Abstract LB-B11: Development and validation of ColoScape™: A new colorectal cancer mutation detection assay™

Introduction: Colorectal cancer is a highly preventable disease as early detection increases rates of patient survival to near 100%. Herein we report the development and validation of ColoScape™, a highly sensitive test powered by XNA technology to detect novel multigene mutation biomarkers in CRC by real-time PCR. The assay allows the sensitive detection of the presence or absence of mutations in the targeted regions of the genes interrogated in tissue biopsy (FFPE) and plasma samples.Methodology: The high sensitivity of this multigene biomarker assay is achieved due to xenonucleic acid (XNA) probe technology. XNA probes are novel backbone modified oligomers with natural nucleoside bases (A, T, C and G) that hybridize by Watson-Crick base pairing to natural DNA and RNA with much higher binding affinity. XNA probes are designed that bind to the selected wild-type sequences at the respective genetic loci in the target genes. These XNA probes cannot be extended by DNA polymerase thus suppress amplification of WT DNA templates and only allow amplification of the target mutant DNA templates in the sample. For each of the selected mutation sites, primers and FAM-labeled TaqMan probes were designed and tested together with the selected XNA oligomers. An internal PCR control selected in the Human -Actin (ACTB) gene was employed utilizing an HEX-labeled TaqMan probe.Results: The ColoScape™ kit was demonstrated to have robust analytical performance and clinical accuracy for FFPE, stool and plasma samples. The rapid, precise and sensitive molecular assay for mutation detection in colon cancer has key benefits listed below. The assays showed a sensitivity of as low as 0.1% mutation in 5-10 ng of WT DNA/well. No cross-reactivity was observed with wild-type up to 320ng purified gDNA and up to 20ng FFPE DNA per reaction demonstrating high specificity of the ColoScape™ assay. Intra-assay, inter-assay, lot-to-lot and operator variation comparison showed CV% between 3% and 8%. Excluding pre-cancer samples, the assay clinical specificity and sensitivity were 95% and 100%, respectively. Pre-cancer detection sensitivity was 60% and 62.5% for stool samples. For tested FFPE clinical samples, the assay specificity and sensitivity were 95% and 91% respectively while the assay clinical specificity and sensitivity were both 100% for plasma samples.Conclusion: The ColoScape™ Colorectal Cancer Mutation Detection qPCR assay is shown to be a sensitive tool intended to aide in the early detection of colorectal cancer, disease monitoring and therapeutic interventions. Citation Format: Michael J. Powell, Elena Peletskaya, Qing Sun, Larry Pastor, Aiguo Zhang, Kamila Koprowska, Walter Bodmer. Development and validation of ColoScape™: A new colorectal cancer mutation detection assay™ [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2017 Oct 26-30; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Ther 2018;17(1 Suppl):Abstract nr LB-B11.