Guo Wenting, Li Xiaojia, Fan Jingyu, Li Hongwei, Wen Yan, Meng Chunyan, Chen Haitao, Zhao Zhipeng, Zhang Yuling, Du Yushen, Wu Baixing
Guangdong Provincial Key Laboratory of Malignant Tumor Epigenetics and Gene Regulation, Guangdong-Hong Kong Joint Laboratory for RNA Medicine, Medical Research Center, Sun Yat-Sen Memorial Hospital, Sun Yat-Sen University, Guangzhou 510120, China.
Cancer Institute (Key Laboratory of Cancer Prevention and Intervention, China National Ministry of Education), The Second Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, Zhejiang, China.
iScience. 2023 Aug 18;26(9):107672. doi: 10.1016/j.isci.2023.107672. eCollection 2023 Sep 15.
Non-natural nucleobase isocytosine (IC) is the isomer of cytosine; its chemical derivate 5-fluoroisocytosine (5-FIC) together with the isocytosine-specific deaminase (ICD) VCZ was suggested to be potential practical enzyme/prodrug pair for cancer therapy through gene-directed enzyme-prodrug therapy (GDEPT) method. In this study, we have determined the crystal structures of apo-VCZ and its complex with 5-FU. We identified the critical residues for substrate binding and catalytic reaction. We also captured the substrate-induced conformational changes of VCZ, then proposed the conjectural reaction procedures of VCZ for converting the IC into the uracil. Moreover, we evaluated the therapeutic effect of wildtype or the mutated VCZ protein in the colorectal cancer cell lines. Our studies will shed light on optimizing the ICD/5-FIC pairs by modifying either the enzyme or the prodrug based on the structural observations, thereby improving the possibility of applying the ICD/5-FIC pair in clinical trials.
非天然核碱基异胞嘧啶（IC）是胞嘧啶的异构体；其化学衍生物5-氟异胞嘧啶（5-FIC）与异胞嘧啶特异性脱氨酶（ICD）VCZ一起，被认为是通过基因导向酶-前药疗法（GDEPT）用于癌症治疗的潜在实用酶/前药对。在本研究中，我们确定了脱辅基VCZ及其与5-FU复合物的晶体结构。我们鉴定了底物结合和催化反应的关键残基。我们还捕捉到了底物诱导的VCZ构象变化，进而提出了VCZ将IC转化为尿嘧啶的推测反应过程。此外，我们评估了野生型或突变型VCZ蛋白在结肠癌细胞系中的治疗效果。我们的研究将基于结构观察结果，通过修饰酶或前药来优化ICD/5-FIC对，从而提高ICD/5-FIC对在临床试验中应用的可能性。
