Macrophage Based Gene Therapy for Hereditary Pulmonary Alveolar Proteinosis

描述（由申请方提供）：遗传性肺泡蛋白沉积症（hPAP）是一种表面活性物质蓄积增加导致呼吸衰竭的疾病，目前尚无药物治疗。它是由分别编码GM-CSF受体（GM-R）α和β亚基的CSF 2 RA或CSF 2 RB突变破坏GM-CSF向肺泡巨噬细胞（AM）的信号传导引起的。长期目标是开发基因治疗以恢复AM中的GM-CSF信号传导，从而恢复GM-CSF依赖性AM功能，这些功能对于表面活性剂清除、肺泡稳态、肺功能和宿主防御至关重要。目的是评价一种新的治疗方法，肺巨噬细胞移植（PMT）。初步数据显示，GM-R缺陷（GM-RKO）小鼠发生与人类hPAP相同的hPAP肺病，包括肺GM-CSF水平升高，这赋予具有功能性GM-R的AM选择性存活优势。中心假设是，安全性增强的慢病毒载体介导的造血干细胞/前体细胞（HSPC）中的GM-R表达，扩增成巨噬细胞，以及基因校正细胞的自体PMT（无骨髓消融）作为hPAP的治疗将是有效和安全的。初步数据显示，单次PMT治疗可以在GM-RKO小鼠中校正hPAP至少一年（评估的最长时间）而没有相关的不良事件，并且人HSPC可以在申请人的实验室中使用完善的方法容易地转导并扩增成表达功能性GM-R的巨噬细胞。理由是预期结果将为未来临床试验的设计提供信息，并提供临床前安全性和有效性数据，以及GMP 获得监管批准以在人体中测试该方法所需的制造程序和验证数据。将在四个具体目标中测试该假设：1）确定GM-RKO小鼠中hPAP的基因转移/PMT疗法的效率和动力学; 2）优化具有最大PMT植入潜力的人HSPC衍生的基因校正的巨噬细胞的扩增; 3）在与基因转移、来自HSPC的巨噬细胞扩增、PMT、GM-RKO小鼠中转导细胞的药理学消耗; 4）开发和验证用于生产HSPC衍生的基因校正的巨噬细胞的方案，并编写用于hPAP的基因转移/PMT治疗的临床方案和研究性新药申请。该方法是创新性的，因为它明显不同于目前通过全肺灌洗物理去除表面活性剂的低效、高侵入性方法，而是使用一种新的方法来恢复AM功能。这项拟议的研究意义重大，因为它有望建立一种针对hPAP儿童的特异性疗法和一种可能对其他疾病有用的新型疗法（PMT）的可行性，并评估多种安全性改进以降低基因治疗的风险。这些结果将为GM-CSF调节AM群体大小的基本机制提供信息，提供AM寿命的估计，并为基于巨噬细胞的治疗的发展奠定基础。

DESCRIPTION (provided by applicant): Hereditary pulmonary alveolar proteinosis (hPAP) is a disorder of increased surfactant accumulation resulting in respiratory failure for which no pharmacologic therapy exists. It is caused by disruption of GM-CSF signaling to alveolar macrophages (AMs) by mutations in CSF2RA or CSF2RB, which encode GM-CSF receptor (GM-R) α and β subunits, respectively. The long-term goal is to develop gene therapy to restore GM-CSF signaling in AMs and, thereby, GM-CSF-dependent AM functions critical to surfactant clearance, alveolar homeostasis, lung function and host defense. The objective here is to evaluate a novel therapeutic approach, pulmonary macrophage transplantation (PMT). Preliminary data show that GM-R deficient (GM-RKO) mice develop hPAP lung disease that is identical to hPAP in humans including an increased level of pulmonary GM-CSF, which confers a selective survival advantage to AMs with functional GM-Rs. The central hypothesis is that safety-enhanced, lentiviral vector-mediated GM-R expression in hematopoietic stem/precursor cells (HSPCs), expansion into macrophages, and autologous PMT of gene-corrected cells without myeloablation will be effective and safe as therapy of hPAP. Preliminary data show that a single PMT treatment can correct hPAP in GM-RKO mice for at least one year (the longest time evaluated) without associated adverse events, and that human HSPCs can be readily transduced using well-established methods in the applicants' laboratory and expanded into macrophages expressing functional GM-Rs. The rationale is that anticipated results will inform the design of future a clinical trial and provide the preclinical safety and efficacy data, and GMP manufacturing procedures and validation data required to obtain regulatory approval to test this approach in humans. The hypothesis will be tested in four specific aims: 1) determine the efficiency and kinetics of gene transfer/PMT therapy of hPAP in GM-RKO mice; 2) optimize the expansion of human HSPC-derived, gene- corrected macrophages with maximum PMT engraftment potential; 3) determine the safety of gene transfer/PMT in preclinical studies related to gene transfer, macrophage expansion from HSPCs, PMT, and pharmacologic depletion of transduced cells in GM-RKO mice; 4) develop and validate protocols for the manufacture of HSPC-derived, gene-corrected macrophages, and write a clinical protocol and investigational new drug application for gene transfer/PMT therapy of hPAP. The approach is innovative because it departs markedly from the current inefficient, highly invasive method of physically removing surfactant by whole lung lavage and in- stead uses a novel approach to restore AM function. The proposed research is significant because it is expected to establish the feasibility of a specific therapy for children with hPAP and a new type of therapy (PMT) that may be useful for other diseases, and evaluate multiple safety improvements to reduce the risks of gene therapy. Results will inform a fundamental mechanism by which GM-CSF regulates AM population size, pro- vide an estimate of AM lifespan, and lay the foundation for the development of macrophage-based therapy.