Abstract 1508: A highly sensitive and reliable methodology (ARMS-PM system) for detection of K-ras mutations in FFPE samples

Abstract

The association between the activating mutations of K-ras gene and poor clinical response to current targeted therapies against epidermal growth factor receptor (EGFR ) such as cetuximab (Erbitux®) and panitumumab (Vectibix®) in colorectal cancer (CRC) has been well documented. Therefore detection of K-ras mutation status will guide effective therapeutical options for patients with CRC. Here we developed a new method for the rapid and reliable detection of K-ras mutations by using a combination of the modified mutation-specific ARMS primers, peptide nucleic acid (PNA) clamping and taqman-MGB probes, named ARMS-PM system. In the ARMS-PM system, the wild-type K-ras DNA is completely blocked by PNA at a higher annealing temperature, and the mutant gene is selectively amplified by the modified ARMS primers and the amplicon extending signals are collected by the taqman-MGB probe at the same time. The ARMS-PM system can detect approximately 10 copies of the mutant gene stably in somatic cells at the background of 10ng genomic DNA. In addition, the premixed reaction was introduced into this system to allow the assay to be completed in only one step of sample loading, which minimizes the potential contamination significantly. By using this assay, we analyzed the K-ras mutations in 1014 Chinese CRC and 205 Chinese lung cancer FFPE samples in comparison with Sanger sequencing. Our results showed that the positive rates were 38.6% and 10.2% 38.0% in CRC and lung cancer samples respectively, which were higher than that measured by Sanger sequencing (38.0% and 7.1%). The coincidence rates of the two methods were 97.3% for the positive samples and 97.7 for the negative samples. The total coincidence rate was 97.8%. In conclusion, our study provides a basis for the detection of somatic mutations in K-ras in a routine clinical setting. The ARMS-PM assay allows the detection of the activating K-ras mutations in FFPE samples with better sensitivity and feasibility than the current assays used in clinic. <!–EndFragment–> Citation Format: Wei-Tao Duan, Xiujuan Wang, Lihong Hu, Feng Zhu, Dehua Yu. A highly sensitive and reliable methodology (ARMS-PM system) for detection of K-ras mutations in FFPE samples. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 1508. doi:10.1158/1538-7445.AM2014-1508