A cytochrome P450 2B6 meditated gene therapy strategy to enhance the effects of radiation or cyclophosphamide when combined with the bioreductive drug AQ4N.
BACKGROUND
AQ4N is metabolised in hypoxic cells by cytochrome P450s (CYPs) to the cytotoxin AQ4. Most solid tumours are known to contain regions of hypoxia whereas levels of CYPs have been found to vary considerably. Enhancement of CYP levels may be obtained using gene-directed enzyme prodrug therapy (GDEPT). We have therefore examined the potential of a CYP2B6-mediated GDEPT strategy to enhance the anti-tumour effect of the combination of AQ4N with radiation or cyclophosphamide (CPA).


METHODS
In vitro and in vivo transient transfection of human CYP2B6 +/- CYP reductase (CYPRED) was investigated in RIF-1 mouse tumours. Efficacy in vitro was assessed using the alkaline comet assay (ACA). In vivo, the time to reach 4x the treatment volume (quadrupling time; VQT) was used as the end point.


RESULTS
When CYP2B6 was transfected into RIF-1 cells and treated with AQ4N under hypoxic conditions there was a significant increase in DNA damage (measured by the ACA) compared with non-transfected cells. In vivo, a single intra-tumoural injection of a CYP2B6 vector construct significantly enhanced tumour growth delay in combination with AQ4N (100 mg/kg) and 10 Gy X-rays. AQ4N (100 mg/kg) and CPA (100 mg/kg) with CYP2B6 and CYPRED also enhanced tumour growth delay; this effect became significant when the schedule was repeated 14 days later (p = 0.0197).


CONCLUSIONS
The results show the efficacy of a CYP2B6-mediated GDEPT strategy for bioreduction of AQ4N; this may offer an additional approach to target radiation- and chemo-resistant hypoxic tumours that should enhance overall tumour control.
McErlane
Verna
V
Radiation Science Research Group, School of Biomedical Sciences, University of Ulster, Cromore Road, Coleraine, Co. Londonderry BT52 1SA, UK.
Yakkundi
Anita
A
McCarthy
Helen O
HO
Hughes
Ciara M
CM
Patterson
Laurence H
LH
Hirst
David G
DG
Robson
Tracy
T
McKeown
Stephanie R
SR
D000818:Animals; D000880:Anthraquinones / Q000378:metabolism / Q000494:pharmacology; D001189:Aryl Hydrocarbon Hydroxylases / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology; D015687:Cell Hypoxia / Q000187:drug effects; D003131:Combined Modality Therapy; D003520:Cyclophosphamide / Q000008:administration & dosage / Q000627:therapeutic use; D065702:Cytochrome P-450 CYP2B6; D004249:DNA Damage / Q000187:drug effects / Q000528:radiation effects; D004273:DNA, Neoplasm / Q000187:drug effects; D005354:Fibrosarcoma / Q000628:therapy; D015316:Genetic Therapy / Q000379:methods*; D006801:Humans; D051379:Mice; D008809:Mice, Inbred C3H; D009251:NADPH-Ferrihemoprotein Reductase / Q000378:metabolism; D010089:Oxidoreductases, N-Demethylating; D011355:Prodrugs / Q000378:metabolism* / Q000494:pharmacology; D011878:Radiotherapy; D011994:Recombinant Proteins / Q000032:analysis; D014162:Transfection; D019076:Transgenes; D014407:Tumor Cells, Cultured
D000880:Anthraquinones; D004273:DNA, Neoplasm; D011355:Prodrugs; D011994:Recombinant Proteins; C094428:AQ4N; D003520:Cyclophosphamide; D001189:Aryl Hydrocarbon Hydroxylases; C585599:CYP2B6 protein, human; D065702:Cytochrome P-450 CYP2B6; D010089:Oxidoreductases, N-Demethylating; D009251:NADPH-Ferrihemoprotein Reductase