Virus-DNA-Virus hybrid vaccines for the prevention of Influenza - PROJECT SUMMARY AND ABSTRACT
The ability to use a virus as a carrier particle to deliver DNA vaccines directly to antigen presenting
cells has not been previously explored. Our proposal seeks to develop a virus-DNA-virus complex
vaccine that will be preferentially taken up by dendritic cells (DCs) and other important Antigen
Presenting Cells (APC) in vivo, that will deliver intracellular nucleic acid, activate DCs by engaging
toll-receptor- like (TLR) signaling pathways, and deliver a vaccine stimulus that should promote
robust and long- term immune activation against Influenza A pathogens. Targeted antigen
presenting cell (APC) uptake and activation needs to occur to foster an appropriate immune activation
sequence to pathogen subunit vaccine antigens that are by themselves weakly immunogenic. We
have exploited the stability, and APC uptake properties of Tobacco Mosaic Virus (TMV) to deliver
DNA by macropinocytosis. Our preliminary data demonstrate that TMV virus is actively taken up DCs,
and induce DC and other APC activation pathways. TMV has been engineered to deliver DNA
encoding Ovalbumin, expressed from a CMV promoter (TMV-OvaD). Preparation of TMV-OvaD
particles, is a simple two step process, and engineered to deliver intracellular DNA after uptake.
Based on immunogenicity in mice, TMV-OvaD can induce significantly improved IgG and cytotoxic
T lymphocyte (CTL) immune responses to Ova protein or SIINFEKL peptide, compared to CMV-
Ova intramuscular injection. However, there are several crucial steps to optimize TMV-DNA antigen
delivery, and boost immune response activation. CMV promoter-driven expression is limited to nuclear
RNA transcription, and may limit antigen expression. First, we want to explore the use of a CMV
promoter to drive expression of a small self-replicating RNA, which will promote and amplify
cytoplasmic RNA transcription, boost antigen delivery, and provide a second TLR signal. Second, we
will move from model antigen expression (Ova and GFP), to expressing influenza virus
Hemagglutinin (HA) antigens relevant to pandemic and pre-pandemic influenza (H5N1 and H7N9
HAs). Third, we will test TMV-DNA formulations in Influenza pathogen challenge studies to determine
protective efficacy, and initiate safety testing in mice. We believe that TMV-DNA vaccines will target
APC uptake, provide several sources of TLR signaling and activation, and stimulate appropriate
antigen presentation context that stimulates broadly neutralizing antibodies. Our goal is to actualize the
promise of safe and effective rapid response influenza vaccines against potentially pandemic
disease. Development of similar vaccines against other pathogens, where complex mixtures of
antigens may be required for stimulating adequate immunity, may broaden the utility of this approach.