Transcriptome Analysis Describing New Immunity and Defense Genes in Peripheral Blood Mononuclear Cells of Rheumatoid Arthritis Patients

Background: Large-scale gene expression profiling of peripheral blood mononuclear cells from Rheumatoid Arthritis (RA) patients could provide a molecular description that reflects the contribution of diverse cellular responses associated with this disease. The aim of our study was to identify peripheral blood gene expression profiles for RA patients, using Illumina technology, to gain insights into RA molecular mechanisms. Methodology/Principal Findings: The Illumina Human-6v2 Expression BeadChips were used for a complete genome-wide transcript profiling of peripheral blood mononuclear cells (PBMCs) from 18 RA patients and 15 controls. Differential analysis per gene was performed with one-way analysis of variance (ANOVA) and P values were adjusted to control the False Discovery Rate (FDR < 5%). Genes differentially expressed at significant level between patients and controls were analyzed using Gene Ontology (GO) in the PANTHER database to identify biological processes. A differentially expression of 339 Reference Sequence genes (238 down-regulated and 101 up-regulated) between the two groups was observed. We identified a remarkably elevated expression of a spectrum of genes involved in Immunity and Defense in PBMCs of RA patients compared to controls. This result is confirmed by GO analysis, suggesting that these genes could be activated systemically in RA. No significant down-regulated ontology groups were found. Microarray data were validated by real time PCR in a set of nine genes showing a high degree of correlation. Conclusions/Significance: Our study highlighted several new genes that could contribute in the identification of innovative clinical biomarkers for diagnostic procedures and therapeutic interventions

Tumor-Induced Cholesterol Efflux from Macrophages Drives IL-4 Mediated Reprogramming and Tumor Progression

Tumor-associated macrophages (TAM) have been shown to have important roles in the malignant progression of various cancers. However, macrophages also posses intrinsic tumoricidal activity and can promote the activity of cytotoxic lymphocytes, but they rapidly adopt an alternative phenotype within tumors, associated with immune-suppression and trophic functions that support tumor growth. The mechanisms that promote TAM polarization in the tumor-microenvironment remain poorly understood, these mechanisms may represent important therapeutic targets to block the tumor-promoting functions of TAM and restore their anti-tumor potential. Here we have characterized TAM in a mouse model of metastatic ovarian cancer. We show that ovarian cancer cells promote membrane-cholesterol efflux and the depletion of lipid rafts from macrophages. Increased cholesterol efflux promoted IL-4 mediated reprogramming while inhibiting IFNγ-induced gene expression. These studies reveal an unexpected role for tumor-induced membrane-cholesterol efflux in driving the IL-4 signaling and the tumor-promoting functions of TAM, while rendering them refractory to pro-inflammatory stimuli. Thus, preventing cholesterol efflux in TAM could represent a novel therapeutic strategy to block pro-tumor functions and restore anti-tumor immunity. Biopharmaceutic

Naïve rat umbilical cord matrix stem cells significantly attenuate mammary tumor growth through modulation of endogenous immune responses

Background: Un-engineered human and rat umbilical cord matrix stem cells (rUCMSC) attenuate growth of several types of tumors in mice and rats. However, the mechanism by which UCMSC attenuate tumor growth has not been studied rigorously. Methods- The possible mechanisms of tumor growth attenuation by rUCMSC were studied using orthotopic Mat B III rat mammary tumor grafts in female F344 rats. Tumor-infiltrating leukocytes were identified and quantified by immunohistochemical image analysis. Potential cytokines involved in lymphocyte infiltration in the tumors were determined by microarray and Western blot analysis. The Boyden chamber migration assay was performed for the functional analysis of identified cytokines. Results: rUCMSC markedly attenuated the tumor growth; this attenuation was accompanied by considerable lymphocyte infiltration. Immunohistochemical analysis revealed that the majority of infiltrating lymphocytes in the rUCMSC-treated tumors were CD3+ T cells. In addition, treatment with rUCMSC significantly increased infiltration of CD 8+ and CD4+ T cells and NK cells throughout tumor tissue. CD68+ monocytes/macrophages and FoxP3+ regulatory T cells were scarcely observed, only in the tumors of the PBS control group. Microarray analysis of rUCMSC identified that monocyte chemotactic protein (MCP)-1 is involved in rUCMSCinduced lymphocyte infiltration in the tumor tissues. Discussion: These results suggest that naïve rUCMSC attenuated mammary tumor growth at least in part by enhancing host anti-tumor immune responses. Thus, naïve UCMSC can be used as powerful therapeutic cells for breast cancer treatment, and MCP-1 may be a key molecule to enhance the effect of UCMSC at the tumor site

