CLDN3

Gene Summary

Gene:CLDN3; claudin 3
Aliases: RVP1, HRVP1, C7orf1, CPE-R2, CPETR2
Location:7q11.23
Summary:Tight junctions represent one mode of cell-to-cell adhesion in epithelial or endothelial cell sheets, forming continuous seals around cells and serving as a physical barrier to prevent solutes and water from passing freely through the paracellular space. These junctions are comprised of sets of continuous networking strands in the outwardly facing cytoplasmic leaflet, with complementary grooves in the inwardly facing extracytoplasmic leaflet. The protein encoded by this intronless gene, a member of the claudin family, is an integral membrane protein and a component of tight junction strands. It is also a low-affinity receptor for Clostridium perfringens enterotoxin, and shares aa sequence similarity with a putative apoptosis-related protein found in rat. [provided by RefSeq, Jul 2008]
Databases:VEGA, OMIM, HGNC, Ensembl, GeneCard, Gene
Protein:claudin-3
Source:NCBIAccessed: 15 March, 2017

Ontology:

What does this gene/protein do?
Show (8)
Pathways:What pathways are this gene/protein implicaed in?
Show (3)

Cancer Overview

Research Indicators

Publications Per Year (1992-2017)
Graph generated 15 March 2017 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.

  • Claudin-1
  • Adenocarcinoma
  • DNA Methylation
  • Tissue Array Analysis
  • Vimentin
  • Neoplasm Invasiveness
  • RT-PCR
  • Down-Regulation
  • siRNA
  • Reproducibility of Results
  • Ovarian Cancer
  • Cystadenocarcinoma, Serous
  • Stomach Cancer
  • Polymerase Chain Reaction
  • Vulvar Cancer
  • Messenger RNA
  • Enterotoxins
  • Gene Expression Profiling
  • Breast Cancer
  • Claudins
  • Lung Cancer
  • Biomarkers, Tumor
  • Neoplastic Cell Transformation
  • Tight Junctions
  • Claudin-3
  • Survival Rate
  • Oligonucleotide Array Sequence Analysis
  • Cervical Cancer
  • RTPCR
  • Esophageal Cancer
  • Chromosome 7
  • Cancer Gene Expression Regulation
  • Squamous Cell Carcinoma
  • Immunoenzyme Techniques
  • Immunohistochemistry
  • Membrane Proteins
  • Cell Proliferation
  • Pathology, Molecular
  • Western Blotting
  • Clostridium perfringens
  • Claudin-4
Tag cloud generated 15 March, 2017 using data from PubMed, MeSH and CancerIndex

Specific Cancers (8)

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: CLDN3 (cancer-related)

Wahdan-Alaswad R, Harrell JC, Fan Z, et al.
Metformin attenuates transforming growth factor beta (TGF-β) mediated oncogenesis in mesenchymal stem-like/claudin-low triple negative breast cancer.
Cell Cycle. 2016; 15(8):1046-59 [PubMed] Free Access to Full Article Related Publications
Mesenchymal stem-like/claudin-low (MSL/CL) breast cancers are highly aggressive, express low cell-cell adhesion cluster containing claudins (CLDN3/CLDN4/CLDN7) with enrichment of epithelial-to-mesenchymal transition (EMT), immunomodulatory, and transforming growth factor-β (TGF-β) genes. We examined the biological, molecular and prognostic impact of TGF-β upregulation and/or inhibition using in vivo and in vitro methods. Using publically available breast cancer gene expression databases, we show that upregulation and enrichment of a TGF-β gene signature is most frequent in MSL/CL breast cancers and is associated with a worse outcome. Using several MSL/CL breast cancer cell lines, we show that TGF-β elicits significant increases in cellular proliferation, migration, invasion, and motility, whereas these effects can be abrogated by a specific inhibitor against TGF-β receptor I and the anti-diabetic agent metformin, alone or in combination. Prior reports from our lab show that TNBC is exquisitely sensitive to metformin treatment. Mechanistically, metformin blocks endogenous activation of Smad2 and Smad3 and dampens TGF-β-mediated activation of Smad2, Smad3, and ID1 both at the transcriptional and translational level. We report the use of ID1 and ID3 as clinical surrogate markers, where high expression of these TGF-β target genes was correlated to poor prognosis in claudin-low patients. Given TGF-β's role in tumorigenesis and immunomodulation, blockade of this pathway using direct kinase inhibitors or more broadly acting inhibitors may dampen or abolish pro-carcinogenic and metastatic signaling in patients with MCL/CL TNBC. Metformin therapy (with or without other agents) may be a heretofore unrecognized approach to reduce the oncogenic activities associated with TGF-β mediated oncogenesis.

Madaras L, Balint N, Gyorffy B, et al.
BRCA Mutation-Related and Claudin-Low Breast Cancer: Blood Relatives or Stepsisters.
Pathobiology. 2016; 83(1):1-12 [PubMed] Related Publications
BACKGROUND: BRCA mutation-associated (BRCAmut) breast cancer represents a heterogeneous group displaying certain molecular features. Claudin-low breast cancers (CLBC) overlap with characteristics of BRCAmut tumors; therefore, we have investigated whether these are identical subtypes.
METHODS: Using public gene expression data, CLDN, CDH1, 9-cell line claudin-low predictor (9CLCLP) and PAM50 expression was evaluated in BRCAmut and BRCA wild-type (BRCAwt) breast cancer cases focusing on their possible overlap with the CLBC subtype. A separate formalin-fixed, paraffin-embedded (FFPE) cohort of 22 BRCAmut and 19 BRCAwt tumor tissues was used for immunohistochemical examination of AR, CD24, CD44, CK5/6, claudin-1, -3, -4 and -7, E-cadherin, EGFR, estrogen receptor (ER), EZH2, HER2, Ki67, p53, progesterone receptor (PgR) and vimentin expression.
RESULTS: In the data sets, CLDN1 (ROC = 0.785, p < 0.001), CDH1 (ROC = 0.785, p < 0.001), CLDN7 (ROC = 0.723, p < 0.001), CLDN3 (ROC = 0.696, p = 0.020) and CLDN4 (ROC = 0.685, p = 0.027) were expressed at higher level in BRCAmut than BRCAwt tumor tissue. The PAM50 subtype differed from the assigned immunohistochemistry (IHC)-based subtype in 30%. Based on accessible 9CLCLP predictor genes, BRCAmut breast cancer does not display the claudin-low phenotype. Utilizing FFPE samples, claudins were evidently expressed in both BRCAmut and BRCAwt cases. However, at the protein level, only claudin-3 expression was higher in BRCAmut tumors, while claudin-1, -4 and -7 and E-cadherin expression was lower compared to BRCAwt cases. A CD24low/CD44high phenotype was found in BRCAmut tumors upon comparison with BRCAwt cases (p < 0.001 and p = 0.001, respectively).
CONCLUSIONS: There is a prominent correlation between the genes under focus herein and BRCA mutation status. BRCAmut tumors bear stem cell characteristics displaying a distinct cell adhesion molecule profile characterized by high expression of CDH1 and CLDN4 according to public gene expression data set analysis, and higher claudin-3 expression as detected by IHC; thus, BRCAmut breast carcinomas are not identical with the previously identified claudin-low subtype of breast cancer.

