Defining the therapeutic efficacy, tolerogenic potential, and genotoxicity of liver-targeted AAV gene therapy for hemophilia A

定义肝脏靶向 AAV 基因疗法治疗血友病 A 的疗效、耐受潜力和遗传毒性

• 批准号: 10704568
• 负责人: Graca Duarte Almeida-Porada
• 金额: $ 67.39万
• 依托单位: WAKE FOREST UNIVERSITY HEALTH SCIENCES
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-09-15 至 2026-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10704568
• 关键词: Address; Adult; Animal Model; Biological; Birth; Blood Coagulation Disorders; Blood Coagulation Factor; Capsid; Cell Proliferation; Cells; Clinical; Clinical Trials; Clonal Expansion; Cross Presentation; Data; Dose; Early treatment; Enzyme-Linked Immunosorbent Assay; Event; Exhibits; Exposure to; F8 gene; Fetal Development; Fetal Sheep; Fetus; Fibrosis; Gene Transfer; Gene therapy trial; Genetic Diseases; Genome; Genomics; Goals; Hemophilia A; Hemorrhage; Hepatocyte; Hepatotoxicity; Human; Immune; Immune Tolerance; Immune response; Immunity; Immunologics; Incidence; Inflammatory Response; Inherited; Innate Immune Response; Intrauterine Blood Transfusion; Life; Liver; Location; Mediating; Neonatal; Newborn Infant; Onset of illness; Organ; Patients; Plasma; Primary carcinoma of the liver cells; Production; Proteins; Recording of previous events; Risk; Safety; Serotyping; Sheep; System; Testing; Therapeutic; Thymic epithelial cell; Tissues; Transgenes; Treatment Efficacy; adeno-associated viral vector; biological adaptation to stress; cell type; clinically relevant; cytokine; fetal; gene therapy; gene therapy clinical trial; genomic locus; genotoxicity; in utero; in utero transplantation; in vivo; inhibitor; liver function; liver inflammation; manufacturing run; minimal risk; neonatal period; neonate; neutralizing antibody; participant enrollment; postnatal; pre-clinical; prenatal; prenatal exposure; prevent; safety testing; tissue tropism; transgene expression; tumorigenesis; vector; vector genome; vector induced

PROJECT SUMMARY: Gene therapy (GT) for the hemophilias is poised to fulfill the promise of a safe, affordable, lifelong correction following a single treatment. Yet clinical trials employing AAV vectors to treat adult hemophilia A (HA) patients have underscored the risk of hepatoxicity and genotoxicity in adult patients receiving GT. In addition, the use of AAV-based GT is prevented by pre-existing AAV immunity in a high percentage of patients. While FVIII inhibitors have not yet been seen in GT trials, inhibitors usually appear within the first 50 exposure days to FVIII, and since all enrolled patients were selected based upon an inhibitor-free clinical history, it is likely that the patients were not “inhibitor-prone”, leaving unanswered the critical question of whether GT will avoid inhibitor induction. GT to treat HA at birth provides the advantage of correcting bleeding events sooner, but whether neonatal GT would avoid FVIII inhibitor induction and hepatic inflammation is, at present, unknown. Recent studies have shown a 59% incidence of neutralizing antibodies to AAV at birth. Thus, treatment during the neonatal period will likely be hampered by anti-AAV immunity. One approach that could circumvent both AAV- and FVIII-related immune hurdles while enabling correction prior to disease onset is in utero GT (IUGT). Using the same preclinical animal model (sheep) used to delineate conditions to cure SCID patients prenatally, we showed that the highly proliferative state and relative immune-immaturity of the fetus can be exploited to achieve clinically relevant levels of gene transfer to multiple tissues and induce durable tolerance to vector- encoded proteins. Besides its inherent advantages as a GT recipient, its rapid cellular turnover makes the fetus a highly sensitive system in which to define the potential genotoxicity and risk of tumorigenesis following AAV GT. This rapid cellular proliferation also provides a rigorous testbed to define the incidence and locations of AAV genomic integration, determine the duration of AAV persistence, and identify the optimal target cell(s) within the liver to achieve long-term expression of a fVIII transgene and resultant therapeutic benefit. The overall goal of this proposal is to use fetal sheep to address critical unanswered biological and safety questions that plague liver-targeted AAV-based GT for HA and test the hypothesis that performing AAV-based IUGT will overcome the immune hurdles inherent to postnatal GT, transduce sufficient numbers of the appropriate hepatic cells to mediate long-term therapeutic plasma FVIII levels, and induce durable tolerance to FVIII. We propose 3 Aims: i) Define the in vivo tissue tropism of two AAV serotypes in use in clinical trials for HA (AAV5 and AAV3B) following administration into fetal and neonatal recipients; ii) Define the impact of fVIII-encoding AAV transduction on fetal vs. neonatal liver function; and iii) Determine the duration of fVIII-AAV persistence and fVIII transgene expression, and whether durable immune tolerance is induced to FVIII and/or AAV following in utero vs. neonatal GT. We hope that these studies will elucidate whether IUGT or neonatal GT for HA will be safe, and whether or not the therapy will be long-lasting, prevent breakthrough bleeds, and avoid FVIII immune responses.

