以四環素及其衍生物為第二型登革熱病毒之抑制劑

Abstract

世界衛生組織估計每年發生約5000萬的登革熱病例，而目前仍沒有商業化且有效的治療方式及疫苗。先前實驗室與生資所楊進木老師合作進行以登革病毒2型的外膜蛋白的結構進行virtual screening篩選出可與其docking的臨床藥物清單，其中含有tetracycline及其衍生物rolitetracycline。本研究以此為基礎，針對tetracycline及其衍生物在培養的細胞上進行測試，以確認其對登革病毒的抑制能力。利用登革熱二型PL046病毒株進行溶斑試驗來測試，最後得到三個四環素衍生物在溶斑試驗中對登革熱二型病毒展現出抑制能力。 而在此初步結論下，為進一步推測其機制，於是進行時間點的測試。而其中doxycycline與rolitetracycline表現出來的是在加入病毒前加入和與病毒同時加入這兩個時間點可以觀察到抑制的現象，而chlortetracycline則是持續表現出抑制能力，直至加入病毒顆粒後的10小時後。此外，為了建構登革病毒之expression clone以進一步探討其機制，便設計將本實驗室所擁有的cDNA clone做修飾，在3’UTR端後面加上ribozyme使得此cDNA clone所產生的RNA可自行將因cloning過程產生的多餘序列移除，以期可以利用修正過的clone來進一步探討藥物的作用機制，因此建構HCV ribozyme之construct。

The WHO has estimated there are about 50 million dengue cases each year globally. Currently there is no commercial available vaccine and treatment. In previous study, this laboratory has obtained a list of compound candidates by applying virtual screening based on molecular docking and cluster analysis to select for commercial available medical compounds interacting with dengue E protein and, in theory, blocking the infection in cell cultures. Among them, there are tetracycline derivatives. This has raised the interest in other tetracycline derivatives. In this study, I tested the effectiveness of those tetracycline compounds were tested in cell culture system. In all, seven tetracycline compounds were assayed for their inhibitory effect on DV2 PL046. Three have showed inhibitory activities on DV2 PL046 in cell cultures. In addition, I have also constructed an HCV-ribozyme plasmid for the purpose of attaching to the 3’end of a DV2 cDNA clone to remove extra sequence when expressed in RNA form.