Clonal Derivation in Biopharmaceutical Cell Line Development – a Regulatory Perspective

19th November 2019: Early last month saw publication of the latest perspectives from the FDA in a paper entitled:

Considering “clonality”: A regulatory perspective on the importance of the clonal derivation of mammalian cell banks in biopharmaceutical development authored by Joel Welch and Sarah Arden (USFDA).

Key Points

As a major supplier of technologies for cell line development and a thought leader in this area, we decided it was valuable to pick out some key points from this latest paper to provide further clarity and confidence for our customers.

The first point to note is a shift in terminology from previous ‘proof of clonality’ or more recently ‘assurance of clonality’ to the term ‘clonally-derived population’ to describe the original cell bank.  This is now preferable and scientifically more accurate.

The paper talks to the importance of imaging to provide supportive data for clonal derivation, but also based I think on some of our own arguments, outlines some of the potential pitfalls with imaging namely “their utility (and “accuracy”) may be affected by factors, such as calibration, focus, illumination, focal depth, and/or resolution issues. In addition, these imaging systems present other, non-optical based, considerations as well, such as the probability that the cell is on the bottom of the well, and the probability of locating the cell within the well during imaging.” Indeed, it is these very shortcomings of relying solely on well imaging to support clonal derivation, that Solentim developed the VIPS.  Within the VIPS, the use of accurate low volume dispensing and smart real-time imaging leads to a greater confidence of seeing a single cell deposited in the well.

The paper is at pains to explain that the need to carry out cloning steps is not just for the sake of regulatory expectation, but rather is also driven by scientific necessity and operational excellence with the outcome on the patient being top of mind.

The paper reflects upon the natural drift within CHO cells (see below).  There is emphasis how variability is magnified within an unintentionally non clonally-derived cell bank (Figure 1, graph A) caused by seeding of 2 cells during development of the cell bank, versus 1 cell seeded (Figure 1, graph B) and the potential impact on manufacturing change and lifecycle of a biopharma product:

Figure 1. Variability of unintentionally non clonally-derived cell bank and potential effect on manufacturing change (orange arrows depict range of values at time of harvest, blue squares depict mean of range). Time zero on y-axis reflects single cell isolation. Figure taken from paper by Welch and Arden.

“During culturing, phenotypic drift occurs, impacting the genetic population mean at the time of harvest which results in that population having a mean (depicted as a blue square), and a range (depicted by orange arrows) for a given attribute. Critically, though the nonclonally derived bank may result in the same CQA mean value at the time of harvest as the clonally derived bank, its divergent starting point during cloning could potentially result in greater variability of one or more CQA's, and potentially higher susceptibility to drift, shift, and unforeseen selective pressures during manufacturing lifetime, depicted by the manufacturing change shown as the blue arrow. Such variations within a cell bank may have impacts that could be observed (if quantifiable) in product performance.”

Consistent with prior public presentations, the authors repeat that clonal derivation should not be considered separately from all other elements comprising the control strategy. They highlight that regulatory emphasis should be placed on ensuring the quality of the product administered and, with it, “ensuring the continued availability of medically necessary drugs to patients (e.g., reducing the risk of drug shortages). Moreover, a determination of the probability of a master cell bank reflecting “a clonally-derived population” and the nature of the corresponding risk is not a determination made in a vacuum, rather it is considered in the context of the overall control strategy proposed by the sponsor”.  With greater complexity in the market lifecycle of drugs. It is important to be sure in the consistency of the cell bank to allow flexibility in changing manufacturing scales, or even moving to perfusion cultures to meet market demand. 

Precision cell imaging for analytical clonality

Importantly with respect to imaging, the authors note that “regulators should request sufficient details to evaluate the process and procedures associated with the clonal derivation. Data, such as images in support of what happened during cell bank generation should be available for review and consideration.” This is the focus Solentim has taken with its Cell Metric and VIPS products, using high resolution cell imaging system to give an analytical assurance of clonal derivation, and ultimately being able to provide users a harmonised Clonality Report.

Finally, in the concluding remarks they highlight that “clonal derivation of production cell lines is a factor that has real and tangible potential impact on reproducible manufacture and product quality attributes.”

For full copy of the paper: Please cite this article as: Joel T. Welch and N. Sarah Arden, Biologicals,