Mesangial cells-mediated glomerulonephritis is a frequent cause of end-stage renal disease. Here, we show that celastrol is effective in treating both reversible and irreversible mesangioproliferative glomerulonephritis in rat models, but find that its off-target distributions cause severe systemic toxicity. We thus target celastrol to mesangial cells using albumin nanoparticles. Celastrol-albumin nanoparticles crosses fenestrated endothelium and accumulates in mesangial cells, alleviating proteinuria, inflammation, glomerular hypercellularity, and excessive extracellular matrix deposition in rat anti-Thy1.1 nephritis models. Celastrol-albumin nanoparticles presents lower drug accumulation than free celastrol in off-target organs and tissues, thereby minimizing celastrol-related systemic toxicity. Celastrol-albumin nanoparticles thus represents a promising treatment option for mesangioproliferative glomerulonephritis and similar glomerular diseases.Mesangial cell-mediated glomerulonephritis is a frequent cause of kidney disease. Here the authors show that celastrol loaded in albumin nanoparticles efficiently targets mesangial cells, and is effective in rat models.

Sustained release and non-parental formulations of peptides and protein drugs are highly desirable because of enhanced therapeutic effects as well as improved patient compliance. This is especially true for small peptides such as thymopentin (TP5). To this end, implantable sandwich poly (hydroxybutyrate-co-hydroxyhexanoate) (PHBHHx) films were designed to prolong release time and to inhibit burst release phenomenon of TP5 by a simple volatilization method. In vitro release studies revealed that sandwich films had nearly no burst release. In vivo release time of sandwich films was prolonged to 42 days. Pharmacodynamic evaluation demonstrated that TP5 sandwich films significantly increased survival rates in a rat immunosuppressive model and normalized CD4+/CD8+ values. These results suggest that TP5 released from sandwich films can attenuate cyclophosphamide's immunosuppressive activity, and possibly achieve results comparable to daily TP5 injection therapy. Thus, sandwich PHBHHx films show excellent potential as a sustained, burst-free release system for small molecular weight, hydrophilic peptide drugs.

Transdermal drug delivery systems for rheumatoid arthritis (RA) have been receiving increasing attention as they can potentially overcome drawbacks which exist in traditional oral or injection strategies, including low patient compliance and serious gastrointestinal side effects. However, transdermal delivery of RA drugs especially biological drugs suffers from low drug delivery efficiency due to the robust skin barrier. Herein, we fabricated melittin-loaded hyaluronic acid (HA) microneedles and investigated their capacity for inhibiting RA. We showed that melittin-loaded HA microneedles possessed high mechanical strength for successful delivery of melittin into the skin and effectively inhibited RA progression in adjuvant induced both rodent and murine models, as shown by results in histological, paw swelling and arthritis score. Furthermore, after modifying HA with cross-linkable groups, the fabricated microneedles with sustained release properties could further improve the therapeutic potency. Cytokine and T cell analysis in the paws and lymphatic organs indicated that the application of microneedles suppressed the levels of pro-inflammation cytokines including IL-17 and TNF-α, and increased the percentage of regulatory CD4 T cells. Our study revealed that polymeric microneedle-mediated transdermal delivery of melittin could serve as a new therapy with high compliance and good therapeutic efficacy for RA and other autoimmune diseases.

