PAK1 is a Therapeutic Target in Acute Myeloid Leukemia and Myelodysplastic Syndrome
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ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE70791 | GEO | 2015/07/11
SECONDARY ACCESSION(S): PRJNA289538
REPOSITORIES: GEO
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Project description:Poor clinical outcome of Acute Myeloid Leukemia (AML) and Myelodysplastic Syndrome (MDS) has been attributed to failure of current chemotherapeutic regimens to target leukemic stem cells. We recently identified p21-activated kinase (PAK1) as a downstream effector molecule of H2.0-like homeobox (HLX), a gene functionally relevant for AML pathogenesis. In this study, we find that inhibition of PAK1 activity by small molecule inhibitors or by RNA interference leads to profound leukemia-inhibitory effects both in vitro and in vivo. Inhibition of PAK1 induces differentiation and apoptosis of AML cells through downregulation of MYC oncogene and a core network of MYC target genes. Moreover, we find that PAK1 up-regulation occurs during disease progression and is relevant for patient survival in MDS. Importantly, we find that inhibition of PAK1 inhibits primary human leukemic cells including immature leukemic stem cell-enriched populations. Our studies highlight PAK1 as a novel target in AML and MDS, and support the use of PAK1 inhibitors as a therapeutic strategy in these diseases. To obtain insight into the molecular mechanism for the induction of apoptosis and differentiation resulting from PAK1 inhibition in AML, we performed gene expression microarrays following treatment with either IPA-3 or FRAX-597. RNA was isolated from THP-1 cells after 5 hours of treatment with IPA-3 (6 ug/mL), FRAX-597 (2 ug/mL) or an equal volume of DMSO using Trizol Reagent (Invitrogen).
2015-07-11 | E-GEOD-70791 | biostudies-arrayexpress
Project description:Hypomethylating agents (HMAs) are frontline therapies effective at altering the natural course of Myelodysplastic Neoplasms (MDS). However, acquired resistance and treatment failure are hallmarks of HMA therapy. Developing effective and rational HMA-focused combinatorial therapies is challenging as the underlying mechanisms driving HMA efficacy are complex. To address this clinical need, we performed a genome-wide CRISPR-Cas9 screen in a human MDS-derived cell line, MDS-L, and characterized TOPORS as a highly ranked target that synergizes with HMAs to reduce leukemic burden and improve survival in xenograft models. We demonstrated that the depletion of TOPORS mediates sensitivity to HMAs by predisposing leukemic blasts to an impaired DNA damage response (DDR) accompanied by an accumulation of SUMOylated DNMT1 in HMA-treated TOPORS-depleted cells. Importantly, the combination of HMAs with targeting of TOPORS did not functionally impair healthy hematopoiesis. While inhibitors of TOPORS are currently unavailable, we show that inhibition of SUMOylation (upstream of TOPORS functions) with TAK-981 partially phenocopies HMA-sensitivity and DDR impairment. Overall, our data suggest that the combination of HMAs with the inhibition of SUMOylation demonstrates a favourable therapeutic index and represents a rational framework towards the treatment of High-Risk MDS (HR-MDS) or Acute Myeloid Leukemia (AML).
