青少年肌阵挛癞痫患者全外显子组测序研究及其临床意义

发布时间：2018-06-10 11:55

本文选题：青少年肌阵挛癫痫 + 全外显子组测序　；参考：《第四军医大学》2017年硕士论文

【摘要】：目的通过对确诊为青少年肌阵挛癫痫(juvenile myoclonic epilepsy,JME)的患者进行全外显子组测序(whole-exome sequencing,WES),明确JME患者的基因诊断,为JME的发病机制及诊疗提供依据。方法对2015年6月-2016年12月就诊于第四军医大学西京医院神经内科癫痫专科门诊的确诊为JME的29例患者进行WES,通过与数据库比对及生物信息学分析初步筛选基因突变位点,对该位点进行一代验证和相关亲属验证,依据2015年美国医学遗传学与基因组学学会(ACMG)联合分子病理学学会发布的《序列变异解读标准和指南》筛选出致病或可能致病的突变,明确这些JME患者的基因诊断,并结合临床用药疗效分析不同变异的治疗效果。结果在29例行WES的JME患者中,发现23例患者存在致病或可能致病的基因突变,共检出21个突变位点(其中3例患者为同一位点突变),包括20个单基因错义突变和1个单基因移码突变。在20个单基因错义突变中,CLCN2突变位点6个:c.1834GAp.R612C, c.1991AT p.E664V, c.1141CG p.P381A, c.2006GAp.A669V,c.1141GC p.P381A,c.1705GA p.G569S; CACNA1H突变位点 3 个:c.1735GA p.D579N,c.5468GA p.R1823H,c.3646GA p.D1216N; CHD2突变位点 2 个:c.927CGp.I309M,c.2291ACp.H764P;以下突变位点各 1 个:SCN2Ac.1571GA p.R524Q, LAMC3 c.1675AG p.I559V, GRIN2A c.2636AG p.K879R, EFHC1с.268AG p.M90V, CHRNA2 c.1073GT p.S358I, GABRA1 c.116CT p.T39I, CACNB4 c.1550CTp.R517Q,KCNT1 c.3317GAp.R1106Q, CACNA1D c.2206AGp.M736V)。1个单基因移码突变为DEPDC5c.3225_c.3226insG p.V1076Gfs*85。发现6例患者基因突变的位点尚未见既往文献报道,包括5例单基因错义突变(CLCN2 c.1834GAр.R612C, EFHC1 c.268AG p.M90V, GABRA1 c.116CT p.T39I, CACNB4 c.1550CT p.R517Q,CHD2c.2291AC p.H764P)和 1 例单基因移码突变(DEPDC5 c.3225_c.3226insGp.V1076Gfs*85),5例单基因错义突变经软件预测结果显示突变导致的蛋白质翻译的变化对蛋白功能均有影响,1例单基因移码突变因导致DEPDC5基因编码的蛋白从第1076位氨基酸开始算起,后面再翻译85个氨基酸即终止,且这85个氨基酸是移码的,序列和原来的序列完全不同,因此该位点突变风险较高,致病可能性较大。另有8例患者的突变基因为目前研究报道中与其他类型癫痫相关的基因,尚未见这些基因与JME发病相关的报道。6例CLCN2基因突变的患者中,3例为肌阵挛发作(myoclonic seizures,MS)伴全面强直-阵挛发作(generalized tonic-clonic seizures,GTCS),2 例为 MS 伴 GTCS 和失神发作(absence seizures,AS),1例患者为单纯MS。这6例患者中,有4例患者接受丙戊酸治疗,3例总发作减少100%,1例总发作减少75%-99%; 2例患者接受左乙拉西坦治疗,总发作减少均在50%-74%。发现5例患者存在CACNA1H基因突变,其发作类型均为MS伴GTCS,其中有2例患者接受丙戊酸单药治疗,3例接受双药联合治疗,所有患者的GTCS均减少100%,MS减少均在75%-99%,其中有2例患者曾在MS未得到控制时尝试减药,但均在减药过程中(未完全停药)出现GTCS。结论本研究在29例行WES的JME患者中发现23例患者存在致病或可能致病的基因突变,其中6例为新发现的突变,突变位点尚未见相关文献报道。另有8例患者的突变基因为目前研究报道的与其他类型癫痫(非JME)相关的基因,推测JME可能和其他类型癫痫拥有共同的基因基础。CACNA1H突变的JME患者的表型可能多为MS合并GTCS,抗癫痫治疗后GTCS较MS易得到控制,同时该类患者须慎重减停药。总之,WES为JME患者的基因诊断提供了线索,同时也为该病的发病机制及精准治疗提供了一定的依据。
[Abstract]:Objective to carry out all exon sequencing (whole-exome sequencing, WES) in patients with juvenile myoclonic epilepsy (JME) and to clarify the genetic diagnosis of JME patients and provide basis for the pathogenesis and diagnosis and treatment of JME. Methods in December of June 2015 -2016 year in Xijing Hospital of The Fourth Military Medical University. WES was performed in 29 patients with JME in the medical clinic of the medical department of internal medicine. By screening the gene mutation sites with database comparison and bioinformatics analysis, the site was verified by one generation and related relative verification, according to the sequence of the United molecular pathology Association of the American Medical Genetics and genomics Society (ACMG) in 2015. Mutation interpretation criteria and guidelines > screening out pathogenic or possible pathogeny mutations, identifying the genetic diagnosis of these JME patients and analyzing the therapeutic effects of different variations in combination with clinical efficacy. Results in 29 cases of WES's JME patients, 23 patients were found to have pathogenic or possibly pathogenic gene mutations, and 21 mutation sites were detected (3 of