Cargo Release Research Articles

Stabilizing Entropically Driven Self-Assembly of Self-Immolative Polyurethanes in Water: A Strategy for Tunable Encapsulation Stability and Controlled Cargo Release

Stimuli-responsive amphiphilic polymer assemblies are of great interest in the area of targeted drug delivery applications as they can sequester guest molecules in one set of conditions and release them under another. Hence, developing a strategy of molar mass controlled synthesis, in-depth understanding of the thermodynamics of the self-assembly process, stabilization, and triggered guest release in a controlled fashion would be highly anticipated in the field of drug delivery applications. As polymer molar mass has a significant impact on the self-assembly process or material property of polymers, we focused on a methodology of controlled molar mass polyurethane synthesis using recycled plastic waste and synthesized a series of self-immolative amphiphilic polyurethanes of varying molar masses and polydispersity indexes. All of the polymers were equipped with periodically grafted triethyleneglycol monomethyl ether as a pendant, a redox-responsive disulfide bond, a tertiary amine, and an aromatic moiety on the backbone. In aqueous milieu, these polymers are found to form entropically driven nanoassemblies (ΔS > 0), which were further stabilized by supramolecular cross-linking via the synergistic effect of π–π stacking (aromatic moiety), H-bonding (urethane functionality), and hydrophobic interactions, which eventually amplifies the guest encapsulation stability. A guest release profile in the presence of a redox environment shows ∼65% release in a controlled fashion. Furthermore, the tertiary amine on the polymer backbone leads to the formation of positively charged nanoassemblies at the tumor-relevant pH (pH ∼ 6.5–6.8), which could potentially enhance the cellular uptake of nanocarriers in tumor cells. Thus, a strategy for molar mass controlled polyurethane synthesis, understanding the effect of polymer molar mass on thermodynamics of self-assembly, establishing a stable micellar nanostructure endowed with environment-specific surface charge modulation, and controlled guest release, we believe, will significantly contribute to the development of robust chemotherapeutics.

A light-activated magnetic bead strategy utilized in spatio-temporal controllable exosomes isolation.

Tumor-derived exosomes are considered as a key biomarker in the field of liquid biopsy. However, conventional separation techniques such as ultracentrifugation, co-precipitation and column chromatography cannot isolate samples with high throughput, while traditional immunomagnetic separation techniques, due to steric effect of magnetic beads, reducing sensitivity of exosomes optical detection. Herein, we provide a novel and simple nanoplatform for spatiotemporally controlling extraction and elution of exosomes via magnetic separation and light-activated cargo release. In this system, magnetic beads are co-modified by photoresponsive groups -nitrobenzyl group and aptamers that are compatible with CD63-a highly expressed exosomal surface-specific protein. Through exosomes extracted from cell model and nude mice xenograft tumor model morphological characterization and proteomic analysis, results showed that our novel magnetic bead system outperformed current ultracentrifugation in serum exosome extraction in terms of extraction time, yield, and proportion of populations with high CD63 expression. This strategy may be a powerful tool for exosome isolation in clinical liquid biopsies of cancer disease.

Optical control of neuronal activities with photoswitchable nanovesicles.

Precise modulation of neuronal activity by neuroactive molecules is essential for understanding brain circuits and behavior. However, tools for highly controllable molecular release are lacking. Here, we developed a photoswitchable nanovesicle with azobenzene-containing phosphatidylcholine (azo-PC), coined 'azosome', for neuromodulation. Irradiation with 365 nm light triggers the trans-to-cis isomerization of azo-PC, resulting in a disordered lipid bilayer with decreased thickness and cargo release. Irradiation with 455 nm light induces reverse isomerization and switches the release off. Real-time fluorescence imaging shows controllable and repeatable cargo release within seconds (< 3 s). Importantly, we demonstrate that SKF-81297, a dopamine D1-receptor agonist, can be repeatedly released from the azosome to activate cultures of primary striatal neurons. Azosome shows promise for precise optical control over the molecular release and can be a valuable tool for molecular neuroscience studies.

