Conjugates of Porphyrins and Hyperbranched Polyglycerol for an Application in Photodynamic Therapy

This thesis consists of four different parts of the synthesis and investigation of various porphyrins containing different types of linkers for the connection to substrates, conjugates of these porphyrins with hyperbranched polyglycerol (hPG) in nanoparticles and nanogels, hPG-porphyrin conjugates additionally loaded with mannose and investigation of their antibacterial activity, and cleavable porphyrin-hPG-conjugates: A) The reaction of pentafluorophenyl-substituted A3B-porphyrins with amines has been used to obtain diverse reactive groups which are useful for the further functionalization and/or conjugation to different substrates and/or materials. With this method, porphyrins with alkenyl-, alkynyl-, amino-, azido-, cyclooctyne-, epoxide-, hydroxyl- and maleimido-groups have been obtained. The conjugation of an alkynyl porphyrin to an hPG-azide via the copper(I)-catalyzed 1,3-dipolar cycloaddition (CuAAC) is shown. For a possible application in the photodynamic therapy (PDT) the photocytotoxicity of porphyrin-hPG-conjugates has been demonstrated against human epidermoid carcinoma A-253 and squamous carcinoma CAL-27 cells. To allow cleavage of the porphyrins from a carrier system, examples of porphyrins containing disulfide (for reductive cleavage) and acetal (for pH-triggered cleavage) linkers are presented. B) The multivalency effect has been investigated using hPG loaded with zinc porphyrins as photosensitizer (PS) and a systematically varied number of mannose units as targeting groups. These conjugates have been tested on their antibacterial photodynamic activity. hPG-azide with a core molecular weight (MW) of 19.5 kDa was functionalized with approx. 14 groups of the PS using the CuAAC. Subsequently, the conjugates have been further functionalized with systematically different loadings of mannose in the range of approx. 20 to 110 units. For the conjugates with approx. 58-110 mannose groups with this method the water insoluble zinc-porphyrin have been transformed to water soluble formulation. For these conjugates, the antibacterial phototoxicity against S. aureus was investigated in phosphate-buffered saline (PBS). Conjugates with approx. 69 to 110 mannose groups showed a significantly higher antibacterial activity, whereas conjugates with approx. 20 to 58 groups exhibited no activity at all. These in vitro tests showed promising results shedding light on the multivalency effect in combination with PDT. With the addition of serum, the antibacterial activity is completely quenched. We investigated this behavior with fluorescence studies performed with bovine serum albumin (BSA) suggesting protein-conjugate association as a possible cause for the loss of antibacterial activity. C) Another insight into multivalency effect came from the preparation of hPG with systematically different MW loaded with zinc porphyrins as PS and mannose groups as targeting functionality which have been tested on their antibacterial photodynamic activity. hPG-azides with a core MW between 3.7 and 100 kDa have been functionalized with approx. 4 to 108 groups of the PS [[5,10,15-tris(3-hydroxyphenyl)-20-[4-(prop-2-yn-1-ylamino)tetrafluorophenyl]porphyrinato]]-zinc(II) using the CuAAC. Additionally, polyglycerol based nanogel (nPG) with azide functionalities have been synthesized using a modified inverse nanoprecipitation. With this new method, it was possible to obtain nPGs with excess azide functionalities on the surface. The nPG-azides were further transformed to the porphyrin-nPG-conjugates via CuAAC. Subsequently, the conjugates have been further functionalized with mannose in the range of approx. 