Defective Immune Surveillance Research Articles

Elevated levels of cell-free NKG2D-ligands modulate NKG2D surface expression and compromise NK cell function in severe COVID-19 disease.

It is now clear that coronavirus disease 19 (COVID-19) severity is associated with a dysregulated immune response, but the relative contributions of different immune cells is still not fully understood. SARS CoV-2 infection triggers marked changes in NK cell populations, but there are contradictory reports as to whether these effector lymphocytes play a protective or pathogenic role in immunity to SARS-CoV-2. To address this question we have analysed differences in the phenotype and function of NK cells in SARS-CoV-2 infected individuals who developed either very mild, or life-threatening COVID-19 disease. Although NK cells from patients with severe disease appeared more activated and the frequency of adaptive NK cells was increased, they were less potent mediators of ADCC than NK cells from patients with mild disease. Further analysis of peripheral blood NK cells in these patients revealed that a population of NK cells that had lost expression of the activating receptor NKG2D were a feature of patients with severe disease and this correlated with elevated levels of cell free NKG2D ligands, especially ULBP2 and ULBP3 in the plasma of critically ill patients. In vitro, culture in NKG2DL containing patient sera reduced the ADCC function of healthy donor NK cells and this could be blocked by NKG2DL-specific antibodies. These observations of reduced NK function in severe disease are consistent with the hypothesis that defects in immune surveillance by NK cells permit higher levels of viral replication, rather than that aberrant NK cell function contributes to immune system dysregulation and immunopathogenicity.

The Limitations of Current T Cell-Driven Anticancer Immunotherapies Can Be Overcome with an Original Extracellular-Vesicle-Based Vaccine Strategy.

The emergence of tumors associated with defects in immune surveillance often involve the impairment of key functions of T lymphocytes. Therefore, several anticancer immunotherapies have focused on the induction/strengthening of the tumor-specific activity of T cells. In particular, strategies based on immune checkpoint inhibitors, CAR-T cells, and mRNA vaccines share a common goal of inducing/recovering an effective antitumor cytotoxic activity, often resulting in either exhausted or absent in patients' lymphocytes. In many instances, these approaches have been met with success, becoming part of current clinic protocols. However, the most practiced strategies sometimes also pay significant tolls in terms of adverse events, a lack of target specificity, tumor escape, and unsustainable costs. Hence, new antitumor immunotherapies facing at least some of these issues need to be explored. In this perspective article, the characteristics of a novel CD8+ T cell-specific anticancer vaccine strategy based on in vivo-engineered extracellular vesicles are described. How this approach can be exploited to overcome at least some of the limitations of current antitumor immunotherapies is also discussed.

Safety and efficacy of immune checkpoint inhibitors after allogeneic hematopoietic cell transplantation.

The immune system plays a major role in preventing infections and cancers. Impairment in immunity may facilitate the development of neoplasia owing to defective immune surveillance, among other mechanisms. Immune evasion plays a significant role in relapse after allogeneic hematopoietic cell transplantation (alloHCT); one purported mechanism is through immune checkpoint signaling pathways. Checkpoint inhibitors (CPIs) are FDA approved for relapsed classical Hodgkin's Lymphoma (cHL), primary mediastinal large B cell Lymphoma (PMBCL) and other solid tumors. Retrospective studies evaluating the outcomes of alloHCT after prior exposure to CPIs showed favorable survival outcomes but high rates of graft-versus-host disease (GVHD); the risk appears to be lower when using post-transplant cyclophosphamide as GVHD prophylaxis. CPIs have increasingly been used to prevent or treat post-alloHCT relapse. Available data, albeit limited, supports the clinical activity of CPIs in post-alloHCT relapse; however, serious and even fatal cases of GVHD have been reported. The optimal timing, schedule, dosing, and patients likely to benefit from this strategy are yet to be identified. In this review, we highlight the immune system's role in cancer surveillance and relapse prevention and discuss the current clinical evidence of CPIs use in post-alloHCT relapse.

Role of cellular senescence in the pathogenesis of systemic sclerosis.

Systemic sclerosis (SSc) is a chronic rheumatic disease that is characterized by immune activation, vasculopathy and fibrosis of the skin and internal organs. It has been proposed that premature onset of ageing pathways and associated senescent changes in cells contribute to the clinical and pathological features of SSc. The aim of this review is to critically review recent insights into the involvement of cellular senescence in SSc. Cellular senescence plays a critical role in SSc pathogenesis, particularly involving endothelial cells and fibroblasts. Immunosenescence could also contribute to SSc pathogenesis by direct alteration of cellular functions or indirect promotion of defective immune surveillance. Molecular studies have shed some light on how cellular senescence contributes to fibrosis. Recent and planned proof-of-concept trials using senotherapeutics showed promising results in fibrotic diseases, including SSc. There is increasing evidence implicating cellular senescence in SSc. The mechanisms underlying premature cellular senescence in SSc, and its potential role in pathogenesis, merit further investigation. Emerging drugs targeting senescence-related pathways might be potential therapeutic options for SSc.

