Attenuating the growth of tumors by intratumoral administration of DNA encoding Pseudomonas exotoxin via cationic liposomes.
A gene therapy approach was taken to inhibit tumor growth by transfecting tumor cells with a plasmid encoding a truncated but active form of Pseudomonas exotoxin A (PE), using cationic lipids as the transfection reagent. Cells transfected with this plasmid express PE intracellularly and undergo apoptosis. Transfection was optimized in vitro using two cationic lipids, DOGS and DOSPER. A ratio of between 1:4 and 1:10 (wt/wt) was found to be optimal for DOSPER, and the ratio 1:4 was used for the in vivo study when a smaller injection volume was desired. Estimating the activity of the PE-encoding plasmid was done both directly, by counting cells in vitro after transfection, and by using a cytotoxicity assay, and indirectly, by cotransfecting the plasmid with a plasmid carrying a reporter beta-galactosidase gene and observing a reduction in beta-galactosidase activity with increasing amounts of the PE-encoding plasmid. The cotransfection method was found to be very sensitive, and showed transfection of cells even with 1-2 ng of the PE-encoding plasmid per 10(5) cells. Complexes of the PE-encoding plasmid together with cationic lipid were injected into tumor xenografts in athymic nude mice. The tumor growth of transfected tumors was attenuated compared with control untreated tumors or tumors transfected with a nontoxin-expressing vector. These results indicate the potential of such a treatment for attenuating solid tumor growth in vivo.
Yerushalmi
N
N
Laboratory of Molecular Biology, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892-4255, USA.
Brinkmann
U
U
Brinkmann
E
E
Pai
L
L
Pastan
I
I
D036002:ADP Ribose Transferases; D000818:Animals; D001427:Bacterial Toxins; D004269:DNA, Bacterial; D004195:Disease Models, Animal; D004337:Drug Carriers; D005098:Exotoxins / Q000008:administration & dosage / Q000235:genetics* / Q000627:therapeutic use; D015316:Genetic Therapy; D006801:Humans; D050356:Lipid Metabolism; D008081:Liposomes; D051379:Mice; D008819:Mice, Nude; D009368:Neoplasm Transplantation; D009374:Neoplasms, Experimental / Q000235:genetics / Q000628:therapy*; D014162:Transfection / Q000379:methods; D014407:Tumor Cells, Cultured; D037521:Virulence Factors; D000097668:Pseudomonas aeruginosa Exotoxin A
D036002:ADP Ribose Transferases; D001427:Bacterial Toxins; D004269:DNA, Bacterial; D004337:Drug Carriers; D005098:Exotoxins; D008081:Liposomes; D037521:Virulence Factors; D000097668:Pseudomonas aeruginosa Exotoxin A