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Although inhibitors targeting the PD1/PD-L1 immune checkpoint are showing comparably good outcomes, a significant percentage of head and neck squamous cell carcinoma (HNSCC) patients do not respond to treatment. Apart from using different treatment strategies, another possibility would be to target other immune checkpoints operating in these non-responding tumors. To obtain an overview of which checkpoint ligands are expressed on HNSCC tumor cells and if these ligands are affected by HGF/MET signaling, we used mRNA sequencing and antibody-based techniques for identifying checkpoint ligands in six HNSCC tumor cell lines. Furthermore, we compared our results to mRNA sequencing data. From the checkpoint ligands we investigated, VISTA was expressed the highest at the RNA level and was also the most ubiquitously expressed. PD-L2 and B7-H3 were expressed comparably lower and were not present in all cell lines to the same extent. B7-H4, however, was only detectable in the Detroit 562 cell line. Concerning the effect of HGF on the ligand levels, PD-L2 expression was enhanced with HGF stimulation, whereas other checkpoint ligand levels decreased with stimulation. B7-H4 levels in the Detroit 562 cell line drastically decreased with HGF stimulation. This is of interest because both the checkpoint ligand and the growth factor are reported to be connected to epithelial–mesenchymal transition in the literature. [ABSTRACT FROM AUTHOR]

The inducible T cell co-stimulator ligand (ICOSL), expressed by antigen presenting cells, binds to the inducible T cell co-stimulator (ICOS) on activated T cells. Improper function of the ICOS/ICOSL pathway has been implicated in several autoimmune diseases, including multiple sclerosis (MS). Previous studies showed that ICOS-knockout (KO) mice exhibit severe experimental autoimmune encephalomyelitis (EAE), the animal model of MS, but data on ICOSL deficiency are not available. In our study, we explored the impact of both ICOS and ICOSL deficiencies on MOG35-55 -induced EAE and its associated immune cell dynamics by employing ICOSL-KO and ICOS-KO mice with a C57BL/6J background. During EAE resolution, MOG-driven cytokine levels and the immunophenotype of splenocytes were evaluated by ELISA and multiparametric flow cytometry, respectively. We found that both KO mice exhibited an overlapping and more severe EAE compared to C57BL/6J mice, corroborated by a reduction in memory/regulatory T cell subsets and interleukin (IL-)17 levels. It is noteworthy that an unsupervised analysis showed that ICOSL deficiency modifies the immune response in an original way, by affecting T central and effector memory (TCM, TEM), long-lived CD4+ TEM cells, and macrophages, compared to ICOS-KO and C57BL/6J mice, suggesting a role for other binding partners to ICOSL in EAE development, which deserves further study. [ABSTRACT FROM AUTHOR]

Abstract Background Immunotherapy has become the fastest‐adopting treatment paradigm for lung cancer with improved survival. By binding with its ligand (inducible T‐cell co‐stimulator and its ligand [ICOSL]), an inducible T‐cell co‐stimulator (ICOS) could contribute to reversing immunosuppression and improving immune response and thus be a potential target for cancer immunotherapy. Methods We selected 54 formalin‐fixed, paraffin‐embedded tumor tissues from cases with stage I–III lung adenocarcinoma cancer. Immunohistochemical expression of ICOS and ICOSL was evaluated. The correlation with clinical parameters in Chinese patients was also compared with TCGA results. Results The positive rates of ICOS and ICOSL were 68% and 81.5%, respectively, in lung tumor tissues. Of these, 9 cases had a low expression of ICOS, and 22 cases had a high expression of ICOS; ICOSL expression was low in 20 cases and high in 24 cases. According to the International Association for the Study of Lung Cancer (8th edition), phase I lesions were detected in 21 cases, phase II lesions in 15 cases, and phase III lesions in 18 cases. The median survival time of all patients was 44.5 months, and the median disease‐free survival was 32 months. Univariate analysis showed that the factors significantly associated with overall survival were tumor size, regional lymph node involvement, stage, and expression level of ICOS/ICOSL. Survival analysis using log‐rank test indicated that the lower ICOS+ cell infiltration may predict poor prognosis, whereas lower ICOSL protein expression may be associated with better prognosis, but ICOSL data need further validation in larger samples due to inconsistency in TCGA mRNA prediction. Conclusion ICOS/ICOSL might be associated with prognosis of lung cancer, and ICOS and its ligand may be potential therapeutic targets in non‐small cell lung cancer.

Introduction: Malignant mesothelioma is a rare and aggressive form of cancer. Despite improvements in cancer treatment, there are still no curative treatment modalities for advanced stage of the malignancy. The aim of this study was to evaluate the anti-tumor efficacy of a novel combinatorial therapy combining AdV5/ 3-D24-ICOSL-CD40L, an oncolytic vector, with an anti-PD-1monoclonal antibody. Methods: The efficacy of the vector was confirmed in vitro in three mesothelioma cell lines -- H226, Mero-82, and MSTO-211H, and subsequently the antineoplastic properties in combination with anti-PD-1 was evaluated in xenograft H226 mesothelioma BALB/c and humanized NSG mouse models. Results and discussion: Anticancer efficacy was attributed to reduced tumour volume and increased infiltration of tumour infiltrating lymphocytes, including activated cytotoxic T-cells (GrB+CD8+). Additionally, a correlation between tumour volume and activated CD8+ tumour infiltrating lymphocytes was observed. These findings were confirmed by transcriptomic analysis carried out on resected human tumour tissue, which also revealed upregulation of CD83 and CRTAM, as well as several chemokines (CXCL3, CXCL9, CXCL11) in the tumour microenvironment. Furthermore, according to observations, the combinatorial therapy had the strongest effect on reducing mesothelin and MUC16 levels. Gene set enrichment analysis suggested that the combinatorial therapy induced changes to the expression of genes belonging to the "adaptive immune response" gene ontology category. Combinatorial therapy with oncolytic adenovirus with checkpoint inhibitors may improve anticancer efficacy and survival by targeted cancer cell destruction and triggering of immunogenic cell death. Obtained results support further assessment of the AdV5/3-D24-ICOSL-CD40L in combination with checkpoint inhibitors as a novel therapeutic perspective for mesothelioma treatment. [ABSTRACT FROM AUTHOR]

Background: Bullous pemphigoid (BP) is an autoimmune skin-blistering disease. Systemic corticosteroids remain the first line treatment for moderate-to-severe BP with the potential for severe adverse events. Dupilumab has emerged as an alternative option for BP patients. Objective: We evaluated the efficiency and safety of dupilumab on BP treatment and explored a mode of drug action in depth. Methods and results: A multicenter retrospective cohort included 20 BP patients who received dupilumab with or without systemic corticosteroid in dupilumab group, and 20 matched BP patients who received corticosteroid alone in conventional group. Serum samples were collected from 20 patients (10 from dupilumab group and 10 from conventional group) at baseline and week 4. Compared to systemic corticosteroid alone, dupilumab with or without systemic corticosteroid was similarly efficacious in clinical remission at week4 (complete remission plus partial remission: 100%) and week24 (complete remission plus partial remission:100%), but allowing significant decreases in the cumulative doses of corticosteroids with reducing the incidence of adverse events. However, dupilumab did not decrease BP180 antibody despite an obvious clinical improvement. Comparative plasma proteomic analysis performed before and after treatment in 3 BP patients from dupilumab group revealed that drug use was associated with 30 differentially expressed proteins, including 26 down-regulated and 4 up-regulated proteins. The former consisted of immune related proteins involved in T/B cell interactions (inducible T-cell costimulator ligand, ICOSL) and in the activation of eosinophils (PRG2), mast cells (S100A12), and complement (CR2). TARC and ICOSL levels correlated with BP severity in patients who received either dupilumab or conventional treatment. Conclusion: Dupilumab has similar efficacy in treating BP as conventional drugs, by inhibiting the activities of many types of immune cells and complement, and regulating the interactions between T and B cells. [ABSTRACT FROM AUTHOR]

