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In the context of ex vivo T cell elimination from bone marrow, the anti-T cell cytotoxic potential of immunotoxins (IT) prepared by conjugation of the monoclonal antibodies (MoAb) WT32 (CD3). T101 (CD5), and WT1 (CD7) to ricin A chain was evaluated. The cytotoxicity of IT was based on protein synthesis inhibition in human T cell lines: CH1, CEM, HPB-ALL, and Jurkat, and appeared closely related to the antigen density and internalization rate of the IT. Normal unstimulated T cells appeared to be rather insensitive to IT not due to a low antigen density or decreased internalization. The cytotoxicity of IT to T cells could be enhanced considerably by NH4Cl. Treatment of T cells with a cocktail of IT (10-8 M) and 20 mM NH4Cl resulted in a 5000-fold cytoreduction as measured by clonogenic assays of limiting T cell dilutions, whereas the haematopoietic progenitor cells remained unaltered. Stimulation of T cells with phytohaemagglutinin (PHA) prior to incubation with IT considerably increased the sensitivity to IT treatment. Thus, normal I cells are less sensitive to anti-T cell IT than T cell lines and activated T cells. This suggests that a low protein synthesis is responsible for the resistance to IT. However, a high specific cytotoxicity of 11 to normal 1 cells can be achieved in the presence of 20 mM ammonium chloride. [ABSTRACT FROM AUTHOR]

WT 1, an IgG2a subclass monoclonal antibody, recognizes a human T lineage specific antigen (mol. wt 40,000). This antigen is strongly expressed on thymic T blasts, and on peripheral T cells activated by phytohaemagglutinin, whereas cortical thymocytes and peripheral T cells are moderately positive for WT 1, In contrast, B lymphocytes, myeloid and erythroid cells, including the progenitor cells of these lineages, and terminal deoxynucleotidyl transferase positive cells in the bone marrow, arc all WT 1 negative. Binding of WT 1 to T cells is blocked by a previously described antibody (3A1) suggesting that both antibodies bind to the same antigen present on human T cells. WT 1, however, is also reactive with T lymphocytes from rhesus monkeys whereas 3A1 is not. Therefore, the biological effects of WT 1 could be studied in a monkey model. In a skin allograft model. WT 1 was immunosuppressive and induced a marked prolongation of graft survival. [ABSTRACT FROM AUTHOR]