Potential tumorsuppressive role of microRNA99a3p in sunitinibresistant renal cell carcinoma cells through the regulation of RRM2

Sunitinib is the most common primary moleculartargeted agent for metastatic clear cell renal cell carcinoma (ccRCC); however, intrinsic or acquired sunitinib resistance has become a significant problem in medical practice. The present study focused on microRNA (miR)99a3p, which was significantly downregulated in clinical sunitinibresistant ccRCC tissues in previous screening analyses, and investigated the molecular network associated with it. The expression levels of miR99a3p and its candidate target genes were evaluated in RCC cells, including previously established sunitinibresistant 786o (SUR786o) cells, and clinical ccRCC tissues, using reverse transcriptionquantitative polymerase chain reaction. Gainoffunction studies demonstrated that miR99a3p significantly suppressed cell proliferation and colony formation in RCC cells, including the SUR786o cells, by inducing apoptosis. Based on in silico analyses and RNA sequencing data, followed by luciferase reporter assays, ribonucleotide reductase regulatory subunitM2 (RRM2) was identified as a direct target of miR99a3p in the SUR786o cells. Lossoffunction studies using small interfering RNA against RRM2 revealed that cell proliferation and colony growth were significantly inhibited via induction of apoptosis, particularly in the SUR786o cells. Furthermore, the RRM2 inhibitor Didox (3,4dihydroxybenzohydroxamic acid) exhibited anticancer effects in the SUR786o cells and other RCC cells. To the best of our knowledge, this is the first report demonstrating that miR99a3p directly regulates RRM2. Identifying novel genes targeted by tumorsuppressive miR99a3p in sunitinibresistant RCC cells may improve our understanding of intrinsic or acquired resistance and facilitate the development of novel therapeutic strategies.