Selective Uptake Into Inflamed Human Intestinal Tissue and Immune Cell Targeting by Wormlike Polymer Micelles.

Inflammatory bowel disease (IBD) has become a globally prevalent chronic disease with no causal therapeutic options. Targeted drug delivery systems with selectivity for inflamed areas in the gastrointestinal tract promise to reduce severe drug-related side effects. By creating three distinct nanostructures (vesicles, spherical, and wormlike micelles) from the same amphiphilic block copolymer poly(butyl acrylate)-block-poly(ethylene oxide) (PBA-b-PEO), the effect of nanoparticle shape on human mucosal penetration is systematically identified. An Ussing chamber technique is established to perform the ex vivo experiments on human colonic biopsies, demonstrating that the shape of polymeric nanostructures represents a rarely addressed key to tissue selectivity required for efficient IBD treatment. Wormlike micelles specifically enter inflamed mucosa from patients with IBD, but no significant uptake is observed in healthy tissue. Spheres (≈25 nm) and vesicles (≈120 nm) enter either both normal and inflamed tissue types or do not penetrate any tissue. According to quantitative image analysis, the wormlike nanoparticles localize mainly within immune cells, facilitating specific targeting, which is crucial for further increasing the efficacy of IBD treatment. These findings therefore demonstrate the untapped potential of wormlike nanoparticles not only to selectively target the inflamed human mucosa, but also to target key pro-inflammatory cells.

Switchable Electrostatically Templated Polymerization.

We report a switchable, templated polymerization system where the strength of the templating effect can be modulated by solution pH and/or ionic strength. The responsiveness to these cues is incorporated through a dendritic polyamidoamine-based template of which the charge density depends on pH. The dendrimers act as a template for the polymerization of an oppositely charged monomer, namely sodium styrene sulfonate. We show that the rate of polymerization and maximum achievable monomer conversion are directly related to the charge density of the template, and hence the environmental pH. The polymerization could effectively be switched "ON" and "OFF" on demand, by cycling between acidic and alkaline reaction environments. These findings break ground for a novel concept, namely harnessing co-assembly of a template and growing polymer chains with tunable association strength to create and control coupled polymerization and self-assembly pathways of (charged) macromolecular building blocks.

Tuneable Time Delay in the Burst Release from Oxidation-Sensitive Polymersomes Made by PISA.

Reactive polymersomes represent a versatile artificial cargo carrier system that can facilitate an immediate release in response to a specific stimulus. The herein presented oxidation-sensitive polymersomes feature a time-delayed release mechanism in an oxidative environment, which can be precisely adjusted by either tuning the membrane thickness or partial pre-oxidation. These polymeric vesicles are conveniently prepared by PISA allowing the straightforward and effective in situ encapsulation of cargo molecules, as shown for dyes and enzymes. Kinetic studies revealed a critical degree of oxidation causing the destabilization of the membrane, while no release of the cargo is observed beforehand. The encapsulation of glucose oxidase directly transforms these polymersomes into glucose-sensitive vesicles, as small molecules including sugars can passively penetrate their membrane. Considering the ease of preparation, these polymersomes represent a versatile platform for the confinement and burst release of cargo molecules after a precisely adjustable time span in the presence of specific triggers, such as H2 O2 or glucose.

Influence of Core Cross-Linking and Shell Composition of Polymeric Micelles on Immune Response and Their Interaction with Human Monocytes.

Block copolymer micelles have received increasing attention in the last decades, in particular for their appealing properties in nanomedicine. However, systematic investigations of the interaction between polymeric micelles and immune cells are still rare. Therefore, broader studies comparing the structural effects remain inevitable for a comprehensive understanding of the immune response and for the design of efficient, nonimmunogenic delivery systems. Here, we present novel block copolymer micelles with the same hydrophobic core, based on a copolymer of BA and VDM, and various hydrophilic shells ranging from common PEG derivatives to morpholine-based materials. The influence of these shells on innate immune responses was studied in detail. In addition, we investigated the impact of micelle stability by varying the cross-linking density in the micellar core. Surprisingly, whereas different shells had only a minor impact on immune response, micelles with reduced cross-linking density considerably enhanced the release of cytokines from isolated human monocytes. Moreover, the uptake of non-cross-linked micelles by monocytes was significantly higher as compared to cross-linked materials. Our study emphasizes the importance of the micellar stability on the interaction with the immune system, which is the key for any stealth properties in vivo. Polymers based on morpholines result in a similar low response as the PEG derivative and may represent an interesting alternative to the common PEGylation.

