Evaluation of methodologies for risk assessment of combined toxic actions of chemical substances and establishment of PBTK/TD models for pesticides

摘要

Humans are simultaneously exposed to a number of chemicals via food and environment.These chemicals may have a combined action that causes a lower or higher toxic effect thanwould be expected from knowledge about the single compounds. Therefore, combined actionsneed to be addressed in the risk assessment process.This Ph.D.-thesis provides an overview of the current knowledge on methods for riskassessment of combined actions of chemicals focussing on pesticides. Some of the methodsare based on knowledge on the whole mixture and others are based on data on the singlecompounds in the mixture. The whole mixture approaches would be the ideal choice forassessment of e.g. pesticide residues in food. However, they are normally not applicable sincethey require a large number of experimental data that are rarely available. This leaves thesingle compound approaches as the more realistic ones.The first step in the risk assessment of a mixture is to evaluate whether a group of compoundscan be identified that induce a common toxic effect by a common mechanism of toxicity andtherefore is suited for a cumulative risk assessment based on additivity. Ideally theidentification of a group of pesticides for cumulative risk assessment should be based oncriteria providing the best and most robust grouping such as chemical structure, mechanismof action, common toxic mode of action or common toxic effect. Unfortunately, such data areseldom available for all of the compounds of concern. Instead for pragmatic reasons, it is oftenmore appropriate to consider the individual compounds as possible candidates for one (ormore) cumulative assessment group(s).The cumulative risk assessment of this group will then be performed assuming simple similaraction using one of the single compound approaches. The hazard index based on a healthbased guidance value e.g. the acceptable daily intake (ADI) would normally be sufficient.However, the point of departure index is the most preferably method because it does notmake use of a policy driven uncertainty factor and instead it is based on the most relevanttoxicity data.In case that more than one common mechanism group based on different simple similaractions are identified, they should be assessed separately. In addition, the potential forinteractions between the groups (or single compounds) has to be considered. If nointeractions are identified, simple dissimilar action can be anticipated and the responseaddition method should be used to assess the effect of the mixture.In many cases the evaluators will probably tend to use very pragmatic approaches if lack ofinteraction between the compounds at the actual dose level can be assumed. This includesassuming all compounds in the mixture show dose additivity (simple similar actions). Thehazard index or point of departure index would then be the preferred methods.A crucial point in the assessment is whether there is interaction or no interaction between thecompounds in the mixture. Although interactions among chemicals at high doses are wellknown,no single simple approach is currently available to judge upon potential interactionsat the low dose levels of pesticide residues that humans are exposed to in food. For this purpose, physiologically based toxicokinetic/toxicodynamic (PBTK/TD) modelling has beenrecommended as a tool to assess combined tissue doses and to help predict potentialinteractions including thresholds for such effects. Therefore, this thesis also focuses on suchmodels and their applicability for use in risk assessment. This type of model has been used forseveral years in the area of pharmacology but the use in the area of toxicology is relativelynew.In a PBTK model the animal or man is described as a set of tissue compartments which iscombined by mathematical descriptions of biological tissues and physiological processes inthe body. Thereby it is possible to quantitatively simulate the absorption, distribution,metabolism and excretion of chemicals and to predict the internal dose after exposure to thechemical (or metabolite) of concern.The PBTK models make it possible to extrapolate between species, from high-dose to lowdose,from route-to-route and between exposure scenarios. In this way the risk assessor cansimulate various scenarios including scenarios which cannot be studied experimentally.Models can be developed for subpopulations such as children and this may help the riskassessor determine whether special care should be taken for such groups.It is also possible to incorporate mechanistic information on interactions in the model and asmentioned above interaction threshold can be determined. This would provide a helpful toolin the risk assessment of combined actions of chemicals.The PBTK model can be coupled with a toxicodynamic part in which the model attempts toestimate the effect resulting from the internal dose. The output of a PBTK model is linked to atoxicodynamic model by mathematical descriptions of the hypothesis of how compoundscontribute to the initiation of cellular changes leading to the toxic responses.In the present Ph.D. project a PBTK/TD model was established based on a previous publishedmodel. The model describes the organophosphorus pesticide chlorpyrifos and its metabolismto chlorpyrifos-oxon and 3,5,6-trichloro-2-pyridinol as well as the toxicodynamic of thechlorpyrifos-oxon i.e. inhibition of acetylcholinesterase activity in various tissues. This paperwas chosen because the model is on a relevant compound (a pesticide that is widely used) andone had the impression that the model work was described in details.The work in establishing this model clearly pointed out the importance that authors of suchpublications report their results with a high degree of transparency in order to enablecolleagues to reproduce their work and e.g. evaluate it for further developing the model. Themodel description should include model structure and equations as well as documentation ofthe choice of parameters and their origin. At present there is a lack of adequate data for use inthe PBTK models and further studies in order to determine parameters for use in PBTKmodels are needed. The model developer is also forced to make assumptions andextrapolations. It is of great importance that these are biologically based and explained.The PBTK/TD model on chlorpyrifos in the present project was used to illustrate how a noobserved adverse effect level (NOAEL) can be established by the model and how to makeextrapolations between species (rats and humans). The model underestimated the inhibition improvement before it can be used in risk assessment.The PBTD modelling is still in its infancy and it will probably be better to put more effort intoimproving the toxicokinetic part especially including establishment of internationallyacceptable reference values for various parameters before extending the model with atoxicodynamic part.PBTK models can be used to evaluate combined actions of a mixture of compounds. In case ofa mixture of compounds that do not interact (e.g. simple similar action) the PBTK modellingtool is useful to predict the combined doses in the target organ taking metabolism of thecompounds into account. Such compounds should be dealt with in the PBTK models in thesame way as single compounds.In case of a mixture of interacting compounds mechanistic information on interactions can beincorporated in the PBTK/TD model and thereby it can be used e.g. to determine theinteraction threshold. Such a model will consists of sets of identical equations, one set for eachchemical as well as equations that specifically accounts for the interactions (e.g. competitiveinhibition of metabolism in liver or induction of hepatic metabolism).The development of PBTK models is complex and should only be used when it is consideredessential. If adequate models are developed, they can provide better knowledge andunderstanding of the effects of mixtures in the organism and provide improved informationon tissue dose levels and variations between species and within a population. Moreover,scientifically supportable results about possible combined actions in humans after exposureto mixtures of pesticide residues in food would help making more reliable risk assessment.The PBTK models thus have a potential as an important tool in the risk assessment. Adequatedocumentation of the model is fundamental in order to increase the credibility of PBTKmodelling. Such credibility is crucial for a spreading of its use in risk assessment.This Ph.D.-project constitutes the initial work on implementing PBTK/TD models in the riskassessment of combined toxic action of chemical substances in food at the DTU National FoodInstitute. The work has revealed some major problems and pitfalls in the developing process.However if reliable, these models will provide knowledge of the relationship between internaldoses of the chemicals and the observed toxic effects and this knowledge will reduce theuncertainty in the risk assessment. Therefore, the work will continue implementing thesemodels as a helpful tool in future risk assessment.