Nanoparticles Containing Oxidized Cholesterol Deliver mRNA to the Liver Microenvironment at Clinically Relevant Doses

Using mRNA to produce therapeutic proteins is a promising approach to treat genetic diseases. However, systemically delivering mRNA to cell types besides hepatocytes remains challenging. Fast identification of nanoparticle delivery (FIND) is a DNA barcode‐based system designed to measure how over 100 lipid nanoparticles (LNPs) deliver mRNA that functions in the cytoplasm of target cells in a single mouse. By using FIND to quantify how 75 chemically distinct LNPs delivered mRNA to 28 cell types in vivo, it is found that an LNP formulated with oxidized cholesterol and no targeting ligand delivers Cre mRNA, which edits DNA in hepatic endothelial cells and Kupffer cells at 0.05 mg kg^?1. Notably, the LNP targets liver microenvironmental cells fivefold more potently than hepatocytes. The structure of the oxidized cholesterols added to the LNP is systematically varied to show that the position of the oxidative modification may be important; cholesterols modified on the hydrocarbon tail associated with sterol ring D tend to outperform cholesterols modified on sterol ring B. These data suggest that LNPs formulated with modified cholesterols can deliver gene‐editing mRNA to the liver microenvironment at clinically relevant doses.     

Synthesis and Biological Evaluation of Ionizable Lipid Materials for the In Vivo Delivery of Messenger RNA to B Lymphocytes

B lymphocytes regulate several aspects of immunity including antibody production, cytokine secretion, and T‐cell activation; moreover, B cell misregulation is implicated in autoimmune disorders and cancers such as multiple sclerosis and non‐Hodgkin's lymphomas. The delivery of messenger RNA (mRNA) into B cells can be used to modulate and study these biological functions by means of inducing functional protein expression in a dose‐dependent and time‐controlled manner. However, current in vivo mRNA delivery systems fail to transfect B lymphocytes and instead primarily target hepatocytes and dendritic cells. Here, the design, synthesis, and biological evaluation of a lipid nanoparticle (LNP) system that can encapsulate mRNA, navigate to the spleen, transfect B lymphocytes, and induce more than 60 pg of protein expression per million B cells within the spleen is described. Importantly, this LNP induces more than 85% of total protein production in the spleen, despite LNPs being observed transiently in the liver and other organs. These results demonstrate that LNP composition alone can be used to modulate the site of protein induction in vivo, highlighting the critical importance of designing and synthesizing new nanomaterials for nucleic acid delivery.     

Ionizable Amino‐Polyesters Synthesized via Ring Opening Polymerization of Tertiary Amino‐Alcohols for Tissue Selective mRNA Delivery

The utility of messenger RNA (mRNA) as a therapy is gaining a broad interest due to its potential for addressing a wide range of diseases, while effective delivery of mRNA molecules to various tissues still poses a challenge. This study reports on the design and characterization of new ionizable amino‐polyesters (APEs), synthesized via ring opening polymerization (ROP) of lactones with tertiary amino‐alcohols that enable tissue and cell type selective delivery of mRNA. With a diverse library of APEs formulated into lipid nanoparticles (LNP), structure‐activity parameters crucial for efficient transfection are established and APE‐LNPs are identified that can preferentially home to and elicit effective mRNA expression with low in vivo toxicity in lung endothelium, liver hepatocytes, and splenic antigen presenting cells, including APE‐LNP demonstrating nearly tenfold more potent systemic mRNA delivery to the lungs than vivo‐jetPEI. Adopting tertiary amino‐alcohols to initiate ROP of lactones allows to control polymer molecular weight and obtain amino‐polyesters with narrow molecular weight distribution, exhibiting batch‐to‐batch consistency. All of which highlight the potential for clinical translation of APEs for systemic mRNA delivery and demonstrate the importance of employing controlled polymerization in the design of new polymeric nanomaterials to improve in vivo nucleic acid delivery.     

