Chemical Disruption of Wnt-dependent Cell Fate Decision-making Mechanisms in Cancer and Regenerative Medicine

Abstract

Cell-to-cell signaling molecules such as the Wnt proteins that directly influence the expression of cell-type specific transcriptional programs are essential for tissue generation in metazoans. The mechanisms supporting cellular responses to these molecules represent potential points of intervention for directing cell fate outcomes in therapeutic contexts. Small molecules that modulate Wnt-mediated cellular responses have proven to be powerful probes for Wnt protein function in diverse biological settings including cancer, development, and regeneration. Whereas efforts to develop these chemicals as therapeutic agents have dominated conversation, the unprecedented modes-of-action associated with these molecules and their implications for drug development deserve greater examination. In this review, we will discuss how medicinal chemistry efforts focused on first in class small molecules targeting two Wnt pathway components--the polytopic Porcupine (Porcn) acyltransferase and the cytoplasmic Tankyrase (Tnks) poly-ADP-ribosylases--have contributed to our understanding of the druggable genome and expanded the armamentarium of chemicals that can be used to influence cell fate decision-making.

Fig. (1). Overview of the Wnt/β-catenin signal transduction pathway

Reviews of Wnt-mediated cellular responses that do not utilize β-catenin can be found in [12, 27].

Fig. (2). Mechanism of action for Porcn and Tnks inhibitors

Left: Inhibition of endoplasmic reticulum-localized Porcn results in loss of Wnt fatty acylation. Wnt proteins devoid of their lipid moiety are not recognized by the Wntless (Wls) chaperone resulting in their sequestration in the secretory pathway. Wnt molecules in addition to regulating β-catenin/TCF activity control other cellular responses not depicted here. Right: Disruption of Tnks1 & 2 activity with chemicals results in loss of Axin protein PARylation, a biochemical change that promotes Axin destruction by ubiquitinylation. Thus in cells treated with Tnks inhibitors, Axin accumulates and accelerates the rate of β-catenin turnover. Without a sufficient abundance of β-catenin, the TCF/LEF proteins are unable to elicit a meaningful transcriptional response. The turnover rate of other proteins in addition to β-catenin that are regulated by Tnks and Axin are not depicted here but discussed in Section 3.

Fig. (3). Pre-clinical cancer therapeutic studies of Porcn and Tnks inhibitors in mice

C59 and JW55 are predecessor molecules of LGK-974 and G007-LK, respectively. MMTV-Wnt1 transgenic mice develop ductal hyperplasia in mammary gland tissue due to introduction of a viral transcriptional enhancer that induces Wnt1 expression. Liu et al has shown that loss of Notch signaling in head and neck squamous cell carcinoma cell lines induce cell growth-promoting Wnt/β-catenin signaling. Thioacetamide is a chemical that induces dysregulated growth in the biliary tract of animals that are compromised for the p53 tumor suppressor. Juan et al. has shown that lung cancers initiated by a mutation in the protooncogenic Braf protein (V600E) are dependent upon β-catenin signaling.

Fig. (4). Summary of cell engineering achievements using the IWP and IWR classes of Porcn and Tnks inhibitors, respectively

IWP-1, -2, and -4 inhibit Porcn whereas IWR-1 compound inhibits Tnks1 & 2. mES cells=mouse embryonic stem cells; hES cells=human embryonic stem cells; hiPS cells= human induced pluripotent stem cells.

Fig. (5). Summary of selected classes of Tnks inhibitors and their mode of binding to Tnks enzymes

NAD+ is the substrate for Tnks enzymes. Chemical components of small molecules that occupy the adenosine-binding pocket are aggregated into head, spacer, and tail for discussion purposes. Although representative molecules targeting various active site pockets are discussed here, a more extensive overview that includes references to additional Tnks inhibitors can be found in these excellent reviews [68, 89, 90].

Fig. (6). The Porcn active site brings together cytoplasmic palmitoleoyl-CoA and ER luminal Wnt proteins to produce fatty acylated Wnt proteins

Various Porcn inhibitors presumably target the Porcn active site to prevent the formation of a functionally meaningful tri-partite complex.