Current tumor vaccines mobilize the immune system to eradicate malignant cells and represent a promising complement to conventional therapies. Nonetheless, their efficacy is constrained by the need for multiple doses, intrinsically low immunogenicity and sub-optimal targeted delivery. Here, we developed a stepwise exfoliated microneedle (MN) patch for immediate and concomitant targeted booster vaccination. The patch comprised a rapidly dissolving outer shell loaded with melittin and a slowly degrading inner core that encapsulated biomineralized nanovaccine (NV) composed of ovalbumin (OVA), CpG and Mn ions. Swift melittin release generated a local inflammatory milieu that recruited and matured dendritic cells (DCs), whereas the gradual liberation of NVs sustained antigen presentation and prolonged immune activation by concurrently engaging Toll-like receptor-9 (TLR9) and the cGAS-STING pathway. In a prophylactic B16-OVA melanoma model, this stepwise exfoliated MN patch reduced dosing frequency, markedly enhanced DCs recruitment, maturation and antigen-specific immunity: treated mice exhibited superior tumor prevention, elevated T-cell responses-including memory subsets-and durable immunological memory. In therapeutic studies, the Melittin+NV MN patch likewise produced strong tumor-suppressive effects. Owing to its programmable release profile, the patch coupled immediate immune activation with sustained protection, thereby reducing dosing frequency, improving patient compliance, substantially elevating overall vaccine efficacy. Therefore, the clinical potential of this strategy was underscored for both tumor prevention and therapy.

In order to overcome the high costs and logistical issues associated with nitric oxide (NO) storage tanks, a portable tankless prototype device has been developed for precise controlled release of inhaled nitric oxide (iNO) gas from a solid NO donor, S-nitroso-N-acetyl penicillamine (SNAP) stored within "coffee pod" style replaceable cartridges. The device utilizes LED light to trigger the release of NO from SNAP, and the concentration of the delivered NO gas is continuously monitored with amperometric sensors and regulated by a feedback control system via modulating the light intensity. The system enables facile adjustment of NO concentrations, allowing for the reliable delivery of low-dose NO (up to 10 ppm, as demonstrated here) across clinically relevant air flow rates, with the prototype specifically designed for air flow up to 4 L/min. Importantly, NO generation is achieved without the need for nitrogen (N₂) as a carrier gas, enabling safe and simple operation using ambient air even in home settings. To ensure the safety of this device, the key factors affecting the formation of the major undesired impurity, nitrogen dioxide (NO), during NO generation were carefully considered and minimized to levels suitable for NO inhalation therapy (<1 ppm). The gas phase NO generated by this device significantly reduced the biofilm density as measured by crystal violet staining and the viable bacterial counts in the mucoid P. aeruginosa biofilm established on a plastic surface in vitro and in sputum isolated from cystic fibrosis (CF) patients ex vivo. With the capacity to maintain 10 ppm NO at up to 4 L/min air flow for over five hours, this portable system demonstrates promise for safe, low-cost, in-home iNO delivery as an adjunctive therapy to reduce biofilm-associated bacterial burden in the lungs of cystic fibrosis patients.

Gout is a chronic inflammatory arthritis caused by the deposition of monosodium urate (MSU) crystals in joints and other areas, and the MSU crystals is a result of the progressive long-term hyperuricemia. Therefore, long-term gout management requires a combination of on-demand urate lowering therapy and anti-inflammatory treatment. And the long-term combined administration urgently requires a simplified and effective administration method. Here, we developed a smart hydrogel microneedle system featuring urate-responsive drug release and rapid bubble separation. This system utilized the rapid bubble separation to retain the hydrogel drug reservoir that simultaneously carried the urate lowering drug uricase and the anti-inflammatory drug colchicine in the skin. In hyperuricemia, the ROS-responsive hydrogel formed by the spontaneous cross-linking of dopamine-modified β-cyclodextrin, polyvinyl alcohol and phenylboric acid linker was promoted for degradation by the HO released from the decomposition of uric acid by uricase, thereby achieving the urate-responsive release. In addition, colchicine loading was increased by forming inclusion complexes with dopamine-modified β-cyclodextrin. Thus, this smart microneedle system retained the drug reservoir with urate-responsive release in the skin through transdermal delivery. It not only dynamically controlled the uric acid concentration and decomposed MSU crystals, but also has a powerful real-time anti-inflammatory effect on gout flares. Accordingly, we believe the smart microneedle system has great potential for long-term gout management.

