This system for generating AAV2, or recombinant Adeno Associated Virus stereotype 2, lowers the presence of herapin sulfate receptor sites by mutating viral capsid proteins. Recent developments in gene therapy have shown great potential of recombinant AAV2 as a vector, due to its low immunogenicity and simple DNA structure. Ideally, recombinant AAV2 vectors will bind to a host cell via herapin sulfate proteoglycan (HSPG) sites, allowing vectors’ DNA to integrate with the host genome. This causes the host to produce necessary proteins that will help treat a targeted genetic skeletal or brain disorder, such as Parkinson’s disease. Due to the large amount of HSPG binding sites, rAAV’s target specificity is diminished, permitting the vector to bind and infect non-targeted cells. To increase target specificity, current vector production methods decrease the amount of HSPG sites by inserting peptide ligands onto the virus’s capsid open reading frame. However, these methods result in peptide insertion sites to be imbedded within the less abundant capsid proteins, VP1 and VP2, decreasing their availability. Researchers at the University of Florida have developed an efficient system for constructing rAAV vectors by mutating specific capsid proteins. These capsid mutants increase the amount of peptide insertions available within a single capsid protein, which decreases the vector’s overall affinity for HSPG. With a lower affinity for HSPG, more specific rAAV vectors can be developed that will treat a specific hereditary disease without risks of infecting non-targeted cell populations.
Genetic recombination system to increase recombinant AAV2 specificity by changing capsid protein expression
This capsid mutant system alters the genetic structure of a specific capsid protein, increasing the availability of peptide insertions. Nucleic acid segments found on the pIM45 plasmids are adjusted to change the expression of different functional capsid proteins VP1, VP2, and VP3, found on the external shell of the virus. Consequently, mutations will change the expression of these proteins so that some are expressed more prominently than others. This increases the flexibility and control among manufacturers when inserting various peptide ligands onto the viral vector. Inserting peptide ligands of various lengths and shapes will decrease a vector’s affinity to HSPG, changing the chemical responses to different cell populations. Typical external structure alterations also allow for larger peptide ligands to be inserted without any significant changes to vector structure.
Patent Issued US 8,802,080
