Discovery of Critical Host Genes Enabling Resistance to HIV-1 Infection as Therap

描述（由申请人提供）：人类免疫缺陷病毒（HIV-1）是对公共卫生的全球性威胁，对药物开发提出了前所未有的挑战。直接针对HIV-1病毒的药物往往由于耐药性的迅速出现而失败。功能遗传学（FGI）使用了一种专有技术，随机纯合基因扰动（RHGP），发现了新的宿主基因靶点，可以阻断HIV-1感染，而不会对细胞存活产生明显的有害影响。与病毒靶点不同，依赖于细胞靶点的治疗有望对包括耐药病毒在内的所有艾滋病毒变体保持广谱疗效。此外，通过消除对病毒的选择性压力，宿主靶向可以规避耐药性的发展。在I期研究中确定的靶点使用siRNA方法进行了验证，并且也被证明是CXCR4和CCR5热带hiv病毒感染所必需的。这些结果证明了FGI专有的RHGP技术在人类基因组中识别阻断病毒感染同时无毒的宿主靶标的能力。在II期提案中，我们将使用单克隆抗体疗法开发针对HIV的新型靶向干预措施。我们将重点关注一个特定的宿主靶点，“Robo1”，它对HIV-1的生命周期至关重要。为此，我们发现在HIV-1感染的原发CD4+ T淋巴细胞中，Robo1的表达被激活。因此，Robo1独特地出现在产生病毒的活细胞表面。Robo1抗体在人PBMC检测中显示出抗hiv -1活性。因此，rob1可以被单克隆抗体靶向，阻断HIV生命周期和/或通过正常的宿主防御机制，如抗体依赖性细胞毒性（ADCC）和/或补体依赖性细胞毒性（CDC）消除感染细胞。我们将对人类scFv文库进行基于噬菌体的筛选，以确定选择性识别HIV-1感染而非未感染细胞的候选抗体。这些候选抗体将被构建成全长人IgG1抗体，利用分子进化方法，候选抗体的结合亲和力将被提高几个数量级。然后将评估它们的广谱抗病毒能力，包括在体外直接抑制病毒复制和/或介导ADCC和CDC效应，以及在人源化小鼠中的功效。抗体导联将通过基于细胞、动物和组织的交叉反应性分析进一步仔细检查。基于抗病毒活性和迭代改进，我们将提名一个主要的Robo1抗体候选物用于ind研究。我们相信选择性靶向HIV感染细胞的概念具有令人兴奋的前景，并且开发针对广谱HIV分离株的治疗性单克隆抗体是可行的。

DESCRIPTION (provided by applicant): Human Immunodeficiency Virus (HIV-1) is a global threat to public health and poses an unprecedented challenge to drug development. HIV-1 drugs that directly target virus often fail due to rapid emergence of drug resistance. Functional Genetics (FGI) used a proprietary technology, Random Homozygous Gene Perturbation (RHGP) and discovered novel host gene targets that block HIV-1 infection without observable deleterious effect on cell survival. Unlike viral-targets, treatments relying on cellular targets are expected to retain broad spectrum efficacy against all HIV variants including drug-resistant viruses. In addition, by removing the selective pressure on the virus, host targeting circumvents drug resistance development. The identified targets in Phase I studies were validated using a siRNA approach and also shown to be commonly necessary for infection by both CXCR4 and CCR5 tropic HIV1 viruses. These results demonstrate the power of FGI proprietary RHGP technology to identify host targets(s) in human genome that block viral infection and at the same time are non-toxic. In the Phase II proposal, we will develop novel targeted intervention against HIV using monoclonal antibody therapeutics. We will focus on one particular host target, "Robo1", which is essential for the life cycle of HIV-1. To this end, we have discovered that Robo1 expression is activated in primary CD4+ T lymphocytes by HIV-1 infection. As a result, Robo1 uniquely appears on the surface of live, viral producing cells. Robo1 antibodies exhibited anti-HIV-1 activity in human PBMC assays. Robo1 thus can be targeted by monoclonal antibodies to block HIV life cycle and/or to eliminate infected cells via normal host defense mechanisms such as antibody-dependent cellular cytotoxicity (ADCC) and/or complement-dependent cytotoxicity (CDC). We will conduct phage-based screening of human scFv libraries to identify antibody candidates that selectively recognize HIV-1 infected but not uninfected cells. These leading scFv candidates will be built out into full- length human IgG1 antibodies and using molecular evolution approach, the binding affinities of lead antibody candidates will be enhanced by several orders of magnitude. Their broad spectrum anti-viral abilities will then be evaluated including direct inhibition of viral replication and/or mediation of ADCC and CDC effects in vitro and efficacy in humanized mice. Antibody lead(s) will be further scrutinized through cell-based, animal-based and tissue cross-reactivity assays. Based on antiviral activities and iterative improvements, we will nominate a lead Robo1 antibody candidate for IND-enabling studies. We believe the concept of selective targeting of HIV- infected cells has exciting prospects and that it is feasible to develop a therapeutic monoclonal antibody against a broad spectrum of HIV isolates. PUBLIC HEALTH RELEVANCE: Most current AIDS drugs target the HIV virus and therefore enable the development of drug resistance through viral mutation. The Phase I project seeks to identify human host targets (as opposed to HIV virus targets) that will prevent the HIV virus from using the host's cellular mechanism for its life cycle. The overall goal of Phase II is to develop Host Oriented Therapeutic antibodies using targets identified from Phase I. These antibodies are expected to be active against all variants of HIV, including those with resistance to current antiviral therapies.