Affect T-cell Responses Research Articles

The pro-tumorigenic cytokine IL-32 has a high turnover in multiple myeloma cells due to proteolysis regulated by oxygen-sensing cysteine dioxygenase and deubiquitinating enzymes.

IL-32 is a pro-inflammatory cytokine expressed by several types of cancer cells and immune cells. Currently, no treatment targeting IL-32 is available, and its intracellular and exosomal localization make IL-32 less accessible to drugs. We previously showed that hypoxia promotes IL-32 expression through HIF1α in multiple myeloma cells. Here, we demonstrate that high-speed translation and ubiquitin-dependent proteasomal degradation lead to a rapid IL-32 protein turnover. We find that IL-32 protein half-life is regulated by the oxygen-sensing cysteine-dioxygenase ADO and that deubiquitinases actively remove ubiquitin from IL-32 and promote protein stability. Deubiquitinase inhibitors promoted the degradation of IL-32 and may represent a strategy for reducing IL-32 levels in multiple myeloma. The fast turnover and enzymatic deubiquitination of IL-32 are conserved in primary human T cells; thus, deubiquitinase inhibitors may also affect T-cell responses in various diseases.

Inosine, gut microbiota, and cancer immunometabolism.

This article briefly reviews cancer immunity and the role of gut microbiota in carcinogenesis, followed by an understanding of mechanisms by which inosine is involved in cancer immunometabolism. The immune system plays a paradoxical role in cancer treatment. Antitumor immunity depends on the T-cell priming against tumor antigens, whereas inflammatory mediators trigger the protumor signaling in the tumor microenvironment. Studies link the microbiome with metabolism and immunity-two main factors implicated in carcinogenesis. Gut microbiota has been shown to affect both antitumor immunity and protumor immune signaling. There is mounting evidence that the human microbiome can play a role in the immunotherapeutic effects, both response and resistance. Inosine-5'-monophosphate dehydrogenase (IMPDH) is a highly conservative enzyme widely expressed in mammals. Cell signaling pathways use molecular inosine, a crucial secondary metabolite in purine metabolism and a molecular messenger. Recent research has identified inosine as a critical regulator of immune checkpoint inhibition (ICI) therapeutic response in various tumor types. Some bacterial species were found to produce inosine or its metabolite hypoxanthine and induce T-helper 1 differentiation and effector functions via the inosine-A2AR-cAMP-PKA pathway upon ICI therapy. Also, inosine acts as a substitute carbon source for T-cell metabolism in glucose-restricted environments, i.e., the tumor microenvironment, assisting T-cell proliferation and differentiation while enhancing sensitivity to ICI, reinforcing the notion that inosine metabolism might contribute to antitumor immunity. Also, inosine is a potent agonist of the adenosine receptor, A2AR, and A2AR signaling can affect T-cell responses and antitumor immunity, making the inosine-A2AR pathway blockage a candidate for cancer treatment. Further research is required to investigate inosine as a cancer immunometabolism therapy.

Blue Lagoon Algae Improve Uneven Skin Pigmentation: Results from in vitro Studies and from a Monocentric, Randomized, Double-Blind, Vehicle-Controlled, Split-Face Study

Introduction: Bathing in the Blue Lagoon (BL) in Iceland benefits patients with psoriasis. Accordingly, the BL water contains algae with biological activities that improve skin barrier function and affect T-cell responses relevant for psoriasis. Bathing in the BL is also becoming increasingly popular among healthy individuals and anecdotal evidence suggests positive effects on uneven skin pigmentation. Objective: The aim of the study was to address the impact of BL algae on skin pigmentation. Methods: In this work, in vitro gene expression studies in melanocytes and a noninvasive in vivo study were conducted. Results: We here report that normal human epidermal melanocytes, which had been treated with nontoxic concentrations of BL algae, show a significantly reduced expression of α melanocyte-stimulating hormone-induced expression of genes important for melanin synthesis, such as tyrosinase, tyrosinase-related protein 1, dopachrome tautomerase, melan A protein, and pre-melanosome protein. This in vitro observation prompted us to conduct a randomized, double-blind, intra-individual, comparative split-face in vivo study, in which 60 volunteers with pre-existing facial pigment spots were treated twice daily with a BL algae containing serum or a vehicle control. We found that constitutive skin pigmentation as determined by colorimetry (individual typology angle and luminescence) did not differ significantly between vehicle- and serum-treated skin sites. In marked contrast, digital photography under cross-polarized lighting and RBX technology (VISIA CR) revealed that the number of pigment spots in the serum-treated face decreased significantly compared to the vehicle-treated side. Conclusion: Thus, BL algae can affect human melanocyte function in vitro and reduce uneven facial skin pigmentation in vivo.

