Department of Breast Surgery, The First Affiliated Hospital of Zhengzhou University, Zhengzhou City, Henan Province, China.
Cell Transplant. 2020 Jan-Dec;29:963689720929983. doi: 10.1177/0963689720929983.
Triple-negative breast cancer (TNBC) is one of the most aggressive cancer types with high recurrence, metastasis, and drug resistance. Recent studies report that long noncoding RNAs (lncRNAs)-mediated competing endogenous RNAs (ceRNA) play an important role in tumorigenesis and drug resistance of TNBC. Although elevated lncRNA DLX6 antisense RNA 1 (DLX6-AS1) has been observed to promote carcinogenesis in various cancers, the role in TNBC remained unclear. In this study, expression levels of DLX6-AS1 were increased in TNBC tissues and cell lines when compared with normal tissues or breast fibroblast cells which were determined by quantitative real-time PCR (RT-qPCR). Then, CCK-8 assay, cell colony formation assay and western blot were performed in CAL-51 cells transfected with siRNAs of DLX6-AS1 or MDA-MB-231 cells transfected with DLX6-AS1 over expression plasmids. Knock down of DLX6-AS1 inhibited cell proliferation, epithelial-mesenchymal transition (EMT), decreased expression levels of BCL2 apoptosis regulator (Bcl-2), Snail family transcriptional repressor 1 (Snail) as well as N-cadherin and decreased expression levels of cleaved caspase-3, γ-catenin as well as E-cadherin, while up regulation of DLX6-AS1 had the opposite effect. Besides, knockdown of DLX6-AS1 in CAL-51 cells or up regulation of DLX6-AS1 in MDA-MB-231 cells also decreased or increased cisplatin resistance of those cells analyzed by MTT assay. Moreover, by using dual luciferase reporter assay, RNA immunoprecipitation and RNA pull down assay, a ceRNA which was consisted by lncRNA DLX6-AS1, microRNA-199b-5p (miR-199b-5p) and paxillin (PXN) was identified. And DLX6-AS1 function through miR-199b-5p/PXN in TNBC cells. Finally, results of xenograft experiments using nude mice showed that DLX6-AS1 regulated cell proliferation, EMT and cisplatin resistance by miR-199b-5p/PXN axis in vivo. In brief, DLX6-AS1 promoted cell proliferation, EMT, and cisplatin resistance through miR-199b-5p/PXN signaling in TNBC in vitro and in vivo.
三阴性乳腺癌（TNBC）是一种侵袭性最强的癌症类型，具有高复发、转移和耐药性。最近的研究报告称，长链非编码 RNA（lncRNA）介导的竞争性内源 RNA（ceRNA）在 TNBC 的肿瘤发生和耐药中发挥重要作用。尽管在各种癌症中观察到上调的 lncRNA DLX6 反义 RNA 1（DLX6-AS1）促进了致癌作用，但在 TNBC 中的作用仍不清楚。在这项研究中，通过定量实时 PCR（RT-qPCR）确定，与正常组织或乳腺成纤维细胞相比，TNBC 组织和细胞系中 DLX6-AS1 的表达水平升高。然后，在转染 siRNAs 的 CAL-51 细胞或转染 DLX6-AS1 过表达质粒的 MDA-MB-231 细胞中进行 CCK-8 测定、细胞集落形成测定和 Western blot。DLX6-AS1 的敲低抑制细胞增殖、上皮-间充质转化（EMT），降低 BCL2 凋亡调节因子（Bcl-2）、Snail 家族转录阻遏物 1（Snail）以及 N-钙粘蛋白的表达水平，并降低 cleaved caspase-3、γ-catenin 和 E-cadherin 的表达水平，而上调 DLX6-AS1 则有相反的效果。此外，通过 MTT 测定分析，在 CAL-51 细胞中敲低 DLX6-AS1 或在 MDA-MB-231 细胞中上调 DLX6-AS1 也降低或增加了这些细胞对顺铂的耐药性。此外，通过双荧光素酶报告基因检测、RNA 免疫沉淀和 RNA 下拉实验，鉴定了一个由 lncRNA DLX6-AS1、microRNA-199b-5p（miR-199b-5p）和桩蛋白（PXN）组成的 ceRNA。并且 DLX6-AS1 通过 TNBC 细胞中的 miR-199b-5p/PXN 发挥作用。最后，使用裸鼠进行的异种移植实验结果表明，DLX6-AS1 通过体内 miR-199b-5p/PXN 轴调节细胞增殖、EMT 和顺铂耐药性。总之，DLX6-AS1 通过 miR-199b-5p/PXN 信号通路在体外和体内促进 TNBC 细胞的增殖、EMT 和顺铂耐药性。
