DARTpaths, an in silico platform to investigate molecular mechanisms of compounds.

SUMMARY: Xpaths is a collection of algorithms that allow for the prediction of compound-induced molecular mechanisms of action by integrating phenotypic endpoints of different species; and proposes follow-up tests for model organisms to validate these pathway predictions. The Xpaths algorithms are applied to predict developmental and reproductive toxicity (DART) and implemented into an in silico platform, called DARTpaths. AVAILABILITY AND IMPLEMENTATION: All code is available on GitHub https://github.com/Xpaths/dartpaths-app under Apache license 2.0, detailed overview with demo is available at https://www.vivaltes.com/dartpaths/. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Report of the First ONTOX Stakeholder Network Meeting: Digging Under the Surface of ONTOX Together With the Stakeholders.

The first Stakeholder Network Meeting of the EU Horizon 2020-funded ONTOX project was held on 13-14 March 2023, in Brussels, Belgium. The discussion centred around identifying specific challenges, barriers and drivers in relation to the implementation of non-animal new approach methodologies (NAMs) and probabilistic risk assessment (PRA), in order to help address the issues and rank them according to their associated level of difficulty. ONTOX aims to advance the assessment of chemical risk to humans, without the use of animal testing, by developing non-animal NAMs and PRA in line with 21st century toxicity testing principles. Stakeholder groups (regulatory authorities, companies, academia, non-governmental organisations) were identified and invited to participate in a meeting and a survey, by which their current position in relation to the implementation of NAMs and PRA was ascertained, as well as specific challenges and drivers highlighted. The survey analysis revealed areas of agreement and disagreement among stakeholders on topics such as capacity building, sustainability, regulatory acceptance, validation of adverse outcome pathways, acceptance of artificial intelligence (AI) in risk assessment, and guaranteeing consumer safety. The stakeholder network meeting resulted in the identification of barriers, drivers and specific challenges that need to be addressed. Breakout groups discussed topics such as hazard versus risk assessment, future reliance on AI and machine learning, regulatory requirements for industry and sustainability of the ONTOX Hub platform. The outputs from these discussions provided insights for overcoming barriers and leveraging drivers for implementing NAMs and PRA. It was concluded that there is a continued need for stakeholder engagement, including the organisation of a 'hackathon' to tackle challenges, to ensure the successful implementation of NAMs and PRA in chemical risk assessment.

Effects of exercise during chemo- or radiotherapy on immune markers - a systematic review.

Introduction Patients with cancer receiving radio- or chemotherapy undergo many immunological stressors. Chronic regular exercise was shown to positively influence the immune system in several populations, while exercise overload may have negative effects. Exercise is currently recommended for all patients with cancer. However, knowledge regarding the effects of exercise on immune markers in patients undergoing chemo- or radiotherapy is limited. The aim of this study is to systematically review the effects of moderate and high intensity exercise interventions in patients with cancer during chemotherapy or radiotherapy, on immune markers. Methods For this review, a search was performed in PubMed and EMBASE, until March 2023. Methodological quality was assessed with the Pedro tool and best-evidence syntheses were performed both per immune marker and for the inflammatory profile. Results Methodological quality of the 15 included articles was rated fair to good. The majority of markers was unaltered, but observed effects included a suppressive effect of exercise during radiotherapy on some pro-inflammatory markers, a preserving effect of exercise during chemotherapy on NK cell degranulation and cytotoxicity, a protective effect on the decrease in thrombocytes during chemotherapy, and a positive effect of exercise during chemotherapy on IgA. Discussion/conclusion Although exercise only influenced a few markers, the results are promising. Exercise did not negatively influence immune markers, and some were positively affected since suppressed inflammation might have positive clinical implications. For future research, consensus is needed regarding a set of markers that are most responsive to exercise. Next, differential effects of training types and intensities on these markers should be further investigated, as well as their clinical implications.

A practical guide for probiotics applied to the case of antibiotic-associated diarrhea in The Netherlands.

