RESUMO
Plasmodium falciparum merozoite surface protein (PfMSP)2 is a target of parasite-neutralizing Abs. Inclusion of recombinant PfMSP2 (rPfMSP2) as a component of a multivalent malaria vaccine is of interest, but presents challenges. Previously, we used the highly immunogenic PfMSP8 as a carrier to enhance production and/or immunogenicity of malaria vaccine targets. In this study, we exploited the benefits of rPfMSP8 as a carrier to optimize a rPfMSP2-based subunit vaccine. rPfMSP2 and chimeric rPfMSP2/8 vaccines produced in Escherichia coli were evaluated in comparative immunogenicity studies in inbred (CB6F1/J) and outbred (CD1) mice, varying the dose and adjuvant. Immunization of mice with both rPfMSP2-based vaccines elicited high-titer anti-PfMSP2 Abs that recognized the major allelic variants of PfMSP2. Vaccine-induced T cells recognized epitopes present in both PfMSP2 and the PfMSP8 carrier. Competition assays revealed differences in Ab specificities induced by the two rPfMSP2-based vaccines, with evidence of epitope masking by rPfMSP2-associated fibrils. In contrast to aluminum hydroxide (Alum) as adjuvant, formulation of rPfMSP2 vaccines with glucopyranosyl lipid adjuvant-stable emulsion, a synthetic TLR4 agonist, elicited Th1-associated cytokines, shifting production of Abs to cytophilic IgG subclasses. The rPfMSP2/8 + glucopyranosyl lipid adjuvant-stable emulsion formulation induced significantly higher Ab titers with superior durability and capacity to opsonize P. falciparum merozoites for phagocytosis. Immunization with a trivalent vaccine including PfMSP2/8, PfMSP1/8, and the P. falciparum 25 kDa sexual stage antigen fused to PfMSP8 (Pfs25/8) induced high levels of Abs specific for epitopes in each targeted domain, with no evidence of antigenic competition. These results are highly encouraging for the addition of rPfMSP2/8 as a component of an efficacious, multivalent, multistage malaria vaccine.
Assuntos
Antígenos de Protozoários/imunologia , Vacinas Antimaláricas/imunologia , Malária/imunologia , Merozoítos/metabolismo , Plasmodium falciparum/imunologia , Proteínas de Protozoários/imunologia , Células Th1/imunologia , Animais , Anticorpos Neutralizantes/metabolismo , Anticorpos Antiprotozoários/metabolismo , Antígenos de Protozoários/genética , Mapeamento de Epitopos , Feminino , Glucosídeos , Epitopos Imunodominantes , Imunoglobulina G/metabolismo , Lipídeo A , Vacinas Antimaláricas/genética , Masculino , Merozoítos/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Fagocitose , Proteínas de Protozoários/genéticaRESUMO
Challenges with the production and suboptimal immunogenicity of malaria vaccine candidates have slowed the development of a Plasmodium falciparum multiantigen vaccine. Attempting to resolve these issues, we focused on the use of highly immunogenic merozoite surface protein 8 (MSP8) as a vaccine carrier protein. Previously, we showed that a genetic fusion of the C-terminal 19-kDa fragment of merozoite surface protein 1 (MSP119) to P. falciparum MSP8 (PfMSP8) facilitated antigen production and folding and the induction of neutralizing antibodies to conformational B cell epitopes of MSP119 Here, using the PfMSP1/8 construct, we further optimized the recombinant PfMSP8 (rPfMSP8) carrier by the introduction of two cysteine-to-serine substitutions (CΔS) to improve the yield of the monomeric product. We then sought to test the broad applicability of this approach using the transmission-blocking vaccine candidate Pfs25. The production of rPfs25-based vaccines has presented challenges. Antibodies directed against the four highly constrained epidermal growth factor (EGF)-like domains of Pfs25 block sexual-stage development in mosquitoes. The sequence encoding mature Pfs25 was codon harmonized for expression in Escherichia coli We produced a rPfs25-PfMSP8 fusion protein [rPfs25/8(CΔS)] as well as unfused, mature rPfs25. rPfs25 was purified with a modest yield but required the incorporation of refolding protocols to obtain a proper conformation. In comparison, chimeric rPfs25/8(CΔS) was expressed and easily purified, with the Pfs25 domain bearing the proper conformation without renaturation. Both antigens were immunogenic in rabbits, inducing IgG that bound native Pfs25 and exhibited potent transmission-reducing activity. These data further demonstrate the utility of PfMSP8 as a parasite-specific carrier protein to enhance the production of complex malaria vaccine targets.
