Intravenous Delivery of a Gene Therapy Vector that Expresses an Antitransferrin Receptor 1 Nanobody-I2S Fusion Protein Rescued Central Nervous System Lysosomal Burden in Mucopolysaccharidosis II Mice

Hunter syndrome, also known as mucopolysaccharidosis type II, is an X-linked lysosomal storage disease caused by the deficiency of functional iduronate-2-sulfatase (I2S) enzyme, leading to the accumulation of lysosomal glycosaminoglycans (GAGs) affecting multiple organs. Two-thirds of patients have central nervous system (CNS) manifestations. The current standard of care, enzyme replacement therapy (ERT) via weekly intravenous delivery of recombinant human I2S (rhI2S), does not address the neuropathy in the CNS due to its inability to cross the blood–brain barrier (BBB). Next-generation ERTs consisting of systemically administered rhI2S linked to antibodies that target the transferrin receptor (TfR) have shown clinical efficacy in addressing CNS and peripheral manifestations of disease. We demonstrate here that systemic administration of recombinant AAV9 gene therapy vectors encoding human I2S fusion protein with a TfR1-targeted Variable Heavy chain domain of Heavy chain (VHH) nanobody at the N-terminus normalized brain and cerebrospinal fluid GAGs in symptomatic Ids knockout ( Ids KO) mice. This ability to correct toxic substrate accumulation in the CNS was superior to gene therapy vectors expressing I2S with a C-terminal VHH tag or untagged I2S control. The VHH-I2S transgene product demonstrated a broader distribution in the brain parenchyma, coincident with a significant reduction of lysosomal-associated membrane protein 1 immunoreactivity, unlike untagged I2S and I2S-VHH transgene products. These data illuminate strategies to enhance AAV gene therapy vector design and leverage receptor-mediated transcytosis to strategize BBB-penetrating gene therapy for addressing the unmet medical needs of neuronopathic Hunter syndrome.