1. Proteasome inhibitors abolish cell death downstream of caspase activation during anti-microtubule drug-induced apoptosis in leukemia cells.
- Author
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Nagy K, Petak I, Imre G, Barna G, Gezane-Csorba M, Sebestyen A, Houghton JA, Mihalik R, and Kopper L
- Subjects
- Amino Acid Chloromethyl Ketones administration & dosage, Amino Acid Chloromethyl Ketones pharmacology, Apoptosis physiology, Caspase Inhibitors, Cell Cycle drug effects, Cysteine Proteinase Inhibitors administration & dosage, Drug Interactions, Enzyme Activation, Etoposide administration & dosage, Etoposide pharmacology, HL-60 Cells, HSP70 Heat-Shock Proteins biosynthesis, Humans, Leukemia, Promyelocytic, Acute enzymology, Leukemia, Promyelocytic, Acute pathology, Leupeptins administration & dosage, Leupeptins pharmacology, Microtubules drug effects, Nocodazole administration & dosage, Up-Regulation drug effects, Antineoplastic Combined Chemotherapy Protocols pharmacology, Apoptosis drug effects, Caspases metabolism, Cysteine Proteinase Inhibitors pharmacology, Leukemia, Promyelocytic, Acute drug therapy, Nocodazole pharmacology, Proteasome Inhibitors
- Abstract
Purpose: Anti-microtubule drugs and proteasome inhibitors are currently among the most intensively studied anti-tumor agents, however little is known about their pharmacological interactions at the cellular level., Materials and Methods: The human promyelocytic leukemia cell line, HL-60, was exposed to nocodazole or etoposide in combination with proteasome or caspase inhibitors. Apoptotic cell death was detected by flow cytometry as sub-G1 population. Caspase and proteasome activities were monitored by the fluorogenic substrates Ac-DEVD-AMC and Suc-LLVY-AMC, respectively, in cell lysate. Heat shock protein 70 (HSP70) expression was determined by Western blotting., Results: Nocodazole, a microtubule inhibitor, induced caspase-dependent apoptosis in the HL-60 cell line. At sub-cytotoxic concentrations, proteasome inhibitors, including MG-132 or clasto-beta-lactone, decreased nocodazole-induced apoptotic DNA fragmentation without affecting the induction of caspase-3 activity. In contrast, MG-132 decreased both DNA fragmentation and caspase activation induced by etoposide, a topoisomerase-II inhibitor. HSP70 had previously been found to inhibit apoptosis independently from caspase activation. In this study, MG-132 up-regulated HSP70 protein expression, both in the presence or absence of nocodazole., Conclusion: Proteasome inhibitors decreased anti-microtubule agent-induced apoptotic DNA fragmentation downstream of caspase-3 activation, possibly due to increased HSP70 expression. The results indicate that combination treatment with these novel anti-tumor agents in leukemia requires careful evaluation of their molecular interaction at the level of apoptosis induction.
- Published
- 2005