The expression of β-catenin in different subtypes of breast cancer and its clinical significance

The Wnt/β-catenin signaling pathway is implicated in mammary oncogenesis. Reports of β-catenin expression and its association with outcome in breast cancer are controversial. This study was performed to address the distribution of β-catenin expression in invasive breast cancer and the correlation between β-catenin expression and survival of breast cancer patients, and to determine whether β-catenin was specifically activated in any molecular subtypes. Immunohistochemistry was performed on a tissue microarray containing 169 invasive breast cancers to detect expression of β-catenin. One hundred thirty one of the 169 patients were followed up. Correlation between β-catenin expression and different molecular subtypes was determined using chi-square analysis. Overall survival (OS) was analyzed by Kaplan-Meier method with log-rank test. The invasive breast cancer displayed the different patterns of β-catenin expression from normal tissues with significantly increased cytoplasmic and nuclear staining of β-catenin. Aberrant β-catenin expression was observed in 109 in the 169 cases (64.50 %), and there was no difference in β-catenin expression in the four molecular subtypes. Furthermore, aberrant β-catenin expression was significantly associated with adverse outcome not only in the entire cohort but also in each of the different molecular subtypes. β-catenin activation is preferentially found and is associated with a poor clinical outcome in invasive breast cancer independent of molecular subtype.     

Prognostic significance of Wnt-1, β-catenin and E-cadherin expression in advanced colorectal carcinoma

Wnt/β-catenin pathway plays an important role in initiation and progression of colorectal oncogenesis. The aim of this study was to determine expression and localization of E-cadherin, β-catenin and Wnt-1 proteins in colorectal tumors. Expression of β-catenin, E-cadherin and Wnt-1 was determined by immunohistochemistry on advanced colorectal cancers. Abnormal expression of E-cadherin, β-catenin, Wnt-1 was observed. Additionally, we revealed correlations between levels of studied proteins and histoclinical data. In multivariate analysis nuclear β-catenin, higher carcinoembryonic antigen serum level before treatment, female sex and tumor localized in colon or rectum were independent unfavorable prognostic factors. These findings support the hypothesis that Wnt/β-catenin pathway plays an important role in advanced colorectal carcinoma.     

Clinical–pathologic significance of cancer stem cell marker expression in familial breast cancers

Human breast cancer cells with a CD44⁺/CD24^?/low or ALDH1+ phenotype have been demonstrated to be enriched for cancer stem cells (CSCs) using in vitro and in vivo techniques. The aim of this study was to determine the association between CD44⁺/CD24^?/low and ALDH1 expression with clinical–pathologic tumor characteristics, tumor molecular subtype, and survival in a well characterized collection of familial breast cancer cases. 364 familial breast cancers from the Ontario Familial Breast Cancer Registry (58 BRCA1-associated, 64 BRCA2-associated, and 242 familial non-BRCA1/2 cancers) were studied. Each tumor had a centralized pathology review performed. TMA sections of all tumors were analyzed for the expression of ER, PR, HER2, CK5, CK14, EGFR, CD44, CD24, and ALDH1. The Chi square test or Fisher’s exact test was used to analyze the marker associations with clinical–pathologic tumor variables, molecular subtype and genetic subtype. Analyses of the association of overall survival (OS) with marker status were conducted using Kaplan–Meier plots and log-rank tests. The CD44⁺/CD24^?/low and ALDH1+ phenotypes were identified in 16% and 15% of the familial breast cancer cases, respectively, and associated with high-tumor grade, a high-mitotic count, and component features of the medullary type of breast cancer. CD44⁺/CD24^?/low and ALDH1 expression in this series were further associated with the basal-like molecular subtype and the CD44⁺/CD24^?/low phenotype was independently associated with BRCA1 mutational status. The currently accepted breast CSCs markers are present in a minority of familial breast cancers. Whereas the presence of these markers is correlated with several poor prognostic features and the basal-like subtype of breast cancer, they do not predict OS.     

