Human adenovirus (HAdV) mainly causes minor illnesses, but can lead to severe disease and death in both immunocompromised and immunocompetent patients. In such cases, the current standards of treatment often do not improve disease outcome and no approved antiviral therapy against HAdV exists. Since HAdV relies on cellular machinery to assist in the progression of the virus lifecycle, we hypothesized that small molecules targeting certain cellular proteins/pathways, without severely affecting cell health, may serve as effective anti-HAdV compounds. Thus, we aimed to identify novel inhibitors of HAdV, and investigate the molecular mechanism to determine new therapeutic targets for intervention in HAdV infection. We first examined the antiviral properties of pan-histone deacetylase (HDAC) inhibitor SAHA and found that the drug affects multiple stages of the HAdV lifecycle, resulting in significant reductions in virus yield. SAHA was effective in decreasing gene expression from clinically relevant HAdV serotypes. Subsequent investigations on the role of HDACs in HAdV infection led us to determine that class I HDAC activity, mainly HDAC2, is necessary for optimal viral gene expression. Using a wildtype-like HAdV reporter construct that allows us to monitor virus replication by fluorescence microscopy, we then designed an efficient system for screening small molecules to identify novel HAdV inhibitors. We screened over 1300 small molecules, and the screen was sensitive enough to detect compounds with both robust and modest antiviral activity. Several positive hits were validated to reduce HAdV gene expression and yield from infected cells. Further investigation on the efficacy of these compounds and the mechanism behind their inhibition of HAdV can lead to the discovery of new pharmacological targets and the development of more effective antivirals.

Bibliography: leaves 306-353. / xiii, 353 leaves : ill. ; 30 cm. / Title page, contents and abstract only. The complete thesis in print form is available from the University Library. / Utilizing a paired transcoronary sampling technique, serial determination of myocardial drug content was determined following intravenous bolus injection in patients undergoing diagnostic cardiac catheterization for the investigation of chest pain. There was significant modulation of haemodynamic effects of all three drugs according to changes in systolic interval. / Thesis (Ph.D.)--University of Adelaide, Dept. of Medicine, Queen Elizabeth Hospital, Cardiology Unit

Diss. (sammanfattning) Stockholm : Karol. inst., 2003. / Härtill 5 uppsatser.

Thesis (Ph.D.)--Medical University of Ohio, 2005. / "In partial fulfillment of the requirements for the degree of Doctor of Philosophy in Medical Sciences." Major advisor: Joseph I. Shapiro. Includes abstract. Document formatted into pages: v, 265 p. Title from title page of PDF document. Bibliography: pages 52-59,94-100,129-134,171-176,200-263.

