Unveiling the Role of miR-183-5p and miR-187-3p in Breast Cancer

Abstract

Background: Breast cancer (BC) is the most common type of cancer in incidence and mortality among women both worldwide and in Lebanon, which renders it a major health problem. Despite extensive research in this field, incidence and mortality rates are significantly increasing. BC is a heterogeneous disease composed of several subtypes characterized by distinct clinical outcomes. Thus, unveiling new players in BC development might lead to better understanding of its tumorigenesis. Studies have reported important roles of microRNAs (miRNAs) in the tumorigenesis and progression of cancer. It has been shown that the dysregulation of these small, endogenously expressed noncoding RNA molecules in BC participates in its tumorigenesis. Recent data in our lab has shown that miR-183-5p is one of the most upregulated miRNAs in estrogen-receptor positive (ER+) Lebanese BC tissues compared to normal adjacent tissues, by a microarray profiling analysis. In addition, an ongoing project in the lab has shown a significant upregulation of miR-187-3p in BC. Thus, our aim was to assess the roles of miR-183-5p and miR-187-3p in ER+ BC tumorigenesis.

Methods and Results: Dysregulation of miR-183-5p and miR-187-3p was validated in ER+ BC tissues downloaded from TCGA database compared to normal adjacent tissues (NAT) using DEseq2 tool. It was shown that both miRNAs are significantly upregulated in ER+ BC. Then, potential shared targets and target pathways between miR-183-5p and miR-187-3p were uncovered using miRTargetLink 2.0 tool. Then, some targets shown to be implicated in BC or in cancer in general were chosen for further investigation. These include MBNL1, BCL6, SMAD4, FGF9, and DAB1. After that, the correlations between miR-183-5p, miR-187-3p and the chosen shared targets expression with overall survival (OS) in ER+ BC were assessed using Kaplan-Meier (KM) plot analysis. An inverse correlation was observed between the effects of miR-183-5p and miR-187-3p expression with that of MBNL1, SMAD4, FGF9, and BCL6 on OS in ER+ BC where a low expression of the miRNAs of interest while a high expression of the targets was correlated with a better OS. In contrast, a direct correlation was observed between the effects of miRNAs and DAB1 expression on OS. This indicated that DAB1 may not be a direct target for these miRNAs in ER+ BC. Accordingly, the study was focused on the other targets (MBNL1, SMAD4, BCL6, and FGF9). After that, the dysregulation of miR-183-5p and miR-187-3p in BC cell lines was assessed by RT-qPCR where MCF-7 was further chosen as an in vitro model. miR-183-5p was shown to be upregulated in MCF-7 cell line while miR-187-3p was shown to be downregulated. So, the expression of the miRNAs of interest was modulated in MCF-7 cells by transfection with miR-183-5p inhibitor and miR-187-3p mimic. MTT assay and PI analysis were done to determine the effects of miR-183-5p downregulation and miR-187-3p overexpression on proliferation and cell cycle progression of MCF-7 cells. Both the downregulation of miR-183-5p and the overexpression of miR-187-3p had no effects on the proliferation of the cells. No effect was detected on cell cycle progression with downregulation of miR-183-5p whereas a slight increase in G2-M phase was observed with overexpression of miR-187-3p. Finally, RT-qPCR data showed that miR-183-5p downregulation upregulated the expression of SMAD4 and MBNL1 genes, while the overexpression of miR-187-3p unexpectedly upregulated MBNL1 mRNA levels in MCF-7 cells.

Conclusion: Overall, our study suggests the potential of miR-183-5p and miR-187-3p to be active players in estrogen-receptor (ER+) BC through the modulation of different mRNA targets. However, further studies are required to validate their role.