The Experts below are selected from a list of 252 Experts worldwide ranked by ideXlab platform
Francesco Marchetti - One of the best experts on this subject based on the ideXlab platform.
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chemically induced mutations in a Mutamouse reporter gene inform mechanisms underlying human cancer mutational signatures
Communications biology, 2020Co-Authors: Marc A. Beal, Matthew J. Meier, Carole L. Yauk, Danielle P. Leblanc, Clotilde Maurice, Jason M Obrien, Francesco MarchettiAbstract:Transgenic rodent (TGR) models use bacterial reporter genes to quantify in vivo mutagenesis. Pairing TGR assays with next-generation sequencing (NGS) enables comprehensive mutation pattern analysis to inform mutational mechanisms. We used this approach to identify 2751 independent lacZ mutations in the bone marrow of Mutamouse animals exposed to four chemical mutagens: benzo[a]pyrene, N-ethyl-N-nitrosourea, procarbazine, and triethylenemelamine. We also collected published data for 706 lacZ mutations from eight additional environmental mutagens. We report that lacZ gene sequencing generates chemical-specific mutation signatures observed in human cancers with established environmental causes. For example, the mutation signature of benzo[a]pyrene, a carcinogen present in tobacco smoke, matched the signature associated with tobacco-induced lung cancers. Our results suggest that the analysis of chemically induced mutations in the lacZ gene shortly after exposure provides an effective approach to characterize human-relevant mechanisms of carcinogenesis and propose novel environmental causes of mutation signatures observed in human cancers.
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Chemically induced mutations in a Mutamouse reporter gene inform mechanisms underlying human cancer mutational signatures
Communications Biology, 2020Co-Authors: Marc A. Beal, Matthew J. Meier, Carole L. Yauk, Danielle P. Leblanc, Clotilde Maurice, Jason M. O’brien, Francesco MarchettiAbstract:Transgenic rodent (TGR) models use bacterial reporter genes to quantify in vivo mutagenesis. Pairing TGR assays with next-generation sequencing (NGS) enables comprehensive mutation pattern analysis to inform mutational mechanisms. We used this approach to identify 2751 independent lacZ mutations in the bone marrow of Mutamouse animals exposed to four chemical mutagens: benzo[a]pyrene, N -ethyl- N -nitrosourea, procarbazine, and triethylenemelamine. We also collected published data for 706 lacZ mutations from eight additional environmental mutagens. We report that lacZ gene sequencing generates chemical-specific mutation signatures observed in human cancers with established environmental causes. For example, the mutation signature of benzo[a]pyrene, a carcinogen present in tobacco smoke, matched the signature associated with tobacco-induced lung cancers. Our results suggest that the analysis of chemically induced mutations in the lacZ gene shortly after exposure provides an effective approach to characterize human-relevant mechanisms of carcinogenesis and propose novel environmental causes of mutation signatures observed in human cancers. Marc A. Beal, Matt J. Meier et al. use a transgenic rodent model with NGS to inform mutational signature analyses. They analyze lacZ mutations from the bone marrow of Mutamouse animals exposed to chemical mutagens. They report that lacZ sequencing generates chemical-specific mutation signatures observed in human cancers with established environmental causes.
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Sequencing Chemically Induced Mutations in the Mutamouse Lacz Reporter Gene Identifies Human Cancer Mutational Signatures
2019Co-Authors: Marc A. Beal, Matthew J. Meier, Carole L. Yauk, Danielle P. Leblanc, Clotilde Maurice, Jason M. O'brien, Francesco MarchettiAbstract:ABSTRACT Transgenic rodent (TGR) models use bacterial reporter genes to quantify in vivo mutagenesis. Pairing TGR assays with next-generation sequencing (NGS) enables comprehensive mutation spectrum analysis to inform mutational mechanisms. We used this approach to identify 2,751 independent lacZ mutations in the bone marrow of Mutamouse animals exposed to four chemical mutagens: benzo[a]pyrene, N-ethyl-N-nitrosourea, procarbazine, and triethylenemelamine. We also collected published data for 706 lacZ mutations from eight additional environmental mutagens. We demonstrate that lacZ gene sequencing generates chemical-specific mutation signatures observed in human cancers with established environmental causes. For example, the mutation signature of benzo[a]pyrene, a potent carcinogen in tobacco smoke, matched the signature associated with tobacco-induced lung cancers. Our results show that the analysis of chemically induced mutations in the lacZ gene shortly after exposure provides an effective approach to characterize human-relevant mechanisms of carcinogenesis and identify novel environmental causes of mutation signatures observed in human cancers.
