RESUMO
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.
Assuntos
Antígeno B7-H1 , Antígenos CD58 , Indolamina-Pirrol 2,3,-Dioxigenase , Linfoma Difuso de Grandes Células B , Humanos , Linfoma Difuso de Grandes Células B/imunologia , Linfoma Difuso de Grandes Células B/genética , Linfoma Difuso de Grandes Células B/patologia , Antígeno B7-H1/genética , Antígeno B7-H1/metabolismo , Indolamina-Pirrol 2,3,-Dioxigenase/genética , Indolamina-Pirrol 2,3,-Dioxigenase/metabolismo , Antígenos CD58/genética , Antígenos CD58/metabolismo , Feminino , Masculino , Mutação , Animais , Pessoa de Meia-Idade , Camundongos , Linhagem Celular Tumoral , IdosoRESUMO
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.
Assuntos
Doenças Autoimunes , Lúpus Eritematoso Sistêmico , Esclerose Múltipla , Humanos , Doenças Autoimunes/genética , Predisposição Genética para Doença , Estudo de Associação Genômica Ampla , Lúpus Eritematoso Sistêmico/genética , Esclerose Múltipla/genética , Esclerose Múltipla/metabolismo , Polimorfismo de Nucleotídeo Único , Antígenos CD58/metabolismoRESUMO
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.
Assuntos
Aterosclerose , Doenças Cardiovasculares , Infecções por Citomegalovirus , Infecções por HIV , Animais , Humanos , Antígenos CD28/metabolismo , Infecções por HIV/tratamento farmacológico , Citomegalovirus , Antígenos CD58/metabolismo , Macaca mulatta , Linfócitos T CD8-Positivos , Linfócitos T CD4-Positivos , Aterosclerose/metabolismoRESUMO
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.
Assuntos
Neoplasias Colorretais , Detecção Precoce de Câncer , Humanos , Antígenos CD2/química , Antígenos CD2/metabolismo , Antígenos CD58/química , Antígenos CD58/metabolismo , Simulação de Dinâmica Molecular , Neoplasias Colorretais/tratamento farmacológicoRESUMO
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.
Assuntos
Glioma , Adulto , Humanos , Glioma/genética , Glioma/terapia , Imunidade Celular , Estimativa de Kaplan-Meier , Ativação Linfocitária , Prognóstico , Antígenos CD2/metabolismo , Antígenos CD58/metabolismoRESUMO
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.
Assuntos
Carcinoma Hepatocelular , Neoplasias Hepáticas , Humanos , beta Catenina/metabolismo , Carcinógenos , Carcinoma Hepatocelular/patologia , Linhagem Celular Tumoral , Proliferação de Células , Glicogênio Sintase Quinase 3 beta , Neoplasias Hepáticas/patologia , Proteínas Proto-Oncogênicas c-akt/metabolismo , Antígenos CD58/metabolismoRESUMO
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.
Assuntos
Antígeno B7-H1 , Melanoma , Camundongos , Animais , Antígeno B7-H1/genética , Linfócitos T , Antígenos CD58/química , Antígenos CD58/metabolismo , Melanoma/genética , Melanoma/metabolismo , Ativação LinfocitáriaRESUMO
In psoriasis and other inflammatory skin diseases, keratinocytes (KCs) secrete chemokines that attract T cells, which, in turn, cause epidermal hyperplasia by secreting proinflammatory cytokines. To date, it remains unclear whether skin-homing T cells, particularly memory T cells, can also be activated by direct cell contact with KCs. In this study, we demonstrated the ability of primary human KCs to activate human memory T cells directly by transmitting costimulatory signals through the CD6/CD166/CD318 axis. Interestingly, despite being negative for CD80/CD86, KCs initiate a metabolic shift within T cells. Blockade of the CD6/CD166/CD318 axis prevents mammalian target of rapamycin activation and T cell proliferation but promotes oxidative stress and aerobic glycolysis. In addition, it diminishes formation of central memory T cells. Importantly, although KC-mediated costimulation by CD2/CD58 also activates T cells, it cannot compensate for the lack of CD6 costimulation. Therefore, KCs likely differentially regulate T cell functions in the skin through two distinct costimulatory receptors: CD6 and CD2. This may at least in part explain the divergent effects observed when treating inflammatory skin diseases with antibodies to CD6 versus CD2. Moreover, our findings may provide a molecular basis for selective interference with either CD6/CD166/CD318, or CD2/CD58, or both to specifically treat different types of inflammatory skin diseases.
