P53 signaling in response to increased DNA damage sensitizes AML1-ETO cells to stress-induced death

Division of Experimental Hematology, Cincinnati Children's Hospital Medical Center, University of Cincinnati College of Medicine, OH 45226, USA.
Blood (Impact Factor: 10.45). 03/2008; 111(4):2190-9. DOI: 10.1182/blood-2007-06-093682
Source: PubMed


Chromosomal translocation (8;21) is present in 10% to 15% of patients with acute myeloid leukemia. Expression of the AML1-ETO (AE) fusion protein alone is not sufficient to induce leukemia, but the nature of the additional genetic alterations is unknown. It is unclear whether AE facilitates acquisition of these cooperating events. We show that AE down-regulates genes involved in multiple DNA repair pathways, potentially through a mechanism involving direct binding at promoter elements, and increases the mutation frequency in vivo. AE cells display increased DNA damage in vitro and have an activated p53 pathway. This results in increased basal apoptosis and enhanced sensitivity to DNA damaging agents. Intriguingly, microarray data indicate that t(8;21) patient samples exhibit decreased expression of DNA repair genes and increased expression of p53 response genes compared with other acute myeloid leukemia (AML) patient samples. Inhibition of the p53 pathway by RNAi increases the resistance of AE cells to DNA damage. We thus speculate that AML1-ETO may facilitate accumulation of genetic alterations by suppressing endogenous DNA repair. It is possible that the superior outcome of t(8;21) patients is partly due to an activated p53 pathway, and that loss of the p53 response pathway is associated with disease progression.

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Available from: James C Mulloy, Feb 27, 2015
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    • "Additionally to ROS-mediated mutagenesis, the expression of leukemia-associated fusion genes may interfere with DNA repair as convincingly demonstrated for breakpoint cluster region/ Abelson 1 (BCR/ABL1) leukemia [21]. DNA repair is also disturbed in leukemic cells characterized by translocation products such as RUNX1/ ETO (AML1/ETO) [22] [23] and in cases of ETV6/RUNX1 + leukemia [12]. ETV6/RUNX1 + cells also display a disrupted spindle checkpoint that may result in aneuploidy [24]. "
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    ABSTRACT: The t(12;21)(p13;q22) chromosomal translocation is the most frequent translocation in childhood B cell precursor-acute lymphoblastic leukemia and results in the expression of an ETV6/RUNX1 fusion protein. The frequency of ETV6/RUNX1 fusions in newborns clearly exceeds the leukemia rate revealing that additional events occur in ETV6/RUNX1-positive cells for leukemic transformation. Hitherto, the mechanisms triggering these second hits remain largely elusive. Thus, we generated a novel ETV6/RUNX1 transgenic mouse model where the expression of the fusion protein is restricted to CD19(+) B cells. These animals harbor regular B cell development and lack gross abnormalities. We established stable pro-B cell lines carrying the ETV6/RUNX1 transgene that allowed us to investigate whether ETV6/RUNX1 itself favors the acquisition of second hits. Remarkably, these pro-B cell lines as well as primary bone marrow cells derived from ETV6/RUNX1 transgenic animals display elevated levels of reactive oxygen species (ROS) as tested with ETV6/RUNX1 transgenic dihydroethidium staining. In line, intracellular phospho-histone H2AX flow cytometry and comet assay revealed increased DNA damage indicating that ETV6/RUNX1 expression enhances ROS. On the basis of our data, we propose the following model: the expression of ETV6/RUNX1 creates a preleukemic clone and leads to increased ROS levels. These elevated ROS favor the accumulation of secondary hits by increasing genetic instability and double-strand breaks, thus allowing preleukemic clones to develop into fully transformed leukemic cells.
    Neoplasia (New York, N.Y.) 11/2013; 15(11):1292-300. DOI:10.1593/neo.131310 · 4.25 Impact Factor
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    • "The original AE cell lines, derived from 4 different cord blood donors, are designated AE30, AE31, AE33, and AE42. The cell lines derived from AE30 and AE31 cultures that are described here were transduced with a retrovirus containing the HPV E6/E7 viral oncogenes (a gift from Dr. Susanne Wells and Dr. Denise Galloway [38]), the pBABEneo-p53 DD plasmid (which has a deletion mutation of mouse p53 that acts in a dominant fashion and transforms human cells [39] [40]), a p53 specific shRNA, or if not otherwise specified, either a scrambled shRNA or the empty LXSN vector [22] [38]. Knockdown of p53 levels was confirmed by Western Blot. "
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    ABSTRACT: The mutation rate (μ) is likely to be a key parameter in leukemogenesis, but historically, it has been difficult to measure in humans. The PIG-A gene has some advantages for the detection of spontaneous mutations because it is X-linked, and therefore only one mutation is required to disrupt its function. Furthermore, the PIG-A-null phenotype is readily detected by flow cytometry. Using PIG-A, we have now provided the first in vitro measurement of μ in myeloid cells, using cultures of CD34+ cells that are transduced with either the AML-ETO or the MLL-AF9 fusion genes and expanded with cytokines. For the AML-ETO cultures, the median μ value was ∼ 9.4×10(-7) (range ∼ 3.6 to 23×10(-7)) per cell division. In contrast, few spontaneous mutations were observed in the MLL-AF9 cultures. Knockdown of p53 or introduction of mutant NRAS or FLT3 alleles did not have much of an effect on μ. Based on these data, we provide a model to predict whether hypermutability must occur in the process of leukemogenesis.
    Mutation Research/Fundamental and Molecular Mechanisms of Mutagenesis 06/2013; 749(1-2). DOI:10.1016/j.mrfmmm.2013.05.004 · 3.68 Impact Factor
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    • "p53, a well characterized human tumor suppressor gene involved in genotoxic stress response and DNA damage repair, also participates in autophagy regulation[30]. Intriguingly, the role of p53 in autophagy seems to be paradoxical depending on its Subcellular localization, which may dictate whether p53 contributes to cancer cell survival or death[31]. In the nucleus, p53 can activate AMPK to inhibit mTOR and induce the autophagic process. "
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    Chinese journal of cancer 07/2012; 32(3). DOI:10.5732/cjc.012.10010 · 2.16 Impact Factor
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