Direct retroviral delivery of human cytochrome P450 2B6 for gene-directed enzyme prodrug therapy of cancer.
Human cytochrome P450 2B6 (CYP2B6) metabolizes the prodrug cyclophosphamide (CPA) to produce phosphoramide mustard that cross-links DNA leading to cell death. We have constructed a novel retroviral vector encoding CYP2B6 (designated "MetXia-P450") and used it to transduce the human tumor cell lines HT29 and T47D. MetXia-P450 transduction sensitised these cells to the cytotoxic effects of the prodrug CPA. Results from in vitro experiments demonstrated adverse effects on the clonogenic survival of cyclophosphamide-treated cells transduced with MetXia-P450. Cytotoxic activity accompanied by bystander effect was particularly evident in 3-D multicellular spheroid models suggesting that this in vitro system may be a more appropriate model for assessing the efficacy of gene directed-enzyme prodrug therapy (GDEPT). We have applied this approach in a clinically relevant gene therapy protocol on established subcutaneous tumor xenografts. These studies show for the first time the efficacy of a P450-based GDEPT strategy mediated by a direct retroviral gene transfer in vivo.
Kan
O
O
Oxford Biomedica (UK) Ltd., Medawar Centre, Oxford Science Park, Oxford OX4 4GA, UK.
Griffiths
L
L
Baban
D
D
Iqball
S
S
Uden
M
M
Spearman
H
H
Slingsby
J
J
Price
T
T
Esapa
M
M
Kingsman
S
S
Kingsman
A
A
Slade
A
A
Naylor
S
S
D000818:Animals; D018906:Antineoplastic Agents, Alkylating / Q000008:administration & dosage; D001189:Aryl Hydrocarbon Hydroxylases; D015153:Blotting, Western; D003432:Cross-Linking Reagents / Q000494:pharmacology; D003520:Cyclophosphamide / Q000008:administration & dosage / Q000378:metabolism; D065702:Cytochrome P-450 CYP2B6; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics*; D004247:DNA / Q000378:metabolism; D018014:Gene Transfer Techniques; D015316:Genetic Therapy / Q000379:methods*; D005822:Genetic Vectors / Q000378:metabolism; D006801:Humans; D020287:In Situ Nick-End Labeling; D051379:Mice; D008819:Mice, Nude; D008861:Microsomes / Q000378:metabolism; D009368:Neoplasm Transplantation; D009369:Neoplasms / Q000628:therapy*; D010089:Oxidoreductases, N-Demethylating / Q000235:genetics*; D010957:Plasmids / Q000378:metabolism; D011355:Prodrugs / Q000627:therapeutic use*; D012190:Retroviridae / Q000235:genetics*; D013997:Time Factors; D014161:Transduction, Genetic; D014407:Tumor Cells, Cultured
D018906:Antineoplastic Agents, Alkylating; D003432:Cross-Linking Reagents; D011355:Prodrugs; D003520:Cyclophosphamide; D004247:DNA; D003577:Cytochrome P-450 Enzyme System; D001189:Aryl Hydrocarbon Hydroxylases; C585599:CYP2B6 protein, human; D065702:Cytochrome P-450 CYP2B6; D010089:Oxidoreductases, N-Demethylating