High-titer manufacturing of SARS-CoV-2 Spike-pseudotyped VSV in stirred-tank bioreactors

• Published in: Molecular therapy. Methods & clinical development Vol. 32; no. 1; p. 101189
• Main Authors: Todesco, Hayley M., Gafuik, Chris, John, Cini M., Roberts, Erin L., Borys, Breanna S., Pawluk, Alexis, Kallos, Michael S., Potts, Kyle G., Mahoney, Douglas J.
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 14.03.2024; American Society of Gene & Cell Therapy; Elsevier
• Subjects: COVID-19; Cytodex 1; microcarrier; omicron; Original; SARS-CoV-2; spinner flask; vaccine; Vero; VSV; COVID-19; SARS-CoV-2; vaccine; omicron; Cytodex 1; spinner flask; Vero; VSV; microcarrier
• ISSN: 2329-0501; 2329-0501
• DOI: 10.1016/j.omtm.2024.101189

The severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2) pandemic highlighted the importance of vaccine innovation in public health. Hundreds of vaccines built on numerous technology platforms have been rapidly developed against SARS-CoV-2 since 2020. Like all vaccine platforms, an important bottleneck to viral-vectored vaccine development is manufacturing. Here, we describe a scalable manufacturing protocol for replication-competent SARS-CoV-2 Spike-pseudotyped vesicular stomatitis virus (S-VSV)–vectored vaccines using Vero cells grown on microcarriers in a stirred-tank bioreactor. Using Cytodex 1 microcarriers over 6 days of fed-batch culture, Vero cells grew to a density of 3.95 ± 0.42 ×106 cells/mL in 1-L stirred-tank bioreactors. Ancestral strain S-VSV reached a peak titer of 2.05 ± 0.58 ×108 plaque-forming units (PFUs)/mL at 3 days postinfection. When compared to growth in plate-based cultures, this was a 29-fold increase in virus production, meaning a 1-L bioreactor produces the same amount of virus as 1,284 plates of 15 cm. In addition, the omicron BA.1 S-VSV reached a peak titer of 5.58 ± 0.35 × 106 PFU/mL. Quality control testing showed plate- and bioreactor-produced S-VSV had similar particle-to-PFU ratios and elicited comparable levels of neutralizing antibodies in immunized hamsters. This method should enhance preclinical and clinical development of pseudotyped VSV-vectored vaccines in future pandemics. [Display omitted] Todesco and colleagues describe a scalable manufacturing protocol for high-titer replication-competent SARS-CoV-2 Spike-pseudotyped vesicular stomatitis virus (S-VSV)–vectored vaccines using Vero cells grown on microcarriers in a stirred-tank bioreactor. This method should enhance preclinical and clinical development of pseudotyped VSV-vectored vaccines in future pandemics.