Monoterpene-Containing Substituted Coumarins as Inhibitors of Respiratory Syncytial Virus (RSV) Replication

Respiratory syncytial virus (RSV) is a critical cause of infant mortality. However, there are no vaccines and adequate drugs for its treatment. We showed, for the first time, that O-linked coumarin–monoterpene conjugates are effective RSV inhibitors. The most potent compounds are active against both RSV serotypes, A and B. According to the results of the time-of-addition experiment, the conjugates act at the early stages of virus cycle. Based on molecular modelling data, RSV F protein may be considered as a possible target.

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GENETIC AND ANTIGENIC CHARACTERISTICS OF RESPIRATORY SYNCYTIAL VIRUS STRAINS ISOLATED IN ST. PETERSBURG IN 2013-2016

Voprosy Virusologii ◽

10.18821/0507-4088-2017-62-6-273-282 ◽

2017 ◽

Vol 62 (6) ◽

pp. 273-282

Author(s):

V. Z. Krivitskaya ◽

K. S. Sintsova ◽

E. R. Petrova ◽

M. V. Sverlova ◽

E. V. Sorokin ◽

...

Keyword(s):

Monoclonal Antibodies ◽

Respiratory Syncytial Virus ◽

Reference Strain ◽

Antigenic Site ◽

Genetic Characteristics ◽

F Protein ◽

Reference Virus ◽

Syncytial Virus ◽

First Time ◽

Virus Strains

Antigenic and genetic characteristics of Russian RSV isolates are presented for the first time. Of the 69 strains isolated in St. Petersburg, 93% belonged to the RSV-A antigenic group. The antigenic variations in the F-protein RSV were analyzed using a panel from 6 monoclonal antibodies by the method of micro-cultural ELISA. Depending on the decrease in the effectiveness of interaction with monoclonal antibodies (relative to the reference strain Long), RSV-A isolates were divided into 4 antigenic subgroups. The results of 24 isolates sequencing showed that more than 60% of them had substitutions in significant F-protein sites compared to the ON67-1210A reference strain of the current RSV genotype ON1/GA2. The most variable were the signal peptide and antigenic site II. When comparing the results of ELISA and sequencing, it was not possible to identify any specific key substitutions in the amino acid sequence of the F-protein that affect the interaction of the virus with antibodies. The nucleotide sequence of the F-gene from 19 of the 24 characterized isolates was close to that of ON67-1210A reference virus and was significantly different from RSV-A Long and A2 viruses. A separate group consisted of 5 strains, in which the F-protein structure was approximated to RSV Long.

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Shifting immunodominance pattern of two cytotoxic T-lymphocyte epitopes in the F glycoprotein of the Long strain of respiratory syncytial virus

Journal of General Virology ◽

10.1099/vir.0.80219-0 ◽

2004 ◽

Vol 85 (11) ◽

pp. 3229-3238 ◽

Cited By ~ 28

Author(s):

Carolina Johnstone ◽

Patricia de León ◽

Francisco Medina ◽

José A. Melero ◽

Blanca García-Barreno ◽

...

Keyword(s):

