Cadherin Expression Profiles Define Glioblastoma Differentiation and Patient Prognosis.

Cadherins are cell-cell adhesion proteins which have been strongly implicated in cancer invasion, dissemination and metastasis capacity; thus, they are key players in the epithelial-to-mesenchymal transition (EMT) program. However, their role in glioblastoma (GBM), a primary central nervous system aggressive tumor, remains to be clarified. N-, E- and P-cadherin expression was analyzed on a large series of GBMs, characterized with clinical, imaging and neuropathological parameters, as well as with patients' survival data. In addition, cadherins' expression was studied in match-recurrent cases. Using TCGA data, cadherin expression profiles were also evaluated according to GBM transcription subtypes. N-cadherin expression was observed in 81.5% of GBM, followed by E-cadherin in 31% and P-cadherin in 20.8%. Upon tumor recurrence, P-cadherin was the only significantly upregulated cadherin compared with the primary tumor, being positive in 65.8% of the cases. Actually, P-cadherin gain was observed in 51.4% of matched primary-recurrent cases. Cadherins' co-expression was also explored. Interestingly, E- and N-cadherin co-expression identified a GBM subgroup with frequent epithelial differentiation and a significant survival benefit. On the other hand, subgroups with P-cadherin expression carried the worse prognosis. P- and N-cadherin co-expression correlated with the presence of a mesenchymal phenotype. Expressions of isolated P-cadherin or E- and P-cadherin co-expression were associated with imaging characteristics of aggressiveness, to highly heterogeneous tumors, an d to worse patient survival. Classical cadherins co-expression subgroups present consistent clinical, imaging, neuropathological and survival differences, which probably reflect different states of an EMT-like program in GBM.

Cardiovascular risk assessment in Portugal's primary health care system: SCORE vs. SCORE2.

INTRODUCTION AND OBJECTIVES: The 2021 European Society of Cardiology guidelines on cardiovascular disease (CVD) prevention introduced the more accurate SCORE2 risk model as a replacement for the earlier SCORE, which is still used in primary care software in Portugal. Our objective is to determine whether the difference between risk assessment using SCORE and SCORE2, in the same patient population, is statistically significant. METHODS: A total of 1642 patients aged 40-65 without previous CVD, from the medical records of two Family Health Units, were included in this cross-sectional study. SCORE and SCORE2 were calculated using the variables gender, age, smoking status, lipid profile and systolic blood pressure. A statistical analysis was performed on the results. RESULTS: Using SCORE, 98% of the patients were in the low-moderate risk categories and 2% in the high or very high risk categories. When using SCORE2, the corresponding percentages were 55% and 45%, respectively. Reclassification with SCORE2 into higher categories was more often observed in younger (under 50 years of age) and male patients. With SCORE, 38.61% of patients were within the LDL-C target range; this figure fell to 20.28% with SCORE2. These differences are statistically significant (p<0.0001). CONCLUSION: Our findings show that a significant number of patients in this cohort who were classified through SCORE at lower risk levels were reclassified into higher risk categories with SCORE2. Similarly, the number of patients within the LDL-C target range for LDL-C was also lower using SCORE2.

Severe Pediatric Sleep Apnea: Drug-Induced Sleep Endoscopy Based Surgery.

