Prognostic and Functional Relevance of Aberrant Microrna-9/9* Expression in Acute Myeloid Leukemia.

Abstract 2542

Acute myeloid leukemia (AML) is a highly diverse disease characterized by various cytogenetic and molecular abnormalities, resulting in increased cell proliferation and block in myeloid differentiation. Several studies have demonstrated that distinct microRNA (miRNA) expression profiles are associated with cytogenetic and molecular abnormalities and reveal miRNAs with a suggested functional role in AML biology and with prognostic value for treatment outcome.

Expression levels of miR-9/9* in different stages of normal human granulopoiesis are low or not detectable by Q-RT-PCR. In the current study we demonstrated abnormal expression of both miRNA-9 and miRNA-9* (miR-9/9*) in a cohort of 647 patients with AML. MiR-9/9* expression profiles displayed distinct distributions in genotypic AML subtypes. In contrast with the highly elevated expression values of miR-9/9* in the majority of the AMLs, AML with t(8;21) and AML with double mutations in CEBPA gene showed low or undetectable miR-9/9* expression levels. Subsets of AML with 11q23 abnormalities, AML with NPM1 gene mutations as well as cases with FLT3- ITD mutations were significantly associated with high miR-9/9* expression. In order to investigate the effect of abnormal expression of miR-9/9* in relation to survival in AML, multivariable analysis was performed using different survival endpoints. The quartile of patients with AML with the highest miR-9 expression values showed highly significant improved overall survival (P= 0.005, HR=0.672), event-free survival (P=0.012, HR=0.72), and relapse-free survival (P=0.004, HR=0.585).

Enforced expression of miR-9/9* in both murine 32D myeloid cell line and mouse hematopoietic stem and progenitors (HSPCs), that both lack endogenous miR-9/9* expression, inhibited granulocytic cell development. Ectopic miR-9/9* expression in 32D cells, induced a block in G-CSF-induced cell differentiation and enhanced cell expansion. Overexpression of miR-9/9* in HSPCs caused reduced G-CSF- and GM-CSF-induced colony formation capacity resulting in reduced colony numbers and size.

MiR-9/9* target gene analysis with Affymetrix gene expression data of primary AML cases and prediction algorithms (TargetScan and microCosm) suggested ERG, a transcription factor of the ets family, involved in hematopoiesis, as a most prominent candidate target of miR-9. ERG was validated as miR-9 targets on protein level by Western blot analyses and luciferase assays.

In conclusion, we demonstrate aberrant expression of miRNA-9/9* with prognostic relevance in AML. Enforced expression of miR-9/9* in both the murine 32D myeloid cell line and primary HSPCs inhibited granulocytic cell development. Furthermore, these data suggest that miR-9/9* may be functionally involved in the pathogenesis of AML by translational deregulation of ERG.