1. Promoter Optimization Circumvents Bcl-2 Transgene-Mediated Suppression of Lentiviral Vector Production
- Author
-
Cindy Y. Kok, Lauren M. MacLean, Renuka Rao, Shinya Tsurusaki, and Eddy Kizana
- Subjects
lentivirus ,promoter ,apoptosis ,gene therapy ,cardiotoxicity ,Microbiology ,QR1-502 - Abstract
Lentiviral vectors are a robust gene delivery tool for inducing transgene expression in a variety of cells. They are well suited to facilitate the testing of therapeutic candidate genes in vitro, due to relative ease of packaging and ability to transduce dividing and non-dividing cells. Our goal was to identify a gene that could be delivered to the heart to protect against cancer-therapy-induced cardiotoxicity. We sought to generate a lentivirus construct with a ubiquitous CMV promoter driving expression of B-cell lymphocyte/leukemia 2 gene (Bcl-2), a potent anti-apoptotic gene. Contrary to our aim, overexpression of Bcl-2 induced cell death in the producer HEK293T cells, resulting in failure to produce usable vector titre. This was circumvented by exchanging the CMV promoter to the cardiac-specific NCX1 promoter, leading to the successful production of a lentiviral vector which could induce cardioprotective expression of Bcl-2. In conclusion, reduced expression of Bcl-2 driven by a weaker promoter improved vector yield, and led to the production of functional cardioprotective Bcl-2 in primary cardiomyocytes.
- Published
- 2023
- Full Text
- View/download PDF