Antisense BCR-ABL oligonucleotides induce apoptosis in the Philadelphia chromosome-positive cell line BV173.
BCR-ABL antisense oligonucleotides can specifically reduce colony formation of early hematopoietic progenitor cells from chronic myeloid leukemia (CML) patients. Little is known about the mechanism of this inhibition. We studied the inhibition of the bcr-abl oncogene using fluorescein-labeled phosphorothioate oligonucleotides in the Philadelphia chromosome-positive cell line BV173. Oligonucleotide stability, uptake, bcr-abl mRNA degradation, inhibition of cell proliferation, and cell death were studied. The oligonucleotide uptake was directly dependent on the extracellular concentration and was constant over the first 18 h of incubation. After that the uptake rate decreased. We detected a decrease in bcr-abl mRNA after 3 days of treatment with antisense oligonucleotides, but much less in controls. The controls used in the experiments were the sense oligonucleotide, equimolar amounts of sense and antisense, and an untreated control. Antisense oligonucleotides completely inhibited cell growth of BV173 cells and did not inhibit growth of HL-60 cells, whereas control oligonucleotides had no such effect on either cell line. An oligonucleotide specific for the other CML breakpoint was also effective in reducing cell growth of BV173. By the use of a DNA double staining technique to discriminate between necrotic and apoptotic cells, we detected a large number of apoptotic cells in antisense treated BV173 cultures after 5 days of treatment as compared to controls. We conclude that antisense BCR-ABL oligonucleotides reduce bcr-abl mRNA expression in BV173 cells mainly in a sequence-specific manner and induce apoptosis.
Smetsers
T F
TF
Department of Internal Medicine, University Hospital St. Radboud, Nijmegen, The Netherlands.
Skorski
T
T
van de Locht
L T
LT
Wessels
H M
HM
Pennings
A H
AH
de Witte
T
T
Calabretta
B
B
Mensink
E J
EJ
D017209:Apoptosis / Q000187:drug effects* / Q000502:physiology; D001483:Base Sequence; D016923:Cell Death; D002455:Cell Division / Q000187:drug effects; D004249:DNA Damage; D005452:Fluoresceins; D016044:Fusion Proteins, bcr-abl / Q000235:genetics*; D006801:Humans; D007700:Kinetics; D015464:Leukemia, Myelogenous, Chronic, BCR-ABL Positive / Q000188:drug therapy* / Q000378:metabolism / Q000473:pathology*; D008969:Molecular Sequence Data; D016376:Oligonucleotides, Antisense / Q000378:metabolism / Q000493:pharmacokinetics / Q000494:pharmacology*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D012680:Sensitivity and Specificity; D014407:Tumor Cells, Cultured
D005452:Fluoresceins; D016376:Oligonucleotides, Antisense; D012333:RNA, Messenger; D016044:Fusion Proteins, bcr-abl