Limits...
Antitumor monoclonal antibodies enhance cross-presentation ofcCellular antigens and the generation of myeloma-specific killer T cells by dendritic cells.

Dhodapkar KM, Krasovsky J, Williamson B, Dhodapkar MV - J. Exp. Med. (2002)

Bottom Line: The mechanism of antitumor effect of monoclonal antibodies (mAbs) is not fully understood.Importantly, mAbs-coated tumor-loaded DCs were consistently superior to DCs loaded with peptides or dying cells for eliciting tumor-specific killer T cells.Cross-presentation was inhibited by pretreatment of DCs with Fc gamma receptor blocking antibodies.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Tumor Immunology and Immunotherapy, The Rockefeller University, New York, NY 10021, USA.

ABSTRACT
The mechanism of antitumor effect of monoclonal antibodies (mAbs) is not fully understood. Here we show that coating myeloma cells with anti-syndecan-1 antibody promotes cross-presentation of cellular antigens by dendritic cells (DCs) to autologous T cells from healthy donors. The tumor cells treated with anti-syndecan-1 or isotype-matched control antibody were fed to HLA-mismatched monocyte-derived immature DCs. Tumor cell-loaded mature DCs induced a strong CD8(+) T cell response that was specific for the cancer-testis (C-T) antigens expressed in the tumor. The CD8(+) T cells killed peptide-pulsed targets, as well as myeloma tumor cells. Importantly, mAbs-coated tumor-loaded DCs were consistently superior to DCs loaded with peptides or dying cells for eliciting tumor-specific killer T cells. This enhanced cross-presentation was not due to enhanced tumor cell uptake or to DC maturation. When mixtures of NY-Eso-1-positive and -negative myeloma cells were captured by DCs, the anti-syndecan-1 antibody had to be on the NY-Eso-1-positive cells to elicit NY-Eso-1-specific response. Cross-presentation was inhibited by pretreatment of DCs with Fc gamma receptor blocking antibodies. Targeting of mAb-coated tumors to DCs may contribute to the efficacy of tumor-reactive mAb and offers a new strategy for immunotherapy.

Show MeSH

Related in: MedlinePlus

Effects of antitumor antibodies on cross presentation of cellular antigens from dying tumor cells. (A) HLA A2.1−ve myeloma cells were killed either by γ-irradiation (apoptosis) or freeze thaw (necrosis), and either left untreated, or coated with anti–syndecan-1 antibody (mAb) or isotype. HLA A2.1+ DCs were fed with tumor cells, matured with cytokine cocktail and then used to stimulate autologous T cells. After two stimulations, generation of HLA A*0201-restricted MAGE-3 or NY-Eso-1 peptide-specific T cells was quantified in a 16 h ELISPOT using autologous peptide-pulsed mature DCs as APCs. (B) T cells from experiment in A were tested for killing of T2 cells pulsed with HLA A*0201-restricted MAGE3/NY-Eso-1 peptide or HLA A*0201-positive (U266) or -negative (ark) myeloma cells, at a E/T ratio of 20:1, with a standard 5 h 51Cr release assay.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2196013&req=5

fig5: Effects of antitumor antibodies on cross presentation of cellular antigens from dying tumor cells. (A) HLA A2.1−ve myeloma cells were killed either by γ-irradiation (apoptosis) or freeze thaw (necrosis), and either left untreated, or coated with anti–syndecan-1 antibody (mAb) or isotype. HLA A2.1+ DCs were fed with tumor cells, matured with cytokine cocktail and then used to stimulate autologous T cells. After two stimulations, generation of HLA A*0201-restricted MAGE-3 or NY-Eso-1 peptide-specific T cells was quantified in a 16 h ELISPOT using autologous peptide-pulsed mature DCs as APCs. (B) T cells from experiment in A were tested for killing of T2 cells pulsed with HLA A*0201-restricted MAGE3/NY-Eso-1 peptide or HLA A*0201-positive (U266) or -negative (ark) myeloma cells, at a E/T ratio of 20:1, with a standard 5 h 51Cr release assay.

Mentions: In the experiments described above, we cocultured DCs with live antibody-coated tumor cells after low dose (3 Gy) irradiation. However, several studies have shown that DCs can cross-present antigen from apoptotic or necrotic tumor cells. Therefore, we next examined if antibody coating also enhanced cross-presentation of antigen from dying cells. We first confirmed the expression of syndecan-1 on tumor cells rendered apoptotic after γ-irradiation, by double staining for Annexin V and CD138 (data not shown). Next, we fed DCs with apoptotic or necrotic tumor cells that had been stained with anti–syndecan-1 antibody, isotype control, or left unstained. These DCs were then matured using an inflammatory cytokine cocktail and used as APCs to stimulate autologous T cells. The DCs that were fed with antibody-coated apoptotic tumor cells expanded antigen-specific T cells more efficiently relative to DCs fed with apoptotic cells alone (Fig. 5 A). No enhancement of presentation of necrotic cells was observed by antibody-mediated targeting, which is likely due to the lack of effective binding of the antibody to necrotic fragments. T cells expanded using DCs loaded with antibody-coated apoptotic cells were also superior at killing peptide-pulsed or tumor cell targets (Fig. 5 B). Thus antibody coating of tumor cells also leads to enhanced cross-presentation of dying cells by DCs.


