Using adenoviral vectors to express broadly neutralizing anti-gp160 antibodies

描述（由申请人提供）：最近的估计表明，美国有110万人感染人类免疫缺陷病毒（HIV），每年有超过56,000例新感染。在这些新感染中，12%是静脉注射毒品（IDU，共用受污染的针头）的直接结果，32%是由于与IDU有关的行为（与IDU者的异性恋和同性恋接触）。根据疾病控制中心的数据，IDU被认为是艾滋病毒感染的第三大危险因素，仅次于男男性行为者和无保护的异性恋者。虽然预防和治疗药物使用可以减轻艾滋病毒的发病率，但鉴于注射过量者的人数和社会经济地位，目前可能无法实现这一目标，因为资金和获得资源的机会不足；因此，在此期间，疾病的治疗是关键。对于美国和发展中国家的艾滋病毒感染者来说，高效抗逆转录病毒疗法（HAART）已成为主要的治疗方法。HAART的新进展导致越来越多的患者无法检测到病毒血症，CD4+ t细胞计数更高，生活质量更好，总体生存率提高。然而，许多问题仍然存在，如逃逸突变体的出现，患者无法获得和不依从性，不良副作用以及需要持续治疗。重要的是，HAART不影响继续产生病毒的细胞。因此，大多数接受HAART治疗的患者都是低水平的病毒血症，必须无限期地继续治疗以防止病毒反弹。为了从治疗上解决这一问题，我们建议使用第一代辅助腺病毒载体编码抗HIV gp160的广泛中和抗体（bnAbs）。这种bnAbs可以靶向表达抗原的感染细胞，被宿主免疫系统消除，并且使用腺病毒载体可以高效地转导不同类型的细胞进行长期的转基因表达。首先，在第一代和依赖帮助者的腺病毒系统中，将对具有良好特征的bnAbs进行编码。其次，我们将生产和纯化高滴度的载体颗粒，用于体外表征。第三，将使用人源化的HIV感染小鼠模型对载体颗粒进行体内测试，以确定bnab减少病毒复制的有效性。在这个提议中，载体编码的bnAbs将与纯化的重组bnAbs并行测试，并将第一代腺病毒与依赖辅助的腺病毒进行比较。将测试编码bnab的各种组合，以确定针对不同的表位是否能提高疗效。这些目标的完成应有助于深入了解腺病毒介导的载体递送和宿主中和性抗艾滋病毒抗体反应，并提供新的载体介导的持久治疗手段的潜力，最终可能减少感染者（包括使用药物的人）所需的HAART频率和持续时间。

DESCRIPTION (provided by applicant): Recent estimates have suggested there are 1.1 million individuals living with human immunodeficiency virus (HIV) in the United States with over 56,000 new infections arising annually. Of these new infections, 12% are a direct result of intravenous drug use (IDU, sharing of contaminated needles) and 32% are due to IDU-associated behaviors (heterosexual and homosexual contact with an IDU individual). According to the Centers for Disease Control, IDU is considered the third most important risk factor for HIV infection, surpassed by the risk groups of men who have sex with men and individuals engaging in unprotected heterosexual contact. While drug use prevention and treatment can alleviate the incidence of HIV, this may not be currently achievable due to inadequate funding and access to resources given the number and socioeconomic status of IDU individuals; thus in the meantime treatment of the disease is key. For individuals in the US and developing world who live with HIV, highly active antiretroviral therapy (HAART) has become the mainstay of treatment. New developments in HAART have given rise to increasing numbers of patients with undetectable viremia, higher CD4+ T-cell counts, better quality of life, and overall improved survival. However, many issues remain by way of emergence of escape mutants, patient inaccessibility and non-adherence, adverse side effects and the need for ongoing treatment. Importantly, HAART does not affect cells that continue to produce virus. As a result, most patients on HAART have low-level viremia and must remain on treatment indefinitely to prevent viral rebound. To therapeutically address this issue, we propose the use of first generation and helper-dependent adenoviral vectors encoding broadly neutralizing antibodies (bnAbs) against HIV gp160. Such bnAbs can target antigen-expressing, infected cells for elimination by the host immune system, and the use of adenoviral vectors gives high efficiency transduction of diverse cell types for long-term transgene expression. First, well-characterized bnAbs will be encoded in the context of first generation and helper-dependent adenoviral systems. Second, we will produce and purify high-titer stocks of vector particles for in vitro characterization. Third, vectr particles will be tested in vivo with the use of a humanized mouse model of HIV infection to determine the effectiveness of the bnAbs in reducing viral replication. In this proposal vector-encoded bnAbs will be tested in parallel against purified recombinant bnAbs, and first generation adenoviruses compared with helper-dependent adenoviruses. Various combinations of encoded bnAbs will be tested to determine if targeting different epitopes improves efficacy. The completion of these objectives should provide insight into adenoviral-mediated vector delivery and host-neutralizing anti-HIV antibody responses, as well as offer the potential for new vector-mediated means of long-lasting therapy that may eventually reduce the frequency and duration of HAART required for infected individuals, including those who use substances.