The inducible T cell co-stimulator ligand (ICOSL), expressed by antigen presenting cells, binds to the inducible T cell co-stimulator (ICOS) on activated T cells. Improper function of the ICOS/ICOSL pathway has been implicated in several autoimmune diseases, including multiple sclerosis (MS). Previous studies showed that ICOS-knockout (KO) mice exhibit severe experimental autoimmune encephalomyelitis (EAE), the animal model of MS, but data on ICOSL deficiency are not available. In our study, we explored the impact of both ICOS and ICOSL deficiencies on MOG35-55 -induced EAE and its associated immune cell dynamics by employing ICOSL-KO and ICOS-KO mice with a C57BL/6J background. During EAE resolution, MOG-driven cytokine levels and the immunophenotype of splenocytes were evaluated by ELISA and multiparametric flow cytometry, respectively. We found that both KO mice exhibited an overlapping and more severe EAE compared to C57BL/6J mice, corroborated by a reduction in memory/regulatory T cell subsets and interleukin (IL-)17 levels. It is noteworthy that an unsupervised analysis showed that ICOSL deficiency modifies the immune response in an original way, by affecting T central and effector memory (TCM, TEM), long-lived CD4+ TEM cells, and macrophages, compared to ICOS-KO and C57BL/6J mice, suggesting a role for other binding partners to ICOSL in EAE development, which deserves further study.

Inducible costimulator (ICOS) and its ligand (ICOSL) are critical to regulate the immune response in autoimmune diseases. The participation of B lymphocytes exhibits pathogenic potential in the disease process of rheumatoid arthritis (RA). However, the precise role of ICOSL in RA remains unclear. In this study, we aimed to explore the regulatory effects of CD19+ICOSL+ B cells in the pathogenesis of RA. We demonstrated the increased expression of ICOS and ICOSL in patients with RA and collagen-induced arthritis (CIA) mice. The population of CD19+ICOSL+ B-cell subset was significantly correlated with clinicopathological characteristics of RA patients and CIA mice. Adoptive transfer of CD19+ICOSL+ B cells aggravated arthritic progression in CIA mice. Moreover, microarray analysis revealed that CD19+ICOSL+ cells could exert pivotal effect in pathological process of RA. Further blocking of ICOSL significantly inhibited proinflammatory responses and ameliorated arthritic progression. Therefore, CD19+ICOSL+ B-cell subset could be defined as a specific pathogenic cell subpopulation involved in immunopathological damage of RA. Blockade of ICOSL is promising to be a potential new approach for RA therapy. [ABSTRACT FROM AUTHOR]

Inducible costimulator (ICOS) and its ligand (ICOSL) are critical to regulate the immune response in autoimmune diseases. The participation of B lymphocytes exhibits pathogenic potential in the disease process of rheumatoid arthritis (RA). However, the precise role of ICOSL in RA remains unclear. In this study, we aimed to explore the regulatory effects of CD19+ICOSL+ B cells in the pathogenesis of RA. We demonstrated the increased expression of ICOS and ICOSL in patients with RA and collagen-induced arthritis (CIA) mice. The population of CD19+ICOSL+ B-cell subset was significantly correlated with clinicopathological characteristics of RA patients and CIA mice. Adoptive transfer of CD19+ICOSL+ B cells aggravated arthritic progression in CIA mice. Moreover, microarray analysis revealed that CD19+ICOSL+ cells could exert pivotal effect in pathological process of RA. Further blocking of ICOSL significantly inhibited proinflammatory responses and ameliorated arthritic progression. Therefore, CD19+ICOSL+ B-cell subset could be defined as a specific pathogenic cell subpopulation involved in immunopathological damage of RA. Blockade of ICOSL is promising to be a potential new approach for RA therapy.

Purposes: Effective and selective T-cell activation and proliferation during the T-cell expansion phase of a cellular adoptive immunotherapy method, challenging because recent studies revealed the importance of each subtype of T-cells in different immunologic strategies against tumors, like CAR-T cell therapies. Artificial antigen presenting cells (aAPCs) regarded as a natural way to manipulate T-cell subtypes activation and specific proliferation. In the current study, we utilized K562 cells based aAPC method expressing the ICOSL molecule, to evaluate T-cell subtypes differentiation rate and functional status. Methods: CD3+T-cells isolated and, co-cultured with ICOSL expressing K562 cells. After 4, 6, and 10 days selective CD markers of T-cell subtypes and each subtype’s activity-related genes levels evaluated by qPCR methods. Results: During the culture period, CD4+ Th related phenotype reduced continuously, and in day 10th of culture CD4+ T-cell’s population significantly reduced (P=0.029). In contrast, the CD8+ population ratio was ascending during the study period but was not statistically significant. FoxP3+CD25-, Treg population ratio was significantly increased during the time in comparison with the control group, as well as memory T-cell phenotypic marker, CD127+, expressing cells ratio. T-cell subpopulations activity-related genes expression levels evaluated too, and the Th1 related IL-2 and INF-γ reductions observed alongside regulatory T-cells gene (IL-10) and Cytotoxic T-cell’s related gene (Geranzym-A) elevations. Conclusion: We concluded that the K562-ICOSL based aAPC system is working and effective in T-cell short to medium culture periods, and this approach preparing relatively selective milieu for CD8+ T-Cell differentiation and much less Treg differentiation.

Negative selection of developing T cells plays a significant role in T‐cell tolerance to self‐antigen. This process relies on thymic antigen‐presenting cells which express both self‐antigens and cosignaling molecules. Inducible T‐cell costimulator (ICOS) belongs to the CD28 family of cosignaling molecules and binds to ICOS ligand (ICOSL). The ICOS signaling pathway plays important roles in shaping the immune response to infections, but its role in central tolerance is less well understood. Here we show that ICOSL is expressed by subsets of thymic dendritic cells and medullary thymic epithelial cells as well as thymic B cells. ICOS expression is upregulated as T cells mature in the thymus and correlates with T‐cell receptor signal strength during thymic selection. We also provide evidence of a role for ICOS signaling in mediating negative selection. Our findings suggest that ICOS may fine‐tune T‐cell receptor signals during thymic selection contributing to the generation of a tolerant T‐cell population. [ABSTRACT FROM AUTHOR]

Background: Pulmonary hypertension (PH) is a life-threatening complication of connective tissue diseases (CTD); in this study, we aimed at investigating the potential role of inducible co-stimulator (ICOS) and its ligand (ICOS-L) as biomarkers of PH in CTD. Materials and Methods: We recruited 109 patients: 84 CTD patients, 13 patients with CTD complicated by pulmonary arterial hypertension (PAH), and 12 subjects with PAH alone. All recruited patients underwent a complete clinical and instrumental assessment along with quantitative measurement of serum ICOS and ICOS-L. Results: Independently of the underlying cause, patients with PAH were older and had a lower glomerular filtration rate. Interestingly, patients with both CTD-related and CTD-unrelated PAH had higher ICOS and ICOS-L serum concentrations than CTD patients (0.0001 for both). When compared to CTD patients, those affected by CTD-PAH showed higher ICOS (440 (240–600) vs. 170 (105–275) pg/mL, p = 0.0001) and ICOS-L serum concentrations (6000 (4300–7000) vs. 2450 (1500–4100) pg/mL; p = 0.0001). In a logistic regression, ICOS and ICOS-L were associated with a diagnosis of PAH, independently from age, gender, and renal function. The corresponding receiver operating characteristic (ROC) curves demonstrated a good diagnostic performance for both ICOS and ICOS-L. Conclusions: ICOS and ICOS-L are increased in patients with PAH, irrespectively from the underlying cause, and represent promising candidate biomarkers for the diagnostic screening for PAH among CTDs patients. [ABSTRACT FROM AUTHOR]

