Amphiphilic Peptide Self-Assembly: Expansion to Hybrid Materials

The design of functional systems with sizes in the nanometer range is a key challenge in fields such as biomedicine, nanotechnology, and engineering. Some of the most promising materials nowadays consist of self-assembling peptides or peptide–polymer hybrid materials because of their versatility and the resulting properties that can be achieved with these structures. Self-assembly of pure amphiphilic peptides or in combination with block copolymers results in a large variety of nanostructures (micelles, nanoparticles (NPs), compartments, planar membranes) each with different characteristics and tunable properties. Here, we describe such novel peptide- or peptide–polymer-based supramolecular nanostructures and emphasize their functionality and various promising applications

Current Perspectives on Synthetic Compartments for Biomedical Applications

Nano- and micrometer-sized compartments composed of synthetic polymers are designed to mimic spatial and temporal divisions found in nature. Self-assembly of polymers into compartments such as polymersomes, giant unilamellar vesicles (GUVs), layer-by-layer (LbL) capsules, capsosomes, or polyion complex vesicles (PICsomes) allows for the separation of defined environments from the exterior. These compartments can be further engineered through the incorporation of (bio)molecules within the lumen or into the membrane, while the membrane can be decorated with functional moieties to produce catalytic compartments with defined structures and functions. Nanometer-sized compartments are used for imaging, theranostic, and therapeutic applications as a more mechanically stable alternative to liposomes, and through the encapsulation of catalytic molecules, i.e., enzymes, catalytic compartments can localize and act in vivo. On the micrometer scale, such biohybrid systems are used to encapsulate model proteins and form multicompartmentalized structures through the combination of multiple compartments, reaching closer to the creation of artificial organelles and cells. Significant progress in therapeutic applications and modeling strategies has been achieved through both the creation of polymers with tailored properties and functionalizations and novel techniques for their assembly

From spherical compartments to polymer films: exploiting vesicle fusion to generate solid supported thin polymer membranes

Solid supported polymer membranes as scaffold for the insertion of functional biomolecules provide the basis for mimicking natural membranes. They also provide the means for unraveling biomolecule-membrane interactions and engineering platforms for biosensing. Vesicle fusion is an established procedure to obtain solid supported lipid bilayers but the more robust polymer vesicles tend to resist fusion and planar membranes rarely form. Here, we build on vesicle fusion to develop a refined and efficient way to produce solid supported membranes based on poly(dimethylsiloxane)-poly(2-methyl-2-oxazoline) (PMOXA-b-PDMS-b-PMOXA) amphiphilic triblock copolymers. We first create thiol-bearing polymer vesicles (polymersomes) and anchor them on a gold substrate. An osmotic shock then provokes polymersome rupture and drives planar film formation. Prerequisite for a uniform amphiphilic planar membrane is the proper combination of immobilized polymersomes and osmotic shock conditions. Thus, we explored the impact of the hydrophobic PDMS block length of the polymersome on the formation and the characteristics of the resulting solid supported polymer assemblies by quarz crystal microbalance with dissipation monitoring (QCM-D), atomic force microscopy (AFM) and spectroscopic ellipsometry (SE). When the PDMS block is short enough, attached polymersomes restructure in response to osmotic shock, resulting in a uniform planar membrane. Our approach to rapidly form planar polymer membranes by vesicle fusion brings many advantages to the development of synthetic planar membranes for bio-sensing and biotechnological applications

Tailoring a Solvent-Assisted Method for Solid-Supported Hybrid Lipid-Polymer Membranes

Combining amphiphilic block copolymers and phospholipids opens new opportunities for the preparation of artificial membranes. The chemical versatility and mechanical robustness of polymers together with the fluidity and biocompatibility of lipids afford hybrid membranes with unique properties that are of great interest in the field of bioengineering. Owing to its straightforwardness, the solvent-assisted method (SA) is particularly attractive for obtaining solid-supported membranes. While the SA method was first developed for lipids and very recently extended to amphiphilic block copolymers, its potential to develop hybrid membranes has not yet been explored. Here, we tailor the SA method to prepare solid-supported polymer-lipid hybrid membranes by combining a small library of amphiphilic diblock copolymers poly(dimethyl siloxane)-poly(2-methyl-2-oxazoline) and poly(butylene oxide)- block -poly(glycidol) with phospholipids commonly found in cell membranes including 1,2-dihexadecanoyl- sn -glycero-3-phosphocholine, 1-palmitoyl-2-oleoyl- sn -glycero-3-phosphoethanolamine, sphingomyelin, and 1,2-dioleoyl- sn -glycero-3-phosphoethanolamine- N -(glutaryl). The optimization of the conditions under which the SA method was applied allowed for the formation of hybrid polymer-lipid solid-supported membranes. The real-time formation and morphology of these hybrid membranes were evaluated using a combination of quartz crystal microbalance and atomic force microscopy. Depending on the type of polymer-lipid combination, significant differences in membrane coverage, formation of domains, and quality of membranes were obtained. The use of the SA method for a rapid and controlled formation of solid-supported hybrid membranes provides the basis for developing customized artificial hybrid membranes

