Telomerase Inhibition Research Articles

The Role of Alternative RNA Splicing in the Regulation of hTERT, Telomerase, and Telomeres: Implications for Cancer Therapeutics.

Alternative RNA splicing impacts the majority (>90%) of eukaryotic multi-exon genes, expanding the coding capacity and regulating the abundance of gene isoforms. Telomerase (hTERT) is a key example of a gene that is alternatively spliced during human fetal development and becomes dysregulated in nearly all cancers. Approximately 90% of human tumors use telomerase to synthesize de novo telomere repeats and obtain telomere-dependent cellular immortality. Paradigm shifting data indicates that hTERT alternative splicing, in addition to transcription, plays an important role in the regulation of active telomerase in cells. Our group and others are pursuing the basic science studies to progress this emerging area of telomerase biology. Recent evidence demonstrates that switching splicing of hTERT from the telomerase activity producing full-length hTERT isoform to alternatively spliced, non-coding isoforms may be a novel telomerase inhibition strategy to prevent cancer growth and survival. Thus, the goals of this review are to detail the general roles of telomerase in cancer development, explore the emerging regulatory mechanisms of alternative RNA splicing of the hTERT gene in various somatic and cancer cell types, define the known and potential roles of hTERT splice isoforms in cancer cell biology, and provide insight into new treatment strategies targeting hTERT in telomerase-positive cancers.

6-Dithio-2′-deoxyguanosine analogs induce reactive oxygen species-mediated tumor cell apoptosis via bi-targeting thioredoxin 1 and telomerase

Thioredoxin 1 (Trx1) and telomerase play key roles in the development and progression process of most tumors, and they both are promising drug therapy targets. We have, for the first time, discovered that Trx1 and telomerase had a dual-target synergistic effect. Based on that results, we designed a series of 6-dithio-2′-deoxyguanosine analogs (named as YLS00X) and verified whether they can inhibit Trx1 and telomerase simultaneously. TrxR1/Trx1 system activity and telomerase expression were significantly inhibited by 6-dithio-2′-deoxyguanosine analogs, especially YLS004. YLS004 can also cause ROS accumulation, and induce tumor cell apoptosis. The vitro antitumor activity of 6-dithio-2′-deoxyguanosine analogs using MTT assay on 11 different human cancer cells and found that human colon cancer cells(HCT116) and melanoma cells (A375) were the most sensitive cells to 6-dithio-2′-deoxyguanosine analogs treatment and vivo xenografts models also confirmed that. The serum biochemical parameters and multiple organs HE staining results of subacute experiments indicated that YLS004 might be mildly toxic to immune organs, including the thymus, spleen, and hematopoietic system. Besides, YLS004 was rapidly metabolized in the rats' blood. Our study revealed that YLS004, a Trx1 and telomerase inhibitor, has strong anti-tumor effects to colon cancer and melanoma cells and is a promising new candidate drug.

Azidothymidine "Clicked" into 1,2,3-Triazoles: First Report on Carbonic Anhydrase-Telomerase Dual-Hybrid Inhibitors.

Cancer cells rely on the enzyme telomerase (EC 2.7.7.49) to promote cellular immortality. Telomerase inhibitors (i.e., azidothymidine) can represent promising antitumor agents, although showing high toxicity when administered alone. Better outcomes were observed within a multipharmacological approach instead. In this context, we exploited the validated antitumor targets carbonic anhydrases (CAs; EC 4.2.1.1) IX and XII to attain the first proof of concept on CA–telomerase dual-hybrid inhibitors. Compounds 1b, 7b, 8b, and 11b showed good in vitro inhibition potency against the CAs IX and XII, with KI values in the low nanomolar range, and strong antitelomerase activity in PC-3 and HT-29 cells (IC50 values ranging from 5.2 to 9.1 μM). High-resolution X-ray crystallography on selected derivatives in the adduct with hCA II as a model study allowed to determine their binding modes and thus to set the structural determinants necessary for further development of compounds selectively targeting the tumoral cells.

Identifying Predictive Gene Expression and Signature Related to Temozolomide Sensitivity of Glioblastomas.