Although adenotonsillectomy is the recommended treatment of obstructive sleep apnea (OSA) in children, some patients with preoperative severe OSA (Apnea-hypopnea index/AHI > 10) remain symptomatic after surgery and may need further workup. This study aims to: (1) analyse preoperative factors and its relation with surgical failure/persistent OSA (AHI > 5 after adenotonsillectomy) in severe pediatric OSA; (2) determine the levels of airway collapse during DISE (drug induced sleep endoscopy) in cases of surgical failure; (3) evaluate the efficacy of targeted surgery based on DISE findings. This retrospective study was conducted between August and September 2020. Across 9 years (from 2011 to 2020), all children diagnosed with severe OSA in our Hospital underwent adenotonsillectomy and repeated type 1 polysomnography (PSG) 3 months after surgery. Cases of surgical failure underwent DISE for planning eventual directed surgery. Chi-square test was used to assess the relationship between persistent OSA and preoperative patients' characteristics. 80 cases of severe pediatric OSA were diagnosed (68.8% males; mean age: 4.3 years-standard deviation: 2.49; mean AHI: 16.3-standard deviation 7.14) in the aforementioned period. We found a significant association between surgical failure (11.3% of cases; mean AHI: 6.9-SD 0.91) and obesity (p = 0.002; confidence level of 95%). Neither preoperative AHI nor other PSG parameters were associated with surgical failure. In cases of surgical failure, epiglottis collapse was present in every DISEs and adenoid tissue was present in 66% of children. All cases of surgical failure had directed surgery and surgical cure (AHI ≤ 5) was obtained in 100% of cases. This study suggests that obesity is the strongest predictor of surgical failure in children with severe OSA who undergo adenotonsillectomy. Epiglottis collapse and presence of adenoid tissue are the most common findings in postoperative DISEs of children with persistent OSA after primary surgery. DISE based surgery seems a safe and effective tool to manage persistent OSA after adenotonsillectomy.

Epithelial-Mesenchymal Plasticity Induced by Discontinuous Exposure to TGFß1 Promotes Tumour Growth.

Transitions between epithelial and mesenchymal cellular states (EMT/MET) contribute to cancer progression. We hypothesize that EMT followed by MET promotes cell population heterogeneity, favouring tumour growth. We developed an EMT model by on and off exposure of epithelial EpH4 cells (E-cells) to TGFß1 that mimics phenotypic EMT (M-cells) and MET. We aimed at understanding whether phenotypic MET is accompanied by molecular and functional reversion back to epithelia by using RNA sequencing, immunofluorescence (IF), proliferation, wound healing, focus formation and mamosphere formation assays as well as cell xenografts in nude mice. Phenotypic reverted epithelial cells (RE-cells) obtained after MET induction presented epithelial morphologies and proliferation rates resembling E cells. However, the RE transcriptomic profile and IF staining of epithelial and mesenchymal markers revealed a uniquely heterogeneous mixture of cell subpopulations with a high self-renewal ability. RE cell heterogeneity was stably maintained for long periods after TGFß1 removal both in vitro and in large tumours derived from the nude mice. Overall, we show that phenotypic reverted epithelial cells (RE cells) do not return to the molecular and functional epithelial state and present mesenchymal features related to aggressiveness and cellular heterogeneity that favour tumour growth in vivo. This work strengthens epithelial cell reprogramming and cellular heterogeneity fostered by inflammatory cues as a tumour growth-promoting factor in vivo.

Characterisation of an Atrx Conditional Knockout Mouse Model: Atrx Loss Causes Endocrine Dysfunction Rather Than Pancreatic Neuroendocrine Tumour.

ATRX is a chromatin remodeller that maintains telomere homeostasis. Loss of ATRX is described in approximately 10% of pancreatic neuroendocrine tumours (PanNETs) and associated with poorer prognostic features. Here, we present a genetically engineered mouse model (GEMM) addressing the role of Atrx loss (AtrxKO) in pancreatic ß cells, evaluating a large cohort of ageing mice (for up to 24 months (mo.)). Atrx loss did not cause PanNET formation but rather resulted in worsening of ageing-related pancreatic inflammation and endocrine dysfunction in the first year of life. Histopathological evaluation highlighted an exacerbated prevalence and intensity of pancreatic inflammation, ageing features, and hepatic steatosis in AtrxKO mice. Homozygous floxed mice presented hyperglycaemia, increased weights, and glucose intolerance after 6 months, but alterations in insulinaemia were not detected. Floxed individuals presented an improper growth of their pancreatic endocrine fraction that may explain such an endocrine imbalance. A pilot study of BRACO-19 administration to AtrxKO mice resulted in telomere instability, reinforcing the involvement of Atrx in the maintenance of ß cell telomere homeostasis. Thereby, a non-obese dysglycaemic GEMM of disrupted Atrx is here presented as potentially useful for metabolic studies and putative candidate for inserting additional tumourigenic genetic events.

