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1.
Environ Health Perspect ; 108 Suppl 2: 215-23, 2000 May.
Article in English | MEDLINE | ID: mdl-10807553

ABSTRACT

This article addresses the evidence that trichloroethylene (TCE) or its metabolites might mediate tumor formation via a mutagenic mode of action. We review and draw conclusions from the published mutagenicity and genotoxicity information for TCE and its metabolites, chloral hydrate (CH), dichloroacetic acid (DCA), trichloroacetic acid (TCA), trichloroethanol, S-(1, 2-dichlorovinyl)-l-cysteine (DCVC), and S-(1, 2-dichlorovinyl) glutathione (DCVG). The new U.S. Environmental Protection Agency proposed Cancer Risk Assessment Guidelines provide for an assessment of the key events involved in the development of specific tumors. Consistent with this thinking, we provide a new and general strategy for interpreting genotoxicity data that goes beyond a simple determination that the chemical is or is not genotoxic. For TCE, we conclude that the weight of the evidence argues that chemically induced mutation is unlikely to be a key event in the induction of human tumors that might be caused by TCE itself (as the parent compound) and its metabolites, CH, DCA, and TCA. This conclusion derives primarily from the fact that these chemicals require very high doses to be genotoxic. There is not enough information to draw any conclusions for trichloroethanol and the two trichloroethylene conjugates, DCVC and DCVG. There is some evidence that DCVC is a more potent mutagen than CH, DCA, or TCA. Unfortunately, definitive conclusions as to whether TCE will induce tumors in humans via a mutagenic mode of action cannot be drawn from the available information. More research, including the development and use of new techniques, is required before it is possible to make a definitive assessment as to whether chemically induced mutation is a key event in any human tumors resulting from exposure to TCE.


Subject(s)
Carcinogens, Environmental/adverse effects , Mutagens/adverse effects , Neoplasms/chemically induced , Trichloroethylene/adverse effects , Animals , Carcinogens, Environmental/metabolism , Dose-Response Relationship, Drug , Genes/drug effects , Humans , Mutagens/metabolism , Neoplasms/genetics , Risk Assessment , Trichloroethylene/metabolism , United States , United States Environmental Protection Agency
2.
Environ Mol Mutagen ; 35(3): 185-90, 2000.
Article in English | MEDLINE | ID: mdl-10737953

ABSTRACT

The Mouse Lymphoma Assay (MLA) Workgroup addressed and reached consensus on a number of issues. Discussion focused on five areas: (1) acceptable assay versions; (2) cytotoxicity measure; (3) 24-hr treatment; (4) microwell colony counting and sizing; and (5) data acceptability/statistical analysis. Although the International Conference on Harmonisation (ICH) indicated a preference for the microwell over the soft agar method, all of the workgroup members agreed that both versions of the MLA are equally acceptable. The workgroup agreed that it is desirable for both assay versions to use the same measure of cytotoxicity to define the acceptable and required concentration range. Currently, laboratories using the microwell version use the relative survival (RS) determined by cloning immediately after the treatment. Laboratories using the soft agar method do not obtain an RS but use the relative total growth (RTG), a combination of the relative suspension growth (RSG) during the expression period and the relative cloning efficiency determined at the time of mutant selection. The workgroup agreed to investigate the RSG, the RS, and the RTG and to develop further guidance. In the interim, the workgroup reached consensus that the RTG be used as the standard measure of cytotoxicity. The ICH recommended a 24-hr treatment in the absence of S9 when negative results are obtained with short (3-4 hr) treatments. The workgroup agreed to retain this requirement but acknowledged that more data are needed prior to making final recommendations concerning the need for and the specific protocol for the 24-hr treatment. Environ. Mol. Mutagen. 35:185-190, 2000 Published 2000 Wiley-Liss, Inc.


