RESUMO
Respiratory syncytial virus (RSV) is one of the most important etiologies of acute respiratory infections that cause bronchiolitis in children under 5 years of age. Treatments are expensive, no vaccine is available, and this is an important cause of hospitalization. Costimulatory molecules have been reported to be good inducers of antiviral type 1 immune response. This study aimed to generate a recombinant vaccine with the RSV F protein fused to 4-1BBL to evaluate the activation of an antiviral response in vitro and the production of neutralizing antibodies in vivo. The codon-optimized F gene was subcloned into an expression vector as follows: streptavidin core, gene F, and costimulatorytumor necrosis factor receptor superfamily member 9 -TNFRS9- ligand (4-1BBL). After the induction of expression in Escherichia coli C43, the recombinant protein (SA-F3x-4-1BBL, denominated SF4) was purified and verified by western blotting. Cultured RAW264.7 macrophages were stimulated with SF4 protein, then tumor necrosis factor (TNF) receptor-associated factor 2 (TRAF2), p38, and nuclear factor κ-light-chain-enhancer of activated B cells (NF-κB) were analyzed by western blot, and mRNA cytokines were analyzed by RT-qPCR. Finally, male C57BL/6 mice were inoculated with SF4, and the generation of anti-RSV neutralizing antibodies and serum cytokines was examined. SF4 had a size of 84.4 kDa with a 5.6% yield. SA-F-4-1BBL upregulated TRAF2, TNF-α, and interferon (IFN)-γ expression levels and activated p38 mitogen-activated protein kinase and NF-κΒ pathways in RAW264.7 cells. Importantly, antibodies capable of neutralizing RSV infection and producing type 1 cytokines were detected in the sera of immunized animals. These results suggest that the fusion protein SF4 activates the 4-1BBL signaling pathway, resulting in an effective antiviral response mediated by neutralizing antibodies and antiviral cytokines.
Assuntos
Anticorpos Neutralizantes , Anticorpos Antivirais , Infecções por Vírus Respiratório Sincicial , Vacinas contra Vírus Sincicial Respiratório , Vacinas Sintéticas , Proteínas Virais de Fusão , Animais , Proteínas Virais de Fusão/imunologia , Proteínas Virais de Fusão/genética , Camundongos , Infecções por Vírus Respiratório Sincicial/prevenção & controle , Infecções por Vírus Respiratório Sincicial/imunologia , Anticorpos Neutralizantes/sangue , Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/sangue , Anticorpos Antivirais/imunologia , Vacinas contra Vírus Sincicial Respiratório/imunologia , Vacinas contra Vírus Sincicial Respiratório/genética , Vacinas Sintéticas/imunologia , Vacinas Sintéticas/genética , Vacinas Sintéticas/administração & dosagem , Vírus Sincicial Respiratório Humano/imunologia , Vírus Sincicial Respiratório Humano/genética , Células RAW 264.7 , Camundongos Endogâmicos BALB C , Feminino , Citocinas/metabolismo , Macrófagos/imunologia , Escherichia coli/genética , Imunização , Proteínas Recombinantes de Fusão/imunologia , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes/imunologia , Proteínas Recombinantes/genéticaRESUMO
Antivenoms are the only effective treatment for snakebite envenomation and have saved countless lives over more than a century. Despite their value, antivenoms present risks of adverse reactions. Current formulations contain a fraction of nonspecific antibodies and serum proteins. While new promising candidates emerge as the next generation of antivenoms, it remains clear that animal-derived antivenoms will still play a critical role for years to come. In this study, we improved the bothropic antivenom (BAv), by capturing toxin-specific antibodies through affinity chromatography using immobilized Bothrops jararaca venom toxins. This process produced an improved antivenom (iBAv) enriched in neutralizing antibodies and depleted of serum proteins. Proteomic analysis showed that iBAv was 87% depleted in albumin and 37-83% lower in other serum proteins compared to BAv. Functional evaluation demonstrated that iBAv had a 2.9-fold higher affinity for venom toxins by surface plasmon resonance and a 2.8-fold lower ED50 in vivo, indicating enhanced potency. Our findings indicate that enriching specific antibodies while depleting serum proteins reduces the total protein dose required and increases the potency of antivenom. Although technical and economic considerations remain for large-scale implementation, this affinity-enriched antivenom represents a significant advancement in improving antivenom efficacy against B. jararaca envenomations.
Assuntos
Antivenenos , Bothrops , Venenos de Crotalídeos , Antivenenos/imunologia , Antivenenos/farmacologia , Animais , Venenos de Crotalídeos/imunologia , Proteômica/métodos , Cromatografia de Afinidade/métodos , Camundongos , Anticorpos Neutralizantes/imunologia , Mordeduras de Serpentes/tratamento farmacológico , Ressonância de Plasmônio de SuperfícieRESUMO
Yellow fever virus (YFV) circulates in a sylvatic cycle between non-human primates (NHPs) and arboreal mosquitoes in Brazil. Passive monitoring of ill or deceased NHPs is a key component of the Brazilian yellow fever (YF) surveillance program. Samples from NHPs carcasses are usually suitable for molecular tests but not for serological assays. As an alternative to the conventional plaque reduction neutralization test (PRNT) based on sera, we tested the utility of liver homogenates from experimentally infected (YFV, Mayaro virus [MAYV], chikungunya virus [CHIKV], or mock) mice to quantify PRNTs. Although homogenates from mock-infected mice showed a low level of nonspecific virus neutralization against all three viruses, homogenates from YFV-, MAYV- and CHIKV-infected mice demonstrated significantly higher levels of virus neutralization compared to controls. Receiver operating characteristic (ROC) curves analyses were performed using the median neutralization values of three technical replicates for each infected group separately or collectively. Results showed scores ≥0.97 (95% CI ≥ 0.89-1.0) for the area under the curve at dilutions 1:20 to 1:80, suggesting that median virus neutralization values effectively differentiated infected mice from controls. Liver homogenates obtained from 25 NHP carcasses (collected during the 2017 YF outbreak in Brazil) were also tested using the adapted PRNT as well as rapid lateral flow tests to investigate anti-YFV IgM. Neutralization activity was detected in six NHP samples that were also positive by PCR and anti-YFV IgM tests and one sample that tested negative by PCR and IgM test. Our results demonstrate the feasibility of using liver homogenates as an alternative approach for serological investigation in viral epidemiologic surveillance.
