Abstract A2-41: The Amplinome™ Test – A droplet digital PCR (ddPCR) based pan-cancer test to assess gene amplification status in solid tumors

Abstract

Abstract Gene amplifications constitute >30% of all actionable mutation types present in cancer. However, despite their prevalence and the availability of targeted drug therapies, the development of molecular assays to detect gene amplifications has been limited by technical challenges such as limited DNA quantity and tumor heterogeneity. To address this, we describe the Amplinome™ test, a test that leverages the high precision and accuracy of droplet digital PCR (ddPCR) to determine the amplification status of 12 commonly amplified genes targeted by FDA approved drugs. Twelve taqman assays were designed to target AURKA, BRAF, CDK4, CDK6, EGFR, ERBB2/HER2, ERBB3/HER3, FGFR1, JAK2, MET, SRC, and VEGFA, and duplexed with the reference gene RNaseP (Epoch, Bothell, WA). Each duplexed assay was then assembled with 2 μL of purified DNA and analyzed via ddPCR (Bio-rad QX-200 ddPCR Platform, Hercules CA). Genes were identified as “amplified” if the observed target:reference ratio was >1.48 (p < 0.005), which adjusts the ASCO-CAP threshold of 2.2 recommended for HER2 amplification for a minimum tumor content of 40%. The Amplinome test identified 29 of 49 patients having actionable amplifications in at least one of the 12 assayed genes. In contrast, only 6 of 49 patients were identified as having amplifications in these genes through a commercially available sequencing test. An overall concordance rate of 90% (532/588 calls) between the two tests was observed. Discordant calls (56/588) arose from amplifications identified by the Amplinome test but not sequencing (55/56). One discordant call (HER3) was identified as amplified via sequencing, but confirmed to be unamplified via FISH. The observed 10-fold increase in sensitivity versus a sequencing-based clinical test suggests that the prevalence of actionable amplifications may be significantly underestimated in cancer. This sensitivity, combined with its ability to provide results within an actionable timeframe (<1 week), the underlines the potential value of Amplinome™ test in providing treatment options for oncologists and their patients. Citation Format: Amy Wong, Jennifer Pecson, Girish Putcha, Lincoln Nadauld, Austin P. So. The Amplinome™ Test – A droplet digital PCR (ddPCR) based pan-cancer test to assess gene amplification status in solid tumors. [abstract]. In: Proceedings of the AACR Special Conference on Translation of the Cancer Genome; Feb 7-9, 2015; San Francisco, CA. Philadelphia (PA): AACR; Cancer Res 2015;75(22 Suppl 1):Abstract nr A2-41.