Gene therapy of urea cycle deficirncy

尿素循环缺陷的基因治疗

• 批准号: 06454610
• 负责人: MATSUDA Ichiro
• 金额: $ 4.54万
• 依托单位: Kumamoto University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for General Scientific Research (B)
• 财政年份: 1994
• 资助国家: 日本
• 起止时间: 1994 至 1995
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-06454610/
• 关键词: Gene therapy; Urea cycle; Ornithine transcarbamylase; Adenovirus vector; OTC欠損症; CAGプロモーター; spf^<ash>マウス

Ornithine transcarbamylase (OTC) deficiency, the most commom and severe inborn error of the urea cycle in humans, remains without adequate treatment, and ortality rates are high. Adenoviral vectors provide an efficient system for gene delivery, but there are problems, including toxicity. Efficient promoters that reduce the amount of vector required for treatment need to be developed. We constructed two recombinant adenoviral vectors, AdexCAGhOTC and AdexSRalphahOTC,which harbor the human OTC gene under transcriptional control of CAG (a modified chicken beta-actin promoter with CMV-IE enhancer) and SRalpha (the SV 40 early prommoter with the R segment and part of the U5 segment of the HTLV-1 LTR) , respectively. Each was tested in adult spf^<ash> mice, an animal model of human OTC deficiency, and in primary human hepatocytes with OTC deficiency. Spf^<ash> mice have a pronounced orotic aciduria as seen in humans. A complete recovery of hepatic OTC activity with minimal tissue damage was observed in these animals following the intravenous administration of AdexCAGhOTC alone. Western blot analysis confirmed hepatic OTC expression and normalization of orotic aciduria was evident for 60 days. Enzyme activities of primary human hepatocytes infected with AdexCAGhOTC were 10-40 times higher than those with AdexSRalphahOTC.Thus, the adenoviral vector with an efficient promoter such as CAG,can be given further considerartion for possible gene therapy in humans with OTC deficiency.

鸟氨酸经钙化酶（OTC）缺乏症，人类尿素周期中最常见和最严重的天生误差，没有适当的治疗，并且正常率很高。腺病毒载体为基因递送提供了有效的系统，但是存在问题，包括毒性。需要开发减少治疗所需载体量的有效启动子。我们构建了两个重组腺病毒载体，Adexcaghotc和Adexsralphahotc，它们在CAG的转录控制下（带有CMV-IE增强剂的改性鸡β-肌动蛋白启动子）和Sralpha（SV 40的早期前推者）（SV 40早期引起者与R r s sember of R r s eft of U5和u5）。每个人都在成年SPF^<ash>小鼠（人类OTC缺乏症的动物模型）和OTC缺乏症的原发性肝细胞中进行了测试。 SPF^<ash>小鼠具有人类所见的明显果酸。单独静脉内给药后，这些动物在这些动物中观察到了肝脏OTC活性的完全恢复，仅在这些动物中观察到了最小的肝脏活性。蛋白质印迹分析证实了肝脏的OTC表达和牛毒尿尿液的归一化60天。经adexcaghotc感染的原代人肝细胞的酶活性比具有Adexsralphahotc的患者高10-40倍。因此，可以进一步考虑具有有效启动子（例如CAG）的腺病毒载体，以进一步考虑OTC缺乏症的人类可能的基因治疗。

项目成果

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“使用重组腺病毒载体的神经细胞类型特异性表达系统。”

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