This represents rather a large chapter 287 and is based on the concept that agents produce biological effects by interfering with the action of naturally occurring, physiologically active compounds or with their enzymic transformation, or by blocking the pharmacodynamic action of drugs. It comprises, apart from enzymic events (cf. reference 248), pharmacodynamic (anti-acetylcholine, anti-epinephrine, antihistamine compounds, etc.), hormonal (anti-hormones) and immunological aspects. Since the first clinical demonstration 88 of the effectiveness of folic acid analogues, the study and use of 'antimetabolites' in the cancer field has increased enormously and every day new representatives of this class of therapeutics are disclosed, as reviewed from a clinical angle by Farber et al.,89 from a chemical point of view by Montgomery 183 and Timmis,236 and speculated upon by Baker.20 It can be assumed that most of these agents achieve their chemotherapeutic effects by interfering in one way or another (often overlapping) with the enzymic processes of the cell. In view of this and of Baker's argument with regard to irreversible inhibitors,20 it is desirable to look in a general, but brief manner into the subject of enzyme antagonists although there is an excellent treatment of this topic in Dixon and Webb's book.83 One could start with an oversimplified scheme where E is the enzyme, S the substrate, P the product, and KM (Michaelis constant) expresses the substrate concentration at half-maximal velocity (V) and K8 the dissociation constant of ES, the enzyme-substrate complex. The latter can be formed in absence or presence of a coenzyme and/or metal activators dependent on E in question. Prevention of the formation of P is due to inhibitors (I) (see Fig. 17) acting with E or ES reversibly or irreversibly. In the case of the latter type of antagonism, the agent progressively inhibits the catalytic process which, after a time, is completely arrested. This is very often achieved by chemical interaction between I and E with the formation of co-valent bonds and might involve the catalytic site of E or an adjacent site. A similar but less permanent interaction occurs with reversible inhibitors, leading to an equilibrium between E and I (Kt being the reciprocal affinity constant of I). But if such an inhibitor reacts with the catalytic site it is called a 'competitive' inhibitor, producing an increase of KM, and if combining with an adjacent site, 'non-competitive,' reducing V of the enzyme reaction. As it can be seen further on, the inhibitor may also compete or interfere with the coenzyme or the metal activators, if they are present in and necessary for a specific enzyme system (see Fig. 17).

In consequence, antagonism can be produced by substrate-like, product or product-like, substrate-and product-unlike and coenzyme- or coenzyme precursor-like compounds. In addition removal or changes of metal-activators can cause inactivity. A classical example of a substrate-like inhibitor is that of malonic acid which interferes competitively with oxaloacetic acid decarboxylase and succinic dehydrogenase;205 another is that of eserine or neostigmine, blocking cholinesterases. Of greater interest for cancer research are the numerous pyrimidine and purine analogues72 which were discussed recently by Montgomery188 (see also references 89, 236). Most of them act in the cancer cell in form of nucleotides. Quoting one example, namely that of 5-fluorouracil,1291227 it competes after transformation into the deoxyribotide intracellularly with deoxyuridylic acid for the enzyme system which normally methylates the natural intermediate to thymidylic acid. In this manner, the fluoro-compound blocks this reaction and hence interferes with DNA synthesis. At the same time like other 'fraudulent' or 'rogue' NA-bases, it is incorporated into oligo- and poly-nucleo-tides and as such may exert a product-like antagonism by accumulation and hindering a related normal EP-complex (see Scheme) from splitting into E and P. The old example of fructose, as a product of invertase action on saccharose, inhibiting further formation of itself, illustrates this even more clearly. As mentioned before an increasing amount of product, either left intracellularly unchanged because of a deficiency of its degrading enzyme or introduced into the biological system from outside, can inhibit the activity of the enzyme necessary for its own synthesis and thus cause a process of negative feedback. This might lead in the end to suppression or repression of one or more EFS which is of course the opposite to induction. At the same time this interdependence of a whole series of enzymic events, physico-chemically governed by the exponential law of autocatalysis, as described by Hinshelwood,131 is probably responsible for the fact that a 'biochemical lesion' in the cell, involving originally only one event, will change or block a whole pathway or even a number of metabolic chains, bringing about the death of the cell or of the total organism.

Fig. 17. Mechanism of action of enzyme inhibitors and antagonists (--- interaction with sites,........interaction with type of antagonist).

Enzyme antagonists, however, could be unlike substrates or products if they are capable of interacting either with the catalytic or adjacent site or with any parts essential for enzymic activity. Reversible inhibitors in this group are, for instance, aromatic acids, such as salicylic acid, antagonizing and stabilizing bovine xanthine oxidase 27,28 and m-toluic acid 245 doing the same with d-amino-acid oxidase. Organic phosphates block cholinesterases, chymotrypsin, etc. irreversibly by phosphorylation of one amino-acid of the active center,138 although such inhibition can be overcome by the use of compounds such as pyridine aldoxime methiodide.184 Vulnerable points of many enzymes are thiol groups (-SH) which can be blunted by heavy metals (silver, mercury or lead), organo-metal compounds (p-chloromercuribenzoate), arsenicals and a number of other substances (quinones, halogeno-acids and ketones, etc.). The toxic effects of some of them can be counteracted by BAL (2, 3-dimercaptopropanol) .233 While certain alkylating agents, in addition to their effects on nucleotides, may interact with functional proteins, it has been demonstrated by Roberts and Warwick 218 that one of the principal actions of myleran (1,4-dimethanesul-phonyloxybutane) represents very likely a process of de-thiolation of sulfhydryl-containing key substances, such as glutathione, etc. with the in vivo production of 3-hydroxy-tetrahydrothiophene sul-phone. This may also occur with enzymes and could be the mechanism of the anti-leukemic activity of the drug. Another group of potential antagonists may be that of enzyme antibodies. These, according to the review by Cinader,78 are effective in vitro inhibitors of enzymes only if their interaction with the enzymic antigen involves either the catalytic site or a site nearby which, when blocked by bulky antibodies, could sterically hinder the catalytic process. Other enzyme-antibody complexes show a normal or even a higher enzymic activity. Nevertheless, enzymes, such as pyruvic acid carboxylase, tryptophanase, hexokinase, glyceraldehyde 3-phosphate dehydrogenase, bacterial proteinase, ribonuclease, hyaluronidase, etc. have been inhibited up to 100% by the corresponding antiserum under certain conditions. It is interesting to note that cross-reactions have never been observed with functionally similar enzymes from different species but did occur with enzymes from closely related organs, this being dependent solely on taxonomic factors but not on catalytic properties. Such circumstances might diminish considerably the usefulness of some of these anti-enzymes for neutralizing those enzymes which reside in tumor cells at a higher level than in normal cells. However, a limited but systematic study of this immunological group of inhibitors under in vivo conditions would be a not unprofitable undertaking.