Rejection of bone marrow transplant and resistance of alloantigen reactive cells to in vivo deoxyadenosine in adenosine deaminase deficiency

Severe combined immunodeficiency disease (SCID) in patients with adenosine deaminase (ADA) deficiency is thought to result from increased levels of purine metabolites. We attempted to immunosuppress a patient with ADA deficiency and SCID using a continuous infusion of deoxyadenosine to obtain engraftment of a T cell-depleted haplocompatible parental bone marrow graft. Before administering the drug in vivo, we investigated hematopoietic colony formation in two children with ADA deficiency (including the potential recipient), the obligate heterozygote donor (father), and normal controls using deoxyadenosine and erythro-9-(2-hydroxy-3-nanyl)adenosine (EHNA), and inhibitor of ADA. Deoxyadenosine alone in concentrations as high as 100 microM had no significant affect on erythroid (BFU-E) or myeloid (CFU-c) colony formation. However, in the presence of EHNA there was a significant reduction in BFU-E and CFU-c growth in all subjects and controls. Increasing doses of deoxyadenosine were given to one patient with ADA deficiency and SCID as a continuous 24-hr intravenous infusion. We found that there was a linear relationship between the dose administered and the plasma level; however, doses greater than 100 mg/day were required to increase erythrocyte dATP levels. We were able to raise intracellular dATP levels to more than three times baseline with doses of deoxyadenosine of 200 mg/day. However, there were no significant effects on the absolute lymphocyte counts or the lymphocyte responses to mitogen or alloantigen, and the haploidentical marrow failed to engraft. Our results suggest that the bone marrow of ADA-deficient patients is normal with respect to standard colony formation, that inhibitors of ADA do not adequately model the deficient state, and that the immunodeficiency in ADA deficiency is not proportionately related to either the deoxyadenosine or dATP levels, both of which were significantly elevated at the time of transplantation.     

Resistance of pokeweed mitogen-stimulated B cells to inhibition by deoxyadenosine.

Deoxyadenosine (dAdo) levels above 2 microM inhibit plasma cell (PC) differentiation by human blood lymphocytes in pokeweed mitogen (PWM) stimulated cultures containing deoxycoformycin (dCF), a potent inhibitor of adenosine deaminase (ADA). ADA inhibition by dCF alone did not suppress PC differentiation. Thymidine uptake by T cell blasts continuously cultured in conditioned medium was inhibited by dAdo and dCF; two of five EBV-infected B cell lines were also inhibited while three were resistant. Inhibition of PWM-induced PC differentiation of B cells by dCF and dAdo was reversed when conditioned medium (a source of T cell helper factors) was added to the cultures, and dAdo and dCF added to PWM-stimulated cultures 48 hr after their initiation did not inhibit PC differentiation, though thymidine uptake and the total number of cells recovered from the cultures were reduced. Removal of T cells after 48 hr of culture slightly reduced the numbers of PC in PWM-stimulated lymphocyte cultures but no further inhibition was obtained when dCF and dAdo were added to these T-depleted cultures, nor was their thymidine uptake further reduced. These results suggest that the in vitro suppression of B cell differentiation by dAdo in PWM-stimulated cultures is not due to direct toxicity of purine nucleosides to B cells but may be due to interference with T cell help. This is consistent with the view that a relative lack of helper activity by T cells contributes to the antibody deficiency of patients with ADA deficiency.     

Interleukin 2 responsive lymphocytes in patients with adenosine deaminase deficiency

We evaluated the effects of recombinant interleukin 2 (IL-2) on the proliferative responses to mitogens of peripheral blood mononuclear cells (PBMC) from three adenosine deaminase (ADA)-deficient patients. There was significant enhancement by IL-2 of the proliferative responses to phytohemagglutinin (PHA) and pokeweed mitogen (PWM) of PBMC from all three patients. We found that normal PBMC respond with increased numbers of CD3-positive cells when exposed to PHA or PWM and that the response by normal CD8-positive cells was greater than that by CD4-positive cells. In contrast, we found that in ADA-deficient cells the response is almost entirely due to the CD3/CD4-positive population of lymphocytes. These results could not be explained by either the culture conditions or the possibility of a mixed chimeric state. When we evaluated an in vitro cell model of ADA deficiency using an ADA inhibitor, erythro-9-(2-hydroxy-3-nonyl)adenine (EHNA), we found that the inhibitory effect of EHNA plus deoxyadenosine on mitogen-stimulated PBMC could not be prevented by IL-2. These results suggest that the immunodeficiency in ADA deficiency includes the absence or failure of a subset of T cells to make IL-2 and the failure of the CD8-positive subset to respond to IL-2. Also, the in vitro cell model of ADA deficiency using EHNA as the ADA inhibitor is limited in its use in understanding the pathogenesis of this disease.     

