S. Khleif - Ovarian cancer - General lecture on vaccine

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  • Vaccines based on DNA constructs, viral vectors, cytokines…etc …already proven safe
  • Related % = all g3/4 toxicities possibly/probably/defiantly related to vaccine except for local reactions, constitutional symptoms, and adjuvant related events
  • Related % = all g3/4 toxicities possibly/probably/defiantly related to vaccine except for local reactions, constitutional symptoms, and adjuvant related events
  • Autologous – Stated by us (3), stated by them (4), statistically significant (1)Allogeneic – stated by us (2), stated by them (1)Synthetic – stated by us (13), stated by them (5), statistically significant (1), unknown (1)Non phase 1 – stated by us (2), stated by them (4), statistically significant (1)
  • S. Khleif - Ovarian cancer - General lecture on vaccine

    1. 1. Cancer Vaccine- What is the Current State of the Art? Samir N. Khleif Chief, Cancer Vaccine Section, NCIProfessor of Medicine, Uniformed Services University for Health Sciences
    2. 2. VaccinesProphylactic Therapeutic preventive Treatment
    3. 3. VaccinesProphylactic Therapeutic preventive Treatment
    4. 4. VaccinesProphylactic Therapeutic preventive Treatment
    5. 5. Cancer Vaccines• Prophylactic: prevention or elimination of the causative agents• Preventive: elimination of premalignant lesion or prevention of recurrence• Treatment: established tumor
    6. 6. VaccinesProphylactic TherapeuticHPV preventive Treatment Provenge
    7. 7. Prophylactic
    8. 8. Infectious agents and Cancer• HPV• HBV• HCV• EBV• HIV• HHV8• HTLV1• Helicobactore
    9. 9. Infectious agents and Cancer• HPV• HBV• HCV• EBV• HIV• HHV8• HTLV1• Helicobactore
    10. 10. Cervical Cancer• 2nd most common cancer worldwide• ~ 510,000 new cases / year• ~ 274,000 deaths / year• Most common cancer in the developing world
    11. 11. What is HPV? • Small DNA Tumor Virus • 55 nm in diameter • 8000-base double stranded circular DNA • Causes an array of benign and malignant pathology
    12. 12. L1/2 E5/2/4 E1 E6/7
    13. 13. L1/2 E5/2/4 E1 E6/7
    14. 14. L1/2 E5/2/4 E1 E6/7
    15. 15. Woodman et al. Nature Reviews Cancer 7, 11–22 (January 2007) | doi:10.1038/nrc2050
    16. 16. HPV InfectionTransient PersistentInfection Infection • 1 year Low Grade Dysplasia CIN 1 • 5 years High Grade Dysplasia CIN 2/3 • Decades Invasive Cancer
    17. 17. HPV InfectionTransient PersistentInfection Infection Low Grade Dysplasia CIN 1 High Grade Dysplasia CIN 2/3 Invasive Cancer
    18. 18. HPV InfectionTransient PersistentInfection Infection Low Grade Dysplasia CIN 1 Integration High Grade Dysplasia CIN 2/3 Invasive Cancer
    19. 19. HPV InfectionTransient PersistentInfection Infection Low Grade Dysplasia CIN 1 Integration High Grade Dysplasia CIN 2/3 Invasive Cancer
    20. 20. HPV InfectionTransient PersistentInfection Infection Low Grade Dysplasia CIN 1 Integration High Grade Dysplasia CIN 2/3 Invasive Cancer
    21. 21. Viral Proteins Expressed HPV Infection L1, L2Transient PersistentInfection Infection Low Grade Dysplasia CIN 1 Integration High Grade Dysplasia CIN 2/3 Invasive Cancer
    22. 22. L1/2 E5/2/4 E1 E6/7
    23. 23. Humoral Vaccines• Killed virus (influenza)• Live attenuated (MMR)• Sub-units
    24. 24. Humoral Vaccines• Killed virus (influenza)• Live attenuated (MMR)• Sub-units
    25. 25. Live Attenuated/killed Viruses Are Not Suitable For an HPV Prophylactic Vaccine• Papillomavirus cannot be efficiently grown incultured cells• The viral genomes contain oncogenes
    26. 26. Humoral Vaccines• Killed virus (influenza)• Live attenuated (MMR)• Sub-units
    27. 27. L1/2 E5/2/4 E1 E6/7
    28. 28. L1/2 E5/2/4 E1 E6/7
    29. 29. HPV L1 VLP Vaccine SynthesisL1 gene Empty viralon HPV capsids DNA Yeast cell DNA Transcription L1 gene inserted Capsid proteins into genome of mRNA yeast cell tRNA Translation rRNA Yeast Cell
    30. 30. HPV L1 VLP Vaccine SynthesisL1 gene Empty viralon HPV capsids DNA Elicits immune response in host Yeast cell DNA Transcription L1 gene inserted Capsid proteins into genome of mRNA yeast cell tRNA Translation rRNA Yeast Cell
    31. 31. Vaccine AvailablecompanyName Vaccine TypeMerck: Gardasil Quadrivalent (yeast) FDA Approved 2006GSK: Cervarix Bivalent (baculovirus) FDA Approved 2009
    32. 32. Clinical Trials: HPV Prophylactic Vaccine Study Vaccine No of Subjects End points Efficacy Vaccine Control % (confidence limits)Koutsky Gardasil 6087 6080 CIN 2/3 AIS 98 (86-100)Garland Gardasil 2241 2258 CIN 2/3 AIS 100 (94-100) 2261 2279 GW VIN 100 (94-100) VAIN Joura Gardasil 7811 7785 VIN 2/3 100 (72-100) VAIN 2/3Harper Cervarix 481 470 CIN 100 (42-100) CIN 2/3Paavonen Cervarix 7788 7838 CIN 2/3 90 (53-99) MITT Analysis
    33. 33. Duration of Follow up in Phase II Studies Harper DM, Expert Rev Vaccines. 2009
    34. 34. Reduction in LEEP for CIN 2/3 Harper DM, Expert Rev Vaccines. 2009
    35. 35. US Recommendations• FDA approved the vaccine for girls and women ages 9 to 26 for the prevention of Cervical, Vulvar and vaginal cancers and precancerous lesions• FDA approved the vaccine for men and women ages 9 to 26 for the prevention of anal cancers and precancerous lesions• FDA approved the vaccine for boys and men ages 9 to 26 to prevent genital warts• Federal Advisory Committee on Immunization Practices recommends that girls routinely receive the vaccine between the ages of 11 and 12.• The HPV vaccine is most effective when administered to girls and women before the onset of sexual activity. Vaccination is also recommended for women up to age 26, regardless of sexual activity.
