Small molecule inhibitors of Wnt/beta-catenin/lef-1 signaling induces apoptosis in chronic lymphocytic leukemia cells in vitro and in vivo.

BACKGROUND: Lymphoid enhancer factor-1 (lef-1) is overexpressed in B-cell chronic lymphocytic leukemia (CLL) when compared with normal B cells and transcribes several genes implicated in the pathogenesis of CLL. We therefore hypothesize that antagonism of lef-1 might lead to killing of CLL cells. We used two small molecule inhibitors of Wnt/beta-catenin/lef-1 signaling (CGP049090 and PKF115-584) to test our hypothesis. DESIGN AND METHODS: Enriched CLL cells and healthy B cells were used in this study. Small interfering RNA (siRNA)-mediated knockdown of lef-1 in primary CLL cells was done using nucleofection, and 50% lethal concentration (LC(50)) of two small molecules was assessed using ATP-based cell viability assay. Apoptotic response was investigated in time course experiments with different apoptotic markers. Specificity of the small molecules was demonstrated by coimmunoprecipitation experiments for the lef-1/beta-catenin interaction. In vivo studies were done in JVM-3 subcutaneous xenograft model. RESULTS: Inhibition of lef-1 by siRNA leads to increased apoptosis of CLL cells and inhibited proliferation of JVM-3 cell lines. The two small molecule inhibitors (CGP049090 and PKF115-584) efficiently kill CLL cells (LC(50)<1 microM), whereas normal B cells were not significantly affected. Coimmunoprecipitation showed a selective disruption of beta-catenin/lef-1 interaction. In vivo studies exhibited tumor inhibition of 69% with CGP049090 and 57% with PKF115-584 when compared with vehicle-treated controls, and the intervention was well tolerated. CONCLUSIONS: We have demonstrated that targeting lef-1 is a new and selective therapeutic approach in CLL. CGP049090 or PKF115-584 may be attractive compounds for CLL and other malignancies that deserve further (pre)clinical evaluation.

Small molecule inhibitors of WNT signaling effectively induce apoptosis in acute myeloid leukemia cells.

In a significant proportion of acute myeloid leukemia (AML) cases the canonical WNT pathway is upregulated and targeting the WNT/LEF1 signaling cascade in AML may be a promising approach to develop new treatments for this entity. Recently two compounds (CGP049090 and PFK115-584) have been identified, which specifically inhibit complexation of beta-catenin (CTNNB1) and lymphoid enhancer-binding factor 1 (LEF1) leading to transcriptional inactivation of LEF1 in colon carcinoma cell lines. To evaluate the effect of WNT inhibition utilizing theses compounds with regard to their effectivity in AML we treated the AML cell lines Kasumi-1 and HL-60, primary AML blasts and healthy peripheral blood mononuclear cells (PBMCs) with varying concentrations of both substances. Treatment with both compounds for 24 h resulted in a significant killing of AML cell lines and primary AML blasts with 50% effective concentration doses (EC(50)) within the submicromolar range. PBMCs were not significantly affected as indicated by EC(50)-values 100-fold higher than for AML cells. Cell kill was mediated by apoptosis as indicated by induction of caspases 3 and 7 and cleavage of poly(ADP-ribose) polymerase (PARP) upon treatment. Furthermore, we could show that both compounds substantially decrease expression of CTNNB1/LEF1 target genes c-myc, cyclin D1 and survivin, proofing the specificity of the substances. This was shown in both, AML cell lines and most of the tested primary samples. Our data demonstrate that targeting this pathway seems to be an innovative approach in the treatment of AML.