核苷類衍生物之合成及其生物活性的應用

Abstract

近年來細菌具有抗藥性的比例逐漸上升，為了要阻止細菌對人類所造成的威脅，設計新的抗生素為首要的課題之一，而細菌細胞壁是細菌存活很重要的結構之一。細菌壁的生合成中有許多的酵素參與，MraY為其中很重要的磷酸轉移酶 (phosphate transferase)，核苷類似物的分子對此具有高度的生物活性，因此我們以此為主架構，設計一系列的核苷類分子。 在我的實驗中，建構方便及快速的合成方法是相當重要的，為了簡化複雜的天然物結構，尤其是繁瑣的立體中心的建立，我們使用三唑 (triazole) 片段來嫁接分子，並且使用醯胺鍵 (amide bond) 或是異氰酸酯基團 (isocyanate) 快速製造一系列的分子，以取代嫁接胺基核醣 (amino ribose) 所需要用到複雜的醣基化反應，因此可以利用平行合成的方式、機器的輔助，以及固相萃取得到分子庫。在觀察細菌生長的型態中，我們選用枯草桿菌 (B. subtilis 168)，可以發現部分分子會影響細菌的生長及分裂。 在含有胺基核醣 (amino ribose) 的系列分子中，我們設計了脂質鏈片段的修飾以及立體中心的改變，並且進行抑制酵素活性以及最低抑制濃度 (MIC) 的測試，發現具有長碳鏈的分子對於革蘭氏陽性菌 (Gram-positive bacteria) 或是革蘭氏陰性菌 (Gram-negative bacteria) 都有較好的最低抑制濃度。 另一方面，MraY的受質含有核苷鹼基的結構，因此我們想要在鹼基上做修飾，並且進行生物抑制活性的探討。分子設計方面，我們可以在脲嘧啶 (uracil) 五號位的碳 (C-5 position) 上利用胺化反應 (amination) 轉換成胺基，進行醯胺鍵的連接，或是利用薗頭耦合反應 (Sonogashira reaction) 得到碳-碳鍵的連結，再進行修飾，而在六號位的碳 (C-6 position) 則可以利用強鹼置換出連接橋梁，以此得到相對應以UMP為骨架的小型分子庫，並測試MraY的抑制活性。

Due to serious antibacterial resistance, new antibacterial drugs, new targets or new treatments are urgently needed to combat the global spread of multi-drug resistant bacteria, such as MRSA. Translocase MraY, an integral membrane protein, plays an important role in bacterial cell wall biosynthesis (or peptidoglycan biosynthesis). Thus, it is considered a potential and promising antibacterial target. In natural products, several structurally complicated nucleoside analogues are identified to target MraY and exhibit interesting antibacterial activity. Their structures contain several feature moieties including a uridine, an aminoribose, a lipid side chain and multi-stereogenic centers, which become a hurdle to efficiently prepare these molecules or molecular libraries for further biological or antibacterial studies. Herein, we at first plan to simplify these complicated structures and investigate the essential moieties, followed by developing a convenient approach to prepare nucleoside analogue-based libraries for bacterial morphology study and antibacterial evaluation. In this study, my research work can be divided into three parts. Firstly, in order to develop a general and flexible synthetic approach to prepare nucleoside derivatives, we designed to efficiently assemble feature moieties through a triazole formation (the click chemistry), an urea formation, and an amide bond formation to build up the backbone template and install a diverse substituent. Notably, the original glycosidic bond and an aminoribose moiety were replaced with a simplified fragment. The proper application of semi-automatic such as solution-phase synthesizer, multichannel liquid handler, and vacuum centrifuge can allow us to prepare the high quality products efficiently. Interestingly, several synthetic molecules with simplified structures are capable of affecting bacterial morphology and growth. Secondly, we planned to investigate a varied lipophilic side chain and the chiral center linked to the aminoribosyluridine moiety. The antibacterial results suggested that the proper length of the lipid chain in nucleosides is required against Gram-positive and Gram-negative bacteria. Besides, the preliminary results in our laboratory suggest modifications of UMP (Uridine monophosphate) might affect the enzyme-based Lipid I synthesis catalyzed by MraY. In the third part, we plan to develop a method to build a UMP -based library with a substituent diversity on the C-5 or C-6 position. Through my effort, a more convenient approach has been established to build up these nucleoside molecules. In addition, several essential moieties have been identified. Importantly, the proper length of the lipid chain has been discovered. This valuable information is a foundation to allow us for further modifications to improve inhibition potency against MraY or antibacterial activity.