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Journal ArticleDOI

A wide role for NOTCH1 signaling in acute leukemia

TL;DR: The expression of genes belonging to NOTCH pathway, in acute leukemia primary samples and lymphoblastoid cell lines, shows high levels of JAGGED1 expression despite the low NOTCH1 pathway activation, consistent with an autonomous JAGgED1 signaling in myeloid leukemogenesis.
About: This article is published in Cancer Letters.The article was published on 2005-02-28 and is currently open access. It has received 77 citations till now. The article focuses on the topics: Acute leukemia & Leukemia.

Summary (2 min read)

1. Introduction

  • NOTCH gene family encodes single pass transmembrane receptors able to transduce intercellular signals involved in cell-fate determination [1].
  • The fundamental role played by the NOTCH pathway in T-lineage proliferation and differentiation has a counterpart in the involvement of NOTCH signaling alterations in the pathogenesis of T-cell acute lymphoblastic leukemia (T-ALL).
  • By contrast in AML the authors found high levels of JAGGED1 expression despite of the low expression of NOTCH1 pathway genes, consistent with a potential role of autonomous JAGGED1 signaling in myeloid leukemogenesis.

2.1. Cell cultures and blood samples

  • Under informed consent by the parents or the guardians, 32 children with T-ALL, 25 with B-cell precursor ALL and 20 with AML were included in the study.
  • All patients have been referred to a single Institution (Clinica Pediatrica of Monza, Italy) and treated according to the protocols of the ‘Associazione Italiana Ematologia ed Oncologia Pediatrica’ .
  • Peripheral blood mononuclear cells (PBMC) have been separated on Ficoll-Hypaque gradient, frozen as viable cells and kept at K80 8C.
  • Cell lines were cultured as previously reported [12] in 5% CO2 atmosphere in complete RPMI-1640 medium supplemented with 10% foetal calf serum.

2.2. Retrotranscription and amplification

  • Total RNA was extracted according to Chomczynski and Sacchi [13].
  • RNA samples were retrotranscripted by M-MLV Reverse Transcriptase (Gibco BRL-Life Technologies) in the conditions suggested.
  • Results of electrophoresis were acquired by the image acquisition system EDAS 2400 and following analysed by 1D image analysis system.

2.3. Statistical analysis

  • All values resulting from image analysis of PCR products were normalized against the corresponding values obtained from GAPDH amplification.
  • The resulting values were then statistically analysed by ANOVA (analysis of variance) test to reveal if there is any statistical difference in gene expression associated with the different leukemia cell samples, T-ALL, BCP–ALL and AML.
  • Analysis was independently performed for each gene.
  • The rejection of the null hypothesis (H0) (P-value!0.01) drives to the conclusion that the analysed gene expression levels are different in, at least, two among the analysed populations.
  • A following multiple comparison test (Tukey test) was performed to determine which particular population differs from the others: the difference between values distributions of two samples groups is significant when P!0.05; is not significant when PO0.05.

3.1. Identification of NOTCH pathway target genes

  • The authors have previously reported the concordance between HES1 and NOTCH1 genes expression both in T-cell ALL and B-cell lymphoma cell lines [12].
  • To identify further NOTCH pathway target genes the authors have expanded the analysis to the genes reported to be modulated by NOTCH pathway: pTa (the gene for preTCR-a), DELTEX, HERP2, HES5, MELTRINb (an ADAM family metallo-protease), IFI16 and MNDA [14,15].
  • Only the expression of pTa gene correlates with NOTCH1 and HES1 expression (data not shown).
  • Accordingly, the subsequent analyses on primary leukemic cells were limited to NOTCH1, HES1 and pTa as marker of NOTCH1 signaling.

3.2. NOTCH1 and ligands expression in T-ALL cell lines

  • To assess a possible contribution of the ligands in the NOTCH pathway activation the authors performed RTPCR analyses on T-ALL leukemia cell lines of transcripts for all the DSL ligands.
  • PBMC isolated from normal get genes, HES1 and pTa, and the ligands JAGGED1 and DLL-4, in ML.
  • The house keeping gene GAPDH has been used for normalization.
  • Donors and the EW36 B lymphoma cell line were used as controls.

