1. Small Molecule PD and Imaging – Models and Assay Development DCTD Phase 0 Primer, September 5, 2007 Ralph E Parchment, PhD SAIC-Frederick, Inc., NCI-Frederick, Frederick, Maryland 21702 Funded by Contract N01-CO-12400

2. Session Speakers and Topics Dr. Robert Kinders, Principal Scientist, PD Assay Development & Implementation Section, SAIC-Frederick ¬ ¬Assay Development/Validation ¬ ¬Phase 0 Clinical Trial Modeling Using Preclinical Models Dr. Melinda Hollingshead, Chief, BTB/DTP/DCTD/NCI ¬ ¬Implementing Clinical Biopsy Procedures in Small Animal Models Dr. Susan Galbraith, VP of Clinical Discovery Research at BMS ¬ ¬Phase 0 Trials-Why Arent They More Widely Used by Industry? Dr. David Mankoff, University of Washington ¬ ¬PET Imaging Assessment of Therapeutics

3. Building Blocks of the DCTD Program in Clinical Pharmacodynamics Phase I dose escalation with PD to establish surrogate tissue for tumor Establish reliable supply of key materials with QC and batch equivalency Phase 0 clinical trial PD assay gives 1 o endpoint surrogate tissues also evaluated Preclinical modeling of tumor-surrogate relationship eIND filing Transfer validation to clinical lab Create real-time reporting Preclinical modeling of Phase 0 design to prove suitability of drug-target pair Validate specimen handling SOPs Validate an SOP-driven PD assay Select clinical procedures specimen collection, processing, storage, extraction and dilution Test candidate PD assay for dose effect and specificity (inactive control) Identify 1 o /2 o assay endpoints Identify technology platform(s)/assay Identify surrogate tissue candidate(s) Schedule supply of specimens Scientific knowledge base mechanism of action, target(s), signal transduction pathways

4. Clinical scientists Veterinarians Lab scientists Phase II trials using surrogate tissue for PD assessments Phase I dose escalation with PD to establish surrogate tissue for tumor Establish reliable supply of key materials with QC and batch equivalency Phase 0 clinical trial PD assay gives 1 o endpoint surrogate tissues also evaluated Preclinical modeling of tumor-surrogate relationship eIND filingTransfer validation to clinical lab Create real-time reporting Preclinical modeling of Phase 0 design to prove suitability of drug-target pair Validate specimen handling SOPs Validate an SOP-driven PD assay Select clinical procedures specimen collection, processing, storage, extraction and dilution Test candidate PD assay for dose effect and specificity (inactive control) Identify 1 o /2 o assay endpoints Identify technology platform(s)/assay Identify surrogate tissue candidate(s) Schedule supply of specimens Scientific knowledge base mechanism of action, target(s), signal transduction pathways

5. Select Clinical Procedures: Specimen Collection, Processing, Storage, Extraction and Dilution Current limitations in PD… ¬ ¬Replicating human medicine procedures in the animal models ¬ ¬Implementing PD measurements within clinical constraints of tissue yield, procedure time, logistics of specimen handling, anesthesia effects, etc., because method can affect PD measurement ¬ ¬Processing strategy is hurrying to stabilize the signal after tissue is removed from the body ¬ ¬Limited to a very static measurement of PD via biopsy at 1-2 time points This area will be addressed in detail by Dr. Hollingshead

6. Future improvements in PD…. ¬ ¬Change philosophy of specimen handling to create procedures and devices that stabilize molecular profile prior to tissue removal (FY2008 SBIR Contract Solicitation, Topic #250) ¬ ¬Create methods for repeat sampling to achieve more dynamic pharmacodynamics, such a circulating tumor cells, permanent placement of biopsy access port, etc ¬ ¬If we all use inhibitors of enzymes that degrade the product of the molecular target during tissue extraction, why dont we also use inhibitors of the enzymes that produce them? ¬ ¬Develop methods to directly assess target enzyme activity in biopsies Select Clinical Procedures: Specimen Collection, Processing, Storage, Extraction and Dilution

7. 1:20 rule for dynamic range ¬ ¬assumes 33-50% yield ¬ ¬assumes that a 80-90% inhibition level will need to be measured ¬ ¬achievable with 3 of 3 platforms (immunosandwich assays, immunofluorescent assays on tissue sections, and qRT-PCR) Assess influence of: ¬ ¬biomatrix on assay signal (dilution non-linearity) ¬ ¬tumor heterogeneity on PD variability and required drug effect ¬ ¬effect of prior Bx on subsequent Bx, including drug effect ¬ ¬time required after dosing to find PD effect in tumor Bx This area will be addressed in detail by Dr. Kinders Preclinical Modeling of Phase 0 Design to Prove Suitability of Drug-Target Pairs Validate Specimen-Handling SOPs

8. Dose or Exposure Target Function Dose or Exposure Target Function Dose or Exposure Target Function Dose or Exposure Target Function Non-toxic dose range Candidate Phase 0 PairsCandidate Phase 1 Pairs Non-toxic dose range Non-toxic dose range Non-toxic dose range Preclinical Modeling of Phase 0 Design to Prove Suitability of Drug-Target Pairs

9. The Next Speaker is: Dr. Melinda Hollingshead