Gene Summary

Gene:AXIN1; axin 1
Aliases: AXIN, PPP1R49
Summary:This gene encodes a cytoplasmic protein which contains a regulation of G-protein signaling (RGS) domain and a dishevelled and axin (DIX) domain. The encoded protein interacts with adenomatosis polyposis coli, catenin beta-1, glycogen synthase kinase 3 beta, protein phosphate 2, and itself. This protein functions as a negative regulator of the wingless-type MMTV integration site family, member 1 (WNT) signaling pathway and can induce apoptosis. The crystal structure of a portion of this protein, alone and in a complex with other proteins, has been resolved. Mutations in this gene have been associated with hepatocellular carcinoma, hepatoblastomas, ovarian endometriod adenocarcinomas, and medullablastomas. Two transcript variants encoding distinct isoforms have been identified for this gene. [provided by RefSeq, Dec 2010]
Databases:OMIM, VEGA, HGNC, Ensembl, GeneCard, Gene
Source:NCBIAccessed: 21 August, 2015


What does this gene/protein do?
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Pathways:What pathways are this gene/protein implicaed in?
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Cancer Overview

Research Indicators

Publications Per Year (1990-2015)
Graph generated 21 August 2015 using data from PubMed using criteria.

Literature Analysis

Mouse over the terms for more detail; many indicate links which you can click for dedicated pages about the topic.

  • Base Sequence
  • Loss of Heterozygosity
  • Tumor Markers
  • beta Catenin
  • Single Nucleotide Polymorphism
  • Genetic Predisposition
  • Hepatocellular Carcinoma
  • Liver Cancer
  • Trans-Activators
  • Cancer Gene Expression Regulation
  • APC
  • Adenocarcinoma
  • Gene Expression Profiling
  • Polymerase Chain Reaction
  • Cytoskeletal Proteins
  • Chromosome 16
  • DNA Mutational Analysis
  • Adolescents
  • Wnt Proteins
  • Proteins
  • Signal Transduction
  • Cancer DNA
  • Intercellular Signaling Peptides and Proteins
  • Transcription Factors
  • Axin Protein
  • DNA Primers
  • DNA Methylation
  • Single-Stranded Conformational Polymorphism
  • Colorectal Cancer
  • DNA Sequence Analysis
  • Adenomatous Polyposis Coli Protein
  • Repressor Proteins
  • Western Blotting
  • Proto-Oncogene Proteins
  • Mutation
  • Cell Nucleus
  • TCF Transcription Factors
  • Missense Mutation
  • Immunohistochemistry
Tag cloud generated 21 August, 2015 using data from PubMed, MeSH and CancerIndex

Specific Cancers (2)

Data table showing topics related to specific cancers and associated disorders. Scope includes mutations and abnormal protein expression.

Note: list is not exhaustive. Number of papers are based on searches of PubMed (click on topic title for arbitrary criteria used).

Latest Publications: AXIN1 (cancer-related)

Schulze K, Imbeaud S, Letouzé E, et al.
Exome sequencing of hepatocellular carcinomas identifies new mutational signatures and potential therapeutic targets.
Nat Genet. 2015; 47(5):505-11 [PubMed] Related Publications
Genomic analyses promise to improve tumor characterization to optimize personalized treatment for patients with hepatocellular carcinoma (HCC). Exome sequencing analysis of 243 liver tumors identified mutational signatures associated with specific risk factors, mainly combined alcohol and tobacco consumption and exposure to aflatoxin B1. We identified 161 putative driver genes associated with 11 recurrently altered pathways. Associations of mutations defined 3 groups of genes related to risk factors and centered on CTNNB1 (alcohol), TP53 (hepatitis B virus, HBV) and AXIN1. Analyses according to tumor stage progression identified TERT promoter mutation as an early event, whereas FGF3, FGF4, FGF19 or CCND1 amplification and TP53 and CDKN2A alterations appeared at more advanced stages in aggressive tumors. In 28% of the tumors, we identified genetic alterations potentially targetable by US Food and Drug Administration (FDA)-approved drugs. In conclusion, we identified risk factor-specific mutational signatures and defined the extensive landscape of altered genes and pathways in HCC, which will be useful to design clinical trials for targeted therapy.

Yu N, Kakunda M, Pham V, et al.
HSP105 recruits protein phosphatase 2A to dephosphorylate β-catenin.
Mol Cell Biol. 2015; 35(8):1390-400 [PubMed] Article available free on PMC after 01/10/2015 Related Publications
The Wnt/β-catenin pathway causes accumulation of β-catenin in the cytoplasm and its subsequent translocation into the nucleus to initiate the transcription of the target genes. Without Wnt stimulation, β-catenin forms a complex with axin (axis inhibitor), adenomatous polyposis coli (APC), casein kinase 1α (CK1α), and glycogen synthase kinase 3β (GSK3β) and undergoes phosphorylation-dependent ubiquitination. Phosphatases, such as protein phosphatase 2A (PP2A), interestingly, also are components of this degradation complex; therefore, a balance must be reached between phosphorylation and dephosphorylation. How this balance is regulated is largely unknown. Here we show that a heat shock protein, HSP105, is a previously unidentified component of the β-catenin degradation complex. HSP105 is required for Wnt signaling, since depletion of HSP105 compromises β-catenin accumulation and target gene transcription upon Wnt stimulation. Mechanistically, HSP105 depletion disrupts the integration of PP2A into the β-catenin degradation complex, favoring the hyperphosphorylation and degradation of β-catenin. HSP105 is overexpressed in many types of tumors, correlating with increased nuclear β-catenin protein levels and Wnt target gene upregulation. Furthermore, overexpression of HSP105 is a prognostic biomarker that correlates with poor overall survival in breast cancer patients as well as melanoma patients participating in the BRIM2 clinical study.

Zhou J, Zheng B, Ji J, et al.
LYTAK1, a novel TAK1 inhibitor, suppresses KRAS mutant colorectal cancer cell growth in vitro and in vivo.
Tumour Biol. 2015; 36(5):3301-8 [PubMed] Related Publications
KRAS mutation in colorectal cancer (CRC) activates transforming growth factor-β (TGF-β)-activated kinase 1 (TAK1) to promote tumor progression. In the current study, we explored the potential effect of LYTAK1, a novel TAK1 inhibitor, against KRAS mutant CRC cells in vitro and in vivo. We found that LYTAK1 dose-dependently inhibited KRAS mutant CRC cell (HT-29 and SW-620 lines) growth, and induced cell cycle G1-S arrest. Further, LYTAK1 activated apoptosis in HT-29 cells and SW-620 cells, and apoptosis inhibitors almost reversed LYTAK1-mediated growth inhibition. While in KRAS wild-type (WT) CRC cell lines (DLD-1 and HCT-116), LYTAK1 had almost no effect on cell growth, cell cycle progression, or cell apoptosis. In KRAS mutant HT-29 cells and SW-260 cells, LYTAK1 blocked TAK1 activation or phosphorylation at Thr-184/187. Activation of nuclear factor κB (NF-κB) in these cells, detected by phosphorylations of p65 and IκB kinase α (IKKα) as well as expression of NF-κB-regulated gene cyclin D1, was significantly inhibited by LYTAK1. Further, LYTAK1 treatment resulted in downregulation of β-catenin and Wnt response gene Axin 2, indicating Wnt inactivation. In vivo, oral LYTAK1 significantly inhibited HT-29 xenograft growth in nude mice. Together, these results show that LYTAK1 inhibits KRAS mutant CRC cell growth both in vitro and in vivo. LYTAK1 might be investigated as a novel agent against CRC with KRAS mutation.

