Association between activation of atypical NF‐κB1 p105 signaling pathway and nuclear β‐catenin accumulation in colorectal carcinoma

Recent studies have demonstrated that increased expression of coding region determinant‐binding protein (CRD‐BP) in response to β‐catenin signaling leads to the stabilization of β‐TrCP1, a substrate‐specific component of SCF E3 ubiquitin ligase complex, resulting in an accelerated degradation of IκBα and activation of canonical nuclear factor‐κB (NF‐κB) pathway. Here, we show that the noncanonical NF‐κB1 p105 pathway is constitutively activated in colorectal carcinoma specimens, being particularly associated with β‐catenin‐mediated increased expression of CRD‐BP and β‐TrCP1. In the carcinoma tissues exhibiting high levels of nuclear β‐catenin the phospho‐p105 levels were increased and total p105 amounts were decreased in comparison to that of normal tissue indicating an activation of this NF‐κB pathway. Knockdown of CRD‐BP in colorectal cancer cell line SW620 resulted in significantly higher basal levels of both NF‐κB inhibitory proteins, p105 and IκBα. Furthermore decreased NF‐κB binding activity was observed in CRD‐BP siRNA‐transfected SW620 cells as compared with those transfected with control siRNA. Altogether, our findings suggest that activation of NF‐κB1 p105 signaling in colorectal carcinoma might be attributed to β‐catenin‐mediated induction of CRD‐BP and β‐TrCP1. © 2009 Wiley‐Liss, Inc.     

Hsp27 regulates EGF/β‐catenin mediated epithelial to mesenchymal transition in prostate cancer

Increased expression of the molecular chaperone Hsp27 is associated with the progression of prostate cancer (PCa) to castration‐resistant disease, which is lethal due to metastatic spread of the prostate tumor. Metastasis requires epithelial to mesenchymal transition (EMT), which endows cancer cells with the ability to disseminate from the primary tumor and colonize new tissue sites. A wide variety of secreted factors promote EMT, and while overexpression and constitutive activation of epidermal growth factor (EGF) signaling is associated with poor prognosis of PCa, a precise role of EGF in PCa progression to metastasis remains unclear. Here, we show that Hsp27 is required for EGF‐induced cell migration, invasion and MMPs activity as well as the expression of EMT markers including Fibronectin, Vimentin and Slug with concomitant decrease of E‐cadherin. Mechanistically, we found that Hsp27 is required for EGF‐induced AKT and GSK3β phosphorylation and β‐catenin nuclear translocation. Moreover, silencing Hsp27 decreases EGF dependent phosphorylation of β‐catenin on tyrosine 142 and 654, enhances β‐catenin ubiquitination and degradation, prevents β‐catenin nuclear translocation and binding to the Slug promoter. These data suggest that Hsp27 is required for EGF‐mediated EMT via modulation of the β‐catenin/Slug signaling pathway. Together, our findings underscore the importance of Hsp27 in EGF induced EMT in PCa and highlight the use of Hsp27 knockdown as a useful strategy for patients with advanced disease.     

Tumor inhibition by sodium selenite is associated with activation of c‐Jun NH2‐terminal kinase 1 and suppression of β‐catenin signaling

Epidemiological and clinical studies suggest that an increased intake of dietary selenium significantly reduces overall cancer risk, but the anticancer mechanism of selenium is not clear. In this study, we fed intestinal cancer mouse model. Muc2/p21 double mutant mice with a selenium‐enriched (sodium selenite) diet for 12 or 24 weeks, and found that sodium selenite significantly inhibited intestinal tumor formation in these animals (p < 0.01), which was associated with phosphorylation of JNK1 and suppression of β‐catenin and COX2. In vitro studies showed that sodium selenite promoted cell apoptosis and inhibited cell proliferation in human colon cancer cell lines HCT116 and SW620. These effects were dose‐ and time course‐dependent, and were also linked to an increase of JNK1 phosphorylation and suppression of β‐catenin signaling. Reduced JNK1 expression by small RNA interference abrogated sufficient activation of JNK1 by sodium selenite, leading to reduced inhibition of the β‐catenin signaling, resulting in reduced efficacy of inhibiting cell proliferation. Taken together, our data demonstrate that sodium selenite inhibits intestinal carcinogenesis in vivo and in vitro through activating JNK1 and suppressing β‐catenin signaling, a novel anticancer mechanism of selenium.     

