Background: Access to molecular testing for metastatic NSCLC (mNSCLC) patients has been improved by circulating tumor DNA (ctDNA) based liquid biopsies (LB), which can obviate invasive procedures, expedite results, and enable serial testing. Emerging clinical applications for LB include therapeutic efficacy monitoring and minimal residual disease (MRD) detection, which demand a ctDNA assay with high sensitivity, specificity, and appropriate genomic coverage. Here we demonstrate that SafeSEQ next-generation sequencing (NGS) LB delivers equivalent performance to OncoBEAM digital PCR, a sensitive ctDNA approach that has been extensively clinically validated in pivotal trials (e.g., AURA, TIGER-X). Importantly, SafeSEQ delivers significantly expanded genomic coverage to address the need for expanding targeted therapy indications, monitoring treatment response, and detecting MRD. Methods: Whole blood samples (n=176) were collected from mNSCLC patients prior to/during treatment or at disease progression and transported to a CLIA laboratory for OncoBEAM analysis to detect mutations in EGFR (exon 19 del, L858R, T790M, C797S), KRAS (codons 12, 13, 61) and BRAF V600E. Replicate plasma aliquots were analyzed with SafeSEQ to interrogate clinically relevant regions in BRAF, EGFR, ERBB2, KRAS, MET, NRAS, PIK3CA, and TP53. Mutation level concordance between the methods was assessed, where only genomic alterations interrogated by both platforms were considered. For mutations detected by both methods, correlation analysis of mutant allelic frequency (MAF) was performed. Results: Concordance analysis of the mutation results from OncoBEAM and SafeSEQ testing of 176 replicate patient samples demonstrated an overall percent agreement (OPA) of 99.6%, with a positive percent agreement (PPA) of 78.1% and a negative percent agreement (NPA) of 99.9%. The mean MAF for discordant mutations (n=16) was 0.06% (range: 0.04-0.12%). When considering mutations with MAF >0.1%, PPA and OPA increased to 96.0% and 99.9%, respectively. MAF levels for mutations detected by both methods demonstrated a strong linear correlation (R2=0.98). Of 124 patient samples having no mutation detected by OncoBEAM, 75 (60%) showed ≥1 alteration with SafeSEQ. Panel-wide, 76% and 30% of all mutations were detected at <1% and <0.1% MAF, respectively. Conclusions: SafeSEQ demonstrates clinical sensitivity comparable to OncoBEAM, with a strong positive correlation between MAF values across a broad dynamic range. SafeSEQ also provides expanded coverage across broader genomic regions, which - when combined with robust clinical performance - should better inform treatment selection, improve high resolution monitoring of therapeutic efficacy, and enable MRD detection and surveillance for NSCLC patients.

Citation Format: Hillary Sloane, Priya Sathyanarayan, Daniel Edelstein, Frederick Jones, Jennifer Preston, Sam Wu, Jenna Los, Lara Duchstein, Johannes Fredebohm, Katharina Wichner, Denise Heim, Frank Holtrup, Hannah Quinn, David Feller-Kopman. Clinical evaluation of NGS-based liquid biopsy genotyping in non-small cell lung cancer (NSCLC) patients [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr LB053.