Cancer Immunology, Immunotherapy

, Volume 60, Issue 8, pp 1137–1146 | Cite as

CTLA-4 blockade increases antigen-specific CD8+ T cells in prevaccinated patients with melanoma: three cases

  • Jianda Yuan
  • Brian Ginsberg
  • David Page
  • Yanyun Li
  • Teresa Rasalan
  • Humilidad F. Gallardo
  • Yinyan Xu
  • Sylvia Adams
  • Nina Bhardwaj
  • Klaus Busam
  • Lloyd J. Old
  • James P. Allison
  • Achim Jungbluth
  • Jedd D. WolchokEmail author
Focussed Research Review



Anti-cytotoxic T-lymphocyte antigen-4 (CTLA-4) antibodies, such as ipilimumab, have generated measurable immune responses to Melan-A, NY-ESO-1, and gp100 antigens in metastatic melanoma. Vaccination against such targets has potential for immunogenicity and may produce an effector-memory T-cell response.


To determine the effect of CTLA-4 blockade on antigen-specific responses following vaccination, in-depth immune monitoring was performed on three ipilimumab-treated patients prevaccinated with gp100 DNA (IMF-24), gp100209–217 and tyrosinase peptides plus GM-CSF DNA (IMF-32), or NY-ESO-1 protein plus imiquimod (IMF-11); peripheral blood mononuclear cells were analyzed by tetramer and/or intracellular cytokine staining following 10-day culture with HLA-A*0201-restricted gp100209–217 (ITDQVPFSV), tyrosinase369–377 (YMDGTMSQV), or 20-mer NY-ESO-1 overlapping peptides, respectively. Tumors from IMF-32 were analyzed by immunohistochemistry to help elucidate mechanism(s) underlying tumor escape.


Following vaccination, patients generated weak to no CD4+ or CD8+ T-cell response specific to the vaccine antigen but demonstrated increases in effector-memory (CCR7loCD45RAlo) tetramer+CD8+ T cells. After ipilimumab induction, patients experienced a robust, although sometimes transient, antigen-specific response for gp100 (IMF-32 and IMF-24) or NY-ESO-1 (IMF-11) and produced polyfunctional intracellular cytokines. Primary and metastatic tumors expressed tyrosinase but not gp100 or class I/II MHC molecules.


Vaccination induced a measurable antigen-specific T-cell response that increased following CTLA-4 blockade, potentially “boosting” the vaccine-primed response. Tumor escape may be related to antigen loss or lack of MHC expression necessary for immune activity. These results in a limited number of patients support the need for further research into combining vaccination with ipilimumab and provide insight into mechanisms underlying tumor escape.


Melanoma Cytotoxic T-lymphocyte antigen-4 Ipilimumab T-cell response Vaccines PIVAC 10 



Absolute lymphocyte count


Cytotoxic T-lymphocyte antigen-4


Granulocyte macrophage colony-stimulating factor


Hematoxylin and eosin


Inducible co-stimulator


Intracellular cytokine staining




Peripheral blood mononuclear cell


White blood cell



We thank Dr. Immanuel Luescher from Tetramer Core, Lausanne Branch, Ludwig Institute of Cancer Research, for providing the tetramers; Cora Mariano from the MSKCC Departments of Pathology for the collection of tumor tissues; and Hematology for the lymphocyte counts. This work was supported by Swim Across America, the Experimental Therapeutics Center of MSKCC and Ludwig Foundation. JDW was supported by a Damon Runyon-Lilly Clinical Investigator Award. JY was supported by MSKCC Pilot Grant of Grant number P50AT002779 from the National Center for complementary and Alternative Medicine (NCCAM) and the Office of Dietary Supplements (ODS). Editorial and writing assistance was provided by StemScientific with funding from Bristol-Myers Squibb Company.