Respiratory Syncytial Virus ◽

Vaccinia Virus ◽

T Lymphocyte ◽

Cytotoxic T Lymphocyte ◽

Recombinant Vaccinia Virus ◽

Recombinant Vaccinia ◽

F Protein ◽

Syncytial Virus ◽

Lymphocyte Responses

Human respiratory syncytial virus (RSV) is a major cause of respiratory infection in children and in the elderly. The RSV fusion (F) glycoprotein has long been recognized as a vaccine candidate as it elicits cytotoxic T-lymphocyte (CTL) and antibody responses. Two murine H-2Kd-restricted CTL epitopes (F85–93 and F92–106) are known in the F protein of the A2 strain of RSV. F-specific CTL lines using BCH4 fibroblasts that are persistently infected with the Long strain of human RSV as stimulators were generated, and it was found that in this strain only the F85–93 epitope is conserved. Motif based epitope prediction programs and an F2 chain deleted F protein encoded in a recombinant vaccinia virus enabled identification of a new epitope in the Long strain, F249–258, which is presented by Kd as a 9-mer (TYMLTNSEL) or a 10-mer (TYMLTNSELL) peptide. The results suggest that the 10-mer might be a naturally processed endogenous Kd ligand. The CD8+ T-lymphocyte responses to epitopes F85–93 and F249–258 present in the F protein of RSV Long were found to be strongly skewed to F85–93 in in vitro multispecific CTL lines and in vivo during a secondary response to a recombinant vaccinia virus that expresses the entire F protein. However, no hierarchy in CD8+ T-lymphocyte responses to F85–93 and F249–258 epitopes was observed in vivo during a primary response.

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Immunogenicity and efficacy of codon optimized DNA vaccines encoding the F-protein of respiratory syncytial virus

Vaccine ◽

10.1016/j.vaccine.2007.07.025 ◽

2007 ◽

Vol 25 (41) ◽

pp. 7271-7279 ◽

Cited By ~ 47

Author(s):

Nicola Ternette ◽

Bettina Tippler ◽

Klaus Überla ◽

Thomas Grunwald

Keyword(s):

Respiratory Syncytial Virus ◽

Dna Vaccines ◽

F Protein ◽

Syncytial Virus

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Reduction of Respiratory Syncytial Virus (RSV) in Tracheal Aspirates in Intubated Infants by Use of Humanized Monoclonal Antibody to RSV F Protein

The Journal of Infectious Diseases ◽

10.1086/314523 ◽

1998 ◽

Vol 178 (6) ◽

pp. 1555-1561 ◽

Cited By ~ 148

Author(s):

Richard Malley ◽

John DeVincenzo ◽

Octavio Ramilo ◽

Penelope H. Dennehy ◽

H. Cody Meissner ◽

...

Keyword(s):

Monoclonal Antibody ◽

Respiratory Syncytial Virus ◽

F Protein ◽

Humanized Monoclonal Antibody ◽

Syncytial Virus ◽

Tracheal Aspirates

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Enhanced Neutralizing Antibody Response Induced by Respiratory Syncytial Virus Prefusion F Protein Expressed by a Vaccine Candidate

Journal of Virology ◽

10.1128/jvi.01373-15 ◽

2015 ◽

Vol 89 (18) ◽

pp. 9499-9510 ◽

Cited By ~ 37

Author(s):

Bo Liang ◽

Sonja Surman ◽

Emerito Amaro-Carambot ◽

Barbora Kabatova ◽

Natalie Mackow ◽

...

Keyword(s):

Respiratory Syncytial Virus ◽

Clinical Evaluation ◽

Genetic Stability ◽

Vaccine Candidate ◽

Early Passage ◽

F Protein ◽

Syncytial Virus ◽

Human Parainfluenza Virus ◽

Type 3

ABSTRACTRespiratory syncytial virus (RSV) and human parainfluenza virus type 3 (HPIV3) are the first and second leading viral agents of severe respiratory tract disease in infants and young children worldwide. Vaccines are not available, and an RSV vaccine is particularly needed. A live attenuated chimeric recombinant bovine/human PIV3 (rB/HPIV3) vector expressing the RSV fusion (F) glycoprotein from an added gene has been under development as a bivalent vaccine against RSV and HPIV3. Previous clinical evaluation of this vaccine candidate suggested that increased genetic stability and immunogenicity of the RSV F insert were needed. This was investigated in the present study. RSV F expression was enhanced 5-fold by codon optimization and by modifying the amino acid sequence to be identical to that of an early passage of the original clinical isolate. This conferred a hypofusogenic phenotype that presumably reflects the original isolate. We then compared vectors expressing stabilized prefusion and postfusion versions of RSV F. In a hamster model, prefusion F induced increased quantity and quality of RSV-neutralizing serum antibodies and increased protection against wild-type (wt) RSV challenge. In contrast, a vector expressing the postfusion F was less immunogenic and protective. The genetic stability of the RSV F insert was high and was not affected by enhanced expression or the prefusion or postfusion conformation of RSV F. These studies provide an improved version of the previously well-tolerated rB/HPIV3-RSV F vaccine candidate that induces a superior RSV-neutralizing serum antibody response.IMPORTANCERespiratory syncytial virus (RSV) and human parainfluenza virus type 3 (HPIV3) are two major causes of pediatric pneumonia and bronchiolitis. The rB/HPIV3 vector expressing RSV F protein is a candidate bivalent live vaccine against HPIV3 and RSV. Previous clinical evaluation indicated the need to increase the immunogenicity and genetic stability of the RSV F insert. Here, we increased RSV F expression by codon optimization and by modifying the RSV F amino acid sequence to conform to that of an early passage of the original isolate. This resulted in a hypofusogenic phenotype, which likely represents the original phenotype before adaptation to cell culture. We also included stabilized versions of prefusion and postfusion RSV F protein. Prefusion RSV F induced a larger quantity and higher quality of RSV-neutralizing serum antibodies and was highly protective. This provides an improved candidate for further clinical evaluation.

