scholarly journals Parainfluenza Virus 5 Expressing Wild-Type or Prefusion Respiratory Syncytial Virus (RSV) Fusion Protein Protects Mice and Cotton Rats from RSV Challenge

2017 ◽  
Vol 91 (19) ◽  
Author(s):  
Shannon I. Phan ◽  
James R. Zengel ◽  
Huiling Wei ◽  
Zhuo Li ◽  
Dai Wang ◽  
...  
Keyword(s):  
Respiratory Syncytial Virus ◽  
Fusion Protein ◽  
Vaccine Candidate ◽  
Parainfluenza Virus ◽  
Wild Type ◽  
Rsv Infection ◽  
Cotton Rats ◽  
Parainfluenza Virus 5 ◽  
Syncytial Virus ◽  
Vectored Vaccine

ABSTRACT Human respiratory syncytial virus (RSV) is the leading cause of pediatric bronchiolitis and hospitalizations. RSV can also cause severe complications in elderly and immunocompromised individuals. There is no licensed vaccine. We previously generated a parainfluenza virus 5 (PIV5)-vectored vaccine candidate expressing the RSV fusion protein (F) that was immunogenic and protective in mice. In this work, our goal was to improve the original vaccine candidate by modifying the PIV5 vector or by modifying the RSV F antigen. We previously demonstrated that insertion of a foreign gene at the PIV5 small hydrophobic (SH)–hemagglutinin-neuraminidase (HN) junction or deletion of PIV5 SH increased vaccine efficacy. Additionally, other groups have demonstrated that antibodies against the prefusion conformation of RSV F have more potent neutralizing activity than antibodies against the postfusion conformation. Therefore, to improve on our previously developed vaccine candidate, we inserted RSV F at the PIV5 SH-HN gene junction or used RSV F to replace PIV5 SH. We also engineered PIV5 to express a prefusion-stabilized F mutant. The candidates were tested in BALB/c mice via the intranasal route and induced both humoral and cell-mediated immunity. They also protected against RSV infection in the mouse lung. When they were administered intranasally or subcutaneously in cotton rats, the candidates were highly immunogenic and reduced RSV loads in both the upper and lower respiratory tracts. PIV5-RSV F was equally protective when administered intranasally or subcutaneously. In all cases, the prefusion F mutant did not induce higher neutralizing antibody titers than wild-type F. These results show that antibodies against both pre- and postfusion F are important for neutralizing RSV and should be considered when designing a vectored RSV vaccine. The findings also that indicate PIV5-RSV F may be administered subcutaneously, which is the preferred route for vaccinating infants, who may develop nasal congestion as a result of intranasal vaccination. IMPORTANCE Despite decades of research, human respiratory syncytial virus (RSV) is still a major health concern for which there is no vaccine. A parainfluenza virus 5-vectored vaccine expressing the native RSV fusion protein (F) has previously been shown to confer robust immunity against RSV infection in mice, cotton rats, and nonhuman primates. To improve our previous vaccine candidate, we developed four new candidates that incorporate modifications to the PIV5 backbone, replace native RSV F with a prefusion-stabilized RSV F mutant, or combine both RSV F and PIV5 backbone modifications. In this work, we characterized the new vaccine candidates and tested their efficacies in both murine and cotton rat models of RSV infection. Most importantly, we found that PIV5-based RSV vaccine candidates were efficacious in preventing lower respiratory tract infection as well as in reducing the nasal viral load when administered via the subcutaneous route.

scholarly journals A Single-Dose Recombinant Parainfluenza Virus 5-Vectored Vaccine Expressing Respiratory Syncytial Virus (RSV) F or G Protein Protected Cotton Rats and African Green Monkeys from RSV Challenge

2017 ◽  
Vol 91 (11) ◽  
Author(s):  
Dai Wang ◽  
Shannon Phan ◽  
Daniel J. DiStefano ◽  
Michael P. Citron ◽  
Cheryl L. Callahan ◽  
...  
Keyword(s):  
Single Dose ◽  
Respiratory Syncytial Virus ◽  
G Protein ◽  
Parainfluenza Virus ◽  
Lung Pathology ◽  
Rsv Infection ◽  
Cotton Rats ◽  
Parainfluenza Virus 5 ◽  
Syncytial Virus ◽  
Green Monkeys

