Desmocollin 3 Research Articles

A Novel In Vivo Active Pemphigus Model Targeting Desmoglein1 and Desmoglein3: A Tool Representing All Pemphigus Variants.

Background: Pemphigus is a life-threatening blistering autoimmune disease. Several forms, characterized by the presence of autoantibodies against different autoantigens, have been described. In Pemphigus Vulgaris (PV), autoantibodies target the cadherin Desmoglein 3 (DSG3), while in Pemphigus foliaceous (PF) autoantibodies target the cadherin Desmoglein 1 (DSG1). Another variant, mucocutaneous Pemphigus, is characterized by the presence of IgG against both DSG1 and DSG3. Moreover, other forms of Pemphigus characterized by the presence of autoantibodies against other autoantigens have been described. With regard to animal models, one can distinguish between passive models, where pathological IgG are transferred into neonatal mice, and active models, where B cells deriving from animals immunized against a specific autoantigen are transferred into immunodeficient mice that develop the disease. Active models recreate PV and a form of Pemphigus characterized by the presence of IgG against the cadherin Desmocollin 3 (DSC3). Further approaches allow to collect sera or B/T cells from mice immunized against a specific antigen to evaluate the mechanisms underlying the onset of the disease. Objective: To develop and characterize a new active model of Pemphigus where mice express auto antibodies against either DSG1 alone, or DSG1 and DSG3, thereby recapitulating PF and mucocutaneous Pemphigus, respectively. In addition to the existing models, with the active models reported in this work, it will be possible to recapitulate and mimic the main forms of pemphigus in adult mice, thus allowing a better understanding of the disease in the long term, including the benefit/risk ratio of new therapies. Results: The new DSG1 and the DSG1/DSG3 mixed models were developed as proposed. Immunized animals, and subsequently, animals that received splenocytes from the immunized donors produce a high concentration of circulating antibodies against the specific antigens. The severity of the disease was assessed by evaluating the PV score, evidencing that the DSG1/DSG3 mixed model exhibits the most severe symptoms among those analyzed. Alopecia, erosions, and blistering were observed in the skin of DSG1, DSG3 and DSG1/DSG3 models, while lesions in the mucosa were observed only in DSG3 and DSG1/DSG3 animals. The effectiveness of the corticosteroid Methyl-Prednisolone was evaluated in the DSG1 and DSG1/DSG3 models, that showed only partial responsiveness.

In-Silico Analysis of Differentially Expressed Genes and Their Regulating microRNA Involved in Lymph Node Metastasis in Invasive Breast Carcinoma

Axillary nodal metastasis is related to poor prognosis in breast cancer (BC). Key candidate genes in BC lymph node metastasis have been identified from Gene Expression Omnibus datasets and explored through functional enrichment database for annotation, visualization and integrated discovery (DAVID) , protein-protein interaction by Search Tool for the Retrieval of Interacting Genes and proteins (STRING), network visualization (Cytoscape), survival analysis (GEPIA, KM Plotter), and target prediction (miRNet). A total of 102 overlapping differentially expressed genes were found. In-silico survival and expression analyses revealed six candidate hub genes, Desmocollin 3 (DSC3), KRT5, KRT6B, KRT17, KRT81, and SERPINB5, to be significantly associated with nodal metastasis and overall survival, and 83 MicroRNA (miRNAs), which may be potential diagnostic markers and therapeutic targets in BC patients.

Human Desmocollin 3‒Specific IgG Antibodies Are Pathogenic in a Humanized HLA Class II Transgenic Mouse Model of Pemphigus

Pemphigus is a potentially lethal autoimmune bullous skin disorder, which is associated with IgG autoantibodies against desmoglein (DSG) 3 and DSG1. Notably, a subset of patients with pemphigus presents with a similar clinical phenotype in the absence of anti-DSG IgG, suggesting the presence of serum IgG reactive with desmosomal components other than DSG1 or DSG3. We and others have previously shown that such patients have serum IgG autoantibodies against desmocollin 3 (DSC3), a component of desmosomes, which induce loss of keratinocyte adhesion exvivo. Moreover, DSC3 hypomorphic mice show a severe blistering phenotype of the mucous membrane, which is highly characteristic of pemphigus. These findings prompted us to study the induction and regulation of anti-human DSC3 IgG in humanized mice transgenic for HLA-DRB1∗04:02, which is a highly prevalent haplotype in pemphigus. We show that IgG from sera of immunized mice induces acantholysis in a dispase-based keratinocyte dissociation assay through the activation of p38 MAPKs and EGFR. Passive IgG transfer from mice immunized with recombinant human DSC3 into neonates did not induce intraepidermal loss of adhesion presumably owing to the lack of homology between human and mouse DSC3. Exvivo stimulation of splenocytes from DSC3-immunized mice with human DSC3 leads to a significant proliferative IFN-γ and IL-4 T-cell response, which is restricted by HLA-DR/HLA-DQ. These findings suggest that the induction of pathogenic anti-DSC3 IgG is associated with DSC3-specific T cells that recognize DSC3 in association with HLA-DRB1∗04:02.