Tang Q, Zou Z, Zou C, et al.
MicroRNA-93 suppress colorectal cancer development via Wnt/β-catenin pathway downregulating.
Tumour Biol. 2015; 36(3):1701-10 [PubMed] Related Publications
MicroRNA-93 (miR-93) is involved in several carcinoma progressions. It has been reported that miR-93 acts as a promoter or suppressor in different tumors. However, till now, the role of miR-93 in colon cancer is unclear. Herein, we have found that expression of miR-93 was lower in human colon cancer tissue and colorectal carcinoma cell lines compared with normal colon mucosa. Forced expression of miR-93 in colon cancer cells inhibits colon cancer invasion, migration, and proliferation. Furthermore, miR-93 may downregulate the Wnt/β-catenin pathway, which was confirmed by measuring the expression level of the β-catenin, axin, c-Myc, and cyclin-D1 in this pathway. Mothers against decapentaplegic homolog 7 (Smad7), as an essential molecular protein for nuclear accumulation of β-catenin in the canonical Wnt signaling pathway, is predicted as a putative target gene of miR-93 by the silico method and demonstrated that it may be suppressed by targeting its 3'UTR. These findings showed that miR-93 suppresses colorectal cancer development via downregulating Wnt/β-catenin, at least in part, by targeting Smad7. This study revealed that miR-93 is an important negative regulator in colon cancer and suggested that miR-93 may serve as a novel therapeutic agent that offers benefits for colon cancer treatment.

Xu W, Wang Z, Zhang W, et al.
Mutated K-ras activates CDK8 to stimulate the epithelial-to-mesenchymal transition in pancreatic cancer in part via the Wnt/β-catenin signaling pathway.
Cancer Lett. 2015; 356(2 Pt B):613-27 [PubMed] Related Publications
Cyclin-dependent kinase 8 (CDK8), a gene encoding the cyclin-dependent kinase (CDK) component of the Mediator complex, is known as a colon cancer oncogene. Our recent study showed that CDK8 plays an important role in the formation of pancreatic cancer, but the CDK8 expression levels were not completely identical in different pancreatic cancer samples. The level of CDK8 expression depended on whether the K-ras gene was mutated; its expression was much higher in samples carrying a K-ras mutation than in wild-type K-ras samples. Moreover, CDK8 expression was reduced following mutated K-ras knockdown in K-ras-mutated pancreatic cancer cells, whereas CDK8 expression was increased following expression of mutated K-ras in wild-type K-ras cells. Our study demonstrates that mutated K-ras stimulates CDK8 expression, possibly by regulating HIF-1α, and both CDK8 and mutated K-ras were confirmed to promote cell proliferation and prevent apoptosis in vitro. Additionally, we found that both CDK8 and mutated K-ras promote the invasion and migration of pancreatic cancer cells via the positive regulation of the Wnt/β-catenin signaling pathway, thereby increasing the expression of Snail1 and ZEB1, which act as important stimulating factors of the epithelial-to-mesenchymal transition (EMT). Finally, knockdown of either CDK8 or mutated K-ras contributed to attenuated pancreatic cancer growth in BALB/c nude mice. In conclusion, these findings demonstrate that mutated K-ras promotes CDK8 expression and that the regulatory effects of CDK8 on the EMT are partially attributed to the Wnt/β-catenin signaling pathway.

Nomura R, Saito T, Mitomi H, et al.
GNAS mutation as an alternative mechanism of activation of the Wnt/β-catenin signaling pathway in gastric adenocarcinoma of the fundic gland type.
Hum Pathol. 2014; 45(12):2488-96 [PubMed] Related Publications
Gastric adenocarcinoma of the fundic gland type (GAFG) is a rare variant of gastric tumor. We have recently reported the frequent accumulation of β-catenin in GAFGs and showed that approximately half of the cases studied harbored at least 1 mutation in CTNNB1/AXINs/APC, leading to the constitutive activation of the Wnt/β-catenin pathway. However, the mechanisms of Wnt signaling activation in the remaining cases are unknown. Accumulating evidence showed that the activating mutation in GNAS promotes tumorigenesis via the activation of the Wnt/β-catenin pathway or the ERK1/2 MAPK pathway. Therefore, we analyzed the mutations in GNAS (exons 8 and 9) and in KRAS (exon 2) in 26 GAFGs. Immunohistochemistry revealed nuclear β-catenin expression in 22 of 26 GAFGs, and 10 (38.5%) of 26 cases harbored at least 1 mutation in CTNNB1/AXINs/APC. Activating mutations in GNAS were found in 5 (19.2%) of 26 GAFGs, all of which harbored R201C mutations. Activating mutations in KRAS were found in 2 (7.7%) of 26 GAFGs, and both of these also contained GNAS activating mutations. Four of 5 cases with GNAS mutation showed nuclear β-catenin expression, and presence of GNAS mutation was associated with β-catenin nuclear expression (P = .01). Furthermore, 3 of these 4 cases did not harbor mutations in CTNNB1, APC, or AXINs, suggesting that mutations in the Wnt component genes and those in GNAS occur almost exclusively. These results suggest that GNAS mutation might occur in a small subset of GAFG as an alternative mechanism of activating the Wnt/β-catenin signaling pathway.

Mazzoni SM, Fearon ER
AXIN1 and AXIN2 variants in gastrointestinal cancers.
Cancer Lett. 2014; 355(1):1-8 [PubMed] Article available free on PMC after 01/10/2015 Related Publications
Mutations in the APC (adenomatous polyposis coli) gene, which encodes a multi-functional protein with a well-defined role in the canonical Wnt pathway, underlie familial adenomatous polypsosis, a rare, inherited form of colorectal cancer (CRC) and contribute to the majority of sporadic CRCs. However, not all sporadic and familial CRCs can be explained by mutations in APC or other genes with well-established roles in CRC. The AXIN1 and AXIN2 proteins function in the canonical Wnt pathway, and AXIN1/2 alterations have been proposed as key defects in some cancers. Here, we review AXIN1 and AXIN2 sequence alterations reported in gastrointestinal cancers, with the goal of vetting the evidence that some of the variants may have key functional roles in cancer development.

