Development of an in vitro Plasmodium parasite killing assay for the evaluation of cell-mediated immune responses following vaccination with pre-erythrocytic malaria vaccine candidates

ABSTRACT Immunogenicity testing of Plasmodium falciparum antigens being considered as malaria vaccine candidates was undertaken in rabbits. The antigens compared were recombinant baculovirus MSP-119 and five Pichia pastoris candidates, including two versions of MSP-119, AMA-1 (domains I and II), AMA-1+MSP-119, and fused AMA-1/MSP-119). Animals were immunized with equimolar amounts of each antigen, formulated in Montanide ISA720. The specificities and titers of antibodies were compared using immunofluorescence assays and enzyme-linked immunosorbent assay (ELISA). The antiparasite activity of immunoglobulin G (IgG) in in vitro cultures was determined by growth inhibition assay, flow cytometry, lactate dehydrogenase assay, and microscopy. Baculovirus MSP-119 immunizations produced the highest parasite-specific antibody titers in immunofluorescence assays. In ELISAs, baculovirus-produced MSP-119 induced more antibodies than any other single MSP-119 immunogen and three times more MSP-119 specific antibodies than the AMA-1/MSP-119 fusion. Antibodies induced by baculovirus MSP-119 gave the highest levels of growth inhibition in HB3 and 3D7 parasite cultures, followed by AMA-1+MSP-119 and the AMA-1/MSP-119 fusion. With the FCR3 isolate (homologous to the AMA-1 construct), antibodies to the three AMA-1-containing candidates gave the highest levels of growth inhibition at high IgG concentrations, but antibodies to baculovirus MSP-119 inhibited as well or better at lower IgG concentrations. The two P. pastoris-produced MSP-119-induced IgGs conferred the lowest growth inhibition. Comparative analysis of immunogenicity of vaccine antigens can be used to prioritize candidates before moving to expensive GMP production and clinical testing. The assays used have given discriminating readouts but it is not known whether any of them accurately reflect clinical protection.

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Chemical Attenuation in the Development of a Whole-Organism Malaria Vaccine

Infection and Immunity ◽

10.1128/iai.00062-17 ◽

2017 ◽

Vol 85 (7) ◽

Cited By ~ 8

Author(s):

Amber I. Raja ◽

Danielle I. Stanisic ◽

Michael F. Good

Keyword(s):

Subunit Vaccine ◽

Malaria Parasite ◽

Malaria Vaccine ◽

Vaccine Development ◽

Liver Stage ◽

Content Type ◽

Vaccine Candidates ◽

Limited Efficacy

ABSTRACT Malaria vaccine development has been dominated by the subunit approach; however, many subunit vaccine candidates have had limited efficacy in settings of malaria endemicity. As our search for an efficacious malaria vaccine continues, the development of a whole-organism vaccine is now receiving much scrutiny. One strategy currently being explored in the development of a whole-organism vaccine involves chemical attenuation of the malaria parasite. In vivo and in vitro chemical attenuation of both liver-stage and blood-stage Plasmodium parasites has been investigated. Here, we discuss both approaches of chemical attenuation in the development of a whole-organism vaccine against malaria.

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Reduced Protective Efficacy of a Blood-Stage Malaria Vaccine by Concurrent Nematode Infection

Infection and Immunity ◽

10.1128/iai.74.4.2138-2144.2006 ◽

2006 ◽

Vol 74 (4) ◽

pp. 2138-2144 ◽

Cited By ~ 62

Author(s):

Zhong Su ◽

Mariela Segura ◽

Mary M. Stevenson

Keyword(s):

