A tetravalent dengue virus-like particle vaccine induces high levels of neutralizing antibodies and reduces dengue replication in non-human primates.

Dengue virus (DENV) represents a significant global health burden, with 50% of the world's population at risk of infection, and there is an urgent need for next-generation vaccines. Virus-like particle (VLP)-based vaccines, which mimic the antigenic structure of the virus but lack the viral genome, are an attractive approach. Here, we describe a dengue VLP (DENVLP) vaccine which generates a neutralizing antibody response against all four DENV serotypes in 100% of immunized non-human primates for up to 1 year.

Safety and immunogenicity of VLPCOV-02, a SARS-CoV-2 self-amplifying RNA vaccine with a modified base, 5-methylcytosine.

Continuing emergence of variants of concern resulting in reduced SARS-CoV-2 vaccine efficacy necessitates additional prevention strategies. The structure of VLPCOV-01, a lipid nanoparticle-encapsulated, self-amplifying RNA COVID-19 vaccine with a comparable immune response to BNT162b2, was revised by incorporating a modified base, 5-methylcytosine, to reduce reactogenicity, and an updated receptor-binding domain derived from the Brazil (gamma) variant. Interim analyses of a phase 1 dose-escalation booster vaccination study with the resulting construct, VLPCOV-02, in healthy, previously vaccinated Japanese individuals (N = 96) are reported (jRCT2051230005).

Incorporation of 5 methylcytidine alleviates innate immune response to self-amplifying RNA vaccine.

In order to improve vaccine effectiveness and safety profile of existing synthetic RNA-based vaccines, we have developed a self-amplifying RNA (saRNA)-based vaccine expressing membrane-anchored receptor binding domain (RBD) of SARS-CoV-2 S protein (S-RBD) and have demonstrated that a minimal dose of this saRNA vaccine elicits robust immune responses. Results from a recent clinical trial with 5-methylcytidine (5mC) incorporating saRNA vaccine demonstrated reduced vaccine-induced adverse effects while maintaining robust humoral responses. In this study, we investigate the mechanisms accounting for induction of efficient innate and adaptive immune responses and attenuated adverse effects induced by the 5mC-incorporated saRNA.

Safety and immunogenicity of SARS-CoV-2 self-amplifying RNA vaccine expressing an anchored RBD: A randomized, observer-blind phase 1 study.

VLPCOV-01 is a lipid nanoparticle-encapsulated self-amplifying RNA (saRNA) vaccine that expresses a membrane-anchored receptor-binding domain (RBD) derived from the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike protein. A phase 1 study of VLPCOV-01 is conducted (jRCT2051210164). Participants who completed two doses of the BNT162b2 mRNA vaccine previously are randomized to receive one intramuscular vaccination of 0.

saRNA vaccine expressing membrane-anchored RBD elicits broad and durable immunity against SARS-CoV-2 variants of concern.

Several vaccines have been widely used to counteract the global pandemic caused by SARS-CoV-2. However, due to the rapid emergence of SARS-CoV-2 variants of concern (VOCs), further development of vaccines that confer broad and longer-lasting protection against emerging VOCs are needed. Here, we report the immunological characteristics of a self-amplifying RNA (saRNA) vaccine expressing the SARS-CoV-2 Spike (S) receptor binding domain (RBD), which is membrane-anchored by fusing with an N-terminal signal sequence and a C-terminal transmembrane domain (RBD-TM).

Reassessing therapeutic antibodies for neglected and tropical diseases.

In the past two decades there has been a significant expansion in the number of new therapeutic monoclonal antibodies (mAbs) that are approved by regulators. The discovery of these new medicines has been driven primarily by new approaches in inflammatory diseases and oncology, especially in immuno-oncology. Other recent successes have included new antibodies for use in viral diseases, including HIV.

A virus-like particle vaccine prevents equine encephalitis virus infection in nonhuman primates.

Western, Eastern, and Venezuelan equine encephalitis viruses (WEEV, EEEV, and VEEV, respectively) are important mosquito-borne agents that pose public health and bioterrorism threats. Despite considerable advances in understanding alphavirus replication, there are currently no available effective vaccines or antiviral treatments against these highly lethal pathogens. To develop a potential countermeasure for viral encephalitis, we generated a trivalent, or three-component, EEV vaccine composed of virus-like particles (VLPs).

Structural studies of Chikungunya virus maturation.

Cleavage of the alphavirus precursor glycoprotein p62 into the E2 and E3 glycoproteins before assembly with the nucleocapsid is the key to producing fusion-competent mature spikes on alphaviruses. Here we present a cryo-EM, 6.8-Å resolution structure of an "immature" Chikungunya virus in which the cleavage site has been mutated to inhibit proteolysis.

