Sebastian Reiter

Demonstration of a combined deficiency of xanthine oxidase and aldehyde oxidase in xanthinuric patients not forming oxipurinol

Genetic heterogeneity has been suggested in xanthinuria from the hitherto unexplained ability of some patients with this hereditary disorder to convert allopurinol to its active metabolite oxipurinol--an activity generally attributed to xanthine oxidase. This study provides evidence that the enzyme aldehyde oxidase is also deficient in xanthinuric patients not converting allopurinol to oxipurinol, whereas a xanthinuric patient with normal formation of oxipurinol had normal aldehyde oxidase activity. It is concluded that the enzyme aldehyde oxidase is the principal enzyme responsible for the formation of oxipurinol in man.

Influence of Dietary Purines on Allopurinol Metabolism and Allopurinol Induced Oroticaciduria

The allopurinol induced oroticaciduria is markedly diminished by oral administration of purines.1 The underlying mechanism is still unknown. We performed nutrition experiments to determine whether this effect could be due to an influence of dietary purines on either absorption or metabolism of allopurinol. A change of allopurinol metabolism by intravenous administration of adenine in the pig was previously reported.2

Activity of Adenine Phosphoribosyltransferase (Aprt) in Patients with Renal Failure and Urolithiasis

The common clinical manifestation of complete APRT-deficiency is 2,8-dihydroxyadenine (2,8-DHA) urolithiasis. Acute and chronic renal failure have also been reported1. On the other hand, homozygotes, either with late onset2or without symptoms3, have been described. Heterozygous patients are usually symptom-free and only occasionally form stones. It has been suggested that APRT-deficiency might not be a rare disease and is often overlooked in patients with renal failure or urolithiasis1. Thus, screening for APRT-deficiency in these patients seemed worthwhile.

Adeninephosphoribosyltransferase (APRT)-Activity in Patients with Nephrolithiasis or Renal Failure

The homozygous state of APRT-deficiency is associated with high urinary levels of 2, 8-dihydroxyadenine (2, 8-DHA) and may lead to nephrolithiasis and renal failure (Cartier and Barnet 1974; van Acker et al., 1977). Heterozygous patients are usually without symptoms (Fox et al., 1977). Recently Kuroda et al. (1981) reported of heterozygous patients with elevated 2, 8-DHA excretion and 2, 8-DHA stones.

RNA Environment Is Responsible for Decreased Photostability of Uracil

UV light can induce chemical reactions in nucleic acids and thereby damage the genetic code. Like all of the five primary nucleobases, the isolated RNA base uracil exhibits ultrafast, nonradiative relaxation after photoexcitation, which helps to avoid photodamage most of the time. Nevertheless, within RNA and DNA strands, commonly occurring photolesions have been reported and are often ascribed to long-lived and delocalized excited states. Our quantum dynamical study now shows that excited-state longevity can also occur on a single nucleobase, without the need for delocalization. We include the effects of an atomistic RNA surrounding in wave packet simulations and explore the photorelaxation of uracil in its native biological environment. This reveals that steric hindrance through embedding in an RNA strand can inhibit the ultrafast relaxation mechanism of uracil, promoting excited-state longevity and potential photodamage. This process is nearly independent from the specific combination of neighboring bases.

DOSE DEPENDENCY OF THE URIC ACID LOWERING EFFECTS OF ALLOPURINOL WITH AND WITHOUT ORAL PURINES: 111

Healthy young volunteers received a purine-free formula diet for 18 days. Allopurinol (A) was given from day 11-18 in single oral doses of 125, 250, 375 and 500 mg/m2 body surface to 4, 8, 4 and 8 subjects, respectively. In a second study 4 of the 8 subjects on 250 and 500 mg/m2 took the same doses with the formula diet being supplemented by 4 g RNA/d. Maximum reduction of renal uric acid excretion was 55% and 70% during purine-free diet and RNA intake, respectively. On 250 and 500 mg/m2 reduction of endogenous renal uric acid was 36 and 46%, while exogenous uric acid was reduced by 74 and 86%, respectively in the 4 subjects. Renal uric acid clearance was increased by A during purine-free diet, but was not during RNA loading. In a different series of experiments A was given alone first followed by additional purine intake. On this regimen uric acid excretion did not increase during purine ingestion in most instances. This suggests previous inhibition by A or stimulation by oral purines of xanthine oxydase to be a factor in determining the influence of A on exogenous uric acid production.