Unilateral vocal fold paralysis as a port complication.

Unilateral vocal fold paralysis (UVFP) is a frequent finding in otorhinolaryngology practice, but its occurrence as a port complication was very rarely described in English Literature. The authors report a 55-year-old woman with a pancreatic adenocarcinoma who presented a left vocal fold paralysis that occurred concurrently with a venous thrombosis of the left subclavian vein, where a totally implantable venous-access had been previously placed. Although the patient's oncologic disease, that could mislead to a neoplastic cause of the UVFP, the authors came across with an unusual etiology and to their best knowledge, it is the first case of irreversible UVFP associated with onsite thrombosis of the vessel where a port was implanted. The objective of this article is to present and discuss this rare case of UVFP secondary to a port complication and to review the main mechanisms of iatrogenic vocal fold paralysis related to these devices.

Biotinylated Polymer-Ruthenium Conjugates: In Vitro and In Vivo Studies in a Triple-Negative Breast Cancer Model.

The need for new therapeutic approaches for triple-negative breast cancer is a clinically relevant problem that needs to be solved. Using a multi-targeting approach to enhance cancer cell uptake, we synthesized a new family of ruthenium(II) organometallic complexes envisaging simultaneous active and passive targeting, using biotin and polylactide (PLA), respectively. All compounds with the general formula, [Ru(η5-CpR)(P)(2,2'-bipy-4,4'-PLA-biotin)][CF3SO3], where R is -H or -CH3 and P is P(C6H5)3, P(C6H4F)3 or P(C6H4OCH3)3, were tested against triple-negative breast cancer cells MDA-MB-231 showing IC50 values between 2.3-14.6 µM, much better than cisplatin, a classical chemotherapeutic drug, in the same experimental conditions. We selected compound 1 (where R is H and P is P(C6H5)3), for further studies as it was the one showing the best biological effect. In a competitive assay with biotin, we showed that cell uptake via SMVT receptors seems to be the main transport route into the cells for this compound, validating the strategy of including biotin in the design of the compound. The effects of the compound on the hallmarks of cancer show that the compound leads to apoptosis, interferes with proliferation by affecting the formation of cell colonies in a dose-dependent manner and disrupts the cell cytoskeleton. Preliminary in vivo assays in N: NIH(S)II-nu/nu mice show that the concentrations of compound 1 used in this experiment (maximum 4 mg/kg) are safe to use in vivo, although some signs of liver toxicity are already found. In addition, the new compound shows a tendency to control tumor growth, although not significantly. In sum, we showed that compound 1 shows promising anti-cancer effects, bringing a new avenue for triple-negative breast cancer therapy.

Upregulation of tRNA-Ser-AGA-2-1 Promotes Malignant Behavior in Normal Bronchial Cells.

Serine tRNAs (tRNASer) are frequently overexpressed in tumors and associated with poor prognosis and increased risk of recurrence in breast cancer. Impairment of tRNA biogenesis and abundance also impacts proteome homeostasis, and activates protein quality control systems. Herein, we aimed at testing whether increasing tRNASer abundance could foster tumor establishment through activation of the UPR. In order to do so, firstly we confirmed that the expression of tRNA-Ser-AGA-2-1 [hereafter tRNASer(AGA)] was upregulated by 1.79-fold in Stage I NSCLC tumors when compared to normal adjacent tissue. To study the impact of tRNASer(AGA) in early stage tumorigenesis, we induced its upregulation in a non-tumoral bronchial cell line, BEAS-2B. Upregulation of this tRNA increased cellular proliferation and protein synthesis rate, driven by eIF2α dephosphorylation and ATF4 activation downstream of PERK signaling. Futhermore, tRNASer(AGA) enhanced transformation potential in vitro, and promoted the establishment of slow growing tumors with aggressive features in nude mice. Our work highlights the importance of studying tRNA deregulation on early stage tumorigenesis, as they may be potential malignancy and aggressiveness biomarkers.

