Дисертації з теми "Drug screening model"
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Psaroudakis, G. "Virtual screening in drug design and model evaluation." Thesis, University of Essex, 2005. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.422234.
Повний текст джерелаWu, Yuelong Ph D. Massachusetts Institute of Technology. "A high-throughput antiepileptic drug screening system based on chemically Induced zebrafish behavioral model." Thesis, Massachusetts Institute of Technology, 2014. http://hdl.handle.net/1721.1/93816.
Повний текст джерелаCataloged from PDF version of thesis.
Includes bibliographical references (pages 53-59).
Epilepsy, which has the largest worldwide impacts among all nervous system diseases expect for stroke and dementia, is a group of long-term neurological disorders characterized by epileptic seizures. AED medications are the mainstay for epileptic seizure management. However, the existing AEDs cannot fit the needs for every patient due to the efficacy and side effect issues. In this thesis, a high-throughput system to screen new antiepileptic drug is built up. Chemically induced zebrafish larvae are used as a seizure model. The change in fishes' behavior patterns serves as an indicator of the fishes' nervous system condition. The design of the behavior data acquisition setup as well as the requirements of its components is described. A fish tracking program that tracks the locomotion variables like the head position, the tail movement and sideway orientation etc. is developed. The tracking results are treated either by simply computing the statistics of the tracking variables or implementing behavior pattern classifications. Two test datasets involving two different convulsants and one known AED are acquired and analyzed. The results coincide with the existing knowledge about the chemicals' effects on the human nerve system, which suggests the system described in this thesis is promising to help with the actual AED development.
by Yuelong Wu.
S.M.
Wu, Calvin. "In Vitro Cortical Networks for Disease Modeling and Drug Evaluation." Thesis, University of North Texas, 2013. https://digital.library.unt.edu/ark:/67531/metadc407860/.
Повний текст джерелаAldhumani, Ali Hamed. "Pharmacophore Model Development: Targeting Noncoding RNA for Antibacterial/Antiviral Drug Discovery." Ohio University / OhioLINK, 2021. http://rave.ohiolink.edu/etdc/view?acc_num=ohiou1610705872573225.
Повний текст джерелаGuzman, Castro Gustavo Adolfo [Verfasser], and Stephan [Akademischer Betreuer] Reichl. "Human Hemicornea Model for Drug Transport Testing and Screening of Excipients / Gustavo Adolfo Guzman Castro ; Betreuer: Stephan Reichl." Braunschweig : Technische Universität Braunschweig, 2017. http://d-nb.info/1175816949/34.
Повний текст джерелаZraikat, Manar Saleh Ali. "Development of in vitro models of invasion for the pharmacological investigation of small molecule inhibitors of tumour progression : development and validation of a 3-dimensional tumour spheroid invasion model to evaluate the pharmacological effects of novel small molecule β3 integrin antagonists". Thesis, University of Bradford, 2015. http://hdl.handle.net/10454/7511.
Повний текст джерелаLee, Bill. "Preclinical antimicrobial drug discovery : development and evaluation of a platform for high-throughput screening in vitro and an immunocompromised animal model." Thesis, McGill University, 2006. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=100745.
Повний текст джерелаRobinson, Clayt Austin. "Development of an in vitro three-dimensional model for colon cancer study and drug efficacy analysis." Connect to resource, 2005. http://rave.ohiolink.edu/etdc/view?acc%5Fnum=osu1124223577.
Повний текст джерелаTitle from first page of PDF file. Document formatted into pages; contains xvi, 204 p.; also includes graphics (some col.). Includes bibliographical references (p. 196-204). Available online via OhioLINK's ETD Center
Lindquist, Tera M. "The development of zebrafish (Danio rerio) as a rapid and efficient model system for therapeutic drug screening for Spinal Muscular Atrophy." The Ohio State University, 2011. http://rave.ohiolink.edu/etdc/view?acc_num=osu1311694979.
Повний текст джерелаMosaad, Eman Mohamed Othman. "Three dimensional prostate cancer model systems." Thesis, Queensland University of Technology, 2018. https://eprints.qut.edu.au/118287/1/Eman%20Mohamed%20Othman_Mosaad_Thesis.pdf.
Повний текст джерелаWan, Xiao. "Development of advanced three-dimensional tumour models for anti-cancer drug testing." Thesis, University of Oxford, 2014. http://ora.ox.ac.uk/objects/uuid:5342fe46-c676-4fe8-8b6e-96d17a18d17d.
Повний текст джерелаMalvezzi, Alberto. "Modelos de virtual Screening de inibidores da cruzaína: desenvolvimento e validação experimental." Universidade de São Paulo, 2008. http://www.teses.usp.br/teses/disponiveis/46/46135/tde-10082016-115529/.
Повний текст джерелаIn order to search and identify new cruzain inhibitor(s) - a cysteine-protease of Trypanosoma cruzi, the etiologic agent of Chagas disease - two virtual screening schemes(Models I and II) were proposed, validated- and applied to the ZINC database (3.294.714 compounds). The proposed virtual screening models, bearing a sequence of different physicalchemical, pharmacophore and docking filters, as well as a visual inspection filter, were built from information taken from 13 cruzain complexes and from 20 complexes of other cysteine proteases, having their structures available in PDB. In a first step, a detailed recognition of the cruzain structural features and characteristics was performed through visual inspection of the enzyme environment; followed by the analysis of GRID generated molecular interaction fields; through the identification of molecular interaction properties exposed at the enzyme cavity surface, generated by the CAVBASE program; and by molecular dynamics simulations. The virtual screening Model I, - generated from the structural characteristics recognized from 13 PDB cruzain complexes - when applied to the ZINC database selected 10 compounds. For the experimental validation ofthe model, six ofthese compounds have been acquired and were tested as cruzain inhibitors. It was observed that three of the tested compounds (ZINC02470662, ZINC02682879 and ZINC03192044, respectively) did not show any significant cruzain inhibition, up to 7 mM. Meanwhile the other three tested compounds (ZINC02663001, ZINC01936854 and ZINC03326243, respectively) showed an unspecific cruzain inhibition, suggesting that an enzyme inhibition by promiscuous mechanism occurred. This mechanism was verified by the addition of 0.1% Triton X-100 on the enzymatic assay with a concomitant loss of cruzain inhibition activity. For these compounds, the confirmation of the promiscuous mechanism was also done, observing the loss of enzyme inhibition, after a ten times increase in the cruzain concentration on the enzymatic assay. The virtual screenmg Model II - generated from the structural characteristics recognized from 13 cruzain complexes and 20 complexes of other cysteine proteases, that have been identified on a search for cavities similar to cruzain - selected 55 compounds, when applied to the ZINC database. In order to experimentally validate the model, nineteen compounds have been acquired and were tested as cruzain inhibitors. It has been observed that one compound, ZINC01794422, showed a specific cruzain inhibition (Ki = 21 µM), while the other eighteen showed no significant inhibition, up to 592 µM concentration. The promiscuous mechanism of enzymatic inhibition was not observed, since 0.1% of Triton X-100 was added in ali assays. Additionally, Model II identified two other cruzain inhibitors (ZINC04899534 and ZINC01547017). However, these compounds have not been acquired or tested, since they are known cruzain inhibitors - already described in the literature and are structurally similar to the inhibitors used in the construction of the mode!. Referring to new inhibitors found, Model II showed a hit rate of 5,3%. This value is in agreement with those found in the literature, which ranges from 1 to 50%.
Babahosseini, Hesam. "Single Cell Biomechanical Phenotyping using Microfluidics and Nanotechnology." Diss., Virginia Tech, 2016. http://hdl.handle.net/10919/64502.
Повний текст джерелаPh. D.
Wang, Yuanyuan (Marcia). "Statistical Methods for High Throughput Screening Drug Discovery Data." Thesis, University of Waterloo, 2005. http://hdl.handle.net/10012/1204.
