Peptide-major Histocompatibility Complex Research Articles

Cooperative binding of T cell receptor and CD4 to peptide-MHC enhances antigen sensitivity

Antigen recognition by the T cell receptor (TCR) of CD4+ T cells can be greatly enhanced by the coreceptor CD4. Yet, understanding of the molecular mechanism is hindered by the ultra-low affinity of CD4 binding to class-II peptide-major histocompatibility complexes (pMHC). Here we show, using two-dimensional (2D) mechanical-based assays, that the affinity of CD4–pMHC interaction is 3-4 logs lower than that of cognate TCR–pMHC interactions, and it is more susceptible to increased dissociation by forces (slip bond). In contrast, CD4 binds TCR-pre-bound pMHC at 3-6 logs higher affinity, forming TCR–pMHC–CD4 tri-molecular bonds that are prolonged by force (catch bond), and modulated by protein mobility on the cell membrane, indicating profound TCR-CD4 cooperativity. Consistent with a tri-crystal structure, using DNA origami as a molecular ruler to titrate spacing between TCR and CD4 we show that 7-nm proximity optimizes TCR–pMHC–CD4 tri-molecular bond formation with pMHC. Our results thus provide deep mechanistic insight into CD4 enhancement of TCR antigen recognition.

Nilabh Shastri – Towards understanding classical and non-classical MHC-I antigen processing and presentation

Major histocompatibility complex (MHC-I) peptide antigen processing and presentation has experienced a revived interest in the context of immuno oncology, immune surveillance escape by pathogen mutations and technical advances that accelerate vaccine design. This sheds new light on the discoveries made by Nilabh Shastri and colleagues that includes the characterisation of cryptic MHC-I peptide antigen epitopes derived from untranslated regions and the N-terminal trimming of peptide antigen precursors by the aminopeptidase ERAAP (ERAP1/2 / ARTS1/LRAP) in the endoplasmic reticulum (ER) prior to the complete assembly of MHC-I complexes and their subsequent exposure to the cell surface. These scientific findings have important implications for developing novel therapeutic approaches in immunotherapy and modern vaccine design.

Time required for commitment to T cell proliferation depends on TCR affinity and cytokine response.

T cell activation and effector functions are determined by the affinity of the interaction between T cell receptor (TCR) and its antigenic peptide MHC (pMHC) ligand. A better understanding of the quantitative aspects of TCR-pMHC affinity-dependent T cell activation is critical for the development of new immunotherapeutic strategies. However, the role of TCR-pMHC affinity in regulating the kinetics of CD8+ T cell commitment to proliferation and differentiation is unknown. Here, we show that the stronger the TCR-pMHC affinity, the shorter the time of T cell-APC co-culture required to commit CD8+ T cells to proliferation. The time threshold for T cell cytokine production is much lower than that for cell proliferation. There is a strong correlation between affinity-dependent differences in AKT phosphorylation and T cell proliferation. The cytokine IL-15 increases the poor proliferation of T cells stimulated with low affinity pMHC, suggesting that pro-inflammatory cytokines can override the affinity-dependent features of T cell proliferation.

Subcellular location of source proteins improves prediction of neoantigens for immunotherapy.

Antigen presentation via the major histocompatibility complex (MHC) is essential for anti-tumor immunity. However, the rules that determine which tumor-derived peptides will be immunogenic are still incompletely understood. Here, we investigated whether constraints on peptide accessibility to the MHC due to protein subcellular location are associated with peptide immunogenicity potential. Analyzing over 380,000 peptides from studies of MHC presentation and peptide immunogenicity, we find clear spatial biases in both eluted and immunogenic peptides. We find that including parent protein location improves the prediction of peptide immunogenicity in multiple datasets. In human immunotherapy cohorts, the location was associated with a neoantigen vaccination response, and immune checkpoint blockade responders generally had a higher burden of neopeptides from accessible locations. We conclude that protein subcellular location adds important information for optimizing cancer immunotherapies.

Extending the Mass Spectrometry-Detectable Landscape of MHC Peptides by Use of Restricted Access Material.