Assuntos
Antígenos CD , Ativação Linfocitária , Humanos , Antígenos CD/metabolismo , Antígenos CD58/metabolismo , Queratinócitos , Estresse Oxidativo , Serina-Treonina Quinases TOR/metabolismo , Linfócitos T/metabolismoRESUMO
BACKGROUND/AIM: CD58 is an immune adhesion molecule on the cellular surface. It was previously found that a high expression of CD58 predicted a poor prognosis of patients with lower-grade gliomas. Therefore, the aim of this paper was to investigate the association between CD58 and breast cancer. MATERIALS AND METHODS: CD58 gene expression data downloaded from cBioPortal was compared between the different subtypes of breast cancer. Clinical prognosis was examined using Kaplan-Meier analysis and multivariable Cox regression analysis. The association between CD58 expression and immune cell infiltration was estimated using the TIMER 2.0 web platform. Finally, the tumour sphere formation of aldehyde dehydrogenase 1 (ALDH1)high basal-like breast cancer stem cells in which CD58 was knocked down using siRNA was measured. RESULTS: CD58 mRNA was mainly enriched in claudin-low and basal-like subtypes. The high expression of CD58 predicted a good prognosis in patients with luminal A and luminal B breast cancer. This prediction may be due to the association of immune cell infiltration with CD58. Notably, patients with luminal A breast cancer with a high expression of CD58 in association with ALDH1A3 exhibited a good prognosis; however, this did not apply to patients with basal-like breast cancer. The in vitro experiments revealed that knockdown of CD58 inhibited the tumour sphere formation ability of ALDH1high basal-like cancer cells. CONCLUSION: CD58 may function as a potential prognostic biomarker and therapeutic target in ALDH-positive basal-like cancer stem cells.
Assuntos
Neoplasias da Mama , Feminino , Humanos , Família Aldeído Desidrogenase 1 , Biomarcadores Tumorais/metabolismo , Neoplasias da Mama/patologia , Claudinas , Prognóstico , Retinal Desidrogenase/genética , Retinal Desidrogenase/metabolismo , RNA Mensageiro , RNA Interferente Pequeno , Antígenos CD58/metabolismoRESUMO
CD2 is largely described to promote T cell activation when engaged by its ligands, CD48 in mice and CD58 in humans, that are present on antigen-presenting cells (APCs). However, both CD48 and CD58 are also expressed on T cells. By generating new knockout mouse strains lacking CD2 or CD48 in the C57BL/6 background, we determined that whereas CD2 was necessary on T cells for T cell activation, its ligand CD48 was not required on APCs. Rather, CD48 was also needed on T cells. One exception was during cytotoxicity, which required CD48 on T cells and APCs. Fluorescence resonance energy transfer (FRET) studies in nonimmune cells provided evidence that cis interactions between CD2 and CD48 existed within individual cells. CD2-CD48 interactions on T cells enabled more robust T cell receptor (TCR) signals, including protein tyrosine phosphorylation. Using T cells from a CD2 knock-in mouse in which a tag was inserted at the carboxyl terminus of CD2, mass spectrometry analyses revealed that the role of CD2 in T cell activation correlated with its ability to interact with components of the TCR complex and the protein tyrosine kinase Lck. CD2-CD58 provided a similar function in human T cells. Thus, our data imply that T cell-intrinsic cis interactions of CD2 with its ligands are required for TCR signaling and T cell activation. Interactions with ligands on APCs contribute during cytotoxicity.
