Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:1.10.3.1 (tyrosinase)
9,065 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Many peptide epitopes for cytotoxic T lymphocytes (CTLs) have been identified from melanocytic differentiation proteins. Vaccine trials with these peptides have been limited mostly to those associated with HLA-A2, and immune responses have been detected inconsistently. Cases of clinical regression have been observed after peptide vaccination in some trials, but melanoma regressions have not correlated well with T-cell responses measured in peripheral blood lymphocytes (PBLs). We vaccinated stage IV melanoma patients with a mixture of gp100 and tyrosinase peptides restricted by HLA-A1 (DAEKSDICTDEY), HLA-A2 (YLEPGPVTA and YMDGTMSQV) and HLA-A3 (ALLAVGATK) in an emulsion with GM-CSF and Montanide ISA-51 adjuvant. CTL responses were assessed in PBLs and in a lymph node draining a vaccine site (sentinel immunized node, SIN). We found CTL responses to vaccinating peptides in the SIN in 5/5 patients (100%). Equivalent assays detected peptide-reactive CTLs in PBLs of 2 of these 5 patients (40%). CTLs expanded from the SIN lysed melanoma cells naturally expressing tyrosinase or gp100. We demonstrated immunogenicity for peptides restricted by HLA-A1 and -A3 and for 1 HLA-A2 restricted peptide, YMDGTMSQV. Immune monitoring of clinical trials by evaluation of PBLs alone may under-estimate immunogenicity; evaluation of SIN provides a new and sensitive approach for defining responses to tumor vaccines and correlating these responses with clinical outcomes. This combination of an immunogenic vaccine strategy with a sensitive analysis of CTL responses demonstrates the potential for inducing and detecting anti-tumor immune responses in the majority of melanoma patients.
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PMID:Evaluation of peptide vaccine immunogenicity in draining lymph nodes and peripheral blood of melanoma patients. 1134 May 76

In previous studies CD8+ T cells specific for melanocyte antigens have been frequently found in melanoma patients responding to interleukin-2 (IL-2)-based therapies. In our study we analyzed the suitability of using circulating T cells from melanoma patients with clinical response after IL-2-based therapy to identify novel T-cell epitopes from defined tumor antigens. Using unstimulated peripheral blood mononuclear cells and the interferon-gamma (IFN-gamma) ELISPOT assay, we studied CD8(+) T-cell responses against 5 peptides from the tumor antigen tyrosinase (Tyr) selected by epitope prediction using an HLA-A1-binding computer algorithm. T cells specifically secreting IFN-gamma in response to 3 of these 5 peptides, namely, Tyr (454-463), Tyr (146-156) and Tyr (243-251), could be detected in 4 of 4 HLA-A1-positive patients with clinical response. In contrast, no T-cell responses against these peptides were seen in 6 HLA-A1-positive melanoma patients with progressive disease and in 8 healthy subjects. We could generate specific cytotoxic T lymphocytes (CTL) against Tyr (454-463) using peptide-pulsed autologous dendritic cells as antigen-presenting cells. The induced CTLs efficiently killed melanoma cells that express HLA-A1 and tyrosinase. The peptides Tyr (146-156) and Tyr (243-251) had recently been identified as CTL epitopes by other groups. Further ex vivo characterization of the T cells reactive against the novel epitope Tyr (454-463) in 1 patient by multicolor flow cytometry showed specific CD3+/CD8+/IFN-gamma+ T cells with frequencies of up to 0.41% of the CD3+/CD8+ T-cell population. Most of this T-cell population also expressed granzyme B. Our data confirm that in patients with tumor regressions induced by immunotherapy or chemoimmunotherapy circulating T cells reactive with tyrosinase epitopes can frequently be detected. Peripheral blood T cells from such patients are a valuable source for screening peptides selected by epitope prediction This strategy facilitates the rapid identification of immunogenic T-cell epitopes that are probable targets of immune-mediated tumor rejection.
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PMID:Identification of known and novel immunogenic T-cell epitopes from tumor antigens recognized by peripheral blood T cells from patients responding to IL-2-based treatment. 1192 May 92

