Antibody immunotherapeutics for biodefense against Lassa virus

描述（由申请人提供）：该项目将开发针对拉沙病毒（LASV）的生物防御免疫疗法，拉沙病毒是拉沙热（LF）的病原体。LASV是生物安全4级和NIAID生物防御a类药剂。LF是一个重大的公共卫生问题，在西非造成广泛的生命损失和社会混乱。本提案的目标是在治疗LF的新型治疗方法的表征和临床前开发中执行关键步骤。这项工作结合了杜兰大学和德克萨斯大学医学分部的病毒学家和免疫学家的专业知识，以及Zalgen的科学家。Zalgen是一家生物技术公司，专门为具有全球影响的新出现的病毒性疾病开发免疫疗法、快速诊断和新疫苗平台。这里提出的研究将利用我们的最新进展，包括鉴定主要候选人类单克隆抗体（huMAbs），以中和LASV传染性并保护豚鼠免受LASV诱导的致命疾病。Zalgen拥有快速开发（<3个月）高产抗体表达稳定细胞系的独家技术，确保为一级生物防御市场和二级公共卫生市场生产治疗性抗体的成本降到最低。在MILESTONE 1中提出的研究中，我们将在适应哺乳动物细胞瞬时表达平台的无血清培养基中产生数毫克量的IgG1和IgG4 LASV huMAb亚型，用于纯化以进行体外测试。我们还将使用huMAb进行一系列特性分析，重点是鉴定具有广泛LASV中和潜力和低IC50的huMAb或huMAb组合。进一步的研究将建立与相关的新大陆和旧大陆沙粒病毒蛋白的交叉反应性，以及LASV人源抗体与人细胞膜磷脂的反应性。在MILESTONE 2中，我们将在化学定义的无血清培养基中生成CHO或NS0稳定细胞系，并选择高产分离株。限制性稀释细胞克隆（LDCC）将用于分离稳定的、生产级CD-SFM-CHO或CD-SFM-NS0克隆。我们还计划为感兴趣的克隆细胞系生成接入细胞库（ACB），并通过QA/QC转换为生产，以生成克数的候选huMAbs用于体内研究。在MILESTONE 3中，我们将对LASV huMAb亚型在豚鼠（GPs）中的药代动力学进行评估。我们还将在gp中使用两种主要候选LASV huMAb或huMAb组合进行挑战治疗剂量发现研究，通过最佳给药途径进行单剂量给药。接下来，我们将在良好实验室规范（GLP） GP研究中，通过最佳确定的给药途径，确定LASV huMAb或huMAb组合在致死性感染时间轴上的治疗效果。GP研究的结果将指导在GLP非人灵长类动物（NHP）研究中，单次静脉、腹腔或皮下给药后，评估两种单LASV huMAb （IgG1或IgG4）或huMAb组合的药代动力学。我们还将在GLP NHP研究中进行LASV huMAb或huMAb联合治疗的疗效研究，在致死性感染的整个过程中，在感染后的不同时间，通过最佳给药途径进行两种剂量的治疗。这一提议的努力将产生一类新的

DESCRIPTION (provided by applicant): This project will develop Immunotherapeutics for Biodefense against Lassa virus (LASV), an arenavirus that is the causative agent of Lassa fever (LF). LASV is a Biosafety Level 4 and NIAID Biodefense category A agent. LF is a major public health concern causing widespread loss of life and social disruption across West Africa. The goal of this proposal is to perform critical steps in the characterization and pre-clinical development of a new class of therapeutics to treat LF. This effort combines the expertise of virologists and immunologists at Tulane University and the University of Texas Medical Branch with scientists at Zalgen, a biotechnology company specialized in the development of immunotherapeutics, rapid diagnostics, and new vaccine platforms for emerging viral diseases with global impact. Studies proposed here will leverage our recent advances, including identification of lead candidate human monoclonal antibodies (huMAbs) that neutralize LASV infectivity and protect guinea pigs against fatal disease induced by LASV. Zalgen's exclusive technology for rapid development (<3 months) of high yield antibody expressing stable cell lines ensures that costs of producing therapeutic antibodies is minimized for primary Biodefense markets and secondary Public Health markets. In studies proposed in MILESTONE 1 we will generate multi-milligram quantities of IgG1 and IgG4 LASV huMAb subtypes in serum-free medium adapted mammalian cell transient expression platforms for purification to perform in vitro testing. We will also perform a panel of characterization assays with the huMAbs focusing on identification of a huMAb or combinations of huMAbs with broad LASV neutralization potential and low IC50. Additional studies will establish cross-reactivity profiles with related Ne and Old World arenaviral proteins and reactivity of LASV huMAbs with human cell membrane phospholipids. In MILESTONE 2 we will generate CHO or NS0 stable cell lines in chemically defined serum free medium and select high producing isolates. Limiting dilution cell cloning (LDCC) will be performed for isolation of stable, production grade CD-SFM-CHO or CD-SFM-NS0 clones. We also plan to generate Accession Cell Banks (ACB) for clonal cell lines of interest, and to convert to manufacturing with QA/QC to generate gram quantities of candidate huMAbs for in vivo studies. In MILESTONE 3 we will perform evaluation of pharmacokinetics of LASV huMAb subtypes in guinea pigs (GPs). We will also perform challenge - therapeutic dose finding studies with two lead candidate LASV huMAbs or huMAb combinations in GPs, following a single dose via optimal delivery route. Next, we will determine the therapeutic efficacy with LASV huMAbs or huMAb combinations in Good Laboratory Practice (GLP) GP studies throughout the course of a lethal infection timeline, via the optimally determined route of administration. Results of the GP studies will guide an evaluation of pharmacokinetics of two single LASV huMAbs (IgG1 or IgG4) or huMAb combinations in a GLP nonhuman primate (NHP) study, following a single intravenous, intraperitoneal, or subcutaneous dose. We will also perform therapeutic efficacy studies with LASV huMAbs or huMAb combinations in GLP NHP studies throughout the course of a lethal infection timeline, following two doses via optimal delivery route, at varying times post infection. This proposed effort will result in a new class of immunotherapeutics for LF.