Celecoxib leads to G2/M arrest by induction of p21 and down-regulation of cyclin B1 expression in a p53-independent manner.

A unique in vitro system has been developed in our lab that consists of normal enterocytes derived from the rat ileum (IEC-18 cells) and their transformed derivatives with c-K-ras (R1 cells), anti-sense bak (B3 cells) and cyclin D1 (D1 cells). R1 and B3 cells express high level of COX-2 protein and PGE2. IEC 18 and D1 cells express negligible amount of COX-2, and produce very low level of PGE2. A relatively low dose of celecoxib (5-10 microM) induced G2/M arrest, followed by induction of apoptosis in the transformed but not in the normal cells. Down-regulation of cyclin B1 and up-regulation of p21 expressions independent of p53 might have cause this cell cycle block. Growth inhibition was related to COX-2 function with 90-95% reduction in PGE2 production. These findings may be of clinical importance, since low concentration of celecoxib can be achieved in human serum following standard anti-inflammatory (100-200 mg bid) regime.

Attenuation of the p53 response to DNA damage by high cell density.

The p53 tumor suppressor is critical for preventing cancer progression. Numerous observations suggest that p53 function can be modulated by the cells' microenvironment. We addressed specifically the impact of cell crowding on the induction of p53 by DNA damage. We report that cell crowding attenuates markedly p53 upregulation, transcriptional activation and subsequent p53-dependent apoptosis following exposure to genotoxic stress. The p53 protein remains short-lived in confluent cultures regardless of the extent of DNA damage, even though it undergoes efficient phosphorylation on the mouse equivalent of human p53 serine 15. This inhibitory effect of cell crowding is not a secondary consequence of density-dependent cell cycle arrest (contact inhibition). Microscopic examination indicates that dense cultures display prominent cadherin-mediated cell-cell junctions, and only poor cell-matrix focal adhesions, whereas sparse cells possess conspicuous matrix adhesions and essentially no cell-cell contacts. High-density cell culture might recapitulate the microenvironment of cells in a living organism, where the response of p53 to DNA damage is reported to be low in some organs and ages. The impact of cell density on p53 activation may have important bearings on the involvement of p53 in tumor suppression and the cellular response to anticancer therapy.