Multifunctional Human Anti-HIV Antibodies

描述(由申请人提供)：在本申请中，我们建议通过将HIV-1融合抑制剂与异常广泛的中和抗体(BNAbs)相结合来创建一种新型的抗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，因为它们没有形成耐药性的压力。不幸的是，HIV包膜蛋白(Env)突变很容易进化来逃避小分子受体的阻断。同样，已经发现bNAbs可以中和高达90%的测试病毒株，但当HIV用抗体攻击时，体内很快就会产生耐药性。在这两种情况下，耐药性都是由于环境病毒的突变，尽管为了逃避一个分子而发生的突变会使艾滋病毒对另一个分子更加敏感。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，并充当单分子“鸡尾酒”。这种化合物将使艾滋病毒的耐药性进化变得非常困难，因为它需要许多突变才能逃脱bNAbs的结合部位和空间上被阻止的细胞受体部位。我们建议合成针对辅助受体CCR5和CXCR4的有效小分子HIV抑制剂，以及结合和破坏CD4/gp120融合的小分子。这些化合物将被合成连接到蛋白质活性部分，这将允许对bNAbs进行特定的修饰。我们的实验室最近为此目的开发了两种不同的针对酪氨酸残基的特定靶向方法。具有良好特性和非常广泛的中和抗体PG9和VRCO1将作为修饰的支架。除了化学修饰，我们还将研究创造基因融合，将PG9和VRCO1的Fv结合到单个分子上，以及改造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药性的株。然后将对成功的候选抗体进行半衰期和毒性测试。最后，多特异性抗体将用于艾滋病毒逃逸试验，以检验我们的假设，即这些分子可以构成艾滋病毒无法回答的进化挑战。我们预计，这项研究的产品将是非常有效和广泛有效的艾滋病毒-1免疫疗法和预防性药物。 与公共卫生相关：这项提案旨在开发特别有效和广泛有效的抗艾滋病毒-1疗法和预防性药物。尽管目前的逆转录病毒药物鸡尾酒在控制大多数人的艾滋病毒-1方面非常有效，但艾滋病毒-1最终会对所有药物产生抗药性。我们建议创造一种新型的化学修饰的抗HIV抗体，通过多种机制阻止HIV-1进入。这种多特异性免疫疗法将提供有效的长寿抗艾滋病毒治疗/预防，并且艾滋病毒-1将很难产生抗药性。

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. PUBLIC HEALTH RELEVANCE: This proposal aims to develop exceptionally potent and broadly active anti-HIV-1 therapeutics and prophylactics. Although current retroviral drug cocktails are very effective in controlling HIV-1 in most individuals, HIV-1 eventually evolves resistance to all drugs. We propose to create a novel type of chemically modified anti-HIV antibody that blocks HIV-1 entry through multiple mechanisms. This multi-specific immunotherapeutic would provide potent long-lived anti-HIV therapy/prophylaxis and would be very hard for HIV-1 to evolve resistance against.