Sun Haohang, Chen Qi, Zhang Xiwei, Chen Mengze, Dai Ji, Yan Meidi
Department of General Surgery, Zhenhai District People's Hospital, No. 718, South 2nd West Road, Camel Street, Zhenhai District, Ningbo City, 315200, Zhejiang, China.
Biochem Genet. 2025 Sep 5. doi: 10.1007/s10528-025-11242-9.
Triple-negative breast cancer (TNBC), a particularly aggressive cancer, significantly menaces women's health. Recently, a novel form of cell death known as cuproptosis has been identified, with the key gene FDX1 emerging as a potential oncogenic factor. We analyzed the heterogeneity of breast cancer (BC) epithelial cells using available single-cell RNA sequencing (scRNA-seq) datasets. We developed knockdown cell lines in vitro and verified the knockdown efficiency with qPCR. The malignant phenotypes of the cells were assessed through cell counting kit-8, colony formation, Transwell, and scratch healing assays. We also co-cultured the cells with CD8 T cells and evaluated their activation using Transwell, CFSE, lactate dehydrogenase release assay, and enzyme-linked immunosorbent assay. IHC analysis was conducted to reveal the impact of FDX1 on tumor growth in mice. Based on scRNA-seq data, we discovered that in TNBC, epithelial cells were more abundant, and T-cell infiltration was less frequent compared to other subtypes of BC. FDX1 + epithelial cells, which are associated with cuproptosis, were highly enriched in TNBC. The expression of FDX1, a key gene in cuproptosis, upregulated in these cells. This upregulation is essential for sustaining the growth, invasion, and migration of TNBC cells. Co-culture experiments revealed that FDX1 expression could modulate the activation and cytotoxicity of T cells. Tumor growth in mice was largely curbed by the knockdown of FDX1 expression. In TNBC, FDX1 expression aids in the survival and proliferation of cancer cells while dampening the immune response of CD8 T cells.
三阴性乳腺癌（TNBC）是一种侵袭性特别强的癌症，严重威胁着女性健康。最近，一种名为铜死亡的新型细胞死亡形式被发现，关键基因FDX1成为一种潜在的致癌因素。我们利用现有的单细胞RNA测序（scRNA-seq）数据集分析了乳腺癌（BC）上皮细胞的异质性。我们在体外构建了基因敲低细胞系，并通过qPCR验证了敲低效率。通过细胞计数试剂盒-8、集落形成、Transwell和划痕愈合试验评估细胞的恶性表型。我们还将细胞与CD8 T细胞共培养，并使用Transwell、CFSE、乳酸脱氢酶释放试验和酶联免疫吸附试验评估它们的活化情况。进行免疫组化分析以揭示FDX1对小鼠肿瘤生长的影响。基于scRNA-seq数据，我们发现，与其他BC亚型相比，TNBC中的上皮细胞更为丰富，T细胞浸润频率更低。与铜死亡相关的FDX1+上皮细胞在TNBC中高度富集。铜死亡关键基因FDX1在这些细胞中的表达上调。这种上调对于维持TNBC细胞的生长、侵袭和迁移至关重要。共培养实验表明，FDX1表达可调节T细胞的活化和细胞毒性。FDX1表达的敲低在很大程度上抑制了小鼠的肿瘤生长。在TNBC中，FDX1表达有助于癌细胞的存活和增殖，同时抑制CD8 T细胞的免疫反应。
