Department of Neurology, Xinxiang Central Hospital, Xinxiang, Henan province, China.
Department of Critical Care Medicine, Qingpu Branch of Zhongshan Hospital Affiliated to Fudan University, Shanghai, China.
J Biochem Mol Toxicol. 2021 Sep;35(9):e22848. doi: 10.1002/jbt.22848. Epub 2021 Jul 30.
Temozolomide (TMZ) is the first-line chemotherapy drug for glioblastoma (GBM) but acquired TMZ resistance is frequently observed. Thus, a TMZ resistant GBM cell line U87-R was established to search for potential long noncoding RNAs (lncRNAs) used in TMZ resistance. In our study, LINC00511 was identified as a TMZ resistance-associated lncRNA in U87-R cells by transcriptome RNA sequencing. The potential functions of LINC00511 were evaluated by quantitative real-time polymerase chain reaction, cell viability assay, colony formation assay, western blot, soft agar assay, flow cytometry, tumor xenograft model, immunofluorescence, sphere formation assay, fluorescent in situ hybridization, luciferase reporter assay, and RNA pull-down assay. We found that LINC00511 was upregulated in U87-R cells and GBM samples, and correlated with poor prognosis of GBM patients. Silencing LINC00511 impaired TMZ tolerance of U87-R cells, while LINC00511 overexpression increased TMZ resistance of sensitive GBM cells. Wnt/β-catenin signaling was activated in U87-R cells, and inhibiting Wnt/β-catenin signaling enhanced TMZ sensitivity. Furthermore, LINC00511 was mainly distributed in the cytoplasm of GBM cells and regulated Wnt/β-catenin activation by acting as a molecular sponge for miR-126-5p. Multiple genes of Wnt/β-catenin signaling such as DVL3, WISP1, and WISP2 were targeted by miR-126-5p. MiR-126-5p restoration impaired TMZ resistance of GBM cells. In conclusion, our results provided a novel insight into acquired TMZ resistance of GBM cells and suggested LINC00511 as a potential biomarker or therapeutic target for GBM patients.
替莫唑胺（TMZ）是胶质母细胞瘤（GBM）的一线化疗药物，但经常观察到获得性 TMZ 耐药。因此，建立了 TMZ 耐药 GBM 细胞系 U87-R，以寻找用于 TMZ 耐药的潜在长非编码 RNA（lncRNA）。在我们的研究中，通过转录组 RNA 测序鉴定 LINC00511 是 U87-R 细胞中与 TMZ 耐药相关的 lncRNA。通过定量实时聚合酶链反应、细胞活力测定、集落形成测定、western blot、软琼脂测定、流式细胞术、肿瘤异种移植模型、免疫荧光、球体形成测定、荧光原位杂交、荧光素酶报告基因测定和 RNA 下拉测定评估 LINC00511 的潜在功能。我们发现 LINC00511 在 U87-R 细胞和 GBM 样本中上调，并与 GBM 患者的不良预后相关。沉默 LINC00511 削弱了 U87-R 细胞对 TMZ 的耐受性，而 LINC00511 的过表达增加了敏感 GBM 细胞对 TMZ 的耐药性。Wnt/β-catenin 信号在 U87-R 细胞中被激活，抑制 Wnt/β-catenin 信号增强了 TMZ 的敏感性。此外，LINC00511 主要分布在 GBM 细胞的细胞质中，并通过作为 miR-126-5p 的分子海绵来调节 Wnt/β-catenin 激活。Wnt/β-catenin 信号的多个基因，如 DVL3、WISP1 和 WISP2，是 miR-126-5p 的靶标。miR-126-5p 的恢复损害了 GBM 细胞对 TMZ 的耐药性。总之，我们的结果为 GBM 细胞获得性 TMZ 耐药提供了新的见解，并表明 LINC00511 可作为 GBM 患者的潜在生物标志物或治疗靶点。
