Radiolabeling of defined polymer architectures with fluorine-18 and iodine-131 for ex vivo and in vivo evaluation : visualization of structure-property relationships

Nuclear medicine imaging techniques such as PET are of increasing relevance in pharmaceutical research being valuable (pre)clinical tools to non-invasively assess drug performance in vivo. Therapeutic drugs, e.g. chemotherapeutics, often suffer from a poor balance between their efficacy and toxicity. Here, polymer based drug delivery systems can modulate the pharmacokinetics of low Mw therapeutics (prolonging blood circulation time, reducing toxic side effects, increasing target site accumulation) and therefore leading to a more efficient therapy. In this regard, poly-N-(2-hydroxypropyl)-methacrylamide (HPMA) constitutes a promising biocompatible polymer. Towards the further development of these structures, non-invasive PET imaging allows insight into structure-property relationships in vivo. This performant tool can guide design optimization towards more effective drug delivery. Hence, versatile radiolabeling strategies need to be developed and establishing 18F- as well as 131I-labeling of diverse HPMA architectures forms the basis for short- as well as long-term in vivo evaluations. By means of the prosthetic group [18F]FETos, 18F-labeling of distinct HPMA polymer architectures (homopolymers, amphiphilic copolymers as well as block copolymers) was successfully accomplished enabling their systematic evaluation in tumor bearing rats. These investigations revealed pronounced differences depending on individual polymer characteristics (molecular weight, amphiphilicity due to incorporated hydrophobic laurylmethacrylate (LMA) segments, architecture) as well as on the studied tumor model. Polymers showed higher uptake for up to 4 h p.i. into Walker 256 tumors vs. AT1 tumors (correlating to a higher cellular uptake in vitro). Highest tumor concentrations were found for amphiphilic HPMA-ran-LMA copolymers in comparison to homopolymers and block copolymers. Notably, the random LMA copolymer P4* (Mw=55 kDa, 25% LMA) exhibited most promising in vivo behavior such as highest blood retention as well as tumor uptake. Further studies concentrated on the influence of PEGylation (‘stealth effect’) in terms of improving drug delivery properties of defined polymeric micelles. Here, [18F]fluoroethylation of distinct PEGylated block copolymers (0%, 1%, 5%, 7%, 11% of incorporated PEG2kDa) enabled to systematically study the impact of PEG incorporation ratio and respective architecture on the in vivo performance. Most strikingly, higher PEG content caused prolonged blood circulation as well as a linear increase in tumor uptake (Walker 256 carcinoma). Due to the structural diversity of potential polymeric carrier systems, further versatile 18F-labeling strategies are needed. Therefore, a prosthetic 18F-labeling approach based on the Cu(I)-catalyzed click reaction was established for HPMA-based polymers, providing incorporation of fluorine-18 under mild conditions and in high yields. On this basis, a preliminary µPET study of a HPMA-based polymer – radiolabeled via the prosthetic group [18F]F-PEG3-N3 – was successfully accomplished. By revealing early pharmacokinetics, 18F-labeling enables to time-efficiently assess the potential of HPMA polymers for efficient drug delivery. Yet, investigating the long-term fate is essential, especially regarding prolonged circulation properties and passive tumor accumulation (EPR effect). Therefore, radiolabeling of diverse HPMA copolymers with the longer-lived isotope iodine-131 was accomplished enabling in vivo evaluation of copolymer P4* over several days. In this study, tumor retention of 131I-P4* could be demonstrated at least over 48h with concurrent blood clearance thereby confirming promising tumor targeting properties of amphiphilic HPMA copolymer systems based on the EPR effect.

Teoria del consenso e applicazione al problema del coordinamento del moto di robot.

