Cell cycle stage-dependent variations in drug-induced topoisomerase II mediated DNA cleavage and cytotoxicity

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Abstract
The DNA cleavage produced by 4''-(9-acridinylamino)methanesulfon-m-anisidide (m-AMSA) in mammalian cells in putatively mediated by topoisomerase II. We found that in synchronized HeLa cells the frequency of such cleavage was 4.sbd.15-fold greater in mitosis than in S while the DNA of G1 and G2 cells exhibited an intermediate susceptibility to cleavage. The hypersensitivity of mitotic DNA to m-AMSA-induced cleavage. The hypersensitivity of mitotic DNA to m-AMSA-induced cleavage was required relatively abruptly in late G2 and was lost similarly abruptly in early G1. The susceptibility of mitotic cells to m-AMSA-induced DNA cleavage was not clearly paralleled by an increase in topoisomerase II activity (decatenation of kinetoplast DNA) in 350 mM NaCl extracts from mitotic cells compared to similar extracts from cells in G1, S, or G2. Furthermore, equal amounts of decatenating activity from cells in mitosis and S produced equal amounts of m-AMSA-induced cleavage of simian virus 40 (SV40) DNA; i.e., the interaction between m-AMSA and extractable enzyme was similar in mitosis and S. The DNA of mitotic cells was also hypersensitive to cleavage by 4''-demethylepipodophyllotoxin 4-(4,6-O-ethylidene-.beta.-D-glucopyranoside) (etoposide), a drug that produces topoisomerase II mediated DNA cleavage without binding to DNA. Thus, alterations in the drug-chromatin interaction during the cell cycle seem in an unlikely explanation for results in whole cells. Cell cycle stage dependent fluctuations in m-AMSA-induced DNA cleavage may result from fluctuations in the structure of chromatin per se that occur during the cell cycle. Alternatively, they may refelct fluctuations in the structure of chromatin per se that occur during the cell cycle. Alternatively, they may reflect fluctuations in enzyme activity occurring during the cell cycle that were masked by cell cycle stage dependent differences in extractability at 350 mM NaCl either of the enzyme or of substances that influence enzyme activity. Surprisingly, cell cycle stage dependent differences in m-AMSA-induced DNA cleavage did not correlate with differences in the susceptibility to the cytotoxic effects of the drug. In fact, cells in S were most sensitive to these effects. These results are in exception to the previously observed parallel between the susceptibility of mammalian cells to drug-induced DNA cleavage and the susceptibility of the cells to drug-induced cytotoxicity and indicate the complexity of any relation between the two phenomena.

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