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Whole Genome Sequencing of the Mutamouse Model Reveals Strain- and Colony-Level Variation, and Genomic Features of the Transgene Integration Site
Scientific Reports, 2019Co-Authors: Matthew J. Meier, Marc A. Beal, Andrew Schoenrock, Carole L. Yauk, Francesco MarchettiAbstract:The Mutamouse transgenic rodent model is widely used for assessing in vivo mutagenicity. Here, we report the characterization of Mutamouse’s whole genome sequence and its genetic variants compared to the C57BL/6 reference genome. High coverage (>50X) next-generation sequencing (NGS) of whole genomes from multiple Mutamouse animals from the Health Canada (HC) colony showed ~5 million SNVs per genome, ~20% of which are putatively novel. Sequencing of two animals from a geographically separated colony at Covance indicated that, over the course of 23 years, each colony accumulated 47,847 (HC) and 17,677 (Covance) non-parental homozygous single nucleotide variants. We found no novel nonsense or missense mutations that impair the Mutamouse response to genotoxic agents. Pairing sequencing data with array comparative genomic hybridization (aCGH) improved the accuracy and resolution of copy number variants (CNVs) calls and identified 300 genomic regions with CNVs. We also used long-read sequence technology (PacBio) to show that the transgene integration site involved a large deletion event with multiple inversions and rearrangements near a retrotransposon. The Mutamouse genome gives important genetic context to studies using this model, offers insight on the mechanisms of structural variant formation, and contributes a framework to analyze aCGH results alongside NGS data.
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Integrated In Vivo Genotoxicity Assessment of Procarbazine Hydrochloride Demonstrates Induction of Pig-a and LacZ Mutations, and Micronuclei, in Mutamouse Hematopoietic Cells.
Environmental and Molecular Mutagenesis, 2019Co-Authors: Clotilde Maurice, Carole L. Yauk, Stephen D. Dertinger, Francesco MarchettiAbstract:Procarbazine hydrochloride (PCH) is a DNA-reactive hematopoietic carcinogen with potent and well-characterized clastogenic activity. However, there is a paucity of in vivo mutagenesis data for PCH, and in vitro assays often fail to detect the genotoxic effects of PCH due to the complexity of its metabolic activation. We comprehensively evaluated the in vivo genotoxicity of PCH on hematopoietic cells of male Mutamouse transgenic rodents using a study design that facilitated assessments of micronuclei and Pig-a mutation in circulating erythrocytes, and lacZ mutant frequencies in bone marrow. Mice were orally exposed to PCH (0, 6.25, 12.5, and 25 mg/kg/day) for 28 consecutive days. Blood samples collected 2 days after cessation of treatment exhibited significant dose-related induction of micronuclei in both immature and mature erythrocytes. Bone marrow and blood collected 3 and 70 days after cessation of treatment also showed significantly elevated mutant frequencies in both the lacZ and Pig-a assays even at the lowest dose tested. PCH-induced lacZ and Pig-a (immature and mature erythrocytes) mutant frequencies were highly correlated, with R2 values ≥0.956, with the exception of lacZ vs. Pig-a mutants in mature erythrocytes at the 70-day time point (R2 = 0.902). These results show that PCH is genotoxic in vivo and demonstrate that the complex metabolism and resulting genotoxicity of PCH is best evaluated in intact animal models. Our results further support the concept that multiple biomarkers of genotoxicity, especially hematopoietic cell genotoxicity, can be readily combined into one study provided that adequate attention is given to manifestation times. Environ. Mol. Mutagen. 60:505-512, 2019. © 2018 Her Majesty the Queen in Right of Canada.
Carole L. Yauk - One of the best experts on this subject based on the ideXlab platform.
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chemically induced mutations in a Mutamouse reporter gene inform mechanisms underlying human cancer mutational signatures
Communications biology, 2020Co-Authors: Marc A. Beal, Matthew J. Meier, Carole L. Yauk, Danielle P. Leblanc, Clotilde Maurice, Jason M Obrien, Francesco MarchettiAbstract:Transgenic rodent (TGR) models use bacterial reporter genes to quantify in vivo mutagenesis. Pairing TGR assays with next-generation sequencing (NGS) enables comprehensive mutation pattern analysis to inform mutational mechanisms. We used this approach to identify 2751 independent lacZ mutations in the bone marrow of Mutamouse animals exposed to four chemical mutagens: benzo[a]pyrene, N-ethyl-N-nitrosourea, procarbazine, and triethylenemelamine. We also collected published data for 706 lacZ mutations from eight additional environmental mutagens. We report that lacZ gene sequencing generates chemical-specific mutation signatures observed in human cancers with established environmental causes. For example, the mutation signature of benzo[a]pyrene, a carcinogen present in tobacco smoke, matched the signature associated with tobacco-induced lung cancers. Our results suggest that the analysis of chemically induced mutations in the lacZ gene shortly after exposure provides an effective approach to characterize human-relevant mechanisms of carcinogenesis and propose novel environmental causes of mutation signatures observed in human cancers.
