Insect Cells as a Factory to Produce Adeno-Associated Virus Type 2 Vectors

TLDR: This robust system provides a simple, cost-effective method for AAV vector production in invertebrate cells and is indistinguishable from 293 cell-produced rAAV, as determined on the basis of physical properties and biologic activities.

Abstract: Recombinant adeno-associated viruses (rAAV) are produced transiently in mammalian cells usually by cotransfecting two or three plasmids containing AAV genes, adenovirus helper genes, and a vector genome. Expansion and transfection of adherent cells limit the scale of rAAV production. Efficient transfection is performed with cells on solid support media such as tissue culture plates. A large animal study or a human clinical trial may require 1015 particles of vector, depending on dose. To generate this quantity of rAAV by transfection, more than 1011 HEK293 cells may be needed, which would require about 5000 × 175 cm2 flasks. The ability to scale up rAAV production by these methods severely restricts the commercialization and use of AAV vectors. A recombinant baculovirus derived from the Autographa californica nuclear polyhedrosis virus is widely employed for large-scale production of heterologous proteins in cultured insect cells and may provide an attractive alternative. Toward this goal, we have explore...