Figure 6. K-7174 and bortezomib exert additive cytotoxicity against MM cell.

A. We treated RPMI8226 cells with K-7174 (10 µM), bortezomib (5 nM) or both agents, and determined chymotrypsin-like, caspase-like and trypsin-like activities. Results are represented as relative fluorescence units (RFU) with vehicle controls set at 100%. The means ± S.D. (bars) of three independent experiments are shown. Asterisks indicate p<0.01 by paired Student’s t-test. B. We cultured KMS12-BM, U266 and RPMI8226 cells in the absence or presence of K-7174, bortezomib or both agents for 48 hours at the indicated doses. Whole cell lysates were subjected to immunoblotting for ubiquitinated proteins and GAPDH (internal control). Data are representative of multiple independent experiments. C. Isobolograms of simultaneous exposure of three MM cell lines to K-7174 and bortezomib. The concentrations that produced 80% growth inhibition are expressed as 1.0 on the ordinate and abscissa of isobolograms. The envelope of additivity, surrounded by solid and broken lines, was constructed from dose–response curves of bortezomib and K-7174. The combination is regarded as additive, because all data points fall within the envelope of additivity. The isobolograms shown are representative of at least three independent experiments. Each point represents the mean of at least three independent experiments; standard deviations were less than 25% and were omitted.