A Pan-Canadian Validation Study for the Detection of EGFR T790M Mutation Using Circulating Tumor DNA From Peripheral Blood

Abstract

Introduction Genotyping circulating tumor DNA (ctDNA) is a promising noninvasive clinical tool to identify the EGFR T790M resistance mutation in patients with advanced NSCLC with resistance to EGFR inhibitors. To facilitate standardization and clinical adoption of ctDNA testing across Canada, we developed a 2-phase multicenter study to standardize T790M mutation detection using plasma ctDNA testing. Methods In phase 1, commercial reference standards were distributed to participating clinical laboratories, to use their existing platforms for mutation detection. Baseline performance characteristics were established using known and blinded engineered plasma samples spiked with predetermined concentrations of T790M, L858R, and exon 19 deletion variants. In phase II, peripheral blood collected from local patients with known EGFR activating mutations and progressing on treatment were assayed for the presence of EGFR variants and concordance with a clinically validated test at the reference laboratory. Results All laboratories in phase 1 detected the variants at 0.5 % and 5.0 % allele frequencies, with no false positives. In phase 2, the concordance with the reference laboratory for detection of both the primary and resistance mutation was high, with next-generation sequencing and droplet digital polymerase chain reaction exhibiting the best overall concordance. Data also suggested that the ability to detect mutations at clinically relevant limits of detection is generally not platform-specific, but rather impacted by laboratory-specific practices. Conclusions Discrepancies among sending laboratories using the same assay suggest that laboratory-specific practices may impact performance. In addition, a negative or inconclusive ctDNA test should be followed by tumor testing when possible.

Can Liquid Biopsy Based on ctDNA/cfDNA Replace Tissue Biopsy for the Precision Treatment of EGFR-Mutated NSCLC?

“…Of course, the impact of false negatives from the different detection methods also needs to be considered [ 43 ]. The limited yield and complex kinetics of ctDNA carry the risk of false negative results, even when using sensitive and well-validated molecular detection methods [ 44 ]. When possible, tumor tissues should be tested when ctDNA test results are negative or inconclusive.…”

Liquid Biopsy in Clinical Oncology

“…Так, например, в клиническую практику повсеместно вошло генотипирование цоДНК из периферической крови больных с немелкоклеточным раком легкого, прогрессирующих на фоне анти-EGFR терапии. В ряде масштабных клинических испытаний было показано, что у значительной доли пациентов (24-37%) формирование резистентности к таргетному препарату сопровождается появлением в плазме фрагментов опухолевой ДНК, несущих приобретенную мутацию EGFR T790M [63][64]. Для поиска активирующих мутаций в гене EGFR (exon 19del, L858R) и отслеживания момента появления мутации резистентности EGFR T790M в цоДНК у пациентов с раком легкого был адаптирован метод цифровой капельной ПЦР (droplet-digital PCR, ddPCR).…”