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Targeted gene panel and genotype-phenotype

correlation in children with developmental and

epileptic encephalopathy

Ara Ko

Department of Medicine

The Graduate School, Yonsei University

[UCI]I804:11046-000000516567[UCI]I804:11046-000000516567




Ara Ko






Directed by Professor Hoon-Chul Kang

The Master's Thesis

submitted to the Department of Medicine,

the Graduate School of Yonsei University

in partial fulfillment of the requirements for the degree of Master of

Medical Science

Ara Ko

June 2018

ACKNOWLEDGEMENTS

I’d like to show my appreciation to my thesis advisor, Dr. Hoon-Chul

Kang for directing me through this work and being my role model as a

doctor, researcher, and educator.

For everything else, I thank my parents for literally everything.

<TABLE OF CONTENTS>

ABSTRACT ····································································· 1

I. INTRODUCTION ···························································· 2

II. PATIENTS AND METHODS ·············································· 2

1. Patients ···································································· 2

2. Targeted NGS gene panel ·············································· 3

3. Clnical characteristics ··················································· 3

4. Statistical analysis ······················································· 4

III. RESULTS ·································································· 4

1. Demographics and general characteristics ························· 4

2. Genotype-phenotype correlations ···································· 7

3. SCN1A ··································································· 8

4. CDKL5 ··································································· 8

5. CHD2 ···································································· 9

6. KCNQ2 ··································································· 9

7. STXBP1 ·································································· 10

8. SCN2A ··································································· 10

9. SCN8A ··································································· 10

10. SYNGAP1 ······························································ 11

IV. DISCUSSION ······························································ 13

V. CONCLUSION ····························································· 15

REFERENCES ································································· 16

APPENDICES ·································································· 19

ABSTRACT (IN KOREAN) ················································ 28

PUBLICATION LIST ························································ 29

LIST OF FIGURES

Figure 1. Age distribution of seizure onset in 103 patients with

identified pathogenic mutations ··································· 6

Figure 2. Positivity rate of targeted gene-panel sequencing for

developmental and epileptic encephalopathy according to

epilepsy syndromes ·················································· 6

Figure 3. Genotype-phenotype correlations ····················· 7

Figure 4. Age of seizure onset according to genes identified with

pathogenic mutations ················································ 9

LIST OF TABLES

Table 1. Identified pathogenic mutations ························ 5

Table 2. Demographic characteristics of patients with develop-

mental and epileptic encephalopathy and comparison between

patients with positive and negative results from targeted gene-

panel sequencing ······················································ 5

Table 3. Clinical characteristic of the eight most frequently iden-

tified genotypes ························································ 12

1

ABSTRACT

Targeted gene panel and genotype-phenotype correlation in children with

developmental and epileptic encephalopathy

Ara Ko



(Directed by Professor Hoon-Chul Kang)

We performed targeted gene-panel sequencing for children with developmental and

epileptic encephalopathy (DEE) and evaluated the clinical implications of

genotype–phenotype correlations.

We assessed 278 children with DEE using a customized gene panel that included 172

genes, and extensively reviewed their clinical characteristics, including therapeutic

efficacy, according to genotype.

In 103 (37.1%) of the 278 patients with DEE, 35 different disease-causing monogenic

mutations were identified. The diagnostic yield was higher among patients who were

younger at seizure onset, especially those whose seizures started during the neonatal

period, and in patients with drug-resistant epilepsy. According to epilepsy syndromes,

the diagnostic yield was the highest among patients with West syndrome (WS) with a

history of neonatal seizures and mutations in KCNQ2 and STXBP1 were most frequently

identified. On the basis of genotypes, we evaluated the clinical progression and seizure

outcomes with specific therapeutic regimens; these were similar to those reported

previously. In particular, sodium channel blockers were effective in patients with

mutations in KCNQ2 and SCN2A in infancy, as well as SCN8A, and interestingly, the

ketogenic diet also showed diverse efficacy for patients with SCN1A, CDKL5, KCNQ2,

STXBP1, and SCN2A mutations. Unfortunately, quinidine was not effective in 2 patients

with migrating focal epilepsy in infancy related to KCNT1 mutations.

Targeted gene-panel sequencing is a useful diagnostic tool for DEE in children, and

genotype–phenotype correlations are helpful in anticipating the clinical progression and

treatment efficacy among these patients.

----------------------------------------------------------------------------------------

Key words: epileptic encephalopathy, NGS, genotype-phenotype

2

Targeted gene panel and genotype-phenotype correlation in children with

developmental and epileptic encephalopathy

Ara Ko



(Directed by Professor Hoon-Chul Kang)

I. INTRODUCTION

Epileptic encephalopathy refers to a group of severe pediatric epilepsies in which

epileptic activities contribute to cognitive delay or regression.1 Recently, developmental and

epileptic encephalopathy (DEE) was introduced as a new concept, because cognitive

deterioration can also derive from genetic etiologies, irrespective of epileptic activity.1 With

advances in sequencing methods, monogenic mutations responsible for DEE have been

identified, suggesting underlying genetic etiologies in DEE patients who were previously

included in the unknown etiology group.2 Advanced sequencing methods have shortened the

diagnostic process, and potential benefits from gene-based determination of clinical progression

and therapeutic regimens might be expected in this era of emerging precision medicine.

Therefore, here, we sought to elaborate our single-center experience of using targeted

gene-panel sequencing to diagnose the genetic etiology of DEE and to reveal the clinical

implications of gene-panel studies by extensively reviewing genotypephenotype correlations in

such patients.

II. PATIENTS AND METHODS

II-1. Patients

A total of 280 unrelated pediatric patients with early-onset DEE of unknown etiology

were recruited from the epilepsy clinic of Severance Children’s Hospital between March 2015

and June 2017. All patients met the following criteria: (1) seizure onset before the age of 3

years; (2) multiple epileptiform discharges with severely disorganized background activity on

electroencephalography (EEG); (3) progressive developmental deterioration or a known

developmental and epileptic encephalopathy syndrome; (4) no significant structural lesion

detected on brain magnetic resonance imaging; (5) no metabolic abnormalities; and (6) no

abnormalities detected on previous genetic tests. This study was approved by the Institutional

3

Review Board of Severance Hospital.

II-2. Targeted NGS gene panel

A total of 172 genes known to be related to DEE were included in our gene panel; the

genes are listed in Appendix 1. Briefly, the processes for analyzing the sequencing data were as

follows. Genomic DNA was extracted from the leukocytes of whole-blood samples using the

QIAamp Blood DNA mini kit (Qiagen, Hilden, Germany). The pooled libraries were sequenced

using a MiSeq sequencer (Illumina, San Diego, CA, USA) and the MiSeq Reagent Kit v2 (300

cycles). Sequencing data were aligned against appropriate reference sequences and analyzed

using Sequencher 5.3 software (Gene Codes Corp., Ann Arbor, MI, USA).

Parental studies were performed by Sanger sequencing on a 3730 DNA Analyzer with

the BigDye Terminator v3.1 Cycle Sequencing kit (Applied Biosystems, Foster City, CA, USA)

if needed. Large exonic deletions and duplications were confirmed using the MLPA kit (MRC

Holland). Classification of the variants followed a three-step approach: (1) conventional

bioinformatics analysis, based on the nature of the mutation and frequencies in the normal

population; (2) in silico analysis, with a literature review of the same variant and other variants

in the same amino acid position; (3) a consensus discussion of genotypephenotype correlations

between geneticists and epileptologists, along with family studies and other confirmatory assays.