Nepomuceno AI, Shao H, Jing K, et al.
In-depth LC-MS/MS analysis of the chicken ovarian cancer proteome reveals conserved and novel differentially regulated proteins in humans.
Anal Bioanal Chem. 2015; 407(22):6851-63 [PubMed] Free Access to Full Article Related Publications
Ovarian cancer (OVC) remains the most lethal gynecological malignancy in the world due to the combined lack of early-stage diagnostics and effective therapeutic strategies. The development and application of advanced proteomics technology and new experimental models has created unique opportunities for translational studies. In this study, we investigated the ovarian cancer proteome of the chicken, an emerging experimental model of OVC that develops ovarian tumors spontaneously. Matched plasma, ovary, and oviduct tissue biospecimens derived from healthy, early-stage OVC, and late-stage OVC birds were quantitatively characterized by label-free proteomics. Over 2600 proteins were identified in this study, 348 of which were differentially expressed by more than twofold (p ≤ 0.05) in early- and late-stage ovarian tumor tissue specimens relative to healthy ovarian tissues. Several of the 348 proteins are known to be differentially regulated in human cancers including B2M, CLDN3, EPCAM, PIGR, S100A6, S100A9, S100A11, and TPD52. Of particular interest was ovostatin 2 (OVOS2), a novel 165-kDa protease inhibitor found to be strongly upregulated in chicken ovarian tumors (p = 0.0005) and matched plasma (p = 0.003). Indeed, RT-quantitative PCR and Western blot analysis demonstrated that OVOS2 mRNA and protein were also upregulated in multiple human OVC cell lines compared to normal ovarian epithelia (NOE) cells and immunohistochemical staining confirmed overexpression of OVOS2 in primary human ovarian cancers relative to non-cancerous tissues. Collectively, these data provide the first evidence for involvement of OVOS2 in the pathogenesis of both chicken and human ovarian cancer.

Yang Y, Cheon S, Jung MK, et al.
Interleukin-18 enhances breast cancer cell migration via down-regulation of claudin-12 and induction of the p38 MAPK pathway.
Biochem Biophys Res Commun. 2015; 459(3):379-86 [PubMed] Related Publications
Interleukin-18 (IL-18) was recently reported to have a pro-tumor effect in various cancers. Increased IL-18 levels in the serum of cancer patients correlated with malignancy, and IL-18 acts a crucial factor for cell migration in gastric cancer and melanoma. Claudins, which are the most important tight junction proteins, are also linked with cancer progression and metastasis. However, the relationship between claudins and IL-18 is not well-understood. Here, we show that the migratory ability of MCF-7 cells was reduced when endogenous IL-18 expression was inhibited with IL-18 siRNA. Moreover, exogenous IL-18 enhanced breast cancer cell migration and suppressed the expression of the tight junction proteins claudin-1, claudin-3, claudin-4, and claudin-12 in MCF-7 cells. Knockdown of claudin-3, claudin-4, and claudin-12, but not claudin-1, increased breast cancer migration with maximal effects observed in claudin-12 siRNA-transfected cells. To investigate whether the mitogen-activated protein kinase (MAPK) signaling pathway is involved in IL-18-induced cell migration and claudin-12 expression, cells were pretreated with SB203580 (an inhibitor of p38 MAPK) or PD98059 (an inhibitor of ERK1/2) prior to the addition of IL-18. Although pretreatment of MCF-7 cells with SB203580 blocked both the enhanced cell migration and the decreased claudin-12 expression, PD98059 only blocked cell migration and did not affect claudin-12 expression. In addition, exogenous IL-18 induced rapid phosphorylation of p38 MAPK. These results suggest that IL-18 is an important factor inducing breast cancer cell migration through down-regulation of claudin-12 and activation of the p38 MAPK pathway.

Vidal M, Peg V, Galván P, et al.
Gene expression-based classifications of fibroadenomas and phyllodes tumours of the breast.
Mol Oncol. 2015; 9(6):1081-90 [PubMed] Related Publications
Fibroepithelial tumors (FTs) of the breast are a heterogeneous group of lesions ranging from fibroadenomas (FAD) to phyllodes tumors (PT) (benign, borderline, malignant). Further understanding of their molecular features and classification might be of clinical value. In this study, we analysed the expression of 105 breast cancer-related genes, including the 50 genes of the PAM50 intrinsic subtype predictor and 12 genes of the Claudin-low subtype predictor, in a panel of 75 FTs (34 FADs, 5 juvenile FADs, 20 benign PTs, 5 borderline PTs and 11 malignant PTs) with clinical follow-up. In addition, we compared the expression profiles of FTs with those of 14 normal breast tissues and 49 primary invasive ductal carcinomas (IDCs). Our results revealed that the levels of expression of all breast cancer-related genes can discriminate the various groups of FTs, together with normal breast tissues and IDCs (False Discovery Rate < 5%). Among FTs, the levels expression of proliferation-related genes (e.g. CCNB1 and MKI67) and mesenchymal/epithelial-related (e.g. CLDN3 and EPCAM) genes were found to be most discriminative. As expected, FADs showed the highest and lowest expression of epithelial- and proliferation-related genes, respectively, whereas malignant PTs showed the opposite expression pattern. Interestingly, the overall profile of benign PTs was found more similar to FADs and normal breast tissues than the rest of tumours, including juvenile FADs. Within the dataset of IDCs and normal breast tissues, the vast majority of FADs, juvenile FADs, benign PTs and borderline PTs were identified as Normal-like by intrinsic breast cancer subtyping, whereas 7 (63.6%) and 3 (27.3%) malignant PTs were identified as Claudin-low and Basal-like, respectively. Finally, we observed that the previously described PAM50 risk of relapse prognostic score better predicted outcome in FTs than the morphological classification, even within PTs-only. Our results suggest that classification of FTs using gene expression-based data is feasible and might provide clinically useful biological and prognostic information.

Jiang L, Yang YD, Fu L, et al.
CLDN3 inhibits cancer aggressiveness via Wnt-EMT signaling and is a potential prognostic biomarker for hepatocellular carcinoma.
Oncotarget. 2014; 5(17):7663-76 [PubMed] Free Access to Full Article Related Publications
Hepatocellular carcinoma (HCC) is one of the most common fatal malignancies but the molecular genetic basis of this disease remains unclear. By using genome-wide methylation profiling analysis, we identified CLDN3 as an epigenetically regulated gene in cancer. Here, we investigated its function and clinical relevance in human HCC. CLDN3 downregulation occurred in 87/114 (76.3%) of primary HCCs, where it was correlated significantly with shorter survival of HCC patients (P=0.021). Moreover, multivariate cyclooxygenase regression analysis showed that CLDN3 was an independent prognostic factor for overall survival (P=0.014). Absent expression of CLDN3 was also detected in 67% of HCC cell lines, which was significantly associated with its promoter hypermethylation. Ectopic expression of CLDN3 in HCC cells could inhibit cell motility, cell invasiveness, and tumor formation in nude mice. Mechanistic investigations suggested through downregulation of GSK3B, CTNNB1, SNAI2, and CDH2, CLDN3 could significantly suppress metastasis by inactivating the Wnt/β-catenin-epithelial mesenchymal transition (EMT) axis in HCC cells. Collectively, our findings demonstrated that CLDN3 is an epigenetically silenced metastasis suppressor gene in HCC. A better understanding of the molecular mechanism of CLDN3 in inhibiting liver cancer cell metastasis may lead to a more effective management of HCC patients with the inactivation of CLDN3.

