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

Abstract

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.

Chronic lithium administration in a mouse model for Krabbe disease

“… 36 Unfortunately, however, in all GT studies KD mice at a certain point died with uncertain complications. 36 , 68 , 69 , 70 Two very recently published papers suggest that GT could not be able to efficiently act on a long time due to the exhaustion of therapeutic AAV episomal DNA in specific regions, 23 or to the onset of additional pathological events unrelated to KD, as hepatocellular carcinoma (HCC). 22 Additionally, a very intriguing possibility is that there could be also additionally pathogenic mechanisms, not directly related to GALC deficiency, which could significantly contribute to the pathogenesis and the bad progression of KD.…”

“…In all studies, however, this approach did not allow completely curing the disease. Although GT was successful in increasing the GALC activity and reducing PSY accumulation, the lifespan of treated KD animals 22,23 could not reach the one of the non‐affected controls. This might be attributed to the fact that these therapies need time to engraft and create therapeutic effects, not allowing the effective prevention of the very early nervous system damage.…”

Non-Viral Delivery of RNA Gene Therapy to the Central Nervous System

“…The mice demonstrated decreased central and peripheral neuropathology and prolonged survival, but at ages of 6-8 months began to show small focal demyelinating regions in the brain. This suggested dysregulation of therapeutic GALC, likely due to exhaustion of the tranduced episomal DNA [20]. Work will continue on systems using viral vectors to improve their safety in light of the potential benefit of a single curative treatment.…”

Krabbe Disease: Prospects of Finding a Cure Using AAV Gene Therapy

“…These results were reflected in the low levels of psychosine in the brain, spinal cord, and sciatic nerve of treated mice at P40. However, at the terminal timepoint, levels of psychosine in each of these regions was significantly high ( 34 ), likely indicative of decreasing GALC levels due to episomal dilution in proliferating oligodendrocyte progenitor cells and glia over time ( 125 ).…”