Expression profiling of BRAFV600E melanoma cell lines upon RAF inhibition
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ABSTRACT:
ORGANISM(S): Homo sapiens
SUBMITTER: Joseph EW
PROVIDER: S-ECPF-GEOD-20051 | biostudies-other | 2010 Aug
REPOSITORIES: biostudies-other
Similar Datasets
Project description:Microarray expression analysis to identify global changes in transcription in response to RAF inhibition. Genes under RAF control were identified in a panel of BRAFV600E tumor cells, following the short-term inhibition of RAF using a pan-RAF kinase inhibitor, PLX4032 (Plexxikon). For comparison with changes in gene expression in response to MEK inhibition using PD0325901 (Pfizer), the following array data was referenced: (http://www.ncbi.nlm.nih.gov/geo/ (accession no. GSE10086)).
2010-07-18 | GSE20051 | GEO
Project description:Microarray expression analysis to identify global changes in transcription in response to RAF inhibition. Genes under RAF control were identified in a panel of BRAFV600E tumor cells, following the short-term inhibition of RAF using a pan-RAF kinase inhibitor, PLX4032 (Plexxikon). For comparison with changes in gene expression in response to MEK inhibition using PD0325901 (Pfizer), the following array data was referenced: (http://www.ncbi.nlm.nih.gov/geo/ (accession no. GSE10086)). Cell lines growing in culture (n=5) were treated with the RAF inhibitor PLX4032 (250nM or 1000nM) or vehicle alone (0.1% DMSO) as control, for eight hours.
2010-07-18 | E-GEOD-20051 | biostudies-arrayexpress
Project description:Blocking oncogenic signaling induced by the BRAFV600E mutation is a promising approach for melanoma treatment. We tested the anti-tumor effects of a specific inhibitor of Raf protein kinases, PLX4032/RG7204, in melanoma cell lines. PLX4032 decreased signaling through the MAPK pathway only in cell lines with the BRAFV600E mutation. Seven out of 10 BRAFV600E mutant cell lines displayed sensitivity based on cell viability assays and three were resistant at concentrations up to 10 muM. Among the sensitive cell lines, four were highly sensitive with IC50 values below 1 muM, and three were moderately sensitive with IC50 values between 1 and 10 muM. There was evidence of MAPK pathway inhibition and cell cycle arrest in both sensitive and resistant cell lines. Genomic analysis by sequencing, genotyping of close to 400 oncogeninc mutations by mass spectrometry, and SNP arrays demonstrated no major differences in BRAF locus amplification or in other oncogenic events between sensitive and resistant cell lines. However, metabolic tracer uptake studies demonstrated that sensitive cell lines had a more profound inhibition of FDG uptake upon exposure to PLX4032 than resistant cell lines. In conclusion, BRAFV600E mutant melanoma cell lines displayed a range of sensitivities to PLX4032 and metabolic imaging using PET probes can be used to assess sensitivity.
| S-EPMC2876068 | biostudies-literature
Project description:Oncogenic mutations in BRAF are common in melanoma and thyroid carcinoma and drive constitutive activation of the MAPK pathway. Molecularly targeted therapies of this pathway improves survival compared to chemotherapy; however, responses tend to be short-lived as resistance invariably occursCell line models of melanoma and thyroid carcinoma, +/- BRAF(V600E) activating mutation, were treated with the MEK inhibitor PD0325901. Treated and naive samples were assayed for expression of key members of the MAPK pathway. Global microRNA expression profiling of naive and resistant cells was performed via next generation sequencingand indicated pluripotency pathways in resistance. Parental cell lines were progressed to holoclones to confirm the miRNA stemness profileMembers of the MIR302/373/374/520 family of embryonic stem cell specific cell cycle regulating (ESCC) microRNAs were identified as differentially expressed between resistant BRAF(V600E) melanoma and thyroid cell lines. Upregulated expression of gene and protein stemness markers, upregulated expression of MAPK pathway genes and downregulation of the ESCC MIR302 cluster in BRAF(V600E) melanoma indicated an increased stem-like phenotype in resistant BRAF(V600E) melanoma. Conversely, downregulated expression of gene and protein stemness markers, downregulated expression of MAPK pathway genes, upregulation of the ESCC MIR520 cluster, reeexpression of cell surface receptors, and induced differentiation-associated morphology in resistant BRAF(V600E) indicate a differentiated phenotype associated with MEK inhibitor resistance in BRAF(V600E) thyroid cellsThe differential patterns of resistance observed between BRAF(V600E) melanoma and thyroid cell lines may reflect tissue type or de novo differentiation, but could have significant impact on the response of primary and metastatic cells to MEK inhibitor treatment. This study provides a basis for the investigation of the cellular differentiation/self-renewal access and its role in resistance to MEK inhibition.