Molina-Pinelo S, Gutiérrez G, Pastor MD, et al.
MicroRNA-dependent regulation of transcription in non-small cell lung cancer.
PLoS One. 2014; 9(3):e90524 [PubMed] Free Access to Full Article Related Publications
Squamous cell lung cancer (SCC) and adenocarcinoma are the most common histological subtypes of non-small cell lung cancer (NSCLC), and have been traditionally managed in the clinic as a single entity. Increasing evidence, however, illustrates the biological diversity of these two histological subgroups of lung cancer, and supports the need to improve our understanding of the molecular basis beyond the different phenotypes if we aim to develop more specific and individualized targeted therapy. The purpose of this study was to identify microRNA (miRNA)-dependent transcriptional regulation differences between SCC and adenocarcinoma histological lung cancer subtypes. In this work, paired miRNA (667 miRNAs by TaqMan Low Density Arrays (TLDA)) and mRNA profiling (Whole Genome 44 K array G112A, Agilent) was performed in tumor samples of 44 NSCLC patients. Nine miRNAs and 56 mRNAs were found to be differentially expressed in SCC versus adenocarcinoma samples. Eleven of these 56 mRNA were predicted as targets of the miRNAs identified to be differently expressed in these two histological conditions. Of them, 6 miRNAs (miR-149, miR-205, miR-375, miR-378, miR-422a and miR-708) and 9 target genes (CEACAM6, CGN, CLDN3, ABCC3, MLPH, ACSL5, TMEM45B, MUC1) were validated by quantitative PCR in an independent cohort of 41 lung cancer patients. Furthermore, the inverse correlation between mRNAs and microRNAs expression was also validated. These results suggest miRNA-dependent transcriptional regulation differences play an important role in determining key hallmarks of NSCLC, and may provide new biomarkers for personalized treatment strategies.

Liu L, Gou M, Yi T, et al.
Antitumor effects of heparin-polyethyleneimine nanogels delivering claudin-3-targeted short hairpin RNA combined with low-dose cisplatin on ovarian cancer.
Oncol Rep. 2014; 31(4):1623-8 [PubMed] Related Publications
Cisplatin is normally administered in chemotherapy for ovarian cancer, but is accompanied by severe dose-dependent toxicity. The combination of cisplatin with other antitumor agents may be a useful alternative for achieving higher antitumor efficiency and lower toxicity. Claudin-3 (CLDN3), a commonly upregulated gene in 90% of ovarian cancers, has been identified as a novel therapeutic target of ovarian cancer. Therefore, in the present study, we constructed a recombinant plasmid carrying an shRNA targeting CLDN3 (pshCLDN3), and investigated the antitumor effects of the combination therapy of pshCLDN3 and a low-dose of cisplatin for the treatment of ovarian cancer. Heparin-polyethyleneimine (HPEI) nanogel, a novel gene carrier with superior biodegradability, excellent blood compatibility and low-toxicity, was used to deliver pshCLDN3 into ovarian cancer cells. The knockdown efficiency was determined by western blot analysis and CLDN3 immunostaining. Nude mice bearing intraperitoneal ovarian carcinomas were treated with pshCLDN3/HPEI complexes, low-dose cisplatin, pshCLDN3/HPEI plus low-dose cisplatin or control agents, respectively. The results showed that pshCLDN3/HPEI effectively suppressed the expression of CLDN3 in ovarian cancer. The combination therapy of pshCLDN3/HPEI and low-dose cisplatin exhibited enhanced antitumor activity, when compared with either agent alone, as evidenced by mean tumor weight analysis, Ki-67 immunostaining analysis and TUNEL assay, without obvious systemic toxicity. These results indicate that pshCLDN3/HPEI combined with low-dose cisplatin demonstrates apparent synergistic antitumor activity without marked toxicity. Our study offers a novel therapeutic strategy for the treatment of ovarian cancer.

Amaro A, Esposito AI, Gallina A, et al.
Validation of proposed prostate cancer biomarkers with gene expression data: a long road to travel.
Cancer Metastasis Rev. 2014; 33(2-3):657-71 [PubMed] Free Access to Full Article Related Publications
Biomarkers are important for early detection of cancer, prognosis, response prediction, and detection of residual or relapsing disease. Special attention has been given to diagnostic markers for prostate cancer since it is thought that early detection and surgery might reduce prostate cancer-specific mortality. The use of prostate-specific antigen, PSA (KLK3), has been debated on the base of cohort studies that show that its use in preventive screenings only marginally influences mortality from prostate cancer. Many groups have identified alternative or additional markers, among which PCA3, in order to detect early prostate cancer through screening, to distinguish potentially lethal from indolent prostate cancers, and to guide the treatment decision. The large number of markers proposed has led us to the present study in which we analyze these indicators for their diagnostic and prognostic potential using publicly available genomic data. We identified 380 markers from literature analysis on 20,000 articles on prostate cancer markers. The most interesting ones appeared to be claudin 3 (CLDN3) and alpha-methysacyl-CoA racemase highly expressed in prostate cancer and filamin C (FLNC) and keratin 5 with highest expression in normal prostate tissue. None of the markers proposed can compete with PSA for tissue specificity. The indicators proposed generally show a great variability of expression in normal and tumor tissue or are expressed at similar levels in other tissues. Those proposed as prognostic markers distinguish cases with marginally different risk of progression and appear to have a clinically limited use. We used data sets sampling 152 prostate tissues, data sets with 281 prostate cancers analyzed by microarray analysis and a study of integrated genomics on 218 cases to develop a multigene score. A multivariate model that combines several indicators increases the discrimination power but does not add impressively to the information obtained from Gleason scoring. This analysis of 10 years of marker research suggests that diagnostic and prognostic testing is more difficult in prostate cancer than in other neoplasms and that we must continue to search for better candidates.

Christgen M, Geffers R, Kreipe H, Lehmann U
IPH-926 lobular breast cancer cells are triple-negative but their microarray profile uncovers a luminal subtype.
Cancer Sci. 2013; 104(12):1726-30 [PubMed] Related Publications
Human primary breast cancers and breast cancer cell lines are classified by microarray-defined molecular subtypes, which reflect differentiation characteristics. Estrogen receptor (ER) expression is indicative of the luminal molecular subtype. We have previously established IPH-926, the first well-characterized cell line from infiltrating lobular breast cancer. IPH-926 displays an ER/PR/ErbB2 triple-negative immunophenotype, which is due to a loss of ER expression in its in vivo clonal ancestry. Loss of ER might indicate a fundamental change of cellular differentiation and it is unclear whether a luminal subtype is preserved beyond ER conversion. Using Affymetrix microarray analysis, seven different classifier gene lists (PAM305, DISC256, TN1288, PAM50, UNC1300, LAB704, INT500) and a background population of 50 common mammary carcinoma cell lines, we have now determined the molecular subtype of IPH-926. Strikingly, the IPH-926 expression profile is highly consistent with a luminal subtype. It is nearest to luminal/ER-positive breast cancer cell lines and far apart from basal breast cancer cell lines. Quantitative real-time RT-PCR confirmed enhanced expression of luminal marker genes (AGR2, CLU, CA12, EMP2, CLDN3) and low or absent expression of basal marker genes (KRT5, CD44, CAV1, VIM). Moreover, IPH-926 lacked androgen receptor (AR) expression, a transcription factor previously associated with luminal-like gene expression in a subset of triple-negative or molecular apocrine breast cancers. In conclusion, IPH-926 is triple-negative but belongs to the luminal subtype. Luminal differentiation characteristics can be preserved beyond ER conversion and might not require a compensatory expression of AR.

