Center for Human Genetic Research and Department of Molecular Biology, Massachusetts General Hospital, 185 Cambridge Street, Sixth Floor, Boston, MA 02114, USA.
Sci Transl Med. 2012 Jan 25;4(118):118ra10. doi: 10.1126/scitranslmed.3003310.
Advances in next-generation sequencing (NGS) promise to facilitate diagnosis of inherited disorders. Although in research settings NGS has pinpointed causal alleles using segregation in large families, the key challenge for clinical diagnosis is application to single individuals. To explore its diagnostic use, we performed targeted NGS in 42 unrelated infants with clinical and biochemical evidence of mitochondrial oxidative phosphorylation disease. These devastating mitochondrial disorders are characterized by phenotypic and genetic heterogeneity, with more than 100 causal genes identified to date. We performed "MitoExome" sequencing of the mitochondrial DNA (mtDNA) and exons of ~1000 nuclear genes encoding mitochondrial proteins and prioritized rare mutations predicted to disrupt function. Because patients and healthy control individuals harbored a comparable number of such heterozygous alleles, we could not prioritize dominant-acting genes. However, patients showed a fivefold enrichment of genes with two such mutations that could underlie recessive disease. In total, 23 of 42 (55%) patients harbored such recessive genes or pathogenic mtDNA variants. Firm diagnoses were enabled in 10 patients (24%) who had mutations in genes previously linked to disease. Thirteen patients (31%) had mutations in nuclear genes not previously linked to disease. The pathogenicity of two such genes, NDUFB3 and AGK, was supported by complementation studies and evidence from multiple patients, respectively. The results underscore the potential and challenges of deploying NGS in clinical settings.
下一代测序（NGS）的进步有望促进遗传性疾病的诊断。尽管在研究环境中，NGS 通过大的家族中的分离已经确定了因果等位基因，但临床诊断的关键挑战是应用于单个个体。为了探索其诊断用途，我们对 42 名具有临床和生化证据表明患有线粒体氧化磷酸化疾病的非相关婴儿进行了靶向 NGS。这些破坏性的线粒体疾病的特点是表型和遗传异质性，迄今为止已经确定了 100 多个因果基因。我们对线粒体 DNA（mtDNA）和编码线粒体蛋白的~1000 个核基因的外显子进行了“线粒体外显子组”测序，并优先考虑了预测会破坏功能的罕见突变。由于患者和健康对照个体都携带有相似数量的杂合等位基因，因此我们无法优先考虑显性作用的基因。然而，患者携带的可能导致隐性疾病的两种此类突变的基因富集了五倍。总的来说，42 名患者中有 23 名（55%）携带此类隐性基因或致病性 mtDNA 变体。在 10 名（24%）以前与疾病相关的基因发生突变的患者中能够确定明确的诊断。13 名（31%）患者的核基因突变以前与疾病无关。对两个此类基因 NDUFB3 和 AGK 的致病性，分别通过互补研究和来自多个患者的证据得到了支持。这些结果强调了在临床环境中部署 NGS 的潜力和挑战。
