Metabolism-based category formation for the prioritisation of genotoxicity hazard assessment for plant protection product residues (Part 4): α-Chloroacetamides.

In dietary risk assessment of plant protection products, residues of active ingredients and their metabolites need to be evaluated for their genotoxic potential. The European Food Safety Authority recommend a tiered approach focussing assessment and testing on classes of similar chemicals. To characterise similarity, in terms of metabolism, a metabolic similarity profiling scheme has been developed from an analysis of 69 α-chloroacetamide herbicides for which either Ames, chromosomal aberration or micronucleus test results are publicly available. A set of structural space alerts were defined, each linked to a key metabolic transformation present in the α-chloroacetamide metabolic space. The structural space alerts were combined with covalent chemistry profiling to develop categories suitable for chemical prioritisation via read-across. The method is a robust and reproducible approach to such read-across predictions, with the potential to reduce unnecessary testing. The key challenge in the approach was identified as being the need for metabolism data individual groups of plant protection products as the basis for the development of the structural space alerts.

Metabolism-based category formation for the prioritisation of genotoxicity hazard assessment for plant protection product residues (part 3): Strobilurins.

In dietary risk assessment of plant protection products, residues of active ingredients and their metabolites need to be evaluated for their genotoxic potential. The European Food Safety Authority recommend a tiered approach focussing assessment and testing on classes of similar chemicals. To characterise similarity, in terms of metabolism, a metabolic similarity profiling scheme has been developed from an analysis of 46 chemicals of strobilurin fungicides and their metabolites for which either Ames, chromosomal aberration or micronucleus test results are publicly available. This profiling scheme consists of a set of ten sub-structures, each linked to a key metabolic transformation present in the strobilurin metabolic space. This metabolic similarity profiling scheme was combined with covalent chemistry profiling and physico-chemistry properties to develop chemical categories suitable for chemical prioritisation via read-across. The method is a robust and reproducible approach to such read-across predictions, with the potential to reduce unnecessary testing. The key challenge in the approach was identified as being the need for metabolism data and individual groups of plant protection products as the basis for the development of such profiling schemes.

Sub-structure-based category formation for the prioritisation of genotoxicity hazard assessment for pesticide residues (part 2): Triazoles.

In dietary risk assessment, residues of pesticidal ingredients or their metabolites need to be evaluated for their genotoxic potential. The European Food Safety Authority recommend a tiered approach focussing assessment and testing on classes of similar chemicals. To characterise similarity and to identify structural alerts associated with genotoxic concern, a set of chemical sub-structures was derived for an example dataset of 66 triazole agrochemicals for which either Ames, chromosomal aberration or micronucleus test results are publicly available. This analysis resulted in a set of ten structural alerts that define the chemical space, in terms of the common parent and metabolic scaffolds, associated with the triazole chemical class. An analysis of the available profiling schemes for DNA and protein reactivity shows the importance of investigating the predictivity of such schemes within a well-defined area of structural space. Structural space alerts, covalent chemistry profiling and physico-chemistry properties were combined to develop chemical categories suitable for chemical prioritisation. The method is a robust and reproducible approach to such read-across predictions, with the potential to reduce unnecessary testing. The key challenge in the approach was identified as being the need for pesticide-class specific metabolism data as the basis for structural space alert development.

Sub-structure-based category formation for the prioritisation of genotoxicity hazard assessment for pesticide residues: Sulphonyl ureas.

In dietary risk assessment, residues of pesticidal ingredients or their metabolites need to be evaluated for their genotoxic potential. The European Food Safety Authority recommend a tiered approach focussing assessment and testing on classes of similar chemicals. To characterise similarity and to identify structural alerts associated with genotoxic concern, a set of chemical sub-structures was derived for an example dataset of 74 sulphonyl urea agrochemicals for which either Ames, chromosomal aberration or micronucleus test results are publicly available. This analysis resulted in a set of seven structural alerts that define the chemical space, in terms of the common parent and metabolic scaffolds, associated with the sulphonyl urea chemical class. An analysis of the available profiling schemes for DNA and protein reactivity shows the importance of investigating the predictivity of such schemes within a well-defined area of structural space. Structural space alerts, covalent chemistry profiling and physico-chemistry properties were combined to develop chemical categories suitable for chemical prioritisation. The method is a robust and reproducible approach to such read-across predictions, with the potential to reduce unnecessary testing. The key challenge in the approach was identified as being the need for pesticide-class specific metabolism data as the basis for structural space alert development.