He ZY, Wei XW, Luo M, et al.
Folate-linked lipoplexes for short hairpin RNA targeting claudin-3 delivery in ovarian cancer xenografts.
J Control Release. 2013; 172(3):679-89 [PubMed] Related Publications
Ovarian cancers highly overexpress folate receptor α (FRα) and claudin3 (CLDN3), both of which are associated with tumor progression and poor prognosis of patients. Downregulation of FRα and CLDN3 in ovarian cancer may suppress tumor growth and promote benign differentiation of tumor. In this study, F-P-LP/CLDN3, a FRα targeted liposome loading with short hairpin RNA (shRNA) targeting CLDN3 was prepared and the pharmaceutical properties were characterized. Then, the antitumor effect of F-P-LP/CLDN3 was studied in an in vivo model of advanced ovarian cancer. Compared with Control, F-P-LP/CLDN3 promoted benign differentiation of tumor and achieved about 90% tumor growth inhibition. In the meantime, malignant ascites production was completely inhibited, and tumor nodule number and tumor weight were significantly reduced (p<0.001). FRα and CLDN3 were downregulated together in tumor tissues treated by F-P-LP/CLDN3. The antitumor mechanisms were achieved by promoting tumor cell apoptosis, inhibiting tumor cell proliferation and reducing microvessel density. Finally, safety evaluation indicated that F-P-LP/CLDN3 was a safe formulation in intraperitoneally administered cancer therapy. We come to a conclusion that F-P-LP/CLDN3 is a potential targeting formulation for ovarian cancer gene therapy.

Siar CH, Abbas SA
Claudin expression and tight junction protein localization in the lining epithelium of the keratocystic odontogenic tumors, dentigerous cysts, and radicular cysts.
Oral Surg Oral Med Oral Pathol Oral Radiol. 2013; 115(5):652-9 [PubMed] Related Publications
OBJECTIVE: The aim of this study was to evaluate the expression and localization of tight junction proteins (TJPs) or claudins in the keratocystic odontogenic tumor (KCOT) and to correlate with its biological behavior.
STUDY DESIGN: Five claudins (-1, -3, -4, -5, and 7) were examined immunohistochemically in 25 KCOTs and compared with 10 dentigerous cysts (DCs) and 10 radicular cysts (RCs).
RESULTS: Marked claudin-3 loss of expression in KCOT basal layer (n=24/25; 96%) compared with DCs (n=1/10; 10%) and RCs (n=5/10; 50%) (P<.05) suggests that claudin-3 downregulation may indicate altered or loss of basal cell polarity and impaired barrier function of KCOT lining epithelium and this might contribute indirectly to its biological behavior. In contrast, claudins-1, -4, -5, and -7 distribution patterns were less distinctive in all three entities, suggesting that these TJP molecules probably play limited roles in influencing their different growth potentials.
CONCLUSION: Present findings suggest that differential claudin expressions in the lining epithelium of KCOTs, DCs, and RCs probably reflect their neoplastic or nonneoplastic nature.

Nordfors K, Haapasalo J, Sallinen PK, et al.
Expression of claudins relates to tumour aggressivity, location and recurrence in ependymomas.
Histol Histopathol. 2013; 28(9):1137-46 [PubMed] Related Publications
The aim of our study was to assess the nature and importance of claudin expression in grade I-III ependymomas. The expression of claudins 2-5, 7, 10, TWIST, and ZEB1 were investigated in a series of 61 ependymomas using immunohistochemistry. All the claudins were expressed in ependymomas, except for CLDN4. CLDN5 positive tumours were associated with higher grade (p=0.049), whereas CLDN10 was lower in higher grade tumours (p=0.039). CLDN5 and CLDN3 were overexpressed in ependymomas of cerebral location (p=0.036, p=0.007, respectively). CLDN5 positive tumours showed more nuclear atypia, endothelial proliferation, mitosis, and hypercellularity (p=0.007, p=0.018, p=0.041, p=0.010, respectively). CLDN5 positivity correlated to higher proliferation (p=0.015). CLDN7 was more often positive in primary tumours (p=0.041). Positive ZEB1 expression was associated with CLDN2 negativity (p=0.031). TWIST-negative tumours were more often also CLDN5 and 10 negative (p=0.013, p=0.017, respectively). CLDN5 was related to more aggressive tumours compared to CLDN2 and 10, which tended to display a better degree of differentiation and a better prognosis. CLDN2 and CLDN5 were expressed commonly in ependymomas, while the parental ependymal cells in the central nervous system were usually negative. Evidently, claudins influence growth and differentiation in ependymomas.

Shang X, Lin X, Alvarez E, et al.
Tight junction proteins claudin-3 and claudin-4 control tumor growth and metastases.
Neoplasia. 2012; 14(10):974-85 [PubMed] Free Access to Full Article Related Publications
The extent of tight junction (TJ) formation is one of many factors that regulate motility, invasion, and metastasis. Claudins are required for the formation and maintenance of TJs. Claudin-3 (CLDN3) and claudin-4 (CLDN4) are highly expressed in the majority of ovarian cancers. We report here that CLDN3 and CLDN4 each serve to constrain the growth of human 2008 cancer xenografts and limit metastatic potential. Knockdown of CLDN3 increased in vivo growth rate by 2.3-fold and knockdown of CLDN4 by 3.7-fold in the absence of significant change in in vitro growth rate. Both types of tumors exhibited increase in birth rate as measured by Ki67 staining and decrease in death rate as reflected by terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining. Knockdown of either claudin did not alter expression of other TJ protein but did reduce TJ formation as measured by transepithelial resistance and paracellular flux of dextran, enhance migration and invasion in in vitro assays, and increase lung colonization following intravenous injection. Knockdown of CLDN3 and CLDN4 increased total lung metastatic burden by 1.7-fold and 2.4-fold, respectively. Loss of either CLDN3 or CLDN4 resulted in down-regulation of E-cadherin mRNA and protein, increased inhibitory phosphorylation of glycogen synthase kinase-3β (GSK-3β), and activation of β-catenin pathway signaling as evidenced by increases in nuclear β-catenin, the dephosphorylated form of the protein, and transcriptional activity of β-catenin/T-cell factor (TCF). We conclude that both CLDN3 and CLDN4 mediate interactions with other cells in vivo that restrain growth and metastatic potential by sustaining expression of E-cadherin and limiting β-catenin signaling.

