Rapid detection of nucleophosmin (NPM1) mutations in acute myeloid leukemia by denaturing HPLC.

Application of the erythrocyte glutathione reductase assay in evaluating riboflavin nutritional status in a high school student population. Riboflavin, flavin mononucleotide, and flavin adenine dinucleotide in human plasma and erythrocytes at baseline and after low-dose riboflavin supple-mentation. Recovery from impaired dark adaptation in nightblind pregnant Nepali women who receive small daily doses of vitamin A as amaranth leaves, carrots, goat liver, vitamin A-fortified rice, or retinyl palmitate. during the course of pregnancy of rural and urban women in northeast Thailand. et al. Riboflavin, folate and vitamin C status of Gambian women during pregnancy: a comparison between urban and rural communities. blood cell glutathione reductase and pyridoxine phosphate oxidase activities not related to dietary riboflavin: selection by malaria? Nucleophosmin (NPM1) is a multifunctional, highly conserved protein found most frequently in nucleoli. NPM1 acts as a molecular chaperone (1) and is thought to participate in preribosome maturation and centrosome duplication (2); in addition, it has been implicated in the regulation of the ArF-p53 tumor suppressor pathway (3, 4). NPM1 mutations have recently been reported to occur at high frequency in acute myeloid leukemia (AML), for which they currently represent the most common detectable genetic lesion (ϳ35% of cases). This abnormality is strongly associated with normal-karyotype AML and has never been detected in AMLs bearing major cytogenetic abnormalities. It has not been observed in other hemato-poietic tumors (5). Two main types of mutations have been described to date. The first and most frequent consists of a 4-nucleotide (nt) insertion (YWTG; YUPAC code) downstream from nucleotide 959; the second is deletion of a GGAGG sequence at positions 965 through 969 and substitution with 9 extra nt (GenBank accession no. NM_002520). Both mutations lead to aberrant cyto-plasmic localization of NPM1 as shown after immuno-staining with anti-NPM1 monoclonal antibodies. In addition to their high frequency and clustering with normal karyotype, NPM1 mutations may identify a subset of AMLs with distinct response to therapy (5). Together, these findings may have repercussions in AML classification and suggest that analysis of NPM1 mutational status should integrate modern genetic characterization of AML. We describe here a rapid and reproducible method for screening NPM1 mutations by reverse transcription-PCR followed by denaturing HPLC (DHPLC). Bone marrow samples showing at least 70% bone marrow infiltration by leukemic cells were collected at diagnosis from 56 patients with newly diagnosed AML observed at the Department of Biopathology at the University Tor Vergata (Rome). According to …