Toxicology in the 21 st Century

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Leona Garrett
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1 Toxicology in the 1 st Century A Road Map for the National Toxicology Program Dr. Christopher J. Portier Associate Director, National Toxicology Program National Institute of Environmental Health Sciences Department of Health and Human Services

2 5 Years of the Basic Research Research In Toxicology Testing And Evaluation Public Health

3 5 Years of the Basic Research Research In Toxicology Testing And Evaluation Public Health Improved toxicological sciences Design Implementation Interpretation Utilization of new tools

4 5 Years of the Basic Research Research In Toxicology Testing And Evaluation Public Health Innovative reviews ROC and mechanism CERHR Alternatives Major issues of concern

5 5 Years of the Basic Research Research In Toxicology Testing And Evaluation Public Health Toxicology databases Cancer Immunotoxicology Genetic toxicology Tissue blocks/slides Others

6 The Challenge for the Our expanding concerns Chemicals Pharmaceuticals Physical and biological agents Mixtures

7 The Challenge for the Our expanding concerns Chemicals Pharmaceuticals Physical and biological agents Mixtures Our expanding knowledge Genomics/Genetics/Mo. Bio. Toxicity mechanisms

8 The Challenge for the Our expanding concerns Chemicals Pharmaceuticals Physical and biological agents Mixtures Our expanding knowledge Genomics/Genetics/Mo. Bio. Toxicity mechanisms Our improved science Robotics and computers Better cellular and molecular tools Newer laboratory models

9 The for the Next 5 Years Continue to lead Mechanism-based toxicology Faster screening Interpretation

10 The for the Next 5 Years Continue to lead Mechanism-based toxicology Faster screening Interpretation Continue to build Databases and analysis tools Scientific foundation for a transformation of toxicology

11 The for the Next 5 Years Continue to lead Mechanism-based toxicology Faster screening Interpretation Continue to build Databases and analysis tools Scientific foundation for a transformation of toxicology Continue to improve Standard toxicological assays Public health decisions

13 Use of Mechanistic Toxicology Studies in Public Health Decision Making

14 Use of Mechanistic Toxicology Studies in Public Health Decision Making

15 Approaching Toxicology Today First level screen High-Throughput Fairly inexpensive Lot s of possible mechanistic links Second Level Screen Integrated living organism Medium throughput Complex inter-related mechanistic information Definitive Evaluation Bioassay of the future

16 The Bioassay of the Future Species/strain/genetics Dose spacing/timing/age of exposure Use of sub-chronic/pre-chronic/other Toxicokinetics Digital pathology and non-invasive methods Addition of mechanistic endpoints Presentation and interpretation

17 Chronics, Cancer and Genomics Biomarkers Biomarkers Cancer No Cancer Biomarker Biomarker Cancer No Cancer

18 Second Level Screens Shorter-term whole animal assays C. elegans Zebra fish Transgenics and others Molecular screening assays Gene chips with large numbers of genes Proteomic screens Metabolomic screens Combined assays Reproductive/developmental/cancer/cardiotox/other

endpoints Subtle changes in individual neurons Alterations in egg sac

19 C. Elegans and developmental neurotoxicity Develop the screen 00 /- agents Reproduction, development, behaviour, movement Expand the tool to expand endpoints Subtle changes in individual neurons Alterations in egg sac function and number Evaluate mechanism Knock down every gene for a few chemicals

20 High-Throughput Screening Develop capacity Two to five thousand agents initially Obvious toxicity targets Develop data handling tools Build a validation data set Expand as warranted Broader number of assays Broader number of agents Wider number of targeted toxicities

assays 30 grants per year on new methods 10K agents screened 153 well

21 High-Throughput Screening Fully integrated with the NIH Molecular Libraries Initiative 10 screening centers 100 grants per year on new assays 30 grants per year on new methods 10K agents screened 153 well format 10 plus dilutions Multiple replicates Cross-laboratory validation

