Ultra-deep sequencing confirms immunohistochemistry as a highly sensitive and specific method for detecting BRAFV600E mutations in colorectal carcinoma
Ultra-deep sequencing confirms immunohistochemistry as a highly sensitive and specific method for detecting BRAFV600E mutations in colorectal carcinoma
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DOI：
10.1007/s00428-013-1492-3
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发表时间：
2013-11-01
期刊：
影响因子：
3.5
通讯作者：
Rechsteiner, Markus P.
中科院分区：
文献类型：
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作者：
Roessle, Matthias;Sigg, Michele;Rechsteiner, Markus P.
激活BRAF （V600）突变是转移性结直肠癌（CRC）中公认的阴性预后生物标志物。最近开发的一种单克隆小鼠抗体（克隆VE1）已被证明可以通过免疫组化（IHC）可靠地检测BRAF （V600E）突变蛋白。在本研究中，我们旨在比较IHC、Sanger测序（SaS）和超深度测序（UDS）在结直肠癌中检测BRAF （V600E）突变的情况。在68个KRAS野生型crc队列中建立了VE1-IHC。通过SaS和UDS评估，VE1-IHC仅在3例已知BRAF （V600E）突变的患者中呈阳性。试验队列由265个非选择的连续CRC样本组成。265例VE1-IHC阳性39例（14.7%）。随机选择20例IHC阴性肿瘤的SaS显示BRAF野生型（20/20）。SaS检测ihc阳性24例（24/39;61.5%），SaS检测ihc阳性15例（15/39;38.5%）为BRAF野生型。UDS在这15例不一致病例中检测到13例BRAF （V600E）突变。在一个肿瘤中，突变频率低于我们的UDS阳性阈值，而在另一个肿瘤中，由于DNA量低，无法进行UDS。统计分析显示，与SaS和UDS联合结果相比，VE1-IHC和SaS的敏感性分别为100%和63%，特异性分别为95%和100%。我们的数据表明，使用抗braf (V600E) （VE1）抗体的UDS和IHC之间存在高度一致性。因此，VE1免疫组化是检测结直肠癌BRAF （V600E）突变的一种高度敏感和特异性的方法。
The activating BRAF (V600) mutation is a well-established negative prognostic biomarker in metastatic colorectal carcinoma (CRC). A recently developed monoclonal mouse antibody (clone VE1) has been shown to detect reliably BRAF (V600E) mutated protein by immunohistochemistry (IHC). In this study, we aimed to compare the detection of BRAF (V600E) mutations by IHC, Sanger sequencing (SaS), and ultra-deep sequencing (UDS) in CRC. VE1-IHC was established in a cohort of 68 KRAS wild-type CRCs. The VE1-IHC was only positive in the three patients with a known BRAF (V600E) mutation as assessed by SaS and UDS. The test cohort consisted of 265 non-selected, consecutive CRC samples. Thirty-nine out of 265 cases (14.7 %) were positive by VE1-IHC. SaS of 20 randomly selected IHC negative tumors showed BRAF wild-type (20/20). Twenty-four IHC-positive cases were confirmed by SaS (24/39; 61.5 %) and 15 IHC-positive cases (15/39; 38.5 %) showed a BRAF wild-type by SaS. UDS detected a BRAF (V600E) mutation in 13 of these 15 discordant cases. In one tumor, the mutation frequency was below our threshold for UDS positivity, while in another case, UDS could not be performed due to low DNA amount. Statistical analysis showed sensitivities of 100 % and 63 % and specificities of 95 and 100 % for VE1-IHC and SaS, respectively, compared to combined results of SaS and UDS. Our data suggests that there is high concordance between UDS and IHC using the anti-BRAF(V600E) (VE1) antibody. Thus, VE1 immunohistochemistry is a highly sensitive and specific method in detecting BRAF (V600E) mutations in colorectal carcinoma.