Adenomatous Polyposis Coli Expression Research Articles

Upregulation of miR-3607 promotes lung adenocarcinoma proliferation by suppressing APC expression

Lung cancer is the leading cause of worldwide cancer-related deaths, although many drugs and new therapeutic approaches have been used, the 5-years survival rate is still low for lung cancer patients. microRNAs have been shown to regulate lung cancer initiation and development, here we studied the role of miR-3607 in lung cancer cell proliferation. We found miR-3607 was upregulated in lung cancer tissues and cells, miR-3607 overexpression promoted lung cancer cell A549 proliferation determined by MTT assay, colony formation assay, anchorage-independent growth ability assay and bromodeoxyuridine incorporation assay, while the opposite phenotypes were shown when miR-3607 was knocked down. Predicted analysis suggested a Wnt signaling pathway regulator adenomatous polyposis coli (APC) was the target of miR-3607, miR-3607 could directly bind to the 3′UTR of APC, and promoted Cyclin D1 and c-Myc expression which can be suppressed by APC. Double knockdown of miR-3607 and APC copied the phenotypes of miR-3607 overexpression, suggesting miR-3607 promoted lung cancer cell A549 proliferation by targeting APC. In conclusion, our study suggested miR-3607 contributes to lung cancer cell proliferation by inhibiting APC.

The effect of EBV on WIF1, NLK, and APC gene methylation and expression in gastric carcinoma and nasopharyngeal cancer.

Epstein-Barr virus (EBV) is an important DNA tumor virus that is associated with approximately 10% of gastric carcinomas and 99% of nasopharyngeal cancers (NPC). DNA methylation and microRNAs (miRNAs) are the most studied epigenetic mechanisms that can prompt disease susceptibility. This study aimed to detect the effect of EBV on Wnt inhibitory factor 1 (WIF1), Nemo-like kinase (NLK), and adenomatous polyposis coli (APC) gene methylation, and expression in gastric carcinoma and NPC. The WIF1, NLK, and APC gene mRNA expression levels were measured by real-time quantitative RT-PCR in four EBV-positive cell lines and four EBV-negative cell lines. Bisulfite genomic sequencing or methylation-specific PCR was used to detect the methylation status of the WIF1, NLK, and APC promoters. All cell lines were treated with 5-azacytidine (5-aza-dC), miR-BART19-3p mimics or an inhibitor, and analyzed by flow cytometry and MTT cell proliferation assays. The WIF1, NLK, and APC promoters were hypermethylated in all eight cell lines. 5-Aza-dC displayed a growth inhibitory effect on cells . After transfection with miR-BART19-3p mimics, the expression of WIF1, and APC decreased, and the cellular proliferation rate increased. After transfection with the miR-BART19-3p inhibitor, the expression levels were higher, and the cell growth was inhibited. In the NPC and GC cell lines, the promoters of WIF1, NLK, and APC are highly methylated, and the expression of these three genes is regulated by miR-BART19-3p. The activity of the Wnt pathway in EBV-associated tumors may be enhanced by miR-BART19-3p.

Adenomatous polyposis coli nucleates actin assembly to drive cell migration and microtubule-induced focal adhesion turnover.

Cell motility depends on tight coordination between the microtubule (MT) and actin cytoskeletons, but the mechanisms underlying this MT-actin cross talk have remained poorly understood. Here, we show that the tumor suppressor protein adenomatous polyposis coli (APC), which is a known MT-associated protein, directly nucleates actin assembly to promote directed cell migration. By changing only two residues in APC, we generated a separation-of-function mutant, APC (m4), that abolishes actin nucleation activity without affecting MT interactions. Expression of full-length APC carrying the m4 mutation (APC (m4)) rescued cellular defects in MT organization, MT dynamics, and mitochondrial distribution caused by depletion of endogenous APC but failed to restore cell migration. Wild-type APC and APC (m4) localized to focal adhesions (FAs), and APC (m4) was defective in promoting actin assembly at FAs to facilitate MT-induced FA turnover. These results provide the first direct evidence for APC-mediated actin assembly in vivo and establish a role for APC in coordinating MTs and actin at FAs to direct cell migration.

