Targeting telomeres and telomerase (original) (raw)
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Reevaluation of telomerase inhibition by quadruplex ligands and their mechanisms of action
Proceedings of the National Academy of Sciences, 2007
Quadruplex ligands are often considered as telomerase inhibitors. Given the fact that some of these molecules are present in the clinical setting, it is important to establish the validity of this assertion. To analyze the effects of these compounds, we used a direct assay with telomerase-enriched extracts. The comparison of potent ligands from various chemical families revealed important differences in terms of effects on telomerase initiation and processivity. Although most quadruplex ligands may lock a quadruplex-prone sequence into a quadruplex structure that inhibits the initiation of elongation by telomerase, the analysis of telomerase-elongation steps revealed that only a few molecules interfered with the processivity of telomerase (i.e., inhibit elongation once one or more repeats have been incorporated). The demonstration that these molecules are actually more effective inhibitors of telomeric DNA amplification than extension by telomerase contributes to the already growing suspicion that quadruplex ligands are not simple telomerase inhibitors but, rather, constitute a different class of biologically active molecules. We also demonstrate that the popular telomeric repeat amplification protocol is completely inappropriate for the determination of telomerase inhibition by quadruplex ligands, even when PCR controls are included. As a consequence, the inhibitory effect of many quadruplex ligands has been overestimated.
Journal of Biological Chemistry, 2003
Ligands that stabilize the telomeric G-rich singlestranded DNA overhang into G-quadruplex can be considered as potential antitumor agents that block telomere replication. Ligand 12459, a potent G-quadruplex ligand that belongs to the triazine series, has been previously shown to induce both telomere shortening and apoptosis in the human A549 cell line as a function of its concentration and time exposure. We show here that A549 clones obtained after mutagenesis and selected for resistance to the short term effect of ligand 12459 frequently displayed hTERT transcript overexpression (2-6-fold). Overexpression of hTERT was also characterized in two resistant clones (JFD10 and JFD18) as an increase in telomerase activity, leading to an increase in telomere length. An increased frequency of anaphase bridges was also detected in JFD10 and JFD18, suggesting an alteration of telomere capping functions. Transfection of either hTERT or DN-hTERT cDNAs into A549 cells did not confer resistance or hypersensitivity to the short term effect of ligand 12459, indicating that telomerase expression is not the main determinant of the antiproliferative effect of ligand 12459. In contrast, transfection of DN-hTERT cDNA into resistant JFD18 cells restored sensitivity to apoptotic concentrations of ligand 12459, suggesting that telomerase does participate in the resistance to this G-quadruplex ligand. This work provides evidence that telomerase activity is not the main target for the 12459 G-quadruplex ligand but that hTERT functions contribute to the resistance phenotype to this class of agents.
Cancer research, 2003
The molecular mechanisms induced by G-quadruplex ligands to trigger senescence in mammalian cells are still unknown, although the critical role of telomerase is highly suspected. JFA2 cells selected for resistance to senescence induced by the G-quadruplex ligand 12459 presented an overexpression of hTERT transcript that correlated to a functional increase in telomerase activity and telomere length. Consistently, treatment with 12459 failed to trigger senescence and telomere shortening in JFA2 cells. Resistant cells also presented cross-resistance for senescence induction to telomestatin, another G-quadruplex ligand from a different series, but not to other anticancer agents, indicating the selectivity of the resistance mechanism. We, thus, provide evidence that telomerase activity and telomere length are key cellular determinants of the resistance to G-quadruplex ligands.
Telomeres and Telomerase: Pharmacological Targets for New Anticancer Strategies?
Current Cancer Drug Targets, 2006
Telomeres are located at the ends of eukaryotic chromosomes. Human telomerase, a cellular reverse transcriptase, is a ribonucleoprotein enzyme that catalyzes the synthesis and extension of telomeric DNA. It is composed of at least, a template RNA component (hTR; human Telomerase RNA) and a catalytic subunit, the telomerase reverse transcriptase (hTERT). The absence of telomerase is associated with telomere shortening and aging of somatic cells, while high telomerase activity is observed in over 85% of human cancer cells, strongly indicating its key role during tumorigenesis. Several details regarding telomere structure and telomerase regulation have already been elucidated, providing new targets for therapeutic exploitation. Further support for anti-telomerase approaches comes from recent studies indicating that telomerase is endowed of additional functions in the control of growth and survival of tumor cells that do not depend only on the ability of this enzyme to maintain telomere length. This observation suggests that inhibiting telomerase or its synthesis may have additional anti-proliferative and apoptosis inducing effect, independently of the reduction of telomere length during cell divisions. This article reviews the basic information about the biology of telomeres and telomerase and attempts to present various approaches that are currently under investigation to inhibit its expression and its activity. We summarize herein distinct anti-telomerase approaches like antisense strategies, reverse transcriptase inhibitors, and G-quadruplex interacting agents, and also review molecules targeting hTERT expression, such as retinoids and evaluate them for their therapeutic potential.