4. Discussion

  • During T-cell lineage development NOTCH1 expression is finely regulated [16]: NOTCH1 drives cell fate decision (the choice between TCRg/d or a/b and between CD4C or CD8C) by inductive interactions from thymic stromal cells [17]; its expression is physiologically down-regulated to permit successful maturation by interfering with TCR signal strength [18].
  • Recent evidences further support the hypothesis that several gands.
  • Both ligands were selected since DLL-4 overexpression in mouse bone marrow results in T Cell leukemia [9].
  • The authors have found that JAGGED1 is highly expressed only in AML samples, thus confirming a selective role of this ligand in AML [11].
  • In conclusion the expression analysis of genes involved in NOTCH1 signaling suggests that NOTCH1 pathway exerts a wide role in acute leukemia.

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Citations
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Journal ArticleDOI
15 Mar 2006-Blood
TL;DR: This review highlights recent findings regarding the molecular and functional aspects of Notch-mediated neoplastic transformation and cellular mechanisms that potentially explain the complex role of notch in tumorigenesis.

516 citations

Journal ArticleDOI
TL;DR: Restoration of Ikaros activity in tumor cells leads to a rapid and specific downregulation of Notch target gene expression and proliferation arrest, which may play a critical role in defining the tumor suppressor function of this factor.
Abstract: The Ikaros transcription factor is both a key regulator of lymphocyte differentiation and a tumor suppressor in T lymphocytes. Mice carrying a hypomorphic mutation (Ik(L/L)) in the Ikaros gene all develop thymic lymphomas. Ik(L/L) tumors always exhibit strong activation of the Notch pathway, which is required for tumor cell proliferation in vitro. Notch activation occurs early in tumorigenesis and may precede transformation, as ectopic expression of the Notch targets Hes-1 and Deltex-1 is detected in thymocytes from young Ik(L/L) mice with no overt signs of transformation. Notch activation is further amplified by secondary mutations that lead to C-terminal truncations of Notch 1. Strikingly, restoration of Ikaros activity in tumor cells leads to a rapid and specific downregulation of Notch target gene expression and proliferation arrest. Furthermore, Ikaros binds to the Notch-responsive element in the Hes-1 promoter and represses Notch-dependent transcription from this promoter. Thus, Ikaros-mediated repression of Notch target gene expression may play a critical role in defining the tumor suppressor function of this factor.

167 citations


Cites background from "A wide role for NOTCH1 signaling in..."

  • ...Recently, activating mutations in the Notch 1 gene have been found in more than 50% of T-ALLs (55), and T-ALL samples frequently express the Notch target genes Hes-1 and pT (11)....

    [...]

Journal ArticleDOI
TL;DR: It is proposed that the Notch signaling pathway may represent novel therapeutic targets and will be a welcome asset to the cancer therapeutic arena.

163 citations

Journal ArticleDOI
TL;DR: Comparing gene expression profiles between normal hematopoietic cells from 38 healthy donors and leukemic blasts from 26 AML patients found that microarray studies could identify previously unrecognized expression changes that occur only in AML blasts.
Abstract: Acute myeloid leukemia (AML) is one of the most common and deadly forms of hematopoietic malignancies. We hypothesized that microarray studies could identify previously unrecognized expression changes that occur only in AML blasts. We were particularly interested in those genes with increased expression in AML, believing that these genes may be potential therapeutic targets. To test this hypothesis, we compared gene expression profiles between normal hematopoietic cells from 38 healthy donors and leukemic blasts from 26 AML patients. Normal hematopoietic samples included CD34+ selected cells (N = 18), unselected bone marrows (N = 10), and unselected peripheral bloods (N = 10). Twenty genes displayed AML-specific expression changes that were not found in the normal hematopoietic cells. Subsequent analyses using microarray data from 285 additional AML patients confirmed expression changes for 13 of the 20 genes. Seven genes (BIK, CCNA1, FUT4, IL3RA, HOMER3, JAG1, WT1) displayed increased expression in AML, while 6 genes (ALDHA1A, PELO, PLXNC1, PRUNE, SERPINB9, TRIB2) displayed decreased expression. Quantitative RT/PCR studies for the 7 over-expressed genes were performed in an independent set of 9 normal and 21 pediatric AML samples. All 7 over-expressed genes displayed an increased expression in the AML samples compared to normals. Three of the 7 over-expressed genes (WT1, CCNA1, and IL3RA) have already been linked to leukemogenesis and/or AML prognosis, while little is known about the role of the other 4 over-expressed genes in AML. Future studies will determine their potential role in leukemogenesis and their clinical significance.

161 citations


Cites background from "A wide role for NOTCH1 signaling in..."

  • ...There has been a recent flurry of studies examining the JAG1/Notch pathway in normal hematopoiesis and stem cell renewal (Tohda and Nara 2001; Alcalay et al., 2003; Chiaramonte et al., 2005; Zweidler-McKay et al., 2005)....

