Glucocorticoid-receptor-gene defects and resistance to glucocorticoid-induced apoptosis in human leukemic cell lines.
The application of glucocorticoids (GC3) in human leukemia is based on apoptosis induction but is often hampered by GC resistance. To delineate resistance mechanisms, we examined 5 GC-resistant leukemic cell lines, termed CEM-C7.R1-R5, isolated from the GC-sensitive human acute-T-cell-leukemic line, CCRF-CEM-C7, by selection in GC-containing medium. GC resistance was ascertained by analyzing cell-cycle progression, proliferation, and apoptosis. Radioreceptor assays revealed absence of ligand-binding activity in all clones, suggesting that defects in GC-receptor(GR) expression cause GC resistance. Analyses of the GR gene revealed that all but one (CEM-C7.R5) of the clones were heterozygous for the previously described L753F mutation. CEM-C7.R5 was either hemi- or homozygous for the L753F mutation and, hence, lacked a functional GR. Sequencing of the allele not carrying the L753F mutation of the other GC-resistant sub-lines revealed additional mutations in the GR gene in 3 cases: CEM-C7.R1 and R2 had a base-pair deletion in exon 9 (deltaT740) that resulted in a reading-frame shift and a pre-terminal in-frame stop. Translation of this mutant mRNA would produce a protein lacking 32 amino acids and expressing 4 altered residues at its new C terminus. CEM-C7.R3 harbored a non-sense mutation (Q710X) in exon 8, and its mRNA would be translated into a protein lacking 67 residues. Only CEM-C7.R4 cells were devoid of mutations in the coding region of the L753F negative allele. These data suggest that, in the CCRF-CEM acute-lymphatic-leukemia model, mutations in the GR-gene coding region represent one, but not the only, cause of GC resistance.
Hala
M
M
Institute for General and Experimental Pathology, Division of Molecular Pathophysiology, University of Innsbruck, School of Medicine, Austria.
Hartmann
B L
BL
Böck
G
G
Geley
S
S
Kofler
R
R
D000483:Alleles; D000595:Amino Acid Sequence; D017209:Apoptosis / Q000187:drug effects*; D019008:Drug Resistance, Neoplasm; D015972:Gene Expression Regulation, Neoplastic; D005938:Glucocorticoids / Q000008:administration & dosage*; D006801:Humans; D007938:Leukemia / Q000235:genetics* / Q000473:pathology; D008969:Molecular Sequence Data; D009154:Mutation; D011965:Receptors, Glucocorticoid / Q000235:genetics*; D014407:Tumor Cells, Cultured
D005938:Glucocorticoids; D011965:Receptors, Glucocorticoid