NAM journal最新文献

Traditional ecotoxicity testing relies heavily on vertebrate animal models, which are resource-intensive, low-throughput, and raise ethical concerns under the principle of the 3Rs (Reduction, Refinement, Replacement). New Approach Methodologies (NAMs), including in silico, in chemico, in vitro and ex vivo, offer promising alternatives for environmental chemical hazard assessment. While NAMs have gained traction in human health risk assessment, their regulatory acceptance for environmental safety remains limited due to technical, scientific and policy barriers. This paper presents an overview of a literature review related to the status, challenges and future strategies for broader adoption of NAMs for environmental safety assessment of chemicals. The review´s objectives were to: (1) systematically review the current state of NAMs for aquatic toxicity assessment, (2) identify gaps in NAM readiness and endpoint coverage, and (3) provide recommendations to accelerate regulatory acceptance and integration of NAMs into environmental safety frameworks. A systematic literature review was conducted using PubMed followed by the SWIFT-Active Screener tool, focusing on NAMs applicable to aquatic organisms (fish, amphibians, marine mammals). The review excluded terrestrial and invertebrate models. Search terms considered within the scope of the review included in silico, in chemico, in vitro and ex vivo. While embryo assays may not be considered a NAM by all geographical regions and stakeholders, information from existing embryo assays was also reviewed to reduce the reliance on protected vertebrate life stages. The screening identified 4275 unique articles and 1310 were selected for full-text review. A database containing all the reviewed information is freely available and contains 18 metadata fields including for example the captured assay type, species, endpoints, and readiness level. This review provides a comprehensive roadmap for transitioning toward animal-free environmental safety assessment. By addressing gaps in readiness, standardization, and mechanistic integration, NAMs can become central to next-generation risk assessment, supporting global initiatives to phase out vertebrate testing.

Pharmaceutical scientists develop and apply increasingly complex technologies from an expanding toolbox to create more effective and safer drugs. Consequently, the increasing human specificity of new modalities has highlighted the limitations of conventional animal studies. Recently, new approach methodologies (NAMs) have attracted attention for their potential to promote the 3Rs and human-relevant evaluation in pharmaceutical research and development, and have been applied not only in early discovery research but also in regulatory review. In April 2025, the US Food and Drug Administration released a roadmap outlining a stepwise transition from animal studies to NAMs and highlighting drug development tool qualification, raising the urgency for a coordinated response in Japan. The Japan Pharmaceutical Manufacturers Association (JPMA) conducted a survey of JPMA member companies in July 2025 to provide an overview of the status of NAMs in the Japanese pharmaceutical industry and to generate evidence for policy recommendations. While most responding companies were aware of the FDA roadmap, concrete internal responses were limited, and approximately half had not conducted NAMs research. The results indicate strong interest in promoting the 3Rs and improving non-clinical development, but limited experience with NAMs, especially for regulatory applications. Both internationally harmonized guidelines and the establishment of a domestic qualification system, including early consultation channels and a transparent public database, are required to promote the regulatory use of NAMs in Japan. The JPMA could serve as a central hub by facilitating proactive communication among domestic and international stakeholders, thereby promoting the broader implementation of NAMs in regulatory submissions.

Reliable, mechanism-based in vitro assays are essential for Next Generation Risk Assessment of non-genotoxic carcinogens, including those operating through the induction of oxidative stress. This study evaluated how oxygen tension, metabolic capacity and exposure scenarios influence oxidative stress quantification in human hepatocytes, aiming to support the development of in vitro assays relevant for chronic toxicity. We compared chemically-induced oxidative stress in HepG2 cells under atmospheric (20% O₂) and physiological (3% O₂) conditions by measuring reactive oxygen species (ROS) and the ratio between reduced and oxidized glutathione (GSH/GSSG). Lower oxygen tension reduced cell clustering, slowed proliferation, and mildly decreased hepatocyte marker expression and cytochrome P450 activity. Notably, the potency to induce ROS of most chemicals was lower at 3% oxygen, while the potency to perturb GSH/GSSG was chemical dependent. Maturation of HepG2 cells (mHepG2) yielded a more hepatocyte-like phenotype, with increased expression of metabolic genes and cytochrome P450 activity. Six of the eight tested chemicals showed greater susceptibility to chemically-induced ROS in mHepG2 cells. However, mHepG2 cells had a seemingly better recovery capacity based on the chemical's potencies to perturb GSH/GSSG. Importantly, mHepG2 cells can be cultured over extended periods, allowing to study the effects of repeated exposure. Repeated exposure with or without recovery periods resulted in increased potency to induce oxidative stress. These findings highlight the importance of oxygen tension, metabolic competence, and exposure frequency for hazard characterization of oxidative stress-inducing chemicals, supporting the development of more predictive in vitro assays for chronic toxicity.