URINARY OXIPURINOL-1-RIBOSIDE EXCRETION AND ALLOPURINOL-INDUCED OROTICACIDURIA: 169

Allopurinol-induced oroticaciduria which is due to inhibition of OMP-decarboxylase by exipurinol-1- and -7-ribotide is markedly reduced by dietary purines. The underlying mechanism possibly is an influence of dietary purines on the formation of oxipurinol-ribotides. The latter being excreted as their corresponding nucleosides we first measured the urinary excretion of oxipurinol-7-riboside which was found not to be altered by dietary purines. Therefore we searched for oxipurinol-1-riboside which had not yet been detected in human urine. By HPLC we were able to demonstrate the presence of this metabolite. In the absence of dietary purines its excretion was similar to that of oxipurinol-7-riboside, but was strongly reduced by purine supplement. Having shown the indirect formation of allopurinol-1-riboside via allopurinol-1-ribotide in vivo(cf. the accompanying paper) we assume that the excretion of oxipurinol-1-riboside parallels the formation of oxipurinol-1-ribotide as well. We conclude from our data that dietary purines reduce allopurinol-induced oroticaciduria by inhibition of oxipurinol-1-ribotide synthesis.

ALLOPURINOL-1-RIBOSIDE FORMATION AND HGPRT-DEFICIENCY: 170

Allopurinol-1-riboside has been thought to be synthesized in vivo rather directly by purine nucleoside phosphorylase(PNP) than indirectly via the ribotide by hypoxanthine guanine phosphoribosyl-transferase(HGPRT). Consequently the lack of allopurinol-1-riboside formation in HGPRT-deficiency has been attributed to competitive inhibition of PNP by increased hypoxanthine levels(Elion, Handb.Exp.Pharm.51). Having demonstrated elevated excretion of allopurinol-1-riboside in PNP-deficiency we studied the influence of hypoxantbine on allopurinol-1-riboside formation in healthy volunteers and a patient with partial(5% residual activity) HGPRT-deficiency. Allopurinol was given as a single dose of 5mmol, the volunteers received additionally 7,5mmol of different purines. The volunteers shoved a reciprocal relationship between allopurinol-1-riboside excretion(0,615-0,034 mmol) per 24hrs and hypoxanthine excretion(0,082-1,22mmol) during the first 5hrs following allopurinol and purine intake - the period of allopurinol-1-riboside formation. The corresponding values of our patient were 0,014mmol allopurinol-1-riboside and 0,24mmol hypoxanthine. These data indicate that the very low excretion of allopurinol-1-riboside in HGPRT-deficiency must be attributed to the enzyme defect itself as it cannot be explained by the moderate elevation of hypoxanthine formation.

UNPROPORTIONAL INCREASE IN HEMOLYSATE OF THE ACTIVITY OF A MUTANT HPRT DURING HEMODIALYSIS: 112

A patient having been treated by hemodialysis because of gouty nephropathy since 1973 was diagnosed to have partial HPRT deficiency (19 nmol/mg proteinxh in hemolysate) in 1978. His grandson presented with acut renal failure at the age of 3 weeks in 1982. After an initial value of 31 nmol/ mgxh after previous transfusion his enzyme activity consistently was below 1 nmol/mgxh. Intact red cell studies revealed a twofold activity in the propositus compared with his grandson (12.2 and 6.4 nmol/ml packed cells per h). In lysates of cultured fibroblasts both patients showed the same activity (0.8 nmol/mgxh). In both patients kinetic studies of the enzyme gave normal results and heat inactivation showed it to be more labile than normal controls.From family studies as well as activity in cultured fibroblasts it can be concluded that both patients inherited the same enzyme defect. Comparing the results in these two patients the twofold activity in intact red cells of the propositus can be attributed to a shortened half-life of his erythrocytes, whereas the fortyfold increase (range 20-60) in hemolysate is unexplained.