Induction of apoptosis by apicidin, a histone deacetylase inhibitor, via the activation of mitochondria-dependent caspase cascades in human Bcr-Abl-positive leukemia cells.
OBJECTIVE
Apicidin, a histone deacetylase inhibitor, is a novel cyclic tetrapeptide that exhibits potent antiproliferative activity against various cancer cell lines. The aim of this study was to examine the potential of apicidin to induce apoptosis in human Bcr-Abl-positive leukemia cells and to assess the mechanism of apicidin-induced apoptosis.


METHODS
Cells were exposed to various concentrations of apicidin for 2-72 h, after which the levels of apoptosis, histone acetylation, mitochondrial damage, caspase activation, and Bcr-Abl expression were assessed.


RESULTS
Apicidin induced apoptosis in K562 cells in a concentration- and time-dependent manner. Similarly, apicidin notably induced the apoptosis in the primary leukemic blasts obtained from chronic myelogenous leukemia patients in blast crisis. The acetylated histone H4 levels increased in a concentration-dependent manner in the K562 cells. However, the timing of cell death caused by apicidin did not exactly correlate with the histone deacetylase inhibitory effect. The disruption of the mitochondrial membrane potential, cytochrome c release into the cytosol, and the mitochondrial Bax translocation were notably demonstrated after the apicidin treatment. Apicidin induced the proteolytic cleavage of procaspase-9, -3, -8, and poly(ADP-ribose) polymerase. Pretreatment of the K562 cells with the caspase-3 inhibitor, DEVD-CHO, completely inhibited the apicidin-induced apoptosis, suggesting that apicidin-induced apoptosis was caspase-dependent. The Fas/Fas ligand death receptor pathway was not involved in the apicidin-mediated apoptosis in K562 cells. Pretreatment of the cells with the caspase-9 inhibitor LEHD-fmk abrogated the apicidin- induced cleavage of procaspase-3, -8, and poly(ADP-ribose) polymerase. The p210 Bcr-Abl protein levels were notably decreased after the apicidin treatment, with near complete loss after 48 h. Reverse transcription-PCR assay demonstrated that the Bcr-Abl mRNA level was also remarkably decreased in a time-dependent manner.


CONCLUSIONS
These results indicate that apicidin effectively induces the apoptosis of Bcr-Abl-positive leukemia cells through the activation of the mitochondrial pathway-dependent caspase cascades. The down-regulation of Bcr-Abl mRNA might also be one of the mechanisms implicated in the apicidin-mediated apoptosis in the K562 cells. This study provides the rationale to additionally investigate apicidin as a potential therapeutic agent for the drug-resistant Bcr-Abl-positive leukemia cells.
Department of Internal Medicine, Yonsei University College of Medicine, Seoul 120-752, Korea.
June-Won
JW
Cheong
So Young
SY
Chong
Ji Yeon
JY
Kim
Ju In
JI
Eom
Hoi Kyung
HK
Jeung
Ho Young
HY
Maeng
Seung Tae
ST
Lee
Yoo Hong
YH
Min
D017304:Annexin A5 / Q000494:pharmacology; D017209:Apoptosis; D001692:Biological Transport; D015153:Blotting, Western; D053148:Caspase 3; D053181:Caspase 8; D053453:Caspase 9; D020169:Caspases / Q000378:metabolism*; D002453:Cell Cycle; D045744:Cell Line, Tumor; D004396:Coloring Agents / Q000494:pharmacology; D045304:Cytochromes c / Q000378:metabolism; D003600:Cytosol / Q000378:metabolism; D053938:DNA Fragmentation; D015536:Down-Regulation; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D016044:Fusion Proteins, bcr-abl / Q000378:metabolism*; D016313:Genes, abl / Q000235:genetics*; D056572:Histone Deacetylase Inhibitors; D006657:Histones / Q000378:metabolism; D006801:Humans; D020014:K562 Cells; D007938:Leukemia / Q000378:metabolism*; D008928:Mitochondria / Q000378:metabolism; D009842:Oligopeptides / Q000494:pharmacology; D010456:Peptides, Cyclic / Q000378:metabolism / Q000494:pharmacology*; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism; D021381:Protein Transport; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D019253:Proto-Oncogene Proteins c-bcl-2; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013997:Time Factors; D051028:bcl-2-Associated X Protein
D017304:Annexin A5; C494826:BAX protein, human; D004396:Coloring Agents; D004791:Enzyme Inhibitors; D056572:Histone Deacetylase Inhibitors; D006657:Histones; D009842:Oligopeptides; D010456:Peptides, Cyclic; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; C102351:apicidin; C108637:aspartyl-glutamyl-valyl-aspartal; D051028:bcl-2-Associated X Protein; D045304:Cytochromes c; D011065:Poly(ADP-ribose) Polymerases; D016044:Fusion Proteins, bcr-abl; C505407:CASP3 protein, human; C505530:CASP8 protein, human; C506433:CASP9 protein, human; D053148:Caspase 3; D053181:Caspase 8; D053453:Caspase 9; D020169:Caspases