22 Data Analysis and Interpretation Evaluate current and future information technology needs Methods for new assays and HTS Interaction with public health decision makers How and when to use new results Steps as we proceed Staffing and expansion of skills staff Decision makers International community

23 Environmental Systems Biology Arterial Venous Blood Metabolite to Feces Liver Metabolite CYP1A- 80% 0% Metabolite to Urine Oral Dose GI Tract Lumen Fat Kidney Rapidly Perfused Muscle Topical Dose Local Skin Remote Skin

24 Environmental Systems Biology Arterial Venous Blood Biochemistry Topical Dose Liver CYP1A- Fat Metabolite to Feces Metabolite to Urine Rapidly Perfused Metabolite Local Skin 80% 0% Lumen GI Tract Oral Dose Metabolite Metabolism Cell Lysis Kidney Bound Capillary (CyP1A) Blood Muscle Liver Remote Skin Ah Growth Signal EGFR Ah. Growth Peptide - CYP1A1 ER ER.E CYP1A CYP1B1 Liver Interstitium E DNA Damage E OH

25 Environmental Systems Biology Topical Dose Arterial Venous Blood Two-Stage Cancer Model Liver CYP1A- Fat Metabolite to Feces Metabolite to Urine Rapidly Perfused Metabolite Local Skin 80% 0% Lumen Oral Dose Metabolite GI Normal Tract Cells Metabolism Cell Lysis Initiated Kidney Bound Cells Capillary (CyP1A) Blood Muscle Single Cell Liver Malignant Clones Remote Skin Ah Growth Signal EGFR Ah. Growth Peptide Biochemistry - CYP1A1 ER ER.E CYP1A CYP1B1 Liver Interstitium E DNA Damage E OH

26 Environmental Systems Biology Topical Dose Arterial Venous Blood Two-Stage Cancer Model Liver CYP1A- Fat Metabolite to Feces Metabolite to Urine Rapidly Perfused Metabolite Local Skin 80% 0% Lumen Oral Dose Metabolite GI Normal Tract Cells Metabolism Cell Lysis Initiated Kidney Bound Cells Capillary (CyP1A) Blood Muscle Single Cell Liver Malignant Clones Remote Skin 0.7 Ah Growth 0.4 Signal EGFR Ah. Growth Peptide Biochemistry - CYP1A1 ER ER.E CYP1A CYP1B1 Liver Interstitium E DNA Damage E OH

27 Environmental Systems Biology Topical Dose Arterial Venous Blood Two-Stage Cancer Model Liver CYP1A- Fat Metabolite to Feces Metabolite to Urine Rapidly Perfused Metabolite Local Skin 80% 0% Lumen Oral Dose Metabolite GI Normal Tract Cells Metabolism Cell Lysis Initiated Kidney Bound Cells Capillary (CyP1A) Blood Muscle Single Cell Liver Malignant Clones Remote Skin 0.7 Ah Growth 0.4 Signal EGFR Ah. Growth Peptide Biochemistry - CYP1A1 ER ER.E CYP1A CYP1B1 Liver Interstitium E DNA Damage E OH

28 Environmental Systems Biology Topical Dose Arterial Venous Blood Two-Stage Cancer Model Liver CYP1A- Fat Metabolite to Feces Metabolite to Urine Rapidly Perfused Metabolite Local Skin 80% 0% Lumen Oral Dose Metabolite GI Normal Tract Cells Metabolism Cell Lysis Initiated Kidney Bound Cells Capillary (CyP1A) Blood Muscle Single Cell Liver Malignant Clones Remote Skin Ah Growth Signal EGFR Ah. Growth Peptide Biochemistry - CYP1A1 ER ER.E CYP1A CYP1B1 Liver Interstitium E DNA Damage E OH

