pH-cleavable and hyperbranched polyether architectures: from novel synthesis strategies to applications in nanotechnology and biomedicine
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Abstract
The use of polyethylene glycol (PEG) is diverse, ranging from food technology and cosmetics to applications in nanotechnology and pharmaceutical industries. However, despite the various favorable properties of PEG, its non-degradability and the low number of functional groups can be a drawback to many applications. The aim of this thesis is to establish cleavability of PEG and PEG-derived lipids and surfactants by exploring novel synthetic pathways to incorporate pH-hydrolyzable moieties into the polyether structures. A strong emphasis is put on elucidating the hydrolysis kinetics of the novel materials to evaluate their potential for future applications. Furthermore, hyperbranched polyglycerol (hbPG) architectures are investigated that contain a large number of hydroxyl functionalities, which render hbPGs interesting PEG substitutes if multifunctionality of the polyether is a requirement.
Chapter 1.1 serves as an introduction to the chemistry of PEG and presents an overview on various routes to incorporate functional moieties into the backbone or at the chain ends of PEG. A particular focus is directed at the anionic ring-opening polymerization (AROP) of epoxide monomers, which constitutes the fundamental polymerization technique employed in this thesis. Chapter 1.2 represents an introduction into liposome research, which highlights the development of sterically stabilized liposomes (“stealth liposomes”), active cell targeting and stimuli-responsive drug release strategies. Especially, the usefulness of pH-cleavable PEG-lipids in liposomal formulations is illustrated. In Chapter 1.3 block copolymers composed of poly(propylene oxide) (PPO) and PEG (denoted as PEO) are introduced, focusing on the broad range of applications for PEO-b-PPO-b-PEO copolymers (poloxamers) as surfactants and reflecting recent trends in this area. An overview on cleavable PEG architectures is given in Chapter 1.4 summarizing current achievements in addressing the non-degradability of PEG. Problems arising from PEG’s biopersistence are discussed that, for instance, set an upper molecular weight limit of PEG for the use in biomedical applications. Chapters 1.3 and 1.4 are sections of a comprehensive review article that was published in Chemical Reviews.
In Chapter 2, the design of pH-cleavable poloxamer analogs (PEO-b-PPO-b-PEO copolymers) is presented that contain acid-sensitive acetal linkages at the block junctions. In a proof-of-concept experiment, the novel amphiphilic polymers were used as surfactants for the miniemulsion polymerization of styrene to prepare well-defined polystyrene nanoparticles. pH-triggered precipitation of these nanoparticles from stable miniemulsions allowed efficient removal of surfactant fragments to obtain additive-free nanoparticles. Separation of surfactants is a major challenge with MEPs, however, is desirable due to environmental concerns and potential surfactant-mediated changes of particle properties.
To address the non-degradability of PEG, in Chapter 3 the anionic ring-opening copolymerization of ethylene oxide and 3,4-epoxy-1-butene (EPB) was explored to introduce multiple pH-sensitive vinyl ether moieties in the PEG backbone after catalytic isomerization of the allylic double bonds. Well-defined, acid-cleavable copolymers of tailorable molecular weight with low contents of EPB (4 mol%) were synthesized in an efficient two-step procedure. Analysis of degradation products via SEC indicated moderate molecular weight distribution (Ð = 1.6 – 1.8). Hydrolysis profiles of pH-cleavable P(EPB-co-EG) copolymers were monitored by 1H NMR spectroscopy in a physiologically relevant pH range of 4.4 to 5.4, revealing promising cleavage kinetics with respect to biomedical application, e.g. for cleavable, long-circulating polymer-drug conjugates.
The synthesis of pH-sensitive PEG-lipids with in-chain cleavable ketal and acetal units for the application in pH-responsive “stealth” liposomes is reported in Chapter 4. To this end, a novel synthesis strategy towards asymmetric ketals in polymers was developed providing access to PEG-lipids of tunable molecular weights and with a defined hydroxyl end group facilitating post-modification reactions. In situ 1H NMR kinetic studies on the hydrolysis rates of the pure lipids suggested significantly faster cleavage of ketals compared to acetals. The behavior of the lipids in liposomal formulations was further analyzed via time-resolved fluorescence spectroscopy and gel electrophoresis assays to monitor pH-dependent detachment of PEG from the liposomal surface. Highly promising PEG shedding properties in mildly acidic media (pH 5.5 to 6.5) were found for the ketal-functional PEG-lipids that suggest great potential for pH-triggered drug release strategies from liposomes e.g. in tumor therapy.
In liposomal research, hbPGs are promising alternatives to PEG as they can convey a multivalence effect in active cell targeting. In order to study the effect of the polyether architecture on liposomal properties, in Chapter 5 dialkyl-functional hbPG- and PEG-lipids were synthesized with molecular weights of 3 to 7 kg mol-1. A novel linPG-lipid precursor offered access to the hbPG-lipids using slow monomer addition of the latent AB2-type monomer glycidol. Liposomes containing linear or hyperbranched lipids were prepared by dual centrifugation. Tuning of the diameter of liposomes in a range below 100 nm could be achieved, which has been proposed as an optimum for antitumor therapy. First in vivo biodistribution studies of 18F-radiolabeled lipids by positron emission tomography (PET) imaging were performed that suggest efficient renal clearance of the PEG-/hbPG-lipids. In addition, blood circulation of radiolabeled, polyether-modified liposomes for more than 1 h was observed, providing evidence for hbPG-mediated “stealth” properties of liposomes in vivo.