Deficiency of purine nucleoside phosphorylase activity in thymocytes from the immunodeficient diabetic BB rat.

The spontaneously diabetic BB (BBd) rat displays marked T lymphopenia. The present study was designed to investigate whether the immunodeficiency in this animal may be associated with deficiency of purine nucleoside phosphorylase (PNP) and possibly adenosine deaminase (ADA). The activities of these two enzymes were measured in lymphoid and non-lymphoid cells from both non-diabetes-prone (BBn) and BBd rats as well as from streptozotocin-induced diabetic (STZ) BBn rats. There were no significant differences between BBn and BBd rats in ADA activities in thymocytes, skeletal muscle or brain. However, ADA activity was increased (P less than 0.01) by 50% in BBd mesenteric lymph node lymphocytes and splenocytes as compared with BBn cells, but was not altered in cells from STZ-BBn rats. On the other hand, the PNP activity in BBd thymocytes was only 61% (P less than 0.01) of that observed in BBn cells. This PNP deficiency was not the consequence of diabetes per se, as its activity was normal in thymocytes from STZ-BBn rats. There were no significant differences in PNP activities between BBn and BBd rats in all other cell types examined. The diabetic BB rat may be a novel source of PNP-deficient thymocytes (mainly immature T cells) for studying biochemical mechanisms of immunodeficiency in association with decreased PNP activity. The findings also raise the question of whether a causal relationship exists between PNP deficiency and the recently demonstrated abnormality in T cell maturation in the thymus of the BBd rat.     

Activities of purine nucleotide metabolizing enzymes in human thymocytes and peripheral blood T lymphocytes: in vitro effect of thymic hormones

Adenosine deaminase (ADA, E.C. 3.5.4.4) and purine nucleoside phosphorylase (PNP, E.C. 2.4.2.1) activities are essential for the normal development and function of T lymphocytes. A comparison of the specific activity of these two enzymes and of ecto-5'-nucleotidase (5'-N, E.C. 3.1.3.5) of human thymocytes with that of peripheral T lymphocytes of children and young adults shows that significant differences exist between the activities of ADA and 5'-N in thymocytes and peripheral T cells. ADA activity is six-fold higher in thymocytes than in peripheral T cells whereas 5'-N activity is approximately one-fourth lower in thymocytes than in T lymphocytes. PNP activity is two-fold higher in T cells. The apparent Km and Vmax values for the three enzymes in thymocytes and peripheral T lymphocytes have been determined. The observed differences in enzyme activity are probably not entirely due to differences in the affinity of the enzymes for their substrates. The enzyme activities of a thymoma removed from a patient with myasthenia gravis had intermediate enzyme levels for ADA and PNP, but 5'-N was similar to peripheral T cells. Exposure of thymocytes in short-term culture to the thymic hormones thymopoietin (the pentapeptide TP5), thymosin fraction V or serum thymic factor (FTS) had no significant stimulatory or inhibitory effect on the activities of the three enzymes under our experimental conditions.     

Enzymes of purine metabolism in human peripheral lymphocyte subpopulations.