Subject(s)
Lymphoma/genetics , Thymidine Kinase/genetics , Animals , Guidelines as Topic , Lymphoma/enzymology , Mice , Mutagenicity Tests , Tumor Cells, Cultured
6.
Mutat Res ; 431(2): 247-57, 1999 Dec 17.
Article in English | MEDLINE | ID: mdl-10635991

ABSTRACT

A pilot biomarker study was conducted to investigate the feasibility of using the hypoxanthine guanine phosphoribosyltransferase (HPRT) gene in peripheral blood lymphocytes as a biomarker for detecting genetic effects of arsenic exposure. Blood and urine samples were obtained from workers highly exposed to arsenic in a copper roasting plant in Antofagasta, Chile. Individuals were classified according to their job titles into three potential exposure groups: high, medium, and low. To confirm exposure, arsenic concentration was determined in urine samples. The HPRT mutant frequencies were measured in lymphocytes from 15 individuals ranging in age from 24 to 66 years. The mean mutant frequencies for the three exposure groups were: low (9 x 10(-6)), medium (11 x 10(-6)), and high (24 x 10(-6)). An increased mutant frequency was observed in the highly exposed group, but the response was so slight that it is not likely that this assay will be capable of providing dose-response information across a range of lower, more typical environmental arsenic levels.


Subject(s)
Arsenic/toxicity , Hypoxanthine Phosphoribosyltransferase/drug effects , Hypoxanthine Phosphoribosyltransferase/genetics , Mutation , Occupational Exposure , Aged , Biomarkers , Chile , Chromosome Aberrations , DNA Mutational Analysis , Dose-Response Relationship, Drug , Genetics, Population , Humans , Lymphocytes/drug effects , Lymphocytes/physiology , Male , Metallurgy , Middle Aged , Pilot Projects
7.
Mutagenesis ; 13(5): 427-34, 1998 Sep.
Article in English | MEDLINE | ID: mdl-9800187

ABSTRACT

The thymidine kinase locus (Tk1) in Tk(+/-)-3.7.2C mouse lymphoma cells is widely used to identify mutagenic agents. Because Trp53 (the mouse homolog of human TP53) is located with Tk1 on chromosome 11 and is critical in regulating cellular responses following exposure to DNA damaging agents, we wanted to determine if these mouse lymphoma cells harbor mutations in Trp53. Single-stranded conformation polymorphism (SSCP) analysis of PCR-amplified exons 4-9 of Trp53 indicated mutations in both exons 4 and 5. We sequenced exons 4-9 from isolated clones of Tk(+/-)-3.7.2C cells and a Tk-/- mutant (G4). Mutant G4 has two copies of the chromosome carrying the Tk1- allele and no copy of the chromosome carrying the Tk1+ allele and thus could establish linkage of the individual Trp53 and Tk1 alleles. DNA sequence analysis revealed no mutations in exons 6-9 in any Tk(+/-)-3.7.2C or G4 clones. As suggested by SSCP, there was a nonsense mutation in exon 4 at bp 301 (codon 101) in one Trp53 allele. Tk(+/-)-3.7.2C clones have both mutant and wild-type sequences at bp 301; G4 clones have wild-type exon 4 sequence. These data allow assignment of the Trp53 exon 4 mutated allele to chromosome 11 carrying the Tk1+ allele. The exon 4 mutation leads to a stop codon early in translation, thus functionally deleting the Trp53 allele on the Tk1(+)-bearing chromosome. As previously reported, we find a missense mutation in exon 5 at bp 517 (codon 173) in one Trp53 allele. Using the G4 clones we determined that the exon 5 mutation is linked to the Tk1- allele. Thus the Tk +/-(-)3.7.2C mouse lymphoma cells have two mutant Trp53 alleles, likely accounting for their rapid cell growth and contributing to their ability to detect the major types of mutational damage associated with the etiology of tumor development. This ability to integrate across the mutational events seen in the multiple stages of tumor development further supports the use of the assay in chemical and drug safety studies and its recommendation as part of the required screening battery for regulatory agency submissions.


