Mafosfamide induces DNA fragmentation and apoptosis in human T-lymphocytes. A possible mechanism of its immunosuppressive action.
Cyclophosphamide, an alkylating agent belonging to the family of nitrogen mustards, is commonly used to treat progressive autoimmune diseases in humans. At the molecular level, its cytotoxicity results from DNA double strand crosslinks and, at higher concentrations, from DNA strand breaks. At the cellular level, cyclophosphamide may selectively affect mature lymphocytes with relative sparing of the respective precursor cells. In this study, we show that 4-hydroxycyclophosphamide (4-OH-CP), the active metabolite of cyclophosphamide, induces apoptosis in mature human lymphocytes at concentrations that are achieved in vivo. Since cyclophosphamide requires enzymatic conversion in the liver to yield its active metabolite, 4-OH-CP was generated in vitro by non-enzymatic hydrolysis of mafosfamide. Apoptotic cell death of lymphocytes was characterized by typical morphological changes, nucleosomal DNA fragmentation, and quantified by 3'-OH end labeling of fragmented DNA. The percentage of apoptotic cells both depended on drug concentration and time of exposure. Cycloheximide or ZnSO4 did not suppress 4-OH-CP induced apoptosis. Etoposide, a topoisomerase II inhibitor known to induce apoptosis in human tumor cell lines like 4-OH-CP, did induce detectable DNA fragmentation in only a minor proportion of T-lymphocytes but suppressed T-cell proliferation.
Pette
M
M
Neurologische Klinik, Julius-Maximilians-Universität, Würzburg, Germany.
Gold
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R
Pette
D F
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Hartung
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HP
Toyka
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KV
D000328:Adult; D000478:Alkylation; D017209:Apoptosis / Q000276:immunology*; D003513:Cycloheximide / Q000494:pharmacology; D003520:Cyclophosphamide / Q000031:analogs & derivatives* / Q000633:toxicity; D004249:DNA Damage / Q000276:immunology*; D004254:DNA Nucleotidyltransferases / Q000032:analysis; D004591:Electrophoresis, Polyacrylamide Gel; D006131:Growth Inhibitors / Q000494:pharmacology*; D006801:Humans; D007166:Immunosuppressive Agents / Q000633:toxicity*; D008213:Lymphocyte Activation / Q000187:drug effects; D008297:Male; D013431:Sulfates / Q000494:pharmacology; D013601:T-Lymphocytes / Q000187:drug effects*; D017967:Zinc Compounds / Q000494:pharmacology; D019287:Zinc Sulfate
D006131:Growth Inhibitors; D007166:Immunosuppressive Agents; D013431:Sulfates; D017967:Zinc Compounds; C048341:mafosfamide; D019287:Zinc Sulfate; D003520:Cyclophosphamide; D003513:Cycloheximide; D004254:DNA Nucleotidyltransferases; C056773:telomere terminal transferase