Mutagenicity of diol-epoxides and tetrahydroepoxides of benz(a)acridine and benz(c)acridine in bacteria and in mammalian cells.

The mutagenic activities of benz(a)acridine, benz(c)acridine, and a number of their derivatives, including 12 epoxides and diol-epoxides, were examined in bacterial and mammalian cells to determine the importance of bay-region activation of azapolycyclic aromatic hydrocarbons. In Salmonella typhimurium strain TA98, the diastereomeric bay-region 3,4-diol-1,2-epoxides of benz(c)acridine, in which the benzylic hydroxyl group is either cis (diol-epoxide 1) or trans (diol-epoxide 2) to the epoxide oxygen, had equivalent mutagenic potency (250 to 300 His+ revertants/nmol), while in S. typhimurium strain TA100, diolepoxide 1 induced 5100 His* revertants/nmol and was approxi mately twice as active as was diol-epoxide 2. In Chinese hamster V79 cells, the diol-epoxide 2 isomer of benz(c)acridine 3,4-diol1,2-epoxide was approximately twice as mutagenic (4.5 8-azaguanine-resistant colonies/105 surviving cells/nmol) as diol-epox ide 1. In all three test systems, the bay-region diol-epoxides of benz(c)acridine were from 1 to 4 orders of magnitude more mutagenic than were the non-bay-region diol-epoxides in which the epoxide was in the 3,4-, 8,9-, or 10,11-positions of the molecule. With the benz(a)acridine diol-epoxides, the 3,4-diol1,2-epoxide 2 diastereomer was 2 to 4 times more active than was the 3,4-diol-1,2-epoxide 1 diastereomer in the bacterial systems, but both diastereomers had less than 10% of the activity of their benz(c)acridine counterparts. The bay-region tetrahydro-1,2-epoxides of both benzacridines were exception ally mutagenic, and the benz(c)acridine epoxide was 4to 11fold more active than was the benz(a)acridine epoxide. The bayregion diol-epoxide diastereomers of benz(c)acridine and its bayregion tetrahydro-epoxide were not metabolized to nonmutagenic products by highly purified epoxide hydrolase (EC 3.3.2.3). Metabolic activation experiments using the cytochrome P-450dependent monooxygenase system and the dihydrodiol precur sors of the bay-region diol-epoxides were consistent with the intrinsic mutagenicity data, in that benz(c)acridine 3,4-dihydrodiol was metabolized to mutagenic products to a greater extent than was benz(a)acridine 3,4-dihydrodiol. The benz(c)acridine 1,2-, 5,6-, 8,9-, and 10,11-dihydrodiols were not metabolically acti vated to mutagenic metabolites. These results provide initial evidence that the bay-region theory can be extended to certain azapolycyclic aromatic hydrocarbons and indicate that the posi tion of the nitrogen heteroatom can markedly affect mutagenic activity. INTRODUCTION Polycyclic aromatic hydrocarbons are a large, widespread, and well-characterized class of environmental carcinogens formed during the incomplete combustion of organic matter (7, 10). Under appropriate combustion conditions, nitrogen may be in corporated into the aromatic ring systems to form nitrogen heterocycles. Significant concentrations of these compounds have been identified in coal-derived fuels (27), urban air particulates (25, 29), and aquatic sediments (2, 32), and several of the azaaromatics are known carcinogens (10,15). Numerous investigations over the past 8 years have indicated that bay-region diol-epoxides (Chart 1) play a predominant role in mediating the mutagenic and carcinogenic activity of at least 12 unsubstituted and alkyl-substituted polycyclic aromatic hydro carbons (8, 14, 21, 39). These diol-epoxides are formed in mammalian cells via oxidation of the angular benzo-ring of the hydrocarbon to form an arene oxide with a bay-region double bond, hydration of the arene oxide to a frans-dihydrodiol,3 and finally oxidation of the olefinic double bond to form the diolepoxide. Cytochrome P-450-dependent monooxygenases cata lyze the 2 oxidative reactions, and another microsomal enzyme, epoxide hydrolase, catalyzes dihydrodiol formation (19). Despite the voluminous literature on the metabolic fate and biological activity of polycyclic aromatic hydrocarbons and their metabolites, few, if any, comparable studies have been reported for the nitrogen heterocycles. We have therefore undertaken an evaluation of the mutagenic potential of 2 nitrogen heterocycles, B(a)ACR and B(c)ACR, as well as a number of their epoxide derivatives (Chart 1). The choice of these 2 heterocycles was predicated on: (a) their presence in our environment; (b) the reported carcinogenicity of several of their methyl derivatives; (c) the expectation that several diol-epoxides and related derivatives of both heterocycles could be unequivocally synthesized; and (d) the availability of a large number of comparable derivatives of BA, the analogous polycyclic aromatic hydrocarbon. Thus, we ' Supported in part by Grant CA 22985 from the National Cancer Institute. 2To whom requests for reprints should be addressed. Received August 16.1982; accepted January 5,1983. 3The abbreviations used are: dihydrodiol, rrans-dihydroxydihydro derivatives of benz(a)acridine, benz(c)acridine, or benz(a)anthracene, in which the hydroxyl groups are either at the 1,2-, 3,4-, 5,6-, 8,9-, or 10,11-positions; B<a)ACR, benz(a)acridine; B(c)ACR, benz(c)acridine; BA, benz(a)anthracene; H4-1,2-epoxide, 1,2-epoxy-1,2,3,4-tetrahydroderivative of benz(a)acridine, benz(c)acridine, or benz(a)anthracene; H4-3,4-epoxide, 3,4-epoxy-1,2,3,4-tetrahydro deriv; live of benz(c)acridine or benz(a)anthracene; DMSO, dimethyl sulfoxide; 3,4-diol-1,2-epox ide 1, (±)-3«,4fi-dihydroxy-10,2/}-epoxy-1,2,3,4-tetrahydrobenz(a)acridine. benz(c)acridine, or benz(a)anthracene diol-epoxide diastereomer, in which the benzylic hydroxyl and the epoxide oxygen are cis; 3,4-diol-1,2-epoxide 2, (±)-3«,4iJdihydroxy-1a,2tt-epoxy-1,2,3,4-tetrahydrobenz(a)acridine, benz(c)acridine, or benz(a)anthracene diol-epoxide diastereomer, in which these groups are trans; other diol-epoxides are similarly designated; «and tÃ-, relative stereochemistry, and all compounds are racemic mixtures where enantiomers are possible. 1656 CANCER RESEARCH VOL. 43 on May 28, 2017. © 1983 American Association for Cancer Research. cancerres.aacrjournals.org Downloaded from Mutagenicity of Benzacridine Epoxides