EVI1 Inhibits Apoptosis Induced by Antileukemic Drugs via Upregulation of CDKN1A/p21/WAF in Human Myeloid Cells

Anna Rommer,Kadin Karakaya,Ilse Schwarzinger,Katharina Hartl,Birgit Steinmetz,Martin Filipits,Hanno Glimm,Daniela Heilos,Friederike Herbst,Hubert Hackl,Shayda Hemmati,Petra Heffeter,Irene Herbacek,Rotraud Wieser,Alwin Krämer,Katarina Steinleitner,Walter Berger,Pieter Rondou,Ivan Cruz Moura

doi:10.1371/journal.pone.0056308

Abstract

Overexpression of ecotropic viral integration site 1 (EVI1) is associated with aggressive disease in acute myeloid leukemia (AML). Despite of its clinical importance, little is known about the mechanism through which EVI1 confers resistance to antileukemic drugs. Here, we show that a human myeloid cell line constitutively overexpressing EVI1 after infection with a retroviral vector (U937_EVI1) was partially resistant to etoposide and daunorubicin as compared to empty vector infected control cells (U937_vec). Similarly, inducible expression of EVI1 in HL-60 cells decreased their sensitivity to daunorubicin. Gene expression microarray analyses of U937_EVI1 and U937_vec cells cultured in the absence or presence of etoposide showed that 77 and 419 genes were regulated by EVI1 and etoposide, respectively. Notably, mRNA levels of 26 of these genes were altered by both stimuli, indicating that EVI1 regulated genes were strongly enriched among etoposide regulated genes and vice versa. One of the genes that were induced by both EVI1 and etoposide was CDKN1A/p21/WAF, which in addition to its function as a cell cycle regulator plays an important role in conferring chemotherapy resistance in various tumor types. Indeed, overexpression of CDKN1A in U937 cells mimicked the phenotype of EVI1 overexpression, similarly conferring partial resistance to antileukemic drugs.

Highlights

Aberrant expression of the ecotropic viral integration site 1 (EVI1) gene, which in healthy individuals is transcribed in hematopoietic stem and progenitor cells but not in mature blood cells [1,2,3,4], is associated with a poor prognosis in myeloid leukemias [2,5,6,7,8,9,10,11,12,13]
In the present report we describe an in vitro model that recapitulates the resistance of EVI1 overexpressing cells to drugs used in the chemotherapy of acute myeloid leukemia (AML), and show that this effect may in part be mediated by upregulation of CDKN1A/p21/WAF
In order to establish an in vitro model for human myeloid leukemias constitutively overexpressing EVI1, the human myeloid cell line U937 was infected with a retroviral vector containing the full length human EVI1 cDNA, or with empty vector as a control

Summary

Introduction

Aberrant expression of the ecotropic viral integration site 1 (EVI1) gene, which in healthy individuals is transcribed in hematopoietic stem and progenitor cells but not in mature blood cells [1,2,3,4], is associated with a poor prognosis in myeloid leukemias [2,5,6,7,8,9,10,11,12,13]. EVI1 overexpression correlates with shorter survival in some solid tumors like ovarian carcinoma [14] and estrogen receptor negative breast cancer [15]. The phosphatase and tensin homolog (PTEN) gene appears to be of particular interest, as direct repression of PTEN by EVI1 lead to activation of the AKT/ mTOR pathway in murine bone marrow cells, and rapamycin prolonged survival of mice with EVI1 overexpressing leukemias [17]. Recent results indicate that the opposite may be true [20,21,22], so that it is presently unclear to which extent activation of the AKT pathway can explain the poor prognosis associated with EVI1 overexpression in AML

Methods

Results

Conclusion