設計及合成對抗SARS冠狀病毒 之抑制劑: (一) alpha-羥基醯胺與alpha-酮基醯胺 (二) 甘草素衍生物

Abstract

本篇論文主要在研發對抗嚴重呼吸道症候群(severe acute respiratory syndrome, SARS ) 冠狀病毒的試劑。包括兩部分: 第一部份，合成出alpha羥基醯胺與alpha酮基醯胺衍生物，第二部份，合成出甘草素衍生物。 為了模擬SARS病毒主要蛋白酶催化胜肽水解時，所形成的過渡態結構的特性，而合成了alpha-羥基醯胺與alpha-酮基醯胺等衍生物，如化合物12和13，並使用螢光分析方法去檢測其抑制主要蛋白酶活性的效力，發現抑制效果並沒有先前預想的結果好，高達10 microM。 最近國外學者發現甘草素( Glycyrrhizin ) 能夠抑制SARS冠狀病毒的複製，但是所需的劑量太高，而且甘草素在細胞內的抑制機制仍然不明確，因此希望能找到更有效的抑制劑。甘草素的結構主要分為兩個部份: 18β甘草次酸( Glycyrrhizic acid ) 和雙葡萄糖酸的部份。本論文就甘草次酸進行化學修飾，與胺基酸作偶合反應。利用細胞病變效應 (cytopathic effect assay)的檢測，來進一步找到很有效的SARS冠狀病毒抑制劑。 我們發現甘草次酸衍生物( JMF479 ) 對於抑制SARS冠狀病毒抑制效果甚佳，其 IC50與EC50分別是17和3.3 microM。

The aim of this thesis is to discover the antiviral agents against severe acute respiratory syndrome coronavirus (SARS-CoV). This thesis consists of two parts: (i) synthesis of alpha-hydroxy amide and alpha-keto amide derivatives, and (ii) synthesis of glycyrrhetinic acid derivatives. The derivatives of alpha-hydroxy amide and alpha-keto amide, such as 12 and 13 were synthesized, in order to mimic the tetrahedral transition state in the hydrolysis of the peptide substrate catalyzed by the SARS-CoV main protease. The inhibition assays were carried out using a peptide substrate that contains a fluorophore Edans and a quencher Dabcyl. It has been reported that Glycyrrhizin has the activity to inhibit the replication of the SARS coronavirus, but the inhibition concentration is still high. The mechanism of glycyrrhizic acid in vivo is still unclear. It is thus desirable to find better SARS-CoV inhibitors. The structure of Glycyrrhizin contains two parts: 18 beta glycyrrhetinic acid (GA) and disaccharide acid. A series of GA derivatives were prepared by coupling reactions with amino acid esters, and their anti-SARS activities were measured using the CPE (cytopathic effect) assay. We found that a glycyrrhetinic acid derivative (JMF479) was particularly effective against the SARS coronavirus with the IC50 and CPE values of 17 and 3.3 mucroM.