them). The patient was the same mutation), including 20 monogenic missense mutations and 1 single gene shift mutation. In 20 single gene missense mutations, 6 CLCN2 mutation sites were c.1834GAp.R612C, c.1991AT p.E664V, c.1141CG p.P381A, c.2006GAp.A669V, c.1141GC p.P381A, c.1705GA p.G569S, and 3 CACNA1H mutation sites. A p.R1823H, c.3646GA p.D1216N, CHD2 mutation site 2: c.927CGp.I309M, c.2291ACp.H764P; the following mutation loci are 1: SCN2Ac.1571GA p.R524Q, LAMC3 c.1675AG p.I559V. CACNA1D c.2206AGp.M736V).1 single gene shift mutation to DEPDC5c.3225_c.3226insG p.V1076Gfs*85. found that 6 cases of gene mutation have not been reported in previous literature, including 5 single gene missense mutations (EFHC1 c.268AG p.M90V.R612C, EFHC1 c.268AG p.M90V, GABRA1) 764P) and 1 single gene transcoding mutation (DEPDC5 c.3225_c.3226insGp.V1076Gfs*85). 5 cases of single gene missense mutation show that the changes of protein translation caused by the mutation have influence on the protein function. 1 single gene transcoding mutations cause the protein of the DEPDC5 gene to start from 1076th amino acids. The 85 amino acids are terminated, and the 85 amino acids are transcoding, and the sequence is completely different from the original sequence, so the mutation risk is higher and the pathogenicity is greater. In addition, 8 of the mutant genes are related to other types of epilepsy in the present study, and there has not been a report of these genes related to the pathogenesis of JME.6 Among the patients with CLCN2 gene mutation, 3 were myoclonic seizures (MS) with total tonic clonic seizure (generalized tonic-clonic seizures, GTCS). 2 cases were MS accompanied by GTCS and absence seizures (absence seizures), and 1 patients were only 6 cases, 4 patients received valproic acid treatment, 3 cases decreased 1 of total attack. 00%, 1 cases of total seizures decreased 75%-99%, 2 patients received levetiracetam treatment, total seizure reduction was found in 5 cases of CACNA1H mutations in 5 patients, the type of attack was MS with GTCS, of which 2 patients received valproic acid single drug treatment, 3 cases were treated with double drug combination, all patients were reduced by 100%, MS decreased in 75%- 99%, 2 of them had tried to reduce the drug when MS was not controlled, but all in the process of reducing the drug (not completely stopped) GTCS. conclusion the study was found in 29 cases of JME patients with WES, and 23 of the patients were found to have pathogenetic or potentially pathogenic mutations, of which 6 were newly discovered, and the mutation sites had not yet been reported in the literature. And 8 The mutant gene of the patient is currently reported to be related to other types of epileptic (non JME) related genes. It is presumed that the phenotype of JME may be MS combined with GTCS in the JME patients with common genetic basis.CACNA1H mutations in other types of epilepsy, and GTCS is more easily controlled than MS after antiepileptic treatment. WES provides clues for gene diagnosis of JME patients, and provides a basis for the pathogenesis and precise treatment of the disease.
【学位授予单位】：第四军医大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R742.1

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