High toxic efficiency of ricin immunotoxins specific for the T-cell antigen receptor of a human leukemia T-cell line.
Immunotoxins (ITs) were prepared by covalently coupling ricin to monoclonal antibodies (MAbs) directed against: (a) 2 different epitopes of the T-cell receptor (TcR) expressed by the Jurkat leukemia T-cell line (JTi2 and JTi4 MAb), (b) 2 epitopes of the CD3 complex (SpV-T3b and 11D8 MAb), (c) the CD2 and the CD8 cell-surface molecules. Conjugates were assayed for their cytotoxic activity by pre-incubating the Jurkat cell line with different concentrations (10-250 ng/ml) of each IT for 2 hr at 37 degrees C in the presence of 0.1 M lactose. After washing, cells were cultured for 24 hr and their protein synthesis and proliferative capacities were assessed. Dose-response experiments indicated that JTi2, JTi4 and anti-CD3 (11D8) ITs inhibited by greater than 90% the cell line proliferation at 50 ng/ml, a 5-fold lower concentration than that required to achieve a similar effect when anti-CD2 and anti-CD3 (SpVT3b) were used. After 4 hr of culture subsequent to treatment with JTi2 or JTi4 ITs (250 ng/ml), protein synthesis was inhibited (greater than 80%). By limiting dilution analysis (LDA) we estimated that the frequency of proliferating Jurkat cells (1/1.5) was reduced to 1/20, 1/460 and 1/300 after treatment with anti-CD3 (SpVT3b), JTi4 and JTi2 ITs, respectively. Phenotypic analysis of 13 clones derived from JTi2 IT-treated Jurkat cells showed that 50% were CD7+ CD3- JTi- variants. When bone-marrow mononuclear cells, previously mixed with low concentrations of Jurkat cells, were treated with anti-JTi ITs, the toxic efficiency estimated by LDA was maintained whereas the growth of CFU-GM remained unaltered.
Izquierdo
M
M
Department of Immunology, Hospital de la Princesa, Madrid, Spain.
Balboa
M A
MA
Lamana
M L
ML
López-Botet
M
M
D000911:Antibodies, Monoclonal / Q000276:immunology / Q000302:isolation & purification / Q000633:toxicity; D000918:Antibody Specificity; D000945:Antigens, Differentiation, T-Lymphocyte / Q000276:immunology; D000954:Antigens, Surface / Q000276:immunology; D018801:CD2 Antigens; D017252:CD3 Complex; D002460:Cell Line; D004305:Dose-Response Relationship, Drug; D000939:Epitopes / Q000276:immunology; D006801:Humans; D000922:Immunotoxins / Q000276:immunology / Q000302:isolation & purification / Q000633:toxicity*; D015458:Leukemia, T-Cell / Q000276:immunology*; D011948:Receptors, Antigen, T-Cell / Q000187:drug effects* / Q000276:immunology; D011971:Receptors, Immunologic / Q000276:immunology; D012276:Ricin / Q000276:immunology / Q000302:isolation & purification / Q000633:toxicity*; D013601:T-Lymphocytes / Q000276:immunology; D014407:Tumor Cells, Cultured
D000911:Antibodies, Monoclonal; D000945:Antigens, Differentiation, T-Lymphocyte; D000954:Antigens, Surface; D018801:CD2 Antigens; D017252:CD3 Complex; D000939:Epitopes; D000922:Immunotoxins; D011948:Receptors, Antigen, T-Cell; D011971:Receptors, Immunologic; D012276:Ricin