Повний текст джерелаClassification methods are commonly proposed as solutions to this problem. However, regarding drug discovery, researchers are more interested in ranking compounds by predicted activity than in the classification itself. This feature makes my approach distinct from common classification techniques.
In this thesis, two AIDS data sets from the National Cancer Institute (NCI) are mainly used. Local methods, namely K-nearest neighbours (KNN) and classification and regression trees (CART), perform very well on these data in comparison with linear/logistic regression, neural networks, and Multivariate Adaptive Regression Splines (MARS) models, which assume more smoothness. One reason for the superiority of local methods is the local behaviour of the data. Indeed, I argue that conventional classification criteria such as misclassification rate or deviance tend to select too small a tree or too large a value of k (the number of nearest neighbours). A more local model (bigger tree or smaller k) gives a better performance in terms of drug discovery.
Because off-the-shelf KNN works relatively well, this thesis takes this promising method and makes several novel modifications, which further improve its performance. The choice of k is optimized for each test point to be predicted. The empirically observed superiority of allowing k to vary is investigated. The nature of the problem, ranking of objects rather than estimating the probability of activity, enables the k-varying algorithm to stand out. Similarly, KNN combined with a kernel weight function (weighted KNN) is proposed and demonstrated to be superior to the regular KNN method.
High dimensionality of the explanatory variables is known to cause problems for KNN and many other classifiers. I propose a novel method (subset KNN) of averaging across multiple classifiers based on building classifiers on subspaces (subsets of variables). It improves the performance of KNN for HTS data. When applied to CART, it also performs as well as or even better than the popular methods of bagging and boosting. Part of this improvement is due to the discovery that classifiers based on irrelevant subspaces (unimportant explanatory variables) do little damage when averaged with good classifiers based on relevant subspaces (important variables). This result is particular to the ranking of objects rather than estimating the probability of activity. A theoretical justification is proposed. The thesis also suggests diagnostics for identifying important subsets of variables and hence further reducing the impact of the curse of dimensionality.
In order to have a broader evaluation of these methods, subset KNN and weighted KNN are applied to three other data sets: the NCI AIDS data with Constitutional descriptors, Mutagenicity data with BCUT descriptors and Mutagenicity data with Constitutional descriptors. The k-varying algorithm as a method for unbalanced data is also applied to NCI AIDS data with Constitutional descriptors. As a baseline, the performance of KNN on such data sets is reported. Although different methods are best for the different data sets, some of the proposed methods are always amongst the best.
Finally, methods are described for estimating activity rates and error rates in HTS data. By combining auxiliary information about repeat tests of the same compound, likelihood methods can extract interesting information about the magnitudes of the measurement errors made in the assay process. These estimates can be used to assess model performance, which sheds new light on how various models handle the large random or systematic assay errors often present in HTS data.
Sleigh, James Nicholas. "Model systems for exploring new therapeutic interventions and disease mechanisms in spinal muscular atrophies (SMAs)." Thesis, University of Oxford, 2012. http://ora.ox.ac.uk/objects/uuid:378416c5-a586-4a2a-980c-81dfff6803df.
Повний текст джерелаLarsson, Dhana E. "Analyses of Dose-Response and Mechanistic Action of Different Anti-Cancer Drugs for Neuroendocrine Tumor Cell Lines." Doctoral thesis, Uppsala universitet, Institutionen för medicinska vetenskaper, 2011. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-158833.
Повний текст джерелаTitle corrected from: Analyses of Dos-Response and Mechanistic Action of Different Anti-Cancer Drugs for Neuroendocrine Tumor Cell Lines
Modak, Swananda Rajan. "Effects of Aβ42 on the human proteome and compound library screening using cellular models of Alzheimer's disease". Thesis, University of Manchester, 2013. https://www.research.manchester.ac.uk/portal/en/theses/effects-of-a42-on-the-human-proteome-and-compound-library-screening-using-cellular-models-of-alzheimers-disease(4cca38b1-fd3f-4ef1-bde1-e16cf227ef68).html.
Повний текст джерелаCrutchley, James E. B. "Automation and scale-up of human induced pluripotent stem cell models of cardiovascular disease for drug screening." Thesis, University of Nottingham, 2016. http://eprints.nottingham.ac.uk/32207/.
Повний текст джерелаSantiago, Daniel Navarrete. "Use and Development of Computational Tools in Drug Discovery: From Small Molecules to Cyclic Peptides." Scholar Commons, 2012. http://scholarcommons.usf.edu/etd/4398.
Повний текст джерелаHe, Shanshan. "Neglected Tropical Disease Chemotherapy: Mechanistic Characterization of Antitrypanosomal Dihydroquinolines and Development of a High Throughput Antileishmanial Screening Assay." The Ohio State University, 2012. http://rave.ohiolink.edu/etdc/view?acc_num=osu1337980540.
Повний текст джерелаAbdulHameed, Mohamed Diwan Mohideen. "COMPUTATIONAL DESIGN OF 3-PHOSPHOINOSITIDE DEPENDENT KINASE-1 INHIBITORS AS POTENTIAL ANTI-CANCER AGENTS." UKnowledge, 2009. http://uknowledge.uky.edu/gradschool_diss/757.
Повний текст джерелаRivera, Maria [Verfasser], Roland [Akademischer Betreuer] Lauster, Roland [Gutachter] Lauster, Wolfgang [Gutachter] Walther, and Jens [Gutachter] Kurreck. "Colorectal cancer patient-derived xenograft (PDX) models as platform for drug screening, molecular and response analysis / Maria Rivera ; Gutachter: Roland Lauster, Wolfgang Walther, Jens Kurreck ; Betreuer: Roland Lauster." Berlin : Technische Universität Berlin, 2016. http://d-nb.info/1156013364/34.
Повний текст джерелаPasquet, Vivian [Verfasser], Klaus [Gutachter] Brehm, August [Gutachter] Stich, Jörg [Gutachter] Vogel, and Gustavo [Gutachter] Salinas. "Characterization of thioredoxin and glutathione reductase activities of Mesocestoides vogae, a flatworm parasite useful as a laboratory model for the screening of drugs. / Vivian Pasquet. Gutachter: Klaus Brehm ; August Stich ; Jörg Vogel ; Gustavo Salinas." Würzburg : Universität Würzburg, 2014. http://d-nb.info/1102828955/34.
Повний текст джерелаFernandes, William Borges. "Determinação do modo de interação de inibidores reversíveis da cruzaína de Trypanosoma cruzi via cristalografia de raios X." Universidade de São Paulo, 2015. http://www.teses.usp.br/teses/disponiveis/75/75133/tde-28082015-081328/.
Повний текст джерелаCruzain, the major Trypanosoma cruzi cysteine protease, is a validated therapeutic target for the search of new medicines for the treatment of Chagas disease. A myriad of inhibitors of this enzyme consists of covalent irreversible peptidomimetics whose development has been impaired due to potential off target effect and undesirable pharmacokinetic profile. Modern cheminformatic methods employed by The Medicinal Chemistry Group (NEQUIMED/IQSC/USP) were used to identify cruzain reversible inhibitors. Their optimization for more efficient compounds can be accomplished by the use of data and information gathered from computational and structural modes of interaction (MOB). In this doctoral thesis, the X-ray crystallography was used to describe in detail the MOB of three important new cruzain reversible inhibitors: the dipeptidil-nitrile Neq0409 and the two molecular fragmentos Neq0147 and Neq0176. In order to avoid cruzain self-proteolysis during crystallization with low affinity reversible inhibitors such as the identified fragments, two new mutant and inactive constructs of cruzain (the C25S and C25A) were designed to upholding the same properties of the wild type catalytic site. The C25S was validated as representative crystallographic model given the coherence of the MOB elucidated for the best-known and studied cruzain inhibitor, the K777. The description of the presence, orientation, conformation and mode of action at the site, besides the complete pattern of bimolecular interactions, provided by these crystalline structures, orthogonally validated the predicted in silico MOB and allowed the identification of other potent inhibitors analogous to the Neq0147 and the Neq0409. The 2-acetamidothiophene-3-carboxamide moiety (Neq0176) and heterocyclic five-membered ring based on the 1,3,4-oxadiazole (Neq0147) were thereby identified as attractive alternatives to traditional peptidomimetics. The impact of proteolytic effect on the quality and resolution of crystallographic structures in cruzain was best understood from two high-resolution structures obtained for the native cruzain in complex with methyl methanethiolsulfonate (MMTS) and iodoacetamide. These results allow the understanding of experimental basis for the crystallization with reversible inhibitors of low affinity such as fragments. All structural results obtained by X-ray crystallography together with the catalytic site depiction using GRID and SuperStar methods are useful for mapping the essential structural basis for the design of future more potent and selective cruzain inhibitors.