    36. 36. Incidence of Cervical cancer/ 100,000 cases• US : 7• Jordan 2.6• Egypt 2.7• Israel (Arabs) 2.5• Israel (Jews) 5.3• Cyprus 3.7• Turkey 4.76 Komodiki et al, MECC Monograph Hatipoglu/Ozgul, MOH Turkey
    37. 37. Issues to be addressed• Cross sub-types protection• Duration of protection• Production cost• Time frame for public health impact
    38. 38. Normal Cancer
    39. 39. Normal Cell Tumor Cell
    40. 40. Tumor Cell
    41. 41. 1 2 1
    42. 42. Dendritic CellsB-cellsMacrophage
    43. 43. Normal Cell KILL
    44. 44. L1/2 E5/2/4 E1 E6/7
    45. 45. Viral Proteins Expressed HPV Infection L1, L2Transient PersistentInfection Infection E1, E2, L1, L2 Low Grade E1, E2 Dysplasia CIN 1 Integration High Grade Dysplasia CIN 2/3 Invasive Cancer
    46. 46. • Prevention of HPV infection Viral Proteins Expressed• Elimination of HPV infection HPV Infection L1, L2 Transient Persistent Infection Infection E1, E2, L1, L2 Low Grade• Elimination of low grade dysplasia E1, E2/ E6, E7 Dysplasia CIN 1 Integration High Grade High Grade E6, E7/ E1, E2• Elimination of high grade dysplasia Dysplasia CIN 2/3 CIN 3• Preventions of recurrence Invasive E6, E7 Cancer Treatment of invasive disease
    47. 47. •Prevention of HPV infection Viral Proteins Expressed•Elimination of HPV infection HPV Infection L1, L2 Transient Persistent Infection Infection E1, E2, L1, L2 Low Grade•Elimination of low grade dysplasia E1, E2/ E6, E7 Dysplasia CIN 1 Integration High Grade High Grade E6, E7/ E1, E2•Elimination of high grade dysplasia Dysplasia CIN 2/3 CIN 3•Preventions of recurrence Invasive E6, E7 Cancer Treatment of invasive disease
    48. 48. •Prevention of HPV infection Viral Proteins Expressed•Elimination of HPV infection HPV Infection L1, L2 Transient Persistent Infection Infection E1, E2, L1, L2 Low Grade•Elimination of low grade dysplasia E1, E2/ E6, E7 Dysplasia CIN 1 Integration High Grade High Grade E6, E7/ E1, E2•Elimination of high grade dysplasia Dysplasia CIN 2/3 CIN 3•Preventions of recurrence Invasive E6, E7 Cancer Treatment of invasive disease
    49. 49. Clinical Trials: Preventive Vaccine for CIN Vaccine Type Phase Antigen Lesion Patient Number Immunologic Clinical response Authors responseProtein-based II Fusion protein CIN 3 58 Increased 13/58 CR Einstein 2007 HPV-16 E7- inflammation in 32/58 PR HSP65 responders 11/58 SD 2/58 PD II Fusion protein High grade CIN 21 9/17 7/20 CR Roman 2007 HPV-16 E7- 1/20 PR HSP65 11/20 SD 1/20 PD Viral vector- I/II MVA E2 CIN 1, 2 and 3 36 100% Ab 34/36 CR Gutierez 2004 based particles induction 2/36 PR 100% CTL II MVA E2 CIN 2 or 3 34 100% Ab 19/34 CR Garcia- particles induction 15/34 PR Hernandez CTL response 2006 (data not shown) II Recombinant Anogenital (non- 29 NA 1/29 CR Fiander 2006 HPV-16 E6E7L2 cervical) 5/29 PR protein (TA- neoplasia 3 CIN) and TA- (AGIN 3) HPVChimeric virus- I/II HPV-16L1E7 CIN 2 or 3 39 100% Ab 9/23 PR Kaufmann like particle induction (p>0.05) 2007 5/23 E7-specific T cells DNA I Plasmid CIN 2 or 3 15 Low immune 3/9 PR Trimble 2009 expressing a Sig- response (p>0.05) E7(detox)-HSP 70 fusion protein
    50. 50. Preventive HPV E7 Vaccine / VINKenter GG et al. NEJM 2009
    51. 51. Preventive HPV E7 Vaccine / VIN Kenter GG et al. NEJM 2009
    52. 52. A combined prophylactic therapeutic vaccines
    53. 53. L1/2 E5/2/4 E1 E6/7
    54. 54. Targeting HPV16 E2 Using Chimeric VLPs and E2 Peptides Chimeric VLP: HPV16 VLP-E7E2 L2 L1 E7 E2 HPV16 L1/L2-E7-E2 Chimeric VLP
    55. 55. Conclusion• E2 vaccine induce CTL that lyse the tumor cell line expressing HPV 16 E2.• Modified E2 sequence showed higher binding affinity , elicited strong IFN- production and can, also, induce CTL to lyse B16-AAD-E2.• Combination with VLP priming enhances the E2 CTL• E2 vaccine has a potential for the treatment of CINs.
