CD74 supports accumulation and function of regulatory T cells in tumors.

Regulatory T cells (Tregs) are plastic cells playing a pivotal role in the maintenance of immune homeostasis. Tregs actively adapt to the microenvironment where they reside; as a consequence, their molecular and functional profiles differ among tissues and pathologies. In tumors, the features acquired by Tregs remains poorly characterized. Here, we observe that human tumor-infiltrating Tregs selectively overexpress CD74, the MHC class II invariant chain. CD74 has been previously described as a regulator of antigen-presenting cell biology, however its function in Tregs remains unknown. CD74 genetic deletion in human primary Tregs reveals that CD74KO Tregs exhibit major defects in the organization of their actin cytoskeleton and intracellular organelles. Additionally, intratumoral CD74KO Tregs show a decreased activation, a drop in Foxp3 expression, a low accumulation in the tumor, and consistently, they are associated with accelerated tumor rejection in preclinical models in female mice. These observations are unique to tumor conditions as, at steady state, CD74KO-Treg phenotype, survival, and suppressive capacity are unaffected in vitro and in vivo. CD74 therefore emerges as a specific regulator of tumor-infiltrating Tregs and as a target to interfere with Treg anti-tumor activity.

Structural insights into human MHC-II association with invariant chain.

The loading of processed peptides on to major histocompatibility complex II (MHC-II) molecules for recognition by T cells is vital to cell-mediated adaptive immunity. As part of this process, MHC-II associates with the invariant chain (Ii) during biosynthesis in the endoplasmic reticulum to prevent premature peptide loading and to serve as a scaffold for subsequent proteolytic processing into MHC-II-CLIP. Cryo-electron microscopy structures of full-length Human Leukocyte Antigen-DR (HLA-DR) and HLA-DQ complexes associated with Ii, resolved at 3.0 to 3.1 Å, elucidate the trimeric assembly of the HLA/Ii complex and define atomic-level interactions between HLA, Ii transmembrane domains, loop domains, and class II-associated invariant chain peptides (CLIP). Together with previous structures of MHC-II peptide loading intermediates DO and DM, our findings complete the structural path governing class II antigen presentation.

Immunohistochemical study of CD74 biomarker in normal and malignant breast tissues.

OBJECTIVE: Aim: To detect the role of CD74 expression in breast carcinoma as a predictive marker for identifying the biological behavior of malignancy in Iraqi women.. PATIENTS AND METHODS: Materials and Methods: The study used technique of immunohistochemistry for detection CD74 protein role in breast cancer, and its expression in breast cancer tissue samples. Samples were collected in Al-Najaf city in Iraq, from Al-Forat Al-Awsat Oncology Center. The study was achieved at the Laboratories of the Faculty of Science in the University of Kufa. Fifty samples of breast cancer tissue, and twenty controls benign tissue were included in the study. The study has investigated relationship between expression of biomarker with grade, age of patient and tumor size. RESULTS: Results: The study showed that the cytoplasmic expression of CD74 with more clear and intensive staining in the cytoplasm, and reported that CD74 positivity rate was 52%. A significant association between CD74 expression and grade and size of tumor, so CD74 can be considered as a biomarker for prediction of breast cancer in women. No association was found between CD74 expression and each of patients' age and node metastasis. CONCLUSION: Conclusions: The study represents an important step in our region because there are a few studies about this topic; more efforts are required to approve the function of this biomarker.

MIF-Modulated Spinal Proteins Associated with Persistent Bladder Pain: A Proteomics Study.