Ecto-5'nucleotidase (5'NT), adenosine deaminase (ADA), purine nucleoside phosphorylase (PNP) and deoxycytidine (CdR), deoxyguanosine (GdR), deoxyadenosine (AdR) and adenosine (AR) kinases have been measured in subpopulations of peripheral blood lymphocytes of eight healthy volunteers. The separation of B, T, T helper/inducer and T suppressor/cytotoxic cells was performed by means of density gradient centrifugation, E rosetting, passage through a nylon-wool column and antibody affinity chromatography utilising OKT8 and OKT4 monoclonal antibodies. ADA was significantly higher in T lymphocytes and 5'NT in B lymphocytes. Among T cell subpopulations, 5'NT activity was significantly higher (P less than 0 . 01) in T suppressor/cytotoxic (OKT8+) cells (32 . 9 units/10(6) cells than in T helper/inducer (OKT4+) cells (9 . 7 units/10(6) cells). Indeed, the 5'NT activity in T suppressor cells was similar to that in B cells. T helper cells tended, however, to have higher PNP and ADA activities than T suppressor cells but the differences were not statistically significant. No major differences were noted in kinase activities between any of the lymphocyte subpopulations.     

Decreased 5'-nucleotidase activity in suppressor (OKT8) T lymphocytes from homosexuals with AIDS-related complex: nonassociation with enhanced deoxynucleoside toxicity

The purine metabolic enzymes adenosine deaminase (ADA), purine nucleoside phosphorylase (PNP), and 5'nucleotidase (5NT) have been shown to be important for normal lymphocyte maturation. Abnormalities of these enzymes have been associated with hereditary as well as acquired immunodeficiency states. Enzyme activity was measured in helper (OKT4) and suppressor (OKT8) lymphocyte subsets from 10 homosexuals with AIDS-related complex (ARC) and in 10 healthy controls. There were no significant differences in either mean ADA activity or mean PNP activity between ARC OKT4 cells and control OKT4 cells and between ARC OKT8 cells and control OKT8 cells. By contrast, mean 5NT activity was slightly decreased in OKT4 cells from ARC patients compared with that of controls and more significantly diminished in ARC OKT8 cells compared with that of controls. Both deoxyadenosine and deoxyguanosine, when incubated separately with OKT4 and OKT8 cells in the presence of EHNA, an ADA inhibitor, did not significantly inhibit lymphocyte blastogenesis to a greater extent in ARC patients than in controls. Hence, the decreases in 5NT activity most likely reflect lymphocyte immaturity and are not associated with biochemical abnormalities leading to increased deoxynucleoside toxicity.     

Differences in the activity of adenosine deaminase and of purine nucleoside phosphorylase and in the sensitivity to deoxypurine nucleosides between subpopulations of mouse thymocytes

The activities of adenosine deaminase (ADA) and of purine nucleoside phosphorylase (PNP) were measured in thymocyte subpopulations separated by peanut agglutinin (PNA), in unseparated thymocytes, in lymph node and in spleen cells. The PNA+ thymocyte subpopulation exhibited the highest ADA activity of all cells studied. The lowest PNP activity was found in the PNA- subpopulation of thymocytes. PNA+ cells, moreover, exhibited a more intensive DNA synthesis than the PNA- cells, and a greater sensitivity to deoxyadenosine toxicity in the presence of erythro-9-(2-hydroxy-3-nonyl)-adenine (EHNA). The two thymocyte subpopulations exhibited a similar sensitivity to deoxyguanosine (dG) toxicity.     

Protection by various deoxynucleosides against deoxyadenosine-induced DNA damage in adenosine deaminase-inactivated lymphocytes

Adenosine deaminase deficiency is an inborn error resulting in immunodeficiency. The pathogenesis of the lymphopenia is not fully understood. Intracellular increases in dATP in the absence of deamination retard DNA repair in human resting lymphocytes and results in the slow accumulation of DNA strand breaks. We focused on the relationship between DNA damage and DNA precursor pools in cultures of deoxycoformycin-treated, ADA-inhibited resting lymphocytes. The addition of 10 microM deoxyadenosine led to a substantial number of DNA strand breaks within 12 h, breaks equivalent to those which occur with about 190 rad irradiation. Addition of any of the other deoxynucleosides used partially prevented this dAdo-induced DNA damage and promoted DNA repair. However, the preventive effects did not correlate inversely with intracellular dATP levels. Resting lymphocytes have very small dNTP pools. Treatment with dAdo slightly reduced dTTP and dCTP. Three kinds of deoxynucleosides, other than dAdo, restored or raised the corresponding dNTP level but the pool imbalance was only minimally corrected. Regarding the toxic effects of dAdo in ADA deficiency, not only dATP levels but also dNTP pool balance has a crucial role in the pathogenesis. Pool sizes of dTTP, dCTP, and possibly dGTP must be maintained at normal levels, if dAdo-induced DNA damage is to be avoided.     