Homozygous Nonsense Mutation in DSC3 Resulting in Skin Fragility and Hypotrichosis

Homozygous Nonsense Mutation in DSC3 Resulting in Skin Fragility and Hypotrichosis

Desmocollin 3 has a tumor suppressive activity through inhibition of AKT pathway in colorectal cancer

Desmocollin 3 (DSC3) is a transmembrane adhesion protein of desmosomes and involved in carcinogenesis in various cancer types. Downregulation of DSC3 has been reported in colorectal cancer (CRC). However, the function of DSC3 in CRC has not yet been elucidated. In this study, we performed cell-based functional analysis after DSC3 overexpression by stable transfection and knockdown by siRNA in CRC cells. It turned out that overexpression of DSC3 reduced cell proliferation, colony forming ability, induced G0/G1 cell cycle arrest and promoted apoptosis. Further pathway analysis showed that overexpression of DSC3 significantly inhibited the activity of AKT pathway and increased the expression of E-cadherin as well as p53 and p21. In contrast, siRNA-mediated knockdown of DSC3 increased cell proliferation and colony formation, activated the AKT pathway and decreased the expression of E-cadherin as well as p53 and p21. Additionally, in primary CRC patient samples, the expression of DSC3 protein was significantly related to the expression of desmocollin 1 (DSC1) and desmocollin 2 (DSC2) as well as E-cadherin (p < 0.001 respectively). Taken together, our data reveal that DSC3 suppresses CRC cell growth through inhibition of AKT pathway and regulation of E-cadherin. DSC3 may serve as a novel therapeutic target for CRC.

A Homozygous Nonsense Mutation in the DSG3 Gene Causes Acantholytic Blisters in the Oral and Laryngeal Mucosa

A Homozygous Nonsense Mutation in the DSG3 Gene Causes Acantholytic Blisters in the Oral and Laryngeal Mucosa

Association of DSC3 mRNA Down-Regulation in Prostate Cancer with Promoter Hypermethylation and Poor Prognosis

BackgroundDesmocollin 3 (DSC3), a member of the cadherin gene superfamily, is associated with pathogenesis of some cancers, but its role in prostate cancer (PCa) remains largely unknown.MethodsDSC3 gene expression level in available PCa microarray dataset was examined using the Oncomine database. DSC3 transcript expression in prostate cell line panel and an independent tissue cohort (n = 52) was estimated by quantitative PCR (Q-PCR). Epigenetic status of DSC3 gene promoter in PCa was investigated by uploading three dataset (ENCODE Infinium 450K array data and two methylation sequencing) in UCSC genome browser. While pyrosequencing analysis measured promoter DNA methylation, Q-PCR estimates were obtained for DSC3 transcript re-expression after 5-Aza-deoxycytidine (5-Aza) treatment. Clinical relevance of DSC3 expression was studied by Kaplan-Meier survival analysis. Finally, functional studies monitoring cell proliferation, migration and invasion were performed in prostate cell lines after siRNA mediated DSC3 knockdown or following 5-Aza induced re-expression. EMT markers Vimentin and E-cadherin expression was measured by Western Blot.ResultsMicroarray data analyses revealed a significant decrease in DSC3 transcript expression in PCa, compared to benign samples. Q-PCR analysis of an independent cohort revealed DSC3 transcript down-regulation, both in PCa cell lines and tumor tissues but not in their benign counterpart. Examination of available NGS and Infinium data identified a role for epigenetic regulation DSC3 mRNA reduction in PCa. Pyrosequencing confirmed the increased DSC3 promoter methylation in cancer cell lines and restoration of transcript expression upon 5-Aza treatment further corroborated this epigenetic silencing mechanism. Importantly Kaplan-Meier analysis of an outcome cohort showed an association between loss of DSC3 expression and significantly increased risk of biochemical recurrence. Functional studies indicate a role for epithelial–mesenchymal transition in DSC3 regulated cell migration/invasion.ConclusionTaken together, our data suggests that DNA methylation contributes to down-regulation of DSC3 in prostate cancer, and loss of DSC3 predicts poor clinical outcome.

