Department of Clinical Laboratory, Huashan Hospital, Fudan University, 200040, China.
Baoshan Hospital of Integrated Traditional Chinese and Western Medicine, Shanghai, 201999, China.
Biomed Res Int. 2018 Oct 10;2018:8727941. doi: 10.1155/2018/8727941. eCollection 2018.
KRAS genotyping in tumor samples is a decisive clinical test for the anti-EGFR therapy management. However, the complexity of KRAS mutation landscape across different cancer types and the mosaic effect caused by cancer cellularity and heterogeneity make the choice of KRAS genotyping method a challenging topic in the clinical practice.
We depicted the landscape of somatic KRAS mutation in 7,844 primary tumors and 10,336 metastatic tumors across over 30 types of cancer using the Cancer Genome Atlas (TCGA) and Integrated Mutation Profiling of Actionable Cancer Targets (MSKCC-IMPACT) databases, respectively. A snapback primer assay based on melting curve analysis was developed to detect the most common somatic mutations in KRAS codons 12 and 13. The sensitivity and accuracy of the method was validated by genotyping 100 colorectal cancer (CRC) samples, in comparison with Sanger sequencing and T-A cloning sequencing.
Pancreas adenocarcinoma (somatic mutation frequency 90.6%), colorectal adenocarcinoma (42.5%), and lung adenocarcinoma (32.6%) are the top three most KRAS mutant primary cancer types. The metastatic tumors showed a higher prevalence (90.99% versus 66.31%) and diversity of KRAS mutation compared with the primary tumors. Mutations in codons 12 and 13 are the predominant genetic alteration in KRAS (84.15% for TCGA and 86.13% for MSK-IMPACT). Moreover, KRAS mutation is highly correlated with the overall survival of patients with metastatic cancer. The snapback primer assay showed a more favorable performance in enriching and detecting the KRAS codon 12 and 13 mutation (1% mutation load) compared with Sanger sequencing (20% mutation load and 7% false-negative rate).
KRAS mutation pattern is highly diverse among different cancer types and is associated with the survival of patients with metastatic cancers. The snapback primer assay is a reliable, sensitive method to detect the major mutant KRAS alleles, which might facilitate the effective cancer treatment decisions.
KRAS 基因分型是肿瘤样本中用于决定抗 EGFR 治疗管理的关键临床检测。然而，不同癌症类型中 KRAS 突变景观的复杂性以及由肿瘤细胞异质性和镶嵌性引起的马赛克效应，使得 KRAS 基因分型方法的选择成为临床实践中的一个具有挑战性的课题。
我们分别使用癌症基因组图谱（TCGA）和综合 actionable 癌症靶点突变分析（MSKCC-IMPACT）数据库，描绘了 7844 例原发性肿瘤和 10336 例转移性肿瘤中体细胞 KRAS 突变的景观。开发了一种基于熔解曲线分析的 snapback 引物检测方法，用于检测 KRAS 密码子 12 和 13 中最常见的体细胞突变。通过对 100 例结直肠癌（CRC）样本进行基因分型，与 Sanger 测序和 T-A 克隆测序进行比较，验证了该方法的灵敏度和准确性。
胰腺腺癌（体细胞突变频率 90.6%）、结直肠腺癌（42.5%）和肺腺癌（32.6%）是 KRAS 突变最常见的三种原发性癌症类型。与原发性肿瘤相比，转移性肿瘤中 KRAS 突变的发生率（90.99%比 66.31%）和多样性更高。密码子 12 和 13 的突变是 KRAS 中的主要遗传改变（TCGA 为 84.15%，MSK-IMPACT 为 86.13%）。此外，KRAS 突变与转移性癌症患者的总生存期高度相关。与 Sanger 测序（突变负荷 20%，假阴性率 7%）相比，snapback 引物检测法在富集和检测 KRAS 密码子 12 和 13 突变（1%突变负荷）方面表现出更好的性能。
KRAS 突变模式在不同癌症类型之间高度多样化，与转移性癌症患者的生存相关。Snapback 引物检测法是一种可靠、敏感的检测主要突变 KRAS 等位基因的方法，可能有助于做出有效的癌症治疗决策。
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