COSMOS next generation - A public knowledge base leveraging chemical and biological data to support the regulatory assessment of chemicals.

The COSMOS Database (DB) was originally established to provide reliable data for cosmetics-related chemicals within the COSMOS Project funded as part of the SEURAT-1 Research Initiative. The database has subsequently been maintained and developed further into COSMOS Next Generation (NG), a combination of database and in silico tools, essential components of a knowledge base. COSMOS DB provided a cosmetics inventory as well as other regulatory inventories, accompanied by assessment results and in vitro and in vivo toxicity data. In addition to data content curation, much effort was dedicated to data governance - data authorisation, characterisation of quality, documentation of meta information, and control of data use. Through this effort, COSMOS DB was able to merge and fuse data of various types from different sources. Building on the previous effort, the COSMOS Minimum Inclusion (MINIS) criteria for a toxicity database were further expanded to quantify the reliability of studies. COSMOS NG features multiple fingerprints for analysing structure similarity, and new tools to calculate molecular properties and screen chemicals with endpoint-related public profilers, such as DNA and protein binders, liver alerts and genotoxic alerts. The publicly available COSMOS NG enables users to compile information and execute analyses such as category formation and read-across. This paper provides a step-by-step guided workflow for a simple read-across case, starting from a target structure and culminating in an estimation of a NOAEL confidence interval. Given its strong technical foundation, inclusion of quality-reviewed data, and provision of tools designed to facilitate communication between users, COSMOS NG is a first step towards building a toxicological knowledge hub leveraging many public data systems for chemical safety evaluation. We continue to monitor the feedback from the user community at support@mn-am.com.

Molecular fingerprint-derived similarity measures for toxicological read-across: Recommendations for optimal use.

Computational approaches are increasingly used to predict toxicity due, in part, to pressures to find alternatives to animal testing. Read-across is the "new paradigm" which aims to predict toxicity by identifying similar, data rich, source compounds. This assumes that similar molecules tend to exhibit similar activities i.e. molecular similarity is integral to read-across. Various of molecular fingerprints and similarity measures may be used to calculate molecular similarity. This study investigated the value and concordance of the Tanimoto similarity values calculated using six widely used fingerprints within six toxicological datasets. There was considerable variability in the similarity values calculated from the various molecular fingerprints for diverse compounds, although they were reasonably concordant for homologous series acting via a common mechanism. The results suggest generic fingerprint-derived similarities are likely to be optimally predictive for local datasets, i.e. following sub-categorisation. Thus, for read-across, generic fingerprint-derived similarities are likely to be most predictive after chemicals are placed into categories (or groups), then similarity is calculated within those categories, rather than for a whole chemically diverse dataset.

Mechanism-Based QSAR Modeling of Skin Sensitization.

Many chemicals can induce skin sensitization, and there is a pressing need for non-animal methods to give a quantitative indication of potency. Using two large published data sets of skin sensitizers, we have allocated each sensitizing chemical to one of 10 mechanistic categories and then developed good QSAR models for the seven categories that have a sufficient number of chemicals to allow modeling. Both internal and external validation checks showed that each model had good predictivity.

Chemical determinants of occupational hypersensitivity pneumonitis.

BACKGROUND: Workplace inhalational exposures to low molecular weight (LMW) chemicals cause hypersensitivity pneumonitis (HP) as well as the more common manifestation of respiratory hypersensitivity, occupational asthma (OA). AIMS: To explore whether chemical causation of HP is associated with different structural and physico-chemical determinants from OA. METHODS: Chemical causes of human cases of HP and OA were identified from searches of peer-reviewed literature up to the end of 2011. Each chemical was categorized according to whether or not it had been the attributed cause of at least one case of HP. The predicted asthma hazard was determined for each chemical using a previously developed quantitative structure-activity relationship (QSAR) model. The chemicals in both sets were independently and 'blindly' analysed by an expert in mech anistic chemistry for a qualitative prediction of protein cross-linking potential and determination of lipophilicity (log K ow). RESULTS: Ten HP-causing chemicals were identified and had a higher median QSAR predicted asthma hazard than the control group of 101 OA-causing chemicals (P < 0.01). Nine of 10 HP-causing chemicals were predicted to be protein cross-linkers compared with 24/92 controls (P < 0.001). The distributions of log K ow indicated higher values for the HP list (median 3.47) compared with controls (median 0.81) (P < 0.05). CONCLUSIONS: These findings suggest that chemicals capable of causing HP tend to have higher predicted asthma hazard, are more lipophilic and are more likely to be protein cross-linkers than those causing OA.