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Respiratory Syncytial Virus (RSV) Diseases

10.33442/vt202160 ◽

2022 ◽

Author(s):

Heinz-Josef Schmitt ◽

Khrystyna Hrynkevych

Keyword(s):

Respiratory Syncytial Virus ◽

Rna Virus ◽

Passive Immunization ◽

Active Immunization ◽

F Protein ◽

Hospitalization Rates ◽

Syncytial Virus ◽

Repeated Infections ◽

Long Acting ◽

Underlying Diseases

The respiratory syncytial virus (RSV) is an RNA virus that causes annual ARI outbreaks during winter with mild URTI in the general population, but with severe LRTI particularly among young children (bronchiolitis), patients with underlying diseases and people >65 years of age. RSV does not induce a long-lasting protective immunity and repeated infections throughout life are the norm. Basically, all children have been infected by 2 years of age and of those hospitalized, >50% are <3 months and 75% are <6 months of age. The overall CFR is 1/500. For adults ≥65 years, RSV hospitalization rates are 90–250/105. There is no specific therapy, general preventive measures include general hygiene and isolation/separation of patients. A monoclonal anti-F-protein antibody is available for passive immunization of selected high-risk children. It requires monthly injections, comes at a high cost and has limited efficacy (50% against RSV hospitalization). Active immunization failed in the past, probably as the post-fusion conformation of the F-protein was used. Long-acting monoclonal antibodies (for infants) as well as stabilized pre-fusion F-protein vaccines (for immunization of pregnant women, children, older adults) produced on various platforms are in late stages of clinical development.

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Characterization of Pre-F-GCN4t, a Modified Human Respiratory Syncytial Virus Fusion Protein Stabilized in a Noncleaved Prefusion Conformation

Journal of Virology ◽

10.1128/jvi.02437-16 ◽

2017 ◽

Vol 91 (13) ◽

Cited By ~ 18

Author(s):

Normand Blais ◽

Martin Gagné ◽

Yoshitomo Hamuro ◽

Patrick Rheault ◽

Martine Boyer ◽

...

Keyword(s):

