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Lookup NU author(s): Professor Paul RaceORCiD
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Escherichia coli nitroreductase (NTR) activates the prodrug CB1954 to a cytotoxic derivative, allowing selective sensitization of NTR-expressing cells or tumors to the prodrug. This is one of several enzyme-prodrug combinations that are under development for cancer gene therapy, and the system has now entered clinical trials. Enhancing the catalytic efficiency of NTR for CB1954 could improve its therapeutic potential. From the crystal structure of an enzyme-ligand complex, we identified nine amino acid residues within the active site that could directly influence prodrug binding and catalysis. Mutant libraries were generated for each of these residues and clones screened for their ability to sensitize E. coli to CB1954. Amino acid substitutions at six positions conferred markedly greater sensitivity to CB1954 than did the WT enzyme; the best mutants, at residue F124, resulted in ∼5-fold improvement. Using an adenovirus vector, we introduced the F124K NTR mutant into human SK-OV-3 ovarian carcinoma cells and showed it to be ∼5-fold more potent in sensitizing the cells to CB1954 at the clinically relevant prodrug concentration of 1 μM than was the WT enzyme. Enhanced mutant NTRs such as F124K should improve the efficacy of the NTR/CB1954 combination in cancer gene therapy.
Author(s): Grove JI, Lovering AL, Guise C, Race PR, Wrighton CJ, White SA, Hyde EI, Searle PF
Publication type: Article
Publication status: Published
Journal: Cancer Research
Year: 2003
Volume: 63
Issue: 17
Pages: 5532-5537
Print publication date: 01/09/2003
ISSN (print): 0008-5472
Publisher: American Association for Cancer Research
URL: https://aacrjournals.org/cancerres/article/63/17/5532/510458/Generation-of-Escherichia-Coli-Nitroreductase
PubMed id: 14500391