Effects of dual modified resistant indica rice starch on azoxymethane-induced incipient colon cancer in mice.

In this study, the effects of different doses of dual modification-treated (DMT) indica rice resistant starch (IR-RS) on azoxymethane (AOM)-induced early colon cancer in mice were investigated. The investigated factors included body weight, gastrointestinal emptying rate, the number and morphology of aberrant crypt foci (ACFs) and the specific expressions of adenomatous polyposis coli (APC), B-cell lymphoma 2 (Bcl-2), Bcl-2-associated X protein (Bax) and cytochrome c genes. The results demonstrated that DMT IR-RS controlled the increase in the body weights of the mice, increased the gastrointestinal emptying rates and reduced the numbers of ACFs and aberrant crypts. Reverse transcription-polymerase chain reaction revealed that DMT IR-RS promoted the expression of APC, Bax and cytochrome c and inhibited the expression of Bcl-2. These results demonstrate that a DMT IR-RS diet may induce apoptosis and has beneficial health effects in AOM-induced early colon cancer in mice.

Characterization of the number and site of APC mutations in sporadic colorectal cancer.

630 Background: Truncating mutations in the adenomatous polyposis coli ( APC) gene are well-described events in the carcinogenesis of colorectal carcinomas (CRC) and may impact one or both APC alleles. These aberrations often fall within the mutation cluster region (MCR) of the APC gene to preserve a “just right” number of beta-catenin binding sites in the resulting mutant APC protein. Further clinical and genotypic characterization of CRCs based on number and site of mutations in APC determined using modern next generation sequencing methods is needed. Methods: Next generation sequencing of 70 CRC tumors was performed at a single institution via UNCseq to determine mutations in a panel of 247 oncogenes and tumor suppressors, including APC. RNASeq, DNA sequencing, and clinical characteristics from 224 colon and rectal cancer samples in The Cancer Genome Atlas (TCGA) project were also obtained. Results: In the UNCseq cohort, 58 patients (83%) had at least one inactivating APC mutation, and 33 (47%) had two mutations. Of those with at least one mutation, 81% had a mutation in the MCR (residues 1281-1556), but only 5/33 (15%) with two mutations had both in the MCR. In the TCGA cohort, 162 (72%) had at least one inactivating APC mutation, and 52 (23%) had two mutations. Of those with at least one mutation, 59% had a mutation in the MCR, but only 3/52 (6%) with two mutations had both in the MCR. Gene expression of APC was higher in those with no APC mutations vs. 1-2 mutations (p = 0.015), but was not significantly different between those with one vs. two mutations (p = 0.29). The absence of APC mutations was associated with microsatellite instability (p < 0.001) and with right-sided primary tumors (p = 0.001 by chi-square). Conclusions: CRCs have high frequency of biallelic APC mutations, and the majority of tumors with APC mutations had a single mutation within the MCR region of the APC gene. These genotypic factors may impact tumor biology and clinical features.

WIF-1 gene inhibition and Wnt signal transduction pathway activation in NSCLC tumorigenesis