Background: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the novel coronavirus responsible for the COVID-19 pandemic. The viral protein of SARS-CoV-2, spike protein (SP), mediates entry into host cells, contributing to pathogenesis of COVID-19. Prostate cancer is the most common cancer among men in the United States. Inducible T-cell costimulator ligand (ICOSL) and intercellular cell adhesion molecule 2 (ICAM-2) are expressed in cancer cells and their roles in cancer growth remain controversial. It is unknown if SP can affect the expression of ICAM-2 or ICOSL in prostate cancer. This study investigated the effects of SARS-CoV-2 SP on the expression of ICAM-2 and ICOSL and the time-dependent effect of SP on growth and survival of prostate cancer cells. Methods: The effect of SARS-CoV-2 SP on the survival of a widely-used prostate cancer cell line, LNCaP, was assessed using clonogenic cell survival assay and quick cell proliferation assay. Reverse transcription-polymerase chain reaction (RT-PCR) and immunohistochemistry (IHC) were performed to investigate the expression of ICAM-2 and ICOSL. The survival of an additional prostate cancer cell line, PC-3, was also evaluated by clonogenic survival assay. Results: After 3 days, a significant decrease in the percentage of colonies in LNCaP cells treated with SP was found, which was paralleled by a decrease in optical density (OD) value in LNCaP cells in the presence of SP. A significant decrease in the percentage of colonies treated with SP was also found in PC-3 cells evaluated by clonogenic survival assay. In addition, the mRNA expression of ICAM-2 was lower, whereas the mRNA expression of ICOSL was higher in SP-treated LNCaP cells. This was supported by protein expressions for ICAM-2 and ICOSL evaluated with IHC. Conclusions: In LNCaP cells, SARS-CoV-2 SP downregulates the expression of ICAM-2 but upregulates the expression of ICOSL. SARS-CoV-2 SP inhibits growth of prostate cancer cells in a time-dependent manner. Further studies are needed to fully address the roles of ICAM-2 and ICOSL in the inhibition prostate cancer growth by SARS-CoV-2 SP.

Purposes: Effective and selective T-cell activation and proliferation during the T-cell expansion phase of a cellular adoptive immunotherapy method, challenging because recent studies revealed the importance of each subtype of T-cells in different immunologic strategies against tumors, like CAR-T cell therapies. Artificial antigen presenting cells (aAPCs) regarded as a natural way to manipulate T-cell subtypes activation and specific proliferation. In the current study, we utilized K562 cells based aAPC method expressing the ICOSL molecule, to evaluate T-cell subtypes differentiation rate and functional status. Methods: CD3+T-cells isolated and, co-cultured with ICOSL expressing K562 cells. After 4, 6, and 10 days selective CD markers of T-cell subtypes and each subtype's activity-related genes levels evaluated by qPCR methods. Results: During the culture period, CD4+ Th related phenotype reduced continuously, and in day 10th of culture CD4+ T-cell's population significantly reduced (P=0.029). In contrast, the CD8+ population ratio was ascending during the study period but was not statistically significant. FoxP3+CD25-, Treg population ratio was significantly increased during the time in comparison with the control group, as well as memory T-cell phenotypic marker, CD127+, expressing cells ratio. T-cell subpopulations activity-related genes expression levels evaluated too, and the Th1 related IL-2 and INF-γ reductions observed alongside regulatory T-cells gene (IL-10) and Cytotoxic T-cell's related gene (Geranzym-A) elevations. Conclusion: We concluded that the K562-ICOSL based aAPC system is working and effective in T-cell short to medium culture periods, and this approach preparing relatively selective milieu for CD8+ T-Cell differentiation and much less Treg differentiation. [ABSTRACT FROM AUTHOR]

Viral infections are controlled, and very often cleared, by activated T lymphocytes. The inducible co-stimulator (ICOS) mediates its functions by binding to its ligand ICOSL, enhancing T-cell activation and optimal germinal center (GC) formation. Here, we show that ICOSL is heavily downmodulated during infection of antigen-presenting cells by different herpesviruses. We found that, in murine cytomegalovirus (MCMV), the immunoevasin m138/fcr-1 physically interacts with ICOSL, impeding its maturation and promoting its lysosomal degradation. This viral protein counteracts T-cell responses, in an ICOS-dependent manner, and limits virus control during the acute MCMV infection. Additionally, we report that blockade of ICOSL in MCMV-infected mice critically regulates the production of MCMV-specific antibodies due to a reduction of T follicular helper and GC B cells. Altogether, these findings reveal a novel mechanism evolved by MCMV to counteract adaptive immune surveillance, and demonstrates a role of the ICOS:ICOSL axis in the host defense against herpesviruses.

Genome-wide association studies have identified ICOSLG, which encodes the inducible costimulator ligand (ICOSLG or ICOSL) as a susceptibility locus for inflammatory bowel disease. ICOSL has been implicated in the enhancement of pattern recognition receptor signaling in dendritic cells, induction of IL-10 production by CD4 T cells, and the generation of high-affinity antibodies to specific antigens--all of which can potentially explain its involvement in gastrointestinal inflammation. Here, we show that murine ICOSL deficiency results in significant enrichment of IL-10-producing CD4 T cells particularly in the proximal large intestine. Transient depletion of IL-10-producing cells from adult ICOSL-deficientmice induced severe colonic inflammation that was prevented when mice were first treated with metronidazole. ICOSL-deficient mice displayed reduced IgA and IgG antibodies in the colon mucus and impaired serum antibody recognition of microbial antigens, including flagellins derived from mucus-associated bacteria of the Lachnospiraceae family. Confirming the synergy between ICOSL and IL-10, ICOSL deficiency coupled with CD4-specific deletion of the Il10 gene resulted in juvenile onset colitis that was impeded when pups were fostered by ICOSL-sufficient dams. In this setting, we found that both maternally acquired and host-derived antibodies contribute to the life anti-commensal antibody repertoire that mediates this protection in early life. Collectively, our findings reveal a partnership between ICOSL-dependent anti-commensal antibodies and IL-10 in adaptive immune regulation of the microbiota in the large intestine. Furthermore, we identify ICOSL deficiency as an effective platform for exploring the functions of anti-commensal antibodies in host-microbiota mutualism. [ABSTRACT FROM AUTHOR]

The role of the inducible costimulatory of T cells (ICOS) has been shown to be important for many different T cell outcomes and is indispensable for follicular helper T cell (TFH) responses. Since its discovery, there have been several studies on the regulation of ICOS at a transcriptional level. However, the post-translational regulation of ICOS has not been well characterized. Here, we demonstrate that ICOS is internalized following ligation. We show that costimulation with CD3 results in differential internalization and fate than stimulation of ICOS alone. Additionally, we show that ICOS internalization is PI3K and clathrin mediated. The studies presented here not only increase the mechanistic understanding of ICOS post-translational regulation but also give insight into the potential mechanisms by which T cells expressing high affinity receptors may be preferentially chosen to become TFH cells with increased ICOS levels.