| S-EPMC4910922 | biostudies-literature
Project description:Cross-feedback activation of MAPK and AKT pathways is implicated as a resistance mechanism for cancer therapeutic agents targeting either RAF/MEK or PI3K/AKT/mTOR. It is thus important to have a better understanding of the molecular resistance mechanisms to improve patient survival benefit from these agents. Here we show that BRAFV600E is a negative regulator of the AKT pathway. Expression of BRAFV600E in NIH3T3 cells significantly suppresses MEK inhibitor (RG7167) or mTORC1 inhibitor (rapamycin) induced AKT phosphorylation (pAKT) and downstream signal activation. Treatment-induced pAKT elevation is found in BRAF wild type melanoma cells but not in a subset of melanoma cell lines harboring BRAFV600E. Knock-down of BRAFV600E in these melanoma cells elevates basal pAKT and downstream signals, whereas knock-down of CRAF, MEK1/2 or ERK1/2 or treatment with a BRAF inhibitor have no impact on pAKT. Mechanistically, we show that BRAFV600E interacts with rictor complex (mTORC2) and regulates pAKT through mTORC2. BRAFV600E is identified in mTORC2 after immunoprecipitation of rictor. Knock-down of rictor abrogates BRAFV600E depletion induced pAKT. Knock-down of BRAFV600E enhances cellular enzyme activity of mTORC2. Aberrant activation of AKT pathway by PTEN loss appears to override the negative impact of BRAFV600E on pAKT. Taken together, our findings suggest that in a subset of BRAFV600E melanoma cells, BRAFV600E negatively regulates AKT pathway in a rictor-dependent, MEK/ERK and BRAF kinase-independent manner. Our study reveals a novel molecular mechanism underlying the regulation of feedback loops between the MAPK and AKT pathways.
| S-EPMC3411810 | biostudies-literature
Project description:Cancer cell differentiation is an important field of discussion in the light of cancer stem cells. In a recent study by Herr et al. (2015) "B-RAF inhibitors induce epithelial differentiation in BRAF-mutant colorectal cancer cells" we described how inhibition of mutant BRAF in colorectal cancer cell lines induces cell re-differentiation that is correlated with the loss of tumor growth in vitro and in vivo. We used Illumina HumanHT-12 v4 Expression BeadChip to characterize the gain of differentiation of PLX4720-treated 3D cultures of HT29 and Colo-205 cells. Here, we describe the experimental design and statistical analysis that were performed on the data set leading to the above hypothesis. The data are publicly available at the Gene Expression Omnibus (GEO) database under the accession number GSE50791.
| S-EPMC4535967 | biostudies-literature
Project description:Cerebral metastases occur in a majority of metastatic melanoma patients and are a major cause of mortality. Despite this, there is a poor understanding of the molecules/pathways that lead to the brain-metastatic phenotype. Studies designed to address this deficiency and test novel therapeutic approaches have until recently been slowed by an absence of preclinical models of spontaneous CNS metastatic melanoma disease. To address this, we isolated two variants of the human melanoma cell line WM239 (named 131/4-5B1 and 131/4-5B2) which can metastasize spontaneously to brain parenchyma from an orthotopic primary transplant. We have performed gene expression profiling on both brain metastatic cell lines (131/4-5B1 and 131/4-5B2) and compared to the poorly metastatic parental cell line WM239A and a derived highly metastatic variant 113/6-4L in order to examine the mechanisms that influence the progression of malignant melanoma to a brain-metastatic phenotype. Two-condition experiment, brain metastatic cell lines (131/4-5B1 and 131/4-5B2) and compared to the poorly metastatic parental cell line WM239A and a derived highly metastatic variant 113/6-4L. Biological replicates: 4 independently grown and harvested cell line passages. Two technical replicate per condition (including dye swap).