Regulation of invasion and peritoneal dissemination of ovarian cancer by mesothelin manipulation.

Peritoneal dissemination is a particular form of metastasis typically observed in ovarian cancer and the major cause for poor patient's outcome. Identification of the molecular players involved in ovarian cancer dissemination can offer an approach to develop treatment strategies to improve clinical prognosis. Here, we identified mesothelin (MSLN) as a crucial protein in the multistep process of peritoneal dissemination of ovarian cancer. We demonstrated that MSLN is overexpressed in primary and matched peritoneal metastasis of high-grade serous carcinomas (HGSC). Using several genetically engineered ovarian cancer cell lines, resulting in loss or gain of function, we found that MSLN increased cell survival in suspension and invasion of tumor cells through the mesothelial cell layer in vitro. Intraperitoneal xenografts established with MSLNhigh ovarian cancer cell lines showed enhanced tumor burden and spread within the peritoneal cavity. These findings provide strong evidences that MSLN is a key player in ovarian cancer progression by triggering peritoneal dissemination and provide support for further clinical investigation of MSLN as a therapeutic target in HGSC.

Expression of CD44v6-Containing Isoforms Influences Cisplatin Response in Gastric Cancer Cells.

CD44v6-containing isoforms are frequently de novo expressed in gastric cancer (GC). Whether CD44v6 has a central role in GC transformation and/or progression, whether it conditions response to therapy or whether it is only a bystander marker is still not known. Therefore, we aimed to clarify the role of CD44v6 in GC. We generated GC isogenic cell lines stably expressing CD44s or CD44v6 and tested them for different cancer hallmarks and response to cisplatin, and we further confirmed our findings in cells that endogenously express CD44v6. No correlation between overexpression of CD44v6 and the tested cancer hallmarks was observed, suggesting CD44v6 is not a driver of GC progression. Upon cisplatin treatment, CD44v6+ cells survive better and have lower apoptosis levels than CD44v6- cells, possibly due to concomitant activation of STAT3 and P38. In co-culture experiments, we discovered that CD44v6+ cells are involved in GC cell overgrowth after cisplatin treatment. In conclusion, we show that CD44v6 expression increases cell survival in response to cisplatin treatment in GC cells and that these cells override CD44v6-negative cells after cisplatin-treatment. This suggests that tumor expression of CD44v6-containing variants may condition the outcome of GC patients treated with chemotherapy.

O contributo da educação física para a a formação de indivíduos fisicamente cultos: uma perspetiva portuguesa / The contribution of physical education to the training of physically educated individuals: a portuguese perspective / Las aportaciones de la educación física para la formación de individuos físicamente cultos: una perspectiva portuguesa

Resumo: Este trabalho, realizado com 150 indivíduos no final da escolaridade obrigatória e candidatos a uma licenciatura em Ciências do Desporto, pretende perceber se a Educação Física está a contribuir para a formação de indivíduos fisicamente cultos. Seguimos uma metodologia quantitativa e optámos por um questionário com três partes: a primeira de questões fechadas e de caracterização sociodemográfica da amostra; a segunda, também de questões fechadas, sobre o entendimento acerca das Finalidades da Educação Física; e a terceira composta por 25 afirmações de escolha dicotómica (verdadeiro/falso), sobre conteúdos fundamentais para uma prática autónoma de atividade física, constantes nos Programas Nacionais de Educação Física (Currículos Nacionais) para o ensino secundário. Os resultados revelam fragilidades nos conhecimentos dos indivíduos relativamente ao desenvolvimento das capacidades motoras, aos efeitos da prática de exercício físico, a procedimentos de segurança e à composição e funcionamento corporal, afastando-se do preconizado por aqueles Programas Nacionais.

