THE PROPOSED SBIR PROJECT IS DESIGNED TO DEVELOP AN ASSAY WHICH WILL ALLOW THE DIRECT MEASUREMENT OF A COMMON CARCINOGEN BOUND TO DNA (DNA-ADDUCT) AND ESTABLISH THE CORRELATION BETWEEN THE BURDEN OF THIS ADDUCT AND THE INDUSTRIAL EXPOSURE TO THE

THE PROPOSED SBIR PROJECT IS DESIGNED TO DEVELOP AN ASSAY WHICH WILL ALLOW THE DIRECT MEASUREMENT OF A COMMON CARCINOGEN BOUND TO DNA (DNA-ADDUCT) AND ESTABLISH THE CORRELATION BETWEEN THE BURDEN OF THIS ADDUCT AND THE INDUSTRIAL EXPOSURE TO THE CARCINOGEN. THE PROJECT WILL BE PERFORMED IN THREE PHASES: ASSAY DEVELOPMENT; ASSAY OPTIMIZATION, AND CORRELATION OF ADDUCT BURDEN WITH EXPOSUREINDEX; COMMERCIALIZATION. THE THIRD PHASE WILL BE FUNDED BYA COMMERCIAL PARTNER. COMMERCIALIZATION WILL ENTAIL A SERVICE WHICH MEASURES THE DNA-ADDUCT BURDEN IN INDIVIDUALS.THE OBJECTIVE OF THIS PHASE I RESEARCH IS TO DEVELOP AND DEMONSTRATE AN ASSAY SYSTEM THAT IS CAPABLE OF MEASURING IN HUMANS THE DNA ADDUCTS OF THE PREVALENT CHEMICAL CARCINOGEN,BENZO(A)PYRENE (B(A)P). B(A)P BECOMES MUTAGENIC AND CARCINOGENIC AFTER METABOLIC ACTIVATION. THE COVALENT BINDING OF THE ACTIVATED METABOLITE TO CELLULAR DNA IS CONSIDERED TO BE AN IMPORTANT STEP IN THE INITIATION OF MALIGNANT TRANSFORMATION. IF THESE COVALENT ADDUCTS TO DNA CAN BE DIRECTLY MONITORED IN PEOPLE AT RISK TO CARCINOGEN EXPOSURE, ACTIONS CAN BE TAKEN TO PREVENT FURTHEREXPOSURE AND POSSIBLY AVOID THE INDUCTION OF CANCER AT A OF BIFUNCTIONAL REAGENTS (EDTA-PH-B[OH]2) CONTAINING A POWERFUL METAL CHELATING EDTA MOIETY LINKED TO GROUP THAT REACTS WITH THE CIS-8, 9-DIHYDRODIOL PORTION OF THE GUANOSINE ADDUCT (BPDE-DG) OF A B(A)P METABOLITE. BLOOD (10 ML) IS DRAWN FROM B(A)P EXPOSED HUMANS, LYMPHOCYTES ARE SEPARATED, AND DNA IS EXTRACTED AND ENZYMATICALLY HYDROLYSEDTO NUCLEOSIDES BY CONVENTIONAL METHODS. THE EDTA-PH-B(OH)2 IS COMPLEXED WITH A SHORT-LIVED METAL ISOTOPE (E.G.(111)IN, (99)TC) AND THEN CONDENSED WITH THE BPDE-DG TO FORM A CYCLICBORONATE ESTER. THE STABLE COMPLEX OF THE RADIOACTIVE METALEDTA-PH(OH)2 - BPDE-DG IS CHROMATOGRAPHICALLY SEPARATED FROMOTHER IMPURITIES AND DETECTED. WITH A DETECTABILITY CRITERION OF 100 DPM, 5 X 10(4) MOLECULES OF THE DNA ADDUCT CAN BE DETECTED IF (99)TC IS USED. THE GOAL OF THE PHASE I RESEARCH IS TO ATTAIN A DETECTABILITY LEVEL OF 10(7) MOLECULES SO THAT ONE B(A)P ADDUCT CAN BE DETECTED PER CELL FROM 10 ML OF BLOOD. WHEN THE CHEMICAL ASSAY IS DEMONSTRATED "IN VITRO", IT WILL BE USED TO MEASURE THE ADDUCTS IN BLOOD SAMPLES DRAWN FROM TWO COKE OVEN WORKERS. EPIDEMIOLOGY STUDIES CORRELATING THE ADDUCT BURDEN WITH CUMULATION OCCUPATION DOSE WILL BE CONDUCTED IN PHASE II.