The major goal of therapeutic cancer vaccine trials is to mediate tumor regression. However, it is critically important to devise in vitro immunological assays that correlate with clinical outcome, for use as surrogate markers of vaccine efficacy. To date, clinical emphasis has been placed on peptide vaccines, but trends towards the use of more complex immunogens such as whole proteins require the development of efficient and sensitive methods for monitoring their immunological effects. In the context of a vaccination trial using full-length tyrosinase (Ty) to immunize patients with metastatic melanoma, a monitoring technique was developed in which autologous dendritic cells (DC) infected with a recombinant adenovirus encoding the Ty protein were used to assess the Ty-specific reactivity of fresh peripheral blood lymphocytes (PBL) collected from patients at different intervals during therapy. Quantitative real-time RT-PCR (qRT-PCR) was used to measure the production of cytokine mRNA by T cells following a 2.5-h incubation with Ty-expressing DC. Two out of ten patients studied demonstrated Ty protein-specific reactivity that increased during and after the period of vaccination. While one of these patients also reacted to an HLA-A1-compatible Ty peptide, the second did not recognize any of the known Ty epitopes, highlighting the importance of this technique for monitoring the effects of complex vaccines.
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PMID:Quantitative real-time RT-PCR as a method for monitoring T lymphocyte reactivity to full-length tyrosinase protein in vaccinated melanoma patients. 1213 25

Immune therapy has provided a significant breakthrough in the treatment of metastatic melanoma. Despite the remarkable clinical efficacy and established involvement of effector CD8 T cells, the knowledge of the exact peptide-MHC complexes recognized by T cells on the tumor cell surface is limited. Many melanoma-associated T-cell epitopes have been described, but this knowledge remains largely restricted to HLA-A2, and we lack understanding of the T-cell recognition in the context of other HLA molecules. We selected six melanoma-associated antigens (MAGE-A3, NY-ESO-1, gp100, Mart1, tyrosinase and TRP-2) that are frequently recognized in patients with the aim of identifying novel T-cell epitopes restricted to HLA-A1, -A3, -A11 and -B7. Using in silico prediction and in vitro confirmation, we identified 127 MHC ligands and analyzed the T-cell responses against these ligands via the MHC multimer-based enrichment of peripheral blood from 39 melanoma patients and 10 healthy donors. To dissect the T-cell reactivity against this large peptide library, we used combinatorial-encoded MHC multimers and observed the T-cell responses against 17 different peptide-MHC complexes in the patient group and four in the healthy donor group. We confirmed the processing and presentation of HLA-A3-restricted T-cell epitopes from tyrosinase (TQYESGSMDK) and gp100 (LIYRRRLMK) and an HLA-A11-restricted T-cell epitope from gp100 (AVGATKVPR) via the cytolytic T-cell recognition of melanoma cell lines and/or K562 cells expressing the appropriate antigen and HLA molecule. We further found T-cell reactivity against two of the identified sequences among tumor-infiltrating lymphocytes from melanoma patients, suggesting a potential clinical relevance of these sequences.
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PMID:Broadening the repertoire of melanoma-associated T-cell epitopes. 2585 82