Submitted by Viviane Lima da Cunha (viviane@biblioteca.ufpb.br) on 2017-03-30T13:07:06Z No. of bitstreams: 1 arquivototal.pdf: 3105346 bytes, checksum: 743a49eb2c327b5bb4a178343286d03e (MD5) / Made available in DSpace on 2017-03-30T13:07:06Z (GMT). No. of bitstreams: 1 arquivototal.pdf: 3105346 bytes, checksum: 743a49eb2c327b5bb4a178343286d03e (MD5) Previous issue date: 2016-06-16 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - CAPES / Ouabain (OUA), a potent Na+/K+ ATPase inhibitor, was identified as an endogenous substance present in human plasma. It was shown that OUA is able to interfere in several aspects of the immune system and inflammation, but the effect of this substance in allergic lung inflammation had not been previously investigated. Lower airways inflammation process is characterized by cell migration and bronchial hyperresponsiveness, as a result of sensitization by protein antigens such as ovalbumin (OVA). Aim: Evaluate the effect of ouabain on the model of allergic pulmonary inflammation induced by OVA. Methods: BALB/c mice (n = 6) were sensitized and challenged with OVA and pretreated intraperitoneally (ip) with OUA (0.56 mg/kg) two days before the sensitization and one hour before them, or dexamethasone (2 mg/kg) subcutaneously 1 hour before to each challenge. Allergic inflammation parameters, such as cell migration, and production of cytokines in bronchoalveolar lavage fluid (BALF), mucus production and pulmonary histopathologic remodeling, and IgE quantitation were analyzed posteriorly. Results: Ouabain reduced (60%) BALF cells as a reflex of polymorphonuclear leukocyte inhibition and T CD3+ lymphocytes, as well as it reduced the production of Th2 profile cytokines, IL-4 and IL-13. Furthermore, ouabain also reduced histopathologic parameters of inflammation, such as hyperplasia of goblet cells and with consequent mucus decrease, besides it reduced OVA-specific IgE titer. Conclusion: These results suggests that OUA presents anti-inflammatory effect on pulmonary allergic inflammation, modulating Th2 phenotype parameters. / A ouabaína (OUA), um potente inibidor da Na+/K+ ATPase, foi identificada como uma substância endógena presente no plasma humano. Nos últimos anos, foi evidenciado que a OUA é capaz de interferir em diversos aspectos do sistema imunológico e em diferentes modelos de inflamação, porém o seu efeito na inflamação alérgica pulmonar não tinha sido investigado previamente. Processos inflamatórios alérgicos das vias aéreas inferiores são caracterizados por migração celular e hiperresponsividade brônquica, em decorrência da sensibilização por antígenos proteicos como a ovalbumina (OVA). Objetivo: Avaliar o efeito da ouabaína no modelo de inflamação alérgica pulmonar induzida por OVA. Métodos: Camundongos BALB/c fêmeas (n = 6) foram sensibilizados e desafiados com OVA e pré-tratados via intraperitoneal (i.p.) com OUA (0,56 mg/kg) dois dias antes das sensibilizações e uma hora antes destas, ou com dexametasona (2 mg/kg) subcutânea 1 hora antes de cada desafio. Parâmetros da inflamação alérgica, como migração celular e produção de citocinas no fluido do lavado broncoalveolar (BALF), produção de muco e remodelamento histopatológico pulmonar quantificação de IgE sérica, foram analisados posteriormente. Resultados: A ouabaína reduziu em 60% o número total de células presentes no BALF como um reflexo da inibição de leucócitos polimorfonucleares e linfócitos T CD3+, bem como a produção de citocinas do fenótipo Th2, IL-4 e IL-13. Ademais, a ouabaína reduziu os parâmetros inflamatórios histopatológicos, como hiperplasia das células caliciformes com consequente diminuição de muco, além de reduzir o título de OVA-IgE específica. Conclusão: Estes dados sugerem que a OUA apresenta efeito anti-inflamatória no modelo de inflamação alérgica pulmonar, modulando parâmetros do fenótipo Th2.

