A framework to characterize uncertainty for dose extrapolation from in vitro bioassays

New approach methods (NAMs) can accelerate chemical assessment and reduce reliance on animal testing, but their application requires quantitative extrapolation of bioactive in vitro concentrations to equivalent in vivo doses. Confidence in quantitative in vitro-in vivo extrapolation (Q-IVIVE) is impacted by uncertainty in both the in vitro concentration and the toxicokinetic parameters used for reverse dosimetry. We propose a framework to: (1) define the processes used to derive an administered equivalent dose (AED); (2) identify uncertainty sources; (3) characterize uncertainty in the data and models; and (4) propagate these uncertainties to the AED. The framework distinguishes nominal assay concentrations from blood- or plasma-equivalent concentrations that account for chemical disposition and compositional differences between in vitro and in vivo media. Parameter-specific uncertainty factors can be derived from experimental variability, model performance statistics, simulations, or expert judgement and combined using a semi-probabilistic approach. Illustrative chemical-space analyses show that nominal and blood-equivalent concentrations may be comparable for some hydrophilic, non-volatile chemicals but can differ by more than two orders of magnitude for hydrophobic or volatile chemicals. The framework provides a systematic, and tiered basis for evaluating confidence in calculated Q-IVIVE doses and supports the application of NAM data in chemical hazard and risk assessment. Ongoing chemical-specific case studies will evaluate implementation of the framework, verify the validity of the simplifying assumptions, and identify opportunities for refinement.