Potent Antigen-specific Immune Responses Research Articles

A bacterial outer membrane vesicle-based click vaccine elicits potent immune response against Staphylococcus aureus in mice.

Staphylococcus aureus infection is a severe public health concern with the growing number of multidrug-resistant strains. S. aureus can circumvent the defense mechanisms of host immunity with the aid of multiple virulence factors. An efficacious multicomponent vaccine targeting diverse immune evasion strategies developed by S. aureus is thus crucial for its infection control. In this study, we exploited the SpyCatcher-SpyTag system to engineer bacterial outer membrane vesicles (OMVs) for the development of a multitargeting S. aureus click vaccine. We decorated OMVs with surface exposed SpyCatcher via a truncated OmpA(a.a 1-155)-SpyCatcher fusion. The engineered OMVs can flexibly bind with various SpyTag-fused S. aureus antigens to generate an OMV-based click vaccine. Compared with antigens mixed with alum adjuvant, the click vaccine simultaneously induced more potent antigen-specific humoral and Th1-based cellular immune response, which afforded protection against S. aureus Newman lethal challenge in a mouse model. Our study provided a flexible and versatile click vaccine strategy with the potential for fighting against emerging S. aureus clinical isolates.

Direct Presentation of Tumor-Associated Antigens to Induce Adaptive Immunity by Personalized Dendritic Cell-Mimicking Nanovaccines.

Dendritic cells (DCs)-based vaccines are an approved method for inducing potent antigen-specific immune responses to eliminate tumor cells. However, this promising strategy still faces challenges such as tumor-associated antigens (TAAs) loading, lymph node homing, qualitycontrol, and other limitations. Here, a personalized DC-mimicking nanovaccine (nanoDC) for stimulation of TAAs-specific T cell populations is developed. The nanoDCs are fabricated using nanoparticles with dendritic structure and membranes from mature bone-marrow-derived cells (BMDCs). Mature BMDCs are stimulated by nanostructures assembled from Escherichia coli and tumor cells to efficiently deliver TAAs and induce BMDCs maturation through the stimulator of interferon genes (STING) pathway. By maintaining co-stimulatory markers, molecules class I (MHC-I) antigen complexes and lymphocyte homing receptors, nanoDCs efficiently migrate to lymph nodes and generate potent antigen-specific T cell responses. Consequently, vaccination with nanoDCs strongly inhibits the tumor growth and metastases formation in vivo. In particular, nanoDCs can also induce memory T cells for long-term protective immunity. This study demonstrates that nanoDCs can trigger adaptive immune protection against tumors for personalized immunotherapy and precision medicine.

Smart combination of aluminum hydroxide and MF59 to induce strong cellular immune responses.

As two of the most widely used adjuvants, aluminum hydroxide and the oil-in-water emulsion MF59 have their intrinsic limitations: classical aluminum gel induces only weak cellular immune responses while MF59 cannot be used as an antigen delivery system due to its poor physical interaction with antigen molecules. Herein, we combined these two adjuvants and constructed a novel nano-vaccine delivery system by inserting aluminum hydroxide into the surface of a modified MF59 nano-emulsion (AlNEs). A model antigen ovalbumin (OVA) and an immune potentiator CpG were adsorbed on the surface of AlNEs (hereinafter AlNEs-OVA-CpG) through a facile mixing step. After subcutaneous injection, AlNEs-OVA-CpG effectively drained to lymph nodes, delivered both cargos into lymph node-resident antigen presenting cells (APCs), and escaped from lysosomes into the cytoplasm, resulting in enhanced antigen cross-presentation. Finally, AlNEs-OVA-CpG induced potent antigen-specific humoral and cellular immune responses, which significantly inhibited tumor growth and prolonged mice survival in a EG7-OVA tumor model. In sum, our results suggested that AlNEs have a great prospect to induce CD8+ T cell responses for subunit antigens.

