BACKGROUND: Checkpoint inhibitors have recently been approved for the treatment of patients with hepatocellular carcinoma (HCC). However, biomarkers, which will help identify patients responding to therapy, are missing. We recently tested the combination of anti-CTLA4 treatment (tremelimumab) with loco-regional therapy in patients with HCC and reported a partial response rate of 26%.
METHODS: Here, we report updated survival analyses and results from our immune monitoring studies on peripheral blood mononuclear cells (PBMCs) and tumors from these patients.
RESULTS: Tremelimumab therapy increased CD4
CONCLUSION: In summary, we observed a clear activation of T cell responses in HCC patients treated with tremelimumab and identified potential biomarkers which will help identify patients responding to immunotherapy with anti-CTLA4.

A better characterization of T-cell subsets in the microenvironment of classical Hodgkin lymphoma (cHL) would help to develop immunotherapies. Using multicolor flow cytometry, we identified in 6 of 43 cHL tissue samples a previously unrecognized subset of CD8 T cells coexpressing CXCR5 and inducible T-cell costimulator (ICOS) molecules (CD8

Cutaneous T-cell lymphoma (CTCL) develops from clonally expanded CD4

Background: Positive and negative co-stimulatory molecules are important factors determining the outcome of immune responses to the presence of tumors. Since co-stimulatory molecule expression may be affected by gene polymorphisms, we aimed to investigate associations between variants of PD.1 and ICOS and susceptibility to colon cancer. Material and methods: ICOS (-693A/G), ICOS (+1720C/T) and PD.1 (-538G/A) gene polymorphisms were evaluated by the PCR-RFLP method in 76 colon cancer patients and 73 healthy controls. Results: The frequencies of the GG genotype and the G allele at position -693 of the ICOS gene were significantly higher in the patient group (P=0.014 and p=0.0002), while the AA genotype was significantly more common in controls (P=0.0016). At position -538 of PD.1, GG genotype and G allele frequencies were higher in the patient group (P<0.0001and P<0.0001). Again, AA and also AG genotypes significantly predominated in controls (P<0.0001 and P=0.012). Regarding genotypes and alleles of ICOS at position +1720. Frequencies of GCG and GTG haplotypes were higher in patients compared to those of controls (P=0.016 and P<0.0001), while, frequencies of GTA, ATA and ATG haplotypes were higher in controls (P=0.0017, P<0.0001 and P=0.015). GTG/GTG and GTG/GCG double haplotypes were more frequent in patients compared to controls (P=0.0147 and P=0.0071). Conclusion: Our study clarified that PD.1 (-538G/A) and ICOS (-693A/G) gene polymorphisms can be considered as genetic risk factors for the development of colon cancer among Iranian patients.

Polymorphisms within a gene's 3'-UTR may modulate posttranscriptional regulation of gene expression, and may explain individual sensitivity of chemotherapy. To investigate the correlation between single nucleotide polymorphisms (SNPs) in 3'-UTRs of B7/CD28 family genes and the response of capecitabine-based chemotherapy in colon cancer, 16 SNPs were identified in 274 advanced colon cancer patients. Statistical analysis indicated that ICOS rs1559931, rs4404254, and rs4675379 were in complete linkage disequilibrium and significantly associated with chemotherapy response. Heterozygous patients with rs1559931 G/A (31.34% vs 48.29%; P = 0.016), rs4404254 T/C (30.43% vs 48.77%; P = 0.011), or rs4675379 G/C (28.13% vs 49.04%; P = 0.004) genotypes showed poorer response to chemotherapy compared to wildtype patients. Moreover, three SNPs, including ICOSL rs15927, ICOSL rs3804033 and CD28 rs3181113, were significantly associated with the occurrence of side effects of chemotherapy. In addition, patients with ICOSL rs15927 G/G (78.26%), ICOSL rs3804033 G/G (76.00%), or CD28 rs3181113 T/T (82.05%) were more prone to enduring adverse events compared to patients bearing other polymorphisms. Taken together, our findings demonstrated that polymorphisms in the 3'-UTRs of genes in the ICOS/CD28-ICOSL pathway may influence the efficacy and occurrence of adverse events of capecitabine-based chemotherapy in advanced colon cancer patients.

