High-efficiency gene transfer and high-level expression of wild-type p53 in human lung cancer cells mediated by recombinant adenovirus.
A replication-defective and helper-independent recombinant p53 adenovirus was generated. The virus, Ad5CMV-p53, carries an expression cassette that contains human cytomegalovirus E1 promoter, human wild-type p53 cDNA, and SV40 early polyadenylation signal. Four human non-small-cell lung cancer cell lines representing differences in p53 configuration were used to evaluate the Ad5CMV-p53 virus. In the H358 cell line, which has a homozygous deletion of p53, the p53 gene was transferred with 97% to 100% efficiency, as detected by immunohistochemical analysis, when the cells were infected with Ad5CMV-p53 at a multiplicity of infection of 30 to 50 plaque-forming units/cell. Western blots showed that the p53 protein was expressed at a high level. The protein expression peaked at day 3 after infection and lasted for at least 15 days. Growth of the Ad5CMV-p53 virus-infected H358 cells was inhibited 79%, whereas that of noninfected cells or the cells infected with the control virus was not inhibited. Growth of cell line H322, which has a point mutation in p53, was inhibited 72% by Ad5CMV-p53, while that of cell line H460 containing wild-type p53 was less affected (28% inhibition). Tests in nude mice demonstrated that tumorigenicity of the Ad5CMV-p53-treated H358 cells was greatly inhibited. In a mouse model of orthotopic human lung cancer, the tumorigenic H226Br cells, with a point mutation in p53, were inoculated intratracheally 3 days before the virus treatment. Intratracheal instillation of Ad5CMV-p53 prevented tumor formation.(ABSTRACT TRUNCATED AT 250 WORDS)
Zhang
W W
WW
Department of Thoracic and Cardiovascular Surgery, University of Texas M.D. Anderson Cancer Center, Houston 77030, USA.
Fang
X
X
Mazur
W
W
French
B A
BA
Georges
R N
RN
Roth
J A
JA
D000260:Adenoviruses, Human / Q000235:genetics*; D000818:Animals; D002289:Carcinoma, Non-Small-Cell Lung / Q000378:metabolism / Q000473:pathology* / Q000517:prevention & control / Q000628:therapy; D003587:Cytomegalovirus / Q000235:genetics; D015967:Gene Expression Regulation, Viral; D016158:Genes, p53; D015316:Genetic Therapy / Q000379:methods*; D005822:Genetic Vectors; D006801:Humans; D008175:Lung Neoplasms / Q000378:metabolism / Q000473:pathology* / Q000517:prevention & control / Q000628:therapy; D051379:Mice; D008819:Mice, Nude; D009368:Neoplasm Transplantation; D017354:Point Mutation; D011401:Promoter Regions, Genetic; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis* / Q000235:genetics; D014162:Transfection; D014407:Tumor Cells, Cultured; D016159:Tumor Suppressor Protein p53 / Q000096:biosynthesis* / Q000235:genetics
D011993:Recombinant Fusion Proteins; D016159:Tumor Suppressor Protein p53