Aqueous extracts from cultivated Cistanche deserticola Y.C. Ma as polysaccharide adjuvant promote immune responses via facilitating dendritic cell activation

Ethnopharmacological relevanceHerbal polysaccharides have exhibited great immune-enhancing potential. Adjuvants are a key tool for developing efficacious vaccines. In our previous study, a water-soluble polysaccharide extracted from wild Cistanche deserticola Y.C. Ma showed potent immunostimulatory activity. Aim of studyIn this study, the immune profiles and efficacy of aqueous extracts of cultivated Cistanche deserticola Y.C. Ma (AECCD) on ICR mice against ovalbumin (OVA) were investigated. In vitro experiments, the possible DC activation mechanism by AECCD was evaluated. Materials and methodsAECCD were extracted using hot water after which the crude polysaccharides were precipitated by ethanol. Mice were firstly immunized subcutaneously with OVA (10 μg per mouse) alone or OVA (10 μg per mouse) respectively containing different dose of AECCD (200, 400 and 800 μg per mouse) on Days 1 and 14 and the magnitude and kinetics of antibodies and cell-mediated responses were then assessed. ResultsAECCD elicited vigorous and long-term IgG responses with mixed Th1/Th2 responses and up-regulated levels of Th-associated cytokines (CD4+IL-4, CD4+IFN-γ and CD8+IFN-γ). Moreover, AECCD induced the strong cellular immune response characterized by increased splenocyte proliferation as well as the activated T cell response. Notably, AECCD significantly enhanced the maturation of dendritic cells (DCs) and inhibited Tregs. In vitro experiments, Preliminary tests indicated that AECCD induced DC activation by promoting phenotypic maturation, cytokine section and allostimulatory activity. Toll-like receptor 4 (TLR4) was an essential receptor for DCs to directly bind AECCD. The inhibitors of NF-κB decreased the expression levels of CD40, CD80, CD86 and MHC-II and the production of IFN-γ, TNF-α and IL-6 through DCs. ConclusionsFinally, these findings suggested that AECCD could elicit potent and durable antigen specific immune responses through DC activation, which was involved in the regulation of maturation markers and cytokine expression via TLR4-related NF-κB pathway. The study indicates that AECCD is a potential immunomodulator.

Self-amplifying mRNA-Based Vaccine Technology and Its Mode of Action.

Self-amplifying mRNAs derived from the genomes of positive-strand RNA viruses have recently come into focus as a promising technology platform for vaccine development. Non-virally delivered self-amplifying mRNA vaccines have the potential to be highly versatile, potent, streamlined, scalable, and inexpensive. By amplifying their genome and the antigen encoding mRNA in the host cell, the self-amplifying mRNA mimics a viral infection, resulting in sustained levels of the target protein combined with self-adjuvanting innate immune responses, ultimately leading to potent and long-lasting antigen-specific humoral and cellular immune responses. Moreover, in principle, any eukaryotic sequence could be encoded by self-amplifying mRNA without the need to change the manufacturing process, thereby enabling a much faster and flexible research and development timeline than the current vaccines and hence a quicker response to emerging infectious diseases. This chapter highlights the rapid progress made in using non-virally delivered self-amplifying mRNA-based vaccines against infectious diseases in animal models. We provide an overview of the unique attributes of this vaccine approach, summarize the growing body of work defining its mechanism of action, discuss the current challenges and latest advances, and highlight perspectives about the future of this promising technology.

Bio-membrane adhesive poly(choline phosphate l-glutamate)-based nanoparticles as vaccine delivery systems for cancer immunotherapy

