Tesis por compendio / [ES] La presente tesis doctoral titulada "Sistemas de liberacio'n controlada de fa'rmacos diseñados para mejorar el tratamiento de Enfermedad Inflamatoria Intestinal" se centra en el diseño, preparación, caracterización y evaluación in vivo de distintos sistemas de liberación controlada de fármacos en colon (CDDS, por sus siglas en inglés) utilizando como soporte micropartículas de silice mesoporosa, funcionalizadas con puertas moleculares. En conclusión, los estudios realizados demuestran que los materiales de silice mesoporosa, en combinación con puertas moleculares sensibles a estímulos específicos, tienen un gran potencial para el desarrollo de nuevos sistemas de liberación controlada de fármacos en el colon, dirigidos a mejorar el arsenal terapéutico disponible para el tratamiento de EII. La posibilidad de adaptar o personalizar la carga y las puertas moleculares hace que estos soportes de sílice mesoporosa sean una opción interesante para el desarrollo de nuevos sistemas de liberación controlada de fármacos en diferentes aplicaciones biomédicas. Finalmente, esperamos que los resultados obtenidos en esta tesis doctoral sirvan de inspiración para el desarrollo de sistemas de liberación controlada de fármacos innovadores y cada vez más inteligentes, para su aplicación tanto en medicina como en otras áreas. / [CA] La present tesi doctoral titulada "Sistemes d'alliberament controlat de farmacs dissenyats per a millorar el tractament de Malaltia Inflamatoria Intestinal" se centra en el disseny, preparacio, caracteritzacio i avaluacio in vivo de diferents sistemes d'alliberament controlat de farmacs en colon (*CDDS, per les seues sigles en angles) utilitzant com a suport microparticules de si'lice mesoporosa, funcionalitzades amb portes moleculars. En conclusio, els estudis realitzats demostren que els materials de si'lice mesoporosa, en combinacio amb portes moleculars sensibles a estimuls especifics, tenen un gran potencial per al desenvolupament de nous sistemes d'alliberament controlat de farmacs en el colon, dirigits a millorar l'arsenal terapeutic disponible per al tractament de MII. La possibilitat d'adaptar o personalitzar la carrega i les portes moleculars, fa que aquests suports de silice mesoporosa siguen una opcio interessant per al desenvolupament de nous sistemes d'alliberacio controlada de farmacs en diferents aplicacions biomediques. Finalment, esperem que els resultats obtinguts en aquesta tesi doctoral servisquen d'inspiracio per al desenvolupament de sistemes d'alliberament controlat de farmacs innovadors i cada vegada mes intel·ligents, per a la seua aplicacio tant en medicina com en altres arees. / [EN] This PhD thesis entitled "Smart drug delivery systems designed to improve Inflammatory Bowel Disease therapy" is focused on the design, synthesis, characterization and in vivo evaluation of several Colon Drug Delivery Systems (CDDS) using hybrid mesoporous silica microparticles as scaffolds containing molecular gates. In conclusion, the studies shown in this Thesis demonstrate that mesoporous silica materials in combination with responsive molecular gates have great potential in the design and preparation of new CDDS to improve the therapeutic options available for IBD. The possibility to adapt the cargo and the molecular gate makes mesoporous silica support especially appealing for similar controlled drug delivery applications in the biomedical field. We hope that the obtained results could inspire the development of new innovative smart drug delivery systems in this or other fields. / We thank the Spanish Government (projects MAT2015-64139-C4-1-R and AGL2015-70235-C2-2-R (MINECO/FEDER)) and the Generalitat Valenciana (project PROMETEOII/2014/047) for support. AHT thanks to the Spanish MEC for his FPU grant. We thank the Generalitat Valenciana (Project PROMETEO2018/024) / Hernández Teruel, A. (2019). Smart drug delivery systems designed to improve Inflammatory Bowel Disease therapy [Tesis doctoral]. Universitat Politècnica de València. https://doi.org/10.4995/Thesis/10251/129863 / Compendio

Yes / Understanding the structure-property relationships of drug delivery system (DDS) components is critical for their development and the prediction of bodily performance. This study investigates the effects of introducing polyethylene oxide (PEO) homopolymers, over a wide range of molecular weights, into Pluronic injectable smart hydrogel formulations. These smart DDSs promise to enhance patient compliance, reduce adverse effects and dosing frequency. Pharmaceutically, Pluronic systems are attractive due to their unique sol-gel phase transition in the body, biocompatibility, safety and ease of injectability as solutions before transforming into gel matrices at body temperature. This paper presents a systematic and comprehensive evaluation of gelation and the interplay of microscopic and macroscopic properties under both equilibrium and non-equilibrium conditions in controlled environments, as measured by rheology in conjunction with time-resolved Small Angle Neutron Scattering (SANS). The non-equilibrium conditions investigated in this work offer a better understanding of the two polymeric systems’ complex interactions affecting the matrix thermo-rheological behaviour and structure and therefore the future release of an active pharmaceutical ingredient from the injectable DDS.

