Zeolitic imidazolate frameworks activate endosomal Toll-like receptors and potentiate immunogenicity of SARS-CoV-2 spike protein trimer.

Nanomaterials offer unique opportunities to engineer immunomodulatory activity. In this work, we report the Toll-like receptor agonist activity of a nanoscale adjuvant zeolitic imidazolate framework-8 (ZIF-8). The accumulation of ZIF-8 in endosomes and the pH-responsive release of its subunits enable selective engagement with endosomal Toll-like receptors, minimizing the risk of off-target activation. The intrinsic adjuvant properties of ZIF-8, along with the efficient delivery and biomimetic presentation of a severe acute respiratory syndrome coronavirus 2 spike protein receptor-binding domain trimer, primed rapid humoral and cell-mediated immunity in a dose-sparing manner. Our study offers insights for next-generation adjuvants that can potentially impact future vaccine development.

A microneedle vaccine printer for thermostable COVID-19 mRNA vaccines.

Decentralized manufacture of thermostable mRNA vaccines in a microneedle patch (MNP) format could enhance vaccine access in low-resource communities by eliminating the need for a cold chain and trained healthcare personnel. Here we describe an automated process for printing MNP Coronavirus Disease 2019 (COVID-19) mRNA vaccines in a standalone device. The vaccine ink is composed of lipid nanoparticles loaded with mRNA and a dissolvable polymer blend that was optimized for high bioactivity by screening formulations in vitro. We demonstrate that the resulting MNPs are shelf stable for at least 6 months at room temperature when assessed using a model mRNA construct. Vaccine loading efficiency and microneedle dissolution suggest that efficacious, microgram-scale doses of mRNA encapsulated in lipid nanoparticles could be delivered with a single patch. Immunizations in mice using manually produced MNPs with mRNA encoding severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike protein receptor-binding domain stimulate long-term immune responses similar to those of intramuscular administration.

Recent advances in nano- and micro-scale carrier systems for controlled delivery of vaccines.

Vaccines provide substantial safety against infectious diseases, saving millions of lives each year. The recent COVID-19 pandemic highlighted the importance of vaccination in providing mass-scale immunization against outbreaks. However, the delivery of vaccines imposes a unique set of challenges due to their large molecular size and low room temperature stability. Advanced biomaterials and delivery systems such as nano- and mciro-scale carriers are becoming critical components for successful vaccine development. In this review, we provide an updated overview of recent advances in the development of nano- and micro-scale carriers for controlled delivery of vaccines, focusing on carriers compatible with nucleic acid-based vaccines and therapeutics that emerged amid the recent pandemic. We start by detailing nano-scale delivery systems, focusing on nanoparticles, then move on to microscale systems including hydrogels, microparticles, and 3D printed microneedle patches. Additionally, we delve into emerging methods that move beyond traditional needle-based applications utilizing innovative delivery systems. Future challenges for clinical translation and manufacturing in this rapidly advancing field are also discussed.

Topical application of Lactobacilli successfully eradicates Pseudomonas aeruginosa biofilms and promotes wound healing in chronic wounds.

Chronic wounds are difficult to treat due to the presence of biofilm which prevents wound healing. Pseudomonas aeruginosa is one of the most common pathogens found in chronic wounds and conventional treatment strategies have been ineffective in the eradication of its biofilm, without harming the surrounding healthy tissue at the same time. Here, we introduced an innovative approach applying the probiotic product Bio-K+ (containing three lactobacilli) topically as an antimicrobial and antibiofilm agent. We identified lactic acid as the main active component. While antibiotics and antiseptics such as silver-ions only demonstrated limited efficacy, Bio-K+ was able to completely eradicate mature P. aeruginosa biofilms established in an in-vitro and ex-vivo human skin model. Furthermore, it demonstrated biocompatibility in the co-culture with human dermal fibroblasts and accelerated the migration of fibroblasts in a cell migration assay promoting wound healing. To enhance clinical practicability, we introduced Bio-K+ into the hydrocolloid dressing Aquacel, achieving sustained release of lactic acid and biofilm eradication. This new treatment approach applying probiotics could represent a major improvement in the management of chronic wounds and can be extended in treating other biofilm-associated infections.

