环状 RNA EXOC6B 通过 miR-376c-3p/FOXO3 轴抑制卵巢癌细胞增殖、迁移并增加其对紫杉醇的敏感性。

CircEXOC6B Suppresses the Proliferation and Motility and Sensitizes Ovarian Cancer Cells to Paclitaxel Through miR-376c-3p/FOXO3 Axis.

机构信息

Department of Gynecologic Oncology, Qingdao Central Hospital, Qingdao, China.

Department of Reproductive Center, Qingdao Hospital for Women and Children, Qingdao, China.

出版信息

Cancer Biother Radiopharm. 2022 Nov;37(9):802-814. doi: 10.1089/cbr.2020.3739. Epub 2020 Oct 2.

DOI:10.1089/cbr.2020.3739

PMID:33006481

Abstract

Circular RNAs (circRNAs) are regarded as important regulators in the tumorigenesis of multiple cancers. However, the characterization of circRNA exocyst complex component 6B (circEXOC6B) in ovarian cancer is barely known. Quantitative real-time polymerase chain reaction (qRT-PCR) was utilized to detect the enrichment of circEXOC6B, microRNA-376c-3p (miR-376c-3p), and forkhead box O3 (FOXO3). Cell proliferation was examined by Cell Counting Kit-8 (CCK8) assay and colony formation assay. Cell metastasis was measured by transwell assays. Western blot assay was conducted to examine the expression of proliferation and metastasis-related proteins and FOXO3. The chemoresistance of ovarian cancer cells was analyzed by CCK8 assay. Flow cytometry was used to detect cell apoptosis. The activities of caspase3 and caspase9 were analyzed through using colorimetric assay kits. The direct interaction between miR-376c-3p and circEXOC6B or FOXO3 was predicted by StarBase software and confirmed by dual-luciferase reporter assay and RNA immunoprecipitation (RIP) assay. Murine xenograft assay was conducted to verify the role of circEXOC6B on the paclitaxel (PTX) resistance of ovarian cancer cells . The level of circEXOC6B was notably decreased in ovarian cancer tissues. Low level of circEXOC6B was associated with malignant pathological characteristics in ovarian cancer patients. CircEXOC6B suppressed the proliferation and motility and decreased the chemoresistance of ovarian cancer cells to PTX. CircEXOC6B functioned through directly targeting and downregulating miR-376c-3p. FOXO3 was a direct target of miR-376c-3p, and the abundance of FOXO3 was regulated by circEXOC6B/miR-376c-3p axis. CircEXOC6B accelerated the PTX sensitivity of ovarian cancer cells through acting as a decoy of miR-376c-3p to upregulate FOXO3 . CircEXOC6B suppressed the progression and PTX resistance of ovarian cancer cells through sequestering miR-376c-3p, thus enhancing FOXO3 level.

摘要

环状 RNA（circRNAs）被认为是多种癌症发生肿瘤的重要调节因子。然而，关于卵巢癌中环状 exocyst 复合物成分 6B（circEXOC6B）的特征几乎未知。定量实时聚合酶链反应（qRT-PCR）用于检测 circEXOC6B、微小 RNA-376c-3p（miR-376c-3p）和叉头框蛋白 O3（FOXO3）的富集。通过细胞计数试剂盒-8（CCK8）测定和集落形成测定来检测细胞增殖。通过 Transwell 测定来测量细胞转移。通过 Western blot 测定来检测增殖和转移相关蛋白和 FOXO3 的表达。通过 CCK8 测定分析卵巢癌细胞的化疗耐药性。通过流式细胞术检测细胞凋亡。通过比色法测定试剂盒分析 caspase3 和 caspase9 的活性。通过 StarBase 软件预测 miR-376c-3p 与 circEXOC6B 或 FOXO3 的直接相互作用，并通过双荧光素酶报告基因测定和 RNA 免疫沉淀（RIP）测定进行验证。进行小鼠异种移植实验以验证 circEXOC6B 对卵巢癌细胞紫杉醇（PTX）耐药性的作用。circEXOC6B 在卵巢癌组织中明显下调。circEXOC6B 水平低与卵巢癌患者恶性病理特征相关。circEXOC6B 抑制卵巢癌细胞的增殖和运动，并降低卵巢癌细胞对 PTX 的化疗耐药性。circEXOC6B 通过直接靶向和下调 miR-376c-3p 发挥作用。FOXO3 是 miR-376c-3p 的直接靶标，FOXO3 的丰度受 circEXOC6B/miR-376c-3p 轴调节。circEXOC6B 通过充当 miR-376c-3p 的诱饵来上调 FOXO3，从而加速卵巢癌细胞对 PTX 的敏感性。circEXOC6B 通过隔离 miR-376c-3p 来抑制卵巢癌细胞的进展和 PTX 耐药性，从而提高 FOXO3 水平。