Abstract: This study was carried out with 150 male students at the end of their mandatory education and with candidates to a degree in Sports Science and Physical Education. It aims to understand if Physical Education contributed to the education of physically educated individuals. We followed a quantitative methodology and administered a three-part questionnaire: close-ended questions for the sociodemographic characterization of the sample; close-ended questions on participants' understanding of the purposes of Physical Education; true-or-false questions on fundamental contents for an autonomous practice of physical activity, which are included in the National Curricula of Physical Education for secondary school. The results reveal individuals' weak knowledge of motor skills development, effects of physical exercise, safety procedures, and body composition and functioning, which is not in accordance with National Curricula.

Resumen: Este trabajo, realizado con 150 estudiantes al final de su escolaridad obligatoria y aspirantes a cursar la licenciatura en Ciencias del Deporte, pretende averiguar si la Educación Física está contribuyendo con la formación de individuos físicamente cultos. Seguimos una metodología cuantitativa y optamos por un cuestionario con tres partes: la primera con cuestiones cerradas y de caracterización sociodemográfica de los participantes; la segunda, también con cuestiones cerradas y sobre su entendimiento acerca de las finalidades de la Educación Física; y la tercera, compuesta por 25 afirmaciones de selección dicotómica (verdadero y falso), sobre contenidos fundamentales para una práctica autónoma de la actividad física y que constan en los Programas Nacionales de Educación Física (Currículos Nacionales) para el bachillerato. Los resultados revelan fragilidades en los conocimientos de los participantes en lo referente al desarrollo de las capacidades motoras, los efectos de la práctica de ejercicio físico, los procedimientos de seguridad y la composición y funcionamiento corporal, alejándose de lo preconizado en aquellos Programas Nacionales.

The Antitumor Activity of a Lead Thioxanthone is Associated with Alterations in Cholesterol Localization.

The search for novel anticancer small molecules and strategies remains a challenge. Our previous studies have identified TXA1 (1-{[2-(diethylamino)ethyl]amino}-4-propoxy-9H- thioxanthen-9-one) as a hit compound, with in vitro antitumor potential by modulating autophagy and apoptosis in human tumor cell lines. In the present study, the mechanism of action and antitumor potential of the soluble salt of this molecule (TXA1.HCl) was further investigated using in vitro and mouse xenograft tumor models of NSCLC. Our results showed that TXA1.HCl affected steroid biosynthesis, increased RagD expression, and caused abnormal cellular cholesterol localization. In addition, TXA1.HCl treatment presented no toxicity to nude mice and significantly reduced the growth of human NSCLC cells xenografts in mice. Overall, this work provides new insights into the mechanism of action of TXA1, which may be relevant for the development of anticancer therapeutic strategies, which target cholesterol transport.

Afadin Downregulation by Helicobacter pylori Induces Epithelial to Mesenchymal Transition in Gastric Cells.