    56. 56. Provenge/Sipuleucel-T
    57. 57. Prostate antigen: PAP48 hours incubation with GM
    58. 58. Disease Behavior Kantoff PW, N Engl J Med. 2010 Jul 29;363(5):411-22
    59. 59. Sipuleucel-T (Provenge)• Sipuleucel-T (Provenge) is an autologous, dendritic cell- based vaccine (CD541) that is pulsed with a selective prostate antigen: prostatic acid phosphatase.• the FDA has recently approved sipuleucel-T in patients with asymptomatic or minimally symptomatic metastatic hormone-refractory prostate cancer.• Side effects include chills, fatigue, fever, and joint aches.
    60. 60. Framing the Problem Fact I• Cancer Vaccine article
    61. 61. Framing the Problem Fact I• Cancer Vaccine article 18,543
    62. 62. Framing the Problem Fact I• Cancer Vaccine article 18,543• Drug Approved 1
    63. 63. Fact II• Most of the people who received cancer the Vaccine did not benefit
    64. 64. Effectors vs. Suppressors TumorVaccine CD8+ CTL CTL CTL CTL CTL APC CD4+
    65. 65. Ideal Antigenic target• Expressed uniquely in cancer cells• Important for the maintenance of the malignant phenotype• Immunogenic
    66. 66. Mutant Ras
    67. 67. Solid tumor with Ras mutation Sequencing Ras Mutant Ras peptide Toubaji et al, 2008
    68. 68. Post 3 vax.
    69. 69. Post 3 vax.
    70. 70. Patients responses
    71. 71. Effectors vs. Suppressors TumorVaccine CD8+ CTL CTL CTL CTL CTL APC CD4+
    72. 72. Tumor Immune Modulation Network (TIMN) Co-inhibitory-Ligands Tumor CD8+ CTL CTL CTL Cytokines/factors APC CTL CTL Treg Treg Treg CD4+ Treg MDSC Other inhibitory mechanism TAM
    73. 73. PD-1/PD-L1/PD-L2 Blank, Cancer Immunol Immunother, 2005
    74. 74. PD-1/PD-L1 Engagement Suppresses Effector T cells PD-L1 PD-1 Tumor cell MDSC Suppressed/Apoptotic Activated T cell CD28 B7 TCR MHC/ APC Treg peptide
    75. 75. PD-1/PD-L1 Engagement Suppresses Effector T cells PD-L1 PD-1 Tumor cell MDSC Activated T cell CD28 B7 Anti-PD-1 TCR MHC/ APC Treg peptide
    76. 76. Effectors vs. Suppressors CTLA4 CTL Tumor CD8+ CTL CTL CytokinesAPC Treg Treg CTLA4 CTL Treg CD4+ Treg CTL MDSC Other inhibitory mechanism TAM
    77. 77. Improved Overall Survival in a Phase II Randomized Controlled Trial of a Poxviral-Based PSA-Targeted Immunotherapy in Metastatic Castration-Resistant Prostate CancerKantoff, et al. JCO, 2010
    78. 78. Effectors vs. Suppressors PD1-PDL1 CTL Tumor CD8+ CTL CTL CytokinesAPC Treg Treg PD1-PDL1 CTL Treg CD4+ Treg CTL MDSC Other inhibitory mechanism TAM
    79. 79. Humanized mAb PD-L1 PD-L2 to PD-1 PD-1 tumor DCsActivated T cells anti-PD-1 antiboby
    80. 80. Evaluation of therapeutic efficacy of vaccine in combination with anti-PD1 Days 0 7 8 15 22 Monitoring of tumor growth and survival TC-1 E7+aPD1 E7+aPD1 E7+aPD1 Day 7 after implantation of 50,000 TC-1 cell
    81. 81. 1.5 1.5 1.5 Tumor Growth1.2 1.2 1.20.9 0.9 0.90.6 0.6 0.60.3 0.3 0.3 Non-immunized E7 a-PD1 0 0 0 5 8 15 22 26 29 31 35 38 5 8 15 22 26 29 31 35 38 5 8 15 22 26 29 31 35 381.5 1.5 1.5 E7+CPM1.2 1.2 1.20.9 0.9 0.90.6 0.6 0.60.3 0.3 0.3 CPM a-PD1 +CPM 0 0 0 5 8 15 22 26 29 31 35 38 5 8 15 22 26 29 31 35 38 5 8 15 22 26 29 31 35 381.5 1.5 E7+aPD11.2 1.20.90.6 0.9 Tumor Volume, cm30.3 0.6 E7+CPM+aPD1 0 5 8 15 22 26 29 31 35 38 0.3 0 5 8 15 22 26 29 31 35 38 Days after tumor implantation
    82. 82. Enhancing T cell Enhancing Memory response responseEnhancing Agpresentation + + Long Lasting Ag I. R. I. R. Effective I. R.
    83. 83. Enhancing T cell Enhancing Memory response responseEnhancing Agpresentation + + Long Lasting Ag I. R. I. R. _Better Targets Inhibiting negative Effective regulators I. R.
    84. 84. A Pilot Study of CT-011+ Provenge (Sipuleucel- T)+ Cyclophosphamidein Patients with Metastatic Castrate Resistant Prostate Cancer
    85. 85. Study Design Part 1, Run In phase, up to 12 patients CPM 250 mg/m2 (Day -1 only) + Sipuleucel –T (Day 0) >>> Q 2 wk X 3 CPM 125 mg/m2 Part 2, Randomized, total 45 patients • Sipuleucel-T Q 2 wk X 3 • Sipuleucel-T Day 0 + CT-011 (3 mg/kg Day 2) • CPM (Day -1 only) + Sipuleucel-T Day 0 + CT-011 (3 mg/kg Day 2)CPM = Low Dose CyclophosphamideApheresis 2-3 days prior to each dose of Sipuleucel-T for cell generation
    86. 86. ObjectivesPrimary endpoint Secondary endpoint• Feasibility Provenge™+ • Progression-free CPM survival• Immune efficacy on • Overall survival PA2024-specific IFN-γ • Toxicity
    87. 87. Inclusion Criteria• mCRPC with progression, testosterone < 50 ng/dL.• PSA over nadir at least 2X, 3 weeks apart.• No prior chemotherapy.
