Analytic validation of anFGFR-focused cell-free DNA liquid biopsy assay (FGFR-Dx)

Abstract

ABSTRACTCommercial liquid biopsy assays are routinely used by oncologists to monitor disease response and resistance to therapy. Additionally, in cases where tumor tissue is not available, clinicians may rely on cell-free DNA (cfDNA) testing as a surrogate for comprehensive tumor testing. While some gene rearrangements are well detected, current commercial liquid biopsy assays exhibit low sensitivity for fibroblast growth factor receptor (FGFR) rearrangements.FGFRsare altered in ∼2.5% of all cancers, includingFGFR2rearrangements in 10% of intrahepatic cholangiocarcinoma andFGFR3point mutations and rearrangements in 10-15% of urothelial carcinoma. Therefore, we developed and analytically validated FGFR-Dx, anFGFR-focused cfDNA assay with improved sensitivity forFGFRrearrangements. FGFR-Dx comprehensively targets the introns inFGFR1-3previously shown to be involved in gene fusions as well as all coding exons. CustomFGFRsynthetic reference standards representing both single nucleotide variants (SNVs) and gene rearrangements were utilized at a range of variant frequencies and revealed a detection limit of 0.5% with sensitivities of 97.2% and 92.9% for SNVs and rearrangements, respectively. Furthermore, FGFR-Dx detected rearrangements and identified the intronic breakpoints from cfDNA collected from 13 of 15 patients with known FGFR fusions.