A Novel Approach for Glioblastoma Treatment by Combining Apoptosis Inducers (TMZ, MTX, and Cytarabine) with E.V.A. (Eltanexor, Venetoclax, and A1210477) Inhibiting XPO1, Bcl-2, and Mcl-1.

Adjuvant treatment for Glioblastoma Grade 4 with Temozolomide (TMZ) inevitably fails due to therapeutic resistance, necessitating new approaches. Apoptosis induction in GB cells is inefficient, due to an excess of anti-apoptotic XPO1/Bcl-2-family proteins. We assessed TMZ, Methotrexate (MTX), and Cytarabine (Ara-C) (apoptosis inducers) combined with XPO1/Bcl-2/Mcl-1-inhibitors (apoptosis rescue) in GB cell lines and primary GB stem-like cells (GSCs). Using CellTiter-Glo® and Caspase-3 activity assays, we generated dose-response curves and analyzed the gene and protein regulation of anti-apoptotic proteins via PCR and Western blots. Optimal drug combinations were examined for their impact on the cell cycle and apoptosis induction via FACS analysis, paralleled by the assessment of potential toxicity in healthy mouse brain slices. Ara-C and MTX proved to be 150- to 10,000-fold more potent in inducing apoptosis than TMZ. In response to inhibitors Eltanexor (XPO1; E), Venetoclax (Bcl-2; V), and A1210477 (Mcl-1; A), genes encoding for the corresponding proteins were upregulated in a compensatory manner. TMZ, MTX, and Ara-C combined with E, V, and A evidenced highly lethal effects when combined. As no significant cell death induction in mouse brain slices was observed, we conclude that this drug combination is effective in vitro and expected to have low side effects in vivo.

Reciprocal crosstalk between Th17 and mesothelial cells promotes metastasis-associated adhesion of ovarian cancer cells.

BACKGROUND: IL-17A and TNF synergistically promote inflammation and tumorigenesis. Their interplay and impact on ovarian carcinoma (OC) progression are, however, poorly understood. We addressed this question focusing on mesothelial cells, whose interaction with tumor cells is known to play a pivotal role in transcoelomic metastasis formation. METHODS: Flow-cytometry and immunohistochemistry experiments were employed to identify cellular sources of IL-17A and TNF. Changes in transcriptomes and secretomes were determined by bulk and single cell RNA sequencing as well as affinity proteomics. Functional consequences were investigated by microscopic analyses and tumor cell adhesion assays. Potential clinical implications were assessed by immunohistochemistry and survival analyses. RESULTS: We identified Th17 cells as the main population of IL-17A- and TNF producers in ascites and detected their accumulation in early omental metastases. Both IL-17A and its receptor subunit IL-17RC were associated with short survival of OC patients, pointing to a role in clinical progression. IL-17A and TNF synergistically induced the reprogramming of mesothelial cells towards a pro-inflammatory mesenchymal phenotype, concomitantly with a loss of tight junctions and an impairment of mesothelial monolayer integrity, thereby promoting cancer cell adhesion. IL-17A and TNF synergistically induced the Th17-promoting cytokines IL-6 and IL-1ß as well as the Th17-attracting chemokine CCL20 in mesothelial cells, indicating a reciprocal crosstalk that potentiates the tumor-promoting role of Th17 cells in OC. CONCLUSIONS: Our findings reveal a novel function for Th17 cells in the OC microenvironment, which entails the IL-17A/TNF-mediated induction of mesothelial-mesenchymal transition, disruption of mesothelial layer integrity and consequently promotion of OC cell adhesion. These effects are potentiated by a positive feedback loop between mesothelial and Th17 cells. Together with the observed clinical associations and accumulation of Th17 cells in omental micrometastases, our observations point to a potential role in early metastases formation and thus to new therapeutic options.

Subpopulations of regulatory T cells are associated with subclinical atherosclerotic plaques, levels of LDL, and cardiorespiratory fitness in the elderly.