29 Environmental Systems Biology Topical Dose Arterial Venous Blood Two-Stage Cancer Model Liver CYP1A- Fat Metabolite to Feces Metabolite to Urine Rapidly Perfused Metabolite Local Skin 80% 0% Lumen Oral Dose Metabolite GI Normal Tract Cells Metabolism Cell Lysis Initiated Kidney Bound Cells Capillary (CyP1A) Blood Muscle Single Cell Liver Malignant Clones Remote Skin Ah Growth Signal EGFR Ah. Growth Peptide Biochemistry - CYP1A1 ER ER.E CYP1A CYP1B1 Liver Interstitium E DNA Damage E OH

30 Environmental Systems Biology Arterial Venous Blood Biochemistry Topical Dose Liver CYP1A- Fat Metabolite to Feces Metabolite to Urine Rapidly Perfused Metabolite Local Skin 80% 0% Lumen GI Tract Oral Dose Metabolite Metabolism Cell Lysis Kidney Bound Capillary (CyP1A) Blood Muscle Liver Remote Skin Ah Growth Signal EGFR Ah. Growth Peptide - CYP1A1 ER ER.E CYP1A CYP1B1 Liver Interstitium E DNA Damage E OH

31 Environmental Systems Biology Arterial Venous Blood Biochemistry serum CRABPI Polar Topical Dose Liver CYP1A- retinol Fat retin(ol/al)-crbpii CRBP CRABPII metabolites Metabolite to Feces Metabolite to Urine Capillary rreh Blood Rapidly Perfused apo-crbp RA-CRABP1 UDPGT CYP1A1 ZNF4 Metabolite Local Skin RA-RARB retinyl esters retinol-crbp RARB 80% 0% (ARAT) LRAT ADH CRBP-retinal CYP1A1 Retinoic acid (ATRA) THR VitDR ALDH6 9c-RA RXR Lumen RXR/PPAR ACOX1 GI Tract Oral Dose Metabolite Metabolism Cell Lysis Hsp70 Kidney Bound AhR Capillary (CyP1A) Blood Hsp90/p3/XAP - AhR/ARNT Muscle ALDH3 ALDH10 Laminins Liver - Remote IRF4 Skin -/ LIFR-IL6ST LIF src IFN - EGFR EGFR-P Rap1 Ras cadherin- LAMA Laminin5 catenin Ah Integrins Egr-1 DUSP1 - Cell adhesion Proliferation p38 MAPK STK4(MST1) Growth Thrombin PAR-1 (FR/TR) Apoptosis Signal VEGF ARNT/HIF1a IFIT1 IFT ISG15 MX1 EGF Mek-P Erk-P Ras-P Terminal Differentiation Mek Erk Cell Migration Immunomodulation -/ The Inflammation National Institutes of Health Angiogenesis Cell Adhesion Genes altered in HPL1A Cells EGFR Ah. Growth Peptide - CYP1A1 ER ER.E CYP1A CYP1B1 Liver Interstitium E DNA Damage E OH

32 AHR Activated Pathways serum CRABPI retinol rreh ZNF4 retinyl esters retinol-crbp RARB (ARAT) LRAT apo-crbp ADH retin(ol/al)-crbpii CRBP-retinal CRBP CRABPII Polar metabolites RA-CRABP1 UDPGT CYP1A1 RA-RARB CYP1A1 Retinoic acid (ATRA) THR VitDR ALDH6 9c-RA RXR RXR/PPAR ACOX1 Hsp70 AhR Hsp90/p3/XAP - AhR/ARNT ALDH3 ALDH10 Laminins - -/ LIF LIFR-IL6ST -/ src IFN - IRF4 Inflammation EGFR EGFR-P Ras Rap1 DUSP1 - Thrombin ARNT/HIF1a LAMA Laminin5 cadherin- IFIT1 IFT ISG15 MX1 EGF p38 MAPK STK4(MST1) catenin Ras-P Mek-P Erk-P Integrins Cell adhesion Proliferation PAR-1 (FR/TR) Terminal Differentiation Mek Erk VEGF Angiogenesis Cell Migration Cell Adhesion Immunomodulation Egr-1 Apoptosis Genes altered in HPL1A Cells

