A Research On The Role Of SDF-1/CXCR4/CXCR7 Axis In Hormone Refractory Prostate Cancer

Prostate cancer is a common malignant tumor in men.With the improvement of living standard,the incidence of prostate cancer is increasing.Prostate cancer is mainly developed by androgen receptors,so androgen-free endocrine therapy is standard therapy.Endocrine therapy can make the patient's tumor shrink and PSA decline,it can alleviate the symptoms to some extent.But after 18-30 months of median remission,Almost all patients develop drug resistance,and then develop into hormone refractory prostate cancer.Endocrine therapy has no effect on these patients and the survival rate is low,their survival time is generally less than 20 months.Therefore,the transformation of hormone refractory prostate cancer has become a difficult point in the treatment of prostate cancer.The stromal cell-derived factor 1(SDF-1)is a chemotactic cytokine that regulates many biological processes,including heart and nerve development,angiogenesis,stem cell movement and tumorigenesis.The receptors for SDF-1 are CXCR4 and CXCR7.The SDF-1/CXCR4/CXCR7 biological axis has extensive biological effects.The main manifestations are mobilization and homing of hematopoietic stem cells,metastasis and growth of tumors,etc.The discussion of SDF-1/CXCR4/CXCR7 biological axis and tumor is also a hot issue.In the earlier research work of our group,steroid-resistant prostate cancer cells was induced by androgen removal in vitro,and transcriptome was detected by microarray.The results showed that CXCR7 and SDF-1/CXCR related regulatory genes were upregulated significantly.It is speculated that SDF-1/CXCR axis may play an important role in hormone resistance of prostate cancer.Therefore,a hormone refractory cell model was used,which derived from LNCap,and has the same genetic background and transfer characteristics.Based on transcriptome and phosphorylation proteomics analysis,the role of SDF-1/CXCR4/CXCR7 biological axis in the pathogenesis of HRPC was discussed,it provides a new direction for the study of the transformation mechanism of HRPC.In this study,RNA-seq method was used to compare the difference of gene expression profile between LNCaP cells and C4-2 cells.Then 293 differentially expressed genes were obtained,of which 118 were up-regulated and 175 were down-regulated.According to cluster analysis,up-regulated genes is mainly involved in cell proliferation,apoptosis,gene regulation,expression,metabolism and other functions,down-regulation genes is mainly involved in membrane protein,transmembrane,intercellular signal transduction,gene expression regulation and other functions.The expression of CXCR7 gene was up-regulated,and the downstream signal transduction pathway regulated by SDF-1 was significantly different in gene expression.Therefore,it was speculated that CXCR7 might play a role in the formation of HRPC.Western blot was used to detect differentially expressed gene related proteins.The results showed that the CXCR4 and CXCR7 in C4-2 was significantly higher than that in LNCaP cells.It suggested that the SDF-1/CXCR4/CXCR7 axis might play an important role in the development of steroid-resistant prostate cancer.In order to understand the specific mechanism of hormone resistance in prostate cancer,LNCaP and C4-2 cells were analyzed by quantitative proteomics with phosphorylated iTRAQ.A total of 63 up-regulated phosphorylated proteins and 101 down-regulated proteins were obtained.By cluster analysis,the up-regulation protein is mainly involved in the function of cell adhesion,migration,invasion,cytoskeleton and so on,which is also consistent with SDF-1/CXCR4/CXCR7 mediated effects.Among the up-regulated phosphorylated proteins,28 proteins have clear upstream regulatory kinases,including PKA,MEK,ERK,catenin.Based on the analysis of known signaling pathways and protein interactions,it is further confirmed that SDF-1/CXCR4/CXCR7 may be involved in MAPK multicascade reaction,and that SDF-1/CXCR-PKA-ERK pathway may be involved in hormone resistance and malignant biological behavior of prostate cancer.In order to verify the role of SDF-1/CXCR4/CXCR7 axis,we used CXCR receptor activator SDF-1 and CXCR4,CXCR7 inhibitor AMD3100 and neutralizing antibody ab72100 to treat C4-2 cells,and tested the effects on the biological behavior of hormone refractory prostate cancer.The results showed that SDF-1 could significantly promote the proliferation and invasion of C4-2 cells,but blocking CXCR4\CXCR7 could inhibit the proliferation and invasion of C4-2,and blocking CXCR4 and CXCR7 showed the most significant effect.These results suggest that CXCR4 and CXCR7 play a synergistic role in regulating the proliferation,invasion and metastasis of steroid-resistant prostate cancer C4-2 cells.Western blot was used to verify the pathway of SDF-1/CXCR4/CXCR7 axis,the results showed that the expression of apoptosis-related proteins such as P53,Caspase3,mTOR was not affected by SDF-1/CXCR4/CXCR7.The expression of MAPK cascade proteins such as MEK?pMEK?ERK1/2?pERK1/2in SDF-1 was significantly lower than that in C4-2 after treatment with CXCR4 and CXCR7 inhibiters,especially in blocking the CXCR4 and CXCR7 simultaneously.The expression of MAPK cascade regulatory protein PKA?pPKA? Catenin was also affected by SDF-1/CXCR4/CXCR7.Compared with C4-2,the expression of pPKA? Catenin increased significantly after treatment with CXCR4 and CXCR7 inhibiters.These results suggest that the SDF-1/CXCR4/CXCR7 axis affects the formation of MAPK networks and playsan important role in hormone refractory prostate cancer through the regulation of the SDF-1/CXCR4/CXCR7-PKA-MEK-ERK pathway.It has been reported that SDF-1 / CXCR4 / CXCR7 axis plays an important role in liver cancer,breast cancer,cervical cancer and other tumors.However,there are no reports of hormone refractory prostate cancer based on C4-2.This study may provide a new direction for the clinical treatment of HRPC.