Enzymes of therapeutic value

Enzymes are used for a variety of therapeutic purposes, the most significant of which are listed in Table 9.12. A number of specific examples have already been discussed in detail within this chapter, including tPA, urokinase and factor IXa. The additional therapeutic enzymes now become the focus of the remainder of the chapter. 

Liposomes, in particular, may offer the future possibility of delivering enzymes (and other drugs) to specific body tissues. This could be attained by incorporating target molecules (e.g. specific receptors, glycoproteins or antibodies) in the liposome membrane, which selectively 

HSA is used therapeutically as an aqueous solution it is available in concentrated form (15-25 per cent protein) or as an isotonic solution (4-5 per cent protein). In both cases, in excess of 95 per cent of the protein present is albumin. It can be prepared by fractionation from normal plasma or serum, or purified from placentas. The source material must first be screened for the presence of indicator pathogens. After purification, a suitable stabilizer (often sodium caprylate) is added, but no preservative. The solution is then sterilized by filtration and aseptically filled into final sterile containers. The relative heat stability of HSA allows a measure of subsequent heat treatment, which further reduces the risk of accidental transmission of viable pathogens (particularly viruses). This treatment normally entails heating the product to 60 °C for 10 h. It is then normally incubated at 30-32 °C for a further 14 days and subsequently examined for any signs of microbial growth. 

HSA is used as a plasma expander in the treatment of haemorrhage, shock, burns and oedema, as well as being administered to some patients after surgery. For adults, an initial infusion containing at least 25 g of albumin is used. The annual world demand for HSA exceeds 3001, representing a market value of the order of US 1 billion. 

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A potential for endogenous excitotoxin produced in the CNS via tryptophan metabolism has been localised to microglia.97 Microglia contain indoleamine-2,3-dioxy-genase (the first enzyme in this pathway), which converts tryptophan to kynurenine, and which is induced in microglia and macrophages by IL-1 and infections.98 Inhibitors of quinolinic acid production such as 4-chloro-3-hydroxyanthranilate and ///-nitrobenzylalanine99 100 could probably be of therapeutic value. 

The role of therapeutic inhibitors of elastase is to prevent the degradation of extracellular matrix proteins following the release of elastase from activated neutrophils. To determine which inhibitors of elastase may be of therapeutic value, there are several properties that must be considered. These include rate of proteinase inactivation, enzyme selectivity, size, susceptibility to inactivation, and their potential for antigenicity. Some of these properties are reviewed in the following section. 

DNase, a well characterized and biologically specific enzyme with therapeutic value in the treatment of cystic fibrosis, was chosen as a model agent for studying the effects on an enzymatic bio/molecule of surface immobilization on nanocrystalline ceramic particulates. As the data show, and consistent with previous observations, DNase exhibits a marked retention of biological activity when surface immobilized on the solid phase of a colloid comprised of polyhydroxyl oligomeric films investing degradable calcium-phosphate nanoparticles. 

An impressive example of the application of structure-based methods was the design of a inhibitor of the HIV protease by a group of scientists at DuPont Merck [Lam et al. 1994 This enzyme is crucial to the replication of the HIV virus, and inhibitors have bee shown to have therapeutic value as components of anti-AIDS treatment regimes. The star1 ing point for their work was a series of X-ray crystal structures of the enzyme with number of inhibitors boimd. Their objective was to discover potent, novel leads whid were orally available. Many of the previously reported inhibitors of this enzyme possessei substantial peptide character, and so were biologically unstable, poorly absorbed am rapidly metabolised. 

The molecule 7-aminopyrazolopyrimidine is related to the DNA base adenine. It is the base attached to ribose in formycin A, which is believed to have potential therapeutic value. It is also shown in Figure 5. There is a paradox in this system. This molecule is deactivated by the enzyme adenosine deaminase (ADA). In solution the N7H tautomer predominates. This structure however inhibits ADA, and this tautomer of formycin A would not be deactivated by the enzyme. 

Compared to baseline saquinavir pharmacokinetic parameters obtained in period 1, the use of garlic reduced the mean saquinavir area under the concentration-time curve (AUC) by 51%, and the maximum (Cmax) and minimum (Cmin) saquinavir concentrations by 54% and 49%, respectively. After a 10-day washout, the AUC, Cmax, and Cmin values were within a range of 60% to 70% of baseline values. The magnitude of the decline in concentration might result in therapeutic failure and viral rebound in patients with HIV. Based on the pharmacokinetic parameters obtained in period 3, it also appears that garlic might have a prolonged, albeit lesser, effect on saquinavir exposure. The effects of combined treatment with other protease inhibitors that are also potent cytochrome P-450 (CYP) enzymes modulators need to be further evaluated. 

The most powerful and specific inhibitors of carbonic anhydrase are certain aromatic or heterocyclic sulfonamides containing an unsubstituted R—SO2NH2 group (28). Inhibitors of this type are of great value in biochemical and physiological studies of carbonic anhydrase action and have found some therapeutic applications (28). As the various factors involved in their interaction with the enzyme have been extensively discussed at a recent symposium (51, 52) they will only be briefly mentioned here. 

Fourth, the demonstration that many of these enzymes have prognostic value in cancer may qualify them as novel therapeutic targets. For example, similar to many other proteases, these enzymes may participate in the digestion of extracellular matrix, thus facilitating tumor invasion and metastasis. The identification of highly specific inhibitors may reveal new therapeutic opportunities. 

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