Expression Patterns And Functional Study Of Cancer-Testis Gene PAGE5 In Breast Cancer

Objective:Breast cancer remains the most common malignant neoplasms and one of the leading causes of cancer deaths in women around the world.Owing to the development of earlier detection and adjuvant therapy,the mortality has significantly reduced,however,breast cancer is an extremely heterogeneous disease that still lacks a standard therapy strategy suitable for all types of breast cancer.Consequently,further research of new targets for breast cancer diagnosis and therapy is still urgently needed.Cancer-Testis antigen(CT antigen)as a kind of tumor-associated antigens that are normally expressed in the testis and can be induced in cancer cells.CT genes as tumor markers will play important roles in cancer screening,diagnosis,disease progression,treatment and prognosis assessment.Previous research reported that CT gene,PAGE5,was highly expressed in melanoma,renal,and lung cancer cells;moreover,it could regulate the expression of apoptotic and anti-apoptotic genes and promote the survival of melanoma cells.However,the biological roles of PAGE5 in cancer development especially in breast cancer remains unclear.The aim of this study was to investigate the roles of PAGE5 in breast cancer proliferation and clinical significance,exploring the functions of PAGE5 in breast cancer progress.Methods:The GTEx(Genotype-Tissue Expression),our multi-tissue kit and TCGA(The Cancer Genome Atlas)project all proved the expression patterns of PAGE5 in normal tissues and breast cancers.Subsequently,the immunohistochemical(IHC)analysis were used to detect the expression levels of PAGE5 in breast cancer tissues and adjacent tissues,as well as analyzing the correlation with clinicopathological characteristics.We also analyzed the association between PAGE5 expression and prognosis in TCGA project.Moreover,we explored the function of PAGE5 in breast cancer cells proliferation through Cell Counting Kit-8(CCK8)and colony formation analysis.Flow cytometry analyses detected the influence of cell cycle when overexpressed PAGE5.Finally,we observed the expression of cell cycle inhibitors P15 and P57 when overexpressed PAGE5 through RT-qPCR(Real-time Quantative Polymerse Chin Rection)and Western Blot.Results:The GTEx and our multi-tissue kit all showed that PAGE5 was predominantly expressed in the testis for all the normal tissues.We discovered that the expression of PAGE5 was high in breast cancer tissues while almost no expression in adjacent tissues of TCGA project.Additionally,the IHC analysis revealed that PAGE5 expression was significantly higher in breast cancer tissues compared to adjacent tissues and the percentage of PAGE5-positive breast cancer tissues was 66.3%,while adjacent tissues was only 10.5%.Furthermore we explored the clinical significance of PAGE5 and found its expression levels were remarkably associated with histological grade and clinical TNM stage.PAGE5 expression was significantly increased in high-grade breast cancers compared to low-grade tumors;TCGA project also showed that PAGE5 activation rates were significantly correlated with breast cancer pathologic stage,as activation rates increased at higher stages.Moreover,the expression levels of PAGE5 were negatively associated with prognosis.Subsequently,we explored the biological function of PAGE5 in breast cancer progression in vitro.CCK8 and clone forming array showed that overexpression of PAGE5 promoted,whereas knockdown suppressed breast cancer cells proliferation.Further analysis revealed that overexpression of PAGE5 promoted cell cycle G1-S phase transition by inhibiting cell cycle inhibitors P15 and P57.Conclusions:PAGE5 was a CT gene which promotes breast cancer cells proliferation through negatively regulating the expression of P15 and P57 and plays important roles in the progression of breast cancer,demonstrating that PAGE5 may be a promising maker for breast cancer diagnosis,treatment and prognostic evaluation.