scholarly journals An Alphavirus Replicon Particle Chimera Derived from Venezuelan Equine Encephalitis and Sindbis Viruses Is a Potent Gene-Based Vaccine Delivery Vector

2003 ◽  
Vol 77 (19) ◽  
pp. 10394-10403 ◽  
Author(s):  
Silvia Perri ◽  
Catherine E. Greer ◽  
Kent Thudium ◽  
Barbara Doe ◽  
Harold Legg ◽  
...  
Keyword(s):  
Sindbis Virus ◽  
Semliki Forest Virus ◽  
Cultured Cells ◽  
Heterologous Gene Expression ◽  
Envelope Glycoproteins ◽  
Venezuelan Equine Encephalitis ◽  
Vaccine Vectors ◽  
Equine Encephalitis Virus ◽  
Replicon Particle

ABSTRACT Alphavirus replicon particle-based vaccine vectors derived from Sindbis virus (SIN), Semliki Forest virus, and Venezuelan equine encephalitis virus (VEE) have been shown to induce robust antigen-specific cellular, humoral, and mucosal immune responses in many animal models of infectious disease and cancer. However, since little is known about the relative potencies among these different vectors, we compared the immunogenicity of replicon particle vectors derived from two very different parental alphaviruses, VEE and SIN, expressing a human immunodeficiency virus type 1 p55Gag antigen. Moreover, to explore the potential benefits of combining elements from different alphaviruses, we generated replicon particle chimeras of SIN and VEE. Two distinct strategies were used to produce particles with VEE-p55 gag replicon RNA packaged within SIN envelope glycoproteins and SIN-p55 gag replicon RNA within VEE envelope glycoproteins. Each replicon particle configuration induced Gag-specific CD8+ T-cell responses in murine models when administered alone or after priming with DNA. However, Gag-specific responses varied dramatically, with the strongest responses to this particular antigen correlating with the VEE replicon RNA, irrespective of the source of envelope glycoproteins. Comparing the replicons with respect to heterologous gene expression levels and sensitivity to alpha/beta interferon in cultured cells indicated that each might contribute to potency differences. This work shows that combining desirable elements from VEE and SIN into a replicon particle chimera may be a valuable approach toward the goal of developing vaccine vectors with optimal in vivo potency, ease of production, and safety.

scholarly journals Lack of Interference with Immunogenicity of a Chimeric Alphavirus Replicon Particle-Based Influenza Vaccine by Preexisting Antivector Immunity

2012 ◽  
Vol 19 (7) ◽  
pp. 991-998 ◽  
Author(s):  
Yasushi Uematsu ◽  
Michael Vajdy ◽  
Ying Lian ◽  
Silvia Perri ◽  
Catherine E. Greer ◽  
...  
Keyword(s):  
Influenza Virus ◽  
Sindbis Virus ◽  
Antibody Responses ◽  
Target Antigen ◽  
Structural Components ◽  
Venezuelan Equine Encephalitis ◽  
Vaccine Platform ◽  
Lethal Challenge ◽  
Equine Encephalitis Virus ◽  
Replicon Particle

ABSTRACTAntivector immunity has been recognized as a potential caveat of using virus-based vaccines. In the present study, an alphavirus-based replicon particle vaccine platform, which has demonstrated robust immunogenicity in animal models, was tested for effects of antivector immunity on immunogenicity against hemagglutinin of influenza virus as a target antigen and efficacy for protection against lethal challenge with the virus. Chimeric alphavirus-based replicon particles, comprising Venezuelan equine encephalitis virus nonstructural and Sindbis virus structural components, induced efficient protective antibody responses, which were not adversely influenced after multiple immunizations with the same vector expressing various antigens.

scholarly journals Vaccination of Mice with Gonococcal TbpB Expressed In Vivo from Venezuelan Equine Encephalitis Viral Replicon Particles

