Long-term efficacy of microwave ablation for multifocal papillary thyroid microcarcinoma: a 5-year follow-up study.

OBJECTIVES: Microwave ablation (MWA) has achieved excellent long-term efficacy in treating unifocal papillary thyroid microcarcinoma (UPTMC). The therapeutic effect of this treatment on multifocal papillary thyroid microcarcinoma (MPTMC) is unknown. Therefore, we evaluated the long-term efficacy of MWA for low-risk MPTMC and to provide evidence-based medicine for the revision of clinical guidelines. METHODS: This study included 66 MPTMC patients with a total of 158 lesions, all of whom received MWA. We collected and retrospectively analyzed the patients' follow-up data before MWA, at 1, 3, 6, and 12 months posttreatment and every 6 months thereafter until 5 years posttreatment. We evaluated the MWA complication rate, technical success rate (TSR), lesion volume reduction rate (VRR), and complete disappearance rate (CDR) during follow-up and in those patients with tumor progression and delayed surgery. RESULTS: After 60 months of follow-up, all 158 lesions disappeared in 66 patients, and the volume was reduced from 43.82 mm3 to 0.00 mm3. The TSR and VRR were both 100%. The CDRs at 1 year, 2 years, and 3 years were 57.59%, 93.67%, and 100%, respectively. The complication rate was 3.03% (2/66), and the incidence of tumor progression was 3.03% (2/66), including one new intrathyroidal lesion and one cervical lymph node metastasis (LNM). These lesions were retreated with MWA, and the lesions disappeared during the follow-up period. CONCLUSIONS: Ultrasound-guided MWA for low-risk MPTMC is safe and effective and may serve as an alternative option for patients who refuse surgery or active surveillance (AS). CLINICAL RELEVANCE STATEMENT: This study concludes that ultrasound-guided microwave ablation for low-risk multifocal papillary thyroid microcarcinoma is safe and effective and may serve as an alternative option for patients who refuse surgery or active surveillance. KEY POINTS: • Ultrasound-guided microwave ablation for low-risk multifocal papillary thyroid microcarcinoma is safe and effective. • During 5 years of follow-up, multifocal papillary thyroid microcarcinoma patients treated with microwave ablation had a favorable prognosis. • To provide evidence-based medicine for the revision of clinical guidelines.

Speckle suppression in holographic phase fringe patterns with different level noises based on FFDNet.

In this paper, an ANLVENet speckle suppression method in holographic phase fringe patterns with different level noises is proposed based on FFDNet, combined with asymmetric pyramid non-local block with a verge extraction module. The experimental results are compared to three network models and several representative algorithms. It is shown that the ANLVENet method not only has better superiority in the speckle suppression with different noise levels, but also preserves more details of the image edge. In addition, another speckle noise model is applied in the phase fringe patterns to prove the stronger generalization of the ANLVENet algorithm. The proposed method is suitable for suppressing the speckle with different levels in a large noise range under complex environmental conditions.

Value of CEUS combined with feeding artery ablation in the microwave ablation of large solid benign thyroid nodules.

OBJECTIVES: To investigate the value of contrast-enhanced ultrasound (CEUS) combined with feeding artery ablation in the microwave ablation (MWA) of large solid benign thyroid nodules (LSBTNs) with a diameter ≥ 4 cm. METHODS: We retrospectively analyzed 122 patients with LSBTN ≥ 4 cm in diameter treated with MWA. During evaluations before and after MWA, 53 patients who underwent conventional ultrasound examination were classified as the routine group, and 69 patients who underwent CEUS combined with feeding artery ablation were classified as the union group. The differences in ablation energy required per milliliter (AERPM), complication rate, regrowth rate, and volume reduction rate (VRR) were compared between the two groups. RESULTS: The AERPM of the union group was significantly less than that of the routine group (956.3 ± 38.5 J/mL vs. 1025.9 ± 121.5 J/mL, p < 0.001). The complication rate of the routine group was significantly higher than that of the union group (13.2% vs. 2.9%, p = 0.031). The regrowth rate of the routine group (22.6%, 12/53) was significantly higher than that of the union group (7.2%, 5/69) (p = 0.015). At the 1st, 3rd, 6th, 12th, 18th, and 24th month after ablation, the mean VRRs of the routine group were significantly less than those of the union group, with p values of < 0.001, < 0.001, 0.002, 0.007, 0.013, and < 0.001, respectively. CONCLUSIONS: The application of CEUS combined with feeding artery ablation in the MWA of LSBTNs is helpful to reduce the regrowth rate, improve the ablation efficiency, and reduce bleeding. KEY POINTS: • CEUS combined with feeding artery ablation in MWA of LSBTNs is helpful to reduce regrowth rate. • CEUS combined with feeding artery ablation can help improve ablation efficiency than conventional ultrasound in LSBTNs. • CEUS combined with feeding artery ablation helps reduce the incidence of bleeding during MWA.