Il sempre crescente numero di applicazioni di reti di sensori, robot cooperanti e formazioni di veicoli, ha fatto sì che le problematiche legate al coordinamento di sistemi multi-agente (MAS) diventassero tra le più studiate nell’ambito della teoria dei controlli. Esistono numerosi approcci per affrontare il problema, spesso profondamente diversi tra loro. La strategia studiata in questa tesi è basata sulla Teoria del Consenso, che ha una natura distribuita e completamente leader-less; inoltre il contenuto informativo scambiato tra gli agenti è ridotto al minimo. I primi 3 capitoli introducono ed analizzano le leggi di interazione (Protocolli di Consenso) che permettono di coordinare un Network di sistemi dinamici. Nel capitolo 4 si pensa all'applicazione della teoria al problema del "loitering" circolare di più robot volanti attorno ad un obiettivo in movimento. Si sviluppa a tale scopo una simulazione in ambiente Matlab/Simulink, che genera le traiettorie di riferimento di raggio e centro impostabili, a partire da qualunque posizione iniziale degli agenti. Tale simulazione è stata utilizzata presso il “Center for Research on Complex Automated Systems” (CASY-DEI Università di Bologna) per implementare il loitering di una rete di quadrirotori "CrazyFlie". I risultati ed il setup di laboratorio sono riportati nel capitolo 5. Sviluppi futuri si concentreranno su algoritmi locali che permettano agli agenti di evitare collisioni durante i transitori: il controllo di collision-avoidance dovrà essere completamente indipendente da quello di consenso, per non snaturare il protocollo di Consenso stesso.

From defined polymer architectures to structure-property relationships in vivo

Makromolekulare Wirkstoffträgersysteme sind von starkem Interesse bezüglich der klinischen Anwendung chemotherapeutischer Agenzien. Um ihr klinisches Potential zu untersuchen ist es von besonderer Bedeutung das pharmakokinetische Profil in vivo zu bestimmen. Jede Veränderung der Polymerstruktur beeinflusst die Körperverteilung des entsprechenden Makromoleküls. Aufgrund dessen benötigt man detailliertes Wissen über Struktur-Eigenschaftsbeziehungen im lebenden Organismus, um das Nanocarrier System für zukünftige Anwendungen einzustellen. In dieser Beziehung stellt das präklinische Screening mittels radioaktiver Markierung und Positronen-Emissions-Tomographie eine nützliche Methode für schnelle sowie quantitative Beobachtung von Wirkstoffträgerkandidaten dar. Insbesondere poly(HPMA) und PEG sind im Arbeitsgebiet Polymer-basierter Therapeutika stark verbreitet und von ihnen abgeleitete Strukturen könnten neue Generationen in diesem Forschungsbereich bieten.rnDie vorliegende Arbeit beschreibt die erfolgreiche Synthese verschiedener HPMA und PEG basierter Polymer-Architekturen – Homopolymere, Statistische und Block copolymere – die mittels RAFT und Reaktivesterchemie durchgeführt wurde. Des Weiteren wurden die genannten Polymere mit Fluor-18 und Iod-131 radioaktiv markiert und mit Hilfe von microPET und ex vivo Biodistributionsstudien in tumortragenden Ratten biologisch evaluiert. Die Variation in Polymer-Architektur und darauffolgende Analyse in vivo resultierte in wichtige Schlussfolgerungen. Das hydrophile / lipophile Gleichgewicht hatte einen bedeutenden Einfluss auf das pharmakokinetische Profil, mit besten in vivo Eigenschaften (geringe Aufnahme in Leber und Milz sowie verlängerte Blutzirkulationszeit) für statistische HPMA-LMA copolymere mit steigendem hydrophoben Anteil. Außerdem zeigten Langzeitstudien mit Iod-131 eine verstärkte Retention von hochmolekularen, HPMA basierten statistischen Copolymeren im Tumorgewebe. Diese Beobachtung bestätigte den bekannten EPR-Effekt. Hinzukommend stellen Überstrukturbildung und damit Polymergröße Schlüsselfaktoren für effizientes Tumor-Targeting dar, da Polymerstrukturen über 200 nm in Durchmesser schnell vom MPS erkannt und vom Blutkreislauf eliminiert werden. Aufgrund dessen wurden die hier synthetisierten HPMA Block copolymere mit PEG Seitengruppen chemisch modifiziert, um eine Verminderung in Größe sowie eine Reduktion in Blutausscheidung zu induzieren. Dieser Ansatz führte zu einer erhöhten Tumoranreicherung im Walker 256 Karzinom Modell. Generell wird die Körperverteilung von HPMA und PEG basierten Polymeren stark durch die Polymer-Architektur sowie das Molekulargewicht beeinflusst. Außerdem hängt ihre Effizienz hinsichtlich Tumorbehandlung deutlich von den individuellen Charakteristika des einzelnen Tumors ab. Aufgrund dieser Beobachtungen betont die hier vorgestellte Dissertation die Notwendigkeit einer detaillierten Polymer-Charakterisierung, kombiniert mit präklinischem Screening, um polymere Wirkstoffträgersysteme für individualisierte Patienten-Therapie in der Zukunft maßzuschneidern.rn

