Evolution of heterogeneous mechanisms of acquired resistance to cetuximab-based therapy in colorectal cancer.

3526 Background: Anti-EGFR drug resistance is a major challenge in metastatic colorectal cancer (mCRC). The aim of this study was to elucidate molecular mechanisms of acquired resistance in mCRC patients treated with cetuximab-based therapy. Methods: 37 mCRC patients with acquired resistance to cetuximab-based therapy with paired tumor samples (prior and at progression to treatment) were prospectively included. In three patients, multiple biopsies were obtained over the course of the disease. Mutations in EGFR, KRAS, NRAS, BRAF and PIK3CA genes were evaluated by next-generation pyrosequencing. Gene amplification was evaluated by FISH. Mutations were also assessed in plasma in 7 representative cases. Results: Eighty-one percent of patients harbored a molecular event (comprising mutations and amplifications) in post-treatment biopsies. In one third of patients, multiple molecular events (range 2-5) coexisted in the same sample. RAS mutations were the most frequent event (40% of patients) and mostly affected exons 3 and 4, followed by mutations in PIK3CA (19%), BRAF (11%), EGFR S492R (8%) and novel mutations in EGFR ectodomain (5%). Of note, in 9 patients the mutations were already detected at diagnosis by next-generation sequencing. Multiple repeat biopsies revealed that the percentage of mutant alleles increased under drug pressure and became undetectable following drug withdrawal. Mutations were detected in plasma samples in 62% of tested cases. Conclusions: We provide comprehensive evidence for the evolution of multiple mechanisms of acquired resistance to cetuximab-based therapy in mCRC. This data establishes the need of routine molecular studies at disease progression to individualize further therapeutic decisions.