HEPATOCYTE GENE THERAPY FOR CITRULLINEMIA

瓜氨酸血症的肝细胞基因治疗

• 批准号: 6434947
• 负责人: Brendan Lee
• 金额: $ 13.19万
• 依托单位: BAYLOR COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-03-09 至 2001-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6434947
• 关键词: Adenoviridae; alpha 1 antitrypsin; ammonia; antigen presenting cell; arginosuccinate synthetase deficiency; blood chemistry; chemical kinetics; citrulline; cow; disease /disorder model; dosage; gene expression; gene therapy; human morbidity; immune tolerance /unresponsiveness; immunocytochemistry; laboratory mouse; liver; liver cells; neutralizing antibody; newborn animals; nonhuman therapy evaluation; reporter genes

The long-term goal of this proposal is to correct an inborn error of metabolism, citrullinemia, by hepatocyte gene therapy using the adenoviral delivery system. Citrullinemia, an autosomal recessive enzymatic deficiency of argininosuccinate synthetase (ASS), is representative of the group of urea cycle defects. They present in the neonatal period with hyperammonemic coma and result in long-term neurological and developmental deficits. Plasma, metabolites, ammonia, and citrulline can be easily quantitated to monitor the efficacy of therapeutic intervention, and both murine and bovine models are available. Protocols derived from this study are expected to be applicable to other genetic disorders characterized by primary metabolic dysfunction of hepatocytes. Transient correction of the ASS deficiency will first be attempted by in vivo transduction of hepatocytes after intravenous delivery of an E1-deleted type 5 adenovirus (Ad-ASS) into a neonatal murine model. Once the optimal dose and pattern of delivery is determined and efficacy is established by direct enzymatic assay of liver isolates and by achieving normalized growth rates and prolonged survival, the protocol will be scaled up and applied to the larger bovine model. At first, a recombinant adenovirus (Ad-AAT) expressing a serum reported gene product, alpha-antitrypsin (alpha1AT), will be delivered intravenously into normal neonatal calves. The extent, efficacy, and kinetics of heterologous gene expression and morbidity associated with infection will be determined by serial measurements of serum alpha1AT, adenovirus specific neutralizing antibodies, blood chemistries, and immuno-histologic analysis of liver isolates. Subsequently, Ad-ASS will be delivered into citrullinemic newborn calves using optimized dosage and delivery protocols. Once transient correction of the metabolic defect is demonstrated in the bovine model, human trials will be proposed. Even a transient correction may abbreviate the hyperammonemic state and reduce long-term central nervous system morbidity. Concurrent studies directed at minimizing the role of host immune response in terminating the initial period of heterologous gene expression and in limiting subsequent reinfection will be carried out in the bovine model. The ability to induce immune tolerance will be investigated by delivering the reporter construct Ad-AAT in the neonatal and in utero period. In addition, new adenoviral vectors mutated in the E2a and E4 domains and designed to minimize adenoviral protein expression and host antigen presentation will be tested.

该提议的长期目标是纠正原始错误 代谢，瓜氨酸血症，通过肝细胞基因治疗使用腺病毒 交付系统。 瓜氨酸血症，一种常染色体隐性酶促酶促 精氨酸偶联合成酶（ASS）的缺乏代表 一组尿素周期缺陷。 他们出现在新生儿时期 高症昏迷，导致长期神经和发育 缺陷。 血浆，代谢产物，氨和瓜氨酸可以很容易被 定量以监测治疗干预的功效，均 可以使用鼠和牛型号。 从这项研究得出的协议 预计将适用于其他遗传疾病的特征 肝细胞的原代代谢功能障碍。 瞬态校正 屁股缺乏将首先通过体内转导 静脉输送E1删除5型腺病毒后的肝细胞 （ad-ass）进入新生儿鼠模型。 一旦最佳剂量和图案 确定输送的确定，并通过直接酶促确定功效 通过达到归一化的生长速率和 长时间的生存期，该方案将缩放并应用于 较大的牛模型。 首先，重组腺病毒（AD-AAT） 表达血清报道的基因产物α-抗胰蛋白酶（alpha1at）， 将静脉注射到正常的新生儿犊牛中。 程度， 异源基因表达和发病率的功效和动力学 与感染相关的将由串行测量确定 血清α1AT，腺病毒特异性中和抗体，血液 化学和肝分离株的免疫历史分析。 随后，Ad-Ass将被交付到Citrullinmemic Newborn Calves中 使用优化的剂量和输送协议。 一旦短暂校正 在牛模型中证明了代谢缺陷的人类试验 将提出。 即使是短暂的校正也可能缩写 高ammonememementem并降低长期中枢神经系统 发病率。 并发研究旨在最大程度地减少宿主的作用 终止异源基因的初始时期的免疫反应 表达和限制后随后的再感染将在 牛模型。 诱导免疫耐受性的能力将是 通过在新生儿中交付记者构造来调查 在子宫时期。 此外，新的腺病毒载体在 E2A和E4结构域，旨在最小化腺病毒蛋白表达 将测试主机抗原表现。