Abstract 4936: Development of NTRK reference materials for global assay standardization

Abstract

Abstract Tropomyosin receptor kinases (TRK) are a family of transmembrane proteins (TRKA, B, and C) encoded by NTRK1, NTRK2 and NTRK3 genes. Fusions involving NTRK genes result in uncontrolled TRK signaling and oncogenesis. Cancers driven by TRK fusions are rare but occur in a broad range of tumor types in both pediatric and adult patients. Accurate testing for NTRK fusions has become critically important because of precision biopharmaceutical treatments, such as Larotrectinib. This drug, which was granted Breakthrough Therapy Designation by the FDA and is currently undergoing review, is a promising treatment option for patients with locally advanced or metastatic solid tumors harboring NTRK fusions. NGS based assays have the potential to accurately detect NTRK fusions; however, the majority of tests currently available are laboratory-developed tests, which vary substantially in their workflow, chemistry, sensitivity, and bioinformatics analyses. In order to achieve precision diagnostics for personalized therapy, testing standardization is needed worldwide. Towards this end, we developed Seraseq® FFPE NTRK Fusion RNA Reference Material for standardization of NTRK fusion testing by targeted NGS panels. The reference material contains 15 clinically-relevant NTRK fusions, selected based on prevalence data, in a single reference sample. The five fusion RNAs each for NTRK1, NTRK2, and NTRK3 are all incorporated in the well characterized GM24385 human reference cell line. The reference material challenges detection across multiple break points for each NTRK gene, as well as 12 different amino terminal fusion partners. Each fusion is quantified by digital PCR with concentrations ranging from 150 - 600 fusion copies per nanogram of total extractable RNA. The reference material was prepared in a formalin fixed paraffin embedded (FFPE) format, which allows evaluation of the full technical workflow. In our laboratory, one 10-micron FFPE curl yields more than 400 ng of extractable RNA when using the Agencourt Formapure® extraction kit. NGS analysis on the RNA-based ArcherDx FusionPlex Solid Tumor assay correctly identified all 15 fusions, demonstrating compatibility with a leading commercial assay. Highly characterized, patient-like reference materials help standardize testing across clinical trial sites and can speed test adoption, training and validation at clinical labs. The Seraseq FFPE NTRK Fusion RNA Reference Material is designed to aid standardization of NTRK fusion testing to identify patients that will benefit from new precision therapeutics. Note: This abstract was not presented at the meeting. Citation Format: Sebastian Bender, Patricia Carrigan, Kara O’Brien, Catherine Huang, Rajeswari Vemula, Praveena Kamineni, Omo Clement, Russell Garlick, Bharathi Anekella. Development of NTRK reference materials for global assay standardization [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 4936.