2006 ◽  
Vol 74 (3) ◽  
pp. 1612-1620 ◽  
Author(s):  
Christopher E. Thomas ◽  
Weiyan Zhu ◽  
Cornelius N. Van Dam ◽  
Nancy L. Davis ◽  
Robert E. Johnston ◽  
...  
Keyword(s):  
Recombinant Protein ◽  
Specific Antibody ◽  
Serum Antibody ◽  
Venezuelan Equine Encephalitis ◽  
Equine Encephalitis Virus ◽  
Secretion Signal ◽  
Replicon Particle ◽  
Intact Surface ◽  
Antibody Levels

ABSTRACT We investigated the immunogenicity of gonococcal transferrin binding protein B (TbpB) expressed with and without a eukaryotic secretion signal from a nonpropagating Venezuelan equine encephalitis virus replicon particle (VRP) delivery system. TbpB was successfully expressed in baby hamster kidney (BHK) cells, and the presence of the eukaryotic secretion signal not only apparently increased the protein's expression but also allowed for extracellular localization and glycosylation. Mice immunized with VRPs produced significant amounts of serum antibody although less than the amounts produced by mice immunized with recombinant protein. The response of mice immunized with VRPs encoding TbpB was consistently more Th1 biased than the response of mice immunized with recombinant protein alone. Boosting with recombinant protein following immunization with TbpB VRPs resulted in higher specific-antibody levels without altering the Th1/Th2 bias. Most of the immunization groups produced significant specific antibody binding to the intact surface of the homologous Neisseria gonorrhoeae strain. Immunization with TbpB VRPs without a eukaryotic secretion signal generated no measurable specific antibodies on the genital mucosal surface, but inclusion of a eukaryotic secretion signal or boosting with recombinant protein resulted in specific immunoglobulin G (IgG) and IgA in mucosal secretions after TbpB VRP immunization. The TbpB VRP system has potential for an N. gonorrhoeae vaccine.

scholarly journals An alphavirus replicon-based vaccine expressing a stabilized Spike antigen induces protective immunity and prevents transmission of SARS-CoV-2 between cats

npj Vaccines ◽  
2021 ◽  
Vol 6 (1) ◽  
Author(s):  
Martijn A. Langereis ◽  
Irina C. Albulescu ◽  
Judith Stammen-Vogelzangs ◽  
Morindy Lambregts ◽  
Ken Stachura ◽  
...  
Keyword(s):  
Protective Immunity ◽  
Animal Species ◽  
Close Contact ◽  
Neutralizing Antibody ◽  
Antibody Responses ◽  
Venezuelan Equine Encephalitis ◽  
Equine Encephalitis Virus ◽  
Replicon Particle ◽  
Nasal Swabs ◽  
Animal Hosts

AbstractEarly in the SARS-CoV-2 pandemic concerns were raised regarding infection of new animal hosts and the effect on viral epidemiology. Infection of other animals could be detrimental by causing clinical disease, allowing further mutations, and bares the risk for the establishment of a non-human reservoir. Cats were the first reported animals susceptible to natural and experimental infection with SARS-CoV-2. Given the concerns these findings raised, and the close contact between humans and cats, we aimed to develop a vaccine candidate that could reduce SARS-CoV-2 infection and in addition to prevent spread among cats. Here we report that a Replicon Particle (RP) vaccine based on Venezuelan equine encephalitis virus, known to be safe and efficacious in a variety of animal species, could induce neutralizing antibody responses in guinea pigs and cats. The design of the SARS-CoV-2 spike immunogen was critical in developing a strong neutralizing antibody response. Vaccination of cats was able to induce high neutralizing antibody responses, effective also against the SARS-CoV-2 B.1.1.7 variant. Interestingly, in contrast to control animals, the infectious virus could not be detected in oropharyngeal or nasal swabs of vaccinated cats after SARS-CoV-2 challenge. Correspondingly, the challenged control cats spread the virus to in-contact cats whereas the vaccinated cats did not transmit the virus. The results show that the RP vaccine induces protective immunity preventing SARS-CoV-2 infection and transmission. These data suggest that this RP vaccine could be a multi-species vaccine useful to prevent infection and spread to and between animals should that approach be required.

