Mazurier F, Moreau-Gaudry F, Salesse S, Barbot C, Ged C, Reiffers J, de Verneuil H
Laboratoire de Pathologie Moléculaire et Thérapie Génique, Université Victor Ségalen Bordeaux 2, France.
J Inherit Metab Dis. 1997 Jun;20(2):247-57. doi: 10.1023/a:1005365008147.
Congenital erythropoietic porphyria (CEP) is an inherited metabolic disorder characterized by an overproduction and accumulation of porphyrins in bone marrow. This autosomal recessive disease results from a deficiency of uroporphyrinogen III synthase (UROIIIS), the fourth enzyme of the haem biosynthetic pathway. It is phenotypically heterogeneous: patients with mild disease have cutaneous involvement, while more severely affected patients are transfusion dependent. The cloning of UROIIIS cDNA and genomic DNA has allowed the molecular characterization of the genetic defect in a number of families. To date, 22 different mutations have been characterized. Allogeneic bone marrow transplantation is the only curative treatment available for the severe, transfusion-dependent, cases. When bone marrow transplantation cannot be performed owing to the absence of a suitable donor, the autografting of genetically modified cells is an appealing alternative. The best approach to somatic gene therapy in this disease involves the use of recombinant retroviral vectors to transduce cells ex vivo, followed by autologous transplantation of the genetically modified cells. We investigated retroviral transfer in deficient human fibroblasts, immortalized lymphoblasts as well as bone marrow cells, and obtained a complete restoration of the enzymatic activity and full metabolic correction. Using K562 cells, an erythroleukaemic cell line, the expression of the transgene remained stable during 3 months and during erythroid differentiation of the cells. Finally, a 1.6- to 1.9-fold increase in enzyme activity compared to the endogenous level was found in normal CD34+ cells, a population of heterogeneous cells known to contain the progenitor/stem cells for long-term expression. The future availability of a mouse model of the disease will permit ex vivo gene therapy experiments on the entire animal.
先天性红细胞生成性卟啉病（CEP）是一种遗传性代谢紊乱疾病，其特征是骨髓中卟啉过度产生和积累。这种常染色体隐性疾病是由于血红素生物合成途径的第四个酶——尿卟啉原III合成酶（UROIIIS）缺乏所致。它在表型上具有异质性：轻症患者有皮肤受累，而病情较重的患者则依赖输血。UROIIIS cDNA和基因组DNA的克隆使得许多家族中遗传缺陷的分子特征得以明确。迄今为止，已鉴定出22种不同的突变。同种异体骨髓移植是严重的、依赖输血的病例唯一可用的治愈性治疗方法。当由于缺乏合适的供体而无法进行骨髓移植时，基因改造细胞的自体移植是一种有吸引力的替代方法。在这种疾病中，体细胞基因治疗的最佳方法是使用重组逆转录病毒载体在体外转导细胞，然后对基因改造细胞进行自体移植。我们研究了逆转录病毒在缺陷的人成纤维细胞、永生化淋巴细胞以及骨髓细胞中的转移情况，并实现了酶活性的完全恢复和全面的代谢纠正。使用红白血病细胞系K562细胞，转基因的表达在3个月内以及细胞的红系分化过程中保持稳定。最后，在正常CD34 +细胞中发现酶活性比内源性水平增加了1.6至1.9倍，CD34 +细胞是一群已知含有长期表达祖细胞/干细胞的异质性细胞。该疾病小鼠模型未来的可用性将允许在整个动物身上进行体外基因治疗实验。
