Excessive T-cell Research Articles

The Treg/Th17 Axis: A Dynamic Balance Regulated by the Gut Microbiome

T-helper 17 (Th17) and T-regulatory (Treg) cells are frequently found at barrier surfaces, particularly within the intestinal mucosa, where they function to protect the host from pathogenic microorganisms and to restrain excessive effector T-cell responses, respectively. Despite their differing functional properties, Th17 cells and Tregs share similar developmental requirements. In fact, the fate of antigen-naïve T-cells to either Th17 or Treg lineages is finely regulated by key mediators, including TGFβ, IL-6, and all-trans retinoic acid. Importantly, the intestinal microbiome also provides immunostimulatory signals, which can activate innate and downstream adaptive immune responses. Specific components of the gut microbiome have been implicated in the production of proinflammatory cytokines by innate immune cells, such as IL-6, IL-23, IL-1β, and the subsequent generation and expansion of Th17 cells. Similarly, commensal bacteria and their metabolites can also promote the generation of intestinal Tregs that can actively induce mucosal tolerance. As such, dysbiosis of the gut microbiome may not solely represent a consequence of gut inflammation, but rather shape the Treg/Th17 commitment and influence susceptibility to inflammatory bowel disease. In this review, we discuss Treg and Th17 cell plasticity, its dynamic regulation by the microbiome, and highlight its impact on intestinal homeostasis and disease.

Effect of allergen-specific immunotherapy on CD4+ T cells.

The aims of this review are to discuss the impact of chronic high-dose allergen exposure on allergen-specific CD4(+) T-cell subset during allergen-specific immunotherapy (AIT) and discuss recent advances supporting novel mechanisms for desensitization and tolerance induction during AIT. (Figure is included in full-text article.) New technologies for direct molecular and cellular analysis have now provided an unprecedented opportunity to compare the functions and phenotypes of allergen-specific T cells at a single cell level, both in the context of disease and clinical intervention. Recent studies have demonstrated that AIT may restore tolerance by transiently inducing interleukin (IL)-10 producing T cells followed by selective deletion of allergen-specific TH2 cell subset. With antigen-specific TH2 cells at the core of the allergic process in atopic individuals, the duration and dose of antigen exposure can be the driving force behind current AIT protocol. Mechanisms modulating allergen-specific CD4(+) T-cell responses may include autocrine IL-10 production to limit excessive TH2 cell effector responses, T-cell exhaustion, and preferential proallergic TH2 cell deletion allowing concurrent downregulating T-cell responses to emerge. Administration of AIT in the context of immune modulating strategies able to induce counter-regulatory immune response may lead to improved AIT with durable clinical benefit.

Abstract CN04-02: High priority immunotherapy agents in development to treat and prevent cancer

Abstract The key insight driving the development of effective cancer immunotherapy is that all cancers have mutated proteins, and that many of these mutated proteins are recognizable as foreign by the patient's own T cells. Cancers evolve and become more and more aggressive by continuously accumulating additional mutations. As cancers evolve, T cells co-evolve the capacity to recognize and kill the cancer cells by recognition of the additional mutated proteins. Unfortunately, cancer cells also co-evolve mechanisms to escape T cell. Many cancer cell escape mechanisms are similar or identical to escape mechanisms exploited by infectious disease pathogens. Equally important for cancer cell escape from immune attack is that the immune system itself is programed to dampen down and limit ongoing T-cell responses. In order to protect the body from excessive T-cell responses and autoimmunity, multiple overlapping and redundant feedback mechanisms limit the size -- and thus effectiveness -- of immune responses, including attacks against cancer. Fortunately, basic scientists have identified many of the negative influences and have invented and constructed immunotherapeutic agents that counteract the natural/intrinsic programed dampening of T-cell actions that otherwise would allow the subsequent escape of cancer cells from immune attack by T-cells. Accordingly, the rules of cancer therapy are rapidly evolving. The most important agents are the check-point inhibitors, in particular anti-PD1 and anti-PD-L1. The publication of clinical data with these agents provides a spectacular tipping point towards cancer immunotherapy. In general, subsets of patients with a chronic T cell immune or inflammatory infiltrate are likely to respond to PD1:PD-L1 checkpoint inhibitors whereas patients without an existent immune response or T cell infiltrate are unlikely to respond. PD1:PD-L1 checkpoint inhibitors are not alone. There are many other agents in development that hit their intended targets and together represent agents to boost a comprehensive variety of mechanisms essential to developing optimal immunotherapy regimens that include amongst others: • Dendritic cell activators (anti-CD40) • Dendritic cell growth factors (Flt3L) • Vaccine adjuvants (CpG, MPL/GLA, poly ICLC, resiquimod) • T-cell stimulators (anti-OX40, CD137, anti-GITR) • T-cell growth factors (IL-7, IL-15) • Immune checkpoint inhibitors (anti-PD-1, anti-PD-L1, anti-LAG3, anti-B7-H4) • Agents to neutralize or inhibit suppressive cells, cytokines, and enzymes (IDO inhibitor, anti-IL-10, anti-TGF-beta) The current major task of cancer immunotherapists is to learn how to convert anti-PD1 and anti-PD-L1 non-responders into responders. The major task for employing the same agents for cancer prevention is to learn how and when to apply the agents in a non-toxic fashion at the correct stage of tumor development. Some possibilities will be discussed. Citation Format: Martin A. “Mac” Cheever. High priority immunotherapy agents in development to treat and prevent cancer. [abstract]. In: Proceedings of the Thirteenth Annual AACR International Conference on Frontiers in Cancer Prevention Research; 2014 Sep 27-Oct 1; New Orleans, LA. Philadelphia (PA): AACR; Can Prev Res 2015;8(10 Suppl): Abstract nr CN04-02.

