Sialyl Tn-Expressing Bladder Cancer Cells Induce a Tolerogenic Phenotype in Innate and Adaptive Immune Cells

Despite the wide acceptance that glycans are centrally implicated in immunity, exactly how they contribute to the tilt immune response remains poorly defined. In this study, we sought to evaluate the impact of the malignant phenotype-associated glycan, sialyl-Tn (STn) in the function of the key orchestrators of the immune response, the dendritic cells (DCs). In high grade bladder cancer tissue, the STn antigen is significantly overexpressed and correlated with the increased expression of ST6GALNAC1 sialyltransferase. Bladder cancer tissue presenting elevated expression of ST6GALNAC1 showed a correlation with increased expression of CD1a, a marker for bladder immature DCs and showed concomitant low levels of Th1-inducing cytokines IL-12 and TNF-α. In vitro, human DCs co-incubated with STn+ bladder cancer cells, had an immature phenotype (MHC-IIlow, CD80low and CD86low) and were unresponsive to further maturation stimuli. When contacting with STn+ cancer cells, DCs expressed significantly less IL-12 and TNF-α. Consistent with a tolerogenic DC profile, T cells that were primed by DCs pulsed with antigens derived from STn+ cancer cells were not activated and showed a FoxP3high IFN-γlow phenotype. Blockade of STn antigens and of STn+ glycoprotein, CD44 and MUC1, in STn+ cancer cells was able to lower the induction of tolerance and DCs become more mature. Overall, our data suggest that STn-expressing cancer cells impair DC maturation and endow DCs with a tolerogenic function, limiting their capacity to trigger protective anti-tumour T cell responses. STn antigens and, in particular, STn+ glycoproteins are potential targets for circumventing tumour-induced tolerogenic mechanisms.

Tumors of the Brachial Plexus in a Tertiary Referral Center: a Case Series and Literature Review

Introduction: Brachial plexus (BP) tumors are very rare tumors, with less than 800 cases been described in the literature worldwide since 1970. These tumors often present as local or radicular pain, with scant or no neurological deficits. These symptoms are shared by many other more common rheumatologic diseases, thus making their diagnosis difficult in most cases. Additionally, these tumors often present as lumps and are therefore biopsied, which carries a significant risk of iatrogenic nerve injury. Material and Methods: In this paper the authors describe their experience with the management of 5 patients with BP tumors followed up for at least 2 years. There were 4 males and 1 female. Median follow-up time was 41 ± 21 months. Average age at diagnosis was 40,0 ± 19,9 years. The most common complaints at presentation were pain and sensibility changes. All patients had a positive Tinel sign when the lesion was percussed. In all patients surgery was undertaken and the tumors removed. In 4 patients nerve integrity was maintained. In one patient with excruciating pain a segment of the nerve had to be excised and the nerve defect was bridged with sural nerve grafts. Results: Pathology examination of the resected specimens revealed a Schwannoma in 4 cases and a neurofibroma in the patient submitted to segmental nerve resection. Two years postoperatively, no recurrences were observed. All patients revealed clinical improvement. The patient submitted to nerve resection had improvement in pain, but presented diminished strength and sensibility in the involved nerve territory. Conclusion: Surgical excision of BP tumors is not a risk free procedure. Most authors suggest surgery if the lesion is symptomatic or progressing in size. If the tumor is stationary and not associated with neurological dysfunction a conservative approach should be taken.

Inflammatory Pseudotumor of the Urinary Bladder

We report a case of an inflammatory pseudotumor of the urinary bladder in a 31 year-old woman. She presented at the emergency room with low abdominal pain and urinary symptoms. Abdominal ultrasound, computed tomography and magnetic resonance imaging were performed and revealed asymmetric thickening of the urinary bladder wall. Cystoscopy with urinary cytology revealed a benign nature of the process. The patient underwent partial cystectomy and the pathologic examination of the specimen revealed an inflammatory pseudotumor. We reviewed the clinical, imaging and pathological features of the inflammatory pseudotumor of the urinary bladder and discussed its differential diagnosis.