Jhunjhunwala S, Jiang Z, Stawiski EW, et al.
Diverse modes of genomic alteration in hepatocellular carcinoma.
Genome Biol. 2014; 15(8):436 [PubMed] Article available free on PMC after 01/10/2015 Related Publications
BACKGROUND: Hepatocellular carcinoma (HCC) is a heterogeneous disease with high mortality rate. Recent genomic studies have identified TP53, AXIN1, and CTNNB1 as the most frequently mutated genes. Lower frequency mutations have been reported in ARID1A, ARID2 and JAK1. In addition, hepatitis B virus (HBV) integrations into the human genome have been associated with HCC.
RESULTS: Here, we deep-sequence 42 HCC patients with a combination of whole genome, exome and transcriptome sequencing to identify the mutational landscape of HCC using a reasonably large discovery cohort. We find frequent mutations in TP53, CTNNB1 and AXIN1, and rare but likely functional mutations in BAP1 and IDH1. Besides frequent hepatitis B virus integrations at TERT, we identify translocations at the boundaries of TERT. A novel deletion is identified in CTNNB1 in a region that is heavily mutated in multiple cancers. We also find multiple high-allelic frequency mutations in the extracellular matrix protein LAMA2. Lower expression levels of LAMA2 correlate with a proliferative signature, and predict poor survival and higher chance of cancer recurrence in HCC patients, suggesting an important role of the extracellular matrix and cell adhesion in tumor progression of a subgroup of HCC patients.
CONCLUSIONS: The heterogeneous disease of HCC features diverse modes of genomic alteration. In addition to common point mutations, structural variations and methylation changes, there are several virus-associated changes, including gene disruption or activation, formation of chimeric viral-human transcripts, and DNA copy number changes. Such a multitude of genomic events likely contributes to the heterogeneous nature of HCC.

Liu D, Li L, Yang Y, et al.
The Axin2 rs2240308 polymorphism and susceptibility to lung cancer in a Chinese population.
Tumour Biol. 2014; 35(11):10987-91 [PubMed] Related Publications
Axis inhibition protein 2 (Axin2) is a negative regulator of the canonical Wnt/β-catenin signaling pathway, and functions as a tumor suppressor in a number of human cancers. Previous pilot studies have suggested an association between Axin2 exon1 148 (rs2240308) SNP polymorphism and risk for lung cancer. In the present study, we aimed to investigate the Axin2 exon1 148 polymorphism and its association with lung cancer susceptibility in Han Chinese population. The Axin2 exon1 148 SNP was genotyped in 555 controls and 520 lung cancer patients using TaqMan SNP Genotyping Assays. Unconditional logistic regression analysis was used to calculate adjusted odds ratios (ORs) and 95% confidence intervals (CIs). We observed that the genotype frequencies of TC, TT, and CC were significantly different between controls and cases (χ(2) = 6.849, P = 0.03256, df = 2). Subjects carrying T allele (TC + TT genotypes) had decreased susceptibility to lung cancer as compared to those carrying CC genotype (OR = 0.733, 95% CI = 0.5726-0.9393, P = 0.01382). No significant association was found between rs2240308 polymorphism and histological subtypes of lung cancers. Findings from this study suggest that Axin2 exon1 T148C polymorphism (rs2240308) contributes to increased susceptibility to lung cancer in Chinese population. This further implicates Axin2 as a lung cancer-related gene.

Zhao Z, Lu P, Zhang H, et al.
Nestin positively regulates the Wnt/β-catenin pathway and the proliferation, survival and invasiveness of breast cancer stem cells.
Breast Cancer Res. 2014; 16(4):408 [PubMed] Article available free on PMC after 01/10/2015 Related Publications
INTRODUCTION: We investigated Nestin expression in triple-negative breast cancer and examined how the modulation of Nestin expression affects cell cycle progression, survival, invasion and regulatory signaling in breast cancer stem cells (CSC) in vitro.
METHODS: Nestin expression in 150 triple-negative breast cancer specimens were examined by immunohistochemistry. The role of Nestin expression in tumorigenesis was examined by assaying naturally occurring Nestinhigh/Nestinlow CSC from 12 breast cancer tissues, as well as CSC from 26 clinical specimens, where Nestin overexpression and silencing was achieved by genetic manipulation, for their ability to form mammospheres and induce solid tumors. Cell cycle progression, spontaneous apoptosis and invasiveness of Nestin-silenced breast CSC were investigated by flow cytometry and transwell assays. The relative levels of expression of epithelial-mesenchymal transition (EMT) and Wnt/β-catenin pathway-related molecules were determined by western blotting.
RESULTS: Nestin expression was significantly associated with poor survival in patients with triple-negative breast cancer (P = 0.01). Nestinhigh breast CSC rapidly formed typical mammospheres in vitro. Nestinhigh, but not Nestinlow CSC, efficiently formed solid tumors in vivo. Nestin silencing induced cell cycle arrest at G2/M (52.03% versus 19.99% in controls) and promoted apoptosis (36.45% versus 8.29% in controls). Nestin silencing also inhibited breast CSC invasiveness, and was associated with significantly upregulated E-cadherin, while N-cadherin, vimentin, a-smooth muscle actin (a-SMA), matrix metalloproteinase-2 (MMP-2), MMP-9 and vascular endothelial growth factor (VEGF) expression was downregulated (P <0.05 for all). Nestin silencing also upregulated Axin, glycogen synthase kinase-3 beta (GSK-3β), adenomatous polyposis coli (APC), and peroxisome proliferator-activated receptor alpha (PPARa), and downregulated β-catenin, c-Myc, cyclin D and MMP-7 expression in CSC. Inhibition of the Wnt/β-catenin pathway mitigated mammosphere formation in Nestinhigh CSC, while inhibition of GSK-3β promoted the mammosphere formation in Nestinlow CSC (P <0.05 for all).
CONCLUSIONS: Our data indicates that Nestin positively regulates the proliferation, survival and invasiveness of breast CSC by enhancing Wnt/β-catenin activation.

Lee DG, Kim HS, Lee YS, et al.
Helicobacter pylori CagA promotes Snail-mediated epithelial-mesenchymal transition by reducing GSK-3 activity.
Nat Commun. 2014; 5:4423 [PubMed] Related Publications
Cytotoxin-associated gene A (CagA) is an oncoprotein and a major virulence factor of H. pylori. CagA is delivered into gastric epithelial cells via a type IV secretion system and causes cellular transformation. The loss of epithelial adhesion that accompanies the epithelial-mesenchymal transition (EMT) is a hallmark of gastric cancer. Although CagA is a causal factor in gastric cancer, the link between CagA and the associated EMT has not been elucidated. Here, we show that CagA induces the EMT by stabilizing Snail, a transcriptional repressor of E-cadherin expression. Mechanistically we show that CagA binds GSK-3 in a manner similar to Axin and causes it to shift to an insoluble fraction, resulting in reduced GSK-3 activity. We also find that the level of Snail protein is increased in H. pylori infected epithelium in clinical samples. These results suggest that H. pylori CagA acts as a pathogenic scaffold protein that induces a Snail-mediated EMT via the depletion of GSK-3.

