Development of a production method for scaling up lentiviral vector manufacture

DESCRIPTION (provided by applicant): Gene therapy vectors derived from lentiviruses offer a number of advantages over other gene transfer vectors, and they represent a promising approach for treating a variety of diseases, such as age-related macular degeneration, Parkinson's disease, and blood cell malignancies. Unfortunately, lentiviral vectors are difficult to produce in large numbers due to the lack of stable packaging cell lines and to inefficiencies associated with standard methods of transient transfection. This Phase 1 proposal describes the development of a scalable process for lentiviral vector production that uses MaxCyte's proprietary flow electroporation technology to transfect plasmid DNAs encoding components of bovine immunodeficiency virus (BIV) gene therapy vectors into mammalian cells. The workplan includes optimization of electroporation parameters for loading BIV component plasmids into adherent HEK 293T cells at both small and large scales. These experiments are intended to demonstrate that flow electroporation is superior to other methods (e.g., calcium phosphate precipitation) for transfecting cells that are commonly used in lentivector production. In addition, protocols will be developed for transfecting BIV component plasmids into suspension cells, which offer a number of advantages over adherent cells for manufacturing biological products. The goal is to generate an efficient, suspension cell-based method for lentivirus production that can be scaled up accommodate the needs of clinical-scale testing as well as commercial manufacturing of a lentivirus-based therapeutic product. The results of these studies will provide the foundation for Phase 2 studies to optimize manufacturing protocols that can by used in a GMP facility for producing lentiviral gene therapy vectors on a commercial scale.