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Aside from process improvements focussed on media and feed strategies the CHO host remains largely unchanged from that which was used thirty years ago. Here I will present Horizon Discovery’s approach to improving our GS knockout CHO K1 cell line using genome engineering approaches such as CRISPR screening and rAAV to provide FTO in biomanufacturing.
We describe our CHO expression platform for the rapid production of various recombinant proteins. We use the cumate-inducible CHO expression system that allows the generation of stable and scalable pools expressing high levels of monoclonal antibodies or r-proteins in 2-3 weeks post-transfection. The pools are also used to derive stable and high-expressing CHO clones for manufacturing therapeutic candidates.
David Shaw*, Ali AlBarakat, Daniel Tran, Joni Tsukuda, Mandy Yim
Early Stage Cell Culture, Pharma Technical Development, Genentech, Inc.
Keywords: cell line development automation, clonally-derived, single-cell printer
In order to increase efficiency, consistency and flexibility, we embarked on completely automating the clinical and commercial cell line development (CLD) process using custom automated solutions. Automated single-cell cloning (SCC) and hit-picking workflows have been in use for several years and automate the most tedious steps in CLD. The SCC workflow uses plate imaging to document that our cell lines are clonally-derived. We added single-cell deposition to our plate imaging workflow to gain significant efficiencies over limiting dilution SCC. We use existing hit-picking automation to pick and scale-up single-cell clones from 384-well plates to 96-well plates, 24-well plates, 6-well plates and 24-well deep well blocks. We developed custom seed train automation to passage seed train cultures and set-up and sample fed-batch cultures in an unattended operation. Using the same seed train automation and custom consumables we are able to scale up to 300mL cultures in an automated operation. The implementation of the entire automated CLD workflow and our projected capacity of 50 – 200 cell lines per year with a very limited staff will be discussed.
The concept of high stringency selection of cells producing high amounts of a desired protein of interest is well known in the art and different solutions exist. However, many existing solutions exhibit different drawbacks e.g. need to engineer a host cell line or use of an antibiotic. The presentation will introduce a high stringency selection system using an endogenous metabolic selection marker without the need for cell line engineering.
Currently there is a logjam of candidate HIV vaccines awaiting clinical testing . HIV subunit vaccines have been more difficult to manufacture than other biopharmaceuticals due to extensive heterogeneity in N-linked glycosylation. Here we describe the use of gene editing and robotic selection to create an MGAT1- CHO cell line that improves upstream and downstream manufacturing processes.