The aim of the present study is to explore the differential expression of key molecules associated with Wnt signaling in both clinical non-small cell lung cancer (NSCLC) tissue and adjacent normal lung tissue, and to discuss the tumorigenic role of the activation of Wnt signaling pathways in NSCLC. A total of 52 NSCLC patients were employed in the present study. Lung cancer tissue samples and paracarcinoma tissue samples were obtained from these patients, who had undergone surgical resection of their primary cancer. The cases were diagnosed by hematoxylin and eosin staining. Using reverse transcription-quantitative polymerase chain reaction and immunohistochemical straining, the messenger RNA (mRNA) and protein expression levels of Wnt inhibitory factor-1 (WIF-1) and important molecules associated with Wnt signaling pathways were detected. Compared with normal tissues, a marked decreased in the mRNA and protein expression levels of WIF-1, and an increase in β-catenin and cyclin D1 expression, were observed in tumor tissues. This suggests that the activation of the Wnt/β-catenin signaling pathway may be closely associated with lymph nodal metastasis and lower pathological classification. However, no obvious difference could be observed in adenomatous polyposis coli (APC) expression levels between lung cancer tissues and adjacent tissues to the carcinoma. The activation of the Wnt/β-catenin signaling pathway in NSCLC could be initiated by WIF-1 gene inhibition without APC expression changes, and this may be different to the mechanism in other tumors.

Inherited Variants in Wnt Pathway Genes Influence Outcomes of Prostate Cancer Patients Receiving Androgen Deprivation Therapy.

Aberrant Wnt signaling has been associated with many types of cancer. However, the association of inherited Wnt pathway variants with clinical outcomes in prostate cancer patients receiving androgen deprivation therapy (ADT) has not been determined. Here, we comprehensively studied the contribution of common single nucleotide polymorphisms (SNPs) in Wnt pathway genes to the clinical outcomes of 465 advanced prostate cancer patients treated with ADT. Two SNPs, adenomatous polyposis coli (APC) rs2707765 and rs497844, were significantly (p ≤ 0.009 and q ≤ 0.043) associated with both prostate cancer progression and all-cause mortality, even after multivariate analyses and multiple testing correction. Patients with a greater number of favorable alleles had a longer time to disease progression and better overall survival during ADT (p for trend ≤ 0.003). Additional, cDNA array and in silico analyses of prostate cancer tissue suggested that rs2707765 affects APC expression, which in turn is correlated with tumor aggressiveness and patient prognosis. This study identifies the influence of inherited variants in the Wnt pathway on the efficacy of ADT and highlights a preclinical rationale for using APC as a prognostic marker in advanced prostate cancer.

Activation of the Wnt/β-catenin pathway is common in wilms tumor, but rarely through β-catenin mutation and APC promoter methylation.

The Wnt/β-catenin pathway is known to be crucial for the regulation of embryogenesis and cell differentiation, and its constitutive activation is associated with a wide range of malignancies. There are two major principles for an activated Wnt/β-catenin pathway. The first is caused by the failure of the destruction complex, mainly due to the decreased expression of the tumor suppressor gene adenomatous polyposis coli (APC); the second is the mutation of the β-catenin (CTNNB1) protein itself. Wilms tumors (WTs) are also thought to be malignancies with a high rate of Wnt/β-catenin pathway activation. The aim of this study was to analyze a large cohort of WT for activated Wnt/β-catenin pathway. The transcription of axis inhibition protein 2 (AXIN2) and APC was analyzed by real-time PCR. Expression was compared with those in healthy renal tissues as a control. Methylation status of the APC promoter was measured by pyrosequencing and correlated with APC expression. Finally, the mutations of CTNNB1 itself were detected by Sanger sequencing. The analysis was done in a cohort of 103 WTs, treated in our institution. There was a significant overexpression of AXIN2 in WTs (P<0.0001), with 33 (32%) tumors showing higher expression (median+3× SD) than normal kidney tissue. In contrast, the expression of APC as well as its promoter methylation did not differ from control (P=0.78; P=0.82). Finally, there were only seven (6.8%) mutations detectable in CTNNB1, and five out of seven were seen in WTs with AXIN2 overexpression. The finding that AXIN2, one of the major Wnt target genes, is overexpressed in our cohort of WTs, is indicative for the activation of the Wnt/β-catenin pathway. However, neither the alteration of APC nor frequent CTNNB1 mutations were seen in our analyses. Therefore, other mechanisms might be responsible for the common activation of the Wnt/β-catenin pathway.

Suppression of β-catenin Signaling Pathway in Human Prostate Cancer PC3 Cells by Delphinidin.

