Publications by authors named "Kaloian Koynov"

113 Publications

Fluorescence correlation spectroscopy to unravel the interactions between macromolecules in wine.

Food Chem 2021 Feb 20;352:129343. Epub 2021 Feb 20.

Max Planck Institute for Polymer Research, Ackermannweg 10, 55128 Mainz, Germany. Electronic address:

In this work, the interaction of wine macromolecules with a bovine serum albumin (BSA) was investigated using fluorescence correlation spectroscopy (FCS). FCS offers the opportunity to study molecular and macromolecular aggregation without disturbing the wine by introducing only very small amounts of fluorescently labelled molecules to the system. It was observed that the diffusion coefficient of fluorescently labelled BSA varies with the addition of wine macromolecules, indicating changes in the protein conformation and the formation of complexes and aggregates. The addition of a wine polysaccharide rhamnogalacturonan II-enriched fraction led to aggregation, while addition of a mannoprotein-enriched fraction exhibited a protective effect on protein aggregation. Proteins strongly interacted with tannins, leading to the precipitation of protein-tannin complexes, while the presence of polysaccharides prevented this precipitation. Finally, the application of FCS was demonstrated in real wines, to investigate the problem of protein haze formation through live monitoring of heat-induced aggregation in wine.
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http://dx.doi.org/10.1016/j.foodchem.2021.129343DOI Listing
February 2021

Lipid-Polyglutamate Nanoparticle Vaccine Platform.

ACS Appl Mater Interfaces 2021 Feb 28;13(5):6011-6022. Epub 2021 Jan 28.

Department of Pharmaceutics, Ghent University, 9000 Ghent, Belgium.

Peptide-based subunit vaccines are attractive in view of personalized cancer vaccination with neo-antigens, as well as for the design of the newest generation of vaccines against infectious diseases. Key to mounting robust antigen-specific immunity is delivery of antigen to antigen-presenting (innate immune) cells in lymphoid tissue with concomitant innate immune activation to promote antigen presentation to T cells and to shape the amplitude and nature of the immune response. Nanoparticles that co-deliver both peptide antigen and molecular adjuvants are well suited for this task. However, in the context of peptide-based antigen, an unmet need exists for a generic strategy that allows for co-encapsulation of peptide and molecular adjuvants due to the stark variation in physicochemical properties based on the amino acid sequence of the peptide. These properties also strongly differ from those of many molecular adjuvants. Here, we devise a lipid nanoparticle (LNP) platform that addresses these issues. Key in our concept is poly(l-glutamic acid) (PGA), which serves as a hydrophilic backbone for conjugation of, respectively, peptide antigen (Ag) and an imidazoquinoline (IMDQ) TLR7/8 agonist as a molecular adjuvant. Making use of the PGA's polyanionic nature, we condensate PGA-Ag and PGA-IMDQ into LNP by electrostatic interaction with an ionizable lipid. We show in vitro and in vivo in mouse models that LNP encapsulation favors uptake by innate immune cells in lymphoid tissue and promotes the induction of Ag-specific T cells responses both after subcutaneous and intravenous administration.
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http://dx.doi.org/10.1021/acsami.0c20607DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7116839PMC
February 2021

From Macro to Mesoporous ZnO Inverse Opals: Synthesis, Characterization and Tracer Diffusion Properties.

Nanomaterials (Basel) 2021 Jan 14;11(1). Epub 2021 Jan 14.

Institute for Materials Science, University of Stuttgart, 70569 Stuttgart, Germany.

Oxide inverse opals (IOs) with their high surface area and open porosity are promising candidates for catalyst support applications. Supports with confined mesoporous domains are of added value to heterogeneous catalysis. However, the fabrication of IOs with mesoporous or sub-macroporous voids (<100 nm) continues to be a challenge, and the diffusion of tracers in quasi-mesoporous IOs is yet to be adequately studied. In order to address these two problems, we synthesized ZnO IOs films with tunable pore sizes using chemical bath deposition and template-based approach. By decreasing the size of polystyrene (PS) template particles towards the mesoporous range, ZnO IOs with 50 nm-sized pores and open porosity were synthesized. The effect of the template-removal method on the pore geometry (spherical vs. gyroidal) was studied. The infiltration depth in the template was determined, and the factors influencing infiltration were assessed. The crystallinity and photonic stop-band of the IOs were studied using X-Ray diffraction and UV-Vis, respectively. The infiltration of tracer molecules (Alexa Fluor 488) in multilayered quasi-mesoporous ZnO IOs was confirmed via confocal laser scanning microscopy, while fluorescence correlation spectroscopy analysis revealed two distinct diffusion times in IOs assigned to diffusion through the pores (fast) and adsorption on the pore walls (slow).
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http://dx.doi.org/10.3390/nano11010196DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7828802PMC
January 2021

Adaptation of a Styrene-Acrylic Acid Copolymer Surface to Water.

Langmuir 2021 Feb 13;37(4):1571-1577. Epub 2021 Jan 13.

Max Planck Institute for Polymer Research, Ackermannweg 10, 55128 Mainz, Germany.

