Systemic Delivery of Tyrosine-Mutant AAV Vectors Results in Robust Transduction of Neurons in Adult Mice

Abstract

Recombinant adeno-associated virus (AAV) vectors are powerful tools for both basic neuroscience experiments and clinical gene therapies for neurological diseases. Intravascularly administered self-complementary AAV9 vectors can cross the blood-brain barrier. However, AAV9 vectors are of limited usefulness because they mainly transduce astrocytes in adult animal brains and have restrictions on foreign DNA package sizes. In this study, we show that intracardiac injections of tyrosine-mutant pseudotype AAV9/3 vectors resulted in extensive and widespread transgene expression in the brains and spinal cords of adult mice. Furthermore, the usage of neuron-specific promoters achieved selective transduction of neurons. These results suggest that tyrosine-mutant AAV9/3 vectors may be effective vehicles for delivery of therapeutic genes, including miRNAs, into the brain and for treating diseases that affect broad areas of the central nervous system.

Benefits of Neuronal Preferential Systemic Gene Therapy for Neurotransmitter Deficiency

“…11 We analyzed the expression of AADC in the brain using immunohistochemistry (IHC). Under low magnification, AADC-positive cells were observed throughout the brain of AAV9-AADC-treated mice …”

“…11 We also examined AADC expression in mice after gene therapy. Scattered AADC-positive cells were observed in the liver, adrenal medulla, and superior mesenteric ganglion in both AAV9-AADC-and AAVN-AADC-treated mice ( Figure 8).…”

“…In this A c c e p t e d m a n u s c r i p t study, we extended the treatment model of AADC deficiency to systemic treatment in young mice by employing vectors containing either a ubiquitous promoter (i.e., AAV9-AADC) or a promoter for preferential neuronal expression (i.e., yfAAV9/3-Syn-I-hAADC, AAVN-AADC). 11 …”

“…The AAVN vector consisted of a tyrosine-mutant AAV9/3-mAADC vector driven by the synapsin I (Syn-I) promoter for preferential neuronal expression (yfAAV9/3-Syn-I-mAADC; AAVN-AADC). This vector consisted of virus posttranscriptional regulatory element, as previously reported 11. The expression cassette was cloned into a plasmid with AAV serotype 3 inverted terminal repeats and then packaged into an AAV type 9 capsid with an amino acid substitution from tyrosine to phenylalanine at position 446 of the AAV9 capsid protein.…”

Systemic Vascular Transduction by Capsid Mutant Adeno-Associated Virus After Intravenous Injection

“…1A). Specifically, site-directed tyrosine-to-phenylalanine (Y-F) and threonine-to-valine (T-V) substitutions have been shown to effectively prevent phosphorylation and subsequent ubiquitin-mediated proteolysis of the capsid, resulting in substantially improved transduction efficiency and altered tropism in several tissues, including the eye, [24][25][26] brain, 27 and muscle. 28 Building on these studies, herein we evaluate a number of AAV serotype 2 (AAV2)-based vectors containing combinations of single-amino acid substitutions at specific sites throughout the shared C-terminal domain (Y272F, Y444F, T491V, Y500F, Y730F; see Fig.…”