Targeting necroptosis as an alternative strategy in tumor treatment: From drugs to nanoparticles.

Over last decades, most antitumor therapeutic strategies have focused on apoptosis, however, apoptosis resistance and immunological silence usually led to treatment failure. In this sense, triggering other programmed cell death such as necroptosis may achieve a better therapeutic efficacy and has gained widespread attentions in tumor therapy. Studies in this field have identified several types of necroptosis modulators and highlighted the therapeutic potential of necroptotic cell death in cancer. Nanoparticles further provide possibilities to improve therapeutic outcomes as an efficient drug delivery system, facilitating tumor targeting and controlled cargo release. Furthermore, some nanoparticles themselves can trigger/promote programmed necrosis through hyperthermia, ultrasound and autophagy blockage. These investigations have entered necroptosis for consideration as a promising strategy for tumor therapy, though numerous challenges remain and clinical applications are still distant. In this review, we would briefly introduce molecular mechanism and characteristics of necroptosis, and then summarize recent progress of programmed necrosis and their inducers in tumor therapy. Furthermore, the antitumor strategies that take advantages of nanoparticles to induce necroptosis are also discussed.

MRNA lipid nanoparticle phase transition

Crucial for mRNA-based vaccines are the composition, structure, and properties of lipid nanoparticles (LNPs) as their delivery vehicle. Using all-atom molecular dynamics simulations as a computational microscope, we provide an atomistic view of the structure of the Comirnaty vaccine LNP, its molecular organization, physicochemical properties, and insight in its pH-driven phase transition enabling mRNA release at atomistic resolution. At physiological pH, our simulations suggest an oil-like LNP core that is composed of the aminolipid ALC-0315 and cholesterol (ratio 72:28). It is surrounded by a lipid monolayer formed by distearoylphosphatidylcholine, ALC-0315, PEGylated lipids, and cholesterol at a ratio of 22:9:6:63. Protonated aminolipids enveloping mRNA formed inverted micellar structures that provide a shielding and likely protection from environmental factors. In contrast, at low pH, the Comirnaty lipid composition instead spontaneously formed lipid bilayers that display a high degree of elasticity. These pH-dependent lipid phases suggest that a change in pH of the environment upon LNP transfer to the endosome likely acts as trigger for cargo release from the LNP core by turning aminolipids inside out, thereby destabilizing both the LNP shell and the endosomal membrane.

Designer Liposomic Nanocarriers Are Effective Biofilm Eradicators.

Drug delivery via nanovehicles is successfully employed in several clinical settings, yet bacterial infections, forming microbial communities in the form of biofilms, present a strong challenge to therapeutic treatment due to resistance to conventional antimicrobial therapies. Liposomes can provide a versatile drug-vector strategy for biofilm treatment, but are limited by the need to balance colloidal stability with biofilm penetration. We have discovered a liposomic functionalization strategy, using membrane-embedded moieties of poly[2-(methacryloyloxy)ethyl phosphorylcholine], pMPC, that overcomes this limitation. Such pMPCylation results in liposomic stability equivalent to current functionalization strategies (mostly PEGylation, the present gold-standard), but with strikingly improved cellular uptake and cargo conveyance. Fluorimetry, cryo-electron, and fluorescence microscopies reveal a far-enhanced antibiotic delivery to model Pseudomonas aeruginosa biofilms by pMPC-liposomes, followed by faster cytosolic cargo release, resulting in significantly greater biofilm eradication than either PEGylation or free drug. Moreover, this combination of techniques uncovers the molecular mechanism underlying the enhanced interaction with bacteria, indicating it arises from bridging by divalent ions of the zwitterionic groups on the pMPC moieties to the negatively charged lipopolysaccharide chains emanating from the bacterial membranes. Our results point to pMPCylation as a transformative strategy for liposomal functionalization, leading to next-generation delivery systems for biofilm treatment.

Protein-Delivering Nanocomplexes with Fenton Reaction-Triggered Cargo Release to Boost Cancer Immunotherapy.

