In 2021 the European Food Safety Authority (EFSA) concluded that "A concern for genotoxicity of TiO2 particles that may be present in E 171 could therefore not be ruled out.". A detailed review of the genotoxicity of titanium dioxide (TiO) was subsequently published by Kirkland et al. (2022) using a comprehensive weight of evidence (WoE) approach in which test systems and endpoints were allocated different levels of relevance. At that time only 34 publications met the reliability and quality criteria for being most relevant in the evaluation of genotoxicity, and based on these it was concluded that the existing evidence did not support a direct DNA damaging mechanism for TiO. Recently a number of regulatory opinions have been published, in which papers were cited that described in vitro DNA damage (mainly comet), mode of action, and cellular uptake studies that were not discussed in Kirkland et al. (2022). Furthermore, a number of additional papers have been published recently or have been identified from the regulatory opinions as a result of using extended search criteria. A total of 70 publications not previously reviewed in Kirkland et al. (2022) have been reviewed here, and again show that the published data on the genotoxicity of TiO are inconsistent, often of poor quality, and in some cases difficult to interpret. The cellular uptake studies show some evidence of cytoplasmic uptake, particularly in cells treated in vitro, but there is no convincing evidence of nuclear uptake. In terms of genotoxicity, the conclusions of Kirkland et al. (2022) that existing evidence does not support a direct DNA damaging mechanism for titanium dioxide (including nano forms), and that the main mechanism leading to TiO genotoxicity is most likely indirect damage to DNA through generation of reactive oxygen species (ROS), are still valid.

Malaria is an acute, debilitating parasitic illness. There were 249 million cases of malaria in 2022, resulting in 608,000 deaths globally, 76% of which were children ≤5 years. The unique nature of this disease (recurrences leading to re-treatments and numerous organ systems affected), specific clinical treatment regimens, poor compliance, and diversity of affected populations (predominantly pediatrics, women of childbearing potential, pregnant and lactating women), often makes standard testing approaches inadequate, and tailor-made safety assessments are more appropriate. We provide best practice recommendations based on company experience for the non-clinical safety assessment of antimalarial drugs, with a focus on small molecules since they represent the majority of drug candidates for this illness. We focus on specific testing considerations for repeat dose toxicity studies, including combination toxicity assessments, since new drug treatment regimens typically foresee short treatment durations to improve compliance (i.e., 1 day) with combinations of compounds to improve efficacy and limit potential resistance. Due to the target population, the timing of reproductive, developmental, and juvenile toxicity studies may be earlier than general testing roadmaps for other small molecule drugs. In conclusion, key recommendations presented should enable a more effective and efficient development path whilst protecting clinical trial participants and patients.

Endocrine-disrupting chemicals (EDCs), including substances used in plant protection products (PPPs), are a source of ongoing concern for the EU society. Under the EC Regulation 1107/2009, the endocrine-disrupting (ED) properties of active substances, safeners, and synergists used in PPPs shall be investigated. The scientific criteria established by the Regulation (EU) 2018/605 and the joint guidance of the European Chemicals Agency (ECHA)/European Food Safety Authority (EFSA) provide the basis for this assessment. Data requirements for the approval of safeners and synergists have been recently published in Commission Regulation (EU) 2024/1487, allowing a consistent assessment of these substances. The approach to assessing co-formulant hazards is currently a subject of EU-wide discussion. It outlines the necessity to take into account information or evaluation data from other than pesticides' EU regulatory frameworks, such as REACH or SCCS applications for cosmetic ingredients. This paper outlines: a) current EU approach applied for identification of endocrine disrupting properties of pesticides; b) issues related to European regulations that may have an indirect impact on the safe use of plant protection products and c) an analysis of the European Commission's activities aimed to limit exposure to EDCs associated with use of PPPs in the society.

In response to the current disparity in risk assessment values for PFOA from different agencies and countries, an international effort facilitated by the Alliance for Risk Assessment (ARA) was recently undertaken to characterize the range of scientifically supportable safe dose estimates. In this assessment (Burgoon et al., 2023), an evaluation of the evidence regarding the potential modes of action (MOA) for PFOA toxicity was performed first, so that it could be used to inform subsequent decisions regarding potential critical effects and studies. This review describes the evidence considered in the MOA evaluations that were performed as part of the ARA effort. The overall conclusions of this evaluation are that the available mechanistic data do not support any conclusion that reported epidemiological associations of blood concentrations of PFOA as low as 10 ng/mL with various health effects should be considered causal. It is more likely that the reported associations may instead reflect reverse causality/pharmacokinetic confounding. These conclusions are consistent with the opinions of the World Health Organization (WHO, 2022).

