Pyrogenic/pro-inflammatory activities
Pyrogens are fever-causing agents which contaminate medicines and vaccines as a consequence of the manufacturing process.
Pyrogen-contaminated parenteral preparations can cause mild to severe clinical reactions including fever, rash, headache, myalgia, nausea, vomiting and hypotension. It is therefore vital that the level of pyrogenic contamination of parenteral preparations is carefully controlled to ensure their safety.
The most common pyrogen is endotoxin which is shed from the cell wall of Gram-negative bacteria. Medicines and vaccines produced by a process involving such bacteria are therefore at high risk of being contaminated with endotoxin.
The amount of endotoxin in a product can be quantified using one of the various methods of the bacterial endotoxins test (BET). In line with the 3Rs principle – the replacement, reduction and refinement of animal testing, the BET was a replacement for the rabbit pyrogen test (RPT). However, unlike the RPT, the BET does not detect non-endotoxin pyrogens or other pro-inflammatory contaminants of parenteral preparations which can stimulate unwanted clinical reactions.
For this reason a third test – the monocyte activation test (MAT) – was developed as a replacement for the RPT for quantifying pro-inflammatory/non-endotoxin pyrogenic contaminants of medicines and vaccines.
All three tests are described in the British and European Pharmacopoeias.
What we do
Our laboratory has considerable expertise in the detection of pyrogenic contaminants of parenteral preparations using the monocyte activation test and bacterial endotoxins test for the purposes of medicines control and research and development.
Standardisation
Our area was responsible for preparing the 2nd and 3rd International Standards (IS) for endotoxin as well as the new standard for the BET described in the US and European Pharmacopoeias.
The 3rd IS for endotoxin also serves as the standard for the European Pharmacopoeial MAT Methods A and B. A total of 3 x 25,000 vials of endotoxin – all with the potency of 10 000 IU of endotoxin per vial – were filled to preserve global harmonisation of unitage for this important standard.
Control
As an the UK’s National Control Laboratory our area is responsible for:
- applying the ISO 17025-compliant, United Kingdom Accreditation Service (UKAS)- accredited, semi-quantitative gel-clot BET and the ISO 17025-compliant, quantitative kinetic chromogenic BET to approximately 40 different products – including products for batch release – amounting to approximately 500 samples a year
- control/batch release testing of vaccines using the MAT, specifically developed for each type of vaccine
- applying the BET/MAT to investigate medicines/vaccines already released but which have been associated with adverse drug reactions
- applying the BET to released medicines and vaccines as part of market surveillance
The role of the pyrogen testing area in medicines control is much wider than carrying out routine testing. We advise about product testing, methodology and regulatory requirements, including setting pharmacopoeial specifications, to the pharmaceutical industry, regulatory organisations and other Official Medicines Control Laboratories (OMCLs) to develop new policies and practices.
In particular, we work with the European Directorate for the Quality of Medicines (EDQM) through the Expert Groups for the MAT and the BET and the OMCL network.
To promote high-quality safety testing of parenteral products in the pharmaceutical industry and at other OMCLs in Europe and in China, we provide support and training for laboratories wanting to use the MAT and novel cell-based assays for the detection of unwanted pro-inflammatory activities of parenteral preparations.
Find out more about NIBSC control testing.
Research
Our work in this area includes developing novel bioassays capable of detecting unwanted intrinsic activities of parenteral preparations including vaccines and therapeutic monoclonal antibodies.
Other research areas are:
- developing existing bioassays/tests to detect pyrogenic/pro-inflammatory contaminants of parenteral preparations
- applying the BET/MAT to research samples such as potential vaccine candidates submitted by pharmaceutical companies and by various groups within NIBSC
- conducting investigations aimed at improving our understanding of the mechanisms of action underlying the pro-inflammatory activity of components from various microorganisms and of parenteral preparations resulting from contamination or as a result of the intrinsic nature of the medicine
- conducting investigations using the MAT on pharmaceutical products which have passed the BET but which have failed the RPT, suggesting contamination with a non-endotoxin pyrogen.
An example of this is a study we carried out in collaboration with a vaccine manufacturer to identify which component (and its batch) of a multivalent vaccine was causing problems with the vaccine’s pro-inflammatory activity.