Development of an Alternative Modified Live Influenza B Virus Vaccine

ABSTRACT Influenza B virus (IBV) is considered a major human pathogen, responsible for seasonal epidemics of acute respiratory illness. Two antigenically distinct IBV hemagglutinin (HA) lineages cocirculate worldwide with little cross-reactivity. Live attenuated influenza virus (LAIV) vaccines have been shown to provide better cross-protective immune responses than inactivated vaccines by eliciting local mucosal immunity and systemic B cell- and T cell-mediated memory responses. We have shown previously that incorporation of temperature-sensitive (ts) mutations into the PB1 and PB2 subunits along with a modified HA epitope tag in the C terminus of PB1 resulted in influenza A viruses (IAV) that are safe and effective as modified live attenuated (att) virus vaccines (IAV att). We explored whether analogous mutations in the IBV polymerase subunits would result in a stable virus with an att phenotype. The PB1 subunit of the influenza B/Brisbane/60/2008 strain was used to incorporate ts mutations and a C-terminal HA tag. Such modifications resulted in a B/Bris att strain with ts characteristics in vitro and an att phenotype in vivo. Vaccination studies in mice showed that a single dose of the B/Bris att candidate stimulated sterilizing immunity against lethal homologous challenge and complete protection against heterologous challenge. These studies show the potential of an alternative LAIV platform for the development of IBV vaccines. IMPORTANCE A number of issues with regard to the effectiveness of the LAIV vaccine licensed in the United States (FluMist) have arisen over the past three seasons (2013–2014, 2014–2015, and 2015–2016). While the reasons for the limited robustness of the vaccine-elicited immune response remain controversial, this problem highlights the critical importance of continued investment in LAIV development and creates an opportunity to improve current strategies so as to develop more efficacious vaccines. Our laboratory has developed an alternative strategy, the incorporation of 2 amino acid mutations and a modified HA tag at the C terminus of PB1, which is sufficient to attenuate the IBV. As a LAIV, this novel vaccine provides complete protection against IBV strains. The availability of attenuated IAV and IBV backbones based on contemporary strains offers alternative platforms for the development of LAIVs that may overcome current limitations.

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Influenza B Virus NS1-Truncated Mutants: Live-Attenuated Vaccine Approach

Journal of Virology ◽

10.1128/jvi.01213-08 ◽

2008 ◽

Vol 82 (21) ◽

pp. 10580-10590 ◽

Cited By ~ 73

Author(s):

Rong Hai ◽

Luis Martínez-Sobrido ◽

Kathryn A. Fraser ◽

Juan Ayllon ◽

Adolfo García-Sastre ◽

...

Keyword(s):

Reverse Genetics ◽

Influenza Viruses ◽

Influenza B Virus ◽

Influenza B ◽

Type I ◽

Live Attenuated Vaccine ◽

Live Virus ◽

B Virus

ABSTRACT Type B influenza viruses can cause substantial morbidity and mortality in the population, and vaccination remains by far the best means of protection against infections with these viruses. Here, we report the construction of mutant influenza B viruses for potential use as improved live-virus vaccine candidates. Employing reverse genetics, we altered the NS1 gene, which encodes a type I interferon (IFN) antagonist. The resulting NS1 mutant viruses induced IFN and, as a consequence, were found to be attenuated in vitro and in vivo. The absence of pathogenicity of the NS1 mutants in both BALB/c and C57BL/6 PKR−/− mice was confirmed. We also provide evidence that influenza B virus NS1 mutants induce a self-adjuvanted immune response and confer effective protection against challenge with both homologous and heterologous B virus strains in mice.

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FluB-RAM and FluB-RANS: Genome Rearrangement as Safe and Efficacious Live Attenuated Influenza B Virus Vaccines

Vaccines ◽

10.3390/vaccines9080897 ◽

2021 ◽

Vol 9 (8) ◽

pp. 897

Author(s):

Stivalis Cardenas-Garcia ◽

C. Joaquín Cáceres ◽

Aarti Jain ◽

Ginger Geiger ◽

Jong-Suk Mo ◽

...