Solid surfaces, in particular polymer surfaces, are able to adapt upon contact with a liquid. Adaptation results in an increase in contact angle hysteresis and influences the mobility of sliding drops on surfaces. To study adaptation and its kinetics, we synthesized a random copolymer composed of styrene and 11-25 mol% acrylic acid (PS/PAA). We measured the dynamic advancing (θ) and receding (θ) contact angles of water drops sliding down a tilted plate coated with this polymer. We measured θ ≈ 87° for velocities of the contact line <20 μm/s. At higher velocities, θ gradually increased to ∼98°. This value is similar to θ of a pure polystyrene (PS) film, which we studied for comparison. We associate the gradual increase in θ to the adaptation process to water: The presence of water leads to swelling and/or an enrichment of acid groups at the water/polymer interface. By applying the latest adaptation theory (Butt et al. 2018, 34, 11292), we estimated the time constant of this adaptation process to be ≪1 s. For sliding water drops, θ is ∼10° lower compared to the reference PS surface for all tested velocities. Thus, at the receding side of a sliding drop, the surface is already enriched by acid groups. For a water drop with a width of 5 mm, the increase in contact angle hysteresis corresponds to an increase in capillary force in the range of 45-60 μN, depending on sliding velocity.
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http://dx.doi.org/10.1021/acs.langmuir.0c03226DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7880564PMC
February 2021

Influence of Riboflavin Targeting on Tumor Accumulation and Internalization of Peptostar Based Drug Delivery Systems.

Bioconjug Chem 2020 12 25;31(12):2691-2696. Epub 2020 Nov 25.

Institute for Experimental Molecular Imaging, University Hospital Aachen, Forckenbeckstrasse 55, 52074 Aachen, Germany.

Riboflavin carrier protein (RCP) and riboflavin transporters (RFVTs) have been reported to be highly overexpressed in various cancer cells. Hence, targeting RCP and RFVTs using riboflavin may enhance tumor accumulation and internalization of drug delivery systems. To test this hypothesis, butyl-based 3-arm peptostar polymers were synthesized consisting of a lysine core (10 units per arm) and a sarcosine shell (100 units per arm). The end groups of the arms and the core were successfully modified with riboflavin and the Cy5.5 fluorescent dye, respectively. While in phosphate buffered saline the functionalized peptostars showed a bimodal behavior and formed supramolecular structures over time, they were stable in the serum maintaining their hydrodynamic diameter of 12 nm. Moreover, the polymers were biocompatible and the uptake of riboflavin targeted peptostars in A431 and PC3 cells was higher than in nontargeted controls and could be blocked competitively. In vivo, the polymers showed a moderate passive tumor accumulation, which was not significantly different between targeted and nontargeted peptostars. Nonetheless, at the histological level, internalization into tumor cells was strongly enhanced for the riboflavin-targeted peptostars. Based on these results, we conclude that passive accumulation is dominating the accumulation of peptostars, while tumor cell internalization is strongly promoted by riboflavin targeting.
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http://dx.doi.org/10.1021/acs.bioconjchem.0c00593DOI Listing
December 2020

Glass Transition of Disentangled and Entangled Polymer Melts: Single-Chain-Nanoparticles Approach.

Macromolecules 2020 Sep 20;53(17):7312-7321. Epub 2020 Aug 20.

Max Planck Institute for Polymer Research, Ackermannweg 10, D-55128 Mainz, Germany.

We study the effect of entanglements on the glass transition of high molecular weight polymers, by the comparison of single-chain nanoparticles (SCNPs) and equilibrated melts of high-molecular weight polystyrene of identical molecular weight. SCNPs were prepared by electrospraying technique and characterized using scanning electron microscopy and atomic force microscopy techniques. Differential scanning calorimetry, Brillouin light spectroscopy, and rheological experiments around the glass transition were compared. In parallel, entangled and disentangled polymer melts were also compared under cooling from molecular dynamics simulations based on a bead-spring polymer model. While experiments suggest a small decrease in the glass transition temperature of films of nanoparticles in comparison to entangled melts, simulations do not observe any significant difference, despite rather different chain conformations.
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http://dx.doi.org/10.1021/acs.macromol.0c00550DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7482400PMC
September 2020

Brownian Diffusion of Individual Janus Nanoparticles at Water/Oil Interfaces.

ACS Nano 2020 Aug 6;14(8):10095-10103. Epub 2020 Aug 6.

Max Planck Institute for Polymer Research, Ackermannweg 10, 55128, Mainz, Germany.

Janus nanoparticles could exhibit a higher interfacial activity and adsorb stronger to fluid interfaces than homogeneous nanoparticles of similar sizes. However, little is known about the interfacial diffusion of Janus nanoparticles and how it compares to that of homogeneous ones. Here, we employed fluorescence correlation spectroscopy to study the lateral diffusion of ligand-grafted Janus nanoparticles adsorbed at water/oil interfaces. We found that the diffusion was significantly slower than that of homogeneous nanoparticles. We carried out dissipative particle dynamic simulations to study the mechanism of interfacial slowdown. Good agreement between experimental and simulation results has been obtained only provided that the flexibility of ligands grafted on the nanoparticle surface was taken into account. The polymeric ligands were deformed and oriented at an interface so that the effective radius of Janus nanoparticles is larger than the nominal one obtained by measuring the diffusion in bulk solution. These findings highlight further the critical importance of the ligands grafted on Janus nanoparticles for applications involving nanoparticle adsorption at an interface, such as oil recovery or two-dimensional self-assembly.
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http://dx.doi.org/10.1021/acsnano.0c03291DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7458482PMC
August 2020

DNA-Polymer Nanostructures by RAFT Polymerization and Polymerization-Induced Self-Assembly.

Angew Chem Int Ed Engl 2020 09 17;59(36):15474-15479. Epub 2020 Jun 17.

Synthesis of Macromolecules, Max Planck Institute for Polymer Research, Ackermannweg 10, 55128, Mainz, Germany.