Regulation of Desmocollin3 Expression by Promoter Hypermethylation is Associated with Advanced Esophageal Adenocarcinomas.

BACKGROUND: Desmocollin3 (DSC3) is a member of the cadherin superfamily of calcium-dependent cell adhesion molecules and plays an important role in tumor invasion and metastasis. In this study, we investigated the epigenetic mechanism that regulates DSC3 expression in esophageal adenocarcinomas (EACs). METHODS: Expression of DSC3 was analyzed by quantitative real-time polymerase chain reaction (qRT-PCR). The promoter DNA methylation level of DSC3 was examined using quantitative bisulfite pyrosequencing. RESULTS: The qRT-PCR analysis demonstrated significant down-regulation of the DSC3 mRNA levels in human EAC cell lines and tissue samples (P<.001). In addition, the EAC cell lines and tumor samples have aberrant promoter hypermethylation as compared to normal esophageal samples (P<.001). DSC3 promoter hypermethylation (>10% methylation level) was detected in 97.5% (39/40) of EAC samples whereas none of the normal tissue samples showed hypermethylation (P<.0001). There was a significant inverse correlation between promoter DNA methylation levels and mRNA expression folds for DSC3 (coefficient r=-0.685, P<.0001). Treatment of FLO-1 and SKGT4 EAC cells with 5-Aza-deoxytidine led to a significant reduction in the promoter DNA methylation levels with restoration of the DSC3 expression, suggesting that promoter DNA methylation is a key epigenetic mechanism regulating DSC3 expression. High DSC3 promoter DNA methylation levels were significantly correlated with advanced tumor stage (P<.001) and lymph node metastasis (P<.001). CONCLUSION: Taken together, our results demonstrate that epigenetic silencing of DSC3 is a frequent finding in EAC that is possibly associated with advanced stages.

Heterogeneous and abnormal localization of desmosomal proteins in oral intraepithelial neoplasms.

To study the relationship between the biological and morphological characteristics of oral intraepithelial neoplasms (OINs), we examined the localization of desmosome-related proteins. Twenty-seven cases of OIN3 were tentatively classified as basaloid (14 cases) or differentiated (13 cases), and the latter were further subdivided into verrucous (five cases) and acanthotic (eight cases) subtypes. All samples were stained using antibodies against desmoglein 1 (DSG1), desmocollin 3 (DSC3), junction plakoglobin (JUP) and serine peptidase inhibitor Kazal type 5 (SPINK5) domain. All variants of OIN3 showed significantly high rates of positivity for DSG1 in the basal layer (basaloid 57%; differentiated 85%), DSC3 in the surface layer (basaloid 93%; differentiated 77%) and JUP in the basal and parabasal layers (basaloid 93%; differentiated 62%). Interestingly, even the basaloid type showed areas of alternating DSG1 positivity and negativity, reflecting keratinocyte maturation. Therefore, most cases of OIN appear to have the characteristics of well differentiated squamous epithelium.

Differential expression of cell adhesion molecules in an ionizing radiation-induced breast cancer model system