128: DECIPHERING THE IMMUNOLOGICAL DETERMINANTS OF RESPONSE TO NEOADJUVANT CHEMO-RADIATION IN ESOPHAGEAL ADENOCARCINOMA

Abstract Background and aim The standard therapy for esophageal adenocarcinoma (EAC) is pre-operative neoadjuvant chemo-radiation (NACR) and surgery. Less that 30% of patients achieve a pathological complete response (pCR) and increased 5-year survival after NACR. Understanding the mechanisms of response to NACR is pivotal to better stratify patients and inform the design of more efficacious therapies. Cancer progression partly depends on defective immune surveillance, and the presence of tumor infiltrating T lymphocytes (TILs) is associated with better prognosis of several malignancies. We hypothesize that NACR induces immunogenic cell death that stimulates immune responses against tumor neoantigens (TNAs), derived from cancer somatic mutations, in the responders. We thus compared responders and non-responders for tumor immune contexture, immune molecular pathways, circulating immune markers and TNAs-specific T cells responses. Methods Pre-NACR tumor biopsies prospectively collected from consecutive EAC patients were subjected to DNA whole exome and RNA sequencing, high dimensional (HD) multiparametric flow cytometry and immune contexture analysis by histology and by proteome digital spatial profiling (n = 53 targets), while their peripheral blood mononuclear cells and plasma collected pre- and post- NACR were analyzed by HD flow cytometry and cytokine multiplexing assay respectively. Results Treatment-naïve EACs of NACR responders showed higher TNA load and enriched immune signatures compared to non-responders. Moreover, pre-treatment EACs of responders demonstrate augmented effector T cell infiltration and reduced Treg cells and M2-like pro-tumoral macrophages. High M2-like macrophages score associated with lower overall survival. Increased frequencies of circulating myeloid suppressor populations were detected both at baseline and after NACR in non-responder patients. These observations were associated with circulating levels of n = 30 cytokines/chemokines and with spatial expression of immune markers in tumor biopsies. Conclusion Collectively, these results support pre-existing immune-mediated mechanisms of anti-tumor response stimulated by NACR in the fraction of responder EAC patients, suggesting possible ways to stratify patients and direct them to more tailored neoadjuvant and/or adjuvant treatments, possibly in combination with personalized immunotherapy approaches.

The Inflammatory Feed-Forward Loop Triggered by the Complement Component C3 as a Potential Target in Endometriosis.

The complement system is a major component of humoral innate immunity, acting as a first line of defense against microbes via opsonization and lysis of pathogens. However, novel roles of the complement system in inflammatory and immunological processes, including in cancer, are emerging. Endometriosis (EM), a benign disease characterized by ectopic endometrial implants, shows certain unique features of cancer, such as the capacity to invade surrounding tissues, and in severe cases, metastatic properties. A defective immune surveillance against autologous tissue deposited in the peritoneal cavity allows immune escape for endometriotic lesions. There is evidence that the glandular epithelial cells found in endometriotic implants produce and secrete the complement component C3. Here, we show, using immunofluorescence and RT-qPCR, the presence of locally synthesized C3 in the ectopic endometriotic tissue, but not in the eutopic tissue. We generated a murine model of EM via injection of minced uterine tissue from a donor mouse into the peritoneum of recipient mice. The wild type mice showed greater amount of cyst formation in the peritoneum compared to C3 knock-out mice. Peritoneal washings from the wild type mice with EM showed more degranulated mast cells compared to C3 knock-out mice, consistent with higher C3a levels in the peritoneal fluid of EM patients. We provide evidence that C3a participates in an auto-amplifying loop leading to mast cell infiltration and activation, which is pathogenic in EM. Thus, C3 can be considered a marker of EM and its local synthesis can promote the engraftment of the endometriotic cysts.

Evasion of Innate Immunity Contributes to Small Cell Lung Cancer Progression and Metastasis.

Small cell lung cancer (SCLC) is a pulmonary neuroendocrine cancer with very poor prognosis and limited effective therapeutic options. Most patients are diagnosed at advanced stages, and the exact reason for the aggressive and metastatic phenotype of SCLC is completely unknown. Despite a high tumor mutational burden, responses to immune checkpoint blockade are minimal in patients with SCLC. This may reflect defects in immune surveillance. Here we illustrate that evading natural killer (NK) surveillance contributes to SCLC aggressiveness and metastasis, primarily through loss of NK-cell recognition of these tumors by reduction of NK-activating ligands (NKG2DL). SCLC primary tumors expressed very low level of NKG2DL mRNA and SCLC lines express little to no surface NKG2DL at the protein level. Chromatin immunoprecipitation sequencing showed NKG2DL loci in SCLC are inaccessible compared with NSCLC, with few H3K27Ac signals. Restoring NKG2DL in preclinical models suppressed tumor growth and metastasis in an NK cell-dependent manner. Likewise, histone deacetylase inhibitor treatment induced NKG2DL expression and led to tumor suppression by inducing infiltration and activation of NK and T cells. Among all the common tumor types, SCLC and neuroblastoma were the lowest NKG2DL-expressing tumors, highlighting a lineage dependency of this phenotype. In conclusion, these data show that epigenetic silencing of NKG2DL results in a lack of stimulatory signals to engage and activate NK cells, highlighting the underlying immune avoidance of SCLC and neuroblastoma. SIGNIFICANCE: This study discovers in SCLC and neuroblastoma impairment of an inherent mechanism of recognition of tumor cells by innate immunity and proposes that this mechanism can be reactivated to promote immune surveillance.