Poor distribution of antigen/adjuvant to target sites and inadequate induction of T cell responses remain major challenges in cancer immunotherapy because of the lack of appropriate delivery systems. Nanocarrier-based antigen delivery systems have emerged as an innovative strategy to improve vaccine efficacy. Here we present polymeric hybrid micelles (PHMs) as a simple and potent antigen/adjuvant co-delivery system with highly tunable properties. PHMs consist of two amphiphilic diblock copolymers, polycaprolactone-polyethylenimine (PCL-PEI) and polycaprolactone-polyethyleneglycol (PCL-PEG). PHMs with different proportions of cationic PCL-PEI were prepared and loaded with tyrosinase-related protein 2 (Trp2) peptide and adjuvant CpG oligodeoxynucleotide to generate the Trp2/PHM/CpG co-delivery system. Lymphatic and intracellular antigen delivery as a function of PCL-PEI proportion was evaluated in vitro and in vivo. PHMs containing 10% (w/w) PCL-PEI (Trp2/PHM10/CpG) showed the optimal balance of good distribution to lymph nodes, strong immunization effect after subcutaneous administration, and low toxicity to dendritic cells. In a mouse model of B16F10 melanoma, Trp2/PHM10/CpG showed significantly higher antigen-specific cytotoxic T lymphocyte activity and greater anticancer efficacy than Trp2/PHM0/CpG without PCL-PEI or a mixture of free Trp2 and CpG. These results provide new insights into how cationic segments affect the efficiency of antigen delivery by cationic nanocarriers. They also suggest that PHMs can serve as a structurally simple and highly tunable platform for co-delivery of antigen and adjuvant in cancer immunotherapy.

Facing the threat of highly variable virus infection, versatile vaccination systems are urgently needed. Intranasal mRNA vaccination provides a flexible and convenient approach. However, the nasal epithelium remains a major biological barrier to deliver antigens to nasal associated lymphoid tissue (NALT). To address this issue, a potent polymer-based intranasal mRNA vaccination system for HIV-1 treatment was synthesized using cationic cyclodextrin-polyethylenimine 2k conjugate (CP 2k) complexed with anionic mRNA encoding HIV gp120. The delivery vehicle containing CP 2k and mRNA overcame the epithelial barrier by reversibly opening the tight junctions, enhanced the paracellular delivery of mRNA and consequently minimized absorption of toxins in the nasal cavity. Together with the excellent intracellular delivery and prolonged nasal residence time, strong system and mucosal anti-HIV immune responses as well as cytokine productions were achieved with a balanced Th1/Th2/Th17 type. Our study provided the first proof of evidence that cationic polymers can be used as safe and potent intranasal mRNA vaccine carriers to overcome the nasal epithelial barrier. The safe and versatile polymeric delivery system represents a promising vaccination platform for infectious diseases.

Macromolecular prodrugs obtained by covalently conjugating small molecular drugs with polymeric carriers were proven to accomplish controlled and sustained release of the therapeutic agents in vitro and in vivo. Polyethylene glycol (PEG) has been extensively used due to its low toxicity, low immunogenicity and high biocompatibility. However, for linear PEG macromolecules, the number of available hydroxyl groups for drug coupling does not change with the length of polymeric chain, which limits the application of PEG for drug conjugation purposes. To increase the drug loading and prolong the retention time of 5-fluorouracil (5-Fu), a macromolecular prodrug of 5-Fu, 5-fluorouracil-1 acid-PAE derivative (5-FA-PAE) was synthesized and tested for the antitumor activity in vivo.

Due to the critical correlation between inflammation and carcinogenesis, a therapeutic candidate with anti-inflammatory activity may find application in cancer therapy. Here, we report the therapeutic efficacy of celastrol as a promising candidate compound for treatment of pancreatic carcinoma via naïve neutrophil membrane-coated poly(ethylene glycol) methyl ether-block-poly(lactic-co-glycolic acid) (PEG-PLGA) nanoparticles. Neutrophil membrane-coated nanoparticles (NNPs) are well demonstrated to overcome the blood pancreas barrier to achieve pancreas-specific drug delivery in vivo. Using tumor-bearing mice xenograft model, NNPs showed selective accumulations at the tumor site following systemic administration as compared to nanoparticles without neutrophil membrane coating. In both orthotopic and ectopic tumor models, celastrol-loaded NNPs demonstrated greatly enhanced tumor inhibition which significantly prolonged the survival of tumor bearing mice and minimizing liver metastases. Overall, these results suggest that celastrol-loaded NNPs represent a viable and effective treatment option for pancreatic carcinoma.

