| 1. |
- Borell, Anton, 1991-
(författare)
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In Between Competing Ideals : On the Relationships among Accounting, NPM, and Welfare
- 2021
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- We know that public organizations today are expected to manage activities on multiple, and somewhat incongruent ideals. Since the development of the New Public Management (NPM) wave, public organizations have been increasingly subject to governance by markets, economic frugality and outputs for the purpose of improving efficiency. At the same time, they are also required to manage their daily work according to the ideals of needs, equity and professional expertise, which have traditionally been foundational to the realization of a welfarian agenda. Together, these requirements make up dual expectations of public organizations as well as the accounting technologies used to manage them: a form of paradoxical management that seeks to measure, evaluate and document both efficiency and welfare. But how do actors realize these plural ideals in practice? And what is the role of accounting in this? These are some of the aspects and problems with control that lie at the very heart of this dissertation.Based on 41 interviews and 28 observations in a Swedish school context, this dissertation shows how the paradoxical expectations become manifest in the design and use of accounting technologies. On the one hand, it shows how school actors attempt to integrate welfare and NPM in practice by relying on accounting technologies, which results in hybrid outcomes. On the other hand, the dissertation also shows the fragility of such a style of governance, which continuously leads to failures and breakdowns that actors have to readdress as a consequence of attempting to achieve competing ideals by relying on accounting.By leaning on governmentality as a metatheoretical framework, this dissertation contributes to some new understandings of control in public sector contexts. First, in contrast to the literature on colonization, which suggests that the expansive use of accounting is a direct threat to welfarian ideals, this dissertation shows that accounting can equally serve as a means for welfarian ideals to eclipse NPM. Second, in contrast to the stream of separation that treats the loose or minimal use of accounting as a mechanism for sealing off welfare from NPM-led intrusions, the dissertation shows that even a marginal or loose application of accounting can in fact be conceived as a threat to welfarian ideals. Third, this dissertation also provides additional understanding in relation to the hybridity approach, not only by illuminating how NPM and welfare were brought together through various hybrids, but also by showing how these hybrids were facilitated by accounting technologies, whose functions served to make integration a practically viable outcome amenable to both breakdowns and reintegration. Overall, the dissertation thus contributes to current knowledge on public sector governance in general, and more specifically about the relationships among accounting, NPM, and welfare, in terms of their entanglement, disentanglement, interdependencies, and modes of reproduction in public sector life, a gap also echoed by recent calls in research.
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| 2. |
- Nunez-Otero, Carlos, 1992-
(författare)
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Novel inhibitors of Chlamydia trachomatis virulence
- 2020
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Chlamydia trachomatis is an obligate intracellular bacterium that infects over 100 million people globally every year. Chlamydia infections can be persistent, cause infertility and blindness, adding an economical burden in the healthcare systems. Moreover, Chlamydia infections are treated with broad-spectrum antibiotics that contribute to the selection of antibiotic resistant bacteria in the commensal flora. For this reason, novel compounds with specificity against C. trachomatis would be important for treatment of Chlamydia infections.We have developed a new class of substituted 2-pyridone amides that inhibited development of C. trachomatis. While bacterial growth was only affected to a limited extent, the produced progeny bacteria had impaired capacity to infect new cells. The compounds presented no toxicity in human or mouse cell lines and they did not inhibit growth of bacteria from the normal flora. Structure activity relationship (SAR) development of 2-pyridones lead to compounds with effect at nanomolar concentrations. Further modifications of the C3 part of the molecules resulted in isostere compounds with even a higher potency. By exploring the C8 position, we observed that methylsulfonamide substituents improved the pharmacokinetic properties and enabled oral uptake in mice. This discovery opens the door for oral treatment.Among 2-pyridone amides, KSK213 was one of the most potent and we investigated the mode of action on the life cycle of C. trachomatis. KSK213 reduced transcription by the end of the developmental cycle and upon infection of new host cells. Mutations in RNA helicase and RNAse III genes, involved in transcription, mediated resistance to KSK213. It also attenuated the infectivity in a mouse vaginal infection model. To further explore the molecular target for 2-pyridone amides in Chlamydia, we used a custom synthesized probe for affinity chromatography approaches.Here we show that 2-pyridones are potent non-toxic inhibitors of C. trachomatis that can be chemically modified to increase potency and enable oral bioavailability. These molecules have the potential to treat and prevent Chlamydia infections without affecting the normal flora.
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| 3. |
- Knutsson, Sofie, 1983-
(författare)
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Towards Mosquitocides for Prevention of Vector-Borne Infectious Diseases : discovery and Development of Acetylcholinesterase 1 Inhibitors
- 2016
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Diseases such as malaria and dengue impose great economic burdens and are a serious threat to public health, with young children being among the worst affected. These diseases are transmitted by mosquitoes, also called disease vectors, which are able to transmit both parasitic and viral infections. One of the most important strategies in the battle against mosquito-borne diseases is vector control by insecticides and the goal is to prevent people from being bitten by mosquitoes. Today’s vector control methods are seriously threatened by the development and spread of insecticide-resistant mosquitos warranting the search for new insecticides. This thesis has investigated the possibilities of vector control using non-covalent inhibitors targeting acetylcholinesterase (AChE); an essential enzyme present in mosquitoes as well as in humans and other mammals. A key requirement for such compounds to be considered safe and suitable for development into new public health insecticides is selectivity towards the mosquito enzyme AChE1. The work presented here is focused on AChE1 from the disease transmitting mosquitoes Anopheles gambiae (AgAChE1) and Aedes aegypti (AaAChE1), and their human (hAChE) and mouse (mAChE) counterparts. By taking a medicinal chemistry approach and utilizing high throughput screening (HTS), new chemical starting points have been identified. Analysis of the combined results of three different HTS campaigns targeting AgAChE1, AaAChE1, and hAChE allowed the identification of several mosquito-selective inhibitors and a number of compound classes were selected for further development. These compounds are non-covalent inhibitors of AChE1 and thereby work via a different mechanism compared to current anti-cholinergic insecticides, whose activity is the result of a covalent modification of the enzyme. The potency and selectivity of two compound classes have been explored in depth using a combination of different tools including design, organic synthesis, biochemical assays, protein X-ray crystallography and homology modeling. Several potent inhibitors with promising selectivity for the mosquito enzymes have been identified and the insecticidal activity of one new compound has been confirmed by in vivo experiments on mosquitoes. The results presented here contribute to the field of public health insecticide discovery by demonstrating the potential of selectively targeting mosquito AChE1 using non-covalent inhibitors. Further, the presented compounds can be used as tools to study mechanisms important in insecticide development, such as exoskeleton penetration and other ADME processes in mosquitoes.
