Orientador: Antonio Wilson Sallum / Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Odontologia de Piracicaba / Made available in DSpace on 2018-08-26T23:52:25Z (GMT). No. of bitstreams: 1 Moura_LucasAlves_D.pdf: 4231687 bytes, checksum: ec30a84348074c7f0f52d37535f57bbe (MD5) Previous issue date: 2015 / Resumo: O objetivo deste estudo foi avaliar o efeito da associação da administração local de nanoesferas contendo doxiciclina (DOX) com o debridamento periodontal no tratamento da periodontite crônica avançada. As nanoesferas foram preparadas pelo método da dupla emulsão (W/O/W), e caracterizadas quanto à morfologia através da microscopia eletrônica de varredura (MEV), e quanto à interação polímero (PLGA) e DOX através da espectroscopia infravermelha da transformada de Fourier (FTIR). A avaliação da liberação controlada de DOX foi feita através da cromatografia líquida de alta eficiência (HPLC), a partir do fluido gengival sulcular (GCF) coletado nos períodos: 2, 5, 7, 10, 15 e 20 dias após aplicação das nanoesferas. Foi realizado um ensaio clínico randomizado controlado duplo cego, incluindo 30 pacientes diagnosticados com periodontite crônica que apresentavam sete sítios com sangramento à sondagem e profundidade de sondagem (PS) ? 5 mm, sendo 2 dentes com PS ? 7 mm. Os pacientes foram aleatoriamente divididos em 2 grupos e receberam os seguintes tratamentos: debridamento periodontal por 45 minutos associado à administração local de nanoesferas contendo doxiciclina nos sítios com PS ? 5 mm (DB+DOX) ou debridamento periodontal por 45 minutos associado à aplicação de nanoesferas vazias (DB). Foram avaliados os seguintes parâmetros clínicos: Índice de Placa (IP), Sangramento à Sondagem (SS), Profundidade de Sondagem (PS) e Nível de Inserção Clínica (NIC), no baseline, 3 e 6 meses após a terapia. A avaliação microbiológica foi feita por meio da reação de cadeia de polimerase ¿ tempo real ("real time" - PCR) para detecção das bactérias: Porphyromonas gingivalis, Tannerella forsythia, Treponema denticola, Aggregatibacter actinomycetemcomitans e Prevotella intermedia, provenientes do biofilme subgengival coletado no baseline, 1, 3 e 6 meses após o tratamento. Os resultados foram comparados por meio do teste de análise de variância com medidas repetidas, com nível de significância de 5%. Foram obtidas nanoesferas variando de 700 nm a 4 um. Através do FTIR observou-se boa interação da DOX com o PLGA, sem alterações das propriedades químicas de nenhum dos compostos. Após a aplicação das nanoesferas observou-se uma liberação de DOX constante no GCF até o vigésimo dia pós-tratamento, acima da concentração inibitória mínima. No grupo DB+DOX houve redução significativa da PS e ganho de inserção clínica nas bolsas profundas e moderadas em relação ao baseline e entre grupos. Observou-se redução significativa dos níveis bacterianos pós-tratamento, sendo o grupo DB+DOX mais eficaz em manter os níveis mais baixos após 6 meses do tratamento. Dentro das limitações deste trabalho, os resultados sugerem que as nanoesferas de PLGA são efetivas como sistema de liberação controlada local de DOX e quando associadas ao debridamento periodontal podem promover resultados superiores em relação à terapia mecânica isoladamente / Abstract: The aim of this study was to assess the effect of the association of local administration of nanospheres loading doxycycline (DOX) with the periodontal debridement treating advanced chronic periodontitis. Nanospheres were made by double-emulsion method (W/O/W) and characterized regarding morphology, by scaning eléctron microscopy (SEM); and drug/polymer interaction, by Fourier transform infrared spectroscopy (FTIR). The DOX controlled release was assessed by high performance liquid chromatography (HPLC) of gingival crevicular fluid (GCF) samples collected 2, 5, 7, 10, 15 and 20 days after treatment. It was performed randomized double-blinded clinical trial, with thirty patients diagnosed with chronic periodontitis presenting at least seven sites with bleeding on probing and probing depth (PD) ? 5 mm, and 2 sites with PD ? 