Vaccination is a widely used efficient method to prevent or to treat a series of diseases. However, current widely-used vaccines usually could not induce strong cellular immunity, which limits their clinical applications in tumor immunotherapy. Therefore, to design and develop effective vaccine delivery systems to induce strong antigen-specific immune response has attracted more and more attention. In this work, a novel bio-membrane adhesive biodegradable cationic polymer poly(choline phosphate l-glutamate) (pCPGlu) was synthesized and evaluated as a vaccine carrier to deliver model antigen protein ovalbumin (OVA). To prepare the pCPGlu, a biodegradable poly(l-glutamate) was functionalized with a choline phosphate (CP) group which could strongly adsorb to cell membranes and then rapidly enter cells. Then, cationic nanoparticles pCPGlu/OVA were formed via electrostatic attraction between the polycationic pCPGlu and negatively charged OVA. The pCPGlu/OVA nanoparticles were efficiently internalized by immature dendritic cells and escaped from lysosomes into the cytoplasm to induce cross-presentation. As a result, the pCPGlu/OVA nanoparticles could provide adequate initial and long-term antigen exposure to stimulate immune response. After subcutaneously immunized with the pCPGlu/OVA nanoparticles in C57BL/6 mice, the antigen-specific immune responses were investigated. The results indicate that, the pCPGlu/OVA nanoparticles could induce more potent antigen-specific humoral and cellular immune response with significantly higher level of activation of dendritic cells, OVA-specific IgG antibodies, splenocytes proliferation, cytokine secretion, and activation of effector memory T cells. Moreover, subcutaneous administration of the pCPGlu/OVA nanoparticles to the E.G7-OVA tumor-bearing mice significantly increased the amount of cytotoxic T lymphocytes, inhibited the tumor growth, and prolonged the mice survival. Taken together, the cationic cell membrane adhesive pCPGlu could be a safe and potent antigen delivery system for efficient cancer immunotherapy.

MMP-7 derived peptides with MHC class-I binding motifs from canine mammary tumor tissue elicit strong antigen-specific T-cell responses in BALB/c mice.

Matrix Metalloproteinases (MMPs)-induced altered proteolysis of extracellular matrix proteins and basement membrane holds the key for tumor progression and metastasis. Matrix metalloproteinases-7 (Matrilysin), the smallest member of the MMP family also performs quite alike; thus serves as a potential candidate for anti-tumor immunotherapy. Conversely, being an endogenous tumor-associated antigen (TAA), targeting MMP-7 for immunization is challenging. But MMP-7-based xenovaccine can surmount the obstacle of poor immunogenicity and immunological tolerance, often encountered in TAA-based conventional vaccine for anti-tumor immunotherapy. This paves the way for investigating the potential of MMP-7-derived major histocompatibility complex (MHC)-binding peptides to elicit precise epitope-specific T-cell responses towards their possible inclusion in anti-tumor vaccine formulations. Perhaps it also ushers the path of achieving multiple epitope-based broad and universal cellular immunity. In current experiment, an immunoinformatics approach has been employed to identify the putative canine matrix matelloproteinases-7 (cMMP-7)-derived peptides with MHC class-I-binding motifs which can elicit potent antigen-specific immune responses in BALB/c mice. Immunization with the cMMP-7 DNA vaccine induced a strong CD8+ cytotoxic T lymphocytes (CTLs) and Th1- type response, with high level of gamma interferon (IFN-γ) production in BALB/c mice. The two identified putative MHC-I-binding nonameric peptides (Peptide32-40 and Peptide175-183) from cMMP-7 induced significant lymphocyte proliferation along with the production of IFN-γ from CD8+ T-cells in mice immunized with cMMP-7 DNA vaccine. The current observation has depicted the immunogenic potential of the two cMMP-7-derived nonapeptides for their possible exploitation in xenovaccine-mediated anti-tumor immunotherapy in mouse model.

Oral immunization of carps with chitosan–alginate microcapsule containing probiotic expressing spring viremia of carp virus (SVCV) G protein provides effective protection against SVCV infection

Spring viremia of carp (SVC) is highly contagious and lethal disease in cyprinid fish, in particular common carps (Cyprinus carpio), causing numerous economic losses to the aquaculture industry. SVC is presently endemic disease in Europe, America, and several countries in Asia and its causative agent is spring viremia of carp virus (SVCV). In this study, a chitosan-alginate microcapsule probiotic vaccine expressing G protein of SVCV was prepared, and the immunogenicity in carps of orally administrated with the microcapsule probiotic vaccine was evaluated. Our results showed that the microcapsule probiotic vaccine can induce potent antigen-specific immune responses in carps via oral vaccination, and provide effective anti-SVCV protection for carps. Significantly, the microcapsule probiotic vaccine is suitable for mass fish immunization, suggesting a promising vaccine strategy for fish.

