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MELOE-1 is a new antigen overexpressed in melanomas and involved in adoptive T cell transfer efficiency.

Godet Y, Moreau-Aubry A, Guilloux Y, Vignard V, Khammari A, Dreno B, Jotereau F, Labarriere N - J. Exp. Med. (2008)

Bottom Line: Remarkably, the structure of meloe was unusual, with multiple short open reading frames (ORFs).The peptide recognized by the CTL clone was encoded by one of these ORFs, called MELOE-1.Using a specific HLA-A2/peptide tetramer, we showed a correlation between the infusion of TILs containing MELOE-1-specific T cells and relapse prevention in HLA-A2 patients.

View Article: PubMed Central - PubMed

Affiliation: Institut National de Santé et de Recherche Médicale, Unité Mixte de Recherche 892, 44093 Nantes, France.

ABSTRACT
A cytotoxic T lymphocyte (CTL) clone was derived from a tumor-infiltrating lymphocyte (TIL) population infused to a melanoma patient who remained relapse free for 10 yr after this adoptive transfer. This clone recognized all melanoma cell lines tested and, to a lower extent, melanocytes, in the context of human histocompatibility leukocyte antigen A2 (HLA-A2), but it did not recognize other tumor cell types. The gene coding for the antigen recognized by this clone was identified by the screening of a melanoma complementary DNA expression library. This antigen is overexpressed in melanomas, compared with other cancer cell lines and healthy tissues, and was thus called melanoma-overexpressed antigen (meloe). Remarkably, the structure of meloe was unusual, with multiple short open reading frames (ORFs). The peptide recognized by the CTL clone was encoded by one of these ORFs, called MELOE-1. Using a specific HLA-A2/peptide tetramer, we showed a correlation between the infusion of TILs containing MELOE-1-specific T cells and relapse prevention in HLA-A2 patients. Indeed, 5 out of 9 patients who did not relapse were infused with TILs that contained MELOE-1-specific T cells, whereas 0 out of the 21 patients who relapsed was infused with such TIL-containing lymphocytes. Overall, our results suggest that this new antigen is involved in immunosurveillance and, thus, represents an attractive target for immunotherapy protocols of melanoma.

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T cell clone selection and characterization. (A) Percentage of TNF-producing T cells and of HLA-A2/Melan-AA27L tetramer–positive T cells in the M170 TIL population in response to the autologous melanoma cell line. 105 TILs and 2 × 105 melanoma cells were incubated for 5 h in the presence of Brefeldin A, stained with HLA-A2/Melan-AA27L tetramer, fixed, and stained with anti-TNF antibody in a permeabilization buffer. 104 T cells were then analyzed by flow cytometry. (B) TNF secretion by the M170.48 T cell clone in response to the autologous melanoma cell line. 104 CTLs were added to 3 × 104 M170 melanoma cells in the presence of blocking antibodies directed against class I, A2, and B/C HLA, diluted to 1:50 (shaded bars), 1:500 (hatched bars), and 1:5,000 (open bars). CTL clone reactivity was assessed by a TNF release assay. (C) TNF response of the M170.48 CTL clone to HLA-A*0201 tumor cell lines. The M6 cell line (HLA-A2 negative) was used as a negative control. (D) IFN-γ response of the M170.48 CTL clone (shaded bars) and of a Melan-A/A2–specific CTL clone (hatched bars) to HLA-A*0201 melanocytes. The M170 cell line was added as a positive control.
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fig1: T cell clone selection and characterization. (A) Percentage of TNF-producing T cells and of HLA-A2/Melan-AA27L tetramer–positive T cells in the M170 TIL population in response to the autologous melanoma cell line. 105 TILs and 2 × 105 melanoma cells were incubated for 5 h in the presence of Brefeldin A, stained with HLA-A2/Melan-AA27L tetramer, fixed, and stained with anti-TNF antibody in a permeabilization buffer. 104 T cells were then analyzed by flow cytometry. (B) TNF secretion by the M170.48 T cell clone in response to the autologous melanoma cell line. 104 CTLs were added to 3 × 104 M170 melanoma cells in the presence of blocking antibodies directed against class I, A2, and B/C HLA, diluted to 1:50 (shaded bars), 1:500 (hatched bars), and 1:5,000 (open bars). CTL clone reactivity was assessed by a TNF release assay. (C) TNF response of the M170.48 CTL clone to HLA-A*0201 tumor cell lines. The M6 cell line (HLA-A2 negative) was used as a negative control. (D) IFN-γ response of the M170.48 CTL clone (shaded bars) and of a Melan-A/A2–specific CTL clone (hatched bars) to HLA-A*0201 melanocytes. The M170 cell line was added as a positive control.

