Receptor-binding domain recombinant protein on alum-CpG induces broad protection against SARS-CoV-2 variants of concern

We conducted preclinical studies in mice using a yeast-produced SARS-CoV-2 RBD subunit vaccine candidate formulated with aluminum hydroxide (alum) and CpG deoxynucleotides. This formulation is equivalent to the Corbevax^TM vaccine that recently received emergency use authorization by the Drugs Controller General of India. We compared the immune response of mice vaccinated with RBD/alum to mice vaccinated with RBD/alum + CpG. We also evaluated mice immunized with RBD/alum + CpG and boosted with RBD/alum. Mice were immunized twice intramuscularly at a 21-day interval. Compared to two doses of the /alum formulation, the RBD/alum + CpG vaccine induced a stronger and more balanced Th1/Th2 cellular immune response, with high levels of neutralizing antibodies against the original Wuhan isolate of SARS-CoV-2 as well as the B.1.1.7 (Alpha), B.1.351 (Beta), B.1.617.2 and (Delta) variants. Neutralizing antibody titers against the B.1.1.529 (BA.1, Omicron) variant exceeded those in human convalescent plasma after Wuhan infection but were lower than against the other variants. Interestingly, the second dose did not benefit from the addition of CpG, possibly allowing dose-sparing of the adjuvant in the future. The data reported here reinforces that the RBD/alum + CpG vaccine formulation is suitable for inducing broadly neutralizing antibodies against SARS-CoV-2, including variants of concern.     

Sublingual immunization with a subunit influenza vaccine elicits comparable systemic immune response as intramuscular immunization,but also induces local IgA and TH17 responses

Influenza is a vaccine-preventable disease that remains a major health problem world-wide. Needle and syringe are still the primary delivery devices, and injection of liquid vaccine into the muscle is still the primary route of immunization. Vaccines could be more convenient and effective if they were delivered by the mucosal route. Elicitation of systemic and mucosal innate and adaptive immune responses, such as pathogen neutralizing antibodies (including mucosal IgA at the site of pathogen entry) and CD4⁺ T-helper cells (especially the Th17 subset), have a critical role in vaccine-mediated protection. In the current study, a sublingual subunit influenza vaccine formulated with or without mucosal adjuvant was evaluated for systemic and mucosal immunogenicity and compared to intranasal and intramuscular vaccination. Sublingual administration of adjuvanted influenza vaccine elicited comparable antibody titers to those elicited by intramuscular immunization with conventional influenza vaccine. Furthermore, influenza-specific Th17 cells or neutralizing mucosal IgA were detected exclusively after mucosal immunization.     

Induction of multispecific Th-1 type immune response against HCV in mice by protein immunization using CpG and Montanide ISA 720 as adjuvants

Recent studies demonstrate that Th1-type immune responses against a broad spectrum of hepatitis C virus (HCV) gene products are crucial to the resolution of acute HCV infection. We investigated new vaccine approaches to augment the strength of HCV-specific Th1-type immune responses. ELISPOT assay revealed that single or multiple protein immunization using both CpG ODN and Montanide ISA 720 as adjuvants induced much stronger IFN-gamma-producing Th1 responses against core, NS3 and NS5b targets than did the formulation without these adjuvants. Protein vaccination using CpG ODN and Montanide ISA 720 as adjuvants also greatly enhanced humoral responses to HCV core, E1/E2 and NS3. When specific IgG isotypes were assayed, protein immunization using CpG ODN and Montanide ISA 720 as adjuvants produced higher titers of IgG2a dominant antibodies than did protein immunization alone, indicating a more Th1-biased pathway. This increase in IgG2a is consistent with the induction of Th1 cells secreting IFN-gamma demonstrated by ELISPOT assay. In conclusion, protein immunization using CpG ODN and Montanide ISA 720 as adjuvants greatly enhanced cellular (Th1 type) as well as humoral immune responses against HCV in Balb/c mice. The use of adjuvants appears critical to the induction of Th1 immune responses during HCV vaccination with recombinant proteins.     

