Figure 7. A combination of PI3K and MEK inhibitors reduces myeloid colony formation by KMX BM.
(A–C and E) Number of total colonies observed 7 days after plating 1 × 105 BM cells from pIpC-treated KMX (A and C) or FAKMX mice (B and E) in M3231 growth factor–free methylcellulose media. The final concentration of each inhibitor in methylcellulose is as follows: BYL719, 1 μM; GS1101, 1 μM; TGX221, 1 μM; BKM120, 1 μM; NVS-PI3-5, 100 nM; MEK162, 100 nM. PI3K inhibitors were used at the highest dose at which isoform selectivity could be expected for each. Each experiment was performed 3 times. *P < 0.05, **P < 0.01, ***P < 0.001, 1-way ANOVA with Bonferroni multiple-comparison post-test. (D) Freshly harvested KMX BM cells were cultured for 1 hour in the presence of the same inhibitors, stimulated with 1 ng/ml GM-CSF for 5 minutes, lysed, and subjected to Western blotting. Lanes were run on the same gel but were noncontiguous. The experiment was performed 3 times. The membrane was stripped and incubated with anti-AKT and anti-ERK antibodies. See Supplemental Figure 5B for quantification of signal intensities.