An Envelope-Modified Tetravalent Dengue Virus-Like-Particle Vaccine Has Implications for Flavivirus Vaccine Design.

Dengue viruses (DENV) infect 50 to 100 million people each year. The spread of DENV-associated infections is one of the most serious public health problems worldwide, as there is no widely available vaccine or specific therapeutic for DENV infections. To address this, we developed a novel tetravalent dengue vaccine by utilizing virus-like particles (VLPs).

Development of a Novel Virus-Like Particle Vaccine Platform That Mimics the Immature Form of Alphavirus.

Virus-like particles (VLPs) are noninfectious multiprotein structures that are engineered to self-assemble from viral structural proteins. Here, we developed a novel VLP-based vaccine platform utilizing VLPs from the chikungunya virus. We identified two regions within the envelope protein, a structural component of chikungunya, where foreign antigens can be inserted without compromising VLP structure.

Structural Studies of Chikungunya Virus-Like Particles Complexed with Human Antibodies: Neutralization and Cell-to-Cell Transmission.

Unlabelled: Chikungunya virus is a positive-stranded RNA alphavirus. Structures of chikungunya virus-like particles in complex with strongly neutralizing antibody Fab fragments (8B10 and 5F10) were determined using cryo-electron microscopy and X-ray crystallography. By fitting the crystallographically determined structures of these Fab fragments into the cryo-electron density maps, we show that Fab fragments of antibody 8B10 extend radially from the viral surface and block receptor binding on the E2 glycoprotein.

Cryo-EM structures elucidate neutralizing mechanisms of anti-chikungunya human monoclonal antibodies with therapeutic activity.

Chikungunya virus (CHIKV) is a mosquito-transmitted alphavirus that causes severe acute and chronic disease in humans. Although highly inhibitory murine and human monoclonal antibodies (mAbs) have been generated, the structural basis of their neutralizing activity remains poorly characterized. Here, we determined the cryo-EM structures of chikungunya virus-like particles complexed with antibody fragments (Fab) of two highly protective human mAbs, 4J21 and 5M16, that block virus fusion with host membranes.

Safety and tolerability of chikungunya virus-like particle vaccine in healthy adults: a phase 1 dose-escalation trial.

Background: Chikungunya virus--a mosquito-borne alphavirus--is endemic in Africa and south and southeast Asia and has recently emerged in the Caribbean. No drugs or vaccines are available for treatment or prevention. We aimed to assess the safety, tolerability, and immunogenicity of a new candidate vaccine.

Development of a highly protective combination monoclonal antibody therapy against Chikungunya virus.

Chikungunya virus (CHIKV) is a mosquito-transmitted alphavirus that causes global epidemics of a debilitating polyarthritis in humans. As there is a pressing need for the development of therapeutic agents, we screened 230 new mouse anti-CHIKV monoclonal antibodies (MAbs) for their ability to inhibit infection of all three CHIKV genotypes. Four of 36 neutralizing MAbs (CHK-102, CHK-152, CHK-166, and CHK-263) provided complete protection against lethality as prophylaxis in highly susceptible immunocompromised mice lacking the type I IFN receptor (Ifnar(-/-) ) and mapped to distinct epitopes on the E1 and E2 structural proteins.

Structural analyses at pseudo atomic resolution of Chikungunya virus and antibodies show mechanisms of neutralization.

A 5.3 Å resolution, cryo-electron microscopy (cryoEM) map of Chikungunya virus-like particles (VLPs) has been interpreted using the previously published crystal structure of the Chikungunya E1-E2 glycoprotein heterodimer. The heterodimer structure was divided into domains to obtain a good fit to the cryoEM density.

A specific domain of the Chikungunya virus E2 protein regulates particle formation in human cells: implications for alphavirus vaccine design.

Virus-like particles (VLPs) can be generated from Chikungunya virus (CHIKV), but different strains yield variable quantities of particles. Here, we define the genetic basis for these differences and show that amino acid 234 in E2 substantially affects VLP production. This site is located within the acid-sensitive region (ASR) known to initiate a major conformational change in E1/E2.

A virus-like particle vaccine for epidemic Chikungunya virus protects nonhuman primates against infection.

Chikungunya virus (CHIKV) has infected millions of people in Africa, Europe and Asia since this alphavirus reemerged from Kenya in 2004. The severity of the disease and the spread of this epidemic virus present a serious public health threat in the absence of vaccines or antiviral therapies. Here, we describe a new vaccine that protects against CHIKV infection of nonhuman primates.

Different vaccine vectors delivering the same antigen elicit CD8+ T cell responses with distinct clonotype and epitope specificity.

