Peptide vaccines

An alternative approach to the production of subunit vaccines entails their direct chemical synthesis. Peptides identical in sequence to short stretches of pathogen-derived polypeptide antigens can be easily and economically synthesized. The feasibility of this approach was first verified in the 1960s, when a hexapeptide purified from the enzymatic digest of tobacco mosaic virus was found to confer limited immunological protection against subsequent administration of the intact virus. (The...

Recombinant veterinary vaccines

Amongst the limited number of biopharmaceuticals approved for animal use, recombinant vaccines represent the single largest subgroup. Several such products target pigs, including 'Porcilis pesti' and 'Bayovac CSF E2'. Porcilis pesti, for example, contains a recombinant form of the classical swine fever virus E2 antigen, the immunodominant surface antigen associated with this viral pathogen. It is used to immunize young pigs. An overview of its manufacture is presented in Figure 13.14. The...

Additional means of insulin administration

Issues surrounding protein delivery via means alternative to parenteral routes have been outlined in Chapter 4. One such product (inhalable insulin, tradename Exubera) has gained marketing approval (Box 11.2). An additional approach that may mimic more closely the normal changes in blood insulin levels entails the use of infusion systems that constantly deliver insulin to the patient. The simplest design in this regard is termed an 'open-loop system'. This consists of an infusion pump that...

Additional recombinant hormones now approved

Three additional recombinant hormones have recently gained marketing approval thyroid-stimulating hormone, parathyroid hormone and calcitonin. Structurally, thyroid-stimulating hormone (TSH or thyrotrophin) is classified as a member of the gonadotrophin family, although functionally it targets the thyroid gland as opposed to the gonads. As with other gonadotrophins, it is a heterodimeric glycoprotein displaying a common a-subunit and a unique P-subunit. The P-subunit shows less homology to that...

Cell disruption

Disruption of microbial cells is rendered difficult due to the presence of the microbial cell wall. Despite this, a number of very efficient systems exist that are capable of disrupting large quantities of microbial biomass (Table 6.1). Disruption techniques, such as sonication or treatment with the enzyme lysozyme, are usually confined to laboratory-scale operations, due either to equipment limitations or on economic grounds. Protein extraction procedures employing chemicals such as detergents...

Engineered insulins

Recombinant DNA technology facilitates not only production of human insulin in microbial systems, but also facilitates generation of insulins of modified amino acid sequences. The major aims of generating such engineered insulin analogues include Identification of insulins with altered pharmacokinetic properties, such as faster-acting or slower-acting insulins. Identification of super-potent insulin forms (insulins with higher receptor affinities). This is due to commercial considerations,...

The insulin receptor and signal transduction

The insulin receptor is a tetrameric integral membrane glycoprotein consisting of two 735 amino acid a-chains and two 620 amino acid P-chains. These are held together by disulfide linkages (Figure 11.2). The a-chain resides entirely on the extracellular side of the plasma membrane and contains the cysteine-rich insulin-binding domain. Each P-subunit is composed of three regions the extracellular domain, the transmembrane domain and a large cytoplasmic domain that displays tyrosine kinase...

Therapeutic application of monoclonal antibodies

The unrivalled specificity of monoclonal antibodies, coupled to their relatively straightforward production and their continuity of supply, renders them attractive biochemical tools. Therapeu-tically, they represent by far the single largest category of biopharmaceutical substances under investigation. Several hundred such preparations are currently undergoing preclinical and clinical trials. Throughout the 1980s the focus of attention rested upon their use either as in vivo imaging (i.e....

Traditional vaccine preparations

For the purposes of this discussion, the term 'traditional' refers to those vaccines whose development predated the advent of recombinant DNA technology. Approximately 30 such vaccines Table 13.5 Some diseases against which effective more effective vaccines are urgently required. Diseases more prevalent in developing world regions differ from those that are most common in developed countries Table 13.5 Some diseases against which effective more effective vaccines are urgently required. Diseases...

AGalactosidase urate oxidase and laronidase

Recombinant a-galactosidase, urate oxidase and laronidase represent additional biopharmaceuti-cals recently approved for general medical use. a-Galactosidase is approved for long-term enzyme replacement therapy in patients with Fabry disease. Like Gaucher's disease, Fabry disease is a genetic disease of lipid metabolism. Sufferers display little or no liposomal a-galactosidase-A activity. This results in the progressive accumulation of glycosphingolipids in several body cell types. Resultant...

Product case study Humalog

Humalog (tradename, also known as insulin lispro) was the first recombinant fast-acting insulin analogue to gain marketing approval (in 1996). It is indicated for the treatment of diabetes mellitus, for the control of hyperglycaemia and is used in conjunction with long-acting insulins (see main text). It is administered subcutaneously. The product displays an amino acid sequence identical to native human insulin with one alteration, i.e. an inversion of the natural proline-lysine sequence found...