Genome-wide association study in an admixed case series reveals IL12A as a new candidate in Behçet disease.

IntroductionThe etiology of Behçet’s disease (BD) is unknown, but widely considered an excessive T-cell mediated inflammatory response in a genetically susceptible host. Recent genome-wide association studies (GWAS) have shown limited number of novel loci-associations. The rarity and unequal distribution of the disease prevalence amongst different ethnic backgrounds have hampered the use of GWAS in cohorts of mixed ethnicity and sufficient sample size. However, novel statistical approaches have now enabled GWAS in admixed cohorts.MethodsWe ran a GWAS on 336 BD cases and 5,843 controls. The cases consisted of Western Europeans, Middle Eastern and Turkish individuals. Participants from the Generation R study, a multiethnic birth cohort in Rotterdam, The Netherlands were used as controls. All samples were genotyped and data was combined. Linear regression models were corrected for population stratification using Genomic Principal Components and Linear Mixed Modelling. Meta-analysis was performed on selected results previously published.ResultsWe identified SNPs associated at genome-wide significant level mapping to the 6p21.33 (HLA) region. In addition to this known signal two potential novel associations on chromosomes 6 and 18 were identified, yet with low minor allele frequencies. Extended meta-analysis reveal a GWS association with the IL12A variant rs17810546 on chromosome 3.DiscussionWe demonstrate that new statistical techniques enable GWAS analyses in a limited sized cohort of mixed ethnicity. After implementation, we confirmed the central role of the HLA region in the disease and identified new regions of interest. Moreover, we validated the association of a variant in the IL2A gene by meta-analysis with previous work. These findings enhance our knowledge of genetic associations and BD, and provide further justification for pursuing collective initiatives in genetic studies given the low prevalence of this and other rare diseases.

Abstract T P182: Peripheral Immune Protein Profiles Can Differentiate Ischemic Stroke from Transient Ischemic Attack and are Correlated with Stroke Severity

Introduction: Inflammatory signals released from ischemic brain tissue recruits peripheral immune cells to the brain following ischemic stroke (IS). The mechanisms behind this crosstalk between the brain and periphery are not well understood. Studies suggest that excessive T-cell activation and migration into the brain may contribute to a greater infarct volume and increased stroke severity. The goal of the project was to evaluate the protein expression of a variety of T cell related cytokines in the peripheral blood to determine their differential expression in ischemic stroke (IS) and transient ischemic attack (TIA). Methods: Blood was drawn from IS and TIA patients within 24 hours of last known normal. Serum was processed from whole blood centrifuged within one hour of collection then frozen at -80C until analysis. Serum cytokine concentrations of 35 analytes were quantified using a Multi-Analyte Profiling Kit (R&D Systems®) on the Luminex® 100 platform. Random forest methodology and logistic regression were used to identify cytokines and demographic characteristics that distinguish stroke from TIA. Univariate analyses were performed to describe relationships between cytokine levels and NIHSS. Results: We analyzed data from n=17 IS and n=12 TIA patients, with an average age of 69 years and a median NIHSS of 3. Age, NIHSS, dyslipidemia, CD40, and CD27 were identified as important predictors of stroke versus TIA (Error rate = 0.233, sensitivity =0.765, specificity = 0.769). In IS patients CD27 and CD40 were significantly correlated with NIHSS (r=0.823, p=0.006; r=0.66; p=0.03 respectively). Conclusion: The peripheral blood protein expression of CD27 and CD40 correlates with stroke severity, and in combination with demographic data may be used to differentiate IS from TIA in the acute care setting. Future studies are needed to assess the functional role of CD27 and CD40 following stroke for potential therapeutic applications.