BACKGROUND: Antibiotic-associated diarrhea (AAD) is a side-effect frequently associated with the use of broad spectrum antibiotics. Although a number of clinical studies show that co-administration of specific probiotics reduces the risk for AAD, there is still unclarity among healthcare professionals on the recommendation of probiotic products. This paper aims at a practical guide to inform healthcare professionals, patients and consumers about the exact product characteristics of available probiotics with a proven efficacy to prevent AAD. METHODS: The workflow in this paper includes three consecutive steps: 1) systematic review of relevant clinical studies for effective probiotics by a meta-analysis, 2) compilation of a list of available probiotic products, and 3) recommendation of probiotic products that match effective formulations. Our systematic review on the efficacy of probiotics for the prevention of AAD included only studies with randomized, double blind placebo-controlled trials, a clear definition of antibiotic associated diarrhea, and a probiotic administration regime for at least the duration of the antibiotic therapy. RESULTS: Using our inclusion criteria, we selected 32 out of 128 identified trials and pooled the results of these studies for each specific dairy product and food supplement. The results indicate a total of seven single or multiple-strain formulations favoring the probiotic treatment group, with the strain Lactobacillus rhamnosus GG being the most effective [relative risk ratio of probiotic versus placebo 0.30 (95% CI 0.16-0.5)]. We selected products for recommendation from a compiled list of all probiotic dairy products and food supplements available in The Netherlands and categorized them into groups of products showing effects against the incidence of AAD in at least one, two or three independent clinical studies. We excluded all products which did not unambiguously declare on the label the specific probiotic strain(s) and the number of colony forming units. CONCLUSION: Here we present a practical guide that informs healthcare professionals and patients on the availability of probiotic products with a proven efficacy for the prevention of AAD.

Regional Expression Levels of Drug Transporters and Metabolizing Enzymes along the Pig and Human Intestinal Tract and Comparison with Caco-2 Cells.

Intestinal transporter proteins and metabolizing enzymes play a crucial role in the oral absorption of a wide variety of drugs. The aim of the current study was to characterize better available intestinal in vitro models by comparing expression levels of these proteins and enzymes between porcine intestine, human intestine, and Caco-2 cells. We therefore determined the absolute protein expression of 19 drug transporters and the mRNA expression of 12 metabolic enzymes along the pig intestinal tract (duodenum, jejunum, ileum; N = 4), in human intestine (jejunum; N = 9), and Caco-2 cells. Expression of the included transporters and enzymes was in general well comparable between porcine and human intestinal tissue, although breast cancer resistance protein, monocarboxylate transporter 5, multidrug resistance protein (MRP) 1, MRP1, MRP3 (∼2-fold), and organic anion-transporting polypeptide (OATP) 4A1 (∼6-fold) was higher expressed in pig compared with human jejunum. Alternatively, expression level of relevant transporter proteins (glucose transporter 1, OATP4A1, MRP2, MRP1, and OATP2B1) was significantly higher (3- to 130-fold) in Caco-2 cells compared with human jejunum. Moreover, all examined CYPs showed at least a fivefold lower gene expression in Caco-2 cells compared with human jejunum, with the smallest differences for CYP1A1 and CYP3A5 and the largest difference for CYP3A4 (871-fold higher expression in human jejunum compared with Caco-2 cells). In conclusion, a comprehensive overview is provided of the expression levels of clinically relevant transporter proteins and metabolic enzymes in porcine and human intestinal tissue and Caco-2 cells, which may assist in deciding upon the most suitable model to further improve our understanding of processes that determine intestinal absorption of compounds.

The Isolated Chicken Eye test to replace the Draize test in rabbits.

In 1944, Draize et al., published a paper entitled "Methods for the study of irritation and toxicity of substances applied topically to the skin and mucous membranes". The Organization for Economic Co-operation and Development published their first guideline on eye irritation in 1981, using rabbits. In the early eighties the development of alternative non-animal tests to replace the Draize eye test started. The first attempts to validate alternative tests for eye irritation were considered to be relatively simple by comparing in vitro and in vivo irritation index scores. In the early nineteen-eighties, we introduced the use of isolated eyes as an alternative test for the Draize eye irritation test. What was expected to be a process of several years, however, turned out to be a decades spanning process still not fully completed. For a large part, this can be attributed to the nature of the in vivo test in rabbits, which is more complicated and compromised than originally believed. This paper describes, most chronologically, the development, performance, validation and application of the Isolated Eye Test and, in broader perspective, the international validation and acceptance of this alternative test by regulatory authorities and agencies.

Prediction of carcinogenic potential of chemicals using repeated-dose (13-week) toxicity data.

