Use of human cancer cell lines mitochondria to explore the mechanisms of BH3 peptides and ABT-737-induced mitochondrial membrane permeabilization

Current limitations of chemotherapy include toxicity on healthy tissues and multidrug resistance of malignant cells. A number of recent anti-cancer strategies aim at targeting the mitochondrial apoptotic machinery to induce tumor cell death. In this study, we set up protocols to purify functional mi...

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Bibliographic Details
Published in:PloS one 2010-03, Vol.5 (3), p.e9924
Main Authors: Buron, Nelly, Porceddu, Mathieu, Brabant, Magali, Desgué, Diana, Racoeur, Cindy, Lassalle, Myriam, Péchoux, Christine, Rustin, Pierre, Jacotot, Etienne, Borgne-Sanchez, Annie
Format: Article
Language:English
Subjects:
Analysis
Animals
Antagonists
Apoptosis
Apoptosis-inducing factor
Bax protein
Bcl-2 protein
Bcl-x protein
BID protein
BIM protein
Biochemistry/Drug Discovery
Biotechnology
Biotechnology/Protein Chemistry and Proteomics
Biphenyl Compounds - pharmacology
Cancer
Capital punishment
Cell Biology/Cellular Death and Stress Responses
Cell death
Cell Line, Tumor
Cells (Biology)
Chelerythrine
Chemotherapy
Computer Science
Copsychus
Cross-Linking Reagents - pharmacology
Cytochrome
Cytochrome c
DIABLO protein
Female
Gene Expression Regulation, Neoplastic
Gossypol
Humans
Laboratories
Life Sciences
Major outer membrane protein
Membrane Potentials
Membranes
Mice
Mice, Inbred BALB C
Microbial drug resistance
Mitochondria
Mitochondria - metabolism
Mitochondrial DNA
Mitochondrial Membranes - drug effects
Monomers
Multidrug resistance
Nitrophenols - pharmacology
Non-Clinical Medicine/Research Methods
Oligomerization
Oligomers
Oncology
Oncology/Oncology Agents
Peptide Fragments - chemistry
Peptides
Permeability
Piperazines - pharmacology
Proteins
Proto-Oncogene Proteins - chemistry
Proto-Oncogene Proteins c-bcl-2 - metabolism
Rodents
Sulfonamides - pharmacology
Telomerase
Tissues
Toxicity
Tumor cell lines
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Summary:Current limitations of chemotherapy include toxicity on healthy tissues and multidrug resistance of malignant cells. A number of recent anti-cancer strategies aim at targeting the mitochondrial apoptotic machinery to induce tumor cell death. In this study, we set up protocols to purify functional mitochondria from various human cell lines to analyze the effect of peptidic and xenobiotic compounds described to harbour either Bcl-2 inhibition properties or toxic effects related to mitochondria. Mitochondrial inner and outer membrane permeabilization were systematically investigated in cancer cell mitochondria versus non-cancerous mitochondria. The truncated (t-) Bid protein, synthetic BH3 peptides from Bim and Bak, and the small molecule ABT-737 induced a tumor-specific and OMP-restricted mitochondrio-toxicity, while compounds like HA-14.1, YC-137, Chelerythrine, Gossypol, TW-37 or EM20-25 did not. We found that ABT-737 can induce the Bax-dependent release of apoptotic proteins (cytochrome c, Smac/Diablo and Omi/HtrA2 but not AIF) from various but not all cancer cell mitochondria. Furthermore, ABT-737 addition to isolated cancer cell mitochondria induced oligomerization of Bax and/or Bak monomers already inserted in the mitochondrial membrane. Finally immunoprecipatations indicated that ABT-737 induces Bax, Bak and Bim desequestration from Bcl-2 and Bcl-xL but not from Mcl-1L. This study investigates for the first time the mechanism of action of ABT-737 as a single agent on isolated cancer cell mitochondria. Hence, this method based on MOMP (mitochondrial outer membrane permeabilization) is an interesting screening tool, tailored for identifying Bcl-2 antagonists with selective toxicity profile against cancer cell mitochondria but devoid of toxicity against healthy mitochondria.
ISSN:1932-6203
1932-6203
DOI:10.1371/journal.pone.0009924