Internal ribosome entry site-based attenuation of a flavivirus candidate vaccine and evaluation of the effect of beta interferon coexpression on vaccine properties

Michael FRESE, Eva LEE, Maximilian Larena, Chloe LIM, Sudha RAO, Klaus Matthaei, Alex Khromykh, Ian Ramshaw, Mario Lobigs

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Abstract

Infectious clone technologies allow the rational design of live attenuated viral vaccines with the possibility of vaccine-driven coexpression of immunomodulatory molecules for additional vaccine safety and efficacy. The latter could lead to novel strategies for vaccine protection against infectious diseases where traditional approaches have failed. Here we show for the flavivirus Murray Valley encephalitis virus (MVEV) that incorporation of the internal ribosome entry site (IRES) of Encephalomyocarditis virus between the capsid and prM genes strongly attenuated virulence and that the resulting bicistronic virus was both genetically stable and potently immunogenic. Furthermore, the novel bicistronic genome organization facilitated the generation of a recombinant virus carrying an beta interferon (IFN-β) gene. Given the importance of IFNs in limiting virus dissemination and in efficient induction of memory B and T cell antiviral immunity, we hypothesized that coexpression of the cytokine with the live vaccine might further increase virulence attenuation without loss of immunogenicity. We found that bicistronic mouse IFN-β coexpressing MVEV yielded high virus and IFN titers in cultured cells that do not respond to the coexpressed IFN. However, in IFN response-sufficient cell cultures and mice, the virus produced a self-limiting infection. Nevertheless, the attenuated virus triggered robust innate and adaptive immune responses evidenced by the induced expression of Mx proteins (used as a sensitive biomarker for measuring the type I IFN response) and the generation of neutralizing antibodies, respectively. IMPORTANCE The family Flaviviridae includes a number of important human pathogens, such as Dengue virus, Yellow fever virus, Japanese encephalitis virus, West Nile virus, and Hepatitis C virus. Flaviviruses infect large numbers of individuals on all continents. For example, as many as 100 million people are infected annually with Dengue virus, and 150 million people suffer a chronic infection with Hepatitis C virus. However, protective vaccines against dengue and hepatitis C are still missing, and improved vaccines against other flaviviral diseases are needed. The present study investigated the effects of a redesigned flaviviral genome and the coexpression of an antiviral protein (interferon) on virus replication, pathogenicity, and immunogenicity. Our findings may aid in the rational design of a new class of well-tolerated and safe vaccines.

Original languageEnglish
Pages (from-to)2056-2070
Number of pages15
JournalJournal of Virology
Volume88
Issue number4
DOIs
Publication statusPublished - 2014

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interferon-beta
Flavivirus
Interferon-beta
ribosomes
Vaccines
vaccines
viruses
Murray Valley encephalitis virus
Viruses
Flaviviridae
Virulence
Dengue virus
Dengue Virus
Hepatitis C virus
live vaccines
interferons
Hepacivirus
Interferons
Antiviral Agents
Japanese Encephalitis Virus

