Assessment of chemicals, using and developing New Approach Methodologies (NAMs)

ECHA’s approach to incorporate NAMs into regulatory framework

Tomasz Sobanski

European Chemicals Agency

Why ECHA is interested in NAMs?

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We understand NAMs as all methods (in silico, in chemico, in vitro and in vivo) that (potentially) can enhance the pace of our work, to have better informed, more relevant decisions and reduce/replace the need for studies on (vertebrate) animals, with a main focus on high tier human health and environmental ‘endpoints’.

Enhancement mentioned above can be in terms of:

• Throughput and/or

• Robustness and/or;

• Bringing mechanistic knowledge and/or;

• Providing appropriate protection levels for human health and environment

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Projection of data availability (REACH)

• Overall, during the 10 years of evaluation, ECHA checked the compliance of 1780 dossier (Annex IX,X). In the vast majority of the cases (overall 71%) the compliance checks have confirmed one or more non-compliances (Art 54 Report 2018).

• Most frequent deficiencies related to alternatives:

• lack of mechanistic evidence to support predictions (toxicokinetics)

• shortcomings in the toxicological hypothesis.

ECHA Strategic Plan 2019-2023 (draft)

• Strategic Priority 1: Identification and risk management of substances of concern

• Increase data availability for prioritising data poor substances with an aligned strategy for further generation and use of data from new approach methodologies (NAMs).

• ECHA considerations on the World Summit of Sustainable Development (WSSD) 2020 goal

• Success factor 1. Robust data is available on all chemicals in Europe:

b). Hazard data is generated using non-animal testing methods and new approaches wherever possible.

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Outcome from the 3rd Report on The Use of Alternatives to Testing on Animals for the REACH Regulation (Article 117.3 Report)

• The findings from the analysis in this report of the extensive use of adaptations, particularly for high tier endpoints, when taken together with deficiencies identified in their use, will be used to help ECHA further refine its efforts to promote the proper use of alternative methods and to support further scientific development. As was mentioned before, to support proper use of adaptations for high tier human health and environmental endpoints, additional supporting evidence related to mechanisms of toxicity or toxicokinetics will often need to be generated.

• ECHA sees potential in new approach methodologies (NAMs) in the longer term, as these methods are based on models able to detect specific mechanisms of toxicity, provide kinetic information and be run in a high-throughput manner. Therefore, ECHA will actively seek to benefit from, and support, the scientific developments of methods that could ultimately limit or replace the need for new studies in (vertebrate) animals in the long term.

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NAM related activities in ECHA

• NAM integration into screening activities

• Some assays already used (endocrine disruption)

• Application of NAMs to support Chemical Categorization

• Focus on hazard, but also exploring Exposure ‘NAM’

• EUToxRisk http://www.eu-toxrisk.eu/

• ECHA follows and contributes where appropriate

• Focus on Read-Across Case studies

• In addition supporting (via the JRC) the design of an ‘Ab initio’ case study

• Accelerating the Pace of Chemical Risk

• ECHA follows and contributes to several case studies

• Currently focus on hazard and ‘mapping’ between NAM and classical methods

http://www.eu-toxrisk.eu/

Application of NAMs to support Chemical Categorization

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Problem space: Grouping and read-across are frequently used for human health and environmental risk assessments. Traditionally, substances are grouped based on structural / physical-chemical parameters, not using any mechanistic biological response.

Purpose of this project: Evaluate the effectiveness of omics technologies and machine learning to derive molecular data for mechanism-driven substance grouping. Specifically, to use similarities/dissimilarities of the molecular responses to substantiate (or not) a grouping hypothesis derived from traditional QSAR approaches.

Substantiating Chemical Categories with Omics-derived Mechanistic Evidence*

*SC-01 under ECHA framework contract “Services related to metabolomics measurements and multi-omics data interpretation”, Contract notice: 2018/S 159-363671 (September 2018)

Key features

Instrumentation: liquid

handling robots, 7 UHPLC-MS

for discovery metabolomics, 3

UHPLC-MS/MS for targeted

metabolite analysis, DI-MSn for

metabolite identification, 2 NMR

spectrometers for discovery

metabolomics

Standardised workflows: for

both sample analysis and

computational work, with

rigorous QA/QC

Scale: capacity for 20,000

targeted and 30,000 non-targeted

assays per year.

http://www.wikiwand.com/en/Daphnia_magna

Chemical grouping project utilises Daphnia as an invertebrate test organism

Project leaders at UoB:Prof Ben Brown – biostatistics, machine learning, artificial intelligenceProf John Colbourne – environmental genomics, transcriptomics, phylogenetics, pathway conservationProf Mark Viant – metabolomics, toxicology, QA/QC, reporting standards

1. Substance selection:

Structure based grouping to generate a similarity/dissimilarity map of a large number of substances;

2. Dose range-finding experiments in Daphnia:

Range-finding studies with Daphnia to determine the “equi-effective” concentrations

3. Exposures, transcriptomics and metabolomics analyses:

Obtain mechanistic evidence indicating weather category of substances is consistent for aquatic toxicity (invertebrates).