Assuntos
Anticorpos Neutralizantes , Anticorpos Antivirais , Fígado , Testes de Neutralização , Animais , Anticorpos Neutralizantes/imunologia , Anticorpos Neutralizantes/sangue , Fígado/virologia , Camundongos , Anticorpos Antivirais/sangue , Anticorpos Antivirais/imunologia , Brasil/epidemiologia , Vírus da Febre Amarela/imunologia , Vírus da Febre Amarela/isolamento & purificação , Feminino , Arbovírus/imunologia , Arbovírus/isolamento & purificação , Alphavirus/imunologia , Alphavirus/isolamento & purificação , Vírus Chikungunya/imunologiaRESUMO
Classical swine fever (CSF) is endemic in Cuba and is one of the major health problems of the Cuban swine industry. The current efforts to control the disease in Cuba include vaccination with Porvac®, a subunit marker vaccine. Although the efficacy of Porvac against CSF virus (CSFV) subgenotype 1.4 has been extensively documented, little is known about the ability of the antibodies induced by this vaccine to neutralize other CSFV genotypes. In this study, sera collected from three pigs vaccinated with Porvac were able to efficiently neutralize CSFV strains belonging to genotypes 1, 2, and 3. The findings from this study indicate that additional in vivo studies are warranted to confirm the ability of this vaccine to protect pigs against CSFV genotypes 2 and 3.
Assuntos
Anticorpos Neutralizantes , Anticorpos Antivirais , Vírus da Febre Suína Clássica , Peste Suína Clássica , Genótipo , Vacinas de Subunidades Antigênicas , Vacinas Virais , Animais , Vírus da Febre Suína Clássica/imunologia , Vírus da Febre Suína Clássica/genética , Suínos , Anticorpos Neutralizantes/imunologia , Anticorpos Neutralizantes/sangue , Peste Suína Clássica/prevenção & controle , Peste Suína Clássica/imunologia , Peste Suína Clássica/virologia , Vacinas de Subunidades Antigênicas/imunologia , Vacinas Virais/imunologia , Vacinas Virais/genética , Anticorpos Antivirais/imunologia , Anticorpos Antivirais/sangue , Cuba , VacinaçãoRESUMO
Studies have suggested an association between polymorphisms in class I genes of the major histocompatibility complex, specifically the human leukocyte antigen (HLA), and susceptibility to SARS-CoV-2. To explore this, 135 individuals with positive serological tests for SARS-CoV-2 were recruited. All the samples were collected before the advent of vaccines, avoiding immunization effects. Participants were divided into high and low neutralizing antibody titer groups, and polymorphisms in HLA-A, HLA-B, and HLA-DRB1 genes were examined using PCR-SSO. Allele prevalence in the study population was compared to the National Bone Marrow Volunteer Donors Register (REDOME) in São Paulo and between the high and low titer groups within the study population. Results indicated that the HLA-B*15 polymorphism was more prevalent in the COVID-19 positive group compared to the control population (COVID-19 = 0.1370; Control = 0.0875; p = 0.0067). The HLA-B*18 polymorphism was less prevalent in the COVID-19 group (COVID-19 = 0.0185; Control = 0.0534; p = 0.0064). Additionally, the HLA-A*30 polymorphism was more prevalent in the high titer group within the (high = 0.10937; low = 0.02816; p = 0.0125). Other polymorphisms showed no significant differences. These findings align with international studies, suggesting these genes plays a role in COVID-19 pathophysiology, however, further research is required to fully understand their impact.
Assuntos
COVID-19 , SARS-CoV-2 , Humanos , COVID-19/epidemiologia , COVID-19/imunologia , COVID-19/genética , COVID-19/virologia , Brasil/epidemiologia , Masculino , Feminino , Adulto , SARS-CoV-2/genética , SARS-CoV-2/imunologia , Pessoa de Meia-Idade , Predisposição Genética para Doença , Polimorfismo Genético , Frequência do Gene , Anticorpos Neutralizantes/sangue , Anticorpos Neutralizantes/imunologia , Alelos , Idoso , Antígeno HLA-B15/genética , Adulto Jovem , Anticorpos Antivirais/sangueRESUMO
The global impact of the SARS-CoV-2 (severe acute respiratory syndrome coronavirus 2) pandemic in 2019-2020 has led to significant changes in worldwide vaccination and immune prophylactic approaches. In this study, our research delves into a new immunization strategy that does not involve the use of additional adjuvants or preservatives, focusing on the effects of virus fusion with a bacterial nanostructure. The experimental procedures outlined in this paper involved the cultivation of SARS-CoV-2, the production, extraction, and nanocharacterization of outer membrane vesicles (OMV) from Neisseria meningitidis, immunization of mice with two doses of OMV combined with SARS-CoV-2, and the use of mesoporous silica SBa15 and SBa16 adsorbed to the same virus. The immune response was assessed through an indirect elisa method, analysis of cytokine expression profiles, and seroneutralization of the SARS-CoV-2 strain. The characterizations of associated OMV - SARS-CoV-2 and adsorption SBa15 and SBa16 were performed using Nanosight Tracking Analysis (NTA), which showed a high density of particles in the formulation. mice were then immunized, resulting in an immune response that produced high levels of neutralizing antibodies in IgG and IgG1 mouse immunoglobulins. In addition, expressions of IL-2, IL-4, and IL-23 in spleen cells were reinforced after the vaccination process. The comparative study of these three vaccine formulations has shown that the development of new vaccines for SARS-CoV-2 should take into consideration the production of neutralizing antibodies and the maintenance of immunological memory.
