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Meta-analysis to better integrate human variability in toxicokinetic: CYP2D6-related uncertainty factors
Abstracts / Toxicology Letters 238S (2015) S56–S383
S105
P03-055 Tobacco smoking induces specifically the expansion of a subpopulation of T cells expressing the homing and HIV co-receptor GPR15
P03-056 Novel analytical method to measure formaldehyde release from heated hair straightening cosmetic products: Impact on risk assessment
M. Bauer 1,∗ , G. Linsel 2 , K. Offenberg 1 , U. Sack 3 , H. Knaak 3 , G. Herberth 1
C.L. Galli 1,∗ , F. Bettin 2 , P. Metra 2 , P. Fidente 1 , M. Marinovich 1 , D. Dominicis 2
1
1
Helmholtz Centre for Environmental Research, Environmental Immunology, Leipzig, Germany 2 Federal Institute for Occupational Safety and Health, Berlin, Germany 3 Medical Faculty, University of Leipzig, Institute of Clinical Immunology, Leipzig, Germany Purpose: Tobacco smoking seriously raises mortality and morbidity worldwide. It increases risk for diseases in people living with HIV and worsens the severity of colon-restricted Crohn’s disease. Understanding the mechanisms through which smoking increases health risk to these disorders may establish new prevention or treatment strategies. Methods: Healthy adult subjects of two independent cohorts (Duck Operations (n = 93) and Blood Donor (n = 123) Cohorts) were implemented to study gene and protein expression in peripheral white blood cells by PCR and flow cytometry, respectively. Specimens of bronchoalveolar lavage were used to clarify smoking induced homing into lung. Results: Analyzing the lymphocyte homing and HIV co-receptor GPR15 on leucocytes in blood we found a high specific 3.6 fold (p < 0.0001) enrichment of GPR15-positive T cells in smokers compared to nonsmokers. The difference was independent on gender and age but reversal after cessation. In contrast to female smokers of the Duck Operations Cohort, the GPR15 gene expression in male followed an age-dependency (correlation coefficient r2 = 0.48, p < 0.00001). With regard to other known organ-specific homing receptors the GPR15 + T cells expressed partially CCR4 and CCR7 and to high extent the lymphnode homing CD62L. The assumption that GPR15 may serve additionally as a homing receptor for lymphocyte trafficking into human lung has not been confirmed analyzing GPR15 expression in leucocytes of bronchoalveolar lavage. Conclusion: For the first time we have illuminated a very sensitive and specific tobacco smoke induced enrichment of a specialized T cell type in blood as a result of an adaptive immune response toward tobacco smoke pollution of the human body. This finding may explain the tobacco-smoke promoted affection of the hereby mentioned diseases by helping HIV-virus to spread or by maintaining of inflammation in colon, into which this cell type specifically home, via tobacco smoking induced excess of GPR15+ cells in blood. http://dx.doi.org/10.1016/j.toxlet.2015.08.344
Università degli Studi di Milano, Dipartimento di Scienze Farmacologiche e Biomolecolari, Milano, Italy 2 R&D Department Chelab-Silliker a Mérieux Nutriscience Company, Resana, Italy In Europe, formaldehyde (FA) is permitted for use in personal care products at concentrations ≤ 0.2 g/100. Our method captures and collects the FA released into the air from heated cosmetic products by derivatization with 2,4-dinitrophenylhydrazine and analysis by HPLC/DAD instrument. 42% of 72 market samples analyzed showed FA concentrations very close to or above the threshold value (0.074 g/100 g calculated as FA), whereas 11 samples, negative using the official method of reference, were close to or above the threshold value. This may pose a health problem for occasional users and professional hair stylists. The values of FA equivalents released from the selected hair straightening products transformed into ppm (FA mg/1.23) extrapolated to the hypothesized volume of 1 m3 (FA mg/1.23/1 m3 ), for the exposure of the consumer and the operator, are mostly far above the limits established by DFG (0.3 ppm) and OSHA STEL (2 ppm, 15 min) for both stylists and customers receiving a 100 g (median 67.48 ppm) or a 35 g (median 23.62 ppm) treatment, respectively, even when the FA concentration in the product is below the 0.074% limit. http://dx.doi.org/10.1016/j.toxlet.2015.08.345
