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.


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.
Gandhirajan
Rajesh Kumar
RK
Department I of Internal Medicine I, University at Cologne, Cologne, Germany.
Staib
Peter Anton
PA
Minke
Katharina
K
Gehrke
Iris
I
Plickert
Günther
G
Schlösser
Axel
A
Schmitt
Esther Katharina
EK
Hallek
Michael
M
Kreuzer
Karl-Anton
KA
D000818:Animals; D000970:Antineoplastic Agents / Q000737:chemistry / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects*; D006801:Humans; D015451:Leukemia, Lymphocytic, Chronic, B-Cell / Q000473:pathology*; D051821:Lymphoid Enhancer-Binding Factor 1 / Q000037:antagonists & inhibitors* / Q000378:metabolism / Q000502:physiology; D051379:Mice; D008819:Mice, Nude; D008970:Molecular Weight; D010569:Perylene / Q000031:analogs & derivatives* / Q000737:chemistry / Q000494:pharmacology; D015398:Signal Transduction / Q000187:drug effects; D014407:Tumor Cells, Cultured; D015854:Up-Regulation / Q000187:drug effects; D051153:Wnt Proteins / Q000037:antagonists & inhibitors* / Q000378:metabolism / Q000502:physiology; D023041:Xenograft Model Antitumor Assays; D051176:beta Catenin / Q000037:antagonists & inhibitors* / Q000378:metabolism / Q000502:physiology
D000970:Antineoplastic Agents; C538948:CGP049090; D051821:Lymphoid Enhancer-Binding Factor 1; C538949:PKF115-584; D051153:Wnt Proteins; D051176:beta Catenin; D010569:Perylene