山奈酚通过调节胶质瘤中 Ku80 的稳定性抑制非同源末端连接修复。

Kaempferol inhibits non-homologous end joining repair via regulating Ku80 stability in glioma cancer.

机构信息

Laboratory of Experimental Hematology, School of Basic Medical Sciences, Binzhou Medical University, No. 346 Guanhai Road, Yantai, Shandong 264003, China.

Institute of Integrated Medicine, Binzhou Medical University, Yantai 264003, China.

出版信息

Phytomedicine. 2023 Jul 25;116:154876. doi: 10.1016/j.phymed.2023.154876. Epub 2023 May 12.

DOI:10.1016/j.phymed.2023.154876

PMID:37210962

Abstract

BACKGROUND

Targeting DNA damage response and DNA repair proficiency of cancers is an important anticancer strategy. Kaempferol (Kae), a natural flavonoid, displays potent antitumor properties in some cancers. However, the precise underlying mechanism of Kae regulates DNA repair system are poorly understood.

PURPOSE

We aim to evaluate the efficacy of Kae in the treatment of human glioma as well as the molecular mechanism regarding DNA repair.

STUDY DESIGN

Effects of Kae on glioma cells were detected using CCK-8 and EdU labeling assays. The molecular mechanism of Kae on glioma was determined using RNAseq. The inhibition effects of Kae on DNA repair were verified using Immunoprecipitation, immunofluorescence, and pimEJ5-GFP report assays. For in vivo study, orthotopic xenograft models were established and treated with Kae or vehicle. Glioma development was monitored by bioluminescence imaging, Magnetic Resonance Imaging (MRI), and brain sections Hematoxylin/Eosin (HE) staining. Immunohistochemical (IHC) analysis was used to detect expression of Ku80, Ki67 and γH2AX in engrafted glioma tissue.

RESULTS

We found that Kae remarkably inhibits viability of glioma cells and decreases its proliferation. Mechanistically, Kae regulates multiple functional pathways associated with cancer, including non-homologous end joining (NHEJ) repair. Further studies revealed that Kae inhibits release of Ku80 from the double-strand breaks (DSBs) sites via reducing ubiquitylation and degradation of Ku80. Therefore, Kae significantly suppresses NHEJ repair and induces accumulation of DSBs in glioma cells. Moreover, Kae displays a dramatic inhibition effects on glioma growth in an orthotopic transplantation model. These data demonstrate that Kae can induce deubiquitination of Ku80, suppress NHEJ repair and inhibit glioma growth.

CONCLUSION

Our findings indicate that inhibiting release of Ku80 from the DSBs by Kae may be a potential effective approach for glioma treatment.

摘要

背景

针对癌症的 DNA 损伤反应和 DNA 修复能力是一种重要的抗癌策略。山奈酚（Kae）是一种天然类黄酮，在一些癌症中显示出强大的抗肿瘤特性。然而，Kae 调节 DNA 修复系统的确切潜在机制尚不清楚。

目的

我们旨在评估 Kae 在治疗人类脑胶质瘤中的疗效以及与 DNA 修复相关的分子机制。

研究设计

使用 CCK-8 和 EdU 标记测定法检测 Kae 对神经胶质瘤细胞的影响。使用 RNAseq 确定 Kae 对神经胶质瘤的分子机制。使用免疫沉淀、免疫荧光和 pimEJ5-GFP 报告测定法验证 Kae 对 DNA 修复的抑制作用。在体内研究中，建立了原位异种移植模型，并使用 Kae 或载体进行治疗。通过生物发光成像、磁共振成像（MRI）和脑切片苏木精/伊红（HE）染色监测胶质瘤的发展。免疫组织化学（IHC）分析用于检测植入性胶质瘤组织中 Ku80、Ki67 和 γH2AX 的表达。

结果

我们发现 Kae 可显著抑制神经胶质瘤细胞的活力并降低其增殖。从机制上讲，Kae 调节与癌症相关的多个功能途径，包括非同源末端连接（NHEJ）修复。进一步的研究表明，Kae 通过减少 Ku80 的泛素化和降解来抑制 Ku80 从双链断裂（DSBs）位点的释放。因此，Kae 可显著抑制神经胶质瘤细胞中的 NHEJ 修复并导致 DSBs 积累。此外，Kae 在原位移植模型中对神经胶质瘤生长显示出显著的抑制作用。这些数据表明，Kae 可以诱导 Ku80 的去泛素化，抑制 NHEJ 修复并抑制神经胶质瘤的生长。