Osteoarthritis (OA), the most prevalent degenerative joint disorder, is emerging as a significant medical and socioeconomic burden worldwide, representing a major challenge in the field of public health for the coming decades. Although intra-articular drug administration remains a cornerstone of OA treatment, its therapeutic efficacy is substantially constrained by multiple barriers posed by intricate joint anatomy, including rapid joint clearance, poor tissue penetration, non-specific biodistribution, and suboptimal pharmacokinetics. Recently, biomaterials have surfaced as promising platforms for advanced drug delivery in OA management. Through appropriate design strategies, biomaterial-based drug delivery systems (DDSs) can significantly prolong intra-articular retention while enabling spatiotemporal control over drug release with optimized pharmacokinetics. These enhancements help overcome the barriers of conventional intra-articular drug delivery, potentially revolutionizing OA treatment paradigms. Herein, this review first describes the anatomical structure of OA joints and elucidates the associated intra-articular drug delivery barriers arising from cartilage, synovial membrane and synovial fluid characteristics. Subsequently, this review delves into the corresponding cutting-edge biomaterial-based DDSs design strategies to overcome these barriers, including size modulation, targeted delivery, stimulus-responsive systems, and multifunctional platforms. Finally, a comprehensive discussion about the current limitations and future directions toward the design and development of advanced biomaterial-based intra-articular DDSs was sketched out. This review establishes a systematic barrier-strategy matching paradigm for biomaterial-based DDSs development, providing actionable insights to address this urgent global health burden of OA.

Osteoarthritis (OA) is a prevalent degenerative joint disease characterized by the progressive breakdown of cartilage, which lacks the capacity for self-repair. However, the negatively charged nature and compactness of the extracellular matrix present obstacles for drugs administered via intra-articular injection to effectively permeate the cartilage matrix and reach the intended target sites. Despite advancements in drug delivery systems, achieving prolonged drug retention and efficient penetration into cartilage tissue remains a significant challenge. In response, this study presents a novel dual physiological signal-responsive KPP@PLEL nanohydrogel system designed to enhance cartilage repair by targeting bone marrow mesenchymal stem cells (BMSCs) and overcoming cartilage permeability barriers. The KPP@PLEL system uniquely combines a polyamidoamine (PAMAM) dendrimer modified with MMP-13 responsive peptides and kartogenin (KGN), which is encapsulated within a thermosensitive hydrogel, PLEL. This approach overcomes the limitations of previous delivery systems by leveraging both body temperature sensitivity and MMP-13 enzyme responsiveness, which are specifically upregulated in OA-affected tissues. This dual-response mechanism enables the sustained release and enhanced delivery of KGN to the deep cartilage matrix while maintaining extended retention in the joint cavity. In vitro and in vivo studies demonstrated that the KPP@PLEL system has excellent biocompatibility, effectively reduces inflammation, and promotes the differentiation of BMSCs into chondrocytes. Additionally, the system showed superior cartilage penetration and extended retention time compared with those of free KGN or the hydrogel alone. In vivo, significant improvements in cartilage regeneration and substantial reductions in osteoarthritis progression were observed following intra-articular administration. This research offers a promising dual-response nanohydrogel platform that addresses key challenges in osteoarthritis treatment by combining efficient drug delivery, prolonged retention, and potent regenerative effects.

Bioactive peptides have become important ingredients in upmarket cosmetics due to their excellent biocompatibility and skincare effects. However, relatively low skin penetration and instability retard their applications. In this study, a palmitoyl peptide-based "bioactive ingredient reservoir" is established to enhance skin penetration and peptide stability, and the persistent ingredient release is endowed. The experimental results identify the Pal-KVK as superior molecules to form helical ribbons and realize multiple skincare efficacies. Pal-KVK shows aggregate transitions from lamella into double-layer lamellae and final helical superstructures (hydrogel state), driven by combined hydrophobic interaction, electrostatic force, and hydrogen bonding. This kind of self-assembled structures allow for the sustained release of active Pal-KVK ingredients through disassembly. In addition, the Pal-KVK aggregates are embedded within a gelatin matrix to prepare a composite hydrogel mask, implementing long-lasting skincare efficacy. Transdermal evaluation and cellular toxicity assay for Pal-KVK system confirm its excellent skin penetration ability and biocompatibility. Consumer dermatological evaluations have conclusively proven that Pal-KVK/gelatin composite hydrogel mask has excellent anti-wrinkle and moisturizing effects, thus developing an effective and advanced skincare method.

