Department of Pharmacy, The First Affiliated Hospital, Sun Yat-sen University, 510080 Guangzhou, China.
Department of Pharmacy, The First Affiliated Hospital, Sun Yat-sen University, 510080 Guangzhou, China.
Phytomedicine. 2021 Jun;86:153563. doi: 10.1016/j.phymed.2021.153563. Epub 2021 Apr 4.
Hepatocellular carcinoma (HCC) is one of the leading causes of cancer-associated mortality worldwide. Sorafenib (SORA), as a first-line therapeutic drug, has been used to treat HCC, but resistance poses a major limitation on the efficacy of SORA chemotherapy. Pristimerin (PRIS), a natural bioactive component isolated from various plant species in the Celastraceae and Hippocrateaceae families, has been reported to exhibit outstanding antitumor effects in several types of cells in vitro.
The aim of this study was to investigate whether PRIS can exert synergistic anti-tumor effects with the combination of SORA, and if so, through what mechanism.
Conditionally reprogrammed patient derived-primary hepatocellular carcinoma cells (CRHCs) were isolated from human liver cancer tissues and treated with SORA and PRIS. Cell proliferation, apoptosis, migration and tube formation ability were detected by DNA content quantification, flow cytometry, transwell assay and Matrigel-based angiogenesis assay. Gene and protein expression were assessed by qRT-PCR and Western blot respectively.
Initially, we observed that the combination of the two drugs had a much stronger inhibitory effect on CRHCs growth than either drug alone. Moreover, the combination of 2 µM SORA and 1 µM PRIS exhibited a significant anti‑migrative and anti-invaded effect on CRHCs, and remarkably inhibited capillary structure formation of Human Umbilical Vein Endothelial Cells (HUVECs). Furthermore, the combined treatment with SORA and PRIS synergistically induced intrinsic apoptosis in CRHCs, involving a caspase-4-dependent mechanism paralleled by an increased Bax/Bcl-xL ratio. These activities were mediated through ROS generation and the induction of endoplasmic reticulum (ER) stress and mitochondrial dysfunction. GRP78 silencing or ER stress inhibitor 4-phenylbutyric acid administration was revealed to abolish the anticancer effects of PRIS, indicating the critical role of GRP78 in mediating the bioactivity of PRIS. The present study also provides mechanistic evidence that PRIS modulated the Akt/FoxO1/p27 signaling pathway, which is required for mitochondrial-mediated intrinsic apoptosis, activation of ER stress, and stimulation of caspase-4 induced by PRIS, and, consequently resulting in suppressed cell viability, migration and angiogenesis co-treated with SORA in CRHCs.
Our results suggest the use of PRIS as sensitizers of chemotherapy paving the way for innovative and promising targeted chemotherapy-based therapeutic strategies in human HCC.
肝细胞癌（HCC）是全球癌症相关死亡的主要原因之一。索拉非尼（SORA）作为一线治疗药物，已被用于治疗 HCC，但耐药性严重限制了 SORA 化疗的疗效。普瑞巴林（PRIS）是从 Celastraceae 和 Hippocrateaceae 科的各种植物中分离得到的天然生物活性成分，已被报道在体外的几种细胞类型中具有出色的抗肿瘤作用。
本研究旨在探讨 PRIS 是否可以与 SORA 联合发挥协同抗肿瘤作用，如果可以，其作用机制是什么。
从人肝癌组织中分离出条件重编程的患者衍生原代肝癌细胞（CRHCs），并用 SORA 和 PRIS 处理。通过 DNA 含量定量、流式细胞术、Transwell 测定和基于 Matrigel 的血管生成测定检测细胞增殖、凋亡、迁移和管形成能力。通过 qRT-PCR 和 Western blot 分别评估基因和蛋白表达。
首先，我们观察到两种药物联合使用对 CRHCs 的生长抑制作用强于单独使用任何一种药物。此外，2μM SORA 和 1μM PRIS 的联合用药对 CRHCs 具有显著的抗迁移和抗侵袭作用，并显著抑制人脐静脉内皮细胞（HUVECs）的毛细血管结构形成。此外，SORA 和 PRIS 的联合治疗协同诱导 CRHCs 发生内在凋亡，涉及 caspase-4 依赖性机制，并伴有 Bax/Bcl-xL 比值增加。这些活性是通过 ROS 生成以及内质网（ER）应激和线粒体功能障碍的诱导介导的。沉默葡萄糖调节蛋白 78（GRP78）或给予 ER 应激抑制剂 4-苯基丁酸可消除 PRIS 的抗癌作用，表明 GRP78 在介导 PRIS 的生物活性中起关键作用。本研究还提供了机制证据，表明 PRIS 调节 Akt/FoxO1/p27 信号通路，该通路对于线粒体介导的内在凋亡、ER 应激的激活以及 PRIS 诱导的 caspase-4 的激活是必需的，从而导致 SORA 共同处理的 CRHCs 细胞活力、迁移和血管生成受到抑制。
我们的研究结果表明，PRIS 可用作化疗增敏剂，为 HCC 的创新和有前景的靶向化疗治疗策略铺平道路。
