title: Caspase activation is required for gemcitabine activity in multiple myeloma cell lines.
abstract: The objective of this study was to determine potential mechanisms of apoptotic activity of gemcitabine, a pyrimidine nucleoside analogue, in the MM1.S multiple myeloma (MM) cell line. A MM cell line that is sensitive to glucocorticoids (MM1.S) was used for this study. Immunoblotting analysis, cell cycle assays, and annexin V staining were performed to determine whether gemcitabine induced apoptosis in this model. Furthermore, we attempted to delineate the apoptotic pathway by measuring caspase-8 and -9 activity using fluorometric assays. Loss of mitochondrial membrane potential was measured by flow cytometry. Gemcitabine treatment caused apoptosis in MM cell lines as measured by an increase in DNA cleavage, an increase in annexin V binding, a decrease in the mitochondrial membrane potential, and activation of caspase activity. Furthermore, cleavage of the caspase substrate poly(ADP-ribose) polymerase and caspase-3 activation were documented as early as 8 h after treatment with gemcitabine. Caspase-8 and -9 were activated by gemcitabine treatment in this cell line, suggesting several mechanisms of action including death receptor pathway and mitochondrial damage. The addition of interleukin 6 to MM1.S cells treated with gemcitabine offered no protection against gemcitabine-induced cell death. Gemcitabine induced apoptosis in the MM1.S cell line, and its activity required caspase activation. There is a suggestion that mitochondrial integrity is being affected with gemcitabine in this system. Gemcitabine acts independently of interleukin 6, suggesting potential important therapeutic implications in MM patients.
authors.affiliation: Division of Hematology and Oncology, Department of Medicine, Robert H. Lurie Comprehensive Cancer Center, Feinberg School of Medicine, Northwestern University Medical School, Chicago, Illinois 60611, USA.
authors.forename: Chadi
authors.initials: C
authors.lastname: Nabhan
authors.forename: Devika
authors.initials: D
authors.lastname: Gajria
authors.forename: Nancy L
authors.initials: NL
authors.lastname: Krett
authors.forename: Varsha
authors.initials: V
authors.lastname: Gandhi
authors.forename: Kulsoom
authors.initials: K
authors.lastname: Ghias
authors.forename: Steven T
authors.initials: ST
authors.lastname: Rosen
mesh_terms: D017304:Annexin A5 / Q000378:metabolism; D000964:Antimetabolites, Antineoplastic / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects; D053148:Caspase 3; D053181:Caspase 8; D053453:Caspase 9; D020169:Caspases / Q000378:metabolism*; D002453:Cell Cycle / Q000187:drug effects; D002455:Cell Division / Q000187:drug effects; D003841:Deoxycytidine / Q000031:analogs & derivatives* / Q000494:pharmacology*; D004338:Drug Combinations; D004789:Enzyme Activation; D005434:Flow Cytometry; D006801:Humans; D015151:Immunoblotting; D015850:Interleukin-6 / Q000494:pharmacology; D008564:Membrane Potentials / Q000187:drug effects; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism; D009101:Multiple Myeloma / Q000188:drug therapy* / Q000201:enzymology / Q000473:pathology; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism; D014407:Tumor Cells, Cultured / Q000187:drug effects / Q000378:metabolism / Q000473:pathology; D000093542:Gemcitabine
chemical_list: D017304:Annexin A5; D000964:Antimetabolites, Antineoplastic; D004338:Drug Combinations; D015850:Interleukin-6; D003841:Deoxycytidine; D011065:Poly(ADP-ribose) Polymerases; C505407:CASP3 protein, human; C505530:CASP8 protein, human; C506433:CASP9 protein, human; D053148:Caspase 3; D053181:Caspase 8; D053453:Caspase 9; D020169:Caspases; D000093542:Gemcitabine