Bladder pain is a prominent symptom in Interstitial Cystitis/Bladder Pain Syndrome (IC/BPS). We studied spinal mechanisms of bladder pain in mice using a model where repeated activation of intravesical Protease Activated Receptor-4 (PAR4) results in persistent bladder hyperalgesia (BHA) with little or no bladder inflammation. Persistent BHA is mediated by spinal macrophage migration inhibitory factor (MIF), and is associated with changes in lumbosacral proteomics. We investigated the contribution of individual spinal MIF receptors to persistent bladder pain as well as the spinal proteomics changes associated with relief of persistent BHA by spinal MIF antagonism. Female mice with persistent BHA received either intrathecal (i.t.) MIF monoclonal antibodies (mAb) or mouse IgG1 (isotype control antibody). MIF antagonism temporarily reversed persistent BHA (peak effect: 2 h), while control IgG1 had no effect. Moreover, i.t. antagonism of the MIF receptors CD74 and C-X-C chemokine receptor type 4 (CXCR4) partially reversed persistent BHA. For proteomics experiments, four separate groups of mice received either repeated intravesical scrambled peptide and sham i.t. injection (control, no pain group) or repeated intravesical PAR4 and: sham i.t.; isotype IgG1 i.t. (15 µg); or MIF mAb (15 µg). L6-S1 spinal segments were excised 2 h post-injection and examined for proteomics changes using LC-MS/MS. Unbiased proteomics analysis identified and relatively quantified 6739 proteins. We selected proteins that showed significant changes compared to control (no pain group) after intravesical PAR4 (sham or IgG i.t. treatment) and showed no significant change after i.t. MIF antagonism. Six proteins decreased during persistent BHA (V-set transmembrane domain-containing protein 2-like confirmed by immunohistochemistry), while two proteins increased. Spinal MIF antagonism reversed protein changes. Therefore, spinal MIF and MIF receptors mediate persistent BHA and changes in specific spinal proteins. These novel MIF-modulated spinal proteins represent possible new targets to disrupt spinal mechanisms that mediate persistent bladder pain.

Indirect CD4+ T cell protection against mouse gamma-herpesvirus infection via interferon gamma.

CD4+ T cells play a key role in γ-herpesvirus infection control. However, the mechanisms involved are unclear. Murine herpesvirus type 4 (MuHV-4) allows relevant immune pathways to be dissected experimentally in mice. In the lungs, it colonizes myeloid cells, which can express MHC class II (MHCII), and type 1 alveolar epithelial cells (AEC1), which lack it. Nevertheless, CD4+ T cells can control AEC1 infection, and this control depends on MHCII expression in myeloid cells. Interferon-gamma (IFNγ) is a major component of CD4+ T cell-dependent MuHV-4 control. Here, we show that the action of IFNγ is also indirect, as CD4+ T cell-mediated control of AEC1 infection depended on IFNγ receptor (IFNγR1) expression in CD11c+ cells. Indirect control also depended on natural killer (NK) cells. Together, the data suggest that the activation of MHCII+ CD11c+ antigen-presenting cells is key to the CD4+ T cell/NK cell protection axis. By contrast, CD8+ T cell control of AEC1 infection appeared to operate independently. IMPORTANCE: CD4+ T cells are critical for the control of gamma-herpesvirus infection; they act indirectly, by recruiting natural killer (NK) cells to attack infected target cells. Here, we report that the CD4+ T cell/NK cell axis of gamma-herpesvirus control requires interferon-γ engagement of CD11c+ dendritic cells. This mechanism of CD4+ T cell control releases the need for the direct engagement of CD4+ T cells with virus-infected cells and may be a common strategy for host control of immune-evasive pathogens.

MHCII-peptide presentation: an assessment of the state-of-the-art prediction methods.

Major histocompatibility complex Class II (MHCII) proteins initiate and regulate immune responses by presentation of antigenic peptides to CD4+ T-cells and self-restriction. The interactions between MHCII and peptides determine the specificity of the immune response and are crucial in immunotherapy and cancer vaccine design. With the ever-increasing amount of MHCII-peptide binding data available, many computational approaches have been developed for MHCII-peptide interaction prediction over the last decade. There is thus an urgent need to provide an up-to-date overview and assessment of these newly developed computational methods. To benchmark the prediction performance of these methods, we constructed an independent dataset containing binding and non-binding peptides to 20 human MHCII protein allotypes from the Immune Epitope Database, covering DP, DR and DQ alleles. After collecting 11 known predictors up to January 2022, we evaluated those available through a webserver or standalone packages on this independent dataset. The benchmarking results show that MixMHC2pred and NetMHCIIpan-4.1 achieve the best performance among all predictors. In general, newly developed methods perform better than older ones due to the rapid expansion of data on which they are trained and the development of deep learning algorithms. Our manuscript not only draws a full picture of the state-of-art of MHCII-peptide binding prediction, but also guides researchers in the choice among the different predictors. More importantly, it will inspire biomedical researchers in both academia and industry for the future developments in this field.