Temozolomide (TMZ) is considered a standard chemotherapeutic agent for glioblastoma (GBM). Characterizing the biological molecules and signaling pathways involved in TMZ sensitivity would be helpful for selecting therapeutic schemes and evaluating prognosis for GBM. Thus, in the present study, we selected 34 glioma cell lines paired with specific IC50 values of TMZ obtained from CancerRxGene and RNA-seq data downloaded from the Cancer Cell Line Encyclopedia to identify genes related to TMZ sensitivity. The results showed that 1,373 genes were related to the response of GBM cells to TMZ. Biological function analysis indicated that epithelial–mesenchymal transition, Wnt signaling, and immune response were the most significantly activated functions in TMZ-resistant cell lines. Additionally, negative regulation of telomere maintenance via telomerase was enriched in TMZ-sensitive glioma cell lines. We also preliminarily observed a synergistic effect of combination treatment comprising TMZ and a telomerase inhibitor in vitro. We identified six genes (MROH8, BET1, PTPRN2, STC1, NKX3-1, and ARMC10) using the random survival forests variable hunting algorithm based on the minimum error rate of the gene combination and constructed a gene expression signature. The signature was strongly related to GBM clinical characteristics and exhibited good prognosis accuracy for both The Cancer Genome Atlas (TCGA) and Chinese Glioma Genome Atlas (CGGA) datasets. Patients in the high score group had a shorter survival time than those in the low score group (11.2 vs. 22.2 months, hazard ratio = 7.31, p = 4.59e−11) of the TCGA dataset. The CGGA dataset was selected as a validation group with 40 patients in the high score set and 43 patients in the low score set (12.5 vs. 28.8 months, hazard ratio = 3.42, p = 8.61e−5). Moreover, the signature showed a better prognostic value than MGMT promoter methylation in both datasets. We also developed a nomogram for clinical use that integrated the TMZ response signature and four other risk factors to individually predict patient survival after TMZ chemotherapy. Overall, our study provides promising therapeutic targets and potential guidance for adjuvant therapy of GBM.

A small molecule compound IX inhibits telomere and attenuates oncogenesis of drug-resistant leukemia cells.

Drug resistance is a common obstacle in leukemia treatment and failing to eradicate leukemia stem cells is the main cause of leukemia relapse. Previous studies have demonstrated that telomerase activity is associated with deregulated self-renewal of leukemia stem cells (LSCs). Here, we identified a novel compound IX, an imatinib derivative with a replacement fragment of a telomerase inhibitor, which can effectively eradicate LSCs but had no influence on normal hematopoietic stem cells (HSCs) survival. We showed that compound IX can decrease the viability of drug-resistant K562/G cells and blast crisis CML primary patient cells. Besides, IX can affect LSC survival, inhibit the colony-forming ability, and reduce LSC frequency. In vivo results showed that IX can relieve the tumor burden in patient-derived xenograft (PDX) model and prolong the lifespan. We observed that compound IX can not only decrease telomerase activity, but also affect the alternative lengthening of telomeres. In addition, IX can inhibit both the canonical and non-canonical Wnt pathways. Our data suggested this novel compound IX as a promising candidate for drug-resistant leukemia therapy.

Targeting telomerase for its advent in cancer therapeutics.

Telomerase has emerged as an important primary target in anticancer therapy. It is a distinctive reverse transcriptase enzyme, which extends the length of telomere at the 3' chromosomal end, and uses telomerase reverse transcriptase (TERT) and telomerase RNA template-containing domains. Telomerase has a vital role and is a contributing factor in human health, mainly affecting cell aging and cell proliferation. Due to its unique feature, it ensures unrestricted cell proliferation in malignancy and plays a major role in cancer disease. The development of telomerase inhibitors with increased specificity and better pharmacokinetics is being considered to design and develop newer potent anticancer agents. Use of natural and synthetic compounds for the inhibition of telomerase activity can lead to an opening of new vistas in cancer treatment. This review details about the telomerase biochemistry, use of natural and synthetic compounds; vaccines and oncolytic virus in therapy that suppress the telomerase activity. We have discussed structure-activity relationships of various natural and synthetic telomerase inhibitors to help medicinal chemists and chemical biology researchers with a ready reference and updated status of their clinical trials. Suppression of human TERT (hTERT) activity through inhibition of hTERT promoter is an important approach for telomerase inhibition.

