Adenosine Deaminase-deficient Mice Generated Using a Two-stage Genetic Engineering Strategy Exhibit a Combined Immunodeficiency

Abstract

Adenosine deaminase (ADA) deficiency in humans leads to a combined immunodeficiency. The mechanisms involved in the lymphoid specificity of the disease are not fully understood due to the inaccessibility of human tissues for detailed analysis and the absence of an adequate animal model for the disease. We report the use of a two-stage genetic engineering strategy to generate ADA-deficient mice that retain many features associated with ADA deficiency in humans, including a combined immunodeficiency. Severe T and B cell lymphopenia was accompanied by a pronounced accumulation of 2-deoxyadenosine and dATP in the thymus and spleen, and a marked inhibition of S-adenosylhomocysteine hydrolase in these organs. Accumulation of adenosine was widespread among all tissues examined. ADAdeficient mice also exhibited severe pulmonary insufficiency, bone abnormalities, and kidney pathogenesis. These mice have provided in vivo information into the metabolic basis for the immune phenotype associated with ADA deficiency.Genetic defects in purine metabolism in humans result in serious metabolic disorders, often with pronounced tissue-specific phenotypes (1). A striking example of this is adenosine deaminase (ADA) 1 deficiency, which results in impaired lymphoid development and a severe combined immunodeficiency disease (SCID) (2). ADA-deficient SCID was the first of the inherited immunodeficiencies for which the underlying molecular defect was identified (3); however, despite over 20 years of subsequent research, a satisfactory explanation for the lymphoid specificity of this metabolic disease has not emerged. This is largely due to the inaccessibility of human tissue for detailed phenotypic and metabolic analysis and the absence of an animal model which retains features of ADA deficiency in humans. The availability of a genetic animal model for ADA deficiency would make possible a wide range of biochemical and immunological experiments that are not permissible with humans. Additional interest in ADA deficiency stems from recent attempts to use novel therapeutic strategies, including enzyme therapy (4) and gene therapy (5, 6), to treat the condition in humans. Although the results of these therapeutic approaches are encouraging, unexpected outcomes have raised numerous important questions regarding the efficacy of specific treatment protocols (4, 7). The pace with which new enzyme and gene therapy protocols can be tested would be greatly increased by the availability of an animal model for ADA deficiency.Successful attempts to generate ADA-deficient mice were initially reported by two groups (8, 9), resulting in animals with independent sites of Ada gene disruption. In each case, a similar phenotype was observed. ADA-deficient fetuses died perinatally due to severe liver damage (8, 9). This phenotype was accompanied by profound disturbances in purine metabolism, including marked increases in the ADA substrates adenosine and 2Ј-deoxyadenosine. Both adenosine and 2Ј-deoxyadenosine are biologically active purines that can have profou...

Metabolites from apoptotic thymocytes inhibit thymopoiesis in adenosine deaminase–deficient fetal thymic organ cultures

“…Conventional ADA-deficient mice die of liver failure in the immediate perinatal period (6,7), and normal mice treated with the specific and potent ADA inhibitor 2′-deoxycoformycin (dCF) (8) show signs of hepatic and adrenal toxicity (9). The problem of liver toxicity in ADA-deficient mice has been solved by the introduction of an Ada minigene expressed in the placenta (10). These mice display profound T-and B-cell lymphopenia and reduced concentrations of serum immunoglobulins by 2 weeks of age.…”

High Levels of Adenosine Deaminase on Dendritic Cells Promote Autoreactive T Cell Activation and Diabetes in Nonobese Diabetic Mice

“…ADA KO mice were available under FVB/N founder background (21,26). To transfer the KO phenotype into NOD mice, a two-step rescue scheme is necessary to produce the adult mice.…”

“…ADA-deficient newborns are further rescued by weekly i.p injection of PEG-conjugated ADA. Previous reports on these strains showed a largely normal phenotype in the peripheral and central immune cells (21,26). To produce ADA KO NOD mice from the current FVB/N background, the minigene and the ADA-deficient allele were bred separately into NOD mice for 10 generations.…”

Impaired Germinal Center Maturation in Adenosine Deaminase Deficiency

“…A mouse model for the human disease is now available. The model allows studies of lymphoid organs and immune function in ADA deficiency that are not accessible in humans (2). Because ADA deficiency in humans has served as a testing ground for enzyme replacement and gene therapies in humans, the mouse model is key in assessing the in vivo pathophysiological relevance of these therapies.…”