Filters
Technology
Platform
SRP127564
Mus musculus
79 Downloadable Samples
Illumina HiSeq 2500
Description
Purpose: Eliciting effective anti-tumor immune responses in patients who fail checkpoint inhibitor therapy is a critical challenge in cancer immunotherapy, and in such patients, tumor-associated myeloid cells and macrophages (TAMs) are promising therapeutic targets. We demonstrate in an autochthonous, poorly immunogenic mouse model of melanoma that combination therapy with an agonistic anti-CD40 mAb and CSF1R inhibitor potently suppressed tumor growth. Microwell assays to measure multiplex protein secretion by single cells identified that untreated tumors have distinct TAM subpopulations secreting MMP9 or co-secreting CCL17/22, characteristic of an M2-like state. Combination therapy reduced the frequency of these subsets, while simultaneously inducing a separate polyfunctional inflammatory TAM subset co-secreting TNF?, IL-6, and IL-12. Tumor suppression by this combined therapy was partially dependent on T cells, TNF? and IFN?. Together, this study demonstrates the potential for targeting TAMs to convert a “cold” into an “inflamed” tumor microenvironment capable of eliciting protective T cell responses. Methods: Total RNA was purified with the use of QIAzol and RNeasy Mini kit (QIAGEN), in which an on-column DNase treatment was included. Purified RNA was submitted to the Yale Center for Genomic Analysis where it was subjected to mRNA isolation and library preparation. Non-strand specific libraries were generated from 50ng total RNA using the SMARTer Ultra Low Input RNA for Illumina Sequencing kit. Libraries were pooled, six samples per lane, and sequenced on an Illumina HiSeq 2500 (75-bp paired end reads), and aligned using STAR to the GRCm38 (mm10) reference genome. A count-based differential expression protocol was adapted for this analysis(Anders et al., 2013); mappable data were counted using HTSeq, and imported into R for differential expression analysis using the DESeq2.To find differentially regulated sets of genes for signature generation, a 1.5-Log2 fold-change difference between samples and p-adjusted (Holm-Sidak) = 0.01 was used. Results: To begin to understand how these treatments modulated T cells to control tumor growth, and to possibly illuminate additional biomarkers of response, we examined the transcriptomes of CD11b+ Ly6G- cells treated with CD40 or CSF1Ri, alone or in combination, relative to control, using high throughput RNA-sequencing. Principal components analysis (PCA) on the genome-wide dataset demonstrated that treating with CD40 and CSF1Ri individually caused largely non-overlapping changes in transcription, as indicated by their movement along orthogonal principal components (PC) relative to the control. Importantly, combination therapy was visualized as a systems-level combination of each individual treatment in PC space. We then examined the mRNAs most altered by either treatment alone or in combination relative to Controls (Log2FC>1.5, p<.01) by unsupervised hierarchical clustering. Five major gene patterns emerged from the clustering of genes. Cluster #1 comprises genes that are upregulated by CD40 and CSF1Ri+CD40 treatment but are mostly unaffected by CSF1Ri, suggesting that CD40 is the primary driver of this cluster in the combination treatment. Notable genes in this cluster include Tnfa, Ifng??Il12b and Cxcl9; interestingly, for Tnfa and Il12b, CSF1Ri+CD40 appears to have a synergistic effect on expression. In contrast to Cluster #1, Cluster #5 contains genes substantially downregulated by CSF1Ri and CSF1Ri+CD40 treatments, but are largely unaffected by CD40, suggesting that CSF1Ri is the driver of this cluster in the combination treatment. Cluster #5 genes include Cd36 and Fabp4, suggesting alterations in lipid homeostasis in the TAMs after treatment. Cluster #2 includes genes that are modestly upregulated by CD40 and CSF1Ri individually, leading to a stronger upregulation when combined. Finally, Clusters #3 and #4 include, for the most part, genes that are differentially affected by CD40 versus CSF1Ri and for which the combination treatment yields an intermediate response. In summary, these data show that CSF1Ri and CD40 agonism elicit predominantly distinct changes in gene expression in the CD11b+ cells, indicating they target different biological processes in myeloid cells. The net result of the changes in myeloid gene expression from the combination of CSF1Ri+CD40 