Single-cell transcriptomics of NRAS-mutated melanoma transitioning to drug resistance: elevated P2RX7 levels indicate response to targeted therapy
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ABSTRACT:
PROVIDER: PRJNA961723 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR24302000_1.fastq.gz | Fastqsanger.gz | |||
| SRR24302000_2.fastq.gz | Fastqsanger.gz | |||
| SRR24302001_1.fastq.gz | Fastqsanger.gz | |||
| SRR24302001_2.fastq.gz | Fastqsanger.gz | |||
| SRR24302002_1.fastq.gz | Fastqsanger.gz |
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Project description:While effective therapy is available for melanoma patients harbouring BRAF mutations, to date, the efficient therapeutic approaches for NRAS-mutant melanoma patients are limited. Therapeutic regimen involving combinatorial inhibition of MEK1/2 and CDK4/6 prove to be beneficial as targeted therapy for NRAS-mutant melanoma. Yet, only a subset of melanoma patients responded to this treatment, whereas acquired resistance eventually emerge in responders. Upon prolonged therapy with MEK1/2 and CDK4/6 inhibitors cells switch to fully drug-resistant or senescent phenotype in vitro. Consequently, there is a necessity to portray cell populations sensitive to targeted therapy, but also cell transitions associated with NRAS-mutant melanoma resistance. To define transcriptional states arising under indicated therapy and cell fate decisions governing resistance, we performed single-cell RNA sequencing at four distinguished time points during treatment. In this study, we identified a cell population highly enriched in calcium signalling, among others, associated with positive drug response. In particular, prolonged expression of ATP-gated ion channel P2RX7 was related to a senescent-like phenotype. Next, we described an immune-like melanoma state that acquired drug resistance and re-enter the cell cycle under treatment, as well as cell populations with intrinsic potential to evade drug pressure.
2023-04-26 | GSE230538 | GEO
Project description:Ten-eleven translocation (Tet) family-mediated DNA oxidation represents a novel epigenetic modification capable of converting 5-methylcytosine (5-mC) to 5-hydroxymethylcytosine (5-hmC) to regulate various biological processes. However, it is unknown whether the Tet family affects mesenchymal stem cells (MSCs) or the skeletal system. Here we show that depletion of Tet1 and Tet2 resulted in impaired self-renewal and differentiation of bone marrow MSCs (BMMSCs) and a significant osteopenia phenotype. Mechanistically, Tet1 and Tet2 deficiency reduced demethylation of the P2rX7 promoter and thus downregulated exosome release, leading to intracellular accumulation of miR-297a-5p, miR-297b-5p, and miR-297c-5p. These miRNAs inhibited Runx2 signaling to impair BMMSC function. We show that overexpression of P2rX7 consistently rescued the impaired BMMSCs and osteoporotic phenotype in Tet1 and Tet2 double knockout mice. These results indicate that Tet1 and Tet2 play a critical role in maintaining BMMSC and bone homeostasis through epigenetic regulation of P2rX7 to control exosome and miRNA release. This newly identified Tet/P2rX7/Runx2 cascade may serve as a target for the development of novel therapies for osteopenia disorders.
2018-04-11 | GSE108872 | GEO
Project description:Extracellular adenosine triphosphate (eATP) is a signaling molecule that affects T cell function via the ionotropic P2X7 receptor. The study of effector/memory T cells isolated from mice with deletion of P2rx7, the gene encoding for P2X7, allowed understanding the impact of P2X7 activity on T cell function in the eATP-rich tumor microenvironment. To explore the the transcriptional impact of the lack of P2rx7 in CD4+ naïve and TEM cells, we performed genome-wide expression profiling of ex vivo purified CD4+ naïve and TEM cells from WT and P2rx7-/- mice
2020-07-24 | GSE118146 | GEO
