Comparison of testing methods for the detection of BRAF V600E mutations in malignant melanoma: pre-approval validation study of the companion diagnostic test for vemurafenib.
Comparison of testing methods for the detection of BRAF V600E mutations in malignant melanoma: pre-approval validation study of the companion diagnostic test for vemurafenib.
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DOI：
10.1371/journal.pone.0053733
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发表时间：
2013
期刊：
影响因子：
3.7
通讯作者：
de Castro DG
中科院分区：
文献类型：
--
作者：
Lopez-Rios F;Angulo B;Gomez B;Mair D;Martinez R;Conde E;Shieh F;Vaks J;Langland R;Lawrence HJ;de Castro DG
cobas 4800 BRAF V600 Mutation Test是一种经CE认证和FDA批准的体外诊断试验，用于选择转移性黑色素瘤患者接受选择性BRAF抑制剂vemurafenib治疗。我们描述了该检测在两个外部实验室进行的批准前验证。在两个实验室使用cobas BRAF突变检测、ABI BRAF检测和双向直接测序检测黑色素瘤标本的BRAF V600突变。确定cobas检测和其他检测之间的阳性（PPA）和阴性（NPA）一致率。采用大规模平行焦磷酸测序法对结果不一致的标本进行检测（454）。测试具有5%突变等位基因的DNA混合物以评估检出率。在桑格检测的8/116份标本（6.9%）、ABI BRAF检测的10/116份标本（8.6%）和cobas BRAF检测的0/232份标本（0%）中观察到无效结果。对于cobas BRAF试验和桑格试验，V600 E突变的PPA为97.7%，NPA为95.3%。cobas BRAF和ABI BRAF的PPA为71.9%，NPA为83.7%。对于16份cobas BRAF检测阴性/ABI BRAF阳性标本，454测序在12份中未检测到密码子600突变，在4份中检测到变异密码子600突变。对于8份cobas BRAF检测阳性/ABI BRAF阴性标本，通过454测序，4份为V600 E，4份为V600 K。对于5%突变混合物，cobas BRAF试验的检出率为100%，桑格试验的检出率为33%，ABI BRAF试验的检出率为21%。两个研究中心之间cobas BRAF试验的重现性为111/116（96%）。在关键临床试验验证的同时，在外部实验室评价潜在的伴随诊断检测是可行的。卫生监管部门批准的测定方法的性能特征明显优于其他两种方法。cobas BRAF测试的总体成功是未来生物标志物开发的概念证明。
The cobas 4800 BRAF V600 Mutation Test is a CE-marked and FDA-approved in vitro diagnostic assay used to select patients with metastatic melanoma for treatment with the selective BRAF inhibitor vemurafenib. We describe the pre-approval validation of this test in two external laboratories. Melanoma specimens were tested for BRAF V600 mutations at two laboratories with the: cobas BRAF Mutation Test; ABI BRAF test; and bidirectional direct sequencing. Positive (PPA) and negative (NPA) percent agreements were determined between the cobas test and the other assays. Specimens with discordant results were tested with massively parallel pyrosequencing (454). DNA blends with 5% mutant alleles were tested to assess detection rates. Invalid results were observed in 8/116 specimens (6·9%) with Sanger, 10/116 (8·6%) with ABI BRAF, and 0/232 (0%) with the cobas BRAF test. PPA was 97·7% for V600E mutation for the cobas BRAF test and Sanger, and NPA was 95·3%. For the cobas BRAF test and ABI BRAF, PPA was 71·9% and NPA 83·7%. For 16 cobas BRAF test-negative/ABI BRAF-positive specimens, 454 sequencing detected no codon 600 mutations in 12 and variant codon 600 mutations in four. For eight cobas BRAF test-positive/ABI BRAF-negative specimens, four were V600E and four V600K by 454 sequencing. Detection rates for 5% mutation blends were 100% for the cobas BRAF test, 33% for Sanger, and 21% for the ABI BRAF. Reproducibility of the cobas BRAF test was 111/116 (96%) between the two sites. It is feasible to evaluate potential companion diagnostic tests in external laboratories simultaneously to the pivotal clinical trial validation. The health authority approved assay had substantially better performance characteristics than the two other methods. The overall success of the cobas BRAF test is a proof of concept for future biomarker development.
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影响因子：
11.5
作者：
Luke, Jason J.;Hodi, F. Stephen
通讯作者：
Hodi, F. Stephen
影响因子：
7.3
作者：
Angulo, B.;Suarez-Gauthier, A.;Sanchez-Cespedes, M.
通讯作者：
Sanchez-Cespedes, M.
影响因子：
3.7
作者：
Yancovitz M;Litterman A;Yoon J;Ng E;Shapiro RL;Berman RS;Pavlick AC;Darvishian F;Christos P;Mazumdar M;Osman I;Polsky D
通讯作者：
Polsky D
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4.1
作者：
Angulo, Barbara;Garcia-Garcia, Elena;Lopez-Rios, Fernando
通讯作者：
Lopez-Rios, Fernando
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64.8
作者：
Margulies, M;Egholm, M;Rothberg, JM
通讯作者：
Rothberg, JM