4. Elucidating mechanism:

Attempt to identify perturbed molecular signatures by mapping to human genes.

5. Cross species:

If a “hit” is observed in both Daphnia and human, It will trigger suitable regulatory process based on hypothesis that this toxicity pathway is conserved in both invertebrate and vertebrate animals and therefore evidence from daphnia might be used to trigger hazard concern beyond aquatic toxicity.

Project Plan 2018-19

APCRA: What is it, and where are we now?

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APCRA main objectives & partners

• Initiated by Bob Kavlock (EPA, now retired)

• To bring together international government regulators and researchers to discuss progress and barriers in applying new approach methodologies (NAMs) to prioritization, screening, and quantitative risk assessment of differing levels of complexity.

• Participating organisations: ECHA, EFSA, JRC, INERIS, RIVM, EPA, NTP, NICNAS, NITE (Japan), OECD, Health Canada, Environment and Climate Change Canada, A*STAR, SAHTECH, Seoul National University (Korea), University of Birmingham

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Key questions which are motivating APCRA projects:

• What are the current barriers to acceptance for successful use of NAMs in regulatory decision-making?

• What are near-term efforts that can improve use of NAM data?

• What is needed to lead to acceptance of NAMs by regulators and the public?

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Two APCRA projects in the spotlights

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APCRA Retrospective Study

APCRA Prospective Study

Examining the Utility of In Vitro Bioactivity as a Conservative

Point of Departure(APCRA Retrospective Study)

Katie Paul Friedman and Russell Thomas

Based on collaboration with A*STAR, ECHA, EFSA, EPA-OLEM, EPA-ORD, Health Canada, and the JRC

The views expressed in this presentation are those of the authors and do not necessarily reflect the views or policies of theU.S. EPA

Why is this case study important?

• Clear need to demonstrate in practical terms, for as many chemicals as possible, how preliminary screening level risk assessment using a new approach methodologies (NAM) based approach would perform when compared to traditional approaches to deriving points-of-departure (PODs)

• Illustrate the current state-of-the-science

• Evaluate the specific strengths and weaknesses of rapid, screening level risk assessment using NAMs

• Approach: Take a retrospective look at the traditional and NAM data for as many chemicals as possible.

The big question:

Can in vitro bioactivity be used to derive a conservative point-of-departure (POD) for prioritization and screening level risk assessment?

See the forest for the trees

PODtrad

EPA - ToxValDB

Health Canada

EFSA

ECHA

PODNAM

ToxCast AC50s (µM)

Apply high-throughput

toxicokinetics(httk) to get mg/kg/day

Exposure

EPA - ExpoCast

Health Canada

Bioactivity-exposure ratio

PODtrad : PODNAM ratio

Is POD ratio > 0 for most chemicals?Can we learn from POD ratio < 0?

Is BER useful for prioritization?Are there addressable weaknesses?

• NOEL, LOEL, NOAEL, or LOAEL

• Oral exposures

• Mg/kg/day

• If the sum of hitcalls across the ToxCast DB > 5, then the 5th percentile on the distribution of AC50 values was used.

• If the sum of hitcalls across the ToxCast DB ≤ 5, the lowest AC50 was used.• Flag-filtering by removing AC50 values from fits with 3+ caution flags and

hitpct

34/448 chemicals = 8% where PODNAM > PODtraditional

414/448 chemicals = 92% of the time this naïve approach appears conservative

PODNAM < PODtraditional

(most of the time)

Figure 2, Paul Friedman et al. in prep.

Next steps:

• To finalise the retrospective work

• To review the cases where PODNAM > PODTraditional(8% where NAM estimates are not conservative)

• To review the cases where PODNAM

Two APCRA projects in the spotlights

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APCRA Retrospective Study

APCRA Prospective Study

Quantitative and qualitative comparison of NAMs and traditional animal toxicity testing for data poor chemicals

• Case Leader: ECHA (Tomasz Sobanski, Mike Rasenberg)

• Project partners: A*STAR, ECHA, EPA, HC, JRC, NTP, RIVM, UoB

• Prospective case study to evaluate the qualitative and quantitative concordance of NAMs and traditional animal toxicity testing

• Will build off of the retrospective case study described before but data will be generated for data poor chemicals

• Use for hazard characterization and quantitative analysis if possible (in vivo part)

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APCRA prospective study

Main Case study questions:

• How far can we go with the NAM technologies currently available? Could we seriously consider application for hazard assessment at systemic toxicity level (high tier endpoints)?