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Chemically induced mutations in a Mutamouse reporter gene inform mechanisms underlying human cancer mutational signatures
Communications Biology, 2020Co-Authors: Marc A. Beal, Matthew J. Meier, Carole L. Yauk, Danielle P. Leblanc, Clotilde Maurice, Jason M. O’brien, Francesco MarchettiAbstract:Transgenic rodent (TGR) models use bacterial reporter genes to quantify in vivo mutagenesis. Pairing TGR assays with next-generation sequencing (NGS) enables comprehensive mutation pattern analysis to inform mutational mechanisms. We used this approach to identify 2751 independent lacZ mutations in the bone marrow of Mutamouse animals exposed to four chemical mutagens: benzo[a]pyrene, N -ethyl- N -nitrosourea, procarbazine, and triethylenemelamine. We also collected published data for 706 lacZ mutations from eight additional environmental mutagens. We report that lacZ gene sequencing generates chemical-specific mutation signatures observed in human cancers with established environmental causes. For example, the mutation signature of benzo[a]pyrene, a carcinogen present in tobacco smoke, matched the signature associated with tobacco-induced lung cancers. Our results suggest that the analysis of chemically induced mutations in the lacZ gene shortly after exposure provides an effective approach to characterize human-relevant mechanisms of carcinogenesis and propose novel environmental causes of mutation signatures observed in human cancers. Marc A. Beal, Matt J. Meier et al. use a transgenic rodent model with NGS to inform mutational signature analyses. They analyze lacZ mutations from the bone marrow of Mutamouse animals exposed to chemical mutagens. They report that lacZ sequencing generates chemical-specific mutation signatures observed in human cancers with established environmental causes.
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Sequencing Chemically Induced Mutations in the Mutamouse Lacz Reporter Gene Identifies Human Cancer Mutational Signatures
2019Co-Authors: Marc A. Beal, Matthew J. Meier, Carole L. Yauk, Danielle P. Leblanc, Clotilde Maurice, Jason M. O'brien, Francesco MarchettiAbstract:ABSTRACT Transgenic rodent (TGR) models use bacterial reporter genes to quantify in vivo mutagenesis. Pairing TGR assays with next-generation sequencing (NGS) enables comprehensive mutation spectrum analysis to inform mutational mechanisms. We used this approach to identify 2,751 independent lacZ mutations in the bone marrow of Mutamouse animals exposed to four chemical mutagens: benzo[a]pyrene, N-ethyl-N-nitrosourea, procarbazine, and triethylenemelamine. We also collected published data for 706 lacZ mutations from eight additional environmental mutagens. We demonstrate that lacZ gene sequencing generates chemical-specific mutation signatures observed in human cancers with established environmental causes. For example, the mutation signature of benzo[a]pyrene, a potent carcinogen in tobacco smoke, matched the signature associated with tobacco-induced lung cancers. Our results show that the analysis of chemically induced mutations in the lacZ gene shortly after exposure provides an effective approach to characterize human-relevant mechanisms of carcinogenesis and identify novel environmental causes of mutation signatures observed in human cancers.
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Whole Genome Sequencing of the Mutamouse Model Reveals Strain- and Colony-Level Variation, and Genomic Features of the Transgene Integration Site
Scientific Reports, 2019Co-Authors: Matthew J. Meier, Marc A. Beal, Andrew Schoenrock, Carole L. Yauk, Francesco MarchettiAbstract:The Mutamouse transgenic rodent model is widely used for assessing in vivo mutagenicity. Here, we report the characterization of Mutamouse’s whole genome sequence and its genetic variants compared to the C57BL/6 reference genome. High coverage (>50X) next-generation sequencing (NGS) of whole genomes from multiple Mutamouse animals from the Health Canada (HC) colony showed ~5 million SNVs per genome, ~20% of which are putatively novel. Sequencing of two animals from a geographically separated colony at Covance indicated that, over the course of 23 years, each colony accumulated 47,847 (HC) and 17,677 (Covance) non-parental homozygous single nucleotide variants. We found no novel nonsense or missense mutations that impair the Mutamouse response to genotoxic agents. Pairing sequencing data with array comparative genomic hybridization (aCGH) improved the accuracy and resolution of copy number variants (CNVs) calls and identified 300 genomic regions with CNVs. We also used long-read sequence technology (PacBio) to show that the transgene integration site involved a large deletion event with multiple inversions and rearrangements near a retrotransposon. The Mutamouse genome gives important genetic context to studies using this model, offers insight on the mechanisms of structural variant formation, and contributes a framework to analyze aCGH results alongside NGS data.