Abstract 3112: Prospective immune phenotyping of neuroblastoma children at diagnosis reveals specific immune defects related to the aggressiveness of the disease

Abstract CONTEXT: Neuroblastoma is the most common extracranial solid tumor in children. Patients with low- and intermediate-risk neuroblastoma (LIRNB) have favorable prognosis and an excellent five-year survival rate of more than 90%. However, in the case of high-risk neuroblastoma (HRNB; ~50% of cases), the prognosis of treatment remains unfavorable and the five-year survival rate remains under 40% despite aggressive multi modal therapy. QUESTION: We wanted to know if there was a defect in immune surveillance that could explain part of the aggressive phenotype of HRNB. METHODS: We performed a prospective multi-center study on thirty two new cases of neuroblastoma at all stages. We performed a wide phenotyping of immune cells by flow cytometry in fresh whole blood, and fresh whole bone marrow at diagnosis of children diagnosed for neuroblastoma. Cell counts were performed in order to be able to reason in absolute values (G/L) rather than only percentages. RESULTS: Blood and bone marrow from 20 LIRNB and 12 HRNB were prospectively analyzed. We found that specific subsets of circulating immune cells such as myeloid dendritic cells BDCA-1+, plasmacytoid dendritic cells, Teff/Treg CD4+ T-cells, were decreased in HRNB blood compared to LIRNB groups as opposed to other subsets (e.g iNKT, g9d2 T-cells,⋯). More surprisingly, these differences were not due to defects in cell production as there was no difference found for the same cell populations in the bone marrow between LIRNB and HRNB children. CONCLUSION: Specific immune cell defects are found in the blood from High risk neuroblastoma children that are not present in the blood of low & intermediate risk neuroblastoma children. These defects are not due to bone marrow production impairment. These results suggest that high risk neuroblastoma disease generates an impairment in the immune homeostasis of children. Gene expression analysis of cytokine/chemokine from these neuroblastoma tumors is underway (NCT01295762). Citation Format: Sandrine Susini, Isabelle Rochet, Estelle Verronèse, Christine Bardin, Chantal Rigal, Cécile Conter, Perrine Marec-Bérard, Christophe Bergeron, Audrey Lardy-Cleaud, Séverine Neymarc, Séverine Metzger, Jean-Louis Stephan, Dominique Plantaz, Christophe Caux, Christine Ménétrier-Caux, Aurelien Marabelle. Prospective immune phenotyping of neuroblastoma children at diagnosis reveals specific immune defects related to the aggressiveness of the disease [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3112.

Carcinogenesis with mitochondrial respiratory degeneration was testified using a specific herbal medicine

Background: Oncologists continue to dispute whether the main factors underlying carcinogenesis are related to genetic abnormalities or rather protracted mitochondrial respiratory dysfunction, both are core reasons for the occurrence of carcinogenesis. Thus, the aim of this article is to demonstrate that mitochondrial respiratory degeneration might be the true causal factor underlying carcinogenesis by presenting evidence obtained from experiments with a specific herbal medicine to inhibit oxygen respiration in cancer cells. Methods: The tested herbal medicine (Sun Advance) was used to selectively impact mitochondrial respiration of cancer cells, which is a hallmark of cancer, the results would thereby obtain evidence that carcinogenesis originates from mitochondrial respiratory degeneration. Results: In cellular and animal experiments, the Sun Advance had good effects against most hallmarks of cancer. Conclusion: Carcinogenesis is a form of disease initiation due to mitochondrial respiratory degeneration testified using a specific herbal medicine. This demonstrate that mitochondrial respiratory degeneration might be re-differentiating factor with the use of herbal medicine. Therefore, primary cancer prevention programs should advocate for life-style changes and dietary energy restriction. Keywords: Genetic abnormalities, Protracted mitochondrial respiratory degeneration, Genetic instability, Herbal medicine, Cancer cell respiration, Defective immune-surveillance.

Interferon-alfa2 Treatment of Patients with Polycythemia Vera and Related Neoplasms Influences Deregulated Inflammation and Immune Genes in Polycythemia Vera and Allied Neoplasms