NUBPL,a novel metastasis‐related gene,promotes colorectal carcinoma cell motility by inducing epithelial–mesenchymal transition

Nucleotide binding protein‐like, NUBPL, is an assembly factor for human mitochondrial complex I, which is the biggest member of the mitochondrial respiratory chain. However, the relationship between NUBPL and carcinoma progression remains unknown. In this study, NUBPL was characterized for its role in colorectal cancer (CRC) and the underlying molecular mechanisms. Data (n = 197) from the Oncomine database revealed that mRNA levels of NUBPL were remarkably overexpressed in CRC tissues compared with normal tissues. In addition, immunohistochemical analysis of 75 pairs of CRC and non‐tumor tissues showed that the expression level of NUBPL was significantly higher in CRC tissues, and its expression level was positively associated with lymph node metastasis (P = 0.028) and advanced staging (P = 0.030). Expression of NUBPL in metastatic lymph nodes of CRC patients was also detected by immunohistochemical staining and high expression levels of NUBPL were observed. Overexpression of NUBPL significantly promoted the migration and invasion ability of CRC cell lines SW480 and SW620, whereas knockdown of NUBPL lead to an opposite effect. Our further study found that NUBPL could induce epithelial–mesenchymal transition (EMT), characterized by downregulation of epithelial markers (E‐cadherin) and upregulation of mesenchymal markers (N‐cadherin and vimentin). Moreover, NUBPL was able to activate ERK, which is believed to promote EMT and tumor metastasis. Inhibition of ERK suppressed the NUBPL‐induced changes in EMT and cell motility. These data showed that NUBPL plays a vital role in CRC migration and invasion by inducing EMT and activating ERK. It might be a novel therapeutic target for CRC.     

Retracted: EphB3‐targeted regulation of miR‐149 in the migration and invasion of human colonic carcinoma HCT116 and SW620 cells

microRNAs play key roles during various crucial cell processes such as proliferation, migration, invasion and apoptosis. Also, microRNAs have been shown to possess oncogenic and tumor‐suppressive functions in human cancers. Here, we describe the regulation and function of miR‐149 in colorectal cancer cell lines. miR‐149 expression patterns were detected in human colorectal cell lines and tissue samples, and then focused on its role in regulation of cell growth, migration, invasion, and its target gene identification. Furthermore, the function of the target gene of miR‐149 was analyzed in vitro and in vivo. miR‐149 expression was downregulated in human colorectal cancer HCT116 and SW620 cell lines compared to the normal colon epithelial NCM460 cell line using quantitative real‐time polymerase chain reaction methods. Further studies indicated that introduction of miR‐149 was able to suppress cell migration and invasion. Then, EphB3 was identified as a direct target gene of miR‐149 in colorectal cancer cells. Moreover, experiments in vitro showed that knockdown expression of EphB3 could suppress cell proliferation and invasion, and ectopic expression of EphB3 restored the phenotypes of CRC cell lines transfected with miR149. In addition, silencing of EphB3 significantly affected cycle progression distribution and increased apoptosis in CRC cell lines. Finally, in vivo results demonstrated that knockdown of EphB3 by siRNA inhibited tumor growth. In conclusion,the important role of miR‐149 in colorectal cancer progression suggesting that miR‐149 may serve as a therapeutic target for colorectal cancer treatment.     

Significance of the glycolytic pathway and glycolysis related-genes in tumorigenesis of human colorectal cancers

We investigated gene expressions involved in the glycolytic pathways in colorectal cancer. The study was designed to use gene ontology and its relevant bioinformatics tools to analyze the microarray data obtained from CRC tissues and their corresponding normal tissues, in order to explore the correlation between the glycolytic metabolic pathway and possible pathogenesis of this disease. The overexpression of glycolysis-related genes was observed in over 76% of CRC tissues. In addition, we stimulated the SW480 and SW620 CRC cell lines with 15 mM D-(+)-glucose and 10 mM 2-deoxy-D-glucose respectively. The results indicate that the proliferation response of both the SW480 and SW620 cell lines increased remarkably with a time-dependent effect by D-(+)-glucose administration. In contrast, the proliferation response of both the SW480 and SW620 cell lines was significantly inhibited by 2-DG administration. Likewise, further analyses of the expression of related genes triggered by the D-(+)-glucose in vivo show that the activation process of these eight genes - GLUT1, HK1, GPI, GAPD, PGK1, PGK2, ENO2, PKM2 - prominently increased with a time-dependent effect. In conclusion, this study demonstrates that the glycolytic pathway and glycolysis-related genes may play an important role in the tumorigenesis of CRC, but their molecular mechanisms need further investigation to verify this.     

