E-cadherin: A determinant molecule associated with ovarian cancer progression, dissemination and aggressiveness

Ovarian cancer (OC) is the fifth cancer death cause in women worldwide. The malignant nature of this disease stems from its unique dissemination pattern. Epithelial-to-mesenchymal transition (EMT) has been reported in OC and downregulation of Epithelial cadherin (E-cadherin) is a hallmark of this process. However, findings on the relationship between E-cadherin levels and OC progression, dissemination and aggressiveness are controversial. In this study, the evaluation of E-cadherin expression in an OC tissue microarray revealed its prognostic value to discriminate between advanced-and early-stage tumors, as well as serous tumors from other histologies. Moreover, E-cadherin, Neural cadherin (N-cadherin), cytokeratins and vimentin expression was assessed in TOV-112, SKOV-3, OAW-42 and OV-90 OC cell lines grown in monolayers and under anchorage-independent conditions to mimic ovarian tumor cell dissemination, and results were associated with cell aggressiveness. According to these EMT-related markers, cell lines were classified as mesenchymal (M; TOV-112), intermediate mesenchymal (IM; SKOV-3), intermediate epithelial (IE; OAW-42) and epithelial (E; OV-90). M-and IM-cells depicted the highest migration capacity when grown in monolayers, and aggregates derived from M-and IM-cell lines showed lower cell death, higher adhesion to extracellular matrices and higher invasion capacity than E- antigen 125 levels more than 500 U/mL and platinum-free intervals less than 6 months. Altogether, E-cadherin expression levels were found relevant for the assessment of OC progression and aggressiveness.and IE-aggregates. The analysis of E-cadherin, N-cadherin, cytokeratin 19 and vimentin mRNA levels in 20 advanced-stage high-grade serous human OC ascites showed an IM phenotype in all cases, characterized by higher proportions of N-to E-cadherin and vimentin to cytokeratin 19. In particular, higher E-cadherin mRNA levels were associated with cancer antigen 125 levels more than 500 U/mL and platinum-free intervals less than 6 months. Altogether, E-cadherin expression levels were found relevant for the assessment of OC progression and aggressiveness

CDH1/E-cadherin and solid tumors. An updated gene-disease association analysis using bioinformatics tools

Cancer is a group of diseases that causes millions of deaths worldwide. Among cancers, Solid Tumors (ST) stand-out due to their high incidence and mortality rates. Disruption of cell–cell adhesion is highly relevant during tumor progression. Epithelial-cadherin (protein: E-cadherin, gene: CDH1) is a key molecule in cell–cell adhesion and an abnormal expression or/and function(s) contributes to tumor progression and is altered in ST. A systematic study was carried out to gather and summarize current knowledge on CDH1/E-cadherin and ST using bioinformatics resources. The DisGeNET database was exploited to survey CDH1-associated diseases. Reported mutations in specific ST were obtained by interrogating COSMIC and IntOGen tools. CDH1 Single Nucleotide Polymorphisms (SNP) were retrieved from the dbSNP database. DisGeNET analysis identified 609 genes annotated to ST, among which CDH1 was listed. Using CDH1 as query term, 26 disease concepts were found, 21 of which were neoplasms-related terms. Using DisGeNET ALL Databases,172 disease concepts were identified. Of those, 80 ST disease-related terms were subjected to manual curation and 75/80 (93.75%) associations were validated. On selected ST, 489 CDH1 somatic mutations were listed in COSMIC and IntOGen databases. Breast neoplasms had the highest CDH1- mutation rate. CDH1 was positioned among the 20 genes with highest mutation frequency and was confirmed as driver gene in breast cancer. Over 14,000 SNP for CDH1 were found in the dbSNP database. This report used DisGeNET to gather/compile current knowledge on gene-disease association for CDH1/E-cadherin and ST; data curation expanded the number of terms that relate them. An updated list of CDH1 somatic mutations was obtained with COSMIC and IntOGen databases and of SNP from dbSNP. This information can be used to further understand the role of CDH1/E-cadherin in health and disease.Fil: Abascal, Maria Florencia. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental. Fundación de Instituto de Biología y Medicina Experimental. Instituto de Biología y Medicina Experimental; ArgentinaFil: Besso, María José. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental. Fundación de Instituto de Biología y Medicina Experimental. Instituto de Biología y Medicina Experimental; ArgentinaFil: Rosso, Marina. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental. Fundación de Instituto de Biología y Medicina Experimental. Instituto de Biología y Medicina Experimental; ArgentinaFil: Mencucci, Maria Victoria. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental. Fundación de Instituto de Biología y Medicina Experimental. Instituto de Biología y Medicina Experimental; ArgentinaFil: Aparicio, Evangelina. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental. Fundación de Instituto de Biología y Medicina Experimental. Instituto de Biología y Medicina Experimental; ArgentinaFil: Szapiro, Gala. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental. Fundación de Instituto de Biología y Medicina Experimental. Instituto de Biología y Medicina Experimental; ArgentinaFil: Furlong, Laura Ines. Universitat Pompeu Fabra; EspañaFil: Vazquez, Monica Hebe. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental. Fundación de Instituto de Biología y Medicina Experimental. Instituto de Biología y Medicina Experimental; Argentin

