The document discusses several case studies highlighting the importance of bidirectional information flow between clinical, preclinical, and discovery research: 1) A study of rituximab immunotherapy in lymphoma patients found that allowing longer B cell recovery time before vaccination improved responses, informed by preclinical studies. 2) Development of an anti-IgVH monoclonal antibody for lymphoma was guided by preclinical toxicology in monkeys to explore depletion of target B cells. 3) Gene expression analysis of pediatric IBD patient biopsies generated hypotheses tested with discovery research and preclinical models. 4) Increased IP-10 levels correlated with clinical response in UC patients treated with anti-CD3 visilizumab, informing the mechanism of

1. Cancer and Inflammatory Diseases Immunotherapy Building Bridges Between Discovery, Preclinical, and Clinical Research

3. B Cell Lymphoma Immunotherapy

7. Clinical Study Design Immunization efficiency following rituximab Rituximab 4 Doses (N=90)* Restage at 8 Weeks: CR, CRu, PR Patients who failed chemotherapy Study 2002-09 Sampling times for T and B cells and immune response *Third cohort (N=16) of immunized patients were not eligible for either 3M or 6M Rest groups. Screen failures (N=33) not eligible for immunization. Leonard J.P. et al., ASCO 2006 Follow- up 6-month Rest Period (N=23) 8 Immunizations Q 2 weeks Follow- up 3-month Rest Period (N=18) 8 Immunizations Q 2 weeks

8. Patients treated with rituximab have dramatically reduced B cell numbers B Cells (CD19 + ) T Cells (CD3 + ) 3M rest period 3M rest period 6M rest period 6M rest period Leonard J.P. et al., ASCO 2006 Post-R Post-R

9. Patients treated with rituximab have significantly lower humoral response against KLH Leonard J.P. et al., ASCO 2006 Analysis of variance (treatment effect) 9902B 2002-09 3M Rest p < 0.0001 9902B 2002-09 6M Rest p < 0.0001 2002-09 3M Rest 2002-09 6M Rest p < 0.05

13. Concept of a selective mAb immunotherapy of B cell lymphomas Sornasse T. et al., ASH 2007

14. mAbs specific for Ig variable region framework directly kill lymphoma cell line in vitro Anti-Ig VL mAb: Cell line A Anti-Ig VH mAb: Cell line B Lymphoma cell lines were incubated for 48 hours in the presence of mAbs. Dead cells were identified by flow cytometry as 7-AAD-positive cells.

17. Anti human Ig VH 3.23 mAb does not significantly affect the frequencies of B and T lymphocytes in vivo Control Group Dose Group: 10 mg/kg Dose Group: 40 mg/kg Dose Group: 100 mg/kg Legend Infusions Flow cytometry Schedule of events Sornasse T. et al., ASH 2007 Study Days 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15

18. Anti human Ig VH 3.23 mAb depletes target B cells in vivo (Study 2)          Sornasse T. et al., ASH 2007

20. Inflammatory Diseases

23. Basic Differences Between CD and UC Ulcerative Colitis Crohn’s Disease Lesion characteristics : - Strictly mucosal. - Continuous and diffuse. - Progress from distal portion of GI tract. Lesion characteristics : - Transmural ( may involve layers as deep as adventitia ). - Scattered and focal. - May appear anywhere along lower GI tract.

25. Legend Apoptosis IFN-  pathway Chemokines Cytokines Tissue remodeling

26. Summary of Observations Tissue repair (Nidogen, IGFBP-5) Proteases (MMP-12, Serpin) Tissue Remodeling Cellular Recruitment MCP-1, MIP3-  Inflammation IFN-  , IL-1  Caspases Apoptosis Inhibitors (Ets-2, BIRC3) IBD Pathology Collaborate with clinical research to obtain critical resources for discovery research

29. In vitro mode of action: experiment design OKT3: anti-CD3 mAb that binds to FcR  strong activator of T cell proliferation and cytokine release Microarray

30. Results: Cytokines Time RNA Levels (Arbitrary Units)

31. Results: CXC Chemokines Time RNA Levels (Arbitrary Units)

34. Rapid increase in serum IP-10 levels in patients treated with visilizumab Woo J. et al., DDW 2006

40. Daclizumab does not significantly affect  CD3/  CD28-induced proliferation of human PBMC The PBMC of adult volunteers were stimulated with  CD3/  CD28 beads for 72 hours in the presence of graded doses of daclizumab. Results are presented as the Average of the relative proliferation to no-daclizumab control ± SEM Sornasse T. et al., AAAAI 2005 Potential effect on cytokine production is not affected by T cell number

41. Daclizumab strongly inhibits the secretion of pro-asthmatic Th2 cytokines The PBMC of healthy adult volunteers were stimulated with  CD3/  CD28 beads for 72 hours in the presence of graded doses of daclizumab. Results are presented as the Average of the relative cytokine levels to no-daclizumab control ± SEM Sornasse T. et al., AAAAI 2005

42. Daclizumab inhibits the secretion of pro-asthmatic Th0 cytokines The PBMC of healthy adult volunteers were stimulated with  CD3/  CD28 beads for 72 hours in the presence of graded doses of daclizumab. Results are presented as the Average of the relative cytokine levels to no-daclizumab control ± SEM Sornasse T. et al., AAAAI 2005

43. Daclizumab partially inhibits the secretion of pro-inflammatory Th1 cytokines The PBMC of healthy adult volunteers were stimulated with  CD3/  CD28 beads for 72 hours in the presence of graded doses of daclizumab. Results are presented as the Average of the relative cytokine levels to no-daclizumab control ± SEM Sornasse T. et al., AAAAI 2005

46. Flow of information in research The old view

47. Flow of information in research The new view