Extra-Large Pore Mesoporous Silica Nanoparticles Enabling Co-Delivery of High Amounts of Protein Antigen and Toll-like Receptor 9 Agonist for Enhanced Cancer Vaccine Efficacy

Cancer vaccine aims to invoke antitumor adaptive immune responses to detect and eliminate tumors. However, the current dendritic cells (DCs)-based cancer vaccines have several limitations that are mostly derived from the <i>ex vivo</i> culture of patient DCs. To circumvent the limitations, direct activation and maturation of host DCs using antigen-carrying materials, without the need for isolation of DCs from patients, are required. In this study, we demonstrate the synthesis of extra-large pore mesoporous silica nanoparticles (XL-MSNs) and their use as a prophylactic cancer vaccine through the delivery of cancer antigen and danger signal to host DCs in the draining lymph nodes. Extra-large pores of approximately 25 nm and additional surface modification of XL-MSNs resulted in significantly higher loading of antigen protein and toll-like receptor 9 (TLR9) agonist compared with conventional small-pore MSNs. <i>In vitro</i> study showed the enhanced activation and antigen presentation of DCs and increased secretion of proinflammatory cytokines. <i>In vivo</i> study demonstrated efficient targeting of XL-MSNs co-delivering antigen and TLR9 agonist to draining lymph nodes, induction of antigen-specific cytotoxic T lymphocytes (CTLs), and suppression of tumor growth after vaccination. Furthermore, significant prevention of tumor growth after tumor rechallenge of the vaccinated tumor-free mice resulted, which was supported by a high level of memory T cells. These findings suggest that mesoporous silica nanoparticles with extra-large pores can be used as an attractive platform for cancer vaccines

Photoresponsive Fluorescent Reduced Graphene Oxide by Spiropyran Conjugated Hyaluronic Acid for in Vivo Imaging and Target Delivery

This present article demonstrates the strategy to prepare photoresponsive reduced graphene oxide with mussel inspired adhesive material dopamine (DN) and photochromic dye spiropyran (SP) conjugated to the backbone of the targeting ligand hyaluronic acid (HA; HA-SP). Graphene oxide (GO) was reduced by prepared HA-SP accepting the advantages of catechol chemistry under mildly alkaline condition enabling to achieve functionalized graphene (rGO/HA-SP) as fluorescent nanoparticles. Due to containing HA, rGO/HA-SP can bind to the CD44 cell receptors. The prepared rGO/HA-SP is able to retain its photochromic features and can be converted to merocyanine (MC) form upon irradiation with UV light (wavelength: 365 nm) displaying purple color. Photochromic behavior of rGO/HA-SP was monitored by UV–vis and fluorescence spectroscopy. In vitro fluorescence behavior, examined by confocal laser scanning microscope (CLSM), of rGO/HA-SP in cancerous A549 cell lines assured that efficient delivery of rGO/HA-SP was gained due to HA as targeting ligand. In this work, we have shown that in vivo fluorescence image of spiropyran is possible by administrating MC form solution of rGO/HA-SP using Balb/C mice as in vivo modal. Accumulation of rGO/HA-SP in tumor tissue from biodistribution analysis strongly supports the specific delivery of prepared graphene to the target destination. The well tuned drug release manner from the surface of rGO/HA-SP strongly recommends the developed material not only as fluorescent probe for diagnosis but also as a drug carrier in drug delivery system

VEGF siRNA Delivery System Using Arginine-Grafted Bioreducible Poly(disulfide amine)

Small interfering RNAs (siRNAs) are able to silence their target genes when they are successfully delivered intact into the cytoplasm. Delivery systems that enhance siRNA localization to the cytoplasm can facilitate gene silencing by siRNA therapeutics. We describe an arginine-conjugated poly(cystaminebisacrylamide-diaminohexane) (poly(CBA-DAH-R)), a bioreducible cationic polymer, as an siRNA carrier for therapeutic gene silencing for cancer. After intracellular uptake of the siRNA/poly(CBA-DAH-R) polyplexes, the reductive environment of the cytoplasm cleaves the disulfide linkages in the polymeric backbone, resulting in decomplexing of the siRNA/poly(CBA-DAH-R) polyplexes and release of siRNA molecules throughout the cytoplasm. The siRNA/poly(CBA-DAH-R) polyplexes, which demonstrate increased membrane permeability with arginine modification, have a similar level of cellular uptake as siRNA/bPEI polyplexes. The VEGF siRNA/poly(CBA-DAH-R) polyplexes, however, inhibit VEGF expression to a greater degree than VEGF siRNA/bPEI in various human cancer cell lines. The improved RNAi activity demonstrated by the VEGF siRNA/poly(CBA-DAH-R) polyplexes is due to enhanced intracellular delivery and effective localization to the cytoplasm of the VEGF siRNAs. These results demonstrate that the VEGF siRNA/poly(CBA-DAH-R) polyplex delivery system may useful for siRNA-based approaches for cancer therapy