Mass spectrometry-based immunopeptidomics enables the comprehensive identification of major histocompatibility complex (MHC) peptides from a cell culture as well as from tissue or tumor samples and is applied for the identification of tumor-specific and viral T-cell epitopes. Although mass spectrometry is generally considered an "unbiased" method for MHC peptide identification, the physicochemical properties of MHC peptides can greatly influence their detectability. Here, we demonstrate that highly hydrophobic peptides are lost during sample preparation when C18 solid-phase extraction (SPE) is used for separating MHC peptides from proteins. To overcome this limitation, we established an optimized protocol involving restricted access material (RAM). Compared to C18-SPE, RAM-SPE improved the overall MHC peptide recovery and extended the landscape of mass spectrometry-detectable MHC peptides toward more hydrophobic peptides.

Influenza-Induced CD103+ T Resident Memory Cells Exhibit Enhanced Functional Avidity over CD103- Memory T Cells in the Mediastinal Lymph Node.

Influenza virus–specific tissue-resident memory CD8 T cells (Trms) targeting conserved viral proteins provide strain-transcending heterosubtypic immunity to infection. Trms in the lung combat reinfection through rapid cytolytic function and production of inflammatory cytokines to recruit other immune cells. Influenza-specific Trms are also generated in the lung draining mediastinal lymph node (mLN) and can provide immunity to heterologous virus infection in this tissue, although their role in combating influenza infection is less well defined. Functional avidity, a measure of T cell sensitivity to Ag stimulation, correlates with control of viral infection and may be important for immune detection of recently infected cells, when low numbers of surface peptide–MHC complexes are displayed. However, the functional avidity of influenza-specific Trms has not been previously compared with that of other memory CD8 T cell subsets. In this article, a methodology is presented to compare the functional avidity of CD8 T cell subsets across murine tissues, with a focus on influenza-specific mLNs compared with splenic CD8 T cells, by stimulating both populations in the same well to account for CD8 T cell–extrinsic variables. The functional avidity of influenza-specific mLN effector CD8 T cells is slightly increased relative to splenic effector CD8 T cells. However, CD103+ mLN Trms display increased functional avidity compared with splenic memory CD8 T cells and CD103− memory CD8 T cells within the mLN. In contrast, lung-derived CD103+ Trms did not exhibit enhanced functional avidity. mLN CD103+ Trms also exhibit increased TCR expression, providing a potential mechanism for their enhanced functional avidity.

T-cell receptor variable region usage in Chagas disease: A systematic review of experimental and human studies.

T cells recognize their ligand, the peptide major histocompatibility complex (MHC), via the T-cell receptor (TCR), which is composed of covalently linked α and β or γ and δ chains. This recognition is critical for T-cell ontogeny and controls the selection, activation, and function of T lymphocytes. Specific TCR αβ variable regions have been associated with immunopathogenesis of Chagas disease. Here, we present a systematic review that compiles experimental in vivo and human data regarding the preferential expression of variable alpha (Vα) and variable beta (Vβ) chain regions in Trypanosoma cruzi infection. The original studies indexed in PubMed/Medline, Scopus, and Web of Science databases were screened according to the PRISMA strategy. The analysis showed that expression of TCR Vα subfamilies were evaluated in one human study, and, unlike TCR Vβ, TCR Vα presented a more restricted usage. Despite the great variability in the usage of TCR Vβ regions in human Chagas disease, a down-regulation of TCR Vβ5 expression by T cells from patients in the acute phase of the disease was shown. Opposingly, this TCR region was found overly expressed in CD4+ T cells from chronic Chagas patients. It was also demonstrated that murine Vβ9+ T cells derived from nonlymphoid organs of T. cruzi-infected animals had a modulatory profile, while splenic Vβ9+ T cells produced inflammatory cytokines, indicating that although they display the same TCR Vβ region usage, these cells are functionally distinct. Despite the limitations of few papers and year of publication of the studies, compiling the data derived from them reveals that further investigation of TCR usage will point to their potential role in protective or pathogenic responses, as biomarkers of disease progression, and in the search for dominant peptides potentially useful for the development of vaccines or therapies.

