9-(2-phosphonylmethoxyethyl) derivatives of purine nucleotide analogs: A comparison of their metabolism and interaction with cellular DNA synthesis

Incubation of CEM cells for 24 h with the guanine, 2,6-diaminopurine, and a denine nucleotide analogs of the 9-(2-phosphonylmethoxyethyl) series, 9-(2- phosphonylmethoxyethyl)guanine (PMEG), 9-(2- phosphonylmethoxyethyl)-2,6-di aminopurine (PMEDAP), and 9-( 2-phosphonylmethoxyethyl) adenine (PMEA), was found to inhibit DNA synthesis 50% at concentrations of 1, 6, and 25 mu M, respectively. Possible reasons for the marked differences were investigate d, including cellular transport of the analogs, different efficiencies of i ntracellular phosphorylation, differential effects on 2'-deoxynucleotide (d NTP) pools, and differences in the affinities of the cellular DNA polymeras es for the diphosphate derivatives of the drugs. No significant differences in cellular uptake were found among the analogs; however, they did differ in the efficiency of phosphorylation, i.e., CEM cells were found to accumul ate higher levels of PMEG-diphosphate (PMEGpp) than PMEDAP-diphosphate (PME DAPpp) or PMEA-diphosphate (PMEApp). Treatment of cells with any of the nuc leotide analogs resulted in increased dNTP pools, with PMEG producing the g reatest increase. All three analogs had the greatest effect on the dATP poo l size, whereas the dGTP pool size was not significantly affected. Comparis on of the ratios of nucleotide analog diphosphates to their corresponding d NTPs under conditions where DNA synthesis is inhibited 50% suggested that c ellular DNA polymerases were approximately twice as sensitive to PMEGpp tha n to PMEDAPpp and 5-fold more sensitive to PMEGpp than to PMEApp. Consisten t with this hypothesis, examination of the efficiencies with which the repl icative DNA polymerases alpha, delta, and epsilon incorporated the analogs showed that DNA polymerase delta, the most sensitive of the DNA polymerases , incorporated PMEGpp twice as efficiently as PMEDAPpp and 7-fold more effi ciently than PMEApp.