Utility of a next‐generation framework for assessment of genomic damage: A case study using the pharmaceutical drug candidate etoposide

We present a hypothetical case study to examine the use of a next-generation framework developed by the Genetic Toxicology Technical Committee of the Health and Environmental Sciences Institute for assessing the potential risk of genetic damage from a pharmaceutical perspective. We used etoposide, a genotoxic carcinogen, as a representative pharmaceutical for the purposes of this case study. Using the framework as guidance, we formulated a hypothetical scenario for the use of etoposide to illustrate the application of the framework to pharmaceuticals. We collected available data on etoposide considered relevant for assessment of genetic toxicity risk. From the data collected, we conducted a quantitative analysis to estimate margins of exposure (MOEs) to characterize the risk of genetic damage that could be used for decision-making regarding the predefined hypothetical use. We found the framework useful for guiding the selection of appropriate tests and selecting relevant endpoints that reflected the potential for genetic damage in patients. The risk characterization, presented as MOEs, allows decision makers to discern how much benefit is critical to balance any adverse effect(s) that may be induced by the pharmaceutical. Interestingly, pharmaceutical development already incorporates several aspects of the framework per regulations and health authority expectations. Moreover, we observed that quality dose response data can be obtained with carefully planned but routinely conducted genetic toxicity testing. This case study demonstrates the utility of the next-generation framework to quantitatively model human risk based on genetic damage, as applicable to pharmaceuticals

Use of in vitro 3D tissue models in genotoxicity testing: Strategic fit, validation status and way forward. Report of the working group from the 7th International Workshop on Genotoxicity Testing (IWGT)

Use of three-dimensional (3D) tissue equivalents in toxicology has been increasing over the last decade as novel preclinical test systems and as alternatives to animal testing. In the area of genetic toxicology, progress has been made with establishing robust protocols for skin, airway (lung) and liver tissue equivalents. In light of these advancements, a “Use of 3D Tissues in Genotoxicity Testing” working group (WG) met at the 7th IWGT meeting in Tokyo in November 2017 to discuss progress with these models and how they may fit into a genotoxicity testing strategy. The workshop demonstrated that skin models have reached an advanced state of validation following over 10 years of development, while liver and airway model-based genotoxicity assays show promise but are at an early stage of development. Further effort in liver and airway model-based assays is needed to address the lack of coverage of the three main endpoints of genotoxicity (mutagenicity, clastogenicity and aneugenicity), and information on metabolic competence. The IWGT WG believes that the 3D skin comet and micronucleus assays are now sufficiently validated to undergo an independent peer review of the validation study, followed by development of individual OECD Test Guidelines

Étude des mécanismes et cibles cellulaires de la phototoxicité des fluoroquinolones (l'approche microspectrofluorimétrique sur cellules vivantes)

Les fluoroquinolones sont des antibiotiques très utilisés mais phototoxiques. Pour élucider les mécanismes de cette phototoxicité, nous avons mené une étude microspectrofluorimétrique de leur localisation intracellulaire dans des fibroblastes humains en culture, afin d'identifier les sites cellulaires de cette photosensibilisation, ainsi que les dommages causés au niveau de ces sites. Les expériences de co-localisation réalisées avec le jaune de Lucifer ou le rouge neutre (des marqueurs lysosomiaux fluorescents), ont permis d'identifier les lysosomes comme site préférentiel de localisation des fluoroquinolones, bien qu'on les retrouve dans tout le cytoplasme. Une altération de la perméabilité de la membrane lysosomiale a été démontrée en suivant la re-localisation du jaune de Lucifer ou du rouge neutre dans des fibroblastes photosensibilisés par ces antibiotiques. Des dommages ont aussi été détectés au niveau des mitochondries en utilisant la rhodamine 123 comme sonde fluorescente, bien que l'on n'observe pas de localisation très importante des fluoroquinolones dans ces organites. La perturbation des mécanismes de l'endocytose dans les fibroblastes traités avec les fluoroquinolones a été démontrée en utilisant les LDL comme ligand modèle. La fixation et l'internalisation plus la dégradation des LDL furent suivies au moyen du phénomène de F.R.E.T. (transfert d'énergie de fluorescence de type Forster), grâce à deux carbocyanines incorporées dans le cœur lipidique des LDL et formant un couple accepteur-donneur. Une dépolymerisation des filaments d'actine marqués au alexa 568 phalloidine a aussi été révélée par microscopie de fluorescence. La phototoxicité des fluoroquinolones induite par l'UVA, résulte donc de l'altération de cibles biologiques multiplesPARIS-Museum Hist.Naturelle (751052304) / SudocSudocFranceF

