Lessons Learned, Challenges, and Opportunities: The US EDSP | April 23

- 24, 2013| RTP, North Carolina 

### Predictive Toxicology Summit | May 22

– 23 2012 | Boston MA FIFRA SAP to review EDSP Tier 1 Assays and Battery Performance| May 21

- 13, 2013| Arlington, VA 

# The Future of Endocrine Screening: An Animal Welfare Perspective

Catherine Willett, PhD Director, Regulatory Testing Risk Assessment and Alternatives kwillett@humanesociety.orgFIFRA SAP to review EDSP Tier 1 Assays and Battery Performance| May 21

- 13, 2013| Arlington, VA 

# Outline

Animal use in the current EDSP design Suggestion for an alternate approach:

– Begin with modified tiered system

– + weight

- of

- evidence

– Move toward pathway

- based approachFIFRA SAP to review EDSP Tier 1 Assays and Battery Performance| May 21

- 13, 2013| Arlington, VA 

# Tier 1 Animal UseFIFRA SAP to review EDSP Tier 1 Assays and Battery Performance| May 21

- 13, 2013| Arlington, VA 

# Putative Tier 2 tests

Assay # animals/ test
Mammalian 2

- Generation 1600

- 2400 Amphibian 2

- Generation 400 P, plus progeny Avian 2

- Generation 1024 Fish Lifecycle ~1000* Total > 4,000

*number varies by speciesFIFRA SAP to review EDSP Tier 1 Assays and Battery Performance| May 21

- 13, 2013| Arlington, VA 

# Animals Used in Phase I
Data submitted and generated for 52 chemicals in Phase I: OSRI was submitted for all or some assays for 47 chemicals*

* Bishop, Sullivan and Willett, submittedFIFRA SAP to review EDSP Tier 1 Assays and Battery Performance| May 21

- 13, 2013| Arlington, VA 

# Need for a better approach

Current program design is resource, labor and animal

intensive Provides much data, only some of which may have have regulatory use Provides insufficient opportunity for chemically

- relevant tailoring Does not link mechanism to adverse outcome Is not predictiveFIFRA SAP to review EDSP Tier 1 Assays and Battery Performance| May 21

- 13, 2013| Arlington, VA 

# First step: a more refined tiered system

Initial tier takes into account all existing information, including physicochemcial Second tier assesses potential mechanisms of action Following tiers address potential effects in more complex systems, on multiple modes of action Highest tiers address adverse outcome and dose

- response WoE with clearly articulated criteria applied at each tier

• Defines positive and negative cut

- offs at the outset for each test and each Tier or Level

• Results are used to design the strategy for further testing Strategy is designed to address information needsFIFRA SAP to review EDSP Tier 1 Assays and Battery Performance| May 21

- 13, 2013| Arlington, VA 

# OECD Conceptual Framework

Level 1 

### Sorting and prioritization based on existing information

• physical and chemical properties

• exposure

• available toxicological data Level 2 

### In vitro assays providing mechanistic data

• QSARs

• binding and transcription assays

• aromatase, steroidogenesis, thyroid function

• high

- throughput screens Level 3 

### In vivo assays addressing single mechanisms and effects

• uterotrophic

• Hershberger

• amphibian metamorphosis

• fish VTG (e.g. TG 230) Level 4 

### In vivo assays addressing multiple mechanisms and effects

• enhanced 407

• male and female pubertal assays

• adult intact male

• fish short

- term reproductive assay (e.g. TG 299)
Level 5 

### In vivo assays providing data on effects from endocrine + other mechanisms

• 1 and 2 generation reproduction in rodents, birds, fishFIFRA SAP to review EDSP Tier 1 Assays and Battery Performance| May 21

- 13, 2013| Arlington, VA Level 3: Mechanistic **in vivo**  screens/ metabolism considerations • Negative

• Positive: MoA + existing adverse effects data

• Positive and

• More information required for HA /RA

Level 4: Targeted **in vivo** testing to assess specific adverse effects

Level 5: Targeted **in vivo** testing to

address specific adverse effects more completely More extensive characterization required for HA /RA Clear positive

or negative Continue? Yes Level 1: Physical/chemical and known/estimated information

Level 2: Mechanistic **in vitro** and **in silico** screens Lower Priority for

Further Testing

• Activity in more than one related assay

• Activity at low concentrations Continue? Yes

Continue? Yes Higher Priority for Further Testing

• No activity or

activity only at

high concs

. • Activity in few, unrelated

assays

Clear positive

or negative 

# Refined tiered system with integrated assessment

Willett et al. 2011. Toxicol . Sci.123(1):15

– 25.FIFRA SAP to review EDSP Tier 1 Assays and Battery Performance| May 21

- 13, 2013| Arlington, VA 

# The future of EDSP: a pathway

- based approach

A conceptual construct that portrays existing knowledge of linkages between a direct molecular initiating event and an adverse outcome at a level of biological organization relevant to risk assessment From: Ankley et al. Environ.Toxicol.Chem . 2010. 29 (3): 730

– 741.FIFRA SAP to review EDSP Tier 1 Assays and Battery Performance| May 21

- 13, 2013| Arlington, VA 

# ER

- mediated reproductive impairment

Developed from:

• an understanding of the MIE plus

• a need for a strategy for measuring endocrine

- mediated effects.

