Mutations in the F protein of the live-attenuated respiratory syncytial virus vaccine candidate ΔNS2/Δ1313/I1314L increase the stability of infectivity and content of prefusion F protein

• Published in: PloS one 2024-04, Vol.19 (4), p.e0301773-e0301773
• Main Authors: Alamares-Sapuay, Judith, Kishko, Michael, Lai, Charles, Parrington, Mark, Delagrave, Simon, Herbert, Richard, Castens, Ashley, Swerczek, Joanna, Luongo, Cindy, Yang, Lijuan, Collins, Peter L, Buchholz, Ursula J, Zhang, Linong
• Format: Article
• Language: English
• Subjects: Animals; Antibodies, Neutralizing; Antibodies, Viral; Biology and Life Sciences; Child; Chlorocebus aethiops; Humans; Medicine and Health Sciences; Mutation, Missense; Physical Sciences; Research and Analysis Methods; Respiratory Syncytial Virus Infections; Respiratory Syncytial Virus Vaccines - genetics; Respiratory Syncytial Virus, Human - genetics; Vero Cells; Viral Fusion Proteins - genetics
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0301773

Respiratory syncytial virus (RSV) is the leading viral cause of bronchiolitis and pneumonia in infants and toddlers, but there currently is no licensed pediatric vaccine. A leading vaccine candidate that has been evaluated for intranasal immunization in a recently completed phase 1/2 clinical trial is an attenuated version of RSV strain A2 called RSV/ΔNS2/Δ1313/I1314L (hereafter called ΔNS2). ΔNS2 is attenuated by deletion of the interferon antagonist NS2 gene and introduction into the L polymerase protein gene of a codon deletion (Δ1313) that confers temperature-sensitivity and is stabilized by a missense mutation (I1314L). Previously, introduction of four amino acid changes derived from a second RSV strain "line 19" (I79M, K191R, T357K, N371Y) into the F protein of strain A2 increased the stability of infectivity and the proportion of F protein in the highly immunogenic pre-fusion (pre-F) conformation. In the present study, these four "line 19" assignments were introduced into the ΔNS2 candidate, creating ΔNS2-L19F-4M. During in vitro growth in Vero cells, ΔNS2-L19F-4M had growth kinetics and peak titer similar to the ΔNS2 parent. ΔNS2-L19F-4M exhibited an enhanced proportion of pre-F protein, with a ratio of pre-F/total F that was 4.5- to 5.0-fold higher than that of the ΔNS2 parent. The stability of infectivity during incubation at 4°C, 25°C, 32°C and 37°C was greater for ΔNS2-L19F-4M; for example, after 28 days at 32°C, its titer was 100-fold greater than ΔNS2. ΔNS2-L19F-4M exhibited similar levels of replication in human airway epithelial (HAE) cells as ΔNS2. The four "line 19" F mutations were genetically stable during 10 rounds of serial passage in Vero cells. In African green monkeys, ΔNS2-L19F-4M and ΔNS2 had similar growth kinetics, peak titer, and immunogenicity. These results suggest that ΔNS2-L19F-4M is an improved live attenuated vaccine candidate whose enhanced stability may simplify its manufacture, storage and distribution, which merits further evaluation in a clinical trial in humans.