NexGen Risk Assessment

Transcription

1 NexGen Risk Assessment Daniel Krewski, PhD, MHA Professor and Director McLaughlin Centre for Population Heath Risk Assessment & Risk Sciences International January 12, 2011

2 McLaughlin Centre for Population Health Risk Assessment

3 Cornerstone #1: Toxicity Testing in the 21 st Century McLaughlin Centre for Population Health Risk Assessment

4 Toxicity Testing in the 21 st Century: A Vision and A Strategy Committee on Toxicity Testing and Assessment of Environmental Agents Board on Environmental Studies and Toxicology Institute for Laboratory Animal Research Division on Earth and Life Studies National Research Council

5 Components of the Vision

6 Chemical Characterization

7 Toxicity Testing

8 Endorsement by the Scientific Community Collins, F.S., Gray, G.M. & Bucher, J.R. (2008), Science (Policy Forum). Vol pp

9 US Environmental Protection Agency Strategic Plan and Strategic Goals

10 Reaction from Experts in Risk Assessment Suresh Moolgavkar, our Area Editor for Health Risk Assessment, asked six experts with different perspectives to comment on the paper. Each praises the vision and offers suggestions for making it more useful. Michael Greenberg & Karen Lowrie, Editors

11 Reaction from the Legal Community

12 Reaction from the Animal Law Community International Symposia on Challenges and Opportunities in Implementation June 29-30, 2009 September 12, 2009 November 5, 2009 June 21-23, 2010 There is widespread support for the NAS vision. There are also real but surmountable challenges in moving the vision into routine regulatory practice. Progress is being made in producing the necessary science and knowledge base we need to redouble our efforts to see that these insights carry over into the worlds of law and policy. Paul Locke, Johns Hopkins University Center for Alternatives to Animal Testing

13 Cornerstone #2: Population Health Risk Assessment McLaughlin Centre for Population Health Risk Assessment

14 McLaughlin Centre for Population Health Risk Assessment

15 Canadian Institute for Advanced Research (1989) Individual Response biology behaviour Social Environment Physical Environment Genetic Endowment Health & Function Disease Health Care Interventions Well-being Prosperity

16 What is Population Health Risk Assessment? Risk assessment is a process to characterize risk using scientific methods. Population health risk assessment is the comprehensive assessment of health risks in the general population based on genetic, environmental, social & behavioural determinants of health. This forms the basis for evidence-based population health risk policy analysis, and, ultimately, costeffective population health risk management decisions. McLaughlin Centre for Population Health Risk Assessment

17 Population Health Regulatory Economic Advisory Community Technological Multiple Interventions Health Risk Policy Analysis Evidence Based Policy Health Risk Science Determinants and Interactions Biology-environment interactions Environment-social interactions Biology and Genetics Environment and Occupation Social and Behavioural Biology-social interactions

18 Cornerstone #3: Science and Decisions: Advancing Risk Assessment McLaughlin Centre for Population Health Risk Assessment

19 KEY MESSAGES Enhanced framework Formative focus Four steps still core Matching analysis to decisions Clearer estimates of population risk Advancing cumulative assessments People and capacity building

20 Risk-Based Decision-Making Framework Phase I Formulating and Scoping Problem For environmental condition: What s the problem? What are the options for altering? What assessments are needed to evaluate options? Not OK Phase II Planning and Risk Assessing Stage 1: Planning for: Options Assessment Uncertainty and Variability Analysis Stage 2: Assessing Stage 3: Confirming Utility of Assessment OK Phase III Risk Management Relative benefits of proposed options? How are other factors (e.g., costs) affected by options? Which option is chosen? What s the uncertainty and justification? How to communicate it? Should decision effectiveness be evaluated? If so, how? Stakeholder involvement at each phase

21 Background Paper #1 McLaughlin Centre for Population Health Risk Assessment

22 NRC Vision for the Future of Toxicity Testing McLaughlin Centre for Population Health Risk Assessment

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25 Interim and Final Reports

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27 Original Editorial 8 +1 Invited Commentaries McLaughlin Centre for Population Health Risk Assessment

28 Recurring Themes in the Commentaries Definition of adversity Predicting in vivo results from in vitro toxicity pathway assay results Setting standards from results of in vitro assays How can the change from current practices to a new paradigm occur?