Subject(s)
Genes, p53 , Leukemia L5178/genetics , Mutagenicity Tests/standards , Mutation, Missense , Neoplasm Proteins/genetics , Point Mutation , Thymidine Kinase/genetics , Alleles , Amino Acid Sequence , Animals , Base Sequence , Chromosome Mapping , Codon/genetics , Codon, Nonsense/genetics , DNA, Neoplasm/genetics , Exons/genetics , Genetic Linkage , Mice , Molecular Sequence Data , Polymerase Chain Reaction , Polymorphism, Single-Stranded Conformational
8.
Mutat Res ; 413(3): 265-76, 1998 Mar 30.
Article in English | MEDLINE | ID: mdl-9651541

ABSTRACT

The disinfection of water, required to make it safe for human consumption, leads to the presence of halogenated organic compounds. Three of these carcinogenic 'disinfection by-products', dichloroacetic acid (DCA), trichloroacetic acid (TCA) and chloral hydrate (CH) have been widely evaluated for their potential toxicity. The mechanism(s) by which they exert their activity and the steps in the etiology of the cancers that they induce are important pieces of information that are required to develop valid biologically-based quantitative models for risk assessment. Determining whether these chemicals induce tumors by genotoxic or nongenotoxic mechanisms (or a combination of both) is key to this evaluation. We evaluated these three chemicals for their potential to induce micronuclei and aberrations as well as mutations in L5178Y/TK +/- (-)3.7.2C mouse lymphoma cells. TCA was mutagenic (only with S9 activation) and is one of the least potent mutagens that we have evaluated. Likewise, CH was a very weak mutagen. DCA was weakly mutagenic, with a potency (no. of induced mutants/microgram of chemical) similar to (but less than) ethylmethanesulfonate (EMS), a classic mutagen. When our information is combined with that from other studies, it seems reasonable to postulate that mutational events are involved in the etiology of the observed mouse liver tumors induced by DCA at drinking water doses of 0.5 to 3.5 g/l, and perhaps chloral hydrate at a drinking water dose of 1 g/l. The weight-of-evidence for TCA suggest that it is less likely to be a mutagenic carcinogen. However, given the fact that DCA is a weak mutagen in the present and all of the published studies, it seems unlikely that it would be mutagenic (or possibly carcinogenic) at the levels seen in finished drinking water.


Subject(s)
Chloral Hydrate/toxicity , Dichloroacetic Acid/toxicity , Disinfectants/toxicity , Leukemia L5178/genetics , Mutagens/toxicity , Trichloroacetic Acid/toxicity , Water Purification , Animals , Chromosome Aberrations , Mice , Mutagenicity Tests , Thymidine Kinase/genetics , Tumor Cells, Cultured/drug effects
9.
Mutat Res ; 397(2): 119-36, 1998 Feb 02.
Article in English | MEDLINE | ID: mdl-9541637