Constrained Nanoparticles Deliver siRNA and sgRNA to T Cells In Vivo without Targeting Ligands

T cells help regulate immunity, which makes them an important target for RNA therapies. While nanoparticles carrying RNA have been directed to T cells in vivo using protein‐ and aptamer‐based targeting ligands, systemic delivery to T cells without targeting ligands remains challenging. Given that T cells endocytose lipoprotein particles and enveloped viruses, two natural systems with structures that can be similar to lipid nanoparticles (LNPs), it is hypothesized that LNPs devoid of targeting ligands can deliver RNA to T cells in vivo. To test this hypothesis, the delivery of siRNA to 9 cell types in vivo by 168 nanoparticles using a novel siGFP‐based barcoding system and bioinformatics is quantified. It is found that nanomaterials containing conformationally constrained lipids form stable LNPs, herein named constrained lipid nanoparticles (cLNPs). cLNPs deliver siRNA and sgRNA to T cells at doses as low as 0.5 mg kg^?1 and, unlike previously reported LNPs, do not preferentially target hepatocytes. Delivery occurs via a chemical composition‐dependent, size‐independent mechanism. These data suggest that the degree to which lipids are constrained alters nanoparticle targeting, and also suggest that natural lipid trafficking pathways can promote T cell delivery, offering an alternative to active targeting approaches.     

Ionizable Lipid with Supramolecular Chemistry Features for RNA Delivery In Vivo

Lipid nanoparticles (LNPs) and ribonucleic acid (RNA) technology are highly versatile tools that can be deployed for diagnostic, prophylactic, and therapeutic applications. In this report, supramolecular chemistry concepts are incorporated into the rational design of a new ionizable lipid, C3-K2-E14, for systemic administration. This lipid incorporates a cone-shaped structure intended to facilitate cell bilayer disruption, and three tertiary amines to improve RNA binding. Additionally, hydroxyl and amide motifs are incorporated to further enhance RNA binding and improve LNP stability. Optimization of messenger RNA (mRNA) and small interfering RNA (siRNA) formulation conditions and lipid ratios produce LNPs with favorable diameter (<150 nm), polydispersity index (<0.15), and RNA encapsulation efficiency (>90%), all of which are preserved after 2 months at 4 or 37 °C storage in ready-to-use liquid form. The lipid and formulated LNPs are well-tolerated in animals and show no deleterious material-induced effects. Furthermore, 1 week after intravenous LNP administration, fluorescent signal from tagged RNA payloads are not detected. To demonstrate the long-term treatment potential for chronic diseases, repeated dosing of C3-K2-E14 LNPs containing siRNA that silences the colony stimulating factor-1 (CSF-1) gene can modulate leukocyte populations in vivo, further highlighting utility.     

Polydispersity characterization of lipid nanoparticles for siRNA delivery using multiple detection size-exclusion chromatography

The development of lipid nanoparticle (LNP) based small interfering RNA (siRNA) therapeutics presents unique pharmaceutical and regulatory challenges. In contrast to small molecule drugs that are highly pure and well-defined, LNP drug products can exhibit heterogeneity in size, composition, surface property, or morphology. The potential for batch heterogeneity introduces a complexity that must be confronted in order to successfully develop and ensure quality in LNP pharmaceuticals. Currently, there is a lack of scientific knowledge in the heterogeneity of LNPs as well as high-resolution techniques that permit this evaluation. This article reports a size-exclusion chromatography (SEC) method that permits the high-resolution analysis of LNP size distribution in its native solution condition. When coupled with multiple detection systems including UV-vis, multi-angle light scattering, and refractive index, on-line characterization of the distributions in size, molecular weight, and siRNA cargo loading of LNPs could be achieved. Six LNPs with sizes in the rang of 60-140 nm were evaluated and it was found that the SEC separation is efficient, highly reproducible, and can be broadly applied to a diverse range of LNPs. A comparison between the current SEC method and asymmetric field flow fractionation (FFF) shows that the current method provides similar size distribution results on LNPs compared to FFF. Two representative LNPs with similar bulk properties were evaluated in-depth using the SEC method along with two other sizing techniques-dynamic light scattering and cryo-TEM. Profound differences in batch polydispersity were observed between them. Despite the similarity in the particle assembly process, it was found that one LNP (A) possessed a narrow size and molecular weight distribution while the other (B) was polydisperse. The present results suggest that LNP drug products are highly complex and diverse in nature, and care should be taken in examining and understanding them to ensure quality and consistency. The method developed here can not only serve as a method for understanding LNP product property, permitting control on product quality, but also could serve as a potential manufacturing method for product purification. Understandings obtained in this work can help to facilitate the development of LNPs as a well-defined pharmaceutical product.     

