Abstract
Adeno-associated virus (AAV) vectors have been successfully applied in clinical trials for hemophilic patients. Although promising, the clinical results suggest that the capsid-specific CD8+T cell response has a negative effect on therapeutic success. In an in vitro analysis using an engineered AAV virus carrying immune-dominant SIINFEKL peptide in the capsid backbone, we have previously demonstrated that capsid antigen presentation from full (genome containing) AAV capsids requires endosome escape and is proteasome dependent and that no capsid antigen presentation is induced from empty virions. In the present study, we examined capsid antigen presentation from administration of empty virions in animal models. In wild-type mice, similar to AAV full particles, capsid antigen presentation from AAV empty virion infection was dose dependent, and the kinetics studies showed that antigen presentation was detected from 2 to 40 days after AAV empty virion administration. In the transporter associated with antigen processing 1 deficient (TAP−/−) mice, capsid antigen presentation was inhibited from both AAV full and empty virions, but higher inhibition was achieved from AAV full particle administration than that from empty virions. This indicates that the pathway of capsid antigen presentation from AAV transduction is dependent on proteasome-mediated degradation of AAV capsids (mainly for full particles) and that the endosomal pathway may also play a role in antigen presentation from empty particles but not full virions. The capsid antigen presentation efficiency from AAV preparations was positively correlated with the amount of empty virions contaminated with full particles. Collectively, the results indicate that contamination of AAV empty virions induces efficient antigen presentation in vivo and the mechanism of capsid antigen presentation from empty virions involves both endosomal and proteasomal pathways. The elucidation of capsid antigen presentation from AAV empty virions may allow us to rationally design effective strategies to prevent elimination of AAV transduced target cells by capsid specific CD8+ T cells.
Highlights
Adeno-associated virus (AAV) vectors have been successfully used to transduce hepatocytes in Phase I clinical trials in patients with hemophilia B [1,2,3,4]
We used an engineered AAV2 capsid by swapping the immune-domain epitope, SIINFEKL, from ovalbumin protein into the AAV virion HI loop and demonstrated that capsid antigen presentation is elicited from AAV2-transduced liver cells in mice [17]
While the TAP pathway plays a major role in capsid antigen presentation from AAV full particles, both TAP and endosomal pathways may be involved in the capsid antigen presentation from empty virions in vivo
Summary
Adeno-associated virus (AAV) vectors have been successfully used to transduce hepatocytes in Phase I clinical trials in patients with hemophilia B [1,2,3,4]. Clinical results have suggested that capsid specific cytotoxic T lymphocytes (CTLs) eliminated AAV-transduced hepatocytes, which resulted in therapeutic failure [1,2,3,4]. These observations pose an outstanding concern regarding. Two main intracellular pathways for cross-presentation have been described: endocytic (TAP and proteasome-independent) and cytosolic (TAP and proteasome-dependent) MHC class I peptide loading [5]. Using pharmacological agents and AAV mutants, we have demonstrated that the classic MHC class I antigen presentation pathway plays a major role in AAV capsid antigen cross-presentation in AAV-transduced cells in vitro [6]. The mechanism of capsid antigen crosspresentation from AAV-transduced cells in vivo is perhaps different from that in vitro due to the far more complex environment
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