Pre-existing heterologous T-cell immunity and neoantigen immunogenicity.

Neoantigens are tumor‐specific mutated proteins that are exempt from central tolerance and are therefore capable of efficiently eliciting effective T‐cell responses. The identification of immunogenic neoantigens in tumor‐specific mutated proteins has promising clinical implications for cancer immunotherapy. However, the factors that may contribute to neoantigen immunogenicity are not yet fully understood. Through molecular mimicry of antigens arising during cancer progression, the gut microbiota and previously encountered pathogens potentially have profound impacts on T‐cell responses to previously unencountered tumor neoantigens. Here, we review the characteristics of immunogenic neoantigens and how host exposure to microbes may affect T‐cell responses to neoantigens. We address the hypothesis that pre‐existing heterologous memory T‐cell immunity is a major factor that influences neoantigen immunogenicity in individual cancer patients. Accumulating data suggest that differences in individual histories of microbial exposure should be taken into account during the optimisation of algorithms that predict neoantigen immunogenicity.

Abstract A200: Targeting ICOS receptor for development of cancer immunotherapy

Abstract Recently, checkpoint blockade-mediated immune reinvigoration acts as potential strategies to cure cancer. In addition to checkpoint blockade, a number of studies have shown that immune co-stimulators also act as crucial players in the immunotherapy. Inducible co-stimulator (ICOS), a homodimeric protein that belongs to CD28 superfamily, is expressed on activated T-cells and resting memory T-cells. Upon stimulation, ICOS and its ligand ICOSL (B7RP-1, B7H-2) have been shown to affect T-cell responses via ICOS-mediated PI3-kinase signaling to enhance the T-cells’ development and activity. In addition, previous studies have shown that ICOS/ICOSL pathway may play an important role in CTLA4 inhibitor-induced tumor immunity. In this study, we aim to develop and implement the Fc-fusion recombinant ICOSL proteins to induce T-cells’ activation for therapeutic use. Our data suggested that recombinant ICOSL-Fc can enhance the proliferation and cytotoxicity effect of human T-cells by increasing INF-γ and IL10 secretion. Moreover, ICOSL-Fc can also inhibit tumor growth in xenograft animal models. Based on our study, ICOS/ICOSL pathway could be a promising therapeutic target for the immunotherapy. Citation Format: Yu-Hsun Lo, Cheng-Chou Yu, Shu-Han Yu, Tsung-Hang Hseih, Mei-Chi Chan, Tsai Shih-Chong. Targeting ICOS receptor for development of cancer immunotherapy [abstract]. In: Proceedings of the Fourth CRI-CIMT-EATI-AACR International Cancer Immunotherapy Conference: Translating Science into Survival; Sept 30-Oct 3, 2018; New York, NY. Philadelphia (PA): AACR; Cancer Immunol Res 2019;7(2 Suppl):Abstract nr A200.

Human Mesenchymal Stem Cells Impact Th17 and Th1 Responses Through a Prostaglandin E2 and Myeloid-Dependent Mechanism.

Although animal studies have generated a growing interest in the anti-inflammatory potential of mesenchymal stem cells (MSCs) for the treatment of autoimmune diseases, MSCs possess the capacity to both limit and promote immune responses. Yet relatively little is known about human-MSC modulation of human disease-implicated T-cell responses, or the mechanisms underlying such modulation. The current study reveals a novel prostaglandin E2-dependent and myeloid cell-mediated mechanism by which human MSCs can reciprocally regulate human Th17 and Th1 responses, with implications for the use of MSCs as a potential therapeutic for patients with multiple sclerosis and other immune-mediated diseases.

Type I IFN signaling triggers immunopathology in tuberculosis-susceptible mice by modulating lung phagocyte dynamics.

