- 學位論文
抗金黃色葡萄球菌感染生物活性藥物的篩選
Screening Strategies for Bioactive Compounds against Staphylococcus aureus Infections
摘要
中文摘要 研究背景 金黃色葡萄球菌是造成皮膚及軟組織感染最常見的致病菌之一，抗藥性細菌的快速崛起擴及全世界，新型抗生素的缺乏，使得感染的症狀變成臨床治療一大考驗，這也是一項值得重視的公共衛生議題。開發新的抗生素至少需花費十年的時間，且由於抗生素的使用療程短、投資效益不高，導致新藥研發缺乏投資管道。因此，評估當前藥物用於抗感染治療的潛力，似乎是縮短新藥研發時程的可行途徑。 實驗方法 研究過程中，藉由文獻搜尋方式，找到金黃色葡萄球菌中可能具抗菌潛力的標的蛋白基因: thioredoxin reductase (TrxR)，以此當作藥物結合標的，利用模擬TrxR分子結構，對已核准的藥物資料庫，以docking的技術進行電腦模擬做虛擬藥物篩選，觀察化合物與標的蛋白質的結合情形。篩選出可能具有抑菌潛力的候選藥物，進行後續的生物活性實驗，以評估藥物對金黃色葡萄球菌的抑菌能力，並利用體外細胞毒性試驗，測試藥物對人類皮膚角質細胞 (human keratinocyte) 及人類單核性細胞 (human monocyte) 活性的影響，評估藥物作為局部給藥途徑使用的發展潛力。 結果 利用電腦虛擬篩選藥物技術篩選出具抗菌潛力的藥物，包括抗精神病、降血糖、降血壓等慢性疾病用藥。抗金黃色葡萄球菌活性的試驗結果顯示thioridazine, telmisartan與glibenclamide在MIC濃度以下就有抑制金黃色葡萄球菌生物膜形成的效果，而glimepiride隨藥物濃度增加，生物膜生成總量也有增加的趨勢。硫氧化還原蛋白還原酶活性試驗中，risperidone, glimepiride與glipizide增加還原酶活性。Glimepiride, glipizide與glibenclamide屬同藥理機轉分類藥物，但對金黃色葡萄球菌生物膜形成的抑制活性及硫氧化還原蛋白還原酶活性的有不同的表現。體外細胞毒性試驗結果顯示，除thioridazine在50 μg/ml以上對THP-1細胞有毒性，其他藥物以細胞對溶媒的最高耐受度配製濃度以下，對人類單核球細胞沒有毒性。相較於人類單核球細胞，藥物對人類角質細胞活性的影響較明顯。 結論 以抗菌標的蛋白分子為基礎的藥物篩選策略，可促進抗金黃色葡萄球菌治療的發展。
並列摘要
Abstract Background Staphylococcus aureus is one of the most common causes of skin and soft-tissue infections. The rapid emergence of resistant bacteria is occurring worldwide. The lack of new antibiotics is a serious public health problem. Developing new antibiotics requires decades, and the drug development pipeline is running dry due to the low market profit for antibiotics. Evaluating currently available drugs for their potential as part of an anti-infection therapy is plausible approach to accelerate the drug development process. Materials and Methods We did literature search for potential anti-staphylococcal targets and performed in silico screenings using ZINC database- US Food and Drug Administration (FDA)-approved drug database for inhibitors of those targets. Candidate compounds with favorable scores and binding orientation were further evaluated experimentally for their ability to inhibit S. aureus growth and to interfere with infection. The toxicity assays including human keratinocyte viability and human monocyte viability were performed to evaluate the potential of these candidate compounds to be applied topically. Results Our data showed that thioridazine, telmisartan and glibenclamide inhibited S. aureus biofilm formation at sub-MIC concentrations, while glimepiride increased biofilm formation. Rispiridone, glimepiride and glipizide increased thioredoxin reductase activity. All the tested drugs except thioridazine showed no significant toxicity to THP-1 cells at the maximum concentration which can be performed. Despite structurally similar, glimepiride, glipizide and glibenclamide had distinct properties on S. aureus biofilm and thioredoxin reductase activity. Conclusions This target based screening strategy can facilitate the development of antistaphylococcal therapy.