Purine oxidative metabolism

Xanthine oxidoreductase (XOR) is a molybdenum-containing complex homodimeric 300-kDa cytosolic enzyme. Each subunit contains a molybdopterin cofactor, two nonidentical iron-sulfur centers, and FAD (89). The enzyme has an important physiologic role in the oxidative metabolism of purines, e.g., it catalyzes the sequence of reactions that convert hypoxanthine to xanthine then to uric acid (Fig. 4.36). 

Mercaptopurine and thioguanine are both given orally (Table 55-3) and excreted mainly in the urine. However, 6-MP is converted to an inactive metabolite (6-thiouric acid) by an oxidation catalyzed by xanthine oxidase, whereas 6-TG requires deamination before it is metabolized by this enzyme. This factor is important because the purine analog allopurinol, a potent xanthine oxidase inhibitor, is frequently used with chemotherapy in hematologic cancers to prevent hyperuricemia after tumor cell lysis. It does this by blocking purine oxidation, allowing excretion of cellular purines that are relatively more soluble than uric acid. Nephrotoxicity and acute gout produced by excessive uric acid are thereby prevented. Simultaneous therapy with allopurinol and 6-MP results in excessive toxicity unless the dose of mercaptopurine is reduced to 25% of the usual level. This effect does not occur with 6-TG, which can be used in full doses with allopurinol. 

Fig. 3.2 Classification of "metabolism ESTs. Tentative unique genes (TUGs) from each species that clustered into the metabolism category (see Fig. 3.1J were further subdivided. Here, carbohydrate incorporates all glycoside hydrolases, lipid includes lipases, B-oxidation and steroid metabolizing enzymes, protein includes predominantly proteases and their precursors, and amino acid includes all enzymes involved in the interconversion of amino acids. Esterase includes those esterase-like enzymes with unknown substrates, and other contains those TUGs that do not sort into other categories. These include predominantly oxidoreductases and purine/pyrimidine metabolizing enzymes.

In addition to metabolizing some aldehydes, aldehyde oxidase also oxidizes a variety of azaheterocycles but not thia- or oxaheterocycles. Of the various purine nucleosides metabolized by aldehyde oxidase, the 2-hydroxy- and 2-amino derivatives are more efficiently metabolized, and for the N -substituents, the typical order of preference is the acyclic nucleosides is as follows 9-[(hydroxy-alkyloxy)methyl]-purines) > 2 -deoxyribofuranosyl > ribofuranosyl > arabinofuranosyl > H. The kinetic rate constants for purine analogues revealed that the pyrimidine portion of the purine ring system is more important for substrate affinity than the imidazole portion. Aldehyde oxidase is inhibited by potassium cyanide and menadione (synthetic vitamin K). 

In vivo and in vitro studies have shown that oxidations play a major role in purine drug metabolism. Electrochemical oxidation pathways of purine drugs were, therefore, evaluated. 

Fig. 7. Model for the subcellular localization of reactions of purine synthesis and ureide biogenesis in nodules of ureide-exportlng legumes. The model is based on results of subcellular fractionation and ultrastructural studies. The processes (shown in the hatched boxes) involved in ureide biogenesis (i.e., nitrogen fixation, ammonium assimilation, precursor synthesis, purine synthesis, energy-yielding metabolism, and purine oxidation and catabolism) may occur in more than one subcellular compartment. The location of the enzymes involved in the conversion of IMP to xanthine is not certain. We have proposed that in soybean nodules these reactions [shown in bold-face type with bold arrows] occur in the plastid while in other species such as cowpea these reactions may take place in the ground cytoplasm. In all cases the intermediate exported from the plastid is uncertain. This uncertainty is indicated with the dashed lines and question marks.

The mechanism for the production of O2" in ischaemic tissue appears to involve changes in purine metabolism within ischaemic cells. Sublethal hypoxia decelerates mitochondrial oxidative phosphorylation, rendering the production of ATP dependent upon the... 