Analysis of PAX2 gene alterations in renal cell tumors

Dissertation for applying to a Master’s Degree in Molecular Genetics and Biomedicine submitted to the Sciences and Technology Faculty of New University of Lisbon

Thomsen-Friedenreich antigens in bladder cancer: evaluation of their prognostic value

RESUMO: Introdução. O cancro de bexiga é uma patologia comum que representa o 6° e o 5° cancro mais incidente em Portugal e na Itália, respetivamente. Em mais de metade dos casos ocorre reincidência durante o primeiro ano, requerendo acompanhamento clínico ao longo da vida. A instilação intravesical de Bacillus Calmette-Guérin (BCG) (uma estirpe atenuada do Mycobacterium bovis) representa uma imunoterapia eficaz no combate ao cancro de bexiga, no entanto, muitos aspetos da interação de BCG com as células tumorais bem como com as células do sistema imunitário permanecem por desvendar. As células tumorais de bexiga expressam frequentemente as formas sialiladas dos antigénios de Thomsen-Friedenreich (TF), i.e., sialil-T (sT) e sialil-Tn (sTn). Contudo ainda se desconhece o significado da sua expressão na malignidade tumoral e se afeta a eficácia da terapêutica BCG. Objetivo do estudo. Investigar o papel dos antigénios sT e sTn no fenótipo maligno de células de cancro de bexiga bem como na resposta mediada pelo sistema imunitário à terapia com BCG. Metodologia. Para tal, foram utilizadas as linhas celulares de cancro da bexiga HT1376 e MCR, geneticamente modificadas por transdução com vetores codificantes para as sialiltransferases ST3GAL1 ou ST6GALNAC1, de forma a expressar homogeneamente os antigénios sT ou sTn respetivamente. Estes modelos celulares foram estudados após confronto com BCG. O nível de BCG internalizado foi avaliado por citometria de fluxo. O perfil global de expressão genética dos modelos celulares antes e após incubação com BCG foi analisado pela tecnologia de microarray. O perfil de citocinas secretadas pelos modelos celulares após incubação com BCG, bem como de macrófagos estimulados pelo secretoma de células de cancro de bexiga que por sua vez foram estimuladas previamente por BCG, foi estudado pelo sistema multiplex de “imuno-esferas”. Resultados. A análise do transcritoma dos modelos celulares revelou que grupos de genes envolvidos em funções específicas foram modulados em paralelo nos dois modelos celulares, após transdução, independentemente da sialiltransferase expressa. Ou seja, em células que expressavam a sialiltransferase ST3GAL1 ou ST6GALNAC1, os genes envolvidos na regulação da segregação cromossómica e na reparação do DNA foram consistentemente regulados negativamente. Genes descritos na literatura como marcadores para o cancro de bexiga foram também modulados. A incubação com BCG resultou numa tendência ao aumento da expressão de genes relevantes na preservação e estabilidade genómica e menor malignidade, no entanto, apenas em células que expressavam sT ou sTn. Entre as dez citocinas testadas, apenas a IL-6 e IL-8 foram expressas pelas linhas celulares de cancro da bexiga, com indução destas após estimulação com BCG, e principalmente em células que expressavam ST3GAL1 ou ST6GALNAC1. Em macrófagos, citocinas inflamatórias, tais como IL-1β, IL-6 e TNFα, e a citocina anti-inflamatória IL-10, foram induzidas apenas pelo secretoma de células de cancro da bexiga confrontadas com BCG, com maior relevância quando estas expressavam ST3GAL1 ou ST6GALNAC1, prevendo a estimulação de macrófagos semelhantes aos de tipo M1 e uma melhor resposta à terapia com BCG. Conclusões. O efeito geral da expressão destas sialiltransferases e dos produtos enzimáticos sT ou sTn nas células de cancro de bexiga conduz a um fenótipo de maior malignidade. Contudo, a maior avidez de estas na produção de citocinas inflamatórias após confronto com BCG, bem como a maior capacidade de estimulação de macrófagos, predirá uma resposta à terapia com BCG mais eficaz em tumores que expressem os antigénios de TF sialilados. Tais conclusões são totalmente concordantes com os nossos mais recentes dados clínicos obtidos em colaboração, que mostram que em doentes com cancro de bexiga que expressam sTn respondem melhor a terapia BCG. ----------ABSTRACT: Background. Bladder cancer is a common malignancy representing the 6th and the 5th most incident cancer in Portugal and in Italy, respectively. More than half of the cases relapse within one year, requiring though a lifelong follow-up. Intravesical instillation of Bacillus Calmette-Guérin (BCG) (an attenuated strain of Mycobacterium bovis) represents an effective immunotherapy of bladder cancer, although many aspects of the interaction of BCG with cancer cells and host immune cells remain obscure. Bladder cancer cells often express the sialylated forms of the Thomsen-Friedenreich (TF), i.e., sialil-T (sT) e sialil-Tn (sTn). However, it’s still unknown the sense of such expression in tumour malignancy and in the BCG therapy efficacy. Aim of the study. To investigate the role of the sT and sTn antigens on the malignant phenotype of bladder cancer cells and the immune mediated response to BCG therapy. Experimental. We have utilized populations of the bladder cancer cell lines HT1376 and MCR, genetically modified by transduction with the sialyltransferases ST3GAL1 or ST6GALNAC1 to express homogeneously sT or sTn antigens. The level of BCG internalized was assessed by flow cytometry. The whole gene expression profile of BCG-challenged or unchallenged bladder cancer cell lines was studied by microarray technology. The profile of cytokines secreted by BCG-challenged bladder cancer cells and that of macrophages challenged by the secretome of BCG-challenged bladder cancer cells was studied by multiplex immune-beads assay. Results. Transcriptome analysis of the sialyltransferase-transduced cells revealed that groups of genes involved in specific functions were regulated in parallel in the two cell lines, regardless the sialyltransferase expressed. Namely, in sialyltransferase-expressing cells, genes involved in the proper chromosomal segregation and in the DNA repair were consistently down-regulated, while genes reported in literature as markers for bladder cancer were modulated. BCG-challenging induced a tendency to up-regulation of the genes preserving genomic stability and reducing malignancy, but only in cells expressing either sT or sTn. Among the ten cytokines tested, only IL-6 and IL-8 were expressed by bladder cancer cell lines and up-regulated by BCG-challenging, mainly in sialyltransferases-expressing cells. In macrophages, inflammatory cytokines, such as IL-1β, IL-6 and TNFα, and the antinflammatory IL-10 were induced only by the secretome of BCG-challenged bladder cancer cells, particularly when expressing either sialyltransferase, predicting the stimulation of M1-like macrophages and a better response to BCG therapy. Conclusions. The general effect of the expression of the two sialyltransferases and their products in the bladder cancer cells is toward a more malignant phenotype. However, the stronger ability of sialyltransferase expressing cells to produce inflammatory cytokines upon BCG-challenging and to stimulate macrophages predicts a more effective response to BCG in tumours expressing the sialylated TF antigens. This is fully consistent with our recent clinical data obtained in collaboration, showing that patients with bladder cancer expressing sTn respond better to BCG therapy.

