Narcolepsy Type 1 (NT1) is a neurological sleep disorder, characterized by the loss of hypocretin/orexin signaling in the brain. Genetic, epidemiological and experimental data support the hypothesis ...that NT1 is a T-cell-mediated autoimmune disease targeting the hypocretin producing neurons. While autoreactive CD4
T cells have been detected in patients, CD8
T cells have only been examined to a minor extent. Here we detect CD8
T cells specific toward narcolepsy-relevant peptides presented primarily by NT1-associated HLA types in the blood of 20 patients with NT1 as well as in 52 healthy controls, using peptide-MHC-I multimers labeled with DNA barcodes. In healthy controls carrying the disease-predisposing HLA-DQB1*06:02 allele, the frequency of autoreactive CD8
T cells was lower as compared with both NT1 patients and HLA-DQB1*06:02-negative healthy individuals. These findings suggest that a certain level of CD8
T-cell reactivity combined with HLA-DQB1*06:02 expression is important for NT1 development.
CD8
T cell reactivity towards tumor mutation-derived neoantigens is widely believed to facilitate the antitumor immunity induced by immune checkpoint blockade (ICB). Here we show that broadening in ...the number of neoantigen-reactive CD8
T cell (NART) populations between pre-treatment to 3-weeks post-treatment distinguishes patients with controlled disease compared to patients with progressive disease in metastatic urothelial carcinoma (mUC) treated with PD-L1-blockade. The longitudinal analysis of peripheral CD8
T cell recognition of patient-specific neopeptide libraries consisting of DNA barcode-labelled pMHC multimers in a cohort of 24 patients from the clinical trial NCT02108652 also shows that peripheral NARTs derived from patients with disease control are characterised by a PD1
Ki67
effector phenotype and increased CD39 levels compared to bystander bulk- and virus-antigen reactive CD8
T cells. The study provides insights into NART characteristics following ICB and suggests that early-stage NART expansion and activation are associated with response to ICB in patients with mUC.
Human endogenous retroviruses (HERV) form a substantial part of the human genome, but mostly remain transcriptionally silent under strict epigenetic regulation, yet can potentially be reactivated by ...malignant transformation or epigenetic therapies. Here, we evaluate the potential for T cell recognition of HERV elements in myeloid malignancies by mapping transcribed HERV genes and generating a library of 1169 potential antigenic HERV-derived peptides predicted for presentation by 4 HLA class I molecules. Using DNA barcode-labeled MHC-I multimers, we find CD8
T cell populations recognizing 29 HERV-derived peptides representing 18 different HERV loci, of which HERVH-5, HERVW-1, and HERVE-3 have more profound responses; such HERV-specific T cells are present in 17 of the 34 patients, but less frequently in healthy donors. Transcriptomic analyses reveal enhanced transcription of the HERVs in patients; meanwhile DNA-demethylating therapy causes a small and heterogeneous enhancement in HERV transcription without altering T cell recognition. Our study thus uncovers T cell recognition of HERVs in myeloid malignancies, thereby implicating HERVs as potential targets for immunotherapeutic therapies.
Adoptive cell therapy may be based on isolation of tumor-specific T cells, e.g. autologous tumor infiltrating lymphocytes (TIL), in vitro activation and expansion and the reinfusion of these cells ...into patients upon chemotherapy induced lymphodepletion. Together with high-dose interleukin (IL)-2 this treatment has been given to patients with advanced malignant melanoma and impressive response rates but also significant IL-2 associated toxicity have been observed. Here we present data from a feasibility study at a Danish Translational Research Center using TIL adoptive transfer in combination with low-dose subcutaneous IL-2 injections.
This is a pilot trial (ClinicalTrials.gov identifier: NCT00937625) including patients with metastatic melanoma, PS ≤1, age <70, measurable and progressive disease and no involvement of the central nervous system. Six patients were treated with lymphodepleting chemotherapy, TIL infusion, and 14 days of subcutaneous low-dose IL-2 injections, 2 MIU/day.
Low-dose IL-2 considerably decreased the treatment related toxicity with no grade 3-4 IL-2 related adverse events. Objective clinical responses were seen in 2 of 6 treated patients with ongoing complete responses (30+ and 10+ months), 2 patients had stable disease (4 and 5 months) and 2 patients progressed shortly after treatment. Tumor-reactivity of the infused cells and peripheral lymphocytes before and after therapy were analyzed. Absolute number of tumor specific T cells in the infusion product tended to correlate with clinical response and also, an induction of peripheral tumor reactive T cells was observed for 1 patient in complete remission.
Complete and durable responses were induced after treatment with adoptive cell therapy in combination with low-dose IL-2 which significantly decreased toxicity of this therapy.
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Novel single-cell-based technologies hold the promise of matching T cell receptor (TCR) sequences with their cognate peptide-MHC recognition motif in a high-throughput manner. Parallel capture of TCR ...transcripts and peptide-MHC is enabled through the use of reagents labeled with DNA barcodes. However, analysis and annotation of such single-cell sequencing (SCseq) data are challenged by dropout, random noise, and other technical artifacts that must be carefully handled in the downstream processing steps. We here propose a rational, data-driven method termed ITRAP (improved T cell Receptor Antigen Paring) to deal with these challenges, filtering away likely artifacts, and enable the generation of large sets of TCR-pMHC sequence data with a high degree of specificity and sensitivity, thus outputting the most likely pMHC target per T cell. We have validated this approach across 10 different virus-specific T cell responses in 16 healthy donors. Across these samples, we have identified up to 1494 high-confident TCR-pMHC pairs derived from 4135 single cells.