A Machine Learning-optimized system for on demand, pulsatile, photo- and chemo-therapeutic treatment using near-infrared responsive MoS 2 -based microparticles in a breast cancer model.

Cancer therapy research is of high interest because of the persistence and mortality of the disease and the side effects of traditional therapeutic methods, while often multimodal treatments are necessary based on the patient's needs. The development of less invasive modalities for recurring treatment cycles is thus of critical significance. Herein, a light-activatable microparticle system was developed for localized, pulsatile delivery of anticancer drugs with simultaneous thermal ablation, by applying controlled ON-OFF thermal cycles using near-infrared laser irradiation. The system is composed of poly(caprolactone) microparticles of 200 µm size with incorporated molybdenum disulfide (MoS 2 ) nanosheets as the photothermal agent and hydrophilic doxorubicin or hydrophobic violacein, as model drugs. Upon irradiation the nanosheets heat up to ≥50 °C leading to polymer matrix melting and release of the drug. MoS 2 nanosheets exhibit high photothermal conversion efficiency and allow for application of low power laser irradiation for the system activation. A Machine Learning algorithm was applied to acquire optimal laser operation conditions; 0.4 W/cm 2 laser power at 808 nm, 3-cycle irradiation, for 3 cumulative minutes. In a mouse subcutaneous model of 4T1 triple-negative breast cancer, 25 microparticles were intratumorally administered and after 3-cycle laser treatment the system conferred synergistic phototherapeutic and chemotherapeutic effect. Our on-demand, pulsatile synergistic treatment resulted in increased median survival up to 40 days post start of treatment compared to untreated mice, with complete eradication of the tumors at the primary site. Such a system could have potential for patients in need of recurring cycles of treatment on subcutaneous tumors.

Enhanced stability and clinical absorption of a form of encapsulated vitamin A for food fortification.

Food fortification is an effective strategy to address vitamin A (VitA) deficiency, which is the leading cause of childhood blindness and drastically increases mortality from severe infections. However, VitA food fortification remains challenging due to significant degradation during storage and cooking. We utilized an FDA-approved, thermostable, and pH-responsive basic methacrylate copolymer (BMC) to encapsulate and stabilize VitA in microparticles (MPs). Encapsulation of VitA in VitA-BMC MPs greatly improved stability during simulated cooking conditions and long-term storage. VitA absorption was nine times greater from cooked MPs than from cooked free VitA in rats. In a randomized controlled cross-over study in healthy premenopausal women, VitA was readily released from MPs after consumption and had a similar absorption profile to free VitA. This VitA encapsulation technology will enable global food fortification strategies toward eliminating VitA deficiency.

Experimental and computational understanding of pulsatile release mechanism from biodegradable core-shell microparticles.

Next-generation therapeutics require advanced drug delivery platforms with precise control over morphology and release kinetics. A recently developed microfabrication technique enables fabrication of a new class of injectable microparticles with a hollow core-shell structure that displays pulsatile release kinetics, providing such capabilities. Here, we study this technology and the resulting core-shell microstructures. We demonstrated that pulsatile release is governed by a sudden increase in porosity of the polymeric matrix, leading to the formation of a porous path connecting the core to the environment. Moreover, the release kinetics within the range studied remained primarily independent of the particle geometry but highly dependent on its composition. A qualitative technique was developed to study the pattern of pH evolution in the particles. A computational model successfully modeled deformations, indicating sudden expansion of the particle before onset of release. Results of this study contribute to the understanding and design of advanced drug delivery systems.

Mild Acidosis-Directed Signal Amplification in Tumor Microenvironment via Spatioselective Recruitment of DNA Amplifiers.

DNA biotechnology offers intriguing opportunities for amplification-based sensitive detection. However, spatiotemporally-controlled manipulation of signal amplification for in situ imaging of the tumor microenvironment remains an outstanding challenge. Here, we demonstrate a DNA-based strategy that can spatial-selectively amplify the acidic signal in the extracellular milieu of the tumor to achieve specific imaging with improved sensitivity. The strategy, termed mild acidosis-targeted amplification (MAT-amp), leverages the specific acidic microenvironment to engineer tumor cells with artificial DNA receptors through a pH (low) insertion peptide, which permits controlled recruitment of fluorescent amplifiers via a hybridization chain reaction. The acidosis-responsive amplification cascade enables significant fluorescence enhancement in tumors with a reduced background signal in normal tissues, leading to improved signal-to-background ratio. These results highlight the utility of MAT-amp for in situ imaging of the microenvironment characterized by pH disequilibrium.