Nanostructures derived from amphiphilic DNA-polymer conjugates have emerged prominently due to their rich self-assembly behavior; however, their synthesis is traditionally challenging. Here, we report a novel platform technology towards DNA-polymer nanostructures of various shapes by leveraging polymerization-induced self-assembly (PISA) for polymerization from single-stranded DNA (ssDNA). A "grafting from" protocol for thermal RAFT polymerization from ssDNA under ambient conditions was developed and utilized for the synthesis of functional DNA-polymer conjugates and DNA-diblock conjugates derived from acrylates and acrylamides. Using this method, PISA was applied to manufacture isotropic and anisotropic DNA-polymer nanostructures by varying the chain length of the polymer block. The resulting nanostructures were further functionalized by hybridization with a dye-labelled complementary ssDNA, thus establishing PISA as a powerful route towards intrinsically functional DNA-polymer nanostructures.
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http://dx.doi.org/10.1002/anie.201916177DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7496909PMC
September 2020

Metallopolymer Organohydrogels with Photo-Controlled Coordination Crosslinks Work Properly Below 0 °C.

Adv Mater 2020 Apr 24;32(14):e1908324. Epub 2020 Feb 24.

CAS Key Laboratory of Soft Matter Chemistry, Hefei National Laboratory for Physical Sciences at the Microscale, Department of Polymer Science and Engineering, University of Science and Technology of China, Hefei, 230026, China.

Controlling the structures and functions of gels is important for both fundamental research and technological applications. Introducing photoresponsive units into gels enables remote control of their properties with light. However, existing gels show photoresponsiveness only at room temperature or elevated temperatures. The development of photoresponsive gels that work below 0 °C can expand their usage in cold environments. Here, photoresponsive metallopolymer organohydrogels that function even at -20 °C are reported. The organohydrogels are prepared using photoresponsive Ru-thioether coordination bonds as reversible crosslinks to form polymer networks. A water/glycerol mixture is used as an anti-freezing solvent. At -20 °C, the Ru-thioether coordination bonds are dissociated under light irradiation and reformed reversibly in the dark, which result in alternating crosslinking densities in the polymer networks. This process enables inducing reversible gel-to-sol transitions, healing damaged gels, controlling the mechanical properties and volumes of the gels, and rewriting microstructures on the gels below 0 °C.
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http://dx.doi.org/10.1002/adma.201908324DOI Listing
April 2020

Precision Anisotropic Brush Polymers by Sequence Controlled Chemistry.

J Am Chem Soc 2020 01 26;142(3):1332-1340. Epub 2019 Dec 26.

Max Planck Institute for Polymer Research , Ackermannweg 10 , 55128 Mainz , Germany.

The programming of nanomaterials at molecular length-scales to control architecture and function represents a pinnacle in soft materials synthesis. Although elusive in synthetic materials, Nature has evolutionarily refined macromolecular synthesis with perfect atomic resolution across three-dimensional space that serves specific functions. We show that biomolecules, specifically proteins, provide an intrinsic macromolecular backbone for the construction of anisotropic brush polymers with monodisperse lengths via grafting-from strategy. Using human serum albumin as a model, its sequence was exploited to chemically transform a single cysteine, such that the expression of said functionality is asymmetrically placed along the backbone of the eventual brush polymer. This positional monofunctionalization strategy was connected with biotin-streptavidin interactions to demonstrate the capabilities for site-specific self-assembly to create higher ordered architectures. Supported by systematic experimental and computational studies, we envisioned that this macromolecular platform provides unique avenues and perspectives in macromolecular design for both nanoscience and biomedicine.
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http://dx.doi.org/10.1021/jacs.9b10491DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6978811PMC
January 2020

Water-dispersed semiconductor nanoplatelets with high fluorescence brightness, chemical and colloidal stability.

J Mater Chem B 2020 01 29;8(1):146-154. Epub 2019 Nov 29.

Max-Planck-Institute for Polymer Research, Ackermannweg 10, 55128 Mainz, Germany.

Quasi-two dimensional semiconductor nanoplatelets (NPLs) exhibit high spectral brightness and large absorption cross sections, making them promising for various applications including bioimaging. However, the synthesis of NPLs takes place in organic solvents, therefore they require phase transfer in order to use them in aqueous environments. The phase transfer of NPLs has so far been challenging with few examples in literature. This is likely due to the facile agglomeration of materials with plate-like geometries during the coating procedure. Here we demonstrate how to overcome agglomeration and transfer NPLs, individually coated with amphiphilic polymer chains, to aqueous phase. Upon one and two-photon excitation the water transferred NPLs exhibit more than two fold higher fluorescent brightness relative to commercially available quantum dots. Additionally, the polymer coating increase the stability of nanoparticles in physiological conditions (pH 4.5-7.4, [NaCl] 5.8-11.7 g L, and in human serum). Our experiments with NPL labeled RAW264.7 cells demonstrate the capabilities of NPLs as next generation ultra-bright fluorescent labels for bioimaging.
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http://dx.doi.org/10.1039/c9tb02377aDOI Listing
January 2020

Tackling the Limitations of Copolymeric Small Interfering RNA Delivery Agents by a Combined Experimental-Computational Approach.

Biomacromolecules 2019 12 4;20(12):4389-4406. Epub 2019 Nov 4.

Institute of Organic Chemistry and Macromolecular Chemistry , Friedrich Schiller University Jena , Lessingstraße 8 , 07743 Jena , Germany.