Unmethylated CpG motif-containing genomic DNA fragments of bacillus calmette-guerin improves immune response towards a DNA vaccine for COVID-19

The development of an effective vaccine to control the global coronavirus disease-2019 (COVID-19) pandemic caused by the severe acute respiratory syndrome coronavirus- 2 (SARS-CoV-2) is of utmost importance. In this study, a synthetic DNA-based vaccine candidate, known as pSV10-SARS-CoV-2, expressing the SARS-CoV-2 spike protein was designed and tested in 39 BALB/c mice with BC01, an adjuvant derived from unmethylated CpG motif-containing DNA fragments from the Bacillus Calmette-Guerin genome. Mice vaccinated with pSV10-SARS-CoV-2 with BC01 produced early neutralizing antibodies and developed stronger humoral and cellular immune responses compared to mice that received the DNA vaccine only. Moreover, sera from mice vaccinated with pSV10-SARS-CoV-2 with BC01 can neutralize certain variants, including 614G, 614G + 472 V, 452R, 483A, 501Y.V2, and B.1.1.7. The results of this study demonstrate that the addition of BC01 to a DNA-vaccine for COVID-19 could elicit more effective neutralizing antibody titers for disease prevention.     

Oral, intrarectal and intranasal immunizations using CpG and non-CpG oligodeoxynucleotides as adjuvants

We have previously demonstrated that synthetic oligodeoxynucleotides (ODN) containing immunostimulatory CpG motifs (CpG ODN) are potent adjuvants in mice when delivered by intramuscular, intranasal and subcutaneous routes. Herein, using tetanus toxoid (TT) as a model antigen in BALB/c mice, we compared the ability of CpG ODN to induce mucosal and systemic humoral immune responses when antigen was delivered by three different routes: intrarectal, intranasal and oral. Results showed differences in immune responses with the three routes and also revealed that non-CpG "control" ODN had adjuvant effects when used at mucosal sites. This was unexpected since non-CpG ODN do not have such immunostimulatory effects in vitro or after parenteral immunization. These findings were further investigated after oral delivery of a killed influenza vaccine on its own as well as combined with TT and hepatitis B surface antigen. Our findings demonstrate that with mucosal delivery, there is a Th2 immunostimulatory effect associated with the phosphorothioate ODN backbone, and that the presence of CpG motifs shifts this towards a Th1 response.     

Homologous prime-boost with Zika virus envelope protein and poly (I:C) induces robust specific humoral and cellular immune responses

The recent outbreaks of Zika virus (ZIKV) infection and the potential association with Guillain-Barré syndrome in adults and with congenital abnormalities have highlighted the urgency for an effective vaccine. The ZIKV Envelope glycoprotein (E_ZIKV) is the most abundant protein on the virus surface, and has been evaluated together with the pre-membrane protein (prM) of the viral coat as a vaccine candidate in clinical trials. In this study, we performed a head-to-head comparison of the immune response induced by different E_ZIKV-based vaccine candidates in mice. We compared different platforms (DNA, recombinant protein), adjuvants (poly (I:C), CpG ODN 1826) and immunization strategies (homologous, heterologous).The hierarchy of adjuvant potency showed that poly (I:C) was a superior adjuvant than CpG ODN. While poly (I:C) assisted immunization reached a plateau in antibody titers after two doses, the CpG ODN group required an extra immunization dose. Besides, the administration of poly (I:C) induced higher E_ZIKV-specific cellular immune responses than CpG ODN. We also show that immunization with homologous prime-boost E_ZIKV protein + poly (I:C) regimen induced a more robust humoral response than homologous DNA (pVAX-E_ZIKV) or heterologous regimens (DNA/protein or protein/DNA). A detailed analysis of cellular immune responses revealed that homologous (E_ZIKV + poly (I:C)) and heterologous (pVAX-E_ZIKV/E_ZIKV + poly (I:C)) prime-boost regimens induced the highest magnitude of IFN-γ secreting cells and cytokine-producing CD4⁺ T cells.Overall, our data demonstrate that homologous E_ZIKV + poly (I:C) prime-boost immunization is sufficient to induce more robust specific-E_ZIKV humoral and cellular immune responses than the other strategies that contemplate homologous DNA (pVAX-E_ZIKV) or heterologous (pVAX-E_ZIKV/E_ZIKV + poly (I:C), and vice-versa) immunizations.     

