Electron paramagnetic resonance spectrometry evidence for bioreduction of tirapazamine to oxidising free radicals under anaerobic conditions.
Tirapazamine (SR 4233) is a bioreductive antitumour drug in Phase III clinical trial which is activated in hypoxic tumour regions to generate a cytotoxic species. Electron paramagnetic resonance (EPR) spectrometry was used to investigate directly the formation of free radicals as the result of tirapazamine reduction by NADPH-supplemented liver microsomes. Under anaerobic conditions, the tirapazamine nitroxide free radical EPR signal was not evident over a range of rat or human liver microsomal protein (1-5 mg) concentrations. However, in combination with 1,1',5, 5'-dimethylpyrolline-1-N-oxide (DMPO), a spin trap for short-lived free radicals, tirapazamine resulted in formation of a 1:1:1:1:1:1 spectrum with hyperfine splitting A(N) = 15.8 G A(H) = 22.3 G consistent with generation of DMPO-R, a carbon-centered radical adduct. Addition of DMSO increased the signal intensity of the carbon-centred radical by at least twofold. The hyperfine splitting constants associated with DMPO-R could be indicative of a tirapazamine carbon-centred radical per se or, more likely, carbon radicals from endogenous materials (or DMSO) in the biological matrix as a result of oxidative attack by the tirapazamine primary radical. Formation of DMPO-OH, the hydroxyl radical spin adduct, by tirapazamine in the absence of air indicates that liberation of a hydroxyl radical may be a consequence of tirapazamine bioreduction under anaerobic conditions. The reactivity of tirapazamine free radicals with endogenous microsomal substances to generate reactive carbon-centred radicals indicates that tirapazamine may disrupt a wide range of cellular activities.
Patterson
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The School of Pharmacy, University of London, WC1N 1AX, London, UK. medchem@cub.ulsop.ac.uk
Taiwo
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D000693:Anaerobiosis; D000818:Animals; D000970:Antineoplastic Agents / Q000378:metabolism*; D004578:Electron Spin Resonance Spectroscopy / Q000379:methods; D005609:Free Radicals / Q000378:metabolism*; D066298:In Vitro Techniques; D008297:Male; D008862:Microsomes, Liver / Q000378:metabolism*; D010084:Oxidation-Reduction; D051381:Rats; D017207:Rats, Sprague-Dawley; D013997:Time Factors; D000077704:Tirapazamine; D014227:Triazines / Q000378:metabolism*
D000970:Antineoplastic Agents; D005609:Free Radicals; D014227:Triazines; D000077704:Tirapazamine