Cite this

FRESE, Michael ; LEE, Eva ; Larena, Maximilian ; LIM, Chloe ; RAO, Sudha ; Matthaei, Klaus ; Khromykh, Alex ; Ramshaw, Ian ; Lobigs, Mario. / Internal ribosome entry site-based attenuation of a flavivirus candidate vaccine and evaluation of the effect of beta interferon coexpression on vaccine properties. In: Journal of Virology. 2014 ; Vol. 88, No. 4. pp. 2056-2070.
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abstract = "Infectious clone technologies allow the rational design of live attenuated viral vaccines with the possibility of vaccine-driven coexpression of immunomodulatory molecules for additional vaccine safety and efficacy. The latter could lead to novel strategies for vaccine protection against infectious diseases where traditional approaches have failed. Here we show for the flavivirus Murray Valley encephalitis virus (MVEV) that incorporation of the internal ribosome entry site (IRES) of Encephalomyocarditis virus between the capsid and prM genes strongly attenuated virulence and that the resulting bicistronic virus was both genetically stable and potently immunogenic. Furthermore, the novel bicistronic genome organization facilitated the generation of a recombinant virus carrying an beta interferon (IFN-β) gene. Given the importance of IFNs in limiting virus dissemination and in efficient induction of memory B and T cell antiviral immunity, we hypothesized that coexpression of the cytokine with the live vaccine might further increase virulence attenuation without loss of immunogenicity. We found that bicistronic mouse IFN-β coexpressing MVEV yielded high virus and IFN titers in cultured cells that do not respond to the coexpressed IFN. However, in IFN response-sufficient cell cultures and mice, the virus produced a self-limiting infection. Nevertheless, the attenuated virus triggered robust innate and adaptive immune responses evidenced by the induced expression of Mx proteins (used as a sensitive biomarker for measuring the type I IFN response) and the generation of neutralizing antibodies, respectively. IMPORTANCE The family Flaviviridae includes a number of important human pathogens, such as Dengue virus, Yellow fever virus, Japanese encephalitis virus, West Nile virus, and Hepatitis C virus. Flaviviruses infect large numbers of individuals on all continents. For example, as many as 100 million people are infected annually with Dengue virus, and 150 million people suffer a chronic infection with Hepatitis C virus. However, protective vaccines against dengue and hepatitis C are still missing, and improved vaccines against other flaviviral diseases are needed. The present study investigated the effects of a redesigned flaviviral genome and the coexpression of an antiviral protein (interferon) on virus replication, pathogenicity, and immunogenicity. Our findings may aid in the rational design of a new class of well-tolerated and safe vaccines.",
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Internal ribosome entry site-based attenuation of a flavivirus candidate vaccine and evaluation of the effect of beta interferon coexpression on vaccine properties. / FRESE, Michael; LEE, Eva; Larena, Maximilian; LIM, Chloe; RAO, Sudha; Matthaei, Klaus; Khromykh, Alex; Ramshaw, Ian; Lobigs, Mario.

In: Journal of Virology, Vol. 88, No. 4, 2014, p. 2056-2070.

Research output: Contribution to journalArticle

TY - JOUR

T1 - Internal ribosome entry site-based attenuation of a flavivirus candidate vaccine and evaluation of the effect of beta interferon coexpression on vaccine properties

AU - FRESE, Michael

AU - LEE, Eva

AU - Larena, Maximilian

AU - LIM, Chloe

AU - RAO, Sudha

AU - Matthaei, Klaus

AU - Khromykh, Alex

AU - Ramshaw, Ian

AU - Lobigs, Mario

PY - 2014

Y1 - 2014

N2 - Infectious clone technologies allow the rational design of live attenuated viral vaccines with the possibility of vaccine-driven coexpression of immunomodulatory molecules for additional vaccine safety and efficacy. The latter could lead to novel strategies for vaccine protection against infectious diseases where traditional approaches have failed. Here we show for the flavivirus Murray Valley encephalitis virus (MVEV) that incorporation of the internal ribosome entry site (IRES) of Encephalomyocarditis virus between the capsid and prM genes strongly attenuated virulence and that the resulting bicistronic virus was both genetically stable and potently immunogenic. Furthermore, the novel bicistronic genome organization facilitated the generation of a recombinant virus carrying an beta interferon (IFN-β) gene. Given the importance of IFNs in limiting virus dissemination and in efficient induction of memory B and T cell antiviral immunity, we hypothesized that coexpression of the cytokine with the live vaccine might further increase virulence attenuation without loss of immunogenicity. We found that bicistronic mouse IFN-β coexpressing MVEV yielded high virus and IFN titers in cultured cells that do not respond to the coexpressed IFN. However, in IFN response-sufficient cell cultures and mice, the virus produced a self-limiting infection. Nevertheless, the attenuated virus triggered robust innate and adaptive immune responses evidenced by the induced expression of Mx proteins (used as a sensitive biomarker for measuring the type I IFN response) and the generation of neutralizing antibodies, respectively. IMPORTANCE The family Flaviviridae includes a number of important human pathogens, such as Dengue virus, Yellow fever virus, Japanese encephalitis virus, West Nile virus, and Hepatitis C virus. Flaviviruses infect large numbers of individuals on all continents. For example, as many as 100 million people are infected annually with Dengue virus, and 150 million people suffer a chronic infection with Hepatitis C virus. However, protective vaccines against dengue and hepatitis C are still missing, and improved vaccines against other flaviviral diseases are needed. The present study investigated the effects of a redesigned flaviviral genome and the coexpression of an antiviral protein (interferon) on virus replication, pathogenicity, and immunogenicity. Our findings may aid in the rational design of a new class of well-tolerated and safe vaccines.