Shang X, Lin X, Manorek G, Howell SB
Claudin-3 and claudin-4 regulate sensitivity to cisplatin by controlling expression of the copper and cisplatin influx transporter CTR1.
Mol Pharmacol. 2013; 83(1):85-94 [PubMed] Related Publications
Claudin-3 (CLDN3) and claudin-4 (CLDN4) are the major structural molecules that form tight junctions (TJs) between epithelial cells. We found that knockdown of the expression of either CLDN3 or CLDN4 produced marked changes in the phenotype of ovarian cancer cells, including an increase in resistance to cisplatin (cDDP). The effect of CLND3 and CLDN4 on cDDP cytotoxicity, cDDP cellular accumulation, and DNA adduct formation was compared in the CLDN3- and CLDN4-expressing parental human ovarian carcinoma 2008 cells and CLDN3 and CLDN4 knockdown sublines (CLDN3KD and CLDN4KD, respectively). Knockdown of CLDN3 or CLDN4 rendered human ovarian carcinoma 2008 cells resistant to cDDP in both in vitro culture and in vivo xenograft model. The net accumulation of platinum (Pt) and the Pt-DNA adduct levels were reduced in CLDN3KD and CLDN4KD cells. The endogenous mRNA levels of copper influx transporter CTR1 were found to be significantly reduced in the knockdown cells, and exogenous expression of CTR1 restored their sensitivity to cDDP. Reexpression of an shRNAi-resistant CLDN3 or CLDN4 up-regulated CTR1 levels, reversed the cDDP resistance, and enhanced TJ formation in the knockdown cells. Baseline copper (Cu) level, Cu uptake, and Cu cytotoxicity were also reduced in CLDN3KD and CLDN4KD cells. Cu-dependent tyrosinase activity was also markedly reduced in both types of CLDN knockdown cells when incubated with the substrate l-DOPA. These results indicate that CLDN3 and CLDN4 affect sensitivity of the ovarian cancer cells to the cytotoxic effect of cDDP by regulating expression of the Cu transporter CTR1.

Hung SW, Chiu CF, Chen TA, et al.
Recombinant viral protein VP1 suppresses HER-2 expression and migration/metastasis of breast cancer.
Breast Cancer Res Treat. 2012; 136(1):89-105 [PubMed] Related Publications
Breast cancer is one of the most common cancers in women worldwide and metastasis is the major cause of breast cancer death. Development of new therapeutic agents for inhibiting breast cancer metastasis is therefore an urgent need. We previously demonstrated that recombinant DNA-derived viral capsid protein VP1 (rVP1) of foot-and-mouth disease virus-induced apoptosis of MCF-7 breast cancer cells in vitro. Here, we investigated whether rVP1 exhibits any inhibitory effects on migration/metastasis and human epidermal growth factor receptor 2 (HER-2), a well-known biomarker for poor prognosis of breast cancer. The effects of rVP1 on cancer cell migration/invasion and metastasis were evaluated using Transwell migration assay and animal cancer models of metastasis. Western blotting, RT-PCR, flow cytometry, immunohistochemistry, and immunofluorescence staining techniques were used to investigate the effects of rVP1 on HER-2 and signal transduction mediators. Non-cytotoxic concentrations of rVP1-induced mesenchymal-epithelial transition and significantly suppressed AP-2α and HER-2 expression as well as the migration and invasion of a variety of breast cancer cell lines in a β1-integrin-dependent manner in vitro. Gross and histopathologic examinations showed that rVP1 also suppressed metastasis of several breast cancer cell lines, including HER-2-overexpressing SK-BR-3 and BT-474 cells to lung, liver, or peripheral lymph node in orthotopic allograft/xenograft murine models. In addition, rVP1 significantly prolonged survival in breast cancer-bearing mice. Notably, no apparent side effects of rVP1 were detected, as shown by normal complete blood count levels and serum biochemistry profiles, including AST, ALT, BUN, and creatine. This study demonstrates that rVP1 suppresses the migration, invasion, and metastasis of breast cancer cells via binding to β1 integrin receptor and down-regulation of AP-2α and HER-2 expression. The effectiveness of rVP1 on inhibiting migration/metastasis of breast cancer and HER-2 expression suggests that it may be suitable for serving as potential therapeutics for metastatic breast cancer particularly HER-2-overexpressing cancer.

Ricardo S, Gerhard R, Cameselle-Teijeiro JF, et al.
Claudin expression in breast cancer: high or low, what to expect?
Histol Histopathol. 2012; 27(10):1283-95 [PubMed] Related Publications
The evaluation of claudins (CLDNs) expression pattern in tumours can be important to understand breast carcinogenesis. The study of CLDNs became more appealing since it was found that CLDN3 and CLDN4 are putative therapeutic targets for Clostridium perfrigens enterotoxin (CPE), as well as for monoclonal antibody-based therapy. Moreover, the recently characterized CLDN-low molecular subgroup of breast tumours increased the interest in these molecules. Based on these facts, our aim was to explore the pattern of expression of CLDNs among a large series of invasive breast carcinomas. We also analysed the correlation between the combinatorial expression of CLDN3/CLDN4 and classical prognostic factors and biological markers. In addition, we also compared the characteristics of tumours with low expression of CLDN3, CLDN4 and CLDN7, assessed by immunohistochemistry (IHC), and the ones from CLDN-low subgroup of tumours previously defined by genomic assays. The combinatorial analysis of the expression of CLDN3/CLDN4 showed a significant association between high CLDN3/CLDN4 levels and triple-negative tumours, as well as with worse patient outcome. This combined analysis may provide useful information for breast carcinomas, since these two CLDN members are putative therapeutic targets. Comparing tumours with low expression of CLDN3, CLDN4 and CLDN7 with tumours previously referred to as CLDN-low by genomic assays, we demonstrated that the single IHC evaluation of these three specific CLDNs is insufficient to identify the CLDN-low molecular subtype of breast tumours. The analysis of several other molecular markers, such as EMT (epithelial-to-mesenchymal transition) and CSC (cancer stem cell) markers should probably be added to improve the identification of this subgroup of tumours by IHC, which probably are enriched in carcinomas with metaplastic differentiation.

Park HS, Kim GY, Choi IW, et al.
Inhibition of matrix metalloproteinase activities and tightening of tight junctions by diallyl disulfide in AGS human gastric carcinoma cells.
J Food Sci. 2011; 76(4):T105-11 [PubMed] Related Publications
The effect of diallyl disulfide (DADS), a major component of an oil-soluble allyl sulfide garlic (Allium sativum) derivative, on the correlation between anti-invasive activity and tightening of tight junctions (TJs) was investigated in human gastric adenocarcinoma AGS cells. Our data indicated that the inhibitory effects of DADS on cell motility and invasiveness were found to be associated with increased tightness of the TJs, which was demonstrated by an increase in transepithelial electrical resistance. Activities of matrix metalloprotease (MMP)-2 and -9 in AGS cells were dose-dependently inhibited by treatment with DADS, and this was also correlated with a decrease in expression of their mRNA and proteins; however, tissue inhibitor of metalloproteinase (TIMP)-1 and -2 mRNA levels and proteins were increased. Additionally, immunoblotting results indicated that DADS repressed the levels of claudin proteins (claudin-2, -3, and -4), major components of TJs that play key roles in control and selectivity of paracellular transport. Although further studies are needed, these results suggest that DADS treatment may inhibit tumor cell motility and invasion and, therefore, act as a dietary source to decrease the risk of cancer metastasis.

