Risk assessment of parabens in a transcriptomics-based in vitro test.

Parabens have been used for decades as preservatives in food, drugs and cosmetics. The majority however, were banned in 2009 and 2014 leaving only methyl-, ethyl-, propyl-, and butyl-derivates available for subsequent use. Methyl- and propylparaben have been extensively tested in vivo, with no resulting evidence for developmental and reproductive toxicity (DART). In contrast, ethylparaben has not yet been tested for DART in animal experiments, and it is currently debated if additional animal studies are warranted. In order to perform a comparison of the four currently approved parabens, we used a previously established in vitro test based on human induced pluripotent stem cells (iPSC) that are exposed to test substances during their differentiation to neuroectodermal cells. EC50 values for cytotoxicity were 906 µM, 698 µM, 216 µM and 63 µM for methyl-, ethyl-, propyl- and butylparaben, respectively, demonstrating that cytotoxicity increases with increasing alkyl chain length. Genome-wide analysis demonstrated that FDR-adjusted significant gene expression changes occurred only at cytotoxic or close to cytotoxic concentrations, for example 1720 differentially expressed genes (DEG) at 1000 µM ethylparaben, 1 DEG at 316 µM, and no DEG at 100 µM or lower concentrations. The highest concentration of ethylparaben that did not induce any cytotoxicity nor DEG was 1670-fold above the highest concentration reported in biomonitoring studies (60 nM ethylparaben in cord blood). In conclusion, cytotoxicity and gene expression alterations of ethylparaben occurred at concentrations of approximately three orders of magnitude above human blood concentrations; moreover, the substance fitted well into a scenario where toxicity increases with the alkyl chain length, and gene expression changes only occur at cytotoxic or close to cytotoxic concentrations. Therefore, no evidence was obtained suggesting that in vivo DART with ethylparaben would lead to different results as the methyl- or propyl derivates.

Read-across and new approach methodologies applied in a 10-step framework for cosmetics safety assessment - A case study with parabens.

Parabens are esters of para-hydroxybenzoic acid that have been used as preservatives in many types of products for decades including agrochemicals, pharmaceuticals, food and cosmetics. This illustrative case study with propylparaben (PP) demonstrates a 10-step read-across (RAX) framework in practice. It aims at establishing a proof-of-concept for the value added by new approach methodologies (NAMs) in read-across (RAX) for use in a next-generation risk assessment (NGRA) in order to assess consumer safety after exposure to PP-containing cosmetics. In addition to structural and physico-chemical properties, in silico information, toxicogenomics, in vitro toxicodynamic, toxicokinetic data from PBK models, and bioactivity data are used to provide evidence of the chemical and biological similarity of PP and analogues and to establish potency trends for observed effects in vitro. The chemical category under consideration is short (C1-C4) linear chain n-alkyl parabens: methylparaben, ethylparaben, propylparaben and butylparaben. The goal of this case study is to illustrate how a practical framework for RAX can be used to fill a hypothetical data gap for reproductive toxicity of the target chemical PP.

The Grouping and Assessment Strategy for Organic Pigments (GRAPE): Scientific evidence to facilitate regulatory decision-making.

This article presents the Grouping and Assessment Strategy for Organic Pigments (GRAPE). GRAPE is driven by the hypotheses that low (bio)dissolution and low permeability indicate absence of systemic bioavailability and hence no systemic toxicity potential upon oral exposure, and, for inhalation exposure, that low (bio)dissolution (and absence of surface reactivity, dispersibility and in vitro effects) indicate that the organic pigment is a 'poorly soluble particle without intrinsic toxicity potential'. In GRAPE Tier 1, (bio)solubility and (bio)dissolution are assessed, and in Tier 2, in vitro Caco-2 permeability and in vitro alveolar macrophage activation. Thereafter, organic pigments are grouped by common properties (further considering structural similarity depending on the regulatory requirements). In Tier 3, absence of systemic bioavailability is verified by limited in vivo screening (rat 28-day oral and 5-day inhalation toxicity studies). If Tier 3 confirms no (or only very low) systemic bioavailability, all higher-tier endpoint-specific animal testing is scientifically not-relevant. Application of the GRAPE can serve to reduce animal testing needs for all but few representative organic pigments within a group. GRAPE stands in line with the EU REACH Regulation (Registration, Evaluation, Authorisation and Restriction of Chemicals). An ongoing research project aims at establishing a proof-of-concept of the GRAPE.