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

Background and Objectives. N-ras mutations are the most commonly detected m olecular abnormalities in hematologic malignancies, especially in those of myeloid origin. Different techniques have been used to detect N-ras mutatio ns; however, most of them are either labor intensive or provide sequence da ta for only a limited number of codons. Consequently, study of the N-ras on cogene has not been convenient in every day clinical practice being restric ted, 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 p rognostic significance of N-ras mutations in 77 Greek patients with acute l eukemia; myelodysplastic syndromes and chronic myeloproliferative disorders , both. at presentation and during relapse or progression of the disease. Results. Activating N-ras mutations were detected in 7 patients and our res ults were confirmed by direct sequencing. Interestingly, two novel alterati ons were identified, a mutation at codon 8 (characterized by a substitution of valine by leucine) in a patient with chronic myeloid leukemia during he matologic 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 a nalysis of N-ras sequences has been developed. This reverse transcription-p olymerase chain reaction (RT-PCR)/NIRCA protocol can allow the study of thi s proto-oncogene in every day clinical practice; rapidly facilitating the v alidation of the diagnostic and prognostic value of N-ras mutational analys es in patients with hematologic malignancies. (C) 2001, Ferrata Storti Foun dation.