Our long term goal is to identify markers of chemical exposure in biological fluids and to develop procedure(s) by which such marker(s) can be measured in a population exposed occupationally to chemicals and thus can be advised for possible adverse effects. In the proposed research we plan to develop spectral and immunochemical methods to measure chemical exposure in an occupationally exposed population. Spectral methods will involve identification and quantitation of chemically modified amino acids in blood proteins such as al-proteinase inhibitor and hemoglobin using gas chromatography-mass spectrometry. The modified amino acid(s) will be obtained from the protein either by complete hydrolysis of the intact protein to its amino acids or components from peptide fragment(s) of partially cleaved protein obtained through peptide mapping method. The immunochemical method will involve development of enzymelinked immunosorbent assays (ELISA) using monoclonal antibodies developed either against intact covalently modified proteins or their peptide fragment(s) containing covalently modified amino acid(s). Both these methods will have the sensitivity of detection of at least in the p mole/g protein range. Industrial chemicals containing functional groups such as aldehyde (acrolein, formaldehyde, and glutaraldehyde), epoxide (ethylene oxide, propylene oxide, and styrene oxide), and hydrocarbon (benzene, styrene, and butadiene) will be used as model compounds for this study. The methods thus developed will eventually be used for human biomonitoring and could also be utilized for medical surveillance as well as for risk assessment, though not included in the present grant proposal of Chinese workers exposed to benzidine to identify confirmed and presumptive cases of bladder cancer and conduct a risk factor analysis. The screening of exposed workers will include occupational, medical, and smoking histories to identify exogenous risk factors, a limited physical examination, urinary quantitative fluorescence image analysis (QFIA) to detect DNA hyperploidy, Papanicolaou (Pap) urinary cytology, measurement of urine pH, and determination of acetylator phenotype. Individuals with a positive, suspicious, or atypical Pap or equivalent OFIA cytology outcome will undergo a urologic diagnostic evaluation with clinically indicated biopsies for localization and pathologic confirmation of genitourinary malignancy. Urine pH and acetylator phenotype vill be investigated, in conjunction with exogenous risk factors such as benzidine exposure and smoking history, as possible risk factors for bladder cancer. DNA hyperploidy, quantifiable by QFIA, is associated with bladder cancer and has previously been shown to correlate with exposure to beta-naphthyl, another potent bladder carcinogen. OFIA and Pap cytology results will be compared with each other and correlated with exogenous and endogenous risk factors to evaluate their usefulness as biological markers of bladder cancer risk. The data will be analyzed to determine whether any or all of the tests are useful as intermediate endpoint markers for bladder cancer and, if they are, results will be employed to develop individual and group profiles of bladder cancer risk. A secondary objective of this study is to determine the effectiveness of chemopreventive therapy for halting or reversing cytologic findings indicative of premalignant changes associated with bladder cancer. Individuals with positive QFIA results, which we believe to be an intermediate endpoint marker for malignancy, or positive Pap cytology will be selected to participate in a two-year chemoprevention clinical trial in which they will be administered either 4-HPR or a placebo and monitored at 6 month intervals with OFIA and Pap cytology. The proposed screening and chemoprevention studies to be conducted in China will be a collaborative project by investigators in China and the United States.
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