PDBu sensitizes myeloid cells to TRAIL. (A) Expression of CD41, CD61, and glycophorin antigens on the surface of TF-1 cells cultured with or without 5 nM PDBu in the presence or absence of 25 ng/mL TRAIL for 3 days. Data from 5 experiments are expressed as means plus or minus SD of absolute numbers of surface antigens expressed per cell (MESF). *P < .05 versus control; #P < .05 TRAIL plus PDBu versus PDBu alone. (B) Apoptosis of leukemic cell lines treated for 3 days with increasing doses of PDBu (□), TRAIL (○), or both agents (▴). Cell death was determined PI staining. Data from 3 independent experiments are expressed as means plus or minus SD. The combination index (CI) was expressed as the average of the CI values obtained at the ED₅₀, ED₇₅, and ED₉₀ and indicates synergism between TRAIL and PDBu in the induction of cell death. (C) Expression of CD41, CD14, CD33, and CD34 antigens on the surface of AML cells cultured with or without 20 nM PDBu in the presence or absence of 25 ng/mL TRAIL for 3 days. Top panels show the absolute numbers of surface antigens expressed per cell (MESF). Bottom panels show the percentage of positive cells. Tendency lines are calculated as means plus or minus SD. *P < .05 versus control; #P < .05 TRAIL plus PDBu versus PDBu alone. (D) Light microscopy of primary AML cells cultured in the presence of TRAIL plus PDBu. Cells progressively acquired an enlarged and multinucleated morphology, typical of differentiating megakaryocytes (Giemsa staining; original magnification, ×40; insets, ×100). (E) Relative number of platelets in the culture of AML cells at day 7 of treatment with TRAIL, PDBu, or their combination. The number of platelets was calculated by flow cytometry on the gated CD41⁺/calcein AM⁺ population analyzed in combination with a known number of calibration beads. Data from 3 independent experiments are shown (means ± SD). *P < .05 versus control; #P < .05 TRAIL plus PDBu versus PDBu alone. (F) Apoptosis of AML cells treated for 3 days with or without 20 nM PDBu in the presence or absence of 25 ng/mL TRAIL. Cell viability is shown as percentage of the control (DMSO). Data from 11 patients with AML are expressed as means plus or minus SD. *P < .05 versus control; #P < .05 TRAIL plus PDBu versus PDBu alone. (G) Cell viability of AML cells cultured with or without 20 nM PDBu in the presence or absence of 25 ng/mL TRAIL. A representative flow cytometric detection of PI incorporation by death cells is shown.