Multifunctional polyether architectures : versatile materials for surface attachment and functionalization

Chapter 1 of this thesis comprises a review of polyether polyamines, i.e., combinations of polyether scaffolds with polymers bearing multiple amino moieties. Focus is laid on controlled or living polymerization methods. Furthermore, fields in which the combination of cationic, complexing, and pH-sensitive properties of the polyamines and biocompatibility and water-solubility of polyethers promise enormous potential are presented. Applications include stimuli-responsive polymers with a lower critical solution temperature (LCST) and/or the ability to gel, preparation of shell cross-linked (SCL) micelles, gene transfection, and surface functionalization.rnIn Chapter 2, multiaminofunctional polyethers relying on the class of glycidyl amine comonomers for anionic ring-opening polymerization (AROP) are presented. In Chapter 2.1, N,N-diethyl glycidyl amine (DEGA) is introduced for copolymerization with ethylene oxide (EO). Copolymer microstructure is assessed using online 1H NMR kinetics, 13C NMR triad sequence analysis, and differential scanning calorimetry (DSC). The concurrent copolymerization of EO and DEGA is found to result in macromolecules with a gradient structure. The LCSTs of the resulting copolymers can be tailored by adjusting DEGA fraction or pH value of the environment. Quaternization of the amino moieties by methylation results in polyelectrolytes. Block copolymers are used for PEGylated gold nanoparticle formation. Chapter 2.2 deals with a glycidyl amine monomer with a removable protecting group at the amino moiety, for liberation of primary amines at the polyether backbone, which is N,N-diallyl glycidyl amine (DAGA). Its allyl groups are able to withstand the harsh basic conditions of AROP, but can be cleaved homogeneously after polymerization. Gradient as well as block copolymers poly(ethylene glycol)-PDAGA (PEG-PDAGA) are obtained. They are analyzed regarding their microstructure, LCST behavior, and cleavage of the protecting groups. rnChapter 3 describes applications of multi(amino)functional polyethers for functionalization of inorganic surfaces. In Chapter 3.1, they are combined with an acetal-protected catechol initiator, leading to well-defined PEG and heteromultifunctional PEG analogues. After deprotection, multifunctional PEG ligands capable of attaching to a variety of metal oxide surfaces are obtained. In a cooperative project with the Department of Inorganic and Analytical Chemistry, JGU Mainz, their potential is demonstrated on MnO nanoparticles, which are promising candidates as T1 contrast agents in magnetic resonance imaging. The MnO nanoparticles are solubilized in aqueous solution upon ligand exchange. In Chapter 3.2, a concept for passivation and functionalization of glass surfaces towards gold nanorods is developed. Quaternized mPEG-b-PqDEGA diblock copolymers are attached to negatively charged glass surfaces via the cationic PqDEGA blocks. The PEG blocks are able to suppress gold nanorod adsorption on the glass in the flow cell, analyzed by dark field microscopy.rnChapter 4 highlights a straightforward approach to poly(ethylene glycol) macrocycles. Starting from commercially available bishydroxy-PEG, cyclic polymers are available by perallylation and ring-closing metathesis in presence of Grubbs’ catalyst. Purification of cyclic PEG is carried out using α-cyclodextrin. This cyclic sugar derivative forms inclusion complexes with remaining unreacted linear PEG in aqueous solution. Simple filtration leads to pure macrocycles, as evidenced by SEC and MALDI-ToF mass spectrometry. Cyclic polymers from biocompatible precursors are interesting materials regarding their increased blood circulation time compared to their linear counterparts.rnIn the Appendix, A.1, a study of the temperature-dependent water-solubility of polyether copolymers is presented. Macroscopic cloud points, determined by turbidimetry, are compared with microscopic aggregation phenomena, monitored by continuous wave electron paramagnetic resonance (CW EPR) spectroscopy in presence of the amphiphilic spin probe and model drug (2,2,6,6-tetramethylpiperidin-1-yl)oxyl (TEMPO). These thermoresponsive polymers are promising candidates for molecular transport applications. The same techniques are applied in Chapter A.2 to explore the pH-dependence of the cloud points of PEG-PDEGA copolymers in further detail. It is shown that the introduction of amino moieties at the PEG backbone allows for precise manipulation of complex phase transition modes. In Chapter A.3, multi-hydroxyfunctional polysilanes are presented. They are obtained via copolymerization of the acetal-protected dichloro(isopropylidene glyceryl propyl ether)methylsilane monomer. The hydroxyl groups are liberated through acidic work-up, yielding versatile access to new multifunctional polysilanes.

