Nucleophosmin (NPM1) mutations in exon 12 are the most common genetic alternation in cytogenetically normal AML (CN-AML). Although mutation types A, B, and D represent the majority of cases, rare mutation variants of the NPM1 gene in individual patients do occur. In this study, we have evaluated a novel, DNAbased real-time quantitative polymerase chain reaction (RQ-PCR) for the detection of three of the most commonly occurring mutations and for six rare patient-specific mutation types, which represent 28% of all of the NPM1 mutations in our group of 25 CN-AML patients. Furthermore, the prognostic relevance of NPM1-based monitoring of minimal residual disease (MRD) in peripheral blood (PB), bone marrow (BM), and in specific cell subsets (CD34 1 , CD34 2 , CD34 dim ) of BM were evaluated. In 80% of the evaluable patients, a molecular relapse preceded a hematological relapse. Moreover, in this subset of patients, the molecular relapse occurred at a median of 97 days before the hematological relapse. Our compartment analysis showed a strong correlation between BM and PB (r 5 0.907, P < 0.001) as well as a high copy number of mutated NPM1 in CD341 BM cells.In conclusion, we have demonstrated applicability of our presented RQ-PCR method for a large percentage of mutated NPM1 patients with CN-AML as well as the usefulness for long-term follow-up monitoring of MRD and the prediction of hematological relapse. Am. J. Hematol. 85:926-929, 2010. V V C 2010 Wiley-Liss, Inc.
IntroductionThe Nucleophosmin (NPM1) gene is one of the most commonly mutated genes in acute myeloid leukemia (AML), occurring in approximately 35% of AML patients at the time of diagnosis and in approximately 60% of adult cytogenetically normal AML (CN-AML). On the basis of its unique molecular, genotypic, immunophenotypic, and prognostic features, AML with an NPM1 mutation was included as a separate provisional entity in the 2008 World Health Organization classification of myeloid neoplasms [1].Somatically acquired mutations in the NPM1 gene usually occur in exon 12, which encodes the COOH-terminal region, and in most cases results in the net insertion of four base pairs. More than 50 molecular variants of mutant NPM1 have been identified to date [2]. The most common variant, Type A, represents approximately 80% of all cases, and Types B and D account for 10% and 5% of cases, respectively. Other mutations rarely occur and vary from case to case. NPM1 mutations appear to be stable, since in the majority of patients, the same mutation is detectable at the time of diagnosis and at relapse. Molecular monitoring using real-time quantitative polymerase chain reaction (RQ-PCR) in a group of CN-AML patients in complete remission has the potential for detecting subclinical levels of residual disease to allow for early treatment intervention and prevent an overt hematological relapse.We developed DNA-based RQ-PCR with mutation-specific reverse primers and a common minor groove binding (MGB) probe for nine different types of NPM1 mutations. Therefore, we can moni...