Liu Xue, Lin Ye, Zhang Jingzhi
Department of Gastroenterology, The Affiliated Ganzhou Hospital of Nanchang University, Ganzhou City, Jiangxi Province, China.
J Biochem Mol Toxicol. 2025 Sep;39(9):e70472. doi: 10.1002/jbt.70472.
Cuproptosis, a recently characterized form of programmed cell death, has been implicated in tumor progression; however, its specific role in colon cancer remains poorly understood. This study aims to elucidate the potential involvement of cuproptosis-related genes in the development and progression of colon cancer. Differentially expressed genes (DEGs) associated with cuproptosis in colon cancer were identified through bioinformatics analysis of the GSE4183 and GSE74602 datasets. Gain-of-function experiments were performed in HT29 and HCT116 colon cancer cell lines to evaluate their effects on cellular proliferation, migration, and invasion. Functional assays, including JC-1 staining, copper (Cu²⁺) quantification, and lactate/pyruvate measurements, were employed to assess mitochondrial membrane potential and metabolic reprogramming. The involvement of hypoxia-inducible factor-1 alpha (HIF-1α) was further explored through overexpression and rescue assays. To confirm the dependency of the observed effects on cuproptosis, tetrathiomolybdate (TTM) was used as a copper chelator. Additionally, vasoactive intestinal peptide receptor 1 (VIPR1) signaling was activated using its agonist, vasoactive intestinal peptide (VIP). Five downregulated cuproptosis-related hub genes (VIPR1, PYY, NPY, VIP, and SST) were identified as potential diagnostic biomarkers for colon cancer. Among them, VIPR1 overexpression significantly suppressed the proliferation, migration, and invasion of colon cancer cells, accompanied by upregulation of cuproptosis-associated proteins FDX1 and DLST. These effects were markedly attenuated by HIF1A overexpression. The application of the copper chelator TTM abolished the antitumor effects mediated by VIPR1, confirming cuproptosis dependency. Furthermore, treatment with the VIPR1 agonist VIP enhanced VIPR1 signaling, further inhibited malignant cellular behaviors, and downregulated HIF-1α activity. Dual-luciferase reporter assays showed that HIF-1α overexpression reduced the transcriptional activity of wild-type FDX1 and DLST promoters. VIPR1 acts as a tumor suppressor in colon cancer by promoting cuproptosis and disrupting cellular metabolic fitness through inhibition of HIF-1α signaling, thereby representing a promising target for therapeutic intervention.
铜死亡是一种最近被发现的程序性细胞死亡形式，与肿瘤进展有关；然而，其在结肠癌中的具体作用仍知之甚少。本研究旨在阐明铜死亡相关基因在结肠癌发生发展中的潜在作用。通过对GSE4183和GSE74602数据集进行生物信息学分析，确定了与结肠癌铜死亡相关的差异表达基因（DEG）。在HT29和HCT116结肠癌细胞系中进行功能获得实验，以评估其对细胞增殖、迁移和侵袭的影响。采用包括JC-1染色、铜（Cu²⁺）定量和乳酸/丙酮酸测量在内的功能分析方法，评估线粒体膜电位和代谢重编程。通过过表达和挽救实验进一步探究缺氧诱导因子-1α（HIF-1α）的作用。为了证实观察到的效应对铜死亡的依赖性，使用四硫代钼酸盐（TTM）作为铜螯合剂。此外，使用血管活性肠肽受体1（VIPR1）激动剂血管活性肠肽（VIP）激活VIPR1信号通路。鉴定出五个下调的铜死亡相关枢纽基因（VIPR1、PYY、NPY、VIP和SST）作为结肠癌的潜在诊断生物标志物。其中，VIPR1过表达显著抑制结肠癌细胞的增殖、迁移和侵袭，同时铜死亡相关蛋白FDX1和DLST上调。HIF1A过表达显著减弱了这些效应。铜螯合剂TTM的应用消除了VIPR1介导的抗肿瘤作用，证实了对铜死亡的依赖性。此外，用VIPR1激动剂VIP处理增强了VIPR1信号通路，进一步抑制恶性细胞行为，并下调HIF-1α活性。双荧光素酶报告基因实验表明，HIF-1α过表达降低了野生型FDX1和DLST启动子的转录活性。VIPR1通过促进铜死亡和抑制HIF-1α信号通路破坏细胞代谢适应性，在结肠癌中发挥肿瘤抑制作用，因此是一个有前景的治疗干预靶点。
