AAV Core

载体核心的主要目的是提供高质量的自身互补（sc）和单链（ss）rAAV 1载体，并对适用于rAAV 1抗病毒研究的NHP动物进行血清学筛选。我们将通过以下具体目标实现这些目标： 目标1。设计，创建，生产和质量控制测试scAAV 1载体批次在不同规模的各种转基因和表达盒，以满足本计划项目的其他研究者的特定需求。更具体地说，120只恒河猴将在5年内入组不同的研究，数百只小鼠将用于猕猴前评价。平均而言，我们估计每年将生产15-20个载体批次，以满足这些研究的需要。 目标二。通过使用体外和体内中和抗体（NAB）试验筛选NHP人群中预先存在的抗AAV 1免疫力，以选择无AAV 1-NAB的动物进行疫苗和治疗研究。在rAAV 1介导的抗HIV免疫粘附素基因转移中，需要对NHP群体进行预筛选，以选择不存在rAAV 1中和抗体（NAB）的动物。我们的数据表明，灵长类动物来源的AAV在NHP人群中的血清学流行率为60- 80%。这意味着我们可能必须筛选超过360只动物以鉴定120只无AAV 1 NAB的动物。 目标3。开发新型和可扩展的rAAV生产方法，用于更大规模的翻译NHP研究和基于rAAV 1的抗HIV疫苗和治疗剂的未来临床开发。我们目前的AAV生产系统应能满足本项目前期的载体需求。然而，大规模的载体生产可能会成为更大的翻译NHP研究和未来临床开发的瓶颈。我们将利用我们在开发各种载体包装细胞系和基于感染的载体生产系统方面的丰富经验，开发基于293细胞感染的新型可扩展生产方法，以克服这一限制。

The main objectives of the Vector Core are to provide high quality self-complementary (sc) and single-stranded (ss) rAAV1 vectors and serologically screen NHP animals suitable for rAAV1 anti-viral studies. We will accomplish these goals through the following specific aims: Aim 1. To design, create, produce and quality control test scAAV1 vector lots at different scales with a variety of transgenes and expression cassettes to serve the specific needs of other investigators of this program project. More specifically, 120 rhesus macaques will be enrolled for different studies over 5 years and hundreds of mice will used for pre-macaque evaluation. In average, we estimate that 15-20 vector lots will be produced annually to meet the needs of those studies. Aim 2. To screen NHP populations for pre-existing immunity against AAV1 by using both in vitro and in vivo neutralizing antibody (NAB) assays to select AAV1-NAB free animals for vaccine and therapeutic studies. Pre-screening of NHP population to select the animals without preexisting neutralizing antibody (NAB) to rAAV1 is essential for rAAV1-mediated anti-HIV immunoadhesin gene transfer. Our data suggested that the serological prevalence of primate-derived AAVs in NHP populations ranges from 60-80%. This implies that we may have to screen more than 360 animals to identify 120 animals free of AAV1 NAB. Aim 3. To develop novel and scalable rAAV production method for larger scale translational NHP studies and future clinical development of rAAV1-based anti- HIV vaccine and therapeutics. Our current AAV production system should meet the vector needs in the early stage of this program project. However, large scale vector production may become a bottle neck for larger translational NHP studies and future clinical development as well. We will utilize our extensive experience in developing various vector packaging cell lines and infection-based vector production system to develop a 293 cell infection-based novel and scalable production method to overcome this limitation.