Absence of Cytogenetic Abnormalities of PRV-1, TPO and C-MPL Genes Detected by Fluorescence In Situ Hybridization in Essential Thrombocythemia.

Abstract

Introduction. Essential thrombocythemia (ET) is a chronic myeloproliferative disorder (CMPD) with heterogeneous features and no specific diagnostic markers. Consequently, its diagnosis is based on exclusion of other CMPD and secondary thrombocytosis. The role of PRV-1 (Polycythemia rubra vera-1), TPO (Thrombopoietin) and c-Mpl (Myeloproliferative leukemia virus oncogene) in ET pathogenesis has been studied in order to find new molecular targets which would help in ET diagnosis. PRV-1 gene is overexpressed in granulocytes from polycythemia vera (PV) and in some ET patients. TPO serum levels are not diagnostically useful and c-Mpl expression in megakaryocytes and platelets are generally decreased in ET. Mutations in TPO and c-MPL genes have been detected in familial thrombocythemia, but not in patients with acquired ET. The aim of the present study was to analyse PRV-1, TPO and c-MPL genes status by fluorescence in situ hybridization (FISH) technique in order to find new molecular markers in ET patients.Patients and Methods. Thirty bone marrow samples of ET patients (7M/23F) diagnozed by PVSG criteria with a normal karyotype and 10 bone marrow samples of normal healthy donors were included in the study. All samples were studied by three locus-specific probes for PRV-1, TPO and c-MPL genes as follows: 1. PRV-1 gene (BAC RP11-160A19, 157 Kb, located at 19q13.12-2) labeled in red cohybridized with the 19p telomeric probe (D19S238E, Vysis) labeled in green. 2. TPO gene (BAC RP11-45NP16, 183 Kb, located at 3q27) labeled in green cohybridized with the centromeric probe for chromosome 3 (D3Z1, Vysis) labeled in red. 3. c-MPL gene (BAC RP11-297L5, 190 Kb, located at 1p34) labeled in green cohybridized with the centromeric probe for chromosome 1 (D1Z5, Vysis) labeled in orange. A minimum of 100 interphase nuclei were analyzed.Results. FISH study showed no PRV-1, TPO and c-MPL cytogenetic abnormalities in any of the analyzed cases, except for one patient in which 21% of interphase nuclei presented a trisomy for the TPO gene region. The monosomy and trisomy thresholds were 6.1% and 4.7% for PRV-1, 4.9% and 3.4% for TPO, and 5.4% and 3.71% for c-MPL, respectively.Conclusions. Our results suggest a lack of structural and numerical rearrangements of PRV-1, TPO and c-MPL genes in ET patients. The PRV-1 gene FISH results are in line with the previously reported by Najfeld et al (Exp Hematol 2003; 31:118–21); regarding TPO and c-MPL results, this is the first FISH study reported in the literature in ET.