscript. NZ reviewed related articles and extracted the data. All authors have read and approved the final manuscript.

Corresponding author

Correspondence to Ning Zhong, [email protected]

Ethics declarations

Ethics approval and consent to participate

This study was approved by the Institutional Ethics Committee of Kaiser Permanente Foundation Hospital and written informed consent has been obtained from the patient.

Consent for publication

The written informed consent for publication was obtained from the patient.

Competing interests

The authors declare that they have no competing interests.

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Tables 1 and 2 are available in the Supplementary Files section.

Tables.docx

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Familial Epilepsy Associated with Concurrent CHRNB2 Mutation and RBFOX1 Exon Deletion

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Abstract

Figures

Background

Case Presentation

Clinical Characteristics

Genetic Testing

VUS Protein Structure Analysis

Discussion

Conclusions

Abbreviations

Declarations

References

Tables

Supplementary Files

Status:

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