7 mm. The patients were randomly allocated in two groups receiving interventions as followed: 45 minutes periodontal debridement + subgengival nanospheres loading DOX (DB+DOX); and 45 minutes periodontal debridement + subgengival void nanospheres (DB). Plaque index (IP), bleeding on probing (BP), probing depth (PD) and clinical attachment level (CAL) were evaluated on baseline, 3, and 6 months after therapy. Real-time polymerase chain reaction (real-time PCR) was used to quantify Porphyromonas gingivalis, Tannerella forsythia, Treponema denticola, Aggregatibacter actinomycetemcomitans and Prevotella intermedia from subgingival biofilm samples collected on baseline, 1, 3 and 6 months after treatment. The results were compared by variance analyses test for repeated measures, with significance of 5%. Nanospheres varying from 700 nm to 4 um were obtained. There was a good interaction between DOX and PLGA, with no chemical properties alterations. After local administration of the nanospheres, it was observed constant DOX release in the GCF until 20th day post-treatment, with concentration above the minimum inhibitory concentration. DB+DOX group showed significant PD reduction and CAL gain in deep and moderates pockets comparing to baseline and between groups. It was observed significant reduction of bacteria levels along follow up period, and DB+DOX group was more eficiente in keeping lower levels of bacteria after 6 months from treatment. Within limitations of this study, the results can suggest that PLGA nanospheres are effective carriers for controlled release of DOX and when used adjunctively to periodontal debridement, improved results can be achieved compared to mechanical therapy alone / Doutorado / Periodontia / Doutor em Clínica Odontológica

A incidência de casos de melanoma tem aumentado em todo o mundo sendo que, apesar do diagnóstico precoce e do advento das terapias moleculares, o número de pacientes que morrem com a doença em estágio avançado continua em elevação. Deste modo as pesquisas atuais têm focado no desenvolvimento de diferentes estratégias para a disponibilização de terapias eficazes e acessíveis. Nesse contexto, a via de administração transdérmica constitui uma alternativa promissora para aumentar a eficácia local e sistêmica de fármacos, incluindo agentes antitumorais. Diversos métodos têm sido desenvolvidos para maximizar a permeação cutânea de fármacos, destacando-se, entre esses, a ablação térmica por radiofrequência (RF). Esse processo resulta na criação de vários microcanais entre a epiderme e a derme, pelos quais diversas moléculas podem passar em direção às camadas mais profundas da pele. Nesse estudo, a eficácia da infusão transdérmica de etoposídeo por dispositivo de radiofrequência fracionada foi avaliada em modelo de melanoma murino. Camundongos da linhagem C56BL/6 foram divididos nos seguintes grupos experimentais: 1) controle; 2) tratados com radiofrequência; 3) tratados com a aplicação tópica de etoposídeo; e 4) tratados com radiofrequência e posterior aplicação tópica de etoposídeo. Os tratamentos foram realizados durante o período de 28 dias. O peso corpóreo, o volume tumoral e o perfil hematológico foram avaliados semanalmente. Ao término do tratamento os animais foram eutanasiados e procedeu-se a coleta da massa tumoral e dos órgãos (pulmão, baço, rins, linfonodos e fígado) para análise histopatológica. As células tumorais obtidas das massas tumorais foram analisadas quanto às alterações do ciclo celular e do potencial transmembrânico mitocondrial. Os resultados demonstraram que o tratamento com etoposídeo isolado reduziu a sobrevida dos animais e ocasionou alterações histológicas indicativas de toxicidade. Em contrapartida, a infusão transdérmica do etoposídeo por dispositivo de radiofrequência promoveu redução significativa do volume tumoral, em comparação com todos os grupos experimentais, sem ocasionar mortalidade. Esse tratamento também diminuiu a plaquetocitose e elevou o número de eritrócitos em comparação com os outros grupos. A análise histopatológica dos órgãos dos animais tratados com RF + etoposídeo evidenciou que não houveram alterações significativas na arquitetura tecidual. Ainda, o grupo tratado com RF + etoposídeo foi o que apresentou o maior percentual de células estacionadas na fase S/G2M e com mitocôndrias inativas, evidenciando o aumento