Front Toxicol. 2025 Dec 17;7:1690491. doi: 10.3389/ftox.2025.1690491. eCollection 2025.
ABSTRACT
Read-across has matured from an expert-driven extrapolation based largely on structural analogy into a rigorously documented, mechanistically informed cornerstone of next-generation risk assessment. Three pivotal frameworks are compared that now shape its regulatory use: the European Food Safety Authority's (EFSA) 2025 guidance for food and feed safety, the European Chemicals Agency's (ECHA) Read-Across Assessment Framework (RAAF) for industrial chemicals under REACH, and the community-driven Good Read-Across Practice (GRAP) principles. Using five analytical lenses-conceptual structure, scientific rigor, implementation tools, regulatory acceptance, and practical impact-we identified areas of complementarity and divergence. EFSA provides a seven-step, uncertainty-anchored workflow that actively embeds new approach methodologies (NAMs) and adverse outcome pathway reasoning, offering applicants a transparent "how-to" template. RAAF, in contrast, operates as an evaluator's rubric: six scenario types and associated assessment elements delineate what evidence must be delivered, thereby standardizing regulatory scrutiny but leaving dossier construction to the registrant. GRAP supplies the conceptual glue, emphasizing mechanistic plausibility, exhaustive analogue selection, explicit uncertainty characterization, and the strategic use of NAMs; its influence is evident in both EFSA's and ECHA's evolving expectations. (Terminology note: the acronym "NAM" was popularized at an ECHA workshop in 2016; earlier documents such as RAAF and initial GRAP papers therefore may not use the term explicitly). Regulatory experience under REACH demonstrates that dossier quality and acceptance rates rise markedly when RAAF criteria are met, while EFSA's new guidance is poised to catalyze similar gains in food and feed assessments. Globally, the convergence of these frameworks-reinforced by OECD initiatives and NAM-enhanced case studies-signals an emerging international consensus on what constitutes defensible read-across. In conclusion, harmonizing EFSA's procedural roadmap with RAAF's evaluative rigor and GRAP's best-practice ethos can mainstream reliable, animal-saving read-across across regulatory domains, paving the way for fully mechanistic, AI-enabled chemical safety assessment.
PMID:41479674 | PMC:PMC12753468 | DOI:10.3389/ftox.2025.1690491
Open Res Eur. 2025 Dec 9;4:194. doi: 10.12688/openreseurope.18439.2. eCollection 2024.
ABSTRACT
Metabolism disrupting chemicals (MDCs) elicit negative effects on metabolically active organs such as the liver and the pancreas, altering normal metabolic processes. Chemicals that are known, or suspected MDCs include compounds found in everyday consumer products and food, making low-dose, continuous exposure inevitable for humans. Through the discovery of chemically induced metabolic disruption, a concern has surfaced whether and how MDCs impact human health and the development of metabolic diseases. This has accelerated research around the topic, and it has been found that exposure to MDCs is linked to increased incidence of metabolic diseases including obesity and liver steatosis. Effective regulatory action is hindered by the lack of accurate methods to identify MDCs. The NEMESIS project addresses this regulatory gap by investigating the mechanisms through which MDCs cause metabolic disruption. The project aims at identifying novel biomarkers of exposure and link exposure to disease outcomes. As chemical toxicity testing is rapidly moving towards new approach methodologies (NAMs), NEMESIS promotes non-animal methodologies by employing state-of-the-art in vitro methods, epidemiological data, systems biology approaches, and seeks to replace mammalian in vivo experiments with alternative models. By understanding mechanisms of MDC-induced metabolic health effects, and through the development of reliable effect biomarkers and testing strategies, the NEMESIS project aims to facilitate more effective regulatory measures to improve and protect the health and well-being of EU citizens. The project is particularly focused on maximizing its impact through effective dissemination and communication efforts, to ensure that the project's message and results reach a broad audience and are tailored to different population groups. These actions will improve the risk assessment of MDCs and ensure that the EU citizens are informed and protected from the harmful effects of MDCs and can adapt their consumer patterns and behaviors to prevent exposure.
PMID:41477515 | PMC:PMC12749563 | DOI:10.12688/openreseurope.18439.2
Toxics. 2025 Dec 3;13(12):1048. doi: 10.3390/toxics13121048.
ABSTRACT
The rapid growth in manufacturing and use of electrical and electronic equipment has led to unprecedented volumes of poorly managed e-waste, posing serious ecological risks. Although data on individual chemical substances in e-waste are available, evidence of ecotoxicity from actual e-waste materials remains scattered. This review consolidates organism-level ecotoxicity data on real e-waste samples (mixed fractions, fragments, leachates) and samples collected near e-waste facilities (soil, sediments, dust, water) across aquatic and terrestrial environments. It critically examines how methodological approaches influence reported outcomes and outlines research priorities. In aquatic environments, toxic responses vary with increased amounts of toxicants (dissolved metals, particles from dismantling operations) that mobilise to surface waters, while hydrophobic organic compounds cause sublethal behavioural and genotoxic effects. The few studies on terrestrial environments show impaired invertebrate growth and reproduction, along with changes in soil and "plastisphere" microbiota. However, tested concentrations, material complexity, and incomplete reporting of exposure chemistry, among other factors, limit the environmental relevance and comparability of the data. Uniformised procedures, combined with thorough chemical characterisation, environmentally realistic conditions, and cross-system bioassays (including different exposure routes and cumulative assessments), may provide mechanistic insights into e-waste toxicity, supporting evidence-based risk management strategies while contributing towards the development and validation of robust new approach methodologies (NAMs).
PMID:41441269 | PMC:PMC12737757 | DOI:10.3390/toxics13121048
Endocrinology. 2025 Dec 20:bqaf186. doi: 10.1210/endocr/bqaf186. Online ahead of print.
ABSTRACT
Endocrine mediators are essential for pregnancy maintenance, and their functional withdrawal is associated with normal term and preterm birth (PTB). Therefore, the disruption to endocrine functions or agents that can disrupt endocrine functions are naturally suspected as contributors to PTB. One of the well-studied endocrine-disrupting compounds is Polybrominated diphenyl ether(s) (PBDE). PBDE is a flame-retardant compound that is contained in several products and is a ubiquitous environmental contaminant. PBDE exists in several different congeners, many harmless compounds, but a few PBDE congeners are linked as endocrine disruptors contributing to adverse pregnancy outcomes like PTB. However, data ambiguity suggests that current platforms are insufficient to conclude PBDE's mechanisms of action as an endocrine disruptor at the feto-maternal interface (placenta/fetal membranes). The development of microfluidic-based new approach methods (NAMs) is being introduced to study PBDE and other environmental pollutants. Organs-on-a-chip (OOC) are an emerging class of NAMs that can replicate human organ-level functions in vitro. OOCs are microfluidic systems comprising multiple cell types from an organ that mimics the environment of a physiological organ. These devices are interconnected through microchannels to maintain intercellular interactions. OOC-based testing and development have accelerated globally as regulatory agencies now emphasize the need for reliable, humanized alternatives to traditional animal models. Multiple reproduction-associated OOCs are being developed, and their utility has been tested in assessing mechanisms of action and toxicological parameters of environmental pollutants. This review provides an overview of feto-maternal interface OOCs and uses PBDE as an example to demonstrate how OOCs can study endocrine-disrupting compounds.
PMID:41420538 | DOI:10.1210/endocr/bqaf186
The National Asset Management Agency has announced its final surplus transfer of €450m to the State and said its overall lifetime contribution had increased by €100m more than projected, to €5.6 billion.
The project Computational characterisation of ecological hazard and risk of environmental mixtures focused on two key priorities in advancing a next‐generation risk assessment (NGRA) workflow: (i) transitioning to assessing mixtures and (ii) utilising mechanism‐based hazard assessment based on new approach methodology (NAM) data.
New approach methodologies (NAMs) are gaining increasing attention within the scientific community as strategies to reduce the use of animal testing. Their integration into regulatory frameworks for chemical hazard and risk assessment in food and feed would directly support the goals of the European Food Safety Authority (EFSA).
Recently, I’m ashamed to admit, I received an email that initially made me feel warm, human, even grateful: a rejection for a job I’d applied to. But my thankful feelings quickly curdled into self-loathing—the nausea one gets when looking back over pathetic, paragraphs-long texts to an ex, whose monosyllabic responses suggested they’d clearly moved on. The rejection was a form letter, not even a late-round, personalized “we gave you serious consideration but ultimately decided to hire a VP’s nephew” message. I was so accustomed to being treated with indifference, I realized, that the barest acknowledgment of my existence felt like a win.