This review is focused on research within three different areas of tumor immunology: discovery of new T-cell epitopes and a new immunological antigen (reported in Paper I and II), elucidation of the immunological effects of treatment with a hypomethylating drug (reported in Paper III) and discovery of new conditional ligands (reported in Paper IV). Many melanoma-associated T-cell epitopes have been described, but 45% of these are restricted to human leukocyte antigen (HLA)-A2, leaving the remaining 36 different HLA molecules with only a few described T-cell epitopes each. Therefore we wanted to expand the number of T-cell epitopes restricted to HLA-A1, -A3, -A11 and -B7, all HLA molecules frequently expressed in Caucasians in Western Europe and Northern America. In Paper I we focused on the proteins gp100, Mart1, MAGE-A3, NY-ESO-1, tyrosinase and TRP-2, all melanoma-associated antigens frequently recognized by T cells from HLA-A2 patients. On contrary, in Paper II we wanted to investigate the protein Nodal as a novel immunological target. We took advantage of a T-cell epitope mapping platform in which HLA ligands are predicted by computer-based algorithms, further tested in the laboratory by an ELISA-based method and used for flow cytometry-based detection of specific T-cell responses by use of combinatorial encoded major histocompatibility (MHC) class I multimers. This procedure resulted in 127 (Paper I) and 32 (Paper II) confirmed HLA ligands, respectively, which we used for screening of the T-cell recognition within peripheral blood mononuclear cell samples from melanoma patients. As spontaneous tumor-specific T-cell responses tend to be of very low frequency and probably below the detection threshold of the method, we incorporated a T-cell enrichment step prior to the detection of these responses. Our screening of 39 melanoma patients resulted in 26 (17 different) T-cell responses against the common melanoma-associated antigens and 10 (8 different) T-cell responses against Nodal. We were further able to show processing and presentation on the cell-surface in K562 and melanoma cells expressing relevant protein and HLA molecules of four of these peptide sequences from tyrosinase, gp100 (2 peptides) and Nodal, respectively. However, one of the gp100 peptides has previously been described as a T-cell epitope. In addition to identifying new melanoma-associated T-cell epitopes we could thus describe Nodal as a new immunological antigen found of relevance in melanoma patients. In Paper III we wanted to investigate if the hypomethylating drug 5-azactytidine (Vidaza, Celgene Inc.) modulates the immune system in patients with myeloproliferative diseases. It has previ-ously been shown that 5-azacytidine-mediated demethylation of gene promoter regions results in enhanced transcription and expression of tumor suppressor genes and cancer-testis antigens. Cancer-testis antigens have frequently been recognized by T-cells in many cancers, and we hypothesized that 5-azacytidine treat-ment in the clinic would increase their frequency with resulting enhanced anti-tumor reactivity. We investigated separately the effect on T cells and tumor cells, and found that tumor cells af-fected by the treatment were better recognized, resulting in higher numbers of activated T cells, than tumor cells not exposed to 5-azacytidine. No effects were observed on the T-cell population. A screen of the T-cell recognition of 43 cancer-testis antigens in blood from our patients revealed increased T-cell recognition upon start of therapy which, though, stabilized or declined at later time points. We further investigated the general immune effector and inhibitory cell populations and found only minor effects of drug exposure, suggesting that 5-azacytidine primarily affects the tumor cells. From these results we are currently initiating a phase I clinical trial of cancer-testis antigen-peptide vaccination in combination with 5-azacytidine therapy for patients with myeloproliferative diseases. In Paper IV we wanted to expand the library of conditional ligands for use with the UV light-mediated peptide-exchange method. This method enables high-throughput generation of MHC class I molecules with different peptide-specificities. These MHC monomers can be multimerized and used for detection of specific T cell populations by flow or mass cytometry. The HLA molecules are highly genetically variable and this necessitates unique design of conditional ligands for each HLA molecule. Thus, to screen for the T-cell recognition in a given setting within all patients or healthy donors present in a cohort, a broad library of conditional ligands is needed. We designed and evaluated conditional ligands for HLA-B*08:01, HLA-B*35:01 and HLA-B*44:02/03/05, all HLA-B molecules present in high frequency among Caucasians. In addition, we provided proof for the use of a conditional ligand first designed for HLA-B*15:02 in complex with HLA-B*15:01. We compared the staining patterns of HLA-B*15:01 and HLA-B*15:02 MHC multimers and found remarkable dissimilarities, although the two heavy chains in these MHC molecules only differ in a few amino acid positions.
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PMID:Elucidating the immunological effects of 5-azacytidine treatment in patients with myelodysplastic syndrome and identifying new conditional ligands and T-cell epitopes of relevance in melanoma. 2623 96


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