ABSTRACT

A survey of glycophorin A (gpa) in vivo somatic cell mutation in a population of 394 healthy people from 8 to 77 years of age (mean age +/- SD 41 +/- 15 years) revealed a subset of 37 individuals with stably elevated allele-loss and/or allele-loss with duplication variant erythrocyte frequencies (Vf) exceeding 30 x 10(-6). These 37 individuals with gpa outlier Vf are significantly older (P < 0.001) than the remainder of the larger study population from which they were drawn reflecting a highly significant increase in the prevalence of these individuals in the population beyond age 40 years. A study of hpt mutant frequencies (Mf) in the peripheral blood T-lymphocytes of 27 of these individuals, together with 15 matched control individuals with unremarkable gpa Vf, was undertaken to determine if these subjects also displayed elevated mutation frequencies at this independent locus indicative of globally elevated somatic mutation. The hprt Mf in these 27 subjects (geometric mean 11.5 x 10(-6)(dispersion interval 5.8 x 10(-6) to 22.8 x 10(-6)) was not significantly different from that observed in the 15 controls (geometric mean 12.1 x 10(-6)(dispersion interval 5.7 x 10(-6) to 25.5 x 10(-6)). These Mf are higher than typically reported values reflecting the older age distribution of these individuals (arithmetic mean age +/- SD 53 +/- 12 and 50 +/- 16 years for the subjects and controls, respectively). Taken together, these data suggest that several genetic mechanisms may be responsible for producing the gpa outlier Vf observed in these subjects. The observation that hprt Mf were not increased indicates that the majority did not arise by a genome-wide increased rate of somatic mutation detectable at both loci. The fixation and subsequent expansion of 'jackpot' mutations at the gpa locus occurring early in embryonic/fetal development also does not appear to be a predominant mechanism. Some cases may result from a stable over-representation of gpa variant cells, perhaps associated with a marked age-dependent decrease in the number of contributing erythroid stem cells in the bone marrow. The subset that displays elevated allele-loss with duplication Vf involving both gpa alleles may represent individuals with increased rates of somatic recombination. Elevations arising by this mechanism are not detected in the hprt assay, but could be confirmed using a autosomal locus in vivo somatic cell mutation endpoint such as the hla-a assay. Of primary biological significance, these results demonstrate that genetics/stochastic processes leading to the loss of heterozygosity of somatic cells occur ubiquitously in humans and in some individuals this level of somatic mosaicism can approach a frequency of 10(-3) at the gpa locus in erythroid lineage cells.


Subject(s)
Erythrocytes/metabolism , Glycophorins/genetics , Hypoxanthine Phosphoribosyltransferase/genetics , Mutation , T-Lymphocytes/enzymology , Adolescent , Adult , Aged , Child , Female , Humans , Male , Middle Aged
10.
Mutat Res ; 386(3): 279-90, 1997 Jun.
Article in English | MEDLINE | ID: mdl-9219565

ABSTRACT

Arsenic is one of the few identified human carcinogens that has yet to be shown to cause cancer in rodents when the standard bioassay protocols are used. The reasons for this apparent interspecies difference are unclear but may be related to differences between humans and rodents in their detoxification capabilities. Detoxification of arsenic may occur through a methylation pathway. If, in fact, methylation does detoxify arsenic, one would predict that the methylated arsenicals might be less genotoxic than the inorganic arsenicals. To evaluate the hypothesis that the inorganic arsenicals are more mutagenic than the organic arsenicals, we tested sodium arsenite, sodium arsenate, monomethylarsonic acid (MMA) and dimethylarsinic acid (DMA) for their relative mutagenic and clastogenic potentials. We used the L5178Y/TK+/- mouse lymphoma assay which allows the detection of chemicals inducing a broad spectrum of different types of genetic damage. Sodium arsenite and sodium arsenate were active at concentrations of 1-2 micrograms/ml and 10-14 micrograms/ml, respectively. MMA was active between 2500-5000 micrograms/ml; while DMA required almost 10000 micrograms/ml to induce a genotoxic response. The organic arsenicals are thus much less potent as mutagenic agents than the inorganic arsenicals. All four of these arsenicals appear to act by mechanisms that cause chromosomal mutations.


Subject(s)
Arsenic/toxicity , Chromosome Aberrations , Lymphoma/genetics , Poisons/toxicity , Animals , Arsenic/chemistry , Humans , Methylation , Mice , Tumor Cells, Cultured
11.
Mutat Res ; 348(3): 105-10, 1995 Nov.
Article in English | MEDLINE | ID: mdl-8524361

ABSTRACT

3-chloro-4-(dichloromethyl)-5-hydroxy-2(5H)-furanone (MX) was tested without exogenous activation in L5178Y/TK+/(-)-3.7.2C mouse lymphoma cells for mutation at the thymidine kinase locus and for clastogenicity. At a concentration of 0.75 micrograms/ml, the induced mutant frequency was 1027 per 10(6) survivors (survival = 11%). A concentration-related increase of large and small colony mutants was observed, but the majority of the MX induced mutants formed small colonies, consistent with the positive clastogenic response that was observed. MX primarily induced chromatid breaks and rearrangements (30 chromatid and 4 chromosome aberrations per 100 cells) at the 0.75 microgram/ml dose. These studies indicate that MX induces a broad spectrum of genetic damage.