Despite the first successful applications of nonviral delivery vectors for small interfering RNA in the treatment of illnesses, such as the respiratory syncytial virus infection, the preparation of a clinically suitable, safe, and efficient delivery system still remains a challenge. In this study, we tackle the drawbacks of the existing systems by a combined experimental-computational in-depth investigation of the influence of the polymer architecture over the binding and transfection efficiency. For that purpose, a library of diblock copolymers with a molar mass of 30 kDa and a narrow dispersity (Đ < 1.12) was synthesized. We studied in detail the impact of an altered block size and/or composition of cationic diblock copolymers on the viability of each respective structure as a delivery agent for polynucleotides. The experimental investigation was further complemented by a computational study employing molecular simulations as well as an analytical description of systemic properties. This is the first report in which molecular dynamics simulations of RNA/cationic polymer complexes have been performed. Specifically, we developed and employed a coarse-grained model of the system at the molecular level to study the interactions between polymer chains and small interfering RNA. We were further able to confirm a threshold length/length ratio, which is required for efficient complexation of siRNA, and it was possible to find a correlation between the length of the cationic block and the size of the resulting polyplex. Hence, the combined insights from the experiments and the theoretical investigation resulted in a wealth of information about the properties of cationic diblock copolymers employed as RNA delivery agents, in particular regarding the molecular and mechanistic details of the interaction between the two components of a polyplex.
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http://dx.doi.org/10.1021/acs.biomac.9b01061DOI Listing
December 2019

Overcoming the barrier of CD8T cells: Two types of nano-sized carriers for siRNA transport.

Acta Biomater 2019 12 4;100:338-351. Epub 2019 Oct 4.

Department of Dermatology, University Medical Center of the Johannes Gutenberg-University Mainz, Langenbeckstr. 1, 55131 Mainz, Germany; Max Planck Institute for Polymer Research, Ackermannweg 10, 55128 Mainz, Germany. Electronic address:

Bioengineering immune cells via gene therapy offers treatment opportunities for currently fatal viral infections. Also cell therapeutics offer most recently a breakthrough technology to combat cancer. These primary human cells, however, are sensitive to toxic influences, which make the utilization of optimized physical transfection techniques necessary. The otherwise commonly applied delivery agents such as Lipofectamine or strongly cationic polymer structures are not only unsuitable for in vivo experiments, but are also highly toxic to immune cells. This study aimed to improve the design of polymeric carrier systems for small interfering RNA, which would allow efficient internalization into CD8T-cells without affecting their viability and thereby removing the current limitations in the field. Here, two new carrier systems for small interfering RNA were tested. One is a cationic diblock copolymer, in which less than 10% of the monomers were modified with triphenylphosphonium cations. This moiety is lipophilic, promotes uptake and it is mostly known for its mitotropic properties. Furthermore, cationic nanohydrogel particles were synthesized in exceedingly small sizes (R < 14 nm). After full physicochemical characterization of the two carriers, extensive cytotoxicity studies were performed and the concentration dependent uptake into CD8T-cells was tested in correlation to incubation time and protein content of the surrounding medium. Both carriers facilitated efficient complexation of siRNA as well as significant internalization into primary human cells in less than three hours of incubation. In addition, neither of the delivery systems reduced cell viability making them good candidates to transport siRNA into CD8T-cells efficiently. STATEMENT OF SIGNIFICANCE: This study provides insights into the design of polymeric delivery agents as the method of choice for overcoming the limitations of cell manipulation. Until now, CD8T-cells, which have become a treatment tool for currently fatal diseases, have not yet been made accessible for gene silencing by polymeric siRNA carrier systems. Choosing appropriate modification approaches for two chemically different polymer structures, we were, in both cases, able to achieve significant uptake in these cells even at low concentrations and without inducing cytotoxicity. These results remove current limitations and pave the way for bioengineering via gene therapy.
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http://dx.doi.org/10.1016/j.actbio.2019.10.006DOI Listing
December 2019

HPMA-Based Nanoparticles for Fast, Bioorthogonal iEDDA Ligation.

Biomacromolecules 2019 10 19;20(10):3786-3797. Epub 2019 Sep 19.

Johannes Gutenberg University Mainz , Institute of Organic Chemistry , Duesbergweg 10-14 , 55128 Mainz , Germany.

Fast and bioorthogonally reacting nanoparticles are attractive tools for biomedical applications such as tumor pretargeting. In this study, we designed an amphiphilic block copolymer system based on HPMA using different strategies to introduce the highly reactive click units 1,2,4,5-tetrazines (Tz) either at the chain end (Tz-CTA) or statistical into the hydrophobic block. This reactive group undergoes a rapid, bioorthogonal inverse electron-demand Diels-Alder reaction (iEDDA) with -cyclooctenes (TCO). Subsequently, this polymer platform was used for the preparation of different Tz-covered nanoparticles, such as micelles and colloids. Thereby it was found that the reactivity of the polymeric micelles is comparable to that of the low molar mass tetrazines. The core-cross-linked micelles can be successfully conjugated at rather low concentrations to large biomacromolecules like antibodies, not only in physiological buffer, but also in human blood plasma, which was confirmed by fluorescence correlation spectroscopy (FCS).
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http://dx.doi.org/10.1021/acs.biomac.9b00868DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6794642PMC
October 2019

Covalently Binding of Bovine Serum Albumin to Unsaturated Poly(Globalide-Co-ε-Caprolactone) Nanoparticles by Thiol-Ene Reactions.

Macromol Biosci 2019 10 6;19(10):e1900145. Epub 2019 Sep 6.

Max Planck Institute for Polymer Research, Ackermannweg 10, 55128, Mainz, Germany.

