BRCA somatic and germline mutation detection in paraffin embedded ovarian cancers by next-generation sequencing.
BRCA somatic and germline mutation detection in paraffin embedded ovarian cancers by next-generation sequencing.
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DOI：
10.18632/oncotarget.6834
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发表时间：
2016-01-12
期刊：
影响因子：
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通讯作者：
Scarpa A
中科院分区：
文献类型：
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作者：
Mafficini A;Simbolo M;Parisi A;Rusev B;Luchini C;Cataldo I;Piazzola E;Sperandio N;Turri G;Franchi M;Tortora G;Bovo C;Lawlor RT;Scarpa A
BRCA突变的卵巢癌对铂类药物和最近批准的PARP抑制剂的反应更好。有必要使用福尔马林固定石蜡包埋（FFPE）组织和商业上可获得的技术来检测体细胞和种系突变的有效和及时的方法。我们使用商业试剂盒探索BRCA 1和BRCA 2基因的所有外显子和50 bp外显子-内含子连接，并对47例高级别浆液性卵巢癌FFPE样本的DNA进行半导体下一代测序（NGS）。在13/47（28%）的癌症中发现了致病性突变：8个在BRCA 1中，5个在BRCA 2中。所有BRCA 1和两个BRCA 2突变都是生殖系的;三个BRCA 2突变是体细胞的。通过桑格测序证实所有突变。为了评价NGS panel的性能，我们使用可调用性分析评估了其检测ClinVar和COSMIC数据库中BRCA 1和BRCA 2所描述的6，953种变体的能力。软件自动识别了6，059个（87.1%）变体; 829个（12.0%）需要目视验证。其余65例（0.9%）变异为不可调用，需要15例桑格反应才能解决。因此，NGS板的灵敏度为99.1%。总之，使用商业试剂盒进行的NGS对于使用常规FFPE组织检测BRCA基因的生殖系和体细胞突变是高效的。
BRCA mutated ovarian cancers respond better to platinum-based therapy and to the recently approved PARP-inhibitors. There is the need for efficient and timely methods to detect both somatic and germline mutations using formalin-fixed paraffin-embedded (FFPE) tissues and commercially available technology. We used a commercial kit exploring all exons and 50bp exon-intron junctions of BRCA1 and BRCA2 genes, and semiconductor next-generation sequencing (NGS) on DNA from 47 FFPE samples of high-grade serous ovarian cancers. Pathogenic mutations were found in 13/47 (28%) cancers: eight in BRCA1 and five in BRCA2. All BRCA1 and two BRCA2 mutations were germline; three BRCA2 mutations were somatic. All mutations were confirmed by Sanger sequencing. To evaluate the performance of the NGS panel, we assessed its capability to detect the 6,953 variants described for BRCA1 and BRCA2 in ClinVar and COSMIC databases using callability analysis. 6,059 (87.1%) variants were identified automatically by the software; 829 (12.0%) required visual verification. The remaining 65 (0.9%) variants were uncallable, and would require 15 Sanger reactions to be resolved. Thus, the sensitivity of the NGS-panel was 99.1%. In conclusion, NGS performed with a commercial kit is highly efficient for detection of germline and somatic mutations in BRCA genes using routine FFPE tissue.