Activation of mitochondria and release of mitochondrial apoptogenic factors by betulinic acid.
Different classes of anticancer drugs may trigger apoptosis by acting on different subcellular targets and by activating distinct signaling pathways. Here, we report that betulinic acid (BetA) is a prototype cytotoxic agent that triggers apoptosis by a direct effect on mitochondria. In isolated mitochondria, BetA directly induces loss of transmembrane potential independent of a benzyloxycarbonyl-Val-Ala-Asp-fluoromethyl ketone-inhibitable caspase. This is inhibited by bongkrekic acid, an agent that stabilizes the permeability transition pore complex. Mitochondria undergoing BetA-induced permeability transition mediate cleavage of caspase-8 (FLICE/MACH/Mch5) and caspase-3 (CPP32/Yama) in a cell-free system. Soluble factors such as cytochrome c or apoptosis-inducing factor released from BetA-treated mitochondria are sufficient for cleavage of caspases and nuclear fragmentation. Addition of cytochrome c to cytosolic extracts results in cleavage of caspase-3, but not of caspase-8. However, supernatants of mitochondria, which have undergone permeability transition, and partially purified apoptosis-inducing factor activate both caspase-8 and caspase-3 in cytosolic extracts and suffice to activate recombinant caspase-8. These findings show that induction of mitochondrial permeability transition alone is sufficient to trigger the full apoptosis program and that some cytotoxic drugs such as BetA may induce apoptosis via a direct effect on mitochondria.
Fulda
S
S
University Children's Hospital, Prittwitzstrasse 43, D-89075 Ulm, Germany.
Scaffidi
C
C
Susin
S A
SA
Krammer
P H
PH
Kroemer
G
G
Peter
M E
ME
Debatin
K M
KM
D000972:Antineoplastic Agents, Phytogenic / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects* / Q000502:physiology; D001865:Bongkrekic Acid / Q000494:pharmacology; D053148:Caspase 3; D053181:Caspase 8; D053453:Caspase 9; D020169:Caspases / Q000378:metabolism*; D003574:Cytochrome c Group / Q000378:metabolism; D053938:DNA Fragmentation; D006801:Humans; D007425:Intracellular Membranes / Q000187:drug effects / Q000502:physiology; D007700:Kinetics; D008564:Membrane Potentials / Q000187:drug effects; D008928:Mitochondria / Q000187:drug effects / Q000502:physiology*; D009447:Neuroblastoma; D053978:Pentacyclic Triterpenes; D010539:Permeability; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D014315:Triterpenes / Q000494:pharmacology*; D014407:Tumor Cells, Cultured; D051020:bcl-X Protein; D000094062:Betulinic Acid
D000972:Antineoplastic Agents, Phytogenic; C494811:BCL2L1 protein, human; D003574:Cytochrome c Group; D053978:Pentacyclic Triterpenes; D019253:Proto-Oncogene Proteins c-bcl-2; D014315:Triterpenes; D051020:bcl-X Protein; D001865:Bongkrekic Acid; C505407:CASP3 protein, human; C505530:CASP8 protein, human; C506433:CASP9 protein, human; D053148:Caspase 3; D053181:Caspase 8; D053453:Caspase 9; D020169:Caspases; D000094062:Betulinic Acid