Subject(s)
Furans/toxicity , Mutagens/toxicity , Water Pollutants/toxicity , Animals , CHO Cells , Clone Cells , Cricetinae , Lymphoma , Mice , Thymidine Kinase/genetics , Thymidine Kinase/metabolism , Tumor Cells, Cultured
12.
Carcinogenesis ; 15(7): 1463-6, 1994 Jul.
Article in English | MEDLINE | ID: mdl-8033326

ABSTRACT

The DNA sequence of 11 in vivo-arising intragenic deletion junctions occurring in the hypoxanthine-guanine phosphoribosyltransferase (hprt) gene of human T-lymphocytes was determined. These deletions ranged in size from 16 bp to 4057 bp. Extensive homology was not found at the deletion breaksites, indicating that non-homologous recombination was responsible for these deletions. Short regions of homology (1-3 nucleotides) at the deletion termini, which may direct the recombination event, were found in seven of the mutations. Only one mutation had an unaccounted for nucleotide at the junction. V(D)J recombinase recognition sequences, previously identified at other hprt deletion breaksites, were not present. Such features are also found at the deletion and translocation junctions of rearranged oncogenes and suppressor oncogenes. The ability to isolate and molecularly analyze deletion mutations occurring in vivo in peripheral human T-lymphocytes allows the assay of DNA breakage/rejoining events. Such a system may serve as a biomarker of exposure to environmental and occupational agents which may be important in the etiology of cancer.


Subject(s)
Gene Deletion , Gene Rearrangement, T-Lymphocyte , Hypoxanthine Phosphoribosyltransferase/genetics , Neoplasms/genetics , T-Lymphocytes/enzymology , Adult , Base Sequence , Biomarkers , Humans , Male , Middle Aged , Molecular Sequence Data , Mutation
13.
Environ Mol Mutagen ; 23(2): 89-95, 1994.
Article in English | MEDLINE | ID: mdl-8143706

ABSTRACT

A multiplex polymerase chain reaction (PCR) procedure was adapted for the rapid and efficient evaluation of deletions of the hypoxanthine guanine phosphoribosyltransferase (hprt) gene in human T-lymphocytes. The hprt clonal assay was used to isolate in vivo-arising hprt-deficient T-cells from six healthy males. Mutant frequencies ranged from 9-27 x 10(-6). Simple crude cellular extracts from 223 mutants were analyzed for hprt gene deletion. Sixteen (7.2%) were found to be due to total gene deletion and 22 (9.9%) were due to partial gene deletion. The relatively high frequency of total gene deletions was caused by replicate isolates of a single mutational event as shown by single-strand conformation polymorphism (SSCP) analysis of rearranged T-cell receptor (TCR)-gamma genes. Eighteen of the 22 partial hprt gene deletion mutants were determined to be of independent origin based on a unique hprt mutation or SSCP-TCR -gamma pattern. One-half (9/18) of the partial deletion mutants involved all or part of exon 4 alone, suggesting that this region of the hprt gene is prone to deletion. The small deletions effecting exon 1 (1 mutant), exon 2 (2 mutants), and exon 4 (6 mutants) would not have been detected by conventional Southern blot analysis and may represent a new, previously unrecognized class of mutations. The ready isolation of such intragenic deletions will allow the characterization of breakpoint junctions and may provide insights into the important processes of DNA breakage and rejoining.


