title: Requirement of apoptotic protease-activating factor-1 for bortezomib-induced apoptosis but not for Fas-mediated apoptosis in human leukemic cells.
abstract: Bortezomib is a highly selective inhibitor of the 26S proteasome and has been approved for clinical use in the treatment of relapsing and refractory multiple myeloma and mantle cell lymphoma. Clinical trials are also underway to assess the role of bortezomib in several other human malignancies, including leukemia. However, the mechanism(s) by which bortezomib acts remain to be fully understood. Here, we studied the molecular requirements of bortezomib-induced apoptosis using the human T-cell leukemic Jurkat cells stably transfected with or without shRNA against apoptotic protease-activating factor-1 (Apaf-1). The Apaf-1-deficient Jurkat T cells were resistant to bortezomib-induced apoptosis, as assessed by caspase-3 activity, poly(ADP-ribose) polymerase cleavage, phosphatidylserine externalization, and hypodiploid DNA content. In contrast, Apaf-1-deficient cells were sensitive to Fas-induced apoptosis. Bortezomib induced an upregulation of the pro-apoptotic protein Noxa, loss of mitochondrial transmembrane potential, and release of cytochrome c in cells expressing or not expressing Apaf-1. Transient silencing of Apaf-1 expression in RPMI 8402 T-cell leukemic cells also diminished bortezomib-induced apoptosis. Fas-associated death domain (FADD)-deficient Jurkat cells were resistant to Fas-mediated apoptosis yet remained sensitive to bortezomib. Our results show that bortezomib induces apoptosis by regulating pathways that are mechanistically different from those activated upon death receptor ligation. Furthermore, in silico analyses of public transcriptomics databases indicated elevated Apaf-1 expression in several hematologic malignancies, including acute lymphoblastic and myeloid leukemia. We also noted variable Apaf-1 expression in a panel of samples from patients with acute lymphoblastic leukemia. Our results suggest that the expression of Apaf-1 may be predictive of the response to proteasome inhibition.
authors.affiliation: Division of Molecular Toxicology, Institute of Environmental Medicine, Nobels väg 13, Karolinska Institutet, 171 77 Stockholm, Sweden.
authors.forename: Astrid
authors.initials: A
authors.lastname: Ottosson-Wadlund
authors.forename: Rebecca
authors.initials: R
authors.lastname: Ceder
authors.forename: Giulio
authors.initials: G
authors.lastname: Preta
authors.forename: Katja
authors.initials: K
authors.lastname: Pokrovskaja
authors.forename: Roland C
authors.initials: RC
authors.lastname: Grafström
authors.forename: Mats
authors.initials: M
authors.lastname: Heyman
authors.forename: Stefan
authors.initials: S
authors.lastname: Söderhäll
authors.forename: Dan
authors.initials: D
authors.lastname: Grandér
authors.forename: Ingrid
authors.initials: I
authors.lastname: Hedenfalk
authors.forename: John D
authors.initials: JD
authors.lastname: Robertson
authors.forename: Bengt
authors.initials: B
authors.lastname: Fadeel
mesh_terms: D000293:Adolescent; D000970:Antineoplastic Agents / Q000494:pharmacology*; D017209:Apoptosis; D053477:Apoptotic Protease-Activating Factor 1 / Q000235:genetics / Q000378:metabolism*; D001897:Boronic Acids / Q000494:pharmacology*; D000069286:Bortezomib; D053148:Caspase 3 / Q000378:metabolism; D002648:Child; D002675:Child, Preschool; D045304:Cytochromes c / Q000378:metabolism; D053200:Fas-Associated Death Domain Protein / Q000235:genetics / Q000502:physiology; D020868:Gene Silencing; D006801:Humans; D019169:Jurkat Cells; D007951:Leukemia, Myeloid / Q000378:metabolism; D053078:Membrane Potential, Mitochondrial / Q000187:drug effects; D054198:Precursor Cell Lymphoblastic Leukemia-Lymphoma / Q000188:drug therapy / Q000378:metabolism / Q000473:pathology; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D011719:Pyrazines / Q000494:pharmacology*; D034741:RNA, Small Interfering / Q000235:genetics; D059467:Transcriptome; D019014:fas Receptor / Q000502:physiology*
chemical_list: D000970:Antineoplastic Agents; D053477:Apoptotic Protease-Activating Factor 1; D001897:Boronic Acids; D053200:Fas-Associated Death Domain Protein; C498702:PMAIP1 protein, human; D019253:Proto-Oncogene Proteins c-bcl-2; D011719:Pyrazines; D034741:RNA, Small Interfering; D019014:fas Receptor; D000069286:Bortezomib; D045304:Cytochromes c; D053148:Caspase 3