Nuclear AURKA Enhances Breast Cancer Stem Cell Phenotype

BackgroundOne of the most common malignant tumor in female is breast cancer,of which incidence still maintains a rapid increase.Although the clinic therapy for breast cancer has achieved good outcome,tumor relapse and metastasis are still difficult to overcome.Tumor cell's drug resistance is leading cause of tumor relapse.The chemoresistant tumor cells will be stronger after relapse.Cancer stem cell is a cancer cell population that has self-renewal ability and multipotent capability.Tumor chemoresistance has been shown to be associated with cancer stem cell,which can pump out drugs and is not sensitive to chemotherapy.The latant,dormant,but self-renewal cancer stem cell also contributes to tumor relapse.Aurora kinase A is a oncoprotein that overexpresses in various tumors,containing breast cancer.Aberrant activation of AURKA will promote tumorigenesis through inducing genome instability and aneuploid,disrupting cell porlarity,overcoming DNA damage checkpoint under DNA damage and spindle checkpoint upon G2/M arrest,therefore aberrant cell proliferation.AURKA overexpression in tumor cells is associated with poor prognosis,consistant with its roles in promoting tumor proliferation,migration and invision.AURKA kinase inhibitors showed significant cell cycle arrest and tumor inhibition effect.However,there is no considerable outcome in clinic therapy.ObjectiveThis project is aimed to investigate the oncogenic effect of nuclear localized AURKA on breast cancer stem cell.We want to show the nuclear localization of AURKA with multiple molecular biology methods and look for its role in tumor development.From this point,we try to uncover the kinase independent effect of AURKA,therefore present a combination therapy that involves AURKA kinase inhibition and nuclear localization inhibition,providing a new effective strategy for AURAK targeted therapy.MethodsWe test the localization of AURKA with the method of nuclear protein extraction,immunofluorescence in cells and immunohistochemistry in tissue.Flow cytometry was used to test the molecular surface markers of breast cancer stem cell.ALDEFLUOR assay and microsphere formation assay were performed to investigate the stemness of breast cancer cells.We performed the gene set enrichment analysis to analyze the regulation of AURKA on cancer stem cell related genes.Tumor formation in nude mice was conduct to test the tumor-promoting effect of nuclear localized AURKA and the sensibilization effect of AURKA nuclear localization inhibition on AURKA kinase inhibitor.Results1.We showed the nuclear localization of AURKA in breast cancer tissue and cell lines.Malignant transformation promotes AURKA to enter nuclear.2.Overexpressed AURKA increased breast cancer stemness.AURKA knockdown inhibit breast cancer stem cell(BCSC).3.Nuclear localized AURKA,but not cytoplasmic AURKA,is regulator of breast cancer stem cell.4.Blocking AURKA nuclear localization inhibit tumor growth and enhanced the anticancer effect of AURKA kinase inhibitor in xenograft.