Sulindac Sulfide Reverts the Activation of the WNT-Signaling and Aberrant Self Renewal Induced by the AML-Associated Fusion Proteins PML/RARα and PLZF/RARα.

Leukemia-specific translocations - t(15;17), t(11;17), or t(8;21) - lead to the expression of leukemia associated fusion proteins (LAFP) such as PML/RAR, PLZF/RAR and AML-1/ETO. LAFP induce and maintain a leukemic phenotype by blocking terminal differentiation of hematopoietic progenitors and by increasing the self renewal potential of the leukemic stem cells (LSC). The key mechanims by which LAFP increase LSC self renewal is the activation of the Wnt-signaling pathway by up-regulating of γ-catenin and β-catenin at a transcriptional level. The aberrant activation of Wnt-signaling by the LAFP decisively contributes to the pathogenesis of AML. To disclose whether a “leukemic stem cell therapy” is effective we targeted the Wnt-signaling by Sulindac Sulfid (SuSu) in PML/RAR- or PLZF/RAR- (X-RAR) positive stem cell models. SuSu represents the active metabolite of Sulindac, a nonsteroidal anti-inflammatory drug (NSAID), known to inactivate the Wnt-signaling. SuSu was used at a clinically achievable concentration of 50–100 μM. As leukemia models we used U937 cells stably expressing the X-RAR and NB4 cells. As stem cell models we used i.) the aldehyde dehydrogenase positive/CD34+/CD38- fraction of the KG-1 cells stably expressing the X-RAR; ii.) Sca-1+/lin- murine HSC retrovirally transduced with the X-RAR in semi solid medium. The amount of total γ-catenin and β-catenin and activated β-catenin was determined by immunoblotting. We report that SuSu i.) down-regulated not only β-catenin but also γ-catenin in X-RAR expressing U937 and KG-1 cells; ii.) reduced the active form of β-catenin in the presence of X-RAR; iii.) induced a high apoptosis rate in PML/RAR-positive NB4 cells; iv.) reduced the “stem cell fraction” of KG-1 cells expressing X-RAR but not of mock transfected controls; iv.) reduced the self renewal potential of X-RAR-positive HSC as revealed by a significantly reduced replating efficiency. Here we provide evidence that the exposure to therapeutically achievable dosages of a NSAID revert the aberrant activation of the Wnt-signaling by LAFP. The significant reduction of the aberrant self renewal potential of HSC in the presence of X-RAR further support that the inhibition of the aberrantly activated Wnt signaling in AML might be a valid molecular therapy approach which has to further validated in in vivo leukemia models and in a more clinical setting.