Cytotoxicity, DNA cross-linking, and DNA single-strand breaks induced by cyclophosphamide in a rat leukemia in vivo.

Abstract

A study of cyclophosphamide (CP)-induced DNA damage and repair occurring in vivo was conducted in the brown Norway rat myelocytic leukemia (BNML) model. DNA single-strand breaks (SSB), DNA-DNA interstrand cross-links (DIC), DNA-protein cross-links (DPC), and DNA double-strand breaks (DSB) were measured by alkaline and neutral elution. After i.p. injection of 50 mg/kg CP, DIC were detectable at 1 h and peaked at 8 h. DPC were detectable at 2 h and peaked at 6 h. Both DIC and DPC persisted at a relatively high level until 28 h. Dose-response curves for both DIC and DPC were determined at 4 h after CP injection over the dose range of 25-150 mg/kg. These doses ranged from the minimally effective dose to doses curative for rats bearing this leukemia (1- to 9-log kill of leukemia cells). No SSB or DSB was observed at 4 h after CP injection over the dose range of 15-250 mg/kg, but a low level of SSB was observed at 18-28 h after CP treatment. These data suggest that the cytotoxic effect of CP in vivo is mediated mostly by DIC and DPC. SSB appearing late after CP injection in vivo may be a reflection of repair of DIC and DPC and an indication of the optimal timing for administration of DNA-repair inhibitors. This observation is of interest since our earlier work demonstrated that hydroxyurea can potentiate the therapeutic benefit of CP in this model when it is given over the 4-day period immediately after CP treatment.