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CYTOKINES: INTERLEUKINS AND TUMOUR NECROSIS FACTOR 243
provide a means of treating or alleviating chronic or inflammatory conditions associated, at least in part, with eosinophilia (increased eosinophil counts).
A number of strategies may be adopted to inhibit IL-5 activity. Methods currently under investigation include:
• inhibition of IL-5 synthesis/release by T-lymphocytes;
• use of anti-IL-5 monoclonal antibodies or soluble IL-5 receptor polypeptides;
• development of a mutant IL-5 capable of binding its receptor but without triggering signal transduction;
• identification of a low molecular mass substance which, by interacting with either IL-5 or its receptor, would prevent direct interaction of the cytokine and its receptor and hence prevent IL-5 signalling.
A soluble receptor has been identified in vivo which, when produced by recombinant means and administered at higher levels, does act as an effective receptor antagonist. Fab fragments of a humanized anti-IL-5 monoclonal antibody (Chapter 10) have also been produced, which can reduce IL-5-mediated effects in animal models.
Alanine scans (use of genetic engineering to generate a series of mutant proteins in which each residue in the native protein is systematically changed to alanine) of IL-5 have identified several residues important in ligand-receptor interaction. Replacement of IL-5 residue No. 13 (usually Glu) with alanine, while having no effect upon receptor binding, almost totally prevents subsequent initiation of signal transduction.
The ongoing (chronic) nature of many inflammatory conditions renders most attractive the development of a drug which would:
• exhibit a long circulatory half-life;
• be relatively inexpensive;
• be capable of being administered orally.
All the protein-based potential therapeutics discussed above fail to meet any of these criteria. Low molecular mass antagonists of IL-5 or its receptor may prove of most use. A number of research teams are screening for such substances. The development strategies pursued include:
• computer-aided drug design; attempts are being made to logically design molecules capable of binding to specific regions of IL-5 (or its receptor) and hence blocking their function;
• traditional screening methods; the screening of microbial, plant or other extracts for substances capable of interfering with IL-5 ligand-receptor interaction.
Thus far, at least one class of compound — isothiazolone derivatives — has been identified as potent IL-5 inhibitors. These substances react with a free sulphydryl group on the surface of the IL-5 receptor a-subunit, thereby preventing ligand binding. Isothiazolones are already used as anti-microbial agents, but are quite toxic.
Overall, therefore, inhibition of IL-5-mediated biological activities may be achieved, at least in part, by a number of means. All studies to date, however, have been undertaken in vitro. Clinical trials will be necessary to assess whether any of these strategies are effective in treating chronic asthma or other inflammatory conditions.
Interleukin 12 represents a cytokine that has just begun to generate substantial clinical interest. This is due to its ability to generate a very strong cell-mediated immune response. IL-12 is a heterodimeric structure composed of two polypeptide subunits, termed p35 and p40. p35 is a 196 amino acid glycoprotein exhibiting a molecular mass in the 30-35 kDa range. p40, containing 306 amino acids, is also glycosylated and exhibits a molecular mass in the 35-45 kDa range. Neither chain on its own exhibits biological activity . Monocytes/macrophages appear to represent the major cellular source of IL-12. Sensitive cells express a cell surface 180 kDa glycoprotein, which serves as the IL-12 receptor. The mechanism of signal transduction remains to be characterized. Whatever its mechanism of signal transduction, IL-12 induces a number of important biological effects, including:
• stimulation of IFN-g synthesis and release from T lymphocytes and NK cells;
• activation of NK cells;
• induction of the growth and differentiation of precursor T lymphocytes specifically producing TH1 cells.
Such biological activities suggest various clinical scenarios in which this cytokine may be positively employed (Table 5.8).
TH1 cells (i.e. T helper type 1 lymphocytes) are the cellular population that triggers a cell-mediated immune response. This is the most effective means by which the immune system can identify and destroy many foreign agents, particularly viruses, bacteria or parasites, that have entered host cells.
In many instances, the unaided immune response to such infections can be inadequate, leading to prolonged and/or serious illness. Examples include the malaria and tuberculosis agents, the HIV virus (which has infected at least 14 million people worldwide), leishmaniasis (caused by the protozoan Leishmania, from which 12 million people suffer) and schistosomiasis, (caused by the Schistoma mansoni helminth, which currently infects approximately 250 million people worldwide).