**= .008. The proportion of FoxP3+ cells was also analyzed by FACS analyses for CD4+CD25+FoxP3+, CTLA-4, and CD45RB in the peripheral blood (PB) and lymphoid organs of ADA+/+ and ADA?/? mice. and the underlying mechanisms causing defective peripheral tolerance in ADA-SCID. Trials were registered at www.clinicaltrials.gov as “type”:”clinical-trial”,”attrs”:”text”:”NCT00598481″,”term_id”:”NCT00598481″NCT00598481/”type”:”clinical-trial”,”attrs”:”text”:”NCT00599781″,”term_id”:”NCT00599781″NCT00599781. Introduction CD4+CD25+FoxP3+ regulatory T cells (Tregs) actively suppress pathologic and physiologic immune responses to maintain peripheral immune self-tolerance and prevent autoimmunity.1,2 Several Tigecycline molecules and mechanisms have been shown to operate in Treg-mediated control of immune responses.3 In vitro functional studies have shown that Tregs suppress effector cells via the secretion of immunosuppressive cytokines, by cell contact-dependent Treg/antigen-presenting cells (APCs) or Treg/T-cell interactions, and functional modification or killing of APCs.4 Extracellular adenosine produced by Tregs has been described as one of the mechanisms mediating their suppressive activity. Concordant expression of the ectoenzymes CD39 and CD73 has been reported both for murine and human Tregs.5C7 The CD39 ectoenzyme produces AMP from ATP or ADP, which is subsequently converted into extracellular adenosine by the CD73 ectoenzyme.8 Treg function requires the coordinated expression of the adenosine 2A receptor (Adora2A) on activated T effector cells to enable adenosine-mediated immunosuppression.2 Moreover, human Tregs have been shown to express low levels of adenosine deaminase (ADA), the enzyme responsible for adenosine breakdown. In contrast, T effector cells are enriched in ADA but express low levels of CD39 and CD73.7 This molecular profile of human Treg (CD39+CD73+ADAlow) has functional significance, as it confers Tregs the capability to Tigecycline produce and sustain relatively high concentrations of extracellular adenosine. 7 ADA catalyzes the Tigecycline irreversible deamination of adenosine and deoxyadenosine into inosine and deoxyinosine, respectively. Further conversion of these deaminated nucleosides leads to hypoxanthine, which can be either transformed irreversibly into uric acid or salvaged into mononucleosides.9 In the absence of ADA, adenosine and deoxyadenosine accumulate in plasma and cells, forming the metabolic basis underlying immune cell defects and severe combined immunodeficiency (ADA-SCID). Immunologic defects associated with this disease include impaired T-, B-, NK-cell development and function and complete absence of cellular and humoral immunity.9,10 In ADA-deficient patients, as in other forms of SCID, bone marrow transplantation (BMT) from an HLA-identical sibling donor is the treatment of choice, but available only for a minority of patients.11 The administration of enzyme replacement therapy with pegylated bovine ADA (PEG-ADA) results in clinical improvement with approximately 70% to 80% survival but often incomplete immunologic reconstitution. Recent trials represented the first demonstration of long-term clinical efficacy of hematopoietic stem cell gene therapy (GT) for ADA-SCID, underlining that GT has a favorable safety profile and is Tigecycline effective in restoring purine metabolism and immune function.12 Manifestations of immune dysregulation, autoimmunity, and allergic manifestations are key findings in milder forms of the disease but have also been reported in patients under PEG-ADA treatment.13C16 Little is known on their pathogenesis or to what extent some mechanisms of tolerance might be specifically affected NMDAR2A by ADA deficiency.10 Because peripheral tolerance is primarily mediated by Tregs, which possess a unique biochemical signature among T cells in that they generate and sustain high adenosine concentrations, we hypothesized that the autoimmune manifestations associated with ADA deficiency might be the result of an altered purine metabolism interfering with normal Treg function. Herein we provide evidence for specific defects in the regulatory T-cell compartment of ADA-deficient mice and patients. Furthermore, we show that enzyme replacement treatment with PEG-ADA, but not GT or BMT, severely interferes with normal Treg function. Methods.