Molecular Variability of FLT3/ITD Mutants and Their Impact on the Differentiation Program of 32D Cells: Implications for the Biological Properties of AML Blasts

FLT3 is the most frequently mutated gene in AML – up to 40% of AML harbor an activating mutation within FLT3 gene. Though AML is a relatively rare disease, such a high mutability rate, as observed with FLT3 gene, is striking. To elucidate the molecular background of this phenomenon, we have established nine unique FLT3/ITD - carrying 32D cell lines and a set of controls, and subjected them to whole genome expression analysis and 2DE LC/MS proteomics. Data obtained on this so far largest set of ITD mutants indicates that FLT3/ITD positive cells exhibit less mature expression profiles resembling ST-HSC and MkEP/CMP/LMPP progenitors. FLT3/ITD-positive 32D cells overexpress a combination of genes of pluripotency (Etv5, endoglin, EpoR, Mpl, Trfr2, GATA.2, Hemgn, Myb, Dok2, Gli1, Pim3, PML, Ikaros, Pum1, Dlx4, Hox-c8, HoxB9, Runx1, c-Myc, Shh, FL) and early erythroid precursors (EpoR, Trfr2, GATA.1, Klf1), implicating their less mature expression profile than 32D cell lines harboring wild type FLT3 or 32D controls. We hypothesize that FLT3/ITD might contribute not only to the proliferative advantage of FLT3/ITD positive cells, but also to their reprogramming towards less differentiated stages, thus strengthening their malignant properties. This finding might explain the pronounced mutation rate of the aberrantly expressed FLT3 gene in AML, and, also, the inferior prognosis of FLT3/ITD positive AML patients. Moreover, the microarray data has revealed biological differences among individual ITD variants – a finding supporting the recent clinical data on the prognostic impact of the size of individual ITDs.