Data on MPXV-specific T-cell responses at the single-peptide level remain limited in patients with mpox and in MVA-BN vaccinees. Here, we characterized the breadth and specificity of MPXV-specific T-cell responses at the single-peptide level after in vitro expansion with overlapping 20-mer peptides spanning the MPXV proteins H3L, A35R, and B6R. The study included 28 adult males: 15 with a history of mpox (including 6 with additional MVA-BN vaccination), 7 MVA-BN-only vaccinees, and 6 unexposed participants. All mpox and MVA-BN participants responded to at least one H3L peptide, indicating broad immunogenicity, while responses to A35R and B6R were more common in the mpox group. Notably, the breadth of B6R-specific CD4+ T-cell responses correlated with hybrid immunity (r = 0.6; p = 0.02). Interestingly, MVA-BN and mpox individuals demonstrated distinct immunodominant patterns: H3L_251-270 and H3L_211-230 were mainly recognized among mpox individuals, whereas MVA-BN recognized H3L_221-240 more frequently. High-affinity HLA binding to multiple H3L peptides suggests broad population coverage. Additional immunogenetic analysis revealed a shared TRBV15 clonotype in about 50% of mpox cases. In summary, these findings highlight H3L as a potential vaccine target, guiding the development of next-generation multi-antigen MPXV vaccines to elicit comprehensive T-cell immunity.

Human leukocyte antigen (HLA) class I molecules are highly polymorphic, restricting peptide binding to narrow sequence subsets. Designing peptides that bind multiple HLA supertypes-termed superbinders-offers a promising strategy for broad-spectrum T cell vaccines and immunotherapies. Here, we present superHLA, a computational framework that combines Markov Chain Monte Carlo optimization with state-of-the-art major histocompatibility complex binding predictors to design synthetic 9-mer peptides with broad HLA-binding profiles. Using superHLA, we generated over 190,000 candidate superbinders predicted to bind 8 to 12 HLA class I alleles across distinct supertypes. A multitier filtering pipeline-incorporating sequence clustering, synthesis feasibility, cross-predictor validation, and self-peptidome exclusion-yielded a final panel of 100 peptides for experimental testing. Of these, 21 bound 4 to 9 supertypes in vitro. Superbinders displayed distinct anchor residue preferences and showed minimal similarity to human peptides. These results suggest that HLA superbinders are more abundant than previously recognized and can be rationally designed at scale. This approach supports development of pan-HLA immunogens with broad population coverage and may inform applications in vaccine research, neoantigen discovery, and immunotherapy.

BACKGROUND: Atherosclerosis is a chronic inflammatory disease with a strong autoimmune component, marked by the detection of autoreactive T cells and autoantibodies. Recent single-cell RNA sequencing studies have shown that atherosclerotic plaques contain clonally expanded CD8 + T cells. One of the known atherosclerosis autoantigens is APOB (apolipoprotein B). However, autoreactive CD8 + T cells to APOB in humans have not been described. METHODS: We studied CD8 + T-cell reactivity to human leukocyte antigen-A*02:01-restricted APOB epitopes, starting with in silico epitope prediction. We used peripheral blood mononuclear cells from human leukocyte antigen-A02:01+ healthy subjects to test the top 64-ranked peptides for their potential to elicit a CD8 + T-cell response. Antigen-specific responses were assessed using activation-induced marker assays, intracellular cytokine staining, and IFNγ (interferon gamma) ELISpot assays. RESULTS: Some APOB peptides triggered robust CD8 + T-cell activation with effector memory features, and expression of cytokines and cytotoxic molecules. Five immunodominant epitopes spanning 2 APOB regions accounted for most of the response and elicited significant T-cell activation in healthy donors that was increased in clinical samples from patients with severe coronary artery disease. CONCLUSIONS: The discovery of immunodominant major histocompatibility complex class I-restricted APOB epitopes suggests a new perspective for immune-based interventions to mitigate atherosclerosis.

