John L. Seed

A report of the U.S. environmental protection agency gene-tox program: Part I Host-mediated assay☆

Abstract The methodologies and status of the Host-Mediated Assay were reviewed using the published literature available up to June 1980. The Working Group reviewed 274 documents, including abstracts, research articles, review articles, and publicly available contracts and grant final reports. From this group, abstracts and reviews were rejected from critical evaluation. 77 documents were accepted and reviewed by the Working Group and the test results summarized. These selected documents yielded 208 chemicals that were evaluated in the host-mediated assay. Of these chemicals, 133 were mutagenic in this assay with one or more indicators. 76 chemicals, several of which are not considered to be carcinogenic, were not detected by any of the indicators. Of the 208 chemicals, 125 had been tested in carcinogenicity assay in rodents. 90, or 71%, of the carcinogens were detected as mutagens in the Host-Mediated Assay. In several cases, those carcinogens not detected may have been negative because of improper selection of the indicator. The Working Group concluded that the Host-Mediated Assay is an important test in mutagenicity/carcinogenicity research and that, by proper selection of protocols and indicators, valuable information can be gained that otherwise would be overlooked by strict, in vitro assays.The methodologies and status of the Host-Mediated Assay were reviewed using the published literature available up to June 1980. The Working Group reviewed 274 documents, including abstracts, research articles, review articles, and publicly available contracts and grant final reports. From this group, abstracts and reviews were rejected from critical evaluation. 77 documents were accepted and reviewed by the Working Group and the test results summarized. These selected documents yielded 208 chemicals that were evaluated in th host-mediated assay. Of these chemicals, 133 were mutagenic in this assay with one or more indicators. 76 chemicals, several of which are not considered to be carcinogenic, were not detected by any of the indicators. Of the 208 chemicals, 125 had been tested in carcinogenicity assay in rodents. 90, or 71%, of the carcinogens were detected as mutagens in the Host-Mediated Assay. In several cases, those carcinogens not detected may have been negative because of improper selection of the indicator. The Working Group concluded that the Host-Mediated Assay is an important test in mutagenicity/carcinogenicity research and that, by proper selection of protocols and indicators, valuable information can be gained that otherwise would be overlooked strict, in vitro assays.

SINGLET OXYGEN INDUCED MUTAGENESIS OF BENZO[α]PYRENE DERIVATIVES

Abstract— Singlet oxygen activates the mutagenicity of several benzo[α]pyrene (BP) derivatives in the absence of mammalian metabolic action. This has been demonstrated using a separated‐surface‐sensitizer system for generating chemically pure singlet oxygen, eliminating most of the complications that arise with singlet oxygen generation by conventional photosensitization. Salmonella typhimurium bacteria were exposed to singlet oxygen in the presence of certain BP derivatives and the mutation frequency determined with an azaguanine forward mutation assay. The mutation frequency was increased by exposure to singlet oxygen compared to light‐only controls for those BP derivatives that were saturated at either the 7,8 or 9,10 positions but not both. The increase in mutuation frequency depends on both the concentration of BP derivative and on the dose of singlet oxygen. Mutation frequency was also significantly increased when bacteria were treated with a solution of trans‐7,8‐dihydrodiol‐BP that had been separately exposed to singlet oxygen, unequivocally demonstrating that the mutagenicity is due to the formation of a product of BP derivative oxidation by singlet oxygen and that this product has a lifetime at least on the order of minutes in acetonitrile. The requirement for singlet oxygen rather than some other form of reactive oxygen was confirmed by determination of the gas phase lifetime of the intermediate responsible for activating mutagenicity. This was performed by measuring the dependence of the mutation frequency on the distance separating the sensitizer from the target. This gives a value of 88 ± 35 ms, which is in excellent agreement with the mean value of 89 ms calculated from previous independent determinations of the gas phase lifetime of singlet oxygen reported in the literature.

Activation of Xenobiotics by Human Polymorphonuclear Leukocytes via Reactive Oxygen-Dependent Reactions

Reactive oxygen intermediates have become widely implicated in various pathologic states, chemical-induced tissue injury and chemical carcinogenesis (1–5). While much of the interest in the role of oxy-radicals in these processes has centered on the direct interaction of reactive oxygen metabolites with biomolecules, it is becoming increasingly apparent that molecular oxygen-derived oxidants can also participate in the metabolic activation of chemicals (6,7). It has been hypothesized that polymorphonuclear leukocytes (PMNs) may be a useful cellular model to study the interaction and possible activation of compounds by reactive oxygen species (Figure 1)(8,9). Resting PMNs release measurable quantities of superoxide \({{O}_{2}}\overline{.}\) ), hydrogen peroxide (H2O2) and hydroxyl radical (•OH), while activation of their redox metabolism by both particulate and soluble stimulants results in an increased rate in the generation of these molecular oxygen-derived oxidants (10,11). The utilization of H2O2 by the PMN enzyme myeloperoxidase (MPO) results in the formation of hypochlorous acid and an O2 metabolite or complex with singlet oxygen (1O2)-like reactivity (11). The data presented in this study demonstrate that bleomycin A2 and benzo[a]pyrene-7,8-dihydrodiol (BP-7,8-dihydrodiol) are activated to genotoxic derivatives as a result of their interaction with PMN-derived oxidants. Such an activation mechanism could provide an explanation as to how neoplasms often develop at sites of ongoing inflammation (12).

Effects of anethole dithiolthione and 2(3)‐tert‐butyl‐4‐hydroxyanisole on schistosome granuloma formation

Summary Administration of the antioxidants 2(3)‐tert‐butyl‐4‐hydroxyanisole (BHA) or 5‐(P‐methoxyphenyl)‐3H‐1.2‐dithiol‐3‐thione (ADT) to female CD‐1 mice starting 4 weeks after infection with 70 cercariae of Schistosoma mansoni resulted in a decrease in the size of the inner fibrotic region of the hepatic granuloma. The cellular composition of the granuloma was not altered by treatment with these two compounds. The administration of the specific superoxide scavenger copper diisopropylsalicylate (CuDIPS) resulted in a similar decrease in granuloma size, suggesting a role of superoxide radicals in the granulomatous response.

A report of the U.S. environmental protection agency gene-tox program: Part II Body fluid analysis

Abstract The methodologies and status of Body Fluid Analysis were reviewed using the published literature available up to June 1980. 98 documents were reviewed by the Working Group. These included abstracts, research articles, and review articles. From this selection, abstracts and reviews were rejected from critical evaluation by the Working Group, who accepted and reviewed 38 documents and summarized the results. The conclusion reached by the working Group was that, although the Body Fluid Analysis could not be fully assessed because of the limited number of compounds tested, tests of body fluids may identify mutagens that are not detected by in vitro methods.