Co-inhibitory Molecules CTLA-4 Research Articles

Abstract 7551: Combined epigenetic therapy and immune checkpoint blockade drive reshaping of the tumor microenvironment of platinum resistant ovarian cancer

Abstract Introduction: Immunotherapy has demonstrated modest effectiveness for patients with platinum-resistant ovarian cancer, which is in part related to the immunosuppressive tumor microenvironment (TME) of ovarian cancers. Epigenetic modulators in combination with immune checkpoint inhibitors (ICIs) may represent a potential way to sensitize ovarian cancer to immunotherapy by TME reprogramming. Methods: We evaluated the effect of epigenetic modulation in combination with ICI by comprehensive transcriptomic analyses of serially biopsied platinum-resistant/refractory ovarian cancer. Leveraging serial tumor samples from 30 patients that received oral azacitidine and pembrolizumab in the TRIO026 phase II clinical trial (NCT02900560), we performed bulk RNA sequencing and direct digital counting of 770 target genes using molecular barcodes (nCounter, NanoString) for 72 serial tumor samples, prior to treatment initiation and 6 weeks on-therapy. RNA sequencing of ~200 million reads for each sample was used for differential gene expression and gene set enrichment analysis (GSEA) and cell type deconvolution. Target gene expression data was normalized and analyzed for differential gene and pathway expression. Results: Differential gene expression analyses revealed an upregulation of inflammatory and cytolytic genes (IFNG, GZMA and GZMH; FDR-adjusted p=0.0184) as well as the co-inhibitory molecule CTLA-4 (FDR-adjusted p=0.0184). GSEA revealed an enrichment of IFNG response, natural killer cell mediated cytotoxicity, neutrophil degranulation, inflammatory response, and antigen processing/presentation related gene sets on-therapy (FDR-adjusted p < 0.001). Target gene expression analyses orthogonally confirmed these findings, which were also concordant with pathway scoring analyses showing an increased score for immune inflammatory pathways such as INFG signaling (Wilcoxon rank sum test p=0.01). Immune cell subset deconvolution suggested TME reshaping driven by a greater density of intra-tumoral T cells on-therapy (Wilcoxon rank sum test p=0.012). Higher IFNG, CXCL13, CXCR5 expression, CD8 T cell and NK cell density were associated with longer time on treatment, both at baseline and at the on-therapy timepoint. GSEA revealed an upregulation of interferon gamma, adaptive immunity, antigen presentation as well as conserved immune response gene sets on-therapy, for the patients attaining longer time on-treatment. Conclusion: Combination epigenetic and immunotherapy induces an inflammatory response and reshaping of the tumor microenvironment that may enhance their clinical efficacy, highlighting the therapeutic potential of utilizing epigenetic modulators as a way to sensitize platinum-resistant ovarian cancer to immune checkpoint inhibition. Citation Format: Blair V. Landon, Guneet Kaleka, Archana Balan, Julia L. Boland, Christopher Cherry, Gavin Pereira, Cynthia Zahnow, Boris Winterhoff, Stephen Baylin, Victor E. Velculescu, Gottfried E. Konecny, John A. Glaspy, Valsamo Anagnostou. Combined epigenetic therapy and immune checkpoint blockade drive reshaping of the tumor microenvironment of platinum resistant ovarian cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 7551.

Correlation between “Qing Fei San Jie Wan” and key gene CTLA-4 in lung cancer

Lung cancer is the most threatening malignant tumor to the health and life of the population. In the past fifty years, the incidence and mortality of lung cancer have increased significantly, and Chinese residents are facing a serious burden of lung cancer. The co-inhibitory molecule CTLA-4 on the surface of T cells is a glycoprotein expressed on the surface of active CD4 and CD8 positive T cells, which transmits inhibitory signals to T cells, has the effect of down-regulating or terminating T cell activation, and participates in Negative regulation of the immune system. “QingFeiSanJieWan” has the effect of enhancing the function and effect of the immune system, which is of great significance to the human body's fight against lung cancer, and has a certain promoting effect on the activation of T cells and other lymphocytes. The drug has different inhibitory effects on the expression of co-inhibitory molecule CTLA-4 on the surface of CD4+ and CD8+ T cells. Preventing CTLA-4 and then inhibiting the immune response effect of T cells, thereby enhancing the immune function, fighting against lung cancer cell antigens in the body, and providing new immunological ideas for the treatment and drug mechanism of lung cancer. Using “QingFeiSanJieWan” to treat CTLA The anti-resistance effect of -4 proves that the drug is of great significance to the therapeutic mechanism of lung cancer.

