探討人類免疫缺乏病毒非核苷酸類似物抑制劑抗藥性突變L100V/L100I與多型性突變之功能性影響

Functional characterization of HIV-1 NNRTI-resistance mutation L100V/L100I and related polymorphic mutations

張庭瑋(Ting-Wei Chang)

指導教授 ： 張淑媛

國立臺灣大學/醫學院/醫學檢驗暨生物技術學研究所/碩士(2016年)

https://doi.org/10.6342/NTU201601261

摘要

HIV-1高效率抗反轉錄病毒治療(highly active antiretroviral therapy, HAART)能有效降低病人的致死率與發病率。然而因有些病人服藥順從性不佳及病毒本身高突變率的因素，使抗藥性病毒株的盛行率也增加。在台灣，HARRT第一線用藥主要是兩種NRTI(nucleoside reverse transcriptase inhibitor)藥物加一種NNRTI(non-nucleoside reverse transcriptase inhibitor)藥物。因為NNRTI的基因屏障比較低，且目前已知主要的NNRTI藥物抗藥性突變會造成多種NNRTI藥物感受性下降，所以NNRTI藥物抗藥性的發生率也逐年增加。除了主要的抗藥性位點之外，相關的突變也隨之被觀察到，但是這些相關突變對病毒生長與藥物感受性之影響仍不清楚。
　　本實驗室首先針對2008-2015台灣地區5114條HIV-1反轉錄酶序列進行分析。NNRTI主要突變以反轉錄酶上K103、V179、V106與L100為多數。本實驗目的為確認L100I/V是否確實對NNRTI有抗性，並想進一步了解多型性突變I202V之影響。首先我們比較病毒生長能力，發現HXB2/L100V與HXB2/L100V+I202V病毒複製能力明顯低於原生型HXB2病毒；同時發現HXB2/L100V與HXB2/L100V+I202V反轉錄酶活性也顯著低於帶有其它突變位點之病毒反轉錄酶活性。HXB2/L100V與HXB2/L100I對於NNRTI藥物 NVP、EFV與ETR皆具有抗性。HXB2/L100V/I加入I202V多型性突變後，會增加對NVP與ETR的感受性；相反的HXB2/L100V/I+I202V卻會降低對EFV的感受性。
　　總結本研究結果，反轉錄酶上L100V與L100I點突變，的確會降低反轉錄酶活性，進而影響病毒複製的能力，也會造成病毒對於NNRTI的感受性下降。

關鍵字

非核?酸類似物抑制劑 ； L100V ； L100I

並列摘要

The morbidity and mortality of HIV-1-infected indviduals has reduced after introduction of highly active antiretroviral therapy (HAART). However, the poor adherence of HIV patients and the high mutation rate of HIV-1 result in the increased prevalence of drug-resistant viruses. In Taiwan, the first line regimen of HAART consists of 2 nucleoside reverse transcriptase inhibitors (NRTIs), and one non-nucleoside reverse transcriptase inhibitor (NNRTI). Due to the lower genetic barrier of NNRTI and the major NNRTI mutations often caused cross resistance to other drugs in the same category, the prevalence of NNRTIs resistant virus has increased recently. Nevertheless, the impacts of some NNRTI related mutations and polymorphisms on drug susceptibility and virus fitness are still unclear.
In this study, a total of 5114 HIV-1 reverse transcriptase sequences amplified from HIV-1 infected, treatment-naive patients from 2008 to 2015 were analyzed. Among these NNRTI-related major mutations, K103, V179, V106 and L100 were dominant.We aimed to determine the effects of L100V/I mutation and its compensatory mutations on virus fitness and susceptibility to NNRTIs. First, the HXB2/L100V and HXB2/L100V+I202V viruses showed slight impairments in replication as well as reverse transcriptase activity when compared to that of the parental wild-type HXB2 viruses. In virus susceptibility to NNRTIs, HXB2/L100V and HXB2/L100I had reduced susceptibility to NVP, EFV and ETR. These virus harbored I202V mutations also showed increased susceptibility to NVP and ETR. Neverless, HXB2/L100V+I202V and HXB2/L100I+I202V reduced virus susceptibility to EFV when compared to that of the single L100-mutant viruses. In addition, HXB2 / L100V and HXB2 / L100I mutations had no effect on viral susceptibility to RPV.
In summary, the presence of L100V/I mutationreduced reverse transcriptase activity and virus replication capacity. The virus susceptibility to NNRTIs was also decreased.

並列關鍵字

NNRTI ； L100V ； L100I

參考文獻

1. S. J. Flint , V.R.Racaniello, L. W. Enquist , A. M. Skalka, Principles of virology. human immunodeficiency virus pathogenesis. Vol. 2. 2009.

2. UNAIDS, Global AIDS Update 2016, 2016.

3. de Bethune, M.P., Non-nucleoside reverse transcriptase inhibitors (NNRTIs), their discovery, development, and use in the treatment of HIV-1 infection: a review of the last 20 years (1989-2009). Antiviral Res, 2010. 85(1): p. 75-90.

4. Flexner, C., HIV drug development: the next 25 years. Nat Rev Drug Discov, 2007. 6(12): p. 959-66.

5. Singh, K., B. Marchand, K.A. Kirby, E. Michailidis, and S.G. Sarafianos, Structural Aspects of Drug Resistance and Inhibition of HIV-1 Reverse Transcriptase. Viruses, 2010. 2(2): p. 606-638.