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(PP149) Exploiting cytochrome p450 reductase-mediated enhancement of NLCQ-1 hypoxic cytotoxicity for use in combination with radiotherapy.
Cowen, Rachel1, Wind, Natasha1, Telfer, Brian1, Stratford, Ian1, Papadopoulou, Maria*,2, Bloomer, William2, Williams, Kaye1, 1 School of Pharmacy and Pharmaceutical Sciences, Manchester, Greater Manchester, UK2 Department of Radiation Medicine, Evanston, Illinois, USA
ABSTRACT- 4-[3-(2-nitro-1-imidazolyl)-propylamino]-7-chloroquinoline hydrochloride (NLCQ-1, NSC 709257) is a weak DNA-intercalating bioreductive compound about to enter a Phase I trial. Studies using rat liver microsomes have suggested a role for cytochrome p450 reductase (P450R) in NLCQ-1 bio-activation. As human tumour levels of P450R are heterogeneous and not substantially elevated versus normal tissue, exploitation of P450R in a therapeutic context requires a gene therapy based approach. These studies have focused upon using an adenoviral vector encoding P450R (Ad-P450R) to achieve overexpression in human tumour cell lines. MDA231 (breast) and HT1080 (fibrosarcoma) cells were infected with Ad-P450R (MOI 100) 48h prior to NLCQ-1 exposure in air or hypoxia (3h). NLCQ-1 cytotoxicity was determined 3-days later by MTT assay. Tirapazamine (TPZ) was evaluated in parallel. Infection of MDA231 and HT1080 cells with Ad-P450R resulted in P450R activities of 44±16 and 178±6 nmol cytochrome c reduced/min/mg protein that were 12 and 24 fold higher than in control cells. The NLCQ-1 hypoxic cytotoxicity ratio (HCR) increased from 7 (both cell lines) to 51 and 27-fold for MDA231 and HT1080 cells compared with uninfected controls. Ad-P450R pre-treatment yielded an increase in the TPZ HCR only in the MDA231 cells (26 versus 11 in uninfected controls). The hypoxic NLCQ-1 IC50 values following Ad-P450R treatment were 5M for both MDA231 and HT1080 cells versus 69 and 75M in uninfected controls. For TPZ the comparative values were 6M (both lines) versus 50 and 22M respectively. Preliminary in vivo studies have established that NLCQ-1 treatment (15mg/kg dailyx4) yields a 4-day growth delay in MDA231 xenografts and that direct intratumoural injection of 108 Ad-P450R particles results in a 5-fold elevation in tumour P450R activity 3 days post infection. We now aim to evaluate Ad-P450R in a combined strategy with radiotherapy to enhance the efficacy of NLCQ-1 as a radiopotentiating agent.
Key words: NLCQ-1, bioreductive, hypoxia, cytochrome p450 reductase
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