14 to 527 units. With this method the complete water insoluble zinc-porphyrin could be transformed into water-soluble formulations. Investigation of the antibacterial phototoxicity of these conjugates against S. aureus has revealed that conjugates with approx. 14 to 108 mannose groups exhibit an increasing antibacterial activity, whereas conjugates with approx. 108 to 527 groups exhibit a complete eradication of the bacteria. It has been shown that the increasing size of the porphyrin-mannose-hPG conjugates has a direct effect of antibacterial phototoxicity. With the results of the conjugates with a fixed polymer size and systematically varied mannose loadings from part B these in vitro tests show another aspect of the multivalency effect in combination with PDT. D) Porphyrin-hPG-conjugates connected via reductive and acid-cleavable linkers have been synthesized for application in PDT using CuAAC and strain-promoted alkyne-azide cycloaddition (SPAAC). Employing nucleophilic aromatic substitution (SNAr), a pentafluorophenyl substituted porphyrin has been functionalized with 1,6-diaminohexane and cystamine. In the next step, these porphyrins have been further reacted with propiolic acid or bicycle[6.1.0]non-4-yn-9-ylmethyl(4-nitrophenyl)carbonate. Porphyrin-hPG-conjugates with hPG-azide and/or hPG with acid-labile benzacetal linkers have been obtained using CuAAC or SPAAC and the previously synthesized porphyrins. With this method, the final porphyrin-hPG conjugates with acidic and/or reductive cleavable linkers were synthesized. The successful cleavage of the PS from these conjugates has been shown in different release studies. To illustrate the possible application of these conjugates selected compounds have been tested for their phototoxicity in four tumor cell lines and on the typical Gram-positive germ S. aureus.Die vorliegende Dissertation umfasst vier separate welche zum Teil aufeinander aufbauen. Darunter fällt die Synthese und Funktionalisierung diverser Porphyrine für die zukünftige Verlinkung mit Substraten, Porphyrin-hPG Konjugate mit systematisch variierter Mannose-Beladung, Porphyrin-hPG Konjugate funktionalisiert mit Mannose mit systematisch variierter Polymergröße und Porphyrin-hPG Konjugate, in welchen die Porphyrine über spaltbare Linker mit dem Polymer verbunden sind: 1. Die Reaktion von Aminen mit Pentafluorphenyl-substituierten A3B-Porphyrinen wurde benutzt um verschiedene reaktive Gruppen für die zukünftige weitere Funktionalisierung und/oder Konjugation dieser Porphyrine an andere Substrate oder Materialien einzuführen. Porphyrine mit Alkenyl, Alkinyl, Amino, Azido, Epoxid, Hydroxyl und Maleimido-Gruppen konnten so synthetisiert werden. Zum Beispiel wurde ein Alkinyl-substituiertes Porphyrin mittels CuAAC an hPG-Azid als einem möglichen Trägersystem für die PDT konjugiert. Die Photocytotoxizität von ausgewählten Porphyrinen wie auch von den Porphyrin-hPG-Konjugaten wurde in Zell Assays mit zwei Tumorzellkulturen untersucht. Für verschiedene biomedizinische Anwendungen ist eine Freisetzung der aktiven Substanz und/oder des Fluoreszenzfarbstoffes erwünscht. Deswegen wurden zusätzlich A3B-Porphyrine synthetisiert, welche spaltbare Linker enthalten, speziell Disulfid (spaltbar unter reduktiven Bedingungen) und Acetal Linker (spaltbar unter sauren pH Bedingungen). 