Specific features of endothelial cells in primary tumors

RESUMO: As células endoteliais definem e delineiam todo o sistema vascular...Nesta tese procurámos explorar o papel que o ambiente tumoral exerce sobre as células endoteliais. ... Avaliamos também a capacidade anti-angiogénica de alguns derivados do estrogénio... Em suma os nossos resultados mostram a importância de um controlo rigoroso da regulação transcricional...

The use of botulinum toxin type A in the treatment of HTLV-1-associated overactive bladder refractory to conventional therapy

Urinary symptoms occur in 19% of human T-cell lymphotropic virus type 1 (HTLV-1)-infected patients who do not fulfill criteria for HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP) and in almost 100% of HAM/TSP patients. Few studies have evaluated therapies for overactive bladder (OAB) caused by HTLV-1 infection. This case report describes the effect of onabotulinum toxin A on the urinary manifestations of three patients with HAM/TSP and OAB symptoms. The patients were intravesically administered 200 units of Botox®. Their incontinence episodes improved, and their OAB symptoms scores (OABSS) reduced significantly. These data indicate that Botox® should be a treatment option for OAB associated with HTLV-1 infection.

Detection of codon 12 mutation in the k-ras oncogene in pancreatic tumors

Mutations at codons 12, 13, or 61 of the H-ras, K-ras, and N-ras have been detected in human neoplasias by a variety of techniques. Some of these techniques are very sensitive and can detect K-ras mutation in 90% of the cases of pancreatic adenocarcinomas. We analyzed 11 samples of pancreatic adenocarcinoma, three samples of pancreatic mucinous cystadenoma, and two samples without tumors in formalin-fixed paraffin embedded tissue sections. K-ras mutations at codon 12 were detected by a two-step PCR-enriched technique in all the samples of pancreatic adenocarcinoma, but not in cystadenoma or control samples. This technique may be useful for early detection of pancreatic cancer.