Antitumor immunity by small extracellular vesicles collected from activated dendritic cells through effective induction of cellular and humoral immune responses

Dendritic cell-derived small extracellular vesicles (DC-sEVs) are proposed as a novel candidate for tumor antigen-based cancer immunotherapy. In order to improve the DC-sEV-induced antitumor immunity, production of DC-sEVs capable of inducing potent antigen-specific humoral and cellular immune responses is necessary. Here, we collected sEVs from DCs and added ovalbumin (OVA), which was used as model antigen, as well as LPS and IFN-γ, to prepare DC-sEVs with high immune activity. After confirming that the collected sEVs, named activated-DCOVA-sEVs, contained OVA and possessed immunologically relevant components (MHC class I molecule displaying antigen epitopes and co-stimulatory molecules, as well as sEV marker proteins), we found that activated-DCOVA-sEV stimulated macrophages and DCs through Toll-like receptor 4 signaling and boosted innate immunity in the tumor microenvironment. Moreover, activated-DCOVA-sEVs induced potent antigen-specific humoral and cellular immune responses both in vitro and in vivo. Finally, immunization with activated-DCOVA-sEVs exhibited stronger in vivo antitumor effects in tumor-bearing mice induced by inoculation with OVA-expressing tumor cells.

Ex vivo-generated dendritic cell-based vaccines in melanoma: the role of nanoparticulate delivery systems.

Melanoma is a poor immunogenic cancer and many treatment strategies have been used to enhance specific or nonspecific immunity against it. Dendritic cell (DC)-based cancer vaccine is the most effective therapies that have been used so far. Meanwhile, the efficacy of DC-based immunotherapy relies on critical factors relating to DCs such as the state of maturation and proper delivery of antigens. In this regard, the use of nanoparticulate delivery systems for effective delivery of antigen to ex vivo-generated DC-based vaccines that also poses adjuvanticity would be an ideal approach. In this review article, we attempt to summarize the role of different types of nanoparticulate antigen delivery systems used in the development of ex vivo-generated DC-based vaccines against melanoma and describe their adjuvanticity in mediation of DC maturation, cytoplasmic presentation of antigens to MHC class I molecules, which led to potent antigen-specific immune responses. As were represented, cationic liposomes were the most used approach, which suggest its potential applicability as delivery systems for further experiments in combination with either adjuvants or monoclonal antibodies.

Dissolving undercut microneedle arrays for multicomponent cutaneous vaccination

The skin is an attractive tissue target for vaccination, as it is readily accessible and contains a dense population of antigen-presenting and immune-accessory cells. Microneedle arrays (MNAs) are emerging as an effective tool for in situ engineering of the cutaneous microenvironment to enable diverse immunization strategies. Here, we present novel dissolving undercut MNAs and demonstrate their application for effective multicomponent cutaneous vaccination. The MNAs are composed of micron-scale needles featuring pyramidal heads supported by undercut stem regions with filleted bases to ensure successful skin penetration and retention during application. Prior efforts to fabricate dissolving undercut microstructures were limited and required complex and lengthy processing and assembly steps. In the current study, we strategically combine three-dimensional (3D) laser lithography, an emerging micro-additive manufacturing method with unique geometric capabilities and nanoscale resolution, and micromolding with favorable materials. This approach enables reproducible production of dissolving MNAs with undercut microneedles that can be tip-loaded with multiple biocargos, such as antigen (ovalbumin) and adjuvant (Poly(I:C)). The resulting MNAs fulfill the geometric (sharp tips and smooth edges) and mechanical-strength requirements for failure-free penetration of human and murine skin to simultaneously deliver multicomponent (antigen plus adjuvant) vaccines to the same cutaneous microenvironment. Cutaneous vaccination of mice using these MNAs induces more potent antigen-specific cellular and humoral immune responses than those elicited by traditional intramuscular injection. Together, the unique geometric features of these undercut MNAs and the associated manufacturing strategy, which is compatible with diverse drugs and biologics, could enable a broad range of non-cutaneous and cutaneous drug delivery applications, including multicomponent vaccination.