Sub-chronic toxicity studies of 163 non-genotoxic chemicals were evaluated in order to predict the tumour outcome of 24-month rat carcinogenicity studies obtained from the EFSA and ToxRef databases. Hundred eleven of the 148 chemicals that did not induce putative preneoplastic lesions in the sub-chronic study also did not induce tumours in the carcinogenicity study (True Negatives). Cellular hypertrophy appeared to be an unreliable predictor of carcinogenicity. The negative predictivity, the measure of the compounds evaluated that did not show any putative preneoplastic lesion in de sub-chronic studies and were negative in the carcinogenicity studies, was 75%, whereas the sensitivity, a measure of the sub-chronic study to predict a positive carcinogenicity outcome was only 5%. The specificity, the accuracy of the sub-chronic study to correctly identify non-carcinogens was 90%. When the chemicals which induced tumours generally considered not relevant for humans (33 out of 37 False Negatives) are classified as True Negatives, the negative predictivity amounts to 97%. Overall, the results of this retrospective study support the concept that chemicals showing no histopathological risk factors for neoplasia in a sub-chronic study in rats may be considered non-carcinogenic and do not require further testing in a carcinogenicity study.

Regulatory acceptance and use of the Extended One Generation Reproductive Toxicity Study within Europe.

The two-generation study (OECD TG 416) is the standard requirement within REACH to test reproductive toxicity effects of chemicals with production volumes >100 tonnes. This test is criticized in terms of scientific relevance and animal welfare. The Extended One Generation Reproductive Toxicity Study (EOGRTS), incorporated into the OECD test guidelines in 2011 (OECD TG 443) has the potential to replace TG 416, while using only one generation of rats and being more informative. However, its regulatory acceptance proved challenging. This article reconstructs the process of regulatory acceptance and use of the EOGRTS and describes drivers and barriers influencing the process. The findings derive from literature research and expert interviews. A distinction is made between three sub-stages; The stage of Formal Incorporation of the EOGRTS into OECD test guidelines was stimulated by retrospective analyses on the value of the second generation (F2), strong EOGRTS advocates, animal welfare concern and changing US and EU chemicals legislation; the stage of Actual Regulatory Acceptance within REACH was challenged by legal factors and ongoing scientific disputes, while the stage of Use by Industry is influenced by uncertainty of registrants about regulatory acceptance, high costs, the risk of false positives and the manageability of the EOGRTS.

Development of a panel of high-throughput reporter-gene assays to detect genotoxicity and oxidative stress.

The lack of toxicological information on many of the compounds that humans use or are exposed to, intentionally or unintentionally, poses a big problem in risk assessment. To fill this data gap, more emphasis is given to fast in vitro screening tools that can add toxicologically relevant information regarding the mode(s) of action via which compounds can elicit adverse effects, including genotoxic effects. By use of bioassays that can monitor the activation of specific cellular signalling pathways, many compounds can be screened in a high-throughput manner. We have developed two new specific reporter-gene assays that can monitor the effects of compounds on two pathways of interest: the p53 pathway (p53 CALUX) for genotoxicity and the Nrf2 pathway (Nrf2 CALUX) for oxidative stress. To exclude non-specific effects by compounds influencing the luciferase reporter-gene expression non-specifically, a third assay was developed to monitor changes in luciferase expression by compounds in general (Cytotox CALUX). To facilitate interpretation of the data and to avoid artefacts, all three reporter-gene assays used simple and defined reporter genes and a similar cellular basis, the human U2OS cell line. The three cell lines were validated with a range of reference compounds including genotoxic and non-genotoxic agents. The sensitivity (95%) and specificity (85%) of the p53 CALUX was high, showing that the assay is able to identify various types of genotoxic compound, while avoiding the detection of false positives. The Nrf2 CALUX showed specific responses to oxidants only, enabling the identification of compounds that elicit part of their genotoxicity via oxidative stress. All reporter-gene assays can be used in a high-throughput screening format and can be supplemented with other U2OS-based reporter-gene assays that can profile nuclear receptor activity, and several other signalling pathways.

Regulatory acceptance and use of 3R models for pharmaceuticals and chemicals: expert opinions on the state of affairs and the way forward.