Do immature T cells accumulate in advanced age?

The hypothesis that decreased T cell function in the elderly involves an increased number of less differentiated T cells was examined. Three markers known to change during thymocyte development were analyzed; ratio of adenosine deaminase (ADA) to purine nucleoside phosphorylase (PNP), lactate dehydrogenase (LD) H/M subunit ratios and the T cell associated antigens, T3, T4, T8 and T10. Cells tested were from 10 old (greater than 75 years) and 10 young (less than 35 years) persons with equal numbers of males and females in each group. Before analysis, cells were purified into three groups; unfractionated, and monocyte depleted T cell and B cell enriched populations. Results for ADA/PNP ratios showed no significant differences between old and young in any of the fractions analyzed. H/M ratios however, were significantly reduced in all three fractions from old donors when compared with young. Surface marker distribution pattern as illustrated by the T3 - (T4 + T8) difference was lower in samples from old donors but not significantly so. There was a very significant reduction in percent cells positive for T3 in all three fractions from old persons. Although some of the changes seen in these markers could be due to a failure of normal differentiation, they could also be caused by the general phenomenon of altered gene expression known to occur with advanced age in a variety of non-lymphoid cells. The absence of any difference in the ADA/PNP ratio suggests that T cell dysfunction in the elderly may not be due to increased numbers of less differentiated cells as a result of thymic involution.     

Functional and mechanistic studies on the toxicity of deoxyguanosine for the in vitro proliferation and differentiation of human peripheral blood B lymphocytes

Deoxyguanosine (dGuo) has been implicated as the toxic metabolite causing a severe impairment of cellular immunity in children with a genetic deficiency of purine nucleoside phosphorylase (PNP). In peripheral blood T cells of normal donors both the pathway which leads to phosphorylation of dGuo (ultimately resulting in deoxyguanosine triphosphate, dGTP) and the salvage pathway which starts with degradation of dGuo by PNP (resulting in the formation of guanosine triphosphate, GTP) contribute to the inhibition of proliferation. In normal peripheral blood B cells, addition of dGuo leads to an inhibition of proliferation and differentiation. The concentrations of dGuo needed to cause a 50% inhibition are equivalent for peripheral blood T cells and B cells. Inhibition of B cell differentiation can be observed at the level of intracytoplasmic as well as secreted Ig and concerns all Ig isotypes. The early phase of B cell activation which takes place during a 24-h preculture with formalinized Cowan I Staphylococci is not affected by dGuo; it is not until proliferation and differentiation of B cells, brought about by culturing in the presence of crude concanavalin A supernatant, occurs that inhibitory effects of dGuo become evident. Addition of dGuo to B cell cultures results in an intracellular accumulation of GTP and dGTP. Addition of 8-aminoguanosine, a PNP inhibitor, next to dGuo, completely prevents the dGuo-mediated inhibition. Under these circumstances the dGuo-mediated increase in intracellular GTP is abrogated while dGTP accumulation still occurs. This indicates that the inhibitory effect of dGuo on the proliferation and differentiation of peripheral blood B lymphocytes of normal donors is independent of dGTP accumulation.     

Heterogeneity of biochemical, clinical and immunological parameters in severe combined immunodeficiency due to adenosine deaminase deficiency.