Background: Pulmonary hypertension (PH) is a life-threatening complication of connective tissue diseases (CTD); in this study, we aimed at investigating the potential role of inducible co-stimulator (ICOS) and its ligand (ICOS-L) as biomarkers of PH in CTD. Materials and Methods: We recruited 109 patients: 84 CTD patients, 13 patients with CTD complicated by pulmonary arterial hypertension (PAH), and 12 subjects with PAH alone. All recruited patients underwent a complete clinical and instrumental assessment along with quantitative measurement of serum ICOS and ICOS-L. Results: Independently of the underlying cause, patients with PAH were older and had a lower glomerular filtration rate. Interestingly, patients with both CTD-related and CTD-unrelated PAH had higher ICOS and ICOS-L serum concentrations than CTD patients (0.0001 for both). When compared to CTD patients, those affected by CTD-PAH showed higher ICOS (440 (240–600) vs. 170 (105–275) pg/mL, p = 0.0001) and ICOS-L serum concentrations (6000 (4300–7000) vs. 2450 (1500–4100) pg/mL; p = 0.0001). In a logistic regression, ICOS and ICOS-L were associated with a diagnosis of PAH, independently from age, gender, and renal function. The corresponding receiver operating characteristic (ROC) curves demonstrated a good diagnostic performance for both ICOS and ICOS-L. Conclusions: ICOS and ICOS-L are increased in patients with PAH, irrespectively from the underlying cause, and represent promising candidate biomarkers for the diagnostic screening for PAH among CTDs patients.

T cells edited by chimeric antigen receptors (CAR) have shown great potential in the treatment of tumors, especially malignant blood tumors. However, there remain many obstacles in the CAR-T therapy against solid tumors, such as the expansion of CAR-T cells ex vivo and the exhaustion of CAR-T cells in vivo. In order to solve these problems, we described a novel CAR which is targeting GPC3 by expressing CD28 co-stimulation domain and CD3z ITAM (G328z), meanwhile co-expressing ICOSL extracellular and transmembrane region fused with 41BB cytoplasmic domain (G328z-ICOSL-41BB). Compared with G328z, G328z-ICOSL-41BB fusion protein significantly reinforced the expansion ability of CAR-T cells ex vivo, and prolonged the survival time of mice with hepatocellular carcinoma. We now demonstrate that the enhancement of CAR-T cell activity is dependent on the enhanced PI3K signaling pathway and up-regulated expression of Bcl2 to inhibit apoptosis and promote proliferation of CAR-T cells. Besides, the CAR with ICOSL-41BB fusion protein have been strengthened significantly in comparison with fusing ICOSL protein only, which might be caused by the fact that ICOSL-41BB not only supplies ICOS signal for other cells, but also provides 41BB signal for itself. Consequently, CARs with ICOSL-41BB fusion protein could increase the therapeutic efficacy against solid tumors in vivo compared with the G328z CAR, which might further assist the development of potent and durable T cell therapeutics.

Recently, we demonstrated that inducible T-cell costimulator (ICOS) shares its unique ligand (ICOSL) with osteopontin (OPN), and OPN/ICOSL binding promotes tumor metastasis and angiogenesis in the 4T1 breast cancer model. Literature showed that OPN promotes melanoma metastasis by suppressing T-cell activation and recruiting myeloid suppressor cells (MDSC). On the opposite, ICOS/ICOSL interaction usually sustains an antitumor response. Here, we engineered murine B16F10 melanoma cells, by transfecting or silencing ICOSL. In vitro data showed that loss of ICOSL favors anchorage-independent growth and induces more metastases in vivo, compared to ICOSL expressing cells. To dissect individual roles of the three molecules, we compared data from C57BL/6 with those from OPN-KO, ICOS-KO, and ICOSL-KO mice, missing one partner at a time. We found that OPN produced by the tumor microenvironment (TME) favors the metastasis by interacting with stromal ICOSL. This activity is dominantly inhibited by ICOS expressed on TME by promoting Treg expansion. Importantly, we also show that OPN and ICOSL highly interact in human melanoma metastases compared to primary tumors. Interfering with this binding may be explored in immunotherapy either for nonresponding or patients resistant to conventional therapies.

Background/Aims: The aim of this study was to investigate the involvement of inducible co-stimulatory ligand (ICOSL) expression in stimulation of mast cells (MCs) by TNF-α and the ability of TNF-α stimulation of MCs to influence CD4+ T cell differentiation and function. The mechanisms underlying TNF-α stimulation of MCs were also explored. Methods: Mast cells and CD4+ T cells were prepared from C57BL/6 mice (aged 6–8 weeks). ICOSL expression by MCs was measured by real-time PCR and flow cytometry, and levels of IL-4, IL-10 and IFN-γ were measured by ELISA. Results: ICOSL expression by MCs was increased by TNF-α stimulation, and resulted in interaction with CD4+ T cells. The IL-4 and IL-10 levels in the co-culture system increased, while IFN-γ levels decreased. Furthermore, CD4+CD25+Foxp3+ T cell proliferation was induced by co-culture with TNF-α-stimulated MCs. The mechanism by which TNF-α stimulated MCs was dependent on the activation of the MAPK signaling pathway. Conclusion: TNF-α upregulated the expression of ICOSL on mast cells via a mechanism that is dependent on MAPK phosphorylation. TNF-α-treated MCs promoted the differentiation of regulatory T cells and induced a shift in cytokine expression from a Th1 to a Th2 profile by up-regulation ICOSL expression and inhibition of MC degranulation. Our study reveals a novel mechanism by which mast cells regulate T cell function.

Malignant melanoma, an aggressive form of skin cancer, has a low five-year survival rate in patients with advanced disease. Immunotherapy represents a promising approach to improve survival rates among patients at advanced stage. Herein, the aim of the study was to design and produce, by using engineering tools, a novel oncolytic adenovirus AdV-D24- inducible co-stimulator ligand (ICOSL)-CD40L expressing potent co-stimulatory molecules enhancing clinical efficacy through the modulation of anti-cancer immune responses. Firstly, we demonstrated the vector’s identity and genetic stability by restriction enzyme assay and sequencing, then, by performing in vitro and in vivo pre-clinical studies we explored the anti-cancer efficacy of the virus alone or in combination with anti PD-1 inhibitor in human melanoma cell lines, i.e., MUG Mel-1 and MUG Mel-2, and in immunocompetent C57BL/6 melanoma B16V mouse model. We showed that both monotherapy and combination approaches exhibit enhanced anti-cancer ability and immunogenic cell death in in vitro settings. Furthermore, AdV-D24-ICOSL-CD40L combined with anti PD-1 revealed a fall in tumor volume and 100% survival in in vivo context, thus suggesting enhanced efficacy and survival via complementary anti-cancer properties of those agents in melanoma therapy. Collectively, the novel oncolytic vector AdV-D24-ICOSL-CD40L alone or in combination with anticancer drugs, such as check point inhibitors, may open novel therapeutic perspectives for the treatment of melanoma.

Background Inducible co-stimulator (ICOS) is a CD28-related molecule exclusively expressed on activated T cells and plays a critical role in modulating the immune response in breast cancer. The blockage of ICOS pathway has been shown to inhibit the activity of Type 2 T helper cells, thus potentially protecting against cancer growth. The current study aims to investigate the correlation between inducible co-stimulator ligand (ICOSL) expression in tumor tissues and the prognoses of patients with invasive breast cancer. Methods Tumor samples from 562 Chinese patients with invasive breast carcinomas were collected between 2003 and 2010. The expression of ICOSL on breast tumor and adjacent non-cancerous tissue was determined via immunohistochemistry. The overall survival (OS) of patients with positive and negative ICOSL expression were described using Kaplan–Meier curves, respectively. Parametric correlation method was used to analyze the correlation between ICOSL expression and other clinicopathological parameters. ICOSL was selected as a dependent variable for multivariate analysis. Results Positive ICOSL expression was identified on the plasma membrane in both cytoplasm and the nucleus of breast cancer cells. Membrane-expressed ICOSL is determined as an independent prognostic factor for OS in breast cancer but without significantly correlating with other clinicopathologic parameters such as age, menopausal status, depth of invasion, lymph node metastasis status, histologic classification, etc. Conclusion Our study suggests that the up-regulated expression of ICOSL protein in breast tumor cells can be associated with poor prognoses in invasive breast carcinomas.

