Hu Yunlong, Tan Xiao, Zhang Lin, Zhu Xiang, Wang Xiangyao
Department of Gastroenterology and Hepatology, The First Medical Center of Chinese PLA General Hospital, Beijing, China.
Department of Emergency and Intensive Care Unit, The 966th Hospital of Joint Logistic Support Force of PLA, Dandong, China.
Discov Oncol. 2023 Apr 20;14(1):45. doi: 10.1007/s12672-023-00656-9.
WD repeat domain 76 (WDR76) has been reported in multiple tumors, while without relation to chemotherapy resistance. 5-fluorouracil (5-FU) is widely adopted in treating colon cancer. However, the resistance of WDR76 and 5-FU in colon cancer remains unclear.
Limma package in R software was employed to analyze the differentially expressed genes. Western blot or quantitative real-time PCR (qRT-PCR) were run to assessed the gene expression. The cytotoxic effect was determined according to cell viability assay, colony formation assay in vitro. Cell apoptosis was assayed using flow cytometry. GSEA analysis was performed to identify pathways related to the target gene. Xenografted mice model was employed to evaluate the tumor growth.
Bioinformatic analysis revealed the higher expression of WDR76 in 5-FU sensitive colon cancer cells compared to resistant colon cancer cells, accompanied by the decreased mRNA expression of WDR76 in 5-FU resistant colon cancer cells. The overexpressed WDR76 resulted in the apoptosis and the downregulated colony numbers in 5-FU resistant colon cancer cells, leading to the elevated sensitivity of 5-FU. Meanwhile, knockdown of WDR76 enhances the resistance of 5-FU in colon cancer both in vitro and vivo, which was reversed by a specific inhibitor of HRAS, Kobe006. An important molecular mechanism of 5-FU resistance lies the degradation of HRAS induced by WDR76.
Our findings demonstrated a role of WDR76 as a promising target for reversing the resistance of colon cancer to 5-FU.
WD重复结构域76（WDR76）已在多种肿瘤中被报道，但与化疗耐药性无关。5-氟尿嘧啶（5-FU）被广泛用于治疗结肠癌。然而，WDR76与5-FU在结肠癌中的耐药性仍不清楚。
使用R软件中的Limma包分析差异表达基因。通过蛋白质免疫印迹或定量实时聚合酶链反应（qRT-PCR）评估基因表达。根据细胞活力测定、体外集落形成测定确定细胞毒性作用。使用流式细胞术检测细胞凋亡。进行基因集富集分析（GSEA）以鉴定与靶基因相关的通路。采用异种移植小鼠模型评估肿瘤生长。
生物信息学分析显示，与5-FU耐药的结肠癌细胞相比，5-FU敏感的结肠癌细胞中WDR76表达更高，同时5-FU耐药的结肠癌细胞中WDR76的mRNA表达降低。过表达WDR76导致5-FU耐药的结肠癌细胞凋亡增加且集落数量减少，从而提高了5-FU的敏感性。同时，敲低WDR76增强了结肠癌在体外和体内对5-FU的耐药性，而HRAS的特异性抑制剂Kobe006可逆转这种耐药性。5-FU耐药的一个重要分子机制在于WDR76诱导的HRAS降解。
我们的研究结果表明WDR76作为逆转结肠癌对5-FU耐药性的一个有前景的靶点发挥了作用。
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