Folate inhibition

In view of the well-documented inhibition of dihydrofolate reductase by aminopterin (325), methotrexate (326) and related compounds it is generally accepted that this inhibitory effect constitutes the primary metabolic action of folate analogues and results in a block in the conversion of folate and dihydrofolate (DHF) to THF and its derivatives. As a consequence of this block, tissues become deficient in the THF derivatives, and this deficiency has many consequences similar to those resulting from nutritional folate deficiency. The crucial effect, however, is a depression of thymidylate synthesis with a consequent failure in DNA synthesis and arrest of cell division that has lethal results in rapidly proliferating tissues such as intestinal mucosa and bone marrow (B-69MI21604, B-69MI21605). 

Methotrexate (MTX, chemical structure shown in Fig. 1.) competitively inhibits the dehyrofolate reductase, an enzyme that plays an essential role in purine synthesis. The dehydrofolate reductase regenerates reduced folates when thymidine monophosphate is formed from deoxyuridine monophosphate. Without reduced folates cells are unable to synthesize thymine. Administration of N-5 tetrahydrofolate or N-5 formyl-tetrahydrofolate (folinic acid) can bypass this block and rescue cells from methotrexate activity by serving as antidote. 

Ralitrexed is a folate analog with greater selectivity. It easily crosses the cell membrane and undergoes polyglutamation. Within tissues, ralitrexed may be stored up to 29 days. It directly inhibits thymidylate synthase, the key enzyme for synthesizing thymidine triphosphate (TTP). The drug has been described to induce apoptosis in tumor cells. Ralitrexed is used for the treatment of colon carcinomas. 

Fluorouracil (5-fluorouracil, 5-FU, Fig. 5) represents an early example of rational drag design in that it originated from the observation that tumor cells, especially from gut, incorporate radiolabeled uracil more efficiently into DNA than normal cells. 5-FU is a fluorinated pyrimidine analog that must be activated metabolically. In the cells 5-FU is converted to 5-fluoro-2>deoxyuridine-monophosphate (FdUMP). This metabolite inhibits thymidilate synthase which catalyses the conversion of uridylate (dUMP) to thymidilate (dTMP) whereby methylenetetrahydrofo-late plays the role of the carbon-donating cofactor. The reduced folate cofactor occupies an allosteric site of... 

The folate antagonists, pyrimethamine and sulfadiazine, inhibit the parasite s DHFR/TS synthase enzyme complex and the DHPS, respectively (Fig. 4) (see antimalarial drugs). To avoid deficiency of folic acid in patients treated with antifolate antagonists, folinic acid supplementation is recommended to reduce bone-marrow suppression. 

Several reactions of IMP biosynthesis require folate derivatives and glutamine. Consequently, antifolate drugs and glutamine analogs inhibit purine biosynthesis. 

Figure 45-14. Homocysteinuria and the folate trap. Vitamin 6,2 deficiency leads to inhibition of methionine synthase activity causing homocysteinuria and the trapping of folate as methyltetrahydrofolate.

Folate metabolism Sulphonamides (also ) Trimethoprim Pyrimethamine Trimetrexate / Inhibit folate synthesis Inhibits dihydrofolate reductase Inhibits dihydrofolate reductase Inhibits dihydrofolate reductase Not present in mammalian cells Mammalian enzyme not inhibited Mammalian enzyme not inhibited Toxicity overcome with leucovorin... 

Sulphonamides are structural analogues of PABA. They competitively inhibit the incorporation of PABA into dihydropteroic acid and there is some evidence for their incorporation into false folate analogues which inhibit subsequent metabolism. The presence of excess PABA will reverse the inhibitory action of sulphonamides, as will thymine, adenine, guanine and methionine. However, these nutrients are not normally available at the site of infections for which the sulphonamides are used. 

Folic acid antagonist inhibits dihydrofolate reductase (DHFR) blocks reduction of folate to tetrahydrofolate inhibits de novo purine synthesis results in arrest of DNA, RNA, and protein synthesis... 

TK), 5-FU is activated to 5-fluorodeoxyuridine monophosphate (5-FdUMP). Potent inhibition of thymidylate synthase (TS) by 5-FdUMP is considered critical for 5-FU cytotoxicity. TS catalyzes the rate-limiting step of DNA synthesis, such as the conversion of dUMP into dTMP. Optimal TS function requires the formation of a covalent ternary complex consisting of TS, the folate cofactor 5,10-methylenetetra-hydrofolate (CH2THF), and 5-FdUMP. Inadequate cellular levels of 5,10-methyle-netetrahydrofolate reduce the stability of the ternary complex and consequently the inhibition of TS by 5-FdUMP. For this reason, 5-FU is administered in association with folinic acid, a precursor of 5,10-methylenetetrahydrofolate [40]. 

Fig. 14.10 Folate metabolism and role of MTHFR. Genetically reduced MTHFR activity affects the distribution between folate species required for protein and DNA synthesis. Higher availabil ity of 5,10-methylenetetrahydrofolate (CH2THF) potentiates the TS inhibition by 5-FdUMP, the active metabolite of 5-FU. Hey, homocysteine Met, methionine CH3HF, 5-methyltetrahydrofolate TS, thymidylate synthase 5-FdUMP, fluorodeoxyuridine monophosphate.

It is the role of jV5-methyl THF which is key to understanding the involvement of cobalamin in megaloblastic anaemia. The metabolic requirement for N-methyl THF is to maintain a supply of the amino acid methionine, the precursor of S-adenosyl methionine (SAM), which is required for a number of methylation reactions. The transfer of the methyl group from jV5-methyl THF to homocysteine is cobalamin-dependent, so in B12 deficiency states, the production of SAM is reduced. Furthermore, the reaction which brings about the formation of Ns-methyl THF from N5,N10-methylene THF is irreversible and controlled by feedback inhibition by SAM. Thus, if B12 is unavailable, SAM concentration falls and Ah -methyl THF accumulates and THF cannot be re-formed. The accumulation of AT-methyl THF is sometimes referred to as the methyl trap because a functional deficiency of folate is created. 

W. J. Chang, K. G. Rothberg, B. A. Kamen, and R. G. Anderson. Lowering the cholesterol content of MA104 cells inhibits receptor-mediated transport of folate. J. Cell Biol. 118 63-69 (1992). 

Methotrexate acid produced Antineoplastic (Cancer) Inhibits dihydrofolate reductase, enzyme that converts folate... 

Inhibit Enzymes Many drugs are competitive inhibitors of key enzymes in pathways. The statin drugs (lovastatin, simvastatin), used to control blood cholesterol levels, competitively inhibit 3-hvdroxy-3-methylglutaryl coenzyme A (HMG CoA) reductase in cholesterol biosynthesis. Methotrexate, an antineoplastic drug, competitively inhibits dihydrofolate reductase, depriving the cell of active folate needed for purine and deoxythymidine synthesis, thus interfering with DNA replication during S phase. 

Megaloblastic anemia results from insufficient active THF to support cell division in the bone marrow. Methotrexate inhibits DHF reductase, making it a useful antineoplastic drug. Folate deficiencies may be seen during pregnancy and in alcoholism. 

Gene polymorphisms in MTX transporters and enzymes in the folate and adenosine pathways inhibited by MTX have been studied in RA patients. 

---