A dynamic CD2-rich compartment at the outer edge of the immunological synapse boosts and integrates signals.

The CD2-CD58 recognition system promotes adhesion and signaling and counters exhaustion in human T cells. We found that CD2 localized to the outer edge of the mature immunological synapse, with cellular or artificial APC, in a pattern we refer to as a 'CD2 corolla'. The corolla captured engaged CD28, ICOS, CD226 and SLAM-F1 co-stimulators. The corolla amplified active phosphorylated Src-family kinases (pSFK), LAT and PLC-γ over T cell receptor (TCR) alone. CD2-CD58 interactions in the corolla boosted signaling by 77% as compared with central CD2-CD58 interactions. Engaged PD-1 invaded the CD2 corolla and buffered CD2-mediated amplification of TCR signaling. CD2 numbers and motifs in its cytoplasmic tail controlled corolla formation. CD8+ tumor-infiltrating lymphocytes displayed low expression of CD2 in the majority of people with colorectal, endometrial or ovarian cancer. CD2 downregulation may attenuate antitumor T cell responses, with implications for checkpoint immunotherapies.

Cytomegalovirus Infection Facilitates the Costimulation of CD57+CD28- CD8 T Cells in HIV Infection and Atherosclerosis via the CD2-LFA-3 Axis.

CD8 T cells are emerging as important mediators in atherosclerosis and cardiovascular disease (CVD). Immune activation may play a particular role in people with HIV (PWH) who are at an increased risk of CVD, even after controlling for known CVD risk factors. Latent CMV infection is associated with increased CVD risk for both PWH and people without HIV, and human CMV-specific CD4 and CD8 T cells are enriched for an immunosenescent phenotype. We previously showed that CMV coinfection in PWH promotes vascular homing and activation of inflammatory CD4 T cells through the CD2-LFA-3 axis. However, the role of CD2/LFA3 costimulation of CD8 T cells in PWH with CMV has yet to be described. In the present study, we demonstrate that CD2 expression on CX3CR1+CD57+CD28- inflammescent CD8 T cells is increased on cells from CMV-seropositive PWH. In vitro CD2/LFA-3 costimulation enhances TCR-mediated activation of these inflammatory CD8 memory T cells. Finally, we show that LFA-3 is highly expressed in aortas of SIV-infected rhesus macaques and in atherosclerotic plaques of people without HIV. Our findings are consistent with a model in which CMV infection enhances CD2 expression on highly proinflammatory CD8 T cells that can then be stimulated by LFA-3 expressed in the vasculature, even in the absence of CD28 costimulation. This model, in which CMV infection exacerbates toxic cytokine and granzyme production by CD8 T cells within the vasculature, highlights a potential therapeutic target in atherosclerosis development and progression, especially for PWH.

rs10924104 in the expression enhancer motif of CD58 confers susceptibility to human autoimmune diseases.

CD58 plays roles in cell adhesion and co-stimulation with antigen presentation from major histocompatibility complex class II on antigen-presenting cells to T-cell antigen receptors on naïve T cells. CD58 reportedly contributes to the development of various human autoimmune diseases. Recently, genome-wide association studies (GWASs) identified CD58 as a susceptibility locus for autoimmune diseases such as systemic lupus erythematosus (SLE), multiple sclerosis (MS), and primary biliary cholangitis (PBC). However, the primary functional variant and molecular mechanisms of susceptibility to autoimmune diseases in the CD58 locus were not clarified. Here, rs10924104, located in the ZNF35-binding motif within the gene expression regulatory motif, was identified as the primary functional variant for SLE, MS, and PBC among genetic variants showing stronger linkage disequilibrium (LD) with GWAS-lead variants in the CD58 locus. Expression-quantitative trait locus (e-QTL) data for each distinct blood cell type and in vitro functional analysis using the CRISPR/Cas9 system corroborated the functional role of rs10924104 in the upregulation of CD58 transcription by the disease-risk allele. Additionally, the strength of disease susceptibility observed in the CD58 locus could be accounted for by the strength of LD between rs10924104 and each GWAS-lead variant. In conclusion, the present study demonstrated for the first time the existence of a shared autoimmune disease-related primary functional variant (i.e., rs10924104) that regulates the expression of CD58. Clarifying the molecular mechanism of disease susceptibility derived from such a shared genetic background is important for understanding human autoimmune diseases and human immunology.