Abstract Introduction Atherosclerosis, contributing to ∼one-third of global deaths, is a chronic inflammatory disorder with autoimmune features. Atherosclerosis related antigen - apolipoprotein B (APOB) specific human CD4 T cells have been characterized, and CD8 T cell infiltration and oligoclonal expansion have been observed in human atherosclerotic lesions. Despite studies in mouse atherosclerosis models, APOB-specific CD8 T cells remain unexplored in humans. Methods Through in silico analysis, ∼0.1% of top-ranked APOB peptides predicted to bind HLA-A02:01 were selected. Using APOB peptide mesopools, PBMCs from HLA-A02:01+ donors underwent expansion-based restimulation, followed by flow cytometry. Responses to individual peptides were assessed via IFNγ ELISpot. Six immunodominant peptides were used to generate tetramers for tracking APOB-specific CD8+ T cells in human PBMCs. APOB tetramer-specific CD8 T cells from atherosclerosis patients were analyzed using single-cell RNA and TCR sequencing. Results APOB peptide stimulation induced the expression of activation markers (4-1BB, CD69, and CD25), proinflammatory cytokines (IFNγ and TNFα), and cytotoxic molecules (granzyme B and perforin) and enriched for effector memory phenotype in CD8 T cells. APOB epitopes that elicited the strongest IFNγ ELISpot response were mapped to 2 major antigenic regions in APOB. Tetramers detected APOB-specific circulating CD8 T cells in healthy humans and atherosclerosis patients. APOB-specific CD8 T cells from atherosclerotic humans were activated, as evidenced by high CD45RO and PD-1 expression. APOB-specific CD8 T cells had higher clonal expansion and lower diversity as compared to APOB non-specific CD8 T cells. Conclusion Human APOB harbors dominant HLA-I-restricted epitopes that trigger autoreactive CD8 T responses. Future assessment of transcriptomic and TCR data, followed by functional assays, will help characterize the nature and role of APOB-specific CD8 T cells in atherosclerosis. Funding Source n/a Topic Categories Immune Mechanisms of Human Disease (HUM)

T cell immunity has a crucial role in vaccine-induced protection against respiratory viruses, yet a detailed characterization of T cell responses and epitopes in Syrian hamsters, a highly utilized preclinical, small animal model for SARS-CoV-2 research, is lacking. In this study, using an intranasal Chimpanzee adenoviral vectored vaccine (ChAd-SARS-CoV-2-S), we characterized the T cell response to the spike protein of SARS-CoV-2 in Syrian hamsters and identified immunogenic CD4+ and CD8+ T cell epitopes using IFN-γ ELISpot assays and cell depletions. The mucosal ChAd-SARS-CoV-2-S vaccine elicited strong T cell responses, with evidence of CD4+ and CD8+ T cell activation in both lymphoid and mucosal tissues. Responses were directed toward the non-receptor-binding domain regions of the spike protein, indicating that dominant T cell epitopes for hamsters reside elsewhere in this structural protein. Six different T cell epitopes (4 for CD4 and 2 for CD8) were identified in the spike protein, and epitope-specific responses were detected in hamsters from 2 vendors, suggesting genetic similarity in terms of major histocompatibility complex allele expression. Identifying T cell epitopes and characterizing T cell responses in lymphoid and mucosal compartments enhances the utility of Syrian hamsters as a preclinical model for SARS-CoV-2 vaccine studies.

Human CD8+ T cells undergo significant metabolic and transcriptional shifts during activation and differentiation. The orphan nuclear receptor Nur77 plays a role in modulating these processes and has been linked to T cell dysfunction. However, few studies have addressed its role in the memory potential and functionality of human CD8+ T cells. Here, we evaluated the expression of Nur77 in human CD8+ T cells, focusing on its relationship with their differentiation profile and functionality. Our findings indicate that Nur77 is associated with an early-differentiated, T cell factor 1+ (TCF-1+) memory-like phenotype in both total and virus-specific human CD8+ T cells across contexts of acute resolved or chronic viral infection and vaccination. Nur77 expression was associated with cytokine polyfunctionality and increased proliferative capacity in long-lived antigen-responsive cells. Moreover, the modulation of Nur77 activity in vitro enhanced the functionality of chronic human immunodeficiency virus (HIV) and hepatitis B virus (HBV)-specific CD8+ T cells. These results suggest that Nur77 is associated with polyfunctional properties in virus-specific CD8+ T cell responses in humans. In addition, this study provides insights into novel strategies for enhancing CD8+ T cell functionality in settings of chronic antigen stimulation.