29 McLaughlin Centre for Population Health Risk Assessment.

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31 JTEH Special Issue on Future Directions in Toxicity Testing Part A: NRC Report on Toxicity Testing in the 21 st Century (reprint with permission) Part B: U.S. EPA Strategic Plan for Toxicity Testing (reprint) Part C: Individual contributions on future directions in toxicity testing McLaughlin Centre for Population Health Risk Assessment

32 Building the Scientific Toolbox (Andersen et al., 2010)

33 Case Study Prototypes McLaughlin Centre for Population Health Risk Assessment

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35 Tool Lung Injury and Ozone Functional genomics Microarray analysis of both in vivo and in vitro airway epithelial cells exposed to clean air and ozone Proteomic changes will be monitored and compared with mrna changes Comparison of altered toxicity pathways between in vivo and in vitro cell systems Bioinformatics Mining of known toxicity pathways, key biological events, biomarkers and relevant endpoints related to ozone toxicity Systems biology Systems biology approach to integrate information for modeling NFκB inflammatory pathway due to ozone exposure Goals are to understand network mechanisms behind disease process Computational systems biology Structure-activity relationships Molecular epidemiology Development of BBDR models Framework to illustrate key events and MOA due to ozone exposure Ozone structure linked to perturbation pathways Examination of a number of inflammatory markers to assess inflammatory response in lungs such as neutrophils, cytokines, prostaglandins, LDH, etc. Genetic epidemiology Comparison of toxicity pathways from in vitro and in vivo systems come from the same subjects therefore the variability in response across all the subjects will be taken into account McLaughlin Centre for Population Health Risk Assessment

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37 Tool High throughput screens Developmental Impairment and Thyroid Hormone Disruptors ToxCast TM : screened for endpoints relevant to thyroid hormone disruption 90 HTS assays were identified as thyroid hormone pathway relevant : 1) endpoints had a direct impact on thyroid hormone pathways, 2) 85 had impacts related to indirect thyroid hormone homeostasis and neurodevelopment Functional genomics HPT axis is well mapped and points of possible disruption due to chemical exposure are shown in Figure 1 Figure 2 shows thyroid hormone control pathways and points of disruption 90 assays were identified that measured endpoints and had dose- response curves related to HPT pathway The identification of a signature profile could involve the combined analysis of a diverse set of assays endpoints Bioinformatics: ToxCast database, developed by NCCT and NIEHS Mining of this database identified relevant endpoints found in thyroid hormone disruption Quality analysis and summarization of data entered into a database is needed for effective predictive modeling Simplification of data into binary formats are highly dependant on appropriate criteria and may be inconsistent across different assays Systems biology 90 assays identified with endpoints related to thyroid hormone disruptions; pathways and cellular responses include hepatic stimulation and metabolism, thyroid hormone clearance and thyroid hormone pathway Computational systems biology Structure-activity relationships ToxCast database: analysis of dose-response curves by visual inspection and modeling 1) species concordance, 2) effect identification, and 3) relative potency. Chemical classification is highly dependant on quality data analysis and summarization for predictive modeling Table 1 identifies IRIS chemicals that have direct effects on thyroid hormone pathway Biomarkers Table 2 identifies several assay targets to measure thyroid hormone disruption and homeostasis Table 3 identifies possible endpoints that still need to be evaluated and measured in assays CAR, PXR nuclear receptor activation: activation linked directly to reduced thyroid hormone Other possible nuclear receptors related to reduced thyroid hormone: PPAR, LXR McLaughlin Centre for Population Health Risk Assessment

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39 Tool High throughput screens Cancer and Polycyclic Aromatic Hydrocarbons ToxCast TM : HTS assays for genotoxicity and detecting genes in the p53 signaling pathway Three HTS assays were evaluated: 1) the GreenScreen HC assay using a (TK6 human cell line with a GADD45a-GFP (green fluorescent protein) gene reporter) 2) the p53 endpoint in the CellCiphr Cytotox Profiling Panel uses (HepG2 cells and an anti-p53 antibody) 3) the Invitrogen CellSensor p53re-bla HCT-116 assay (HCT-116 colon cancer cells with a beta-lactamase reporter gene controlled by p53 response elements) Functional genomics Table 3.2 evaluates data for key mechanistic events from omics literature: 1) High carcinogenic potency of dibenzo[a,l]pyrene potentially related to genotoxic key events (i.e., DNA adduct formation, ras-oncogene mutation, and tumor initiation) 2) Carcinogenicity of benzo[a]pyrene could be due to direct interaction of metabolites with DNA (i.e., adduct formation, oncogene and tumor suppressor gene mutation, tumor initiation) and possible epigenetic key events (AhR-mediated effects, inhibition of gap junctional intracellular communication [GJIC]) 3) dibenzo[a,l]pyrene and dibenzo[a,h]anthracene induced changes in genes involved in apoptosis Pathway analysis demonstrated that the potent PAHs modulated several pathways whereas less potent PAHs (fluoranthene and 1-methylphenanthrene) did not significantly affect any pathways. Proteomic changes were limited to evaluation of p53 accumulation Venn diagrams showing overlap in gene expression changes Bioinformatics Micro array data from many studies were analyzed and summarized in several tables, for example Table 3-4 list various genes affected by PAHs and significantly correlated with cancer potency, DNA adduct formation, or Ah induction in HepG2 cells Systems biology PAHs have the ability to bind to AhR and this is suspected to suppress estrogen response element-controlled gene expression through crosstalk between the AhR and ERα. McLaughlin Centre for Population Health Risk Assessment

