Dataset Information

Targeting Wnt-driven cancer through the inhibition of Porcupine by LGK974.

ABSTRACT: Wnt signaling is one of the key oncogenic pathways in multiple cancers, and targeting this pathway is an attractive therapeutic approach. However, therapeutic success has been limited because of the lack of therapeutic agents for targets in the Wnt pathway and the lack of a defined patient population that would be sensitive to a Wnt inhibitor. We developed a screen for small molecules that block Wnt secretion. This effort led to the discovery of LGK974, a potent and specific small-molecule Porcupine (PORCN) inhibitor. PORCN is a membrane-bound O-acyltransferase that is required for and dedicated to palmitoylation of Wnt ligands, a necessary step in the processing of Wnt ligand secretion. We show that LGK974 potently inhibits Wnt signaling in vitro and in vivo, including reduction of the Wnt-dependent LRP6 phosphorylation and the expression of Wnt target genes, such as AXIN2. LGK974 is potent and efficacious in multiple tumor models at well-tolerated doses in vivo, including murine and rat mechanistic breast cancer models driven by MMTV-Wnt1 and a human head and neck squamous cell carcinoma model (HN30). We also show that head and neck cancer cell lines with loss-of-function mutations in the Notch signaling pathway have a high response rate to LGK974. Together, these findings provide both a strategy and tools for targeting Wnt-driven cancers through the inhibition of PORCN.

SUBMITTER: Liu J

PROVIDER: S-EPMC3864356 | biostudies-literature | 2013 Dec

REPOSITORIES: biostudies-literature

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Targeting Wnt-driven cancer through the inhibition of Porcupine by LGK974.

Liu Jun J Pan Shifeng S Hsieh Mindy H MH Ng Nicholas N Sun Fangxian F Wang Tao T Kasibhatla Shailaja S Schuller Alwin G AG Li Allen G AG Cheng Dai D Li Jie J Tompkins Celin C Pferdekamper AnneMarie A Steffy Auzon A Cheng Jane J Kowal Colleen C Phung Van V Guo Guirong G Wang Yan Y Graham Martin P MP Flynn Shannon S Brenner J Chad JC Li Chun C Villarroel M Cristina MC Schultz Peter G PG Wu Xu X McNamara Peter P Sellers William R WR Petruzzelli Lilli L Boral Anthony L AL Seidel H Martin HM McLaughlin Margaret E ME Che Jianwei J Carey Thomas E TE Vanasse Gary G Harris Jennifer L JL

Proceedings of the National Academy of Sciences of the United States of America 20131125 50

Wnt signaling is one of the key oncogenic pathways in multiple cancers, and targeting this pathway is an attractive therapeutic approach. However, therapeutic success has been limited because of the lack of therapeutic agents for targets in the Wnt pathway and the lack of a defined patient population that would be sensitive to a Wnt inhibitor. We developed a screen for small molecules that block Wnt secretion. This effort led to the discovery of LGK974, a potent and specific small-molecule Porcu ...[more]

