Análise comparativa de duas traduções de Tropic of Cancer de Henry Miller

Orientadora: Maria Helena Anacleto-Matias

Synthesis, characterization and antiproliferative activity on cancer cell lines of new ionic liquids from ampicillin

Ionic Liquids (ILs) are ionic compounds that possess melting temperature below 100ºC and they have been a topic of great interest since the mid-1990s due to their unique properties. The range of IL uses has been broadened, due to a significant increase in the variety of physical, chemical and biological ILs properties. They are now used as Active Pharmaceutical Ingredients (APIs) and recent interests are focused on their application as innovative solutions in new medical treatment and delivery options.1 In this work, our principal objective was the synthesis and investigation of physicochemical and medical properties of ionic liquids (ILs) and organic salts from ampicillin. This approach is of huge interest in pharmaceutical industry as cation and anion composition of ILs and organic salts can greatly alter their desired properties, namely the melting temperature and even synergistic effects can be obtained.2,3 For the synthesis of these compounds we used a recently developed method proposed by Ohno et al.4 for the preparation of quaternary ammonium and phosphonium hydroxides, that were neutralized by ampicillin. After purification we obtained pure ILs and salts in good yields. These ILs shows good antimicrobial and antifungal activities. As it is well known that some ionic liquids containing phosphonium and ammonium cation also shows anti-cancer activity1,5 we also decided to study these compounds against some cancer cell lines.

Relatório de estágio: intervenção em fisioterapia na sindrome da cirurgia lombar falhada

Introdução: A síndrome da cirurgia lombar falhada (SCLF) caracteriza-se pela persistência ou recidiva da dor ou aparecimento de novos sintomas após discectomia, descompressão de canal estreito ou artrodese lombares. Objectivos: O objectivo deste estudo foi descrever a abordagem em fisioterapia de um caso com Síndrome da Cirurgia Lombar Falhada e avaliar os seus efeitos na funcionalidade e sintomatologia dolorosa neste indivíduo. Metodologia: A amostra foi constituída por um sujeito com 41 anos, trabalhador da construção civil, apresentando dor lombar com irradiação para o membro inferior, após ter sido submetido a discectomia lombar e artrodese, que realizou sessões de fisioterapia convencional antes e depois da cirurgia, e que apresentava sinais compatíveis com compressão radicular. A intensidade da dor foi medida através da escala visual analógica (EVA), as amplitudes com o goniómetro universal, a funcionalidade foi avaliada utilizando o Questionário de Incapacidade Roland-Morris (RMDQ), o estado psicológico foi avaliado através da “Escala de Desânimo Associado à Dor” e do “Inventário de Convicções e Percepções Relacionadas com a Dor”. Resultados: A dor era inicialmente em média de 5,5/10 EVA, sendo de 3,4/10 EVA no final do tratamento. Quanto à funcionalidade (RMDQ) esta variou de 14/24 no início para 17/24 no final. De uma forma geral, após a aplicação de técnicas para diminuir a tensão do piriforme, técnicas neuurodinâmicas e um programa de exercícios para melhoria do controlo motor, o paciente apresentou alívio das queixas de dor na nádega esquerda e anca, aumento da amplitude de rotação medial da coxo-femoral, ausência das parestesias na face plantar do pé esquerdo e melhoria das queixas de dor na face anterior e lateral da coxa. Verificamos que houve diminuição dos índices de dor e melhorias dos níveis de funcionalidade. Conclusão: Neste estudo de caso, foi salientado o processo de raciocínio clínico desenvolvido pelo fisioterapeuta: a interpretação dos dados da história e levantamento das primeiras hipóteses e a realização dos testes no exame objectivo, permitiram estabelecer um diagnóstico funcional e elaborar um plano de intervenção através do qual o utente recuperou parcialmente a sua funcionalidade e diminuiu os seus índices de dor.

