本文選題：組蛋白去乙酰化酶 + 組蛋白去乙�；敢种苿���； 參考：《大連理工大學(xué)》2014年碩士論文

【摘要】：在腫瘤的發(fā)生和發(fā)展過程中,表觀遺傳學(xué)修飾起著非常重要的作用。組蛋白修飾是其中的主要方式之一,主要包括乙�；�、甲基化、磷酸化、泛素化等。組蛋白修飾主要由兩種拮抗酶共同調(diào)控：組蛋白乙酰基轉(zhuǎn)移酶(Histone Acetyl Transferases, HATs)、組蛋白去乙�；�(Histone Deacetylases, HDACs)、HDACs將組蛋白N-端賴氨酸殘基側(cè)鏈上的ε-氨基去乙�；�,因此,電荷量增多會促進(jìn)組蛋白與DNA鏈之間的相互作用,從而影響染色質(zhì)結(jié)構(gòu)及基因的表達(dá)。HDACs的高表達(dá)會導(dǎo)致平衡失調(diào)從而引起腫瘤的發(fā)生,尋找高效抑制HDACs為靶點(diǎn)的抑制劑成為研發(fā)新型抗癌藥物的熱點(diǎn)。組蛋白去乙酰化酶抑制劑(Histone Deacetylases Inhibitors, HDACIs)的結(jié)構(gòu)分為三部分：金屬結(jié)合區(qū)(Zinc Binding Group, ZBG)、連接區(qū)(Linker Group)和表面識別區(qū)(Surface Recognition Group)。通過對三部分的結(jié)構(gòu)進(jìn)行設(shè)計(jì),尋找高活性與高選擇性的抑制劑。 本論文根據(jù)上市藥物SAHA、FK228及報道的抑制劑結(jié)構(gòu)特點(diǎn),設(shè)計(jì)了兩個系列化合物。第一系列化合物基于2-氨基-7溴庚酸為骨架,引入不同的疏水結(jié)構(gòu)作為表面識別區(qū)；5個亞甲基作為連接區(qū)；硫代乙�；鳛榻饘俳Y(jié)合區(qū)。第二系列化合物主要以對苯硝基作為金屬結(jié)合區(qū)�；谝陨辖Y(jié)構(gòu)特點(diǎn),設(shè)計(jì)出300余種抑制劑小分子,通過Autodock4.2分子對接軟件初步篩選出13種化合物,并采用多肽液相合成法成功合成了13種化合物10A1-10A6、11B1、12C1-12C2、13-1至16-1。通過了HPLC、ESI-MS、1H NMR與13C NMR的結(jié)構(gòu)驗(yàn)證,與目標(biāo)產(chǎn)物結(jié)構(gòu)一致。 目標(biāo)化合物均顯示很好的酶抑制活性。其中第一系列的12C1(IC50=0.04μm)與12C2(IC50=0.092μm)活性最好,在表面識別區(qū)引入2-氨基-4-苯基噻唑可以提高抑制劑活性。對HDAC1與HDAC6的抑制活性,與對HDAC的抑制活性呈現(xiàn)相同趨勢。 在目標(biāo)化合物對腫瘤細(xì)胞的抗增殖活性實(shí)驗(yàn)中,對HeLa(宮頸癌細(xì)胞系)的選擇性要高于MCF-7(乳腺癌細(xì)胞系)與SMMC-7721(肝癌細(xì)胞系)�；衔�12C1(IC50=40.3μM)對HeLa的抑制作用優(yōu)于TSA (IC50=48.5μM)。12C1、12C2較對照TSA的酶抑制活性低10倍,但在細(xì)胞活性實(shí)驗(yàn)中,12C1活性優(yōu)于TSA1.21倍。測試化合物中的硫代乙�；惢衔锱c對照TSA分別進(jìn)行抗增殖活性水平與酶抑制活性水平比較,硫代乙酰基類化合物在抗增殖活性水平上比對照TSA有更高的提升。說明硫代乙�；诩�(xì)胞內(nèi)水解為巰基,促進(jìn)與Zn2+發(fā)生作用,達(dá)到與TSA相同的抑制水平。由于脂溶性,滲透性的原因造成其它化合物的抑制活性不高。 通過對接實(shí)驗(yàn)中的結(jié)合能分析,化合物中可旋轉(zhuǎn)的單鍵數(shù)目過多,與復(fù)合物之間的靜電能作用太大,都不利于與酶的相互作用。HDAC4-HDACIs復(fù)合物之間的對接能量與HDAC2-HDACIs復(fù)合物具有相同的分布趨勢。在目標(biāo)化合物與HDAC2的對接實(shí)驗(yàn)中,12C1與HDAC2形成的氫鍵數(shù)目多于其它,且只有12C1表面識別區(qū)的2-氨基-4-苯基噻唑中的N原子參與氫鍵的形成。在目標(biāo)化合物與HDAC4的對接實(shí)驗(yàn)中,TSA中異羥肟酸基形成的氫鍵多于硫代乙�；�,與體外酶抑制活性一致。巰基與酶的相互作用高于硫代乙�；�,因此,硫代乙�；诩�(xì)胞內(nèi)水解為巰基后會提高與酶的相互作用,這與體外抗增殖活性實(shí)驗(yàn)較酶抑制活性有較大提高相符。
[Abstract]:Epigenetic modification plays a very important role in the development and development of tumor. Histone modification is one of the main ways, including acetylation, methylation, phosphorylation, and ubiquitination. Histone modification is mainly regulated by two kinds of antagonists: histone acetyltransferase (Histone Acetyl Transferases, H) ATs), the histone deacetylation enzyme (Histone Deacetylases, HDACs), HDACs deacetylation of the epsilon amino group on the side chain of the protein N- terminal lysine residue, therefore, the increase of the charge will promote the interaction between the histone and the DNA chain, thus affecting the chromatin structure and the gene expression of the.HDACs, resulting in the imbalance of the balance resulting from the expression of.HDACs. The occurrence of tumor and the search for the inhibitors that target the HDACs as the target are the hot spots in the development of new anticancer drugs. The structure of Histone Deacetylases Inhibitors (HDACIs) is divided into three parts: the metal binding zone (Zinc Binding Group, ZBG), the connection area (Linker Group) and the surface recognition area (Surface) Tion Group). Through the design of the three part of the structure, we look for inhibitors with high activity and high selectivity.