Immunotherapeutic efficacy of tumors based on immune checkpoint blockade (ICB) therapy is frequently limited by an immunosuppressive tumor microenvironment and cross-reactivity with normal tissues. Herein, we develop reactive oxygen species (ROS)-responsive nanocomplexes with the function of ROS production for delivery and triggered release of anti-mouse programmed death ligand 1 antibody (αPDL1) and glucose oxidase (GOx). GOx and αPDL1 were complexed with oligomerized (-)-epigallocatechin-3-O-gallate (OEGCG), which was followed by chelation with Fe3+ and coverage of the ROS-responsive block copolymer, POEGMA-b-PTKDOPA, consisting of poly(oligo(ethylene glycol)methacrylate) (POEGMA) and the block with thioketal bond-linked dopamine moieties (PTKDOPA) as the side chains. After intravenous injection, the nanocomplexes show prolonged circulation in the bloodstream with a half-life of 8.72 h and efficient tumor accumulation. At the tumor sites, GOx inside the nanocomplexes can produce H2O2 via oxidation of glucose for Fenton reaction to generate hydroxyl radicals (•OH) which further trigger the release of the protein cargos through ROS-responsive cleavage of thioketal bonds. The released GOx improves the production efficiency of •OH to kill cancer cells for release of tumor-associated antigens via chemodynamic therapy (CDT). The enhanced immunogenic cell death (ICD) can activate the immunosuppressive tumor microenvironment and improve the immunotherapy effect of the released αPDL1, which significantly suppresses primary and metastatic tumors. Thus, the nanocomplexes with Fenton reaction-triggered protein release show great potentials to improve the immunotherapeutic efficacy of ICB via combination with CDT.

Inflammation-triggered dual release of nitroxide radical and growth factor from heparin mimicking hydrogel-tissue composite as cardiovascular implants for anti-coagulation, endothelialization, anti-inflammation, and anti-calcification

Cardiovascular implants made from heterogeneous tissues (HT) often clinically face premature failures such as thrombosis, inflammation, and calcification. Herein, we report a hydrogel-tissue composite exhibiting inflammation instructive release of multiple components towards preventing coagulation, promoting endothelial growth, and modulating reactive oxygen species (ROS) homeostasis. The hydrogel composed of MMP-responsive segment-crosslinked heparin mimicking polymer was loaded with a nitroxide radical via ROS cleavable boronic ester bonds and vascular endothelial growth factor (VEGF) via electrostatic attraction. Matrix metalloproteinase (MMP), which reportedly showed elevated expression in inflammation response to foreign implant degraded the hydrogel and led to the release of heparin mimicking polymer and VEGF, enhancing its anti-coagulation capacity and accelerating the growth of endothelial cells on it. In addition, the composite could sense oxidation biosignal present in the inflammation environment and subsequently release a ROS scavenger for auto-regulation of ROS balance. Subcutaneous implantation in mice suggested that the composite could steer the immune response toward an anti-inflammation state and subcutaneous implantation in rats suggested an anti-calcification effect of it. The enhanced hemocompatibility and endothelialization effects in vivo were further confirmed by the endovascular implantation of tissues via membrane-covered stent delivery. The current findings demonstrate that the incorporation of functional hydrogel into the tissue sophistically exploiting host response for controlled release of multiple active cargos is a feasible approach to boost the anticoagulant, endothelialization, anti-inflammatory, and anti-calcification functions of HT-based cardiovascular implants.

