Process development for pandemic influenza VLP vaccine production using a baculovirus expression system

Abstract

BackgroundInfluenza viruses cause hundreds of thousands of respiratory diseases worldwide each year, and vaccination is considered the most effective approach for preventing influenza annual epidemics or pandemics. Since 1950, chicken embryonated eggs have been used as the main method for producing seasonal influenza vaccines. However, this platform has the main drawback of a lack of scale-up flexibility, and thus, egg-based vaccine manufacturers cannot supply sufficient doses within a short period for use for pandemic prevention. As a result, strategies for reducing the manufacturing time and increasing production capacity are urgently needed. Non-virion vaccine methods have been considered an alternative strategy against an influenza pandemic, and the purpose of maintaining an immunogenic capsule structure with infectious properties appears to be met by the virus-like particle (VLP) platform.ResultsAn influenza H7N9-TW VLP production platform using insect cells, which included the expression of hemagglutinin (HA), NA, and M1 proteins, was established. To scale up H7N9-TW VLP production, several culture conditions were optimized to obtain a higher production yield. A high level of dissolved oxygen (DO) could be critical to H7N9-TW VLP production. If the DO was maintained at a high level, the HA titer obtained in the spinner flask system with ventilation was similar to that obtained in a shake flask. In this study, the HA titer in a 5-L bioreactor with a well-controlled DO level was substantially improved by 128-fold (from 4 HA units (HAU)/50 μL to 512 HAU/50 μL).ConclusionsIn this study, a multigene expression platform and an effective upstream process were developed. Notably, a high H7N9-TW VLP yield was achieved using a two-step production strategy while a high DO level was maintained. The upstream process, which resulted in high VLP titers, could be further used for large-scale influenza VLP vaccine production.

Integrating high cell density cultures with adapted laboratory evolution for improved Gag‐HA virus‐like particles production in stable insect cell lines

“…Different expression systems and/or delivery vectors have been employed for production of influenza VLPs (Haynes et al, 2009;Thompson et al, 2015;Tretyakova et al, 2016;Venereo-Sanchez et al, 2016). In insect cells, influenza VLPs have been mostly produced using the IC-BEVS, with HA titers varying from 16 to 300 HA titer/ml Lai et al, 2019;Park & Song, 2017). In mammalian cells, these particles are mainly produced using transient transfection, with values reportedly lower than those achieved with IC-BEVS (Venereo-Sanchez et al, 2016).…”

Improving Influenza HA-Vlps Production in Insect High Five Cells via Adaptive Laboratory Evolution

“…Different cell lines and/or expression systems have been employed for the production of influenza HA-VLPs, yielding 128 HA titer.mL −1 [ 26 ], 16 HA titer.mL −1 [ 9 ], or even above 300 HA titer.mL −1 [ 46 , 47 , 48 ]. Noteworthy, the production process herein developed was shown to yield an extracellular HA titer similar to those reported when using non-adapted cells (192 HA titer.mL −1 ) or higher when using adapted cells (768 HA titer.mL −1 ).…”

Virus-like particles: preparation, immunogenicity and their roles as nanovaccines and drug nanocarriers

“…After cell harvesting, the nature of the first stage in the purification process depends on the ability of the VLPs to be released into the extracellular medium. While in some reported cases such as influenza VLPs produced in insect cell culture the particles are released into the medium without the need for special measures, if the VLP is not released effectively, cell lysis or an alternative extraction method may be required to disrupt the cells [110]. The generally adopted approach is to design a cloned gene to express a protein containing an effective signal peptide that will be recognized by the secretory pathway to facilitate release [55,60].…”