SPICE2 network thermodynamic simulation of antifolate effects on purine and pyrimidine biosynthesis: Exploring the role of tetrahydrofolate cofactor depletion versus dihydrofolate feedback inhibition

Utilizing the network thermodynamic approach, it is possible to simulate large, nonlinear dynamic systems on SPICE2. For this purpose SPICE2 can become a general systems simulator in spite of the fact that it was originally designed for simulating integrated electrical circuits. As an example of the utility of SPICE2 for large metabolic systems which involve nonlinear kinetics, feedback inhibition of a number of kinds and branching pathways, the cellular folate metabolism scheme is modeled. This scheme is a source of intermediates for de novo purine and pyrimidine metabolism utilized in DNA and RNA synthesis. The action of antifolates that specifically inhibit the enzyme dihydrofolate reductase (DHFR), is simulated in some detail. In particular, the potential of feedback inhibition of secondary sites by dihydrofolate (DHF) polyglutamates that accumulate in the presence of antifolates is modeled to investigate the possible role of such inhibition as opposed, or in addition to, the classical role of tetrahydrofolate (THF) cofactor depletion in the suppression of purine and pyrimidine biosynthesis after blockage of DHFR. The interplay of mathematical modeling, computer simulation, network thermodynamics and experimental observations gave rise to the following conclusions: (1) THF cofactor depletion alone can account for the rapid cessation of purine and pyrimidine synthesis in S-phase cells by antifolates; (2) feedback inhibition by DHF polyglutamates need not be implemented in the mechanism of antifolate inhibition; and (3) the lack of complete interconversion of THF cofactors to DHF does not appear to be related to inhibition of thymidylate synthase by DHF polyglutamates, since this would only slow the accumulation of but not change the final DHF and THF levels in cells with antifolates.