Using available in vitro metabolite identification and time course kinetics for ß-chloroprene and its metabolite, (1-chloroethenyl) oxirane, to include reactive oxidative metabolites and glutathione depletion in a PBPK model for ß-chloroprene.

Introduction: ß-chloroprene (2-chloro-1,3-butadiene; CP) causes lung tumors after inhalation exposures in rats and mice. Mice develop these tumors at lower exposures than rats. In rats CP exposures cause depletion of lung glutathione (GSH). Methods: PBPK models developed to relate the appearance of mouse lung tumors with rates of CP metabolism to reactive metabolites or total amounts metabolized during exposures have been expanded to include production of reactive metabolites from CP. The extended PBPK model describes both the unstable oxirane metabolite, 2-CEO, and metabolism of the more stable oxirane, 1-CEO, to reactive metabolites via microsomal oxidation to a diepoxide, and linked production of these metabolites to a PK model predicting GSH depletion with increasing CP exposure. Key information required to develop the model were available from literature studies identifying: 1) microsomal metabolites of CP, and 2) in vitro rates of clearance of CP and 1-CEO from active microsomal preparations from mice, rats, hamsters and humans. Results: Model simulation of concentration dependence of disproportionate increases in reactive metabolite concentrations as exposures increases and decreases in tissue GSH are consistent with the dose-dependence of tumor formation. At the middle bioassay concentrations with a lung tumor incidence, the predicted tissue GSH is less than 50% background. These simulations of reduction in GSH are also consistent with the gene expression results showing the most sensitive pathways are Nrf2-regulation of oxidative stress and GSH metabolism. Discussion: The PBPK model is used to correlate predicted tissue exposure to reactive metabolites with toxicity and carcinogenicity of CP.

Objective ranges of soil-to-dust transfer coefficients for lead-impacted sites.

Residential yard soil and indoor dust datasets from eight communities near historical mining, smelting, and refining operations were used to quantify soil track-in, an important factor in evaluating indoor exposures to soil metals and to set residential soil cleanup levels. Regression analyses were used to derive slopes that represent mass soil-to-dust transfer coefficients or MSDs. Lead concentration data were available for all datasets. Arsenic data were available for six of the eight datasets. Cadmium and zinc data were available for one dataset, allowing limited comparison of MSDs for lead with other metals. Covariates that could indicate potential indoor sources of metals, such as house age and indoor heating source, were examined by multivariate regression analysis when available (three datasets). Covariates that could affect soil track-in, such as the amount of bare soil in the yard or having pets, were examined by stratified linear regression analysis when available (two datasets). Most of the R-squared values for lead, cadmium and zinc indicate a good to moderate fit (≥0.25), but for arsenic most indicate a poor fit (<0.25). Significant MSDs for models with a good to moderate fit range from 0.14 to 0.47 for lead, and 0.12 to 0.43 for the other metals (arsenic, cadmium, and zinc). The treatment of outliers was a significant methodological factor affecting the slope of the regressions. Substantial variability is expected among soils at residences due to both physical characteristics of each property and the ways in which residents interact with their home. Survey data providing information on various factors affecting soil track-in help to refine MSD estimates. For three of the datasets, covariate data were available that improved model fit by multivariate or stratified regression analysis for lead. When multivariate or stratified regression analyses were performed, the estimated MSD varied as little as <1% to as great as 200% depending on the dataset, but all estimates were below 0.4. Notably, the MSDs were lowest for the three datasets with the highest soil lead concentrations, i.e., those with average soil lead concentrations greater than 300 mg/kg after outlier removal. For five of the six datasets that had both arsenic and lead sampled, arsenic MSDs were much less than the lead MSDs; however, only two of the sites' arsenic models had significant MSDs and adequate fit. Cadmium and zinc were only included in one dataset, limiting our ability to draw any conclusions from comparison to those MSDs. The results of our study are consistent with prior studies suggesting that MSDs for metals without internal sources are 0.3-0.4, and application of MSDs in that range will provide more reliable exposure estimates than the 0.7 default value used by the United States Environmental Protection Agency in the Integrated Exposure Uptake Biokinetic (IEUBK) Model.

Assessing children's exposure to manganese in drinking water using a PBPK model.