da eficácia demonstrada no estudo in vivo. O conjunto de resultados sugere que o tratamento com a radiofrequência seguida do etoposídeo resulta em melhor resposta antitumoral do quimioterápico, com baixos índices de toxicidade sistêmica / The incidence of melanoma cases has increased worldwide and, despite early diagnosis and targeted molecular therapy, the number of patients dying from metastatic disease continues to rise. Thus, current research has focused on the development of different treatment strategies to provide efficient and accessible solutions. In this sense, transdermal delivery is a promising alternative enhancing the local and systemic efficacy of drugs, including antitumor agents. Several methods have been developed to improve the skin permeation of drugs, highlighting, among those, the radiofrequency thermal ablation (RFA). This process results in the creation of many microchannels between the epidermis and the dermis through which several molecules can pass towards the deeper layers of the skin. In this study, the efficacy of the transdermal delivery of etoposide by a fractional radiofrequency device was evaluated in a murine melanoma model. C56BL/6 lineage mice were divided into the following experimental groups: 1) control; 2) treated with radiofrequency; 3) treated with topical applications of etoposide; and 4) treated with radiofrequency followed by topical applications of etoposide. The animals were treated for 28 days and the body weight, tumor volume and hematological profile were analyzed weekly. At the end of the treatments, the animals were euthanized and the tumor mass and organs (lung, spleen, kidneys, lymph nodes and liver) were collected for histopathological analysis. Tumor cells obtained from the tumor masses were analyzed for changes in the cell cycle and mitochondrial transmembrane potential. The results showed that the treatment with etoposide alone reduced the survival of the animals and caused histological changes indicating toxicity. On the other hand, the transdermal delivery of etoposide by a radiofrequency device resulted in a significant reduction of the tumor volume, in comparison with all the experimental groups, not causing mortality. This treatment also decreased thrombocytosis and increased the number of red blood cells compared to the other groups. The histopathological analysis of the organs from animals treated with RFA + etoposide demonstrated that there was no significant change in tissue architecture. Furthermore, the group treated with RFA + etoposide presented the highest percentage of cells with inactive mitochondria and interruption at the S/G2M stage, corroborating the increased efficacy of the in vivo study. The set of results indicates that the treatment with radiofrequency followed by etoposide results in better antitumor responses of chemotherapy, with low toxicity rates

Thèse réalisée dans le cadre d'une collaboration avec UCB Pharma et la région Wallonne s'inscrivant dans le cadre du projet SAS. Le but de ce projet était de développer des implants sous-cutanés imprimés en trois dimensions pour permettre une libération d’anticorps thérapeutique de manière prolongée au cours du temps. En effet, les thérapies disponibles sont souvent administrées par voie intraveineuse, ce qui peut réduire la compliance des patients dû à l’inconfort et à la fréquence de ces administrations. Les systèmes de délivrance, tels que des implants, peuvent limiter les fréquences d’administration grâce à l’insertion d’un dispositif qui libèrera le principe actif au cours du temps durant une période donnée. Les implants s’inscrivent comme une alternative aux microsphères qui sont également des dispositifs développés et investigués en vue de favoriser l’adhésion et la compliance des patients. L’avènement du 3D dans le milieu pharmaceutique a montré une certaine frénésie liée au développement de la médecine personnalisée et à l’innovation du procédé dans ce secteur. La sélection d’un matériau biocompatible et biorésorbable tel que le PLGA représente une véritable plus-value dans le développement d’implant. Etant donné que ces implants sont biodégradables, le retrait n’est pas à envisager, ce qui limite les désagréments du