The swine CD81 enhances E2-based DNA vaccination against classical swine fever

Classical swine fever (CSF) is a highly contagious and economically important viral disease that affects the pig industry worldwide. The glycoprotein E2 of CSFV can induce neutralizing antibodies and protective immunity, and is widely used for novel vaccine development. The objective of this study was to explore whether a tetraspanin molecule CD81 could improve the immune responses of an E2-based DNA vaccine. Plasmids pVAX-CD81, pVAX-E2 and pVAX-CD81-E2 were constructed and the expression of target proteins was confirmed in BHK-21 cells by indirect immunofluorescence assay. BALB/c mice were divided into 5 groups and immunized with different plasmids (pVAX-E2, pVAX-CD81-E2, pVAX-E2 + pVAX-CD81, pVAX-CD81 and PBS) three times with two weeks interval. The results showed that the introduction of CD81 promoted higher humoral and cellular immune responses than E2 expression alone (P < 0.05). In addition, immunization with pVAX-CD81-E2 induced stronger immune responses than pVAX-E2 + pVAX-CD81. Furthermore, four groups of pigs were immunized with pVAX-E2, pVAX-CD81-E2, pVAX-CD81 and PBS, respectively. Humoral and cellular immune responses detection showed similar results with those in mice. Compared to pVAX-E2, pVAX-CD81-E2 induced higher titers of neutralizing antibodies after viral challenge and conferred stronger protection. These results confirmed the capacity of swine CD81 enhancing the humoral and cellular responses with an adjuvant effect on CSFV DNA vaccine. This is the first report demonstrating the adjuvant effect of CD81 to enhance the DNA vaccination for swine pathogen.     

Induction of protective immunity by vaccination against Chlamydia trachomatis using the major outer membrane protein adjuvanted with CpG oligodeoxynucleotide coupled to the nontoxic B subunit of cholera toxin

The present study was undertaken to test the efficacy of immunization with the native major outer membrane protein (nMOMP) of Chlamydia trachomatis mouse pneumonitis (MoPn) serovar in combination with a novel immunostimulatory adjuvant consisting of CpG oligodeoxynucleotide (ODN) linked to the nontoxic B subunit of cholera toxin (CTB-CpG) to elicit a protective immune response to C. trachomatis. High levels of Chlamydia-specific IgG antibodies were detected in the sera from BALB/c mice immunized intramuscularly and subcutaneously (i.m. + s.c.) with the nMOMP/CTB-CpG vaccine or with nMOMP adjuvanted with a mixture of CT and CpG ODN (CT + CpG). Further, these immunization schemes gave rise to significant T-cell-mediated Chlamydia-specific immune responses. No Chlamydia-specific humoral or cell-mediated immune responses were detected in the control mice vaccinated with ovalbumin together with either CTB-CpG or CT + CpG. Following an intranasal challenge with C. trachomatis the groups of mice immunized with nMOMP plus CTB-CpG, CT + CpG or live C. trachomatis were found to be protected based on their change in body weight and lung weight as well as number of inclusion forming unit recovered from the lungs, as compared with control groups immunized with ovalbumin plus either adjuvants. Interestingly, IFN-γ-producing CD4⁺, but not CD8⁺, T-cells showed a significant correlation with the outcomes of the challenge. In conclusion, nMOMP in combination with the novel adjuvant CTB-CpG elicited a significant antigen-specific antibody and cell-mediated immune responses as well as protection against a pulmonary challenge with C. trachomatis.     

CpG oligonucleotide,host defense peptide and polyphosphazene act synergistically,inducing long-lasting,balanced immune responses in cattle

