Insights into Health Canada's TTC-Based Approach

1. 1) Overview of the Health Canada Threshold of Toxicological Concern (TTC)-based Approach for Certain Substances under the Third Phase of the CMP2) Recent application and development related to Eco-TTC at Environment and Climate Change Canada Canadian Network for Human Health and the Environment (CNHHE) Webinar May 16, 2019

2. TTC is an approach to establish a human exposure threshold value for a chemical, below which there is a low probability of risk to human health (Kroes et al. 2004). Makes use of relatively large historical databases of toxicity information, including distributions of critical effect levels (e.g. NOAELs, TD05) and structural information to derive the threshold values for classes of chemicals. The TTC concept was first applied as the Threshold of Regulation for food contact materials by the Unites States Food and Drug Administration (US FDA, 1995). This concept evolved over time with the development of multiple TTCs based on the analysis of additional classes of chemicals and additional health effects, including cancer and non-cancer effects. What is the Threshold of Toxicological Concern (TTC)?

3. The TTC concept has been acknowledged to be a science-based prioritization and risk assessment tool by different organizations and is currently used or proposed in several areas of chemical risk assessment internationally, including: • Food contact articles (US FDA 1995), • Food flavouring agents under the Joint FAO/WHO Expert Committee on Food Additives (JECFA) (WHO 1995; WHO 1997), • Mutagenic impurities in pharmaceutical preparations (ICH 2017) The European Food Safety Authority (EFSA) Scientific Committee and TTC Expert Workshop (WHO/EFSA) have recommended the approach as a scientifically-valid method for deciding if the exposure to a substance is so low that the probability of adverse health effects is low; and accordingly, an effective tool to prioritize further testing and assessment (EFSA 2012, 2016). TTC Use Internationally

4. 2003 Europe ILSI workshop Produced a decision framework for trace substances in food, integrating cancer & non-cancer effects Incorporates 5 high potency structural groups alerts that were not suitable for TTC approach (Cohort of Concern) Explored applicability to potentially sensitive endpoints (immunotoxicity, teratogenicity, neurotoxicity, allergenicity) • Established an organophosphate-specific TTC value for neurotox • Excluded allergens from TTC approach • Concluded that acceptable protection was provided for other endpoints Establishing TTC Values from Databases of Toxicity Data (Kroes et. al. 2004 reproduced with permission)

5. Cancer Potency Database 700+ Substances (CPDB 1999) Establishment of the dose giving a 50% tumour incidence (TD50) using data for the most sensitive species and most sensitive site. Simple linear extrapolation from the TD50 to a 1 in 106 lifetime tumor risk level in exposed individuals (virtual safe dose). The majority of these chemicals with TD50 values would not exceed the level of 1 in 106 risk level at intakes of 0.0025 ug/kg bw/day. TTC Values for Potential Genotoxic Carcinogens (Cheeseman et al., 1999 reproduced with permission)

6. Grouped the database based on structural alerts for carcinogenicity and mutagenicity (Ashby and Tennant 1991 and others) While looking at the structural alerts for carcinogenicity, Kroes et al. (2004) identified a “cohort of concern” Five groups had a significant fraction of their members that may still be of concern at an intake of 0.15 µg per person per day (0.0025 µg/kg bw/day); therefore proposed as exclusion groups: • Aflatoxin-like compounds • Azoxy compounds • N-nitroso compounds • Steroids • Tetrahalogenateddibenzodioxins and dibenzofurans High Potency Carcinogens “Cohort of Concern”

7. Establishment of tiered thresholds to account for repeat dose, developmental and reproductive effects based on chemical structure (Munro et al. 1996). Assumes non-genotoxic carcinogens are also covered (i.e. threshold effects). 613 chemicals in reference database, which contained ~2900 NOELs • Substances included industrial/agricultural/consumer chemicals, pharmaceuticals, food substances • Database included oral sub-chronic, chronic, reproductive and developmental toxicity studies in rodents and rabbits • The most conservative (species/sex/endpoint) NOEL was identified for each substance in the reference database • Substances were then separated on the basis of Cramer structural class and a UF of 100 (300 for sub-chronic studies) was applied to at TTC values for non-carcinogenic effects for each Cramer class Cramer Chemical classification scheme (for defined organics) consists of 33 ordered questions intended to help predict toxicity (Cramer et al. 1978) • Class I: suggests low oral toxicity (“innocuous”), simple structures with efficient metabolism • Class II: “less innocuous” but no structural features suggesting toxicity • Class III: not strongly presumed to be safe; may suggest toxicity or have reactive functional groups TTC Values for Other Effects

8. TTC Values for Other Effects • These derived TTC values are explicitly limited to structurally defined chemicals with no evidence of being a potential carcinogen with a genotoxic MOA (i.e DNA-reactive mutagens) • Munro et. al (1999) and Kroes et al. (2004) conducted an analysis of organophosphates (OPs) based on neurotoxicity and specified a threshold for these chemicals that was lower than Class III. • Papers published after Kroes et al (2004) that consider different databases support the values published by Kroes et al. as being similar or conservative. • Kalkhof, H et al. 2011. Threshold of toxicological concern values for non-genotoxic effects in industrial chemicals: re-evaluation of the Cramer classification. Arch Toxicol • Tluczkiewicz, I et al. 2011. Improvement of the Cramer classification for oral exposure using the database TTC RepDose – A strategy description. Regul. Toxicol. Pharmacol.

