Chemosensitization of Drug and Rituximab-Resistant Daudi B-NHL Clones to Drug-Induced Apoptosis by the Proteasome Inhibitor NPI-0052. (2005)

Abstract Rituximab (chimeric anti-CD20) has been used clinically, alone or in combination with chemotherapy, in the treatment of B-NHL with a good response rate. However, some patients initially do not respond to such treatment and others develop resistance to further treatments. The mechanisms governing unresponsiveness are not known. We have proposed that responsiveness to rituximab treatment may be influenced by the response of the tumor cells to rituximab-mediated cell signaling. In addition, we have reported that rituximab sensitizes B-NHL cell lines to chemotherapy-induced drug apoptosis (see review Jazirehi and Bonavida, 2005, Oncogene24:2121–2843). In an effort to investigate the resistance of NHL response to rituximab treatment, we have generated rituximab resistant clones from the B-NHL cell line Daudi. For these studies, one such clone was examined, Daudi RR1. We have recently reported that rituximab signals the wild type (wt) Daudi and inhibits the constitutively activated NF-ΚB signaling pathway; this resulted in the selective downregulation of the anti-apoptotic Bcl-xl gene expression and sensitization of the tumor cells to various chemotherapeutically drug-induced apoptosis (Jazirehi, et al., 2005, Cancer Research 65: 264–76). In contrast to wt Daudi, treatment of Daudi RR1 with rituximab failed to modify the NF-ΚB signaling pathway. Further, examination of Daudi RR1 revealed that the cells exhibited hyperactivation of the NF-ΚB signaling pathway and overexpression of Bcl-xl and also exhibited higher degree of drug resistance as compared to wt Daudi. We investigated whether inhibition of NF-ΚB activity by a novel marine-derived orally active proteasome inhibitor, NPI-0052 (Nereus Pharmaceuticals), can sensitize both wt Daudi and Daudi RR1 cells to drug-induced apoptosis. NPI-0052 inhibits all three major proteolytic functions of the proteasome with a significantly different profile compared to Velcade and is active on Velcade resistant multiple myeloma cells (Chauhan, D., et al., Proc Amer Cancer Assoc, 2005, 46: [Abstract#: 6153]). In addition, NPI-0052 inhibits NF-ΚB activity, cytokine synthesis and exhibit cytostatic and cytotoxic activity in a variety of tumor cell lines. Both Daudi and Daudi RR1 cells were treated with various concentrations of NPI-0052 (1–30 nM for 1 h) and then treated with CDDP (10–30 ug/ml for 20 h). Cell viability was determined microscopically and apoptosis was determined by PI staining for DNA hypoploidy. The findings reveal that treatment of Daudi and Daudi RR1 with single agent was not cytotoxic; however, combination treatment resulted in significant potentiation of cytotoxicity and synergy was achieved. Current studies are comparing the findings obtained with NPI-0052 with other NF-ΚB inhibitors for optimal sensitization and synergy with various chemotherapeutic drugs. The present findings demonstrate that NPI-0052 is a potent sensitizing agent that can reverse both rituximab and drug resistance of B-NHL cell lines when used in combination with conventional chemotherapeutic drugs.