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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.

van Elsas A, Hurwitz AA, Allison JP - J. Exp. Med. (1999)

Bottom Line: We examined the effectiveness of cytotoxic T lymphocyte-associated antigen 4 (CTLA-4) blockade, alone or in combination with a granulocyte/macrophage colony-stimulating factor (GM-CSF)-expressing tumor cell vaccine, on rejection of the highly tumorigenic, poorly immunogenic murine melanoma B16-BL6.The same treatment regimen was found to be therapeutically effective against outgrowth of preestablished B16-F10 lung metastases, inducing long-term survival.Depigmentation was found to occur in CD4-depleted mice, strongly suggesting that the effect was mediated by CTLs.

View Article: PubMed Central - PubMed

Affiliation: Howard Hughes Medical Institute, Cancer Research Laboratory, Department of Molecular and Cellular Biology, University of California, Berkeley 94720-3200, USA.

ABSTRACT
We examined the effectiveness of cytotoxic T lymphocyte-associated antigen 4 (CTLA-4) blockade, alone or in combination with a granulocyte/macrophage colony-stimulating factor (GM-CSF)-expressing tumor cell vaccine, on rejection of the highly tumorigenic, poorly immunogenic murine melanoma B16-BL6. Recently established tumors could be eradicated in 80% (68/85) of the cases using combination treatment, whereas each treatment by itself showed little or no effect. Tumor rejection was dependent on CD8(+) and NK1.1(+) cells but occurred irrespective of the presence of CD4(+) T cells. Mice surviving a primary challenge rejected a secondary challenge with B16-BL6 or the parental B16-F0 line. The same treatment regimen was found to be therapeutically effective against outgrowth of preestablished B16-F10 lung metastases, inducing long-term survival. Of all mice surviving B16-BL6 or B16-F10 tumors after combination treatment, 56% (38/68) developed depigmentation, starting at the site of vaccination or challenge and in most cases progressing to distant locations. Depigmentation was found to occur in CD4-depleted mice, strongly suggesting that the effect was mediated by CTLs. This study shows that CTLA-4 blockade provides a powerful tool to enhance T cell activation and memory against a poorly immunogenic spontaneous murine tumor and that this may involve recruitment of autoreactive T cells.

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Anti–CTLA-4 enhances IFN-γ production by B16-specific T cells induced in vivo. Mice (four per group) were vaccinated with irradiated BL6/g (106 per mouse) and cotreated with control hamster IgG (A) or anti–CTLA-4 (B). After 4 wk, mice were challenged with 2 × 104 B16-BL6, and 10 d later, splenocytes were pooled and restimulated in vitro using B16-BL6/B7.1 (open bars) or a mixture of B16-F10 and DC2.4 dendritic cells (filled bars). On day 8, cultures were tested for tumor-specific IFN-γ release as described in Materials and Methods. Targets included B16 sublines -F0, -F10, and -BL6, as well as unrelated H-2b tumors EL4 and MC38.
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Figure 3: Anti–CTLA-4 enhances IFN-γ production by B16-specific T cells induced in vivo. Mice (four per group) were vaccinated with irradiated BL6/g (106 per mouse) and cotreated with control hamster IgG (A) or anti–CTLA-4 (B). After 4 wk, mice were challenged with 2 × 104 B16-BL6, and 10 d later, splenocytes were pooled and restimulated in vitro using B16-BL6/B7.1 (open bars) or a mixture of B16-F10 and DC2.4 dendritic cells (filled bars). On day 8, cultures were tested for tumor-specific IFN-γ release as described in Materials and Methods. Targets included B16 sublines -F0, -F10, and -BL6, as well as unrelated H-2b tumors EL4 and MC38.

Mentions: To determine if tumor-reactive T cells were induced by the combination therapy, mice were immunized with BL6/g plus anti–CTLA-4 or control IgG and challenged with B16-BL6 after 4 wk. 10 d after challenge, spleens from four mice in each group were pooled and restimulated with B16-BL6/B7.1 or a mixture of B16-F10 and the dendritic cell line DC2.4 19. After one round of restimulation in vitro, specific IFN-γ release was tested using different variants of B16 and two unrelated tumor cell lines expressing the H-2b haplotype, the thymoma EL4 and the colorectal carcinoma MC38. As shown in Fig. 3T cells from mice vaccinated with BL6/g in the presence of control hamster IgG produced very low levels of IFN-γ in this assay. T cells from mice treated with anti–CTLA-4 in vivo had greatly enhanced B16-specific IFN-γ secretion. These results indicate that CTLA-4 blockade during vaccination with BL6/g specifically enhances reactivity toward an antigen (or antigens) expressed by B16 and its variants. In addition, all splenocyte cultures established from mice that were long-term (3–10 mo) survivors after combination treatment were found to specifically react with B16 and its variants, as tested by IFN-γ release after one round of restimulation in vitro (data not shown). Successful rejection of B16-BL6 coincides with the generation of tumor-specific T cell activity.