The Role of Co-Signaling Molecules in Psoriasis and Their Implications for Targeted Treatment.

Psoriasis is a chronic, systemic immune-mediated inflammatory disease manifesting in the skin, joint or both. Co-signaling molecules are essential for determining the magnitude of the T cell response to the antigen. According to the function of co-signaling molecules, they can be divided into co-stimulatory molecules and co-inhibitory molecules. The role of co-signaling molecules in psoriasis is recognized, mainly including the co-stimulatory molecules CD28, CD40, OX40, CD27, DR3, LFA-1, and LFA-3 and the co-inhibitory molecules CTLA-4, PD-1, and TIM-3. They impact the pathological process of psoriasis by modulating the immune strength of T cells, regulating the production of cytokines or the differentiation of Tregs. In recent years, immunotherapies targeting co-signaling molecules have made significant progress and shown broad application prospects in psoriasis. This review aims to outline the possible role of co-signaling molecules in the pathogenesis of psoriasis and their potential application for the treatment of psoriasis.

Review of immune checkpoint inhibitors in immuno-oncology.

Tumor cells predominantly express self-antigens and overcoming self-tolerance is the primary challenge to effective immunotherapy. Tumors also express ligands for co-inhibitory molecules on immune cells, in order to suppress anti-tumor immunity. Over a decade ago, the first antibodies generated to block the co-inhibitory molecule CTLA-4 was tested in patients with metastatic melanoma. Results from this landmark trial have informed not only the current landscape of checkpoint blockade but also the way in which immunotherapy trial outcomes are determined. Antibodies targeting PD-1 and its ligand, PD-L1, soon followed and use of these checkpoint inhibitors (ICIs) have expanded exponentially. ICI treatment has shown long-lasting clinical benefit in several tumor types and patients refractory to other treatments can often respond to ICI therapy. On the other hand, in some tumor types, the response to ICI is short-lived and tumors eventually recur. Current clinical trials are focused on enhancing anti-tumor effects through combinations of multiple ICIs with agents which cause tumor death, particularly in solid tumors, in order to enhance antigen presentation. It is also important to define which patients will respond to therapy with ICIs as over half of all patients suffer from immune-related adverse events (irAE), some of which are severe and long-lasting.

Age-related changes in the immune phenotype of human memory CD4 T cells differ between colon and peripheral blood

Abstract Gut lamina propria (LP) CD4 T cells are primarily tissue resident memory cells that are critical in maintaining gut homeostasis and responding to enteric pathogens. We established that aging altered the phenotypic and functional profiles of human small intestinal CD4 T cells. However, given site-specific differences in immune cell composition in the gut, it is difficult to extrapolate these findings to other locations. To probe the effect of aging on large intestine CD4 T cells, we used flow cytometry to analyze immune phenotypes of colon LP CD4 T cells from younger (34±2yrs mean±SEM; N=6) and older (80±2yrs, N=6) adults. Tissue-specificity of these profiles was determined by comparing to memory (CD45RA−) peripheral blood (mPB) CD4 T cells in a separate cohort of similarly aged persons (N=6/age group). Among younger samples, LP CD4 T cells expressed higher levels of co-inhibitory molecules CTLA4 and PD1, but had lower turnover (Ki67+) and IL2 receptor (CD25) expression relative to mPB CD4 T cells. Expression of activation (CD38, HLA-DR), senescence (CD57) and survival (Bcl2) markers were similar. Regarding age effects, frequencies of LP CD4 T cells (% of CD3 T cells) were similar between older and younger samples; mPB CD4 T cell frequencies were higher in older vs younger samples. Older LP CD4 T cells had lower CTLA4, PD1, CD38, CD25 and Ki67 and higher Bcl2 expression than younger LP CD4 T cells. Among mPB CD4 T cells, greater age was only associated with a higher CD57+ fraction. Thus, age effects differed between colon LP and mPB CD4 memory T cells. Loss of co-inhibitory markers, decreased turnover and increased survival of colon LP CD4 T cells may contribute to altered homeostasis, loss of protective immunity, and chronic inflammation in older people.

Age-related alterations in human gut CD4 T cell phenotype, T helper cell frequencies, and functional responses to enteric bacteria.

