St John's Institute of Dermatology, King's College London (Guy's Campus), London, U.K; Department of Dermatology, Nagoya University Graduate School of Medicine, Nagoya, Japan.
Br J Dermatol. 2015 Jan;172(1):94-100. doi: 10.1111/bjd.13190. Epub 2014 Nov 19.
Subtypes of inherited epidermolysis bullosa (EB) vary significantly in their clinical presentation and prognosis. Establishing an accurate diagnosis is important for genetic counselling and patient management. Current approaches in EB diagnostics involve skin biopsy for immunohistochemistry and transmission electron microscopy, and Sanger sequencing of candidate genes. Although informative in most cases, this approach can be expensive and laborious and may fail to identify pathogenic mutations in ~15% of cases.
Next-generation DNA sequencing (NGS) technologies offer a fast and efficient complementary diagnostic strategy, but the value of NGS in EB diagnostics has yet to be explored. The aim of this study was to undertake whole-exome sequencing (WES) in nine cases of EB in which established diagnostic methods failed to make a genetic diagnosis.
Whole-exome capture was performed using genomic DNA from each case of EB, followed by massively parallel sequencing. Resulting reads were mapped to the human genome reference hg19. Potentially pathogenic mutations were subsequently confirmed by Sanger sequencing.
Analysis of WES data disclosed biallelic pathogenic mutations in each case, with all mutations occurring in known EB genes (LAMB3, PLEC, FERMT1 and COL7A1). This study demonstrates that NGS can improve diagnostic sensitivity in EB compared with current laboratory practice.
With appropriate diagnostic platforms and bioinformatics support, WES is likely to increase mutation detection in cases of EB and improve EB diagnostic services, although skin biopsy remains an important diagnostic investigation in current clinical practice.
遗传性大疱性表皮松解症 (EB) 的亚型在临床表现和预后方面差异很大。准确诊断对于遗传咨询和患者管理非常重要。目前的 EB 诊断方法包括皮肤活检进行免疫组织化学和透射电子显微镜检查，以及候选基因的 Sanger 测序。虽然在大多数情况下都很有帮助，但这种方法可能既昂贵又繁琐，并且可能无法在~15%的病例中识别出致病性突变。
下一代 DNA 测序 (NGS) 技术提供了一种快速有效的互补诊断策略，但 NGS 在 EB 诊断中的价值尚未得到探索。本研究旨在对 9 例经传统诊断方法未能确定遗传诊断的 EB 病例进行全外显子组测序 (WES)。
使用来自每个 EB 病例的基因组 DNA 进行全外显子捕获，然后进行大规模平行测序。将产生的读数映射到人类基因组参考 hg19。随后通过 Sanger 测序确认潜在的致病性突变。
WES 数据分析显示每个病例均存在双等位基因致病性突变，所有突变均发生在已知的 EB 基因 (LAMB3、PLEC、FERMT1 和 COL7A1) 中。本研究表明，与当前的实验室实践相比，NGS 可以提高 EB 的诊断灵敏度。
在适当的诊断平台和生物信息学支持下，WES 可能会增加 EB 病例中的突变检测，并改善 EB 诊断服务，尽管皮肤活检在当前临床实践中仍然是一项重要的诊断研究。