Assuntos
Vacinas contra COVID-19 , COVID-19 , Neisseria meningitidis , SARS-CoV-2 , Dióxido de Silício , Animais , COVID-19/prevenção & controle , COVID-19/imunologia , Vacinas contra COVID-19/imunologia , SARS-CoV-2/imunologia , Camundongos , Neisseria meningitidis/imunologia , Imunidade Humoral/imunologia , Imunidade Celular , Feminino , Camundongos Endogâmicos BALB C , Anticorpos Antivirais/sangue , Citocinas/imunologia , Citocinas/metabolismo , Anticorpos Neutralizantes/imunologia , Anticorpos Neutralizantes/sangue , VacinaçãoRESUMO
The high antigenic variability of the foot-and-mouth disease virus (FMDV) represents a challenge for developing prophylactic strategies, stressing the need for research into vaccines offering broad protection against a range of virus strains. Here, the heterotypic cross-reaction using different vaccine schemes against serotype O strains was studied, evaluating the impact of revaccination, antigen dose, and incorporation of additional FMDV serotypes. Naïve cattle were immunized with seven distinct FMDV vaccines, receiving three doses of the same formulation at 0, 28, and 56 days post-primary vaccination (dpv). Serum samples were collected up to 70 dpv and tested by a virus-neutralizing test against serotype O strains from a South American lineage and two strains representative of two Asian lineages. Our results showed that vaccines containing the ME-SA topotype O1/Campos strain developed cross-neutralizing responses against the two Asian viruses after the first vaccination. In contrast, significant heterotypic neutralizing antibody titers against the homologous topotype strain were only found after the second vaccination, indicating that the phylogenic relationship may differ from the antigenic profiles for these two viruses. The amount of the O1/Campos strain and the revaccination were essential factors for neutralization against the homologous- and heterologous-type O FMDV viruses. The strain composition of the vaccine was only relevant for cross-neutralization against one of the Asian strains, suggesting potential intra-serotypic divergences for this pattern.
Assuntos
Anticorpos Neutralizantes , Anticorpos Antivirais , Doenças dos Bovinos , Reações Cruzadas , Vírus da Febre Aftosa , Febre Aftosa , Sorogrupo , Vacinação , Vacinas Virais , Animais , Bovinos , Vírus da Febre Aftosa/imunologia , Vírus da Febre Aftosa/classificação , Febre Aftosa/prevenção & controle , Febre Aftosa/imunologia , Febre Aftosa/virologia , Anticorpos Neutralizantes/imunologia , Anticorpos Neutralizantes/sangue , Vacinas Virais/imunologia , Anticorpos Antivirais/sangue , Anticorpos Antivirais/imunologia , Doenças dos Bovinos/prevenção & controle , Doenças dos Bovinos/virologia , Doenças dos Bovinos/imunologia , Reações Cruzadas/imunologia , Vacinação/veterinária , Testes de Neutralização , Proteção Cruzada/imunologia , FilogeniaRESUMO
Conventional live virus research on severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causal agent of coronavirus disease-19 (COVID-19), requires Biosafety Level 3 (BSL-3) facilities. SARS-CoV-2 pseudotyped viruses have emerged as valuable tools in virology, mimicking the entry process of the SARS-CoV-2 virus into human cells by expressing its spike glycoprotein in a surrogate system using recombinant plasmids. One significant application of this tool is in functional assays for the evaluation of neutralizing antibodies. Pseudotyped viruses have the advantage of being competent for only a single cycle of infection, providing better safety and versatility and allowing them to be studied in BSL-2 laboratories. Here, we describe three protocols for the detection of SARS-CoV-2 neutralizing antibodies through a pseudotyped virus assay. First, SARS-CoV-2 S pseudotyped viruses (PV SARS-CoV-2 S) are produced using a Moloney murine leukemia virus (MuLV) three-plasmid system. The plasmids are designed to express the GagPol packing proteins, enhanced green fluorescent protein (eGFP) as a readout system, and the SARS-CoV-2 S protein modified to remove the endoplasmic reticulum retention domain and to improve infection. Next, the internalization of PV SARS-CoV-2 S protein in human embryonic kidney 293T (HEK-293T) cells overexpressing angiotensin-converting enzyme 2 (HEK-293T-ACE2) is confirmed by fluorescence microscopy and quantified using flow cytometry. Finally, PV SARS-CoV-2 S is used to screen neutralizing antibodies in serum samples from convalescent COVID-19 patients; it can also be used for studying the cell entry mechanisms of different SARS-CoV-2 variants, evaluating antiviral agents, and designing vaccines. © 2024 Wiley Periodicals LLC. Basic Protocol 1: Generation of PV SARS-CoV-2 S pseudotyped virus Basic Protocol 2: Assay of PV SARS-CoV-2 S internalization in target cells. Basic Protocol 3: Detection of neutralizing antibodies in serum samples.