IL10-Deficiency in CD4+ T Cells Exacerbates the IFNγ and IL17 Response During Bacteria Induced Colitis

Background/Aims: IL10 is a key inhibitor of effector T cell activation and a mediator of intestinal homeostasis. In addition, IL10 has emerged as a key immunoregulator during infection with various pathogens, ameliorating the excessive T-cell responses that are responsible for much of the immunopathology associated with the infection. Because IL10 plays an important role in both intestinal homeostasis and infection, we studied the function of IL10 in infection-associated intestinal inflammation. Methods: Wildtype mice and mice deficient in CD4⁺ T cell-derived or regulatory T cells-derived IL10 were infected with the enteric pathogen Citrobacter (C.) rodentium and analyzed for the specific immune response and pathogloy in the colon. Results: We found that IL10 expression is upregulated in colonic tissue after infection with C. rodentium, especially in CD4⁺ T cells, macrophages and dendritic cells. Whereas the deletion of IL10 in regulatory T cells had no effect on C. rodentium induced colitis, infection of mice deficient in CD4⁺ T cell-derived IL10 exhibited faster clearance of the bacterial burden but worse colitis, crypt hyperplasia, and pathology than did WT mice. In addition, the depletion of CD4⁺ T cell-derived IL10 in infected animals was accompanied by an accelerated IFNγ and IL17 response in the colon. Conclusion: Thus, we conclude that CD4⁺ T cell-derived IL10 is strongly involved in the control of C. rodentium-induced colitis. Interference with this network could have implications for the treatment of infection-associated intestinal inflammation.

Extracellular adenosine is a therapeutic target for limiting graft-versus-host disease and enhancing the graft-versus-tumor effect against hematopoietic malignancy

Allogeneic hematopoietic stem cell transplantation is performed in patients with hematologic malignancies refractory to chemotherapy. However, its efficacy is often limited by the development of graft-versus-host disease (GVHD) secondary to the allogeneic interaction of donor T cells with host dendritic cells. On the other hand, the antihost cytotoxicity of donor T cells enhances the graft-versus-tumor (GVT) effect. Extracellular adenosine generated by CD73/ecto-5'-nucleotidase from ATP via AMP plays pleiotropic roles under physiological and pathological conditions by engaging four adenosine receptors. One study recently demonstrated that ATP released from damaged cells exacerbates GVHD by activating the P2X7 receptor on host dendritic cells. In this review, we summarize our recent findings on the immunosuppressive role of extracellular adenosine in GVHD and the GVT effect. We have shown that in MHC-mismatched bone marrow transplantation, CD73 deficiency, particularly in the recipient, enhanced GVHD severity because of excessive donor T-cell expansion. Severe GVHD was enhanced by repeated administration of a CD73 inhibitor or an adenosine receptor antagonist. A competitive engraftment assay identified endogenous A2AAR signaling in donor T cells as part of a regulatory mechanism by CD73-generated adenosine. Pharmacological inhibition of CD73 enhanced the GVT effect against B-cell lymphoma and improved survival in tumor-relapsing mice after transplantation. Along with our findings, we herein introduce a novel concept that CD73-generated adenosine counteracts the ATP-evoked allogeneic immune reaction as a negative regulatory mechanism in GVHD. Pharmacological manipulation of CD73 activity could be a therapeutic strategy to limit GVHD and to preserve the GVT effect against hematopoietic malignancy.