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| 4. |
- Kulén, Martina, 1988-
(författare)
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New alternatives to combat Listeria monocytogenes and Chlamydia trachomatis : Design, synthesis, and evaluation of substituted ring-fused 2-pyridones as anti-virulent agents
- 2018
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Antibiotic resistance has become a global health burden with the number of resistant bacteria continuously increasing. Antibiotic drugs act by being either bactericidal (killing bacteria) or bacteriostatic (inhibiting growth of bacteria). However, these modes of action increase the selective pressure on the bacteria. An alternative treatment strategy to antibiotics is anti-virulence therapies that inhibits virulence of the pathogenic bacteria. The term “virulence” summarises a number of factors that the bacteria need to colonise a new niche and as a consequence its ability to infect and cause diseases. By inhibiting virulence, instead of killing, the selective pressure on the bacteria can be reduced and consequently decreases the rapid development of resistance. This thesis describes two projects focusing on development of anti-virulence agents, with the ring-fused 2-pyridone scaffold as the central character, targeting the bacteria Listeria monocytogenes and Chlamydia trachomatis.The first project is targeting L. monocytogenes, which is the cause for listeriosis in humans. This can develop into life-threatening encephalitis and meningitis as well as cause severe complications for developing fetus. The target in L. monocytogenes is the transcriptional regulator PrfA that control almost all virulence factors in this bacterium. We have designed and synthesised potent substituted ring-fused 2-pyridones, which at low micromolar concentrations block activation of the virulence regulator PrfA and thus attenuate the bacterial infection. Co-crystallisation of the active ring-fused 2-pyridones with PrfA resulted in determination of the exact substance interaction site in the protein. This facilitated further structure-based design that resulted in improved compounds capable of attenuating L. monocytogenes in an in vivo model.The second project targets C. trachomatis, which is the causative agent behind the most common sexually transmitted infection as well as the eye infection trachoma. By structure-activity relationship analysis of previously tested ring-fused 2-pyridones, we have designed and synthesised non-hydrolysable ring-fused 2-pyridone amide isosteres. The most potent analogues inhibit C. trachomatis infectivity at low nanomolar concentrations, without showing host cell toxicity or affecting the viability of commensal microbiota. Introduction of heteroatom substituents at specific sites of the ring-fused 2-pyridone scaffold, resulted in improved pharmacokinetic properties of the analogues and further evaluation in vivo was performed.
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| 5. |
- Oelker, Melanie, 1988-
(författare)
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Disarming bacteria : a structure-based approach to design an anti-virulence drug against Listeria monocytogenes
- 2021
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Antibiotic resistances are one of the biggest threats to global health and if we don’t change our behavior and way of using antibiotics we will end up in a ‘post-antibiotic era’, in which common infections and minor injuries can once kill again and up to 10 million deaths per year may occur by 2050. Therefore, there is a high need for new anti-bacterial drugs, especially of alternatives to existing antibiotics with already described resistances. Classical antibiotics target the essential processes of survival and growth in bacteria and therefore put a high selective pressure on them to develop resistances. In contrast, the ability to infect or damage a host, the virulence, is less essential for bacteria. Thus, targeting the virulence is supposed to cause a lower selective pressure and this alternative mode-of-action could help to decelerate the development of antibiotic resistances.The aims in this work were to proceed with the structure-based design of an anti-virulence drug against the food-borne pathogen Listeria monocytogenes, but also to deepen our understanding of the complex regulation system for the virulence of this bacterium. PrfA, the master regulator of virulence in Listeria monocytogenes, is a member of a large family of bacterial transcription factors, which are regulated by a conformational change and allosteric modulation by different regulator molecules. Furthermore, its critical role in virulence regulations makes is a suitable target for an anti-virulence drug. In this work new lead compounds based on the previously identified ring-fused 2-pyridone scaffold were designed, synthesized and analyzed by different biological, biophysical, computational and structural biology methods. Three new binding sites and binding modes of these compounds in PrfA were evaluated for their potential use in future designs and a compound with improved activity was identified. In a second study another structurally different lead compound was discovered to inhibit PrfA. Furthermore, the studies on proposed natural regulators of PrfA uncovered the underlying mechanism for the virulence regulation by the peptide signature of the environment and in a follow-up study the structural basis of the binding of inhibitory peptides to PrfA was further investigated. Finally, a structural review on all available structure of PrfA provided more insights into the allosteric regulation mechanism of PrfA activity.This work will hopefully support in the successful development of an anti-virulence drug against Listeria monocytogenes and thus contribute to the reduction of the problem of antibiotic resistances.
