We treated two children who had adenosine deaminase deficiency and severe combined immunodeficiency disease by injecting bovine adenosine deaminase modified by conjugation with polyethylene glycol. The modified enzyme was rapidly absorbed after intramuscular injection and had a half-life in plasma of 48 to 72 hours. Weekly doses of approximately 15 U per kilogram of body weight maintained plasma adenosine deaminase activity at two to three times the level of erythrocyte adenosine deaminase activity in normal subjects. The principal biochemical consequences of adenosine deaminase deficiency were almost completely reversed. In erythrocytes, adenosine nucleotides increased and deoxyadenosine nucleotides decreased to less than 0.5 percent of total adenine nucleotides. The activity of S-adenosylhomocysteine hydrolase, which is inactivated by deoxyadenosine, increased to normal in red cells and nucleated marrow cells. Neither toxic effects nor hypersensitivity reactions were observed. In vitro tests of the cellular immune function of each patient showed marked improvement, along with an increase in circulating T lymphocytes. Clinical improvement was indicated by absence of infection and resumption of weight gain. We conclude that from the standpoints of efficacy, convenience, and safety, polyethylene glycol-modified adenosine deaminase is preferable to red-cell transfusion as a treatment for adenosine deaminase deficiency. Patients with other inherited metabolic diseases in which accumulated metabolites equilibrate with plasma could benefit from treatment with the appropriate polyethylene glycol-modified enzyme.
S-Adenosylhomocysteine (AdoHcy) hydrolase regulates all adenosylmethionine-(AdoMet) dependent transmethylations by hydrolyzing the potent feedback inhibitor AdoHcy to homocysteine and adenosine. The crystallographic structure determination of a selenomethionyl-incorporated AdoHcy hydrolase inhibitor complex was accomplished using single wavelength anomalous diffraction data and the direct methods program, Snb v2.0, which produced the positions of all 30 crystallographically distinct selenium atoms. The mode of enzyme-cofactor binding is unique, requiring interactions from two protein monomers. An unusual dual role for a catalytic water molecule in the active site is revealed in the complex with the adenosine analog 2'-hydroxy, 3'-ketocyclopent-4'-enyladenine.
Polyethylene glycol (PEG)-modified bovine adenosine deaminase (ADA) is used for replacement therapy of severe combined immunodeficiency disease due to inherited ADA deficiency. We monitored IgG anti-ADA antibody in 17 patients treated by intramuscular injections of PEG-ADA for 1 to > 5.5 yr. ELISA-detectable anti-ADA IgG appeared in 10 patients, usually between the third and eighth months oftreatment. Anti-ADA levels did not correlate with trough plasma ADA activity, which averaged 1.8-5 times normal blood (erythrocyte) ADA activity, depending on dose (15-60 U/kg per wk). ELISA-detectable anti-ADA antibodies were directed primarily at bovine-specific peptide (rather than PEG-containing) epitopes. Enhanced enzyme clearance, mediated by antibody that directly inhibited native and PEG-modified bovine ADA, and native, but not PEG-modified human ADA, occurred in two patients. In one, tolerance was induced; in the second, twice weekly injections of PEG-ADA compensated for accelerated clearance. We speculate that inhibitory antibodies recognize conserved, relatively PEG-free epitope(s) encompassing the active site, and that in human, but not bovine, ADA a PEG-attachment site "shields" the active site from immune recognition. We conclude that PEG-modification largely prevents the development of high affinity, or high levels of, clearing antibodies to bovine ADA, and that PEG-modified human ADA should be further investigated as a possible treatment for ADA deficiency.
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