Single-chain mono- and bispecific antibody derivatives with novel biological properties and antitumour activity from a COS cell transient expression system.
Single-chain antibody molecules were expressed from modified eukaryotic expression vectors as individual protein domains encoded on interchangeable cDNA cassettes. Two different single-chain antibody derivatives were constructed by linking individual light- and heavy-chain variable domains. The first was specific for the L6 tumour-associated antigen and the second was specific for human CD3. Each single-chain variable domain was genetically fused with an Fc 'tag' and expressed as a fusion protein in a COS cell transient transfection system. These single-chain antibody derivatives demonstrated specific binding to cells expressing appropriate antigen and bound with affinities similar to native antibody. The CD3 single chain molecule mediated stronger activation of PLC gamma 1 and similar levels of T-cell proliferation compared with native antibody. A bispecific Fv single-chain cassette was created by fusing the expression cassettes encoding the binding domains for L6 and CD3 single-chain molecules using oligonucleotide primers encoding a short 27-residue 'helical' peptide linker. The CD3-L6 variable domains were fused to the Fc tag and expressed in COS cells. The CD3-L6FvIg bispecific fusion protein mediated adhesion between T cells and L6-positive tumour cells, and stimulated potent T-cell proliferation and cytotoxicity against tumour cells expressing the L6 antigen.
Hayden
M S
MS
Bristol-Myers Squibb Pharmaceutical Research Institute, Department of Autoimmunity and Transplantation, Seattle, WA 98121, USA.
Linsley
P S
PS
Gayle
M A
MA
Bajorath
J
J
Brady
W A
WA
Norris
N A
NA
Fell
H P
HP
Ledbetter
J A
JA
Gilliland
L K
LK
D000595:Amino Acid Sequence; D000818:Animals; D018033:Antibodies, Bispecific / Q000235:genetics*; D000911:Antibodies, Monoclonal / Q000235:genetics* / Q000276:immunology; D000912:Antibodies, Neoplasm / Q000276:immunology*; D000918:Antibody Specificity; D000951:Antigens, Neoplasm / Q000276:immunology; D001483:Base Sequence; D017252:CD3 Complex / Q000276:immunology; D002448:Cell Adhesion; D002522:Chlorocebus aethiops; D017931:DNA Primers / Q000737:chemistry; D005822:Genetic Vectors; D006801:Humans; D000922:Immunotoxins / Q000235:genetics*; D008213:Lymphocyte Activation; D008969:Molecular Sequence Data; D011993:Recombinant Fusion Proteins / Q000235:genetics*; D013601:T-Lymphocytes / Q000276:immunology
D018033:Antibodies, Bispecific; D000911:Antibodies, Monoclonal; D000912:Antibodies, Neoplasm; D000951:Antigens, Neoplasm; D017252:CD3 Complex; D017931:DNA Primers; D000922:Immunotoxins; D011993:Recombinant Fusion Proteins