Development of a CHO cell-free protein synthesis platform for accelerated antibody screening

Chiara Josephine Heide, Kleio Kontoravdi, Karen Polizzi, Oscar Ces, London Imperial College
2021
Over 80% of therapeutic monoclonal antibody (mAb) products are expressed in Chinese Hamster Ovary (CHO) cells. While cell-based expression platforms are traditionally used for the exploration of novel mAb therapeutics in the discovery phase, cell-based technologies remain particularly time- and resource intensive for mAb screening. To circumvent these shortcomings, cell-free protein synthesis (CFPS) platforms have emerged as versatile alternative allowing for rapid and flexible production of
more » ... rapeutic proteins. In contrast to in vivo systems, CFPS platforms do not require intact host cells, which make them completely independent of host cell metabolism in determining product yield and quality. Although the open-production system bears several advantages over the traditional cell-based platforms, CFPS platforms still face limitations such as low product yields, challenges with post-translational modifications (PTMs), and poor cost efficiency of CFPS. It is therefore of great interest, to develop a simple, more cost-efficient and active CHO CFPS platform for accelerated screening of therapeutic mAbs. This work addresses the current limitations by developing a cell-free protein synthesis platform using CHO extracts for the rapid production and evaluation of industrially relevant mAbs. This study is divided into three main parts: 1) Development of a CHO CFPS system, 2) Yield optimization of CHO CFPS, 3) Production and screening of therapeutic mAbs. Our platform was able to express and characterise four functional mAbs in the supernatant fraction of 25 μl coupled batch reactions including two low in vivo expressers and the blockbuster drug Trastuzumab. Total synthesis yields were increased up to 50-fold by supplementing the system with two accessory proteins, GADD34 and K3L. Using our optimized platform, mAb yields of up to 31.06 μg/ml could be achieved. The trend in cell-free expressed functional mAbs replicated previously reported results from cell-based expression. Based on the success of our platform, we suggest [...]
doi:10.25560/90485 fatcat:5djvftqarrcfhcbhlyn5zfytqm