P03-057 Meta-analysis to better integrate human variability in toxicokinetic: CYP2D6-related uncertainty factors C. Béchaux 1,∗ , B. Amzal 2 , A. Crépet 1 , J.-L. Dorne 3 1
ANSES, DER, Paris, France LASER ANALYTICA, London, France 3 EFSA, Parma, Italy 2
Conventionally, a 3.16-fold default uncertainty factor has been used in risk assessment to cover human variability in toxicokinetics. The objective of this work is to show how data collection and meta-analysis can be used to inform human variability in tokicokinetic and refine uncertainty factors. This work focuses on CYP2D6 which a major human polymorphic metabolic pathway. Published pharmacokinetic studies in Caucasian and Asian extensive (EMs) intermediate (IMs) and poor metabolizers (PMs) for CYP2D6 have been analyzed using data primarily related to chronic exposure (oral clearance and area under the plasma concentration time curve) and acute exposure (peak concentration). A multi-level hierarchical Bayesian model has been proposed to meta-analyze these data integrating all quantifiable sources of variability, including inter-compound and inter-study variability. This work highlights large inter-individual variability in kinetic within the Caucasian EMs (coefficients of variation above 50%). A slightly weaker interindividual variability was found within Caucasian PM (coefficients of variation around 35%) and within Asian population (coefficients of variation below 40% for EMs and IMs). Comparisons between
S106
Abstracts / Toxicology Letters 238S (2015) S56–S383
Caucasian EMs and PMs and Asian EMs and IMs revealed an increase internal dose for PMs and IMs with a ratio compared to EMs that ranges from 1 to 34 and from 1 to 5. Uncertainty factors were calculated for each subgroup and for the two populations taking the prevalence of PMs and IMs into account through Monte-Carlo simulations. It results that none of the subgroup would be cover by the 3.16-fold default factor. Indeed, a 11-fold CYP2D6-related factors would be necessary to cover 95% of Caucasian and Asian populations. Moreover, an exponential relationship was found between the percentage of CYP2D6 metabolism in EMs and the uncertainty factors for PMs. Such relation combined with in vitro method could be used to establish chemical-specific uncertainty factors or as input to for kinetic models used in risk assessment. This work highlights how published data can be combined into a weight of evidence approach to better assess variability and uncertainty in human risk assessment. Indeed, pathway related uncertainty factors derived in this work could be proposed to replace the default uncertainty factor in case of single exposure. Such approach could also be extended to investigate human variability in case of exposure to chemical mixture. http://dx.doi.org/10.1016/j.toxlet.2015.08.346
P03-058 What are the best metabolites of gaseous polycyclic aromatic hydrocarbons to perform occupational biomonitoring? S. Lutier 1,∗ , D. Barbeau 1,2,∗ , R. Persoons 2 , M. Marques 1 , A. Maître 1,2 1 Université Grenoble Alpes, Laboratoire TIMC-IMAG, Equipe Environnement et Prédictions de la Santé des Populations, La Tronche, France 2 CHU de Grenoble, DBTP, IBP, Laboratoire de Toxicologie Professionnelle et Environnementale, Grenoble, France
