Telomerase protein rather than its RNA is the target of phosphorothioate-modified oligonucleotides

Human telomerase is a ribonucleoprotein which uses its internal RNA moiety as a template for telomeric DNA synthesis. This enzyme is up-regulated in m ost malignant tumors and is therefore considered as a possible cancer targe t. Here we examined the effects of differently modified oligomers on telome rase activity from HL-60 cell extracts (TRAP-eze(TM) assay), Phosphorothioa te-modified oligonucleotides (PS-ODNs) inhibited telomerase activity at sub nanomolar concentrations and proved to be more efficient than peptide nucle ic acids. In contrast to all the investigated oligomers, PS-ODNs were found to bind to the protein motif of telomerase called the primer binding site but poorly to its RNA, This is suggested by kinetic investigations demonstr ating a competitive interaction of PS-ODNs and TS primer at the primer bind ing site. The K-m value of the TS primer was 10.8 nM, the K-i value of a 20 mer PS-ODN was 1.6 nM, When the TS primer was PS-modified a striking increa se in the telomerase activity was found which correlates with the number of phosphodiesters replaced. The K-m value of a completely PS-modified TS pri mer was 0.56 nM, Based on these results the design of chimeric ODNs is prop osed consisting of a 5'-PS-modified part targeting the primer binding site and a 3'-terminus part targeting the telomerase RNA.