A Biodegradation Study of SBA-15 Microparticles in Simulated Body Fluid and <i>in Vivo</i>

Mesoporous silica has received considerable attention as a drug delivery vehicle because of its large surface area and large pore volume for loading drugs and large biomolecules. Recently, mesoporous silica microparticles have shown potential as a three-dimensional vaccine platform for modulating dendritic cells via spontaneous assembly of microparticles in a specific region after subcutaneous injection. For further <i>in vivo</i> applications, the biodegradation behavior of mesoporous silica microparticles must be studied and known. Until now, most biodegradation studies have focused on mesoporous silica nanoparticles (MSNs); here, we report the biodegradation of hexagonally ordered mesoporous silica, SBA-15, with micrometer-sized lengths (∼32 μm with a high aspect ratio). The degradation of SBA-15 microparticles was investigated in simulated body fluid (SBF) and in mice by analyzing the structural change over time. SBA-15 microparticles were found to degrade in SBF and <i>in vivo</i>. The erosion of SBA-15 under biological conditions led to a loss of the hysteresis loop in the nitrogen adsorption/desorption isotherm and fingerprint peaks in small-angle X-ray scattering, specifically indicating a degradation of ordered mesoporous structure. Via comparison to previous results of degradation of MSNs in SBF, SBA-15 microparticles degraded faster than MCM-41 nanoparticles presumably because SBA-15 microparticles have a pore size (∼8 nm) and a pore volume larger than those of MCM-41 mesoporous silica. The surface functional groups, the residual amounts of organic templates, and the hydrothermal treatment during the synthesis could affect the rate of degradation of SBA-15. In <i>in vivo</i> testing, previous studies focused on the evaluation of toxicity of mesoporous silica particles in various organs. In contrast, we studied the change in the physical properties of SBA-15 microparticles depending on the duration after subcutaneous injection. The pristine SBA-15 microparticles injected into mice subcutaneously slowly degraded over time and lost ordered structure after 3 days. These findings represent the possible <i>in vivo</i> use of microsized mesoporous silica for drug delivery or vaccine platform after local injection

In Vitro and In Vivo Tumor Targeted Photothermal Cancer Therapy Using Functionalized Graphene Nanoparticles

Despite the tremendous progress that photothermal therapy (PTT) has recently achieved, it still has a long way to go to gain the effective targeted photothermal ablation of tumor cells. Driven by this need, we describe a new class of targeted photothermal therapeutic agents for cancer cells with pH responsive bioimaging using near-infrared dye (NIR) IR825, conjugated poly­(ethylene glycol)-<i>g</i>-poly­(dimethylaminoethyl methacrylate) (PEG-<i>g</i>-PDMA, PgP), and hyaluronic acid (HA) anchored reduced graphene oxide (rGO) hybrid nanoparticles. The obtained rGO nanoparticles (PgP/HA-rGO) showed pH-dependent fluorescence emission and excellent near-infrared (NIR) irradiation of cancer cells targeted in vitro to provide cytotoxicity. Using intravenously administered PTT agents, the time-dependent in vivo tumor target accumulation was exactly defined, presenting eminent photothermal conversion at 4 and 8 h post-injection, which was demonstrated from the ex vivo biodistribution of tumors. These tumor environment responsive hybrid nanoparticles generated photothermal heat, which caused dominant suppression of tumor growth. The histopathological studies obtained by H&E staining demonstrated complete healing from malignant tumor. In an area of limited successes in cancer therapy, our translation will pave the road to design stimulus environment responsive targeted PTT agents for the safe eradication of devastating cancer

