DNA topoisomerase II inhibition by peroxisomicine A(1) and its radical metabolite induces apoptotic cell death of HL-60 and HL-60/MX2 human leukemia cells.
Peroxisomicine A(1) (T-514) is a dimeric anthracenone first isolated from the plant Karwinskia humboldtiana. The compound presents a high and selective toxicity toward liver and skin cell cultures and is currently the subject of preclinical studies as an antitumor drug. To date, the molecular basis for its diverse biological effects remains poorly understood. To elucidate its mechanism of action, we studied its interaction with DNA and its effects on human DNA topoisomerases. Practically no interaction with DNA was detected. Peroxisomicine was found to inhibit topoisomerase II but not topoisomerase I. DNA relaxation and decatenation assays indicated that the drug interferes with the catalytic activity of topoisomerase II but does not stimulate DNA cleavage, in contrast to conventional topoisomerase poisons such as etoposide. Two human leukemia cell lines sensitive or resistant to mitoxantrone were used to assess the cytotoxicity of the toxin and its effect on the cell cycle. In both cases, peroxisomicine treatment was associated with a loss of cells from every phase of the cell cycle and was accompanied by a large increase in the sub-G1 region which is characteristic of apoptotic cells. The cell cycle changes were more pronounced with the sensitive HL-60 cells than with the resistant HL-60/MX2 cells (with reduced topoisomerase II activity), in agreement with the cytotoxicity measurements. Treatment of HL-60 cells with T-514 stimulated the cleavage of the poly(ADP-ribose) polymerase by intracellular proteases such as caspase-3. The cytometry and Western blot analyses reveal that peroxisomicine induces apoptosis in leukemia cells. In addition, we characterized a catabolite of peroxisomicine, named T-510R, in the form of a highly stable radical metabolite. The electron spin resonance and mass spectrometry data are consistent with the formation of an anionic semiquinonic radical. The oxidized product T-510R inhibits topoisomerase II with a reduced efficiency compared to the parent toxin and was found to be about 3-4 times less toxic to both the sensitive and resistant leukemia cell lines than T-514. Collectively, the results suggest that topoisomerase II inhibition plays a role in the cytotoxicity of the plant toxin peroxisomicine. Inhibition of topoisomerase II may serve as an inducing signal triggering the apoptotic cell death of leukemia cells exposed to the toxin. The dihydroxyanthracenone unit may represent a useful chemotype for the preparation of topoisomerase II-targeted anticancer agents.
Lansiaux
A
A
INSERM U-524 et Laboratoire de Pharmacologie Antitumorale du Centre Oscar Lambret, IRCL, Place de Verdun, 59045 Lille, France.
Laine
W
W
Baldeyrou
B
B
Mahieu
C
C
Wattez
N
N
Vezin
H
H
Martinez
F J
FJ
Piñeyro
A
A
Bailly
C
C
D000873:Anthracenes / Q000378:metabolism* / Q000633:toxicity*; D000972:Antineoplastic Agents, Phytogenic / Q000378:metabolism* / Q000633:toxicity*; D017209:Apoptosis / Q000187:drug effects*; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism; D004273:DNA, Neoplasm / Q000378:metabolism; D004278:DNA, Superhelical / Q000378:metabolism; D019008:Drug Resistance, Neoplasm; D004578:Electron Spin Resonance Spectroscopy; D004789:Enzyme Activation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000378:metabolism* / Q000633:toxicity*; D005609:Free Radicals / Q000378:metabolism / Q000633:toxicity; D018922:HL-60 Cells / Q000166:cytology / Q000187:drug effects / Q000201:enzymology; D006801:Humans; D007700:Kinetics; D008942:Mitoxantrone / Q000494:pharmacology; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D059004:Topoisomerase I Inhibitors; D059005:Topoisomerase II Inhibitors
D000873:Anthracenes; D000972:Antineoplastic Agents, Phytogenic; D004273:DNA, Neoplasm; D004278:DNA, Superhelical; D004791:Enzyme Inhibitors; D005609:Free Radicals; D059004:Topoisomerase I Inhibitors; D059005:Topoisomerase II Inhibitors; C075149:Karwinskia toxin T-514; D008942:Mitoxantrone; C505407:CASP3 protein, human; D053148:Caspase 3; D020169:Caspases