Putting aside the question of whether the job market itself is in good or bad shape right now (it’s bad), the code of what behavior is and isn’t acceptable seems to have broken down. Ghosting has become more rampant not just by employers but also by job seekers. In 2024, candidates reviewing employers on the website Glassdoor used the term ghosting nearly three times as much as they did in 2020. And a 2023 Indeed survey of job seekers found that 62 percent of respondents planned to ghost a prospective employer in a future job search, compared with only 37 percent in 2019. The disappearing act is not just in the early rounds, either. Employers routinely ask applicants to do multiple interviews and time-consuming test work, and are never heard from again; a survey this year from Greenhouse, a recruiting-software company, found that nearly two out of every three candidates in the United States had been ghosted after an interview. Meanwhile, some applicants who make it through the onerous hiring process and accept jobs never show up for their first day. One California recruiter told me that some of the candidates who ditched had even signed offers for positions that paid six-figure salaries. Today, many people on both sides of the hiring equation—whether because of convenience, self-protection, or resentment—have abandoned even the pretense of courtesy, resulting in a job market that’s as rude as it is dysfunctional.
[Read: The great ghosting paradox]
It might sound glib to blame a simple loss of manners for employment woes. The way cultural norms tell people to comport themselves can seem silly—like empty formality (Smile even if you don’t mean it) or even hypocrisy, because this behavior often hides the way people actually feel or what they actually want to say. But try going a single day telling a boss or a landlord, an employee or a co-worker, what you really think, and why everybody obscures their true opinions all the time would quickly become apparent.
Manners, it turns out, have long been smoothing over a lot of rough edges in the hiring process, hiding the uglier realities that can emerge from the unequal relationship between employers and the employed. In a society where many workers have little power, the fig leaf of societal niceties is often the only difference between a functioning system built on trust and a polarized one roiled by humiliation, disgust, and retribution.
Before a code of conduct was something that HR had you read and initial when you started a job, it was the main deterrent against open class warfare. Throughout the 18th and 19th centuries, in Europe and America, manners essentially boiled down to “Good fences make good neighbors.” People avoided class conflict mainly by avoiding each other; associating outside your class was strongly discouraged, according to Informalization, a history of manners by the sociologist Cas Wouters. When “class mixing” was necessary, people maintained psychological distance by acting with extreme formality, often making elaborate shows of haughty superiority or craven deference. When people clashed, norms dictated that the privileged classes were afforded the prerogative of violence; retribution from lower classes tended to be penalized. An 1859 guide to courtesy, The Habits of Good Society, for instance, advised that if a gentleman were confronted with a “dishonest cabman,” then “one well-dealt blow settles the whole matter.” The author goes on to specify that violence is only for pu
Comput Struct Biotechnol J. 2025 Nov 16;29:305-327. doi: 10.1016/j.csbj.2025.11.032. eCollection 2025.
ABSTRACT
Traditional in vivo methodologies have long formed the foundation of chemical and material safety assessment, yet they are increasingly inadequate to meet modern regulatory, ethical, and sustainability demands. These conventional approaches are resource-intensive, ethically questionable, and often fail to accurately predict human or environmental toxicity, particularly for emerging pollutants such as PFAS, (nano-) pesticides, and 2D materials. In response, the EU has launched initiatives like the Chemical Strategy for Sustainability and the Zero Pollution Action Plan under the European Green Deal to promote innovation in safer, and more sustainable chemicals. Central to this transformation is the Safe and Sustainable by Design (SSbD) framework, developed by the European Commission's Joint Research Center, which provides structured methodologies and metrics to integrate safety and sustainability into material innovation from the earliest stages of design. Building on this vision, the CHIASMA project aims to advance Next generation Safety Assessment (NGSA) by developing innovative New Approach Methodologies (NAMs) that combine experimental, computational, and Life Cycle Assessment (LCA) tools. Focusing on key biological systems and exposure routes, CHIASMA integrates Artificial Intelligence (AI), Machine Learning (ML), and Knowledge Graph (KG) technologies to enhance data interoperability and predictive accuracy. By embedding FAIR data principles and aligning with Good Laboratory Practice (GLP) standards, CHIASMA promotes transparency and regulatory acceptance. Fully aligned with SSbD principles, CHIASMA establishes a digital, interoperable infrastructure for predictive safety evaluation, that leverage on state-of-art experimental New Approach Methodologies (NAMs) bridging critical data gaps and supporting the transition towards sustainable, science-driven, and ethically responsible chemical and material innovation in Europe and beyond.
PMID:41362652 | PMC:PMC12682023 | DOI:10.1016/j.csbj.2025.11.032
Northern Ireland's former First Minister has given evidence to the NAMA trial in Belfast.
Front Toxicol. 2025 Nov 19;7:1665163. doi: 10.3389/ftox.2025.1665163. eCollection 2025.
ABSTRACT
INTRODUCTION: Per- and polyfluoroalkyl substances (PFAS) are persistent environmental pollutants with potential immunotoxic effects. Most toxicological studies have focused on long-chain PFAS such as perfluorooctanesulfonic acid (PFOS) and perfluorooctanoic acid (PFOA). However, short-chain and ultra-short-chain alternatives, including trifluoroacetic acid (TFA), are increasingly used despite limited toxicological data.
METHODS: This study evaluated and compared the immunotoxic effects of PFAS with varying chain lengths-long-, short-, ultra-short-chain compounds, and fluoropolymer representatives (polytetrafluoroethylene, PTFE)-using human-relevant new approach methodologies (NAMs). Two complementary in vitro models were employed. Peripheral blood mononuclear cells (PBMCs) from healthy donors to assess antibody (IgG and IgM) production. THP-1-derived dendritic cells (DCs) to evaluate maturation marker expression (CD83, CD86, HLA-DR). Environmentally and occupationally relevant PFAS concentrations were tested.
RESULTS: PFOS, PFOA, and perfluorononanoic acid (PFNA) suppressed antibody production and impaired DC maturation in a concentration-dependent manner, consistent with previous in vivo and epidemiological data. Short-chain PFAS (PFHxS, PFBS, PFHxA, PFBA) showed modest to intermediate immunomodulatory activity, with subtle immunosuppressive trends in female donors. Notably, TFA reduced antibody production at levels comparable to PFOS, indicating that chain length alone is not a reliable predictor of immunotoxic potential. PTFE exhibited no suppressive effects; instead, increased antibody release was observed in female donors, suggesting possible sex-dependent immunostimulation.
DISCUSSION: Findings support a nuanced, compound-specific approach to PFAS risk assessment rather than a simple long- vs. short-chain distinction. In vitro NAMs provided mechanistic, human-relevant insights and reinforce their integration into regulatory frameworks.
PMID:41347043 | PMC:PMC12672224 | DOI:10.3389/ftox.2025.1665163
Toxicol Sci. 2025 Dec 4:kfaf166. doi: 10.1093/toxsci/kfaf166. Online ahead of print.
ABSTRACT
The EPA continues to evaluate strategies to implement new approach methods (NAMs) for screening chemicals that disrupt the thyroid endocrine system. Validation of NAMs is a critical milestone toward establishing confidence in data sources that could be used in a regulatory decision-making context. The objective of this study was to conduct an interlaboratory validation of the human thyroid microtissue assay to evaluate its relevance and reliability. In coordination with the U.S. validation authority, NICEATM, and collaboration with industry partners (LifeNet Health, Bayer Crop Science, Corteva Agrisciences), the study aims were to 1) define the study design and establish standard operating procedures, 2) conduct test method transfer, training, and within-laboratory model performance evaluation, 3) perform interlaboratory reference chemical testing and assay performance evaluation. Progress was independently monitored by a validation management team comprised of an international group of experts in thyroid physiology, in vitro test methods, and regulatory toxicology. Results indicated the thyroid microtissue model could be reliably transferred to new laboratories with reproducible effects on thyroid hormone synthesis. Interlaboratory testing of four blinded reference chemicals (three true positive, one true negative) across three independent human donors revealed consistent bioactivity across the reference set and performance metrics (dynamic range, precision, screening quality) that met acceptance criteria and shed insight into areas for improvement. In the context of a NAM-based testing strategy, a validated human thyroid microtissue assay enables direct measurement of thyroid hormone synthesis perturbations, reducing reliance on animal testing and addressing a critical mode-of-action that is of regulatory concern.