There was considerable heterogeneity of the biochemical, clinical and immunological findings in 12 patients and two fetuses from 16 kindreds affected by severe combined immunodeficiency (SCID) due to a complete deficiency of the enzyme adenosine deaminase (ADA). Despite this heterogeneity a consistent pattern was observed, in which levels of abnormal purine metabolites paralleled the severity of the immunodeficiency. A high level of urinary deoxyadenosine was a universal finding for homozygous ADA deficiency. ATP depletion, in association with raised deoxy-ATP (dATP) levels, was found in the erythrocytes of nine infants with profound cellular and humoral immunodeficiency. There was no erythrocyte ATP depletion in two patients with some residual immunity, who presented later, but adenosine accumulated in their plasma and urine. This finding, together with the presence of some T and normal B-lymphocytes in less severely affected patients, suggests that adenosine is relatively non-toxic. The other results are consistent with the hypothesis that the sequence of deoxyadenosine accumulation, dATP formation and ATP depletion represents the major mechanism of toxicity to the immune system. Low numbers of T lymphocytes and dATP accumulation were also found in the blood of affected fetuses at 18 weeks gestation. Since extreme instability of erythrocyte ADA was demonstrated in some heterozygotes, and heterozygote ADA levels were detected in one infant with SCID, simultaneous immunological and biochemical analysis of fetal blood are important for precise antenatal diagnosis.     

Enzymes of purine metabolism in lymphoid neoplasms,clinical relevance for treatment with enzyme inhibitors

Summary A few enzymes of the purine degradative pathway have proved valuable in diagnosis and treatment of lymphomas and lymphocytic leukemia. Of particular interest are the enzymes adenosine deaminase (ADA), purine nucleoside phosphorylase (PNP) and ecto-5-nucleotidase (5NT). Intact activities of ADA and PNP have been shown to be vital for lymphoid cells. During development, lymphoid precursors go through remarkable changes in the concentrations of these enzymes and the neoplasma derived from them show a frozen biochemical profile similar to the corresponding normal cell of origin. Knowledge of the role of these enzymes has led to the pharmacological use of enzyme inhibitors for the specific treatment of lymphoid neoplasms. This review concerns the enzymatic make-up of normal and neoplastic lymphocytes and exploitation of this knowledge for the treatment of lymphomas. Special emphasis will be put on the clinical use of an ADA-inhibitor, deoxycoformycin.Abbreviations ADA Adenosine deaminase - ALL Acutelymphocytic leukemia - CdA 2-Chloradeoxyadenosine - CLL Chronic lymphocytic leukemia - dATP Deoxyadenosine triphosphate - dCTP Deoxycytidine triphosphate - dGTP Deoxyguanosine triphosphate - 5NT Ecto-5-nucleotidase - PNP Purine nucleoside phosphorylase - SAH S-adenosylhomocysteine - SAM S-adenosyl-methionine - SCID Severe combined immunodeficiency disease Dedicated to Professor Werner Hunstein on the occasion of his 60th birthday     

In vitro modulation of purine enzyme metabolism and lymphocyte surface marker expression by thymosin fraction 5 in homosexual males

Thymosin fraction 5 (Thymosin) has numerous immunoregulatory activities including modulation of enzymes involved in lymphocyte maturation. The effect of Thymosin on the purine metabolic enzymes adenosine deaminase (ADA), purine nucleoside phosphorylase (PNP), and 5' nucleotidase (5'NT) in null and T-enriched peripheral blood lymphocytes from sexually active asymptomatic homosexual males (AS), patients with the AIDS-related symptom complex (ARC), and those with acquired immune deficiency syndrome (AIDS) was examined and compared to its effect on lymphocytes from healthy heterosexual controls. Mean ADA activity was significantly higher in null cells from fourteen AIDS patients than in five asymptomatic homosexuals, ten ARC patients, or 27 controls. Mean PNP activity was significantly elevated in null-enriched lymphocytes from ten ARC and fourteen AIDS patients compared to controls. No differences in these enzymes were found in T-enriched cells from any group. 5'NT was markedly decreased in both null and T lymphocytes in all homosexual groups relative to controls. Homosexuals had significantly elevated percentages of OKT10 positive and Ia positive lymphocytes compared to controls. Thymosin at an optimal concentration of 150 micrograms/ml caused significant decreases in mean ADA and PNP activity in null lymphocytes from ARC + AIDS patients along with a significant decrease in the percentage of OKT10 positive lymphocytes. No phenotypic changes were seen in AS or control lymphocytes. The data suggest that Thymosin has a maturational effect in vitro on immature T cells from symptomatic homosexuals.     

