發(fā)布時(shí)間：2018-06-10 11:55

  本文選題：青少年肌陣攣癲癇 + 全外顯子組測(cè)序�。� 參考：《第四軍醫(yī)大學(xué)》2017年碩士論文

【摘要】：目的通過對(duì)確診為青少年肌陣攣癲癇(juvenile myoclonic epilepsy,JME)的患者進(jìn)行全外顯子組測(cè)序(whole-exome sequencing,WES),明確JME患者的基因診斷,為JME的發(fā)病機(jī)制及診療提供依據(jù)。方法對(duì)2015年6月-2016年12月就診于第四軍醫(yī)大學(xué)西京醫(yī)院神經(jīng)內(nèi)科癲癇�？崎T診的確診為JME的29例患者進(jìn)行WES,通過與數(shù)據(jù)庫比對(duì)及生物信息學(xué)分析初步篩選基因突變位點(diǎn),對(duì)該位點(diǎn)進(jìn)行一代驗(yàn)證和相關(guān)親屬驗(yàn)證,依據(jù)2015年美國醫(yī)學(xué)遺傳學(xué)與基因組學(xué)學(xué)會(huì)(ACMG)聯(lián)合分子病理學(xué)學(xué)會(huì)發(fā)布的《序列變異解讀標(biāo)準(zhǔn)和指南》篩選出致病或可能致病的突變,明確這些JME患者的基因診斷,并結(jié)合臨床用藥療效分析不同變異的治療效果。結(jié)果在29例行WES的JME患者中,發(fā)現(xiàn)23例患者存在致病或可能致病的基因突變,共檢出21個(gè)突變位點(diǎn)(其中3例患者為同一位點(diǎn)突變),包括20個(gè)單基因錯(cuò)義突變和1個(gè)單基因移碼突變。在20個(gè)單基因錯(cuò)義突變中,CLCN2突變位點(diǎn)6個(gè):c.1834GAp.R612C, c.1991AT p.E664V, c.1141CG p.P381A, c.2006GAp.A669V,c.1141GC p.P381A,c.1705GA p.G569S; CACNA1H突變位點(diǎn) 3 個(gè):c.1735GA p.D579N,c.5468GA p.R1823H,c.3646GA p.D1216N; CHD2突變位點(diǎn) 2 個(gè):c.927CGp.I309M,c.2291ACp.H764P;以下突變位點(diǎn)各 1 個(gè):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個(gè)單基因移碼突變?yōu)镈EPDC5c.3225_c.3226insG p.V1076Gfs*85。發(fā)現(xiàn)6例患者基因突變的位點(diǎn)尚未見既往文獻(xiàn)報(bào)道,包括5例單基因錯(cuò)義突變(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 例單基因移碼突變(DEPDC5 c.3225_c.3226insGp.V1076Gfs*85),5例單基因錯(cuò)義突變經(jīng)軟件預(yù)測(cè)結(jié)果顯示突變導(dǎo)致的蛋白質(zhì)翻譯的變化對(duì)蛋白功能均有影響,1例單基因移碼突變因?qū)е翫EPDC5基因編碼的蛋白從第1076位氨基酸開始算起,后面再翻譯85個(gè)氨基酸即終止,且這85個(gè)氨基酸是移碼的,序列和原來的序列完全不同,因此該位點(diǎn)突變風(fēng)險(xiǎn)較高,致病可能性較大。另有8例患者的突變基因?yàn)槟壳把芯繄?bào)道中與其他類型癲癇相關(guān)的基因,尚未見這些基因與JME發(fā)病相關(guān)的報(bào)道。6例CLCN2基因突變的患者中,3例為肌陣攣發(fā)作(myoclonic seizures,MS)伴全面強(qiáng)直-陣攣發(fā)作(generalized tonic-clonic seizures,GTCS),2 例為 MS 伴 GTCS 和失神發(fā)作(absence seizures,AS),1例患者為單純MS。這6例患者中,有4例患者接受丙戊酸治療,3例總發(fā)作減少100%,1例總發(fā)作減少75%-99%; 2例患者接受左乙拉西坦治療,總發(fā)作減少均在50%-74%。發(fā)現(xiàn)5例患者存在CACNA1H基因突變,其發(fā)作類型均為MS伴GTCS,其中有2例患者接受丙戊酸單藥治療,3例接受雙藥聯(lián)合治療,所有患者的GTCS均減少100%,MS減少均在75%-99%,其中有2例患者曾在MS未得到控制時(shí)嘗試減藥,但均在減藥過程中(未完全停藥)出現(xiàn)GTCS。結(jié)論本研究在29例行WES的JME患者中發(fā)現(xiàn)23例患者存在致病或可能致病的基因突變,其中6例為新發(fā)現(xiàn)的突變,突變位點(diǎn)尚未見相關(guān)文獻(xiàn)報(bào)道。另有8例患者的突變基因?yàn)槟壳把芯繄?bào)道的與其他類型癲癇(非JME)相關(guān)的基因,推測(cè)JME可能和其他類型癲癇擁有共同的基因基礎(chǔ)。CACNA1H突變的JME患者的表型可能多為MS合并GTCS,抗癲癇治療后GTCS較MS易得到控制,同時(shí)該類患者須慎重減停藥�？傊�,WES為JME患者的基因診斷提供了線索,同時(shí)也為該病的發(fā)病機(jī)制及精準(zhǔn)治療提供了一定的依據(jù)。
[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.
【學(xué)位授予單位】：第四軍醫(yī)大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類號(hào)】：R742.1

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