Development of monoclonal antibodies to the malondialdehyde-deoxyguanosine adduct, pyrimidopurinone

Malondialdehyde (MDA), an endogenous product of lipid peroxidation and prostaglandin biosynthesis, is mutagenic in bacterial and mammalian cells and carcinogenic in rats. In order to determine whether MDA-modified bases are formed in nucleic acids in vivo, sensitive immunoassays to detect MDA-DNA and MDA-RNA adducts are being developed in our laboratory. Murine monoclonal antibodies reactive with the MDA-deoxyguanosine adduct 3-beta-D-erythro-pentofuranosylpyrimido[1,2-alpha]purin-10(3H)-one (M(1)G-R) were prepared and characterized. Several MDA-modified nucleosides and deoxynucleosides and structural analogs were synthesized and characterized and were compared as competitive inhibitors in enzymelinked immunosorbent assays (ELISAs). Less than 5 fmol of M(1)G in MDA-modified DNA was detected in a direct ELISA, and antibody binding to the modified DNA was competitively inhibited by free M(1)G-dR. DNA from Salmonella typhimurium treated with concentrations of MDA that induce reversion to histidine prototrophy was enzymatically digested, and M(1)G-dR was quantitated by competitive ELISA. Over a range of MDA concentrations from 10 to 40 mM, the level of M(1)G residues in bacterial DNA increased from 0.2 to 2.5/10(6) base pairs.