DNMT3A Mutations in Tunisian Patients with Acute Myeloid Leukemia

Special Article - Myeloid Leukemia

J Blood Disord. 2015; 2(3): 1032.

DNMT3A Mutations in Tunisian Patients with Acute Myeloid Leukemia

Mechaal Amal¹*, Safra Ines¹, Ben Neji Hind², Rezgui Ichraf¹, Menif Samia¹, Fouzai Chaker¹, Meddeb Balkis² and Abbes Salem¹

1Dpartment of Molecular and Cellular Hematology, University of Tunis El Manar, Tunisia

2Aziza Othmana Hospital, Tunisia

*Corresponding author: Mechaal A, Department of Molecular and Cellular Hematology, Pasteur Institute of Tunis, University of Tunis El Manar, Tunisia

Received: September 14, 2015; Accepted: October 21, 2015; Published: October 28, 2015


DNA Methyl Transferase 3A (DNMT3A) is one of two human de novo DNA methyltransferases essential for the regulation of gene expression. DNMT3A mutations were recently linked to hematologic malignancies prognosis. In fact, numerous mutations in this gene were reported in patients with Acute Myeloid Leukaemia (AML), pointing DNMT3A as an important oncogenic role in AML patients. In the present study, 84 patients were analyzed, at the first diagnosis for DNMT3A mutations. Exons 18, 19, 20, 21, 22 and 23 were screened by Polymerase Chain Reaction (PCR) and direct sequencing. The results demonstrated that 33.33% (28/84) de novo AML patients presented DNMT3A mutations. 9 missense mutations including 7 novel single nucleotide polymorphism resulting in amino acid substitution, one silence mutation and 1 nonsense mutation. These mutations are associated with an intermediate-risk cytogenetics (Normal Karyotype (KN-AML)), younger age, higher WBC count, bone marrow infiltration at diagnosis and lower plated count. In conclusion, we retain that the DNMT3A gene is highly mutated in the AML subgroup, its role as a prognostic factor needs to be further elucidated by correlation studies with other molecular prognosis factors and survival.

Keywords: Acute myeloid leukemia; PCR and Direct sequencing; DNMT3A gene


A: Alanine; AML: Acute Myeloid Leukaemia; bp: Base Pair; C: Cysteine; D: Acide Aspartique; DNA: Desoxyribo Nucleic Acid; DNMT3A: DNA Methyl Transferase 3A; E: Acide Glutamique; F: Phenylalanine; FAB: Franco-Américano-Britannique; g/dl: Grams Per Deciliter; H: Histidine; HSCs: Hematopoietic Stem Cells; I: Isoleucine; K: Lysine; L: Leucine; L: Liter; M: Methionine; N: Asparagines; KN: Normal Karyotype; P: Proline; PCR: Polymerase Chain Reaction; R: Arginine; RNA: Ribo Nucleic Acid; SPSS: Statistical Package for the Social Sciences; V: Valine; WBC : White Cell Count


Acute Myeloid Leukemia (AML) is a malignancy of Hematopoietic Stem Cells (HSCs) characterized by the expansion of undifferentiated myeloid progenitors (blasts) with great variability in clinical course and response to therapy. The development of AML is associated with accumulation of acquired genetic alterations and epigenetic changes in hematopoietic progenitor cells that alter normal mechanisms of cell growth, proliferation, and differentiation [1-6]. DNA methylation plays a key role in the pathophysiology of AML [7-10].

DNA Methyltransferase 3A (DNMT3A) is one of two human de novo DNA methyltransferases essential for regulating gene expression during cellular development and differentiation [11]. DNMT3A mutations are generally present in the clones of AML samples, suggesting that they may initiate leukemia [5,6,9,12]. However, the mechanisms by which they contribute to leukemogenesis are not yet clear. Analysis of mutated DNMT3A gene has an important clinical and pathologic application. DNMT3A mutations may become a new tool for target therapy [13, 14].

DNMT3A is located in chromosomal band 2p23 and, is a member of a DNA Methyltransferase (Mtase) family [15]. It has three conserved domains: the PWWP (a highly conserved prolinetryptophan- tryptophan-proline motif) domain targeting the enzyme to nucleic acids, the cystein-rich PHD zinc-finger domain interacting with unmodified histone H3, and the highly conserved catalytic domain representing the Mtase domain in the c-terminal region. DNMT3A has high ubiquitous expression in embryonic tissues and undifferentiated embryonic stem cells. By catalyzing the conversion of cytosine to 5- methylcytosine, DNMT3A add methyl groups to unmodified DNA. DNMT3A mutations, most commonly are heterozygous, and almost at codon R882 in exon 23 DNMT3A Mtase domain modify its enzymatic activity. DNMT3A mutations are significantly enriched in patients with intermediate risk cytogenetics with a normal caryotype [14,16-18].

This work was designed to study the prevalence and nature of the DNMT3A gene mutations in novo AML patients in a Tunisian group.

Patients and Methods

Patients and Bone Marrow Mononuclear Cell (BMMC) collection

We have studied 84 samples of bone marrow from patients with AML, at diagnosis and prior to any chemotherapy. Patients’ samples had been collected from January 2014 to August 2015. Bone marrow mononuclear cells were isolated with Ficoll gradient separation.

The patients’ clinical information’s were obtained from the AML database of different Tunisian hospitals retrospectively. Patients were stratified according to the FAB (Frensh American British) classification.

The diagnosis of the AML subgroup was based on the standard clinic morphologic and phenotype criteria. Cytogenetic information was available for only 78 patients (Table 1).