Antitumor monoclonal antibodies enhance cross-presentation ofcCellular antigens and the generation of myeloma-specific killer T cells by dendritic cells.

Dhodapkar KM, Krasovsky J, Williamson B, Dhodapkar MV - J. Exp. Med. (2002)

Effects of antitumor antibodies on cross presentation of cellular antigens from dying tumor cells. (A) HLA A2.1−ve myeloma cells were killed either by γ-irradiation (apoptosis) or freeze thaw (necrosis), and either left untreated, or coated with anti–syndecan-1 antibody (mAb) or isotype. HLA A2.1+ DCs were fed with tumor cells, matured with cytokine cocktail and then used to stimulate autologous T cells. After two stimulations, generation of HLA A*0201-restricted MAGE-3 or NY-Eso-1 peptide-specific T cells was quantified in a 16 h ELISPOT using autologous peptide-pulsed mature DCs as APCs. (B) T cells from experiment in A were tested for killing of T2 cells pulsed with HLA A*0201-restricted MAGE3/NY-Eso-1 peptide or HLA A*0201-positive (U266) or -negative (ark) myeloma cells, at a E/T ratio of 20:1, with a standard 5 h 51Cr release assay.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2196013&req=5

fig5: Effects of antitumor antibodies on cross presentation of cellular antigens from dying tumor cells. (A) HLA A2.1−ve myeloma cells were killed either by γ-irradiation (apoptosis) or freeze thaw (necrosis), and either left untreated, or coated with anti–syndecan-1 antibody (mAb) or isotype. HLA A2.1+ DCs were fed with tumor cells, matured with cytokine cocktail and then used to stimulate autologous T cells. After two stimulations, generation of HLA A*0201-restricted MAGE-3 or NY-Eso-1 peptide-specific T cells was quantified in a 16 h ELISPOT using autologous peptide-pulsed mature DCs as APCs. (B) T cells from experiment in A were tested for killing of T2 cells pulsed with HLA A*0201-restricted MAGE3/NY-Eso-1 peptide or HLA A*0201-positive (U266) or -negative (ark) myeloma cells, at a E/T ratio of 20:1, with a standard 5 h 51Cr release assay.
Mentions: In the experiments described above, we cocultured DCs with live antibody-coated tumor cells after low dose (3 Gy) irradiation. However, several studies have shown that DCs can cross-present antigen from apoptotic or necrotic tumor cells. Therefore, we next examined if antibody coating also enhanced cross-presentation of antigen from dying cells. We first confirmed the expression of syndecan-1 on tumor cells rendered apoptotic after γ-irradiation, by double staining for Annexin V and CD138 (data not shown). Next, we fed DCs with apoptotic or necrotic tumor cells that had been stained with anti–syndecan-1 antibody, isotype control, or left unstained. These DCs were then matured using an inflammatory cytokine cocktail and used as APCs to stimulate autologous T cells. The DCs that were fed with antibody-coated apoptotic tumor cells expanded antigen-specific T cells more efficiently relative to DCs fed with apoptotic cells alone (Fig. 5 A). No enhancement of presentation of necrotic cells was observed by antibody-mediated targeting, which is likely due to the lack of effective binding of the antibody to necrotic fragments. T cells expanded using DCs loaded with antibody-coated apoptotic cells were also superior at killing peptide-pulsed or tumor cell targets (Fig. 5 B). Thus antibody coating of tumor cells also leads to enhanced cross-presentation of dying cells by DCs.

Bottom Line: The mechanism of antitumor effect of monoclonal antibodies (mAbs) is not fully understood.Importantly, mAbs-coated tumor-loaded DCs were consistently superior to DCs loaded with peptides or dying cells for eliciting tumor-specific killer T cells.Cross-presentation was inhibited by pretreatment of DCs with Fc gamma receptor blocking antibodies.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Tumor Immunology and Immunotherapy, The Rockefeller University, New York, NY 10021, USA.

ABSTRACT
The mechanism of antitumor effect of monoclonal antibodies (mAbs) is not fully understood. Here we show that coating myeloma cells with anti-syndecan-1 antibody promotes cross-presentation of cellular antigens by dendritic cells (DCs) to autologous T cells from healthy donors. The tumor cells treated with anti-syndecan-1 or isotype-matched control antibody were fed to HLA-mismatched monocyte-derived immature DCs. Tumor cell-loaded mature DCs induced a strong CD8(+) T cell response that was specific for the cancer-testis (C-T) antigens expressed in the tumor. The CD8(+) T cells killed peptide-pulsed targets, as well as myeloma tumor cells. Importantly, mAbs-coated tumor-loaded DCs were consistently superior to DCs loaded with peptides or dying cells for eliciting tumor-specific killer T cells. This enhanced cross-presentation was not due to enhanced tumor cell uptake or to DC maturation. When mixtures of NY-Eso-1-positive and -negative myeloma cells were captured by DCs, the anti-syndecan-1 antibody had to be on the NY-Eso-1-positive cells to elicit NY-Eso-1-specific response. Cross-presentation was inhibited by pretreatment of DCs with Fc gamma receptor blocking antibodies. Targeting of mAb-coated tumors to DCs may contribute to the efficacy of tumor-reactive mAb and offers a new strategy for immunotherapy.

Show MeSH
Related in: MedlinePlus