ABSTRACT Human respiratory syncytial virus (RSV) is a common cause of severe respiratory disease among infants, immunocompromised individuals, and the elderly. No licensed vaccine is currently available. In this study, we evaluated two parainfluenza virus 5 (PIV5)-vectored vaccines expressing RSV F (PIV5/F) or G (PIV5/G) protein in the cotton rat and African green monkey models for their replication, immunogenicity, and efficacy of protection against RSV challenge. Following a single intranasal inoculation, both animal species shed the vaccine viruses for a limited time but without noticeable clinical symptoms. In cotton rats, the vaccines elicited RSV F- or G-specific serum antibodies and conferred complete lung protection against RSV challenge at doses as low as 103 PFU. Neither vaccine produced the enhanced lung pathology observed in animals immunized with formalin-inactivated RSV. In African green monkeys, vaccine-induced serum and mucosal antibody responses were readily detected, as well. PIV5/F provided nearly complete protection against RSV infection in the upper and lower respiratory tract at a dose of 106 PFU of vaccine. At the same dose levels, PIV5/G was less efficacious. Both PIV5/F and PIV5/G were also able to boost neutralization titers in RSV-preexposed African green monkeys. Overall, our data indicated that PIV5/F is a promising RSV vaccine candidate. IMPORTANCE A safe and efficacious respiratory syncytial virus (RSV) vaccine remains elusive. We tested the recombinant parainfluenza virus 5 (PIV5) vectors expressing RSV glycoproteins for their immunogenicity and protective efficacy in cotton rats and African green monkeys, which are among the best available animal models to study RSV infection. In both species, a single dose of intranasal immunization with PIV5-vectored vaccines was able to produce systemic and local immunity and to protect animals from RSV challenge. The vaccines could also boost RSV neutralization antibody titers in African green monkeys that had been infected previously. Our data suggest that PIV5-vectored vaccines could potentially protect both the pediatric and elderly populations and support continued development of the vector platform.

scholarly journals Parainfluenza virus 5 (PIV5) amplifying virus-like particles expressing respiratory syncytial virus (RSV) antigens protect mice against RSV infection

Vaccine ◽  
2019 ◽  
Vol 37 (22) ◽  
pp. 2925-2934 ◽  
Author(s):  
María Cristina Huertas-Díaz ◽  
Shannon Phan ◽  
Andrew Elson ◽  
Ivette Nuñez ◽  
Huiling Wei ◽  
...  
Keyword(s):  
Respiratory Syncytial Virus ◽  
Parainfluenza Virus ◽  
Virus Like Particles ◽  
Rsv Infection ◽  
Parainfluenza Virus 5 ◽  
Syncytial Virus

scholarly journals Enhancing the Thermostability and Immunogenicity of a Respiratory Syncytial Virus (RSV) Live-Attenuated Vaccine by Incorporating Unique RSV Line19F Protein Residues

2017 ◽  
Vol 92 (6) ◽  
Author(s):  
Christina A. Rostad ◽  
Christopher C. Stobart ◽  
Sean O. Todd ◽  
Samuel A. Molina ◽  
Sujin Lee ◽  
...  
Keyword(s):  
Respiratory Syncytial Virus ◽  
Vaccine Candidate ◽  
Nonstructural Protein ◽  
Wild Type ◽  
Live Attenuated Vaccine ◽  
F Protein ◽  
Attenuated Vaccine ◽  
Cotton Rats ◽  
Syncytial Virus ◽  
Tract Infections

ABSTRACTRespiratory syncytial virus (RSV) is the leading cause of lower respiratory tract infections in infants, and an effective vaccine is not yet available. We previously generated an RSV live-attenuated vaccine (LAV) candidate, DB1, which was attenuated by a low-fusion subgroup B F protein (BAF) and codon-deoptimized nonstructural protein genes. DB1 was immunogenic and protective in cotton rats but lacked thermostability and stability of the prefusion conformation of F compared to strains with the line19F gene. We hypothesized that substitution of unique residues from the thermostable A2-line19F strain could thermostabilize DB1 and boost its immunogenicity. We therefore substituted 4 unique line19F residues into the BAF protein of DB1 by site-directed mutagenesis and rescued the recombinant virus, DB1-QUAD. Compared to DB1, DB1-QUAD had improved thermostability at 4°C and higher levels of prefusion F as measured by enzyme-linked immunosorbent assays (ELISAs). DB1-QUAD was attenuated in normal human bronchial epithelial cells, in BALB/c mice, and in cotton rats but grew to wild-type titers in Vero cells. In mice, DB1-QUAD was highly immunogenic and generated significantly higher neutralizing antibody titers to a panel of RSV A and B strains than did DB1. DB1-QUAD was also efficacious against wild-type RSV challenge in mice and cotton rats. Thus, substitution of unique line19F residues into RSV LAV DB1 enhanced vaccine thermostability, incorporation of prefusion F, and immunogenicity and generated a promising vaccine candidate that merits further investigation.IMPORTANCEWe boosted the thermostability and immunogenicity of an RSV live-attenuated vaccine candidate by substituting 4 unique residues from the RSV line19F protein into the F protein of the heterologous vaccine strain DB1. The resultant vaccine candidate, DB1-QUAD, was thermostable, attenuatedin vivo, highly immunogenic, and protective against RSV challenge in mice and cotton rats.