Subject(s)
Hypoxanthine Phosphoribosyltransferase/genetics , Mutagenicity Tests/methods , Polymerase Chain Reaction/methods , Sequence Deletion , T-Lymphocytes/enzymology , Adult , Aged , Environmental Monitoring , Humans , Male , Middle Aged
14.
Environ Health Perspect ; 101 Suppl 3: 219-24, 1993 Oct.
Article in English | MEDLINE | ID: mdl-8143621

ABSTRACT

Somatic mutations have been implicated as critical early events in carcinogenesis. Point mutations, deletions, and translocation events have been shown to activate oncogenes or inactivate suppressor oncogenes. In human population monitoring, quantitative analysis of mutation events that affect gene function is limited to those genes whose cellular phenotypes can be identified by selection procedures and to those tissues (like blood) that are accessible for analysis. In an effort to determine the frequency and types of mutations that can be detected at the hypoxanthine guanine phosphoribosyltransferase (hprt) gene, we have used the T-cell cloning assay and have developed a strategy to propagate mutants and screen for point mutations and breakage events. Early in the clonal expansion of mutants, 1-2 x 10(4) cells are prepared as a crude cell lysate, and a sample is analyzed using the multiplex polymerase chain reaction (PCR). Those mutants that yield altered DNA fragments are then expanded for Southern blot hybridization, PCR, flanking probe isolation, and DNA sequencing. To date we have found presumed point mutations, intragenic deletions, and deletions that extend outside of the hprt gene. By analyzing mutations in selectable, nonessential gene markers, it should be possible to understand mechanisms of both spontaneous and induced genetic damage. An association of these specific genetic events with human diseases and the evaluation of the ability of environmental chemicals to induce these specific types of mutations will lead to a rational basis for evaluating risks from various chemical exposures.


Subject(s)
Mutation , T-Lymphocytes/physiology , Cells, Cultured , Chromosome Mapping , Gene Deletion , Humans
15.
Mutat Res ; 301(1): 57-63, 1993 Jan.
Article in English | MEDLINE | ID: mdl-7677945

ABSTRACT

The L5178Y mouse lymphoma assay was used to examine the potential mutagenicity of three halogenated pyridine compounds. Position effects of the halogen moiety and the role of metabolic activation were analyzed based on induced mutant frequency, gross chromosome aberrations, and micronuclei. Without activation, 2-chloropyridine, 3-chloropyridine, and 2-chloro-5-trifluoromethylpyridine produced a small increase in mutant frequency; only the 2-chloropyridine activity was significantly increased with activation. All three compounds were also clastogenic as demonstrated by increases in chromosome aberrations and micronuclei (except for 2-chloro-5-trifluoromethylpyridine which did not induce micronuclei either with or without activation).


Subject(s)
Chromosome Aberrations , Hydrocarbons, Halogenated/toxicity , Mutagenicity Tests , Mutagens/toxicity , Pyridines/toxicity , Animals , Lymphoma , Mice , Micronucleus Tests , Tumor Cells, Cultured/drug effects
16.
Mutat Res ; 283(4): 255-62, 1992 Dec.
Article in English | MEDLINE | ID: mdl-1383798

ABSTRACT

A cloning assay was used to recover hprt- T-lymphocytes from adult human males. Analysis of crude cellular extracts by polymerase chain reactions (PCRs) demonstrated that 7% (16/218) of the hprt mutations were due to total deletion of the hprt gene. 14 of the 16 mutants were examined by PCR for the presence of flanking DNA to determine the extent of the deletions. The deletion mutation in 13 mutants was at least 350 kb with 5 of these deletions being at least 700 kb. The largest deletions were greater than 15 times the size of the hprt gene. Therefore, large deletions are tolerated at the hprt locus of human T-lymphocytes.