Differences in F pocket impact on HLA I genetic associations with autoimmune diabetes.

Introduction: Human leukocyte antigen (HLA) I molecules present antigenic peptides to activate CD8+ T cells. Type 1 Diabetes (T1D) is an auto-immune disease caused by aberrant activation of the CD8+ T cells that destroy insulin-producing pancreatic ß cells. Some HLA I alleles were shown to increase the risk of T1D (T1D-predisposing alleles), while some reduce this risk (T1D-protective alleles). Methods: Here, we compared the T1D-predisposing and T1D-protective allotypes concerning peptide binding, maturation, localization and surface expression and correlated it with their sequences and energetic profiles using experimental and computational methods. Results: T1D-predisposing allotypes had more peptide-bound forms and higher plasma membrane levels than T1D-protective allotypes. This was related to the fact that position 116 within the F pocket was more conserved and made more optimal contacts with the neighboring residues in T1D-predisposing allotypes than in protective allotypes. Conclusion: Our work uncovers that specific polymorphisms in HLA I molecules potentially influence their susceptibility to T1D.

Selection, engineering, and in vivo testing of a human leukocyte antigen-independent T-cell receptor recognizing human mesothelin.

BACKGROUND: Canonical α/ß T-cell receptors (TCRs) bind to human leukocyte antigen (HLA) displaying antigenic peptides to elicit T cell-mediated cytotoxicity. TCR-engineered T-cell immunotherapies targeting cancer-specific peptide-HLA complexes (pHLA) are generating exciting clinical responses, but owing to HLA restriction they are only able to target a subset of antigen-positive patients. More recently, evidence has been published indicating that naturally occurring α/ß TCRs can target cell surface proteins other than pHLA, which would address the challenges of HLA restriction. In this proof-of-concept study, we sought to identify and engineer so-called HLA-independent TCRs (HiTs) against the tumor-associated antigen mesothelin. METHODS: Using phage display, we identified a HiT that bound well to mesothelin, which when expressed in primary T cells, caused activation and cytotoxicity. We subsequently engineered this HiT to modulate the T-cell response to varying levels of mesothelin on the cell surface. RESULTS: The isolated HiT shows cytotoxic activity and demonstrates killing of both mesothelin-expressing cell lines and patient-derived xenograft models. Additionally, we demonstrated that HiT-transduced T cells do not require CD4 or CD8 co-receptors and, unlike a TCR fusion construct, are not inhibited by soluble mesothelin. Finally, we showed that HiT-transduced T cells are highly efficacious in vivo, completely eradicating xenografted human solid tumors. CONCLUSION: HiTs can be isolated from fully human TCR-displaying phage libraries against cell surface-expressed antigens. HiTs are able to fully activate primary T cells both in vivo and in vitro. HiTs may enable the efficacy seen with pHLA-targeting TCRs in solid tumors to be translated to cell surface antigens.

Antigen presentation plays positive roles in the regenerative response to cardiac injury in zebrafish.

In contrast to adult mammals, adult zebrafish can fully regenerate injured cardiac tissue, and this regeneration process requires an adequate and tightly controlled immune response. However, which components of the immune response are required during regeneration is unclear. Here, we report positive roles for the antigen presentation-adaptive immunity axis during zebrafish cardiac regeneration. We find that following the initial innate immune response, activated endocardial cells (EdCs), as well as immune cells, start expressing antigen presentation genes. We also observe that T helper cells, a.k.a. Cd4+ T cells, lie in close physical proximity to these antigen-presenting EdCs. We targeted Major Histocompatibility Complex (MHC) class II antigen presentation by generating cd74a; cd74b mutants, which display a defective immune response. In these mutants, Cd4+ T cells and activated EdCs fail to efficiently populate the injured tissue and EdC proliferation is significantly decreased. cd74a; cd74b mutants exhibit additional defects in cardiac regeneration including reduced cardiomyocyte dedifferentiation and proliferation. Notably, Cd74 also becomes activated in neonatal mouse EdCs following cardiac injury. Altogether, these findings point to positive roles for antigen presentation during cardiac regeneration, potentially involving interactions between activated EdCs, classical antigen-presenting cells, and Cd4+ T cells.