The SARS-CoV-2 pandemic has highlighted the need for society, as a whole, to be prepared against potential pandemics caused by a variety of different viral families of concern. Here, we describe a roadmap towards the identification and validation of conserved T cell epitope regions from Viral Families of Pandemic Potential (VFPP). For each viral family, we select a prototype virus, the sequence of which could be utilized in epitope identification screens. Examples of viral families considered and their respective prototypes (species/ subspecies) are Coronaviridae (Severe Acute Respiratory Syndrome-related Coronavirus/ SARS-CoV-2), Flaviviridae (Dengue virus/DENV2), Togaviridae (Chikungunya virus/ CHIKV), Paramyixoviridae (Morbillivirus/measles), Arenaviridae (Mammarenavirus/ Lassa), and Picornaviridae (Enterovirus C/poliovirus). The peptide sequences encoded in each prototype virus are then analyzed to determine their conservation across different viral taxonomic groups and viral variants derived from each of the VFPP. We outline available methodologies for epitope discovery based on panels of overlapping peptides and bioinformatics- based predictions of HLA-peptide binding, along with high-throughput in vitro assays, with emphasis on addressing coverage of the general worldwide population. Validation can be achieved by a variety of methodologies, including determining HLA restriction and recognition in samples from volunteers convalescent from previous infections or immunized with approved or experimental vaccines, and immunophenotyping of responding T cells. The capacity of these regions to induce crossreactive T cell responses can be tested experimentally with homologous peptides derived from the various viral species of interest. Importantly, they could be considered as a component of pan-viral family vaccines. Conversely, immunogenic regions that are highly specific to a given virus could be of interest for diagnostic applications.

Abstract Introduction Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disorder characterized by progressive motor neuron loss and eventual paralysis. While autoimmune mechanisms have long been suspected to contribute to ALS pathology, disease-associated antigen targets have remained elusive. Methods We analyzed if ALS was associated with increased autoreactive T cell responses towards TDP-43, SOD1, and C9orf72, compared to healthy controls. Peptide megapools consisting of 15 amino acid peptides overlapping by 10 residues spanning the entire sequence was synthesized for each protein. Fluorospot assays were used to measure T cell responses to the peptide pools by measuring release of interferon-γ (IFNγ), interleukins 5 (IL-5) and 10 (IL-10). Results We demonstrate that the ALS-associated protein C9orf72 is a major antigenic target in ALS. Epitope mapping revealed multiple immunogenic regions across the C9orf72 protein. While the reactivity was found broadly in ALS subjects, the responses were particularly high ALS donors who carried the C9orf72 hexanucleotide repeat expansion mutation in the non-coding regions of the gene. Importantly, IL-10-mediated responses were significantly higher in individuals with longer predicted survival, suggesting a regulatory and potentially protective influence on disease progression. Conclusion Together, these findings identify the first defined target of autoreactive T cell responses in ALS, and support the hypothesis that disease trajectory is in part shaped by the balance of inflammatory and counter-inflammatory T cell activity. Funding Source Supported by LJI & Kyowa Kirin, Inc. (KKNA- Kyowa Kirin North America), the Swedish Research Council (salary for E.J., grant references 2024-00175), the Freedom Together Foundation (to D.S.), and in part by the Intramural Research Program, NINDS, NIH (A. Topic Categories Neuroimmunology (NEUR)

Gene fusions (GFs) are critical events in pediatric oncology, often serving as oncogenic drivers. However, fusion proteins and their derived neoantigens (GF-NEOs) remain underexplored for targeted immunotherapy. We developed ProteoFusioNEO, a computational tool for the in silico translation of transcriptomic data, analyzing 5,190 pediatric patients with cancer and 935 cell lines, yielding 382 and 446 fusion proteins. We highlight that GFs generate multiple translational outcomes, with 97% being in-frame in patients. Fusion junctions exhibit the sequence motif [KQE][DG], which partly reflects the nature of exon-exon junctions, albeit with additional hydrophilicity. Moreover, GF-NEOs' abundance may be shaped by the motif, offering insights into fusion protein biology. Finally, a multipronged validation strategy using in vitro and in vivo systems confirms the GF-NEOs presentation through mass spectrometry-based proteomics and immunopeptidomics. Multiple GF-NEOs encoded by two versions of the ETV6-RUNX1 fusion were validated, paving the way for targeted immunotherapy development.

