In this work, we demonstrate that there is crosstalk between integrin beta1 and c-MET and this crosstalk regulates EGFR TKIs resistance in NSCLC. We provide evidence that integrin beta1/MET crosstalk is a key factor of EGFR TKIs resistance, thus rendering integrin beta1/c-MET a suitable double-target for adjuvant therapy in combination with anti-EGFR agents currently used in clinic.
Although the patients with mutant EGFR display dramatic response to EGFR TKIs, duration of response is typically only 9 to 10 months and then most patients eventually acquire resistance to the agents [20, 21], leading to treatment failure. Mechanisms for acquired resistance to EGFR TKIs have been widely studied. T790M mutation and c-MET gene amplification have been found to be related to acquired resistance to EGFR TKIs in NSCLC [22–24]. Except for the above two mechanisms, others that account for the remaining about 30% of acquired resistance are still unclear. Some papers showed that integrin beta1 signaling has been implicated in the progression and metastasis of various cancers, and shown to facilitate resistance to radiation therapy  and drug resistance . Moreover, our previous research had confirmed that integrin beta1 was responsible for EGFR TKI resistance.
Integrin beta1, that associates with the adhesion and migration capability of tumor cells and has a key role in the growth and metastasis of tumors, is an important molecular of the adhesion-mediated drug resistance [27–29]. The FN receptor (α5β1-integrin) binds to fibronectin to anchor cells and activates non receptor tyrosine kinases, FAK and Src, which play an important role in tumorigenesis by promoting the proliferation and invasion of cancer and endothelial cells [30, 31]. In our previous research, we have established the cell lines with stable down- and up- expression of integrin beta1 by transfecting siRNA or integrin beta1 cDNA plasmid into PC9/AB2 cells or PC9 cells, respectively. After down-regulation of integrin beta1, PC9/AB2 cells partially restored sensitivity to gefitinib while up-regulation of integrin beta1 led to resistance of PC9 cells to gefitinib. Expression level of integrin beta1 was negatively correlated with gefitinib sensitivity in these two cell lines. These data identified that integrin beta1 is an important factor of EGFR TKIs resistance. Morello also found that the integrin beta1-silenced cells showed a defective activation of the EGFR signaling cascade, leading to decreased in vitro proliferation, enhanced sensitivity to gefitinib, impaired migration and invasive behavior .
Our results showed that both integrin beta1 and c-MET were expressed in these cell lines, and their ligands can enhance cell proliferation synergistically. Importantly, inhibition of both receptors led to growth inhibition and apoptosis, and down-regulation of phosphorylation of molecules in their downstream signal transduction (such as the AKT, FAK pathways) in a synergistic fashion. Ligand-dependent activation of integrin beta1 induced c-MET and it’s downstream signals activation (FAK, and AKT). Mitra et al. also reported that inhibition of α(5)β(1)-integrin decreased the phosphorylation of c-Met, FAK and Src, both in vitro and in vivo. Activation of c-Met by its ligand, HGF/SF, or overexpression of a constitutively active FAK in HeyA8 cells could overcome the effect of α(5)β(1)-integrin inhibition on tumor cell invasion, indicating that α(5)β(1)-integrin is upstream of c-Met, Src and FAK. Inhibition of α(5)β(1)-integrin on cancer cells in two xenograft models of ovarian cancer metastasis resulted in a significant decrease of tumor burden, which was independent of the effect of α(5)β(1)-integrin on angiogenesis . These data shows that there is a crosstalk between integrin beta1 and c-MET signaling pathways, and it reaches consensus with Beviglia’s results that the two signaling pathways, integrin/ECM and c-MET/HGF, cooperate synergistically to induce FAK activation in an adhesion-dependent manner, leading to enhanced cell adhesion and motility .
In conclusion, we identified that the crosstalk between integrin beta1 and c-MET via AKT and FAK signaling pathways is very important in EGFR TKI resistance. Our findings identified a new molecular mechanism of EGFR TKIs resistance, which will provide an effective therapeutic intervention of EGFR TKIs resistance.