PMID:41344865 | DOI:10.1093/toxsci/kfaf166
A judge has ruled that the trial of one of two men accused of fraud linked to the sale of NAMA's Northern Ireland loan book cannot continue.
The National Asset Management Agency (NAMA) was "broadly successful" in how it managed the process of selling off its assets, according to a new report.
Environ Sci Technol. 2025 Nov 21. doi: 10.1021/acs.est.5c11122. Online ahead of print.
ABSTRACT
The global restriction of legacy per- and polyfluoroalkyl substances (PFASs), such as perfluorooctanoic acid (PFOA), has led to increased production and environmental occurrence of emerging alternatives. This study applied a next generation risk assessment (NGRA) framework to evaluate and compare the local environmental risks of eight PFOA alternatives using multiple lines of evidence from new approach methodologies (NAMs). Targeted chemical analysis was conducted to quantify environmental concentrations in wastewater and surface waters near an industrial source. Hazard characterization integrated in silico modeling, in vitro assays using zebrafish liver cells, in vivo zebrafish embryo toxicity tests, and transcriptomic profiling. Multiple points of departure (PODs), including conventional effect concentrations, benchmark concentrations (BMC), transcriptomic POD (tPOD), and species sensitivity distributions (SSD), were derived. The results showed that HFPO-TA and HFPO-TeA were cytotoxic, while HFPO-DA, HFPO-TA, and HFPO-TeA caused significant embryo toxicity. HFPO-TeA emerged as the most potent alternative, with transcriptomic signatures implicating disruption of metabolic and cardiovascular functions. Risk quotients (RQ) for HFPO-DA exceeded concern thresholds across end points, while those for HFPO-TA and HFPO-TeA declined with appropriate assessment factor adjustments. These findings highlight potential local environmental risks from PFOA alternatives and demonstrate the utility of NAM-based NGRA frameworks.
PMID:41270243 | DOI:10.1021/acs.est.5c11122
Toxicol Sci. 2025 Nov 19:kfaf162. doi: 10.1093/toxsci/kfaf162. Online ahead of print.
ABSTRACT
Zebrafish (danio rerio) are an ideal system for understanding developmental toxicity as they display similar toxicity outcomes to other vertebrates. Further, many molecules have been tested for developmental toxicity in zebrafish providing an opportunity for machine learning model development. We curated 1345 small molecules from ToxCast, flame retardant compounds, per- and polyfluoroalkyl substances (PFAS), and industrial chemicals published by the Superfund Research Program (SRP). Following curation, we trained machine learning models on the zebrafish toxicity endpoints ANY_ = any effect including mortality, ANY_BUT_MORT = any effect excluding mortality, MORT = mortality ie did the embryo die, EDEM = did an edema form, CRAN = Craniofacial malformation. We demonstrated that these models were better than random when compared to shuffled data. We also fine-tuned the molecular SMILES encoder MolBART to predict on all zebrafish toxicity endpoints and found it generally matched the performance of classical machine learning models for ANY_BUT_MORT, CRAN, and EDEM endpoints. We present new toxicity data for Proteolysis Targeting Chimeras (PROTACs) in Zebrafish and machine learning models for these data by fingerprinting different parts of the molecule individually, yielding predictive performance (AUROC 0.6-0.7). If we are to reduce animal testing with new approach methodologies (NAMs) like these Zebrafish toxicity models they need to be able adapt to new molecular classes like PROTACs.
PMID:41259055 | DOI:10.1093/toxsci/kfaf162
International Journal of Toxicology, Ahead of Print.
The U.S. Food and Drug Administration (FDA), Center for Drug Evaluation and Research (CDER), and Office of New Drugs (OND) has continuously encouraged the submission of nonclinical tests utilizing new approach methodologies (NAMs) and sponsor engagement ...
International Journal of Toxicology, Ahead of Print.
The U.S. Food and Drug Administration (FDA), Center for Drug Evaluation and Research (CDER), and Office of New Drugs (OND) has continuously encouraged the submission of nonclinical tests utilizing new approach methodologies (NAMs) and sponsor engagement ...
J Hazard Mater. 2025 Oct 22;499:140194. doi: 10.1016/j.jhazmat.2025.140194. Online ahead of print.
ABSTRACT
Microplastics (MPs) are emerging environmental contaminants, yet their impact on autoimmune diseases such as rheumatoid arthritis (RA) remains unclear. We report that polystyrene microplastics (PS-MPs) are detectable in synovial fluid samples from RA patients and that exposure to 5 μm PS-MPs directly promotes the pathogenic activation of RA fibroblast-like synoviocytes (RA-FLSs), key effector cells in synovial inflammation and joint destruction. High-resolution imaging confirmed PS-MPs internalization into the cytoplasm of RA-FLSs, accompanied by cytoskeletal changes and mitochondrial cristae disruption indicative of intracellular stress. PS-MPs exposure activated NF-κB and MAPK (JNK/p38) signaling and induced the expression of IL-6, IL-8, CCL2, MMP3, MMP9, NAMPT, and TWIST1. These changes coincided with enhanced migration, invasion, and monocyte adhesion via increased VCAM-1 and ICAM-1. In vivo, chronic PS-MPs exposure aggravated inflammation in CFA-induced arthritis, with fluorescent particles accumulating in inflamed synovium. In humanized SCID co-implantation model, PS-MPs-treated RA-FLSs triggered greater cartilage erosion and macrophage infiltration. Importantly, pharmacologic inhibition of NF-κB and p38, as well as treatment with Ginsenoside Compound K (GCK), significantly reduced PS-MPs-induced cytokine production in vitro. Together, these findings demonstrate that MPs can directly activate synovial fibroblasts and aggravate RA pathology. This study identifies MPs as a previously unrecognized environmental cofactor in autoimmune joint disease.
PMID:41167161 | DOI:10.1016/j.jhazmat.2025.140194
Front Toxicol. 2025 Oct 8;7:1657708. doi: 10.3389/ftox.2025.1657708. eCollection 2025.
ABSTRACT
Scientific confidence frameworks (SCFs) are alternatives to traditional validation for new approach methodologies (NAMs). The SCFs adapted by the Interagency Coordinating Committee for the Validation of Alternative Methods (ICCVAM) and the American Chemistry Council (ACC) both address inference performance-the ability of NAMs to predict or inform the biological effect of interest. Inference performance is a distinct evaluation procedure in ACC's SCF but is blended into several steps of ICCVAM's SCF. Here, we first reproduce the previously derived human relevant potency threshold (HRPT) for the estrogen receptor alpha (ERα) agonism of Borgert et al. (2018) using guideline and guideline-like studies; we found that a HRPT of 1 to 10-1 positively and consistently predicted clinical endometrial and endocervical effects. We next mapped inference performance to ICCVAM's SCF and found that it can be used as an effective initial screen prior to performing more detailed characterizations in their SCF. We first conclude that a HRPT for ERα agonism of 10-2 to 10-4 is a health-protective NAM based on an established mode of action that could potentially be used in early screening, much like the threshold of toxicological concern. We then conclude that inference performance is a core requirement for SCFs.
PMID:41132625 | PMC:PMC12540326 | DOI:10.3389/ftox.2025.1657708
Mar Pollut Bull. 2025 Oct 17;222(Pt 3):118790. doi: 10.1016/j.marpolbul.2025.118790. Online ahead of print.
ABSTRACT
Coastal marine environments receive large influxes of anthropogenic contaminants from land-based sources such as metal(loid) s and emerging persistent organic pollutants (POPs). These contaminants can accumulate in marine sediments, seagrass and herbivorous marine megafauna such as dugongs, manatees and green turtles. Using the Preferred Reporting Items for Systematic reviews and Meta-Analyses (PRISMA) approach, we evaluated 131 publications on the bioaccumulation and effects of metal(loid)s and organic contaminants in seagrass, dugongs, manatees and green turtles. The global distribution of these publications was mapped, with concentration and effects data extracted to calculate chemical risk quotients (RQ). Metal(loid)s were the most studied contaminant class, while research on other chemicals including per - and polyfluoroalkyl substances (PFAS) and pharmaceuticals and personal care products (PPCP) were under-represented. The link between the concentration of a contaminant measured in tissue and its biological effect was often lacking. New Approach Methods (NAMs), which include in vitro assays and omics approaches, were highlighted as valuable but not widely used. The reported bioaccumulated concentrations of contaminants in the literature varied greatly, with little data on the cumulative risk of mixtures. The calculation of RQ suggest a risk to many wild populations to the adverse effects of some contaminants. This review emphasises the need for standardised methods and data sharing, which will foster a more unified and global approach to better assess the risk contaminants pose to seagrass and the megafauna they support.