patient à un seul acte chirurgical lors de l’implantation. Au cours de ce travail, une approche pragmatique a d’abord été abordée sur les procédés d’extrusion à chaud et de l’impression 3D en utilisant un polymère couramment employé dans l’impression grand public, le PLA. L’investigation des paramètres d’impressions (température d’impression, epaisseur de couche et vitesse d’impression) et l’usage de divers plastifiants (la triacétine (TA), le polyethylène glycol 400 (PEG 400), le citrate de triéthyle (TEC) et l’acétyle citrate de triéthyle (ATEC)) pour faciliter les procédés à chaud et dans l’idée de réduire les températures d’extrusion et d’impression du matériau ont été évalués. Ces essais ont démontré l’effet de la température d’impression sur la qualité de l’impression et principalement sur les propriétés du matériau comme la force de traction et la ductilité. De plus, l’ajout de plastifiant à la matrice du PLA a permis de diminuer sa température de transition vitreuse. Par exemple, la température de transition vitreuse du PLA a été diminuée de 53 °C à 34 °C par l’ajout de PEG 400. Cette approche avait pour but d’évaluer la possibilité de diminuer les températures d’impression dans l’optique d’encapsuler à chaud un anticorps sensible à la chaleur pour la suite de ce travail.Ensuite, le développement de filaments imprimables contenant des anticorps a été abordé et mis en place à l’aide d’un modèle d’anticorps polyclonal disponible en grandes quantités et à des coûts relativement faibles. Un anticorps à l’état solide a été favorisé dans le procédé car il est largement accepté que les protéines sous forme solide sont plus stables au cours du temps en comparaison aux solutions d’anticorps. De plus, cet état solide facilite les manipulations précédant l’extrusion comme l’étape de mélange. Pour la réalisation des filaments, différents types de PLGA ont été investigués afin d’atteindre les propriétés nécessaires à l’impression en termes de diamètre mais également de comportement physique. Ces dérivés étaient caractérisés par des masses moléculaires différentes comme pour le PDLG5004 (44 kDa), le RG502 (7-17 kDa) et parmis eux, un copolymère PEG-PLGA (2 kDa-20 kDa). Un PLGA de faible masse moléculaire a été sélectionné pour développer ce filament. En effet, les extrusions étaient réalisables à une température maximum de 90 °C et les impressions à 113 °C minimum. L’un des enjeux cruciaux du développement de filament imprimable contenant un anticorps à haute concentration, au minimum 15% (w/w), était d’en assurer l’homogénéité. Cependant, l’usage de températures aussi élevées lors de l’impression a induit la dégradation de l’anticorps par la formation d’agrégats et principalement de fragments. Ces derniers sont généralement produits lors de procédé à haute température ou par l’usage de conditions drastiques telles que l’acidification du milieu. Cette plateforme a été adaptée à l’encapsulation d’anticorps thérapeutique fournit par UCB Pharma. L’usage d’un anticorps monoclonal possédant une stabilité supérieure à celle du modèle initialement utilisé permettrait d’identifier l’impact du procédé sur l’intégrité de l’anticorps. La formulation de l’anticorps a été réalisée en utilisant différents stabilisants conventionnels (sucrose (Suc), trehalose (Tre), 2-Hydroxypropyl-beta-cyclodextrine (HP-β-CD), inuline (Inu) et sorbitol (Sor)) et reconnus pour la stabilisation des protéines. A côté des excipients ajoutés, différentes quantités d’excipients ont été investigués. Ces manipulations ont montré que la stabilité de l’anticorps était privilégiée à l’aide du sucrose et du tréhalose à un ratio anticorps monoclonal:excipient de 2.0:1. En gardant ce ratio, l’ajout d’un acide aminé (leucine) aux deux disaccharides précédemment cités, a amélioré la stabilité de l’anticorps vis-à-vis des procédés à chaud (extrusion et impression 3D). L’homogénéité au sein des filaments imprimables et des pièces 3D a été confirmée tout au long du procédé. En effet, les charges en anticorps étaient similaires à la charge théorique de 15% (w/w). Aucune fragmentation de l’anticorps n’a été observée à l’issue des procédés à chaud. Cependant, une augmentation des agrégats de 2.6% en solution à 3.6% après