• Includes use of biomarker (vitellogenin) that is (in the case of females) or is not (in the case of males) directly linked to the adverse outcomeFIFRA SAP to review EDSP Tier 1 Assays and Battery Performance| May 21

- 13, 2013| Arlington, VA 

# ER

- mediated reproductive impairment integrated testing strategy

Adverse Outcome Pathway Chemicals Receptor Binding 

### ER Binding Liver Cells Altered Protein Expression 

### Vitellogenin Liver 

### Altered proteins

Gonad 

### Ova

- testis; Sex

- reversed; Fecundity Sex reversal ; Altered behavior;

Repro. **In vivo**

MOLECULAR

Target CELLULAR Response TISSUE/ORGAN INDIVIDUAL


### Skewed Sex Ratios; Population POPULATION



## Toxicity Pathway

P. Schmieder , McKim conference 2008. 

### QSAR/ **In vitro In**  **vitro**FIFRA SAP to review EDSP Tier 1 Assays and Battery Performance| May 21

- 13, 2013| Arlington, VA 

# ER and AR pathway efforts

US EPA:

• AOP development

• HTS and other in vitro assays 

# The Hamner Institutes: “Tier 1 and Done” CAAT: Pathways of Toxicity OECD Validation Management Group

– Non

- animal

• Binding and TA assays 

# OECD Advisory Group on Molecular Screening and Toxicogenomics

• AOP developmentFIFRA SAP to review EDSP Tier 1 Assays and Battery Performance| May 21

- 13, 2013| Arlington, VA 

# Thyroid pathway efforts

Development of AOPs and relevant assays

• OECD Thyroid Scoping Effort Expert Group

• OECD Validation Management Group

– Non

animal

• OECD Advisory Group on Molecular Screening and Toxicogenomics

• US EPAFIFRA SAP to review EDSP Tier 1 Assays and Battery Performance| May 21

- 13, 2013| Arlington, VA 

### • Build Biological and adverse

- outcome “pathways”

• OECD integration of AOPs into the Test Guidelines program

• Guidance



### • Improve predictive tools

• NIH National Center for Advancing Translational Sciences

• EPA’s Computational Toxicology Research

• OECD QSAR tool box

• Hamner Institute



### • Develop assessment systems for complex endpoints

• Reconstructed tissues and organ systems

• Human skin, eye, lung

• Liver

- on
- a
- chip

• Stem

- cell derived



### • Integrate absorption, metabolism and distribution information

• QSAR QIVIVE

-
• Liver cells, tissues, extracts, reconstructed tissues



### • Integrated databases and “knowledge bases”

• ACToR and MetaPath : EPA

– all available chemical toxicity data on over 500,000 environmental chemicals searchable by chemical name and structure

• Kegg pathway database : collection of manually drawn pathway maps representing current knowledge on the molecular interaction and reaction networks

• AOP Wiki: EPA

- European Commission joint project to house developing AOPs 

# Other necessary elementsFIFRA SAP to review EDSP Tier 1 Assays and Battery Performance| May 21

- 13, 2013| Arlington, VA 

# Conclusions

T he current EDSP design could be improved in terms of efficiency and utility of data generated Could begin with:

• a more refined organization of tiered tests

• c ombined with an iterative weight

- of

- evidence analysis As AOPs and tests are developed, move toward pathway

based approachFIFRA SAP to review EDSP Tier 1 Assays and Battery Performance| May 21

- 13, 2013| Arlington, VA 

# Thank You

Catherine Willett, PhD
Director, Regulatory Testing Risk Assessment and Alternatives Coordinator, Human Toxicology Project Consortium kwillett@humanesociety.org www.humantoxicologyproject.orgFIFRA SAP to review EDSP Tier 1 Assays and Battery Performance| May 21

- 13, 2013| Arlington, VA 

# Main concerns from Animal Protection Perspective High impact on animal use due to range finding

• Better prediction paradigms
> One assay to another
> From existing information (e.g. Part 158 data) 

# N eed to move away from MTD

• Confounding of specific results 

# U se of human recombinant receptors in binding studies U se of the BG1luc as alternate to STTA

• Both agonist and antagonist

• Concordance with uterotrophic 

# E limination of the Hershberger and uterotrophic assays

• Not adding value 

# Need to tie results back to mechanistic interpretation

• MoA or AOP

• To explain patterns of complementarity and redundancy

• As well as “non

- concordance”