When nanoparticles (NPs) are introduced to a biological fluid, different proteins (and other biomolecules) rapidly get adsorbed onto their surface, forming a protein corona capable of giving to the NPs a new "identity" and determine their biological fate. Protein-nanoparticle conjugation can be used in order to promote specific interactions between living systems and nanocarriers. Non-covalent conjugates are less stable and more susceptible to desorption in biological media, which makes the development of engineered nanoparticle surfaces by covalent attachment an interesting topic. In this work, the surface of poly(globalide-co-ε-caprolactone) (PGlCL) nanoparticles containing double bonds in the main polymer chain is covalently functionalized with bovine serum albumin (BSA) by thiol-ene chemistry, producing conjugates which are resistant to dissociation. The successful formation of the covalent conjugates is confirmed by flow cytometry (FC) and fluorescence correlation spectroscopy (FCS). Transmission electron microscopy (TEM) allows the visualization of the conjugate formation, and the presence of a protein layer surrounding the NPs can be observed. After conjugation with BSA, NPs present reduced cell uptake by HeLa and macrophage RAW264.7 cells, in comparison to uncoated NP. These results demonstrate that it is possible to produce stable conjugates by covalently binding BSA to PGlCL NP through thiol-ene reaction.
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http://dx.doi.org/10.1002/mabi.201900145DOI Listing
October 2019

Preparation of Monodisperse Giant Unilamellar Anchored Vesicles Using Micropatterned Hydrogel Substrates.

ACS Omega 2019 May 29;4(5):9393-9399. Epub 2019 May 29.

Max Planck Institute for Polymer Research, Ackermannweg 10, 55128 Mainz, Germany.

Giant unilamellar vesicles (GUVs) are model membrane systems consisting of a single lipid bilayer separating an inner lumen from the outer solution, with dimensions comparable to that of eukaryotic cells. The importance of these biomimetic systems has recently grown with the development of easy and safe methods to assemble GUVs from complex biorelevant compositions. However, size and position control is still a key challenge for GUV formation and manipulation. Here, a gel-assisted formation method is introduced, able to produce arrays of giant unilamellar anchored vesicles (GUAVs) with a predetermined narrow size distribution. The approach based on micropatterned gel substrates of cross-linked poly(-isopropylacrylamide) allows performing parallel measurements on thousands of immobile unilamellar vesicles. Such power and flexibility will respond to the growing need for developing platforms of biomimetic constructs from cell-sized single bilayers.
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http://dx.doi.org/10.1021/acsomega.9b00912DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6648857PMC
May 2019

Multifunctional Cationic PeptoStars as siRNA Carrier: Influence of Architecture and Histidine Modification on Knockdown Potential.

Macromol Biosci 2020 01 20;20(1):e1900152. Epub 2019 Aug 20.

Institute of Organic Chemistry, Johannes Gutenberg University Mainz, Duesbergweg 10-14, 55128, Mainz, Germany.

RNA interference provides enormous potential for the treatment of several diseases, including cancer. Nevertheless, successful therapies based on siRNA require overcoming various challenges, such as poor pharmacokinetic characteristics of the small RNA molecule and inefficient cytosolic accumulation. In this respect, the development of functional siRNA carrier systems is a major task in biomedical research. To provide such a desired system, the synthesis of 3-arm and 6-arm PeptoStars is aimed for. The different branched polypept(o)idic architectures share a stealth-like polysarcosine corona for efficient shielding and a multifunctional polylysine core, which can be independently varied in size and functionality for siRNA complexation-, transport and intra cellular release. The special feature of star-like polypept(o)ides is in their uniform small size (<20 nm) and a core-shell structure, which implies a high stability and stealth-like properties and thus, they may combine long circulation times and a deep penetration of cancerous tissue. Initial toxicity and complement studies demonstrate well tolerated cationic PeptoStars with high complexation capability toward siRNA (N/P ratio up to 3:1), which can lead to potent RNAi for optimized systems. Here, the synthetic development of 3-arm and 6-arm polypept(o)idic star polymers, their modification with endosomolytic moieties, and first in vitro insights on RNA interference are reported on.
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http://dx.doi.org/10.1002/mabi.201900152DOI Listing
January 2020

Effects of Spacers on Photoinduced Reversible Solid-to-Liquid Transitions of Azobenzene-Containing Polymers.

Chemistry 2019 Aug 25;25(46):10946-10953. Epub 2019 Jul 25.

CAS Key Laboratory of Soft Matter Chemistry, Hefei National Laboratory for Physical Sciences at the Microscale, Department of Polymer Science and Engineering, University of Science and Technology of China, Jinzhai Road 96, Hefei, 230026, P.R. China.

Photoisomerization in some azobenzene-containing polymers (azopolymers) results in reversible solid-to-liquid transitions because trans- and cis-azopolymers have different glass transition temperatures. This property enables photoinduced healing and processing of azopolymers with high spatiotemporal resolution. However, a general lack of knowledge about the influence of the polymer structure on photoinduced reversible solid-to-liquid transitions hinders the design of such novel polymers. Herein, the synthesis and photoresponsive behavior of new azopolymers with different lengths of spacers between the polymer backbone and the azobenzene group on the side chain are reported. Azopolymers with no and 20 methylene spacers did not show photoinduced solid-to-liquid transitions. Azopolymers with 6 or 12 methylene spacers showed photoinduced solid-to-liquid transitions. This study demonstrates that spacers are essential for azopolymers with photoinduced reversible solid-to-liquid transitions, and thus, gives an insight into how to design azopolymers for photoinduced healing and processing.
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http://dx.doi.org/10.1002/chem.201902273DOI Listing
August 2019

Surfactants Mediate the Dewetting of Acrylic Polymer Films Commonly Applied to Works of Art.