Blocking the MIF-CD74 axis augments radiotherapy efficacy for brain metastasis in NSCLC via synergistically promoting microglia M1 polarization.

BACKGROUND: Brain metastasis is one of the main causes of recurrence and death in non-small cell lung cancer (NSCLC). Although radiotherapy is the main local therapy for brain metastasis, it is inevitable that some cancer cells become resistant to radiation. Microglia, as macrophages colonized in the brain, play an important role in the tumor microenvironment. Radiotherapy could activate microglia to polarize into both the M1 and M2 phenotypes. Therefore, searching for crosstalk molecules within the microenvironment that can specifically regulate the polarization of microglia is a potential strategy for improving radiation resistance. METHODS: We used databases to detect the expression of MIF in NSCLC and its relationship with prognosis. We analyzed the effects of targeted blockade of the MIF/CD74 axis on the polarization and function of microglia during radiotherapy using flow cytometry. The mouse model of brain metastasis was used to assess the effect of targeted blockade of MIF/CD74 axis on the growth of brain metastasis. RESULT: Our findings reveals that the macrophage migration inhibitory factor (MIF) was highly expressed in NSCLC and is associated with the prognosis of NSCLC. Mechanistically, we demonstrated CD74 inhibition reversed radiation-induced AKT phosphorylation in microglia and promoted the M1 polarization in combination of radiation. Additionally, blocking the MIF-CD74 interaction between NSCLC and microglia promoted microglia M1 polarization. Furthermore, radiation improved tumor hypoxia to decrease HIF-1α dependent MIF secretion by NSCLC. MIF inhibition enhanced radiosensitivity for brain metastasis via synergistically promoting microglia M1 polarization in vivo. CONCLUSIONS: Our study revealed that targeting the MIF-CD74 axis promoted microglia M1 polarization and synergized with radiotherapy for brain metastasis in NSCLC.

Upregulation of HLA-II related to LAG-3+CD4+ T cell infiltration is associated with patient outcome in human glioblastoma.

Glioblastoma (GBM) is the most common malignant diffuse glioma of the brain. Although immunotherapy with immune checkpoint inhibitors (ICIs), such as programmed cell death protein (PD)-1/PD ligand-1 inhibitors, has revolutionized the treatment of several cancers, the clinical benefit in GBM patients has been limited. Lymphocyte-activation gene 3 (LAG-3) binding to human leukocyte antigen-II (HLA-II) plays an essential role in triggering CD4+ T cell exhaustion and could interfere with the efficiency of anti-PD-1 treatment; however, the value of LAG-3-HLA-II interactions in ICI immunotherapy for GBM patients has not yet been analyzed. Therefore, we aimed to investigate the expression and regulation of HLA-II in human GBM samples and the correlation with LAG-3+CD4+ T cell infiltration. Human leukocyte antigen-II was highly expressed in GBM and correlated with increased LAG-3+CD4+ T cell infiltration in the stroma. Additionally, HLA-IIHighLAG-3High was associated with worse patient survival. Increased interleukin-10 (IL-10) expression was observed in GBM, which was correlated with high levels of HLA-II and LAG-3+ T cell infiltration in stroma. HLA-IIHighIL-10High GBM associated with LAG-3+ T cells infiltration synergistically showed shorter overall survival in patients. Combined anti-LAG-3 and anti-IL-10 treatment inhibited tumor growth in a mouse brain GL261 tumor model. In vitro, CD68+ macrophages upregulated HLA-II expression in GBM cells through tumor necrosis factor-α (TNF-α). Blocking TNF-α-dependent inflammation inhibited tumor growth in a mouse GBM model. In summary, T cell-tumor cell interactions, such as LAG-3-HLA-II, could confer an immunosuppressive environment in human GBM, leading to poor prognosis in patients. Therefore, targeting the LAG-3-HLA-II interaction could be beneficial in ICI immunotherapy to improve the clinical outcome of GBM patients.

