The Effect C-Met Inhibitor PHA-655752Combined With5-FU, CDDP On Colorectal Xenograft Tumor

Background: Colorectal cancer is one of the most commonlygastrointestinal malignant tumor and its incidence and mortality showed arapid upward trend in our country. Most patients have been diagnosed asmiddle-late stage. Traditional treatment methods include surgery,chemotherapy and radiotherapy. In recent years, targeted drugs have emergedand been used in clinical VEGF antagonists(Bevacizumab), EGFRantagonists(Cetuximab and Panitumimab). But either chemotherapy (5-FU,CDDP, etc.) or targeted drugs are likely to produce drug resistance, and hascertain side effects. Thus, finding new colorectal cancer treatment strategy isbecoming a hot topic currently.C-Met, a proto-oncogene encoding by c-met, is an important class oftyrosine kinase receptor. It highly expressed in colorectal cancer cells, andclosely related to tumor proliferation, invasion, angiogenesis and lymph nodemetastases. Previous studies have found that high expression of c-Met tumorswere resistant to radiotherapy, chemotherapy and EGFR TKIs targeted drugs.Currently, c-Met inhibitor shows good efficacy in clinical trials in solid tumorssuch as non-small cell lung cancer. c-Met inhibitor is also likely to be used inthe treatment of colorectal cancer. However, c-Met inhibitor combined withtraditional chemotherapy on the growth of colorectal cancer has not beenreported.In vitro experiments of the previous studies, we have established a stableand inducible c-Met expression in human colon cancer cell line SW620.Appropriate knocked-down c-Met expression can inhibit the proliferation ofSW620and can be capable of sensitizing5-FU inhibition of proliferation ofSW620. This study is aims to detect the effect of SW620tumor growth byc-Met inhibitor combined with5-FU, CDDP and the possible mechanisms. Objective: To explore the effect of the c-Met inhibitor combined with5-FU, CDDP on SW620xenograft tumor growth and the possiblemechanisms.Methods:1. Cultured human colon cancer cells SW620, which was inoculatedsubcutaneously and established (BALB/c-nu,5weeks old, male) SW620nudemice xenograft tumor model.2. There were six groups in the experiment and every group of eightmices: control group (DMSO group),2.5%DMSO diluent medication everyother day administer; simple fluorouracil group (5-FU group),40mg/kg everythird day administer; simple cisplatin group (CDDP group),3mg/kg everythird day administer; simple c-Met inhibitor (PHA-665752group),25mg/kgevery other day administer; fluorouracil+c-Met inhibitor group (5-FU+PHA-665752group), the same with single medication, respectively; cisplatin+c-Met inhibitor group (CDDP+PHA-665752group), the same with singlemedication, respectively. All groups were administered continuous threeweeks by intraperitoneal injection. Observe the physiological state of the nudemice and tumor volume changes during the experiment.3. The tumor specimens paraffin-embedded and sectioned every group,morphological changes of SW620was observed by HE staining; theexpression of E-cadherin, Vimentin protein and Ki-67protein was detected byimmunohistochemistry of every group xenograft tumor; the tumor apoptosiswas detected by situ terminal transferase enzyme labeling (TUNEL).Results:1. Physiological state of nude mice: expect CDDP group, nude mice werein good condition before and after treatment each group and no significantchange. CDDP can produce toxicity for nude mice. However, CDDPcombined with PHA can reduce the toxicity of CDDP.5-FU did not producesignificant toxicity and5-FU combined with PHA did not increase the toxicityfor nude mice.2. Tumor growth in each group: the experimental groups compared with DMSO group, the average volume of SW620tumor is smaller than DMSOgroup, the differences were statistically significant (P <0.01). The averagevolume of5-FU+PHA group was smaller than5-FU group (P <0.01); CDDP+PHA group was smaller than CDDP group(P <0.01).3. HE staining:5-FU+PHA group and CDDP+PHA group tumor hadmore serious necrosis than other groups, and a large number of inflammatorycells were observed in these two groups.4. Immunohistochemical results:the experimental groups E-cadherinprotein expression was significantly higher than DMSO group (P <0.01);5-FU+PHA group was significantly higher than5-FU group (P <0.01) andCDDP+PHA group was significantly higher than CDDP group (P <0.01).The experimental groups Vimentin protein expression was lower than DMSOgroup; the CDDP group lower than the DMSO group (P <0.05), the otherexperimental groups was lower than DMSO group (P <0.01)5-FU+PHAgroup was lower than5-FU group (P <0.01) and CDDP+PHA group waslower than CDDP group (P <0.01). The experimental groups Ki-67proteinexpression was lower than DMSO group (P <0.01);5-FU+PHA group waslower than5-FU group (P <0.01); CDDP+PHA group was lower than CDDPgroup (P <0.01).5. TUNEL assay results: the average of apoptotic cells rate in each groupwas (7.53±2.16)%、(18.39±1.90)%、(19.70±1.97)%、(24.38±3.79)%、(36.37±3.40)%、(30.69±1.24)%. The experimental groups significantlyhigher than DMSO group (P <0.01);5-FU+PHA group was significantlyhigher than5-FU group and PHA group (P <0.01), CDDP+PHA group wassignificantly higher than CDDP group and PHA group (P <0.01).Conclusion:1. C-Met inhibitor can increase5-FU, CDDP sensitivity for colon cancercells; the mechanism may be inhibited epithelial-mesenchymal transition(EMT). 2. C-Met inhibitor combined with5-FU, CDDP does not increase toxicityand side effects.