Future changes in the incidence and prevalence of OA are difficult to predict. As incidence and prevalence rise with increasing age, extending life expectancy will result in greater numbers with OA. Actually, usual therapeutic approaches are really restricted because of important side effects with long-term use. Therefore, there is a need to develop improved formulations which are well tolerated, biocompatible and biodegradable. Ideally, these new treatments should be able to deliver locally sufficient amount of anti-inflammatory or analgesic drugs into the site of arthritic inflammation while stabilizing or better restoring the mechanical integrity of the joint. In this way, the objective of this project is to develop slow-release gels that are sterile, injectable, characterized by viscoelastic properties and capable to sustain the in situ release of both hydrophilic and lipophilic drugs. The intraarticular delivery combined to sustained-release property should be interesting to reduce the number of injection required while prolonging the local drug activity over weeks. For that purpose, glycerol monooleate (GMO), also called “monolein” was selected for its capacity to form highly viscous crystalline phase structures upon contact with an aqueous fluid (e.g. synovial fluid). <p>In the first step of this work, it was decided to develop and characterize hydro-lipidic gels based on the use of monolein and hyaluronic acid in order to provide in vitro sustained release of hydrophilic drugs such as clonidine and lipophilic drugs such as betamethasone. Initially, a compatibility study was performed on the main ingredients selected in order to check that there were not physico-chemical incompatibilities, which could be deleterious regarding to their stability in formulation. Then, the development of hydro-lipidic gels was initiated by considering on the first hand the solubility of each ingredient and on the other hand the syringeability, the rheological properties and the in vitro dissolution profiles obtained for the developed formulations. The objective of this preformulation program was to identify potential candidates that presented suitable syringeability while being able to sustain the release of drugs over weeks and being characterized by interesting viscoelastic properties for the long-term management of osteoarthritis. Moreover, several methods of quantification and characterization were developed in order to allow the physico-chemical properties (rheology, syringeability, water uptake, stability and dissolution profiles) of the developed formulations to be studied.<p>Results of the compatibility study showed that the concomitant use of monolein, hyaluronic acid and clonidine/betamethasone is not contraindicated. Next, the preformulation program allowed many injectable drug delivery systems to be prepared. However, the carrier that best meets our needs was composed of 10,0 % (wt/wt) absolute ethanol ;15,0 % propylene glycol (wt/wt) ;15,0 % (wt/wt) water ;55,0 % (wt/wt) de monolein ;5,0 % (wt/wt) purified soybean oil ;0,03 % (wt/wt) α-tocophérol and 7,5 mg/g sodium hyaluronate (1.9 MDa). This carrier assured suitable syringeability and rheological properties. Indeed, it presented marked pseudoplastic flow behavior that allowed relatively fast injection through a narrow needle, followed by an increase in viscosity upon contact with aqueous fluids to obtain an in vitro sustained release of hydrophilic and lipophilic drugs over a few weeks. As a consequence, it was assumed that this carrier should be able to jellify in situ upon contact with physiological fluid such as synovial fluid. Then, according to EMA recommendations, a fast and easy manufacturing process that could be applied in a cleanroom at industrial scale was validated in our Laboratory. Finally, according to these promising results obtained in vitro, a stability study was performed on the carrier alone and containing clonidine or betamethasone according to ICH recommendations described for products intended for storage in a refrigerator. In that purpose, several parameters such as the quantification of drugs, the pH, the molecular weight of hyaluronic acid, the dissolution profiles of drugs and the rheological properties of the formulations were recorded depending on time and conditions of storage. This stability study showed clearly the importance to adjust the pH value of the formulation. Indeed, it was demonstrated that a pH value of 6.5, adjusted with diluted NaOH, allowed the stability of the formulation to be significantly improved. During this first step of this project, our Laboratory initiated two new collaborations. On the first hand, collaboration with the Laboratory of Professor Siepmann (University of Lille 2 – Faculty of Pharmacy) was started for their expertise on mathematical modeling. On the other hand, collaboration with the Laboratory of Professor Jerôme (ULg – Faculty of sciences) was started for their expertise on macromolecular chemistry and more particularly on rheological properties.<p>In the second step of this work, it was decided to evaluate in vitro the safety and the efficiency of the developed carrier and formulations containing clonidine or betamethasone. In this way, it was suggested to test selected drugs and potential candidates formulations on equine polymorphonuclear leukocytes (PMN) by measuring the production of reactive oxygen species (ROS) by PMNs stimulated or not with phorbol 12-myristate 13-acetate (PMA). For that purpose, our Laboratory initiated a new collaboration with the Laboratory of Professor Serteyn (ULg – Faculty of veterinary) for their expertise on equine PMNs and quantification of (ROS) produced in particular in inflammatory diseases.