Pharmaceuticals and chemicals are subjected to regulatory safety testing accounting for approximately 25% of laboratory animal use in Europe. This testing meets various objections and has led to the development of a range of 3R models to Replace, Reduce or Refine the animal models. However, these models must overcome many barriers before being accepted for regulatory risk management purposes. This paper describes the barriers and drivers and options to optimize this acceptance process as identified by two expert panels, one on pharmaceuticals and one on chemicals. To untangle the complex acceptance process, the multilevel perspective on technology transitions is applied. This perspective defines influences at the micro-, meso- and macro level which need alignment to induce regulatory acceptance of a 3R model. This paper displays that there are many similar mechanisms within both sectors that prevent 3R models from becoming accepted for regulatory risk assessment and management. Shared barriers include the uncertainty about the value of the new 3R models (micro level), the lack of harmonization of regulatory requirements and acceptance criteria (meso level) and the high levels of risk aversion (macro level). In optimizing the process commitment, communication, cooperation and coordination are identified as critical drivers.

Beyond Animal Testing Index: Benchmarking tool for a world beyond animal testing.

While the original definition of replacement focuses on the replacement of the use of animals in science, a more contemporary definition focuses on accelerating the development and use of predictive and robust models, based on the latest science and technologies, to address scientific questions without the use of animals. The transition to animal free innovation is on the political agenda in and outside the European Union. The Beyond Animal Testing Index (BATI) is a benchmarking instrument designed to provide insight into the activities and contributions of research institutes to the transition to animal free innovation. The BATI allows participating organizations to learn from each other and stimulates continuous improvement. The BATI was modelled after the Access to Medicine Index, which benchmarks pharmaceutical companies on their efforts to make medicines widely available in developing countries. A prototype of the BATI was field-tested with three Dutch aca­demic medical centers and two universities in 2020-2021. The field test demonstrated the usability and effectiveness of the BATI as a benchmarking tool. Analyses were performed across five different domains. The participating institutes concluded that the BATI served as an internal as well as an external stimulus to share, learn, and improve institutional strategies towards the transition to animal free innovation. The BATI also identified gaps in the development and implementation of 3R technologies. Hence, the BATI might be a suitable instrument for monitoring the effectiveness of policies. BATI version 1.0 is ready to be used for benchmarking at a larger scale.

The use of animals for research is being scrutinized by the public. The transition to animal-free methods is on the political agenda in and outside the European Union. This requires accelerating the development and use of useful and reliable animal-free methods. The Beyond Animal Testing Index (BATI) is designed to provide insight into the activities and contributions of research institutes to the transition to animal-free innovation. The BATI allows participating organizations to learn from each other and stimulates continuous improvement. A prototype of the BATI was field-tested with three Dutch academic medical centers and two universities in 2020-2021. The field test showed that the BATI could be used to monitor how effective policies are and to show where more work is needed towards the full replacement of animals in research.

How to optimize communication about animal and animal-free research methods.

There is a long-standing debate about experimental non-human animals and animal-free methods in scientific research. Among the various stakeholders involved in the debate are the scientists. During media broadcasts we, animal researchers and animal-free methods researchers, were positioned as 'opponents'. In this essay we describe our initial rational thoughts and emotions after these events, and how we came together to explore our common ground on animal(-free) experimentation. Realizing that all models have advantages and limitations, our common ground lies in the principles of good scientific research and responsible experimentation. Our communication emanating from the broadcasts has been instrumental in improving communication on animal(-free) experimentation issues by teaming up. We strongly believe that this is essential for making well-informed decisions for the methods we are using now and will be using in the future.

Comet assay in reconstructed 3D human epidermal skin models--investigation of intra- and inter-laboratory reproducibility with coded chemicals.

Reconstructed 3D human epidermal skin models are being used increasingly for safety testing of chemicals. Based on EpiDerm™ tissues, an assay was developed in which the tissues were topically exposed to test chemicals for 3h followed by cell isolation and assessment of DNA damage using the comet assay. Inter-laboratory reproducibility of the 3D skin comet assay was initially demonstrated using two model genotoxic carcinogens, methyl methane sulfonate (MMS) and 4-nitroquinoline-n-oxide, and the results showed good concordance among three different laboratories and with in vivo data. In Phase 2 of the project, intra- and inter-laboratory reproducibility was investigated with five coded compounds with different genotoxicity liability tested at three different laboratories. For the genotoxic carcinogens MMS and N-ethyl-N-nitrosourea, all laboratories reported a dose-related and statistically significant increase (P < 0.05) in DNA damage in every experiment. For the genotoxic carcinogen, 2,4-diaminotoluene, the overall result from all laboratories showed a smaller, but significant genotoxic response (P < 0.05). For cyclohexanone (CHN) (non-genotoxic in vitro and in vivo, and non-carcinogenic), an increase compared to the solvent control acetone was observed only in one laboratory. However, the response was not dose related and CHN was judged negative overall, as was p-nitrophenol (p-NP) (genotoxic in vitro but not in vivo and non-carcinogenic), which was the only compound showing clear cytotoxic effects. For p-NP, significant DNA damage generally occurred only at doses that were substantially cytotoxic (>30% cell loss), and the overall response was comparable in all laboratories despite some differences in doses tested. The results of the collaborative study for the coded compounds were generally reproducible among the laboratories involved and intra-laboratory reproducibility was also good. These data indicate that the comet assay in EpiDerm™ skin models is a promising model for the safety assessment of compounds with a dermal route of exposure.