CD58 acts as a tumor promotor in hepatocellular carcinoma via activating the AKT/GSK-3ß/ß-catenin pathway.

BACKGROUND: Hepatocellular carcinoma (HCC) is one of the most prevalent malignancies worldwide because of rapid progression and high incidence of metastasis or recurrence. Accumulating evidence shows that CD58-expressing tumor cell is implicated in development of various cancers. The present study aimed to reveal the functional significance of CD58 in HCC progression and the underlying mechanisms. METHODS: Immunohistochemical staining (IHC), and western blotting were used to detect the expression of CD58 in HCC tissues and cells. The levels of sCD58 (a soluble form of CD58) in the cell supernatants and serum were assessed by ELISA. CCK-8, colony formation, and xenograft assays were used to detect the function of CD58 on proliferation in vitro and in vivo. Transwell assay and sphere formation assay were performed to evaluate the effect of CD58 and sCD58 on metastasis and self-renewal ability of HCC cells. Western blotting, immunofluorescence (IF), TOP/FOP Flash reporter assay, and subcellular fractionation assay were conducted to investigate the molecular regulation between CD58/sCD58 and AKT/GSK-3ß/ß-catenin axis in HCC cells. RESULTS: CD58 was significantly upregulated in HCC tissues. Elevation of CD58 expression correlated with more satellite foci and vascular invasion, and poorer tumor-free and overall survival in HCC patients. Higher sCD58 levels were in HCC patients' serum compared to healthy individuals. Functionally, CD58 promotes the proliferation of HCC cells in vitro and in vivo. Meanwhile, CD58 and sCD58 induce metastasis, self-renewal and pluripotency in HCC cells in vitro. Mechanistically, CD58 activates the AKT/GSK-3ß/ß-catenin signaling pathway by increasing phosphorylation of AKT or GSK3ß signaling, promoting expression of Wnt/ß-catenin target proteins and TCF/LEF-mediated transcriptional activity. Furthermore, AKT activator SC-79 or inhibitor LY294002 abolished the inhibitory effect of CD58 silencing on the proliferation, metastasis, and stemness of HCC cells. CONCLUSIONS: Taken together, CD58 promotes HCC progression and metastasis via activating the AKT/GSK-3ß/ß-catenin pathway, suggesting that CD58 is a novel prognostic biomarker and therapeutic target for HCC.

Virtual Screening and Binding Analysis of Potential CD58 Inhibitors in Colorectal Cancer (CRC).

Human cell surface receptor CD58, also known as lymphocyte function-associated antigen 3 (LFA-3), plays a critical role in the early stages of immune response through interacting with CD2. Recent research identified CD58 as a surface marker of colorectal cancer (CRC), which can upregulate the Wnt pathway and promote self-renewal of colorectal tumor-initiating cells (CT-ICs) by degradation of Dickkopf 3. In addition, it was also shown that knockdown of CD58 significantly impaired tumor growth. In this study, we developed a structure-based virtual screening pipeline using Autodock Vina and binding analysis and identified a group of small molecular compounds having the potential to bind with CD58. Five of them significantly inhibited the growth of the SW620 cell line in the following in vitro studies. Their proposed binding models were further verified by molecular dynamics (MD) simulations, and some pharmaceutically relevant chemical and physical properties were predicted. The hits described in this work may be considered interesting leads or structures for the development of new and more efficient CD58 inhibitors.

Targeting protein-protein interaction for immunomodulation: A sunflower trypsin inhibitor analog peptidomimetic suppresses RA progression in CIA model.

