1. Nonclinical Studies Subcommittee Advisory Committee for Pharmaceutical Science Introduction & FDA Objectives Gaithersburg Hilton December 14, 1999 T:\NCSS\121499.PPT

2. Nonclinical Studies Subcommittee Functions:  To provide advice on improved scientific approaches to nonclinical drug development  A means to foster scientific collaboration among FDA, industry, academia, and the public

3. Scientific Advances & Opportunities  Genomics/proteomics  Information technology  High throughput technologies  Mechanistic knowledge  Cancer, inflammation, cell signalling, etc.  Artificial intelligence  Noninvasive imaging

4. An example of accelerating scientific information and technology:  Safety Assessment

5. Toxicology in the last millennium ~~ B.C. knowledge & use of poisons 1500’s Exptl. physiology, toxic vs therapeutic dose-response (Paracelsus) 1600 Compound microscope 1600-1700 Anatomic microscopy (Hooke, Malphigi, van Leeuwenhoek) 1700’s Comparative anatomy (Hunter, Cuvier); Occupational toxicology (Ramazzini); Environmental cancer (Potts) 1800’s Exptl. physiology & toxicolgy (CO, strychnine, curare) Chemical teratogenesis (St. Hilare) Cellular pathology (Virchow, 1860’s) 1900’s Modern regulatory toxicolgy Cell biochemistry and molecular biology

6. Toxicology in the last half century ~~ 1906/1938 Food and Drug Acts; first flight 1940s Chemical carcinogenesis (Millers) & mutagenesis (Auerbach); DNA is the genetic material 1950 DNA structure; Clinical chemistry (serum biomarkers); Biochemistry (e.g., Krebs cycle); current approach to systemic toxicity evaluation 1960Toxicology as a discipline (SOT) Thalidomide, teratogensis testing 1970 EPA, OSHA formed; man on moon 1970’s Genetic toxicology testing; EMS (19070; TSCA (1978) 1980’s Genetic engineering; oncogenes; noninvasive imaging 1990’s Molecular damage response and defense Genome sequenced; era of genetic links to disease; high-throughput genetic tools

7. Current “Biomarkers” of Systemic Toxicity  Markers of:  Cellular integrity (AST, ALT, etc.)  Homeostasis (BUN, electrolytes,cell type, etc.)  Morpohologic evidence of damage  Host defense responses  Behavior/appearance/body weight of organism

8. Current approach to safety evaluation  Clinical Chemistry/Hematology  Markers of cellular integrity (AST, ALT, etc.)  Markers of homeostasis (BUN, electolytes, etc.)  Alterations in circulating cell populations  Histopathology  Visible morphologic or staining change  Host defense cell infiltration  Behavior/appearance/body weight of organism  Special tests: cancer, mutation, reproduction, neurotoxicology, immunotoxicology, etc.

9. Some Opportunities for New/Improved Toxicological Practices  Damage-specific responses  Objective biochemical assays for host-defense cell signaling and infiltration  Better biomarkers of integrity/homeostasis  e.g., troponins  Biochemical markers of cell death  In vivo genetic markers of mutational damage, oncogene activation, and suppressor inactivation  Noninvasive technologies  “Humanized” and/or transgenic animals  Cell culture & microengineering

10. Molecular Evolution of Defense Systems  Systems have evolved to protect and repair each major function  Defense systems are often inducible  Molecules often evolve from function to repair  Key defense systems are conserved  Understanding these systems will provide the next generation of surrogate biomarkers for monitoring damage to cells and tissues

12. Some Damage- or Agent-Inducible Genes Damage Type Functional Class /Inducer Class Example Genes Protein structure Protein denaturation HSP70, clpB DNA integrity DNA damage dinD, recA, GADD153 Oxidative Protectants Redox balance katG, soi28, GST Growth Control Cell proliferators FOS, JUN DNA damage Metal Inducible Toxic metals merR Xenobiotic Inducible Xenobiotics CYP1A1, CYP2E1

13. DNA damage-response genes Protein damage-response genes Intracellular free radical-response genes DNA Microarray or “Gene Chip” with Multiple Probes Containing scores of potential biomarkers, e.g.

14. Genomics to Proteomics Microsphere color identifies the protein captured Intensity of tagged Ab reflects amount of captured protein Sample mixed with Ab-coated spheres and labeled Ab Biomarker proteins are in patient serum sample

15. What do we need to know?  Relationship of endpoint to health  Relationship to outcomes in established assays  Relationship between laboratory models and man  Reproducibility, accuracy, sensitivity, robustness

16. Application of Safety Biomarkers to Human Studies Secreted proteins that are upregulated following toxic insult Tissue/organ-specific proteins that signal loss of cellular integrity Inaccessible upregulated membrane proteins that bind specific non- invasively monitorable probes

17. Biochemical Markers of Pathology Damage/Insult Defense Response Cell death Caspase-mediated cell death Cell death/tissue damage Chemokine/cytokine-mediated inflammatory response Foreign/damaged protein Immune responses/NK receptor

19. Better biomarkers of cell integrity and homeostasis

20. Immunofluorescent Labeling of Cardiac Troponin I (cTnI) in Doxorubicin- induced Cardiomyopathy Intercalated disks (arrow), I-bands (circle), and staining intensity (box) are reduced, while vacuoles (triangle) are increased in DXR- treated animals. ControlDoxorubicin

21. Effect of Doxorubicin (DXR) on Serum Cardiac Troponin T (TnT) Cumulative Dose DXR (mg/kg) Serum Cardiac TnT (ng/kg)

22. Influence of the Severity of Doxorubicin Induced Cardiomyopathy of the Concentration of Serum Cardiac Troponin T (TnT) Cardiomyopathy Score Serum Cardiac TnT Concentration (ng/ml)

23. How to focus resources among opportunities? ACPS Nonclinical Studies Subcommittee: Identify and recommend focus areas Identify experts in focus areas; form expert working groups (EWGs) with nominations from: Federal Register announcements (Public) FDA and “Stakeholders” (Collaborators) Professional Societies Steering committee to collaborative projects Support workshops and facilitate reports

24. Potential collaborators  FDA  CDER  CBER  Industry  PhRMA  BIO  Academia  Public Institutions

25. Objectives 1. To recommend approaches and mechanisms to improve:  Nonclinical information for effective drug development  Predictivity of nonclinical tests for human outcomes  Linkage between nonclinical and clinical studies and 2. To facilitate collaborative approaches to advancing the scientific basis of drug development and regulation

26. History and Next Steps  NCSS Subcommittee meeting 8/31/99 –Define objectives and operating principles –Define focus areas –Discuss initial focus areas and mechanisms for implementation –Define operating structure  ACPS meeting 9/24/99 –ACPS endorsed concept  Select initial focus areas  Form EWGs  EWGs identify collaborators and identify resources