Influência da cirurgia abdominal na função pulmonar e capacidade de tosse

Introdução: A cirurgia abdominal acarreta grande risco de complicações pulmonares pós-operatórias. As alterações pós-cirúrgicas abdominais, reflectem-se na dinâmica ventilatória, de modo particular nos volumes e capacidades pulmonares, e na capacidade de tosse. Objectivos: Compreender qual a variação dos volumes e capacidades pulmonares e da capacidade de tosse antes e depois da cirurgia abdominal (estômago e cólon), e qual a correlação dessa variação com o nível de dor percepcionada. Desenho do estudo: Unicêntrico, prospectivo e observacional. Amostra: 10 indivíduos, propostos para cirurgia abdominal – estômago e cólon. Metodologia: Dois momentos de avaliação: um nas 24h préoperatórias em que se mediu a capacidade vital forçada (CVF) e o volume expirado máximo no primeiro segundo (VEMS1) com espirometria, e do pico de fluxo de tosse (PCF); e um segundo momento nas 24h pós-operatórias onde se repetiram as medições do primeiro momento com o acréscimo da avaliação da dor. Resultados: No pós-operatorio imediato há uma diminuição significativa da CVF de 44,30%±17,24 (p=0,005), do VEMS1 de 35,50%±28,47 (p=0,009) e do PCF de 38,97%±38,66 (p=0,012). Não se verificou nenhuma relação entre a dor percepcionada na realização das manobras de espirometria e tosse com diminuição a da CVF e do VEMS1 e do PCF respectivamente. O sexo apresentou uma relação significativa com a variação da CRF e do VEMS1 (p=0,046 e p=0,008 respectivamente). A frequência respiratória apresentou um aumento significativo no pós-operatório de 10±11,22 cpm (p=0,019). A saturação periférica de oxigénio apresentou uma diminuição significativa no pós-operatório de 3,52±2,47 (p=0,011) Conclusão: No estudo efectuado fica demonstrado o impacto negativo da cirurgia abdominal na dinâmica respiratória. A diminuição dos valores da CVF, do VEMS1 e do PCF podem contribuir de forma significativa para o aumento do risco de complicações respiratória pós-operatórias. No entanto seria importante a realização deste estudo com uma amostra maior.

A experiência da unidade autónoma de gestão de cirurgia do Centro Hospitalar de São João E.P.E modelos de gestão intermédia hospitalar

As organizações de saúde são muito particulares devido à sua missão, aos recursos que mobilizam, aos processos que dinamizam, à produção que realizam e ainda à envolvente externa onde se inserem (Reis, 2007). Os sucessivos esforços que têm sido utilizados na reforma na saúde, sobretudo a partir de 1988, têm sido uma constante da agenda política na tentativa de aumentar a eficiência dos serviços prestados, a efetividade dos resultados e a responsabilidade dos profissionais. A empresarialização do Hospital de São João operada a partir de 2006, com a publicação do Dec.Lei 233/05 de 29 de Dezembro, tornou como imperativo estratégico a alteração profunda do modelo de gestão até então praticado. Este era caracterizado por uma forte componente administrativa, de cariz burocrática, e sob ponto de vista económico assentava em sucessivos deficits e no permanente aumento e descontrolo da despesa. Tomando como pressuposto que a única via de modificar esse padrão passava entre outras medidas pela efetivação de uma gestão descentralizada, vieram a ser criadas seis estruturas intermédias de gestão designadas por “Unidades Autónomas de Gestão”. Estas tinham como objetivo aumentar o valor em saúde, melhorar a gestão dos serviços clínicos, potenciando desse modo a qualidade e efetividade dos cuidados prestados, bem como a eficiência dos recursos utilizados. Neste sentido, o propósito deste trabalho centra-se em demonstrar que a implementação de um modelo de gestão descentralizado como é o caso da Unidade Autónoma de Gestão de Cirurgia, doravante designada por UAGC, constituiu uma opção gestionária eficaz e altamente promissora na governação clínica, desmistificando o mito da “ingovernabilidade dos hospitais centrais” como era apanágio do Hospital S. João. Cremos que a descentralização da gestão enquanto forma de reengenharia da organização interna dos hospitais constitui um importante instrumento no sentido de orientar e motivar o comportamento dos gestores (sejam eles clínicos ou não) para o cumprimento dos objetivos institucionais, através da implementação de políticas de desconcentração de poderes, competências e responsabilidades. Embora existam outros modelos de organização ao nível da gestão intermédia, na verdade, a implementação destas estruturas descentralizadas traduziu-se numa inegável mais valia organizativa e gestionária do CHSJ. como os indicadores de desempenho mais à frente tentarão demonstrar. Temos consciência que este modelo está longe de ser perfeito, e que por vezes não é corretamente entendido pelos profissionais, que o encaram como uma necessidade de cariz exclusivamente económica. Porém o caminho já percorrido pela UAGC ao longo destes 5 anos permite-nos afirmar que é possível “fazer mais” com “os mesmos recursos”, desde que exista uma clara estratégia de ação suportada em programas concretos e exequíveis, praticados num clima social participado e responsabilizante.