General interest in the biological functions of IFN type I in Mycobacterium tuberculosis (Mtb) infection increased after the recent identification of a distinct IFN gene expression signature in tuberculosis (TB) patients. Here, we demonstrate that TB-susceptible mice lacking the receptor for IFN I (IFNAR1) were protected from death upon aerogenic infection with Mtb. Using this experimental model to mimic primary progressive pulmonary TB, we dissected the immune processes affected by IFN I. IFNAR1 signaling did not affect T-cell responses, but markedly altered migration of inflammatory monocytes and neutrophils to the lung. This process was orchestrated by IFNAR1 expressed on both immune and tissue-resident radioresistant cells. IFNAR1-driven TB susceptibility was initiated by augmented Mtb replication and in situ death events, along with CXCL5/CXCL1-driven accumulation of neutrophils in alveoli, followed by the discrete compartmentalization of Mtb in lung phagocytes. Early depletion of neutrophils rescued TB-susceptible mice to levels observed in mice lacking IFNAR1. We conclude that IFN I alters early innate events at the site of Mtb invasion leading to fatal immunopathology. These data furnish a mechanistic explanation for the detrimental role of IFN I in pulmonary TB and form a basis for understanding the complex roles of IFN I in chronic inflammation.

DNA plasmid HIV vaccine design, number of doses, participant gender, and body mass index affect T-cell responses across HIV vaccine clinical trials

Background Numerous studies have evaluated individual DNA plasmid vaccines. We evaluated data from 10 HIV vaccine clinical trials that utilized common, validated immunogenicity assays to objectively investigate factors that influence DNA plasmid-induced T-cell responses. Methods This retrospective analysis included data from 1218 healthy, HIV-1 uninfected adults enrolled in 10 DNA HIV vaccine clinical trials conducted within the HIV Vaccine Trials Network. HIV-specific T-cell responses from peripheral blood mononuclear cells were measured using validated IFN-g ELISpot and intracellular cytokine staining assays. The effects of DNA vaccine HIV antigens, number of doses, gender, body mass index (BMI), and age were evaluated. Results When plasmids expressing Gag, Env and Pol were coadministered, the highest T-cell response rate was against Env (38.1%) and much less to Gag (4.6%) or Pol (4.5%). Comparing 2, 3, and 4 DNA injections, 3 vaccinations compared to 2 improved the magnitude of Env-specific CD8+ T-cell response (p=0.048), but not CD4+ T-cell responses, and a 4th vaccination had no additional effect. HIV-specific CD4+ T-cell response rates were higher in females (50.9%) than in males (27.7%, p=0.0002). Having lower BMI (p=0.025) was independently associated with higher HIV-specific CD4+ T-cell response rates. There were no significant differences in HIV-specific CD8+ T-cell response frequencies by gender or BMI. Lower HIV-specific CD4+ T-cell responses were seen with the 18-20 (36.4%) and 41-50 (23.3%) age groups compared to the 21-30 (44.0%) and 31-40 (41.%) groups, but these differences were not significant (p=0.0565) and no differences were seen in HIV-specific CD8+ T-cell responses by age. Conclusion Pooled data across DNA HIV vaccine clinical trials indicate that plasmid inserts, number of doses, gender, and BMI can affect T-cell responses. These factors should be considered in vaccine research.

The role of T‐regulatory cells and Toll‐like receptors in the pathogenesis of human inflammatory bowel disease

Two related chronic inflammatory diseases, Crohn's disease and ulcerative colitis, are together often referred to as inflammatory bowel disease (IBD). Current treatment options are not curative, and patients face lifelong therapy and debilitation. IBD is thought to be the product of a combination of genetic and environmental factors that result in the abnormal regulation of immune responses. Experimental models have demonstrated that normal CD4+ T-regulatory (Treg) cell responses and commensal bacteria are required for the maintenance of gut immune homeostasis. Recent evidence that CD4+ T cells express Toll-like receptors (TLRs) and respond directly to TLR ligands, suggests that signals from commensal bacteria may directly affect T-cell responses in the gut. In this review, we focus on evidence that defects in Treg cells may underlie IBD in humans. In addition, we discuss evidence that direct signaling via TLRs to T cells can affect IBD and that T-cell-dependent responses to bacterial proteins, such as flagellin, are central to the aetiology of this disease.

