RNAi-mediated silencing of MLL-AF9 reveals leukemia-associated downstream targets and processes

Katrin K Fleischmann,Irene Schmid,Adelbert A Roscher,Philipp Pagel

doi:10.1186/1476-4598-13-27

Abstract

BackgroundThe translocation t(9;11)(p22;q23) leading to the leukemogenic fusion gene MLL-AF9 is a frequent translocation in infant acute myeloid leukemia (AML). This study aimed to identify genes and molecular processes downstream of MLL-AF9 (alias MLL-MLLT3) which could assist to develop new targeted therapies for such leukemia with unfavorable prognosis.MethodsIn the AML cell line THP1 which harbors this t(9;11) translocation, endogenous MLL-AF9 was silenced via siRNA while ensuring specificity of the knockdown and its efficiency on functional protein level.ResultsThe differential gene expression profile was validated for leukemia-association by gene set enrichment analysis of published gene sets from patient studies and MLL-AF9 overexpression studies and revealed 425 differentially expressed genes. Gene ontology analysis was consistent with a more differentiated state of MLL-AF9 depleted cells, with involvement of a wide range of downstream transcriptional regulators and with defined functional processes such as ribosomal biogenesis, chaperone binding, calcium homeostasis and estrogen response. We prioritized 41 gene products as candidate targets including several novel and potentially druggable effectors of MLL-AF9 (AHR, ATP2B2, DRD5, HIPK2, PARP8, ROR2 and TAS1R3). Applying the antagonist SCH39166 against the dopamine receptor DRD5 resulted in reduced leukemic cell characteristics of THP1 cells.ConclusionBesides potential new therapeutic targets, the described transcription profile shaped by MLL-AF9 provides an information source into the molecular processes altered in MLL aberrant leukemia.

Highlights

The translocation t(9;11)(p22;q23) leading to the leukemogenic fusion gene MLL-AF9 is a frequent translocation in infant acute myeloid leukemia (AML)
Certain partner genes are associated with distinct leukemia subtypes, e.g. MLL-AF4 with pro B acute lymphoblastic leukemia (ALL) and MLL-AF9, -AF6 and -AF10 with acute myeloid leukemia (AML) of M4 and M5 subtypes (French–American–British classification) [1]
The dopamine receptor antagonist SCH39166 exerts effects on malignant cell characteristics of THP1 cells As a proof of principle for the functional relevance of the suggested therapeutic targets, we examined the effects of the DRD1- and DRD5-specific dopamine receptor antagonist SCH39166 on THP1 cells

Summary

Introduction

The translocation t(9;11)(p22;q23) leading to the leukemogenic fusion gene MLL-AF9 is a frequent translocation in infant acute myeloid leukemia (AML). This study aimed to identify genes and molecular processes downstream of MLL-AF9 (alias MLL-MLLT3) which could assist to develop new targeted therapies for such leukemia with unfavorable prognosis. MLL-AF9 (alias MLL-MLLT3) results from the translocation t(9;11) (p22;q23) and is sufficient to initiate acute leukemia in murine models with potential secondary mutations being. An additional concern is that the wildtype functions of the involved proteins would be abrogated as well, leading to toxicity [7]. For these reasons, we explored downstream effects of MLL-AF9 in order to identify new alternative drug targets for MLL-AF9 positive AML. Functional relevance of one of these, the dopamine receptor DRD5, was confirmed as an antagonist resulted in reduced leukemic cell characteristics of THP1 cells

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