Homo sapiens
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PROVIDER: PRJNA93301 | ENA |
REPOSITORIES: ENA
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Project description:The diterpene ester PEP005 is a novel anticancer agent that activates PKC and cures subcutaneous murine melanoma by topical application. We now describe the in vitro cytostatic effects of PEP005 and the diterpene ester TPA, observed in 20% of human melanoma cell lines. Primary cultures of normal human neonatal fibroblasts were uniformly resistant to growth arrest, indicating a potential for tumor selectivity. Sensitive cells were induced to senesce and exhibited a G1 and G2/M arrest. There was sustained expression of p21WAF1/CIP1, irreversible dephosphorylation of the retinoblastoma gene product (Rb) and transcriptional silencing of E2F-responsive genes in sensitive cell lines. Activation of MEK1/2 by PKC was required for diterpene ester-induced senescence. Expression profiling revealed that the MAPK inhibitor HREV107 was expressed at a higher transcript level in resistant compared to sensitive cell lines. We propose that activation of PKC over-stimulates the Ras/Raf/MEK/ERK pathway, resulting in sustained induction of p21WAF1/CIP1, dephosphorylation of Rb and transcriptional silencing of E2F-responsive genes required for DNA synthesis and mitosis. Keywords: expression profile following treatment
2006-01-20 | GSE3484 | GEO
Project description:We previously identified the induction of growth arrest with phenotypic characteristics of senescence in melanoma cell lines sensitive to diterpene esters, indicating a therapeutic potential. Here we compared the cytostatic effects of two diterpene esters namely TPA (12-O-tetradecanoylphorbol-13-acetate) and PEP008 (20-O-acetyl-ingenol-3-angelate) in sensitive and resistant cell lines derived from melanoma, breast cancer and colon cancer. We showed the diterpene esters to induce senescence-like growth arrest in the sensitive cells at 100-1000 ng/ml. Use of the pan-PKC inhibitor bisindolylmaleimide-l demonstrated that activation of PKC was required for growth arrest. Full genome expression profiling revealed that pivotal genes involved in DNA synthesis and cell cycle control were down-regulated by treatment in all three sensitive solid tumor models. At the protein level, prolonged down-regulation of E2F-1 and proliferating cell nuclear antigen (PCNA), sustained expression of p21WAF1/CIP1 and dephosphorylation of retinoblastoma (Rb) occurred in the sensitive cells. Although activation of extracellular signal-related kinase (ERK) 1/2 by the diterpene esters occurred in both sensitive and resistant cell lines, the HRASLS3 type II tumor suppressor, which appears to have a role in MAPK pathway suppression, was constitutively elevated in the resistant cell lines compared to their sensitive counterparts. Together, these results demonstrate the ability of the PKC activating drugs TPA and PEP008 to induce growth arrest with characteristics of senescence in solid tumor cell lines derived from a variety of tissue types through a similar mechanism. PKC-activating diterpene esters may therefore have therapeutic potential in a range of solid tumors. Experiment Overall Design: We analyzed the transcriptional profiles of the diterpene ester sensitive cell lines MCF7, COLO-205 and SK-MEL-5 following treatment with PEP008 using full genome expression profiling (Affymetrix, U133 Plus 2.0). Cells were treated for 24 h and 24 h plus 72 h recovery with 1000 ng/ml of the drug, before harvesting RNA for analysis. From the cell growth assays, all three cell lines demonstrated permanent growth arrest with diterpene ester treatments at the 1000 ng/ml dose. Mock controls were treated with solvent alone for 24 h. SK-MEL-5 cells were also treated with 1000 ng/ml TPA for 24 h.
2008-06-16 | E-GEOD-8742 | biostudies-arrayexpress
Project description:The diterpene ester PEP005 is a novel anticancer agent that activates PKC and cures subcutaneous murine melanoma by topical application. We now describe the in vitro cytostatic effects of PEP005 and the diterpene ester TPA, observed in 20% of human melanoma cell lines. Primary cultures of normal human neonatal fibroblasts were uniformly resistant to growth arrest, indicating a potential for tumor selectivity. Sensitive cells were induced to senesce and exhibited a G1 and G2/M arrest. There was sustained expression of p21WAF1/CIP1, irreversible dephosphorylation of the retinoblastoma gene product (Rb) and transcriptional silencing of E2F-responsive genes in sensitive cell lines. Activation of MEK1/2 by PKC was required for diterpene ester-induced senescence. Expression profiling revealed that the MAPK inhibitor HREV107 was expressed at a higher transcript level in resistant compared to sensitive cell lines. We propose that activation of PKC over-stimulates the Ras/Raf/MEK/ERK pathway, resulting in sustained induction of p21WAF1/CIP1, dephosphorylation of Rb and transcriptional silencing of E2F-responsive genes required for DNA synthesis and mitosis. To investigate the molecular changes associated with the senescent phenotype, we examined the unique transcriptional changes occurring in sensitive melanoma cell lines treated with TPA or PEP005. Initially a time-course cDNA microarray analysis was conducted on one sensitive and one resistant cell line treated with 1 µg/ml of TPA for 6, 24 h and 24 h recovery following 24 h treatment, to determine the earliest time point at which the most significant changes in transcription occurred. The results provided support for conducting array experiments with 24 h treatment, in which three sensitive and four resistant melanoma cell lines were treated with 1 µg/ml of either diterpene ester. Our primary objective was to identify those genes which were uniquely up or down-regulated in sensitive or resistant cell lines in response to treatment, which could reflect the phenotypic outcome. Through applying a series of stringent selective criteria (see Material and Methods) we found that the most significant changes occurred in the transcriptional repression of genes required for DNA synthesis and mitosis in cell lines sensitive to treatment (Table 1). To confirm that these changes were reflected at the protein level, western blot analysis was conducted on three sensitive and three resistant cell lines following 6 and 24 h treatment with TPA or PEP005. We also included a 24 h recovery time point to determine the irreversibility of the change.
