The Effect And Mechanism Of SDPR During Breast Carcinogenesis And Progression

Background:Breast cancer is one of the most common malignancies in women worldwide and the second leading cause of cancer-related diseases in women,especially middle and old aged women,the incidence of breast cancer is still very high.Although improvements in medical technology have decreased breast cancer mortality rates in recent years,prevention and therapy of breast cancer remain an important research topic for clinicians.Unfortunately,the incomplete understanding of its carcinogenic mechanisms leads to the less targeted treatment and contributes to a low survival rate for patients with breast cancer.Serum deprivation response factor(SDPR),also known as Caveolae Associated Protein 2(Cavin-2),is encoded in chromosome 2nd,q32-q33 location,with the function of coding purified phospholipids binding protein of platelets(PS-P68)and was first purified from human platelets as a phosphatidylserine(PS)binding protein,while its expression in serum-starved cells increased.SDPR has been considered as a key substrate for protein kinase C(PKC)phosphorylation and the intermediate structure of SDPR can be combined with the regulatory domain of PKC.In cells lacking SDPR-PKC interaction,it was found that the cell positioning and the substrate specificity changed significantly and the interaction between the two determines the relationship between PKC and caveolae.SDPR was further confirmed to function in inducing membrane curvature and participate in the caveolae formation.The caveolae is a specialized cystic structure formed by the inward depression of the plasma membrane,while main function is to participate in the transport of the transmembrane substance,and as the hub of cell signaling molecules enrichment and signal transduction,impacting the cell proliferation,migration and invasion ability.Caveolae is calcium channel related to gut electrophysiological pacing function.Previous studies showed that the expression of SDPR gene is downregulated in multiple tumors,suggesting that SDPR is a potential tumor suppressor gene.However,the specific roles and mechanism of SDPR in breast cancer development and progression is still not clear.Objectives and methods:1.The expression of SDPR m RNA in 30 cases of paired breast cancer and normal tissues was detected by RT-q PCR.The normalized SDPR m RNA expression were analyzed based on the expression profile data in Cancer Genome Atlas tumor genome(TCGA),and the Kaplan-Meier survival analysis was used to analyze the disease-free survival of breast cancer patients with different expression level of SDPR by using GOBO database,to understand the differential expression of SDPR in breast cancer and normal tissue.2.Western blot was used to detect the expression level of SDPR in different breast cancer cells,and further to construct SDPR-overexpressing plasmids or si RNAs.After transfection of breast cancer cells,the effects of SDPR expression on the biological behavior of breast cancer cells were evaluated by MTT,clone formation,EDU,Transwell assay,flow cytometry et al.3.The effect of SDPR on EMT in breast cancer was evaluated by RT-q PCR,Western blot,flow cytometry and cell morphology observations,while the effect of SB431542,an inhibitor of TGF-? on EMT-like Phenotypes in breast cancer cell was evaluated by RT-q PCR,MTT,clone formation,EDU,Transwell et al.4.The effects of SDPR on TGF-? signaling pathway were evaluated by luciferase assay,ELISA,immunofluorescence staining,MTT,clone formation,EDU,Transwell assay,RT-q PCR,Western blot,and flow cytometry.Result:1.The SDPR m RNA expression was downregulated in 29/30 collected breast cancer tissues as compared with paired adjacent normal breast tissue.The TCGA database confirmed that SDPR m RNA expression was downregulated in breast cancer tissues.Moreover,the GOBO database data show that low expression of SDPR is associated with poorer prognosis in breast cancer patients.2.The MCF-10 A cell line with the highest SDPR expression level and the MDA-MB-231 cell line with the lowest SDPR expression level were selected as the follow-up study object.It is proved that SDPR can inhibit the proliferation and invasion of breast cancer cells and promote apoptosis by transfection of SDPR overexpression plasmids in MDA-MB-231 and transfection of si RNAs in MCF10 A to inhibit SDPR expression.3.SDPR upregulates the expression of epithelial markers E-cadherin and ?-catenin,while the expression of mesenchymal markers vimentin and N-cadherin were descreased.SDPR inhibits EMT in breast cancer cells,and affects the cell cycle and cell morphology.In addition,TGF-?inhibitor SB431542 can inhibit the EMT-like phenotype of breast cancer cells.4.The EMT-like phenotype in breast cancer cells was retored in SDPR-overexpressed cells after treatment with TGF-?1,in addition,interference with SDPR expression could increase the expression of TGF-?1,activate TGF-? signaling pathway,induce EMT in breast cancer,while it was inhibited after treatment with inhibitor SB431542.Conclusion:This study confirms that SDPR expression was downregulated in breast cancer tissues,and low expression of SDPR is associated with a poor prognosis in breast cancer patients.SDPR inhibits the EMT phenotype in breast cancer cells by blocking TGF-?signaling pathway,thereby inhibiting the proliferation and invasion of breast cancer cells and induceing apoptosis.