| S-ECPF-GEOD-23601 | biostudies-other
Project description:A detailed understanding of the mechanisms by which tumors acquire resistance to targeted anticancer agents should speed the development of treatment strategies with lasting clinical efficacy. RAF inhibition in BRAF-mutant melanoma exemplifies the promise and challenge of many targeted drugs; although response rates are high, resistance invariably develops. Here, we articulate overarching principles of resistance to kinase inhibitors, as well as a translational approach to characterize resistance in the clinical setting through tumor mutation profiling. As a proof of principle, we performed targeted, massively parallel sequencing of 138 cancer genes in a tumor obtained from a patient with melanoma who developed resistance to PLX4032 after an initial dramatic response. The resulting profile identified an activating mutation at codon 121 in the downstream kinase MEK1 that was absent in the corresponding pretreatment tumor. The MEK1(C121S) mutation was shown to increase kinase activity and confer robust resistance to both RAF and MEK inhibition in vitro. Thus, MEK1(C121S) or functionally similar mutations are predicted to confer resistance to combined MEK/RAF inhibition. These results provide an instructive framework for assessing mechanisms of acquired resistance to kinase inhibition and illustrate the use of emerging technologies in a manner that may accelerate personalized cancer medicine.
| S-EPMC3157968 | biostudies-literature
Project description:BRAF-inhibitor (BRAFi)-resistance compromises long term survivorship of malignant melanoma patients, and mutant NRAS is a major mediator of BRAFi-resistance. We have employed NanoString nCounterTM transcriptomic analysis of isogenic human malignant melanoma cells that differ only by NRAS mutational status (BRAFi-sensitive A375-BRAFV600E/NRASQ61 versus BRAFi-resistant A375-BRAFV600E/NRASQ61K), identifying modulation of specific gene expression networks as a function of NRASQ61K-status.
2021-09-23 | GSE184633 | GEO
Project description:BackgroundGene expression profiling has revolutionized our ability to molecularly classify primary human tumors and significantly enhanced the development of novel tumor markers and therapies; however, progress in the diagnosis and treatment of melanoma over the past 3 decades has been limited, and there is currently no approved therapy that significantly extends lifespan in patients with advanced disease. Profiling studies of melanoma to date have been inconsistent due to the heterogeneous nature of this malignancy and the limited availability of informative tissue specimens from early stages of disease.Methodology/principle findingsIn order to gain an improved understanding of the molecular basis of melanoma progression, we have compared gene expression profiles from a series of melanoma cell lines representing discrete stages of malignant progression that recapitulate critical characteristics of the primary lesions from which they were derived. Here we describe the unsupervised hierarchical clustering of profiling data from melanoma cell lines and melanocytes. This clustering identifies two distinctive molecular subclasses of melanoma segregating aggressive metastatic tumor cell lines from less-aggressive primary tumor cell lines. Further analysis of expression signatures associated with melanoma progression using functional annotations categorized these transcripts into three classes of genes: 1) Upregulation of activators of cell cycle progression, DNA replication and repair (CDCA2, NCAPH, NCAPG, NCAPG2, PBK, NUSAP1, BIRC5, ESCO2, HELLS, MELK, GINS1, GINS4, RAD54L, TYMS, and DHFR), 2) Loss of genes associated with cellular adhesion and melanocyte differentiation (CDH3, CDH1, c-KIT, PAX3, CITED1/MSG-1, TYR, MELANA, MC1R, and OCA2), 3) Upregulation of genes associated with resistance to apoptosis (BIRC5/survivin). While these broad classes of transcripts have previously been implicated in the progression of melanoma and other malignancies, the specific genes identified within each class of transcripts are novel. In addition, the transcription factor NF-KB was specifically identified as being a potential "master regulator" of melanoma invasion since NF-KB binding sites were identified as consistent consensus sequences within promoters of progression-associated genes.Conclusions/significanceWe conclude that tumor cell lines are a valuable resource for the early identification of gene signatures associated with malignant progression in tumors with significant heterogeneity like melanoma. We further conclude that the development of novel data reduction algorithms for analysis of microarray studies is critical to allow for optimized mining of important, clinically-relevant datasets. It is expected that subsequent validation studies in primary human tissues using such an approach will lead to more rapid translation of such studies to the identification of novel tumor biomarkers and therapeutic targets.
| S-EPMC1895889 | biostudies-literature