Strand breaks in DNA from normal and transformed human cells treated with 1,3-bis(2-chloroethyl)-1-nitrosourea.

determine the molecular basis for the cytotoxic action that produces the antitumor effect. Unfortunately, there is pres ently no direct way to identify with certainty the critical molecular lesions. It is, however, possible to obtain comrel ative evidence between molecular effects and cytotoxic effects in various cell types, which could lead to a probable identification of the critical lesions. In the current work, we studied DNA single-strand breaks and/or alkali-labile sites in BCNU-treated normal human embryo fibroblasts (Wl-38) and in an SV4O-transformed derivative (VA-13) that exhibited increased sensitivity to BCNU. Nitrosoureas react with biological macromolecules by 2 mechanisms: alkylation, which affects both proteins and nucleic acids; and carbamoylation, which affects proteins but not nucleic acids (3, 22). Since certain nitrosoureas, e.g., chlorozotocin, specifically lack the carbamoylation effect and yet retain antitumor potency (10), it is probably the alkylating activity that is primarily responsible for the antitumor action. Our working hypothesis is that the crucial activity stems from alkylation of DNA and that this activity may be reflected by effects on DNA macromolecular struc ture. Simple alkylnitrosoureas, such as methylor ethylnitro sourea, alkylate DNAat multiple sites (16, 20). Alkylations at guanmne-N-7and adenine-N-3lead to depurination (15), and the resulting apuninic sites are susceptible to phospho diester cleavage by specific repair enzymes, as well as to cleavage by alkali (21). Alkali lability can also result from alkylation of DNA phosphate groups (19). Thus alkylation by simple nitrosoureas produces DNA lesions that can lead to enzymatically produced single-strand breaks and to al kali-labile lesions that can be converted to single-strand breaks by alkali. The sites of DNA alkylation by chloroe thylnitrosoureas, such as BCNU and CCNU, however, have not been fully determined.The production of single-strand breaks or alkali-labile sites by BCNU and CCNU have been reported previously in abstracts (6, 11).