<p>This in vitro study has shown that no pro-inflammatory effect appeared by incubating carrier with unstimulated PMNs in comparison with the control assay. However, the production of ROS was quickly and considerably decreased when stimulated cells were placed in contact with carrier regardless on the incorporation of clonidine or betamethasone. This observation demonstrated that developed carrier provided a strong antioxidant effect, certainly by trapping the ROS produced. These results were very promising because that antioxidant effect of carrier could inhibit oxidative damages and might consequently potentiate the prevention of inflammatory conditions. Concerning the clonidine and betamethasone, only the last one provided significant inhibition of the ROS activity.<p>Finally, by considering the very promising results obtained with the in vitro study on PMNs, an in vivo study on rabbits, which seemed to be the most appropriate small animal model for this kind of intraarticular formulations, was performed to evaluate the toxicity and the efficiency of the developed carrier and formulation containing betamethasone. Therefore, our Laboratory started collaboration with the unit of research in osteo-articular pathologies (UROC) of Pr. Henrotin (ULg) for their expertise in animal models, in particular rabbits with osteoarticular pathologies such as osteoarthritis. For this purpose, this in vivo study was outsourced by TNO (Delft, Holland) and was designed as follow: (i) 0.9 % saline buffered (n=8); (ii) carrier (n=8); (iii) formulation containing betamethasone (n=8); (iv) Durolane® (n=8) a marketed product of HA. Surprisingly, it seemed that the control group (saline buffered) presented macroscopical and histological scores that were globally low according to literature. As a consequence, it was difficult to conclude about the efficiency of the developed treatments by considering only this pilot study. However, it is important to note that it seemed that the expected viscoelastic protection of the carrier to prevent the degradation of articular cartilage was not optimal regardless on the incorporation of betamethasone. Nevertheless, the histological analyses of synovial membranes from each treated groups demonstrated that there was no pro-inflammatory reaction. This meant that all formulations tested were well tolerated despite of the apparition of lumps (in 37.5 % of treated rabbits) that are probably due to both the high volume injected (900 µL) and an excessive and unexpected in situ water uptake of developed formulations based on GMO. However, this lack of rejection of the developed carrier could be very important since it allowed new perspectives to be considered. For example, other articular disorders could be targeted by incorporating drugs, for which in situ sustained release or mechanical protection could be beneficial. <p>Our laboratory is member of a collaborative project "JOINT-AIC" from BioWin and is supported by a grant from the Walloon Region. The development of analytical methods, the evaluation of physico-chemical properties and finally the preparation of sterile batches of formulations based on GMO intended for in vitro and in vivo studies were performed in the Laboratory of Galenic and Biopharmacy of the Faculty of Pharmacy of ULB./L’arthrose est une pathologie dont la prévalence et le coût ne font qu’augmenter dans notre société vieillissante. Les moyens thérapeutiques actuels étant fort limités suite à de sérieux effets secondaires à long terme, il existe réellement un besoin médical important de développer de nouveaux traitements locaux qui soient bien tolérés, biocompatibles et biodégradables. Idéalement, ceux-ci devraient être actifs au niveau du processus inflammatoire ou de la douleur tout en étant capable de stabiliser, voire de restaurer, l’intégrité mécanique de l’articulation. <p>Dans cette optique, l’objectif de ce projet a été de développer des systèmes hydrolipidiques stériles, injectables et viscoélastiques qui soient capables de prolonger in situ la libération de principes actifs hydrophiles et lipophiles. Cette caractéristique devait permettre de réduire le nombre d’injections nécessaires dans le cadre du traitement symptomatique de l’arthrose et de maintenir l’effet des composés sur un minimum de quatre à six semaines. Cette étude entre dans le cadre du projet JOINT-AIC entièrement financé par le programme BioWin de la Région Wallonne. Le développement, la validation des méthodes analytiques, l’évaluation des propriétés physico-chimiques ainsi que la préparation stérile des lots de formulation destinés aux tests in vitro et in vivo ont été réalisés au sein du Laboratoire de Galénique et Biopharmacie de la Faculté de Pharmacie de l’ULB. <p>Au cours de ce projet, il a donc fallu dans un premier temps développer et caractériser des formulations hydrolipidiques à base de monoléine et d’acide hyaluronique permettant une libération in vitro prolongée de principes actifs tels que la clonidine (hydrophile) et le dipropionate de bétaméthasone (lipophile). Une étude de compatibilité a ainsi été préalablement réalisée afin de s’assurer qu’aucun des constituants principaux de la formulation ne présentaient d’incompatibilité physico-chimique qui pourrait être délétère vis-à-vis de leur stabilité en formulation. Ensuite, le développement de préparations hydro-lipidiques a été initié en tenant compte, d’une part de la solubilité des différents composants et, d’autre part de l’injectabilité, des propriétés rhéologiques et des profils de libération de la clonidine obtenus à partir des gels développés. Cette étude visait à obtenir une composition de référence qui soit à la fois injectable et capable de libérer un principe actif hydrophile sur plusieurs jours, voire plusieurs semaines, tout en possédant des propriétés rhéologiques intéressantes dans le cadre d’une viscosupplémentation articulaire. Enfin, un protocole de fabrication en milieu aseptique a été développé et plusieurs méthodes pour étudier les propriétés physico-chimiques des gels développés telles que la rhéologie, l’injectabilité, l’indice de gonflement, la stabilité et les profils de libérations ont été mises en place. <p>Les résultats ont montré qu’aucune incompatibilité ne semblait exister entre les trois