George S. Morris

Allopurinol in renal failure and the tumour lysis syndrome

This paper illustrates several important points relating to the use of allopurinol in renal failure, or situations of purine overproduction: It is very easy to give too much allopurinol. Most of the side effects (bone marrow depression, exfoliative dermatitis, etc) are the result of overdosage due to the retention of oxipurinol, an effect exaggerated by thiazide diuretics. Monitoring of plasma oxipurinol levels (ideally less than 100 mumol/l) by high-pressure liquid chromatography is helpful for adjusting dosage in renal failure. Some estimate of the anticipated purine excess is equally vital in deciding dosage during tumour lysis if the risk of urate nephropathy is not to be substituted for the certainty of xanthine nephropathy. In this situation the use of allopurinol may even be questioned. Patients with HGPRT deficiency are exquisitely sensitive to allopurinol, and careful monitoring of the effect on urinary purine levels is essential if xanthine colic is to be avoided.

Use of a fundamental elution protocol for the development of reversed-phase high-performance liquid chromatography enabling rapid simultaneous determination of purines, pyrimidines and allied compounds commonly found in human biological fluids

Many high-performance liquid chromatographic methods for the analysis of nucleosides and bases have been developed for a specific requirement. Most are not directly applicable to the separation of all the purines and pyrimidines found in biological fluids. A method has been developed to separate a wide range of compounds found in human urine and plasma in a single run without unduly long run times. Special attention has been paid to sample preparation. Isocratic systems derived from this method for the analysis of specific compounds and enzyme assay mixtures over even shorter periods are described.

dATP accumulation and ATP depletion in platelets in adenosine deaminase deficiency: Significance for the immune response?

High levels of dATP and dADP, accompanied by ATP depletion, were found in the platelets of two ADA-deficient children with severe combined immunodeficiency (SCID). In vitro studies demonstrated that even normal platelets had the ability to make dATP from deoxyadenosine (dAR) under physiological conditions. This capability was greatly enhanced by conditions simulating ADA deficiency. These results question whether the platelet has a specific role in the normal immune response.

CORRELATIONS BETWEEN PURINE LEVELS, CLINICAL AND.IMMUNOLOGICAL STATUS IN ADA DEFICIENCY: 192

Nine cases of adenosine deaminase (ADA) deficiency have been found amongst fifty infants investigated for immunodeficiency. Prenatal diagnosis identified a further two affected foetuses in six pregnancies at risk. Absence of ADA activity and high dATP levels in foetal blood,with low lymphoid cell numbers and few T-cells, indicated early intrauterine onset.High dATP levels, accompanied by varying degrees of ATP depletion were also found in the red cells of the nine infants. Two late presenters had normal ATP levels, normal lymphocyte numbers and immunoglobulin levels, with some detectable ADA in mononuclear cells (PBMs). dATP was found in PBMs in only one case where heparinised blood contaminated with platelets was used. The novel finding in the two latest cases was the presence of ATP depletion accompanied by dATP accumulation in the platelets but not PBMs from defibrinated blood. This explains the earlier results and questions the possible significance for the immune response.Deoxyadenosine was found in the urine in all cases.S-adenosylhomocysteine was not; adenosine was identified in some. The results suggest that the degree of ATP depletion may reflect the clinical status most closely,but that any therapeutic approaches are likely to be ineffective because of the early toxicity to T-lymphocyte precursors.

B-LYMPHOCYTES THYMOCYTES AND PLATELETS ACCUMULATE HIGH dATP LEVELS IN SIMULATED ADA DEFICIENCY: 69

Novel findings in vitro and in vivo obtained previously in simulated and inherited ADA deficiency were investigated using [8-14C] deoxyadenosine (dAR) in short-term experiments in intact human cells of the myeloid and lymphoid series.The studies produced several interesting results. (1) Tonsil-derived B-lymphocytes,thymocytes and platelets all accumulated detectable amounts of dATP even without ADA inhibition, and together with erythrocytes,extremely high dATP levels when ADA was inhibited by deoxycoformycin (dCF): varying amounts of dCF (20-60μm) were needed to completely inhibit ADA depending on the cell type. (2) By contrast,dATP accumulation by peripheral blood lymphocytes,granulocytes and macrophages was negligible without, and extremely low even with dCF. (3) B-lymphocytes showed a capacity equal to that of thymocytes in their ability to sustain the elevated dATP levels accumulated in ADA deficiency conditionsThe results support earlier findings which question the hypothesis that B-cells,compared with T-cells, have an inherent resistance to the toxic effects of dAR because of a lower ability to accumulate and sustain elevated dATP levels. They underline the difficulty in extrapolating from lysed or cultured cells to the situation in vivo in the peripheral blood. They suggest that the severe combined immunodeficiency in this disorder may be due to an equal sensitivity of B-lymphocytes and T-lymphocyte precursors to the toxic effects of dATP accumulation.

THE USE AND ABUSE OF ALLOPURINOL IN RENAL FAILURE THE TUMOUR LYSIS SYNDROME AND UROLITHIASIS: 24

Despite widespread use, side effects of allopurinol have been rare. Problems arising in chronic renal failure (CRF) have been well documented,but necessity to reduce dosage in such cases is often ignored. Not so well known is the acute sensitivity to xanthine oxidase inhibition in situations of gross purine overproduction and the attendant risk of xanthine stones, or acute renal failure (ARF) from xanthine nephropathy.To underline potential hazards and their avoidance, purine levels found in collaborative studies on allopurinol-treated patients are reported. 90% of referrals involved monitoring of plasma oxipurinol levels in CRF. Results frequently exceeded the desirable level of 100μmol/l, with values as high as 310μmol/l in those receiving 300mg/day and still 60μmol/l three days after stopping therapy in one case complicated by exfoliative dermatitis. One death from bone marrow aplasia had occurred prior to general awareness of the marrow depressant effects of sustained oxipurinol levels in CRF.Four cases of ARF due to xanthine nephropathy occurred during therapy for conditions resulting in rapid cell lysis where plasma xanthine (normally <1μmol/1) and oxipurinol levels of 650μmol/l and 642μmol/l,respectively,were recorded. Two Lesch-Nyhan patients developed xanthine stones and required careful monitoring.The results demonstrate the effects of allopurinol overdose which should be avoidable through knowledge of purine metabolism in the particular situation.

A SINGLE SYSTEM FOR THE EVALUATION OF PURINE AND PYRIMIDINE NUCLEOSIDES AND BASES TOGETHER WITH THEIR ANALOGUES IN BIOLOGICAL FLUIDS: 137

Procedures for the HPLC analysis of nucleosides and bases have generally been designed to fulfil specific requirements. No single method exists capable of separating all the purines and pyrimidines found in the biological fluids of patients in the different inherited disorders in a single run and within a reasonable time. Because of a need for such a system an appropriate method has been devised.The system has been in continuous use for over two years and three to four thousand samples have been analysed. It has proved reliable and reproducible throughout the life of many columns. Problems encountered during this period have resulted from artifacts and errors inherent to different isolation/extraction procedures. Pitfalls frequently noted have in the main been due to diet and/or drug metabolites with similar chromatographic behaviour to endogenous purines and pyrimidines.These problems are exaggerated in renal failure and during antibiotic therapy.Data derived from the chromatography of body fluids of patients with various purine and pyrimidine defects,illustrate the adaptability of the system. The method is particularly applicable to the separation of endogenously derived adenosine based compounds. An abbreviated form of the system has been used successfully for enzyme assays and other in vitro studies.