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High toxic efficiency of ricin immunotoxins specific for the t‐cell antigen receptor of a human leukemia t‐cell line
Author(s) -
Izquierdo Manuel,
Balboa María A.,
Lamana María L.,
LöpezBotet Miguel
Publication year - 1989
Publication title -
international journal of cancer
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.475
H-Index - 234
eISSN - 1097-0215
pISSN - 0020-7136
DOI - 10.1002/ijc.2910430426
Subject(s) - jurkat cells , immunotoxin , microbiology and biotechnology , cd3 , monoclonal antibody , antigen , biology , t cell , cell culture , epitope , t cell receptor , ricin , leukemia , t lymphocyte , cd8 , cytotoxic t cell , antibody , biochemistry , immunology , immune system , in vitro , toxin , genetics
Abstract 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 (JTi 2 and JTi 4 MAb), (b) 2 epitopes of the CD3 complex (SpV‐T3b and IID8 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°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. Doseresponse experiments indicated that JTi 2 , JTi 4 and anti‐CD3 (IID8) ITs inhibited by >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 JTi 2 or JTi 4 ITs (250 ng/ml), protein synthesis was inhibited (>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), JTi 4 and JTi 2 ITs, respectively. Phenotypic analysis of 13 clones derived from JTi 2 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.