Okugawa T, Oshima T, Chen X, et al.
Down-regulation of claudin-3 is associated with proliferative potential in early gastric cancers.
Dig Dis Sci. 2012; 57(6):1562-7 [PubMed] Related Publications
BACKGROUND: Claudins are tight junction (TJ) proteins, and the relationship between the level of expression and localization of TJ protein, and tumor aggressiveness in early gastric cancer (GC) is still far from clear.
AIMS: To investigate the expression of claudins and Ki-67 in early GC cells and surrounding normal gastric mucosa.
METHODS: A total of 53 early GC lesions removed via endoscopic mucosal resection or endoscopic submucosal resection were evaluated. All of the GCs were characterized as well to moderately differentiated adenocarcinoma. The labeling index (LI) of Ki-67 was calculated for each sample. To assess the prevalence of epithelial TJs, immunofluorescent staining for claudin-3, claudin-4, and claudin-7 was performed. The immunoreactivity was graded according to the percentage of stained cells.
RESULTS: Claudin-3, claudin-4, and claudin-7 expression at TJs in GC and intestinal metaplasia were significantly higher than that in gastric mucosa with no intestinal metaplasia. The Ki-67 LI of GC specimens was inversely correlated with claudin-3 expression, but not with claudin-4 or claudin-7 expression. Claudin-3 expression was significantly lower at the submucosal invasive front of GCs.
CONCLUSIONS: The down-regulation of claudin-3 was associated with the proliferative potential of GC cells, indicating that claudins may have a pivotal role in the progression of GC.

Riski M, Santala M, Soini Y, Talvensaari-Mattila A
Claudins 1, 3M, 3S, 4, 5 and 7 in vulvar neoplasms compared with vulvar squamous cell carcinoma.
Tumour Biol. 2012; 33(2):537-42 [PubMed] Related Publications
The purpose of this study was to evaluate the expression of claudins 1, 3M (membrane-bound), 3S (cytoplasmic), 4, 5 and 7 in vulvar epithelial neoplasia (VIN I-III) and to compare those with invasive vulvar squamous cell carcinoma. Paraffin tissue sections from 73 vulvar neoplasms (12 VIN I, 12 VIN II-III and 49 vulvar carcinomas) were studied by immunohistochemistry for the expression of claudins 1, 3M, 3S, 4, 5 and 7. Claudin 1 stained strongly in all groups, whereas claudin 3M, 3S and 4 immunostaining were moderate in all groups. Claudin 7 stained strongly in all groups. Claudin 3M expression was higher in VIN I compared to carcinoma, while no difference was found between VIN I and VIN II-III or between VIN II-III and carcinoma. Claudin 1 and claudin 3S expressions also showed the same decreasing tendency from VIN towards vulvar carcinoma. Claudin 5 showed only weak staining in VIN I and VIN II-III, and positive expression was also low in the carcinoma group. Expressions of claudins 1, 3M, 3S, 4 and 7 were found in VIN and vulvar carcinoma. Changes in claudin 1 and claudin 3 expression during progression from VIN to vulvar carcinoma suggests a connection with claudin expression and differentiation of vulvar squamous cells. Claudin 5 does not seem to be important in VIN or vulvar carcinoma.

Walther W, Petkov S, Kuvardina ON, et al.
Novel Clostridium perfringens enterotoxin suicide gene therapy for selective treatment of claudin-3- and -4-overexpressing tumors.
Gene Ther. 2012; 19(5):494-503 [PubMed] Related Publications
Bacterial toxins are known to be effective for cancer therapy. Clostridium perfringens enterotoxin (CPE) is produced by the bacterial Clostridium type A strain. The transmembrane proteins claudin-3 and -4, often overexpressed in numerous human epithelial tumors (for example, colon, breast, pancreas, prostate and ovarian), are the targeted receptors for CPE. CPE binding to them triggers formation of membrane pore complexes leading to rapid cell death. In this study, we aimed at selective tumor cell killing by CPE gene transfer. We generated expression vectors bearing the bacterial wild-type CPE cDNA (wtCPE) or translation-optimized CPE (optCPE) cDNA for in vitro and in vivo gene therapy of claudin-3- and -4-overexpressing tumors. The CPE expression analysis at messenger RNA and protein level revealed more efficient expression of optCPE compared with wtCPE. Expression of optCPE showed rapid cytotoxic activity, hightened by CPE release as bystander effect. Cytotoxicity of up to 100% was observed 72 h after gene transfer and is restricted to claudin-3-and -4-expressing tumor lines. MCF-7 and HCT116 cells with high claudin-4 expression showed dramatic sensitivity toward CPE toxicity. The claudin-negative melanoma line SKMel-5, however, was insensitive toward CPE gene transfer. The non-viral intratumoral in vivo gene transfer of optCPE led to reduced tumor growth in MCF-7 and HCT116 tumor-bearing mice compared with the vector-transfected control groups. This novel approach demonstrates that CPE gene transfer can be employed for a targeted suicide gene therapy of claudin-3- and -4-overexpressing tumors, leading to the rapid and efficient tumor cell killing in vitro and in vivo.

Casagrande F, Cocco E, Bellone S, et al.
Eradication of chemotherapy-resistant CD44+ human ovarian cancer stem cells in mice by intraperitoneal administration of Clostridium perfringens enterotoxin.
Cancer. 2011; 117(24):5519-28 [PubMed] Free Access to Full Article Related Publications
BACKGROUND: Emerging evidence has suggested that the capability to sustain tumor formation, growth, and chemotherapy resistance in ovarian as well as other human malignancies exclusively resides in a small proportion of tumor cells termed cancer stem cells. During the characterization of CD44(+) ovarian cancer stem cells, we found a high expression of the genes encoding for claudin-4. Because this tight junction protein is the natural high-affinity receptor for Clostridium perfringens enterotoxin (CPE), we have extensively investigated the sensitivity of ovarian cancer stem cells to CPE treatment in vitro and in vivo.
METHODS: Real-time polymerase chain reaction and flow cytometry were used to evaluate claudin-3/-4 expression in ovarian cancer stem cells. Small interfering RNA knockdown experiments and MTS assays were used to evaluate CPE-induced cytotoxicity against ovarian cancer stem cell lines in vitro. C.B-17/SCID mice harboring ovarian cancer stem cell xenografts were used to evaluate CPE therapeutic activity in vivo.
RESULTS: CD44(+) ovarian cancer stem cells expressed claudin-4 gene at significantly higher levels than matched autologous CD44(-) ovarian cancer cells, and regardless of their higher resistance to chemotherapeutic agents died within 1 hour after exposure to 1.0 μg/mL of CPE in vitro. Conversely, small-interfering RNA-mediated knockdown of claudin-3/-4 expression in CD44(+) cancer stem cells significantly protected cancer stem cells from CPE-induced cytotoxicity. Importantly, multiple intraperitoneal administrations of sublethal doses of CPE in mice harboring xenografts of chemotherapy-resistant CD44(+) ovarian cancer stem cells had a significant inhibitory effect on tumor progression leading to the cure and/or long-term survival of all treated animals (ie, 100% reduction in tumor burden in 50% of treated mice; P < .0001).
CONCLUSIONS: CPE may represent an unconventional, potentially highly effective strategy to eradicate chemotherapy-resistant cancer stem cells.