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Integrated In Vivo Genotoxicity Assessment of Procarbazine Hydrochloride Demonstrates Induction of Pig-a and LacZ Mutations, and Micronuclei, in Mutamouse Hematopoietic Cells.
Environmental and Molecular Mutagenesis, 2019Co-Authors: Clotilde Maurice, Carole L. Yauk, Stephen D. Dertinger, Francesco MarchettiAbstract:Procarbazine hydrochloride (PCH) is a DNA-reactive hematopoietic carcinogen with potent and well-characterized clastogenic activity. However, there is a paucity of in vivo mutagenesis data for PCH, and in vitro assays often fail to detect the genotoxic effects of PCH due to the complexity of its metabolic activation. We comprehensively evaluated the in vivo genotoxicity of PCH on hematopoietic cells of male Mutamouse transgenic rodents using a study design that facilitated assessments of micronuclei and Pig-a mutation in circulating erythrocytes, and lacZ mutant frequencies in bone marrow. Mice were orally exposed to PCH (0, 6.25, 12.5, and 25 mg/kg/day) for 28 consecutive days. Blood samples collected 2 days after cessation of treatment exhibited significant dose-related induction of micronuclei in both immature and mature erythrocytes. Bone marrow and blood collected 3 and 70 days after cessation of treatment also showed significantly elevated mutant frequencies in both the lacZ and Pig-a assays even at the lowest dose tested. PCH-induced lacZ and Pig-a (immature and mature erythrocytes) mutant frequencies were highly correlated, with R2 values ≥0.956, with the exception of lacZ vs. Pig-a mutants in mature erythrocytes at the 70-day time point (R2 = 0.902). These results show that PCH is genotoxic in vivo and demonstrate that the complex metabolism and resulting genotoxicity of PCH is best evaluated in intact animal models. Our results further support the concept that multiple biomarkers of genotoxicity, especially hematopoietic cell genotoxicity, can be readily combined into one study provided that adequate attention is given to manifestation times. Environ. Mol. Mutagen. 60:505-512, 2019. © 2018 Her Majesty the Queen in Right of Canada.
Werner Lutz - One of the best experts on this subject based on the ideXlab platform.
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the viracept nelfinavir ethyl methanesulfonate case a threshold risk assessment for human exposure to a genotoxic drug contamination
Toxicology Letters, 2009Co-Authors: Werner LutzAbstract:Abstract In May 2007, the F. Hoffmann-La Roche Company became aware of a contamination of Viracept (nelfinavir) tablets by the mutagenic DNA-ethylating agent ethyl methanesulfonate (EMS) as a result of a production incident. HIV-patients could have been exposed for 3 months to daily doses of up to 2.75 mg EMS, i.e., about 50 μg/kg per day. In this special issue, 12 manuscripts have been assembled to provide comprehensive insight in what happened and how the incident was managed by Roche and handled by the regulatory agencies. In the first four papers, the course of events and the toxicological information available at the outset are summarized and a traditional cancer risk assessment on the basis of a linear default dose–response is made. Three articles then report on the experiments performed for an improved risk assessment. A standard 4-week toxicity study with EMS in the rat indicated an NOAEL of 20 mg/kg per day. Extensive studies on the genotoxicity showed threshold-like dose responses for both chromosome damage (bone marrow micronucleus test) and gene mutation (lacZ transgenic Mutamouse test) in various organs of mice treated for up to 4 weeks, whereas ethylation of hemoglobin at the N-terminal valine increased linearly with dose. The difference between adduct formation in DNA and protein was interpreted by repair of DNA adducts that becomes saturated above a threshold concentration of EMS, regarded as the metrics for the rate of DNA ethylation. Elaborate toxicokinetic investigations in various animal species, coupled to appropriate modeling, were performed in order to extrapolate the animal data to humans. Using a threshold risk assessment based on estimated cmax of EMS, a safety factor of 370 was derived for maximum doses ingested by Viracept patients. A number of critical points are addressed in this editorial, concerning (i) definitions and types of “thresholds”, (ii) estimation of a confidence limit for a slope below the threshold dose, interpreted as an increment within background variation, (iii) implementation for other mutagens and for human risk assessment.