Vaccines consisting of subunit or protein antigens are less immunogenic than traditional vaccines, and therefore require formulation with an adjuvant. Conventional adjuvants, however, often cause undesirable injection site reactions and Th2-biased immune responses. Therefore, novel vaccine adjuvants which can safely enhance and selectively bias the resulting immune response are required. Here the adjuvant combination of CpG ODN, indolicidin and polyphosphazene (CpG + indol + PP) was evaluated for its ability to enhance and modulate the immune response when formulated with the antigen hen egg lysozyme (HEL). Cattle immunized with HEL co-adjuvanted with CpG + indol + PP developed higher antigen-specific humoral responses, and long-lasting cell-mediated immune responses, as evidenced by elevated levels of IFN-γ secretion by re-stimulated PBMCs, that were superior even to EMULSIGEN^®, an oil-in-water based adjuvant that was used as positive control. Physical characterization of the vaccines indicated that formulation of HEL with CpG + indol + PP resulted in the formation of antigen-adjuvant complexes, which may have contributed to their enhanced immunogenicity. Furthermore, the addition of polyphosphazene to CpG ODN and indolicidin dose-dependently enhanced the secretion of the cytokines IFN-α, TNF-α and IFN-γ in vitro, indicating that polyphosphazene can also synergize with CpG ODN and indolicidin to stimulate innate immune responses.     

Intranasal c-di-GMP-adjuvanted plant-derived H5 influenza vaccine induces multifunctional Th1 CD4+ cells and strong mucosal and systemic antibody responses in mice

Vaccination is the best available measure of limiting the impact of the next influenza pandemic. Ideally, a candidate pandemic influenza vaccine should be easy to administer and should elicit strong mucosal and systemic immune responses. Production of influenza subunit antigen in transient plant expression systems is an alternative to overcome the bottleneck in vaccine supply during influenza pandemic. Furthermore, a needle-free intranasal influenza vaccine is an attractive approach, which may provide immunity at the portal of virus entry.The present study investigated the detailed humoral and cellular immune responses in mice vaccinated intranasally or intramuscularly with plant-derived influenza H5N1 (A/Anhui/1/05) antigen alone or formulated with bis-(3′,5′)-cyclic dimeric guanosine monophosphate (c-di-GMP) as adjuvant. The use of c-di-GMP as intramuscular adjuvant did not enhance the immune response to plant-derived influenza H5 antigen. However, intranasal c-di-GMP-adjuvanted vaccine induced strong mucosal and systemic humoral immune responses. Additionally, the intranasal vaccine elicited a balanced Th1/Th2 profile and, most importantly, high frequencies of multifunctional Th1 CD4⁺ cells. Our results highlight that c-di-GMP is a promising mucosal adjuvant for pandemic influenza vaccine development.     

The potential of oligodeoxynucleotides as mucosal and parenteral adjuvants

Synthetic oligodeoxynucleotides (ODN) containing immunostimulatory CpG motifs (CpG ODN) are potent adjuvants in mice when delivered by parenteral (intramuscular, subcutaneous) and mucosal (intranasal, oral and intrarectal) routes. We have recently shown that with mucosal delivery non-CpG ODN can also have immunostimulatory properties which, in contrast to the Th1-bias characteristic of CpG ODN, are predominantly Th2-like. Herein, using hepatitis B surface antigen (HBsAg) and tetanus toxoid (TT) as model antigens in BALB/c mice, we have examined a number of different ODN (CpG, non-CpG, poly-T, poly-CG) to determine their effects on immune responses after mucosal (oral) and parenteral (IM) immunizations. Our findings demonstrate that with mucosal delivery, there is a Th2-biased immunostimulatory effect that is associated with non-CpG ODN, and that the presence of CpG motifs can shift this towards a Th1 response. The adjuvant effect of non-CpG ODN was much less evident after parenteral immunization.     

Development of a subunit vaccine containing recombinant Riemerella anatipestifer outer membrane protein A and CpG ODN adjuvant

Riemerella anatipestifer, a Gram-negative bacillus, causes septicemia that can result in high mortality for ducklings. In this study, we evaluated the immune response and protective efficacy provided by a subunit vaccine containing recombinant outer membrane protein A (rOmpA) and plasmid constructs containing CpG oligodeoxynucleotides (ODN). Results showed that CpG ODN enhanced both humoral and cell-mediated immunity elicited by rOmpA as early as two weeks after primary immunization. When compared to ducks immunized with rOmpA, ducks immunized with rOmpA + CpG ODN showed higher levels (p < 0.05) of antibody titer, T cell proliferation, and percentages of CD4⁺ and CD8⁺ T cell in peripheral blood mononuclear cells (PBMCs). The relative fold inductions of mRNA expression of Th1-type (IFN-γ and IL-12), and Th2-type (IL-6) cytokines in PBMCs isolated from ducks immunized with rOmpA + CpG ODN were significantly higher than those of the rOmpA group. Homologous challenge result showed that the rOmpA + CpG ODN vaccine reduced the pathological score by 90% in comparison with the saline control. In conclusion, our study found that CpG ODN can enhance both humoral and cellular immunity elicited by a rOmpA vaccine. The rOmpA + CpG ODN vaccine can be further developed as a subunit vaccine against R. anatipestifer.     

