School of Pharmacy, College of Medicine, National Taiwan University, Taipei, Taiwan.
Department of Pharmacy, New Taipei Municipal TuCheng Hospital, Chang Gung Memorial Hospital, New Taipei City, Taiwan.
Prostate. 2023 Dec;83(16):1549-1563. doi: 10.1002/pros.24613. Epub 2023 Aug 15.
Castration-resistant prostate cancer (CRPC) is refractory to hormone treatment and the therapeutic options are continuously advancing. This study aims to discover the anti-CRPC effects and underlying mechanisms of small-molecule compounds targeting topoisomerase (TOP) II and cellular components of DNA damage repair.
Cell proliferation was determined in CRPC PC-3 and DU-145 cells using anchorage-dependent colony formation, sulforhodamine B assay and flow cytometric analysis of CFSE staining. Flow cytometric analyses of propidium iodide staining and JC-1 staining were used to examine the population of cell-cycle phases and mitochondrial membrane potential, respectively. Nuclear extraction was performed to detect the nuclear localization of cellular components in DNA repair pathways. Protein expressions were determined using Western blot analysis.
A series of azathioxanthone-based derivatives were synthesized and examined for bioactivities in which WC-A13, WC-A14, WC-A15, and WC-A16 displayed potent anti-CRPC activities in both PC-3 and DU-145 cell models. These WC-A compounds selectively downregulated both TOP IIα and TOP IIβ but not TOP I protein expression. WC-A13, WC-A14, and WC-A15 were more potent than WC-A16 on TOP II inhibition, mitochondrial dysfunction, and induction of caspase cascades indicating the key role of amine-containing side chain of the compounds in determining anti-CRPC activities. Furthermore, WC-A compounds induced an increase of γH2AX and activated ATR-Chk1 and ATM-Chk2 signaling pathways. P21 protein expression was also upregulated by WC-A compounds in which WC-A16 showed the least activity. Notably, WC-A compounds exhibited different regulation on Rad51, a major protein in homologous recombination of DNA in double-stranded break repair. WC-A13, WC-A14, and WC-A15 inhibited, whereas WC-A16 induced, the nuclear translocation of Rad51.
The data suggest that WC-A compounds exhibit anti-CRPC effects through the inhibition of TOP II activities, leading to mitochondrial stress-involved caspase activation and apoptosis. Moreover, WC-A13, WC-A14, and WC-A15 but not WC-A16 display inhibitory activities of Rad51-mediated DNA repair pathway which may increase apoptotic effect of CRPC cells.
去势抵抗性前列腺癌（CRPC）对激素治疗有抗药性，治疗选择在不断推进。本研究旨在发现针对拓扑异构酶（TOP）II 和 DNA 损伤修复细胞成分的小分子化合物的抗 CRPC 作用及其潜在机制。
用依赖于附着的集落形成、磺酰罗丹明 B 测定和 CFSE 染色的流式细胞术分析来确定 CRPC PC-3 和 DU-145 细胞中的细胞增殖。用碘化丙啶染色和 JC-1 染色的流式细胞术分析分别检测细胞周期各期的细胞群体和线粒体膜电位。进行核提取以检测 DNA 修复途径中细胞成分的核定位。使用 Western blot 分析来确定蛋白表达。
合成了一系列氮杂噻唑酮类衍生物，并对其生物活性进行了检测，其中 WC-A13、WC-A14、WC-A15 和 WC-A16 在 PC-3 和 DU-145 细胞模型中均显示出有效的抗 CRPC 活性。这些 WC-A 化合物选择性地下调了 TOP IIα 和 TOP IIβ 但不调 TOP I 蛋白的表达。在 TOP II 抑制、线粒体功能障碍和半胱天冬酶级联诱导方面，WC-A13、WC-A14 和 WC-A15 比 WC-A16 更有效，这表明化合物中含胺侧链在决定抗 CRPC 活性方面起着关键作用。此外，WC-A 化合物诱导 γH2AX 的增加，并激活了 ATR-Chk1 和 ATM-Chk2 信号通路。WC-A 化合物还上调了 P21 蛋白的表达，其中 WC-A16 的活性最低。值得注意的是，WC-A 化合物对 Rad51 表现出不同的调节作用，Rad51 是双链断裂修复中同源重组的主要蛋白。WC-A13、WC-A14 和 WC-A15 抑制，而 WC-A16 诱导 Rad51 的核易位。
数据表明，WC-A 化合物通过抑制 TOP II 活性来发挥抗 CRPC 作用，导致涉及半胱天冬酶激活和凋亡的线粒体应激。此外，WC-A13、WC-A14 和 WC-A15 但不是 WC-A16 显示出抑制 Rad51 介导的 DNA 修复途径的活性，这可能会增加 CRPC 细胞的凋亡效应。