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41 Tool Cancer and Polycyclic Aromatic Hydrocarbons (Part 2) Structure-activity relationships QSAR models capable of predicting mechanisms and/or cancer potency for PAHs based on structural features: 1) PAHs having four or more benzene rings exhibit greater carcinogenic potency 2) PAHs with two or three benzene rings have less carcinogenic potency 3) carcinogenicity is related to the specific arrangement of the benzene rings 4) PAH metabolites resistant to detoxification due to stereochemical effects are more likely to be mutagenic and cause cancer 5) Dihydrodiol epoxides formed at certain positions on the PAH molecule are more accessible to glutathione transferase detoxification and are less potent mutagens and carcinogens Table 3-1 tabulates chemical with three in vivo pathways linked to differing potencies of carcinogenesis Table 3-4 shows the genes affected by PAHs and significantly correlated with cancer potency, DNA adduct formation, or Ah induction in HepG2 cells Biomarkers Possible biomarkers could be found from microarray data and Venn diagrams Biomarkers of AhR, estrogen response, and p53 pathway Exposure assessment Extracts from soils contaminated with PAHs were exposed to HepG2 cells and various proteins levels related to different pathways were measured; p53, Cdk2, cyclin D1, p21, Chk2 Thr68, Mdm2 Ser166, perk Tyr204,53BP1, γ-h2ax, and Mdm2 2A10 McLaughlin Centre for Population Health Risk Assessment

42 Tool Cancer and Benzene Functional genomics Genome-wide expression profiles of human promyelocytic leukemia HL-60 cells exposed to VOCs was analyzed using a microarray Yeast screening approach identified highly conserved genes involved in susceptibility to HQ, CAT, and BT. NF1 (neurofibromin) was identified as a candidate susceptibility gene to the benzene metabolite, hydroquinone (HQ) Microarray platforms to identify global gene expression changes associated with occupational benzene exposure in the peripheral blood mononuclear cells (PBMC) of a population of shoefactory workers. Four genes (CXCL16, ZNF331, JUN and PF4were among the top 100 genes identified by both platforms Carcinogenesis epigenomics data showed effects of benzene on the DNA methylation of specific genes and on mirna expression Systems biology The roles of the human homologs of selected genes in benzene toxicity were examined through mechanistic studies in human cell lines. WRN protein using sirna in HeLa cells and examined sensitivity to toxicity following exposure to the benzene metabolite, HQ Biomarkers Biomarkers to assess benzene exposure: Urinary trans,trans-muconic acid (t,t-ma), S-phenylmercapturic acid, and benzene (U-benzene) Molecular epidemiology 125 workers exposed to a range of benzene exposures : microarray mrna transcriptome analysis of peripheral blood mononuclear cells identified genes and pathways (apoptosis, immune response, and inflammatory response) 239 workers recruited among traffic policemen, taxi drivers and gasoline pump attendants of the city of Parma (Italy) to study benzene exposures at low levels: Study investigates nucleic acid oxidation and biomarkers associated with exposure. Gas station attendants, urban policemen, bus drivers, and two groups of controls (415 subjects): Biomarkers such as urinary trans, trans-muconic acid (t,t-ma), S-phenylmercapturic acid, and benzene (U-benzene) were measured to assess benzene exposure Genetic epidemiology Genotyping studies of 250 benzene-exposed workers and 140 unexposed controls in China: analyzed 1395 SNPs in 411 potential carcinogenesis-related genes and associations between DNA repair, genomic maintenance and development of benzene hematotoxicity were found McLaughlin Centre for Population Health Risk Assessment

43 Tool Lung Injury and Ozone Developmental Impairment and Thyroid Hormone Disruptors Cancer and Polycyclic Aromatic Hydrocarbons Cancer and Benzene High throughput screens Stem cell biology Functional genomics Bioinformatics Systems biology Computational systems biology Physiologically-based pharmacokinetic models Structure-activity relationships Biomarkers Molecular and genetic epidemiology. Exposure assessment McLaughlin Centre for Population Health Risk Assessment

44 Background Paper #2 McLaughlin Centre for Population Health Risk Assessment

45 Cornerstone #1: Toxicity Testing in the 21 st Century McLaughlin Centre for Population Health Risk Assessment

46 Toxicity Testing and Risk Assessment (from Krewski et al., 2010, Annual Review of Public Health, in press)

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52 Cornerstone #2: Population Health Risk Assessment McLaughlin Centre for Population Health Risk Assessment