PAD4 controls tumor immunity via restraining the MHC class II machinery in macrophages.

Tumor-associated macrophages (TAMs) shape tumor immunity and therapeutic efficacy. However, it is poorly understood whether and how post-translational modifications (PTMs) intrinsically affect the phenotype and function of TAMs. Here, we reveal that peptidylarginine deiminase 4 (PAD4) exhibits the highest expression among common PTM enzymes in TAMs and negatively correlates with the clinical response to immune checkpoint blockade. Genetic and pharmacological inhibition of PAD4 in macrophages prevents tumor progression in tumor-bearing mouse models, accompanied by an increase in macrophage major histocompatibility complex (MHC) class II expression and T cell effector function. Mechanistically, PAD4 citrullinates STAT1 at arginine 121, thereby promoting the interaction between STAT1 and protein inhibitor of activated STAT1 (PIAS1), and the loss of PAD4 abolishes this interaction, ablating the inhibitory role of PIAS1 in the expression of MHC class II machinery in macrophages and enhancing T cell activation. Thus, the PAD4-STAT1-PIAS1 axis is an immune restriction mechanism in macrophages and may serve as a cancer immunotherapy target.

Major Histocompatibility Complex II Expression on Oral Langerhans Cells Differentially Regulates Mucosal CD4 and CD8 T Cells.

In murine periodontitis, the T helper (Th)17 response against Porphyromonas gingivalis in cervical lymph node is abrogated by diphtheria toxin-driven depletion of Langerhans cells (LCs). We determined the impact of major histocompatibility complex class II (MHC-II) presentation in LCs on Th17 cells in the oral mucosa of mice. Using an established human-Langerin promoter-Cre mouse model, we generated LC-specific deletion of the H2-Ab1 (MHC-II) gene. MHC-II expression was ablated in 81.2% of oral-resident LCs compared with >99% of skin-resident LCs. MHC-II (LCΔMHC-II) depletion did not reduce the number of CD4 T cells nor the frequency of Th17 cells compared with that in wild-type mice. However, the frequencies of Th1 cells decreased, and Helios+ T-regulatory cells increased. In ligature-induced periodontitis, the numbers of CD4 T cells and Th17 cells were similar in LCΔMHC-II and wild-type mice. Normal numbers of Th17 cells can therefore be sustained by as little as 18.8% of MHC-II-expressing LCs in oral mucosa. Unexpectedly, oral mucosa CD8 T cells increased >25-fold in LCΔMHC-II mice. Hence, these residual MHC-II-expressing LCs appear unable to suppress the local expansion of CD8 T cells while sufficient to sustain a homeostatic CD4 T-cell response. Reducing the expression of MHC-II on specific LC subpopulations may ultimately boost CD8-mediated intraepithelial surveillance at mucosal surfaces.

Accurate modeling of peptide-MHC structures with AlphaFold.

Major histocompatibility complex (MHC) proteins present peptides on the cell surface for T cell surveillance. Reliable in silico prediction of which peptides would be presented and which T cell receptors would recognize them is an important problem in structural immunology. Here, we introduce an AlphaFold-based pipeline for predicting the three-dimensional structures of peptide-MHC complexes for class I and class II MHC molecules. Our method demonstrates high accuracy, outperforming existing tools in class I modeling accuracy and class II peptide register prediction. We validate its performance and utility with new experimental data on a recently described cancer neoantigen/wild-type peptide pair and explore applications toward improving peptide-MHC binding prediction.

Ubiquitination of Major Histocompatibility Complex II Changes Its Immunological Recognition Structure.

