10 The HER2 and EGFR alliance in cancer: defects in system control

Abstract

Cyclophosphamides are pro-drugs whose killing agent is produced from an aldehyde that is formed by the action of a P450 oxidation step. The mustard from the aldehyde can destroy bone marrow cells as well as the tumor. Aldehyde dehydrogenase (EC 1.2.1.3) can oxidize the aldehyde and hence inactivate the cytotoxic intermediate but bone marrow has little, if any, of the enzyme. Others have shown that over-expression of the enzyme can afford protection of the marrow. A T186S mutant of the human stomach enzyme (ALDH3) that we developed has increased activity against the aldehyde compared to the native enzyme and HeLa cells transformed with the point mutant are better protected against the killing effect of the drug. It took threefold more drug to kill 90% of the cells transformed with the mutant compared to the native enzyme (15.8 compared to 5.1 mM of a precursor of the toxic aldehyde). Analysis of molecular models makes it appear that removing the methyl group of threonine in the T186S mutant allows the bulky aldehyde to bind better. The mutant was found to be a poorer enzyme when small substrates such as benzaldehyde derivatives were investigated. Thus, the enzyme appears to be better only with large substrates such as the one produced by cyclophosphamide.