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the viracept nelfinavir ethyl methanesulfonate case a threshold risk assessment for human exposure to a genotoxic drug contamination
Toxicology Letters, 2009Co-Authors: Werner LutzAbstract:Abstract In May 2007, the F. Hoffmann-La Roche Company became aware of a contamination of Viracept (nelfinavir) tablets by the mutagenic DNA-ethylating agent ethyl methanesulfonate (EMS) as a result of a production incident. HIV-patients could have been exposed for 3 months to daily doses of up to 2.75 mg EMS, i.e., about 50 μg/kg per day. In this special issue, 12 manuscripts have been assembled to provide comprehensive insight in what happened and how the incident was managed by Roche and handled by the regulatory agencies. In the first four papers, the course of events and the toxicological information available at the outset are summarized and a traditional cancer risk assessment on the basis of a linear default dose–response is made. Three articles then report on the experiments performed for an improved risk assessment. A standard 4-week toxicity study with EMS in the rat indicated an NOAEL of 20 mg/kg per day. Extensive studies on the genotoxicity showed threshold-like dose responses for both chromosome damage (bone marrow micronucleus test) and gene mutation (lacZ transgenic Mutamouse test) in various organs of mice treated for up to 4 weeks, whereas ethylation of hemoglobin at the N-terminal valine increased linearly with dose. The difference between adduct formation in DNA and protein was interpreted by repair of DNA adducts that becomes saturated above a threshold concentration of EMS, regarded as the metrics for the rate of DNA ethylation. Elaborate toxicokinetic investigations in various animal species, coupled to appropriate modeling, were performed in order to extrapolate the animal data to humans. Using a threshold risk assessment based on estimated cmax of EMS, a safety factor of 370 was derived for maximum doses ingested by Viracept patients. A number of critical points are addressed in this editorial, concerning (i) definitions and types of “thresholds”, (ii) estimation of a confidence limit for a slope below the threshold dose, interpreted as an increment within background variation, (iii) implementation for other mutagens and for human risk assessment.
Clotilde Maurice - One of the best experts on this subject based on the ideXlab platform.
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Chemically induced mutations in a Mutamouse reporter gene inform mechanisms underlying human cancer mutational signatures
Communications Biology, 2020Co-Authors: Marc A. Beal, Matthew J. Meier, Carole L. Yauk, Danielle P. Leblanc, Clotilde Maurice, Jason M. O’brien, Francesco MarchettiAbstract:Transgenic rodent (TGR) models use bacterial reporter genes to quantify in vivo mutagenesis. Pairing TGR assays with next-generation sequencing (NGS) enables comprehensive mutation pattern analysis to inform mutational mechanisms. We used this approach to identify 2751 independent lacZ mutations in the bone marrow of Mutamouse animals exposed to four chemical mutagens: benzo[a]pyrene, N -ethyl- N -nitrosourea, procarbazine, and triethylenemelamine. We also collected published data for 706 lacZ mutations from eight additional environmental mutagens. We report that lacZ gene sequencing generates chemical-specific mutation signatures observed in human cancers with established environmental causes. For example, the mutation signature of benzo[a]pyrene, a carcinogen present in tobacco smoke, matched the signature associated with tobacco-induced lung cancers. Our results suggest that the analysis of chemically induced mutations in the lacZ gene shortly after exposure provides an effective approach to characterize human-relevant mechanisms of carcinogenesis and propose novel environmental causes of mutation signatures observed in human cancers. Marc A. Beal, Matt J. Meier et al. use a transgenic rodent model with NGS to inform mutational signature analyses. They analyze lacZ mutations from the bone marrow of Mutamouse animals exposed to chemical mutagens. They report that lacZ sequencing generates chemical-specific mutation signatures observed in human cancers with established environmental causes.
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chemically induced mutations in a Mutamouse reporter gene inform mechanisms underlying human cancer mutational signatures
Communications biology, 2020Co-Authors: Marc A. Beal, Matthew J. Meier, Carole L. Yauk, Danielle P. Leblanc, Clotilde Maurice, Jason M Obrien, Francesco MarchettiAbstract:Transgenic rodent (TGR) models use bacterial reporter genes to quantify in vivo mutagenesis. Pairing TGR assays with next-generation sequencing (NGS) enables comprehensive mutation pattern analysis to inform mutational mechanisms. We used this approach to identify 2751 independent lacZ mutations in the bone marrow of Mutamouse animals exposed to four chemical mutagens: benzo[a]pyrene, N-ethyl-N-nitrosourea, procarbazine, and triethylenemelamine. We also collected published data for 706 lacZ mutations from eight additional environmental mutagens. We report that lacZ gene sequencing generates chemical-specific mutation signatures observed in human cancers with established environmental causes. For example, the mutation signature of benzo[a]pyrene, a carcinogen present in tobacco smoke, matched the signature associated with tobacco-induced lung cancers. Our results suggest that the analysis of chemically induced mutations in the lacZ gene shortly after exposure provides an effective approach to characterize human-relevant mechanisms of carcinogenesis and propose novel environmental causes of mutation signatures observed in human cancers.
