Anti-angiogenic therapies targeting vascular endothelial growth factor (VEGF) and its receptor (VEGF-R) are important treatments for a number of human malignancies， including colorectal cancers. However， there is increasing evidence that VEGF/VEGF-R inhibitors promote the adaptive and evasive resistance of tumor cells to the therapies. The mechanism by which the cancer cells become resistant remains unclear. One potential mechanism is that VEGF/VEGF-R blockers directly act on tumor cells independently of anti-angiogenic effects. In this study， the direct effects of an anti-VEGF antibody (bevacizumab) and a VEGF-R tyrosine kinase inhibitor (sunitinib) on the evasive adaptation of colon cancer cells were co... More
Anti-angiogenic therapies targeting vascular endothelial growth factor (VEGF) and its receptor (VEGF-R) are important treatments for a number of human malignancies， including colorectal cancers. However， there is increasing evidence that VEGF/VEGF-R inhibitors promote the adaptive and evasive resistance of tumor cells to the therapies. The mechanism by which the cancer cells become resistant remains unclear. One potential mechanism is that VEGF/VEGF-R blockers directly act on tumor cells independently of anti-angiogenic effects. In this study， the direct effects of an anti-VEGF antibody (bevacizumab) and a VEGF-R tyrosine kinase inhibitor (sunitinib) on the evasive adaptation of colon cancer cells were compared. HCT116 and RKO human colon cancer cell lines were chronically exposed (3 months) to bevacizumab or sunitinib in vitro to establish bevacizumab- and sunitinib-adapted cells， respectively. Transwell migration and invasion assays， western blotting， reverse transcription-quantitative polymerase chain reaction， co-immunoprecipitation analysis， cell survival assays and ELISAs were conducted to analyze the adapted cells. Compared with the control vehicle-treated cells， the two cell models exhibited increased migration and invasion activities to different degrees and through different mechanisms. The bevacizumab-adapted cells， but not in the sunitinib-adapted cells， exhibited redundantly increased expression levels of VEGF/VEGF-R family members， including VEGF-A， placental growth factor， VEGF-C， VEGF-R1 and VEGF-R3. In addition， the phosphorylation levels of VEGF-R1 and VEGF-R3 were increased in the bevacizumab-adapted cells compared with the control cells. Thus， the inhibition of VEGF-R1 and VEGF-R3 decreased the evasive activities of the cells， suggesting that they remained dependent on redundant VEGF/VEGF-R signaling. By contrast， the sunitinib-adapted cells exhibited increased neuropilin-1 (NRP1) expression levels compared with the control cells. In the sunitinib-adapted cells， NRP1 interacted with phosphorylated cMet， and the cMet activation was dependent on NRP1. Thus， NRP1 or cMet blockade suppressed the evasive activation of the sunitinib-adapted cells. These results suggest that the sunitinib-adapted cells switched from a VEGF-R-dependent pathway to an alternative NRP1/cMet-dependent one. The findings of the present study indicate that VEGF/VEGF-R inhibitors directly act on colon cancer cells and activate their evasive adaptation via different mechanisms.