Subsequently, the variants determined to be pathogenic or to be likely pathogenic on the basis of

the American College of Medical Genetics and Genomics and the Association for Molecular

Pathology classification were considered as causative mutations for DEE.3

II-3. Clinical characteristics

We reviewed the clinical features of DEE patients investigated using targeted

gene-panel sequencing. The clinical characteristics included demographic profiles, classification

of epilepsy syndromes, and seizure outcomes after therapeutic regimens and diet therapy. For

epilepsy syndromes, the patients were classified according to the 2010 International League

Against Epilepsy classification.4 If the patient’s syndromic diagnoses changed over time, the

first syndromic diagnosis was selected. Patients who developed seizures during the neonatal

period and could not readily be categorized according to a syndrome but later developed West

syndrome (WS) were grouped separately as “WS with neonatal seizures.” Drug-resistant

epilepsy was defined as failure to achieve seizure freedom after adequate trials with 2

antiepileptic drugs. With regard to the efficacy of the therapeutic regimens, including diet

therapy, we considered the regimens as effective if they contributed to more than 50% decrease

in the seizure frequency from the baseline.

4

II-4. Statistical analysis

Data from statistical analyses are expressed as medians and interquartile ranges (IQRs)

for continuous and ordinal variables, and as counts and percentages for categorical variables.

The 2 groups were compared using the Chi-square test or Fisher’s exact tests for categorical and

ordinal data, or the Mann–Whitney U-test for non-parametric continuous data. A p value of <

0.05 was considered significant. The Statistical Package for the Social Sciences (version 23.0;

SPSS Inc., Chicago, IL, USA) was used for all analysis.

III. RESULTS

III-1. Demographics and general characteristics

Among the 278 patients (268 Koreans, 1 Mongolian, and 9 Caucasians),

pathogenic monogenic mutations were identified in 103 (37.1%). Thirty-five different causative

genes were found, with SCN1A being the most frequent (n = 11, 10.7%), followed by CDKL5 (n

= 9, 8.7%), CHD2 (n = 8, 7.8%), KCNQ2 (n = 7, 6.8%), STXBP1 (n = 7, 6.8%), SCN2A (n = 5,

4.9%), SCN8A (n = 5, 4.9%), SYNGAP1 (n = 5, 4.9%), and others (Table 1). Thirty-four (33.0%)

of 103 patients with identified mutations carried variants that had not been previously reported,

which were therefore confirmed to be de novo mutations by trio sequencing. All pathogenic

variants detected in this study are listed in the Appendix 2.

When the 103 patients with identified mutations were compared with the 175 patients

who showed negative findings in the gene panel screen, the age of seizure onset was

significantly earlier (p = 0.039) and the proportion of patients showing drug-resistant epilepsy

was significantly larger (p < 0.001) among patients with identified mutations (Table 2). After

controlling with age at seizure onset, presence of drug-resistant epilepsy, sex, and epilepsy

syndrome, age of seizure onset (OR 0.977, 95% CI 0.957-0.996, p = 0.019) and presence of

drug-resistant epilepsy (OR 3.036, 95% CI 1.544-5.970, p = 0.001) were still significant

predictors of achieving genetic diagnosis with gene panel study. The diagnostic yield was

highest in patients with seizure onset during the neonatal period, with 80.0% (20 of 25) of

patients proven to have causative mutations. This was significantly higher when compared to

the patients whose seizures began after 30 days of age who shows a diagnostic yield of 32.8% (p

< 0.001). The proportions of identified gene mutations decreased as the age at seizure onset

increased (Figure 1).

5

Table 1. Identified pathogenic mutations (n = 103, 35 genes)

Pathogenic gene n (%) Pathogenic gene n (%)

ALDH7A1 2 (1.9) KCNT1 3 (2.9)

ARX 1 (1.0) MECP2 5 (4.9)

BRAT1 3 (2.9) PCDH19 3 (2.9)

CACNA1A 1 (1.0) PRODH 1 (1.0)

CACNB4 1 (1.0) SCN1A 11 (10.7)

CASK 1 (1.0) SCN1B 1 (1.0)

CDKL5 9 (8.7) SCN2A 5 (4.9)

CHD2 8 (7.8) SCN3A 1 (1.0)

DNM1 2 (1.9) SCN8A 5 (4.9)

EEF1A2 2 (1.9) SLC6A1 3 (2.9)

GNAO1 1 (1.0) SLC9A6 2 (1.9)

GRIN2A 1 (1.0) STXBP1 7 (6.8)

HCN1 1 (1.0) SYN1 1 (1.0)

IQSEC2 1 (1.0) SYNGAP1 5 (4.9)

KANSL1 1 (1.0) UBE3A 2 (1.9)

KCNA1 1 (1.0) WWOX 1 (1.0)

KCNB1 2 (1.9) ZEB2 2 (1.9)

n, number

Table 2. Demographic characteristics of patients with developmental and epileptic

encephalopathy and comparison between patients with positive and negative results from

targeted gene-panel sequencing

Total

(n = 278)

Identified pathogenic

mutations (n = 103)

Negative results

(n = 175)

p

Age at seizure

onset (months)

7 (318) 6 (218) 7 (419) 0.039

Sex (male) 155 (55.8%) 51 (49.5%) 104 (59.4%) 0.108

Drug-resistant

epilepsy

161 (57.9%) 74 (71.8%) 87 (49.7%) < 0.001

Prior genetic

tests

107 (38.5%) 43 (41.7%) 64 (36.6%) 0.392

Data are presented as median (interquartile range) or number (percentage).

6

Figure 2. Positivity rate of targeted gene-panel sequencing for developmental and

epileptic encephalopathy according to epilepsy syndromes.

(WS, West syndrome; EIMFS, epilepsy of infancy with migrating focal seizures; GE,

generalized epilepsy; EMAS, epilepsy with myoclonic atonic seizures; LGS,

Lennox-Gastaut syndrome; LKS, Landau-Kleffner syndrome)

Figure 1. Age distribution of seizure onset in 103 patients with identified pathogenic

mutations (m, months). The right pie chart shows seizure onset age distribution of 0-12m

group in more detail.

7

III-2. Genotype-phenotype correlations

The 278 patients could be classified into 10 groups according to the epilepsy

syndromes; 119 (42.8%) were classified as showing WS. The diagnostic yield of the gene-panel

study differed significantly among patients with the different epilepsy syndromes (p < 0.001,),

with the highest diagnostic yield in patients with WS with neonatal seizures (100.0%), followed

by those with Ohtahara syndrome (85.7%), and others (Figure 2).

Figure 3 shows the range of ages at seizure onset in our patients and the mutations

identified in each epilepsy syndrome. For patients with WS with neonatal seizures, KCNQ2 and

STXBP1 were the most frequently identified disease-causing genes; for those with Ohtahara

syndrome, KCNQ2; for epilepsy of infancy with migrating focal seizures (EIMFS), KCNT1; for

WS, CDKL5; for Dravet syndrome, SCN1A; for Lennox-Gastaut syndrome (LGS), SYNGAP1;

for epilepsy with myoclonic atonic seizures (EMAS), SLC6A1; for unspecified generalized

epilepsy, CDH2; and for unspecified focal epilepsy, PCDH19.