Responsive Supramolecular Devices Assembled from Pillar[5]arene Nanogate and Mesoporous Silica for Cargo Release

Responsive Supramolecular Devices Assembled from Pillar[5]arene Nanogate and Mesoporous Silica for Cargo Release

Diverse cellular strategies for the export of leaderless proteins

<p indent="0mm">Unconventional protein export/secretion (UPE/UPS), in contrast to the classical ER-Golgi-dependent export/secretion of proteins with a leader sequence (signal peptide), employs multiple means to release leaderless cargoes (and in some special cases, cargoes with a leader sequence) to the extracellular space. By far, two major types of UPE have been classified, vesicle-independent UPE and vesicle-dependent UPE. In the former, UPE cargoes can directly translocate across the plasma membrane from the cytoplasm without the assistance of a vesicle carrier. In the latter, UPE cargoes translocate into the lumen of a vesicle which then delivers them out of the cell through membrane trafficking. Both types of UPE require multiple unconventional solutions to complete secretion. Here, we briefly discuss the multiple strategies for a UPE cargo release, focusing on two key steps of leaderless cargoes release in UPE: protein translocation and membrane trafficking.

Nematic Colloidal Micro‐Robots as Physically Intelligent Systems

AbstractPhysically intelligent micro‐robotic systems exploit information embedded in micro‐robots, their colloidal cargo, and their milieu to interact, assemble, and form functional structures. Nonlinear anisotropic fluids such as nematic liquid crystals (NLCs) provide untapped opportunities to embed interactions via their topological defects, complex elastic responses, and ability to dramatically restructure in dynamic settings. Here a four‐armed ferromagnetic micro‐robot is designed and fabricated to embed and dynamically reconfigure information in the nematic director field, generating a suite of physical interactions for cargo manipulation. The micro‐robot shape and surface chemistry are designed to generate a nemato‐elastic energy landscape in the domain that defines multiple modes of emergent, bottom‐up interactions with passive colloids. Micro‐robot rotation expands the ability to sculpt interactions; the energy landscape around a rotating micro‐robot is dynamically reconfigured by complex far‐from‐equilibrium dynamics of the micro‐robot's companion topological defect. These defect dynamics allow transient information to be programmed into the domain and exploited. Robust micro‐robotic manipulation strategies are demonstrated that exploit these diverse modes of nemato‐elastic interaction to achieve cargo docking, transport, release, and assembly of complex reconfigurable structures at multi‐stable sites. Such structures are of great interest to future developments of LC‐based advanced optical device and micro‐manufacturing in anisotropic environments.

Hesperetin Nanoparticle Targeting Neutrophils for Enhanced TBI Therapy

AbstractTraumatic brain injury (TBI) is the main cause of death and disability in people of all ages worldwide. Neuroinflammation plays beneficial and harmful roles in secondary brain injury. Neutrophils play an important role in chemically mediated inflammatory responses through myeloperoxidase (MPO) and inflammation triggered by TBI. Herein, a nanodrug targeting neutrophils and MPO through chemical and biological functions after TBI is designed to enhance the retention and sustained release of drug cargos for improved TBI therapy. 5‐Hydroxytryptamide (5‐HT) is modified on nanoparticles (NPs) loaded with an anti‐inflammatory and antioxidant natural product, hesperetin, to obtain MPO‐ and neutrophil‐targeting NPs, denoted as T‐Hes. In a mouse TBI model, it is confirmed that neutrophil‐targeting NPs can quickly accumulate and remain in the brain tissue, reduce the secretion of inflammatory factors, and the level of microglia and astrocytes, subsequently inducing the transformation of microglia from pro‐inflammatory M1 to anti‐inflammatory M2 cells and promoting the infiltration of regulatory T cells (Tregs). T‐Hes significantly inhibits neuroinflammation and improves neurological deficits through the sustained release of hesperetin in the brain. The findings may open up new avenues for designing clinically translatable probes for TBI treatment.