impression 3D a été constatée à la fin du processus. Après avoir stabilisé l’anticorps, le but premier étant d’en promouvoir une libération prolongée au cours du temps. Les profils ont révélé une libération en trois phases au cours du temps mais avec un relargage après 24h relativement faible (< 5%) dû à la densité des matrices polymériques. Ensuite, la dégradation du polymère représente l’élément limitant la libération de l’anticorps au cours du temps. En effet, l’érosion du polymère joue un rôle clé dans la libération de l’anticorps encapsulé. La libération au cours du temps a été démontrée sur une période allant jusqu’à 15 semaines. La stabilité de l’anticorps dans le milieu de dissolution a été évaluée et une dégradation de celui-ci au cours du temps a été observée. Cette dégradation est principalement liée à l’érosion du polymère et à l’acidification du milieu au cours du test de dissolution. Après avoir optimisé la formulation de l’anticorps et avoir démontré la libération prolongée de celui-ci, son affinité restait à être étudiée. La capacité de l’anticorps à se lier à sa cible a pu être démontrée après 24h de dissolution mais cette affinité s’est réduite au cours de la durée de la dissolution avec une augmentation de l’agrégation et de la fragmentation de l’anticorps. Une étude de stabilité a également démontré que les implants imprimés en 3D sont stables à une température 5 °C sur une durée de 6 mois. Aucun élément de dégradation n’a été observé au cours du temps et l’affinité de l’anticorps a été préservée au cours de l’étude. Finalement, cette plateforme a également été évaluée pour l’encapsulation d’une troisième molécule biologique, un fragment d’anticorps monoclonal, pour d’une part en estimer la stabilité et l’applicabilité et d’autre part envisager une prochaine étude pré-clinique sur rongeurs. Le fragment d’anticorps a montré une stabilité supérieure à celle de l’anticorps monoclonal avec une faible agrégation après l’extrusion et l’impression. La libération prolongée du fragment a été évaluée sur 8 semaines et une libération du fragment de 79% a été observée avec une formulation contenant du tréhalose et de la leucine. En effet, les fragments d’anticorps ont une demi-vie plasmatique relativement faible, de l’ordre de 28 minutes, ce qui donne tout son sens à des systèmes à libération prolongée. Pour finir, la réalisation d’une étude pré-clinique permettrait de valider le modèle. En conclusion, ce travail a démontré la faisabilité de l’usage de l’impression 3D en vue de développer des systèmes à libération prolongée contenant des anticorps et en utilisant des procédés à hautes températures. Ces implants ont été caractérisés par une stabilité favorable et une libération intéressante qui feront l’objet d’investigation lors d’études pharmacocinétiques. / Doctorat en Sciences biomédicales et pharmaceutiques (Pharmacie) / info:eu-repo/semantics/nonPublished

First part of this thesis was focused on the previously overlooked field of C1'-branched acyclic nucleoside phosphonates (ANPs). Five diverse synthetic approaches were developed/optimized affording key 6-chloropurine intermediates bearing N9 -phosphonomethoxyethyl (PME) branched at C1' position in 2-4 steps. It was demonstrated that these intermediates can be further vastly diversified into ANPs bearing both natural and unnatural nucleobases. Single enantiomers as well as racemates of final C1'-branched ANPs (overall 48 final compounds) were prepared and selected compounds were evaluated with respect to their biological properties. The aforementioned ANPs showed no antiviral potency against studied viruses and only weak to moderate cytostatic activity. Adenine C1'-branched ANPs proved to be the most potent currently known inhibitors of Trypanosoma brucei adenine phosphoribosyl transferase (TbrAPRT), an enzyme involved in purine salvage pathway (PSP) of T. brucei. Further biological evaluation of prepared compounds is in progress. Second part of this thesis was focused on development of novel prodrug moieties with higher selectivity index (i.e. toxicity/potency ratio - SI) based on so-called ProTide prodrugs where phenol (present in ProTides) was replaced by tyrosine derivatives. Tenofovir was...