Elitt, Matthew S. "DISEASE MODELING AND THERAPEUTIC DEVELOPMENT FOR PELIZAEUS-MERZBACHER DISEASE." Case Western Reserve University School of Graduate Studies / OhioLINK, 2019. http://rave.ohiolink.edu/etdc/view?acc_num=case1536687505814955.
Повний текст джерелаFerreira, Luís Pedro Correia Pinto. "Development of multicelular 3D cancer testing platforms for evaluation of new anti-cancer therapies." Master's thesis, Universidade de Aveiro, 2017. http://hdl.handle.net/10773/22713.
Повний текст джерелаO cancro do pulmão (CP) é um dos cancros mais diagnosticados a nível mundial e também um dos mais mortíferos. Atualmente, as terapias administradas a nível clínico para o tratamento do CP são ainda extremamente ineficazes e limitadas no que diz respeito ao aumento da taxa de sobrevivência dos pacientes oncológicos. Esta realidade demonstra a necessidade de investigar ativamente novas terapias para o tratamento desta neoplasia. No entanto a validação pré-clínica de terapias inovadoras para o CP tem-se revelado extremamente difícil devido à inexistência de plataformas que sejam adequadas para testes a nível laboratorial, uma vez que as culturas celulares in vitro bidimensionais (2D), recomendadas pelas agências regulatórias são incapazes de mimetizar as caraterísticas principais dos tumores humanos. Estas limitações têm originado uma fraca correlação entre a performance das terapias nos estudos in vitro e a obtida em ensaios clínicos controlados. Neste contexto, os modelos de tumores tridimensionais (3D) in vitro têm vindo a ser reconhecidos como uma solução para este problema, pois podem recapitular várias componentes do microambiente tumoral. Das várias plataformas 3D in vitro de CP investigadas atualmente muito poucas avaliaram o papel da inclusão de células estaminais mesenquimais (MSCs). Para colmatar esta lacuna, o trabalho de investigação desenvolvido no âmbito desta dissertação descreve a produção e otimização de novos modelos hétero-celulares 3D in vitro. Estas plataformas são compostas por células tumorais do CP (A549) e do seu estroma, nomeadamente fibroblastos da pele e células estaminais mesenquimais derivadas da medula óssea (BM-MSCs). Estes três tipos de células foram co-cultivadas em micropartículas poliméricas de policaprolactona revestidas por ácido hialurónico, com o objetivo de incluir este componente da matriz extracelular que se encontra presente no microambiente do CP. Esta abordagem permitiu formar a nível laboratorial microtecidos multicelulares 3D híbridos que melhor mimetizam a heterogeneidade celular das neoplasias pulmonares. Os resultados obtidos demonstraram que os microtumores formados através da técnica de sobreposição-líquida são reprodutíveis em termos de morfologia e tamanho, apresentaram núcleos necróticos, organização celular 3D e produziram proteínas do microambiente tumoral. Além destas caraterísticas, os dados obtidos através de microscopia de fluorescência revelaram que as BM-MSCs migram para o interior dos microtumores ao longo do tempo. A avaliação da citotoxicidade da Doxorubicina, um fármaco anti-tumoral rotineiramente utilizado a nível clínico, demonstrou que a inclusão de micropartículas aumenta a resistência das células tumorais em modelos homotípicos. Nos modelos tri-cultura heterotípicos a citotoxicidade foi comparável à obtida em microtumores sem micropartículas. Estes resultados evidenciam assim o papel importante dos fibroblastos e das BM-MSCs na resposta dos microtumores. Numa visão global, os modelos 3D formados recapitulam com mais exatidão o microambiente do cancro do pulmão e poderão servir no futuro como plataformas de teste para descobrir ou aperfeiçoar novas terapias, ou combinações de terapêuticas, para este tipo de neoplasia.
Lung cancer (LC) is one the most commonly diagnosed cancers worldwide, being also one of the deadliest. Currently, clinically administered therapies for treatment of LC are still extremely ineffective and limited in increasing oncologic patients survival rates. This reality evidences the necessity of actively investigating novel therapies for the treatment of LC. However, preclinical validation of novel therapies as revealed itself as an extremely arduous process, due to the lack of suitable laboratory testing platforms since the recommend in vitro bi-dimensional (2D) cell cultures are unable to fully mimic the main hallmarks of human tumors. In this context, in vitro tridimensional (3D) tumor models are being increasingly recognized as a solution due to their ability to correctly recapitulate several characteristics of the tumor microenvironment (TME). Amongst currently developed 3D in vitro platforms for the study of LC, few have included or studied the role of mesenchymal stem cells (MSCs). To provide further insights into this hypothesis, the research work developed in this thesis describes the production and optimization of novel heterotypic in vitro 3D models, comprised by non-small-cell lung cancer cells (A549) and stromal cells, namely skin fibroblasts (HFs), and bone-marrow derived mesenchymal stem cells (BM-MSCs). These three diverse cell populations were co-cultured in hyaluronic acid coated polymeric polycaprolactone microparticles (LbL-MPs) as to include this key extracellular matrix component of LC TME. This approach allowed the formation of 3D multicellular heterotypic microtissues (3D-MCTS) that better recapitulate the cellular heterogeneity of LC TME in the laboratory. The obtained findings demonstrate that these models formed via the liquid-overlay technique were reproducible in terms of morphology and size, presented necrotic core formation, 3D cellular organization, and deposited matrix proteins in a similar manner as in the TME. Besides this, fluorescence microscopy data revealed that BM-MSCs migrated overtime into the microtumors core . Performed doxorubicin in vitro cytotoxicity assays revealed that the inclusion of LbL-MPs lead to an increased resistance of homotypic A549 monoculture models against this anti-cancer drug commonly used in clinical treatments. Alongside, the cytotoxicity obtained in triculture heterotypic models was comparable to that of microtumors without LbL-MPs inclusion, showcasing the role of HFs and BM-MSCs in microtumors response to therapy. Globally, the herein bioengineered 3D models were able to recapitulate with an increased precision the TME of LC, making them suitable test platforms for development or improvement of standalone or combinatorial therapies for this type of neoplasia.
Durán, Alcaide Ángel. "Development of high-performance algorithms for a new generation of versatile molecular descriptors. The Pentacle software." Doctoral thesis, Universitat Pompeu Fabra, 2010. http://hdl.handle.net/10803/7201.
Повний текст джерелаEl trabajo que se presenta en esta tesis se ha centrado en el desarrollo de algoritmos de altas prestaciones para la obtención de una nueva generación de descriptores moleculares, con numerosas ventajas con respecto a sus predecesores, adecuados para diversas aplicaciones en el área del diseño de fármacos, y en su implementación en un programa científico de calidad comercial (Pentacle). Inicialmente se desarrolló un nuevo algoritmo de discretización de campos de interacción molecular (AMANDA) que permite extraer eficientemente las regiones de máximo interés. Este algoritmo fue incorporado en una nueva generación de descriptores moleculares independientes del alineamiento, denominados GRIND-2. La rapidez y eficiencia del nuevo algoritmo permitieron aplicar estos descriptores en cribados virtuales. Por último, se puso a punto un nuevo algoritmo de codificación independiente de alineamiento (CLACC) que permite obtener modelos cuantitativos de relación estructura-actividad con mejor capacidad predictiva y mucho más fáciles de interpretar que los obtenidos con otros métodos.