Respiratory Syncytial Virus ◽

Antibody Response ◽

Neutralizing Antibodies ◽

Neutralizing Antibody ◽

Human Respiratory Syncytial Virus ◽

Vaccine Antigen ◽

Significant Advance ◽

F Protein ◽

Syncytial Virus

ABSTRACT The human respiratory syncytial virus (hRSV) fusion (F) protein is considered a major target of the neutralizing antibody response to hRSV. This glycoprotein undergoes a major structural shift from the prefusion (pre-F) to the postfusion (post-F) state at the time of virus-host cell membrane fusion. Recent evidences suggest that the pre-F state is a superior target for neutralizing antibodies compared to the post-F state. Therefore, for vaccine purposes, we have designed and characterized a recombinant hRSV F protein, called Pre-F-GCN4t, stabilized in a pre-F conformation. To show that Pre-F-GCN4t does not switch to a post-F conformation, it was compared with a recombinant post-F molecule, called Post-F-XC. Pre-F-GCN4t was glycosylated and trimeric and displayed a conformational stability different from that of Post-F-XC, as shown by chemical denaturation. Electron microscopy analysis suggested that Pre-F-GCN4t adopts a lollipop-like structure. In contrast, Post-F-XC had a typical elongated conical shape. Hydrogen/deuterium exchange mass spectrometry demonstrated that the two molecules had common rigid folding core and dynamic regions and provided structural insight for their biophysical and biochemical properties and reactivity. Pre-F-GCN4t was shown to deplete hRSV-neutralizing antibodies from human serum more efficiently than Post-F-XC. Importantly, Pre-F-GCN4t was also shown to bind D25, a highly potent monoclonal antibody specific for the pre-F conformation. In conclusion, this construct presents several pre-F characteristics, does not switch to the post-F conformation, and presents antigenic features required for a protective neutralizing antibody response. Therefore, Pre-F-GCN4t can be considered a promising candidate vaccine antigen. IMPORTANCE Human respiratory syncytial virus (RSV) is a global leading cause of infant mortality and adult morbidity. The development of a safe and efficacious RSV vaccine remains an important goal. The RSV class I fusion (F) glycoprotein is considered one of the most promising vaccine candidates, and recent evidences suggest that the prefusion (pre-F) state is a superior target for neutralizing antibodies. Our study presents the physicochemical characterization of Pre-F-GCN4t, a molecule designed to be stabilized in the pre-F conformation. To confirm its pre-F conformation, Pre-F-GCN4t was analyzed in parallel with Post-F-XC, a molecule in the post-F conformation. Our results show that Pre-F-GCN4t presents characteristics of a stabilized pre-F conformation and support its use as an RSV vaccine antigen. Such an antigen may represent a significant advance in the development of an RSV vaccine.

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The respiratory syncytial virus (RSV) prefusion F‐protein functional antibody repertoire in adult healthy donors

EMBO Molecular Medicine ◽

10.15252/emmm.202114035 ◽

2021 ◽

Author(s):

Emanuele Andreano ◽

Ida Paciello ◽

Monia Bardelli ◽

Simona Tavarini ◽

Chiara Sammicheli ◽

...

Keyword(s):

Respiratory Syncytial Virus ◽

Antibody Repertoire ◽

F Protein ◽

Healthy Donors ◽

Syncytial Virus

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Respiratory syncytial virus co-opts host mitochondrial function to favour infectious virus production

eLife ◽

10.7554/elife.42448 ◽

2019 ◽

Vol 8 ◽

Cited By ~ 8

Author(s):

MengJie Hu ◽

Keith E Schulze ◽

Reena Ghildyal ◽

Darren C Henstridge ◽

Jacek L Kolanowski ◽

...

Keyword(s):

Respiratory Syncytial Virus ◽

Membrane Potential ◽

Mitochondrial Membrane Potential ◽

Mitochondrial Membrane ◽

Quantitative Imaging ◽

Virus Production ◽

Dynein Motor ◽

Mitochondrial Ros ◽

Syncytial Virus ◽

First Time

Although respiratory syncytial virus (RSV) is responsible for more human deaths each year than influenza, its pathogenic mechanisms are poorly understood. Here high-resolution quantitative imaging, bioenergetics measurements and mitochondrial membrane potential- and redox-sensitive dyes are used to define RSV’s impact on host mitochondria for the first time, delineating RSV-induced microtubule/dynein-dependent mitochondrial perinuclear clustering, and translocation towards the microtubule-organizing centre. These changes are concomitant with impaired mitochondrial respiration, loss of mitochondrial membrane potential and increased production of mitochondrial reactive oxygen species (ROS). Strikingly, agents that target microtubule integrity the dynein motor protein, or inhibit mitochondrial ROS production strongly suppresses RSV virus production, including in a mouse model with concomitantly reduced virus-induced lung inflammation. The results establish RSV’s unique ability to co-opt host cell mitochondria to facilitate viral infection, revealing the RSV-mitochondrial interface for the first time as a viable target for therapeutic intervention.

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