Can inhibition of telomerase increase pancreatic cancer cell's susceptibility to chemotherapeutic reagents?

摘要

Objectives: To clarify the inhibition of pancreatic can-cer cells by interference with the hTR component of thetelomerase reverse transcriptase enzymatic complexand evaluate susceptibility of antisense hTR pancreaticcancer cells to chemotherapeutic reagents.Methods: A 593 bp of full length hTR cDNA was sub-cloned into a mammalian expression vector pcDNA3.1(-) in antisense orientation to construct an antisensehTR expression plasmid. The plasmids were introducedinto Pancl cells, a human pancreatic carcinoma cellline, by lipofectin, and G418-resistant stable trans-formants were expanded. Resulting stable clones werescreened for the presence of hTR insert by PCR withT7 and BGH reverse primers located on the flanks ofthe multiclonal site of pcDNA3.1 vector. Cell growth rate,hTR expression, telomerase activity, and anchorage-in-dependent growth property were analyzed. Finally, sus-ceptibility of antisense hTR cells to chemotherapeuticreagents was evaluated.Results: Significant downregulation of endogenous hTRwas evident in the antisense-hTR transformed cells,and telomerase activity was markedly decreased com-pared to control cells in standard TRAP assays. Fur-thermore, the proliferation and the anchorage-inde-pendent growth ability in antisense-hTR expressingcells were significantly decreased compared with thecontrol parental cells. However, no crisis or senescencephenomena was observed. Antisense hTR appears toincrease Pancl cell’s susceptibility to chemotherapeuticreagent cDDP, but not to differentiation reagent DM-SO, COX2 inhibitor sulinbac, NS-398, curcumin, andchemotherapeutic reagent adriamycin(ADM).Conclusions: These data indicate that hTR is probablya critical component of human telomerase activity andthat downregulation of the RNA component of humantelomerase is an effective target for anticancer strategyand antisense hTR can increase Pancl cell’s susceptibilityto cDDP.