Micromolding of Thermoplastic Polymers for Direct Fabrication of Discrete, Multilayered Microparticles.

Soft lithography provides a convenient and effective method for the fabrication of microdevices with uniform size and shape. However, formation of an embossed, connective film as opposed to discrete features has been an enduring shortcoming associated with soft lithography. Removing this residual layer requires additional postprocessing steps that are often incompatible with organic materials. This limits adaptation and widespread realization of soft lithography for broader applications particularly in drug discovery and drug delivery fields. A novel and versatile approach is demonstrated that enables fabrication of discrete, multilayered, fillable, and harvestable microparticles directly from any thermoplastic polymer, even at very high molecular weights. The approach, isolated microparticle replication via surface-segregating polymer blend mold, utilizes a random copolymer additive, designed with a highly fluorinated segment that, when blended with the mold's matrix, spontaneously orients to the surface conferring an extremely low surface energy and nonwetting properties to the template. The extremely nonwetting properties of the mold are further utilized to load soluble biologics directly into the built-in microwells in a rapid and efficient manner using an innovative screen-printing approach. It is believed that this approach holds promise for fabrication of large-array, 3D, complex microstructures, and is a significant step toward clinical translation of microfabrication technologies.

Vaccine delivery systems toward lymph nodes.

Strategies of improving vaccine targeting ability toward lymph nodes have been attracting considerable interest in recent years, though there are remaining delivery barriers based on the inherent properties of lymphatic systems and limited administration routes of vaccination. Recently, emerging vaccine delivery systems using various materials as carriers are widely developed to achieve efficient lymph node targeting and improve vaccine-triggered adaptive immune response. In this review, to further optimize the vaccine targeting ability for future research, the design principles of lymph node targeting vaccine delivery based on the anatomy of lymph nodes and vaccine administration routes are first summarized. Then different designs of lymph node targeting vaccine delivery systems, including vaccine delivery systems in clinical applications, are carefully surveyed. Also, the challenges and opportunities of current delivery systems for vaccines are concluded in the end.

Stimuli-responsive transdermal microneedle patches.

Microneedle (MN) patches consisting of miniature needles have emerged as a promising tool to perforate the stratum corneum and translocate biomolecules into the dermis in a minimally invasive manner. Stimuli-responsive MN patches represent emerging drug delivery systems that release cargos on-demand as a response to internal or external triggers. In this review, a variety of stimuli-responsive MN patches for controlled drug release are introduced, covering the mechanisms of action toward different indications. Future opportunities and challenges with respect to clinical translation are also discussed.

Modeling, design, and machine learning-based framework for optimal injectability of microparticle-based drug formulations.

Inefficient injection of microparticles through conventional hypodermic needles can impose serious challenges on clinical translation of biopharmaceutical drugs and microparticle-based drug formulations. This study aims to determine the important factors affecting microparticle injectability and establish a predictive framework using computational fluid dynamics, design of experiments, and machine learning. A numerical multiphysics model was developed to examine microparticle flow and needle blockage in a syringe-needle system. Using experimental data, a simple empirical mathematical model was introduced. Results from injection experiments were subsequently incorporated into an artificial neural network to establish a predictive framework for injectability. Last, simulations and experimental results contributed to the design of a syringe that maximizes injectability in vitro and in vivo. The custom injection system enabled a sixfold increase in injectability of large microparticles compared to a commercial syringe. This study highlights the importance of the proposed framework for optimal injection of microparticle-based drugs by parenteral routes.

Engineered PLGA microparticles for long-term, pulsatile release of STING agonist for cancer immunotherapy.

