STING agonists as promising vaccine adjuvants to boost immunogenicity against SARS-related coronavirus derived infection: possible role of autophagy.

As a major component of innate immunity and a positive regulator of interferons, the Stimulator of interferon gene (STING) has an immunotherapy potential to govern a variety of infectious diseases. Despite the recent advances regarding vaccines against COVID-19, nontoxic novel adjuvants with the potential to enhance vaccine efficacy are urgently desired. In this connection, it has been well-documented that STING agonists are applied to combat COVID-19. This approach is of major significance for boosting immune responses most likely through an autophagy-dependent manner in susceptible individuals against infection induced by severe acute respiratory syndrome Coronavirus (SARS­CoV­2). Given that STING agonists exert substantial immunomodulatory impacts under a wide array of pathologic conditions, these agents could be considered novel adjuvants for enhancing immunogenicity against the SARS-related coronavirus. Here, we intend to discuss the recent advances in STING agonists' recruitment to boost innate immune responses upon vaccination against SARS-related coronavirus infections. In light of the primordial role of autophagy modulation, the potential of being an antiviral vaccine adjuvant was also explored.

The synthesis and preliminary immunological evaluation of a dual-adjuvant SARS-CoV-2 RBD vaccine: Covalent integration of TLR7/8 and iNKT cell agonists.

Covalently linking an adjuvant to an antigenic protein enhances its immunogenicity by ensuring a synergistic delivery to the immune system, fostering a more robust and targeted immune response. Most adjuvant-protein conjugate vaccines incorporate only one adjuvant due to the difficulties in its synthesis. However, there is a growing interest in developing vaccines with multiple adjuvants designed to elicit a more robust and targeted immune response by engaging different aspects of the immune system for complex diseases where traditional vaccines fall short. Here, we pioneer the synthesis of a dual-adjuvants protein conjugate Vaccine 1 by assembling a toll-like receptor 7/8 (TLR7/8) agonist, an invariant natural killer T cell (iNKT) agonist with a clickable bicyclononyne (BCN). The BCN group can bio-orthogonally react with azide-modified severe acute respiratory syndrome coronavirus-2 receptor-binding domain (SARS-CoV-2 RBD) trimer antigen to give the three-component Vaccine 1. Notably, with a mere 3 µg antigen, it elicited a balanced subclass of IgG titers and 20-fold more IgG2a than control vaccines, highlighting its potential for enhancing antibody-dependent cellular cytotoxicity. This strategy provides a practicable way to synthesize covalently linked dual immunostimulants. It expands the fully synthetic self-adjuvant protein vaccine that uses a single adjuvant to include two different types of adjuvants.

Design, Synthesis, and Biological Evaluation of New 2,6,7-Substituted Purine Derivatives as Toll-like Receptor 7 Agonists for Intranasal Vaccine Adjuvants.

TLR7/8 agonists are versatile immune stimulators capable of treating various diseases such as viral infections, autoimmune, and cancer. Despite the structural similarity of TLR7/8, their immune stimulation mechanisms and time-course responses significantly differ. In this study, a new series of TLR7-selective agonists was synthesized utilizing the economical building block 2,6-dichloropurine. Compound 27b showed the most potent activity on hTLR7 with an EC50 of 17.53 nM and demonstrated high hTLR7 selectivity (224 folds against TLR8). 27b effectively stimulated the secretion of proinflammatory cytokines in mouse macrophages and enhanced intranasal vaccine efficacy against influenza A virus in vivo. Assessment of humoral and mucosal antibody titers confirmed that 27b elevates IgG and IgA levels, protecting against both homologous and heterologous influenza viral infections. These findings suggest that 27b is a promising candidate as a vaccine adjuvant to prevent viral infections or as a robust immunomodulator with prolonged activity for treating immune-suppressed diseases.

Monophosphoryl Lipid A-Rhamnose Conjugates as a New Class of Vaccine Adjuvants.

Adjuvant is an integral part of all vaccine formulations but only a few adjuvants with limited efficacies or application scopes are available. Thus, developing more robust and diverse adjuvants is necessary. To this end, a new class of adjuvants having α- and ß-rhamnose (Rha) attached to the 1- and 6'-positions of monophosphoryl lipid A (MPLA) was designed, synthesized, and immunologically evaluated in mice. The results indicated a synergistic effect of MPLA and Rha, two immunostimulators that function via interacting with toll-like receptor 4 and recruiting endogenous anti-Rha antibodies, respectively. All the tested MPLA-Rha conjugates exhibited potent adjuvant activities to promote antibody production against both protein and carbohydrate antigens. Overall, MPLA-α-Rha exhibited better activities than MPLA-ß-Rha, and 6'-linked conjugates were slightly better than 1-linked ones. Particularly, MPLA-1-α-Rha and MPLA-6'-α-Rha were the most effective adjuvants in promoting IgG antibody responses against protein antigen keyhole limpet hemocyanin and carbohydrate antigen sTn, respectively.