In light of widespread digitization endeavors and ever-growing textual data generation, developing efficient academic Natural Language Processing (NLP) infrastructures, which can deal with large amounts of data, is of particular importance. Novel computation technologies allow tools that support big data and heavy computation while performing timely and cost-effective data processing. This development has led researchers to demand that knowledge be extracted from ever-increasing textual data before it is outdated. Cluster computation is a modern technology for handling big data efficiently. It provides distribution of computing and data over a number of machines in a cluster, as well as efficient use of resources, which are key requirements to process big data in a timely manner. It also assures applications’ high availability and fault tolerance, which are fundamental concerns when dealing with vast amounts of data. In addition, it provides load balancing of data during the execution of tasks, which results in optimal use of resources and enhances efficiency. Data-oriented parallelization is an effective solution to enable the currently available academic NLP infrastructures to process big data. This approach offers a solution to parallelize the NLP tools which comprise identical non-complicated tasks without the expense of changing NLP algorithms. This thesis presents the adaption of cluster computation technology to academic NLP infrastructures to address the notable features that are essential to process vast quantities of text materials efficiently, in terms of both resources and time. Apache Spark on top of Apache Hadoop and its ecosystem have been utilized to develop a set of NLP tools that provide a distributed environment to execute the NLP tasks. Many experiments were conducted to assess the functionality of the designated strategy. This thesis shows that using cluster computation technology and data-oriented parallelization enables academic NLP infrastructures to execute large amounts of textual data in a timely manner while improving the performance of the NLP tools. Moreover, these experiments provide information that brings a more realistic and transparent estimation of workflows’ costs (required hardware resources) and execution time, along with the fastest, optimum, or feasible resource configuration for each individual workflow. This knowledge can be employed by users to trade-off between run-time, size of data, and hardware, and it enables them to design a strategy for data storage, duration of data retention, and delivery time. This has the potential to enhance researchers’ satisfaction when using academic NLP infrastructures. The thesis also shows that a cluster computation approach provides the capacity to adapt NLP services with JIT delivery systems. The proposed strategy assures the reliability and predictability of the services, which are the main characteristics of the services in JIT delivery systems. Defining the relevant parameters, recording the behavior of the services, and analyzing the generated data resulted in the provision of knowledge that can be utilized to create a service catalog—a fundamental requirement for the services in JIT delivery systems—for each service offered. This knowledge also helps to generate the performance profiles for each item mentioned in the service catalog and to update them continuously to cover new experiments and improve service quality.