Background: Immunotherapy has become the fastest-adopting treatment paradigm for lung cancer with improved survival. By binding with its ligand (inducible T-cell co-stimulator and its ligand [ICOSL]), an inducible T-cell co-stimulator (ICOS) could contribute to reversing immunosuppression and improving immune response and thus be a potential target for cancer immunotherapy., Methods: We selected 54 formalin-fixed, paraffin-embedded tumor tissues from cases with stage I-III lung adenocarcinoma cancer. Immunohistochemical expression of ICOS and ICOSL was evaluated. The correlation with clinical parameters in Chinese patients was also compared with TCGA results., Results: The positive rates of ICOS and ICOSL were 68% and 81.5%, respectively, in lung tumor tissues. Of these, 9 cases had a low expression of ICOS, and 22 cases had a high expression of ICOS; ICOSL expression was low in 20 cases and high in 24 cases. According to the International Association for the Study of Lung Cancer (8th edition), phase I lesions were detected in 21 cases, phase II lesions in 15 cases, and phase III lesions in 18 cases. The median survival time of all patients was 44.5 months, and the median disease-free survival was 32 months. Univariate analysis showed that the factors significantly associated with overall survival were tumor size, regional lymph node involvement, stage, and expression level of ICOS/ICOSL. Survival analysis using log-rank test indicated that the lower ICOS+ cell infiltration may predict poor prognosis, whereas lower ICOSL protein expression may be associated with better prognosis, but ICOSL data need further validation in larger samples due to inconsistency in TCGA mRNA prediction., Conclusion: ICOS/ICOSL might be associated with prognosis of lung cancer, and ICOS and its ligand may be potential therapeutic targets in non-small cell lung cancer., (© 2023 The Authors. Animal Models and Experimental Medicine published by John Wiley & Sons Australia, Ltd on behalf of The Chinese Association for Laboratory Animal Sciences.)

Summary: Follicular T‐helper (TFH) cells play a crucial role in three aspects of the germinal center (GC) response. They promote GC formation, arbitrate competition among GC B cells to determine the outcome of affinity maturation, and regulate GC output of memory and plasma cells to shape the long‐lived humoral immune memory. Of fundamental importance are dynamic physical interactions between TFH and B cells, which are the main platform for TFH cells to deliver "help" factors to B cells and also for reciprocal signaling from B cells to maintain the helper state of TFH cells. Recent work has significantly expanded our understanding of how T‐B interactions are spatiotemporally regulated and molecularly orchestrated to fulfill those TFH functions. In this review, we elaborate two modes of T‐B interactions, the antigen‐specific or cognate mode in which TFH cells engage individual antigen‐presenting B cells and the antigen nonspecific bystander mode in which TFH cells are engaged with the ensemble of follicular B cells. We discuss findings that indicate how short‐lived cognate T‐B contacts coupled with an intercellular positive feedback drive affinity‐based selection and how bystander interactions between T and B cells regulate follicular T‐cell recruitment and maintenance of an appropriate helper state. We argue that this combination of bystander and cognate interactions with B cells constantly shapes the internal state of TFH cells and provides the platform to execute their helper functions. [ABSTRACT FROM AUTHOR]

Background: Bullous pemphigoid (BP) is an autoimmune skin-blistering disease. Systemic corticosteroids remain the first line treatment for moderate-to-severe BP with the potential for severe adverse events. Dupilumab has emerged as an alternative option for BP patients., Objective: We evaluated the efficiency and safety of dupilumab on BP treatment and explored a mode of drug action in depth., Methods and Results: A multicenter retrospective cohort included 20 BP patients who received dupilumab with or without systemic corticosteroid in dupilumab group, and 20 matched BP patients who received corticosteroid alone in conventional group. Serum samples were collected from 20 patients (10 from dupilumab group and 10 from conventional group) at baseline and week 4. Compared to systemic corticosteroid alone, dupilumab with or without systemic corticosteroid was similarly efficacious in clinical remission at week4 (complete remission plus partial remission: 100%) and week24 (complete remission plus partial remission:100%), but allowing significant decreases in the cumulative doses of corticosteroids with reducing the incidence of adverse events. However, dupilumab did not decrease BP180 antibody despite an obvious clinical improvement. Comparative plasma proteomic analysis performed before and after treatment in 3 BP patients from dupilumab group revealed that drug use was associated with 30 differentially expressed proteins, including 26 down-regulated and 4 up-regulated proteins. The former consisted of immune related proteins involved in T/B cell interactions (inducible T-cell co-stimulator ligand, ICOSL) and in the activation of eosinophils (PRG2), mast cells (S100A12), and complement (CR2). TARC and ICOSL levels correlated with BP severity in patients who received either dupilumab or conventional treatment., Conclusion: Dupilumab has similar efficacy in treating BP as conventional drugs, by inhibiting the activities of many types of immune cells and complement, and regulating the interactions between T and B cells., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Yan, Xie, Liu, Shan, Wu, Wang, Zuo and Zhang.)

Systemic sclerosis (SSc), an autoimmune disorder, is characterized by vasculopathy, inflammation, progressive perivascular and interstitial fibrosis. Its pathogenesis is largely unknown, however strong evidences suggest that genetic predisposition may contribute to SSc development. Several gene polymorphisms involved in regulatory T cell function have been identified in many autoimmune diseases, including SSc. Moreover, dysregulation of co-stimulatory and/or co-inhibitory signals, including ICOS signalling, can lead to autoimmunity. The aim of the present study was to investigate the association of the FOXP3 rs2294020, ICOS rs6726035 and ICOSL rs378299 SNPs with both the susceptibility and the progression to SSc in an Italian case-series of patients. SNP genotyping results were successfully obtained from a total of 350 subjects including 166 individuals with SSc and 184 healthy controls. Although analysis tests did not show any significant associations between the SNPs under study and susceptibility to SSc, the occurrence of FOXP3 rs2294020 in female patients was associated with decreased time to progression from early to definite SSc (allelic model: HR = 1.43; CI = 1.03-1.99; p = 0.03; dominant model: HR = 1.54; CI = 1.04-2.28; p = 0.03). The inclusion of presence of ACA autoantibodies in the model did not significantly change the estimates. No conclusions can be drawn for the susceptibility to the disease or the time to progression in men due to the low statistical power. This study provides evidence of the association of rs2294020 with SSc evolution in female patients, modulating the time of progression from the diagnosis of early SSc to the diagnosis of definite SSc, while no effect on SSc susceptibility per se was found. rs2294020 may be considered a disease-modifying gene-variant rather than a disease-susceptibility SNP in SSc. [ABSTRACT FROM AUTHOR]

Background/Aims: The aim of this study was to investigate the involvement of inducible co-stimulatory ligand (ICOSL) expression in stimulation of mast cells (MCs) by TNF-α and the ability of TNF-α stimulation of MCs to influence CD4+ T cell differentiation and function. The mechanisms underlying TNF-α stimulation of MCs were also explored. Methods: Mast cells and CD4+ T cells were prepared from C57BL/6 mice (aged 6-8 weeks). ICOSL expression by MCs was measured by real-time PCR and flow cytometry, and levels of IL-4, IL-10 and IFN-γ were measured by ELISA. Results: ICOSL expression by MCs was increased by TNF-α stimulation, and resulted in interaction with CD4+ T cells. The IL-4 and IL-10 levels in the coculture system increased, while IFN-γ levels decreased. Furthermore, CD4+CD25+Foxp3+ T cell proliferation was induced by co-culture with TNF-α-stimulated MCs. The mechanism by which TNF-α stimulated MCs was dependent on the activation of the MAPK signaling pathway. Conclusion: TNF-α upregulated the expression of ICOSL on mast cells via a mechanism that is dependent on MAPK phosphorylation. TNF-α-treated MCs promoted the differentiation of regulatory T cells and induced a shift in cytokine expression from a Th1 to a Th2 profile by up-regulation ICOSL expression and inhibition of MC degranulation. Our study reveals a novel mechanism by which mast cells regulate T cell function. [ABSTRACT FROM AUTHOR]

