T cell egress via lymphatic vessels limits the intratumoral T cell repertoire in melanoma
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ABSTRACT:
ORGANISM(S): Mus musculus
PROVIDER: GSE218455 | GEO | 2023/01/20
REPOSITORIES: GEO
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Project description:CXCL12 is a chemokine that binds to its cognate receptors CXCR4 and ACKR3 (CXCR7). An increase in CXCL12 concentrations has been used as a pharmacodynamic biomarker to assess ACKR3 antagonism in healthy adults. Furthermore, increased CXCL12 concentrations have been observed in various human pathologies. To date, CXCL12 concentrations have typically been quantified using antibody-based assays with overlapping or unclear specificity for the various existing CXCL12 proteoforms. Only the N-terminal full-length CXCL12 proteoform is biologically active and can engage CXCR4 and ACKR3. However, this proteoform could so far not be quantified in healthy adults. Here, we describe a new and validated fit-for-purpose immunoaffinity mass spectrometry (IA-MS) biomarker assay for specific measurement of five CXCL12α proteoforms in human plasma, including the biologically active CXCL12α proteoform. The assay was employed in a Phase 1 clinical study with the ACKR3 antagonist ACT-1004-1239 to quantify CXCL12α proteoforms. At baseline levels, 1.00 nM total CXCL12α and 0.10 nM biologically active CXCL12α was quantified in placebo treated adults. The concentrations of both proteoforms increased up to two-fold in healthy adults following drug administration. At all doses, 10% of CXCL12α was biologically active and the simultaneous increase of all proteoforms suggests that a new equilibrium has been reached 24 h following dosing. Hence, this IA-MS biomarker assay can be used to specifically measure active CXCL12 proteoform concentrations in clinical trials. Specific quantification of active chemokines can support decision making in clinical trial and thus successful drug development.
2024-01-27 | PXD038814 | Pride
Project description:Chemotaxis is an essential physiological process, often harnessed by tumors for metastasis. CXCR4, its ligand CXCL12 and the atypical receptor ACKR3 are overexpressed in many human cancers. Interfering with this axis by ACKR3 deletion impairs lymphoma cell migration towards CXCL12. Here, we propose a model of how ACKR3 controls the migration of the diffused large B-cell lymphoma VAL cells in vitro and in vivo in response to CXCL12. VAL cells expressing full-length ACKR3, but not a truncated version missing the C-terminus, can support the migration of VAL cells lacking ACKR3 (VAL-ko) when allowed to migrate together. This migration of VAL-ko cells is pertussis toxin-sensitive suggesting the involvement of a Gi-protein coupled receptor. RNAseq analysis indicate the expression of chemotaxis-mediating LTB4 receptors in VAL cells. We found that LTB4 acts synergistically with CXCL12 in stimulating the migration of VAL cells. Pharmacologic or genetic inhibition of BLT1R markedly reduces chemotaxis towards CXCL12 suggesting that LTB4 enhances in a contact-independent manner the migration of lymphoma cells. The results unveil a novel mechanism of cell-to-cell-induced migration of lymphoma.
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Project description:The goal of this study is to analyze differences in gene expression of CD8 T cells that are antigen-educated by lymphatic stromal cells versus traditional antigen-presenting cells. Naive antigen-specific CD8 T cells reactive against the model antigen ovalbumin (OVA) were isolated from spleens of OT-I transgenic mice. They were then co-cultured with OVA-pulsed mature dendritic cells (mDCs) versus lymphatic endothelial cells (LECs) for up to 3d. Each day, OT-I cells were isolated and prepared for RNAseq analysis to determine differences in gene expression as a function of antigen-presenting cell. Naive, LEC-educated, or DC-educated OT-I transcriptomes are provided here. Grant ID - AdG-323053 Agency - European Research Council (ERC) Grant Title - LYMPHIMMUNE - Flow in the tumor microenvironment: Linking mechanobiology with immunology Grant ID - R01CA219304 Agency - US National Institutes of Health (NIH) Grant Title - Paradoxical roles of tumor lymphangiogenesis on tumor immunity and implications for immunotherapy
2017-12-31 | GSE88830 | GEO
Project description:Nathaniel L. Coggins, Danielle Trakimas, S. Laura Chang, Anna Ehrlich, Paramita Ray, Kathryn E. Luker, Jennifer J. Linderman & Gary D. Luker. CXCR7 controls competition for recruitment of β-arrestin 2 in cells expressing both CXCR4 and CXCR7. PLoS ONE 9, 6 (2014).