Porphyrin-polymer nanocompartments: singlet oxygen generation and antimicrobial activity

A new water-soluble photocatalyst for singlet oxygen generation is presented. Its absorption extends to the red part of the spectrum, showing activity up to irradiation at 660 nm. Its efficiency has been compared to that of a commercial analogue (Rose Bengal) for the oxidation of L-methionine. The quantitative and selective oxidation was promising enough to encapsulate the photocatalyst in polymersomes. The singlet oxygen generated in this way can diffuse and remain active for the oxidation of L-methionine outside the polymeric compartment. These results made us consider the use of these polymersomes for antimicrobial applications. E. Coli colonies were subjected to oxidative stress using the photocatalyst-polymersome conjugates and nearly all the colonies were damaged upon extensive irradiation while under the same red LED light irradiation, liquid cultures in the absence of porphyrin or porphyrin-loaded polymersomes were unharme

Porphyrin Containing Polymersomes with Enhanced ROS Generation Efficiency: in vitro evaluation

Abstract Porphyrins are molecules possessing unique photophysical properties making them suitable for application in photodynamic therapy. The incorporation of porphyrins into natural or synthetic nano-assemblies such as polymersomes is a strategy to improve and prolong their therapeutic capacities and to overcome their limitations as therapeutic and diagnostic agents. Here, 5,10,15,20-tetrakis(1-(6-ethoxy-6-oxohexyl)-4-pyridin-1-io)-21H,23H-porphyrin tetrabromide porphyrin is inserted into polymersomes in order to demonstrate that the encapsulation enhances its ability to generate highly reactive singlet oxygen (1O2) upon irradiation in vitro. The photoactivation of the free and polymersome-encapsulated porphyrin is evaluated by electron spin resonance and cell viability assays on three different mammalian cell lines. The results indicate that by encapsulating the porphyrin, a controlled ROS delivery within the cells is achieved, at the same time avoiding side effects such as dark toxicity, non-specific porphyrin release and over time decreased activity in vitro. This work focuses on showing a not-toxic model system for modern therapeutic nanomedicine, which works under mild irradiation and dosage conditions

Biomolecules Turn Self-Assembling Amphiphilic Block Co-polymer Platforms Into Biomimetic Interfaces

Biological membranes constitute an interface between cells and their surroundings and form distinct compartments within the cell. They also host a variety of biomolecules that carry out vital functions including selective transport, signal transduction and cell-cell communication. Due to the vast complexity and versatility of the different membranes, there is a critical need for simplified and specific model membrane platforms to explore the behaviors of individual biomolecules while preserving their intrinsic function. Information obtained from model membrane platforms should make invaluable contributions to current and emerging technologies in biotechnology, nanotechnology and medicine. Amphiphilic block co-polymers are ideal building blocks to create model membrane platforms with enhanced stability and robustness. They form various supramolecular assemblies, ranging from three-dimensional structures (e.g., micelles, nanoparticles, or vesicles) in aqueous solution to planar polymer membranes on solid supports (e.g., polymer cushioned/tethered membranes,) and membrane-like polymer brushes. Furthermore, polymer micelles and polymersomes can also be immobilized on solid supports to take advantage of a wide range of surface sensitive analytical tools. In this review article, we focus on self-assembled amphiphilic block copolymer platforms that are hosting biomolecules. We present different strategies for harnessing polymer platforms with biomolecules either by integrating proteins or peptides into assemblies or by attaching proteins or DNA to their surface. We will discuss how to obtain synthetic structures on solid supports and their characterization using different surface sensitive analytical tools. Finally, we highlight present and future perspectives of polymer micelles and polymersomes for biomedical applications and those of solid-supported polymer membranes for biosensing