Next generation risk assessment of human exposure to estrogens using safe comparator compound values based on in vitro bioactivity assays.

In next generation risk assessment (NGRA), the Dietary Comparator Ratio (DCR) can be used to assess the safety of chemical exposures to humans in a 3R compliant approach. The DCR compares the Exposure Activity Ratio (EAR) for exposure to a compound of interest (EARtest) to the EAR for an established safe exposure level to a comparator compound (EARcomparator), acting by the same mode of action. It can be concluded that the exposure to a test compound is safe at a corresponding DCR ≤ 1. In this study, genistein (GEN) was selected as a comparator compound by comparison of reported safe internal exposures to GEN to its BMCL05, as no effect level, the latter determined in the in vitro estrogenic MCF7/Bos proliferation, T47D ER-CALUX, and U2OS ERα-CALUX assay. The EARcomparator was defined using the BMCL05 and EC50 values from the 3 in vitro assays and subsequently used to calculate the DCRs for exposures to 14 test compounds, predicting the (absence of) estrogenicity. The predictions were evaluated by comparison to reported in vivo estrogenicity in humans for these exposures. The results obtained support in the DCR approach as an important animal-free new approach methodology (NAM) in NGRA and show how in vitro assays can be used to define DCR values.

Research on Recognition of Coal and Gangue Based on Laser Speckle Images.

Coal gangue image recognition is a critical technology for achieving automatic separation in coal processing, characterized by its rapid, environmentally friendly, and energy-saving nature. However, the response characteristics of coal and gangue vary greatly under different illuminance conditions, which poses challenges to the stability of feature extraction and recognition, especially when strict illuminance requirements are necessary. This leads to fluctuating coal gangue recognition accuracy in industrial environments. To address these issues and improve the accuracy and stability of image recognition under variable illuminance conditions, we propose a novel coal gangue recognition method based on laser speckle images. Firstly, we studied the inter-class separability and intra-class compactness of the collected laser speckle images of coal and gangue by extracting gray and texture features from the laser speckle images, and analyzed the performance of laser speckle images in representing the differences between coal and gangue minerals. Subsequently, coal gangue recognition was achieved using an SVM classifier based on the extracted features from the laser speckle images. The fusion feature approach achieved a recognition accuracy of 94.4%, providing further evidence of the feasibility of this method. Lastly, we conducted a comparative experiment between natural images and laser speckle images for coal gangue recognition using the same features. The average accuracy of coal gangue laser speckle image recognition under various lighting conditions is 96.7%, with a standard deviation of the recognition accuracy of 1.7%. This significantly surpasses the recognition accuracy obtained from natural coal and gangue images. The results showed that the proposed laser speckle image features can facilitate more stable coal gangue recognition with illumination factors, providing a new, reliable method for achieving accurate classification of coal and gangue in the industrial environment of mines.

PBK modelling of topical application and characterisation of the uncertainty of Cmax estimate: A case study approach.

