Mucopolysaccharidosis (MPS) IIIB is a neuropathic lysosomal storage disease characterized by the deficient activity of a lysosomal enzyme obligate for the degradation of the glycosaminoglycan (GAG) heparan sulfate (HS). The pathogenesis of neurodegeneration in MPS IIIB is incompletely understood. Large animal models are attractive for pathogenesis and therapeutic studies due to their larger size, outbred genetics, longer lifespan, and naturally occurring MPS IIIB disease. However, the temporospatial development of neuropathologic changes has not been reported for canine MPS IIIB. Here we describe lesions in 8 brain regions, cervical spinal cord, and dorsal root ganglion (DRG) in a canine model of MPS IIIB that includes dogs aged from 2 to 26 months of age. Pathological changes in the brain included early microscopic vacuolation of glial cells initially observed at 2 months, and vacuolation of neurons initially observed at 10 months. Inclusions within affected cells variably stained positively with PAS and LFB stains. Quantitative immunohistochemistry demonstrated increased glial expression of GFAP and Iba1 in dogs with MPS IIIB compared to age-matched controls at all time points, suggesting neuroinflammation occurs early in disease. Loss of Purkinje cells was initially observed at 10 months and was pronounced in 18- and 26-month-old dogs with MPS IIIB. Our results support the dog as a replicative model of MPS IIIB neurologic lesions and detail the pathologic and neuroinflammatory changes in the spinal cord and DRG of MPS IIIB-affected dogs.

Mucopolysaccharidosis type IIIB (MPS IIIB; Sanfilippo syndrome B; OMIM #252920) is a lethal, pediatric, neuropathic, autosomal recessive, and lysosomal storage disease with no approved therapy. Patients are deficient in the activity of N-acetyl-alpha-glucosaminidase (NAGLU; EC 3.2.150), necessary for normal lysosomal degradation of the glycosaminoglycan heparan sulfate (HS). Tralesinidase alfa (TA), a fusion protein comprised of recombinant human NAGLU and a modified human insulin-like growth factor 2, is in development as an enzyme replacement therapy that is administered via intracerebroventricular (ICV) infusion, thus circumventing the blood brain barrier. Previous studies have confirmed ICV infusion results in widespread distribution of TA throughout the brains of mice and non-human primates. We assessed the long-term tolerability, pharmacology, and clinical efficacy of TA in a canine model of MPS IIIB over a 20-month study. Long-term administration of TA was well-tolerated as compared to administration of vehicle. TA was widely distributed across brain regions, which was confirmed in a follow-up 8-week pharmacokinetic/pharmacodynamic study. MPS IIIB dogs treated for up to 20 months had near-normal levels of HS and HS-NRE in CSF and central nervous system (CNS) tissues. TA-treated MPS IIIB dogs performed better on cognitive tests, had improved CNS pathology and decreased cerebellar volume loss relative to vehicle-treated MPS IIIB dogs. These findings demonstrate the ability of TA to prevent or limit the biochemical, pathological, and cognitive manifestations of canine MPS IIIB disease, thus providing support of its potential long-term tolerability and efficacy in MPS IIIB subjects.

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Recombinant human alpha-N-acetylglucosaminidase-insulin-like growth factor-2 (rhNAGLU-IGF2) is an investigational enzyme replacement therapy for Sanfilippo B, a lysosomal storage disease. The fusion protein with IGF2 permits binding to the cation-independent mannose 6-phosphate receptor, because recombinant human NAGLU (rhNAGLU) is poorly mannose 6-phosphorylated. We previously administered rhNAGLU-IGF2 intracerebroventricularly to Sanfilippo B mice. We demonstrated therapeutic restoration of NAGLU, normalization of lysosomal storage, and improvement in markers of neurodegeneration and inflammation. Here, we studied intracerebroventricular rhNAGLU-IGF2 in murine and canine Sanfilippo B to determine potential effects on the behavioral phenotype and survival. Heparan sulfate (HS) levels in brain and heart were reduced following treatment with rhNAGLU-IGF2 or rhNAGLU. Treated mice showed improvement in disease markers such as beta-hexosaminidase, CD68, and LAMP1. We found a more normal number of stretch attend postures, a fear pose, in mice treated with either rhNAGLU or rhNAGLU-IGF2 vs vehicle-treated Sanfilippo B mice on elevated plus maze testing (p<0.001). We found a more normal dark/light activity pattern in Sanfilippo B mice treated with rhNAGLU-IGF2 compared to vehicle-treated Sanfilippo B mice (p=0.025). We found a 61% increase in survival in Sanfilippo B mice treated with rhNAGLU-IGF2 (mean 53d, median 48d) compared to vehicle-treated Sanfilippo B mice (mean 33d, median 37d; p<0.001). In canine Sanfilippo B, we found that rhNAGLU-IGF2 administered into cerebrospinal fluid normalized heparan sulfate and beta-hexosaminidase activity in gray and white matter brain regions. Proteomics analysis of cerebral cortex showed restoration of protein concentrations in pathways relevant to cognitive function, synapse, and the lysosome. Treatment with rhNAGLU-IGF2 may improve the phenotype of Sanfilippo B disease.

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