Detection of FLT3 Oncogene Mutations in Acute Myeloid Leukemia Using Conformation Sensitive Gel Electrophoresis

Mamdooh Gari,Sahira Lary,Adeel Chaudhary,Waseem Ahmad,Adel Abuzenadah,Fatin Al-Sayes,Huda Banni,Mohammed Al-Qahtani,Ghazi Damanhouri

doi:10.3390/ijms9112194

Abstract

FLT3 (fms-related tyrosine kinase 3) is a receptor tyrosine kinase class III that is expressed on by early hematopoietic progenitor cells and plays an important role in hematopoietic stem cell proliferation, differentiation and survival. FLT3 is also expressed on leukemia blasts in most cases of acute myeloid leukemia (AML). In order to determine the frequency of FLT3 oncogene mutations, we analyzed genomic DNA of adult de novo acute myeloid leukemia (AML). Polymerase chain reaction (PCR) and conformation-sensitive gel electrophoresis (CSGE) were used for FLT3 exons 11, 14, and 15, followed by direct DNA sequencing. Two different types of functionally important FLT 3 mutations have been identified. Those mutations were unique to patients with inv(16), t(15:17) or t(8;21) and comprised fifteen cases with internal tandem duplication (ITD) mutation in the juxtamembrane domain and eleven cases with point mutation (exon 20, Asp835Tyr). The high frequency of the flt3 proto-oncogene mutations in acute myeloid leukemia AML suggests a key role for the receptor function. The association of FLT3 mutations with chromosomal abnormalities invites speculation as to the link between these two changes in the pathogenesis of acute myeloid leukemiaAML. Furthermore, CSGE method has shown to be a rapid and sensitive screening method for detection of nucleotide alteration in FLT3 gene. Finally, this study reports, for the first time in Saudi Arabia, mutations in the human FLT3 gene in acute myeloid leukemia AML patients.

Highlights

IntroductionThe FLT3 gene (fms-like tyrosine kinase), is a member of the class III tyrosine kinase receptor family that includes c-kit, c-fms and the platelet-derived growth factor receptors (PDGFRs) [1,2,3]
The FLT3 gene, is a member of the class III tyrosine kinase receptor family that includes c-kit, c-fms and the platelet-derived growth factor receptors (PDGFRs) [1,2,3].Class III receptor tyrosine kinases (RTKs) share sequence homology and have a similar overall structure, with five immunoglobulin-like repeats in the extracellular domain, a single transmembrane domain (TM), a juxtamembrane domain (JM), two intracellular tyrosine kinase domains (TK1 and TK2) divided by a kinase insert domain (KI), and a C-terminal domain [4]
In order to screen for the FLT3-internal tandem duplication (ITD) mutation, exons 14 and 15 of the FLT3 gene were amplified from genomic DNA of 129 acute myeloid leukemia (AML) patients using Polymerase chain reaction (PCR), followed by conformation sensitive gel electrophoresis (CSGE) analysis

Summary

Introduction

The FLT3 gene (fms-like tyrosine kinase), is a member of the class III tyrosine kinase receptor family that includes c-kit, c-fms and the platelet-derived growth factor receptors (PDGFRs) [1,2,3]. Class III receptor tyrosine kinases (RTKs) share sequence homology and have a similar overall structure, with five immunoglobulin-like repeats in the extracellular domain, a single transmembrane domain (TM), a juxtamembrane domain (JM), two intracellular tyrosine kinase domains (TK1 and TK2) divided by a kinase insert domain (KI), and a C-terminal domain [4]. Data from different sources suggest a pathogenic role of FLT3 in acute myeloid leukemia (AML). Functional analysis of FLT3 expression has been documented in many cases of AML, besides of immortalized human myeloid and monocytic cell lines [3, 6,7,8].

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