2. In diesem Teil der Arbeit wurde die antibakterielle photodynamische Aktivität von hPG beladen mit Zink Porphyrin als PS und Mannose Gruppen untersucht. hPG mit einer molekularen Masse MW von 19,5 kDa wurde mittels der CuAAC mit ~14 Molekülen des PS {5,10,15-Tris(3-hydroxyphenyl)-20-[4-(prop-2-yn-1-ylamino)tetrafluorphenyl]porphyrinato}Zink(II) funktionalisiert. Diese Nanopartikel Konjugate wurden weiterhin systematisch mit verschiedenen Beladungen von Mannose (zwischen ~20 und 110) Gruppen funktionalisiert. Mit höheren Mannose Beladungen (zwischen ~58 und 110 Gruppen) konnte der wasserunlösliche Zink-Porphyrin-PS in eine wasserlösliche Form überführt werden. Anschließend wurde die antibakterielle Phototoxizität der Konjugate gegen S. aureus (als typischer Gram positiver Keim) in PBS untersucht. Es zeigte sich, dass Konjugate mit ~70 bis 110 Mannose Einheiten eine signifikante antibakterielle Aktivität aufwiesen, wohingegen Konjugate mit ~20 bis 60 Einheiten nicht antibakteriell aktiv waren. Die Resultate geben einen Einblick in den Multivalenzeffekt in Kombination mit PDT. Bei Zugabe von Serum zu den Bakterienkulturen verschwand die antibakterielle Phototoxizität vollständig. Fluoreszenzstudien der Konjugate in Gegenwart von zunehmenden BSA Konzentrationen deuteten auf eine Protein-Konjugat-Assoziation als mögliche Ursache hin. 3. hPGs mit unterschiedlichen molekularen Massen bzw. Größen wurden mit Zink-Porphyrin (als PS) und Mannose Gruppen funktionalisiert. Diese Konjugate wurden anschließend auf ihre antibakterielle Phototoxizität untersucht. hPGs zwischen 3,7 und 100 kDa wurden mit ~4 bis ~108 Molekülen des PS {5,10,15-Tris(3-hydroxyphenyl)-20-[4-(prop-2-yn-1-ylamino)tetrafluorphenyl]porphyrinato}-Zink(II) mittels CuAAC funktionalisiert. Zusätzlich zu diesen Konjugaten wurde ein Polyglycerol Nanogel mit Hilfe einer neuen inversen Nanofällung hergestellt. Diese modifizierte Technik erlaubt die Synthese von nPGs mit einem Überschuss von Aziden auf der Oberfläche. Die erhaltenen nPG-Azide wurden dann weiter benutzt um die Porphyrin-nPG Konjugate mittels CuAAC zu erhalten. Diese Nanopartikel wurden anschließend weiter mit zwischen ~14 und ~527 Mannose Einheiten (als Targeting Gruppen) funktionalisiert. Auch mit dieser Methode konnte der wasserunlösliche Zink-Porphyrin-PS bei allen Konjugaten in eine wasserlösliche Formulierung überführt werden. Die antibakterielle Phototoxizität der Konjugate gegen Kulturen von S. aureus wurde in PBS untersucht. Es zeigte sich, dass Konjugate zwischen ~14 und ~108 Mannose Einheiten eine zunehmende antibakterielle Aktivität aufwiesen. Die Konjugate mit ~108 und ~527 Gruppen zeigten eine vollständige Auslöschung der Bakterien. Diese Resultate weisen einen weiteren Aspekt des Multivalenzeffektes hin: Der Einfluss unterschiedlicher molekularer Massen der Konjugaten in Kombination mit der PDT. 4. Dieser letzte Teil der Arbeit befasst sich mit hPG-Porphyrin-Konjugaten mit spaltbaren Linkern. Dafür wurde ein Syntheseweg für solche Porphyrin-hPG Konjugate mittels CuAAC und SPAAC entwickelt. Im ersten Schritt erfolgte eine Funktionalisierung von 10,15,20-Tris(3-hydroxyphenyl)-5-(2,3,4,5,6-pentafluorphenyl)porphyrin durch nukleophile aromatische Substitution mit 1,6-Diaminohexan bzw. Cystamin funktionalisiert. Die erhaltenen Porphyrine wurden weiter mit Propiolsäure oder Bicyclo[6.1.0]non-4-yn-9-ylmethyl(4-nitrophenyl)karbonat umgesetzt. Parallel wurden Azid-funktionalisierte hPGs und hPGs mit säurelabilen Benzacetal Linkern synthetisiert und mit den Porphyrinen und Mannose mittels CuAAC und SPAAC verknüpft. Auf diese Weise konnten Porphyrin-hPG Konjugate mit sauer und/oder reduktiv spaltbaren Linkern erhalten werden. Die Abspaltung des PS vom Carrier konnte in verschiedenen Freisetzungsstudien gezeigt werden. Um eine mögliche Anwendung für die PDT zu prüfen, wurde die Phototoxität von ausgewählten Konjugaten an verschiedenen Tumorzelllinien und Kulturen von S. aureus getestet. Hierbei erwiesen sich einige der Konjugate als gut wirksam gegen S. aureus, insbesondere eines der Konjugate mit einer Kombination aus sauer und reduktiv spaltbarem Linker

