Foamy Viral Gene Therapy for X-linked Severe Combined Immune Deficiency

描述(由申请人提供)：我们提出了一个基础和翻译研究的综合计划，以开发基于泡沫病毒(FV)载体的基因替代疗法，用于患有遗传性严重联合免疫缺陷SCID-X1的患者。该计划涉及三个科学项目和四个核心。其中两个科学项目的重点是利用动物模型研究FV载体SCID-X1的基因治疗：项目1-针对小鼠和人SCID-X1的FV基因治疗的临床前建模，以及项目2-FV介导的犬SCID-X1模型的基因治疗。这些项目将测试有关FV载体与先进的非清髓性调节方案联合应用于？C基因替代治疗的安全性和有效性的关键假设。另一个科学项目，项目3-第二代FV载体SCID-X1基因治疗，将测试与FV载体系统和技术相关的关键假设，目标是进一步提高FV载体的安全性和性能。这三个项目将由四个核心提供支助，以有效利用共同的方法和技术。在五年的支持期内，该计划旨在为SC1D-X1产生一种集成的基因替代疗法，该疗法由具有良好特性的第一代临床C抗体载体和完全准备好转化为人类SCID-X1临床试验的先进调节方案组成。此外，该计划将增进我们对FV载体系统和FV载体技术的了解；并启动第二代FV载体系统和综合治疗的临床前评估，预计将导致未来在安全性和有效性方面的进一步进步。总体而言，我们的工作有望带来安全和新颖的未来治疗方法，包括直接体内FV载体基因传递的有效方法， 人类SCID-X1以及其他遗传性疾病所需的。 相关性：该项目将为患有灾难性免疫缺陷SC1D-X1的患者创造一种新型的基因疗法。它还将在基因治疗和干细胞移植方面产生进展，有可能导致对许多其他类型的血液和免疫系统遗传性疾病的更安全和更有效的治疗。 项目1-泡沫病毒基因治疗的临床前建模 项目负责人(PL)：戴维J.罗林斯 (申请人提供的描述)：SCID-X1是一种由常见伽马链(？C)基因突变引起的灾难性免疫缺陷疾病。虽然使用匹配的兄弟姐妹捐赠者进行干细胞移植可以治愈，但大多数患者缺乏理想的捐赠者，导致结果较差。基因替代作为SCID-X1的一种替代治疗方法有许多理论上的优势；使用伽玛逆转录病毒C进行的开创性临床研究既带来了显著的好处，也由于病毒增强剂引发的白血病而导致了意想不到的不良事件。这种PPG的主要假设是，使用基于重组泡沫病毒(FV)的载体可以显著提高？C基因的有效性和安全性。项目1中的研究旨在检验这样的假设，即不含病毒增强剂的FV载体(在转录盒两侧有或没有额外的增强子阻断元件)将显示出足以在体内进行功能挽救的转基因表达水平，同时显示出较低的遗传毒性。项目1的目的是通过详细的表型、功能和分子分析来验证这些假设：1)SCID-X1的小动物模型和2)来自SCID-X1患者的造血干细胞(HSC)。我们的具体研究将包括：1)EF1？-Hu-？C Fv载体在去髓和非去髓小鼠SCID-X1受者体内的有效性和安全性评估；2)在SCID-X1患者CD34+BM细胞中转导的临床前和GMP级第一代？C Fv；以及3)候选绝缘的第二代？C Fv载体在SCID-X1小鼠和人类患者的HSC中的有效性和安全性评估。项目1将利用所有4个核心，并将在多个层面上与项目2和项目3的工作对接。结合项目2和项目3的数据，我们的研究将为未来的第一代和第二代SCID-X1 FV载体临床试验提供有关有效性、安全性和最佳载体设计的关键数据。 相关性：SCID-X1是一种由？C基因突变引起的严重免疫疾病。预计基因治疗将为SCID-X1提供一种有益的治疗方法。然而，以前的方法虽然有益，但也会导致高频率的不良事件。该PPG的目标是利用基于重组泡沫病毒(FV)的载体开发安全有效的SCID-X1基因载体。项目1中的研究旨在测试候选临床SCID-X1FV载体将在小鼠和人类模型中挽救SCID-X1功能的想法，同时显示出降低的遗传毒性。

DESCRIPTION (provided by applicant): We propose an integrated program of basic and translational research to develop foamy virus (FV) vector- based gene replacement therapy for patients who suffer from the inherited severe combined immunodeficiency SCID-X1. The program involves three scientific Projects and four Cores. Two of the scientific projects are focused on studying FV vector SCID-X1 gene therapy using animal models: Project 1 - Pre-clinical modeling of FV gene therapy for murine and human SCID-X1, and Project 2 - FV mediated gene therapy in the canine SCID-X1 model. These projects will test key hypotheses regarding the safety and efficacy of FV vectors applied in conjunction with advanced non-myeloablative conditioning regimens for ?c gene replacement therapy. The other scientific project, Project 3 - Second generation approaches to FV vector SCID-X1 gene therapy, will test key hypotheses related to FV vector systems and technology, with the goal of further enhancing FV vector safety and performance. The three Projects will be supported by four Cores to provide for efficient use of common methods and technology. Over the five year period of support, the program is designed to yield an integrated gene replacement therapy for SC1D-X1 consisting of a well characterized 1st generation clinical ?c FV vector and an advanced conditioning regimen that is fully ready for translation to a human SCID-X1 clinical trial. In addition, the program wil advance our knowledge of FV vector systems and FV vector technology; and initiate pre-clinical evaluation of 2nd generation ?c FV vector systems and integrated therapies anticipated to lead to future further advances