Combined with in vitro bioactivity data, physiologically based kinetic (PBK) models has increasing applications in next generation risk assessment for animal-free safety decision making. A tiered framework of building PBK models for such application has been developed with increasing complexity and refinements, as model parameters determined in silico, in vitro, and with human pharmacokinetic data become progressively available. PBK modelling has been widely applied for oral/intravenous administration, but less so on topically applied chemicals. Therefore, building PBK models for topical applications and characterizing their uncertainties in the tiered approach is critical to safety decision making. The purpose of this study was to assess the confidence of PBK modelling of topically applied chemicals following the tiered framework, using non-animal methods derived parameters. Prediction of maximum plasma concentration (Cmax) and area under the curve were compared to observed kinetics from published dermal clinical studies for five chemicals (diclofenac, salicylic acid, coumarin, nicotine, caffeine). A bespoke Bayesian statistical model was developed to describe the distributions of Cmax errors between the predicted and observed data. We showed a general trend that confidence in model predictions increases when more quality in vitro data, particularly those on hepatic clearance and dermal absorption, are available as model input. The overall fold error distributions are useful for characterizing model uncertainty. We concluded that by identifying and quantifying the uncertainties in the tiered approach, we can increase the confidence in using PBK modelling to help make safety decisions on topically applied chemicals in the absence of human pharmacokinetic data.

Towards best use and regulatory acceptance of generic physiologically based kinetic (PBK) models for in vitro-to-in vivo extrapolation (IVIVE) in chemical risk assessment.

With an increasing need to incorporate new approach methodologies (NAMs) in chemical risk assessment and the concomitant need to phase out animal testing, the interpretation of in vitro assay readouts for quantitative hazard characterisation becomes more important. Physiologically based kinetic (PBK) models, which simulate the fate of chemicals in tissues of the body, play an essential role in extrapolating in vitro effect concentrations to in vivo bioequivalent exposures. As PBK-based testing approaches evolve, it will become essential to standardise PBK modelling approaches towards a consensus approach that can be used in quantitative in vitro-to-in vivo extrapolation (QIVIVE) studies for regulatory chemical risk assessment based on in vitro assays. Based on results of an ECETOC expert workshop, steps are recommended that can improve regulatory adoption: (1) define context and implementation, taking into consideration model complexity for building fit-for-purpose PBK models, (2) harmonise physiological input parameters and their distribution and define criteria for quality chemical-specific parameters, especially in the absence of in vivo data, (3) apply Good Modelling Practices (GMP) to achieve transparency and design a stepwise approach for PBK model development for risk assessors, (4) evaluate model predictions using alternatives to in vivo PK data including read-across approaches, (5) use case studies to facilitate discussions between modellers and regulators of chemical risk assessment. Proof-of-concepts of generic PBK modelling approaches are published in the scientific literature at an increasing rate. Working on the previously proposed steps is, therefore, needed to gain confidence in PBK modelling approaches for regulatory use.

Clinical Application of Ultrasound Guidance for Parametrial Treatment of Advanced Cervical Cancer.

OBJECTIVES: To evaluate the accuracy of ultrasound (US) in determining the positions of parametrial implants by comparing US with magnetic resonance imaging (MRI) for advanced cervical cancer. METHODS: Patients undergoing brachytherapy with parametrial implantation for cervical cancer from February 2017 to February 2019 were involved in the study. The transverse section of the cervix (surface S1 ) and the transverse section 1 cm above the external cervix (surface S2 ) were selected from MRI and US images as the observation planes. In the MRI observation plane, the distances between the uterine titanium needles and the uterine tube/implanter were set as M1 to M4 ; in the US observation plane, the distances between the uterine titanium needles and the uterine tube/implanter were set as D1 to D4 . The differences and consistency in M and D of each group were then compared. RESULTS: There were no significant differences between M and D in each group (P = .058; P = .821; P = .870; and P = .936, respectively). The intraclass correlation coefficients of M and D in each group were 0.970, 0.968, 0.952, and 0.956. A regression analysis showed that the relationships between M and D in each group were as follows: M1 = 0.9449D1 + 0.1812; M2 = 0.9463D2 + 0.0965; M3 = 0.9176D3 + 0.1233; and M4 = 0.9253D4 + 0.1224. CONCLUSIONS: In parametrial brachytherapy for cervical cancer, US can accurately determine the positions of parametrial implantation needles, which is already applicable on MRI, and can provide assistance in parametrial brachytherapy for advanced cervical cancer.