Introduction: The Philadelphia-negative myeloproliferative neoplasms are associated with deregulation of inflammation and immune genes implying a gene signature of a chronic inflammatory state and immune deregulation. Several studies have demonstrated interferon-alpha2 (IFN) to be highly efficacious in normalizing elevated blood cell counts and in inducing molecular remissions as well. Using whole blood gene expression, we herein for the first time show that IFN profoundly influences the deregulated inflammation- and immune genes giving rise to a gene signature indicative of decreased inflammation and improved regulation of immune genes during IFN treatment.Material and Methods: Gene expression microarray studies have been performed on eight patients with ET, 21 patients with PV, and 4 patients with PMF. All patients received treatment with IFN, in the large majority in a dosage ranging from 45-90 ug x 1 sc/week. Gene expression profiles were generated using Affymetrix HG-U133 2.0 Plus microarrays recognizing 54.675 probe sets (38.500 genes). Total RNA was purified from whole-blood and amplified to biotin-labeled aRNA and hybridized to microarray chips.Results: We identified 6261, 10,008, and 2828 probe sets to be significantly differentially expressed in ET, PV, and PMF, respectively, during treatment with IFN (pvalue <0.05). Previously, we have found 123 inflammation and immune genes to be significantly deregulated in patients with MPN compared to healthy subjects. Here, we extend the gene list with 19 inflammation and immune genes, recently shown to be involved in other cancer. Several of these 142 genes were significantly deregulated during IFN therapy. In response to treatment with IFN, 20 genes were upregulated in ET including CX3CR1, CCL2, CCR1, CCR5, CCR7, HGF, HLA-G, IL1RN, IL10RA, PTX3, TP53, and TGFB1, and 6 genes were downregulated including MAPK1, ORM1, and IL1R1. In patients with PV, ABCF1, CCL2, CCR1, CCR2, CCR5, CCR7, CD3E, CD3G, CD4, CD40LG, CX3CR1, HLA-G, IL1RN, IL8, IL10RA, PTX3, TP53, TGFB1, TNFAIP3 were among the 32 upregulated genes, and BCL6, IL1R1, MAPK1, and ORM1 were among the 10 downregulated genes. During treatment with IFN, 11 genes were upregulated in PMF including CCL2, CCR1, CCR5, CX3CR1, IL1RN, PTX3, and TNFAIP3 and 5 genes were downregulated including BCL6, IL1R1, and ORM1.Discussion and Conclusion: Interferon-alpha2 is increasingly been recognized as a highly efficacious and promising agent in the treatment of MPNs. During recent years several studies have shown that chronic inflammation and immunoderegulation is likely involved in the pathogenesis of MPNs. Thus, elevated blood levels of circulating inflammatory cytokines have been recorded, some even having a prognostic impact and predicting imminent leukemic transformation. Chronic inflammation impairs IFN-signaling and likely therefore the efficacy of IFN in MPNs, although this notion has to be confirmed in MPNs. Immunoderegulation with defective immune surveillance may contribute to the increased risk of second cancers both before and after the MPN-diagnosis. Accordingly, there is an urgent need to explore if IFN might be able to restore deregulated inflammation and immune genes in MPNs. In this study, we have convincingly shown that several genes of significance for inflammation and immune surveillance are positively influenced by IFN treatment implying a significant downregulation of upregulated inflammation genes (e.g. BCL6, IL1R1, MAPK1, ORM1,) and a significant upregulation of downregulated immune genes (e.g. ABCF1, CCR2, CCR5, CCR7, CD3D, CD3E, CD3G, CD4, CD40LG, CXCR1, HLA-G, IL10RA, IL8, TFGB1, TNFAIP3, and TP53). Recently, we have shown significant downregulation of CCR9, CREB1, FAS, LSP1, LTB, NFKB1, SCYE1, SELPG, STAT3, and TNFAIP8L1, and significant upregulation of CCL25, CCL7, CSF3, CXCL9, FOXP3, IL17A, IL17C, IL1F10, IL1F6, IL22, IL4, IL5, ITGB3, in ET, PV, and PMF compared to controls. These genes were no longer significantly deregulated during IFN-treatment. In conclusion, our results have added highly important information on the impact of IFN upon deregulated inflammation and immune genes in MPNs, thereby substantiating the beneficial effects of IFN and its major role as the cornerstone in the future treatment of MPNs. Our study opens the avenue for larger studies exploring the genomic landscape during treatment of patients with MPNs. DisclosuresHasselbalch:Novartis: Research Funding.

Histoprognostic Factors in Bladder Cancer: A Case Series of 156 Patients

Background: For a long time, phenomenon of defective immune-surveillance in cancer patients from pathological standpoint was not elucidated. I have reported 1985 that lowering seral cyclic AMP is biochemical reason of defective immunesurveillance from adoptive lymphocyte immunotherapy experiment among cancer patients. Recently, Honjo Tasku et al., reported why T cell don’t attack cancer cells by the covering of PD-L1 on its receptor of PD-1. This is also one of defective immune-surveillance from cellular side. I have recently surveyed the correlation between seral cyclic AMP concentration and immune activity in cancer patients. I have confirmed the biochemical reason of lowering cyclic AMP leads to seral defective immune-surveillance in cancer patient’s serum. Methods: I have surveyed seral cyclic AMP concentration and T cell number and NK cell among 40 cancer patients and preclinical cancer patients. Results: The averaged seral cyclic AMP has a positive tendency between lower immune activity and lower cyclic AMP in the serum when cancer is developing beforehand. Conclusion: As cyclic AMP is elevated after fasting therapy, adoptive lymphocyte immunotherapy become useful as immuno-potentiation even if in advanced cancer patients.