Preparation of self-assembled nanoparticles of ε-polylysine-sodium alginate: A sustained-release carrier for antigen delivery.

Low immunogenicity prohibits the widespread use of subunit vaccine against infectious diseases and cancers. Hence, a new generation of adjuvants and delivery systems is indispensable for more potent antigen-specific immune responses. Predominantly, nanoparticles formulated from biodegradable polymers are being widely explored as carriers of novel vaccines owing to their outstanding natural properties. We fabricated a model antigen - bovine serum albumin (BSA) encapsulated ε-polylysine (ε-PL) - sodium alginate (SA) nanoparticles (PSNPs), which were self-assembled by ionotropic complexation method, a very simple and mild process, as a result of the electrostatic interaction between oppositely charged polyelectrolyte complexes (PEC). After the preparation, various in vitro parameters were characterized. Scanning electron microscope and dynamic light scattering were employed to study the morphology, size, zeta potential and optimize formulation. Forming mechanism of PSNPS was analyzed and verified by infrared absorption spectra and thermal analysis. Delivery behavior of PSNPs was assessed via release study, cytotoxicity measurement and cellular uptake. BSA-PSNPs with a mean particle diameter 133.2 ± 0.5 nm, narrow size distribution and negatively charged surface had been synthesized successfully by this method. The results of in vitro studies demonstrated that the nanosuspension was able to prevent burst release of loaded BSA and presented sustained-release behavior. It was no cytotoxicity by the bio-assessment using macrophage cells, and was observed significantly higher uptake compared with BSA free solution. Herein, ε-polylysine - sodium alginate nanoparticles had been found to be a potential candidate for vaccine delivery.

Toll-like receptor 4 signalling regulates antibody response to adenoviral vector-based vaccines by imprinting germinal centre quality.

Adenoviral vectors (AdV) are considered promising candidates for vaccine applications. A prominent group of Toll-like receptors (TLRs) participate in the adenovirus-induced adaptive immune response, yet there is little information regarding the role of TLR4 in AdV-induced immune responses in recent literature. We investigated the function of TLR4 in both adaptive and innate immune responses to an AdV-based anthrax vaccine. By immunizing wild-type and TLR4 knockout (TLR4-KO) mice, we revealed the requirement of TLR4 in AdV-induced innate responses. We also showed that TLR4 functions are required for germinal centre responses in immunized mice, as expression of the apoptosis-related marker Fas was down-regulated on germinal centre B cells from TLR4-KO mice. Likewise, decreased expression of inducible costimulator on follicular T helper cells was observed in immunized TLR4-KO mice. Moreover, a potent protective antigen-specific humoral immune response was mimicked using an adjuvant system containing the TLR4 agonist monophosphoryl lipid A. Overall, our findings showed that very rapid antigen-specific antibody production is correlated with the TLR4-imprinted germinal centre response to AdV-based vaccine. These results provide additional evidence for the use of the AdV and a TLR agonist to induce humoral responses. Our findings offer new insights into rational vaccine design.

Chitosan stabilized nasal emulsion delivery system for effective humoral and cellular response against recombinant tetravalent dengue antigen

Nasal vaccine delivery systems are emerging alternatives to the conventional sub unit vaccine delivery systems owing to their ability to stimulate potent antigen specific humoral and cellular immune responses. Additional virtue of nasal delivery is its close proximity of immune cells to external epithelial layer which is the route of entry to pathogens. Toxicity of emulsion based vaccine delivery systems may be attributed to the presence of high quantities of surfactants used for stabilizing the emulsions. A safer approach would be to reduce physiologically unwanted surfactant burden in the emulsion to the bare limit to necessity. Oleic acid was used as oil phase due to its ability to enhance penetration of system in nasal mucosa. This emulsion was designed with the purpose that it activates the innate (TLR 4) and adaptive immune systems apart from performing its antigen delivery function. Proving the hypothesis, emulsion when immunized along with recombinant tetravalent dengue antigen has elicited a profound antigen specific humoral and cellular response. Antigen cross presenting and sustained release of antigen by emulsion is the key factor in shaping this immune response. Moreover, the dose sparing effect of emulsion has also been proven which has a crucial role in modern day vaccine delivery. This significant humoral and cellular response elicited proves the suitability of this emulsion system for enhancing the protective effect of vaccines against various intracellular pathogens.