Myocardial ischemia-reperfusion (MI/R) injury remains a significant clinical challenge, primarily attributed to excessive oxidative stress and sterile inflammation resulting from dysregulated activation of the cyclic GMP-AMP synthase-stimulator of interferon genes (cGAS-STING) pathway. Therapeutic strategies that target single pathological factors demonstrate limited efficacy, highlighting a critical need for integrative approaches targeting the complex ischemic microenvironment. Herein, this study develops a therapeutic platform termed "Hydro-guarder", integrating mesenchymal stem cells (MSCs)-mediated delivery of hydrogen sulfide (H₂S) donor 5-(4-Hydroxyphenyl)-3H-1,2-dithiole-3-thione (ADTOH) via ROS-responsive nanoparticles. Specifically, Hydro-guarder utilizes bioorthogonal click chemistry to covalently anchor reactive oxygen species (ROS)-responsive nanoparticles loaded with the advanced H₂S donor, ADTOH, onto MSCs. Upon MSCs homing to infarcted myocardium, elevated ROS enabling the spatiotemporally controlled, on-demand release of H₂S. This targeted release protects MSCs from oxidative damage, enhancing their survival and paracrine function. In turn, revitalized MSCs amplify H₂S-mediated suppression of the cGAS-STING inflammatory cascade. In murine MI/R models, Hydro-guarder significantly preserves mitochondrial integrity, improves cardiac function, and reduces infarct size and fibrosis. Furthermore, Hydro-guarder induced a pronounced immunoregulatory shift, characterized by restoration of cytokine homeostasis and robust polarization of macrophages from the M1 to the M2 phenotype. Mechanistically, we found that Hydro-guarder potently suppressed the cGAS-STING inflammatory cascade. This study provides the first systematic demonstration that co-delivery of H₂S and MSCs via a bioorthogonally engineered platform can synergistically attenuate MI/R-induced cGAS-STING inflammation. The "Hydro-guarder" system thus represents a promising and potentially translatable therapeutic strategy for ischemic cardiovascular diseases.

Radiation-induced jaw injury, characterized by inflammation, bone damage, and infection, is a frequent complication of radiotherapy for head and neck malignancies. However, effective biomaterials designed based on the microenvironmental characteristics of irradiated jaws for alleviating bone regeneration disorders remain lacking. Taurine (Tau) and hypotaurine metabolism are significantly upregulated in irradiated mandibular stromal cells, and orally administered Tau can markedly promote jaw regeneration. In order to increase local taurine accumulation and augment therapeutic outcomes, this study aimed to develop covalent organic frameworks (COFs) loaded with Tau (Tau-COFs) for localized application to irradiated jaws. Notably, Tau-COFs modulated bone homeostasis and downregulated tartrate-resistant acid phosphatase in the jaws (86.73 %, p<0.0001). Additionally, they promoted macrophage recruitment (49.34 %, p = 0.0026) and M2 polarization (725.11 %, p<0.0001) within irradiated tissues. Irradiated bone marrow mesenchymal stromal cells and macrophages exhibited endocytosis but unirradiated cells did not, suggesting its distinct role in post-irradiation bone repair. The results of plate count assays revealed that 50 μg/mL COFs significantly inhibited two specific pathogens of osteoradionecrosis of the jaws, Staphylococcus aureus (79.20 %, p = 0.0008) and Prevotella intermedia (84.27 %, p = 0.0238), demonstrating its antibacterial properties. Tau-COFs at the same concentration exhibited superior antibacterial abilities on S. aureus (92.80 %, p = 0.0004) and P. intermedia (95.63 %, p = 0.0006) than COFs. In vivo experiments further validated the osteogenic effects and biosafety of Tau-COFs. Overall, the findings of this study identify Tau-COFs as a promising biomaterial for improving radiation-induced bone regeneration disorders, offering significant potential for future clinical applications.

