Small molecule inhibitors of Wnt/beta-catenin/lef-1 signaling induces apoptosis in chronic lymphocytic leukemia cells in vitro and in vivo.
Small molecule inhibitors of Wnt/beta-catenin/lef-1 signaling induces apoptosis in chronic lymphocytic leukemia cells in vitro and in vivo.
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发表时间：
2010
期刊：
影响因子：
4.8
通讯作者：
R. Gandhirajan;P. Staib;Katharina Minke;I. Gehrke;Günther Plickert;Axel Schlösser;E. Schmitt;M. Hallek;K. Kreuzer
中科院分区：
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作者：
R. Gandhirajan;P. Staib;Katharina Minke;I. Gehrke;Günther Plickert;Axel Schlösser;E. Schmitt;M. Hallek;K. Kreuzer
与正常B细胞相比，类磷脂增强因子-1（lef-1）在B细胞慢性淋巴细胞白血病（CLL）中过表达，并转录与CLL发病机制有关的几个基因。因此，我们假设lef-1的拮抗作用可能导致CLL细胞的杀伤。我们使用两种Wnt/β-连环蛋白/lef-1信号传导的小分子抑制剂（CGP 049090和PKF 115 -584）来检验我们的假设。设计与方法采用富集的CLL细胞和健康的B细胞。使用核转染进行小干扰RNA（siRNA）介导的原代CLL细胞中lef-1的敲低，并使用基于ATP的细胞活力测定评估两种小分子的50%致死浓度（LC（50））。在时间进程实验中用不同的凋亡标志物研究凋亡反应。通过lef-1/β-连环蛋白相互作用的免疫共沉淀实验证明了小分子的特异性。在JVM-3皮下异种移植模型中进行体内研究。结果siRNA抑制lef-1可导致CLL细胞凋亡增加和JVM-3细胞增殖抑制。两种小分子抑制剂（CGP 049090和PKF 115 -584）有效地杀死CLL细胞（LC（50）<1 μ M），而正常B细胞没有受到显著影响。免疫共沉淀显示β-连环蛋白/lef-1相互作用的选择性破坏。体内研究显示，与溶剂处理对照相比，CGP 049090和PKF 115 -584的肿瘤抑制率分别为69%和57%，并且干预耐受性良好。结论：我们已经证明了靶向lef-1是一种新的选择性治疗CLL的方法。CGP 049090或PKF 115 -584可能是治疗CLL和其他恶性肿瘤的有吸引力的化合物，值得进一步（预）临床评估。
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.