TY - JOUR
T1 - Identification of tumor-associated, MHC class II-restricted phosphopeptides as targets for immunotherapy
AU - Depontieu, Florence R.
AU - Qian, Jie
AU - Zarling, Angela L.
AU - McMiller, Tracee L.
AU - Salay, Theresa M.
AU - Norris, Andrew
AU - Michelle English, A.
AU - Shabanowitz, Jeffrey
AU - Engelhard, Victor H.
AU - Hunt, Donald F.
AU - Topalian, Suzanne L.
PY - 2009/7/21
Y1 - 2009/7/21
N2 - The activation and recruitment of CD4+ T cells are critical for the development of efficient antitumor immunity and may allow for the optimization of current cancer immunotherapy strategies. Searching for more optimal and selective targets for CD4+ T cells, we have investigated phosphopeptides, a new category of tumor-derived epitopes linked to proteins with vital cellular functions. Although MHC I-restricted phosphopeptides have been identified, it was previously unknown whether human MHC II molecules present phosphopeptides for specific CD4+ T cell recognition. We first demonstrated the fine specificity of human CD4+ T cells to discriminate a phosphoresidue by using cells raised against the candidate melanoma antigen mutant B-Raf or its phosphorylated counterpart. Then, we assessed the presence and complexity of human MHC II-associated phosphopeptides by analyzing 2 autologous pairs of melanoma and EBV-transformed B lymphoblastoid lines. By using sequential affinity isolation, biochemical enrichment, mass spectrometric sequencing, and comparative analysis, a total of 175 HLA-DR-associated phosphopeptides were characterized. Many were derived from source proteins that may have roles in cancer development, growth, and metastasis. Most were expressed exclusively by either melanomas or transformed B cells, suggesting the potential to define cell type-specific phosphatome "fingerprints." We then generated HLA-DRβ1*0101-restricted CD4+ T cells specific for a phospho-MART-1 peptide identified in both melanoma cell lines. These T cells showed specificity for phosphopeptide-pulsed antigen-presenting cells as well as for intact melanoma cells. This previously undescribed demonstration of MHC II-restricted phosphopeptides recognizable by human CD4+ T cells provides potential new targets for cancer immunotherapy.
AB - The activation and recruitment of CD4+ T cells are critical for the development of efficient antitumor immunity and may allow for the optimization of current cancer immunotherapy strategies. Searching for more optimal and selective targets for CD4+ T cells, we have investigated phosphopeptides, a new category of tumor-derived epitopes linked to proteins with vital cellular functions. Although MHC I-restricted phosphopeptides have been identified, it was previously unknown whether human MHC II molecules present phosphopeptides for specific CD4+ T cell recognition. We first demonstrated the fine specificity of human CD4+ T cells to discriminate a phosphoresidue by using cells raised against the candidate melanoma antigen mutant B-Raf or its phosphorylated counterpart. Then, we assessed the presence and complexity of human MHC II-associated phosphopeptides by analyzing 2 autologous pairs of melanoma and EBV-transformed B lymphoblastoid lines. By using sequential affinity isolation, biochemical enrichment, mass spectrometric sequencing, and comparative analysis, a total of 175 HLA-DR-associated phosphopeptides were characterized. Many were derived from source proteins that may have roles in cancer development, growth, and metastasis. Most were expressed exclusively by either melanomas or transformed B cells, suggesting the potential to define cell type-specific phosphatome "fingerprints." We then generated HLA-DRβ1*0101-restricted CD4+ T cells specific for a phospho-MART-1 peptide identified in both melanoma cell lines. These T cells showed specificity for phosphopeptide-pulsed antigen-presenting cells as well as for intact melanoma cells. This previously undescribed demonstration of MHC II-restricted phosphopeptides recognizable by human CD4+ T cells provides potential new targets for cancer immunotherapy.
KW - Tumor antigen
KW - Tumor immunology
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U2 - 10.1073/pnas.0903852106
DO - 10.1073/pnas.0903852106
M3 - Article
C2 - 19581576
AN - SCOPUS:67749145750
SN - 0027-8424
VL - 106
SP - 12073
EP - 12078
JO - Proceedings of the National Academy of Sciences of the United States of America
JF - Proceedings of the National Academy of Sciences of the United States of America
IS - 29
ER -