Volume 7: Medicines and Cosmetics
2. The operation of medicines licensing
Annex 2 to Chapter 2: Outline of production processes for vaccines and injectable medicinal products
Vaccines
Injectable medicines derived from bovine tissues

Vaccines

2.124 Administered to healthy individuals, vaccines protect against infection caused by pathogens, eg, viruses, bacteria, by inducing immunity. Some vaccines contain preparations of an infective agent which are non-pathogenic and do not cause disease. For example, killed vaccines are preparations of a pathogen that has been killed or inactivated. Other vaccines contain attenuated (reduced infectivity) live strains of the agent which do not cause disease but which still induce specific immunity. Some vaccines are produced using so-called recombinant DNA technology where only the antigenic parts of the pathogen, ie, those parts which induce immunity in an individual, are used in the vaccine. Only in this latter class of vaccines are bovine products used during manufacture.

2.125 Recombinant vaccines are produced by isolating the DNA sequence that specifies the particular antigenic (ie, capable of inducing a specific immune response) portion of the pathogen and integrating it into bacterial or animal cells growing in culture. The cells then manufacture the foreign antigenic molecule as if it were a normal constituent of the cell. The antigen can then be purified away from the bacterial or animal cells and used in a vaccine. Alternatively, the antigen can be integrated into a non-pathogenic virus strain, which is grown in animal cells. In this case, the viruses are harvested from the animal cells and suitably treated to become vaccines.

2.126 As illustrated by the examples in Table 2 of Annex 1, various bovine materials are used as ingredients in cell cultures for the production of vaccines. These are most commonly blood products, milk derivatives and peptone.

2.127 In order for animal cells to survive in culture, it is necessary to grow them in nutrient-rich media. These nutrients are generally supplied from bovine serum derived from foetal or new-born calves - foetal calf serum (FCS) and new-born calf serum (NCS) respectively. In these rapidly developing animals, serum is packed with nutrients and important growth factors. Serum from older animals is sometimes used, though it is much less rich. A further bovine blood product, bovine serum albumin (BSA), is often added to serum for nutritional purposes.

2.128 In the production of FCS, blood is removed from the foetus by insertion of a needle into its heart following the death of the dam. Blood is removed under vacuum (rarely by gravity flow) and is drawn directly into a bag to reduce any contamination of the blood. This process occurs at the abattoir. Serum is separated from the blood by centrifugation, filtered to remove large particulates and sterile-filtered to remove particulates 1 micron in size or larger. The serum is then bottled and frozen. NCS is prepared in the same way, except that the animals are between 10 and 14 days old when the blood is removed. Donor serum is removed from animals up to the age of three years old. The animals are not killed, but are periodically 'tapped' for blood. ¹

2.129 When the cells are harvested, the growth medium is removed and the cells washed to remove the serum. Similarly, viruses used for vaccine production are purified away from both the cells and culture medium before being treated further.

2.130 Bacterial cells are not grown in serum-containing media, but in nutrient broths based on peptone. Peptone is obtained through the chemical treatment of milk or beef, which releases the building blocks of proteins that can be utilised by the bacteria. In some circumstances, beef extract is used in a nutrient broth to feed the bacteria in the same way. Unlike serum, these nutrient additives are very stable and can therefore be sterilised by autoclaving. Digests of calf brain and ox liver have also been constituents of special culture media used in the production of allergens. ² Allergens are similar to vaccines in that they invoke an allergic reaction in an individual as a means of raising general immunity to a particular antigen upon environmental exposure. Whatever the culture medium used, the bacteria are ultimately recovered from the broth and subjected to separation procedures to isolate the component to be used in the vaccine.

2.131 Thus bovine ingredients are not a constituent of the final vaccine product but are used in a peripheral way, for the nourishment of cells used to grow vaccines. The question of risk of transmission of BSE through vaccines was, therefore, linked to the likelihood of any BSE infectivity present in the starting material surviving through to the final product.

Injectable medicines derived from bovine tissues

2.132 Unlike vaccines, in which bovine products are used peripherally during manufacture, there are several injectable medicinal products derived directly from a bovine source. Such products include hormones such as glucagon, insulin and corticosteroids or protein products such as aprotinin and heparin. The tissues from which these products are derived are obtained directly from the abattoir. Following slaughter, the cattle viscera, the larger internal organs present in the chest and abdominal cavities, are removed by slaughtermen. These are then passed either to in-house workers in a separate 'clean' part of the slaughterhouse called the gut room, or to contractors, who separate the organs according to requirements. The appropriate tissues are cleaned and extraneous material removed in accordance with the specifications of the pharmaceutical company. The tissues are frozen on site before transportation to the pharmaceutical company, to prevent breakdown of the required products.

2.133 The products required from the tissues are extracted in the laboratories of the pharmaceutical companies using purification techniques involving many steps. Initially these steps are crude but increase in specificity as more extraneous materials are removed. These later, more specific purification steps involve stages which are appropriate for only the product of interest, and as such utilise particular physical or biochemical properties that would be shared with few other compounds. In this way it is possible to obtain an homogeneous product free from contaminating molecules.

2.134 It might be expected, therefore, that any BSE present in a tissue would be reduced during purification steps. Indeed, the infective agent of BSE is known to be 'sticky' in the absence of detergents, and easily removed from tissue extracts by centrifugation (the process of separating particles based on size and density under centrifugal force) since the agent would remain attached to cellular particles and proteins and be removed in the sedimented material. Filtration is also effective, as the agent absorbs to the filters themselves. ³