Simultaneous identification of 36 mutations in KRAS codons 61and 146, BRAF, NRAS, and PIK3CA in a single reaction by multiplex assay kit

BMC Cancer

Hideaki Bando⁴  Takayuki Yoshino⁴  Eiji Shinozaki²  Tomohiro Nishina⁷  Kentaro Yamazaki⁵  Kensei Yamaguchi³  Satoshi Yuki⁹  Shinya Kajiura⁶  Satoshi Fujii¹  Takeharu Yamanaka⁸  Katsuya Tsuchihara⁸  Atsushi Ohtsu⁸ 

[1] Pathology Division, Research Center for Innovative Oncology, National Cancer Center Hospital East, Chiba, Japan

[2] Cancer Institute Hospital of Japanese Foundation for Cancer Research, Tokyo, Japan

[3] Division of Gastroenterology, Saitama Cancer Center, Saitama, Japan

[4] Department of Gastroenterology and Gastrointestinal Oncology, National Cancer Center Hospital East, 6-5-1 Kashiwanoha, Kashiwa, Chiba 277-8577, Japan

[5] Division of Gastrointestinal Oncology, Shizuoka Cancer Center, Shizuoka, Japan

[6] The Third Department of Internal Medicine, University of Toyama, Toyama, Japan

[7] National Hospital Organization Shikoku Cancer Center, Ehime, Japan

[8] Exploratory Oncology Research & Clinical Trial Center, National Cancer Center, Chiba, Japan

[9] Department of Gastroenterology, Hokkaido University Graduate School of Medicine, Hokkaido, Japan

关键词: Epidermal growth factor;    PIK3CA;    NRAS;    BRAF;    KRAS;    Luminex assay;   

Others  :  1079586
DOI  :  10.1186/1471-2407-13-405

 received in 2013-04-30, accepted in 2013-08-30,  发布年份 2013

【 摘 要 】

Background

Retrospective analyses in the West suggest that mutations in KRAS codons 61 and 146, BRAF, NRAS, and PIK3CA are negative predictive factors for cetuximab treatment in colorectal cancer patients. We developed a novel multiplex kit detecting 36 mutations in KRAS codons 61 and 146, BRAF, NRAS, and PIK3CA using Luminex (xMAP) assay in a single reaction.

Methods

Tumor samples and clinical data from Asian colorectal cancer patients treated with cetuximab were collected. We investigated KRAS, BRAF, NRAS, and PIK3CA mutations using both the multiplex kit and direct sequencing methods, and evaluated the concordance between the 2 methods. Objective response, progression-free survival (PFS), and overall survival (OS) were also evaluated according to mutational status.

Results

In total, 82 of 83 samples (78 surgically resected specimens and 5 biopsy specimens) were analyzed using both methods. All multiplex assays were performed using 50 ng of template DNA. The concordance rate between the methods was 100%. Overall, 49 (59.8%) patients had all wild-type tumors, 21 (25.6%) had tumors harboring KRAS codon 12 or 13 mutations, and 12 (14.6%) had tumors harboring KRAS codon 61, KRAS codon 146, BRAF, NRAS, or PIK3CA mutations. The response rates in these patient groups were 38.8%, 4.8%, and 0%, respectively. Median PFS in these groups was 6.1 months (95% confidence interval (CI): 3.1–9.2), 2.7 months (1.2–4.2), and 1.6 months (1.5–1.7); median OS was 13.8 months (9.2–18.4), 8.2 months (5.7–10.7), and 6.3 months (1.3–11.3), respectively. Statistically significant differences in both PFS and OS were found between patients with all wild-type tumors and those with KRAS codon 61, KRAS codon 146, BRAF, NRAS, or PIK3CA mutations (PFS: 95% CI, 0.11–0.44; P < 0.0001; OS: 95% CI, 0.15–0.61; P < 0.0001).

Conclusions

Our newly developed multiplex kit is practical and feasible for investigation of a range of sample types. Moreover, mutations in KRAS codon 61, KRAS codon 146, BRAF, NRAS, or PIK3CA detected in Asian patients were not predictive of clinical benefits from cetuximab treatment, similar to the result obtained in European studies.

【 授权许可】

   
2013 Bando et al.; licensee BioMed Central Ltd.

【 图 表 】

Figure 1.

Figure 2.

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