The validation of in vitro New Approach Methodologies (NAMs) requires the use of well-characterized reference chemicals to assess assay performance, reproducibility, and relevance. However, selecting such chemicals is often labor-intensive and lacks standardization. We developed a semi-automated, evidence-based workflow that integrates systematic literature review, AI-assisted data extraction, and quantitative evidence scoring to improve the efficiency and transparency of chemical selection. We conducted a structured EMBASE search, followed by AI-assisted abstract screening and data extraction, in a case study to select reference chemicals for the validation of an in vitro adipogenesis assay. Chemicals were evaluated for in vitro, in vivo, and human evidence of adipogenic or obesogenic effects. From 11,648 screened publications, 236 studies met the inclusion criteria, resulting in the identification of 243 candidate reference chemicals, of which 50 were prioritized based on scoring. The final selection encompassed a set of 22 chemicals with different levels of potency and diverse structures. This workflow illustrates how AI-assisted evidence synthesis can accelerate and standardize the selection of reference chemicals while maintaining expert oversight and ensuring regulatory relevance. It provides a reproducible and adaptable framework for future NAMs validation studies across diverse toxicological endpoints.

The concept of benchmark materials in the personal care and cosmetic industry is often based on the positioning of prototypes relative to the leading brand within a given product category. The main points of comparison are the market ranking, performance regarding consumers' perception, aesthetics, cost, complexity of the manufacturing process and patents covering the product and/or its components. Even though benchmark materials are often included when conducting safety tests of cosmetic and personal care products, the concept is rather loosely defined in toxicological profiling. The use of benchmarks becomes of particular interest when safety testing is based on New Approach Methodologies (NAMs) (in silico, in vitro or ex vivo technologies) that allow for toxicity ranking of the prototypes and provide mechanistic insights without having an established prediction model. It is therefore important to define the properties and performance of the benchmark materials to better guide the safety testing of candidate formulations. In this review manuscript we discuss the criteria to select benchmarks, and provide specific examples of such finished products incorporated in the safety assessment studies of a wide range of cosmetic and personal care product classes. We also discuss strategies using benchmarks to evaluate the predictive capacity of test systems and methodologies that do not rely on the use of animals for safety assessments.

Mechanistically grounded, human-relevant new approach methodologies (NAMs) are needed to support AOP-informed, animal-free strategies for kidney safety assessment and to improve the prediction of nephrotoxic risk. Here, we developed a mechanistically anchored in vitro assay battery to quantify functional readouts aligned with key events (KEs) central to kidney adverse outcome pathways (AOPs). Using human conditionally immortalised proximal tubule epithelial cells (ciPTECs), we assessed mitochondrial depolarisation, reactive oxygen species (ROS) generation, lysosomal enzyme release, membrane integrity, and cell viability in response to cisplatin, carboplatin, and oxaliplatin. All three compounds produced concentration- and time-dependent effects consistent with their clinical nephrotoxic profiles. Mitochondrial depolarisation emerged as the most sensitive and consistent key event (KE) in ciPTECs, while ROS generation was compound-dependent, robust for cisplatin, moderate for oxaliplatin, and minimal for carboplatin. Later endpoints, including increased β‑N‑acetylglucosaminidase (NAG) activity, lactate dehydrogenase (LDH) release, and loss of cell viability, required higher concentrations to reach significance. These findings identify an apparent sensitivity order among KE-aligned endpoints and provide quantitative data consistent with established mechanisms of nephrotoxicity. These data provide quantitative, KE-aligned response profiles that may support future key event relationship (KER) and quantitative (qAOP) development and demonstrate the utility of an AOP-informed assay battery for mechanistic screening and prioritisation within a focused platinum nephrotoxicity case study.

Transcriptomic points of departure (tPODs) from in vitro studies are being explored as potential alternatives to animal studies. This study aimed to derive tPODs for 19 pesticides of concern using two human cell lines relevant to oral exposure, Caco-2 and HepG2, and to compare the results with in vivo toxicity data and regulatory thresholds. tPODs were successfully obtained for all pesticides, ranging from 0.032 to 880 µM. Pathway analysis indicated more stress-related responses in Caco-2 cells, whereas more metabolic pathways were identified in HepG2 cells. Following mass balance modeling and in vitro-to-in vivo extrapolation, tPODs were converted to human administered equivalent doses (t-AEDs). Approximately 50% and 70% of t-AEDs derived from Caco-2 and HepG2 cells, respectively, were within one order of magnitude of corresponding PODs from chronic animal studies, suggesting moderate concordance. The overall results suggest that tPODs from short-term in vitro studies may provide conservative estimates relative to apical endpoints from animal studies. Overall, this study demonstrates the potential of in vitro transcriptomics to generate quantitative and mechanistic data to support chemical screening and prioritization activities.