Inherited disorders of purine metabolism — Underlying molecular mechanisms

Summary An overview of inherited disorders of purine metabolism, concentrating on well established enzyme defects is given. Included are HPRT and the LNS, APRT and 2,8-dihydroxyadenine lithiasis, hyperactivity of PRPP synthetase, ADA and PNP and immunodeficiencies. Emphasis is put on underlying molecular mechanisms on the gene-, enzyme-, or metabolite level for a better understanding of the events leading from the genotype to the clinical phenotype. Finally some aspects of extracellular purine nucleotide metabolism catalyzed by cell surface-bound ectoenzymes are discussed.Abbreviations Ado adenosine - AMP adenosine monophosphate - ADP adenosine diphosphate - ATP adenosine triphosphate - Guo guanosine - GMP guanosine monophosphate - GDP guanosine diphosphate - GTP guanosine triphosphate - Ino inosine - IMP inosine monophosphate - XMP xanthosine monophosphate - AMP-S adenylosuccinate - dIno deoxyinosine - dAdo deoxyadenosine - dATP deoxyadenosine triphosphate - dGuo deoxyguanosine - dGTP deoxyguanosine triphosphate - dCTP deoxycytidine triphosphate - dTTP thymidine triphosphate - Rib-5-P ribose-5-phosphate - dRib-1-P deoxyribose-1-phosphate - 2,3-DPG 2,3-diphosphoglycerate - PRPP 5-phosphorylribose-1-pyrophosphate - SAM S-adenosylmethionine - SAH S-adenosylhomocysteine - P_i inorganic phosphate - PP_i inorganic pyrophosphate - APRT adenine phosphoribosyl-transferase - HPRT hypoxanthine-phosphoribosyltransferase - ADA adenosine deaminase - PNP purine nucleoside phosphorylase - PRA phosphoribosyl amidotransferase - XOD xanthine oxidase - ATPase adenosine triphosphatase - ADPase adenosine diphosphatase - SAHH S-adenosylhomocysteine-hydrolase - BamH 1 restriction endonuclease BamH 1 - Taq 1 restriction endonuclease Taq 1 - K_M Michaelis constant - CRM cross-reacting material - cDNA complementary DNA - LNS Lesch-Nyhan syndrome Parts of this review were presented at the 42nd European Atherosclerosis Group Meeting, Munich, 5–6 March 1984     

Inhibition of Lymphocyte-Mediated Cytolysis and Cyclic amp Phosphodiesterase by Erythro-9–(2-Hydroxy-3-Nonyl)Adenine

Abstract

The adenosine deaminase (ADA) inhibitor erythro-9–(2-hydroxy-3-nonyl)adenine (EHNA), at low concentrations (<10 µM), enhances the inhibitory activity of adenosine against lymphocyte-mediated cytolysis (LMC) without itself being inhibitory. At higher concentrations, EHNA alone is inhibitory to LMC with an IC₅₀ of 160 µM. This inhibition is reversible upon washout, appears to affect an early stage of the lytic process, and does not appear to involve changes in basal levels of cyclic AMP (cAMP), ribonucleoside 5′-triphosphate pool sizes, S-adenosylhomocysteine levels, or protein carboxy-methylation. EHNA does enhance the cAMP response of cytolytic lymphocytes (CL) to activators of adenylate cyclase such as prostaglandin E_l. EHNA inhibits lymphocyte high-affinity cAMP phosphodiesterase at immunosuppressive levels, exhibiting hyperbolic mixed-type inhibition (K_i = 83 µM, α = 0.47, β = 0.18). Whereas inhibition of intralymphocytic ADA is complete at low concentrations (<25 µM) of EHNA, inhibition of LMC and intralymphocytic cAMP phosphodiesterase increases linearly with EHNA concentration to at least 200 µM. The presence of 200 µM EHNA during the centrifugation of mixtures of CL and EL4 leukemia target cells leads to increased CL cAMP levels. 2′-Deoxycoformycin, a more potent ADA inhibitor than EHNA, is not inhibitory to LMC and shows none of these cAMP-related effects. These results suggest that CL-target cell contact stimulates adenylate cyclase in the CL and that EHNA inhibits LMC due to its enhancement of this target cell-stimulated elevation of cAMP.     

