RT Journal Article
SR Electronic
T1 Diflavin Oxidoreductases Activate the Bioreductive Prodrug PR104A Under Hypoxia
JF Molecular Pharmacology
JO Mol Pharmacol
FD American Society for Pharmacology and Experimental Therapeutics
SP mol.111.073759
DO 10.1124/mol.111.073759
A1 Christopher P Guise
A1 Maria R Abbattista
A1 Smitha R Tipparaju
A1 Neil K Lambie
A1 Jiechuang Su
A1 Dan Li
A1 William R Wilson
A1 Gabi U Dachs
A1 Adam V Patterson
YR 2011
UL http://molpharm.aspetjournals.org/content/early/2011/10/07/mol.111.073759.abstract
AB The clinical agent PR-104 is converted systemically to PR-104A, a nitrogen mustard prodrug designed to target tumour hypoxia. Reductive activation of PR-104A is initiated by one-electron oxidoreductases in a process reversed by oxygen. The identity of these oxidoreductases is unknown, with the exception of cytochrome P450 reductase (POR). To identify other hypoxia-selective PR-104A reductases, nine candidate oxidoreductases were expressed in HCT116 cells. Increased PR-104A-cytotoxicity was observed in cells expressing methionine synthase reductase (MTRR), novel diflavin oxidoreductase 1 (NDOR1) and inducible nitric oxide synthase (NOS2A), in addition to POR. Plasmid-based expression of these diflavin oxidoreductases also enhanced bioreductive metabolism of PR104A in an anoxia-specific manner. Diflavin oxidoreductase-dependent PR104A metabolism was suppressed >90% by pan-flavoenzyme inhibition with diphenyliodonium chloride. Antibodies were used to quantify endogenous POR, MTRR, NDOR1 and NOS2A expression in 23 human tumour cell lines; however only POR protein was detectable and its expression correlated with anoxic PR-104A reduction (r2 = 0.712). An anti-POR monoclonal antibody was used to probe expression using human tissue microarrays; 13 of 19 cancer types expressed detectable POR with 21% of cores (185 of 874) staining positive; this heterogeneity suggests POR as a useful biomarker for PR-104A activation. Immunostaining for carbonic anhydrase 9 (CAIX), reportedly an endogenous marker of hypoxia, revealed only moderate co-expression (9.6%) of both CAIX and POR across a subset of five cancer types.