Expression of T-cell KV1.3 potassium channel correlates with pro-inflammatory cytokines and disease activity in ulcerative colitis

Background and aimsPotassium channels, KV1.3 and KCa3.1, have been suggested to control T-cell activation, proliferation, and cytokine production and may thus constitute targets for anti-inflammatory therapy. Ulcerative colitis (UC) is a chronic inflammatory bowel disease characterized by excessive T-cell infiltration and cytokine production. It is unknown if KV1.3 and KCa3.1 in the inflamed mucosa are markers of active UC. We hypothesized that KV1.3 and KCa3.1 correlate with disease activity and cytokine production in patients with UC. MethodsMucosal biopsies were collected from patients with active UC (n=33) and controls (n=15). Protein and mRNA expression of KV1.3 and KCa3.1, immune cell markers, and pro-inflammatory cytokines were determined by quantitative-real-time-polymerase-chain-reaction (qPCR) and immunofluorescence, and correlated with clinical parameters of inflammation. In-vitro cytokine production was measured in human CD3+ T-cells after pharmacological blockade of KV1.3 and KCa3.1. ResultsActive UC KV1.3 mRNA expression was increased 5-fold compared to controls. Immunofluorescence analyses revealed that KV1.3 protein was present in inflamed mucosa in 57% of CD4+ and 23% of CD8+ T-cells. KV1.3 was virtually absent on infiltrating macrophages. KV1.3 mRNA expression correlated significantly with mRNA expression of pro-inflammatory cytokines TNF-α (R2=0.61) and IL-17A (R2=0.51), the mayo endoscopic subscore (R2=0.13), and histological inflammation (R2=0.23). In-vitro blockade of T-cell KV1.3 and KCa3.1 decreased production of IFN-γ, TNF-α, and IL-17A. ConclusionsHigh levels of KV1.3 in CD4 and CD8 positive T-cells infiltrates are associated with production of pro-inflammatory IL-17A and TNF-α in active UC. KV1.3 may serve as a marker of disease activity and pharmacological blockade might constitute a novel immunosuppressive strategy.

PTPN2 attenuates T-cell lymphopenia-induced proliferation

When the peripheral T-cell pool is depleted, T cells undergo homoeostatic expansion. This expansion is reliant on the recognition of self-antigens and/or cytokines, in particular interleukin-7. The T cell-intrinsic mechanisms that prevent excessive homoeostatic T-cell responses and consequent overt autoreactivity remain poorly defined. Here we show that protein tyrosine phosphatase N2 (PTPN2) is elevated in naive T cells leaving the thymus to restrict homoeostatic T-cell proliferation and prevent excess responses to self-antigens in the periphery. PTPN2-deficient CD8(+) T cells undergo rapid lymphopenia-induced proliferation (LIP) when transferred into lymphopenic hosts and acquire the characteristics of antigen-experienced effector T cells. The enhanced LIP is attributed to elevated T-cell receptor-dependent, but not interleukin-7-dependent responses, results in a skewed T-cell receptor repertoire and the development of autoimmunity. Our results identify a major mechanism by which homoeostatic T-cell responses are tuned to prevent the development of autoimmune and inflammatory disorders.

Epigenetic Modification of FOXP3 in Patients With Chronic HIV Infection

HIV-1 modulates host cell epigenetic machinery to control its own replication and induce immune suppression. HIV-1 infection leads to activation of T regulatory cell (T(reg)), but the mechanism underlying this immune modulation is unclear. T(reg) plays a prominent role in gut-mucosal immune tolerance by restraining excessive effector T-cell responses, a mechanism that is known to be disturbed in chronic HIV-1 infection. DNA methylation plays a major role in T(reg) lineage commitment and immune homeostasis, which may be regulated by HIV. To investigate the mechanisms of aberrant methylation of the T(reg) marker FOXP3 in HIV-1 infection, we evaluated the expression pattern of methylation-related enzymes and its correlation to FOXP3 methylation. FOXP3 promoter methylation in the colon mucosa and peripheral blood from HIV-infected patients and control subjects was measured using Pyrosequencing. Gene expression pattern of DNA methylation enzymes in the colon mucosa was investigated by Microarray and quantitative reverse transcriptase-polymerase chain reaction analysis in the same subjects. FOXP3 promoter was significantly (P ≤ 0.0001) demethylated in HIV-infected patients compared with control subjects in both tissues. Expression of DNA methyltransferase 1 (DNAMT1), DNA methyltransferase 1-associated protein 1(DMAP1), methyltransferase-like 7B (METTL7B), and methyltransferase-like 10 (METTL10) were significantly down regulated in HIV-infected patients compared with controls and had a significant positive correlation to FOXP3 promoter methylation. We present evidence suggesting that altered methylation pattern of FOXP3 and accordingly higher T(reg) frequency in gut mucosa of HIV-infected patients may be because of aberrant methylation processing in HIV.