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| 6. |
- Adolfsson, Dan E., 1989-
(författare)
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Synthesis of Ring-fused Peptidomimetics : Interacting with Amyloid Fibrils
- 2020
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Parkinson's and Alzheimer's disease are the two most common neurological disorders in humans. Both conditions involve progressive death of neurons in the central nervous system, decline in bodily functions and eventually (and invariably), death. So far, no cure exists and the available treatments can only ease symptoms. Despite substantial investments in research, the biomolecular processes are still far from fully understood. However, both diseases are associated with formation of fibrillar protein aggregates called amyloid deposits. Whereas Alzheimer’s disease involves aggregation of the Tau and Amyloid β proteins, α-Synuclein fibrilization plays a key role in Parkinson's disease. Although they are chemically distinct, the deposits consist of protein fibres with similar morphology and fold. Small molecules, such as the thiazoline fused 2-pyridones herein presented, can interfere with the formation of amyloid fibres, or bind to them. Besides having potential for diagnostication and treatment, such small molecules constitute valuable tool compounds in future research, to unravel the mechanisms of amyloid formation and pathology. The first step towards successful treatment, diagnostication and prevention of Alzheimer's and Parkinson's disease is understanding the causes and underlying mechanisms better. This thesis narrates the synthesis and development of novel chemical structures: multi ring fused peptidomimetics with the ability to bind mature amyloid fibrils, consisting of α-Synuclein or Amyloid β. The first project (articles I, III and VI) describes method development for the extension of bicyclic thiazolino 2-pyrdiones by fusion with aromatic nitrogen heterocycles, which enables the desired amyloid binding properties. Derivatisations of the newly generated central scaffold, and variation of the multiple attached substituents, were subsequently performed in efforts to improve binding strength and solubility, and gain selectivity towards certain fibrils. One of the most promising amyloid fibril binders was evaluated in a human cell line and in mice, and found to be protective against accelerator induced neurotoxicity. One pyrimidine fused compound moreover indicated potent inhibition of Amyloid b aggregation. The second project (articles II, IV and V) focuses on development of methods to modify the thiazoline ring. Ring opening induced by electrophiles generates N-alkenyl 2-pyridones but decreases amyloid binding potency. Introduction of a cyclobutane moiety fused with the thiazoline ring is better tolerated, and adds a terminal alkene moiety that can be exploited in future chemical modifications. Expansion of the five membered thiazoline ring to a six membered dihydrothiazine ring, equipped with a nitrophenyl substituent, provides compounds with enhanced fibril binding capacity, which further inhibits Amyloid β fibril formation in vitro. Taken together, the synthetic methodologies allow construction and late stage modification of complex fused heterocycles, with several points of variation. Thus, the developed methods may be of future value in our laboratories and elsewhere.
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| 7. |
- Aliashkevich, Alena, 1990-
(författare)
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Molecular mechanisms and biological consequences of the production of non-canonical D-amino acids in bacteria
- 2021
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Most bacteria possess a vital net-like macromolecule – peptidoglycan (PG). PG encases bacteria around the cytoplasmic membrane to withstand the high internal turgor pressure and thereby protect the cell from bursting. In addition, PG is a major morphological determinant of bacteria being both required and sufficient to maintain cell shape. During cell growth PG hydrolysis and synthesis are tightly controlled to keep proper cell shape and integrity at all times. Given the essentiality of PG for bacterial growth and survival, the synthesis of this polymer is a major target of many natural and synthetic antibiotics (e.g. penicillins, glycopeptides).For a long time, PG composition was considered to be conserved and static, however it’s now being recognized as a dynamic and plastic macromolecule. The structure and chemistry of PG is influenced by a myriad of environmental cues that include interkingdom/interspecies interactions. Recently, it was found that a wide set of non-canonical D-amino acids (D-amino acids different from D-Ala and D-Glu, NCDAAs) are produced and released to the extracellular milieu by diverse bacteria. In Vibrio cholerae these NCDAAs are produced by broad-spectrum racemase enzyme (BsrV) and negatively regulate PG synthesis through their incorporation into PG. We have shown that in addition to D-Met and D-Leu, which were reported previously, V. cholerae also releases high amounts of D-Arg, which inhibits a broader range of phylogenetically diverse bacteria. Thus, NCDAAs affect not only the producer, but might target other species within the same environmental niche. However, in contrast to D-Met, D-Arg targets cell wall independent pathways. We have shown that non-proteinogenic amino acids also can be racemized by Bsr. A plant amino acid L-canavanine (L-CAN) is converted into D-CAN by a broad-spectrum amino acid racemase (BSAR) of the soil bacterium Pseudomonas putida and subsequently released to the environment. D-CAN gets highly incorporated into the PG of Rhizobiales (such as Agrobacterium tumefaciens, Sinorhizobium meliloti) thereby affecting the overall PG structure, bacterial morphogenesis and growth fitness. We found that detrimental effect of D-CAN in A. tumefaciens can be suppressed by a single amino acid substitution in the cell division PG transpeptidase penicillin-binding protein 3a (PBP3a). Rhizobiales are a polar-growing species that encode multiple LD-transpeptidases (LDTs), enzymes that normally perform PG crosslinking, but that can also incorporate NCDAAs into termini of the PG peptides. As these species incorporate high amounts of D-CAN in their PG, we hypothesized that LDTs might represent the main path used by NCDAAs to edit A. tumefaciens’ PG and cause their detrimental effects. Therefore, we decided to further explore the significance of LDT proteins for growth and morphogenesis in A. tumefaciens. While in the Gram-negative model organism E. coli LDT proteins are non-essential under standard laboratory conditions, we found that A. tumefaciens needs at least one LDT for growth out of the 14 putative LDTs encoded in its genome. Moreover, clustering the LDT proteins based on their sequence similarity revealed that A. tumefaciens has 7 LDTs that are exclusively present among Rhizobiales. Interestingly, the loss of this group of LDTs (but not the rest) leads to reduced growth, lower PG crosslinkage and rounded cell phenotype, which suggests that this group of Rhizobiales- specific LDTs have a major role in maintaining LD-crosslinking homeostasis, which in turn is important for cell elongation and proper shape maintenance in A. tumefaciens.