ACS Appl Mater Interfaces 2019 Jul 20;11(30):27288-27296. Epub 2019 Jun 20.

Department of Chemistry , University of Florence and CSGI , Via della Lastruccia 3 , I-50019 Sesto Fiorentino , Firenze , Italy.

The removal of hydrophobic polymer coatings from artistic surfaces is a ubiquitous challenge in art restoration. Over the years, nanostructured fluids (NSFs), aqueous surfactant solutions containing a good solvent for the polymer, have been successfully applied in polymer removal interventions; however, the precise role of the surfactant in promoting polymer film dewetting is not fully understood. This contribution addresses the interaction of a NSF of water/propylene carbonate containing a nonionic surfactant with an acrylic polymer film commonly used in art conservation. Combining confocal microscopy and fluorescence correlation spectroscopy, we monitored the penetration of the fluid into the polymer film, defining its compositional changes and following the polymer swelling. The ensemble of results highlights that the surfactant role is twofold: (i) at the polymer-support interface, it promotes the detachment of the polymer film from the underlying support; (ii) inside the polymer film, it accelerates polymer swelling by increasing the chains' mobility.
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http://dx.doi.org/10.1021/acsami.9b04912DOI Listing
July 2019

Noncovalent Hydrogen Bonds Tune the Mechanical Properties of Phosphoester Polyethylene Mimics.

ACS Omega 2019 May 28;4(5):9324-9332. Epub 2019 May 28.

Max-Planck-Institut für Polymerforschung, Ackermannweg 10, 55128 Mainz, Germany.

Polyethylene mimics of semicrystalline polyphosphoesters (PPEs) with an adjustable amount of noncovalent cross-links were synthesized. Acyclic diene metathesis copolymerization of a phosphoric acid triester () with a novel phosphoric acid diester monomer () was achieved. PPEs with different co-monomer ratios and 0, 20, 40, and 100% of phosphodiester content were synthesized. The phosphodiester groups result in supramolecular interactions between the polymer chains, with the P-OH functionality as an H-bond donor and the P=O group as an H-bond acceptor. A library of unsaturated and saturated PPEs was prepared and analyzed in detail by NMR spectroscopy, size exclusion chromatography, differential scanning calorimetry, thermogravimetry, rheology, and stress-strain measurements. The introduction of the supramolecular cross-links into the aliphatic and hydrophobic PPEs showed a significant impact on the material properties: increased glass-transition and melting temperatures were observed and an increase in the storage modulus of the polymers was achieved. This specific combination of a flexible aliphatic backbone and a supramolecular H-bonding interaction between the chains was maximized in the homopolymer of the phosphodiester monomer, which featured additional properties, such as shape-memory properties, and polymer samples could be healed after cutting. The P-OH groups also showed a strong adhesion toward metal surfaces, which was used together with the shape-memory function in a model device that responds to a temperature stimulus with shape change. This systematic variation of phosphodiesters/phosphotriesters in polyethylene mimics further underlines the versatility of the phosphorus chemistry to build up complex macromolecular architectures.
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http://dx.doi.org/10.1021/acsomega.9b01040DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6545546PMC
May 2019

The mechanics of single cross-links which mediate cell attachment at a hydrogel surface.

Nanoscale 2019 Jun 6;11(24):11596-11604. Epub 2019 Jun 6.

INM - Leibniz Institute for New Materials, Campus D2 2, 66123 Saarbrücken, Germany.

The response of cultured cells to the mechanical properties of hydrogel substrates depends ultimately on the response of single crosslinks to external forces exerted at cell attachment points. We prepared hydrogels by co-polymerization of poly(ethylene glycol diacrylate) (PEGDA) and carboxy poly(ethylene glycol) acrylate (ACPEG-COOH) and confirmed fibroblast spreading on the hydrogel after the ACPEG linker was functionalized with the RGD cell adhesive motif. We performed specific force spectroscopy experiments on the same ACPEG linkers in order to probe the mechanics of single cross-links which mediate the cell attachment and spreading. Measurements were performed with tips of an atomic force microscope (AFM) functionalized with streptavidin and ACPEG linkers functionalized with biotin. We compared hydrogels of varying elastic modulus between 4 and 41 kPa which exhibited significant differences in cell spreading. An effective spring constant for the displacement of single cross-links at the hydrogel surface was derived from the distributions of rupture force and molecular stiffness. A factor of ten in the elastic modulus E of the hydrogel corresponded to a factor of five in the effective spring constant k of single crosslinks, indicating a transition in scaling with the mesh size ξ from the macroscopic E∝ξ to the molecular k∝ξ. The quantification of stiffness and deformation at the molecular length scale contributes to the discussion of mechanisms in force-regulated phenomena in cell biology.
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http://dx.doi.org/10.1039/c9nr01784dDOI Listing
June 2019

Monitoring drug nanocarriers in human blood by near-infrared fluorescence correlation spectroscopy.

Nat Commun 2018 12 13;9(1):5306. Epub 2018 Dec 13.

Max Planck Institute for Polymer Research, Ackermannweg 10, 55128, Mainz, Germany.

Nanocarrier-based drug delivery is a promising therapeutic approach that offers unique possibilities for the treatment of various diseases. However, inside the blood stream, nanocarriers' properties may change significantly due to interactions with proteins, aggregation, decomposition or premature loss of cargo. Thus, a method for precise, in situ characterization of drug nanocarriers in blood is needed. Here we show how the fluorescence correlation spectroscopy that is a well-established method for measuring the size, loading efficiency and stability of drug nanocarriers in aqueous solutions can be used to directly characterize drug nanocarriers in flowing blood. As the blood is not transparent for visible light and densely crowded with cells, we label the nanocarriers or their cargo with near-infrared fluorescent dyes and fit the experimental autocorrelation functions with an analytical model accounting for the presence of blood cells. The developed methodology contributes towards quantitative understanding of the in vivo behavior of nanocarrier-based therapeutics.
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http://dx.doi.org/10.1038/s41467-018-07755-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6294246PMC
December 2018

Impact of Branching on the Solution Behavior and Serum Stability of Starlike Block Copolymers.