Keyword(s):

Genome Rearrangement ◽

Natural Infection ◽

The Elderly ◽

Respiratory Pathogen ◽

Influenza B Virus ◽

Influenza B ◽

B Virus ◽

Cellular Immune

Influenza B virus (IBV) is considered a major respiratory pathogen responsible for seasonal respiratory disease in humans, particularly severe in children and the elderly. Seasonal influenza vaccination is considered the most efficient strategy to prevent and control IBV infections. Live attenuated influenza virus vaccines (LAIVs) are thought to induce both humoral and cellular immune responses by mimicking a natural infection, but their effectiveness has recently come into question. Thus, the opportunity exists to find alternative approaches to improve overall influenza vaccine effectiveness. Two alternative IBV backbones were developed with rearranged genomes, rearranged M (FluB-RAM) and a rearranged NS (FluB-RANS). Both rearranged viruses showed temperature sensitivity in vitro compared with the WT type B/Bris strain, were genetically stable over multiple passages in embryonated chicken eggs and were attenuated in vivo in mice. In a prime-boost regime in naïve mice, both rearranged viruses induced antibodies against HA with hemagglutination inhibition titers considered of protective value. In addition, antibodies against NA and NP were readily detected with potential protective value. Upon lethal IBV challenge, mice previously vaccinated with either FluB-RAM or FluB-RANS were completely protected against clinical disease and mortality. In conclusion, genome re-arrangement renders efficacious LAIV candidates to protect mice against IBV.

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A Reverse Genetics Approach for Recovery of Recombinant Influenza B Viruses Entirely from cDNA

Journal of Virology ◽

10.1128/jvi.76.22.11744-11747.2002 ◽

2002 ◽

Vol 76 (22) ◽

pp. 11744-11747 ◽

Cited By ~ 53

Author(s):

David Jackson ◽

Andrew Cadman ◽

Thomas Zurcher ◽

Wendy S. Barclay

Keyword(s):

Amino Acid ◽

Influenza A ◽

Amino Acid Change ◽

Reverse Genetics ◽

Single Gene ◽

Neuraminidase Inhibitor ◽

Influenza B Virus ◽

Influenza B ◽

B Virus

ABSTRACT The recovery of recombinant influenza A virus entirely from cDNA was recently described (9, 19). We adapted the technique for engineering influenza B virus and generated a mutant bearing an amino acid change E116G in the viral neuraminidase which was resistant in vitro to the neuraminidase inhibitor zanamivir. The method also facilitates rapid isolation of single-gene reassortants suitable as vaccine seeds and will aid further investigations of unique features of influenza B virus.

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The NB Protein of Influenza B Virus Is Not Necessary for Virus Replication In Vitro

Journal of Virology ◽

10.1128/jvi.77.10.6050-6054.2003 ◽

2003 ◽

Vol 77 (10) ◽

pp. 6050-6054 ◽

Cited By ~ 46

Author(s):

Masato Hatta ◽

Yoshihiro Kawaoka

Keyword(s):

Cell Culture ◽

Virus Replication ◽

Wild Type Virus ◽

Influenza B Virus ◽

Influenza B ◽

Type Virus ◽

Wild Type ◽

B Virus

ABSTRACT The NB protein of influenza B virus is thought to function as an ion channel and therefore would be expected to have an essential function in viral replication. Because direct evidence for its absolute requirement in the viral life cycle is lacking, we generated NB knockout viruses by reverse genetics and tested their growth properties both in vitro and in vivo. Mutants not expressing NB replicated as efficiently as the wild-type virus in cell culture, whereas in mice they showed restricted growth compared with findings for the wild-type virus. Thus, the NB protein is not essential for influenza B virus replication in cell culture but promotes efficient growth in mice.