Although pyroptosis has emerged as a promising strategy for immunotherapy in triple-negative breast cancer (TNBC), its clinical translation remains hindered by systemic off-target toxicity and pronounced immune exclusion. To address these limitations, we developed a high-throughput double nanosecond pulsed microfluidic electroporation system to generate engineered extracellular vesicle (GDEV). These extracellular vesicles co-encapsulate gasdermin E N-terminal (GSDME-N) mRNA to induce pyroptosis and discoidal domain receptor 1 (DDR1) shRNA to suppress immune exclusion, respectively. Proteomics analysis of clinical TNBC specimens identified CD156 as a TNBC-specific surface biomarker. To achieve tumor-specific delivery and minimize off-target effects, GDEV was further functionalized with anti-CD156 antibody and the GALA pH-sensitive peptide, yielding pTGDEV. In the acidic tumor microenvironment, pTGDEV facilitates the precise release of its therapeutic cargo, simultaneously activating pyroptotic signaling and mitigating immune exclusion. In both orthotopic murine TNBC models and patient-derived xenograft models, pTGDEV significantly inhibited tumor progression and prolonged median survival. Treatment with pTGDEV also reduced the population of myeloid-derived suppressor cells while enhancing the infiltration of dendritic cells, CD4 T cells, and CD8 T cells, effectively reprogramming the tumor microenvironment from immunosuppressive to immunostimulatory. These results demonstrate that pTGDEV offers a potent and targeted approach for TNBC immunotherapy, with strong potential for clinical translation.

Colorectal cancer (CRC) characterized by microsatellite stability (MSS) exhibits a low response rate to immunotherapy, attributed to its inherently poor immunogenicity and the exacerbated immunosuppression. The FOX regimen based on 5-fluorouracil (5-FU) and oxaliplatin (OX), a standard therapy for CRC, paradoxically induces both detrimental upregulation of CD47 and beneficial microsatellite instability (MSI). To capitalize on the beneficial MSI while mitigating the adverse effects of CD47 upregulation, we have developed a CD47 antibody-armored liposome (αL-FOX) loaded with 5-FU and OX. This formulation leverages the chemotherapeutically induced MSI and CD47 upregulation to enhance the immunotherapy efficacy. αCD47 guidance facilitates the positive feedback targeting strategy, selectively delivering αL-FOX to cells that overexpress CD47. The FOX-induced upregulation of CD47 further amplifies this targeted delivery. By blocking the CD47 pathway, αCD47-mediated therapy is expected to reactivate infiltrating immune cells, thereby strengthening innate/adaptive responses. In vivo assays have validated the self-targeting capability of αL-FOX, demonstrating significant tumor suppression, reduced recurrence rates, and extended survival time in metastatic CRC-bearing mice. Notably, these effects are achieved at half the FOX dosage with minimal systemic toxicity. This study highlights the potential of nano-immunotherapeutic drugs that function as chemotherapeutic feedback modulators, offering a promising avenue for heterogeneous and immunotherapy-resistant MSS-CRC.

Intracellular protein therapeutics offer great promise for treating diseases involving intracellular targets, yet their clinical translation remains limited by inefficient delivery systems. Here, we report the development of a simple, versatile, and biocompatible protein delivery platform based on tea polyphenol nanoparticles (TPNs) formed via the oxidative self-polymerization of epigallocatechin gallate (EGCG). Through noncovalent interactions between EGCG and proteins, TPNs enable efficient encapsulation of proteins with diverse molecular weights and isoelectric points under mild conditions. Using urate oxidase (UOx) as a model therapeutic protein, we demonstrate that TPNs protect UOx from enzymatic degradation, preserve its native structure and enzymatic activity, and achieve effective cytosolic delivery via glutathione-triggered release. In disease models of hyperuricemia and gouty arthritis, TPNs/UOx significantly prolonged UOx circulation, lowered serum uric acid levels, reduced joint inflammation, and improved liver and kidney function, with favorable biocompatibility. This work establishes TPNs as a versatile and stimuli-responsive platform for intracellular protein delivery, offering new strategies for the clinical application of protein therapeutics.

Protein therapeutics offer high specificity and potency but face clinical limitations such as poor stability, rapid clearance, and suboptimal patient compliance. To address these challenges, we engineered an elastin-like polypeptide (ELP)-based delivery system by genetically fusing the N-terminal domain of Annexin A1 (ANXA1-N) with ELP to generate ANXA1-N-ELP fusion proteins. These proteins self-assembled into nanoparticles at physiological temperature, preserving ANXA1-N activity while markedly improving its pharmacokinetics. The resulting nanoparticles exhibited homogeneous composition, prolonged circulation, and favorable biodistribution. Both in vitro and in vivo studies further confirmed their potent anti-inflammatory activity. In a murine model of inflammatory bowel disease (IBD), ANXA1-N-ELP treatment effectively alleviated inflammation, restored immune homeostasis, and improved disease outcomes. Importantly, the fully genetically encoded ELP carrier enables precise molecular design, tunable biophysical properties, and controlled biosynthesis. Collectively, this study establishes a versatile protein delivery platform with significant therapeutic potential for IBD and other inflammation-related disorders, advancing the clinical translation of protein-based therapeutics.