scholarly journals Replicon Vectors Derived from Sindbis Virus and Semliki Forest Virus That Establish Persistent Replication in Host Cells

2000 ◽  
Vol 74 (20) ◽  
pp. 9802-9807 ◽  
Author(s):  
Silvia Perri ◽  
David A. Driver ◽  
Jason P. Gardner ◽  
Scott Sherrill ◽  
Barbara A. Belli ◽  
...  
Keyword(s):  
Sindbis Virus ◽  
Semliki Forest Virus ◽  
Cultured Cells ◽  
Nonstructural Protein ◽  
Host Cells ◽  
Heterologous Gene ◽  
High Level Expression ◽  
Genomic Rnas ◽  
Bhk Cells ◽  
High Level

ABSTRACT Alphavirus replicon vectors are well suited for applications where transient, high-level expression of a heterologous gene is required. Replicon vector expression in cells leads to inhibition of host macromolecular synthesis, culminating in eventual cell death by an apoptotic mechanism. For many applications, including gene expression studies in cultured cells, a longer duration of transgene expression without resulting cytopathic effects is useful. Recently, noncytopathic Sindbis virus (SIN) variants were isolated in BHK cells, and the mutations responsible were mapped to the protease domain of nonstructural protein 2 (nsP2). We report here the isolation of additional variants of both SIN and Semliki Forest virus (SFV) replicons encoding the neomycin resistance gene that can establish persistent replication in BHK cells. The SIN and SFV variant replicons resulted from previously undescribed mutations within one of three discrete regions of the nsP2 gene. Differences among the panel of variants were observed in processing of the nonstructural polyprotein and in the ratios of subgenomic to genomic RNAs. Importantly, high-level expression of a heterologous gene was retained with most replicons. Finally, in contrast to previous studies, efficient packaging was obtained with several of the variant replicons. This work expands the utility of noncytopathic replicons and the understanding of how alphavirus replicons establish persistent replication in cultured cells.

scholarly journals Replication and Clearance of Venezuelan Equine Encephalitis Virus from the Brains of Animals Vaccinated with Chimeric SIN/VEE Viruses

2006 ◽  
Vol 80 (6) ◽  
pp. 2784-2796 ◽  
Author(s):  
Slobodan Paessler ◽  
Haolin Ni ◽  
Olga Petrakova ◽  
Rafik Z. Fayzulin ◽  
Nadezhda Yun ◽  
...  
Keyword(s):  
Venezuelan Equine Encephalitis Virus ◽  
Encephalitis Virus ◽  
Venezuelan Equine Encephalitis ◽  
Challenge Virus ◽  
Equine Encephalitis Virus ◽  
Adult Mice ◽  
The Central Nervous System ◽  
Different Strains

ABSTRACT Venezuelan equine encephalitis virus (VEEV) is an important, naturally emerging zoonotic pathogen. Recent outbreaks in Venezuela and Colombia in 1995, involving an estimated 100,000 human cases, indicate that VEEV still poses a serious public health threat. To develop a safe, efficient vaccine that protects against disease resulting from VEEV infection, we generated chimeric Sindbis (SIN) viruses expressing structural proteins of different strains of VEEV and analyzed their replication in vitro and in vivo, as well as the characteristics of the induced immune responses. None of the chimeric SIN/VEE viruses caused any detectable disease in adult mice after either intracerebral (i.c.) or subcutaneous (s.c.) inoculation, and all chimeras were more attenuated than the vaccine strain, VEEV TC83, in 6-day-old mice after i.c. infection. All vaccinated mice were protected against lethal encephalitis following i.c., s.c., or intranasal (i.n.) challenge with the virulent VEEV ZPC738 strain (ZPC738). In spite of the absence of clinical encephalitis in vaccinated mice challenged with ZPC738 via i.n. or i.c. route, we regularly detected high levels of infectious challenge virus in the central nervous system (CNS). However, infectious virus was undetectable in the brains of all immunized animals at 28 days after challenge. Hamsters vaccinated with chimeric SIN/VEE viruses were also protected against s.c. challenge with ZPC738. Taken together, our findings suggest that these chimeric SIN/VEE viruses are safe and efficacious in adult mice and hamsters and are potentially useful as VEEV vaccines. In addition, immunized animals provide a useful model for studying the mechanisms of the anti-VEEV neuroinflammatory response, leading to the reduction of viral titers in the CNS and survival of animals.

