Clustered CD20 Induced Apoptosis: SRC-Family Kinase, the Proximal Regulator of Tyrosine Phosphorylation, Calcium Influx, and Caspase 3-dependent Apoptosis

Blood Cells Mol Dis. 2000 Apr;26(2):133-43. doi: 10.1006/bcmd.2000.0287.

Abstract

Anti-CD20 antibodies may reduce or eliminate non-Hodgkin's lymphoma B cells in patients, although the mechanism of action is not clear. To explore mechanism(s), we examined the induction of signal transduction events using anti-CD20 monoclonal antibodies (mAb) in the human non-Hodgkin's lymphoma Ramos B cell line. We found that while Rituximab (a human-mouse hybrid mAb) alone induced apoptotic cell death, other murine anti-CD20 mAbs induced apoptosis of Ramos B cells only upon clustering with a secondary antibody. CD20 clustering was accompanied by activation of tyrosine protein kinase activity, PLCgamma2 phosphorylation, influx of Ca(2+), and activation of caspase 3. All signaling events, as well as the subsequent apoptosis, were blocked by PP2, a selective inhibitor of Src-family kinases. Treatment of Ramos with EGTA and BAPTA to block changes in cytoplasmic Ca(2+) likewise prevented CD20-induced apoptosis. Our findings support a model in which CD20 clustering activates members of the Src family of protein tyrosine kinases, leading to phosphorylation of PLCgamma2 and increased cytoplasmic Ca(2+). These early signal transduction events activate caspase 3 to promote apoptotic cell death of NHL B cells.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Antibodies, Monoclonal / immunology
  • Antigens, CD20 / immunology*
  • Apoptosis / immunology*
  • Calcium / immunology
  • Caspase 3
  • Caspases / immunology*
  • Enzyme Activation
  • Humans
  • Lymphoma, B-Cell / immunology*
  • Lymphoma, B-Cell / metabolism
  • Lymphoma, B-Cell / pathology*
  • Phosphorylation
  • Signal Transduction / immunology*
  • Tumor Cells, Cultured
  • Tyrosine
  • src-Family Kinases / immunology*

Substances

  • Antibodies, Monoclonal
  • Antigens, CD20
  • Tyrosine
  • src-Family Kinases
  • CASP3 protein, human
  • Casp3 protein, mouse
  • Caspase 3
  • Caspases
  • Calcium