Respiratory syncytial virus (RSV) is a major cause of severe respiratory tract disease worldwide, and a pediatric vaccine is not available. We generated a filamentous RSV-based virus-like particle (VLP) that presents the central conserved region of the attachment protein G. This was achieved by co-expressing the matrix protein, phosphoprotein, nucleoprotein, and a hybrid fusion protein in which the F ectodomain was replaced with the G central region (GCR). The latter is relatively conserved and contains a receptor binding site and hence is a logical vaccine target. The immunogenicity and efficacy of the resulting VLP, termed VLP-GCR, were examined in mice using intranasal application without adjuvant. VLP-GCR induced substantial anti-N antibody levels but very low anti-G antibody levels, even after three vaccinations. In contrast, a VLP presenting prefusion-stabilized fusion (preF) protein instead of GCR induced both high anti-F and anti-nucleoprotein antibody levels, suggesting that our GCR antigen was poorly immunogenic. Challenge of VLP-GCR-vaccinated mice caused increased weight loss and lung pathology, and both VLPs induced mucus in the lungs. Thus, neither VLP is suitable as a vaccine for RSV-naive individuals. However, VLP-preF enhanced the proportion of preF antibodies and could serve as a multi-antigen mucosal booster vaccine in the RSV-experienced population.

The human respiratory syncytial virus (RSV) is a major cause of severe respiratory tract disease, and a vaccine is not available. We previously reported a novel live vaccine expressing prefusion-stabilized fusion protein (preF) in place of the native F protein (RSV-preF^ΔCT). As preF is non-functional, RSV-preF^ΔCT was amplified in a production line expressing a functional substitute, and exhibited a single-cycle replication phenotype, which holds several unique potential advantages. RSV-preF^ΔCT prevented shedding and lung pathology after viral challenge in mice, but induced low levels of anti-attachment protein (G) antibodies (Abs). Given the significant contributions of anti-G Abs toward disease prevention, we generated modifications to RSV-preF^ΔCT in an effort to induce higher anti-G Ab levels. The Ab levels were monitored after the prime-boost vaccination of mice with modified vaccines. The most successful modification for enhancing induced anti-G Abs was seen with the placement of G in the first genome position. This vaccine also reduced the pathology after challenge with a high dose of wt RSV, and outperformed the sera from wt RSV-vaccinated mice in in vitro neutralization. Thus, raising the anti-G Ab levels induced by RSV-preF^ΔCT enhanced efficacy in vitro and in vivo, and constitutes an important next step in developing a live, single-cycle, efficacious vaccine for the human population.

Respiratory syncytial virus (RSV) is a major cause of severe lower respiratory tract disease in infants and children, and a pediatric vaccine is not available. Among various approaches, live vaccines are attractive as they induce broad multi-antigenic responses and have never been associated with vaccine-enhanced disease in RSV-naïve children. We previously reported a live prefusion F (preF) expressing single-cycle RSV vaccine, which was deficient in cell–cell transmission and yet induced protective responses in mice. To improve efficacy, we focused the anti-attachment protein (G) response on the relatively conserved G central region (GCR). Anti-GCR antibodies were shown to cross-protect against different RSV strains, and the GCR contains a receptor-binding domain. Focusing on the GCR was done by replacing the G open reading frame with that of a hybrid protein displaying the GCR, while also retaining stabilized preF expression. The resulting virus, RSV-preF-GCR, was readily produced in complementing cells, expressed preF and GCR at the infected cell surface, and maintained its single-cycle phenotype. In mice, intranasal prime-boost vaccination induced serum anti-preF and moderate levels of anti-G antibodies, which were specific for the GCR. RSV-preF-GCR also significantly reduced lung pathology after challenge with wild-type RSV. However, relative to a vaccine expressing full-length G, in vitro neutralization by RSV-preF-GCR-induced antibodies was poor, and lung IgA responses were unexpectedly low. Thus, RSV-preF-GCR is promising in uniquely combining single-cycle replication and induction of both anti-preF and anti-GCR immunity but likely requires efficacy improvements to enhance anti-G antibody levels and induce mucosal immunity.