Efficient and stable transduction of dopaminergic neurons in rat substantia nigra by rAAV 2/1, 2/2, 2/5, 2/6.2, 2/7, 2/8 and 2/9

Abstract

Dysfunction of the nigrostriatal system is the major cause of Parkinson's disease (PD). This brain region is therefore an important target for gene delivery aiming at disease modeling and gene therapy. Recombinant adeno-associated viral (rAAV) vectors have been developed as efficient vehicles for gene transfer into the central nervous system. Recently, several serotypes have been described, with varying tropism for brain transduction. In light of the further development of a viral vector-mediated rat model for PD, we performed a comprehensive comparison of the transduction and tropism for dopaminergic neurons (DNs) in the adult Wistar rat substantia nigra (SN) of seven rAAV vector serotypes (rAAV 2/1, 2/2, 2/5, 2/6.2, 2/7, 2/8 and 2/9). All vectors were normalized by titer and volume, and stereotactically injected into the SN. Gene expression was assessed non-invasively and quantitatively in vivo by bioluminescence imaging at 2 and 5 weeks after injection, and was found to be stable over time. Immunohistochemistry at 6 weeks following injection revealed the most widespread enhanced green fluorescence protein expression and the highest number of positive nigral cells using rAAV 2/7, 2/9 and 2/1. The area transduced by rAAV 2/8 was smaller, but nevertheless almost equal numbers of nigral cells were targeted. Detailed confocal analysis revealed that serotype 2/7, 2/9, 2/1 and 2/8 transduced at least 70% of the DNs. In conclusion, these results show that various rAAV serotypes efficiently transduce nigral DNs, but significant differences in transgene expression pattern and level were observed.

Serotype-dependent transduction efficiencies of recombinant adeno-associated viral vectors in monkey neocortex

“…Other studies with injections in the SN of rats reported higher transduced brain volumes using rAAV2/7, rAAV2/9, and rAAV2/1 than with rAAV2/8 and rAAV2/5, indicating species-dependent transduction efficiencies of the viral vectors. 30,33,62 In our study, the rAAV2/1-CaMKIIα0. 4 showed efficient transduction at the injection site but the transgene expression pattern was limited to about 1.2 × 1.0 mm 2 .…”

“…About 100 ng of genomic DNA was used for the determination of the rAAV genomic copies using a real-time PCR, as described earlier. 33 Each dilution of the viral vectors was used three times and this experiment was done in two different assays.…”

“…[30][31][32] Unfortunately, the latter properties are species specific and even brain region specific. 30,33 Moreover, information about transduction and expression properties obtained in rodents does not always translate to primates. [34][35][36][37][38] In macaque monkeys, a comparison between the transduction properties of different rAAVs in subcortical structures, including the substantia nigra, striatum, 39,40 and basal ganglia, 41 revealed that rAAV2/1 and rAAV2/5 were most efficient when under the control of a synthetic chicken ß-actin promoter.…”

Preclinical Evaluation of a P2X7 Receptor–Selective Radiotracer: PET Studies in a Rat Model with Local Overexpression of the Human P2X7 Receptor and in Nonhuman Primates

“…To obtain efficient gene transfer, we applied viral vector technology, engineering a hP2X7R reporter system using HIV-based lentiviral vectors (LVs) for validation in cell culture settings, and recombinant adeno-associated viral (rAAV) vectors in the brain in vivo (19). JNJ-54173717 (20,21), which has nanomolar affinity for rat P2X7R and hP2X7R, was radiolabeled with 11 C and evaluated in this rat model using small-animal PET.…”

Viral Vectors in Neurobiology: Therapeutic and Research Applications

“…This is the case for gene therapy approaches in epilepsy, multiple sclerosis, Alzheimer disease, diabetes, Parkinson's disease, chronic pain, lysosomal storage disorders, amyotrophic lateral sclerosis, brain ischemia, seizure, Huntington disease, and others. HSV vectors have also been used for some of these indications, but currently, they have been frequently applied for gene therapy approaches in brain tumors, as is the case for adenovirus vectors as well (Maingay et al, 2005;Wong et al, 2006;Robinson et al, 2007;Lowery et al, 2009;Shih et al, 2009;Björklund et al, 2010;Manfredsson & Mandel, 2010;Eskenazi & Neumaier, 2011;Jacobs & Wang, 2011;Van der Perren et al, 2011;Vande Velde et al, 2011;Thaci et al, 2011).…”