Assuntos
Anticorpos Neutralizantes , Anticorpos Antivirais , COVID-19 , SARS-CoV-2 , Humanos , Anticorpos Neutralizantes/imunologia , Anticorpos Neutralizantes/sangue , SARS-CoV-2/imunologia , Anticorpos Antivirais/imunologia , Anticorpos Antivirais/sangue , COVID-19/virologia , COVID-19/imunologia , COVID-19/diagnóstico , COVID-19/sangue , Testes de Neutralização/métodos , Células HEK293 , Pseudotipagem Viral , Glicoproteína da Espícula de Coronavírus/imunologia , Glicoproteína da Espícula de Coronavírus/genéticaRESUMO
Understanding the immune response generated by SARS-CoV-2 is critical for assessing efficient therapeutic protocols and gaining insights into the durability of protective immunity. The current work was aimed at studying the specific humoral responses against SARS-CoV-2 in Cuban COVID-19 convalescents. We developed suitable tools and methods based on ELISA methodology, for supporting this evaluation. Here, we describe the development of an ELISA for the quantification of anti-RBD IgG titers in a large number of samples and a similar test in the presence of NH4SCN as chaotropic agent for estimating the RBD specific antibody avidity. Additionally, a simple and rapid ELISA based on antibody-mediated blockage of the binding RBD-ACE2 was implemented for detecting, as a surrogate of conventional test, the levels of anti-RBD inhibitory antibodies in convalescent sera. In a cohort of 273 unvaccinated convalescents, we identified higher anti-RBD IgG titer (1 : 1,330, p < 0.0001) and higher levels of inhibitory antibodies blocking RBD-ACE2 binding (1 : 216, p < 0.05) among those who had recovered from severe illness. Our results suggest that disease severity, and not demographic features such as age, sex, and skin color, is the main determinant of the magnitude and neutralizing ability of the anti-RBD antibody response. An additional paired longitudinal assessment in 14 symptomatic convalescents revealed a decline in the antiviral antibody response and the persistence of neutralizing antibodies for at least 4 months after the onset of symptoms. Overall, SARS-CoV-2 infection elicits different levels of antibody response according to disease severity that declines over time and can be monitored using our homemade serological assays.
Assuntos
Anticorpos Neutralizantes , Anticorpos Antivirais , COVID-19 , Ensaio de Imunoadsorção Enzimática , Imunidade Humoral , Imunoglobulina G , SARS-CoV-2 , Humanos , COVID-19/imunologia , SARS-CoV-2/imunologia , Anticorpos Antivirais/sangue , Anticorpos Antivirais/imunologia , Cuba , Masculino , Feminino , Imunoglobulina G/sangue , Imunoglobulina G/imunologia , Pessoa de Meia-Idade , Adulto , Anticorpos Neutralizantes/imunologia , Anticorpos Neutralizantes/sangue , Glicoproteína da Espícula de Coronavírus/imunologia , Idoso , Enzima de Conversão de Angiotensina 2/metabolismo , Afinidade de Anticorpos/imunologiaRESUMO
The emergence of new SARS-CoV-2 variants of concern associated with waning immunity induced by natural infection or vaccines currently in use suggests that the COVID-19 pandemic will become endemic. Investing in new booster vaccines using different platforms is a promising way to enhance protection and keep the disease under control. Here, we evaluated the immunogenicity, efficacy, and safety of the SpiN-Tec vaccine, based on a chimeric recombinant protein (SpiN) adjuvanted with CTVad1 (MF59-based adjuvant), aiming at boosting immunity against variants of concern of SARS-CoV-2. Immunization of K18-hACE-2 transgenic mice and hamsters induced high antibody titers and cellular immune response to the SpiN protein as well as to its components, RBD and N proteins. Importantly in a heterologous prime/boost protocol with a COVID-19 vaccine approved for emergency use (ChAdOx1), SpiN-Tec enhanced the level of circulation neutralizing antibodies (nAb). In addition to protection against the Wuhan isolate, protection against the Delta and Omicron variants was also observed as shown by reduced viral load and lung pathology. Toxicity and safety tests performed in rats demonstrated that the SpiN-Tec vaccine was safe and, based on these results, the SpiN-Tec phase I/II clinical trial was approved.