Predictive Strength of Photophysical Measurements for in Vitro Photobiological Activity in a Series of Ru(II) Polypyridyl Complexes Derived from π-Extended Ligands.

This study investigates the correlation between photocytotoxicity and the prolonged excited-state lifetimes exhibited by certain Ru(II) polypyridyl photosensitizers comprised of π-expansive ligands. The eight metal complexes selected for this study differ markedly in their triplet state configurations and lifetimes. Human melanoma SKMEL28 and human leukemia HL60 cells were used as in vitro models to test photocytotoxicity induced by the compounds when activated by either broadband visible or monochromatic red light. The photocytotoxicities of the metal complexes investigated varied over 2 orders of magnitude and were positively correlated with their excited-state lifetimes. The complexes with the longest excited-state lifetimes, contributed by low-lying 3IL states, were the most phototoxic toward cancer cells under all conditions.

How To Tune the Gene Delivery and Biocompatibility of Poly(2-(4-aminobutyl)-2-oxazoline) by Self- and Coassembly.

Despite their promising potential in gene transfection, the toxicity and limited efficiency of cationic polymers as nonviral vectors are major obstacles for their broader application. The large amount of cationic charges, for example, in poly(ethylene imine) (PEI) is known to be advantageous in terms of their transfection efficiency but goes hand-in-hand with a high toxicity. Consequently, an efficient shielding of the charges is required to minimize toxic effects. In this study, we use a simple mixed-micelle approach to optimize the required charge density for efficient DNA complex formation and to minimize toxicity by using a biocompatible polymer. In detail, we coassembled mixed poly(2-oxazoline) nanostructures ( d ≈ 100 nm) consisting of a hydrophobic-cationic block copolymer (P(NonOx52- b-AmOx184)) and a hydrophobic-hydrophilic stealth block copolymer (P(EtOx155- b-NonOx76) in ratios of 0, 20, 40, 60, 80, and 100 wt % P(NonOx52- b-AmOx184). All micelles with cationic polymers exhibited a very good DNA binding efficiency and dissociation ability, while the bio- and hemocompatibility improved with increasing EtOx content. Analytics via confocal laser scanning microscopy and flow cytometry showed an enhanced cellular uptake, transfection ability, and biocompatibility of all prepared micelleplexes compared to AmOx homopolymers. Micelleplexes with 80 or 100 wt % revealed a similar transfection efficiency as PEI, while the cell viability was significantly higher (80 to 90% compared to 60% for PEI).

Anti-Inflammatory Potential of Beclometasone-Loaded Filomicelles on Activated Human Monocytes.

Polymeric micelles with a hydrophobic core represent versatile nanostructures for encapsulation and delivery of water-insoluble drugs. Here, water-insoluble beclometasone dipropionate (BDP) which is a potent anti-inflammatory therapeutic agent but limited to topical applications so far, is encapsulated. Therefore, this work used an amphiphilic block copolymer self-assembling into flexible polymeric filomicelles, which have recently proven to selectively target inflamed areas in patients with inflammatory bowel disease (IBD). The small diameter and flexibility of these filomicelles is considered beneficial for transepithelial passages, while their length minimizes the unspecific uptake into nontargeted cells. This work successfully establishes a protocol to load the water-insoluble BDP into the core of the filomicelles, while maintaining the particle stability to prevent any premature drug release. The anti-inflammatory efficacy of BDP-loaded filomicelles is further investigated on lipopolysaccharide (LPS) stimulated human monocytes. In these ex vivo assays, the BDP-loaded filomicelles significantly reduce TNF-α, IL-6, IL-1ß, IL-12p70, IL-17a, and IL-23 release after 24 h. Additional time course study of drug-loaded filomicelles and their comparison with a common water-soluble and unspecific corticosteroid demonstrate promising results with significant immune response suppression in stimulated monocytes after 2 and 6 h. These findings demonstrate the potential of polymeric filomicelles as a vehicle for potent water-insoluble corticosteroids.

Efficient Transfection via an Unexpected Mechanism by Near Neutral Polypiperazines with Tailored Response to Endosomal pH.