Protein-protein interactions (PPI) of co-stimulatory molecules CD2-CD58 are important in the early stage of an immune response, and increased expression of these co-stimulatory molecules is observed in the synovial region of joints in rheumatoid arthritis (RA) patients. A CD2 epitope region that binds to CD58 was grafted on to sunflower trypsin inhibitor (SFTI) template structure to inhibit CD2-CD58 PPI. The peptide was incorporated with an organic moiety dibenzofuran (DBF) in its structure. The designed peptidomimetic was studied for its ability to inhibit CD2-CD58 interactions in vitro, and its thermal and enzymatic stability was evaluated. Stability studies indicated that the grafted peptidomimetic was stable against trypsin cleavage. In vivo studies using the collagen-induced arthritis (CIA) model in mice indicated that the peptidomimetic was able to slow down the progress of arthritis, an autoimmune disease in the mice model. These studies suggest that with the grafting of organic functional groups in the stable peptide template SFTI stabilizes the peptide structure, and these peptides can be used as a template to design stable peptides for therapeutic purposes.

Cytomegalovirus Coinfection Is Associated with Increased Vascular-Homing CD57+ CD4 T Cells in HIV Infection.

Cytotoxic CD4 T cells are linked to cardiovascular morbidities and accumulate in both HIV and CMV infections, both of which are associated with increased risk of cardiovascular disease (CVD). In this study, we identify CMV coinfection as a major driver of the cytotoxic phenotype, characterized by elevated CD57 expression and reduced CD28 expression, in circulating CD4 T cells from people living with HIV infection, and investigate potential mechanisms linking this cell population to CVD. We find that human CD57+ CD4 T cells express high levels of the costimulatory receptor CD2 and that CD2/LFA-3 costimulation results in a more robust and polyfunctional effector response to TCR signals, compared with CD28-mediated costimulation. CD57+ CD4 T cells also express the vascular endothelium-homing receptor CX3CR1 and migrate toward CX3CL1-expressing endothelial cells in vitro. IL-15 promotes the cytotoxic phenotype, elevates CX3CR1 expression, and enhances the trafficking of CD57+ CD4 T cells to endothelium and may therefore be important in linking these cells to cardiovascular complications. Finally, we demonstrate the presence of activated CD57+ CD4 T cells and expression of CX3CL1 and LFA-3 in atherosclerotic plaque tissues from HIV-uninfected donors. Our findings are consistent with a model in which cytotoxic CD4 T cells contribute to CVD in HIV/CMV coinfection and in atherosclerosis via CX3CR1-mediated trafficking and CD2/LFA-3-mediated costimulation. This study identifies several targets for therapeutic interventions and may help bridge the gap in understanding how CMV infection and immunity are linked to increased cardiovascular risk in people living with HIV infection.

A genetic variant associated with multiple sclerosis inversely affects the expression of CD58 and microRNA-548ac from the same gene.

Genome-wide association studies have identified more than 200 genetic variants to be associated with an increased risk of developing multiple sclerosis (MS). Still, little is known about the causal molecular mechanisms that underlie the genetic contribution to disease susceptibility. In this study, we investigated the role of the single-nucleotide polymorphism (SNP) rs1414273, which is located within the microRNA-548ac stem-loop sequence in the first intron of the CD58 gene. We conducted an expression quantitative trait locus (eQTL) analysis based on public RNA-sequencing and microarray data of blood-derived cells of more than 1000 subjects. Additionally, CD58 transcripts and mature hsa-miR-548ac molecules were measured using real-time PCR in peripheral blood samples of 32 MS patients. Cell culture experiments were performed to evaluate the efficiency of Drosha-mediated stem-loop processing dependent on genotype and to determine the target genes of this underexplored microRNA. Across different global populations and data sets, carriers of the MS risk allele showed reduced CD58 mRNA levels but increased hsa-miR-548ac levels. We provide evidence that the SNP rs1414273 might alter Drosha cleavage activity, thereby provoking partial uncoupling of CD58 gene expression and microRNA-548ac production from the shared primary transcript in immune cells. Moreover, the microRNA was found to regulate genes, which participate in inflammatory processes and in controlling the balance of protein folding and degradation. We thus uncovered new regulatory implications of the MS-associated haplotype of the CD58 gene locus, and we remind that paradoxical findings can be encountered in the analysis of eQTLs upon data aggregation. Our study illustrates that a better understanding of RNA processing events might help to establish the functional nature of genetic variants, which predispose to inflammatory and neurological diseases.

