Targeted Optimization Of An Upstream AAV8 Process For Enhanced Downstream Performance

By Petra Dekleva, Ivana Petrović Koshmak, Janja Merkelj Koren, Rok Žigon, Andrej Mihevc, Mirjam Krašna, Kaja Bažec, Valentina Novak, Marko Šnajder, Sandra Potušek, Maja Leskovec, and Aleš Štrancar

While several recombinant adeno-associated viral vector (rAAV)-mediated therapies have received approval, and many more are undergoing clinical trials, establishing an efficient rAAV process platform remains a significant hurdle. One primary challenge stems from the frequent practice of developing upstream (USP) and downstream (DSP) processes separately. USP optimization activities often prioritize achieving a high viral titer, which may not always result in maximal purity and recovery during DSP.

A second obstacle is the lack of suitable analytical methodology capable of delivering the reliable and unbiased results essential for integrating USP and DSP into a single bioprocess. Additionally, rAAV-based gene therapy vectors necessitate the elimination of both process- and product-specific impurities, as these can pose serious safety risks.

Here, we show an example of AAV8 USP optimization directed towards enhancing DSP performance where AAV8 is generated in HEK293 suspension cells through plasmid transfection.