Tetraspanin CD63 acts as a pro‐metastatic factor via β‐catenin stabilization

The tetraspanin CD63 is implicated in pro‐metastatic signaling pathways but, so far, it is unclear, how CD63 levels affect the tumor cell phenotype. Here, we investigated the effect of CD63 modulation in different metastatic tumor cell lines. In vitro, knock down of CD63 induced a more epithelial‐like phenotype concomitant with increased E‐cadherin expression, downregulation of its repressors Slug and Zeb1, and decreased N‐cadherin. In addition, β‐catenin protein was markedly reduced, negatively affecting expression of the target genes MMP‐2 and PAI‐1. β‐catenin inhibitors mimicked the epithelial phenotype induced by CD63 knock down. Inhibition of β‐catenin upstream regulators PI3K/AKT or GSK3β could rescue the mesenchymal phenotype underlining the importance of the β‐catenin pathway in CD63‐regulated cell plasticity. CD63 knock down‐induced phenotypical changes correlated with a decrease of experimental metastasis whereas CD63 overexpression enhanced the tumor cell‐intrinsic metastatic potential. Taken together, our data show that CD63 is a crucial player in the regulation of the tumor cell‐intrinsic metastatic potential by affecting cell plasticity.     

转凝蛋白在结直肠癌组织中的表达及其对SW480细胞恶性生物学行为的影响

目的:探讨转凝蛋白(transgelin,TAGLN)在结直肠癌(colorectal cancer,CRC)组织中的表达及其对SW480细胞增殖、迁移及侵袭的影响.方法:选取郑州大学附属肿瘤医院2015年5月至2016年8月收治的97例CRC患者的癌及配对的癌旁组织标本,以及人CRC细胞系SW620、SW480、HC...     

Dkk3, downregulated in cervical cancer,functions as a negative regulator of β‐catenin

The Wnt/β‐catenin signaling pathway is activated during the malignant transformation of keratinocytes that originate from the human uterine cervix. Dkk1, 2 and 4 have been shown to modulate the Wnt‐induced stabilization of the β‐catenin signaling pathway. However, the function of Dkk3 in this pathway is unknown. Comparison of the Dkk3 gene expression profiles in cervical cancer and normal cervical tissue by cDNA microarray and subsequent real‐time PCR revealed that the Dkk3 gene is frequently downregulated in the cancer. Methylation studies showed that the promoter of Dkk3 was methylated in cervical cancer cell lines and 22 (31.4%) of 70 cervical cancer tissue specimens. This promoter methylation was associated with reduced expression of Dkk3 mRNA in the paired normal and tumor tissue samples. Further, the reintroduction of Dkk3 into HeLa cervical cancer cells resulted in reduced colony formation and retarded cell growth. The forced expression of Dkk3 markedly attenuated β‐catenin‐responsive luciferase activity in a dose‐dependent manner and decreased the β‐catenin levels. By utilizing a yeast two‐hybrid screen, βTrCP, a negative regulator of β‐catenin was identified as a novel Dkk3‐interacting partner. Coexpression with βTrCP synergistically enhanced the inhibitory function of Dkk3 on β‐catenin. The stable expression of Dkk3 blocks the nuclear translocation of β‐catenin, resulting in downregulation of its downstream targets (VEGF and cylcin D), whereas knockdown of Dkk3 abrogates this blocking. We conclude from our finding that Dkk3 is a negative regulator of β‐catenin and its downregulation contribute to an activation of the β‐catenin signaling pathway. © 2008 Wiley‐Liss, Inc.     

lncRNA MALAT1在结直肠癌中的表达、生物学功能及其与患者预后的关系

目的:探讨lncRNA MALAT1在结直肠癌（colorectal cancer,CRC）中的表达、生物学功能及其与患者预后的关系。方法:采用生物信息方法在TCGA数据库和Oncomine数据库中分析比较MALAT1在CRC患者癌组织和癌旁组织中的差异表达情况,并比较高低表达组患者总生存期（OS）和无病生存期（DFS）是否存在差异。采用cBioPortal在线分析TCGA数据库中MALAT1突变种类及频率。采用实时荧光定量聚合酶链反应(real-time PCR)技术检测我院收治并手术治疗的30例CRC患者手术切除标本的癌组织、癌旁组织、肠癌细胞系(SW620、LOVO、HCT116)及正常人肠上皮细胞(NCM460)中MALAT1的相对表达水平。小干扰RNA敲低SW620细胞中MALAT1表达后观察MALAT1细胞迁移和侵袭能力改变情况。siRNA下调SW620细胞MALAT1,利用免疫印迹实验检测上皮间质化标志物E-cadherin、N-cadherin和Vimentin的表达水平。结果:Oncomine数据库中,CRC患者癌组织中高表达MALAT1的研究有6项;30对人配对CRC组织标本中,相比癌旁组织,MALAT1 在46.7%（14/30）的CRC患者组织中表达增高（P＜0.05）;Real-time PCR结果显示,肠癌细胞系(SW620、LOVO、HCT116)中MALAT1的相对表达量显著高于正常人肠上皮细胞(NCM460),差异有统计学意义（P＜0.05）。MALAT1基因突变类型主要为融合突变、扩增突变及深度缺失突变,其在各种肿瘤中的总体突变率为1.1%,在CRC组织中的突变率为0.34%。小干扰RNA抑制MALAT1表达可明显抑制SW620细胞的划痕迁移与小室侵袭能力。抑制MALAT1表达后,SW620细胞中E-cadherin的表达水平升高,同时N-cadherin和Vimentin的表达水平明显降低,表明上皮间质化过程受到明显抑制。MALAT1表达水平与结肠癌患者DFS存在明显的关系,高表达患者DFS显著低于低表达患者（HR=1.6,P=0.044）。结论:lncRNA MALAT1在肠癌中表达上调,并通过抑制上皮间质化抑制细胞迁移和侵袭能力。MALAT1高表达结肠癌患者预后不良,可能成为结肠癌预后分子标志物。     