The use of ex vivo human skin tissue for genotoxicity testing.

As a result of the chemical legislation concerning the registration, evaluation, authorization and restriction of chemicals (REACH), and the Seventh Amendment to the Cosmetics Directive, which prohibits animal testing in Europe for cosmetics, alternative methods for safety evaluation of chemicals are urgently needed. Current in vitro genotoxicity assays are not sufficiently predictive for the in vivo situation, resulting in an unacceptably high number of misleading positives. For many chemicals and ingredients of personal care products the skin is the first site of contact, but there are no in vitro genotoxicity assays available in the skin for additional evaluation of positive or equivocal responses observed in regulatory in vitro genotoxicity assays. In the present study ex vivo human skin tissue obtained from surgery was used for genotoxicity evaluation of chemicals by using the comet assay. Fresh ex vivo human skin tissue was cultured in an air-liquid interface and topically exposed to 20 chemicals, including true positive, misleading positive and true negative genotoxins. Based on the results obtained in the present study, the sensitivity, specificity and accuracy of the ex vivo skin comet assay to predict in vivo genotoxicity were 89%, 90% and 89%, respectively. Donor and experimental variability were mainly reflected in the magnitude of the response and not the difference between the presence and absence of a genotoxic response. The present study indicates that human skin obtained from surgery is a promising and robust model for safety evaluation of chemicals that are in direct contact with the skin.

The in vivo rat skin photomicronucleus assay: phototoxicity and photogenotoxicity evaluation of six fluoroquinolones.

An in vivo photomicronucleus test (MNT) using rat skin, the target organ for photoirritancy and carcinogenicity, was recently described. The assay was evaluated using fluoroquinolone (FQ) antibiotics with varying degrees of phototoxic potency (i.e. sparflocacin [SPFX], lomefloxacin [LOFX], ciprofloxacin [CIFX], levofloxacin [LEFX], gemifloxacin [GEFX] and gatifloxacin [GAFX]) using a solar simulator producing both UVA and UVB (ratio 23:1). Experiments were performed at The Netherlands Organisation for Applied Scientific Research (TNO) and GlaxoSmithKline (GSK) to investigate interlaboratory variability, including evaluation of phototoxicity (clinical signs), micronucleus induction and histopathology. The potency of micronuclei (MN) formation in rat skin induced by the FQs was SPFX = LOFX > CIFX = LEFX, however, MN induction was only statistically significant for SPFX and LOFX. In both laboratories, GEFX and GAFX did not increase the MN frequencies compared to the irradiated vehicle control. Signs of phototoxicity, including clinical and histopathological changes, were observed with SPFX and LOFX to a similar degree as the positive control, 8-methoxypsoralen. In addition, there were some clinical signs of phototoxicity seen with CIFX, LEFX, GEFX and GAFX, but not always in both laboratories for CIFX, GEFX and GAFX and when observed, these were considered only mild. Of these, only LEFX also showed histopathological changes. In all studies, photogenotoxic potency correlated with photocarcinogenic potential and moreover, photogenotoxicity was not observed in the absence of phototoxicity. The results of the TNO/GSK study indicate that the in vivo rat skin photoMNT may be a promising tool for detection of photoclastogencity and photoirritancy in the skin/eye in the same animal. Given the association between the MNT and cancer, the skin photoMNT may also provide a promising tool for the early detection of photocarcinogenesis and help bridge the gap in the existing photosafety testing paradigm.

The 3T3 neutral red uptake phototoxicity test: practical experience and implications for phototoxicity testing--the report of an ECVAM-EFPIA workshop.

