Proteolytic Antibody HIVcides

描述(由申请人提供)：一种用于艾滋病毒预防的特定、不可逆转和具有成本效益的局部杀微生物剂将有助于减缓大流行的性传播。在这里，我们建议开发能够降解HIV包膜蛋白gp120的催化抗体作为候选HIV杀菌剂。抗体有希望的特征是永久破坏包膜蛋白，重复使用单个催化剂分子来降解数千个gp120分子，以及识别保守的gp120区域，即超抗原区，允许中和不同的HIV毒株。来自非感染者和HIV感染者的多克隆抗体研究表明，高水平的蛋白水解性和HIV中和活性是IgA类抗体的一个值得注意的特性。在R21项目阶段，我们提出了现有抗体及其单链抗体(ScFv)、IgA和Ig G变体的特征。现有的抗体是通过用亲电的gp120类似物免疫获得的，该类似物诱导具有增强的亲核反应能力的抗体，这是催化反应的先决条件。使用针对gp120超抗原位点的亲电探针，从狼疮患者的免疫谱系克隆的单链抗体中获得了额外的蛋白水解性抗体，这些患者倾向于产生针对gp120超抗原位点的具有蛋白水解性的抗体。这些抗体将根据HIV降解效率、HIV中和的特异性、效力和广度以及在阴道环境中执行这些功能的能力来表征。为了分离新的、改进的抗体，我们将在R21项目阶段从HIV阴性和HIV阳性的受试者中筛选唾液和血清IgA的蛋白分解和HIV中和活性。在R33阶段，基于亲电gp120肽类似物的共价结合，通过细胞分离从淋巴细胞中克隆出具有所需特性的单抗，这是一种与蛋白水解性抗体产生细胞相关的特性。体内抗体效力的原则证明将在R33阶段使用SHIV-猕猴感染模型获得。此外，还将在模拟性交后阴道环境的条件下，测试这些抗体作为杀微生物剂辅料的体外活性。这些研究可能会确定适合进一步开发为局部HIV杀伤性药物的蛋白水解性抗体。

DESCRIPTION (provided by applicant): A specific, irreversible and cost-effective topical microbicide for HIV prophylaxis will help slow sexual transmission of the pandemic. Here, we propose the development of catalytic antibodies capable of degrading the HIV envelope protein gp120 as candidate HIVcides. The promising features of the antibodies are the permanent destruction of the envelope protein, reuse of a single catalyst molecule to degrade thousands of gp120 molecules and the recognition of a conserved gp120 region, the superantigen region, permitting neutralization of diverse HIV strains. Polyclonal antibody studies from uninfected and HIV infected subjects indicated that high level proteolytic and HIV neutralizing activities is a noteworthy property of IgA class antibodies. In the R21 project phase, characterization of our existing antibodies and their single chain Fv (scFv), IgA and IgG variants is proposed. The existing antibodies were obtained as IgGs by immunization with an electrophilic gp120 analog that induces antibodies with enhanced nucleophilic reactivity, a prerequisite for the catalytic reaction. Using an electrophilic probe to the gp120 superantigen site, additional proteolytic antibodies were obtained as scFv constructs cloned from the immune repertoire of lupus patients, who tend to produce antibodies with proteolytic activity directed to the gp120 superantigen site. The antibodies will be characterized with respect to HIV degrading efficiency; specificity, potency and breadth of HIV neutralization; and the ability to perform these functions in the vaginal milieu. To isolate novel, improved antibodies, we will screen the proteolytic and HIV neutralizing activity of salivary and serum IgAs from HIV-negative and HIV-positive subjects in the R21 project phase. In the R33 phase, monoclonal IgAs with the desired properties will be cloned from lymphocytes by cell-fractionation based on covalent binding of an electrophilic gp120 peptide analog, a property associated with proteolytic antibody-producing cells. Proof-of-principle for in vivo antibody efficacy will be obtained in the R33 phase using the SHIV-macaque model of infection. The Abs will also be examined for activity in vitro as microbicide excipient formulations under conditions simulating the vaginal environment following sexual intercourse. These studies may identify proteolytic antibodies suitable for further development as a topical HIVcide.