Bispecific CD3 x CD19 diabody for T cell-mediated lysis of malignant human B cells.
For the treatment of minimal residual disease in patients with leukemias and malignant lymphomas, we constructed a heterodimeric diabody specific for human CD19 on B cells and CD3epsilon chain of the T cell receptor complex. The bispecific diabody was expressed in Escherichia coli using a vector containing a dicistronic operon for co-secretion of V(H)3-V(L)19 and V(H)19-V(L)3 single-chain Fv fragments (scFv). It was purified in one step by immobilized metal affinity chromatography (IMAC) from the periplasmic extract and culture medium. Flow cytometry experiments revealed specific interactions of the diabody with both CD3 and CD19 positive cells, to which it bound with affinities close to those of the parental scFvs. It was less stable than anti-CD3 scFv but more stable than anti-CD19 scFv when incubated in human serum at 37 degrees C. In cytotoxicity tests, the diabody proved to be a potent agent for retargeting peripheral blood lymphocytes to lyse tumor cells expressing the CD19 antigen. The efficiency of cell lysis compared favorably with that obtained with a bispecific antibody (BsAb) of the same dual specificity that was prepared by the quadroma technique.
Recombinant Antibody Research Group, Diagnostics and Experimental Therapy Program, German Cancer Research Center (DKFZ), Heidelberg.
S M
SM
Kipriyanov
G
G
Moldenhauer
G
G
Strauss
M
M
Little
D000595:Amino Acid Sequence; D018033:Antibodies, Bispecific / Q000302:isolation & purification; D000911:Antibodies, Monoclonal / Q000302:isolation & purification; D000918:Antibody Specificity; D015703:Antigens, CD / Q000032:analysis / Q000276:immunology*; D001483:Base Sequence; D017252:CD3 Complex / Q000032:analysis / Q000276:immunology*; D003001:Cloning, Molecular; D003602:Cytotoxicity, Immunologic; D017931:DNA Primers; D000939:Epitopes / Q000737:chemistry / Q000276:immunology; D004926:Escherichia coli; D006801:Humans; D007128:Immunoglobulin Fragments / Q000276:immunology / Q000302:isolation & purification; D007135:Immunoglobulin Variable Region / Q000276:immunology / Q000302:isolation & purification; D016393:Lymphoma, B-Cell / Q000276:immunology*; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000302:isolation & purification; D013601:T-Lymphocytes / Q000276:immunology*
D018033:Antibodies, Bispecific; D000911:Antibodies, Monoclonal; D015703:Antigens, CD; D017252:CD3 Complex; D017931:DNA Primers; D000939:Epitopes; D007128:Immunoglobulin Fragments; D007135:Immunoglobulin Variable Region; D011994:Recombinant Proteins; C055966:immunoglobulin Fv