Targeting Subtype-Independent Immune Responses Against Influenza A Virus
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Philipps-Universität Marburg
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Abstract
Influenza A virus (IAV) is a major burden for public health. Besides seasonal, human
IAV, which circulates during the cold seasons and is responsible for hundreds of
thousands of deaths each year, its potential to reassort with avian IAV fuels the fear
of future pandemics. Currently licensed vaccines are standardized to elicit antibodies
directed against the head domain of the hemagglutinin (HA) on the viral surface and
binding of these can effectively neutralize virus particles. However, the antigenic
plasticity of HA in consecutive IAV seasons and the above-mentioned potential for
upcoming reassorted viruses make it necessary to update IAV vaccines on an annual
basis and still, these vaccines cannot protect against outbreaks of zoonotic origin. It
is therefore a major objective to develop new subtype-independent approaches
against IAV via immunization as a first line of defense or via antiviral drugs.
This thesis is divided into two different projects. The first one investigates the
protection against heterologous challenge after vaccination with single or combined
IAV proteins and the underlying humoral immune response. The second project aims
at characterizing the effect of ADAR1 (adenosine deaminase acting on RNA 1), a
potential target for antiviral compounds, on IAV.
In the first project, the internal proteins NP and M1 derived from the live-attenuated
influenza vaccine were investigated regarding their potential to elicit protective
immunity against heterologous virus challenge and compared to currently used
approaches based on the stem region of HA (HAstem) or the membrane-integral M2
protein. Furthermore, underlying immune responses were characterized. We
demonstrate that VSV-vectored immunization with the internal protein NP and M2,
but also H3stem can remarkably reduce IAV-induced disease in a heterologous manner
and that this effect is independent of detectable T cell responses. Analysis of humoral
immunity revealed high IgG antibodies, distinct IgG subclass profiles against
different viruses, and most importantly, activation of the murine FcγRIV, known to
mediate antibody-dependent cell-mediated cytotoxicity and –phagocytosis via
alveolar macrophages. Furthermore, we showed that the absence of humoral
immunity after viral vector-based immunization with M1 correlated with the lack of
protection against IAV challenge in mice, which suggests absence of protective, but
undetected, T cell responses, and strengthens our assumption of antibody-mediated
protection. Humoral immunity against internal proteins of IAV have first been
described decades ago but are often considered inferior to those directed against
surface antigens. The main reason for this is the localization of internal proteins, as
it raises questions about the capability to provide protection through humoral
immune responses. Therefore, they were often neglected or ignored in the past. While
these antibodies do not mediate neutralization, they can in fact activate Fc-mediated
effector functions and protect from homologous as well as heterologous disease. Our
results add further insights in these mechanisms and correlate with previously
described capability of NP and M2 to induce protective antibodies. Our results
therefore imply that these immune responses should not be ignored in the rational
design of future IAV vaccines.
In the second project, we used two different cell culture systems to investigate the
effect of different ADAR1 isoforms on the replication of IAV, in order to consider its
potential as a target for antiviral therapies. We demonstrated that ADAR1p150 is a
proviral factor for IAV infection and is required for efficient viral protein expression
in HeLa cells, which is in line with previous publications describing this in other
eukaryotic cell lines. This finding consolidates the concept of proviral ADAR1p150
for IAV, as it was described for other RNA viruses. Furthermore, we generated
MDCK cells deficient for ADAR1p150 or complete ADAR1 using a
CRISPR/Cas9n system and confirmed the proviral effect of ADAR1p150 on viral
replication. We showed that absence of ADAR1 resulted in strikingly decreased IAVinduced
cell death, which may be an essential factor for targeting it with antiviral
compounds. Individuals passing away because of an IAV infection show severe
damage of lung tissue, resulting from exaggerated innate immune responses, a proinflammatory
milieu, and the subsequent collapse of the epithelial barrier in the lung.
Our results in HeLa cells indicate that a potential knock-down or inhibition of
ADAR1 does not necessarily lead to activated innate immunity pathways, as it could
be expected from other RNA viruses, and in combination with the above-mentioned
decreased cell death in infected MDCK ADAR1KO cells builds a promising basis for
the further investigation of ADAR1 as a target for anti-IAV treatment therapies.
Review
Metadata
Contributors
Supervisor:
Dates
Created: 2022Issued: 2023-03-09Updated: 2023-03-09
Faculty
Medizin
Publisher
Philipps-Universität Marburg
Language
ger
Data types
DoctoralThesis
Keywords
heterologous immunityvaccinenon-neutralizing antibodiesinnate immunity
DFG-subjects
Angeborene ImmunitätNicht-neutralisierende AntikörperADAR1Heterologe ImmunitätVakzinInfluenza A VirusImpfstoff
DDC-Numbers
610
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Wittwer, Kevin: Targeting Subtype-Independent Immune Responses Against Influenza A Virus. : Philipps-Universität Marburg 2023-03-09. DOI: https://doi.org/10.17192/z2023.0038.
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This item has been published with the following license: In Copyright