Non-linear in cis-spliced bacterial MHC class II epitopes are formed in human monocyte-derived dendritic cells and are recognized by human CD4+ T cells

Pathogen-derived spliced MHC class II epitopes have not been reported so far. Earlier we studied HLA-DRB*0101-presented peptides derived from P1.7-2,4, a major outer membrane porin (PorA) serosubtype from Neisseria meningitidis, using stable isotope-tagging of the antigens. We found evidence for +114 Da and +270 Da in cis-spliced peptide variants, but could not unequivocally identify their sequence at the time. Here we aimed to confirm this biological phenomenon and determine the identity and immunogenicity of the in-cis-spliced pathogen-derived epitopes. Metabolically 14N and 15N-labeled P1.7-2,4 preps were isolated and mixed 1:1 (protein mass) before pulsing monocyte-derived HLA-DRB*0101 homozygous dendritic cells for 48 hr. MHC peptides were acid-eluted after HLA-DR pull-down, fractionated and analysed using nanoLC-EThcD- MS-MS. Mass spectra of P1.7-2,4-derived peptides were algorithm-searched based on their 14N/15N-ion doublets. Candidate ions were sequenced by using EThcD. Linear and spliced peptides were tested ASVG+114 Da and IGNYTQINAASVG+270 Da were confirmed, present in full 14N-and 15N-labeled form, indicating in-cis transpeptidation. The improved sequence coverage enabled by using EThcD identified the in cis -spliced peptides as IGNYTQINAASVG, C-terminally extended with Gly-Gly or Gly- Gly-Arg. Both linear and in cis -spliced region 8 peptides stimulated clone JS-63 in an HLA-DR-restricted manner. This is the first evidence for the formation of pathogen-derived in cis -spliced epitopes that are recognized by human CD4+ T cells. The mechanism behind the in cis transpeptidation remains to be elucidated, although the overall P1.7-2,4 sequence may inform a protease specificity. Further studies are ongoing to evaluate the biological consequences of these findings.

Effects of Graphene Oxide and Reduced Graphene Oxide Nanostructures on CD4+ Th2 Lymphocytes.

The activation of T helper (Th) lymphocytes is necessary for the adaptive immune response as they contribute to the stimulation of B cells (for the secretion of antibodies) and macrophages (for phagocytosis and destruction of pathogens) and are necessary for cytotoxic T-cell activation to kill infected target cells. For these issues, Th lymphocytes must be converted into Th effector cells after their stimulation through their surface receptors TCR/CD3 (by binding to peptide-major histocompatibility complex localized on antigen-presenting cells) and the CD4 co-receptor. After stimulation, Th cells proliferate and differentiate into subpopulations, like Th1, Th2 or Th17, with different functions during the adaptative immune response. Due to the central role of the activation of Th lymphocytes for an accurate adaptative immune response and considering recent preclinical advances in the use of nanomaterials to enhance T-cell therapy, we evaluated in vitro the effects of graphene oxide (GO) and two types of reduced GO (rGO15 and rGO30) nanostructures on the Th2 lymphocyte cell line SR.D10. This cell line offers the possibility of studying their activation threshold by employing soluble antibodies against TCR/CD3 and against CD4, as well as the simultaneous activation of these two receptors. In the present study, the effects of GO, rGO15 and rGO30 on the activation/proliferation rate of these Th2 lymphocytes have been analyzed by studying cell viability, cell cycle phases, intracellular content of reactive oxygen species (ROS) and cytokine secretion. High lymphocyte viability values were obtained after treatment with these nanostructures, as well as increased proliferation in the presence of rGOs. Moreover, rGO15 treatment decreased the intracellular ROS content of Th2 cells in all stimulated conditions. The analysis of these parameters showed that the presence of these GO and rGO nanostructures did not alter the response of Th2 lymphocytes.

