Conditions for combining gene therapy with bone marrow transplantation in murine Krabbe disease

Introduction: Krabbe disease (KD) is an autosomal recessive lysosomal disorder caused by mutations in the galactocerebrosidase (GALC) gene. This results in defective myelination in the peripheral and central nervous systems due to low GALC activity. Treatment at this time is limited to hematopoietic stem cell transplantation (HSCT) in pre-symptomatic individuals. While this treatment extends the lives of treated individuals, most have difficulty walking by the end of the first decade due to peripheral neuropathy. Studies in the murine model of KD, twitcher (twi) combining bone marrow transplantation (BMT) with AAVrh10-mGALC showed a great extension of life from 40 days to about 400 days, with some living a full life time. Methods: In order to find the optimum conditions for dosing and timing of this combined treatment, twi mice were injected with five doses of AAVrh10-mGALC at different times after BMT. Survival, as well as GALC expression were monitored along with studies of sciatic nerve myelination and possible liver pathology. Results: Dosing had a pronounced effect on survival and measured GALC activity. There was window of time after BMT to inject the viral vector and see similar results, however delaying both the BMT and the viral injection shortened the lifespans of the treated mice. Lowering the viral dose too much decreased the correction of the sciatic nerve myelination. There was no evidence for hepatic neoplasia. Conclusion: These studies provide the conditions optimum for successfully treating the murine model of KD. There is some flexibility in dosing and timing to obtain a satisfactory outcome. These studies are critical to the planning of a human trial combining the standard of care, HSCT, with a single iv injection of AAVrh10-GALC.