title: Synergistic effects of chemotherapeutic drugs in lymphoma cells are associated with down-regulation of inhibitor of apoptosis proteins (IAPs), prostate-apoptosis-response-gene 4 (Par-4), death-associated protein (Daxx) and with enforced caspase activation.
abstract: Cytotoxic drugs mediate apoptotic tumor cell death by influencing key regulator proteins of programmed cell death. In clinical practice cytotoxic drug combinations are desired to potentiate tumor cell kill and to minimize side effects. Nevertheless, the molecular mechanisms underlying synergistic and antagonistic effects on tumor cells are still poorly understood. In order to elucidate these molecular mechanisms we established models of synergistic and antagonistic drug combinations within the same lymphoma cell lines. By combination index method we demonstrated that bendamustine in combination with either doxorubicin or mitoxantrone caused antagonistic effects on disruption of mitochondrial membrane potential as well as on the rate of apoptosis. In contrast the combination of bendamustine with cladribine acted synergistically on these parameters. By using the IC(50) (dosages causing 50% rate of apoptosis) the synergistic effect of the combination of bendamustine and cladribine was associated with an enhanced mitochondrial release of cytochrome c and Smac/DIABLO, by down-regulation of x-linked inhibitor of apoptosis (XIAP), cIAP1, Par-4 and Daxx as well as by a significantly increased activation of caspases-3, -6, -7, -8 and -9. At the same rate of apoptosis (IC(50)), the antagonistic combinations did not increase the release of cytochrome c or Smac/DIABLO, nor down-regulate the expression of XIAP, cIAP1, Par-4 and Daxx, nor increase the activation of caspases. The role of down-regulation of IAPs and of enforced caspase activation for synergism in this model was supported by the observation, that broad spectrum inhibition of caspases re-established expression of XIAP. Our study is the first to outline the molecular alterations caused by synergistic and antagonistic drug combinations within the same lymphoma cell model. The above described mechanisms were already assessable at a point where the effects of synergistic or antagonistic combinations could not yet be discriminated quantitatively by the level of apoptosis rate of the lymphoma cells.
authors.affiliation: Department of Internal Medicine III, Hematology and Oncology, Johann Wolfgang Goethe-University Hospital, 60590 Frankfurt am Main, Germany. chow@em.uni-frankfurt.de
authors.forename: Kai Uwe
authors.initials: KU
authors.lastname: Chow
authors.forename: Daniel
authors.initials: D
authors.lastname: Nowak
authors.forename: Simone
authors.initials: S
authors.lastname: Boehrer
authors.forename: Martin
authors.initials: M
authors.lastname: Ruthardt
authors.forename: Andrea
authors.initials: A
authors.lastname: Knau
authors.forename: Dieter
authors.initials: D
authors.lastname: Hoelzer
authors.forename: Paris S
authors.initials: PS
authors.lastname: Mitrou
authors.forename: Eckhart
authors.initials: E
authors.lastname: Weidmann
mesh_terms: D048868:Adaptor Proteins, Signal Transducing; D000971:Antineoplastic Combined Chemotherapy Protocols / Q000494:pharmacology*; D017209:Apoptosis; D051017:Apoptosis Regulatory Proteins; D000069461:Bendamustine Hydrochloride; D002352:Carrier Proteins / Q000378:metabolism*; D020169:Caspases / Q000378:metabolism*; D017338:Cladribine / Q000494:pharmacology; D056970:Co-Repressor Proteins; D003574:Cytochrome c Group / Q000378:metabolism; D003598:Cytoskeletal Proteins / Q000037:antagonists & inhibitors; D015536:Down-Regulation / Q000187:drug effects; D004317:Doxorubicin / Q000494:pharmacology; D004357:Drug Synergism; D004789:Enzyme Activation; D006801:Humans; D051034:Inhibitor of Apoptosis Proteins; D047908:Intracellular Signaling Peptides and Proteins; D008223:Lymphoma / Q000378:metabolism*; D024101:Mitochondrial Proteins / Q000378:metabolism; D008942:Mitoxantrone / Q000494:pharmacology; D018832:Molecular Chaperones; D009588:Nitrogen Mustard Compounds / Q000494:pharmacology; D009687:Nuclear Proteins / Q000378:metabolism*; D011506:Proteins / Q000378:metabolism; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000037:antagonists & inhibitors* / Q000378:metabolism; D014407:Tumor Cells, Cultured; D016159:Tumor Suppressor Protein p53 / Q000037:antagonists & inhibitors* / Q000378:metabolism; D044767:Ubiquitin-Protein Ligases; D051636:X-Linked Inhibitor of Apoptosis Protein
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D051017:Apoptosis Regulatory Proteins; D002352:Carrier Proteins; D056970:Co-Repressor Proteins; D003574:Cytochrome c Group; D003598:Cytoskeletal Proteins; C106985:DAXX protein, human; C412403:DIABLO protein, human; D051034:Inhibitor of Apoptosis Proteins; D047908:Intracellular Signaling Peptides and Proteins; D024101:Mitochondrial Proteins; D018832:Molecular Chaperones; D009588:Nitrogen Mustard Compounds; D009687:Nuclear Proteins; D011506:Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D016159:Tumor Suppressor Protein p53; D051636:X-Linked Inhibitor of Apoptosis Protein; C496866:XIAP protein, human; C101296:prostate apoptosis response-4 protein; D017338:Cladribine; D004317:Doxorubicin; D000069461:Bendamustine Hydrochloride; D008942:Mitoxantrone; C494833:BIRC2 protein, human; D044767:Ubiquitin-Protein Ligases; D020169:Caspases