in safety and efficacy. Overall, our work is predicted to lead to safe and novel future therapies, including efficient methods for direct in vivo FV vector gene delivery, needed for human SCID-X1 as well other genetic disorders. RELEVANCE: This project will create a new type of gene therapy for patients who suffer from the catastrophic immunodeficiency SC1D-X1. It will also generate advances in gene therapy and stem cell transplantation that have the potential to lead to safer and more effective therapies for many other types of inherited diseases of the blood and immune system. PROJECT 1 - Pre-clinical Modeling of Foamy Viral Gene Therapy Project Leader (PL): David J. Rawlings (Description as provided by applicant): SCID-X1 is catastrophic immunodeficiency disorder caused by mutations within the common gamma chain (?c) gene. While stem cell transplantation using a matched sibling donor can be curative, most patients lack optimal donors leading to poorer outcomes. Gene replacement has many theoretical advantages as an alternative therapeutic approach for SCID-X1; and pioneering clinical studies using gammaretroviral ?c delivery lead to both significant benefit as well as unanticipated adverse events due to viral enhancer triggered leukemogenesis. The overarching hypothesis of this PPG is that both the efficacy and safety of ?c gene delivery can be significantly improved using recombinant foamy virus (FV) based vectors. Studies in Project 1 are designed to test the hypotheses that ?c FV vectors devoid of viral enhancers (with or without additional enhancer blocking elements flanking the transcriptional cassette) will exhibit levels of transgene expression sufficient for functional rescue in vivo while concurrently showing reduced genotoxicity. The aims of Project 1 are designed to test these hypotheses via detailed phenotypic, functional, and molecular analysis in both: 1) a small animal model of SCID-X1 and 2) hematopoietic stem cells (HSC) derived from SCID-X1 patients. Our specific studies will include efficacy and safety assessment of 1) EF1?-hu-?c FV vectors in vivo in myeloablated vs. non-myeloablated murine SCID-X1 recipients; and in alternative in vitro transactivation assays; 2) Preclinical and GMP-grade 1st generation ?c FV in transduced SCID-X1 patient CD34+ BM cells; and 3) Candidate insulated 2nd generation ?c FV vectors in HSC from SCID-X1 mice and human patients. Project 1 will utilize all 4 Cores and will interface on multiple levels with work within both Projects 2 and 3. In conjunction with data derived from Projects 2 and 3, our studies will provide key data regarding efficacy, safety and optimal vector design for future 1st and 2nd generation SCID-X1 FV vector clinical trials. RELEVANCE: SCID-X1 is severe immune disorder caused by mutations in the ?c gene. Gene therapy is predicted to provide a beneficial treatment for SCID-X1. However, previous approaches, while beneficial, also lead to a high frequency of adverse events. The goal of this PPG is develop safe and effective SCID-X1 gene delivery using recombinant foamy virus (FV) based vectors. Studies in Project 1 are designed to test the idea that candidate clinical SCID-X1 FV vectors will rescue function in mouse and human models of SCID-X1 while concurrently showing reduced genotoxicity.