Polycyclic aromatic hydrocarbons (PAHs) are ubiquitous carcinogenic pollutants emitted in complex mixtures as gaseous (gPAHs) and particulate (pPAHs) states. While urinary metabolites of pPAHs are classically monitored to estimate occupational exposure in industries using coal tar pitch, gPAHs biomonitoring could be useful for low exposure assessment. Question: The aim of this study was to determine the best biomarkers to perform gPAHs biomonitoring in electrometallurgy among the metabolites of naphthalene, fluorene and phenanthrene. Methods: Urine of five non-smoking electrometallurgy plant workers were collected at the pre- and post-shift of the last workday, then all urine were recovered during weekend. 1- and 2-hydroxynaphthalene (1- and 2-Naph), 2-, 3- and 9-hydroxyfluorene (2-,3- and 9-Flu), 1-, 2-, 3, 4- and 9-hydroxyphenanthrene (1-, 2-, 3-, 4- and 9-Phe) were analyzed using GC–MS. Occupational exposure was investigated by linear regression of log-transformed metabolites concentrations from the maximum excretion to the 24 h after. For biomarkers showing significant first order elimination for at least 4 workers, a non-linear mixed effects model not requiring the whole urinary volumes was developed to calculate their half-lives. Results: While 43% of 9-Flu and 85% of 4-Phe levels were below the limit of quantification, the concentrations of 1-, 2-Naph, 3-Flu, and 1-, 2-, 9-Phe were close to individual background. Occupational exposure was only be demonstrated for fluorene and phenanthrene thanks to 2-Flu and 3-Phe. Half-lives of 2-Flu and 3-Phe calculated by the non-linear mixed effects model were 6.7 h and 6.0 h, respectively. Conclusions: To perform gPAHs biomonitoring in electrometallurgy, urinary analysis of 2-Flu and 3-Phe seems to be the most relevant
biomarkers. However, the usefulness of these biomarkers must yet be assessed in other industrial sectors. Modelling approach allowed to calculate their urinary elimination half-lives which are close to those reported with more conventional approach. Grant from Region Rhône-Alpes and fund from ANSES. http://dx.doi.org/10.1016/j.toxlet.2015.08.347
P03-059 Climate change impact on the PAH ecotoxicity in Mediterranean soils J. Domingo ∗ , M. Marquès, M. Mari, M. Schuhmacher, J. Sierra, M. Nadal Universitat Rovira i Virgili, School of Medicine, Reus, Spain One of the consequences of climate change is its capacity to alter the occurrence and biological effects of chemical toxicants. As climate change is already altering temperature and solar radiation, among other parameters, an impact on the fate, transport and distribution pathways of semivolatile organic chemicals, such as polycyclic aromatic hydrocarbons (PAHs), is estimated. PAHs are important airborne pollutants which are deposited in soils. This study was aimed at assessing the ecotoxicological effects of temperature and solar radiation exposure in PAH-polluted soils and evaluating the potential generation of PAH metabolites. Ten grams of the A horizon of two typical Mediterranean soils were contaminated by using a mixture of the 16 US EPA priority PAHs. Soil samples were subjected to light radiation in a climate chamber under 2 different climate scenarios, according to IPCC projections: (1) a current Mediterranean (20 ◦ C and 9.6 W/m2 ), and (2) an RCP 8.5 scenario (24 ◦ C and 24 W/m2 ). Simultaneously, dark control samples, covered by aluminum foil, were concurrently irradiated. Ecotoxicity levels were daily controlled with Microtox® by measuring potential changes in the activity of the bioluminescent bacterium Vibrio fischeri in organic phases. Soil samples became more detoxified over time in the current climate scenario, showing a higher detoxification rate in fine-textured soil. Soil detoxification is likely to be dependent on soil texture and climate conditions. A close chromatographic analysis of the data revealed the presence of PAH metabolites, whose toxicity could be even higher than that of their parental compounds. http://dx.doi.org/10.1016/j.toxlet.2015.08.348
P03-060 Assessment of primary DNA damage in HepG2 cell line after acute exposure to alpha-cypermethrin and imidacloprid V. Kasuba ∗ , D. Zeljezic, M. Mladinic, N. Kopjar, R. Rozgaj Institute for Medical Research and Occupational Health, Mutagenesis Unit, Zagreb, Croatia Question: Genotoxic effect of alpha cypermethrin (type II pyrethroid insecticide) and Imidacloprid (neonicotinoid insecticide) were evaluated on human hepatoma HepG2 cell line. Concentrations used corresponded to real-life exposures will be evaluated based on residential exposure levels (REL␣ cypermethrin = 0.003643 g/mL; RELImidacloprid = 0.314 g/mL), occupational exposure limits (OEL) for alpha cypermethrin 0.262 g/mL, and AOEL (acceptable operator exposure level) for Imidacolprid,