Intestinal lamina propria (LP) CD4 T cells play critical roles in maintaining intestinal homeostasis and in immune responses to enteric microbes, yet little is known regarding whether they contribute to age-associated intestinal immune dysfunction. In this study, we evaluated the direct ex vivo frequency, activation/inhibitory phenotype, death profiles, and in vitro functional responses of human jejunum LP CD4 T cells, including Th1, Th17, and Th22 subsets isolated from younger (<45 years) and older (>65years) persons. Expression of the co-inhibitory molecule CTLA-4 was significantly lower in older CD4 T cells, whereas expression of HLA-DR, CD38, CD57, and PD-1 were not significantly different between groups. Total CD4 T cell frequencies were similar between age groups, but lower frequencies and numbers of Th17 cells were observed directly ex vivo in older samples. Older Th17 and Th1 cells proliferated to a lesser degree following in vitro exposure to bacterial antigens vs. their younger counterparts. Levels of spontaneous cell death were increased in older CD4 T cells; however, cellular death profiles following activation did not differ based on age. Thus, small intestinal CD4 T cells from older persons have altered phenotypic and functional profiles including reduced expression of a co-inhibitory molecule, increased spontaneous cell death, and both reduced frequencies and altered functional responses of specific Th cell subsets. These changes may contribute to altered intestinal homeostasis and loss of protective gut immunity with aging.

Deficiency of programmed cell death 4 affects the balance of T cell subsets in hyperlipidemic mice

Programmed cell death 4 (Pdcd4) was found to be related to apoptosis upon first discovery. It was later found to play the role of tumor suppressor gene in a variety of tumors by inhibiting transcription and translation. Recently, it has been proposed that it may play an important role in some inflammatory diseases and in the immune response. In our previous study, deficiency of Pdcd4 was found to attenuate the formation of atherosclerotic plaques. This might be because deficiency of Pdcd4 may increase IL-10 expression and lipoautophagy by macrophages and attenuate the formation of foam cells. However, the effect of Pdcd4 on the subsets of T cells in hyperlipidemic mice still remained unclear. In the present study, results showed that Pdcd4 deficiency decreased the percentage of CD8+ T cells and increased that of regulatory T cells (Tregs) under hyperlipidemic conditions both in vitro and in vivo, which may be due to the reduced expression of co-stimulatory molecules CD28 and CD137, and the enhancive expression of co-inhibitory molecules CTLA-4. These results indicated that endogenous Pdcd4 promotes immune response mediated by T cells through regulation of the co-stimulatory molecules expression, which may contribute to the development of advanced atherosclerotic plaques. The current work provides new data to understand the role of Pdcd4 in different T cell subsets under hyperlipidemic microenvironment.

Differential expression pattern of co-inhibitory molecules on CD4+ T cells in uncomplicated versus complicated malaria

The immune response of malaria patients is a main factor influencing the clinical severity of malaria. A tight regulation of the CD4+ T cell response or the induction of tolerance have been proposed to contribute to protection from severe or clinical disease. We therefore compared the CD4+ T cell phenotypes of Ghanaian children with complicated malaria, uncomplicated malaria, asymptomatic Plasmodium falciparum (Pf) infection or no infection. Using flow cytometric analysis and automated multivariate clustering, we characterized the expression of the co-inhibitory molecules CTLA-4, PD-1, Tim-3, and LAG-3 and other molecules implicated in regulatory function on CD4+ T cells. Children with complicated malaria had higher frequencies of CTLA-4+ or PD-1+ CD4+ T cells than children with uncomplicated malaria. Conversely, children with uncomplicated malaria showed a higher proportion of CD4+ T cells expressing CD39 and Granzyme B, compared to children with complicated malaria. In contrast, asymptomatically infected children expressed only low levels of co-inhibitory molecules. Thus, different CD4+ T cell phenotypes are associated with complicated versus uncomplicated malaria, suggesting a two-sided role of CD4+ T cells in malaria pathogenesis and protection. Deciphering the signals that shape the CD4+ T cell phenotype in malaria will be important for new treatment and immunization strategies.

66P - Systematic evaluation of the immune microenvironment of neuroendocrine tumours (NET)

