microRNA‐99a‐5p induces cellular senescence in gemcitabine‐resistant bladder cancer by targeting SMARCD1

• Published in: Molecular oncology 2022-03, Vol.16 (6), p.1329-1346
• Main Authors: Tamai, Motoki, Tatarano, Shuichi, Okamura, Shunsuke, Fukumoto, Wataru, Kawakami, Issei, Osako, Yoichi, Sakaguchi, Takashi, Sugita, Satoshi, Yonemori, Masaya, Yamada, Yasutoshi, Nakagawa, Masayuki, Enokida, Hideki, Yoshino, Hirofumi
• Format: Article
• Language: English
• Subjects: Apoptosis; Bladder cancer; Cancer; Cell Line, Tumor; Cell Proliferation; cellular senescence; Cellular Senescence - genetics; Chemotherapy; Chromosomal Proteins, Non-Histone - genetics; Chromosome 5; Comparative analysis; Deoxycytidine - analogs & derivatives; Drug resistance; Gemcitabine; gemcitabine resistance; Gene Expression Regulation, Neoplastic; Humans; Kinases; Laboratories; Metastasis; MicroRNA; MicroRNAs; MicroRNAs - genetics; miRNA; miR‐99a‐5p; RNA sequencing; Scientific equipment and supplies industry; Senescence; Sequence analysis; SMARCD1; Tumors; Urinary Bladder Neoplasms - drug therapy; Urinary Bladder Neoplasms - genetics
• ISSN: 1574-7891; 1878-0261
• DOI: 10.1002/1878-0261.13192

Patients with advanced bladder cancer are generally treated with a combination of chemotherapeutics, including gemcitabine, but the effect is limited due to acquisition of drug resistance. Thus, in this study, we investigated the mechanism of gemcitabine resistance. First, gemcitabine‐resistant cells were established and resistance confirmed in vitro and in vivo. Small RNA sequencing analyses were performed to search for miRNAs involved in gemcitabine resistance. miR‐99a‐5p, selected as a candidate miRNA, was downregulated compared to its parental cells. In gain‐of‐function studies, miR‐99a‐5p inhibited cell viabilities and restored sensitivity to gemcitabine. RNA sequencing analysis was performed to find the target gene of miR‐99a‐5p. SMARCD1 was selected as a candidate gene. Dual‐luciferase reporter assays showed that miR‐99a‐5p directly regulated SMARCD1. Loss‐of‐function studies conducted with si‐RNAs revealed suppression of cell functions and restoration of gemcitabine sensitivity. miR‐99a‐5p overexpression and SMARCD1 knockdown also suppressed gemcitabine‐resistant cells in vivo. Furthermore, β‐galactosidase staining showed that miR‐99a‐5p induction and SMARCD1 suppression contributed to cellular senescence. In summary, tumor‐suppressive miR‐99a‐5p induced cellular senescence in gemcitabine‐resistant bladder cancer cells by targeting SMARCD1. Here, we established gemcitabine‐resistant bladder cancer (BC) cell line by stepwise exposure to gemcitabine in vitro. We also confirmed gemcitabine resistance in a xenograft mouse model. Our data showed downregulation of miR‐99a‐5p and upregulation of the miR‐99a‐5p target SMARCD1 in gemcitabine‐resistant BC cells, following cellular senescence inhibition. In summary, miR‐99a‐5p induced cellular senescence in gemcitabine‐resistant BC by targeting SMARCD1.