Consumers may be exposed via foods to a diverse range of substances that could be considered as contaminants. However, it is not always straightforward to understand the definition of a 'contaminant'. The present review evaluates how various categories of food-relevant substances are considered in terms of being 'contaminants'. To this end these categories of food borne constituents are evaluated against the various criteria encountered in the available definitions of a food contaminant, including unintentional presence, harmful, existence of regulatory limits, and stakeholder perception. The categories of chemicals considered include: phytotoxins, mycotoxins, (heavy) metals, persistent organic pollutants (POPs), processing aids, process related contaminants, food contact materials (FCMs), pesticides and veterinary drugs. The evaluation revealed that usage of the term appears complex, and may differ between stakeholders. A common proposed definition of the term 'contaminant' could be 'a substance considered to require control measures due to the unacceptability of its context within a food'. Use of a dimension of harm results in equivocal outcomes because risk depends on the level of exposure. As the term 'contaminant' has influence on risk management including public policy, the motivations for applying the term should be subject to more detailed analysis and understanding.

Improving regulatory confidence and acceptance of in silico toxicology methods for chemical risk assessment requires assessment of associated uncertainties. Therefore, there is a need to identify and systematically categorize sources of uncertainty relevant to the methods and their predictions. In the present study, we analyzed studies that have characterized sources of uncertainty across commonly applied in silico toxicology methods. Our study reveals variations in the kind and number of uncertainty sources these studies cover. Additionally, the studies use different terminologies to describe similar sources of uncertainty; consequently, a majority of the sources considerably overlap. Building on an existing framework, we developed a new uncertainty categorization framework that systematically consolidates and categorizes the different uncertainty sources described in the analyzed studies. We then illustrate the importance of the developed framework through a case study involving QSAR prediction of the toxicity of five compounds, as well as compare it with the QSAR Assessment Framework (QAF). The framework can provide a structured (and potentially more transparent) understanding of where the uncertainties reside within in silico toxicology models and model predictions, thus promoting critical reflection on appropriate strategies to address the uncertainties.

Pharmaceutical and personal care products, including syrups and toothpastes, extensively use glycerin, sorbitol, and propylene glycol. However, past incidents of ethylene glycol (EG) and diethylene glycol (DEG) contamination in these products have raised serious health concerns. Recently, several child deaths linked to contaminated cough syrup consumption have heightened concerns regarding the safety of Indian pharmaceuticals. In response, Indian drug regulatory authorities and the Indian Pharmacopoeia have implemented several measures to enhance the quality, safety, and efficacy of pharmaceuticals manufactured in India. These measures encompass risk-based inspections of manufacturing facilities, rigorous quality control checks of medicinal products intended for export, and increased transparency in the supply chain of excipients prone to EG and DEG contamination. Further, the Indian Pharmacopoeia has updated monographs for five high-risk excipients: glycerin, propylene glycol, sorbitol solution (70%, both crystallizing and non-crystallizing), and liquid maltitol. These efforts are consistent with global regulatory standards and aim to ensure the overall quality and safety of pharmaceuticals produced in India.

Plant protection products (PPPs) undergo rigorous safety assessments. In Europe, non-dietary risk assessments for operators, workers, bystanders and residents are highly conservative as this area of exposure science has historically been data poor. CropLife Europe (CLE) companies have collaborated to generate new data and pool existing data to refine the approaches prescribed by the European Food Safety Authority (EFSA) guidance on non-dietary exposure (2022). This article summarises key activities, beginning with the development of the Agricultural Operator Exposure Model (AOEM) and covers projects which refine current approaches to bystander, resident and re-entry worker assessment, including the Bystander Resident Orchards Vineyards (BROV) project, improvements to the Bystander and Resident Exposure Assessment Model for spray drift (BREAM), proposals for refined vapour inhalation assessments, and a meta-analysis of Dislodgeable Foliar Residue (DFR) data. A study quantifying the benefits of using closed transfer systems, an appraisal of the inherent compounded conservatism in current risk assessment paradigms and the development of a new seed treatment model by the SeedTROPEX taskforce are also introduced. These industry-led activities underscore the critical role of non-dietary exposure in the registration process for PPPs and reflect an ongoing commitment to provide farmers with effective crop protection solutions while ensuring safety.