Testing in Big Data: An Architecture Pattern for a Development Environment for Innovative, Integrated and Robust Applications

Big Data is a crucial pillar for many of today’s newly emerging business models. Areas of application range from consumer analysis over medicine to fraud detection. All of those domains require reliable software. Even though imperfect results are accepted in Big Data software, bugs and other defects can have drastic consequences. Therefore, in this paper, the software engineering sub discipline of testing is addressed. Big Data exhibits characteristics which differentiate its processing software from those that process traditional workloads. Consequently, an architecture pattern for testing that can be integrated into development environments for Big Data software is proposed. The paper features a detailed description of the artifact as well as a preliminary plan for evaluation

Facing Big Data System Architecture Deployments: Towards an Automated Approach Using Container Technologies for Rapid Prototyping

Within the last decade, big data became a promising trend for many application areas, offering immense potential and a competitive edge for various organizations. As the technical foundation for most of today´s data-intensive projects, not only corresponding infrastructures and facilities but also the appropriate knowledge is required. Currently, several projects and services exist that not only allow enterprises to utilize but also to deploy related technologies and systems. However, at the same time, the use of these is accompanied by various challenges that may result in huge monetary expenditures, a lack of modifiability, or a risk of vendor lock-ins. To overcome these shortcomings, in the contribution at hand, modern container and task automation technologies are used to wrap complex big data technologies into re-usable and portable resources. Those are subsequently incorporated in a framework to automate the deployment of big data architectures in private and limited resources

The Linkage to Business Goals in Data Science Projects

Modern data analytics equips businesses to make data-driven decisions by revealing patterns and insights that enhance strategic planning, operational efficiency, and process optimization. Its applications encompass personalized marketing through customer segmentation, predictive modelling for fraud detection, and enhancing security. A significant methodology in this realm is the Cross-Industry Standard Process for Data Mining (CRISP-DM), where the Business Understanding phase aims to ensure data science projects align with overarching business goals. However, challenges arise when these business objectives are ambiguous, ill-defined, or evolving. The complexity of data analytics projects underscores the need for domain expertise and robust collaboration between data scientists, business stakeholders, and domain experts. The imperative is to bridge the technical and business perspectives, manage expectations, and define project scopes. The short paper at hand addresses the question how data analytic goals can systematically align with business objectives in data science projects. By incorporating methods from Enterprise Architecture Management, we propose a structured approach for goal determination in data science projects, ensuring business and data mining objectives are seamlessly integrated

Drivers of bat fly diversity and prevalence of six Rhinolophus bat species in southern Africa.

Masters of Science in Zoology. University of KwaZulu-Natal. Durban, 2016.Demographic and ecological characteristics render obligate cave roosting bats highly susceptible to infestation by ectoparasites. However, the patterns and factors of ectoparasite loads among bat host species are understudied, particularly in the Old World. I tested predictions of habitat heterogeneity, host sex, body size hypotheses to explain parasitic bat fly (Streblidae and Nycteribiidae) abundance, morphospecies richness and prevalence on six Rhinolophus bat species in southern Africa. I sampled and classified 930 bat flies to six morphospecies (3 streblids and 3 nycteribiids) captured on 333 bats at 20 sites in eight biomes. In support of the habitat heterogeneity hypothesis, there were significantly positive relationships between habitat heterogeneity and bat fly abundance, morphospecies richness and prevalence. In support of the host body size hypothesis, there were significantly positive relationships between host body condition and bat fly abundance, morphospecies richness and prevalence. By contrast, there was little evidence that parasitic flies preferred either male or female bats. Recursive partitioning analysis showed that the most significant predictor of bat fly abundance and morphospecies richness was habitat heterogeneity, specifically the number of land cover classes surrounding bat roosts. My results suggest that land use and biome characteristics at the meso-scale, and to a lesser degree biotic processes at the local scale, mediate bat fly abundance and morphospecies richness on rhinolophid bats. Specifically, structurally heterogeneous and complex habitats increase the number of niches available for bat species as well as their prey, which, in turn, may favour diverse bat fly populations. Thus, factors responsible for driving bat diversity may also drive bat fly diversity. Future studies should focus on other families of cave-roosting bats, as well as endoparasites, to better understand the mechanisms responsible for ectoparasitism in Old World bats

