Adenosine kinase inhibition promotes survival of fetal adenosine deaminase–deficient thymocytes by blocking dATP accumulation

Thymocyte development past the CD4 -CD8 -stage is markedly inhibited in adenosine deaminase-deficient (ADA-deficient) murine fetal thymic organ cultures (FTOCs) due to the accumulation of ADA substrates derived from thymocytes failing developmental checkpoints.Such cultures can be rescued by overexpression of Bcl-2, suggesting that apoptosis is an important component of the mechanism by which ADA deficiency impairs thymocyte development.Consistent with this conclusion, ADA-deficient FTOCs were partially rescued by a rearranged T cell receptor transgene that permits virtually all thymocytes to pass the -selection checkpoint.ADA-deficient cultures were also rescued by the adenosine kinase inhibitor 5-amino-5-deoxyadenosine (5A5dAdo), indicating that the metabolite responsible for the inhibition of thymocyte development is not adenosine or deoxyadenosine, but a phosphorylated derivative of an ADA substrate.Correction of ADA-deficient FTOCs by 5A5dAdo correlated with reduced accumulation of dATP, implicating this compound as the toxic metabolite.In ADA-inhibited FTOCs rescued with a Bcl-2 transgene, however, dATP levels were superelevated, suggesting that cells failing positive and negative selection continued to contribute to the accumulation of ADA substrates.Our data are consistent with dATP-induced mitochondrial cytochrome c release followed by apoptosis as the mechanism by which ADA deficiency leads to reduced thymic T cell production.