Figure 3. Genotype-phenotype correlations. The bar indicates the range of seizure onset

age (months) observed in our cohort for each syndrome, and genes identified to have

disease-causing mutations for each syndrome are listed in an order of decreasing frequency.

The numbers before the genes indicate the identified frequency for each gene.

(WS, West syndrome; EIMFS, epilepsy of infancy with migrating focal seizures; LGS,

Lennox-Gastaut syndrome; EMAS, epilepsy with myoclonic atonic seizures; GE,

generalized epilepsy; FE, focal epilepsy)

8

Age of seizure onset for each genotype was relatively consistent, and the median onset

age of seizures was 3 days in patients with KCNQ2-related encephalopathy; 7 days in

STXBP1-related conditions; 1 month in KCNT1-related conditions; 3 months in CDKL5-,

SCN8A-, and BRAT1-related conditions; and 6 months in SCN1A-related conditions. The age at

seizure onset in patients with CHD2 and SYNGAP1 encephalopathy was relatively high, with a

median seizure onset age of 19 and 26 months, respectively (Figure 4).

The details of the clinical characteristics, including seizure outcomes according to

therapeutic regimens, are described below.

III-3. SCN1A (Sodium channel, neuronal type 1, alpha subunit)

Eleven patients were identified with mutations in SCN1A (sodium channel, neuronal

type 1, alpha subunit gene). A summary of the clinical characteristics are shown in Table 3. All

patients showed phenotypes that were concordant with Dravet syndrome. These 11 patients

comprised 61.1% of those with total Dravet syndrome (n = 18), with another 6 patients without

identified pathogenic variants and 1 with a mutation in SCN1B.

III-4. CDKL5 (Cyclin-dependent kinase-like 5)

Nine patients had mutations in the CDKL5 (cyclin-dependent kinase-like 5 gene) in our

study. Five patients presented with tonic seizures first, and shortly afterwards developed spasms

and later showed hypsarrhythmia on EEG and were diagnosed with WS. Two patients presented

with spasms with hypsarrhythmia on EEG, and were also diagnosed with WS. The remaining 2

Figure 4. Age of seizure onset according to genes identified with pathogenic mutations.

9

patients who had an earlier seizure onset age (15 and 50 days, respectively) presented with tonic

seizures with a burst-suppression pattern on EEG, and were diagnosed with Ohatahara

syndrome; the condition in both evolved into WS at the age of 3 months. All these patients

showed early developmental delay before seizure onset, and showed profound intellectual

disability (ID); 7 (77.8%) were unable to make eye contact or control their heads. All patients

also showed drug-resistant epilepsy, and a ketogenic diet (KD) was attempted in all 9 patients,

but was effective in only 1 patient. In 2 patients who were followed up for more than 10 years,

seizures evolved into LGS. Corpus callosotomy was performed in both of them, which resulted

in 50% and 75% seizure reduction after the surgery, respectively.

III-5. CHD2 (Chromodomain helicase DNA-binding protein 2)

Eight patients showed mutations in CHD2 (chromodomain helicase DNA-binding

protein 2 gene). All patients presented predominantly with myoclonic seizures, and clinical

photosensitivity was observed in 5 patients (62.5%). The EEGs of all these patients showed

generalized epileptiform discharges, and 2 patients were diagnosed with EMAS, 1 with LGS,

and 5 were classified as having unspecified generalized epilepsy. All patients showed normal

development until seizure onset, had their first seizures during childhood at a median age of 19

months, and showed developmental impairment after seizure onset. Five patients, with disease

duration of less than 10 years, currently show mild ID with relatively near normal social

quotients on the social maturity scale. However, 2 patients with longer follow-up periods have

shown continued developmental regression to severe ID. In 6 (75.0%) patients, valproate was

the most effective medication, while 2 (25.0%) patients have drug-resistant epilepsy.

III-6. KCNQ2 (Potassium channel, voltage-gated, KQT-like subfamily, member 2)

Seven patients were found to harbor mutations in KCNQ2 (potassium channel, voltage-gated,

KQT-like subfamily, member 2 gene). Patients with KCNQ2 encephalopathy showed the earliest

seizure onset, and all patients had seizure onset within the first 9 days of life. All patients

presented with tonic seizures, but EEGs of 4 patients showed a burst-suppression pattern, while

the other 3 patients showed focal epileptiform discharges. The former 4 patients were diagnosed

with Ohtahara syndrome, and the condition in all 7 patients later evolved into WS. All patients

showed profound ID, with none showing any signs of development. All patients had

drug-resistant epilepsy, but the KD had favorable responses in 6 patients. Sodium channel

blockers were administered in 4 patients and these were effective in controlling their seizures.

10

III-7. STXBP1 (Syntaxin-binding protein 1)

Seven patients had mutations in STXBP1 (syntaxin-binding protein 1 gene). The

median seizure onset age was 7 days, which was relatively early. Five patients presented with

focal or generalized seizures during the neonatal period, 2 were diagnosed with Ohtahara

syndrome with a burst-suppression pattern on EEG, and 3 showed focal epileptiform discharge.

The remaining 2 patients with a later seizure onset age, at 2 and 8 months, presented with

spasms, hypsarrhythmia on EEG, and were diagnosed with WS. The condition in the former 5

patients also evolved into WS. Seizures were drug resistant in half of the patients, and various

degrees (mild to severe) of ID were observed. Ketogenic diet was effective in seizure reduction

in all 4 patients who tried the dietary therapy.


Five patients had pathogenic variants in SCN2A (sodium channel, neuronal

type 2, alpha subunit gene). The age of seizure onset had a bimodal distribution; 3 patients had

seizure onset at 1 day, 2 patients at 5 months, and 2 patients at 30 and 36 months each. The first

3 patients presented with Ohtahara syndrome and WS, which later evolved into LGS with a

significant developmental impact (3 were unable to make eye contact or control their heads).

Two patients with seizure onset at 1 day and 5 months, respectively, were given sodium channel

blockers and showed significant seizure reduction. One patient with seizure onset age of 5

months was not tried with sodium channel blockers. The other 2 patients with later seizure onset

age of 30 and 36 months were both diagnosed with LGS and unspecified focal epilepsy. Both

patients showed normal development before seizure onset, and regressed thereafter, currently

showing mild ID. Sodium channel blockers were administered to both patients without effect

and were discontinued.


Five patients were identified with pathogenic variants in SCN8A (sodium

channel, neuronal type 8, alpha subunit gene). One patient presented with neonatal seizures that

later developed into WS, and the other 4 all presented with WS. Later, the condition in all 5

patients evolved into LGS, and the patients showed significant developmental delay. Three

patients (60.0%) had drug-resistant epilepsy; sodium channel blockers were administered to 4

patients, with a favorable response in 3 (75.0%) patients.

11

III-10. SYNGAP1 (Synaptic Ras-GTPase-activating protein 1)

Five patients had disease-causing variants in SYNGAP1 (synaptic

Ras-GTPase-activating protein 1 gene). The median seizure onset age was 26 months, which

was relatively late. All patients had generalized seizures, such as myoclonic, atonic, or atypical

absence seizures, with EEGs showing predominantly generalized epileptiform discharges. Four

patients were diagnosed with LGS and 1 with EMAS. In 1 (20.0%) patient, seizures were

intractable to medication. All patients showed developmental delay prior to seizure onset, and

currently show severe ID, except for 1 patient with moderate ID.