composés majeurs de notre préparation, la monoléine, l’acide hyaluronique et la clonidine. Le développement des formulations nous a ensuite permis d’obtenir de nouveaux systèmes hydrolipidiques stériles et injectables à délivrance prolongée. Le véhicule qui remplissait au mieux nos objectifs était composé de 10,0% (m/m) d’éthanol ;de 15,0% de propylène glycol (m/m) ;de 15,0% (m/m) d’eau ;de 55,0% (m/m) de monoléine ;5,0% (m/m) d’huile de soja purifiée ;0,03% (m/m) d’α-tocophérol, de 7,5 mg/g d’HA et son pH était ajusté à 6,5 avec du NaOH 1N. Ce véhicule a montré un intérêt réel dans le cadre du développement de préparations biodégradables et biocompatibles pour le traitement de pathologies articulaires.En effet, cette composition présentait un écoulement de type pseudoplastique et des propriétés rhéologiques qui lui procuraient une bonne injectabilité. De plus, cette formulation a démontré in vitro une excellente capacité à gélifier au contact de fluides aqueux et à ralentir efficacement sur plusieurs semaines la libération des différents principes actifs incorporés (clonidine et dipropionate de bétaméthasone). Nous pouvions, dès lors, envisager que celle-ci serait capable de gélifier in situ au contact d’un fluide physiologique tel que le liquide synovial. Ensuite, suivant les recommandations de l’EMA, nous avons décidé d’utiliser l’association d’une filtration stérilisante et d’une préparation en milieu aseptique pour obtenir des formulations qui répondaient aux exigences en matière de préparation parentérale. C’est ainsi qu’un protocole de fabrication stérile de nos gels a été développé par nos soins en vue d’une éventuelle mise à l’échelle industrielle. Enfin, une étude de stabilité sur une année, suivant les normes ICH décrites pour des formulations destinées à être conservées au frigo, a été réalisée sur différents véhicules développés et contenant soit la clonidine, soit le dipropionate de bétaméthasone. Dans cette optique, plusieurs paramètres, tels que le dosage en principe actif, l’évolution du pH et du poids moléculaire de HA, le profil de libération ainsi que la rhéologie des formulations ont été évalués au cours du temps aux différentes conditions de conservation testées. Cette étude a permis de démontrer toute l’importance d’ajuster le pH de la préparation pour prévenir l’hydrolyse de l’HA, et cela indépendamment de l’incorporation de principe actif. Ainsi, il a pu être montré que l’ajustement du pH du véhicule à 6,5 à partir de NaOH dilué permettait d’améliorer considérablement la stabilité de la formulation puisqu’aucune modification significative de ses différents paramètres physico-chimiques et teneurs n’a été observée après un an de conservation à 5 et à 25 °C (60% HR) mais également après six mois à 30 °C (65% HR). Au cours de cette première partie, deux collaborations ont été initiées, l’une avec le Laboratoire du Prof. Siepmann de l’Université de Lille 2 et l’autre avec le Prof. Jerôme de l’Université de Liège. Avec l’aide du Prof. Siepmann, il a été possible de mettre au point un modèle mathématique pour caractériser les profils de libération des principes actifs à partir des différents véhicules développés. Le Prof. Jerôme a, quant à elle, mis à notre disposition un rhéomètre qui a permis d’approfondir nos connaissances sur les propriétés rhéologiques et viscoélastiques des formulations.<p>Ensuite, la seconde partie de notre travail a consisté à évaluer la tolérance, ainsi que l’efficacité des principes actifs sélectionnés et des formulations développées, à travers un modèle in vitro de cellules de l’inflammation (neutrophiles équins). Cette étude avait pour objectif d’évaluer deux aspects importants de la formulation :d’une part vérifier l’absence de réaction pro-inflammatoire qui pourrait être in vivo destructrice vis-à-vis du véhicule ainsi que des tissus environnants, et d’autre part vérifier l’effet anti-inflammatoire propre à la clonidine et au dipropionate de bétaméthasone seuls et en formulation. Cette étude a été réalisée avec la collaboration du Laboratoire du Prof. Serteyn de l’Université de Liège.Cette étude in vitro a démontré que les cellules restaient viables au moins pendant quatre heures lorsqu’elles étaient exposées à la matrice épurée de ses solvants. Ensuite, de manière surprenante, il a même pu être démontré que le véhicule permettait à la fois de prévenir et de réduire significativement la production des espèces réactives de l’oxygène (ROS) par les neutrophiles équins lorsque ceux-ci étaient stimulés au phorbol 12-myristate 13-acetate (PMA). Cette propriété peut être d’un grand intérêt dans le cadre de la prise en charge de l’arthrose car cette activité antioxydante pourrait permettre d’inhiber les dommages oxydatifs générés par les ROS et ainsi prévenir les dommages liés au développement du processus inflammatoire et qui peut, à long terme, s’avérer délétère pour les tissus environnants tels que le cartilage. Cette propriété du véhicule semble trouver son origine dans la monoléine qui, de par sa composition en alpha-tocophérol (200 ppm), présente également une activité antioxydante vis-à-vis des ROS. Toutefois, une action synergique liée à l’HA, à l’huile de soja ou à l’alpha-tocophérol incorporés aux formulations, n’est pas à exclure. Enfin, parmi les deux principes actifs sélectionnés, seul le dipropionate de bétaméthasone a montré une inhibition significative de la production des ROS.<p>Enfin, en tenant compte des résultats obtenus sur cellules, une étude in vivo pilote a été réalisée sur base d’un modèle de lapins. Cette étude visait à vérifier la tolérance ainsi que l’efficacité en prophylaxie de l’arthrose du véhicule développé ainsi que de la formulation contenant le dipropionate de bétaméthasone. Dans ce but, quatre groupes d’animaux (n=8) ont été constitués pour chacun des traitements testés :(i) groupe témoin :0,9 % tampon salin pH 7,4 ;(ii) véhicule à base de GMO développé; (iii) véhicule contenant du dipropionate de bétaméthasone ;(iv) groupe référence :Durolane®. Cette étude a été réalisée avec l’aide du Laboratoire du Prof. Henrotin de l’Université de Liège. L’hébergement des animaux ainsi que les actes chirurgicaux ont, quant à eux, été sous-traités par TNO (Delft, Pays-Bas).