Molecular imprinted nanoelectrodes for ultra sensitive detection of ovarian cancer marker

The relentless discovery of cancer biomarkers demands improved methods for their detection. In this work, we developed protein imprinted polymer on three-dimensional gold nanoelectrode ensemble (GNEE) to detect epithelial ovarian cancer antigen-125 (CA 125), a protein biomarker associated with ovarian cancer. CA 125 is the standard tumor marker used to follow women during or after treatment for epithelial ovarian cancer. The template protein CA 125 was initially incorporated into the thin-film coating and, upon extraction of protein from the accessible surfaces on the thin film, imprints for CA 125 were formed. The fabrication and analysis of the CA 125 imprinted GNEE was done by using cyclic voltammetry (CV), differential pulse voltammetry (DPV) and electrochemical impedance spectroscopy (EIS) techniques. The surfaces of the very thin, protein imprinted sites on GNEE are utilized for immunospecific capture of CA 125 molecules, and the mass of bound on the electrode surface can be detected as a reduction in the faradic current from the redox marker. Under optimal conditions, the developed sensor showed good increments at the studied concentration range of 0.5–400 U mL−1. The lowest detection limit was found to be 0.5 U mL−1. Spiked human blood serum and unknown real serum samples were analyzed. The presence of non-specific proteins in the serum did not significantly affect the sensitivity of our assay. Molecular imprinting using synthetic polymers and nanomaterials provides an alternative approach to the trace detection of biomarker proteins.

Ultrasensitive detection of ovarian cancer marker using immunoliposomes and gold nanoelectrodes

Mucin-16 (MUC16) is the established ovarian cancer marker used to follow the disease during or after treatment for epithelial ovarian cancer. The emerging science of cancer markers also demands for the new sensitive detection methods. In this work, we have developed an electrochemical immunosensor for antigen MUC16 using gold nanoelectrode ensemble (GNEE) and ferrocene carboxylic acid encapsulated liposomes tethered with monoclonal anti-Mucin-16 antibodies ( MUC16). GNEEs were fabricated by electroless deposition of the gold within the pores of polycarbonate track-etched membranes. Afterwards, MUC16 were immobilized on preformed self-assembled monolayer of cysteamine on the GNEE via cross-linking with EDC-Sulfo-NHS. A sandwich immunoassay was performed on MUC16 functionalized GNEE with MUC16 and immunoliposomes. The differential pulse voltammetry was employed to quantify the faradic redox response of ferrocene carboxylic acid released from immunoliposomes. The dose–response curve for MUC16 concentration was found between the range of 0.001–300 U mL−1. The lowest detection limit was found to be 5 × 10−4 U mL−1 (S/N = 3). We evaluated the performance of this developed immunosensor with commercial ELISA assay by comparing results obtained from spiked serum samples and real blood serum samples from volunteers.