Immune Responses ◽

Malaria Vaccine ◽

Transforming Growth Factor ◽

Protective Efficacy ◽

Nematode Infection ◽

Blood Stage ◽

Interleukin 4 ◽

Spleen Cells ◽

The Impact

ABSTRACT Helminth infections, which are prevalent in areas where malaria is endemic, have been shown to modulate immune responses to unrelated pathogens and have been implicated in poor efficacy of malaria vaccines in humans. We established a murine coinfection model involving blood-stage Plasmodium chabaudi AS malaria and a gastrointestinal nematode, Heligmosomoides polygyrus, to investigate the impact of nematode infection on the protective efficacy of a malaria vaccine. C57BL/6 mice immunized with crude blood-stage P. chabaudi AS antigen in TiterMax adjuvant developed strong protection against malaria challenge. The same immunization protocol failed to induce strong protection in H. polygyrus-infected mice. Immunized nematode-infected mice produced significantly lower levels of malaria-specific antibody than nematode-free mice produced. In response to nematode and malarial antigens, spleen cells from immunized nematode-infected mice produced significantly lower levels of gamma interferon but more interleukin-4 (IL-4), IL-13, and IL-10 in vitro than spleen cells from immunized nematode-free mice produced. Furthermore, H. polygyrus infection also induced a strong transforming growth factor β1 response in vivo and in vitro. Deworming treatment of H. polygyrus-infected mice before antimalarial immunization, but not deworming treatment after antimalarial immunization, restored the protective immunity to malaria challenge. These results demonstrate that concurrent nematode infection strongly modulates immune responses induced by an experimental malaria vaccine and consequently suppresses the protective efficacy of the vaccine against malaria challenge.

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Implications of Dengue Virus Maturation on Vaccine Induced Humoral Immunity in Mice

Viruses ◽

10.3390/v13091843 ◽

2021 ◽

Vol 13 (9) ◽

pp. 1843

Author(s):

Connor A. P. Scott ◽

Alberto A. Amarilla ◽

Summa Bibby ◽

Natalee D. Newton ◽

Roy A. Hall ◽

...

Keyword(s):

Immune Responses ◽

Dengue Virus ◽

Humoral Immunity ◽

Vaccine Candidate ◽

Serum Concentrations ◽

Vaccine Candidates ◽

Virus Maturation ◽

Potential Safety ◽

Increased Risk

The use of dengue virus (DENV) vaccines has been hindered by the complexities of antibody dependent enhancement (ADE). Current late-stage vaccine candidates utilize attenuated and chimeric DENVs that produce particles of varying maturities. Antibodies that are elicited by preferentially exposed epitopes on immature virions have been linked to increased ADE. We aimed to further understand the humoral immunity promoted by DENV particles of varying maturities in an AG129 mouse model using a chimeric insect specific vaccine candidate, bDENV-2. We immunized mice with mature, partially mature, and immature bDENV-2 and found that immunization with partially mature bDENV-2 produced more robust and cross-neutralizing immune responses than immunization with immature or mature bDENV-2. Upon challenge with mouse adapted DENV-2 (D220), we observed 80% protection for mature bDENV-2 vaccinated mice and 100% for immature and partially mature vaccinated mice, suggesting that protection to homotypic challenge is not dependent on maturation. Finally, we found reduced in vitro ADE at subneutralising serum concentrations for mice immunized with mature bDENV-2. These results suggest that both immature and mature DENV particles play a role in homotypic protection; however, the increased risk of in vitro ADE from immature particles indicates potential safety benefits from mature DENV-based vaccines.

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RelCoVax®, a two antigen subunit protein vaccine candidate against SARS-CoV-2 induces strong immune responses in mice

10.1101/2022.01.07.475330 ◽

2022 ◽

Author(s):

Abhishek Phatarphekar ◽

GEC Vidyadhar Reddy ◽

Abhiram Gokhale ◽

Gopala Karanam ◽

Pushpa Kuchroo ◽

...