A previously published human PBPK model for manganese (Mn) in infants and children has been updated with Mn in drinking water as an additional exposure source. Built upon the ability to capture differences in Mn source-specific regulation of intestinal uptake in nursing infants who are breast-fed and formula-fed, the updated model now describes the bioavailability of Mn from drinking water in children of ages 0-18. The age-related features, including the recommended age-specific Mn dietary intake, age-specific water consumption rates, and age-specific homeostasis of Mn, are based on the available human data and knowledge of the biology of essential-metal homeostasis. Model simulations suggest that the impact of adding drinking-water exposure to daily Mn exposure via dietary intake and ambient air inhalation in children is not greater than the impacts in adults, even at a drinking-water concentration that is 2 times higher than the USEPA's lifetime health advisory value. This conclusion was also valid for formula-fed infants who are considered at the highest potential exposure to Mn from drinking water compared to all other age groups. Our multi-route, multi-source Mn PBPK model for infants and children provides insights about the potential for Mn-related health effects on growing children and will thereby improve the level of confidence in properly interpreting Mn exposure-health effects relationships in children in human epidemiological studies.

A global human health risk assessment for octamethylcyclotetrasiloxane (D4).

Octamethylcyclotetrasiloxane (D4) is a low-molecular-weight volatile cyclic siloxane, primarily used as an intermediate in the production of some widely-used industrial and consumer silicone based polymers and may be present as a component in a variety of consumer products. A global "harmonized" risk assessment was conducted to meet requirements for substance-specific risk assessments conducted by regulatory agencies such as USEPA's Integrated Risk Information System (IRIS), Health Canada's Chemical Management Program (CMP) and various independent scientific committees of the European Commission (e.g. the Scientific Committee on Consumer Safety (SCCS), the Scientific Committee on Health and Environmental Risks (SCHER)), as well as to provide guidance for chemical safety assessments under REACH in Europe. This risk assessment incorporates global exposure information combined with a Monte Carlo analysis to determine the most significant routes of exposure. Utilization of a multi-species, multi-route physiologically based pharmacokinetic (PBPK) model was included to estimate internal dose metrics, benchmark modeling was used to determine a point of departure (POD), and a margin of safety (MOS) evaluation was used to compare the estimates of intake with the POD. Because of the specific pharmacokinetic behaviors of D4 including high lipophilicity, high volatility with low blood-to-air partition coefficients and an extensive metabolic clearance that regulates tissue dose after exposure, the use of a PBPK model was essential to provide a comparison of a dose metric that reflects these processes. The characterization of the potential for adverse effects after exposure to D4 using a MOS approach based on an internal dose metric removes the subjective application of varying uncertainty factors from various regulatory agencies and allows examination of the differences between internal dose metrics associated with exposure and those associated with adverse effects.

A constrained maximum likelihood approach to evaluate the impact of dose metric on cancer risk assessment: application to ß-chloroprene.

ß-Chloroprene (2-chloro-1,3-butadiene, CD) is used in the manufacture of polychloroprene rubber. Chronic inhalation studies have demonstrated that CD is carcinogenic in B6C3F1 mice and Fischer 344 rats. However, epidemiological studies do not provide compelling evidence for an increased risk of mortality from total cancers of the lung. Differences between the responses observed in animals and humans may be related to differences in toxicokinetics, the metabolism and detoxification of potentially active metabolites, as well as species differences in sensitivity. The purpose of this study was to develop and apply a novel method that combines the results from available physiologically based kinetic (PBK) models for chloroprene with a statistical maximum likelihood approach to test commonality of low-dose risk across species. This method allows for the combined evaluation of human and animal cancer study results to evaluate the difference between predicted risks using both external and internal dose metrics. The method applied to mouse and human CD data supports the hypothesis that a PBK-based metric reconciles the differences in mouse and human low-dose risk estimates and further suggests that, after PBK metric exposure adjustment, humans are equally or less sensitive than mice to low levels of CD exposure.

Acrylamide: review of toxicity data and dose-response analyses for cancer and noncancer effects.

