MiR-494 Inhibits Epithelial Ovarian Cancer Growth by Targeting c-Myc

Abstract

BACKGROUND Epithelial ovarian cancer (EOC) is the most lethal malignant gynecological cancer. MicroRNAs (miRNAs) play important roles in the pathogenesis of ovarian cancer. The role of miR-494 in EOC has not been fully investigated. MATERIAL AND METHODS MiR-494 levels in ovarian cancer tissues and cells were tested by qRT-PCR. Cells were transfected with miR-494 mimics or miR-494 ASO by Lipofectamine. Bioinformatics algorithms from TargetScanHuman were used to predict the target genes of miR-494. The c-Myc protein level was assayed by Western blot. The interaction between miR-494 and c-Myc was confirmed by dual luciferase assays. RESULTS MiR-494 showed low levels in EOC tissues and cells. Overexpression of miR-494 inhibited cell growth and migration of EOC cells and vice versa. c-Myc is the targeted gene of miR-494. CONCLUSIONS MiR-494 has an anti-tumor role in EOC via c-Myc.

Figure 1

The miR-494 level in epithelial ovarian cancer tissues. The miR-494 levels in the 15 EOC tissues and matched adjacent normal cancer tissues were tested by qRT-PCR. U6 snRNA was used as blank control (A). The mean expression of miR-494 in the 15 EOC tissues and matched adjacent normal cancer tissues were calculated, and the mean level of 15 normal ovarian tissues were arbitrarily defined as 100% (B). The qRT-PCR experiments were performed 3 times. Data are mean ±S.D. * P<0.05

Figure 2

Overexpression of miR-494 inhibited cells growth and invasion and vice versa. The miR-494 levels in normal ovarian tissues, HEK293, SKOV3, and HO8910 cells were tested by qRT-PCR. The miR-494 level in normal ovarian tissues was deliberately treated as 100% (A). After miR-494 mimics transfection, the miR-494 levels in SKOV3 and HO8910 cells were examined. The miR-494 level in negative control was deliberately treated as 100% (B). After miR-494 ASO transfection, the miR-494 levels in SKOV3 and HO8910 cells were examined. The miR-494 level in negative control transfected cells was deliberately treated as 100% (C). After miR-494 mimics or miR-494 ASO transfection, the cellular proliferation of SKOV3 and HO8910 were tested by MTT analysis (D). After miR-494 mimics or miR-494 ASO transfection, cells were collected for migration test (E). The experiments were performed 3 times. Data are mean ±S.D. * P<0.05

Figure 3

c-Myc was targeted by miR-494. Alterations of c-Myc were visualized by cBioPortal for Cancer Genomics. Mutation, deletion, amplification, and other alterations are shown in different colors. The most frequent alteration of c-Myc in different types of cancer is amplification. The CAN data stand for copy number alteration data (A). The binding site for miR-494 in c-Myc was mutated (B). HEK293 cells were co-transfected with miR-494 mimics or control and reporter plasmid or the mutant 3′UTR of c-Myc, together with the controls. At 48 h after transfection, the luciferase activity was measured (C). MiR-494 mimics was transfected into HEK293 cells. At 48 h later, the c-Myc protein level was tested by Western blot (D). The mRNA levels of c-Myc in 15 EOC tissues were assayed by qRT-PCR (E). The experiments were performed 3 times. Data are mean±S.D. *P<0.05