Biochemical Pharmacology. Vol. 36. No. 11. pp. 17654768. 1987. GUO6-295@7$3.00 + 0.00 Printed in Great Britain. Per~amon Journals Ltd. RAPID COMMUNICATION zyxwvutsrqponmlkjihgfedcbaZYXW INITIAL STUDIES ON THE CELLULAR PHARMACOLOGY OF 2'.,3'-DIDEOXYADENOSINE, AN INHIBITOR OF HTLV-III INFECTIVITY David A. Cooney*, Gurpreet Ahluwalla*, Hiroaki Mltsuya+. Arnold Fridland*, Mark Johnson*, Zhang Hao*, Maha Dalal*. Jan Balzarinit, Samuel Broder+ and David 6. Johns*5 *DevelopmentalTherapeutics Program and +Clinical Oncology Program Division of Cancer Treatment, Natlonal Cancer Institute, NIH Bethesda, MD 20892, U.S.A. 'Division of Bfochemicaland Clinical Pharmacology St. Jude Children's Research Hospital, Memphis, TN 38101, U.S.A. (Received Y March 1987; accepted 19 March 1987) A number of 2',3'-dideoxynucleosides and related compounds inhibit the in vitro infec- _- tivity and cytopathic effect of the HTLV-III (HIV) retrovirus (11; this virus is the etio- logic agent of the acqufred innnunodeficlency syndrome (AIDS) (2,31. Of the 2',3'-dideoxy- nucleosides studied to date, the most potent in the ATH8 test system is 2',3'-dideoxycytf- dine (ddCyd1 (11. Superior to ddCyd in terms of therapeutic index, however, is the purine nucleoside analogue 2',3'-dideoxyadenoslne (ddAdo): whlle the concentration of ddAdo requ- Ired for maximal anti-HTLV activity in a 7-day assay is some 20-fold higher than that for ddcyd (i.e. 10 UM vs 0.5 PM), ddAdo becomes cytotoxic to ATH8 cells only at >2OD IM, vs >5 PM for ddCyd (1). In thfs respect, ddAdo is also superior to the agent 3'-azido-3'- deoxythymidine (AZT), which has a therapeutic ratio of&?. 5 In the ATH8 test system (41. In view of the favorable therapeutic index of ddAdo, we have examined some of its pharmacb- logical properties and, in particular, its ability to generate the corresponding 5'-nucleo- side triphosphate (ddATP1, putatively (by analogy to ddCTP and AZTTP) the active form of the drug. Initial studies were carried out with ddAdo labeled with tritium in the 2 and 8 posi- tions of the purine base (Moravek Biochemicals.specific activity 56 Ci/mmol, radiochemical purity > 99%). The cell line utilized was an HTLV-III/HIV-sensftive DXT4+ T-cell clone (ATH81, selected on the basis of its rapid growth (in the presence of fnterleukin 21 and sensitivity to the cytopathic effects of the virus (11. ATH8 cells (2 x lo7 cells) were ex- posed to HTLV-III at a dose of 3000 virus partfcles per cell; 24 hr later,C2,8-3HlddAdo was added at a drug concentration of 1 #l, a level sufficient to result in detectable, al- though submaximal, inhibition of the cytopathic effect, infectivfty and replication of the virus and without cytotoxic effect on the host cells (1). The incubation was terminated after 24 hr of drug exposure, and the methanol:aceticacid (pH 41 fraction of the cell lysate was resolved on HPLC, utilizing a radial compressfon column of Partisil-10 SAX with a gradient of ammonium phosphate (Fig. 11. It was observed that ddAdo, In marked contrast to ddCyd (51, was catabolized extenslve- ly under these conditions. In a typical experiment, only a low level of the dideoxyrlbo- nucleotide ddATP (0.037 PM) was detected, and the predominant (>98%1 radiolabeledmetabo- lites were ribonucleotidescoeluting with ADP and ATP (Fig. 1). This result is in general agreement with the early observation of Plunkett and Cohen (61 that ddAdo is readily converted to 2',3'-dideoxyinosfne (ddIno1 by adenosine deaminase, and the more recent 'Author to whom reprint requests should be sent. 1765