Assuntos
Proteínas de Transporte/imunologia , Vacinas Antimaláricas/imunologia , Plasmodium falciparum/imunologia , Proteínas de Protozoários/imunologia , Proteínas Recombinantes/imunologia , Adjuvantes Imunológicos/farmacologia , Animais , Anticorpos Antiprotozoários/imunologia , Antígenos de Protozoários/imunologia , Culicidae/parasitologia , Epitopos de Linfócito B/imunologia , Escherichia coli/metabolismo , Masculino , Proteína 1 de Superfície de Merozoito/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , CoelhosAssuntos
Betacoronavirus , Infecções por Coronavirus , Pandemias , Pneumonia Viral , Anticorpos Monoclonais , COVID-19 , Humanos , SARS-CoV-2RESUMO
COVID-19 remains a major public health concern. Monoclonal antibodies have received emergency use authorization (EUA) for pre-exposure prophylaxis against COVID-19 among high-risk groups for treatment of mild to moderate COVID-19. In addition to recombinant biologics, engineered synthetic DNA-encoded antibodies (DMAb) are an important strategy for direct in vivo delivery of protective mAb. A DMAb cocktail was synthetically engineered to encode the immunoglobulin heavy and light chains of two different two different Fc-engineered anti-SARS-CoV-2 antibodies. The DMAbs were designed to enhance in vivo expression and delivered intramuscularly to cynomolgus and rhesus macaques with a modified in vivo delivery regimen. Serum levels were detected in macaques, along with specific binding to SARS-CoV-2 spike receptor binding domain protein and neutralization of multiple SARS-CoV-2 variants of concern in pseudovirus and authentic live virus assays. Prophylactic administration was protective in rhesus macaques against signs of SARS-CoV-2 (USA-WA1/2020) associated disease in the lungs. Overall, the data support further study of DNA-encoded antibodies as an additional delivery mode for prevention of COVID-19 severe disease. These data have implications for human translation of gene-encoded mAbs for emerging infectious diseases and low dose mAb delivery against COVID-19.
Assuntos
COVID-19 , Profilaxia Pré-Exposição , Animais , Macaca mulatta , COVID-19/prevenção & controle , SARS-CoV-2/genética , Anticorpos Antivirais , Anticorpos Monoclonais , Macaca fascicularis , DNA , Anticorpos Neutralizantes , Glicoproteína da Espícula de Coronavírus/genéticaRESUMO
Ovarian cancer (OC) is a lethal gynecologic malignancy, with modest responses to CPI. Engagement of additional immune arms, such as NK cells, may be of value. We focused on Siglec-7 as a surface antigen for engaging this population. Human antibodies against Siglec-7 were developed and characterized. Coculture of OC cells with PBMCs/NKs and Siglec-7 binding antibodies showed NK-mediated killing of OC lines. Anti-Siglec-7 mAb (DB7.2) enhanced survival in OC-challenged mice. In addition, the combination of DB7.2 and anti-PD-1 demonstrated further improved OC killing in vitro. To use Siglec-7 engagement as an OC-specific strategy, we engineered an NK cell engager (NKCE) to simultaneously engage NK cells through Siglec-7, and OC targets through FSHR. The NKCE demonstrated robust in vitro killing of FSHR+ OC, controlled tumors, and improved survival in OC-challenged mice. These studies support additional investigation of the Siglec-7 targeting approaches as important tools for OC and other recalcitrant cancers.