Megabladder in experimental Chagas disease: pathological features of the bladder wall

Mega-organs, primarily in the digestive tract, are well known to occur in chronic Chagas disease. Acute experimental infection with Trypanosoma cruzi results in parasitism of a wide range of cells, tissues, and organs, including the urinary bladder. Infection of BALB/c mice with 100,000 bloodstream forms of the Y strain of T. cruzi induced acute infection with intense parasitism of all layers of the urinary bladder. Parasites were found in the mucosa, lamina propria, muscular, adventitial connective, and fat tissue. Desquamate epithelial cells with amastigotes in the bladder lumen were also found. After 60 days of infection, mice inoculated with 50 bloodstream forms developed dilated, thin-walled bladders that had inflammatory infiltrates and foci of fibrosis replacing areas of damaged muscular layer. These lesions result from direct damage to the muscle fibers by the T. cruzi, leading to myosites, muscle damage, and scarring. Direct damage of paraganglia cells secondary to parasitism, leading to dilatation, damage of muscle fibers, and scarring with replacement of muscular tissue with connective tissue, should also be considered as a cause of functional disturbance of the urinary bladder.

Unusual abdominal tumors with intracardiac extension. Two cases with successful surgical resection

Abdominal tumors that can grow through vascular lumen and spread to the right heart are rare. Although these tumors have different histologic aspects, they may cause similar abdominal and cardiac symptoms and are a serious risk factor for pulmonary embolism and sudden death when they reach the right atrium and tricuspid valve. The best treatment is radical surgical resection of the entire tumor using cardiopulmonary bypass with or without deep hypothermia and total circulatory arrest. We report the cases of two patients, the first with leiomyosarcoma of the inferior vena cava and the other with intravenous leiomyomatosis of the uterus that showed intravascular growth up to right atrium and ventricle, who underwent successful radical resection in a one-stage procedure with the use of cardiopulmonary bypass. We discuss the clinical and histologic aspects and imaging diagnosis and review the literature.

Evaluation and treatment of the overactive bladder

The overactive bladder is characterized by symptoms of frequency, urgency, and urge incontinence, substantially affecting the quality of life of millions of people throughout the world. The symptoms are associated with significant social, psychological, occupational, domestic, physical, and sexual problems. Despite the considerable impact of this condition on quality of life, sufferers are often unwilling to discuss their problem with family members or health care professionals. This situation is unfortunate, for much can be done to alleviate the symptoms of this distressing condition. It is therefore of utmost importance that medical education about symptoms of the overactive bladder and other related problems be improved to help health care professionals identify and treat patients who will benefit from therapy. This article reviews current thinking regarding definition, epidemiology, quality of life effects, evaluation, and management of the overactive bladder.

Adrenocortical tumors: results of treatment and study of Weiss's score as a prognostic factor

PURPOSE: The differential diagnosis between benign and malignant adrenal cortical tumors circumscribed to the gland is controversial. One hundred and seven patients with adrenal cortex tumors (excluding those with primary hyperaldosteronism) were studied to assess the 5-year survival rate of adults, children, patients stratified by pathological stage, and patients stratified according to Weiss's score of <3 or >3. METHODS: The patients were evaluated both clinically and biochemically. One hundred and five patients underwent surgery and were classified pathologically as stages I, II, III, or IV. The tumors were weighed, measured, and classified according to Weiss's criteria and divided into 2 groups: <3 and >3. RESULTS: After 5 years, the survival rate was 77.5% for the whole group, 74.61% for the adults, 84.3% for the children, 100% for stage I, 83.9% for stage II, 33% for stage III, and 11.7% for stage IV groups. Additionally, after 5 years, 100% of the patients with tumors with Weiss's score <3 were alive compared to 61.65% of those with Weiss's score >3. The average weights of the tumors of score <3 and >3 were 23.38 g ± 41.36 g and 376.3 ± 538.76 g, respectively, which is a statistically significant difference. The average sizes of tumors of Weiss's score <3 and >3 were 3.67 ± 2.2 cm and 9.64 ± 5.8 cm, respectively, which is also a statistically significant difference. CONCLUSIONS: Weiss's score may be a good prognostic factor for tumors of the adrenal cortex. Additionally, there was a statistically significant difference between the average weight and size of tumors with benign behavior (Weiss's score <3) and those with malignant behavior (Weiss's score >3).