Cell Penetrating Peptide-Based Redox-Sensitive Vaccine Delivery System for Subcutaneous Vaccination.

In immunotherapy, induction of potent cellular immunity by vaccination is essential to treat intracellular infectious diseases and tumors. In this work, we designed a new synthetic peptide carrier, Cys-Trp-Trp-Arg8-Cys-Arg8-Cys-Arg8-Cys, for vaccine delivery by integrating a redox-responsive disulfide bond cross-linking and cell-penetrating peptide arginine octamer. The carrier peptide bound to the antigen protein ovalbumin (OVA) via electrostatic self-assembly to form peptide/OVA nanocomposites. Then, the spontaneous oxidization of the thiols of the cysteine residues induced interpeptide disulfide bond cross-linking to construct denser peptide/OVA condensates. The cell-penetrating peptides incorporated in the carrier peptide could increase antigen uptake by antigen presenting cells. After being internalized by antigen presenting cells, the antigen could be rapidly released in cytoplasm along with degradation of the disulfide bonds by intracellular glutathione, which could promote potent CD8+ T cell immunity. The cross-linked peptide/OVA condensates were used for subcutaneous vaccination. The results showed that the peptide carrier mediated potent antigen-specific immune response by significantly increasing IgG titer; splenocyte proliferation; the secretion level of cytokines INF-γ, IL-12, IL-4, and IL-10; immune memory function, and the activation and maturation of dendritic cells. From the results, the low-molecular weight vaccine-condensing peptide with definite chemical composition could be developed as a novel class of vaccine delivery systems.

Mast cell activator compound 48/40 is not an effective adjuvant for UV-attenuated Toxoplasma gondii vaccine.

Toxoplasma gondii (T. gondii, Tg) is a globally distributed parasitic protozoan causing different forms of toxoplasmosis in humans. Mast cells (MCs) play a role during T. gondii infection. Several studies suggest that MC activator compound 48/80 (C48/80) may be an effective vaccine adjuvant resulting in a potent and protective antigen-specific immune response against bacteria or virus infections. The present study was performed to determine whether C48/80 had adjuvant activity for ultraviolet (UV)-attenuated T. gondii vaccine to induce protective immune responses against T. gondii in mouse model. Kunming mice were divided into the following groups: naive mice, naive mice administrated with C48/80 intraperitoneal (i.p.) injection, mice infected by i.p. injection of 104 T. gondii RH strain alone (Tg group), mice infected with 104 RH tachyzoites plus C48/80 administration (Tg+C48/80), mice immunized with UV-Tg alone, and mice immunized with UV-Tg plus C48/80 administration (UV-Tg+C48/80). All the vaccinated mice were challenged with 104 tachyzoites of T. gondii RH strain at the same time as the primary infection. The survival rates, liver histopathologies, liver parasite burdens, and mRNA expression levels of Th1 and Th2 cytokines in the livers and spleens detected by quantitative real-time reverse transcription-polymerase chain reaction (qRT-PCR) were compared among the aforementioned groups after primary infection or challenge infection. The results showed that, compared to the Tg group or Tg+C48/80 group, the UV-Tg+Tg group and UV-Tg+C48/80+Tg group had significantly prolonged survival time, lower liver histopathological scores, decreased liver parasite burdens, and increased levels of Th1 and Th2 cytokines in the livers and spleens. There was no significant difference of survival time between the UV-Tg+Tg group and the UV-Tg+C48/80+Tg group; however, the UV-Tg+C48/80+Tg group showed higher parasite burden, more severe liver histopathology, and decreased IL-4 level compared to the UV-Tg+Tg group. These results indicate that C48/80 had no adjuvant activity for the immunization induced by UV-attenuated T. gondii vaccine.

