Continuing on from our series of blogs discussing E&L assessment in the production of CGTs, we present a case study on single-use components. This is ahead of an online Q&A session with a panel of experts on this area of E&L testing, which you can watch on demand here. This is a valuable chance to access know-how and find out what’s going on in the industry.

Parts 1 and part 2 of this series can be found by following the links.

Simulation Studies

In this blog we will look extractable data designed to simulate the in-use conditions more closely than a traditional controlled extraction study, these studies are commonly referred to as simulation studies.

Due to the batch sizes, it can be difficult to perform leachable studies on CGTs (especially those products developed for rare diseases) it might be appropriate to further understand the risk of patient exposure to leachables through a well-designed simulation study.

This particular study was designed to answer the following question:

What effect does long term frozen storage have on the leachable profile?

We might assume that:

  • Solid-solid interactions are generally considered low risk
  • Thawing processes considered to have the biggest impact

The study was designed to look at the long term storage of drug product in a single layer polyolefin cryogenic bag with Ethylene-vinyl acetate (EVA) line and Polyvinyl Chloride (PVC) leads. The bag was gamma-irradiated and a surface area to volume ratio of 0.588 cm2/mL was used.

The following solvent was used as a worst-case simulation solution for the product (Cells, 5% DMSO, 2.5% HSA in saline):

Excipient Concentration
IPA 10% (v/v)
DMSO 10% (v/v)
NaCl 1% (w/v)
Water QS

Samples were stored under the following conditions:

  • Room temperature for 1 month
  • -20°C for 1 year
  • -20°C + 1 week at RT for 1 year
  • -80°C for 1 year
  • -80°C + 1 week at RT for 1 year

Analysis was performed using headspace GC-MS, direct injection GC-MS, LC-UV-MS and ICP-MS.

Two extractables were detected above the reporting threshold via LC-UV-MS and silicone was detected via ICP-MS.

  • 2,​4-​dioxo-tridecanoic acid observed at MN1 (RT)
  • Low levels of Silicon were detected at MN9 and 1 year

The table below shows the concentration of Polyethoxylated alcohols detected at various time points.

The data appears to show that 1-month extraction at RT the analyte levels are greatest.

Storing frozen drug product slows the migration of extractables to negligible levels, whilst the thawing process accelerates the amount of analyte detected.

Storage at -20°C or -80°C did not have a significant effect of the amount of compound detected.

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This concludes our series on extractable and leachable studies in the field of cell and gene therapy. Please, join the free Q&A to find out more about the subject, join the discussion, and have your questions answered by a panel of experts.

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