Application of the Hepatic Transit Time (HTT) in Evaluation of Portal Vein Pressure in Gastroesophageal Varices Patients.

OBJECTIVES: To analyze the clinical significance of using hepatic transit time (HTT) to evaluate portal vein pressure in gastroesophageal varices patients. METHODS: For the observation group, we enrolled 50 gastroesophageal varices patients who had received esophagogastric variceal embolization in our hospital between January 2015 and February 2018. Patients without liver disease populated the control group and were recruited during the same time period. All patients underwent contrast-enhanced sonography. In the observation group, free portal pressure (FPP) was detected during esophagogastric variceal embolization with ultrasound guidance. Differences in hepatic artery-hepatic vein transit time (HA-HVTT), portal vein-hepatic vein transit time (PV-HVTT), and parenchyma-hepatic vein transit time (PA-HVTT) were compared between groups. Correlations between HA-HVTT, PV-HVTT, PA-HVTT, and FPP in the observation group were analyzed using the Pearson coefficient and linear regression analysis. RESULTS: HA-HVTT (t = 5.078; P < .001), PV-HVTT (t = 12.163; P < .001), and PA-HVTT (t = 2.649; P = .009) within the observation group were significantly lower than those of the control group. The areas under the curve of HTT were 0.771 (HA-HVTT), 0.951 (PV-HVTT), and 0.652 (PA-HVTT), and the sensitivity and specificity of PV-HVTT at 7.99 seconds were 86.0% and 88.0%, respectively. The HA-HVTT (r = -0.799; P < .001), PV-HVTT (r = -0.554; P < .001), and PA-HVTT (r = -0.735; P < .001) negatively correlated to FPP in the observation group. Linear regression analysis showed y = -0.410x + 7.254 (HA-HVTT and FPP), y = -0.335x + 4.983 (PV-HVTT and FPP), and y = -0.566x + 4.997 (PA-HVTT and FPP) in the observation group. CONCLUSION: Compared with the control patients, the HTT of patients with portal hypertension-esophagogastric varices was significantly shorter, and showed an inverse relationship with FPP.

Extended evaluation on the ES-D3 cell differentiation assay combined with the BeWo transport model, to predict relative developmental toxicity of triazole compounds.

The mouse embryonic stem D3 (ES-D3) cell differentiation assay is based on the morphometric measurement of cardiomyocyte differentiation and is a promising tool to detect developmental toxicity of compounds. The BeWo transport model, consisting of BeWo b30 cells grown on transwell inserts and mimicking the placental barrier, is useful to determine relative placental transport velocities of compounds. We have previously demonstrated the usefulness of the ES-D3 cell differentiation assay in combination with the in vitro BeWo transport model to predict the relative in vivo developmental toxicity potencies of a set of reference azole compounds. To further evaluate this combined in vitro toxicokinetic and toxicodynamic approach, we combined ES-D3 cell differentiation data of six novel triazoles with relative transport rates obtained from the BeWo model and compared the obtained ranking to the developmental toxicity ranking as derived from in vivo data. The data show that the combined in vitro approach provided a correct prediction for in vivo developmental toxicity, whereas the ES-D3 cell differentiation assay as stand-alone did not. In conclusion, we have validated the combined in vitro approach for developmental toxicity, which we have previously developed with a set of reference azoles, for a set of six novel triazoles. We suggest that this combined model, which takes both toxicodynamic and toxicokinetic aspects into account, should be further validated for other chemical classes of developmental toxicants.

Modulation of (-)-epicatechin metabolism by coadministration with other polyphenols in Caco-2 cell model.