The water solubility of drug molecules plays an important role in consideration of formulation development to treat a wide range of diseases. In this project, two kinds of drug delivery systems, cyclodextrins and liposomes, were developed for insoluble drug delivery to treat Alzheimer's disease (AD) and colorectal cancer (CRC), respectively. AD is an irreversible neurodegenerative disorder associated with the accumulation of amyloid-beta (A??) fibrils. Approximately 10% of people aged 65 and above have AD. Crocetin (CRT) is an active compound isolated from the fruits of gardenia (Gardenia jasminoides Ellis) and the stigmas of saffron (Crocus sativus L.). It has been reported to show various neuroprotective activities. However, poor water solubility and bioavailability are the major obstacles in developing pharmaceutical formulations of CRT. To address the issues, CRT liposomal formulations and CRT-cyclodextrin inclusion complexes were developed and evaluated. CRT-cyclodextrin inclusion complexes significantly increased the water solubility of CRT from the range ??g/mL to mg/mL. The CRT-??-cyclodextrin inclusion complex (1:3 molar ratio of CRT/??-cyclodextrin) was chosen for further studies as it showed the highest encapsulation efficiency (94.73 ?? 0.86%). The formulation had no toxicity to neuronal cells nor AD model cells within the experimental concentration range (0.625 to 100 ??M of CRT). It could downregulate the expression of C-terminus fragments and decrease both intracellular and extracellular levels of A??, which are hallmarks of AD. It also showed dose-dependent neuroprotective and antioxidant effects against H2O2-induced cell death. Pharmacokinetics and biodistribution studies showed that this CRT-??-cyclodextrin inclusion complex was suitable for intravenous administration. The formulation significantly increased the bioavailability of CRT and facilitated CRT crossing the blood-brain barrier to enter the brain. Similar to AD, CRC is increasingly prevalent with aging populations. Approximately 60% of CRC patients are aged 70 and above. Halofuginone (HF) is an active pharmaceutical ingredient (API) originated from Chinese quinine (Dichroa febrifuga Lour.) and artesunate (ART) is a semi-synthetic derivative of artemisinin (ATS) extracted from annual wormwood (Artemisia annua L.). Both APIs show anticancer activities by inhibiting the growth of CRC. However, low aqueous stability limits their applications. Liposome formulation with surface functionalization by CPP2 cell-penetrating peptide was developed to deliver HF and ART for targeted CRC therapy. CPP2 is a peptide that can selectively penetrate colon cancer cells. The liposomal drug formulations had uniform particle size (about 100 nm), high encapsulation efficiency (over 80%) and good stability upon 14 days of storage. In cellular uptake study, CPP2-modified liposome showed stronger permeability and selectivity to colon cancer lines without inducing lysosomal degradation. CPP2 surface-modified liposomal drugs demonstrated greater anticancer activities than free form of drugs or conventional liposomal drugs. Combinations of HF and ART formulations notably decreased cancer cell viability as compared to single formulation alone, which indicated that HF and ART formulations exhibited synergistic anticancer effects at specific ratios. To conclude, the drug delivery systems, cyclodextrins and peptide-modified liposomes, which were developed for AD and CRC treatment, successfully improved the aqueous solubility of insoluble APIs extracted from Chinese medicinal plants.

La thèse est intitulée « Modifications chimiques de polysaccharides pour l'obtention de nouveaux systèmes polymère-principe actif ». Le thème de la recherche porte sur trois axes principaux, dont l’objectif majeur est la synthèse et la caractérisation de nouveaux systèmes pour la libération contrôlée de principes actifs. Le premier chapitre traite de l’optimisation de la méthode pour préparer des émulsions stables simples inverses (eau/huile) et multiples (eau/huile/eau) à base de polysaccharides (amidon et 2-hydroxyethyle cellulose). On a étudié le processus de relargage de la caféine, l’influence de la présence d’amidon dans la phase aqueuse sur la stabilité et cinétique de libérations. Le second chapitre a pour objet l’obtention de nouveaux principes actifs à partir d’aminoacides et leur immobilisation sur polysaccharides par couplage chimique covalent. Les conjugues polymère-médicament obtenus ont été caractérisés par spectroscopie, physico-chimie. Leur activité antimicrobienne, leur toxicité in vivo, l’efficacité de l’immobilisation et la cinétique de relargage