Autophagic degradation of BCR-ABL contributes to the generation of the antileukemic effects of As2O3 on primitive leukemic progenitors from CML patients. (A) Circulating leukemia cells from a CML patient were treated with As2O3 (2μM) for 24 hours. Before collection, cells were stained with quenched probe DQ-BSA (red), and after collection stained with either anti-ABL (blue) or anti-p62/SQSTM1 (green) as indicated, and signals were detected by confocal microscopy. Merged panels indicate overlapping images of the 3 fluorescing signals, and arrows show the colocalization of p62/SQSTM1, BCR-ABL, and lysosomal probe DQ-BSA. (B) Effects of As2O3 (0.5μM) or E64d/Pepstatin (10μM/10μM) or the indicated combinations on primitive leukemic progenitor (CFU-GM) colony formation from different CML patients were examined in clonogenic assays in methylcellulose. Data are expressed as percent control of CFU-GM colony numbers for control untreated cells and represent means ± SE of 6 independent experiments using samples from different patients. Paired t test analysis comparing the effects of As2O3 in the absence or presence of E64d/pepstatin demonstrated a paired value of P = .0017. (C) Effects of As2O3 (0.5μM) or CA-074 or the indicated combinations on primitive leukemic progenitor (CFU-GM) colony formation from different CML patients were examined in clonogenic assays in methylcellulose. Data are expressed as percent control of CFU-GM colony numbers for control untreated cells and represent means ± SE of 4 independent experiments using samples from different patients. Paired t test analysis comparing the effects of As2O3 in the absence or presence of E64d/pepstatin demonstrated a paired value of P = .0027.