Control of peripheral T‐cell tolerance and autoimmunity via the CTLA‐4 and PD‐1 pathways

Classically, the CD28/cytotoxic T-lymphocyte antigen-4 (CTLA-4) and B7 families of cell surface molecules regulate complex signaling pathways that profoundly affect T-cell responses. The recent identification and characterization of additional CD28 and B7 family members including programmed death-1 (PD-1), programmed death ligand-1 (PD-L1) (B7-H1), and PD-L2 (B7-DC) has added to the complexity and greater appreciation of how surface molecules control T-cell activation and peripheral tolerance. CD28/B7 interactions mediate co-stimulation and significantly enhance peripheral T-cell responses. CTLA-4, in contrast, interacting with the same B7 molecules, results in decreased T-lymphocyte activity and regulates the immune response. Similarly, PD-1 interactions with PD-L1 and PD-L2 downmodulate T-cell immune responses. Despite these similarities, the regulatory roles of the CTLA-4 and PD-1 pathways are distinct. This may be due, at least in part, to the differential expression patterns of the CTLA-4 and PD-1 ligands both temporally and spatially. This article examines the role of CTLA-4 and PD-1 in limiting autoreactivity and establishing peripheral self-tolerance with the hypothesis that CTLA-4 signals are required early in the lymph node during initiation of an immune response and PD-1 pathways act late at the tissue sites to limit T-cell activity.

Mesenchymal stem cells exert differential effects on alloantigen and virus-specific T-cell responses

AbstractMesenchymal stem cells (MSCs) suppress alloantigen-induced T-cell functions in vitro and infusion of third-party MSCs seems to be a promising therapy for graft-versus-host disease (GVHD). Little is known about the specificity of immunosuppression by MSCs, in particular the effect on immunity to pathogens. We have studied how MSCs affect T-cell responses specific to Epstein-Barr virus (EBV) and cytomegalovirus (CMV). We found that EBV- and CMV-induced proliferation and interferon-γ (IFN-γ) production from peripheral blood mononuclear cells (PBMCs) was less affected by third-party MSCs than the response to alloantigen and that MSCs had no effect on expansion of EBV and CMV pentamer-specific T cells. Established EBV-specific cytotoxic T cells (CTL) or CMV-CTL cultured with MSCs retained the ability to proliferate and produce IFN-γ in response to their cognate antigen and to kill virally infected targets. Finally, PBMCs from 2 patients who received MSCs for acute GVHD showed persistence of CMV-specific T cells and retained IFN-γ response to CMV after MSC infusion. In summary, MSCs have little effect on T-cell responses to EBV and CMV, which contrasts to their strong immunosuppressive effects on alloreactive T cells. These data have major implications for immunotherapy of GVHD with MSCs and suggest that the effector functions of virus-specific T cells may be retained after MSC infusion.

Analysis of T-Cell Assays to Measure Autoimmune Responses in Subjects With Type 1 Diabetes

Type 1 diabetes is a chronic autoimmune disease mediated by autoreactive T-cells. Several experimental therapies targeting T-cells are in clinical trials. To understand how these therapies affect T-cell responses in vivo, assays that directly measure human T-cell function are needed. In a blinded, multicenter, case-controlled study conducted by the Immune Tolerance Network, we tested responses in an immunoblot and T-cell proliferative assay to distinguish type 1 diabetic patients from healthy control subjects. Peripheral blood cells from 39 healthy control subjects selected for DR4 and 23 subjects with recently diagnosed type 1 diabetes were studied. Autoantibody responses were measured in serum samples. Positive responses in both assays were more common in peripheral blood mononuclear cells from new-onset type 1 diabetic patients compared with control subjects. The proliferative, immunoblot, and autoantibody assays had sensitivities of 58, 91, and 78% with specificities of 94, 83, and 85%, respectively. When cellular assays were combined with autoantibody measurements, the sensitivity of the measurements was 75% with 100% specificity. We conclude that cellular assays performed on peripheral blood have a high degree of accuracy in discriminating responses in subjects with type 1 diabetes from healthy control subjects. They may be useful for assessment of cellular autoimmune responses involved in type 1 diabetes.