BACKGROUND: This study determined the regulatory effects of inducible T-cell co-stimulators (ICOS) in human hepatocellular carcinoma HepG2 cells using a RNA interference (RNAi) technique.
METHODS: A RNAi technique was used to knockdown the expression of ICOS. ICOS expression after knockdown was detected as mRNA and protein levels by RT-PCR and Western blot, respectively. A MTT colorimetric assay was used to detect cell proliferation, and the Transwell assay was used to detect cell invasion. Western blot was carried out to detect the level of Bcl-2, AKT, and PI3K protein expression in different groups.
RESULTS: The proliferation of HepG2 cells were significantly decreased after ICOS siRNA transfection (EG group). Similarly, the results of the Transwell experiment showed that invasion of HepG2 cells in the EG group was clearly reduced compared to the negative control (NC) and blank control groups (CON). Western blot analysis showed that knockdown of ICOS expression reduced the levels of Bcl-2 and AKT, and also significantly up-regulated the level of PI3K phosphorylation (P < 0.01).
CONCLUSION: Down-regulating ICOS expression in HepG2 cells suppressed cell proliferation and invasion. The underlying mechanism may be related to the expression of the downstream factor, PI3K/AKT.

Angioimmunoblastic T-cell lymphoma (AITL) has been classified as a subtype of mature T-cell neoplasms. The recent revision of the WHO classification proposed a new category of nodal T-cell lymphoma with follicular helper T (TFH)-cell phenotype, which was classified into three diseases: AITL, follicular T-cell lymphoma, and nodal peripheral T-cell lymphoma with TFH phenotype. These lymphomas are defined by the expression of TFH-related antigens, CD279/PD-1, CD10, BCL6, CXCL13, ICOS, SAP, and CXCR5. Although recurrent mutations in TET2, IDH2, DNMT3A, RHOA, and CD28, as well as gene fusions, such as ITK-SYK and CTLA4-CD28, were not diagnostic criteria, they may be considered as novel criteria in the near future. Notably, premalignant mutations, tumor-specific mutations, and mutations specific to tumor-infiltrating B cells were identified in AITL. Thus, multi-step and multi-lineage genetic events may lead to the development of AITL.

The first generation of immune checkpoint inhibitors (anti-CTLA-4 and anti-PD-1/PD-L1) targeted natural immune homeostasis pathways, co-opted by cancers, to drive anti-tumor immune responses. These agents led to unprecedented results in patients with previously incurable metastatic disease and may become first-line therapies for some advanced cancers. However, these agents are efficacious in only a minority of patients. Newer strategies are becoming available that target additional immunomodulatory mechanisms to activate patients' own anti-tumor immune responses. Herein, we present a succinct summary of emerging immune targets with reported pre-clinical efficacy that have progressed to active investigation in clinical trials. These emerging targets include co-inhibitory and co-stimulatory markers of the innate and adaptive immune system. In this review, we discuss: 1) T lymphocyte markers: Lymphocyte Activation Gene 3 [LAG-3], T-cell Immunoglobulin- and Mucin-domain-containing molecule 3 [TIM-3], V-domain containing Ig Suppressor of T cell Activation [VISTA], T cell ImmunoGlobulin and ITIM domain [TIGIT], B7-H3, Inducible T-cell Co-stimulator [ICOS/ICOS-L], CD27/CD70, and Glucocorticoid-Induced TNF Receptor [GITR]; 2) macrophage markers: CD47/Signal-Regulatory Protein alpha [SIRPα] and Indoleamine-2,3-Dioxygenase [IDO]; and 3) natural killer cell markers: CD94/NKG2A and the Killer Immunoglobulin-like receptor [KIR] family. Finally, we briefly highlight combination strategies and potential biomarkers of response and resistance to these cancer immunotherapies.

