Formulation and Antiviral Treatments

Formulation
In order that biomaterials, and particularly biopharmaceutical proteinbased therapeutic drugs, can be maintained and delivered in a biologically active state, it is often necessary to formulate purified materials in excipients and additives that protect the product from deleterious reactions and degradations that compromise the authenticity of such products. This can be achieved in either the solid (lyophilised) or solution state and both are routinely used for the preparation of such products.
Current methods for determining the best formulation(s) for preservation, stability and delivery rely heavily upon trial-and-error approaches. Preservation is then usually investigated using higher temperature and varying pH studies in order to ‘force’ stability issues and stability is then investigated routinely using isoelectric focusing, SDS-PAGE and fluorescence methodology. Particularly with regard to antibody-based therapies where relatively large amounts of material are required per dose,there is interest in the development of high-concentration liquid formulations (4100 mgmL^-1) destined for clinical use. In particular, there is muchacademic and industrial interest in developing our ability to predict appropriate formulation strategies and compositions for particular proteins that would negate the need for time-consuming trial-and-error studies.
 Antiviral Treatments
Ever since the transmission of HIV infection to haemophiliac patients through contaminated blood-based products in the early 1980s, there have been stricter controls and scrutiny of the methods of viral inactivation utilised during bioprocessing. This is of particular concern when using mammalian cells to produce biotherapeutic products, as there is clear potential to host harmful viruses. Current good practice therefore dictates that products destined for use as therapeutics are submitted to antiviral treatments with two different viral-inactivation steps each of which gives a four to five log reduction in virus load. To achieve this level of reduction, several steps with viral removal functionality are required, although some of these steps may have dual functions. The methods employed to achieve this are varied and the regulatory bodies do not require specific viral inactivation treatments but rather judge the validity of the approach used for each product on its own merits.
The most commonly used methods to reduce viral load are the heating of protein solutions and lyophilisates, nanofiltration, low-pH hold steps, UV irradiation, solvent/detergent treatments and those process removal steps that are part of the purification methodology or the production of the protein of interest. Of these treatments, heat treatment is the current ‘gold standard’ and many therapeutic protein preparations are treated in this way (e.g. albumin, Factor VIII, Factor IX, thrombin, fibrinogen, IgG). Further, heat treatment has the advantage of being one of the few methods that can be applied to a given product in the vial without the need for disruption of the product. Typical heat treatment in the liquid state consists of 60 1C for 10 h, whereas in the lyophilised state samples are typically subjected to temperatures of 90 1C for 10 h or 80 1C for 72 h. The disadvantage of heat treatment is that as proteins denature and undergo deleterious reactions upon heat stress, it is necessary to formulate the protein of interest in stabilising excipients such as sucrose to prevent such reactions. It has been shown that the excipients themselves can break down and react with the protein of interest under certain heat
treatment conditions, resulting in disadvantageous side reactions that compromise the authenticity of the protein product.