Covalent TCR-peptide-MHC interactions induce T cell activation and redirect T cell fate in the thymus

Interactions between a T cell receptor (TCR) and a peptide-major histocompatibility complex (pMHC) ligand are typically mediated by noncovalent bonds. By studying T cells expressing natural or engineered TCRs, here we describe covalent TCR-pMHC interactions that involve a cysteine-cysteine disulfide bond between the TCR and the peptide. By introducing cysteines into a known TCR-pMHC combination, we demonstrate that disulfide bond formation does not require structural rearrangement of the TCR or the peptide. We further show these disulfide bonds still form even when the initial affinity of the TCR-pMHC interaction is low. Accordingly, TCR-peptide disulfide bonds facilitate T cell activation by pMHC ligands with a wide spectrum of affinities for the TCR. Physiologically, this mechanism induces strong Zap70-dependent TCR signaling, which triggers T cell deletion or agonist selection in the thymus cortex. Covalent TCR-pMHC interactions may thus underlie a physiological T cell activation mechanism that has applications in basic immunology and potentially in immunotherapy.

A Review of Suggested Mechanisms of MHC Odor Signaling.

Simple SummaryImmune genes of the vertebrate MHC vary among individuals. Each individual collection is optimally diverse to provide resistance against some infectious diseases but not too diverse to cause autoimmune diseases. MHC-dependent mate choice aims for optimally complementary MHC alleles. Each potential partner signals through body odor his/her MHC alleles. Identifying the signal molecules was a long-lasting puzzle solved only recently after many deviations as described. Commensal microbiota which are controlled by the individual MHC genes differ among individuals. They were suspected repeatedly to provide the signal, though mice raised germ-free could still smell MHC genes. Carrier hypotheses came in various versions, centered around the specificity of each MHC molecule for binding peptides from diseases, shown to T lymphocytes to induce the immune response. Volatiles of various origins were suggested to fill the place of the peptide and thus reflect the identity of the MHC molecule. Finally, the bound peptides themselves were identified as the sought info-chemicals. Synthesized peptides affect mate choice as predicted. Specific olfactory neurons were shown to react to these peptides but only to the anchors that define the binding specificity. Even eggs choose sperm to produce offspring with optimal MHC, though the signaling pathway needs further research.Although an individual’s mix of MHC immune genes determines its resistance, finding MHC-dependent mate choice occurred by accident in inbred mice. Inbred mice prefer MHC dissimilar mates, even when the choice was restricted to urine. It took decades to find the info-chemicals, which have to be as polymorphic as the MHC. Microbiota were suggested repeatedly as the origin of the odor signal though germ-free mice maintained normal preference. Different versions of the ‘carrier hypothesis’ suggested MHC molecules carry volatiles after the bound peptide is released. Theory predicted an optimal individual MHC diversity to maximize resistance. The optimally complementary mate should be and is preferred as several studies show. Thus, the odor signal needs to transmit the exact information of the sender’s MHC alleles, as do MHC ligand peptides but not microbiota. The ‘MHC peptide hypothesis’ assumes that olfactory perception of the peptide ligand provides information about the MHC protein in a key-lock fashion. Olfactory neurons react only to the anchors of synthesized MHC peptides, which reflect the binding MHC molecule’s identity. Synthesized peptides supplemented to a male’s signal affect choice in the predicted way, however, not when anchors are mutated. Also, the human brain detects smelled synthesized self-peptides as such. After mate choice, the lottery of meiosis of randomly paired oocyte and sperm haplotypes would often produce MHC non-optimal offspring. In sticklebacks, eggs select MHC-compatible sperm, thus prefer the best combination close to the population optimum.

T-cell-engaging antibodies for the treatment of solid tumors: challenges and opportunities.