From anionic polymerization in continuous flow to macromolecular architectures and stimuli-responsive surfaces

The thesis can be divided in four parts and summarized as follows:(i) The investigation and development of a continuous flow synthesis procedure affording end-functional polymers by anionic polymerization and subsequent termination in one reaction step and on a multigram scale was carried out. Furthermore, the implementation of not only a single hydroxyl but multiple orthogonal functionalities at the chain terminus was achieved by utilizing individually designed, functional epoxide-based end-capping reagents.(ii) In an additional step, the respective polymers were used as macroinitiators to prepare in-chain functionalized block copolymers and star polymers bearing intriguing novel structural and material properties. Thus, the second part of this thesis presents the utilization of end-functional polymers as precursors for the synthesis of amphiphilic complex and in some cases unprecedented macromolecular architectures, such as miktoarm star polymers based on poly(vinyl pyridine), poly(vinyl ferrocene) and PEO.(iii) Based on these structures, the third part of this thesis represents a detailed investigation of the preparation of stimuli-responsive ultrathin polymer films, using amphiphilic junction point-reactive block copolymers. The single functionality at the block interface can be employed as anchor group for the covalent attachment on surfaces. Furthermore, the change of surface properties was studied by applying different external stimuli.(iv) An additional topic related to the oxyanionic polymerizations carried out in the context of this thesis was the investigation of viscoelastic properties of different hyperbranched polyethers, inspired by the recent and intense research activities in the field of biomedical applications of multi-functional hyperbranched materials.

Stato dell'arte della progettazione automatica di programmi per robot

La comprensione delle proprietà e delle meccaniche di funzionamento dei sistemi complessi e dinamici che ci circondano è uno degli obiettivi principali delle discipline ingegneristiche e scientifiche. Lo sviluppo nel campo dell'Intelligenza Artificiale ha permesso la creazione di entità robotiche sempre più intelligenti, in grado di manifestare comportamenti notevoli e di raggiungere gli obiettivi prefissati: la progettazione di software di controllo ha velocemente ricevuto una crescente responsabilità. Questo elaborato si propone di inquadrare in generale la situazione della progettazione di software di controllo robotici, focalizzandosi in particolare sulle tecniche di progettazione automatica di programmi per robot: dopo aver inquadrato il background su cui si basa la progettazione automatica ed aver mostrato una panoramica dei principali metodi di progettazione diretta, vengono elencate e spiegate le tecniche più utilizzate ed interessanti della progettazione automatica di programmi per robot.