Submitted by Luciana Ferreira (lucgeral@gmail.com) on 2017-02-13T10:28:30Z No. of bitstreams: 2 Tese - Ana Paula Araújo Costa - 2016.pdf: 2728666 bytes, checksum: 2cd43bd911125e81ab1a9483baff4302 (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) / Approved for entry into archive by Luciana Ferreira (lucgeral@gmail.com) on 2017-02-13T10:29:52Z (GMT) No. of bitstreams: 2 Tese - Ana Paula Araújo Costa - 2016.pdf: 2728666 bytes, checksum: 2cd43bd911125e81ab1a9483baff4302 (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) / Made available in DSpace on 2017-02-13T10:29:52Z (GMT). No. of bitstreams: 2 Tese - Ana Paula Araújo Costa - 2016.pdf: 2728666 bytes, checksum: 2cd43bd911125e81ab1a9483baff4302 (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) Previous issue date: 2016-12-16 / Conselho Nacional de Pesquisa e Desenvolvimento Científico e Tecnológico - CNPq / Dilated cardiomyopathy (DCM) is a disease of the heart muscle that culminates in dilatation of the left ventricle, or both, and myocardial contractile dysfunction. The clinical phase of the disease is characterized by congestive heart failure signs (CHF), with or without arrhythmias. The treatment involves the use of drugs aimed at reducing the signs of CHF and arrhythmias, with diuretics, positive inotropic, vasodilator and antiarrhythmic. A new drug candidate (LASSBio 294), capable of promoting combined positive inotropic and vasodilating effects, has recently been developed, and have been tested in pre-clinical study in healthy Beagle dogs with promising results. Therefore, this study proposed to verify the action of the drug prototype LASSBio 294, at a dose of 2mg/Kg on cardiovascular parameters of rabbits with DCM experimentally induced by doxorubicin, using as positive control the pimobendan at a dose of 0.3mg/kg. The DCM was induced by intravenous administration of 1 mg/kg of doxorubicin, at a concentration of 2mg/ml, twice a week, for three weeks, and then weekly until it reached fractional shortening less or equal to 25%. As methods of evaluating the LASSBio 294 action on the cardiovascular system of rabbits and monitor the induction of DMC, the following tests were performed: electrocardiography, echodopplercardiography, measurement of blood pressure, chest radiograph, dosage of cardiac lesions, and kidney and liver function biomarkers and hematologic evaluation. At the end of the induction protocol the animals were randomly divided into two groups A (LASSBio 294) and B (pimobendan) and underwent treatment for 30 days, twice a day. At the end of the study it was concluded that the DCM model induced by doxorubicin is a good model to study the disease and its consequences, leading to systolic and diastolic dysfunction with dilatation of the left ventricle. However the time for induction of DCM is inaccurate and the occurrence of multisystemic toxicity, such as nephrotoxicity and myelosuppression, contributes to high mortality rate in this model (35%). It can be concluded that LASSBio 294 is effective in increasing systolic function, improving diastolic function, without altering rabbits blood pressure, has no pro-arrhythmogenic or toxic effect, and reduced the serum creatinine concentration of the animals, but it does not prevent the evolution of the congestive condition. / A cardiomiopatia dilatada (CMD) é uma enfermidade do músculo cardíaco que culmina em dilatação do ventrículo esquerdo, e/ou direito, e disfunção contrátil do miocárdio, sendo a fase clínica da doença caracterizada por sinais de insuficiência cardíaca congestiva (ICC), com ou sem a presença de arritmias. O tratamento envolve a utilização de fármacos que visem diminuir os sinais de ICC e as arritmias, sendo os diuréticos, inotrópicos positivos, vasodilatadores e antiarrítmicos os mais utilizados. Recentemente foi desenvolvido um novo candidato a fármaco (LASSBio 294) capaz de promover os efeitos vasodilatador e inotrópico positivo combinados, tendo sido testado em estudo pré-clínico em cães saudáveis da raça Beagle, com resultados promissores. Propôs-se neste estudo verificar a ação do protótipo a fármaco LASSBio 294, na dose de 2mg/kg, sobre os parâmetros cardiovasculares de coelhos com CMD experimentalmente induzida por doxorrubicina, utilizando como controle positivo o tratamento com a pimobendana, na dose de 0,3mg/Kg. A CMD foi induzida por meio da administração endovenosa de 1mg/Kg de doxorrubicina, na concentração de 2mg/mL, duas vezes na semana, por três semanas e depois semanalmente até que alcançada fração de encurtamento igual ou inferior a 25%. Como métodos de avaliação da ação do LASSBio 294 sobre o sistema cardiovascular de coelhos e monitoramento da indução da CMD, foram realizados os seguintes exames: eletrocardiografia, ecodopplercardiografia, mensuração da pressão arterial, radiografia torácica, dosagem de biomarcadores de lesão cardíaca, de função renal e hepática e avaliação hematológica. Ao final do protocolo de indução os animais foram distribuídos aleatoriamente em dois grupos, A (LASSBio 294) e B (pimobendana), e foram submetidos a tratamento durante trinta dias, duas vezes ao dia. Ao final do estudo foi possível concluir que o modelo de CMD induzida por doxorrubicina é um bom modelo para estudo da doença e suas consequências, induzindo disfunção sistólica e diastólica do miocárdico, com dilatação do ventrículo esquerdo. Porém, o tempo para indução da CMD é inexato e a ocorrência de toxicidade multissistêmica, como a mielossupressão e nefrotoxicidade, contribui para a elevada taxa de mortalidade dos animais (35%). Ainda, conclui-se que o LASSBio 294 é eficiente em incrementar a função sistólica, melhorar a função diastólica, sem alterar a pressão arterial dos coelhos, não apresentando efeito pró arritmogênico ou tóxico, e reduzindo a concentração sérica de creatinina dos animais, porém não impede a evolução do quadro congestivo.