Human telomere double G-quadruplex recognition by berberine-bisquinolinium imaging conjugates in vitro and in cells

Molecular tools for double or multimeric G-quadruplexes have recently received high attention due to the possibility to increase detection sensitivity, thermal stabilization and cell imaging opportunities, in oder to determine roles and functions of G-quadruplex aggregation in medical context. In this study, two smart berberine-bisquinolinium conjugates (Ber-360A and Ber-PDS), obtained by linking the berberine fluorophore to established G-quadruplex binders (with bisquinolinium scaffold), have been synthesized and evaluated for their G-quadrulex affinity, as well as to establish mechanisms of aggregation. The conjugates are two highly selective, sensitive and fluorescent sensors able to distinguish human telomere double G-quadruplexes from other type of G-quadruplexes and ds DNA. These ligands are the first example of fluorescent tools that bind to two adjacent G-quadruplex units and optically recognize double structures. Furthermore, microscopy experiments demonstrate that the conjugate Ber-PDS enters the nucleoli and targets G-quadruplex DNA in these organelles, by simultaneously displaying a strong telomerase inhibition and antitumor activity.

Selected ellipticine derivatives, known to target topoisomerase II, suppress the alternative lengthening of telomere (ALT) pathway in telomerase-negative cells.

DNA topoisomerase and telomerase enzymes are popular targets of several anti-tumor drugs. Smooth proceeding of telomeric recombination requires Topoisomerase II (Top2), which is involved in telomere-telomere recombination through functioning in relaxation of positive supercoils among the cells adopting telomerase-independent Alternative lengthening of telomere (ALT) pathway. Most of the inhibitors reported so far have been designed to targetsolely telomerase-positive cells, which can potentially lead to therapeutic failure because tumor cells treated with telomerase inhibitors can activate the ALT pathway for telomere maintenance. Knowing that ALT cells are more sensitive against a Top2 inhibitor, ICRF-93 agent, compared to telomerase-positive cells, we analyzed two selected ellipticine derivatives that we recently reported as TopII-targeting compounds, to assess their effects on the formation of DNA breaks and suppression of ALT pathway. Cell viability, Comet, C-Circle assays, dot blot, immunofluorescence staining, and telomere fluorescence in situ hybridization (FISH) staining were used for determining the effect of the compounds on ALT status of tumor cells. Treatment of ALT cells with ellipticine derivatives resulted in the formation of DNA breaks and suppression of ALT-associated phenotypes in vitro. Our results will contribute to the development of therapeutic strategies combining telomerase and ALT pathway inhibitors.

JAK2 inhibition in JAK2V617F-bearing leukemia cells enriches CD34+ leukemic stem cells that are abolished by the telomerase inhibitor GRN163L

The activating-mutation of JAK2V617F drives the development of myeloproliferative neoplasms (MPNs). Several JAK2 inhibitors such as ruxolitinib and gandotinib (LY2784544) currently in clinical trials and, provide improvements in MPNs including myelofibrosis. However, JAK2 inhibitors are non-curative and murine experiments show that JAK2 inhibitors don’t eradicate MPN stem cells and it is currently unclear how they escape. We thus determined the effect of the specific JAK2V617F inhibitor LY2784544 on leukemic stem (CD34+) cells (LSCs) using the JAK2V617F-bearing erythroleukemia cell line HEL. The LY2784544 treatment caused a transient proliferation inhibition and apoptosis of HEL cells, but a recovery occurred within a week. Thereafter, the continuous LY2784544 exposure induced the accumulation of CD34+ LSCs, and the CD34+ cells increased from 2% to >90% by week 9, which was accompanied by increased clonogenic potentials. LY2784544 was capable of stimulating CD34 expression even in CD34− HEL cells, which indicated cellular de-differentiation. A significantly enhanced expression of the stem cell factor KLF4 was observed in LY2784544-treated HEL cells. Inhibiting KLF4 expression attenuated LY2784544-mediated accumulation of CD34+ LSCs. Moreover, the telomerase inhibitor GRN163L abolished the LY2784544-effect. JAK2 inhibitors thus cause enrichment of LSCs and are unlikely to cure MPN as a monotherapy. Simultaneously targeting JAK2V617F and KLF4 or telomerase may be a novel strategy for MPN therapy, which should be of significance both biologically and clinically.

Investigation on Quantitative Structure-Activity Relationships of 1,3,4-Oxadiazole Derivatives as Potential Telomerase Inhibitors