Isoform specificity of progesterone receptor antibodies

Progesterone receptors (PR) are prognostic and predictive biomarkers in hormone-dependent cancers. Twomain PR isoforms have been described, PRB and PRA, that differ only in that PRB has 164 extra N-terminalamino acids. It has been reported that several antibodies empirically exclusively recognize PRA in formalinfixedparaffin-embedded (FFPE) tissues. To confirm these findings, we used human breast cancer xenograftmodels, T47D-YA and -YB cells expressing PRA or PRB, respectively, MDA-MB-231 cells modified to synthesizePRB, and MDA-MB-231/iPRAB cells which can bi-inducibly express either PRA or PRB. Cells were injectedinto immunocompromised mice to generate tumours exclusively expressing PRA or PRB. PR isoform expressionwas verified using immunoblots. FFPE samples from the same tumours were studied by immunohistochemistryusing H-190, clone 636, clone 16, and Ab-6 anti-PR antibodies, the latter exclusively recognizing PRB. Exceptfor Ab-6, all antibodies displayed a similar staining pattern. Our results indicate that clones 16, 636, and theH-190 antibody recognize both PR isoforms. They point to the need for more stringency in evaluating thetrue specificity of purported PRA-specific antibodies as the PRA/PRB ratio may have prognostic and predictivevalue in breast cancer.Fil: Fabris, Victoria Teresa. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental. Fundación de Instituto de Biología y Medicina Experimental. Instituto de Biología y Medicina Experimental; ArgentinaFil: Abascal, Maria Florencia. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental. Fundación de Instituto de Biología y Medicina Experimental. Instituto de Biología y Medicina Experimental; ArgentinaFil: Giulianelli, Sebastian Jesus. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Centro Nacional Patagónico. Instituto de Biología de Organismos Marinos; ArgentinaFil: May, María. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica; ArgentinaFil: Sequeira, Gonzalo Ricardo. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental. Fundación de Instituto de Biología y Medicina Experimental. Instituto de Biología y Medicina Experimental; ArgentinaFil: Jacobsen, Britta. University of Colorado Anschutz Medical Campus; Estados UnidosFil: Lombès, Marc. Inserm; FranciaFil: Han, Julie. Moore´s Cancer Center; Estados UnidosFil: Tran, Luan. Moore´s Cancer Center; Estados UnidosFil: Molinolo, Alfredo. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental. Fundación de Instituto de Biología y Medicina Experimental. Instituto de Biología y Medicina Experimental; ArgentinaFil: Lanari, Claudia Lee Malvina. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental. Fundación de Instituto de Biología y Medicina Experimental. Instituto de Biología y Medicina Experimental; Argentin

E-cadherin: A determinant molecule associated with ovarian cancer progression, dissemination and aggressiveness

Ovarian cancer (OC) is the fifth cancer death cause in women worldwide. The malignant nature of this disease stems from its unique dissemination pattern. Epithelial-to-mesenchymal transition (EMT) has been reported in OC and downregulation of Epithelial cadherin (E-cadherin) is a hallmark of this process. However, findings on the relationship between E-cadherin levels and OC progression, dissemination and aggressiveness are controversial. In this study, the evaluation of E-cadherin expression in an OC tissue microarray revealed its prognostic value to discriminate between advanced-and early-stage tumors, as well as serous tumors from other histologies. Moreover, E-cadherin, Neural cadherin (N-cadherin), cytokeratins and vimentin expression was assessed in TOV-112, SKOV-3, OAW-42 and OV-90 OC cell lines grown in monolayers and under anchorage-independent conditions to mimic ovarian tumor cell dissemination, and results were associated with cell aggressiveness. According to these EMT-related markers, cell lines were classified as mesenchymal (M; TOV-112), intermediate mesenchymal (IM; SKOV-3), intermediate epithelial (IE; OAW-42) and epithelial (E; OV-90). M-and IM-cells depicted the highest migration capacity when grown in monolayers, and aggregates derived from M-and IM-cell lines showed lower cell death, higher adhesion to extracellular matrices and higher invasion capacity than E- antigen 125 levels more than 500 U/mL and platinum-free intervals less than 6 months. Altogether, E-cadherin expression levels were found relevant for the assessment of OC progression and aggressiveness.and IE-aggregates. The analysis of E-cadherin, N-cadherin, cytokeratin 19 and vimentin mRNA levels in 20 advanced-stage high-grade serous human OC ascites showed an IM phenotype in all cases, characterized by higher proportions of N-to E-cadherin and vimentin to cytokeratin 19. In particular, higher E-cadherin mRNA levels were associated with cancer antigen 125 levels more than 500 U/mL and platinum-free intervals less than 6 months. Altogether, E-cadherin expression levels were found relevant for the assessment of OC progression and aggressiveness