Background: Immune checkpoint (ICK) expression might represent a surrogate measure of tumor-infiltrating T cell (CTL) exhaustion and therefore be a more accurate prognostic biomarker for colorectal cancer (CRC) patients than CTL enumeration as measured by the Immunoscore.
Methods: The expression of ICKs, Th1, CTLs, cytotoxicity-related genes, and metagenes, including Immunoscore-like metagenes, were evaluated in three independent cohorts of CRC samples (260 microsatellite instable [MSI], 971 non-MSI). Their associations with patient survival were analyzed by Cox models, taking into account the microsatellite instability (MSI) status and affiliation with various Consensus Molecular Subgroups (CMS). PD-L1 and CD8 expression were examined on a subset of tumors with immunohistochemistry. All statistical tests were two-sided.
Results: The expression of Immunoscore-like metagenes was statistically significantly associated with improved outcome in non-MSI tumors displaying low levels of both CTLs and immune checkpoints (ICKs; CMS2 and CMS3; hazard ratio [HR] = 0.63, 95% confidence interval [CI] = 0.43 to 0.92, P = .02; and HR = 0.55, 95% CI = 0.34 to 0.90, P = .02, respectively), but clearly had no prognostic relevance in CRCs displaying higher levels of CTLs and ICKs (CMS1 and CMS4; HR = 0.46, 95% CI = 0.10 to 2.10, P = .32; and HR = 1.13, 95% CI = 0.79 to 1.63, P = .50, respectively), including MSI tumors. ICK metagene expression was statistically significantly associated with worse prognosis independent of tumor staging in MSI tumors (HR = 3.46, 95% CI = 1.41 to 8.49, P = .007). ICK expression had a negative impact on the proliferation of infiltrating CD8 T cells in MSI neoplasms (median = 0.56 in ICK low vs median = 0.34 in ICK high, P = .004).
Conclusions: ICK expression cancels the prognostic relevance of CTLs in highly immunogenic colon tumors and predicts a poor outcome in MSI CRC patients.

Tumor-infiltrating immune cells are closely associated with clinical outcome. However, immunohistochemistry-based analysis of tumor infiltrates can be misleading as the representative marker of an immune subpopulation might be expressed in other cell types. In this study, based on a metagene approach (known as CIBERSORT) and an online databse, The Cancer Immunome Atlas (https://tcia.at/), we comprehensively analyzed the tumor-infiltrating immune cells present in lung adenocarcinoma (LUAD) and lung squamous cell carcinoma (LUSC). A total of 22 types of both adaptive and innate tumor-infiltrating immune cells were evaluated in LUAD (n=492) and LUSC (n=488). As a result, tumors lacking memory B cells or with increased number of M0 macrophages were associated with the poor prognosis in LUAD at early clinical stage. In LUSC, T follicular helper cells were associated with favorable outcome, while increased number of neutrophils predicted a poor outcome. Moreover, Kaplan-Meier analysis of the prognostic value of immune checkpoint molecules revealed that expression of ICOS was positively correlated the clinical outcome of patients with LUAD. Collectively, our data suggest that tumor-infiltrating immune cells in lung cancer are likely to be important determinants of both prognosis and response to immunotherapies.

BACKGROUND: Tremelimumab is an antibody that blocks CTLA-4 and demonstrates clinical efficacy in a subset of advanced melanoma patients. An unmet clinical need exists for blood-based response-predictive gene signatures to facilitate clinically effective and cost-efficient use of such immunotherapeutic interventions.
METHODS: Peripheral blood samples were collected in PAXgene® tubes from 210 treatment-naïve melanoma patients receiving tremelimumab in a worldwide, multicenter phase III study (discovery dataset). A central panel of radiologists determined objective response using RECIST criteria. Gene expression for 169 mRNA transcripts was measured using quantitative PCR. A 15-gene pre-treatment response-predictive classifier model was identified. An independent population (N = 150) of refractory melanoma patients receiving tremelimumab after chemotherapy enrolled in a worldwide phase II study (validation dataset). The classifier model, using the same genes, coefficients and constants for objective response and one-year survival after treatment, was applied to the validation dataset.
RESULTS: A 15-gene pre-treatment classifier model (containing ADAM17, CDK2, CDKN2A, DPP4, ERBB2, HLA-DRA, ICOS, ITGA4, LARGE, MYC, NAB2, NRAS, RHOC, TGFB1, and TIMP1) achieved an area under the curve (AUC) of 0.86 (95% confidence interval 0.81 to 0.91, p < 0.0001) for objective response and 0.6 (95% confidence interval 0.54 to 0.67, p = 0.0066) for one-year survival in the discovery set. This model was validated in the validation set with AUCs of 0.62 (95% confidence interval 0.54 to 0.70 p = 0.0455) for objective response and 0.68 for one-year survival (95% confidence interval 0.59 to 0.75 p = 0.0002).
CONCLUSIONS: To our knowledge, this is the largest blood-based biomarker study of a checkpoint inhibitor, tremelimumab, which demonstrates a validated pre-treatment mRNA classifier model that predicts clinical response. The data suggest that the model captures a biological signature representative of genes needed for a robust anti-cancer immune response. It also identifies non-responders to tremelimumab at baseline prior to treatment.