Ionic liquids (ILs) have emerged as a transformative platform in drug delivery, overcoming key limitations of conventional systems such as poor drug solubility, low bioavailability, and inadequate targeting. Through rational cation-anion engineering, ILs enable versatile drug loading strategies via ionic/covalent bonding, physical mixing, and nanocarrier encapsulation, while concurrently enhancing biotherapeutic stability, potentiating vaccine immunogenicity, and facilitating transport across critical biological barriers. Recent advances incorporate molecular targeting ligands, pharmacokinetic optimization, and stimuli-responsive mechanisms to achieve spatiotemporally controlled drug release. Growing preclinical and clinical evidence underscores ILs' efficacy in addressing unmet therapeutic needs across diseases such as oncology, neurological disorders, and metabolic diseases. While remarkable progress has been achieved, clinical translation necessitates addressing key challenges in long-term biosafety, scalable manufacturing, and regulatory harmonization. The convergence of ILs technology with artificial intelligence, nanomedicine, and additive manufacturing presents unprecedented opportunities to develop personalized therapeutic platforms. This comprehensive review critically examines the evolution of ILs-based drug delivery systems from fundamental principles to clinical implementation, providing a strategic roadmap for their integration into next-generation precision medicine.

Lipid nanoparticles (LNPs) enable efficient mRNA delivery, yet their potential for ocular gene editing remains largely unexplored. Here, we systematically evaluated three LNP formulations containing distinct ionizable lipids, DLin-MC3-DMA, ALC0315, and SM102, for gene delivery to ocular tissues. Among them, SM102-based LNP encapsulating GFP mRNA (SM102-GFP) exhibited the highest transfection efficiency across three cultured ocular cells in vitro. Following intravitreal injection in mice, SM102-GFP achieved selective and robust expression in the trabecular meshwork (TM) without detectable retinal transfection. GFP expression in TM peaked at one week post-injection, declined by three weeks, and could be effectively re-induced by a second dosing of the same vector. Compared with adeno-associated viral (AAV) and adenoviral (Ad) vectors, SM102-GFP showed superior TM specificity and reduced retinal inflammation. Co-delivery of SpCas9 mRNA and sgRNA via SM102-based LNPs enabled efficient CRISPR-mediated knockout of Matrix Gla Protein (Mgp), a key inhibitor of TM calcification. Mgp knockout induced sustained intraocular pressure elevation and anterior chamber deepening with open angles, recapitulating features of primary open-angle glaucoma. Chronic ocular hypertension further led to Müller gliosis and ganglion cell complex thinning, indicative of progressive retinal stress. These findings establish SM102-based LNPs as a safe and efficient platform for TM-targeted gene editing and glaucoma modeling.

Fibroblast activation protein (FAP)-targeted radioligands have shown superior performance in detecting FAP-expressing cancers. However, the quinoline-based agent Ga-FAPI-46 exhibits only moderate in vivo stability and suboptimal pharmacokinetics. To overcome these limitations, a FAP inhibitor, GP01 was designed by introducing N-methyl-benzenesulfonic acid side chain on FAPI-46 with enhanced binding affinity and improved in vivo stability. Ga-GP01 demonstrated high radiochemical purity, favorable binding energy, and excellent in vitro stability. It specifically bound to FAP on A549-hFAP and U87MG cells, exhibiting greater uptake, enhanced internalization, and slower efflux compared to Ga-FAPI-46. In vivo PET/CT imaging revealed that Ga-GP01 accumulated specifically in FAP-positive tumor models, with improved pharmacokinetic behavior and a higher tumor-to-background ratio than Ga-FAPI-46. Tumor uptake correlated significantly with both ex vivo biodistribution and immunohistochemical FAP expression. Clinical evaluation in 35 patients with solid tumors revealed that Ga-GP01 has optimal pharmacokinetics, persistent target engagement and promising lesion detection, delivering a low effective dose with no adverse events within 6 h post-injection. Furthermore, in a direct comparison with F-FDG, Ga-GP01 detected more lesions and achieved higher uptake and tumor-to-background ratios. These data support Ga-GP01 as a clinically translatable FAP-targeted PET tracer and motivate prospective trials in indications where stromal imaging augments or surpasses metabolic imaging.