CD133 and nuclear β-catenin: The marker combination to detect high risk cases of low stage colorectal cancer

Nuclear β-catenin and CD133 are linked with two hallmarks of colon cancer, wingless-type mouse mammary tumour virus integration site (WNT)-pathway dysregulation and colon cancer stem cells (Co-CSCs), respectively. Both molecules may be related, as Co-CSCs were proposed to require activated WNT-signalling and as CD133 was postulated as a WNT/β-catenin target gene. Herein, we investigated the expression of these markers on serial sections of 162 stage IIA colonic adenocarcinomas. We found that the expression of these molecules is statistically independent and that they mark distinct but overlapping subpopulations of the tumour cells. Moreover, we show that their combined evaluation can identify colon cancer cases with vastly reduced survival (hazard ratio (HR) 13.4, 95% confidence interval (CI): 4.7–38.2) and a high risk of tumour progression (HR 6.8, 95%CI: 3.1–15.0). In conclusion, the independence of these markers may on the one hand have implications for their presumed value to identify Co-CSCs; on the other hand it allows their combined analysis to become a powerful tool to identify high risk cases of stage IIA colon cancer.     

Xanthine oxidoreductase – Clinical significance in colorectal cancer and in vitro expression of the protein in human colon cancer cells

Xanthine oxidoreductase (XOR) is a key enzyme in degradation of DNA and RNA, and has previously been shown to be decreased in aggressive breast and gastric cancer. In this study, XOR expression was assessed in tissue microarray specimens of 478 patients with colorectal cancer and related to clinical parameters. In addition, we performed in vitro studies of XOR activity, protein and mRNA in colon cancer cells (Caco-2). Results from the tissue expression analyses show that XOR was decreased in 62% and undetectable in 22% of the tumours as compared to normal tissue. Loss of XOR was associated with poor grade of differentiation (p = 0.006) and advanced Dukes stage (p = 0.03). In multivariate survival analysis, XOR was a prognostic factor (p = 0.008), independent of Dukes stage, histological grade, age and tumour location. The in vitro analyses show that XOR is not measurable in undifferentiated Caco-2 cells, but appears and increases with differentiation. We conclude that XOR expression is associated with histological grade of differentiation and extent of disease in colorectal cancer, and it provides significant prognostic information independently of established factors.     

Clinical significance of axin and β-catenin protein expression in primary hepatocellular carcinomas

The aim of the present research was to investigate clinicopathologic correlations of immunohistochemically- demonstrated axin (axis inhibition) and β-catenin expression in primary hepatocellular carcinomas (HCCs), in comparison with paraneoplastic, cirrhotic and normal liver tissues. Variation in Axin expression across groups were significant (P < 0.01), correlating with alpha fetoprotein (AFP), HBsAg, cancer plugs in the portal vein, and clinical stage of HCCs(P < 0.05); however, there were no links with sex, age, and tumour size (P > 0.05). Differences in cell membrane β-catenin expression were also statistically significant (P < 0.01), again correlated with AFP, HBsAg, cancer plugs in the portal vein, and clinical stage in HCCs (P < 0.05) but not with sex, age, and tumour size (P > 0.05). Axin expression levels in tissues with reduced membrane β-catenin were low (P < 0.05), also being low with nuclear β-catenin expression (P < 0.05). Axin and β-catenin may play an important role in the genesis and progression of HCC via the Wnt signal transmission pathway. Simultaneous determination of axin, β-catenin, AFP, and HBsAg may be useful for early diagnosis, and metastatic and clinical staging of HCCs.     

Impact of mutant β-catenin on ABCB1 expression and therapy response in colon cancer cells

Background:

Colorectal cancers are often chemoresistant toward antitumour drugs that are substrates for ABCB1-mediated multidrug resistance (MDR). Activation of the Wnt/β-catenin pathway is frequently observed in colorectal cancers. This study investigates the impact of activated, gain-of-function β-catenin on the chemoresistant phenotype.

Methods:

The effect of mutant (mut) β-catenin on ABCB1 expression and promoter activity was examined using HCT116 human colon cancer cells and isogenic sublines harbouring gain-of-function or wild-type β-catenin, and patients'' tumours. Chemosensitivity towards 24 anticancer drugs was determined by high throughput screening.

Results:

Cell lines with mut β-catenin showed high ABCB1 promoter activity and expression. Transfection and siRNA studies demonstrated a dominant role for the mutant allele in activating ABCB1 expression. Patients'' primary colon cancer tumours shown to express the same mut β-catenin allele also expressed high ABCB1 levels. However, cell line chemosensitivities towards 24 MDR-related and non-related antitumour drugs did not differ despite different β-catenin genotypes.

Conclusion:

Although ABCB1 is dominantly regulated by mut β-catenin, this did not lead to drug resistance in the isogenic cell line model studied. In patient samples, the same β-catenin mutation was detected. The functional significance of the mutation for predicting patients'' therapy response or for individualisation of chemotherapy regimens remains to be established.     

