2',3'-Dideoxyadenosine killing of TdT-positive cells is due to a trace contaminant

We have previously reported that the chain-terminating nucleoside analogue 2',3'-dideoxyadenosine (ddA) is specifically cytotoxic for TdT-positive cells in the co-presence of the adenosine deaminase (ADA) inhibitor coformycin (CF). Further studies with ddA/CF revealed that cytotoxicity occurs only if ddA is supplied from the Calbiochem or Fluka companies. ddA supplied from other commercial sources (Pharmacia, Sigma) or the NCI Pharmaceutical Resources Branch is non-cytotoxic. To explore the basis for this difference, ddA from various sources was subjected to reverse-phase high-pressure liquid chromatography (HPLC) analysis. The Calbiochem and Fluka ddA had a unique HPLC peak, with a retention time of 12.8 min, representing a contaminant of less than 0.1% of the bulk material applied to the C-18 HPLC column. Study of all HPLC peaks resolved from the bulk material showed cytotoxic activity in only the 12.8 min peak. To identify the nature of the unknown compound, we compared HPLC characteristics of the synthetic intermediates and byproducts of ddA synthesis to the peak eluting at 12.8 min. Of these, only 3'-deoxyadenosine (cordycepin) had similar HPLC characteristics. In addition, in the co-presence of CF, cordycepin was specifically cytotoxic (IC50 < 0.5 microM) for all TdT-positive cell lines tested. Cytotoxicity was seen in TdT-negative cells only at concentrations 10-100-fold higher. We conclude that our previous report on ddA/CF as a TdT-specific cytotoxic combination was due to contamination of the ddA supplied by Calbiochem by cordycepin. ddA itself is non-cytotoxic for TdT-positive cells. Cordycepin in the co-presence of an ADA inhibitor may be effective in the treatment of TdT-positive hematological malignancies.