Characterization of the bortezomib-adapted cell line I-45-BTZ-R. A: I-45 and I-45-BTZ-R cells were treated with bortezomib (3.13 nM to 400 nM) for 72 hours. Cell viability was determined after treatment using the XTT assay. Control cells were treated with PBS and their viability was set as 100%. Values are the mean ± SD of triplicate assays from two experiments. B: I-45 and I-45-BTZ-R cells were treated with 40 nM bortezomib for 24, 48, or 72 hours. Caspase-3 and PARP activation (cleavage) were analyzed by Western blot analysis. C: I-45 and I-45-BTZ-R cells were treated with bortezomib (25 nM to 200 nM) for 72 hours. Percentages of sub-G1 cells and cell cycle distribution were determined by flow cytometry analysis. Values are the mean ± SD of two experiments. D: I-45 and I-45-BTZ-R cells were seeded into 20 cm cell culture dishes (1 × 106 cells per dish). At days 2, 4, 6, or 8, the cells were trypsinized and stained with trypan blue. Viable cells were counted under a microscope using a hemocytometer. Values are the mean ± SD of three experiments.