Dendritic cell-derived exosomes elicit tumor regression in autochthonous hepatocellular carcinoma mouse models

Dendritic cell (DC)-derived exosomes (DEXs) form a new class of vaccines for cancer immunotherapy. However, their potency in hepatocellular carcinoma (HCC), a life-threatening malignancy with limited treatment options in the clinic that responds poorly to immunotherapy, remains to be investigated. Exosomes derived from α-fetoprotein (AFP)-expressing DCs (DEXAFP) were investigated in three different HCC mouse models systemically. Tumor growth and microenvironment were monitored. DEXAFP elicited strong antigen-specific immune responses and resulted in significant tumor growth retardation and prolonged survival rates in mice with ectopic, orthotopic and carcinogen-induced HCC tumors that displayed antigenic and pathological heterogeneity. The tumor microenvironment was improved in DEXAFP-treated HCC mice, demonstrated by significantly more γ-interferon (IFN-γ)-expressing CD8+ T lymphocytes, elevated levels of IFN-γ and interleukin-2, and fewer CD25+Foxp3+ regulatory T (Treg) cells and decreased levels of interleukin-10 and transforming growth factor-β in tumor sites. Lack of efficacy in athymic nude mice and CD8+ T cell-depleted mice showed that T cells contribute to DEXAFP-mediated antitumor function. Dynamic examination of the antitumor efficacy and the immune microenvironment in DEXAFP-treated orthotopic HCC mice at different time-points revealed a positive correlation between tumor suppression and immune microenvironment. Our findings provide evidence that AFP-enriched DEXs can trigger potent antigen-specific antitumor immune responses and reshape the tumor microenvironment in HCC mice and thus provide a cell-free vaccine option for HCC immunotherapy. Lay summary: Dendritic cell (DC)-derived exosomes (DEXs) form a new class of vaccines for cancer immunotherapy. However, their potency in hepatocellular carcinoma (HCC) remains unknown. Here, we investigated exosomes from HCC antigen-expressing DCs in three different HCC mouse models and proved their feasibility and capability of treating HCC, and thus provide a cell-free vaccine for HCC immunotherapy.

Thermo-sensitive hydrogel PLGA-PEG-PLGA as a vaccine delivery system for intramuscular immunization.

In this work, we explored the potential of thermo-sensitive PLGA-PEG-PLGA with sol-gel transition temperature around 32℃ as an intramuscular vaccine delivery system by using ovalbumin as a model antigen. First, invitro release test showed that the PLGA-PEG-PLGA-deriving hydrogels could release ovalbumin invitro in a more sustainable way. From fluorescence living imaging, 50-200 mg/mL of PLGA-PEG-PLGA formulations could release antigen in a sustainable manner invivo, suggesting that the PLGA-PEG-PLGA hydrogel worked as an antigen-depot. Further, the sustainable antigen release from the PLGA-PEG-PLGA hydrogels increased antigen availability in the spleens of the immunized mice. The intramuscular immunization results showed that 50-200 mg/mL of PLGA-PEG-PLGA formulations promoted significantly more potent antigen-specific IgG immune response. In addition, 200 mg/mL of PLGA-PEG-PLGA formulation significantly enhanced the secretion of both Th1 and Th2 cytokines. From invitro splenocyte proliferation assay, 50-200 mg/mL of PLGA-PEG-PLGA formulations all initiated significantly higher splenocyte activation. These results indicate that the thermo-sensitive and injectable PLGA-PEG-PLGA hydrogels (particularly, 200 mg/mL of PLGA-PEG-PLGA-based hydrogel) own promising potential as an intramuscular vaccine delivery system.

