Abstract 3886: The G-quadruplex induced DNA damage at telomeres is regulated by ATM

Abstract

Abstract Quadruplex-forming DNA sequences are present throughout the genome, including in the telomeres. Our group has shown that quadruplex-forming oligonucleotides selectively inhibit proliferation of transformed cells. One of these oligonucleotides, AS1411, is currently in a Phase IIb clinical trial. We have recently shown that the c-myc promoter quadruplex-forming sequence, Pu-27, also inhibits growth of malignant cells. We have hypothesized that one potential mechanism of this effect is competition for telomere-binding proteins. If this is correct, we would expect this effect to be mediated by the DNA repair response. Telomere integrity is maintained by the shelterin protein complex which is required to prevent chromosomal ends from being recognized as DNA double strand breaks. Like many other DNA damage response proteins, ATM has been shown to play a role in the maintenance of telomere integrity. We have shown that ATM deficient cells are more sensitive to growth inhibition by Pu-27 whereas ATM proficient cells were relatively resistant to Pu-27. Co-immunostaining with γ-H2AX and TRF1 revealed that the sensitivity to Pu-27 in ATM deficient cells was associated with phosphorylation of H2AX, an indicator of DNA double strand break and induction of TIFs (Telomere-dysfunction Induced Foci). There was induction of γ-H2AX and downregulation of shelterin proteins TRF2 and TRF1 in Pu-27 treated ATM deficient cells (as determined by western blotting). U937 leukemia cells, which were sensitive to Pu-27, became relatively resistant when they were stably transfected with a vector expressing a dominant negative mutant of the shelterin protein TRF2 (dn-TRF2). This suggests that Pu-27 may act by altering the telomere-shelterin complex. This idea was further substantiated by the fact that ALT (Alternative Lengthening of Telomere) cells were also relatively resistant to Pu27. A549 cells, which were sensitive to Pu-27, also showed abundant presence of TIF compared to untreated cells. Lastly, we showed that both p53 wild type and deficient colorectal cells (HCT116 cell line) were relatively resistant to Pu-27. Taken together, this data suggests that ATM plays a pivotal role in Pu-27 induced DNA damage at telomere ends, which occurs in response to a Pu-27 induced alteration in telomere-shelterin homeostasis. It is likely that this response is induced in a p53 independent manner. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3886. doi:10.1158/1538-7445.AM2011-3886