Background: Immunotherapy is currently being explored in many tumour types with encouraging results, but has not yet been evaluated in neuroendocrine tumours (NET). Our aim is to characterise the immune landscape of NET and determine which immune-modulatory pathways control the tumour infiltrating lymphocytes (TILs) in order to develop a rational approach for immunotherapy in this tumour type. Methods: Peripheral blood and fresh tissue was obtained from consenting patients with NET, and subjected to multicolour flow-cytometry to determine the abundance of CD8+, CD4+FoxP3- effector (CD4eff) and CD4+FoxP3+ regulatory (Treg) T cell subsets and the expression of co-inhibitory and co-stimulatory checkpoint molecules on these subsets. Additionally, matched FFPE tissue was obtained for multiparametric immunohistochemistry to investigate the distribution of the immune infiltrate. Results: Tissue from 23 NET patients including 19 mid-gut tumours (13: G1 and 6: G2) was analysed. Overall the tumours contained a higher proportion of Tregs compared with peripheral blood (8.5% vs 5%, P = 0.02) and had a CD8:Treg ratio of 18.1:24.3 respectively (P = 0.036). The co-inhibitory molecules CTLA-4 and TIM-3 showed highest expression on Tregs, while PD-1 and LAG-3 expression was similar across all T cell subsets. Co-stimulatory molecules, including ICOS, 41BB and OX-40, were also highest on Tregs, as was the recently identified co-stimulatory receptor TIGIT. Immunohistochemistry revealed that the majority of cases have <1% intratumoural CD4+ and CD8+ T cells but a higher number of peritumoural T cells from all subsets. Where present, T cells were predominantly CD8+ and intratumoural CD163+ macrophages were also identified. Conclusions: These preliminary results provide novel insight into the immune landscape of NET, and may inform the development of targeted combination immunotherapies. Initial results suggest that checkpoint molecules, such as PD1 and LAG-3, may be potential targets in this tumour type and work is ongoing to further elucidate the immunogenic potential of NET. Legal entity responsible for the study: University College London Funding: None Disclosure: All authors have declared no conflicts of interest.

A cellular platform for the evaluation of immune checkpoint molecules.

Blockade of the T cell coinhibitory molecules CTLA-4 and PD-1 has clinical utility to strengthen T cell responses. In addition to these immune checkpoints an ever-growing number of molecules has been implicated in generating coinhibitory signals in T cells. However, investigating coinhibitory molecules in primary human cells is complicated by the restricted expression and promiscuity of both coinhibitory receptors and their ligands. Here we have evaluated the potential of fluorescence-based transcriptional reporters based on the human Jurkat T cell line in conjunction with engineered T cell stimulator cell lines for investigating coinhibitory pathways. CTLA-4, PD-1, TIGIT, BTLA and 2B4 expressing reporter cells were generated and activated with T cell stimulator cells expressing cognate ligands of these molecules.All accessory molecules tested were functional in our reporter system. Engagement of CTLA-4, PD-1, BTLA and TIGIT by their ligands significantly inhibited T cell activation, whereas binding of 2B4 by CD48 resulted in enhanced responses. Mutational analysis revealed intracellular motifs that are responsible for BTLA mediated T cell inhibition and demonstrates potent reporter inhibition by CTLA-4 independent of cytoplasmic signaling motifs. Moreover, considerably higher IC50 values were measured for the CTLA-4 blocker Ipilimumab compared to the PD-1 antibody Nivolumab.Our findings show that coinhibitory pathways can be evaluated in Jurkat-based transcriptional reporters and yield novel insights on their function. Results obtained from this robust reductionist system can complement more time consuming and complex studies of such pathways in primary T cells.

What's Hot, What's New From the 2016 American Transplant Congress.

From June 11-15, 2016 the American Transplant Congress, the joint meeting of the American Society of Transplantation and the American Society of Transplant Surgeons, held its annual meeting in Boston, MA. The meeting, attended by 5200 registrants, included pre-meeting conferences, focused topic sessions, and hundreds of high-quality presentations from the transplant field. This meeting report highlights key findings from specific basic science, translational, and clinical research presentations deemed to have notable impact in thematic areas. In particular, there were a number of transformative studies indicating important advances in the understanding of alloimmunity, chronic rejection, tolerance, and organ-specific outcomes. Many of these results are discussed in the context of the published literature to showcase rapid advances in the transplant field.

Abstract 545: Differential regulation of human T-cells by TGR-1202, a novel PI3Kδ inhibitor