Abstract CT032: A first-in-human phase I/II clinical trial assessing novel mRNA-lipoplex nanoparticles for potent cancer immunotherapy in patients with malignant melanoma

Abstract Immunotherapeutic approaches have evolved as promising and valid alternatives to available conventional cancer treatments. Amongst others, vaccination with tumor antigen-encoding RNAs by local administration is currently successfully employed in various clinical trials. To allow for a more efficient targeting of antigen-presenting cells (APCs) and to overcome potential technical challenges associated with local administration, we have developed a novel RNA immunotherapeutic for systemic application based on a fixed set of four liposome complexed RNA drug products (RNA(LIP)), each encoding one shared melanoma-associated antigen. The novel RNA(LIP) formulation was engineered (i) to protect RNA from degradation by plasma RNases and (ii) to enable directed in vivo targeting of APCs in lymphoid compartments, thus (iii) allowing for intravenous administration of multiple RNA products advancing from local to systemic targeting of APCs. Here, RNA(LIP) products trigger a Toll-like receptor (TLR)-mediated Interferon-α (IFN-α) release from plasmacytoid dendritic cells (DCs) and macrophages stimulating DC maturation and hence inducing innate immune mechanisms as well as potent vaccine antigen-specific immune responses. Notably, BioNTech RNA Pharmaceuticals′ RNA(LIP) formulation is a universally applicable potent novel vaccine class for intravenous APC targeting and the induction of potent synchronized adaptive and type-I interferon-mediated innate immune responses for cancer immunotherapy. Similar to other liposomal drugs, the ready-to-use RNA(LIP) products are prepared individually in a straight-forward manner directly prior to use from three components, namely solutions containing RNA drug product, NaCl diluent, and liposome excipient, that are provided as a kit. A multi-center phase I/II trial to clinically validate this pioneering RNA(LIP) formulation for the treatment of malignant melanoma was initiated in 2015 (NCT02410733). The objective of the clinical trial is to study the feasibility, safety, tolerability, immunogenicity and evaluate potential clinical activity of the RNA(LIP) immunotherapy concept. Detailed information on the ongoing trial, the recruitment and treatment status as well as data on the assessment of vaccine-induced immune responses will be presented. Citation Format: Robert A. Jabulowsky, Carmen Loquai, Mustafa Diken, Lena M. Kranz, Heinrich Haas, Sebastian Attig, Nicole Bidmon, Janina Buck, Evelyna Derhovanessian, Jan Diekmann, Daniel Fritz, Veronika Jahndel, Alexandra Kemmer-Brueck, Klaus Kuehlcke, Andreas N. Kuhn, Peter Langguth, Ulrich Luxemburger, Martin Meng, Felicitas Mueller, Richard Rae, Fatih Sari, Doreen Schwarck-Kokarakis, Christine Seck, Kristina Spieß, Meike Witt, Jessica C. Hassel, Jochen Utikal, Roland Kaufmann, Sebastian Kreiter, Christoph Huber, Oezlem Tuereci, Ugur Sahin. A first-in-human phase I/II clinical trial assessing novel mRNA-lipoplex nanoparticles for potent cancer immunotherapy in patients with malignant melanoma. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr CT032.

Spontaneous Protein Adsorption on Graphene Oxide Nanosheets Allowing Efficient Intracellular Vaccine Protein Delivery.

Nanomaterials hold potential of altering the interaction between therapeutic molecules and target cells or tissues. High aspect ratio nanomaterials in particular have been reported to possess unprecedented properties and are intensively investigated for their interaction with biological systems. Graphene oxide (GOx) is a water-soluble graphene derivative that combines high aspect ratio dimension with functional groups that can be exploited for bioconjugation. Here, we demonstrate that GOx nanosheets can spontaneously adsorb proteins by a combination of interactions. This property is then explored for intracellular protein vaccine delivery, in view of the potential of GOx nanosheets to destabilize lipid membranes such as those of intracellular vesicles. Using a series of in vitro experiments, we show that GOx nanosheet adsorbed proteins are efficiently internalized by dendritic cells (DCs: the most potent class of antigen presenting cells of the immune system) and promote antigen cross-presentation to CD8 T cells. The latter is a hallmark in the induction of potent cellular antigen-specific immune responses against intracellular pathogens and cancer.