Glioma, one of the most common brain tumors, remains a challenge worldwide. Due to the specific biological barriers such as blood-brain barrier (BBB), cancer stem cells (CSCs), tumor associated macrophages (TAMs), and vasculogenic mimicry channels (VMs), a novel versatile targeting delivery for anti-glioma is in urgent need. Here, we designed a hyaluronic acid (HA) ion-pairing nanoparticle. Then, these nanoparticles were encapsulated in liposomes, termed as DOX-HA-LPs, which showed near-spherical morphology with an average size of 155.8 nm and uniform distribution (PDI = 0.155). HA was proven to specifically bind to CD44 receptor, which is over-expressed on the surface of tumor cells, other associated cells (such as CSCs and TAMs) and VMs. We systematically investigated anti-glioma efficacy and mechanisms in vivo and in vitro. The strong anti-glioma efficacy could attribute to the accumulation in glioma site and the regulation of tumor microenvironment with depletion of TAMs, inhibition of VMs, and elimination of CSCs.

Acute liver injury (ALI) has an elevated fatality rate due to untimely and ineffective treatment. Although, schisandrin B (SchB) has been extensively used to treat diverse liver diseases, its therapeutic efficacy on ALI was limited due to its high hydrophobicity. Palmitic acid-modified serum albumin (PSA) is not only an effective carrier for hydrophobic drugs, but also has a superb targeting effect via scavenger receptor-A (SR-A) on the M1 macrophages, which are potential therapeutic targets for ALI. Compared with the common macrophage-targeted delivery systems, PSA enables site-specific drug delivery to reduce off-target toxicity. Herein, we prepared SchB-PSA nanoparticles and further assessed their therapeutic effect on ALI. In vitro, compared with human serum albumin encapsulated SchB nanoparticles (SchB-HSA NPs), the SchB-PSA NPs exhibited more potent cytotoxicity on lipopolysaccharide (LPS) stimulated Raw264.7 (LAR) cells, and LAR cells took up PSA NPs 8.79 times more than HSA NPs. As expected, the PSA NPs also accumulated more in the liver. Moreover, SchB-PSA NPs dramatically reduced the activation of NF-κB signaling, and significantly relieved inflammatory response and hepatic necrosis. Notably, the high dose of SchB-PSA NPs improved the survival rate in 72 h of ALI mice to 75%. Hence, SchB-PSA NPs are promising to treat ALI.

An ideal cancer vaccine should contain both strongly immunogenic cancer-specific antigen and potent adjuvant for stimulating robust cellular immunity which are pivotal for clearance of cancer cells. However, most of commercially available adjuvants such as aluminum phosphate gel cannot stimulate robust cellular immune response. In the current study, we reformed microscale aluminum phosphate gel adjuvant into nanoscale and fabricated CpG loaded and B16F10 tumor cell membrane coated aluminum phosphate nanoparticles (APMC). The resultant nano-vaccines showed a size of around 60 nm and a negative surface charge of -40 mV. Tumor cell membrane not only served as tumor antigens but also effectively improved the colloidal dispersion of aluminum phosphate nanoparticles. Subcutaneously injected APMC were efficiently drained to mouse lymph nodes, significantly increased co-uptake of tumor antigen and CpG by lymph node resident antigen presenting cells, promoted maturation of these cells and enhanced lysosomal antigen escape. After immunizing mice, they triggered robust cellular immunity, including potent IFN-γ+CD4+ T cells, IFN-γ+CD8+ T cells, cytotoxic T lymphocytes and cytokine excretion in spleen and lymph node cells. The elicited responses significantly suppressed tumor growth and prolonged survival of mice in both prophylactic and therapeutic melanoma models. This promising vaccine delivery system shows great potential to clinical transformation and can be further developed for personalized cancer vaccines.