Assuntos
Produtos Biológicos , Neoplasias Ovarianas , Feminino , Humanos , Camundongos , Animais , Produtos Biológicos/metabolismo , Células Matadoras Naturais , Neoplasias Ovarianas/terapia , Neoplasias Ovarianas/metabolismo , Antígenos CD/metabolismo , Lectinas Semelhantes a Imunoglobulina de Ligação ao Ácido Siálico/metabolismoRESUMO
Despite numerous clinically available vaccines and therapeutics, aged patients remain at increased risk for COVID-19 morbidity. Furthermore, various patient populations, including the aged can have suboptimal responses to SARS-CoV-2 vaccine antigens. Here, we characterized vaccine-induced responses to SARS-CoV-2 synthetic DNA vaccine antigens in aged mice. Aged mice exhibited altered cellular responses, including decreased IFNγ secretion and increased TNFα and IL-4 secretion suggestive of TH2-skewed responses. Aged mice exhibited decreased total binding and neutralizing antibodies in their serum but significantly increased TH2-type antigen-specific IgG1 antibody compared to their young counterparts. Strategies to enhance vaccine-induced immune responses are important, especially in aged patient populations. We observed that co-immunization with plasmid-encoded adenosine deaminase (pADA)enhanced immune responses in young animals. Ageing is associated with decreases in ADA function and expression. Here, we report that co-immunization with pADA enhanced IFNγ secretion while decreasing TNFα and IL-4 secretion. pADA expanded the breadth and affinity SARS-CoV-2 spike-specific antibodies while supporting TH1-type humoral responses in aged mice. scRNAseq analysis of aged lymph nodes revealed that pADA co-immunization supported a TH1 gene profile and decreased FoxP3 gene expression. Upon challenge, pADA co-immunization decreased viral loads in aged mice. These data support the use of mice as a model for age-associated decreased vaccine immunogenicity and infection-mediated morbidity and mortality in the context of SARS-CoV-2 vaccines and provide support for the use of adenosine deaminase as a molecular adjuvant in immune-challenged populations.
Assuntos
COVID-19 , SARS-CoV-2 , Humanos , Animais , Camundongos , Vacinas contra COVID-19 , Fator de Necrose Tumoral alfa , Interleucina-4 , Adenosina Desaminase , Imunização , Anticorpos Antivirais , Modelos Animais de DoençasRESUMO
Immune responses diminish with age resulting in an increased susceptibility of the elderly to infectious agents and an inability to mount protective immune responses to vaccines. Immunosenescence affects multiple aspects of the immune system, including CD8(+) T cells, which control viral infections and are assumed to prevent the development of cancers. In this study, we tested if CD8(+) T cell responses in aged mice could be enhanced through a vaccine that concomitantly expresses Ag and a molecule that blocks an immunoinhibitory pathway. Specifically, we tested a vaccine based on a replication-defective chimpanzee-derived adenovirus vector expressing the nucleoprotein (NP) of influenza A virus as a fusion protein with the HSV type 1 glycoprotein D, which through binding to the herpes virus entry mediator, blocks the immunoinhibitory herpes virus entry mediator B and T lymphocyte attenuator/CD160 pathways. Our results show that the vaccine expressing a fusion protein of NP and glycoprotein D induces significantly higher NP-specific CD8(+) T cell responses in young and aged mice compared with the vaccine expressing NP only.