The landscape of drug product development and regulatory sciences is evolving, driven by the increasing application of systems thinking and modeling and simulation (M&S) techniques, especially from a biopharmaceutics perspective. Patient-centric quality standards can be achieved within this context through the application of quality by design (QbD) principles and M&S, specifically by defining clinically relevant dissolution specifications (CRDS). To this end, it is essential to bridge in vitro results to drug product in vivo performance, emphasizing the need to explore the translational capacity of biopharmaceutics tools. Physiologically based M&S analyses offer a unique avenue for integrating the drug, drug product, and biological properties of a target organism to study their interactions on the pharmacokinetic response. Accordingly, Physiologically Based Biopharmaceutics Modeling (PBBM) has seen increasing use to support drug development and regulatory applications globally. In Brazil, a Model-Informed Drug Development (MIDD) policy and strategic project are not yet established, limiting applicability of M&S techniques. Drawing from the experience of the ANVISA-Academia PBBM Working Group (WG), this article assesses the opportunities and challenges for pharmacometrics (PMx) in Brazil and proposes strategies to advance the adoption of M&S analyses into regulatory decision-making.

HSK21542 is a peripherally-restricted kappa opioid receptor (KOR) agonist developed for pain treatment. Because of the CNS pharmacological concern of opioid receptor activation, such as physical dependence and addiction potential, an assessment of abuse potential of HSK21542 was required prior to marketing approval. The preclinical abuse potential assessments for HSK21542 included the following studies: 1) intravenous self-administration study to explore the relative reinforcing efficacy in rats self-administering remifentanil; 2) rat drug discrimination study to examine the pharmacological similarity of the interoceptive or subjective effects of HSK21542 in rats discriminating pentazocine; 3) rat conditioned place preference (CPP) paradigm to test the rewarding effects; 4) rat natural physical dependence-spontaneous withdrawal study in rats chronically treated with HSK21542; 5) naloxone-precipitated withdrawal assay following chronic HSK21542 exposure to evaluate its physical dependence potential. The results showed that HSK21542 was devoid of behavioral evidence of positive reinforcing effect and did not share similar discriminative stimulus effects with pentazocine. HSK21542 also did not produce CPP in rats. In addition, HSK21542 did not produce spontaneous withdrawal or naloxone-precipitated withdrawal in rats with chronic treatments. Collectively, these preclinical findings suggest that HSK21542 has no abuse potential in animals, which demonstrate low abuse potential in humans.

Vanadium is used in alloys, batteries as well as catalyst and is a known impurity in medical devices and pharmaceuticals. The present work describes the calculation of a parenteral tolerable intake (TI) for vanadium by chronic exposure in implantable medical devices per ISO 10993-17:2023, the applicable standard. The 2023 update of ISO 10993-17 [1] introduces new uncertainty factors (UFs) for calculating a TI. Therefore, we noted differences between the ISO guidance and the ICH Q3D guidance on Permissible Daily Exposure (PDE) for parental elemental pharmaceutical impurities. We derived a TI of 0.20 μg V/kg/day based on the updated ISO guidance, and a PDE of 0.24 μg V/kg/day based on ICH guidance. The latter is considered a more realistic estimate.

In this paper we present methodological and experimental details and results from an OECD Test Guideline 474 and GLP-compliant in vivo micronucleus study on sodium molybdate dihydrate in Sprague Dawley rats. Prior to the conduct of this study, there was a data-gap for reliable in vivo genotoxicity data for molybdenum substances. The presentation of the new study is complemented by a review of other available in vitro and in vivo data on the genotoxicity of molybdenum substances, focussing on substances where the contained or released molybdate ion, MoO, is considered the responsible moiety for any toxicological effect (grouping/category approach). After consideration of the relevance and reliability of all available data, the absence of a concern for genotoxicity of molybdate in vitro and in vivo is concluded.

