Identification of FLT3 and NPM1 Mutations in Patients with Acute Myeloid Leukaemia

Objective: The most frequent acquired molecular abnormalities and important prognostic indicators in patients
with Acute Myeloid Leukaemia (AML) are fms-like tyrosine kinase-3 gene (FLT3) and nucleophosmin-1 (NPM1)
mutations. Our study aims to develop a cost effective and comprehensive in-house conventional PCR method for
detection of FLT3-ITD, FLT3-D835 and NPM1 mutations and to evaluate the frequency of these mutations in patients
with cytogenetically normal (CN) AML in our population. Methods: A total of 199 samples from AML patients (95
women, 104 men) were included in the study. Mutation analyses were performed using polymerase chain reaction
(PCR) and gene sequencing. Result: Sixty-eight patients were positive for the mutations. FLT3-ITD mutations were
detected in 32 patients (16.1%), followed by FLT3-D835 in 5 (2.5%) and NPM1 in 54 (27.1%). Double mutations of
NPM1 and FLT3-ITD were detected in 23 cases (11.6%). Assays validation were performed using Sanger sequencing
and showed 100% concordance with in house method. Conclusion: The optimized in-house PCR assays for the
detection of FLT3-ITD, FLT3-D835 and NPM1 mutations in AML patients were robust, less labour intensive and cost
effective. These assays can be used as diagnostic tools for mutation detection in AML patients since identification of
these mutations are important for prognostication and optimization of patient care.