Induction of RKIP and Inhibition of the Transcription Repressor YY1 in B-NHL by the Novel Proteasome Inhibitor NPI-0052 and Sensitization to TRAIL-Induced Apoptosis.

Proteasome inhibitors have been shown to exert multiple effects including direct cytotoxic activity in sensitive cancer cells. In addition, proteasome inhibitors have recently been used therapeutically for certain cancers such as multiple myeloma and mantle cell lymphoma. We have recently reported that the novel proteasome inhibitor, NPI-0052 (Nereus Pharmaceuticals), can sensitize drug-resistant B-NHL tumor cells to apoptosis by various chemotherapeutic drugs. Also, NPI-0052 inhibits the NF-κB survival pathway and inhibition of NF-κB sensitizes tumor cells to TRAIL-induced apoptosis. We hypothesized that inhibition of NF-κB by NPI-0052 may also lead to sensitization of TRAIL-resistant B-NHL cells to TRAIL-mediated apoptosis. Human Burkitt’s lymphoma,Ramos cells, were treated with various concentrations of NPI-0052 (0–10ng/ml for 24h) and were then treated with recombinant human TRAIL (0–100ng/ml for 24h). The cells were examined for apoptosis by Annexin V/PI. The combination treatment resulted in significant potentiation of cytotoxicity and synergy in apoptosis was achieved. We have then examined a potential underlying mechanism of NPI-0052-mediated sensitization. The transcription repressor YY1 was recently shown to negatively regulate the transcription of the TRAIL death receptor DR5 and thus overexpression of YY1 is a resistant factor for TRAIL-induced apoptosis. Hence, treatment of Ramos cells with NPI-0052 resulted in significant inhibition of YY1 expression and upregulation of surface DR5 expression. We have also examined the effect of NPI-0052 on RKIP (Raf-kinase inhibitor protein) (Yeung et al., Molecular Cellular Biology; 21:7207, 2001) expression. Since both NPI-0052 and RKIP inhibit the NF-κB pathway, we hypothesized that treatment of Ramos cells with NPI-0052 may result in significant upregulation of RKIP expression. The findings, indeed, demonstrate that NPI-0052 significantly induced RKIP expression in Ramos cells. Overall, the findings show, for the first time, that NPI-0052 sensitizes B-NHL tumor cells to TRAIL-mediated apoptosis. Sensitization by NPI-0052 was correlated with both the induction of RKIP expression and inhibition of YY1 and upregulation of DR5 expression. Currently, TRAIL or agonist monoclonal antibodies against DR4 or DR5 are being examined clinically for anti-tumor activity. Therefore, our findings suggest the potential application of the combination of NPI-0052 and TRAIL or agonistic monoclonal antibodies against DR4 or DR5 in the treatment of tumor cells that are resistant to drugs and TRAIL.