The concurrent control group is the most important reference for the interpretation of toxicity studies. However, pooled information on control animals from independent studies, i.e., historical control data (HCD), is also used for the interpretation of results. Currently, an overview on actual HCD use in regulatory toxicology is lacking. Therefore, we evaluated the HCD use of the Joint FAO/WHO Meeting on Pesticide Residues from 2004 to 2021 and compared it with recommendations in regulatory guidelines and in the literature. We found that HCD was used routinely and exclusively to avoid potential false positive decisions regarding the treatment-relatedness of effects, mostly using the HCD range, i.e., the most extreme values, as a benchmark. HCD were not used to avoid potential false negative decisions or for quality control of the index study. The central assumption of the HCD use, namely that the HCD and control group of the index study follow the same underlying distribution because they are samples of the same data generation process, was not investigated, although numerous factors potentially contribute to effect variation between the different control groups pooled in the HCD. We recommend that the existing guidelines be revised to improve the robustness and transparency of toxicological assessments.

In November 2023, the International Agency for Research on Cancer (IARC) classified PFOA as "carcinogenic to humans" (Group 1) and PFOS as "possibly carcinogenic to humans" (Group 2B). We evaluated these classifications, considering the epidemiology, experimental animal, and mechanistic evidence. It is our opinion that the IARC Working Group overstated the available evidence for the carcinogenicity of PFOA and PFOS. Epidemiology studies have shown weak and inconsistent associations across studies. Studies reporting increased incidences of tumors in experimental animals exposed to PFOA or PFOS had statistically significant results that were driven by the presence of benign adenomas. The IARC Working Group used the key characteristics of carcinogens (KCCs, which comprise 10 chemical and/or biological properties of known human carcinogens) approach to upgrade the carcinogenicity classifications for PFOA and PFOS from initially lower classifications that were based on the strength of the epidemiology and experimental animal evidence. However, this is not a robust assessment of mechanistic evidence, as it fails to consider the quality, external validity, and relevance of the evidence. Rather than use the KCCs as a checklist of potential carcinogenic mechanisms, IARC should use a rigorous method to evaluate the plausibility and human relevance of mechanistic evidence.

Nitrosamine drug substance-related impurities (NDSRIs) are a sub-category of N-nitrosamine drug impurities that share structural similarity to the corresponding active pharmaceutical ingredient. The mutagenicity of NDSRIs is poorly understood. We previously tested a series of NDSRIs using the Enhanced Ames Test (EAT). In this follow-up study, we further examined the genotoxicity and mutagenicity of 15 of these NDSRIs in human TK6 cells. Seven EAT-positive NDSRIs, including N-nitroso-nortriptyline, N-nitroso-fluoxetine, N-nitroso-desmethyl-diphenhydramine, N-nitroso-duloxetine, N-nitroso-lorcaserin, N-nitroso-varenicline, and N-nitroso-sertraline, induced concentration-dependent increases in micronuclei after bioactivation with hamster liver S9. These NDSRIs were also mutagenic in the TK and HPRT gene mutation assays, consistent with their positive EAT results. In the presence of hamster liver S9, the eight EAT-negative NDSRIs were negative in the micronucleus assay and negative for mutation induction. Using TK6 cells endogenously expressing a single human cytochrome P450 (CYP), we found that CYP2C19, CYP2B6, CYP2A6, and CYP3A4 are key enzymes activating the genotoxicity and mutagenicity of these NDSRIs. Overall, the hamster S9-mediated TK6 cell mutagenicity results agreed with those observed in the EAT, indicating consistency in the mutagenic responses produced by NDSRIs across different testing systems. These data support the use of EAT for hazard identification and safety assessment of NDSRIs.

Chemical characterization of medical devices uses the analytical evaluation threshold (AET) to determine reportable organic extractables, as these chemicals may be of toxicological concern and should be addressed via toxicological risk assessment. The AET is not applicable to metal extractables due to the exclusion of toxicity data on inorganics from the dataset used to derive dose-based threshold (DBT) values. This results in minimal guidance for reporting metal extractables. Herein, an AET for metals, or mAET, is proposed as a reporting threshold for individual metal extractables. The mAET can ensure metals are reported that are at quantities that may present a patient safety risk. This may reduce the number of metals reported in a chemical characterization report, improving the efficiency of the overall biocompatibility evaluation by removing unneeded effort and resource time. Conversely, an analytical method's ability to report all metals at toxicologically relevant levels can be confirmed by comparing method sensitivity to mAET values. DBTs were developed for 70 metals, permitting mAET values to be determined. These mAET values were then compared to metal reporting limits from 13 previously conducted chemical characterization studies, which used varying extraction designs and analytical methods, to determine the impact of the mAET.

