α- l-Iduronidase transport in neurites

Abstract

Effective therapeutic strategies for mucopolysaccharidosis type I (MPSI) rely on mannose-6-phosphate receptor-mediated uptake of extracellular α- l-iduronidase (IDUA), the missing lysosomal enzyme in this disease, by deficient cells. Intravenously infused recombinant human IDUA does not reach the central nervous system, whereas neuropathology and neurological manifestations are prominent in Hurler syndrome, the most severe and most frequent form of MPSI. The creation of a single intracerebral source of IDUA by gene therapy was proved efficient to deliver enzyme throughout the brain of MPSI mice. IDUA spreading far beyond areas where the enzyme was synthesized suggested transport along neuronal processes. To examine the mechanisms of IDUA spreading in the brain, we constructed a chimeric protein in which GFP is fused at the C-terminus of IDUA. The fusion protein was expressed in rat primary neurons using lentivirus vectors. Fluorescent IDUA retained full catalytic activity including on natural substrates, interacted with mannose-6-phosphate receptors and was appropriately addressed to lysosomes. Fluorescent vesicles were broadly distributed over neuronal soma and processes. Time-lapse fluorescent video-microscopy showed that 54% of fluorescent vesicles exhibited either retrograde or anterograde displacements along neurites. Most moving organelles showed complex movements with frequent direction changes and arrests. Motility depended on microtubule integrity. Efficient axono-dendritic transport of IDUA provides a rationale for gene therapy based on the release of therapeutic enzyme at discrete locations within the central nervous system of patients with severe form of MPSI.