【作者】 李雯；

【导师】 方浩；

【作者基本信息】 山东大学 ， 药物化学， 2021， 硕士

【摘要】 组蛋白去乙酰化酶(Histone deacetylases,HDACs)广泛存在,通过影响基因转录,对机体表观遗传进行调控。HDAC参与了多种生理过程,并有望成为各种病理条件的药物靶标,例如癌症,神经退行性疾病,炎症,代谢和免疫疾病,病毒和寄生虫感染等。但是,目前上市的HDAC抑制药物均为泛抑制剂,由于它们靶向多种HDAC亚型酶,所以均伴随不可避免的不良反应,因此亚型选择性抑制剂的研究越来越受到关注,尤其是在癌症疾病领域。组蛋白去乙酰化酶8(HDAC8)属于HDACI亚族,是一种锌离子依赖的酶,广泛分布于细胞核与细胞质中。除了组蛋白外,HDAC8的底物多种多样,例如SMC3、雌激素相关受体α(ERRα)、p53和ARID1a等,因此它能参与各项生命活动,它的异常的高表达水平会伴随恶性肿瘤在内的多种疾病。本文通过分析HDAC8与PCI-34051晶体结构,发现在活性位点附近存在“foot pocket”,能够容纳疏水性基团,因此我们以PCI-34051为先导化合物,通过使用不同取代酰肼结构替换异羟肟酸,设计A系列化合物。结合课题组前期四氢异喹啉类HDAC8抑制剂的研究,利用四氢喹啉环作为中间连接基团,取代酰肼或者异羟肟酸基团作为ZBG,取代苄基等作为酶表面识别基团,设计B系列目标化合物。在目标物的合成中,我们合成新目标物27个,新的中间体20个,共计新化合物47个。所有的目标物经过氢谱、碳谱以及高分辨质谱进行结构确证。我们对目标化合物进行了体外的HDAC8酶活性抑制实验,选择抑酶活性相对较好的目标化合物进行了 HDAC亚型选择性实验、蛋白印迹实验以及体外的抗肿瘤细胞增殖实验。初步生物活性评价结果表明,所有目标化合物对HDAC8的抑制活性高于已上市的泛HDAC抑制剂SAHA。在蛋白印迹实验中,目标化合物Lw-3能够有效提高SMC3的乙酰化水平,对Tubulin和H3的乙酰化水平影响较小。在HDAC亚型选择性抑制实验中,目标化合物显示出相对于HDAC1、HDAC2、HDAC6的HDAC8选择性。在对六株肿瘤细胞的体外抗增殖实验中,目标化合物Lw-18、Lw-22显示出优于PCI-34051的抗肿瘤增殖活性,这进一步说明该系列抑制剂设计的选择性具备一定合理性。更多还原

【Abstract】 Histone deacetylases(HDACs)are wide existed in human body and is related with removing acetyl group on the ε-amino group of the acetylated lysine of histones and performing epigenetic regulation.It becomes a drug target for various pathological conditions,such as cancer,metabolic and immune diseases,inflammation,viral and parasitic infections,neurodegenerative diseases,etc.At present,five drugs have been approved for cancer treatment,and a large number of new HDAC inhibitors are also in the clinical and preclinical research stage.However,the currently marketed HDAC inhibitors are all pan-inhibitors and lead to inevitable adverse reactions.Therefore,the research of subtype selective inhibitors has attracted more and more attention,especially in the field of developing anticancer agents.As a zinc ion-dependent enzyme,histone deacetylase 8(HDAC8)belongs to the HDAC class I subfamily and is widely distributed in the nucleus and cytoplasm.HDAC8 has many kind of substrates,such as histones,SMC3,estrogen-related receptor alpha(ERRα),p53 and ARID1a,etc.,so it can participate in various life activities,and i abnormal expression can cause cancers.In this paper,it is found that there is a "foot pocket" near the active site by analyzing the crystal structure of HDAC8 and PCI-34051,which can accommodate the hydrophobic group.Therefore,we use PCI-34051 as the lead compound to design series A compounds,replacing the hydroxamic acid group with different substituted hydrazides.Combining with the previous research on HDAC8 inhibitors based on the tetrahydroisoquinoline,target series B compounds were designed according to following strategies:(1)tetrahydroquinoline was used as the linking group;(2)hydrazide or hydroxamic acid group were used as ZBG;(3)benzyl group was used as the enzyme surface recognition group.In this thesis,twenty new intermediates were synthezied and totally forty-seven new compounds were obtained.Structures of all target compounds were identified.Preliminary biological evaluation were performed in vitro on HDAC8 enzyme inhibition on target compounds.Then target compounds with good inhibitory activities were tested their HDAC subtype selectivities,anti-proliferative activities and western blot experiments.The preliminary biological activity evaluation results show that the inhibitory activity of all compound against HDAC8 is significantly higher than that of the pan-HD AC inhibitor SAHA.In Western blotting experiments,the target compound Lw-3 can effectively increase the acetylation level of SMC3,but has little effect on the acetylation level of Tubulin and H3.In the HDAC subtype selective inhibition experiment,which confirmed its HDAC8 selectivities in celluar level.Target compounds Lw-18 and Lw-22 showed better anti-proliferative activities against six tumor cell lines compared with PCI-34051,which could be helpful in our further step on drug discovery.更多还原