Acrylamide (ACR) is used in the manufacture of polyacrylamides and has recently been shown to form when foods, typically containing certain nutrients, are cooked at normal cooking temperatures (e.g., frying, grilling or baking). The toxicity of ACR has been extensively investigated. The major findings of these studies indicate that ACR is neurotoxic in animals and humans, and it has been shown to be a reproductive toxicant in animal models and a rodent carcinogen. Several reviews of ACR toxicity have been conducted and ACR has been categorized as to its potential to be a human carcinogen in these reviews. Allowable levels based on the toxicity data concurrently available had been developed by the U.S. EPA. New data have been published since the U.S. EPA review in 1991. The purpose of this investigation was to review the toxicity data, identify any new relevant data, and select those data to be used in dose-response modeling. Proposed revised cancer and noncancer toxicity values were estimated using the newest U.S. EPA guidelines for cancer risk assessment and noncancer hazard assessment. Assessment of noncancer endpoints using benchmark models resulted in a reference dose (RfD) of 0.83 microg/kg/day based on reproductive effects, and 1.2 microg/kg/day based on neurotoxicity. Thyroid tumors in male and female rats were the only endpoint relevant to human health and were selected to estimate the point of departure (POD) using the multistage model. Because the mode of action of acrylamide in thyroid tumor formation is not known with certainty, both linear and nonlinear low-dose extrapolations were conducted under the assumption that glycidamide or ACR, respectively, were the active agent. Under the U.S. EPA guidelines (2005), when a chemical produces rodent tumors by a nonlinear or threshold mode of action, an RfD is calculated using the most relevant POD and application of uncertainty factors. The RfD was estimated to be 1.5 microg/kg/day based on the use of the area under the curve (AUC) for ACR hemoglobin adducts under the assumption that the parent, ACR, is the proximate carcinogen in rodents by a nonlinear mode of action. When the mode of action in assumed to be linear in the low-dose region, a risk-specific dose corresponding to a specified level of risk (e.g., 1 x 10-5) is estimated, and, in the case of ACR, was 9.5 x 10-2 microg ACR/kg/day based on the use of the AUC for glycidamide adduct data. However, it should be noted that although this review was intended to be comprehensive, it is not exhaustive, as new data are being published continuously.

Comparison of the EU T25 single point estimate method with benchmark dose response modeling for estimating potency of carcinogens.

The T25 single-point estimate method of evaluating the carcinogenic potency of a chemical, which is currently used by the European Union (EU) and is denoted the EU approach, is based on the selection of a single dose in a chronic bioassay with an incidence rate that is significantly higher than the background rate. The T25 is determined from that single point by a linear extrapolation or interpolation to the chronic dose (in mg/kg/day), at which a 25% increase in the incidence of the specified tumor type is expected, corrected for the background rate. Another method used to obtain a carcinogenic potency value based on a 25% increase in incidence above the background rate is the estimation of a T25 derived from a benchmark dose (BMD) response model fit to the chronic bioassay data for the specified tumor type. A comparison was made between these two methods using 276 chronic bioassays conducted by the National Toxicology Program. In each of the 2-year bioassays, a tumor type was selected based on statistical and biological significance, and both EU T25 and BMD T25 estimates were determined for that end point. In addition, simulations were done using underlying cumulative probability distributions to examine the effect of dose spacing, the number of animals per dose group, the possibility of a dose threshold, and variation in the background incidence rates on the EU T25 and BMD estimates. The simulations showed that in the majority of cases the EU T25 method underestimated the true T25 dose and overestimated the carcinogenic potency. The BMD estimate is generally less biased and has less variation about the true T25 value than the EU estimate.

Benchmark concentrations for methylmercury obtained from the Seychelles Child Development Study.

Methylmercury is a neurotoxin at high exposures, and the developing fetus is particularly susceptible. Because exposure to methylmercury is primarily through fish, concern has been expressed that the consumption of fish by pregnant women could adversely affect their fetuses. The reference dose for methylmercury established by the U.S. Environmental Protection Agency was based on a benchmark analysis of data from a poisoning episode in Iraq in which mothers consumed seed grain treated with methylmercury during pregnancy. However, exposures in this study were short term and at much higher levels than those that result from fish consumption. In contrast, the Agency for Toxic Substances and Disease Registry (ATSDR) based its proposed minimal risk level on a no-observed-adverse-effect level (NOAEL) derived from neurologic testing of children in the Seychelles Islands, where fish is an important dietary staple. Because no adverse effects from mercury were seen in the Seychelles study, the ATSDR considered the mean exposure in the study to be a NOAEL. However, a mean exposure may not be a good indicator of a no-effect exposure level. To provide an alternative basis for deriving an appropriate human exposure level from the Seychelles study, we conducted a benchmark analysis on these data. Our analysis included responses from batteries of neurologic tests applied to children at 6, 19, 29, and 66 months of age. We also analyzed developmental milestones (age first walked and first talked). We explored a number of dose-response models, sets of covariates to include in the models, and definitions of background response. Our analysis also involved modeling responses expressed as both continuous and quantal data. The most reliable analyses were considered to be represented by 144 calculated lower statistical bounds on the benchmark dose (BMDLs; the lower statistical bound on maternal mercury hair level corresponding to an increase of 0.1 in the probability of an adverse response) derived from the modeling of continuous responses. The average value of the BMDL in these 144 analyses was 25 ppm mercury in maternal hair, with a range of 19 to 30 ppm.

Determination of a site-specific reference dose for methylmercury for fish-eating populations.

