Susceptibility of naturally human papillomavirus type 16 major capsid protein L1 variants to vaccines and predictions of future evolutionary trends.

Human papillomavirus type 16 (HPV16) causes over 50% of HPV-related cervical cancers. With the widespread use of virus-like particle (VLP)-based HPV prophylactic vaccines, investigating the differences in viral assembly, infectivity and antigenic properties among different variants is highly important for further eliminating diseases caused by HPV. We analyzed HPV16 L1 protein sequences from the NCBI database and identified 14 representative clusters. Pseudovirus (PsV) formation, infectivity, and susceptibility to vaccines were assessed for these variants, and molecular dynamics (MD) simulations and modified enzyme-linked immunosorbent assays (ELISAs) were ultimately employed to investigate the underlying mechanisms. The sequence identical to P03101 in the UniProt database served as the reference (P16_WT). Among the 13 variants, five presented reduced PsV packaging efficiency to varying degrees, which correlated with diminished L1 protein expression. Compared with P16_WT, the remaining eight variants successfully produced infectious PsVs with high titers and exhibited equivalent infectivity in epithelial cells. Neutralization assays revealed that four variants demonstrated varying levels of decreased susceptibility to vaccines; notably, P16_5 and P16_13 exhibited significantly reduced sensitivity to all three vaccines. MD simulation and modified ELISA indicated that this reduction results from the unstable binding interactions between the L1 variants and the antibodies. Importantly, key residues in these less susceptible variants, such as P16_5 and P16_13, were under positive selection prior to the commercialization of these vaccines. Our findings demonstrated that a significant proportion of globally circulating HPV16 L1 variants currently display diminished susceptibility to available vaccines. Consequently, ongoing surveillance of HPV variants is imperative.