2. NW Symposium in Melanoma May 22, 2010 Adoptive T Cell Therapy Cassian Yee MD Member Program in Immunology Fred Hutchinson Cancer Research Center cyee@fhcrc.org

3. T Cell Melanoma Cell

5. T Cell Melanoma Cell

6. T Cell Tumor Cell TCR Target Antigen

8. RBCs platelets WBCs B cells T cells WBCs

9. 1 in 100,000 T Cell Tumor Cell TCR Target Antigen CD8+

10. DC CD8 CD4 Normal Tumor Tumor Immune Surveillance

11. Immune evasion T cell frequency : Tumor burden Possible Reasons For Failure of Tumor Immune Surveillance Highly functional T cells More of them

12. Vaccine Therapy Adoptive Therapy

13. DC CD8 CD4 Tumor Adoptive T Cell Therapy

14. WBCs LEUKAPHERESIS

18. Study in Patients with Advanced Melanoma Treated with T Cell Therapy Recruited patients with  metastatic disease  progressive  failing 2nd and 3rd attempts to treat cancer

19. Patient 1017-6 Patient 1017-8 Before After T cell infusion

20. Study in Patients with Advanced Melanoma Treated with T Cell Therapy 30% Failed 30% Partial Response 30% Stabilized Response 10% near Complete Response 3 to > 30 months

21. 20 Lymphoid Homeostasis

22. Increase 'space' for transferred T cells Eliminate 'suppressor cells' Supply Growth Factors Increase 'space' for transferred T cells Eliminate 'suppressor cells' Supply Growth Factors Lymphodepletion building a better environment

23. PROTOCOL # 2140 Adoptive T Cell Therapy following Cyclophosphamide Lymphodepletion Objectives :Eligibility Criteria : - Evaluate Safety - Stage IV (Metastatic) - Evaluate T Cell Persistence - HLA-A2 - Evaluate anti-tumor efficacy T Cell Infusion: - Antigen-specific CD8+ T cell clones - Targeting MART-1, gp100 - Dose: 10 10 cells / m 2 CY 60 mg/kg x 2 Low-Dose IL-2 (250,000 U s.c q12 h)

25. T cell persistence in vivo 5.3% 1.1% D18 2140-1 2140-2

28. Clinical Response PatientTargetToxicityPersistenceDisease SitesResponse 2140-1TyrosinaseF,N,R>290 daysCervical,supraclavicular LN, Chest Wall, Breast Pulmonary nodules MR 2140-2TyrosinaseF 16 daysMediastinal, Pulmonary nodulesPD 2140-3gp100F,N,R>85 daysMesenteric LN, scapular subcutaneous dz CR (> 12 mths) 2140-4MART-1F, N, Rn.d.Pulmonary, inguinal, subcutaneous SD 2140-5MART-1F, N,Rn.d.Right and left kidneys, adrenal, liver PR 2140-6MART-1F, N, Rn.d.Mediastinal, supra clavicular, mammary chain, periportal, portacaval nodes. PR

29. Future Directions

30. Future of Adoptive T Cell Therapy LATE-STAGE DISEASE EARLY-STAGE DISEASE COMBINATION THERAPY WHICH CANCER TYPES?

32. 31

33. Adoptive Therapy following Lymphodepletion FLU 25 mg/m 2 x 5 HD IL-2 720K u/kg TID CY 60 mg/kg x 2 TBI High-Dose IL-2 (600,000 u./kg q8) Low-Dose IL-2 (250,000 U s.c q12 h)

34. Adoptive Therapy following Cytoxan Lymphodepletion Protocol 2140 HD IL-2 720K u/kg TID CY 60 mg/kg x 2 High-Dose IL-2 (600,000 u./kg q8) Low-Dose IL-2 (250,000 U s.c q12 h)

35. Isolate/EnrichClone/Select Expand Translational Strategies to Augment the CD8 Effector Response CD8-FITC Tet-PE Genetically Modify Pre-infusion Immunomodulation Post-infusion Immunomodulation Intrinsic Extrinsic  Phenotype - CD8/CD4 - Memory phenotype  Cytokine modulation  Lymphodepletion - Chemotherapy/TBI  Cytokine help - Low-dose IL-2 - High-dose IL-2 - Other  -chain receptor cytokines  Vaccine + adoptive therapy  TCR  Chimeric receptor  Costimulatory/Inhbitory modification  Suicide gene Ê CD4 Fludarabine Hematol Oncol Clin North Am. 2006 Jun;20(3):711-3

36. T Cell Expansion & Infusion Adoptive Therapy Using Antigen-Specific T cells Transferred Receptor Tumor Cell Chimeric TCR + zeta Chimeric Ig + zeta Receptor Transfer