Upregulation of MicroRNA-1246 Is Associated with BRAF Inhibitor Resistance in Melanoma Cells with Mutant BRAF

Abstract

PurposeIntrinsic and acquired resistance limit the therapeutic benefits of inhibitors of oncogenic BRAF in melanoma. To identify microRNAs (miRNAs) associated with resistance to a BRAF inhibitor, we compared miRNA expression levels in three cell lines with different BRAF inhibitor sensitivity.Materials and MethodsmiRNA microarray analysis was conducted to compare miRNA expression levels. Real-time quantitative reverse-transcription polymerase chain reaction (qRT-PCR) was performed to confirm the expression of differentially expressed miRNAs. The cellular effects of miR-1246 were further examined by MTT assay, immunoblotting analysis, cell cycle analysis, flow cytometric assay of apoptosis, and autophagy assay.ResultsThe miRNA microarray analysis and qRT-PCR identified five miRNAs (miR-3617, miR-92a-1, miR-1246, miR-193b-3p, and miR-17-3p) with expression that was consistently altered in two BRAF inhibitor-resistant cell lines. Among the five miRNAs, a miR-1246 mimic significantly reduced the antiproliferative effects of the BRAF inhibitor PLX4720 in BRAF inhibitor–resistant A375P (A375P/Mdr) cells, suggesting that miR-1246 upregulation confers acquired resistance to BRAF inhibition. In particular, apoptosis was identified as a major type of cell death in miR-1246–transfected cells; however, necrosis predominated in mimic-control-transfected cells, indicating that the resistance to PLX4720 in miR-1246 mimic-transfected cells is predominantly due to a reduction in necrosis. Furthermore, we found that miR-1246 promoted G2/M arrest through autophagy as a way to escape cell death by necrosis and apoptosis in response to PLX4720. The promotion of BRAF inhibitor resistance by miR-1246 was associated with lowered levels of p-ERK.ConclusionThese results suggest that miR-1246 may be a potential therapeutic target in melanoma with acquired resistance to BRAF inhibitors.

Mechanisms of Acquired BRAF Inhibitor Resistance in Melanoma: A Systematic Review

“…Resistance is also mediated at a post-transcriptional level through effects on translation mediated by RNA binding proteins (e.g., human antigen R and 4E-BP) [ 143 , 144 ], translation initiation complexes (e.g., eIF4F or eIF4E) [ 144 , 145 ], micro-RNAs ( Table 2 ) [ 59 , 146 , 147 , 148 , 149 , 150 , 151 , 152 , 153 , 154 ], and by modifications in wobble tRNA [ 155 ]. Some of these micro-RNAs (e.g., miR-199b-5p) are regulators of cell-cell signaling via VEGF and HIF-1α, while others (e.g., miR-4488 and miR-4443) are involved in autophagy regulation [ 149 ].…”

<p>MicroRNA-1246 Promotes Melanoma Progression Through Targeting FOXA2</p>

“…The upregulation of miR-1246 was related to the resistance of BRAF inhibitors. 26 However, it is unclear how miR-1246 regulates melanoma tumorigenesis. Therefore, we designed this study to further elucidate the regulatory mechanism of miR-1246 in melanoma.…”

Circulating miRNAs in Small Extracellular Vesicles Secreted by a Human Melanoma Xenograft in Mouse Brains

“…However, these results warrant further investigation addressing the specific functions of these two rarely characterized miRNAs. Interestingly, unlike the miRNAs identified in the total plasma, which are mostly tumor suppressors, many miRNAs contained in the sEVs have previously shown oncogenic functions (miR-1268a, miR-4258, miR-1246, miR-660, miR373) [61][62][63][64][65][66]. Among the multiple strategies developed by cancer cells to gain a survival advantage, sEVs are, indeed, one of the most concealed.…”