Proposal of an in silico profiler for categorisation of repeat dose toxicity data of hair dyes.

This study outlines the analysis of 94 chemicals with repeat dose toxicity data taken from Scientific Committee on Consumer Safety opinions for commonly used hair dyes in the European Union. Structural similarity was applied to group these chemicals into categories. Subsequent mechanistic analysis suggested that toxicity to mitochondria is potentially a key driver of repeat dose toxicity for chemicals within each of the categories. The mechanistic hypothesis allowed for an in silico profiler consisting of four mechanism-based structural alerts to be proposed. These structural alerts related to a number of important chemical classes such as quinones, anthraquinones, substituted nitrobenzenes and aromatic azos. This in silico profiler is intended for grouping chemicals into mechanism-based categories within the adverse outcome pathway paradigm.

Experimental verification of structural alerts for the protein binding of sulfur-containing compounds.

As often noted by Dr. Gilman Veith, a major barrier to advancing any model is defining its applicability domain. Sulfur-containing industrial organic chemicals can be grouped into several chemical classes including mercaptans (RSH), sulfides (RSR'), disulfides (RSSR'), sulfoxides (RS(=O)R'), sulfones (RS(=O)(=O)R'), sulfonates (ROS(=O)(=O)R') and sulfates (ROS(=O)(=O)OR'). In silico expert systems that predict protein binding reactions from 2D structure sub-divide these chemical classes into a variety of chemical reactive mechanisms and reactions which have toxic consequences. Using the protein binding profilers in version 3.1 of the OECD QSAR Toolbox, a series of sulfur-containing chemicals were profiled for protein binding potential. From these results it was hypothesized which sulfur-containing chemicals would be reactive or non-reactive in an in chemico glutathione assay and whether if reactive they would exhibit toxicity in excess of baseline in the Tetrahymena pyriformis population growth impairment assay. Subsequently, these hypotheses were tested experimentally. The in chemico data show that the in silico profiler predictions were generally correct for all chemical categories, where testing was possible. Mercaptans could not be assessed for GSH reactivity because they react directly with the chromophore 5,5'-dithiobis-(2-nitrobenzoic acid). With some exceptions, the major being disulfides, the in vitro toxicity data supported the in chemico findings.

Evaluation criteria for the quality of published experimental data on nanomaterials and their usefulness for QSAR modelling.

Nowadays nanotechnology is one of the most promising areas of science. The number and quantity of synthesized nanomaterials increase exponentially, therefore it is reasonable to expect that comprehensive risk assessment based only on empirical testing of all novel engineered nanoparticles (NPs) will very soon become impossible. Hence, the development of computational methods complementary to experimentation is very important. Quantitative structure-property relationship (QSPR) and quantitative structure-activity relationship (QSAR) models widely used in pharmaceutical chemistry and environmental science can also be modified and adopted for nanotechnology to predict physico-chemical properties and toxicity of empirically untested nanomaterials. All QSPR/QSAR modelling activities are based on experimentally derived data. It is important that, within a given data set, all values should be consistent, of high quality and measured according to a standardized protocol. Unfortunately, the amount of such data available for engineered nanoparticles in various data sources (i.e. databases and the literature) is very limited and seldom measured with a standardized protocol. Therefore, we have proposed a framework for collecting and evaluating the existing data, with the focus on possible applications for computational evaluation of properties and biological activities of nanomaterials.

Experimental verification of structural alerts for the protein binding of cyclic compounds acting as Michael acceptors.

