Abstract
Although activated macrophages destroy cancer cells more effectively than normal cells, the facility to escape activated macrophages is a characteristic of tumor cells. One of the mechanisms responsible for the specific killing of tumor cells by macrophages is the production of the cytokine tumor necrosis factor alpha (TNF). Therefore, resistance to TNF may provide such cancer cells a selective advantage against host elimination. In the present work we explore the possibility that cyclin D1 overrides the cytostatic effect of TNF. We show that TNF induces p21^waf1^ protein in malignant melanoma A375 cells and its binding to CDK2/4 and 6 proteins, and thereby inhibiting the activity of these complexes. This inhibition leads the cells to a G1 arrest. Overexpression of cyclin D1 in these cells makes them insensitive to TNF treatment with the recovery of CDK activity, however, is unable to overcome the inhibitory action of etoposide blocking the cells on G2/M. The bypass of TNFโinduced G1 arrest seems to be related to the increase in the stability of cyclin D bound CDK complexes, increasing the total amount of CDK2/4 and 6 complexes and leading to a functional down titration of the p21^waf1^ molecules. In these conditions the TNFโinduced increase of p21^waf1^ is not sufficient to inhibit the high amount of cyclin Dโbound complexes. This hypothesis is supported by the fact that a reduction in the levels of p21^waf1^ protein, induced by the expression of a mRNA antisense against p21^waf1^, is also able to bypass of TNFโinduced arrest. Our results confirm that p21^waf1^ has an essential role in TNFโinduced arrest and that the deregulation of cyclin D1 may be one of the mechanisms to escape physiological signals to restrict tumoral growth. ยฉ 2003 WileyโLiss, Inc.