Cooperative epigenetic remodeling by TET2 loss and NRAS mutation drives myeloid transformation and MEK inhibitor sensitivity
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA379581 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR5352410.fastq.gz | Fastqsanger.gz | |||
| SRR5352420_1.fastq.gz | Fastqsanger.gz | |||
| SRR5352420_2.fastq.gz | Fastqsanger.gz | |||
| SRR5352430_1.fastq.gz | Fastqsanger.gz | |||
| SRR5352430_2.fastq.gz | Fastqsanger.gz |
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Project description:Recent studies using next-generation sequencing technology have uncovered mutational landscapes of various myeloid malignancies (Cancer Genome Atlas Research Network, 2013; Yoshida et al., 2011). These genetic data revealed novel classes of mutations that commonly occur in patients with myeloid malignancies, including epigenetic regulators and spliceosomal genes. In addition, co-occurrence and mutual exclusivity of these disease alleles suggest convergent cooperative roles or common biological effects of specific alleles in myeloid leukemogenesis (Shih et al., 2012). Somatic deletions and loss-of-function mutations in TET2, a member of the TET family which regulates DNA methylation, were identified in 10-20% of myelodysplastic syndromes (MDS) and myeloproliferative neoplasms (MPN) patients, 12-24% of acute myeloid leukemia (AML) patients and 40-50% of chronic myelomonocytic leukemia (CMML) patients (Abdel-Wahab et al., 2009; Delhommeau et al., 2009; Jankowska et al., 2009; Langemeijer et al., 2009). Analysis carried out in large AML cohorts has shown that TET2 mutations are associated with adverse outcome in patients with intermediate-risk, cytogenetically normal AML (Patel et al., 2012). TET proteins (TET1-3) are Fe(II)- and a-ketoglutarate-dependent enzymes that catalyze the conversion of 5-methylcytosine (5-mC) to 5-hydroxymethylcytosine (5-hmC) leading to DNA demethylation (Guo et al., 2011; Tahiliani et al., 2009). Consistent with this notion, TET2 mutant AML patient samples show decreased 5-hmC levels and hypermethylation phenotype (Figueroa et al., 2010; Ko et al., 2010). Oncometabolite 2-hydroxyglutarate produced by IDH1/2 mutations inhibits TET2 function (Figueroa et al., 2010). Moreover, WT1 recruits TET2 to regulate its target gene expression (Wang et al., 2015) and WT1 mutations lead to reduced TET2 function and decreased 5-hmC levels (Rampal et al., 2014). Together, IDH1/2 mutations and WT1 mutations share, at least partially, common epigenetic pathogenesis in AML as TET2 mutations through altered DNA hydroxymethylation. A number of studies have shown that TET2 is a key regulator of hematopoietic stem cell (HSC) self-renewal and myeloid differentiation. Conditional loss of Tet2 in mouse hematopoietic compartment leads to expansion of hematopoietic stem/progenitor cells (HSPCs, LSK, lin- Sca1+ cKit+) and enhanced repopulating capacity in vivo (Li et al., 2011; Moran-Crusio et al., 2011; Quivoron et al., 2011). In addition, Tet2-mutant mice show myeloproliferation in vivo and TET2-silenced human cord blood cells show skewed differentiation towards CD14+ myelomonocytic lineage in vitro (Pronier et al., 2011), which is compatible with a high frequency of TET2 mutations in CMML patients. More recently, mutations in TET2 and in other epigenetic regulators, such as DNMT3A and ASXL1, are reported in individuals with clonal hematopoiesis without hematological malignancies (Busque et al., 2012; Xie et al., 2014). Analysis of AML and patients with nonhematologic malignancies suggest mutations in TET2 and in other epigenetic modifiers may represent early premalignant events that cause clonal hematopoietic expansion (Genovese et al., 2014; Jaiswal et al., 2014; Jan et al., 2012). These data indicate that TET2 inactivation contributes to selective clonal advantages in early leukemia initiation but requires additional genetic events to induce myeloid transformation. Ras proteins are small GTPases that cycle between active guanosine triphosphate (GTP)-bound (Ras-GTP) and inactive guanosine diphosphate (GDP)-bound (Ras-GDP) conformations (Schubbert et al., 2007). Ras-GTP binds to and activates downstream signaling effectors, such as Raf and phosphoinositide 3-kinase (PI3K), thereby transducing signals from