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Expression data of HEL cells with or without LY2784544 and GRN163L

ABSTRACT: The mutant JAK2v617F is a driver of myeloproliferative neoplasms(MPNs). Consequently, several JAK2 inhibitors are currently being studied in clinical trials. The most investigated JAK2 inhibitor, Ruxolitinib, provides significant and sustained improvements in spleen related and constitutional symptoms secondary to the disease and represents a milestone in the treatment of myelofibrosis. However, JAK2 inhibitors are non-curative and murine experiments have shown that JAK2 inhibitors don?t eradicate MPN stem cells. In the present study, we determined the effect of the specific JAK2V617F inhibitor LY2784544 on leukemic stem (CD34+) cells (LSCs) using the JAK2V617F-bearing erythroleukemia cell line HEL. The LY2784544 treatment caused a transient proliferation inhibition and apoptosis of HEL cells, but a recovery occurred within a week. Thereafter, the continuous exposure of HEL cells to LY2784544 induced the accumulation of CD34+ LSCs, and the CD34+ cell fraction increased from 8% to over 90% by week 9, which was accompanied by substantially increased clonogenic potentials. A whole-transcript expression analysis revealed a significantly enhanced expression of the stem cell factor KLF4 in LY2784544 treated HEL cells. Inhibiting KLF4 expression attenuated LY2784544-mediated accumulation of CD34+ LSCs. Moreover, we further identified that the telomerase inhibitor GRN163L abolished the LY2784544-mediated CD34+ cell enrichment. Our findings collectively suggest that JAK2 inhibitors may cause enrichment of LSCs and are therefore unlikely to cure MPN as a monotherapy. Moreover, simultaneously targeting JAk2V617F and KLF4 or telomerase may be a novel strategy for MPN treatment.

ORGANISM(S): Homo sapiens

PROVIDER: GSE141602 | GEO | 2019/12/07

REPOSITORIES: GEO

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The HDAC inhibitor Givinostat modulates key hematopoietic genes in JAK2V617F cells, affecting proliferation, differentiation and apoptosis

Project description:We investigated the mechanism of action of the HDAC inhibitor Givinostat in JAK2V617F cells. We confirm that the drug inhibits colony formation and proliferation and induces apoptosis at doses 2-3 fold lower in JAK2V617F (HEL, UKE1 and SET2) compared to JAK2 wild type cell lines (K562, KU812, THP1 and KG1). By global gene expression analysis, we observed 293 common genes in HEL and UKE1 modulated at 6 hour by Givinostat (179 up and 114 down), of which 8/8 were validated by RTQ-PCR. 25, 28 and 33 modulated genes are implicated in the regulation of proliferation, apoptosis and hematopoiesis, respectively. Interestingly, 9 genes, known to be deregulated in MPN (myeloproliferative neoplasms) patients cells, were normalized by Givinostat. The hematopoietic transcription factors NFE2 and C-MYB were downmodulated by the drug specifically in JAK2V617F cells, and ETS1 was upregulated in all cell lines, at both the RNA and protein levels. Modulation of NFE2 and C-MYB was JAK2 dependent, as shown by use of the JAK2 inhibitor AG490. Finally, we suggest that the inhibition of NFE2 and induction of ETS1, also observed in freshly isolated CD34+ cells from MPN patients, may be at least in part responsible for the observed inhibition of erythroid differentiation by the drug. Gene expression profiling, JAK2V617F cell lines, ITF2357. This series of microarray experiments contains the gene expression profiles of independent triplicates of HEL and UKE1 erythroleukemia cell lines bearing the JAK2V617F mutation, before and after ITF2357 treatment. 100 nanograms of total RNA were processed, and fragmented biotin-labelled single-stranded DNA target was hybridized to the GeneChipÂ® Gene 1.0 ST array following the Affymetrix manufacturer's instructions.

