Abstract 2409: Ultrasensitive detection of genomic alterations in cell-free DNA by Droplet Digital PCR

Abstract

Abstract As cancer cells die, they release small pieces of DNA into the bloodstream. These molecules of DNA are called circulating tumor DNA (ctDNA). ctDNA carry somatic aberrations that are found in the originating tumor tissue and can serve as excellent biomarkers. They can be extremely valuable as tools for disease monitoring, and samples for predictive and prognostic biomarkers. However, limited quality and quantity of formalin-fixed paraffin-embedded (FFPE) tissues and circulating cell-free DNA (cfDNA) samples available from patients pose a big challenge to performing DNA quantification, mutation profiling and copy number variation analysis. We explored the feasibility of using Droplet Digital PCR (ddPCR) technology to address this challenge. Using ddPCR, we studied the ability to perform high-sensitivity mutation analysis and accurate quantification of allele frequencies, and compared this approach to alternate methods such as next-generation sequencing (NGS). For mutation profiling, we examined a total of 22 mutation detection ddPCR assays across 7 oncogenes - KRAS, PIK3CA, EGFR, BRAF, NRAS, cKIT and AKT1. The assay sensitivity for the majority of mutation detection assays was found to be approximately 0.03%. We have identified/reconfirmed various mutations using matching FFPE/plasma samples from patients in various cancer types. Moreover, we have adapted the previously published HER2 CNV ddPCR assay to detect HER2 copy number variation in cfDNA. We will present data showing validation of this platform using commercially available DNA control, archival FFPE tissue and cfDNA samples and demonstrate improved sensitivity and specificity. The platform is being routinely used in our labs to analyze samples from multiple clinical trials. These efforts enable the development of a non-invasive method to overcome existing challenges to provide molecular understanding of patient's tumor evolution, and aid in the development of personalized therapies for cancer patients. Citation Format: Nga Wan Rachel Tam, Teiko Sumiyoshi, Rajesh Patel, Sundari Sarma, Astrid Kiermaier, Rajiv Raja. Ultrasensitive detection of genomic alterations in cell-free DNA by Droplet Digital PCR. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2409. doi:10.1158/1538-7445.AM2015-2409