HGPRT deficiency

Remondelli P, Mugnoz B, Della Morte R, et al. 1986. Evaluation of the mutagenic potential of thirteen pesticides by measuring both his+ and HGPRT-deficient mutants in Salmonella typhimurium. Med Biol Environ 14 377-386. 

HGPRT deficiency (l.esch-Nyhan syndrome) Hypoxanthine-guanine phosphor ibosy 111ansferase... 

The relative importance of the de novo and salvage pathways is unclear. However, the severe symptoms of hereditary HGPRT deficiency indicate that the purine salvage pathway is vitally important. In addition, investigations of purine nucleotide synthesis inhibitors for treating cancer indicate that both pathways must be inhibited for significant tumor growth suppression. 

Dramatic Consequences of a Severe HGPRT Deficiency Lesch-Nyhan Syndrome... 

Recently a new cause of supposed uric acid stones in children was identified independently by two groups (2, 3), a defect of the companion salvage enzyme adenine phos-phoribosyltransferase (APRT). This results in an inability to salvage the purine base adenine, which is then picked up by xanthine oxidase and converted to the extremely insoluble analogue of uric acid, 2,8-dihydroxyadenine (2,8-DHA) (Fig. 1). In contrast to HGPRT deficiency, other than the urolithiasis, these children are normal and heterozygotes have no clinical abnormality. 

In some recent studies [1-4, 6], Cleaver et al. have questioned the validity of poly-(ADP-ribose) polymerase studies that are based on the use of inhibitors of ribosyla-tion such as SAB. The high concentrations of SAB commonly used might cause other cellular effects unrelated to poly(ADP-ribose) synthesis. For example, SAB has been reported to inhibit de novo nucleotide synthesis [1-4, 6]. Inhibition of de novo nucleotide synthesis could slow cell growth and lead to imbalances of nucleotide pools. Cleaver [4] has recently reported that a 6TG -CHO line, unable to incorporate exogenous purines and therefore HGPRT-deficient, was more sensitive to the cytotoxic effects of SAB as compared to a CHO line proficient in HGPRT activity. The increased sensitivity is believed to be due to the reported SAB-mediated inhibition of de novo nucleotide synthesis [1-4,6]. 

The spectrum of manifestations in both is broad with similarities, but also important differences Both are a sociated with urinary calculi and can cause severe renal damage Severe neurological problems are also found in complete HGPRT deficiency but have not been noted in APRT deficiency We have investigated 2 children presenting in renal failure that emphasise the problems of diagnosis and treatment of these 2 deficiencies and the particular difficulties encountered in the presence of impaired renal function. 

HGPRT activity in L.W. was less than 0.01 nmol/mg HbA in lysed red cells, less than 0.9 nrool/mg proteinA in skin fibroblasts and less than 0 2% of normal in intact erythrocytes. Table 2. Incorporation of labelled hypoxanthine into purine nucleotides by intact fibroblasts was approximately 6% of control which is low on the criteria of Page et al. These results confirm severe HGPRT deficiency. Heterozygote levels were found in the Mother s fibroblasts... 

These 2 patients phasise some of the diagnostic and therapeutic difficulties encountered when renal failure complicates APRT or HGPRT deficiency, particularly in childhood. 

Treatment in both children involved the use of allopurinol - but at a lesser dosage (5 mg/kg/2hh) because of the retention of the principal metabolite oxipurinol in severe renal failure Careftil monitoring of plasma oxipurinol levels is desirable For the HGPRT deficient child alkali was also given to enhance uric acid solubility It was not prescribed in the APRT deficient child since 2 8-dihyroxy adenine solubility is unaffected by alkali and its use may even be contraindicated ... 

Finally, the difficulty of heterozygote detection in HGPRT deficiency is exemplified in this study Erythrocyte lysates from the mother and her sisters had normal HGPRT activity, the low fibroblast levels exclusively in the mother suggest a new mutation By contrast the mother of the APRT deficient child had erythrocyte lysate levels much less than the anticipated 30% of the normal mean (ca 23%) This is characteristic of the defect and will be discussed elsewhere (Wilson et al - this symposium). 

This patient illustrates some of the difficulties in the diagnosis of HGPRT deficiency when the patient presents in acute renal failure, the clinical syndrome falls short of the full Lesch-Nyhan syndrome, and especially highlights the problems attendant on the evaluation of HGPRT activity from erythrocyte lysates alone. 

HYPOXANTHINE-GUANINE PHOSPHORIBOSYL TRANSFERASE (HGPRT) DEFICIENCY IN A GIRL... 

Genetic Mechanism of HGPRT Deficiency in the Girl, Whose Mother is... 

