MicroRNA-215 acts as a tumor suppressor in breast cancer by targeting AKT serine/threonine kinase 1

Abstract

There are accumulating reports that microRNAs are dysregulated in a number of human cancer types, and that they may function as tumor suppressors or oncogenes in tumorigenesis and tumor development. microRNA-215 (miR-215) has been identified as a tumor suppressor in epithelial ovarian, pancreatic, non-small cell lung and colon cancer, whereas it may act as an oncogene in gastric and cervical cancer. The role of miR-215 in breast cancer carcinogenesis and progression has yet to be elucidated. In the present study, the expression level of miR-215 was determined in breast cancer tissues and cell lines using the reverse transcription-quantitative polymerase chain reaction. The effects of miR-215 overexpression on proliferation and the invasive capacity of breast cancer cells were assessed using MTT and cell invasion assays. The results revealed that miR-215 was significantly downregulated in breast cancer tissues and cell lines. Restoration of miR-215 expression inhibited the proliferation and invasion of breast cancer cells. The underlying molecular mechanism for the suppression of proliferation and invasion by miR-215 was investigated. AKT serine/threonine kinase 1 (AKT1) was validated as a novel direct target of miR-215, and the effect of AKT1 small interfering RNA mimicked the effect of miR-215 overexpression in breast cancer cells. These results indicated that miR-215 acted as a tumor suppressor, and that its downregulation in tumor tissues may contribute to the carcinogenesis and progression of breast cancer, indicating that miR-215 may be a novel therapeutic target for the treatment of breast cancer.

Keywords: AKT serine/threonine kinase; breast cancer; microRNA-215; targeted therapy; tumor suppressor.

Figure 1.

miR-215 was downregulated in breast cancer tissues and cell lines. (A) Expression levels of miR-215 were significantly decreased in the breast cancer tissues compared with matched adjacent non-tumor breast tissues. Data are presented as the mean ± standard deviation. *P<0.05 vs. adjacent non-tumor breast tissues. (B) Relative expression of miR-215 was lower in all breast cancer cell lines examined compared with MCF-10A. *P<0.05 compared with MCF-10A. miR-215, microRNA-215.

Figure 2.

Effects of miR-215 overexpression on the proliferation and invasion of breast cancer cells. (A) Overexpression of miR-215 was simulated by transfecting miR-215 mimic into MCF-7A and MDA-MB-231 cells. miR-215 expression was measured using the reverse transcription-quantitative polymerase chain reaction 48 h after transfection. (B) Cell proliferation was determined using an MTT assay. MCF-7A and MDA-MB-231 cells transfected with miR-215 mimic exhibited a decreased rate of proliferation compared with cells transfected with NC. (C) miR-215 overexpression decreased the invasive capacity of MCF-7A and MDA-MB-231 cells compared with NC groups. *P<0.05 compared with NC-transfected cells. mirR-215, microRNA-215; NC, negative control.

Figure 3.

miR-215 overexpression decreased AKT1 expression in breast cancer cells. (A) miR-215-binding sites in the 3′UTR of AKT1. (B) The miR-215 mimic inhibited the expression of AKT1 at the protein level in MCF-7A and MDA-MB-231 cells. *P<0.05 compared with NC-transfected cells. miR-215, microRNA-215; AKT1, AKT serine/threonine kinase 1; 3′UTR, 3′-untranslated region; Wt, wild-type; mut, mutant; NC, negative control.

Figure 4.

Relative luciferase activities measured in HEK293T cells following co-transfection with the luciferase report vectors and miR-215 mimic or NC. *P<0.05 compared with NC-transfected cells. miR-215, microRNA-215; NC, negative control; AKT1, AKT serine/threonine kinase 1; 3′UTR, 3′-untranslated region; Wt, wild-type; mut, mutant.

Figure 5.

Effects of decreased AKT1 expression on the proliferation and invasion of breast cancer cells. (A) Western blot analysis revealed that AKT1 was markedly downregulated in MCF-7A and MDA-MB-231 cells transfected with AKT1 siRNA. (B) MCF-7A and MDA-MB-231 cells transfected with AKT1 siRNA exhibited a decreased rate of proliferation compared with cells transfected with scrambled siRNA. (C) Underexpression of AKT1 decreased the relative invasive capacity of MCF-7A and MDA-MB-231 cells. *P<0.05 compared with scrambled siRNA-transfected cells. AKT1, AKT serine/threonine kinase 1; siRNA, small interfering RNA.