BACKGROUND: Atherosclerosis forms the pathological basis for the development of cardiovascular disease. Since pathological processes initially develop without clinically relevant symptoms, the identification of early markers in the subclinical stage plays an important role for initiating early interventions. There is evidence that regulatory T cells (Tregs) are involved in the development of atherosclerosis. Therefore, the present study aimed to identify and investigate associations with Tregs and their subsets in a cohort of healthy elderly individuals with and without subclinical atherosclerotic plaques (SAP). In addition, various lifestyle and risk factors, such as cardiorespiratory fitness, were investigated as associated signatures. METHODS: A cross-sectional study was performed in 79 participants (male: n = 50; age = 63.6 ± 3.7 years; body mass index = 24.9 ± 3.1 kg/m²; mean ± SD) who had no previous diagnosis of chronic disease and were not taking medication. Ultrasound of the carotids to identify SAP, cardiovascular function measurement for vascular assessment and a cardiorespiratory fitness test to determine peak oxygen uptake were performed. Additionally, tests were conducted to assess blood lipids and determine glucose levels. Immunophenotyping of Tregs and their subtypes (resting (rTregs) and effector/memory (mTregs)) was performed by 8-chanel flow cytometry. Participants were categorized according to atherosclerotic plaque status. Linear and logistic regression models were used to analyze associations between parameters. RESULTS: SAP was detected in a total of 29 participants. The participants with plaque were older (64.5 ± 3.6 years vs. 62.9 ± 3.5 years) and had higher peripheral systolic blood pressure (133.8 ± 14.7 mmHg vs. 125.8 ± 10.9 mmHg). The participants with SAP were characterized by a lower percentage of rTregs (28.8% ± 10.7% vs. 34.6% ± 10.7%) and a higher percentage of mTregs (40.3% ± 14.7% vs. 30.0% ± 11.9%). Multiple logistic regression identified age (odds ratio (OR) = 1.20 (95% confidence interval (95%CI): 1.01-1.42)) and mTregs (OR = 1.05 (95%CI: 1.02-1.10)) as independent risk factors for SAP. Stepwise linear regression could reveal an association of peak oxygen uptake (ß = 0.441), low-density lipoprotein (LDL) (ß = -0.096), and SAP (ß = 6.733) with mTregs and LDL (ß = 0.104) with rTregs. CONCLUSION: While at an early stage of SAP, the total proportion of Tregs gives no indication of vascular changes, this is indicated by a shift in the Treg subgroups. Factors such as serum LDL or cardiopulmonary fitness may be associated with this shift and may also be additional diagnostic indicators. This could be used to initiate lifestyle-based preventive measures at an early stage, which may have a protective effect against disease progression.

T-cell-derived TNF-α and a cluster of immunological parameters from plasma allow a separation between SARS-CoV-2 convalescent versus vaccinated elite athletes.

Guidelines for medical clearing after SARS-CoV-2 infection in elite athletes do not include T-cell immunity aspects despite its relevance in the course of COVID-19 disease. Therefore, we aimed to analyze T-cell-related cytokines before and after in-vitro activation of CD4+ T-cells. We sampled professional indoor sports athletes at medical clearing after SARS-CoV-2 infection obtaining clinical, fitness data, and serological data including CD4+ T-cell cytokines. All data were analyzed by principal component analysis and 2 × 2 repeated measures ANOVA. CD4+ T-cells were sampled for cell culture activation with anti-CD3/anti-CD28 tetramers. At medical clearing, CD4+ T-cells from convalescent athletes secreted increased levels of TNF-α 72 h after in-vitro activation compared to vaccinated athletes. IL-18 levels in plasma were elevated and a cluster of parameters differentiated convalescent from vaccinated athletes by 13 parameters at the timepoint of medical clearing. All clinical data indicate infection is resolved, while increased TNF-α may reflect altered proportions of peripheral T-cells as a hangover of infection.

IL-17A-producing CD8+ T cells promote PDAC via induction of inflammatory cancer-associated fibroblasts.