Assuntos
Anticorpos Neutralizantes , Anticorpos Antivirais , Vacinas contra COVID-19 , COVID-19 , SARS-CoV-2 , Vacinas Sintéticas , Animais , Vacinas contra COVID-19/imunologia , SARS-CoV-2/imunologia , COVID-19/prevenção & controle , COVID-19/imunologia , Anticorpos Antivirais/sangue , Anticorpos Antivirais/imunologia , Anticorpos Neutralizantes/sangue , Anticorpos Neutralizantes/imunologia , Camundongos , Vacinas Sintéticas/imunologia , Vacinas Sintéticas/administração & dosagem , Cricetinae , Camundongos Transgênicos , Glicoproteína da Espícula de Coronavírus/imunologia , Glicoproteína da Espícula de Coronavírus/genética , Linfócitos T/imunologia , Feminino , Imunogenicidade da Vacina , Humanos , Ratos , Modelos Animais de Doenças , Adjuvantes de Vacinas , Imunização Secundária , Eficácia de VacinasRESUMO
Scorpionism is Brazil's most prevalent envenomation. Treatment typically involves the use of heterologous antivenoms derived from the immunization of horses with crude T. serrulatus venom (TsV). Due to the high toxicity of this immunogen, as well as the particular challenges associated with the use of venoms as antigens, there is interest in exploring new alternatives for reducing their use in antivenom production. In this study, ten linear B-cell epitopes from hyaluronidase and voltage-gated sodium channel toxins, previously identified using the SPOT-synthesis method, were selected to construct a recombinant chimeric MultiEpitopic Protein from T. serrulatus scorpion venom (TsMEP). We demonstrated that TsMEP is non-toxic, and antibodies elicited in rabbits against this antigen exhibited reactivity in ELISA with Brazilian T. serrulatus, T. bahiensis, T. obscurus, and T. stigmurus venoms, as well as with Peruvian Hadruroides lunatus and North African Androctonus australis hector scorpion venoms. In vivo and in vitro neutralization assays revealed that rabbit anti-TsMEP antibodies partially neutralize the hyaluronidase activity of T. serrulatus venom and its lethality. Data presented here suggests that this multiepitopic protein could be a promising candidate in experimental immunization approaches for antivenom production for clinical use against Tityus spp. venoms.
Assuntos
Antivenenos , Hialuronoglucosaminidase , Proteínas Recombinantes , Venenos de Escorpião , Escorpiões , Animais , Antivenenos/imunologia , Antivenenos/farmacologia , Antivenenos/biossíntese , Antivenenos/química , Venenos de Escorpião/imunologia , Venenos de Escorpião/química , Coelhos , Proteínas Recombinantes/imunologia , Proteínas Recombinantes/farmacologia , Hialuronoglucosaminidase/imunologia , Epitopos de Linfócito B/imunologia , Antígenos/imunologia , Testes de Neutralização , Camundongos , Anticorpos Neutralizantes/imunologia , Animais PeçonhentosRESUMO
Vaccination has played a critical role in mitigating COVID-19. Despite the availability of licensed vaccines, there remains a pressing need for improved vaccine platforms that provide high protection, safety, and versatility, while also reducing vaccine costs. In response to these challenges, our aim is to create a self-adjuvanted vaccine against SARS-CoV-2, utilizing Virus-Like Particles (VLPs) as the foundation. To achieve this, we produced bacteriophage (Qß) VLPs in a prokaryotic system and purified them using a rapid and cost-effective strategy involving organic solvents. This method aims to solubilize lipids and components of the cell membrane to eliminate endotoxins present in bacterial samples. For vaccine formulation, Receptor Binding Domain (RBD) antigens were conjugated using chemical crosslinkers, a process compatible with Good Manufacturing Practice (GMP) standards. Transmission Electron Microscopy (TEM) confirmed the expected folding and spatial configuration of the QßVLPs vaccine. Additionally, vaccine formulation assessment involved SDS-PAGE stained with Coomassie Brilliant Blue, Western blotting, and stereomicroscopic experiments. In vitro and in vivo evaluations of the vaccine formulation were conducted to assess its capacity to induce a protective immune response without causing side effects. Vaccine doses of 20 µg and 50 µg stimulated the production of neutralizing antibodies. In in vivo testing, the group of animals vaccinated with 50 µg of vaccine formulation provided complete protection against virus infection, maintaining stable body weight without showing signs of disease. In conclusion, the QßVLPs-RBD vaccine has proven to be effective and safe, eliminating the necessity for supplementary adjuvants and offering a financially feasible approach. Moreover, this vaccine platform demonstrates flexibility in targeting Variants of Concern (VOCs) via established conjugation protocols with VLPs.
Assuntos
Vacinas contra COVID-19 , COVID-19 , SARS-CoV-2 , Vacinas de Partículas Semelhantes a Vírus , Vacinas de Partículas Semelhantes a Vírus/imunologia , Vacinas de Partículas Semelhantes a Vírus/administração & dosagem , Vacinas contra COVID-19/imunologia , Vacinas contra COVID-19/administração & dosagem , Animais , COVID-19/prevenção & controle , SARS-CoV-2/imunologia , Camundongos , Humanos , Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/imunologia , Glicoproteína da Espícula de Coronavírus/imunologia , Feminino , Adjuvantes Imunológicos/administração & dosagem , Adjuvantes de Vacinas , Vacinação/métodos , Camundongos Endogâmicos BALB CRESUMO
Chikungunya fever is a re-emerging mosquito-borne disease caused by the chikungunya virus (CHIKV) and produces acute arthritis that can progress to chronic disease with arthralgia. The first approved live-attenuated chikungunya vaccine has only recently become available for use in humans in the USA, but the access in endemic regions remains unmet. Here, we exploited the baculovirus display technology to develop a vectored vaccine candidate that exposes the CHIKV membrane proteins E1 and E2 on the baculovirus surface. Using recombinant baculovirus as vector vaccines has both productive and regulatory advantages: they are safe for handling and easy to produce in high titers and are non-pathogenic and non-replicative in mammals but have strong adjuvant properties by inducing humoral and cellular immune responses. CHIKV E1 and E2 envelope proteins with their own signal and transmembrane sequences were expressed on the surface of budded baculovirus virions. Immunization of C57BL/6 mice with non-adjuvanted recombinant baculovirus induced IgG antibodies against E2 with a predominant IgG2c subtype, neutralizing antibodies and a specific IFN-γ CD8+ T-cell response. Immunization with a second dose significantly boosted the antibody response, and mice immunized with two doses of the vaccine candidate were completely protected against challenge with CHIKV showing no detectable viremia or signs of disease. Altogether, baculovirus display of CHIKV envelope proteins served as an efficient vaccine platform against CHIKV.IMPORTANCEThe global spread of chikungunya virus (CHIKV) has disproportionately impacted the Americas that experienced a fourfold increase in 2023 in cases and deaths compared with the same period in 2022. The disease is characterized by acute fever and debilitating joint pain that can become chronic. Despite the socioeconomic burden related to the high morbidity rates of CHIKV infection, a vaccine for CHIKV is currently approved only in the USA. Vaccines are the most effective preventive measure against viral diseases, and advances in the development of different vaccine platforms such as nucleic acids and viral vectors have prompted the rapid deployment of vaccines to contain the COVID-19 pandemic. Here, we report the use of baculovirus display as a strategy for the design of a novel vaccine that provides sterilizing immunity in a mouse model of chikungunya disease. Our results encourage further research regarding the potential of baculovirus as platforms for human vaccine design.