Wang, Yun-Lin, and 王允麟. "PPP2R2B: Epigenetic Study and Establishment of Drug Screening Model." Thesis, 2011. http://ndltd.ncl.edu.tw/handle/92422773542420098613.
Повний текст джерела國立臺灣師範大學
生命科學研究所
99
Protein phosphatase 2A (PP2A) is a major serine/threonine phosphatase expressed in all eukaryotic cells. The regulatory B subunit confers substrate specificity and sub-cellular localization of the PP2A holoenzyme. PPP2R2B is a regulatory B subunit expressed throughout the brain. The brain specific PPP2R2B regulates PP2A dephosphorylation activity for microtubule-associated protein tau. The association of pathological hyperphosphorylated tau and reduced PP2A activity with Alzheimer's disease (AD) has been established. Our case-control study and reporter assay have also revealed that the rare low transcriptional activity alleles of PPP2R2B are associated with AD. In the first part of the study, PPP2R2B-driven EGFP construct was used to generate human embryonic kidney (HEK)-293 and neuroblastoma SK-N-SH cell lines. SP1 and CREB1 co-transfection did not enhance PPP2R2B expression. In the second part of the study, epigenetic control of DNA methylation affecting AD susceptivity was explored. Bisulfite sequencing was performed to assess the DNA methylation using lymphocyte DNA from five AD patients and age- and gender-matched controls. The results of increased DNA methylation (although not significantly) in the PPP2R2B gene 5' region, especially -311 and -310, suggest that the epigenetic change may alter the PPP2R2B expression in AD patients. Further direct bisulfite-sequencing of HEK-293 cells revealed increased DNA methylation in the PPP2R2B gene 5' region. However, real-time PCR quantitation of 5-aza-dC treated HEK-293 cells did not show increased PPP2R2B expression. Chromatin IP with MeCP2 antibody and PCR also did not support MeCP2 binding to PPP2R2B promoter.
Mathews, Bobby. "A zebrafish model system for drug screening in diabetes." Thesis, 2019. http://urn.kb.se/resolve?urn=urn:nbn:se:his:diva-17847.
Повний текст джерелаTeng, Wei-Lin, and 滕暐林. "Generation of a Zebrafish Model for Anti-Intrahepatic Cholangiocarcinoma Drug Screening." Thesis, 2015. http://ndltd.ncl.edu.tw/handle/23208518555809201768.
Повний текст джерела國立臺灣海洋大學
生命科學暨生物科技學系
103
Intrahepatic cholangiocarcinoma (ICC) is the second most common type of primary liver cancer. It is a rare malignant tumor that not easy to detect at early stage. Today, there is no effective adjuvant treatment for ICC. Therefore, to establish a platform for drug screening is extremely important. To develop high throughput drug screening platform for anti-ICC drug screening, we developed a xenograft model by transplant fluorescent-labeled human bile duct cancer cells into the yolk sac of 2-day-old zebrafish embryos. The established ICC model of HBx+HCP transgenic zebrafish was used to evaluate candidate drugs. According to the results of pathway maps of zebrafish ICC, Sorafenib (Nexavar), XAV-939, LY2109761 and SB431542 were used to test their anti-ICC ability in ICC cells, xenografts and HBx+HCP transgenic zebrafish, respectively. In the analysis of cell cycle, results revealed significant cell population in sub G1 area when cells were treated with Sorafenib, XAV-939, LY2109761, and SB431542, respectively. In migration assay, the results showed that LY2109761 and SB431542 reduced the ICC cells migration ability; they also inhibited TGF1 pathway downstream gene and metastasis marker mmp9 and mmp2 gene expression. In ICC xenografts, dissemination rate were also reduced by LY2109761 and SB431542 treatment. Compare the results above shows LY2109761 is the more effective drug than others. In the inhibition of ICC in HBx and HCP transgenic zebrafish, LY2109761 inhibited 86% ICC formation compared with control. Taken together, this data demonstrates the potential of LY2109761 to inhibit ICC formation.
Wu, Bo-Kai, and 吳百凱. "Establishment of zebrafish as an animal model for Tauopathy research and drug screening." Thesis, 2016. http://ndltd.ncl.edu.tw/handle/90527521055809637995.
Повний текст джерела國立臺灣大學
生物科技研究所
104
Tau protein is a tubulin-binding protein, which plays important roles in the formation and stability of the microtubule. Mutations in the tau gene are associated with familial forms of frontotemporal dementia with Parkinsonism linked to chromosome-17 (FTDP-17). Neurofibrillary tangles (NFTs) of Tau and extracellular plaques containing amyloid-β (Aβ) are found in the brain of Alzheimer’s disease (AD) patients. Transgenic models, including those of zebrafish, have been employed to elucidate the mechanisms by which Tau protein causes neurodegeneration. In this study, a transient expression system was established to express GFP fusion proteins of zebrafish and human Tau under the control of a neuron-specific HuC promoter. The expression of Tau-GFP was observed to cause high levels of neuronal death which could be directly traced in vivo. Multiple signaling factors, such as Bcl2-L1, Nrf2, and GDNF, were found to effectively protect neuronal cells expressing Tau-GFP from death. Treatment with compounds that induce anti-oxidative or neurotrophic effects also resulted in a similar neuronal protective effect and maintained human Tau-GFP protein in a phosphorylated state, as detected by antibodies pT212 and AT8. Therefore, we used this model to screen 400 herbal extracts and found 45 of them to be effective on reducing Tau-GFP-induced neuronal death. One of the effective herbal extracts is the Tripterygium wilfordii. HPLC analysis and functional assay demonstrated that epicatechin (EC) is the major compound of Tripterygium wilfordii stem extract to decrease the neurotoxicity induced by Tau-GFP. ARE (antioxidant response elements)-luciferase reporter gene assay is usually used to detect the activity of Nrf2. We used the assay to demonstrate that EC could increase the activity of Nrf2 in zebrafish. These data suggest that EC from the Tripterygium wilfordii stem extract could diminish Tau-GFP-induced neuronal death through the activation of Nrf2.
(8803004), Logan C. Ganzen. "Drug Screening Utilizing the Visual Motor Response of a Zebrafish Model of Retinitis Pigmentosa." Thesis, 2020.
Знайти повний текст джерелаYen, Hsiao-Hsiu, and 顏孝修. "Establishment of an Inducible Cell Model System for Spinocerebellar Ataxia Type 3 Potential Drug Screening." Thesis, 2009. http://ndltd.ncl.edu.tw/handle/06681769489654323319.
Повний текст джерела國立臺灣師範大學
生命科學研究所
98
Spinocerebellar ataxia type 3 (SCA3), also known as Machado-Joseph disease (MJD), is the most common autosomal dominant inherited ataxia. SCA3 is caused by a CAG trinucleotide expansion in the MJD1 gene, which is located on chromosome 14q24.3-q31, and translated into a polyglutamine (polyQ) stretch containing ataxin-3 (AT3) protein. Expansion of the glutamine domain in AT3, as well as in eight other members of polyQ neurodegenerative disease family, increases protein misfolding, results in aggregation and formation of nuclear and cytoplasmic inclusions. Inclusions of polyQ proteins are ubiquitylated and contain proteasomes, suggesting an attempt by the ubiquitin proteasome system (UPS) to degrade the misfolded protein. However, little is known about the correlation between aggregate formation and cell death. To further study the role of AT3 in SCA3 neuropathology, we have established AT3-inducible PC12 cells. This cell model should allow us to characterize the aggregation of full-length AT3 protein in living cells. Our results showed that AT3 mutant protein (75Q) expression in the PC12 cells intend to cause the formation of the nuclear or peri-nuclear aggregation. Futhermore, cell survival declined with the expression of extended polyQ AT3, especially under differentiated condition induced by NGF treatment. In addition, we found that neurotoxicity of expanded AT3 (Q75) could further cause unhealthy morphology and high sensitivity to oxidative stress. With this cell model, we have evaluated the neuron protective effect of several histone deacetylase (HDAC) inhibitors and antioxidant compounds. All of these HDAC inhibitors could sustain cell viability significantly when cells were treated with low dosage of these compounds. These results suggest that regulation of gene expression is involved in the neuropathology of SCA3. In conclusion, we have established an SCA3 inducible cell model which could be used as a platform for screening of potential therapeutic strategies for SCA3 and other polyQ-mediated neurodegenerative diseases.
