Inhibition of human telomerase by 7-deaza-2'-deoxyguanosine nucleoside triphosphate analogs: potent inhibition by 6-thio-7-deaza-2'-deoxyguanosine 5'-triphosphate.
We have examined analogs of the previously reported 7-deaza-2'-deoxypurine nucleoside triphosphate series of human telomerase inhibitors. Two new telomerase-inhibiting nucleotides are reported: 6-methoxy-7-deaza-2'-deoxyguanosine 5'-triphosphate (OMDG-TP) and 6-thio-7-deaza-2'-deoxyguanosine 5'-triphosphate (TDG-TP). In particular, TDG-TP is a very potent inhibitor of human telomerase with an IC(50) of 60 nM. TDG-TP can substitute for dGTP as a substrate for telomerase, but only at relatively high concentrations. Under conditions in which TDG-TP is the only available guanosine substrate, telomerase becomes nonprocessive, synthesizing short products that appear to contain only one to three TDG residues. Similarly, the less potent telomerase inhibitor OMDG-TP gives rise to short telomerase products, but less efficiently than TDG-TP. We show here that TDG-TP, and to a lesser extent OMDG-TP, can serve as substrates for both templated (Klenow exo) and nontemplated (terminal transferase) DNA polymerases. For either polymerase, the products arising from TDG-TP are relatively short, and give rise to bands of unusual mobility under PAGE conditions. These anomalous bands revert, under treatment with DTT, to normal mobility bands, indicating that these products may contain thiol-labile disulfide linkages involving the incorporated TDG residues. This observation of potential TDG-crosslinks may have bearing on the mechanism of telomerase inhibition by this nucleotide analog.
Fletcher
T M
TM
College of Pharmacy, University of Texas, Austin, Texas 78712-1074, USA.
Cathers
B E
BE
Ravikumar
K S
KS
Mamiya
B M
BM
Kerwin
S M
SM
D004256:DNA Polymerase I / Q000037:antagonists & inhibitors; D003848:Deoxyguanine Nucleotides / Q000138:chemical synthesis* / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000138:chemical synthesis* / Q000494:pharmacology*; D006801:Humans; D020287:In Situ Nick-End Labeling; D013329:Structure-Activity Relationship; D019098:Telomerase / Q000037:antagonists & inhibitors*
C445702:6-thio-7-deaza-2'-deoxyguanosine 5'-triphosphate; C445703:7-deaza-2'-deoxyguanosine nucleoside triphosphate; D003848:Deoxyguanine Nucleotides; D004791:Enzyme Inhibitors; D019098:Telomerase; D004256:DNA Polymerase I