Protein mode of action and pharmacodynamics

Different protein therapeutics bring about their therapeutic effect in different ways (Figure 4.8). Hormones and additional regulatory molecules invariably achieve their effect by binding to a specific cell surface receptor, with receptor binding triggering intracellular signal transduction event(s) that ultimately mediate the observed physiological effect(s). Many antibodies, on the other hand, bring about their effect by binding to their specific target molecule, which either inactivates...

Sources and medical uses of folliclestimulating hormone luteinizing hormone and human chorionic gonadotrophin

Although the human pituitary is the obvious source of human gonadotrophins, it also constitutes an impractical source of medically useful quantities of these hormones. However, the urine of post-menopausal women does contain both FSH and LH activity. Up until relatively recently, this has served as the major source used medically, particularly of FSH. Menotrophin (human menopausal gonadotrophin) is the name given to FSH-enriched extracts from human urine. Such preparations contain variable...

Vaccine vectors

An alternative approach to the development of novel vaccine products entails the use of live vaccine vectors. The strategy followed involves incorporation of a gene cDNA coding for a pathogen-derived antigen into a non-pathogenic species. If the resultant recombinant vector expresses the gene product on its surface, then it may be used to immunize against the pathogen of interest (Figure 13.12). Most vaccine vectors developed to date are viral based, with poxviruses (as well as picorna viruses...

The impact of genetic engineering on vaccine technology

The advent of recombinant DNA technology has rendered possible the large-scale production of polypeptides normally present on the surface of virtually any pathogen. These polypeptides, when purified from the producer organism (e.g. E. coli, Saccharomyces cerevisiae) can then be used as 'subunit' vaccines. This method of vaccine production exhibits several advantages over conventional vaccine production methodologies. These include Production of a clinically safe product the pathogen-derived...

Insulin production

Traditionally, commercial insulin preparations were produced by direct extraction from pancreatic tissue of slaughterhouse pigs and cattle, followed by multistep chromatographic purification. However, the use of animal-derived product had a number of potential disadvantages, including Figure 11.2 Structure of the insulin receptor (a). Binding of insulin promotes autophosphorylation of the p-subunits, where each p-subunit phosphorylates the other p-subunit. Phosphate groups are attached to three...

Production of human insulin by recombinant DNA technology

Human insulin produced by recombinant DNA technology was first approved for general medical use in 1982, initially in the USA, West Germany, the UK and The Netherlands. As such, it was the first product of recombinant DNA technology to be approved for therapeutic use in humans. From the 1990s on, several engineered insulin products (discussed later) also gained approval (Table 11.3). The initial approach to recombinant insulin production taken entailed inserting the nucleotide sequence coding...

The insulin molecule

Insulin was first identified as an anti-diabetic factor in 1921, and was introduced clinically the following year. Its complete amino acid sequence was determined in 1951. Although mature insulin is a dimeric structure, it is synthesized as a single polypeptide precursor, i.e. preproinsulin. This 108 amino acid polypeptide contains a 23 amino acid signal sequence at its amino terminal end. This guides it through the endoplasmic reticulum membrane, where the signal sequence is removed by a...

Formulation of insulin products

Intermediate Acting Insulin Peak

Insulin, whatever its source, may be formulated in a number of ways, generally in order to alter its pharmacokinetic profile. Fast (short)-acting insulins are those preparations that yield an elevated blood insulin concentration relatively quickly after their administration (which is usually by s.c. or, less commonly, by i.m. injection). Slow-acting insulins, on the other hand, enter the circulation Table 11.3 Native and engineered human insulin preparations that have gained approval for...

Interferon biotechnology

The antiviral and anti-proliferative activity of interferons, as well as their ability to modulate the immune and inflammatory response renders obvious their potential medical application. This has culminated in the approval for clinical use of several interferon preparations (Table 8.8). Ongoing clinical trials are likely to expand the medical uses of these regulatory molecules further over the next few years. While at least some of these potential therapeutic applications were appreciated as...

Protein stability and folding

Upon biosynthesis, a polypeptide folds into its native conformation, which is structurally stable and functionally active. The conformation adopted ultimately depends upon the polypeptide's amino acid sequence, explaining why different polypeptide types have different characteristic conformations. We have previously noted that stretches of secondary structure are stabilized by short-range interactions between adjacent amino acid residues. Tertiary structure, on the other hand, is stabilized by...

Protein posttranslational modification

Many polypeptides undergo covalent modification after or sometimes during their ribosomal assembly. The most commonly observed such PTMs are listed in Table 2.7. Such modifications generally influence either the biological activity or the structural stability of the polypeptide. The majority of therapeutic proteins bear some form of PTM. Although glycosylation represents the most common such modification, additional PTMs important in a biopharmaceutical context include carboxylation,...