OBJECTIVE: Pancreatic ductal adenocarcinoma (PDAC) is characterised by an abundant desmoplastic stroma composed of cancer-associated fibroblasts (CAF) and interspersed immune cells. A non-canonical CD8+ T-cell subpopulation producing IL-17A (Tc17) promotes autoimmunity and has been identified in tumours. Here, we evaluated the Tc17 role in PDAC. DESIGN: Infiltration of Tc17 cells in PDAC tissue was correlated with patient overall survival and tumour stage. Wild-type (WT) or Il17ra-/- quiescent pancreatic stellate cells (qPSC) were exposed to conditional media obtained from Tc17 cells (Tc17-CM); moreover, co-culture of Tc17-CM-induced inflammatory (i)CAF (Tc17-iCAF) with tumour cells was performed. IL-17A/F-, IL-17RA-, RAG1-deficient and Foxn1nu/nu mice were used to study the Tc17 role in subcutaneous and orthotopic PDAC mouse models. RESULTS: Increased abundance of Tc17 cells highly correlated with reduced survival and advanced tumour stage in PDAC. Tc17-CM induced iCAF differentiation as assessed by the expression of iCAF-associated genes via synergism of IL-17A and TNF. Accordingly, IL-17RA controlled the responsiveness of qPSC to Tc17-CM. Pancreatic tumour cells co-cultured with Tc17-iCAF displayed enhanced proliferation and increased expression of genes implicated in proliferation, metabolism and protection from apoptosis. Tc17-iCAF accelerated growth of mouse and human tumours in Rag1-/- and Foxn1nu/nu mice, respectively. Finally, Il17ra-expressed by fibroblasts was required for Tc17-driven tumour growth in vivo. CONCLUSIONS: We identified Tc17 as a novel protumourigenic CD8+ T-cell subtype in PDAC, which accelerated tumour growth via IL-17RA-dependent stroma modification. We described a crosstalk between three cell types, Tc17, fibroblasts and tumour cells, promoting PDAC progression, which resulted in poor prognosis for patients.

IL18 Receptor Signaling Regulates Tumor-Reactive CD8+ T-cell Exhaustion via Activation of the IL2/STAT5/mTOR Pathway in a Pancreatic Cancer Model.

Intratumoral cytotoxic CD8+ T cells (CTL) enter a dysfunctional state characterized by expression of coinhibitory receptors, loss of effector function, and changes in the transcriptional landscape. Even though several regulators of T-cell exhaustion have been identified, the molecular mechanisms inducing T-cell exhaustion remain unclear. Here, we show that IL18 receptor (IL18R) signaling induces CD8+ T-cell exhaustion in a murine pancreatic cancer model. Adoptive transfer of Il18r-/- OT-1 CD8+ CTLs resulted in enhanced rejection of subcutaneous tumors expressing ovalbumin (OVA) as a model antigen (PancOVA), compared with wild-type OT-1 CTLs. Transferred intratumoral IL18R-deficient CTLs expressed higher levels of effector cytokines TNF and IFNγ and had reduced expression of coinhibitory receptors (PD-1, TIM-3, 2B4, LAG-3) and the transcription factors Eomes and TOX. Lower expression of coinhibitory receptors and TOX on IL18R-deficient versus IL18R-sufficient CD8+ T cells were confirmed in an orthotopic KPC model. IL18R-induced T-cell exhaustion was regulated by IL2/STAT5 and AKT/mTOR pathways, as demonstrated in an in vitro exhaustion assay. Concordantly, mice deficient in NLRP3, the molecular complex activating IL18, had decreased expression of coinhibitory receptors on intratumoral T cells and similar changes in signaling pathways at the transcriptome level. Thus, molecular pathways promoting T-cell exhaustion indicate an involvement of an NLRP3-expressing tumor microenvironment, which mediates IL18 release. The Cancer Genome Atlas analysis of patients with pancreatic carcinoma showed an association between NLRP3-mediated IL18 signaling and shorter survival. These findings indicate NLRP3-mediated IL18R signaling as a regulator of intratumoral T-cell exhaustion and a possible target for immunotherapy. See related Spotlight by Stromnes, p. 400.

Lactate induces metabolic and epigenetic reprogramming of pro-inflammatory Th17 cells.

