Abstract
Genetic screening of somatic mutations in circulating free DNA (cfDNA) opens up new opportunities for personalized medicine. In this study, we aim to illustrate the implementation of NGS-based liquid biopsy in clinical practice for the detection of somatic alterations in selected genes. Our work is particularly relevant for the diagnosis and treatment of NSCLC. Beginning in 2020, we implemented the use of Roche’s Avenio ctDNA expanded panel in our diagnostic routine. In this study, we retrospectively review NGS-based clinical genetic tests performed in our laboratory, focusing on key analytical parameters. Avenio ctDNA kits demonstrated 100% sensitivity in detecting single nucleotide variants (SNVs) at >0.5% variant allele frequency (VAF), and high consistency in reproducibility. Since 2020, we performed cfDNA genotyping test in 86 NSCLC patients, and we successfully sequenced 96.5% (83/86) of samples. We observed consistency in sequencing performance based upon sequencing depth and on-target rate. At least one gene variant was identified in 52 samples (63%), and one or more actionable variants were detected in 21 out of 83 (25%) of analysed patients. We demonstrated the feasibility of implementing an NGS-based liquid biopsy assay for routine genetic characterization of metastatic NSCLC patients.
Highlights
Recent advances in molecular biology and cancer genomics improved knowledge of the molecular landscape of cancer, highlighting its heterogeneity and, in parallel, introducing the era of targeted therapies and personalized medicine [1]
The Avenio circulating tumor DNA (ctDNA) Expanded kit consists in a 77-panel genes containing 17 cancer driver genes in U.S National Comprehensive Cancer Network (NCCN) guidelines and additional 60 frequently mutated genes mainly selected for non-smallcell-lung cancer (NSCLC) and colorectal cancer
The panel exploits unique molecular indexes (UMIs) to tag individual fragments in order to provide a better quantification of copy number and in silico error suppression, and it is capable of detecting four mutation classes: single nucleotide variants (SNVs), fusions, copy number variants (CNVs), and indel mutations (InDels)
Summary
Recent advances in molecular biology and cancer genomics improved knowledge of the molecular landscape of cancer, highlighting its heterogeneity and, in parallel, introducing the era of targeted therapies and personalized medicine [1]. Under the pressure of targeted therapies, cancer cells may select for specific somatic mutations that confer drug resistance, allowing them to survive and drive cancer progression [2]. In this context, early assessment of somatic mutations acquired during disease progression became crucial to offer specific therapeutic options for newly activated signalling pathways or for escape mechanisms that require alternative specific inhibitors. The genetic characterization of circulating free DNA (cfDNA) has emerged as a non-invasive method for the detection of somatic mutations for research, diagnosis, and prognosis of solid tumors. cfDNA analysis can potentially identify actionable alterations in tumor-derived DNA and capture intra-tumor heterogeneity [3]
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