Modulation of immunogenicity and allergenicity by controlling the number of immunostimulatory oligonucleotides linked to Amb a 1

Immunostimulatory DNA sequences (ISS) are potent immunomodulators that can drive T(H)1 responses to antigens or allergens. This effect can be dramatically enhanced by direct linkage of ISS to the protein. Evaluate the effects of the number of ISS bound to the major ragweed allergen Amb a 1 on immunogenicity and allergenicity. Immunogenicity in mice and allergenicity using PBMC or sera from subjects with ragweed allergy were assayed. Both antibody induction in vivo and antibody recognition in vitro were highly sensitive to the number of ISSs linked. IgE recognition of Amb a 1 in competitive ELISA or histamine release assays was inhibited by ISS linkage and showed an inverse relationship to the number of ISSs bound. Type and magnitude of antibody induction in mice was also highly dependent on the number of ISS bound. At the highest ISS to protein ratios, antibody induction was very low. Moderate ISS to protein ratios induced high antibody responses in which IgG(2a) generally predominated. Low ISS to protein ratios produced the highest overall antibody responses in which IgG(1) predominated. In contrast, varied ISS to protein ratios did not affect T-cell responses. In both in vivo mouse studies and in vitro human PBMC studies, all ISS to protein ratios evaluated induced similar responses represented by high levels of IFN-gamma and low levels of T(H)2 cytokines. Controlling the number of ISS bound to a protein allows manipulation of antibody recognition and induction while retaining the potent T(H)1 properties of an ISS-linked protein. Immunostimulatory DNA sequence-linked Amb a 1 conjugate represents a safe, novel therapeutic approach for treating ragweed allergy.

Inhibition of T‐cell responses by CD4 on an antigen‐presenting cell

In addition to T-helper cells, CD4 is expressed on monocytes, macrophages and dendritic cells. To see whether CD4 molecules on antigen-presenting cells affect T-cell responses, we expressed CD4 on Raji cells, and compared positive and negative cells as stimulator cells for peripheral blood mononuclear cells in mixed lymphocyte reactions. We found that expression of CD4 on Raji had an inhibitory effect on production of IFN-gamma and other cytokines.

Salmonella entericaSerovar Typhimurium Infection of Dendritic Cells Leads to Functionally Increased Expression of the Macrophage-Derived Chemokine

Gene expression in murine dendritic cells (DCs) infected with green fluorescent protein-expressing Salmonella enterica serovar Typhimurium BRD509 was studied by mRNA differential display. Infected DCs were sorted from uninfected cells by flow cytometry. The mRNA expression patterns of infected and uninfected cells revealed a number of differentially expressed transcripts, which included the macrophage-derived chemokine (MDC). Up-regulation of MDC transcription in infected DCs was confirmed by Northern blotting, and the kinetics of MDC expression was examined by real-time reverse transcription-PCR, with which 31- and 150-fold increases were detected at 2 and 6 h postinfection, respectively. The increased release by DCs of MDC into culture media was detected by an enzyme-linked immunosorbent assay. The biological activity of MDC was investigated in in vitro and in vivo assays. In vitro, supernatants from S. enterica serovar Typhimurium-infected DCs were chemoattractive to T cells, and neutralization of MDC in these supernatants inhibited T-cell migration. Passive transfer of anti-MDC antibody to mice infected with BRD509 revealed that neither growth of the bacterium nor resistance of the mice to reinfection was affected and that in vivo inhibition of MDC did not affect T-cell responses, as measured by the gamma interferon ELISPOT method 3 days after challenge infection.

Imatinib inhibits the activation and proliferation of normal T lymphocytes in vitro.