A Combinatorial Library of Lipid Nanoparticles for RNA Delivery to Leukocytes

Lipid nanoparticles (LNPs) are the most advanced nonviral platforms for small interfering RNA (siRNA) delivery that are clinically approved. These LNPs, based on ionizable lipids, are found in the liver and are now gaining much attention in the field of RNA therapeutics. The previous generation of ionizable lipids varies in linker moieties, which greatly influences in vivo gene silencing efficiency. Here novel ionizable amino lipids based on the linker moieties such as hydrazine, hydroxylamine, and ethanolamine are designed and synthesized. These lipids are formulated into LNPs and screened for their efficiency to deliver siRNAs into leukocytes, which are among the hardest to transfect cell types. Two potent lipids based on their in vitro gene silencing efficiencies are also identified. These lipids are further evaluated for their biodistribution profile, efficient gene silencing, liver toxicity, and potential immune activation in mice. A robust gene silencing is also found in primary lymphocytes when one of these lipids is formulated into LNPs with a pan leukocyte selective targeting agent (β₇ integrin). Taken together, these lipids have the potential to open new avenues in delivering RNAs into leukocytes.     

An Iron Oxide Nanocarrier for dsRNA to Target Lymph Nodes and Strongly Activate Cells of the Immune System

The success of nanoparticle‐based therapies will depend in part on accurate delivery to target receptors and organs. There is, therefore, considerable potential in nanoparticles which achieve delivery of the right drug(s) using the right route of administration to the right location at the right time, monitoring the process by non‐invasive molecular imaging. A challenge is harnessing immunotherapy via activation of Toll‐like receptors (TLRs) for the development of vaccines against major infectious diseases and cancer. In immunotherapy, delivery of the vaccine components to lymph nodes (LNs) is essential for effective stimulation of the immune response. Although some promising advances have been made, delivering therapeutics to LNs remains challenging. It is here shown that iron‐oxide nanoparticles can be engineered to combine in a single and small (<50 nm) nanocarrier complementary multimodal imaging features with the immunostimulatory activity of polyinosinic‐polycytidylic acid (poly (I:C)). Whilst the fluorescence properties of the nanocarrier show effective delivery to endosomes and TLR3 in antigen presenting cells, MRI/SPECT imaging reveals effective delivery to LNs. Importantly, in vitro and in vivo studies show that, using this nanocarrier, the immunostimulatory activity of poly (I:C) is greatly enhanced. These nanocarriers have considerable potential for cancer diagnosis and the development of new targeted and programmable immunotherapies.     

Development of “CLAN” Nanomedicine for Nucleic Acid Therapeutics

Nucleic acid‐based macromolecules have paved new avenues for the development of therapeutic interventions against a spectrum of diseases; however, their clinical translation is limited by successful delivery to the target site and cells. Therefore, numerous systems have been developed to overcome delivery challenges to nucleic acids. From the viewpoint of clinical translation, it is highly desirable to develop systems with clinically validated materials and controllability in synthesis. With this in mind, a cationic lipid assisted PEG‐b‐PLA nanoparticle (CLAN) is designed that is capable of protecting nucleic acids via encapsulation inside the aqueous core, and delivers them to target cells, while maintaining or improving nucleic acid function. The system is formulated from clinically validated components (PEG‐b‐PLA and its derivatives) and can be scaled‐up for large scale manufacturing, offering potential for its future use in clinical applications. Here, the development and working mechanisms of CLANs, the ways to improve its delivery efficacy, and its application in various disease treatments are summarized. Finally, a prospective for the further development of CLAN is also discussed.     

Preparation and Characterization of Dentin Phosphophoryn‐Derived Peptide‐Functionalized Lignin Nanoparticles for Enhanced Cellular Uptake

The surface modification of nanoparticles (NPs) using different ligands is a common strategy to increase NP?cell interactions. Here, dentin phosphophoryn‐derived peptide (DSS) lignin nanoparticles (LNPs) are prepared and characterized, the cellular internalization of the DSS‐functionalized LNPs (LNPs‐DSS) into three different cancer cell lines is evaluated, and their efficacy with the widely used iRGD peptide is compared. It is shown that controlled extent of carboxylation of lignin improves the stability at physiological conditions of LNPs formed upon solvent exchange. Functionalization with DSS and iRGD peptides maintains the spherical morphology and moderate polydispersity of LNPs. The LNPs exhibit good cytocompatibility when cultured with PC3‐MM2, MDA‐MB‐231, and A549 in the conventional 2D model and in the 3D cell spheroid morphology. Importantly, the 3D cell models reveal augmented internalization of peptide‐functionalized LNPs and improve antiproliferative effects when the LNPs are loaded with a cytotoxic compound. Overall, LNPs‐DSS show equal or even superior cellular internalization than the LNPs‐iRGD, suggesting that DSS can also be used to enhance the cellular uptake of NPs into different types of cells, and release different cargos intracellularly.     

