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Lessons learned from contamination with endotoxin originated from the supplement in the cell culture medium
Koaki Uehara1, Eriko Oshiro1, Atsushi Ochiai1
1Social Medical Corporation Yuuaikai, Yuuai Medical Center, Advanced Medical Research Center, 50-5, Yone, Tomigusuku-shi, Okinawa 901-0224, Japan.
Introduction:
Endotoxin is a typical pyrogen derived from the outer membrane of Gram-negative bacteria. In fabricating cell-based medicinal products, it is necessary to control endotoxin in the process and the products. In the quality control tests of our clinical study, endotoxin concentration in the culture supernatant of autologous oral mucosal epithelial cell sheets exceeded the criterion value. Therefore, endotoxin measurements were conducted to clarify the cause of the endotoxin contamination.
Methods:
The reagents used to prepare the culture medium, the unused culture medium, and the culture supernatants were diluted with pure water. Endotoxin concentrations in the diluted samples were measured.
Results:
Endotoxin was detected in both the unused culture medium and the culture supernatant of the epithelial cell sheets at higher concentrations than the criterion value. Therefore, endotoxin concentrations in the reagents used to prepare the culture medium were measured and were found to be below the criterion value, except for cholera toxin. On the other hand, three lots of cholera toxin products were used for the measurement, and the endotoxin concentrations were higher than the criterion value. The results indicate that the endotoxin contamination is caused by the cholera toxin product.
Conclusions:
To prevent endotoxin contamination in cell-based medicinal products, endotoxin concentrations in reagents used for the fabrication should be measured in the facility conducting clinical research or confirmed by an adequate certificate of analysis from the manufacturers of the reagents.
Insights
Endotoxin contamination in cell-based products was traced to cholera toxin reagents. Implementing reagent testing or validated certificates of analysis is crucial for ensuring product safety and quality.
Area of Science:
- Microbiology
- Biotechnology
- Cell Therapy
Background:
- Endotoxins, pyrogens from Gram-negative bacteria, necessitate strict control in cell-based medicinal product fabrication.
- Elevated endotoxin levels were detected in the culture supernatant of autologous oral mucosal epithelial cell sheets during a clinical study.
Purpose of the Study:
- To investigate the source of endotoxin contamination in autologous oral mucosal epithelial cell sheets used in a clinical study.
- To identify specific reagents responsible for endotoxin contamination in cell culture processes.
Main Methods:
- Endotoxin concentrations were measured in culture supernatants, unused culture media, and reagents used for media preparation.
- Samples included epithelial cell sheets, culture media components, and cholera toxin products.
Main Results:
- Endotoxin exceeded acceptable limits in both unused culture medium and epithelial cell sheet supernatant.
- Reagents for culture medium preparation were below limits, except for cholera toxin.
- Three lots of cholera toxin products showed high endotoxin concentrations, identifying it as the contamination source.
Conclusions:
- Cholera toxin products were identified as the primary source of endotoxin contamination.
- Reagent quality control, including endotoxin testing or Certificates of Analysis, is vital for preventing contamination in cell-based product manufacturing.
- Proactive measures in clinical research facilities are essential for ensuring the safety of cell-based medicinal products.
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