Amaddeo G, Cao Q, Ladeiro Y, et al.
Integration of tumour and viral genomic characterizations in HBV-related hepatocellular carcinomas.
Gut. 2015; 64(5):820-9 [PubMed] Article available free on PMC after 01/10/2015 Related Publications
BACKGROUND AND AIM: Hepatocellular carcinoma (HCC) is the most common liver cancer. We characterised HCC associated with infection compared with non-HBV-related HCC to understand interactions between viral and hepatocyte genomic alterations and their relationships with clinical features.
METHODS: Frozen HBV (n=86) or non-HBV-related (n=90) HCC were collected in two French surgical departments. Viral characterisation was performed by sequencing HBS and HBX genes and quantifying HBV DNA and cccDNA. Nine genes were screened for somatic mutations and expression profiling of 37 genes involved in hepatocarcinogenesis was studied.
RESULTS: HBX revealed frequent non-sense, frameshift and deletions in tumours, suggesting an HBX inactivation selected in HCC. The number of viral copies was frequently lower in tumour than in non-tumour tissues (p=0.0005) and patients with low HBV copies in the non-tumour liver tissues presented additional risk factor (HCV, alcohol or non-alcoholic steato-hepatitis, p=0.006). P53 was the most frequently altered pathway in HBV-related HCC (47%, p=0.001). Furthermore, TP53 mutations were associated with shorter survival only in HBV-related HCC (p=0.02) whereas R249S mutations were identified exclusively in migrants. Compared with other aetiologies, HBV-HCC were more frequently classified in tumours subgroups with upregulation of genes involved in cell-cycle regulation and a progenitor phenotype. Finally, in HBV-related HCC, transcriptomic profiles were associated with specific gene mutations (HBX, TP53, IRF2, AXIN1 and CTNNB1).
CONCLUSIONS: Integrated genomic characterisation of HBV and non-HBV-related HCC emphasised the immense molecular diversity of HCC closely related to aetiologies that could impact clinical care of HCC patients.

Muto Y, Maeda T, Suzuki K, et al.
DNA methylation alterations of AXIN2 in serrated adenomas and colon carcinomas with microsatellite instability.
BMC Cancer. 2014; 14:466 [PubMed] Article available free on PMC after 01/10/2015 Related Publications
BACKGROUND: Recent work led to recognize sessile serrated adenomas (SSA) as precursor to many of the sporadic colorectal cancers with microsatellite instability (MSI). However, comprehensive analyses of DNA methylation in SSA and MSI cancer have not been conducted.
METHODS: With an array-based methylation sensitive amplified fragment length polymorphism (MS-AFLP) method we analyzed 8 tubular (TA) and 19 serrated (SSA) adenomas, and 14 carcinomas with (MSI) and 12 without (MSS) microsatellite instability. MS-AFLP array can survey relative differences in methylation between normal and tumor tissues of 9,654 DNA fragments containing all NotI sequences in the human genome.
RESULTS: Unsupervised clustering analysis of the genome-wide hypermethylation alterations revealed no major differences between or within these groups of benign and malignant tumors regardless of their location in intergenic, intragenic, promoter, or 3' end regions. Hypomethylation was less frequent in SSAs compared with MSI or MSS carcinomas. Analysis of variance of DNA methylation between these four subgroups identified 56 probes differentially altered. The hierarchical tree of this subset of probes revealed two distinct clusters: Group 1, mostly composed by TAs and MSS cancers with KRAS mutations; and Group 2 with BRAF mutations, which consisted of cancers with MSI and MLH1 methylation (Group 2A), and SSAs without MLH1 methylation (Group 2B). AXIN2, which cooperates with APC and β-catenin in Wnt signaling, had more methylation alterations in Group 2, and its expression levels negatively correlated with methylation determined by bisulfite sequencing. Within group 2B, low and high AXIN2 expression levels correlated significantly with differences in size (P = 0.01) location (P = 0.05) and crypt architecture (P = 0.01).
CONCLUSIONS: Somatic methylation alterations of AXIN2, associated with changes in its expression, stratify SSAs according to some clinico-pathological differences. We conclude that hypermethylation of MLH1, when occurs in an adenoma cell with BRAF oncogenic mutational activation, drives the pathway for MSI cancer by providing the cells with a mutator phenotype. AXIN2 inactivation may contribute to this tumorigenic pathway either by mutator phenotype driven frameshift mutations or by epigenetic deregulation contemporary with the unfolding of the mutator phenotype.

Yamulla RJ, Kane EG, Moody AE, et al.
Testing models of the APC tumor suppressor/β-catenin interaction reshapes our view of the destruction complex in Wnt signaling.
Genetics. 2014; 197(4):1285-302 [PubMed] Article available free on PMC after 01/10/2015 Related Publications
The Wnt pathway is a conserved signal transduction pathway that contributes to normal development and adult homeostasis, but is also misregulated in human diseases such as cancer. The tumor suppressor adenomatous polyposis coli (APC) is an essential negative regulator of Wnt signaling inactivated in >80% of colorectal cancers. APC participates in a multiprotein "destruction complex" that targets the proto-oncogene β-catenin for ubiquitin-mediated proteolysis; however, the mechanistic role of APC in the destruction complex remains unknown. Several models of APC function have recently been proposed, many of which have emphasized the importance of phosphorylation of high-affinity β-catenin-binding sites [20-amino-acid repeats (20Rs)] on APC. Here we test these models by generating a Drosophila APC2 mutant lacking all β-catenin-binding 20Rs and performing functional studies in human colon cancer cell lines and Drosophila embryos. Our results are inconsistent with current models, as we find that β-catenin binding to the 20Rs of APC is not required for destruction complex activity. In addition, we generate an APC2 mutant lacking all β-catenin-binding sites (including the 15Rs) and find that a direct β-catenin/APC interaction is also not essential for β-catenin destruction, although it increases destruction complex efficiency in certain developmental contexts. Overall, our findings support a model whereby β-catenin-binding sites on APC do not provide a critical mechanistic function per se, but rather dock β-catenin in the destruction complex to increase the efficiency of β-catenin destruction. Furthermore, in Drosophila embryos expressing some APC2 mutant transgenes we observe a separation of β-catenin destruction and Wg/Wnt signaling outputs and suggest that cytoplasmic retention of β-catenin likely accounts for this difference.

