Gene therapy of urea cycle deficirncy

鸟氨酸氨基转移酶(OTC)缺乏症是人类尿素循环中最常见和最严重的先天性错误，至今仍未得到适当的治疗，病死率很高。腺病毒载体为基因传递提供了一种有效的系统，但也存在包括毒性在内的问题。需要开发有效的启动子来减少治疗所需的传播量。我们构建了两个重组腺病毒载体AdexCAGhOTC和AdexSRalphahOTC，它们分别在CAG(带有CMV-IE增强子的修饰的鸡β-肌动蛋白启动子)和SRpha(带有HTLV-1LTR的R片段和部分U5片段的SV 40早期启动子)转录控制下携带人OTC基因。每一种方法都在成年SPF^&lt；ash&gt；小鼠(一种人类OTC缺乏症的动物模型)和原代人类OTC缺乏症肝细胞中进行了测试。像人类一样，SPF；老鼠也有明显的口腔性酸尿。在这些动物中，观察到仅静脉注射AdexCAGhOTC后，肝脏OTC活性完全恢复，组织损伤最小。免疫印迹分析证实肝脏OTC表达，60天后口腔性酸尿明显恢复正常。感染AdexCAGhOTC的原代人肝细胞的酶活性比感染AdexSRalphahOTC的高10-40倍。因此，带有CAG等有效启动子的腺病毒载体可进一步考虑用于OTC缺乏症的基因治疗。

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.