CLINICS 2011;66(5):793-799 CLINICAL SCIENCE DOI:10.1590/S1807-59322011000500014 Screening for hotspot mutations in PI3K, JAK2, FLT3 and NPM1 in patients with myelodysplastic syndromes Joa˜ o Agostinho Machado-Neto, Fabiola Traina, Mariana Lazarini, Paula de Melo Campos, Katia Borgia Barbosa Pagnano, Irene Lorand-Metze, Fernando Ferreira Costa, Sara T Olalla Saad Hematology and Hemotherapy Center, National Institute of Blood, University of Campinas, S a˜o Paulo, Brazil. INTRODUCTION: Myelodysplastic syndromes encompass a heterogeneous group of clonal hematopoietic stem cell disorders characterized by ineffective hematopoiesis, refractory cytopenia and a tendency to progress toward acute myeloid leukemia. The accumulation of genetic alterations is closely associated with the progression of myelodysplastic syndromes toward acute myeloid leukemia. OBJECTIVE: To investigate the presence of mutations in the points most frequent for mutations (hotspot mutations) in phosphatidylinositol-3-kinase (PI3K), Janus kinase 2 (JAK2), FMS-like tyrosine kinase 3 (FLT3) and nucleophosmin (NPM1), which are involved in leukemia and other cancers, in a population of Brazilian MDS patients. METHODS: Fifty-one myelodysplastic syndromes patients were included in the study. According to French-American- British classification, the patients were distributed as follows: 31 with refractory anemia, 8 with refractory anemia with ringed sideroblasts, 7 with refractory anemia with excess blasts, 3 with refractory anemia with excess blasts in transformation and 2 with chronic myelomonocytic leukemia. Bone marrow samples were obtained and screened for the presence of hotspot mutations using analysis based on amplification with the polymerase chain reaction, sequencing, fragment size polymorphisms or restriction enzyme digestion. All patients were screened for mutations at the time of diagnosis, and 5 patients were also screened at the time of disease progression. RESULTS: In the genes studied, no mutations were detected in the patients at the time of diagnosis. One patient with chronic myelomonocytic leukemia was heterozygous for a Janus kinase 2 mutation after disease progression. CONCLUSIONS: These results show that hotspot mutations in the PI3K, JAK2, FLT3 and NPM1 genes are not common in MDS patients; nevertheless, JAK2 mutations may be present in myelodysplasia during disease progression. KEYWORDS: Hematopoietic Disorder; Acute Leukemia; Myelodysplasia; Mutations; Bone Marrow. Machado-Neto JA, Traina F, Lazarini M, Campos PM, Pagnano KBB, Lorand-Metze I, et al. Screening for hotspot mutations in PI3K, JAK2, FLT3 and NPM1 in patients with myelodysplastic syndromes. Clinics. 2011;66(5):793-799. Received for publication on November 30, 2010; First review completed on January 4, 2011; Accepted for publication on February 11, 2011 E-mail: sara@unicamp.br Tel.: 55 19 3289-1089 INTRODUCTION Myelodysplastic syndromes (MDS) encompass a hetero- geneous group of clonal hematopoietic stem cell disorders characterized by ineffective hematopoiesis, refractory cytope- nia and a tendency to progress to acute myeloid leukemia (AML).1 Low-risk MDS present high levels of intramedullar apoptosis, whereas high-risk MDS show a decrease in apop- tosis, an increase in cell proliferation and a high frequency of evolution to AML.2,3 The accumulation of genetic alterations is closely associated with the progression of MDS toward AML, and efforts are being made to determine the significance of Copyright 2011 CLINICS – This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http:// creativecommons.org/licenses/by-nc/3.0/) which permits unrestricted non- commercial use, distribution, and reproduction in any medium, provided the original work is properly cited. various genetic aberrations in patients with MDS.4-6 The same occurs for liver adenomatosis,7 Rubinstein-taybi syndrome8 and hemochromatosis.9 The Phosphatidylinositol-3-kinase (PI3K) and Janus kinase 2 (JAK2) signaling pathways are involved in numerous cellular processes, such as proliferation, apopto- sis and differentiation.10-12 Mutations in the catalytic subunit of PI3K are frequently observed in several cancers, including AML.4,5,13 Hotspot mutations occur in exon 9 (E542 and E545) and in exon 20 (H1047), resulting in increased PI3K/Akt activity.5,14-17 One somatic mutation in the JAK2 gene (V617F) has been identified in myeloproli- ferative disorders such as polycythemia vera (PV) and myelofibrosis.18 FMS-like tyrosine kinase 3 (FLT3) is a tyrosine kinase re- ceptor that plays an important role in the proliferation and dif- ferentiation of hematopoietic progenitors.19 Nucleophosmin (NPM1) is a key regulator of hematopoesis that shuttles 793