13

IV. DISCUSSION

We defined 35 different disease-causing monogenic mutations in 37.1% patients with

DEE. These patients had a lower age at seizure onset and had more intractable seizures than

patients in whom mutations were not identified with the gene panel screening.

With the advent of sequencing methods that enable sequencing of several DNA regions

in a single reaction, there have been significant advances in the identification of epilepsy-related

genes.2 Monogenic epilepsy, in which a single variant with a large effect is considered causative,

is far less common than complex genetic epilepsy, in which a combinatorial effect of multiple

variants is considered causative.5 However, previous studies with NGS gene panels for DEE

have shown substantial diagnostic yields of about 2040%, which is not inferior to the

diagnostic yield of whole exome sequencing.6-16 Some of recent studies include a study with 175

Chinese patients with early-onset epileptic encephalopathy investigated with gene panel

comprising 17 genes which identified disease-causing variants in 32% of patients, a study with

87 patients with epilepsy and developmental delay investigated with 106 genes gene panel in

which 19.5% of patients were identified with pathogenic variants, a study with 349 patients with

drug-resistant epilepsy and seizure onset before 1 year of age who were investigated with

95-genes gene panel and showed 26.6% of diagnostic yield, and a study with 400 patients with

early-onset seizures and severe developmental delay who were investigated with 46-genes gene

panel and showed 18% of diagnostic yield.13-16 Direct comparisons between studies for factors

influencing diagnostic yields are difficult as cohorts and genes included in the panels are

different, but slightly higher diagnostic yield in our study may be attributable to higher

availability of parental samples (90.1% of 131 patients whose parental samples were needed to

investigate variants with unknown significance) to confirm de novo occurrence of variants. In

41.7% of patients in whom disease-causing variants were identified in this study, results from

previous genetic tests prior to the gene-panel study were uninformative. Therefore, monogenic

variants, especially de novo variants, have an important role in DEE, and at present, targeted

gene-panel sequencing is the most cost-effective diagnostic option for epilepsy patients with

suspected genetic etiology.5,17

Here, the proportion of patients with drug-resistant epilepsy was significantly higher

among patients with identified mutations, which is concordant with the results of previous

reports that showed higher diagnostic yields in cohorts with severe drug-resistant epilepsy.5

These observations have not been explained to date; however, possible reasons are that patients

with somatic mutations occurring only in the brain, those with a low rate of mosaic mutations,

or patients with complex genetic epilepsy obtaining negative results in gene panels, may have

14

less severe symptoms than patients carrying monogenic mutations with large effects. The

seizure onset age was significantly lower in patients with identified mutations. This observation

is also in accord with those of previous studies, which showed that seizures with an earlier age

at onset resulted in higher molecular yields.15,17 The timing of seizure onset for each pathogenic

gene was relatively consistent among patients, as seizures started at the age at which the

expression of a gene with the pathogenic mutation is required for physiological neuronal

development.18 Therefore, patients with pathogenic monogenic variants with strong effects may

develop genetic dysfunctions earlier than patients with presumably somatic mosaicism or a

polygenic disease basis. Naturally, the diagnostic yields were also higher in catastrophic

epilepsy syndromes with earlier seizure onset, such as WS with neonatal seizure, Ohtahara

syndrome, and EIMFS.

Factors accounting for phenotypic pleiotropy can include the type and timing of

mutations, including somatic mosaicism or genomic rearrangements; localization of the

mutations in the protein; the loss- or gain-of-function mechanisms caused by the mutations;

epigenetic factors; and modifier genes.19-24 However, patients with the various pathogenic genes

do share some common features, such as the temporal expression of symptoms and the type of

epilepsy syndromes, as described above. This was more evident in some cases, such as in those

with KCNT1 mutations, where all 3 patients presented with EIMFS, and as in Dravet syndrome,

where patients almost exclusively had SCN1A as the causative gene (91.7% of genetically

diagnosed patients).

The ultimate goal of genetic diagnosis is targeted therapy. However, available therapies

targeted to known genetic mutations are still limited to a few genes, such as KD for SCL2A1,

retigabine for KCNQ2, memantine for GRIN2A or GRIN2B, and quinidine for KCNT1.25-29

Three EIMFS patients in our cohort who had pathogenic variants in KCNT1 showed intractable

seizures, and 2 of these were administered quinidine; this was discontinued for 1 patient before

the therapeutic level was reached due to QT prolongation, while another patient, in whom the

therapeutic serum concentration was reached, did not show any effects in seizure reduction.

Therefore, more studies are warranted for achieving targeted therapy, including reprogramming

of stem cells or gene therapy, based on molecular diagnoses.

In other cases, patients with each genetic mutation in this study showed clinical

characteristics that were similar to those described previously, and also similar responses to

specific therapies. Besides retigabine, sodium channel blockers are also known to be effective in

seizure control in KCNQ2 encephalopathy patients, probably through modulation of the sodium

channel that affects the function of the channel complex including the potassium channel.30,31 In

this study, sodium channel blockers were administered to 4 patients, and were effective but did

15

not completely control the seizures in all 4 patients. Wolff et al. reported that mutations in

SCN2A cause 2 distinct phenotypes: early infantile onset (<3 months) and infantile/childhood

onset (≥3 months) encephalopathies.32 The early infantile form was associated with

gain-of-function mutations, and therefore showed good response to sodium channel blockers,

while the later onset form was associated with a loss-of function and sodium channel blockers

were rarely effective or sometimes worsened the seizures.32 Also, early infantile form were all

identified with missense mutations, while later onset form was associated with both missense

and nonsense mutations, and patients with nonsense mutations all had seizure onset beyond the

first year of life. In our study, the patients could be distinguished into earlier and later onset

groups; earlier onset patients all had missense mutations and showed a good response to sodium

channel blockers, while they were not effective in later onset patients with non-sense mutations,

resulting in discontinuation of the medication. For mutations in SCN8A, most functional

analyses to date have revealed gain-of-function effects, but some variants also produced

loss-of-function effects in vitro.33 In our study, the majority (75.0%) of the patients showed a

good response to sodium channel blockers.

V. CONCLUSION

Monogenic mutations, especially de novo monogenic variants, are an important

underlying etiology for DEE, and targeted gene-panel sequencing is an effective diagnostic tool

for DEE. The diagnostic yield is higher in drug-resistant epilepsy, and in patients with earlier

seizure onset especially during the neonatal period. Although phenotypic pleiotropy exists, we

could confirm correlation of genotypes with the clinical progress and seizure outcomes to

specific therapeutic regimens that were similar to those described in previous studies.