Adenine phosphoribosyltransferase (APRT) deficiency is an inherited disorder of purine metabolism and is inherited in an autosomal recessive manner (K18, V7). This enzyme deficiency results in an inability to salvage the purine base adenine, which is oxidized via the 8-hydroxy intermediate by xanthine oxidase to 2,8-di-hydroxyadenine (2,8-DHA). This produces crystalluria and the possible formation of kidney stones due to the excretion of excessive amounts of this insoluble purine. Type I, with virtually undetectable enzyme activity, found predominantly in Caucasians, is found in homozygotes or compound heterozygotes for null alleles. Type II, with significant APRT activity, found only in Japan, is related to a missense mu-... 

Uric acid is one of the principal products of purine metabolism in man 12 13). However, in many other organisms further oxidative degradation of the purine molecule occurs. One of the most important enzymes involved in uric acid oxidation is uricase, which has been studied to some extent in vitro. 

Uric acid is the end-product of purine metabolism in humans, other primates, birds and reptiles. It is produced in the liver by the oxidation of xanthine and hypoxanthine (Figure 12.16),... 

Purine metabolism in some mammals is characterized by a further oxidation of uric acid to al-lantoin by the enzyme urate oxidase. Allantoin is significantly more water soluble than uric acid and is also freely excreted via the renal route. 

AO is also effective in metabolizing a wide range of nitrogen-containing heterocycles such as purines, pyrimidines, pteridines, quinolines, and diazanaphthalenes (95). For example, phthalazine is rapidly converted to 1-phthalazinone by AO and the prodrug, 5-ethynyl-2-(l//)-pyrimidone, is oxidized to the dihydropyrimidine dehydrogenase mechanism-based inhibitor, 5-ethynyluracil, by AO (Fig. 4.40) (96). 

Although demethylation, which occurs in the liver, is normally considered to be a catabolic process, it may result in conversion of an inactive form of a drug to the active form. Thus 6-(methylthio)purine (XXXIX) is demethylated by the rat to 6-mercaptopurine [205]. This demethylation occurs in the liver micro-somes and is an oxidative process which converts the methyl group to formaldehyde [204, 207]. The 1-methyl derivative of 4-aminopyrazolo[3,4-d] pyrimidine (XLI) is demethylated slowly, but 6-mercapto-9-methylpurine (XLII) not at all [208]. The A -demethylation of puromycin (XLlIl) [209, 210], its aminonucleoside (XLIV) [211], and a number of related compounds, including V-methyladenine and V,V-dimethyladenine, occurs in the liver microsomes of rodents [212]. In the guinea-pig the rate-limiting step in the metabolism of the aminonucleoside appears to be the demethylation of the monomethyl compound, which is the major urinary metabolite [213]. The relationship of lipid solubility to microsomal metabolism [214], and the induction of these demethylases in rats by pre-treatment with various drugs have been studied [215]. 

Studies on the mechanism of action of 6-mercaptopurine are complicated by the fact that its anabolic product, thioinosinic acid, is further metabolized by oxidation to 6-thioxanthylic acid [219] and by methylation to 6-(methylthio)purine ribonucleotide [206, 296]. the effects of which could be even more important than those of thioinosinic acid itself, since the methylthio compound is about 20 times as potent as a feedback inhibitor [289]. 

The pyrazolo[3, 4-d] pyrimidines are substrates for and inhibitors of xanthine oxidase [266, 267]. 4-Hydroxypyrazolo[3,4-d] pyrimidine was first investigated for its ability to protect 6-mercaptopurine and other analogues from oxidation by xanthine oxidase [384], but it also inhibits the oxidation of the natural purines, hypoxanthine, and xanthine. Its profound effect on uric acid metabolism made it an obvious choice for the treatment of gout and its utility in the control of this disease has been demonstrated [385, 386]. 

Certain derivatives of 6-mercaptopurine, such as 6-(methylthio)purine, 6-mercaptopurine-3-oxide [448a], and 6-mercaptopurine ribonucleoside and its acylated derivatives apparently owe their activity to their in vivo conversion to 6-mercaptopurine [11,13]. It would appear, however, that the 9-alkyl derivatives of 6-mercaptopurine, and its arabinosyl and xylosyl derivatives, are not metabolized-except in the case of the 9-alkyI derivatives, to a limited extent to their 5-glucuronides—and that their mechanism of action is quite different from that of 6-mercaptopurine. 

---