Toll-like receptor 3 adjuvant in combination with virus-like particles elicit a humoral response against HIV

Human Immunodeficiency Virus (HIV) Virus-Like Particles (VLPs) composed of HIV_IIIB Gag and HIV_BaL gp120/gp41 envelope are a pseudovirion vaccine capable of presenting antigens in their native conformations. To enhance the immunogenicity of the HIV Env antigen, VLPs were coupled to VesiVax Conjugatable Adjuvant Lipid Vesicles (CALV) containing one of four toll-like-receptor (TLR) ligands, each activating a receptor with distinct cellular localization and downstream pathways. C57BL/6 mice were vaccinated by intranasal prime followed by two sub-cheek boosts and their sera immunoglobulin and neutralizing potency were measured over a duration of 3 months after vaccination. PBS control, VLPs alone, CALV + VLPs, and VLPs complexed with CALV and ligands for TLR2 (PAM3CAG), TLR3 (dsRNA), TLR4 (MPLA), or TLR7/8 (resiquimod) were evaluated based on antibody titer, IgG1 and IgG2c class switching, germinal center formation, T follicular cells and potency of neutralizing antibodies. Consistently, the TLR3 ligand dsRNA complexed to CALV and in combination with VLPs (CALV(dsRNA) + VLPs) induced the strongest response. CALV(dsRNA) + VLPs induced the highest titers against the recombinant vaccine antigens clade B Bal gp120 and pr55 Gag. Additionally, CALV(dsRNA) + VLPs induced cross-clade antibodies, represented by high titers of antibody to clade c 96ZM651 gp120. CALV(dsRNA) + VLPs induced predominantly IgG2c over IgG1, a response associated with T helper type 1 (Th1)-like cytokines. In turn, CALV(dsRNA) + VLP immunized mice generated the most potent neutralizing antibodies against HIV strain MN.3. Finally, at time of sacrifice, a significant increase in germinal center B cells and T follicular cells was detected in mice which received CALV(dsRNA) + VLPs compared to PBS. Our results indicate that CALV(dsRNA) is a superior adjuvant for HIV VLPs in generating a Th1-like immunoglobulin profile, while prolonging lymph node germinal centers, T follicular cells and generating neutralizing antibodies to a highly sensitive tier 1A variant of HIV.     

Liposomes containing monophosphoryl lipid A and QS-21 serve as an effective adjuvant for soluble circumsporozoite protein malaria vaccine FMP013

Malaria caused by Plasmodium falciparum continues to threaten millions of people living in the tropical parts of the world. A vaccine that confers sterile and life-long protection remains elusive despite more than 30 years of effort and resources invested in solving this problem. Antibodies to a malaria vaccine candidate circumsporozoite protein (CSP) can block invasion and can protect humans against malaria. We have manufactured the Falciparum Malaria Protein-013 (FMP013) vaccine based on the nearly full-length P. falciparum CSP 3D7 strain sequence. We report here immunogenicity and challenge data on FMP013 antigen in C57BL/6 mice formulated with two novel adjuvants of the Army Liposome Formulation (ALF) series and a commercially available adjuvant Montanide ISA 720 (Montanide) as a control. ALF is a liposomal adjuvant containing a synthetic monophosphoryl lipid A (3D-PHAD®). In our study, FMP013 was adjuvanted with ALF alone, ALF containing aluminum hydroxide (ALFA) or ALF containing QS-21 (ALFQ). Adjuvants ALF and ALFA induced similar antibody titers and protection against transgenic parasite challenge that were comparable to Montanide. ALFQ was superior to the other three adjuvants as it induced higher antibody titers with improved boosting after the third immunization, higher serum IgG2c titers, and enhanced protection. FMP013 + ALFQ also augmented the numbers of splenic germinal center-derived activated B-cells and antibody secreting cells compared to Montanide. Further, FMP013 + ALFQ induced antigen-specific IFN-γ ELISPOT activity, CD4⁺ T-cells and a T_H1-biased cytokine profile. These results demonstrate that soluble CSP can induce a potent and sterile protective immune response when formulated with the QS-21 containing adjuvant ALFQ. Comparative mouse immunogenicity data presented here were used as the progression criteria for an ongoing non-human primate study and a regulatory toxicology study in preparation for a controlled human malaria infection (CHMI) trial.     

