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T cells specific for neoantigens encoded by mutated genes in cancers are increasingly recognized as mediators of tumor destruction after immune checkpoint inhibitor therapy or adoptive cell transfer. Unfortunately, most neoantigens result from random mutations and are patient specific, and some cancers contain few mutations to serve as potential antigens. We describe a patient with stage IV acral melanoma who achieved a complete response following adoptive transfer of tumor-infiltrating lymphocytes (TILs). Tumor exome sequencing surprisingly revealed fewer than 30 nonsynonymous somatic mutations, including oncogenic [BRAF.sup.V600E]. Analysis of the specificity of TILs identified rare [CD4.sup.+] T cells specific for [BRAF.sup.V600E] and diverse [CD8.sup.+] T cells reactive to nonmutated self-antigens. These specificities increased in blood after TIL transfer and persisted long-term, suggesting they contributed to the effective antitumor immune response. Gene transfer of the [BRAF.sup.V600E]-specific T cell receptor (TCR) conferred recognition of class II MHC-positive cells expressing the BRAF mutation. Therapy with TCR-engineered [BRAF.sup.V600E]-specific [CD4.sup.+] T cells may have direct antitumor effects and augment [CD8.sup.+] T cell responses to self- and/or mutated tumor antigens in patients with BRAF-mutated cancers., Introduction T cells can eliminate cancer cells through recognition of peptides from nonmutated or mutated proteins bound to cell surface MHC molecules (1). T cells specific for neoantigens derived from [...]

Dominant protection against type 1 diabetes: Polymorphic histocompatibility genes in the HLA locus have a strong influence on genetic susceptibility to type 1 diabetes (T1D). Several high-risk HLA haplotypes increase susceptibility to T1D, whereas the DR1501-DQ6 HLA haplotype confers dominant protection. Wen et al. investigated the mechanistic basis for this protective effect by measuring the frequency of CD4+ T cells reactive with epitopes on islet autoantigens in healthy individuals. Individuals with the protective HLA haplotype had a higher frequency of islet antigen–reactive CD4+ T cells than individuals with high-risk or neutral haplotypes. These CD4+ T cells included both regulatory T cells and effector T cells with a propensity to make IL-10. These findings provide deeper insights into how beneficial subsets of self-reactive T cells maintain prophylaxis against autoimmunity. The DRB1*15:01-DQB1*06:02 (DR1501-DQ6) haplotype is linked to dominant protection from type 1 diabetes, but the cellular mechanism for this association is unclear. To address this question, we identified multiple DR1501- and DQ6-restricted glutamate decarboxylase 65 (GAD65) and islet-specific glucose-6-phosphatase catalytic subunit–related protein (IGRP)–specific T cell epitopes. Three of the DR1501/DQ6-restricted epitopes identified were previously reported to be restricted by DRB1*04:01/DRB1*03:01/DQB1*03:02. We also used specific class II tetramer reagents to assess T cell frequencies. Our results indicated that GAD65- and IGRP-specific effector and CD25+CD127−FOXP3+ regulatory CD4+ T cells were present at higher frequencies in individuals with the protective haplotype than those with susceptible or neutral haplotypes. We further confirmed higher frequencies of islet antigen–specific effector and regulatory CD4+ T cells in DR1501-DQ6 individuals through a CD154/CD137 up-regulation assay. DR1501-restricted effector T cells were capable of producing interferon-γ (IFN-γ) and interleukin-4 (IL-4) but were more likely to produce IL-10 compared with effectors from individuals with susceptible haplotypes. To evaluate their capacity for antigen-specific regulatory activity, we cloned GAD65 and IGRP epitope–specific regulatory T cells. We showed that these regulatory T cells suppressed DR1501-restricted GAD65- and IGRP-specific effectors and DQB1*03:02-restricted GAD65-specific effectors in an antigen-specific fashion. In total, these results suggest that the protective DR1501-DQ6 haplotype confers protection through increased frequencies of islet-specific IL-10–producing T effectors and CD25+CD127−FOXP3+ regulatory T cells. [ABSTRACT FROM AUTHOR]

In spite of tolerance mechanisms, some individuals develop T-cell-mediated autoimmunity. Posttranslational modifications that increase the affinity of epitope presentation and/or recognition represent one means through which self-tolerance mechanisms can be circumvented. We investigated T-cell recognition of peptides that correspond to modified β-cell antigens in subjects with type 1 diabetes. Modified peptides elicited enhanced proliferation by autoreactive T-cell clones. Endoplasmic reticulum (ER) stress in insulinoma cells increased cytosolic calcium and the activity of tissue transglutaminase 2 (tTG2). Furthermore, stressed human islets and insulinomas elicited effector responses from T cells specific for modified peptides, suggesting that ER stress-derived tTG2 activity generated deamidated neoepitopes that autoreactive T cells recognized. Patients with type 1 diabetes had large numbers of T cells specific for these epitopes in their peripheral blood. T cells with these specificities were also isolated from the pancreatic draining lymph nodes of cadaveric donors with established diabetes. Together, these results suggest that self-antigens are enzymatically modified in β-cells during ER stress, giving rise to modified epitopes that could serve to initiate autoimmunity or to further broaden the antigenic repertoire, activating potentially pathogenic CD4+ T cells that may not be effectively eliminated by negative selection. [ABSTRACT FROM AUTHOR]

