Nuclear receptor coactivator SRC-1 promotes colorectal cancer immune escape by enhancing PD-L1 transcription and protein stability
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ABSTRACT:
ORGANISM(S): Mus musculus
PROVIDER: GSE242552 | GEO | 2024/03/06
REPOSITORIES: GEO
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Project description:Nuclear receptor coactivator SRC-1 promotes colorectal cancer immune escape by enhancing PD-L1 transcription and protein stability
| PRJNA1013927 | ENA
Project description:The rapidly growing family of coregulators of nuclear receptors includes coactivators that promote transcription and corepressors that harbor the opposing function. In recent years, coregulators have begun to emerge as important regulators of metabolic homeostasis, including the p160 Steroid Receptor Coactivator (SRC) family. Members of the SRC family have been ascribed important roles in control of gluconeogenesis in liver and fatty acid oxidation in skeletal muscle. To provide a deeper and more granular understanding of the metabolic impact of SRC family members, we have performed targeted metabolomics analysis of key metabolic byproducts of glucose, fatty acid, and amino acid metabolism in mice with global knockout of SRC-1, SRC-2, or SRC-3. We measured amino acids, acyl carnitines, and organic acids in five tissues with key metabolic functions (liver, heart, skeletal muscle, brain, plasma) isolated from SRC-1, -2, or -3 knockout mice and their wild-type littermates in the fed and fasted conditions, thereby unveiling unique metabolic functions of each SRC coactivator. Overall, we observed entire groups or subgroups of metabolites belonging to discrete metabolic pathways that were influenced in different tissues and/or feeding states due to ablation of individual SRC isoforms. Surprisingly, we identified very few metabolites that changed universally across the three SRC knockout models. These findings demonstrate that coactivator function has very limited redundancy even within the homologous SRC coactivator family. Furthermore, this work also demonstrates the use of metabolomics as a means for identifying novel metabolic regulatory functions of transcriptional coregulators.
2013-04-10 | MTBLS30 | MetaboLights
Project description:Primary irradiated MC38 tumors and secondary MC38 non-irradiated tumors. Subcutaneous MC38 tumors in C57BL/6 mice Concurrent treatment with anti-PD-L1.
2023-10-15 | E-MTAB-13389 | biostudies-arrayexpress
Project description:A major component of the cardiac stress response is the simultaneous activation of several gene regulatory networks. Interestingly, the transcriptional regulator steroid receptor coactivator-2, SRC-2 is often decreased during cardiac failure in humans. We postulated that SRC-2 suppression plays a mechanistic role in the stress response and that SRC-2 activity is an important regulator of the adult heart gene expression profile. Genome-wide microarray analysis, confirmed with targeted gene expression analyses revealed that genetic ablation of SRC-2 activates the “fetal gene program” in adult mice as manifested by shifts in expression of a) metabolic and b) sarcomeric genes, as well as associated modulating transcription factors. While these gene expression changes were not accompanied by changes in left ventricular weight or cardiac function, imposition of transverse aortic constriction (TAC) predisposed SRC-2 knockout (KO) mice to stress-induced cardiac dysfunction. In addition, SRC-2 KO mice lacked the normal ventricular hypertrophic response as indicated through heart weight, left ventricular wall thickness, and blunted molecular signaling known to activate hypertrophy. Our results indicate that SRC-2 is involved in maintenance of the steady-state adult heart transcriptional profile, with its ablation inducing transcriptional changes that mimic a stressed heart. These results further suggest that SRC-2 deletion interferes with the timing and integration needed to respond efficiently to stress through disruption of metabolic and sarcomeric gene expression and hypertrophic signaling, the three key stress responsive pathways.
