Abstract
We have previously shown that live-attenuated rabies virus (RABV)-based vaccines infect and directly activate murine and human primary B cells in-vitro, which we propose can be exploited to help develop a single-dose RABV-based vaccine. Here we report on a novel approach to utilize the binding of Intracellular Adhesion Molecule-1 (ICAM-1) to its binding partner, Lymphocyte Function-associated Antigen-1 (LFA-1), on B cells to enhance B cell activation and RABV-specific antibody responses. We used a reverse genetics approach to clone, recover, and characterize a live-attenuated recombinant RABV-based vaccine expressing the murine Icam1 gene (rRABV-mICAM-1). We show that the murine ICAM-1 gene product is incorporated into virus particles, potentially exposing ICAM-1 to extracellular binding partners. While rRABV-mICAM-1 showed 10-100-fold decrease in viral titers on baby hamster kidney cells compared to the parental virus (rRABV), rRABV-mICAM-1 infected and activated primary murine B cells in-vitro more efficiently than rRABV, as indicated by significant upregulation of CD69, CD40, and MHCII on the surface of infected B cells. ICAM-1 expression on the virus surface was responsible for enhanced B cell infection since pre-treating rRABV-mICAM-1 with a neutralizing anti-ICAM-1 antibody reduced B cell infection to levels observed with rRABV alone. Furthermore, 100-fold less rRABV-mICAM-1 was needed to induce antibody titers in immunized mice equivalent to antibody titers observed in rRABV-immunized mice. Of note, only 103 focus forming units (ffu)/mouse of rRABV-mICAM-1 was needed to induce significant anti-RABV antibody titers as early as five days post-immunization. As both speed and potency of antibody responses are important in controlling human RABV infection in a post-exposure setting, these data show that expression of Icam1 from the RABV genome, which is then incorporated into the virus particle, is a promising strategy for the development of a single-dose RABV vaccine that requires only a minimum of virus.
Highlights
Rabies virus (RABV) causes a deadly zoonotic infection that targets and causes dysfunction within the central nervous system (CNS) of infected hosts
To determine whether B cell infection and activation could be enhanced by expressing murine Intracellular Adhesion Molecule-1 (ICAM-1) from a rRABV-based vaccine in-vitro, and whether enhanced B cell infection and activation leads to improved anti-RABV antibody responses in-vivo, we constructed and recovered a recombinant, attenuated vaccine strain of RABV expressing the murine Icam1 gene
Viral titers of supernatants harvested from cells infected with a low multiplicity of infection (MOI) (MOI = 0.01; multi-cycle growth curves) show that rRABV-mICAM-1 is slightly spread-deficient compared to rRABV, as indicated by 10–100-fold lower viral titers (Figure 1E, right panel)
Summary
Rabies virus (RABV) causes a deadly zoonotic infection that targets and causes dysfunction within the central nervous system (CNS) of infected hosts. Most of the disease burden is located in the developing nations of Asia and Africa, where it is estimated that 3.3 billion people live at risk of RABV infection [2]. Of those infected, 40% are under 15-years-of-age [2]. If administered in a timely and appropriate manner, current PEP is nearly 100% successful in preventing human RABV infection. This, together with routine vaccination of domestic animals, has resulted in a dramatic reduction of human RABV infections in developed countries over the last 50–60 years [3]
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