188P - A clinical validation study of RASKET-B: A multiple detection kit for RAS and BRAF gene mutations in colorectal cancer

Abstract

Background: The detection of RAS and BRAF gene mutations has been essential to determine the treatment strategy for metastatic colorectal cancer (CRC). A multiplex kit simultaneously detecting both gene mutations is required, although few genetic assays with high-quality assurance are established. We evaluated RAS/BRAF KIT (RASKET-B), a multiplex assay using PCR-reverse sequence specific oligonucleotide (PCR-rSSO) and xMAP technology to concurrently detect 48 types of RAS mutations as well as the BRAF V600E mutation in a short turnaround time (4.5 h/96 specimens). Methods: Formalin-fixed paraffin-embedded tissues were obtained from 309 consenting patients with histologically-confirmed CRC. The primary endpoint was the concordance rate (CR) between RAS mutations identified with RASKET-B and a PCR-rSSO method kit (RASKET) and between BRAF mutations identified with direct sequencing (DS), respectively. As the secondary endpoints, we evaluated the CR between RASKET-B and DS for RAS mutations and between RASKET-B and the pyrosequencing (PYRO) method for BRAF V600E mutation. Results: We registered 302 samples, all of which were tested using the RASKET-B and reference methods. In RAS genes, 142 samples (47.0%) were detected by RASKET-B: 114 KRAS exon 2 and 28 other RAS mutations. The CR between RASKET-B and RASKET or RASKET-B and DS were 100% (95%CI: 99-100%) and 97.4% (95%CI: 95-99%), respectively. In BRAF genes, 18 samples (6.0%) were detected using RASKET-B. The CR between RASKET-B and DS or RASKET-B and PYRO were 100% (95%CI: 99-100%) and 99.7% (95%CI: 98-100%), respectively. The result of RASKET-B was consistent with that of DS. We also analyzed RAS and BRAF mutation ratios for each tumor cell ratio and tumor cell area ratio (0-25%, 25-50%, 50-75%, 75-100% respectively) and report it. Conclusions: RASKET-B provides rapid, precise and simultaneous detections of RAS and BRAF mutations in CRC. Clinical trial indentification: Study ID: UMIN000022742. Legal entity responsible for the study: Medical & Biological Laboratories Co., Ltd. Funding: Medical & Biological Laboratories Co., Ltd. Disclosure: W. Okamoto: MSD, K. Muro: Takeda, Chugai Pharma, Yakult Honsha, Merck Serono, Taiho Pharmaceutical, Lilly., H. Taniguchi: Takeda, Chugai Pharma, Merck Serono, Taiho Pharmaceutical, Bayer, Lilly Japan, Yakult Honsha., H. Hara: Chugai Pharma, Taiho Pharmaceutical, Merck Serono, Yakult Honsha, Lilly, Ono Pharmaceutical, Chugai Pharma., T. Satoh: Bayer Yakuhin, Lilly, Ono Pharmaceutical, Takara Bio, Merck Serono, Chugai Pharma, Bristol-Myers Squibb, Takeda, Yakult Honsha. All other authors have declared no conflicts of interest.