Immune-checkpoint blockade is able to achieve durable responses in a subset of patients; however, we lack a satisfying comprehension of the underlying mechanisms of anti-CTLA-4- and anti-PD-1-induced tumor rejection. To address these issues, we utilized mass cytometry to comprehensively profile the effects of checkpoint blockade on tumor immune infiltrates in human melanoma and murine tumor models. These analyses reveal a spectrum of tumor-infiltrating T cell populations that are highly similar between tumor models and indicate that checkpoint blockade targets only specific subsets of tumor-infiltrating T cell populations. Anti-PD-1 predominantly induces the expansion of specific tumor-infiltrating exhausted-like CD8 T cell subsets. In contrast, anti-CTLA-4 induces the expansion of an ICOS

Combination therapies have the potential to improve outcomes in melanoma patients but have not yet been clinically efficacious. Here, we used high-throughput flow cytometry-based screening to identify and characterize candidate therapies that might synergize with and augment T-cell immunotherapy efficacy. Two lead therapies, regorafenib (Reg) and NU7441, were selected based on their ability to alter a variety of immunomodulatory proteins, including CD55, CD73, CD155, programmed death-ligand 1 (PD-L1), nerve growth factor receptor (NGFR), and HLA class I in a heterogeneous panel of melanomas. The therapies also upregulated several melanoma antigens, inhibited proliferation, and perturbed activation of oncogenic signaling pathways in melanomas. T cells treated with the therapies proliferated normally and exhibited a favorably altered phenotype, including increased CD25, CD28, inducible T-cell costimulator (ICOS), and reduced expression of coinhibitory receptors. Cytokine production was also increased in treated T cells. When administered in mice, REg suppressed melanoma progression in a CD8

Adult T-cell leukemia/lymphoma (ATL) is a peripheral T-cell neoplasm with a dismal prognosis. It is caused by human T-cell leukemia virus type-1 (HTLV-1) retrovirus. A long latency period from HTLV-1 infection to ATL onset suggests that not only HTLV-1 proteins, such as Tax and HBZ, but also additional genetic and/or epigenetic events are required for ATL development. Although many studies have demonstrated the biological functions of viral genes, alterations of cellular genes associated with ATL have not been fully investigated. Recently, a large-scale integrated genetic analysis revealed the entire landscape of somatic aberrations in ATL. This neoplasm is characterized by frequent gain-of-function alterations in components of the T-cell receptor/NF-κB signaling pathway, including activating mutations in the PLCG1, PRKCB, CARD11 and VAV1 genes, and CTLA4-CD28 and ICOS-CD28 fusions. Importantly, molecules associated with immune surveillance, such as HLA-A/B, CD58 and FAS, are affected recurrently. Among them, one notable lesion occurs as frequent structural variations that truncate the PD-L1 3'-untranslated region, leading to its overexpression. Other genetic targets include transcription factors (IRF4, IKZF2, and GATA3) and chemokine receptors (CCR4, CCR7 and GPR183), which are functionally relevant in normal T cells. A substantial proportion of ATL cases show widespread accumulation of repressive epigenetic changes, such as trimethylation of histone H3 lysine 27 and DNA hypermethylation of CpG islands, which coordinately modulate multiple pathways, including Cys2-His2 zinc finger genes involved in silencing retroelements. Here we review the current understanding of the genetic/epigenetic aberrations in ATL, focusing on their relevance in its molecular pathogenesis.