Park S, Yun E, Hwang IH, et al.
Ilimaquinone and ethylsmenoquinone, marine sponge metabolites, suppress the proliferation of multiple myeloma cells by down-regulating the level of β-catenin.
Mar Drugs. 2014; 12(6):3231-44 [PubMed] Article available free on PMC after 01/10/2015 Related Publications
Deregulation of Wnt/β-catenin signaling promotes the development of a broad range of human cancers, including multiple myeloma, and is thus a potential target for the development of therapeutics for this disease. Here, we used a cell-based reporter system to demonstrate that ilimaquinone and ethylsmenoquinone (formerly smenorthoquinone), sesquiterpene-quinones from a marine sponge, inhibited β-catenin response transcription induced with Wnt3a-conditioned medium, by down-regulating the level of intracellular β-catenin. Pharmacological inhibition of glycogen synthase kinase-3β did not abolish the ilimaquinone and ethylsmenoquinone-mediated β-catenin down-regulation. Degradation of β-catenin was consistently found in RPMI-8226 multiple myeloma cells after ilimaquinone and ethylsmenoquinone treatment. Ilimaquinone and ethylsmenoquinone repressed the expression of cyclin D1, c-myc, and axin-2, which are β-catenin/T-cell factor-dependent genes, and inhibited the proliferation of multiple myeloma cells. In addition, ilimaquinone and ethylsmenoquinone significantly induced G0/G1 cell cycle arrest and apoptosis in RPMI-8266 cells. These findings suggest that ilimaquinone and ethylsmenoquinone exert their anti-cancer activity by blocking the Wnt/β-catenin pathway and have significant potential as therapies for multiple myeloma.

De Robertis A, Mennillo F, Rossi M, et al.
Human Sarcoma growth is sensitive to small-molecule mediated AXIN stabilization.
PLoS One. 2014; 9(5):e97847 [PubMed] Article available free on PMC after 01/10/2015 Related Publications
Sarcomas are mesenchymal tumors showing high molecular heterogeneity, reflected at the histological level by the existence of more than fifty different subtypes. Genetic and epigenetic evidences link aberrant activation of the Wnt signaling to growth and progression of human sarcomas. This phenomenon, mainly accomplished by autocrine loop activity, is sustained by gene amplification, over-expression of Wnt ligands and co-receptors or epigenetic silencing of endogenous Wnt antagonists. We previously showed that pharmacological inhibition of Wnt signaling mediated by Axin stabilization produced in vitro and in vivo antitumor activity in glioblastoma tumors. Here, we report that targeting different sarcoma cell lines with the Wnt inhibitor/Axin stabilizer SEN461 produces a less transformed phenotype, as supported by modulation of anchorage-independent growth in vitro. At the molecular level, SEN461 treatment enhanced the stability of the scaffold protein Axin1, a key negative regulator of the Wnt signaling with tumor suppressor function, resulting in downstream effects coherent with inhibition of canonical Wnt signaling. Genetic phenocopy of small molecule Axin stabilization, through Axin1 over-expression, coherently resulted in strong impairment of soft-agar growth. Importantly, sarcoma growth inhibition through pharmacological Axin stabilization was also observed in a xenograft model in vivo in female CD-1 nude mice. Our findings suggest the usefulness of Wnt inhibitors with Axin stabilization activity as a potentialyl clinical relevant strategy for certain types of sarcomas.

Ahn SM, Jang SJ, Shim JH, et al.
Genomic portrait of resectable hepatocellular carcinomas: implications of RB1 and FGF19 aberrations for patient stratification.
Hepatology. 2014; 60(6):1972-82 [PubMed] Related Publications
UNLABELLED: Hepatic resection is the most curative treatment option for early-stage hepatocellular carcinoma, but is associated with a high recurrence rate, which exceeds 50% at 5 years after surgery. Understanding the genetic basis of hepatocellular carcinoma at surgically curable stages may enable the identification of new molecular biomarkers that accurately identify patients in need of additional early therapeutic interventions. Whole exome sequencing and copy number analysis was performed on 231 hepatocellular carcinomas (72% with hepatitis B viral infection) that were classified as early-stage hepatocellular carcinomas, candidates for surgical resection. Recurrent mutations were validated by Sanger sequencing. Unsupervised genomic analyses identified an association between specific genetic aberrations and postoperative clinical outcomes. Recurrent somatic mutations were identified in nine genes, including TP53, CTNNB1, AXIN1, RPS6KA3, and RB1. Recurrent homozygous deletions in FAM123A, RB1, and CDKN2A, and high-copy amplifications in MYC, RSPO2, CCND1, and FGF19 were detected. Pathway analyses of these genes revealed aberrations in the p53, Wnt, PIK3/Ras, cell cycle, and chromatin remodeling pathways. RB1 mutations were significantly associated with cancer-specific and recurrence-free survival after resection (multivariate P = 0.038 and P = 0.012, respectively). FGF19 amplifications, known to activate Wnt signaling, were mutually exclusive with CTNNB1 and AXIN1 mutations, and significantly associated with cirrhosis (P = 0.017).
CONCLUSION: RB1 mutations can be used as a prognostic molecular biomarker for resectable hepatocellular carcinoma. Further study is required to investigate the potential role of FGF19 amplification in driving hepatocarcinogenesis in patients with liver cirrhosis and to investigate the potential of anti-FGF19 treatment in these patients.

Arend RC, Londoño-Joshi AI, Samant RS, et al.
Inhibition of Wnt/β-catenin pathway by niclosamide: a therapeutic target for ovarian cancer.
Gynecol Oncol. 2014; 134(1):112-20 [PubMed] Related Publications
Objective. The Wnt/β-catenin pathway is known to regulate cellular proliferation and plays a role in chemoresistance. Niclosamide, an FDA approved salicyclamide derivative used for the treatment of tapeworm infections, targets the Wnt/β-catenin pathway. Therefore, the objective of this study was to investigate niclosamide as a potential therapeutic agent for ovarian cancer. Methods. Tumor cells isolated from 34 patients' ascites with primary ovarian cancer were treated with niclosamide (0.1 to 5 μM) ± carboplatin (5 to 150 μM). Cell viability was assessed using the ATP-lite assay. LRP6, Axin 2, Cyclin D1, survivin and cytosolic free β-catenin levels were determined using Western blot analysis. Tumorspheres were treated, and Wnt transcriptional activity was measured by the TOPflash reporter assay. ALDH and CD133 were analyzed by Flow cytometry and IHC. ALDH1A1 and LRP6 were analyzed by IHC in solid tumor and in ascites before and after treatment with niclosamide. Results. Combination treatment produced increased cytotoxicity compared to single agent treatment in 32/34 patient samples. Western blot analysis showed a decrease in Wnt/β-catenin pathway proteins and the expression of target genes. A significant reduction of Wnt/β-catenin signaling was confirmed by TOPflash assay. There was increased staining of ALDH1A1 and LRP6 in ascites compared to solid tumor which decreased after treatment. Conclusion. This study demonstrates that niclosamide is a potent Wnt/β-catenin inhibitor. Targeting the Wnt/β-catenin pathway led to decreased cellular proliferation and increased cell death. These findings warrant further research of this drug and other niclosamide analogs as a treatment option for ovarian cancer.

