Recombinant Adeno Associated Viral (AAV) vector type 9 delivery of Ex1-Q138-mutant huntingtin in the rat striatum as a short-time model for in vivo studies in drug discovery

Abstract

Huntington's disease (HD) is an inherited neurodegenerative disorder characterized by dyskinesia, cognitive impairment and emotional disturbances, presenting progressive neurodegeneration in the striatum and intracellular mutant Huntingtin (mHTT) aggregates in various areas of the brain. Recombinant Adeno Associated Viral (rAAV) vectors have been successfully used to transfer foreign genes to the brain of adult animals. In the present study we report a novel in vivo rat HD model obtained by stereotaxic injection of rAAV serotype2/9 containing Exon1-Q138 mHTT (Q138) and Exon1-Q17 wild type HTT (Q17; control), respectively in the right and in the left striatum, and expressed as C-terminal GFP fusions to facilitate detection of infected cells and aggregate production. Immunohistochemical analysis of brain slices from animals sacrificed twenty-one days after viral infection showed that Q138 injection resulted in robust formation of GFP-positive aggregates in the striatum, increased GFAP and microglial activation and neurodegeneration, with little evidence of any of these events in contralateral tissue infected with wild type (Q17) expressing construct. Differences in the relative metabolite concentrations (N-Acetyl Aspartate/Creatine and Myo-Inositol/Creatine) were observed by H1 MR Spectroscopy. By quantitative RT-PCR we also demonstrated that mHTT induced changes in the expression of genes previously shown to be altered in other rodent HD models. Importantly, administration of reference compounds previously shown to ameliorate the aggregation and neurodegeneration phenotypes in preclinical HD models was demonstrated to revert the mutant HTT-dependent effects in our model. In conclusion, the AAV2/9-Q138/Q17 exon 1 HTT stereotaxic injection represents a useful first-line in vivo preclinical model for studying the biology of mutant HTT exon 1 in the striatum and to provide early evidence of efficacy of therapeutic approaches.

Adeno-Associated Viral Vector Serotype DJ-Mediated Overexpression of N171-82Q-Mutant Huntingtin in the Striatum of Juvenile Mice Is a New Model for Huntington’s Disease

“…However, features of movement disorder were different to that of N171-82Q Tg mice (expressing a mutant N-terminal fragment of HTT ), which displayed loss of coordination, hunched posture, tremors, abnormal gait, and clasping of hindlimbs (Schilling et al, 1999 ). In the present study, HTT aggregates were observed only in the striatum, similar to the in vivo rat HD model obtained by stereotaxic injection of AAV serotype nine containing Exon1-Q138 mutant HTT (AAV-9-Q138) (Ceccarelli et al, 2016 ). However, the expression pattern was different to that of N171-82Q Tg mice (Schilling et al, 1999 ).…”

“…Presently, recombinant AAVs are one of the preferred vectors, because their stable transduction of dividing and non-dividing cells, strong neural tropism, low risk of insertional mutagenesis, and reduced immune responses (Blessing and Déglon, 2016 ; Grieger et al, 2016 ; Saraiva et al, 2016 ). According to many studies that measured the ability of AAV serotypes to target the CNS, when administrated into the brain parenchyma of rodents, most of the serotypes (1, 2, 4, 5, 8 and 9) transduced neurons and glia in the CNS areas including striatum, hippocampus and neocortex (Ruiz and Déglon, 2012 ; Watakabe et al, 2015 ; Ceccarelli et al, 2016 ; Saraiva et al, 2016 ). Other serotypes, such as AAV-DJ, are potentially useful but have been less well examined (Cearley and Wolfe, 2006 ; Klein et al, 2008 ; Aschauer et al, 2013 ; Holehonnur et al, 2014 ).…”

“…Huntington’s disease (HD) is an inherited progressive neurodegenerative disorder characterized by involuntary abnormal movements (chorea), cognitive decline and emotional as well as psychiatric disturbances (Damiano et al, 2010 ; Jacobsen et al, 2011 ; Gil-Mohapel et al, 2014 ). HD typically becomes apparent at 35–45 years of age and progresses toward death within 15–20 years after the appearance of the first clinical symptoms (Ceccarelli et al, 2016 ). HD is caused by an abnormal expansion of a CAG codon ≥36 repeats located in exon one of the huntingtin gene ( HTT ) on chromosome 4 that confers a toxic function to the protein (Damiano et al, 2010 ; Jacobsen et al, 2011 ; Gil-Mohapel et al, 2014 ).…”

“…The model revealed the rapid formation of fibrillar, cytoplasmic and ubiquitinated nuclear polyglutamine aggregates in neurons (Senut et al, 2000 ). Recently, a rat HD model obtained by AAV serotype 2/9 containing Exon 1-Q138 mutant HTT (Q138) was reported as a short-term model for in vivo studies in drug discovery (Ceccarelli et al, 2016 ). Although these models are useful in HD studies, more models would provide more options to researchers.…”

Emerging Diagnostic and Therapeutic Strategies for Tauopathies

“…Viral vectors may be used as well for siRNA delivery, which have the advantage of being able to directly target the nucleus, and such an approach has been used in animal models of Huntington’s disease and amyotrophic lateral sclerosis [173–175]. Intraparenchymal injections have been utilized in rat and non-human primate models of Huntington’s disease to delivery these viral vectors [176, 177]. Other strategies to transiently increase BBB permeability have been investigated as well including a variety of different compounds and most recently focused ultrasound [178–182].…”