Supplementary material

262_2011_1011_MOESM1_ESM.pdf (663 kb)
Supplementary material 1 (PDF 662 kb)


  1. 1.
    Jemal A, Siegel R, Ward E et al (2008) Cancer statistics, 2008. CA Cancer J Clin 58:71–96PubMedCrossRefGoogle Scholar
  2. 2.
    Atkins MB, Lotze MT, Dutcher JP et al (1999) High-dose recombinant interleukin 2 therapy for patients with metastatic melanoma: analysis of 270 patients treated between 1985 and 1993. J Clin Oncol 17:2105–2116PubMedGoogle Scholar
  3. 3.
    Eigentler TK, Caroli UM, Radny P, Garbe C (2003) Palliative therapy of disseminated malignant melanoma: a systematic review of 41 randomised clinical trials. Lancet Oncol 4:748–759PubMedCrossRefGoogle Scholar
  4. 4.
    Chapman PB, Einhorn LH, Meyers ML et al (1999) Phase III multicenter randomized trial of the Dartmouth regimen versus dacarbazine in patients with metastatic melanoma. J Clin Oncol 17:2745–2751PubMedGoogle Scholar
  5. 5.
    Petrella T, Quirt I, Verma S et al (2007) Single-agent interleukin-2 in the treatment of metastatic melanoma: a systematic review. Cancer Treat Rev 33:484–496PubMedCrossRefGoogle Scholar
  6. 6.
    Karbach J, Gnjatic S, Bender A et al (2010) Tumor-reactive CD8+ T-cell responses after vaccination with NY-ESO-1 peptide, CpG 7909 and Montanide ISA-51: association with survival. Int J Cancer 126:909–918PubMedGoogle Scholar
  7. 7.
    Wolchok JD, Yuan J, Houghton AN et al (2007) Safety and immunogenicity of tyrosinase DNA vaccines in patients with melanoma. Mol Ther 15:2044–2050PubMedCrossRefGoogle Scholar
  8. 8.
    Perales MA, Yuan J, Powel S et al (2008) Phase I/II study of GM-CSF DNA as an adjuvant for a multipeptide cancer vaccine in patients with advanced melanoma. Mol Ther 16:2022–2029PubMedCrossRefGoogle Scholar
  9. 9.
    Spaner DE, Astsaturov I, Vogel T et al (2006) Enhanced viral and tumor immunity with intranodal injection of canary pox viruses expressing the melanoma antigen, gp100. Cancer 106:890–899PubMedCrossRefGoogle Scholar
  10. 10.
    Smith CL, Dunbar PR, Mirza F et al (2005) Recombinant modified vaccinia Ankara primes functionally activated CTL specific for a melanoma tumor antigen epitope in melanoma patients with a high risk of disease recurrence. Int J Cancer 113:259–266PubMedCrossRefGoogle Scholar
  11. 11.
    Lacelle MG, Jensen SM, Fox BA (2009) Partial CD4 depletion reduces regulatory T-cells induced by multiple vaccinations and restores therapeutic efficacy. Clin Cancer Res 15:6881–6890PubMedCrossRefGoogle Scholar
  12. 12.
    Nicholaou T, Ebert LM, Davis ID et al (2009) Regulatory T-cell-mediated attenuation of T-cell responses to the NY-ESO-1 ISCOMATRIX vaccine in patients with advanced malignant melanoma. Clin Cancer Res 15:2166–2173PubMedCrossRefGoogle Scholar
  13. 13.
    Krummel MF, Allison JP (1995) CD28 and CTLA-4 have opposing effects on the response of T-cells to stimulation. J Exp Med 182:459–465PubMedCrossRefGoogle Scholar
  14. 14.
    Salomon B, Lenschow DJ, Rhee L et al (2000) B7/CD28 costimulation is essential for the homeostasis of the CD4+CD25+ immunoregulatory T-cells that control autoimmune diabetes. Immunity 12:431–440PubMedCrossRefGoogle Scholar
  15. 15.
    Karandikar NJ, Vanderlugt CL, Walunas TL et al (1996) CTLA-4: a negative regulator of autoimmune disease. J Exp Med 184:783–788PubMedCrossRefGoogle Scholar
  16. 16.
    Brunner MC, Chambers CA, Chan FK et al (1999) CTLA-4-mediated inhibition of early events of T-cell proliferation. J Immunol 162:5813–5820PubMedGoogle Scholar
  17. 17.
    van Elsas A, Hurwitz AA, Allison JP (1999) Combination immunotherapy of B16 melanoma using anti-cytotoxic T lymphocyte-associated antigen 4 (CTLA-4) and granulocyte/macrophage colony-stimulating factor (GM-CSF)-producing vaccines induces rejection of subcutaneous and metastatic tumors accompanied by autoimmune depigmentation. J Exp Med 190:355–366PubMedCrossRefGoogle Scholar
  18. 18.
    Maker AV, Attia P, Rosenberg SA (2005) Analysis of the cellular mechanism of antitumor responses and autoimmunity in patients treated with CTLA-4 blockade. J Immunol 175:7746–7754PubMedGoogle Scholar
  19. 19.
    Quezada SA, Peggs KS, Curran MA, Allison JP (2006) CTLA4 blockade and GM-CSF combination immunotherapy alters the intratumor balance of effector and regulatory T-cells. J Clin Invest 116:1935–1945PubMedCrossRefGoogle Scholar
  20. 20.
    Quezada SA, Peggs KS, Simpson TR et al (2008) Limited tumor infiltration by activated T effector cells restricts the therapeutic activity of regulatory T-cell depletion against established melanoma. J Exp Med 205:2125–2138PubMedCrossRefGoogle Scholar
  21. 21.
    Hodi FS, Mihm MC, Soiffer RJ et al (2003) Biologic activity of cytotoxic T lymphocyte-associated antigen 4 antibody blockade in previously vaccinated metastatic melanoma and ovarian carcinoma patients. Proc Natl Acad Sci USA 100:4712–4717PubMedCrossRefGoogle Scholar
  22. 22.
    Phan GQ, Yang JC, Sherry RM et al (2003) Cancer regression and autoimmunity induced by cytotoxic T lymphocyte-associated antigen 4 blockade in patients with metastatic melanoma. Proc Natl Acad Sci USA 100:8372–8377PubMedCrossRefGoogle Scholar
