MiR-182-5p inhibits colon cancer tumorigenesis, angiogenesis, and lymphangiogenesis by directly downregulating VEGF-C

Highlights

• MiR-182-5p is downregulated in colon cancer and associated with pathological stage.

• MiR-182-5p inhibits proliferation, migration, and invasion of colon cancer cells.

• VEGF-C is downregulated by miR-182-5p via direct binding.

• In turn, this downregulates VEGF-A, VEGFR-2, and -3 and inhibits ERK/AKT pathways.

• Ectopic miR-182-5p reduces tumor growth, angiogenesis and lymphangiogenesis in vivo.

Abstract

MicroRNAs (miRNAs) are gene modulators essential for biological processes. However, the precise functions of miRNAs in growth and development of colon cancer are still elusive. To clarify their role, here we analyzed a miRNA microarray of colon cancer. MiR-182-5p was found markedly downregulated in colon cancer tissues and cells, and strongly correlated with pathological stage, differentiation, and lymphatic metastasis. In vitro, miR-182-5p overexpression repressed colon cancer cell proliferation, colony formation, migration, and invasion, and triggered G1 arrest and apoptosis. MiR-182-5p overexpression also downregulated vascular endothelial growth factor (VEGF)-C and inhibited the activity of a luciferase reporter containing the VEGF-C 3′-untranslated region. Moreover, miR-182-5p overexpression in colon cancer cells and human umbilical vein endothelial cells (HUVECs) downregulated VEGF-A as well as VEGF receptor (VEGFR)-2 and VEGFR-3, thereby inhibiting the phosphorylation of ERK and AKT. In vivo, miR-182-5p overexpression strikingly suppressed oncogenicity of SW620 cells as well as angiogenesis and lymphangiogenesis of xenograft tumors in nude mice. These data indicate that miR-182-5p regulates colon cancer tumorigenesis partially through modulating angiogenesis and lymphangiogenesis by targeting VEGF-C, and inhibiting ERK and AKT signaling pathways.

Introduction

Colon cancer is a very common and high-mortality cancer worldwide [1]. Every year, 1.2 million new cases are diagnosed and ~0.6 million colon cancer-related deaths occur globally [2]. The 5-year survival rate for this cancer ranges between 40% and 60% [3]. Due to high incidence and poor prognosis of colon cancer, it is imperative to elucidate the mechanisms underlying its tumorigenesis and progression to improve diagnosis and therapy. So far, no effective therapy has been developed for metastatic colon cancer, highlighting that this cancer remains a serious health threat.

MicroRNAs (miRNAs) can modulate gene expression by directly targeting mRNAs, leading to their translational suppression or instability [4,5]. Mounting evidence confirms miRNAs as therapeutic targets and markers for cancer diagnosis [[6], [7], [8], [9]]. Several miRNAs are known to act as vital modulators of angiogenesis and lymphangiogenesis in cancer [10,11]. For instance, miR-526b and miR-655 promote tumor-associated angiogenesis and lymphangiogenesis in breast cancer [12]. However, the exact functions of miRNAs in growth and development of colon cancer are still elusive.

During tumor growth, VEGFs function as pivotal modulators of lymphangiogenesis and angiogenesis [13]. VEGFs and their receptors (VEGFRs) are also major targets for the pathological treatment of lymphangiogenic and angiogenic signaling [14]. VEGF-A mainly regulates angiogenesis, whereas VEGF-C functions in lymphangiogenesis [15,16]. Recent evidence indicates that VEGF-C modulates angiogenesis as well as lymphangiogenesis [17,18]. VEGF-C is likely to be a better target than VEGF-A to suppress angiogenesis and lymphangiogenesis, since VEGF-C silencing by RNA interference also causes a decrease in VEGF-A expression [19]. Therefore, short-term anti-VEGF-A treatment might be clinically effective only to a certain degree. VEGF-C facilitates tumor development through several mechanisms, and VEGF-C/VEGFR-3 autocrine signaling correlate with tumor invasion, proliferation, and lymph node metastasis [[20], [21], [22]]. Thus, VEGF-C is a promising target for modulating tumor metastasis and development. Nonetheless, in contrast to abundant data on VEGF-A, literature on VEGF-C remains scarce. Moreover, whether VEGF-C can be a valuable candidate target for colon cancer is unknown.

In the present research, GSE48267 datasets were screened to identify differentially expressed miRNAs in colon cancer tissues and non-tumor colon tissues. MiR-182-5p was identified as differentially expressed in colon cancer. Although this miRNA has been confirmed to exert an inhibitory effect in multiple tumors [[23], [24], [25]], its action mechanism in colon cancer still needs exploration.

Therefore, in this study, the role of miR-182-5p in color cancer tumorigenesis as well as its effects on angiogenesis and lymphangiogenesis were investigated in vitro and in vivo.