Thymidylate synthase inhibition

Fig. 1.4 The increase in thymidylate synthase inhibition activity obtained from an initial hit identified as a disulphide-bound enzyme adduct and optimised to a reversible potent 330nM inhibitor using crystal structure-guided design.

Miller EM, Kinsella TJ. Radiosensitization by fluorodeoxyuridine Effects of thymidylate synthase inhibition and cell synchronization. Cancer Res 1992 52 1687-1694. 

Curtin NJ, Harris AL, Aheme GW. Mechanism of cell death following thymidylate synthase inhibition ... 

M. Perumal, R.G. Pillai, H. Barthel, J. Leyton, J.R. Latigo, M. Forster, F. Mitchell, A. L. Jackman, E.O. Aboagye, Redistribution of nucleoside transporters to the cell membrane provides a novel approach for imaging thymidylate synthase inhibition by positron emission tomography. Cancer Res. 66(17) (2006) 8558-8564. 

Gmeiner, W.H. (2005) Novel chemical strategies for thymidylate synthase inhibition. Curr. Med. Chem. 12, 191-202. 

This drug is metabolized to 5-deoxyuridine monophosphate (5-dUMP), which binds to thymidylate synthase, inhibiting the interaction between the enzyme and its major cofactor. Thus, synthesis of thymidine is inhibited, causing cell death. 

The antimetabolite, raltitrexed, is a folate analogue and is a potent and specific inhibitor of the enzyme thymidylate synthase. Inhibition of this enzyme ultimately interferes with the synthesis of deoxyribonucleic acid (DNA) leading to cell death. The intracellular polyglutamation of raltitrexed leads to the formation within cells of even more potent inhibitors of thymidylate synthase. Folate (methylene tetrahydrofolate) is a co-faetor required by thymidylate synthase and therefore theoretically folinic acid or folic acid may interfere with the aetion of raltitrexed. Clinieal interaction studies have not yet been undertaken to confirm these predieted inter-aetions. ... 

McGinn CJ, Miller EM, Lindstrom MJ, et al (1994) The role of cell cycle redistribution in radiosensitization implications regarding the mechanism of fluorodeoxyuridine radiosensitization. Int J Radiat Oncol Biol Phys 30 851-859 McGinn CJ, Shewach DS, Lawrence TS (1996) Radiosensitiz-ing nucleosides. J Natl Cancer Inst 88 1193-1203 Milas L, Hunter NR, Mason KA, et al (1994) Enhancement of tumor radioresponse of a murine mammary carcinoma by paclitaxel. Cancer Res 54 3506-3510 Milas L, Hunter NR, Mason KA, et al (1995) Role of reoxygenation in induction of enhancement of tumor radioresponse by paclitaxel. Cancer Res 55 3564-3568 Miller EM, Kinsella TJ (1992) Radiosensitization by fluorodeoxyuridine effects of thymidylate synthase inhibition and cell synchronization. Cancer Res 52 1687-1694 Miller SJ, Lavker RM, Sun TT (2005) Interpreting epithelial cancer biology in the context of stem cells tumor properties and therapeutic implications. Biochem Biophys Acta 1756 25-52... 

Marcucci G, Byrd JC, Dai G, et al (2003) Phase 1 and pharmacodynamic studies of G3139, a Bcl-2 antisense oligonucleotide, in combination with chemotherapy in refractory or relapsed acute leukemia. Blood 101 425-432 Miller EM, Kinsella TJ (1992) Radiosensitization byfluorode-oxyuridine effects of thymidylate synthase inhibition and cell synchronization. Cancer Res 52 1687-1694 Nieder C, Petersen S, Petersen C, Thames HD (2000) The challenge o p53 as prognostic and predictive factor in gliomas. Cancer Treat Rev 26 67-73... 

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. 

Proguanil appears to have a dual activity. Part of it is metabolized to cycloguanil, which subsequently inhibits the protozaon dihydrofolate reduc-tase/thymidylate synthase (DHFR/TS) (Fig. 4). In addition, the native form, proguanil itself, exerts a potent antimalarial activity, especially in combination with other antimalarial drugs. The target of proguanil is unknown. 

The methylation of deoxyuridine monophosphate (dUMP) to thymidine monophosphate (TMP), catalyzed by thymidylate synthase, is essential for the synthesis of DNA. The one-carbon fragment of methy-lene-tetrahydrofolate is reduced to a methyl group with release of dihydrofolate, which is then reduced back to tetrahydrofolate by dihydrofolate reductase. Thymidylate synthase and dihydrofolate reductase are especially active in tissues with a high rate of cell division. Methotrexate, an analog of 10-methyl-tetrahydrofolate, inhibits dihydrofolate reductase and has been exploited as an anticancer drug. The dihydrofolate reductases of some bacteria and parasites differ from the human enzyme inhibitors of these enzymes can be used as antibacterial drugs, eg, trimethoprim, and anti-malarial drugs, eg, pyrimethamine. 

