Background: Given the diverse roles of cyclin A2 both in cell cycle regulation and in DNA damage response, identifying small molecule regulators of cyclin A2 activity carries significant potential to regulate diverse cellular processes in both ageing/neurodegeneration and in cancer.

Objective: Based on cyclin A2’s recently discovered role in DNA repair, we hypothesized that small molecule inhibitors that were predicted to bind to both cyclin A2 and CDK2 will be useful as a radiosensitizer of cancer cells. In this study, we used structure-based drug discovery to find inhibitors that target both cyclin A2 and CDK2.

Methods: Molecular dynamics simulations were used to generate diverse binding pocket conformations for application of the relaxed complex scheme. We then used structure-based virtual screening to find potential dual cyclin A2 and CDK2 inhibitors. Based on a consensus score of docked poses from Glide and AutoDock Vina, we identified about 40 promising hit compounds, where all PAINS scaffolds were removed from consideration. A biochemical luminescence assay of cyclin A2-CDK2 function was used for experimental verification.

Results: Four lead inhibitors of cyclin A2-CDK2 complex have been identified using a relaxed complex scheme virtual screen have been verified in a biochemical luminescence assay of cyclin A2- CDK2 function. Two of the four lead inhibitors had inhibitory concentrations in the nanomolar range.

Conclusion: The four cyclin A2-CDK2 complex inhibitors are the first reported inhibitors that were specifically designed not to target the cyclin A2-CDK2 protein-protein interface. Overall, our results highlight the potential of combined advanced computational tools and biochemical verification to discover novel binding scaffolds.

背景：鉴于细胞周期蛋白 A2 在细胞周期调节和 DNA 损伤反应中的多种作用，鉴定细胞周期蛋白 A2 活性的小分子调节因子具有调节衰老/神经变性和癌症中多种细胞过程的巨大潜力。

目的：基于最近发现的细胞周期蛋白 A2 在 DNA 修复中的作用，我们假设预计可与细胞周期蛋白 A2 和 CDK2 结合的小分子抑制剂可用作癌细胞的放射增敏剂。在这项研究中，我们使用基于结构的药物发现来寻找同时靶向细胞周期蛋白 A2 和 CDK2 的抑制剂。

方法：分子动力学模拟用于生成不同的结合口袋构象，以应用松弛复合方案。然后我们使用基于结构的虚拟筛选来寻找潜在的双周期蛋白 A2 和 CDK2 抑制剂。根据 Glide 和 AutoDock Vina 对停靠姿势的一致评分，我们确定了大约 40 种有希望的命中化合物，其中所有 PAINS 支架都被排除在外。细胞周期蛋白 A2-CDK2 功能的生化发光分析用于实验验证。

结果：已使用松弛复合方案虚拟筛选鉴定了四种细胞周期蛋白 A2-CDK2 复合物的先导抑制剂，已在细胞周期蛋白 A2-CDK2 功能的生化发光分析中得到验证。四种先导抑制剂中的两种具有纳摩尔范围内的抑制浓度。

结论：四种细胞周期蛋白 A2-CDK2 复合物抑制剂是首次报道的专门设计不针对细胞周期蛋白 A2-CDK2 蛋白-蛋白界面的抑制剂。总体而言，我们的结果突出了结合先进计算工具和生化验证的潜力，以发现新型结合支架。