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| 8. |
- Andersson, Hans, 1978-
(författare)
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Reaction Between Grignard reagents and Heterocyclic N-oxides : Synthesis of Substituted Pyridines, Piperidines and Piperazines
- 2009
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- This thesis describes the development of new synthetic methodologies for preparation of bioactive interesting compounds, e.g. substituted pyridines, piperidines or piparazines. Thesecompounds are synthesized from commercially available, cheap and easily prepared reagents, videlicet the reaction between Grignard reagents and heterocyclic N-oxides. The first part of this thesis deals with an improvement for synthesis of dienal-oximes and substituted pyridines. This was accomplished by a rapid addition of Grignard reagents to pyridine N-oxides at rt. yielding a diverse set of substituted dienal-oximes. During these studies, it was observed that the obtained dienal-oxmies are prone to ring-close upon heating. By taking advantage of this, a practical synthesis of substituted pyridines was developed. In the second part, an ortho-metalation of pyridine N-oxides using Grignard reagents is discussed. The method can be used for incorporation of a range of different electrophiles, including aldehydes, ketones and halogens. Furthermore, the importance for incorporation of halogens are exemplified through a Suzuki–Miyaura coupling reaction of 2-iodo pyridine N-oxides and different boronic acids. Later it was discovered that if the reaction temperature is kept below -20 °C, the undesired ringopening can be avoided. Thus, the synthesis of 2,3-dihydropyridine N-oxide, by reacting Grignard reagents with pyridine N-oxides at -40 °C followed by sequential addition of aldehyde or ketone, was accomplished. The reaction provides complete regio- and stereoselectivity yielding trans-2,3-dihydropyridine N-oxides in good yields. These intermediate products could then be used for synthesis of either substituted piperidines, by reduction, or reacted in a Diels–Alder cycloaddtion to give the aza-bicyclo compound. In the last part of this thesis, the discovered reactivity for pyridine N-oxides, is applied on pyrazine N-oxides in effort to synthesize substituted piperazines. These substances are obtained by the reaction of Grignard reagents and pyrazine N-oxides at -78 °C followed by reduction and protection, using a one-pot procedure. The product, a protected piperazine, that easily can be orthogonally deprotected, allowing synthetic modifications at either nitrogens in a fast and step efficient manner. Finally, an enantioselective procedure using a combination of PhMgCl and (-)-sparteine is discussed, giving opportunity for a stereoselective synthesis of substituted piperazines.
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| 9. |
- Arja, Katriann, 1985-
(författare)
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Multimodal Porphyrin-Based Conjugates : Synthesis and characterization for applications as amyloid ligands, photodynamic therapy agents and chiroptical materials
- 2018
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Organic compounds that interact both with certain biological targets and display specific photophysical properties can be utilized as molecular tools to visualize and possibly effect disease related processes taking place in living organisms. In this regard, porphyrins are a class of naturally occurring molecules that possess intriguingly interesting photophysical properties where they can act as luminescent probes by emitting detectable light, as well as photosensitizers in the light mediated therapy called photodynamic therapy. In this thesis, the porphyrin structure has been synthetically combined with other molecule classes to achieve compounds with desirable multimodal characteristics.Firstly, luminescent conjugated oligothiophenes (LCOs) that have extensively, and with great success, been utilized as fluorescent ligands for amyloid formations, have been conjugated to porphyrins to render oligothiophene porphyrin hybrids (OTPHs) comprising two optically active modalities. When applied as fluorescent amyloidophilic dyes for visualization of amyloid-β (Aβ), one of the pathological hallmarks in Alzheimer’s disease, an enhanced optical assignment of distinct aggregated forms of Aβ was afforded. Thus, properly functionalized OTPHs could give us more information about pathological processes underlying devastating disorders, such as Alzheimer’s disease. In addition, the OTPHs can be associated with synthetic peptides inducing peptide folding into certain three-dimensional helical structures giving rise to novel optically active materials.Secondly, this thesis also embraces porphyrins’ potential as photosensitizers in photodynamic therapy to kill cancer cells. Grounded on the prerequisites for an optimal photosensitizer, we designed porphyrin-based conjugates equipped with common carbohydrates for improved cancer cell selectivity and with a fluorinated glucose derivative, 2-fluoro 2-deoxy glucose, for advantageous metabolism in cancer cells. Furthermore, incorporation of a radioisotopic fluorine-18 atom into the glycoporphyrins could give the means for diagnostic use of the conjugates in positron emission tomography (PET).In order to tether together the above-mentioned molecular moieties in a controlled fashion, we developed a robust synthetic strategy for asymmetrical functionalization of porphyrin core. The method involves chlorosulfonation of this otherwise inert tetrapyrrolic structure, followed by alkynylation. Parallelly to amide coupling reactions, copper(I)-catalyzed alkyne azide cycloaddition is used for fast and high-yielding late-stage conjugations. Overall, this thesis demonstrates how combining different molecular moieties in synthetic organic chemistry yields novel molecules with combined and improved multimodal properties for biological and medicinal applications, guided by the design-by-function methodology.
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| 10. |
- Bengtsson, Christoffer, 1979-
(författare)
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Synthesis of substituted Ring-Fused 2-Pyridones and applications in chemical biology
- 2013
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Antibiotics have been extensively used to treat bacterial infections since Alexander Fleming’s discovery of penicillin 1928. Disease causing microbes that have become resistant to antibiotic drug therapy are an increasing public health problem. According to the world health organization (WHO) there are about 440 000 new cases of multidrug-resistant tuberculosis emerging annually, causing at least 150 000 deaths. Consequently there is an immense need to develop new types of compounds with new modes of action for the treatment of bacterial infections.Presented herein is a class of antibacterial ring-fused 2-pyridones, which exhibit inhibitory effects against both the pili assembly system in uropathogenic Escherichia coli (UPEC), named the chaperone usher pathway, as well as polymerization of the major curli subunit protein CsgA, into a functional amyloid fibre. A pilus is an organelle that is vital for the bacteria to adhere to and infect host cells, as well as establish biofilms. Inhibition of the chaperone usher pathway disables the pili assembly machinery, and consequently renders the bacteria avirulent.The focus of this work has been to develop synthetic strategies to more efficiently alter the substitution pattern of the aforementioned ring-fused 2-pyridones. In addition, asymmetric routes to enantiomerically enriched key compounds and routes to compounds containing BODIPY and coumarin fluorophores as tools to study bacterial virulence mechanisms have been developed. Several of the new compounds have successfully been evaluated as antibacterial agents. In parallel with this research, manipulations of the core structure to create new heterocycle based central fragments for applications in medicinal chemistry have also been performed.
