组蛋白去乙酰化酶6选择性抑制剂抗肿瘤作用机制及联合治疗研究

Cancer is a leading cause of death all around the world, the burden is increasing due to the growth and aging of the populations. As a result, the development of antitumor drugs appears urgent and more and more efforts have been made in recent years. Targeted drugs are one of the important directions for the development of antitumor drugs, in which, histone deacetylase (HDAC) inhibitors have attracted much attention because of their roles in regulating the expression of genes via deacetylation, which in turn inhibit the proliferation, differentiation and metastasis of tumor cells. However, pan-target inhibitors, such as vorinostat (SAHA), target multiple HDACs, interfere with multiple signaling pathways, and thus resulting in unavoidable side effects. Therefore subtype-selective inhibitors are concerned as effective alternatives to reduce side effects, and HDAC6 is one of the subtype of concern for its unique structure and biochemical functions..Here, we plan to modify the cap structure of SAHA with OBHS (a small antitumor molecule with three-dimensional topological structure), and novel selective HDAC6 inhibitors will be screened out, expecting to efficiently inhibit the growth and metastasis of tumor cells with a higher tumor therapeutic index than SAHA. The further studies will focus on the genome-wide transcriptional profiles of the effect of the candidate compounds on different gastric cancer cells. We plan to use RNA-seq, ChIP-seq as well as other high-throughput sequencing techniques to explore the molecular mechanisms of the candidate compounds regulation on the target genes transcription, especially those participated in the important process of the survival and development of tumor cells (such as cell apoptosis, cell cycle, cell migration and DNA damage, etc.), and then clarify the relationship between the target genes and HDAC6. Furthermore, the studies will also focus on finding new targets and the targeted drugs, and we plan to combine the candidate compounds with those targeted drugs or some other reported targets drugs (such as DNA methylase inhibitors, proteasome inhibitors, etc.) to achieve more efficient combination of antitumor drugs based on the theory of synthetic lethal effect. The underlying molecular mechanisms of combination therapy are also going to be studied..Together, the project is aimed to find new types of selective target drugs for HDAC6 with high efficiency and low toxicity and to achieve a more effective drug combination mode. What’s more, it provides new experimental and theoretical basis for the final application of candidate compounds in clinical oncology treatment.

靶向药物是抗肿瘤药物研发的重要方向之一。组蛋白去乙酰化酶（HDAC）抑制剂通过影响去乙酰化作用而调节基因的表达，进而抑制肿瘤细胞增殖，分化和转移而受到广泛关注。但广谱多靶标抑制剂如伏立诺他(SAHA)靶向于多种HDAC而波及多种信号通路，具有不可避免的副作用。亚型选择性抑制剂是降低副作用的有效选择，HDAC6是其中值得关注的一个亚型。本研究以小分子OBHS修饰SAHA帽子结构，期望得到新型HDAC6选择性抑制剂，能有效抑制肿瘤生长、转移，且相对于SAHA有着较高的肿瘤治疗指数；并采用RNA-seq、ChIP-seq等高通量测序技术着力探究其对胃癌细胞转录组的影响，阐明其调节相关基因表达的分子作用机制及与设计靶点之间的关系，寻找新作用靶点及其他抗肿瘤药物并摸索新的联合治疗组合。本项目力求找到高效低毒新型抗肿瘤靶向药物，实现更为有效的联合用药模式，为最终应用于临床肿瘤治疗提供更多理论依据。

表观遗传修饰可以逆转基因突变给机体造成的影响，因此通过靶向于表观遗传修饰的化合物可以降低化疗副作用及克服抗药性，其中组蛋白乙酰化酶(histone deacetylases, HDAC)业已成为靶向治疗的重要靶点。但临床上使用的广谱多靶标抑制剂如伏立诺他(SAHA)由于靶向于多种HDAC亚型而波及多种信号通路，具有不可避免的副作用。本研究以不同的优势小分子OBHS、OBHSA等基团修饰SAHA帽子结构，从所合成的OBHS-HDACi、OBHSA-HDACi以及ER/HDAC双靶标PROTAC系列化合物中筛选得到具有优异体外治疗指数的苗头分子。从OBHS-HDACi系列缀合物中挑选出具有HDAC6选择性抑制能力的化合物25h，25h对多种肿瘤细胞均有抗增殖能力，尤其对HDAC6高表达的人肝癌细胞HepG2的抑制能力最为显著，25h可将HepG2细胞阻滞在G2/M期，并促进HepG2细胞凋亡；25h能特异性结合HDAC6的催化结构域并抑制HDAC6的酶活，对HDAC6的蛋白质和RNA含量并无明显改变。而OBHSA-HDACi系列缀合物展现了雌激素受体α(ERα)和HDAC的双靶点抑制能力，对于ERα高表达的人乳腺癌细胞MCF-7展现出了最强的抗肿瘤能力；从中挑选出的优势化合物13i能通过泛素蛋白酶体途径降解ERα，13g能与ERα形成稳定构象，从而抑制ERα的核转移或影响其活性。基于OBHS-HDACi以及OBHSA-HDACi的结构基础，设计合成的ERα/HDAC6双靶点PROTAC系列化合物对MCF-7细胞具有显著的抑制增殖的作用，并且显示出突出的针对ERα和HDAC6分子的直接降解能力，可以避免因靶标突变而引起的耐药性；其中苗头分子29f能够诱导MCF-7细胞凋亡。总体来说，OBHS-HDACi、OBHSA-HDACi以及ER/HDAC双靶点PROTAC化合物分别实现了HDAC6选择性抑制、ERα和HDAC双靶标抑制以及双靶标降解的设计目标，满足了不同类型临床肿瘤患者的治疗需求，有望实现精准治疗的目的，本项目的完成具有极大的临床应用价值。.本项目完成过程中一共发表了11篇SCI文章（其中4篇为1区SCI论文），另有2篇SCI文章正在投稿中。共培养博士研究生10名（已毕业6名）。获得国家专利1项。

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