Delphinidin possesses strong anti-oxidant, anti-inflammatory, and anti-cancer properties. Suppression of the Wnt/β-catenin signaling pathway is a potential strategy for chemoprevention and therapy. As aberrant activation of the β-catenin signaling pathway contributes to prostate cancer progression, we evaluated the effect of delphinidin on this pathway in human PC3 prostate cancer cells. An MTT assay showed that treatment with delphinidin (15–180 μM, 72 hours) resulted in a dose-dependent growth inhibition of cells. Treatment with delphinidin increased the phosphorylation of serine or threonine residues on β-catenin and decreased the levels of cytoplasmic β-catenin. Moreover, treatment with delphinidin inhibited the nuclear translocation of β-catenin and the expression of β-catenin target genes such as cyclin D1, c-myc, Axin-2, and T cell factor-1. Delphinidin also induced the phosphorylation of glycogen synthase kinase 3β and the expression of adenomatous polyposis coli and Axin proteins. Our results indicate that inhibition of cell growth by delphinidin is mediated, at least in part, through modulation of the β-catenin signaling pathway. We suggest that delphinidin is a potent inhibitor of Wnt/β-catenin signaling in prostate cancer cells.

Interaction between APC and Fen1 during breast carcinogenesis.

Aberrant DNA base excision repair (BER) contributes to malignant transformation. However, inter-individual variations in DNA repair capacity plays a key role in modifying breast cancer risk. We review here emerging evidence that two proteins involved in BER - adenomatous polyposis coli (APC) and flap endonuclease 1 (Fen1) - promote the development of breast cancer through novel mechanisms. APC and Fen1 expression and interaction is increased in breast tumors versus normal cells, APC interacts with and blocks Fen1 activity in Pol-β-directed LP-BER, and abrogation of LP-BER is linked with cigarette smoke condensate-induced transformation of normal breast epithelial cells. Carcinogens increase expression of APC and Fen1 in spontaneously immortalized human breast epithelial cells, human colon cancer cells, and mouse embryonic fibroblasts. Since APC and Fen1 are tumor suppressors, an increase in their levels could protect against carcinogenesis; however, this does not seem to be the case. Elevated Fen1 levels in breast and lung cancer cells may reflect the enhanced proliferation of cancer cells or increased DNA damage in cancer cells compared to normal cells. Inactivation of the tumor suppressor functions of APC and Fen1 is due to their interaction, which may act as a susceptibility factor for breast cancer. The increased interaction of APC and Fen1 may occur due to polypmorphic and/or mutational variation in these genes. Screening of APC and Fen1 polymorphic and/or mutational variations and APC/Fen1 interaction may permit assessment of individual DNA repair capability and the risk for breast cancer development. Such individuals might lower their breast cancer risk by reducing exposure to carcinogens. Stratifying individuals according to susceptibility would greatly assist epidemiologic studies of the impact of suspected environmental carcinogens. Additionally, a mechanistic understanding of the interaction of APC and Fen1 may provide the basis for developing new and effective targeted chemopreventive and chemotherapeutic agents.

Chemopreventive activity of GEN-27, a genistein derivative, in colitis-associated cancer is mediated by p65-CDX2-β-catenin axis.

