HIV Reverse Transcriptase-Mediated Mutagenesis

描述(申请人提供)：HIV-1具有很高的变异率，这有助于其逃避宿主免疫系统的能力，限制抗逆转录病毒药物的疗效和驱动 出现抗药性。导致耐药性的突变以及其他病毒突变主要归因于逆转录酶(RT)容易出错的性质。故意增加RT介导的突变，通过将突变率增加到不能维持病毒种群生存的水平，从而降低病毒的传染性。它的效力 通过增加RT介导的错误来降低HIV-1的传染性，导致了发现可能增加HIV-1变异率的小分子的倡议。一个跨学科的协作团队已经组建起来，1)进行发现研究，以确定增加RT介导的错误的新的小分子；2)使用分子分析来确定小分子增加艾滋病毒变异率并导致病毒灭绝的机制(S)，以及3)使用生化方法评估RT介导的突变的机制。通过初步研究，我们已经确定了四个增加RT介导的突变的小分子。为了阐明驱动这种增加的结构-活性关系并优化这种活性，我们将首先进行发现研究，以确定可以增加RT介导的错误的小分子。这些分子的抗病毒和诱变活性将在细胞培养中进行评估。其次，我们将使用细胞培养方法研究小分子在HIV-1中诱导突变和导致病毒灭绝的机制。在这里，我们将研究我们已经发现的小分子，以及我们识别的任何铅分子。第三，我们将使用生化方法来研究作用机制，以阐明RT介导的突变增加的机制基础。这些研究的成功完成将对RT介导的病毒突变机制及其对病毒复制和灭绝的影响提供更深入的了解。

DESCRIPTION (provided by applicant): HIV-1 has a high mutation rate, which contributes to its ability to evade the host immune system, limits the efficacy of antiretroviral drugs and drives the emergence of drug resistance. Drug resistance conferring mutations as well as other viral mutations are primarily attributed to the error-prone nature of reverse transcriptase (RT). An intentional increase in RT-mediated mutations decreases virus infectivity by increasing the mutation rate to a level that is not able to maintain survival of the virus population. The potency by which HIV-1 infectivity can be decreased by increasing RT-mediated errors has led to an initiative to discover small molecules that may increase the HIV-1 mutation rate. An interdisciplinary collaborative team has been assembled to 1) conduct discovery studies to identify new small molecules that increase RT-mediated errors, 2) use molecular analyses to identify the mechanism(s) by which small molecules increase the HIV mutation rate and result in virus extinction, and 3) to assess the mechanism of RT-mediated mutation using biochemical methods. Through preliminary studies, we have identified four small molecules that increase RT-mediated mutations. In order to elucidate the structure-activity relationship driving this increase and to optimize this activity, we will first pursue discovery studies to identify small molecules that can increase RT-mediated errors. The antiviral and mutagenic activities of these molecules will be assessed in cell culture. Second, we will examine the mechanism by which small molecules induce mutations and cause virus extinction in HIV-1 using cell culture methodologies. Here we will examine small molecules that we have already discovered as well as any lead molecules that we identify. Third, we will investigate the mechanism of action using biochemical methods to elucidate the mechanistic basis for increased RT-mediated mutation. Successful completion of these studies will provide deeper insight into the mechanisms of RT-mediated viral mutagenesis and its impact on viral replication and extinction.