Pathological Significance And Drug Target Effect Of Aberrant RON And MET Co-overexpression In Pancreatic Cancer

Nowadays,cancer has become one of the leading causes of death in China,and poses a serious threat to human health.As a leading cancer,pancreatic cancer has the characteristics of poor clinical manifestation,poor prognosis and high mortality.Because of pancreatic cancer with high proliferation and chemical resistance,most patients have lost the opportunity for surgery when diagnosed.Current treatment options such as radiotherapy and chemotherapy are almost useless for patients to improve their quality of life and extend the survival time,so there is an urgent need to explore new biomarkers and effective therapeutic targets.Tyrosine kinase receptor RON(also known as macrophage-stimulating 1 receptor[MST1R])and MET(also known as hepatocyte growth factor receptor[HGFR])both belong to MET proto-oncogene family.Various cancers have aberrant RON and MET expression and activation.In this study,RON and MET expression level in human pancreatic cancer tissue samples was analyzed by immunohistochemical staining.Four human pancreatic cancer cell lines expressing variable levels of RON and/or MET and four MET superfamily inhibitors were used.The effect of the four tyrosine kinase inhibitors on pancreatic cell viability,migration,and apoptosis were determined using Cell Counting Kit-8 assays,wound healing assays,and Caspase-Glo 3/7 assays.Cellular signaling pathway and signaling proteins were analyzed by immunoprecipitation and western blotting.The therapeutic efficacy of four tyrosine kinase inhibitors was determined with mouse pancreatic cancer xenograft models in vivo.In the present study,we found that there were widely aberrant RON and MET co-expression in the pancreatic cancer tissues,and their expression levels were significantly related to OS.Patients with RON and MET co-overexpression had poorest OS.TKIs targeting RON and MET signaling pathways can effectively inhibit pancreatic cancer cell viability and migration,promote apoptosis,and further inhibit tumor growth in vivoPart 1 RON and MET expression in pancreatic cancer andtheir relationship with clinicopathological characteristics and OSAimRON(Recepteur d'origine nantais)and MET have abnormal expression and activation in various types of cancer,but the co-expression characteristics of RON and MET in pancreatic cancer and their relationship with patient survival time are still worth further exploring.In this study,we explored the expression of RON and MET in pancreatic cancer and their relationship with clinicopathological characteristics and overall survival time.Material and methodWe enrolled 227 patients who had been pathologically diagnosed with pancreatic cancer with or without liver metastases in the study.The clinical parameters included patient demographics,tumor size,tumor node-metastasis(TNM)stage,tumor di erentiation,lymph node metastasis,distant metastasis and survival time.RON and MET expression level were analyzed by immunohistochemical staining.The relationship between RON and MET expression and clinicopathological characteristics was compared using the chi-square test.Survival data were analyzed by the Kaplan-Meier method and log rank test.The independent prognostic factors of survival were identified using Cox proportional hazard model analysis.ResultsThere was widely aberrant RON and MET expression in the cancer tissues,and elevated RON expression was associated with distant metastasis(P=0.019).Of the 227 pancreatic cancer samples,195(85.9%)samples had positive RON expression,and 75(33%)samples had RON overexpression;207(91.2%)samples had positive MET expression,and 93(41%)samples had MET overexpression.182(80%)samples had RON and MET co-expression,of which RON and MET were co-overexpressed in 35 samples,approximately 50%of the RON overexpressed samples.RON and MET expression levels were highly correlated,and significantly related to OS.Patients with RON and MET co-overexpression had poorer OS.ConclusionRON is an independent predictor of distant metastases.RON and MET expression levels are significantly correlated with the survival time of patients.RON and MET can be important prognosis biomarkers in pancreatic cancer.Part 2 Inhibitory effects of tyrosine receptor inhibitors targeting RON and/or MET in pancreatic cancer in vitro and in vivoAimVarious cancers have aberrant RON and MET expression and activation,which promote cancer cell proliferation,invasiveness and metastasis.In this study,we explored whether there is a cross-talk between RON and MET in pancreatic cancer and their signal transduction mechanism,and further analyzed the significance of overexpressed RON and/or MET as therapeutic targets for anti-pancreatic cancer drugs.Material and methodFour human pancreatic cancer cell lines expressing variable levels of RON and MET were selected as the TKIs targeting model.Four MET superfamily inhibitors(BMS-777607,PHA665752,INCB28060,Tivantinib)were used.The effect of the four tyrosine kinase inhibitors on cell viability,migration,and apoptosis were determined using Cell Counting Kit-8 assays,wound healing assays,and Caspase-Glo 3/7 assays.Cellular signaling pathway and signaling proteins were analyzed by immunoprecipitation and western blotting.The therapeutic efficacy of the tyrosine kinase inhibitors in vivo was determined with mouse pancreatic cancer xenograft models.ResultsBMS-777607 and PHA665752 inhibited pancreatic cancer cell viability and migration,and promoted apoptosis by inhibiting RON and MET phosphorylation and further inhibiting their mediated signaling pathways in vitro.They also inhibited tumor growth and further inhibited phospho-RON and phospho-MET expression in the mouse xenograft models in vivo effectively.INCB28060,which inhibits the MET signaling pathway alone,was not effective.ConclusionRON and MET co-expression and their cross-talk promote the malignant biological behavior of pancreatic cancer cells.The dual inhibition of abnormally expressed RON and MET may be a potentially effective strategy for the treatment of pancreatic cancer.