Fusion assay of the HIV-induced CD4 down-modulation

HIV 诱导的 CD4 下调的融合测定

• 批准号: 7140590
• 负责人: JUAN LAMA
• 金额: $ 32.3万
• 依托单位: LA JOLLA INST FOR MOLECULAR MEDICINE
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-07-01 至 2008-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7140590
• 关键词: CD4 molecule; HIV envelope protein gp120; HIV infections; antiAIDS agent; cell fusion; combinatorial chemistry; human tissue; protein protein interaction; small molecule; technology /technique development; virus infection mechanism

DESCRIPTION (provided by applicant): Currently used antiretrovirals targeting the viral reverse transcriptase, protease, and envelope proteins have reduced mortality and morbidity in HIV-infected patients. However, the emergence of viral strains resistant to the inhibitors, and the appearance of side effects after prolonged drug administration pose important problems to the efficacy of HAART treatments. Characterization and evaluation of novel targets for therapeutic intervention is a priority in HIV/AIDS research. CD4 down-modulation plays an important role in HIV pathogenesis. In vitro, failure to down-modulate CD4 results in the production of HIV-1 particles with severely reduced infectivity whereas in vivo enhanced receptor down-modulation confers a replicative advantage to HIV-1 in primary lymphocytes. To date, no specific inhibitors of this function have been characterized. To facilitate the discovery of new drugs, we propose to develop a fusion-based assay to screen for specific inhibitors of the HIV-induced CD4 down-modulation activity. This assay will rely on the fact that the rate of fusion between gp120-expressing cells and CD4-positive cells can be modulated by the concentration of these proteins on the cell surface. We hypothesize that since HIV down-modulates expression of CD4, interfering with this function in infected cells will result in higher amounts of surface-CD4, and therefore fusion will occur to a higher extent than in cells in which efficient down-modulation occurs. Experiments to evaluate the sensitivity and specificity of this assay are proposed in Specific Aim 1. To validate the assay, we will screen synthetic combinatorial libraries for potential inhibitors of the virus-induced CD4 down-modulation (Specific Aim 2). We hypothesize that drugs inhibiting this function will impair HIV replication. Our goal is to identify lead compounds with defined chemical structures that could evolve into a discovery program for a new class of anti-HIV drugs. The specific aims are: Aim 1, to develop a fusion-based assay for the identification of inhibitors of the HIV-induced CD4 down-modulation, and; Aim 2, to characterize small-molecule inhibitors of the HIV-induced CD4 down-modulation activity.

描述（由申请人提供）： 目前使用的靶向病毒逆转录酶、蛋白酶和包膜蛋白的抗逆转录病毒药物降低了HIV感染患者的死亡率和发病率。然而，对抑制剂耐药的病毒株的出现，以及长期给药后副作用的出现，对HAART治疗的功效造成了重要的问题。描述和评价治疗干预的新靶点是艾滋病毒/艾滋病研究的优先事项。CD 4下调在HIV发病机制中起重要作用。在体外，未能下调CD 4导致HIV-1颗粒的产生，感染性严重降低，而在体内增强受体下调赋予HIV-1在原代淋巴细胞中的复制优势。迄今为止，还没有这种功能的特异性抑制剂的特点。为了促进新药的发现，我们建议开发一种基于融合的检测方法来筛选HIV诱导的CD 4下调活性的特异性抑制剂。该测定将依赖于这样的事实，即表达gp 120的细胞和CD 4阳性细胞之间的融合速率可以通过细胞表面上这些蛋白质的浓度来调节。我们假设，由于HIV下调了CD 4的表达，干扰感染细胞中的这种功能将导致更高量的表面-CD 4，因此融合将比发生有效下调的细胞中发生的程度更高。在具体目标1中提出了评价该测定法灵敏度和特异性的实验。为了验证该测定法，我们将筛选合成组合文库中病毒诱导的CD 4下调的潜在抑制剂（特异性目的2）。我们假设抑制这种功能的药物会损害HIV的复制。我们的目标是确定具有确定化学结构的先导化合物，这些化合物可以发展成为一类新的抗HIV药物的发现计划。 具体目标是：目的1，开发用于鉴定HIV诱导的CD 4下调的抑制剂的基于融合的测定;目的2，表征HIV诱导的CD 4下调活性的小分子抑制剂。