Drug-induced apoptosis in B-cell chronic lymphocytic leukemia: relationship between p53 gene mutation and bcl-2/bax proteins in drug resistance.
We investigated the relationship among chemosensitivity to drug-induced apoptosis in vitro, the presence of p53 gene mutations, and the expression of bcl-2 and bax proteins in B-cells from B-cell chronic lymphocytic leukemia (B-CLL) patients. Apoptosis was induced with a camptothecin analogue, 9-amino-20(s)-camptothecin, or a purine analogue, fludarabine. Cell death was monitored by propidium iodide staining and FACS analysis. Drug-induced apoptosis in B-CLL cells was p53-independent. Immunoblot analysis of bcl-2 and bax expression revealed a correlation between drug-induced apoptosis and the ratio of endogenous levels of bcl-2 to bax proteins. B-CLL cells with none to low bcl-2/bax ratios were drug-sensitive as compared to cells with intermediate to high ratios that were drug-resistant (P = 0.015). Prior to drug treatment, bax protein migrated as a single species of 21 kDa. Following drug-induced apoptosis, anti-bax specific protein complexes of 36-42 kDa were up-regulated. Using two-dimensional gel electrophoresis, bax complexes were disrupted under reducing conditions to reveal homo- and heterodimers of 18 and 21 kDa suggesting that disulfide interactions were required for complex formation. The de novo appearance of the 18 kDa anti-bax specific protein together with its increased expression in drug-sensitive B-CLL B-cells undergoing cell death suggests a role for this protein in the regulation of apoptosis.
Thomas
A
A
Department of Pathology, New York University School of Medicine, New York 10016, USA.
El Rouby
S
S
Reed
J C
JC
Krajewski
S
S
Silber
R
R
Potmesil
M
M
Newcomb
E W
EW
D017209:Apoptosis / Q000187:drug effects / Q000235:genetics / Q000502:physiology*; D019008:Drug Resistance, Neoplasm / Q000502:physiology; D016158:Genes, p53 / Q000235:genetics*; D006801:Humans; D015451:Leukemia, Lymphocytic, Chronic, B-Cell / Q000188:drug therapy / Q000235:genetics / Q000378:metabolism* / Q000473:pathology*; D008623:Mercaptoethanol / Q000494:pharmacology; D009154:Mutation; D011518:Proto-Oncogene Proteins / Q000187:drug effects / Q000378:metabolism*; D019253:Proto-Oncogene Proteins c-bcl-2; D015854:Up-Regulation; D051028:bcl-2-Associated X Protein
C494826:BAX protein, human; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D051028:bcl-2-Associated X Protein; D008623:Mercaptoethanol