AZ-AZIN作为抗肿瘤治疗分子靶点的实验研究

The rapid growth of tumor cells is highly dependent on polyamines, which makes the polyamine metabolic pathway, especially the rate-limiting enzyme--ornithine decarboxylase (ODC), become a novel target for anti-tumor therapy. However, the clinical effect of ODC inhibitor was not good. Recent studies have demonstrated that antizyme (AZ) and antizyme inhibitor (AZIN) play important roles in regulating the activity of ODC and the concentration of cellular polyamine. Our previous studies have showed that the over-expression of AZ and knock-down of AZIN inhibited the proliferation of tumor cells and the cell cycle was arrested in G0/G1 phase. In this project, we want to explore the potential value of AZ/AZIN as a molecular target of anti-tumor therapy. We intend to increase the level of intra-cellular OAZ through multiple methods, including up-regulating the expression of AZ, down-regulating exprssion of AZIN and designing small molecule inhibitor to interfere the interaction between AZ and AZIN. We will further determine if the increased AZ can inhibit the activity of ODC and prevent polyamine in the tumor microenvironment entering into cell, which can ultimately inhibit the growth of tumor cells and induce the cell apopotosis. This project will contribute to providing some theoretical and experimental basis for developing anti-polyamine micromolecular drug, which will offer new methods and strategies for treatment and prevention of tumor.

肿瘤细胞快速生长对多胺的高度依赖，使多胺代谢途径，特别是多胺合成限速酶鸟氨酸脱羧酶（ODC），成为抗肿瘤治疗的新靶点。然而，直接靶向ODC的抑制剂作为抗肿瘤药物用于临床却疗效不佳。近来研究表明，由抗酶（AZ）和抗酶抑制因子（AZIN）构成的网络在调节ODC活性和细胞内多胺含量中发挥重要作用。本课题的前期研究发现，AZ高表达和AZIN表达抑制可抑制肿瘤细胞增殖，使细胞阻滞于G0/G1期。在此基础上，本项目拟探索AZ/AZIN作为抗肿瘤治疗分子靶点的潜在价值，通过在肿瘤细胞中增强AZ表达、阻遏AZIN表达以及设计筛选小分子抑制剂用于干扰AZ/AZIN的相互作用等多种手段，增加细胞内功能性AZ的水平，继而抑制ODC活性和阻止肿瘤微环境中的多胺入胞而耗竭细胞内的多胺，以达到抑制肿瘤细胞增殖和诱导其凋亡的目的。本项目的成功实施，将为临床肿瘤防治提供新思路和新途径，为新型抗肿瘤药物的研发奠定基础。

肿瘤细胞快速生长对多胺的高度依赖，使多胺代谢途径，特别是多胺合成限速酶鸟氨酸脱羧酶（ODC），成为抗肿瘤治疗的新靶点。然而，直接靶向ODC的抑制剂作为抗肿瘤药物用于临床却疗效不佳。近来研究表明，由抗酶（AZ）和抗酶抑制因子（AZIN）构成的网络在调节ODC活性和细胞内多胺含量中发挥重要作用。本项目设计和筛选了小分子抑制剂用于干扰 AZ-AZIN 相互作用并分析其对肿瘤细胞的影响；并通过上调AZ、下调AZIN表达等多种手段，增加细胞内功能性AZ的水平，继而抑制ODC活性，以达到抑制肿瘤细胞增殖和诱导其凋亡的目的。实验结果显示：1）用构建好的药效团模型对含100万个小分子化合物specs小分子数据库进行筛选，并进一步根据对接打分的排序，挑选出58个小分子化合物。挑选出的小分子化合物通过体外细胞水平的活性测试进一步筛选发现19号小分子能明显促进AZ的表达及抑制AZIN、ODC的表达，从而抑制肿瘤细胞生长，诱导肿瘤细胞凋亡。2）促进肿瘤细胞AZ和（或）抑制AZIN的表达，能调节肿瘤细胞多胺及多胺代谢酶的表达水平，进一步抑制肿瘤细胞的生长。本研究成功筛选了干扰AZ-AZIN 相互结合的小分子化合物，为新型抗肿瘤药物的研发奠定了基础。