FGF2 Induces Breast Cancer Growth through Ligand-Independent Activation and Recruitment of ERα and PRB∆4 Isoform to MYC Regulatory Sequences

Progression to hormone-independent growth leading to endocrine therapy resistance occurs in a high proportion of patients with estrogen receptor alpha (ERα) and progesterone receptors (PR) positive breast cancer. We and others have previously shown that estrogen- and progestin-induced tumor growth requires ERα and PR interaction at their target genes. Here, we show that fibroblast growth factor 2 (FGF2)-induces cell proliferation and tumor growth through hormone-independent ERα and PR activation and their interaction at the MYC enhancer and proximal promoter. MYC inhibitors, antiestrogens or antiprogestins reverted FGF2-induced effects. LC?MS/MS identified 700 canonical proteins recruited to MYC regulatory sequences after FGF2 stimulation, 397 of which required active ERα (ERα-dependent). We identified ERα-dependent proteins regulating transcription that, after FGF2 treatment, were recruited to the enhancer as well as proteins involved in transcription initiation that were recruited to the proximal promoter. Also, among the ERα-dependent and independent proteins detected at both sites, PR isoforms A and B as well as the novel protein product PRBΔ4 were found. PRBΔ4 lacks the hormone-binding domain and was able to induce reporter gene expression from estrogen-regulated elements and to increase cell proliferation when cells were stimulated with FGF2 but not by progestins. Analysis of the Cancer Genome Atlas data set revealed that PRBΔ4 expression is associated with worse overall survival in luminal breast cancer patients. This discovery provides a new mechanism by which growth factor signaling can engage nonclassical hormone receptor isoforms such as PRBΔ4, which interacts with growth-factor activated ERα and PR to stimulate MYC gene expression and hence progression to endocrine resistance.Fil: Giulianelli, Sebastian Jesus. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Centro Nacional Patagónico. Instituto de Biología de Organismos Marinos; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental. Fundación de Instituto de Biología y Medicina Experimental. Instituto de Biología y Medicina Experimental; ArgentinaFil: Riggio, Marina. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental. Fundación de Instituto de Biología y Medicina Experimental. Instituto de Biología y Medicina Experimental; ArgentinaFil: Guillardoy, Tomás. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental. Fundación de Instituto de Biología y Medicina Experimental. Instituto de Biología y Medicina Experimental; ArgentinaFil: Pérez Piñero, Cecilia. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental. Fundación de Instituto de Biología y Medicina Experimental. Instituto de Biología y Medicina Experimental; ArgentinaFil: Gorostiaga, María A.. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental. Fundación de Instituto de Biología y Medicina Experimental. Instituto de Biología y Medicina Experimental; ArgentinaFil: Sequeira, Gonzalo Ricardo. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental. Fundación de Instituto de Biología y Medicina Experimental. Instituto de Biología y Medicina Experimental; ArgentinaFil: Pataccini, Gabriela. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental. Fundación de Instituto de Biología y Medicina Experimental. Instituto de Biología y Medicina Experimental; ArgentinaFil: Abascal, María F.. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental. Fundación de Instituto de Biología y Medicina Experimental. Instituto de Biología y Medicina Experimental; ArgentinaFil: Toledo, María Florencia. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental. Fundación de Instituto de Biología y Medicina Experimental. Instituto de Biología y Medicina Experimental; ArgentinaFil: Jacobsen, Britta M.. University of Colorado; Estados UnidosFil: Guerreiro, Ana C.. Universidade de Aveiro; PortugalFil: Barros, António. Universidade de Aveiro; PortugalFil: Novaro, Virginia. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental. Fundación de Instituto de Biología y Medicina Experimental. Instituto de Biología y Medicina Experimental; ArgentinaFil: Monteiro, Fátima L.. Universidade de Aveiro; PortugalFil: Amado, Francisco. Universidade de Aveiro; PortugalFil: Gass, Hugo. Hospital Zonal General de Agudos Magdalena V de Martínez; ArgentinaFil: Abba, Martin. Universidad Nacional de La Plata; ArgentinaFil: Helguero, Luisa A.. Universidade de Aveiro; PortugalFil: Lanari, Claudia. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental. Fundación de Instituto de Biología y Medicina Experimental. Instituto de Biología y Medicina Experimental; Argentin

Schematic representation of the OC dissemination process.