A Novel Polymer Nanoparticle Polydimethyl Diallyl Ammonium Chloride as An Adjuvant Enhances the Immune Response of SARS-CoV-2 Subunit Vaccine.

The Coronavirus Disease 2019 (COVID-19) pandemic caused by SARS-CoV-2 has a significant impact on global health and the economy. It has underscored the urgent need for a stable, easily produced and effective vaccine. This study presents a novel approach using SARS-CoV-2 spike (S) protein-conjugated nanoparticles (NPs) in combination with cyclic GMP-AMP (cGAMP) (S-NPs-cGAMP) as a subunit vaccine. When mice are immunized, the antiserum of S-NPs-cGAMP group exhibits a 16-fold increase in neutralizing activity against a pseudovirus, compared to S protein group. Additionally, S-NPs-cGAMP induces even higher levels of neutralizing antibodies. Remarkably, the vaccine also triggers a robust humoral immune response, as evidenced by a notable elevation in virus-specific IgG and IgM antibodies. Furthermore, after 42 days of immunization, there is an observed increase in specific immune cell populations in the spleen. CD3+CD4+ and CD3+CD8+T lymphocytes, as well as B220+CD19+ and CD3-CD49b+ NK lymphocytes, show an upward trend, indicating a positive cellular immune response. Moreover, the S-NPs-cGAMP demonstrates promising results against the Delta strain and exhibits good cross-neutralization potential against other variants. These findings suggest that pDMDAAC NPs is potential adjuvant and could serve as a versatile platform for future vaccine development.

cGAS-STING pathway agonists are promising vaccine adjuvants.

Adjuvants are of critical value in vaccine development as they act on enhancing immunogenicity of antigen and inducing long-lasting immunity. However, there are only a few adjuvants that have been approved for clinical use, which highlights the need for exploring and developing new adjuvants to meet the growing demand for vaccination. Recently, emerging evidence demonstrates that the cGAS-STING pathway orchestrates innate and adaptive immunity by generating type I interferon responses. Many cGAS-STING pathway agonists have been developed and tested in preclinical research for the treatment of cancer or infectious diseases with promising results. As adjuvants, cGAS-STING agonists have demonstrated their potential to activate robust defense immunity in various diseases, including COVID-19 infection. This review summarized the current developments in the field of cGAS-STING agonists with a special focus on the latest applications of cGAS-STING agonists as adjuvants in vaccination. Potential challenges were also discussed in the hope of sparking future research interests to further the development of cGAS-STING as vaccine adjuvants.

Potentials of saponins-based adjuvants for nasal vaccines.

Respiratory infections are a major public health concern caused by pathogens that colonize and invade the respiratory mucosal surface. Nasal vaccines have the advantage of providing protection at the primary site of pathogen infection, as they induce higher levels of mucosal secretory IgA antibodies and antigen-specific T and B cell responses. Adjuvants are crucial components of vaccine formulation that enhance the immunogenicity of the antigen to confer long-term and effective protection. Saponins, natural glycosides derived from plants, shown potential as vaccine adjuvants, as they can activate the mammalian immune system. Several licensed human vaccines containing saponins-based adjuvants administrated through intramuscular injection have demonstrated good efficacy and safety. Increasing evidence suggests that saponins can also be used as adjuvants for nasal vaccines, owing to their safety profile and potential to augment immune response. In this review, we will discuss the structure-activity-relationship of saponins, their important role in nasal vaccines, and future prospects for improving their efficacy and application in nasal vaccine for respiratory infection.

iNKT cell agonists as vaccine adjuvants to combat infectious diseases.

iNKT cells are a special type of T cell that acts as a link between the innate and adaptive immune systems, with the capacity to stimulate a wide range of cell types. The glycolipid α-galactosylceramide (αGC) is a robust agonist of iNKT cells and induces the secretion of Th1- and Th2-type cytokines. αGC and its analogs are widely used as adjuvants to enhance immune responses against viral, parasitic, and bacterial pathogens. This review first discusses the challenges of using free αGC as a vaccine adjuvant to treat infectious diseases. We next present strategies to realize the potential of the adjuvant effect of iNKT cell glycolipids, including (1) the use of Th1- or Th2-biasing αGC analogs, (2) covalent conjugation of glycolipid with antigen, (3) particulate vehicle-assisted delivery of glycolipid, (4) glycolipid-loaded cellular systems, (5) glycolipid combination with other immunostimulants, and (6) usage as mucosal adjuvants. Finally, we discuss future approaches for the development of iNKT cell agonists used as vaccine adjuvants against infectious diseases.

