485. ZFN-Mediated Liver-Targeting Gene Therapy Corrects Systemic and Neurological Disease of Mucopolysaccharidosis Type I

Abstract

Mucopolysaccharidosis type I (MPS I) is characterized by progressive neurodegeneration, and premature death (<10 years). Caused by α-L-iduronidase (IDUA) deficiency and systemic accumulation of glycosaminoglycans (GAG), current therapies include stem cell transplant (with significant risk of morbidity and mortality), and enzyme replacement therapy (requiring costly and frequent long therapeutic infusion sessions). A promising alternative is genome editing by integration of a therapeutic hIDUA transgene using zinc finger nucleases (ZFN). We have previously demonstrated AAV-mediated in vivo targeting of the albumin locus as a “safe harbor” for coagulation factors, correcting clotting defects in hemophilic mice. Targeted insertion of a transgene into the genome offers multiple advantages. We can exploit the highly active albumin enhancer/promoter, such that stably modified hepatocytes demonstrate long-term transgene expression. Also, utilization of an endogenous promoter allows efficient packaging of the transgene donor into AAV. MPS I mice (n=8 per gender, 4-9 weeks old) were injected with a single dose of AAV2/8 encoding albumin-targeted ZFN and a donor encoding a partial hIDUA cDNA. MiSeq analysis showed that treated mice displayed significant levels of insertions/deletions (indels) (up to 56%) at the target locus, demonstrating efficient delivery and expression of the albumin ZFNs. Significant plasma IDUA activity was also observed in the ZFN+hIDUA donor treated mice, up to 10-fold of wild type levels, throughout the 120-day study. Urine GAG levels serving as a biomarker for hIDUA activity were reduced significantly (up to 90%). IDUA levels in these animals increased significantly in liver (up to 14 fold), heart, lung, muscle and spleen. Tissue GAG levels were significantly reduced in liver (by 91%), heart (85%), lung (86%), muscle (68%) and spleen (84%). Barnes maze tests at the end of the study showed that ZFN+hIDUA donor treated MPS I mice achieved significant neurological benefits compared with untreated MPS I mice.ZFN-mediated genome editing of hepatocytes in vivo thus resulted in high and stable levels of hIDUA expression in treated animals. This enzyme was secreted into plasma and then taken up by secondary tissues, leading to significant GAG reduction. The correction of the observable neurological impairment in MPS I mice suggests that a small but sufficient amount of hIDUA protein crossed the blood-brain barrier in these animals.The study provides a general “proof-of-concept” for treatment of lysosomal diseases.