Mentions: The M170 TIL population contained 16% of melanoma-reactive lymphocytes, among which 5% were specific for the Melan-A/A2 epitope and 11% were of unknown specificity (Fig. 1 A). This TIL population was then tested for recognition of a large panel of known antigens (Table I) transfected into COS cells with the class I HLA molecules of patient M170 (14, 19), and no response aside from the Melan-A/A2 response could be detected (unpublished data), suggesting that this population contained lymphocytes specific for new tumor antigens. To characterize them, we derived tumor-reactive CD8+ T cell clones by limiting dilution. Eight of these CTL clones showed reactivity patterns consistent with the recognition of new antigens, and one of them, hereafter referred to as M170.48, was further characterized to determine the HLA context restricting its recognition. As illustrated by Fig. 1 B, the recognition of the autologous melanoma cell line occurs in the HLA-A2 context. To establish the distribution of the target antigen, we tested M170.48 reactivity toward various HLA-A2 tumor cell lines, including melanomas, ovarian carcinomas, lung carcinomas, breast carcinomas, renal carcinomas, and colon carcinomas, using a TNF release assay. As shown in Fig. 1 C, this T cell clone recognized all of the HLA-A2 melanoma cell lines tested but none of the other HLA-A2 tumor cell types. In addition, the M170.48 T cell clone weakly recognized HLA-A2 melanocytes (Fig. 1 D). However, this reactivity was much lower than that seen with a Melan-A/A2–specific T cell clone (Fig. 1 D, hatched bars).


MELOE-1 is a new antigen overexpressed in melanomas and involved in adoptive T cell transfer efficiency.

Godet Y, Moreau-Aubry A, Guilloux Y, Vignard V, Khammari A, Dreno B, Jotereau F, Labarriere N - J. Exp. Med. (2008)