Hess J, Thomas G, Braselmann H, et al.
Gain of chromosome band 7q11 in papillary thyroid carcinomas of young patients is associated with exposure to low-dose irradiation.
Proc Natl Acad Sci U S A. 2011; 108(23):9595-600 [PubMed] Free Access to Full Article Related Publications
The main consequence of the Chernobyl accident has been an increase in papillary thyroid carcinomas (PTCs) in those exposed to radioactive fallout as young children. Our aim was to identify genomic alterations that are associated with exposure to radiation. We used array comparative genomic hybridization to analyze a main (n = 52) and a validation cohort (n = 28) of PTC from patients aged <25 y at operation and matched for age at diagnosis and residency. Both cohorts consisted of patients exposed and not exposed to radioiodine fallout. The study showed association of a gain on chromosome 7 (7q11.22-11.23) with exposure (false discovery rate = 0.035). Thirty-nine percent of the exposed group showed the alteration; however, it was not found in a single case from the unexposed group. This was confirmed in the validation set. Because only a subgroup of cases in the exposed groups showed gain of 7q11.22-11.23, it is likely that different molecular subgroups and routes of radiation-induced carcinogenesis exist. The candidate gene CLIP2 was specifically overexpressed in the exposed cases. In addition, the expression of the genes PMS2L11, PMS2L3, and STAG3L3 correlated with gain of 7q11.22-11.23. An enrichment of Gene Ontology terms "DNA repair" (PMS2L3, PMS2L5), "response to DNA damage stimulus" (BAZ1B, PMS2L3, PMS2L5, RFC2), and "cell-cell adhesion" (CLDN3, CLDN4) was found. This study, using matched exposed and unexposed cohorts, provides insights into the radiation-related carcinogenesis of young-onset PTC and, with the exposure-specific gain of 7q11 and overexpression of the CLIP2 gene, radiation-specific molecular markers.

Sun C, Yi T, Song X, et al.
Efficient inhibition of ovarian cancer by short hairpin RNA targeting claudin-3.
Oncol Rep. 2011; 26(1):193-200 [PubMed] Related Publications
Ovarian cancer is one of the most lethal gynecologic neoplasms. Even though various new chemotherapeutics have been developed for the treatment of ovarian cancer, drug resistance and undesired serious side effects remain unavoidable obstacles for chemotherapeutic approaches. New strategies to overcome the therapeutic dilemma are needed. Claudin-3 (CLDN3) is a recently discovered gene generally overexpressed in human ovarian cancers but not in normal ovarian tissue. Its high expression has been identified to associate with the invasion, proliferation and survival of cancer cells, making it a promising target for gene therapy of ovarian cancer. However, in gene therapy, traditional gene carriers such as virus or cationic liposomes suffer from distressing shortcomings of potential carcinogenicity, obvious cytotoxicity and immunogenicity. Nanoparticles (NPs) based on PLGA are a novel gene delivery system with good biodegradability, excellent biocompatibility and low toxcity for in vivo gene delivery compared with traditional gene carriers. We constructed a plasmid expressing shRNA targeted CLDN3 (pshCLDN3) encapsulated with PLGA-NPs, and administered it by i.p. injection to nude mice bearing intraperitoneal SKOV3 ovarian cancer, to investigate the antitumor potential of knocking down CLDN3. After 12 times of administration, the tumors of each group were compared. The underlying antitumor mechanisms were revealed by immunostaining of CD31, Ki-67 and TUNEL assay, to exhibit possible alterations in microvessel density, cell proliferation and cell apoptosis. Our study demonstrated that i.p. administration of pshCLDN3 effectively suppressed the expression of CLDN3 and, thus, inhibited the growth of ovarian tumors, significantly reducing tumor weight by 67.4% compared with blank controls (p<0.05). Immunostaining of CD31, Ki-67 and TUNEL assay demonstrated decreased angiogenesis (p<0.05), reduced proliferation (p<0.05) and increased apoptosis (p<0.05) in the pshCLDN3 treated group compared with controls. No obvious toxicity of PLGA-NPs was observed either in vitro or in vivo. Our results indicated that knockdown of CLDN3 by pshCLDN3 encapsulated in PLGA NPs may provide a promising approach for the treatment of ovarian cancer.

Tang W, Dou T, Zhong M, Wu Z
Dysregulation of Claudin family genes in colorectal cancer in a Chinese population.
Biofactors. 2011 Jan-Feb; 37(1):65-73 [PubMed] Related Publications
Claudins play an important role in tumor metastasis and in invasiveness of colorectal cancer (CRC). We have evaluated the relationship between CRC and expression of the claudin genes in Chinese patients with CRC. We measured CLDN1 and CLDN7 mRNA using quantitative PCR, and protein levels with immunohistochemistry in cancer tissues and adjacent normal tissue. Cancer tissues had significantly higher levels of CLDN1, and significantly lower levels of CLDN3, CLDN4, and CLDN7 than did normal tissue. CLDN3, CLDN4, and CLDN7 expression levels were higher in CRC of the protruded type than in CRC of the infiltrative type. Expression of CLDN7 correlated with lymph node metastasis. Stage N0 cancer tissues had higher levels of CLDN7 than did stages N1 and N2, suggesting that CLDN7 expression was closely related to the extent of lymph node metastasis. CLDN1 protein was upregulated, but CLDN7 protein was downregulated in cancer tissues when compared with expression in adjacent normal tissues. In conclusion, CLDN3, CLDN4, and CLDN7 were significantly downregulated, whereas CLDN1 was significantly upregulated in CRC. The altered expression of claudin genes may play a role in the initiation and development of CRC.

Han CL, Chen JS, Chan EC, et al.
An informatics-assisted label-free approach for personalized tissue membrane proteomics: case study on colorectal cancer.
Mol Cell Proteomics. 2011; 10(4):M110.003087 [PubMed] Free Access to Full Article Related Publications
We developed a multiplexed label-free quantification strategy, which integrates an efficient gel-assisted digestion protocol, high-performance liquid chromatography tandem MS analysis, and a bioinformatics alignment method to determine personalized proteomic profiles for membrane proteins in human tissues. This strategy provided accurate (6% error) and reproducible (34% relative S.D.) quantification of three independently purified membrane fractions from the same human colorectal cancer (CRC) tissue. Using CRC as a model, we constructed the personalized membrane protein atlas of paired tumor and adjacent normal tissues from 28 patients with different stages of CRC. Without fractionation, this strategy confidently quantified 856 proteins (≥2 unique peptides) across different patients, including the first and robust detection (Mascot score: 22,074) of the well-documented CRC marker, carcinoembryonic antigen 5 by a discovery-type proteomics approach. Further validation of a panel of proteins, annexin A4, neutrophils defensin A1, and claudin 3, confirmed differential expression levels and high occurrences (48-70%) in 60 CRC patients. The most significant discovery is the overexpression of stomatin-like 2 (STOML2) for early diagnostic and prognostic potential. Increased expression of STOML2 was associated with decreased CRC-related survival; the mean survival period was 34.77 ± 2.03 months in patients with high STOML2 expression, whereas 53.67 ± 3.46 months was obtained for patients with low STOML2 expression. Further analysis by ELISA verified that plasma concentrations of STOML2 in early-stage CRC patients were elevated as compared with those of healthy individuals (p < 0.001), suggesting that STOML2 may be a noninvasive serological biomarker for early CRC diagnosis. The overall sensitivity of STOML2 for CRC detection was 71%, which increased to 87% when combined with CEA measurements. This study demonstrated a sensitive, label-free strategy for differential analysis of tissue membrane proteome, which may provide a roadmap for the subsequent identification of molecular target candidates of multiple cancer types.