Endoplasmic Reticulum (ER) Reorganization and Intracellular Retention of CD58 Are Functionally Independent Properties of the Human Cytomegalovirus ER-Resident Glycoprotein UL148.

The human cytomegalovirus (HCMV) endoplasmic reticulum (ER)-resident glycoprotein UL148 is posited to play roles in immune evasion and regulation of viral cell tropism. UL148 prevents cell surface presentation of the immune cell costimulatory ligand CD58 while promoting maturation and virion incorporation of glycoprotein O, a receptor binding subunit for an envelope glycoprotein complex involved in entry. Meanwhile, UL148 activates the unfolded protein response (UPR) and causes large-scale reorganization of the ER. In order to determine whether the seemingly disparate effects of UL148 are related or discrete, we generated six charged cluster-to-alanine (CCTA) mutants within the UL148 ectodomain and compared them to wild-type UL148, both in the context of infection studies using recombinant viruses and in ectopic expression experiments, assaying for effects on ER remodeling and CD58 surface presentation. Two mutants, targeting charged clusters spanning residues 79 to 83 (CC3) and 133 to 136 (CC4), retained the potential to impede CD58 surface presentation. Of the six mutants, only CC3 retained the capacity to reorganize the ER, but it showed a partial phenotype. Wild-type UL148 accumulates in a detergent-insoluble form during infection. However, all six CCTA mutants were fully soluble, which implies a relationship between insolubility and organelle remodeling. Additionally, we found that the chimpanzee cytomegalovirus UL148 homolog suppresses surface presentation of CD58 but fails to reorganize the ER, while the homolog from rhesus cytomegalovirus shows neither activity. Collectively, our findings illustrate various degrees of functional divergence between homologous primate cytomegalovirus immunevasins and suggest that the capacity to cause ER reorganization is unique to HCMV UL148.IMPORTANCE In myriad examples, viral gene products cause striking effects on cells, such as activation of stress responses. It can be challenging to decipher how such effects contribute to the biological roles of the proteins. The HCMV glycoprotein UL148 retains CD58 within the ER, thereby preventing it from reaching the cell surface, where it functions to stimulate cell-mediated antiviral responses. Intriguingly, UL148 also triggers the formation of large, ER-derived membranous structures and activates the UPR, a set of signaling pathways involved in adaptation to ER stress. We demonstrate that the potential of UL148 to reorganize the ER and to retain CD58 are separable by mutagenesis and, possibly, by evolution, since chimpanzee cytomegalovirus UL148 retains CD58 but does not remodel the ER. Our findings imply that ER reorganization contributes to other roles of UL148, such as modulation of alternative viral glycoprotein complexes that govern the virus' ability to infect different cell types.

T Cell Activation Pathways: B7, LFA-3, and ICAM-1 Shape Unique T Cell Profiles.

Two signals are required for induction of cell proliferation and cytokine production in resting T cells. Occupancy of the T cell receptor by antigen/MHC complexes delivers the first signal to the T cell, while the second signal is provided by interaction with costimulatory ligands on APC. CD2, LFA-1, and CD28 are the major costimulatory and adhesive molecules on T cells and bind to the LFA-3, ICAM-1 and B7 ligands, respectively, on APC. LFA-3 plays a central role for naive and memory T helper cells during the early phase of an immune response. The LFA-3/CD2 pathway initiates strong antigen-independent cell adhesion, substantial expansion of naive T helper cells, and induction of large amounts of IFN-γ in memory cells. The release of IFN-γ may upregulate expression of ICAM-1 and B7 on APC and allows multiple adhesion pathways to amplify the immune response. The LFA- 1/ICAM-l pathway stimulates adhesion and cell proliferation more efficiently in memory T helper cells than in naive cells. Further, the results suggest that naive T helper cells express functionally inactive LFA-1 molecules on the cell surface, which may have a physiological role in keeping these cells in a resting state. B7 costimulation superinduces IL-2 production in both naive and memory T helper cells and generates long-lasting cell proliferation. This permits transition from an autocrine to a paracrine immune response. Coexpression of B7/LFA-3 provides an optimal APC function and enables a vigorous T cell response to minute amounts of antigen. AP-1 and NF-κB transcription factors are involved in the induction of several cytokine gene promoters and play a central role in the regulation of IL-2 gene transcription. LFA-3 costimulation only moderately enhances AP-1 DNA-binding activity and does not influence the NF-κB activity induced by TCR engagement, whereas B7 costimulation induces large amounts of NF-κB and AP-1 activity in T helper cells. The costimulatory ligands represent a family of adhesion molecules with considerable redundancy. Interfamily redundancy of LFA-3, B7, and ICAM ligands offers an opportunity to regulate distinct T cell response profiles in various microenvironments at separate time points of an immune response.