2012-10-13 | GSE41558 | GEO
Project description:A major component of the cardiac stress response is the simultaneous activation of several gene regulatory networks. Interestingly, the transcriptional regulator steroid receptor coactivator-2, SRC-2 is often decreased during cardiac failure in humans. We postulated that SRC-2 suppression plays a mechanistic role in the stress response and that SRC-2 activity is an important regulator of the adult heart gene expression profile. Genome-wide microarray analysis, confirmed with targeted gene expression analyses revealed that genetic ablation of SRC-2 activates the M-bM-^@M-^\fetal gene programM-bM-^@M-^] in adult mice as manifested by shifts in expression of a) metabolic and b) sarcomeric genes, as well as associated modulating transcription factors. While these gene expression changes were not accompanied by changes in left ventricular weight or cardiac function, imposition of transverse aortic constriction (TAC) predisposed SRC-2 knockout (KO) mice to stress-induced cardiac dysfunction. In addition, SRC-2 KO mice lacked the normal ventricular hypertrophic response as indicated through heart weight, left ventricular wall thickness, and blunted molecular signaling known to activate hypertrophy. Our results indicate that SRC-2 is involved in maintenance of the steady-state adult heart transcriptional profile, with its ablation inducing transcriptional changes that mimic a stressed heart. These results further suggest that SRC-2 deletion interferes with the timing and integration needed to respond efficiently to stress through disruption of metabolic and sarcomeric gene expression and hypertrophic signaling, the three key stress responsive pathways. For microarray analysis, 250ng of RNA isolated from total heart (RNeasy kit, Qiagen) for each sample was labeled using the new standard Affymetrix linear amplification protocol using the 3' IVT Express Kit. This was reverse-transcribed and cRNA was produced and biotinylated via in vitro transcription. A hybridization cocktail containing Affymetrix spike-in controls and 15 M-NM-<g fragmented, labeled cRNA was loaded onto a GeneChipM-BM-. Mouse 430 2.0 array. The array was hybridized for 16 hours at 45M-BM-0C with rotation at 60 rpm then washed and stained with a strepavidin, R-phycoerythrin conjugate stain using the FS 450_0001 Fluidics protocol setting. Signal amplification was done using biotinylated antistreptavidin. The stained array was scanned on the Affymetrix GeneChipM-BM-. Scanner 3000. The images were analyzed and quality control metrics recorded using Affymetrix Command Console v3. Experiments were run using Affymetrix MG 430 2.0 chip with 45,101 probesets representing 20,757 unique genes. There were 8 experiments in 2 groups: WT-unstressed M-bM-^@M-^S 4 experiments, and KO-unstressed M-bM-^@M-^S 4 experiments. QC parameters for all experiments were within the acceptable limits. We used the following software packages for data QC, statistical analysis and presentation of the results: Affymetrix Expression Console (www.affymetrix.com), Partek (www.partek.com), BRB Array Tools (linus.nci.nih.gov/BRB-ArrayTools.html), and dChip (biosun1.harvard.edu/complab/dchip). Expressions were estimated using the RMA (Multi-Array Analysis) method [38] with Partek software. Differentially expressed genes were found using the RVM (Random Variance Model) t-test, which is designed for small sample size experiments [39]. We used BRB Array Tools software, developed by Dr. Richard Simon and the BRB-ArrayTools Development Team. All genes were included in the comparison. For the genes represented by more than one probeset, we used the most highly expressed probeset. The cutoffs for differentially expressed genes were False Discovery Rate (FDR) = 0.05 [40].
2012-10-13 | E-GEOD-41558 | biostudies-arrayexpress
Project description:This experiment series addresses the role of coactivator SRC-1/NcoA-1 for the induction of interleukin-6 (IL-6) target genes in HepG2 cells. For that purpose, HepG2 human hepatocellular carcinoma cells were manipulated to stably express an shRNA that knocks down SRC-1 expression yielding the HepG2-âSrc1 cells. Either unmanipulated HepG2 or HepG2-âSrc1 cells were then treated for various periods with IL-6. Experiment Overall Design: HepG2 or HepG2-âSrc1 (with silenced SRC-1/NcoA-1 expression) cells were either left untreated or treated with 10 ng/ml IL-6 for 1 or 4 hours. Every experiment was carried out in replicate.
2008-06-13 | E-GEOD-4885 | biostudies-arrayexpress
Project description:This experiment series addresses the role of coactivator SRC-1/NcoA-1 for the induction of interleukin-6 (IL-6) target genes in HepG2 cells. For that purpose, HepG2 human hepatocellular carcinoma cells were manipulated to stably express an shRNA that knocks down SRC-1 expression yielding the HepG2-∆Src1 cells. Either unmanipulated HepG2 or HepG2-∆Src1 cells were then treated for various periods with IL-6. Keywords: time course, genetic modification
2006-12-31 | GSE4885 | GEO