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| 11. |
- Chorell, Erik, 1980-
(författare)
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Pilicides and Curlicides : Design, synthesis, and evaluation of novel antibacterial agents targeting bacterial virulence
- 2010
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- New strategies are needed to counter the growing problem of bacterial resistance to antibiotics. One such strategy is to design compounds that target bacterial virulence, which could work separately or in concert with conventional bacteriostatic or bactericidal antibiotics. Pilicides are a class of compounds based on a ring-fused 2-pyridone scaffold that target bacterial virulence by blocking the chaperone/usher pathway in E. coli and thereby inhibit the assembly of pili. This thesis describes the design, synthesis, and biological evaluation of compounds based on the pilicide scaffold with the goal of improving the pilicides and expanding their utility. Synthetic pathways have been developed to enable the introduction of substituents at the C-2 position of the pilicide scaffold. Biological evaluation of these compounds demonstrated that some C-2 substituents give rise to significant increases in potency. X-ray crystallography was used to elucidate the structural basis of this improved biological activity. Furthermore, improved methods for the preparation of oxygen-analogues and C-7 substituted derivatives of the pilicide scaffold have been developed. These new methods were used in combination with existing strategies to decorate the pilicide scaffold as part of a multivariate design approach to improve the pilicides and generate structure activity relationships (SARs). Fluorescent pilicides were prepared using a strategy where selected substituents were replaced with fluorophores having similar physicochemical properties as the original substituents. Many of the synthesized fluorescent compounds displayed potent pilicide activities and can thus be used to study the complex interactions between pilicide and bacteria. For example, when E. coli was treated with fluorescent pilicides, it was found that the compounds were not uniformly distributed throughout the bacterial population, suggesting that the compounds are primarily associated to bacteria with specific properties. Finally, by studying compounds designed to inhibit the aggregation of Aβ, it was found that some compounds based on the pilicide scaffold inhibit the formation of the functional bacterial amyloid fibers known as curli; these compounds are referred to as 'curlicides'. Some of the curlicides also prevent the formation of pili and thus exhibit dual pilicide-curlicide activity. The potential utility of such 'dual-action' compounds was highlighted by a study of one of the more potent dual pilicide-curlicides in a murine UTI model were the compound was found to significantly attenuate virulence in vivo.
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| 12. |
- Draxler, Helena
(författare)
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Project Support i svensk socialtjänst : En genomförbarhetsstudie av ett föräldrastödsprogram för våldsutsatta föräldrar och deras barn som utvecklat beteendeproblematik
- 2017
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Licentiatavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Projekt Support (PS) är ett föräldrastödsprogram som har utvecklats i USA för mammor och barn som utsatts för våld i nära relationer där barnen också utvecklat känslomässiga och beteendemässiga problem. Syftet med denna licentiatuppsats är att undersöka hur interventionen mottogs av behandlare och föräldrar inom ramen för svensk socialtjänst med hänsyn till skillnaderna mellan länderna.Två studier genomfördes. I den första studien (studie I) intervjuades behandlare (n 11) och mammor (n 13) från tre kommuner och Sveriges största skyddade boende om sina erfarenheter av att ge och ta emot föräldraintervention. Intervjuerna analyserades separat med hjälp av tematisk analys, vilka sedan sammanslogs i en syntes. Resultaten grupperades i sex teman, vilket indikerade initiala tvivel på interventionene och på deltagarnas egen förmåga. Teman visade också att arbetet med PS ledde till en upplevd positiv förändring av barns beteende, liksom betonades behovet av kompetenta behandlare och anpassning utifrån kulturella skillnader. Den andra studien (studie II) utvärderade interventionens effekt avseende föräldrarnas föräldrakapacitet (n 35) och deras barns psykiska symtom (19 pojkar och 16 flickor, medelålder 6 år). Resultaten visade att effekten som erhållits av PS i Sverige kan i nästan samma omfattning likställas med den effekt som framgår av tidigare studier gjorda i USA. Även barnens känslomässiga symptom, problem och hyperaktivitet minskade, och föräldrarna bemötte sina barn på ett mer positivt sätt. Föräldrarna upplevde också minskad hjälplöshet och rädsla i relationen till sina barn. Det var också en kovarians mellan föräldrarnas känslor (hjälplöshet och rädsla) och barns uppförandeproblem och hyperaktivitet där båda minskade.Slutsatserna i denna licentiatuppsats är för det första att det är möjligt att genomföra PS med föräldrar utsatta för våld i nära relationer och deras barn som har utvecklat känslomässiga och beteendemässiga problem och för det andra att organisatoriska förhållanden behöver anpassas för att interventionen ska kunna implementeras i svensk socialtjänst.
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| 13. |
- Lindfors, Lina
(författare)
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Molecular Imaging of Diabetic Kidney Tissue and Binding Studies of Proinsulin C-peptide
- 2020
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Diabetic kidney disease is a serious complication of diabetes with a complex and incompletely understood pathology. In this work, the molecular changes in diabetic rat kidneys at a very early disease stage were studied using nanospray desorption electrospray ionisation mass spectrometry imaging. Our results demonstrate how disease-relevant metabolites and lipids can be conveniently analysed on intact kidney tissue sections. A number of significantly increased metabolites were identified in the diabetic kidney, revealing disturbances in energy metabolism detectable before histological changes.Proinsulin C-peptide is produced in the pancreas along with insulin and has shown beneficial effects in diabetes, but its mode of action is not yet known. 125I radiolabelled C-peptide was used to study its tissue distribution in healthy and diabetic rats after intravenous injection. The majority of C-peptide accumulated in renal tissues, with lower levels in the diabetic animals, showing that there are significant changes in kidney – C-peptide interactions in early stage diabetes.The interactions of C-peptide with the orphan receptor GPR146, which has been proposed as its receptor, were also investigated using Chinese hamster ovary cells overexpressing human GPR146. Neither dynamic mass redistribution nor β-arrestin recruitment assays showed any significant response to human or murine C-peptides in the GPR146 overexpressing cells compared to controls. Fluorescence confocal microscopy revealed no surface binding or cellular uptake of C-peptides by GPR146 overexpressing cells compared to controls. These combined results refute the suggestion that GPR146 is the C-peptide receptor.To further probe the function of C-peptide, 15N-labelled residues were incorporated into the peptide in preparation for nanoscale secondary ion mass spectrometry imaging of cells and intact kidney tissue sections. A number of crosslinking C-peptides were also designed and synthesised for experiments aimed at identifying its binding target. These studies have not yet been completed. Finally, to investigate the structure-activity relationship of C-peptide, a library of modified pentapeptide analogues was created for medium-throughput testing in a cell assay.