Driver Mutations Affecting Natural Killer/T Cell Lymphoma

Natural killer/T cell lymphoma (NKTCL) is an aggressive subtype of non-Hodgkin lymphoma that is rare overall but has a higher prevalence in Chinese and Hispanic populations. Recent sequencing efforts have improved the understanding of the disease and revealed a number of genes that may drive the pathogenesis of NKTCL. These efforts were largely limited by the number of cases studied. Based on previously reported whole exome sequencing data in NKTCL and other lymphoid malignancies and on the frequency and potential biological significance of the mutant, we designed a 334-gene panel and sequenced 105 NKTCLs. We characterized single nucleotide variants (SNVs), small insertions and deletions (indels), and copy number alterations (CNAs) that may have pathogenic roles.Tumor DNA was isolated from 74 Chinese and 31 Hispanic NKTCL patients. For 7 of these, matched normal DNA was extracted from peripheral blood. The 334 genes in the custom panel were captured using the Agilent SureSelect platform, and the exons of these genes were sequenced on an Illumina HiSeq 2500 instrument. For cases without matched normal samples, we discriminated between somatic and germline variants with an in-house machine learning algorithm which is based on information of public variant databases, variant annotations, and sequencing statistics, and followed by manual inspection. CNAs were determined based on the untargeted sequences using an R package, CopywriteR.We identified a total of 479 SNVs and 113 indels in 155 genes in 102 out of 105 patients. Mutations in STAT3 and JAK3 were identified in 30% and 7% of the cases, respectively. The hotspot mutations Y640F, S614R, and G618R in the SH2 domain of STAT3 and A573V of the JH2 domain of JAK3 were observed in most of these cases. 46% of the cases harbored one or more mutations in epigenetic regulators ARID1A, EP300, MLL2, MLL3, and TET2. Transcription co-repressors BCOR and NCOR2 were mutated in 25 of the cases. 13% of the cases contained mutations in HLA-A or CIITA, most of which were inactivating, suggesting possible defects in immune surveillance. DDX3X and TP53 mutations were detected in 19% and 10% of the cases, respectively. TP53 mutations were mutually exclusive with mutations in DDX3X and, except for one case, with STAT3 also. MGA mutations, which were present in 12% of the cases, were usually concurrent with STAT3 mutations. The mutational profiles of Chinese and Hispanic cases were similar, and only CD58 mutations were significantly more common in Hispanic patients (p=0.03). Comparison between NKTCL with localized and systemic presentation demonstrated that CCR7 was mutated more frequently in systemic cases (p=0.02). Prognostic analysis identified EP300 mutations as a potential marker for poorer survival (p=0.01). Frequent copy loss on chromosome 6q was observed and PRDM1, which is present on 6q, was mutated in 10% of the cases.In summary, we identified the profile of mutated genes in a large series of NKTCL. Most of the genes have been previously reported to be mutated in NKTCL, but the frequencies for some of the mutants were quite different. Interestingly, the mutational profiles of NKTCL in Chinese and Hispanic patients were very similar despite the geographic differences. Mutations leading to abnormal activation of the JAK-STAT3 pathway and abnormal chromatin modification were highly prevalent. BCOR, DDX3X and TP53 mutations were also frequent. Variants in CCR7 and EP300 were potential markers for systemic disease and poor prognosis, respectively. [Display omitted] [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

'Trained immunity': consequences for lymphoid malignancies.

In hematological malignancies complex interactions exist between the immune system, microorganisms and malignant cells. On one hand, microorganisms can induce cancer, as illustrated by specific infection-induced lymphoproliferative diseases such as Helicobacter pylori-associated gastric mucosa-associated lymphoid tissue lymphoma. On the other hand, malignant cells create an immunosuppressive environment for their own benefit, but this also results in an increased risk of infections. Disrupted innate immunity contributes to the neoplastic transformation of blood cells by several mechanisms, including the uncontrolled clearance of microbial and autoantigens resulting in chronic immune stimulation and proliferation, chronic inflammation, and defective immune surveillance and anti-cancer immunity. Restoring dysfunction or enhancing responsiveness of the innate immune system might therefore represent a new angle for the prevention and treatment of hematological malignancies, in particular lymphoid malignancies and associated infections. Recently, it has been shown that cells of the innate immune system, such as monocytes/macrophages and natural killer cells, harbor features of immunological memory and display enhanced functionality long-term after stimulation with certain microorganisms and vaccines. These functional changes rely on epigenetic reprogramming and have been termed 'trained immunity'. In this review the concept of 'trained immunity' is discussed in the setting of lymphoid malignancies. Amelioration of infectious complications and hematological disease progression can be envisioned to result from the induction of trained immunity, but future studies are required to prove this exciting new hypothesis.

Papillary Thyroid Cancer in a Child with Progressive Transformation of Germinal Centers.