S105
P03-055 Tobacco smoking induces specifically the expansion of a subpopulation of T cells expressing the homing and HIV co-receptor GPR15
P03-056 Novel analytical method to measure formaldehyde release from heated hair straightening cosmetic products: Impact on risk assessment
M. Bauer 1,∗ , G. Linsel 2 , K. Offenberg 1 , U. Sack 3 , H. Knaak 3 , G. Herberth 1
C.L. Galli 1,∗ , F. Bettin 2 , P. Metra 2 , P. Fidente 1 , M. Marinovich 1 , D. Dominicis 2
1
1
Helmholtz Centre for Environmental Research, Environmental Immunology, Leipzig, Germany 2 Federal Institute for Occupational Safety and Health, Berlin, Germany 3 Medical Faculty, University of Leipzig, Institute of Clinical Immunology, Leipzig, Germany Purpose: Tobacco smoking seriously raises mortality and morbidity worldwide. It increases risk for diseases in people living with HIV and worsens the severity of colon-restricted Crohn’s disease. Understanding the mechanisms through which smoking increases health risk to these disorders may establish new prevention or treatment strategies. Methods: Healthy adult subjects of two independent cohorts (Duck Operations (n = 93) and Blood Donor (n = 123) Cohorts) were implemented to study gene and protein expression in peripheral white blood cells by PCR and flow cytometry, respectively. Specimens of bronchoalveolar lavage were used to clarify smoking induced homing into lung. Results: Analyzing the lymphocyte homing and HIV co-receptor GPR15 on leucocytes in blood we found a high specific 3.6 fold (p < 0.0001) enrichment of GPR15-positive T cells in smokers compared to nonsmokers. The difference was independent on gender and age but reversal after cessation. In contrast to female smokers of the Duck Operations Cohort, the GPR15 gene expression in male followed an age-dependency (correlation coefficient r2 = 0.48, p < 0.00001). With regard to other known organ-specific homing receptors the GPR15 + T cells expressed partially CCR4 and CCR7 and to high extent the lymphnode homing CD62L. The assumption that GPR15 may serve additionally as a homing receptor for lymphocyte trafficking into human lung has not been confirmed analyzing GPR15 expression in leucocytes of bronchoalveolar lavage. Conclusion: For the first time we have illuminated a very sensitive and specific tobacco smoke induced enrichment of a specialized T cell type in blood as a result of an adaptive immune response toward tobacco smoke pollution of the human body. This finding may explain the tobacco-smoke promoted affection of the hereby mentioned diseases by helping HIV-virus to spread or by maintaining of inflammation in colon, into which this cell type specifically home, via tobacco smoking induced excess of GPR15+ cells in blood. http://dx.doi.org/10.1016/j.toxlet.2015.08.344
Università degli Studi di Milano, Dipartimento di Scienze Farmacologiche e Biomolecolari, Milano, Italy 2 R&D Department Chelab-Silliker a Mérieux Nutriscience Company, Resana, Italy In Europe, formaldehyde (FA) is permitted for use in personal care products at concentrations ≤ 0.2 g/100. Our method captures and collects the FA released into the air from heated cosmetic products by derivatization with 2,4-dinitrophenylhydrazine and analysis by HPLC/DAD instrument. 42% of 72 market samples analyzed showed FA concentrations very close to or above the threshold value (0.074 g/100 g calculated as FA), whereas 11 samples, negative using the official method of reference, were close to or above the threshold value. This may pose a health problem for occasional users and professional hair stylists. The values of FA equivalents released from the selected hair straightening products transformed into ppm (FA mg/1.23) extrapolated to the hypothesized volume of 1 m3 (FA mg/1.23/1 m3 ), for the exposure of the consumer and the operator, are mostly far above the limits established by DFG (0.3 ppm) and OSHA STEL (2 ppm, 15 min) for both stylists and customers receiving a 100 g (median 67.48 ppm) or a 35 g (median 23.62 ppm) treatment, respectively, even when the FA concentration in the product is below the 0.074% limit. http://dx.doi.org/10.1016/j.toxlet.2015.08.345