FASL polymorphism is associated with response to bacillus Calmette-Guérin immunotherapy in bladder cancer

Objective Deregulation of FAS/FASL system may lead to immune escape and influence bacillus Calmette-Guérin (BCG) immunotherapy outcome, which is currently the gold standard adjuvant treatment for high-risk non–muscle invasive bladder tumors. Among other events, functional promoter polymorphisms of FAS and FASL genes may alter their transcriptional activity. Therefore, we aim to evaluate the role of FAS and FASL polymorphisms in the context of BCG therapy, envisaging the validation of these biomarkers to predict response. Patients and methods DNA extracted from peripheral blood from 125 patients with bladder cancer treated with BCG therapy was analyzed by Polymerase Chain Reaction—Restriction Fragment Length Polymorphism for FAS-670 A/G and FASL-844 T/C polymorphisms. FASL mRNA expression was analyzed by real-time Polymerase Chain Reaction. Results Carriers of FASL-844 CC genotype present a decreased recurrence-free survival after BCG treatment when compared with FASL-844 T allele carriers (mean 71.5 vs. 97.8 months, P = 0.030) and have an increased risk of BCG treatment failure (Hazard Ratio = 1.922; 95% Confidence Interval: [1.064–3.471]; P = 0.030). Multivariate analysis shows that FASL-844 T/C and therapeutics scheme are independent predictive markers of recurrence after treatment. The evaluation of FASL gene mRNA levels demonstrated that patients carrying FASL-844 CC genotype had higher FASL expression in bladder tumors (P = 0.0027). Higher FASL levels were also associated with an increased risk of recurrence after BCG treatment (Hazard Ratio = 2.833; 95% Confidence Interval: [1.012–7.929]; P = 0.047). FAS-670 A/G polymorphism analysis did not reveal any association with BCG therapy outcome. Conclusions Our results suggest that analysis of FASL-844 T/C, but not FAS-670 A/G polymorphisms, may be used as a predictive marker of response to BCG immunotherapy.

Cancro da mama: avaliação do gânglio sentinela em exame extemporâneo

A maximização do potencial da abordagem cirúrgica conservadora da axila para o cancro da mama, como um método minimamente invasivo para a avaliação de metastização axilar, visando diminuir a morbilidade associada ao esvaziamento axilar completo, requer um método preciso para avaliação patológica intraoperatória. Esse método não foi ainda estabelecido. Imprints e corte de congelação do gânglio sentinela são os procedimentos comummente utilizados, apesar de uma sensibilidade e especificidade inferior à desejada. Actualmente, novas técnicas estão a ser desenvolvidas, que apesar da sua optimização, ainda não ultrapassam os resultados das utilizadas. Um total de 138 mulheres com cancro da mama, submetidas a cirurgia mamária por abordagem conservadora da axila, cuja utilização de imprints e cortes de congelação foram os métodos de avaliação intraoperatória do gânglio sentinela. Os diagnósticos dados pela observação dos cortes dos fragmentos do exame extemporâneo foram comparados com os obtidos nos cortes histológicos definitivos dos fragmentos fixados em formol e incluídos em parafina. Os resultados obtidos da avaliação do exame extemporâneo demonstraram sensibilidade de 79,1%, especificidade de 96,9%, com uma precisão de 91,4%. Não se obteve correlação entre os resultados do extemporâneo e os parâmetros de caracterização do exame extemporâneo e tumores. Os métodos, imprints e cortes de congelação, actualmente utilizados na nossa instituição apresentam bons resultados, mas a adopção de apenas um ou outro necessita de uma análise mais aprofundada dos dados relativos à metodologia utilizada nos extemporâneos, de forma a verificar a especificidade e sensibilidade individualizadas dos imprints e dos cortes de congelação. Se os imprints se revelarem semelhantes aos cortes de congelação, a sua utilização é preferencial uma vez que acarreta menores custos e são menos morosos. A implementação de novas técnicas será uma metodologia adoptar, pelos benefícios acrescidos, porém mais estudos e a optimização