Cationic pH-responsive polymers promise to overcome critical challenges in cellular delivery. Ideally, the polymers become selectively charged along the endosomal pathway disturbing only the local membrane and avoiding unintended interactions or cytotoxic side effects at physiological conditions. Polypiperazines represent a novel, hydrophilic class of pH-responsive polymers whose response can be tuned within the relevant pH range (5-7.4). The authors discovered that the polypiperazines are effectively binding plasmid DNA (pDNA) and demonstrate high efficiency in transfection. By design of experiments (DoE), a wide parameter space (pDNA and polymer concentration) is screened to identify the range of effective concentrations for transfection. An isopropyl modified polypiperazine is highly efficient over a wide range of concentrations outperforming linear polyethylenimine (l-PEI, 25 kDa) in regions of low N*/P ratios. A quantitative polymerase chain reaction (qPCR) surprisingly revealed that the pDNA within the piperazine-based polyplexes can be amplified in contrast to polyplexes based on l-PEI. The pDNA must therefore be more accessible and bound differently than for other known transfection polymers. Considering the various opportunities to further optimize their structure, polypiperazines represent a promising platform for designing effective soluble polymeric vectors, which are charge-neutral at physiological conditions.

ROS-Sensitive Polymer Micelles for Selective Degradation in Primary Human Monocytes from Patients with Active IBD.

Inflammatory bowel disease (IBD) is characterized by increased levels of reactive oxygen species (ROS) in inflamed areas of the gastrointestinal tract and in circulating immune cells, providing novel opportunities for targeted drug delivery. In the recent experiments, oxidation-responsive polymeric nanostructures selectively degrade in the presence of H2 O2 . Based on these results, it is hypothesized that such degradation process can be triggered in a similar way by the incubation with stimulated monocytes isolated from patients with IBD. A first indication is given by a significant correlation between excessive ROS and degradation of micelles in monocytes isolated from healthy individuals after phorbol 12-myristate 13-acetate (PMA) stimulation. But even if the ROS-sensitive micelles are incubated with nonstimulated monocytes from patients with active IBD, a spontaneous degradation is observed in contrast to micelles incubated with monocytes from healthy donors. The findings indicate that the thioether-based micelles are indeed promising for selective drug release in the presence of activated immune cells.

Correlation between Protonation of Tailor-Made Polypiperazines and Endosomal Escape for Cytosolic Protein Delivery.

Responsive polymers, which become protonated at decreasing pH, are considered a milestone in the development of synthetic cell entry vectors. Exact correlations between their properties and their ability to escape the endosome, however, often remain elusive due to hydrophobic interactions or limitations in the design of water-soluble materials with suitable basicity. Here, we present a series of well-defined, hydrophilic polypiperazines, where systematic variation of the amino moiety facilitates an unprecedented fine-tuning of the basicity or pKa value within the physiologically relevant range (pH 6-7.4). Coincubation of HEK 293T cells with various probes, including small fluorophores or functioning proteins, revealed a rapid increase of endosomal release for polymers with pKa values above 6.5 or 7 in serum-free or serum-containing media, respectively. Similarly, cytotoxic effects became severe at increased pKa values (>7). Although the window for effective transport appears narrow, the discovered correlations offer a principal guideline for the design of effective polymers for endosomal escape.

One polymer composition, various morphologies: the decisive influence of conditions on the polymerization-induced self-assembly (PISA) of N-acryloyl thiomorpholine.

Polymerization-induced self-assembly (PISA) represents a powerful technique for the preparation of nanostructures comprising various morphologies. Herein, we demonstrate that the recently introduced monomer N-acryloylthiomorpholine (NAT) features a unique self-assembly behaviour during an aqueous PISA. The one-pot, aqueous RAFT dispersion polymerization starting from short poly(N-acryloylmorpholine) (PNAM) enables access to all common solution morphologies including spheres, worms, vesicles and lamellae, at very low molar masses (< 8 kDa). Moreover, all these structures can be obtained for the same polymer composition and size by the variation of the polymerization temperature and concentration of the monomer. This exceptional self-assembly behavior is associated with the combination of a high glass transition temperature, excellent water solubility of the monomer, and the early onset of aggregation during the polymerization, which stabilizes the morphology at different stages. This PISA system opens up new opportunities to reproducibly create versatile, functional nanostructures and enables an independent evaluation of morphology-property relationships, as it is exemplarily shown for the oxidative degradation of spherical and wormlike micelles, as well as vesicles.