PMID:41109174 | DOI:10.1016/j.marpolbul.2025.118790
Toxicol Sci. 2025 Sep 29:kfaf136. doi: 10.1093/toxsci/kfaf136. Online ahead of print.
ABSTRACT
Environmental exposure to industrial chemicals, endocrine disruptors, and pharmaceuticals has been increasingly linked to the global decline in male reproductive health. To address the urgent need for efficient and mechanistically informed toxicity screening, we developed a high-throughput, high-content screening (HTS/HCA) platform using a 3D in vitro mini-testis model. This system was used to evaluate 87 structurally diverse compounds from the National Toxicology Program (NTP) chemical library. The model incorporates murine-derived spermatogonia, Sertoli, and Leydig cells embedded in an extracellular matrix, providing a physiologically relevant environment for mechanistic toxicology. Each compound was tested across ten phenotypic endpoints, including nuclear morphology, cytoskeletal integrity (F-actin), DNA damage (γH2AX), and cell viability by using high-content imaging. Quantitative Points of Departures (PODs) were calculated and integrated into a High-content Assay Index (HCAI). Toxicological Priority Index (ToxPi) scores, derived from the PODs, enabled compound ranking and clustering. Compared to existing in vivo reproductive toxicity data, the 3D model demonstrated 91.5% sensitivity, 93.8% specificity, and 93.6% concordance (n = 64 compounds). Notably, 22 compounds lacking reproductive toxicity data were identified as potentially reproductive toxicants. Mechanistic analyses revealed that nuclear morphology, F-actin intensity, and γH2AX were the most sensitive indicators of reproductive toxicity. Cluster and category-level analysis showed that flame retardants and pesticides ranked highest in toxicity. The integration of multi-parametric data via ToxPi facilitated high-resolution chemical prioritization. Given current ethical and technical challenges in sourcing human testicular tissue or differentiating stem cells into testicular cell types, murine cells provide a reproducible and practical alternative for complex multicellular testis modeling. Our results demonstrate that the HCA-integrated 3D mini-testis model offers a robust, scalable, and mechanistically insightful platform for male reproductive toxicity screening, supporting its adoption as New Approach Methodologies (NAMs) aligned with regulatory and ethical testing goals.
PMID:41017718 | DOI:10.1093/toxsci/kfaf136
Front Endocrinol (Lausanne). 2025 Sep 12;16:1654965. doi: 10.3389/fendo.2025.1654965. eCollection 2025.
ABSTRACT
INTRODUCTION: Intrauterine exposure to endocrine disrupting chemicals (EDCs), particularly anti-androgens, has been implicated in hypospadias by disrupting fetal masculinization of the genital tubercle (GT). Other pathways, including estrogen signaling, may also contribute but remain poorly characterized, especially in rats - a key model in chemical toxicity testing. Estrogen signaling has also been linked to hypospadias in mice, raising questions about androgen-estrogen interactions in guiding GT differentiation.
METHODS: We induced hypospadias in male rat offspring via intrauterine exposure to the antiandrogenic drug flutamide and characterized androgen and estrogen receptor expression.
RESULTS: We observed key structural and transcriptional changes in the developing penis, including altered estrogen receptor a (ERa, Esr1) expression. Notably, beyond this established androgen-estrogen relationship in hormone-sensitive tissues, anti-androgenic exposure also induced spatial changes in Esr1 expression in specific regions of the GT.
DISCUSSION: Future toxicological testing using new approach methodologies (NAMs) should consider androgen-estrogen balance and crosstalk in reproductive tissues as a mechanism of action.
PMID:41019341 | PMC:PMC12464887 | DOI:10.3389/fendo.2025.1654965
Mol Reprod Dev. 2025 Sep;92(9):e70058. doi: 10.1002/mrd.70058.
ABSTRACT
Visfatin, a multifunctional adipokine, plays a crucial role in ovarian physiology. This study examines its effects on the level of selected angiogenesis and apoptosis factors in the porcine corpus luteum (CL). Using in vitro cultures of luteal cells, we analyzed the effects of visfatin on key angiogenic factors (VEGF family, PDGF, bFGF2, ANG, and iNOS) and their receptors. Additionally, we assessed luteal cell viability and apoptosis by measuring caspase activity, DNA fragmentation, and transcript levels of apoptotic factors. We noted that visfatin mostly decreases the expression of angiogenic factors and their receptors during the luteal phase except for VEGFC, VEGFR3 on Days 2-3 of the estrous cycle, and VEGFB, VEGFD, VEGFR1, VEGFR2, FGFR2 on Days 14-16 of the estrous cycle, which were upregulated. Moreover, visfatin inhibits apoptosis by downregulating caspase-3, -8, -9, BAX, and caspase 3/7 activity while increasing BCL-2 and luteal cells viability. Treatment with FK866, nicotinamide phosphoribosyltransferase (NAMPT) inhibitor, largely disrupted observed effects, highlighting the NAMPT-dependent function of visfatin. Moreover, visfatin suppressed VEGF-A levels via insulin receptor and MAPK, while insulin receptor, MAPK, and AKT pathways mediated the inhibition of caspase 3/7 activity. These findings suggest that visfatin regulates apoptosis and angiogenesis in the porcine CL.
PMID:41015905 | DOI:10.1002/mrd.70058
Environ Sci Process Impacts. 2025 Sep 19. doi: 10.1039/d5em00084j. Online ahead of print.
ABSTRACT
To explore the use of molecular docking as a high throughput in silico screening tool for identifying chemicals of environmental health concern, we conducted a case study to assess endocrine disruption effects due to targeting of nuclear receptors (NRs) by chemicals with backbone structures like bisphenols, but with varied functional groups. The molecular docking analysis elucidates how functional groups of the chemicals, such as NH2, Cl, and OCH3, influence their interaction with the human estrogen receptor alpha (hERα), a key player in endocrine regulation. Through comparative docking analysis, we examined how bisphenol analogs interact with three distinct conformations of hERα: the apo structure and two structures with bound agonist and antagonist ligands. Water molecules within the protein and surrounding the ligand binding domain (LBD) were found to have little impact on the affinity of compounds binding to the receptor across various conformations. This can be attributed to the hydrophobic nature of the ligand-binding pocket, which consists mainly of hydrophobic amino acid residues and binding sites. In the assessment of bisphenol analogs compared to well established endocrine disrupting chemicals (EDCs), it was observed that these analogs exhibit characteristics commonly associated with endocrine disruptors. While compounds like BPA and BPF exhibited partial agonist activity, stimulating hERα activity to varying degrees, other compounds displayed non-agonist behavior, suggesting a different mode of interaction with the receptor. Further analysis revealed the significance of specific functional groups, such as hydroxyl or amine groups, on the aromatic ring of these compounds in modulating their binding affinity to hERα. Within the ligand binding site of hERα, amino acid residues Glu353, Arg394, and His524 have the capacity to form hydrogen bonds with hydroxyl or amine groups. Protonation or deprotonation of these groups can further alter their binding affinity, thereby influencing their interaction with estrogen receptors and subsequent estrogenic effects. Via this case study, we demonstrate the potential and provide best practices of using molecular docking as a new approach methodology (NAM) for chemical assessments and regulations.
PMID:40970403 | DOI:10.1039/d5em00084j
International Journal of Toxicology, Volume 44, Issue 6, Page 499-506, November/December 2025.
Technological advances and the desire to reduce dependence on animal models have brought human-relevant models to the forefront of drug development. This paradigm shift is leveraging the advances inin vitrosystems and new approach methodologies (NAMs), ...
International Journal of Toxicology, Ahead of Print.
Technological advances and the desire to reduce dependence on animal models have brought human-relevant models to the forefront of drug development. This paradigm shift is leveraging the advances inin vitrosystems and new approach methodologies (NAMs), ...