T cell clone selection and characterization. (A) Percentage of TNF-producing T cells and of HLA-A2/Melan-AA27L tetramer–positive T cells in the M170 TIL population in response to the autologous melanoma cell line. 105 TILs and 2 × 105 melanoma cells were incubated for 5 h in the presence of Brefeldin A, stained with HLA-A2/Melan-AA27L tetramer, fixed, and stained with anti-TNF antibody in a permeabilization buffer. 104 T cells were then analyzed by flow cytometry. (B) TNF secretion by the M170.48 T cell clone in response to the autologous melanoma cell line. 104 CTLs were added to 3 × 104 M170 melanoma cells in the presence of blocking antibodies directed against class I, A2, and B/C HLA, diluted to 1:50 (shaded bars), 1:500 (hatched bars), and 1:5,000 (open bars). CTL clone reactivity was assessed by a TNF release assay. (C) TNF response of the M170.48 CTL clone to HLA-A*0201 tumor cell lines. The M6 cell line (HLA-A2 negative) was used as a negative control. (D) IFN-γ response of the M170.48 CTL clone (shaded bars) and of a Melan-A/A2–specific CTL clone (hatched bars) to HLA-A*0201 melanocytes. The M170 cell line was added as a positive control.
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fig1: T cell clone selection and characterization. (A) Percentage of TNF-producing T cells and of HLA-A2/Melan-AA27L tetramer–positive T cells in the M170 TIL population in response to the autologous melanoma cell line. 105 TILs and 2 × 105 melanoma cells were incubated for 5 h in the presence of Brefeldin A, stained with HLA-A2/Melan-AA27L tetramer, fixed, and stained with anti-TNF antibody in a permeabilization buffer. 104 T cells were then analyzed by flow cytometry. (B) TNF secretion by the M170.48 T cell clone in response to the autologous melanoma cell line. 104 CTLs were added to 3 × 104 M170 melanoma cells in the presence of blocking antibodies directed against class I, A2, and B/C HLA, diluted to 1:50 (shaded bars), 1:500 (hatched bars), and 1:5,000 (open bars). CTL clone reactivity was assessed by a TNF release assay. (C) TNF response of the M170.48 CTL clone to HLA-A*0201 tumor cell lines. The M6 cell line (HLA-A2 negative) was used as a negative control. (D) IFN-γ response of the M170.48 CTL clone (shaded bars) and of a Melan-A/A2–specific CTL clone (hatched bars) to HLA-A*0201 melanocytes. The M170 cell line was added as a positive control.
Mentions: The M170 TIL population contained 16% of melanoma-reactive lymphocytes, among which 5% were specific for the Melan-A/A2 epitope and 11% were of unknown specificity (Fig. 1 A). This TIL population was then tested for recognition of a large panel of known antigens (Table I) transfected into COS cells with the class I HLA molecules of patient M170 (14, 19), and no response aside from the Melan-A/A2 response could be detected (unpublished data), suggesting that this population contained lymphocytes specific for new tumor antigens. To characterize them, we derived tumor-reactive CD8+ T cell clones by limiting dilution. Eight of these CTL clones showed reactivity patterns consistent with the recognition of new antigens, and one of them, hereafter referred to as M170.48, was further characterized to determine the HLA context restricting its recognition. As illustrated by Fig. 1 B, the recognition of the autologous melanoma cell line occurs in the HLA-A2 context. To establish the distribution of the target antigen, we tested M170.48 reactivity toward various HLA-A2 tumor cell lines, including melanomas, ovarian carcinomas, lung carcinomas, breast carcinomas, renal carcinomas, and colon carcinomas, using a TNF release assay. As shown in Fig. 1 C, this T cell clone recognized all of the HLA-A2 melanoma cell lines tested but none of the other HLA-A2 tumor cell types. In addition, the M170.48 T cell clone weakly recognized HLA-A2 melanocytes (Fig. 1 D). However, this reactivity was much lower than that seen with a Melan-A/A2–specific T cell clone (Fig. 1 D, hatched bars).

Bottom Line: Remarkably, the structure of meloe was unusual, with multiple short open reading frames (ORFs).The peptide recognized by the CTL clone was encoded by one of these ORFs, called MELOE-1.Using a specific HLA-A2/peptide tetramer, we showed a correlation between the infusion of TILs containing MELOE-1-specific T cells and relapse prevention in HLA-A2 patients.

View Article: PubMed Central - PubMed

Affiliation: Institut National de Santé et de Recherche Médicale, Unité Mixte de Recherche 892, 44093 Nantes, France.

ABSTRACT
A cytotoxic T lymphocyte (CTL) clone was derived from a tumor-infiltrating lymphocyte (TIL) population infused to a melanoma patient who remained relapse free for 10 yr after this adoptive transfer. This clone recognized all melanoma cell lines tested and, to a lower extent, melanocytes, in the context of human histocompatibility leukocyte antigen A2 (HLA-A2), but it did not recognize other tumor cell types. The gene coding for the antigen recognized by this clone was identified by the screening of a melanoma complementary DNA expression library. This antigen is overexpressed in melanomas, compared with other cancer cell lines and healthy tissues, and was thus called melanoma-overexpressed antigen (meloe). Remarkably, the structure of meloe was unusual, with multiple short open reading frames (ORFs). The peptide recognized by the CTL clone was encoded by one of these ORFs, called MELOE-1. Using a specific HLA-A2/peptide tetramer, we showed a correlation between the infusion of TILs containing MELOE-1-specific T cells and relapse prevention in HLA-A2 patients. Indeed, 5 out of 9 patients who did not relapse were infused with TILs that contained MELOE-1-specific T cells, whereas 0 out of the 21 patients who relapsed was infused with such TIL-containing lymphocytes. Overall, our results suggest that this new antigen is involved in immunosurveillance and, thus, represents an attractive target for immunotherapy protocols of melanoma.

Show MeSH
Related in: MedlinePlus