Schneikert J, Ruppert JG, Behrens J, Wenzel EM
different Roles for the axin interactions with the SAMP versus the second twenty amino acid repeat of adenomatous polyposis coli.
PLoS One. 2014; 9(4):e94413 [PubMed] Article available free on PMC after 01/10/2015 Related Publications
Wnt signalling is prevented by the proteosomal degradation of β-catenin, which occurs in a destruction complex containing adenomatous polyposis coli (APC), APC-like (APCL), Axin and Axin2. Truncating mutations of the APC gene result in the constitutive stabilisation of β-catenin and the initiation of colon cancer, although tumour cells tolerate the expression of wild-type APCL. Using the colocalisation of overexpressed Axin, APC and APCL constructs as a readout of interaction, we found that Axin interacted with the second twenty amino acid repeat (20R2) of APC and APCL. This interaction involved a domain adjacent to the C-terminal DIX domain of Axin. We identified serine residues within the 20R2 of APCL that were involved in Axin colocalisation, the phosphorylation of truncated APCL and the down-regulation of β-catenin. Our results indicated that Axin, but not Axin2, displaced APC, but not APCL, from the cytoskeleton and stimulated its incorporation into bright cytoplasmic dots that others have recognised as β-catenin destruction complexes. The SAMP repeats in APC interact with the N-terminal RGS domain of Axin. Our data showed that a short domain containing the first SAMP repeat in truncated APC was required to stimulate Axin oligomerisation. This was independent of Axin colocalisation with 20R2. Our data also suggested that the RGS domain exerted an internal inhibitory constraint on Axin oligomerisation. Considering our data and those from others, we discuss a working model whereby β-catenin phosphorylation involves Axin and the 20R2 of APC or APCL and further processing of phospho-β-catenin occurs upon the oligomerisation of Axin that is induced by binding the SAMP repeats in APC.

Goksel G, Bilir A, Uslu R, et al.
WNT1 gene expression alters in heterogeneous population of prostate cancer cells; decreased expression pattern observed in CD133+/CD44+ prostate cancer stem cell spheroids.
J BUON. 2014 Jan-Mar; 19(1):207-14 [PubMed] Related Publications
PURPOSE: Established cancer cell lines contain cancer stem cells (CSCs) which can propagate to form three dimensional (3D) tumor spheroids in vitro. Aberrant activation of WNT signaling is strongly implicated in the progression of cancer and controls CSCs properties. In this study we hypothesized that when cells were maintained as spheroids, the structure of CSCs could show differentiation between CSCs and non- CSCs.
METHODS: CD133+/CD44+ cancer-initiating cells were isolated from DU-145 human prostate cancer cell line monolayer cultures, propagated as tumor spheroids and compared with the remaining heterogeneous cancer cells bulk population. The expression levels of WNT1, FZD1, ADAR, APC, AXIN, BTRC, FRAT1 and PPARD genes were measured by polymerase chain reaction (PCR) array assay and the protein expression levels of WNT1, FZD and AXIN by immunohistochemistry.
RESULTS: The expression levels of WNT pathway-related molecules were found to increase in both CSCs and non- CSCs when CSCs were maintained as spheroids. However, different expression profiles were observed when CSCs and non-CSCs were compared. In spheroids, the expression levels of FZD1, APC, ADAR, WNT1, PPARD genes in CSCs decreased when compared to non-CSCs. Interestingly, when CSCs from spheroids were compared with CSCs from monolayers the most significant decrease was observed in FZD1 and increase in APC genes.
CONCLUSION: It is possible to assume that intracellular signaling of WNT-related molecules in the nucleus and/or cytoplasm might play an important role but it is independent from increased ligand expression and this expression strongly differentiate CSCs and non-CSCs population. This unexpected expression could be important for CSCs behavior and targeting this pathway could have therapeutic implications in cancer.

Herbst A, Jurinovic V, Krebs S, et al.
Comprehensive analysis of β-catenin target genes in colorectal carcinoma cell lines with deregulated Wnt/β-catenin signaling.
BMC Genomics. 2014; 15:74 [PubMed] Article available free on PMC after 01/10/2015 Related Publications
BACKGROUND: Deregulation of Wnt/β-catenin signaling is a hallmark of the majority of sporadic forms of colorectal cancer and results in increased stability of the protein β-catenin. β-catenin is then shuttled into the nucleus where it activates the transcription of its target genes, including the proto-oncogenes MYC and CCND1 as well as the genes encoding the basic helix-loop-helix (bHLH) proteins ASCL2 and ITF-2B. To identify genes commonly regulated by β-catenin in colorectal cancer cell lines, we analyzed β-catenin target gene expression in two non-isogenic cell lines, DLD1 and SW480, using DNA microarrays and compared these genes to β-catenin target genes published in the PubMed database and DNA microarray data presented in the Gene Expression Omnibus (GEO) database.
RESULTS: Treatment of DLD1 and SW480 cells with β-catenin siRNA resulted in differential expression of 1501 and 2389 genes, respectively. 335 of these genes were regulated in the same direction in both cell lines. Comparison of these data with published β-catenin target genes for the colon carcinoma cell line LS174T revealed 193 genes that are regulated similarly in all three cell lines. The overlapping gene set includes confirmed β-catenin target genes like AXIN2, MYC, and ASCL2. We also identified 11 Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways that are regulated similarly in DLD1 and SW480 cells and one pathway - the steroid biosynthesis pathway - was regulated in all three cell lines.
CONCLUSIONS: Based on the large number of potential β-catenin target genes found to be similarly regulated in DLD1, SW480 and LS174T cells as well as the large overlap with confirmed β-catenin target genes, we conclude that DLD1 and SW480 colon carcinoma cell lines are suitable model systems to study Wnt/β-catenin signaling and associated colorectal carcinogenesis. Furthermore, the confirmed and the newly identified potential β-catenin target genes are useful starting points for further studies.

Saito T, Mitomi H, Imamhasan A, et al.
Downregulation of sFRP-2 by epigenetic silencing activates the β-catenin/Wnt signaling pathway in esophageal basaloid squamous cell carcinoma.
Virchows Arch. 2014; 464(2):135-43 [PubMed] Related Publications
Basaloid squamous cell carcinoma (BSCC) of the esophagus is a rare variant of typical squamous cell carcinoma (SCC) associated with poor survival. A characteristic feature is nuclear accumulation of β-catenin, without a mutation of the gene. We studied the methylation status of Wnt antagonist genes, such as secreted frizzled-related protein (sFRP) gene family members, Wnt inhibitory factor-1 (WIF-1), Dickkopf-1 (Dkk-1), and human Dapper protein-1 (HDPR-1), and alterations of the APC, Axin1, and Axin2 genes in 30 cases of esophageal BSCC. β-catenin and sFRP (sFRP-1, sFRP-2, sFRP-4, sFRP-5) protein expression was examined by immunohistochemistry. APC, Axin1, and Axin2 gene mutations were detected in 3, 2, and 2 cases, respectively, and 6 cases (20 %) harbored at least 1 alteration in these genes. Methylation of the sFRP-2 promoter region was observed in all cases, and methylation was frequent in sFRP-1 and sFRP-5, but infrequent in Dkk-1, WIF-1, sFRP-4, and HDPR-1. sFRP-2 expression was almost completely absent in 25 cases (83 %), consistent with the methylation status. Nuclear accumulation of β-catenin was observed in all cases. sFRP-5 expression was associated with a low nuclear β-catenin labeling index. These results show that sFRP-2 is a target gene of hypermethylation in esophageal BSCC and suggest that sFRP-2 might contribute to BSCC tumorigenesis through the Wnt/β-catenin signaling pathway.