Objectives. To describe the presentation and management of a child with Progressive Transformation of Germinal Centers (PTGC), an uncommon condition characterized by significant persistent lymphadenopathy, who developed papillary thyroid carcinoma and to explore and review potential links between PTGC and neoplastic processes in the head and neck. Methods. Case presentation and literature review are used. Results. A 10-year-old female presented with a right parotid mass and cervical lymphadenopathy. Multiple biopsies revealed PTGC without malignancy. Two years later, she developed fatigue and weight gain, and a thyroid nodule was found. Fine needle aspiration was strongly suggestive of papillary thyroid carcinoma. The patient underwent total thyroidectomy and central neck dissection without surgical management of the longstanding right lateral neck lymphadenopathy. Final pathology confirmed papillary thyroid carcinoma. She was treated with radioactive iodine therapy postoperatively and remains free of disease at three years of follow-up. Conclusions. PTGC is considered a benign condition but has previously been associated with Nodular Lymphocyte Predominant Hodgkin Lymphoma (NLPHL). This is the first reported case of papillary thyroid cancer in a child with preexisting cervical PTGC and no defined risk factors for thyroid malignancy. No link has been established with thyroid carcinoma, but patients with PTGC may have a defect in immune surveillance that predisposes them to malignancy.

Inducible expression of antigen processing and antigen presentation molecules and cancer testis antigens in human prostate cancer

Defects in immune surveillance have been correlated with tumorigenesis and poor clinical outcomes in cancer patients. Given recent advances in vaccine therapies and checkpoint inhibition, therapeutic targeting of defective antigen processing may play a critical role in improving the broad utility of immunotherapies. Epigenetic alterations have been found to alter expression of genes involved in antigen processing and presentation. We evaluated the ability of epigenetic modifying agents to increase expression of antigen presentation machinery (APM) molecules ex vivo drug culture using primary human prostate tumor tissue biopsies. Prostate tissue biopsies were collected from patients with advanced, localized prostate cancer undergoing radical prostatectomy. These tumor biopsies sectioned into 200uM slices and cultured with the hypomethylating agent decitabine (5AZA) and/or histidine-deacetylase inhibitor panobinostate (LBH589). After 72 hours of treatment, tissue slices were harvested and subjected to qRT-PCR analysis. 5AZA alone or in combination with LBH589 have increased expression of MHC Class I molecules (HLA-ABC, B2M) and APM elements including TAP2, Tapasin and LMP7. Increased expression of MHC class I molecules was identified in human prostate cancer treated with 5AZA and the combination of 5AZA and LBH589. Significant inter-patient heterogeneity was also observed, suggesting other mechanisms by which prostate cancer downregulates MHC class I expression. We further identify increased expression of Cancer-Testis Antigens in these samples, indicating multiple mechanisms by which epigenetic modifying agents may improve immune recognition of human prostate cancer. These include NY-ESO, SSX2 and NYSAR35. These results suggest novel therapeutic strategies that can be employed with other immune based therapies for men with localized and advanced prostate cancer.

Abstract 1295: Role of glycine N-methyltransferase in the regulation of T cell responses in experimental autoimmune encephalomyelitis

Abstract Glycine N-methyltransferase (GNMT) is known for its function as a tumor suppressor gene. Since 100% of female Gnmt−/− mice developed hepatocellular carcinoma, we hypothesized that Gnmt−/− mice may have defective immune surveillance. In this study, we examined the immune modulation of GNMT in T cell responses using experimental autoimmune encephalomyelitis (EAE). The results showed that EAE severity was significantly reduced in Gnmt−/− mice. Pathological examination of the spinal cords revealed that Gnmt−/- mice had significantly lower level of mononuclear cell infiltration and demyelination than the wild-type mice. In addition, quantitative real-time PCR showed that expression levels of pro-inflammatory cytokines, including IFN-gamma and IL-17A, were much lower in the spinal cord of Gnmt−/- than in that of wild-type mice. Accordingly, myelin oligodendrocyte glycoprotein (MOG)-specific T cell proliferation and induction of T helper (Th) 1 and Th17 cells were markedly suppressed in MOG35-55 induced Gnmt−/- mice. Moreover, the amount of regulatory T cells was significantly increased in these mice. When the T cell receptor was stimulated, the proliferative capacity and the activation status of mTOR-associated downstream signaling were significantly decreased in GNMT−/- CD4+ T cells via an IL-2 and CD25-independent manner. Moreover, GNMT deficiency enhanced the differentiation of regulatory T cells without affecting the differentiation of Th1 and Th17 cells. Furthermore, the severity of EAE in mice adoptive transferred with GNMT-deficient CD4+ T cells was much milder than in those with wild-type CD4+ T cells. In summary, our findings suggest that GNMT is involved in the pathogenesis of EAE and plays a crucial role in the regulation of CD4+ T cell functions. Citation Format: Chung-Hsien Li, Ming-Hong Lin, Yen-Fu Chen, Shih-Han Chu, Pang-Hsien Tu, Cheng-Chieh Fang, Chia-Hung Yen, Peir-In Liang, Jason C. Huang, Yu-Chia Su, Huey-Kang Sytwu, Yi-Ming A. Chen. Role of glycine N-methyltransferase in the regulation of T cell responses in experimental autoimmune encephalomyelitis. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 1295. doi:10.1158/1538-7445.AM2015-1295

Incidence and Survival Outcomes of Secondary Malignancies Among Survivors of Waldenström Macroglobulinemia