Multiple effects of the special AT‐rich binding protein 1 (SATB1) in colon carcinoma

SATB1 (special AT‐rich binding protein 1) is a global chromatin organizer regulating the expression of a large number of genes. Overexpression has been found in various solid tumors and positively correlated with prognostic and clinicopathological properties. In colorectal cancer (CRC), SATB1 overexpression and its correlation with poor differentiation, invasive depth, TNM (tumor, nodes, metastases) stage and prognosis have been demonstrated. However, more detailed studies on the SATB1 functions in CRC are warranted. In this article, we comprehensively analyze the cellular and molecular role of SATB1 in CRC cell lines with different SATB1 expression levels by using RNAi‐mediated knockdown. Using siRNAs with different knockdown efficacies, we demonstrate antiproliferative, cell cycle‐inhibitory and proapoptotic effects of SATB1 knockdown in a SATB1 gene dose‐dependent manner. Tumor growth inhibition is confirmed in vivo in a subcutaneous tumor xenograft mouse model using stable knockdown cells. The in‐depth analysis of cellular effects reveals increased activities of caspases‐3, ‐7, ‐8, ‐9 and other mediators of apoptotic pathways. Similarly, the analysis of E‐ and N‐cadherin, slug, twist, β‐catenin and MMP7 indicates SATB1 effects on epithelial–mesenchymal transition (EMT) and matrix breakdown. Our results also establish SATB1 effects on receptor tyrosine kinases and (proto‐)oncogenes such as HER receptors and Pim‐1. Taken together, this suggests a more complex molecular interplay between tumor‐promoting and possible inhibitory effects in CRC by affecting multiple pathways and molecules involved in proliferation, cell cycle, EMT, invasion and cell survival.     

ATP‐P2Y2‐β‐catenin axis promotes cell invasion in breast cancer cells

Extracellular adenosine 5′‐triphosphate (ATP), secreted by living cancer cells or released by necrotic tumor cells, plays an important role in tumor invasion and metastasis. Our previous study demonstrated that ATP treatment in vitro could promote invasion in human prostate cancer cells via P2Y2, a preferred receptor for ATP, by enhancing EMT process. However, the pro‐invasion mechanisms of ATP and P2Y2 are still poorly studied in breast cancer. In this study, we found that P2Y2 was highly expressed in breast cancer cells and associated with human breast cancer metastasis. ATP could promote the in vitro invasion of breast cancer cells and enhance the expression of β‐catenin as well as its downstream target genes CD44, c‐Myc and cyclin D1, while P2Y2 knockdown attenuated above ATP‐driven events in vitro and in vivo. Furthermore, iCRT14, a β‐catenin/TCF complex inhibitor, could also suppress ATP‐driven migration and invasion in vitro. These results suggest that ATP promoted breast cancer cell invasion via P2Y2‐β‐catenin axis. Thus blockade of the ATP‐P2Y2‐β‐catenin axis could suppress the invasive and metastatic potential of breast cancer cells and may serve as potential targets for therapeutic interventions of breast cancer.     

The leukocyte protein L-plastin induces proliferation, invasion and loss of E-cadherin expression in colon cancer cells

L-plastin, a gene that codes for an actin-bundling protein, is upregulated in the metastatic colon cancer cell line SW620, when compared to its premetastatic counterpart SW480. The aim of our study was to characterise the effect of L-plastin overexpression on SW480 cells in the context of the acquisition of a metastatic phenotype. SW480 cell lines overexpressing L-plastin were established (SW480-LPL). Analysis of these cell lines revealed significantly higher rates of proliferation and invasion than the control cell line (SW480-Ctrl). In addition, the expression of E-cadherin was lost from SW480-LPL cells. Treatment of SW480-LPL cells with cytochalasin B, an inhibitor of endocytosis, attenuated the loss of E-cadherin expression in these cells. The association of L-plastin overexpression with an increased rate of proliferation and invasion, and loss of E-cadherin expression in the SW480 colon cancer cell line indicates that L-plastin plays an important mechanistic role in colorectal cancer metastasis (supplementary material for this article can be found on the International Journal of Cancer website at http://www.interscience.wiley.com/jpages/0020-7136/suppmat/index.html).