Enzyme Microb Technol. 2025 Jul 24;191:110724. doi: 10.1016/j.enzmictec.2025.110724. Online ahead of print. ABSTRACT Poly and per fluorinated substances (PFAS) are emerging contaminants of concern that are thought to be involved in causing numerous adverse health effects, such as immunosuppression, increased chance of cancer development, and altered levels of hepatic enzyme levels in humans. However, PFAS are considered highly persistent and resistant to biodegradation given the fact that the C-F bond can have a bond dissociation energy of up to 544 kJ/mol. Though many studies have reported PFAS biodefluorination by bacterial isolates and microbial communities, little is known regarding the molecular foundations for biodefluorination. In this study, a novel defluorinase was identified, that is responsible for the biodefluorination of 6:2 fluorotelomer carboxylic acid (6:2 FTCA) in R.jostii RHA1 using the combination of transposome-based insertional mutagenesis and heterologous expression. From a library of 417 R.jostii RHA1. mutants, 3 individual mutants lost their ability for defluorination when they were exposed to 6:2 FTCA (mutant # 15, 32 and 38 - Table S2). The disruption of the genetic locus in all 3 non-defluorinating mutants was identified coding for a putative MhPC superfamily protein. The MhPC superfamily of proteins is known to harbor other proteins such as fluoroacetate dehalogenase (UniProt - Q6NAM1) that are capable of -C-F bond cleavage. This identified gene was cloned into the heterologous expression host M. smegmatis MC2-155. After induction, the M. smegmatis MC2-155 transformant exhibited the ability to defluorinate 6:2 FTCA at a rate of 13 µmol/h (Vmax = 80.9 µmol/min and Km = 5.04 mM in Michaelis-Menten models). In contrast, defluorination was not observed in either abiotic or biotic controls. Further characterization of the novel defluorinase indicated that it could moderately defluorinate the unsaturated PFAS compound 6:2 FTCUA (4.9 µmol/h fluoride) and minimally defluorinate 5:2 sFTOH (1.3 µmol/h fluoride). The novel defluorinase indicated a maximal specific activity of 12.9 ± 1.9 µmol F/hr/g protein, against its primary PFAS substrate, 6:2 FTCA. However, it showed no activity with 5:3 FTCA or sulfonated PFAS compounds such as 6:2 FTS and 8:2 FTS. The wild-type Rhodococcus could defluorinate 6:2 FTCA at a rate of 2.2 µmol/h. The discovery of this MhPC class novel defluorinase in WT R.jostii RHA1. has substantial value since it is responsible for the critical step that initiates defluorination of PFAS compounds such as 6:2 FTCA. PMID:40714482 | DOI:10.1016/j.enzmictec.2025.110724 J Toxicol Environ Health B Crit Rev. 2025 Jul 6:1-69. doi: 10.1080/10937404.2025.2521615. Online ahead of print. ABSTRACT Per- and polyfluoroalkyl substances (PFAS) comprise a large class of human-made chemicals that are in widespread use and present concerns for persistence, bioaccumulation and toxicity. Whilst a handful of PFAS have been characterized for their hazard profiles, the vast majority of PFAS have not been extensively studied. A comprehensive evaluation to characterize the hazard profiles of the thousands of available PFAS would require extensive resources in terms of cost, number of animals and time. An alternative and more efficient approach is to develop a structural chemical categorization approach to prioritize which PFAS or categories of PFAS should be subject to additional study. To that end, the U.S. Environmental Protection Agency (EPA), in collaboration with the National Institute of Environmental Health Sciences (NIEHS) Division of Translational Toxicology (DTT), initiated a research project in 2018 to screen approximately 150 PFAS through a battery of alternative model organisms, in vitro cell and biochemical assays, and in vitro toxico kinetic (TK) assays in order to inform chemical category and read-across approaches. The aim of this review summarizes the experimental testing undertaken, how data were processed, what insights were derived from a category perspective and how these might potentially inform subsequent tiered testing. PMID:40619890 | DOI:10.1080/10937404.2025.2521615 J Hypertens. 2025 Jun 13. doi: 10.1097/HJH.0000000000004085. Online ahead of print. ABSTRACT OBJECTIVES: Hypertension-induced endothelial damage in cerebral microvessels is a key factor contributing to vascular cognitive impairment (VCI). Endothelial function stabilization considerably depends on the endothelial nitric oxide synthase (eNOS)/nitrogen monoxide (NO) pathway. Furthermore, the eNOS/NO signaling pathway plays a role in stabilizing the vascular endothelium. Although betaine (bet) has been shown to improve cognitive dysfunction, its underlying mechanisms remain unclear. Therefore, this study aimed to determine whether betaine protects cognitive function by regulating eNOS/NO activity. METHODS: Male 7-month-old spontaneously hypertensive rats (SHR) were randomly assigned to four groups: SHR, Bet, Bet and N(G)-Nitroarginine methyl ester hydrochloride (L-NAME), and L-NAME groups. Male 7-month-old Wistar Kyoto rats (WKY) served as controls. All animals received treatment or saline for 4 weeks. In-vitro experiments were conducted using rat brain microvascular endothelial cells (RBMECs) treated with either homocysteine (Hcy) or betaine. Behavioral experiments, western blotting, pathological tissue staining, Doppler ultrasound technique, and ELISA were employed to assess the impact of hypertension on cognitive and endothelial functions. RESULTS: Hypertension led to cognitive decline in SHR by causing endothelial dysfunction, blood-brain barrier disruption, inflammation, oxidative stress, and apoptosis. Bet administration significantly improved these pathological indicators of cognitive impairment; however, the eNOS inhibitor L-NAME reversed its effects. CONCLUSION: Our findings suggest that betaine protects vascular endothelium and improves VCI by modulating the eNOS/NO signaling pathway. PMID:40530574 | DOI:10.1097/HJH.0000000000004085 Regul Toxicol Pharmacol. 2025 Jun 6;162:105883. doi: 10.1016/j.yrtph.2025.105883. Online ahead of print. ABSTRACT Exposure to endocrine disruptors (EDs) are associated with significant risks to human health and the environment. The European Union (EU) thus prioritizes their identification and regulation and is developing a roadmap to phase out animal testing in chemical safety assessments while advancing New Approach Methodologies (NAMs). This review outlines EU's practices for ED identification, focusing on the use of NAMs, as well as Defined Approaches and read-across. We assessed the current EU framework under the Classification, Labelling and Packaging (CLP) Regulation, the Registration, Evaluation, Authorisation and Restriction of Chemicals (REACH), the Plant Protection Products Regulation (PPPR), and the Biocidal Products Regulation (BPR), evaluating current use of NAMs and reflection on potential future use. We find that EU legislation and guidance documents allow the use of NAMs in ED identification, including for assessment of endocrine activity and adversity. However, guidance on predicting adversity using NAMs remains limited, and ED identifications have largely depended on animal data to assess endocrine-mediated adversity. Continued in vivo testing until reliable methodologies are accepted as alternatives and routinely applied is required. The report concludes with short- and long-term recommendations for updates to the information requirements across regulations and further development of methods to predict endocrine-mediated adversity. PMID:40484110 | DOI:10.1016/j.yrtph.2025.105883 Regul Toxicol Pharmacol. 2025 Nov;162:105884. doi: 10.1016/j.yrtph.2025.105884. Epub 2025 Jun 6. ABSTRACT Read-across is a widely used adaptation to address information requirements under REACH. However, registrants submitting for the 2010 and 2013 deadline have often failed to satisfy regulatory requirements from ECHA's point of view. Due to their complex composition and unique physicochemical properties, surfactants are posing major challenges in this respect. With the aim to improve future submissions and prevent unnecessary animal testing, read-across-related discussions of 72 ECHA Final Decisions on Compliance Checks and Testing Proposal Evaluations of 24 major surfactant groups were analysed in-depth, and causes for acceptance or rejection were identified. Key drivers of regulatory acceptance/rejection were presence or absence of composition information, considerations on structural similarity as well as availability and nature of bridging studies. Several elements were identified that may be easily improved in future REACH dossiers without the need for additional animal testing. Other cases revealed uncertainty of expectations by ECHA, highlighting the need for improved communication during the dossier preparation. Notably, no example for acceptance of read-across based on non-animal New Approach Methodologies (NAMs) was identified in this analysis. Owing to the benefits that non-animal NAMs may present as supporting information, all stakeholders are encouraged to contribute to an increase of regulatory acceptance. PMID:40484111 | DOI:10.1016/j.yrtph.2025.105884 The American Chemistry Council’s Long-Range Research Initiative (LRI) is proud to announce its participation in the NIH Common Fund’s Complement Animal Research In Experimentation (Complement-ARIE) public-private partnership (PPP) to accelerate the scientific development and evaluation of New Approach Methodologies (NAMs). Sci Total Environ. 