Liu, Chi-Yin, and 劉琦殷. "The Establishment of Platelet-Collagen Adhesion Assay Model for Screening of Novel Anti-GPVI Drug." Thesis, 2012. http://ndltd.ncl.edu.tw/handle/11203743931076978554.
Повний текст джерела國立陽明大學
藥理學研究所
100
The platelet activation at the site of vascular damage is crucial for development of arterial thrombosis. The anti-platelet drugs are widely used in prevention of thrombotic diseases. However, side effects of current anti-platelet drugs are still unsolved. Glycoprotein VI (GPVI) is a platelet-specific collagen receptor. GPVI not only participates in platelet adhesion on collagen of subendothelial matrix but also activates the signal through FcRγ-chain mediated tyrosine phosphorylation-dependent pathway. Recent reports show that GPVI inhibition has no side effect of bleeding. Thus, inhibitors blocking GPVl-collagen interaction may help in antithrombotic therapy. In addition, the collagen-related anti-platelet agents have not been developed. To explore the novel anti-GPVI drugs, the convenient platelet-collagen adhesion assay model was established. The result found that the adherence of calcein-loaded platelets on 0.2 mg/ml of collagen-coated plates was detected by fluorescence analysis at 30 minutes. In parallel, the inhibition of losartan (GPVI-antagonist) on convulxin (GPVI agonist) induced platelet aggregation assay was also tested. Our results found that losartan can dose-dependent suppress both calcein-loaded platelets adherence and convulxin-induced platelet aggregation. These data indicated that the feasibility of the analytical model for GPVIcollagen drug searching. In the future, we will use this model can be used to search the potential lead compounds from Chinese herbal medicine and the inhibitory effect will be further confirmed by aggregation assay. Caihu extracts (Bupleurum falcatum) was supplied by Dr. Lu, Chung-Kuang in National Research Institute of Chinese Medicine. Caihu induces platelets lysis that will lead to destroy platelets structure, and the fluorescence decreases in adhesion assay, and the transmission increases in aggregation assay. Yejiaoteng induces platelets shape change under 100 g/ml. And it can explain the fluorescence increasing highly in adhesion assay, because the surface area of platelet is spreading that elevated the binding between collagen and platelets, and it induces GPVI signaling and activates intergrin 21, then form stable adhesion. It interestly focuses that the transmission is higher pretreated CV3988 (PAR antagonist) then add Yejiaoteng to induce platelet aggregation compared with control. It suggests that PAF receptor is indispensable in the signal transduction of Yejiaoteng. In platelet-collagen adhesion model, we found that Caihu doesn’t have inhibition ability of platelets adhesion, and Yejiaoteng has mildly increasing platelets shape change.
Fried, Sabrina Liora. "Drug screening in gastro-esophageal adenocarcinoma and the advantages of the organoid model: a literature review." Thesis, 2020. https://hdl.handle.net/2144/41236.
Повний текст джерелаYang, Miao-Chia, and 楊苗佳. "Using CRISPR/Cas9-Mediated GLA-null Cell Lines as An In Vitro Drug Screening Model for Fabry Disease." Thesis, 2016. http://ndltd.ncl.edu.tw/handle/49926843078539991979.
Повний текст джерела國立陽明大學
藥理學研究所
104
Fabry disease is a hereditary, X-linked lysosomal storage disease resulting from deficient activity of the lysosomal α-galactosidase A. It leads to progressive accumulation of glycosphingolipids particularly globotriaosylceramide (GL-3) in lysosomes of the heart, kidneys, skin and various tissues. Regular administration of recombinant human alpha Gal A (rh-α-GLA), termed enzyme replacement therapy (ERT) is currently available as the only effective treatment for the Fabry patients with GL-3 accumulation. However, the rh-α-GLA driven GL-3 clearance has the limitations, i.e. rh-α-GLA is physiologically instable and quickly degraded in cells. Moreover, lacking of an appropriate in vitro disease model restricted the pharmaceutical studies for improving the ERT treatment. Therefore, it is worth to establish a cell model of Fabry disease (FD) as the platform to screen the potential candidates for prolonging its potency. By utilizing the CRISPR/Cas9 genome editing system, we generated the GLA disruption in HEK293T cells that was completely devoid of detectable GLA protein expression and enzyme activity, providing a clear background to investigate rh-α-GLA cellular pharmacokinetics. The administrated rh-α-GLA was decreased with time and had a half-life of 24 hrs in the GLA-null cells. Base on the GLA deficient cell line, we applied to discover the potential drug or small molecular to restore rh-α-GLA activity. Co-treatment of chaperone drug, N-butyldeoxygalactonojirimycin (NB-DGJ), and protease inhibitor, E64, with ERT significantly prolonged rh-α-GLA activity by over two-folds compared to ERT alone. In addition, NB-DGJ and E64 significantly decreased GL-3 accumulation in the Fabry patients-derived fibroblast. Next, we expanded the screening range of drug and identified the activity for discovering other potential drugs. We screened 64 drugs combining ERT in GLA-null cells and discovered that Calpain inhibitor II, E64C, 2-NBDG, β-D-Galactose pentapivalate, 2-Deoxy-D-galactose, Finasteride, Diazepam, Theophylline, Trazodone, Benzamidine, 3-Methyladenine, Carbamazepine, Selegiline, Sulpiride and Fluorouracil could prolong rh-α-GLA activity. By creating this model, we provide a novel in vitro tool with which to screen potential compounds to avoid short period of GLA activity in human body.
Monteiro, Maria Vinhas. "Development of biomimetic pancreatic cancer 3D in vitro models for preclinical drug screening." Master's thesis, 2020. http://hdl.handle.net/10773/30418.
Повний текст джерелаO adenocarcinoma ductal pancreático (ADP) é uma doença com uma das maiores taxas de mortalidade e com uma incidência crescente a nível mundial. Atualmente, as terapias administradas na clínica para o tratamento do ADP são ainda extremamente ineficazes e de acesso limitado. Perante este cenário torna-se urgente a investigação e validação de novas terapias para o tratamento desta neoplasia. O ADP é um cancro com uma bioarquitetura estratificada única e caracterizado por uma exacerbada reação desmoplásica envolvendo fibroblastos associados ao cancro, células imunes e proteínas da matriz extracelular (MEC), que desempenham um papel significativo na progressão tumoral e na resistência às terapias utilizadas atualmente em contexto clínico. A ausência de modelos de celulares capazes de reproduzir in vitro o microambiente desmoplásico e a histo-morfologia do ADP origina uma baixa correlação entre a performance de novas terapias obtida em ensaios pré-clínicos e aquela observada em ensaios clínicos controlados. Neste sentido, os modelos de tumores tridimensionais (3D) in vitro surgem como uma solução mais adequada para a avaliação pré-clínica quando comparados com as recomendadas culturas celulares bidimensionais 2D. Os modelos 3D representam modelos mais biomiméticos pois permitem recapitular de uma forma mais robusta e realista o microambiente tumoral, contribuindo para a descoberta de novos biomarcadores e para a avaliação pré-clínica de novos fármacos de uma forma mais precisa. Das plataformas 3D in vitro de ADP desenvolvidas atualmente, poucas são as que recapitulam de forma precisa a heterogeneidade celular, a arquitetura tumoral e o estroma fibrótico. Com o objetivo de colmatar estas limitações, a presente dissertação foca na bioengenharia e caracterização de um novo modelo 3D de ADP, consistindo numa co-cultura biomimética de células cancerígenas pancreáticas (PANC-1) e fibroblastos associados ao cancro (FACs). Este modelo 3D demonstrou recapitular os componentes celulares, a sua distribuição espacial e a resistência a terapias farmacológicas de uma forma semelhante à encontrada nos tumores humanos.