<p>De manière étonnante, il s’est avéré que le groupe contrôle présentait des scores macroscopique et histologique globalement peu élevés par rapport à ce qui est rapporté dans la littérature. Compte tenu de cette observation, il est difficile de se prononcer, sur base uniquement de cette étude, sur l’efficacité des différents traitements testés. Toutefois, il faut reconnaître que l’effet protecteur attendu pour le véhicule vis-à-vis de la dégradation du cartilage ne semble pas optimal et cela indépendamment de l’incorporation de dipropionate de bétaméthasone. Par ailleurs, l’étude des membranes synoviales a permis de démontrer qu’il n’y avait aucune différence significative en termes d’inflammation et de structure entre le groupe contrôle et les différents groupes traités. Ce qui signifie qu’aucun rejet n’a été observé vis-à-vis des formulations et que celles-ci ont, par conséquent, été bien tolérées malgré la formation de masses liées probablement au volume important injecté (900 µL) et au gonflement in situ du produit chez 37,5 % des lapins. Cette observation est importante puisqu’elle permet d’envisager de nouvelles perspectives telles que l’incorporation d’autres principes actifs pouvant éventuellement viser d’autres pathologies articulaires et pour lesquels une libération prolongée ou une protection mécanique du principe actif in situ serait bénéfique. <p><p><p><p><p><p> / Doctorat en Sciences biomédicales et pharmaceutiques / info:eu-repo/semantics/nonPublished

Σήμερα η μελέτη των βιοϋλικών προσανατολίζεται σε δύο κατευθύνσεις, την ανάπτυξη συστημάτων μεταφοράς φαρμάκων και συστημάτων κατάλληλων να διεγείρουν κυτταρικές λειτουργίες. Η έρευνά μας έχει σχέση με την συγκριτική μελέτη συστημάτων μεταφοράς φαρμάκων κατάλληλων για εφαρμογή σε οστικούς καρκίνους. Τέτοιου είδους συστήματα θα πρέπει, αρχικά να είναι ικανά να μεταφέρουν τα φαρμακευτικά μόρια και στη συνέχεια να μπορούν να επάγουν οστεογένεση. Η δεύτερη λειτουργικότητα είναι ιδιαιτέρως σημαντική καθώς έχει σαν αποτέλεσμα την πλήρωση του οστικού ελλείμματος που προκαλείται από την δράση των καρκινικών κυττάρων. Για τον σκοπό αυτό, διερευνήθηκε η ικανότητα του υαλώδους δικτύου να μεταφέρει φαρμακευτικά μόρια μέσω παραδοσιακών συστημάτων μεταφοράς. Συνεπώς, ακολούθησε η ex vitro μελέτη pH-ευαίσθητων τροποποιημένων πυριτικών ξηρών πηκτών στα οποία είχε συνδεθεί το αντικαρκινικό φάρμακο δοξορουπμυσίνη. Συγκεκριμένα, πυριτικά ξηρά πηκτώματα συντέθηκαν με την μέθοδο sol-gel και τροποποιήθηκαν περαιτέρω με χημεία καρβοδιϊμιδίου. Η τροποποίηση είχε σαν αποτέλεσμα την επιφανειακή σύνδεση υδροπηκτών δεξτράνης που παρουσιάζουν ευαισθησία στο pH. Στη συνέχεια, ακολούθησε η σύνθεση των ανόργανων βιοενεργών νανοσφαιρών. Για την σύνθεση των υαλωδών νανοσφαιρών με εσωτερική κοιλότητα ακολουθήθηκε η διαδικασία επικάλυψης sol-gel, κατά την οποία έγινε η ηλεκτροστατική επικάλυψη νανοσωματιδίων πολυστυρενίου με αποτέλεσμα την σύνθεση ανόργανων πυριτικών και φωσφοπυριτικών νανοσφαιρών. Επιπλέον, μελετήθηκε η εφαρμογή του συμπολυμερούς του πολυμεθακρυλικού μεθυλεστέρα – υδρομεθακρυλικού προπυλεστέρα ως υποστρώματος καθώς το PMMA αποτελεί βασικό συστατικό των οστικών τσιμέντων και παρουσιάζει βελτιωμένες μηχανικές ιδιότητες. Προκειμένου να ολοκληρωθεί η συγκριτική μελέτη μας, ακολούθησε η ex vitro ανάλυση των παραπάνω υβριδικών φωσφοπυριτικών νανοσφαιρών καθώς επίσης των πυριτικών νανοσφαιρών PS και PMMA-co-HPMA. Η επώαση σε διάλυμα SBF οδήγησε στον σχηματισμό ανθρακικού πυριτικού υδροξυαπατίτη με το μέγεθος των κρυσταλλιτών να μην υπερβαίνει τα 45 nm και έντονη παρουσία συσσωματωμάτων. Βέλτιστες ιδιότητες βιοενεργότητας παρουσιάζουν οι τροποποιημένες με αμίνες υβριδικές νανοσφαίρες PMMA-co-HPMA, οι οποίες έχουν επίσης την δυνατότητα να χρησιμοποιηθούν ως μεταφορείς φαρμακευτικών μορίων σε όξινο καθώς επίσης και σε φυσιολογικό pH με παρατεταμένη δυνατότητα αποδέσμευσης. / Recently the study of biomaterials has moved in two directions, the evolution of drug delivery systems and of systems that can stimulate specific cellular responses. Our investigation aims to the study of drug delivery systems for bone cancer therapy. These systems must fulfill two important functionalities. At first, they should be able to deliver drug molecules to bone cancer environment through loading or surface conjugation and subsequently to cause osteogenesis. Their second functionality is especially important since it leads to substitution of bone defects caused from the action of cancer cells. For this purpose, the ability of the glassy network to deliver drug molecules was studied. For this purpose, expanded ex vitro research was followed in DOX conjugated pH-sensitive functionalized silica xerogels. Specifically, silica xerogels were synthesized through a sol-gel process and further functionalized with carbodiimide chemistry. The functionalization process resulted in pH-sensitive dextran hydrogels. The study of the enhanced properties of glassy substrates was followed by the synthesis of amorphous bioactive nanospheres. Moreover, the change of the organic core and the use of PMMA-co-HPMA were advantageous due to the enhanced mechanical properties of PMMA-co-HPMA and its use in bone cements. In order to accomplish our comparative investigation, we followed the ex vitro study of the above hybrid binary silicate, ternary and quaternary phosphosilicate nanospheres as well as the silicate PS and PMMA-co-HPMA nanospheres. The incubation in SBF solution resulted in the formation of a silica-substituted carbonate hydroxyapatite (Si-HCA) a with crystallite size of around 45 nm and extended surface aggregates. The amino-modified PMMA-co-HPMA hybrid nanospheres present enhanced biocompatible properties, with prolonged release ability as drug delivery systems, in acidic as well as physiological pH.