P03-057 Meta-analysis to better integrate human variability in toxicokinetic: CYP2D6-related uncertainty factors C. Béchaux 1,∗ , B. Amzal 2 , A. Crépet 1 , J.-L. Dorne 3 1
ANSES, DER, Paris, France LASER ANALYTICA, London, France 3 EFSA, Parma, Italy 2
Conventionally, a 3.16-fold default uncertainty factor has been used in risk assessment to cover human variability in toxicokinetics. The objective of this work is to show how data collection and meta-analysis can be used to inform human variability in tokicokinetic and refine uncertainty factors. This work focuses on CYP2D6 which a major human polymorphic metabolic pathway. Published pharmacokinetic studies in Caucasian and Asian extensive (EMs) intermediate (IMs) and poor metabolizers (PMs) for CYP2D6 have been analyzed using data primarily related to chronic exposure (oral clearance and area under the plasma concentration time curve) and acute exposure (peak concentration). A multi-level hierarchical Bayesian model has been proposed to meta-analyze these data integrating all quantifiable sources of variability, including inter-compound and inter-study variability. This work highlights large inter-individual variability in kinetic within the Caucasian EMs (coefficients of variation above 50%). A slightly weaker interindividual variability was found within Caucasian PM (coefficients of variation around 35%) and within Asian population (coefficients of variation below 40% for EMs and IMs). Comparisons between
S106
Abstracts / Toxicology Letters 238S (2015) S56–S383
Caucasian EMs and PMs and Asian EMs and IMs revealed an increase internal dose for PMs and IMs with a ratio compared to EMs that ranges from 1 to 34 and from 1 to 5. Uncertainty factors were calculated for each subgroup and for the two populations taking the prevalence of PMs and IMs into account through Monte-Carlo simulations. It results that none of the subgroup would be cover by the 3.16-fold default factor. Indeed, a 11-fold CYP2D6-related factors would be necessary to cover 95% of Caucasian and Asian populations. Moreover, an exponential relationship was found between the percentage of CYP2D6 metabolism in EMs and the uncertainty factors for PMs. Such relation combined with in vitro method could be used to establish chemical-specific uncertainty factors or as input to for kinetic models used in risk assessment. This work highlights how published data can be combined into a weight of evidence approach to better assess variability and uncertainty in human risk assessment. Indeed, pathway related uncertainty factors derived in this work could be proposed to replace the default uncertainty factor in case of single exposure. Such approach could also be extended to investigate human variability in case of exposure to chemical mixture. http://dx.doi.org/10.1016/j.toxlet.2015.08.346
P03-058 What are the best metabolites of gaseous polycyclic aromatic hydrocarbons to perform occupational biomonitoring? S. Lutier 1,∗ , D. Barbeau 1,2,∗ , R. Persoons 2 , M. Marques 1 , A. Maître 1,2 1 Université Grenoble Alpes, Laboratoire TIMC-IMAG, Equipe Environnement et Prédictions de la Santé des Populations, La Tronche, France 2 CHU de Grenoble, DBTP, IBP, Laboratoire de Toxicologie Professionnelle et Environnementale, Grenoble, France
Polycyclic aromatic hydrocarbons (PAHs) are ubiquitous carcinogenic pollutants emitted in complex mixtures as gaseous (gPAHs) and particulate (pPAHs) states. While urinary metabolites of pPAHs are classically monitored to estimate occupational exposure in industries using coal tar pitch, gPAHs biomonitoring could be useful for low exposure assessment. Question: The aim of this study was to determine the best biomarkers to perform gPAHs biomonitoring in electrometallurgy among the metabolites of naphthalene, fluorene and phenanthrene. Methods: Urine of five non-smoking electrometallurgy plant workers were collected at the pre- and post-shift of the last workday, then all urine were recovered during weekend. 