Afadin is a cytoplasmic protein of the adherens junctions, which regulates the formation and stabilization of both the adherens and the tight junctions. Aberrant expression of Afadin has been shown in cancer and its loss has been associated with epithelial-to-mesenchymal transition (EMT). EMT is characterized by the change from an epithelial to a mesenchymal phenotype, with modifications on the expression of adhesion molecules and acquisition of a migratory and invasive cell behavior. While it is known that Helicobacter pylori disrupts the tight and the adherens junctions and induces EMT, the effect of the bacteria on Afadin is still unknown. The aim of this study was to disclose the effect of H. pylori on Afadin and its impact in the induction of an EMT phenotype in gastric cells. Using two different cell lines, we observed that H. pylori infection decreased Afadin protein levels, independently of CagA, T4SS, and VacA virulence factors. H. pylori infection of cell lines recapitulated several EMT features, displacing and downregulating multiple proteins from cell-cell junctions, and increasing the expression of ZEB1, Vimentin, Slug, N-cadherin, and Snail. Silencing of Afadin by RNAi promoted delocalization of junctional proteins from the cell-cell contacts, increased paracellular permeability, and decreased transepithelial electrical resistance, all compatible with impaired junctional integrity. Afadin silencing also led to increased expression of the EMT marker Snail, and to the formation of actin stress fibers, together with increased cell motility and invasion. Finally, and in line with our in vitro data, the gastric mucosa of individuals infected with H. pylori showed decrease/loss of Afadin membrane staining at cell-cell contacts significantly more frequently than uninfected individuals. In conclusion, Afadin is downregulated by H. pylori infection in vitro and in vivo, and its downregulation leads to the emergence of EMT and to the acquisition of an aggressive phenotype in gastric cells, which can contribute to gastric carcinogenesis.

SRC inhibition prevents P-cadherin mediated signaling and function in basal-like breast cancer cells.

BACKGROUND: Basal-like breast cancer (BLBC) is a poor prognosis subgroup of triple-negative carcinomas that still lack specific target therapies and accurate biomarkers for treatment selection. P-cadherin is frequently overexpressed in these tumors, promoting cell invasion, stem cell activity and tumorigenesis by the activation of Src-Family kinase (SRC) signaling. Therefore, our aim was to evaluate if the treatment of BLBC cells with dasatinib, the FDA approved SRC inhibitor, would impact on P-cadherin induced tumor aggressive behavior. METHODS: P-cadherin and SRC expression was evaluated in a series of invasive Breast Cancer and contingency tables and chi-square tests were performed. Cell-cell adhesion measurements were performed by Atomic Force Microscopy, where frequency histograms and Gaussian curves were applied. 2D and 3D cell migration and invasion, proteases secretion and self-renew potential were evaluated in vitro. Student's t-tests were used to determine statistically significant differences. The cadherin/catenin complex interactions were evaluated by in situ proximity-ligation assay, and statistically significant results were determined by using Mann-Whitney test with a Bonferroni correction. In vivo xenograft mouse models were used to evaluate the impact of dasatinib on tumor growth and survival. ANOVA test was used to evaluate the differences in tumor size, considering a confidence interval of 95%. Survival curves were estimated by the Kaplan-Meier's method, using the log-rank test to assess significant differences for mice overall survival. RESULTS: Our data demonstrated that P-cadherin overexpression is significantly associated with SRC activation in breast cancer cells, which was also validated in a large series of primary tumor samples. SRC activity suppression with dasatinib significantly prevented the in vitro functional effects of P-cadherin overexpressing cells, as well as their in vivo tumorigenic and metastatic ability, by increasing mice overall survival. Mechanistically, SRC inhibition affects P-cadherin downstream signaling, rescues the E-cadherin/p120-catenin complex to the cell membrane, recovering cell-cell adhesion function. CONCLUSIONS: In conclusion our findings show that targeting P-cadherin/SRC signaling and functional activity may open novel therapeutic opportunities for highly aggressive and poor prognostic basal-like breast cancer.

Mucins and Truncated O-Glycans Unveil Phenotypic Discrepancies between Serous Ovarian Cancer Cell Lines and Primary Tumours.