    [...]

  • ...normal hematopoiesis and stem cell renewal (Tohda and Nara 2001; Alcalay et al., 2003; Chiaramonte et al., 2005; Zweidler-McKay et al., 2005)....

    [...]

Journal ArticleDOI
TL;DR: Activating Notch with a Notch agonist peptide induces apoptosis in AML patient samples and this work has implications for drug discovery and clinical practice.
Abstract: Although aberrant Notch activation contributes to leukemogenesis in T cells, its role in acute myelogenous leukemia (AML) remains unclear. Here, we report that human AML samples have robust expression of Notch receptors; however, Notch receptor activation and expression of downstream Notch targets are remarkably low, suggesting that Notch is present but not constitutively activated in human AML. The functional role of these Notch receptors in AML is not known. Induced activation through any of the Notch receptors (Notch1–4), or through the Notch target Hairy/Enhancer of Split 1 (HES1), consistently leads to AML growth arrest and caspase-dependent apoptosis, which are associated with B cell lymphoma 2 (BCL2) loss and enhanced p53/p21 expression. These effects were dependent on the HES1 repressor domain and were rescued through reexpression of BCL2. Importantly, activated Notch1, Notch2, and HES1 all led to inhibited AML growth in vivo, and Notch inhibition via dnMAML enhanced proliferation in vivo, thus revealing the physiological inhibition of AML growth in vivo in response to Notch signaling. As a novel therapeutic approach, we used a Notch agonist peptide that led to significant apoptosis in AML patient samples. In conclusion, we report consistent Notch-mediated growth arrest and apoptosis in human AML, and propose the development of Notch agonists as a potential therapeutic approach in AML.

136 citations

References
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Journal ArticleDOI
TL;DR: A new method of total RNA isolation by a single extraction with an acid guanidinium thiocyanate-phenol-chloroform mixture is described, providing a pure preparation of undegraded RNA in high yield and can be completed within 4 h.

65,881 citations


"A wide role for NOTCH1 signaling in..." refers methods in this paper

  • ...Total RNA was extracted according to Chomczynski and Sacchi [13]....

    [...]

Journal ArticleDOI
23 Aug 1991-Cell
TL;DR: It is shown that the locus on chromosome 9 contains a gene highly homologous to the Drosophila gene Notch, which may be important for normal lymphocyte function and that alteration of TAN-1 may play a role in the pathogenesis of some T cell neoplasms.

1,781 citations

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01 May 1999-Immunity
TL;DR: It is suggested that Notch1 plays an obligatory and selective role in T cell lineage induction in mice with a neonatally induced loss of Notch 1 function.

1,442 citations

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TL;DR: It is shown that five different T cell oncogenes are often aberrantly expressed in the absence of chromosomal abnormalities, and HOX11L2 activation is identified as a novel event in T cell leukemogenesis.

1,063 citations

Journal ArticleDOI
TL;DR: Results show that TAN1 is an oncoprotein and suggest that truncation and overexpression are important determinants of transforming activity in hematopoietic cells.
Abstract: Notch is a highly conserved transmembrane protein that is involved in cell fate decisions and is found in organisms ranging from Drosophila to humans. A human homologue of Notch, TAN1, was initially identified at the chromosomal breakpoint of a subset of T-cell lymphoblastic leukemias/lymphomas containing a t(7;9) chromosomal translocation; however, its role in oncogenesis has been unclear. Using a bone marrow reconstitution assay with cells containing retrovirally transduced TAN1 alleles, we analyzed the oncogenic potential of both nuclear and extranuclear forms of truncated TAN1 in hematopoietic cells. Although the Moloney leukemia virus long terminal repeat drives expression in most hematopoietic cell types, retroviruses encoding either form of the TAN1 protein induced clonal leukemias of exclusively immature T cell phenotypes in approximately 50% of transplanted animals. All tumors overexpressed truncated TAN1 of the size and subcellular localization predicted from the structure of the gene. These results show that TAN1 is an oncoprotein and suggest that truncation and overexpression are important determinants of transforming activity. Moreover, the murine tumors caused by TAN1 in the bone marrow transplant model are very similar to the TAN1-associated human tumors and suggest that TAN1 may be specifically oncotropic for T cells.

720 citations

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Q1. What have the authors contributed in "A wide role for notch1 signaling in acute leukemia" ?

The authors analyzed the expression of genes belonging to NOTCH pathway, in acute leukemia primary samples and lymphoblastoid cell lines.