Center for Drug Discovery, Graduate School of Pharmaceutical Sciences (J.-i.S., H.I., K.M., A.A.) and Laboratory of Organic Chemistry, School of Pharmaceutical Sciences (M.S., Y.S.), University of Shizuoka, Shizuoka, Japan.
Center for Drug Discovery, Graduate School of Pharmaceutical Sciences (J.-i.S., H.I., K.M., A.A.) and Laboratory of Organic Chemistry, School of Pharmaceutical Sciences (M.S., Y.S.), University of Shizuoka, Shizuoka, Japan
Mol Pharmacol. 2019 Nov;96(5):609-618. doi: 10.1124/mol.119.116624. Epub 2019 Aug 30.
In the research field of tubulin-binding agents for the development of anticancer agents, hidden targets are emerging as a problem in understanding the exact mechanisms of actions. The quinazoline derivative 1-(4-methoxyphenyl)-1-(quinazolin-4-yl)ethan-1-ol (PVHD121) has anti-cell proliferative activity and inhibits tubulin polymerization by binding to the colchicine site of tubulin. However, the molecular mechanism of action of PVHD121 in cells remains unclear. Here, we demonstrate that PVHD121 delays mitotic entry and efficiently causes mitotic arrest with spindle checkpoint activation, leading to subsequent cell death. The dominant phenotype induced by PVHD121 was aberrant spindles with robust microtubules and unseparated centrosomes. The microtubules were radially distributed, and their ends appeared to adhere to kinetochores, and not to centrosomes. Extensive inhibition by high concentrations of PVHD121 eliminated all microtubules from cells. PVHD277 [1-(4-methoxyphenyl)-1-(2-morpholinoquinazolin-4-yl)ethan-1-ol], a PVHD121 derivative with fluorescence, tended to localize close to the centrosomes when cells prepared to enter mitosis. Our results show that PVHD121 is an antimitotic agent that selectively disturbs microtubule formation at centrosomes during mitosis. This antimitotic activity can be attributed to the targeting of centrosome maturation in addition to the interference with microtubule dynamics. Due to its unique bioactivity, PVHD121 is a potential tool for studying the molecular biology of mitosis and a potential lead compound for the development of anticancer agents. SIGNIFICANCE STATEMENT: Many tubulin-binding agents have been developed as potential anticancer agents. The aim of this study was to understand the subcellular molecular actions of a quinazoline derivative tubulin-binding agent, 1-(4-methoxyphenyl)-1-(quinazolin-4-yl)ethan-1-ol (PVHD121). As expected from its binding activity to tubulin, PVHD121 caused aberrant spindles and inhibited mitotic progression. However, in addition to tubulin, PVHD121 also targeted an unexpected biomolecule involved in centrosome maturation. Due to targeting the biomolecule just before entering mitosis, PVHD121 preferentially inhibited centrosome-derived microtubules rather than chromosome-derived microtubules during spindle formation. This study not only revealed the molecular action of PVHD121 in cells but also emphasized the importance of considering possible tubulin-independent effects of tubulin-binding agents via hidden targeted biomolecules for future use.
在开发抗癌药物的微管结合剂的研究领域中，隐藏靶点作为理解确切作用机制的一个问题而出现。喹唑啉衍生物 1-(4-甲氧基苯基)-1-(喹唑啉-4-基)乙醇（PVHD121）具有抗细胞增殖活性，并通过结合微管的秋水仙碱结合位点抑制微管聚合。然而，PVHD121 在细胞中的分子作用机制仍不清楚。在这里，我们证明 PVHD121 延迟有丝分裂进入，并通过纺锤体检验点激活有效地导致有丝分裂停滞，从而导致随后的细胞死亡。由 PVHD121 诱导的主要表型是具有强壮微管和未分离的中心体的异常纺锤体。微管呈放射状分布，其末端似乎与动粒结合，而不是与中心体结合。高浓度的 PVHD121 广泛抑制会从细胞中消除所有微管。PVHD277[1-(4-甲氧基苯基)-1-(2-吗啉代喹唑啉-4-基)乙醇]是 PVHD121 的衍生物，具有荧光，当细胞准备进入有丝分裂时，它倾向于定位在中心体附近。我们的结果表明，PVHD121 是一种抗有丝分裂剂，它在有丝分裂期间选择性地扰乱中心体处的微管形成。这种抗有丝分裂活性可以归因于除了干扰微管动力学之外，还靶向中心体成熟。由于其独特的生物活性，PVHD121 是研究有丝分裂分子生物学的潜在工具，也是开发抗癌药物的潜在先导化合物。
许多微管结合剂已被开发为有潜力的抗癌药物。本研究的目的是了解喹唑啉衍生物微管结合剂 1-(4-甲氧基苯基)-1-(喹唑啉-4-基)乙醇（PVHD121）的亚细胞分子作用。正如其与微管的结合活性所预期的那样，PVHD121 导致异常纺锤体并抑制有丝分裂进程。然而，除了微管之外，PVHD121 还针对参与中心体成熟的意想不到的生物分子。由于在进入有丝分裂之前靶向该生物分子，因此在纺锤体形成过程中，PVHD121 优先抑制中心体衍生的微管，而不是染色体衍生的微管。这项研究不仅揭示了 PVHD121 在细胞中的分子作用，还强调了在考虑未来使用的潜在微管结合剂的隐藏靶向生物分子时，考虑可能的微管非依赖性效应的重要性。