2024-07-25 | PXD050539 | Pride
Project description:Transcriptional repressor Growth factor independence 1 (GFI1) is a key regulator of haematopoiesis. We previously established that the germline variant GFI1-36N promotes acute myeloid leukemia (AML) development, however the mechanism is not full elucidated. Here using multi-omics approach, we show GFI1-36N expression impedes DNA repair in leukemic cells. We demonstrate the presence of GFI1-36N is associated with increased frequency of chromosomal aberrations and mutational burden in murine and human AML cells. In particular, GFI1-36N modulates DNA repair pathways, O6-methylguanine-DNA-methyltransferase (MGMT) and homologous recombination repair (HR). Mechanistically, GFI1-36N exhibits impaired binding to Ndrg1 promoter element compared to GFI1-36S (wild type), causing decreased NDRG1 levels consequently leading to suppression of MGMT expression, imprinted at the transcriptome and proteome, thus leaving the AML cells vulnerable to DNA damaging agents. Targeting MGMT via temozolomide and HR via olaparib caused specifically extensive lethality in in vitro and ex vivo human and AML samples expressing GFI1-36N. Whereas the effects were insignificant on non-malignant GFI1-36S or GFI1-36N cells. Further, mice transplanted with GFI1-36N leukemic cells treated with combination of temozolomide and olaparib had a significantly longer AML-free survival than mice transplanted with GFI1-36S leukemic cells. In summary, we show that GFI1-36N disturbs DNA repair activity via NDRG1-MGMT axis and thus provides critical insights into novel therapeutic option for AML presented with GFI1-36N variant. Key Points Presence of GFI1-36N impedes Homologous DNA and MGMT DNA repair selectively in AML cells via the NDRG1-MGMT axis. Use of temozolomide and olaparib allows selectively targeting GFI1-36N leukemic cells. Introduction Gfi1 is a transcription factor which regulates the development of haematopoietic cells as well as neuronal and intestinal epithelial cells 1-5. We reported that a variant of GFI1, denominated GFI1-36N (characterized by an exchange of serine to asparagine at position 36), has a prevalence of 5-7% in a healthy control population but is found at an increased frequency of 10-15% among MDS and AML patients 6,7. The expression of germline variant GFI1-36N predisposes the carriers to develop de novo AML and MDS and correlates with a poor prognosis 6,7. Recently, we and other showed that malignant cells with GFI1-36N variant have increased H3K9-acetylation at target genes resulting in higher expression of genes required for cell survival and proliferation 8. GFI1 exerts its repressive role by recruiting histone-modifying enzymes (deacetylases HDAC1-3, demethylase LSD1, methyl transferase G9a) and regulates the accessibility of DNA to its target genes such as Hoxa9, Pbx1, Meis1, CSF1 and CSFR1 9-15. We also showed that GFI1 regulates apoptosis through its regulation of p53 in lymphoblastic leukemia 16 and we have demonstrated that GFI1 facilitates DNA repair 17. However, it is not known how these activities are affected in the GFI1-36N variant and whether the ability of GFI1 to regulate DNA repair pathways is maintained and how this might affect the development of myeloid malignancies. In this study, we leveraged multi-omics profiling to gain mechanistic insights into the molecular architecture that drives leukemia in the presence of GFI1-36N. We provide evidence that GFI1-36N interferes with DNA in leukemic myeloid cells, which leads to a higher frequency of genetic aberrations in MDS/AML cells. We also show that GFI1-36N myeloid leukemic cells are more sensitive to targeting MGMT and HR repair deficient cells, which opens a new selective therapeutic window to treat AML/MDS.
2024-01-26 | PXD037433 | Pride
Project description:Epigenetic pathways regulate gene expression by controlling and interpreting chromatin modifications. Cancer cells are characterized by altered epigenetic landscapes and commonly exploit the chromatin regulatory machinery to enforce oncogenic gene expression programs. While chromatin alterations are, in principle, reversible and often amendable to drug intervention, the promise of targeting such pathways therapeutically has been hampered by our limited understanding of cancer-specific epigenetic dependencies. Here we describe a non-biased approach to probe epigenetic vulnerabilities in acute myeloid leukemia (AML) – an aggressive hematopoietic malignancy often associated with aberrant chromatin states. By screening a custom shRNA library targeting known chromatin regulators in a genetically defined AML mouse model, we identify the bromodomain-containing protein Brd4 as a critical requirement for disease maintenance. Suppression of Brd4 using shRNAs or the small-molecule inhibitor JQ1 led to robust anti-leukemic effects in vitro and in vivo, accompanied by terminal myeloid differentiation. Extensive evaluation of JQ1-sensitivity in primary human leukemia samples and in established cell lines revealed a broad activity of this compound against diverse AML subtypes. These effects are, at least in part, due to a requirement for Brd4 in maintaining Myc expression and promoting aberrant self-renewal. Together, our results indicate that Brd4 is a promising therapeutic target in AML and identify a small molecule that efficiently targets Myc. These findings also highlight the utility of RNAi screening as a discovery platform for revealing epigenetic vulnerabilities for direct pharmacologic intervention in cancer. In order to understand downstream targets of Brd4, we performed array in murine or human MLL-AF9/NrasG12D cell line under the condition that Brd4 was suppressed by using shRNAs or the small molecule inhibitor JQ1. To test the hypothesis that Myc might be an important target of Brd4, we performed arrary on murine ectopic Myc overexpression MLL-AF9/NrasG12D cell under JQ1 treatment.