Nonresolving inflammation in the intestine predisposes individuals to colitis-associated colorectal cancer (CAC), which leads to high morbidity and mortality. Here we show that genistein-27 (GEN-27), a derivative of genistein, inhibited proliferation of human colorectal cancer cells through inhibiting β-catenin activity. Our results showed that GEN-27 increased expressions of adenomatous polyposis coli (APC) and axis inhibition protein 2 (AXIN2), and reduced β-catenin nuclear localization, which resulted from the inhibition of NF-κB/p65 nuclear localization and up-regulation of caudal-related homeobox transcription factor 2 (CDX2). Furthermore, GEN-27 decreased binding of p65 to the silencer region of CDX2 and increased binding of CDX2 to the promoter regions of APC and AXIN2, thus inhibiting the activation of β-catenin induced by TNF-α. Importantly, GEN-27 protected mice from azoxymethane (AOM)/dextran sodium sulfate (DSS)-induced colon carcinogenesis, with reduced mortality, tumor number and tumor volume. Histopathology, immunohistochemistry and flow cytometry revealed that dietary GEN-27 significantly decreased secretion of proinflammatory cytokines and macrophage infiltration. Moreover, GEN-27 inhibited AOM/DSS-induced p65 and β-catenin nuclear translocation, while promoted the expression of CDX2, APC, and AXIN2. Taken together, our findings demonstrate that the anti-proliferation effect of GEN-27 in vitro and the prevention of CAC in vivo is mediated by p65-CDX2-β-catenin axis via inhibiting β-catenin target genes. Our results imply that GEN-27 could be a promising candidate for the chemoprevention of CAC.

Adenomatous polyposis coli regulates radial axonal sorting and myelination in the PNS.

The tumor suppressor protein adenomatous polyposis coli (APC) is multifunctional - it participates in the canonical Wnt/β-catenin signal transduction pathway as well as modulating cytoskeleton function. Although APC is expressed by Schwann cells, the role that it plays in these cells and in the myelination of the peripheral nervous system (PNS) is unknown. Therefore, we used the Cre-lox approach to generate a mouse model in which APC expression is specifically eliminated from Schwann cells. These mice display hindlimb weakness and impaired axonal conduction in sciatic nerves. Detailed morphological analyses revealed that APC loss delays radial axonal sorting and PNS myelination. Furthermore, APC loss delays Schwann cell differentiation in vivo, which correlates with persistent activation of the Wnt signaling pathway and results in perturbed extension of Schwann cell processes and disrupted lamellipodia formation. In addition, APC-deficient Schwann cells display a transient diminution of proliferative capacity. Our data indicate that APC is required by Schwann cells for their timely differentiation to mature, myelinating cells and plays a crucial role in radial axonal sorting and PNS myelination.

Immunohistochemical analysis of the Wnt/β-catenin signaling pathway in pancreatic neuroendocrine neoplasms.

To investigate the role of the Wnt/β-catenin pathway in pancreatic neuroendocrine neoplasms (PanNENs). Tissue microarrays containing 88 PanNENs were immunohistochemically labeled with antibodies to β-catenin, E-cadherin, adenomatous polyposis coli (APC), chromogranin and synaptophysin. One case had only metastatic tumors resected, whereas others (n = 87) received pancreatectomy with or without partial hepatectomy. Pathology slides, demographic, clinicopathologic, and follow up data were reviewed. Patients' demographics, clinicopathologic features, and immunohistochemical results from 87 primary tumors were compared between patients with low stage (stage I/II) and high stage (stage III/IV) tumors. In addition, correlation of immunohistochemical results from primary tumors with disease-specific survival (DSS) was evaluated. Strong membranous β-catenin staining in the primary tumor was observed in all 13 stage III/IV PanNENs as compared to 47% (35/74) of stage I/II tumors (P < 0.01). However, the strong membranous β-catenin staining was unassociated with tumor grade or DSS. Decreased membranous β-catenin staining was associated with decreased membranous E-cadherin labeling. Nuclear β-catenin staining was seen in 15% (2/13) of stage III/IV PanNENs as compared to 0% (0/74) of stage I/II tumors (P = 0.02). The case with metastasectomy only also showed nuclear β-catenin staining. Two of the three cases with nuclear β-catenin staining were familial adenomatous polyposis (FAP) patients. Lack of APC expression was seen in 70% (57/81) of the cases, including the 3 cases with nuclear β-catenin staining. Expression of E-cadherin and APC in primary tumor was not correlated with tumor grade, tumor stage, or disease specific survival. The Wnt/β-catenin pathway was altered in some PanNENs, but did not Impact DSS. PanNENs in FAP patients demonstrated nuclear β-catenin accumulation and loss of APC.