CD2-CD58 interactions are pivotal for the activation and function of adaptive natural killer cells in human cytomegalovirus infection.

The existence and expansion of adaptive NK-cell subsets have been linked to HCMV infection. Phenotypically, a majority of adaptive NK cells expresses the activating receptor NKG2C and CD57. Some of the molecular factors driving the expansion of NKG2C+ CD57+ NK cells in HCMV infection have been identified. The direct interaction of adaptive NK cells with HCMV-infected cells, preceding the expansion, however, remains less studied. Recently, adaptive NK cells were reported to express higher levels of the co-activating receptor CD2. We explored whether CD2 was directly involved in the response of adaptive NK cells to HCMV. In a co-culture system of human PBMCs and productively infected fibroblasts, we observed an upregulation of CD69, CD25, and HLA-DR on all NK cells. However, only in adaptive NK cells was this increase largely blocked by antibodies against CD2 and CD58. Functionally, this blockade also resulted in diminished production of IFN-γ and TNF-α by adaptive human NK cells in response to HCMV-infected cells. Our results demonstrate that binding of CD2 to upregulated CD58 on infected cells is a critical event for antibody-mediated activation and subsequent effector functions of adaptive NKG2C+ CD57+ NK cells during the antiviral response.

Alterations of the CD58 gene in classical Hodgkin lymphoma.

Immune evasion plays a central role in the pathophysiology of classical Hodgkin lymphoma (cHL). As mutations of the CD58 gene contribute to immune evasion of diffuse large B cell lymphoma tumor cells, we studied whether alterations of the CD58 gene also occur in Hodgkin and Reed/Sternberg (HRS) cells of cHL. Single nucleotide polymorphism chip analysis revealed homozygous deletions within the CD58 gene in two cHL cell lines (SUP-HD1 and U-HO1). Sequencing of the CD58 gene in seven cHL cell lines disclosed in addition a homozygous splice site mutation in cell line KM-H2. None of the three mutated lines expressed CD58 protein on their surface. Thus, three of seven cHL cell lines analyzed harbor destructive CD58 mutations. Molecular analysis of isolated HRS cells from 10 primary cases of cHL; however, did not reveal any case with a CD58 mutation. A FICTION study indicated heterozygous deletions of CD58 in 3 of 13 cHL analyzed. Overall, we report frequent inactivating mutations of CD58 in cHL cell lines, but their rare occurrence in primary HRS cells. As the three cHL cell lines with CD58 mutations were all established from HRS cells located in pleural effusions, i.e., outside the normal lymph node microenvironment, in end-stages of the disease, CD58 inactivation in cHL might be predominantly prevalent to such situations.

The novel immunotherapeutic molecule T11TS modulates glioma-induced changes of key components of the immunological synapse in favor of T cell activation and glioma abrogation.