The extremely low red cell NAD levels also indicate involvement of pyridine nucleotide metabolism in the clinical expression of this disorder. PP-ribose-P is require for the formation of the pyridine nucleotides NAD and NADP both are vi al to the erythrocyte. However, high, rather than low NAD levels would be anticipated and have been found by us in both HGPRT and PNP deficient red cells it is possible that the raised NAD in the latter reflect the raised PP-ribose-P levels found in PNP and HGPRT deficiency. PP-ribose-P levels were not raised in NB s red cells and the NAD values were extremely low. The latter finding may also be associated with the observed inability of the intact red cells to stimulate the re-cycling of hypoxanthine or adenine at high phosphate and suggest that NAD may be necessary for the stabilisation of erythrocyte PP-ribose-P. This would in turn... 

These studies show that diagnosis of the aberrant enzyme may be impossible from red cells alone but that altered nucleotide levels together with the raised uric acid values in plasma and urine in both mother and child, provide useful diagnostic tools. This again distinguishes e defect from HGPRT deficiency where mothers are rarely affected. 

Increased intracellular levels of PP-ribose-P have been implicated in the cause of certain hyperuricemic states associated with uric acid overproduction. Fibroblasts from two patients with the Lesch-Nyhan syndrome were found previously to have an elevated intracellular concentration of PP-ribose-P with a normal rate of PP-ribose-P production (Rosenbloom, et al., 1968). Green and Seegmiller (1969) subsequently reported a mean PP-ribose-P value of 47.1 in erythrocytes from seven patients with HGPRT deficiency. We have confirmed these elevated PP-ribose-P levels in three additional patients with the Lesch-Nyhan syndrome with values of 20.5, 39.4 and 49.5 juM (Table 1). The mothers of these patients are obligate heterozygotes and have normal PP-ribose-P levels. Two diseases associated with a deficiency of other PRT enzymes are not associated with altered erythrocyte PP-ribose-P levels (Table 1). PP-ribose-P levels were in the normal range in one patient with a partial deficiency of adenine phosphoribosyltransferase (APRT) and in one patient with orotic aciduria, which is due to a deficiency... 

In patients with nearly complete deficiency of erythrocyte HGPRT activity (either Gout or LESH NYHAN syndrome) thiopurinol has no effect on plasma and urinary excretion of uric acid (DELBARRE et al. 1970), while in the same patients treated with allopurinol there is a rapid andimportant decrease of uric acid balanced by nearly stochiometric increase of oxypurines. Gouty patients with HGPRT deficiency have higher urinary oxypurine excretion with a more important contribution of hypoxanthine (H/X = 2,36) than... 

The per cent of control values in partial HGPRT deficiency is the same as well as with hypoxanthine and guanine or with alio and thiopurinol as substrat. In the particular enzymatic deficiency LUG, who has nearly normal PRT activity with hypoxanthine as substrat, rate of synthesis of alio and thiopurinol are about 30 % of normal value. 

Cultured human skin fibroblasts are commonly utilized in the detection of hemizygosity and heterozygosity for hypoxanthine-guanine phosphoribosyltrans-ferase (HGPRT) deficiency. Two obstacles are encountered in the determination of HGPRT and adenine phosphoribosyl-transferase (APRT) in extracts of cultured skin fibroblasts the sensitivity of these enzymes in dilute cell suspension to freezing and thawing (l), and the presence of nucleotidase activity (2). 

The specific activities of HGPRT and APRT were determined in extracts of cultured skin fibroblasts from both normal and HGPRT-deficient subjects, comparing the results obtained with and without TTP (Table l). 

It is noteworthy that for the Lesch-Nyhan syndrome fibroblasts, the assay in the presence of TTP furnished the bulk of the radioactivity in the nucleoside fraction rather than in the nucleotide fraction, in contrast to normal cell extracts in which the bulk of the radioactivity is found in the nucleotide fraction. Two possible explanations may be entertained to explain this observation. First, in the HGPRT-deficient system the nucleotidase activity is far in excess over the HGPRT activity, and presumably the residual nucleotidase activity, not inhibited by TTP, is sufficient to degrade the relatively small amount of nucleotide formed. Secondly, the possibility should be considered that the accumulation of radioactivity in the nucleoside fraction reflects the anabolic activity of nucleoside phospho-rylase reacting the purine base with ribose-l-phosphate to form the nucleotide by an alternative pathway. However, this latter explanation seems to be invalid in view of the absence of the suitable substrate, ribose-l-phosphate, from the incubation system, and by the linearity... 

TABLE 1. SPECIFIC ACTIVITIES OP HGPRT AND APRT IN NORMAL AND HGPRT DEFICIENT CULTURED SKIN FIBROBLASTS, ASSAYED WITH AND WITHOUT THYMIDINE-5 -TRIPHOSPHATE... 

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