Impact of smoking on circulating immune cells with emphasis on CD26hiCD161hi MAIT cells and ICOSL+ pDCs, in healthy controls and MS patients. Upon chronic cigarette smoke exposure, inhaled antigens and irritants cause altered lung immune homeostasis. Circulating immune cells are affected, and smoking is associated with an increased risk of developing various disorders, including multiple sclerosis (MS). This study was conducted to determine the impact of smoking on circulating immune cell subsets. Furthermore, we determined whether any smoking‐associated changes were related to MS. With the use of flow cytometry, CFSE assays, and ELISpot assays, we analyzed circulating immune cell phenotypes and quantified antigen‐induced proliferation and cytokine secretion in smokers and nonsmokers in a cohort of 100 healthy individuals (HI). In addition, we analyzed immune cell subsets associated with smoking in 2 independent cohorts of patients with MS. In HI smokers compared with nonsmokers, we found increased blood cell counts of granulocytes, monocytes, and lymphocytes. These cells were not more proinflammatory, autoreactive, or EBV reactive compared with cells from nonsmokers. Phenotypic differences were seen in plasmacytoid dendritic cells (pDCs) and CD8+ T cells as higher percentages of ICOS ligand (ICOSL)+ pDCs and lower percentages of CD26hiCD161hi CD8+ T cells and CCR6+ CD8+ T cells in smokers compared with nonsmokers. In supplemental analyses, we showed that CD26hiCD161hi CD8+ T cells were mainly mucosal‐associated invariant T cells (MAITs). Comparable frequencies of ICOSL+ pDCs, CCR6+ CD8+ T cells, and CD26hiCD161hi CD8+ T cells were found between HI and MS patients who were nonsmokers. Our findings suggest general proinflammatory effects from smoking combined with skewing of specific cell populations in HI and MS patients. The function of these cell populations needs further investigation. [ABSTRACT FROM AUTHOR]

Abrogation of ICOS/ICOS ligand (ICOSL) costimulation prevents the onset of diabetes in the non-obese diabetic (NOD) mouse but, remarkably, yields to the development of a spontaneous autoimmune neuromyopathy. At the pathological level, ICOSL-/- NOD mice show stronger protection from insulitis than their ICOS-/- counterparts. Also, the ICOSL-/- NOD model carries a limited C57BL/6 region containing the Icosl nul mutation, but, in contrast to ICOS-/- NOD mice, no gene variant previously reported as associated to NOD diabetes. Therefore, we aimed at providing a detailed characterization of the ICOSL-/- NOD model. The phenotype observed in ICOSL-/- NOD mice is globally similar to that observed in ICOS-/- and ICOS-/-ICOSL-/- double-knockout NOD mice, manifested by a progressive locomotor disability first affecting the front paws as observed by catwalk analysis and a decrease in grip test performance. The pathology remains limited to peripheral nerve and striated muscle. The muscle disease is characterized by myofiber necrosis/regeneration and an inflammatory infiltrate composed of CD4+ T-cells, CD8+ T-cells, and myeloid cells, resembling human myositis. Autoimmune neuromyopathy can be transferred to NOD.scid recipients by CD4+ but not by CD8+ T-cells isolated from 40-week-old female ICOSL-/- NOD mice. The predominant role of CD4+ T-cells is further demonstrated by the observation that neuromyopathy does not develop in CIITA-/-ICOSL-/- NOD in contrast to β2microglobulin-/-ICOSL-/- NOD mice. Also, the cytokine profile of CD4+ T-cells infiltrating muscle and nerve of ICOSL-/- NOD mice is biased toward a Th1 pattern. Finally, adoptive transfer experiments show that diabetes development requires expression of ICOSL, in contrast to neuromyopathy. Altogether, the deviation of autoimmunity from the pancreas to skeletal muscles in the absence of ICOS/ICOSL signaling in NOD mice is strictly dependent on CD4+ T-cells, leads to myofiber necrosis and regeneration. It provides the first mouse model of spontaneous autoimmune myopathy akin to human myositis. [ABSTRACT FROM AUTHOR]

Objective This study aims to investigate the role of T follicular helper (TFH) cells in the immunopathogenesis of pediatric immune thrombocytopenia (ITP), as well as differences in TFH expansion and its regulation between newly diagnosed ITP (nITP) and chronic pediatric ITP (cITP). Methods Eighty-five children with ITP and 20 age-matched healthy controls were enrolled into this study. TFH cell frequencies and TFH cell-associated regulatory factors before and after treatment were analyzed by flow cytometry, RT-PCR and ELISA. Results The percentages of TFH cells were significantly elevated in both nITP and cITP compared with controls. RT-PCR revealed significant differences in Bcl-6, c-Maf, Blimp-1, ICOSL, TACI and BAFFR mRNA expression in CD4 + T or CD19 + B cells between patients and controls, and further between nITP and cITP, before and after treatment. Moreover, there were significant differences in serum IL-4, IL-21 and BAFF between patients and controls. Conclusion The overactivation of TFH cells may contribute to the immunopathogenesis of pediatric ITP. IL-21 and IL-4 serum levels may affect the differentiation of TFH cells in ITP patients. The aberrant balance between BAFFR-BAFF/TACI-BAFF may be a factor that caused the persistent high expression of ICOSL in pediatric cITP, which consequently lead to the over activation of TFH cells in pediatric cITP. [ABSTRACT FROM AUTHOR]

ICOSL (B7-H2, CD275), a co-stimulatory molecule of the B7 superfamily, functions as a positive signal in immune response. To investigate whether ICOSL could be released into sera and the possible biological function of soluble ICOS (sICOSL), we generated and characterized a functional anti-human ICOSL monoclonal antibody (mAb), 20B10, and developed a novel enzyme-linked immunosorbent assay (ELISA) based on two anti-human ICOSL antibodies with different epitope specificities. Using the ELISA system, we found that sICOSL in the serum of healthy donors increases in an age-dependent manner and that the matrix metalloproteinase inhibitor (MMPI) could suppress sICOSL production. Together, these data demonstrate that the existence of circulating sICOSL in human serum might play an important role in immunoregulation. [ABSTRACT FROM AUTHOR]

Inducible costimulator (ICOS) and its ligand (ICOSL) are critical to regulate the immune response in autoimmune diseases. The participation of B lymphocytes exhibits pathogenic potential in the disease process of rheumatoid arthritis (RA). However, the precise role of ICOSL in RA remains unclear. In this study, we aimed to explore the regulatory effects of CD19 + ICOSL + B cells in the pathogenesis of RA. We demonstrated the increased expression of ICOS and ICOSL in patients with RA and collagen-induced arthritis (CIA) mice. The population of CD19 + ICOSL + B-cell subset was significantly correlated with clinicopathological characteristics of RA patients and CIA mice. Adoptive transfer of CD19 + ICOSL + B cells aggravated arthritic progression in CIA mice. Moreover, microarray analysis revealed that CD19 + ICOSL + cells could exert pivotal effect in pathological process of RA. Further blocking of ICOSL significantly inhibited proinflammatory responses and ameliorated arthritic progression. Therefore, CD19 + ICOSL + B-cell subset could be defined as a specific pathogenic cell subpopulation involved in immunopathological damage of RA. Blockade of ICOSL is promising to be a potential new approach for RA therapy., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Ding, Sun, Jiang, Chang, Shen, Gu and Liu.)

