Table 1.

Applications of in silico toxicology

In silico toxicology application	Discussion
1. Alternative to test data.	The use of non-animal alternative methods including in silico approaches, may substitute for other types of tests in regulatory submissions in certain cases. Acceptable alternative methods for filling data gaps are outlined in Annex XI of the European Union’s REACH regulation (EU 2006). In the United States, Frank R. Lautenberg Chemical Safety for the 21st Century Act revised the Toxic Substances Control Act (TSCA) to include predictive models and expert review as part of an overall assessment (TSCA 2016). The United States Food and Drug Administration (US FDA) Center for Devices and Radiological Health (CDRH) issued a guidance for industry and FDA staff. This guidance is on the use of International Standard ISO 10993-1 for biological evaluation of medical devices and indicates in the absence of experimentally derived carcinogenicity information, structure activity relationship modeling for these materials may be needed (CDRH 2016). The FDA draft guidance on Electronic Nicotine Delivery Devices (ENDS) also discusses the use of computational toxicology models in the absence of toxicological data for potential toxicants created by the aerosolization process (PMTA/FDA 2016). When chemicals with limited toxicity data are required to be classified and labeled for shipping or other purposes, in silico toxicology provides an alternative method for quickly filling the data gaps in the toxicity/safety information, such as predictions of acute toxicity to support assignment to the Globally Harmonized System of Classification and Labelling category (Freidig et al., 2007; ECHA 2015).
2. As part of the weight-of-evidence in regulatory submissions.	There are currently several regulatory frameworks where only specific laboratory tests for an endpoint of concern may be submitted (such as for drugs or food additives). However, in such cases, in silico predictions can be submitted alongside standard toxicological data to complement the assessment. This may include in silico assessments provided as supporting data or adjuncts to the primary in vivo or in vitro studies to give a mechanistic understanding of the observed results and/or allow a better definition of experimental needs. Additionally, in silico methods may be used to guide or prioritize in vitro testing (EU 2012). The European Union’s Cosmetics Regulation (EU 2009a) prohibits the use of animal testing for products or ingredients and a complete marketing ban of such products tested as a whole or containing tested ingredients. This requires the use of alternative methods, such as IST, in the assessment of new cosmetics ingredients. In a recent memorandum, the European Commission’s Scientific Committee for Consumer Safety (SCCS), which is responsible for the risk assessment of cosmetic ingredients, acknowledged the importance and limitations of in silico methods; the SCCS recommended that in silico methods be used either for internal decision making or as part of a weight-of-evidence (WOE) approach to estimate toxicity risks before embarking on any experimental testing (SCCS 2016).
3. Mixtures assessment.	Most exposures are not to a single chemical but rather to complex mixtures of chemicals that may be found in food, beverages, the environment, cigarette smoke, electronic nicotine delivery systems (ENDS) aerosols, botanical drugs or natural products. In certain situations, it may be possible to use in silico methods to assess individual components since today’s in silico analysis can only be performed on discrete identifiable chemicals. While preliminary analytical work is required to identify all chemicals in the mixture above appropriate Analytical Evaluation Thresholds (AET) (Ball and Norwood 2012), leveraging in silico approaches may avoid having to synthesize or purify each of the potentially large number of mixture components to perform standard toxicological tests (Mumtaz et al., 2010). Careful consideration is required for mixtures when there are multiple chemicals for interactions, such as synergistic or additive effects that may have the same, similar or different mechanisms of action (MOA).
4. Assessment of impurities and degradation products.	Chemicals, such as pharmaceuticals or plant protection products, may contain low levels of impurities produced during manufacturing and degradation. Many such substances, when present at levels above accepted thresholds, need to be assessed. In most cases, mutagenicity evaluation of the impurity under question is required as a first step of the risk assessment. (Harvey et al., 2017) The ICH M7 guideline provides specific recommendations for assessing drug impurities (ICH M7, 2017(R1)), including the use of two complementary computational toxicology methodologies (i.e., statistical-based and expert rule-based models) to predict bacterial mutagenicity.
5. Residues of plant protection products.	Residues of plant protection products may be evaluated as a part of residue definition for dietary risk assessment of plant protection products (EU 2009b). In this context, in silico methods provide a useful alternative approach. (EFSA 2016)
6. Assessment of extractables and leachables.	Medical devices, such as inhaled aerosols, food-contact substances, and consumer product packaging materials may pose a risk for human health due to release of potentially harmful chemicals that are used in the production of the components (Bossuyt et al., 2017). These include plasticizers, copolymers, vulcanization additives, etc. for which toxicological data is often lacking but where a risk assessment must be performed. A migration or leachables study supports the discovery, identification, and quantification of any leachables. An in silico toxicological assessment, in certain situations, can provide sufficient data for the risk assessment.
