Thymidine phosphorylase

BVdU is degraded by thymidine phosphorylase more rapidly than the natural substrate, thymidine. This rapid enzymic degradation may present a problem in its clinical use. Moreover, herpes vimses develop resistance to BVdU, apparendy because of mutant vimses that have lower thymidine kinase activity. G. D. Seade has dropped further development of BVdU because of increased animal tumor incidence induced by prolonged dosing (1). 

After intravenous administration, about 80-90% of the dose is catabolized in the liver by dihydropyrimidine dehydrogenase (DPD) [38] (Figure 14.3). The formation of the inactive 5-fluoro-5,6-dihydrouracil (5-FUH2) by DPD is the rate-limiting step of 5-FU catabolism [39]. DPD is widely distributed among tissues, with the highest levels found in the liver. Once 5-FU entered tumor cells, its antitumor effect is mainly dependent on the extent of 5-FU anabolism. After two sequential anabolic steps involving thymidine phosphorylase (TP) and thymidine kinase... 

Fig. 14.3 5 -FU catabolism, anabolism and mechanism of action. 5-FUH2, 5-fluoro-5,6-dihydrouracil 5-FdUMP, 5-fluorodeoxyuridine monophosphate TP, thymidine phosphorylase TK, thymidine kinase TS, thymidylate synthase CH2THF, 5,10-methylenetetrahydrofolate.

Like many reported A -transglycosylations, this reaction uses uncharacterized nucleoside phosphorylases from whole cells held at 50-60 °C, a temperature well above the range for viability of the parent microorganism. Remarkable temperature stability has been reported for three well-known NPs of E. coli purine nucleoside phosphorylase (PNP), uridine phosphorylase (URDP) and thymidine phosphorylase. ... 

Salonga, D., Danenberg, K. D., Johnson, M., et al. (2000) Colorectal tumors responding to 5-fluorouracil have low gene expression levels of drhydropyrimidine dehydrogenase, thymi-dylate synthase, and thymidine phosphorylase. Clin. Cancer Res. 6, 1322-1327. 

Y.L. Chung, H. Troy, I.R. Judson, R. Leek, M.O. Leach, M. Stubbs, A.L. Harris, J.R. Griffiths, Noninvasive measurements of capecitabine metabolism in bladder tumors overexpressing thymidine phosphorylase by fluorine-19 magnetic resonance spectroscopy, Clin. Cancer Res. 10 (2004) 3863-3870. 

A number of fluoropyrimidines other than 5-FU have been s mthesized, most of which act as prodrugs for 5-FU. Capecitabine (Xeloda ) is an orally administered fluorop5Tim-idine that is converted to 5-FU by the enzyme thymidine phosphorylase (TP), which is often over-expressed in malignant compared to normal tissues (29). Capecitabine has been shown to be equivalent to 5-FU/LV and was the first oral agent to be approved by the FDA for the treatment of mCRC (30). 

Abbreviations TS (thymidylate synthase) DPD (dihydropyrimidine dehydrogenase) TP (thymidine phosphorylase) RR (response rate) OS (overall survival) mCRC (metastatie coloreetal eaneer). 

Fig. lA. Anabolic and catabolic pathways of 5-FU. DPD dihydropyrimidine dehydrogenase, DP di-hydropyrimidinase, pUP beta-ureidopropionase, UP uridine phosphorylase, OPRT orotate phospho-ribosyl transferase, UK uridine kinase, TP thymidine phosphorylase, TK thymidine kinase, RNR ribonucleotide reductase. The three active metabolites (shown in rectangles) are FdUMP (5-fluoro-2 -deoxyuridine 5 -monophosphate) inhibiting TS (thymidylate synthase), and FUTP (5-fluorouridine 5 -triphosphate) and FdUTP (5-fluoro 2 -deoxyuridine 5 -triphosphate) interfering with RNA and DNA, respectively. 

Fig. IB. Metabolic pathway of the oral pro-drug capecitabine. The dmg has been designed to generate specifically 5-FU in tumor cells over-expressing thymidine phosphorylase.

