Adenovirus MART-1-engineered autologous dendritic cell vaccine for metastatic melanoma - PubMed (original) (raw)

Clinical Trial

. 2008 Apr;31(3):294-309.

doi: 10.1097/CJI.0b013e31816a8910.

Begonya Comin-Anduix, Lazar Vujanovic, Yohan Lee, Vivian B Dissette, Jin-Quan Yang, Hong T Vu, Elizabeth Seja, Denise K Oseguera, Douglas M Potter, John A Glaspy, James S Economou, Antoni Ribas

Affiliations

Clinical Trial

Adenovirus MART-1-engineered autologous dendritic cell vaccine for metastatic melanoma

Lisa H Butterfield et al. J Immunother. 2008 Apr.

Abstract

We performed a phase 1/2 trial testing the safety, toxicity, and immune response of a vaccine consisting of autologous dendritic cells (DCs) transduced with a replication-defective adenovirus (AdV) encoding the full-length melanoma antigen MART-1/Melan-A (MART-1). This vaccine was designed to activate MART-1-specific CD+8 and CD4+ T cells. Metastatic melanoma patients received 3 injections of 10(6) or 10(7) DCs, delivered intradermally. Cell surface phenotype and cytokine production of the DCs used for the vaccines were tested, and indicated intermediate maturity. CD8+ T-cell responses to MART-1 27-35 were assessed by both major histocompatibility complex class I tetramer and interferon (IFN)-gamma enzyme-linked immunosorbent spot (ELISPOT) before, during, and after each vaccine and CD4+ T-cell responses to MART-1 51-73 were followed by IFN-gamma ELISPOT. We also measured antigen response breadth. Determinant spreading from the immunizing antigen MART-1 to other melanoma antigens [gp100, tyrosinase, human melanoma antigen-A3 (MAGE-A3)] was assessed by IFN-gamma ELISPOT. Twenty-three patients were enrolled and 14 patients received all 3 scheduled DC vaccines. Significant CD8+ and/or CD4+ MART-1-specific T-cell responses were observed in 6/11 and 2/4 patients evaluated, respectively, indicating that the E1-deleted adenovirus encoding the cDNA for MART-1/Melan-A (AdVMART1)/DC vaccine activated both helper and killer T cells in vivo. Responses in CD8+ and CD4+ T cells to additional antigens were noted in 2 patients. The AdVMART1-transduced DC vaccine was safe and immunogenic in patients with metastatic melanoma.

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Figures

FIGURE 1

FIGURE 1

AdVMART1/DC vaccine phenotypes. After DC harvest, cells were stained to analyze cell surface phenotype. DCs were gated on (as large, granular lymphocytes) by forward and side scatter, and the MFI of each marker was determined. Examples of DC vaccines from three different patients, B2, B6 and B13, are shown. DC indicates dendritic cell; MFI, mean fluorescence intensity.

FIGURE 2

FIGURE 2

Vaccine DC cytokine production. Cell-free culture supernatants were saved from each DC vaccine culture and tested for the presence of cytokines and chemokines. Results are shown as pg/mL of cytokine. A, Multiple vaccine preparations from several patients were tested by ELISA for IL-15, IL-1β, IL-12p70, TNF-α, IL-10, and IL-6. B, Batched DC vaccine preparation supernatants were tested by Luminex multiplex analysis for IL-12p70, TNF-α, IL-10, IL-6, IFN-α, IFN-γ, and chemokines MIG, IP-10, IL-8, and MCP-1. DC indicates dendritic cell; IFN, interferon; IP, human interferon-inducible protein; IL, interleukin; MCP-1, monocyte chemotactic protein-1; MIG, monokine induced by IFN-γ.

FIGURE 3

FIGURE 3

Examples of IFN-γ ELISPOT responses from CD8+ and CD4+ T cells in 4 patients. Thawed PBMCs from each time point were purified for CD8+ and CD4+ T cells, and these cells were plated overnight with peptide-pulsed antigen presenting cells and controls overnight, in IFN-γ coated plates. Net MART-127-35 or MART-151-73–specific responses per million cells plated (after subtraction of “no peptide” controls) are shown. A, CD8 and CD4 T-cell responses from the first patient vaccinated, A1, are shown. These responses met the statistical criteria for positive responses. B, CD8 and CD4 T-cell responses from patient, B6, are shown. These responses did not meet the statistical criteria for positive responses. C, CD8 T-cell responses from the 2 HLA-DR4-negative patients vaccinated, B3 and B4, are shown. These responses did not meet the statistical criteria for positive responses. ELISPOT indicates enzyme-linked immunosorbent spot; HLA, human leukocyte antigen; IFN, interferon; MART-1, melanoma antigen MART-1/Melan-A; PBMC, peripheral blood mononuclear cells.

FIGURE 4

FIGURE 4

MHC tetramer analysis of CD8+ T-cell frequencies. Examples of 4 different patient MHC tetramer responses are shown. Data are presented as the frequency of tetramer-positive cells among the CD8+ T cells (after gating on live lymphocytes). The frequencies are shown over each available time point tested, from baseline (day 0) to day +56 or day +112. The statistically significant responses shown are for A1 and B6. B4 had a high preexisting level of tetramer-positive cells. B13 had no increase. MHC indicates major histo-compatibility complex.

FIGURE 5

FIGURE 5

ELISPOT analysis of determinant spreading. IFN-γ ELISPOT frequencies of CD8+ and CD4+ T cells specific for MART-1, gp100, tyrosinase, and MAGE-A3–derived peptides for 5 of the 9 tested patients are shown. Solid black and gray bars show the vaccine encoded MART-1 CD8+ and CD4+ responses, respectively. Striped bars show commonly expressed antigens not present in the vaccines. Arrows show any detected increases from baseline to postvaccine time points in immunity to nonvaccine encoded antigens. B2 increased the CD8+ T-cell response to tyrosinase, B4 had a slight increase in CD8+ immune response to gp100, and B6 increased CD8+ immunity to gp100, activated newly detectable immunity to MAGE-A3 as well as showed increased CD4+ T-cell response to gp100. A1 and B1 had detectable responses to MART-1 and other antigens, but these frequencies decreased during the enrollment period. Other tested patients not graphed are described in Tables 3 and 4. ELISPOT indicates enzyme-linked immunosorbent spot; IFN, interferon; MAGE, melanoma antigen; MART-1, MART-1/Melan-A.

FIGURE 6

FIGURE 6

Phenotypic analysis of circulating NK cells. The CD8−/CD4− lymphocytes thawed in batches from several patients were stained for the CD56hi/CD16neg (regulatory or cytokine-producing) and CD56dim/CD16hi (cytotoxic) NK subset markers, and also for frequency changes, activation marker (CD25, CD69), and intracellular granzyme B content changes. The MFI for each of the NK subsets is shown, over time, from 2 patients. A, NK phenotype results for patient B3. B, NK phenotype results for patient B6. MFI indicates mean fluorescence intensity; NK, natural killer.

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