Increased lactate levels in the tissue microenvironment are a well-known feature of chronic inflammation. However, the role of lactate in regulating T cell function remains controversial. Here, we demonstrate that extracellular lactate predominantly induces deregulation of the Th17-specific gene expression program by modulating the metabolic and epigenetic status of Th17 cells. Following lactate treatment, Th17 cells significantly reduced their IL-17A production and upregulated Foxp3 expression through ROS-driven IL-2 secretion. Moreover, we observed increased levels of genome-wide histone H3K18 lactylation, a recently described marker for active chromatin in macrophages, in lactate-treated Th17 cells. In addition, we show that high lactate concentrations suppress Th17 pathogenicity during intestinal inflammation in mice. These results indicate that lactate is capable of reprogramming pro-inflammatory T cell phenotypes into regulatory T cells.

Eltanexor Effectively Reduces Viability of Glioblastoma and Glioblastoma Stem-Like Cells at Nano-Molar Concentrations and Sensitizes to Radiotherapy and Temozolomide.

Current standard adjuvant therapy of glioblastoma multiforme (GBM) using temozolomide (TMZ) frequently fails due to therapy resistance. Thus, novel therapeutic approaches are highly demanded. We tested the therapeutic efficacy of the second-generation XPO1 inhibitor Eltanexor using assays for cell viability and apoptosis in GBM cell lines and GBM stem-like cells. For most GBM-derived cells, IC50 concentrations for Eltanexor were below 100 nM. In correlation with reduced cell viability, apoptosis rates were significantly increased. GBM stem-like cells presented a combinatorial effect of Eltanexor with TMZ on cell viability. Furthermore, pretreatment of GBM cell lines with Eltanexor significantly enhanced radiosensitivity in vitro. To explore the mechanism of apoptosis induction by Eltanexor, TP53-dependent genes were analyzed at the mRNA and protein level. Eltanexor caused induction of TP53-related genes, TP53i3, PUMA, CDKN1A, and PML on both mRNA and protein level. Immunofluorescence of GBM cell lines treated with Eltanexor revealed a strong accumulation of CDKN1A, and, to a lesser extent, of p53 and Tp53i3 in cell nuclei as a plausible mechanism for Eltanexor-induced apoptosis. From these data, we conclude that monotherapy with Eltanexor effectively induces apoptosis in GBM cells and can be combined with current adjuvant therapies to provide a more effective therapy of GBM.

Selected commensals educate the intestinal vascular and immune system for immunocompetence.

BACKGROUND: The intestinal microbiota fundamentally guides the development of a normal intestinal physiology, the education, and functioning of the mucosal immune system. The Citrobacter rodentium-carrier model in germ-free (GF) mice is suitable to study the influence of selected microbes on an otherwise blunted immune response in the absence of intestinal commensals. RESULTS: Here, we describe that colonization of adult carrier mice with 14 selected commensal microbes (OMM12 + MC2) was sufficient to reestablish the host immune response to enteric pathogens; this conversion was facilitated by maturation and activation of the intestinal blood vessel system and the step- and timewise stimulation of innate and adaptive immunity. While the immature colon of C. rodentium-infected GF mice did not allow sufficient extravasation of neutrophils into the gut lumen, colonization with OMM12 + MC2 commensals initiated the expansion and activation of the visceral vascular system enabling granulocyte transmigration into the gut lumen for effective pathogen elimination. CONCLUSIONS: Consortium modeling revealed that the addition of two facultative anaerobes to the OMM12 community was essential to further progress the intestinal development. Moreover, this study demonstrates the therapeutic value of a defined consortium to promote intestinal maturation and immunity even in adult organisms. Video Abstract.

IRF4 deficiency vulnerates B-cell progeny for leukemogenesis via somatically acquired Jak3 mutations conferring IL-7 hypersensitivity.