This study outlines how a combination of and in vitro data can be used to define the applicability domain of selected structural alerts within the protein binding profilers of the Organisation for Economic Co-operation (OECD) Quantitative Structure-Activity Relationship (QSAR) Toolbox. Thirty chemicals containing a cyclic moiety were profiled for reactivity using the OECD and Optimised Approach based on Structural Indices Set (OASIS) protein binding profilers. The profiling results identified 22 of the chemicals as being reactive towards proteins. Analysis of the experimentally data showed 19 of these chemicals to be reactive. Subsequent analysis allowed refinements to be suggested to improve the applicability domain of the structural alerts investigated. The accurate definition of the applicability domain for structural alerts within in silico profilers is important due to their use in chemical category in predictive and regulatory toxicology.

Hepatotoxicity: a scheme for generating chemical categories for read-across, structural alerts and insights into mechanism(s) of action.

The ability of a compound to cause adverse effects to the liver is one of the most common reasons for drug development failures and the withdrawal of drugs from the market. Such adverse effects can vary tremendously in severity, leading to an array of possible drug-induced liver injuries (DILIs). As a result, it is not surprising that drug development has evolved into a complex and multifaceted process including methods aiming to identify potential liver toxicities. Unfortunately, hepatotoxicity remains one of the most complex and poorly understood areas of human toxicity; thus it is a significant challenge to identify potential hepatotoxins. The performance of existing methods to identify hepatotoxicity requires improvement. The current study details a scheme for generating chemical categories and the development of structural alerts able to identify potential hepatotoxins. The study utilized a diverse 951-compound dataset and used structural similarity methods to produce a number of structurally restricted categories. From these categories, 16 structural alerts associated with observed human hepatotoxicity were developed. Furthermore, the mechanism(s) by which these compounds cause hepatotoxicity were investigated and a mechanistic rationale was proposed, where possible, to yield mechanistically supported structural alerts. Alerts of this nature have the potential to be used in the screening of compounds to highlight potential hepatotoxicity, whilst the chemical categories themselves are important in applying read-across approaches. The scheme presented in this study also has the potential to act as a knowledge generator serving as an excellent starting platform from which to conduct additional toxicological studies.

The definition of the toxicologically relevant applicability domain for the SNAr reaction for substituted pyridines and pyrimidines.

This study outlines how results from a glutathione reactivity assay (so-called in chemico data) can be used to define the applicability domain for the nucleophilic aromatic substitution (SNAr) reaction for nitrogen-containing aromatic compounds. SNAr is one of the six mechanistic domains that have been shown to be important in toxicological endpoints in which the ability to bind covalently to a protein is a key molecular initiating event. This study has analysed experimental data (2 h RC50 values), allowing a clear and interpretable structure-activity relationship to be developed for pyridines and pyrimidines which reside within the SNAr domain. The in-ring nitrogen(s) act as activating groups in the SNAr reaction. The position(s) of the in-ring nitrogen(s) as well as other activating groups, especially in relationship to the leaving group, affect reactive potency. The experimentally defined applicability domain has resulted in a series of structural alerts. These results build on early work on the benzene derivatives residing in the SNAr domain. The definition of the applicability domain for the SNAr reaction and the resulting structural alerts are likely to be beneficial in the development of computational tools for category formation and read-across in hazard identification, and the development of adverse outcome pathways.

Experimental verification, and domain definition, of structural alerts for protein binding: epoxides, lactones, nitroso, nitros, aldehydes and ketones.

This study outlines how a combination of in chemico and Tetrahymena pyriformis data can be used to define the applicability domain of selected structural alerts within the profilers of the OECD QSAR Toolbox. Thirty-three chemicals were profiled using the OECD and OASIS profilers, enabling the applicability domain of six structural alerts to be defined, the alerts being: epoxides, lactones, nitrosos, nitros, aldehydes and ketones. Analysis of the experimental data showed the applicability domains for the epoxide, nitroso, aldehyde and ketone structural alerts to be well defined. In contrast, the data showed the applicability domains for the lactone and nitro structural alerts needed modifying. The accurate definition of the applicability domain for structural alerts within in silico profilers is important due to their use in the chemical category in predictive and regulatory toxicology. This study highlights the importance of utilizing multiple profilers in category formation.

Predicting skin sensitization potency for Michael acceptors in the LLNA using quantum mechanics calculations.