Applicazione del linguaggio ad agenti Jason per la programmazione di alto livello di robot

I sistemi software rivestono nel campo dell'informatica un ruolo di fondamentale importanza. Negli ultimi anni una caratteristica richiesta ai sistemi è la decentralizzazione del controllo, nell'ottica di un sistema visto come connessioni di parti che possono interagire, e dove ciascuna parte possiede un certo grado di autonomia nella scelta delle attività che devono essere compiute. In tale contesto si introduce il paradigma degli agenti, in quanto include sia aspetti relativi ai modelli computazionali, sia aspetti relativi ai linguaggi. Nella tesi si esplora l'applicazione del linguaggio di programmazione Jason applicato alla programmazione dei controllori di robot. In particolare, si esplora ciò che riguarda il comportamento individuale dell'agente.

Extracting FSA descriptions of robot behaviours from the dynamics of automatically designed controller networks

Automatic design has become a common approach to evolve complex networks, such as artificial neural networks (ANNs) and random boolean networks (RBNs), and many evolutionary setups have been discussed to increase the efficiency of this process. However networks evolved in this way have few limitations that should not be overlooked. One of these limitations is the black-box problem that refers to the impossibility to analyze internal behaviour of complex networks in an efficient and meaningful way. The aim of this study is to develop a methodology that make it possible to extract finite-state automata (FSAs) descriptions of robot behaviours from the dynamics of automatically designed complex controller networks. These FSAs unlike complex networks from which they're extracted are both readable and editable thus making the resulting designs much more valuable.

Tesi sperimentale per valutare l'esperienza chirurgica nell'uso del robot Da Vinci Si

Questa tesi si compone di due parti tra loro correlate. Nella prima viene analizzato l’attuale stato dell’arte in ambito di robot chirurgici e, successivamente, sono illustrate le caratteristiche tecniche e funzionali del robot chirurgico Da Vinci Si e ne viene evidenziata la mancanza di sensori in grado di fornire all’utilizzatore un feedback aptico. La seconda parte propone uno studio che ha lo scopo di dimostrare come l’esperienza del chirurgo riesca a sopperire alla mancanza di un sensore in grado di fornire un riscontro della forza impressa sugli strumenti nel campo operatorio. A questo proposito, vengono illustrati i risultati del test appositamente costruito, cui è stato sottoposto un campione di chirurghi di diversi ambiti specialistici, per dimostrare come l’esperienza e la destrezza consentono all’utilizzatore del robot Da Vinci Si di ricevere un’indicazione della forza impressa sui tessuti del campo operatorio tramite un riscontro visivo.

Un’applicazione di elaborazione del linguaggio naturale per il gioco di "Indovina chi?" su robot umanoide

La tesi è stata incentrata sul gioco «Indovina chi?» per l’identificazione da parte del robot Nao di un personaggio tramite la sua descrizione. In particolare la descrizione avviene tramite domande e risposte L’obiettivo della tesi è la progettazione di un sistema in grado di capire ed elaborare dei dati comunicati usando un sottoinsieme del linguaggio naturale, estrapolarne le informazioni chiave e ottenere un riscontro con informazioni date in precedenza. Si è quindi programmato il robot Nao in modo che sia in grado di giocare una partita di «Indovina chi?» contro un umano comunicando tramite il linguaggio naturale. Sono state implementate regole di estrazione e categorizzazione per la comprensione del testo utilizzando Cogito, una tecnologia brevettata dall'azienda Expert System. In questo modo il robot è in grado di capire le risposte e rispondere alle domande formulate dall'umano mediante il linguaggio naturale. Per il riconoscimento vocale è stata utilizzata l'API di Google e PyAudio per l'utilizzo del microfono. Il programma è stato implementato in Python e i dati dei personaggi sono memorizzati in un database che viene interrogato e modificato dal robot. L'algoritmo del gioco si basa su calcoli probabilistici di vittoria del robot e sulla scelta delle domande da proporre in base alle risposte precedentemente ricevute dall'umano. Le regole semantiche realizzate danno la possibilità al giocatore di formulare frasi utilizzando il linguaggio naturale, inoltre il robot è in grado di distinguere le informazioni che riguardano il personaggio da indovinare senza farsi ingannare. La percentuale di vittoria del robot ottenuta giocando 20 partite è stata del 50%. Il data base è stato sviluppato in modo da poter realizzare un identikit completo di una persona, oltre a quello dei personaggi del gioco. È quindi possibile ampliare il progetto per altri scopi, oltre a quello del gioco, nel campo dell'identificazione.

