College of Pharmacy, Chongqing Medical University, Chongqing, P. R. China.
Department of Urology, Southwest Hospital, Army Medical University, Chongqing, P. R. China.
Clin Transl Med. 2022 Jul;12(7):e994. doi: 10.1002/ctm2.994.
Clear cell renal cell carcinoma (ccRCC) is a common malignant tumour of the urinary tract. The major causes of poor prognosis are the lack of early diagnosis and metastasis. Accumulating research reveals that circular RNAs (circRNAs) can play key roles in the development and the progression of cancer. However, the role of circRNAs in ccRCC is still uncertain.
The circRNAs microarray (n = 4) was performed to investigate the circRNAs with differential expression in ccRCC tissues. The candidate circRNA was selected based on the cut-off criteria, such as circRNA expression abundance, circRNA size and the design of divergent primers. The circ-transportin-3 (TNPO3) levels in ccRCC tissues were tested by quantitative real-time (qRT)-PCR (n = 110). The characteristics and subcellular localization of circ-TNPO3 were identified via RNase R assay, qRT-PCR and fluorescence in situ hybridization (FISH). Then, we explored the biological roles of circ-TNPO3 in ccRCC via the function experiments in vitro and in vivo. RNA pull-down, RNA immunoprecipitation, bioinformatic analysis, RNA-FISH assays and rescue assays were applied to validate the interactions between circ-TNPO3, insulin-like growth factor 2 mRNA-binding protein 2 (IGF2BP2) and serpin family H member 1 (SERPINH1) to uncover the underlying molecular mechanisms of circ-TNPO3.
We detected the obvious downregulation of circ-TNPO3 in ccRCC compared to matched adjacent normal tissues (n = 110). The lower circ-TNPO3 expression was found in ccRCC patients with distant metastasis, higher World Health Organization/International Society of Urologic Pathologists (WHO/ISUP) grade and more advanced tumour T stage. In vitro and in vivo, circ-TNPO3 significantly suppressed the proliferation and migration of ccRCC cells. Mechanistically, we elucidated that circ-TNPO3 directly bound to IGF2BP2 protein and then destabilized SERPINH1 mRNA. Moreover, IGF2BP2/SERPINH1 axis was responsible for circ-TNPO3's function of inhibiting ccRCC metastasis. Epithelial splicing regulatory protein 1 (ESRP1) was probably involved in the biogenesis of circ-TNPO3.
Circ-TNPO3 can suppress ccRCC progression and metastasis via directly binding to IGF2BP2 protein and destabilizing SERPINH1 mRNA. Circ-TNPO3 may act as a potential target for ccRCC treatment.
透明细胞肾细胞癌（ccRCC）是一种常见的泌尿道恶性肿瘤。预后不良的主要原因是缺乏早期诊断和转移。越来越多的研究表明，环状 RNA（circRNA）在癌症的发生和发展中起着关键作用。然而，circRNA 在 ccRCC 中的作用仍不确定。
采用 circRNA 微阵列（n=4）研究 ccRCC 组织中差异表达的 circRNAs。根据 circRNA 表达丰度、circRNA 大小和发散引物设计等截断标准，选择候选 circRNA。通过定量实时（qRT）-PCR（n=110）检测 ccRCC 组织中的 circ-transportin-3（TNPO3）水平。通过 RNase R 试验、qRT-PCR 和荧光原位杂交（FISH）鉴定 circ-TNPO3 的特征和亚细胞定位。然后，我们通过体外和体内的功能实验探索 circ-TNPO3 在 ccRCC 中的生物学作用。应用 RNA 下拉、RNA 免疫沉淀、生物信息学分析、RNA-FISH 测定和挽救测定来验证 circ-TNPO3、胰岛素样生长因子 2 mRNA 结合蛋白 2（IGF2BP2）和丝氨酸蛋白酶抑制剂家族 H 成员 1（SERPINH1）之间的相互作用，以揭示 circ-TNPO3 的潜在分子机制。
与匹配的相邻正常组织相比（n=110），我们检测到 ccRCC 中 circ-TNPO3 的明显下调。在有远处转移、更高的世界卫生组织/国际泌尿病理学会（WHO/ISUP）分级和更晚期肿瘤 T 分期的 ccRCC 患者中，circ-TNPO3 表达较低。在体外和体内，circ-TNPO3 显著抑制 ccRCC 细胞的增殖和迁移。在机制上，我们阐明 circ-TNPO3 可直接与 IGF2BP2 蛋白结合，然后使 SERPINH1 mRNA 不稳定。此外，IGF2BP2/SERPINH1 轴是 circ-TNPO3 抑制 ccRCC 转移功能的关键。上皮剪接调节蛋白 1（ESRP1）可能参与了 circ-TNPO3 的生物发生。
circ-TNPO3 通过直接结合 IGF2BP2 蛋白和使 SERPINH1 mRNA 不稳定，抑制 ccRCC 的进展和转移。circ-TNPO3 可能成为 ccRCC 治疗的潜在靶点。
