基于目标序列捕获二代测序技术对结节性硬化症相关肾脏病变患者的基因诊断

Application of targeted second-generation sequencing in the genetic diagnosis of tuberous sclerosis complex associated with renal complications

目的 评估目标序列捕获二代测序技术在结节性硬化症相关肾脏病变患者基因诊断中的应用价值.方法 分析2014年1月至2015年12月北京协和医院泌尿外科收治的43例临床诊断（33例确定诊断和10例可能诊断）为结节性硬化症相关肾脏病变患者的临床资料.男17例,女26例.年龄10 ～48岁,平均28岁.43例中39例肾脏病变表现为多发性肾脏血管平滑肌脂肪瘤,3例表现为多发性肾囊肿,1例表现为肾透明细胞癌.签署知情同意书后,采集外周血5 ml提取基因组DNA,进行目标序列捕获二代测序靶基因TSC1和TSC2,应用Sanger测序技术完成验证.结果 39例（90.7％）检测到与临床表型相符的基因突变,2例检测到临床意义未明的突变,另2例未能检测到任何突变.我们发现了14种新的致病性突变,包括8种框移突变,3种大片段缺失突变,2种无义突变和1种剪接突变.结论 目标序列捕获二代测序技术可快速、准确检测出TSC1和TSC2基因突变,对于结节性硬化症相关肾脏病变患者的基因诊断具有重要价值.

Objective To evaluate the value of targeted second-generation sequencing （NGS） in the genetic diagnosis of tuberous sclerosis complex （TSC） associated with renal complications.Methods The clinical data of 43 patients （with 33 patients of definite diagnosis and 10 patients of possible diagnosis）with tuberous sclerosis complex associated with renal complications were analyzed.There were 26 females and 17 males with a mean age of 28 （10 ～ 48） years ranging from 10 to 48 years old.All patients had renal complications,including 39 renal angiomyolipomas,3 renal cysts and 1 renal cell carcinoma.Written informed consents were signed,and 5ml peripheral blood was drawn for DNA extraction.Mutations of TSC1 and TSC2 genes were detected by the NGS technology,and then confirmed by Sanger sequencing.Results TSC1 or TSC2 pathogenetic mutants were identified in 39 patients （90.7%）,which were consistent with clinical phenotypes and two non-significant mutations were identified in two individuals,while no mutation was detected in the other two cases.Fourteen novel mutations were reported for the first time,including 8 frameshift mutation,3 deletion mutation,2 nonsense mutation and 1 splicing mutation.Conclusion NGS is an accurate and less time-consuming technology in detecting TSC1 or TSC2 gene mutation,which has great value in genetic diagnosis of tuberous sclerosis complex associated with renal complications.