<p>Along epithelial ovarian tumor progression E-cadherin expression levels decrease with tumor FIGO stages, contributing to the shedding of OC cells into the abdominal cavity. Individual cells could then aggregate in suspension and form multicellular structures with different expression levels of E- and N-cadherin, as well as of cytokeratins and vimentin. According to the expression of these EMT-related markers, cell aggregates could be classified as mesenchymal (M; expression of N-cadherin and vimentin, and absence of E-cadherin and cytokeratins detectable levels), intermediate mesenchymal (IM; expression of N-cadherin, E-cadherin, cytokeratins and vimentin, with a high N- to E-cadherin proportion), intermediate epithelial (IE; expression of N-cadherin, E-cadherin, cytokeratins and vimentin, with a high E- to N-cadherin proportion) and epithelial (E; expression of E-cadherin and cytokeratins, and absence of N-cadherin and vimentin detectable levels).Those aggregates with an mesenchymal-like phenotype (M and IM) will be able to survive under anchorage-independent conditions and to adhere and invade the mesothelium lining, leading to metastasis and a worse patient prognosis. However, aggregates with an epithelial-like phenotype (E and IE) will be more prompted to undergo apoptosis.</p

E-cadherin mRNA expression, CA125 levels and PFI in OC ascites.

<p>E-cadherin mRNA expression, CA125 levels and PFI in OC ascites.</p

Assessment of adhesion, disaggregation and invasion capacity of OC cell lines grown under anchorage-independent conditions.

<p><b>(A)</b> Representative phase contrast images of 48 hour-aggregates (black spots) placed onto fibronectin and collagen I matrices (40x magnification) (left). The number (#) of aggregates adhered to each matrix (Fibronectin: white, Collagen I: black) after 2 hour-incubation was plotted (right) (***p<0.001). <b>(B)</b> Representative phase contrast images of the area of an SKOV-3 aggregate placed onto collagen I over time (0, 6, 9, 30 hours) are shown (left) (100x magnification). Estimated area (px²: pixeles²) of 48 hour-aggregates placed onto fibronectin (white) and collagen I (black) for 30 hours (right) (**p<0.01, *p<0.05). <b>(C)</b> Phase contrast images and immunofluorescence analysis of paxillin in 48 hour-aggregates placed onto fibronectin and collagen I for 24 hours (200x and 400x magnification, respectively). <b>(D)</b> Phase contrast images of 48 hour-aggregates 2 and 7 days after placing them into Matrigel^TM (200x magnification).</p

Expression analyses of E-cadherin and EMT-related markers in OC cell lines grown under anchorage-independent conditions.

<p><b>Assessment of their aggregation and survival capacities. (A)</b> Phase contrast images of 48 hour-aggregates (100x and 200x magnifications). <b>(B)</b> Plot of the area (px²: pixeles²) and number (black spots) of 48 hour-aggregates in 4 drops of each cell line. <b>(C)</b> Western immunoblotting analyses of E-cadherin, N-cadherin, pan-cytokeratin and vimentin in 48 hour-aggregates. β-tubulin served as total protein loading control. <b>(D)</b> Quantitative real time PCR analyses of E-cadherin and N-cadherin mRNA expression levels in 48 hour-aggregates. <b>(E)</b> Fluorescent immunocytochemistry analysis of E-cadherin and N-cadherin in 48 hour-aggregates (400x magnification). A merge image of both cadherins is also included. <b>(F)</b> Cell death assessed by means of PI staining in 48 hour-aggregates. Images were taken using an inverted microscope with phase contrast and red fluorescence after PI staining of disaggregated cells (100x magnification) (top). Cell death (%) was plotted (bottom) (***p<0.001). <b>(G)</b> Western immunoblotting analysis of PARP-1 on 48 hour-aggregates (left). Relative expression (%) of cleaved versus FL PARP-1 form (right).</p

E-cadherin, N-cadherin, cytokeratin 19 and vimentin mRNA expression levels in human ovarian tumor-primary cultures.

<p>E-cadherin, N-cadherin, cytokeratin 19 and vimentin mRNA expression levels in human ovarian tumor-primary cultures.</p