Assuntos
Anticorpos Neutralizantes , Anticorpos Antivirais , Baculoviridae , Febre de Chikungunya , Vírus Chikungunya , Camundongos Endogâmicos C57BL , Proteínas do Envelope Viral , Vacinas Virais , Animais , Vírus Chikungunya/imunologia , Vírus Chikungunya/genética , Proteínas do Envelope Viral/imunologia , Proteínas do Envelope Viral/genética , Camundongos , Baculoviridae/genética , Vacinas Virais/imunologia , Vacinas Virais/genética , Febre de Chikungunya/prevenção & controle , Febre de Chikungunya/imunologia , Anticorpos Antivirais/sangue , Anticorpos Antivirais/imunologia , Anticorpos Neutralizantes/sangue , Anticorpos Neutralizantes/imunologia , Feminino , Humanos , Imunoglobulina G/sangue , Imunidade Celular , Vacinas Sintéticas/imunologia , Vacinas Sintéticas/administração & dosagemRESUMO
The COVID-19 pandemic has overwhelmed healthcare systems and triggered global economic downturns. While vaccines have reduced the lethality rate of SARS-CoV-2 to 0.9% as of October 2024, the continuous evolution of variants remains a significant public health challenge. Next-generation medical therapies offer hope in addressing this threat, especially for immunocompromised individuals who experience prolonged infections and severe illnesses, contributing to viral evolution. These cases increase the risk of new variants emerging. This study explores miniACE2 decoys as a novel strategy to counteract SARS-CoV-2 variants. Using in silico design and molecular dynamics, blocking proteins (BPs) were developed with stronger binding affinity for the receptor-binding domain of multiple variants than naturally soluble human ACE2. The BPs were expressed in E. coli and tested in vitro, showing promising neutralizing effects. Notably, miniACE2 BP9 exhibited an average IC50 of 4.9 µg/mL across several variants, including the Wuhan strain, Mu, Omicron BA.1, and BA.2 This low IC50 demonstrates the potent neutralizing ability of BP9, indicating its efficacy at low concentrations.Based on these findings, BP9 has emerged as a promising therapeutic candidate for combating SARS-CoV-2 and its evolving variants, thereby positioning it as a potential emergency biopharmaceutical.
Assuntos
Enzima de Conversão de Angiotensina 2 , Anticorpos Neutralizantes , COVID-19 , Simulação de Dinâmica Molecular , SARS-CoV-2 , Glicoproteína da Espícula de Coronavírus , SARS-CoV-2/efeitos dos fármacos , SARS-CoV-2/imunologia , Humanos , COVID-19/virologia , COVID-19/imunologia , Enzima de Conversão de Angiotensina 2/metabolismo , Enzima de Conversão de Angiotensina 2/química , Anticorpos Neutralizantes/imunologia , Glicoproteína da Espícula de Coronavírus/metabolismo , Glicoproteína da Espícula de Coronavírus/química , Glicoproteína da Espícula de Coronavírus/imunologia , Simulação por Computador , Pandemias , Ligação Proteica , Betacoronavirus/imunologia , Betacoronavirus/efeitos dos fármacos , Testes de NeutralizaçãoRESUMO
Antibodies are an essential component of the antiviral response in many species, but to date, there is no compelling evidence that bats are capable of eliciting a robust humoral immunity, including neutralizing antibodies. Here, we report that infection of Jamaican fruit bats with the bat influenza A virus H18N11 elicits a rapid and stable humoral immune response with a strong neutralizing capacity, associated with no detectable viral shedding after repeat challenge infection. Thus, the neutralizing antibody response of bats might play an important role in the bat immunity.