  23. 23.
    Lynch T, Neal J, Bondarenko I et al (2010) Phase 2 trial of ipilimumab (IPI) and paclitaxel/carboplatin (P/C) in first-line stage IIIb/IV non-small cell lung cancer (NSCLC). Presented at the 2010 European Society for Clinical Oncology (ESMO) Annual Meeting; October 8–12, 2010, Milan, Italy. Abstract 375PDGoogle Scholar
  24. 24.
    Slovin SF, Beer TM, Higano CS et al (2009) Initial phase II experience of ipilimumab (IPI) alone and in combination with radiotherapy (XRT) in patients with metastatic castration resistant prostate cancer (mCRPC). Presented at the 2009 American Society for Clinical Oncology (ASCO) Meeting; May 29–June 2, 2009, Orlando, FL. Abstract 5138Google Scholar
  25. 25.
    Yuan J, Gnjatic S, Li H et al (2008) CTLA-4 blockade enhances polyfunctional NY-ESO-1 specific T-cell responses in metastatic melanoma patients with clinical benefit. Proc Natl Acad Sci USA 105:20410–20415PubMedCrossRefGoogle Scholar
  26. 26.
    Klein O, Ebert LM, Nicholaou T et al (2009) Melan-A-specific cytotoxic T-cells are associated with tumor regression and autoimmunity following treatment with anti-CTLA-4. Clin Cancer Res 15:2507–2513PubMedCrossRefGoogle Scholar
  27. 27.
    Hodi FS, O’Day SJ, McDermott DF et al (2010) Improved survival with ipilimumab in patients with metastatic melanoma. N Engl J Med 363:711–723PubMedCrossRefGoogle Scholar
  28. 28.
    Fong L, Kwek SS, O’Brien S et al (2009) Potentiating endogenous antitumor immunity to prostate cancer through combination immunotherapy with CTLA4 blockade and GM-CSF. Cancer Res 69:609–615PubMedCrossRefGoogle Scholar
  29. 29.
    Ribas A, Comin-Anduix B, Chmielowski B et al (2009) Dendritic cell vaccination combined with CTLA4 blockade in patients with metastatic melanoma. Clin Cancer Res 15:6267–6276PubMedCrossRefGoogle Scholar
  30. 30.
    Ku GY, Yuan J, Page DB et al (2010) Single-institution experience with ipilimumab in advanced melanoma patients in the compassionate use setting: lymphocyte count after 2 doses correlates with survival. Cancer 116:1767–1775PubMedCrossRefGoogle Scholar
  31. 31.
    Wolchok JD, Hoos A, O’Day S et al (2009) Guidelines for the evaluation of immune therapy activity in solid tumors: immune-related response criteria. Clin Cancer Res 15:7412–7420PubMedCrossRefGoogle Scholar
  32. 32.
    Yuan J, Ku GY, Gallardo HF et al (2009) Safety and immunogenicity of a human and mouse gp100 DNA vaccine in a phase I trial of patients with melanoma. Cancer Immun 9:5PubMedGoogle Scholar
  33. 33.
    Barrow C, Browning J, MacGregor D et al (2006) Tumor antigen expression in melanoma varies according to antigen and stage. Clin Cancer Res 12:764–771PubMedCrossRefGoogle Scholar
  34. 34.
    Carthon BC, Wolchok JD, Yuan J et al (2010) Preoperative CTLA-4 blockade: tolerability and immune monitoring in the setting of a presurgical clinical trial. Clin Cancer Res 16:2861–2871PubMedCrossRefGoogle Scholar
  35. 35.
    Romero P, Zippelius A, Kurth I et al (2007) Four functionally distinct populations of human effector-memory CD8+ T lymphocytes. J Immunol 178:4112–4119PubMedGoogle Scholar
  36. 36.
    Casazza JP, Betts MR, Price DA et al (2006) Acquisition of direct antiviral effector functions by CMV-specific CD4+ T lymphocytes with cellular maturation. J Exp Med 203:2865–2877PubMedCrossRefGoogle Scholar
  37. 37.
    Genescà M, Rourke T, Li J et al (2007) Live attenuated lentivirus infection elicits polyfunctional simian immunodeficiency virus Gag-specific CD8+ T-cells with reduced apoptotic susceptibility in rhesus macaques that control virus replication after challenge with pathogenic SIVmac239. J Immunol 179:4732–4740PubMedGoogle Scholar
  38. 38.
    Duvall MG, Precopio ML, Ambrozak DA et al (2008) Polyfunctional T-cell responses are a hallmark of HIV-2 infection. Eur J Immunol 38:350–363PubMedCrossRefGoogle Scholar
  39. 39.
    Precopio ML, Betts MR, Parrino J et al (2007) Immunization with vaccinia virus induces polyfunctional and phenotypically distinctive CD8+ T-cell responses. J Exp Med 204:1405–1416PubMedCrossRefGoogle Scholar
  40. 40.
    Adams S, O’Neill DW, Nonaka D et al (2008) Immunization of malignant melanoma patients with full-length NY-ESO-1 protein using TLR7 agonist imiquimod as vaccine adjuvant. J Immunol 181:776–784PubMedGoogle Scholar
  41. 41.
    Lin Y, Gallardo HF, Ku GY et al (2009) Optimization and validation of a robust human T-cell culture method for monitoring phenotypic and polyfunctional antigen-specific CD4 and CD8 T-cell responses. Cytotherapy 11:1–11CrossRefGoogle Scholar
  42. 42.
    Vence L, Palucka AK, Fay JW et al (2007) Circulating tumor antigen-specific regulatory T-cells in patients with metastatic melanoma. Proc Natl Acad Sci USA 104:20884–20889PubMedCrossRefGoogle Scholar
  43. 43.
    Thomson TM, Real FX, Murakami S et al (1988) Differentiation antigens of melanocytes and melanoma: analysis of melanosome and cell surface markers of human pigmented cells with monoclonal antibodies. J Invest Dermatol 90:459–466PubMedCrossRefGoogle Scholar
  44. 44.