Highly Prolonged Release of the Cancer Vaccine and Immunomodulator via a Two-Layer Biodegradable Microneedle for Prophylactic Treatment of Metastatic Cancer

Simultaneous sustained release of cancer vaccines and immunomodulators may effectively trigger durable immune responses and avoid multiple administrations. Here, we established a biodegradable microneedle (bMN) based on a biodegradable copolymer matrix made of polyethylene glycol (PEG) and poly­(sulfamethazine ester urethane) (PSMEU). This bMN was applied to the skin and slowly degraded in the epidermis/dermis layers. Then, the complexes composed of a positively charged polymer (DA3), cancer DNA vaccine (pOVA), and toll-like receptor 3 agonist poly­(I/C) were synchronously released from the matrix in a pain-free manner. The whole microneedle patch was fabricated with two layers. The basal layer was formed using polyvinyl pyrrolidone/polyvinyl alcohol that could be rapidly dissolved upon applying the microneedle patch to the skin, whereas the microneedle layer was formed by complexes encapsulating biodegradable PEG–PSMEU, which was stuck at the injection site for sustained release of therapeutic agents. According to the results, 10 days is the time for the complexes to be completely released and express specific antigens in antigen-presenting cells in vitro and in vivo. It is noteworthy that this system could successfully elicit cancer-specific humoral immune responses and inhibit metastatic tumors in the lungs after a single shot of immunization

Smart pH-Responsive Nanocube-Controlled Delivery of DNA Vaccine and Chemotherapeutic Drugs for Chemoimmunotherapy

The combination of chemotherapeutic agents with immune stimulating agents for treating degenerative diseases, called chemoimmunotherapy, has emerged as a promising cancer treatment modality. Despite the tremendous potential, chemoimmunotherapy by the combination of drugs and immune stimulators often suffers because of the lack of controlled delivery nanostructures in the microenvironment. To this end, we show that by using pH-responsive smart nanocubes (NCs), cancer cells and tumor-associated immune cells can be precisely targeted with a chemotherapeutic agent (doxorubicin, DOX) and immune stimulating agent (plasmid ovalbumin, pOVA) for enhanced chemoimmunotherapy. The pH-responsive smart NCs protect payloads from nuclease degradation and avoid renal clearance and undergo supersensitive structural change at the extracellular tumor regions that mediate efficient release. Concurrent release of pOVA vaccines encoding tumor-specific antigen laden with polyplexes were loaded on tumor-associated immune cells and produce antigen-specific humoral immune response, whereas DOX enables effective infiltration into the cancer cells and is involved in the eradication of tumor tissues. The amount of anti-OVA IgG1 antibody produced by the intravenous administration of NC formulation was similar to that of free OVA formulation. Importantly, the combined delivery of pDNA and DOX using NCs showed significantly enhanced antitumor efficacy in B16/OVA melanoma tumor xenografts, which remarkably outperforms the monotherapy counterparts. These results suggest that pH-responsive smart NCs laden with pDNA and DOX provide a promising nanostructure for chemoimmunotherapy that simultaneously involves cancer cell killing and stimulates antigen-specific immune response to prevent cancer recurrence

Bioinspired pH- and Temperature-Responsive Injectable Adhesive Hydrogels with Polyplexes Promotes Skin Wound Healing

Despite great potential, the delivery of genetic materials into cells or tissues of interest remains challenging owing to their susceptibility to nuclease degradation, lack of permeability to the cell membrane, and short in vivo half-life, which severely restrict their widespread use in therapeutics. To surmount these shortcomings, we developed a bioinspired in situ-forming pH- and temperature-sensitive injectable hydrogel depot that could control the delivery of DNA-bearing polyplexes for versatile biomedical applications. A series of multiblock copolymer, comprised of water-soluble poly­(ethylene glycol) (PEG) and pH- and temperature-responsive poly­(sulfamethazine ester urethane) (PSMEU), has been synthesized as in situ-forming injectable hydrogelators. The free-flowing PEG–PSMEU copolymer sols at high pH and room temperature (pH 8.5, 23 °C) were transformed to stable gel at the body condition (pH 7.4, 37 °C). Physical and mechanical properties of hydrogels, including their degradation rate and viscosity, are elegantly controlled by varying the composition of urethane ester units. Subcutaneous administration of free-flowing PEG–PSMEU copolymer sols to the dorsal region of Sprague–Dawley rats instantly formed hydrogel depot. The degradation of the hydrogel depot was slow at the beginning and found to be bioresorbable after two months. Cationic protein or DNA-bearing polyplex-loaded PEG–PSMEU copolymer sols formed stable gel and controlled its release over 10 days in vivo. Owing to the presence of urethane linkages, the PEG–PSMEU possesses excellent adhesion strength to wide range of surfaces including glass, plastic, and fresh organs. More importantly, the hydrogels effectively adhered on human skin and peeled easily without eliciting an inflammatory response. Subcutaneous implantation of PEG–PSMEU copolymer sols effectively sealed the ruptured skin, which accelerated the wound healing process as observed by the skin appendage morphogenesis. The bioinspired in situ-forming pH- and temperature-sensitive injectable adhesive hydrogel may provide a promising platform for myriad biomedical applications as controlled delivery vehicle, adhesive, and tissue regeneration

