Action of aminopterin and amethopterin methotrexate

On addition to cells these inhibitors rapidly bring about inhibition of DNA synthesis. Incorporation of [3H]deoxyuridine into DNA is reduced to 3% of control values within 15 min of addition of 10/xM aminopterin and even O.ljuM reduces incorporation to less than 20% of controls though this takes 45 min (Siegers et a]., 1975). Although concentrations of amethopterin between 0.1 and lfiM block synthesis of thymidylate, DNA synthesis continues at a significant rate and pools of thymidylate are slow to dissipate (Adams et al., 1971 Baumunk and Friedman, 1971 Fridland, 1974 Fridland and Brent, 1975). 

Aminopterin and amethopterin are 4-amino analogues of folic acid (Fig. 11.5) and as such are potent inhibitors of the enzyme dihydrofolate reductase (EC 1.5.1.3) (Blakley, 1969). This enzyme catalyses the reduction of folic acid and dihydrofolic acid to tetrahy-drofolic acid which is the level of reduction of the active coenzyme involved in many different aspects of single carbon transfer. As is clear from Fig. 11.6, tetrahydrofolate is involved in the metabolism of (a) the amino acids glycine and methionine (b) the carbon atoms at positions 2 and 8 of the purine ring (c) the methyl group of thymidine and (d) indirectly in the synthesis of choline and histidine. 

The inhibitory action of the antifolates may be reversed either by changing the medium (Adams, 1969a, b), or more conveniently by additions of thymidine to the medium when dTTP is synthesised using the enzymes thymidine and thymidylate kinase (see Fig. 11.7). 

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