    88. 88. Disease Behavior
    89. 89. A Pilot Study to of Gemcitabine andCT-011 in Resected Pancreatic Cancer
    90. 90. Phase I trial of escalating doses of CT-011 incombination with p53 vaccine in adults with advanced solid tumors
    91. 91. Effectors vs. Suppressors CTLA4 CTL Tumor CD8+ CTL CTL CytokinesAPC Treg Treg CTLA4 CTL Treg CD4+ Treg CTL MDSC Other inhibitory mechanism TAM
    92. 92. PD-1 and CTLA-4 combination blockade expands infiltrating T cells and reduces regulatory T and myeloid cells(Curran MA et al, PNAS 2010)
    93. 93. Tumor Growth Pattern with Chemotherapy and Vaccines Fojo et.al. Clin Cancer Res, 2010
    94. 94. treatment Fojo et.al. Clin Cancer Res, 2010
    95. 95. treatment Chemotherapy Fojo et.al. Clin Cancer Res, 2010
    96. 96. treatment Chemotherapy Vaccines Fojo et.al. Clin Cancer Res, 2010
    97. 97. Disease Behavior
    98. 98. Disease Behavior
    99. 99. Pt Age Cancer Pathology Mutation Pre-vaccination Stage on Disease extension Arm A Patient Profile: Il-2 Therapy Enrollment on enrollment Liver, Lung, Para-1A 56 Colon Adenocarcinoma ASP Sx1, Cx3 4 aortic LN2A 57 Colon Adenocarcinoma ASP Sx2, Cx2 4 Lung3A 62 Colon Adenocarcinoma ASP Sx1, Cx3, Rx1 4 Lung Pancreas, Liver, Adrenals, Omentum,4A Peritoneum, Infradiaphragmati 50 Pancreatic Adenocarcinoma ASP Sx1, Cx2 4 c LN5A 60 Lung Adenocarcinoma VAL Cx2, Rx1 4 Chest Wall6A 59 Colon Adenocarcinoma VAL Cx2 4 Liver7A 52 Colon Adenocarcinoma CYS Sx2, Cx2 NED NED Lung, Hilar8A 68 Lung NSCLC CYS Sx1, Cx5, Rx2 4 Adenopathy9A 56 Colon Adenocarcinoma ASP Sx2, Cx3, Rx2 NED NED10A 63 Colon Adenocarcinoma ASP Sx3, Cx3 4 Liver, Lung11A 42 Pancreatic Adenocarcinoma ASP Sx2, Cx1 NED NED12A 39 Colon Adenocarcinoma ASP Sx1, Cx1 NED NED13A 67 Colon Adenocarcinoma CYS Sx1, Cx2 4 Liver14A 51 Colon Adenocarcinoma ASP Sx2, Cx1 NED NED15A 60 Pancreatic Adenocarcinoma VAL Sx1, Cx4, Rx1 4 Liver16A
    100. 100. # Of Off-study reason/DiseasePt Cycles A Clinical Outcome:PFS (ms)# Arm Status Il-2 OS (ms)*1A 2 PD 0.5 5.52A 3 PD 3.9 16.83A 3 PD 5.8 21.54A 3 PD 3.6 6.25A 1 PD 1 2.86A 3 PD 3.5 8.97A 11 Completed 129+ 129+8A 3 PD 3.6 13.19A 10 PPS 15.4 37.210A 3 PD 3.3 19.911A 6 PD 7.5 24.112A 3 PD 6.2 2313A 3 PD 3.5 4.814A 3 PD 7.1 41.3 PPS/Lost to Follow-15A 3 Up 2.7+ 5.3
    101. 101. Study Design Re-staging-7 1 8 15 22 1 8 15 22 1 For 6 cycles or Days until disease progression or SE
    102. 102. ObjectivesPrimary endpoint Secondary endpoint• Feasibility and safety • Clinical response
    103. 103. Inclusion Criteria• Resectable pancreatic CA• Eligible for Gemcitabine• No previous treatment
    104. 104. Schema• •Staging •Staging •Staging evaluation •evaluatio •evaluatio n n 1(0) 2 (3) 3 (6) 4 (9) 5 (12) 6 (15) 7 (18) Until Cycles (Weeks) disease p53 (264-272) progression or CT-011 toxicity
    105. 105. Schema• Dose Escalation Schedule Dose Level Dose of CT-011^ Level 1 0.3 mg/kg every 3 weeks X 2 * Level 2 1.5 mg/kg every 3 weeks X 2 * Level 3 3 mg/kg every 3 weeks X 2 * Level 4 6 mg/kg every 3 weeks X 2 * ^Evaluation of toxicity to define DLT will be performed 3 weeks after the 1st dose (cycle 1). *Evaluation of clinical status and tumor response will be performed after 2 cycles
    106. 106. Improved Overall Survival in a Phase II Randomized Controlled Trial of a Poxviral-Based PSA-Targeted Immunotherapy in Metastatic Castration-Resistant Prostate CancerKantoff, et al. JCO, 2010
    107. 107. Disease Behavior
    108. 108. Disease Behavior Show the difference in vaccine vehcle and then Here add the understanding of the TIRN and Then show CTLA and potentially PD1 for use together
    109. 109. T regs in colon cancer patients p<0.05 * 12 11 %CD4+CD25+FOXP3+ 10 9 8%CD4+CD25+FOXP3+ 7 6 5 4 3 2 1 0 Normal NORMAL Colon Cancer COLON CANCER COLON CANCER WIT H RAS MUT AT ION WIT HOUT RAS MUT AT Ion
    110. 110. Successful cancer immunotherapy requires simultaneous targeting of both arms of immune system Induction of immune Inhibition of response suppression Bhardwaj et al. 2007
    111. 111. Successful cancer immunotherapy requires simultaneous targeting of both arms of immune system Induction of immune Inhibition of response suppression Bhardwaj et al. 2007
    112. 112. Successful cancer immunotherapy requires simultaneous targeting of both arms of immune system Induction of immune Inhibition of response suppression Bhardwaj et al. 2007
    113. 113. Effectors vs. Suppressors TumorVaccine CD8+ CTL CTL CTL CTL CTL APC CD4+
    114. 114. Long LastingAg I. R. I. R. Effective I. R.