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Sequencing Chemically Induced Mutations in the Mutamouse Lacz Reporter Gene Identifies Human Cancer Mutational Signatures
2019Co-Authors: Marc A. Beal, Matthew J. Meier, Carole L. Yauk, Danielle P. Leblanc, Clotilde Maurice, Jason M. O'brien, Francesco MarchettiAbstract:ABSTRACT Transgenic rodent (TGR) models use bacterial reporter genes to quantify in vivo mutagenesis. Pairing TGR assays with next-generation sequencing (NGS) enables comprehensive mutation spectrum analysis to inform mutational mechanisms. We used this approach to identify 2,751 independent lacZ mutations in the bone marrow of Mutamouse animals exposed to four chemical mutagens: benzo[a]pyrene, N-ethyl-N-nitrosourea, procarbazine, and triethylenemelamine. We also collected published data for 706 lacZ mutations from eight additional environmental mutagens. We demonstrate that lacZ gene sequencing generates chemical-specific mutation signatures observed in human cancers with established environmental causes. For example, the mutation signature of benzo[a]pyrene, a potent carcinogen in tobacco smoke, matched the signature associated with tobacco-induced lung cancers. Our results show that the analysis of chemically induced mutations in the lacZ gene shortly after exposure provides an effective approach to characterize human-relevant mechanisms of carcinogenesis and identify novel environmental causes of mutation signatures observed in human cancers.
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Integrated In Vivo Genotoxicity Assessment of Procarbazine Hydrochloride Demonstrates Induction of Pig-a and LacZ Mutations, and Micronuclei, in Mutamouse Hematopoietic Cells.
Environmental and Molecular Mutagenesis, 2019Co-Authors: Clotilde Maurice, Carole L. Yauk, Stephen D. Dertinger, Francesco MarchettiAbstract:Procarbazine hydrochloride (PCH) is a DNA-reactive hematopoietic carcinogen with potent and well-characterized clastogenic activity. However, there is a paucity of in vivo mutagenesis data for PCH, and in vitro assays often fail to detect the genotoxic effects of PCH due to the complexity of its metabolic activation. We comprehensively evaluated the in vivo genotoxicity of PCH on hematopoietic cells of male Mutamouse transgenic rodents using a study design that facilitated assessments of micronuclei and Pig-a mutation in circulating erythrocytes, and lacZ mutant frequencies in bone marrow. Mice were orally exposed to PCH (0, 6.25, 12.5, and 25 mg/kg/day) for 28 consecutive days. Blood samples collected 2 days after cessation of treatment exhibited significant dose-related induction of micronuclei in both immature and mature erythrocytes. Bone marrow and blood collected 3 and 70 days after cessation of treatment also showed significantly elevated mutant frequencies in both the lacZ and Pig-a assays even at the lowest dose tested. PCH-induced lacZ and Pig-a (immature and mature erythrocytes) mutant frequencies were highly correlated, with R2 values ≥0.956, with the exception of lacZ vs. Pig-a mutants in mature erythrocytes at the 70-day time point (R2 = 0.902). These results show that PCH is genotoxic in vivo and demonstrate that the complex metabolism and resulting genotoxicity of PCH is best evaluated in intact animal models. Our results further support the concept that multiple biomarkers of genotoxicity, especially hematopoietic cell genotoxicity, can be readily combined into one study provided that adequate attention is given to manifestation times. Environ. Mol. Mutagen. 60:505-512, 2019. © 2018 Her Majesty the Queen in Right of Canada.
