title: Switching on cytotoxicity by a single mutation at the heavy chain variable region of an anti-ganglioside antibody.
abstract: 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.
authors.affiliation: Department of Immunobiology, Center of Molecular Immunology, P.O. Box 16040, Havana 11600, Cuba.
authors.forename: Yuniel
authors.initials: Y
authors.lastname: Fernández-Marrero
authors.forename: Tays
authors.initials: T
authors.lastname: Hernández
authors.forename: Lourdes
authors.initials: L
authors.lastname: Roque-Navarro
authors.forename: Ariel
authors.initials: A
authors.lastname: Talavera
authors.forename: Ernesto
authors.initials: E
authors.lastname: Moreno
authors.forename: Tania
authors.initials: T
authors.lastname: Griñán
authors.forename: Ana María
authors.initials: AM
authors.lastname: Vázquez
authors.forename: Cristina Mateo
authors.initials: CM
authors.lastname: de Acosta
authors.forename: Rolando
authors.initials: R
authors.lastname: Pérez
authors.forename: Alejandro
authors.initials: A
authors.lastname: López-Requena
mesh_terms: D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000818:Animals; D000888:Antibodies, Anti-Idiotypic / Q000276:immunology; D000911:Antibodies, Monoclonal / Q000235:genetics / Q000276:immunology* / Q000494:pharmacology; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects / Q000276:immunology; D022801:Complementarity Determining Regions / Q000235:genetics / Q000276:immunology; D004305:Dose-Response Relationship, Drug; D005678:G(M2) Ganglioside / Q000276:immunology*; D005679:G(M3) Ganglioside / Q000276:immunology*; D057809:HEK293 Cells; D006801:Humans; D007143:Immunoglobulin Heavy Chains / Q000235:genetics / Q000276:immunology*; D007130:Immunoglobulin Idiotypes / Q000276:immunology; D007135:Immunoglobulin Variable Region / Q000235:genetics / Q000276:immunology; D051379:Mice; D008969:Molecular Sequence Data; D009154:Mutation; D011485:Protein Binding / Q000276:immunology
chemical_list: D000888:Antibodies, Anti-Idiotypic; D000911:Antibodies, Monoclonal; D022801:Complementarity Determining Regions; D005679:G(M3) Ganglioside; D007143:Immunoglobulin Heavy Chains; D007130:Immunoglobulin Idiotypes; D007135:Immunoglobulin Variable Region; D005678:G(M2) Ganglioside