Small activating RNA (saR) has garnered increasing attention in the biomedical field due to its unique RNA activation function. However, as a short-sequence, double-stranded oligonucleotide, saR's efficacy heavily relies on the delivery efficiency of the vehicle, as well as the accuracy and reliability of saR loading and dissociation. The tetrahedral framework nucleic acid has been established as an effective carrier for oligonucleotides, but its application in saR delivery remains limited. In this study, p21-saR, which upregulates the p21 gene to induce cellular senescence, was selected as a model saR. p21-saR was loaded onto the tetrahedral framework nucleic acid (t-saR) via DNA/RNA hybrid sticky ends. Upon efficient uptake by SCC-9 cells, t-saR underwent sticky end shearing triggered by intracellular RNase H, resulting in the dynamic release of saR and the screening of its guide strand. Through targeted activation of the p21/Rb signaling pathway, t-saR promoted senescence in SCC-9 cells and inhibited tumor growth in vivo, demonstrating superior therapeutic efficacy compared to the saR monomer. In conclusion, the bioswitchable t-saR effectively achieves RNA activation and holds significant promise for biomedical applications.

The rapidly growing global aging population is facing a rising burden of chronic age-related diseases, largely driven by oxidative stress resulting from the accumulation of reactive oxygen species (ROS). Nanozymes, as superior artificial enzymes with diverse biological functions, offer substantial therapeutic potential in age-related diseases. This review integrates a 'chemical properties-biological functions-delivery strategies-therapeutic applications' paradigm, demonstrating how the intrinsic chemical properties of nanozymes underpin their diverse biological functions, which are then precisely leveraged for specific therapeutic mechanisms to combat age-related diseases. First, a brief discussion of the ROS-mediated pathologies associated with age-related diseases was introduced. Then, the chemical properties and biological functions of nanozymes were comprehensively highlighted. Further, strategies for nanozyme delivery and activation were introduced to enhance their stability, targeted accumulation, and controlled release of nanozymes at pathological sites, followed by the highlight of their applications in the therapy of various age-related issues-from neurodegenerative and cardiovascular diseases to diabetes, musculoskeletal disorders, cancer, and pulmonary diseases. Finally, we also present our perspective on the current challenges and future directions, aiming to fully realize the therapeutic promise of nanozymes for promoting healthy aging.

Myocardial fibrosis (MF) is a terminal pathological process in various cardiovascular diseases, posing a serious threat to human health. Despite the acknowledged advantages of liposomes in drug delivery for the targeted anti-MF therapy, the adverse effects of cholesterol greatly hinder the application of conventional liposomes. In this study, we designed Astragaloside IV derivative LS102 replacing cholesterol to construct a novel liposome delivering ferulic acid (LS-Lipo), aiming for the precise management of MF. Compared with conventional liposomes (CL-Lipo), LS-Lipo displayed smaller particle size, stronger stability, higher biosafety and enhanced intracellular uptake. As expected, LS-Lipo achieved targeted action to the fibrotic hearts and showed longer in vivo retention time compared to CL-Lipo. This phenomenon might be attributed to the decreased adsorption of plasma immune proteins and complement proteins after LS102 embedding. Meanwhile, LS-Lipo alleviated isoproterenol-induced fibrosis and myocardial injury, mainly involving in relieving the endothelial-mesenchymal transition, reducing the expressions of inflammatory factors (IL-17, IL-1β, IL-6), and inhibiting excessive collagen deposition. Collectively, these findings preliminarily indicated that LS-Lipo, with the advantages of safety, simplicity and efficiency, might replace conventional liposomes for MF therapy, showing good potential for further clinical application and transformation.

