Conformational Sensitive Gel Electrophoresis (CSGE) as a method for NPM1 mutational screening in patients with Acute Myeloid Leukaemia

Abstract

IntroductionNucleophosmin gene (NPM1) exon 12 is the most common mutated gene found in Acute Myeloid Leukaemia (AML), which is present in 25–35%. NPM1 gene encodes for nucleophosmin (NPM) protein, which contains 294 amino acids. The NPM protein is an abundant nucleolar phosphoprotein that shuttles between the nucleus and cytoplasm. It contributes to promotion of ribosome biogenesis, regulation of centrosomal duplication during cell division, interaction with tumor suppressor gene and control of various nuclear proteins. ObjectiveTo screen for NPM1 exon 12 mutationsin AML patients diagnosed at Hospital Universiti Sains Malaysia (HUSM) using Conformational Sensitive Gel Electrophoresis (CSGE). MethodsTotal genomic DNAs were obtained from bone marrow aspirates or peripheral blood samples taken at diagnosis from 67 AML patients diagnosed at HUSM [French-American-British (FAB) subtypes: M0=1, M1=2, M2=10, M3=22, M4=9,M5=12,M6=2,M7=1, Not Otherwise Spesified (NOS)=8]. Patients were screened for NPM1 exon 12 mutations by CSGE. Cases displaying abnormal CSGE profiles were directly sequenced. Results & DiscussionMutation of NPM1exon 12 was present in 7/67 AML cases (10%) and all cases were females. They consisted of two M1, two M5b, one M2, one M3 and one NOS FAB subtypes. Most of the mutations identified as insertional (ins) mutation (5/7), 4 patients having ins 962–963 CTGG (2), CATG (1) and CTGT (1). One patient having ins at 969 CATG and followed with stop codon (TGA). One patient having single base deletion (del) at 966 and became TGA (stop codon) while another patient having ins at 956 TGGA and followed with del 958–969. ConclusionOur data showed high frequency of NPM1 in AML-M1 and M5b, which is highly associated in female patients. The CSGE was found to be an inexpensive, simple and reliable test to be used in screening of NPM1 mutations.