The primary objective of this project is to provide the marketplace with a short-term test that shows a higher accuracy for detection of carcinogens than currently used assays. The goal for the experiments to be carried out at OSIP is to finish automation and to use the automated form to thoroughly validate the yeast DEL assay with 500 different chemicals of known carcinogenic and non-carcinogenic activity. In addition, different strains produced in phase II at Harvard will be validated with a smaller number of chemicals. A modular approach to validation will be utilized with small studies of 25-50 chemicals aimed at addressing specific industries and/or chemical classes. Every batch of compounds will run concurrently with positive and negative controls. This will enable the validation program to release data at intervals during the study, and to correct protocol deficiencies if they arise. The goals for experiments proposed to be carried out at Harvard are to increase the sensitivity of the yeast DEL assay similarly to how the Salmonella assay has been developed over the years. It is propose to further increase the sensitivity of the DEL assay to select carcinogens with strains that are mutated for excision repair and oxidative DNA damage defense and repair. Hew constructs that allow fluorescent of luminescent detection of the deletion events will also be constructed and tested with a number of chemicals. It is likely that these strains will significantly facilitate high-throughput screening by increasing the sensitivity of recombinant detection by several orders of magnitude. PROPOSED COMMERCIAL APPLICATION: A rapid, automated, high-throughput genotoxicology assay with improved prediction of carcinogenic potential for use in early screening of lead compounds and drug candidates.