Purpose of reviewT-cell-engaging antibodies or T-cell engagers (TCEs) can connect a patient's cytotoxic T cells with cancer cells, leading to potent redirected lysis. Until very recently, only one TCE was approved, the CD19/CD3-bispecific blinatumomab. Many new TCEs in late-stage clinical development target various hematopoietic lineage markers like CD20, BCMA, or CD123. Although very compelling single-agent activity of TCEs was observed with various blood-borne cancers, therapy of solid tumor indications has thus far been less successful.Recent findingsThe approval in 2022 of the gp100 peptide-major histocompatibility complex (MHC)/CD3 bispecific TCE tebentafusp in uveal melanoma confirms that TCEs can also efficiently work against solid tumors. TCEs targeting peptide–MHC complexes will expand the target space for solid tumor therapy to intracellular targets. Likewise, early clinical trial data from TCEs targeting DLL3 in small cell lunger cancer showed promising antitumor activity. Various technologies for conditional activation of TCEs in the tumor microenvironment (TME) may expand the scope of conventional surface targets that suffer from a narrow therapeutic window. Finally, pharmacological enhancements for TCE therapies by engagement of certain costimulatory receptors and cytokines, or blockade of checkpoints, are showing promise.SummaryTargeting peptide–MHC complexes, conditional TCE technologies, and concepts enhancing TCE-activated T cells are paving the way towards overcoming challenges associated with solid tumor therapy.

Physical and in silico immunopeptidomic profiling of a cancer antigen prostatic acid phosphatase reveals targets enabling TCR isolation

Tissue-specific antigens can serve as targets for adoptive T cell transfer-based cancer immunotherapy. Recognition of tumor by T cells is mediated by interaction between peptide-major histocompatibility complexes (pMHCs) and T cell receptors (TCRs). Revealing the identity of peptides bound to MHC is critical in discovering cognate TCRs and predicting potential toxicity. We performed multimodal immunopeptidomic analyses for human prostatic acid phosphatase (PAP), a well-recognized tissue antigen. Three physical methods, including mild acid elution, coimmunoprecipitation, and secreted MHC precipitation, were used to capture a thorough signature of PAP on HLA-A*02:01. Eleven PAP peptides that are potentially A*02:01-restricted were identified, including five predicted strong binders by NetMHCpan 4.0. Peripheral blood mononuclear cells (PBMCs) from more than 20 healthy donors were screened with the PAP peptides. Seven cognate TCRs were isolated which can recognize three distinct epitopes when expressed in PBMCs. One TCR shows reactivity toward cell lines expressing both full-length PAP and HLA-A*02:01. Our results show that a combined multimodal immunopeptidomic approach is productive in revealing target peptides and defining the cloned TCR sequences reactive with prostatic acid phosphatase epitopes.

Identification of MHC Class II Immunopeptidomes from Shigella flexneri 2a-infected Macrophages as Potential Vaccine Candidates

BACKGROUND: Shigella is a Gram-negative rod-shaped intracellular bacterial pathogen that causes bacterial dysentery or shigellosis among children under five years old. Antibiotics have been less effective in treating shigellosis due to the multi-drug resistance of Shigella. Therefore, an effective vaccine is urgently needed to prevent this disease. The present study aims to determine the peptides presented by major histocompatibility complex (MHC) class II molecules of Shigella-infected macrophages using mass spectrometry-based immunopeptidomics approaches. The MHC class II-associated peptides derived from Shigella-infected macrophages are candidates for developing subunit-based Shigella vaccine.METHODS: THP-1-derived macrophages were infected with Shigella flexneri 2a at the multiplicity of infection equal to 10. The lysate was immunoprecipitated and analyzed by liquid chromatography–tandem mass spectrometry (LC-MS/MS). The sequences retrieved were analyzed using bioinformatics tools.RESULTS: The Shigella-infected THP-1-derived macrophages contained sample peptides from source proteins of almost all subcellular localizations. Eight peptides from S. flexneri 2a-infected macrophages were predicted to be localized at the outer membrane proteins (OMPs) of S. flexneri 2a by the PSORTb server. Two of the OMP-associated peptides were predicted as antigenic, non-allergenic, and non-toxic by respective bioinformatics tools.CONCLUSION: The findings in this study showed two selected OMPs have great potential for vaccine development against shigellosis.KEYWORDS: immunopeptidomics, mass spectrometry, vaccine development, Shigella, MHC peptides