The processes leading from disturbed B-cell development to adult B-cell progenitor acute lymphoblastic leukemia (BCP-ALL) remain poorly understood. Here, we describe Irf4-/- mice as prone to developing BCP-ALL with age. Irf4-/- preB-I cells exhibited impaired differentiation but enhanced proliferation in response to IL-7, along with reduced retention in the IL-7 providing bone marrow niche due to decreased CXCL12 responsiveness. Thus selected, preB-I cells acquired Jak3 mutations, probably following irregular AID activity, resulting in malignant transformation. We demonstrate heightened IL-7 sensitivity due to Jak3 mutants, devise a model to explain it, and describe structural and functional similarities to Jak2 mutations often occurring in human Ph-like ALL. Finally, targeting JAK signaling with Ruxolitinib in vivo prolonged survival of mice bearing established Irf4-/- leukemia. Intriguingly, organ infiltration including leukemic meningeosis was selectively reduced without affecting blood blast counts. In this work, we present spontaneous leukemogenesis following IRF4 deficiency with potential implications for high-risk BCP-ALL in adult humans.

Effects of Receptor Specificity and Conformational Stability of Influenza A Virus Hemagglutinin on Infection and Activation of Different Cell Types in Human PBMCs.

Humans can be infected by zoonotic avian, pandemic and seasonal influenza A viruses (IAVs), which differ by receptor specificity and conformational stability of their envelope glycoprotein hemagglutinin (HA). It was shown that receptor specificity of the HA determines the tropism of IAVs to human airway epithelial cells, the primary target of IAVs in humans. Less is known about potential effects of the HA properties on viral attachment, infection and activation of human immune cells. To address this question, we studied the infection of total human peripheral blood mononuclear cells (PBMCs) and subpopulations of human PBMCs with well characterized recombinant IAVs differing by the HA and the neuraminidase (NA) but sharing all other viral proteins. Monocytes and all subpopulations of lymphocytes were significantly less susceptible to infection by IAVs with avian-like receptor specificity as compared to human-like IAVs, whereas plasmacytoid dendritic cells (pDCs) and myeloid dendritic cells were equally susceptible to IAVs with avian-like and human-like receptor specificity. This tropism correlated with the surface expression of 2-3-linked sialic acids (avian-type receptors) and 2-6-linked sialic acids (human-type receptors). Despite a reduced infectivity of avian-like IAVs for PBMCs, these viruses were not less efficient than human-like IAVs in terms of cell activation as judged by the induction of cellular mRNA of IFN-α, CCL5, RIG-I, and IL-6. Elevated levels of IFN-α mRNA were accompanied by elevated IFN-α protein secretion in primary human pDC. We found that high basal expression in monocytes of antiviral interferon-induced transmembrane protein 3 (IFITM3) limited viral infection in these cells. siRNA-mediated knockdown of IFITM3 in monocytes demonstrated that viral sensitivity to inhibition by IFITM3 correlated with the conformational stability of the HA. Our study provides new insights into the role of host- and strain-specific differences of HA in the interaction of IAVs with human immune cells and advances current understanding of the mechanisms of viral cell tropism, pathogenesis and markers of virulence.

Flt3L, LIF, and IL-10 combination promotes the selective in vitro development of ESAMlow cDC2B from murine bone marrow.

The development of two conventional dendritic cells (DC) subsets (cDC1 and cDC2) and the plasmacytoid DC (pDC) in vivo and in cultures of bone marrow (BM) cells is mediated by the growth factor Flt3L. However, little is known about the factors that direct the development of the individual DC subsets. Here, we describe the selective in vitro generation of murine ESAMlow CD103- XCR1- CD172a+ CD11b+ cDC2 from BM by treatment with a combination of Flt3L, LIF, and IL-10 (collectively named as FL10). FL10 promotes common dendritic cell progenitors (CDP) proliferation in the cultures, similar to Flt3L and CDP sorted and cultured in FL10 generate exclusively cDC2. These cDC2 express the transcription factors Irf4, Klf4, and Notch2, and their growth is reduced using BM from Irf4-/- mice, but the expression of Batf3 and Tcf4 is low. Functionally they respond to TLR3, TLR4, and TLR9 signals by upregulation of the surface maturation markers MHC II, CD80, CD86, and CD40, while they poorly secrete proinflammatory cytokines. Peptide presentation to TCR transgenic OT-II cells induced proliferation and IFN-γ production that was similar to GM-CSF-generated BM-DC and higher than Flt3L-generated DC. Together, our data support that FL10 culture of BM cells selectively promotes CDP-derived ESAMlow cDC2 (cDC2B) development and survival in vitro.