2025 Jul 1;984:179690. doi: 10.1016/j.scitotenv.2025.179690. Epub 2025 May 24. ABSTRACT Aquatic ecosystems provide essential services, yet they face increasing pressures from anthropogenic activities, including land-use change, eutrophication, browning, and contaminant pollution. While the ecological effects of these stressors are documented, the impacts of complex contaminant mixtures, particularly those from wastewater treatment plant (WWTP) effluents, remain poorly understood. Mixtures effects are typically assessed using traditional species-by-species toxicological approaches, which, though the gold standard, are time-intensive, require test animals, and have limited extrapolability. New Approach Methodologies (NAMs), such as environmental DNA (eDNA), offer a non-invasive alternative, enabling broader assessments of taxa responses across trophic levels. Here, we apply an eDNA approach to assess community-wide responses to effluent discharge in the St. Lawrence River, one of North America's most diverse freshwater ecosystems. We sampled water and aquatic communities along the effluent plume of the Montréal WWTP, analyzing taxa-specific responses across trophic levels using high-throughput sequencing. We evaluated the influence of water physico-chemistry and per- and polyfluoroalkyl substances (PFAS) on aquatic beta diversity and network structure. To validate our eDNA results, we compared fish-specific detections with traditional fishing surveys. Our findings highlight how wastewater-derived contaminants influence biodiversity patterns and species interactions, with taxonomic responses varying across trophic levels. Network analyses revealed shifts in ecological stability, with changes in species connectivity and modularity influenced by effluent exposure. This study demonstrates the value of eDNA for characterizing biodiversity responses to anthropogenic stressors and provides insights into the broader implications of point-source pollution for freshwater ecosystem resilience. PMID:40414060 | DOI:10.1016/j.scitotenv.2025.179690 Neurotoxicology. 2025 Jul;109:11-26. doi: 10.1016/j.neuro.2025.05.006. Epub 2025 May 19. ABSTRACT With the societal interest to decrease experimental animal testing for regulatory purposes, the need for reliable new approach methods (NAMs) has become evident. To ensure the continued safe use of chemicals, NAMs should perform ideally at a comparable or better level of sensitivity and specificity as the in vivo modalities that they aim to replace, especially if they are to be used for hazard assessment. The use of relevant reference substances, selected with transparent criteria, forms a cornerstone of developing and validating (in silico and in vitro) NAMs. Claims on sensitivity and specificity should be based on results generated with reference chemicals that were previously scrutinised by independent expert panels on whether the substance has a hazardous property. CLP (Regulation (EC) No 1272/2008 on the classification, labelling and packaging of substances and mixtures) forms a key pillar in EU chemicals management. The evaluation of all available information by the Committee for Risk Assessment (RAC) and their comparison to CLP classification criteria creates the opportunity to objectively compile lists of positive reference substances. We collated a reference list of neurotoxic substances to aid in the development of neurotoxicity NAMs. We screened CLP Annex VI and reflected on existing reference lists and mode of action data. The identified substances included industrial chemicals and active substances in plant protection products and biocidal products. This list of neurotoxicants is not an exhaustive consensus list, which ideally would be the result of combining this list with those generated by other authorities or expert groups. PMID:40398565 | DOI:10.1016/j.neuro.2025.05.006 Deadline: 2025-06-03
Approximate launch date: 05/06/2025 In 2023 EFSA launched a project to promote the use of New Approach Methods (NAMs) into the risk assessment of pesticide residues metabolites (NAMs4PestRes) through the use of the OECD QSAR toolbox, a free available in silico tool managed by ECHA, to enhance connectivity of IUCLID and MetaPath and to implement embedded in silico models. The result will be an automated and standardized workflow for the assessment of mutagenicity (bacterial gene mutation) of pesticide residue metabolites to be implemented in a... The 2nd Commission Conference on the Roadmap towards Phasing out Animal Testing for Chemical Safety Assessments, held on 25 October 2024, brought together key stakeholders from regulatory bodies, industry, research organisations, and NGOs to discuss the future of chemical safety assessments done using non-animal methods (NAMs) in the EU. This second conference aimed to collect input from various stakeholders on how they address the transition from animal-based testing to innovative, reliable non-animal alternatives and to report progress with the development of the roadmap, which will set out actions and milestones to drive this transformation across chemical safety assessments. Toxicology. 2025 Aug;515:154167. doi: 10.1016/j.tox.2025.154167. Epub 2025 Apr 27. ABSTRACT Chronic liver diseases, including metabolic dysfunction-associated steatosic liver disease (MASLD) and hepatocellular carcinoma (HCC), are on the rise, potentially due to daily exposure to complex mixtures of chemical compounds forming part of the exposome. Understanding the mechanisms involved in hepatotoxicity of these mixtures is essential to identify common molecular targets that may highlight potential interactions at the molecular level. We illustrated this issue with two families of environmental contaminants, per- and polyfluoroalkyl substances (PFAS) and heterocyclic aromatic amines (HAAs), both of which could be involved in the progression of chronic liver diseases, and whose toxicity may be potentiated by interactions at the level of xenobiotic metabolism. In the study of exposome effects on chronic liver disease, New Approach Methodologies (NAMs) including omics analyses and data from various in vitro, in vivo and in silico approaches, are crucial for improving predictivity of toxicological studies in humans while reducing animal experimentation. Additionally, the development of complex in vitro human liver cell models, such as organoids, is essential to avoid interspecies differences that minimize the risk for humans. All together, these approaches will contribute to construct Adverse Outcome Pathways (AOPs) and could be applied not only to PFAS mixtures but also to other chemical families, providing valuable insights into mixture hepatotoxicity prediction in the study of the exposome. A better understanding of toxicological mechanisms will clarify the role of environmental contaminant mixtures in the development of MASLD and HCC, supporting risk assessment for better treatment, monitoring and prevention strategies. PMID:40300710 | DOI:10.1016/j.tox.2025.154167 Front Endocrinol (Lausanne). 2025 Apr 4;16:1472563. doi: 10.3389/fendo.2025.1472563. eCollection 2025. ABSTRACT INTRODUCTION: The Constitutive Androstane Receptor (CAR) (NR1I3), a pivotal member of the xenosensor family, plays a key role in the hepatic detoxification of xenobiotic and endobiotic chemicals through the induction of the expression of drug-metabolizing enzymes and transporters. CAR's involvement extends beyond detoxification, influencing gluconeogenesis, lipogenesis, bile acid regulation, and cellular processes such as proliferation, tissue regeneration, and carcinogenesis. This review explores CAR regulation by various factors, highlighting its role in mediating metabolic changes induced by environmental contaminants. METHODS: A literature search was conducted to identify all articles on the PubMed website in which the CAR-contaminant and CAR-hepatic steatosis relationship is analyzed in both in vitro and in vivo models. RESULTS: Numerous contaminants, such as perfluorooctanoic acid (PFOA), Zearalenone mycotoxin, PCB, triazole fungicide propiconazole can activate hepatic nuclear receptors contributing to the development of steatosis through increased de novo lipogenesis, decreased fatty acid oxidation, increased hepatic lipid uptake, and decreased gluconeogenesis. Indirect CAR activation pathways, particularly involving PFOA, are discussed in the context of PPARα-independent mechanisms leading to hepatotoxicity, including hepatocellular hypertrophy and necrosis, and their implications in nonalcoholic steatohepatitis (NASH) and nonalcoholic fatty liver disease (NAFLD). The prevalence of NAFLD, a significant component of metabolic syndrome, underscores the importance of understanding CAR's role in its pathogenesis. CONCLUSIONS: Experimental and epidemiological data suggest that endocrine disruptors, especially pesticides, play a significant role in NAFLD's development and progression via CAR-regulated pathways. This review advocates for the inclusion of modern toxicological risk assessment tools, such as New Approach Methodologies (NAMs), Adverse Outcome Pathways (AOPs), and Integrated Approaches to Testing and Assessment (IATA), to elucidate CAR-mediated effects and enhance regulatory frameworks. PMID:40255499 | PMC:PMC12005993 | DOI:10.3389/fendo.2025.1472563 This editorial provides an update on research & innovation (R&I) needs that can support EFSA's regulatory science in the coming years. The paper presents research needs for EFSA's work in a number of domains: omics technologies; gut microbiome; new approach methodologies (NAMs); allergenicity risk assessment; aggregate exposure assessment and environmental risk assessment (ERA). In briefly describing R&I needs, the document also addresses emerging challenges and opportunities. J Hazard Mater. 