Gao Y, Song C, Hui L, et al.
Overexpression of RNF146 in non-small cell lung cancer enhances proliferation and invasion of tumors through the Wnt/β-catenin signaling pathway.
PLoS One. 2014; 9(1):e85377 [PubMed] Article available free on PMC after 01/10/2015 Related Publications
Studies have suggested a possible correlation between the newly identified E3 ubiquitin ligase ring finger protein 146 (RNF146) and tumor development. However, until now, studies on RNF146 have been restricted to poly(ADP-ribosyl)ation and ubiquitin ligation, whereas the role of RNF146 in tumor biology has rarely been reported. In the present study, the role of RNF146 in non-small cell lung cancer (NSCLC) was investigated. The results showed that the expression of RNF146 was increased in clinical lung cancer samples and cell lines. RNF146 expression correlated with tumor size, differentiation level, lymphatic metastasis, pTNM staging, and prognosis of patients in stage I. RNF146 expression was negatively correlated with Axin expression but positively correlated with the nuclear expression of β-catenin in NSCLC tissues. RNF146 downregulated the expression of Axin in lung cancer cell lines and induced the expression and nuclear distribution of β-catenin. Overexpression of RNF146 in NSCLC cell lines increased the levels of cyclinD1, cyclinE, and CDK4, promoted cell cycle G0/G1-S transitions, and regulated cell proliferation. Overexpression of RNF146 led to upregulated levels of matrix metalloproteinases 2 and 7 and enhanced lung cancer cell invasiveness, events that were mediated by the classical Wnt/β-catenin signaling pathway. In summary, the data in the present study indicate that RNF146 regulated the development and progression of NSCLC by enhancing cell growth, invasion, and survival, suggesting that RNF146 may be a potential treatment target in NSCLC.

Ju X, Ishikawa TO, Naka K, et al.
Context-dependent activation of Wnt signaling by tumor suppressor RUNX3 in gastric cancer cells.
Cancer Sci. 2014; 105(4):418-24 [PubMed] Related Publications
RUNX3 is a tumor suppressor for a variety of cancers. RUNX3 suppresses the canonical Wnt signaling pathway by binding to the TCF4/β-catenin complex, resulting in the inhibition of binding of the complex to the Wnt target gene promoter. Here, we confirmed that RUNX3 suppressed Wnt signaling activity in several gastric cancer cell lines; however, we found that RUNX3 increased the Wnt signaling activity in KatoIII and SNU668 gastric cancer cells. Notably, RUNX3 expression increased the ratio of the Wnt signaling-high population in the KatoIII cells. although the maximum Wnt activation level of individual cells was similar to that in the control. As found previously, RUNX3 also binds to TCF4 and β-catenin in KatoIII cells, suggesting that these molecules form a ternary complex. Moreover, the ChIP analyses revealed that TCF4, β-catenin and RUNX3 bind the promoter region of the Wnt target genes, Axin2 and c-Myc, and the occupancy of TCF4 and β-catenin in these promoter regions is increased by the RUNX3 expression. These results suggest that RUNX3 stabilizes the TCF4/β-catenin complex on the Wnt target gene promoter in KatoIII cells, leading to activation of Wnt signaling. Although RUNX3 increased the Wnt signaling activity, its expression resulted in suppression of tumorigenesis of KatoIII cells, indicating that RUNX3 plays a tumor-suppressing role in KatoIII cells through a Wnt-independent mechanism. These results indicate that RUNX3 can either suppress or activate the Wnt signaling pathway through its binding to the TCF4/β-catenin complex by cell context-dependent mechanisms.

Pita JM, Figueiredo IF, Moura MM, et al.
Cell cycle deregulation and TP53 and RAS mutations are major events in poorly differentiated and undifferentiated thyroid carcinomas.
J Clin Endocrinol Metab. 2014; 99(3):E497-507 [PubMed] Related Publications
BACKGROUND: Anaplastic thyroid carcinomas (ATCs) are among the most lethal malignancies, for which there is no effective treatment.
OBJECTIVE: In the present study, we aimed to elucidate the molecular alterations contributing to ATC development and to identify novel therapeutic targets.
DESIGN: We profiled the global gene expression of five ATCs and validated differentially expressed genes by quantitative RT-PCR in an independent set of tumors. In a series of 26 ATCs, we searched for pathogenic alterations in genes involved in the most deregulated cellular processes, including the hot spot regions of RAS, BRAF, TP53, CTNNB1 (β-catenin), and PIK3CA genes, and, for the first time, a comprehensive analysis of components involved in the cell cycle [cyclin-dependent kinase (CDK) inhibitors (CDKI): CDKN1A (p21(CIP1)); CDKN1B (p27(KIP1)); CDKN2A (p14(ARF), p16(INK4A)); CDKN2B (p15(INK4B)); CDKN2C (p18(INK4C))], cell adhesion (AXIN1), and proliferation (PTEN). Mutational analysis was also performed in 22 poorly differentiated thyroid carcinomas (PDTCs).
RESULTS: Expression profiling revealed that ATCs were characterized by the underexpression of epithelial components and the up regulation of mesenchymal markers and genes from TGF-β pathway, as well as, the overexpression of cell cycle-related genes. In accordance, the up regulation of the SNAI2 gene, a TGF-β-responsive mesenchymal factor, was validated. CDKN3, which prevents the G1/S transition, was significantly up regulated in ATCs and PDTCs and aberrantly spliced in ATCs. Mutational analysis showed that most mutations were present in TP53 (42% of ATCs; 27% of PDTCs) or RAS (31% of ATCs; 18% of PDTCs). TP53 and RAS alterations showed evidence of mutual exclusivity (P = .0354). PIK3CA, PTEN, and CDKI mutations were present in 14%-20% of PDTCs, and in 10%-14% of ATCs. BRAF, CTNNB1, and AXIN1 mutations were rarely detected.
CONCLUSION: Overall, this study identified crucial roles for TP53, RAS, CDKI, and TGF-β pathway, which may represent feasible therapeutic targets for ATC and PDTC treatment.

