Resistance of tumor cells to chemotherapeutlc agents is a major problem in cancer therapy. Continuous exposure of human histiocytic lymphoma U-937 cells to 9-nitro-camptothecin (9NC), an inhibitor of the nuclear DNA topoisomerase I, induces resistance to this drug. Because of the involvement of the nuclear factor NF-κB in the expression of several growth regulatory genes, we examined the activation of this transcription factor in 9NC-resistant U-937 cells. We found that resistance to increasing concentrations of 9NC correlated with resistance to tumor necrosis factor (TNF)- dependent activation of NF-κB. However, the constitutive synthesis of NF-κB proteins remained unaffected. Cellular resistance was not unique to TNF, as other activators of NFKB, including lnterleukin-1, phorbol ester and hydrogen peroxide, also had no effect. There was no difference between 9NC-sensitive and -resistant cells in the activation of NF-κB by okadaic acid. Other transcription factors, including AP-1 and Oct-1, were not affected in the resistant cells. When examined for the inhibitory subunit of NF-κB (IκBα), resistant cells showed a faster rate of resynthesis than the control. Interestingly, although 9NC resistance correlated with resistance to TNF-dependent NF-κB activation, TNF-dependent cytotoxicity in these cells was enhanced by several hundred fold despite a significant decrease in the number of TNF receptors. In conclusion, our results suggest that NF-κB activation may play a role in tumor cell killing by 9NC but not by TNF.