Assuntos
Envelhecimento/imunologia , Linfócitos T CD8-Positivos/imunologia , Epitopos de Linfócito T/imunologia , Vírus da Influenza A/imunologia , Membro 14 de Receptores do Fator de Necrose Tumoral/antagonistas & inibidores , Transdução de Sinais/imunologia , Regulação para Cima/imunologia , Animais , Linfócitos T CD8-Positivos/metabolismo , Linfócitos T CD8-Positivos/virologia , Linhagem Celular , Epitopos de Linfócito T/genética , Feminino , Vetores Genéticos/imunologia , Herpesvirus Humano 1/genética , Herpesvirus Humano 1/imunologia , Humanos , Epitopos Imunodominantes/genética , Epitopos Imunodominantes/imunologia , Vírus da Influenza A Subtipo H1N1/genética , Vírus da Influenza A Subtipo H1N1/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Proteínas do Nucleocapsídeo , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/imunologia , Receptores Imunológicos/genética , Membro 14 de Receptores do Fator de Necrose Tumoral/fisiologia , Proteínas Recombinantes de Fusão/imunologia , Transdução de Sinais/genética , Membro 14 da Superfamília de Ligantes de Fatores de Necrose Tumoral/antagonistas & inibidores , Membro 14 da Superfamília de Ligantes de Fatores de Necrose Tumoral/fisiologia , Proteínas do Core Viral/genética , Proteínas do Core Viral/imunologiaRESUMO
Despite enormous efforts by the scientific community, an effective HIV vaccine remains elusive. To further address to what degree T cells in absence of antibodies may protect against simian immunodeficiency virus (SIV) disease progression, rhesus macaques were vaccinated intramuscularly with a chimpanzee-derived Ad vector (AdC) serotype 6 and then boosted intramuscularly with a serologically distinct AdC vector of serotype 7 both expressing Gag of SIVmac239. Animals were subsequently boosted intramuscularly with a modified vaccinia Ankara (MVA) virus expressing Gag and Tat of the homologous SIV before mucosal challenge with a high dose of SIVmac239 given rectally. Whereas vaccinated animals showed only a modest reduction of viral loads, their overall survival was improved, in association with a substantial protection from the loss of CD4(+) T cells. In addition, the two vaccinated Mamu-A*01(+) macaques controlled viral loads to levels below detection within weeks after challenge. These data strongly suggest that T cells, while unable to affect SIV acquisition upon high-dose rectal infection, can reduce disease progression. Induction of potent T-cell responses should thus remain a component of our efforts to develop an efficacious vaccine to HIV-1.
Assuntos
Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Macaca mulatta/imunologia , Macaca mulatta/virologia , Vacinas contra a SAIDS/imunologia , Vírus da Imunodeficiência Símia/imunologia , Vírus da Imunodeficiência Símia/patogenicidade , Animais , Feminino , MasculinoRESUMO
The global coronavirus disease 2019 (COVID-19) pandemic has claimed more than 5 million lives. Emerging variants of concern (VOCs) continually challenge viral control. Directing vaccine-induced humoral and cell-mediated responses to mucosal surfaces may enhance vaccine efficacy. Here we investigate the immunogenicity and protective efficacy of optimized synthetic DNA plasmids encoding wild-type severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike protein (pS) co-formulated with the plasmid-encoded mucosal chemokine cutaneous T cell-attracting chemokine (pCTACK; CCL27). pCTACK-co-immunized animals exhibit increased spike-specific antibodies at the mucosal surface and increased frequencies of interferon gamma (IFNγ)+ CD8+ T cells in the respiratory mucosa. pCTACK co-immunization confers 100% protection from heterologous Delta VOC challenge. This study shows that mucosal chemokine adjuvants can direct vaccine-induced responses to specific immunological sites and have significant effects on heterologous challenge. Further study of this unique chemokine-adjuvanted vaccine approach in the context of SARS-CoV-2 vaccines is likely important.
Assuntos
COVID-19 , Vacinas Virais , Adjuvantes Imunológicos/farmacologia , Animais , Anticorpos Antivirais , Linfócitos T CD8-Positivos , COVID-19/prevenção & controle , Vacinas contra COVID-19 , Quimiocinas , Humanos , SARS-CoV-2/genética , Vacinas Virais/genéticaRESUMO
The enhanced transmissibility and immune evasion associated with emerging SARS-CoV-2 variants demands the development of next-generation vaccines capable of inducing superior protection amid a shifting pandemic landscape. Since a portion of the global population harbors some level of immunity from vaccines based on the original Wuhan-Hu-1 SARS-CoV-2 sequence or natural infection, an important question going forward is whether this immunity can be boosted by next-generation vaccines that target emerging variants while simultaneously maintaining long-term protection against existing strains. Here, we evaluated the immunogenicity of INO-4800, our synthetic DNA vaccine candidate for COVID-19 currently in clinical evaluation, and INO-4802, a next-generation DNA vaccine designed to broadly target emerging SARS-CoV-2 variants, as booster vaccines in nonhuman primates. Rhesus macaques primed over one year prior with the first-generation INO-4800 vaccine were boosted with either INO-4800 or INO-4802 in homologous or heterologous prime-boost regimens. Both boosting schedules led to an expansion of T cells and antibody responses which were characterized by improved neutralizing and ACE2 blocking activity across wild-type SARS-CoV-2 as well as multiple variants of concern. These data illustrate the durability of immunity following vaccination with INO-4800 and additionally support the use of either INO-4800 or INO-4802 in prime-boost regimens.
