Molecular Genetic study of the Frequency of DNA Methyl Transferase 3A and Nucleophosmin 1 Mutations in adult Denovo Acute Myeloid Leukemia

Abstract

Background: DNMT3A, FLT3, and NPM1Amutations are among the most common genomic alterations in de novo acute myeloid leukemia (AML) and play a key role in thepathogenesis and evolution of the disease, particularly in the absence of AML-associated recurrent cytogenetic abnormalities. DNA methyltransferase 3A (DNMT3A) mutations were considered to be independently associated with unfavorable prognosis in adults with de novo acute myeloid leukemia (AML). Mutations resulting in constitutive FLT3 activation include internal tandem duplication (FLT3-ITD) mutation, which is a known adverse prognostic indicator in AML patients.NPM1 mutations are generally associated with more favorable outcomes in cytogenetically normal (CN)-AML. Methods:Genomic DNA from 123 de novo AML patients treated at National Cancer Institude (NCI), Cairo University were amplified by Polymerase Chain Reaction (PCR) to detect FLT3-ITD mutation. Allele Specific Polymerase Chain Reaction (AS-PCR) analysis for NPM1A and DNMT3A mutations then followed by fragment analysis of post-PCR products using GeneMapper software and Direct Sequencing technique for the detection of NPM1A andDNMT3A mutations respectively. Results: DNMT3A,FLT3-ITD, and NPM1A, gene mutations were detected in22 (17.9%), 22 (17.9%), and 24(19.5%), patients respectively. DNMT3A/FLT3 combined mutant genotypes were detected in 5 (4.1%) patients, while NPM1A/FLT3 combined mutant genotypes were detected in 9 (7.3%) cases. DNMT3A/NPM1A combinated mutant genotypeswere detected in 3 (2.4%) cases. Two (1.6%) cases have triple mutant genotypes (DNMT3A/FLT3/NPM1A) and 70 (56.9%) have triple wild genotype. The presence of FLT3-ITD mutation was significantly associated with older age (P= 0.029). The presence of NPM1Amutation was associated with a higher overall survival (OS), however the difference did not reach the level of significance. The presence of FLT3-ITD mutation was significantly associated with a lower OS (P= 0.046), also the presence ofDNMT3A mutation was associated with a lower OS, however the difference did not reach the level of significance. The presence of DNMT3A/FLT3 combined mutations were significantly associated with a lower OS (P= 0.016). The presence of FLT3-ITD mutation and DNMT3A mutation was significantly associated with a lower complete remission (CR) rate (P= 0.016 & P=0.016 respectively). Combination of NPM1A/FLT3, DNMT3/FLT3, and NPM1A/DNMT3A mutations were significantly associated with a lower CR rate (P= 0.006, P=0.006 & P=0.023 respectively). Conclusion: DNMT3A, FLT3-ITD, andNPM1A are frequent mutations in Egyptian AML. There is asignificant association between FLT3-ITD mutation in older age.The presence of NPM1A mutation is tendency toward a good prognosis but the presence of FLT3-ITD, and DNMT3A mutations are tendency toward a worse prognosis. Analysis forDNMT3A, FLT3-ITD, andNPM1A mutations should be considered as part of the routine work-up of patients with AML at diagnosis for a better designing of risk-adapted therapy. Keywords: AML, NPM1, FLT3, ITD,DNMT3A, prognosis, AS-PCR