Expression of MMP-9 and integrin on AML cells and binding activity of DDGW peptide. (A) Phage displaying DDGW was allowed to bind to 4 leukemia cell lines in the absence or presence of DDGW or KKGW peptide. Insertless Fd phage was a control. The bound phages were determined by titering in bacteria. (B) Fd, DDGW, or CPCFLLGCC phages were examined for binding to bone marrow smears from AML patients and one healthy donor. The percentage of tumor cells is indicated. The bound phages were determined by bacterial infection. The results show mean ± SD. (C) OCI-AML3 cells were treated with 50 nmol/L PDBu for 30 minutes and stained with anti-α_L TS1/22, anti-α_M MEM170, and anti-β₂ R7E4 antibodies (in green in top, middle, and bottom panels, respectively) and affinity purified antibodies against MMP-9 (in red). Yellow indicates the colocalization of α_L/α_Mβ₂ integrins and MMP-9. (D) OCI-AML3 cells were transferred onto the slides coated with endothelial cells (EaHy926), and the cells were treated with 50 nmol/L PDBu for 2 hours. Coculture was stained with anti-α_M MEM170 (green) and polyclonal anti-MMP-9 H-129 (red) antibodies. Yellow indicates colocalization. (E) Cells were grown in the absence or presence of an endothelial cell monolayer and 50 nmol/L PDBu for 2 hours. The cells were stained with anti-α_L MEM83, anti-α_M MEM170, or anti-MMP-9 H-129 antibodies and analyzed by flow cytometry. Shown are mean ± SD from 3 separate experiments.