T-cell-mediated immune responses are typically low in conditions of malignant glioma which has been known to cause marked immunesuppression and dysregulate major T-cell signaling molecules. Thus, T-cell-based immunotherapies are currently in vogue in the treatment of malignant glioma. The novel glycopeptide, T11TS/S-LFA-3/S-CD58 has previously been shown by our group to be highly efficacious in glioma abrogation in in vivo and in vitro conditions. This glycopeptide ligands to the costimulatory CD2 molecule on T-cells, causing profound immune stimulation leading to glioma abrogation, suggesting probable involvement of T11TS in modulation of the T-cell signaling pathway. The present study offers a multi-targeted approach towards repair of some of the key components of the immunological synapse at the T-cell-APC interface and is therefore the first of its kind to offer a holistic model of restoration of immunological synapse components so as to trigger T-cells towards activation against glioma. The study thus indicates that the totally dysregulated molecular events at the immunological synapse in glioma are restored back to normal levels with the administration of T11TS, which finally culminates in glioma abrogation. The present study thus delineates an important T-cell signaling approach whereby T11TS acts as an anti-neoplastic agent, thus helping to chart out newer avenues in the fight against gliomas.

CD58 Alterations Govern Antitumor Immune Responses by Inducing PDL1 and IDO in Diffuse Large B-Cell Lymphoma.

Recurrent abnormalities in immune surveillance-related genes affect the progression of diffuse large B-cell lymphoma (DLBCL) and modulate the response to therapeutic interventions. CD58 interacts with the CD2 receptor on T cells and NK cells and is recurrently mutated and deleted in DLBCL, suggesting that it may play a role in regulating antitumor immunity. In this study, we comprehensively analyzed the genomic characteristics of CD58 through targeted next-generation sequencing, RNA sequencing (RNA-seq), whole-exome sequencing, and single-cell RNA-seq in patients with newly diagnosed DLBCL. The CD58 mutation rate was 9.1%, and the copy number loss rate was 44.7% among all enrolled patients with DLBCL. Notably, CD58 genetic alterations, along with low CD58 expression, significantly correlated with reduced rates of response to R-CHOP therapy and inferior progression-free survival and overall survival. Single-cell RNA-seq revealed that CD58 expression in tumor cells was negatively correlated with CD8+ T-cell exhaustion/dysfunction status. Insufficient T-cell activation resulting from CD58 alterations could not be attributed solely to CD2 signaling. CD58 inhibited the activity of the JAK2/STAT1 pathway by activating the LYN/CD22/SH2 domain-containing phosphatase 1 (SHP1) axis, thereby limiting PDL1 and IDO expression. Elevated PDL1 and IDO expression in CD58-deficient DLBCL cells led to immune evasion and tumor-intrinsic resistance to chimeric antigen receptor T-cell therapy. Direct activation of CD58-CD2 costimulatory signaling in combination with anti-PDL1 blockade or IDO inhibitor sensitized CD58-deficient DLBCL to chimeric antigen receptor T-cell therapy. Collectively, this work identified the multiple roles of CD58 in regulating antitumor immune responses in DLBCL. Significance: Loss of CD58 mediates immune evasion and therapy resistance in diffuse large B-cell lymphoma by upregulating PDL1 and IDO through LYN/CD22/SHP1 signaling, providing potential targets and therapeutic strategies to improve patient treatment.

Neurogenic factor-induced Langerhans cell activation in diabetic mice with mechanical allodynia.

BACKGROUND: Langerhans cells (LCs) are antigen-presenting dendritic cells located in the skin. It has been reported that LC activation is associated with painful diabetic neuropathy (PDN); however, the mechanism of LC activation is still unclear. METHODS: The db/db mouse, a rodent model of PDN, was used to study the roles of LCs in the development of PDN in type 2 diabetes. Hind foot pads from db/db and control db/+ mice from 5 to 24 weeks of age (encompassing the period of mechanical allodynia development and its abatement) were collected and processed for immunohistochemistry studies. LCs were identified with immunohistochemistry using an antibody against CD207 (Langerin). The intraepidermal nerve fibers and subepidermal nerve plexus were identified by immunohistochemistry of protein gene product 9.5 (PGP 9.5) and tropomyosin-receptor kinase (Trk) A, the high affinity nerve growth factor receptor. RESULTS: CD207-positive LCs increased in the db/db mouse during the period of mechanical allodynia, from 8 to 10 weeks of age, in both the epidermis and subepidermal plexus. At 16 weeks of age, when mechanical allodynia diminishes, LC populations were reduced in the epidermis and subepidermal plexus. Epidermal LCs (ELCs) were positive for Trk A. Subepidermal LCs (SLCs) were positive for CD68, suggesting that they are immature LCs. Additionally, these SLCs were positive for the receptor of advanced glycation end products (RAGE) and were in direct contact with TNF-α-positive nerve fibers in the subepidermal nerve plexus during the period of mechanical allodynia. Intrathecal administration of SB203580, a p38 kinase inhibitor, significantly reduced mechanical allodynia, TNF-α expression in the subepidermal plexus, and increased both ELC and SLC populations during the period of mechanical allodynia. CONCLUSIONS: Our data support the hypothesis that increased LC populations in PDN are activated by p38-dependent neurogenic factors and may be involved in the pathogenesis of PDN.