activated growth factor receptors to the nucleus. Thus, Ras-mediated signal transduction critically regulates fundamental cellular behaviors, including proliferation, differentiation and survival (Schubbert et al., 2007). The three cellular Ras genes encode 4 highly homologous proteins, Hras, Kras4A, Kras4B and Nras, that share conserved effector binding domains and the P-loop, which binds the g-phosphate of GTP (Zhou et al., 2016). Cancer-associated RAS mutations encode proteins that accumulate in the active GTP-bound conformation due to defective intrinsic hydrolysis and resistance to guanosine triphosphatase-activating proteins (Schubbert et al., 2007). Among these isoforms, NRAS is the most common target of oncogenic signaling mutations in myeloid leukemia, including 10-15% of AML and CMML cases (Bacher et al., 2006; Ball et al., 2016). Moreover, recent study identified NRAS as one of the somatic gene mutations with adverse prognostic relevance in CMML (Elena et al., 2016). Consistent with human data, mice with hematopoietic tissue specific conditional NrasG12D allele develop fatal myeloproliferative disease in vivo (Li et al., 2011). These data underscore critical role of oncogenic NRAS mutations in myeloid leukemogenesis. Recent studies have shown that mutations in epigenetic modifiers and signaling factors commonly co-occur in AML, including the co-occurrence of TET2 mutations and NRAS mutations (Papaemmanuil et al., 2016; Patel et al., 2012). Analysis of the clonal architecture in CMML patients suggests TET2/NRAS double-mutant clones expand both within stable disease time course and in relapse after allogeneic stem cell transplantation (Itzykson et al., 2013). These evidences imply TET2 and NRAS mutation likely function together in myeloid transformation. Although previous study has shown that knockdown of Tet2 does not accelerate NrasG12D induced transplant myeloproliferative disease model (Chang et al., 2014), to date there have been no studies of the mechanistic impact caused by the combination of these high risk genetic lesions using physiologic in vivo model. Moreover, how the combination of myeloid leukemia-associated disease alleles affect sensitivity to targeted therapy has not been elucidated. Here we investigate loss of Tet2 together with NrasG12D in CMML development and specifically how they interact to affect sensitivity to targeted therapy.
2018-03-17 | GSE96758 | GEO
Project description:Recent studies using next-generation sequencing technology have uncovered mutational landscapes of various myeloid malignancies (Cancer Genome Atlas Research Network, 2013; Yoshida et al., 2011). These genetic data revealed novel classes of mutations that commonly occur in patients with myeloid malignancies, including epigenetic regulators and spliceosomal genes. In addition, co-occurrence and mutual exclusivity of these disease alleles suggest convergent cooperative roles or common biological effects of specific alleles in myeloid leukemogenesis (Shih et al., 2012). Somatic deletions and loss-of-function mutations in TET2, a member of the TET family which regulates DNA methylation, were identified in 10-20% of myelodysplastic syndromes (MDS) and myeloproliferative neoplasms (MPN) patients, 12-24% of acute myeloid leukemia (AML) patients and 40-50% of chronic myelomonocytic leukemia (CMML) patients (Abdel-Wahab et al., 2009; Delhommeau et al., 2009; Jankowska et al., 2009; Langemeijer et al., 2009). Analysis carried out in large AML cohorts has shown that TET2 mutations are associated with adverse outcome in patients with intermediate-risk, cytogenetically normal AML (Patel et al., 2012). TET proteins (TET1-3) are Fe(II)- and a-ketoglutarate-dependent enzymes that catalyze the conversion of 5-methylcytosine (5-mC) to 5-hydroxymethylcytosine (5-hmC) leading to DNA demethylation (Guo et al., 2011; Tahiliani et al., 2009). Consistent with this notion, TET2 mutant AML patient samples show decreased 5-hmC levels and hypermethylation phenotype (Figueroa et al., 2010; Ko et al., 2010). Oncometabolite 2-hydroxyglutarate produced by IDH1/2 mutations inhibits TET2 function (Figueroa et al., 2010). Moreover, WT1 recruits TET2 to regulate its target gene expression (Wang et al., 2015) and WT1 mutations lead to reduced TET2 function and decreased 