Early or Late-Stage Anti-N-Terminal Huntingtin Intrabody Gene Therapy Reduces Pathological Features in B6.HDR6/1 Mice

Abstract

Huntington disease (HD) is a progressive neurodegenerative disease caused by an expansion of a polyglutamine sequence in mutant huntingtin (mhtt) that produces abnormal folding and aggregation that results in the formation of nuclear and cytoplasmic neuronal inclusion bodies. Although the precise role of mhtt aggregates in the pathogenesis is unclear, attempts to reduce accumulated mhtt protein have ameliorated the phenotype in multiple cellular and in vivo HD models. Here, we provide critical results on intracranial delivery of a single-chain Fv intrabody, C4, which targets the first 17 amino acids of the htt protein, a region of httExon1 that is increasingly being recognized as pivotal. To assess long-term efficacy and safety issues, we used adeno-associated viral vectors (AAV2/1) to deliver intrabody genes to striatum of inbred B6.HDR6/1 mice. Treatment initiation at various stages of the disease showed that early treatment preserved the largest number of cells without nuclear aggregates and that the accumulation of aggregated material could be delayed by several months. Even when intrabody treatment was not initiated until the clinical disease stage, significant, albeit smaller, effects were seen. These data indicate that neuronal intrabodies against critical N-terminal epitopes can be safely and effectively delivered using AAV2/1 to delay the aggregation phenotype over a sustained period of time in this HD model, even when delivery is initiated after disease onset.

Engineering humoral immunity as prophylaxis or therapy

“…Of the iAbs tested, Happ1 ameliorated neuropathology in cell lines and conferred significant beneficial effects in a variety of motor and cognitive assays, significantly prolonging life span by 10 weeks in a mouse model of disease [45]. Another group delivering iAb C4 via intrastriatal AAV1 delivery demonstrated that early treatment of an HD mouse model, prevented cells from exhibiting nuclear aggregates and delayed aggregate accumulation [46]. …”

Intrabodies as Neuroprotective Therapeutics

“…Quantitatively, both the size and number of HTT aggregates were significantly reduced at early-tomiddle stages of disease [48]. Confocal imaging with an anti-HA antibody, used to identify the presence of intrabody, confirmed that most transduced cells lack HTT aggregates initially.…”

“…For continuous production of intrabodies, viral delivery of genes encoding intrabodies has been shown to be effective in a fraction of cells, with persistence of signal over weeks-to-months [48,51]. Newer vectors may offer better spread and more efficient transduction, although localized and systemic immunogenicity remain concerns.…”

Bifunctional Anti-Huntingtin Proteasome-Directed Intrabodies Mediate Efficient Degradation of Mutant Huntingtin Exon 1 Protein Fragments

“…Intrastriatal delivery of scFv-C4, using the adeno-associated virus vector (AAV2/1), resulted in a significant reduction in the size and number of mhtt aggregates in B6.Cg-HDR6/1 transgenic mice. However, the neuroprotective effect weakened both with severity of disease at time of injection, and with age beyond 6 months, although it does not disappear entirely [23]. Additional optimization of scFv-C4 is required for this intrabody to be of future use in clinical applications.…”