2012-07-16 | E-GEOD-27615 | biostudies-arrayexpress

Integrated genomic analysis illustrates the central role of JAK-STAT pathway activation in myeloproliferative neoplasm pathogenesis [HEL cell lines]

Project description:We used expression profiling, SNP arrays, and mutational profiling to investigate a well-characterized cohort of MPN patients. MPN patients with homozygous JAK2V617F mutations were characterized by a distinctive transcriptional profile. Notably, a transcriptional signature consistent with activated JAK2 signaling is seen in all MPN patients regardless of clinical phenotype or mutational status. In addition, the activated JAK2 signature was present in patients with somatic CALR mutations. Conversely, we identified a gene expression signature of CALR mutations; this signature was significantly enriched in JAK2-mutant MPN patients consistent with a shared mechanism of transformation by JAK2 and CALR mutations. We also identified a transcriptional signature of TET2 mutations in MPN patent samples. Our data indicate that MPN patients, regardless of diagnosis or JAK mutational status are characterized by a distinct gene expression signature with upregulation of JAK-STAT target genes, demonstrating the central importance of the JAK-STAT pathway in MPN pathogenesis. [HEL cell lines] We have performed gene expression profiling in the JAK2V617F homozygous mutant HEL cell line following treatment with 2 independent shRNAs targeting JAK2 or 2 different control shRNAs

2014-06-03 | E-GEOD-54645 | biostudies-arrayexpress

The HDAC inhibitor Givinostat modulates key hematopoietic genes in JAK2V617F cells, affecting proliferation, differentiation and apoptosis

2012-07-17 | GSE27615 | GEO

Depletion of Jak2V617F MPN-propagating stem cells by interferon-alpha in a murine model of polycythemia vera

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2013-03-09 | E-GEOD-44961 | biostudies-arrayexpress

Physiological Jak2V617F expression causes a lethal myeloproliferative neoplasm with differential effects on hematopoietic stem and progenitor cells

Project description:We report a Jak2V617F knock-in mouse myeloproliferative neoplasm (MPN) model resembling human polycythemia vera (PV). The MPN is serially transplantable and we demonstrate that the hematopoietic stem cell (HSC) compartment has the unique capacity for disease initiation but does not have a selective competitive advantage over wild type HSCs. In contrast, myeloid progenitor populations are expanded and skewed towards the erythroid lineage, but cannot transplant the disease. Treatment with a JAK2 kinase inhibitor ameliorated the MPN phenotype, but did not eliminate the disease-initiating population. These findings provide insights into the consequences of JAK2 activation on HSC differentiation and function and have the potential to inform therapeutic approaches to JAK2V617F positive MPN. LKS cells were isolated from wild type (n=4) and JAK2V617F mutant mice (n=4). RNA was extracted using Qiagen RNeasy Micro Kit according to manufacturers instruction and amplified using NUGEN amplification kit. cDNA was fragmented and biotinylated before hybridization onto Affymetrix Mouse Expression Array 430 2.0.

2010-05-28 | E-GEOD-21842 | biostudies-arrayexpress

Jak2V617F Reversible Activation Shows an Essential Requirement for Jak2V617F in Myeloproliferative Neoplasms

Project description:Janus kinases (JAKs) mediate cytokine signaling, cell growth and hematopoietic differentiation. Gain-of-function mutations activating JAK2 signaling are seen in the majority of myeloproliferative neoplasm (MPN) patients, most commonly due to the JAK2V617F driver allele. While clinically-approved JAK inhibitors improve symptoms and outcomes in MPNs, remissions are rare, and mutant allele burden does not substantively change with chronic JAK inhibitor therapy in most patients. This has been postulated to be due to incomplete dependence on constitutive JAK/STAT signaling, alternative signaling pathways, and/or the presence of cooperating disease alleles; however we hypothesize this is due to the inability of current JAK inhibitors to potently and specifically abrogate mutant JAK2 signaling. We therefore developed a conditionally inducible mouse model allowing for sequential activation, and then inactivation, of Jak2V617F from its endogenous locus using a Dre-rox/Cre-lox dual orthogonal recombinase system. Deletion of oncogenic Jak2V617F abrogates the MPN disease phenotype, induces mutant-specific cell loss including in hematopoietic stem/progenitor cells, and extends overall survival to an extent not observed with pharmacologic JAK inhibition. Furthermore, reversal of Jak2V617F in MPN cells with antecedent loss of Tet2 abrogates the MPN phenotype and inhibits mutant stem cell persistence suggesting cooperating epigenetic-modifying alleles do not alter dependence on mutant JAK/STAT signaling. Our results suggest that mutant-specific inhibition of JAK2V617F represents the best therapeutic approach for JAK2V617F-mutant MPN and demonstrate the therapeutic relevance of a dual-recombinase system to assess mutant-specific oncogenic dependencies in vivo.

