Concurrent Pemetrexed With EGFR-TKI Slows the Accumulation of De Novo Mutations During In Vitro Exposure

Background/Aim: Resistance to targeted therapy limits its efficacy in non-small cell lung cancer (NSCLC) harboring epidermal growth factor receptor (EGFR) mutations. Although concurrent administration of pemetrexed (PEM) and EGFR-tyrosine kinase inhibitors (TKIs) has yielded clinical benefits, it remains unclear whether concurrent PEM influences de novo mutations in the EGFR-TKI therapy. To address this gap, we aimed to compare the time to acquired resistance and accumulation of de novo tumor mutational burden (TMB) during in vitro exposure to osimertinib (OSI) and gefitinib (GEF), as well as their respective combinations with PEM. Materials and Methods: EGFR-mutated PC-9 cells were continuously exposed to EGFR-TKIs, alone or in combination with PEM, at equimolar concentrations. The drug concentration gradually increased to 1 and 3 μM for OSI and GEF, respectively. Whole-exome sequencing and quantitative PCR were performed to measure TMB and gene expression, respectively. Results: Concurrent PEM with either OSI or GEF extended the treatment duration compared to single EGFR-TKIs, decreased the de novo accumulated TMB per treatment time, and increased the expression of POLE2, POLQ, MLH1, BRCA1, BRCA2, RAD51, and FEN1 compared to that of single EGFR-TKIs. Conclusion: Concurrent PEM with EGFR-TKI treatment slowed TMB accumulation rate and resistance acquisition in EGFR-mutated NSCLC compared to single EGFR-TKIs in vitro.