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. 2004 Sep 15;64(18):6783-90.
doi: 10.1158/0008-5472.CAN-04-1621.

Dendritic cells strongly boost the antitumor activity of adoptively transferred T cells in vivo

Yanyan Lou  1 , Gang Wang, Gregory Lizée, Grace J Kim, Steven E Finkelstein, Chiguang Feng, Nicholas P Restifo, Patrick Hwu
Affiliations

Dendritic cells strongly boost the antitumor activity of adoptively transferred T cells in vivo

Yanyan Lou et al. Cancer Res. .

Abstract

Dendritic cells (DCs) have been well characterized for their ability to initiate cell-mediated immune responses by stimulating naive T cells. However, the use of DCs to stimulate antigen-activated T cells in vivo has not been investigated. In this study, we determined whether DC vaccination could improve the efficacy of activated, adoptively transferred T cells to induce an enhanced antitumor immune response. Mice bearing B16 melanoma tumors expressing the gp100 tumor antigen were treated with cultured, activated T cells transgenic for a T-cell receptor specifically recognizing gp100, with or without concurrent peptide-pulsed DC vaccination. In this model, antigen-specific DC vaccination induced cytokine production, enhanced proliferation, and increased tumor infiltration of adoptively transferred T cells. Furthermore, the combination of DC vaccination and adoptive T-cell transfer led to a more robust antitumor response than the use of each treatment individually. Collectively, these findings illuminate a new potential application for DCs in the in vivo stimulation of adoptively transferred T cells and may be a useful approach for the immunotherapy of cancer.

