Effect of PD184352 on AML cell lines and primary samples. (a) OCI-AML3 cells were exposed to DMSO (C) or PD184352 for 6 hours and analyzed for MAPK phosphorylation. (b) OCI-AML3 cells were cultured in the presence of DMSO or PD184352. After 96 hours, cells were assessed for viability (open circles) and DNA content (filled circles). Results show viable cells in the PD98059-treated group, expressed as percentage of the viable cells in the DMSO-treated group, and percentage of cells with a hypodiploid DNA content, respectively, and are representative of one of three independent experiments. (c) OCI-AML3 (open circles), HL-60 (filled circles), and leukemic blasts from patient 19 (open squares) were cultured in the presence of DMSO or PD184352 for 48 hours and stained for DNA content. Results are expressed as percentage of S-phase cells in the DMSO-treated group. Cell line results are representative of one of three independent experiments. (d) The MAPK phosphorylation status of AML cell lines was assessed by Western blot. The ratio of phosphorylated to total p42^MAPK was then plotted against the IC₅₀ of PD184352 for the corresponding cell lines, and regression analysis was performed (R² = 0.8933, P = 0.015). (e) CD34⁺ cells from healthy donors (n = 3) and blasts from AML patients (n = 7) were analyzed for phosphorylated MAPK by Western blot. Patient samples showing a greater than 25% decrease (from patients 14, 19, 21, and 24) and a less than 25% decrease (from patients 17, 20, and 22) in cell viability after in vitro exposure to MEK inhibitors are labeled as sensitive and resistant, respectively.