Single Copy qPCR-Based Detection of BRAF and KRAS Mutations

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July 30, 2012

Vladimir Makarov, Chief Scientific Officer, Swift Biosciences Inc

Abstract
The development of highly sensitive genotyping assays that are suitable for clinical diagnostics opens new opportunities for the detection, assessment, and management of cancer. Anticipated uses for these assays include profiling tissue biopsies, circulating tumor cells (CTCs), and detecting mutations in circulating cell-free nucleic acids. Swift Biosciences™ has developed myT™ Primers which have unique structural and thermodynamic properties that make them highly sensitive to mismatch discrimination. A myT Primer assay for BRAF V600E/K mutations demonstrated single mutant copy sensitivity in a background of 14,000 wild-type genomic DNA copies. This assay also demonstrated high specificity as indicated by a low percentage (< 3%) of amplification events from 14,000 wild-type genomic copies. myT Primer assays are compatible with multiple qPCR instruments and reaction mixes. myT Primer qPCR assays for seven common KRAS mutations demonstrated similar sensitivity and specificity. When compared to a leading commercially available KRAS mutation qPCR test kit, several orders of improved specificity were observed. The extreme selectivity of myT Primer assays will be especially useful for detection of mutations present at ultra-low copy number and for genotyping difficult samples such as needle biopsies, CTCs, and serum, resulting in better detection, evaluation, and monitoring of cancer.

Drug DiscoveryGenomicsInformatics

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