Previous immunohistochemical (IHC) studies showed controversial data about the prognostic value of tumor-infiltrating lymphocytes (TILs) in follicular lymphoma (FL). To clarify this issue, a large series of FL samples from rituximab-treated patients enrolled in the randomized PRIMA trial was examined. IHC was quantified using automated image analysis in 417, 287, 418, 406, 379, and 369 patients for CD3, CD4, CD8, PD1, ICOS, and FOXP3, respectively. RNAseq analysis was used to quantify TIL-related mRNA transcripts from 148 patients. When each IHC marker was used as a continuous variable in the whole cohort, high CD3 counts were associated with better progression-free survival (PFS) (P = .025). When an optimal IHC cut point was applied to the whole patient population, high CD3 counts and high PD1 counts were associated with better PFS (P = .011 and P = .044, respectively), whereas none of the other TIL markers had any significant correlation with outcome. When a stringent analysis was performed by dividing the whole cohort into a training set and a validation set, none of the TIL markers showed a prognostic significance in both groups. RNAseq analysis showed a significant correlation between high levels of CD3 and CD8 transcripts and better PFS (P = .001 and P = .037, respectively). No prognostic correlation was found as to the level of other immune gene transcripts. These results suggest that the IHC prognostic value of TILs is circumvented by rituximab treatment, although there is a trend for high numbers of CD3+ TILs to correlate with better PFS.

Recent success in cancer immunotherapy (anti-CTLA-4, anti-PD1/PD-L1) has confirmed the hypothesis that the immune system can control many cancers across various histologies, in some cases producing durable responses in a way not seen with many small-molecule drugs. However, only less than 25% of all patients do respond to immuno-oncology drugs and several resistance mechanisms have been identified (e.g. T-cell exhaustion, overexpression of caspase-8 and β-catenin, PD-1/PD-L1 gene amplification, MHC-I/II mutations). To improve response rates and to overcome resistance, novel second- and third-generation immuno-oncology drugs are currently evaluated in ongoing phase I/II trials (either alone or in combination) including novel inhibitory compounds (e.g. TIM-3, VISTA, LAG-3, IDO, KIR) and newly developed co-stimulatory antibodies (e.g. CD40, GITR, OX40, CD137, ICOS). It is important to note that co-stimulatory agents strikingly differ in their proposed mechanism of action compared with monoclonal antibodies that accomplish immune activation by blocking negative checkpoint molecules such as CTLA-4 or PD-1/PD-1 or others. Indeed, the prospect of combining agonistic with antagonistic agents is enticing and represents a real immunologic opportunity to 'step on the gas' while 'cutting the brakes', although this strategy as a novel cancer therapy has not been universally endorsed so far. Concerns include the prospect of triggering cytokine-release syndromes, autoimmune reactions and hyper immune stimulation leading to activation-induced cell death or tolerance, however, toxicity has not been a major issue in the clinical trials reported so far. Although initial phase I/II clinical trials of agonistic and novel antagonistic drugs have shown highly promising results in the absence of disabling toxicity, both in single-agent studies and in combination with chemotherapy or other immune system targeting drugs; however, numerous questions remain about dose, schedule, route of administration and formulation as well as identifying the appropriate patient populations. In our view, with such a wealth of potential mechanisms of action and with the ability to fine-tune monoclonal antibody structure and function to suit particular requirements, the second and third wave of immuno-oncology drugs are likely to provide rapid advances with new combinations of novel immunotherapy (especially co-stimulatory antibodies). Here, we will review the mechanisms of action and the clinical data of these new antibodies and discuss the major issues facing this rapidly evolving field.

Th17 and regulatory T (T

Iron is an essential nutrient for normal cell growth, and reprogramming of iron metabolism is essential to tumor cell survival and progression. HTLV-1-associated adult T-cell leukemia/lymphoma (ATLL) has no effective therapy and high levels of cell surface transferrin receptor 1 (TFR1) expression have been reported in ATLL by us and other groups. In this study, to develop a novel molecular-targeted therapy against TFR1 to modulate iron metabolism, we initially determined the expression pattern of several iron-related genes along with TFR1 and found that ATLL cells presented characteristic of an iron-deficiency state such as high expression of iron-regulatory protein 2 (IRP2) and low expression of its E3 ubiquitin-ligase, FBXL5. Therefore, we developed human IgG monoclonal antibodies to human TFR1 using a phage display method (ICOS method) to block the incorporation of the transferrin (TF)-iron complex into ATLL cells for inhibiting cell growth. One of the mAbs, JST-TFR09, presented its greater affinity to TFR1 on ATLL cells in flow cytometry (FCM) analysis than those of commercially available anti-TFR1 antibodies and identified high expression of TFR1 in most of the acute-type ATLL cells. Moreover, JST-TFR09 could interfere with binding between TFR1 and TF, which resulted in effective blockade of TFR1 internalization and induction of cell apoptosis by the treatment of ATLL cells with JST-TFR09. JST-TFR09 showed dual activities through direct cell cytotoxicity and antibody-dependent cellular cytotoxicity (ADCC), and the treatment of JST-TFR09 significantly suppressed cell growth of ATLL cells with induction of apoptosis in in vitro and in vivo experiments. Thus, JST-TFR09 described here may become a promising therapeutic antibody for the treatment of ATLL.