Chemokine CXCL12 promotes growth and metastasis of more than 20 different human cancers, as well as pathogenesis of other common diseases. CXCL12 binds two different receptors, CXCR4 and CXCR7, both of which recruit and signal through the cytosolic adapter protein β-arrestin 2. Differences in CXCL12-dependent recruitment of β-arrestin 2 in cells expressing one or both receptors remain poorly defined. To quantitatively investigate parameters controlling association of β-arrestin 2 with CXCR4 or CXCR7 in cells co-expressing both receptors, we used a systems biology approach combining real-time, multi-spectral luciferase complementation imaging with computational modeling. Cells expressing only CXCR4 maintain low basal association with β-arrestin 2, and CXCL12 induces a rapid, transient increase in this interaction. In contrast, cells expressing only CXCR7 have higher basal association with β-arrestin 2 and exhibit more gradual, prolonged recruitment of β-arrestin 2 in response to CXCL12. We developed and fit a data-driven computational model for association of either CXCR4 or CXCR7 with β-arrestin 2 in cells expressing only one type of receptor. We then experimentally validated model predictions that co-expression of CXCR4 and CXCR7 on the same cell substantially decreases both the magnitude and duration of CXCL12-regulated recruitment of β-arrestin 2 to CXCR4. Co-expression of both receptors on the same cell only minimally alters recruitment of β-arrestin 2 to CXCR7. In silico experiments also identified β-arrestin 2 as a limiting factor in cells expressing both receptors, establishing that CXCR7 wins the "competition" with CXCR4 for CXCL12 and recruitment of β-arrestin 2. These results reveal how competition for β-arrestin 2 controls integrated responses to CXCL12 in cells expressing both CXCR4 and CXCR7. These results advance understanding of normal and pathologic functions of CXCL12, which is critical for developing effective strategies to target these pathways therapeutically.
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Project description:We characterized high-grade ovarian serous carcinoma TIME based on integrative single-cell transcriptomics analysis of public and in-house datasets. We identified a distinct transcriptomic landscape of both immune and non-immune cells between the dense and more sparse stromal tumors. High stromal tumors contain a lower fraction of infiltrating activated CXCR3+ GRZB+ IFN+ IL2R+ CD8+ T and NCR3+ KLRD1+ natural killer (NK) cells and CXCL1+ macrophages. Next, we determined the potential interplays between non-immune and immune cells. High stromal tumors showed increased expression of CXCL12 in epithelial cancer cells and CA-MSCs. Cell-cell communication analyses suggested that epithelial cancer cells and CA-MSCs secreted CXCL12 interacts with the CXCR4 receptor on NK and CD8+ T cells. Using CXCL12 and/or CXCR4 neutralizing antibodies, we confirmed the immunosuppressive role of the CXCL12-CXCR4 axis in CA-MSC-induced immune suppression.
2024-01-07 | GSE232314 | GEO
Project description:An autochthonous model of pancreatic ductal adenocarcinoma (PDA) permitted the analysis of why immunotherapy is ineffective in this human disease. Despite finding that PDA-bearing mice had cancer cell-specific CD8+ T cells, the mice, like human PDA patients, did not respond to two immunological checkpoint antagonists that promote the function of T cells, α-CTLA-4 and α-PD-L1. Immune control of PDA growth was achieved, however, by depleting carcinoma-associated fibroblasts (CAFs) that express Fibroblast Activation Protein (FAP). The depletion of the FAP+ stromal cell also uncovered the anti-tumor effects of α-CTLA-4 and α-PD-L1, indicating that its immune suppressive activity accounts for the failure of these T cell checkpoint antagonists. Three findings suggested that CXCL12 explained the overriding immunosuppression by the FAP+ cell: T cells were absent from regions of the tumor containing cancer cells; cancer cells were coated with the chemokine, CXCL12; and the FAP+ CAF was the principle source of CXCL12 in the tumor. Administering AMD3100, a CXCL12 receptor (CXCR4) inhibitor, induced rapid T cell accumulation among cancer cells, and acted synergistically with α-PD-L1 to selectively and greatly diminish cancer cells, identified by their loss-of-heterozygosity (LOH) of Trp53. The residual tumor was comprised only of pre-malignant epithelial cells and inflammatory cells. Thus, a single protein, CXCL12, from a single stromal cell type, the FAP+ CAF, may direct tumor immune evasion in a model of human PDA.
2012-11-30 | GSE42605 | GEO
Project description:During inflammation and tissue regeneration, the alarmin High Mobility Group Box 1 (HMGB1), in its reduced isoform, enhances the activity of the chemokine CXCL12, forming a heterocomplex that acts via the chemokine receptor CXCR4. Despite the established roles of both HMGB1 and CXCL12 in tumor progression and metastatic spread to distal sites, the role of the CXCL12/HMGB1 heterocomplex in cancer has never been investigated. By employing a newly established mass spectrometry protocol that allows an unambiguous distinction between reduced (red HMGB1) and oxidized (ox HMGB1) HMGB1 isoforms in cell lysates, we demonstrate that human epithelial cells derived from breast (MCF-7 and MDA-MB-231) and prostate (PC-3) cancer predominantly express red-HMGB1, while primary CD3+ T lymphocytes from peripheral blood express both HMGB1 isoforms. All these cancer cells release HMGB1 in the extracellular microenvironment together with varying concentrations of thioredoxin and thioredoxin reductase. The CXCL12/HMGB1 heterocomplex enhances, via CXCR4, the directional migration and invasiveness of cancer cells characterized by high metastatic potential that possess a fully active thioredoxin system, contributing to maintain red-HMGB1. On the contrary, cancer cells with low metastatic potential, lack thioredoxin reductase, promptly uptake CXCL12 and fail to respond to the heterocomplex. Our study demonstrates that the responsiveness of cancer cells to the CXCL12/HMGB1 heterocomplex, resulting in enhanced cell migration and invasiveness, depends on the maintenance of HMGB1 in its reduced isoform, and suggests disruption of the heterocomplex as a potential therapeutic target to inhibit invasion and metastatic spread in cancer therapies.