in vitro de médicaments couplés ont été étudiées. Le dernier chapitre traite de la synthèse des dérivés de chitosane avec l’anhydride maléique et l’acide acrylique, ainsi que leur utilisation dans la préparation d’hydrogels sous forme de films et de nanoparticules par copolymérisation avec le méthacrylate de 2-hydroxyethyle et l’acrylamide de N- isopropyl. Les dérivés ont été évalués par leur degré de fonctionnalisation. Les nanosystèmes ont été caractérisés par leur potentiel zêta, leur capacité de gonflement, d’adhésion et de relargage in vitro. Tous les résultats expérimentaux ont été soutenus par importantes études bibliographiques / The thesis is entitled “Chemical modifications of polysaccharides for the obtaining of new drug-polymer systems“. The topic of research includes four main working axes, diverted from the major objective to synthesize and characterize new drug delivery systems. The first chapter treats the optimization of the preparation method of stable simple W/O emulsions and multiple W/O/W emulsions, based on polysaccharides (starch and 2-hydroxyethyle cellulose). We also studied the process of caffeine release outside the multiple emulsions particles and the influence of starch in the internal aqueous phase stage upon the stability and the drug release kinetics. The second chapter studies the obtaining of new active principles from aminoacid and their immobilization on polysaccharides by covalent coupling. The new drug-polymer conjugates were characterized by spectral, physicochemical properties, antimicrobial activity, in vivo toxicity, immobilization efficiency and I vitro drug release kinetics. The last chapter treats the synthesis of the chitosan derivatives with maleic anhydride or acrylic acid, and their use in the preparation of hydrogels as film and nanoparticles by copolymerization with 2-hydroxymethacrilate or N-isopropyl acrylamide. The derivatives were evaluated by determining the degree of functionalization and by spectral methods. The nanosystems were characterized by measurements of zeta potential, swelling and adhesion capacity and membership and in vitro drug release kinetics. All experimental results were supported by important bibliographic studies

Current chemotherapeutics pose many di sadvantages due to their lack of specificity and low therapeutic index. To overcome these challenges, research has focused its attention on the development of nano-based delivery systems that can penetrate the leaky vasculature of tumor endothelium, use site-directed ligands that can bind with high affinity and specific ity to tumor cells, physically entrap poorly soluble drugs, and deliver these cytotoxic agents directly to the tumor site. One approach to nanosystem drug delivery is with the use of peptide amphiphiles (PAs) that are conjugated with the Arginine-Glycine-Aspartic Acid (RGD) motif to actively target a αVβ3 integrin receptors on cancer cells or tumor endothelium. The current work is focused on mechanistic studies to evaluate the uptake of novel RGD amphiphi les with varying alkyl chain lengths (palmitic acid : Cl 6 and stearic acid: C 18) and hydrophilic linkers, 8-amino- 3,6-dioxaoctonoic acid (ADA) or glucose, as micellar delivery systems of hydrophobic anticancer agents. PAs were confirmed for their self-assembling properties and further evaluated for their RGD-mediated binding specificity to purified αVβ3 integrin through a competitive binding fluorescence polarization assay (with novel RGD micelles displacing an integrin-bound fluorescent RGD probe by as much as 63.03%). Ultimately, these nanocarriers were assessed for their ability to deliver phys ically entrapped fluorescein isoth iocyanate (FITC) to A2058 cells overexpressing αVβ3 integrin receptors. Results from confocal microscopy indicate that uptake of RGD micelles was driven by an energy-dependent mechanism, as statistically significant levels of FITC internalization was seen at 37°C versus 4°C (p-value<0.05 for all treatment groups); moreover, intracellular fluorescence was notably higher (as much as 4-fold) when delivered through novel RGD conjugates as opposed to its free form. Regardless of chain length and the number of hydrophilic linkers, all RGD PAs showed promising results as micellar carriers that can effectively deliver their payload to the target tumor site via receptor mediated endocytosis.

京都大学 / 0048 / 新制・課程博士 / 博士(工学) / 甲第18573号 / 工博第3934号 / 新制||工||1604(附属図書館) / 31473 / 京都大学大学院工学研究科都市社会工学専攻 / (主査)教授 谷口 栄一, 准教授 宇野 伸宏, 准教授 QURESHIAli Gul / 学位規則第4条第1項該当 / Doctor of Philosophy (Engineering) / Kyoto University / DFAM

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