Telomerase, a reverse transcriptase, maintains telomere and chromosomes integrity of dividing cells, while it is inactivated in most somatic cells. In tumor cells, telomerase is highly activated, and works in order to maintain the length of telomeres causing immortality, hence it could be considered as a potential marker to tumorigenesis.A series of 1,3,4-oxadiazole derivatives showed significant broad-spectrum anticancer activity against different cell lines, and demonstrated telomerase inhibition. This series of 24 N-benzylidene-2-((5-(pyridine-4-yl)-1,3,4-oxadiazol-2yl)thio)acetohydrazide derivatives as telomerase inhibitors has been considered to carry out QSAR studies. The endpoint to build QSAR models is determined by the IC50 values for telomerase inhibition, i.e., the concentration (μM) of inhibitor that produces 50% inhibition. These values were converted to pIC50 (- log IC50) values. We used the most common and transparent method, where models are described by clearly expressed mathematical equations: Multiple Linear Regression (MLR) by Ordinary Least Squares (OLS). Validated models with high correlation coefficients were developed. The Multiple Linear Regression (MLR) models, by Ordinary Least Squares (OLS), showed good robustness and predictive capability, according to the Multi-Criteria Decision Making (MCDM = 0.8352), a technique that simultaneously enhances the performances of a certain number of criteria. The descriptors selected for the models, such as electrotopological state (E-state) descriptors, and extended topochemical atom (ETA) descriptors, showed the relevant chemical information contributing to the activity of these compounds. The results obtained in this study make sure about the identification of potential hits as prospective telomerase inhibitors.

The Role of Alternative Lengthening of Telomeres Mechanism in Cancer: Translational and Therapeutic Implications.

Telomere maintenance mechanisms (i.e., telomerase activity (TA) and the alternative lengthening of telomere (ALT) mechanism) contribute to tumorigenesis by providing unlimited proliferative capacity to cancer cells. Although the role of either telomere maintenance mechanisms seems to be equivalent in providing a limitless proliferative ability to tumor cells, the contribution of TA and ALT to the clinical outcome of patients may differ prominently. In addition, several strategies have been developed to interfere with TA in cancer, including Imetelstat that has been the first telomerase inhibitor tested in clinical trials. Conversely, the limited information available on the molecular underpinnings of ALT has hindered thus far the development of genuine ALT-targeting agents. Moreover, whether anti-telomerase therapies may be hampered or not by possible adaptive responses is still debatable. Nonetheless, it is plausible hypothesizing that treatment with telomerase inhibitors may exert selective pressure for the emergence of cancer cells that become resistant to treatment by activating the ALT mechanism. This notion, together with the evidence that both telomere maintenance mechanisms may coexist within the same tumor and may distinctly impinge on patients’ outcomes, suggests that ALT may exert an unexpected role in tumor biology that still needs to be fully elucidated.

Aloe emodin inhibits telomerase activity in breast cancer cells: transcriptional and enzymological mechanism.

Telomerase plays an essential role in cancer cell proliferation. In this study, we investigated inhibition mechanism of aloe emodin (AE) on three different types of breast cancer cell lines, MDA-MB-453, MDA-MB-231 and MCF-7. The cells were treated with different concentrations of AE. Relative length of telomere and human telomerase reverse-transcriptase (hTERT) mRNA level was analyzed by quantitative PCR (qPCR). Protein level was assayed by Western blot. Sodium bisulfite methylation sequencing was performed to assess the methylation status of gene promoter. Enzymology kinetics was applied to reveal the interaction between AE and telomerase. Ultraviolet-visible titration and fluorescence resonance energy transfer (FRET) melting experiment were carried out to study the interaction between AE and telomeric DNA. Continuous AE exposure of these cells for 48 h results in shortening of telomeres and inhibition of telomerase. The transcription of hTERT was repressed by activation of E2F1 and inactivation of c-myc proteins. Significant demethylation of CpG islands in hTERT gene promoter was observed in MDA-MB-453 and MCF-7 cells. AE competed with dNTP for occupation of the enzyme active site. AE was a telomeric G-quadruplex structure stabilizer as indicated by titration test and FRET experiments. AE was a competitive inhibitor of telomerase and a G-quadruplex structure stabilizer. AE decreased the transcription of hTERT gene in the three breast cancer cell lines via up-regulation E2F1 and down-regulation c-myc expressions. The suppressed transcription was also related to the demethylation of the gene promoter.

Tumor‐Oriented Telomerase‐Terminated Nanoplatform as Versatile Strategy for Multidrug Resistance Reversal in Cancer Treatment