Alopecia areata (AA), an autoimmune disease affecting anagen stage hair follicles, is associated with polymorphisms in immune-related genes and with decreased number of CD4+ CD25+ T regulatory cells (Treg). Treg function is modulated by the forkhead box protein 3 (FOXP3) transcription factor and by inducible costimulator (ICOS), through interaction with the relative ligand, ICOSLG, whose genes are polymorphic. The aim of the study was to investigate whether specific single nucleotide polymorphisms (SNPs) of the rs2294020 FOXP3 and/or rs378299 ICOSLG genes may be associated with AA. A case-control study was performed in 120 AA patients and 84 controls. SNPs were analyzed by gene sequencing. FOXP3 and ICOSLG gene expressions were analyzed by real-time PCR. Increased frequencies of the genotype carrying the FOXP3 rs2294020-3675(A) [ P = 0.002, OR (95 % CI): 2.55 (1.2-2.7)] or the ICOSLG rs378299-509(C) [ P = 0.01, OR (95 % CI): 2.21 (1.1-2.6)] allelic variants were observed in AA patients than in controls. The genotype carrying the combination of the FOXP3 rs2294020-3675(A) and ICOSLG rs378299-509(C) allelic variants with the HLA DQB1*03 allele was more frequently present in AA patients than in controls ( P = 0.04). The presence of the FOXP3 rs2294020-3675(A) or the ICOSLG rs378299-509(C) allelic variant was associated with reduced relative gene expression in AA patients. These data suggest that rs2294020 SNP of FOXP3 gene and rs378299 SNP of ICOSLG gene are associated with AA and with a reduced expression of the FOXP3 and ICOSLG genes in alopecia patients. [ABSTRACT FROM AUTHOR]

Inducible costimulator ligand (ICOSL) expressed on cancer cells has immunoregulatory functions in various malignancies. However, the role of ICOSL in triple-negative breast cancer (TNBC) remains unclear. In this study, the role and expression of ICOSL in TNBC were analyzed using the cBioPortal and GEPIA databases. Then the role of ICOSL in Foxp3+ Treg cell differentiation, reversal of p38 pathway activation and cell proliferation, migration and apoptosis was determined in vitro. Finally, the effect of ICOSL expression on TNBC progression was verified in a nude mouse model of TNBC. We here observed that ICOSL expression in TNBC was found to be related to relapse-free survival, and Treg abundance was positively correlated with ICOSL expression, as demonstrated by database analyses. In vitro experiments showed that ICOSL overexpression (OE) in MDA-MB-231 cells induced cocultured T cells to differentiate into Foxp3+ Treg cells and promoted secretion of the tumor-promoting factors IL-10 and IL-4. Furthermore, in vitro experiments showed that ICOSL reversed p38 phosphorylation and promoted the proliferation, invasion, and metastasis of MDA-MB-231 ICOSL-OE cells. Finally, tumor progression was found to be promoted by ICOSL expression in a TNBC nude mouse model. Together, ICOSL expression can enhance tumor cell growth by inducing Foxp3+ Treg cell differentiation and reversing p38 pathway activation in TNBC., Competing Interests: None., (AJCR Copyright © 2022.)

Recently, we demonstrated that inducible T-cell costimulator (ICOS) shares its unique ligand (ICOSL) with osteopontin (OPN), and OPN/ICOSL binding promotes tumor metastasis and angiogenesis in the 4T1 breast cancer model. Literature showed that OPN promotes melanoma metastasis by suppressing T-cell activation and recruiting myeloid suppressor cells (MDSC). On the opposite, ICOS/ICOSL interaction usually sustains an antitumor response. Here, we engineered murine B16F10 melanoma cells, by transfecting or silencing ICOSL. In vitro data showed that loss of ICOSL favors anchorage-independent growth and induces more metastases in vivo, compared to ICOSL expressing cells. To dissect individual roles of the three molecules, we compared data from C57BL/6 with those from OPN-KO, ICOS-KO, and ICOSL-KO mice, missing one partner at a time. We found that OPN produced by the tumor microenvironment (TME) favors the metastasis by interacting with stromal ICOSL. This activity is dominantly inhibited by ICOS expressed on TME by promoting Treg expansion. Importantly, we also show that OPN and ICOSL highly interact in human melanoma metastases compared to primary tumors. Interfering with this binding may be explored in immunotherapy either for nonresponding or patients resistant to conventional therapies. [ABSTRACT FROM AUTHOR]

Objective. Inducible costimulator (ICOS), expressed on activated T cells, and its ligand, ICOS ligand (ICOSL), expressed on antigen-presenting cells, have been considered a single receptor–ligand pair. Here we investigated the expression of ICOS and ICOSL in patients with SSc.Methods. ICOS expression on peripheral blood T cells, and ICOSL expression on B cells and macrophages was determined by flow cytometry. Expression of ICOS and ICOSL was assessed by immunohistological staining and real-time PCR of lesional skin.Results. ICOS expression levels were specifically increased on both peripheral blood memory T cells and Tregs from early dcSSc patients compared with those from healthy controls. Mean ICOSL expression on B cells or macrophages was comparable between SSc patients and healthy controls. ICOS-expressing T cells, ICOSL-expressing macrophages and mRNA levels of ICOS and ICOSL were increased in the lesional skin of patients with early dcSSc. In vitro ICOS costimulation enhanced production of IFN-γ, IL-4 and IL-17A from T cells in SSc patients vs normal controls. Soluble ICOS levels were significantly increased in SSc patients and were negatively associated with the presence of ACAs and positively associated with CRP values. Serum levels of soluble ICOS were more closely associated with clinical features compared with levels of soluble IL-2 receptor.Conclusion. Augmented ICOS signalling may contribute to the pathogenesis of SSc during early progressive disease. Soluble ICOS levels may be used as a serum marker for the activity and severity of SSc. [ABSTRACT FROM PUBLISHER]

Background: The role of costimulatory molecules expressed on lymphocytes and thyrocytes in hyperthyroidism has attracted increasing attention and research has shown a close correlation between variant expression of these molecules on lymphocytes and thyrocytes and the development of GD. Meterials and Methods: Thyroid tissues were collected from GD patients during surgery and from Hashimoto disease (HT) and non-toxic goiter (NTG) patients as controls. ICOSL expression on infiltrated B cells and TFC was detected by flow cytometry (FCM), reverse transcription polymerase chain reaction (RT-PCR) and immunohistochemistry (IHC). Variation in ICOSL expression on TFC in primary cultures was analyzed in the absence or presence of cytokines using FCM assays. The role of ICOS-ICOSL signaling in proliferation, thyroid hormone production and thyroglobulin (Tg) release was investigated in primary TFC cultures using ICOS gene transfected L929 cells (ICOS-L929 cells) and the blocking ICOSL antibody (11 C4) in MTT assays and radioimmunoassays. Results and Discussion: ICOSL expression on infiltrated B cells and TFC was detected in GD patient tissue. However, ICOSL expression was only detected on infiltrated B cells in control HT and NTG patient tissue. ICOSL expression on TFC was induced in vitro by the proinflammatory cytokines IFN-γ, IL-6 and TNF-α. Compared with mock transfected L929 (mock-L929) control cells, ICOS-L929 cells promoted significant proliferation of primary cultured TFC, with increased thyroid hormone and Tg production (all P < 0.01). TFC proliferation and production of thyroid hormones and Tg were inhibited significantly in the presence of ICOSL blocking antibody (11 C4) (all P < 0.05). Our observations suggest that ICOS-ICOSL signal plays a direct role in proliferation and differentiation of TFC and may exert important effects in the initiation, maintenance and exaggeration of autoimmune responses in local tissue. [ABSTRACT FROM AUTHOR]