The metastasis suppressor,NDRG1, inhibits “stemness” of colorectal cancer via down-regulation of nuclear β-catenin and CD44

N-myc downstream-regulated gene 1 (NDRG1), has been identified as an important metastasis suppressor for colorectal cancer (CRC). In this study, we investigated: (1) the effects of NDRG1 on CRC stemness and tumorigenesis; (2) the molecular mechanisms involved; and (3) the relationship between NDRG1 expression and colorectal cancer prognosis. Our investigation demonstrated that CRC cells with silenced NDRG1 showed more tumorigenic ability and stem cell-like properties, such as: colony and sphere formation, chemoresistance, cell invasion, high expression of CD44, and tumorigenicity in vivo. Moreover, NDRG1 silencing reduced β-catenin expression on the cell membrane, while increasing its nuclear expression. The anti-tumor activity of NDRG1 was demonstrated to be mediated by preventing β-catenin nuclear translocation, as silencing of this latter molecule could reverse the effects of silencing NDRG1 expression. NDRG1 expression was also demonstrated to be negatively correlated to CRC prognosis. In addition, there was a negative correlation between NDRG1 and nuclear β-catenin and also NDRG1 and CD44 expression in clinical CRC specimens. Taken together, our investigation demonstrates that the anti-metastatic activity of NDRG1 in CRC occurs through the down-regulation of nuclear β-catenin and suggests that NDRG1 is a significant therapeutic target.     

Nuclear β-catenin and CD44 upregulation characterize invasive cell populations in non-aggressive MCF-7 breast cancer cells

Background

In breast cancer cells, the metastatic cell state is strongly correlated to epithelial-to-mesenchymal transition (EMT) and the CD44⁺/CD24^- stem cell phenotype. However, the MCF-7 cell line, which has a luminal epithelial-like phenotype and lacks a CD44⁺/CD24^- subpopulation, has rare cell populations with higher Matrigel invasive ability. Thus, what are the potentially important differences between invasive and non-invasive breast cancer cells, and are the differences related to EMT or CD44/CD24 expression?

Methods

Throughout the sequential selection process using Matrigel, we obtained MCF-7-14 cells of opposite migratory and invasive capabilities from MCF-7 cells. Comparative analysis of epithelial and mesenchymal marker expression was performed between parental MCF-7, selected MCF-7-14, and aggressive mesenchymal MDA-MB-231 cells. Furthermore, using microarray expression profiles of these cells, we selected differentially expressed genes for their invasive potential, and performed pathway and network analysis to identify a set of interesting genes, which were evaluated by RT-PCR, flow cytometry or function-blocking antibody treatment.

Results

MCF-7-14 cells had enhanced migratory and invasive ability compared with MCF-7 cells. Although MCF-7-14 cells, similar to MCF-7 cells, expressed E-cadherin but neither vimentin nor fibronectin, β-catenin was expressed not only on the cell membrane but also in the nucleus. Furthermore, using gene expression profiles of MCF-7, MCF-7-14 and MDA-MB-231 cells, we demonstrated that MCF-7-14 cells have alterations in signaling pathways regulating cell migration and identified a set of genes (PIK3R1, SOCS2, BMP7, CD44 and CD24). Interestingly, MCF-7-14 and its invasive clone CL6 cells displayed increased CD44 expression and downregulated CD24 expression compared with MCF-7 cells. Anti-CD44 antibody treatment significantly decreased cell migration and invasion in both MCF-7-14 and MCF-7-14 CL6 cells as well as MDA-MB-231 cells.

Conclusions

MCF-7-14 cells are a novel model for breast cancer metastasis without requiring constitutive EMT and are categorized as a "metastable phenotype", which can be distinguished from both epithelial and mesenchymal cells. The alterations and characteristics of MCF-7-14 cells, especially nuclear β-catenin and CD44 upregulation, may characterize invasive cell populations in breast cancer.

