Tumor Progression Can Occur despite the Induction of Very High Levels of Self/Tumor Antigen-Specific CD8+ T Cells in Patients with Melanoma

Steven A. Rosenberg, Richard M. Sherry, Kathleen E. Morton, William J. Scharfman, James C. Yang, Suzanne L. Topalian, Richard E. Royal, Udai Kammula, Nicholas P. Restifo, Marybeth S. Hughes, Douglas Schwartzentruber, David M. Berman, Susan L. Schwarz, Lien T. Ngo, Sharon A. Mavroukakis, Donald E. White and Seth M. Steinberg
J Immunol November 1, 2005, 175 (9) 6169-6176; DOI: https://doi.org/10.4049/jimmunol.175.9.6169

Article Figures & Data

Figures

  • FIGURE 1.
    FIGURE 1.

    Evidence for the generation of high self/tumor Ag-specific T cells using multiple assays. Results of the ELISPOT assay (upper), the tetramer assay (middle), and the in vitro sensitization boost assay (lower) are shown. Each dot represents results from an individual patient. Lymphocytes were obtained by apheresis before all immunizations and 3 wk following each of the first four courses of immunization as well as 1 year after the last administration of peptide. As noted in the text, increasing immunization was seen throughout three courses but little advantage was seen with a fourth course of immunization. In addition, Arm 3 appeared to be more effective in generating precursor T cells compared with Arms 1 and 2.

  • FIGURE 2.
    FIGURE 2.

    The frequency of patients with evidence of positive immunization in each of the three arms of the protocol. Arm 1 (qw), Arm 2 (q3w), Arm 3 (qdx4). Positive in the ELISPOT assay was defined as ≥10 spots per 105 PBMC and more than or equal to twice background (upper). Positive in the tetramer assay was defined as ≥0.5% of all CD8+ cells (middle). A positive assay was defined as IFN-γ release by postvaccination PBMC >100 pg/ml during incubation with the native peptide and more than two times the IFN-γ released by prevaccination PBMC with native peptide.

  • FIGURE 3.
    FIGURE 3.

    Correlation of assays. The correlation of the ELISPOT and tetramer assays (upper), of the in vitro sensitization and tetramer assays (middle), and of the in vitro sensitization and ELISPOT assays (lower) are shown taking all patients after three and four courses. Each dot represents an individual patient tested with each of these assays. In general, correlations were moderately strong (see Results).

  • FIGURE 4.
    FIGURE 4.

    Survival curves. Actuarial curves of disease-free survival of patients in the three eligibility groups (left) and in the three (right) immunization arms of the protocol.

Tables

  • Table I.

    HLA-A2 adjuvant protocol

    ScheduleArm1 (qw)Arm2 (q3w)Arm3 (qdx4 q3w)
    Number of patients (%)
    31 (100)31 (100)33 (100)
    Sex
     Male17 (55)16 (52)24 (73)
     Female14 (45)15 (48)9 (27)
    Age
     21–301 (3)2 (6)0 (0)
     31–406 (19)3 (10)4 (12)
     41–5011 (35)8 (26)10 (30)
     51–609 (29)12 (39)8 (24)
     61–704 (13)5 (16)10 (30)
     Over 700 (0)1 (3)1 (3)
    Eligibility criteria
     Resected primary6 (19)14 (45)5 (15)
     Positive nodes19 (61)13 (42)23 (70)
     Resected mets6 (19)4 (13)5 (15)
    Number of positive lymph nodes
     012 (39)18 (5)10 (30)
     1–216 (52)9 (29)10 (30)
     3–42 (6)4 (13)5 (15)
     5–60 (0)0 (0)4 (12)
     7–80 (0)0 (0)3 (9)
     9–100 (0)0 (0)1 (3)
     >101 (5)0 (0)0 (0)
  • Table II.

    Results of ELISPOT and tetramer assays before immunization and after three or four courses

    Arm1Arm2Arm3Total
    Pre (n = 31)Post 3–4 (n = 22)Pre (n = 28)Post 3–4 (n = 25)Pre (n = 31)Post 3–4 (n = 16)Pre (n = 90)Post 3–4 (n = 63)
    ELISPOT assay Spots/105 PBMC
     <1027 (87%)4 (18%)23 (82%)7 (28%)30 (97%)1 (6%)80 (89%)12 (19%)
     10–1004 (13%)7 (32%)5 (18%)10 (40%)1 (3%)1 (6%)10 (11%)18 (29%)
     10–1,000010 (45%)08 (32%)014 (88%)032 (51%)
     >1,00001 (5%)000001 (2%)
    Tetramer assay % CD8+ cells
     <0.1%30 (97%)6 (27%)28 (97%)9 (36%)28 (90%)2 (12%)86 (95%)17 (27%)
     0.1–1%1 (3%)3 (14%)1 (3%)4 (16%)2 (7%)04 (4%)7 (9%)
     1.1–10%08 (36%)09 (36%)1 (3%)11 (69%)1 (1%)28 (44%)
     >10%05 (23%)03 (12%)03 (19%)011 (17%)
  • Table III.

    Recognition of peptide and tumor before and after immunization: ELISPOT assay

    PatientPreAfter Three or Four Courses
    CIRA2 + peptideMelanomaCIRA2 + peptideMelanoma
    02802095266248889380280209526624888938
    (A2+)(A2+)(A2−)(A2−)(A2+)(A2+)(A2−)(A2−)
    (Number spots/105 PBMC)
    1242428710610108 290 a 414157
    2108117115173 321 222958
    3261726810512518 1160 432 227 1593
    42117257915275261010 266 22030016163
    51278937388 509 3404
    6333047175595224 489 8937744
    78462792112418241315 577 28411711150
    82492314205853 507 9734069
    93522175306820 710 4217024
    1016151112141656748 636 105361967
    115559111241510 782 299 334 3015
    12121118111418111113 725 88143117
    1313101313131842920 1486 219 234 5038
    14555817201123 377 56523116
    151399682010134 1474 613 574 2524
    16201620202132353315 2082 627 773 4339
    1789101131201863 831 113 165 2823

    • a Values twice both controls are underlined.

  • Table IV.

    Patients with recurrence after ≥2 courses of immunization

    CourseIntensity/% Cells PositiveELISPOT (per 105 PBMC)Tetramer (% of CD8+)
    gp100MART1TyrosinaseMHC1
    Arm1(qw)43/>50%0/00/00/0422.01
    42/>50%2/>50%0/01/<5%122.21
    20/00/02/>50%3/>50%40.29
    23/>50%1/>50%3/>50%1/>50%00
    22/>50%3/>50%3/>50%1/<5%00
    41/5–50%3/>50%3/>50%0/060616.70
    43/>50%3/>50%3/>50%2/>50%1600
    43/>50%3/>50%2>50%2/5–50%30315.2
    Arm2/(q3w)40/00/00/03/>50%2452.56
    41/5–50%0/03/>50%2/>50%240.51
    40/01/>50%0/03/>50%22816.85
    43/>50%3/>50%3/>50%3/>50%90.47
    40/03/>50%3/>50%3/>50%00
    Arm3(qdx4)43/5–50%1/>50%3>50%3>50%1140.5
    42/>50%n.d.0/00/01513.89
    20/00/00/03/<5%160.01
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