Self assembling block copolymers : a versatile tool for creating functional 3D and planar membranes

Driven by self assembly, amphiphilic block copolymers can be used as the basis for the creation of various artificial nano and micro-devices which can: i) deliver therapeutic agents ii) imitate functionalities found in nature and thus give us the opportunity to study and understand them iii) be engineered for biosensing and bioanalytical applications. Highlighting the versatility of block copolymers, this work describes how we can use amphiphilic block copolymers as a to assemble nanometer-sized vesicles i.e., polymersomes for reactive oxygen species (ROS) delivery in vitro, and solid supporting polymer membranes for the development of an applicable platform. First, we used polymersomes to deliver ROS in vitro, in a controlled and biocompatible manner. A water-soluble porphyrin, which generates ROS upon irradiation under red LED light, was encapsulated in the aqueous cavity of polymersomes. The porphyrin-incorporating polymersomes were then co-cultured with Escherichia coli bacteria and three different mammalian cells lines (HeLa, HEK293T and HepG2). Next, they were irradiated to in situ photoactivate the prophryrin to produce ROS. The evaluation of our experimental findings allowed us to optimize the encapsulation and process and most importantly to find out that the polymer membrane has many valuable advantages for our system. More specifically, the low permeability of the polymer membrane limits the intrinsic toxicity of the porphyrin. At the same time, the encapsulation prolongs the ROS generation within cells. Furthermore, the ROS delivery is ’on-demand’ and only occurs upon irradiation. The following significant decrease of viability for both bacteria and mammalian cells in vitro was verified by corresponding viability assays as well as EPR spectroscopy and confocal laser microscopy. The triggerable ROS generating polymersomes proved to be promising nano-carriers for photodynamic therapy. The second part of this thesis moves from 3D polymersomes based on poly(dimethylsiloxane)-poly(2-methyl-2-oxazoline) (PDMS-PMOXA) amphiphilic triblock copolymers to their planar counterpart. Creating a solid supported polymer membrane by using the vesicle fusion method is established for phospholipids but not for polymersomes. In order to overcome this challenge, we propose a simple procedure for creating planar polymer membranes. Here, it’s worth to mention that polymersomes are known to have a robust yet flexible outer membrane which makes their transition from hollow spherical to planar structures a demanding task. We decided to use thiol-modified polymersomes to facilitate their adsorption onto a gold coated surface. Then, we triggered the rupture of the polymersomes and the corresponding membrane formation via Ca 2+ induced osmotic shock. Using sensitive surface characterization techniques such quartz crystal microbalance with dissipation (QCM-D) monitoring, atomic force microscopy (AFM), spectroscopic ellipsometry and brewster angle microscopy (BAM) we gained a deeper understanding of polymer membrane formation requirements. Our findings suggest that i) the length of the single polymer chains and the resulting membrane thickness, ii) the attachment on solid support and iii) the external stimulus for the membrane formation have to be considered as crucial parameters. These results open up new perspectives on the creation of block copolymer based, cellular membrane-mimicking platforms

FAP Targeting of Photosensitizer-Loaded Polymersomes for Increased Light-Activated Cell Killing

As current chemo- and photodynamic cancer therapies are associated with severe side effects due to a lack of specificity and to systemic toxicity, innovative solutions in terms of targeting and controlled functionality are in high demand. Here, we present the development of a polymersome nanocarrier equipped with targeting molecules and loaded with photosensitizers for efficient uptake and light-activated cell killing. Polymersomes were self-assembled in the presence of photosensitizers from a mixture of nonfunctionalized and functionalized PDMS-b-PMOXA diblock copolymers, the latter designed for coupling with targeting ligands. By encapsulation inside the polymersomes, the photosensitizer Rose Bengal was protected, and its uptake into cells was mediated by the nanocarrier. Inhibitor of fibroblast activation protein α (FAPi), a ligand for FAP, was attached to the polymersomes’ surface and improved their uptake in MCF-7 breast cancer cells expressing relatively high levels of FAP on their surface. Once internalized by MCF-7, irradiation of Rose Bengal-loaded FAPi-polymersomes generated reactive oxygen species at levels high enough to induce cell death. By combining photosensitizer encapsulation and specific targeting, polymersomes represent ideal candidates as therapeutic nanocarriers in cancer treatment