Extremely short telomeres and high telomerase activity in T-cell prolymphocytic leukemia (T-PLL): An optimal target for telomerase inhibition with GRN163L?

Abstract

7061 Background: T-cell prolymphocytic leukemia (T-PLL) is an uncommon aggressive lymphoproliferative disease characterized by the expansion of a T-cell clone. We recently reported that leukemic T-PLL cells have extremely short telomeres (n=11, mean ± sd: 1.53 kb ± 0.65 kb) and high levels of telomerase activity (Roeth et al., 2007). We hypothesized that such cells have telomeres that are very close to critically short lengths and thus could be sensitive to telomerase inhibition with high sensitivity and short lag times. Methods: Our aim was to investigate the effect of the selective telomerase inhibitor GRN163L on survival of leukemic cells from T-PLL patients in short-term culture with detection of cell apoptosis by FACS-analysis using Annexin V-PI double staining. Telomere lengths of subsets of leukocytes from the peripheral blood of patients with T-PLL were determined by the automated flow-FISH and telomerase activity was measured by the TRAP-assay. Results: The leukemic cells of these T-PLL patients had short telomere lengths and high telomerase activity. We observed a dose-dependent cytotoxicity in T-PLL cell cultures with GRN163L. Viable cells as a percentage of untreated controls (viability index) after 7 days of cell culture with 0, 1, 3, 10 μM GRN163L were as follows (% mean ± sd, (t-test p-value versus control)): 100 ± 0, 57.3 ± 17.5 (p=0.024), 37.4 ± 25.0 (p=0.022), 25.3 ± 20.0 (p=0.008), respectively, (n=4). No effect on viability was observed in normal, unstimulated T-cells after 7 days with 0, 1, 3, 10 μM GRN163L. Viability indices for normal T- cells were as follows: (% mean ± sd) 100 ± 0.0, 97.9 ± 1.7, 102.4 ± 9.4, 95.1 ± 4.7, respectively, (n=3). In addition, a dose-dependent increase of apoptotic cells with the telomerase inhibitor GRN163L by Annexin V-PI double staining was observed. A mismatch control oligonucleotide at 10 μM had no significant effect on viability of leukemic cells. Conclusions: GRN163L is active in T-PLL cells in vitro and leads to rapid cytotoxicity. Targeting telomerase and telomeres therefore seems an attractive strategy for the future treatment of this disease. These results provide strong rationale for a planned clinical trial of GRN163L in patients with T-PLL. Author Disclosure Employment or Leadership Consultant or Advisory Role Stock Ownership Honoraria Research Expert Testimony Other Remuneration Geron Corporation Geron Corporation Geron Corporation