Subject(s)
Gene Deletion , Hypoxanthine Phosphoribosyltransferase/genetics , T-Lymphocytes/drug effects , X Chromosome/drug effects , Adult , Base Sequence , Cloning, Molecular , Genes/drug effects , Humans , Male , Molecular Sequence Data , Mutagenesis, Site-Directed , Polymerase Chain Reaction , Thioguanine/toxicity
17.
Mutat Res ; 283(1): 13-20, 1992 Sep.
Article in English | MEDLINE | ID: mdl-1380658

ABSTRACT

The hprt T-cell cloning assay allows the detection of mutations occurring in vivo in the hypoxanthine-guanine phosphoribosyltransferase (hprt) gene of T-lymphocytes. We have shown previously that the illegitimate activity of V(D)J recombinase accounts for about 40% of the hprt mutations in T-lymphocytes of human newborns as measured with umbilical cord blood samples (Fuscoe et al., 1991). This mechanism results in deletion of hprt exons 2 + 3. In this report, we examined a collection of 314 HPRT-deficient clones derived from adult humans for evidence that the mutations were caused by this mechanism by analyzing exons 2 + 3 deletion mutations. DNA sequence analysis of deletion breakpoint junctions showed that 8 of the mutations were the result of V(D)J recombinase activity. The frequency of the recombinase-mediated mutations was similar in the adults and newborns (2-4 x 10(-7). However, since the hprt mutant frequency is about 10-fold higher in the adult than in the newborn, the recombinase-mediated mutations account for only a few percent of the adult mutations. These mutations are likely to have occurred during early development and persist into adulthood. Unregulated expression of V(D)J recombinase activity may be an important mechanism for genomic rearrangements in the genesis of cancer.


Subject(s)
Chromosome Deletion , DNA Nucleotidyltransferases/metabolism , Genes , Hypoxanthine Phosphoribosyltransferase/genetics , T-Lymphocytes/enzymology , Adult , Base Sequence , Clone Cells , DNA/genetics , Exons , Humans , Introns , Male , Molecular Sequence Data , Oligodeoxyribonucleotides , Polymerase Chain Reaction/methods , VDJ Recombinases
18.
Mutagenesis ; 6(6): 519-25, 1991 Nov.
Article in English | MEDLINE | ID: mdl-1800900

ABSTRACT

Over the past several years, we have been evaluating the mutagenicity and clastogenicity of compounds capable of Michael-type reactions. These compounds, including acrylamide, several acrylate and methacrylate esters, vinyl sulfones, and phorone, have been evaluated using TK+/- -3.7.2C mouse lymphoma cells. Mutagenic chemicals induced increases in the number of small colony tk- deficient mutants. This suggested a clastogenic mechanism which was confirmed by demonstrating increases in aberrations and micronucleus frequencies in cultured cells. Vinyl sulfone was found to be the most effective chemical mutagen with induction of genotoxic effects at concentrations as low as 0.25 microgram/ml. The other compounds also produced positive results, but at higher concentrations. Since these compounds are known to deplete glutathione, phorone, a model glutathione depleter, was examined and found to produce similar effects as the other compounds in mouse lymphoma cells. These results suggest that the direct-acting Michael-type reaction has activity relevant to producing a genotoxic effect. Since acrylamide has been found to be a potent germ cell mutagen, this mechanism may be also relevant in the induction of heritable mutagenic risk.


Subject(s)
Ketones/toxicity , Mutagens/toxicity , Sulfones/toxicity , Acrylates/metabolism , Acrylates/toxicity , Animals , Biotransformation , Cell Count , Lymphoma , Mice , Mutagenicity Tests/statistics & numerical data , Mutagens/chemistry , Tumor Cells, Cultured/drug effects
19.
Mutat Res ; 253(1): 63-71, 1991 Aug.
Article in English | MEDLINE | ID: mdl-1651446

