Heterogeneity of resistance mutations detectable by next-generation sequencing in TKI-treated lung adenocarcinoma

Abstract

EGFR-mutated lung adenocarcinomas routinely develop resistance to tyrosine kinase inhibitors (TKI). To better characterize the relative frequencies of the resistance mechanisms, we analyzed 48 EGFR-mutated TKI-resistant specimens from 41 patients. Next-generation sequencing of post-treatment specimens detected EGFR p.T790M in 31 (79%) of 39 patients, PIK3CA mutations in 10 (26%), EGFR p.S768_V769delinsIL in one, and KRAS p.G12C in one. Five PIK3CA mutations were outside of codons 542, 545, and 1047. Three of four pre-treatment specimens did not carry the PIK3CA mutation found in the post-treatment sample. Small cell carcinoma transformation was identified in four patients; none had p.T790M, including two where p.T790M was identified in the co-existing adenocarcinoma. In p.T790M-mutated specimens, the allele frequency was less than 5% in 24% of cases. p.T790M allele frequency was usually lower than that of the sensitizing mutation indicating that the resistance mutation was present either in a subset of cells or, if the sensitizing mutation was amplified, in a subset of the sensitizing alleles of a dominant clone. Eight patients had multiple resistance mutations, suggesting either multiple separate resistant clones or a single clone harboring multiple resistance mechanisms. PIK3CA mutations appear to be a more significant resistance mechanism than previously recognized.

Molecular Adequacy of Image-Guided Rebiopsies for Molecular Retesting in Advanced Non–Small Cell Lung Cancer: A Single-Center Experience

“…Third, incomplete data on the technical aspects of each procedure precluded analysis of the potential relationship between incidence of molecular analysis failure and the way in which procedures were performed, which would help define optimal conditions to obtain adequate tissue samples. Finally, instead of single-gene tests performed in parallel or sequentially, many centers have now moved to implementing next-generation sequencing-based molecular genotyping [35][36][37][38] ; therefore, the individual molecular success rate at individual genes may not reflect changes in gene testing methodologies.…”

Drugging the 'undruggable' cancer targets

“…A recent study of epidermal growth factor receptor ( EGFR )-mutated lung carcinomas revealed the existence of multiple resistance mutations in some patients, suggesting the presence of multiple but separate resistant clones or a single cell harbouring multiple resistance mechanisms 50 .…”

Clinical Validation of Coexisting Activating Mutations Within EGFR, Mitogen-Activated Protein Kinase, and Phosphatidylinositol 3-Kinase Pathways in Lung Cancers

“…Next-generation sequencing was conducted by using AmpliSeq Cancer Hotspot Panel (v2) (Life Technologies) for targeted multigene amplification as described previously. 20,21 Sequencing data of the targeted genes were analyzed with Torrent Suite (Life Technologies). Mutations were identified and annotated through both Torrent Variant Caller and direct visual inspection of the binary sequence alignment/map (BAM) file on the Broad Institute's Integrative Genomics Viewer (IGV).…”