Expression of adenomatous polyposis coli in primary gastric lymphoma tissue and its relationship with helicobacter pylori

Objective To investigate the expression and significance of adenomatous polyposis coli (APC) protein in primary gastric lymphoma (PGL) tissue, and explore its relationship with H. pylori. Methods The clinical and pathological data of 45 patients with PGL admitted between June 2006 and December 2011 in Xiangya Hospital and Shaoyang Chinese Medicine Hospital were collected. Expression of APC protein was detected with immunohistochemical staining (SP) in 45 cases (32 cases were male and 13 cases female, 33 cases were less than 60 years old, 12 cases were equal to or more than 60 years old, 35 cases were Hp positive, and 10 cases Hp negative), and 30 samples of normal gastric mucosa tissues. Results The positive expression rate of APC protein was 37.78% (17/45) in PGLs, and 90.0% (27/30) in normal gastric mucosa tissues. There was significant difference between PGLs and normal gastric mucosa tissues (P<0.05). The positive expression rate of APC protein in PGL was 25.71% (9/35) in those with Hp infection, and 80.00% (8/10) without Hp infection. There was significant correlation between the expression of APC and Hp infection (P<0.05). Conclusions The APC protein was lowly expressed in PGL. Because of the significant correlation between low expression of APC protein and Hp infection, Hp infection might be an important factor that impacts on the expression of APC protein. Key words: Lymphoma/PA; Helicobacter infections; Adenomatous polyposis coli/ME; Adenomatous polyposis coli protein/ME

Correlation between mutations and mRNA expression of APC and MUTYH genes: new insight into hereditary colorectal polyposis predisposition.

BackgroundTranscript dosage imbalance may influence the transcriptome. To gain insight into the role of altered gene expression in hereditary colorectal polyposis predisposition, in the present study we analyzed absolute and allele-specific expression (ASE) of adenomatous polyposis coli (APC) and mutY Homolog (MUTYH) genes.MethodsWe analyzed DNA and RNA extracted from peripheral blood mononuclear cells (PBMC) of 49 familial polyposis patients and 42 healthy blood donors selected according similar gender and age. Patients were studied for germline alterations in both genes using dHPLC, MLPA and automated sequencing. APC and MUTYH mRNA expression levels were investigated by quantitative Real-Time PCR (qRT-PCR) analysis using TaqMan assay and by ASE assays using dHPLC-based primer extension.ResultsTwenty out of 49 patients showed germline mutations: 14 in APC gene and six in MUTYH gene. Twenty-nine patients did not show mutations in both genes. Results from qRT-PCR indicated that gene expression of both APC and MUTYH was reduced in patients analyzed. In particular, a significant reduction in APC expression was observed in patients without APC germline mutation vs control group (P < 0.05) while APC expression in the mutation carrier patients, although lower compared to control individuals, did not show statistical significance. On the other hand a significant reduced MUTYH expression was detected in patients with MUTYH mutations vs control group (P < 0.05). Altered ASE of APC was detected in four out of eight APC mutation carriers. In particular one case showed a complete loss of one allele. Among APC mutation negative cases, 4 out of 13 showed a moderate ASE. ASE of MUTYH did not show any altered expression in the cases analyzed. Spearman’s Rho Test analysis showed a positive and significant correlation between APC and MUTYH genes both in cases and in controls (P = 0.020 and P < 0.001).ConclusionsAPC and MUTYH showed a reduced germline expression, not always corresponding to gene mutation. Expression of APC is decreased in mutation negative cases and this appears to be a promising indicator of FAP predisposition, while for MUTYH gene, mutation is associated to reduced mRNA expression. This study could improve the predictive genetic diagnosis of at-risk individuals belonging to families with reduced mRNA expression regardless of presence of mutation.Electronic supplementary materialThe online version of this article (doi:10.1186/s13046-015-0244-4) contains supplementary material, which is available to authorized users.