Assuntos
Antígenos CD , Ativação Linfocitária , Linfócitos T , Animais , Antígenos CD2/química , Antígenos CD2/metabolismo , Antígeno CD48/metabolismo , Antígenos CD58/metabolismo , Humanos , Ligantes , Camundongos , Camundongos Endogâmicos C57BL , Receptores de Antígenos de Linfócitos T/metabolismoRESUMO
The in vivo persistence of adoptively transferred T cells is predictive of antitumor response. Identifying functional properties of infused T cells that lead to in vivo persistence and tumor eradication has remained elusive. We profiled CD19-specific chimeric antigen receptor (CAR) T cells as the infusion products used to treat large B cell lymphomas using high-throughput single-cell technologies based on time-lapse imaging microscopy in nanowell grids (TIMING), which integrates killing, cytokine secretion, and transcriptional profiling. Our results show that the directional migration of CD19-specific CAR T cells is correlated with multifunctionality. We showed that CD2 on T cells is associated with directional migration and that the interaction between CD2 on T cells and CD58 on lymphoma cells accelerates killing and serial killing. Consistent with this, we observed that elevated CD58 expression on pretreatment tumor samples in patients with relapsed or refractory large B cell lymphomas treated with CD19-specific CAR T cell therapy was associated with complete clinical response and survival. These results highlight the importance of studying dynamic T cell-tumor cell interactions in identifying optimal antitumor responses.
Assuntos
Antígenos CD2/metabolismo , Antígenos CD58/metabolismo , Linfoma Difuso de Grandes Células B , Linfócitos T , Antígenos CD19 , Humanos , Imunoterapia Adotiva/métodos , Linfoma Difuso de Grandes Células B/genética , Linfoma Difuso de Grandes Células B/terapia , Receptores de Antígenos de Linfócitos T , Análise de Célula ÚnicaRESUMO
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.
Assuntos
Artrite Experimental , Artrite Reumatoide , Helianthus , Peptidomiméticos , Animais , Artrite Experimental/tratamento farmacológico , Artrite Reumatoide/tratamento farmacológico , Antígenos CD58/química , Antígenos CD58/metabolismo , Helianthus/química , Helianthus/metabolismo , Humanos , Imunidade , Imunomodulação , Camundongos , Peptídeos/farmacologia , Peptídeos Cíclicos/metabolismo , Peptídeos Cíclicos/farmacologia , Peptidomiméticos/farmacologia , Peptidomiméticos/uso terapêutico , Inibidores da Tripsina/uso terapêuticoRESUMO
CD2 is a costimulatory protein expressed in all mature T/NK-cells, in particular memory T-cells. CD58 (or LFA-3) is the receptor for CD2 and is ubiquitously expressed. CD2-CD58 interaction has key functions in T-cell activation and organization of the immunological synapse between T- and antigen-presenting cells. Cancer cells have developed multiple mechanisms to evade immune surveillance. Loss of CD58 expression is one frequently reported in diffuse large B-cell lymphomas (DLBCL). On the other hand, in non-hematological neoplasms, tumor infiltrating lymphocytes (TILs) with reduced expression of CD2 have been associated with defective cytotoxicity and T-cell exhaustion. Here, we reported a case of DLBCL involving the jejunal mucosa associated with a rim of cytotoxic reactive T-cells with features of immune evasion (CD2- and TCR-) and T-cell exhaustion (PD1 + high). This case likely exemplifies a previously unrecognized immune evasion mechanism in lymphoma involving a decreased CD2 expression in the lymphoma-associated T-cells.
