Transient Gene Expression: A Novel Mammalian Cell-Based Technology for Recombinant Protein Production

Abstract

Methods for the transfection of DNA into mammalian cells for transient gene expression were initially developed about 30 years ago. The main purpose was and still remains the molecular analysis of the cell's response to the exogenous protein. The use of transient gene expression for industrial purposes began in 1991 when Bennett and coworkers identified by alanine-scanning mutagenesis several mutants of human tissue-plasminogen activator (rtPA) that were eventually shown to be of higher medical potency than the wild-type protein. One of the molecules, now an approved thrombolytic ``TNKase{\texttrademark}'', had a dramatically extended half-life in patients, a high fibrin-specificity, and was insensitive to the inhibitor of plasminogen-activator PAI. The identification of improved mutants was based on more than 40,000 transient gene expression experiments (Wurm et al. unpublished) executed during the course of one year using calcium-phosphate transfections in human embryonic kidney (HEK) 293 cells (Gorman et al 1990).