Representing more than 50% of a worldwide pharmaceutical market of US$ 400 billions, oral drug delivery systems become naturally the focus of many studies. For almost half a century scientists have attempted to develop a theoretical model capable of predicting oral drug absorption in humans. From steady state or quasi-equilibrium models to complex and computationally intractable dynamic modeling approaches, numerous research efforts tried to address the problem of interest. Surprisingly though, no simple insightful first-principle-based dynamic modeling approaches have been reported in the literature. It is the purpose of the present work to provide a simple dynamic distributed-parameter modeling approach for performance monitoring of oral drug delivery systems. As a consequence of the complexity of the gastrointestinal tract, drug oral bioavailability is influenced by many different parameters. These parameters range from the compound's physicochemical properties, the physiological factors of the environment to other factors inherent in the drug form itself known as encapsulation factors. Physicochemical properties account for parameters such as drug stability, solubility or diffusivity. Furthermore, the environment, namely the gastrointestinal tract, influences the drug delivery process to the body with its pH, intestinal transit time and the different transport mechanisms that take place. From a chemical engineering point of view, the human body's anatomy can be analyzed and conceptually realized as a transport-reaction chemical system. Within the proposed modeling framework, the stomach is modeled as a non-ideal continuous-stirred tank reactor (CSTR) and the small intestine is the place where convection-diffusion occurs. The governing transport equations have been solved at steady state conditions in a small intestine represented by the lumen surrounded by its wall. The present work however develops a systematic dynamic first-principle-based distributed-parameter modeling framework where the time-dependent convection-diffusion-reaction model equations are analytically solved, offering the concentration profile in the small intestine lumen and in the wall from the moment the drug is administered until the complete absorption or disintegration of the drug particles. Once the modeling work is performed, a thorough and insightful sensitivity analysis can be conducted in order to assess the impact of the different process parameters on drug bioavailability.

Made available in DSpace on 2014-12-17T15:42:08Z (GMT). No. of bitstreams: 1 FabianoESG_TESE.pdf: 2562039 bytes, checksum: f41fc74bb604ad5b4eb0383a80628c68 (MD5) Previous issue date: 2010-03-30 / Conselho Nacional de Desenvolvimento Cient?fico e Tecnol?gico / Among the new drugs launched into the market since 1980, up to 30% of them belong to the class of natural products or they have semisynthetic origin. Between 40-70% of the new chemical entities (or lead compounds) possess poor water solubility, which may impair their commercial use. An alternative for administration of poorly water-soluble drugs is their vehiculation into drug delivery systems like micelles, microemulsions, nanoparticles, liposomes, and cyclodextrin systems. In this work, microemulsion-based drug delivery systems were obtained using pharmaceutically acceptable components: a mixture Tween 80 and Span 20 in ratio 3:1 as surfactant, isopropyl mirystate or oleic acid as oil, bidistilled water, and ethanol, in some formulations, as cosurfactants. Self-Microemulsifying Drug Delivery Systems (SMEDDS) were also obtained using propylene glycol or sorbitol as cosurfactant. All formulations were characterized for rheological behavior, droplet size and electrical conductivity. The bioactive natural product trans-dehydrocrotonin, as well some extracts and fractions from Croton cajucara Benth (Euphorbiaceae), Anacardium occidentale L. (Anacardiaceae) e Phyllanthus amarus Schum. & Thonn. (Euphorbiaceae) specimens, were satisfactorily solubilized into microemulsions formulations. Meanwhile, two other natural products from Croton cajucara, trans-crotonin and acetyl aleuritolic acid, showed poor solubility in these formulations. The evaluation of the antioxidant capacity, by DPPH method, of plant extracts loaded into microemulsions evidenced the antioxidant activity of Phyllanthus amarus and Anacardium occidentale extracts. For Phyllanthus amarus extract, the use of microemulsions duplicated its antioxidant efficiency. A hydroalcoholic extract from Croton cajucara incorporated into a SMEDDS formulation showed bacteriostatic activity against colonies of Bacillus cereus and Escherichia coli bacteria. Additionally, Molecular Dynamics simulations were performed using micellar systems, for drug delivery systems, containing sugar-based surfactants, N-dodecylamino-1-deoxylactitol and N-dodecyl-D-lactosylamine. The computational simulations indicated that micellization process for N-dodecylamino-1- deoxylactitol is more favorable than N-dodecyl-D-lactosylamine system. / Dos novos f?rmacos lan?ados no mercado a partir de 1980, cerca de 30% apresentam origem natural ou semissint?tica. Entre 40 e 70% destes novos prot?tipos de f?rmacos (naturais ou sint?ticos) apresentam baixa solubilidade aquosa, o que pode inviabilizar a sua utiliza??o comercial. Uma das alternativas encontrada pela ind?stria farmac?utica foi a veicula??o dessas subst?ncias a partir de ve?culos ou sistemas de libera??o de f?rmacos, tais como: micelas, microemuls?es, lipossomos, nanopart?culas e ciclodextrinas. Neste trabalho, sistemas microemulsionados biologicamente compat?veis foram obtidos utilizando a mistura Tween 80 e Span 20, na propor??o 3:1, como tensoativo, miristato de isopropila ou ?cido ol?ico como fase oleosa, ?gua bidestilada, e em alguns sistemas, etanol como cotensoativo. Tamb?m foram obtidos sistemas auto-microemulsificantes (SMEDDS) utilizando propilenoglicol ou sorbitol como cotensoativo. As microemuls?es obtidas foram caracterizadas quanto ao comportamento reol?gico, tamanho das got?culas e condutividade el?trica. O produto natural bioativo trans-desidrocrotonina (DCTN), bem como extratos e fra??es das esp?cies vegetais Croton cajucara Benth (Euphorbiaceae), Anacardium occidentale L. (Anacardiaceae) e Phyllanthus amarus Schum. & Thonn. (Euphorbiaceae), foram solubilizados satisfatoriamente nos sistemas microemulsionados obtidos, enquanto que outros produtos tamb?m isolados da esp?cie Croton cajucara, trans-crotonina e ?cido acetilaleurit?lico, n?o apresentaram resultados satisfat?rios em termos de solubilidade. A avalia??o da propriedade antioxidante, pelo m?todo do DPPH, dos extratos vegetais incorporados nos sistemas microemulsionados obtidos evidenciou a capacidade antioxidante dos extratos de Phyllanthus amarus e Anacardium occidentale, bem como a atua??o dos sistemas microemulsionados, que para o caso do extrato etan?lico de Phyllanthus amarus dobrou a sua atividade antioxidante quando solubilizado em microemuls?es. O extrato hidroalco?lico de Croton cajucara incorporado em um sistema auto-microemulsificante mostrou atividade bacteriost?tica frente a col?nias de bact?rias Bacillus cereus e Escherichia coli. Adicionalmente, foi realizado um estudo de simula??o computacional por Din?mica Molecular de sistemas micelares para uso farmacol?gico contendo tensoativos derivados de a??cares, N-dodecilamino-1-deoxilactitol e N-dodecil- -lactosilamina, que indicou que o processo de miceliza??o do primeiro sistema ? mais favor?vel que o segundo.