DNA vaccine encoding Middle East respiratory syndrome coronavirus S1 protein induces protective immune responses in mice

The Middle East respiratory syndrome coronavirus (MERS-CoV), is an emerging pathogen that continues to cause outbreaks in the Arabian peninsula and in travelers from this region, raising the concern that a global pandemic could occur. Here, we show that a DNA vaccine encoding the first 725 amino acids (S1) of MERS-CoV spike (S) protein induces antigen-specific humoral and cellular immune responses in mice. With three immunizations, high titers of neutralizing antibodies (up to 1: 10⁴) were generated without adjuvant. DNA vaccination with the MERS-CoV S1 gene markedly increased the frequencies of antigen-specific CD4⁺ and CD8⁺ T cells secreting IFN-γ and other cytokines. Both pcDNA3.1-S1 DNA vaccine immunization and passive transfer of immune serum from pcDNA3.1-S1 vaccinated mice protected Ad5-hDPP4-transduced mice from MERS-CoV challenge. These results demonstrate that a DNA vaccine encoding MERS-CoV S1 protein induces strong protective immune responses against MERS-CoV infection.     

Novel Th1-biased adjuvant, SPO1, enhances mucosal and systemic immunogenicity of vaccines administered intranasally in mice

Oil-in-water emulsions are potent human adjuvants commonly used in effective pandemic influenza vaccines; however, such emulsions that can induce both Th1-biased systemic immune responses and strong mucosal immune responses via an easy method of administration are lacking. To address this need for new adjuvants, we developed a novel oil/water emulsion, SPO1, which allows convenient mucosal immunization via an intranasal spray as well as by parenteral routes. Our report shows that SPO1 was able to boost up immunological resistance by inducing effective mucosal and serum antibodies, and the immune response was polarized to a Th1 pattern, as demonstrated by high IgG2α antibody levels and interferon-gamma production by splenocytes from intranasally (i.n.) immunized mice. Up-regulation of co-stimulatory and antigen-presenting molecules on dendritic cells was also observed in vivo after i.n. immunization, suggesting a possible mechanism for the adjuvant effects of SPO1. Another explanation may simply be a depot of antigen at the immunization site, as evidenced by in vivo imaging of i.n. immunized mice. In conclusion, our results demonstrate that a novel oil/water emulsion, SPO1, is a potent Th1 adjuvant for use in influenza and other vaccines, as it induces strong mucosal and systemic immune responses.     

A novel triple adjuvant formulation promotes strong, Th1-biased immune responses and significant antigen retention at the site of injection

Ovalbumin (OVA) was labeled with a near infra-red dye (*OVA) and formulated with the host defense peptide indolicidin (Indol), CpG oligodeoxynucleotide (ODN) 1826 (CpG) and/or poly(p-dicarboxylatophenoxy)-phosphazene (PP4). The immunogenicity of these *OVA formulations was evaluated in mice. All double and triple adjuvant combinations elicited strong antibody responses. *OVA formulated with CpG ODN in combination with indolicidin, PP4 or both induced only IFN-γ, while formulations with indolicidin and/or PP4 promoted predominantly IL-5 production. Overall, both IgG and IFN-γ production was superior when *OVA was combined with CpG/Indol/PP4. Furthermore, mice injected with *OVA formulated with CpG/Indol/PP4 contained detectable *OVA in the injection site two months post immunization. These results indicate that the CpG/Indol/PP4 combination promotes prolonged antigen retention and strong, antigen-specific Th1-biased immune responses.     