16

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19

APPENDICES

Appendix 1. List of 172 targeted genes included in the developmental and epileptic

encephalopathy panel

Gene OMIM Full name Cytogenetic

location

AARS 601065 ALANYL-tRNA SYNTHETASE 16q22.1

ABAT 137150 4-AMINOBUTYRATE AMINOTRANSFERASE 16p13.2

ACADL 609576 ACYL-CoA DEHYDROGENASE,

LONG-CHAIN

2q34

ACADM 607008 ACYL-CoA DEHYDROGENASE,

MEDIUM-CHAIN

1p31.1

ACADS 606885 ACYL-CoA DEHYDROGENASE,

SHORT-CHAIN

12q24.31

ACY1 104620 AMINOACYLASE 1 3p21.2

ADGRV1 602851 ADHESION G PROTEIN-COUPLED

RECEPTOR V1

5q14.3

ADSL 608222 ADENYLOSUCCINATE LYASE 22q13.1

ALAD 125270 DELTA-AMINOLEVULINATE

DEHYDRATASE

9q32

ALAS2 301300 DELTA-AMINOLEVULINATE SYNTHASE 2 Xp11.21

ALDH4A1 606811 ALDEHYDE DEHYDROGENASE, FAMILY 4,

SUBFAMILY A, MEMBER 1

1p36.13

ALDH7A1 107323 ALDEHYDE DEHYDROGENASE 7 FAMILY,

MEMBER A1

5q23.2

ALG13 300776 ASPARAGINE-LINKED GLYCOSYLATION 13 Xq23

ALPL 171760 ALKALINE PHOSPHATASE, LIVER 1p36.12

AMT 238310 AMINOMETHYLTRANSFERASE 3p21.31

ARHGEF15 608504 RHO GUANINE NUCLEOTIDE EXCHANGE

FACTOR 15

17p13.1

ARHGEF9 300429 RHO GUANINE NUCLEOTIDE EXCHANGE

FACTOR 9

Xq11.1

ARX 300382 ARISTALESS-RELATED HOMEOBOX,

X-LINKED

Xp21.3

ASNS 108370 ASPARAGINE SYNTHETASE 7q21.3

ASPM 605481 ABNORMAL SPINDLE-LIKE,

MICROCEPHALY-ASSOCIATED

1q31.3

ATP13A2 610513 ATPase, TYPE 13A2 1p36.13

ATP6AP2 300556 ATPase, H+ TRANSPORTING, LYSOSOMAL,

ACCESSORY PROTEIN 2

Xp11.4

BRAT1 614506 BRCA1-ASSOCIATED ATM ACTIVATOR 1 7p22.3

BTD 609019 BIOTINIDASE 3p25.1

CACNA1A 601011 CALCIUM CHANNEL,

VOLTAGE-DEPENDENT, P/Q TYPE,

ALPHA-1A SUBUNIT

19p13.13

CACNB4 601949 CALCIUM CHANNEL,

VOLTAGE-DEPENDENT, BETA-4 SUBUNIT

2q23.3

CASK 300172 CALCIUM/CALMODULIN-DEPENDENT

SERINE PROTEIN KINASE

Xp11.4

CASR 601199 CALCIUM-SENSING RECEPTOR 3q13.3-q21.1

20

CBS 613381 CYSTATHIONINE BETA-SYNTHASE 21q22.3

CDKL5 300203 CYCLIN-DEPENDENT KINASE-LIKE 5 Xp22.13

CHD2 602119 CHROMODOMAIN HELICASE

DNA-BINDING PROTEIN 2

15q26.1

CHRNA2 118502 CHOLINERGIC RECEPTOR, NEURONAL

NICOTINIC, ALPHA POLYPEPTIDE 2

8p21.2



20q13.33



15q13.3

CHRNB2 118507 CHOLINERGIC RECEPTOR, NEURONAL

NICOTINIC, BETA POLYPEPTIDE 2

1q21.3

CLCN4 302910 CHLORIDE CHANNEL 4 Xp22.2

CLN3 607042 CLN3 GENE 16p12.1

CLN5 608102 CLN5 GENE 13q22.3

CLN6 606725 CLN6 GENE 15q23

CLN8 607837 CLN8 GENE 8p23.3

CNTNAP2 604569 CONTACTIN-ASSOCIATED PROTEIN-LIKE 2 7q35-q36

COL4A1 120130 COLLAGEN, TYPE IV, ALPHA-1 13q34

CPOX 612732 COPROPORPHYRINOGEN OXIDASE 3q11.2

CPT1A 600528 CARNITINE PALMITOYLTRANSFERASE I,

LIVER

11q13.3

CPT1B 601987 CARNITINE PALMITOYLTRANSFERASE I,

MUSCLE

22q13.33

CPT2 600650 CARNITINE PALMITOYLTRANSFERASE II 1p32.3

CSTB 601145 CYSTATIN B 21q22.3

CTSD 116840 CATHEPSIN D 11p15.5

CTSF 603539 CATHEPSIN F 11q13.2

DNAJC5 611203 DNAJ/HSP40 HOMOLOG, SUBFAMILY C,

MEMBER 5

20q13.33

DNM1 602377 DYNAMIN 1 9q34.11

DOCK7 615730 DEDICATOR OF CYTOKINESIS 7 1p31.3

DYRK1A 600855 DUAL-SPECIFICITY TYROSINE

PHOSPHORYLATION-REGULATED KINASE

1A

21q22.13

EEF1A2 602959 EUKARYOTIC TRANSLATION ELONGATION

FACTOR 1, ALPHA-2

20q13.33

EPM2A 607566 EPM2A GENE 6q24.3

FARS2 611592 PHENYLALANYL-tRNA SYNTHETASE 2,

MITOCHONDRIAL

6p25.1

FECH 612386 FERROCHELATASE 18q21.31

FOLR1 136430 FOLATE RECEPTOR 1, ADULT 11q13.4

FOXG1 164874 FORKHEAD BOX G1 14q12

GABBR2 607340 GAMMA-AMINOBUTYRIC ACID B

RECEPTOR 2

9q22.33

GABRA1 137160 GAMMA-AMINOBUTYRIC ACID

RECEPTOR, ALPHA-1

5q34

GABRB3 137192 GAMMA-AMINOBUTYRIC ACID

RECEPTOR, BETA-3

15q12

GABRG2 137164 GAMMA-AMINOBUTYRIC ACID

RECEPTOR, GAMMA-2

5q34

GAMT 601240 GUANIDINOACETATE

METHYLTRANSFERASE

19p13.3

21

GATM 602360 L-ARGININE:GLYCINE

AMIDINOTRANSFERASE

15q21.1

GCSH 238330 GLYCINE CLEAVAGE SYSTEM H PROTEIN 16q23.2

GLDC 238300 GLYCINE DECARBOXYLASE 9p24.1

GNAO1 139311 GUANINE NUCLEOTIDE-BINDING

PROTEIN, ALPHA-ACTIVATING ACTIVITY

POLYPEPTIDE O

16q13

GOSR2 604027 GOLGI SNAP RECEPTOR COMPLEX

MEMBER 2

17q21.32

GRIN1 138249 GLUTAMATE RECEPTOR, IONOTROPIC,

N-METHYL-D-ASPARTATE, SUBUNIT 1

9q34.3

GRIN2A 138253 GLUTAMATE RECEPTOR, IONOTROPIC,

N-METHYL-D-ASPARTATE, SUBUNIT 2A

16p13.2

GRIN2B 138252 GLUTAMATE RECEPTOR, IONOTROPIC,

N-METHYL-D-ASPARTATE, SUBUNIT 2B

12p13.1

GRN 138945 GRANULIN PRECURSOR 17q21.31

HADH 601609 3-HYDROXYACYL-CoA DEHYDROGENASE 4q25

HADHA 600890 HYDROXYACYL-CoA

DEHYDROGENASE/3-KETOACYL-CoA

THIOLASE/ENOYL-CoA HYDRATASE,

ALPHA SUBUNIT

2p23.3

HCN1 602780 HYPERPOLARIZATION-ACTIVATED CYCLIC

NUCLEOTIDE-GATED POTASSIUM

CHANNEL 1

5p12

HCN4 605206 HYPERPOLARIZATION-ACTIVATED CYCLIC

NUCLEOTIDE-GATED POTASSIUM

CHANNEL 4

15q24.1

HFE 613609 HFE GENE 6p22.2

HLCS 609018 HOLOCARBOXYLASE SYNTHETASE 21q22.13

HMBS 609806 HYDROXYMETHYLBILANE SYNTHASE 11q23.3

HNRNPU 602869 HETEROGENEOUS NUCLEAR

RIBONUCLEOPROTEIN U

1q44

IQSEC2 300522 IQ MOTIF- AND SEC7

DOMAIN-CONTAINING PROTEIN 2

Xp11.22

KANSL1 612452 KAT8 REGULATORY NSL COMPLEX,

SUBUNIT 1

17q21.31

KCNA1 176260 POTASSIUM CHANNEL, VOLTAGE-GATED,

SHAKER-RELATED SUBFAMILY, MEMBER 1