    115. 115. Enhancing T cell Enhancing Memory response responseEnhancing Agpresentation + + Long Lasting Ag I. R. I. R. Effective I. R.
    116. 116. SC Low dose IL-2 and Vaccines
    117. 117. Immune Modulating Therapy
    118. 118. Enhancing effector Enhancing memory T cell response responseEnhancing Agpresentation + + Long Lasting Ag I. R. I. R. Effective I. R.
    119. 119. Enhancing effector Enhancing memory T cell response responseEnhancing Agpresentation + + Long Lasting Ag I. R. I. R. _Immune Evasion Immune Negative Effective regulation I. R.
    120. 120. Enhancing T cell Enhancing Memory response responseEnhancing Agpresentation + + Long Lasting Ag I. R. I. R. _Better Targets Inhibiting negative Effective regulators I. R.
    121. 121. Combination Immune Therapy• Immune Enhancing• Immune Modulating • Targeting inhibitory cells • Blocking inhibitory cytokines/factors (anti- IL10, TGFb etc) • Blocking co-inhibitory molecules (anti-PD1, CTLA-4 etc)
    122. 122. Immune Combination Therapy• Vaccine• Immune modulating and enhancing agents• Antibodies• Targeted Therapy• Chemotherapy• Radiotherapy
    123. 123. Evaluation of therapeutic efficacy of vaccine in combination with anti-PD1 Days 0 8 15 22 Monitoring of tumor growth and survival TC-1 E7+aPD1 E7+aPD1 E7+aPD1Tumor Volume, cm3 Non-treated E7 n=10 n=9 E7+ aPD1 aPD1 n=10 n=8 Days after tumor implantation
    124. 124. Evaluation of therapeutic efficacy of vaccine/CT-011 combination in TC-1 mouse model Days 0 8 15 22 Monitoring of tumor growth and survival TC-1 E7+aPD1 E7+aPD1 E7+aPD1 120 100 Percent Survival 80Tumor Volume, cm3 Non-treated E7 n=10 n=9 60 40 20 0 E7+ CT-011 CT-011 10 15 20 25 30 35 40 45 n=10 n=8 Days after tumor implantation Days after tumor implantation
    125. 125. Treg cell inhibitor-cyclophosphamide (CPM)Low Dose CPM selectively targets Treg cells, leaving other T cellpopulations intact (Lutsiak et al, Blood, 2005; Ikezawa et al, J Dermatol Sci, 2005).
    126. 126. Effectors vs. Suppressors PD-1 PD-L1 CTL Tumor CD8+ CTL CTL CytokinesAPC Treg PD-1 PD-L1 Treg CTL Treg CD4+ Treg CTL MDSC Other inhibitory mechanism TAM
    127. 127. Therapeutic efficacy of vaccine in combination with anti-PD1 and CPM CPM Days 0 7 8 15 22 Monitoring of tumor growth and survival TC-1 E7+aPD1 E7+aPD1 E7+aPD1 Day 7 after implantation of 50,000 TC-1 cell
    128. 128. 1.5 1.5 1.5 Tumor Growth1.2 1.2 1.20.9 0.9 0.90.6 0.6 0.60.3 0.3 0.3 Non-immunized E7 aPD1 0 0 0 5 8 15 22 26 29 31 35 38 5 8 15 22 26 29 31 35 38 5 8 15 22 26 29 31 35 381.5 E7+CPM1.20.90.60.3 CPM 0 5 8 15 22 26 29 31 35 38 E7+CT-011
    129. 129. 1.5 1.5 1.5 Tumor Growth1.2 1.2 1.20.9 0.9 0.90.6 0.6 0.60.3 0.3 0.3 Non-immunized E7 aPD1 0 0 0 5 8 15 22 26 29 31 35 38 5 8 15 22 26 29 31 35 38 5 8 15 22 26 29 31 35 381.5 1.5 1.5 E7+CPM1.2 1.2 1.20.9 0.9 0.90.6 0.6 0.60.3 0.3 0.3 CPM aPD1+CPM 0 0 0 5 8 15 22 26 29 31 35 38 5 8 15 22 26 29 31 35 38 5 8 15 22 26 29 31 35 381.5 E7+aPD11.20.90.60.3 0 5 8 15 22 26 29 31 35 38
    130. 130. Vaccine in combination with anti-PD1 and CPM Synergize in tumor rejection Non-treated (n=15) E7 (n=14) a-PD1 (n=15) CPM (n=15) E7+ a-PD1(n=15) E7+CPM (n=14) a-PD1 +CPM (n=15) 100 E7 + a-PD1 + CPM (n=20) 80Percent Survival 60 40 20 0 8 12 16 20 24 28 32 36 40 44 48 52 56 60 64 68 72 76 Days after tumor implantation
    131. 131. Vaccine in combination with anti-PD1 and CPMinduces potent antigen-specific immune responses in tumor bearing mice Days 0 7 8 15 21 TC-1 tumor CPM E7+a-PD1 TERMINATION 140 ***Number of IFN spots per 106 *** R2=0.8106 120 *** Number of IFN spots per 106 120 P<0.001 100 splenocytes splenocytes 80 80 60 40 40 20 0 0 E7 E7 E7 E7 a-PD1 NT 0.0 0.3 0.6 0.9 1.2 1.5 a-PD1 +CPM a-PD1 +CPM Tumor volume, cm3 ***P<0.001 +CPM