Multidrug resistance is one of the major problems in chemotherapy, and exploiting impactful targets to reverse drug resistance of most tumors remains a difficult problem. In this study, the tumor-oriented nanoparticle, BIBR1532-loaded peptide dendrimeric prodrug nanoassembly (B-PDPN), is used to assist telomerase inhibition for multidrug resistance reversal. B-PDPN possesses the characteristics of an acid-activated histidine to promote cellular uptake, a redox-sensitive poly(ethylene glycol) (PEG) layer to actualize endosomal escape and telomerase inhibitor release, and an acid sensitive chemical bond to facilitate chemotherapeutic drug release. Telomerase termination weakens the protective effect of hTERT protein on mitochondria and enhances reactive oxygen species (ROS) production, which increases DNA damage and apoptosis. The tumor-oriented nanoparticle B-PDPN achieves a broad-spectrum telomerase inhibition to combat multidrug resistance. In vivo experiments support the evidence that B-PDPN accumulates in the tumor site and reduces the expression of hTERT in tumor tissues to inhibit drug resistant tumor growth. This work introduces an innovative strategy of utilizing features of tumor-activated nanoplatform to assist telomerase termination. The nanoplatform enhances intracellular drug concentration and nucleus delivery of doxorubicin (DOX), and promotes DNA damage to combat multidrug resistance.

Telomeres and Telomere Length: A General Overview.

Telomeres are highly conserved tandem nucleotide repeats that include proximal double-stranded and distal single-stranded regions that in complex with shelterin proteins afford protection at chromosomal ends to maintain genomic integrity. Due to the inherent limitations of DNA replication and telomerase suppression in most somatic cells, telomeres undergo age-dependent incremental attrition. Short or dysfunctional telomeres are recognized as DNA double-stranded breaks, triggering cells to undergo replicative senescence. Telomere shortening, therefore, acts as a counting mechanism that drives replicative senescence by limiting the mitotic potential of cells. Telomere length, a complex hereditary trait, is associated with aging and age-related diseases. Epidemiological data, in general, support an association with varying magnitudes between constitutive telomere length and several disorders, including cancers. Telomere attrition is also influenced by oxidative damage and replicative stress caused by genetic, epigenetic, and environmental factors. Several single nucleotide polymorphisms at different loci, identified through genome-wide association studies, influence inter-individual variation in telomere length. In addition to genetic factors, environmental factors also influence telomere length during growth and development. Telomeres hold potential as biomarkers that reflect the genetic predisposition together with the impact of environmental conditions and as targets for anti-cancer therapies.

Telomerase inhibition, telomere attrition and proliferation arrest of cancer cells induced by phosphorothioate ASO-NLS conjugates targeting hTERC and siRNAs targeting hTERT

Telomerase activity has been regarded as a critical step in cellular immortalization and carcinogenesis and because of this, regulation of telomerase represents an attractive target for anti-tumor specific therapeutics. Recently, one avenue of cancer research focuses on antisense strategy to target the oncogenes or cancer driver genes, in a sequence specific fashion to down-regulate the expression of the target gene. The protein catalytic subunit, human telomerase reverse transcriptase (hTERT) and the template RNA component (hTERC) are essential for telomerase function, thus theoretically, inhibition of telomerase activity can be achieved by interfering with either the gene expression of hTERT or the hTERC of the telomerase enzymatic complex. The present study showed that phosphorothioate antisense oligonucleotide (sASO)-nuclear localization signal (NLS) peptide conjugates targeting hTERC could inhibit telomerase activity very efficiently at 5 μM concentration but less efficiently at 1 μM concentration. On the other hand, siRNA targeting hTERT mRNA could strongly suppress hTERT expression at 200 nM concentration. It was also revealed that siRNA targeting hTERT could induce telomere attrition and then irreversible arrest of proliferation of cancer cells.

Pyridine Derivative of the Natural Alkaloid Berberine as Human Telomeric G4-DNA Binder: A Solution and Solid-State Study.

Telomerase is an enzyme deputed to the maintenance of eukaryotic chromosomes; however, its overexpression is a recognized hallmark of many cancer forms. A viable route for the inhibition of telomerase in malignant cells is the stabilization of G-quadruplex structures (G4) at the 3' overhang of telomeres. Berberine has shown in this regard valuable G4 binding properties together with a significant anticancer activity and telomerase inhibition effects. Here, we focused on a berberine derivative featuring a pyridine containing side group at the 13th position. Such modification actually improves the binding toward telomeric G-quadruplexes and establishes a degree of selectivity in the interaction with different sequences. Moreover, the X-ray crystal structure obtained for the complex formed by the ligand and a bimolecular human telomeric quadruplex affords a better understanding of the 13-berberine derivatives behavior with telomeric G4 and allows to draw useful insights for the future design of derivatives with remarkable anticancer properties.

