A potent, broadly protective vaccine against SARS-CoV-2 variants of concern.

Since the first outbreak in December 2019, SARS-CoV-2 has been constantly evolving and five variants have been classified as Variant of Concern (VOC) by the World Health Organization (WHO). These VOCs were found to enhance transmission and/or decrease neutralization capabilities of monoclonal antibodies and vaccine-induced antibodies. Here, we successfully designed and produced a recombinant COVID-19 vaccine in CHO cells at a high yield. The vaccine antigen contains four hot spot substitutions, K417N, E484K, N501Y and D614G, based on a prefusion-stabilized spike trimer of SARS-CoV-2 (S-6P) and formulated with an Alum/CpG 7909 dual adjuvant system. Results of immunogenicity studies showed that the variant vaccine elicited robust cross-neutralizing antibody responses against SARS-CoV-2 prototype (Wuhan) strain and all 5 VOCs. It further, stimulated a TH1 (T Helper type 1) cytokine profile and substantial CD4+ T cell responses in BALB/c mice and rhesus macaques were recorded. Protective efficacy of the vaccine candidate was evaluated in hamster and rhesus macaque models of SARS-CoV-2. In Golden Syrian hamsters challenged with Beta or Delta strains, the vaccine candidate reduced the viral loads in nasal turbinates and lung tissues, accompanied by significant weight gain and relieved inflammation in the lungs. In rhesus macaque challenged with prototype SARS-CoV-2, the vaccine candidate decreased viral shedding in throat, anal, blood swabs over time, reduced viral loads of bronchus and lung tissue, and effectively relieved the lung pathological inflammatory response. Together, our data demonstrated the broadly neutralizing activity and efficacy of the variant vaccine against both prototype and current VOCs of SARS-CoV-2, justifying further clinical development.

Preclinical evaluation of recombinant HFMD vaccine based on enterovirus 71 (EV71) virus-like particles (VLP): Immunogenicity, efficacy and toxicology.

Enterovirus 71 (EV71) is one of the major causative agents for hand, foot and mouth disease (HFMD) in children. Currently, three inactivated EV71 vaccines have been approved by Chinese government. We previously demonstrated that recombinant EV71 virus-like particles (VLP) produced in Pichia pastoris can be produced at a high yield with a simple manufacturing process, and the candidate vaccine elicited protective humoral immune responses in mice. In present study, the nonclinical immunogenicity, efficacy and toxicity of the EV71 vaccine was comprehensively evaluated in rodents and non-human primates. The immunogenicity assessment showed that EV71 VLPs vaccine elicited high and persistent neutralizing antibody responses, which could be comparable with a licensed inactivated vaccine in animals. The immune sera of vaccinated mice also exhibited cross-neutralization activities to the heterologous subtypes of EV71. Both passive and maternal antigen specific antibodies protected the neonatal mice against the lethal EV71 challenge. Furthermore, nonclinical safety assessment of EV71 VLP vaccine showed no signs of systemic toxicity in animals. Therefore, the excellent immunogenicity, efficacy and toxicology data supported further evaluation of the VLP-based EV71 vaccine in humans.

Development and characterization of an enterovirus 71 (EV71) virus-like particles (VLPs) vaccine produced in Pichia pastoris.

Enterovirus 71 (EV71) is one of the major causative agents for hand, foot and mouth disease (HFMD) in children. Although there are three inactivated virus-based HFMD vaccines licensed in China, alternative approaches have been taken to produce an effective and safer vaccine that is easier to manufacture in large scale. Among these, a virus-like particles (VLPs) based EV71 vaccine is under active development. For this purpose, an efficient methodology for the production of EV71-VLPs by recombinant technology is needed. We here report the construction and expression of the P1 and 3C genes of EV71 in Pichia pastoris for producing VLP-based EV71 vaccine antigen with a high yield and simple manufacturing process. Based on codon-optimized P1 and 3C genes, EV71-VLPs were efficiently expressed in Pichia pastoris system, and the expression level reached 270 mg/L. Biochemical and biophysical analyses showed that the produced EV71-VLPs consisted of processed VP0, VP1, and VP3 present as ~35nm spherical particles. The immune response as a function of EV71-VLPs and adjuvant dose ratio was investigated for vaccine development. Immunization with EV71-VLPs of 1-5 µg/dose and adjuvant of 225 µg/dose induced robust neutralizing antibody responses in mice and provided effective protection against lethal challenge in both maternally transferred antibody and passive transfer protection mouse models. Therefore, the yeast produced EV71-VLPs antigen is a promising candidate for the development of a vaccine against HFMD.