O paclitaxel é utilizado amplamente no carcinoma de ovário, nos casos refratários de carcinoma de endométrio e quimioterapia exclusiva para carcinoma avançado de colo uterino. A associação de paclitaxel a uma nanoemulsião rica em colesterol, denominada LDE, mostrou toxicidade menor e aumento da atividade antitumoral do fármaco em cobaias. No presente estudo, investigou-se os parâmetros farmacocinéticos do oleato de LDE-paclitaxel e a habilidade da LDE de concentrar o fármaco no tumor em oito pacientes com câncer do trato genital feminino. O oleate de paclitaxel associado a LDE é estável na circulação e tem uma meia-vida plasmática maior do que o paclitaxel comercial. A LDE concentra 3,6 mais paclitaxel em tecidos tumorais do que nos tecidos normais. Esta associação parece ser uma alternativa no tratamento dos tumores ginecológicos / A cholesterol-rich nanoemulsion termed LDE concentrates in cancer tissues after injection into the bloodstream. The association of a derivatized paclitaxel to LDE showed lower toxicity and increased antitumoral activity as tested in mice. Here, the pharmacokinetics of LDE-paclitaxel oleate and the ability of LDE to concentrate the drug in the tumor were investigated in eight patients with gynecologic cancers. Fractional clearance rate (FCR) and pharmacokinetic parameters were calculated by compartmental analysis. Also, specimens of tumors and the normal tissues were excised during the surgery for radioactivity measurement. LDE concentrates 3.5 more paclitaxel in malignant tissues than in the normal tissues. Therefore, association to LDE is an interesting strategy for using paclitaxel to treat gynecologic cancers

O paclitaxel é utilizado amplamente no carcinoma de ovário, nos casos refratários de carcinoma de endométrio e quimioterapia exclusiva para carcinoma avançado de colo uterino. A associação de paclitaxel a uma nanoemulsião rica em colesterol, denominada LDE, mostrou toxicidade menor e aumento da atividade antitumoral do fármaco em cobaias. No presente estudo, investigou-se os parâmetros farmacocinéticos do oleato de LDE-paclitaxel e a habilidade da LDE de concentrar o fármaco no tumor em oito pacientes com câncer do trato genital feminino. O oleate de paclitaxel associado a LDE é estável na circulação e tem uma meia-vida plasmática maior do que o paclitaxel comercial. A LDE concentra 3,6 mais paclitaxel em tecidos tumorais do que nos tecidos normais. Esta associação parece ser uma alternativa no tratamento dos tumores ginecológicos / A cholesterol-rich nanoemulsion termed LDE concentrates in cancer tissues after injection into the bloodstream. The association of a derivatized paclitaxel to LDE showed lower toxicity and increased antitumoral activity as tested in mice. Here, the pharmacokinetics of LDE-paclitaxel oleate and the ability of LDE to concentrate the drug in the tumor were investigated in eight patients with gynecologic cancers. Fractional clearance rate (FCR) and pharmacokinetic parameters were calculated by compartmental analysis. Also, specimens of tumors and the normal tissues were excised during the surgery for radioactivity measurement. LDE concentrates 3.5 more paclitaxel in malignant tissues than in the normal tissues. Therefore, association to LDE is an interesting strategy for using paclitaxel to treat gynecologic cancers

Cellular internalization of large therapeutic agents such as proteins or nucleic acids is a challenging task because of the presence of the plasma membrane. One strategy to facilitate intracellular drug uptake is to induce transient pores in the cell membrane through physical delivery strategies. Physical approaches are attractive as they offer more generic applicability compared with viral or biochemical counterparts. Pulsed laser light can induce the endothermic carbon-steam reaction in carbon-nanoparticle suspensions to produce explosive photoacoustic effects in the surrounding medium. In this study, for the first time, these photoacoustic forces were used to transiently permeabilize the cell membrane to deliver macromolecules into cells. Intracellular delivery using this method was demonstrated in multiple cell types for uptake of small molecules, proteins and DNA. At optimized conditions, uptake was seen in up to 50% of cells with nearly 100% viability and in 90% of cells with ≥90% viability, which compared favorably with other physical methods of drug delivery. Cellular bioeffects were shown to be a consequence of laser-carbon interaction and correlated with properties of the carbon and laser, such as carbon concentration and size, laser pulse duration, wavelength, intensity and exposure time. Similar results were observed using two different lasers, a femtosecond Ti: Sapphire laser and a nanosecond Nd: YAG laser. Uptake was also shown in murine skeletal muscles in vivo with up to 40% efficiency compared to non-irradiated controls. This synergistic use of nanotechnology with advanced laser technology could provide an alternative to viral and chemical-based drug and gene delivery.