    132. 132. Vaccine in combination with anti-PD1 and CPM increases the levels of tumor-infiltrated CD8+ T cells 9.E+03 8.E+03 *** CD8+ cells *** * Absolute numbers per 7.E+03 10e6 tumor cells 6.E+03 5.E+03 4.E+03 3.E+03 2.E+03 1.E+03 0.E+00 E7 E7 E7 E7+ a-PD1 NT +CPM a-PD1 a-PD1 +CPM +CPM 12 *** * *** 10 * * CD8+/Treg Ratio 8 6 4 2 0 E7 E7 E7 E7+ a-PD1 NT +CPM a-PD1 a-PD1 +CPM +CPM*P<0.05, **P<0.01 and ***P<0.001
    133. 133. Anti-PD1 partially overcomes tumor-induced suppression of stimulated Tconv cell proliferation in vitro Isotype control anti-PD-L1 100 25 *** 25 Percent of maximum *** 20 15 10 5 0 +TC-1 Tconv# +TC-1 +TC-1 +TC-1 +Irr Ab + a-PD1 +PD-L1***P<0.001
    134. 134. Anti-PD1 and CPM synergize to decrease the level splenic and tumor infiltrated Treg cells ** Day 21 40 *Treg - % of CD4+ cells 35 30 25 20 15 10 5 2.5E+03 0 ** Treg cells E7 E7 E7 E7 CT-011 NT * + a-PD1 +CPM +a-PD1 +CPM 2.0E+03 ** +CPM Absolute numbers per * * 10e6 tumor cells 1.5E+03 ** 1.0E+03 5.0E+02 *P<0.05 and **P<0.01 0.0E+00 E7 E7 E7 E7+ aPD1 NT +CPM + a-PD1 a-PD1 +CPM +CPM
    135. 135. Anti-PD1/CPM synergize to decrease and maintain low level of Tregs in periphery 25 % of Tregs within CD4+T cell population 20 NT CPM a-PD1 15 a-PD1 +CPM NS NS 10 * * ** 5 ** ** 0Tumor (days) 7 8 11 14 18 21 26 CPM (days) 0 +1 +4 +7 +11 +14 +19
    136. 136. T cell subsets involved in tumor protection after Combination of Vaccine with anti-PD1/CPM Anti-CD8 Anti-CD8 400ug i.p. 400ug i.p. TC-1 Days 0 5 7 8 15 17 22 24 Monitoring of tumor growth and survival Anti-CD4 Vaccine + Vaccine + Anti-CD4 Vaccine + 300ug i.p. a-PD1 a-PD1 300ug i.p. a-PD1 CPM 100 No treatmentPercent Survival 80 E7+a-PD1+CPM E7+a-PD1+CPM+anti-CD8 60 E7+a-PD1+CPM+anti-CD4 /anti-CD8 40 E7+a-PD1+CPM+anti-CD4 20 0 8 13 18 23 28 33 38 43 48 53 58 63 68 73 78 Days after tumor implantation
    137. 137. Vaccine/anti-PD1/CPM: Mechanism of Action PD-1 PD-L1 CTL Tumor CD8+ CTL CTL CTL CTLCTL TGFAPC Treg CD4+ PD-1 PD-L1 Treg Treg CD4+ CTL CD4+ Treg PD-1 PD-L1
    138. 138. Summary• Combination of anti-PD1 with Treg cell inhibition (CPM) and vaccine is a feasible strategy that results in tumor eradication under challenging conditions• Anit-PD1 enhances CD8+ tumor infiltration when combined with vaccine and CPM• Anti-PD1 synergize with CPM to decrease both peripheral and tumor-infiltrated Treg cells
    139. 139. Current Clinical Trials• A Pilot Study to of Gemcitabine and CT-011 in Resected Pancreatic Cancer• Phase I trial of escalating doses of CT-011 in combination with p53 vaccine in adults with advanced solid tumors• A Pilot Study of CT-011 and Provenge (Sipuleucel-T) in combination with low dose Cyclophosphamide in Patients with Metastatic Castrate Resistant Prostate Cancer
    140. 140. B7-DC-Ig PD-L1 PD-L2 PD-1 Chimera of ECD of murine PD-L2 and Fc portion of IgG tumor DCsActivated T cells
    141. 141. Vaccine in combinations with B7-DC-Ig and CPM induce potent antigen-specific immune responses in tumor bearing miceNumber of IFN spots per 10e6 splenocytes *** E7 E7 E7 E7 B7DCIg NT +AMP-224 +CPM +B7DCIg +CPM +CPM *P<0.05, ***P<0.001
    142. 142. Vaccine in combinations with B7-DC-Ig and CPM induce potent antigen-specific immune responses in tumor bearing miceNumber of IFN spots per 10e6 splenocytes *** 30 Percent of apoptotic TC-1 cells E:T=10:1 25 *** *** E:T=25:1 * E:T=50:1 20 15 10 5 E7 E7 E7 E7 AMP-224 NT 0 E7 E7/ E7/ E7/ CPM/ NT + B7DCIg +CPM + B7DCIg +CPM B7DCIg CPM CPM/ AMP-224 +CPM B7DCIg *P<0.05, ***P<0.001