scholarly journals Evolution of protection after maternal immunization for respiratory syncytial virus in cotton rats

2021 ◽  
Author(s):  
Jorge C.G. Blanco ◽  
Lori McGinnes-Cullen ◽  
Arash Kamali ◽  
Fatoumata Sylla ◽  
Marina Boukhavalova ◽  
...  
Keyword(s):  
Respiratory Syncytial Virus ◽  
Neutralizing Antibody ◽  
Wild Type ◽  
Cotton Rat ◽  
Before And After ◽  
Boost Immunization ◽  
Cotton Rats ◽  
Antibody Titers ◽  
Syncytial Virus ◽  
Efficient Transfer

Maternal anti-respiratory syncytial virus (RSV) antibodies acquired by the fetus through the placenta protect neonates from RSV disease through the first weeks of life.  In the cotton rat model of RSV infections, we previously reported that immunization of dams during pregnancy with virus-like particles assembled with mutation stabilized pre-fusion F protein as well as the wild type G protein resulted in robust protection of their offspring from RSV challenge (Blanco, et al Journal of Virology 93: e00914-19, https://doi.org/10.1128/JVI.00914-19).  Here we describe the durability of those protective responses in dams, the durability of protection in offspring, and the transfer of that protection to offspring of two consecutive pregnancies without a second boost immunization.  We report that four weeks after birth, offspring of the first pregnancy were significantly protected from RSV replication in both lungs and nasal tissues after RSV challenge, but protection was reduced in pups at 6 weeks after birth.   However, the overall protection of offspring of the second pregnancy was considerably reduced, even at four weeks of age.  This drop in protection occurred even though the levels of total anti-pre-F IgG and neutralizing antibody titers in dams remained at similar, high levels before and after the second pregnancy.  The results are consistent with an evolution of antibody properties in dams to populations less efficiently transferred to offspring or the less efficient transfer of antibodies in elderly dams.

scholarly journals Eosinophils contribute to innate antiviral immunity and promote clearance of respiratory syncytial virus

Blood ◽  
2007 ◽  
Vol 110 (5) ◽  
pp. 1578-1586 ◽  
Author(s):  
Simon Phipps ◽  
Chuan En Lam ◽  
Suresh Mahalingam ◽  
Matthew Newhouse ◽  
Ruben Ramirez ◽  
...  
Keyword(s):  
Respiratory Syncytial Virus ◽  
Host Defense ◽  
No Production ◽  
Wild Type ◽  
Interleukin 5 ◽  
Virus Clearance ◽  
Lung Dysfunction ◽  
Rsv Infection ◽  
Syncytial Virus

AbstractEosinophils are recruited to the lungs in response to respiratory syncytial virus (RSV) infection; however, their role in promoting antiviral host defense remains unclear. Here, we demonstrate that eosinophils express TLRs that recognize viral nucleic acids, are activated and degranulate after single-stranded RNA (ssRNA) stimulation of the TLR-7–MyD88 pathway, and provide host defense against RSV that is MyD88 dependent. In contrast to wild-type mice, virus clearance from lung tissue was more rapid in hypereosinophilic (interleukin-5 transgenic) mice. Transfer of wild-type but not MyD88-deficient eosinophils to the lungs of RSV-infected wild-type mice accelerated virus clearance and inhibited the development of airways hyperreactivity. Similar responses were observed when infected recipient mice were MyD88 deficient. Eosinophils isolated from infected hypereosinophilic MyD88-sufficient but not MyD88-deficient mice expressed greater amounts of IFN regulatory factor (IRF)–7 and eosinophil-associated ribonucleases EAR-1 and EAR-2. Hypereosinophilia in the airways of infected mice also correlated with increased expression of IRF-7, IFN-β, and NOS-2, and inhibition of NO production with the NOS-2 inhibitor L-NMA partially reversed the accelerated virus clearance promoted by eosinophils. Collectively, our results demonstrate that eosinophils can protect against RSV in vivo, as they promote virus clearance and may thus limit virus-induced lung dysfunction.

scholarly journals CD4+T Cells Drive Lung Disease Enhancement Induced by Immunization with Suboptimal Doses of Respiratory Syncytial Virus Fusion Protein in the Mouse Model