Microbial short-chain fatty acids modulate CD8+ T cell responses and improve adoptive immunotherapy for cancer.

Emerging data demonstrate that the activity of immune cells can be modulated by microbial molecules. Here, we show that the short-chain fatty acids (SCFAs) pentanoate and butyrate enhance the anti-tumor activity of cytotoxic T lymphocytes (CTLs) and chimeric antigen receptor (CAR) T cells through metabolic and epigenetic reprograming. We show that in vitro treatment of CTLs and CAR T cells with pentanoate and butyrate increases the function of mTOR as a central cellular metabolic sensor, and inhibits class I histone deacetylase activity. This reprogramming results in elevated production of effector molecules such as CD25, IFN-γ and TNF-α, and significantly enhances the anti-tumor activity of antigen-specific CTLs and ROR1-targeting CAR T cells in syngeneic murine melanoma and pancreatic cancer models. Our data shed light onto microbial molecules that may be used for enhancing cellular anti-tumor immunity. Collectively, we identify pentanoate and butyrate as two SCFAs with therapeutic utility in the context of cellular cancer immunotherapy.

PRMT1 promotes the tumor suppressor function of p14ARF and is indicative for pancreatic cancer prognosis.

The p14ARF protein is a well-known regulator of p53-dependent and p53-independent tumor-suppressive activities. In unstressed cells, p14ARF is predominantly sequestered in the nucleoli, bound to its nucleolar interaction partner NPM. Upon genotoxic stress, p14ARF undergoes an immediate redistribution to the nucleo- and cytoplasm, where it promotes activation of cell cycle arrest and apoptosis. Here, we identify p14ARF as a novel interaction partner and substrate of PRMT1 (protein arginine methyltransferase 1). PRMT1 methylates several arginine residues in the C-terminal nuclear/nucleolar localization sequence (NLS/NoLS) of p14ARF . In the absence of cellular stress, these arginines are crucial for nucleolar localization of p14ARF . Genotoxic stress causes augmented interaction between PRMT1 and p14ARF , accompanied by arginine methylation of p14ARF . PRMT1-dependent NLS/NoLS methylation promotes the release of p14ARF from NPM and nucleolar sequestration, subsequently leading to p53-independent apoptosis. This PRMT1-p14ARF cooperation is cancer-relevant and indicative for PDAC (pancreatic ductal adenocarcinoma) prognosis and chemotherapy response of pancreatic tumor cells. Our data reveal that PRMT1-mediated arginine methylation is an important trigger for p14ARF 's stress-induced tumor-suppressive function.

Ush regulates hemocyte-specific gene expression, fatty acid metabolism and cell cycle progression and cooperates with dNuRD to orchestrate hematopoiesis.

The generation of lineage-specific gene expression programmes that alter proliferation capacity, metabolic profile and cell type-specific functions during differentiation from multipotent stem cells to specialised cell types is crucial for development. During differentiation gene expression programmes are dynamically modulated by a complex interplay between sequence-specific transcription factors, associated cofactors and epigenetic regulators. Here, we study U-shaped (Ush), a multi-zinc finger protein that maintains the multipotency of stem cell-like hemocyte progenitors during Drosophila hematopoiesis. Using genomewide approaches we reveal that Ush binds to promoters and enhancers and that it controls the expression of three gene classes that encode proteins relevant to stem cell-like functions and differentiation: cell cycle regulators, key metabolic enzymes and proteins conferring specific functions of differentiated hemocytes. We employ complementary biochemical approaches to characterise the molecular mechanisms of Ush-mediated gene regulation. We uncover distinct Ush isoforms one of which binds the Nucleosome Remodeling and Deacetylation (NuRD) complex using an evolutionary conserved peptide motif. Remarkably, the Ush/NuRD complex specifically contributes to the repression of lineage-specific genes but does not impact the expression of cell cycle regulators or metabolic genes. This reveals a mechanism that enables specific and concerted modulation of functionally related portions of a wider gene expression programme. Finally, we use genetic assays to demonstrate that Ush and NuRD regulate enhancer activity during hemocyte differentiation in vivo and that both cooperate to suppress the differentiation of lamellocytes, a highly specialised blood cell type. Our findings reveal that Ush coordinates proliferation, metabolism and cell type-specific activities by isoform-specific cooperation with an epigenetic regulator.

