结直肠癌 (CRC) 是癌症死亡的第二大原因，约 10% 的恶性肿瘤是结直肠癌。癌症干细胞被认为是结直肠癌治疗耐药和疾病复发的罪魁祸首。本研究探讨了含有 24 (TRIM24) 和 X 连锁锌指蛋白 (ZFX) 的三联基序对 CRC 细胞干性和 5-FU 耐药性的影响。构建了 5-FU 耐药细胞系 (HT29-5-FU)，用于 CRC 5-FU 耐药细胞的功能分析。采用 qRT-PCR 和蛋白质印迹 (WB) 分析 5-FU 耐药细胞和敏感细胞中 ZFX 的 mRNA 和蛋白水平。 WB还用于分析各组干细胞的表面标志物。 CCK-8测定测定5-FU处理的不同细胞组的IC50值。使用肿瘤球测定法测定各组细胞的成球能力。双荧光素酶报告基因测定验证了 ZFX 与 TRIM24 的结合。 ZFX 在 HT29-5-FU 细胞中高表达。沉默ZFX显着降低HT29-5-FU细胞的5-FU耐药性和IC50值，干细胞的表面标志物和细胞球形成能力也显着降低。当ZFX过表达时，HT29细胞的功能相反。在CRC细胞中，TRIM24是ZFX的上游转录因子，并且它们相互作用。 TRIM24激活ZFX的表达来影响细胞的干性和5-FU耐药性。 TRIM24/ZFX 调节轴影响 CRC 细胞的干性及其对 5-FU 的敏感性，为 CRC 的新治疗途径提供潜在的药物靶点。

Colorectal cancer (CRC) is the second leading cause of cancer death, and about 10% of all malignancies are CRC. Cancer stem cells are considered main culprits in CRC treatment resistance and disease recurrence. This study explored the effects of tripartite motif containing 24 (TRIM24) and zinc finger protein, X-linked (ZFX) on CRC cell stemness and 5-FU resistance. A 5-FU-resistant cell line (HT29-5-FU) was constructed for functional analysis of CRC 5-FU-resistant cells. qRT-PCR and western blot (WB) were employed to analyze mRNA and protein levels of ZFX in 5-FU resistant cells and sensitive cells. WB was also utilized to analyze the surface markers of stem cells in each group. CCK-8 assay determined the IC50 values of different cell groups treated with 5-FU. The sphere-forming ability of cells in each group was determined using tumor sphere assay. Dual-luciferase reporter gene assay validated binding of ZFX to TRIM24. ZFX was highly expressed in HT29-5-FU cells. Silencing ZFX significantly reduced the 5-FU resistance and IC50 value of HT29-5-FU cells, and the surface markers and cell sphere-forming ability of stem cells were also significantly reduced. The function of HT29 cells was opposite when ZFX was overexpressed. In CRC cells, TRIM24 was an upstream transcription factor of ZFX, and they interacted with each other. TRIM24 activated the expression of ZFX to influence the stemness and 5-FU resistance of cells. The TRIM24/ZFX regulatory axis affected the stemness of CRC cells and their sensitivity to 5-FU, providing potential drug targets for novel therapeutic avenues for CRC.