Matching complementing functions of transformed cells with stable expression of selected viral genes for production of E1-deleted adenovirus vectors.
Production of E1-deleted adenovirus (rAd) vectors requires complementation by E1A and E1B functions provided by the production cell line. The two cell lines most commonly used for production of rAd vectors, 293 and Per.C6, were derived from human primary cells and contain contiguous E1A and E1B sequences from the Ad genome. As an alternative system, we tested complementation of rAd vectors using sequential transfection of individual E1A and E1B expression cassettes into A549 human lung tumor cells, which support highly efficient replication of wild type adenovirus. We found that E1A function could be complemented in A549 cells by the mutant E1Adl01/07, and that E1B function could be provided in such cells using only the 55K E1B gene. Production yields in the resulting producer cell line, designated SL0003, were similar to those obtained from 293 cells without generation of detectable recombinant replication competent adenovirus.
Howe
John A
JA
Biological Research, Virology, Schering-Plough Research Institute, Kenilworth, NJ 07033, USA. john.howe@spcorp.com
Pelka
Peter
P
Antelman
Doug
D
Wilson
Chris
C
Cornell
Doug
D
Hancock
Wendy
W
Ramachandra
Murali
M
Avanzini
Jenny
J
Horn
Mark
M
Wills
Ken
K
Sutjipto
Suganto
S
Ralston
Robert
R
D000256:Adenoviridae / Q000235:genetics* / Q000254:growth & development*; D017500:Adenovirus E1A Proteins / Q000096:biosynthesis / Q000235:genetics*; D017501:Adenovirus E1B Proteins / Q000096:biosynthesis / Q000235:genetics*; D015153:Blotting, Western; D002460:Cell Line; D004279:DNA, Viral / Q000032:analysis; D017930:Genes, Reporter; D005816:Genetic Complementation Test; D005822:Genetic Vectors; D006801:Humans; D047468:Immunoprecipitation; D008156:Luciferases / Q000032:analysis / Q000235:genetics; D016254:Mutagenesis, Insertional; D011995:Recombination, Genetic; D014162:Transfection; D014764:Viral Proteins / Q000032:analysis; D014779:Virus Replication
D017500:Adenovirus E1A Proteins; D017501:Adenovirus E1B Proteins; D004279:DNA, Viral; D014764:Viral Proteins; D008156:Luciferases