Cocco E, Casagrande F, Bellone S, et al.
Clostridium perfringens enterotoxin carboxy-terminal fragment is a novel tumor-homing peptide for human ovarian cancer.
BMC Cancer. 2010; 10:349 [PubMed] Free Access to Full Article Related Publications
BACKGROUND: Development of innovative, effective therapies against recurrent/chemotherapy-resistant ovarian cancer remains a high priority. Using high-throughput technologies to analyze genetic fingerprints of ovarian cancer, we have discovered extremely high expression of the genes encoding the proteins claudin-3 and claudin-4.
METHODS: Because claudin-3 and -4 are the epithelial receptors for Clostridium perfringens enterotoxin (CPE), and are sufficient to mediate CPE binding, in this study we evaluated the in vitro and in vivo bioactivity of the carboxy-terminal fragment of CPE (i.e., CPE290-319 binding peptide) as a carrier for tumor imaging agents and intracellular delivery of therapeutic drugs. Claudin-3 and -4 expression was examined with rt-PCR and flow cytometry in multiple primary ovarian carcinoma cell lines. Cell binding assays were used to assess the accuracy and specificity of the CPE peptide in vitro against primary chemotherapy-resistant ovarian carcinoma cell lines. Confocal microscopy and biodistribution assays were performed to evaluate the localization and uptake of the FITC-conjugated CPE peptide in established tumor tissue.
RESULTS: Using a FITC-conjugated CPE peptide we show specific in vitro and in vivo binding to multiple primary chemotherapy resistant ovarian cancer cell lines. Bio-distribution studies in SCID mice harboring clinically relevant animal models of chemotherapy resistant ovarian carcinoma showed higher uptake of the peptide in tumor cells than in normal organs. Imunofluorescence was detectable within discrete accumulations (i.e., tumor spheroids) or even single chemotherapy resistant ovarian cancer cells floating in the ascites of xenografted animals while a time-dependent internalization of the FITC-conjugated CPE peptide was consistently noted in chemotherapy-resistant ovarian tumor cells by confocal microscopy.
CONCLUSIONS: Based on the high levels of claudin-3 and -4 expression in chemotherapy-resistant ovarian cancer and other highly aggressive human epithelial tumors including breast, prostate and pancreatic cancers, CPE peptide holds promise as a lead peptide for the development of new diagnostic tracers or alternative anticancer agents.

Jung H, Jun KH, Jung JH, et al.
The expression of claudin-1, claudin-2, claudin-3, and claudin-4 in gastric cancer tissue.
J Surg Res. 2011; 167(2):e185-91 [PubMed] Related Publications
BACKGROUND: The claudins (CLDNs) are a family of functional tight junction proteins, and are involved with the epithelial-to-mesenchymal transition (EMT). The claudin proteins have a significant influence on the biological behavior of tumor progression in several types of cancers. In this study, we aimed to evaluate the expression pattern of claudin-1, claudin-2, claudin-3, and claudin-4 in gastric cancer tissue.
MATERIALS AND METHODS: Tissue was obtained from surgically resected specimens of 72 patients who were diagnosed with gastric adenocarcinoma at a single institution. The expressions of claudin-1, claudin-2, claudin-3, and claudin-4 were determined by immunohistochemical staining with the ABC method.
RESULTS: Claudin-2 demonstrated the highest expression rate (73.6%) and claudin-4 demonstrated the lowest expression rate (44.4%). The expression of claudin-1 was significantly lower in cases of intestinal type adenocarcinoma based on the Lauren classification. The expressions of claudin-3 and claudin-4 were significantly lower in cases with positive lymphatic invasion. The expression of claudin-3 was significantly lower in cases with an advanced T-stage (T3 and T4). The expression of claudin-3 showed significantly positive correlations with the expression of the other claudin proteins. In survival analysis, the expression of claudin-4 was related to good overall survival rate with significance (P = 0.046).
CONCLUSION: We suggest that claudin-3 and claudin-4 represent useful molecular markers for gastric cancer. Claudin-3 and claudin-4 would be the most important proteins related to the lymphatic invasion process, and claudin-4 would be useful with prognostic marker based on our results. Further investigations with a greater number of subjects are required to identify the action mechanism of claudin in gastric cancer.

Kwon MJ, Kim SS, Choi YL, et al.
Derepression of CLDN3 and CLDN4 during ovarian tumorigenesis is associated with loss of repressive histone modifications.
Carcinogenesis. 2010; 31(6):974-83 [PubMed] Free Access to Full Article Related Publications
Unlike epigenetic silencing of tumor suppressor genes, the role of epigenetic derepression of cancer-promoting genes or oncogenes in carcinogenesis remains less well understood. The tight junction proteins claudin-3 and claudin-4 are frequently overexpressed in ovarian cancer and their overexpression was previously reported to promote the migration and invasion of ovarian epithelial cells. Here, we show that the expression of claudin-3 and claudin-4 is repressed in ovarian epithelial cells in association with promoter 'bivalent' histone modifications, containing both the activating trimethylated histone H3 lysine 4 (H3K4me3) mark and the repressive mark of trimethylated histone H3 lysine 27 (H3K27me3). During ovarian tumorigenesis, derepression of CLDN3 and CLDN4 expression correlates with loss of H3K27me3 in addition to trimethylated histone H4 lysine 20 (H4K20me3), another repressive histone modification. Although CLDN4 repression was accompanied by both DNA hypermethylation and repressive histone modifications, DNA methylation was not required for CLDN3 repression in immortalized ovarian epithelial cells. Moreover, activation of both CLDN3 and CLDN4 in ovarian cancer cells was associated with simultaneous changes in multiple histone modifications, whereas H3K27me3 loss alone was insufficient for their derepression. CLDN4 repression was robustly reversed by combined treatment targeting both DNA demethylation and histone acetylation. Our study strongly suggests that in addition to the well-known chromatin-associated silencing of tumor suppressor genes, epigenetic derepression by the conversely related loss of repressive chromatin modifications also contributes to ovarian tumorigenesis via activation of cancer-promoting genes or candidate oncogenes.

Kim SO, Choi BT, Choi IW, et al.
Anti-invasive activity of histone deacetylase inhibitors via the induction of Egr-1 and the modulation of tight junction-related proteins in human hepatocarcinoma cells.
BMB Rep. 2009; 42(10):655-60 [PubMed] Related Publications
The potential anti-metastasis and anti-invasion activities of early growth response gene-1 (Egr-1) and claudin-3, a tight junction (TJ)-related protein, were evaluated using histone deacetylase (HDAC) inhibitors in human hepatocarcinoma cells. The results of wound healing and Transwell assays showed that HDAC inhibitors such as trichostatin A and sodium butyrate inhibited cell migration and invasion. HDAC inhibitors markedly induced Egr-1 expression during the early period, after which expression levels decreased. In addition, the down-regulation of snail and type 1 insulin-like growth factor receptor (IGF-1R) in HDAC inhibitor-treated cells induced the upregulation of thrombospondin-1 (TSP-1), E-cadherin and claudin-3. Cells transfected with Egr-1 and claudin-3 siRNA displayed significant blockage of HDAC inhibitor-induced anti-invasive activity. Collectively, these findings indicate that the up-regulation of Egr-1 and claudin-3 are crucial steps in HDAC inhibitor-induced anti-metastasis and anti-invasion.

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