2019 ◽  
Vol 93 (15) ◽  
Author(s):  
Kirsten Schneider-Ohrum ◽  
Angie Snell Bennett ◽  
Gaurav Manohar Rajani ◽  
Leigh Hostetler ◽  
Sean K. Maynard ◽  
...  
Keyword(s):  
T Cells ◽  
Respiratory Syncytial Virus ◽  
Mouse Model ◽  
Fusion Protein ◽  
Lung Disease ◽  
Rat Model ◽  
Preclinical Models ◽  
Cotton Rat ◽  
Rsv Infection ◽  
Syncytial Virus

ABSTRACTRespiratory syncytial virus (RSV) infection of seronegative children previously immunized with formalin-inactivated (FI) RSV has been associated with serious enhanced respiratory disease (ERD). The phenomenon was reproduced in the cotton rat and the mouse, and both preclinical models have been routinely used to evaluate the safety of new RSV vaccine candidates. More recently, we demonstrated that immunizations with suboptimal doses of the RSV fusion (F) antigen, in its post- or prefusion conformation, and in the presence of a Th1-biasing adjuvant, unexpectedly led to ERD in the cotton rat model. To assess if those observations are specific to the cotton rat and to elucidate the mechanism by which vaccination with low antigen doses can drive ERD post-RSV challenge, we evaluated RSV post-F antigen dose de-escalation in BALB/c mice in the presence of a Th1-biasing adjuvant. While decreasing antigen doses, we observed an increase in lung inflammation associated with an upregulation of proinflammatory cytokines. The amplitude of the lung histopathology was comparable to that of FI-RSV-induced ERD, confirming the observations made in the cotton rat. Importantly, depletion of CD4+T cells prior to viral challenge completely abrogated ERD, preventing proinflammatory cytokine upregulation and the infiltration of T cells, neutrophils, eosinophils, and macrophages into the lung. Overall, low-antigen-dose-induced ERD resembles FI-RSV-induced ERD, except that the former appears in the absence of detectable levels of viral replication and in the context of a Th1-biased immune response. Taken together, our observations reinforce the recent concept that vaccines developed for RSV-naïve individuals should be systematically tested under suboptimal dosing conditions.IMPORTANCERSV poses a significant health care burden and is the leading cause of serious lower-respiratory-tract infections in young children. A formalin-inactivated RSV vaccine developed in the 1960s not only showed a complete lack of efficacy against RSV infection but also induced severe lung disease enhancement in vaccinated children. Since then, establishing safety in preclinical models has been one of the major challenges to RSV vaccine development. We recently observed in the cotton rat model that suboptimal immunizations with RSV fusion protein could induce lung disease enhancement. In the present study, we extended suboptimal dosing evaluation to the mouse model. We confirmed the induction of lung disease enhancement by vaccinations with low antigen doses and dissected the associated immune mechanisms. Our results stress the need to evaluate suboptimal dosing for any new RSV vaccine candidate developed for seronegative infants.

scholarly journals Packaging and Prefusion Stabilization Separately and Additively Increase the Quantity and Quality of Respiratory Syncytial Virus (RSV)-Neutralizing Antibodies Induced by an RSV Fusion Protein Expressed by a Parainfluenza Virus Vector

2016 ◽  
Vol 90 (21) ◽  
pp. 10022-10038 ◽  
Author(s):  
Bo Liang ◽  
Joan O. Ngwuta ◽  
Richard Herbert ◽  
Joanna Swerczek ◽  
David W. Dorward ◽  
...  
Keyword(s):  
Respiratory Syncytial Virus ◽  
Rhesus Monkeys ◽  
Neutralizing Antibodies ◽  
Vaccine Candidate ◽  
Parainfluenza Virus ◽  
F Protein ◽  
Syncytial Virus ◽  
Human Parainfluenza Virus ◽  
Type 3

