Combined inhibition of MET and VEGF enhances therapeutic efficacy of EGFR TKIs in EGFR-mutant non-small cell lung cancer with concomitant aberrant MET activation

Background: Aberrant activation of mesenchymal epithelial transition (MET) has been considered to mediate primary and acquired resistance to epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) in EGFR-mutant non-small cell lung Cancer (NSCLC). However, mechanisms underlying this process are not wholly clear and the effective therapeutic strategy remains to be determined.

Methods: The gefitinib-resistant NSCLC cell lines were induced by concentration increase method in vitro. Western blot and qPCR were used to investigate the relationship between MET and vascular endothelial growth factor (VEGF)/VEGF receptor 2 (VEGFR2/KDR/Flk-1) signaling pathway. Double luciferase reporter gene and co-immunoprecipitation were used to further reveal the regulation mechanism between MET and VEGF/VEGFR2/KDR/Flk-1. The effect of combined inhibition of MET and VEGF/VEGFR2/KDR/Flk-1 signaling pathway on the therapeutic sensitivity of EGFR-TKI in gefitinib resistant cell lines with MET aberration was verified ex vivo and in vivo.

Results: We successfully obtained two gefitinib-resistant NSCLC cell lines with EGFR mutation and abnormal activation of MET. We observed that MET formed a positive feedback loop with the VEGF/VEGFR2/KDR/Flk-1 signaling, leading to persistent downstream signaling activation. Specifically, MET up-regulated VEGFR2/KDR/Flk-1 expression in a MAPK/ERK/ETS1-dependent manner, while VEGF promoted physical interaction between VEGFR2/KDR/Flk-1 and MET, thereby facilitating MET phosphorylation. A Met Inhibitor, crizotinib, combined with an anti-VEGF antibody, bevacizumab, enhanced the sensitivity of NSCLC cells to gefitinib and synergistically inhibited the activation of downstream signaling in vitro. Dual inhibition of MET and VEGF combined with EGFR TKIs markedly restrained tumor growth in both human NSCLC xenograft models and in an EGFR/MET co-altered case.

Conclusions: Our work reveals a positive feedback loop between MET and VEGF/VEGFR2/KDR/Flk-1, resulting in continuous downstream signal activation. Combined inhibition of MET and VEGF/VEGFR2/KDR/Flk-1 signaling pathway may be beneficial for reversing EGFR TKIs resistance.

EGFR TKIs resistance; MET; Non-small cell lung cancer; VEGF/VEGFR2 signaling.