treatment reveal additive effects by the individual treatments, but also synergy in the expression of several pro-inflammatory genes (e.g., Tnfa, Ifng, Il6 and Il12b). We further examined our dataset with Gene Set Enrichment Analysis (GSEA). Although CSF1Ri and CD40 treatments did not closely match any immunological signatures in the immunological database of MSigDb, combined CSF1Ri+CD40 had a strikingly similar signature to myeloid cells exposed to a variety of inflammatory stimulants, most closely reflected by BMDMs treated with lipopolysaccharide (LPS). This motivated us to look specifically at categories of NF-?B target genes that are significantly affected by LPS treatment, including transcription factors, cytokines and chemokines. Indeed, most of these NF-?B target genes associated with inflammation were strongly upregulated by CSF1Ri+CD40 treatment. Finally, Ingenuity Pathway Analysis identified TNFR1 and TNFR2 signaling and Acute phase response signaling among the top genetic signatures produced by the CSF1Ri+CD40 treatment combination, matching what we observed with GSEA. Thus, gene expression analysis not only revealed several biomarkers of response that may be relevant for assessing therapeutic activity in ongoing clinical trials using these drugs, but illuminated lead biological factors that may cause tumor regression. Conclusions: myeloid-targeted immunotherapies anti-CD40+CSF1R inhibition synergistically induce a pro-inflammatory microenviroment Overall design: mRNA profiles of tumor infiltrating lymphocytes (TILs) in mice were generated by deep sequencing, in triplicate, using Illumina.
Publication Title
Myeloid-targeted immunotherapies act in synergy to induce inflammation and antitumor immunity.
Sample Metadata Fields
Specimen part, Cell line, Subject
View Samples- Rattus norvegicus
- 12 Downloadable Samples
Affymetrix Rat Genome 230 2.0 Array (rat2302)
Description
Several different mechanical signals have been proposed to control the extent and pattern of myocardial growth and remodeling, though this has largely been studied using in vitro model systems that are not representative of intact myocardium or in vivo models in which isolating the effects of individual candidate stimuli is exceedigly difficult. We used a unique tissue culture system that allows the simultaneous control of multiple mechanical inputs and other potentially confounding stimuli (e.g., hormonal).
Publication Title
Effects of stretch and shortening on gene expression in intact myocardium.
Sample Metadata Fields
Sex, Age
View Samples- Mus musculus
- 12 Downloadable Samples
- Affymetrix Mouse Gene 2.0 ST Array (mogene20st)
Description
Expression data from antigen-experienced Nfat1+/+ and Nfat1-/- CD4+ T cells following 21 days of Plasmodium yoelii 17XNL infection.
Publication Title
The Transcription Factor NFAT1 Participates in the Induction of CD4<sup>+</sup> T Cell Functional Exhaustion during Plasmodium yoelii Infection.
Sample Metadata Fields
Sex, Specimen part
View Samples- Homo sapiens, Rattus norvegicus
- 2 Downloadable Samples
- Affymetrix Human Genome U133 Plus 2.0 Array (hgu133plus2)
Description
MicroRNAs (miRNAs) regulate many basic aspects of cell biology including neuronal plasticity, but little is known of their roles in drug addiction. Extended access to cocaine can trigger the emergence of compulsive drug-seeking behaviors, but molecular mechanisms regulating this process remain unclear. Here we report that microRNA-212 (miR-212) is upregulated in the dorsal striatum of rats with extended access to cocaine. Striatal overexpression of miR-212 decreases, whereas its inhibition increases cocaine intake in rats with extended but not restricted drug access, suggesting that miR-212 serves as a protective factor against the development of compulsive drug seeking. The transcription factor CREB (cAMP response element-binding protein) is considered a core regulator of cocaine reward. We show that miR-212 controls responsiveness to cocaine by dramatically amplifying striatal CREB signaling. This action occurs through miR-212-enhanced Raf-1 activity, resulting in adenylyl cyclase sensitization and increased expression of the essential CREB co-activator TORC (Transducer of Regulated CREB; also known as CRTC). Our findings suggest that striatal miR-212 signaling plays a key role in vulnerability to addiction, and that noncoding RNAs such as the miRNAs may serve as novel targets for the development of anti-addiction therapeutics.
