Rapid mutational analysis of N-ras proto-oncogene in hematologic malignancies: study of 77 Greek patient

• Published in: Haematologica (Roma) Vol. 86; no. 9; pp. 918 - 927
• Main Authors: Speletas, M, Arvanitidi, K, Tzoanopoulos, D, Tsironidou, V, Pardali, E, Aggeli, C, Tsapogas, P, Kartalis, G, Sideras, P, Ritis, K
• Format: Journal Article
• Language: English
• Published: Italy 01.09.2001
• Subjects: Adolescent; Adult; Aged; Child; DNA Mutational Analysis - methods; Endoribonucleases - metabolism; Genes, ras - genetics; Greece; Hematologic Neoplasms - genetics; Humans; Middle Aged; Mutation; Polymorphism, Genetic; Reverse Transcriptase Polymerase Chain Reaction - standards; Time Factors; Greece
• ISSN: 0390-6078; 1592-8721

First Department of Internal medicine, Democritus University of Thrace, Alexandroupolis, Greece. speletas@otenet.gr BACKGROUND AND OBJECTIVES: N-ras mutations are the most commonly detected molecular abnormalities in hematologic malignancies, especially in those of myeloid origin. Different techniques have been used to detect N-ras mutations; however, most of them are either labor intensive or provide sequence data for only a limited number of codons. Consequently, study of the N-ras oncogene has not been convenient in every day clinical practice being restricted, as a rule, to retrospective analysis of patients. DESIGN AND METHODS: In this study we used a recently developed method that enables rapid and reliable detection of mutations at the cDNA level, namely, the non-isotopic RNase cleavage assay (NIRCA). Using this method we were able to screen the N-ras oncogene rapidly and determine the incidence and prognostic significance of N-ras mutations in 77 Greek patients with acute leukemia, myelodysplastic syndromes and chronic myeloproliferative disorders, both at the presentation and during relapse or progression of the disease. RESULTS: Activating N-ras mutations were detected in 7 patients and our results were confirmed by direct sequencing. Interestingly, two novel alterations were identified, a mutation at codon 8 (characterized by a substitution of valine by leucine) in a patient with chronic myeloid leukemia during hematologic relapse of the disease and a polymorphism at codon 92 (1002T-->C, without amino acid substitution) in a patient with chronic myelomonocytic leukemia. INTERPRETATION AND CONCLUSIONS: A rapid and easy protocol that allows the analyses of N-ras sequences has been developed. This reverse transcription-polymerase chain reaction (RT-PCR)/NIRCA protocol can allow the study of this proto-oncogene in every day clinical practice, rapidly facilitating the validation of the diagnostic and prognostic value of N-ras mutational analyses in patients with hematologic malignancies.