Key Events Dose-Response Framework TCEQ

1. Key Events Dose-Response FrameworkTCEQ/ARA Workshop IBeyond Science and Decisions:From Issue Identification to Dose-Response AssessmentMarch 16-18, 2010Austin, Texas Steve Olin (ILSI Research Foundation) Elizabeth Julien (Consultant) 1

2. Origins QUESTION: Can our increasing understanding of modes of action provide insights for characterizing dose-response relationships (including thresholds) and, if so, how? Not only for chemicals but also for other bioactive agents: nutrients, pathogens, allergens. ? Key Events Dose-Response Framework 2

3. Key Events Dose-Response Framework ILSI Research Foundation established a tripartite, multidisciplinary expert group to develop an integrated framework to incorporate advances in scientific knowledge, based on mode of action concept, focus on understanding the fundamental biology and dose-response (including possible thresholds) at each key event, inform hazard characterization and risk assessment, foster cross-disciplinary discussion. 3

4. ILSI RF Threshold Working Group Chemical Group: Alan Boobis (Imperial College London), George Daston (Procter & Gamble), and Julian Preston (EPA). Nutrient Group: A. Catharine Ross (Pennsylvania State), Robert Russell (Tufts), Joseph Rodricks (ENVIRON), Sanford Miller (U Maryland), Ian Munro (CANTOX), and Elizabeth Yetley (retired NIH). Pathogen Group: Bob Buchanan (U Maryland), Arie Havelaar (RIVM), Mary Alice Smith (U Georgia), and Richard Whiting (Exponent). Allergen Group: Steven Gendel (FDA CFSAN), Geert Houben (TNO), and Steve Taylor (U Nebraska). 4

5. Products and Next Steps 5 papers � Crit Rev Food Sci Nutr, 49 (8), Sept 2009 (Overview, Chemicals, Nutrients, Pathogens, Allergens) � open access: http://www.informaworld.com/smpp/title~db=all~content=g914018566 Next Steps Encourage the development of additional case studies illustrating and evaluating the utility of the Framework Organize small meetings and workshops to work through specific examples Explore application and integration of the Framework into MOA analysis for risk assessment 5

6. Mode of action and key events 6

7. 7 Examining an individual key event Factors that combine to determine outcome of individual events: Dose (level, frequency and duration) Physiological mechanisms (e.g., homeostasis, repair, immune response, compensatory pathways) Host factors (life-stage, disease state, genetic makeup, nutritional status, co-exposure)

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9. Address each key eventsystematically Is a minimum dose level required in order for this key event to occur? What data would be needed to demonstrate this? What response mechanisms (e.g. homeostasis, repair) are involved? At what dose would these be overwhelmed? What modifying factors (e.g. lifestage, disease state, nutritional status) can potentially reduce the effectiveness of response mechanisms? What factors can increase the effectiveness of response mechanisms? Do such modifying factors change the dose level at which response mechanisms become overwhelmed? What data would be needed to demonstrate this? 9

10. Looking at the whole pathway of events � Which events may be �control points� where mechanisms exist to maintain the normal physiological environment (e.g., homeostasis)? Are any control points especially vulnerable (readily overwhelmed by dose? readily modified by host factors? ) Is any particular key event a possible �determining event� � i.e., its outcome disproportionately affects the probability of seeing the outcome of interest? Does any particular key event appear to drive the slope or shape of the overall dose-response relationship? 10

11. First Case Examples Chemicals Non-DNA-reactive carcinogen (chloroform) DNA-reactive carcinogens Endocrine disruptors (binding to estrogen receptor) Nutrients Vitamin A (retinol) toxicity Pathogens General discussion of toxigenic, toxicoinfectious, and invasive bacteria Listeria monocytogenes Food Allergens General discussion of key events for elicitation 11

12. Postulated MOA for CHCl3 In rodent bioassay studies, Chlorofom has been shown to result in liver and kidney tumors. The postulated MOA to explain these tumor responses involves bioactivation through CYP2E1 oxidative metabolism as the rate limiting step in chloroform�s mode of action. Metabolism by this pathway produces cytotoxic metabolite within the target organ, in particular phosgene that injures and kills cells, cytotoxicity is followed by regenerative cell proliferation, and it cytotoxicity/regenerative proliferation is sustained, eventually tumor development. So,these are the key sequence of events that will be considered with respect to chloroform;s included tumorigenesis in the rodent kidney and liver.In rodent bioassay studies, Chlorofom has been shown to result in liver and kidney tumors. The postulated MOA to explain these tumor responses involves bioactivation through CYP2E1 oxidative metabolism as the rate limiting step in chloroform�s mode of action. Metabolism by this pathway produces cytotoxic metabolite within the target organ, in particular phosgene that injures and kills cells, cytotoxicity is followed by regenerative cell proliferation, and it cytotoxicity/regenerative proliferation is sustained, eventually tumor development. So,these are the key sequence of events that will be considered with respect to chloroform;s included tumorigenesis in the rodent kidney and liver.

14. Critical determinant of threshold There is evidence that several of the key events in chloroform carcinogenesis exhibit a threshold. It is not possible to determine which of these is the critical determinant (if any), from available data. For this, studies in which each event was studied in isolation would be necessary. Potential contribution of toxicogenomics Information from all biologically relevant models should be used in such analyses. This is of particular value for more distal key events, which will be less compound specific. 14

15. Individual vs. population thresholds Thresholds will vary among individuals. Once the determining key events are understood, research to study contributions to population variability (including identification of susceptible populations) can be targeted on those events. The goal is to understand how various factors (age, gender, disease state, nutritional status, etc.) may quantitatively affect the doses at which those determining events occur. 15

16. 16 Applying the KEDRF to nutrients Typically, for a given nutrient there will be long-term intake, with a dose that varies day to day. For some nutrients, homeostatic controls exist to regulate blood and tissue levels despite daily intake variation. Control via: One or more kinetic events One or more dynamic events Various intake patterns may lead to adverse effects: acute excess intake, chronic excess intake, chronic deficiency.

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18. 18 Overview of Retinol Pathway

19. 19 Analysis of Events I

20. 20 Analysis of Events II

21. 21 Analysis of Events III Conclusion: Binding activity appears to be only a partially regulated event; not a control point for regulating RA levels.

22. Central events leading to adverse effects from excess dietary vitamin ACentral events leading to adverse effects from excess dietary vitamin A

23. 23 Vitamin A Toxicity:Analysis and Research Needs Saturation of LRAT, leading to excessive RA levels, is likely a �determining event�. Question: how high must circulating RA (or its metabolites) be in order to cause effect? How long must it remain high? Key research needs Quantitative data on dose-response for LRAT saturation Characterize sources of variability at control points Explore the potential role of downstream events

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25. Another ILSI Project �Risk Assessment in the 21st Century (Risk21)� Joint ILSI HESI/ILSI Research Foundation project Just getting underway; probably 3 years Building on recent NAS reports, a Scoping Meeting, August 2009, proposed 4 broad focus areas: Exposure Science Dose Response MOA and high-to-low dose extrapolation In vitro ? in vivo extrapolation in context of HTPS and ToxTesting21 Tiered (Integrated) Testing Cumulative Risk Contact: Michelle Embry (HESI) � membry@ilsi.org 25