OBJECTIVE: Peroxisome proliferator-activated receptor γ (PPARγ) agonists have been shown to have anti-proliferative, anti-angiogenic, and proapoptotic effects, leading to interest in their use as cancer therapeutics. Pioglitazone, a U.S. Food and Drug Administration-approved type II diabetes medication and PPARγ agonist, may have a role in adjuvant head-and-neck squamous cell carcinoma treatment or prevention. Therefore, the purpose of this study was: 1) to treat oral cavity cancer cells with the PPARγ activator, pioglitazone, to analyze gene expression changes; and 2) to compare those changes with our preexisting genomic data for development of hypothesis-driven additional basic and clinical studies.
STUDY DESIGN: Prospective in vitro.
METHODS: We utilized microarray technology, as well as OCPlus (Bioconductor open source software) and Ingenuity Pathway Analysis (Qiagen, Redwood City, CA), to analyze differential gene expression in tumor and pioglitazone-treated tumor cells on a genome-wide level to demonstrate the feasibility of such an approach and determine appropriate sample size for future investigations.
RESULTS: We found that approximately 35 samples are required to adequately power future studies. We next discovered that pioglitazone significantly affects Inducible T-Cell Costimulator (iCOS)-Ligand for the T-cell-specific cell surface receptor ICOS (iCOSL) and type II diabetes mellitus pathways as a putative anti-cancer mechanism.
CONCLUSION: Genome-wide analysis is possible for the exploration of differential pathway modulation and rapid hypothesis generation. Both inflammation and type II diabetes pathways were significantly altered and therefore might provide unique hypothesis-driven pharmacodynamic parameters for future in vitro or in vivo studies utilizing thiazolidinediones. These techniques could be applied to microarray or other high throughput data from a variety of hypothesis-generating research scenarios in otolaryngology (e.g., middle ear proteomics, sinus microbiome studies).
LEVEL OF EVIDENCE: NA. Laryngoscope, 127:E124-E131, 2017.

To cope with immune responses, tumour cells implement elaborate strategies such as adaptive resistance and induction of T-cell exhaustion. T-cell exhaustion has been identified as a state of hyporesponsiveness that arises under continuous antigenic stimulus. Nevertheless, contribution of co-stimulatory molecules to T-cell exhaustion in cancer remains to be better defined. This study explores the role of myeloid leukaemia-derived co-stimulatory signals on CD4

Clinical responses to high-dose IL2 therapy are limited due to selective expansion of CD4

T-box transcription factors, T-box expressed in T cells (T-bet) encoded by Tbx21 and Eomesodermin (Eomes), drive the differentiation of effector/memory T cell lineages and NK cells. The aim of the study was to determine the prognostic influence of the expression of these transcription factors in peripheral blood (pB) in a cohort of 41 metastatic (m) RCC patients before receiving sorafenib treatment and to analyze their association with the immunophenotype in pB. In contrast to Tbx21, in the multivariate analysis including clinical features, Eomes mRNA expression was identified as an independent good prognostic factor for progression-free survival (PFS, p = 0.042) and overall survival (OS, p = 0.001) in addition to a favorable ECOG performance status (p = 0.01 and p = 0.008, respectively). Eomes expression correlated positively not only with expression of Tbx21 and TGFβ1 mRNA, but also with mRNA expression of the activation marker ICOS, and with in vivo activated HLA-DR(+) T cells. Eomes expression was negatively associated with TNFα-producing T cells. On protein level, Eomes was mainly expressed by CD56(+)CD3(-) NK cells in pB. In conclusion, we identified a higher Eomes mRNA expression as an independent good prognostic factor for OS and PFS in mRCC patients treated with sorafenib.

