The production and preclinical characterization of a chimeric anti-breast-cancer antibody, cBC2.
A chimeric (mouse-human) BC2 antibody (cBC2) was produced which may be used in the diagnosis and treatment of breast cancer. The BC2 variable region genes were amplified by polymerase chain reaction (PCR), using oligonucleotide primers homologous to the framework sequences of mouse VH and V kappa genes. The PCR products were used to create cBC2 expression vectors containing the mouse BC2 VH and V kappa and human constant region (IgG1 and K) genes. Chimeric antibody was produced following transfection of these constructs into Sp2/0 myeloma cells. Binding assays in vitro demonstrated that cBC2 had the same specificity for human milk fat globule membrane (HMFGM) and MUC1+ cells as mBC2, and bound antigen with a similar affinity (cBC2, Ka 5.53 +/- 2.09 x 10(8); mBC2, Ka 1.44 +/- 0.98 x 10(9)). Functionally, only cBC2 (5-25 micrograms ml-1), was able to mediate antibody-dependent cellular cytotoxicity (ADCC) with human effector cells, with 25% maximal specific lysis of MUC1+ cells at an E/T ratio of 100:1. Human complement-mediated lysis was minimal (10-15% specific lysis) with both mBC2 and cBC2. Neither cBC2 nor mBC2 was able to inhibit tumour growth in vivo in the absence of covalently coupled anticancer drugs. However, biodistribution studies demonstrated that both antibodies preferentially targeted MUC1+ tumour cells, with 17% of the injected dose of cBC2, as compared to 27% of mBC2, localized to the MUC1+ tumour at 24 h (less than 6% detected in any other tissue).
Sutton
V R
VR
Austin Research Institute, Austin Hospital, Heidelberg, Victoria, Australia.
Burgess
J
J
Pietersz
G A
GA
Li
W J
WJ
McKenzie
I F
IF
Trapani
J A
JA
D016475:3T3 Cells; D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000235:genetics; D000912:Antibodies, Neoplasm / Q000235:genetics* / Q000276:immunology; D000915:Antibody Affinity; D000920:Antibody-Dependent Cell Cytotoxicity; D000951:Antigens, Neoplasm / Q000276:immunology; D001483:Base Sequence; D001943:Breast Neoplasms / Q000276:immunology*; D003001:Cloning, Molecular; D005803:Genes, Immunoglobulin; D006801:Humans; D007143:Immunoglobulin Heavy Chains / Q000235:genetics; D007145:Immunoglobulin kappa-Chains / Q000235:genetics; D008325:Mammary Neoplasms, Experimental / Q000276:immunology; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D018396:Mucin-1 / Q000276:immunology*; D011993:Recombinant Fusion Proteins; D014018:Tissue Distribution
D000911:Antibodies, Monoclonal; D000912:Antibodies, Neoplasm; D000951:Antigens, Neoplasm; D007143:Immunoglobulin Heavy Chains; D007145:Immunoglobulin kappa-Chains; D018396:Mucin-1; D011993:Recombinant Fusion Proteins