Assuntos
COVID-19 , Vacinas de DNA , Vacinas Virais , Animais , Formação de Anticorpos , COVID-19/prevenção & controle , Vacinas contra COVID-19 , Humanos , Macaca mulatta , Camundongos , Camundongos Endogâmicos BALB C , SARS-CoV-2 , VacinaçãoRESUMO
Monoclonal antibody therapy has played an important role against SARS-CoV-2. Strategies to deliver functional, antibody-based therapeutics with improved in vivo durability are needed to supplement current efforts and reach underserved populations. Here, we compare recombinant mAbs COV2-2196 and COV2-2130, which compromise clinical cocktail Tixagevimab/Cilgavimab, with optimized nucleic acid-launched forms. Functional profiling of in vivo-expressed, DNA-encoded monoclonal antibodies (DMAbs) demonstrated similar specificity, broad antiviral potency and equivalent protective efficacy in multiple animal challenge models of SARS-CoV-2 prophylaxis compared to protein delivery. In PK studies, DNA-delivery drove significant serum antibody titers that were better maintained compared to protein administration. Furthermore, cryo-EM studies performed on serum-derived DMAbs provide the first high-resolution visualization of in vivo-launched antibodies, revealing new interactions that may promote cooperative binding to trimeric antigen and broad activity against VoC including Omicron lineages. These data support the further study of DMAb technology in the development and delivery of valuable biologics.
Assuntos
Produtos Biológicos , COVID-19 , Ácidos Nucleicos , Animais , Anticorpos Monoclonais , Anticorpos Neutralizantes , Anticorpos Antivirais , Antivirais/farmacologia , Antivirais/uso terapêutico , COVID-19/prevenção & controle , DNA , SARS-CoV-2 , Glicoproteína da Espícula de Coronavírus/genéticaRESUMO
A universal influenza vaccine, designed to induce broadly cross-reactive immunity against current and future influenza A virus strains, is in critical demand to reduce the need for annual vaccinations with vaccines chosen upon predicting the predominant circulating viral strains, and to ameliorate the threat of cyclically occurring pandemics that have, in the past, killed tens of millions. Here, we describe a vaccine regimen based on sequential immunization with two serologically distinct chimpanzee-derived replication-defective adenovirus (Ad) vectors expressing the matrix-2 protein ectodomain (M2e) from three divergent strains of influenza A virus fused to the influenza virus nucleoprotein (NP) for induction of antibodies to M2e and virus-specific CD8(+) T cells to NP. In preclinical mouse models, the Ad vaccines expressing M2e and NP elicit robust NP-specific CD8(+) T-cell responses and moderate antibody responses to all three M2e sequences. Most importantly, vaccinated mice are protected against morbidity and mortality following challenge with high doses of different influenza virus strains. Protection requires both antibodies to M2e and cellular immune responses to NP.