TREM2 inhibition triggers antitumor cell activity of myeloid cells in glioblastoma

Triggering receptor expressed on myeloid cells 2 (TREM2) plays important roles in brain microglial function in neurodegenerative diseases, but the role of TREM2 in the GBM TME has not been examined. Here, we found that TREM2 is highly expressed in myeloid subsets, including macrophages and microglia in human and mouse GBM tumors and that high TREM2 expression correlates with poor prognosis in patients with GBM. TREM2 loss of function in human macrophages and mouse myeloid cells increased interferon-γ-induced immunoactivation, proinflammatory polarization, and tumoricidal capacity. In orthotopic mouse GBM models, mice with chronic and acute Trem2 loss of function exhibited decreased tumor growth and increased survival. Trem2 inhibition reprogrammed myeloid phenotypes and increased programmed cell death protein 1 (PD-1

Genomic analysis of macrophage gene signatures during idiopathic pulmonary fibrosis development

Idiopathic Pulmonary Fibrosis (IPF) is a chronic, progressive, irreversible lung disease. After diagnosis, the interstitial condition commonly presents 3-5 years of life expectancy if untreated. Despite the limited capacity of recapitulating IPF, animal models have been useful for identifying related pathways relevant for drug discovery and diagnostic tools development. Using these techniques, several immune-related mechanisms have been implicated to IPF. For instance, subpopulations of macrophages and monocytes-derived cells are recognized as centrally active in pulmonary immunological processes. One of the most used technologies is high-throughput gene expression analysis, which has been available for almost two decades now. The “omics” revolution has presented major impacts on macrophage and pulmonary fibrosis research. The present study aims to investigate macrophage dynamics within the context of IPF at the transcriptomic level. Using publicly available gene-expression data, we applied modern data science approaches to (1) understand longitudinal profiles within IPF models; (2) investigate correlation between macrophage genomic dynamics and IPF development; and (3) apply longitudinal profiles uncovered through multivariate data analysis to the development of new sets of predictors able to classify IPF and control samples accordingly. Principal Component Analysis and Hierarchical Clustering showed that our pipeline was able to construct a complex set of biomarker candidates that together outperformed gene expression alone in separating treatment groups in an IPF animal model dataset. We further assessed the predictive performance of our candidates on publicly available gene expression data from IPF patients. Once again, the constructed biomarker candidates were significantly differentiated between IPF and control samples. The data presented in this work strongly suggest that longitudinal data analysis holds major unappreciated potentials for translational medicine research

Immunomodulation of myeloid-derived suppressor cells by particulate b-glucan in cancer.

Myeloid-derived suppressor cells (MDSC) are a heterogeneous population of immature myeloid cells that promote tumor progression. In this study, we investigated the effect of dectin-1 stimulation by yeast-derived particulate β-glucan in MDSC function and differentiation in cancer. In vivo treatment of mice bearing lewis lung carcinoma and mammary cell carcinoma with particulate β-glucan decreased tumor weight and splenomegaly, and reduced the accumulation of polymorphonuclear-MDSC (PMN-MDSC) but not monocytic-MDSC (M-MDSC) in the spleen and tumor. In addition, particulate β-glucan differentially modulated the function of different MDSC subsets; it enhanced PMN-MDSC respiratory burst and apoptosis, and induced the differentiation of M-MDSC into F4/80+CD11c+antigen-presenting cells in a dectin-1 dependent manner. ERK1/2 phosphorylation was also required for the acquisition of APC properties in M-MDSC. Moreover, M-MSDC treated with particulate β-glucan did not promote tumor growth in vivo when inoculated with LLC subcutaneously. To evaluate the effect of particulate β-glucan treatment in humans, patients with non-small cell lung cancer (NSCL) were treated with particulate β-glucan for two weeks prior to any other treatment and surgical excision of the tumor. Strikingly, the frequency of CD14-HLA-DR-CD11b+CD33+ MDSC decreased in the peripheral blood, and arginase-1 expression significantly decreased in a cohort of 15 patients. This study was the first to assess the effect of particulate β-glucan on MDSC in lung cancer patients, towards a future inclusion of particulate β-glucan in combination therapies in lung cancer