A Novel Prophylaxis Strategy Using Liposomal Vaccine Adjuvant CAF09b Protects against Influenza Virus Disease.

The SARS-CoV-2 pandemic caused a massive health and societal crisis, although the fast development of effective vaccines reduced some of the impact. To prepare for future respiratory virus pandemics, a pan-viral prophylaxis could be used to control the initial virus outbreak in the period prior to vaccine approval. The liposomal vaccine adjuvant CAF®09b contains the TLR3 agonist polyinosinic:polycytidylic acid, which induces a type I interferon (IFN-I) response and an antiviral state in the affected tissues. When testing CAF09b liposomes as a potential pan-viral prophylaxis, we observed that intranasal administration of CAF09b liposomes to mice resulted in an influx of innate immune cells into the nose and lungs and upregulation of IFN-I-related gene expression. When CAF09b liposomes were administered prior to challenge with mouse-adapted influenza A/Puerto Rico/8/1934 virus, it protected from severe disease, although the virus was still detectable in the lungs. However, when CAF09b liposomes were administered after influenza challenge, the mice had a similar disease course to controls. In conclusion, CAF09b may be a suitable candidate as a pan-viral prophylactic treatment for epidemic viruses, but must be administered prior to virus exposure to be effective.

Polysaccharides derived from Chinese medicinal herbs: A promising choice of vaccine adjuvants.

Adjuvants have been used in vaccines for a long time to promote the body's immune response, reducing vaccine dosage and production costs. Although many vaccine adjuvants are developed, the use in human vaccines is limited because of either limited action or side effects. Therefore, the development of new vaccine adjuvants is required. Many studies have found that natural polysaccharides derived from Traditional Chinese medicine (TCM) possess good immune promoting effects and simultaneously improve humoral, cellular and mucosal immunity. Recently polysaccharide adjuvants have attracted much attention in vaccine preparation because of their intrinsic characteristics: immunomodulation, biocompatibility, biodegradability, low toxicity and safety. This review article systematically analysed the literature on polysaccharides possessing vaccine adjuvant activity from TCM plants, such as Astragalus polysaccharide (APS), Rehmannia glutinosa polysaccharide (RGP), Isatis indigotica root polysaccharides (IRPS), etc. and their derivatives. We believe that polysaccharide adjuvants can be used to prepare the vaccines for clinical use provided their mechanisms of action are studied in detail.

Cyclodextrin metal-organic framework as vaccine adjuvants enhances immune responses.

It is urgently needed to develop novel adjuvants for improving the safety and efficacy of vaccines. Metal-organic frameworks (MOFs), with high surface area, play an important role in drug delivery. With perfect biocompatibility and green preparation process, the γ-cyclodextrin metal-organic framework (γ-CD-MOF) fabricated with cyclodextrin and potassium suitable for antigen delivery. In this study, we modified γ-CD-MOF with span-85 to fabricate the SP-γ-CD-MOF as animal vaccine adjuvants. The ovalbumin (OVA) as the model antigen was encapsulated into particles to investigate the immune response. SP-γ-CD-MOF displayed excellent biocompatibility in vitro and in vivo. After immunization, SP-γ-CD-MOF loaded with OVA could induce high antigen-specific IgG titers and cytokine secretion. Meanwhile, SP-γ-CD-MOF also significantly improved the proliferation of spleen cells and activated and matured the bone marrow dendritic cells (BMDCs). The study showed the potential of SP-γ-CD-MOF in vaccine adjuvants and provided a novel idea for the development of vaccine adjuvants.

Lipopolysaccharide Derived From the Lymphoid-Resident Commensal Bacteria Alcaligenes faecalis Functions as an Effective Nasal Adjuvant to Augment IgA Antibody and Th17 Cell Responses.

Alcaligenes spp., including A. faecalis, is a gram-negative facultative bacterium uniquely residing inside the Peyer's patches. We previously showed that A. faecalis-derived lipopolysaccharides (Alcaligenes LPS) acts as a weak agonist of toll-like receptor 4 to activate dendritic cells and shows adjuvant activity by enhancing IgG and Th17 responses to systemic vaccination. Here, we examined the efficacy of Alcaligenes LPS as a nasal vaccine adjuvant. Nasal immunization with ovalbumin (OVA) plus Alcaligenes LPS induced follicular T helper cells and germinal center formation in the nasopharynx-associated lymphoid tissue (NALT) and cervical lymph nodes (CLNs), and consequently enhanced OVA-specific IgA and IgG responses in the respiratory tract and serum. In addition, nasal immunization with OVA plus Alcaligenes LPS induced OVA-specific T cells producing IL-17 and/or IL-10, whereas nasal immunization with OVA plus cholera toxin (CT) induced OVA-specific T cells producing IFN-γ and IL-17, which are recognized as pathogenic type of Th17 cells. In addition, CT, but not Alcaligenes LPS, promoted the production of TNF-α and IL-5 by T cells. Nasal immunization with OVA plus CT, but not Alcaligenes LPS, led to increased numbers of neutrophils and eosinophils in the nasal cavity. Together, these findings indicate that the benign nature of Alcaligenes LPS is an effective nasal vaccine adjuvant that induces antigen-specific mucosal and systemic immune responses without activation of inflammatory cascade after nasal administration.