Assuntos
Linfoma Difuso de Grandes Células B , Linfoma de Células T , Antígenos CD2/metabolismo , Antígenos CD58/metabolismo , Humanos , Evasão da Resposta Imune , Ativação Linfocitária , Receptores de Antígenos de Linfócitos TRESUMO
Background: Cytogenetics at diagnosis is the most important prognostic factor for adult acute myeloid leukemia (AML), but nearly 50% of AML patients who exhibit cytogenetically normal AML (CN-AML) do not undergo effective risk stratification. Therefore, the development of potential biomarkers to further define risk stratification for CN-AML patients is worth exploring. Methods: Transcriptome data from 163 cases in the GSE12417-GPL96 dataset and 104 CN-AML patient cases in the GSE71014-GPL10558 dataset were downloaded from the Gene Expression Omnibus database for overall survival (OS) analysis and validation. Results: The combination of Wilms tumor 1 (WT1) and cluster of diffraction 58 (CD58) can predict the prognosis of CN-AML patients. High expression of WT1 and low expression of CD58 were associated with poor OS in CN-AML. Notably, when WT1 and CD58 were used to concurrently predict OS, CN-AML patients were divided into three groups: low risk, WT1lowCD58high; intermediate risk, WT1highCD58high or WT1lowCD58low; and high risk, WT1highCD58low. Compared with low-risk patients, intermediate- and high-risk patients had shorter survival time and worse OS. Furthermore, a nomogram model constructed with WT1 and CD58 may personalize and reveal the 1-, 2-, 3-, 4-, and 5-year OS rate of CN-AML patients. Both time-dependent receiver operating characteristics and calibration curves suggested that the nomogram model demonstrated good performance. Conclusion: Higher expression of WT1 with lower CD58 expression may be a potential biomarker for risk stratification of CN-AML patients. Moreover, a nomogram model constructed with WT1 and CD58 may personalize and reveal the 1-, 2-, 3-, 4-, and 5-year OS rates of CN-AML patients.
Assuntos
Biomarcadores Tumorais , Antígenos CD58/genética , Regulação Leucêmica da Expressão Gênica , Leucemia Mieloide Aguda/genética , Proteínas WT1/genética , Antígenos CD58/metabolismo , Biologia Computacional , Análise Citogenética , Bases de Dados Genéticas , Perfilação da Expressão Gênica , Humanos , Leucemia Mieloide Aguda/diagnóstico , Leucemia Mieloide Aguda/metabolismo , Prognóstico , Modelos de Riscos Proporcionais , Transcriptoma , Proteínas WT1/metabolismo , Fluxo de TrabalhoRESUMO
Resistance to immune checkpoint inhibitors (ICIs) is a key challenge in cancer therapy. To elucidate underlying mechanisms, we developed Perturb-CITE-sequencing (Perturb-CITE-seq), enabling pooled clustered regularly interspaced short palindromic repeat (CRISPR)-Cas9 perturbations with single-cell transcriptome and protein readouts. In patient-derived melanoma cells and autologous tumor-infiltrating lymphocyte (TIL) co-cultures, we profiled transcriptomes and 20 proteins in ~218,000 cells under ~750 perturbations associated with cancer cell-intrinsic ICI resistance (ICR). We recover known mechanisms of resistance, including defects in the interferon-γ (IFN-γ)-JAK/STAT and antigen-presentation pathways in RNA, protein and perturbation space, and new ones, including loss/downregulation of CD58. Loss of CD58 conferred immune evasion in multiple co-culture models and was downregulated in tumors of melanoma patients with ICR. CD58 protein expression was not induced by IFN-γ signaling, and CD58 loss conferred immune evasion without compromising major histocompatibility complex (MHC) expression, suggesting that it acts orthogonally to known mechanisms of ICR. This work provides a framework for the deciphering of complex mechanisms by large-scale perturbation screens with multimodal, single-cell readouts, and discovers potentially clinically relevant mechanisms of immune evasion.
