Jennifer Seed, PhD Office of Pollution Prevention & Toxics US EPA

1. Perspective on the current state-of-knowledge of mode of action as it relates to the dose response assessment of cancer and noncancer toxicity This presentation represents the view of presenter & does not necessarily represent the decisions or stated policies of the EPA. Jennifer Seed, PhD Office of Pollution Prevention & Toxics US EPA

2. Evolution of Risk Assessment 21st Century 1980s 2000s 1990s RA/RM Paradigm Guidelines/Methods Dosimetry/PbPK Mode of Action Susceptible Populations Mixtures Toxicity Pathways Integrated Approaches CompTox 1983 2007 1994 2009

3. Characterize Uncertainty Mode of Action information is core to the risk assessment paradigm and its evolution into the 21st century • Understand • Human Relevance • Susceptibility • Dose Extrapolation • Mode of Action • Toxicity Pathways • Identify Key Events • Promote • Harmonized approach for all endpoints • Inform • Multiple Chemical Risks

4. How do you determine the Weight of Evidence for establishing MoA? • Needs to be based around specific hypothesis judged against data • International Framework • Introduced by USEPA & IPCS (1999, 2001) • ILSI builds on EPA/IPCS work (2003, 2005) •  IPCS continues work (2006, 2008) Based on Bradford Hill Criteria for Causality Distinguishes MoA vs Mechanism Human Relevance (Concordance) Analysis

5. Mode of Action/Human Relevance (MoA/HR) Framework • Why is it important? • Increases predictive capacity and utility of risk assessment • Provides rigor and structure to analysis and consistency of documentation • Provides transparency • Guides and focuses research/testing 5

6. Figure 5.8 New unified process for selecting approach and methods for dose-response assessment for cancer and noncancer end points.

7. Mode of Action Human Relevance (MoA/HR) Framework (v.IPCS) Dose Response & Species Extrapolations Exposure Assessment Compare “Key Events” & relevant biology between animals & humans

8. Question 1: Sufficient Weight of Evidence to establish MoA in animals? • Postulated MoA (hypothesis) • Describe sequence of Key Events (measurable) • Consider other possible MoAs • Experimental support for key events • Concordance of dose-response relationships • Temporal association • Strength, consistency and specificity of association of toxicological effect with key events • Biological plausibility and coherence • Uncertainties, inconsistencies, missing data

9. Cacodylic Acid: Sequence of “Key Events” in MOA Measurable Key Events in Target Tissue DMAIII Metabolite Urothelial Toxicity Urinary bladder from a female F344 rat treated with 100 ppm DMAV Sustained Regenerative Proliferation BrdU Labeling Other MoAs: DNA damage via ROS? Hyperplasia Urinary Bladder Tumors

10. Q1: WoE Sufficient to Establish MoA? Temporal • Postulated mode of action • Sequence of key events • Experimental support • Relationship of key events & adverse effects • Dose-response • Temporal response • Biological plausibility & coherence • Strength, consistency & specificity • Identify uncertainties 1 wk 6 hrs 8 wk 104 wk Dose Response

11. Q.2 & 3 Evaluation of Human Relevance Q1. Weight of evidence sufficient to establish MoA in animals? Q2. Fundamental qualitative differences in key events? Q3. Fundamental quantitative differences in key events? Compare “Key Events” & relevant biology between animals & humans *Use all existing knowledge: chemical specific & generic (e.g., genetic/disease models, related cpds) 11

12. Q. 2 & 3 The Concordance Analysis 12

13. Carrying MoA information forwardinto the dose-response analysis MoA-Based vs Default • Dose extrapolation based on MoA understanding • Key events - shape of the dose-response curve & points of departure Q1. Weight of evidence sufficient to establish MoA in animals? Q2. Fundamental qualitative differences in key events? Q3. Fundamental quantitative differences in key events? 13

14. Point of Departure for Cacodylic Acid: Benchmark dose modeling using key events BMDL10 (mg/kg/day) Key Events BMD10 (mg/kg/day) Urothelial cytotoxicity (3 wk; 10 wk) Proliferation (10 wks) Hyperplasia (10 wks) Tumors (104 wks) 0.7; 0.02 0.9 2.0 7.7 0.2; 0.0008 0.4 1.5 6.0 PoD that provides a level of exposure that will not trigger regenerative proliferation .

15. Carrying MoA information forward -Implications for inter- and intra- species extrapolation Default = 10X Default = 10X Interspecies Dynamics (2.5) Human Variability in Disposition (3) Interspecies Kinetics (4) Human Variability in Sensitivity (3) Data Derived Extrapolation Factors

16. IPCS Harmonization Guidance Relevant to MoA-Based Risk Assessment • Conceptual Framework for Evaluating a Mode of Action for Chemical Carcinogenesis • Human Relevance Framework for Cancer • Non-cancer Human Relevance Framework • Principles of Characterizing and Applying PBPK Models in Risk Assessment • Chemical Specific Adjustment Factors • Combined Exposures to Multiple Chemicals information can be found at: http://www.who.int/ipcs/methods/harmonization/en/index.html

17. Hazard Characterization Risk Characterization 2007 NRC Toxicity Testing in the 21st Century Dose Response Assessment Chemical Characterization Mode of Action Population Based Studies Compounds Dose Response Analysis for Perturbations of Toxicity Pathways Affected Pathway Assess Biological Perturbation Calibrating in vitro and human Dosimetry Exposure Guideline Measures of dose in vitro Metabolite(s) Human Exposure Data Exposure Assessment Fig 3-7 Risk Assessment Components