This is the report from the "ECVAM-EFPIA workshop on 3T3 NRU Phototoxicity Test: Practical Experience and Implications for Phototoxicity Testing", jointly organized by ECVAM and EFPIA and held on the 25-27 October 2010 in Somma Lombardo, Italy. The European Centre for the Validation of Alternative Methods (ECVAM) was established in 1991 within the European Commission Joint Research, based on a Communication from the European Commission (1991). The main objective of ECVAM is to promote the scientific and regulatory acceptance of alternative methods which are of importance to the biosciences and which reduce, refine and replace the use of laboratory animals. The European Federation of Pharmaceuticals Industries and Association (EFPIA) represent the pharmaceutical industry operating in Europe. Through its direct membership of 31 national associations and 40 leading pharmaceutical companies, EFPIA is the voice on the EU scene of 2200 companies committed to researching, developing and bringing to patients new medicines that improve health and the quality of life around the world. The workshop, co-chaired by Joachim Kreysa (ECVAM) and Phil Wilcox (GSK, EFPIA) involved thirty-five experts from academia, regulatory authorities and industry, invited to contribute with their experiences in the field of phototoxicology. The main objectives of the workshop were: -to present 'in use' experience of the pharmaceutical industry with the 3T3 Neutral Red Uptake Phototoxicity Test (3T3 NRU-PT), -to discuss why it differs from the results in the original validation exercise, -to discuss technical issues and consider ways to improve the usability of the 3T3 NRU-PT for (non-topical) pharmaceuticals, e.g., by modifying the threshold of chemical light absorption to trigger photo-toxicological testing, and by modifying technical aspects of the assay, or adjusting the criteria used to classify a positive response. During the workshop, the assay methodology was reviewed by comparing the OECD Test Guideline (TG 432) with the protocols used in testing laboratories, data from EFPIA and JPMA 'surveys' were presented and possible reasons for the outcomes were discussed. Experts from cosmetics and pharmaceutical industries reported on their experience with the 3T3 NRU-PT and evidence was presented for phototoxic clinical symptoms that could be linked to certain relevant molecules. Brainstorming sessions discussed if the 3T3 NRU-PT needed to be improved and whether alternatives to the 3T3 NRU-PT exist. Finally, the viewpoint from EU and US regulators was presented. In the final session, the conclusions of the meeting were summarized, with action points. It was concluded that the 3T3 NRU-PT identifies phototoxicological hazards with a 100% sensitivity, and thus is accepted as the tier one test that correctly identifies the absence of phototoxic potential. Consequently, positive results in the 3T3 NRU-PT often do not translate into a clinical phototoxicity risk. Possible ways to improve the practical use of this assay include: (i) adaptation of changed UV/vis-absorption criteria as a means to reduce the number of materials tested, (ii) reduction of the highest concentration to be tested, and (iii) consideration of modifying the threshold criteria for the prediction of a positive call in the test.

Natural language processing in toxicology: Delineating adverse outcome pathways and guiding the application of new approach methodologies.

Adverse Outcome Pathways (AOPs) are conceptual frameworks that tie an initial perturbation (molecular initiating event) to a phenotypic toxicological manifestation (adverse outcome), through a series of steps (key events). They provide therefore a standardized way to map and organize toxicological mechanistic information. As such, AOPs inform on key events underlying toxicity, thus supporting the development of New Approach Methodologies (NAMs), which aim to reduce the use of animal testing for toxicology purposes. However, the establishment of a novel AOP relies on the gathering of multiple streams of evidence and information, from available literature to knowledge databases. Often, this information is in the form of free text, also called unstructured text, which is not immediately digestible by a computer. This information is thus both tedious and increasingly time-consuming to process manually with the growing volume of data available. The advancement of machine learning provides alternative solutions to this challenge. To extract and organize information from relevant sources, it seems valuable to employ deep learning Natural Language Processing techniques. We review here some of the recent progress in the NLP field, and show how these techniques have already demonstrated value in the biomedical and toxicology areas. We also propose an approach to efficiently and reliably extract and combine relevant toxicological information from text. This data can be used to map underlying mechanisms that lead to toxicological effects and start building quantitative models, in particular AOPs, ultimately allowing animal-free human-based hazard and risk assessment.

Development and characterisation of an in vitro photomicronucleus test using ex vivo human skin tissue.

