Investigation the effect of siRNA against CD133 on the apoptosis, proliferation, and migration in prostate cancer

Abstract

Introduction: One of the major barriers in cancer therapy is resistance to conventional therapies. According to several studies, cancer stem cells (CSCs) are the major cancer-related problem. CSCs are a small subpopulation of cells within tumors that drive chemoresistance and tumor recurrence in various cancers. CD133 (prominin-1) as a CSC surface marker displayed stem cell-like properties, tumorigenic capacity, drug resistance, migration ability, and the capacity of spheroid and colony forming in various cancers. CD133 has been known as an effective target in the field of cancer therapy. However, the exact role and the underlying molecular mechanism of CD133 in prostate cancer (PCa) remains unclear. The purpose of this study was to explore the potential function and mechanism of CD133 siRNA and paclitaxel in the inhibition of migration and reduction of chemoresistance in prostate cancer cells. Methods: In this study, we used quantitative real-time PCR (qRT-PCR) to measure CD133 expression in metastasis-derived prostate cancer cell line, LNCaP. The cells were transfected with CD133 siRNA using transfection reagent. The cytotoxic effects of CD133 siRNA, paclitaxel alone and the combination of CD133 siRNA/paclitaxel on LNCaP cells were determined using an MTT assay. The migration and invasive capacity of LNCaP cells after treatment by CD133 siRNA, paclitaxel alone and the combination of CD133 siRNA/paclitaxel were examined by scratch wound healing assay. Subsequently, in order to investigate the effect of CD133 siRNA on CSCs properties in the LNCaP cells after treatment by CD133 siRNA, paclitaxel alone and the combination of CD133 siRNA/paclitaxel, spheroid assay, colony formation assay, and cancer cell II proliferation were examined. In addition, Flow cytometry and 4′, 6‐diamidino‐2‐phenylindole staining analysis were used to determine that CD133 siRNA increased the paclitaxel‐induced apoptosis. We also used qRT-PCR to study the effect of CD133 silencing on the AKT/mTOR/C-myc axis and the expression levels of pro-metastatic genes and EMT markers in CD133+ prostate CSCs. Results: We find that the CD133 siRNA significantly reduced both mRNA expression levels of CD133 in 48 hours after transfection and dose-dependent manner in LNCaP cells. More importantly, we observed that CD133 siRNA and paclitaxel treatment significantly reduced cell viability and proliferation, also inhibited cells invasion and migration by changing the expression levels of a number of pro-metastatic genes and EMT markers in LNCaP cells in vitro. Moreover, we observed the functional relationship between CD133 knockdown and decreased activation of Akt/mTOR/C-myc axis in LNCaP cells.