Cell-cell adhesion is mediated by members of the cadherin-catenin system and among them E-cadherin and β-catenin are important adhesion molecules for epithelial cell function and preservation of tissue integrity. To investigate the importance of cell adhesion molecules in breast carcinogenesis, we developed an in vitro breast cancer model system wherein immortalized human breast epithelial cell line, MCF-10F, was malignantly transformed by exposure to low doses of high linear energy transfer (LET) α particle radiation (150 keV/µm) and subsequent growth in the presence or absence of 17β-estradiol. This model consisted of human breast epithelial cells in different stages of transformation: i) parental cell line MCF-10F; ii) MCF-l0F continuously grown with estradiol at 10(-8) (Estrogen); iii) a non-malignant cell line (Alpha3); and iv) a malignant and tumorigenic cell line (Alpha5) and the Tumor2 cell line derived from the nude mouse xenograft of the Alpha5 cell line. Expression levels of important cell adhesion molecules such as α-catenin, β-catenin, γ-catenin, E-cadherin and integrin were found to be higher at the protein level in the Alpha5 and Tumor2 cell lines relative to these levels in the non-tumorigenic MCF-10F, Estrogen and Alpha3 cell lines. In corroboration, cDNA expression analysis revealed elevated levels of genes involved in the cell adhesion function [E-cadherin, integrin β6 and desmocollin3 (DSc3)] in the Alpha5 and Tumor2 cell lines relative to the levels in the MCF-10F, Estrogen and Alpha3 cell lines. Collectively, our results suggest that cell adhesion molecules are expressed at higher levels in malignantly transformed breast epithelial cells relative to levels in non-malignant cells. However, reduced levels of adhesion molecules observed in the mouse xenograft-derived Tumor 2 cell line compared to the pre-tumorigenic Alpha5 cell line suggests that the altered expression levels of adhesion molecules depend on the tumor tissue microenvironment.

The p53 target gene desmocollin 3 acts as a novel tumor suppressor through inhibiting EGFR/ERK pathway in human lung cancer

Desmosomes are intercellular junctions that confer strong cell-cell adhesion. Altered expression of desmocollin 3 (DSC3), a member of the desmosomal cadherin family, was found in various cancers; however, its functional involvement in carcinogenesis has not yet been elucidated. Expression/localization of DSC3 was analyzed by real-time reverse transcription-PCR, western blotting, immunofluorescence and immunohistochemistry. Methylation status of DSC3 was examined by demethylation tests, methylation-specific PCR and bisulfite sequencing. It turned out that downregulation of DSC3 in lung cancer cells was associated with DNA hypermethylation. In primary lung tumors, DSC3 was a potential diagnostic marker for lung squamous cell carcinoma, and DSC3 DNA hypermethylation was correlated with poor clinical outcome. To investigate the effect of the tumor suppressor gene p53 on DSC3, transient transfection with a wild-type p53-expression vector was performed. Overexpression of p53 resulted in an increased expression of DSC3 in a DSC3-unmethylated lung cancer cell line H2170, but not in H1299, a DSC3-methylated cell line. However, combination of p53 transfection with demethylation agent 5-aza-2'-deoxycytidine treatment led to increased expression of DSC3 in H1299 cells. Furthermore, functional studies after stable transfection of a DSC3 expression vector showed that ectopic expression of DSC3 inhibited cell proliferation, anchorage-independent growth, migration, as well as invasion, and most interestingly led to reduced phosphorylation levels of extracellular signal-regulated kinase1/2. Taken together, our data suggested that DSC3 acts as a novel tumor suppressor gene through inhibition of epidermal growth factor receptor/extracellular signal-regulated kinase signaling in lung cancer cells.

Loss of desmocollin 3 in skin tumor development and progression

Desmocollin 3 (DSC3) is a desmosomal cadherin that is required for maintaining cell adhesion in the epidermis as demonstrated by the intra-epidermal blistering observed in Dsc3 null skin. Recently, it has been suggested that deregulated expression of DSC3 occurs in certain human tumor types. It is not clear whether DSC3 plays a role in the development or progression of cancers arising in stratified epithelia such as the epidermis. To address this issue, we generated a mouse model in which Dsc3 expression is ablated in K-Ras oncogene-induced skin tumors. Our results demonstrate that loss of Dsc3 leads to an increase in K-Ras-induced skin tumors. We hypothesize that acantholysis-induced epidermal hyperplasia in the Dsc3 null epidermis facilitates Ras-induced tumor development. Further, we demonstrate that spontaneous loss of DSC3 expression is a common occurrence during human and mouse skin tumor progression. This loss occurs in tumor cells invading the dermis. Interestingly, other desmosomal proteins are still expressed in tumor cells that lack DSC3, suggesting a specific function of DSC3 loss in tumor progression. While loss of DSC3 on the skin surface leads to epidermal blistering, it does not appear to induce loss of cell-cell adhesion in tumor cells invading the dermis, most likely due to a protection of these cells within the dermis from mechanical stress. We thus hypothesize that DSC3 can contribute to the progression of tumors both by cell adhesion-dependent (skin surface) and likely by cell adhesion-independent (invading tumor cells) mechanisms.

