Cell line development for biotherapeutics

This is the typical workflow used by groups developing stable cell lines for production of therapeutic recombinant proteins. In many cases, these proteins will be monoclonal antibodies (mAbs), but some companies also use to produce enzymes for enzyme replacement therapies in rare diseases.

Cell Line developments for biotherapeutics

The importance of clonality

A key assessment in the development of any new stable production cell line is the clonality. Historically, this was assessed based on probabilities, but more recently with the development of dedicated imaging, assurance can be provided by a high clarity picture of the single cell in the whole well immediately after seeding.

Importantly, from a CMC perspective for any new biological drug, the FDA and EMA both request assurance of clonality for the Master Cell Bank (MCB) as part of an IND process.

“It is expected that clonal cell lines are developed as referenced by ICH Q5D and EMA/CHMP”
Rachel Novak, US FDA, Jan 2017

Additionally, it is very important to consider the quality and consistency of the protein therapeutic. This is important for batch to batch production but in addition where companies are making biosimilars and they need to show equivalence. The regulator has provided guidance here again relating to clonality:

“Assurance of production cell bank clonality ensures consistency of product quality and process performance throughout the lifecycle of a product”
Rashmi Rawat, US FDA, April 2016

Continued Protein Supply

For the continued supply of protein therapeutic in the clinic to patients, to make the amounts of protein needed and of consistent quality, this can only be achieved by stable cell line production. If the regulator decided to put a hold on the product or did not accept the clonality for a given cell line, then this could lead to shortages of the drug in the market. This was discussed by Dr Audrey Jia in her video interviews.

Cell Line Development Workflow evolution

Several things have caused the workflow to evolve and change over the past 5 years. Firstly, there have been advancements in instrumentation. Second generation dedicated imagers for clonality, such as Cell Metric, have been largely able to remove the need for a second round of cloning which has nearly halved development times. More recently, novel single cell dispensing approaches, like the VIPS, have begun to replace traditional FACS as they can deliver higher cloning efficiencies, meaning potentially far fewer plates per project. Improvements in liquid handling have also meant more customers moving from 96 to 384 well plates.

Secondly, new cell engineering techniques have emerged which have led to a move to more targeted integration for the Gene of Interest (GOI) rather than previous random integration. These approaches have included Zinc Finger nucleases, TALENS, and more recently CRISPR/Cas9. This potentially means far fewer clones need to be screened.

The need now is for dedicated and integrated instruments for cell line development which do not require expensive specialists to run them, and which provide connectivity between respective instruments, ensuring complete audit trail and assurance documentation for the production cell line to the client and ultimately the regulator.