azaserine and Lesch-Nyhan-Syndrome

Genetic defects in purine metabolism are associated with severe immunodeficiency. Adenosine deaminase deficiency impairs the function of both B- and T-lymphocytes whereas in purine nucleoside (inosine) phosphorylase deficiency there is more severe impairment of T-lymphocyte functions than of B-lymphocyte functions. The relative unimportance of the salvage pathway catalysed by hypoxanthine-guanine phosphoribosyltransferase is shown by the normal responses of T-lymphocytes from patients with the Lesch-Nyhan syndrome to antigenic and mitogenic stimulation. A mild deficiency of B-lymphocyte function is found in these patients. Agents inhibiting the de novo pathway of purine synthesis, including azaserine, 6-mercaptopurine and azathioprine in low doses, block the responses of normal human lymphocytes to mitogenic stimulation. These observations emphasize the importance of the de novo pathway of purine synthesis in lymphocyte responses to antigenic and mitogenic stimulation. There is considerable heterogeneity in the amount of labelled uridine incorporated into human and rat lymphocytes. This does not appear to reflect only a difference between T- and B-lymphocytes

Topics: Adenine Phosphoribosyltransferase; Adenosine Deaminase; Adolescent; Adult; Allopurinol; Azaserine; B-Lymphocytes; Child; Child, Preschool; Humans; Hypoxanthine Phosphoribosyltransferase; Immunoglobulins; Lectins; Lesch-Nyhan Syndrome; Lymphocyte Activation; Male; Mercaptopurine; Mitogens; Phosphoribosyl Pyrophosphate; Purine-Nucleoside Phosphorylase; Purines; RNA; T-Lymphocytes; Thymidine; Uridine

Lesch-Nyhan syndrome is a hereditary disorder of purine metabolism causing overproduction of uric acid and neurological problems including spasticity, choreoathetosis, mental retardation, and compulsive self-mutilation. The syndrome is caused by a defect in the enzyme hypoxanthine-guanine phosphoribosyltransferase (HPRT), which converts guanine and hypoxanthine to the nucleotides GMP and IMP. There is evidence that the neurological problems are due to an adverse effect of the HPRT deficiency on the survival and/or development of dopaminergic neurons, specifically. Here we report that HPRT-deficient PC12 mutants that have a normal or near normal dopamine content (55-97% of that of wild-type cells) fail to undergo neuronal differentiation induced by nerve growth factor (NGF) when the de novo pathway of purine synthesis is partially inhibited. However, nerve growth factor-induced differentiation is near normal under these conditions in PC12 HPRT-deficient mutants containing much lower dopamine levels (<8% of that of wild type cells), indicating a neurotoxic effect of the endogenous dopamine in the mutants. The degree of inhibition of the de novo pathway of purine synthesis was the same in both classes of HPRT-deficient mutants. Expression of BCl-2 in a PC12 mutant that has a normal dopamine content allowed partial NGF-induced differentiation suggesting that the apoptotic pathway might be involved in the failure of differentiation when the de novo pathway of purine synthesis is partially inhibited.

Topics: Animals; Antimetabolites, Antineoplastic; Azaserine; Blotting, Northern; Cell Differentiation; Cell Survival; Dopamine; Humans; Hypoxanthine Phosphoribosyltransferase; Lesch-Nyhan Syndrome; Neurites; Neurons; PC12 Cells; Proto-Oncogene Proteins c-bcl-2; Purines; Rats; Tyrosine 3-Monooxygenase

Topics: Azaserine; Cells, Cultured; Genetic Carrier Screening; Humans; Hypoxanthine; Hypoxanthine Phosphoribosyltransferase; Hypoxanthines; Kinetics; Lesch-Nyhan Syndrome; Lymphocyte Activation; Lymphocytes; Male; Reference Values; Thioguanine