scholarly journals A Single-Site Mutant and Revertants Arising in Vivo Define Early Steps in the Pathogenesis of Venezuelan Equine Encephalitis Virus

Virology ◽  
2000 ◽  
Vol 270 (1) ◽  
pp. 111-123 ◽  
Author(s):  
Judith F. Aronson ◽  
Franziska B. Grieder ◽  
Nancy L. Davis ◽  
Peter C. Charles ◽  
Travis Knott ◽  
...  
Keyword(s):  
Venezuelan Equine Encephalitis Virus ◽  
Encephalitis Virus ◽  
Single Site ◽  
Venezuelan Equine Encephalitis ◽  
Equine Encephalitis Virus

scholarly journals Ablation of Programmed −1 Ribosomal Frameshifting in Venezuelan Equine Encephalitis Virus Results in Attenuated Neuropathogenicity

2016 ◽  
Vol 91 (3) ◽  
Author(s):  
Joseph A. Kendra ◽  
Cynthia de la Fuente ◽  
Ashwini Brahms ◽  
Caitlin Woodson ◽  
Todd M. Bell ◽  
...  
Keyword(s):  
Cultured Cells ◽  
Rna Virus ◽  
Venezuelan Equine Encephalitis Virus ◽  
Encephalitis Virus ◽  
Venezuelan Equine Encephalitis ◽  
Ribosomal Frameshifting ◽  
Positive Strand ◽  
Live Attenuated Vaccines ◽  
Equine Encephalitis Virus ◽  
Positive Strand Rna

ABSTRACT The alphaviruses Venezuelan equine encephalitis virus (VEEV), eastern equine encephalitis virus (EEEV), and western equine encephalitis virus (WEEV) are arthropod-borne positive-strand RNA viruses that are capable of causing acute and fatal encephalitis in many mammals, including humans. VEEV was weaponized during the Cold War and is recognized as a select agent. Currently, there are no FDA-approved vaccines or therapeutics for these viruses. The spread of VEEV and other members of this family due to climate change-mediated vector range expansion underscores the need for research aimed at developing medical countermeasures. These viruses utilize programmed −1 ribosomal frameshifting (−1 PRF) to synthesize the viral trans-frame (TF) protein, which has previously been shown to be important for neuropathogenesis in the related Sindbis virus. Here, the alphavirus −1 PRF signals were characterized, revealing novel −1 PRF stimulatory structures. −1 PRF attenuation mildly affected the kinetics of VEEV accumulation in cultured cells but strongly inhibited its pathogenesis in an aerosol infection mouse model. Importantly, the decreased viral titers in the brains of mice infected with the mutant virus suggest that the alphavirus TF protein is important for passage through the blood-brain barrier and/or for neuroinvasiveness. These findings suggest a novel approach to the development of safe and effective live attenuated vaccines directed against VEEV and perhaps other closely related −1 PRF-utilizing viruses. IMPORTANCE Venezuelan equine encephalitis virus (VEEV) is a select agent that has been weaponized. This arthropod-borne positive-strand RNA virus causes acute and fatal encephalitis in many mammals, including humans. There is no vaccine or other approved therapeutic. VEEV and related alphaviruses utilize programmed −1 ribosomal frameshifting (−1 PRF) to synthesize the viral trans-frame (TF) protein, which is important for neuropathogenesis. −1 PRF attenuation strongly inhibited VEEV pathogenesis in mice, and viral replication analyses suggest that the TF protein is critical for neurological disease. These findings suggest a new approach to the development of safe and effective live attenuated vaccines directed against VEEV and other related viruses.

scholarly journals Insertion of EGFP into the replicase gene of Semliki Forest virus results in a novel, genetically stable marker virus