DSC3 expression is regulated by p53, and methylation of DSC3 DNA is a prognostic marker in human colorectal cancer

Background:Desmocollin 3 (DSC3), a member of the cadherin superfamily and integral component of desmosomes, is involved in carcinogenesis. However, the role of DSC3 in colorectal cancer (CRC) has not yet been established.Methods:Desmocollin 3 expression in CRC cell lines was analysed by RT–PCR and western blotting. Methylation status of DSC3 was examined by demethylation tests, methylation-specific PCR, and bisulphite sequencing (BS). The regulatory role of p53 was investigated by transfection.Results:Desmocollin 3 was downregulated in CRC cells at mRNA and protein levels. Desmocollin 3 expression was restored in five out of seven cell lines after 5-aza-2′-deoxycytidine (DAC) treatment. A heterogeneous methylation pattern was detected by BS in promoter region and exon 1 of DSC3. Methylation of DSC3 genomic sequences was found in 41% (41 out of 99) of primary CRC, being associated with poor prognosis (P=0.002). Transfection of p53 alone or in combination of DAC increased the DSC3 expression. Similarly, treatment with p53 inducer adriamycin alone or in combination with DAC enhanced DSC3 expression.Conclusions:DNA methylation contributes to downregulation of DSC3 in CRC cell lines. Methylation status of DSC3 DNA is a prognostic marker for CRC. P53 appears to have an important role in regulating DSC3 expression.

Immunohistochemical subtyping of nonsmall cell lung cancer not otherwise specified in fine‐needle aspiration cytology

Histopathological subtyping of nonsmall cell lung cancer (NSCLC) is currently relevant in treatment decision because of a differential activity of specific therapeutic agents. Immunohistochemistry highlights cell differentiation lineages and, in this study, it was applied to maximize the proportion of accurately subtyped NSCLC not otherwise specified (NOS) on fine-needle aspiration cytology (FNAC) samples. Cell blocks from 103 FNAC samples with a morphological diagnosis of NSCLC-NOS were immunostained for cytokeratin (CK) 7, CK5, TTF1, and p63, whereas p40, napsin A (Naps-A), and desmocollin-3 (DSC-3) were only assessed in a subgroup of cases with discordant (CK7 and TTF1+ for nonsquamous, CK5 and p63+ for squamous) findings. Results were correlated with surgical specimens evaluated by morphology alone. Thirty-seven (36%) tumors with CK7/TTF1+ and CK5/p63- corresponded to 35 cases of adenocarcinoma (ADC) and 2 cases of large cell carcinoma, whereas 9 (9%) cases with the reverse immunoprofile were squamous cell carcinoma (SQCC) at surgery (P < .001). Although the remaining 57 cases had different marker combinations, a correlation was found with ADC histology for TTF1+ samples (independent of other markers) and with SQCC for p63+/TTF1- immunophenotype (P < .001). p40 was never expressed in p63+ ADC, whereas Naps-A was restricted to ADC and DSC-3 to SQCC lineage. The percentage of unclassified NSCLC-NOS decreased from 36% to 14%. Combinations of 2 antibodies (TTF1/DSC-3 or p63/Naps-A) in the same section allowed diagnostic optimization in scant cytological samples. [corrected] This 4-antibody panel approach may contribute to refine lung cancer classification in FNAC cell blocks, remarkably reducing the NSCLC-NOS diagnostic category.

Emerging role of thymidylate synthase for the pharmacogenomic selection of patients with thoracic cancer