Malignant melanoma, a highly dangerous type of skin cancer, is usually resistant to pro-apoptosis agents such as tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) due to low death receptor expression levels. After verifying combination of chemotherapy drug paclitaxel (PTX) and TRAIL could significantly enhance their anti-melanoma effects, we developed a liposomal melanoma target-delivery system with tumor microenvironment responsiveness (TRAIL-[Lip-PTX]C18-TR) to co-deliver TRAIL and PTX. TRAIL is attached to negatively-charged liposome surface while PTX is encapsulated inside, with final surface modification of a stearyl chain (C18) fused pH-sensitive cell-penetrating peptide (TR). Here, C18-TR could specifically binds to melanoma-rich integrin receptors αvβ3 for melanoma targeting, help release TRAIL in low pH microenvironment by reversing the liposomal charge, and facilitate consequent liposome internalization. TRAIL-[Lip-PTX]C18-TR displayed significantly better in vitro half-maximal inhibitory concentration (IC50) than other formulations, and an in vivo tumor inhibition rate of 93.8%. Mechanistic study revealed that this synergistic effect is associated with the upregulation of death receptors DR4/5 by PTX. This co-delivery system significantly improved TRAIL-based therapy against melanoma, and provided a simple platform to co-deliver other drugs/agents for melanoma treatment.

Successful anticancer chemotherapy requires targeting tumors efficiently and further potential to eliminate cancer stem cell (CSC) subpopulations. Since CD44 is present on many types of CSCs, and it binds specially to hyaluronic acid (HA), we tested whether coating solid lipid nanoparticles with hyaluronan (HA-SLNs)would allow targeted delivery of paclitaxel (PTX) to CD44-overexpressing B16F10 melanoma cells. First, we developed a model system based on melanoma stem-like cells for experiments in vitro and in mouse xenografts, and we showed that cells expressing high levels of CD44 (CD44(+)) displayed a strong CSC phenotype while cells expressing low levels of CD44 (CD44(-)) did not. This phenotype included sphere and colony formation, higher proportion of side population cells, expression of CSC-related markers (ALDH, CD133, Oct-4) and tumorigenicity in vivo. Next we showed that administering PTX-loaded HA-SLNs led to efficient intracellular delivery of PTX and induced substantial apoptosis in CD44(+) cells in vitro. In the B16F10-CD44(+) lung metastasis model, PTX-loaded HA-SLNs targeted the tumor-bearing lung tissues well and subsequently exhibited significant antitumor effects with a relative low dose of PTX, which provided significant survival benefit without evidence of adverse events. These findings suggest that the HA-SLNs targeting system shows promise for enhancing cancer therapy.

A rapid, sensitive and selective liquid chromatography-electrospray ionization-tandem mass spectrometric method was developed and validated for the quantification of scopoletin in rat plasma. After the addition of the internal standard xanthotoxin, plasma samples were pretreated by a simple one-step protein precipitation with acetonitrile-methanol (2:1, v/v). Chromatographic separation was achieved on a Diamonsil ODS chromatography column using gradient elution with the mobile phase consisting of acetonitrile and 0.1% formic acid. The determination was performed by positive ion electrospray ionization in multiple reaction monitoring mode. The calibration curve was linear over the concentration range of 5-1000 ng/mL (r = 0.9996). The intra- and inter-day precision (RSD%) was less than 6.1%, and the accuracy (RE%) was from -3.0%-2.5%. This method was successfully applied to the pharmacokinetic research of scopoletin in rats after intravenous (5 mg/kg) or oral (5, 10 and 20 mg/kg) administration. The result showed that oral bioavailability with a dose of 5 mg/kg was 6.62% ± 1.72%, 10 mg/kg, 5.59% ± 1.16%, and 20 mg/kg, 5.65% ± 0.75%.