Personalization of immunotherapies such as cancer vaccines and adoptive T cell therapy depends on identification of patient-specific neo-epitopes that can be specifically targeted. MuPeXI, the mutant ...peptide extractor and informer, is a program to identify tumor-specific peptides and assess their potential to be neo-epitopes. The program input is a file with somatic mutation calls, a list of HLA types, and optionally a gene expression profile. The output is a table with all tumor-specific peptides derived from nucleotide substitutions, insertions, and deletions, along with comprehensive annotation, including HLA binding and similarity to normal peptides. The peptides are sorted according to a priority score which is intended to roughly predict immunogenicity. We applied MuPeXI to three tumors for which predicted MHC-binding peptides had been screened for T cell reactivity, and found that MuPeXI was able to prioritize immunogenic peptides with an area under the curve of 0.63. Compared to other available tools, MuPeXI provides more information and is easier to use. MuPeXI is available as stand-alone software and as a web server at
http://www.cbs.dtu.dk/services/MuPeXI
.
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EMUNI, FZAB, GEOZS, IJS, IMTLJ, KILJ, KISLJ, MFDPS, NUK, OBVAL, OILJ, PNG, SAZU, SBCE, SBJE, SBMB, SBNM, UKNU, UL, UM, UPUK, VKSCE, ZAGLJ
7.
Immune Mechanisms in Myelodysplastic Syndrome Glenthøj, Andreas; Ørskov, Andreas Due; Hansen, Jakob Werner ...
International journal of molecular sciences,
06/2016, Volume:
17, Issue:
6
Journal Article
Peer reviewed
Open access
Myelodysplastic syndrome (MDS) is a spectrum of diseases, characterized by debilitating cytopenias and a propensity of developing acute myeloid leukemia. Comprehensive sequencing efforts have ...revealed a range of mutations characteristic, but not specific, of MDS. Epidemiologically, autoimmune diseases are common in patients with MDS, fueling hypotheses of common etiological mechanisms. Both innate and adaptive immune pathways are overly active in the hematopoietic niche of MDS. Although supportive care, growth factors, and hypomethylating agents are the mainstay of MDS treatment, some patients-especially younger low-risk patients with HLA-DR15 tissue type-demonstrate impressive response rates after immunosuppressive therapy. This is in contrast to higher-risk MDS patients, where several immune activating treatments, such as immune checkpoint inhibitors, are in the pipeline. Thus, the dual role of immune mechanisms in MDS is challenging, and rigorous translational studies are needed to establish the value of immune manipulation as a treatment of MDS.
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Infusion of highly heterogeneous populations of autologous tumor-infiltrating lymphocytes (TIL) can result in tumor regression of exceptional duration. Initial tumor regression has been associated ...with persistence of tumor-specific TILs 1 month after infusion, but mechanisms leading to long-lived memory responses are currently unknown. Here, we studied the dynamics of bulk tumor-reactive CD8
T-cell populations in patients with metastatic melanoma following treatment with TILs.
We analyzed the function and phenotype of tumor-reactive CD8
T cells contained in serial blood samples of 16 patients treated with TILs.
Polyfunctional tumor-reactive CD8
T cells accumulated over time in the peripheral lymphocyte pool. Combinatorial analysis of multiple surface markers (CD57, CD27, CD45RO, PD-1, and LAG-3) showed a unique differentiation pattern of polyfunctional tumor-reactive CD8
T cells, with highly specific PD-1 upregulation early after infusion. The differentiation and functional status appeared largely stable for up to 1 year after infusion. Despite some degree of clonal diversification occurring
within the bulk tumor-reactive CD8
T cells, further analyses showed that CD8
T cells specific for defined tumor antigens had similar differentiation status.
We demonstrated that tumor-reactive CD8
T-cell subsets that persist after TIL therapy are mostly polyfunctional, display a stable partially differentiated phenotype, and express high levels of PD-1. These partially differentiated PD-1
polyfunctional TILs have a high capacity for persistence and may be susceptible to PD-L1/PD-L2-mediated inhibition.
.
...even though the vast majority of the computational tools are open source and thus freely available, most do not have user-friendly interfaces, limiting their use to investigators with programming ...expertise. ...the OpenCyto framework provides open-source tools for analyzing FCM data within an extensible and flexible interface to simplify the construction of re-usable FCM workflows while facilitating comparative analysis against manually gated results in order to enhance user confidence (Finak et al., 2014).
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
The use of fluorescently labeled major histocompatibility complex multimers has become an essential technique for analyzing disease- and therapy-induced T-cell immunity. Whereas classical major ...histocompatibility complex multimer analyses are well-suited for the detection of immune responses to a few epitopes, limitations on human-subject sample size preclude a comprehensive analysis of T-cell immunity. To address this issue, we developed a combinatorial encoding strategy that allows the parallel detection of a multitude of different T-cell populations in a single sample. Detection of T cells from peripheral blood by combinatorial encoding is as efficient as detection with conventionally labeled multimers but results in a substantially increased sensitivity and, most notably, allows comprehensive screens to be performed. We obtained proof of principle for the feasibility of large-scale screening of human material by analysis of human leukocyte antigen A3-restricted T-cell responses to known and potential melanoma-associated antigens in peripheral blood from individuals with melanoma.
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