Filipe Santos-Silva

Expression of intestine-specific transcription factors, CDX1 and CDX2, in intestinal metaplasia and gastric carcinomas.

Intestinal metaplasia (IM) is part of a stepwise sequence of alterations of the gastric mucosa, leading ultimately to gastric cancer, and is strongly associated with chronic Helicobacter pylori infection. The molecular mechanisms underlying the onset of IM remain elusive. The aim of this study was to assess the putative involvement of two intestine‐specific transcription factors, CDX1 and CDX2, in the pathogenesis of gastric IM and gastric carcinoma. Eighteen foci of IM and 46 cases of gastric carcinoma were evaluated by immunohistochemistry for CDX1 and CDX2 expression. CDX1 was expressed in all foci of IM and in 41% of gastric carcinomas; CDX2 was expressed in 17/18 foci of IM and in 54% of gastric carcinomas. In gastric carcinomas, a strong association was observed between the expression of CDX1 and CDX2, as well as between the intestinal mucin MUC2 and CDX1 and CDX2. No association was observed between the expression of CDX1 and CDX2 and the histological type of gastric carcinoma. In conclusion, these results show that aberrant expression of CDX1 and CDX2 is consistently observed in IM and in a subset of gastric carcinomas. The association of CDX1 and CDX2 with expression of the intestinal mucin MUC2, both in IM and in gastric carcinoma, indirectly implies that CDX1 and CDX2 may be involved in intestinal differentiation along the gastric carcinogenesis pathway. Copyright

Helicobacter pylori induces β3GnT5 in human gastric cell lines, modulating expression of the SabA ligand sialyl–Lewis x

Chronic Helicobacter pylori infection is recognized as a cause of gastric cancer. H. pylori adhesion to gastric cells is mediated by bacterial adhesins such as sialic acid-binding adhesin (SabA), which binds the carbohydrate structure sialyl-Lewis x. Sialyl-Lewis x expression in the gastric epithelium is induced during persistent H. pylori infection, suggesting that H. pylori modulates host cell glycosylation patterns for enhanced adhesion. Here, we evaluate changes in the glycosylation-related gene expression profile of a human gastric carcinoma cell line following H. pylori infection. We observed that H. pylori significantly altered expression of 168 of the 1,031 human genes tested by microarray, and the extent of these alterations was associated with the pathogenicity of the H. pylori strain. A highly pathogenic strain altered expression of several genes involved in glycan biosynthesis, in particular that encoding beta3 GlcNAc T5 (beta3GnT5), a GlcNAc transferase essential for the biosynthesis of Lewis antigens. beta3GnT5 induction was specific to infection with highly pathogenic strains of H. pylori carrying a cluster of genes known as the cag pathogenicity island, and was dependent on CagA and CagE. Further, beta3GnT5 overexpression in human gastric carcinoma cell lines led to increased sialyl-Lewis x expression and H. pylori adhesion. This study identifies what we believe to be a novel mechanism by which H. pylori modulates the biosynthesis of the SabA ligand in gastric cells, thereby strengthening the epithelial attachment necessary to achieve successful colonization.

OCT-1 is over-expressed in intestinal metaplasia and intestinal gastric carcinomas and binds to, but does not transactivate, CDX2 in gastric cells

Intestinal metaplasia (IM) is a preneoplastic lesion of the stomach in which there is transdifferentiation of the gastric mucosa to an intestinal phenotype. The caudal‐related homeobox gene CDX2 encodes an intestine‐specific transcription factor crucial for the regulation of proliferation and differentiation of intestinal cells. In addition, CDX2 is involved in the induction of IM in the stomach. The aim of this study was to access the putative involvement of OCT‐1 in the induction of CDX2 expression de novo in gastric mucosa leading to the onset of IM. OCT‐1 protein expression was evaluated by immunohistochemistry in 31 biopsies with chronic gastritis, 15 biopsies with foci of IM and adjacent gastric mucosa and 42 gastric carcinomas. Furthermore, we evaluated OCT‐1 binding by electrophoretic mobility shift assay and activation of the CDX2 promoter by co‐transfecting a CDX2 promoter/reporter construct with an OCT‐1 expression vector in two gastric carcinoma cell lines, GP220 and MKN45. Our results show that OCT‐1 is expressed in chronic gastritis, particularly when it is adjacent to IM and is expressed in 87% of IM foci. Furthermore, 74% of the gastric carcinomas were positive for OCT‐1 and a strong association was observed between OCT‐1 expression and intestinal‐type carcinoma. We identified that OCT‐1 binds to the CDX2 promoter, although we could not see a transactivation effect in gastric carcinoma cell lines. In conclusion, we observed increased OCT‐1 expression in IM and in intestinal gastric carcinomas and identified the capacity of OCT‐1 to bind to the CDX2 promoter, although we could not demonstrate a direct effect of OCT‐1 in the transactivation of CDX2. Copyright