AB - Infectious clone technologies allow the rational design of live attenuated viral vaccines with the possibility of vaccine-driven coexpression of immunomodulatory molecules for additional vaccine safety and efficacy. The latter could lead to novel strategies for vaccine protection against infectious diseases where traditional approaches have failed. Here we show for the flavivirus Murray Valley encephalitis virus (MVEV) that incorporation of the internal ribosome entry site (IRES) of Encephalomyocarditis virus between the capsid and prM genes strongly attenuated virulence and that the resulting bicistronic virus was both genetically stable and potently immunogenic. Furthermore, the novel bicistronic genome organization facilitated the generation of a recombinant virus carrying an beta interferon (IFN-β) gene. Given the importance of IFNs in limiting virus dissemination and in efficient induction of memory B and T cell antiviral immunity, we hypothesized that coexpression of the cytokine with the live vaccine might further increase virulence attenuation without loss of immunogenicity. We found that bicistronic mouse IFN-β coexpressing MVEV yielded high virus and IFN titers in cultured cells that do not respond to the coexpressed IFN. However, in IFN response-sufficient cell cultures and mice, the virus produced a self-limiting infection. Nevertheless, the attenuated virus triggered robust innate and adaptive immune responses evidenced by the induced expression of Mx proteins (used as a sensitive biomarker for measuring the type I IFN response) and the generation of neutralizing antibodies, respectively. IMPORTANCE The family Flaviviridae includes a number of important human pathogens, such as Dengue virus, Yellow fever virus, Japanese encephalitis virus, West Nile virus, and Hepatitis C virus. Flaviviruses infect large numbers of individuals on all continents. For example, as many as 100 million people are infected annually with Dengue virus, and 150 million people suffer a chronic infection with Hepatitis C virus. However, protective vaccines against dengue and hepatitis C are still missing, and improved vaccines against other flaviviral diseases are needed. The present study investigated the effects of a redesigned flaviviral genome and the coexpression of an antiviral protein (interferon) on virus replication, pathogenicity, and immunogenicity. Our findings may aid in the rational design of a new class of well-tolerated and safe vaccines.

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KW - Antibodies, Neutralizing/immunology

KW - Cercopithecus aethiops

KW - Cloning, Molecular/methods

KW - DNA Primers/genetics

KW - Encephalitis Virus, Murray Valley/genetics

KW - Encephalomyocarditis virus/genetics

KW - Genetic Engineering/methods

KW - Immunity, Cellular/immunology

KW - Immunohistochemistry

KW - Interferon-beta/metabolism

KW - Kaplan-Meier Estimate

KW - Mice

KW - Mice, Inbred C57BL

KW - Myxovirus Resistance Proteins/immunology

KW - Reverse Transcriptase Polymerase Chain Reaction

KW - Ribosomes/genetics

KW - Vaccines, Synthetic/genetics

KW - Vero Cells

KW - Viral Vaccines/biosynthesis

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