Perforin deficiency impairs a critical immunoregulatory loop involving murine CD8+ T cells and dendritic cells

AbstractHumans and mice with impaired perforin-dependent cytotoxic function may develop excessive T-cell activation and the fatal disorder hemophagocytic lymphohistiocytosis (HLH) after infection. Though cytotoxic lymphocytes can kill antigen-presenting cells, the physiological mechanism of perforin-mediated immune regulation has never been demonstrated in a disease-relevant context. We used a murine model of HLH to examine how perforin controls immune activation, and we have defined a feedback loop that is critical for immune homeostasis. This endogenous feedback loop involves perforin-dependent elimination of rare, antigen-presenting dendritic cells (DCs) by CD8+ T cells and has a dominant influence on the magnitude of T-cell activation after viral infection. Antigen presentation by a minor fraction of DCs persisted in T-cell– or perforin–deficient animals and continued to drive T-cell activation well beyond initial priming in the latter animals. Depletion of DCs or transfer of perforin-sufficient T cells dampened endogenous DC antigen presentation and T-cell activation, demonstrating a reciprocal relationship between perforin in CD8+ T cells and DC function. Thus, selective cytotoxic “pruning” of DC populations by CD8+ T cells limits T-cell activation and protects against the development of HLH and potentially other immunopathological conditions.

The Immunopathology of Giant Cell Arteritis

Giant cell arteritis (GCA) is an important cause of preventable blindness, most commonly due to anterior ischemic optic neuropathy. Ischemic tissue injury is the end result of a process that begins within the walls of susceptible arteries in which local dendritic cells (DCs) recruit and activate CD4 T cells that, in turn, direct the activity of effector macrophages. In response to the immune attack, the blood vessel forms lumen-stenosing intima. Multiple cascades of excessive T-cell reactivity contribute to the autoimmune features of giant cell arteritis with TH1 and TH17 immunity responsible for the early phase and TH1 immunity promoting chronic-smoldering inflammation. These cascades are only partially overlapping, supporting the concept that a multitude of instigators induce and sustain vascular inflammation. The artery actively participates in the abnormal immune response through endogenous immune sentinels, so-called vascular DCs embedded in the adventitia. Advancing age, the strongest of all risk factors for GCA, contributes to both, the dysfunction of the immune system and the vascular system. Expansion of the therapeutic armamentarium for GCA needs to focus on approaches that mitigate the impact of the aging artery and adapt to the needs of the immunosenescent host.

A decoy receptor 3 analogue reduces localised defects in phagocyte function in pneumococcal pneumonia

BackgroundTherapeutic strategies to modulate the host response to bacterial pneumonia are needed to improve outcomes during community-acquired pneumonia. This study used mice with impaired Fas signalling to examine susceptibility to...

Loss of endothelial thrombomodulin predicts response to steroid therapy and survival in acute intestinal graft-versus-host disease

Steroid-refractory graft-versus-host disease causes significant morbidity and mortality after allogeneic stem cell transplantation. The pathomechanism of steroid resistance is currently not understood, but it has been suggested that endothelial cell dysfunction plays a role. Endothelial thrombomodulin was quantified along with histological markers of epithelial damage and cytotoxic T cells in colon biopsies from 51 allografted patients, and retrospectively correlated with response to steroids and survival. Loss of endothelial thrombomodulin was the strongest predictor of response to steroids (P=0.02) and nonrelapse mortality (P=0.01) in multivariate analyses adjusting for T-cell infiltrates, histological grading, vessel density, disease status, donor type, and conditioning therapy. Our data provide evidence that at disease onset, loss of endothelial thrombomodulin expression rather than excessive T-cell infiltration associates with steroid-refractory graft-versus-host disease and mortality. Prospective histological investigations are now warranted to improve diagnosis and prognostication of this core complication of stem cell transplantation.