Abstract INTRODUCTION: TGR-1202 is a novel, next-generation PI3Kδ inhibitor presenting significant structural and pharmacological differences from prior small-molecule PI3Kδ inhibitors. TGR-1202 has high clinical efficacy in treatment of B cell malignancies with a substantially differentiated adverse event profile compared to previous PI3Kδ inhibitors, specifically concerning hepatotoxicity or colitis which have been minimal or non-existent. It has been postulated that these effects may be due to T cell immune-mediated mechanisms. We hypothesized that TGR-1202 preserves the function of the regulatory T cell (Treg) population, translating to decreased immune-mediated side effects after treatment. Here, we aimed to compare effects of clinically available PI3Kδ inhibitors on T cells with an emphasis on Tregs. METHODS: We compared activity of idelalisib, duvelisib, and TGR-1202 as single agents in vitro in isolated human T cells. Viability, apoptosis, and proliferation were determined using CellTiter Blue®, annexin V/PI and CFSE staining. CBA and qRT PCR were used to measure cytokines and transcription factors. Flow cytometry was used to detect subset ratios and surface marker expression. RESULTS: First, we observed comparable dose-dependent increases in cytotoxicity beginning at 25uM following treatment of isolated T cell populations with idelalisib, duvelisib, or TGR-1202. At this dose, apoptosis was induced between 48 and 72h. Second, all inhibitors reduced cytokine production in CD3+ T cells upon stimulation. Particularly, IFN-γ, IL-10 and IL-17a reduction was less pronounced after TGR-1202 treatment, indicating relative conservation of T cell response. All inhibitors lowered mRNA expression of T-bet (Th1), GATA-3 (Th2) and FoxP3 (Treg), however FoxP3 levels were consistently higher in TGR-1202 treated T cells. Third, we detected normal CD4:CD8 ratio and unaffected proliferative capacity of CD4+ and CD8+ subsets after drug treatment. Finally, all inhibitors decreased total percent of Tregs following stimulation (CD4+ CD25HI FoxP3+) accompanied by decreased expression of co-inhibitory molecules CTLA-4 and PD-1 on the Tregs. Interestingly, TGR-1202 significantly preserved the percent of Tregs closer to normal as well as surface expression of CTLA-4 and PD-1 on Tregs, indicating greater retention of immune checkpoint blockade and suppressive phenotype. CONCLUSIONS: We report herein that TGR-1202 affects human T cells differently than idelalisib and duvelisib. TGR-1202 sustains IL-10 production, FoxP3 mRNA expression, and maintains Treg percentage and expression of immune checkpoint molecules, suggesting relative preservation of numbers and function of Tregs. Data presented begin to provide novel insight into immune mediated cellular mechanisms responsible for lack of side effects in clinical trials of TGR-1202. In vivo models to further characterize effects on the T cell compartment are ongoing. Citation Format: Kamira K. Maharaj, John Powers, Renee Fonseca, Hari Miskin, Dave Maryanski, Eva Sahakian, Javier Pinilla-Ibarz. Differential regulation of human T-cells by TGR-1202, a novel PI3Kδ inhibitor. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 545.

Abstract AS24: Immunomodulatory antibodies for immunotherapy of ovarian cancer

Abstract Although surgical debulking plus platinum-based chemotherapy induces clinical remission in 70+ % of patients with advanced ovarian carcinoma, most of these patients relapse and novel treatments are urgently needed. Although clinical trials of various immunotherapies have not yielded significant benefit for ovarian cancer, the failures can now be better understood as an outcome of the highly immunosuppressive tumor microenvironment so that one needs to both overcome this immunosuppression and concomitantly activate tumor-destructive immunological mechanisms. One way to achieve this is to use monoclonal antibodies (mAbs) to molecules which regulate the immune response. Antibodies blocking the co-inhibitory molecules CTLA-4 or PD-1 have already demonstrated encouraging clinical activity, particularly in melanoma, and the combination of the two mAbs has improved efficacy with long-lasting remission in some patients. We have evaluated the antitumor efficacy of mAbs against multiple co-signaling molecules, both individually and in combinations, in the ID8 syngeneic mouse model of ovarian cancer. In this model, small intraperitoneal tumor nests appear about 10 days after implantation and solid tumors as well as malignant ascites develop leading to death approximately 30 days after transplantation. Intraperitoneal injections of single mAb antagonists against co-inhibitory receptors (PD-1, CTLA-4, TIM-3, and LAG-3) or agonistic mAb targeting co-stimulatory molecules (CD137, OX40, GITR, and CD40) in mice that had been transplanted 10 days earlier with ID8 cells were ineffective. However, injection of a combination of anti-CD137 and anti-PD-1 mAbs significantly prolonged survival, and addition of a mAb to CTLA-4 further enhanced the antitumor efficacy with some mice surviving tumor-free &amp;gt;150 days. The mAb combination greatly increased the number of tumor-associated CD8+ effector T cells in the peritoneal cavity and decreased the number of immunosuppressive Treg and MDSC cells with a prominent increase in the T effector to T suppressive cell ratios, and it rescued the function of exhausted T cells with a PD-1+TIM3+ or PD-1+TIM3– phenotype. Importantly, pretreatment of the ID8-bearing mice with cisplatin, a drug routinely used to treat ovarian carcinoma, synergized with the anti-CD137/anti-CTLA4 mAb combination for a long-lasting and complete tumor regression in 80% of the mice. Our findings, as well as results from preclinical studies and clinical trials at many centers, strengthen the concept of targeting co-stimulatory and co-inhibitory molecules by immunomodulatory mAbs for cancer therapy. A variety of such receptors is known, and their significance as therapeutic targets, individually or in combinations, merits preclinical characterization and clinical testing of the best candidates. For example, we have recently shown that a combination of mAbs to CD137+PD1+CTLA4+CD19 is superior to the anti-CD137+PD1+CTLA4 combination when tested in mouse melanoma and lung carcinoma models and causes permanent regression of melanomas with a surface area of 80-100 mm2 at the onset of treatment, and we are now evaluating this and other combinations in the ID8 model. Although we have not observed significant toxicity, caution must be exercised since CD137 agonistic antibodies have been observed to cause liver toxicity in clinical trials. Restricting the biodistribution of mAbs for treating ovarian carcinoma to the peritoneal cavity and entrapping them in nanoparticles, may favor efficacy and minimize systemic toxicity. “Translation” of the preclinical findings to clinical trials in ovarian cancer patients should be seriously considered. Citation Format: Min Dai, Ingegerd Hellstrom, Yuen Yee Yip, Huafeng Wei, Karl Erik Hellstrom. Immunomodulatory antibodies for immunotherapy of ovarian cancer [abstract]. In: Proceedings of the 10th Biennial Ovarian Cancer Research Symposium; Sep 8-9, 2014; Seattle, WA. Philadelphia (PA): AACR; Clin Cancer Res 2015;21(16 Suppl):Abstract nr AS24.