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| 14. |
- Lindquist, Charlotta, 1968-
(författare)
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Design & Synthesis of Protein Interacting Affinity Ligands and Protease Inhibitors
- 2010
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- There is a growing need of both protein drugs and synthetic drugs in the fight against many life-threatening diseases. The first part of this thesis deals with the design and synthesis of high affinity binding ligands for the purification of protein drugs. The second part describes design and synthesis of protease inhibitors targeting the cysteine protease cathepsin S and the serine protease hepatitis C NS3/4A. In work with parallel synthesis of new affinity chromatography ligands, indole was used as the scaffold for both solid phase and solution phase syntheses. A library of 1,3-disubstituted indoles was prepared via an iterative Mannich reaction sequence. The first Mannich reaction provided 3-aminomethylindoles, while the second Mannich reaction introduced an additional aminomethyl group at the N1 position of the indole ring. A library of 25 substituted indoles was prepared in moderate to good yields and purity. Inhibition of the cysteine protease cathepsin S is an attractive target for drug development of inhibitors having potential for regulation of autoimmune diseases and allergic disorders. Syntheses targeting the cysteine protease cathepsin S were performed by a solid phase approach. The structure-activity-relationships (SAR) of variations in the P3 sulfonamide part of 4-amidofuran-3-one inhibitors are presented. Several highly potent inhibitors were found, in both enzyme and cellular assays. The hepatitis C virus (HCV), causes a chronic liver condition which can lead to cirrhosis and liver cancer. The serine protease hepatitis C NS3/4A is a promising target for development of HCV drugs. In the syntheses of novel HCV NS3/4A inhibitors, four new P2 substituents were first incorporated on a proline-based linear scaffold. The most potent P2 substituent, quinazoline, was evaluated in a larger study yielding more rigidified cyclopentane-based macrocyclic inhibitors. The SAR exercise resulted in several inhibitors with excellent potency in the low nanomolar range.
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| 15. |
- Lindström, Anton, 1976-
(författare)
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A multivariate approach to characterization of drug-like molecules, proteins and the interactions between them
- 2008
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- En sjukdom kan många gånger härledas till en kaskadereaktion mellan proteiner, co-faktorer och substrat. Denna kaskadreaktion blir många gånger målet för att behandla sjukdomen med läkemedel. För att designa nya läkemedelsmoleyler används vanligen datorbaserade verktyg. Denna design av läkemedelsmolekyler drar stor nytta av att målproteinet är känt och då framförallt dess tredimensionella (3D) struktur. Är 3D-strukturen känd kan man utföra så kallad struktur- och datorbaserad molekyldesign, 3D-geometrin (f.f.a. för inbindningsplatsen) blir en vägledning för designen av en ny molekyl. Många faktorer avgör interaktionen mellan en molekyl och bindningsplatsen, till exempel fysikalisk-kemiska egenskaper hos molekylen och bindningsplatsen, flexibiliteten i molekylen och målproteinet, och det omgivande lösningsmedlet. För att strukturbaserad molekyldesign ska fungera väl måste två viktiga steg utföras: i) 3D anpassning av molekyler till bindningsplatsen i ett målprotein (s.k. dockning) och ii) prediktion av molekylers affinitet för bindningsplatsen. Huvudsyftena med arbetet i denna avhandling var som följer: i) skapa modeler för att prediktera affiniteten mellan en molekyl och bindningsplatsen i ett målprotein; ii) förfina molekyl-protein-geometrin som skapas vid 3D-anpassning mellan en molekyl och bindningsplatsen i ett målprotein (s.k. dockning); iii) karaktärisera proteiner och framför allt deras sekundärstruktur; iv) bedöma effekten av olika matematiska beskrivningar av lösningsmedlet för förfining av 3D molekyl-protein-geometrin skapad vid dockning och prediktion av molekylers affinitet för proteiners bindningsfickor. Ett övergripande syfte var att använda kemometriska metoder för modellering och dataanalys på de ovan nämnda punkterna. För att sammanfatta så presenterar denna avhandling metoder och resultat som är användbara för strukturbaserad molekyldesign. De rapporterade resultaten visar att det är möjligt att skapa kemometriska modeler för prediktion av molekylers affinitet för bindningsplatsen i ett protein och att dessa presterade lika bra som andra vanliga metoder. Dessutom kunde kemometriska modeller skapas för att beskriva effekten av hur inställningarna för olika parametrar i dockningsprogram påverkade den 3D molekyl-protein-geometrin som dockingsprogram skapade. Vidare kunde kemometriska modeller andvändas för att öka förståelsen för deskriptorer som beskrev sekundärstrukturen i proteiner. Förfining av molekyl-protein-geometrin skapad genom dockning gav liknande och ickesignifikanta resultat oberoende av vilken matematisk modell för lösningsmedlet som användes, förutom för ett fåtal (sex av 30) fall. Däremot visade det sig att användandet av en förfinad geometri var värdefullt för prediktion av molekylers affinitet för bindningsplatsen i ett protein. Förbättringen av prediktion av affintitet var markant då en Poisson-Boltzmann beskrivning av lösningsmedlet användes; jämfört med prediktionerna gjorda med ett dockningsprogram förbättrades korrelationen mellan beräknad affintiet och uppmätt affinitet med 0,7 (R2).