ABSTRACTHuman respiratory syncytial virus (RSV) and human parainfluenza virus type 3 (HPIV3) are major pediatric respiratory pathogens that lack vaccines. A chimeric bovine/human PIV3 (rB/HPIV3) virus expressing the unmodified, wild-type (wt) RSV fusion (F) protein from an added gene was previously evaluated in seronegative children as a bivalent intranasal RSV/HPIV3 vaccine, and it was well tolerated but insufficiently immunogenic for RSV F. We recently showed that rB/HPIV3 expressing a partially stabilized prefusion form (pre-F) of RSV F efficiently induced “high-quality” RSV-neutralizing antibodies, defined as antibodies that neutralize RSVin vitrowithout added complement (B. Liang et al., J Virol 89:9499–9510, 2015, doi:10.1128/JVI.01373-15). In the present study, we modified RSV F by replacing its cytoplasmic tail (CT) domain or its CT and transmembrane (TM) domains (TMCT) with counterparts from BPIV3 F, with or without pre-F stabilization. This resulted in RSV F being packaged in the rB/HPIV3 particle with an efficiency similar to that of RSV particles. Enhanced packaging was substantially attenuating in hamsters (10- to 100-fold) and rhesus monkeys (100- to 1,000-fold). Nonetheless, TMCT-directed packaging substantially increased the titers of high-quality RSV-neutralizing serum antibodies in hamsters. In rhesus monkeys, a strongly additive immunogenic effect of packaging and pre-F stabilization was observed, as demonstrated by 8- and 30-fold increases of RSV-neutralizing serum antibody titers in the presence and absence of added complement, respectively, compared to pre-F stabilization alone. Analysis of vaccine-induced F-specific antibodies by binding assays indicated that packaging conferred substantial stabilization of RSV F in the pre-F conformation. This provides an improved version of this well-tolerated RSV/HPIV3 vaccine candidate, with potently improved immunogenicity, which can be returned to clinical trials.IMPORTANCEHuman respiratory syncytial virus (RSV) and human parainfluenza virus type 3 (HPIV3) are major viral agents of acute pediatric bronchiolitis and pneumonia worldwide that lack vaccines. A bivalent intranasal RSV/HPIV3 vaccine candidate consisting of a chimeric bovine/human PIV3 (rB/HPIV3) strain expressing the RSV fusion (F) protein was previously shown to be well tolerated by seronegative children but was insufficiently immunogenic for RSV F. In the present study, the RSV F protein was engineered to be packaged efficiently into vaccine virus particles. This resulted in a significantly enhanced quantity and quality of RSV-neutralizing antibodies in hamsters and nonhuman primates. In nonhuman primates, this effect was strongly additive to the previously described stabilization of the prefusion conformation of the F protein. The improved immunogenicity of RSV F by packaging appeared to involve prefusion stabilization. These findings provide a potently more immunogenic version of this well-tolerated vaccine candidate and should be applicable to other vectored vaccines.

scholarly journals Single Immunization of a Vaccine Vectored by a Novel Recombinant Vaccinia Virus Affords Effective Protection Against Respiratory Syncytial Virus Infection in Cotton Rats

2021 ◽  
Vol 12 ◽  
Author(s):  
Marsha S. Russell ◽  
Sathya N. Thulasi Raman ◽  
Caroline Gravel ◽  
Wanyue Zhang ◽  
Annabelle Pfeifle ◽  
...  
Keyword(s):  
Respiratory Syncytial Virus ◽  
Vaccinia Virus ◽  
High Efficiency ◽  
Vaccine Development ◽  
Nasal Administration ◽  
Effective Vaccine ◽  
High Dose ◽  
Cotton Rats ◽  
Syncytial Virus ◽  
Vectored Vaccine

Respiratory syncytial virus (RSV) is a leading cause of respiratory infections worldwide and disease management measures are hampered by the lack of a safe and effective vaccine against the infection. We constructed a novel recombinant RSV vaccine candidate based on a deletion mutant vaccinia virus platform, in that the host range genes E3L and K3L were deleted (designated as VACVΔE3LΔK3L) and a poxvirus K3L ortholog gene was used as a marker for the rapid and efficient selection of recombinant viruses. The safety of the modified vaccinia virus was investigated by intranasal administration of BALB/c mice with the modified vaccinia vector using a dose known to be lethal in the wild-type Western Reserve. Only a minor loss of body weight by less than 5% and mild pulmonary inflammation were observed, both of which were transient in nature following nasal administration of the high-dose modified vaccinia virus. In addition, the viruses were cleared from the lung in 2 days with no viral invasions of the brain and other vital organs. These results suggest that the virulence of the virus has been essentially abolished. We then investigated the efficiency of the vector for the delivery of vaccines against RSV through comparison with another RSV vaccine delivered by the widely used Modified Vaccinia virus Ankara (MVA) backbone. In the cotton rats, we found a single intramuscular administration of VACVΔE3LΔK3L-vectored vaccine elicited immune responses and protection at a level comparable to the MVA-vectored vaccine against RSV infection. The distinct features of this novel VACV vector, such as an E3L deletion for attenuation and a K3L ortholog for positive selection and high efficiency for vaccine delivery, could provide unique advantages to the application of VACV as a platform for vaccine development.

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