Navegando por Palavras-chave "Descoberta De Drogas"

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    Caracterização funcional de um composto capaz de regular o desenvolvimento celular de Trypanosoma cruzi
    (Universidade Federal de São Paulo (UNIFESP), 2019-03-28) Araujo Junior, Adalberto Miguel De [UNIFESP]; Freitas Junior, Lucio Holanda Gondim De [UNIFESP]; http://lattes.cnpq.br/2136191319465692; http://lattes.cnpq.br/4897591409094074; Universidade Federal de São Paulo (UNIFESP)
    Chagas disease, caused by the kinetoplastid parasite Trypanosoma cruzi, remains one of the most neglected tropical illnesses in the world. Approximately 6 to 7 million people worldwide are estimated to be infected with T. cruzi. Benznidazole (BNZ) and nifurtimox (NFX), the only drugs available to treat the disease, are associated with severe side effects and unclear efficacy on the chronic phase of infection. The pathogenesis related to such disease is still poorly understood, which reinforces the importance of researching more about the basic biology of T. cruzi infection, replication and differentiation, once only this way new strategies of chemotherapies will be discovered. In the absence of clinically validated molecular drug targets, development of novel phenotypic screening assays may result in the discovery of compounds that can inducephenotypes related to distinct mechanisms of action not investigated so far. In this report, we attempted to discuss how relevant the phenotypic screening approach can be for the understanding of T. cruzi cell biology. From the high content screening of a library of 4,000 putative kinase inhibitors, we selected some compounds of interest, which showed low cytotoxicity and capacity to induce specific phenotypes of interference in the normal intracellular development of the parasite. One of such compounds (named HF) inhibited the parasite replication without causing loss of cell viability. The functional characterization of the HF activity showed that: (i) the phenotype of intracellular development arrest was not restricted to a host cell type; (ii) the compound was able to inhibit the replication of different T. cruzi strains (Y, Sylvio X10/1 e CL Brener strain); (iii) the phenotype of arrest was reversible upon compound removal, once the parasite from that point grows normally concluding its intracellular cycle by differentiation to trypomastigotes; (iv) the compound did not show cytotoxicity for distinct host cells (U2OS, LLC-MK2, NRK-52E and BHK-51) for up to the maximum concentration of 100 µM for 48 h, being highly selective for the parasite; (v) the molecule also was able to induce arrest in the epimastigote form of T. cruzi without causing loss of viability up to 96 h, but displayed a dose-dependent cidal activity against bloodstream forms of Trypanosoma brucei and promastigote forms of Leishmania donovani; and (vi) the compound caused T. cruzi cell cycle arrest, being the most of the cell population at G1, unlike what was observed in cells treated with benznidazole. Altogether, these results suggest that this compound acts via a regulator of cell cycle that can specifically cause arrest in T. cruzi and not in mammalian cells. Regarding the other trypanosomatids, there may be a regulation of the cell cycle by the compound, however we did not observe the same phenotypic action as seen in T. cruzi. Moreover, this work has shown us the potential of HF to be applied as a chemical probe in investigations regarding T. cruzi replication and development.