9. A group of 237 candidates amongst the 1500 remaining CMP3 priority substances was identified via qualitative characterization of uses and exposure potential. Estimated exposure values were compared to assigned TTC values for each substance. Substances which had exposure below the assigned TTC value were considered to be of low concern for human health. Substances found to have exposure exceeding the TTC value were set aside to be considered further under a different initiative under CMP. Health Canada (TTC)-based Approach

10. Started with the Kroes et al. 2004 decision tree and TTC values Modified based on recent recommendations from EFSA Scientific Committee (EFSA 2012) and/or TTC Expert Workshop (WHO/EFSA, 2016) • Expanded exclusion criteria • Inclusion of carbamates in the threshold for OPs • Using the Cramer III threshold for Cramer II substances due to limited number of Cramer II chemicals in Munro dataset HC approach underwent external written peer review. Final content and outcome of the TTC-based approach remain the responsibility of Health Canada Health Canada Decision Tree to Assign TTC values

11. Health Canada CMP TTC Decision Tree https://www.ec.gc.ca/ese-ees/326E3E17-730A-4878-BC25-D07303A4DC13/HC%20TTC%20SciAD%20EN%202017-03-23.pdf

12. Health Canada CMP TTC Decision Tree GT Alerts / Predictive Modeling Phase GT Data Gathering Phase Flow chart operationalized in computational workflow with OECD Toolbox and OASIS TIMES Models to assign TTC value

13. For purposes of this TTC-based approach, a chemical is considered potentially genotoxic if: • a) positive results in in vivo mammalian studies for gene mutation or chromosomal aberrations (including micronuclei); and/or • b) positive results in an in vitro test for gene mutation or chromosomal aberrations (including micronuclei) and absence of conflicting data from in vivo mammalian tests that covers the same genetic endpoint that gave a positive response in the in vitro test. Where multiple results for a given assay were available within the OECD QSAR Toolbox, the response was assigned based on the most conservative outcome. Compounds that did not have positive empirical data for genotoxicity (or had no data) were evaluated for structural alerts related to DNA reactivity using computational models that included a metabolic simulator (OASIS TIMES (Ames and In vitro Chrom. Ab.) (details Appendix A). The approach was considered conservative resulting in a high proportion of chemicals being assigned a TTC of 0.0025 µg/kg-bw/day. Genotoxicity Assessment

14. After the genotoxicity assessment, the structures of the negative substances were screened using a structural profiler to determine if the compound was an organophosphate or carbamate (note - none of the candidates in this approach were organophosphates or carbamates). Cramer classification under the CMP approach was based on the software implementation of the 33 questions in the OECD QSAR Toolbox (v3.3). Toxtree was also used to verify Cramer class for each substance outside the computational workflow. Discrepancies were manually examined. Cramer Classification

15. The candidates selected for the TTC approach were associated with a narrow range of uses, including those used primarily in industrial processes or found in low concentration in products (exposure to the general population was expected to be limited). Exposure was estimated using same approaches as for any other CMP substance. Exposure sources for the general population included environmental media and through product uses, such as one fragrance ingredient in cosmetics, food flavouring and food packaging, lubricants and one adhesive. Estimated Exposure

16. For environmental media exposure, a fugacity model was used to predict environmental concentrations with conservative parameters (e.g., use of total volume in commerce). When necessary, a refinement such as sewage treatment plant (STP) removal fraction was incorporated. For exposure from the use of consumer products, the primary route of exposure via the dermal route; substancesin products for which use would be associated with significant inhalation exposure potential were not part of this approach. Estimated Exposure

17. Of the 237 candidate substances 89 were considered to have exposures lower than their respective TTC values (~63% fail rate). The remaining 148 substances of the 237 candidates were either excluded or had exposure estimates that exceeded the TTC values; these substances are undergoing further assessment under separate initiatives. High proportion (~35%) of chemicals assigned the lowest TTC value (0.0025 µg/kg-bw/day) indicating the conservative nature of the genotoxicity assessment portion of the workflow (i.e. general bias towards being protective in this screening approach). Approach was applied as a one off rapid screen in CMP3. May be used in post-2020 CMP to facilitate setting priorities but it is envisioned to be an ‘evergreen’ approach that will adjust as TTC improvements are made internationally. Overview of Results

