Abstract SY23-02: Targeting the Wnt pathway

Abstract

Abstract The Wnt signaling cascade is an extremely complex signal transduction system involving 19 extracellular mammalian glycoprotein Wnt ligands, 10 Fzd family 7-TM spanning receptors and the co-receptors LRP 5, 6 as well as additional non-classical Wnt receptors (e.g. Ryk, Ror). Wnt ligands trigger a variety of intracellular responses, broadly associated with canonical (increase in nuclear β-catenin) and non-canonical (planar cell polarity, Ca++/PKC activation). In a gross oversimplification, the former is often associated with proliferation and lack of differentiation (for example as a hallmark of dysregulated Wnt signaling in cancer), whereas the latter is often associated with cell and organismal differentiation. Beyond classical Wnt activated translocation of β-catenin to the nucleus, a number of other factors (growth factors, prostaglandins, etc.) can induce the nuclear localization of β-catenin. β-catenin plays a key role in both aspects of the Wnt signaling cascade (canonical and non-canonical) through both its nuclear functions and cytoskeletal/cytoplasmic membrane interactions. Nuclear β-catenin drives the expression of a Wnt/catenin regulated-cassette of genes, whereas outside of the nucleus, β-catenin plays a critical role in cell-cell interactions and cellular polarization. Wnt signaling plays important roles throughout development. Although most would agree that Wnt signaling is important in stem cell biology, there is no consensus as to whether Wnt signaling is important for proliferation and maintenance of potency (pluri- or multipotency) or differentiation of stem/progenitor cells. Wnt/β-catenin signaling has been demonstrated to maintain pluripotency in ES cells and is critical for the expansion of neural progenitors thereby increasing brain size. However, Wnt/β-catenin signaling is also required for neural differentiation of ES cells, fate decision in neural crest stem cells and Wnt3a has been reported to promote differentiation into the neural and astrocytic lineages by inhibiting neural stem cell maintenance. Clearly, Wnt/β-catenin signaling also plays a critical role in lineage decision/commitment. These dramatically different outcomes upon activation of the Wnt signaling cascade has fueled enormous controversy concerning the role of Wnt signaling in maintenance of potency and induction of differentiation. Until recently, a rationale for the dichotomous behavior of Wnt/β-catenin signaling in controlling both proliferation and differentiation has been unclear. Recently, using a selective antagonist of the CBP/β-catenin interaction ICG-001 that we identified utilizing a chemical genomic approach, we have developed a model to explain the divergent activities of Wnt/β-catenin signaling (Figure 3). Our model highlights the distinct roles of the coactivators CBP and p300 in the Wnt/β-catenin signaling cascade. The critical feature of the model is that CBP/β-catenin mediated transcription is critical for stem cell/progenitor cell maintenance and proliferation, whereas a switch to p300/β-catenin mediated transcription is the initial critical step to initiate differentiation and a decrease in cellular potency. Identification of the small molecule IQ-1 that allows for the Wnt/ß-Catenin-driven long-term expansion of mouse ES cells by blocking the p300/ß-catenin interaction, and thereby prevents spontaneous differentiation in the absence of LIF, is consistent and lends further support to our model. This study demonstrated that IQ-1, in combination with Wnt3a to stimulate nuclear translocation of β-catenin, is sufficient to sustain pluripotency by decreasing the interaction of ß-catenin with p300 and consequently increasing its interaction with CBP. Removal of IQ-1 from the mES cultures rapidly leads to loss of pluripotency, even in the presence of Wnt3a. The switch to p300/β-catenin mediated transcription, whether induced pharmacologically, by removal of IQ-1, or addition of ICG-001, or naturally (e.g. LIF withdrawal), is critical to initiate a differentiative program with a more limited proliferative capacity. Importantly, the ß-catenin coactivator switching mechanism is also critical in the maintenance of hES cell pluripotency and both mouse and human SSC populations. These studies represents additional “proof-of-principle” that complex phenotypes can be controlled by a single small molecule, isolated for its desirable and specific pharmacologic activity. Given the fact that multiple mutations can lead to the aberrant activation of nuclear β-catenin signaling, there is a clear need for drugs that attenuate the nuclear functions of β-catenin. The small molecule antagonist that our laboratory developed, ICG-001, by binding to the coactivator CBP and not to its highly homologous relative p300, specifically downregulates a subset of Wnt/β-catenin-driven genes including S100A4 and survivin the number 1 and 4 transcriptomes up-regulated in cancer. ICG-001 has proven to be an invaluable tool in helping us to dissect the complex signal transduction pathways and interactions involved in the regulation of “stemness”. Aspects of Wnt signaling in stem cells and cancer stem cells will be discussed. Citation Format: Michael Kahn, Jia-Ling Teo, Philipp Manegold, Tomoyo Sasaki, Yi Zhao. Targeting the Wnt pathway [abstract]. In: Proceedings of the AACR 101st Annual Meeting 2010; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr SY23-02