12p13.32

KCNA2 176262 POTASSIUM CHANNEL, VOLTAGE-GATED,

SHAKER-RELATED SUBFAMILY, MEMBER 2

1p13.3

KCNB1 600397 POTASSIUM CHANNEL, VOLTAGE-GATED,

SHAB-RELATED SUBFAMILY, MEMBER 1

20q13.13

KCNC1 176258 POTASSIUM CHANNEL, VOLTAGE-GATED,

SHAW-RELATED SUBFAMILY, MEMBER 1

11p15.1

KCNH5 605716 POTASSIUM CHANNEL, VOLTAGE-GATED,

SUBFAMILY H, MEMBER 5

14q23.2

KCNJ10 602208 POTASSIUM CHANNEL, INWARDLY

RECTIFYING, SUBFAMILY J, MEMBER 10

1q23.2

KCNJ11 600937 POTASSIUM CHANNEL, INWARDLY

RECTIFYING, SUBFAMILY J, MEMBER 11

11p15.1

KCNMA1 600150 POTASSIUM CHANNEL,

CALCIUM-ACTIVATED, LARGE

CONDUCTANCE, SUBFAMILY M, ALPHA

10q22.3

22

MEMBER 1

KCNQ2 602235 POTASSIUM CHANNEL, VOLTAGE-GATED,

KQT-LIKE SUBFAMILY, MEMBER 2

20q13.33

KCNQ3 602232 POTASSIUM CHANNEL, VOLTAGE-GATED,

KQT-LIKE SUBFAMILY, MEMBER 3

8q24.22

KCNT1 608167 POTASSIUM CHANNEL, SUBFAMILY T,

MEMBER 1

9q34.3

KCTD7 611725 POTASSIUM CHANNEL TETRAMERIZATION


7q11.21

KPNA7 614107 KARYOPHERIN ALPHA-7 7q22.1

LGI1 604619 LEUCINE-RICH GENE,

GLIOMA-INACTIVATED, 1

10q23.33

LIAS 607031 LIPOIC ACID SYNTHASE 4p14

MAGI2 606382 MEMBRANE-ASSOCIATED GUANYLATE

KINASE, WW AND PDZ

DOMAINS-CONTAINING, 2

7q21.11

MBD5 611472 METHYL-CpG-BINDING DOMAIN PROTEIN

5

2q23.1

MECP2 300005 METHYL-CpG-BINDING PROTEIN 2 Xq28

MEF2C 600662 MADS BOX TRANSCRIPTION ENHANCER

FACTOR 2, POLYPEPTIDE C

5q14.3

MFSD8 611124 MAJOR FACILITATOR SUPERFAMILY


4q28.2

MMADHC 611935 MMADHC GENE 2q23.2

MTHFR 607093 5,10-METHYLENETETRAHYDROFOLATE

REDUCTASE

1p36.22

MTR 156570 5-METHYLTETRAHYDROFOLATE-HOMOCY

STEINE S-METHYLTRANSFERASE

1q43

MTRR 602568 METHIONINE SYNTHASE REDUCTASE 5p15.31

NECAP1 611623 NECAP ENDOCYTOSIS-ASSOCIATED

PROTEIN 1

12p13.31

NHLRC1 608072 NHL REPEAT-CONTAINING 1 GENE 6p22.3

NRXN1 600565 NEUREXIN I 2p16.3

OPHN1 300127 OLIGOPHRENIN 1 Xq12

PAH 612349 PHENYLALANINE HYDROXYLASE 12q23.2

PC 608786 PYRUVATE CARBOXYLASE 11q13.2

PCDH19 300460 PROTOCADHERIN 19 Xq22.1

PHGDH 606879 PHOSPHOGLYCERATE DEHYDROGENASE 1p12

PIGA 311770 PHOSPHATIDYLINOSITOL GLYCAN

ANCHOR BIOSYNTHESIS CLASS A PROTEIN

Xp22.2

PIGQ 605754 PHOSPHATIDYLINOSITOL GLYCAN

ANCHOR BIOSYNTHESIS CLASS Q

PROTEIN

16p13.3

PLCB1 607120 PHOSPHOLIPASE C, BETA-1 20p12.3

PNKP 605610 POLYNUCLEOTIDE KINASE 3-PRIME

PHOSPHATASE

19q13.33

PNPO 603287 PYRIDOXAMINE 5-PRIME-PHOSPHATE

OXIDASE

17q21.32

POLG 174763 POLYMERASE, DNA, GAMMA 15q26.1

PPOX 600923 PROTOPORPHYRINOGEN OXIDASE 1q23.3

PPT1 600722 PALMITOYL-PROTEIN THIOESTERASE 1 1p34.2

PRICKLE1 608500 PRICKLE, DROSOPHILA, HOMOLOG OF, 1 12q12

PRICKLE2 608501 PRICKLE, DROSOPHILA, HOMOLOG OF, 2 3p14.1

23

PRODH 606810 PROLINE DEHYDROGENASE (OXIDASE) 1 22q11.21

PRRT2 614386 PROLINE-RICH TRANSMEMBRANE

PROTEIN 2

16p11.2

PURA 600473 PURINE-RICH ELEMENT-BINDING PROTEIN

A

5q31.3

QARS 603727 GLUTAMINYL-tRNA SYNTHETASE 3p21.31

SCARB2 602257 SCAVENGER RECEPTOR CLASS B,

MEMBER 2

4q21.1

SCN1A 182389 SODIUM CHANNEL, NEURONAL TYPE I,

ALPHA SUBUNIT

2q24.3

SCN1B 600235 SODIUM CHANNEL, VOLTAGE-GATED,

TYPE I, BETA SUBUNIT

19q13.11

SCN2A 182390 SODIUM CHANNEL, VOLTAGE-GATED,

TYPE II, ALPHA SUBUNIT

2q24.3


TYPE III, ALPHA SUBUNIT

2q24.3


TYPE VIII, ALPHA SUBUNIT

12q13.13


TYPE IX, ALPHA SUBUNIT

2q24.3

SETBP1 611060 SET-BINDING PROTEIN 1 18q12.3

SIK1 605705 SALT-INDUCIBLE KINASE 1 21q22.3

SLC13A5 608305 SOLUTE CARRIER FAMILY 13

(SODIUM-DEPENDENT CITRATE

TRANSPORTER), MEMBER 5

17p13.1

SLC19A3 606152 SOLUTE CARRIER FAMILY 19 (THIAMINE


2q36.3

SLC22A5 603377 SOLUTE CARRIER FAMILY 22 (ORGANIC

CATION TRANSPORTER), MEMBER 5

5q31.1


(CARNITINE/ACYLCARNITINE

TRANSLOCASE), MEMBER 20

3p21.31


(MITOCHONDRIAL CARRIER,

GLUTAMATE), MEMBER 22

11p15.5


(CARNITINE/ACYLCARNITINE

TRANSLOCASE), MEMBER 29

14q32.2

SLC2A1 138140 SOLUTE CARRIER FAMILY 2 (FACILITATED

GLUCOSE TRANSPORTER), MEMBER 1

1p34.2

SLC46A1 611672 SOLUTE CARRIER FAMILY 46 (FOLATE


17q11.2


(NEUROTRANSMITTER TRANSPORTER,

GABA), MEMBER 1

3p25.3


(NEUROTRANSMITTER TRANSPORTER,

CREATINE), MEMBER 8

Xq28

SLC9A6 300231 SOLUTE CARRIER FAMILY 9, MEMBER 6 Xq26.3

SMARCA2 600014 SWI/SNF-RELATED, MATRIX-ASSOCIATED,

ACTIN-DEPENDENT REGULATOR OF

CHROMATIN, SUBFAMILY A, MEMBER 2

9p24.3

SPTAN1 182810 SPECTRIN, ALPHA, NONERYTHROCYTIC 1 9q34.11

24

SRPX2 300642 SUSHI REPEAT-CONTAINING PROTEIN,

X-LINKED, 2

Xq22.1

ST3GAL3 606494 ST3 BETA-GALACTOSIDE

ALPHA-2,3-SIALYLTRANSFERASE 3

1p34.1

ST3GAL5 604402 ST3 BETA-GALACTOSIDE

ALPHA-2,3-SIALYLTRANSFERASE 5

2p11.2

STX1B 601485 SYNTAXIN 1B 16p11.2

STXBP1 602926 SYNTAXIN-BINDING PROTEIN 1 9q34.11

SYN1 313440 SYNAPSIN I Xp11.3-p11.2

SYNGAP1 603384 SYNAPTIC RAS-GTPase-ACTIVATING

PROTEIN 1

6p21.32

SZT2 615463 SEIZURE THRESHOLD 2, MOUSE,

HOMOLOG OF

1p34.2

TBC1D24 613577 TBC1 DOMAIN FAMILY, MEMBER 24 16p13.3

TBL1XR1 608628 TRANSDUCIN-BETA-LIKE 1 RECEPTOR 1 3q26.32

TCF4 602272 TRANSCRIPTION FACTOR 4 18q21.2

TNK2 606994 TYROSINE KINASE, NONRECEPTOR, 2 3q29

TPP1 607998 TRIPEPTIDYL PEPTIDASE I 11p15.4

TSEN54 608755 tRNA SPLICING ENDONUCLEASE 54, S.