Background: Waldenstrom macroglobulinemia (WM) is an indolent malignancy characterized by extended survival, and affects predominantly older individuals, who are at risk for secondary malignancies (SM). The objectives of our study were to characterize incidence and outcomes of SM after WM diagnosis using the Surveillance, Epidemiology and End Results (SEER) database.Methods: Using the SEER-13 data from 1992-2011, we calculated standardized incidence ratios (SIR) with 95% confidence intervals (CI) for rates of solid and hematologic SM in WM patients compared with matched general population, excluding synchronous tumors. We compared SIR in groups defined by attained age, calendar year, sex and race. SIRs from SM with less than 10 cases are not reported. Also, overall survival (OS) after a malignancy diagnosis was compared between patients with or without antecedent WM in Cox proportional-hazard regression models adjusting for attained age, sex, race and tumor stage using SEER-18 data from 2000-2011. The outcome of interest for survival was hazard ratio (HR) with 95% CI.Results: Among the 4,676 WM patients in the SEER-13 database, 681 SM were recorded (Table). The overall SIR was 1.49 (95% CI 1.38-1.61) and the median time to SM was 3.7 years (95% CI 3.2-4.2 years). The cumulative incidence of SM was 9.5% (95% CI 8.6-10.5%) at 5 years, and 16.1% (95% CI 14.8-17.3%) at 10 years. The cumulative incidence at 10 years was 12.2% (95% CI 11.1-13.3%) for solid tumors and 4.2% (95% CI 3.5-4.9%) for hematologic SM. The excess risk of solid tumors peaked between 5-10 years from WM diagnosis (SIR 1.1, 1.4 and 1.1 for latency of 0-5, 5-10 and more than 10 years, respectively), while for hematologic malignancies it grew continuously with time (SIR 3.8, 4.4 and 6.7, respectively). The risk of solid tumors was significantly increased for cancers of the lung, urinary tract, thyroid gland and melanoma, but not for breast, prostate or colorectal cancer. Among lymphomas, diffuse large B-cell (DLBCL) was the most frequent subtype. Patients younger than 65 had a significantly higher SIR for any SM (SIR 2.2, 95% CI 1.9-2.6) than those 65 years or older (SIR 1.4, 95% CI 1.3-1.5). The SIR for SM was similar in the 1990s (SIR 1.3, 95% CI 1.1-1.6) and 2000s (SIR 1.5, 95% CI 1.4-1.7). The SIR for solid tumors was the same (SIR 1.2) among men and women, but for hematologic malignancies the excess risk was significantly higher in women (SIR 5.8, 95% CI 4.6-7.2) than men (SIR 3.4, 95% CI 2.8-4.2). There was no difference for white and non-white patients, either for aggregate SM or for the solid/hematologic categories. Compared with age- and sex-matched population and adjusting for race and stage, OS was worse with antecedent WM for colon cancer (HR, 2.0, 95% CI 1.4-2.7), melanoma (HR, 2.6, 95% CI 1.8-4.0) and DLBCL (HR, 1.9, 95% CI 1.2-3.0). It was not significantly different for acute leukemia (HR 0.9, 95% CI 0.5-1.4), prostate (HR 1.2, 95% CI 0.9-1.6), breast (HR 1.1, 95% CI 0.7-1.9), bladder (HR 1.2, 95% CI 0.8-1.7), thyroid (HR 0.9, 95% CI 0.1-6.0) or lung cancer (HR 1.2, 95% CI 1.0-1.4).Conclusions: Patients with WM have a 49% higher risk of a SM than the general population. Further research is needed to elucidate the increased incidence of leukemia and DLBCL possibly resulting from therapy and transformation, respectively, and melanoma, lung, bladder and thyroid cancers, possibly associated with defective immune surveillance. Comparatively poor survival in WM patients with colon cancer underscores the need for guideline-adherent screening and therapy.TableIncidenceaSIR95% CIExcess casesaAll malignancies6811.51.4-1.695Solid tumors4841.21.1-1.335Lung1011.51.2-1.814Prostate951.00.8-1.31Urinary tract621.41.1-1.88Colorectal480.90.7-1.2-2Other gastrointestinal431.00.7-1.40Breast431.00.8-1.41Melanoma351.91.3-2.77Other gynecologic211.30.8-2.02Head and neck130.90.5-1.60Thyroid102.71.3-4.93Hematologic malignancies1744.23.6-4.956All lymphomas1204.43.6-5.239Diffuse large B-cell314.32.9-6.110Extranodal marginal zone1614.88.5-24.16Other indolent B-cell193.72.2-5.86Myeloma314.73.2-6.610Acute leukemia153.21.8-5.34a per 10,000 person-years DisclosuresNo relevant conflicts of interest to declare.

Wiskott–Aldrich syndrome protein deficiency in natural killer and dendritic cells affects antitumor immunity

Wiskott-Aldrich syndrome (WAS) is a primary immunodeficiency caused by reduced or absent expression of the WAS protein (WASP). WAS patients are affected by microthrombocytopenia, recurrent infections, eczema, autoimmune diseases, and malignancies. Although immune deficiency has been proposed to play a role in tumor pathogenesis, there is little evidence on the correlation between immune cell defects and tumor susceptibility. Taking advantage of a tumor-prone model, we show that the lack of WASP induces early tumor onset because of defective immune surveillance. Consistently, the B16 melanoma model shows that tumor growth and the number of lung metastases are increased in the absence of WASP. We then investigated the in vivo contribution of Was(-/-) NK cells and DCs in controlling B16 melanoma development. We found fewer B16 metastases developed in the lungs of Was(-/-) mice that had received WT NK cells as compared with mice bearing Was(-/-) NK cells. Furthermore, we demonstrated that Was(-/-) DCs were less efficient in inducing NK-cell activation in vitro and in vivo. In summary, for the first time, we demonstrate in in vivo models that WASP deficiency affects resistance to tumor and causes impairment in the antitumor capacity of NK cells and DCs.