Salivary gland tumors in a Brazilian population: a retrospective study of 124 cases

Salivary gland tumors constitute a highly heterogeneous histopathologic group. There are few epidemiological studies of large series of benign and malignant salivary gland tumors in Brazil. MATERIAL AND METHODS: Hospital records of 124 patients with salivary gland tumors diagnosed from January 1993 to December 1999 were reviewed. The patients were analyzed according to gender, age, size, location, and histopathology of the tumor. RESULTS AND CONCLUSIONS: Patients with benign and malignant tumors presented with a mean age of 47.7 and 48.8 years, respectively. The frequency of benign tumors was 80% (n = 99) and malignant tumors 20% (n = 25). Tumors were localized in the parotid gland 71% (n = 88), in the submandibular gland 24% (n = 30), and in the minor salivary glands 5% (n = 6). The most common benign tumors were pleomorphic adenoma in 84% (n = 84) and Warthin's tumor in 13% (n = 13). Among malignant tumors, mucoepidermoid carcinoma was the most common in 52% (n = 13), adenoid cystic carcinoma occurred in 20% (n = 5), and carcinoma ex pleomorphic adenoma was detected in 12% (n = 3).

Color Doppler sonography with contrast in the differentiation of ovarian tumors

The objective of this study was to differentiate benign ovarian tumors from malignant ones before surgery using color and pulsed Doppler sonography, and to compare results obtained before and after use of contrast medium, thereby verifying whether contrast results in an improvement in the diagnostic sensitivity. METHODS: Sixty two women (mean age 49.9 years) with ovarian tumors were studied, 45 with benign and 17 with malignant tumors. All women underwent a transvaginal color Doppler ultrasonographic exam. A study of the arterial vascular flow was made in all tumor areas, as well as an impedance evaluation of arterial vascular flow using the resistance index. RESULT: Localization of the vessels in the tumor revealed a greater proportion of malignant tumors with detectable internal vascular flows (64%) than benign tumors with such flows (22%). There was a considerable overlap of these findings. The use of contrast identified a greater number of vessels with confirmation in the totality of tumors, but did not improve the Doppler capacity in tumoral differentiation. Malignant tumors presented lower values of resistance index than the benign ones, whether or not contrast was used. The cutoff value for resistance index that better maximized the Doppler sensitivity and specificity was 0.55. Through this value, an increase of the sensitivity after contrast use was obtained, varying from 47% to 82%, while specificity remained statistically unchanged. CONCLUSION: Although the injection of a microbubble agent improved the sensitivity of the method detecting vascularization of tumors, a positive finding for vascularization by this method was not clinically useful in the differentiation of benign and malignant ovarian tumors.

Evolving hallmarks in urothelial bladder cancer: unveiling potential biomarkers

Urothelial bladder carcinoma (UBC), the most frequent type (90%) of bladder cancer and the second most common malignancy of the urogenital region, is a relatively well understood type of cancer, with numerous studies concerning pathogenetic pathways, natural history and bladder tumor biology being reported. Despite this, it continues to remain a challenge in the oncology field, mostly due to its relapsing and progressive nature, and to the heterogeneity in the response to cisplatin-containing regimens. Although the formulae based on clinical staging and histopathological parameters are classically used as diagnostic and prognostic tools, they have proven insufficient to characterize the individual biological features and clinical behaviour of the tumours. Understanding the pathobiology of the disease can add important information to these classical criteria, and contribute to accurately predict outcome and individualize therapy for UBC patients. In this line of investigation, we found that tumour angiogenesis and lymphangiogenesis, the process of invasion and metastasis and the energy metabolism reprogramming/tumour microenvironment encompass several potential biomarkers that seem to infl bladder cancer aggressiveness and chemoresistance. We particularly highlight the roles of lymphovascular invasion, and of RKIP, CD147 and MCT1 immunoexpressions, as relevant prognostic and/or predictive biomarkers, and as promising areas of therapeutic intervention, eliciting for the development of additional studies that can validate and further explore these biomarkers.