Mestrado em Biotecnologia
Ahmed, Mohaned S. A., Haneen A. Basheer, J. M. Ayuso, Djevdet S. Ahmet, Marco Mazzini, Roshan Patel, Steven D. Shnyder, Victoria Vinader, and Kamyar Afarinkia. "Agarose Spot as a Comparative Method for in situ Analysis of Simultaneous Chemotactic Responses to Multiple Chemokines." 2017. http://hdl.handle.net/10454/11904.
Повний текст джерелаWe describe a novel protocol to quantitatively and simultaneously compare the chemotactic responses of cells towards different chemokines. In this protocol, droplets of agarose gel containing different chemokines are applied onto the surface of a Petri dish, and then immersed under culture medium in which cells are suspended. As chemokine molecules diffuse away from the spot, a transient chemoattractant gradient is established across the spots. Cells expressing the corresponding cognate chemokine receptors migrate against this gradient by crawling under the agarose spots towards their centre. We show that this migration is chemokine-specific; meaning that only cells that express the cognate chemokine cell surface receptor, migrate under the spot containing its corresponding chemokine ligand. Furthermore, we show that migration under the agarose spot can be modulated by selective small molecule antagonists present in the cell culture medium.
Burns, C. J., E. Fantino, A. K. Powell, Steven D. Shnyder, Patricia A. Cooper, S. Nelson, C. Christophi, et al. "The microtubule depolymerizing agent CYT997 causes extensive ablation of tumor vasculature in vivo." 2011. http://hdl.handle.net/10454/5902.
Повний текст джерела黃慧茹. "Spinocerebellar ataxia type 17: Genetic testing and development of biochemical and yeast models for drug screening." Thesis, 2007. http://ndltd.ncl.edu.tw/handle/94035774752699133983.
Повний текст джерела國立臺灣師範大學
生命科學研究所
95
Autosomal dominant spinocerebellar ataxias (SCAs) are a heterogeneous group of neurodegenerative disorders characterized by progressive cerebellar ataxia of gait and limbs variably associated with dementia, dysarthria, etc. More than 28 SCA types have been described. Among them, SCA type 17 (SCA17) is caused by an expanded polyglutamine (polyQ) in the TATA-box binding protein (TBP), a general transcription initiation factor. In addition to SCA17, neurodegenerative diseases including HD, SBMA, DRPLA and SCA types 1, 2, 3, 6, 7 are also caused by expansion of a CAG repeat tracts encoding expanded polyQ tracts. The expanded polyQ tract causes a conformational change in the polypeptide to promote misfolding and aggregation of the disease protein, leading to the death of neurons. Unrelated expanded alleles characterized in familial and sporadic ataxia patients are found ranging from 43~66 repeats as opposed to 25~42 repeats in the general population. We screened the SCA17 TBP gene in 197 normal controls and in patients with various neurodegenerative diseases: 13 ataxia, 103 Parkinson's disease, 122 dementia, 101 essential tremor, and 29 chorea and dystonia. The most common TBP allele contains 36 repeats and no expanded allele was found. Then biochemical and yeast assays were developed to screen effective chemical compounds which may inhibit polyglutamine protein aggregation. GST fused Qn, tTBP-Qn (truncated N-terminal TBP), nTBPQn (N-terminal TBP) and fTBPQn (full-length TBP) (n = 3 ~ 61) were overexpressed in E. coli BL21 cells and purified by affinity chromatography. After factor Xa cleavage to remove GST, the TBPQn proteins were used in drug screening by filter-trap assay and western blotting. However, neither controls (Congo red and trehalose) nor tested chemical compounds inhibit polyQ protein aggregation efficiently. This might be due to the low solubility of TBP and the suitability of 1C2 antibody. In addition, the expression of tTBPQ20/Q54-EGFP、nTBPQ20/Q48-EGFP under the control of the GAL1 promoter was induced by galactose in erg6 and W303 yeast strains. No growth inhibition was observed in yeast expressing full-length or N-terminal TBPQ54-EGFP proteins. This might be due to that low expression level was induced or the length of polyQ is not long enough to exhibit toxicity in yeast. Nevertheless, the study may provide information for future research design to develop assay to screen novel chemical inhibitors of aggregation.
Chan, Hoi-hung, and 陳海雄. "Establishment of an Orthotopic Hepatoma Model in Rats by Sono-guided Implantation for Preclinical Drugs Screening." Thesis, 2010. http://ndltd.ncl.edu.tw/handle/48793937082805998594.
Повний текст джерела國立中山大學
生物科學系研究所
99
Hepatocellular carcinoma (HCC) is one of the most prevalent cancers in the world and Taiwan. The major factors involved in the molecular pathogenesis for the development of HCC had been explored in recent years. An extensive array of growth factors and their receptors had been identified and may act as positive and negative modulators in different stages of hepatocarcinogenesis. Current therapeutic approaches for HCC include surgical resection (include liver transplantation), trans-arterial embolization (TAE), alcohol injection, etc. However, the effect is limited due to most of the HCC patients present with advanced stages of the disease. Therefore, this underscores the need for the development of novel therapeutic strategies. It is pivotal to set up an orthotopic hepatoma model for the development of novel intervention strategies for HCC. Under the guidance of ultrasound, we are able to create hepatoma in the liver lobe of Sprague-Dawley (SD) rats by injection of Novikoff (N1-S1) hepatoma cells. In addition, sonographic technique was employed for the monitoring of tumor growth in this animal model in the following subprojects. The continuous, non-invasive measurement of orthotopic hepatoma development will be a valuable tool for the evaluation of effects of drugs for treatment of HCC. In Chapter 1, the study employed a relatively non-invasive approach to establish an orthotopic HCC model in immune-competent rats. This was done by ultrasound-guided implantation of cancer cells and the model was used to evaluate the therapeutic efficacy of short-term and low-dose epirubicin chemotherapy. Ultrasound-guided implantation of Novikoff hepatoma cells led to the formation of orthotopic HCC in 60.4% of the SD rats. Moreover, tumor sizes measured by ultrasound significantly correlated with those measured by calipers after sacrificing the animals (P < 0.00001). The rate of tumor induction by ultrasound-guided implantation was comparable to that of laparotomy (55/91, 60.4% vs. 39/52, 75%) and no significant difference in sizes of tumor was noted between the two groups. Moreover, there was a significant correlation in tumor size measurement by ultrasound and computerized tomography. In tumor-bearing rats, short-term and low-dose epirubicin chemotherapy caused a significant reduction in tumor growth, and was found to be associated with enhanced apoptosis and attenuated proliferation as well as a decrease in microvessel density in tumors. In chapter 2, we investigated the chemopreventive effects of celecoxib in the growth of orthotopic rat HCC and the possible signal pathways involved. The status of COX-2 expression in rat Novikoff HCC was consistent with that in human HCC. Both Western blot and PCR tests had proved that N1-S1 was a HCC model presenting with low COX-2 enzymes in tumor cells. Then, low doses of celecoxib was shown to effectively inhibited the proliferation and increased the apoptosis of N1-S1 cells in vitro, which were also safe to the normal hepatocytes. Moreover, chemoprevention by celecoxib inhibiting the HCC tumor growth was shown in rat orthotropic HCC model. Tumor incidence was not affected by the celecoxib prevention, but, tumor weight was found significantly suppressed by the drug. Possible mechanisms of chemoprevention by celecoxib seen in the animal model were thought to be related to the anti-angiogenic, anti-proliferative and anti-hCSC characters of the drug. In chapter 3, we tried to test the combined inhibitory effects of low doses of celecoxib and epirubicin on the growth of HCC. Combined low doses of epirubicin and celecoxib was effective in inhibiting the hepatic cancer stem cells, tumor angiogenesis, tumor cell proliferation, as well as promoting cancer apoptosis. These are compatible with the effects of the individual drugs on HCC growth shown in the previous two chapters. In general, combination therapy expressed more effectiveness in tumor suppression and less bone marrow suppression than the individual drugs used alone. Taken together, ultrasound-guided implantation of Novikoff hepatoma cells is an effective means of establishing orthotopic HCC in SD rats, which is suitable and convenient for therapeutic trial of anti-HCC treatment. In the current study, we had proved the efficacies of low doses of two drugs, epirubicin and celecoxib, acting individually, as well as the combined effects of them in treating HCC in this model.