2011-08-03 | E-GEOD-29799 | biostudies-arrayexpress
Project description:10 to 30% normal karyotype acute myeloid leukemia (AML) patients express mutations in regulators of DNA methylation such as TET2 or DNMT3A in conjunction with activating mutation in receptor tyrosine kinase, FLT3, which can be present in up to 50% of normal karyotype AML patients. These patients have poor prognosis since they do not respond well to established therapies. Here, utilizing mouse models of AML that recapitulate cardinal features of the human disease and bear a combination of loss of function mutations in either Tet2 or Dnmt3a along with expression of Flt3ITD, we show that inhibition of SHP2, a protein tyrosine phosphatase, essential for cytokine receptor signaling, including FLT3, by a small molecule allosteric inhibitor, SHP099, impairs growth and induces differentiation of leukemic cells without impacting normal hematopoietic cells. We further show that SHP099 normalizes gene expression program associated with increased cell proliferation and self-renewal in leukemic cells by down regulating the Myc signature. Our results provide a new and more effective target for treating a subset of AML patients bearing a combination of genetic and epigenetic mutations.
2019-07-26 | GSE134843 | GEO
Project description:Homeobox (HOX) proteins and the receptor tyrosine kinase FLT3 are frequently highly expressed and mutated in acute myeloid leukemia (AML). Aberrant HOX expression is found in nearly all AMLs that harbor a mutation in the Nucleophosmin (NPM1) gene, and FLT3 is concomitantly mutated in approximately 60% of these cases. Little is known how mutant NPM1 (NPM1mut) cells maintain aberrant gene expression. Here, we demonstrate that the histone modifiers MLL1 and DOT1L control HOX and FLT3 expression and differentiation in NPM1mut AML. Using a CRISPR-Cas9 genome editing domain screen, we show NPM1mut AML to be exceptionally dependent on the menin binding site in MLL1. Pharmacological small-molecule inhibition of the menin-MLL protein interaction had profound anti-leukemic activity in human and murine models of NPM1mut AML in vitro and in vivo. Combined pharmacological inhibition of menin-MLL and DOT1L resulted in dramatic suppression of HOX and FLT3 expression, induction of differentiation, and superior activity against NPM1mut leukemia. Together, MLL1 and DOT1L are chromatin regulators that control HOX, MEIS1 and FLT3 expression and are therapeutic targets in NPM1mut AML. Combinatorial small-molecule inhibition has synergistic on target activity and constitutes a novel therapeutic concept for this common AML subtype. RNA sequencing data of the human AML cell lines OCI-AML2 and OCI-AML3 comparing EPZ004777 [10µM] drug treatment versus DMSO vehicle control; experiments performed in biological triplicates.
2016-08-22 | E-GEOD-85107 | biostudies-arrayexpress
Project description:MDS is characterized by a disturbed function of the myeloid lineage of the hematopoietic system that may transform to AML, a malignant disease of the myeloid compartment. Epigenetic and genetic aberrations contribute to the initiation and progression of MDS/AML. GFI1 is a transcriptional repressor, which regulates expression of its target genes by, among other approaches, recruiting HDACs to its target genes to remove histone 3 lysine 9 (H3K9) acetylation, a marker for active gene expression. Low levels of GFI1 expression and deletion of one GFI1 allele contribute to MDS/AML development in human patients and are associated with a specific gene expression signature and inferior prognosis. To explore the mechanism behind this, we used a mouse strain, which expresses GFI1 only at 5-10% of the normal level (GFI1-Knock-down (KD)). Knock-down of GFI1 or loss of one murine Gfi1 allele reduced latency and increased incidence of AML in different murine models of human MDS/AML development. On the epigenetic level, KD of Gfi1 lead to increased amount of H3K9 acetylation, resulting in increased expression of genes involved in AML development. On a translational level both murine as well as human AML cells with low expression of GFI1 are resistant to standard epigenetic therapy. We show that treatment with histone acetyltransferase inhibitors might be a novel treatment approach for low Gfi1-expressing blast cells. GFI1 has a dose dependent role in myeloid malignancies and is a biomarker for therapeutic intervention. We used microarrays to detail the global programme of gene expression in bone marrow cells of Nup98HoxD13 leucemic mice and identified distinct classes of up-regulated genes during this process. Bone marrow cells of leukemic mice (from KI & KD Nup98HoxD13 mice) were collected for RNA extraction and hybridization on Affymetrix microarrays. We sought to obtain homogeneous populations of leukemic cell population, so we took samples of mice which had the same age.
