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biopharmaceuticals biochemistry and biotecnology - Walsh G.

Walsh G. biopharmaceuticals biochemistry and biotecnology - John Wiley & Sons, 2003. - 572 p.
ISBN 0-470-84327-6
Download (direct link): biochemistryandbiotechnology2003.pdf
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Neumega is the trade name given to the IL-11-based product approved for the prevention of thrombocytopenia. The product is produced in engineered E. coli cells and is presented as a purified product in freeze-dried format. Excipients include phosphate buffer salts and glycine. It is reconstituted (with water for injections) to a concentration of 5 mg/ml before s.c. administration.
The biological activities of several other interleukins also render them likely candidates for therapeutic application. IL-5 represents one such candidate. IL-5 is a 115 amino acid glycoprotein produced mainly by activated T lymphocytes and also by mast cells. It functions as a homodimer, exhibiting a molecular mass of 45 kDa. The individual polypeptide chains interact non-covalently and the overall dimeric structure is stabilized by two interchain disulphide linkages between cysteines 42 and 84 of each chain. Removal or alterations of the cytokine’s carbohydrate side-chain does not appear to affect its biological activity.
X-ray diffractional analysis of crystalline human IL-5 reveals a two-domain structure, with each domain consisting of four a-helical stretches and one b-stretch consisting of two antiparallel b-strands.
The human IL-5 receptor consists of two transmembrane glycoprotein subunits. The glycosylated 60 kDa a-chain binds IL-5 but is unable to induce direct signal transduction. A b-chain, unable to bind IL-5 directly, is associated with the a-chain to form an intact receptor complex. This 130 kDa b-chain, which also constitutes part of the IL-3 and GM-CSF receptors, appears to initiate signal transduction. In addition to the form anchored in the plasma membrane, cells are capable of producing a mRNA coding for a soluble form of the receptor a-chain by a process of alternative splicing. The exact physiological significance of this is unclear, as expression/secretion of this alternative mRNA in humans appears very low. When the corresponding cDNA is expressed in animal cell lines, and the soluble receptor purified and subjected to ligand binding studies, it was found to compete effectively with the membrane receptor for IL-5 binding. Clearly, if synthesized and secreted by human cells, this soluble receptor protein could have an antagonistic effect on IL-5 bioactivity. Soluble receptors for a number of additional human cytokines (some already discussed) have also been discovered (Table 5.7).
In humans, IL-5 functions as a growth and differentiation factor for eosinophils. It also stimulates basophil growth and may influence B lymphocyte activity. Eosinophils are leukocytes containing large cytoplasmic granules. The granules contain basic proteins capable of binding to acidic dyes such as eosin. The two main granular proteins are eosinophil cationic protein and major basic protein. Eosinophils play a particularly important role in mediating the destruction
Table 5.7. Cytokines for which soluble receptors have been detected in biological fluids. The exact functional role played by these binding proteins, in modulating cytokine activity, remains to be elucidated in most cases
IL-2 IL-7
IL-4 IFN-g
Figure 5.9. The destruction of parasites by eosinophils. Eosinophils bind to opsonized parasites via IgE-Fc receptors expressed on their surface. Binding induces release of granular contents by exocytosis. Major basic protein, the most abundant granular-derived protein, then initiates destruction of the parasite by lysis
of helminth parasites. Helminths stimulate IgE synthesis, which opsonize the parasite by binding to its surface antigens. Eosinophils bind the opsonized particle and secrete their granular contents, which induces destruction of the parasite by lysis (Figure 5.9). While IL-5 may have therapeutic potential in the treatment of some helminth infections, the role it plays in mediating acute inflammatory disease is currently receiving most attention.
Eosinophils are believed to play a major role in many inflammatory and allergic conditions. These effects are mediated by release of their granular contents, which can induce local host cell damage. One common such condition, affecting 10 million people in the USA alone, is asthma. Most forms of asthma are actually immediate hypersensitivity reactions (i.e. inappropriate overreaction of certain elements of immunity to selected antigens). Asthma can also be characterized as an inflammatory disease, involving certain T helper lymphocytes (which also produce IL-5) and eosinophils in the airway mucosa. The granular eosinophil contents may play an important role in mediating chronic bronchial inflammation.
While eosinophil production can be induced by IL-3, IL-5 and GM-CSF in vitro, only IL-5 plays this role effectively in vivo. It follows that blocking the biological activity of IL-5 may
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