Biomacromolecules 2019 01 11;20(1):375-388. Epub 2018 Dec 11.

Institute of Organic Chemistry , Johannes Gutenberg-Universität Mainz , Duesbergweg 10-14 , 55128 Mainz , Germany.

The size control of nanomedicines for tumor diagnosis and therapy is of high importance, since it enables or disables deep and sufficient tumor penetration. Amphiphilic star-shaped block copolypept(o)ides offer substantial promise to precisely adjust the hydrophobic core and the hydrophilic corona, independent of each other, and therefore simultaneously control the size dimension in the interesting size range from 10 to 30 nm. To gain access to core-shell structures of such sizes, 3-arm and 6-arm PeptoStars, based on poly(γ- tert-butyloxycarbonyl-l-glutamate)- b-polysarcosine (pGlu(O tBu)- b-pSar), were prepared via controlled living ring-opening polymerization (ROP) of the corresponding N-carboxyanhydrides. Moreover, size exclusion chromatography (SEC) proves the presence of well-defined star shaped polymers with molecular weights from 38 to 88 kg/mol with low polymer dispersities of 1.16 to 1.23. By varying the α-helical peptide core and maintain a constant polysarcosine corona, hydrodynamic size analyses revealed the importance of using a sufficiently large and dense hydrophilic shielding corona to prevent aggregation of the hydrophobic core and obtain uniform-sized spherical-shaped particles with hydrodynamic diameters below 24 nm. Fluorescence correlation spectroscopy (FCS) additionally demonstrates the absence of protein adsorption in human plasma for 6-arm polypept(o)ide stars and thus confirms polysarcosine as stealthlike material.
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http://dx.doi.org/10.1021/acs.biomac.8b01545DOI Listing
January 2019

DNA-Polymer Conjugates by Photoinduced RAFT Polymerization.

Biomacromolecules 2019 01 20;20(1):212-221. Epub 2018 Nov 20.

Max Planck Institute for Polymer Research , Ackermannweg 10 , 55128 Mainz , Germany.

Conventional grafting-to approaches to DNA-polymer conjugates are often limited by low reaction yields due to the sterically hindered coupling of a presynthesized polymer to DNA. The grafting-from strategy, in contrast, allows one to directly graft polymers from an initiator that is covalently attached to DNA. Herein, we report blue-light-mediated reversible addition-fragmentation chain-transfer (Photo-RAFT) polymerization from two different RAFT agent-terminated DNA sequences using Eosin Y as the photocatalyst in combination with ascorbic acid. Three monomer families (methacrylates, acrylates and acrylamides) were successfully polymerized from DNA employing Photo-RAFT polymerization. We demonstrate that the length of the grown polymer chain can be varied by altering the monomer to DNA-initiator ratio, while the self-assembly features of the DNA strands were maintained. In summary, we describe a convenient, light-mediated approach toward DNA-polymer conjugates via the grafting-from approach.
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http://dx.doi.org/10.1021/acs.biomac.8b01328DOI Listing
January 2019

FRET Monitoring of Intracellular Ketal Hydrolysis in Synthetic Nanoparticles.

Angew Chem Int Ed Engl 2018 08 25;57(33):10760-10764. Epub 2018 Jul 25.

Department of Pharmaceutics, Ghent University, Belgium.

Degradable synthetic crosslinking is a versatile strategy to harness nanomaterials against disassembly in a complex physiological medium prompted by dilution effects or competitive interaction. In particular, chemical bonds such as ketals that are stable at physiological conditions but are cleaved in response to disease-mediated or intracellular conditions (e.g., a mildly acidic pH) are of great relevance for biomedical applications. Despite the range of spectroscopic or chromatographic analyses methods that allow chemical degradation in solution to be assessed, it is much less straightforward to interrogate synthetic nanomaterials for their degradation state when located inside a living organism. We demonstrate a method based on FRET analysis to monitor intracellular disassembly of block-copolymer-derived nanoparticles engineered with a FRET couple on separate polymer chains, which after self-assembly are covalently crosslinked with a pH-sensitive ketal-containing crosslinker.
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http://dx.doi.org/10.1002/anie.201803847DOI Listing
August 2018

Redox-Responsive and Thermoresponsive Supramolecular Nanosheet Gels with High Young's Moduli.

Macromol Rapid Commun 2018 Aug 13;39(15):e1800282. Epub 2018 Jun 13.

Max Planck Institute for Polymer Research (MPIP), Ackermannweg 10, 55128, Mainz, Germany.

Supramolecular gels made from 2D building blocks are emerging as one of the novel multifunctional soft materials for various applications. This study reports on a class of supramolecular nanosheet gels formed through a reversible self-assembly process involving both intramolecular folding and intermolecular self-assembly of poly[oligo(ethylene glycol)-co-(phenyl-capped bithiophenes)]. Such hierarchical self-assembled structure allows the gels to switch between sol and gel states under either redox or thermostimulus. Moreover, the gels illustrate high Young's moduli, compared to their controls that are made from the same oligo(ethylene glycol) and phenyl-capped bithiophenes blocks but have highly covalent-crosslinked structures. The example might open a window for emerging supramolecular 2D materials to develop mechanically robust and stimuli-responsive soft materials without compromising their intrinsic functions.
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http://dx.doi.org/10.1002/marc.201800282DOI Listing
August 2018

Histidine-rich glycoprotein-induced vascular normalization improves EPR-mediated drug targeting to and into tumors.