With the emergence of novel and more effective drug therapies, increased importance is being placed upon the methods by which these drugs are being delivered to the body. In conventional drug delivery systems, there is very little control over the release of drug. The effective concentration at the target site can be achieved by intermittent administration of grossly excessive doses, which, often results in constantly, unpredictable variations in plasma concentrations, with the risk of reaching levels below or above the therapeutic range leading to marked side effects. A plethora of formulation strategies mainly based on polymeric/lipid nanoparticles, are described in literature. Even though these systems are therapeutically advantageous in comparison to conventional systems, they remain insensitive to the changing metabolic states of the body although the symptoms of most metabolic diseases follow a rhythmic pattern.A more appropriate and effective approach of managing some of these conditions lies in the chronotherapy. This approach allows for pulsed or self-regulated drug delivery which is adjusted to the staging of biological rhythms, since the onset of certain diseases exhibits strong circadian temporal dependence. In order to reach the objective of mimicking the biophysical and biochemical processes of pathological states, many innovations in material design for drug delivery systems (DDS) that are able to release the therapeutic payload-on-demand were done to release the therapeutic agent only when it is required, according to the physiological need. The development of multidisciplinary research teams has brought huge advantages in the design, fabrication and utilization of such smart systems, especially in the pharmaceutical field. Interestingly, numerous smart polymeric materials exhibit a response to a specific stimulus. A step further, the elaboration of purpose-built monomers can give rise to compounds with tunable sensitivities or multi-stimuli responsiveness. These smart polymers demonstrate an active responsiveness to environmental (or external) signals and change their physicochemical properties as designed (e.g. conformation, solubility, shape, charge or size). As far as the stimuli are concerned, they consist of physical (e.g. temperature, ultrasound, light, electricity, magnetic or mechanical stress), chemical (e.g. pH, ionic strength) and biological signals (e.g. enzymes, biomolecules). Due to the intrapersonal variabilities which may make internal stimuli hazardous, externally controlled systems rely on externally applied stimuli that are produced by stimuli-generating devices, which results in pulsed drug delivery. This type of delivery may be rapid and allows a transient release of a determined amount of drug within a short period of time immediately after a pre-determined off-release period. A novel strategy for the formation of multi-stimuli responsive materials endowed with pH, magnetic and light sensitivity was achieved. The approach relied on the incorporation of magnetic tetrahalogenoferrate(III) anions along a polymeric backbone based on poly(2-(N,N-dimethylamino) ethyl meth-acrylate) (PDMAEMA). Starting from the same PDMAEMA, quaternized pending amine groups with various halide derivatives gave rise to magnetic materials after anion metathesis. Measuring the magnetic susceptibility of these materials exhibited that the magnetic susceptibility increased as the substituted group size decreased (become smaller) which was apparently related to the steric hindrance around the ionic pendants. Additionally, a good correlation between the magnetic susceptibility and ferric content was found. Additional experimental and theoretical Raman analyses allowed the determination of the nature of the magnetic species constituting the materials. This strategy further offers the opportunity to tailor the magnetic response through partial ammonium salt formation. In order to merge the magnetic properties of ferric-based materials with another stimuli-responsive functionality, random copolymers containing DMAEMA (D) with diazobenzene (A) unit were prepared. So, three copolymers PDA were synthesized (with targeted D/A ratios 4/6 (PDA4), 6/4 (PDA6) and 8/2 (PDA8)). Meanwhile, different degrees of amine quaternization (10, 50 and 100 %) were applied, which led to the following polymeric salts PDAX/Y where X = 4, 6, 8 (referring to the percentage of the DMAEMA unit) and Y = 10, 50 and 100 (referring to the percentage of quaternized amine groups). Finally, the aforementioned materials were converted into magnetic polymers by anion exchange. As a result, magnetic responses correlated well with amount of iron oxide in these compounds and the amount of ionic pending groups along the backbone. Moreover, the remaining tertiary amines conferred pH sensitivity to the polymers whereas the diazobenzene units ensured light responsiveness through the well-established trans-to-cis isomerization.In order to functionalize these materials in the pharmaceutical field, an intelligent delivery system was prepared. Firstly, an attempt to formulate riboflavin-5’-phosphate sodium (RPS) loaded on PDA8 microspheres was made using double emulsion evaporation method. Meanwhile, prednisolone (PRD) microspheres were prepared using s/o/w emulsion technique. Subsequently, coating systems of cochineal red tablets were developed. These tablets were coated with polymer solution (using each of three types of copolymers: PDA8, PDA6, and PDA4) until the desired percentage of the coating was achieved (10, 15, and 20 % w/w). The cumulative release profiles of cochineal red tablets coated with PDA8, PDA6, and PDA4 showed a pH-sensitive release behavior. The release in the neutral media (pH ≈ 7.0) was very slow (less than 3 % after one hour). Then, after changing the pH to 1.2, an increase in the release of cochineal was observed. Furthermore, the cumulative release of cochineal red was at the highest value for the PDA8 and the lowest for PDA4 depending on the percentage of PDMAEMA moieties. Moreover, by increasing the percentage of the coating from (10, 15 to 20 % w/w), the cumulative release of cochineal decreased. Therefore, the copolymer PDAX can be used for controlling the release of drug by changing the pH value.Finally, the cochineal tablets coated with PDA6 (10 %) showed features of light sensitivity. The release of cochineal red from coated tablets was only due to the switching in the conformational trans/cis isomerization of azobenzene moieties upon irradiation, which was confirmed by comparing the release of coated tablets with uncoated tablets upon irradiation. / Doctorat en Sciences biomédicales et pharmaceutiques (Pharmacie) / info:eu-repo/semantics/nonPublished