KPT-9274 inhibits MM cell growth and survival and overcomes promoting effect of the BM milieu. (A) A panel of 23 HMMCLs was treated with different doses of KPT-9274 for 48 hours, and cell survival was assessed by CTG. IC₅₀ analysis was performed with GraphPad software. (B) CD138⁺ MM cells from 4 MM patients were cultured in the presence of different concentrations of KPT-9274 for 48 hours. Cell viability was assessed by CTG and expressed as percent change from untreated cells (left panel). IC₅₀ analysis is also shown (right panel). (C) H929, MM1S, and OPM2 cells were cultured with and without BMSC, and in the presence of different doses of KPT-9264 for 48 hours. Cell proliferation was assessed by (³H)-thymidine uptake assay and presented as percent change from untreated cells cultured in the absence of BMSC. (D) BM from 6 myeloma patients was diluted with RPMI to seed 400 to 8000 live cells per well into 96-well plates previously prepared with increasing concentration of KPT-9274 (1 nM-10 µM) and DMSO (up to 0.5%) as vehicle and were incubated for 24 to 72 hours. After red cell lysis, cells were stained with annexin V and CD138 mAb to identify viable myeloma cells. Viable cell number was transformed to percent inhibition relative to vehicle control. Dose-response curves were fitted to nonlinear regression analysis (left panel). IC₅₀ analysis was performed using GraphPad analysis software (right panel). (E) Left panel shows dose-response curve fitted to nonlinear regression analysis of both malignant plasma cells and normal BM cells of a representative case (patient 6). Right panel displays dose/response effect in malignant and normal BM cells of 5 myeloma patients. Data are shown as area under the curve (AUC).