Abstract
The ongoing COVID-19 pandemic drew global attention to infectious diseases, attracting numerous resources for development of pandemic preparedness plans and vaccine platforms—technologies with robust manufacturing processes that can quickly be pivoted to target emerging diseases. Newcastle Disease Virus (NDV) has been studied as a viral vector for human and veterinary vaccines, but its production relies heavily on embryonated chicken eggs, with very few studies producing NDV in cell culture. Here, NDV is produced in suspension Vero cells, and analytical assays (TCID50 and ddPCR) are developed to quantify infectious and total viral titer. NDV-GFP and NDV-FLS (SARS-CoV-2 full-length spike protein) constructs were adapted to replicate in Vero and HEK293 suspension cultures using serum-free media, while fine-tuning parameters such as MOI, temperature, and trypsin concentration. Shake flask productions with Vero cells resulted in infectious titers of 1.07 × 108 TCID50/mL for NDV-GFP and 1.33 × 108 TCID50/mL for NDV-FLS. Production in 1 L batch bioreactors also resulted in high titers in culture supernatants, reaching 2.37 × 108 TCID50/mL for NDV-GFP and 3.16 × 107 TCID50/mL for NDV-FLS. This shows effective NDV production in cell culture, building the basis for a scalable vectored-vaccine manufacturing process that can be applied to different targets.
Highlights
Infectious diseases are present throughout history, emerging and reemerging as decades pass [1]
The Vero cell line adapted to suspension was provided by the National Research Council of Canada (NRC), and the adaptation was described in a previous work [24]
Adherent HEK293 (HEK293A, ATCC CRL-1573 [26]) cells were routinely passaged in the same way as adherent Vero cells, but using Dulbecco’s Modified Eagle’s Medium (DMEM) (Thermo Fisher Scientific, Waltham, MA, USA) with 10% Fetal Bovine Serum (FBS) (Gibco, Gaithersburg, MD, USA) and 1% Penicillin-Streptomycin solution instead of VP Serum-Free Medium (VP-SFM)
Summary
Infectious diseases are present throughout history, emerging and reemerging as decades pass [1]. There are well established methods to generate recombinant NDV constructs bearing protective key antigens from other viral pathogens These aspects make NDV a promising vaccine vector that has already been explored in vaccine candidates against H1N1 influenza, SARS-CoV, HIV, among others [2]. Despite having garnered interest as a viral vector for vaccination and for cancer therapy [15], there are still few studies on the development of production processes for NDV This virus is produced in embryonated chicken eggs and collected in the allantoic fluid. This is a cost-effective method that can take advantage of existing manufacturing structures for influenza [18], it presents disadvantages when compared to cell culture technologies and their potential large-scale production under controlled operational conditions in bioreactors. This is an innovative work in exploring and Vaccines 2021, 9, 1335 establishing these essential aspects for robust production and quality assessments of NDV in cell culture
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