5-hmC levels (Rampal et al., 2014). Together, IDH1/2 mutations and WT1 mutations share, at least partially, common epigenetic pathogenesis in AML as TET2 mutations through altered DNA hydroxymethylation. A number of studies have shown that TET2 is a key regulator of hematopoietic stem cell (HSC) self-renewal and myeloid differentiation. Conditional loss of Tet2 in mouse hematopoietic compartment leads to expansion of hematopoietic stem/progenitor cells (HSPCs, LSK, lin- Sca1+ cKit+) and enhanced repopulating capacity in vivo (Li et al., 2011; Moran-Crusio et al., 2011; Quivoron et al., 2011). In addition, Tet2-mutant mice show myeloproliferation in vivo and TET2-silenced human cord blood cells show skewed differentiation towards CD14+ myelomonocytic lineage in vitro (Pronier et al., 2011), which is compatible with a high frequency of TET2 mutations in CMML patients. More recently, mutations in TET2 and in other epigenetic regulators, such as DNMT3A and ASXL1, are reported in individuals with clonal hematopoiesis without hematological malignancies (Busque et al., 2012; Xie et al., 2014). Analysis of AML and patients with nonhematologic malignancies suggest mutations in TET2 and in other epigenetic modifiers may represent early premalignant events that cause clonal hematopoietic expansion (Genovese et al., 2014; Jaiswal et al., 2014; Jan et al., 2012). These data indicate that TET2 inactivation contributes to selective clonal advantages in early leukemia initiation but requires additional genetic events to induce myeloid transformation. Ras proteins are small GTPases that cycle between active guanosine triphosphate (GTP)-bound (Ras-GTP) and inactive guanosine diphosphate (GDP)-bound (Ras-GDP) conformations (Schubbert et al., 2007). Ras-GTP binds to and activates downstream signaling effectors, such as Raf and phosphoinositide 3-kinase (PI3K), thereby transducing signals from activated growth factor receptors to the nucleus. Thus, Ras-mediated signal transduction critically regulates fundamental cellular behaviors, including proliferation, differentiation and survival (Schubbert et al., 2007). The three cellular Ras genes encode 4 highly homologous proteins, Hras, Kras4A, Kras4B and Nras, that share conserved effector binding domains and the P-loop, which binds the g-phosphate of GTP (Zhou et al., 2016). Cancer-associated RAS mutations encode proteins that accumulate in the active GTP-bound conformation due to defective intrinsic hydrolysis and resistance to guanosine triphosphatase-activating proteins (Schubbert et al., 2007). Among these isoforms, NRAS is the most common target of oncogenic signaling mutations in myeloid leukemia, including 10-15% of AML and CMML cases (Bacher et al., 2006; Ball et al., 2016). Moreover, recent study identified NRAS as one of the somatic gene mutations with adverse prognostic relevance in CMML (Elena et al., 2016). Consistent with human data, mice with hematopoietic tissue specific conditional NrasG12D allele develop fatal myeloproliferative disease in vivo (Li et al., 2011). These data underscore critical role of oncogenic NRAS mutations in myeloid leukemogenesis. Recent studies have shown that mutations in epigenetic modifiers and signaling factors commonly co-occur in AML, including the co-occurrence of TET2 mutations and NRAS mutations (Papaemmanuil et al., 2016; Patel et al., 2012). Analysis of the clonal architecture in CMML patients suggests TET2/NRAS double-mutant clones expand both within stable disease time course and in relapse after allogeneic stem cell transplantation (Itzykson et al., 2013). These evidences imply TET2 and NRAS mutation likely function together in myeloid transformation. Although previous study has shown that knockdown of Tet2 does not accelerate NrasG12D induced transplant myeloproliferative disease model (Chang et al., 2014), to date there have been no studies of the mechanistic impact caused by the combination of these high risk genetic lesions using physiologic in vivo model. Moreover, how the combination of myeloid leukemia-associated disease alleles affect sensitivity to targeted therapy has not been elucidated. Here we investigate loss of Tet2 together with NrasG12D in CMML development and specifically how they interact to affect sensitivity to targeted therapy.
