Production of recombinant adeno-associated type 5 (rAAV5) vectors using recombinant herpes simplex viruses containing rep and cap.
We developed a scaleable production system for adeno-associated virus type 5 (AAV5)-based vectors using a replication-defective recombinant herpes simplex type 1 virus (rHSV) containing the rep and cap genes of AAV5. Multiple rHSV isolates containing AAV5 rep and cap with normal or altered p5 promoter elements were constructed and tested in vector production. Compared with rAAV5 vector yields obtained by plasmid transfection, yields of rAAV5 using any of the rHSV isolates were low. Evidence suggests that the low vector yields are a consequence of the extensive and early cytopathology induced by the rHSV isolates. In addition, we found a correlation between the amount of Rep52 or Rep40 proteins and the amount of vector produced by each rHSV isolate, suggesting that packaging of vector DNA into virus particles is rate-limiting when using rHSV to generate rAAV5 vectors.
Wustner
Jason T
JT
Targeted Genetics Corporation, Sharon Hill, Pennsylvania 19079, USA.
Arnold
Susan
S
Lock
Martin
M
Richardson
James C
JC
Himes
Vaughn B
VB
Kurtzman
Gary
G
Peluso
Richard W
RW
D004261:DNA Replication; D000229:Dependovirus; D005822:Genetic Vectors; D006801:Humans; D010957:Plasmids / Q000235:genetics*; D011995:Recombination, Genetic; D018139:Simplexvirus / Q000235:genetics*; D014764:Viral Proteins / Q000235:genetics; D019065:Virus Assembly / Q000235:genetics
D014764:Viral Proteins