Publication Title
Striatal microRNA controls cocaine intake through CREB signalling.
Sample Metadata Fields
Sex, Specimen part, Cell line
View Samples- Homo sapiens
- 12 Downloadable Samples
- Affymetrix Human Gene 1.0 ST Array (hugene10st)
Description
This study characterizes the response of primary human endothelial cells (human umbilical vein endothelial cells, HUVECs) to the relative shear stress changes that occur during the initiation of arteriogenesis at the entrance regions to a collateral artery network. HUVECs were preconditioned to a baseline level of unidirectional shear of 15 dynes/cm2 for 24 hours. After 24 hours preconditioning, HUVECs were subjected to an arteriogenic stimulus that mimics the shear stress changes observed in the opposing entrance regions into a collateral artery network. The arteriogenic stimulus consisted of a 100% step wise increase in shear stress magnitude to a unidirectional 30 dynes/cm2 in either the same or opposite direction of the preconditioned shear stress. This simulates either the feeding entrance to the collateral artery circuit or the region that drains into the vasculature downstream of an obstruction in a major artery, respectively. In vivo analysis of collateral growth in the mouse hindlimb showed enhanced outward remodeling in the re-entrant (direction reversing) region that reconnects to the downstream arterial tree, suggesting reversal of shear stress direction as a key enhancer of arteriogenesis. Transcriptional profiling using microarray techniques identified that the reversal of shear stress direction, but not an increase in shear stress alone, yielded a broad-based enhancement of the mechanotransduction pathways necessary for the induction of arteriogenesis.
Publication Title
Mechanisms of Amplified Arteriogenesis in Collateral Artery Segments Exposed to Reversed Flow Direction.
Sample Metadata Fields
Specimen part
View Samples- Mus musculus
- 4 Downloadable Samples
- Affymetrix Mouse Genome 430 2.0 Array (mouse4302)
Description
Aims: Resident cardiac progenitor cells show homing properties when injected into the injured but not into the healthy myocardium. The molecular background behind this difference in behavior needs to be studied to elucidate how adult progenitor cells can restore cardiac function of the damaged myocardium. Since the brain-derived neurotrophic factor (BDNF) moderates cardioprotection in injured hearts, we focused on delineating its regulatory role in the damaged myocardium.
Publication Title
Brain derived neurotrophic factor contributes to the cardiogenic potential of adult resident progenitor cells in failing murine heart.
Sample Metadata Fields
Age, Specimen part, Disease, Disease stage
View Samples- Homo sapiens
- 16 Downloadable Samples
- Affymetrix Human Genome U133A Array (hgu133a)
Description
Inflammatory hepatocellular adenomas (IHCA) are benign liver tumours defined by the presence of inflammatory infiltrates and by the elevated expression of inflammatory proteins in tumour hepatocytes1,2. Here we show a striking activation of the IL6 signalling pathway in this tumour type, and sequencing candidate genes pinpointed this response to somatic gain-of-function mutations in the IL6ST gene that encodes the signalling co-receptor gp130. Indeed, ~70% of IHCA harbour small in-frame deletions that target the binding site of gp130 for IL6, and expression of the most frequent gp130 mutant, Delta-STVY190, in hepatocellular cells activates STAT3 in absence of ligand. Further, analysis of hepatocellular carcinomas revealed rare gp130 alterations always accompanied by -catenin-activating mutations, suggesting a cooperative effect of these signalling pathways in the malignant conversion of hepatocytes. The recurrent gain-of-function gp130 mutations in these human hepatocellular adenomas explains their inflammatory phenotype, and suggest that similar alterations may occur in other inflammatory epithelial tumours with STAT3 activation.