ABSTRACT

2-Amino-6-N-hydroxyadenine (AHA) treated L5178Y/TK (+/-)-3.7.2C mouse lymphoma cells were evaluated for mutations at the tk, hgprt, and Na+/K+ ATPase loci, as well as for gross chromosome aberrations and induction of micronuclei. In addition, AHA was evaluated for its ability to induce HGPRT mutants in CHO cells. AHA was found to induce mutations at all evaluated loci and in both cell types. The TK mutants were primarily large colonies although a few small colonies were also induced, particularly at the higher concentrations. Preliminary cytogenetic analysis of AHA-treated mouse lymphoma cells indicated that some gross aberrations but not micronuclei were induced. The 20 small-colony TK mutants evaluated by banded karyotype indicate that only a small fraction (2 of 20) showed chromosome 11 abnormalities. From these studies, it appears that AHA may be one of a very few chemicals that is capable of inducing multi-locus point mutations, with only slight clastogenic activity. Particularly at the higher concentrations, some of the mutants may contain multi-locus point mutations that result in slow growth.


Subject(s)
Adenine/analogs & derivatives , Mutagens , Adenine/toxicity , Animals , Chromosome Aberrations , Chromosome Disorders , Cricetinae , Cricetulus , Drug Resistance , Mice , Micronucleus Tests , Mutagenesis , Ouabain/toxicity , Sodium-Potassium-Exchanging ATPase/genetics , Thioguanine/toxicity , Tumor Cells, Cultured
20.
Mutagenesis ; 6(1): 35-46, 1991 Jan.
Article in English | MEDLINE | ID: mdl-2038271

ABSTRACT

Despite their widespread use and potential for significant human exposure, genotoxicity data on anthraquinones and other dyes are limited. In this study, 16 anthraquinones and one azo dye (Solvent Red 1) were selected for testing using the thymidine kinase (tk) locus and micronucleus (MN) analysis in L5178Y/TK(+/-)-3.7.2C mouse lymphoma cells. Six of the dyes were from the same lot used in the NTP rodent bioassay. The dyes used were all production lots and thus varied in their purity. Disperse Blue 7, 2-aminoanthraquinone, 1-amino-2-methylanthraquinone, Disperse Blue 3 and Disperse Red 11 were genotoxic (inducing 1814 mutants/10(6) survivors, 369 MN/1000 cells at 13% survival; 397 mutants/10(6) survivors, 196 MN/1000 cells at 21% survival; 178 mutants/10(6) survivors, 119 MN/1000 cells at 51% survival; 264 mutants/10(6) survivors, 109 MN/1000 cells at 15% survival, respectively). Reactive Blue 19 was weakly mutagenic (inducing 144 mutants/10(6) survivors, but only 8 MN/1000 cells at 13% survival). Vat Yellow 4 and Solvent Red 1, with exogenous activation, were also mutagenic (inducing 300 mutants/10(6) survivors, 18 MN/1000 cells at 57% survival, and 100 mutants/10(6) survivors and 16 MN/1000 cells at 22% survival, respectively). With activation 1-nitro-2-methylanthraquinone was judged to give an equivocal mutagenicity response. The maximum test concentration was limited for some compounds by their solubility. Those chemicals that did not induce mutation or cytotoxicity at the limits of solubility were classified separately. Compounds which were not evaluated without exogenous activation because of insolubility but were evaluated with activation include 1-nitro-2-methylanthraquinone, Solvent Red 1 and Vat Yellow 4. Compounds which were not evaluated either with or without S9 activation because of their insolubility in the culture medium include 1-amino-2,4-dibromoanthraquinone, D&C Green, Disperse Blue 1, Disperse Red 60, Vat Blue 4, Vat Blue 20, Vat Brown 1 and Vat Brown 3.


Subject(s)
Anthraquinones/toxicity , Coloring Agents/toxicity , Mutagenicity Tests , Mutation , Animals , Biotransformation , Lymphoma , Mice , Micronucleus Tests , Thymidine Kinase/genetics , Tumor Cells, Cultured
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