Editorial: T cell regulation by the environment.

T cell responses are initiated by ligation of their cognate T cell receptor by MHC loaded with antigenic peptide, but their response is carefully controlled by a myriad of environmental cues, including co-activation receptors, cytokines, nutrients, growth factors, local oxygen levels, salt concentrations, and microbiome. The complexity of the integration of signals received by T cells is only beginning to be fully understood (1). This research topic in T cell biology aims at highlighting some of the latest research on intrinsic and extrinsic signals regulating T cell responses. The ebook contains 10 articles that encompass key pathways that modulate T cell function and discuss how T cells coordinate their response to environmental cues. One of the first components regulating T cell activation is the expression and subsequent activation of surface receptors. Notably, T cell activation is governed by the co-activation of the TCR and of co-stimulatory or co-inhibitory molecules (2, 3). Expression and activation of co-inhibitory molecules, such as CTLA-4 and PD1 play a key part in turning off effector responses and the balance of expression of co-stimulatory and co-inhibitory receptors needs to be tightly regulated to ensure a proper level of T cell activation (4, 5). The review by Schneider and Rudd nicely illustrates how expression of the co-inhibitory molecule CTLA-4 is regulated, and how that affects T cell activation (6). In recent years, the importance of cellular metabolism in the activation of immune cells, in particular T cells, has been reported. T cell activation requires a change in cellular metabolism to face increased energetic demand. This is an exciting area of research showing that not only changes in metabolism are necessary for cell activation but that they are also actively involved in regulating T cell function and differentiation. This is discussed in several reviews. Craig Byersdorfer focuses on the role of fatty acid oxidation for T cell functions, and notably on its role in graft versus host disease (7). Ramsay and Cantrell discuss the importance of glucose metabolism for T cell function and highlight the role of hypoxia-inducible factor alpha and mTOR in coordinating the responses to the environmental cues (8). They also summarize the importance of the microbiome in regulating T cells, and the key role of the aryl hydrocarbon receptor in sensing microbes. Clovis Palmer and collaborators review glucose metabolism in T cells and also discuss how HIV infection modulates T cell metabolism (9). One of the central questions is how do T cells integrate the multitude of signals received? In their comprehensive review, Chapman and Chi discuss the central role of mTOR in the integration of the many signals received by the T cells that ultimately shape their response (10). Another related aspect of the topic is the impact of infection on T cells, whereby pathogens control the host response, mostly to their advantage, and are able to switch T cell responses to favor their own survival. A study by the group of Francisca Mutapi describes a novel mechanism by which helminth infection downregulates T cell activation, by lowering the level of CD3zeta chain in infected individuals (11). A review from Zaunders further illustrates how HIV infection affects the balance of the various T cell subsets and promotes regulatory T cells and T helper follicular cells (12). Diet can also influence immune homeostasis, with recent studies showing, for instance, how salt can affect Th differentiation (13). The effects of vitamins and their metabolites on T cells are also well described. Colleen Hayes’ group summarizes the latest data on the effects of Vitamin D on T cell responses, and how this can be modulated in autoimmune diseases (14). A review by Leving’s group focuses more particularly on the environmental factors that affect regulatory T cells. This includes cytokines, vitamin A and vitamin D, metabolism, and microbiome (15). Finally, can we therapeutically manipulate the environmental milieu to modulate T cell responses in humans? The study by the group of David Klatzmann highlights the effects of IL-2 in modulating T cell responses in type 1 diabetes (16). While the functions of regulatory CD4+ Tregs have been described years ago, the characterization of CD8+ Tregs is more recent. The authors notably report how injection of low doses of IL-2 modulates the levels of CD8+ Tregs in vivo, in both mice and in patients with type 1 diabetes, following their numbers and phenotype. This study illustrates how exogenous cytokines can modulate in vivo T cell responses, which may prove beneficial for autoimmune diseases. We hope that this compilation of reviews and research that provides an overview on environmental regulators of T cells will give the readers a flavor on the latest development in T cell biology. We would also like to thank all the contributors to this topic and the reviewers for their time in making this ebook possible.