Developmental exposure to environmental toxicants is a cause of skeletal abnormalities. Yet the molecular mechanisms linking early exposure to impaired bone formation remain undefined. Skeletal tissues arise from both neural crest- and mesoderm-derived lineages that rely on shared osteogenic differentiation programs, suggesting that disruption of common regulatory processes may contribute to diverse skeletal outcomes. MicroRNAs (miRNAs) are key post-transcriptional regulators of gene networks and have appeared as indicators of toxicant-induced perturbation. In this study, we examined whether developmentally relevant toxicants are associated with miRNA regulatory networks during osteogenic differentiation. Using a human embryonic stem cell (hESC)-based osteogenic differentiation model, we assessed the effects of nine toxicants spanning distinct primary mechanisms. Toxicant exposure impaired osteogenic differentiation at their IC50, as reflected by altered expression of osteogenic markers and transcriptional remodeling. Global miRNA profiling revealed dysregulation of miRNAs enriched for bone-related biological processes, including regulators of osteogenic commitment and differentiation timing. Integrated miRNA-mRNA network analysis identified a subset of miRNAs linked to core osteogenic and lineage-associated pathways, including RUNX2-dependent transcription and BMP and Wnt signaling. Modulation of representative miRNAs produced osteogenic outcomes consistent with those observed following toxicant exposure and, in some cases, was associated with partial restoration of differentiation in exposed cultures. Collectively, these findings indicate that chemically diverse developmental toxicants are associated with miRNA-mediated regulatory patterns during osteogenic differentiation. Identification of shared miRNA regulatory features provides mechanistic insight into developmental bone toxicity and supports the use of miRNA network analysis as a human-relevant endpoint for skeletal hazard identification.

Predicting metabolism-dependent drug toxicity encompassing both the bioactivation and detoxification processes remains a critical challenge in preclinical safety evaluations. Conventional cardiomyocyte-only assays frequently misclassify compounds such as terfenadine, which require hepatic metabolism to attenuate cardiotoxicity. Current microphysiological systems (MPS) have practical limitations, such as insufficient hepatocyte oxygenation and complex external fluidic setups. We developed a liver-heart co-culture system that integrates three-dimensional human iPS cell-derived engineered heart tissues (hiPSC-EHTs) and cryopreserved primary human hepatocytes (PHHs), using a modified BioStellar™ Plate-a tubeless, stirrer-pump-integrated microfluidic device customized with an oxygen-permeable membrane. Using this system, we evaluated the cardiotoxicity of terfenadine in hiPSC-EHT monocultures and liver-heart co-cultures. Terfenadine exposure induced marked contractile dysfunction in hiPSC-EHT monocultures, whereas co-culturing with PHHs significantly attenuated this effect. Functional monitoring revealed time- and dose-dependent recovery, consistent with a largely reversible pharmacological mechanism. Drug concentration analysis by LC-MS/MS showed a marked increase in the formation of the primary metabolite fexofenadine, in the co-culture system, which was consistent with the reduced cardiotoxic potential of this metabolite. Gene expression analysis confirmed the presence of hepatic CYP3A4, which supports its predominant role in metabolic detoxification. This study demonstrated that a tubeless, stirrer-pump-integrated system with sustained hepatocyte function enables time-resolved evaluation of pharmacokinetic-pharmacodynamic relationships of metabolism-dependent cardiotoxicity on a single platform. This proof-of-concept approach suggests a potential improvement in the translational predictability of human-relevant, metabolism-dependent cardiotoxicity by enabling integrated evaluation of pharmacokinetic and pharmacodynamic responses. These findings highlight the utility of this system as a simple and scalable alternative to conventional perfusion-based MPS.

Environmental risk assessment (ERA) has not yet undergone the shift toward new approach methodologies (NAMs), that has reshaped human health safety assessment. This study adapts the 10-step read across next generation risk assessment (NGRA) framework developed for cosmetic safety, to the environmental domain and applies it to triclosan (TCS), given its data richness and environmental relevance. The framework consists of three tiers: Tier 0 for problem formulation and in silico screening; Tier 1 for bioavailability estimation and mode of action hypothesis formulation; and Tier 2 for refinement, hazard characterisation, risk quantification and uncertainty assessment. Four core principles are proposed for environmental NGRA: the assessments should (1) be exposure-anchored, (2) be mechanism-based, (3) have translatable thresholds, and (4) be both protective and transparent. Exposure was assessed using a two-tier use-based predicted environmental concentration (PEC) approach. The lower tier applied a conservative aggregate cosmetic exposure scenario, yielding a screening PEC of 0.74 µg/L. This value exceeded both the ecological threshold of toxicological concern (ecoTTC) and Tier 0 read across thresholds, triggering refinement using year specific exposure data. Integrated bioactivity, QIVIVE and conservation analysis identified lipid and steroid metabolism as the most sensitive pathways. Tier 2 yielded two NGRA predicted no effect concentrations (PNECs): a species sensitivity distribution (SSD)-based PNEC of 0.020 µg/L and a lowest environmental point of departure (ePoD)-based PNEC of 0.007 µg/L. Risk characterisation ratios (RCRs) were below 1 from 2019 onwards under both routes, supporting an environmental NGRA approach at least as protective as conventional ERA. While the proposed framework has been demonstrated for an insecticide, this study extends it to a cosmetic ingredient, broadening the transferability of the 10-step framework for NAM-based ERA.