Inducible co-stimulator ligand (ICOSL) is a rather newly defined co-stimulatory molecule, which, through interaction with ICOS expressed on T cells, plays an important role in T-cell activation, differentiation and function. Th2-type immune responses are critical for the development and maintenance of allergic responses including asthma. Using knockout (KO) mice, we have assessed the role of ICOSL in allergic airway inflammation and responsiveness using a standard mouse asthma model induced by ovalbumin (OVA) sensitization and challenge. Our data show that OVA-treated ICOSL KO mice exhibit significantly less lung eosinophilic infiltration, histopathology, mucus production and virtually no airway hyperresponsiveness in contrast to wild-type (Wt) counterparts. Serum antibody analysis showed that antigen-specific IgG1, IgG2a and IgE titers in ICOSL KO mice were significantly lower than those of Wt controls. Also, CD4+ T cells isolated from ICOSL KO mice produced less Th2 cytokines (IL-4, IL-5, IL-10 and IL-13) but more Th1 (IFN-γ) and IL-17 than their Wt controls. Taken together, we conclude that ICOSL plays an important role in predisposing individuals to allergic airway hyperresponsiveness by enhancing IgE antibody class switching and Th2 cytokine production and diminishing the Th17 response and airway eosinophilia. [ABSTRACT FROM PUBLISHER]

• Aire-overexpressing DC2.4 cells repress TFH cell differentiation. • Aire inhibits the expression of ICOSL and IL-27 in DC2.4 cells. • Aire inhibits TFH cell differentiation by blocking ICOSL and IL-27 expression in DC2.4 cells. Autoimmune regulator (Aire) is a transcription factor that plays a pivotal role in the maintenance of immune tolerance. However, little is known about its roles in peripheral immune tolerance. Aire is predominantly expressed in dendritic cells (DCs) in the periphery. DCs with higher inducible costimulatory ligand (ICOSL) expression and interleukin (IL)-27 production have been reported highly suggesting its roles in inducing follicular helper T cells (TFH). Here we use Aire-overexpressing DC2.4 cells in a coculture system composed of naïve CD4+ T cells to test whether Aire in DCs affects TFH cell differentiation. We found that the frequency of TFH cells and its specific cytokine IL-21 were decreased in CD4+ T lymphocytes after cocultured with Aire overexpressed DC2.4 cells. In activated DCs, ICOSL expression and IL-27 production were significantly suppressed by Aire. Furthermore, addition of recombinant ICOSL or IL-27 in the coculture system enhanced TFH cell differentiation and IL-21 expression. These results revealed that Aire plays an indispensable role in the repression of dendritic cells on the differentiation and function of TFH cells by inhibiting ICOSL and IL-27 expression. [ABSTRACT FROM AUTHOR]

• AIRE-BMDCs inhibit T FH differentiation through IL-6, IL-27 and ICOSL. • Transplantation of AIRE-BMDCs prevents and attenuates type 1 diabetes. • AIRE-BMDCs inhibit T FH expansion via ICOSL in NOD mice. The strong genetic association between autoimmune regulator (AIRE) and autoimmune diseases indicates its critical role in immune tolerance. AIRE deficiency is thought to promote the development of follicular helper T (T FH) cells, which are considered to be essential in B cell proliferation. Excessive T FH cell generation is a key step towards the development of autoimmune diseases, including type 1 diabetes. However, the potential mechanism by which AIRE contributes to the generation and function of the T FH cell population has remained elusive. We show that AIRE reduced T FH cell generation by inhibiting the expression of inducible costimulatory ligand (ICOSL), interleukin (IL)-6 and IL-27 in dendritic cells (DCs). To understand the precise impact of AIRE-overexpressing bone marrow-derived DCs (AIRE-BMDCs) on type 1 diabetes progression and the associated molecular mechanisms, we transferred AIRE-BMDCs to recipient NOD mice and found that transplantation of AIRE-BMDCs can prevent or delay the onset of diabetes, attenuate diabetes after the establishment of overt hyperglycaemia, and lead to the inhibition of autoreactive pathological T FH cells and germinal centre (GC) B cells. To further determine the potential mechanism underlying this T FH cell depletion, BMDCs were cotransferred with recombinant mouse ICOSL (ICOSLG protein). We demonstrated that NOD mice were more susceptible to diabetes when they received AIRE-BMDCs and ICOSLG than when they received only mock-vehicle BMDCs (GFP-BMDCs). In addition, we did not observe the reversal of diabetes in any mice subjected to this cotransfer system. A single cycle of ICOSLG treatment temporarily promoted T FH cell proliferation and GC development. Our results reveal a mechanistic role of AIRE-BMDCs in the initiation of T FH cell differentiation, and the AIRE-mediated decrease in ICOSL expression in BMDCs plays a critical role. The effect of decreased ICOSL expression in type 1 diabetes will guide the design and evaluation of parallel studies in patients. [ABSTRACT FROM AUTHOR]

Recently, we demonstrated that inducible T-cell costimulator (ICOS) shares its unique ligand (ICOSL) with osteopontin (OPN), and OPN/ICOSL binding promotes tumor metastasis and angiogenesis in the 4T1 breast cancer model. Literature showed that OPN promotes melanoma metastasis by suppressing T-cell activation and recruiting myeloid suppressor cells (MDSC). On the opposite, ICOS/ICOSL interaction usually sustains an antitumor response. Here, we engineered murine B16F10 melanoma cells, by transfecting or silencing ICOSL. In vitro data showed that loss of ICOSL favors anchorage-independent growth and induces more metastases in vivo, compared to ICOSL expressing cells. To dissect individual roles of the three molecules, we compared data from C57BL/6 with those from OPN-KO, ICOS-KO, and ICOSL-KO mice, missing one partner at a time. We found that OPN produced by the tumor microenvironment (TME) favors the metastasis by interacting with stromal ICOSL. This activity is dominantly inhibited by ICOS expressed on TME by promoting Treg expansion. Importantly, we also show that OPN and ICOSL highly interact in human melanoma metastases compared to primary tumors. Interfering with this binding may be explored in immunotherapy either for nonresponding or patients resistant to conventional therapies.

Malignant melanoma, an aggressive form of skin cancer, has a low five-year survival rate in patients with advanced disease. Immunotherapy represents a promising approach to improve survival rates among patients at advanced stage. Herein, the aim of the study was to design and produce, by using engineering tools, a novel oncolytic adenovirus AdV-D24- inducible co-stimulator ligand (ICOSL)-CD40L expressing potent co-stimulatory molecules enhancing clinical efficacy through the modulation of anti-cancer immune responses. Firstly, we demonstrated the vector's identity and genetic stability by restriction enzyme assay and sequencing, then, by performing in vitro and in vivo pre-clinical studies we explored the anti-cancer efficacy of the virus alone or in combination with anti PD-1 inhibitor in human melanoma cell lines, i.e., MUG Mel-1 and MUG Mel-2, and in immunocompetent C57BL/6 melanoma B16V mouse model. We showed that both monotherapy and combination approaches exhibit enhanced anti-cancer ability and immunogenic cell death in in vitro settings. Furthermore, AdV-D24-ICOSL-CD40L combined with anti PD-1 revealed a fall in tumor volume and 100% survival in in vivo context, thus suggesting enhanced efficacy and survival via complementary anti-cancer properties of those agents in melanoma therapy. Collectively, the novel oncolytic vector AdV-D24-ICOSL-CD40L alone or in combination with anticancer drugs, such as check point inhibitors, may open novel therapeutic perspectives for the treatment of melanoma. [ABSTRACT FROM AUTHOR]