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integration of sperm dna damage assessment into oecd test guidelines for genotoxicity testing using the Mutamouse model
Toxicology and Applied Pharmacology, 2018Co-Authors: Clotilde Maurice, Carole L. Yauk, Jason M Obrien, Francesco MarchettiAbstract:The Organisation for Economic Co-operation and Development (OECD) endorses test guidelines (TG) for identifying chemicals that are genotoxic, such as the transgenic rodent gene mutation assay (TG 488). Current OECD TG do not include assays for sperm DNA damage resulting in a critical testing gap. We evaluated the performance of the Sperm Chromatin Structure Assay (SCSA) and the Terminal Deoxynucleotidyl Transferase-Mediated Deoxyuridine Triphosphate Nick end Labeling (TUNEL) assay to detect sperm DNA damage within the recommended TG 488 protocol. Mutamouse males received 0, 0.5, 1, or 2 mg/kg/day triethylenemelamine (TEM), a multifunctional alkylating agent, for 28 days orally and tissues were collected two (blood) and three (sperm and bone marrow) days later. TEM significantly increased the frequency of lacZ mutants in bone marrow, and of micronuclei (MN) in both reticulocytes (%MN-RET) and normochromatic erythrocytes (%MN-NCE) in a dose-dependent manner (P < 0.05). The percentage of DNA fragmentation index (%DFI) and %TUNEL positive cells demonstrated dose-related increases in sperm (P < 0.05), and the two assay results were strongly correlated (R = 0.9298). Within the same animal, a good correlation was observed between %MN-NCE and %DFI (R = 0.7189). Finally, benchmark dose modelling (BMD) showed comparable BMD10 values among the somatic and germ cell assays. Our results suggest that sperm DNA damage assays can be easily integrated into standard OECD designs investigating genotoxicity in somatic tissues to provide key information on whether a chemical is genotoxic in germ cells and impact its risk assessment.
Marc A. Beal - One of the best experts on this subject based on the ideXlab platform.
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Chemically induced mutations in a Mutamouse reporter gene inform mechanisms underlying human cancer mutational signatures
Communications Biology, 2020Co-Authors: Marc A. Beal, Matthew J. Meier, Carole L. Yauk, Danielle P. Leblanc, Clotilde Maurice, Jason M. O’brien, Francesco MarchettiAbstract:Transgenic rodent (TGR) models use bacterial reporter genes to quantify in vivo mutagenesis. Pairing TGR assays with next-generation sequencing (NGS) enables comprehensive mutation pattern analysis to inform mutational mechanisms. We used this approach to identify 2751 independent lacZ mutations in the bone marrow of Mutamouse animals exposed to four chemical mutagens: benzo[a]pyrene, N -ethyl- N -nitrosourea, procarbazine, and triethylenemelamine. We also collected published data for 706 lacZ mutations from eight additional environmental mutagens. We report that lacZ gene sequencing generates chemical-specific mutation signatures observed in human cancers with established environmental causes. For example, the mutation signature of benzo[a]pyrene, a carcinogen present in tobacco smoke, matched the signature associated with tobacco-induced lung cancers. Our results suggest that the analysis of chemically induced mutations in the lacZ gene shortly after exposure provides an effective approach to characterize human-relevant mechanisms of carcinogenesis and propose novel environmental causes of mutation signatures observed in human cancers. Marc A. Beal, Matt J. Meier et al. use a transgenic rodent model with NGS to inform mutational signature analyses. They analyze lacZ mutations from the bone marrow of Mutamouse animals exposed to chemical mutagens. They report that lacZ sequencing generates chemical-specific mutation signatures observed in human cancers with established environmental causes.
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chemically induced mutations in a Mutamouse reporter gene inform mechanisms underlying human cancer mutational signatures
Communications biology, 2020Co-Authors: Marc A. Beal, Matthew J. Meier, Carole L. Yauk, Danielle P. Leblanc, Clotilde Maurice, Jason M Obrien, Francesco MarchettiAbstract:Transgenic rodent (TGR) models use bacterial reporter genes to quantify in vivo mutagenesis. Pairing TGR assays with next-generation sequencing (NGS) enables comprehensive mutation pattern analysis to inform mutational mechanisms. We used this approach to identify 2751 independent lacZ mutations in the bone marrow of Mutamouse animals exposed to four chemical mutagens: benzo[a]pyrene, N-ethyl-N-nitrosourea, procarbazine, and triethylenemelamine. We also collected published data for 706 lacZ mutations from eight additional environmental mutagens. We report that lacZ gene sequencing generates chemical-specific mutation signatures observed in human cancers with established environmental causes. For example, the mutation signature of benzo[a]pyrene, a carcinogen present in tobacco smoke, matched the signature associated with tobacco-induced lung cancers. Our results suggest that the analysis of chemically induced mutations in the lacZ gene shortly after exposure provides an effective approach to characterize human-relevant mechanisms of carcinogenesis and propose novel environmental causes of mutation signatures observed in human cancers.