• Can the outcome from the refined in vitro assay battery be used to derive a (conservative) point of departure and qualitative hazard triggers comparable with the outcome from Repeat Dose Toxicity (RDT) 90 day used in hazard assessment?

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APCRA prospective study

Project Goals

• To assess chemicals with limited/unclear toxicological data, which at the same time have significant potential exposure, using both NAM type of data and classical toxicological studies;

• To utilise and inform the further development needs for NAM:

• for screening, prioritisation and first tier assessments

• for conclusive hazard characterisation/assessment and risk management

• To assess chemicals in an international context

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APCRA prospective study

Why 90 day RDT as benchmark

•RDT study is designed to test wide range of effects;

•RDT provides an overview of systemic toxicity profile;

•RDT might trigger additional investigations for reprotox, immunotox, neurotox, carcinogenicity;

•90 day is considered as a conclusive test.

To allow meaningful comparison between NAM tests and RDT study endpoints extrapolation of in vitro concentration to in vivo relevant doses is critical.

In addition in vivo study will be complemented by mechanistic biomarkers (transcriptomics, metabolomics from multiple time points) and toxicokinetics.

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APCRA prospective study

•WP 1: Substance selection Q3 2017- Q3 2018 (finalised)

•WP 2: Phase I (in vitro) testing & in silico modelling Q4 2018- Q2 2020 (work is ongoing);

•WP 3: Phase II (in vivo) testingQ3 2019- Q4 2020 (preparatory work is ongoing);

•WP4: Analysis of the results & communication Q1 2019 – Q1 2021.

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APCRA prospective study: update on progress

Desired outcome: realistic scenario

• Provide a conservative estimate of in vivo LOAEL:

LOAELNAM

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• Provide quantitative estimate of NOAEL and LOAEL;

• For chemicals without a clear predominant mode-of action/MIE, bioactivity will be used as a conservative estimate of NOAEL/LOAEL

• For chemicals with a predominant MIE linked to a relevant AOP, the dose adjusted potency will be used as an estimate of the NOAEL/LOAEL

• Provide (semi) qualitative indications for

• Toxicity to reproduction & DevTox -> Activity in EPA ToxCast endocrine-related assays;

• Immunotoxicity -> Activity in EPA ToxCast immuotoxicity assays (Bioseek)

• Neurotoxicity -> Activity in EPA ToxCast neurotoxicity assays (microelectrode array),

• Carcinogenicity -> Activity in EPA ToxCast assays mapped to the IARC key characteristics of carcinogens.

Desired outcome: ultimate goal

What does this case study aim to achieve?

• Confirmation that NAM test battery can be successfully applied for screening with minimal risk of false negatives

• Verification whether/when NAM test battery can be directly used for quantitative hazard assessment

• Development of optimized assessment protocols aiming at implementation of the ‘NAM type’ of data in the multi-tiered hazard assessment

• Confidence building in application of NAMs for hazard characterisation

• Chemicals assessed at international level

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• This project builds on the retrospective case study, trying to address emerging questions:

• Why for small fraction of cases NAM estimates are not conservative enough;

• Why some NAM estimates are over conservative;

• How NAM will perform for substances with lower bioactivity;

• What are the limitations (applicability domain) of NAM approach;

• This project will indicate how far can we go with the NAM technologies currently available and

• Will help to find out how current NAMs can perform in various regulatory applications in comparison with classical methods.

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APCRA Prospective Study: Summary

Takes home messages

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• Direct transition from classical toxicity tests into vertebrate animal free methods for higher tier hazard assessment is not yet realistic.

• However, significant potential to refine existing vertebrate tests using mechanistic molecular biomarkers

• hybrid approach allowing smoother transition towards adoption of NAMs;

• gain mechanistic knowledge and confidence in molecular biomarkers;

• calibrate response levels from biomarkers to maintain comparable protection levels;

• potentially significant reduction in test durations;

• potential reduction in number of hazard assessment studies per substance as one study using molecular biomarkers might cover wider spectrum of toxicological space and/or increase sensitivity and specificity.

• Also, significant potential for purposeful increase in invertebrate testingusing mechanistic molecular biomarkers

• potential cross-species extrapolation for toxicants acting via conserved toxicological pathways (eco to human, and vice versa, e.g. expanding Daphnia omics study);

• wider re-use of existing data by providing mechanistic knowledge to substantiate chemical categories and read across hypothesis;

• would drive more dramatic improvements in 3Rs.

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Thank you!

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tomasz.sobanski@echa.europa.eu