2024-05-02 | PXD048091 | Pride
Project description:Tumor escape mechanisms for immunotherapy include deficiencies in antigen presentation, diminishing adaptive CD8+ T cell antitumor activity. Although innate NK cells are triggered by loss of MHC class I, their response is often inadequate. To increase tumor susceptibility to both innate and adaptive immune elimination, we performed parallel genome-wide CRISPR-Cas9 screens. This identified all components, RNF31, RBCK1, and SHARPIN, of the linear ubiquitination chain assembly complex (LUBAC). Genetic and pharmacologic ablation of RNF31, an E3 ubiquitin ligase, strongly sensitized melanoma, breast and colorectal cancer cells to both NK and CD8+ T cell killing. This occurred in a TNF-dependent manner, causing loss of A20 and non-canonical IKK complexes from TNF Receptor Complex I. Corroborating this preclinically, a small molecule RNF31 inhibitor sensitized human colon carcinoma organoids to TNF and greatly enhanced immune bystander killing of antigen-loss and antigen presentation machinery-deficient tumor cells. These results merit exploration of RNF31 inhibition as a clinical pharmacological opportunity for immunotherapy-refractory cancers.
2022-05-30 | PXD029489 | Pride
Project description:The chemokine CXCL12 and its receptor CXCR4 play important roles in signaling and migration of T-cells, but little is known about the transcriptional events involved in CXCL12-mediated T-cell migration. In this study we performed microarray analysis on CXCL12- treated T-cells, and found that the Wnt family of proteins was significantly upregulated during CXCL12 treatment. Confirmation of these results by real-time PCR and Western analysis indicated that the non-canonical Wnt pathway was specifically upregulated during CXCL12 treatment. In vitro and in vivo knockdown studies confirm that b-catenin (the key mediator of canonical Wnt signaling) is not involved in the CXCL12-mediated migration of T-cells. However, Wnt5A, a non-canonical Wnt protein, increases signaling through the CXCL12/ CXCR4 axis via Protein Kinase C (PKC). Our results demonstrated that CXCL12 required Wnt5A to mediate T-cell migration, and the treatment of T-cells with recombinant Wnt5A sensitized T-cells to CXCL12 induced migration. Additionally, Wnt5A expression was required for the sustained expression of CXCR4, both transcriptionally and translationally. These results could be translated in vivo, using EL4 thymoma metastasis as a model of T-cell migration. Taken together our data indicate, for the first time, that Wnt5A is a critical mediator of the CXCL12/ CXCR4 signaling axis. Keywords: Wnt5A, CXCL12, CXCL12, CXCR4, T-cell Migration
2009-01-01 | GSE12449 | GEO
Project description:Neutrophil homeostasis is maintained, in part, by the regulated release of neutrophils from the bone marrow. Constitutive expression of the chemokine CXCL12 by bone marrow stromal cells provides a key retention signal for neutrophils in the bone marrow through activation of its receptor CXCR4. Herein, we show that the ELR chemokines CXCL1 and CXCL2 are constitutively expressed by bone marrow endothelial cells and osteoblasts, and CXCL2 expression is induced in endothelial cells during granulocyte colony-stimulating factor (G-CSF)-induced neutrophil mobilization. Neutrophils lacking CXCR2, the receptor for CXCL1 and CXCL2, are preferentially retained in the bone marrow, reproducing a myelokathexis phenotype. Transient disruption of CXCR4 failed to mobilize CXCR2 neutrophils. However, doubly deficient neutrophils (CXCR2-/- CXCR4-/-) displayed constitutive mobilization, showing that CXCR4 plays a dominant role. Collectively, these data suggest that CXCR2 signaling is a second chemokine axis that interacts antagonistically with CXCR4 to regulate neutrophil release from the bone marrow. We used gene expression microarrays to determine the changes in osteoblasts and bone marrow endothelial cells after G-CSF treatment. 3 untreated and G-CSF-treated osteoblast samples and 4 untreated and G-CSF-treated endothelial samples.
2010-07-14 | E-GEOD-22925 | biostudies-arrayexpress