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    Descoberta de novos antivirais para Flavivírus a partir de uma abordagem fenotípica
    (Universidade Federal de São Paulo (UNIFESP), 2019-05-30) Pilger, Denise Regina Bairros De [UNIFESP]; Freitas Junior, Lucio Holanda Gondim De [UNIFESP]; http://lattes.cnpq.br/2136191319465692; http://lattes.cnpq.br/1034340400100837; http://lattes.cnpq.br/1034340400100837; Universidade Federal de São Paulo (UNIFESP)
    The Flavivirus genus comprises more than 70 pathogenic viruses with different pathologies and a significant public health impact in different regions of the world, with potential to emerge in non-endemic regions. Dengue virus (DENV), yellow fever (YFV) and Zika (ZIKV) are some of the most important human pathogenic flaviviruses with similar symptoms ranging from mild fever and malaise to hemorrhage, shock and failure of multiple organs. Despite the existence of protective vaccines for both viruses, their use is contraindicated for immunocompromised individuals, pregnant women and children. The only treatment is the mitigation of symptoms associated with diseases, with analgesics, antipyretics and replacement of body fluid. Specific antiviral chemotherapies could reduce the morbidity and mortality associated with these infections. In addition, during an outbreak, a specific drug could be administered at the onset of any symptoms associated with infections. Therefore, there is an urgent need to establish a more objective, sensitive and reproducible biological assay to search new antiviral therapies. Thus, an image-based phenotypic assay was developed and optimized for the screening of three commercial compound libraries (NIH Clinical Collection, National Institute of Health, LOPAC1280, Sigma, and Pathogen Box, MMV) against a panel of viruses of the genus Flavivirus. We highlight the results obtained through: (i) NIH library, from which we identified two compounds against dengue virus serotype 2 with high selectivity (> 10); (ii) LOPAC1280, from which we identified three compounds with promising activity against the yellow fever virus. In addition, the activity spectrum of these compounds was evaluated against other viruses of the same genus, as well as their ability to induce resistance in yellow fever virus, and how they interact with the viral life cycle. (iii) Pathogen Box library was screened in parallel with six different viruses to stablish an activity profile of the compounds. As a result, 133 compounds were selected, some of which have already been described as having antiviral activity, validating the assay as a viable tool for the discovery of candidates with potential antiviral activity. In summary, the methodology described in this work represents an important contribution to the initial processes of antiviral development. With these experiments it was possible to find new potent and effective compounds for YFV, which provides a solid contribution to the early stages of the discovery of new drugs for diseases caused by flavivirus.
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    Nova estratégia para descoberta de anti-helmínticos utilizando o organismo modelo Caenorhabditis elegans
    (Universidade Federal de São Paulo (UNIFESP), 2019-03-28) Cintra, Giovana Aparecida De Souza [UNIFESP]; Freitas Junior, Lucio Holanda Gondim De [UNIFESP]; Moraes, Carolina Borsoi [UNIFESP]; http://lattes.cnpq.br/2856012942943418; http://lattes.cnpq.br/2136191319465692; http://lattes.cnpq.br/1967524610258147; Universidade Federal de São Paulo (UNIFESP)
    Diseases caused by soil-transmitted helminths are among the most common infections in the world, affecting marginalized, low-income and resource-poor populations. It is estimated that more than one billion people, especially in developing countries in Africa, Asia and America, are infected with one or more species of helminths. The emergence of resistance to the main anthelmintics used highlights the need for the introduction of new therapeutic agents. The free-living nematode Caenorhabditis elegans is widely recognized as a model organism for investigating the biology of parasitic nematodes due to its easy laboratory culture, short generation time and fully sequenced genome. Most commercial anthelmintics are active against C. elegans, levamisole and benzimidazole derivatives, and this animal model has been useful in understanding the mechanism of action of the same, making it a useful system for selection and investigation of compounds with potential anthelmintic activity. The methodologies currently used to identify new compounds are based on the subjective evaluation of the motile worms, or involve transgenic organisms. In this way there is a need for the establishment of a more objective, sensitive and reproducible biological assay. In the present study the development and standardization of a new methodology of in vivo high content screening using the C. elegans is described. The assay is based on viability assessment by differential incorporation of two fluorescent molecular markers, DB-1, derived from the 2,1,3- benzothiadiazole core and propidium iodide. This model stands out from the previous tests for using an automated imaging platform and later analysis of the images by commercial software, thus quantitatively evaluating the viability in an objective way. In addition, it allows the use of animals without genetic modifications. The establishment of a robust and reproducible assay allowed the screening of 2280 compounds, which are part of three libraries: (i) the commercial LOPAC library, from which three compounds with a potency up to 400x greater than levamisole hydrochloride were selected; (ii) the library of compounds / extracts from different collaborators, with new therapeutic and chemical classes; (iii) the Pathogen Box library, from which three previously reported compounds with activity against malaria, schistosomiasis and tuberculosis were identified. The candidates identified in this work are starting point for studies of medicinal chemistry, aiming to improve their potency and effectiveness. It is also worth noting that two of the compounds selected in the collaborator screening reduced the parasitemia in the in vivo model of Ascarissum infection by 61.96% and 59.78%, confirming the translational value of the C. elegans biological model. In conclusion, the assay developed in this work was successfully established and provides a solid contribution to the initial phases of the discovery of new anthelminthic drugs.