2010-07-01 | E-GEOD-3484 | biostudies-arrayexpress
Project description:We previously identified the induction of growth arrest with phenotypic characteristics of senescence in melanoma cell lines sensitive to diterpene esters, indicating a therapeutic potential. Here we compared the cytostatic effects of two diterpene esters namely TPA (12-O-tetradecanoylphorbol-13-acetate) and PEP008 (20-O-acetyl-ingenol-3-angelate) in sensitive and resistant cell lines derived from melanoma, breast cancer and colon cancer. We showed the diterpene esters to induce senescence-like growth arrest in the sensitive cells at 100-1000 ng/ml. Use of the pan-PKC inhibitor bisindolylmaleimide-l demonstrated that activation of PKC was required for growth arrest. Full genome expression profiling revealed that pivotal genes involved in DNA synthesis and cell cycle control were down-regulated by treatment in all three sensitive solid tumor models. At the protein level, prolonged down-regulation of E2F-1 and proliferating cell nuclear antigen (PCNA), sustained expression of p21WAF1/CIP1 and dephosphorylation of retinoblastoma (Rb) occurred in the sensitive cells. Although activation of extracellular signal-related kinase (ERK) 1/2 by the diterpene esters occurred in both sensitive and resistant cell lines, the HRASLS3 type II tumor suppressor, which appears to have a role in MAPK pathway suppression, was constitutively elevated in the resistant cell lines compared to their sensitive counterparts. Together, these results demonstrate the ability of the PKC activating drugs TPA and PEP008 to induce growth arrest with characteristics of senescence in solid tumor cell lines derived from a variety of tissue types through a similar mechanism. PKC-activating diterpene esters may therefore have therapeutic potential in a range of solid tumors. Keywords: time course
2007-10-10 | GSE8742 | GEO
Project description:Activation of oncogene signaling in primary cells generally results in cellular senescence. This process is not only considered to be tumor-protective, but also irreversible. However, if senescence induction can be circumvented, e.g. by additional genetic or epigenetic changes, tumor progression occurs. An in-vivo example for oncogene-induced senescence are human nevus cells, most of which bear oncogenic mutations in RAS/RAF/MAPK pathway genes. Here, we show that expression of the human melanoma oncogene N-RAS61K in cultured pigment cells initially induces OIS characterized by a highly multinucleated phenotype. Surprisingly, after prolonged periods of oncogene expression, mononucleated cells emerge from the multinucleated cells. They are highly proliferative, anoikis-resistent and induce fast growing and metastatic melanoma upon transplantation into nude mice. During long-term oncogene expression and the corresponding development of anoikis iresistance, expression of melanocyte-specific genes is lost. Our data demonstrate that the induction of oncogene-induced senescence is not just a failsafe escape mechanism, but also provides a source for highly aggressive tumor cells. total samples analysed are 4
2014-01-20 | E-GEOD-27010 | biostudies-arrayexpress
Project description:Aberrant activation of the ERK signaling pathway triggers a protective anticancer response characterized by stable growth arrest and activation of tumor suppressors called cellular senescence. Pancreatic adenocarcinomas (PDAC) often possess mutations in K-Ras that activate the ERK pathway. Pancreatic intraepithelial neoplasia of low degree display high levels of phospho-ERK consistent with senescence acting as a barrier for malignant transformation. However, advanced lesions downregulate phospho-ERK levels circumventing the senescence barrier. Restoring ERK hyperactivation in PDAC using an activated allele of the kinase RAF, leads to ERK-dependent growth arrest with senescence biomarkers. Phosphoproteomics analysis of ERK-dependent senescence in PDAC revealed a decrease in several nucleolar phosphoproteins suggesting that high levels of ERK lead to senescence via nucleolar stress. Consistent with this explanation, ERK-dependent senescent cells displayed intranucleolar foci containing RNA polymerase I. Combining ribosome biogenesis inhibitors with ERK hyperactivation reinforced the senescence response of PDAC cells. The drug cocktail FOLFIRINOX, currently the best treatment for PDAC, also triggered ERK hyperactivation and nucleolar stress characterized by nucleolar foci, solid amyloid aggregates and a decrease in 5.8S and 28S rRNAs. We thus suggest that drugs targeting ribosome biogenesis can improve the senescence anticancer response in pancreatic cancer.