A role of the immune system in Parkinson's disease (PD) progression has long been suspected due to the increased frequency of activated glial cells and infiltrating T cells in the substantia nigra. It was previously reported that PD donors have increased T cell responses towards PINK1 and α-synuclein (α-syn), two Lewy body-associated proteins. Further, T cell reactivity towards α-syn was highest closer to disease onset, highlighting that autoreactive T cells might play a role in PD pathogenesis. However, whether T cell autoreactivity is present during prodromal PD is unknown. Here, we investigated T cell responses towards PINK1 and α-syn in donors at high risk of developing PD (i.e. prodromal PD: genetic risk, hyposmia, and or REM sleep behavior disorder), in comparison to PD and healthy control donors. T cell reactivity to these two autoantigens was detected in prodromal PD at levels comparable to those detected in individuals with clinically diagnosed PD. Aligned with the increased incidence of PD in males, we found that males with PD, but not females, had elevated T cell reactivity compared to healthy controls. However, among prodromal PD donors, males and females had elevated T cell responses. These differing trends in reactivity highlights the need for further studies of the impact of biological sex on neuroinflammation and PD progression.

Amyotrophic lateral sclerosis (ALS), like many neurodegenerative disorders, features cytoplasmic inclusion of proteins, with the most prominent component identified being TDP-43. Neuroinflammation is also apparent in affected tissues, including elevated T cell infiltration and enhanced expression of HLA Class I and II by microglia. However, the role of T cells in ALS is still largely unknown. We have previously found that a subset of individuals with Parkinson’s disease (PD) have increased frequencies of T cells recognizing alpha-synuclein, a protein found in Lewy body aggregates in PD patients. We therefore hypothesized that autoreactive T cells could also play a role in ALS pathology. We generated 15-mer peptide pools spanning the three ALS-related proteins TDP-43, SOD1, and C9orf72. Cytokine production (IFNγ, IL-5, IL-10) against these proteins was measured by the fluorospot assay, using PBMCs collected from ALS patients and age- and sex-matched healthy controls (HCs). Intriguingly, we found that ALS patient have significantly elevated T cell responses towards C9orf72 compared to HCs, particularly IL-5 mediated T cell responses. We further found IL-5-mediated responses to be associated with higher ALS Functional Rating Scale (ALSFRS), raising the possibility that these responses exert a regulatory and protective influence on disease manifestations. Importantly, these are the first findings of autoreactive T cells playing a role in ALS pathology. Supported by Kyowa Kirin, Inc. (KKNA-Kyowa Kirin North America) Neuroimmunology (NEUR)

Parkinson's disease (PD) is the second most prevalent neurodegenerative disorder. While there is no curative treatment, the immune system's involvement with autoimmune T cells that recognize the protein alpha-synuclein (alpha-syn) in a subset of individuals suggests new areas for therapeutic strategies. As not all patients with PD have T cells specific for alpha-syn, we explored additional autoantigenic targets of T cells in PD. We generated 15-mer peptides spanning several PD-related proteins implicated in PD pathology, including glucosylceramidase beta 1 (GBA), superoxide dismutase 1 (SOD1), PTEN induced kinase 1 (PINK1), Parkin RBR E3 ubiquitin protein ligase (parkin), oxoglutarate dehydrogenase (OGDH), and leucine rich repeat kinase 2 (LRRK2). Cytokine production (IFN-gamma, IL-5, IL-10) against these proteins was measured using a fluorospot assay and PBMCs from patients with PD and age-matched healthy controls. We identified PINK1, a regulator of mitochondrial stability, as an autoantigen targeted by T cells, as well as its unique epitopes, and their HLA restriction. The PINK1-specific T cell reactivity revealed sex-based differences, as it was predominantly found in male patients with PD, which may contribute to the heterogeneity of PD. Identifying and characterizing PINK1 and other autoinflammatory targets may lead to antigen-specific diagnostics, progression markers, and/or novel therapeutic strategies for PD.