Antunes, Jéssica Alexandra Sá. "Manufacture of three-dimensional (3D) microcapsules through the use of multifunctional biomaterials." Master's thesis, 2018. http://hdl.handle.net/10773/25898.
Повний текст джерелаO cancro da próstata é um dos cancros mais diagnosticados e uma das principais causas de morte entre os homens a nível mundial. Atualmente, apesar de muitos avanços na medicina, os tratamentos desta neoplasia em estágio avançado são bastante ineficazes. O desenvolvimento de modelos in vitro que recapitulam os tumores da próstata humanos podem ajudar na descoberta de novas terapias e fármacos contribuindo assim para um aumento da expectativa de vida do paciente. Até à data, as agências reguladoras recomendam que o teste da eficácia de fármacos anti-tumorais a nível pré-clínico deve ser efetuado em culturas celulares bidimensionais (2D), no entanto, esses modelos não imitam as principais características dos tumores in vivo, tais como a sua distribuição espacial, interações célula-célula e gradientes de nutrientes/oxigénio. Além deste facto as culturas 2D não replicam os componentes da matriz extracelular tumoral (ECM) e a heterogeneidade celular tumoral. Estas limitações são responsáveis pela baixa correlação de resultados entre as culturas 2D in vitro e os dados obtidos em ensaios clínicos. Para superar essas questões, recentemente os modelos tumorais de cultura 3D in vitro têm sido investigados como alternativas valiosas. Estes modelos conseguem reproduzir vários aspetos do microambiente de tumores sólidos humanos, incluindo os seus padrões de expressão génica, interações 3D entre célula-célula, formação de núcleo necrótico e a intrínseca resistência aos fármacos. O trabalho de investigação desenvolvido no âmbito desta dissertação descreve a produção de um novo modelo tumoral 3D in vitro do cancro da próstata que mimetiza a heterogeneidade celular na metástase óssea do cancro da próstata bem como o microambiente da matriz extracelular. O modelo criado é composto por células humanas do cancro da próstata (PC-3) e osteoblastos humanos, encapsuladas em micro-hidrogéis com forma quasi-esférica. Estas microcápsulas, foram produzidas numa superfície quase super-hidrofóbica onde uma mistura de ácido hialurónico metacrilado, gelatina metacrilada, células cancerígenas e osteoblastos foram depositadas e reticuladas com luz U.V. Os resultados demonstram que os microtumores formados são reprodutíveis em termos de morfologia, tamanho e número de células encapsuladas. As formulações de co-cultura HA-MA / Gel-MA apresentaram deposição de cálcio ao fim de 14 dias, quando comparadas às monoculturas, evidenciando assim a importância dos osteoblastos. A avaliação da citotoxicidade da cisplatina nas co-culturas heterotípicas demonstrou que os microgéis 2.5%HA-MA-5%Gel-MA têm maior resistência ao fármaco que os microgéis com 5%HA-MA-5%Gel-MA. Em conclusão, os resultados indicam que as superfícies quase super-hidrofóbicas são úteis para a produção rápida, e sem solventes, de modelos 3D in vitro do cancro da próstata e podem vir a servir de plataforma de testes para a descoberta de novas terapias para o cancro da próstata
Mestrado em Materiais e Dispositivos Biomédicos
Monteiro, Cátia Filipa Rodrigues. "Platelet lysates-based hydrogels for the development of 3D models for bone cancer." Master's thesis, 2018. http://hdl.handle.net/10773/25632.
Повний текст джерелаO cancro é a segunda principal causa de morte a nível mundial e, relativamente ao cancro do osso, o osteossarcoma (OS) é o tumor maligno primário mais comum caracterizado pelo seu elevado potencial metastático, afetando predominantemente crianças e adolescentes. Apesar dos inúmeros esforços que visam o desenvolvimento de novas terapias anti-cancerígenas, vários candidatos a fármacos identificados como eficazes nos testes pré-clínicos falham durante os ensaios clínicos. Os sistemas de cultura de células tridimensionais (3D) têm sido propostos como plataformas in vitro fidedignas para o desenvolvimento de modelos tumorais na tentativa de reproduzir a patofisiologia tumoral e identificar terapias eficazes. Hidrogéis baseados em lisados de plaquetas humanos metacrilatados (PLMA) foram recentemente propostos como plataformas in vitro 3D economicamente viáveis e biologicamente relevantes para o crescimento e proliferação de células humanas. Assim, o objetivo deste trabalho foi, numa primeira abordagem, validar os hidrogéis de PLMA como plataformas 3D para suportar mecanismos de invasão de esferóides e, subsequentemente, explorar esse potencial para estabelecer modelos humanizados 3D de OS em mono- e co-cultura para teste e validação de fármacos. Esferóides de três linhas celulares tumorais (MG-63, SaOS-2 e A549) e células estaminais mesenquimais humanas derivadas da medula óssea (hBM-MSC) foram embebidos em hidrogéis de PLMA (três concentrações diferentes), Matrigel e poli(etileno glicol) diacrilatado. Os hidrogéis de PLMA demonstraram suportar a heterogeneidade fenotípica dos tumores sólidos e a aquisição de uma polaridade celular semelhante à in vivo. Além disso, estes hidrogéis recapitularam perfeitamente a capacidade de invasão celular, demonstrando que a velocidade de invasão pode ser facilmente controlada através da rigidez dos hidrogéis de PLMA. A co-cultura de esferóides de MG-63 com osteoblastos humanos e hBM-MSCs demonstrou que a comunicação entre as células tumorais invasivas e as células estromais foi fielmente recapitulada nos hidrogéis de PLMA. Um tratamento com doxorubicina nos modelos de OS em mono- e co-cultura claramente refletiu o papel protetivo das células estromais na quimioresistência em OS, exibindo uma resposta ao fármaco mais próxima da obtida in vivo. Globalmente, os resultados validaram os hidrogéis humanizados baseados em PLMA como plataformas in vitro 3D fidedignas para suportar uma morfologia e invasão tumoral semelhante à in vivo. Além disso, a complexidade do modelo de OS de co-cultura estabelecido forneceu um ambiente in vitro mais patofisiológico para o teste e validação de agentes anti-metastáticos de modo a prever a resposta do paciente e acelerar a disponibilidade de terapias efetivas.
Mestrado em Biotecnologia
Puscas, Ina. "Développement d’un modèle in vitro de la barrière hémato-encéphalique." Thesis, 2019. http://hdl.handle.net/1866/24000.