Assessing cellular toxicities in fibroblasts upon exposure to lipid-based nanoparticles: a high content analysis approach

Lipid-based nanoparticles (LNPs) are widely used for the delivery of drugs and nucleic acids. Although most of them are considered safe, there is confusing evidence in the literature regarding their potential cellular toxicities. Moreover, little is known about the recovery process cells undergo after a cytotoxic insult. We have previously studied the systemic effects of common LNPs with different surface charge (cationic, anionic, neutral) and revealed that positively charged LNPs ((+)LNPs) activate pro-inflammatory cytokines and induce interferon response by acting as an agonist of Toll-like receptor 4 on immune cells. In this study, we focused on the response of human fibroblasts exposed to LNPs and their cellular recovery process. To this end, we used image-based high content analysis (HCA). Using this strategy, we were able to show simultaneously, in several intracellular parameters, that fibroblasts can recover from the cytotoxic effects of (+)LNPs. The use of HCA opens new avenues in understanding cellular response and nanotoxicity and may become a valuable tool for screening safe materials for drug delivery and tissue engineering.     

Low‐Density Lipoprotein‐Mimicking Nanoparticles for Tumor‐Targeted Theranostic Applications

This study introduces multifunctional lipid nanoparticles (LNPs), mimicking the structure and compositions of low‐density lipoproteins, for the tumor‐targeted co‐delivery of anti‐cancer drugs and superparamagnetic nanocrystals. Paclitaxel (4.7 wt%) and iron oxide nanocrystals (6.8 wt%, 11 nm in diameter) are co‐encapsulated within folate‐functionalized LNPs, which contain a cluster of nanocrystals with an overall diameter of about 170 nm and a zeta potential of about ‐40 mV. The folate‐functionalized LNPs enable the targeted detection of MCF‐7, human breast adenocarcinoma expressing folate receptors, in T₂‐weighted magnetic resonance images as well as the efficient intracellular delivery of paclitaxel. Paclitaxel‐free LNPs show no significant cytotoxicity up to 0.2 mg mL^?1, indicating the excellent biocompatibility of the LNPs for intracellular drug delivery applications. The targeted anti‐tumor activities of the LNPs in a mouse tumor model suggest that the low‐density lipoprotein‐mimetic LNPs can be an effective theranostic platform with excellent biocompatibility for the tumor‐targeted co‐delivery of various anti‐cancer agents.     

Traceable Bioinspired Nanoparticle for the Treatment of Metastatic Breast Cancer via NIR‐Trigged Intracellular Delivery of Methylene Blue and Cisplatin

Cytotoxic T lymphocyte (CTL) eliminates abnormal cells through target recognition‐triggered intracellular toxin delivery. Chimeric antigen receptor T‐cell improves cancer cell recognition of CTL, but its effectiveness and safety in solid tumor treatment are still hampered by poor tumor infiltration, suppressive tumor microenvironment, and severe on‐target off‐tumor toxicity. Given the functionality and challenges of CTL in cancer therapy, herein, a CTL‐inspired nanovesicle (MPV) with a cell membrane–derived shell and a methylene blue (MB) and cisplatin (Pt) loaded gelatin nanogel core is created. The MPV generates contrast for tumor photoacoustic imaging, and produces hyperthermia upon laser irradiation, enabling photothermal imaging and deep tumor penetration. Meanwhile, it releases MB and Pt, and then delivers them into the cytosol of cancer cells, which process can be visualized by imaging the recovery of MB‐derived fluorescence. The localized hyperthermia, photodynamic therapy, and chemotherapy together kill 4T1 breast cancer cells effectively, resulting in primary tumor regression and 97% inhibition of pulmonary metastasis, without significant toxicity to the animals. Taken together, the MPV shows tumor‐specific and stimuli‐triggered intracellular toxin delivery with advantages in traceable accumulation and activation, high tumor penetration, and triple combination therapy, and thus can be an effective nanomedicine for combating metastatic breast cancer.     