The ABL tyrosine kinase inhibitor imatinib mesylate is highly effective in the treatment of CML and is increasingly used in the stem cell transplantation (SCT) setting. Since ABL-dependent intracellular signaling molecules are involved in T-cell activation, imatinib may affect T-cell responses in vivo, thus affecting T-cell function in CML patients, disrupting immune reconstitution after allogeneic SCT and/or impeding the graft-versus-leukemia effect. Here we demonstrate that imatinib inhibits PHA-induced proliferation of normal peripheral blood mononuclear cells at in vitro concentrations (1-5 micromol/l) representative of the pharmacological doses used therapeutically in vivo. The effect is not dependent on antigen-presenting cells because CD3/CD28-induced T-cell stimulation was similarly inhibited by imatinib. Dose-dependent inhibition of the proliferative response of purified CD8+ and CD4+ T lymphocytes to anti-CD3/CD28 was similarly observed and associated with reduction in IFN-gamma production. The inhibitory effect could not be ascribed to an increased rate of apoptosis but the expression of activation markers on CD3+ T cells was significantly reduced in the presence of imatinib (1-5 micromol/L). Inhibition of T-cell proliferation was reversible after removal of the drug from the cultures. Thus, imatinib inhibits T-cell proliferation in vitro, an effect that is APC-independent, reversible, and does not involve apoptosis induction.

The modulation of thymosin alpha 1 in the maturation, differentiation and function of murine bone marrow-derived dendritic cells in the absence or presence of tumor necrosis factor-alpha

Thymosin alpha 1 (Tα1) has immunomodulatory effects on T-cells, NK-cells and macrophages, but its action on dendritic cells (DCs), which are recognized as the sole professional antigen presenting cells (APCs) capable of priming naı̈ve T-cells, is poorly understood. In this study, the effect of Tα1 in vitro on murine bone marrow-derived DCs (BMDCs) maturation, differentiation, and function with or without tumor necrosis factor-alpha (TNF-α), which is one of the important inflammatory parameters, has been investigated. We have shown, through flow cytometry, ELISA and mixed leukocyte reaction (MLR), that Tα1 promoted CD4-expressed DC differentiation and the expression of activation markers, but did not influence IL-12 production and T cell-stimulatory capacity of DCs in the absence of TNFα during BMDCs maturation. Furthermore, in the presence of TNF-α, Tα1 has been shown not only to promote the expression of CD4 on MHC class II + DCs and enhance the up-regulated levels of mature markers induced by TNF-α, but also to suppress the up-regulated IL-12 production. Particularly, thus effects seen were obvious at pharmacological Tα1 concentrations. However, Tα1 did not inhibit TNF-α-induced T-cell stimulating function of DCs. This is the first reported example of a direct Tα1-DC interaction and suggests a mechanism by which Tα1 may in part affect T-cell responses by acting at the DC level and it may play an important role in the modulation of the local inflammatory responses in vivo.

Exploiting Molecular Mimicry: Defining Rules of the Game

Molecular mimicry has been touted as a mean to develop new generation of vaccines to target carbohydrate antigens on pathogens and on tumor cells. Structural and immunological rules governing molecular mimicry require definition for its successful exploitation. Of interest are the kinds of structures that peptides adopt as carbohydrate mimics, the extent to which topological or sequence similarities among peptide mimeotopes define serum cross-reactivity to carbohydrate antigens and the extent to which peptide mimeotopes affect T-cell responses. Rational design concepts can be applied to define how a peptide may mimic carbohydrate antigens, similarities in binding affinities of antibodies for carbohydrate and for peptides, how peptides can mimic core structures on otherwise dissimilar carbohydrate antigens, and how peptide mimeotopes can be used to manipulate cellular responses not achievable by carbohydrate antigens.

Quantitative differences in the expression of parentally-derived H-2 antigens in F1 hybrid mice affect T-cell responses.

Quantitative absorption with specific anti-H-2 sera has shown that the H-2Kb and H-2Dd antigens coded by the B10.A(5R) haplotype are expressed in about fourfold lower amount on the spleen cells of [B10.A(5R) X B10.A(2R)]F1 hybrids than on parental B10.A(5R) cells. In contrast, the H-2Kk and H-2Db antigens of B10.A(2R) are expressed equally on parental and F1 cells. These quantitative differences are reflected in cytotoxic T-cell (Tc-cell) function. Macrophage target cells from F1 mice are killed less efficiently than B10.A(5R) targets by alloreactive or H-2 restricted Tc cells specific for H-2Kb or H-2Dd, and spleen cells of F1 mice are less efficient stimulators of alloreactive Tc cells specific for B10.A(5R) H-2 antigens, whereas F1 and B10.A(2R) cells are equal as targets and stimulators for Tc cells recognizing B10.A(2R) H-2 antigens.