Macrophages as biomarkers in BCG treatment response in bladder cancer

Bladder cancer is a common urologic cancer and the majority has origin in the urothelium. Patients with intermediate and high risk of recurrence/progression bladder cancer are treated with intravesical instillation with Bacillus Calmette-Guérin, however, approximately 30% of patients do not respond to treatment. At the moment, there are no accepted biomarkers do predict treatment outcome and an early identification of patients better served by alternative therapeutics. The treatment initiates a cascade of cytokines responsible by recruiting macrophages to the tumor site that have been shown to influence treatment outcome. Effective BCG therapy needs precise activation of the Th1 immune pathway associated with M1 polarized macrophages. However, tumor-associated macrophages (TAMs) often assume an immunoregulatory M2 phenotype, either immunosuppressive or angiogenic, that interfere in different ways with the BCG induced antitumor immune response. The M2 macrophage is influenced by different microenvironments in the stroma and the tumor. In particular, the degree of hypoxia in the tumors is responsible by the recruitment and differentiation of macrophages into the M2 angiogenic phenotype, suggested to be associated with the response to treatment. Nevertheless, neither the macrophage phenotypes present nor the influence of localization and hypoxia have been addressed in previous studies. Therefore, this work devoted to study the influence of TAMs, in particular of the M2 phenotype taking into account their localization (stroma or tumor) and the degree of hypoxia in the tumor (low or high) in BCG treatment outcome. The study included 99 bladder cancer patients treated with BCG. Tumors resected prior to treatment were evaluated using immunohistochemistry for CD68 and CD163 antigens, which identify a lineage macrophage marker and a M2-polarized specific cell surface receptor, respectively. Tumor hypoxia was evaluated based on HIF-1α expression. As a main finding it was observed that a high predominance of CD163+ macrophage counts in the stroma of tumors under low hypoxia was associated with BCG immunotherapy failure, possibly due to its immunosuppressive phenotype. This study further reinforces the importance the tumor microenvironment in the modulation of BCG responses.

Expanding our therapeutic options: β-blockers for colon cancer?

Supported by U. Porto/Santander Totta (IJUP) (PP-IJUP2011-320)

Enoxacin inhibits growth of prostate cancer cells and effectively restores microRNA processing

Prostate cancer (PCa) is one of the most incident malignancies worldwide. Although efficient therapy is available for early-stage PCa, treatment of advanced disease is mainly ineffective and remains a clinical challenge. microRNA (miRNA) dysregulation is associated with PCa development and progression. In fact, several studies have reported a widespread downregulation of miRNAs in PCa, which highlights the importance of studying compounds capable of restoring the global miRNA expression. The main aim of this study was to define the usefulness of enoxacin as an anti-tumoral agent in PCa, due to its ability to induce miRNA biogenesis in a TRBP-mediated manner. Using a panel of five PCa cell lines, we observed that all of them were wild type for the TARBP2 gene and expressed TRBP protein. Furthermore, primary prostate carcinomas displayed normal levels of TRBP protein. Remarkably, enoxacin was able to decrease cell viability, induce apoptosis, cause cell cycle arrest, and inhibit the invasiveness of cell lines. Enoxacin was also effective in restoring the global expression of miRNAs. This study is the first to show that PCa cells are highly responsive to the anti-tumoral effects of enoxacin. Therefore, enoxacin constitutes a promising therapeutic agent for PCa.

Anti-tumoral effect of the non-nucleoside DNMT inhibitor RG108 in human prostate cancer cells