2007 ◽  
Vol 88 (4) ◽  
pp. 1225-1230 ◽  
Author(s):  
Nele Tamberg ◽  
Valeria Lulla ◽  
Rennos Fragkoudis ◽  
Aleksei Lulla ◽  
John K. Fazakerley ◽  
...  
Keyword(s):  
Fluorescent Protein ◽  
Semliki Forest Virus ◽  
Cultured Cells ◽  
Marker Gene ◽  
Virus Production ◽  
Marker Genes ◽  
Replicase Gene ◽  
Marker Virus

Alphavirus-based vector and replicon systems have been extensively used experimentally and are likely to be used in human and animal medicine. Whilst marker genes can be inserted easily under the control of a duplicated subgenomic promoter, these constructs are often genetically unstable. Here, a novel alphavirus construct is described in which an enhanced green fluorescent protein (EGFP) marker gene is inserted into the virus replicase open reading frame between nsP3 and nsP4, flanked by nsP2 protease-recognition sites. This construct has correct processing of the replicase polyprotein, produces viable virus and expresses detectable EGFP fluorescence upon infection of cultured cells and cells of the mouse brain. In comparison to parental virus, the marker virus has an approximately 1 h delay in virus RNA and infectious virus production. Passage of the marker virus in vitro and in vivo demonstrates good genetic stability. Insertion of different markers into this novel construct has potential for various applications.

scholarly journals Role of Alpha/Beta Interferon in Venezuelan Equine Encephalitis Virus Pathogenesis: Effect of an Attenuating Mutation in the 5′ Untranslated Region

2001 ◽  
Vol 75 (8) ◽  
pp. 3706-3718 ◽  
Author(s):  
Laura J. White ◽  
Jia-Gang Wang ◽  
Nancy L. Davis ◽  
Robert E. Johnston
Keyword(s):  
Untranslated Region ◽  
Mutant Virus ◽  
Venezuelan Equine Encephalitis ◽  
Wild Type ◽  
Average Survival Time ◽  
Equine Encephalitis Virus ◽  
Average Survival ◽  
Alpha Beta

ABSTRACT Venezuelan equine encephalitis virus (VEE) is an important equine and human pathogen of the Americas. In the adult mouse model, cDNA-derived, virulent V3000 inoculated subcutaneously (s.c.) causes high-titer peripheral replication followed by neuroinvasion and lethal encephalitis. A single change (G to A) at nucleotide 3 (nt 3) of the 5′ untranslated region (UTR) of the V3000 genome resulted in a virus (V3043) that was avirulent in mice. The mechanism of attenuation by the V3043 mutation was studied in vivo and in vitro. Kinetic studies of virus spread in adult mice following s.c. inoculation showed that V3043 replication was reduced in peripheral organs compared to that of V3000, titers in serum also were lower, and V3043 was cleared more rapidly from the periphery than V3000. Because clearance of V3043 from serum began 1 to 2 days prior to clearance of V3000, we examined the involvement of alpha/beta interferon (IFN-α/β) activity in VEE pathogenesis. In IFN-α/βR−/− mice, the course of the wild-type disease was extremely rapid, with all animals dying within 48 h (average survival time of 30 h compared to 7.7 days in the wild-type mice). The mutant V3043 was as virulent as the wild type (100% mortality, average survival time of 30 h). Virus titers in serum, peripheral organs, and the brain were similar in V3000- and V3043-infected IFN-α/βR−/− mice at all time points up until the death of the animals. Consistent with the in vivo data, the mutant virus exhibited reduced growth in vitro in several cell types except in cells that lacked a functional IFN-α/β pathway. In cells derived from IFN-α/βR−/− mice, the mutant virus showed no growth disadvantage compared to the wild-type virus, suggesting that IFN-α/β plays a major role in the attenuation of V3043 compared to V3000. There were no differences in the induction of IFN-α/β between V3000 and V3043, but the mutant virus was more sensitive than V3000 to the antiviral actions of IFN-α/β in two separate in vitro assays, suggesting that the increased sensitivity to IFN-α/β plays a major role in the in vivo attenuation of V3043.

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