The EU-ToxRisk method documentation, data processing and chemical testing pipeline for the regulatory use of new approach methods

Abstract

Hazard assessment, based on new approach methods (NAM), requires the use of batteries of assays, where individual tests may be contributed by different laboratories. A unified strategy for such collaborative testing is presented. It details all procedures required to allow test information to be usable for integrated hazard assessment, strategic project decisions and/or for regulatory purposes. The EU-ToxRisk project developed a strategy to provide regulatorily valid data, and exemplified this using a panel of > 20 assays (with > 50 individual endpoints), each exposed to 19 well-known test compounds (e.g. rotenone, colchicine, mercury, paracetamol, rifampicine, paraquat, taxol). Examples of strategy implementation are provided for all aspects required to ensure data validity: (i) documentation of test methods in a publicly accessible database; (ii) deposition of standard operating procedures (SOP) at the European Union DB-ALM repository; (iii) test readiness scoring accoding to defined criteria; (iv) disclosure of the pipeline for data processing; (v) link of uncertainty measures and metadata to the data; (vi) definition of test chemicals, their handling and their behavior in test media; (vii) specification of the test purpose and overall evaluation plans. Moreover, data generation was exemplified by providing results from 25 reporter assays. A complete evaluation of the entire test battery will be described elsewhere. A major learning from the retrospective analysis of this large testing project was the need for thorough definitions of the above strategy aspects, ideally in form of a study pre-registration, to allow adequate interpretation of the data and to ensure overall scientific/toxicological validity.

Establishment of an a priori protocol for the implementation and interpretation of an in‐vitro testing battery for the assessment of developmental neurotoxicity

“…We did this for all media used in this study (assuming a drug concentration of 1 µM), and the difference of nPoD and fPoD was usually < 3%, never larger than 14% (UKN5 protein-rich medium with strongly protein binding test compound). We conclude from this exercise, that using nPoD (instead of the calculated fPoD) is a reasonable and sufficiently exact solution (Krebs et al, 2020b).…”

“…Increasing gene expression of typical dopaminergic neurodevelopmental markers like DAT, DRD2 and TH demonstrate the dopaminergic phenotype of the LUHMES cells (Scholz et al, 2011). Protein and lipid compositions of media used for NCC, iDRG and LUHMES test systems are given in (Krebs et al, 2020b). More details on the test systems relevant for the test methods can be found below.…”

“…Therefore, we modified OECD GD211 to adapt it to the common needs of test developers and regulators. We developed an annotated toxicity test method template (ToxTemp) (i) to fulfil all requirements of GD211, (ii) to guide the user concerning the types of answers and detail of information required, (iii) to include acceptance criteria for test elements, and (iv) to define the cells sufficiently and transparently Krebs et al (2020b) . As basis for a scoring system, a template was developed (ToxTemp, ).…”

“…Approximation formula for estimating free drug concentrations in vitro (Krebs et al, 2020b;Fisher et al 2019).…”

Setting the stage for next-generation risk assessment with non-animal approaches: the EU-ToxRisk project experience

“…In Vitro Toxicology and Biomedicine, Department Inaugurated By the Doerenkamp-Zbinden Foundation at the University of Konstanz, University of Konstanz, 78457 Constance, Germany Introduction A particular focus of EU-ToxRisk is on endpoints for repeated-dose (RDT) and developmental and reproductive toxicity (DART), and on providing guidance for practical implementation (Daneshian et al 2016). This entails the implementation of NAM-based safety science into regulatory toxicology (Krebs et al 2020b). The project initially focused on read-across (RAx) (Escher et al 2019;Rovida et al 2020a) and here we outline key outcomes and general learnings on the use of NAMs for RAx.…”