前列腺癌（PCa）是全世界男性死亡的原因之一。尽管已经制定了治疗策略，但疾病的复发和随之而来的副作用仍然是一个重要问题。 Diospyros rhodocalyx Kurz 是一种传统的泰国药物，具有多种治疗特性，包括抗癌活性。然而，其针对前列腺癌的抗癌活性尚未得到彻底研究。本研究旨在评估 D. rhodocalyx Kurz 的乙酸乙酯提取物 (EADR) 的抗癌活性和与 LNCaP 人前列腺癌细胞系凋亡诱导相关的潜在机制。红花萼库尔兹。使用 MTS 测定评估 EADR 对 LNCaP 和 WPMY-1 细胞（正常人​​前列腺肌成纤维细胞系）的细胞毒性。分别通过碘化丙啶 (PI)、膜联蛋白 V-FITC/PI 和 JC-1 染料染色评估 EADR 对细胞周期、细胞凋亡诱导和线粒体膜电位 (MMP) 改变的影响。使用流式细胞术进行后续分析。通过蛋白质印迹法检查裂解的 caspase-3、BAX 和 Bcl-2 的表达。使用气相色谱-质谱法（GC-MS）对 EADR 进行植物化学分析。EADR 对 LNCaP 细胞表现出剂量依赖性细胞毒作用，24 小时和 48 小时后的 IC50 值分别为 15.43 和 12.35 µg/mL。 。虽然它对WPMY-1细胞也表现出细胞毒作用，但作用相对较低，暴露24和48小时后的IC50值分别为34.61和19.93 µg/mL。细胞周期分析表明，EADR 不会诱导 LNCaP 或 WPMY-1 细胞的细胞周期停滞。然而，它显着增加了 LNCaP 细胞中的亚 G1 群体，表明可能诱导细胞凋亡。 Annexin V-FITC/PI染色表明EADR显着诱导LNCaP细胞凋亡。随后对 EADR 诱导细胞凋亡的潜在机制的研究揭示了 MMP 的减少，如 JC-1 染色所证明的那样。此外，蛋白质印迹表明，EADR 处理导致 LNCaP 细胞中 BAX 上调、BCL-2 下调以及 caspase-3 裂解升高。值得注意的是，经 GC-MS 鉴定，表羽酚是 EADR 中的一种重要化合物。EADR 通过诱导细胞毒性和细胞凋亡，对 LNCaP 人前列腺癌细胞系表现出抗癌活性。我们的研究结果表明，EADR 通过上调促凋亡 BAX 促进细胞凋亡，而下调抗凋亡 Bcl-2 则导致 MMP 减少和 caspase-3 激活。特别令人感兴趣的是 Epilupeol 的存在，这是 EADR 中发现的一种主要化合物，它可能有望成为开发前列腺癌治疗药物的候选药物。© 2024 Phongsuwichetsak 等人。

Prostate cancer (PCa) is one of the causes of death in men worldwide. Although treatment strategies have been developed, the recurrence of the disease and consequential side effects remain an essential concern. Diospyros rhodocalyx Kurz, a traditional Thai medicine, exhibits diverse therapeutic properties, including anti-cancer activity. However, its anti-cancer activity against prostate cancer has not been thoroughly explored. This study aims to evaluate the anti-cancer activity and underlying mechanisms of the ethyl acetate extract of D. rhodocalyx Kurz (EADR) related to apoptosis induction in the LNCaP human prostate cancer cell line.Ethyl acetate was employed to extract the dried bark of D. rhodocalyx Kurz. The cytotoxicity of EADR on both LNCaP and WPMY-1 cells (normal human prostatic myofibroblast cell line) was evaluated using MTS assay. The effect of EADR on the cell cycle, apoptosis induction, and alteration in mitochondrial membrane potential (MMP) was assessed by the staining with propidium iodide (PI), Annexin V-FITC/PI, and JC-1 dye, respectively. Subsequent analysis was conducted using flow cytometry. The expression of cleaved caspase-3, BAX, and Bcl-2 was examined by Western blotting. The phytochemical profiling of the EADR was performed using gas chromatography-mass spectrometry (GC-MS).EADR exhibited a dose-dependent manner cytotoxic effect on LNCaP cells, with IC50 values of 15.43 and 12.35 µg/mL after 24 and 48 h, respectively. Although it also exhibited a cytotoxic effect on WPMY-1 cells, the effect was comparatively lower, with the IC50 values of 34.61 and 19.93 µg/mL after 24 and 48 h of exposure, respectively. Cell cycle analysis demonstrated that EADR did not induce cell cycle arrest in either LNCaP or WPMY-1 cells. However, it significantly increased the sub-G1 population in LNCaP cells, indicating a potential induction of apoptosis. The Annexin V-FITC/PI staining indicated that EADR significantly induced apoptosis in LNCaP cells. Subsequent investigation into the underlying mechanism of EADR-induced apoptosis revealed a reduction in MMP as evidenced by JC-1 staining. Moreover, Western blotting demonstrated that EADR treatment resulted in the upregulation of BAX, downregulation of BCL-2, and elevation of caspase-3 cleavage in LNCaP cells. Notably, the epilupeol was a prominent compound in EADR as identified by GC-MS.The EADR exhibits anti-cancer activity against the LNCaP human prostate cancer cell line by inducing cytotoxicity and apoptosis. Our findings suggest that EADR promotes apoptosis by upregulating pro-apoptotic BAX, whereas downregulation of anti-apoptotic Bcl-2 results in the reduction of MMP and the activation of caspase-3. Of particular interest is the presence of epilupeol, a major compound identified in EADR, which may hold promise as a candidate for the development of therapeutic agents for prostate cancer.© 2024 Phongsuwichetsak et al.