7. Workers’ safety and occupational health.	Chemicals used in the manufacture of a product are assessed for mutagenicity, carcinogenicity, skin and respiratory sensitization, irritation (skin, eye and respiratory), and reproductive and developmental toxicity and possibly acute toxicity. In silico assessments make it possible to estimate the potential toxicity of chemicals and adopt proper engineering controls and personal protective equipment usage to protect workers who could be exposed to these substances during production, transfer, storage, and delivery processes (EU 2006). In silico approaches have been utilized to assess these major toxicological endpoints in the occupational safety setting. In silico methods to predict respiratory sensitization potential of industrial chemicals have recently been reviewed by Seed and Agius (2017).
8. Metabolite analysis.	Metabolites can present an increased or decreased risk of local or systemic toxicity compared with the parent chemical (Mumtaz and Durkin, 1992). While reactive or toxic metabolites may be formed by an organism, their identification, separation as well as possible synthesis for testing purposes may be challenging. In silico methods provide a practical alternative approach to understanding the safety profiles of this potentially large number of chemicals as well as to support the prediction of metabolites.
9. Ecotoxicology.	Various chemicals are discharged into the environment that may cause harm. Furthermore, the parent compounds can be transformed by hydrolysis, redox-reactions, or photolysis into numerous additional chemicals. IST methods often provide the most practical approach to assess the potential effects on the environment and wildlife species of the many chemicals that are discharged. Prediction of physicochemical parameters supports assessment of potential environment exposure to the chemical (e.g., persistence and distribution). As an example, Chen at al., 2015 describes the use of in silico assessment of potentially hazardous contaminants present in water.
10. Green chemistry and safer alternatives.	In silico methods can play an important role when identifying alternative chemicals that may have a safer profile than existing chemicals (Rastogi et al., 2014). This includes, for example, alternatives for use in manufacturing processes, alternative packaging/delivery materials and the use of specific additives. In silico methods can provide insights about structural features responsible for the toxicity of different groups of chemicals and thereby allow for the rational design of intrinsically safer chemicals.
11. Selection of product development candidates.	In early product discovery or development, many thousands of compounds may be evaluated. In silico methods may provide a helpful approach to selecting candidates, since in silico methods are inexpensive, rapid to perform, and high throughput. In addition, in silico methods can suggest which molecular substructures (toxicophores) are responsible for the predicted toxic activity, thereby supporting the optimization of future compounds (Hillisch et al., 2015; Myatt et al., 2016). Later in the product development process, a smaller number of chemicals may be selected as candidates to take forward for further development; in normal situations, preference would be given to the candidate(s) with the most advantageous safety profile(s) (Myatt et al., 2016).
12. Emergency response situations.	When one or more chemicals are unexpectedly released into the environment (e.g., the West Virginia chemical spill (NTP 2016)) or into a production process, it is important to quickly evaluate the potential effects on humans, wildlife, and the environment. In such emergency situations the toxicological profile of the released chemicals needs to be established as quickly as possible to support the proper emergency response and to protect emergency services staff and bystanders (Hochstein et al., 2008; Schilter, et al., 2014). In such a limited timeframe and in the absence of previously generated data, in silico approaches may be a practical option for rapid hazard identification.
13. Prioritizing testing of chemicals.	In silico approaches can help prioritize in vitro and in vivo toxicology testing, based upon the chemical’s exposure and prediction of toxicity; they are an important aspect of the work at several organizations such as the US EPA, National Toxicology Program, Environment and Climate Change Canada and ECHA (Schwetz 1995). In silico methods may be used to prioritize (based on potential toxicological liabilities) the order in which a series of toxicological studies will be performed (Myatt et al., 2016).
14. Rationalization of in vivo or in vitro study results.	As mentioned previously in the description of the in silico application titled “As part of the weight-of-evidence in regulatory studies”, results from quantitative structure-activity relationship (QSAR) models (toxicophore information, chemical fragments or physicochemical properties) may be used in conjunction with biological data to infer a mechanism of action (MOA), molecular initiating event (MIE), or mode of toxicity as part of an adverse outcome pathway (AOP) (Martin et al., 2015; Ellison et al., 2016). Information from in silico methods can also be used to tailor an in vivo study, e.g., by inclusion of additional endpoints. When existing experimental data on a compound are equivocal or when not all relevant safety information are available or accessible, in silico data may be used as additional information as part of the weight- of-evidence approach in reaching a more informed decision (Kruhlak et al., 2012).