This study outlines the development of a series of quantitative mechanistic models enabling skin sensitization potency in the LLNA to be predicted for direct acting Michael acceptors. These models utilized several computational descriptors based on knowledge of the Michael addition reaction mechanism. The key descriptor was calculated using density functional theory and modeled the stability of the reaction intermediate. A second descriptor relating to the available surface area at the site of the reaction was also found to be important. Several poorly predicted compounds were identified, and in all cases, these could be rationalized mechanistically. The analysis of these compounds allowed a well-defined mechanistically driven applicability domain to be developed. The study showed that in silico quantitative mechanistic models, with a well-defined applicability domain, can be used to predict skin sensitization potency in the LLNA. The approach presented has the potential to be of use as part of a weight of evidence approach for predicting skin sensitization without the use of animals in risk assessment.

Development of mechanism-based structural alerts for respiratory sensitization hazard identification.

This study outlines how mechanistic organic chemistry related to covalent bond formation can be used to rationalize the ability of low molecular weight chemicals to cause respiratory sensitization. The results of an analysis of 104 chemicals which have been reported to cause respiratory sensitization in humans showed that most of the sensitizing chemicals could be distinguished from 82 control chemicals for which no clinical reports of respiratory sensitization exist. This study resulted in the development of a set of mechanism-based structural alerts for chemicals with the potential to cause respiratory sensitization. Their potential for use in a predictive algorithm for this purpose alongside an externally validated quantitative structure-activity relationship model is discussed.

The definition of the applicability domain relevant to skin sensitization for the aromatic nucleophilic substitution mechanism.

This study outlines how a glutathione reactivity assay (so-called in chemico data) can be used to define the applicability domain for the nucleophilic aromatic substitution (S(N)Ar) reaction for benzenes. This reaction is one of the six mechanistic domains that have been shown to be important in toxicological endpoints in which the ability to bind covalently to a protein is a key molecular initiating event. This study has analysed the experimental data, allowing a clear and interpretable structure-activity relationship to be developed for the S(N)Ar domain. The applicability domain has resulted in a series of structural alerts. The definition of the applicability domain for the S(N)Ar reaction and the resulting structural alerts are likely to be beneficial in the development of computational tools for category formation and read-across. The study concludes with how this information can be used in the development of adverse outcome pathways.

Development of new structural alerts suitable for chemical category formation for assigning covalent and non-covalent mechanisms relevant to DNA binding.

The need to assess the ability of a chemical to act as a mutagen is one of the primary requirements in regulatory toxicology. Several pieces of legislation have led to an increased interest in the use of in silico methods, specifically the formation of chemical categories and read-across for the assessment of toxicological endpoints. One of the key steps in the development of chemical categories for mutagenicity is defining the mechanistic organic chemistry associated with the formation of a covalent bond between DNA and an exogenous chemical. To this end this study has analysed, by use of a large set of mutagenicity data (Ames test), the mechanistic coverage of a recently published set of in silico structural alerts developed for category formation. The results show that the majority of chemicals with a positive result in the Ames test were assigned at least one covalent binding mechanism related to the formation of a DNA adduct. The remaining chemicals with positive data in the Ames assay were subjected to a detailed mechanistic analysis from which 26 new structural alerts relating to covalent binding mechanisms were developed. In addition, structural alerts for radical and non-covalent intercalation mechanisms were also defined. The structural alerts outlined in this study are not intended to predict mutagenicity but rather to identify mechanisms associated with covalent and non-covalent DNA binding. This mechanistic profiling information can then be used to form chemical categories suitable for filling data gaps via read-across. A strategy for chemical category formation for mutagenicity is also presented.

A review of the electrophilic reaction chemistry involved in covalent protein binding relevant to toxicity.

Several pieces of legislation have led to an increased interest in the use of in silico methods, specifically the formation of chemical categories for the assessment of toxicological endpoints. For a number of endpoints, this requires a detailed knowledge of the electrophilic reaction chemistry that governs the ability of an exogenous chemical to form a covalent adduct. Historically, this chemistry has been defined as compilations of structural alerts without documenting the associated electrophilic chemistry mechanisms. To address this, this article has reviewed the literature defining the structural alerts associated with covalent protein binding and detailed the associated electrophilic reaction chemistry. This information is useful to both toxicologists and regulators when using the chemical category approach to fill data gaps for endpoints involving covalent protein binding. The structural alerts and associated electrophilic reaction chemistry outlined in this review have been incorporated into the OECD (Q)SAR Toolbox, a freely available software tool designed to fill data gaps in a regulatory environment without the need for further animal testing.