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Experimental Infection Using Mouse-Adapted Influenza B Virus in a Mouse Model

Viruses ◽

10.3390/v12040470 ◽

2020 ◽

Vol 12 (4) ◽

pp. 470 ◽

Cited By ~ 1

Author(s):

Elena Prokopyeva ◽

Olga Kurskaya ◽

Ivan Sobolev ◽

Mariia Solomatina ◽

Tatyana Murashkina ◽

...

Keyword(s):

Influenza Vaccine ◽

Drug Testing ◽

Protective Efficacy ◽

Influenza B Virus ◽

Infection Model ◽

Influenza B ◽

Post Inoculation ◽

B Virus

Every year, influenza B viruses (IBVs) contribute to annual illness, and infection can lead to serious respiratory disease among humans. More attention is needed in several areas, such as increasing virulence or pathogenicity of circulating B viruses and developing vaccines against current influenza. Since preclinical trials of anti-influenza drugs are mainly conducted in mice, we developed an appropriate infection model, using an antigenically-relevant IBV strain, for furtherance of anti-influenza drug testing and influenza vaccine protective efficacy analysis. A Victoria lineage (clade 1A) IBV was serially passaged 17 times in BALB/c mice, and adaptive amino acid substitutions were found in hemagglutinin (HA) (T214I) and neuraminidase (NA) (D432N). By electron microscopy, spherical and elliptical IBV forms were noted. Light microscopy showed that mouse-adapted IBVs caused influenza pneumonia on day 6 post inoculation. We evaluated the illness pathogenicity, viral load, and histopathological features of mouse-adapted IBVs and estimated anti-influenza drugs and vaccine efficiency in vitro and in vivo. Assessment of an investigational anti-influenza drug (oseltamivir ethoxysuccinate) and an influenza vaccine (Ultrix®, SPBNIIVS, Saint Petersburg, Russia) showed effectiveness against the mouse-adapted influenza B virus.

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Defective interfering influenza A virus protects in vivo against disease caused by a heterologous influenza B virus

Journal of General Virology ◽

10.1099/vir.0.034132-0 ◽

2011 ◽

Vol 92 (9) ◽

pp. 2122-2132 ◽

Cited By ~ 27

Author(s):

Paul D. Scott ◽

Bo Meng ◽

Anthony C. Marriott ◽

Andrew J. Easton ◽

Nigel J. Dimmock

Keyword(s):

Influenza A Virus ◽

Influenza A ◽

Influenza Viruses ◽

Influenza B Virus ◽

Influenza B ◽

Type I ◽

Type I Ifn ◽

B Virus ◽

The Family

Influenza A and B viruses are major human respiratory pathogens that contribute to the burden of seasonal influenza. They are both members of the family Orthomyxoviridae but do not interact genetically and are classified in different genera. Defective interfering (DI) influenza viruses have a major deletion of one or more of their eight genome segments, which renders them both non-infectious and able to interfere in cell culture with the production of infectious progeny by a genetically compatible, homologous virus. It has been shown previously that intranasal administration of a cloned DI influenza A virus, 244/PR8, protects mice from various homologous influenza A virus subtypes and that it also protects mice from respiratory disease caused by a heterologous virus belonging to the family Paramyxoviridae. The mechanisms of action in vivo differ, with homologous and heterologous protection being mediated by probable genome competition and type I interferon (IFN), respectively. In the current study, it was shown that 244/PR8 also protects against disease caused by a heterologous influenza B virus (B/Lee/40). Protection from B/Lee/40 challenge was partially eliminated in mice that did not express a functional type I IFN receptor, suggesting that innate immunity, and type I IFN in particular, are important in mediating protection against this virus. It was concluded that 244/PR8 has the ability to protect in vivo against heterologous IFN-sensitive respiratory viruses, in addition to homologous influenza A viruses, and that it acts by fundamentally different mechanisms.