Assuntos
Adenoviridae , Vírus da Influenza A , Vacinas contra Influenza , Influenza Humana/prevenção & controle , Nucleoproteínas/metabolismo , Proteínas da Matriz Viral/metabolismo , Adenoviridae/genética , Sequência de Aminoácidos , Animais , Humanos , Vírus da Influenza A/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Dados de Sequência Molecular , Proteínas Recombinantes de Fusão/genéticaRESUMO
Monoclonal antibody (MAb) 2C7 recognizes a lipooligosaccharide epitope expressed by most clinical Neisseria gonorrhoeae isolates and mediates complement-dependent bactericidal activity. We recently showed that a recombinant human IgG1 chimeric variant of MAb 2C7 containing an E430G Fc modification (2C7_E430G), which enhances complement activation, outperformed the parental MAb 2C7 (2C7_WT) in vivo Because natural infection with N. gonorrhoeae often does not elicit protective immunity and reinfections are common, approaches that prolong bacterial control in vivo are of great interest. Advances in DNA-based approaches have demonstrated the combined benefit of genetic engineering, formulation optimizations, and facilitated delivery via CELLECTRA-EP technology, which can induce robust in vivo expression of protective DNA-encoded monoclonal antibodies (DMAbs) with durable serum activity relative to traditional recombinant MAb therapies. Here, we created optimized 2C7-derived DMAbs encoding the parental Fc (2C7_WT) or complement-enhancing Fc variants (2C7_E430G and 2C7_E345K). 2C7 DMAbs were rapidly generated and detected throughout the 4-month study. While all complement-engaging 2C7 variants facilitated rapid clearance following primary N. gonorrhoeae challenge (day 8 after DMAb administration), the complement-enhancing 2C7_E430G variant demonstrated significantly higher potency against mice rechallenged 65 days after DMAb administration. Passive intravenous transfer of in vivo-produced, purified 2C7 DMAbs confirmed the increased potency of the complement-enhancing variants. This study highlights the ability of the DMAb platform to launch the in vivo production of antibodies engineered to promote and optimize downstream innate effector mechanisms such as complement-mediated killing, leading to hastened bacterial elimination.IMPORTANCENeisseria gonorrhoeae has become resistant to most antibiotics in clinical use. Currently, there is no safe and effective vaccine against gonorrhea. Measures to prevent the spread of gonorrhea are a global health priority. A monoclonal antibody (MAb) called 2C7, directed against a lipooligosaccharide glycan epitope expressed by most clinical isolates, displays complement-dependent bactericidal activity and hastens clearance of gonococcal vaginal colonization in mice. Fc mutations in a human IgG1 chimeric version of MAb 2C7 further enhance complement activation, and the resulting MAb displays greater activity than wild-type MAb 2C7 in vivo Here, we utilized a DNA-encoded MAb (DMAb) construct designed to launch production and assembly of "complement-enhanced" chimeric MAb 2C7 in vivo The ensuing rapid and sustained MAb 2C7 expression attenuated gonococcal colonization in mice at 8 days as well as 65 days postadministration. The DMAb system may provide an effective, economical platform to deliver MAbs for durable protection against gonorrhea.
Assuntos
Anticorpos Antibacterianos/administração & dosagem , Anticorpos Monoclonais/administração & dosagem , Vacinas Bacterianas/imunologia , Epitopos/imunologia , Gonorreia/prevenção & controle , Imunização Passiva , Imunoglobulina G/administração & dosagem , Neisseria gonorrhoeae/imunologia , Animais , Anticorpos Antibacterianos/genética , Anticorpos Antibacterianos/imunologia , Anticorpos Monoclonais/genética , Anticorpos Monoclonais/imunologia , Antígenos de Bactérias/imunologia , Vacinas Bacterianas/administração & dosagem , Ativação do Complemento , Feminino , Gonorreia/imunologia , Imunoglobulina G/genética , Imunoglobulina G/imunologia , Camundongos , Camundongos Endogâmicos BALB CRESUMO
More than 100 million people have been infected with severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2). Common laboratory mice are not susceptible to wild-type SARS-CoV-2 infection, challenging the development and testing of effective interventions. Here, we describe the development and testing of a mouse model for SARS-CoV-2 infection based on transduction of the respiratory tract of laboratory mice with an adeno-associated virus vector (AAV6) expressing human ACE-2 (AAV6.2FF-hACE2). We validated this model using a previously described synthetic DNA vaccine plasmid, INO-4800 (pS). Intranasal instillation of AAV6.2FF-hACE2 resulted in robust hACE2 expression in the respiratory tract. pS induced robust cellular and humoral responses. Vaccinated animals were challenged with 105 TCID50 SARS-CoV-2 (hCoV-19/Canada/ON-VIDO-01/2020) and euthanized four days post-challenge to assess viral load. One immunization resulted in 50% protection and two immunizations were completely protective. Overall, the AAV6.2FF-hACE2 mouse transduction model represents an easily accessible, genetically diverse mouse model for wild-type SARS-CoV-2 infection and preclinical evaluation of potential interventions.
