Multifunctional Human Anti-HIV Antibodies

描述（由申请人提供）：在该申请中，我们提议通过将HIV-1融合抑制剂与异常广泛中和抗体（bNABS）偶联来创建一类新的抗HIV-1免疫疗法。HIV-1利用的细胞表面受体（CD4、CCR5、CXCR4）是有吸引力的治疗靶点，因为它们没有进化耐药性的压力。不幸的是，HIV包膜蛋白（Env）突变很容易进化以逃避小分子受体的封锁。同样地，已经发现bNABs可以中和高达90%的测试病毒株，但当HIV受到抗体的挑战时，体内耐药性很快就会产生。在这两种情况下，抗性都是由于Env的突变，尽管逃离一个分子的突变会使艾滋病毒对另一个分子更敏感。我们的假设是，用抗hiv融合抑制剂修饰的bNAB将是一种非常有效的化合物，而充分修饰的bNAB将是多功能的，并作为单分子“鸡尾酒”。这种化合物将使HIV很难进化出耐药性，因为它需要许多突变才能逃离bnab的结合位点和空间阻断的细胞受体位点。我们建议合成经过验证的靶向共受体CCR5和CXCR4的HIV小分子抑制剂，以及结合和破坏CD4/gp120融合的小分子。这些化合物将与蛋白质反应性片段结合合成，这将允许对bNABs进行特异性修饰。我们的实验室最近开发了两种不同的方法，用于酪氨酸残基的特异性靶向。表征良好且具有广泛中和作用的抗体PG9和VRCO1将作为修饰的支架。除了化学修饰，我们还将研究创建遗传融合，将PG9和VRCO1的Fv结合到单个分子上，以及设计Fc结构域以增加体内半衰期。多特异性抗体将被评估其中和广泛的HIV-1分离株的能力，包括对bNABs和小分子融合抑制剂具有抗性的菌株。然后测定成功的候选抗体的半衰期和毒性。最后，多特异性抗体将用于HIV逃逸试验，以验证我们的假设，即这些分子可以构成HIV无法回答的进化挑战。我们预计，这项研究的产品将是特别有效和广泛活跃的HIV-1免疫治疗和预防。

DESCRIPTION (provided by applicant): In this application, we are proposing to create a novel class of anti-HIV-1 immunotherapeutics by conjugating HIV-1 fusion inhibitors to exceptionally broadly neutralizing antibodies (bNABS). The cell surface receptors utilized by HIV-1 (CD4, CCR5, CXCR4) are attractive targets for therapy because they are under no pressure to evolve resistance. Unfortunately, HIV envelope protein (Env) mutations are readily evolved to escape small molecule receptor blockade. Likewise, bNABs have been discovered that neutralize up to 90% of tested viral strains but when HIV is challenged with antibodies, in vivo resistance quickly develops. In both cases, the resistance is due to mutations in the Env, although mutations to escape one molecule can make HIV more sensitive to another. Our hypothesis is that a bNAB modified with anti-HIV fusion inhibitor would be an exceptionally potent compound and that a sufficiently modified bNAB would be multifunctional and act as single molecule "cocktail." Such a compound would make evolution of resistance very difficult for HIV because it would require many mutations to escape the binding sites of the bNABs and the sterically blocked cell receptor sites. We propose to synthesize validated small molecule HIV inhibitors that target the co-receptors CCR5 and CXCR4, as well as, small molecules that bind and disrupt the CD4/gp120 fusion. These compounds will be synthesized linked to protein reactive moieties which will allow for specific modification of bNABs. Our lab has recently developed two different approaches for specific targeting of tyrosine residues for this purpose. The well characterized and very broadly neutralizing antibodies PG9 and VRCO1 will serve as the scaffold for modification. In addition to chemical modification, we will also investigate creating genetic fusions to combine the Fv's of PG9 and VRCO1 onto a single molecule, as well as, engineering the Fc domain to increase in vivo half-life. Multi-specific antibodies will be assessed for their ability to neutralize a broad panel of HIV-1 isolates including strains which are resistant to the bNABs and the small molecule fusion inhibitors. Successful candidate antibodies will then be assayed for half-life and toxicity. Finally, multi-specific antibodies will be used in an HIV escape assay to test our hypothesis that these molecules can pose an evolutionary challenge that HIV cannot answer. We anticipate that the products of this research will be exceptionally potent and broadly active HIV-1 immunotherapeutics and prophylactics.