Department of Colorectal Surgery, the First Affiliated Hospital of Fujian Medical University, Fuzhou, 350001, China; Department of Colorectal Surgery, National Regional Medical Center, Binhai Campus of the First Affiliated Hospital, Fujian Medical University, Fuzhou, 350001, China; Fujian Abdominal Surgery Research Institute, the First Affiliated Hospital, Fujian Medical University, Fuzhou, 350001, China.
Department of Gastrointestinal Surgery, Fujian Provincial Hospital, Fuzhou, 350001, China.
Chem Biol Interact. 2024 Nov 1;403:111239. doi: 10.1016/j.cbi.2024.111239. Epub 2024 Sep 19.
The microsatellite stable (MSS) colon cancer (CC) has long been considered resistant to immunotherapy. Cuproptosis, as a novel form of cell death, may interact with tumor immunity. This project focused on the impact of cuproptosis on the cytotoxicity of CD8T in MSS CC, aiming to provide effective clues for improving the treatment strategy of MSS CC. The study developed an MSS CC cuproptosis model using 50 nM elesclomol and 1 μM CuCl. Cuproptotic SW480 cells were directly co-cultured with CD8 T cells. Cuproptosis levels were assessed via intracellular copper ion detection, Western blot, and confocal laser scanning microscopy. CCK-8, Hochest/PI staining, CFSE cell proliferation assay, LDH cytotoxicity detection, and ELISA were used to evaluate CD8 T cell immune activity and cytotoxicity. Transcriptome sequencing and bioinformatics analysis identified regulated signals in cuproptotic SW480 cells. A rescue experiment utilized a WNT pathway activator (BML-284). PD-L1 expression in cells/membranes was analyzed using qRT-PCR, Western blot, and flow cytometry. NSG mice were immunoreconstituted, and the effects of cuproptosis on immune infiltration and cancer progression in MSS CC mice were assessed using ELISA and immunohistochemistry (IHC). Treatment with 50 nM elesclomol and 1 μM CuCl significantly increased cuproptosis in SW480 cells. Co-culture with CD8 T cells enhanced their cytotoxicity. Sequencing revealed cuproptosis-mediated modulation of immune and inflammatory pathways, including WNT signaling. Rescue experiments showed downregulation of WNT signaling in cuproptotic SW480 cells. Indirectly, CD8 T cell immune function was enhanced by reducing PD-L1 expression. In mice, cuproptosis resulted in increased infiltration of CD8 T cells in tumor tissue, leading to delayed cancer progression compared to the control group. Cuproptosis in MSS CC cells enhances the cytotoxicity of CD8 T cells, which may be achieved through downregulation of the WNT signaling pathway and decreased expression of PD-L1. In the future, drugs that can induce cuproptosis may be a promising approach to improve MSS CC immunotherapy.
微卫星稳定（MSS）结肠癌（CC）长期以来被认为对免疫疗法具有抗性。铜死亡作为一种新型的细胞死亡形式，可能与肿瘤免疫相互作用。本项目重点研究铜死亡对 MSS CC 中 CD8T 细胞细胞毒性的影响，旨在为提高 MSS CC 的治疗策略提供有效线索。该研究使用 50 nM elesclomol 和 1 μM CuCl 开发了 MSS CC 铜死亡模型。将铜死亡的 SW480 细胞直接与 CD8T 细胞共培养。通过细胞内铜离子检测、Western blot 和共聚焦激光扫描显微镜评估铜死亡水平。使用 CCK-8、Hochest/PI 染色、CFSE 细胞增殖测定、LDH 细胞毒性检测和 ELISA 评估 CD8T 细胞免疫活性和细胞毒性。转录组测序和生物信息学分析鉴定了铜死亡的 SW480 细胞中受调控的信号。利用 WNT 途径激活剂（BML-284）进行挽救实验。使用 qRT-PCR、Western blot 和流式细胞术分析细胞/膜中的 PD-L1 表达。免疫重建 NSG 小鼠，使用 ELISA 和免疫组织化学（IHC）评估铜死亡对 MSS CC 小鼠免疫浸润和癌症进展的影响。使用 50 nM elesclomol 和 1 μM CuCl 处理显着增加了 SW480 细胞中的铜死亡。与 CD8T 细胞共培养增强了它们的细胞毒性。测序显示铜死亡介导的免疫和炎症途径的调节，包括 WNT 信号。挽救实验表明铜死亡的 SW480 细胞中 WNT 信号下调。间接地，通过降低 PD-L1 的表达增强了 CD8T 细胞的免疫功能。在小鼠中，与对照组相比，铜死亡导致肿瘤组织中 CD8T 细胞浸润增加，从而延缓癌症进展。MSS CC 细胞中的铜死亡增强了 CD8T 细胞的细胞毒性，这可能是通过下调 WNT 信号通路和降低 PD-L1 表达来实现的。在未来，能够诱导铜死亡的药物可能是改善 MSS CC 免疫疗法的有前途的方法。