Background Peptide vaccines offer a direct way to initiate an immunogenic response to a defined antigen epitope. However, peptide vaccines are unstable in vivo, subject to rapid enzymatic proteolysis. Replacement of an α-amino acid residue with a homologous β-amino acid residue (native side chain, but backbone extended by a single CH2 unit) impairs proteolysis at nearby amide bonds. Therefore, antigen analogues containing α-to-β replacements have been examined for functional mimicry of native all-α antigens. Another group previously took this approach in the ovalbumin (OVA) antigen model by evaluating single α-to-β analogues of the murine major histocompatibility complex (MHC) I-restricted peptide SIINFEKL.Methods We re-examined this set of α/β SIINFEKL antigens. We tested the susceptibility to proteolysis in mouse serum and their ability to activate OVA-antigen-specific CD8 T cells in vitro. Additionally, we tested the α/β antigens in vivo for their ability to induce an antigen-specific immunogenic response in naïve mice and in OVA-expressing tumor-bearing mice.Results The α/β antigens were comparable to the native antigen in their susceptibility to proteolysis in serum. Each α/β antigen was capable of activating antigen-specific CD8 T cells in vitro. However, antigen-specific CD8 T cells induced against α/β antigens in vivo were not cross-reactive to the native antigen. Moreover, immunization with α/β analogues did not elicit anti-tumor effects in tumor-bearing mice.Conclusions We conclude that even though α/β analogues of the SIINFEKL antigen can elicit a T cell-based response, this class of backbone-modified peptides is not promising from the perspective of antitumor vaccine development.

Introduction:No vaccine is currently licensed against human visceral leishmaniasis (VL), a fatal CD4+ T cell immunosupressive disease against which chemotherapy is reduced to a few toxic drugs. The NH36 nucleoside hydrolase is a DNA metabolism vital enzyme present in all Leishmania species. A vaccine based on such a conserved antigen could protect against both VL and cutaneous leishmaniasis, whose epidemics geographically overlap. Increased frequencies of NH36-specific IL-2+TNF-α+IFN-γ+-producing CD4+ T cells were associated with VL immune protection. Methods:the sequences of HLA-Class I and Class II T cell epitopes were predicted in the NH36 peptide sequence using the Tepitope, Propred, IEDB and NetMHCpan EL 4.1 immune informatic tools. The epitopes were synthetized and used to study their reactivity with sera samples, and to stimulate the in vitro response of PBMC of human patients cured from VL, asymptomatic individuals and healthy blood donors of a non-endemic area. Cytokine production was studied intracellularly by flow cytometry (ICS) and cytokine secretion was measured in PBMC supernatants. The HLA typing of DNA patients and the analysis of epitope conservancy in the Leishmania genus were obtained. Two recombinant multiepitope proteins were designed, cloned in E. coli, expressed, purified and used for in vitro stimulation of PBMC of VL cured and asymptomatic patients. Results:We identified in silico fifteen NH36 conserved epitopes that correspond to promiscuous binders of HLA-DR, -DQ, -DP class II molecules, as well as HLA-A, B and C class I molecules. Collectively, these epitopes provide high worldwide population coverage of both class I and II alleles, and bound to alleles associated with VL susceptibility and resistance. VL asymptomatic individuals showed maximal frequencies of CD4+ and CD8+ multifunctional IL-2+TNF-α+IFN-γ+-producing T lymphocytes in response to these epitopes, with secretion of TNF-α, IL-1β and IL-6. Two recombinant multiepitope vaccines were designed using these epitopes linked by AAA or GPGPG spacers. Both proteins promoted CD4+ and CD8+ T cell responses in PBMC of VL cured and asymptomatic individuals. Discussion:Both MultiAAA and MultiGPGPG proteins could be potentially used for universal human vaccination against leishmaniasis.

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease characterized by a progressive loss of motor neurons. Neuroinflammation is apparent in affected tissues, including increased T cell infiltration and activation of microglia, particularly in the spinal cord1,2. Autoimmune responses are thought to have a key role in ALS pathology, and it is hypothesized that T cells contribute to the rapid loss of neurons during disease progression3,4. However, until now there has been no reported target for such an autoimmune response. Here we show that ALS is associated with recognition of the C9orf72 antigen, and we map the specific epitopes that are recognized. We show that these responses are mediated by CD4+ T cells that preferentially release IL-5 and IL-10, and that IL-10-mediated T cell responses are significantly greater in donors who have a longer predicted survival time. Our results reinforce the previous hypothesis that neuroinflammation has an important role in ALS disease progression, possibly because of a disrupted balance of inflammatory and counter-inflammatory T cell responses4. These findings highlight the potential of therapeutic strategies aimed at enhancing regulatory T cells5, and identify a key target for antigen-specific T cell responses that could enable precision therapeutics in ALS.

