BMVC-12C-P誘導白色念珠菌粒線體失能導致之生物效應及相關分子機制探討

Abstract

念珠菌是造成院內感染的重要致病菌，在臨床上念珠菌抗藥性問題也益發嚴重。目前所使用的抗真菌藥物種類不多，加上這些抗真菌劑之副作用強而造成使用上的限制，因此開發新作用機制的抗真菌藥物有其必要與急迫性。目前研究指出粒線體可能是一個良好的抗真菌藥物標的，其原因是由於粒線體參與在真核細胞內廣泛的生理調控角色，如各式各樣的生理代謝，包括了致病因子的調控。在確認小分子化合物BMVC-12C-P是能夠專一的進入粒線體並以粒線體做為主要攻擊標的而造成白色念珠菌粒線體的失能後，我們利用BMVC-12C-P作為工具，並配合白色念珠菌突變菌株庫的使用來探討白色念珠菌粒線體的失能對菌體造成的生物效應。本研究已經建立有效篩選突變菌株庫的篩選模式並在轉錄因子和激酶之突變菌株庫篩選出會受到BMVC-12C-P所影響的突變菌株。由於對於粒線體與致病機制的關係較感興趣，故將篩選出的突變菌株依其所參與的致病機制做歸類，結果以參與在菌絲的生成相關機制為大宗，故本研究進一步挑選菌絲作為探討主軸。我們發現BMVC-12C-P能夠抑制菌絲與生物膜生成之能力，並進一步以Q-PCR確認BMVC-12C-P能透過抑制菌絲轉錄因子EFG1、BRG1、CPH1、CPH2和TEC1的mRNA表現，然而其不透過Hog1MAPK訊息傳遞路徑去調控這些受到BMVC-12C-P所影響的菌絲轉錄因子。未來希望能夠更進一步了解粒線體是透過怎樣的訊息傳遞路徑來調控這些可能參與在菌絲生成的轉錄因子，並且去了解這些轉錄因子在白色念珠菌菌絲生成機制中所扮演的角色，以利未來我們能更深入了解白色念珠菌粒線體參與在菌絲調控的致病機制，更可以做為未來作為新抗真菌藥物發展之重要基石。於此，本研究亦探討BMVC-12C-P對不同標準念珠菌屬及來自臨床檢體中所篩選出來的具抗藥性之念珠菌屬的藥物感受性，發現BMVC-12C-P具有相當優異的殺菌能力，具有發展成良好抗真菌劑之潛力。

Candida species are important pathogens causing nosocomial infections and the prob-lem of antifungal drugs resistance in clinical is also a serious problem. At present, we do not have many choices for antifungal agents to use so it is necessary and urgent to develop new antifungal drugs. In this study, we explored BMVC-12C-P can specifically enter the mito-chondria to cause the mitochondrial dysfunction and we also confirmed the mitochondria is the main target of BMVC-12C-P. Therefore, BMVC-12C-P was used as a tool and com-bined with C. albicans mutant library to investigate the biological effects of mitochondrial dysfunction induced by BMVC-12C-P. We had established the optimal screening model to screen and selected the candidates from mutant library of transcription factors and kinases. Because we are interested in the relationship between mitochondria and pathogenic mecha-nisms, candidates are classified according to the pathogenic mechanisms involved. As a re-sult, the pathogenic mechanism of candidates involved in the formation of hyphae was most, so we selected the regulation of hyphae to explore. We confirmed BMVC-12C-P can inhibit the formation of hyphae and biofilm. We also found the ability of hyphae switch was sup-pressed which correlated with the reduced mRNA expressions of genes involved in hyphal formation (EFG1、BRG1、CPH1、CPH2 and TEC1) and not through Hog1MAPK pathway to regulate them. In this way, we hope to further understand the mechanism of BMVC-12C-P on the regulation of hyphae and can be used as an important cornerstone for the development of new antifungal drugs in the future. Moreover, we also found that BMVC-12C-P displayed the strongest antifungal activities to against Candida species and even Fluconazole-resistant clinical isolates of Candida species that indicated BMVC-12C-P can be used as a highly potential antifungal agent.