Project description:BackgroundIn patients with microsatellite stable (MSS) metastatic colorectal cancer (mCRC), immune checkpoint blockade is ineffective, and combinatorial approaches enhancing immunogenicity need exploration.MethodsWe treated 43 patients with predominantly microsatellite stable RAS/BRAF wild-type mCRC on a phase II trial combining chemotherapy with the epidermal growth factor receptor antibody cetuximab and the programmed cell death ligand 1 (PD-L1) antibody avelumab. We performed next-generation gene panel sequencing for mutational typing of tumors and liquid biopsy monitoring as well as digital droplet PCR to confirm individual mutations. Translational analyses included tissue immunohistochemistry, multispectral imaging and repertoire sequencing of tumor-infiltrating T cells. Detected PD-L1 mutations were mechanistically validated in CRISPR/Cas9-generated cell models using qRT-PCR, immunoblotting, flow cytometry, complement-dependent cytotoxicity assay, antibody-dependent cytotoxicity by natural killer cell degranulation assay and LDH release assay as well as live cell imaging of T cell mediated tumor cell killing.ResultsCirculating tumor DNA showed rapid clearance in the majority of patients mirroring a high rate of early tumor shrinkage. In 3 of 13 patients expressing the high-affinity Fcγ receptor 3a (FcγR3a), tumor subclones with PD-L1 mutations were selected that led to loss of tumor PD-L1 by nonsense-mediated RNA decay in PD-L1 K162fs and protein degradation in PD-L1 L88S. As a consequence, avelumab binding and antibody-dependent cytotoxicity were impaired, while T cell killing of these variant clones was increased. Interestingly, PD-L1 mutant subclones showed slow selection dynamics reversing on avelumab withdrawal and patients with such subclones had above-average treatment benefit. This suggested that the PD-L1 mutations mediated resistance to direct antitumor effects of avelumab, while at the same time loss of PD-L1 reduced biological fitness by enhanced T cell killing limiting subclonal expansion.ConclusionThe addition of avelumab to standard treatment appeared feasible and safe. PD-L1 mutations mediate subclonal immune escape to avelumab in some patients with mCRC expressing high-affinity FcγR3a, which may be a subset experiencing most selective pressure. Future trials evaluating the addition of avelumab to standard treatment in MSS mCRC are warranted especially in this patient subpopulation.Trial registration numberNCT03174405.
| S-EPMC8317124 | biostudies-literature
Project description:BackgroundLong non-coding RNAs (LncRNAs) have been implicated as critical regulators of cancer tumorigenesis and progression. However, their functions and molecular mechanisms in colorectal cancer (CRC) still remain to be further elucidated.MethodsLINC00460 was identified by differential analysis between human CRC and normal tissues and verified by in situ hybridization (ISH) and qRT-PCR. We investigated the biological functions of LINC00460 in CRC by in vitro and in vivo experiments. We predicted the mechanism and downstream functional molecules of LINC00460 by bioinformatics analysis, and confirmed them by dual luciferase reporter gene assay, RNA immunoprecipitation (RIP), RNA pull-down, etc. RESULTS: LINC00460 was found to be significantly overexpressed in CRC and associated with poor prognosis. Overexpression of LINC00460 promoted CRC cell immune escape and remodeled a suppressive tumor immune microenvironment, thereby promoting CRC proliferation and metastasis. Mechanistic studies showed that LINC00460 served as a molecular sponge for miR-186-3p, and then promoted the expressions of MYC, CD47 and PD-L1 to facilitate CRC cell immune escape. We also demonstrated that MYC upregulated LINC00460 expression at the transcriptional level and formed a positive feedback loop.ConclusionsThe LINC00460/miR-186-3p/MYC feedback loop promotes CRC cell immune escape and subsequently facilitates CRC proliferation and metastasis. Our findings provide novel insight into LINC00460 as a CRC immune regulator, and provide a potential therapeutic target for CRC patients.
| S-EPMC11321182 | biostudies-literature
Project description:The two most common antibody targeting principles in oncology are the induction of direct antitumor effects and the release of antitumor T cell immunity by immune checkpoint blockade. These two principles, however, may be overlapping if the targeted checkpoint molecule is not located on the immune cell but on the tumor cell itself. Secondary resistance by epitope escape may therefore remain a challenge in both settings. We previously reported epitope escape through L88S and truncating programmed cell death ligand 1 (PD-L1) gene mutations in colorectal cancer patients on selective pressure with avelumab, a PD-L1-directed checkpoint blocker that-in addition to T cell disinhibition-allows direct tumor cell killing via its unmodified Fc portion. Here, we confirmed this principle by liquid biopsy monitoring in a colorectal cancer patient from an independent clinical trial. In this patient, both PD-L1 L88E and L88fs mutations emerged under selective pressure with avelumab. By ectopically expressing PD-L1 L88E, we show that this mutation leads to a reduction of full-length glycosylated PD-L1 and greatly reduced avelumab surface binding. Further experiments indicated that PD-L1 L88E represents a phosphomimetic variant of PD-L1 L88S leading to loss of protein stability and increased proteasomal degradation. The association of this PD-L1 mutation with the high-affinity FCGR3A single nucleotide polymorphism rs396991 confirms prior evidence that patients harboring this polymorphism experience the strongest selective pressure by avelumab. Together, position 88 of PD-L1 is a hotspot residue critically regulating PD-L1 cell surface expression with clinical significance in the context of immune checkpoint blockade.
| S-EPMC9353709 | biostudies-literature