Poly(γ-glutamic acid) nano-particles combined with mucosal influenza virus hemagglutinin vaccine protects against influenza virus infection in mice

Adding poly(γ-glutamic acid) nano-particles (γ-PGA-NPs), a safe, natural material, to subcutaneous immunization with influenza virus hemagglutinin (HA) vaccine increases the protective immune responses against influenza virus in mice. Here, we examined whether intranasal administration of the HA vaccine with γ-PGA-NPs would induce protection from influenza virus challenge in mice. Intranasal immunization with the mixture of γ-PGA-NPs and HA vaccine from an influenza virus strain A/PR/8/34 (H1N1) or A/New Caledonia/20/99 (H1N1) enhanced protection of mice from A/PR/8/34 infection. Intranasal immunization with A/New Caledonia/20/99 HA vaccine and γ-PGA-NPs induced cell-mediated immune responses and neutralizing antibody production for both A/New Caledonia/20/99 and A/PR/8/34. These data suggest that γ-PGA-NPs may have potential for clinical applications as a mucosal adjuvant.     

Specific memory B cell response and participation of CD4+ central and effector memory T cells in mice immunized with liposome encapsulated recombinant NE protein based Hepatitis E vaccine candidate

BackgroundLiposome encapsulated neutralizing epitope protein of Hepatitis E virus (HEV), rNEp, our Hepatitis E vaccine candidate, was shown to be immunogenic and safe in pregnant and non-pregnant mice and yielded sterilizing immunity in rhesus monkeys.MethodsThe current study in Balb/c mice assessed the levels and persistence of anti-HEV IgG antibodies by ELISA, frequencies of B, memory B, T and memory T cells by flow cytometry and HEV-specific IgG secreting memory B cells by ELISPOT till 420 days post immunization (PI) with 5 μg rNEp encapsulated in liposome based adjuvant (2 doses, 4 weeks apart). Mice immunized with a lower dose (1 μg) were assessed only for anamnestic response post booster dose.ResultsVaccine candidate immunized mice (5 μg dose) elicited strong anti-HEV IgG response that was estimated to persist for lifetime. At day 120 PI, frequency of memory B cells was higher in immunized mice than those receiving adjuvant alone. Anti-HEV IgG titers were lower in mice immunized with 1 μg dose. A booster dose yielded a heightened antibody response in mice with both high (>800 GMT, 5 μg) and low (⩽100 GMT, 1 μg) anti-HEV IgG titers. At day 6th post booster dose, HEV-specific antibody secreting plasma cells (ASCs) were detected in 100% and 50% of mice with high and low anti-HEV IgG titers, respectively, whereas the frequencies of CD4⁺ central and effector memory T cells were high in mice with high anti-HEV IgG titers only.ConclusionsTaken together, the vaccine candidate effectively generates persistent and anamnestic antibody response, elicits participation of CD4⁺ memory T cells and triggers memory B cells to differentiate into ASCs upon boosting. This approach of assessing the immunogenicity of vaccine candidate could be useful to explore the longevity of HEV-specific memory response in future HEV vaccine trials in human.     

Intranasal Protollin-formulated recombinant SARS S-protein elicits respiratory and serum neutralizing antibodies and protection in mice

The feasibility of developing a prophylactic vaccine against SARS was assessed by comparing the immune responses elicited by immunizing mice with a recombinant SARS spike glycoprotein (S-protein) formulated with different adjuvants, given by different routes. In both young and aged mice, an intranasal Protollin-formulated S-protein vaccine elicited high levels of antigen-specific IgG in serum, comparable to those elicited by an intramuscular Alum-adsorbed S-protein vaccine. Serum antibodies were shown to be virus neutralizing. Intranasal immunization of young mice with the Protollin-formulated vaccine elicited significant levels of antigen-specific lung IgA in contrast to mice immunized with the intramuscular vaccine in which no antigen-specific lung IgA was detected. Following live virus challenge of aged mice, no virus was detected in the lungs of intranasally immunized mice, in contrast to intramuscularly immunized mice whose lung virus titers were comparable to those observed in control mice.