Neutrophil‐Based Drug Delivery Systems

White blood cells (WBCs) are a major component of immunity in response to pathogen invasion. Neutrophils are the most abundant WBCs in humans, playing a central role in acute inflammation induced by pathogens. Adhesion to vasculature and tissue infiltration of neutrophils are key processes in acute inflammation. Many inflammatory/autoimmune disorders and cancer therapies have been found to be involved in activation and tissue infiltration of neutrophils. A promising strategy to develop novel targeted drug delivery systems is the targeting and exploitation of activated neutrophils. Herein, a new drug delivery platform based on neutrophils is reviewed. There are two types of drug delivery systems: neutrophils as carriers and neutrophil‐membrane‐derived nanovesicles. It is discussed how nanoparticles hijack neutrophils in vivo to deliver therapeutics across blood vessel barriers and how neutrophil‐membrane‐derived nanovesicles target inflamed vasculature. Finally, the potential applications of neutrophil‐based drug delivery systems in treating inflammation and cancers are presented.     

Efficient Delivery of GSDMD-N mRNA by Engineered Extracellular Vesicles Induces Pyroptosis for Enhanced Immunotherapy (Small 20/2023)

Pyroptosis is a newly discovered inflammatory form of programmed cell death, which promotes systemic immune response in cancer immunotherapy. GSDMD is one of the key molecules executing pyroptosis, while therapeutical delivery of GSDMD to tumor cells is of great challenge. In this study, an extracellular vesicles-based GSDMD-N mRNA delivery system (namely EV^Tx) is developed for enhanced cancer immunotherapy, with GSDMD-N mRNA encapsulated inside, Ce6 (Chlorin e6 (Ce6), a hydrophilic sensitizer) incorporated into extracellular vesicular membrane, and HER2 antibody displayed onto the surface. Briefly, GSDMD-N mRNA is translationally repressed in donor cells by optimized puromycin, ensuring the cell viability and facilitating the mRNA encapsulation into extracellular vesicles. When targeted and delivered into HER2⁺ breast cancer cells by the engineered extracellular vesicles, the translational repression is unleashed in the recipient cells as the puromycin is diluted and additionally inactivated by sonodynamic treatment as the extracellular vesicles are armed with Ce6, allowing GSDMD-N translation and pyroptosis induction. In addition, sonodynamic treatment also induces cell death in the recipient cells. In the SKBR3- and HER2 transfected 4T1- inoculated breast tumor mouse models, the engineered EV^Tx efficiently induces a powerful tumor immune response and suppressed tumor growth, providing a nanoplatform for cancer immunotherapy.     

Single Chain Epidermal Growth Factor Receptor Antibody Conjugated Nanoparticles for in vivo Tumor Targeting and Imaging

Epidermal growth factor receptor (EGFR) targeted nanoparticle are developed by conjugating a single‐chain anti‐EGFR antibody (ScFvEGFR) to surface functionalized quantum dots (QDs) or magnetic iron oxide (IO) nanoparticles. The results show that ScFvEGFR can be successfully conjugated to the nanoparticles, resulting in compact ScFvEGFR nanoparticles that specifically bind to and are internalized by EGFR‐expressing cancer cells, thereby producing a fluorescent signal or magnetic resonance imaging (MRI) contrast. In vivo tumor targeting and uptake of the nanoparticles in human cancer cells is demonstrated after systemic delivery of ScFvEGFR‐QDs or ScFvEGFR‐IO nanoparticles into an orthotopic pancreatic cancer model. Therefore, ScFvEGFR nanoparticles have potential to be used as a molecular‐targeted in vivo tumor imaging agent. Efficient internalization of ScFvEGFR nanoparticles into tumor cells after systemic delivery suggests that the EGFR‐targeted nanoparticles can also be used for the targeted delivery of therapeutic agents.     