Three patients with the Lesch-Nyhan syndrome were found to have normal delayed hypersensitivity, peripheral-blood T-lymphocyte counts, lymphocyte responses to P.H.A., and serum IgM, IgA, and IgE levels. However, the percentages of B-lymphocytes, IgG levels, serum-isohaemagglutinin titres, and lymphocyte responses to pokeweed mitogen (P.W.M.) were subnormal. These observations suggest that activity of the salvage pathway of purine synthesis catalysed by hypoxanthine-guanine phosphoribosyl transferase (H.G.P.R.T.) is not required for the responses of T-lymphocytes to mitogenic or antigenic stimulation, but may contribute to the proliferation and function of B lymphocytes. The major role of the de-novo pathway of purine synthesis in human lymphocyte responses to mitogenic or antigenic stimulation is shown by the effects of inhibitors of this pathway, including immunosuppressive agents, and by the effects of congenital deficiency or inhibition of adenosine deaminase.

Topics: Adenosine Deaminase; Adolescent; Azaserine; B-Lymphocytes; Child; Child, Preschool; Hemagglutinins; Humans; Hypersensitivity, Delayed; Hypoxanthine Phosphoribosyltransferase; Immunoglobulins; Immunologic Deficiency Syndromes; In Vitro Techniques; Lectins; Lesch-Nyhan Syndrome; Lymphocyte Activation; Lymphocytes; Male; Mercaptopurine; Mitogens; Purines; T-Lymphocytes; Thymidine

We have previously described a 14-yr-old boy with hyperuricemia, renal failure, and accelerated purine production resistant in vivo and in vitro to purine analogs. This patient demonstrated normal red cell hypoxanthine-guanine phosphoribosyltransferase (HPRT) heat stability, electrophoresis at high pH, and activity at standard substrate levels. In the present report an abnormal HPRT enzyme was demonstrated by enzyme kinetic study with phosphoribosylpyrophosphate (PRPP) as the variable substrate and inhibitory studies with sodium fluoride. Apparently normal HPRT activity in a patient with hyperuricemia and gout does not exclude a functionally significant HPRT mutation.

Topics: Adolescent; Aminopterin; Azaguanine; Azaserine; Cells, Cultured; Culture Media; Erythrocytes; Fibroblasts; Guanine; Humans; Hydrogen-Ion Concentration; Hypoxanthines; Kinetics; Lesch-Nyhan Syndrome; Male; Mutation; Organophosphorus Compounds; Pentosephosphates; Pentosyltransferases; Phosphoric Acids; Purine-Pyrimidine Metabolism, Inborn Errors; Purines; Temperature; Uric Acid

Topics: Adenine; Azaserine; Carbon Isotopes; Chromatography, Thin Layer; Diphosphates; Fibroblasts; Formates; Glycine; Humans; Lesch-Nyhan Syndrome; Nicotinic Acids; Pentosephosphates; Pentosyltransferases; Purines; Ribonucleotides; Ribose; Skin

Topics: Adenine; Adenosine; Azaserine; Cells, Cultured; Fibroblasts; Guanine; Humans; Hypoxanthines; Inosine; Lesch-Nyhan Syndrome; Phosphotransferases; Skin; Thymidine; Tritium

Cultured fibroblasts established from skin biopsies from patients with the Lesch-Nyhan syndrome are deficient in hypoxanthine-guanine phosphoribosyl-transferase (EC 2.4.2.8) activity. This deficiency makes possible the use of chemicals that select either for or against deficient variants in cultured fibroblasts. Two-way selection has been achieved by the use of 6-thioguanine, which selects for the deficient mutant, and azaserine, which selects to some extent for the normal allele in mixed cultures, as well as in cultures from heterozygotes. Theoretical considerations predict that the phenomenon of metabolic cooperation would tend to reinforce the former and to weaken the latter type of selection, and this is in accordance with the experimental findings.

Topics: Autoradiography; Azaserine; Cell Line; Culture Techniques; Fibroblasts; Genetics, Medical; Guanine Nucleotides; Heterozygote; Humans; Hypoxanthines; Lesch-Nyhan Syndrome; Mutation; Pentosephosphates; Selection, Genetic; Skin; Thioguanine; Time Factors; Transferases; Tritium