1,N6-etheno-2′ -deoxyadenosine and 3,N4-etheno-2′-deoxycytidine detected by monoclonal antibodies in lung and liver DNA of rats exposed to vinyl chloride

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Abstract
1,N6-Etheno-2′-deoxyadenosine (εdAdo) and 3,N4-Etheno-2′-deoxycytidine (εdCyd) are formed in vitro by reaction of DNA with the electrophilic metabolites of vinyl chloride (VC), chloroethylene oxide and chloroacetaldehyde. To detect and quantitate these DNA adducts in vivo, we have raised a series of specific monoclonal antibodies (Mab). Among those, Mab EM-A-1 and Mab EM-C-1, respectively, were used for detection of εdAdo and εdCyd by competitive radioimmunoassay (RIA), following pre-separation of the etheno adducts from DNA hydrolysates by high perfonnance liquid chromatography. At 50% inhibition of tracer-antibody binding, both Mab had a detection limit of 187 fmol and antibody affinity constants (K) of 2 × 109 l/mol. The levels of εdAdo and εdĊyd were quantitated in the DNA of lung and liver tissue of young Sprague-Dawley rats exposed to 2000 p.p.m. of VC for 10 days. The εdAdo/2′-deoxyadenosine and εdCyd/2′-deoxycytidine molar ratios were 1.3 × 10−7 and 3.3 × 10−7 respectively, in lung DNA, and 5.0 × 10−8 and 1.6 × 10−7 in liver DNA. When hydrolysates of 3 mg of DNA were analyzed by RIA at 25% inhibition of tracer-antibody binding, εdAdo and εdCyd were not detected in liver DNA from untreated rats above the limiting εdAdo/2′-deoxyadenosine and εdCyd/2′-deoxycytidine molar ratios of 2.2 × 10−8 and 3.1 × 10−8, respectively.