Transferrin surface‐modified PLGA nanoparticles‐mediated delivery of a proteasome inhibitor to human pancreatic cancer cells

The aim of this study was to develop a drug delivery system based on poly(lactic-co-glycolic acid) (PLGA) nanoparticles for an efficient and targeted action of the proteasome inhibitor bortezomib against pancreatic cancer cells. The PLGA nanoparticles were formulated with a poloxamer, and further surface-modified with transferrin for tumor targeting. The nanoparticles were characterized as polymer carriers of bortezomib, and the cellular uptake and growth inhibitory effects were evaluated in pancreatic cells. Cellular internalization of nanoparticles was observed in normal and cancer cells, but with higher uptake by cancer cells. The sustained release of the loaded bortezomib from PLGA nanoparticles showed cytotoxic effects against pancreatic normal and cancer cells. Noteworthy differential cytotoxicity was attained by transferrin surface-modified PLGA nanoparticles since significant cell growth inhibition by delivered bortezomib was only observed in cancer cells. These findings demonstrate that the ligand transferrin enhanced the targeted delivery of bortezomib-loaded PLGA nanoparticles to pancreatic cancer cells. These in vitro results highlight the transferrin surface-modified PLGA nanoparticles as a promising system for targeted delivery of anticancer drugs.

Impact of MUC1 mucin downregulation in the phenotypic characteristics of MKN45 gastric carcinoma cell line.

Background Gastric carcinoma is the second leading cause of cancer-associated death worldwide. The high mortality associated with this disease is in part due to limited knowledge about gastric carcinogenesis and a lack of available therapeutic and prevention strategies. MUC1 is a high molecular weight transmembrane mucin protein expressed at the apical surface of most glandular epithelial cells and a major component of the mucus layer above gastric mucosa. Overexpression of MUC1 is found in approximately 95% of human adenocarcinomas, where it is associated with oncogenic activity. The role of MUC1 in gastric cancer progression remains to be clarified. Methodology We downregulated MUC1 expression in a gastric carcinoma cell line by RNA interference and studied the effects on cellular proliferation (MTT assay), apoptosis (TUNEL assay), migration (migration assay), invasion (invasion assay) and aggregation (aggregation assay). Global gene expression was evaluated by microarray analysis to identify alterations that are regulated by MUC1 expression. In vivo assays were also performed in mice, in order to study the tumorigenicity of cells with and without MUC1 downregulation in MKN45 gastric carcinoma cell line. Results Downregulation of MUC1 expression increased proliferation and apoptosis as compared to controls, whereas cell-cell aggregation was decreased. No significant differences were found in terms of migration and invasion between the downregulated clones and the controls. Expression of TCN1, KLK6, ADAM29, LGAL4, TSPAN8 and SHPS-1 was found to be significantly different between MUC1 downregulated clones and the control cells. In vivo assays have shown that mice injected with MUC1 downregulated cells develop smaller tumours when compared to mice injected with the control cells. Conclusions These results indicate that MUC1 downregulation alters the phenotype and tumorigenicity of MKN45 gastric carcinoma cells and also the expression of several molecules that can be involved in tumorigenic events. Therefore, MUC1 should be further studied to better clarify its potential as a novel therapeutic target for gastric cancer.

Functionalized gold nanoparticles improve afatinib delivery into cancer cells.

Objectives: A drug delivery system based on colloidal pegylated gold nanoparticles (PEGAuNPs) conjugated with the tyrosine kinase inhibitor afatinib was designed and tested for enhancing the drug activity against pancreatic and NSCLC cells. Methods: PEGAuNPs were synthesized and characterized physicochemically. Confocal imaging was performed to evaluate the nanoparticle (NP) internalization in cancer cells. For cell-cycle distribution analysis, conjugated NPs and afatinib alone were incubated with cells and alterations on the cell-cycle profile subsequently analyzed by total DNA staining. Cancer cell survival and growth inhibition following incubation with afatinib and PEGAuNPs–afatinib (concentrations between 0.007 and 0.500 µM afatinib) were evaluated. Results: A higher cellular uptake of PEGAuNPs was observed by cancer cells. Our data suggest an efficient conjugation of PEGAuNPs with the drug, enhancing the afatinib activity in comparison with afatinib alone. In fact, IC50 and GI50 results obtained show that the PEGAuNPs–afatinib conjugate is ca. 5 and 20 times more potent than afatinib alone in S2-013 and A549 cell lines, respectively. Conclusions: Conjugating PEGAuNPs with afatinib is a promising antitumor delivery system for cancer therapy as it improves drug efficacy, allowing a reduction in drug dose used and minimizing possible toxicity-related side effects.

