Dual-Responsive Enzyme-Polysaccharide Conjugate as a Nanocarrier System for Enzyme Prodrug Therapy.

Biopolymer-based drug delivery systems have gained considerable attention in the field of nanomedicine. In this study, a protein-polysaccharide conjugate was synthesized by covalent conjugation of the enzyme horseradish peroxidase (HRP) with acetalated dextran (AcDex) via a thiol exchange reaction. The resulting bioconjugate shows a dual-responsive behavior in acidic and reductive environments to achieve a controlled release of drugs. The self-assembly of this amphiphilic HRP-AcDex conjugate allows the encapsulation of prodrug indole-3-acetic acid (IAA) into the hydrophobic polysaccharide core. Under slightly acidic conditions, the acetalated polysaccharide reverts to its native hydrophilic form, which triggers the disassembly of micellar nanoparticles and the release of the encapsulated prodrug. The conjugated HRP further activates the prodrug by oxidation of IAA into cytotoxic radicals, which leads to cellular apoptosis. The results indicate that the HRP-AcDex conjugate in combination with IAA has great potential to be used as a novel enzyme prodrug therapy for cancer treatment.

Functional DNA-Polymer Conjugates.

DNA nanotechnology has seen large developments over the last 30 years through the combination of solid phase synthesis and the discovery of DNA nanostructures. Solid phase synthesis has facilitated the availability of short DNA sequences and the expansion of the DNA toolbox to increase the chemical functionalities afforded on DNA, which in turn enabled the conception and synthesis of sophisticated and complex 2D and 3D nanostructures. In parallel, polymer science has developed several polymerization approaches to build di- and triblock copolymers bearing hydrophilic, hydrophobic, and amphiphilic properties. By bringing together these two emerging technologies, complementary properties of both materials have been explored; for example, the synthesis of amphiphilic DNA-polymer conjugates has enabled the production of several nanostructures, such as spherical and rod-like micelles. Through both the DNA and polymer parts, stimuli-responsiveness can be instilled. Nanostructures have consequently been developed with responsive structural changes to physical properties, such as pH and temperature, as well as short DNA through competitive complementary binding. These responsive changes have enabled the application of DNA-polymer conjugates in biomedical applications including drug delivery. This review discusses the progress of DNA-polymer conjugates, exploring the synthetic routes and state-of-the-art applications afforded through the combination of nucleic acids and synthetic polymers.

End Group Dye-Labeled Polycarbonate Block Copolymers for Micellar (Immuno-)Drug Delivery.

Defined conjugation of functional molecules to block copolymer end groups is a powerful strategy to enhance the scope of micellar carriers for drug delivery. In this study, an approach to access well-defined polycarbonate-based block copolymers by labeling their end groups with single fluorescent dye molecules is established. Following controlled polymerization conditions, the block copolymers' primary hydroxy end group can be converted into activated pentafluorophenyl ester carbonates and subsequently aminolyzed with fluorescent dyes that are equipped with primary amines. During a solvent-evaporation process, the resulting end group dye-labeled block copolymers self-assemble into narrowly dispersed ∼25 nm-sized micelles and simultaneously encapsulate hydrophobic (immuno-)drugs. The covalently attached fluorescent tracer can be used to monitor both uptake into cells and stability under biologically relevant conditions, including incubation with blood plasma or during blood circulation in zebrafish embryos. By encapsulation of the toll-like receptor 7/8 (TLR7/8) agonist CL075, immune stimulatory polymeric micelles are generated that get internalized by various antigen-presenting dendritic cells and promote their maturation. Generally, such end group dye-labeled polycarbonate block copolymers display ideal features to permit targeted delivery of hydrophobic drugs to key immune cells for vaccination and cancer immunotherapy.

Multiple Wavelength Photopolymerization of Stable Poly(Catecholamines)-DNA Origami Nanostructures.

