Incorporating human dosimetry and exposure into high-throughput in vitro toxicity screening

Abstract

Many chemicals in commerce today have undergone limited or no safety testing. To reduce the number of untested chemicals and prioritize limited testing resources, several governmental programs are using high-throughput in vitro screens for assessing chemical effects across multiple cellular pathways. In this study, metabolic clearance and plasma protein binding were experimentally measured for 35 ToxCast phase I chemicals. The experimental data were used to parameterize a population-based in vitro-to-in vivo extrapolation model for estimating the human oral equivalent dose necessary to produce a steady-state in vivo concentration equivalent to in vitro AC(50) (concentration at 50% of maximum activity) and LEC (lowest effective concentration) values from the ToxCast data. For 23 of the 35 chemicals, the range of oral equivalent doses for up to 398 ToxCast assays was compared with chronic aggregate human oral exposure estimates in order to assess whether significant in vitro bioactivity occurred within the range of maximum expected human oral exposure. Only 2 of the 35 chemicals, triclosan and pyrithiobac-sodium, had overlapping oral equivalent doses and estimated human oral exposures. Ranking by the potencies of the AC(50) and LEC values, these two chemicals would not have been at the top of a prioritization list. Integrating both dosimetry and human exposure information with the high-throughput toxicity screening efforts provides a better basis for making informed decisions on chemical testing priorities and regulatory attention. Importantly, these tools are necessary to move beyond hazard rankings to estimates of possible in vivo responses based on in vitro screens.

FIG. 1.

Schematic representation for incorporating human dosimetry and exposure information into the high-throughput in vitro toxicity screening process performed as a part of ToxCast.

FIG. 2.

Distribution of (A) plasma protein binding measurements (averaged for both 1 and 10μM) and (B) metabolic clearance measurements (includes both the 1 and the 10μM concentrations) for the 35 ToxCast chemicals analyzed.

FIG. 3.

Comparison of oral equivalent doses and highest estimated human oral exposure levels for the 35 ToxCast chemicals analyzed. The oral equivalent doses (milligram per kilogram per day) for each chemical were estimated for each of the 398 ToxCast assays that possessed a measurable AC₅₀ value using in vitro-to-in vivo extrapolation modeling. In the modeling analysis, Monte Carlo simulation was performed to simulate variability across a cohort of 100 healthy individuals of both sexes from 20 to 50 years of age. From the Monte Carlo simulations, the lower fifth percentile of oral equivalent doses was selected as a conservative estimate for a population. The distribution of the oral equivalent doses is depicted as a box plot showing the median, upper, and lower 95% confidence intervals, with circles indicating data points outside the 95% range. Human exposure estimates for 23 chemicals were obtained from reregistration eligibility documents and NHANES biomonitoring data and reflect the most highly exposed group or subpopulation (green squares). Exposures for dicrotophos, fenamiphos, and methyl parathion were below 1 × 10⁻⁵ mg/kg/day and are therefore not shown on the graph. Chemicals where the highest estimated human oral exposure values fall within the range of predicted oral equivalents are highlighted with arrows.

FIG. 4.

Box plots of AC₅₀ values across the 398 ToxCast assays for the 35 ToxCast chemicals analyzed. The median, upper, and lower 95% confidence intervals with circles indicating points outside the 95% span are indicated as in Figure 3. The chemicals are presented in the same order as Figure 3 for comparison purposes.