Pin2 telomeric repeat factor 1-interacting telomerase inhibitor 1 (PinX1) inhibits nasopharyngeal cancer cell stemness: implication for cancer progression and therapeutic targeting

BackgroundRecurrence and distant metastasis are still the main factors leading to treatment failure for malignant tumors including nasopharyngeal carcinoma (NPC). Therefore, elucidating the molecular mechanisms underlying nasopharyngeal carcinoma metastasis is of great clinical significance for targeted gene therapy and prognostic evaluation. PinX1, a tumor suppressor gene, was previously demonstrated to be a powerful tool for targeting telomerase in order to resist malignant tumor proliferation and migration. The aim of this study was to explore the mechanism through which PinX1 regulates epithelial–mesenchymal transition (EMT) and tumor metastasis in NPC and investigate its clinical significance and biological role with respect to disease progression.MethodsCell Counting Kit-8 (CCK8), Transwell assays, Colony formation analysis and Xenograft tumorigenicity assay were used to measure the nasopharyngeal CD133+ cancer stem cell proliferation, migration, and invasion abilities. Reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and western blot assays were conducted to investigate the underlying mechanism that PinX1 inhibits cell proliferation, migration, and invasion via regulating EMT in nasopharyngeal CD133+ CSCs.ResultsWe found that the overexpression of PinX1 and P53 inhibited cell proliferation, migration, and invasion, but that the inhibition of miR-200b blocked these effects, in nasopharyngeal CD133+ cancer stem cells (CSCs). Mechanistic investigations elucidated that PinX1 inhibits cell proliferation, migration, and invasion by regulating the P53/miR-200b-mediated transcriptional suppression of Snail1, Twist1, and Zeb1, consequently inhibiting EMT in nasopharyngeal CD133+ CSCs.ConclusionsOur findings indicate that PinX1 inhibits cell proliferation, migration, and invasion via P53/miR-200b-regulated EMT in the malignant progression of human NPC, which might suggest novel clinical implications for disease treatment.

Targeted Covalent Inhibition of Telomerase.

Telomerase is a ribonuceloprotein complex responsible for maintaining telomeres and protecting chromosomal integrity. The human telomerase reverse transcriptase (hTERT) is expressed in ∼90% of cancer cells where it confers the capacity for limitless proliferation. Along with its established role in telomere lengthening, telomerase also serves noncanonical extra-telomeric roles in oncogenic signaling, resistance to apoptosis, and enhanced DNA damage response. We report a new class of natural-product-inspired covalent inhibitors of telomerase that target the catalytic active site.

The TERT copy number gain is sensitive to telomerase inhibitors in human melanoma.

Telomerase reverse transcriptase (TERT) copy number gain is frequently observed in Asian melanoma patients. Here, we explored the correlation between TERT copy number and the effect of telomerase inhibitors in melanoma. A total of 78 melanoma cases were enrolled in the study. The TERT copy number was examined by QuantiGene Plex DNA assay. The sensitivity to telomerase inhibitors was evaluated in cell lines and patient-derived xenograft (PDX) models with or without TERT copy number gain. Among the 78 patients, 33.3% showed TERT copy number gain, and the incidence of this gain in acral melanoma (61.5%) was higher than that in other melanoma subtypes (P=0.02). The telomerase inhibitors 6-thio-2'-deoxyguanosine (6-Thio-dG) and epigallocatechin-3-gallate (EGCG) inhibited cell viability and repressed tumor growth in PDX models with TERT copy number gain. TERT copy number gain is frequently observed in Chinese patients with melanoma. Targeting telomerase may benefit melanoma patients with TERT copy number gain.

Intercalation of a Heterocyclic Ligand between Quartets in a G-Rich Tetrahelical Structure.

YES G‐rich oligonucleotide VK2 folds into an AGCGA‐quadruplex tetrahelical structure distinct and significantly different from G‐quadruplexes, even though it contains four G3 tracts. Herein, a bis‐quinolinium ligand 360A with high affinity for G‐quadruplex structures and selective telomerase inhibition is shown to strongly bind to VK2. Upon binding, 360A does not induce a conformational switch from VK2 to an expected G‐quadruplex. In contrast, NMR structural study revealed formation of a well‐defined VK2–360A complex with a 1:1 binding stoichiometry, in which 360A intercalates between GAGA‐ and GCGC‐quartets in the central cavity of VK2. This is the first high‐resolution structure of a G‐quadruplex ligand intercalating into a G‐rich tetrahelical fold. This unique mode of ligand binding into tetrahelical DNA architecture offers insights into the stabilization of an AGCGA‐quadruplex by a heterocyclic ligand and provides guidelines for rational design of novel VK2 binding molecules with selectivity for different DNA secondary structures.