Monitoring the frequency of circulatory CXCR5+ (cCXCR5+) CD4+ T cells in periphery blood provides a potential biomarker to draw inferences about T follicular helper (TFH) activity within germinal center. However, cCXCR5+ T cells are highly heterogeneous in their expression of ICOS, PD1 and CD38 and the relationship between different cCXCR5 subsets as delineated by these markers remains unclear. We applied class II tetramer reagents and mass cytometry to investigate the ontogeny of different subsets of cCXCR5+ T cell following yellow fever immunization. Through unsupervised analyses of mass cytometry data, we show yellow fever virus-specific cCXCR5 T cells elicited by vaccination were initially CD38+ICOS+PD1+, but then transitioned to become CD38+ICOS−PD1+ and CD38−ICOS−PD1+ before coming to rest as a CD38−ICOS−PD1− subset. These results imply that most antigen-specific cCXCR5+ T cells, including the CD38−ICOS−PD1− CXCR5+ T cells are derived from the CXCR5+CD38+ICOS+PD1+ subset, the subset that most resembles preTFH/TFH in the germinal center. [ABSTRACT FROM AUTHOR]

Tumor-specific T cells likely underpin effective immune checkpoint-blockade therapies. Yet, most studies focus on Treg cells and CD8 + tumor-infiltrating lymphocytes (TILs). Here, we study CD4 + TILs in human lung and colorectal cancers and observe that non-Treg CD4 + TILs average more than 70% of total CD4 + TILs in both cancer types. Leveraging high dimensional analyses including mass cytometry, we reveal that CD4 + TILs are phenotypically heterogeneous, within each tumor and across patients. Consistently, we find different subsets of CD4 + TILs showing characteristics of effectors, tissue resident memory (Trm) or exhausted cells (expressing PD-1, CTLA-4 and CD39). In both cancer types, the frequencies of CD39 - non-Treg CD4 + TILs strongly correlate with frequencies of CD39 - CD8 + TILs, which we and others have previously shown to be enriched for cells specific for cancer-unrelated antigens (bystanders). Ex-vivo , we demonstrate that CD39 - CD4 + TILs can be specific for cancer-unrelated antigens, such as HCMV epitopes. Overall, our findings highlight that CD4 + TILs can also recognize cancer-unrelated antigens and suggest measuring CD39 expression as a straightforward way to quantify or isolate bystander CD4 + T cells., Competing Interests: E.W.N is a co-founder, advisor and shareholder of ImmunoScape Pte. Ltd. E.W.N. is an advisor for Neogene Therapeutics and Nanostring Technologies. All other authors declare no potential conflicts of interest., (© 2022 The Author(s). Published with license by Taylor & Francis Group, LLC.)

Peptide-major histocompatibility complex class II (pMHCII) multimers have emerged as a convenient and powerful tool for characterization of CD4 T cell immune responses in a large variety of human diseases. Peptide-MHCII multimers can rapidly identify peptide antigens recognized by CD4 T cells via high-throughput peptide screening procedures. The specificity and phenotype of antigen-specific CD4 T cells can be effectively visualized by pMHCII multimers from unmanipulated immune cell populations. Functional attributes of antigen-specific CD4 T cells can also be defined with the multimer technology in combination with immune functional assays such as intracellular cytokine staining (ICS).

Although CD4 + T cells likely play key roles in antitumor immune responses, most immuno-oncology studies have been limited to CD8 + T-cell responses due to multiple technical barriers and a lack of shared antigens across patients. Merkel cell carcinoma (MCC) is an aggressive skin cancer caused by Merkel cell polyomavirus (MCPyV) oncoproteins in 80% of cases. Because MCPyV oncoproteins are shared across most patients with MCC, it is unusually feasible to identify, characterize, and potentially augment tumor-specific CD4 + T cells. Here, we report the identification of CD4 + T-cell responses against six MCPyV epitopes, one of which included a conserved, essential viral oncogenic domain that binds/disables the cellular retinoblastoma (Rb) tumor suppressor. We found that this epitope (WED LT209-228 ) could be presented by three population-prevalent HLA class II alleles, making it a relevant target in 64% of virus-positive MCC patients. Cellular staining with a WED LT209-228 -HLA-DRB1*0401 tetramer indicated that specific CD4 + T cells were detectable in 78% (14 of 18) of evaluable MCC patients, were 250-fold enriched within MCC tumors relative to peripheral blood, and had diverse T-cell receptor sequences. We also identified a modification of this domain that still allowed recognition by these CD4 + T cells but disabled binding to the Rb tumor suppressor, a key step in the detoxification of a possible therapeutic vaccine. The use of these new tools for deeper study of MCPyV-specific CD4 + T cells may provide broader insight into cancer-specific CD4 + T-cell responses., (©2019 American Association for Cancer Research.)

In spite of tolerance mechanisms, some individuals develop T-cell-mediated autoimmunity. Posttranslational modifications that increase the affinity of epitope presentation and/or recognition represent one means through which self-tolerance mechanisms can be circumvented. We investigated T-cell recognition of peptides that correspond to modified β-cell antigens in subjects with type 1 diabetes. Modified peptides elicited enhanced proliferation by autoreactive T-cell clones. Endoplasmic reticulum (ER) stress in insulinoma cells increased cytosolic calcium and the activity of tissue transglutaminase 2 (tTG2). Furthermore, stressed human islets and insulinomas elicited effector responses from T cells specific for modified peptides, suggesting that ER stress-derived tTG2 activity generated deamidated neoepitopes that autoreactive T cells recognized. Patients with type 1 diabetes had large numbers of T cells specific for these epitopes in their peripheral blood. T cells with these specificities were also isolated from the pancreatic draining lymph nodes of cadaveric donors with established diabetes. Together, these results suggest that self-antigens are enzymatically modified in β-cells during ER stress, giving rise to modified epitopes that could serve to initiate autoimmunity or to further broaden the antigenic repertoire, activating potentially pathogenic CD4 + T cells that may not be effectively eliminated by negative selection., (© 2018 by the American Diabetes Association.)