项目摘要：血友病的基因治疗(GT)有望实现安全、负担得起、 一次治疗后终生矫正。然而，使用AAV载体治疗成人血友病的临床试验 A(HA)患者强调了接受GT的成年患者的肝毒性和遗传毒性的风险。在……里面 此外，在很高比例的患者中，预先存在的AAV免疫阻止了基于AAV的GT的使用。 虽然还没有在GT试验中看到FVIII抑制剂，但抑制剂通常出现在前50次接触中 到FVIII的天数，由于所有入选的患者都是根据无抑制物的临床病史选择的，很可能 这些患者并不是“易受抑制者”，这使得GT能否避免这个关键问题没有得到解答。 抑制物诱导。GT在出生时治疗HA提供了更快纠正出血事件的优势，但 新生儿GT是否能避免FVIII抑制剂的诱导和肝脏炎症，目前尚不清楚。 最近的研究表明，出生时AAV中和抗体的发生率为59%。因此，在治疗过程中 新生儿期可能会受到抗AAV免疫的阻碍。一种方法可以绕过这两种方法 AAV和FVIII相关的免疫障碍在发病前得以纠正是在宫内GT(IUGT)。 使用相同的临床前动物模型(绵羊)来描述产前治愈SCID患者的情况， 我们表明，胎儿的高度增殖状态和相对免疫不成熟可以被利用来 达到临床相关水平的基因转移到多个组织，并诱导对载体的持久耐受- 编码的蛋白质。除了作为GT接受者的固有优势外，它快速的细胞周转使胎儿 一种高度敏感的系统，用于确定AAV后的潜在遗传毒性和肿瘤形成风险 GT.这种快速的细胞增殖也为确定AAV的发病率和位置提供了严格的试验台 基因组整合，确定甲型肝炎病毒持续时间，并确定最优靶细胞(S) 肝脏实现FVIII转基因的长期表达和由此产生的治疗益处。的总目标是 这项提议是利用胚胎绵羊来解决困扰人类的关键的生物和安全问题。 肝靶向AAV为基础的GT治疗HA并检验实施AAV为基础的IUGT将克服的假设 出生后GT固有的免疫障碍，将足够数量的适当肝细胞转导到 调节长期治疗性血浆FVIII水平，并诱导对FVIII的持久耐受。我们提出了三个目标：i) 确定甲型肝炎临床试验中使用的两种AAV血清型(AAV5和AAV3B)的体内组织趋向性 对胎儿和新生儿受体的给药；ii)确定编码FVIII的AAV转导对胎儿的影响 与新生儿肝功能的对比；以及iii)确定FVIII-AAV持续时间和FVIII转基因 表达，以及在子宫和新生儿中是否诱导了对FVIII和/或AAV的持久免疫耐受 GT.我们希望这些研究将阐明用于HA的IUGT或新生儿GT是否安全，以及是否 不治疗将是持久的，防止突破性出血，并避免FVIII免疫反应。