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| 16. |
- Orrling, Kristina M., 1976-
(författare)
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On the Versatility of Microwave-Assisted Chemistry : Exemplified by Applications in Medicinal Chemistry, Heterocyclic Chemistry and Biochemistry
- 2009
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Today, the demand for speed in drug discovery is constantly increasing, particularly in the iterative processes of hit validation and expansion and lead optimization. Irradiation with microwaves (MWs) has been applied in the area of organic synthesis to accelerate chemical reactions and to facilitate the generation of new chemical entities since 1986. In the work presented in this thesis, the use of MW-mediated heating has been expanded to address three fields of drug discovery, namely hit expansion, chemical library generation and genomics. In the first project, potential inhibitors of malaria aspartic proteases were designed and synthesized, partly by MW-assisted organic chemistry, and evaluated with regard to their inhibitory efficacy on five malaria aspartic proteases and their selectivity over two human aspartic proteases. The synthetic work included the development of fast and convenient methods of MW-assisted formation of thiazolidines and epoxy esters. Some of the resulting structures proved to be efficacious inhibitors of the aspartic protease that degrades haemoglobin in all four malaria parasites infecting man. No inhibitor affected the human aspartic proteases. Expedient, two-step, single-operation synthetic routes to heterocycles of medicinal interest were developed in the second and third projects. In the former, the use of a versatile synthon, Ph3PCCO, provided α,β-unsaturated lactones, lactams and amides within 5–10 minutes. In the latter project, saturated lactams were formed from amines and lactones in 35 minutes, in the absence of strong additives. These two MW-mediated protocols allowed the reduction of the reaction time from several hours or days to minutes. In the fourth project, a fully automated MW-assisted protocol for the important enzyme-catalysed polymerase chain reaction (PCR) was established. In addition, the PCR reaction could be performed in unusually large volumes, 2.5 mL and 15 mL, with yields corresponding to those from conventional PCR. Good amplification rates suggested that the thermophilic enzyme, Taq polymerase, was not affected by the MW radiation.
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| 17. |
- Pemberton, Nils, 1978-
(författare)
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Synthesis and functionalization of ring-fused 2-pyridones : Targeting pili formation in E. coli
- 2007
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Bicyclic dihydrothiazolo fused 2-pyridones have been studied as a new class of antibacterial agents, termed pilicides, that target the formation of adhesive bacterial surface organelles (pili) in pathogenic bacteria. Synthetic methods to further functionalize the bicyclic 2-pyridone scaffold have been developed in order to increase water-solubility and thereby facilitate biological evalua-tions. This was accomplished by introducing aminomethylenes at the open position C-6. Tertiary amines were introduced via a microwave–assisted Mannich reaction and a synthetic route based on a formyl intermediate gave access to primary, secondary and tertiary amines, but also to other interesting functionalities. Biological evaluation confirmed that several of the function-alized compounds inhibited pili formation in uropathogenic E. coli., as dem-onstrated by assays of hemagglutination, biofilm formation and adherence to bladder cells. Co-crystallizing one of the pilicides with the target protein gave information about the binding site and based on this a mechanism of action was proposed, which was supported experimentally by surface plas-mon resonance and single point mutations in the protein. Furthermore, the previously developed acylketene imine reaction used to prepare bicyclic thiazolo fused 2-pyridone pilicides has been developed to allow preparation of other ring-fused 2-pyridone systems. Benzo[a]quinolizine-4-ones and indolo[2,3-a]quinolizine-4-ones could be prepared in a fast and simple manner starting from dihydroisoquinolines and a β-carboline. Finally, this method could also be applied for the preparation of heteroatom analogs of the previously studied sulfur containing pilicides. Biological evaluations established that the sulfur atom can be replaced by oxygen and still maintain the ability to prevent pili assembly.
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| 18. |
- Pudelko, Maciej, 1978-
(författare)
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Antigens derived from the mucin MUC1 : Solution and solid-phase synthesis of saccharides, peptides and glycopeptides
- 2008
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Mucin is a term used to describe a large family of heavily glycosylated proteins which are present on the surfaces of secretory epithelial cells and are overexpressed by many carcinomas. Membrane-bound mucin MUC1 is of special interest. Its backbone consists of repeating units of twenty amino acids with five potential glycosylation sites. These sites are expanded to structures like the T (Galβ(1->3)GalNAcα-Ser/Thr) and Tn (GalNAcα-Ser/Thr) antigens by the action of various glycosyltransferases. In different types of carcinomas these epitopes are being terminated by sialic acid residues to form among others: 2,3-sialyl-T and sialyl-Tn structures due to the elevated levels of different sialyltransferases. Solid-phase synthesis of the selected antigens derived from the mucin MUC1 has been developed and optimized. A chemoenzymatic approach has been used to effectively prepare 2,3-sialyl-T and 2,6-sialyl-Tn glycopeptides. The formation of intramolecular sialic acid lactones in presence of acetic acid was investigated. The stability of lactones formed from 2,3-sialyl-T towards water was studied using NMR spectroscopy and it appeared that 1''->2' lactone displayed remarkable strength to hydrolysis and it was suggested as a candidate for cancer vaccine. Gel-phase 19F NMR spectroscopy is known to be a very good tool to characterize resin-bound products using fluorinated protecting groups and linker molecules. The hydrophobic peptide LLLLTVLTV, which is a fragment from the MUC1 signal sequence, was prepared using solid-phase synthesis according to a modified Fmoc protocol with more active coupling reagent, stronger base, and the isopropylidene dipeptide Fmoc-Leu-Thr-(ΨMe,Mepro)-OH. Gel-phase 19F NMR spectroscopy was used to evaluate peptide chain aggregation and coupling and deprotection efficiency. A carbamate linker strategy proved to be effective in solid-phase synthesis of serine-based neoglycolipids with terminal amino functionality. Neoglycolipids were covalently bound to secondary amines in microtiter plates using squaric acid ester methodology. These arrays have potential to study the interactions between carbohydrates and e.g. proteins and microbes. The new fluorinated α-amino protective group [1-(4-(4-fluorophenyl)-2,6-dioxocyclohexylidene)ethyl] Fde was developed. This group is cleaved with hydrazine in DMF solution. By using amino acids protected with this group, it was possible to quantify the efficiency of peptide coupling using gel-phase 19F NMR spectroscopy.
