The Effect And Mechanism Of MiR-575 On The Sensitivity Of Tamoxifen In ER-positive Breast Cancer

Objective: Breast cancer is the most common malignancy and approximately 70% of breast cancers are estrogen receptor-α(ERα)positive.Antiestrogen tamoxifen is a highly effective and commonly used treatment for patients with ER+ breast cancer.However,intrinsic or acquired resistance to tamoxifen presents a major challenge for ER+ breast cancer therapy,in addition,the underlying mechanism of endocrine resistance remains unclear.In recent years,the role of micro RNA(mi RNA)has increased dramatically in the development of ER-positive breast cancer.Although increasing evidence suggest that dysregulation of mi RNA influences the tamoxifen sensitivity,the mechanism of the cross-talk between mi RNA and ERα signaling remain unclear.A large number of studies have shown that mi RNA is involved in the development and progression of malignant tumor,however,the role of mi R-575 in breast tumorigenesis and progression is still limited.Therefore,this paper makes an in-depth study about the effect and mechanism of mi R-575 on the sensitivity of tamoxifen in ER-positive breast cancer.Content: We first compared the overall survival in breast cancer patients with different mi R-575 expression levels by KM-plotter.We detected the expression level of mi R-575 in different breast cancer tissues and cell lines,explored the effect on breast cancer cell proliferation and tamoxifen sensitivity through in vivo and in vitro experiments.Then,this research elucidated the molecular mechanism that the mi R-575 regulated tamoxifen sensitivity in ER-positive breast cancer,which included the correlation between mi R-575 and estrogen in ER-positive breast cancer,the targets of mi R-575 and the regulatory mechanism of the target.Methods: We first compared the overall survival rate of breast cancer patients with different mi R-575 expression levels by Kaplan-Meier plotter.The expression of mi R-575 in breast cancer tissues and cell lines was determined by RT-q PCR.We generated stable mi R-575-overexpressed T47 D cell line,transiently mi R-575 inhibitor to decrease the expression of mi R-575.A series of functional experiments verified the effect of mi R-575 on the proliferation ability of ER positive breast cancer cells.The binding site of ERα in the mi R-575 promoter region was analyzed by Ch IP and luciferase assays.The interaction of ERα with CDKN1B/p27,Cyclin D1 or BRCA1 was determined by IP analysis.The protein expression levels and localization were analyzed by western blotting and immunofluorescence,respectively.Finally,we explored whether the mi R-575-p27 axis in ER + breast cancer cells can affect the tumor response to tamoxifen treatment in vitro and in vivo.Results: We analyzed the expression level of mi R-575 in breast cancer patients by Kaplan-Meier plotter and found that patients with high mi R-575 expression had a significantly poor outcome compared to those with low mi R-575 expression.More importantly,we found that mi R-575 high ER+ breast cancer patients have poor prognosis compared to mi R-575 low patients.However,the expression of mi R-575 was not associated with the prognosis in ER-breast cancer patients.Next,we determined the expression of mi R-575 in breast cancer specimens and breast cancer cell lines by RT-q PCR.We observed upregulation of mi R-575 expression in most ER+ breast cancers.mi R-575 can promote the proliferation of ER + breast cancer both in vivo and in vitro.Estrogen / ERα activates mi R-575 expression.And we explored p27 was the target of mi R-575,the interaction between p27 and ERα,the interaction between BRCA1 and ERα and regulated by mi R-575.Forced expression of mi R-575 decreased the tamoxifen sensitivity by targeting CDKN1B/p27.ERα bound the mi R-575 promoter for its transcriptional activity,whereas tamoxifen treatment down-regulated the expression of mi R-575 in ER+ breast cancer.Furthermore,CDKN1B/p27 or BRCA1 antagonizes the ERα activity by inhibition of nuclear translocation and the interaction with Cyclin D1.Conclusion: Our data uncover the ERα-mi R-575-CDKN1B/p27 feedback loop in ER+ breast cancer,suggesting that mi R-575 can be used as a prognostic biomarker in patients with ER+ breast cancer,as well as a predictor or a promising target for tamoxifen sensitivity.