Role of Glycine N-Methyltransferase in the Regulation of T-Cell Responses in Experimental Autoimmune Encephalomyelitis

Glycine N-methyltransferase (GNMT) is known for its function as a tumor suppressor gene. Since 100% of female Gnmt(-/-) mice developed hepatocellular carcinoma, we hypothesized that Gnmt(-/-) mice may have defective immune surveillance. In this study, we examined the immune modulation of GNMT in T-cell responses using experimental autoimmune encephalomyelitis (EAE). The results showed that EAE severity was reduced significantly in Gnmt(-/-) mice. Pathological examination of the spinal cords revealed that Gnmt(-/-) mice had significantly lower levels of mononuclear cell infiltration and demyelination than the wild-type mice. In addition, quantitative real-time PCR showed that expression levels of proinflammatory cytokines, including interferon (IFN)-γ and interleukin (IL)-17A, were much lower in the spinal cord of Gnmt(-/-) than in that of wild-type mice. Accordingly, myelin oligodendrocyte glycoprotein (MOG)-specific T-cell proliferation and induction of T-helper (Th)1 and Th17 cells were markedly suppressed in MOG(35-55)-induced Gnmt(-/-) mice. Moreover, the number of regulatory T (Treg) cells was increased significantly in these mice. When the T-cell receptor was stimulated, the proliferative capacity and the activation status of mTOR-associated downstream signaling were decreased significantly in Gnmt(-/-) CD4(+) T cells via an IL-2- and CD25-independent manner. Moreover, GNMT deficiency enhanced the differentiation of Treg cells without affecting the differentiation of Th1 and Th17 cells. Furthermore, the severity of EAE in mice adoptive transferred with GNMT-deficient CD4(+) T cells was much milder than in those with wild-type CD4(+) T cells. In summary, our findings suggest that GNMT is involved in the pathogenesis of EAE and plays a crucial role in the regulation of CD4(+) T-cell functions.

Abstract 532: Nod1 functions in T cells to suppress inflammation-associated colorectal cancer

Abstract A major risk factor for the development of colorectal cancer is the presence of chronic inflammation within the colon. Innate immune receptors have emerged as key players in regulating intestinal homeostasis and the development of colorectal cancer in mouse models. There are two major classes of innate immune receptors, the Toll-like receptors and the Nod-like receptors (NLRs), which are important in pathogen recognition and eradication, but are also capable of sensing commensal bacteria within the gut. We have previously demonstrated that the ubiquitously expressed NLR family member, Nod1, acts as a tumor suppressor in the setting of chronic inflammation. The purpose of the current study is to better understand how Nod1 suppresses tumor development, which would be important in developing a chemoprevention strategy for inflammation-associated colon carcinogenesis using available Nod1 ligands. Using a mouse model of colitis-associated colon cancer in which mice are given a single intraperitoneal injection of the carcinogen, azoxymethane, followed by multiple rounds of water containing dextran sulfate sodium to induce chronic colitis, we generated bone marrow chimeras to determine whether Nod1 functions to suppress inflammation-related tumors in the intestinal epithelial versus hematopoietic compartment. We also performed adoptive transfer experiments to specifically determine the role of Nod1 in T cells in tumor suppression. Finally, Nod1 mice were crossed with T-cell deficient, TCRβ-/- mice, to further determine T-cell intrinsic properties of Nod1 in limiting tumor potential. Real-time PCR analysis and cultures of colon lamina propria cells from wildtype and Nod1-deficient mice were performed to determine recruitment of and production of cytokines by T cells within the colon. Our studies suggest that Nod1 functions in the hematopoietic cell compartment to protect against inflammation-induced tumorigenesis. More specifically, adoptive transfer of wildtype T cells was capable of rescuing Nod1-deficient mice from developing significant numbers of tumors, suggesting that Nod1 signaling in T cells is important for tumor suppression. Consistent with a T cell-specific function for Nod1 in tumor suppression, adoptive transfer of Nod1-deficient T cells in either TCRβ-deficient or TCRβ/Nod1 doubly-deficient mice resulted in significant numbers of tumors in contrast to adoptive transfer of wildtype T cells, which resulted in fewer tumors. Cytokine expression analysis of colon tissue as well as colon lamina propria cells demonstrated that Nod1-deficient mice exhibit an impairment in IFN-γ production during inflammation. These studies suggest a T-cell intrinsic role for Nod1 in protecting against inflammation-associated colon neoplasia, which may be related to a defect in immune surveillance. These observations set the stage for development of Nod1 agonists as colon cancer chemopreventive agents. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 532. doi:1538-7445.AM2012-532