The use of antifolate drugs, and particularly of pemetrexed, is emerging as a relevant therapeutic strategy for the treatment of some specific thoracic cancers. Pemetrexed is a novel multitargeted antifolate which has been shown in vitro to inhibit at least three different enzymes in the folate pathway: thymidylate synthase (TS), dihydrofolate reductase (DHFR) and glycinamide ribonucleotide formyl transferase (GARFT) of which TS is its main target [1]. These enzymes are involved in the synthesis of nucleotides and interference with their function will ultimately hinder the synthesis of RNA and DNA. Pre-clinical studies have demonstrated the cytotoxic activity of this agent in a broad range of tumor types including non-small cell lung cancer (NSCLC). In vitro studies indicated that tumor cell lines expressing high levels of TS or DHFR have reduced sensitivity to pemetrexed, suggesting that increased expression levels might correlate with reduced clinical efficacy [2]. In NSCLC, which accounts for more than 80% of all lung cancer, two recent independent Phase III trials showed that pemetrexed alone or in combination with cisplatin demonstrated a statistically superior activity in patients with non-squamous histotypes [3, 4]. Although histology has not consistently been associated with clinical outcomes in almost every trial testing the role of platinum-based chemotherapy in advanced NSCLC, it has now emerged as a potential predictive factor due to the evidence of the differential expression of TS between the different tumor histotypes, which seems to play an important role [3]. Preliminary molecular evidence, in fact, has previously shown that TS expression is significantly higher in squamous cell carcinoma compared with adenocarcinoma by means of both Real-Time PCR gene quantification and of immunohistochemistry for the detection of the protein levels [5]. In undifferentiated large cell carcinoma, a subsequent analysis consistently reported significantly higher TS levels in desmocollin 3 (DSC-3)-positive as compared to DSC-3negative tumors (indicating a squamous and adenocarcinoma origin of the tumors, respectively), reporting that lack of DSC-3 immunoreactivity in this histologic subtype is associated with lower TS expression [8]. In the same experimental setting, TS expression levels were tested in 22 cases having a cytological diagnosis of NSCLC with no clear histological definition, and in the corresponding histological samples obtained at the time of subsequent surgical resection. As a result, a significantly high degree of concordance in terms of TS protein expression in matched specimens was detected. This result raises the possibility of using the protein levels, evaluated by immunohistostaining on cytological specimens, to select the patients with unresectable advanced disease to be treated with TS-inhibiting agents. On the other hand, the small cell carcinoma (SCLC) group of patients, comprising approximately 13% of all lung cancers, has shown minimal response to pemetrexed in several studies including a large Phase III study [6]. Again, the explanation for these results may be found at the molecular level, in that SCLC has been shown to have very high TS expression when compared with either well-differentiated pulmonary neuroendocrine tumors or NSCLC [7]. These data once again support the hypothesis that elevated TS expression decreases the response to pemetrexed, and that the determination of TS levels can be used in to exclude those patients who are most likely to be unaffected from the treatment. A clear example of this is found in a very recent retrospective study from our group carried out on malignant pleural mesothelioma (MPM), a disease in which pemetrexed has been previously shown to be clinically active. We showed that TS protein and gene expression levels are significantly correlated with the clinical outcome in a consecutive series of 60 patients with advanced MPM treated with pemetrexed, with or without plat

A Homozygous Nonsense Mutation in the Human Desmocollin-3 ( DSC3) Gene Underlies Hereditary Hypotrichosis and Recurrent Skin Vesicles

Desmosomes are the major players in epidermis and cardiac muscles and contribute to intercellular binding and maintenance of tissue integrity. Two important constituents of desmosomes are transmembrane cadherins named desmogleins and desmocollins. The critical role of these desmosomal proteins in epithelial integrity has been illustrated by their disruption in mouse models and human diseases. In the present study, we have investigated a large family from Afghanistan in which four individuals are affected with hereditary hypotrichosis and the appearance of recurrent skin vesicle formation. All four affected individuals showed sparse and fragile hair on scalp, as well as absent eyebrows and eyelashes. Vesicles filled with thin, watery fluid were observed on the affected individuals' scalps and on most of the skin covering their bodies. A scalp-skin biopsy of an affected individual showed mild hair-follicle plugging. Candidate-gene-based homozygosity linkage mapping assigned the disease locus to 8.30 cM (8.51 Mbp) on chromosome 18q12.1. A maximum multipoint LOD score of 3.30 (theta = 0.00) was obtained at marker D18S877. Sequence analysis of four desmoglein and three desmocollin genes, contained within the linkage interval, revealed a homozygous nonsense mutation (c.2129T>G [p.Leu710X]) in exon-14 of the desmocollin-3 (DSC3) gene.

Baseline thymidylate synthase expression according to histological subtypes of non-small cell lung cancer

