CHO cell production of a single, enveloped VLP vaccine co-displaying SARS-CoV-2, influenza and RSV antigens

Abstract

Concurrent circulation of SARS-CoV-2, influenza A (IAV), and respiratory syncytial virus (RSV) remains a major public health concern, driving the need for multivalent, combined vaccines. Here, we report the generation of trivalent enveloped virus-like particles (eVLPs) co-displaying SARS-CoV-2 spike (S), influenza A H1, and RSV fusion (F) proteins produced in Chinese hamster ovary (CHO) cells. Using both transient and stable gene expression systems, we achieved intrinsic co-assembly of all three antigens within enveloped particles. Stable CHO eVLP expression yielded antigen volumetric titers of 48 ± 2 mg/L. A prefusogenic F variant showed superior expression and incorporation efficiency compared to wild-type F and prefusion-stabilized forms. The adjuvanted trivalent eVLPs elicited IgG responses against S, H1, and F, together with S- and F-specific T-cell responses and antibodies targeting the palivizumab-binding site II of RSV-F, a clinically relevant neutralizing epitope. Notably, the formulation provided complete protection against homologous A/Puerto Rico/8/1934 (PR8; H1N1) challenge in mice , supporting further evaluation of this CHO-derived platform for combined respiratory-virus vaccination.