J Control Release 2018 07 4;282:25-34. Epub 2018 May 4.

Department of Nanomedicine and Theranostics, Institute for Experimental Molecular Imaging, RWTH Aachen University Clinic and Helmholtz Institute for Biomedical Engineering, Aachen, Germany; Department of Targeted Therapeutics, Biomaterial Science and Technology, University of Twente, Enschede, The Netherlands; Department of Pharmaceutics, Utrecht Institute for Pharmaceutical Sciences, Utrecht University, Utrecht, The Netherlands. Electronic address:

Tumors are characterized by leaky blood vessels, and by an abnormal and heterogeneous vascular network. These pathophysiological characteristics contribute to the enhanced permeability and retention (EPR) effect, which is one of the key rationales for developing tumor-targeted drug delivery systems. Vessel abnormality and heterogeneity, however, which typically result from excessive pro-angiogenic signaling, can also hinder efficient drug delivery to and into tumors. Using histidine-rich glycoprotein (HRG) knockout and wild type mice, and HRG-overexpressing and normal t241 fibrosarcoma cells, we evaluated the effect of genetically induced and macrophage-mediated vascular normalization on the tumor accumulation and penetration of 10-20 nm-sized polymeric drug carriers based on poly(N-(2-hydroxypropyl)methacrylamide). Multimodal and multiscale optical imaging was employed to show that normalizing the tumor vasculature improves the accumulation of fluorophore-labeled polymers in tumors, and promotes their penetration out of tumor blood vessels deep into the interstitium.
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http://dx.doi.org/10.1016/j.jconrel.2018.05.002DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6130770PMC
July 2018

Engineering Proteins at Interfaces: From Complementary Characterization to Material Surfaces with Designed Functions.

Angew Chem Int Ed Engl 2018 09 30;57(39):12626-12648. Epub 2018 Aug 30.

Max Planck Institute for Polymer Research, Ackermannweg 10, 55128, Mainz, Germany.

Once materials come into contact with a biological fluid containing proteins, proteins are generally-whether desired or not-attracted by the material's surface and adsorb onto it. The aim of this Review is to give an overview of the most commonly used characterization methods employed to gain a better understanding of the adsorption processes on either planar or curved surfaces. We continue to illustrate the benefit of combining different methods to different surface geometries of the material. The thus obtained insight ideally paves the way for engineering functional materials that interact with proteins in a predetermined manner.
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http://dx.doi.org/10.1002/anie.201712448DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6391961PMC
September 2018

Site-Specific DBCO Modification of DEC205 Antibody for Polymer Conjugation.

Polymers (Basel) 2018 Feb 2;10(2). Epub 2018 Feb 2.

Institute of Organic Chemistry, Johannes-Gutenberg University Mainz, Duesbergweg 10-14, D-55128 Mainz, Germany.

The design of multifunctional polymer-based vectors, forming pDNA vaccines, offers great potential in cancer immune therapy. The transfection of dendritic immune cells (DCs) with tumour antigen-encoding pDNA leads to an activation of the immune system to combat tumour cells. In this work, we investigated the chemical attachment of DEC205 antibodies (aDEC205) as DC-targeting structures to polyplexes of P(Lys)--P(HPMA). The conjugation of a synthetic block copolymer and a biomacromolecule with various functionalities (aDEC205) requires bioorthogonal techniques to avoid side reactions. Click chemistry and in particular the strain-promoted alkyne-azide cycloaddition (SPAAC) can provide the required bioorthogonality. With regard to a SPAAC of both components, we firstly synthesized two different azide-containing block copolymers, P(Lys)--P(HPMA)-N₃(stat) and P(Lys)--P(HPMA)-N₃(end), for pDNA complexation. In addition, the site-specific incorporation of ring-strained dibenzocyclooctyne (DBCO) moieties to the DEC205 antibody was achieved by an enzymatic strategy using bacterial transglutaminase (BTG). The chemical accessibility of DBCO molecules within aDEC205 as well as the accessibility of azide-functionalities on the polyplex' surface were investigated by various SPAAC experiments and characterized by fluorescence correlation spectroscopy (FCS).
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http://dx.doi.org/10.3390/polym10020141DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6414842PMC
February 2018

A modular approach for multifunctional polymersomes with controlled adhesive properties.

Soft Matter 2018 Feb 5;14(6):894-900. Epub 2018 Jan 5.

Max Planck Institute for Polymer Research (MPIP), 55128 Mainz, Germany.

The bottom-up approach in synthetic biology involves the engineering of synthetic cells by designing biological and chemical building blocks, which can be combined in order to mimic cellular functions. The first step for mimicking a living cell is the design of an appropriate compartment featuring a multifunctional membrane. This is of particular interest since it allows for the selective attachment of different groups or molecules to the membrane. In this context, we report on a modular approach for polymeric vesicles, so-called polymersomes, with a multifunctional surface, namely hydroxyl, alkyne and acrylate groups. We demonstrate that the surface of the polymersome can be functionalized to facilitate imaging, via fluorescent dyes, or to improve the specific adhesion to surfaces by using a biotin functionalization. This generally applicable multifunctionality allows for the covalent integration of various molecules in the membrane of a synthetic cell.
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http://dx.doi.org/10.1039/c7sm01885aDOI Listing
February 2018