Abstract
IntroductionWith the approval of first-line osimertinib treatment in stage IV EGFR-mutated NSCLC, detection of resistance mechanisms will become increasingly important—and complex. Clear guidelines for analyses of these resistance mechanisms are currently lacking. Here, we provide our recommendations for optimal molecular diagnostics in the post-EGFR tyrosine kinase inhibitor (TKI) resistance setting. MethodsWe compared molecular workup strategies from three hospitals of 161 first- or second-generation EGFR TKI–treated cases and 159 osimertinib-treated cases. Laboratories used combinations of DNA next-generation sequencing (NGS), RNA NGS, in situ hybridization (ISH), and immunohistochemistry (IHC). ResultsResistance mechanisms were identified in 72 first-generation TKI cases (51%) and 85 osimertinib cases (57%). RNA NGS, when performed, revealed fusions or exon-skipping events in 4% of early TKI cases and 10% of osimertinib cases. Of the 30 MET and HER2 amplifications, 10 were exclusively detected by ISH or IHC, and not detected by DNA NGS, mostly owing to low tumor cell percentage (<30%) and possibly tumor heterogeneity. ConclusionsOur real-world data support a method for molecular diagnostics, consisting of a parallel combination of DNA NGS, RNA NGS, MET ISH, and either HER2 ISH or IHC. Combining RNA and DNA isolation into one step limits dropout rates. In case of financial or tissue limitations, a sequential approach is justifiable, in which RNA NGS is only performed in case no resistance mechanisms are identified. Yet, this is suboptimal as—although rare—multiple acquired resistance mechanisms may occur.
Highlights
With the approval of first-line osimertinib treatment in stage IV EGFR-mutated NSCLC, detection of resistance mechanisms will become increasingly important—and complex
In first- and second-generation tyrosine kinase inhibitor (TKI) resistance, acquired resistance mechanisms predominantly consist of ontarget mutations in EGFR, mainly T790M,6–9 and D761Y,10 L747S,11 and T854A point mutations and EGFR amplification.[6,7,8,9,12]
We considered a molecular alteration in the resistance biopsy an “acquired resistance mechanism” if (1) the alteration was absent in the pretreatment biopsy and (2) the molecular alteration was considered to be a class 4 or 5 pathogenic mutation, reported to be associated with an acquired EGFR TKI resistance phenotype in previous literature, such as EGFR T790M, KRAS G12C, and BRAF V600E
Summary
With the approval of first-line osimertinib treatment in stage IV EGFR-mutated NSCLC, detection of resistance mechanisms will become increasingly important—and complex. We provide our recommendations for optimal molecular diagnostics in the post-EGFR tyrosine kinase inhibitor (TKI) resistance setting. 11% of all lung adenocarcinomas harbor a driver mutation in the EGFR gene.[1] Most of these EGFR mutations have been targeted with first- and second-generation tyrosine kinase inhibitors (TKIs) for several years, resulting in a substantial improvement of both overall and progression-free survival for these patients.[2,3] In 2017, osimertinib, a third-generation TKI, was approved by the Food and Drug Administration and European Medicines Agency as second line[4] and more recently as first line for the treatment of metastatic EGFR-mutated NSCLC, which further improved survival. In first- and second-generation TKI resistance (such as erlotinib, gefitinib, afatinib), acquired resistance mechanisms predominantly consist of ontarget mutations in EGFR, mainly T790M,6–9 and D761Y,10 L747S,11 and T854A point mutations and EGFR amplification.[6,7,8,9,12] Off-target resistance mechanisms include mutations in BRAF, PIK3CA, and KRAS, amplifications of HER2 and MET, oncogenic fusions in RET, FGFR3, ROS1, and NTRK, and MET- and EGFR-exon skipping and transformation into SCLC.[6,7,8,9,13,14,15,16,17,18] Squamous transformation has so far only been described in case reports after first- and second-generation TKIs.[19]
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