DGG-300273, a novel WNT/ß-catenin inhibitor, induces apoptotic cell death by activating ROS-BIM signaling in a Wnt-dependent manner in colon cancer cells.

Dysregulated Wnt signaling is associated with malignant oncogenic transformation, especially in colon cancer. Recently, numerous drugs have been developed based on tumorigenesis biomarkers, thus having high potential as drug targets. Likewise, WNT/ß-catenin pathway members are attractive therapeutic targets for colon cancer and are currently in various stages of development. However, although inhibitors of proteins regulating the WNT/ß-catenin signaling pathway have been extensively studied, they have yet to be clinically approved, and the underlying molecular mechanism(s) of their anticancer effects remain poorly understood. Herein, we show that a novel WNT/ß-catenin inhibitor, DGG-300273, inhibits colon cancer cell growth in a Wnt-dependent manner due to upregulation of the BCL2-family protein Bim and caspase-dependent apoptotic cell death. Additionally, DGG-300273-mediated cell death occurs by increased reactive oxygen species (ROS), as shown by abrogation of apoptotic cell death and ROS production following pretreatment with the antioxidant N-acetylcysteine. These results suggest that DGG-300273 represents a promising investigational drug for the treatment of Wnt-associated cancer, thus warranting further characterization and study.

Wnt/ß-catenin inhibitor ICG-001 enhances the antitumor efficacy of radiotherapy by increasing radiation-induced DNA damage and improving tumor immune microenvironment in hepatocellular carcinoma.

BACKGROUND AND PURPOSE: Radiotherapy (RT) has a promising anti-tumor effect depending on its effects on both cancer cells and tumor immune microenvironment (TIME). As one of the most common alterations in hepatocellular carcinoma (HCC), wnt/ß-catenin pathway activation, has been reported to induce radioresistance and suppressive TIME. In this study, we aim to explore the effect of wnt/ß-catenin inhibitor ICG-001 on radiosensitivity and RT-related TIME of HCC and the underlying mechanism. MATERIALS AND METHODS: C57BL/6 and nude mouse tumor models were used to evaluate the efficacy of different treatments on tumor growth, recurrence and mice survival. Flow cytometry was performed to assess tumor infiltrating lymphocytes (TILs). DNA damage response (DDR) and radioresistance was investigated by colony formation assays, γ-H2AX and micronuclei measurements. RESULTS: The addition of ICG-001 to RT exhibited better anti-tumor and survival-prolong efficacy in C57BL/6 than nude mice. TILs analysis revealed that ICG-001 plus RT boosted the infiltration and IFN-γ production of TIL CD8+ T cells, meanwhile reduced the number of Tregs. Moreover, mechanistic study demonstrated that ICG-001 increased the radiation-induced DDR of HCC cells by suppressing p53, thus leading to stronger activation of cGAS/STING pathway. Utilization of cGAS/STING pathway inhibitors impaired the therapeutic effect of combination therapy. Furthermore, combination therapy led to stronger immunologic memory and tumor relapse prevention. CONCLUSIONS: Our findings showed that ICG-001 displayed both local and systematic effects by increasing radiosensitivity and improving immunity in HCC, which indicated that ICG-001 might be a potential synergetic treatment for radiotherapy and radioimmunotherapy in HCC patients.

Enhancing insulin sensitivity by dual PPARγ partial agonist, ß-catenin inhibitor: Design, synthesis of new αphthalimido-o-toluoyl2-aminothiazole hybrids.