CEREVISIAE, HOMOLOG OF

17q25.1

UBE2A 312180 UBIQUITIN-CONJUGATING ENZYME E2A Xq24

UBE3A 601623 UBIQUITIN-PROTEIN LIGASE E3A 15q11.2

UROD 613521 UROPORPHYRINOGEN DECARBOXYLASE 1p34.1

UROS 606938 UROPORPHYRINOGEN III SYNTHASE 10q26.2

WDR62 613583 WD REPEAT-CONTAINING PROTEIN 62 19q13.12

WWOX 605131 WW DOMAIN-CONTAINING

OXIDOREDUCTASE

16q23.1-q23.2

ZEB2 605802 ZINC FINGER E BOX-BINDING HOMEOBOX

2

2q22.3

25

Appendix 2. Pathogenic variants identified in this study

Patient Gene Nucleotide Amino acid Zygosity

1 ALDH7A1 c.1279G>C p.Glu427Gln Hetero

ALDH7A1 c.192+3A>T Hetero

2 ALDH7A1 c.192+3A>T Hetero

ALDH7A1 c.1093+5G>T Hetero

3 ARX c.995G>A p.Arg332His Hetero

4 BRAT1 c.1276C>T p.Gln426Ter Hetero

BRAT1 c.707T>G p.Leu236Arg Hetero

5 BRAT1 c.1276C>T p.Gln426Ter Hetero

BRAT1 c.707T>G p.Leu236Arg Hetero

6 BRAT1 Exon 2-3 deletion Hetero

BRAT1 c.1576C>T p.Gln526Ter Hetero

7 CACNA1A c.526G>A p.Val176Met Hetero

8 CACNB4* c.21delC p.Lys8ArgfsTer26 Hetero

9 CASK c.533-1G>C Hemi

10 CDKL5 c.145+2T>A Hemi

11 CDKL5* c.978-1G>A Hetero

12 CDKL5* c.282+1G>A Hetero

13 CDKL5* c.458A>T p.Asp153Val Hetero

14 CDKL5 c.511T>A p.Tyr171Asn Hetero

15 CDKL5 c.403+1G>A Hetero

16 CDKL5 c.175C>T p.Arg59Ter Hemi

17 CDKL5 c.513C>A p.Tyr171Ter Hetero

18 CDKL5* c.2354dupA p.Lys786GlufsTer15 Hetero

19 CHD2 c.361C>T p.Arg121Ter Hetero

20 CHD2* c.3885dupA p.Ile1296AsnfsTer8 Hetero

21 CHD2* c.1269dupA p.Glu424ArgfsTer3 Hetero

22 CHD2* c.1453C>T p.Arg485Ter Hetero

23 CHD2 Exon 5 deletion Hetero

24 CHD2 c.4507C>T p.Arg1503Trp Hetero

25 CHD2* c.3172G>T p.Glu1058Ter Hetero

26 CHD2* c.1897_1898delCT p.Leu633AspfsTer2 Hetero

27 DNM1* c.1195A>G p.Arg399Gly Hetero

28 DNM1* c.632A>T p.Asp211Val Hetero

29 EEF1A2 c.208G>A p.Gly70Ser Hetero

30 EEF1A2* c.294C>G p.Phe98Leu Hetero

31 GNAO1* c.155A>C p.Gln52Pro Hetero

32 GRIN2A c.1592C>T p.Thr531Met Hetero

33 HCN1* Exon 1-5 duplication Hetero

34 IQSEC2* c.136G>T p.Glu46Ter Hetero

35 KANSL1 Exon 2-3 duplication Hetero

36 KCNA1 c.1112C>T p.Thr371Ile Hetero

37 KCNB1 c.1135G>A p.Gly379Arg Hetero

38 KCNB1 c.1135G>A p.Gly379Arg Hetero

39 KCNQ2 c.773A>T p.Asn258Ile Hetero

40 KCNQ2 c.917C>T p.Ala306Val Hetero

41 KCNQ2 c.338C>T p.Ser113Phe Hetero

42 KCNQ2 c.1639C>T p.Arg547Trp Hetero

43 KCNQ2 c.638G>A p.Arg213Gln Hetero

44 KCNQ2 c.794C>T p.Ala265Val Hetero

26

45 KCNQ2 c.593G>A p.Arg198Gln Hetero

46 KCNT1 c.1421G>A p.Arg474His Hetero

47 KCNT1 c.2800G>A p.Ala934Thr Hetero

48 KCNT1* c.1038C>G p.Phe346Leu Hetero

49 MECP2 c.1164_1207delACCTCC

ACCTGAGCCCGAGAG

CTCCGAGGACCCCAC

CAGCCCCC

p.Pro389Ter Hetero

50 MECP2 Whole gene duplication Hetero



53 MECP2 c.502C>T p.Arg168Ter Hetero

54 PCDH19 Whole gene deletion Hetero

55 PCDH19 c.1019A>G p.Asn340Ser Hetero

56 PCDH19 Whole gene deletion Hetero

57 PRODH Whole exon deletion Hetero

58 SCN1A c.1154A>G p.Glu385Gly Hetero

59 SCN1A* c.644_655delTGAGAAC

ATTCA

p.Leu215_Arg219delinsT

er

Hetero

60 SCN1A Exon 20 deletion Hetero

61 SCN1A c.249C>G p.Tyr83Ter Hetero

62 SCN1A c.2210G>A p.Trp737Ter Hetero

63 SCN1A* c.5532dupC p.Met1845HisfsTer5 Hetero

