Evaluation of ricin A-chain immunotoxins directed against human T cells.
We have synthesized four immunotoxins (ITs) by covalently coupling the A chain of ricin to murine monoclonal antibodies that recognize surface antigens on human T cells. Treatment of human peripheral blood lymphocytes with either 10.2-A, directed against the CD5 (Tp67) antigen, or 64.1-A, directed against the CD3 (Tp19) antigen, abolished protein synthesis in cells subsequently cultured with phytohemagglutinin (PHA). In contrast, two other ITs (9.6-A and 35.1-A), both directed against the CD2 (Tp50) antigen, had minimal effects on protein synthesis in PHA-stimulated cells. The binding of each IT to T cells was shown by immunofluorescence with fluorescein-conjugated goat anti-mouse immunoglobulin (FITC-GAMIg) and fluorescein-conjugated rabbit anti-ricin A-chain (FITC-RAR) antibodies. Activity of the ricin A chain in each IT was demonstrated by its ability to inhibit protein synthesis in a cell-free reticulocyte lysate assay. Ultrastructural immunoperoxidase analysis of IT internalization showed that ineffective and effective ITs were endocytosed at the same rate (50% of cells had labeled endosomes after 15 min). However, ineffective IT 35.1-A was more rapidly delivered to lysosomes (15-30 min) than effective ITs (10.2-A and 64.1-A) (greater than or equal to 30 min). The data support the hypothesis that there are several distinct pathways for internalization of ITs and that the ability of ricin A chain to reach and inactivate ribosomes may depend upon the specific membrane receptor involved in binding a given IT, its route of internalization, and the rate of entry of the IT into lysosomes.
D000911:Antibodies, Monoclonal / Q000276:immunology; D000945:Antigens, Differentiation, T-Lymphocyte; D000954:Antigens, Surface / Q000276:immunology; D004705:Endocytosis; D006801:Humans; D000922:Immunotoxins / Q000378:metabolism / Q000494:pharmacology*; D008213:Lymphocyte Activation; D008247:Lysosomes / Q000032:analysis; D012276:Ricin / Q000378:metabolism / Q000494:pharmacology*; D013601:T-Lymphocytes / Q000187:drug effects* / Q000276:immunology / Q000378:metabolism
D000911:Antibodies, Monoclonal; D000945:Antigens, Differentiation, T-Lymphocyte; D000954:Antigens, Surface; D000922:Immunotoxins; D012276:Ricin