    143. 143. Vaccine in combinations with B7-DC-Ig and CPM induce potent antigen-specific immune responses in tumor bearing mice 120 100 Non-treated (n=15)Percent Survival B7DCIg (n=15) 80 E7 (n=15) E7+ B7DCIg (n=13) 60 B7DCIg +CPM (n=15) 40 E7+CPM (n=15) CPM (n=15) 20 E7 + AMP-224 + CPM (n=15) 0 10 15 20 25 30 35 40 45 50 55 60 65 70 Days after tumor implantation
    144. 144. Vaccine in combinations with B7-DC-Ig and CPM induce potent antigen-specific immune responses in tumor bearing mice 120 100 Non-treated (n=15) Percent Survival B7DCIg (n=15) 80 E7 (n=15) E7+ B7DCIg (n=13) 60 B7DCIg +CPM (n=15) 40 E7+CPM (n=15) CPM (n=15) 20 E7 + AMP-224 + CPM (n=15) 0 10 15 20 25 30 35 40 45 50 55 60 65 70 Days after tumor implantation 100 No treatment E7+B7DCIg+CPMPercent Survival 80 E7+B7DCIg+CPM+anti-CD8 60 40 20 0 8 13 18 23 28 33 38 43 48 53 58 63 68 73 78 Days after tumor implantation
    145. 145. T regulatory Cells Treg Anti-PD1 ** Treg cellsNnumbers per 10e6 tumor * ** * * ** cells E7 E7 E7 E7+ +aPD1 NT +CPM +aPD1 +aPD1 +CPM +CPM *P<0.05, **P<0.01, ***P<0.001
    146. 146. T regulatory Cells Treg Anti-PD1 B7-DC-Ig ** Treg cells *** Treg cellsNnumbers per 10e6 tumor * ** ** ** * * * ** ** ** cells ** E7 E7 E7 E7+ +aPD1 NT E7 E7+ E7+ E7+ B7DCIg + NT +CPM +aPD1 +aPD1 +CPM CPM B7DCIg CPM+ CPM +CPM B7DCIg *P<0.05, **P<0.01, ***P<0.001
    147. 147. T Conventional Cells Tconv Anti-PD1 Numbers per 10e6 tumor cells 3500 * 3000 2500 2000 * 1500 1000 500 0 E7 E7 E7 E7+ aPD1 NT +CPM +aPD1 aPD1 +CPM +CPM*P<0.05, ***P<0.001
    148. 148. T Conventional Cells Tconv Anti-PD1 B7-DC-Ig Numbers per 10e6 tumor cells 3500 3500 * *** 3000 3000 2500 2500 2000 * 2000 1500 1500 1000 1000 500 500 0 E7 E7 E7 E7+ aPD1 0 E7 E7+ E7+ E7+ CPM+ NT NT +CPM +aPD1 aPD1 +CPM CPM B7DCIg CPM/ B7DCIg +CPM B7DCIg*P<0.05, ***P<0.001
    149. 149. CD8+ T Cells CD8 Anti-PD1 AMP-224 *** CD8+ cells * CD8+ cells ***Numbers per 10e6 tumor * NS *** cells E7 E7+ E7+ E7+ AMP + NT E7 E7 E7 E7+ aPD1 NT +CPM +aPD1 aPD1 CPM AMP CPM+ CPM +CPM +CPM AMP *P<0.05, ***P<0.001
    150. 150. CD8+ T Cells CD8 Anti-PD1 B7-DC-Ig *** CD8+ cells * CD8+ cells ***Numbers per 10e6 tumor * NS *** cells E7 E7+ E7+ E7+ B7DCIg NT E7 E7 E7 E7+ aPD1 NT +CPM +aPD1 aPD1 CPM B7DCIg CPM+ CPM +CPM +CPM B7DCIg *P<0.05, ***P<0.001
    151. 151. B7-DC-Ig decreases the level of tumor infiltrated PD-1high CD8+T cells 5000Numbers per 10e6 tumor cells 4500 * 4000 NS *** 3500 3000 2500 2000 1500 1000 500 0 E7 E7+ E7+ E7+ CPM B7DCIg B7DCIg + CPM
    152. 152. B7-DC-Ig decreases the level of tumor infiltrated PD-1high CD8+T cells 5000 TI CD8+ PD-1high cells and CD8+ PD-1low cellsNumbers per 10e6 tumor cells 4500 5000 * 4000 NS 4500 *** 3500 4000 3000 3500 2500 3000 2000 2500 1500 2000 1000 1500 500 1000 0 500 E7 E7+ E7+ E7+ 0 CPM B7DCIg B7DCIg E7 E7+ E7+ E7+ + CPM CPM B7DCIg B7DCIg CPM GATED ON CD8+ PD-1high PD-1high 53.81% 25.75% GATED ON CD8+ PD-1high PD-1high 67.82% 23.36%
    153. 153. B7-DC-Ig decreases the level of tumor infiltrated PD-1high CD8+T cells 5000 TI CD8+ PD-1high cells and CD8+ PD-1low cellsNumbers per 10e6 tumor cells 4500 5000 * 4000 NS 4500 *** 3500 4000 3000 3500 2500 3000 2000 2500 1500 2000 1000 1500 500 1000 0 500 E7 E7+ E7+ E7+ 0 CPM B7DCIg B7DCIg E7 E7+ E7+ E7+ TI CD8+ PD-1low cells + CPM 4000 CPM B7DCIg B7DCIg Numbers per 10e6 tumor cells 3500 CPM * GATED ON CD8+ * PD-1high PD-1high 3000 * 2500 53.81% 25.75% 2000 1500 1000 500 GATED ON CD8+ PD-1high PD-1high 0 E7 E7+ E7+ E7+ 67.82% 23.36% CPM B7DCIg B7DCIg + CPM
    154. 154. In contrast to anti-PD1, B7DCIg doesn’t overcome tumor-induced suppression of stimulated Tconv cell proliferation Percent of maximum *** *** *** Tconv +TC-1 +TC-1 +TC-1 +TC-1 +Irr Ab +CT-011 +AMP-224
    155. 155. Immune Modulating Agents• Does it act as we want it to ?• What else would it do within the context of TIMN ?