AIMS: Partial PPARγ agonists attracted substantially heightened interest as safer thiazolidinediones alternatives. On the other hand, Wnt/ß-catenin antagonists have been highlighted as promising strategy for type 2 diabetes management via up-regulating PPARγ gene expression. We aimed at synthesizing novel partial PPARγ agonists with ß-catenin inhibitory activity which could enhance insulin sensitivity and avoid the side effects of full PPARγ agonists. MAIN METHODS: We synthesized novel series of α-phthlimido-o-toluoyl-2-aminothiazoles hybrids for evaluating their antidiabetic activity and discovering its mechanistic pathway. We assessed effect of the new hybrids on PPARγ activation using a luciferase reporter assay system. Moreover, intracellular triglyceride levels, gene levels of c/EBPα, PPARγ and PPARγ targets including GLUT4, adiponectin, aP2 were measured in 3T3-L1 cells. Uptake of 2-DOG together with PPARγ and ß-catenin protein levels were evaluated in 3T3-L1cells. In addition, molecular docking studies with PPARγ LBD, physicochemical properties and structure activity relationship of the novel hybrids were also studied. KEY FINDINGS: Three of the synthesized hybrids showed partial PPARγ agonistic activity and distinct PPARγ binding pattern. These compounds modulated PPARγ gene expression and PPARγ target genes; and increased glucose uptake in 3T3-L1 and slightly induced adipogenesis compared to rosiglitazone. Moreover, these compounds reduced ß-catenin protein level which reflected in increased both PPARγ gene and protein levels that leads to improved insulin sensitivity and increased GLUT4 and adiponectin gene expression. SIGNIFICANCE: Our synthesized compounds act as novel partial PPARγ agonists and ß-catenin inhibitors that have potent insulin sensitizing activity and mitigate the lipogenic side effects of TZDs.

WNT/ß-catenin signal inhibitor IC-2-derived small-molecule compounds suppress TGF-ß1-induced fibrogenic response of renal epithelial cells by inhibiting SMAD2/3 signalling.

Renal fibrosis compromises kidney function, and it is a risk factor for chronic kidney disease (CKD). CKD ultimately progresses to end-stage kidney disease that can be cured only by kidney transplantation. Owing to the increasing number of CKD patients, effective treatment strategies are urgently required for renal fibrosis. TGF-ß is a well-established fibrogenic factor that signals through SMAD2/3 signaling pathway. It was shown that there is a cross-talk between TGF-ß/SMAD and WNT/ß-catenin signaling pathways in renal tubular epithelial cells, and that a WNT/ß-catenin inhibitor, ICG-001, ameliorates TGF-ß1induced renal fibrosis. IC-2, a derivative of ICG-001, has been shown to potently induce hepatocyte differentiation of human mesenchymal stem cells by inhibiting WNT/ß-catenin signaling. In the present study, we examined the effect of ICG-001, IC-2, and IC-2 derivatives (IC-2-506-1, IC-2-506-2, IC-2-506-3, IC-2-Ar-Cl, IC-2-OH, IC-2-OTBS, and IC-2-F) on TGF-ß1-induced SMAD activation and fibrogenic response in immortalized human renal tubular epithelial HK-2 cells. All these compounds inhibited LiCl-induced WNT/ß-catenin reporter activation to a similar extent, whereas ICG-001, IC-2-OTBS, and IC-2-F almost completely suppressed TGF-ß1-induced SMAD reporter activation without apparent cytotoxicity. Phosphorylation of SMAD2/3 by TGF-ß1 was more potently inhibited by IC-2-OTBS and IC-2-F than by ICG-001 and IC-2. IC-2-F suppressed TGF-ß1-induced COL1A1 protein expression, whereas IC-2-506-1 and IC-2-OTBS suppressed TGF-ß1-induced epithelial-mesenchymal transition. These results demonstrated that IC-2 derivatives suppress the TGF-ß1-induced fibrogenic response of tubular epithelial cells and thus could be promising therapeutic agents for the treatment of renal fibrosis.

SHISA6 Confers Resistance to Differentiation-Promoting Wnt/ß-Catenin Signaling in Mouse Spermatogenic Stem Cells.

In the seminiferous tubules of mouse testes, a population of glial cell line-derived neurotrophic factor family receptor alpha 1 (GFRα1)-positive spermatogonia harbors the stem cell functionality and supports continual spermatogenesis, likely independent of asymmetric division or definitive niche control. Here, we show that activation of Wnt/ß-catenin signaling promotes spermatogonial differentiation and reduces the GFRα1+ cell pool. We further discovered that SHISA6 is a cell-autonomous Wnt inhibitor that is expressed in a restricted subset of GFRα1+ cells and confers resistance to the Wnt/ß-catenin signaling. Shisa6+ cells appear to show stem cell-related characteristics, conjectured from the morphology and long-term fates of T (Brachyury)+ cells that are found largely overlapped with Shisa6+ cells. This study proposes a generic mechanism of stem cell regulation in a facultative (or open) niche environment, with which different levels of a cell-autonomous inhibitor (SHISA6, in this case) generates heterogeneous resistance to widely distributed differentiation-promoting extracellular signaling, such as WNTs.