Adult T cell leukemia/lymphoma (ATL) is a peripheral T cell neoplasm of largely unknown genetic basis, associated with human T cell leukemia virus type-1 (HTLV-1) infection. Here we describe an integrated molecular study in which we performed whole-genome, exome, transcriptome and targeted resequencing, as well as array-based copy number and methylation analyses, in a total of 426 ATL cases. The identified alterations overlap significantly with the HTLV-1 Tax interactome and are highly enriched for T cell receptor-NF-κB signaling, T cell trafficking and other T cell-related pathways as well as immunosurveillance. Other notable features include a predominance of activating mutations (in PLCG1, PRKCB, CARD11, VAV1, IRF4, FYN, CCR4 and CCR7) and gene fusions (CTLA4-CD28 and ICOS-CD28). We also discovered frequent intragenic deletions involving IKZF2, CARD11 and TP73 and mutations in GATA3, HNRNPA2B1, GPR183, CSNK2A1, CSNK2B and CSNK1A1. Our findings not only provide unique insights into key molecules in T cell signaling but will also guide the development of new diagnostics and therapeutics in this intractable tumor.

T-cell lymphoma pathogenesis and classification have, until recently, remained enigmatic. Recently performed whole-exome sequencing and gene-expression profiling studies have significant implications for their classification and treatment. Recurrent genetic modifications in antigen ("signal 1"), costimulatory ("signal 2"), or cytokine receptors ("signal 3"), and the tyrosine kinases and other signaling proteins they activate, have emerged as important therapeutic targets in these lymphomas. Many of these genetic modifications do not function in a cell-autonomous manner, but require the provision of ligand(s) by constituents of the tumor microenvironment, further supporting the long-appreciated view that these lymphomas are dependent upon and driven by their microenvironment. Therefore, the seemingly disparate fields of genomics and immunology are converging. A unifying "3 signal model" for T-cell lymphoma pathogenesis that integrates these findings will be presented, and its therapeutic implications briefly reviewed.

Inducible T cell costimulator (ICOS) functions to regulate cell-cell signalling, immune responses and cell proliferation. ICOS single nucleotide polymorphism (SNP) may affect protein expression and functions. This study investigated the association of ICOS SNPs with hepatitis B virus (HBV) infection and outcome in a Chinese population. A total of 1290 Chinese Han individuals were enrolled, including 63 asymptomatic HBV carriers, 220 chronic hepatitis B patients (CHB), 249 HBV-related liver cirrhosis patients (LC), 108 patients with HBV-related hepatocellular carcinoma (HCC), 338 patients with natural HBV clearance and 312 healthy subjects (as controls). DNA samples from these subjects were genotyped for four ICOS SNPs (rs11883722, rs10932029, rs1559931 and rs4675379) using TaqMan SNP Genotyping Assay and analysed. The data showed that genotype and allele frequencies of ICOS SNPs in cases and controls followed the Hardy-Weinberg distribution. The CC genotype of rs4675379 was higher in patients with HBV infection (including AC, CHB, LC and HCC) than in patients with HBV clearance (P = 0.006). Furthermore, the genotype 'GA' and the minor allele 'A' of rs1559931 were associated with a decreased HCC susceptibility (P < 0.001). Haplotype analysis data showed that 'GC' haplotype in block 2 (rs1559931 and rs4675379) had a lower frequency in patients than in HBV-cleared subjects (P = 0.034), although its overall frequency was only 1.6%. Our study found that ICOS rs1559931 SNP was associated with decreased HBV-related HCC risk in the studied Chinese Han population, except for patients with natural clearance of HBV.

To explore the interactions among immunity related genes and the risk of breast cancer (BC), 376 invasive ductal carcinoma (IDC) of the breast cases and 366 healthy controls were selected into our study. Twenty single nucleotide polymorphisms (SNPs) of five immunological genes in the CD28/B7 pathway were genotyped. Overall, five SNPs filtered by the Relief F algorithm were rs733618, rs11889031, rs4553808, rs4675374 and rs10754339. The best model of multifactor dimensionality reduction (MDR) contained rs733618 and rs11889031. The high risk genotype combination contributed to increasing risk of breast cancer (odds ratio (OR), 4.36; 95% confidence interval (CI); 3.15-6.02). The information gain (IG) value of these two SNPs was 8.07%, presented the strongest interaction effect. Five significant multiplicative interactions and seven significant combining effects were found among the filtered SNPs. Moreover, the filtered SNPs were still stable in the groups of ER(+), PR(+), CerbB2(-) and lymph node (LN) involvement positive with the best models including rs733618 and rs11889031. The most frequent haplotype was TACC which significantly increased breast cancer risk (OR, 1.80; 95% CI, 1.43-2.25). These results suggested that interactions among cytotoxic T lymphocyte antigen-4 (CTLA4), inducible co-stimulator (ICOS) and B7H4 might play critical roles on the risk of breast cancer.

