Microtubule-targeting agents can sensitize cancer cells to ionizing radiation by an interphase-based mechanism.
Microtubule-targeting agents can sensitize cancer cells to ionizing radiation by an interphase-based mechanism.
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DOI：
10.2147/ott.s143096
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发表时间：
2017
影响因子：
4
通讯作者：
Symons M
中科院分区：
文献类型：
--
作者：
Markowitz D;Ha G;Ruggieri R;Symons M
微管靶向剂（MTA）的细胞毒性作用通常归因于对有丝分裂细胞的靶向作用。在临床实践中，MTAs 与 DNA 损伤剂（例如电离辐射 (IR)）结合使用，理由是有丝分裂细胞对 DNA 损伤高度敏感。相比之下，最近的研究表明，MTA 通过干扰间期 DNA 损伤反应 (DDR) 蛋白的运输来与 IR 发挥协同作用。然而，这些研究尚未证明在 IR 存在的情况下干扰间期微管的功能后果。为了解决这个问题，我们将 IR 与已建立的 MTA 甲苯咪唑 (MBZ) 相结合，专门在间期治疗神经胶质瘤细胞。为了测试 MTA 是否可以使间期细胞对 IR 敏感，我们在 IR 后 3-9 小时内（当有丝分裂指数为 0% 时）用 MBZ 处理 GL261 和 GBM14 神经胶质瘤细胞。使用 WST-1 测定法测量细胞活力，并使用剂量增强因子 (DEF) 量化放射增敏。通过 H2AX 磷酸化的蛋白质印迹分析研究了 MBZ 对 DDR 的影响。为了检查 MTA 对 DDR 蛋白、Nbs1 和 Chk2 细胞内转运的影响，在用 MBZ 处理神经胶质瘤细胞后进行了细胞质和核分级分离研究。 MBZ 处理使间期细胞对 IR 的影响敏感，GL261 细胞中的最大 DEF 为 1.34，GBM14 细胞中的最大 DEF 为 1.69。用 MBZ 处理间期细胞导致 IR 后 γH2AX 水平更加持续，表明 DDR 延迟。将神经胶质瘤细胞暴露于 MBZ 会导致 Chk2 和 Nbs1 在细胞质中呈剂量依赖性隔离。这项研究表明，MBZ 可以使癌细胞对 IR 敏感，而与有丝分裂停滞的诱导无关。此外，还提供了证据支持以下假设：MTA 诱导的放射增敏是通过抑制 DDR 蛋白积累到细胞核中介导的。
The cytotoxic effects of microtubule-targeting agents (MTAs) are often attributed to targeted effects on mitotic cells. In clinical practice, MTAs are combined with DNA-damaging agents such as ionizing radiation (IR) with the rationale that mitotic cells are highly sensitive to DNA damage. In contrast, recent studies suggest that MTAs synergize with IR by interfering with the trafficking of DNA damage response (DDR) proteins during interphase. These studies, however, have yet to demonstrate the functional consequences of interfering with interphase microtubules in the presence of IR. To address this, we combined IR with an established MTA, mebendazole (MBZ), to treat glioma cells exclusively during interphase. To test whether MTAs can sensitize interphase cells to IR, we treated GL261 and GBM14 glioma cells with MBZ during 3–9 hours post IR (when the mitotic index was 0%). Cell viability was measured using a WST-1 assay, and radiosensitization was quantified using the dose enhancement factor (DEF). The effect of MBZ on the DDR was studied via Western blot analysis of H2AX phosphorylation. To examine the effects of MTAs on intracellular transport of DDR proteins, Nbs1 and Chk2, cytoplasmic and nuclear fractionation studies were conducted following treatment of glioma cells with MBZ. Treatment with MBZ sensitized interphase cells to the effects of IR, with a maximal DEF of 1.34 in GL261 cells and 1.69 in GBM14 cells. Treatment of interphase cells with MBZ led to more sustained γH2AX levels post IR, indicating a delay in the DDR. Exposure of glioma cells to MBZ resulted in a dose-dependent sequestration of Chk2 and Nbs1 in the cytoplasm. This study demonstrates that MBZ can sensitize cancer cells to IR independently of the induction of mitotic arrest. In addition, evidence is provided supporting the hypothesis that MTA-induced radiosensitization is mediated by inhibiting DDR protein accumulation into the nucleus.