Abstract

6-Methylthiopurine ribonucleoside, a cytotoxic nucleoside with antitumor activity, was phosphorylated by cell-free preparations from H.Ep. #2 cells in culture and from yeast. Studies of phosphorylation in the presence of other nucleosides indicated that adenosine kinase was the enzyme catalyzing this phosphorylation and also the phosphorylation of several other cytotoxic nucleosides derived from adenosine by substitution at the 2-position, substitution of other groups for the 6-amino group, or alteration of the ring structure. Cell lines resistant to 6-methylthiopurine ribonucleoside were selected from H.Ep. #2 cells and from a subline resistant to 6-mercaptopurine; a third line resistant to 2-fluoroadenosine was selected from a subline of H.Ep. #2 cells resistant to 2-fluoroadenine. Each of the three resistant cell lines was also resistant to a series of adenosine analogs that were toxic to the parent cell line. The resistant cells were devoid of detectable capacity to phosphorylate 6-methylthiopurine ribonucleoside and adenosine, and this loss persisted when the cells were cultured for many generations in the absence of inhibitor. The resistant cells did not differ from the cell lines from which they were derived in capacity to utilize purines. These results indicate that adenosine kinase is an enzyme essential for the activation of many cytotoxic nucleosides and that loss of this enzyme activity is a mechanism of resistance to these nucleosides.