靶向去乙酰化酶SIRT3抗三阴性乳腺癌的新型激动剂的设计合成及机制研究

Triple negative breast cancer (TNBC) is well known to be an aggressive type characterized by the absence of well-defined molecular targets, including ESR (estrogen receptor), PGR (progesterone receptor), and ERBB2/HER-2 (erbb2 receptor tyrosine kinase 2). Typically, TNBCs have a relatively poor outcome due to the inherently invasive and metastatic clinical behavior, and a lack of effective targeted therapies represents a huge challenge. Studies have shown that the reprogramming of mitochondrial metabolism is very significant in TNBC, and mitochondrial deacetylase SIRT3 plays a very important regulatory role in this process. Based on TCGA combined with tissue chip analysis, the application of biological means, for the first time we confirmed that SIRT3 could be used as a potential new target for TNBC treatment. In the early stage of the work, we predicted the activator site of SIRT3 based on the elastic network method. A lead compound, ZA-17, was discovered through several rounds of modification and screening, which had a high SIRT3 activated activity, cytotoxic activity and moderate migration inhibitory activity and showed good tumor inhibitory activity in vivo. In this project, we will conduct the chemical optimization of ZA-17 to improve its targeting and effectiveness, and reveal the activated mode of SIRT3. Additionally, we will explain the mechanism that ZA-17 targeted SIRT3 and changed mitochondria reprogramming to induce TNBC death at the cellular and molecular level. This study will help to clarify the function and potential of SIRT3 small molecule activators in TNBC therapy, and provide new ideas for the development of novel targeted TNBC drugs.

三阴性乳腺癌（TNBC）由于缺乏有效的内分泌和靶向治疗靶标，目前没有有效的治疗药物，是临床一个重大难题。研究表明线粒体代谢重编程在TNBC中非常显著，而线粒体去乙酰化酶SIRT3在此过程中扮演非常重要的调控作用。申请者基于TCGA结合组织芯片分析，应用生物学手段首次确认了SIRT3可作为TNBC治疗潜在的新靶点。在前期的工作中，我们基于弹性网络的方法预测了SIRT3潜在的激动位点，经过多轮筛选结合药物化学方法得到了一个具有较高SIRT3激动活力，细胞毒活性及中等迁移抑制活性的先导化合物ZA-17，并在体内显示出良好肿瘤抑制活性。在本项目中，我们拟对ZA-17进一步的化学优化，提高其靶向性和有效性，并揭示SIRT3的激动模式，同时在细胞和分子水平上阐明其通过靶向SIRT3影响线粒体重编程从而诱导TNBC死亡的机制，为SIRT3小分子激动剂抗TNBC提供理论和实验基础。

本项目的研究工作基本按照计划执行，在前期研究结果的基础上，我们通过对前期得到的先导化合物进行基于结构的合理药物设计和化学修饰，合成了 40 个系列化合物，并结合多种活性筛选，最终得到一个具有更优活性和靶向性的SIRT3激动剂化合物 33c。进一步我们通过细胞热转变分析（cellular thermal shift assay， CETSA）和 RNAi 等实验验证了化合物 33c 具有良好的靶向性，同时通过免疫荧光，免疫印迹等实验发现化合物 33c 具有诱导三阴性乳腺癌细胞线粒体重编程和自噬相关性细胞死亡的分子机制。进一步我们还通过多种生物学手段，探索了化合物 33c 诱导细胞死亡的潜在新机制，包含33c通过调控SIRT3对TNBC细胞的转移抑制，Warburg 效应瓦解以及自噬和线粒体自噬的激活。另外，体内实验结果表明化合物 33c 具有良好的抗肿瘤活性，同时无明显毒性，具有候选药物的潜质。