Abnormal CD34 Positive Cells Are Present in NPM+ Acute Myeloid Leukemia (AML) Samples at Diagnosis.

Nucleophosmin (NPM) gene mutations are the molecular hallmark of a large primary AML subgroup of patients. NPM mutated cases are commonly associated with FLT3-internal tandem duplications which seem to confer an adverse prognostic meaning. Blasts from NPM+ leukemias have been described as CD34 negative.

Objective: To assess the presence of CD34 positive populations and their phenotypic profile in NPM mutated samples at diagnosis. To investigate whether there are differences between NPM/FLT3- and those with an associated FLT3-ITD.

Patients and methods: Cells from forty-three bone marrow de novo NPM+ AML (26 females/ 17 males) patients enrolled in AML-CETLAM protocols were analyzed at diagnosis by means of multiparameter flow cytometry (MFC). In 18 cases an associated FLT3-ITD was detected (NPM+/FLT3+ group). Immature antigenic markers were investigated by triple combinations of direct conjugated MoAbs to CD15, CD34 and HLA-Dr ; CD34, CD123 and HLA-Dr; CD34, CD33 and CD19; CD15, CD117 and CD45. Minimal residual disease studies (7 cases) were performed in samples in complete remission using flow cytometry and real-time PCR (ABIPRISM 7700).

Results: In all the patients we were able to detect an abnormal CD34 population. CD15 coexpression, abnormal light scatter patterns, CD117 clusterization, antigenic overexpression and abnormal CD123 expression were all detected in these cases. NPM+/FLT3+ patients had 2,24% CD34+ cells(0,12%–42%) whereas for NPM+/FLT3- cases the mean CD34+ value was 1,05% (0,1–92%)(n.s.). Sequential samples to test in parallel the abnormal immunophenotype and NPM transcripts by real-time PCR were available in 7 cases (5 NPM+/FLT3+ and 2 NPM+/FLT3-). Relapses were observed in two NPM+/FLT3+ cases (4% and 42% CD34+ cells at diagnosis) and 1 NPM+/FLT3- patient (0.1% CD34+ cells at diagnosis). No definite pattern of relapse could be established given that in some cases NPM rises preceded the detection of phenotypically abnormal cells.

Conclusions: NPM mutated AML patients may be mostly categorized as CD34 negative following standard flow-cytometry positivity thresholds. Nevertheless, the constant detection of immature abnormal cells (CD34+CD123+CD117++) may help to identify the leukemic stem-cell pool in NPM+ and FLT3+ leukemias. It remains to be investigated whether the size of this compartment has prognostic relevance in some groups of normal karyotype AML.