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| 19. |
- Sellstedt, Magnus, 1981-
(författare)
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Development of 2-Pyridone-based central fragments : Affecting the aggregation of amyloid proteins
- 2012
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- There are many applications of small organic compounds, e.g. as drugs or as tools to study biological systems. Once a compound with interesting biological activity has been found, medicinal chemists typically synthesize small libraries of compounds with systematic differences to the initial “hit” compound. By screening the new ensemble of compounds for their ability to perturb the biological system, insights about the system can be gained. In the work presented here, various ways to synthesize small libraries of ring-fused 2‑pyridones have been developed. Members of this class of peptidomimetic compounds have previously been found to have a variety of biological activities, e.g. as antibacterial agents targeting virulence, and as inhibitors of the aggregation of Alzheimer b‑peptides. The focus in this work has been to alter the core skeleton, the central fragment, of the previously discovered biologically active 2‑pyridones and evaluate the biological effects of these changes. Several new classes of compounds have been constructed and their preparations have included the development of multi-component reactions and a method inspired by diversity-oriented synthesis. Some of the new compounds have been evaluated for their effect on the fibrillation of different amyloid proteins. Both the Parkinson-associated amyloid protein a-synuclein and the bacterial protein CsgA that is involved in bacterial biofilm formation are affected by subtle changes of the compounds’ central fragments. This is an example of the usefulness of central-fragment alterations as a strategy to probe structure-activity relationships, and the derived compounds may be used as tools in further study of the aggregation of amyloid proteins.
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| 20. |
- Draxler, Helena, 1977-
(författare)
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Initial transfer of Project Support from the US to Sweden : Feasibility and clinical aspects as viewed by counsellors and parents
- 2022
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Project Support (PS) is a parenting programme developed and studied in the US for parents and children exposed to intimate partner violence (IPV) and for these children with subsequent behavioural problems. By examining feasibility aspects and using qualitative and quantitative methods, the study aimed to explore the transferability of PS to a new naturalistic context, i.e., Swedish social services. In the qualitative Study I, IPV-exposed mothers and counsellors who received/conducted PS were interviewed about their experiences. Study II explored whether the effects of PS in the US, i.e., reduced children’s psychological symptoms and improved parenting capacity among parents and children exposed to IPV, could be replicated in Sweden. Study III explored group and individual impacts of PS on parents regarding aspects of IPV, global psychiatric symptoms, and post-traumatic stress symptoms. Study IV is a qualitative follow-up study where counsellors were interviewed about how the PS work was or was not continued. The results indicate cultural differences between the US and Sweden in practising parenting skills though role-playing and in how to address and understand child misbehaviour, necessitating adaptation to Swedish values and norms. Results further indicate that PS is a promising intervention for Swedish IPV-exposed parents and their children with behavioural problems, leading to improved parental capacity and decreased child behavioural problems, supporting prior research in an American context. PS generally seems to improve parents’ psychiatric symptoms, but examining each individual revealed that most parents still had dysfunctional levels of symptoms post treatment. Maintenance and use of PS was inconsistent, due to lack of organisational support. The study indicates that PS can be further implemented and evaluated in Sweden, and stresses the importance of conducting feasibility studies when an intervention is to be transferred to a new context.
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| 21. |
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| 22. |
- Larsson, Andreas, 1972-
(författare)
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Antiadhesive agents targeting uropathogenic Escherichia coli : Multivariate studies of protein-protein and protein-carbohydrate interactions
- 2004
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- This thesis describes studies directed towards development of novel antiadhesive agents, with particular emphasis on compounds that prevent attachment of bacteria to a host-cell. Three different proteins involved in the assembly or function of adhesive pili in uropathogenic Escherichia coli have been targeted either by rational structure based design or statistical molecular methods. A library of substituted galabiose (Galα1-4Gal) derivatives was screened for binding to the E. coli adhesin PapG in an assay based on surface plasmon resonance, and for inhibition of Streptococcus suis adhesins PN and PO in a hemagglutination assay. The results were used to generate QSAR models which had good predictive powers and provided further insight in the structural requirements needed for high affinity binding. 2-pyridones and amino acid derivatives were modelled into the binding site of chaperones involved in pilus assembly in E. coli and a heuristic method, VALIDATE, was used for affinity prediction. The affinity of the compounds for the chaperones PapD and FimC were assessed in assays based on surface plasmon resonance and relaxation-edited NMR spectroscopy. Their ability to disrupt chaperone/subunit complexes was investigated in vitro through a FPLC assay and their capacity to inhibit pilus formation in vivo was determined via hemagglutination and confirmed with atomic force microscopy. Statistical molecular design was used to design a diverse peptide library targeting pili subunits, and an ELISA was developed to investigate the ability of the peptides to inhibit chaperone/subunit complexation. The resulting QSAR model provided extensive information regarding binding of the peptides to the subunits. Because the peptides were suggested to bind in an extended β-strand formation, β-strand mimetics consisting of oligomeric enaminones were designed. Finally, new methods to synthesize enaminone building blocks were developed using microwave assisted chemistry. The projects described have generated compounds that besides their value as leads for developing novel antibacterial agents, also constitute new chemical tools to study the mechanisms underlying bacterial virulence.
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| 23. |
- Thurfjell, Barbro, 1951-
(författare)
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Adolescent Eating Disorders in a Sociocultural Context
- 2005
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Adolescence means an increased risk for eating disorders (ED) and the female gender is the most important risk factor. Empirical studies of the perceptions of gender ideals—as potential mediating factors between the socio-cultural context and ED—were this thesis’ primary goals. Further aims implied evaluating the psychometric properties of the Eating Disorders Inventory for Children (EDI-C) and providing normative data for boys and girls concerning self-assessed ED symptoms and related psychological traits.The EDI-C has satisfying psychometric properties, and normative data are now provided. The vast majority of preadolescent and adolescent boys and girls (n=4725) are not preoccupied with weight concerns, but 1–3% of the boys and 9–20% of the girls here indicate feelings of anxiety and problematic attitudes. An increase of self-reported ED symptoms occurs during female preadolescence and early adolescence. Responses to three open-ended questions—addressing perceived and personal gender ideals of adolescent boys (n=347) and girls with (n=204) and without (n=417) an ED—indicate that girls are, compared to boys, more involved in images of ideals. The perceived ideal man and ideal woman differ in body size and strength. This corresponds with different health risk behaviours reported in boys (body building) and girls (dieting). Awareness of media ideals and an ability to discriminate them from individual values was common in the normal groups and in girls who had recovered from an ED. A poor outcome was associated with thinness stated as ideals of the media, without mentioning good looks. Factors that could predict outcome at the 3-year follow-up of an ED in adolescent girls were: good looks (media ideal), maturity fears, profession (individual ideal), sexual debut, and vomiting.The addition of a gender perspective would add a dimension to programmes aiming at preventing and treating ED in adolescence.
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