A molecular dynamics investigation of N-glycosylation effects on T-cell receptor kinetics

The binding interaction between the T-cell receptor (TCR) and peptide-major histocompatibility complex (pMHC) is modulated by several factors (known and unknown), however, investigations into effects of glycosylation are limited. A fully glycosylated computational model of the TCR bound to the pMHC is developed to investigate the effects of glycosylation on dissociation kinetics from the pMHC. Here, we examine the effects of N-glycosylation on TCR-pMHC bond strength using steered molecular dynamic simulations. N-glycosylation is a post-translational modification that adds sugar moieties to molecules and can modulate the activity of several immune molecules. Using a TCR-pMHC pair found in melanoma as a case study, our study demonstrates that N-glycosylation of the TCR-pMHC alters the proteins’ conformation; increases the bond lifetime; and increases the number of hydrogen bonds and Lennard-Jones Contacts involved in the TCR-pMHC bond. We find that weak glycan-protein or glycan-glycan interactions impact the equilibrated structure of the TCR and pMHC leading to an increase in the overall bond strength of the TCR-pMHC complex including the duration and energetic strength under constant load. These results indicate that N-glycosylation plays an important role in the TCR-pMHC bond and should be considered in future computational and experimental studies. Communicated by Ramaswamy H. Sarma

A Simple and Robust Single-Step Method for CAR-Vδ1 γδT Cell Expansion and Transduction for Cancer Immunotherapy.

The γδT cell subset of peripheral lymphocytes exhibits potent cancer antigen recognition independent of classical peptide MHC complexes, making it an attractive candidate for allogeneic cancer adoptive immunotherapy. The Vδ1-T cell receptor (TCR)-expressing subset of peripheral γδT cells has remained enigmatic compared to its more prevalent Vγ9Vδ2-TCR and αβ-TCR-expressing counterparts. It took until 2021 before a first patient was dosed with an allogeneic adoptive Vδ1 cell product despite pre-clinical promise for oncology indications stretching back to the 1980s. A contributing factor to the paucity of clinical progress with Vδ1 cells is the lack of robust, consistent and GMP-compatible expansion protocols. Herein we describe a reproducible one-step, clinically translatable protocol for Vδ1-γδT cell expansion from peripheral blood mononuclear cells (PBMCs), that is further compatible with high-efficiency gene engineering for immunotherapy purposes. Briefly, αβTCR- and CD56-depleted PBMC stimulation with known-in-the-art T cell stimulators, anti-CD3 mAb (clone: OKT-3) and IL-15, leads to robust Vδ1 cell expansion of high purity and innate-like anti-tumor efficacy. These Vδ1 cells can be virally transduced to express chimeric antigen receptors (CARs) using standard techniques, and the CAR-Vδ1 exhibit antigen-specific persistence, cytotoxicity and produce IFN-γ. Practicable, GMP-compatible engineered Vδ1 cell expansion methods will be crucial to the wide-spread clinical testing of these cells for oncology indications.

Naive and memory T cells TCR-HLA-binding prediction.

T cells recognize antigens through the interaction of their T cell receptor (TCR) with a peptide-major histocompatibility complex (pMHC) molecule. Following thymic-positive selection, TCRs in peripheral naive T cells are expected to bind MHC alleles of the host. Peripheral clonal selection is expected to further increase the frequency of antigen-specific TCRs that bind to the host MHC alleles. To check for a systematic preference for MHC-binding T cellsin TCR repertoires, we developed Natural Language Processing-based methods to predict TCR-MHC binding independently of the peptide presented for Class I MHC alleles. We trained a classifier on published TCR-pMHC binding pairs and obtained a high area under curve (AUC) of over 0.90 on the test set. However, when applied to TCR repertoires, the accuracy of the classifier dropped. We thus developed a two-stage prediction model, based on large-scale naive and memory TCR repertoires, denoted TCR HLA-binding predictor (CLAIRE). Since each host carries multiple human leukocyte antigen (HLA) alleles, we first computed whether a TCR on a CD8 T cell binds an MHC from any of the host Class-I HLA alleles. We then performed an iteration, where we predict the binding with the most probable allele from the first round. We show that this classifier is more precise for memory than for naïve cells. Moreover, it can be transferred between datasets. Finally, we developed a CD4-CD8 T cell classifier to apply CLAIRE to unsorted bulk sequencing datasets and showed a high AUC of 0.96 and 0.90 on large datasets. CLAIRE is available through a GitHub at: https://github.com/louzounlab/CLAIRE, and as a server at: https://claire.math.biu.ac.il/Home.

