Yang Xuemei, Chen Weiguang, Sun Haitao, Chen Weicong, Xu Wei, He Chunyu, Liu Yang, Kuang Ying, Ma Yanhao, Zhong Binglian, Li Chaojie, Li Guohuan, Du Qingfeng, He Songqi
Affiliated Zhuhai Hospital, Southern Medical University (Zhuhai Hospital of Integrated Traditional Chinese and Western Medicine), Zhuhai, 519000, China.
School of Traditional Chinese Medicine, Southern Medical University, Guangzhou, 510515, China.
Chin Med. 2025 Aug 13;20(1):126. doi: 10.1186/s13020-025-01146-6.
Programmed death-ligand 1 (PD-L1), a prominent immune checkpoint, interacts with programmed death protein-1 (PD-1) on cytotoxic T cells within tumors and promotes immune evasion. Emodin, which is known to destabilize PD-L1 in breast cancer, has great potential for enhancing anti-tumor immunity. However, whether emodin can modulate PD-L1 levels in hepatocellular carcinoma (HCC) and enhance anti-tumor immune response remains unclear.
PD-L1 levels were assessed by western blot and RT-qPCR, the degradation mechanism was analyzed using specific inhibitors. Network pharmacology, molecular docking, and glycogen synthase kinase-3 beta (GSK-3β) modulation analyzes were performed to validate emodin's target. In vivo anti-tumor effects were evaluated in H subcutaneous tumor model, and CD8 T cells and RNA-seq data were analyzed. The synergistic effects of emodin and an anti-PD-L1 antibody were assessed.
Emodin effectively reduced PD-L1 levels in H cells and increased anti-tumor activity in an H subcutaneous tumor model by promoting CD8 T cells infiltration and TNF-α, IFN-γ, and granzyme B secretion. Mechanistically, emodin accelerated PD-L1 degradation through the proteasome pathway in both mouse and human HCC cell lines, as confirmed by the use of proteasome, lysosome and autophagy inhibitors. Network pharmacology analysis and molecular docking revealed that GSK-3β, a key regulator of PD-L1 degradation, is a target of emodin. Selective inhibitor-mediated suppression of GSK-3β largely reversed the regulatory effect of emodin on PD-L1. In contrast, overexpression of GSK-3β with a plasmid decreased PD-L1 protein levels and augmented emodin's effect on PD-L1. Additionally, RNA-sequencing revealed the role of emodin in improving the immune responses in the tumor microenvironment. Finally, we observed a synergistic effect when the H cell subcutaneous tumor model was treated with emodin and anti-PD-L1 antibody.
Emodin exerts anti-tumor effects by promoting GSK-3β-mediated PD-L1 proteasomal degradation and enhancing the anti-tumor effects of CD8 T cells, indicating that emodin may be a promising therapeutic option for HCC.
程序性死亡配体1（PD-L1）是一种重要的免疫检查点，它与肿瘤内细胞毒性T细胞上的程序性死亡蛋白1（PD-1）相互作用，促进免疫逃逸。已知大黄素可使乳腺癌中的PD-L1不稳定，在增强抗肿瘤免疫方面具有巨大潜力。然而，大黄素是否能调节肝细胞癌（HCC）中的PD-L1水平并增强抗肿瘤免疫反应仍不清楚。
通过蛋白质免疫印迹法和逆转录定量聚合酶链反应（RT-qPCR）评估PD-L1水平，使用特异性抑制剂分析降解机制。进行网络药理学、分子对接和糖原合酶激酶-3β（GSK-3β）调节分析以验证大黄素的靶点。在H皮下肿瘤模型中评估体内抗肿瘤作用，并分析CD8 T细胞和RNA测序数据。评估大黄素与抗PD-L1抗体的协同作用。
大黄素有效降低H细胞中的PD-L1水平，并通过促进CD8 T细胞浸润以及肿瘤坏死因子-α（TNF-α）、干扰素-γ（IFN-γ）和颗粒酶B的分泌，增强H皮下肿瘤模型中的抗肿瘤活性。机制上，大黄素通过蛋白酶体途径加速小鼠和人肝癌细胞系中PD-L1的降解，蛋白酶体、溶酶体和自噬抑制剂的使用证实了这一点。网络药理学分析和分子对接显示，PD-L1降解的关键调节因子GSK-3β是大黄素的靶点。选择性抑制剂介导的GSK-3β抑制在很大程度上逆转了大黄素对PD-L1的调节作用。相反，用质粒过表达GSK-3β可降低PD-L1蛋白水平并增强大黄素对PD-L1的作用。此外，RNA测序揭示了大黄素在改善肿瘤微环境中免疫反应方面的作用。最后，我们观察到在用大黄素和抗PD-L1抗体治疗H细胞皮下肿瘤模型时具有协同作用。
大黄素通过促进GSK-3β介导的PD-L1蛋白酶体降解和增强CD8 T细胞的抗肿瘤作用发挥抗肿瘤作用，表明大黄素可能是HCC的一种有前景的治疗选择。