Assuntos
Antígenos CD58/imunologia , Resistencia a Medicamentos Antineoplásicos/imunologia , Melanoma/patologia , Análise de Célula Única/métodos , Evasão Tumoral , Antígenos CD58/genética , Antígenos CD58/metabolismo , Sistemas CRISPR-Cas , Técnicas de Cocultura , Biologia Computacional/métodos , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/genética , Epitopos/genética , Técnicas de Inativação de Genes , Humanos , Inibidores de Checkpoint Imunológico/farmacologia , Interferon gama/imunologia , Interferon gama/metabolismo , Linfócitos do Interstício Tumoral/patologia , Melanoma/tratamento farmacológico , Melanoma/imunologia , Análise de Sequência de RNA , Evasão Tumoral/genéticaRESUMO
Peptides were designed to inhibit the protein-protein interaction of CD2 and CD58 to modulate the immune response. This work involved the design and synthesis of eight different peptides by replacing each amino acid residue in peptide 6 with alanine as well as grafting the peptide to the sunflower trypsin-inhibitor framework. From the alanine scanning studies, mutation at position 2 of the peptide was shown to result in increased potency to inhibit cell adhesion interactions. The most potent peptide from the alanine scanning was further studied for its detailed three-dimensional structure and binding to CD58 protein using surface plasmon resonance and flow cytometry. This peptide was used to graft to the sunflower trypsin inhibitor to improve the stability of the peptide. The grafted peptide, SFTI-a1, was further studied for its potency as well as its thermal, chemical, and enzymatic stability. The grafted peptide exhibited improved activity compared to our previously grafted peptide and was stable against thermal and enzymatic degradation.
Assuntos
Antígenos CD2/metabolismo , Antígenos CD58/metabolismo , Peptídeos Cíclicos/química , Sequência de Aminoácidos , Sítios de Ligação , Ligação Competitiva , Antígenos CD2/química , Antígenos CD58/química , Adesão Celular/efeitos dos fármacos , Linhagem Celular , Desenho de Fármacos , Humanos , Simulação de Acoplamento Molecular , Peptídeos Cíclicos/metabolismo , Peptídeos Cíclicos/farmacologia , Ligação Proteica , Mapas de Interação de Proteínas/efeitos dos fármacos , Estabilidade Proteica , Linfócitos T/citologia , Linfócitos T/efeitos dos fármacos , Linfócitos T/metabolismo , Inibidores da Tripsina/farmacologiaRESUMO
The oral mucosa is constantly exposed to a plethora of stimuli including food antigens, commensal microbiota and pathogens, requiring distinct immune responses. We previously reported that human oral epithelial cells (OECs) suppress immune responses to bacteria, using H413 and TR146 OEC lines and primary OECs in co-culture with dendritic cells (DCs) and T cells (OEC-conditioned cells). OECs reduced DCs expression of CD80/CD86 and IL-12/TNFα release and impaired T cell activation. Here, we further evaluated the immunosuppression by these OECs and investigated the underlying mechanisms. OEC-conditioned DCs did not induce CD4 T cell polarization towards Treg, judging by the absence of FoxP3 expression. OECs also repressed T-bet/IFNγ expression in CD4 and CD8 T cells activated by DCs or anti-CD3/CD28 antibodies. This inhibition depended on OEC:T cell ratio and IFNγ repression occurred at the transcriptional level. Time-lapse experiments showed that OECs inhibited early steps of T cell activation, consistent with OECs inability to suppress T cells stimulated with PMA/ionomycin. Blocking CD40/CD40L, CD58/CD2 and PD-L1/PD-1 interactions with specific antibodies did not disrupt T cell suppression by OECs. However, preventing prostaglandin E2 (PGE2) synthesis or blocking PGE2 binding to the cognate EP2/EP4 receptors, restored IFNγ and TNFα production in OEC-conditioned T cells. Finally, treating OECs with poly(I:C), which simulates viral infections, limited T cell suppression. Overall, these results point to an inherent ability of OECs to suppress immune responses, which can nonetheless be eluded when OECs are under direct assault.
