Actively dividing cells show progressive loss of telom-eric DNA during successive rounds of replication due to end-replication problem. Telomere shortening has been proposed as a regulatory mechanism that con-trols the replicative capacity of primary cells before undergoing cellular senescence. In immortal cells in-cluding cancer, cellular senescence can be overcome by reactivation of telomerase or by a telomerase-independent mechanism for lengthening telomeres. In this work, we present a novel example of telomere elongation mechanism in a human stomach adenocar-cinoma cell line which was selected for resistance to adriamycin. The resistant cell line (MKN/ADR) had long terminal restriction fragments (TRFs) of up to ~50 kb, while its parent cell line (MKN-45) had the TRFs, consisting of a smear extending from ~4 to ~25 kb. The very large TRFs in MKN/ADR cell line were proven to be telomeric by digestion with the exonucle-ase Bal31. When telomerase activity was examined using the PCR-based telomeric repeat amplification protocol (TRAP) assay, MKN/ADR cell line showed reduced activity to about 10% of that in MKN-45 cell line. The correlation between reduced telomerase ac-tivity and mRNA expression of telomerase subunits in MKN/ADR cell line was assessed by the reverse tran-scriptase-PCR analysis. The level of human telomerase reverse transcriptase (hTERT) mRNA was lower in MKN/ADR cell line than in MKN-45 cell line. This observation correlates with the finding that telomerase activity is reduced about 10-fold in MKN/ADR cell line. Reverse transcriptase-PCR analysis also revealed a close correlation between telomerase-associated pro-tein (TP1) mRNA expression and telomerase activity in MKN/ADR cell line. In contrast, expression levels of human telomerase RNA (hTR) were identical in both MKN/ADR and MKN-45 cell lines. Taken together, these data suggest that telomeres in MKN/ADR cell line may be regulated through a novel mechanism other than telomerase. Although the basis for telomere elongation mechanism in MKN/ADR cell line is not yet understood, the occurrence of alternative mechanism for telomere elongation in drug-resistant cancer cells may have an important implication for use of telom-erase inhibitors in human cancer treatment.

Keywords: T, Immortalization, Adriamycin-resistant Cell Line