     

APC and β-catenin protein expression patterns in HNPCC-related endometrial and colorectal cancers

Objective: The adenomatous polyposis coli (APC) and β-catenin (CTNNB1) genes are the two major components of the Wnt signaling pathway that has been shown to play an important role in the formation of certain cancers. The overactivation of the pathway, which results in abnormal accumulation of β-catenin protein in nuclei, contributes to most colorectal cancers (CRCs), both sporadic and hereditary, as well as sporadic endometrial cancers (ECs). Here, we studied the involvement of APC and β-catenin in hereditary nonpolyposis colorectal cancer (HNPCC)-related ECs, and compared the expression patterns to those in HNPCC-related CRCs. Materials and methods: Nineteen ECs and 31 CRCs derived from HNPCC patients were immunohistochemically stained with anti-APC- and anti-β-catenin –antibodies. Results: Tumor-specific loss of APC was observed in 16 of endometrial cancers (3 of 19) and in 39 of colorectal cancers (12 of 31). Consistently, the loss of APC expression was associated with nuclear β-catenin staining. Altogether, aberrant β-catenin localization was observed in 53 of ECs (10 of 19) as compared to 84 of CRCs (26 of 31) (P=0.02).Conclusion: Our results suggest a frequent overactivation of the Wnt signaling pathway in hereditary endometrial cancer. In accordance with studies on sporadic cancers, abnormal accumulation of β-catenin protein in nuclei occurred much less frequently in HNPCC-related ECs than CRCs, which may reflect organ-specific differences in their pathogenesis.     

Oxymatrine diminishes the side population and inhibits the expression of β-catenin in MCF-7 breast cancer cells

Cancer stem cells (CSCs) play a critical role in both cancer initiation and relapse as they are resistant to most cytotoxic agents and able to proliferate indefinitely. The plant alkaloid oxymatrine has many biological activities including the ability to induce cell cycle arrest and apoptosis, which makes it a potentially useful agent for targeting cancer cells. In order to determine whether it has beneficial pharmacological properties to eradicate CSCs, we analyzed the effects of oxymatrine on MCF-7 breast cancer cells. Cancer stem-like cells' (side population, SP) identification and sorting were performed. The inhibitory effect of oxymatrine was evaluated on the sorted SP and non-SP cells. The results indicated that oxymatrine caused a dose-dependent reduction in the proliferation of MCF-7 cells and a decrease in SP cells. Wnt/β-catenin signaling pathway was also examined by analyzing the expression of total β-catenin and phosphorylated β-catenin in cytoplasm, and the results showed that the growth inhibitory effects of oxymatrine treatment on MCF-7 cells may be due to the inhibition of SP and Wnt/β-catenin signaling pathway. Further work is warranted to explore whether oxymatrine may be a useful novel therapeutic drug for targeting breast CSCs.     

Oxymatrine diminishes the side population and inhibits the expression of β-catenin in MCF-7 breast cancer cells

Cancer stem cells (CSCs) play a critical role in both cancer initiation and relapse as they are resistant to most cytotoxic agents and able to proliferate indefinitely. The plant alkaloid oxymatrine has many biological activities including the ability to induce cell cycle arrest and apoptosis, which makes it a potentially useful agent for targeting cancer cells. In order to determine whether it has beneficial pharmacological properties to eradicate CSCs, we analyzed the effects of oxymatrine on MCF-7 breast cancer cells. Cancer stem-like cells’ (side population, SP) identification and sorting were performed. The inhibitory effect of oxymatrine was evaluated on the sorted SP and non-SP cells. The results indicated that oxymatrine caused a dose-dependent reduction in the proliferation of MCF-7 cells and a decrease in SP cells. Wnt/β-catenin signaling pathway was also examined by analyzing the expression of total β-catenin and phosphorylated β-catenin in cytoplasm, and the results showed that the growth inhibitory effects of oxymatrine treatment on MCF-7 cells may be due to the inhibition of SP and Wnt/β-catenin signaling pathway. Further work is warranted to explore whether oxymatrine may be a useful novel therapeutic drug for targeting breast CSCs.     

Oxymatrine diminishes the side population and inhibits the expression of β-catenin in MCF-7 breast cancer cells

Cancer stem cells (CSCs) play a critical role in both cancer initiation and relapse as they are resistant to most cytotoxic agents and able to proliferate indefinitely. The plant alkaloid oxymatrine has many biological activities including the ability to induce cell cycle arrest and apoptosis, which makes it a potentially useful agent for targeting cancer cells. In order to determine whether it has beneficial pharmacological properties to eradicate CSCs, we analyzed the effects of oxymatrine on MCF-7 breast cancer cells. Cancer stem-like cells’ (side population, SP) identification and sorting were performed. The inhibitory effect of oxymatrine was evaluated on the sorted SP and non-SP cells. The results indicated that oxymatrine caused a dose-dependent reduction in the proliferation of MCF-7 cells and a decrease in SP cells. Wnt/β-catenin signaling pathway was also examined by analyzing the expression of total β-catenin and phosphorylated β-catenin in cytoplasm, and the results showed that the growth inhibitory effects of oxymatrine treatment on MCF-7 cells may be due to the inhibition of SP and Wnt/β-catenin signaling pathway. Further work is warranted to explore whether oxymatrine may be a useful novel therapeutic drug for targeting breast CSCs.