Installazione del navigation stack su rover terrestre e applicazione del kinect nello human-robot interaction.

Progetto SHERPA. Installazione e configurazione del Navigaton Stack su Rover terrestre. Utilizzo e configurazione di LMS151 Sick. Utilizzo e configurazione di Asus Xtion Pro. Progettazione di software per la localizzazione e l'inseguimento di persone tramite camera di profondita.

Analysis of robot dynamics through complexity measures: an application in automatic design

L'applicazione di misure, derivanti dalla teoria dell'informazione, fornisce un valido strumento per quantificare alcune delle proprietà dei sistemi complessi. Le stesse misure possono essere utilizzate in robotica per favorire l'analisi e la sintesi di sistemi di controllo per robot. In questa tesi si è analizzata la correlazione tra alcune misure di complessità e la capacità dei robot di portare a termine, con successo, tre differenti task. I risultati ottenuti suggeriscono che tali misure di complessità rappresentano uno strumento promettente anche nel campo della robotica, ma che il loro utilizzo può diventare difficoltoso quando applicate a task compositi.

Retraining of interjoint arm coordination after stroke using robot-assisted time-independent functional training

We have developed a haptic-based approach for retraining of interjoint coordination following stroke called time-independent functional training (TIFT) and implemented this mode in the ARMin III robotic exoskeleton. The ARMin III robot was developed by Drs. Robert Riener and Tobias Nef at the Swiss Federal Institute of Technology Zurich (Eidgenossische Technische Hochschule Zurich, or ETH Zurich), in Zurich, Switzerland. In the TIFT mode, the robot maintains arm movements within the proper kinematic trajectory via haptic walls at each joint. These arm movements focus training of interjoint coordination with highly intuitive real-time feedback of performance; arm movements advance within the trajectory only if their movement coordination is correct. In initial testing, 37 nondisabled subjects received a single session of learning of a complex pattern. Subjects were randomized to TIFT or visual demonstration or moved along with the robot as it moved though the pattern (time-dependent [TD] training). We examined visual demonstration to separate the effects of action observation on motor learning from the effects of the two haptic guidance methods. During these training trials, TIFT subjects reduced error and interaction forces between the robot and arm, while TD subject performance did not change. All groups showed significant learning of the trajectory during unassisted recall trials, but we observed no difference in learning between groups, possibly because this learning task is dominated by vision. Further testing in stroke populations is warranted.

Iron(III)-Pivalate-Based Complexes with Tetranuclear {Fe-4(mu(3)-O)(2)}(8+) Cores and N-Donor Ligands: Formation of Cluster and Polymeric Architectures

Different synthetic routes have been used for the preparation of a new tetranuclear [Fe4O2(O2CCMe3)(8)(bpm)] cluster (1) and a one-dimensional coordination polymer [Fe4O2-(O2CCMe3)(8)(hmta)](n) (2) (bpm = 2,2'-bipyrimidine and hmta = hexamethylenetetramine). For cluster 1, two structural isomers, 1a and 1b center dot 3MeCN, have been found. X-ray crystallographic analysis showed that all complexes consist of a central {Fe-4(mu(3)-O)(2)}(8+) core. In 1a, metal ions in the core are additionally linked by six bridging pivalates as two other pivalates and a bpm ligand are chelated to Fe-III ions, whereas in cluster 1b, metal ions in the {Fe-4(mu(3)-O)(2)}(8+) core are linked by seven bridging pivalates and only one carboxylate as well as bpm are chelated to the iron centers. In coordination polymer 2, [Fe4O2(O2CCMe3)(8)] clusters are bridged by hmta ligands to form zigzag chains. Magnetic measurements have been carried out to characterize these complexes and revealed antiferromagnetic interactions between Fe-III ions with best-fit parameters of J(wb) = -72.2 (1a) and -88.7 cm(-1) (1b) for wing...body interactions.