Polymer-Free Side-Patched Gold Nanorods Synthesized via Salt-Assisted Anisotropic Structural Tuning of Silica Shells

The precise and selective modification of the silica shell on the anisotropic structures of gold nanorods (AuNRs) is crucial for advancing their applications in areas such as catalysis, sensing, and directional self-assembly. However, a limitation of existing methods to synthesize patched silica shells on the sides of AuNRs is that they require the introduction of polymeric blocking ligands. Herein, we present a novel method for synthesizing polymer-free, side-silica-patched AuNRs (PF/side-SiO2 AuNRs). Utilizing sodium iodide as the key agent, we achieved a yield of approximately 88%. The underlying mechanism involves the curvature-induced, tip-selective chemisorption of iodide, which reduces the charge attraction between negatively charged silica precursors and the AuNR surface. A comparative study of the localized surface plasmon resonance and surface-enhanced Raman scattering (SERS) properties of PF/side-SiO2 AuNRs with those of conventional silica-coated AuNRs was conducted. The SERS signals of the PF/side-SiO2 AuNRs intensified, whereas interference by the polymeric ligand signals was absent to expose the areas near the hotspots. An efficient technique for synthesizing anisotropic silica-coated AuNRs for various applications is presented herein, and our findings offer mechanistic insights related to the core materials and anisotropic deposition of other oxides

Ginsenoside Re protects against kainate-induced neurotoxicity in mice by attenuating mitochondrial dysfunction through activation of the signal transducers and activators of transcription 3 signaling

It was demonstrated that ginsenosides exert anti-convulsive potentials and interleukin-6 (IL-6) is protective from excitotoxicity induced by kainate (KA), a model of temporal lobe epilepsy. Ginsenosides-mediated mitochondrial recovery is essential for attenuating KA-induced neurotoxicity, however, little is known about the effects of ginsenoside Re (GRe), one of the major ginsenosides. In this study, GRe significantly attenuated KA-induced seizures in mice. KA-induced redox changes were more evident in mitochondrial fraction than in cytosolic fraction in the hippocampus of mice. GRe significantly attenuated KA-induced mitochondrial oxidative stress (i.e. increases in reactive oxygen species, 4-hydroxynonenal, and protein carbonyl) and mitochondrial dysfunction (i.e. the increase in intra-mitochondrial Ca2+ and the decrease in mitochondrial membrane potential). GRe or mitochondrial protectant cyclosporin A restored phospho-signal transducers and activators of transcription 3 (STAT3) and IL-6 levels reduced by KA, and the effects of GRe were reversed by the JAK2 inhibitor AG490 and the mitochondrial toxin 3-nitropropionic acid (3-NP). Thus, we used IL-6 knockout (KO) mice to investigate whether the interaction between STAT3 and IL-6 is involved in the GRe effects. Importantly, KA-induced reduction of manganese superoxide dismutase (SOD-2) levels and neurodegeneration (i.e. astroglial inhibition, microglial activation, and neuronal loss) were more prominent in IL-6 KO than in wild-type (WT) mice. These KA-induced detrimental effects were attenuated by GRe in WT and, unexpectedly, IL-6 KO mice, which were counteracted by AG490 and 3-NP. Our results suggest that GRe attenuates KA-induced neurodegeneration via modulating mitochondrial oxidative burden, mitochondrial dysfunction, and STAT3 signaling in mice.</p