Due to its safety and efficacy, aluminum hydroxide is used as an immune adjuvant in human vaccines for over 80 years. Being a Th2 stimulator, the classical gel-like adjuvant, however, fails to generate CD8+ T cell responses, which are important for cancer vaccines. Here, aluminum hydroxide is turned from gel into nano-sized vaccine carriers AlO(OH)-polymer nanoparticles (APNs) to promote their lymphatic migration. After actively uptaken via scavenger receptor-A by antigen-presenting cells (APCs) resident in lymph nodes (LNs), APNs destabilize lysosomes resulting in efficient cytosolic delivery and cross-presentation of antigens. It is demonstrated that administration of APNs loaded with ovalbumin (OVA) and CpG led to the codelivery of both cargos into APCs in LNs, leading to their activation and subsequent adaptive immunity. A prime-boost strategy with low doses of OVA (1.5 µg) and CpG (0.45 µg) induces potent CD8+ T cell responses and dramatically prolongs the survival of B16-OVA tumor-bearing mice. More impressively, when using B16F10 lysates instead of OVA as antigen, substantial antitumor effects on B16F10 tumor model are observed by using APN-CpG. These results suggest the great potential of APNs as vaccine carriers that activate CD8+ T cell responses and the bright prospect of aluminum adjuvant in a nanoparticle formulation.

Photothermal therapy (PTT) can be an effective antitumor therapy, but it may not completely eliminate tumor cells, leading to the risk of recurrence or metastasis. Here we describe nanocarriers that allow combination therapy involving PTT and immunotherapy. Nanocarriers are prepared by coating Al2O3 nanoparticles with non-toxic, biodegradable polydopamine, which shows high photothermal efficiency. A near-infrared laser irradiation can kill the majority of tumor tissues, resulting in the release of tumor-associated antigens. The Al2O3 within the nanoparticles, together with CpG, acts as an adjuvant to trigger robust cell-mediated immune responses that can help eliminate the residual tumor cells and reduce the risk of tumor recurrence. Methods: The characteristics and photothermal performance of polydopamine-coated Al2O3 nanoparticles were examined after one-step preparation. Then we studied their internalization, photothermal toxicity and immunostimulatory activity in vitro. For in vivo experiments, these nanocarriers were injected directly into B16F10 melanoma allografts in mice to ensure specific localization. After photothermal irradiation on day 0, mice were subcutaneously injected with CpG adjuvant on day 1, 3 and 5. Tumor volumes and number of living mice were recorded every two days. Moreover, various immune responses induced by our combined therapy were tested for mechanism research. Results: 50% of mice after our combined treatment successfully achieved the goal of tumor eradication, and survived for 120 days, which was the end point of the experiment. Mechanism studies demonstrated the combined therapy efficiently led to dendritic cell maturation, resulting in the secretion of antibodies and cytokines as well as the proliferation of splenocytes and lymphocytes for anti-tumor immunotherapy. Conclusion: Taken together, these results demonstrated the promise of our combined photothermal therapy and immunotherapy for tumor shrinkage, which merited further research.

To date, cancer phototherapy remains as an unsatisfactory method of cancer treatment due to the high probability of cancer recurrence - an effect that is partly driven by tumor-driven immunosuppression. Therefore, we propose inducing adequate immune responses after photo tumor ablation may be critical to achieve a long term therapeutic effect of phototherapy. Here, we engineered the photosensitizer chlorin e6 (Ce6) and the time-honored immunoadjuvant aluminum hydroxide into bovine serum albumin by albumin-based biomineralization as a novel nanosystem (Al-BSA-Ce6 NPs). After intravenous injection, the nanoparticles not only destroyed tumor cells effectively but also protected animals against tumor rechallenge and metastasis by strongly inducing a systemic anti-tumor immune response. Subsequent analysis demonstrated T cells accumulated in lymph nodes and infiltrated the tumor site, elevating levels of immune indicators including serum antibody, cytokine level and higher proportions of cytotoxic T cells and Th1 cells. These protective effects were not observed with commercially available alumina gels, or when the aluminum hydroxide in the nanoparticles was replaced with ferric hydroxide. Therefore, we present Al-BSA-Ce6 NPs as a novel and unique system for alumina adjuvants that serves as an effective approach for cancer therapy.