Activation of the stimulator of interferon gene (STING) pathway within the tumor microenvironment has been shown to generate a strong antitumor response. Although local administration of STING agonists has promise for cancer immunotherapy, the dosing regimen needed to achieve efficacy requires frequent intratumoral injections over months. Frequent dosing for cancer treatment is associated with poor patient adherence, with as high as 48% of patients failing to comply. Multiple intratumoral injections also disrupt the tumor microenvironment and vascular networks and therefore increase the risk of metastasis. Here, we developed microfabricated polylactic-co-glycolic acid (PLGA) particles that remain at the site of injection and release encapsulated STING agonist as a programmable sequence of pulses at predetermined time points that mimic multiple injections over days to weeks. A single intratumoral injection of STING agonist-loaded microparticles triggered potent local and systemic antitumor immune responses, inhibited tumor growth, and prolonged survival as effectively as multiple soluble doses, but with reduced metastasis in several mouse tumor models. STING agonist-loaded microparticles improved the response to immune checkpoint blockade therapy and substantially decreased the tumor recurrence rate from 100 to 25% in mouse models of melanoma when administered during surgical resection. In addition, we demonstrated the therapeutic efficacy of STING microparticles on an orthotopic pancreatic cancer model in mice that does not allow multiple intratumoral injections. These findings could directly benefit current STING agonist therapy by decreasing the number of injections, reducing risk of metastasis, and expanding its applicability to hard-to-reach cancers.

Chiral Supraparticles for Controllable Nanomedicine.

Chirality is ubiquitous in nature and hard-wired into every biological system. Despite the prevalence of chirality in biological systems, controlling biomaterial chirality to influence interactions with cells has only recently been explored. Chiral-engineered supraparticles (SPs) that interact differentially with cells and proteins depending on their handedness are presented. SPs coordinated with d-chirality demonstrate greater than threefold enhanced cell membrane penetration in breast, cervical, and multiple myeloma cancer cells. Quartz crystal microbalance with dissipation and isothermal titration calorimetry measurements reveal the mechanism of these chiral-specific interactions. Thermodynamically, d-SPs show more stable adhesion to lipid layers composed of phospholipids and cholesterol compared to l-SPs. In vivo, d-SPs exhibit superior stability and longer biological half-lives likely due to opposite chirality and thus protection from endogenous proteins including proteases. This work shows that incorporating d-chirality into nanosystems enhances uptake by cancer cells and prolonged in vivo stability in circulation, providing support for the importance of chirality in biomaterials. Thus, chiral nanosystems may have the potential to provide a new level of control for drug delivery systems, tumor detection markers, biosensors, and other biomaterial-based devices.

Biocompatible near-infrared quantum dots delivered to the skin by microneedle patches record vaccination.

Accurate medical recordkeeping is a major challenge in many low-resource settings where well-maintained centralized databases do not exist, contributing to 1.5 million vaccine-preventable deaths annually. Here, we present an approach to encode medical history on a patient using the spatial distribution of biocompatible, near-infrared quantum dots (NIR QDs) in the dermis. QDs are invisible to the naked eye yet detectable when exposed to NIR light. QDs with a copper indium selenide core and aluminum-doped zinc sulfide shell were tuned to emit in the NIR spectrum by controlling stoichiometry and shelling time. The formulation showing the greatest resistance to photobleaching after simulated sunlight exposure (5-year equivalence) through pigmented human skin was encapsulated in microparticles for use in vivo. In parallel, microneedle geometry was optimized in silico and validated ex vivo using porcine and synthetic human skin. QD-containing microparticles were then embedded in dissolvable microneedles and administered to rats with or without a vaccine. Longitudinal in vivo imaging using a smartphone adapted to detect NIR light demonstrated that microneedle-delivered QD patterns remained bright and could be accurately identified using a machine learning algorithm 9 months after application. In addition, codelivery with inactivated poliovirus vaccine produced neutralizing antibody titers above the threshold considered protective. These findings suggest that intradermal QDs can be used to reliably encode information and can be delivered with a vaccine, which may be particularly valuable in the developing world and open up new avenues for decentralized data storage and biosensing.

A heat-stable microparticle platform for oral micronutrient delivery.

