Epithelial ovarian cancer cells express the chemokine receptor, CXCR4, which may be associated with increased survival and metastatic potential, but the regulation of this receptor is not understood. The inflammatory cytokine tumor necrosis factor-alpha (TNF-alpha) is found in ovarian cancer biopsies and is associated with increased tumor grade. In this report, we show that CXCR4 expression on human epithelial ovarian cancer cells is associated with, and can be modulated by, TNF-alpha. Ovarian cancer cells with high endogenous expression of TNF-alpha expressed higher levels of CXCR4 mRNA and protein than cells with low TNF-alpha expression. Stimulation of ovarian cancer cell lines and primary epithelial cancer cells with TNF-alpha resulted in increased CXCR4 mRNA and protein. The TNF-alpha-stimulated increase in CXCR4 mRNA was due partly to de novo synthesis, and up-regulation of CXCR4 cell surface protein increased migration to the CXCR4 ligand CXCL12. CXCR4 mRNA and protein was down-regulated by anti-TNF-alpha antibody or by targeting TNF-alpha mRNA using RNAi. TNF-alpha stimulation activated components of the nuclear factor kappaB pathway, and overexpression of the inhibitor of kappaB also reduced CXCR4 expression. Coculture of macrophages with ovarian cancer cells also resulted in cancer cell up-regulation of CXCR4 mRNA in a TNF-alpha-dependent manner. Finally, there was a correlation between the levels of TNF-alpha and CXCR4 mRNA in clinical biopsies of ovarian cancer, and TNF-alpha protein was expressed in CXCR4-positive tumor cells. TNF-alpha is a critical mediator of tumor promotion in a number of experimental cancers. Our data suggest that one mechanism may be through nuclear factor kappaB-dependent induction of CXCR4.