Widely consumed beverages such as red wine, tea, and cocoa-derived products are a great source of flavanols. Epidemiologic and interventional studies suggest that cocoa flavanols such as (-)-epicatechin may reduce the risk of cardiovascular diseases. The interaction of (-)-epicatechin with food components including other polyphenols could modify its absorption, metabolism, and finally its bioactivity. In the present study we investigate (-)-epicatechin absorption and metabolism when coexposed with other polyphenols in the intestinal absorptive Caco-2 cell model. Depending on the type of polyphenols coadministered, the total amount of 3'-O-methyl-epicatechin and 3'-O-sulfate-epicatechin conjugates found both in apical and basal compartments ranged from 19 to 801 nM and from 6 to 432 nM, respectively. The coincubation of (-)-epicatechin with flavanols, chlorogenic acid, and umbelliferone resulted in similar amounts of 3'-O-methyl-epicatechin effluxed into the apical compartment relative to control. Coincubation with isorhamnetin, kaempferol, diosmetin, nevadensin, chrysin, equol, genistein, and hesperitin promoted the transport of 3'-O-methyl-epicatechin toward the basolateral side and decreased the apical efflux. Quercetin and luteolin considerably inhibited the appearance of this (-)-epicatechin conjugate both in the apical and basolateral compartments. In conclusion, we could demonstrate that the efflux of (-)-epicatechin conjugates to the apical or basal compartments of Caco-2 cells is modulated by certain classes of polyphenols and their amount. Ingesting (-)-epicatechin with specific polyphenols could be a strategy to increase the bioavailability of (-)-epicatechin and to modulate its metabolic profile.

Morphological Changes and Strong Cytotoxicity in Yarrowia lipolytica by Overexpressing Delta-12-Desaturase.

In this study, delta-12 desaturase was overexpressed in Yarrowia lipolytica using the single-copy integrative vector pINA1312 and multicopy integrative vector pINA1292, resulting in the engineered yeast strains 1312-12 and 1292-12, respectively. The content of intracellular linoleic acid (LA) in the 1292-12 strain was much higher than in the 1312-12 strain and the control group. One interesting finding was that the 1292-12 strain showed obvious changes in surface morphology. The 1292-12 colonies were much smaller and smoother, whereas their single cells became much larger compared to the control strain. In addition, the dry cell weight (DCW) of the 1292-12 strain was obviously increased from 8.5 to 12.7 g/L, but the viable cell number sharply decreased from 107 to 105/mL. These results indicated that increased LA content in Yarrowia lipolytica could induce morphological changes or even oxidative stress-dependent cell death. The reactive oxygen species (ROS) and malondialdehyde (MDA) were accumulated in the 1292-12 strain, while the antioxidant activities of intracellular catalase (CAT) and superoxide dismutase (SOD) were significantly decreased by 27.6 and 32.0%, respectively. Furthermore, it was also revealed that these issues could be ameliorated by the exogenous supplementation of vitamin C, fish and colza oil.

Assessment of an in vitro transport model using BeWo b30 cells to predict placental transfer of compounds.

The human ex vivo placental perfusion model has regularly been used to study the transplacental transport of compounds. However, this method is laborious and dependent on the presence of fresh human placenta, hampering its use for the assessment of large numbers of compounds. An in vitro model for the placental barrier using BeWo b30 cells may provide an alternative to the ex vivo system. The present study aims to assess whether such an in vitro model could be used to reliably predict placental transfer. To this end, BeWo b30 cells, derived from a human choriocarcinoma, were grown on transwell insert to form a cell layer, separating an apical maternal compartment from a basolateral fetal compartment. For a set of nine selected model compounds, including the reference compound antipyrine, the transport velocity from the apical to the basolateral compartment was determined. Relative transport rates obtained were compared with the transfer indices (a measure for the transport relative to antipyrine) of these compounds obtained in ex vivo placental perfusion studies as reported in the literature. The relative transport rates in the in vitro BeWo model were in good correlation (R² = 0.95) with the transfer indices reported for the ex vivo model. This indicates that the BeWo model could be a valuable in vitro model for prediction of placental transfer of compounds.

Risk assessment in the 21st century: where are we heading?