Reflections on MUC1 glycoprotein: the hidden potential of isoforms in carcinogenesis.

Mucin 1 (MUC1) has been described as the renaissance molecule due to the large set of functions it displays in both normal and neoplastic cells. This membrane‐tethered glycoprotein is overexpressed and aberrantly glycosylated in most epithelial cancers, being involved in several processes related with malignant phenotype acquisition. With a highly polymorphic structure, both in the polypeptide and glycan counterparts, MUC1 variability has been associated with susceptibility to several diseases, including cancer. Biochemical features and biological functions have been characterized upon the full‐length MUC1 protein, remaining to clarify the real impact on cell dynamics of the plethora of MUC1 isoforms. This review aims to encompass a detailed characterization of MUC1 role in carcinogenesis, highlighting recent findings in cell differentiation and uncovering new evidences of MUC1 isoforms involvement in malignant phenotype.

Enhancing the efficiency of bortezomib conjugated to pegylated gold nanoparticles: an in vitro study on human pancreatic cancer cells and adenocarcinoma human lung alveolar basal epithelial cells

ABSTRACT Objectives: Gold nanoparticles have become promising vectors for cancer diagnosis and treatment. The present study investigates the effect of bortezomib (BTZ), a proteasome inhibitor, conjugated with pegylated gold nanoparticles (PEGAuNPs) in pancreatic and lung cancer cells. Methods: Synthesized gold nanoparticles (PEGAuNPs) were conjugated with bortezomib antitumor drug. We investigated the cytotoxicity induced by BTZ conjugated with functionalized gold nanoparticles in vitro, in the human pancreatic (S2-013) and lung (A549) cancer cell lines. Results: We found an efficient of conjugation of BTZ with PEGAuNPs. In vitro assays showed that after 72 h’ incubation with PEGAuNPs-BTZ cancer cells revealed alterations in morphology; also for S2-013 and A549 cancer cells, the IC50 value of free BTZ is respectively 1.5 and 4.3 times higher than the IC50 value of PEGAuNPs-BTZ. Furthermore, for TERT-HPNE, the IC50 value is around 63 times lower for free BTZ than the conjugated nanovehicle. Cell growth inhibition results showed a remarkable enhancement in the effect of BTZ when conjugated with AuNPs. Conclusions: Our findings showed that conjugation with PEGAuNPs enhance the BTZ growth-inhibition effect on human cancer cells (S2-013 and A549) and decreases its toxicity against normal cells (TERT-HPNE).

Correction to: The Cancer, Educate to Prevent Model—the Potential of School Environment for Primary Prevention of Cancer

The affiliation address of Dr. Helena Santos should be Faculty of Economics, University of Porto, Portugal and Dr. Luís Moreira’s should be Research Unit in Education and Community Intervention – RECI & Health School of Vila Nova de Gaia – Piaget Institute.

Can Smartphones Promote Cancer Prevention Behaviours in Healthy Young Adults? A Prospective Study

Cancer prevention should start as early as possible. Young adults would benefit largely from the use of a smartphone app aiming at promoting cancer prevention behaviours. The aims of the study described in this paper are to (1) examine the user participation and engagement with a cancer prevention app in real-life settings and (2) assess changes in the users’ cancer prevention behaviours. A cancer prevention smartphone app called Happy was developed and released to the general population. Data from registered app users’ (N = 3326) demographics and baseline cancer prevention behaviours was analysed. App engagement was measured and all behaviour data logged through the app was analysed to assess the effectiveness of the app in the promotion of cancer prevention behaviours. User demographics and baseline behaviour assessment show that the app reached its intended target population: young adults that generally do not comply with cancer prevention guidelines. Logged behavioural data showed an increased frequency in several cancer prevention behaviours and a significant increase in the overall putative cancer prevention level (p < 0.001). However, user engagement with the app was limited and might condition the long-term impact of such changes. Happy reached its intended population and seems to be an effective tool to promote cancer prevention. Further research is needed to fully assess its long-term persuasive potential.