Distinct biological and transcriptional features of FLT3-ITD variants in vitro and in vivo. Mus musculus

Treatment of FLT3-ITD mutated AML remains a clinical challenge as this subset of patients show high incidence of relapse, even after allogeneic stem cell transplantation. Recently, we defined a molecular subset of FLT3-ITD mutations, located in the FLT3-tyrosine kinase domain 1 (TKD1). In multivariable analyses, insertion of ITD variants in the beta1-sheet (TKD1) was identified as an unfavorable prognostic factor for achievement of a complete remission relapse-free survival and overall survival independently of other prognostic factors such as allelic ratio. In the presented analysis we characterize ITD variants located in the TKD1. While transforming capacity and activation of downstream signaling nodes appears comparable to ‘classical’ ITD mutations located in the JM-domain, TKD1-ITDs reveal a distinct gene expression profile and confer decreased sensitivity to kinase inhibitor therapy in vitro and in vivo. This knowledge may impact clinical decisions in regard to TKI therapy versus allogeneic stem cell transplantation. Overall design: RNA was isolated from 32D cells transduced with the respective FLT3-ITD constructs using TRIzol (Life Technologies). Total RNA was converted into double-stranded cDNA by reverse transcription using the 3' IVT Express Kit from Affymetrix and samples were hybridized to Affymetrix GeneChip Mouse Genome 430 2.0 Arrays according to the manufacturer’s recommendations (Affymetrix).