Adeno-Associated Virus (AAV) has emerged as a widely used delivery vehicle in gene therapy. This virus is engineered to deliver genetic material into target cells, offering a promising approach for treating various diseases. The AAV particle’s size significantly influences its effectiveness and utility in therapeutic applications. Understanding its physical characteristics provides insight into its behavior as a gene delivery tool.
The Compact Structure of AAV
The AAV particle has a small, symmetrical structure. It is non-enveloped, lacking an outer lipid membrane, and its genetic material is encased within a protein shell known as a capsid. This capsid measures 20 to 26 nanometers in diameter. A nanometer is one billionth of a meter, making it significantly smaller than many common bacteria.
The capsid is formed from 60 protein subunits, arranged in an icosahedral shape. Inside this compact protein shell, AAV carries a single-stranded DNA genome. This design contributes to the virus’s stability and its ability to withstand various environmental conditions, benefiting its use as a gene therapy vector.
Why AAV Size Matters for Gene Delivery
The compact size of AAV influences its capabilities and limitations in gene delivery. A significant constraint is its limited capacity for carrying genetic material. AAV vectors have a cargo capacity of about 4.7 kilobases (kb) of DNA. Many therapeutic genes are larger than this limit, requiring alternative delivery strategies.
Despite this size limitation, AAV’s small dimensions offer advantages for cellular interactions. Its small size allows efficient entry into various cell types. It also penetrates deeper into tissues, contributing to its broad tropism—its natural preference to infect a wide range of cell types. This supports its potential for systemic delivery, reaching multiple affected areas.
The compact structure of AAV also influences the body’s immune response. AAV elicits a mild immune response compared to larger viral vectors. This lower immunogenicity is advantageous in gene therapy, reducing the likelihood of the immune system neutralizing the vector or destroying gene-modified cells.
Overcoming Size-Related Challenges in AAV Applications
Scientists have developed strategies to address gene packaging limitations. One approach is gene splitting, also known as dual or triple vector systems. In this method, a large therapeutic gene is divided into two or three smaller segments. Each segment is then packaged into a separate AAV vector for individual delivery.
Upon entering a target cell, these multiple vectors deliver their gene halves. The cell’s machinery reassembles these segments into a complete, functional gene product, overcoming the single-vector packaging limit. This strategy can increase packaging capacity to 9 kb for dual AAV systems and up to 14 kb for triple AAV systems.
Researchers also optimize gene expression to ensure sufficient protein production from the limited genetic material. This involves using efficient promoters, which are DNA sequences that initiate gene transcription, and optimizing codon usage. Codon optimization refines the therapeutic gene’s genetic code to match target host cell preferences, leading to more efficient protein translation.
Ongoing research focuses on engineering AAV capsids to modify their properties. Modifications involve altering specific amino acid sequences on the capsid surface. The goal is to accommodate larger genetic payloads or improve targeting specificity for particular cell types, while maintaining AAV’s small size and safety profile.