RESUMO
Coronavirus disease 2019 (COVID-19), caused by the SARS-CoV-2 virus, has had a dramatic global impact on public health and social and economic infrastructures. Here, we assess the immunogenicity and anamnestic protective efficacy in rhesus macaques of an intradermal (i.d.)-delivered SARS-CoV-2 spike DNA vaccine, INO-4800, currently being evaluated in clinical trials. Vaccination with INO-4800 induced T cell responses and induced spike antigen and RBD binding antibodies with ADCP and ADCD activity. Sera from the animals neutralized both the D614 and G614 SARS-CoV-2 pseudotype viruses. Several months after vaccination, animals were challenged with SARS-CoV-2 resulting in rapid recall of anti-SARS-CoV-2 spike protein T cell and neutralizing antibody responses. These responses were associated with lower viral loads in the lung. These studies support the immune impact of INO-4800 for inducing both humoral and cellular arms of the adaptive immune system, which are likely important for providing durable protection against COVID-19 disease.
Assuntos
Anticorpos Antivirais/sangue , Vacinas contra COVID-19/administração & dosagem , COVID-19/prevenção & controle , Pulmão/virologia , Linfócitos T/imunologia , Animais , Anticorpos Neutralizantes/sangue , Vacinas contra COVID-19/uso terapêutico , Feminino , Injeções Intradérmicas , Macaca mulatta , Masculino , SARS-CoV-2/imunologia , SARS-CoV-2/metabolismo , Glicoproteína da Espícula de Coronavírus/imunologia , Vacinas de DNA/administração & dosagem , Vacinas de DNA/uso terapêutico , Carga ViralRESUMO
BACKGROUND: Eradication of Plasmodium falciparum malaria will likely require a multivalent vaccine, but the development of a highly efficacious subunit-based formulation has been challenging. We previously showed that production and immunogenicity of two leading vaccine targets, PfMSP119 (blood-stage) and Pfs25 (sexual stage), could be enhanced upon genetic fusion to merozoite surface protein 8 (PfMSP8). Here, we sought to optimize a Pfs25-based formulation for use in combination with rPfMSP1/8 with the goal of maintaining the immunogenicity of each subunit. METHODS: Comparative mouse studies were conducted to assess the effects of adjuvant selection (Alhydrogel vs. glucopyranosyl lipid adjuvant-stable emulsion (GLA-SE)) and antigen dose (2.5 vs. 0.5 µg) on the induction of anti-Pfs25 immune responses. The antibody response (magnitude, IgG subclass profile, and transmission-reducing activity (TRA)) and cellular responses (proliferation, cytokine production) generated in response to each formulation were assessed. Similarly, immunogenicity of a bivalent vaccine containing rPfMSP1/8 and rPfs25/8 was evaluated. RESULTS: Alum-based formulations elicited strong and comparable humoral and cellular responses regardless of antigen form (unfused rPfs25 or chimeric rPfs25/8) or dose. In contrast, GLA-SE based formulations elicited differential responses as a function of both parameters, with 2.5 µg of rPfs25/8 inducing the highest titers of functional anti-Pfs25 antibodies. Based on these data, chimeric rPfs25/8 was selected and tested in a bivalent formulation with rPfMSP1/8. Strong antibody titers against Pfs25 and PfMSP119 domains were induced with GLA-SE based formulations, with no indication of antigenic competition. CONCLUSIONS: We were able to generate an immunogenic bivalent vaccine designed to target multiple parasite stages that could reduce both clinical disease and parasite transmission. The use of the same PfMSP8 carrier for two different vaccine components was effective in this bivalent formulation. As such, the incorporation of additional protective targets fused to the PfMSP8 carrier into the formulation should be feasible, further broadening the protective response.