Reliance on animal tests for chemical safety assessment is increasingly being challenged, not only because of ethical reasons, but also because they procrastinate regulatory decisions and because of concerns over the transferability of results to humans. New approach methodologies (NAMs) need to be fit for purpose and new thinking is required to reconsider chemical legislation, validation of NAMs and opportunities to move away from animal tests. This article summarizes the presentations from a symposium at the 2022 Annual Congress of the British Toxicology Society on the topic of the future of chemical risk assessment in the 21st century. The symposium included three case-studies where NAMs have been used in safety assessments. The first case illustrated how read-across augmented with some in vitro tests could be used reliably to perform the risk assessment of analogues lacking data. The second case showed how specific bioactivity assays could identify an NAM point of departure (PoD) and how this could be translated through physiologically based kinetic modelling in an in vivo PoD for the risk assessment. The third case showed how adverse-outcome pathway (AOP) information, including molecular-initiating event and key events with their underlying data, established for certain chemicals could be used to produce an in silico model that is able to associate chemical features of an unstudied substance with specific AOPs or AOP networks. The manuscript presents the discussions that took place regarding the limitations and benefits of these new approaches, and what are the barriers and the opportunities for their increased use in regulatory decision making.

ADME characterization and PBK model development of 3 highly protein-bound UV filters through topical application.

Estimating human exposure in the safety assessment of chemicals is crucial. Physiologically based kinetic (PBK) models which combine information on exposure, physiology, and chemical properties, describing the absorption, distribution, metabolism, and excretion (ADME) processes of a chemical, can be used to calculate internal exposure metrics such as maximum concentration and area under the concentration-time curve in plasma or tissues of a test chemical in next-generation risk assessment. This article demonstrates the development of PBK models for 3 UV filters, specifically octyl methoxycinnamate, octocrylene, and 4-methylbenzylidene camphor. The models were parameterized entirely based on data obtained from in vitro and/or in silico methods in a bottom-up modeling approach and then validated based on human dermal pharmacokinetic (PK) data. The 3 UV filters are "difficult to test" in in vitro test systems due to high lipophilicity, high binding affinity for proteins, and nonspecific binding, for example, toward plastic. This research work presents critical considerations in ADME data generation, interpretation, and parameterization to assure valid PBK model development to increase confidence in using PBK modeling to help make safety decisions in the absence of human PK data. The developed PBK models of the 3 chemicals successfully simulated the plasma concentration profiles of clinical PK data following dermal application, indicating the reliability of the ADME data generated and the parameters determined. The study also provides insights and lessons learned for characterizing ADME and developing PBK models for highly lipophilic and protein-bound chemicals in the future.

Next Generation Risk Assessment of the Anti-Androgen Flutamide Including the Contribution of Its Active Metabolite Hydroxyflutamide.

In next generation risk assessment (NGRA), non-animal approaches are used to quantify the chemical concentrations required to trigger bioactivity responses, in order to assure safe levels of human exposure. A limitation of many in vitro bioactivity assays, which are used in an NGRA context as new approach methodologies (NAMs), is that toxicokinetics, including biotransformation, are not adequately captured. The present study aimed to include, as a proof of principle, the bioactivity of the metabolite hydroxyflutamide (HF) in an NGRA approach to evaluate the safety of the anti-androgen flutamide (FLU), using the AR-CALUX assay to derive the NAM point of departure (PoD). The NGRA approach applied also included PBK modelling-facilitated quantitative in vitro to in vivo extrapolation (QIVIVE). The PBK model describing FLU and HF kinetics in humans was developed using GastroPlus™ and validated against human pharmacokinetic data. PBK model-facilitated QIVIVE was performed to translate the in vitro AR-CALUX derived concentration-response data to a corresponding in vivo dose-response curve for the anti-androgenicity of FLU, excluding and including the activity of HF (-HF and +HF, respectively). The in vivo benchmark dose 5% lower confidence limits (BMDL05) derived from the predicted in vivo dose-response curves for FLU, revealed a 440-fold lower BMDL05 when taking the bioactivity of HF into account. Subsequent comparison of the predicted BMDL05 values to the human therapeutic doses and historical animal derived PoDs, revealed that PBK modelling-facilitated QIVIVE that includes the bioactivity of the active metabolite is protective and provides a more appropriate PoD to assure human safety via NGRA, whereas excluding this would potentially result in an underestimation of the risk of FLU exposure in humans.

