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標題: | 以 MutT Homolog 1 結構為基礎且具抗癌可能性之抑 制物開發研究 Development of structure-based inhibitors of MutT Homolog 1 as potential anti-cancer drug |
其他標題: | Development of structure-based inhibitors of MutT Homolog 1 as potential anti-cancer drug |
作者: | 彭正 Cheng Peng |
指導教授: | 鄭貽生 Yi-Sheng Cheng |
關鍵字: | 氧化dNTP,高通量篩選技術,共結晶結構,MTH1 抑制劑,結構分析, Oxidized dNTP,Ultra high throughput screening,Co-crystal structures,MTH1 inhibitors,Structure analysis, |
出版年 : | 2022 |
學位: | 博士 |
摘要: | 過量的活性氧會導致DNA 複製過程中鹼基配對錯誤與DNA 毀損。由活性氧造成的氧化dNTP (oxidized dNTP) 可由細胞中DNA 修復機制中的MutT Homolog 1 (MTH1) 將其水解。MTH1 可以抑制氧化dNTP 在DNA 複製過程中插入。此功能也可以避免觸發細胞凋亡。MTH1 已經被認為可以作為癌細胞清除氧化dNTP 而避免進入細胞凋亡。因此,抑制MTH1 被認為當作一種癌症治療策略。本研究藉由高通量篩選技術,從含有2,313 種化合物之資料庫中篩選出潛在具有抑制MTH1 的化合物。本研究發現化合物MI0320,MI0639,MI0652 和 MI0861 具潛在抑制 MTH1 之能力,他們的IC50 分別為1.2、1.5、7.2、14 μM。利用細胞生長抑制實驗,發現化合物MI0639 具有抑制效果而其IC50 為 6.5 μM。本研究成功解析出MTH1-MI0639,MTH1-MI0320 和MTH1-MI0861 的共結晶結
構,其解析度為1.8 to 2.2 Å。結構資訊顯示MI0639 的氮原子會與MTH1的Asp119、Asp120 和Asn33 形成氫鍵。以及MTH1 的Phe27、Phe72、Phe74 和Trp117 會與MI0639 的methyl 和ethyl substituent 周邊形成疏水性作用。為了改良MTH1 抑制物的選擇性和專一性,本研究基於共結晶結構分析合成出一系列的衍生化合物。在焦磷酸鹽分析試劑量測MTH1 的活性抑制實驗,其結果顯示未有乙基修飾的scaffold 2 系列其IC50 優於 scaffold 1 系列衍生化合物。這結果是由於scaffold 1系列衍生化合物的N-methyl、N-cyclopropyl substituent 會與ethyl substituent 產生空間阻礙導致N-methyl、N-cyclopropyl substituent 在空間中不穩定。在等溫滴定微量熱儀實驗,其結果顯示scaffold 2 系列衍生化合物的焓會大幅提高。這結果是由於N-methyl、N-cyclopropyl substituent 與MTH1 的Asp119 形成穩定氫鍵。在scaffold 2 系列發現兩種化合物具nM 等級的IC50 抑制能力和Kd 值,MI1024與MI1020。其IC50 為 6 nM 和10 nM。基於共結晶結構資訊,本研究可提供選擇性和專一性優化的MTH1 抑制劑發展策略。 The excessive reactive oxygen species can result in base mispairing and DNA damage in DNA replication. MutT Homolog 1 (MTH1) can catalyze the hydrolysis of oxidized nucleoside triphosphates induced by reactive oxygen species from cellular DNA repairing. MTH1 can restrain oxidized nucleoside triphosphates from incorporation into DNA replication. This function also prevents from the triggering of apoptosis. The MTH1 has been recognized to remove oxidized nucleoside triphosphates and prevent apoptosis. Therefore, MTH1 inhibition is regarded as an anti-cancer therapeutic strategy. In this study, the 2,313 compounds from the database were screened and discovered the potential compounds of MTH1 inhibition by ultra high throughput screening. This research found the potential MTH1 inhibitory compounds MI0639, MI0320, MI0652 and MI0861, their IC50 values were 1.2, 1.5, 7.2 and 14 μM. In the cell growth inhibition experiment, the compound MI0639 possessed the effective inhibitory effect and the IC50 was 6.5 μM. This research successfully resolved the co-crystal structures of MTH1-MI0639, MTH1-MI0320 and MTH1-MI0861 by resolution of 1.8 to 2.2 Å . The structural information revealed the nitrogen atom of MI0639 formed hydrogen bonds with the residues Asp119, Asp120, and Asn33 of MTH1 and hydrophobic effects around the methyl and ethyl substituent of MI0639 by interacting with Phe27, Phe72, Phe74, and Trp117 of MTH1. To improve selectivity and specificity, a series of derivatives were synthesized based on the analysis of the co-crystal structures. In pyrophosphate assay of MTH1 inhibition experiments, the IC50 values of scaffold 2 derivatives without ethyl substituent were better than scaffold 1 derivatives. The result was because of the N-methyl, N-cyclopropyl substituent and ethyl substituent of scaffold 1 compounds made steric hindrance and lead N-methyl and N-cyclopropyl substituent to be unstable in space. In isothermal titration calorimeter experiments, the results revealed the enthalpy of scaffold 2 derivatives were significantly increased. This result was due to stable hydrogen bond formation between N-methyl, N-cyclopropyl substituent and the residue Asp119 of MTH1. In scaffold 2 derivatives, that found two compounds possessed nM-level IC50 inhibitory ability and Kd values, MI1024 and MI1020, their IC50 values were 6 nM and 10 nM. Based on co-crystal structure analysis, this study can provide strategies for the development of MTH1 inhibitors with selectivity and specificity optimization. |
URI: | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/83117 |
DOI: | 10.6342/NTU202210003 |
全文授權: | 未授權 |
顯示於系所單位: | 植物科學研究所 |
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