The Ames test is a widely used bacterial mutagenicity assay to evaluate the potential of chemical compounds to induce mutations. In recent years, there has been growing concern regarding the presence of N-nitrosamines in pharmaceuticals, food, and other consumer products. N-Nitrosamines are probable mutagens and carcinogens. To address the reduced sensitivity of the standard Ames test for N-nitrosamines, particularly N-nitrosodimethylamine, the European Medicines Agency (EMA) and United States Food and Drug Administration (FDA) have recently published recommendations for enhanced Ames test (EAT) conditions. However, there is a lack of clear guidance on the selection of N-nitrosamine positive control concentrations, particularly for 1-cyclopentyl-4-nitrosopiperazine, and the amount of solvent to be used in the EAT. This study aims to address the current gap in concentration and volume specifications by providing a comprehensive guide to set up enhanced Ames test conditions specifically for N-nitrosamine compounds using appropriate amounts of solvent, new solvents, and strain-specific positive control concentrations.

Utilization of data from historical control animals to form virtual control groups (VCGs) is an innovative approach to embody the 3Rs (reduce, refine, and replace use of control animals) principle in research. However, there is no available systematic comparison of statistical performance between concurrent control groups (CCGs) and VCGs in nonrodent safety assessment. The optimal selection criteria and combination of VCGs and CCGs also remain unclear. This study retrospectively evaluated VCGs' statistical performance to detect test article effects on body weight and clinical pathology endpoints in dog and nonhuman primate (NHP) systemic toxicity studies. Body weight and six clinical pathology endpoints were analyzed against the reported study findings from a cohort of 22 previously reported nonrodent 1-month oral gavage toxicity using three different methods of generating VCGs. When the fold change from baseline was used, VCGs yielded a similar or higher statistical sensitivity to detect test article relatedness than CCGs. Compared to simple random sampling or using fixed criteria, the propensity score matching by BW, age, and year of study initiation yielded higher sensitivities. Our analysis supports the hypothesis that VCGs can be a viable instrument in nonrodent toxicity studies.

Although uncertainties expressed in texts within QSAR studies can guide quantitative uncertainty estimations, they are often overlooked during uncertainty analysis. Using neurotoxicity as an example, this study developed a method to support analysis of implicitly and explicitly expressed uncertainties in QSAR modeling studies. Text content analysis was employed to identify implicit and explicit uncertainty indicators, whereafter uncertainties within the indicator-containing sentences were identified and systematically categorized according to 20 uncertainty sources. Our results show that implicit uncertainty was more frequent within most uncertainty sources (13/20), while explicit uncertainty was more frequent in only three sources, indicating that uncertainty is predominantly expressed implicitly in the field. The most highly cited sources included Mechanistic plausibility, Model relevance and Model performance, suggesting they constitute sources of most concern. The fact that other sources like Data balance were not mentioned, although it is recognized in the broader QSAR literature as an area of concern, demonstrates that the output from the type of analysis conducted here must be interpreted in the context of the broader QSAR literature before conclusions are drawn. Overall, the method established here can be applied in other QSAR modeling contexts and ultimately guide efforts targeted towards addressing the identified uncertainty sources.

Minipigs are valid nonrodent species infrequently utilized for pharmaceutical research and development (R&D) compared with dogs or nonhuman primates (NHPs). A 2022 IQ DruSafe survey revealed a modest increase in minipig use by pharmaceutical companies compared with a prior 2014 survey, primarily in the development of oral small molecules and parenteral protein molecules. Some companies considered using minipigs more often due to NHP shortages and regional ethical concerns with using NHPs and dogs. However, for most pharmaceutical companies, minipigs still represent ≤5% of their nonrodent animal use. Key challenges noted by companies to wider adoption of minipigs were high test article requirement, limited historical control data, and lack of relevant reagents or assays. Additionally, some companies expressed uncertainties about contract research organization (CRO) capabilities and experience, a perception not shared by respondent CROs. These latest survey results indicate persistence of many concerns previously identified in 2014. Several case studies are included to illustrate areas of expanded minipig use as well as the challenges that hinder broader adoption. Ongoing, focused, and industry-wide initiatives to address the identified or perceived challenges may lead to more frequent or routine consideration of minipigs as a test species in pharmaceutical R&D.