Differential induction of apoptosis in undifferentiated and differentiated HL-60 cells by DNA topoisomerase I and II inhibitors.
The effects of monocytic/macrophage and granulocytic differentiation induced by phorbol myristate acetate (TPA) and all-trans retinoic acid, respectively, were tested on the induction of apoptosis in human promyelocytic leukemia HL-60 cells treated with topoisomerase I and II inhibitors. Using a filter-binding assay, we observed a strong inhibition of DNA fragmentation induced by 3- and 24-hour continuous exposure to camptothecin, VP-16, VM-26, and m-AMSA in TPA-differentiated cells. The inhibition of the typical internucleosomal DNA fragmentation was confirmed by agarose gel electrophoresis. By contrast, drug-induced DNA fragmentation was not inhibited in retinoic acid-differentiated cells, and apoptosis occurred in these cells after 4 to 5 days in the absence of drug treatment. The TPA inhibitory effect was maximal after 24 hours of treatment and was correlated with differentiation, because phorbol dibutyrate ester was active, whereas 4-alpha-TPA, a nontumor promoter that does not induce differentiation, was not active. Using alkaline elution, we observed that TPA and retinoic acid differentiation were associated with changes in topoisomerase-mediated DNA breaks that were not correlated with their differential effects on drug-induced DNA fragmentation. Moreover, TPA also inhibited DNA fragmentation induced by vinblastine, cycloheximide, calphostin C, and x-rays. Using a cell-free system, we observed that DNA fragmentation was not inhibited in nuclei from TPA-differentiated cells. Rather, inhibition of apoptosis seemed to take place in the cytoplasm. We conclude that phenotypic changes associated with TPA-induced differentiation include inactivation of a cytoplasmic activity that can induce DNA fragmentation associated with apoptosis.
Solary
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Laboratory of Molecular Pharmacology, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892.
Bertrand
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R
Kohn
K W
KW
Pommier
Y
Y
D000677:Amsacrine / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects*; D002166:Camptothecin / Q000494:pharmacology; D002454:Cell Differentiation / Q000187:drug effects; D004247:DNA / Q000187:drug effects / Q000378:metabolism; D004249:DNA Damage; D005047:Etoposide / Q000494:pharmacology; D006801:Humans; D015473:Leukemia, Promyelocytic, Acute / Q000473:pathology*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D059004:Topoisomerase I Inhibitors; D059005:Topoisomerase II Inhibitors; D014212:Tretinoin / Q000494:pharmacology; D014407:Tumor Cells, Cultured
D059004:Topoisomerase I Inhibitors; D059005:Topoisomerase II Inhibitors; D000677:Amsacrine; D014212:Tretinoin; D005047:Etoposide; D004247:DNA; D013755:Tetradecanoylphorbol Acetate; D002166:Camptothecin