Genetically Corrected iPSC-Derived Neural Stem Cell Grafts Deliver Enzyme Replacement to Affect CNS Disease in Sanfilippo B Mice

Abstract

Sanfilippo syndrome type B (mucopolysaccharidosis type IIIB [MPS IIIB]) is a lysosomal storage disorder primarily affecting the brain that is caused by a deficiency in the enzyme α-N-acetylglucosaminidase (NAGLU), leading to intralysosomal accumulation of heparan sulfate. There are currently no treatments for this disorder. Here we report that, ex vivo, lentiviral correction of Naglu−/− neural stem cells derived from Naglu−/− mice (iNSCs) corrected their lysosomal pathology and allowed them to secrete a functional NAGLU enzyme that could be taken up by deficient cells. Following long-term transplantation of these corrected iNSCs into Naglu−/− mice, we detected NAGLU activity in the majority of engrafted animals. Successfully transplanted Naglu−/− mice showed a significant decrease in storage material, a reduction in astrocyte activation, and complete prevention of microglial activation within the area of engrafted cells and neighboring regions, with beneficial effects extending partway along the rostrocaudal axis of the brain. Our results demonstrate long-term engraftment of iNSCs in the brain that are capable of cross-correcting pathology in Naglu−/− mice. Our findings suggest that genetically engineered iNSCs could potentially be used to deliver enzymes and treat MPS IIIB.

Brain transplantation of genetically corrected Sanfilippo B Neural Stem Cells induces partial cross-correction of the disease

“…We found a significant increase of CD68 and GFAP positive signal in the forebrain of vehicleinjected Naglu −/− mice (Vehicle -/-), compared with control mice (Unaffected +/-) (Fig. 3A, B), data that is consistent with our previous findings 35 . We found a 72-fold increase of the CD68 staining in vehicle-injected mice compared with unaffected ones (p = <0.0001).…”

“…Previously, we reprogrammed Naglu -/mouse embryonic fibroblasts into iPSCs and corrected them ex vivo to overexpress the human NAGLU enzyme. We showed that corrected NSCs (N-iNSCs) improved the MPS-associated brain neuropathology in Naglu -/mice 35 . As described above, NAGLU-IGFII is under clinical investigation for ERT.…”

Enzyme Replacement Therapy for Mucopolysaccharidosis IIID using Recombinant Human α-N-Acetylglucosamine-6-Sulfatase in Neonatal Mice

“…Neonatal mice (both genders) were toe-clipped and genotyped at birth. Sterile-filtered (0.2 μm), purified rhGNS (5 μL; 13.4 or 40.2 units /μL), or vehicle (5 μL) was injected in the left lateral ventricle as previously described . The injection site was ∼2.5 mm between the bregma and eyes, 2 mm away laterally from the sagittal suture, and 2.5 mm in depth.…”

Recombinant NAGLU-IGF2 prevents physical and neurological disease and improves survival in Sanfilippo B syndrome

“…36 To study mannose 6-phosphorylation of NAGLU in the context of gene therapy, induced pluripotent stem cells derived from Sanfilippo B mouse embryonic fibroblasts were transduced with lentivirus bearing the human NAGLU transgene under a cytomegalovirus promoter. 48 The supernatant from these cells was added to Sanfilippo B patient fibroblasts in the presence or absence of mannose 6-phosphate. The study demonstrated uptake of NAGLU from the supernatant of these lentiviral-modified cells into Sanfilippo B fibroblasts that was nearly completely abolished with the addition of mannose 6-phosphate to the media, suggesting that NAGLU produced by gene therapy may not suffer the same post-translational processing issues as recombinant NAGLU enzyme produced by cells in vitro, as was used in this study.…”