The synthesis of multicomponent polymer hybrids with nanometer precision is chemically challenging in the bottom-up synthesis of complex nanostructures. Here, we leverage the fidelity of the DNA origami technique to install a multiple wavelength responsive photopolymerization system with nanometer resolution. By precisely immobilizing various photosensitizers on the origami template, which are only activated at their respective maximum wavelength, we can control sequential polymerization processes. In particular, the triggered photosensitizers generate reactive oxygen species that in turn initiate the polymerization of the catecholamines dopamine and norepinephrine. We imprint polymeric layers at designated positions on DNA origami, which modifies the polyanionic nature of the DNA objects, thus promoting their uptake into living cells while preserving their integrity. Our herein proposed method provides a rapid platform to access complex 3D nanostructures by customizing material and biological interfaces.

Squaric Ester-Based, pH-Degradable Nanogels: Modular Nanocarriers for Safe, Systemic Administration of Toll-like Receptor 7/8 Agonistic Immune Modulators.

Small-molecular Toll-like receptor 7/8 (TLR7/8) agonists hold promise as immune modulators for a variety of immune therapeutic purposes including cancer therapy or vaccination. However, due to their rapid systemic distribution causing difficult-to-control inflammatory off-target effects, their application is still problematic, in particular systemically. To address this problem, we designed and robustly fabricated pH-responsive nanogels serving as versatile immunodrug nanocarriers for safe delivery of TLR7/8-stimulating imidazoquinolines after intravenous administration. To this aim, a primary amine-reactive methacrylamide monomer bearing a pendant squaric ester amide is introduced, which is polymerized under controlled RAFT polymerization conditions. Corresponding PEG-derived squaric ester amide block copolymers self-assemble into precursor micelles in polar protic solvents. Their cores are amine-reactive and can sequentially be transformed by acid-sensitive cross-linkers, dyes, and imidazoquinolines. Remaining squaric ester amides are hydrophilized affording fully hydrophilic nanogels with profound stability in human plasma but stimuli-responsive degradation upon exposure to endolysosomal pH conditions. The immunomodulatory behavior of the imidazoquinolines alone or conjugated to the nanogels was demonstrated by macrophages in vitro. In vivo, however, we observed a remarkable impact of the nanogel: After intravenous injection, a spatially controlled immunostimulatory activity was evident in the spleen, whereas systemic off-target inflammatory responses triggered by the small-molecular imidazoquinoline analogue were absent. These findings underline the potential of squaric ester-based, pH-degradable nanogels as a promising platform to permit intravenous administration routes of small-molecular TLR7/8 agonists and, thus, the opportunity to explore their adjuvant potency for systemic vaccination or cancer immunotherapy purposes.

DNA Origami Meets Polymers: A Powerful Tool for the Design of Defined Nanostructures.

The combination of DNA origami nanostructures and polymers provides a new possibility to access defined structures in the 100 nm range. In general, DNA origami serves as a versatile template for the highly specific arrangement of polymer chains. Polymer-DNA hybrid nanostructures can either be created by growing the polymer from the DNA template or by attaching preformed polymers to the DNA scaffold. These conjugations can be of a covalent nature or be based on base-pair hybridization between respectively modified polymers and DNA origami. Furthermore, the negatively charged DNA backbone permits interaction with positively charged polyelectrolytes to form stable complexes. The combination of polymers with tuneable characteristics and DNA origami allows the creation of a new class of hybrid materials, which could offer exciting applications for controlled energy transfer, nanoscale organic circuits, or the templated synthesis of nanopatterned polymeric structures.

Photocontrolled Dopamine Polymerization on DNA Origami with Nanometer Resolution.

Temporal and spatial control over polydopamine formation on the nanoscale can be achieved by installing an irradiation-sensitive polymerization system on DNA origami. Precisely distributed G-quadruplex structures on the DNA template serve as anchors for embedding the photosensitizer protoporphyrin IX, which-upon irradiation with visible light-induces the multistep oxidation of dopamine to polydopamine, producing polymeric structures on designated areas within the origami framework. The photochemical polymerization process allows exclusive control over polydopamine layer formation through the simple on/off switching of the light source. The obtained polymer-DNA hybrid material shows significantly enhanced stability, paving the way for biomedical and chemical applications that are typically not possible owing to the sensitivity of DNA.

Photoinduced Amyloid Fibril Degradation for Controlled Cell Patterning.