Molecular identification of an MHC-independent ligand recognized by a human {alpha}/{beta} T-cell receptor.

During an analysis of T-cell responses against human renal cell carcinoma (RCC), we identified a CD4(+) T-cell line that showed TCR-mediated recognition and lysis of nearly all RCC lines regardless of MHC type. We have now elucidated the nature of the ligand for this α/ß TCR, and it contains no MHC-related moiety and does not involve classic peptide processing. First, matrix metalloproteinase 14 (MMP14) expressed on RCC cells releases membrane-bound TRAIL expressed by the T cell; then, soluble TRAIL binds to its receptor DR4 (TRAIL-R1), which is expressed on tumor cells, and this TRAIL-DR4 complex is recognized by the TCR through a complementarity-determining region 3α (CDR3α)-mediated interaction. Direct and specific antigen-TCR interaction was demonstrated when the immobilized recombinant TRAIL/DR4 complex stimulated the TCR. In addition, amino acid substitutions in the CDR3α of the TCR either obliterated or enhanced target-specific recognition. This description of the molecular nature of a non-MHC target structure recognized by a naturally occurring α/ß TCR not only broadens our concept of what the TCR can recognize, but also raises the question of whether such a T cell could be of clinical utility against RCC.

Human CD14hi monocytes and myeloid dendritic cells provide a cell contact-dependent costimulatory signal for early CD40 ligand expression.

CD40L on CD4(+) T cells plays a vital role in the activation of antigen-presenting cells, thus catalyzing a positive feedback loop for T-cell activation. Despite the pivotal juxtaposition of CD40L between antigen-presenting cells and T-cell activation, only a T-cell receptor stimulus is thought to be required for early CD40L surface expression. We show, for the first time, that CD40L expression on peripheral blood CD4(+) T cells is highly dependent on a cell-cell interaction with CD14(hi)CD16(-) monocytes. Interactions with ICAM-1, LFA-3, and to a lesser extent CD80/CD86 contribute to this enhancement of CD40L expression but are not themselves sufficient. The contact-mediated increase in CD40L expression is dependent on new mRNA and protein synthesis. Circulating myeloid dendritic cells also possess this costimulatory activity. By contrast, CD14(lo)CD16(+) monocytes, plasmacytoid dendritic cells, B-cell lymphoma lines, and resting, activated, and Epstein-Barr virus-immortalized primary B cells all lack the capacity to up-regulate early CD40L. The latter indicates that a human B cell cannot activate its cognate T cell to deliver CD40L-mediated help. This finding has functional implications for the role of biphasic CD40L expression, suggesting that the early phase is associated with antigen-presenting cell activation, whereas the late phase is related to B-cell activation.

The CD2-CD58 axis: A novel marker predicting poor prognosis in patients with low-grade gliomas and potential therapeutic approaches.

