Waning efficacy in a long-term AAV-mediated gene therapy study in the murine model of Krabbe disease

Gregory J. Heller, Michael S. Marshall, Yazan Issa, Jeffrey N. Marshall, Duc Nguyen, Emily Rue, Koralege C. Pathmasiri, Miriam S. Domowicz, Richard B. van Breemen, Leon M. Tai, Stephanie M. Cologna, Stephen J. Crocker, Maria I. Givogri, Mark S. Sands, Ernesto R. Bongarzone

Research output: Contribution to journalArticlepeer-review

11 Scopus citations


Neonatal AAV9-gene therapy of the lysosomal enzyme galactosylceramidase (GALC) significantly ameliorates central and peripheral neuropathology, prolongs survival, and largely normalizes motor deficits in Twitcher mice. Despite these therapeutic milestones, new observations identified the presence of multiple small focal demyelinating areas in the brain after 6–8 months. These lesions are in stark contrast to the diffuse, global demyelination that affects the brain of naive Twitcher mice. Late-onset lesions exhibited lysosomal alterations with reduced expression of GALC and increased psychosine levels. Furthermore, we found that lesions were closely associated with the extravasation of plasma fibrinogen and activation of the fibrinogen-BMP-SMAD-GFAP gliotic response. Extravasation of fibrinogen correlated with tight junction disruptions of the vasculature within the lesioned areas. The lesions were surrounded by normal appearing white matter. Our study shows that the dysregulation of therapeutic GALC was likely driven by the exhaustion of therapeutic AAV episomal DNA within the lesions, paralleling the presence of proliferating oligodendrocyte progenitors and glia. We believe that this is the first demonstration of diminishing expression in vivo from an AAV gene therapy vector with detrimental effects in the brain of a lysosomal storage disease animal model. The development of this phenotype linking localized loss of GALC activity with relapsing neuropathology in the adult brain of neonatally AAV-gene therapy-treated Twitcher mice identifies and alerts to possible late-onset reductions of AAV efficacy, with implications to other genetic leukodystrophies.

Original languageEnglish
Pages (from-to)1883-1902
Number of pages20
JournalMolecular Therapy
Issue number5
StatePublished - May 5 2021


  • AAV9
  • demyelination
  • fibrinogen
  • gene therapy
  • globoid cell leukodystrophy
  • lysosomal dysfunction
  • microglia
  • multi-focal sclerosis
  • psychosine
  • tight junction


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