Apoptosis induced by immunotoxins used in the treatment of hematologic malignancies.
The recombinant immunotoxins anti-Tac(Fv)-PE38 (LMB-2), targeting the interleukin-2 receptor alpha subunit (IL-2Ralpha, Tac or CD25), and RFB4(dsFv)-PE38 (BL22), targeting CD22, are being evaluated in clinical trials as treatment for hematologic malignancies. The toxin moiety Pseudomonas exotoxin A (PE) of these recombinant molecules leads to the arrest of protein synthesis due to inactivation of elongation factor 2. Here, we provide evidence that cell lines derived from patients with hematologic malignancies react to immunotoxins not only with inhibition of protein synthesis but also with characteristic hallmarks of apoptosis such as caspase activation, cleavage of the "death substrate poly(ADP)-ribose polymerase and DNA laddering. Anti-Tac(Fv)-PE38 leads to a 10-fold increase in the cleavage of the fluorescent substrate DEVD-AFC, suggesting that a caspase-3-like enzyme is involved. This was verified by cleavage of caspase-3 (CPP32). MT1 cells exhibited DNA laddering after treatment with immunotoxin, which was reversed by pre-treatment with the protease inhibitor zVAD-fmk. This caspase inhibitor led to an at least 5-fold improvement in cell viability without altering inhibition of protein synthesis. Interestingly, HUT-102 cells did not undergo programmed cell death after exposure to immunotoxins that kill these cells. We conclude that immunotoxins may be valuable in the treatment of cancers that are resistant toward apoptosis because their targeted killing is often facilitated by, but not completely dependent on, programmed cell death. Int. J. Cancer 87:86-94, 2000. Published 2000 Wiley-Liss, Inc.
Keppler-Hafkemeyer
A
A
Laboratory of Molecular Biology, Division of Basic Sciences, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892, USA.
Kreitman
R J
RJ
Pastan
I
I
D000590:Amino Acid Chloromethyl Ketones / Q000494:pharmacology; D000906:Antibodies; D015703:Antigens, CD / Q000276:immunology; D000944:Antigens, Differentiation, B-Lymphocyte / Q000276:immunology; D017209:Apoptosis / Q000187:drug effects*; D002051:Burkitt Lymphoma / Q000201:enzymology / Q000628:therapy; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism; D015815:Cell Adhesion Molecules; D002470:Cell Survival / Q000187:drug effects; D003374:Coumarins / Q000494:pharmacology; D053938:DNA Fragmentation / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D019337:Hematologic Neoplasms / Q000628:therapy*; D015151:Immunoblotting; D000922:Immunotoxins / Q000009:adverse effects* / Q000627:therapeutic use*; D037102:Lectins; D007930:Leucine / Q000378:metabolism; D007938:Leukemia / Q000201:enzymology / Q000628:therapy; D016399:Lymphoma, T-Cell / Q000201:enzymology / Q000628:therapy; D009842:Oligopeptides / Q000494:pharmacology; D011480:Protease Inhibitors / Q000494:pharmacology; D011994:Recombinant Proteins / Q000378:metabolism; D051918:Sialic Acid Binding Ig-like Lectin 2; D013778:Tetrazolium Salts / Q000494:pharmacology; D014407:Tumor Cells, Cultured
C110656:2-(4-iodophenyl)-3-(4-nitrophenyl)-5-(2,4-disulfophenyl)-2H-tetrazolium; D000590:Amino Acid Chloromethyl Ketones; D000906:Antibodies; D015703:Antigens, CD; D000944:Antigens, Differentiation, B-Lymphocyte; D015815:Cell Adhesion Molecules; D003374:Coumarins; D000922:Immunotoxins; D037102:Lectins; D009842:Oligopeptides; D011480:Protease Inhibitors; D011994:Recombinant Proteins; D051918:Sialic Acid Binding Ig-like Lectin 2; D013778:Tetrazolium Salts; C407312:antitac(FV)-PE38 recombinant immunotoxin; C115495:aspartyl-glutamyl-valyl-aspartyl-7-amino-4-trifluoromethylcoumarin; C096713:benzyloxycarbonylvalyl-alanyl-aspartyl fluoromethyl ketone; D053148:Caspase 3; D020169:Caspases; D007930:Leucine