Src kinase pathway is a potential candidate for molecular targeted therapy of non-small cell lung cancers (NSCLC) that show exon 19 deletion mutation or other epidermal growth factor receptor (EGFR) abnormalities

Abstract

Molecular targeted therapy using tyrosine kinase inhibitors (TKI) against EGFR is an effective therapy for NSCLC that harbor EGFR mutations. However, tumour resistance due to second EGFR mutations or unknown mechanisms is frequently observed. Src kinase is known to interact with and mutually activate EGFR. The effect of src kinase inhibition on NSCLC has not been fully studied. 4-(4’-Phenoxyanilino)-6,7-dimethoxyquinazolinee (SKI-1) is a novel, potent and highly specific Src kinase inhibitor due to its simultaneous interaction with both the ATP- and peptide-binding sites. To assess the potential usefulness of Src kinase inhibition in treating NSCLC, we studied the effect of SKI-1 on apoptosis and changes in Src kinase and EGFR activation profiles in different NSCLC cells that harbor EGFR wild type (WT) (H358, H1819) or exon 19 deletion mutation (MT) (HCC827). Hoechst 33258 nuclear staining showed that SKI-1 significantly induced apoptosis in both H1819 and HCC827 but not H358 cells in a dose dependent manner. H358 and H1819 cells showed lower basal Src activities than HCC827 cells by immunoblot analysis against phosphorylated Y416. EGFR phosphorylation profiles were also different in the SKI-1 sensitive H1819 and HCC827 cells. While both showed inhibition of phosphorylation at Y416 of Src and Y845 of EGFR in a dose dependent manner, inhibition of EGFR phosphorylation at tyrosine 1068 and 992 was only observed in HCC827 but not H1819 cells. The lack of apoptosis in H358 and sensitivity of HCC827 cells to SKI treatment are consistent with previously reported observations that NSCLC cells harboring WT or MT-EGFR, respectively, show differential sensitivities to SKI due to enhancement of basal src kinase activation by MT EGFR. The inhibition of phosphorylation activation of EGFR at Y845 implicates that induction of apoptosis and sensitivity of HCC827 to SKI treatment is in part related to the involvement of Stat 3/5 pathway downstream of Y845 and, additional, such as the Akt, pathways downstream of EGFR Y1068 and Y992. In view of the WT EGFR and lower basal src kinase activity in H1819, its apoptotic response and sensitivity to SKI-induced inhibition of phosphorylation activation at Src Y416 and EGFR Y845 is unusual. One possible explanation could be related to the high EGFR copy number and expression levels of other ErbB family members in H1819. Overall, the findings indicate that the Src kinase pathway could be a useful target for the treatment of NSCLC. We further suggest that EGFR mutation and basal Src kinase activity are not the only factors that determine SKI sensitivity, but that other forms of abnormalities in EGFR and related family members may also be important determinants.