Application of ToxCast and ToxRefDB to Develop a Quantitative Model of Systemic Toxicity

Application of ToxCast and ToxRefDB to Develop a Quantitative Model of Systemic Toxicit

Quantitative Model of Systemic Toxicity Using ToxCast and ToxRef DB

Presented at the Annual Society of Toxicolog

Framework for a Quantitative Systemic Toxicity Model

Framework for a Quantitative Systemic Toxicity Mode

Ab Initio safety assessment case study : Integration of alternative methods in an applied risk assessment for repeated-dose toxicity

The SEURAT-1 ("Safety Evaluation Ultimately Replacing Animal Testing") Research Initiative aimed at finding alternative approaches for the safety assessment for repeated-dose toxicity (Gocht et al 2013). Within the SEURAT-1 framework of proof-of-concept case studies for applied safety assessment, the ab initio case study set out to identify and subsequently quantify relevant biological pathway concentration effect levels of a cosmetic ingredient for specific exposure scenarios. Based on the SEURAT-1 conceptual framework for safety assessment (White, Knight 2014), a general workflow was developed in an attempt to structuring knowledge and data in a logical sequence for an integrated safety assessment relying specifically on alternative methods. Considerations included the possible application of the Threshold of Toxicological Concern (TTC) approach and read-across assessment. Physiologically-based pharmacokinetic modelling was applied to identify target organs and internal concentrations. In silico structural alerts and QSAR profilers as well as in vitro information including high throughput assays were used to build a weight of evidence, based on an AOP-anchored mode-of-action hypothesis and supported by in vitro to in vivo extrapolation (IVIVE) modelling and refinement, with the aim of concluding on the safety of use regarding repeated-dose toxicity, focusing on liver toxicity. Piperonyl butoxide (PBO) was selected to illustrate the case study in fictional exposure scenarios as a new ingredient introduced in a shampoo and a daily applied body lotion. The supportive alternative data (in vitro and in silico) were generated in the SEURAT-1 projects or obtained from ToxCast. The case study focusing on alternative approaches highlights the challenge in integrating multiple data streams for safety assessment decisions, points out knowledge gaps and proposes a way forward

Integration of alternative methods for an ab initio chemical safety assessment

Assessing chemical safety using only non-animal methods is a major challenge within the European Union, especially considering the ban to market cosmetics with ingredients tested on animals (Regulation 1223/2009). One of the outcomes of the SEURAT-1 initiative (http://www.seurat-1.eu/), co-sponsored by the European Commission 7thFP and Cosmetics Europe, was to develop case studies addressing that issue. Besides a read-across case-study, where "new approach" data was used to strengthen the confidence in reading across data from an already assessed chemical to a structural analogue, an ab initio case study was carried out and is presented here. The ab initio case study was an attempt to structure knowledge and data in a logic workflow which could be used for decision making to predict whether the intended exposure could be considered safe based on data solely from alternative methods. The workflow is includes the following steps: i) identification of the exposure/use scenario; ii) data collection on the chemical; iii) toxicokinetic and toxicodynamic modeling; iv) evaluation of other alternative methods (in vitro or in silico) which are available and could provide evidence for a hypothesis on the chemicals' mode of action; v) confirmation of the hypothesis with targeted testing using selected in vitro or in silico methods; vi) propose a risk valuation, as well as estimating uncertainties and identifying data gaps. The exposure scenario is an initial and essential step in this workflow. Whether exposure is intentional or not it should be considered, and in both cases estimates of the doses, expected routes of exposure, frequency and length of exposure should be made. The ab initio workflow was applied to the piperonyl butoxide (PBO), as a hypothetical case, for which the mode-of-action was assumed to be unknown. The only data on the compound was the molecular structure and a concentration of PBO in a consumer product formulation. An exposure scenario was set up and data were generated for simulation of internal exposure for a “healthy human” using PBPK modeling. The concentration in blood and liver predicted by the model were then used to design in vitro tests (performed by SEURAT-1 partners). The test results were then evaluated and used to estimate whether PBO would be safe to use for the selected exposure scenario. To our knowledge, the SEURAT-1 ab initio case study is a first attempt to shift our minds to a fully non-animal based chemical risk assessment relying only on alternative methods

Deriving Points of Departure and Performance Baselines for Predictive Modeling of Systemic Toxicity using ToxRefDB

Presented at the Annual Society of Toxicology meetin