Figure 6. Effects of SCF in combination with inhibition of BCR-ABL1, KIT, PI3K and/or MEK on Mo7ep210^BCR-ABL1, Mo7e cells and primary CD34⁺ CML-CP cells.

(A) Mo7ep210^BCR-ABL1 and Mo7e cells were treated with 25 ng/ml SCF, 1 μM PPY-A, 1 μM BAW667 or their combination(s) as indicated. Viable cell numbers were measured after 72 hours. Results at 24 hours and 48 hours were comparable (not shown). Error bars represent SEM. ***p<0.001 (Student’s t-test). (B) Total cellular lysates were harvested after 30 and 60 minutes and subjected to immunoblot analysis for pKIT^Y721, pABL^Y402, pERK1/2^Y202/204, pAKT^S473, pSTAT5^Y694 and α-Tubulin (loading control). (C) Mo7ep210^BCR-ABL1 cells were treated with PPY-A in combination with 20 μM PD98059 (MEK inhibitor) or 20 μM LY294002 (PI3K inhibitor), in the presence or absence of 25 ng/mL SCF. Viable cells were measured by MTS assay at 72 hours. Error bars represent SEM. *p<0.05; ***p<0.001 (Student’s t-test). (D, E) CD34⁺ cells from newly diagnosed CML-CP patients (n=3) were incubated with or without PPY-A (1 3M) for 2 hours, followed by SCF (25 ng/mL) for 30 minutes. Aliquots of cells were analyzed for pAKT^S473 and pERK1/2^Y202/204 by immunofluorescence (D) or cultured in semisolid medium, using identical conditions, with CFU-GM colonies assessed after 15 days (E). Error bars represent SEM.*p<0.05 (Student’s t-test).