Gated Organonanoclays for Large Biomolecules: Controlled Release Triggered by Surfactant Stimulus

The low toxicity and high adsorption capacities of clay minerals make them attractive for controlled delivery applications. However, the number of controlled-release studies in the literature using clay minerals is still scarce. In this work, three different clays from the smectite group (Kunipia F, montmorillonite; Sumecton SA, saponite; and Sumecton SWN, hectorite) were successfully loaded with rhodamine B dye and functionalized with oleic acid as a gatekeeper to produce organonanoclays for active and controlled payload-release. Moreover, hematin and cyanocobalamin have also been encapsulated in hectorite gated clay. These organonanoclays were able to confine the entrapped cargos in an aqueous environment, and effectively release them in the presence of surfactants (as bile salts). A controlled delivery of 49 ± 6 μg hematin/mg solid and 32.7 ± 1.5 μg cyanocobalamin/mg solid was reached. The cargo release profiles of all of the organonanoclays were adjusted to three different release-kinetic models, demonstrating the Korsmeyer–Peppas model with release dependence on (i) the organic–inorganic hybrid system, and (ii) the nature of loaded molecules and their interaction with the support. Furthermore, in vitro cell viability assays were carried out with Caco-2 cells, demonstrating that the organonanoclays are well tolerated by cells at particle concentrations of ca. 50 μg/mL.

PEX5 translocation into and out of peroxisomes drives matrix protein import

Peroxisomes are ubiquitous organelles whose dysfunction causes fatal human diseases. Most peroxisomal enzymes are imported from the cytosol by the receptor PEX5, which interacts with a docking complex in the peroxisomal membrane and then returns to the cytosol after monoubiquitination by a membrane-embedded ubiquitin ligase. The mechanism by which PEX5 shuttles between cytosol and peroxisomes and releases cargo inside the lumen is unclear. Here, we use Xenopus egg extract to demonstrate that PEX5 accompanies cargo completely into the lumen, utilizing WxxxF/Y motifs near its N terminus that bind a lumenal domain of the docking complex. PEX5 recycling is initiated by an amphipathic helix that binds to the lumenal side of the ubiquitin ligase. The N terminus then emerges in the cytosol for monoubiquitination. Finally, PEX5 is extracted from the lumen, resulting in the unfolding of the receptor and cargo release. Our results reveal the unique mechanism by which PEX5 ferries proteins into peroxisomes.

Yield‐Stress and Creep Control Depot Formation and Persistence of Injectable Hydrogels Following Subcutaneous Administration

AbstractHydrogels that can be injected into the body using standard needles or catheters enable a minimally invasive strategy to prolong local delivery of therapeutic cargo. In particular, physically cross‐linked hydrogels exhibit shear‐thinning and self‐healing behaviors enabling facile injectability and depot formation upon administration. While prior efforts to characterize these systems have focused on injectability and cargo release behaviors, prediction of cargo release in the body often assumes these materials form a depot rather than spreading out upon administration. Here, it is evaluated how hydrogel rheology correlates with depot formation and persistence following subcutaneous administration in mice with two physiochemically distinct, physically cross‐linked hydrogel systems. Calcium‐alginate and polymer‐nanoparticle hydrogel systems exhibit variable mechanical behaviors across several rheological properties (stiffness, viscoelasticity, yield stress, and creep). By relating measured rheological properties to depot formation and persistence time following subcutaneous administration, it is identified that yield stress is predictive of initial depot formation while creep is predictive of depot persistence for these two gel systems. Indeed, only materials with yield stresses &gt;25 Pa form robust depots, and reduced creep correlates with longer depot persistence. These findings provide predictive insights into design considerations for hydrogel technologies capable of extended controlled release of therapeutic cargo.

Biodegradable particles for protein delivery: Estimation of the release kinetics inside cells.

A methodology to quantify the efficiency of the protein loading and in-vitro delivery for biodegradable capsules with different architectures based on polyelectrolytes (dextran sulfate, poly-L-arginine and polyethylenimine) and SiO2 was developed. The capsules were loaded with model proteins such as ovalbumin and green fluorescent protein (GFP), and the protein release profile inside cells (either macrophages or HeLa cells) after endocytosis was analysed. Both, protein loading and release kinetics were evaluated by analysing confocal laser scanning microscopy images using MatLab and CellProfiler software. Our results indicate that silica capsules showed the most efficient release of proteins as cargo molecules within 48h, as compared to their polymeric counterparts. This developed method for the analysis of the intracellular cargo release kinetics from carrier structures could be used in the future for a better control of drug release profiles.

