208. Sleeping Beauty Transposon-Mediated Long-Term α-L-iduronidase Expression and Correction of Lysosomal Pathology in the Murine Model of Mucopolysaccharidosis (MPS) Type I

Abstract

Top of pageAbstract The purpose of this project was to evaluate the Sleeping Beauty (SB) transposon for delivery of the human |[alpha]|-L-iduronidase ( IDUA) gene to chromosomes in the livers of IDUA-deficient mice for long- term expression and correction of MPS type I. We chose the hydrodynamics-based DNA injection, which targets mainly the liver. We constructed SB transposon, plasmid-based vectors and injected them into idua-/-mice. Because our goal was in part to determine the efficacy of transposition as a way of providing long-term expression of IDUA protein, control groups of MPS I mice did not get the transposase. Blood samples for plasma isolation were collected 1 day after treatment and once every 2 weeks thereafter. Plasma IDUA activities reached >100-fold of wild type levels on day 1 following treatment, but were essentially gone in all mice 4 weeks following gene delivery. IDUA activity was not detected in the livers of these mice 3 months after plasmid administration. We examined the duration of the transposon-delivered human IDUA DNA maintenance and enzyme activity over 6 months in the liver of wild type (WT) C57Bl/6 mice that express IDUA. As in the MPS I mice, plasma and liver IDUA activity reached supra-normal levels on day 1 and remained at this level for the first week, but still dropped dramatically within 2 weeks and by 4 weeks were indistinguishable from background. Using DNA PCR, we found that the maintenance of the IDUA transgene mirrored the IDUA activity time-line. Even though transposition was confirmed by an excision assay, the PCR product was detectable only for the first two weeks, but not thereafter. It appears that cells that express the IDUA gene are cleared from the liver of treated mice by 4 weeks following injection, which suggests induction of an immune response either to the therapeutic protein or/and the cells that express the therapeutic gene. This hypothesis is supported by our observations that in all cyclophosphamide immune-suppressed MPS I mice, the initial 1-day plasma activity levels dropped more than 100-fold by 2 weeks, but then persisted at 20-500% WT activity in some mice. IDUA levels were stable (up to 5-fold higher than in the WT mice) in SB-transposase-treated MPS I mice whereas without SB, IDUA levels declined to near-background over 3 months. These persistent IDUA activity levels were sufficient to reduce the number and size of pathologic inclusions in the liver seen by toluidine blue staining. Secondary elevation of murine |[beta]|-glucuronidase, another lysosomal enzyme, was reduced in the liver, spleen and kidney and correlated with degree of reduction of lysosomal pathology in these organs. Thus, with immune suppression, a single dose of the SB transposon system can result in partial to complete biochemical correction of IDUA activity in the livers of treated adult MPS I mice.