2018-03-17 | GSE96759 | GEO
Project description:In the present work we propose a new therapy for NRAS mutant melanoma. Simultaneous inhibition of MEK and ROCK caused induction of BimEL , PARP, and Puma, and hence apoptosis. In vivo, MEK and ROCK inhibition suppressed growth of established tumors. Our findings warrant clinical investigation of the effectiveness of combinatorial targeting of MAPK/ERK and ROCK in NRAS mutant melanoma.
2015-03-11 | PXD000528 | Pride
Project description:Mutations in epigenetic modifiers and signaling factors often co-occur in myeloid malignancies, including TET2 and NRAS mutations. Concurrent Tet2 loss and NrasG12D expression in hematopoietic cells induced myeloid transformation, with a fully penetrant, lethal chronic myelomonocytic leukemia (CMML), which was serially transplantable. Tet2 loss and Nras mutation cooperatively led to decrease in negative regulators of mitogen-activated protein kinase (MAPK) activation, including Spry2, thereby causing synergistic activation of MAPK signaling by epigenetic silencing. Tet2/Nras double-mutant leukemia showed preferential sensitivity to MAPK kinase (MEK) inhibition in both mouse model and patient samples. These data provide insights into how epigenetic and signaling mutations cooperate in myeloid transformation and provide a rationale for mechanism-based therapy in CMML patients with these high-risk genetic lesions.
| S-EPMC5760367 | biostudies-literature
Project description:This SuperSeries is composed of the SubSeries listed below.
2018-03-17 | GSE96761 | GEO
Project description:NRAS-mutant melanoma is currently a challenge to treat. This is due to an absence of inhibitors directed against NRAS, along with acquired and adaptive resistance of this tumor type to inhibitors in the MAPK pathway. Inhibitors to MEK (mitogen-activated protein kinase kinase) have shown some promise for this tumor type. In this work we explored the use of MEK inhibitors for NRAS-mutant melanoma, and at the same time investigated the impact of the brain micro-environment, specifically astrocytes, on the response of a melanoma brain metastatic cell line to MEK inhibition. This led to the surprising finding that astrocytes enhance the sensitivity of melanoma tumors to MEK inhibitors (MEKi). We show that MEKi cause an upregulation of the transcription factor ID3, but this is blocked by conditioned media from astrocytes. We show that silencing ID3 enhances the sensitivity of melanoma to MEK inhibitors, thus mimicking the effect of the brain microenvironment. Moreover, we report that ID3 is a client protein of the chaperone HSP70, and that HSP70 inhibition causes ID3 to misfold and accumulate in a detergent-insoluble fraction in cells. We show that HSP70 inhibitors synergize with MEK inhibitors against NRAS-mutant melanoma, and that this combination significantly enhances the survival of mice in two different models of NRAS-mutant melanoma. These studies highlight ID3 as a mediator of adaptive resistance, and support the combined use of MEK and HSP70 inhibitors for the therapy of NRAS-mutant melanoma.
2022-06-30 | GSE179235 | GEO
Project description:Nearly 30% of all malignant melanomas harbor somatic mutations in NRAS. However, there are currently no effective targeted therapies for this tumor type. The bromodomain and extra terminal domain (BET) family of proteins are transcriptional regulators that serve as scaffolds to facilitate gene transcription by binding to acetylated lysine residues in the N-terminal tail of histones. BET/BRD proteins have emerged as therapeutic targets in a broad range of tumors. We found that BET proteins are overexpressed in NRAS mutant melanoma, and that high levels of BET family member BRD4 are associated with poor patient survival, suggesting that BRD4 plays a key role in melanoma. Consequently, we hypothesized that these epigenetic regulators constitute potential vulnerabilities that can be exploited for melanoma treatment. We found that genetic or pharmacological inhibition of BET/BRD proteins decreases viability and inhibits proliferation of NRAS mutant melanoma cells, as well as BRAF/MEK-inhibitor resistant melanoma cells harboring concurrent BRAF/NRAS mutations. However, BET inhibitors when used as single agents were either cytostatic (in vitro) or ineffective (in vivo). We therefore evaluated combinations that could maximize the efficacy of BET inhibitors in NRAS mutant melanoma. Here we report that co-targeting BET and MEK synergistically restrained tumor growth and significantly prolonged the survival of NRAS-mutant tumor bearing mice. RNA-sequencing and RPPA analysis revealed that co-treatment with BETi/MEKi synergistically downregulated cell cycle regulators and activated caspase-7. This study demonstrates that combined BET and MEK inhibition elicits robust synergistic therapeutic effects and supports the clinical utility of this combination therapy for NRAS mutant melanoma patients.