Prime-boost immunization with gene-based vectors has been developed to generate more effective vaccines for AIDS, malaria, and tuberculosis. Although these vectors elicit potent T cell responses, the mechanisms by which they stimulate immunity are not well understood. In this study, we show that immunization by a single gene product, HIV-1 envelope, with alternative vector combinations elicits CD8(+) cells with different fine specificities and kinetics of mobilization.

Protein methyltransferase 2 inhibits NF-kappaB function and promotes apoptosis.

The protein arginine methyltransferases (PRMTs) include a family of proteins with related putative methyltransferase domains that modify chromatin and regulate cellular transcription. Although some family members, PRMT1 and PRMT4, have been implicated in transcriptional modulation or intracellular signaling, the roles of other PRMTs in diverse cellular processes have not been fully established. Here, we report that PRMT2 inhibits NF-kappaB-dependent transcription and promotes apoptosis.

Analysis of evolutionary conservation in CD1d molecules among primates.

The hereditary conservation in the genetically encoded CD1D sequences of various primates was analyzed. Genomic CD1D sequences of 17 rhesus macaques with distinct origins, eight Indian and nine Chinese, were examined and differences of only one or two nucleotides were detected and the consensus sequence of rhesus CD1D was determined. CD1D consensus sequences of three African green monkeys (AGMs) and the rhesus monkeys were then compared to study the evolutionary differences among interspecies.

DNA vaccination of macaques by a full-genome SHIV plasmid that has an IL-2 gene and produces non-infectious virus particles.

We previously reported that a mutant full-sized plasmid DNA vaccine regime in macaques was effective against a homologous challenge [Akahata W, Ido E, Shimada T, Katsuyama K, Yamamoto H, Uesaka H, et al. DNA vaccination of macaques by a full genome HIV-1 plasmid which produces non-infectious virus particles. Virology 2000;275:116-24; Akahata W, Ido E, Akiyama H, Uesaka H, Enose Y, Horiuchi R, et al.

Comparative immunogenicity of human immunodeficiency virus particles and corresponding polypeptides in a DNA vaccine.

The immunogenicity of a plasmid DNA expression vector encoding both Gag and envelope (Env), which produced human immunodeficiency virus (HIV) type 1 virus-like particles (VLP), was compared to vectors expressing Gag and Env individually, which presented the same gene products as polypeptides. Vaccination with plasmids that generated VLP showed cellular immunity comparable to that of Gag and cell-mediated or humoral responses similar to those of Env as immunization with separate vectors. These data suggest that DNA vaccines encoding separated HIV polypeptides generate immune responses similar to those generated by viral particles.

DNA vaccination of macaques by a full-genome simian/human immunodeficiency virus type 1 plasmid chimera that produces non-infectious virus particles.

A DNA vaccination regime was investigated previously in rhesus macaques using a full-genome human immunodeficiency virus type 1 (HIV-1) plasmid, which, due to mutations in the nucleocapsid (NC) proteins, produced only non-infectious HIV-1 particles (Akahata et al., Virology 275, 116-124, 2000). In that study, four monkeys were injected intramuscularly 14 times with the plasmid.

Construction and in vivo infection of a new simian/human immunodeficiency virus chimera containing the reverse transcriptase gene and the 3' half of the genomic region of human immunodeficiency virus type 1.

A new simian/human immunodeficiency virus (SHIV) chimera with the reverse transcriptase (RT)-encoding region of pol, in addition to the 3' region encoding vpr, vpu, tat, rev, env and nef of HIV-1, on an SIV(mac) (SIV from a macaque monkey) background was constructed. This new SHIV chimera, named SHIVrt/3rn, could replicate in monkey peripheral blood mononuclear cells (PBMCs) as well as in the human and monkey CD4(+) T-cell lines M8166 and HSC-F. Since SHIVrt/3rn contains the RT gene of HIV-1, replication of the virus in M8166 cells was inhibited by an HIV-1-specific non-nucleoside RT inhibitor, MKC-442, with a sensitivity similar to that of HIV-1.

Mutational analysis of two zinc-finger motifs in the nucleocapsid protein of simian immunodeficiency virus mac239.

To clarify the physiological function of two zinc-finger (ZF) motifs in the nucleocapsid (NC) protein of simian immunodeficiency virus (SIV), we constructed three mutant viruses with alterations in either or both motifs using a molecular clone of SIVmac (SIVmac239). An immunoblot analysis of the cell lysates transfected with DNA mutated in either the first (ZF1) or second (ZF2) motif showed that the amount of partially processed Gag products (Pr46) was greater than that produced by the wild-type (WT). The genomic RNA contents in the viral particles released from the transfected cells were measured by quantitative RT-PCR.