Rheumatoid arthritis (RA) is a common chronic inflammatory disorder that usually affects joints. It was found that roburic acid (RBA), an ingredient from anti-RA herb Gentiana macrophylla Pall., displayed strong anti-inflammatory activity. However, its medical application is limited by its hydrophobicity, lack of targeting capability and unclear functional mechanism. Here, we constructed a pH responsive dual-target drug delivery system hitchhiking RBA (RBA-NPs) that targeted both CD44 and folate receptors, and investigated its pharmacological mechanism. In rat RA model, the nanocarriers effectively delivered RBA to inflammatory sites and significantly enhanced the therapeutic outcomes compared with free RBA, as well as strongly reducing inflammatory cytokine levels and promoting tissue repair. Following analysis revealed that M1 macrophages in the joints were reprogrammed to M2 phenotype by RBA. Since the balance of pro- and anti-inflammatory macrophages play important roles in maintaining immune homeostasis and preventing excessive inflammation in RA, this reprogramming is likely responsible for the anti-RA effect. Furthermore, we revealed that RBA-NPs drove M1-to-M2 phenotypic switch by down-regulating the glycolysis level via blocking ERK/HIF-1α/GLUT1 pathway. Thus, our work not only developed a targeting delivery system that remarkably improved the anti-RA efficiency of RBA, but also identified a potential molecular target to reversely reprogram macrophages though energy metabolism regulation.

The generation of anti-adenovirus neutralizing antibody (NAb) in humans severely restricts the utilization of recombinant adenovirus serotype 5 (Ad5) vectors in gene therapy for a wide range of clinical trials. To overcome this limitation, we ionically complexed Ad5 with a newly synthesized copolymer, which we called APC, making an adenovirus shielded from NAb.

Despite remarkable progress in the development of both viral and nonviral gene delivery vectors for airway disease treatment, poor gene transfer efficiency to the airway epithelium is a major obstacle in clinical application. To take advantage of the unique features of viral and nonviral vectors, we have developed complexes of adenovirus vector and anionic liposomes (AL-Ad5) by the calcium-induced phase change method. In the current study, based on the fact that there are overexpressed folate receptors on the surface of airway epithelia, we further modified the AL-Ad5 complexes with folate (F-AL-Ad5) to improve the transduction ability of Ad5 in airway epithelia. The transduction efficiencies of the obtained F-AL-Ad5 and AL-Ad5 complexes were assessed in primary-cultured airway epithelia in vitro. Our results indicated that compared with naked adenovirus vector, both AL-Ad5 and F-AL-Ad5 could significantly enhance the gene transduction efficiency of adenovirus vector in primary-cultured airway epithelial cells. Moreover, the enhancement mediated by F-AL-Ad5 was more dramatic than that by AL-Ad5. These results suggested that F-AL-Ad5 may be a useful strategy to deliver therapeutic genes to the airway epithelia and is promising in clinical application.

The development of vaccines has always been an essential task worldwide since vaccines are regarded as powerful weapons in protecting the global population. Although the vast majority of currently authorized human vaccinations are administered intramuscularly or subcutaneously, exploring novel routes of immunization has been a prominent area of study in recent years. This is particularly relevant in the face of pandemic diseases, such as COVID-19, where respiratory immunization offers distinct advantages, such as inducing systemic and mucosal responses to prevent viral infections in both the upper and lower respiratory tracts and also leading to higher patient compliance. However, the development of respiratory vaccines confronts challenges due to the physiological barriers of the respiratory tract, with most of these vaccines still in the research and development stage. In this review, we detail the structure of the respiratory tract and the mechanisms of mucosal immunity, as well as the obstacles to respiratory vaccination. We also examine the considerations necessary in constructing a COVID-19 respiratory vaccine, including the dosage form of the vaccines, potential excipients and mucosal adjuvants, and delivery systems and devices for respiratory vaccines. Finally, we present a comprehensive overview of the COVID-19 respiratory vaccines currently under clinical investigation. We hope this review can provide valuable insights and inspiration for the future development of respiratory vaccinations.