Gu DL, Chen YH, Shih JH, et al.
Target genes discovery through copy number alteration analysis in human hepatocellular carcinoma.
World J Gastroenterol. 2013; 19(47):8873-9 [PubMed] Article available free on PMC after 01/10/2015 Related Publications
High-throughput short-read sequencing of exomes and whole cancer genomes in multiple human hepatocellular carcinoma (HCC) cohorts confirmed previously identified frequently mutated somatic genes, such as TP53, CTNNB1 and AXIN1, and identified several novel genes with moderate mutation frequencies, including ARID1A, ARID2, MLL, MLL2, MLL3, MLL4, IRF2, ATM, CDKN2A, FGF19, PIK3CA, RPS6KA3, JAK1, KEAP1, NFE2L2, C16orf62, LEPR, RAC2, and IL6ST. Functional classification of these mutated genes suggested that alterations in pathways participating in chromatin remodeling, Wnt/β-catenin signaling, JAK/STAT signaling, and oxidative stress play critical roles in HCC tumorigenesis. Nevertheless, because there are few druggable genes used in HCC therapy, the identification of new therapeutic targets through integrated genomic approaches remains an important task. Because a large amount of HCC genomic data genotyped by high density single nucleotide polymorphism arrays is deposited in the public domain, copy number alteration (CNA) analyses of these arrays is a cost-effective way to reveal target genes through profiling of recurrent and overlapping amplicons, homozygous deletions and potentially unbalanced chromosomal translocations accumulated during HCC progression. Moreover, integration of CNAs with other high-throughput genomic data, such as aberrantly coding transcriptomes and non-coding gene expression in human HCC tissues and rodent HCC models, provides lines of evidence that can be used to facilitate the identification of novel HCC target genes with the potential of improving the survival of HCC patients.

Okolicsanyi RK, van Wijnen AJ, Cool SM, et al.
Heparan sulfate proteoglycans and human breast cancer epithelial cell tumorigenicity.
J Cell Biochem. 2014; 115(5):967-76 [PubMed] Article available free on PMC after 01/10/2015 Related Publications
Heparan sulfate proteoglycans (HSPGs) are key components of the extracellular matrix that mediate cell proliferation, invasion, and cellular signaling. The biological functions of HSPGs are linked to their co-stimulatory effects on extracellular ligands (e.g., WNTs) and the resulting activation of transcription factors that control mammalian development but also associated with tumorigenesis. We examined the expression profile of HSPG core protein syndecans (SDC1-4) and glypicans (GPC1-6) along with the enzymes that initiate or modify their glycosaminoglycan chains in human breast cancer (HBC) epithelial cells. Gene expression in relation to cell proliferation was examined in the HBC cell lines MCF-7 and MDA-MB-231 following treatment with the HS agonist heparin. Heparin increased gene expression of chain initiation and modification enzymes including EXT1 and NDST1, as well as core proteins SDC2 and GPC6. With HS/Wnt interactions established, we next investigated WNT pathway components and observed that increased proliferation of the more invasive MDA-MB-231 cells is associated with activation of the Wnt signaling pathway. Specifically, there was substantial upregulation (>5-fold) of AXIN1, WNT4A, and MYC in MDA-MB-231 but not in MCF-7 cells. The changes in gene expression observed for HSPG core proteins and related enzymes along with the associated Wnt signaling components suggest coordinated interactions. The influence of HSPGs on cellular proliferation and invasive potential of breast cancer epithelial cells are cell and niche specific. Further studies on the interactions between HSPGs and WNT ligands may yield clinically relevant molecular targets, as well as new biomarkers for characterization of breast cancer progression.

Stewart DJ
Wnt signaling pathway in non-small cell lung cancer.
J Natl Cancer Inst. 2014; 106(1):djt356 [PubMed] Related Publications
Wnt/β-catenin alterations are prominent in human malignancies. In non-small cell lung cancer (NSCLC), β-catenin and APC mutations are uncommon, but Wnt signaling is important in NSCLC cell lines, and Wnt inhibition reduces proliferation. Overexpression of Wnt-1, -2, -3, and -5a and of Wnt-pathway components Frizzled-8, Dishevelled, Porcupine, and TCF-4 is common in resected NSCLC and is associated with poor prognosis. Conversely, noncanonical Wnt-7a suppresses NSCLC development and is often downregulated. Although β-catenin is often expressed in NSCLCs, it was paradoxically associated with improved prognosis in some series, possibly because of E-cadherin interactions. Downregulation of Wnt inhibitors (eg, by hypermethylation) is common in NSCLC tumor cell lines and resected samples; may be associated with high stage, dedifferentiation, and poor prognosis; and has been reported for AXIN, sFRPs 1-5, WIF-1, Dkk-1, Dkk-3, HDPR1, RUNX3, APC, CDX2, DACT2, TMEM88, Chibby, NKD1, EMX2, ING4, and miR-487b. AXIN is also destabilized by tankyrases, and GSK3β may be inactivated through phosphorylation by EGFR. Preclinically, restoration of Wnt inhibitor function is associated with reduced Wnt signaling, decreased cell proliferation, and increased apoptosis. Wnt signaling may also augment resistance to cisplatin, docetaxel, and radiotherapy, and Wnt inhibitors may restore sensitivity. Overall, available data indicate that Wnt signaling substantially impacts NSCLC tumorigenesis, prognosis, and resistance to therapy, with loss of Wnt signaling inhibitors by promoter hypermethylation or other mechanisms appearing to be particularly important. Wnt pathway antagonists warrant exploration clinically in NSCLC. Agents blocking selected specific β-catenin interactions and approaches to increase expression of downregulated Wnt inhibitors may be of particular interest.

Rennoll SA, Konsavage WM, Yochum GS
Nuclear AXIN2 represses MYC gene expression.
Biochem Biophys Res Commun. 2014; 443(1):217-22 [PubMed] Article available free on PMC after 01/10/2015 Related Publications
The β-catenin transcriptional coactivator is the key mediator of the canonical Wnt signaling pathway. In the absence of Wnt, β-catenin associates with a cytosolic and multi-protein destruction complex where it is phosphorylated and targeted for proteasomal degradation. In the presence of Wnt, the destruction complex is inactivated and β-catenin translocates into the nucleus. In the nucleus, β-catenin binds T-cell factor (TCF) transcription factors to activate expression of c-MYC (MYC) and Axis inhibition protein 2 (AXIN2). AXIN2 is a member of the destruction complex and, thus, serves in a negative feedback loop to control Wnt/β-catenin signaling. AXIN2 is also present in the nucleus, but its function within this compartment is unknown. Here, we demonstrate that AXIN2 localizes to the nuclei of epithelial cells within normal and colonic tumor tissues as well as colorectal cancer cell lines. In the nucleus, AXIN2 represses expression of Wnt/β-catenin-responsive luciferase reporters and forms a complex with β-catenin and TCF. We demonstrate that AXIN2 co-occupies β-catenin/TCF complexes at the MYC promoter region. When constitutively localized to the nucleus, AXIN2 alters the chromatin structure at the MYC promoter and directly represses MYC gene expression. These findings suggest that nuclear AXIN2 functions as a rheostat to control MYC expression in response to Wnt/β-catenin signaling.

Liu J, Pan S, Hsieh MH, et al.
Targeting Wnt-driven cancer through the inhibition of Porcupine by LGK974.
Proc Natl Acad Sci U S A. 2013; 110(50):20224-9 [PubMed] Article available free on PMC after 01/10/2015 Related Publications
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.

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