Thymidine 5 - , enzymic synthesis

The more-common 2-acetamido-2-deoxyhexoses have not been found as thymidine 5 -pyrophosphate derivatives. Nevertheless, the enzymic synthesis of thymidine 5 -(2-acetamido-2-deoxy-a-D-gluco-pyranosyl pyrophosphate) and -(2-acetamido-2-deoxy-a-D-galactopy-ranosyl pyrophosphate) has been achieved with enzyme preparations from Pseudomonas aeruginosa,116 Azotobacter vinelandii,52 and gastric mucosa.117... 

MacNutt70 achieved an enzymic synthesis of thymidine and deoxyuri-... 

Methotrexate (e.g., Mexate) Inhibits dihydrofolate reductase, an enzyme that catalyzes the synthesis of thymidine. DNA synthesis is thus impaired. Prevention of organ transplant rejection, severe psoriasis. Bone marrow depression, Gl ulceration, nephrotoxicity, hepatotoxicity, pulmonary infiltration, skin toxicity. 

An enzymic synthesis of cDNA has been carried out on thymidine polynucleotide-cellulose, which is formed when cellulose reacts with thymidine 5 -phosphate in the presence of a carbodi-imide. The product DNA was used in assays of mRNA. 

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. 

Methotrexate belongs to the class of antimetabolites. As a derivative of folic acid it inhibits the enzyme dihydrofolate reductase resulting in a decreased production of thymidine and purine bases essential for RNA and DNA synthesis. This interruption of the cellular metabolism and mitosis leads to cell death. 

Deoxy-5 -fluorothymidine (838) was prepared by Langen and Kowol-jj. 796,797 fpQjyj 5 -0-tosyl precursor by treatment with fluoride. Compound 838 cannot be phosphorylated enzymically owing to the lack of OH-5, but it inhibits the growth of carcinoma cells. This was explained as follows the thymidine 5 -monophosphate (thymidylate) kinase in carcinoma cells, catalyzing the transformation of thymidine 5 -monophosphate into the diphosphate, is inhibited by 838, thus preventing the synthesis of... 

While mammahan cells reutilize few free pyrimidines, salvage reactions convert the ribonucleosides uridine and cytidine and the deoxyribonucleosides thymidine and deoxycytidine to their respective nucleotides. ATP-dependent phosphoryltransferases (kinases) catalyze the phosphorylation of the nucleoside diphosphates 2 "-de-oxycytidine, 2 -deoxyguanosine, and 2 -deoxyadenosine to their corresponding nucleoside triphosphates. In addition, orotate phosphoribosyltransferase (reaction 5, Figure 34-7), an enzyme of pyrimidine nucleotide synthesis, salvages orotic acid by converting it to orotidine monophosphate (OMP). 

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. 

DHFR has been the object of intense research for the last few decades. The enzyme catalyses the NADPH-dependent reduction of 7,8-dihydrofolate to 5,6,7,8 tetrahydrofolate, a chemical which participates in the thymidilate synthesis cycle. Thus, the enzyme is crucial in the synthesis of thymidine monophosphate as well as in various one-carbon unit transfer reactions. 

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]. 

Inhibition of nucleobase synthesis (2). Tetrahydrofolic acid (THF) is required for the synthesis of both purine bases and thymidine. Formation of THF from folic acid involves dihydrofolate reductase (p. 272). The folate analogues aminopterin and methotrexate (ame-thopterin) inhibit enzyme activity as false substrates. As cellular stores of THF are depleted, synthesis of DNA and RNA building blocks ceases. The effect of these antimetabolites can be reversed Ltillmann, Color Atlas of Pharmacology 2000 Thieme All rights reserved. Usage subject to terms and conditions of iicense. 

Both sulfonamides and trimethoprim (not a sulfonamide) sequentially interfere with folic acid synthesis by bacteria. Folic acid functions as a coenzyme in the transfer of one-carbon units required for the synthesis of thymidine, purines, and some amino acids and consists of three components a pteridine moiety, PABA, and glutamate (Fig. 44.1). The sulfonamides, as structural analogues, competitively block PABA incorporation sulfonamides inhibit the enzyme dihydropteroate synthase, which is necessary for PABA to be incorporated into dihydropteroic acid, an intermediate compound in the formation of folinic acid. Since the sulfonamides reversibly block the synthesis of folic acid, they are bacteriostatic drugs. Humans cannot synthesize folic acid and must acquire it in the diet thus, the sulfonamides selectively inhibit microbial growth. 

F]-FLT is not or only marginally incorporated into DNA (<2%) and therefore not a direct measure of proliferation [122]. In vitro studies indicated that [ F]-FLT uptake is closely related to thymidine kinase 1 (TK1) activity and respective protein levels [117,118]. p F]-FLT is therefore considered to reflect TK1 activity and hence, S-phase fraction rather than DNA synthesis. Although being a poor substrate for type 1 equilibrative nucleoside transporters (ENT), cellular uptake of [ F]-FLT is further facilitated by redistribution of nucleoside transporters to the cellular membrane after inhibition of endogenous synthesis of thymidylate (TMP) de novo synthesis of TMP) [125]. However, the detailed uptake mechanism of [ F]-FLT is yet unknown and the influence of membrane transporters and various nucleoside metabolizing enzymes remains to be determined. 

Acyclovir requires three phosphorylation steps for activation. It is converted first to the monophosphate derivative by the virus-specified thymidine kinase and then to the di- and triphosphate compounds by host cell enzymes (Figure 49-3). Because it requires the viral kinase for initial phosphorylation, acyclovir is selectively activated—and the active metabolite accumulates—only in infected cells. Acyclovir triphosphate inhibits viral DNA synthesis by two mechanisms competition with deoxyGTP for the viral... 

Trifluridine (trifluorothymidine) is a fluorinated pyrimidine nucleoside that inhibits viral DNA synthesis in HSV-1, HSV-2, CMV, vaccinia, and some adenoviruses. It is phosphorylated intracellularly by host cell enzymes, and then competes with thymidine triphosphate for incorporation by the viral DNA polymerase (Figure 49-3). Incorporation of trifluridine triphosphate into both viral and host DNA prevents its systemic use. Application of a 1% solution is effective in treating keratoconjunctivitis and recurrent epithelial keratitis due to HSV-1 or HSV-2. Cutaneous application of trifluridine solution, alone or in combination with interferon alfo, has been used successfully in the treatment of acyclovir-resistant HSV infections. 

Several cases of synthesis of a-D-galactosyl nucleotides from a-D-galactopyranosyl phosphate with bacterial enzymes have been reported. These included formation of UDP-Gal through reaction with uridine 5 -triphosphate22 or UDP-Glc,14,44 49 and of dTDP-Gal through interaction with thymidine 5 -triphosphate.50... 

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