In this paper, two series of compounds were designed based on the structure characteristics of SAHA, FK228 and reported inhibitors. The first series of compounds were based on 2- amino -7 bromoheptanic acid as the skeleton, and different hydrophobic structures were introduced as surface recognition areas; 5 methylene was used as the connection area; thioacetyl was used as the metal binding zone. Second series of compounds were used. Based on the above structural characteristics, more than 300 small molecules of inhibitor are designed based on the above structural characteristics. 13 compounds are screened by Autodock4.2 molecular docking software, and 13 compounds, 10A1-10A6,11B1,12C1-12C2,13-1 to 16-1., are successfully synthesized through the synthesis of HPLC, ESI-MS, 1H NMR and 13C. The structural verification of the NMR is consistent with the structure of the target product.
The target compounds all showed good enzyme inhibitory activity. The first series of 12C1 (IC50=0.04 m) and 12C2 (IC50=0.092 mu m) activity was the best. The introduction of 2- amino -4- phenyl thiazole in the surface recognition area could improve the activity of the inhibitor. The inhibitory activity of HDAC1 and HDAC6 showed the same trend as the inhibitory activity to HDAC.
In the experiment of anti proliferation activity of target compounds to tumor cells, the selectivity of HeLa (cervical cancer cell line) was higher than that of MCF-7 (breast cancer cell line) and SMMC-7721 (liver cancer cell line). Compound 12C1 (IC50=40.3 mu M) inhibited HeLa better than TSA (IC50=48.5 micron M).12C1,12C2 10 times lower than that of control TSA enzyme inhibitory activity, but in cell In the activity test, the activity of 12C1 was better than that of TSA1.21. The antiproliferative activity level of the thioacetyl group in the test compound compared with the control TSA was compared with that of the enzyme inhibitory activity. The thioacetyl group had a higher increase in the anti proliferative activity level than that of the control TSA. It can promote the function of Zn2+ and achieve the same inhibition level as TSA. Due to fat solubility and permeability, the inhibitory activity of other compounds is not high.
Through the binding energy analysis in the docking experiments, the number of rotatable single bonds in the compound is too much, and the electrostatic energy between the compounds is too large, and the interaction between the.HDAC4-HDACIs complex and the HDAC2-HDACIs complex has the same distribution trend. In the docking experiment between the target compound and the HDAC2, The number of hydrogen bonds formed by 12C1 and HDAC2 is more than that of the others, and only the N atoms of the 2- amino -4- phenyl thiazole in the 12C1 surface recognition region are involved in the formation of hydrogen bonds. In the docking experiment between the target compound and HDAC4, the hydrogen bonds formed by the hydroxamic acid group in TSA are more than the thioacetyl group, which is consistent with the inhibitory activity of the enzyme in vitro. The interaction of the sulfhydryl group with the enzyme is high. Therefore, the thioacetyl group can increase the interaction with the enzyme after the hydrolysis of thioacetyl in the cell, which is more consistent with the inhibition activity of anti proliferative activity in vitro than in the enzyme inhibition activity.
【學(xué)位授予單位】：大連理工大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2014
【分類號】：R91;O629

【參考文獻(xiàn)】

相關(guān)期刊論文 前6條

1 曾紅梅;陳萬青;;中國癌癥流行病學(xué)與防治研究現(xiàn)狀[J];化學(xué)進(jìn)展;2013年09期

2 周r，

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