Hemophagocytic lymphohistiocytosis

Hemophagocytic lymphohistiocytosis (HLH) is an immune dysregulatory syndrome that is associated with underlying defects of perforin-dependent cytotoxic function. This review seeks to update readers on new scientific insights and evolving clinical concepts related to this rare but fatal disorder. Clinically, HLH is defined by severe inflammation and potentially fatal damage to a variety of organ systems including the bone marrow, liver, or brain. Recent preclinical studies have increasingly defined HLH as a syndrome of abnormal and excessive T-cell activation, which leads to toxic activation of macrophages and other innate immune cells. Although macrophages have long been suspected to be important for disease development, recent studies have for the first time demonstrated their central role in the development of inflammation-associated cytopenias. In addition to defining new therapeutic targets, these scientific insights suggest significant overlap between HLH and severe inflammation in a variety of clinical contexts. Recent clinical observations are also changing how HLH is conceptualized. Increased recognition of HLH in older children and adults, sometimes in association with classic disease-associated mutations, is challenging the traditional view of HLH as either a distinctly familial or a sporadic disorder. Recent scientific and clinical insights are expanding understanding and recognition of HLH, driving an evolution in how it is defined, and suggesting future directions for improving therapy of this disorder.

LRF Maintains HSC Homeostasis by Preventing Lymphoid-Primed LT-HSCs From Excessive Differentiation

Abstract 44Hematopoietic stem cells (HSCs) are the most primitive cells in the hematopoietic system and are under tight regulation for self-renewal and differentiation. Notch signals are essential for the emergence of definitive hematopoiesis in embryos and are critical regulators for lymphoid lineage fate determination. However, their role in adult HSC function is currently under debate.LRF (Leukemia/ Lymphoma Related Factor, also known as Zbtb7a/pokemon) is a transcriptional factor that plays a key role in lymphoid lineage fate determination, erythroid terminal differentiation and germinal center B cell proliferation. LRF loss at HSC/progenitor levels led to excessive differentiation of HSCs into T cells in the BM at the expense of B cell development. Concomitantly, the numbers of LT-HSCs (CD34−CD150+CD48−Flt3−IL7Rα−Lin−Sca-1+c-Kit+) were significantly reduced in LRFflox/floxMx1-Cre+ mice one month after LRF inactivation. Reduced LT-HSC numbers in LRFflox/floxMx1-Cre+ mice were almost completely rescued by the genetic loss of Notch1 (LRFflox/floxNotch1flox/floxMx1-Cre+) or anti-DLL4 antibody treatment, suggesting that the reduction in LT-HSCs numbers was caused by Notch1/DLL4-mediated mechanisms. Furthermore, immunohistochemical (IHC) analysis demonstrated a dramatic increase in DLL4 protein levels in BM hematopoietic cells in LRFflox/floxMx1-Cre+ mice, although the precise mechanisms for this remain unknown. To determine LRF function in HSCs, highly enriched 50 LT-HSCs were transplanted to lethally-irradiated CD45.1+ congenic recipient mice and contributions of donor-derived cells (CD45.2+) in recipients’ peripheral blood (PB) had been examined over 4 months after transplant. Compared to wild-type (WT) cells, LRF-deficient LT-HSCs barely contributed to lymphoid development (both T and B) in the recipients, while myeloid reconstitutions were largely unaffected.Using antibodies raised against the ligand binding domains of Notch1 and Notch2, we found that Notch proteins are expressed in a gradient at the most primitive CD34−LT-HSCs in adult BM. The CD34−LT-HSCs expressing Notch1 at high levels (Notch1high LT-HSCs) were susceptible to LRF inactivation and disappeared upon LRF inactivation. Only Notch1low LT-HSCs remained in the BM of LRFflox/floxMx1-Cre+ mice after pIpC injections. To elucidate the qualitative difference between Notch1high and Notch1low LT-HSCs in normal hematopoiesis, we analyzed cell cycle status, gene expression profiles and in vitro colony forming capacities at the single cell level. We found that the Notch1high LT-HSCs were in more active cell cycle as compared to Notch1low fractions. Single cell q-PCR analysis demonstrated Notch1low LT-HSCs express stem cell-related genes (e.g. Gata2, Mpl and Runx1) at higher levels compared to Notch1high LT-HSCs. Furthermore, Notch1high LT-HSCs reconstituted hematopoietic system more quickly compared to the Notch1low cells when transplanted to lethally-irradiated recipient mice. There is no difference in colony forming capacities between two LT-HSC subtypes. Since Notch1lowLT-HSCs gave rise to Notch1highLT-HSCs (and vice versa) in recipients’ BM, these two LT-HSC fractions are likely to be interchangeable.Taken together, our data suggest that the LT-HSCs expressing Notch1 at high levels (Notch1high LT-HSCs) are “lymphoid-primed”, which are susceptible to LRF loss. We propose a model in which LRF acts as a safeguard to prevent lymphoid-primed LT-HSCs from excessive T-cell differentiation in the BM micro-environment. Our study sheds a new light on the regulatory mechanisms regulating the balance between HSC self-renewal and lymphoid differentiation. Disclosures:Yan:Genentech Inc.: Employment.