2018-02-20 | GSE95153 | GEO
Project description:Since direct pharmacological inhibition of RAS has thus far been unsuccessful, we explored system biology approaches to identify synergistic drug combination(s) that can mimic direct RAS inhibition. Leveraging an inducible mouse model of NRAS-mutant melanoma, we compare pharmacological MEK inhibition to complete NRAS-Q61K extinction in vivo. NRAS-Q61K extinction leads to a complete and durable tumor regression by enhancing both apoptosis and cell cycle arrest. By contrast, MEK inhibition only produces tumor stasis at best and we find that it robustly activates apoptosis but does not significantly impede proliferation. We used microarrays to determine which transcripts were affected by NRAS-Q61K extinction but insufficiently by MEK inhibition. We selected a single comparative timepoint, 4 days post-treatment. Downstream analyses included GSEA and TRAP algorithms, leading to the identification of a differentially affected CDK4-driven proliferation network. Immune genes were also identified as significant, but control experiments determined these to be largely the off-target effects of doxycycline and not of NRAS-Q61K extinction.
2012-08-09 | GSE39984 | GEO
Project description:Since direct pharmacological inhibition of RAS has thus far been unsuccessful, we explored system biology approaches to identify synergistic drug combination(s) that can mimic direct RAS inhibition. Leveraging an inducible mouse model of NRAS-mutant melanoma, we compare pharmacological MEK inhibition to complete NRAS-Q61K extinction in vivo. NRAS-Q61K extinction leads to a complete and durable tumor regression by enhancing both apoptosis and cell cycle arrest. By contrast, MEK inhibition only produces tumor stasis at best and we find that it robustly activates apoptosis but does not significantly impede proliferation. We used microarrays to determine which transcripts were affected by NRAS-Q61K extinction but insufficiently by MEK inhibition. We selected a single comparative timepoint, 4 days post-treatment. Downstream analyses included GSEA and TRAP algorithms, leading to the identification of a differentially affected CDK4-driven proliferation network. Immune genes were also identified as significant, but control experiments determined these to be largely the off-target effects of doxycycline and not of NRAS-Q61K extinction. The iNRAS-475 mouse melanoma cell line was injected intradermally into nude mice which were fed 2mg/ml doxycycline water. Tumors were allowed to reach 200-500mm3 after 6 weeks. Mice were then treated with vehicle or 100mg/kg of the AZD6244 MEK inhibitor, or doxycyline was withdrawn from the diet. Each sample represents a distinct tumor and thus provide six biological, not technical replicates per cohort.
2012-08-08 | E-GEOD-39984 | biostudies-arrayexpress
Project description:Mutations in the NRAS oncogene are present in up to 20% of melanoma. Here, we show that interferon alpha-inducible protein 6 (IFI6) is necessary for NRASQ61K-induced transformation and melanoma growth. IFI6 was transcriptionally upregulated by NRASQ61K, and knockdown of IFI6 resulted in DNA replication stress due to dysregulated DNA replication via E2F2. This stress consequentially inhibited cellular transformation and melanoma growth via senescence or apoptosis induction depending on the RB and p53 pathway status of the cells. NRAS-mutant melanoma were significantly more resistant to the cytotoxic effects of DNA replication stress-inducing drugs, and knockdown of IFI6 increased sensitivity to these drugs. Pharmacological inhibition of IFI6 expression by the MEK inhibitor trametinib, when combined with DNA replication stress-inducing drugs, blocked NRAS-mutant melanoma growth. Collectively, we demonstrate that IFI6, via E2F2 regulates DNA replication and melanoma development and growth, and this pathway can be pharmacologically targeted to inhibit NRAS-mutant melanoma. YUGASP cells stably expressing a non-silencing shRNA or two individual shRNAs against IFI6 were used to prepare the total RNA, which was then used to analyze for gene expression using Illumina expression array.
2016-09-08 | E-GEOD-69933 | biostudies-arrayexpress