Micronutrient deficiencies affect up to 2 billion people and are the leading cause of cognitive and physical disorders in the developing world. Food fortification is effective in treating micronutrient deficiencies; however, its global implementation has been limited by technical challenges in maintaining micronutrient stability during cooking and storage. We hypothesized that polymer-based encapsulation could address this and facilitate micronutrient absorption. We identified poly(butylmethacrylate-co-(2-dimethylaminoethyl)methacrylate-co-methylmethacrylate) (1:2:1) (BMC) as a material with proven safety, offering stability in boiling water, rapid dissolution in gastric acid, and the ability to encapsulate distinct micronutrients. We encapsulated 11 micronutrients (iron; iodine; zinc; and vitamins A, B2, niacin, biotin, folic acid, B12, C, and D) and co-encapsulated up to 4 micronutrients. Encapsulation improved micronutrient stability against heat, light, moisture, and oxidation. Rodent studies confirmed rapid micronutrient release in the stomach and intestinal absorption. Bioavailability of iron from microparticles, compared to free iron, was lower in an initial human study. An organotypic human intestinal model revealed that increased iron loading and decreased polymer content would improve absorption. Using process development approaches capable of kilogram-scale synthesis, we increased iron loading more than 30-fold. Scaled batches tested in a follow-up human study exhibited up to 89% relative iron bioavailability compared to free iron. Collectively, these studies describe a broad approach for clinical translation of a heat-stable ingestible micronutrient delivery platform with the potential to improve micronutrient deficiency in the developing world. These approaches could potentially be applied toward clinical translation of other materials, such as natural polymers, for encapsulation and oral delivery of micronutrients.

Nanofibrillar Patches of Commensal Skin Bacteria.

We demonstrate entrapment of the commensal skin bacteria Staphylococcus epidermidis in mats composed of soft nanotubes made by membrane-templated layer-by-layer (LbL) assembly. When cultured in broth, the resulting nanofibrillar patches efficiently delay the escape of bacteria and their planktonic growth, while displaying high steady-state metabolic activity. Additionally, the material properties and metabolic activity can be further tuned by postprocessing the patches with additional polysaccharide LbL layers. These patches offer a promising methodology for the fabrication of bacterial skin dressings for the treatment of skin dysbiosis while preventing adverse effects due to bacterial proliferation.

Biofilm-Inspired Encapsulation of Probiotics for the Treatment of Complex Infections.

The emergence of antimicrobial resistance poses a major challenge to healthcare. Probiotics offer a potential alternative treatment method but are often incompatible with antibiotics themselves, diminishing their overall therapeutic utility. This work uses biofilm-inspired encapsulation of probiotics to confer temporary antibiotic protection and to enable the coadministration of probiotics and antibiotics. Probiotics are encapsulated within alginate, a crucial component of pseudomonas biofilms, based on a simple two-step alginate cross-linking procedure. Following exposure to the antibiotic tobramycin, the growth and metabolic activity of encapsulated probiotics are unaffected by tobramycin, and they show a four-log survival advantage over free probiotics. This results from tobramycin sequestration on the periphery of alginate beads which prevents its diffusion into the core but yet allows probiotic byproducts to diffuse outward. It is demonstrated that this approach using tobramycin combined with encapsulated probiotic has the ability to completely eradicate methicillin-resistant Staphylococcus aureus and Pseudomonas aeruginosa in coculture, the two most widely implicated bacteria in chronic wounds.

Stabilized single-injection inactivated polio vaccine elicits a strong neutralizing immune response.

Vaccination in the developing world is hampered by limited patient access, which prevents individuals from receiving the multiple injections necessary for protective immunity. Here, we developed an injectable microparticle formulation of the inactivated polio vaccine (IPV) that releases multiple pulses of stable antigen over time. To accomplish this, we established an IPV stabilization strategy using cationic polymers for pH modulation to enhance traditional small-molecule-based stabilization methods. We investigated the mechanism of this strategy and showed that it was broadly applicable to all three antigens in IPV. Our lead formulations released two bursts of IPV 1 month apart, mimicking a typical vaccination schedule in the developing world. One injection of the controlled-release formulations elicited a similar or better neutralizing response in rats, considered the correlate of protection in humans, than multiple injections of liquid vaccine. This single-administration vaccine strategy has the potential to improve vaccine coverage in the developing world.