CD44 ligation with A3D8 antibody induces apoptosis in acute myeloid leukemia cells through binding to CD44s and clustering lipid rafts

CD44 is a cell surface antigen expressed on acute myeloid leukemia cells and is used as a marker to isolate leukemia stem cells. CD44 ligation with the antibody A3D8 has been found to induce apoptosis in human acute promyelocytic leukemia (APL) cells via activation of caspase-8. The mechanism of A3D8-induced caspase-8 activation was studied in APL NB4 cells. A3D8 induces lipid raft clustering which causes Fas aggregation as determined with a confocal microscope. A3D8-induced apoptosis is abrogated by the lipid raft disrupting agent methyl-beta-cyclodextrin and the caspase-8 inhibitor Z-IETD-fmk. Western blot analysis reveals that A3D8 binds to the standard form of CD44 (CD44s). HL-60 cells without detectable CD44s protein are not responsive to A3D8-induced apoptosis. SKNO-1 cells containing higher level of CD44s protein are more sensitive to A3D8-induced apoptosis than NB4 cells. These results indicate that A3D8 induces apoptosis in leukemia cells through caspase-8 activation by binding to CD44s protein and inducing lipid raft clustering.