Photosafety testing is of concern for the evaluation of personal care products and pharmaceuticals. Current regulatory guidance state that photosafety should be evaluated for compounds that absorb radiation between 290 and 700 nm with relevant exposure in the skin or eyes. However, oversensitivity and the occurrence of 'pseudo-effects' with current in vitro photo(geno)toxicity assays have become a major problem. Furthermore, at this moment, there are no relevant in vitro assays available to identify the photocarcinogenic potential of compounds, which might result in unnecessary in vivo photocarcinogenicity studies for pharmaceutical ingredients or unnecessary dropouts in the development of ingredients of personal care products. For these reasons, availability of a relevant and highly predictive in vitro model from human origin to identify the photogenotoxic and/or photocarcinogenic potential of compounds is viewed as high priority. In the present study, human skin tissue obtained from surgery was used for developing a photomicronucleus test. Prior to investigations of the photogenotoxic potential of 8-methoxypsoralen, tissue viability (lactate production and lactate dehydrogenase leakage), cell proliferation (Ki-67 expression) and the effect of ultraviolet (UV) exposure on viability (MTT test), proliferation (Ki-67 expression) and p53 expression were determined. Results of the present study indicate that ex vivo human skin seems to be a relevant method for safety evaluation of compounds that reach the skin in combination with UV exposure.

Considerations on photochemical genotoxicity. II: report of the 2009 International Workshop on Genotoxicity Testing Working Group.

A workshop to reappraise the previous IWGT recommendations for photogenotoxicity testing [E. Gocke, L. Muller, P.J. Guzzie, S. Brendler-Schwaab, S. Bulera, C.F. Chignell, L.M. Henderson, A. Jacobs, H. Murli, R.D. Snyder, N. Tanaka, Considerations on photochemical genotoxicity: report of the International Workshop on Genotoxicity Test Procedures working group, Environ. Mol. Mutagen., 35 (2000) 173-184] was recently held as part of the 5th International Workshop on Genotoxicity Testing (IWGT) meeting in Basel, Switzerland (August 17-19, 2009). An Expert Panel was convened from regulatory, academic and industrial scientists (with several members serving on the original panel) and chaired by Dr Peter Kasper (BfArM, Germany). The aim of the workshop was to review progress made in photo(geno)toxicity testing over the past decade; a period which saw the introduction of several regulatory photosafety guidances in particular in Europe and the USA. Based on current regulatory guidelines a substantial proportion of compounds trigger the requirements for photosafety testing. Moreover, there has been growing concern within industry about the performance of the in vitro photosafety tests in the "real world" of compound development. Therefore, the expert group reviewed the status of the current regulatory guidance's and the impact these have had on compound development in the context of the various triggers for photosafety testing. In addition, the performance of photogenotoxicity assays (old and new) was discussed, particularly in view of reports of pseudophotoclastogencity. The Expert Panel finished with an assessment of the positioning of photogenotoxicity testing within a photosafety testing strategy. The most significant conclusion made by the Expert Panel was that photogenotoxicity testing should no longer be recommended as part of the standard photosafety testing strategy. In addition, progress was made on the refinement of triggers for photosafety testing. For example, there was support for harmonisation of methods to determine the Molar Extinction Coefficient (MEC) and a consensus agreement that there should be no requirement for testing of compounds with a MEC<1000Lmol(-1)cm(-1).

Towards a reporting guideline for developmental and reproductive toxicology testing in C. elegans and other nematodes.

Implementation of reliable methodologies allowing Reduction, Refinement, and Replacement (3Rs) of animal testing is a process that takes several decades and is still not complete. Reliable methods are essential for regulatory hazard assessment of chemicals where differences in test protocol can influence the test outcomes and thus affect the confidence in the predictive value of the organisms used as an alternative for mammals. Although test guidelines are common for mammalian studies, they are scarce for non-vertebrate organisms that would allow for the 3Rs of animal testing. Here, we present a set of 30 reporting criteria as the basis for such a guideline for Developmental and Reproductive Toxicology (DART) testing in the nematode Caenorhabditis elegans. Small organisms like C. elegans are upcoming in new approach methodologies for hazard assessment; thus, reliable and robust test protocols are urgently needed. A literature assessment of the fulfilment of the reporting criteria demonstrates that although studies describe methodological details, essential information such as compound purity and lot/batch number or type of container is often not reported. The formulated set of reporting criteria for C. elegans testing can be used by (i) researchers to describe essential experimental details (ii) data scientists that aggregate information to assess data quality and include data in aggregated databases (iii) regulators to assess study data for inclusion in regulatory hazard assessment of chemicals.