Повний текст джерелаThe blood-brain barrier (BBB), a central nervous system structure, is found in the cerebral capillaries. It represents a major obstacle for the drugs that have to reach the brain in order to exercise their pharmacological effect. In the early stages of the drug development, in vitro cell models are used to evaluate the brain permeability of new drugs. Models assembled using primary endothelial cells (ECs) isolated from mouse brain capillaries are of particular interest for research, as for their ease of obtaining and relevance for the drug screening. Thus, the goal of this project was to build and characterize a primary mouse monolayer model. At the same time, a murine b.End3 cell line monolayer model was investigated. The evaluation of these models was based on the TEER and fluorescent marker permeability values, as well as on the presence of the BBB hallmark proteins. The model validation was established by the correlation of the permeability data obtained with the in vitro model and the data obtained in mice (in vivo). As a result, the primary mouse model showed superior monolayer integrity and higher expression of the tight junction and membrane transporter proteins when compared with the bEnd.3 cell line model. The in vitro/in vivo correlation of the primary model resulted in r2 = 0.765 compared to the bEnd.3 model with r2 = 0.019. This research work shows that the primary monolayer mouse model is a simple and reliable model for predicting the drug permeability across the BBB.
Chuang, Kai-An, and 莊凱安. "Expression of Severe Acute Respiratory Syndrome-Coronavirus S193 in Sf9 Cells and Establishment of A Model for Screening Anti-Viral Drugs." Thesis, 2005. http://ndltd.ncl.edu.tw/handle/01525326612926972774.
Повний текст джерела國立陽明大學
醫學生物技術研究所
93
During 2002 and 2003, a newly discovered coronavirus, SARS-CoV, was contagious and spreaded rapidly in the world, resulting in 8098 people in 26 countries had probable case, 774 of whom have died. At that time, ribavirin, interferon, and corticosteroids were widely used for SARS therapy, but they were ineffective and had many side effects. Searching for an antiviral and potential drugs were needed urgently. The study indicates that angiotensin-converting enzyme 2 (ACE2) is the receptor for the SARS-CoV and the residues 318-510 (S193) of SARS-CoV spike (S) are receptor-binding domain (RBD). The aim of the thesis was to express the S193 protein in eukaryotic cells and establish a model for screening antiviral drugs. We utilize the SARS-CoV S gene to clone the full length S gene into the pAcSecG2T plasmid successfully. Due to the highly glycosylation of S proteins, we used bauculovirus virus expression vector system to express the S193. Using pAcSecG2T-abcd as a template to clone the RBD into the pAcHLT-A transfer vector was applied in protein expression. At present, recombinant baculoviruses had been produced and induced cytopathic effects in sf9 cells; moreover, a 27KDa of histidine tag S193 protein had been detected by Western blot assay. In this study, the insoluble forms of S193 contained in sf9 pellets were solubilized with 8M urea and the efficiency of purification was enhanced. Using 20-30 mM of imidazole could wash out most of the non-specific binding protein and gain more pure S193. In order to apply S193 for binding assay, expression of ACE2 in Vero E6 and Huh7 was detected by RT-PCR, Western blot assays, and immunofluorescence assay. Results of immunofluorescence assay demonstrated that the S193 could bind to the ACE2 expressing cells. Furthermore, The binding of S193 to ACE2 can be blocked by adding anti-ACE2 antibody and S193 bind to ACE2 can be seen by cell-based ELISA. In future, we will use this model which is essential to set up due to the consideration of laboratory safety, drugs development and pharmaceutical mechanisms to screen drugs.
Vaz, Rita Leitão Landeiro 1988. "Screening and validation of molecules with therapeutic properties in in vivo models of parkinson’s disease." Doctoral thesis, 2018. http://hdl.handle.net/10451/37407.
Повний текст джерелаPasquet, Vivian. "Characterization of thioredoxin and glutathione reductase activities of Mesocestoides vogae, a flatworm parasite useful as a laboratory model for the screening of drugs." Doctoral thesis, 2014. https://nbn-resolving.org/urn:nbn:de:bvb:20-opus-106759.
Повний текст джерелаCharakterisierung von Thioredoxin- und Glutathionreduktase Aktivitäten von Mesocestoides vogae, einem parasitären Plattwurm der als Labormodell für die Testung von Arzneistoffen verwendet werden kann
Lee, Ping-Hsung, and 李秉勳. "Expression and Purification of Severe Acute Respiratory Syndrome-Coronavirus Spike Protein S547 from Recombinant Baculovirus Infected Sf9 Cells and Establishment a Cell-Based Model for Anti-viral Drugs Screening." Thesis, 2006. http://ndltd.ncl.edu.tw/handle/07886033683850674782.
Повний текст джерела輔仁大學
生命科學系碩士班
94
In 2002, a new contagious disease had been found in Southern China and the disease spread widely and resulted in 8098 cases and 774 deaths. This disease was characterized as an atypical pneumonia symptom. The World Health Organization defined it as severe acute respiratory syndrome (SARS). The pathogen of SARS is a new coronavirus and named as SARS-coronavirus (SARS-CoV). Combination of ribavirin and corticosteroid is the most frequent antiviral treatment for SARS. However, the unobvious curative and annoying side effects for these medicines, it is urgent to find new anti-SARS drugs. SARS-CoV spreads mainly via respiratory routes and infects host cells by binding of viral spike protein to cellular angiotensin converting enzyme-2 (ACE2). The aim of thesis is to express the spike protein fragment and establish an anti-viral drug screening model. In the present study, I have obtained a recombinant baculovirus that carried a cDNA encoding SARS-CoV spike proteins from 16 a.a. to 547 a.a. Then a histidine Tag fusion protein (85 kDa), named as S547, was purified from recombinant baculoviruses infected sf9 cells by the Ni2+-NTA system. The results of reverse transcription-polymerase chain reaction (RT-PCR) and Western blotting indicated that ACE2 expression could be detected in Vero E6 cells. The data of immunofluorescent staining demonstrated that S547 proteins can bind to Vero E6 cells. Moreover, I have set up the cell-based ELISA to prove that S547 could bind to Vero E6 cells in a dose-dependant manner. In the future, the cell-based ELISA model will be applied for screening anti-viral drugs.
Mc, Innes Gabrielle. "Développement d’un nouveau modèle de criblage tridimensionnel pour la découverte de médicaments épigénétiques contre le cancer du poumon." Thesis, 2019. http://hdl.handle.net/1866/24521.
Повний текст джерелаSmall molecule development in oncology mainly involves high-throughput drug screenings and preclinical validation studies using cancer cells grown in two-dimension (2D). However, classical cell culture methods poorly reflect tumor biology and cell epigenome. Here, our objective is to develop a 3D model that displays key epigenetic features of solid tumors in order to identify new actionable targets. First, we determined culture conditions for long-term expansion of adenocarcinoma spheroids to allow cell adaptation and the occurrence of specific epigenetic features triggered by 3D condition. Our results demonstrate that cells cultivated in 3D spheroids exhibit significant phenotypic and epigenetic changes as compared to 2D monolayers. Notably, we observed numerous expression changes of key epigenetic regulators, all taken place at a different time-point of the 3D cell culture. We observed a decrease in the expression of the KAT3A/KAT3B complex as well as BRG1. HDAC6 expression also increased in 3D. Then, we asked whether epigenetic changes triggered by 3D culture would modify drug sensitivity. We performed a screening of 154 epigenetic drugs on cancer cells cultivated in 2D and in 3D at two different time points. 60% of epigenetic drugs showed significant anticancer activity against 2D monolayers. Interestingly, A549 cells in 3D spheroids became gradually resistant over time. Against 3D spheroids cultivated for 10 days, only 9% of epigenetic drugs in the drug library showed anticancer activity. Against 3D spheroids cultivated for 24 days, only a single epigenetic compound called MS023, a selective agent against type I PRMTs, reduced cell viability significantly. This sensitivity is correlated with an increase of arginine methylation observed within spheroids. Taken together, we show that 3D spheroids trigger a time-dependent epigenetic context that increases lung cancer cells sensitivity to type I PRMT inhibition. 3D spheroids of well-characterized cancer cell lines will improve our understanding of tumor biology and drug discovery and can overcome the high false discovery rate associated with 2D classical models.