2016-07-31 | E-GEOD-72489 | biostudies-arrayexpress
Project description:Hypomethylating agents (HMAs) are frontline therapies effective at altering the natural course of Myelodysplastic Neoplasms (MDS). However, acquired resistance and treatment failure are hallmarks of HMA therapy. Developing effective and rational HMA-focused combinatorial therapies is challenging as the underlying mechanisms driving HMA efficacy are complex. To address this clinical need, we performed a genome-wide CRISPR-Cas9 screen in a human MDS-derived cell line, MDS-L, and characterized TOPORS as a highly ranked target that synergizes with HMAs to reduce leukemic burden and improve survival in xenograft models. We demonstrated that the depletion of TOPORS mediates sensitivity to HMAs by predisposing leukemic blasts to an impaired DNA damage response (DDR) accompanied by an accumulation of SUMOylated DNMT1 in HMA-treated TOPORS-depleted cells. Importantly, the combination of HMAs with targeting of TOPORS did not functionally impair healthy hematopoiesis. While inhibitors of TOPORS are currently unavailable, we show that inhibition of SUMOylation (upstream of TOPORS functions) with TAK-981 partially phenocopies HMA-sensitivity and DDR impairment. Overall, our data suggest that the combination of HMAs with the inhibition of SUMOylation demonstrates a favourable therapeutic index and represents a rational framework towards the treatment of High-Risk MDS (HR-MDS) or Acute Myeloid Leukemia (AML).
2024-07-24 | GSE261338 | GEO
Project description:Hypomethylating agents (HMAs) are frontline therapies effective at altering the natural course of Myelodysplastic Neoplasms (MDS). However, acquired resistance and treatment failure are hallmarks of HMA therapy. Developing effective and rational HMA-focused combinatorial therapies is challenging as the underlying mechanisms driving HMA efficacy are complex. To address this clinical need, we performed a genome-wide CRISPR-Cas9 screen in a human MDS-derived cell line, MDS-L, and characterized TOPORS as a highly ranked target that synergizes with HMAs to reduce leukemic burden and improve survival in xenograft models. We demonstrated that the depletion of TOPORS mediates sensitivity to HMAs by predisposing leukemic blasts to an impaired DNA damage response (DDR) accompanied by an accumulation of SUMOylated DNMT1 in HMA-treated TOPORS-depleted cells. Importantly, the combination of HMAs with targeting of TOPORS did not functionally impair healthy hematopoiesis. While inhibitors of TOPORS are currently unavailable, we show that inhibition of SUMOylation (upstream of TOPORS functions) with TAK-981 partially phenocopies HMA-sensitivity and DDR impairment. Overall, our data suggest that the combination of HMAs with the inhibition of SUMOylation demonstrates a favourable therapeutic index and represents a rational framework towards the treatment of High-Risk MDS (HR-MDS) or Acute Myeloid Leukemia (AML).
2024-07-24 | GSE261337 | GEO
Project description:Firstly, we extracted bone marrow cells from AML mouse models in four different timepoints (T0, T1, T2 and T3). And we used droplet-based single cell RNA sequencing technology to reveal the early-MDS, late-MDS, MDS-AML and AML four-stage landscape in a mouse AML model initiated by Myc overexpression. Also, we extracted one AML patient bone marrow sample to sequence by constructing library following 10X genomics platform. And integrated this data with four AML patients data from 10X genomics database to explain the molecular mechanism of splicing events of TMEM134 in AML. To verify the alternative splicing results, we used smartseq2 to construct the single cell library in T3. And combined TCGA-LAML and TARGET-AML database and our mouse function data, we identify the alternative splicing events of TMEM134 could drive the proliferation function in acute myelogenous leukemogenesis.
2023-01-30 | GSE142645 | GEO