Pillararene-containing polymers with tunable fluorescence properties based on host-guest interactions.

Linear polymers containing pillar[5]arenes as the pendant groups were designed and synthesized via a ring-opening metathesis polymerization. Such polymers could form supramolecular brush polymers and exhibited tunable fluorescence properties based on the host-guest interactions.

Ready for regulatory use: NAMs and NGRA for chemical safety assurance.

New approach methodologies (NAMs) that do not use experimental animals are, in certain settings, entirely appropriate for assuring the safety of chemical ingredients, although regulatory adoption has been slow. In this opinion article we discuss how scientific advances that utilize NAMs to certify systemic safety are available now and merit broader acceptance within the framework of next generation risk assessments (NGRA).

Beyond AOPs: A Mechanistic Evaluation of NAMs in DART Testing.

New Approach Methodologies (NAMs) promise to offer a unique opportunity to enable human-relevant safety decisions to be made without the need for animal testing in the context of exposure-driven Next Generation Risk Assessment (NGRA). Protecting human health against the potential effects a chemical may have on embryo-foetal development and/or aspects of reproductive biology using NGRA is particularly challenging. These are not single endpoint or health effects and risk assessments have traditionally relied on data from Developmental and Reproductive Toxicity (DART) tests in animals. There are numerous Adverse Outcome Pathways (AOPs) that can lead to DART, which means defining and developing strict testing strategies for every AOP, to predict apical outcomes, is neither a tenable goal nor a necessity to ensure NAM-based safety assessments are fit-for-purpose. Instead, a pragmatic approach is needed that uses the available knowledge and data to ensure NAM-based exposure-led safety assessments are sufficiently protective. To this end, the mechanistic and biological coverage of existing NAMs for DART were assessed and gaps to be addressed were identified, allowing the development of an approach that relies on generating data relevant to the overall mechanisms involved in human reproduction and embryo-foetal development. Using the knowledge of cellular processes and signalling pathways underlying the key stages in reproduction and development, we have developed a broad outline of endpoints informative of DART. When the existing NAMs were compared against this outline to determine whether they provide comprehensive coverage when integrated in a framework, we found them to generally cover the reproductive and developmental processes underlying the traditionally evaluated apical endpoint studies. The application of this safety assessment framework is illustrated using an exposure-led case study.

Are Non-animal Systemic Safety Assessments Protective? A Toolbox and Workflow.

An important question in toxicological risk assessment is whether non-animal new approach methodologies (NAMs) can be used to make safety decisions that are protective of human health, without being overly conservative. In this work, we propose a core NAM toolbox and workflow for conducting systemic safety assessments for adult consumers. We also present an approach for evaluating how protective and useful the toolbox and workflow are by benchmarking against historical safety decisions. The toolbox includes physiologically based kinetic (PBK) models to estimate systemic Cmax levels in humans, and 3 bioactivity platforms, comprising high-throughput transcriptomics, a cell stress panel, and in vitro pharmacological profiling, from which points of departure are estimated. A Bayesian model was developed to quantify the uncertainty in the Cmax estimates depending on how the PBK models were parameterized. The feasibility of the evaluation approach was tested using 24 exposure scenarios from 10 chemicals, some of which would be considered high risk from a consumer goods perspective (eg, drugs that are systemically bioactive) and some low risk (eg, existing food or cosmetic ingredients). Using novel protectiveness and utility metrics, it was shown that up to 69% (9/13) of the low risk scenarios could be identified as such using the toolbox, whilst being protective against all (5/5) the high-risk ones. The results demonstrated how robust safety decisions could be made without using animal data. This work will enable a full evaluation to assess how protective and useful the toolbox and workflow are across a broader range of chemical-exposure scenarios.