Fluorouracil (5-FU) acts as a false pyrimidine, inhibiting the formation of the DNA base thymidine.26,35 The main mechanism by which it accomplishes this is by inhibiting the enzyme thymidylate synthase, the rate-limiting step in thymidine formation. 5-FU first must be metabolized to its active metabolite (F-dUMP). Additionally, metabolites of 5-FU may incorporate into RNA, inhibiting its 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.

Capecitabine -converted to 5-FU preferentially by tumor cells pyrimidine analogue antimetabolite inhibits thymidylate synthase -mucocutaneous effects (stomatitis, mucositis) -diarrhea -bone marrow suppression -nausea and vomiting -palmar-plantar erythrodysethesias (hand-foot syndrome) -fatigue... 

Drugs that can be used to control tumour cell proliferation inhibit a variety of enzymes, including thymidylate synthase and topoisomerase (Chapter 20). The enzyme aromatase converts a ring in a steroid to an aromatic ring. It converts, for example, adrenal steroid hormones into female sex hormones, which bind to oestrogenic receptors in the ovary or breast and increase the risk of ovarian or breast cancer. Aromatase inhibitors are used to treat patients with breast or ovarian cancers that are sensitive to oestrogen. Unfortunately, none of the inhibitors is specific for enzymes in tumour cells and they can therefore have severe side-effects (Chapter 21). 

We first applied Tethering to thymidylate synthase (TS). This enzyme converts de-oxyuridine monophosphate (dUMP) to thymidine monophosphate (dTMP), an activity essential for DNA synthesis. The cancer drug 5-fluorouracil irreversibly inhibits TS, and a selective inhibitor of a non-human form of the enzyme could yield a new antibiotic or antifungal drug [23]. 

Costi, M.P., Tondi, D., Rinaldi, M., Barlocco, D., Pecorari, P., Soragni, F., Venturelli, A., Stroud, R.M. Structure-based studies on species-specific inhibition of thymidylate synthase. Biochim. Biophys. Acta 2002, 1587, 206-214. 

Berger SH, Hakala MT. Relationship of dUMP and free FdUMP pools to inhibition of thymidylate synthase by 5-fluorouracil. Mol Pharmacol 1984 25 303-309. 

Administration by experienced physician only pts should be hospitalized for 1st course d/t risk for severe Rxn Uses G1 adenoma, liver, renal cancers colon pancreatic CAs Action Converted to 5-FU inhibits thymidylate synthase DNA synth (S-phase specific) Dose 0.1-0.6 mg/kg/d for 1-6 wk (per protocols) usually intra-arterial for liver mets Caution [D, -] Interaction w/ vaccines Contra BM... 

Although the anti-tumour activity of 5-FU is mainly because of the inhibition of the thymidylate synthase, it is actually multifactor. The incorporation of 5-FU, under F-desoxyuridine form, into various types of RNA generates cytotoxicity. This occurs through the inhibition of the methylation and of the maturation of the ribosomal RNA [86]. The 5-FU is also precursor, by metabolism, of a monofluoroalanine, which is itself toxic. 

Thymidylate synthase (TS) is the enzyme that converts 2-deoxyuridine monophosphate into thymidine monophosphate. This is a key step in the biosynthesis of DNA. This enzymatic reaction of methylation involves the formation of a ternary complex between the substrate, the enzyme, and tetrahydrofolic acid (CH2FAH4). The catalytic cycle involves the dissociation of this complex and the elimination of FAH4. It is initiated by pulling out the proton H-5, thus generating an exocyclic methylene compound. As the release of a F" " ion is energetically forbidden, the fluorine atom that replaces the proton H-5 cannot be pulled out by the base. This leads to inhibition of the enzyme (Figure 7.2). 

Although the antitumor activity of 5 -fluorouracil (5 -FU) is mainly due to the inhibition of thymidylate synthase (after its in vivo glycosylation and its phosphorylation), it... 

Trifluridine, a trifluoromethylthymidine, inhibits the thymidylate synthase (TS) with a mechanism different from that of 5-FU. It is a mechanism-based inhibitor (see Figure 7.52). 

Trifluridine (5-trifluoromethyl-2 -deoxyuridine) (Viroptic ) is marketed for the topical treatment of herpes simplex virus infection in the eyes. This antiviral drug is a mechanism-based inactivator of thymidylate synthase. The mechanism of inhibition and synthesis of trifluoridine are reported in Chapter 7. 

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