1- and 2-hydroxynaphthalene (1- and 2-Naph), 2-, 3- and 9-hydroxyfluorene (2-,3- and 9-Flu), 1-, 2-, 3, 4- and 9-hydroxyphenanthrene (1-, 2-, 3-, 4- and 9-Phe) were analyzed using GC–MS. Occupational exposure was investigated by linear regression of log-transformed metabolites concentrations from the maximum excretion to the 24 h after. For biomarkers showing significant first order elimination for at least 4 workers, a non-linear mixed effects model not requiring the whole urinary volumes was developed to calculate their half-lives. Results: While 43% of 9-Flu and 85% of 4-Phe levels were below the limit of quantification, the concentrations of 1-, 2-Naph, 3-Flu, and 1-, 2-, 9-Phe were close to individual background. Occupational exposure was only be demonstrated for fluorene and phenanthrene thanks to 2-Flu and 3-Phe. Half-lives of 2-Flu and 3-Phe calculated by the non-linear mixed effects model were 6.7 h and 6.0 h, respectively. Conclusions: To perform gPAHs biomonitoring in electrometallurgy, urinary analysis of 2-Flu and 3-Phe seems to be the most relevant
biomarkers. However, the usefulness of these biomarkers must yet be assessed in other industrial sectors. Modelling approach allowed to calculate their urinary elimination half-lives which are close to those reported with more conventional approach. Grant from Region Rhône-Alpes and fund from ANSES. http://dx.doi.org/10.1016/j.toxlet.2015.08.347
P03-059 Climate change impact on the PAH ecotoxicity in Mediterranean soils J. Domingo ∗ , M. Marquès, M. Mari, M. Schuhmacher, J. Sierra, M. Nadal Universitat Rovira i Virgili, School of Medicine, Reus, Spain One of the consequences of climate change is its capacity to alter the occurrence and biological effects of chemical toxicants. As climate change is already altering temperature and solar radiation, among other parameters, an impact on the fate, transport and distribution pathways of semivolatile organic chemicals, such as polycyclic aromatic hydrocarbons (PAHs), is estimated. PAHs are important airborne pollutants which are deposited in soils. This study was aimed at assessing the ecotoxicological effects of temperature and solar radiation exposure in PAH-polluted soils and evaluating the potential generation of PAH metabolites. Ten grams of the A horizon of two typical Mediterranean soils were contaminated by using a mixture of the 16 US EPA priority PAHs. Soil samples were subjected to light radiation in a climate chamber under 2 different climate scenarios, according to IPCC projections: (1) a current Mediterranean (20 ◦ C and 9.6 W/m2 ), and (2) an RCP 8.5 scenario (24 ◦ C and 24 W/m2 ). Simultaneously, dark control samples, covered by aluminum foil, were concurrently irradiated. Ecotoxicity levels were daily controlled with Microtox® by measuring potential changes in the activity of the bioluminescent bacterium Vibrio fischeri in organic phases. Soil samples became more detoxified over time in the current climate scenario, showing a higher detoxification rate in fine-textured soil. Soil detoxification is likely to be dependent on soil texture and climate conditions. A close chromatographic analysis of the data revealed the presence of PAH metabolites, whose toxicity could be even higher than that of their parental compounds. http://dx.doi.org/10.1016/j.toxlet.2015.08.348
P03-060 Assessment of primary DNA damage in HepG2 cell line after acute exposure to alpha-cypermethrin and imidacloprid V. Kasuba ∗ , D. Zeljezic, M. Mladinic, N. Kopjar, R. Rozgaj Institute for Medical Research and Occupational Health, Mutagenesis Unit, Zagreb, Croatia Question: Genotoxic effect of alpha cypermethrin (type II pyrethroid insecticide) and Imidacloprid (neonicotinoid insecticide) were evaluated on human hepatoma HepG2 cell line. Concentrations used corresponded to real-life exposures will be evaluated based on residential exposure levels (REL␣ cypermethrin = 0.003643 g/mL; RELImidacloprid = 0.314 g/mL), occupational exposure limits (OEL) for alpha cypermethrin 0.262 g/mL, and AOEL (acceptable operator exposure level) for Imidacolprid,