Necrotizing enterocolitis (NEC) is one of the most critical gastrointestinal emergency in neonates, characterized by high mortality and frequent long-term complications. Probiotic supplementation provides a targeted approach to modulate the gut microbiota and reduce the risk of NEC. However, the oral delivery of probiotics is significantly compromised by their poor viability and colonization capacity in the hostile gastrointestinal environment, for which nanoencapsulation has evolved as a powerful solution. Current single-cell nanoencapsulation approaches face significant challenges, including complex preparation processes and difficulties in achieving uniform and scalable production. In this study, we pioneered the application of flash nanoencapsulation (FNE) technology to develop a universal nanoencapsulation strategy for diverse probiotic strains. This strategy successfully achieved high-efficiency polyelectrolyte-lipid bilayer single-cell nanoencapsulation of Lactobacillus rhamnosus GG (LGG), thereby significantly enhancing the mucus adhesiveness and oral bioavailability. Moreover, in a neonatal mice model of NEC, encapsulated LGG administration alleviated intestinal inflammation through the TLR4/NF-κB signaling pathway, enhanced intestinal barrier function, and modulated gut microbiota composition, demonstrating promising efficacy and clinical potential in NEC prevention. This work establishes a theoretical foundation for the development of FNE-based probiotic nanoencapsulation technology and the application of encapsulated probiotics in the prevention of NEC.

While lipid nanoparticle (LNP)-mRNA vaccines have shown remarkable clinical success, their broader application remains limited by relatively low stability, dose-dependent toxicity, and suboptimal immune activation efficiency. Here, we describe an innovative glutarimide-derived ionizable lipid, MOP-1, that enables an LNP with high delivery efficiency and a favorable safety profile. MOP-1-LNPs exhibited optimal physicochemical properties, including high colloidal stability and favorable acid dissociation characteristics for efficient endosomal escape. Comprehensive safety evaluations revealed that MOP-1-LNPs maintained exceptional biocompatibility, with negligible in vitro cytotoxicity and no evidence of organ damage in vivo even at high doses. When evaluated as an influenza mRNA vaccine platform, MOP-1-LNPs induced robust humoral and cellular immune responses, including high neutralizing antibody production and CD8 T-cell activation. These immunogenic properties translated to excellent protection against a lethal viral challenge, with a 90 % survival rate and near-complete viral clearance. Notably, this potent immune response was achieved while maintaining an optimized cytokine profile that avoids excessive inflammatory responses associated with current LNP platforms. These findings indicate that MOP-1-LNPs could become a transformative mRNA delivery system that successfully addresses key challenges in delivery efficiency, stability, and safety. The unique properties of this platform may enable broader applications in prophylactic and therapeutic vaccine development.

Lipid nanoparticles (LNPs) have transformed the delivery of nucleic acid therapeutics; however, their natural tropism favors the liver resulting in clearance by the reticuloendothelial system, with less than 1 % of the injected dose reaching challenging targets such as brain. Biocompatible ionic liquids (ILs) are tunable materials that can modulate nanoparticle interactions with blood components. Choline trans-2-hexenoate (C2HA) is an IL known to facilitate red blood cell (RBC) hitchhiking of PLGA polymeric nanoparticles and reduces hepatic uptake and therefore enabling transport to distant non-hepatic organs. We wanted to determine if C2HA coatings can show similar RBC hitchhiking effects with LNPs. We previously demonstrated that IL-coated LNPs reduce serum protein binding to LNPs-a key contributor to rapid clearance via the liver. While LNPs coated with choline trans-2-hexenoate at 1:1 and 1:2 cation: anion ratios decreased mouse serum protein binding and improved cellular uptake into brain endothelial cells (BECs) and motor neurons, they did not show hitchhiking behavior. To identify IL formulations capable of this behavior, we screened higher IL cation: anion ratios (1:3 and 1:5) for LNP coating and optimized IL volumes that allowed stable particle diameters. The resulting IL-coated LNPs successfully hitchhiked on both mouse and human RBCs and significantly enhanced uptake in b.End3 mouse BECs, and NSC-34 neuroblastoma cells compared to uncoated LNPs. 1:3 IL-coated LNPs demonstrated the most pronounced improvement in RBC binding. Whole blood pharmacokinetic and biodistribution analyses demonstrated that IL-coating significantly extends the circulation time of LNPs and results in reduced hepatic uptake alongside increased routing to the brain, compared to standard LNPs. Both standard as well as 1:3 IL-coated LNPs were safely-tolerated at the tested doses without organ-function abnormalities, stable leukocyte profiles with only mild, isolated changes in platelet number and two cytokines. These findings reveal that ILs can be leveraged to re-engineer clinically approved LNP platforms to promote RBC hitchhiking behavior and further be developed for drug delivery to challenging extra-hepatic targets such as the brain.