64 SCN1A c.4306-1G>A Hetero

65 SCN1A* c.5163delC p.Ile1722PhefsTer46 Hetero

66 SCN1A Exon 7-16 deletion Hetero

67 SCN1A* c.2522T>C p.Val841Ala Hetero

68 SCN1A Whole exon deletion Hetero

69 SCN1B Exon 1-2 deletion Hetero

70 SCN2A* c.5308A>G p.Met1770Val Hetero

71 SCN2A* c.5327T>C p.Leu1776Pro Hetero

72 SCN2A c.5317G>A p.Ala1773Thr Hetero

73 SCN2A c.2516C>T p.Ala839Val Hetero

74 SCN2A c.1747C>T p.Arg583Ter Hetero

75 SCN3A Exon 9 duplication Hetero

76 SCN8A* c.424A>G p.Ile142Val Hetero

77 SCN8A c.2549G>A p.Arg850Gln Hetero

78 SCN8A c.5614C>T p.Arg1872Trp Hetero

79 SCN8A* c.782G>T p.Cys261Phe Hetero

80 SCN8A c.4423G>A p.Gly1475Arg Hetero

81 SLC6A1 Whole gene deletion Hetero

82 SLC6A1* c.1435C>T p.Arg479Ter Hetero

83 SLC6A1* c.694G>C p.Gly232Arg Hetero

84 SLC9A6 c.316_325+28delATGAT

TTATGGCAAGTTCCTC

AACCCTTGTCAGCCC

CT

Hemi

85 SLC9A6* c.589delT p.Tyr197IlefsTer3 Hemi

86 STXBP1* c.84G>A p.Trp28Ter Hetero

87 STXBP1 c.733C>G p.His245Asp Hetero

88 STXBP1 c.581_582dupAA p.Tyr195AsnfsTer11 Hetero

89 STXBP1 c.1216C>T p.Arg406Cys Hetero

90 STXBP1* Exon 7-10 deletion Hetero

27

91 STXBP1 c.874C>T p.Arg292Cys Hetero

92 STXBP1* c.1497C>G p.Tyr499Ter Hetero

93 SYN1 Whole gene duplication Hetero

94 SYNGAP1* c.2014delA p.Thr672ArgfsTer2 Hetero

95 SYNGAP1* c.557T>A p.Leu186Ter Hetero

96 SYNGAP1 c.2764C>T p.Arg922Ter Hetero

97 SYNGAP1 c.1735C>T p.Arg579Ter Hetero

98 SYNGAP1 c.980T>C p.Leu327Pro Hetero

99 UBE3A c.2294+1G>A Hetero

100 UBE3A Whole gene deletion Hetero

101 WWOX Exon 6-8 duplication Hetero

WWOX c.1060C>T p.Gln354Ter Hetero

102 ZEB2 c.1956C>A p.Tyr652Ter Hetero

103 ZEB2* c.2348dupC p.Ser784PhefsTer11 Hetero

28

ABSTRACT (IN KOREAN)

뇌전증성 뇌병증 환아에서 차세대 염기서열분석 유전자 패널과

유전형-표현형 연관성 연구

연세대학교 대학원 의학과

(지도교수: 강훈철)

고아라

뇌전증성 뇌병증 환아를 대상으로 차세대 염기서열분석 유전자 패널

검사를 시행하였고, 임상적으로 유전형-표현형 연관성을 조사하는 것이 이

연구의 목적이다.

뇌전증성 뇌병증 환아 278명에 대해 172개의 유전자로 이루어진 유전자

패널 검사를 시행하였고, 유전형에 따라 환자들의 치료 효과를 포함한

임상적 특징을 조사하였다.

278명의 뇌전증성 뇌병증 환아 중 103명 (37.1%)에게서 원인 유전자

변이가 확인되었으며, 총 35개의 유전자가 원인 유전자로 밝혀졌다.

진단율은 첫 경련이 어릴 때에 발생했을 수록, 특히 신생아기에 했을 때,

또한 약물 저항성 뇌전증을 보일수록 높았다. 뇌전증 증후군 별로 보았을

때는, 신생아 경련의 과거력이 있는 웨스트 증후군 환아에게서 진단율이

가장 높았으며, KCNQ2와 STXBP1의 변이가 가장 자주 보였다. 유전형 별로

임상적 경과와 각 치료법에 따른 경련 조절 여부도 조사하였을 때 이전

보고와 비슷한 결과가 나왔으며, KCNQ2, 영아기에 발생한 SCN2A, SCN8A

변이로 인한 뇌병증에서 나트륨통로차단제가 효과적이었으며, SCN1A, CDKL5,

KCNQ2, STXBP1, SCN2A 변이에서 케톤생성식이요법은 다양한 반응을 보였다.

2명의 KCNT1 변이를 보이는 영아기 이주성 부분 뇌전증 환아에서 quinidine

투여를 시도해 보았으나, 경련 발작에 효과를 보이지는 않았다.

차세대 염기서열분석 유전자 패널은 뇌전증성 뇌병증 환아에게 유용한

진단 도구이며, 유전형-표현형 연관성은 이러한 환아에서 임상적 예후 및

치료 효과를 예측하는데 도움이 된다.

----------------------------------------------------------------------------------------

핵심되는 말: 뇌전증성 뇌병증, 차세대 염기서열분석, 유전형-표현형

29

PUBLICATION LIST

Ko, A, Youn, SE, Kim, SH, Lee, JS, Kim, S, Choi, JR, et al. Targeted gene panel and

genotype-phenotype correlation in children with developmental and epileptic

enceaphlopathy. Epilepsy research 2018; 141: 48-55.

targeted gene panel and genotype-phenotype correlation in … · 2020. 7. 3. · of epilepsy...

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