Acute Myeloid Leukemia with Normal Karyotype: Analysis of FLT3, NPM1, MLL and CEBPA Gene Abnormalities.

Acute myeloid leukemia with normal karyotype (AML-NK) is markedly heterogeneous depending on the presence or absence of certain genetic alterations. We studied the prevalence of different abnormalities, namely of the FLT3, NPM1, CEBPA and MLL genes, in AML-NK patients and tried to correlate them with clinicopathological findings. Techniques used were as follows: FLT3-ITD (internal tandem duplications), PCR using fluorescent primers, analysis in automatic sequencer; FLT3-TKD (D385), PCR, fragment cut with EcoRV restriction enzyme run in agarose gel; NPM1, RT-PCR and automatic sequencing of a fragment containing exon 12; CEBPA, PCR and automatic sequencing of the entire coding region; MLL-PTD, RT-PCR with analysis in agarose gel. The following data from patients were recorded: sex, age, FAB classification, WBC and platelet count, hemoglobin and LDH level, response to induction, survival time and cause of death. The incidence of FLT3 alterations (all ITDs) was 10/23 (43.5%), varying from 1 to 10 ITDs; NPM1 was mutated in 12/24 cases (50.0%) and MLL-PTD in 2/24 patients (8.3%); for CEBPA, of the 10 cases in which all 3 amplicons (1A, 2B, 3C) were analysed, 6 had a mutation mostly in amplicon 2B (5 cases). More than one genetic abnormality was present in 9 cases: six showed coexistence of FLT3-ITD and NPM1 mutation, one of MLL and CEBPA mutations, one of FLT3-ITD and CEBPA mutation and one of FLT3-ITD, NPM1 and CEBPA. Regardind correlation with clinical characteristics, we observed that 72.7% of nucleophosmin mutated (NPM1+) cases were CD34 negative and 91.7% of CD34+ patients were NPM1 negative (p=0.003). There is a tendency, although not significant, for CEBPA+ cases to correlate with low WBC counts and for patients with high WBC count to be NPM1+ and FLT3-ITD+ (odds ratio of 3.0 and 3.33, respectively). All NPM1+ patients treated with appropriate induction therapy achieved complete remission; 22.2% of NPM1 negative patients were refractory to therapy. No significant correlation was observed between survival and presence or absence of any of the genetic alterations. This work was funded by an unrestricted grant from Roche Farmaceutica - Portugal.