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Sequencing Chemically Induced Mutations in the Mutamouse Lacz Reporter Gene Identifies Human Cancer Mutational Signatures
2019Co-Authors: Marc A. Beal, Matthew J. Meier, Carole L. Yauk, Danielle P. Leblanc, Clotilde Maurice, Jason M. O'brien, Francesco MarchettiAbstract:ABSTRACT Transgenic rodent (TGR) models use bacterial reporter genes to quantify in vivo mutagenesis. Pairing TGR assays with next-generation sequencing (NGS) enables comprehensive mutation spectrum analysis to inform mutational mechanisms. We used this approach to identify 2,751 independent lacZ mutations in the bone marrow of Mutamouse animals exposed to four chemical mutagens: benzo[a]pyrene, N-ethyl-N-nitrosourea, procarbazine, and triethylenemelamine. We also collected published data for 706 lacZ mutations from eight additional environmental mutagens. We demonstrate that lacZ gene sequencing generates chemical-specific mutation signatures observed in human cancers with established environmental causes. For example, the mutation signature of benzo[a]pyrene, a potent carcinogen in tobacco smoke, matched the signature associated with tobacco-induced lung cancers. Our results show that the analysis of chemically induced mutations in the lacZ gene shortly after exposure provides an effective approach to characterize human-relevant mechanisms of carcinogenesis and identify novel environmental causes of mutation signatures observed in human cancers.
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Whole Genome Sequencing of the Mutamouse Model Reveals Strain- and Colony-Level Variation, and Genomic Features of the Transgene Integration Site
Scientific Reports, 2019Co-Authors: Matthew J. Meier, Marc A. Beal, Andrew Schoenrock, Carole L. Yauk, Francesco MarchettiAbstract:The Mutamouse transgenic rodent model is widely used for assessing in vivo mutagenicity. Here, we report the characterization of Mutamouse’s whole genome sequence and its genetic variants compared to the C57BL/6 reference genome. High coverage (>50X) next-generation sequencing (NGS) of whole genomes from multiple Mutamouse animals from the Health Canada (HC) colony showed ~5 million SNVs per genome, ~20% of which are putatively novel. Sequencing of two animals from a geographically separated colony at Covance indicated that, over the course of 23 years, each colony accumulated 47,847 (HC) and 17,677 (Covance) non-parental homozygous single nucleotide variants. We found no novel nonsense or missense mutations that impair the Mutamouse response to genotoxic agents. Pairing sequencing data with array comparative genomic hybridization (aCGH) improved the accuracy and resolution of copy number variants (CNVs) calls and identified 300 genomic regions with CNVs. We also used long-read sequence technology (PacBio) to show that the transgene integration site involved a large deletion event with multiple inversions and rearrangements near a retrotransposon. The Mutamouse genome gives important genetic context to studies using this model, offers insight on the mechanisms of structural variant formation, and contributes a framework to analyze aCGH results alongside NGS data.
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In Utero Exposure to Benzo[a]pyrene Induces Ovarian Mutations at Doses That Deplete Ovarian Follicles in Mice
Environmental and Molecular Mutagenesis, 2018Co-Authors: Ulrike Luderer, Matthew J. Meier, Marc A. Beal, Carole L. Yauk, Gregory W. Lawson, Francesco MarchettiAbstract:Polycyclic aromatic hydrocarbons like benzo[a]pyrene (BaP) are ubiquitous environmental contaminants formed during incomplete combustion of organic materials. Our prior work showed that transplacental exposure to BaP depletes ovarian follicles and increases prevalence of epithelial ovarian tumors later in life. We used the Mutamouse transgenic rodent model to address the hypothesis that ovarian mutations play a role in tumorigenesis caused by prenatal exposure to BaP. Pregnant Mutamouse females were treated with 0, 10, 20, or 40 mg/(kg day) BaP orally on gestational days 7-16, covering critical windows of ovarian development. Female offspring were euthanized at 10 weeks of age; some ovaries with oviducts were processed for follicle counting; other ovaries/oviducts and bone marrow were processed for determination of lacZ mutant frequency (MF). Mutant plaques were pooled within dose groups and sequenced to determine the mutation spectrum. BaP exposure caused highly significant dose-related decreases in ovarian follicles and increases in ovarian/oviductal and bone marrow mutant frequencies at all doses. Absence of follicles, cell packets, and epithelial tubular structures were observed with 20 and 40 mg/(kg day) BaP. Depletion of ovarian germ cells was inversely associated with ovarian MF. BaP induced primarily G > T and G > C transversions and deletions in ovaries/oviducts and bone marrow cells and produced a mutation signature highly consistent with that of tobacco smoking in human cancers. Overall, our results show that prenatal BaP exposure significantly depletes ovarian germ cells, causes histopathological abnormalities, and increases the burden of ovarian/oviductal mutations, which may be involved in pathogenesis of epithelial ovarian tumors. Environ. Mol. Mutagen. 60:410-420, 2019. © 2018 Her Majesty the Queen in Right of Canada.
