Laboratory of Signaling in Biomolecular Systems, Department of Biochemistry, Institute of Chemistry, University of São Paulo, São Paulo, Brazil.
Laboratory of Signaling in Biomolecular Systems, Department of Biochemistry, Institute of Chemistry, University of São Paulo, São Paulo, Brazil.
Int J Biochem Cell Biol. 2023 Nov;164:106474. doi: 10.1016/j.biocel.2023.106474. Epub 2023 Sep 30.
Resistance to radio and chemotherapy in Glioblastoma (GBM) is correlated with its malignancy, invasiveness, and aggressiveness. The Rho GTPase pathway plays important roles in these processes, but its involvement in the GBM response to genotoxic treatments remains unsolved. Inhibition of this signaling pathway has emerged as a promising approach for the treatment of CNS injuries and diseases, proving to be a strong candidate for therapeutic approaches. To this end, Rho-associated kinases (ROCK), classic downstream effectors of small Rho GTPases, were targeted for pharmacological inhibition using Y-27632 in GBM cells, expressing the wild-type or mutated p53 gene, and exposed to genotoxic stress by gamma ionizing radiation (IR) or cisplatin (PT). The use of the ROCK inhibitor (ROCKi) had opposite effects in these cells: in cells expressing wild-type p53, ROCKi reduced survival and DNA repair capacity (reduction of γH2AX foci and accumulation of strand breaks) after stress promoted by IR or PT; in cells expressing the mutant p53 protein, both treatments promoted longer survival and more efficient DNA repair, responses further enhanced by ROCKi. The target DNA repair mechanisms of ROCK inhibition were, respectively, an attenuation of NHEJ and NER pathways in wild-type p53 cells, and a stimulation of HR and NER pathways in mutant p53 cells. These effects were accompanied by the formation of reactive oxygen species (ROS) induced by genotoxic stress only in mutant p53 cells but potentiated by ROCKi and reversed by p53 knockdown. N-acetyl-L-cysteine (NAC) treatment or Rac1 knockdown completely eliminated ROCKi's p53-dependent actions, since ROCK inhibition specifically elevated Rac-GTP levels only in mutant p53 cells. Combining IR or PT and ROCKi treatments broadens our understanding of the sensitivity and resistance of, respectively, GBM expressing wild-type or mutant p53 to genotoxic agents. Our proposal may be a determining factor in improving the efficiency and assertiveness of CNS antitumor therapies based on ROCK inhibitors. SIGNIFICANCE: The use of ROCK inhibitors in association with radio or chemotherapy modulates GBM resistance and sensitivity depending on the p53 activity, suggesting the potential value of this protein as therapeutic target for tumor pre-sensitization strategies.
胶质母细胞瘤（GBM）的放化疗耐药性与其恶性程度、侵袭性和侵袭性密切相关。Rho GTPase 通路在这些过程中发挥着重要作用，但它在 GBM 对遗传毒性治疗的反应中的作用仍未解决。抑制该信号通路已成为治疗中枢神经系统损伤和疾病的一种有前途的方法，被证明是治疗方法的一个强有力的候选者。为此，使用 Y-27632 靶向 Rho 相关激酶（ROCK），即小 Rho GTPase 的经典下游效应物，对表达野生型或突变型 p53 基因的 GBM 细胞进行药理学抑制，并通过γ离子辐射（IR）或顺铂（PT）暴露于遗传毒性应激下。在这些细胞中，ROCK 抑制剂（ROCKi）的使用有相反的效果：在表达野生型 p53 的细胞中，ROCKi 减少了应激后 IR 或 PT 促进的存活和 DNA 修复能力（减少 γH2AX 焦点和积累链断裂）；在表达突变型 p53 蛋白的细胞中，两种治疗方法均促进了更长的存活时间和更有效的 DNA 修复，ROCKi 进一步增强了这些反应。ROCK 抑制的靶向 DNA 修复机制分别是在野生型 p53 细胞中减弱 NHEJ 和 NER 途径，以及在突变型 p53 细胞中刺激 HR 和 NER 途径。这些效应伴随着仅在突变型 p53 细胞中由遗传毒性应激诱导的活性氧（ROS）的形成，但由 ROCKi 增强并由 p53 敲低逆转。N-乙酰-L-半胱氨酸（NAC）处理或 Rac1 敲低完全消除了 ROCKi 的 p53 依赖性作用，因为 ROCK 抑制仅在突变型 p53 细胞中特异性地提高 Rac-GTP 水平。联合使用 IR 或 PT 和 ROCKi 治疗拓宽了我们对分别表达野生型或突变型 p53 的 GBM 对遗传毒性药物的敏感性和耐药性的理解。我们的建议可能是提高基于 ROCK 抑制剂的中枢神经系统抗肿瘤治疗效率和果断性的决定因素。意义：ROCK 抑制剂与放疗或化疗联合使用可调节 GBM 的耐药性和敏感性，这取决于 p53 的活性，这表明该蛋白作为肿瘤预致敏策略的治疗靶点具有潜在价值。