7521 Background: In non-small cell lung cancer (NSCLC) baseline thymidilate synthase (TS) levels are higher in squamous cell carcinoma (SCC) compared to adenocarcinoma (AC) and randomized clinical trials have shown a selective benefit for patients with non-squamous histology treated with pemetrexed, a TS-inhibiting agent. TS expression in undifferentiated large cell carcinoma (LCC) is unknown. Methods: TS expression at both mRNA (using tissue microdissection and qRT-PCR) and protein (through immunohistochemistry, IHC) levels was tested in 34 surgically resected LCC (stage I=20,II=6,IIIa=8) and compared with TS expression in surgical cases of SCC (n= 31) and AC (n=40). In addition other comparisons were made: a) TS protein expression with Ki-67 index; b) TS mRNA and E2F1 transcription factor mRNA; c) in all histotypes TS protein level with desmocollin-3 (DSC-3) immunostaining, a marker of squamous cell differentiation. TS expression level was assessed in a group of patients (n=22) with cytological diagnosis of NSCLC-NOS (not otherwise specified) and compared with TS data in tissue specimens obtained through subsequent bronchial biopsy or surgical resection. Results: Significantly higher median TS levels in LCC compared to AC (p&lt;0.001 for both mRNA and protein values) and SCC compared to AC (p=0.002 mRNA, p&lt;0.001 protein) were detected. A strong correlation between TS mRNA and protein levels were found (p&lt;0.001) in SCC and AC, but not in LCC. TS and both Ki-67 and E2F1 were significantly correlated in AC and SCC (p=0.003 and p=0.05, respectively), but in LCC no correlation was found. In LCC, significantly higher TS levels were observed in DSC3-positive compared to DSC3-negative tumors (p=0.02). A significant correlation between TS IHC scores in matched cytological and corresponding tissue specimens was observed (p&lt;0.001). Conclusions: This study demonstrates and confirms the: a) differential expression of TS among the NSCLC histotypes; b) lack of DSC-3 immunoreactivity in LCC is associated with lower TS expression; c) assessment of TS by IHC in cytological specimens correlates with the corresponding tissue TS expression. [Table: see text]

Loss of desmocollin 3 in mice leads to epidermal blistering

Desmocollin 3 (DSC3) belongs to a subfamily of cadherins and is a major component of desmosomes in keratinocytes of stratified epithelia, such as the epidermis. Based on its amino acid sequence homology to classical cadherins, such as E-cadherin, it has been postulated that DSC3 functions as a cell-adhesion molecule. To test this hypothesis, we assessed the function of DSC3 in the development and maintenance of stratified epithelia, in particular the epidermis and hair follicles. Using a conditional null allele, we show that loss of Dsc3 function in the epidermis causes impaired cell-cell adhesion, leading to intra-epidermal blistering and telogen hair loss. Furthermore, the lesions in Dsc3-null skin resemble those observed in individuals with pemphigus vulgaris (PV), indicating that impaired Dsc3 function could be a potential cause of PV-like inherited or acquired skin blistering diseases.

5-Aza-2′-deoxycytidine-mediated reductions in G9A histone methyltransferase and histone H3 K9 di-methylation levels are linked to tumor suppressor gene reactivation

The epigenetic silencing of tumor suppressor genes is a common event during carcinogenesis, and often involves aberrant DNA methylation and histone modification of gene regulatory regions, resulting in the formation of a transcriptionally repressive chromatin state. Two examples include the antimetastatic, tumor suppressor genes, desmocollin 3 (DSC3) and MASPIN, which are frequently silenced in this manner in human breast cancer. Treatment of the breast tumor cell lines MDA-MB-231 and UACC 1179 with 5-aza-2'-deoxycytidine (5-aza-CdR) induced transcriptional reactivation of both genes in a dose-dependent manner. Importantly, DSC3 and MASPIN reactivation was closely and consistently linked with significant decreases in promoter H3 K9 di-methylation. Moreover, 5-aza-CdR treatment also resulted in global decreases in H3 K9 di-methylation, an effect that was linked to its ability to mediate dose-dependent, post-transcriptional decreases in the key enzyme responsible for this epigenetic modification, G9A. Finally, small interfering RNA (siRNA)-mediated knockdown of G9A and DNMT1 led to increased MASPIN expression in MDA-MB-231 cells, to levels that were supra-additive, verifying the importance of these enzymes in maintaining multiple layers of epigenetic repression in breast tumor cells. These results highlight an additional, complimentary mechanism of action for 5-aza-CdR in the reactivation of epigenetically silenced genes, in a manner that is independent of its effects on DNA methylation, further supporting an important role for H3 K9 methylation in the aberrant repression of tumor suppressor genes in human cancer.

Differential expression and distribution of epithelial adhesion molecules in non-small cell lung cancer and normal bronchus

Background: Changes in epithelial cell interactions have been implicated in carcinogenesis, tumour invasion and metastasis. Aim: To screen for altered expression of epithelial adhesion genes in lung cancer development. Methods:...