This study compared second-generation chimeric antigen receptors (CAR) encoding signaling domains composed of CD28, ICOS, and 4-1BB (TNFRSF9). Here, we report that certain CARs endow T cells with the ability to undergo long-term autonomous proliferation. Transduction of primary human T cells with lentiviral vectors encoding some of the CARs resulted in sustained proliferation for up to 3 months following a single stimulation through the T-cell receptor (TCR). Sustained numeric expansion was independent of cognate antigen and did not require the addition of exogenous cytokines or feeder cells after a single stimulation of the TCR and CD28. Results from gene array and functional assays linked sustained cytokine secretion and expression of T-bet (TBX21), EOMES, and GATA-3 to the effect. Sustained expression of the endogenous IL2 locus has not been reported in primary T cells. Sustained proliferation was dependent on CAR structure and high expression, the latter of which was necessary but not sufficient. The mechanism involves constitutive signaling through NF-κB, AKT, ERK, and NFAT. The propagated CAR T cells retained a diverse TCR repertoire, and cellular transformation was not observed. The CARs with a constitutive growth phenotype displayed inferior antitumor effects and engraftment in vivo. Therefore, the design of CARs that have a nonconstitutive growth phenotype may be a strategy to improve efficacy and engraftment of CAR T cells. The identification of CARs that confer constitutive or nonconstitutive growth patterns may explain observations that CAR T cells have differential survival patterns in clinical trials.

Polymorphisms within the 3'-untranslated region (3'-UTR) of genes have been proved to contribute to the risk of cancers. Here, we determined 16 putatively functional polymorphisms in the 3'-UTR of 11 B7/CD28 genes in 382 colorectal cancer patients and 714 healthy controls. Statistical analysis revealed that ICOS rs4404254-C-allele carriers (p=0.0014), rs1559931-A-allele carriers (p=0.0027), and rs4675379-C-allele carriers (p=0.026) were significantly fewer in patients than those in controls. B7-H4-rs13505-GG homozygotes were more prevalent in patients (p=0.03). CD80-rs7628626-GT was apparently less in the patients with lymph node metastasis (p=0.004) or in advanced stage (p=0.037). Furthermore, we found that these polymorphisms impacted the regulatory role of miR-21-3p, miR-186-5p, miR-323b-5p, miR-1207-5p, miR-1279, miR-2117, and miR-3692-3p in the expression of the B7/CD28 molecules. Our findings suggest that rs7628626, rs13505, rs4404254, rs1559931, and rs4675379, through disrupting the regulatory role of miRNAs in the expression of B7/CD28 molecules, contribute to the occurrence and progress of colorectal cancer.

Here we describe increased expression of IL6R in the tumoural region of lung tissue from patients affected by lung adenocarcinoma as compared to squamous cell lung carcinoma. Moreover, here we found increased IL6R in the tumour free part of the lung. By using a murine model of lung adenocarcinoma, we discovered that few lung tumour cells expressed IL-6R and CD4+CD25+Foxp-3+ T regulatory cells down-regulated IL-6R in the tumour bearing lungs. Downstream of IL-6R, the Th17 lineage-specification factors: Signal transducer and activator of transcription 3 (STAT3), Basic leucine zipper transcription factor, BATF and a protein encoded by the RORC in human (RAR-related orphan receptor C) (RORγT), were also found induced in the tumoural region of lung tissue from patients affected by lung adenocarcinoma as compared to those carrying squamous cell carcinoma. Moreover, pSTAT3 protein was found phosphorylated and auto-phosphorylated in the tumoural region of patients with adeno cell carcinoma of the lung as compared to the tumoural region of patients with squamous cell carcinoma of the lung. Intranasal application of anti-IL-6R antibodies in a murine model of lung adenocarcinoma, induced T regulatory cell markers such as Foxp3, Ctla4, Icos, Il10, Il21, Folr4 and Lag3 and inhibited Rorc in lung adenocarcinoma.