Blocking the canonical NF-κB pathway with IKKβ inhibitor. (A) Chemical structure of the IKKβ inhibitor UTC. (B) CLL cells were preincubated with or without various concentrations of UTC for 1 hour. Cells were cultured with or without rhBAFF (50 ng/mL) for 24 hours, and cytoplasmic and nucleic cell lysates were prepared. Protein content was normalized to 25 μg for the cytoplasmic fraction and to 12.5 μg for the nuclear fraction. MFIR of CLL cells stained for BCMA, TACI, or BR3 was 1.6, 2.5, or 19.8, respectively. UTC inhibited the BAFF-induced nuclear translocation of p65 but not of p52. (C) Total cell lysates of CLL cells were prepared after the same treatment described. UTC inhibited the BAFF-induced phosphorylation of IκBα. (D) CLL cells were cultured with or without rhBAFF (50 ng/mL) and various concentrations of UTC for 48 hours. Results are from experiments performed on leukemia samples from each of 8 patients. The viability of CLL cells cultured with rhBAFF was significantly higher than that of CLL cells cultured in medium alone (*P < .01; Bonferroni t test). The protective effect of rhBAFF was inhibited by at least 1 μM UTC. (E) CLL cells were cultured with or without rhBAFF (50 ng/mL) and UTC (10 μM) for 48 hours. Results are from experiments performed on leukemia samples from each of 8 patients. The viability of CLL cells cultured with UTC was significantly lower than that of CLL cells cultured in medium alone (P < .001; Bonferroni t test). The capacity of BAFF to promote leukemia cell survival was not observed when the CLL cells were cultured with UTC. (F) Isolated normal B cells of healthy donors were cultured with or without rhBAFF (50 ng/mL) and UTC (10 μM) for 48 hours. Results are from studies performed on samples from each of 8 donors. No significant difference was observed in the viabilities of normal B cells cultured with or without UTC. (D-F) Error bars indicate SEM.