Publication Title
Frequent in-frame somatic deletions activate gp130 in inflammatory hepatocellular tumours.
Sample Metadata Fields
Sex, Specimen part, Disease
View Samples- Homo sapiens
- 46 Downloadable Samples
- Affymetrix Human Genome U133 Plus 2.0 Array (hgu133plus2)
Description
Background: Immunoadsorption with subsequent IgG substitution (IA/IgG) represents a novel therapeutic approach in treatment of dilated cardiomyopathy (DCM) which leads to improvement of left ventricular ejection fraction (LVEF). However, response to this therapeutic intervention shows wide inter-individual variability. In this pilot study, we tested the value of clinical, biochemical and molecular parameters for prediction of the response of patients with DCM to IA/IgG.
Publication Title
Myocardial gene expression profiles and cardiodepressant autoantibodies predict response of patients with dilated cardiomyopathy to immunoadsorption therapy.
Sample Metadata Fields
Sex, Age, Disease
View Samples- Homo sapiens
- 10 Downloadable Samples
- Affymetrix Human Genome U133 Plus 2.0 Array (hgu133plus2)
Description
Objectives: We studied the signal transduction of atrial structural remodelling that contributes to
Publication Title
Rac1-induced connective tissue growth factor regulates connexin 43 and N-cadherin expression in atrial fibrillation.
Sample Metadata Fields
Specimen part
View Samples- Homo sapiens
- 4 Downloadable Samples
- Affymetrix Human Genome U133 Plus 2.0 Array (hgu133plus2)
Description
Activation of the immune system is a way for host tissue to defend itself against tumor growth. Hence, treatment strategies that are based on immunomodulation are on the rise. Conventional cytostatic drugs such as the anthracycline doxorubicin can also activate immune cell functions of macrophages and natural killer cells. In addition, cytotoxicity of doxorubicin can be enhanced by combining this drug with the cytokine IFN-alpha. Although doxorubicin is one of the most applied cytostatics, the molecular mechanisms of its immunomodulation ability are not investigated thoroughly. In microarray analyses of HeLa cells, a set of 19 genes related to interferon signaling was significantly overrepresented among genes regulated by doxorubicin exposure including STAT-1, -2, IRF9, NMI, and caspase 1. Regulation of these genes by doxorubicin was verified with Real-Time PCR and immunoblotting. An enhanced secretion of IFN-alpha was observed when HeLa cells were exposed to doxorubicin as compared to untreated cells. IFN-alpha neutralizing antibodies and inhibitors of JAK-STAT signaling (ATA and AG490) significantly abolished doxorubicin-stimulated expression of interferon signaling-related genes. Furthermore, inhibition of JAK-STAT signaling significantly reduced doxorubicin induced caspase 3 activation and desensitized HeLa cells to doxorubicin cytotoxicity. In conclusion, we demonstrate that doxorubicin induces interferon-responsive genes via IFN-alpha-JAK-STAT1 signaling and that this pathway is relevant for doxorubicins cytotoxicity in HeLa cells. As immunomodulation is a promising strategy in anticancer treatment, this novel mode of action of doxorubicin may help to further improve the use of this drug among different types of anticancer treatment strategies.
Publication Title
Regulation of interferon-inducible proteins by doxorubicin via interferon γ-Janus tyrosine kinase-signal transducer and activator of transcription signaling in tumor cells.
Sample Metadata Fields
Cell line, Treatment
View Samples