REPORTER ASSAYS FOR TUMOR PROMOTERS

THE LONG TERM GOALS ARE TO DEVELOP A SIMPLE, RAPID AND INEXPENSIVE ASSAY FOR THE IDENTIFICATION OF TUMOR PROMOTING COMPOUNDS PRESENT IN THE ENVIRONMENT. THE PROPOSED METHOD CAPITALIZES ON RECENT MOLECULAR FINDINGS ON THE MECHANISM BY WHICH TUMOR PROMOTERS REGULATE THE EXPRESSION OF SPECIFICGENES. THESE GENES CONTAIN PHORBOL ESTER RESPONSIVE ELEMENTS IN THEIR 5' NON-CODING REGIONS. PLASMID CONSTRUCTSWHICH HAVE SPECIFIC REPORTER GENES DOWNSTREAM OF THESE 5' REGIONS ARE AVAILABLE. THE SPECIFIC OBJECTIVES OF THIS PROPOSAL ARE 1) TO DEVELOP SHORT-TERM SCREENING ASSAYS TO DETECT TUMOR PROMOTERS USING CELL LINES STABLY TRANSFECTED WITH SPECIFIC CONSTRUCTS CONTAINING REPORTER GENES UNDER CONTROL OF PHORBOL ESTER RESPONSIVE ELEMENTS, 2) TO ESTABLISH THE VALIDITY OF THE ASSAY USING KNOWN TUMOR PROMOTERS AND OTHER AGENTS, AND 3) TO TEST THE REPORTER ASSAY FOR TUMOR PROMOTERS USING TEST COMPOUNDS AND TO DETERMINE IF POSITIVE AGENTS STIMULATE EXPRESSION THROUGH THE PROTEIN KINASE C PATHWAY. THIS ASSAY SHOULD PROVIDE SHORT-TERM ASSAYS WHICH COULD BE AUTOMATED IN THE FUTURE FORHIGH VOLUME SCREENING. THE REPORTER GENE ASSAY FOR TUMOR PROMOTERS COULD HAVE SIGNIFICANT COMMERCIAL AND SCIENTIFIC VALUE. THIS ASSAY IN COMBINATION WITH SHORT-TERM ASSAYS FORDETECTION OF MUTAGENS (AMES TEST) WOULD BE OF EXTREME VALUE IN ASSESSING THE RISK OF CANCER CAUSING AGENTS.