Cancer immunotherapy: Progress and challenges in the clinical setting

Cancer immunologists have made enormous efforts to prove that innate and adaptive immune cells recognize tumor cells and induce tumor rejection in experimental animal models. These observations provided the rationale to study the role of the immune system in the control of tumor growth in cancer patients and to develop immunotherapeutic strategies to treat patients with cancers. Several lines of evidence suggest that anti-tumor immune responses may correlate with better clinical outcome in patients with cancers. Among them is the correlation between the presence of tumor-infiltrating T cells and the improved clinical outcome for patients with solid tumors [1–3]. In addition, a number of immunotherapies, such as high-dose IL-2 [4] and TA-specific monoclonal antibodies (mAb) [5], have provided long-term clinical benefits to a minority of cancer patients. Most importantly, the recent approval by the US FDA of the mAb ipilimumab directed against the co-inhibitory molecule CTLA-4 for patients with unresectable or metastatic melanoma represents a major breakthrough for mAb-based therapies in oncology practice [6]. The successful outcome of the randomized phase III clinical trial with ipilumumab has provided the much-needed incontrovertible clinical evidence that in humans, as in experimental animal models, the host’s immune system can control tumor growth. Furthermore, it has infused a considerable amount of optimism among tumor immunologists and clinical oncologists about the clinical potential of immunotherapy for the treatment of advanced cancers. However, there are also many examples of spontaneous or vaccine-induced TA-specific T- and B-cell immune responses that do not correlate with improved clinical status [7–9]. This discrepancy between immune and clinical responses underlines the need to better dissect the molecular and cellular events leading to tumor rejection in humans. Such an endeavor has greatly benefited from the molecular identification of TA expressed by human tumor cells, which are recognized by T cells and antibodies [5, 10, 11]. As a result, TA-specific immunotherapies have been implemented in clinical trials with molecularly defined cancer vaccines, TA-specific mAb and adoptive transfer of TA-specific T cells. Novel generations of cancer vaccines with molecularly defined TAs and potent adjuvants like toll-like receptor ligands appear to stimulate strong TA-specific T-cell responses but have shown evidence of clinical benefits in only a minority of patients with advanced cancer [7, 8, 12]. The adoptive transfer of TA-specific T cells remains technically challenging and the promising data obtained in terms of objective clinical responses and durability of responses from small monocentric clinical trials will need to be further confirmed in large multicenter clinical trials [13]. TA-specific mAb are clinically effective in a number of hematological malignancies and solid tumors and are routinely used in the clinic [5]. We now have a better understanding of the multiple mechanisms of tumor-induced immune escape, which are likely to cause the failure of the spontaneous or vaccine-induced immune responses to promote tumor regression in humans. In the tumor microenvironment, a number of negative regulators dampens anti-tumor immune responses and/or their therapeutic efficacy, including the production of cytokines (like TGF-β or IL-10), suppressive cells (regulatory T cells, myelosuppressive dendritic cells), defective antigen presentation by tumor cells (HLA or tumor antigen loss, antigen processing machinery defects), amino-acid catabolizing enzymes (indoleamine-2-3dioxygenase, arginase) and co-inhibitory pathways (like CTLA-4/CD28, PD-1/PD-L1) [14–17]. As a consequence, a number of therapies to specifically target these pathways are being developed to enhance TA-specific immune responses and to increase the likelihood of clinical benefits. In this article, commissioned to recognize National Cancer Survivors Day (the first Sunday in June each year, 5 June in 2011, see www.ncsdf.org), we will comment on the successes of immunotherapy of cancer in the clinical setting. In addition, we will discuss the challenges to optimize the use of cancer immunotherapies in the clinic.