Keyword(s):

Immune Responses ◽

Mammalian Cells ◽

Neutralizing Antibodies ◽

Vaccine Candidate ◽

Protein Vaccine ◽

Vaccine Candidates ◽

N Protein ◽

Subunit Protein ◽

Boost Dose

The COVID-19 pandemic has spurred an unprecedented movement to develop safe and effective vaccines against the SARS-CoV-2 virus to immunize the global population. The first set of vaccine candidates that received emergency use authorization targeted the spike (S) glycoprotein of the SARS-CoV-2 virus that enables virus entry into cells via the receptor binding domain (RBD). Recently, multiple variants of SARS-CoV-2 have emerged with mutations in S protein and the ability to evade neutralizing antibodies in vaccinated individuals. We have developed a dual RBD and nucleocapsid (N) subunit protein vaccine candidate named RelCoVax® through heterologous expression in mammalian cells (RBD) and E. coli (N). The RelCoVax® formulation containing a combination of aluminum hydroxide (alum) and a synthetic CpG oligonucleotide as adjuvants elicited high antibody titers against RBD and N proteins in mice after a prime and boost dose regimen administered 2 weeks apart. The vaccine also stimulated cellular immune responses with a potential Th1 bias as evidenced by increased IFN-γ release by splenocytes from immunized mice upon antigen exposure particularly N protein. Finally, the serum of mice immunized with RelCoVax® demonstrated the ability to neutralize two different SARS-CoV-2 viral strains in vitro including the Delta strain that has become dominant in many regions of the world and can evade vaccine induced neutralizing antibodies. These results warrant further evaluation of RelCoVax® through advanced studies and contribute towards enhancing our understanding of multicomponent subunit vaccine candidates against SARS-CoV-2.

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In silico approach for designing a novel recombinant fusion protein as a Candidate Vaccine against HPV

Current Proteomics ◽

10.2174/1570164617999201014162235 ◽

2020 ◽

Vol 17 ◽

Author(s):

Mohsen Sisakht ◽

Amir Mahmoodzadeh ◽

Mohammadsaeid Zahedi ◽

Davood Rostamzadeh ◽

Amin Moradi Hasan-Abad ◽

...

Keyword(s):

Immune Responses ◽

Fusion Protein ◽

In Silico ◽

Precancerous Lesions ◽

Candidate Vaccine ◽

T Cell Epitopes ◽

Binding Interaction ◽

Hpv16 E6 ◽

E7 Proteins

Background: Human papillomavirus (HPV) is the main biological agent causing sexually transmitted diseases (STDs), including precancerous lesions and several types of prevalent cancers. To date, numerous types of vaccines are designed to prevent high-risk HPV. However, their prophylactic effect is not the same and does not clear previous infections. Therefore, there is an urgent need for developing therapeutic vaccines that trigger cell-mediated immune responses for the treatment of HPV. The HPV16 E6 and E7 proteins are ideal targets for vaccine therapy against HPV. Fusion protein vaccines, which include both immunogenic interest protein and an adjuvant for augmenting the immunogenicity effects, are theoretically capable of guarantee the power of the immune system against HPV. Method: A vaccine construct, including HPV16 E6/E7 proteins along with a heat shock protein GP96 (E6/E7-NTGP96 construct), was designed using in silico methods. By the aid of the SWISS-MODEL server, the optimal 3D model of the designed vaccine was selected, followed by physicochemical and molecular parameters were performed using bioinformatics tools. Docking studies were done to evaluate the binding interaction of the vaccine. Allergenicity, immunogenicity, B, and T cell epitopes of the designed construct were predicted. Results: Immunological and structural computational results illustrated that our designed construct is potentially proper for stimulation of cellular and humoral immune responses against HPV. Conclusion: Computational studies showed that the E6/E7-NTGP96 construct is a promising candidate vaccine that needs further in vitro and in vivo evaluations.

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Lysosomotropic agents including azithromycin, chloroquine and hydroxychloroquine activate the integrated stress response

Cell Death and Disease ◽

10.1038/s41419-020-03324-w ◽

2021 ◽

Vol 12 (1) ◽

Author(s):

Ai-Ling Tian ◽

Qi Wu ◽

Peng Liu ◽

Liwei Zhao ◽

Isabelle Martins ◽

...