Copyright information

© Springer-Verlag 2011

Authors and Affiliations

  • Jianda Yuan
    • 1
  • Brian Ginsberg
    • 1
  • David Page
    • 2
  • Yanyun Li
    • 2
  • Teresa Rasalan
    • 1
  • Humilidad F. Gallardo
    • 1
  • Yinyan Xu
    • 1
  • Sylvia Adams
    • 3
  • Nina Bhardwaj
    • 3
  • Klaus Busam
    • 4
  • Lloyd J. Old
    • 5
  • James P. Allison
    • 1
  • Achim Jungbluth
    • 5
  • Jedd D. Wolchok
    • 1
    • 2
    Email author
  1. 1.Immunology ProgramLudwig Center for Cancer Immunotherapy, Sloan-Kettering InstituteNew YorkUSA
  2. 2.Department of MedicineMemorial Sloan-Kettering Cancer CenterNew YorkUSA
  3. 3.Department of MedicineNew York University Cancer Institute, NYU Langone Medical CenterNew YorkUSA
  4. 4.Department of PathologyMemorial Sloan-Kettering Cancer CenterNew YorkUSA
  5. 5.Ludwig Institute for Cancer Research, New York BranchMemorial Sloan-Kettering Cancer CenterNew YorkUSA

Personalised recommendations