The transcriptional regulator Bcl-3 impaires the function of regulatory T cells leading to the development of intestinal inflammation (BA6P.131)

Abstract The oncoprotein and transcription factor Bcl-3 (B cell leukemia-3) is a member of the IkB family of NF-κB inhibitors and was originally identified as a component of a chromosomal rearrangement in some B cell lymphomas. In contrast to other IkB proteins, Bcl-3 associates with p50 and p52 homodimers and is predominantly a nuclear protein involved in regulating nuclear NF-kB activity. To understand more about the effects of Bcl-3 on T cells, we have generated a new mouse model, where Bcl-3 is overexpressed specifically in T cells. We could show that mice overexpressing Bcl-3 solely in T cells developed spontaneous colitis accompanied by a massive cell infiltration into the colon. Interestingly, the TCRgd+ T cell population was dramatically increased in the colonic IEL population of the Bcl-3 overexpressing mice. By examing Tregs in these mice, we found that overexpression of Bcl-3 strongly impairs Treg cell function, which was accompanied by decreased expression of CD25, Foxp3 and the co-inhibitory molecule CTLA-4. Hence, the loss of suppression of Bcl-3 overexpressing Tregs might result in the development of severe spontaneous colitis. Our data emphasize a novel pathway in the maintenance of viability and suppressive function of Tregs in the gut that critically involves Bcl-3 as a negative regulator, qualifying this protein as an interesting pharmacological target for the treatment of IBD.

Co-inhibitory factor induced NK cell exhaustion and immune escape meachanism in HBV-related hepatocellular carcinoma (INC1P.400)

Abstract The effectiveness of cancer immunotherapies targeting co-inhibitory molecules CTLA-4 and PD-1 have fully validated that reversal of tumor immune tolerance is an important new pathway for the treatment of tumor. Recent evidence shows that NK cell exhaustion in HBV related HCC patients. However, the underlying mechanism remains unclear. In our study, we found that the expression of NKG2A on NK cells from peripheral blood and different tissue regions of HCC was significantly increased, which then impaired NK cell-mediated cytotoxic capacity and interferon-γ production. We further found the same inhibition and exhaustion of NK cell in the different disease-progressing groups of HCC patients. Furthermore, we confirmed that IL-10 amounts in the immune microenvironment of hepatitis and HCC were highly related to the development of HCC. This study will help to understand the mechanism of immune tolerance of NK cells induced by HCC and verify the potential value of NKG2A as a novel intervention target for cancer immunotherapy.

Expression levels of co-inhibitory molecules CTLA-4, LAG-3, PD-1 and CD39 on CD4⁺ T cells correlate with progression of non-small cell lung cancer

