Abstract 772: Performance characteristics of the Oncomine™ Dx Target Test for the detection of IDH1 R132 variants from formalin-fixed, paraffin-embedded (FFPE) cholangiocarcinoma samples

Background: Cholangiocarcinoma (CC) is an aggressive bile-duct cancer that affects 8000 people per year in the US. This silent disease manifests symptoms only in advanced stages, resulting in a 5-year overall survival rate of < 10%. Mutations in the metabolic enzyme isocitrate dehydrogenase 1 (IDH1) have been observed in ~20% of CC. These mutations are implicated in promoting tumorigenesis potentially by generation of an ‘oncometabolite' resulting in changes in DNA methylation. There are currently no approved targeted therapies for CC. Ivosidenib, developed by Agios Pharmaceuticals, is a first-in-class, oral, targeted, small-molecule inhibitor of the mutant IDH1 protein currently being evaluated in patients with advanced CC with an IDH1 mutation (ClarIDHy). The Oncomine Dx Target Test (ODxTT) is an FDA approved in vitro diagnostic test for the detection of somatic alterations in human DNA and RNA isolated from FFPE non-small cell lung cancer samples. Thermo Fisher Scientific has partnered with Agios Pharmaceuticals to validate the ODxTT for the detection of IDH1 R132 mutations in CC. Here we present the results from our analytical studies which establish sensitivity, reproducibility and accuracy of the IUO ODxTT to detect 5 IDH1 R132 variants from FFPE CC specimens. Methods: Analytical studies were performed to determine the limit of detection (LOD) and reproducibility of the assay to detect the variants of interest. LOD was determined for each of the 5 variants of interest using either FFPE clinical sample or FFPE cell line blends. Reproducibility of the assay was evaluated by testing clinical samples blends for 3 of 5 variants at ~2x LOD across 3 sites, 6 operators and 3 lots of reagents. Analytical accuracy of the ODxTT was determined by testing over 340 clinical samples with both the ODxTT and a validated orthogonal method. The orthogonal method was Sanger sequencing coupled with Applied Biosystems™ Minor Variant Finder Software to allow calling minor variants down to 5%. Results: 1. LOD of the ODxTT for all 5 variants of interest (R132C, R132G, R132S, R132L and R132H) was determined to be at an allelic frequency (AF) between 3.7% - 5.5%. 2. Reproducibility of the ODxTT to detect R132C, R132G and R132L variants from clinical sample blends was 100% when each variant was tested at an AF of ~2x LOD. 3. Analytical accuracy: The positive percent agreement (PPA) of the ODxTT to Sanger sequencing was 99.4% (163/164) and the negative percent agreement (NPA) wa