2022-05-25 | GSE203464 | GEO

Physiological Jak2V617F expression causes a lethal myeloproliferative neoplasm with differential effects on hematopoietic stem and progenitor cells

2010-05-15 | GSE21842 | GEO

Depletion of Jak2V617F MPN-propagating stem cells by interferon-alpha in a murine model of polycythemia vera

2013-03-09 | GSE44961 | GEO

Splicing factor Ybx1 maintains persistent Jak2-mutated neoplasms via Mknk1-ERK-signaling

Project description:Janus kinases (Jak) mediate cytokine, hormone and growth factor responses in hematopoietic cells. Jak2 is one of the most frequently mutated genes in the aging hematopoietic system and in hematopoietic cancers. Mutations in Jak constitutively activate downstream signaling and are drivers of myeloproliferative neoplasms (MPN). In clinical use, Jak-inhibitors have incomplete effects on overall disease burden of Jak2 mutated clones, prompting us to investigate the mechanism underlying disease persistence. By in-depth phospho-proteome profiling we here identify proteins involved in mRNA processing as targets of mutant Jak2. Inactivation of the post-translationally modified Jak2-target Ybx1 sensitizes Jak-inhibitor persistent cells to apoptosis and results in RNA mis-splicing, retained intron enrichment and disruption of the transcriptional control of extracellular signal-regulated kinase (ERK) signaling. In combination with pharmacological Jak-inhibition it induces apoptosis in Jak2-dependent murine and primary human cells, leading to in vivo regression of the malignant clones and inducing remission. This identifies and validates a novel cell-intrinsic mechanism how differential protein phosphorylation results in splicing-dependent alterations of Jak2-ERK-signaling and the maintenance of Jak2V617F malignant clones. Therapeutic targeting of Ybx1 dependent ERK-signaling in combination with Jak2-inhibition may eradicate Jak2-mutated cells.

2020-08-20 | PXD006921 | Pride

Integrated genomic analysis illustrates the central role of JAK-STAT pathway activation in myeloproliferative neoplasm pathogenesis [MPN patients]

Project description:We used expression profiling, SNP arrays, and mutational profiling to investigate a well-characterized cohort of MPN patients. MPN patients with homozygous JAK2V617F mutations were characterized by a distinctive transcriptional profile. Notably, a transcriptional signature consistent with activated JAK2 signaling is seen in all MPN patients regardless of clinical phenotype or mutational status. In addition, the activated JAK2 signature was present in patients with somatic CALR mutations. Conversely, we identified a gene expression signature of CALR mutations; this signature was significantly enriched in JAK2-mutant MPN patients consistent with a shared mechanism of transformation by JAK2 and CALR mutations. We also identified a transcriptional signature of TET2 mutations in MPN patent samples. Our data indicate that MPN patients, regardless of diagnosis or JAK mutational status are characterized by a distinct gene expression signature with upregulation of JAK-STAT target genes, demonstrating the central importance of the JAK-STAT pathway in MPN pathogenesis. [MPN patients] We have performed microarray gene expression analysis in 93 patients with MPNs (28 PV, 47 ET, 18 MF) and 11 age-matched normal donors.

2014-06-03 | E-GEOD-54644 | biostudies-arrayexpress

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