Environmental risk-management decisions in the U.S. involving potential exposures to methylmercury currently use a reference dose (RfD) developed by the U.S. Environmental Protection Agency (USEPA). This RfD is based on retrospective studies of an acute poisoning incident in Iraq in which grain contaminated with a methylmercury fungicide was inadvertently used in the baking of bread. The exposures, which were relatively high but lasted only a few months, were associated with neurological effects in both adults (primarily paresthesia) and infants (late walking, late talking, etc.). It is generally believed that the developing fetus represents a particularly sensitive subpopulation for the neurological effects of methylmercury. The USEPA derived an RfD of 0.1 microg/kg/day based on benchmark dose (BMD) modeling of the combined neurological endpoints reported for children exposed in utero. This RfD included an uncertainty factor of 10 to consider human pharmacokinetic variability and database limitations (lack of data on multigeneration effects or possible long-term sequelae of perinatal exposure). Alcoa signed an Administrative Order of Consent for the conduct of a remedial investigation/feasibility study (RI/FS) at their Point Comfort Operations and the adjacent Lavaca Bay in Texas to address the effects of historical discharges of mercury-containing wastewater. In cooperation with the Texas Natural Resource Conservation Commission and USEPA Region VI, Alcoa conducted a baseline risk assessment to assess potential risk to human health and the environment. As a part of this assessment. Alcoa pursued the development of a site-specific RfD for methylmercury to specifically address the potential human health effects associated with the ingestion of contaminated finfish and shellfish from Lavaca Bay. Application of the published USEPA RfD to this site is problematic; while the study underlying the RfD represented acute exposure to relatively high concentrations of methylmercury, the exposures of concern for the Point Comfort site are from the chronic consumption of relatively low concentrations of methylmercury in fish. Since the publication of the USEPA RfD, several analyses of chronic exposure to methylmercury in fish-eating populations have been reported. The purpose of the analysis reported here was to evaluate the possibility of deriving an RfD for methylmercury, specifically for the case of fish ingestion, on the basis of these new studies. In order to better support the risk-management decisions associated with developing a remediation approach for the site in question, the analysis was designed to provide information on the distribution of acceptable ingestion rates across a population, which could reasonably be expected to be consistent with the results of the epidemiological studies of other fish-eating populations. Based on a review of the available literature on the effects of methylmercury, a study conducted with a population in the Seychelles Islands was selected as the critical study for this analysis. The exposures to methylmercury in this population result from chronic, multigenerational ingestion of contaminated fish. This prospective study was carefully conducted and analyzed, included a large cohort of mother-infant pairs, and was relatively free of confounding factors. The results of this study are essentially negative, and a no-observed-adverse-effect level (NOAEL) derived from the estimated exposures has recently been used by the Agency for Toxic Substances and Disease Registry (ATSDR) as the basis for a chronic oral minimal risk level (MRL) for methylmercury. In spite of the fact that no statistically significant effects were observed in this study, the data as reported are suitable for dose-response analysis using the BMD method. Evaluation of the BMD method used in this analysis, as well as in the current USEPA RfD, has demonstrated that the resulting 95% lower bound on the 10% benchmark dose (BMDL) represents a conservative estimate of the traditional NOAEL, and that it is superior to the use of "average" or "grouped" exposure estimates when dose-response information is available, as is the case for the Seychelles study. A more recent study in the Faroe Islands, which did report statistically significant associations between methylmercury exposure and neurological effects, could not be used for dose-response modeling due to inadequate reporting of the data and confounding from co-exposure to polychlorinated biphenyls (PCBs). BMD modeling over the wide range of neurological endpoints reported in the Seychelles study yielded a lowest BMDL for methylmercury in maternal hair of 21 ppm. This BMDL was then converted to an expected distribution of daily ingestion rates across a population using Monte Carlo analysis with a physiologically based pharmacokinetic (PBPK) model to evaluate the impact of interindividual variability. The resulting distribution of ingestion rates at the BMDL had a geometric mean of 1.60 microg/kg/day with a geometric standard deviation of 1.33; the 1st, 5th, and 10th percentiles of the distribution were 0.86, 1.04, and 1.15 microg/kg/day. In place of the use of an uncertainty factor of 3 for pharmacokinetic variability, as is done in the current RfD, one of these lower percentiles of the daily ingestion rate distribution provides a scientifically based, conservative basis for taking into consideration the impact of pharmacokinetic variability across the population. On the other hand, it was felt that an uncertainty factor of 3 for database limitations should be used in the current analysis. Although there can be high confidence in the benchmark-estimated NOAEL of 21 ppm in the Seychelles study, some results in the New Zealand and Faroe Islands studies could be construed to suggest the possibility of effects at maternal hair concentrations below 10 ppm. In addition, while concerns regarding the possibility of chronic sequelae are not supported by the available data, neither can they be absolutely ruled out. The use of an uncertainty factor of 3 is equivalent to using a NOAEL of 7 ppm in maternal hair, which provides additional protection against the possibility that effects could occur at lower concentrations in some populations. Based on the analysis described above, the distribution of acceptable daily ingestion rates (RfDs) recommended to serve as the basis for site-specific risk-management decisions at Alcoa's Point Comfort Operations ranges from approximately 0.3 to 1.1 microg/kg/day, with a population median (50th percentile) of 0.5 microg/kg/day. By analogy with USEPA guidelines for the use of percentiles in applications of distributions in exposure assessments, the 10th percentile provides a reasonably conservative measure. On this basis, a site-specific RfD of 0.4 microg/kg/day is recommended.

