title: Anti-CD33-antibodies labelled with the alpha-emitter Bismuth-213 kill CD33-positive acute myeloid leukaemia cells specifically by activation of caspases and break radio- and chemoresistance by inhibition of the anti-apoptotic proteins X-linked inhibitor of apoptosis protein and B-cell lymphoma-extra large.
abstract: OBJECTIVE
  The emerging interest in radioimmunotherapies employing alpha-emitters for cancer treatment like high risk-leukaemia leads to the question of how these radionuclides exhibit their cytotoxicity. To clarify the molecular mechanisms of cell death induction, we investigated the molecular effects of the alpha-emitter Bismuth-213 (Bi-213) bound to a monoclonal anti-CD33-antibody ([Bi-213]anti-CD33) on the cell cycle and on apoptosis induction in sensitive as well as in beta- and gamma-radiation-resistant CD33-positive acute myeloid leukaemia (AML) cells.
  
  
  METHODS
  The cytotoxic potential of the radioimmunoconjugate [Bi-213]anti-CD33 was analysed in the CD33-expressing human AML cell line HL-60 and in radiation- and chemoresistant HL-60-derived cell lines. Cell cycle and apoptosis induction analyses were performed via flow cytometry. Activation of apoptosis pathways was determined by immunodetection.
  
  
  RESULTS
  [Bi-213]anti-CD33 induced apoptotic cell death in CD33-positive AML cells specifically. Molecular analyses revealed that the intrinsic mitochondrial pathway of apoptosis was activated resulting in caspase-9 activation. In the apoptotic executioner cascade caspase-3 was activated and its substrate poly (ADP-ribose) polymerase (PARP) was cleaved. Notably, [Bi-213]anti-CD33 overcame radio- and chemoresistance by reversing deficient activation of apoptosis pathways in resistant CD33-positive AML cells and by the downregulation of inhibitors of apoptosis B-cell lymphoma-extra large (Bcl-xL) and X-linked inhibitor of apoptosis protein (XIAP) involved in leukaemia resistance.
  
  
  CONCLUSIONS
  [Bi-213]anti-CD33 exhibits its cytotoxic effects specifically in CD33-expressing AML cells via induction of the intrinsic, mitochondrial pathway of apoptosis. The abrogation of chemo- and radioresistances and the reactivation of apoptotic pathways seem to be promising for the treatment of patients with so far untreatable resistant AML and underline the importance of this emerging therapeutic approach of targeted alpha-therapies.
authors.affiliation: Center for Biomedical Research, University Ulm, Helmholtzstraße 8/1, 89081 Ulm, Germany; Institute for Legal Medicine, University Ulm, Prittwitzstraße 6, 89075 Ulm, Germany. Electronic address: claudia.friesen@uni-ulm.de.
authors.forename: Claudia
authors.initials: C
authors.lastname: Friesen
authors.affiliation: Center for Biomedical Research, University Ulm, Helmholtzstraße 8/1, 89081 Ulm, Germany; Institute for Legal Medicine, University Ulm, Prittwitzstraße 6, 89075 Ulm, Germany.
authors.forename: Mareike
authors.initials: M
authors.lastname: Roscher
authors.affiliation: Center for Biomedical Research, University Ulm, Helmholtzstraße 8/1, 89081 Ulm, Germany; Institute for Legal Medicine, University Ulm, Prittwitzstraße 6, 89075 Ulm, Germany.
authors.forename: Inis
authors.initials: I
authors.lastname: Hormann
authors.affiliation: Isotope Technologies Garching GmbH, Lichtenbergstraße 1, 85748 Garching, Germany.
authors.forename: Oliver
authors.initials: O
authors.lastname: Leib
authors.affiliation: Isotope Technologies Garching GmbH, Lichtenbergstraße 1, 85748 Garching, Germany.
authors.forename: Sebastian
authors.initials: S
authors.lastname: Marx
authors.affiliation: Isotope Technologies Garching GmbH, Lichtenbergstraße 1, 85748 Garching, Germany.
authors.forename: Josue
authors.initials: J
authors.lastname: Moreno
authors.affiliation: Center for Biomedical Research, University Ulm, Helmholtzstraße 8/1, 89081 Ulm, Germany; Institute for Legal Medicine, University Ulm, Prittwitzstraße 6, 89075 Ulm, Germany.
authors.forename: Erich
authors.initials: E
authors.lastname: Miltner
mesh_terms: D000911:Antibodies, Monoclonal / Q000008:administration & dosage / Q000276:immunology; D017209:Apoptosis / Q000528:radiation effects; D001729:Bismuth / Q000008:administration & dosage*; D020169:Caspases / Q000378:metabolism*; D045744:Cell Line, Tumor; D019008:Drug Resistance, Neoplasm; D018922:HL-60 Cells; D006801:Humans; D018796:Immunoconjugates / Q000008:administration & dosage*; D015470:Leukemia, Myeloid, Acute / Q000276:immunology / Q000473:pathology / Q000532:radiotherapy*; D016393:Lymphoma, B-Cell / Q000276:immunology / Q000473:pathology / Q000532:radiotherapy*; D016499:Radioimmunotherapy / Q000379:methods; D011868:Radioisotopes / Q000008:administration & dosage*; D063268:Sialic Acid Binding Ig-like Lectin 3 / Q000096:biosynthesis / Q000276:immunology; D051636:X-Linked Inhibitor of Apoptosis Protein / Q000037:antagonists & inhibitors; D051020:bcl-X Protein / Q000037:antagonists & inhibitors
chemical_list: D000911:Antibodies, Monoclonal; C494811:BCL2L1 protein, human; D018796:Immunoconjugates; D011868:Radioisotopes; D063268:Sialic Acid Binding Ig-like Lectin 3; D051636:X-Linked Inhibitor of Apoptosis Protein; C496866:XIAP protein, human; D051020:bcl-X Protein; D020169:Caspases; D001729:Bismuth