Ubiquitination is a process that dictates the lifespan of major histocompatibility complex class II (MHC II)/peptide complexes on antigen-presenting cells. This process is tightly controlled by the levels of ubiquitin ligases, and disruptions in the turnover of MHC II can lead to the improper development of CD4+ T cells within the thymus and hinder the formation of regulatory T cells in the peripheral tissue. To investigate the underlying mechanisms, we utilized dendritic cells lacking the Membrane-associated RING-CH (MARCH) I ubiquitin ligase. We discovered that the overexpression of MARCH I decreases the interaction with LAG-3. Moreover, the MHC II molecules tethered with ubiquitin also showed diminished binding to LAG-3. We employed Diffracted X-ray Blinking (DXB), a technique used for single-molecule X-ray imaging, to observe the protein movements on live cells in real time. Our observations indicated that the normal MHC II molecules moved more rapidly across the cell surface compared to those on the MARCH I-deficient dendritic cells or MHC II KR mutants, which is likely a result of ubiquitination. These findings suggest that the signaling from ubiquitinated MHC II to the T cell receptor differs from the non-ubiquitinated forms. It appears that ubiquitinated MHC II might not be quickly internalized, but rather presents antigens to the T cells, leading to a range of significant immunological responses.

IL-27p28 specifically regulates MHC II expression in macrophages through CIITA.

Antigen-presenting cells (APCs) constantly express major histocompatibility complex II (MHC II), including macrophages and dendritic cells (DCs) which deliver antigens to CD4+ T cells and play an important role in adaptive immunity. The expression of MHC II is controlled by the transcriptional coactivator CIITA. Interleukin-27 (IL-27), a newly discovered IL-12 family cytokine, is composed of p28 and EBI3 subunits. In this study, we used IL-27p28 conditional knock-out mice to investigate the regulatory effects of IL-27p28 on macrophage polarization and the expression of MHC II in macrophages. We found that MHC II expression was upregulated in the bone marrow-derived and peritoneal exudate macrophages (BMDMs; PEMs) from IL-27p28-deficient mice, with their inflammation regulating function unaffected. We also demonstrated that in the APCs, IL-27p28 selectively regulated MHC II expression in macrophages but not in dendritic cells. During Pseudomonas aeruginosa (P. aeruginosa) reinfection, higher survival rate, bacterial clearance, and ratio of CD4+/CD8+ T cells in the spleen during the specific immune phase were observed in IL-27p28 defect mice, as well as an increased MHC II expression in alveolar macrophages (AMs). But these did not occur in the first infection. For the first time we discovered that IL-27p28 specifically regulates the expression of MHC II in macrophages by regulating CIITA, while its absence enhances antigen presentation and adaptive immunity against P. aeruginosa.

Vaccination with post-translational modified, homocitrullinated peptides induces CD8 T-cell responses that mediate antitumor immunity.

BACKGROUND: Post-translational modification of proteins has the potential to alter the ability of T cells to recognize major histocompatibility complex (MHC) class -I and class-II restricted antigens, thereby resulting in altered immune responses. One such modification is carbamylation (homocitrullination) that results in the formation of homocitrulline (Hcit) residues in a non-enzymatic reaction of cyanate with the lysine residues in the polypeptide chain. Homocitrullination occurs in the tumor microenvironment and CD4-mediated immune responses to Hcit epitopes can target stressed tumor cells and provide a potent antitumor response in mouse models. METHODS: Homocitrullinated peptides were identified and assessed in vitro for HLA-A2 binding and in vivo in human leukocyte antigen (HLA) transgenic mouse models for immunogenicity. CD8 responses were assessed in vitro for cytotoxicity and in vivo tumor therapy. Human tumor samples were analyzed by targeted mass spectrometry for presence of homocitrullinated peptides. RESULTS: Homocitrullinated peptides from aldolase and cytokeratin were identified, that stimulated CD8-mediated responses in vivo. Modified peptides showed enhanced binding to HLA-A2 compared with the native sequences and immunization of HLA-A2 transgenic mice generated high avidity modification specific CD8 responses that killed peptide expressing target cells. Importantly, in vivo the homocitrullinated aldolase specific response was associated with efficient CD8 dependent antitumor therapy of the aggressive murine B16 tumor model indicating that this epitope is naturally presented in the tumor. In addition, the homocitrullinated aldolase epitope was also detected in human tumor samples. CONCLUSION: This is the first evidence that homocitrullinated peptides can be processed and presented via MHC-I and targeted for tumor therapy. Thus, Hcit-specific CD8 T-cell responses have potential in the development of future anticancer therapy.