Assuntos
Antígenos de Protozoários/imunologia , Vacinas Antimaláricas/imunologia , Malária Falciparum/prevenção & controle , Plasmodium falciparum/imunologia , Vacinas de Subunidades Antigênicas/imunologia , Animais , Anticorpos Antiprotozoários/imunologia , Formação de Anticorpos , Humanos , Malária Falciparum/imunologia , Masculino , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Proteínas de Protozoários/imunologia , Proteínas Recombinantes/imunologia , Linfócitos T/imunologiaRESUMO
The coronavirus family member, SARS-CoV-2 has been identified as the causal agent for the pandemic viral pneumonia disease, COVID-19. At this time, no vaccine is available to control further dissemination of the disease. We have previously engineered a synthetic DNA vaccine targeting the MERS coronavirus Spike (S) protein, the major surface antigen of coronaviruses, which is currently in clinical study. Here we build on this prior experience to generate a synthetic DNA-based vaccine candidate targeting SARS-CoV-2 S protein. The engineered construct, INO-4800, results in robust expression of the S protein in vitro. Following immunization of mice and guinea pigs with INO-4800 we measure antigen-specific T cell responses, functional antibodies which neutralize the SARS-CoV-2 infection and block Spike protein binding to the ACE2 receptor, and biodistribution of SARS-CoV-2 targeting antibodies to the lungs. This preliminary dataset identifies INO-4800 as a potential COVID-19 vaccine candidate, supporting further translational study.
Assuntos
Antígenos Virais/imunologia , Glicoproteína da Espícula de Coronavírus/imunologia , Vacinas de DNA/imunologia , Vacinas Virais/imunologia , Enzima de Conversão de Angiotensina 2 , Animais , Anticorpos Neutralizantes/imunologia , Antígenos Virais/química , Vacinas contra COVID-19 , Infecções por Coronavirus/imunologia , Infecções por Coronavirus/prevenção & controle , Mapeamento de Epitopos , Cobaias , Imunidade Humoral , Imunoglobulina G/imunologia , Pulmão/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Coronavírus da Síndrome Respiratória do Oriente Médio , Modelos Animais , Peptidil Dipeptidase A/metabolismo , Glicoproteína da Espícula de Coronavírus/química , Vacinas Virais/químicaRESUMO
We analyzed gene expression profiles of young and aged mouse CD8+ T cells specific for the nucleoprotein (NP) of influenza A/PR8/34 virus. CD8+ T cells were stimulated either by the NP antigen expressed in its native form or fused into the herpes virus (HSV)-1 glycoprotein D (gD) protein, which blocks signaling through the immunoinhibitory B and T lymphocyte attenuator (BTLA) and CD160 pathways. We show that NP-specific CD8+ T cells from aged mice exhibit numerous differences in gene expression compared to NP-specific CD8+ T cells from young mice, including a significant reduction of expression in genes involved in T cell receptor (TcR) and CD28 signaling. We also show that these changes can be reversed in a sub-population (~50%) of the aged mice by a BTLA/CD160 checkpoint blockade. These results suggest that BTLA/CD160 checkpoint blockade has potential value as a vaccine additive to induce better CD8+ T cell responses in the aged.
Assuntos
Envelhecimento/fisiologia , Antígenos CD/metabolismo , Linfócitos T CD8-Positivos/fisiologia , Regulação da Expressão Gênica/imunologia , Receptores Imunológicos/metabolismo , Transcriptoma/fisiologia , Animais , Antígenos CD/genética , Feminino , Proteínas Ligadas por GPI/genética , Proteínas Ligadas por GPI/metabolismo , Vírus da Influenza A , Vacinas contra Influenza/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Infecções por Orthomyxoviridae/prevenção & controle , Receptores Imunológicos/genética , VacinaçãoRESUMO
In humans adeno-associated virus (AAV)-mediated gene transfer is followed by expansion of AAV capsid-specific T cells, evidence of cell damage, and loss of transgene product expression, implicating immunological rejection of vector-transduced cells, which may be prevented by immunosuppressive drugs. We undertook this study to assess the effect of immunosuppression (IS) used for organ transplantation on immune responses to AAV capsid antigens. Recipients of liver or kidney transplants were tested before and 4 weeks after induction of IS in comparison with matched samples from healthy human adults and an additional cohort with comorbid conditions similar to those of the transplant patients. Our data show that transplant patients and comorbid control subjects have markedly higher frequencies of circulating AAV capsid-specific T cells compared with healthy adults. On average, IS resulted in a reduction of AAV-specific CD4⺠T cells, whereas numbers of circulating CD8⺠effector and central memory T cells tended to increase. Independent of the type of transplant or the IS regimens, the trend of AAV capsid-specific T cell responses after drug treatment varied; in some patients responses were unaffected whereas others showed decreases or even pronounced increases, casting doubt on the usefulness of prophylactic IS for AAV vector recipients.