53 Population Health Regulatory Economic Advisory Community Technological Multiple Interventions Health Risk Policy Analysis Evidence Based Policy Health Risk Science Determinants and Interactions Biology-environment interactions Environment-social interactions Biology and Genetics Environment and Occupation Social and Behavioural Biology-social interactions

54 Health Determinants Biology & Genetics Genetic Polymorphisms Gender Immune status Metabolism

55 Health Determinants Environment & Occupational Physical environment: air, food, water, soil, built environment Working conditions

56 Health Determinants Social & Behavioural Personal health practices Coping skills Employment/ working conditions Education and literacy Social environment Healthy child development Culture Customs, traditions, and beliefs Social stability

57 Multiple Interventions: REACT Intervention Examples Regulatory Economic Governmental laws and regulations, policies, guidelines, audits, inspections and fines Levies or other cost structures, insurance, investments in future technology, government grants to invest in new technology Advisory Health promotion campaigns, safety awareness workshops Community Action Volunteer public monitoring groups, increases in capacity building Technological Updating equipment to newer technology or repairing old technology, financial investment in research and development of new emerging technologies

58 Cornerstone #3: Science and Decisions: Advancing Risk Assesment McLaughlin Centre for Population Health Risk Assessment

59 Table 4. Potential Modifications of Risk Assessment Approaches in a NexGen Context Risk Issue Current Approach NexGen Approach Defining Adversity Adversity is presently defined in terms of observation of apical endpoints in mammalian systems. Adversity will be defined in terms of critical perturbations of toxicity pathways, ultimately in the absence of information on apical outcomes. Defining adversity will require knowledge of dose response for various pathway assays and in vitro models that assess conditions leading to excessive pathway perturbations in relevant assays.

60 Table 4. Potential Modifications of Risk Assessment Approaches in a NexGen Context Risk Issue Current Approach NexGen Approach Point of departure (PoD) NOAEL or BMD, based on apical endpoints. Dose response curves are presently based on measurement of apical endpoints in animal models, with inter-species extrapolation of the health outcome from animal to human. NexGen risk assessments will still use empirical statistical models to establish dose response relationships for pathway perturbations, and select established points of departure such as the BMD for in vitro responses deemed adverse. Alternative approaches for PoDs may include new methods, such as SNCD (signal to noise crossover dose) or statistical methods to assess likelihood of threshold behaviours. Inter-species extrapolations will no longer be required when human cells are used in HTS in vitro assays.

61 Table 4. Potential Modifications of Risk Assessment Approaches in a NexGen Context Risk Issue Current Approach NexGen Approach Default assumptions Current default assumptions used in risk assessment (such as a 10-fold variation in sensitivity within the human population) are usually based on limited empirical evidence. Understanding toxicity pathways in more depth will permit a move away from default assumptions, towards a more mechanistic approach guided by scientific evidence and knowledge of the behaviour of the toxicity pathway in shifting from basal levels of activity to enhanced function with excessive perturbation. (It will likely be possible to characterize phenotypic variation with some precision using suites of human cell lines representing inherent differences in sensitivity and differences among life stages and through knowledge of pathway components and polymorphisms in these components that affect function).

62 Table 4. Potential Modifications of Risk Assessment Approaches in a NexGen Context Risk Issue Current Approach NexGen Approach Analysis of various life stages In vivo generational studies using mammalian systems can be conducted at different life stages, but are expensive, time consuming and labour intensive. In the absence of evidence to the contrary, it is assumed that individuals at all life stages are equally vulnerable. NexGen risk assessments can make use of systems biology and virtual tissue modeling that simulates human organs in 3 dimensions (e.g., 3D liver model), to determine the effects that could be expected from the exposures of humans of various ages to toxic substances. High throughput in vitro assays using human cells and human cell lines representing different life stages could facilitate a more complete assessment of vulnerability at different life stages.

63 Table 4. Potential Modifications of Risk Assessment Approaches in a NexGen Context Risk Issue Current Approach NexGen Approach Analysis of multiple exposure doses Present risk assessments are based on dose-response curves generated by high dose acute exposures of animal models, and then low dose extrapolation is performed. Laboratory exposures of animals to environmentally-relevant and sublethal doses are not generally performed for the purpose of conducting a risk assessment but may be stated in the risk assessment report. It will be feasible in NexGen risk assessments to conduct exposures to a wide range of doses for risk assessment purposes because of the rapid and cost efficient use of HTS. Using medium throughput, multidimensional tissue surrogates should allow some conditions where in vitro responses will be assessed over days to weeks of exposure.

64 Selected Risk Issues to be Addressed Chemical mixtures Analysis of various life stages Joint effects of multiple stressors Assessment of delayed effects Reversible or transient effects Analysis of multiple exposure doses Assessment of exposures of different durations (e.g., acute, chronic, and intermittent exposures)

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