Integrated molecular profiles of invasive breast tumors and ductal carcinoma in situ (DCIS) reveal differential vascular and interleukin signaling

We use an integrated approach to understand breast cancer heterogeneity by modeling mRNA, copy number alterations, microRNAs, and methylation in a pathway context utilizing the pathway recognition algorithm using data integration on genomic models (PARADIGM). We demonstrate that combining mRNA expression and DNA copy number classified the patients in groups that provide the best predictive value with respect to prognosis and identified key molecular and stromal signatures. A chronic inflammatory signature, which promotes the development and/or progression of various epithelial tumors, is uniformly present in all breast cancers. We further demonstrate that within the adaptive immune lineage, the strongest predictor of good outcome is the acquisition of a gene signature that favors a high T-helper 1 (Th1)/cytotoxic T-lymphocyte response at the expense of Th2-driven humoral immunity. Patients who have breast cancer with a basal HER2-negative molecular profile (PDGM2) are characterized by high expression of protumorigenic Th2/humoral-related genes (24–38%) and a low Th1/Th2 ratio. The luminal molecular subtypes are again differentiated by low or high FOXM1 and ERBB4 signaling. We show that the interleukin signaling profiles observed in invasive cancers are absent or weakly expressed in healthy tissue but already prominent in ductal carcinoma in situ, together with ECM and cell-cell adhesion regulating pathways. The most prominent difference between low and high mammographic density in healthy breast tissue by PARADIGM was that of STAT4 signaling. In conclusion, by means of a pathway-based modeling methodology (PARADIGM) integrating different layers of molecular data from whole-tumor samples, we demonstrate that we can stratify immune signatures that predict patient survival

The inflammatory infiltrate of high-grade serous carcinoma omental metastasis

PhDThe aim of this thesis is to investigate the role of inflammatory infiltrates and chemokines in metastasis of high-grade serous ovarian cancer, HGSC, to the omentum using human tissue biopsies and a 3- dimensional (3D) cell culture model. In ten patients with metastatic HGSC, omental tumour deposits contained a prominent leukocyte infiltrate of CD3+ T cells (9% of total cells) and CD68+ macrophages (11% of total cells). The presence of CD68+ macrophages showed a significant positive correlation with tumour cell proliferation analysed by Ki67 expression. Four ovarian cancer cell lines were co-cultured on a 3D model mimicking the microenvironment of the omentum for two weeks. The model was composed of collagen embedded human fibroblasts covered in a confluent layer of human primary mesothelial cells. The mesothelial cells in the 3D model significantly increased the growth (p = 0.002) and invasion (p = 0.0004) of the ovarian cancer cells. CXCL12 is the macrophage chemoattractant and ligand for the major chemokine receptor expressed on ovarian cancer cells. An association between CXCL12 and extracellular matrix remodelling was identified in two independent gene expression microarrays of ovarian cancer biopsies. The expression of CXCL12 in the HGSC omental metastases measured by quantitative Real Time-PCR positively correlated with decorin expression. Antibody mediated neutralisation of CXCL12 reduced growth (p = 0.012) and invasion (p = 0.029) in the 3D model. Mimicking an infiltrate of CD68+ macrophages in this multicellular 3D in vitro system also produced measurable changes in inflammatory cytokine and chemokine expression. There is currently a demand for more accurate models of HGSC and a necessity to study its metastasis that presents itself as the major clinical problem in patients. Therefore the development of this 3D model to mimic tumour-promoting inflammation in HGSC metastasis will provide researchers with an essential tool for testing novel therapeutic strategies.This work was supported by the BBSRC (Biotechnology and Biological Sciences Research Council) and AstraZeneca in an Industrial Case Studentship, grant number BB/G017867/1