Cyclooxygenase-2, malondialdehyde and pyrimidopurinone adducts of deoxyguanosine in human colon cells

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Abstract
Cyclooxygenases (COX) catalyse the oxygenation of arachidonic acid to prostaglandin (PG) endoperoxides. Activity of one of the COX isoforms, COX-2, results in production of prostaglandin E 2 (PGE 2 ) via the endoperoxide PGH 2 . COX-2 has been implicated in the pathogenesis of colorectal cancer. Malondialdehyde (MDA) is a mutagen produced by spontaneous and enzymatic breakdown of PGH 2 . MDA reacts with DNA to form adducts, predominantly the pyrimidopurinone adduct of deoxyguanosine (M 1 G). Here the hypothesis was tested that COX-2 activity in human colon cells results in formation of MDA and generation of M 1 G adducts. M 1 G was detected in basal cultures of human non-malignant colon epithelial (HCEC) and malignant SW48, SW480, HT29 and HCA-7 colon cells, at levels from 77 to 148 adducts/10 8 nucleotides. Only HCA-7 and HT29 cells expressed COX-2 protein. Levels of M 1 G correlated significantly ( r = 0.98, P < 0.001) with those of intracellular MDA determined colorimetrically in the four malignant cell types, but neither parameter correlated with expression of COX-2 or PG biosynthesis. Induction of COX-2 expression by phorbol 12-myristate 13-acetate in HCEC cells increased PGE 2 production 20-fold and MDA concentration 3-fold. Selective inhibition of COX-2 activity in HCA-7 cells by NS-398 significantly inhibited PGE 2 production, but altered neither MDA nor M 1 G levels. Malondialdehyde treatment of HCEC cells resulted in a doubling of M 1 G levels. These results show for the first time in human colon cells that COX-2 activity is associated with formation of the endogenous mutagen, MDA. Moreover, they demonstrate the correlation between MDA concentration and M 1 G adduct levels in malignant cells.

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