This paper reports the detection of five inherited disorders of purine and one of pyrimidine metabolism using intact red blood cells (RBCs) and compares the findings with those from RBC lysate activity. Two different phosphate levels (1 and 18 mmol L-1 Pi) were used to evaluate endogenous PP-ribose-P levels and their generation by PP-ribose-P synthetase. The importance of this dual approach is demonstrated by the following evidence: (a) Six out of eight patients with no detectable hypoxanthine-guanine phosphoribosyltransferase (HGPRT) RBC lysate activity had up to 25% of normal activity in their intact RBCs. Two Lesch-Nyhan patients showed no detectable activity in intact or lysed RBCs. (b) RBC lysates from two heterozygotes for adenosine deaminase (ADA) deficiency also showed no detectable activity, but up to 60% of normal activity using intact RBCs. (c) The existence of an aberrant enzyme in a kindred with a superactive PP-ribose-P synthetase was evident from the fact that intact RBCs failed to respond normally to phosphate activation, despite normal HGPRT and adenine phosphoribosyltransferase (APRT) RBC lysate activity. (d) Raised endogenous PP-ribose-P levels in intact RBCs were demonstrable only in purine nucleoside phosphorylase (PNP) and HGPRT deficiency; levels were normal in APRT deficiency and hereditary oroticaciduria (OPRT/ODC) deficiency. The results indicate that diagnosis from RBC lysate activity alone may be misleading. Intact RBC studies clearly provide a better indication of the functional capacity of the enzyme in vivo. They also show a closer correlation with the clinical phenotype and allow further insight into the associated biochemical abnormalities in some cases.

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D e o x y a d e n o s i n e (dAdo) is toxic at micromolar concentrations to adenosine deaminase inhibited resting human peripheral blood lymphocytes. 2-chlorodeoxyadenosine (CdA), a metabolically resistant dAdo congener, exhibits similar properties. Four hours of exposure to dAdo or CdA induced the accumulation of strand breaks in the DNA of normal resting lymphocytes, as measured by a DNA unwinding assay. The DNA damage was followed by consumption of NAD. probably mediated by increased poly(ADP-ribose) synthesis. The addition of 1-5mM nicotinamide prevented the dAdo and CdA triggered fall in NAD levels, and rendered the resting lymphocytes resistant to the toxic effects of both compounds. Both dAdo and CdA inhibited the repair of radiation induced DNA damage in resting lymphocytes, by impeding DNA polymerization. CIA was 100 fold more potent than dAdo. These results suggest that (i) a slow rate of DNA polymerization is required to maintain DNA integrity in resting human lymphocytes, (ii) dAdo and CdA inhibit polymerization, and cause DNA strand breaks to accumulate. (iii) the strand breakage triggers poly(ADP-ribose) synthesis, and causes lethal NAD depletion. In addition, the HL-60 promyelocytic leukemia cell line is 300-fold less sensitive to araG than are T lymphoblasts, and the ability of the promyelocytes to accumulate araGTP is similar to that of the 24B3 cell 1 ine. These data suggest that sensitivity to araG is mediated through a cell-specific kinase which is more selective for dGuo analogs than for a deoxycytidine analog such as araC. CHARACTERIZATION OF ARABINOSYLGUANINE (AraG)18890 Mitchell. University of Michigan Medical Cendr.Department of Internal Medicine, Ann Arbor, MI. 5-Amino-4-imi dazolecarboxamide riboside (Z-ri boside) is an intermediate in the purine de novo synthetic pathway which may be metabolized either to I~m f o l a t e -d e p e n d e n t reaction or to the corresponding Z-nucleotides. Accumulation of ZTP in microorganisms has been associated with depletion of folate intermediates and has been postulated to play a regulatory role in cellular metabolism. We have demonstrated the presence of Z-nucleotides in erythrocytes derived from 5 individuals with the Lesch-Nyhan syndrome. Erythrocyte folate levels were within the normal range in these individuals, although GTP and GDP levels were significantly reduced bel6w those in normal control (p<0.001). Lesser amounts of Z-nucleotides were found in 3 individuals with partial deficiency of the enzyme hypoxanthine guanine phosphoribosyltransferase and in 2 individuals with other disorders of purine overproduction. In contrast, no Znucleotides were detected in 13 normal controls or in 3 individuals with gout on allopurinol therapy. We conclude that Znucleotide formation may result from markedly increased rates of de novo purine biosynthesis. Whether or not the further me- Dept. of Med.. U. of Wa.. Seattle, WA, USA. Apparent crystal-negative effusions are sometimes seen in the early course of acute gouty arthritis. Also, newly-prepared urate cry...

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