PMID: 24277854

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Project description:BackgroundWnt signaling plays key roles in cellular and physiological processes, including cell proliferation, differentiation and migration during development and tissue homeostasis in adults. This pathway can be defined as Wnt/β-catenin-dependent or β-catenin-independent or "non-canonical", both signaling are involved in neurite and synapse development/maintenance. Porcupine (PORCN), an acylase that o-acylates Wnt ligands, a major modification in secretion and interaction with its receptors. We use Wnt-C59, a specific PORCN inhibitor, to block the secretion of endogenous Wnts in embryonic hippocampal neurons (DIV 4). Under these conditions, the activity of exogenous Wnt ligands on the complexity of the dendritic tree and axonal polarity were evaluated METHODS: Cultured primary embryonic hippocampal neurons obtained from Sprague-Dawley rat fetuses (E18), were cultured until day in vitro (DIV) 4 (according to Banker´s protocol) and treated with Wnt-C59 for 24 h, Wnt ligands were added to the cultures on DIV 3 for 24 h. Dendritic arbors and neurites were analysis by fluorescence microscopy. Transfection with Lipofectamine 2000 on DIV 2 of plasmid expressing eGFP and KIF5-Cherry was carried out to evaluate neuronal polarity. Immunostaining was performed with MAP1B and Tau protein. Immunoblot analysis was carried out with Wnt3a, β-catenin and GSK-3β (p-Ser9). Quantitative analysis of dendrite morphology was carried out with ImageJ (NIH) software with Neuron J Plugin.ResultsWe report, here, that Wnt-C59 treatment changed the morphology of the dendritic arbors and neurites of embryonic hippocampal neurons, with decreases β-catenin and Wnt3a and an apparent increase in GSK-3β (p-Ser9) levels. No effect was observed on axonal polarity. In sister cultures, addition of exogenous Wnt3a, 5a and 7a ligands rescued the changes in neuronal morphology. Wnt3a restored the length of neurites to near that of the control, but Wnt7a increased the neurite length beyond that of the control. Wnt5a also restored the length of neurites relative to Wnt concentrations.ConclusionsResults indicated that Wnt ligands, added exogenously, restored dendritic arbor complexity in embryonic hippocampal neurons, previously treated with a high affinity specific Porcupine inhibitor. We proposed that PORCN is an emerging molecular target of interest in the search for preclinical options to study and treat Wnt-related diseases. Video Abstract.

Project description:ObjectiveProstate cancer, marked by a high incidence and mortality rate, presents a significant challenge, especially in the context of castration-resistant prostate cancer (CRPC) with limited treatment options due to drug resistance. This study aims to explore the anti-tumor effects of Xihuang Pills (XHP) on CRPC, focusing on metabolic reprogramming and the Wnt/β-catenin pathway.MethodsIn vitro and in vivo biofunctional assays were employed to assess the efficacy and mechanisms of XHP. Subcutaneous xenografts of PC3 in mice served as an in vivo model to evaluate XHP's anti-tumor activity. Tumor volume, weight, proliferation, and apoptosis were monitored. Various assays, including CCK8, TUNEL assay, QRT-PCR, and Western Blotting, were conducted to measure metabolic reprogramming, proliferation, apoptosis, and cell cycle in prostate cancer cells. RNA-seq analysis predicted XHP's impact on prostate cancer, validating the expression of Wnt/β-catenin-related proteins and mRNA. Additionally, 58 compounds in XHP were identified via LC-MS/MS, and molecular docking analysis connected these compounds to key genes.ResultsIn vitro and in vivo experiments demonstrated that XHP significantly inhibited CRPC cell viability, induced apoptosis, and suppressed invasion and migration. mRNA sequencing revealed differentially expressed genes, with functional enrichment analysis indicating modulation of key biological processes. XHP treatment downregulated Wnt signaling pathway-related genes, including CCND2, PRKCG, and CCN4. Moreover, XHP effectively inhibited glucose uptake and lactate production, leading to reduced HIF-1α and glycolytic enzymes (GLUT1, HK2, PKM2), suggesting its potential in attenuating the Warburg effect. Molecular docking analysis suggested a plausible interaction between XHP's active compounds and Wnt1 protein, indicating a mechanism through which XHP modulates the Wnt/β-catenin pathway.ConclusionXHP demonstrated remarkable efficacy in suppressing the growth, proliferation, apoptosis, migration, and invasiveness of prostate tumors. The interaction between XHP's active constituents and Wnt1 was evident, leading to the inhibition of Wnt1 and downstream anti-carcinogenic factors, thereby influencing the β-catenin/HIF-1α-mediated glycolysis.

Dataset Information

Targeting Wnt-driven cancer through the inhibition of Porcupine by LGK974.

Publications

Targeting Wnt-driven cancer through the inhibition of Porcupine by LGK974.

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