Intracellular Communication between Synthetic Macromolecules.

Non-viral delivery is an important strategy for selective and efficient gene therapy, immunization, and RNA interference, which overcomes problems of genotoxicity and inherent immunogenicity associated with viral vectors. Liposomes and polymers are compelling candidates as carriers for intracellular, non-viral delivery, but maximal efficiencies of around 1% have been reported for the most advanced non-viral carriers. Here, we develop a library of dendronized bottlebrush polymers with controlled defects, displaying a level of precision surpassed only by biological molecules like DNA, RNA, and proteins. We test concurrent and competitive delivery of DNA and show for the first time that, while intracellular communication is thought to be an exclusively biomolecular phenomenon, such communication between synthetic macromolecular complexes can also take place. Our findings challenge the assumption that delivery agents behave as bystanders that enable transfection by passive intracellular release of genetic cargo and improve upon coarse strategies in intracellular carrier design lacking control over polymer sequence, architecture, and composition, leading to a hit-or-miss outcome. Understanding the communication that takes place between macromolecules will help improve the design of non-viral delivery agents and facilitate translation of genome engineering, vaccines, and nucleic acid-based therapies.

Brain Co-Delivery of Temozolomide and Cisplatin for Combinatorial Glioblastoma Chemotherapy.

Glioblastoma (GBM) is an intractable malignancy with high recurrence and mortality. Combinatorial therapy based on temozolomide (TMZ) and cisplatin (CDDP) shows promising potential for GBM therapy in clinical trials. However, significant challenges include limited blood-brain-barrier (BBB) penetration, poor targeting of GBM tissue/cells, and systemic side effects, which hinder its efficacy in GBM therapy. To surmount these challenges,new GBM-cell membrane camouflaged and pH-sensitive biomimetic nanoparticles (MNPs) inspired by the fact that cancer cells readily pass the BBB and localize with homologous cells, are developed. This study's results show that MNPs can efficiently co-load TMZ and CDDP, transport these across the BBB to specifically target GBM. Incorporation of pH-sensitive polymer then allows for controlled release of drug cargos at GBM sites for combination drug therapy. Mice bearing orthotopic U87MG or drug-resistant U251R GBM tumor and treated with MNPs@TMZ+CDDP show a potent anti-GBM effect, greatly extending the survival time relative to mice receiving single-drug loaded nanoparticles. No obvious side effects are apparent in histological analyses or blood routine studies. Considering these results, the study's new nanoparticle formulation overcomes multiple challenges currently limiting the efficacy of combined TMZ and CDDP GBM drug therapy and appears to be a promising strategy for future GBM combinatorial chemotherapy.

DNA-Liposome Hybrid Carriers for Triggered Cargo Release.

The design of simple and versatile synthetic routes to accomplish triggered-release properties in carriers is of particular interest for drug delivery purposes. In this context, the programmability and adaptability of DNA nanoarchitectures in combination with liposomes have great potential to render biocompatible hybrid carriers for triggered cargo release. We present an approach to form a DNA mesh on large unilamellar liposomes incorporating a stimuli-responsive DNA building block. Upon incubation with a single-stranded DNA trigger sequence, a hairpin closes, and the DNA building block is allowed to self-contract. We demonstrate the actuation of this building block by single-molecule Förster resonance energy transfer (FRET), fluorescence recovery after photobleaching, and fluorescence quenching measurements. By triggering this process, we demonstrate the elevated release of the dye calcein from the DNA–liposome hybrid carriers. Interestingly, the incubation of the doxorubicin-laden active hybrid carrier with HEK293T cells suggests increased cytotoxicity relative to a control carrier without the triggered-release mechanism. In the future, the trigger could be provided by peritumoral nucleic acid sequences and lead to site-selective release of encapsulated chemotherapeutics.

Superoxide-Responsive Cargo Release of Mesoporous Silica Nanocontainers with Thioketal Linker

Superoxide-Responsive Cargo Release of Mesoporous Silica Nanocontainers with Thioketal Linker