Project description:We noticed that a recently identified poor prognosis stem/serrated molecular subtype of colorectal cancer (CRC) is characterized by up-regulation of transcripts known to be also expressed by stromal cells. To better define the origin of such transcripts, we analyzed RNAseq and microarray datasets from CRC mouse xenografts, where human cancer cells are supported by murine stroma. The analysis revealed that mRNA levels of stem/serrated subtype genes are mostly due to stromal expression, even when the stromal fraction is below 5%. Indeed, a classifier based on genes exclusively expressed by cancer-associated fibroblasts was significantly associated, in multiple datasets, to poor prognosis of CRC and to radioresistance of rectal cancer. Molecular Characterization of 72 primary rectal cancer formalin-fixed, paraffin-embedded (FFPE) specimens including 58 pretreatment specimens, 14 surgical specimens.
Project description:Aromatase inhibitors are first-line postmenopausal agents for estrogen receptor alpha (ERa)-positive breast cancer. However, there is considerable response heterogeneity and women frequently relapse. Estrogen deprivation does not completely arrest ERa activity, and transactivation of the unliganded receptor may continue through cross-talk with growth factor pathways. In contrast with aromatase inhibitors, the selective ER downregulator fulvestrant also abrogates ligand-independent ERa activity. The benefit of fulvestrant as an alternative, combination, or sequential therapy to aromatase inhibitor has been reported, but molecular mechanisms underpinning its relative efficacy remain unclear and biomarkers for patient selection are lacking. This study demonstrates, for the first time, that the overall transcriptional response to fulvestrant is of greater magnitude than estrogen deprivation, consistent with its clinical efficacy and more complete blockade of estrogenic signaling. Using a robust integrative approach, we identify a subset of genes differentially affected by fulvestrant that comprises distinct biologic networks, correlates with antiproliferative response, and has potential utility as predictive biomarkers for fulvestrant. Global gene expression profiles from ERα-positive breast carcinomas before and during presurgical treatment with fulvestrant (n = 38) or anastrozole (n = 81), and corresponding in vitro models, were compared. Transcripts responding differently to fulvestrant and estrogen deprivation were identified and integrated using Gene Ontology, pathway and network analyses to evaluate their potential significance. --------------------------------- This represents the data for fulvestrant only
Project description:Committed precursors of conventional dendritic cells (pre-cDCs) derived from the common DC progenitor which differentiate into cDC subpopulations in peripheral tissues have been identified, but committed precursors for plasmacytoid DCs (pDCs) have not been found. Here we show that CDP-derived ‘CCR9- MHCIIlow BST2+ Siglec-H+ pDCs from murine bone marrow which enter the circulation and peripheral tissues have a common DC precursor function in vivo in the steady state. Upon adoptive transfer the fate of CCR9- pDC-like precursors is governed by the tissues they enter. In the bone marrow and liver most transferred CCR9- pDC-like precursors differentiate into CCR9+ pDCs, whereas in peripheral lymphoid organs, lung and intestine they can give rise to CCR9+ pDCs and cDCs. Thus, CCR9- pDC-like cells are novel CDP-derived circulating DC precursors with pDC and cDC potential, whose final differentiation depends on tissue-specific factors allowing adaptation to local requirements. Total RNA obtained from CCR9- pDC-like common DC progenitors and CCR9+ pDCs was compared for differential gene expression. 3 independent isolations were performed for the 2 samples. 6 arrays were run in total.
Project description:This experiment was carried out in the context of a pharmacogenetic study of long-term (4-year follow-up) response to Interferon-beta treatment in two cohorts of Italian Multiple Sclerosis patients, to identify genetic variants (SNPs) that may influence response to IFN-beta. We integrated results from meta-analysis of the two cohorts with gene expression profiling of IFNβ stimulated PBMCs from 20 healthy controls and eQTL analyses, to look at possible enrichment of IFN-beta induced genes with genes mapped by top-ranking meta-analyzed SNPs. Total RNA obtained from isolated PBMC extracted from healthy subjects and in vitro stimulated or not with Interferon-β (100IU/mL) for 18h
Project description:Phosphoinositide-3-kinase/protein-kinaseB/mammalian target of rapamycin (PI3K/AKT/mTOR) signalling plays an important role in breast cancer (BC). Its interaction with estrogen receptor (ER) signalling becomes more complex and inter-dependent with acquired endocrine resistance. Targeting mTOR combined with endocrine therapy has shown clinical utility, however, a negative feedback-loop exists downstream of PI3K/AKT/mTOR. Direct blockade of AKT together with endocrine therapy may improve BC treatment. AZD5363, a novel pan-AKT kinase catalytic inhibitor, was examined in a panel of ER+ BC cell lines (MCF7, HCC1428, T47D, ZR75.1) adapted to long-term-estrogen-deprivation (LTED) or tamoxifen (TamR). AZD5363 caused a dose-dependent decrease in proliferation in all cell lines tested (GI50<500nM) except HCC1428 and HCC1428-LTED. T47D-LTED and ZR75-LTED were the most sensitive of the lines (GI50~100nM). AZD5363 re-sensitised TamR cells to tamoxifen and acted synergistically with fulvestrant. AZD5363 decreased p-AKT/mTOR targets leading to a reduction in ERα-mediated transcription in a context specific manner and concomitant decrease in recruitment of ER and CREB-binding protein (CBP) to estrogen-response-elements located on the TFF1, PGR and GREB1 promoters. Furthermore, AZD5363 reduced expression of cell-cycle-regulatory proteins. Global gene expression highlighted ERBB2-ERBB3, ERK5 and IGF1 signaling pathways driven by MYC as potential feedback-loops. Combined treatment with AZD5363 and fulvestrant showed synergy in an ER+ patient derived xenograft and delayed tumour progression post-cessation of therapy. These data support the combination of AZD5363 with fulvestrant as a potential therapy for BC that is sensitive or resistant to E-deprivation or tamoxifen and that activated AKT is a determinant of response, supporting the need for clinical evaluation. Cell lines were treated in biological triplicates in the absence of estrogen with or without AZD5363 for 24hours in order to identify gene changes associated with perturbation of AKT signalling
Project description:The aim of the array was to compare iPS-derived CD19+CD10+ B cells with B cells from umbilical cord blood (UCB) and adult peripheral blood (PB) iPS-CD34+ cells and UCB-CD133+ cells were differentiated to the B cell lineage in vitro. B cells were also isolated directly from UCB and PB. RNA was extracted from CD19+CD10+ FACS isolated cells from all 4 B cell samples as well as from undifferentiated iPS cells.
Project description:The present research aimed to investigate peripheral blood gene expression profiling as a minimally invasive surrogate approach to study silica-induced pulmonary toxicity. Rats were exposed to crystalline silica by inhalation (15 mg/m3, 6 hours/day, 5 days). Pulmonary damage and blood gene expression profiles were determined at various latency periods (0 - 16 weeks). Silica exposure resulted in pulmonary toxicity and this was evidenced by histological changes in the lungs and elevation of LDH activity, and total protein and albumin contents in the bronchoalveolar lavage fluid (BALF) of the rats. Microarray analysis of global gene expression profiles in the blood of the rats identified genes that were differentially expressed in response to silica exposure and toxicity. The number of significantly differentially expressed genes in the blood of silica exposed rats correlated with the severity of silica-induced pulmonary toxicity. Genes involved in biological functions such as inflammatory response, cancer, pulmonary damage, oxidative stress, energy metabolism, fibrosis, etc, were found differentially expressed in the blood of the silica exposed rats compared with the controls. Induction of pulmonary inflammation in the silica exposed rats, as suggested by differential expression of inflammatory response genes in the blood, was supported by significant increases in the number of macrophages and infiltrating neutrophils as well as the activity of pro-inflammatory chemokines M-bM-^@M-^S MCP1 and MIP2, observed in the BALF of the silica exposed rats. A silica-responsive blood gene expression signature developed using the gene expression data predicted with significant accuracy the exposure of rats to lower concentrations (1 and 2 mg/m3) of silica. Taken together our findings suggest the potential application of peripheral blood gene expression profiling as a minimally invasive and efficient surrogate approach to detect and study silica-induced pulmonary toxicity. 96 samples were analyzed in this experiment. The RNA from rat blood samples was isolated for gene expression studies. Rats (48) were exposed to crystalline silica by inhalation (15 mg/m3, 6 hours/day, 5 days) and air (24). Blood gene expression profiling was performed using rat blood samples, 8 each for silica exposed and 4 each for air exposed controls at various post-exposure time periods (0, 1, 2, 4, 8, and 16 weeks). A silica-responsive blood gene expression signature was developed using the gene expression data obtained from the 0 week post-exposure group and the control rats. The signature was tested for predicting silica exposure and resulting toxicity in the rats exposed to lower concentrations (1 and 2 mg/m3) of silica (8 rats per group) and air (8 rats).
Project description:Analysis of the cystic fibrosis gene Cftr in the colon and small intestine of Cftr-deficient murine model. The hypothesis was loss of Cftr altered expression of genes important in intestinal homeostasis and oncogenic signaling pathways. The results identified potential roles of Cftr in up- or down-regulating major gene clusters that belong to groups of immune response, ion channel, intestinal stem cell and other growth regulators. Total RNA was isolated from the normal intestine of three Apc wildtype Cftr wildtype and three Apc Cftr-deficient mice. For the colon intestinal epithelia from the same region of the distal colon of each mouse was separated from the rest of the intestine prior to RNA isolation. Therefore RNA was obtained from only epithelial cells. For the small intestine, a section of the mid-duodenum from each mouse was sheared of villi prior to RNA isolation. Therefore RNA was obtained from whole duodenum (minus villi), containing epithelia cells but also stromal and other cells. RNA Seq was then conducted on all samples, with at least two replicates for each biological sample.