Scientific Opinion of the Scientific Panel on Plant Protection Products and their Residues (PPR Panel) on testing and interpretation of comparative in vitro metabolism studies

Abstract

EFSA asked the Panel on Plant Protection Products and their residues to deliver a Scientific Opinion on testing and interpretation of comparative in vitro metabolism studies for both new active substances and existing ones. The main aim of comparative in vitro metabolism studies of pesticide active substances is to evaluate whether all significant metabolites formed in the human in vitro test system, as a surrogate of the in vivo situation, are also present at comparable level in animal species tested in toxicological studies and, therefore, if their potential toxicity has been appropriately covered by animal studies. The studies may also help to decide which animal model, with regard to a particular compound, is the most relevant for humans. In the experimental strategy, primary hepatocytes in suspension or culture are recommended since hepatocytes are considered the most representative in vitro system for prediction of in vivo metabolites. The experimental design of 3 9 3 9 3 (concentrations, time points, technical replicates, on pooled hepatocytes) will maximise the chance to identify unique (UHM) and disproportionate (DHM) human metabolites. When DHM and UHM are being assessed, test item-related radioactivity recovery and metabolite profile are the most important parameters. Subsequently, structural characterisation of the assigned metabolites is performed with appropriate analytical techniques. In toxicological assessment of metabolites, the uncertainty factor approach is the first alternative to testing option, followed by new approach methodologies (QSAR, read-across, in vitro methods), and only if these fail, in vivo animal toxicity studies may be performed. Knowledge of in vitro metabolites in human and animal hepatocytes would enable toxicological evaluation of all metabolites of concern, and, furthermore, add useful pieces of information for detection and evaluation of metabolites in different matrices (crops, livestock, environment), improve biomonitoring efforts via better toxicokinetic understanding, and ultimately, develop regulatory schemes employing physiologically based or physiology-mimicking in silico and/or in vitro test systems to anticipate the exposure of humans to potentially hazardous substances in plant protection products.

Risk assessment for Birds and Mammals

“…Comparative in vitro metabolism studies have been developed using specific mammalian cell types, e.g. hepatocytes (EFSA PPR Panel Opinion, 2021b). Similarly, OECD guidance on the determination of clearance of chemicals in rainbow trout cell lines is also available (OECD, 2018a).…”

Candidate Proficiency Test Chemicals to Address Industrial Chemical Applicability Domains for in vitro Human Cytochrome P450 Enzyme Induction

“…Moreover, for many chemicals, there are significant differences in metabolism between rats and humans, and only rat S9 microsomes are used to produce metabolic transformation in the OECD genotoxicity TGs. In a recent European Food Safety Authority (EFSA) scientific panel opinion paper relating to the toxicological testing of pesticides, the use of in vitro human metabolite data is recommended to identify any potential human relevant metabolites that had not been adequately tested in non-human toxicological studies ( EFSA 2021 ).…”

Setting of import tolerance for tricyclazole in rice

“…The comparative in vitro metabolism study using liver microsomes from F344/DuCrl Rats, Beagle Dogs and Human indicated no significant differences between animal laboratory species and human liver microsomes. EFSA noted that the study does not follow the recent recommendations of the EFSA Panel on Plant Protection Products and their Residues (PPR) Opinion on comparative in vitro metabolism studies (EFSA PPR, 2021 ).…”