Estimates of the proportion of chemicals that were carcinogenic or anticarcinogenic in bioassays conducted by the National Toxicology Program.

Estimates were made of the proportion of chemicals that were carcinogenic, anticarcinogenic, or either in 397 long-term bioassays conducted by the National Toxicology Program (NTP). The estimates were obtained from the global pattern of p-values obtained from statistical tests applied to individual experiments. These tests accounted for multiple comparisons using a randomization procedure and were found to operate at the correct level of significance. Representative estimates of the proportion of carcinogens [with 90% confidence intervals (CI)] compared to the NTP estimates were as follows: male mice, 0.32 (CI, 0.19-0.44), NTP = 0.29; female mice, 0. 28 (CI, 0.15-0.41), NTP = 0.34; male rats, 0.35 (CI, 0.23-0.47), NTP = 0.36; female rats, 0.34 (CI, 0.21-0.46), NTP = 0.28; all sexes and species, 0.59 (CI, 0.49-0.69), NTP = 0.51. Representative estimates of the proportion of anticarcinogens were as follows: male mice, 0. 34; female mice, 0.27; male rats, 0.40; female rats, 0.44; all sexes and species, 0.66. Thus, there was as much or more evidence in this study for anticarcinogenesis as carcinogenesis. Even though the estimators used were negatively biased, it was estimated that 85% of the chemicals were either carcinogenic or anticarcinogenic at some site in some sex-species group. This suggests that most chemicals given at high enough doses will cause some sort of perturbation in tumor rates.

Estimates of the proportions of carcinogens and anticarcinogens in bioassays conducted by the U.S. National Toxicology Program. Application of a new meta-analytic approach.

A meta-analysis was performed in order to estimate the proportion of liver carcinogens, the proportion of chemicals carcinogenic at any site, and the corresponding proportion of anticarcinogens among chemicals tested in 397 long-term cancer bioassays conducted by the U.S. National Toxicology Program (NTP). Although the estimator used was negatively biased, the study provided persuasive evidence for a larger proportion of liver carcinogens (0.43, 90% CI: 0.35, 0.51) than was identified by the NTP (0.28). A larger proportion of chemicals carcinogenic at any site was also estimated (0.59, 90% CI: 0.49, 0.69) than was identified by the NTP (0.51), although this excess was not statistically significant. A larger proportion of anticarcinogens (0.66) was estimated than carcinogens (0.59). Despite the negative bias, it was estimated that 85% of the chemicals were either carcinogenic or anticarcinogenic at some site in some sex-species group. This suggests that most chemicals tested at high enough doses will cause some sort of perturbation in tumor rates.

Calculation of benchmark doses for reproductive and developmental toxicity observed after exposure to isopropanol.

Reproductive, including developmental, toxicity risk assessment has typically relied on estimation of toxicity criteria values derived from no-observed-adverse-effect levels (NOAELs). The benchmark dose (BMD) approach has been proposed as an alternative that avoids problems with NOAELs. In this analysis of the reproductive and developmental toxicity observed in a multigeneration study of rats exposed to isopropanol, the BMD approach has been applied to all effects exhibiting significant dose-response relationships. The BMD estimates were very consistent across models and across end points; they were within the range of doses (100 to 500 mg/kg/day) that has been suggested as being the NOAEL. The use of the BMD approach for analysis of isopropanol reproductive toxicity is shown to avoid the experiment-specific argument of whether a particular treatment has induced statistically significant differences, compared to controls, in favor of the estimation of experiment-independent doses corresponding to risk levels of interest. The consistency of the BMD estimates, with values of about 420 mg/kg/day, suggests that, for isopropanol, the available multigeneration study data may provide a suitable basis for considering safe exposure.