The COVID-19 pandemic highlighted the critical need for vaccine strategies capable of addressing emerging viral threats. Betacoronaviruses, including severe acute respiratory syndrome coronavirus (SARS-CoV), Middle East respiratory syndrome (MERS), and SARS-CoV-2, present significant pandemic risks due to their zoonotic potential and genetic diversity. T cell-mediated immunity has demonstrated durable responses and strong cross-reactivity, offering a promising avenue for achieving broad immunity within a viral family. In this study, we combined comprehensive epitope mapping with sequence conservation analyses to identify conserved T cell epitope regions (CTERs), which constitute 12% of the complete SARS-CoV-2 proteome. We showed that SARS-CoV-2 CTER-specific T cells cross-reactively recognize sequences from multiple Betacoronavirus subgenera. Importantly, incorporating CTERs from non-spike proteins significantly enhanced T cell cross-reactivity potential and human leukocyte antigen (HLA) coverage compared with T cells targeting only spike proteins. Our findings lay the groundwork for a multi-antigen vaccine strategy that includes non-spike proteins to expand cross-reactive immunity across a broader spectrum of Betacoronaviruses.

ABSTRACT Frequent recent spillovers of subtype H5N1 clade 2.3.4.4b highly pathogenic avian influenza (HPAI) virus into poultry and mammals, especially dairy cattle, including several human cases, increased concerns over a possible future pandemic. Here, we performed an analysis of epitope data curated in the Immune Epitope Database (IEDB). We found that the patterns of immunodominance of seasonal influenza viruses circulating in humans and H5N1 are similar. We further conclude that a significant fraction of the T-cell epitopes is conserved at a level associated with cross-reactivity between avian and seasonal sequences, and we further experimentally demonstrate extensive cross-reactivity in the most dominant T-cell epitopes curated in the IEDB. Based on these observations, and the overall similarity of the neuraminidase (NA) N1 subtype encoded in both HPAI and seasonal H1N1 influenza virus as well as cross-reactive group 1 HA stalk-reactive antibodies, we expect that a degree of pre-existing immunity is present in the general human population that could blunt the severity of human H5N1 infections. IMPORTANCE Influenza A viruses (IAVs) cause pandemics that can result in millions of deaths. The highly pathogenic avian influenza (HPAI) virus of the H5N1 subtype is presently among the top viruses of pandemic concern, according to the WHO and the National Institute of Allergy and Infectious Diseases (NIAID). Previous exposure by infection and/or vaccination to a given IAV subtype or clade influences immune responses to a different subtype or clade. Analysis of human CD4 and CD8 T-cell epitope conservation between HPAI H5N1 and seasonal IAV sequences revealed levels of identity and conservation conducive to T cell cross-reactivity, suggesting that pre-existing T cell immune memory should, to a large extent, cross-recognize avian influenza viruses. This observation was experimentally verified by testing responses from human T cells to non-avian IAV and their HPAI H5N1 counterparts. Accordingly, should a more widespread HPAI H5N1 outbreak occur, we hypothesize that cross-reactive T-cell responses might be able to limit disease severity.

Type 1 diabetes (T1D) is an autoimmune disease involving T cell-mediated destruction of the insulin-producing beta cells in the pancreatic islets of Langerhans. CD8+ T cells, responding to beta cell peptides presented by class I major histocompatibility complex (MHC) molecules, are important effectors leading to beta cell elimination. Human leukocyte antigen (HLA) B∗39:06, B∗39:01, and B∗38:01 are closely related class I MHC allotypes that nonetheless show differential association with T1D. HLA-B∗39:06 is the most predisposing of all HLA class I molecules and is associated with early age at disease onset. B∗39:01 is also associated with susceptibility to T1D, but to a lesser extent, though differing from B∗39:06 by only two amino acids. HLA-B∗38:01, in contrast, is associated with protection from the disease. Upon identifying a peptide that binds to both HLA-B∗39:06 and B∗39:01, we determined the respective X-ray structures of the two allotypes presenting this peptide to 1.7 Å resolution. The peptide residues available for T cell receptor contact and those serving as anchors were identified. Analysis of the F pocket of HLA-B∗39:06 and B∗39:01 provided an explanation for the distinct peptide C terminus preferences of the two allotypes. Structure-based modeling of the protective HLA-B∗38:01 suggested a potential reason for its peptide preferences and its reduced propensity to present 8-mer peptides compared to B∗39:06. Notably, the three allotypes showed differential binding to peptides derived from beta cell autoantigens. Taken together, our findings should facilitate identification of disease-relevant candidate T cell epitopes and structure-guided therapeutics to interfere with peptide binding.