Cytotoxic potential of anti-CD7 immunotoxin (WT1-ricin A) to purge ex vivo malignant T cells in bone marrow.
With the perspective of bone marrow purging in autologous transplantation, we investigated the cytotoxicity of the anti-T cell immunotoxin (IT) WT1-ricin A (anti-CD7) to malignant T cells obtained from patients with T cell acute lymphoblastic leukaemia or lymphoma. The cytotoxic efficacy of IT was based on the extent of protein synthesis inhibition. Cytotoxicity of IT to malignant T cells showed a dependency on antigen density comparable to the T cell lines GH1, CEM, Jurkat, HSB-2 and HPB-ALL and was enhanced considerably in the presence of 6 mM ammonium chloride. The ultimate proof of cell kill can only be obtained from clonogenic assays; however, culturing of malignant T cells was not feasible. Therefore these assays were performed with the cell line CEM that expresses comparable amounts of CD7 antigen as malignant T cells of most patients. More than 6-logs of CEM appeared to be eliminated after incubation with 10(-8) M WT1-ricin A. Immunotoxins are only effective after entering the target cell. The pattern of internalization of the IT was determined by means of 125I-WT1. After internalization the CD7 antigen was re-expressed on the cell membrane. This enables a long incubation period resulting in an increased elimination of malignant T cells. Even after 16 h the IT was still accumulated intracellularly. This pattern of continuous uptake of IT was reflected in a gradually increasing cytotoxicity with incubation time. Effective bone marrow purging can be carried out without adverse effects on progenitor cells with 10(-8) M WT1-ricin A. At that concentration the antibody binding capacity was saturated. We showed that the protein synthesis inhibition in malignant T cells by WT1-ricin A is comparable to the inhibition in T cell lines and that high amounts of CEM cells can be killed. These data suggest that cell lines can be used to test the efficacy of IT to malignant T cells. WT1-ricin A appears to be very potent for the purging of autologous bone marrow from patients with T cell malignancies.
Preijers
F W
FW
Department of Haematology, University Hospital Nijmegen, The Netherlands.
De Witte
T
T
Wessels
J M
JM
Meyerink
J P
JP
Haanen
C
C
Capel
P J
PJ
D000643:Ammonium Chloride / Q000494:pharmacology; D000945:Antigens, Differentiation, T-Lymphocyte / Q000032:analysis; D001853:Bone Marrow / Q000378:metabolism / Q000473:pathology*; D002460:Cell Line; D002469:Cell Separation / Q000379:methods; D006801:Humans; D000922:Immunotoxins / Q000378:metabolism / Q000494:pharmacology*; D015459:Leukemia-Lymphoma, Adult T-Cell / Q000276:immunology; D008228:Lymphoma, Non-Hodgkin / Q000276:immunology; D013601:T-Lymphocytes / Q000276:immunology*; D014407:Tumor Cells, Cultured / Q000187:drug effects; D014410:Tumor Stem Cell Assay
D000945:Antigens, Differentiation, T-Lymphocyte; D000922:Immunotoxins; D000643:Ammonium Chloride