Abstract 3069: One-step sensitive PNA mediated real-time PCR clamping assay to detect PIK3CA mutations

Abstract

Abstract Phosphatidylinositol 3-kinases (PI3K) are important regulators of cellular growth, transformation, adhesion, apoptosis, survival and motility. Mutations in the PIK3CA oncogene encoding PI3K have been frequently found in various human cancers. The presence of PIK3CA mutations in a tumor has been suggested as a prognostic factor and may predict responses to select treatments. Therefore, detection of PIK3CA mutations may help predict drug responses and prognosis of cancer patients. We have developed a highly sensitive and simple method to detect 11 mutations in exons 9 and 20 of PIK3CA using one-step PNA-mediated real-time PCR clamping, PNAClampTM. PNAClampTM relies on unique properties of PNA probes that PNA oligomers are not recognized by DNA serving as sequence-selective clamps during PCR amplification. This method can also detect 11 mutations through only three tube reactions each with 10 ng of the target DNA. PNAClampTM is simple and reliable with a detection limit of approximately 0.1% mutant alleles using 10 to 50 ng of normal DNA as a template. The turnaround time for performing this assay was only 2 hours. In clinical studies with breast cancer patients in Korea, PIK3CA mutations were detected utilizing PNAClampTM in 12 (35.3%) of clinical samples, and the results were compared with direct sequencing method. Among 34 patients, 1 (2.94%) had mutations in codon 542, 6 (17.65%) had mutations in codon 545 and 5 (14.71%) had mutations in codon 1047. The results will be discussed in detail in this report including the method development and optimization. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3069. doi:10.1158/1538-7445.AM2011-3069