Signaling events involved in anti-CD20-induced apoptosis of malignant human B cells.
Anti-CD20 monoclonal antibodies have been successfully employed in the clinical treatment of non-Hodgkin's lymphomas in both unmodified and radio-labeled forms. Previous publications have demonstrated that the antitumor effects of unmodified anti-CD20 mAb are mediated by several mechanisms including antibody-dependent cellular cytotoxicity, complement-mediated cell lysis, and induction of apoptosis by CD20 cross-linking. In this report, we demonstrate induction of apoptosis by three anti-CD20 monoclonal antibodies [1F5, anti-B1, and C2B8 (Rituximab)]. The magnitude of apoptosis induction was greater with the chimeric Rituximab antibody than with the murine 1F5 and anti-B1 antibodies. Apoptosis could be enhanced with any of the antibodies by cross-linking with secondary antibodies (or Fc-receptor-bearing accessory cells). The signaling events involved in anti-CD20-induced apoptosis were investigated, including activation of protein tyrosine kinases, increases in intracellular Ca2+ concentrations, caspase activation, and cleavage of caspase substrates. Our results indicate that anti-CD20-induced apoptosis can be attenuated by PP1, an inhibitor of protein tyrosine kinases Lck and Fyn, chelators of extracellular or intracellular Ca2+, and inhibitors of caspases, suggesting that anti-CD20-induced apoptosis may involve modulation of these signaling molecules. We also demonstrated that varying the expression of Bc1-2 did not affect the magnitude of anti-B1-induced apoptosis, possibly because of the sequestering effects of other Bc1-2 family members, such as Bad. These studies identify several of the signal-transduction events involved in the apoptosis of malignant B cells that transpire following ligation of CD20 by anti-CD20 antibodies in the presence of Fc-receptor-expressing cells or secondary goat anti-(mouse Ig) antibodies and which may contribute to the tumor regressions observed in mouse models and clinical trials.
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Department of Biological Structure, University of Washington Medical Center, Seattle 98195-6043, USA. dmshan@u.washington.edu
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D000590:Amino Acid Chloromethyl Ketones / Q000494:pharmacology; D000818:Animals; D000911:Antibodies, Monoclonal / Q000494:pharmacology*; D058846:Antibodies, Monoclonal, Murine-Derived; D018951:Antigens, CD20 / Q000276:immunology / Q000502:physiology*; D017209:Apoptosis / Q000187:drug effects; D001402:B-Lymphocytes / Q000473:pathology*; D002051:Burkitt Lymphoma / Q000276:immunology / Q000473:pathology*; D002118:Calcium / Q000494:pharmacology; D020013:Calcium Signaling; D002352:Carrier Proteins / Q000502:physiology; D020169:Caspases / Q000494:pharmacology; D002614:Chelating Agents / Q000494:pharmacology; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D004789:Enzyme Activation; D006801:Humans; D007141:Immunoglobulin Fc Fragments / Q000276:immunology; D015759:Ionomycin / Q000494:pharmacology; D016393:Lymphoma, B-Cell / Q000276:immunology / Q000473:pathology*; D016399:Lymphoma, T-Cell / Q000276:immunology / Q000473:pathology; D051379:Mice; D010749:Phosphoprotein Phosphatases / Q000494:pharmacology; D010766:Phosphorylation / Q000187:drug effects; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism; D011499:Protein Processing, Post-Translational / Q000187:drug effects; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000502:physiology; D011940:Receptor Aggregation; D011993:Recombinant Fusion Proteins / Q000494:pharmacology; D000069283:Rituximab; D015398:Signal Transduction; D014407:Tumor Cells, Cultured; D051025:bcl-Associated Death Protein; D019014:fas Receptor / Q000502:physiology
D000590:Amino Acid Chloromethyl Ketones; D000911:Antibodies, Monoclonal; D058846:Antibodies, Monoclonal, Murine-Derived; D018951:Antigens, CD20; C494819:BAD protein, human; C494820:Bad protein, mouse; D002352:Carrier Proteins; D002614:Chelating Agents; D015853:Cysteine Proteinase Inhibitors; D007141:Immunoglobulin Fc Fragments; D019253:Proto-Oncogene Proteins c-bcl-2; D011993:Recombinant Fusion Proteins; D051025:bcl-Associated Death Protein; C096713:benzyloxycarbonylvalyl-alanyl-aspartyl fluoromethyl ketone; D019014:fas Receptor; D000069283:Rituximab; D015759:Ionomycin; D011065:Poly(ADP-ribose) Polymerases; D011505:Protein-Tyrosine Kinases; D010749:Phosphoprotein Phosphatases; D020169:Caspases; D002118:Calcium