Microenvironmental Th9 and Th17 lymphocytes induce metastatic spreading in lung cancer.

Immune microenvironment plays a critical role in lung cancer control versus progression and metastasis. In this investigation, we explored the effect of tumor-infiltrating lymphocyte subpopulations on lung cancer biology by studying in vitro cocultures, in vivo mouse models, and human lung cancer tissue. Lymphocyte conditioned media (CM) induced epithelial-mesenchymal transition (EMT) and migration in both primary human lung cancer cells and cell lines. Correspondingly, major accumulation of Th9 and Th17 cells was detected in human lung cancer tissue and correlated with poor survival. Coculturing lung cancer cells with Th9/Th17 cells or exposing them to the respective CM induced EMT in cancer cells and modulated the expression profile of genes implicated in EMT and metastasis. These features were reproduced by the signatory cytokines IL-9 and IL-17, with gene regulatory profiles evoked by these cytokines partly overlapping and partly complementary. Coinjection of Th9/Th17 cells with tumor cells in WT, Rag1-/-, Il9r-/-, and Il17ra-/- mice altered tumor growth and metastasis. Accordingly, inhibition of IL-9 or IL-17 cytokines by neutralizing antibodies decreased EMT and slowed lung cancer progression and metastasis. In conclusion, Th9 and Th17 lymphocytes induce lung cancer cell EMT, thereby promoting migration and metastatic spreading and offering potentially novel therapeutic strategies.

IL-17+ CD8+ T cell suppression by dimethyl fumarate associates with clinical response in multiple sclerosis.

IL-17-producing CD8+ (Tc17) cells are enriched in active lesions of patients with multiple sclerosis (MS), suggesting a role in the pathogenesis of autoimmunity. Here we show that amelioration of MS by dimethyl fumarate (DMF), a mechanistically elusive drug, associates with suppression of Tc17 cells. DMF treatment results in reduced frequency of Tc17, contrary to Th17 cells, and in a decreased ratio of the regulators RORC-to-TBX21, along with a shift towards cytotoxic T lymphocyte gene expression signature in CD8+ T cells from MS patients. Mechanistically, DMF potentiates the PI3K-AKT-FOXO1-T-BET pathway, thereby limiting IL-17 and RORγt expression as well as STAT5-signaling in a glutathione-dependent manner. This results in chromatin remodeling at the Il17 locus. Consequently, T-BET-deficiency in mice or inhibition of PI3K-AKT, STAT5 or reactive oxygen species prevents DMF-mediated Tc17 suppression. Overall, our data disclose a DMF-AKT-T-BET driven immune modulation and suggest putative therapy targets in MS and beyond.

Intestinal development and homeostasis require activation and apoptosis of diet-reactive T cells.

The impact of food antigens on intestinal homeostasis and immune function is poorly understood. Here, we explored the impact of dietary antigens on the phenotype and fate of intestinal T cells. Physiological uptake of dietary proteins generated a highly activated CD44+Helios+CD4+ T cell population predominantly in Peyer patches. These cells are distinct from regulatory T cells and develop independently of the microbiota. Alimentation with a protein-free, elemental diet led to an atrophic small intestine with low numbers of activated T cells, including Tfh cells and decreased amounts of intestinal IgA and IL-10. Food-activated CD44+Helios+CD4+ T cells in the Peyer patches are controlled by the immune checkpoint molecule PD-1. Blocking the PD-1 pathway rescued these T cells from apoptosis and triggered proinflammatory cytokine production, which in IL-10-deficient mice was associated with intestinal inflammation. In support of these findings, our study of patients with Crohn's disease revealed significantly reduced frequencies of apoptotic CD4+ T cells in Peyer patches as compared with healthy controls. These results suggest that apoptosis of diet-activated T cells is a hallmark of the healthy intestine.