Intracellular transport of recombinant adeno-associated virus vectors

Abstract

Recombinant adeno-associated viral vectors (rAAVs) have been widely used for gene delivery in animal models, and are currently evaluated for human gene therapy after successful clinical trials in the treatment of inherited, degenerative or acquired diseases, such as Leber congenital amaurosis, Parkinson disease or heart failure. However, limitations in vector tropism, such as limited tissue specificity and insufficient transduction efficiencies of particular tissues and cell types, still preclude therapeutic applications in certain tissues. Wild-type adeno-associated viruses (AAVs) are defective viruses that require the presence of a helper virus to complete their life cycle. On the one hand, this unique property makes AAV vectors one of the safest available viral vectors for gene delivery. On the other, it also represents a potential obstacle because rAAV vectors have to overcome several biological barriers in the absence of a helper virus to transduce successfully a cell. Consequently, a better understanding of the cellular roadblocks that limit rAAV gene delivery is crucial and, during the last 15 years, numerous studies resulted in an expanding body of knowledge of the intracellular trafficking pathways of rAAV vectors. This review describes our current understanding of the mechanisms involved in rAAV attachment to target cells, endocytosis, intracellular trafficking, capsid processing, nuclear import and genome release with an emphasis on the most recent discoveries in the field and the emerging strategies used to improve the efficiency of AAV-derived vectors.

Figure 1

Model of Entry and Intracellular Trafficking of AAV Vectors. Following binding to a receptor/co-receptor complex, rAAV enters target cell through endocytosis via one or more of the following pathways: CLIC/GEEC endocytosis, clathrin-mediated endocytosis (CCP) or caveolar endocytosis (CAV). Virions are sorted towards the trans-Golgi network (TGN) along a retrograde transport pathway presumably involving trafficking via early endosomes (EE), followed by late endosomes (LE), perinuclear recycling endosomes (PNRE) or both. Capsid conformation changes and exposure of the PLA2 domain (spikes) allows cytoplasmic release from the Golgi apparatus or the Endoplasmic Reticulum (ER), and nuclear import via the nuclear pore complex (NPC). After nuclear import, intact capsids accumulate in the nucleolus (No) before mobilization into the nucleoplasm (NP) and genome release by partial uncoating. Some of the steps indicated are hypothetical and have not been conclusively proven yet.