Amyloid-like fibrils are a special class of self-assembling peptides that emerge as a promising nanomaterial with rich bioactivity for applications such as cell adhesion and growth. Unlike the extracellular matrix, the intrinsically stable amyloid-like fibrils do not respond nor adapt to stimuli of their natural environment. Here, a self-assembling motif (CKFKFQF), in which a photosensitive o-nitrobenzyl linker (PCL) is inserted, is designed. This peptide (CKFK-PCL-FQF) assembles into amyloid-like fibrils comparable to the unsubstituted CKFKFQF and reveals a strong response to UV-light. After UV irradiation, the secondary structure of the fibrils, fibril morphology, and bioactivity are lost. Thus, coating surfaces with the pre-formed fibrils and exposing them to UV-light through a photomask generate well-defined areas with patterns of intact and destroyed fibrillar morphology. The unexposed, fibril-coated surface areas retain their ability to support cell adhesion in culture, in contrast to the light-exposed regions, where the cell-supportive fibril morphology is destroyed. Consequently, the photoresponsive peptide nanofibrils provide a facile and efficient way of cell patterning, exemplarily demonstrated for A549, Chinese Hamster Ovary, and Raw Dual type cells. This study introduces photoresponsive amyloid-like fibrils as adaptive functional materials to precisely arrange cells on surfaces.

Nanoscale patterning of polymers on DNA origami.

The combination of DNA-origami and synthetic polymers paves the way to a new class of structurally precise biohybrid nanomaterials for diverse applications. Herein, we introduce the grafting to method with high conversions (70-90%) under ambient conditions to generate DNA-polymer conjugates, which can hybridized precisely to DNA-origami architectures. We generated homo and block copolymers from three different polymer families (acrylates, methacrylates and acrylamides), coupled them to single stranded DNA (ssDNA) and pattern different DNA-origami architectures to demonstrate the formation of precise surface nanopatterns.

DNA-Programmed Chemical Synthesis of Polymers and Inorganic Nanomaterials.

DNA nanotechnology, based on sequence-specific DNA recognition, could allow programmed self-assembly of sophisticated nanostructures with molecular precision. Extension of this technique to the preparation of broader types of nanomaterials would significantly improve nanofabrication technique to lower nanometer scale and even achieve single molecule operation. Using such exquisite DNA nanostructures as templates, chemical synthesis of polymer and inorganic nanomaterials could also be programmed with unprecedented accuracy and flexibility. This review summarizes recent advances in the synthesis and assembly of polymer and inorganic nanomaterials using DNA nanostructures as templates, and discusses the current challenges and future outlook of DNA templated nanotechnology.

Supramolecular Toxin Complexes for Targeted Pharmacological Modulation of Polymorphonuclear Leukocyte Functions.

The targeted pharmacological modulation of polymorphonuclear leukocytes (PMNs) is of major medical interest. These innate immune cells play a central role in the defense against pathogenic microorganisms. However, their excessive chemotactic recruitment into tissues after traumatic injury is detrimental due to local and systemic inflammation. Rho-GTPases, being the master regulators of the actin cytoskeleton, regulate migration and chemotaxis of PMNs, are attractive pharmacological targets. Herein, supramolecular protein complexes are assembled in a "mix-and-match" approach containing the specific Rho-inhibiting clostridial C3 enzyme and three PMN-binding peptides using an avidin platform. Selective delivery of the C3 Rho-inhibitor with these complexes into the cytosol of human neutrophil-like NB-4 cells and primary human PMNs ex vivo is demonstrated, where they catalyze the adenosine diphosphate (ADP) ribosylation of Rho and induce a characteristic change in cell morphology. Notably, the complexes do not deliver C3 enzyme into human lung epithelial cells, A549 lung cancer cells, and immortalized human alveolar epithelial cells (hAELVi), demonstrating their cell type-selectivity. The supramolecular complexes represent attractive molecular tools to decipher the role of PMNs in infection and inflammation or for the development of novel therapeutic approaches for diseases that are associated with hyperactivity and reactivity of PMNs such as post-traumatic injury.