     

Alterations of E-cadherin and β-catenin in gastric cancer

Background  

The E-cadherin-catenin complex plays a crucial role in epithelial cell-cell adhesion and in the maintenance of tissue architecture. Perturbation in the expression or function of this complex results in loss of intercellular adhesion, with possible consequent cell transformation and tumour progression.     

Tumour-infiltrating CD68+ and CD57+ cells predict patient outcome in stage II–III colorectal cancer

Background:

The aim of our study was to evaluate the prognostic role of immunological microenvironnement in stage II–III CRC patients.

Methods:

We constructed a tissue microarray from 196 consecutive patients with stage II–III CRC and compared CD3, CD4, CD8, CD57, CD68, CXCL9/MIG, CXCL13, and PPARγ immunoreactivity in tumour samples and their matched non-tumour tissue. We assessed their association with relapse-free survival (RFS; primary endpoint) and overall survival (OS) in multivariate Cox models.

Results:

Low densities of CD57+ and CD68+ tumour-infiltrating cells (TIC) independently predicted worse outcomes. A prognostic score combining CD57 (+, > vs −, ⩽2 cells per spot) and CD68 (+, >0 vs −, =0 cells per spot) TIC density discriminated CRC patients at low (CD68+/CD57+), intermediate (CD68+/CD57−), or high (CD68−/CD57−) risk, with hazard ratios for the intermediate-risk and high-risk groups of 2.7 (95% confidence interval (CI): 1.3–5.8) and 9.0 (3.2–25.4) for RFS, and 2.5 (1.2–5.1) and 10.6 (3.8–29.2) for OS, respectively, as compared with the low-risk group. Corresponding 5-year survival rates (95% CI) in the low-, moderate- and high-risk groups were 84% (71–91), 65% (54–74), and 12% (2–47), respectively, for RFS, and 91% (80–96), 76% (66–84), and 25% (7–59), respectively, for OS.

Conclusion:

Tumour CD57+ and CD68+ TIC density assessment independently predicts survival in patients with stage II–III CRC. If validated, our score based on a quick, inexpensive, and well-established method such as point counting on diagnostic tissue sections could be used routinely as a prognostic tool in CRC patients.     

Regulation of β-catenin by t-DARPP in upper gastrointestinal cancer cells

Background  

Truncated dopamine and cyclic-AMP-regulated phosphoprotein (t-DARPP) is frequently overexpressed in gastrointestinal malignancies. In this study, we examined the role of t-DARPP in regulating β-catenin.     

The clinical significance evaluation of serum β2-microglobulin for thyroid cancer patients

Objective

The aim of this study was to investigate the clinical value and relevance on the serum β₂-microglobulin (β₂-MG) of patients with thyroid cancer.

Methods

One thousand and two normal cases, 95 thyroid cancer patients and 243 nodular goiter patients were selected to measure serum β₂-MG levels using double-antibody sandwich enzyme-linked immunosorbent assay (ELISA).

Results

The positive rate of 7.78% in normal population (78/1002) and 31.57% in thyroid cancer patients (30/95). There were significant differences between the normal population and thyroid cancer patients (χ ² = 55.352; P = 0.000). The positive rate of 7.81% in nodular goiter patients (19/243) and there were no significant differences between the normal population and nodular goiter patients (χ ² = 0.0004; P = 0.986), but significant differences between nodular goiter patients and thyroid cancer patients (χ ² = 31.106; P = 0.000). Meanwhile, the significant difference of the positive rate existed in between the various pathological types of thyroid cancer (χ ² = 10.015; P = 0.007), anaplastic thyroid cancer patients with the highest positive rate and The significant difference was found between the positive lymph node metastasis groups and negative lymph node metastasis groups (χ ² = 4.441; P = 0.035), the presence of distant metastasis group and absence of distant metastasis group (χ ² = 9.795; P = 0.002).

Conclusion

Serum β₂-MG levels and prognosis of thyroid cancer patients was negatively correlated. It showed important clinical value to detect the level of β₂-MG in the early diagnosis, prognosis and the clinical observation for thyroid cancer patients.