    156. 156. Final Rankings of Agents with High Potential for Use in Treating CancerNCI Immunotherapy Agent Workshop, 2007
    157. 157. Houot R and Levy R, Blood, 2011Combinations of anti- OX40, CTLA4, GITR, and 4 (FR4)) with Intratumoral CpG Houot R and Levy R, Blood, 2011 Houot R and Levy R, Blood, 2011
    158. 158. Vaccine and Anti-Transforming Growth Factor-βAnti–TGF-β (1D11) synergistically enhanced the efficacy of a CTL-inducing peptide tumor vaccine consisting of HPV16 E749-57peptide, emulsified in incomplete Freunds adjuvant togetherwith Terabe M et al, Clin Can Res 2009
    159. 159. Synergistic antitumor effect of anti-GITR and anti-CTLA4DTA-1 is an anti-GITR mAb, 4F10 is an anti-CTLA4 mAb Ko et al, JEM, 2005
    160. 160. What did we learn?• Lack of understanding of simple biology--- Simplistic approach, “single agent approach”• Difference in tumor behavior--- “wrong monitoring”• Clinical trial design--- chemotherapy based approach
    161. 161. Immune Combination Therapy• Science
    162. 162. Immune Combination Therapy• Science• Toxicity
    163. 163. Immune Combination Therapy• Science• Toxicity• Regulatory/ Clinical Trial Design
    164. 164. Immune Combination Therapy• Science• Toxicity• Regulatory/ Clinical Trial Design• Business/IP and Data Sharing
    165. 165. Immune Combination Therapy• Combination• Should understand the agent• Regulatory/ Clinical Trial Design• Business/IP and Data Sharing
    166. 166. What did we learn?• Lack of understanding of simple biology--- Simplistic approach, “single agent approach”• Difference in tumor behavior--- “wrong monitoring”• Clinical trial design--- chemotherapy based approach
    167. 167. Improved Overall Survival in a Phase II Randomized Controlled Trial of a Poxviral-Based PSA-Targeted Immunotherapy in Metastatic Castration-Resistant Prostate CancerKantoff, et al. JCO, 2010
    168. 168. Improved Overall Survival in a Phase II Randomized Controlled Trial of a Poxviral-Based PSA-Targeted Immunotherapy in Metastatic Castration-Resistant Prostate CancerKantoff, et al. JCO, 2010
    169. 169. What did we learn?• Lack of understanding of simple biology--- Simplistic approach, “single agent approach”• Difference in tumor behavior--- “wrong monitoring”• Clinical trial design--- chemotherapy based approach
    170. 170. Dose determination and Phase I need?
    171. 171. Clinical Trial Design1. Do cancer vaccines have significant toxicity?2. Does dose escalation effect toxicity?3. Does dose escalation effect response and how is it measured?
    172. 172. Phase 1 for Cancer Therapy• 3+3 Cohort Expansion with Dose escalation• Patients with Late Stage Disease• Determine DLT and MTD• Cancer vaccines – Vaccines are inherently safe, if have toxicity seen in long term – Patients will have weak immune response b/c of advanced disease
    173. 173. Do phase 1 trials for cancer vaccines have significant toxicity?• Reviewed phase 1 trials for therapeutic cancer vaccines – 1990/2011 – 241 studies, 4,952 patients – Excluding all combination therapies
    174. 174. Do phase 1 trials for cancer vaccines have significant toxicity? Toxicity vs. patient number # of Trials Patients Grade 3/4 rxns Percentage Related (%)Autologous 88 1692 37 2.19% 1.36% DC 58 922 9 0.98% 0.33% Tumor 30 770 28 3.64% 2.60%Allogeneic Cells 17 407 22 5.41% 1.23%Synthetic 136 2853 108 3.79% 1.23% Peptide 68 1333 40 3.00% 0.83% DNA 17 311 1 0.32% 0.32% RNA 2 36 0 0% 0% Virus/Bacteria 36 761 50 6.57% 2.63% Anti-idiotypic 10 362 15 4.14% 0% Liposomal 3 45 2 4.44% 4.44% TOTAL 241 4952 167 3.37% 1.25%
    175. 175. Do phase 1 trials for cancer vaccines have significant toxicity? Toxicity vs Vaccine number # of Trials Patients Vaccines Grade 3/4 rxns Percentage Related (%)Autologous 73 1301 5722 20 0.35% 0.14% DC 51 796 3424 9 0.26% 0.09% Tumor 22 505 2298 11 0.48% 0.22%Allogenic Cells 16 347 1874 22 1.17% 0.26%Synthetic 117 2376 14239 78 0.55% 0.21% Peptide 61 1183 7637 37 0.48% 0.12% DNA 15 259 1388 1 0.07% 0.07% RNA 2 36 335 0 0% 0% Virus/Bacteria 30 594 2714 31 1.14% 0.66% Anti-idiotypic 7 266 1938 7 0.36% 0% Liposomal 2 38 227 2 0.88% 0.88% TOTAL 206 4024 21835 120 0.55% 0.20%
    176. 176. Does dose escalation effect toxicity?• Correlation between dose and toxicity• Dose escalation trials – 127 trials – Grade 3/4 Toxicities # of Trials Patients Grade 3/4 rxns Trials w/DLT Autologous 40 847 11 0 DC 27 466 0 0 Tumor 13 381 11 0 Allogenic Cells 5 130 20 1 Synthetic 83 2008 67 2 Peptide 36 852 10 0 DNA 12 208 1 0 Virus/Bacteria 26 592 47 2 Anti-idiotypic 8 339 7 0 Liposomal 1 17 2 0 TOTAL 127 2985 98 3

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