AR通过招募LSD1调控ZEB2的转录活性促进骨肉瘤转移的分子机制研究

There are more men than women in cases of osteosarcoma metastasis, which suggests that androgen/androgen receptors (AR) may be involved in the regulation of osteosarcoma metastasis, but we still know little about its molecular mechanism. In our study, we found that the activated AR could potentiate the promoter activity of the Zinc finger E-box Binding homeobox 2 (ZEB2) in osteosarcoma cells. We also found that the Lysine Specific Demethylase 1 (LSD1) could be recruited by AR to regulate the downstream target genes. However whether AR promotes the osteosarcoma metastasis via the regulation of ZEB2, and whether or not LSD1 is involved in the regulation of AR on the ZEB2, as well as the interactions mechanism between the three of AR, LSD1 and ZEB2 is still unknown. So in this project we will confirm the binding site of AR on the ZEB2 promoter and test the hypothesis that AR could promote osteosarcoma metastasis through recruitment of LSD1 to regulate the expression of ZEB2 via epigenetic modifications at the cellular and molecular lever. We also will study the effect of the AR-blocked drugs on this pathway and assess the therapeutic effect of the drugs on the osteosarcoma metastasis in animals. All of these will provide some new targets and ideas for clinical diagnosis and treatment of osteosarcoma.

骨肉瘤异位转移病例中男性多于女性，提示雄激素/雄激素受体（AR）可能参与介导骨肉瘤异位转移过程，但其分子机制知之甚少。预实验发现，在骨肉瘤细胞中雄激素激活的AR能够显著增强转录因子E-box锌指结合蛋白-2（ZEB2）的启动子活性，同时活化的AR可以招募组蛋白去甲基化酶-1（LSD1）调控靶基因的转录活性。然而AR是否通过调控ZEB2影响骨肉瘤的异位转移，AR调控ZEB2过程中又是否有LSD1的参与，以及AR、LSD1与ZEB2三者间的相互作用机制依然不明。本课题将在骨肉瘤细胞中确定AR与ZEB2启动子的相互作用位点；在分子和细胞水平验证AR通过招募LSD1调控ZEB2的转录活性进而促进骨肉瘤异位转移的假说；在动物及组织水平观察抗AR药物治疗对上述分子通路的影响及其体内治疗效果，为未来临床骨肉瘤的诊治提供新的靶点和思路。

背景：骨肉瘤是最常见的原发性恶性骨肿瘤，好发于儿童和青少年，由于其极易发生早期异位转移，且有较高的致死率，成为严重威胁儿童和青少年生命健康的病患之一。骨肉瘤早期异位转移最常见的靶器官是肺，约占90%左右，且男性患者明显多于女性，这让我们意识到雄激素可能是影响骨肉瘤发生发展的始动或驱动因素之一。.主要研究内容：通过本项目的研究确定雄激素受体AR与ZEB2启动子的相互作用位点；阐明AR通过招募LSD1影响ZEB2启动子的甲基化水平，调控ZEB2的表达，进而调节骨肉瘤转移的分子机制；分析雄激素、LSD1与骨肉瘤转移发生率及预后的相关性，为临床骨肉瘤的诊治提供新的靶点和思路。.实验结果：我们明确了AR、LSD1和ZEB2对骨肉瘤细胞迁移和侵袭的影响；证实了AR可以直接与ZEB2启动子区域相结合，并确定了AR与ZEB2启动子的相互作用位点；确定了AR能够通过招募LSD1在ZEB2启动子的H3K9位富集并去甲基化，激活ZEB2的转录，上调ZEB2的表达，从而促进骨肉瘤细胞发生迁移和侵袭；在动物水平揭示了AR和LSD1在骨肉瘤转移中的作用及抗AR药物治疗的效果；在组织水平确定AR、LSD1与骨肉瘤恶性程度的相关性。.结论及科学意义：我们阐明了AR与LSD1通过调控ZEB2的转录活性进而调节骨肉瘤转移的分子机制，完善了AR的调控网络，为未来临床骨肉瘤的诊治提供了新的靶点和思路，为未来临床治疗和药物开发提供了理论基础。