Effect of miRNA-135b on proliferation, invasion and migration of triple-negative breast cancer by targeting APC

To explore the expression of miRNA-135b in a variety of breast cancer cell lines and its function on the proliferation, invasion and migration in triple-negative breast cancer cell lines by targeting adenomatous polyposis coli (APC). Quantitative real-time (RT)--PCR was used to detect the expression of miRNA-135b in seven breast cancer cell lines and one normal breast cell line. The three-negative breast cancer cell lines MDA-MB-231 and MDA-MB-468 with miRNA-135b high-expression were divided into three groups: the normal growth group, the negative control group (transfected with negative counter-part) and the experimental group (transfected with miRNA-135b inhibitors). RT- PCR and Western blot were used to explore the expression of APC. The proliferation was detected by Cell Counting Kit-8 (CCK-8) method and the invasion and migration were detected by Transwell method. The levels of miRNA-135b were higher in triple-negative breast cancer cell lines than other types, especially MDA-MB-231 (P=0.001) and MDA-MB-468 (P=0.036). Compared with normal growth group and negative control group, the mRNA and protein of APC were up-regulated in experimental group MDA-MB-231 (P=0.013, P=0.021), MDA-MB-468 (P=0.017, P=0.014). CCK-8 results showed that the proliferation rate of experimental group was decreased compared with negative control and normal growth group MDA-MB-231 (P=0.00 ), MDA-MB-468 (P=0.01). The invasion and migration ability of MDA-MB-231 (P=0.002, P=0.00) and MDA-MB-468 (P=0.01, P=0.00) were obviously decreased after transfected miRNA-135b inhibitors. The expressions of miRNA-135b were higher in most triple-negative breast cancer cell lines than others. miRNA-135b could promote the proliferation, invasion and migration in triple-negative breast cancer cell lines MDA-MB-231 and MDA-MB-468, and APC was one of the target genes of miRNA- 135b by participating in the process of regulation.

Adenomatous polyposis coli mutants dominantly activate Hsf1-dependent cell stress pathways through inhibition of microtubule dynamics.

Cancer cells up-regulate cell stress pathways, including the protein chaperone Hsp90. Increases in Hsp90 are believed "buffer" mutant protein activities necessary for cancer phenotypes. Activation of the cell stress pathway also alters the transcriptional landscape of cells in ways that are critical for cancer progression. However, it is unclear when and how the cell stress pathway is de-regulated during cancer progression. Here we report that mutations in adenomatous polyposis coli (APC) found in colorectal cancer activate cell stress pathways in mouse intestinal crypt cells, prior to loss of heterozygosity at APC or to the appearance of canonical intestinal cancer markers. Hsp90 levels are elevated in normal APC heterozygote crypt cells and further elevated in non-cancer cells adjacent to dysplasias, suggesting that the Hsp90 stress pathway marks the "cancer-field" effect. Expression of mutant APC in normal human epithelial cells is sufficient to activate a cell stress pathway via perturbations in microtubule dynamics. Inhibition of microtubule dynamics is sufficient to activate an Hsf1-dependent increase in gene transcription and protein levels. We suggest that the early activation of this Hsf1 dependent cell stress pathway by mono-allelic mutations in APC can affect cell programming in a way that contributes to cancer onset.

MicroRNA-663 activates the canonical Wnt signaling through the adenomatous polyposis coli suppression

Rheumatoid arthritis (RA) is a symmetrical polyarticular autoimmune disease of unknown etiology. In this present study, we observed that the adenomatous polyposis coli (APC) expression is down-regulated and the expression of microRNA (miR)-663 increased significantly in synovium from RA patients compared with control. Target gene prediction for miR-663 revealed that the mRNA of APC gene, which is a member of the canonical Wnt signaling pathway, has a miR-663 binding site in its 3′-untranslated region (3′UTR). The result showed that increased miR-663 suppressed the APC expression significantly, and this down-regulation of APC expression triggered the activation of canonical Wnt signaling through accumulation of β-catenin in fibroblast-like synoviocytes (FLS). In addition, increased miR-663 induced the FLS proliferation and the expression MMP3 and fibronectin during disease development. Therefore, miR-663 can be considered as a critical regulator of RA pathogenesis and can be utilized for developing miRNA-based therapeutic agents for RA patients.