INTRODUCTION: Low-grade gliomas (LGGs) are currently considered a premalignant condition for high-grade gliomas (HGGs) and are characterized by a relatively intact immune system. Immunotherapeutic modalities may offer a safe and effective treatment option for these patients. However, the CD2-CD58 axis, an important component of the immunological synapse, remains unknown in LGG. METHODS: RNA-seq data from TCGA databases were analyzed. Immune cell infiltration was determined using a single-sample gene set enrichment analysis (ssGSEA) based on integrated immune gene sets from published studies. Kaplan-Meier survival analysis, univariate and multivariate logistic analysis, and the ESTIMATE algorithm were employed to evaluate the impact of the CD2-CD58 axis on adult LGG patients. RESULTS: The expression of the CD2-CD58 axis was found to be elevated with increasing of WHO grade (p < .05). Uni- and multi-variable logistic analysis demonstrated that age, WHO grade, and CD58 levels were associated with poor prognosis in LGG patients with (p < .01). MetaSape pathways analysis revealed the involvement of CD58 in regulating T cell activation, leukocyte-mediated immunity, and the positive regulation of cell activation in WHO grade II and III. CD58 expression correlated with infiltrations of CD4+ lymphocytes, NK cells, and macrophages cells. The ESTIMATE algorithm indicated that patients with high CD58 expression had significantly higher immune scores compared with low CD58 expression in WHO grade II/III, but no statistical difference was observed in WHO grade IV (p < .05). Furthermore, correlation analysis demonstrated the significant association between CD58 and CD274 (r = 0.581, p < .001), HAVCR2 (r = 0.58i7, p < .001), and LGALS9 (r = 0.566, p < .001). Immunohistochemical staining further confirmed the relationship of CD58, HAVCR2, WHO grade, and prognosis in grade II and III patients. CONCLUSION: Overall, our findings highlight the significant association between the CD2-CD58 axis and poor survival in LGG patients. High CD58 expression is implicated in T cell-mediated immune responses as an immunosuppressive factor and affect inhibitory immune checkpoint genes.

The CD58-CD2 axis is co-regulated with PD-L1 via CMTM6 and shapes anti-tumor immunity.

The cell-autonomous balance of immune-inhibitory and -stimulatory signals is a critical process in cancer immune evasion. Using patient-derived co-cultures, humanized mouse models, and single-cell RNA-sequencing of patient melanomas biopsied before and on immune checkpoint blockade, we find that intact cancer cell-intrinsic expression of CD58 and ligation to CD2 is required for anti-tumor immunity and is predictive of treatment response. Defects in this axis promote immune evasion through diminished T cell activation, impaired intratumoral T cell infiltration and proliferation, and concurrently increased PD-L1 protein stabilization. Through CRISPR-Cas9 and proteomics screens, we identify and validate CMTM6 as critical for CD58 stability and upregulation of PD-L1 upon CD58 loss. Competition between CD58 and PD-L1 for CMTM6 binding determines their rate of endosomal recycling over lysosomal degradation. Overall, we describe an underappreciated yet critical axis of cancer immunity and provide a molecular basis for how cancer cells balance immune inhibitory and stimulatory cues.

Ligation of the CD2 co-stimulatory receptor enhances IL-2 production from first-generation chimeric antigen receptor T cells.

T cells bearing chimeric antigen receptors (CARs) are broadly categorised into first- and second-generation receptors. Second-generation CARs contain a co-stimulatory signalling molecule and have been shown to secrete IL-2, undergo greater proliferation and to have enhanced persistence in vivo. However, we have shown that T cells bearing a first-generation CAR containing a CD19-targeting scFv (single-chain variable fragment) and the CD3ζ-signalling domain are able to produce IL-2 upon co-culture with CD19(+) B-cell lymphomas independent of CD28 activity. Here, we report that signalling through endogenous CD2 following ligation with its ligands, CD48 in mouse and CD58 in humans, drives IL-2 production by first-generation CD19-specific CAR. Moreover, the high levels of IL-2 produced by human T cells engrafted with a second-generation CD28-containing CAR during target-cell recognition are dependent to a degree upon CD2 receptor activity. These observations highlight the fact that the functional activity induced by T-cell-expressed CARs is dependent upon endogenous 'natural' receptor interactions. A deeper understanding of the role of these activities will serve to further refine the design of future CARs to either exploit or avoid these interactions.