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Figures

Fig. 1
Fig. 1
Cytokine production by pmel-1 T cells following antigen-specific DC vaccination in vivo. Mice were immunized intravenously with DCs pulsed with either the melanoma peptide antigen hgp10025–33 or the irrelevant influenza nucleoprotein-derived peptide NP366–374 plus IL-2 administration immediately following adoptive transfer of cultured pmel-1 T lymphocytes. Three days later, pooled lymphocytes from treated mice (four mice/group) were isolated from the indicated lymphoid organs, and pmel-1 T cells were evaluated for intracellular production of IFN-γ in the absence of additional in vitro stimulation. PLN, peripheral lymph nodes; MLN, mesenteric lymph nodes.
Fig. 2
Fig. 2
Enhanced proliferation of pmel-1 T cells following antigen-specific DC vaccination in vivo. In vitro stimulated pmel-1 T cells were labeled with CFSE, washed, and adoptively transferred into B16 tumor-bearing recipient mice, followed immediately by immunization with DCs pulsed with either hgp10025–33 or NP366–374 peptides plus IL-2 administration. After 5 days, pooled lymphocytes from treated mice (four mice/group) were recovered from the indicated lymphoid organs and (A) analyzed for proliferation by flow cytometry or (B) quantitated for absolute number of pmel-1 cells present within each lymphoid compartment. PLN, peripheral lymph nodes; MLN, mesenteric lymph nodes.
Fig. 3
Fig. 3
Enhanced antitumor activity by combining DC vaccination with adoptive transfer of pmel-1 T cells and IL-2 injection. C57BL/6 mice were subcutaneously inoculated with 5 × 105 B16 tumor cells. Seven days later, tumor-bearing mice were intravenously injected with 6 × 106 cultured pmel-1 T cells, followed immediately by intravenous vaccination with DCs pulsed with either the hgp10025–33 or NP366–374 peptides. IL-2 was administrated twice daily for a total of six doses. (A) Tumor growth and (B) mouse survival rate were monitored for up to 5 weeks following treatment. *P = 0.006; **P = 0.003 for DC/hgp100 versus DC/NP. Results shown are representative of three independent experiments.
Fig. 4
Fig. 4
Sequential DC immunization increases antitumor response, mouse survival, and pmel-1 T-cell persistence. Eight-day B16 tumor-bearing mice were intravenously injected with 4 × 106 pmel-1 T cells, followed by vaccination with DCs pulsed with either the hgp10025–33 or NP366–374 peptides. Indicated groups of mice received booster DC immunizations at 6-day intervals with peptide-pulsed DCs for one, two, or three total vaccinations. IL-2 was administered intraperitoneally immediately following each vacci-nation. (A) Tumor growth and (B) survival rate of mice were monitored to compare vaccination regimens. (C) Peripheral blood was analyzed by flow cytometry for percentages of pmel-1 cells (Vβ13+Thy1.1+) in total PBLs at the indicated time points. *P = 0.014; **P = 0.003; ***P = 0.021 for one versus three sequential DC/hgp100 vaccinations. Data are representative of two independent experiments with a total of five mice per group. (◆) No treatment; pmel-1 + (■) DC/hgp100 + IL-2 (1), (□) DC/NP + IL-2 (1), (▲) DC/hgp100 + IL-2 (2), (△) DC/NP + IL-2 (2), (●) DC/hgp100 + IL-2 (3), (○) DC/NP + IL-2 (3).
Fig. 5
Fig. 5
Irradiation before adoptive pmel-1 T cells transfer and DC immunization enhances antitumor response. Seven-day subcutaneous B16 tumor-bearing mice were irradiated with 500 rad, followed 1 day later by adoptive transfer of pmel-1 T cells and immunization with DCs bearing either of the hgp10025–33 or NP366–374 peptides plus IL-2 administration. Tumor growth was monitored and compared with that of nonirradiated mice. *P = 0.009 for irradiated versus nonirradiated hosts receiving DC/hgp100 immunization. Data are representative of two independent experiments with a total of five mice per group. Non-irradiation: (◆) no treatment, (■) DC/NP + IL-2, (●) DC/hgp100 + IL-2. Irradiation: (◇) no treatment, (□) DC/NP + IL-2, (○) DC/hgp100 + IL-2.
Fig. 6
Fig. 6
Sequential DC vaccination plus irradiation in combination with adoptive transfer of pmel-1 T cells provides an optimal antitumor response. Seven-day subcutaneous B16 tumor-bearing mice were irradiated with 500 rad, followed 1 day later by adoptive transfer of pmel-1 T cells and immunization with DCs bearing either of the hgp10025–33 or NP366–374 peptides plus IL-2 administration. Thereafter, indicated mice received one or two booster DC vaccinations at 6-day intervals. (A) Tumor growth of mice was monitored following treatment. (B) Peripheral blood was analyzed by flow cytometry for percentages of pmel-1 cells (Vβ13+Thy1.1+) in total PBLs at the indicated time points. *P = 0.013; **P = 0.021 for one versus three sequential DC/hgp100 vaccinations. Data are representative of two independent experiments with a total of five mice per group. (◆) No treatment; pmel-1 + (■) DC/hgp100 + IL-2 (1), (□) DC/NP + IL-2 (1), (▲) DC/hgp100 + IL-2 (2), (△) DC/NP + IL-2 (2), (●) DC/hgp100 + IL-2 (3), (○) DC/NP + IL-2 (3).
Fig. 7
Fig. 7
Antigen-specific DC vaccination enhances the pmel-1 T-cell infiltration into tumors. Seven-day B16 tumor-bearing mice were infused intravenously with pmel-1 T cells, followed by immunization with DCs pulsed with either the hgp10025–33 or NP366–374 peptide plus IL-2 administration. Five days later, blood, spleen, and tumors were harvested, and lymphocytes were isolated and analyzed by flow cytometry. (A) percentage of total tumor-infiltrating lymphocytes that are pmel-1 (Vβ13+Thy1.1+) T cells. (B) number of total lymphocytes and number of pmel-1 cells per milligram of tumor. For (B) each data point represents one individual mouse of four per group. (C and D) expression of activation/homing markers CD62L and CD44 on adoptively transferred pmel-1 T cells. Cells were stained with mAbs against Thy1.1 and Vβ13 to identify adoptively transferred pmel-1 cells and then stained with CD62L and CD44 mAbs, respectively. Histograms are gated on pmel-1 cells (Vβ13+Thy1.1+). Specific staining is depicted as a filled histogram with isotype control as an open histogram.
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