Project description:Background: Ductal carcinoma in situ (DCIS) is the most common type of in situ premalignant breast cancers. What drives DCIS to invasive breast cancer is unclear. Basal-like invasive breast cancers are aggressive. We have previously shown that N-Ras is highly expressed selectively in basal-like subtypes of invasive breast cancers and can promote their growth and progression. In this study, we investigated whether NRAS expression at the DCIS stage can control transition from luminal DCIS to basal-like invasive breast cancers. Methods: Wilcoxon rank-sum test was performed to assess expression of NRAS in DCIS compared to invasive breast tumors in patients. NRAS levels were also determined by fluorescence in situ hybridization in patient tumor microarrays (TMAs) with concurrent normal, DCIS, and invasive breast cancer, and association of NRAS mRNA levels with DCIS and invasive breast cancer was assessed by paired Wilcoxon signed-rank test. Pearson’s correlation was calculated between NRAS mRNA levels and basal biomarkers in the TMAs, as well as in patient datasets. RNA-seq data was generated in cell lines and unsupervised hierarchical clustering was performed after combining with RNA-seq data from a previously published patient cohort. Results: Invasive breast cancers showed higher NRAS mRNA levels compared to DCIS samples. These NRAShigh lesions were also enriched with basal-like features, such as basal gene expression signatures, lower ER, and higher p53 protein and Ki67 levels. We have shown previously that N-Ras drives aggressive features in DCIS-like and basal-like SUM102PT cells. Here, we found that NRAS-silencing induced a shift to a luminal gene expression pattern. Conversely, NRAS overexpression in the luminal DCIS SUM225 cells induced a basal-like gene expression pattern, as well as an epithelial-to-mesenchymal transition signature. Furthermore, these cells formed disorganized mammospheres containing cell masses with an apparent reduction in adhesion. Conclusions: These data suggest that elevated NRAS levels in DCIS is not only a marker but can also control the emergence of basal-like features leading to more aggressive tumor activity, thus supporting the therapeutic hypothesis that targeting N-Ras and/or downstream pathways may block disease progression for a subset of DCIS patients with high NRAS.
2022-10-18 | GSE215407 | GEO
Project description:Melanoma is often driven by NRAS mutations. Although oncogenic NRAS activates MAPK signaling, inhibition of this pathway has limited efficacy in NRAS-mutant tumors. As there are limited options to treat these tumors, there is a pressing need to identify NRAS-mutant-specific vulnerabilities that can be therapeutically exploited. Metabolic reprogramming is one of the hallmarks of RAS-mutant tumors, creating a dependency on key metabolic pathways. This dependency could be leveraged to combat these tumors. We discovered that abrogation of S6K2, a mTORC1 effector, triggers accumulation of unsaturated fatty acids, lipid peroxidation and cell death selectively in NRAS-mutant melanoma cells that are resistant to MAPK inhibition. S6K2 depletion induced upregulation of S6K1 and PPAR, triggering apoptotic and ferroptosis-like cell death. Conversely, depletion of S6K1 diminished lipid synthesis and cell proliferation, without eliciting cell death. Collectively, our data indicate that a fine balance between S6K1 and S6K2 activity and metabolic homeostasis is required to prompt cell death of NRAS-mutant melanoma. S6K2 is hence a novel therapeutic target for MAPKi-resistant NRAS-mutant melanoma.
2020-03-06 | GSE127916 | GEO
Project description:Duchenne muscular dystrophy (DMD) is a debilitating and typically fatal X-linked progressive neuromuscular disorder that results in progressive muscle degeneration aggravated by sterile inflammation. The P2RX7 purinoceptor is an extracellular ATP-gated ion channel expressed in immune cells, and has been targeted in treatment of infectious and inflammatory diseases. In particular, P2xr7 receptor abnormalities have been demonstrated in mdx dystrophic mice, a model for DMD lacking expression of the full length dystrophin transcript through a single point mutation in exon 23. Here, we looked at the differential effects in gene expression regulation in whole muscle in dystrophic mdx mice with or without ablation of the P2rx7 purinoceptor.