Keyword(s):

Stress Response ◽

Immune Responses ◽

Initiation Factor ◽

Integrated Stress Response ◽

Serine Residue ◽

Eif2α Phosphorylation ◽

Lysosomotropic Agents ◽

Cultured Human Cells

AbstractThe integrated stress response manifests with the phosphorylation of eukaryotic initiation factor 2α (eIF2α) on serine residue 51 and plays a major role in the adaptation of cells to endoplasmic reticulum stress in the initiation of autophagy and in the ignition of immune responses. Here, we report that lysosomotropic agents, including azithromycin, chloroquine, and hydroxychloroquine, can trigger eIF2α phosphorylation in vitro (in cultured human cells) and, as validated for hydroxychloroquine, in vivo (in mice). Cells bearing a non-phosphorylatable eIF2α mutant (S51A) failed to accumulate autophagic puncta in response to azithromycin, chloroquine, and hydroxychloroquine. Conversely, two inhibitors of eIF2α dephosphorylation, nelfinavir and salubrinal, enhanced the induction of such autophagic puncta. Altogether, these results point to the unexpected capacity of azithromycin, chloroquine, and hydroxychloroquine to elicit the integrated stress response.

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Intestinal mesenchymal cells regulate immune responses and promote epithelial regeneration in vitro and in dextran sulfate sodium‐induced experimental colitis in mice

Acta Physiologica ◽

10.1111/apha.13699 ◽

2021 ◽

Author(s):

Laura Hidalgo‐Garcia ◽

José Alberto Molina‐Tijeras ◽

Francisco Huertas‐Peña ◽

Antonio Jesús Ruiz‐Malagón ◽

Patricia Diez‐Echave ◽

...

Keyword(s):

Immune Responses ◽

Dextran Sulfate ◽

Dextran Sulfate Sodium ◽

Experimental Colitis ◽

Mesenchymal Cells ◽

Epithelial Regeneration ◽

Regeneration In Vitro

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Evaluation of the Safety and Immune Efficacy of Recombinant Human Respiratory Syncytial Virus Strain Long Live Attenuated Vaccine Candidates

Virologica Sinica ◽

10.1007/s12250-021-00345-3 ◽

2021 ◽

Author(s):

Li-Nan Wang ◽

Xiang-Lei Peng ◽

Min Xu ◽

Yuan-Bo Zheng ◽

Yue-Ying Jiao ◽

...

Keyword(s):

Respiratory Syncytial Virus ◽

Gene Deletion ◽

Human Respiratory Syncytial Virus ◽

Temperature Sensitive ◽

T7 Promoter ◽

Vaccine Candidates ◽

Rsv Infection ◽

Syncytial Virus

AbstractHuman respiratory syncytial virus (RSV) infection is the leading cause of lower respiratory tract illness (LRTI), and no vaccine against LRTI has proven to be safe and effective in infants. Our study assessed attenuated recombinant RSVs as vaccine candidates to prevent RSV infection in mice. The constructed recombinant plasmids harbored (5′ to 3′) a T7 promoter, hammerhead ribozyme, RSV Long strain antigenomic cDNA with cold-passaged (cp) mutations or cp combined with temperature-sensitive attenuated mutations from the A2 strain (A2cpts) or further combined with SH gene deletion (A2cptsΔSH), HDV ribozyme (δ), and a T7 terminator. These vectors were subsequently co-transfected with four helper plasmids encoding N, P, L, and M2-1 viral proteins into BHK/T7-9 cells, and the recovered viruses were then passaged in Vero cells. The rescued recombinant RSVs (rRSVs) were named rRSV-Long/A2cp, rRSV-Long/A2cpts, and rRSV-Long/A2cptsΔSH, respectively, and stably passaged in vitro, without reversion to wild type (wt) at sites containing introduced mutations or deletion. Although rRSV-Long/A2cpts and rRSV-Long/A2cptsΔSH displayed temperature-sensitive (ts) phenotype in vitro and in vivo, all rRSVs were significantly attenuated in vivo. Furthermore, BALB/c mice immunized with rRSVs produced Th1-biased immune response, resisted wtRSV infection, and were free from enhanced respiratory disease. We showed that the combination of ΔSH with attenuation (att) mutations of cpts contributed to improving att phenotype, efficacy, and gene stability of rRSV. By successfully introducing att mutations and SH gene deletion into the RSV Long parent and producing three rRSV strains, we have laid an important foundation for the development of RSV live attenuated vaccines.

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