To detect the expression levels of co-inhibitory molecules, including CTLA-4, LAG-3, PD-1 and CD39, on CD4⁺ T cells in peripheral blood or tumor tissues from NSCLC patients and to investigate their potential internal relationships with the progression of NSCLC. Eighty-eight patients including 53 NSCLC, 17 disease control cases and 18 healthy controls were studied. All the peripheral blood and 13 cases of tumor and tumor-adjacent tissues from surgically treated NSCLC patients were obtained. The expression levels of co-inhibitory molecules CTLA-4, LAG-3, PD-1 and CD39 were assayed by flow cytometry (FCM). The ratios of CD4⁺ CTLA-4⁺ T cells, CD4⁺ LAG-3⁺ T cells, CD4⁺ PD-1⁺ T cells and CD4⁺ CD39⁺ T cells in the peripheral blood of NSCLC patients were (2.49 ± 2.43)%, (2.47 ± 3.50)%, (12.94 ± 5.96)% and (6.78 ± 5.21)%, respectively, the ratio of CD4⁺ CTLA-4⁺ T cells was significantly higher in the peripheral blood of NSCLC patients than that in the disease controls and healthy controls (P < 0.05) . The ratio of CD4(+)PD-1⁺ T cells was also highly raised in the peripheral blood of NSCLC patients than that in the healthy controls (P < 0.05). Further stratified analysis indicated that the ratio of CD4⁺ PD-1⁺ T cells was (13.21 ± 5.96)% in NSCLC patients entering stages III and IV, also significantly increased as compared with that of (11.06 ± 3.42)% in the patients undergoing stages I and II (P < 0.05). More CD4⁺ CTLA-4⁺ T cells, CD4⁺ LAG-3⁺ T cells and CD4⁺ PD-1⁺ T cells were verified in the cancer tissues (5.07 ± 2.11)%, (7.86 ± 3.24)% and (40.20 ± 18.84)%, respectively, than those in their matched peripheral blood (3.13 ± 1.01)%, (2.65 ± 1.48)% and (15.79 ± 5.69)%, (P < 0.05 for all), and especially, CD4⁺ CTLA-4⁺ T cells and CD4⁺ PD-1⁺ T cells were also highly increased than those in matched cancer-adjacent tissues (P < 0.05 for all). The increased expression levels of co-inhibitory molecules CTLA-4, LAG-3 and PD-1 on CD4⁺ T cells in peripheral blood and tumor tissues may be one of the mechanisms related to immune escape of tumor cells, acceleration of disease progression and poor prognosis in NSCLC patients.

The transcription factor Bcl-3 represses the function of regulatory T cells leading to the development of colitis (BA15P.223)

Abstract The proto-oncogene Bcl-3 (B cell leukemia-3) is a member of the IκB family of NF-κB inhibitors and was originally identified as a component of a chromosomal rearrangement in some B cell lymphomas. In contrast to other IκB proteins, Bcl-3 associates with p50 and p52 homodimers and is predominantly a nuclear protein involved in regulating nuclear NF-κB activity. To understand more about the effects of Bcl-3 on T cells, we have generated a new mouse model, where Bcl-3 is overexpressed specifically in T cells. We could show that mice overexpressing Bcl-3 solely in T cells developed spontaneous colitis accompanied by a massive cell infiltration into the colon. Interestingly, by analyzing regulatory T cells we could demonstrate that these cells were not able to suppress an inflammatory response. Due to the suppressive dysfunction, Tregs of Bcl-3 overexpressing mice expressed less CD25, Foxp3 as well as less of the co-inhibitory molecule CTLA-4. Hence, the loss of suppression of Bcl-3 overexpressing Tregs might result in the development of severe spontaneous colitis. Our data emphasize a novel pathway in the maintenance of viability and suppressive function of Tregs in the gut that critically involves Bcl-3 as a negative regulator, qualifying this protein as an interesting pharmacological target for the treatment of IBD.

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.

Loss of IL-17–Producing CD8 T Cells during Late Chronic Stage of Pathogenic Simian Immunodeficiency Virus Infection

Progressive disease caused by pathogenic SIV/HIV infections is marked by systemic hyperimmune activation, immune dysregulation, and profound depletion of CD4(+) T cells in lymphoid and gastrointestinal mucosal tissues. IL-17 is important for protective immunity against extracellular bacterial infections at mucosa and for maintenance of mucosal barrier. Although IL-17-secreting CD4 (Th17) and CD8 (Tc17) T cells have been reported, very little is known about the latter subset for any infectious disease. In this study, we characterized the anatomical distribution, phenotype, and functional quality of Tc17 and Th17 cells in healthy (SIV-) and SIV+ rhesus macaques. In healthy macaques, Tc17 and Th17 cells were present in all lymphoid and gastrointestinal tissues studied with predominance in small intestine. About 50% of these cells coexpressed TNF-α and IL-2. Notably, ∼50% of Tc17 cells also expressed the co-inhibitory molecule CTLA-4, and only a minority (<20%) expressed granzyme B suggesting that these cells possess more of a regulatory than cytotoxic phenotype. After SIV infection, unlike Th17 cells, Tc17 cells were not depleted during the acute phase of infection. However, the frequency of Tc17 cells in SIV-infected macaques with AIDS was lower compared with that in healthy macaques demonstrating the loss of these cells during end-stage disease. Antiretroviral therapy partially restored the frequency of Tc17 and Th17 cells in the colorectal mucosa. Depletion of Tc17 cells was not observed in colorectal mucosa of chronically infected SIV+ sooty mangabeys. In conclusion, our results suggest a role for Tc17 cells in regulating disease progression during pathogenic SIV infection.