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Anticorpos Neutralizantes , Anticorpos Antivirais , Quirópteros , Infecções por Orthomyxoviridae , Quirópteros/virologia , Quirópteros/imunologia , Animais , Anticorpos Neutralizantes/imunologia , Infecções por Orthomyxoviridae/imunologia , Infecções por Orthomyxoviridae/virologia , Infecções por Orthomyxoviridae/veterinária , Anticorpos Antivirais/imunologia , Vírus da Influenza A/imunologia , Eliminação de Partículas Virais/imunologiaRESUMO
The COVID-19 pandemic was characterized by the emergence and succession of SARS-CoV-2 variants able to evade the antibody response induced by natural infection and vaccination. To evaluate the IgG reactivity and neutralizing capacity of the serum of individuals vaccinated with Sputnik V (105 volunteers vaccinated) against different viral variants. IgG reactivity to the Spike protein (S) was evaluated by ELISA. A plaque reduction neutralization test was performed using different viral variant isolates. At 42 days post-vaccination, the frequency of recognition and reactivity to the S protein of the Omicron variant was lower compared to that of the other variants. In general, a higher average neutralization titer was seen against the ancestral variant compared to the variants, especially Omicron. However, some sera exhibited a higher neutralization titer to the Gamma variant compared to the ancestral variant, suggesting unapparent exposure during the clinical trial. Antibodies induced by Sputnik V can recognize, persist, and neutralize SARS-CoV-2 variants, with Omicron being the one that best evades this response. These results represent a unique report on the humoral response induced by a globally lesser-studied vaccine in terms of efficacy and immune escape, offering insights into developing vaccines targeting unknown coronaviruses.
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Anticorpos Neutralizantes , Anticorpos Antivirais , COVID-19 , Imunoglobulina G , Testes de Neutralização , SARS-CoV-2 , Glicoproteína da Espícula de Coronavírus , Humanos , SARS-CoV-2/imunologia , SARS-CoV-2/genética , Anticorpos Antivirais/sangue , Anticorpos Antivirais/imunologia , COVID-19/imunologia , COVID-19/prevenção & controle , COVID-19/virologia , COVID-19/epidemiologia , Anticorpos Neutralizantes/imunologia , Anticorpos Neutralizantes/sangue , Imunoglobulina G/sangue , Imunoglobulina G/imunologia , Glicoproteína da Espícula de Coronavírus/imunologia , Glicoproteína da Espícula de Coronavírus/genética , Venezuela/epidemiologia , Vacinas contra COVID-19/imunologia , Vacinas contra COVID-19/administração & dosagem , Adulto , Feminino , Masculino , Vacinação , Pessoa de Meia-IdadeRESUMO
To achieve global herd immunity, widespread vaccination is the most effective strategy. Vaccines stimulate the immune system, generating cytokines and chemokines, isotype antibodies, and neutralizing antibodies; all these molecules collectively provide a more comprehensive characterization of the immune response post-vaccination. We conducted a longitudinal study in northwestern Mexico, involving 120 individuals before vaccination and after the first dose of the SARS-CoV-2 vaccine, and 46 individuals after their second dose. Our findings reveal that antibody levels stabilize over time; cytokine levels generally increase following the first dose but decrease after the second dose and higher than normal levels in IgG1 and IgG3 concentrations are present. Most of the innate cytokines determined in this study were higher after the first dose of the vaccine. Regardless of previous infection history, this finding suggests that the first dose of the vaccine is crucial and may stimulate immunity by enhancing the innate immune response. Conversely, increased levels of IL-4, indicative of a Th2 response, were found in individuals without prior exposure to the virus and in those vaccinated with CoronaVac. These results suggest that the immune response to COVID-19 vaccines is multi-faceted, with preexisting immunity potentiating a more robust innate response. Vaccine type plays a critical role, with genetic vaccines favoring a Th1 response and inactivated vaccines like CoronaVac skewing toward a Th2 profile.
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Anticorpos Neutralizantes , Anticorpos Antivirais , Vacina BNT162 , Vacinas contra COVID-19 , COVID-19 , ChAdOx1 nCoV-19 , Citocinas , SARS-CoV-2 , Humanos , COVID-19/imunologia , COVID-19/prevenção & controle , Anticorpos Antivirais/sangue , Anticorpos Antivirais/imunologia , Masculino , Vacinas contra COVID-19/imunologia , Vacinas contra COVID-19/administração & dosagem , Citocinas/imunologia , Feminino , Adulto , Pessoa de Meia-Idade , Anticorpos Neutralizantes/sangue , Anticorpos Neutralizantes/imunologia , Vacina BNT162/imunologia , Vacina BNT162/administração & dosagem , México , Estudos Longitudinais , ChAdOx1 nCoV-19/imunologia , ChAdOx1 nCoV-19/administração & dosagem , SARS-CoV-2/imunologia , Células Th2/imunologia , Células Th1/imunologia , Imunoglobulina G/sangue , Vacinação , Vacinas de Produtos Inativados/imunologia , Vacinas de Produtos Inativados/administração & dosagem , Adulto Jovem , IdosoRESUMO
BACKGROUND: The immunological response to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection and immunisation is variable. OBJECTIVES: To describe the humoral immune response by correlating IgA and IgG antibodies with NAbs titration following CoronaVac® immunisation and an mRNA (Comirnaty®) booster among healthcare workers (HCWs) and to compare the cytokine and interleukin profiles between HCWs vaccinated with CoronaVac and coronavirus disease 2019 (COVID-19) infected patients. METHODS: Samples from 133 HCWs collected at 20 (T1) and 90 (T2) days after CoronaVac immunisation and 15 (T3) days after a booster dose with the Comirnaty vaccine were analysed for IgA and IgG EIA and neutralisation assay. Cytokine levels from vaccinated individuals at T1 day and COVID-19 patients were compared. FINDINGS: Neutralising antibodies (NAbs) were observed in 81.7% of participants at T1, but only 49.2% maintained detectable NAbs after 90 days. The booster dose increased NAbs response in all participants. The cytokines with the highest levels post-vaccination were IL-6 and MCP-1. The MCP-1, IL-18, and IFN- γ levels were higher in COVID-19 patients than in vaccinated HCWs, while IL-22 levels increased in the vaccinated HCWs group. MAIN CONCLUSIONS: The neutralisation titres in the T2 samples decreased, and antibody levels detected at T2 showed a more significant reduction than the neutralisation. The higher IL-22 expression in immunised individuals compared to those with COVID-19 suggests that IL-22 may be beneficial in protecting against severe disease.
