The nucleotide-permeable Escherichia coli cell, a sensitive DNA repair indicator for carcinogens, mutagens, and antitumor agents binding covalently to DNA.

Ether-permeabilized (nucleotide-permeable) Escherichia coli cells respond to alkylating and arylalkylating carcinogens with DNA excision repair, as assessed by their stimulation of DNA repair synthesis. In the present work, we have investigated whether DNA repair synthesis in ether-treated E. coli cells can serve as a general indicator to monitor the DNA-binding of carcinogens, mutagens and antitumor agents. Therefore, a standard assay was developed and comparative analyses were performed on 11 ultimate carcinogens, 10 proximate carcinogens, 2 tumor promoters, 6 mutagens, and 12 antitumor agents. All ultimate carcinogens (alkylating, acylating, arylalkylating agents) and mutagens (e.g., hydrogeen peroxide, acridine derivatives) caused DNA excision repair in wild type cells as measured by [3H] dTMP incorporation and simultaneously inhibited replicative DNA synthesis to various extents. Control experiments with the mutant cells uvrA and uvrB were performed to determine whether the pyrimidine-dimer-specific UV-endonuclease was involved in the removal of DNA damage. This was found to be true for the ultimate carcinogens (Ac)2 ONFln, mitomycin C, and for very reactive alkylating carcinogens. None of the ultimate carcinogens induced repair polymerization in mutant cells lacking the 5'-3' exonucleolytic activity of DNA polymerase I. Proximate carcinogens, such as Me2NNO, 4-nitroquinoline-1-oxide and aflatoxins, did not induce excision repair in the standard assay, probably because of the inability of E. coli to perform the activation steps necessary for covalent DNA-binding. However, Me2NNO, when pretreated with Udenfriend's hydroxylating mixture, gave rise to a low level of repair polymerization in ether-treated cells. Intercalating mutagens, such as quinacrine and ethidum bromide, inhibited replicative DNA synthesis. However, they were not found to be repair-inducers. THE TUMOR PROMOters TPA and phorbol-12,13-didecanoate did not cause excision repair, even when applied at high concentrations, nor did they inhibit repair synthesis stimulated by MeNOUr or (Ac)2 ONFln. The antitumor agents may be classified into two groups on the basis of the influence they exert on DNA synthesis: members of the first group (involving BCNU and bleomycin) stimulate repair polymerization and, in addition, inhibit DNA replication. These compounds are known to bind covalently to DNA. The second group of drugs (including adriamycin and cis-Pt(II)diammine complexes) inhibits DNA replication without stimulating repair synthesis. The predominant DNA-interaction of these compounds is known to be a non-covalent (i.e., intercalative, electrostatic) binding. Our experiments show that the ether-permeabilized E. coli cell can be successfully used to test ultimate carcinogens, mutagens and antitumor agents for repair-inducing and replication-inhibiting activity. The standard test might be extended to pre- and proximate carcinogens, provided these can be suitably activated.