Switching on cytotoxicity by a single mutation at the heavy chain variable region of an anti-ganglioside antibody

Mol Immunol. 2011 Apr;48(8):1059-67. doi: 10.1016/j.molimm.2011.01.008.


Gangliosides are sialic acid-containing glycosphingolipids present in the plasma membrane of most mammalian cells. In humans, the expression of the N-glycolylated (Neu5Gc) variant of the sialic acid has been associated with malignant transformation, constituting therefore an attractive target for cancer immunotherapy. P3 monoclonal antibody (mAb) recognizes Neu5Gc-containing gangliosides, as well as sulfatides. Heavy chain CDR3 (H-CDR3) arginine residues have been shown to be crucial for ganglioside recognition, but less important for anti-idiotypic antibody binding. Here, we describe the effect on antibody reactivity of different mutations involving a single H-CDR3 acid residue. Substitution of glutamate 99 (Kabat numbering) by arginine, aspartate or serine residues resulted in no differences in anti-idiotype binding. However, the first mutation caused increased reactivity with the antigen, including a cytotoxic effect of the antibody on ganglioside-expressing cells previously unseen for the wild type antibody. Another antibody that recognizes N-glycolyl-GM3 ganglioside (GM3(Neu5Gc)), but not other glycolipids, named 14F7, exhibits also an arginine-enriched H-CDR3 and a complement-independent cell death activity. Unlike 14F7 mAb, the cytotoxicity of the P3 E(99)→R mutant antibody did not exclusively depend on ganglioside expression on tumor cells.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Amino Acid Substitution
  • Animals
  • Antibodies, Anti-Idiotypic / immunology
  • Antibodies, Monoclonal / genetics
  • Antibodies, Monoclonal / immunology*
  • Antibodies, Monoclonal / pharmacology
  • Cell Line, Tumor
  • Cell Survival / drug effects
  • Cell Survival / immunology
  • Complementarity Determining Regions / genetics
  • Complementarity Determining Regions / immunology
  • Dose-Response Relationship, Drug
  • G(M2) Ganglioside / immunology*
  • G(M3) Ganglioside / immunology*
  • HEK293 Cells
  • Humans
  • Immunoglobulin Heavy Chains / genetics
  • Immunoglobulin Heavy Chains / immunology*
  • Immunoglobulin Idiotypes / immunology
  • Immunoglobulin Variable Region / genetics
  • Immunoglobulin Variable Region / immunology
  • Mice
  • Molecular Sequence Data
  • Mutation*
  • Protein Binding / immunology


  • Antibodies, Anti-Idiotypic
  • Antibodies, Monoclonal
  • Complementarity Determining Regions
  • G(M3) Ganglioside
  • Immunoglobulin Heavy Chains
  • Immunoglobulin Idiotypes
  • Immunoglobulin Variable Region
  • G(M2) Ganglioside