18. References Barlow S. 2005. Threshold of toxicological concern (TTC). ILSI Europe Concise Monograph Series. p.1– 31. Available from: http://ilsi.eu/wp-content/uploads/sites/3/2016/06/C2005Thres_Tox.pdf Cheeseman MA, Machuga EJ, Bailey AB. 1999. A tiered approach to threshold of regulation. Food ChemToxicol 37:387–412. Cramer GM, Ford RA, Hall RL. 1978. Estimation of toxic hazard – a decision tree approach. Food CosmetToxicol 16:255–276. [EFSA] European Food Safety Authority. 2012. Scientific opinion on exploring options for providing advice about possible human health risks based on the concept of Threshold of Toxicological Concern (TTC). EFSA Journal 10(7):2750. Available from: http://www.efsa.europa.eu/sites/default/files/scientific_output/files/main_documents/2750.pdf [EFSA] European Food Safety Authority / [WHO] World Health Organisation. 2016. Review Threshold of toxicological concern (TTC) and development of new TTC decision tree. Available fr [ICH] International Conference on Harmonisation. 2017. ASSESSMENT AND CONTROL OF DNA REACTIVE (MUTAGENIC) IMPURITIES IN PHARMACEUTICALS TO LIMIT POTENTIAL CARCINOGENIC RISK. International Conference on Harmonisation of Technical Requirements for Registration of Pharmaceuticals for human use. http://www.ich.org/fileadmin/Public_Web_Site/ICH_Products/Guidelines/Multidisciplinary/M7/M7_R1_Addendum_Step_4_31Mar2017.pdf Kroes R, Renwick AG, Cheeseman M, Kleiner J, Mangelsdorf I, Piersma A, Schilter B, Schlatter J, van Schothorst F, Vos JG, Wurtzen G. 2004. Structure-based thresholds of toxicological concern (TTC): guidance for application to substances present at low levels in the diet. Food ChemToxicol 42:65–83 Munro IC, Kennepohl E, Kroes R 1999. A procedure for the safety evaluation of flavouring substances. Food ChemToxicol 37:207–232. [US FDA] Food and Drug Administration.1995. Federal Register Food Additives; Threshold of Regulation for substances used in food contact articles. Department of Health and Human Services, Food and Drug Administration, 21 CFR Parts 5, 25, 170,171, and 174. Docket Nos 77P-0122, 92N-0181. Available from: http://www.gpo.gov/fdsys/pkg/FR-1995-07-17/pdf/95-17435.pdf [WHO] World Health Organization. 1995. Evaluation of certain food additives and contaminants (Fortyfourth report of the Joint FAO/WHO Expert Committee on Food Additives). WHO Technical Report Series 859. Geneva, Switzerland. [WHO] World Health Organization. 1997. Evaluation of certain food additives and contaminants (Fortysixth report of the Joint FAO/WHO Expert Committee on Food Additives). WHO Technical Report Series 868. Geneva, Switzerland.

19. 2) Recent application and development related to Eco-TTC at Environment and Climate Change Canada

20. Environment and Climate Change Canada (ECCC) implemented a tissue-residue based eco-TTC into a prioritization approach known as the Ecological Risk Classification (ERC) approach (v1.0 and v2.0) ECCC has also been involved in the development of eco-TTC tools and science via HESI Subcommittee on Animal Alternatives* • Ottawa Workshop fall 2017 • Online eco-TTC tool with curated aquatic database Environment and Climate Change Canada and Eco-TTC *Health and Environmental Sciences Institute (HESI) https://hesiglobal.org/animal-alternatives-in-environmental-risk-assessment/

21. An eco-TTC can also be viewed on a whole body tissue residue basis The eco-TTCinternal is associated with median lethality and has been derived from hundreds of data points for select modes of action (MoA) For chemicals acting via a narcotic MoA, the critical body residue associated with lethality (CBR50) ranges from: • 2-8 mmol/kg acute (McCarty and Mackay 1993) • 0.2-0.8 mmol/kg chronic (McCarty and Mackay 1993) CBRs for other eco-MoAs are known (Esher et al. 2011) ERC v1 (ECCC 2016) used an inverse modeling approach to determine eco-MoA according to the estimated CBR from aquatic acute lethality data (narcosis vs specifically acting) ERC and Tissue Residue Eco-TTC

22. Tissue eco-TTCs are also known for Lethal Chemical Activity (LA50) and Critical Membrane Concentrations (CMC50) CBR50, LA50 and CMC50 have been calculated to determine MoA and compared to a consensus QSAR-based MoA to determine an overall consensus MoA Calculated for ~12000 organic chemicals ERC v2 and Tissue Residue Eco-TTC

23. HESI Eco-TTC

24. Eco-TTC & EnviroTox Platform www.EnviroToxDatabase.org Database of ~91K curated aquatic toxicity records User-friendly database filtering interface • Freely available analysis tools: • PNEC calculator (US & Europe) • ecoTTC distribution tool • Chemical Toxicity Distribution (CTD) tool Developed via a global, collaborative partnership with government, academia, and industry, managed by HESI

25. Eco-TTC Calculation ecoTTC

26. Eco-TTC Calculation Application Factors ecoTTC Predicted No Effect Concentration

27. Chemical Toxicity Distribution No Application Factors 50% 25% 10% 5% 1% Effect Concentration

28. EnviroToxDatabase.org membry@hesiglobal.org Search filters Target data PNEC Calculator Application Factors PNEC Calculator No Application Factors CTD Calculator CTD Calculator ecoTTC

29. Publications / Links

30. Questions?

31. Appendix A Models used to identify substances (including predicted metabolites) with structural alerts for genotoxicity