α- L-iduronidase gene-based therapy using the phiC31 system to treat mucopolysaccharidose type I mice.

BACKGROUND: Mucopolysaccharidose type I (MPSI) is a lysosomal monogenic disease caused by mutations in the gene for α- L-iduronidase (IDUA). MPSI patients need a constant supply of IDUA to alleviate progression of the disease. IDUA gene transfer using integrative vectors might provide a definitive solution and support advancement to clinical trials, although studies have not yet been satisfactory. To achieve a stable IDUA gene expression in vivo, phiC31 was tested in the present study. METHODS: Several plasmid vectors were constructed and IDUA-/- mice were treated with cyclophosphamide and transfected with these vectors hydrodynamically via tail veins. IDUA expression was monitored over time. Treated and nontreated mice underwent an open-field test at age 8 months, and IDUA activity and glycosaminoglycan (GAG) content of tissues were evaluated. RESULTS: High levels of IDUA activity were detected initially (>1000 U/ml), although these levels decayed over time. The reinjection of vectors produced a similar profile of IDUA decay. Three out of six treated mice had IDUA activity in the livers, and also showed lower GAG content, reduced lysosomes and better locomotion. To investigate unsustained IDUA production, wild-type mice were submitted to the same gene therapy procedure, which generated a similar profile of IDUA decay. Anti-IDUA antibody was detected in the sera of these animals. In addition, we also found three methylated sites in the cytomegalovirus promoter region. CONCLUSIONS: phiC31-mediated gene therapy resulted in an important improvement in IDUA-/- mice, including locomotion, although the obstacles that need to be overcome to enable long-term gene therapy for MPSI are also noted.

Mesenchymal stem cells do not prevent antibody responses against human α-L-iduronidase when used to treat mucopolysaccharidosis type I.

Mucopolysaccharidosis type I (MPSI) is an autosomal recessive disease that leads to systemic lysosomal storage, which is caused by the absence of α-L-iduronidase (IDUA). Enzyme replacement therapy is recognized as the best therapeutic option for MPSI; however, high titers of anti-IDUA antibody have frequently been observed. Due to the immunosuppressant properties of MSC, we hypothesized that MSC modified with the IDUA gene would be able to produce IDUA for a long period of time. Sleeping Beauty transposon vectors were used to modify MSC because these are basically less-immunogenic plasmids. For cell transplantation, 4×10(6) MSC-KO-IDUA cells (MSC from KO mice modified with IDUA) were injected into the peritoneum of KO-mice three times over intervals of more than one month. The total IDUA activities from MSC-KO-IDUA before cell transplantation were 9.6, 120 and 179 U for the first, second and third injections, respectively. Only after the second cell transplantation, more than one unit of IDUA activity was detected in the blood of 3 mice for 2 days. After the third cell transplantation, a high titer of anti-IDUA antibody was detected in all of the treated mice. Anti-IDUA antibody response was also detected in C57Bl/6 mice treated with MSC-WT-IDUA. The antibody titers were high and comparable to mice that were immunized by electroporation. MSC-transplanted mice had high levels of TNF-alpha and infiltrates in the renal glomeruli. The spreading of the transplanted MSC into the peritoneum of other organs was confirmed after injection of 111In-labeled MSC. In conclusion, the antibody response against IDUA could not be avoided by MSC. On the contrary, these cells worked as an adjuvant that favored IDUA immunization. Therefore, the humoral immunosuppressant property of MSC is questionable and indicates the danger of using MSC as a source for the production of exogenous proteins to treat monogenic diseases.