Benzo[a]pyrene up-regulates cyclooxygenase-2 gene expression in oral epithelial cells.

Cyclooxygenase may be important in the pathogenesis of smoking-related cancer because it activates carcinogens and catalyzes prostaglandin biosynthesis. We determined the effects of benzo[a]pyrene (B[a]P), a polycyclic aromatic hydrocarbon in tobacco smoke, on cyclooxygenase-2 (Cox-2) mRNA, protein and synthesis of prostaglandin E2 (PGE2) in normal and transformed oral epithelial cells. Treatment with B[a]P caused a dose-dependent increase in production of PGE2, with a maximal increase of approximately 100%. Enhanced synthesis of PGE2 was associated with increased amounts of Cox-2 protein. B[a]P also caused a two-fold increase in Cox-2 mRNA in both normal and transformed cells. Transient transfections with a Cox-2 promoter construct showed that B[a]P-mediated induction of Cox-2 mRNA reflected increased transcription. Levels of Cox-1 were unaffected by B[a]P. B[e]P did not affect the synthesis of PGE2 or amounts of Cox-2. These data are important because B[a]P-mediated induction of Cox-2 may predispose to carcinogenesis by enhancing the production of mutagens and the synthesis of prostaglandins.

Transcription of cyclooxygenase-2 is enhanced in transformed mammary epithelial cells.

Cancers form more prostaglandins than the normal tissues from which they arise. Cyclooxygenase-2 (prostaglandin H synthase-2, PGHS-2, EC 1.14.99.1), an enzyme that catalyzes the formation of prostaglandins from arachidonic acid, is inducible in epithelial cells. We investigated whether transformation of mammary cells was associated with up-regulation of Cox-2 as a basis for increased production of prostaglandin E2 (PGE2) by these cells. This hypothesis was tested in two pairs of mammary cell lines between which the mode of transformation (viral versus oncogene) differed. Virally transformed RIII/Pr1 cells, which are highly tumorigenic in mice, produced markedly increased amounts of PGE2 compared to virally initiated RIII/MG cells, a weakly tumorigenic strain. Cox-2 mRNA and protein were increased concomitantly in RIII/Pr1 cells. Similarly, Ras-induced transformation of C57/MG cells resulted in increased levels of Cox-2 mRNA and protein and increased production of PGE2. Nuclear run-offs revealed increased rates of Cox-2 transcription in the virally transformed and oncogene-transformed cell lines. Transient transfection experiments demonstrated that the oncogenes src and ras up-regulated Cox-2 promoter activity. Src-mediated up-regulation of Cox-2 promoter activity was suppressed by dominant negative ras. Our data indicate that cellular transformation is associated with enhanced transcription of Cox-2 and increased production of PGE2.

Cyclooxygenase-2 overexpression and tumor formation are blocked by sulindac in a murine model of familial adenomatous polyposis.

Inducible cyclooxygenase (Cox-2), also known as prostaglandin H synthase 2 (PGH-2) is a key enzyme in the formation of prostaglandins and thromboxanes. Cox-2 is the product of an immediate-early gene that is expressed in response to growth factors, tumor promoters, or cytokines. Overexpression of Cox-2 is associated with both human colon cancers and suppression of apoptosis in cultured epithelia] cells, an activity that is reversed by the nonsteroidal anti-inflammatory drug, sulindac sulfide. To address the relationship between Cox-2, apoptosis, and tumor development in vivo, we studied C57BL/6J-Min/+(Min) mice, a strain containing a fully penetrant dominant mutation in the Apc gene, leading to the development of gastrointestinal adenomas by 110 days of age. Min mice were fed AIN-76A chow diet and given sulindac (0.5 +/- 0.1 mg/day) in drinking water. Control Min mice and homozygous C57BL/6J-+/+ normal littermates lacking the Apc mutation (+/+) were fed AIN-76A diet and given tap water to drink. At 110 days of age, all mice were sacrificed, and their intestinal tracts were examined. Control Min mice had 11.9 +/- 7.8 tumors per mouse compared to 0.1 +/- 0.1 tumors for sulindac-treated Min mice. As expected, +/+ littermates had no macroscopic tumors. Examination of histologically normal-appearing small bowel from Min animals revealed increased amounts of Cox-2 and prostaglandin E(2) compared to +/+ littermates. Using two different in situ techniques, terminal transferase-mediated dUTP nick end labeling and a direct immunoperoxidase method, Min animals also demonstrated a 27-47% decrease in enterocyte apoptosis compared to +/+ animals. Treatment with sulindac not only inhibited tumor formation but decreased small bowel Cox-2 and prostaglandin E(2) to baseline and restored normal levels of apoptosis. These data suggest that overexpression of Cox-2 is associated with tumorigenesis in the gastrointestinal epithelium, and that both are inhibited by sulindac administration.