The intercalative acridine derivative 4'-(9-acridinylamino)methanesulfon-m-anisidide (m-AMSA), but not its isomer o-AMSA, is a potent antitumor drug that in mammalian cells stimulates the formation of DNA strand breaks that are characterized by tightly bound proteins. Using purified mammalian DNA topoisomerases, we have analyzed the effects of these antitumor drugs on topoisomerase-DNA interactions. The antitumor drug m-AMSA dramatically stimulates the formation of a topoisomerase II-DNA complex that is detected on protein-denaturant treatment: both single-and double-stranded DNA breaks are produced and a topoisomerase II subunit is linked covalently to each 5' end of the broken DNA strands. The noncytotoxic isomer, o-AMSA, which does not induce significant amounts of DNA breaks in cultured cells, exhibits a correspondingly smaller effect in stimulating formation of the complex in vitro. The agreement between in vitro and in vivo studies suggests that mammalian DNA topoisomerase II may be the primary target of m-AMSA and that the drug-induced complex formation between topoisomerase II and DNA may be the cause of cytotoxicity and other effects such as DNA sequence rearrangements and sister-chromatid exchange.A large number of antitumor drugs are known to interact with DNA in vitro (for review, see ref. 1). Despite their success in treating certain tumors, the cellular mechanisms of the actions of these drugs are still unknown (1). Among these antitumor drugs, many of them interact with DNA by an intercalative mode (1). A cellular phenomenon has been noted that seems to be unique to these intercalative antitumor drugs. After treatment of mammalian cells with these antitumor drugs [e.g., daunomycin, adriamycin, ellipticine, actinomycin D, and 4'-(9-acridinylamino)methanesulfon-m-anisidide (m-AMSA)], DNA breakage can be demonstrated by several techniques (2-7). Both single-and double-strand breaks have been observed and their ratio varies depending on the particular drug used. Usually, DNA cleavage is quickly reversed on removal of the drug. The cleavage products have protein tightly (perhaps covalently) associated with the broken DNA ends (2-7). The synthetic antitumor drug m-AMSA, being a weak DNA intercalator (1), is particularly interesting because similar DNA strand breaks can be shown to occur in both cultured mammalian cells and isolated nuclei (6, 8-12). Furthermore, it has been shown that the 5' ends of the cleavage product formed by m-AMSA treatment are blocked by proteins (13). The suggestion that a DNA topoisomerase may be involved has been made (3,5,6,13).Recently, it was shown that a tight complex is formed between purified eukaryotic DNA topoisomerase II and DNA (14,15). Mammalian DNA topoisomerase II and DNA can form a tight complex that, on protein-denaturant treatment, produces protein-linked DNA breaks (15). This complex (henceforth referred to as the "cleavable complex") is operationally defined by the detection method used. One of its unusual features is its reversibility with respect t...