2025 Mar 17;491:137949. doi: 10.1016/j.jhazmat.2025.137949. Online ahead of print. ABSTRACT A tiered in vitro/in silico approach was developed to screen 12,654 per- and polyfluoroalkyl substances (PFAS) for their potential to disrupt the thyroid hormone transport. Initially, a set of 45 PFAS was tested using TTR-TRβ-CALUX bioassay, which was subsequently employed to develop a classification model, distinguishing active and inactive PFAS. The model fulfills all good practices for QSAR model validation and can predict whether a given PFAS can disrupt plasma transport of the thyroid hormone (T4). Subsequently, active compounds were used to develop two regression approaches: (i) multiple linear regression MLR, and (ii) second approach aimed at identifying multiple valid QSAR models based on different data-splitting strategies. Finally, a comprehensive virtual screening of a large PFAS dataset was conducted to assess their potency in disrupting thyroid hormone transport. The predictions indicated that more than 7500 compounds were active with over 100 PFAS potentially causing even greater adverse effects than PFOA. These findings highlight the critical role of integrating New Approach Methodologies (NAM)-based in vitro toxicity testing with multifaceted molecular modeling in assessing the risks associated with PFAS contamination in environmental matrices. PMID:40120279 | DOI:10.1016/j.jhazmat.2025.137949 Toxicology. 2025 Mar 5;514:154104. doi: 10.1016/j.tox.2025.154104. Online ahead of print. ABSTRACT New Approach Methodologies (NAMs), including cell culture and multi-level omics analyses, are promising alternatives to animal testing. To become useable for risk assessment purposes, they have to be applicable for different substance groups. One important group of substances is food contaminants, including synthetic chemicals, such as perfluorooctanesulfonic acid (PFOS) and perfluorooctanoic acid (PFOA), and natural compounds, such as mycotoxins and pyrrolizidine alkaloids. We tested five known contaminants affecting the liver and/or the kidney - PFOS, PFOA, Aflatoxin B1 (AB1), lasiocarpine (Las), and cadmium chloride - using HepaRG and RPTEC/TERT1 cells at non-cytotoxic concentrations for 36 and 72 h. Our NAM-based testing protocol included marker protein analysis for cellular functions and targeted transcriptomics followed by bioinformatics pathway analysis. The effects observed were compared with established in vivo results. Protein analysis indicated various affected pathways in HepaRG cells, with generally fewer effects in RPTEC/TERT1 cells. The strongest transcriptional impact was noted for Las in HepaRG cells. This study demonstrated the test protocol's applicability across different substances, revealing significant differences between HepaRG and RPTEC/TERT1 cell lines. RPTEC/TERT1 cells, while expressing renal-specific CYP enzymes, were less suitable for detecting effects of substances requiring hepatic metabolic activation, like Las and AB1. Our data supports the concept of specific pathway toxicity, with pathway analysis enabling the prediction of effects based on mechanism rather than target organ. Employing multiple omics techniques provided comprehensive insights into various compound effects, including steatosis, reactive oxygen species production and DNA damage, highlighting the potential of an extended omics approach for advancing toxicological assessments. PMID:40054833 | DOI:10.1016/j.tox.2025.154104 Aquat Toxicol. 2025 Apr;281:107302. doi: 10.1016/j.aquatox.2025.107302. Epub 2025 Feb 21. ABSTRACT Effective and reliable prediction for ecotoxicity, especially when affecting different levels of trophic chains, including humans, is increasingly gaining even more prominence as ecosystems face new threats and challenges, as that posed by the per- and poly-fluoroalkyl substances (PFAS). Toxicological prediction of PFAS in aquatic organisms, such as zebrafish, can be efficiently achieved through computational ecotoxicological approaches which are fully aligned with the state-of-the-art of new approach methodologies (NAMs) and current regulatory recommendations. Specifically in this work, the PFAS toxicodynamics interaction on the zebrafish mitochondrial voltage-dependent anion channel (zfVDAC2) was evaluated, mimicking in silico the PFAS bioaccumulation in low-concentration by integrating structure-based virtual screening (SB-VS) and predictive quantitative structure-activity(mitotoxicity) relationship (QSAR) methodologies (e.g., 2D/3D-QSAR) to address mechanistic aspects of PFAS toxicity. The best ranked PFAS pose docked in zfVDAC2 exhibits a ΔG-binding affinity higher than the ATP, i.e., the native substrate of the zfVDAC2 channel, with prevalence of van der Waal interactions, followed by fluorine (F)-halogen-bonds and finally hydrogen-bonds interactions. Mitochondrial ATP-transport blocking is thus suggested to be linked with local-flexibility perturbations in the zfVDAC2. Similarly, the obtained 2D/3D- QSAR models point out the packing density index as the most significant PFAS molecular descriptor to induce toxicity in the zfVDAC2, and mainly involving van der Waal interactions. The predictive and statistical performance of these models further indicate its NAM relevance regarding PFAS risk assessment while highlighting its interoperability and extrapolation capability for the ecotoxicological evaluation of other families of compounds. PMID:40024016 | DOI:10.1016/j.aquatox.2025.107302 J Environ Manage. 2025 Mar;377:124654. doi: 10.1016/j.jenvman.2025.124654. Epub 2025 Feb 20. ABSTRACT This study investigates waste management behaviors and public awareness of persistent organic pollutants (POPs) like PBDEs and PFAS in coastal communities of Newfoundland, Canada. Protecting these unique environments requires responsible waste disposal practices. Using an integrated theoretical framework combining the Theory of Planned Behavior (TPB), the Value-Belief-Norm (VBN) theory, and the Norm Activation Model (NAM), we conducted a mixed-methods study employing a pretested survey with open- and closed-ended questions. Although a larger sample was planned, 86 adult residents completed the survey. Our analysis revealed significant differences in waste management behaviors across community types (cities, big towns, and small towns). For example, cities showed higher engagement in e-waste recycling (82%) compared to smaller towns (68%), while smaller towns were more consistent in composting (78% vs. 50% in cities) and hazardous waste disposal (χ2 = 33.97, p = 0.0021). Higher education and income levels were positively correlated with increased recycling and proper waste disposal. However, despite a general awareness of environmental issues, knowledge of specific environmental contaminants was limited (45% for PBDEs, 33% for PFAS). These findings highlight the urgent need for targeted public education campaigns and improved waste management services tailored to the unique needs of diverse coastal communities. This study provides valuable insights for policymakers and environmental managers, emphasizing the importance of targeted interventions to promote sustainable practices and protect fragile coastal ecosystems. PMID:39983581 | DOI:10.1016/j.jenvman.2025.124654 Comput Toxicol. 2024 Jun;30:1-15. doi: 10.1016/j.comtox.2024.100304. ABSTRACT Read-across is a well-established data-gap filling technique used within analogue or category approaches. Acceptance remains an issue, mainly due to the difficulties of addressing residual uncertainties associated with a read-across prediction and because assessments are expert-driven. Frameworks to develop, assess and document read-across may help reduce variability in read-across results. Data-driven read-across approaches such as Generalised Read-Across (GenRA) include quantification of uncertainties and performance. GenRA also affords opportunities on how New Approach Method (NAM) data can be systematically incorporated to support the read-across hypothesis. Herein, a systematic investigation of differences in expert-driven read-across with data-driven approaches was pursued in terms of establishing scientific confidence in the use of read-across. A dataset of expert-driven read-across assessments that made use of registration data as disseminated in the public International Uniform Chemical Information Database (IUCLID) (version 6) of Registration, Evaluation, Authorisation and Restriction of Chemicals (REACH) Study Results were compiled. A dataset of ~5000 read-across cases pertaining to repeated dose and developmental toxicity was extracted and mapped to content within EPA's Distributed Structure Searchable Toxicity database (DSSTox) to retrieve chemical name and structural identification information. Content could be mapped to ~3600 cases which when filtered for unique cases with curated quantitative structure-activity relationship-ready SMILES resulted in 389 target-source analogue pairs. The similarity between target and the source analogues on the basis of different contexts - from structural similarity using chemical fingerprints to metabolic similarity using predicted metabolic information was evaluated. An attempt was also made to quantify the relative contribution each similarity context played relative to the target-source analogue pairs by deriving a model which predicted known analogue pairs. Finally, point of departure values (PODs) were predicted using the GenRA approach underpinned by data extracted from the EPA's Toxicity Values Database (ToxValDB). The GenRA predicted PODs were compared with those reported within the REACH dossiers themselves. This study offers generalisable insights on how read-across is already applied for regulatory submissions and expectations on the levels of similarity necessary to make decisions. PMID:38993812 | PMC:PMC11235147 | DOI:10.1016/j.comtox.2024.100304 Front Toxicol. 