Expression of wingless-type mouse mammary tumor virus integration site family pathway effectors in lymphatic and hepatic metastases of patients with colorectal cancer: Associations with the primary tumor.

The wingless-type mouse mammary tumor virus integration site family (Wnt) pathway plays a major role in the carcinogenesis of colorectal cancer (CRC). Its most important effector, the nuclear β-catenin, influences not only transcription but also the proliferation and dedifferentiation of the colonic mucosa. This induces an epithelial-mesenchymal transition which ultimately can lead to the development of cancer and the formation of metastases. However, little is known about the exact interaction and context-sensitive expression of Wnt-pathway effectors in the primary tumor and corresponding metastasis. Therefore, this study assessed the expression of the three most important effectors of the Wnt pathway, β-catenin, adenomatous polyposis coli (APC) and Wnt-1, in the primary tumor and corresponding metastasis of patients with CRC. Immunohistochemical staining of β-catenin, APC and Wnt-1 was performed in paraffin-embedded tissue samples of the primary tumor, and the corresponding hepatic and nodal metastasis samples from 24 patients with metastatic CRC. Isotype antibodies were used as negative controls. The results were visualized using the ABC-method. Analysis of the primary tumor comprised of a separate evaluation of the central tumor area as well as the invasion front. There was a significant overexpression of nuclear β-catenin at the tumor invasion front (P<0.001). Compared to normal colonic mucosa, expression of cytoplasmic β-catenin was significantly higher in the primary tumor (P<0.001) as well as all the corresponding hepatic and lymphatic metastases (hepatic metastases, P=0.001; nodal metastases, P=0.017). By contrast, APC expression was significantly lower in all analyzed tumor compartments compared with normal colonic mucosa (primary tumor, P=0.022; hepatic metastases, P=0.006; nodal metastases, P=0.012). Finally, Wnt-1 protein expression was significantly lower in liver metastases but not in the primary tumor or lymphatic metastases compared with normal colonic mucosa (P=0.003). The present study demonstrates that the major Wnt-effector proteins, β-catenin, APC and Wnt-1, are heterogeneously expressed in the primary tumor and corresponding hepatic as well as nodal metastases of patients with CRC. This context-sensitive diverse expression of Wnt-effector proteins may be important for future individualized targeted therapies.

LGR5 promotes the proliferation of colorectal cancer cells via the Wnt/β-catenin signaling pathway.

Leucine-rich repeat-containing G-protein coupled receptor 5 (LGR5) is an established cancer stem cell marker and is a target gene of the Wnt/β-catenin signaling pathway, a critical pathway in the process of tumor initiation and growth. In the present study, the mRNA expression levels of LGR5, adenomatous polyposis coli (APC) and β-catenin were detected in 20 colorectal cancer (CRC) tissues and matched healthy mucosa samples using reverse transcription-quantitative polymerase chain reaction. HT-29 CRC cell line was treated with siRNA-Lgr5; the APC, β-catenin and LGR5 RNA expressions were detected and cell viability was measured using a CCK8 assay. The results revealed that LGR5 was significantly overexpressed in CRC tissue compared with healthy mucosa (P<0.05). Furthermore, knockdown of LGR5 by small interfering RNA decreased the expression of APC and β-catenin in HT29 colon cancer cells as well as inhibited the proliferation of HT29 cells. These findings demonstrated that LGR5 expression is critical for the promotion of neoplastic CRC cell proliferation, indicating that LGR5 may be a novel therapeutic target for CRC.