Rekindling RNAi Therapy: Materials Design Requirements for In Vivo siRNA Delivery

With the recent FDA approval of the first siRNA‐derived therapeutic, RNA interference (RNAi)‐mediated gene therapy is undergoing a transition from research to the clinical space. The primary obstacle to realization of RNAi therapy has been the delivery of oligonucleotide payloads. Therefore, the main aims is to identify and describe key design features needed for nanoscale vehicles to achieve effective delivery of siRNA‐mediated gene silencing agents in vivo. The problem is broken into three elements: 1) protection of siRNA from degradation and clearance; 2) selective homing to target cell types; and 3) cytoplasmic release of the siRNA payload by escaping or bypassing endocytic uptake. The in vitro and in vivo gene silencing efficiency values that have been reported in publications over the past decade are quantitatively summarized by material type (lipid, polymer, metal, mesoporous silica, and porous silicon), and the overall trends in research publication and in clinical translation are discussed to reflect on the direction of the RNAi therapeutics field.     

Role of toll-like receptors 3, 4 and 7 in cellular uptake and response to titanium dioxide nanoparticles

Innate immune response is believed to be among the earliest provisional cellular responses, and mediates the interactions between microbes and cells. Toll-like receptors (TLRs) are critical to these interactions. We hypothesize that TLRs also play an important role in interactions between nanoparticles (NPs) and cells, although little information has been reported concerning such an interaction. In this study, we investigated the role of TLR3, TLR4 and TLR7 in cellular uptake of titanium dioxide NP (TiO₂ NP) agglomerates and the resulting inflammatory responses to these NPs. Our data indicate that TLR4 is involved in the uptake of TiO₂ NPs and promotes the associated inflammatory responses. The data also suggest that TLR3, which has a subcellular location distinct from that of TLR4, inhibits the denaturation of cellular protein caused by TiO₂ NPs. In contrast, the unique cellular localization of TLR7 has middle-ground functional roles in cellular response after TiO₂ NP exposure. These findings are important for understanding the molecular interaction mechanisms between NPs and cells.     

Ceria Nanoparticles Meet Hepatic Ischemia‐Reperfusion Injury: The Perfect Imperfection

The mononuclear phagocyte system (MPS, e.g., liver, spleen) is often treated as a “blackbox” by nanoresearchers in translating nanomedicines. Often, most of the injected nanomaterials are sequestered by the MPS, preventing their delivery to the desired disease areas. Here, this imperfection is exploited by applying nano‐antioxidants with preferential liver uptake to directly prevent hepatic ischemia‐reperfusion injury (IRI), which is a reactive oxygen species (ROS)‐related disease. Ceria nanoparticles (NPs) are selected as a representative nano‐antioxidant and the detailed mechanism of preventing IRI is investigated. It is found that ceria NPs effectively alleviate the clinical symptoms of hepatic IRI by scavenging ROS, inhibiting activation of Kupffer cells and monocyte/macrophage cells. The released pro‐inflammatory cytokines are then significantly reduced and the recruitment and infiltration of neutrophils are minimized, which suppress subsequent inflammatory reaction involved in the liver. The protective effect of nano‐antioxidants against hepatic IRI in living animals and the revealed mechanism herein suggests their future use for the treatment of hepatic IRI in the clinic.     

In vivo Gold Nanoparticle Delivery of Peptide Vaccine Induces Anti‐Tumor Immune Response in Prophylactic and Therapeutic Tumor Models

Gold nanoparticles (AuNPs) are promising vehicles for cancer immunotherapy, with demonstrated efficacy in immune delivery and innate cell stimulation. Nevertheless, their potential has yet to be assessed in the in vivo application of peptide cancer vaccines. In this study, it is hypothesized that the immune distribution and adjuvant qualities of AuNPs could be leveraged to facilitate delivery of the ovalbumin (OVA) peptide antigen and the CpG adjuvant and enhance their therapeutic effect in a B16‐OVA tumor model. AuNP delivery of OVA (AuNP‐OVA) and of CpG (AuNP‐CpG) enhanced the efficacy of both agents and induced strong antigen‐specific responses. In addition, it is found that AuNP‐OVA delivery alone, without CpG, is sufficient to promote significant antigen‐specific responses, leading to subsequent anti‐tumor activity and prolonged survival in both prophylactic and therapeutic in vivo tumor models. This enhanced therapeutic efficacy is likely due to the adjuvant effect of peptide coated AuNPs, as they induce inflammatory cytokine release when cultured with bone marrow dendritic cells. Overall, AuNP‐mediated OVA peptide delivery can produce significant therapeutic benefits without the need of adjuvant, indicating that AuNPs are effective peptide vaccine carriers with the potential to permit the use of lower and safer adjuvant doses during vaccination.