Syn-GCaMP6f AAV (Serotype 8)

Recombinant adeno-associated virus (rAAV) is the leading gene therapy vector for monogenic diseases. There are currently two FDA-approved treatments, and hundreds more are in clinical trials. Advantages include low immunotoxicity and broad cytotrophy. AAV is a member of the parvovirus family, one of the smallest and simplest viruses. It is approximately 25 nanometers in diameter and consists of a capsid surrounding a packaged genome. The capsid contains 60 proteins, a mixture of viral proteins VP1, VP2, and VP3, arranged in a pseudo-icosahedral symmetry. The three capsid proteins are generated from overlapping reading frames in the cap gene. They all contain a common VP3 sequence at the C-terminus. VP2 lacks residues 1-137 of the VP1 sequence, while VP3 lacks an additional 66 residues (i.e., 1-203) of the VP1 sequence. The VP1 unique region contains a motif homologous to the phospholipase A2 domain. The VP1 unique region and the VP1/VP2 common region are initially internalized, but at some point during infection conformational changes promote their exposure.

It is thought that each site on the icosahedral T = 1 capsid is randomly populated with VP1, VP2, or VP3, in a ratio that largely reflects their expression stoichiometry. The overall ratio for rAAV from human embryonic kidney (HEK) cells is approximately 1:1:10. Capsids from Spodoptera frugiperda (Sf9) cells expressed using baculovirus appear to be less heterogeneous (i.e., they contain more VP3 and less VP1 and/or VP2). The genome is single-stranded DNA, 4.7 kb for wild-type (WT) virus, with identical T-shaped inverted terminal repeats at the ends. The + and - strands are packaged with equal frequency. Incomplete genomes can be packaged by both WT-AAV particles and rAAV vectors. Small DNA fragments as well as heterogeneous DNA can also be packaged.