2019-09-26 | E-MTAB-7698 | biostudies-arrayexpress
Project description:Purpose: Emerging evidence in the field of RAS biology is beginning to illustrate that RAS mutations are functionally distinct. To elucidate functional differences between NRAS mutants we performed RNA-sequencing on mouse embryonic fibroblasts (MEFs) expressing different NRAS codon 61 variants from the endogenous gene locus Methods: MEFs were isolated from Tyr::CRE-ERT2 LSL-Nras mouse models containing different Nras variants in the endogeous gene locus. NRAS expression was induced in MEFs using an Adenoviral Cre delivery system. RNA was isolated from MEFs six days post-NRAS induction and submitted for RNA-sequencing Results: Differential expression and gene set enrichment analysis (GSEA) between NRAS variants revealed enrichment of transcripts associated with cellular proliferation between all mutant Nras alleles versus wild-type Nras. Further, MEFs expressing NRAS mutants capable of driving melanoma formation were enriched for transcripts associated with RAS-MYC signaling axis. Conclusion: While all NRAS mutants are capable of enhancing cellular proliferation versus wild-type NRAS, the melanomagenic potential of these mutants correlates with the ability to enhance activation of the RAS-MYC signaling axis
2022-04-30 | GSE162124 | GEO
Project description:The p21 RAS subfamily of small GTPases, including KRAS, HRAS, and NRAS, regulates cell proliferation, cytoskeletal organization and other signaling networks, and is the most frequent target of activating mutations in cancer. Activating germline mutations of KRAS and HRAS cause severe developmental abnormalities leading to Noonan, cardio-facial-cutaneous and Costello syndrome, but activating germline mutations of NRAS have not been reported. Autoimmune lymphoproliferative syndrome (ALPS) is the most common genetic disease of lymphocyte apoptosis and causes autoimmunity as well as excessive lymphocyte accumulation, particularly of CD4-, CD8- ab T cells. Mutations in ALPS typically affect CD95 (Fas/APO-1)-mediated apoptosis, one of the extrinsic death pathways involving tumor necrosis factor receptor (TNFR) superfamily proteins, but certain ALPS individuals have no such mutations. We show here that the salient features of ALPS as well as a predisposition to hematological malignancies can be caused by a heterozygous germline Gly13Asp activating mutation of the NRAS oncogene that does not impair CD95-mediated apoptosis. The increase in active, GTP-bound NRAS augments RAF/MEK/ERK signaling which markedly decreases the pro-apoptotic protein BIM and attenuates intrinsic, nonreceptor-mediated mitochondrial apoptosis. Thus, germline activating mutations in NRAS differ from other p21 Ras oncoproteins by causing selective immune abnormalities without general developmental defects. Our observations on the effects of NRAS activation indicate that RAS-inactivating drugs, such as farnesyl-transferase inhibitors (FTIs) should be examined in human autoimmune and lymphocyte homeostasis disorders. Keywords: NRAS
2008-03-21 | GSE7345 | GEO
Project description:To study the role of NRAS mutations in cell proliferation and self-renewal in acute myeloid leukemia (AML), the human AML cell line, THP1, was modified to replace its naturally occurring heterozygous NRAS-G12D mutation with a doxycycline(dox)-inducible heterozygous NRAS-G12V mutation. The endogenous copies of the NRAS-G12D allele were deleted using CRISPR/Cas9 after a dox-inducible, CRISPR resistant, NRAS-G12V transgene was introduced into the THP1 cell line. The resulting cell line was named B11. RNA-seq data confirmed that endogenous NRAS G12D was successfully replaced by dox-inducible exogenous NRAS G12V in the B11 cell line. As expected, depletion of dox induced G1 cell cycle arrest. Interestingly, the B11 cells experienced ten-times higher expression of NRAS induced G2/S-phase cell cycle arrest. Forty-nine genes were identified as signaling responsible genes associated with high expression of NRAS.
2022-07-25 | GSE115911 | GEO
Project description:Targeted therapies have the potential to revolutionize cancer care by providing personalized treatment strategies that are less toxic and more effective but it is clear that for most solid tumors suppression of a single target is not sufficient to prevent development of resistance. A powerful method to identify mechanisms of resistance and targets for combination therapy is to use an in vivo genetic approach. We have developed a novel retroviral gene delivery mouse model of melanoma that permits control of gene expression post-delivery using the tetracycline (tet)-regulated system. In this study we used this melanoma model to select for resistant tumors following genetic inhibition of mutant NRAS. Analysis of tumors that became resistant to NRAS suppression revealed that the most common mechanism of resistance was overexpression of the Met receptor tyrosine kinase (RTK). Importantly, inhibition of Met overcomes NRAS resistance in this context. Analysis of NRAS mutant human melanoma cells revealed that inhibition of MEK is also associated with adaptive RTK signaling. Furthermore, co-inhibition of RTK signaling and MEK overcomes acquired MEK inhibitor resistance in NRAS mutant melanoma. These data suggest that combined inhibition of RTK and MEK signaling is a rational therapeutic strategy in mutant NRAS driven melanoma. Reversible NRAS Q61R expression in the melanocytes of DCT-TVA;Ink4a/Arf lox/lox mice (FVB/n) was achieved by transducing the animals with Tet-off and TRE-NRASQ61R-IRES-Cre avian leukosis viruses. After tumor initiation, the expression of NRAS Q61R was turned off by administrating doxycycline. Despite initial regression, tumors in 40% of mice developed resistance to NRAS Q61R withdraw. Seven resistant tumors and one control tumor where NRAS Q61R expression was not interrupted were subjected to genome-wide gene expression profiling.
2014-10-31 | E-GEOD-51906 | biostudies-arrayexpress