Performance comparison of three BRAF V600E detection methods in malignant melanoma and colorectal cancer specimens.
Performance comparison of three BRAF V600E detection methods in malignant melanoma and colorectal cancer specimens.
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DOI：
10.1007/s13277-014-2711-5
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发表时间：
2015-02
期刊：
影响因子：
--
通讯作者：
Knappskog, Stian
中科院分区：
文献类型：
--
作者：
Loes, Inger Marie;Immervoll, Heike;Angelsen, Jon-Helge;Horn, Arild;Geisler, Juergen;Busch, Christian;Lonning, Per Eystein;Knappskog, Stian
个性化的癌症护理需要可靠的生物标志物。虽然BRAF V600 E突变在临床中实施，但迄今为止尚未建立其检测方法作为参考。我们的目的是对目前用于临床样本中V600 E检测的三种方法进行全面比较。我们通过桑格测序和基于单探针的高分辨率熔解分析（LightMix）分析了127例恶性黑色素瘤（77例患者）和141例结直肠癌患者的389个肿瘤（383个肝转移和6个原发性肿瘤）的基因组DNA。使用V600 E特异性抗体VE 1通过免疫组织化学（IHC）分析了来自这些病变亚组（分别为n = 77和304）的福尔马林固定石蜡包埋（FFPE）组织。在野生型DNA中V600 E突变DNA的稀释系列中，LightMix测定的检测限为1：1000突变等位基因，而桑格测序的检测限为1：10。与此一致，我们通过LightMix测定与桑格测序相比检测了另外15个突变的黑色素瘤样品和另外两个突变的转移性结肠直肠癌样品。对于黑色素瘤样本，我们观察到基于DNA的方法和IHC分析之间的高度一致性。然而，在结直肠样本中，IHC表现不佳，12个样本被IHC专门评分为V600 E阳性，9个样本被IHC专门评分为V600 E阴性。总之，VE 1抗体不能用于结直肠癌样品的临床试验。对于黑色素瘤样本，免疫组化可能是有用的筛选工具，指导进一步的分析方法。本文的在线版本（doi：10.1007/s13277-014-2711-5）包含补充材料，可供授权用户使用。
Personalized cancer care requires reliable biomarkers. While the BRAF V600E mutation is implemented in the clinic, no method for its detection has so far been established as reference. We aimed to perform a comprehensive comparison of three methods currently being used for V600E detection in clinical samples. We analysed genomic DNA from 127 malignant melanomas (77 patients) and 389 tumours from 141 colorectal cancer patients (383 liver metastases and 6 primary tumours) by Sanger sequencing and a single probe-based high-resolution melting assay (LightMix). Formalin-fixed paraffin-embedded (FFPE) tissue from a subset of these lesions (n = 77 and 304, respectively) was analysed by immunohistochemistry (IHC) using the V600E-specific antibody VE1. In a dilution series of V600E-mutated DNA in wild-type DNA, the detection limit for the LightMix assay was 1:1000 mutated alleles while it was 1:10 for Sanger sequencing. In line with this, we detected 15 additional mutated melanoma samples and two additional mutated metastatic colorectal cancer samples by the LightMix assay compared to Sanger sequencing. For the melanoma samples, we observed high concordance between DNA-based methods and analysis by IHC. However, in colorectal samples, IHC performed poorly with 12 samples being scored as V600E positive exclusively by IHC and nine samples being scored as V600E negative exclusively by IHC. In conclusion, the VE1 antibody is not recommendable for clinical tests of colorectal cancer samples. For melanoma samples, IHC may be useful as a screening tool guiding further analytical approaches. The online version of this article (doi:10.1007/s13277-014-2711-5) contains supplementary material, which is available to authorized users.
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影响因子：
3.7
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
通讯作者：
de Castro DG
影响因子：
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影响因子：
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影响因子：
6.4
作者：
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影响因子：
4.1
作者：
Colomba, Emeline;Helias-Rodzewicz, Zofia;Emile, Jean-Francois
通讯作者：
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