Assuntos
Linfócitos T CD4-Positivos/metabolismo , Linfócitos T CD8-Positivos/metabolismo , Dinoprostona/metabolismo , Células Epiteliais/imunologia , Células Epiteliais/metabolismo , Linfócitos T Reguladores/metabolismo , Antígenos CD2/imunologia , Antígenos CD2/metabolismo , Linfócitos T CD4-Positivos/imunologia , Antígenos CD40/imunologia , Antígenos CD40/metabolismo , Ligante de CD40/imunologia , Ligante de CD40/metabolismo , Antígenos CD58/metabolismo , Linfócitos T CD8-Positivos/imunologia , Linhagem Celular , Células Dendríticas/imunologia , Células Dendríticas/metabolismo , Dinoprostona/imunologia , Humanos , Tolerância Imunológica/imunologia , Imunidade/imunologia , Interferon gama/imunologia , Interferon gama/metabolismo , Interleucina-12/imunologia , Interleucina-12/metabolismo , Ativação Linfocitária/imunologia , Linfócitos T Reguladores/imunologia , Transcrição Gênica/imunologiaRESUMO
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.
Assuntos
Antígenos CD2/metabolismo , Antígenos CD58/metabolismo , Linfócitos T CD8-Positivos/metabolismo , Sinapses Imunológicas/metabolismo , Linfócitos do Interstício Tumoral/metabolismo , Neoplasias/metabolismo , Receptor de Morte Celular Programada 1/metabolismo , Adesão Celular , Células Cultivadas , Humanos , Tolerância Imunológica , Ativação Linfocitária , Ligação Proteica , Receptor Cross-Talk , Receptores de Antígenos de Linfócitos T/metabolismo , Transdução de Sinais , Análise de Célula ÚnicaRESUMO
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.
Assuntos
Vasos Sanguíneos/fisiologia , Linfócitos T CD4-Positivos/imunologia , Infecções por Citomegalovirus/imunologia , Citomegalovirus/fisiologia , Infecções por HIV/imunologia , HIV-1/fisiologia , Placa Aterosclerótica/imunologia , Antígenos CD28/metabolismo , Antígenos CD57/metabolismo , Antígenos CD58/metabolismo , Receptor 1 de Quimiocina CX3C/metabolismo , Movimento Celular , Quimiocina CX3CL1/metabolismo , Coinfecção , Citotoxicidade Imunológica , Humanos , Receptores CXCR3/metabolismo , RiscoRESUMO
The interplay between keratinocytes and immune cells, especially T cells, plays an important role in the pathogenesis of chronic inflammatory skin diseases. During psoriasis, keratinocytes attract T cells by releasing chemokines, while skin-infiltrating self-reactive T cells secrete proinflammatory cytokines, e.g., IFNγ and IL-17A, that cause epidermal hyperplasia. Similarly, in chronic graft-versus-host disease, allogenic IFNγ-producing Th1/Tc1 and IL-17-producing Th17/Tc17 cells are recruited by keratinocyte-derived chemokines and accumulate in the skin. However, whether keratinocytes act as nonprofessional antigen-presenting cells to directly activate naive human T cells in the epidermis remains unknown. Here, we demonstrate that under proinflammatory conditions, primary human keratinocytes indeed activate naive human T cells. This activation required cell contact and costimulatory signaling via CD58/CD2 and CD54/LFA-1. Naive T cells costimulated by keratinocytes selectively differentiated into Th1 and Th17 cells. In particular, keratinocyte-initiated Th1 differentiation was dependent on costimulation through CD58/CD2. The latter molecule initiated STAT1 signaling and IFNγ production in T cells. Costimulation of T cells by keratinocytes resulting in Th1 and Th17 differentiation represents a new explanation for the local enrichment of Th1 and Th17 cells in the skin of patients with a chronic inflammatory skin disease. Consequently, local interference with T cell-keratinocyte interactions may represent a novel strategy for the treatment of Th1 and Th17 cell-driven skin diseases.