2024-06-10 | PXD032257 | Pride
Project description:Cutaneous melanoma is the deadliest form of skin cancer, frequently driven by hyperactivation of the RAS/mitogen-activated protein kinase (MAPK) pathway. Cyclin-dependent kinase 12 (CDK12), a downstream effector of MAPK signaling, has emerged as a therapeutic target due to its essential role in transcriptional regulation and DNA damage repair. To uncover vulnerabilities associated with CDK12 inhibition, we performed a genome-wide CRISPR/Cas9 screen and identified the Runt-related transcription factor RUNX1 and its cofactor CBFβ as synthetic lethal partners of CDK12. RUNX1 inhibition enhanced melanoma sensitivity to CDK12 inhibitors in a p53-independent manner, resulting in DNA damage accumulation and impaired repair capacity. Combined inhibition of CDK12 and RUNX1 suppressed melanoma growth in vivo. These findings uncover RUNX1/CBFβ as a compensatory mechanism in CDK12-inhibited melanoma and establish a novel synthetic lethal interaction with translational potential for combinatorial therapy.
2025-10-01 | GSE308214 | GEO
Project description:The eIF4F translation initiation complex plays a critical role in melanoma resistance to clinical BRAF and MEK inhibitors. In this study, we uncover a novel function of eIF4F in the negative regulation of the RAS/RAF/MEK/ERK mitogen-activated protein kinase (MAPK) signaling pathway. We demonstrate that eIF4F is essential for maintaining optimal ERK signaling intensity in treatment-naïve melanoma cells harboring BRAF or NRAS mutations. Specifically, the dual-specificity phosphatase DUSP6/MKP3, which acts as a negative feedback regulator of ERK activity, requires continuous production in an eIF4F-dependent manner to limit excessive ERK signaling driven by oncogenic RAF/RAS mutations. Treatment with small molecule eIF4F inhibitors disrupts the negative feedback control of MAPK signaling, leading to ERK hyperactivation and EGR1 overexpression in melanoma cells in vitro and in vivo. Furthermore, our quantitative analyses reveal a high spare signaling capacity in the ERK pathway, suggesting that eIF4F-dependent feedback keeps the majority of ERK molecules inactive under normal conditions. Overall, our findings highlight the crucial role of eIF4F in regulating ERK signaling flux and suggest that pharmacological eIF4F inhibitors can disrupt the negative feedback control of MAPK activity in melanomas with BRAF and NRAS activating mutations.
2024-10-04 | GSE273721 | GEO
Project description:The tumor suppressive miR-29 family of microRNAs is encoded by two clusters, miR-29b1~a and miR-29b2~c, and is regulated by several oncogenic and tumor suppressive stimuli. Here we investigated whether oncogenic MAPK hyperactivation regulates miR-29 abundance and how this signaling axis impacts melanoma development. Using mouse embryonic fibroblasts and human melanocytes, we found that oncogenic MAPK signaling stimulates p53-independent and p53-dependent transcription of pri-miR-29b1~a and pri-miR-29b2~c, respectively. Expression analyses revealed that while pri-miR-29a~b1 remains elevated, pri-miR-29b2~c levels decrease during melanoma progression. Using a rapid mouse modeling platform, we showed that inactivation of miR-29 in vivo accelerates the development of frank melanomas and decreases overall survival. We identified MAFG as a relevant miR-29 target that has oncogenic potential in melanocytes and is required for growth of melanoma cells. Our findings suggest that MAPK-driven miR-29 induction constitutes a tumor suppressive barrier by targeting MAFG, which is overcome by attenuation of miR-29b2~c expression.
2023-04-10 | GSE148552 | GEO
Project description:Activation of oncogene signaling in primary cells generally results in cellular senescence. This process is not only considered to be tumor-protective, but also irreversible. However, if senescence induction can be circumvented, e.g. by additional genetic or epigenetic changes, tumor progression occurs. An in-vivo example for oncogene-induced senescence are human nevus cells, most of which bear oncogenic mutations in RAS/RAF/MAPK pathway genes. Here, we show that expression of the human melanoma oncogene N-RAS61K in cultured pigment cells initially induces OIS characterized by a highly multinucleated phenotype. Surprisingly, after prolonged periods of oncogene expression, mononucleated cells emerge from the multinucleated cells. They are highly proliferative, anoikis-resistent and induce fast growing and metastatic melanoma upon transplantation into nude mice. During long-term oncogene expression and the corresponding development of anoikis iresistance, expression of melanocyte-specific genes is lost. Our data demonstrate that the induction of oncogene-induced senescence is not just a failsafe escape mechanism, but also provides a source for highly aggressive tumor cells.
2014-01-20 | GSE27010 | GEO