RRC ID 68852
Author Paweletz CP, Sacher AG, Raymond CK, Alden RS, O'Connell A, Mach SL, Kuang Y, Gandhi L, Kirschmeier P, English JM, Lim LP, Jänne PA, Oxnard GR.
Title Bias-Corrected Targeted Next-Generation Sequencing for Rapid, Multiplexed Detection of Actionable Alterations in Cell-Free DNA from Advanced Lung Cancer Patients.
Journal Clin Cancer Res
Abstract PURPOSE:Tumor genotyping is a powerful tool for guiding non-small cell lung cancer (NSCLC) care; however, comprehensive tumor genotyping can be logistically cumbersome. To facilitate genotyping, we developed a next-generation sequencing (NGS) assay using a desktop sequencer to detect actionable mutations and rearrangements in cell-free plasma DNA (cfDNA).
EXPERIMENTAL DESIGN:An NGS panel was developed targeting 11 driver oncogenes found in NSCLC. Targeted NGS was performed using a novel methodology that maximizes on-target reads, and minimizes artifact, and was validated on DNA dilutions derived from cell lines. Plasma NGS was then blindly performed on 48 patients with advanced, progressive NSCLC and a known tumor genotype, and explored in two patients with incomplete tumor genotyping.
RESULTS:NGS could identify mutations present in DNA dilutions at ≥ 0.4% allelic frequency with 100% sensitivity/specificity. Plasma NGS detected a broad range of driver and resistance mutations, including ALK, ROS1, and RET rearrangements, HER2 insertions, and MET amplification, with 100% specificity. Sensitivity was 77% across 62 known driver and resistance mutations from the 48 cases; in 29 cases with common EGFR and KRAS mutations, sensitivity was similar to droplet digital PCR. In two cases with incomplete tumor genotyping, plasma NGS rapidly identified a novel EGFR exon 19 deletion and a missed case of MET amplification.
CONCLUSIONS:Blinded to tumor genotype, this plasma NGS approach detected a broad range of targetable genomic alterations in NSCLC with no false positives including complex mutations like rearrangements and unexpected resistance mutations such as EGFR C797S. Through use of widely available vacutainers and a desktop sequencing platform, this assay has the potential to be implemented broadly for patient care and translational research.
Volume 22(4)
Pages 915-22
Published 2016-2-15
DOI 10.1158/1078-0432.CCR-15-1627-T
PII 1078-0432.CCR-15-1627-T
PMID 26459174
PMC PMC4755822
MeSH Carcinoma, Non-Small-Cell Lung / blood* Carcinoma, Non-Small-Cell Lung / genetics Carcinoma, Non-Small-Cell Lung / secondary Cell Line, Tumor DNA Mutational Analysis DNA, Neoplasm / blood* DNA, Neoplasm / genetics Female High-Throughput Nucleotide Sequencing Humans Lung Neoplasms / blood* Lung Neoplasms / genetics Lung Neoplasms / pathology Male Middle Aged Neoplasm Staging
IF 10.107
Human and Animal Cells LC-2/ad(RCB0440)