Adenosine deaminase and purine nucleoside phosphorylase activity in spleen cells of aged mice

The activities of two enzymes of purine metabolism, adenosine deaminase (ADA) and purine nucleoside phosphorylase (PNP), were determined in spleen lymphocytes from mice of various ages. We found that in the older animals, which have depressed responses to concanavalin A and phytohemagglutinin, there is a decrease in the activity of PNP but normal activity of ADA. The decline of PNP activity was seen at 7.5 months of age and appears to be concurrent with a decline in T-cell function. These results suggest that a decrease in PNP activity may be a contributing factor in the immunodeficient state of the aged.     

Lymphocyte dysfunction caused by deficiencies in purine metabolism

Two inborn errors of purine metabolism have been associated with autosomally inherited human immunodeficiency diseases. A lack of adenosine deaminase (ADA) produces severe lymphopenia and a combined immunodeficiency syndrome. A deficiency of purine nucleoside phosphorylase (PNP) is associated with a selective cellular immune depth. This article discusses the probable biochemical basis for lymphocyte-specific toxicity in these disorders.     

Activities of purine metabolising enzymes in lymphocytes of neonates and young children: correlates with immune function

Depressed activities of the following purine enzymes have been shown to result in immunodeficiencies: adenosine deaminase (ADA), hypoxanthine-guanine phosphoribosyltransferase (HGPRT), and purine nucleoside phosphorylase (PNP). These enzymes and adenosine kinase (AK) were measured in cord blood lymphocytes of premature and small-for-gestational age infants since they have partial immunodeficiencies of unknown biochemical etiology which can persist for many years. We also measured these enzymes in 3 infants with various immunodeficiencies. Activities were compared with appropriate matched control groups. The results indicated normal ADA and PNP but significantly depressed AK (P less than 0.05) and HGPRT (P less than 0.001) activities in 10 premature/SGA infants when compared to 35 full-term normal infants. In the 3 immunodeficient children the results were as follows: Child 1 had a 2- to 3-fold decrease in ADA with normal PNP and AK activities; Child 2 had a 2- to 3-fold decrease in AK, 4-fold decrease in HGPRT with normal PNP and ADA activities; Child 3 had confirmed AIDS and a 4-fold decrease in ADA, 6-fold decrease in HGPRT with normal PNP activity. The possible role of these depressed purine enzyme activities found in lymphocytes is discussed in relation to the imparied immunity seen in these infants.     

Somatic mosaicism caused by monoallelic reversion of a mutation in T cells of a patient with ADA-SCID and the effects of enzyme replacement therapy on the revertant phenotype

Patients with adenosine deaminase (ADA) deficiency exhibit spontaneous and partial clinical remission associated with somatic reversion of inherited mutations. We report a child with severe combined immunodeficiency (T‐B‐ SCID) due to ADA deficiency diagnosed at the age of 1 month, whose lymphocyte counts including CD4+ and CD8+ T and NK cells began to improve after several months with normalization of ADA activity in Peripheral blood lymphocytes (PBL), as a result of somatic mosaicism caused by monoallelic reversion of the causative mutation in the ADA gene. He was not eligible for haematopoietic stem cell transplantation (HSCT) or gene therapy (GT); therefore he was placed on enzyme replacement therapy (ERT) with bovine PEG‐ADA. The follow‐up of metabolic and immunologic responses to ERT included gradual improvement in ADA activity in erythrocytes and transient expansion of most lymphocyte subsets, followed by gradual stabilization of CD4+ and CD8+ T (with naïve phenotype) and NK cells, and sustained expansion of TCRγδ+ T cells. This was accompanied by the disappearance of the revertant T cells as shown by DNA sequencing from PBL. Although the patient’s clinical condition improved marginally, he later developed a germinal cell tumour and eventually died at the age of 67 months from sepsis. This case adds to our current knowledge of spontaneous reversion of mutations in ADA deficiency and shows that the effects of the ERT may vary among these patients, suggesting that it could depend on the cell and type in which the somatic mosaicism is established upon reversion.