P0081 Src inhibitors act through different mechanisms to cooperate with EGFR or MEK inhibitors in NSCLC models sensitive or resistant to erlotinib

Abstract

Background The EGFR tyrosine kinase inhibitors gefitinib and erlotinib are first-line therapy for NSCLCs harbouring EGFR-activating mutations. The intrinsic and acquired resistance to these agents is a relevant clinical issue. Although Src tyrosine kinase has been involved in such resistance in preclinical models, clinical development of these agents has been so far limited. Methods We used a panel of human NSCLC cell lines: PC9 and HCC827 (EGFR-activating mutation; highly sensitive to erlotinib), Calu3 (EGFR/Ras wild-type, wt; moderately sensitive), Calu3-ER (with acquired resistance), H1299 and A549 (Ras mutant; resistant), H1975 (EGFR T790M mutant; resistant). In these models, we tested three different Src inhibitors (saracatinib, dasatinib, and bosutinib), both in vitro and in vivo. Findings NSCLC cell lines showed different activation of EGFR-dependent and Src-dependent pathways and variable sensitivity to Src inhibitors. A kinase assay demonstrated that all the compounds are able to directly inhibit EGFR tyrosine kinase variants. In cell lysates only saracatinib and bosutinib efficiently reduced EGFR activation, while dasatinib was the more effective agent in inhibiting Src tyrosine kinase. Consistently, in EGFR-activating mutant, erlotinib sensitive cells, saracatinib and bosutinib showed anti-proliferative effects related to simultaneous EGFR/Src inhibition. In EGFR wt/Ras mutant cells Src inhibition by dasatinib interfered with cell proliferation and signal transduction. Since Src inhibitors had only moderate effects as single agents we tested the combination of saracatinib with EGFR inhibitors (erlotinib or cetuximab) in EGFR-addicted cells, and of dasatanib with MEK inhibitors (selumetinib) in Ras mutant, erlotinib resistant models. These combinations were effective both in vitro and in nude mice, inhibiting tumour growth, prolonging mice survival, and interfering with signal transduction. Importantly, the combination of saracatanib and cetuximab was effective also in the erlotinib resistant, EGFR T790M mutant model. Interpretation Src inhibitors may act with different mechanisms in NSCLC cell lines, depending on EGFR/Ras mutational profile. Integration of anti-Src agents with EGFR or MEK inhibitors could represent effective therapeutic options for different cohorts of NSCLC patients.