Optimal research results rely on the selection of cellular models capable of recapitulating the characteristics of primary tumours from which they originate. The expression of mucins (MUC16 and MUC1) and truncated O-glycans (Tn, STn and T) represents a characteristic footprint of serous ovarian carcinomas (SOCs). Therefore, selecting ovarian cancer (OVCA) cell lines that reflect this phenotype is crucial to explore the putative biological role of these biomarkers in the SOC setting. Here, we investigated a panel of OVCA cell lines commonly used as SOC models, and tested whether, when cultured in 2D and 3D conditions, these recapitulate the mucin and O-glycan expression profiles of SOCs. We further explored the role of truncating the O-glycosylation capacity in OVCAR3 cells through knockout of the COSMC chaperone, using in vitro and in vivo assays. We found that the majority of OVCA cell lines of serous origin do not share the mucin and truncated O-glycan footprint of SOCs, although 3D cultures showed a higher resemblance. We also found that genetic truncation of the O-glycosylation capacity of OVCAR3 cells did not enhance oncogenic features either in vitro or in vivo. This study underscores the importance of well-characterized cellular models to study specific features of ovarian cancer.

Codon misreading tRNAs promote tumor growth in mice.

Deregulation of tRNAs, aminoacyl-tRNA synthetases and tRNA modifying enzymes are common in cancer, raising the hypothesis that protein synthesis efficiency and accuracy (mistranslation) are compromised in tumors. We show here that human colon tumors and xenograft tumors produced in mice by two epithelial cancer cell lines mistranslate 2- to 4-fold more frequently than normal tissue. To clarify if protein mistranslation plays a role in tumor biology, we expressed mutant Ser-tRNAs that misincorporate Ser-at-Ala (frequent error) and Ser-at-Leu (infrequent error) in NIH3T3 cells and investigated how they responded to the proteome instability generated by the amino acid misincorporations. There was high tolerance to both misreading tRNAs, but the Ser-to-Ala misreading tRNA was a more potent inducer of cell transformation, stimulated angiogenesis and produced faster growing tumors in mice than the Ser-to-Leu misincorporating tRNA. Upregulation of the Akt pathway and the UPR were also observed. Most surprisingly, the relative expression of both misreading tRNAs increased during tumor growth, suggesting that protein mistranslation is advantageous in cancer contexts. These data highlight new features of protein synthesis deregulation in tumor biology.

Porphyrin modified trastuzumab improves efficacy of HER2 targeted photodynamic therapy of gastric cancer.

Gastric cancer (GC) is the 3rd deadliest cancer worldwide, due to limited treatment options and late diagnosis. Human epidermal growth factor receptor-2 (HER2) is overexpressed in ∼20% of GC cases and anti-HER2 antibody trastuzumab in combination with conventional chemotherapy, is recognized as standard therapy for HER2-positive metastatic GC. This strategy improves GC patients' survival by 2-3 months, however its optimal results in breast cancer indicate that GC survival may be improved. A new photoimmunoconjugate was developed by conjugating a porphyrin with trastuzumab (Trast:Porph) for targeted photodynamic therapy in HER2-positive GC. Using mass spectrometry analysis, the lysine residues in the trastuzumab structure most prone for porphyrin conjugation were mapped. The in vitro data demonstrates that Trast:Porph specifically binds to HER2-positive cells, accumulates intracellularly, co-localizes with lysosomal marker LAMP1, and induces massive HER2-positive cell death upon cellular irradiation. The high selectivity and cytotoxicity of Trast:Porph based photoimmunotherapy is confirmed in vivo in comparison with trastuzumab alone, using nude mice xenografted with a HER2-positive GC cell line. In the setting of human disease, these data suggest that repetitive cycles of Trast:Porph photoimmunotherapy may be used as an improved treatment strategy in HER2-positive GC patients.

In Vivo Performance of a Ruthenium-cyclopentadienyl Compound in an Orthotopic Triple Negative Breast Cancer Model.