Anti-CD40 Antibody Fused to CD40 Ligand Is a Superagonist Platform for Adjuvant Intrinsic DC-Targeting Vaccines.

CD40 is a potent activating receptor expressed on antigen-presenting cells (APCs) of the immune system. CD40 regulates many aspects of B and T cell immunity via interaction with CD40L expressed on activated T cells. Targeting antigens to CD40 via agonistic anti-CD40 antibody fusions promotes both humoral and cellular immunity, but current anti-CD40 antibody-antigen vaccine prototypes require co-adjuvant administration for significant in vivo efficacy. This may be a consequence of dulling of anti-CD40 agonist activity via antigen fusion. We previously demonstrated that direct fusion of CD40L to anti-CD40 antibodies confers superagonist properties. Here we show that anti-CD40-CD40L-antigen fusion constructs retain strong agonist activity, particularly for activation of dendritic cells (DCs). Therefore, we tested anti-CD40-CD40L antibody fused to antigens for eliciting immune responses in vitro and in vivo. In PBMC cultures from HIV-1-infected donors, anti-CD40-CD40L fused to HIV-1 antigens preferentially expanded HIV-1-specific CD8+ T cells versus CD4+ T cells compared to analogous anti-CD40-antigen constructs. In normal donors, anti-CD40-CD40L-mediated delivery of Influenza M1 protein elicited M1-specific T cell expansion at lower doses compared to anti-CD40-mediated delivery. Also, on human myeloid-derived dendritic cells, anti-CD40-CD40L-melanoma gp100 peptide induced more sustained Class I antigen presentation compared to anti-CD40-gp100 peptide. In human CD40 transgenic mice, anti-CD40-CD40L-HIV-1 gp140 administered without adjuvant elicited superior antibody responses compared to anti-CD40-gp140 antigen without fused CD40L. In human CD40 mice, compared to the anti-CD40 vehicle, anti-CD40-CD40L delivery of Eα 52-68 peptide elicited proliferating of TCR I-Eα 52-68 CD4+ T cells producing cytokine IFNγ. Also, compared to controls, only anti-CD40-CD40L-Cyclin D1 vaccination of human CD40 mice reduced implanted EO771.LMB breast tumor cell growth. These data demonstrate that human CD40-CD40L antibody fused to antigens maintains highly agonistic activity and generates immune responses distinct from existing low agonist anti-CD40 targeting formats. These advantages were in vitro skewing responses towards CD8+ T cells, increased efficacy at low doses, and longevity of MHC Class I peptide display; and in mouse models, a more robust humoral response, more activated CD4+ T cells, and control of tumor growth. Thus, the anti-CD40-CD40L format offers an alternate DC-targeting platform with unique properties, including intrinsic adjuvant activity.

Programming Multifaceted Pulmonary T Cell Immunity by Combination Adjuvants.

Induction of protective mucosal T cell memory remains a formidable challenge to vaccinologists. Using a combination adjuvant strategy that elicits potent CD8 and CD4 T cell responses, we define the tenets of vaccine-induced pulmonary T cell immunity. An acrylic-acid-based adjuvant (ADJ), in combination with Toll-like receptor (TLR) agonists glucopyranosyl lipid adjuvant (GLA) or CpG, promotes mucosal imprinting but engages distinct transcription programs to drive different degrees of terminal differentiation and disparate polarization of TH1/TC1/TH17/TC17 effector/memory T cells. Combination of ADJ with GLA, but not CpG, dampens T cell receptor (TCR) signaling, mitigates terminal differentiation of effectors, and enhances the development of CD4 and CD8 TRM cells that protect against H1N1 and H5N1 influenza viruses. Mechanistically, vaccine-elicited CD4 T cells play a vital role in optimal programming of CD8 TRM and viral control. Taken together, these findings provide further insights into vaccine-induced multifaceted mucosal T cell immunity with implications in the development of vaccines against respiratorypathogens, including influenza virus and SARS-CoV-2.