Let-7c regulated epithelial-mesenchymal transition leads to osimertinib resistance in NSCLC cells with EGFR T790M mutations

Epidermal growth factor receptor- tyrosine kinase inhibitors (EGFR-TKIs) have shown promise against non-small cell lung cancers (NSCLCs) in clinics but the utility is often short-lived because of T790M mutations in EGFR that help evade TKIs’ action. Osimertinib is the third and latest generation TKI that targets EGFRs with T790M mutations. However, there are already reports on acquired resistance against Osimertinib. Recent work has revealed the role that miRNAs, particularly tumor suppressor let-7c, play in the invasiveness and acquired resistance of NSCLCs, but the mechanistic details, particularly in Osimertinib resistance, remain elusive. Using two cells lines, H1975 (endogenous T790M mutation) and HCC827-T790M (with acquired T790M mutation), we found that let-7c is a regulator of EMT, as well as it affects CSC phenotype. In both the cell lines, transfection with pre-let-7c led to reversal of EMT as studied through EMT markers e-cadherin and ZEB1. This resulted in reduced proliferation and invasion. Conversely, reduced expression of let-7c through anti-let-7c transfections significantly increased proliferation and invasion of lung cancer cells. Expression of let-7c was functionally relevant as EMT correlated with resistance to Osimertinib. High let-7c expression reversed EMT and made cells sensitive to Osimertinib, and vice versa. WNT1 and TCF-4 were found to be two targets of let-7c which were epigenetic suppressed by let-7c through increased methylation. In vivo, pre-let-7c inhibited while anti-let-7c potentiated tumor growth and WNT1 and TCF-4 were downregulated in xenografts with pre-let-7c. Silencing of both WNT1 and TCF-4 resulted in potentiation of Osimertinib action. Our results suggest an important role of let-7c in regulating EMT and the resulting Osimertinib resistance in T790M NSCLCs. More clinical studies need to be performed to fully understand the translational relevance of this novel mechanism.