33 AHR Activated Pathways serum CRABPI retinol rreh ZNF4 retinyl esters retinol-crbp RARB (ARAT) LRAT apo-crbp ADH retin(ol/al)-crbpii CRBP-retinal CRBP CRABPII Polar metabolites RA-CRABP1 UDPGT CYP1A1 RA-RARB CYP1A1 Retinoic acid (ATRA) THR VitDR ALDH6 9c-RA RXR RXR/PPAR ACOX1 Hsp70 AhR Hsp90/p3/XAP - AhR/ARNT ALDH3 ALDH10 Laminins - -/ LIF LIFR-IL6ST -/ src IFN - IRF4 Inflammation EGFR EGFR-P Ras Rap1 DUSP1 - Thrombin ARNT/HIF1a LAMA Laminin5 cadherin- IFIT1 IFT ISG15 MX1 EGF p38 MAPK STK4(MST1) catenin Ras-P Mek-P Erk-P Integrins Cell adhesion Proliferation PAR-1 (FR/TR) Terminal Differentiation Mek Erk VEGF Angiogenesis Cell Migration Cell Adhesion Immunomodulation Egr-1 Apoptosis Genes altered in HPL1A Cells

34 AHR Activated Pathways CYP1A1 ALDH6 ALDH3 ALDH10 CRABPI RARB Gene Interaction Network (GIN) ZNF4

35 Hypothetical Network elf3 aldh3 aldh6 rarb aldh10 crabp coa znf4 ncoa c1a1 cdk

36 Hypothetical Network elf3 aldh3 β 0 rarb β 1 β 7 aldh6 β 6 β β 3 β 4 aldh10 β 13 β 1 β 14 coa β11 β 8 crabp β 9 β 5 znf4 β15 β 16 β 10 ncoa β 18 c1a1 cdk β 17

37 Hypothetical Network elf3 aldh3 β 0 rarb β 1 β 7 aldh6 β 6 β β 3 β 4 aldh10 β 13 β 1 β 14 coa β11 β 8 crabp β 9 β 5 znf4 β15 β 16 β 10 ncoa β 18 c1a1 cdk β 17

38 Hypothetical Network elf3 aldh3 β 0 rarb β 1 β 7 aldh6 β 6 β β 3 β 4 aldh10 β 13 β 1 β 14 coa β11 β 8 crabp β 9 β 5 znf4 β15 β 16 β 10 ncoa β 18 c1a1 cdk β 17

39 The Next 5 Years of the Expanding toxicology databases New assays High-Throughput Screening Genetics/genomics and toxicology Testing the BIG hypotheses Continuing innovative reviews Traditional toxicology screens Genetics/genomics/HTS/new assays and ROC/CERHR Validation on a broader scale Improved toxicological sciences Design Implementation Interpretation

40 1 st Century Toxicology Leading the Way Collaborative Participation at All Levels MULTI-TIERED Testing Program MECHANISTIC Linkage in All Tiers SCIENCE-BASED Priority Setting and Interpretation EFFICIENT Utilization of Resources Developing the Best Science to Achieve the Best Decisions

41 1 st Century Toxicology Leading the Way Collaborative Participation at All Levels MULTI-TIERED Testing Program Basic Research MECHANISTIC Linkage in All Tiers Research Testing SCIENCE-BASED In Priority Setting And and Toxicology Evaluation Interpretation EFFICIENT Utilization of Resources Public Health Developing the Best Science to Achieve the Best Decisions

Using EGFR signaling networks to guide therapeutic strategies

Using EGFR signaling networks to guide therapeutic strategies Erica A. Golemis Program in Developmental Therapeutics Fox Chase Cancer Center November 13, 2010 Problem: the limited efficacy of therapies