2024 Apr 26;6:1359507. doi: 10.3389/ftox.2024.1359507. eCollection 2024. ABSTRACT In the European regulatory context, rodent in vivo studies are the predominant source of neurotoxicity information. Although they form a cornerstone of neurotoxicological assessments, they are costly and the topic of ethical debate. While the public expects chemicals and products to be safe for the developing and mature nervous systems, considerable numbers of chemicals in commerce have not, or only to a limited extent, been assessed for their potential to cause neurotoxicity. As such, there is a societal push toward the replacement of animal models with in vitro or alternative methods. New approach methods (NAMs) can contribute to the regulatory knowledge base, increase chemical safety, and modernize chemical hazard and risk assessment. Provided they reach an acceptable level of regulatory relevance and reliability, NAMs may be considered as replacements for specific in vivo studies. The European Partnership for the Assessment of Risks from Chemicals (PARC) addresses challenges to the development and implementation of NAMs in chemical risk assessment. In collaboration with regulatory agencies, Project 5.2.1e (Neurotoxicity) aims to develop and evaluate NAMs for developmental neurotoxicity (DNT) and adult neurotoxicity (ANT) and to understand the applicability domain of specific NAMs for the detection of endocrine disruption and epigenetic perturbation. To speed up assay time and reduce costs, we identify early indicators of later-onset effects. Ultimately, we will assemble second-generation developmental neurotoxicity and first-generation adult neurotoxicity test batteries, both of which aim to provide regulatory hazard and risk assessors and industry stakeholders with robust, speedy, lower-cost, and informative next-generation hazard and risk assessment tools. PMID:38742231 | PMC:PMC11089904 | DOI:10.3389/ftox.2024.1359507 Sci Total Environ. 2024 Feb 20;912:168738. doi: 10.1016/j.scitotenv.2023.168738. Epub 2023 Nov 27. ABSTRACT Per- and polyfluoroalkyl substances (PFAS) are ubiquitously distributed in the aquatic environment. They include persistent, mobile, bioaccumulative, and toxic chemicals and it is therefore critical to increase our understanding on their adsorption, distribution, metabolism, excretion (ADME). The current study focused on uptake of seven emerging PFAS in zebrafish (Danio rerio) and their potential maternal transfer. In addition, we aimed at increasing our understanding on mixture effects on ADME by developing a physiologically based kinetic (PBK) model capable of handling co-exposure scenarios of any number of chemicals. All studied chemicals were taken up in the fish to varying degrees, whereas only perfluorononanoate (PFNA) and perfluorooctanoate (PFOA) were quantified in all analysed tissues. Perfluorooctane sulfonamide (FOSA) was measured at concerningly high concentrations in the brain (Cmax over 15 μg/g) but also in the liver and ovaries. All studied PFAS were maternally transferred to the eggs, with FOSA and 6:2 perfluorooctane sulfonate (6,2 FTSA) showing significant (p < 0.02) signs of elimination from the embryos during the first 6 days of development, while perfluorobutane sulfonate (PFBS), PFNA, and perfluorohexane sulfonate (PFHxS) were not eliminated in embryos during this time-frame. The mixture PBK model resulted in >85 % of predictions within a 10-fold error and 60 % of predictions within a 3-fold error. At studied levels of PFAS exposure, competitive binding was not a critical factor for PFAS kinetics. Gill surface pH influenced uptake for some carboxylates but not the sulfonates. The developed PBK model provides an important tool in understanding kinetics under complex mixture scenarios and this use of New Approach Methodologies (NAMs) is critical in future risk assessment of chemicals and early warning systems. PMID:38030006 | DOI:10.1016/j.scitotenv.2023.168738 Arch Toxicol. 2023 Dec;97(12):3075-3083. doi: 10.1007/s00204-023-03601-5. Epub 2023 Sep 27. ABSTRACT In Registration, Evaluation, Authorisation and Restriction of Chemicals (REACH) the criterion for deciding the studies that must be performed is the annual tonnage of the chemical manufactured or imported into the EU. The annual tonnage may be considered as a surrogate for levels of human exposure but this does not take into account the physico-chemical properties and use patterns that determine exposure. Chemicals are classified using data from REACH under areas of health concern covering effects on the skin and eye; sensitisation; acute, repeated and prolonged systemic exposure; effects on genetic material; carcinogenicity; and reproduction and development. We analysed the mandated study lists under REACH for each annual tonnage band in terms of the information they provide on each of the areas of health concern. Using the European Chemicals Agency (ECHA) REACH Registration data base of over 20,000 registered substances, we found that only 19% of registered substances have datasets on all areas of health concern. Information limited to acute exposure, sensitisation and genotoxicity was found for 62%. The analysis highlighted the shortfall of information mandated for substances in the lower tonnage bands. Deploying New Approach Methodologies (NAMs) at this lower tonnage band to assess health concerns which are currently not covered by REACH, such as repeat and extended exposure and carcinogenicity, would provide additional information and would be a way for registrants and regulators to gain experience in the use of NAMs. There are currently projects in Europe aiming to develop NAM-based assessment frameworks and they could find their first use in assessing low tonnage chemicals once confidence has been gained by their evaluation with data rich chemicals. PMID:37755502 | PMC:PMC10567824 | DOI:10.1007/s00204-023-03601-5 Regul Toxicol Pharmacol. 2022 Dec;136:105278. doi: 10.1016/j.yrtph.2022.105278. Epub 2022 Oct 21. ABSTRACT The Registration, Evaluation, Authorisation and Restriction of Chemicals (REACH) regulation was created to protect human health and the environment through the better and earlier identification of harmful intrinsic properties of chemical substances on the European market. One of its central aims was the promotion of alternatives to animal testing, yet it has instead become a long tick-box list of in vivo experiments questionable relevance to human health outcomes despite a global trend towards new approach methods (NAMs) in chemical safety assessment. The Chemicals Strategy for Sustainability (CSS), proposed by the European Commission in 2020, is a golden opportunity to revise REACH in a significant and impactful way, yet proposals presented so far have significant negative animal welfare consequences. There is still time to correct the course of the ongoing REACH revision - proposals made herein offer a path towards the promising future intended by the CSS. These proposals are anchored in three vectors of action, varying in level of complexity - from changes that ECHA can implement to improve existing processes, through technical changes aimed at minimizing animal testing and increasing NAM acceptance, to deeper structural changes to establish non-animal testing strategies as the basis for risk assessment. PMID:36280152 | DOI:10.1016/j.yrtph.2022.105278 Regul Toxicol Pharmacol. 2022 Nov;135:105261. doi: 10.1016/j.yrtph.2022.105261. Epub 2022 Sep 11. ABSTRACT New Approach Methodologies (NAMs) are considered to include any in vitro, in silico or chemistry-based method, as well as the strategies to implement them, that may provide information that could inform chemical safety assessment. Current chemical legislation in the European Union is limited in its acceptance of the widespread use of NAMs. The European Partnership for Alternative Approaches to Animal Testing (EPAA) therefore convened a 'Deep Dive Workshop' to explore the use of NAMs in chemical safety assessment, the aim of which was to support regulatory decisions, whilst intending to protect human health. The workshop recognised that NAMs are currently used in many industrial sectors, with some considered as fit for regulatory purpose. Moreover, the workshop identified key discussion points that can be addressed to increase the use and regulatory acceptance of NAMs. These are based on the changes needed in frameworks for regulatory requirements and the essential needs in education, training and greater stakeholder engagement as well the gaps in the scientific basis of NAMs. PMID:36103951 | DOI:10.1016/j.yrtph.2022.105261
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