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Hsa-miR-30e-3p inhibits influenza B virus replication by targeting viral NA and NP genes

Experimental Biology and Medicine ◽

10.1177/1535370220953151 ◽

2020 ◽

Vol 245 (18) ◽

pp. 1664-1671

Author(s):

Kritsada Khongnomnan ◽

Suthat Saengchoowong ◽

Oraphan Mayuramart ◽

Pattaraporn Nimsamer ◽

Trairak Pisitkun ◽

...

Keyword(s):

Virus Infection ◽

Influenza A ◽

Influenza Virus Infection ◽

Influenza B Virus ◽

Influenza B ◽

Cellular Processes ◽

In Vitro Investigation ◽

B Virus ◽

Non Coding Rnas

Influenza B virus is a member of the Orthomyxoviridae family which can infect humans and causes influenza. Although it is not pandemic like influenza A virus, it nevertheless affects millions of people worldwide annually. MicroRNAs are small non-coding RNAs regulating gene expression at posttranscriptional level. They play various important roles in cellular processes including response to viral infection. MiRNA profiles from our previous study suggested that miR-30e-3p was one of the upregulated miRNAs that responded to influenza B virus infection. In this study, in silico prediction and in vitro investigation proved that this miRNA can directly target NA and NP genes of the influenza B virus and inhibit its replication. This finding might be useful for using miRNA as an alternative therapeutics for influenza virus infection.

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TMPRSS2 Independency for Haemagglutinin Cleavage In Vivo Differentiates Influenza B Virus from Influenza A Virus

Scientific Reports ◽

10.1038/srep29430 ◽

2016 ◽

Vol 6 (1) ◽

Cited By ~ 10

Author(s):

Kouji Sakai ◽

Yasushi Ami ◽

Noriko Nakajima ◽

Katsuhiro Nakajima ◽

Minori Kitazawa ◽

...

Keyword(s):

Influenza A Virus ◽

Influenza A ◽

Influenza B Virus ◽

Influenza B ◽

B Virus

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Comparative Study of the Temperature Sensitive, Cold Adapted and Attenuated Mutations Present in the Master Donor Viruses of the Two Commercial Human Live Attenuated Influenza Vaccines

Viruses ◽

10.3390/v11100928 ◽

2019 ◽

Vol 11 (10) ◽

pp. 928 ◽

Cited By ~ 6

Author(s):

Laura Rodriguez ◽

Pilar Blanco-Lobo ◽

Emma C. Reilly ◽

Tatsuya Maehigashi ◽

Aitor Nogales ◽

...

Keyword(s):

Influenza A ◽

Viral Disease ◽

The United States ◽

Influenza Viruses ◽

Influenza Vaccines ◽

Temperature Sensitive ◽

Cold Adapted ◽

Ann Arbor

Influenza viruses cause annual, seasonal infection across the globe. Vaccination represents the most effective strategy to prevent such infections and/or to reduce viral disease. Two major types of influenza vaccines are approved for human use: inactivated influenza vaccines (IIVs) and live attenuated influenza vaccines (LAIVs). Two Master Donor Virus (MDV) backbones have been used to create LAIVs against influenza A virus (IAV): the United States (US) A/Ann Arbor/6/60 (AA) and the Russian A/Leningrad/134/17/57 (Len) H2N2 viruses. The mutations responsible for the temperature sensitive (ts), cold-adapted (ca) and attenuated (att) phenotypes of the two MDVs have been previously identified and genetically mapped. However, a direct comparison of the contribution of these residues to viral attenuation, immunogenicity and protection efficacy has not been conducted. Here, we compared the In vitro and in vivo phenotype of recombinant influenza A/Puerto Rico/8/34 H1N1 (PR8) viruses containing the ts, ca and att mutations of the US (PR8/AA) and the Russian (PR8/Len) MDVs. Our results show that PR8/Len is more attenuated in vivo than PR8/AA, although both viruses induced similar levels of humoral and cellular responses, and protection against homologous and heterologous viral challenges. Our findings support the feasibility of using a different virus backbone as MDV for the development of improved LAIVs for the prevention of IAV infections.

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