Dataset Information

AAV9 transduction mediated by systemic delivery of vector via retro-orbital injection in newborn, neonatal and juvenile mice.

ABSTRACT: Adeno-associated virus (AAV)-based gene therapy is gaining popularity owing to its excellent safety profile and effective therapeutic outcomes in a number of diseases. Intravenous (IV) injection of AAV into the tail vein, facial vein and retro-orbital (RO) venous sinus have all been useful strategies to infuse the viral vector systemically. However, tail vein injection is technically challenging in juvenile mice, and injection at young ages (≤ postnatal day-(P)21) is essentially impossible. The temporal or facial vein is localized anterior to the ear bud and is markedly visible in the first couple of days postnatally. However, this method is age-dependent and requires a dissecting microscope. Retro-orbital injection (ROI), on the other hand, is suitable for all murine ages, including newborn and older mice, and is relatively less stressful to animals compared to tail vein injection. Although many reports have shown ROI as an effective route of AAV delivery, herein we aim to highlight and summarize the methods and benefits of ROI. To capture the full spectrum of transduction efficiency mediated by ROI, we transduced the editing-dependent reporter mice (Ai9 Cre reporter mice) with the AAV9 vector, which targets a wide range of peripheral tissues with exceptional brain tropism. We also provide a comprehensive description of the ROI technique to facilitate viral vector administration without complications.

SUBMITTER: Prabhakar S

PROVIDER: S-EPMC8614020 | biostudies-literature |

REPOSITORIES: biostudies-literature

ACCESS DATA

Similar Datasets

Project description:Several neurodevelopmental and neurodegenerative disorders affecting the central nervous system are potentially treatable via viral vector-mediated gene transfer. Adeno-associated viral (AAV) vectors have been used in clinical trials because of their desirable properties including a high degree of safety, efficacy, and stability. Major factors affecting tropism, expression level, and cell type specificity of AAV-mediated transgenes include encapsidation of different AAV serotypes, promoter selection, and the timing of vector administration. In this study, we evaluated the ability of single-stranded AAV2 vectors pseudotyped with viral capsids from serotype 9 (AAV2/9) to transduce the brain and target gene expression to specific cell types after intracerebroventricular injection into mice. Titer-matched AAV2/9 vectors encoding the enhanced green fluorescent protein (eGFP) reporter, driven by the cytomegalovirus (CMV) promoter, or the neuron-specific synapsin-1 promoter, were injected bilaterally into the lateral ventricles of C57/BL6 mice on postnatal day 5 (neonatal) or 21 (juvenile). Brain sections were analyzed 25 days after injection, using immunocytochemistry and confocal microscopy. eGFP immunohistochemistry after neonatal and juvenile administration of viral vectors revealed transduction throughout the brain including the striatum, hippocampus, cerebral cortex, and cerebellum, but with different patterns of cell-specific gene expression. eGFP expression was seen in astrocytes after treatment on postnatal day 5 with vectors carrying the CMV promoter, expanding the usefulness of AAVs for modeling and treating diseases involving glial cell pathology. In contrast, injection of AAV2/9-CMV-eGFP on postnatal day 21 resulted in preferential transduction of neurons. Administration of AAV2/9-eGFP with the synapsin-1 promoter on either postnatal day 5 or 21 resulted in widespread neuronal transduction. These results outline efficient methods and tools for gene delivery to the nervous system by direct, early postnatal administration of AAV vectors. Our findings highlight the importance of promoter selection and age of administration on the intensity, distribution, and cell type specificity of AAV transduction in the brain.

Project description:Recombinant adeno-associated virus (rAAV) vectors have been widely used as favored delivery vehicles for the treatment of inherited diseases in clinical trials, including neurological diseases. However, the noninvasive systemic delivery of rAAV to the central nervous system is severely hampered by the blood-brain barrier (BBB). Several approaches have been exploited to enhance AAV vector brain transduction after systemic administration, including genetic modification of AAV capsids and physical methods. However, these approaches are not always predictive of desirable outcomes in humans and induce complications. It is imperative to explore novel strategies to increase the ability of AAV9 to cross the BBB for enhanced brain transduction. Herein, we have conducted a combinatorial in vivo/in vitro phage display library screening in mouse brains and purified AAV9 virions to identify a customized BBB shuttle peptide, designated as PB5-3. The PB5-3 peptide specifically bound to AAV9 virions and enhanced widespread transduction of AAV9 in mouse brains, especially in neuronal cells, after systemic administration. Further study demonstrated that systemic administration of AAV9 vectors encoding IDUA complexed with PB5-3 increased the phenotypic correction in the brains of MPS I mice. Mechanistic studies revealed that the PB5-3 peptide effectively increased AAV9 trafficking and transcytosis efficiency in the human BBB model hCMEC/D3 cell line but did not interfere with AAV9 binding to the receptor terminal N-linked galactosylated glycans. Additionally, the PB5-3 peptide slowed the clearance of AAV9 from blood without hepatic toxicity. This study highlights, for the first time, the potential of this combinatorial approach for the isolation of peptides that interact with specific AAV vectors for enhanced and targeted AAV transduction. This promising approach will open new combined therapeutic avenues and shed light on the potential applications of peptides for the treatment of human diseases in future clinical trials with AAV vector-mediated gene delivery.

Dataset Information

AAV9 transduction mediated by systemic delivery of vector via retro-orbital injection in newborn, neonatal and juvenile mice.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets