RNAse-free manufacture of Venezuelan Equine Encephalitis Virus (VEEV) plasmid DNA vaccine

Abstract

The long shelf-life and stability of DNA makes this platform highly attractive for low-cost, rapid delivery of pandemic response vaccines. Protocols utilized for clinical grade plasmid manufacture by contract development and manufacturing organizations are not readily accessible to academic and public research laboratories engaged in early-phase plasmid vaccine development. We present here the framework for DNA manufacturing using 3L-scale fermentation, anion-exchange chromatography and tangential flow filtration (TFF) leading to RNAase-free manufacture of plasmid DNA. The Venezuelan Equine Encephalitis Virus vaccine plasmid pWRG/VEEV, encoding the glycoprotein (E)3, E2, 6K and E1 genes was used as the prototype for this process development. The current effort yielded >95% pure and >80% supercoiled pWRG/VEEV plasmid preparations at 50-g wet cell weight scale. These data showed feasibility of manufacturing, highly pure pWRG/VEEV plasmid DNA using a cGMP compliant manufacturing process. Distribution StatementApproved for public release: distribution is unlimited