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Abasic sites in HeLa cell DNA were increased in frequency by exposing the cells to lucanthone. Cell growth in the presence of lucanthone caused progressive accumulation of abasic sites and loss of cellular DNA. After 2 hr in 8 microM lucanthone, the abundance of abasic sites was 2.4 fold greater than the background of 9.9 +/- 2.0 SE abasic sites/10(6) nucleotides; 80 microM lucanthone in the growth medium increased the level 12.6 +/- 2.5 SE fold and decreased the DNA content in HeLa cells to one-half of the value obtained in untreated cells. The frequency of abasic sites in cellular DNA was determined by the aldehyde reactive probe method, with reference to abasic sites created in plasmid pBR322. The ability of lucanthone to inhibit the normal repair of abasic sites might reflect inhibition of apurinic/apyrimidinic endonuclease (HAP1) by the drug, thereby preventing an early step in the base excision repair pathway. Unrepaired abasic sites prevalent after ionizing radiation are cytotoxic lesions that promote DNA strand breaks. These results suggest a rationale for the joint lethal effects of lucanthone and ionizing radiation in cells and the accelerated tumor regression observed in cancer patients who received the combined therapy.

Hycanthone, the most potent mutagen in a series of nine thiaxanthenones, is a potent inducer of nuclear immunoreactivity to antinucleoside antibodies in HeLa cells. This response indicates exposure of single-stranded DNA regions. All classes of mutagens thus far tested share this property with hycanthone. Immunoreactivity to antinucleoside antibodies was also induced by brief exposure to hycanthone, 3 microgram/ml, in human fibroblasts from three normal subjects and in fibroblasts from seven patients with DNA repair deficiencies. Unlike those of many other mutagens, the metabolic effects and immunoreactivity induction of hycanthone were readily reversible. No evidence for covalent attachment of [3H]hycanthone to HeLa macromolecules could be found. Induction of DNA repair synthesis could not be detected by autoradiography after exposure of cells to hycanthone. Exposure of single-stranded DNA regions appears to be an important feature of the mechanism of action of hycanthone as a mutagen. Both hycanthone and lucanthone intercalate with DNA, but hycanthone was much less active than was lucanthone in reducing the rapid sedimentation of cell lysate DNA in alkaline sucrose gradients. Similarities and differences, therefore, have been found in the way the potent and the weak mutagen affect DNA of HeLa cells. This may provide clues to understanding the mechanism of mutagenesis by thiaxanthenones and other mutagens.

The DNA and histone content of HeLa S-3 cell cultures was measured by direct mass assays 21 hours after 1000 rad of X-irradiation, when the cells were arrested in G2 phase. The nuclear DNA content of such cultures was found to be deficient (73% of control values). In contrast, the synthesis of nuclear histones persisted, and the total histone content was close to 100% of control values. When synchronously-growing cultures were irradiated in mid-S phase and examined 3-5 hours later in G2 phase, both DNA and histone content were equal to control values.