Background: Current therapeutic strategies for advanced prostate cancer (PCa) are largely ineffective. Because aberrant DNA methylation associated with inappropriate gene-silencing is a common feature of PCa, DNA methylation inhibitors might constitute an alternative therapy. In this study we aimed to evaluate the anti-cancer properties of RG108, a novel non-nucleoside inhibitor of DNA methyltransferases (DNMT), in PCa cell lines. Methods: The anti-tumoral impact of RG108 in LNCaP, 22Rv1, DU145 and PC-3 cell lines was assessed through standard cell viability, apoptosis and cell cycle assays. Likewise, DNMT activity, DNMT1 expression and global levels of DNA methylation were evaluated in the same cell lines. The effectiveness of DNA demethylation was further assessed through the determination of promoter methylation and transcript levels of GSTP1, APC and RAR-β2, by quantitative methylation-specific PCR and RT-PCR, respectively. Results: RG108 led to a significant dose and time dependent growth inhibition and apoptosis induction in LNCaP, 22Rv1 and DU145. LNCaP and 22Rv1 also displayed decreased DNMT activity, DNMT1 expression and global DNA methylation. Interestingly, chronic treatment with RG108 significantly decreased GSTP1, APC and RAR-β2 promoter hypermethylation levels, although mRNA re-expression was only attained GSTP1 and APC. Conclusions: RG108 is an effective tumor growth suppressor in most PCa cell lines tested. This effect is likely mediated by reversion of aberrant DNA methylation affecting cancer related-genes epigenetically silenced in PCa. However, additional mechanism might underlie the anti-tumor effects of RG108. In vivo studies are now mandatory to confirm these promising results and evaluate the potential of this compound for PCa therapy.

Regulation of histone H2A.Z expression is mediated by sirtuin 1 in prostate cancer

Histone variants seem to play a major role in gene expression regulation. In prostate cancer, H2A.Z and its acetylated form are implicated in oncogenes’ upregulation. SIRT1, which may act either as tumor suppressor or oncogene, reduces H2A.Z levels in cardiomyocytes, via proteasome-mediated degradation, and this mechanism might be impaired in prostate cancer cells due to sirtuin 1 downregulation. Thus, we aimed to characterize the mechanisms underlying H2A.Z and SIRT1 deregulation in prostate carcinogenesis and how they interact. We found that H2AFZ and SIRT1 were up- and downregulated, respectively, at transcript level in primary prostate cancer and high-grade prostatic intraepithelial neoplasia compared to normal prostatic tissues. Induced SIRT1 overexpression in prostate cancer cell lines resulted in almost complete absence of H2A.Z. Inhibition of mTOR had a modest effect on H2A.Z levels, but proteasome inhibition prevented the marked reduction of H2A.Z due to sirtuin 1 overexpression. Prostate cancer cells exposed to epigenetic modifying drugs trichostatin A, alone or combined with 5-aza-2’-deoxycytidine, increased H2AFZ transcript, although with a concomitant decrease in protein levels. Conversely, SIRT1 transcript and protein levels increased after exposure. ChIP revealed an increase of activation marks within the TSS region for both genes. Remarkably, inhibition of sirtuin 1 with nicotinamide, increased H2A.Z levels, whereas activation of sirtuin 1 by resveratrol led to an abrupt decrease in H2A.Z. Finally, protein-ligation assay showed that exposure to epigenetic modifying drugs fostered the interaction between sirtuin 1 and H2A.Z. We concluded that sirtuin 1 and H2A.Z deregulation in prostate cancer are reciprocally related. Epigenetic mechanisms, mostly histone post-translational modifications, are likely involved and impair sirtuin 1-mediated downregulation of H2A.Z via proteasome-mediated degradation. Epigenetic modifying drugs in conjunction with enzymatic modulators are able to restore the normal functions of sirtuin 1 and might constitute relevant tools for targeted therapy of prostate cancer patients

Role of SPARC in Bone Remodeling and Cancer‐Related Bone Metastasis

There is a growing socioeconomic recognition that clinical bone diseases such as bone infections, bone tumors and osteoporotic bone loss mainly associated with ageing, are major issues in today0s society. SPARC (secreted protein, acidic and rich in cysteine), a matricellular glycoprotein, may be a promising therapeutic target for preventing or treating bone‐related diseases. In fact, SPARC is associated with tissue remodeling, repair, development, cell turnover, bone mineralization and may also participate in growth and progression of tumors, namely cancer‐related bone metastasis. Yet, the function of SPARC in such biological processes is poorly understood and controversial. The main objective of this work is to review the current knowledge related to the activity of SPARC in bone remodeling, tumorigenesis, and bone metastasis. Progress in understanding SPARC biology may provide novel strategies for bone regeneration and the development of anti‐angiogenic, anti‐proliferative, or counter‐adhesive treatments specifically against bone metastasis.