COX-2, Cell Proliferation and PMA in Head-and-Neck Cancer Cells

Abstract

Phorbol 12-myristate 13-acetate (PMA) affects a variety of cell processes, including proliferation and differentiation. In most normal cell types, PMA enhances proliferation by a protein kinase C (PKC)- dependent mechanism, while the proliferation of cancer cell lines is inhibited by PMA. PMA has been shown to induce cyclooxygenase-2 (COX-2) production in many cell lines. We examined the effect of PMA on func- tions of induced COX-2 on proliferation of head-and-neck UMSCC-22B cancer cells. PMA was found to in- hibit head-and-neck cancer cell proliferation in a cell viability assay and to induce apoptosis. Abundance of p21 protein and p53 in these cells was increased by PMA treatment. The phorbol ester increased p21 expres- sion in 22B cells through the classical PKC pathway and independently of activation of p53. In addition, PMA increased expression of COX-2 in a time-dependent, PKC-requiring manner. COX-2 siRNA increased abundance of both p53 and p21 in 22B cells and importantly potentiated the anti-proliferative effect of PMA. Confocal microscopy revealed that PMA increased nuclear accumulation of COX-2, but not COX-1, and that COX-2 formed complexes with p53, p21 and the coactivator, p300. Chromatin immunoprecipitation showed that nuclear fraction COX-2 and p53 bound to the promoter region of p21. PMA did not significantly increase PGE 2 levels in cultured medium of 22B cells, but did increase by 13-fold the levels of PGE 2 in the medium of immortalized normal (293T) cells. In conclusion, our results show that PMA inhibits cell proliferation in head-and-neck cancer cells by a mechanism that is potentiated by COX-2 siRNA. The latter results in an inc- rease of p53 and p21 in PMA-treated cancer cells. In the absence of its siRNA, COX-2 is increased in PMA- exposed cancer cells and becomes a nucleoprotein, where it complexes with p53 and p21. Thus, the anti- proliferative activity of PMA in 22B cells is moderated by concurrent induction by PMA of COX-2 accumu- lation; this moderation may be due to reduction by COX-2 of availability of p21 and p53 in the nucleus.