In June of 1936 the United States Pharmacopeia (USP) implemented the sterility test to better ensure the sterility of parenteral drug products. The test utilized one type of media and applied only to sterile liquids. Since then slight modifications have been made to include a broader range of product types and to increase the overall sensitivity of the method. However, 80 years later the test remains virtually unchanged. Due to advances in microbial detection technologies and a strong initiative from regulatory authorities, alternative sterility testing methods are beginning to take flight.
The addition of chapter <1223> validation of alternative microbiological methods to the USP provides stakeholders with a clear path to validate alternative testing methods. This chapter is clearly the result of the accelerated pace at which alternative detection technologies are advancing and allows manufacturers and testing laboratories alike to validate alternative methodologies in accordance with the outlined criteria. As long as the alternative method yields results equivalent to or better than the compendial method, the proposed alternative method may be validated and used for routine product release.
The current incubation requirements outlined in USP <71> mandate a 14-day incubation period. As many manufacturers contract out their sterility testing, it’s possible that up to 16-17 days (testing + quality assurance time) may pass prior to the release of a final report. Although this time to result has little impact on some industries/product types, certain industries that manufacture short shelf life products require a more rapid release test.
The Need for Rapid Methods
Pharmacy compounding is a practice where licensed personnel combine drug ingredients to create a personalized medicine. Unlike commercially manufactured products that have multi-year expiration dates, compounded preparations typically have beyond use dates (BUD) in days or months. This makes it difficult for manufacturers to maintain an inventory of high shelf life products. To further complicate things, the recent proposed changes to the California Code of Regulations states that sterile compounded drug products may no longer be able to be dispensed until an acceptable sterility test result is in hand. The inherent short shelf life of the products and proposed California regulatory changes further highlight the necessity for a more rapid method.
A similar issue exists for products that need to be administered to patients shortly after manufacturing. Cell therapy manufacturers continue to struggle with comprehending the possibility of obtaining a 14-day sterility test result prior to drug administration. Although some companies are able to freeze their product prior to obtaining batch release results, certain manufacturers greatly benefit from the ability to administer fresh, non-frozen drug product. To achieve this preferred product delivery, obtaining a 14-day sterility test result prior to drug administration is not possible. As per the Food and Drug Administration (FDA) Chemistry Manufacturing and Controls (CMC) guidance document “Information for Human Somatic Cell Therapy Investigational New Drug Applications (INDs)”, manufacturers are given the task to develop an approach to demonstrate sterility assurance for products that must be administered to patients prior to the results of a 14-day sterility test being available. The document recommends an in-process sterility test to be performed on samples collected 48 to 72 hours prior to final harvest or after the last re-feeding of cultures. Gibraltar has developed several of these in-process tests utilizing our rapid sterility technology to provide a high level of sterility assurance for the full panel of USP <71> organisms in only 48 hours in order to help our customers meet the required criteria.
About the Technology
The Milliflex® Rapid Test System by EMD Millipore is a rapid microbial detection instrument capable of detecting low levels of microbial contamination. The innovative system utilizes a novel membrane filtration system where microbial adenosine triphosphate (ATP) is detected in the form of a bioluminescent signal. Following sample filtration, a media cassette is combined with the filter and incubated for a set period of time. Once incubation is complete, an automated system is used to detect ATP bioluminescence. If the level of ATP detected supersedes the calibrated background threshold, the luminescence is recorded as a CFU.
Validation Strategy and Findings
The Milliflex® instrument underwent an extensive validation plan to demonstrate equivalence to the compendial USP <71> and EP 2.6.1 sterility test. Ruggedness, robustness, specificity, limit of detection and other criteria were assessed to demonstrate the effectiveness of the system. Ultimately, a five-day detection plan was administered as that was the minimum amount of time required to recover growth of a wide variety of environmental and clinical isolates including the notoriously slow growing anaerobe Propionibacterium acnes (4-days + 1-day for assurance). We performed a similar validation at Gibraltar and achieved comparable results. To truly understand the detection power of the instrument we attempted to recover <10 CFUs of all USP <71> organisms at varying incubation times. We were able to detect the presence of all bacteria (Staphylococcus aureus ATCC# 6538, Pseudomonas aeruginosa ATCC# 9027, Bacillus subtilis ATCC# 6633 and Clostridium sporogenes ATCC# 11437 and yeast (Candida albicans ATCC# 10231) in as few as 12 hours and mold (Aspergillus brasiliensis ATCC# 16404) in as few as 24 hours when utilizing the instrument. Despite the powerful results, we generally use a five-day incubation period to allow for any slow growing or stressed contaminants to grow. However, as indicated by our early detection data, the Milliflex® rapid system remains a viable option to provide reliable sterility assurance data for short shelf life products that need to be administered or released before a 5- or 14-day sterility test result is available.
The Milliflex® rapid sterility system is a non-destructive instrument that provides fully automated 21 CFR part 11 compliant data interpretation. Results are dependable and reproducible with a LOD of 1 CFU. The instrument is compatible with a wide variety of membrane filterable products and provides growth-based results consistent with compendial methodology in five days or less. Therefore, this is a quantitative or qualitative growth based method where identification of a positive result is possible. It is important to note that other rapid sterility test systems available do not allow for the microbial identification of a positive result but instead rely on non-growth methods. This means that in the event of a sterility failure, a repeat of the test with the same number of units from the same lot is most likely the only option and the identity of that particular failure will never be known. Gibraltar advises potential users to inquire about the ability of the proposed system to be used for microbial identifications prior to signing on with a rapid sterility test system. We invite any potential users to contact Gibraltar Laboratories to learn more about rapid sterility testing.