酵素抑制及抗體藥物複合體的研究

Abstract

在論文的第一部分中，我們合成一系列有潛力的抑制劑，其結構與 GlgE催化的起始物以及過渡型態相似，GlgE 是結核分枝桿菌的重要酵素。然而，我們合出來的化合物，包括文獻報導最有效的兩個分子，沒有展現出對分枝桿菌 GlgE的抑制作用。到目前為止，我們依然還不確定哪些因素產生了這樣的問題。 在第二部分中，我們開發了一種利用雙硫化物-炔反應的新方法來修飾含有雙硫鍵的蛋白質。一開始，我們著重於抗體的修飾。儘管小分子雙硫化合物進行的很順利，胱胺酸衍生物卻完全沒有反應。我們接著將注意力轉向利用芳基重氮鹽。藉由甲酸鈉以及藍光發光二極體的引發，產生芳基自由基，進而和包含胱胺酸衍生物的雙硫化合物反應。將其運用於抗體分子中，只有重氮鹽基團無法完全捕獲雙硫鍵的兩個硫原子。 因此，我們合成了另一個新的重氮鹽，其鄰位含有炔丙基醚，以捕獲雙硫鍵的兩個硫原子。我們證明了這個利用新的重氮鹽的方法可用於修飾曲妥珠單抗(賀癌平)，一種人源化單株抗體，儘管由於兩對鏈間的雙硫鍵距離較短而產生半抗體。最後，我們也提出了一種芳基重氮鹽，含有炔丙基醯胺基團，可經由銅催化炔烴-疊氮化物環加成反應來安裝藥物分子。

In the first part of thesis, we synthesized a series of potential inhibitors, which share similar structure with the substrate or transition-state of GlgE, a crucial enzyme for Mycobacterium tuberculosis. However, our synthesized compounds including the two of the most potent molecules reported in literature did not show any inhibitory effect against M. smegmatis GlgE. So far, we have not identified which factors that caused the problem. In the second part of thesis, we developed a new method using disulfide-yne reaction for modification of proteins containing disulfide bonds. At first, we focused on the decoration of an antibody. Although disulfide-yne reaction proceeded smoothly with small disulfide molecules, the reaction of cystine derivatives did not give any yield. We then turned our attention to using aryl diazonium salts. Upon initiation by sodium formate and blue LED light, aryl radicals were generated and reacted with disulfide compounds including cystine derivatives. In application to antibody, the diazonium group alone could not fully trap both sulfur atoms of disulfide bonds. Therefore, we prepared a new aryl diazonium salt with a propargyl ether at the ortho position to catch both sulfur atoms in a disulfide bond. We proved that the method using this new diazonium salt could be applied to modify the disulfide bonds in trastuzumab (Herceptin), a humanized monoclonal antibody, although the half antibody was obtained due to the short distance between two pairs of interchain disulfide bonds. Finally, we proposed an aryl diazonium salt containing a propargyl amide group, which can be utilized for installation of a drug molecule via the copper-catalyzed alkyne-azide cycloaddition reaction.