Exploring the Applicability of Test Driven Development in the Big Data Domain

Big data analytics and the according applications have gained huge importance in daily life. This results on the one hand from their versatility and on the other hand from their capability to greatly improve an organization’s performance when utilized appropriately. However, despite their prevalence and the corresponding attention through practitioners as well as the scientific world, the actual implementation still remains a challenging task. Therefore, without the adequate testing, the reliability of the systems and thus the obtained outputs is uncertain. This might reduce their utilization, or even worse, lead to a diminished decision-making quality. The publication at hand explores the adoption of test driven development as a potential approach for addressing this issue. Subsequently, using the design science research methodology, a microservice-based test driven development concept for big data (MBTDD-BD) is proposed. In the end, possible avenues for future research endeavours are indicated

Towards a Blockchain Technology Framework – Literature Review on components in blockchain implementations

The goal of this work is to obtain a framework that represents the technological core aspects of blockchain, separated into components, their subcategories and related basic technologies. In order to gain a holistic view of blockchain, with the help of the framework, technologies constructs should be made identifiable as blockchain. For this purpose, a literature review will be conducted to investigate previous approaches to the component-wise division of blockchain technologies. Subsequently, a literature analysis will be conducted in which five established blockchain systems will be analysed and their implementations will be assigned to the general components. For evaluation, a further sixth blockchain technology is used to confirm the basic framework. It becomes apparent that the framework allows a classification of blockchain systems into technologies. The framework has potential for expansion by adding further technology features to make the framework even more useful

Decentralised Autonomous Organisations in Organisational Design Theory

As Decentralised Autonomous Organisation (DAO) is a new emerging form of organisation with unrevealed characteristics, this study examines how DAO can be classified in terms of organisational design theory and provides an overview of its characteristics. The investigation could further provide guidance on what types of organisations can be easily transformed into a DAO. A deeper look into the knowledge base on organisational design theory and organisational forms is essential as well as characteristic properties. In regard of DAO, the underlying concepts of blockchain and smart contracts are marked as they serve a better understanding, which is needed to characterise DAO. An analysis of DAO will specify how they are classified in terms of organisational design and how they differ compared to traditional organisation forms, which will be displayed in an overview. The results are discussed and concluded, also comprising the highlighting of potential paths for future work

Low-code Development Platform Usage: Towards Bringing Citizen Development and Enterprise IT into Harmony

The ongoing digitization of our world leads to many areas of our lives being more pleasant and improved. New technologies and paradigms are emerging to support the development of software and systems. Their proliferation not only leads to higher complexity of potential solutions, but also to the problem of finding qualified people. Especially enterprises, which are constantly confronted with this problem, are increasingly considering low-code development platforms (LCDP) to allow the development of software by inexperienced and untrained citizen developers. However, at this point, non-functional requirements, such as performance and security, can require a thorough system understanding. In this work, we identify issues that may occur when citizen developers use LCDPs, allowing to deduce success factors for their implementation. Eventually, this shall help decision makers when introducing LCDPs into their environments

Automation of Customer Initiated Back Office Processes: A Design Science Research Approach to link Robotic Process Automation and Chatbots

While the emerging technology of robotic process automation is primarily suitable for back office processes, companies use traditional chatbots to support customer interaction in the front office. However, customer requests that require more than written information usually demand an employee to execute an internal process. This paper summarizes the results of a technical design process for a combination of both technologies. After an introduction on both topics, the findings of a literature review regarding existing approaches are outlined. The development of the IT artefact is then carried out according to the design science research methodology. In particular, the research focuses on the constitution of a design theory in consideration of criteria that are found to be important for a purposeful appearance to the external user. After a proof of concept by testing the developed artefact and a summary of the results, an outlook on possible future developments is provided