Authors G. Eisenbrand, B. Pool-Zobel, V. Baker, M. Balls, B.J. Blaauboer, A. Boobis, A. Carere, S. Kevekordes, J.-C. Lhuguenot, R. Pieters, J. Kleiner
Abstract
In vitro methods are common and widely used for screening and ranking chemicals, and have also been taken into account sporadically for risk assessment purposes in the case of food additives. However, the range of food-associated compounds amenable to in vitro toxicology is considered much broader, comprising not only natural ingredients, including those from food preparation, but also compounds formed endogenously after exposure, permissible/authorised chemicals including additives, residues, supplements, chemicals from processing and packaging and contaminants. A major promise of in vitro systems is to obtain mechanism-derived information that is considered pivotal for adequate risk assessment. This paper critically reviews the entire process of risk assessment by in vitro toxicology, encompassing ongoing and future developments, with major emphasis on cytotoxicity, cellular responses, toxicokinetics, modelling, metabolism, cancer-related endpoints, developmental toxicity, prediction of allergenicity, and finally, development and application of biomarkers.
It describes in depth the use of in vitro methods in strategies for characterising and predicting hazards to the human. Major weaknesses and strengths of these assay systems are addressed, together with some key issues concerning major research priorities to improve hazard identification and characterisation of food-associated chemicals. Research Needs It is anticipated that in the future more and more emphasis will be placed on in vitro assays to study toxicity. For this it will be necessary to develop, improve, evaluate and validate systems specifically for studying effects of food associated chemicals. This applies especially to:
- New endpoints of toxicity
a.Identification of molecular markers based on detecting effects at levels of exposure lower than those, which cause pathological response
b. Identification of markers of different stages of the neoplastic process in the target cells of somatic tissues in which (food associated) tumours arise, including genotoxic effects / mutations in relevant tumour associated genes or in DNA repair genes as well as intermediated endpoints (i.e. proliferation, apoptosis, differentiation)
c. establishment of specific toxicity profiles in order to predict acute, subacute and chronic toxicity, including the effects of repeated, low dose exposure
d. establishment of (batteries of) in vitro systems for endpoint that are relevant for target organ or tissue-specific effects (i.e. by measuring changes in functional parameters of cells and tissues or by using transcriptomics/proteomics).
e. use of such information to develop reliable biomarkers of effect or toxicological endpoints for use in the toxicological evaluation of food chemicals
f. Specific target effects should also include effects on developmental processes
g. Development and evaluation of methods to study allergenicity of food components
i. potential effects by low weight molecular compounds, i.e.. on the mucosal immune system of the gastrointestinal tract.
ii. new screening essays for high molecular weight compounds and elucidation of mechanism i.e. interaction with mucosal immune system
iii. development of ex vivo tests to study (peptide specific) memory T cell responses
2. Toxicokinetics
a. in vitro systems for evaluating the biotransformation of food compounds
b. reliable models of barrier functions (i.e. gastrointestinal tract, blood/brain, placenta ) including transporter-functions involved in absorption and excretion of compounds
c. models to study food matrix effects on the absorption of chemicals
3. Human-based systems of increasing hierarchy, such as
a. subcellular systems,
b. cellular systems:primary cells from somatic tissues such as non-transformed cells and cells from preneoplastic foci, tumour cells, embryonic cells, genetically modified cells
c. whole tissues (i.e. surgical samples),
4. Structured approaches for prediction:
a. Integrated strategies combining
i. Prior knowledge on structural alerts
ii. In vitro assays addressing toxicokinetics
b. Parallelogram approach based on comparing in vitro assays with animal and human cells, supported by
i. human exposure data
ii. biomarker-based analysis in animals and humans
5. Reference libraries and in-silico systems containing information on
a. structure, structural alerts for toxic effects, chemical functionalitites and biotransformation;
b. prior knowledge on toxicity data, i.e. on the profile of toxic activities with relevance for food
c. known susceptibility factors and their influence on the impact of risks.
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