To Explore The Mechanism Of Liushen Pill Inhibiting Colorectal Cancer Metastasis By SRC-3 Pathway

ObjectiveTo clarify the anti-tumor metastasis effect of Liushen pill,explore the role of SRC-3 pathway in colorectal cancer metastasis,and then explore the mechanism of Liushen pill in regulating SRC-3 pathway to inhibit colorectal cancer metastasis.Methods:Test cell activity by CCK-8 assay.Test cell proliferation by the clone forming assay.Using Transwell assay to test cell invasion and migration.Detect the interference factors influence on the growth of colorectal cancer by subcutaneous tumor animal models.Detect the interference factors influence on the metastasis of colorectal cancer by orthotopic transplantation tumor animal models of liver metastasis.Test the formation of metastases by in vivo imaging of luciferase.The expression of mir-497 was detected by RT-PCR.HE staining was used to detect the formation of tumor focis in liver tissue.Verified the miR-497 target gene by the dual luciferase reporter gene assay.Lentivirus infection method was used to construct cell lines with stable expression of target genes,and the plasmid transient transfection method was used to increase the expression level of target genes in cells.Results:(1)Liushen pill can inhibit the proliferation and migration of inflammatory tumor cells and down-regulate the expression of SRC-3 protein in cells.(2)Liushen pill can inhibit cell proliferation and metastasis of inflammatory tumor tissue.(3)SRC-3 promotes invasion and migration of colorectal cancer cells,and knockdown of SRC-3 inhibits liver metastasis of colorectal cancer.(4)SRC-3 down-regulates the expression of miR-497,which inhibits liver metastasis of colorectal cancer,and Cyclin D2 is a direct functional target of miR-497.(5)SRC-3/miR-497/Cyclin D2 signal mediated EMT promotes the metastasis of colorectal cancer.(6)Liushen pill inhibited the activity of colorectal cancer cell lines and regulated the expression of EMT-related proteins mediated by the SRC-3 pathway.(7)Liushen pill inhibits the growth and liver metastasis of colorectal cancer with high expression of SRC-3.(8)Liushen pill has certain inhibitory effect on PDX animal model of colorectal cancer,but the difference is not statistically significant,indicating that Liushen pill alone has no definite therapeutic effect on PDX animal model with rich microenvironment.Conclusion:Liushen pill inhibited the inflammatory proliferation and metastasis of colorectal cancer and down-regulated the expression of SRC-3.SRC-3/miR-497/Cyclin D2 mediated EMT may be one of the mechanisms of liver metastasis in colorectal cancer.Liushen pill inhibits the growth and liver metastasis of colorectal cancer with high expression of SRC-3,which may be related to the EMT mediated SRC-3 pathway.Liushen pill alone has no definite therapeutic effect on PDX animal model with rich microenvironment.