Introduction: Hypertension is an important public health challenge worldwide, being the leading cause of cardiovascular disease, morbidity and mortality. It is particularly prevalent in people in sub-Saharan Africa, especially in urban areas. There is an urgent need to develop strategies to prevent, detect, treat, and control hypertension effectively in the African region. Helicobacter pylori, a gram-negative bacterium responsible for many gastric disorders worldwide, has been associated with hypertension in some previous studies; where blood pressure of patients with Helicobacter pylori infection did not subside after hypertensive treatment, when compared to patients without Helicobacter pylori infections. This effect was suggested to be due to Helicobacter pylori produced and modified cardiotonic steroids that are found in elevated concentrations in hypertensive patients. Cardiotonic steroids are positive inotropic agents which are known to increase blood pressure. A sensitive analytical method is needed to detect and quantify the low concentrations of cardiotonic steroids in biological samples. Materials and Methods: An extraction method was optimised using reversed phase Solid Phase Extraction. A targeted liquid chromatography tandem mass spectrometry method using an Agilent binary series 1100/1200 LC system with a Kinetex C18 RP column (100 x 2.1 mm, 2.6 µm) coupled to a Sciex 4000QTRAP tandem mass spectrometer was developed and validated for the detection and quantitation of 9 different cardiotonic steroids in both solvent and whole blood. The method was validated according to the International Conference on Harmonization guidelines with regards to precision, accuracy, sensitivity, selectivity, linearity, range, limit of detection, limit of quantification, reproducibility, recovery, carry-over and stability. Media from Helicobacter pylori cultures and faecal samples from human and different normo- and hypertensive rat strains were analysed. Data analysis was performed with Analyst® Software (version 1.5.2) and multiple t-test and Kruskal Wallis test using GraphPad Prism 8 software. Results and Discussion: The calibration curves of tested cardiotonic steroids were linear over a concentration range of 0.1-40 ng/mL with coefficients of determination greater than 0.990 except for telocinobufagin. The analytical method was selective with an estimated limit of detection and limit of quantification between 0.02-0.5 ng/mL and 0.1-2 ng/mL respectively. All tested cardiotonic steroids showed good recovery of over 70%. Accuracy and precision were found to be within acceptable limits of 15% and 20% at lowest limit of quantification for almost all the analytes and their stability in blood and solvent at room temperature, 4°C, -20°C and -80°C was tested for a month. Cardiotonic steroids were detected in Helicobacter pylori cultures and faecal samples with the exception of ouabain and proscillaridin A which were not detected at all. Although Helicobacter pylori were shown to produce cardiotonic steroids in vitro, no evidence of the effect of Helicobacter pylori on cardiotonic steroids production was detected in different normo- and hypertensive rat groups. Conclusion: The quantitative analytical method was successfully validated, over expected in vivo concentration ranges for 8 different cardiotonic steroids. The extraction and analytical methods were both successfully applied to Helicobacter pylori cultures and faecal rat samples where cardiotonic steroids were detected. / Dissertation (MSc)--University of Pretoria 2020. / National Research Foundation Student bursary / Pharmacology / MSc (Pharmacology) / Unrestricted

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