Inhibition of Human Telomerase by a G-Quadruplex-Interactive Compound
Journal of Medicinal Chemistry, 1997
The ends of chromosomes have specialized sequences, termed telomeres, comprising tandem repeats of simple DNA sequences. Human telomeres consist of the sequence 5′-TTAGGG. 1,2 Telomeres have several functions apart from protecting the ends of chromosomes, the most important of which appear to be associated with senescence, replication, and the cell cycle clock. 3 Progressive rounds of cell division result in a shortening of the telomeres by some 50-200 nucleotides per round. Almost all tumor cells have shortened telomeres, which are maintained at a constant length 4,5 and are associated with chromosome instability and cell immortalization. The enzyme telomerase adds the telomeric repeat sequences onto telomere ends, ensuring the net maintenance of telomere length in tumor cells commensurate with successive rounds of cell division. Telomerase is a DNA polymerase with an endogenous RNA template, 6 on which the nascent telomeric repeats are synthesized. A significant recent finding has been that approximately 85-90% of all human cancers are positive for telomerase, both in cultured tumor cells and primary tumor tissue, whereas most somatic cells appear to lack detectable levels of telomerase. 7 This finding has been extended to a wide range of human tumors (see, for example, refs 8 and 9) and is likely to be of use in diagnosis. Human telomerase has been proposed as a novel and potentially highly selective target for antitumor drug design. This hypothesis is supported by experiments with antisense constructs against telomerase RNA in HeLa cells, which show that telomere shortening is produced, together with the death of these otherwise immortal cells. 6 Sequence-specific peptide nucleic acids directed against telomerase RNA have also been found to exert an inhibitory effect on the enzyme. Our laboratories have initiated a structure-based approach to discovering non-nucleoside compounds that will selectively inhibit human telomerase by targeting the nucleic acid structures, such as G-quadruplexes, 13,14 that may be associated with human telomeres or telomerase. In this report we first demonstrate inhibition of human telomerase by the 2,6-diamidoanthraquinone (compound 1). We also show by 1 H-NMR the stabilization of a G-quadruplex by compound 1 and, finally, provide evidence that this compound inhibits the telomerase enzyme by a mechanism consistent with interaction with G-quadruplex structures. Although Gquadruplexes have been suggested as possible molecular targets for telomerase inhibitors, 15 this is the first published report of a molecule that acts in this way and is also the first report of a non-nucleoside, small molecule inhibitor.
Telomerase and telomere dynamics in ageing and cancer: current status and future directions
Clinical and Translational Oncology, 2007
This review will focus on the clinical utilities of telomerase for human cancer diagnosis and prognosis. Much attention has been focused on control of telomerase activity in early and late stage tumours. Telomerase stabilisation may be required for cells to escape replicative senescence and to proliferate indefinitely. Because of a very strong association between telomerase and malignancy, both clinicians and pathologists expect this molecule to be a useful diagnostic and prognostic marker and a new therapeutic target. These data have greatly inspired the development of various strategies to target telomere and telomerase for cancer therapy. Finally, evidence is now emerging that G-quadruplex ligands produce rapid senescence and selective cell death. A summary of recent experimental works with new small molecules as potential inhibitors of telomerase is presented.
Therapeutic Anticancer Approaches Targeting Telomerase and Telomeres
Multi-Targeted Approach to Treatment of Cancer, 2014
Telomeres and telomerase are attractive targets for anticancer therapy. This is evidenced with the facts that majority of human cancers express the enzyme telomerase which is utmost important to maintain the telomere length, thereby to ensure indefinite cell proliferation -a hallmark of cancer. In human cells, a structure referred to as telomere has been identified to cap the terminal regions of chromosomes which can protect the ends of DNA strands from degradation and fusion, whereas telomerase plays a pivotal role in cellular immortality and tumorigenesis. Henceforth, strategies have been made to induce telomerase inhibition target virtually all of the major components of ribonucleoprotein holoenzyme and related cell signal pathways that regulates its activity which includes telomerase reverse transcriptase (hTERT) catalytic subunit, the telomere RNA component (hTERC), and associated proteins. It is noteworthy here that most of the cancers have an alternative lengthening of telomere termed as ALT even in the absence of telomerase activity. In these cases, there is an urgent need to understand the cell signaling pathways for ALT mechanism which can be used as therapeutic targets. Other strategies have been developed to target the protein associated with telomerase at the telomeric ends of chromosomes such as tankyrase. Increasing evidences suggest that directly targeting telomeric DNA using agents directed against the shelterin complex may also have anticancer activity. The limitations of strategies remain to be resolved to facilitate the clinical applications. In this chapter, recent development of strategies against these targets shall be discussed.
Cancer research, 2002
The telomeric repeat amplification protocol (TRAP) is commonly used to evaluate telomerase activity in tissues or cell extracts and also to determine the inhibitory properties of small molecules against telomerase. The recent discovery of G-quadruplex ligands as potent telomerase inhibitors prompted us to examine the accuracy of TRAP to be used to screen such class of molecules. Because of the specific feature of the TS primer, TRAP only allows the detection of G-quadruplex-induced telomerase inhibition after the synthesis of four TTAGGG repeats by telomerase and may thus lead to misinterpretations during screening assays. We have developed a TRAP-G4 assay that will allow the unambiguous detection of the inhibitory properties of a G-quadruplex ligand on telomerase activity and is able to discriminate them from other telomerase inhibitors.