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Anticorpos Neutralizantes , Anticorpos Antivirais , Vacinas contra COVID-19 , COVID-19 , Citocinas , Pessoal de Saúde , Imunização Secundária , Imunoglobulina G , SARS-CoV-2 , Humanos , COVID-19/imunologia , COVID-19/prevenção & controle , Masculino , Feminino , Anticorpos Antivirais/sangue , Adulto , Anticorpos Neutralizantes/sangue , Anticorpos Neutralizantes/imunologia , SARS-CoV-2/imunologia , Vacinas contra COVID-19/imunologia , Vacinas contra COVID-19/administração & dosagem , Imunoglobulina G/sangue , Imunoglobulina G/imunologia , Pessoa de Meia-Idade , Citocinas/imunologia , Citocinas/sangue , Imunoglobulina A/sangue , Imunoglobulina A/imunologia , Imunoglobulina A/análise , Vacinação , Adulto Jovem , Imunidade Humoral/imunologia , Vacinas de Produtos InativadosRESUMO
Introduction: There are no reports in LATAM related to longitudinal humoral and cellular response to adenovirus based COVID-19 vaccines in people with Multiple Sclerosis (pwMS) under different disease modifying therapies (DMTs) and neutralization of the Omicron and Wuhan variants of SARS-COV-2. Methods: IgG anti- SARS-COV-2 spike titer were measured in a cohort of 101 pwMS under fingolimod, dimethyl fumarate, cladribine and antiCD20, as well as 28 healthy controls (HC) were measured 6 weeks after vaccination with 2nd dose (Sputnik V or AZD1222) and 3nd dose (homologous or heterologous schedule). Neutralizing capacity was against Omicron (BA.1) and Wuhan (D614G) variants and pseudotyped particles and Cellular response were analyzed. Results: Multivariate regression analysis showed anti-cd20 (ß= -,349, 95% CI: -3655.6 - -369.01, p=0.017) and fingolimod (ß=-,399, 95% CI: -3363.8 - -250.9, p=0.023) treatments as an independent factor associated with low antibody response (r2 adjusted=0.157). After the 2nd dose we found a correlation between total and neutralizing titers against D614G (rho=0.6; p<0.001; slope 0.8, 95%CI:0.4-1.3), with no differences between DMTs. Neutralization capacity was lower for BA.1 (slope 0.3, 95%CI:0.1-0.4). After the 3rd dose, neutralization of BA.1 improved (slope: 0.9 95%CI:0.6-1.2), without differences between DMTs. A fraction of pwMS generated anti-Spike CD4+ and CD8+ T cell response. In contrast, pwMS under antiCD20 generated CD8+TNF+IL2+ response without differences with HC, even in the absence of humoral response. The 3rd dose significantly increased the neutralization against the Omicron, as observed in the immunocompetent population. Discussion: Findings regarding humoral and cellular response are consistent with previous reports.
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Anticorpos Neutralizantes , Anticorpos Antivirais , Vacinas contra COVID-19 , COVID-19 , Imunossupressores , Esclerose Múltipla , SARS-CoV-2 , Humanos , Masculino , Feminino , Imunossupressores/uso terapêutico , Vacinas contra COVID-19/imunologia , Vacinas contra COVID-19/administração & dosagem , SARS-CoV-2/imunologia , Pessoa de Meia-Idade , Esclerose Múltipla/imunologia , Esclerose Múltipla/tratamento farmacológico , COVID-19/imunologia , COVID-19/prevenção & controle , Adulto , Anticorpos Antivirais/sangue , Anticorpos Antivirais/imunologia , Anticorpos Neutralizantes/imunologia , Anticorpos Neutralizantes/sangue , Argentina , Adenoviridae/genética , Adenoviridae/imunologia , Imunidade Humoral , Glicoproteína da Espícula de Coronavírus/imunologiaRESUMO
Multivalent live-attenuated or inactivated vaccines are often used to control the bovine viral diarrhea disease (BVD). Still, they retain inherent disadvantages and do not provide the expected protection. This study developed a new vaccine prototype, including the external segment of the E2 viral protein from five different subgenotypes selected after a massive screening. The E2 proteins of every subgenotype (1aE2, 1bE2, 1cE2, 1dE2, and 1eE2) were produced in mammalian cells and purified by IMAC. An equimolar mixture of E2 proteins formulated in an oil-in-water adjuvant made up the vaccine candidate, inducing a high humoral response at 50, 100, and 150 µg doses in sheep. A similar immune response was observed in bovines at 50 µg. The cellular response showed a significant increase in the transcript levels of relevant Th1 cytokines, while those corresponding to the Th2 cytokine IL-4 and the negative control were similar. High levels of neutralizing antibodies against the subgenotype BVDV1a demonstrated the effectiveness of our vaccine candidate, similar to that observed in the sera of animals vaccinated with the commercial vaccine. These results suggest that our vaccine prototype could become an effective recombinant vaccine against the BVD.