Herpes simplex virus thymidine kinase/ganciclovir-induced cell death is enhanced by co-expression of caspase-3 in ovarian carcinoma cells.
There is a need to enhance the efficacy of genetic prodrug activation therapy using herpes simplex virus thymidine kinase (tk) and ganciclovir (GCV) following disappointing results in early clinical trials. tk/GCV has been shown to lead to the activation of caspase-3, a potent executor of apoptosis. We demonstrate that co-expression of pro-caspase-3 with tk/GCV leads to enhanced cell death in ovarian carcinoma cells in vitro. Following transfection with recombinant adenoviral vectors encoding tk, GCV treatment leads to greater cell death in pro-caspase-3-expressing clones of SKOV3 and IGROV1 than control cells, as well as more rapid activation of caspase-3 and more rapid cleavage of PARP. Flow cytometry suggests that there is a greater degree of S-phase block in the pro-caspase-3-expressing clones than in control cells following treatment with tk/GCV. None of these effects is seen following transfection with a control adenovirus that does not encode tk. The increased cell death, early caspase-3 activation and PARP cleavage, and flow cytometric changes seen in pro-caspase-3-expressing cells can be partially inhibited by treatment with benzyloxycarbonyl-val-ala-asp fluoromethylketone, a synthetic caspase inhibitor. Our data suggest that co-expression of pro-caspase-3 may lead to a significant enhancement of the efficacy of tk/GCV therapy.
McNeish
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IA
ICRF Molecular Oncology Unit, Imperial College School of Medicine, Hammersmith Hospital, London, UK. i.mcneish@icrf.icnet.uk
Tenev
T
T
Bell
S
S
Marani
M
M
Vassaux
G
G
Lemoine
N
N
D000256:Adenoviridae / Q000235:genetics; D000590:Amino Acid Chloromethyl Ketones / Q000494:pharmacology; D000998:Antiviral Agents / Q000494:pharmacology*; D017209:Apoptosis; D015153:Blotting, Western; D053148:Caspase 3; D061945:Caspase Inhibitors; D020169:Caspases / Q000378:metabolism*; D002470:Cell Survival / Q000187:drug effects; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D017931:DNA Primers / Q000737:chemistry; D019008:Drug Resistance, Neoplasm; D005260:Female; D005434:Flow Cytometry; D015774:Ganciclovir / Q000494:pharmacology*; D015316:Genetic Therapy / Q000379:methods; D005822:Genetic Vectors; D049452:Green Fluorescent Proteins; D006801:Humans; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D008164:Luminescent Proteins / Q000378:metabolism; D010051:Ovarian Neoplasms / Q000201:enzymology / Q000473:pathology*; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D018139:Simplexvirus / Q000201:enzymology* / Q000235:genetics; D013937:Thymidine Kinase / Q000378:metabolism*; D014162:Transfection; D014407:Tumor Cells, Cultured / Q000201:enzymology*
D000590:Amino Acid Chloromethyl Ketones; D000998:Antiviral Agents; D061945:Caspase Inhibitors; D015853:Cysteine Proteinase Inhibitors; D017931:DNA Primers; D008164:Luminescent Proteins; C096713:benzyloxycarbonylvalyl-alanyl-aspartyl fluoromethyl ketone; D049452:Green Fluorescent Proteins; D007770:L-Lactate Dehydrogenase; D011065:Poly(ADP-ribose) Polymerases; D013937:Thymidine Kinase; C505407:CASP3 protein, human; D053148:Caspase 3; D020169:Caspases; D015774:Ganciclovir