Ciccolini J, Evrard A, Cuq P. Thymidine phosphorylase and fluoropyrimidines efficacy a Jekyll and Hyde story. Curr Med Chem Anticancer Agents 2004 4. 71-S1. 

Ciaparrone M, Quirino M, Schinzari G et al. Predictive role of thymidylate synthase, dihydropyrimidine dehydrogenase and thymidine phosphorylase expression in colorectal cancer patients receiving adjuvant 5-fluorouraeil. Oncology 2006 70 366-377. 

Ishikawa T, Sekiguchi F, Fukase Y et al. Positive correlation between the efficacy of capecitabine and doxifluridine and the ratio of thymidine phosphorylase to dihydropyrimidine dehydrogenase activities in tumors in human cancer xenografts. Cancer Res 1998 58 685-690. 

Capecitabine is a fluoropyrimidine carbamate prodrug with 70-80% oral bioavailability. It undergoes extensive metabolism in the liver by the enzyme carboxylesterase to an intermediate, 5 -deoxy-5-fluorocytidine. This is converted to 5 -deoxy-5-fluorouridine by the enzyme cytidine deaminase. These two initial steps occur mainly in the liver. The 5 -deoxy-5-fluorouridine metabolite is then hydrolyzed by thymidine phosphorylase to 5-FU directly in the tumor. The expression of thymidine phosphorylase has been shown to be significantly higher in a broad range of solid tumors than in corresponding normal tissue, particularly in breast cancer and colorectal cancer. 

MNGIE [21] Intermittent diarrhea and intestinal pseudo-obstruction (myoneurogastrointestinal encephalopathy). Mutations in the thymidine-phosphorylase-encoding gene leads to multiple mtDNA deletion and anomalies in mtDNA synthesis. 

Table 7.1.4 Concentration range of purine and pyrimidine metabolites in urine (pmol/mmol creatinine) from patients. ADA Adenosine deaminase, APRT adenine phosphoribosyltransferase, ASA adenylosuccinate lyase, DHP dihydropyrimidinase, DPD dihydropyrimidine dehydrogenase, HGPRT hypoxanthine-guanine phosphoribosyltransferase, PNP purine nucleoside phosphorylase, TP thymidine phosphorylase, UMPS uridine monophosphate synthase, / -UP fi-ureidopropionase... 

The activity thymidine phosphorylase can be detected in leukocytes using a non-radiochemical assay in which thymine is detected at 265 nm after separation with reverse-phase HPLC [9]. 

Fairbanks LD, Marinaki AM, Carrey EA, Hammans SR, Duley JA (2002) Deoxyuridine accumulation in urine in thymidine phosphorylase deficiency (MNGIE) J Inherit Metab Dis 25 603-604... 

Van Kuilenburg ABP, Zoetekouw L (2005) Determination of thymidine phosphorylase activity by a non-radiochemical assay using reversed-phase high-performance liquid chromatography. J Chromatogr Biomed Sci Appl 820 271-275... 

Just as orotic acid is converted to a ribonucleotide in step e of Fig. 25-14, other free pyrimidine and purine bases can react with PRPP to give monoribonucleotides plus PP . The reversible reactions, which are catalyzed by phosphoribosyltransferases (ribonucleotide pyrophosphorylases), are important components of the salvage pathways by which purine and pyrimidine bases freed by the degradation of nucleic acids are recycled.273 However, thymine is usually not reused. Thymine will react with deoxribose 1-P to form thymidine plus inorganic phosphate (thymidine phosphorylase), and thymidine is rapidly... 

Kenny (1973) recommends a Soy peptone-yeast dialysate medium (Appendix 4) for culture of mycoplasma. Inclusion of arginine (16 mM) and 0.4 mg% phenol red indicates the presence of arginine deaminase by formation of alkali (purple coloration). Alternatively, incubation with tritiated thymidine and analysis of the culture medium for tritiated thymine can be used to detect thymidine phosphorylase (House and Waddell, 1967). 

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