BACKGROUND: Ruthenium-based anti-cancer compounds are proposed as viable alternatives that might circumvent the disadvantages of platinum-based drugs, the only metallodrugs in clinical use for chemotherapy. Organometallic complexes in particular hold great potential as alternative therapeutic agents since their cytotoxicity involves different modes of action and present reduced toxicity profiles. OBJECTIVE: During the last few years our research group has been reporting on a series of organometallic ruthenium(II)- cyclopentadienyl complexes with important cytotoxicity against several cancer cell lines, surpassing cisplatin in activity. We report herein preliminary in vivo studies with one representative compound of this family, with exceptional activity against several human cancer cell lines, including the glycolytic and highly metastatic MDAMB231 cell line used in this study. METHOD: The anti-tumor activity of our compound was studied in vivo on N:NIH(S)II-nu/nu nude female mice bearing triple negative breast cancer (TNBC) orthotopic tumors. Administration of 2.5 mg/kg/day during ten days caused cell death mostly by necrosis (in vitro and in vivo), inducing tumor growth suppression of about 50% in treated animals when compared to controls. RESULTS: The most remarkable result supporting the effectiveness and potential of this drug was the absence of metastases in the main organs of treated animals, while metastases were present in the lungs of all control mice, as revealed by histopathological and immunohistochemical analysis. CONCLUSION: These in vivo studies suggest a dual effect for our drug not only by suppressing growth at the primary tumor tissue but also by inhibiting its metastatic behavior. Altogether, these results represent a benchmark and a solid starting point for future studies.

Morphological features and mucin expression profile of breast carcinomas with signet-ring cell differentiation.

Signet-ring cells are relatively common in breast cancers but are frequently overlooked. Although previously defined as a subtype of mucin producing carcinomas, breast carcinomas with signet-ring cell (SRC) differentiation nowadays are not considered a distinct entity. The objective of the present study was to characterize the morphological features and mucin expression profile of breast carcinomas with SRC differentiation. All breast carcinomas diagnosed at Centro Hospitalar S. Joao between 1996 and 2006 in which the pathology report mentioned the presence of SRCs (n=11) and four mucinous carcinomas were included in the study. The frequency of SRCs and immunohistochemistry expression of MUC1/MUC2/MUC5AC/MUC6 were evaluated. We confirmed that SRC differentiation can occur in different histological types, including ductal, lobular, mucinous and metaplastic carcinomas. The proportion of SRCs was highly variable (range: 8-70%). Tumors encompassed SRCs of intracytoplasmic lumina and goblet-cell type. A higher percentage of SRCs was associated with lymphovascular invasion (p=0.047). All tumors expressed cytoplasmic and membranous MUC1. Secretory mucins were more frequent in mucinous carcinomas and in carcinomas with extensive SRC differentiation. We conclude that besides the usefulness of mucin immunodetection for the differential diagnosis of carcinomas with SRC differentiation of breast origin, it is important to report SRC differentiation regardless of histological type because of its intrinsic prognostic value.

Anti-influenza neuraminidase inhibitor oseltamivir phosphate induces canine mammary cancer cell aggressiveness.

Oseltamivir phosphate is a widely used anti-influenza sialidase inhibitor. Sialylation, governed by sialyltransferases and sialidases, is strongly implicated in the oncogenesis and progression of breast cancer. In this study we evaluated the biological behavior of canine mammary tumor cells upon oseltamivir phosphate treatment (a sialidase inhibitor) in vitro and in vivo. Our in vitro results showed that oseltamivir phosphate impairs sialidase activity leading to increased sialylation in CMA07 and CMT-U27 canine mammary cancer cells. Surprisingly, oseltamivir phosphate stimulated, CMT-U27 cell migration and invasion capacity in vitro, in a dose-dependent manner. CMT-U27 tumors xenograft of oseltamivir phosphate-treated nude mice showed increased sialylation, namely α2,6 terminal structures and SLe(x) expression. Remarkably, a trend towards increased lung metastases was observed in oseltamivir phosphate-treated nude mice. Taken together, our findings revealed that oseltamivir impairs canine mammary cancer cell sialidase activity, altering the sialylation pattern of canine mammary tumors, and leading, surprisingly, to in vitro and in vivo increased mammary tumor aggressiveness.