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.

van Elsas A, Hurwitz AA, Allison JP - J. Exp. Med. (1999)

Anti–CTLA-4 enhances IFN-γ production by B16-specific T cells induced in vivo. Mice (four per group) were vaccinated with irradiated BL6/g (106 per mouse) and cotreated with control hamster IgG (A) or anti–CTLA-4 (B). After 4 wk, mice were challenged with 2 × 104 B16-BL6, and 10 d later, splenocytes were pooled and restimulated in vitro using B16-BL6/B7.1 (open bars) or a mixture of B16-F10 and DC2.4 dendritic cells (filled bars). On day 8, cultures were tested for tumor-specific IFN-γ release as described in Materials and Methods. Targets included B16 sublines -F0, -F10, and -BL6, as well as unrelated H-2b tumors EL4 and MC38.
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Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2195583&req=5

Figure 3: Anti–CTLA-4 enhances IFN-γ production by B16-specific T cells induced in vivo. Mice (four per group) were vaccinated with irradiated BL6/g (106 per mouse) and cotreated with control hamster IgG (A) or anti–CTLA-4 (B). After 4 wk, mice were challenged with 2 × 104 B16-BL6, and 10 d later, splenocytes were pooled and restimulated in vitro using B16-BL6/B7.1 (open bars) or a mixture of B16-F10 and DC2.4 dendritic cells (filled bars). On day 8, cultures were tested for tumor-specific IFN-γ release as described in Materials and Methods. Targets included B16 sublines -F0, -F10, and -BL6, as well as unrelated H-2b tumors EL4 and MC38.
Mentions: To determine if tumor-reactive T cells were induced by the combination therapy, mice were immunized with BL6/g plus anti–CTLA-4 or control IgG and challenged with B16-BL6 after 4 wk. 10 d after challenge, spleens from four mice in each group were pooled and restimulated with B16-BL6/B7.1 or a mixture of B16-F10 and the dendritic cell line DC2.4 19. After one round of restimulation in vitro, specific IFN-γ release was tested using different variants of B16 and two unrelated tumor cell lines expressing the H-2b haplotype, the thymoma EL4 and the colorectal carcinoma MC38. As shown in Fig. 3T cells from mice vaccinated with BL6/g in the presence of control hamster IgG produced very low levels of IFN-γ in this assay. T cells from mice treated with anti–CTLA-4 in vivo had greatly enhanced B16-specific IFN-γ secretion. These results indicate that CTLA-4 blockade during vaccination with BL6/g specifically enhances reactivity toward an antigen (or antigens) expressed by B16 and its variants. In addition, all splenocyte cultures established from mice that were long-term (3–10 mo) survivors after combination treatment were found to specifically react with B16 and its variants, as tested by IFN-γ release after one round of restimulation in vitro (data not shown). Successful rejection of B16-BL6 coincides with the generation of tumor-specific T cell activity.

Bottom Line: We examined the effectiveness of cytotoxic T lymphocyte-associated antigen 4 (CTLA-4) blockade, alone or in combination with a granulocyte/macrophage colony-stimulating factor (GM-CSF)-expressing tumor cell vaccine, on rejection of the highly tumorigenic, poorly immunogenic murine melanoma B16-BL6.The same treatment regimen was found to be therapeutically effective against outgrowth of preestablished B16-F10 lung metastases, inducing long-term survival.Depigmentation was found to occur in CD4-depleted mice, strongly suggesting that the effect was mediated by CTLs.

View Article: PubMed Central - PubMed

Affiliation: Howard Hughes Medical Institute, Cancer Research Laboratory, Department of Molecular and Cellular Biology, University of California, Berkeley 94720-3200, USA.

ABSTRACT
We examined the effectiveness of cytotoxic T lymphocyte-associated antigen 4 (CTLA-4) blockade, alone or in combination with a granulocyte/macrophage colony-stimulating factor (GM-CSF)-expressing tumor cell vaccine, on rejection of the highly tumorigenic, poorly immunogenic murine melanoma B16-BL6. Recently established tumors could be eradicated in 80% (68/85) of the cases using combination treatment, whereas each treatment by itself showed little or no effect. Tumor rejection was dependent on CD8(+) and NK1.1(+) cells but occurred irrespective of the presence of CD4(+) T cells. Mice surviving a primary challenge rejected a secondary challenge with B16-BL6 or the parental B16-F0 line. The same treatment regimen was found to be therapeutically effective against outgrowth of preestablished B16-F10 lung metastases, inducing long-term survival. Of all mice surviving B16-BL6 or B16-F10 tumors after combination treatment, 56% (38/68) developed depigmentation, starting at the site of vaccination or challenge and in most cases progressing to distant locations. Depigmentation was found to occur in CD4-depleted mice, strongly suggesting that the effect was mediated by CTLs. This study shows that CTLA-4 blockade provides a powerful tool to enhance T cell activation and memory against a poorly immunogenic spontaneous murine tumor and that this may involve recruitment of autoreactive T cells.

Show MeSH
Related in: MedlinePlus