Development and characterization of a bispecific single-chain antibody directed against T cells and ovarian carcinoma.
Bispecific antibodies with specificity for tumor antigen and CD3 have been shown to redirect the cytotoxicity of T cells against relevant tumor. Our objective was to generate single-chain bispecific antibodies (bsSCA) that could retarget mouse cytotoxic T lymphocytes (CTL) to destroy human ovarian carcinoma in a xenogeneic setting. A bsSCA, 2C11 x B43.13, was constructed by genetic engineering and expressed in mammalian cells. Molecular characteristics, binding properties, and ability to retarget CTL were studied. Western blot analysis showed that the product is a 65-kDa protein. Purification of antibodies could be done by single-step affinity chromatography using protein L-agarose with an unoptimized yield of 200 microg/L. BsSCA 2C11 x B43.13 was capable of binding to mouse CD3 and human CA125 as detected by FACS analysis of EL4 and OVCAR Nu3H2 cells, respectively. It could also bridge activated splenic T cells and human ovarian carcinoma as demonstrated by a bridge FACS assay. Redirected mouse CTL could mediate human target cell lysis in a 20-h 51Cr release assay despite that they are xenogeneic. Prolonged incubation of redirected CTL and tumor targets resulted in a dramatic reduction in tumor cell number. CD28 co-stimulation enhanced redirected CTL function in both types of assays. BsSCA 2C11 x B43.13 thus can be used as a preclinical immunotherapeutic model for human ovarian cancer in a xenogeneic setting.
Kriangkum
J
J
Faculty of Pharmacy and Pharmaceutical Sciences, University of Alberta, Edmonton, Canada.
Xu
B
B
Gervais
C
C
Paquette
D
D
Jacobs
F A
FA
Martin
L
L
Suresh
M R
MR
D000818:Animals; D018033:Antibodies, Bispecific / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification* / Q000633:toxicity; D000911:Antibodies, Monoclonal / Q000096:biosynthesis / Q000235:genetics; D058846:Antibodies, Monoclonal, Murine-Derived; D018394:CA-125 Antigen / Q000276:immunology; D017252:CD3 Complex / Q000276:immunology; D018920:Coculture Techniques; D003602:Cytotoxicity, Immunologic; D005260:Female; D006131:Growth Inhibitors / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification / Q000633:toxicity; D006801:Humans; D051379:Mice; D008807:Mice, Inbred BALB C; D008819:Mice, Nude; D010051:Ovarian Neoplasms / Q000276:immunology* / Q000473:pathology / Q000628:therapy; D013601:T-Lymphocytes / Q000276:immunology*; D014407:Tumor Cells, Cultured
D018033:Antibodies, Bispecific; D000911:Antibodies, Monoclonal; D058846:Antibodies, Monoclonal, Murine-Derived; D018394:CA-125 Antigen; D017252:CD3 Complex; D006131:Growth Inhibitors; C107428:oregovomab