Induction of broad multifunctional CD8+ and CD4+ T cells by hepatitis B virus antigen-based synthetic long peptides ex vivo.

Introduction: Therapeutic vaccination based on synthetic long peptides (SLP®) containing both CD4+ and CD8+ T cell epitopes is a promising treatment strategy for chronic hepatitis B infection (cHBV). Methods: We designed SLPs for three HBV proteins, HBcAg and the non-secreted proteins polymerase and X, and investigated their ability to induce T cell responses ex vivo. A set of 17 SLPs was constructed based on viral protein conservation, functionality, predicted and validated binders for prevalent human leukocyte antigen (HLA) supertypes, validated HLA I epitopes, and chemical producibility. Results: All 17 SLPs were capable of inducing interferon gamma (IFNÉ£) production in samples from four or more donors that had resolved an HBV infection in the past (resolver). Further analysis of the best performing SLPs demonstrated activation of both CD8+ and CD4+ multi-functional T cells in one or more resolver and patient sample(s). When investigating which SLP could activate HBV-specific T cells, the responses could be traced back to different peptides for each patient or resolver. Discussion: This indicates that a large population of subjects with different HLA types can be covered by selecting a suitable mix of SLPs for therapeutic vaccine design. In conclusion, we designed a set of SLPs capable of inducing multifunctional CD8+ and CD4+ T cells ex vivo that create important components for a novel therapeutic vaccine to cure cHBV.

CCK2-receptor deficiency impairs immune balance by influencing CD4+ T cells development by inhibiting cortical-thymic-epithelial-cells.

Immune balance is crucial for an organism's survival and is inseparable from the regulation of the nervous system. Accumulating evidence indicates that cholecystokinin (CCK) plays an important role in mediating the immune response through the activation of cholecystokinin receptors (CCKRs). However, it remains unclear whether CCKRs deficiency may impair immune balance. Here, we showed that CCK2R-deficient adult mice were immunocompromised and had an increased risk of shock and even death in an endotoxemia (ETM)/endotoxin shock (ES) model. In addition, in both adult and juvenile mice, CCK2R deficiency not only influenced the development of CD4 single-positive (SP) thymocytes in thymic positive selection but also decreased the population of CD3+ CD4+ T cells in the spleen. More importantly, CCK2R deficiency inhibited the expression of major histocompatibility complex class II (MHC II) and CD83 on cortical thymic epithelial cells (cTECs) in juvenile and adult mice. Overall, our study suggests that CCK2R is essential for maintaining CD4+ T cell development in the thymus and reveals that CCK2R plays an important role in maintaining immune balance.

The ligand-dependent suppression of TCR signaling by the immune checkpoint receptor LAG3 depends on the cytoplasmic RRFSALE motif.

Lymphocyte activation gene 3 (LAG3) is an inhibitory immune checkpoint receptor that restrains autoimmune and antitumor responses, but its evolutionarily conserved cytoplasmic tail lacks classical inhibitory motifs. Major histocompatibility complex class II (MHC class II) is an established LAG3 ligand, and fibrinogen-like protein 1 (FGL1), lymph node sinusoidal endothelial cell C-type lectin (LSECtin), and Galectin-3 have been proposed as alternative binding partners that play important roles in LAG3 function. Here, we used a fluorescent human T cell reporter system to study the function of LAG3. We found that LAG3 reduced the response to T cell receptor stimulation in the presence of MHC class II molecules to a lesser extent compared with the receptor programmed cell death protein 1. Analysis of deletion mutants demonstrated that the RRFSALE motif in the cytoplasmic tail of LAG3 was necessary and sufficient for LAG3-mediated inhibition. In this system, FGL1, but not LSECtin or Galectin-3, acted as a LAG3 ligand that weakly induced inhibition. LAG3-blocking antibodies attenuated LAG3-mediated inhibition in our reporter cells and enhanced reporter cell activation even in the absence of LAG3 ligands, indicating that they could potentially enhance T cell responses independently of their blocking effect.