Opening opportunities for Kd determination and screening of MHC peptide complexes

An essential element of adaptive immunity is selective binding of peptide antigens by major histocompatibility complex (MHC) class I proteins and their presentation to cytotoxic T lymphocytes. Using native mass spectrometry, we analyze the binding of peptides to an empty disulfide-stabilized HLA-A*02:01 molecule and, due to its unique stability, we determine binding affinities of complexes loaded with truncated or charge-reduced peptides. We find that the two anchor positions can be stabilized independently, and we further analyze the contribution of additional amino acid positions to the binding strength. As a complement to computational prediction tools, our method estimates binding strength of even low-affinity peptides to MHC class I complexes quickly and efficiently. It has huge potential to eliminate binding affinity biases and thus accelerate drug discovery in infectious diseases, autoimmunity, vaccine design, and cancer immunotherapy.

Tim-3 mediates T cell trogocytosis to limit antitumor immunity.

T cell immunoglobulin mucin domain-containing protein 3 (Tim-3) negatively regulates innate and adaptive immunity in cancer. To identify the mechanisms of Tim-3 in cancer immunity, we evaluated the effects of Tim-3 blockade in human and mouse melanoma. Here, we show that human programmed cell death 1–positive (PD-1+) Tim-3+CD8+ tumor-infiltrating lymphocytes (TILs) upregulate phosphatidylserine (PS), a receptor for Tim-3, and acquire cell surface myeloid markers from antigen-presenting cells (APCs) through transfer of membrane fragments called trogocytosis. Tim-3 blockade acted on Tim-3+ APCs in a PS-dependent fashion to disrupt the trogocytosis of activated tumor antigen–specific CD8+ T cells and PD-1+Tim-3+ CD8+ TILs isolated from patients with melanoma. Tim-3 and PD-1 blockades cooperated to disrupt trogocytosis of CD8+ TILs in 2 melanoma mouse models, decreasing tumor burden and prolonging survival. Deleting Tim-3 in dendritic cells but not in CD8+ T cells impeded the trogocytosis of CD8+ TILs in vivo. Trogocytosed CD8+ T cells presented tumor peptide–major histocompatibility complexes and became the target of fratricide T cell killing, which was reversed by Tim-3 blockade. Our findings have uncovered a mechanism Tim-3 uses to limit antitumor immunity.

High-resolution profiling of MHC II peptide presentation capacity reveals SARS-CoV-2 CD4 T cell targets and mechanisms of immune escape.

Understanding peptide presentation by specific MHC alleles is fundamental for controlling physiological functions of T cells and harnessing them for therapeutic use. However, commonly used in silico predictions and mass spectroscopy have their limitations in precision, sensitivity, and throughput, particularly for MHC class II. Here, we present MEDi, a novel mammalian epitope display that allows an unbiased, affordable, high-resolution mapping of MHC peptide presentation capacity. Our platform provides a detailed picture by testing every antigen-derived peptide and is scalable to all the MHC II alleles. Given the urgent need to understand immune evasion for formulating effective responses to threats such as SARS-CoV-2, we provide a comprehensive analysis of the presentability of all SARS-CoV-2 peptides in the context of several HLA class II alleles. We show that several mutations arising in viral strains expanding globally resulted in reduced peptide presentability by multiple HLA class II alleles, while some increased it, suggesting alteration of MHC II presentation landscapes as a possible immune escape mechanism.