Effects of Anti-B7.1/B7.2 Antibodies on LPS-Stimulated Macrophages

Abstract － T-cell activation depends on signals received by the T-cell receptor and CD28 co-stimulatory receptor. Since B7.1 and B7.2 molecules expressed on the surface of antigen presenting cells provide co-stimulatory signals through CD28 to T-cells, an inhibitor of CD28-B7.1/B7.2 binding has been proposed as a therapeutic agent for suppression of excessive T-cell activity. Although anti-B7.1/B7.2 antibodies are known to block B7.1 and B7.2 molecules, their effects on intracellular events in antigen presenting cells remain unclear. In this study, anti-B7.1/B7.2 antibodies decreased secretion of nitric oxide and pro-inflammatory cyto-kines such as TNF-α, IL-1β, and IL-12 in LPS-activated RAW264.7 macrophage-like cells and peritoneal macrophages. Moreover, anti-B7.1/B7.2 antibodies inhibited IκBα phosphorylation and down-regulated expression of co-stimulatory molecules including B7.1, B7.2, and PD-L1 in LPS-stimulated peritoneal macro-phages. These findings suggest that CTLA4-Ig and anti-B7.1/B7.2 antibodies may be candidates to treat chronic inflammatory diseases and autoimmune responses caused by excessive activation of both T-cells and macrophages.Keywords: Anti-B7.1/B7.2 antibodies, RAW264.7, Peritoneal macrophage, Co-stimulatory molecule, Pro- inflammatory cytokine

IL‐6: Regulator of Treg/Th17 balance

IL-6 is a pleiotropic cytokine involved in the physiology of virtually every organ system. Recent studies have demonstrated that IL-6 has a very important role in regulating the balance between IL-17-producing Th17 cells and regulatory T cells (Treg). The two T-cell subsets play prominent roles in immune functions: Th17 cell is a key player in the pathogenesis of autoimmune diseases and protection against bacterial infections, while Treg functions to restrain excessive effector T-cell responses. IL-6 induces the development of Th17 cells from naïve T cells together with TGF-beta; in contrast, IL-6 inhibits TGF-beta-induced Treg differentiation. Dysregulation or overproduction of IL-6 leads to autoimmune diseases such as multiple sclerosis (MS) and rheumatoid arthritis (RA), in which Th17 cells are considered to be the primary cause of pathology. Given the critical role of IL-6 in altering the balance between Treg and Th17 cells, controlling IL-6 activities is potentially an effective approach in the treatment of various autoimmune and inflammatory diseases. Here, we review the role of IL-6 in regulating Th17/Treg balance and describe the critical functions of IL-6 and Th17 in immunity and immune-pathology.

CD4 +CD25 + regulatory T cells reverse established allergic airway inflammation and prevent airway remodeling

CD4(+)CD25(+) regulatory T cells can inhibit excessive T-cell responses in vivo. We have previously demonstrated that prophylactic administration of CD4(+)CD25(+) regulatory T cells suppresses the development of acute allergen-induced airway inflammation in vivo. We sought to determine the effect of therapeutic transfer of CD4(+)CD25(+) regulatory T cells on established pulmonary inflammation and the subsequent development of airway remodeling. CD4(+)CD25(+) cells were transferred after the onset of allergic inflammation, and airway challenges were continued to induce chronic inflammation and airway remodeling. Administration of CD4(+)CD25(+) regulatory T cells reduced established lung eosinophilia, T(H)2 infiltration, and expression of IL-5, IL-13, and TGF-beta. Moreover, subsequent mucus hypersecretion and peribronchial collagen deposition were reduced after prolonged challenge. In contrast, transfer of CD4(+)CD25(+) regulatory T cells had no effect on established airway hyperreactivity either 7 days or 4 weeks after transfer. In this study we demonstrate for the first time that therapeutic transfer of CD4(+)CD25(+) regulatory T cells can resolve features of chronic allergen-induced inflammation and prevent development of airway remodeling.