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Multicharged Phthalocyanines as Selective Ligands for G-Quadruplex DNA Structures
摘要： The stabilization of G-Quadruplex DNA structures by ligands is a promising strategy for telomerase inhibition in cancer therapy since this enzyme is responsible for the unlimited proliferation of cancer cells. To assess the potential of a compound as a telomerase inhibitor, selectivity for quadruplex over duplex DNA is a fundamental attribute, as the drug must be able to recognize quadruplex DNA in the presence of a large amount of duplex DNA, in the cellular nucleus. By using different spectroscopic techniques, such as ultraviolet-visible, fluorescence and circular dichroism, this work evaluates the potential of a series of multicharged phthalocyanines, bearing four or eight positive charges, as G-Quadruplex stabilizing ligands. This work led us to conclude that the existence of a balance between the number and position of the positive charges in the phthalocyanine structure is a fundamental attribute for its selectivity for G-Quadruplex structures over duplex DNA structures. Two of the studied phthalocyanines, one with four peripheral positive charges (ZnPc1) and the other with less exposed eight positive charges (ZnPc4) showed high selectivity and affinity for G-Quadruplex over duplex DNA structures and were able to accumulate in the nucleus of UM-UC-3 bladder cancer cells.
关键词： telomerase inhibition,selectivity,salmon sperm DNA,UV-Vis,circular dichroism,hyperchromism,G-Quadruplexes,G4-FID,multicharged phthalocyanines
更新于2025-11-21 11:08:12
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Plasmonic Colorimetric Biosensor for Visual Detection of Telomerase Activity Based on Horseradish Peroxidase-Encapsulated Liposomes and Etching of Au Nanobipyramids
摘要： Telomerase aberrant activation is a critical feature in the vast majority of cancers. To visualize telomerase expression level in tumor cells, we developed a plasmonic colorimetric sensor for highly sensitive detection of telomerase activity by integrating an excellent etching substrate Au nanobipyramids (Au NBPs) with a liposome-based signal amplification strategy. Upon telomerase-triggered extension, the telomerase activity was converted into the amount of the attached horseradish peroxidase-encapsulated liposomes (HRP-Ls) on the surfaces of magnetic beads. Afterwards, HRP was liberated from the liposomes following the addition of H2O2-3,3′,5,5′-tetramethylbenzidine sulfate (TMB) substrate, and then the oxidation reaction between H2O2 and TMB was initiated to form TMB2+. The morphological evolution of Au NBPs relied on the TMB2+-mediated etching reaction, which gave rise to tremendous localized surface plasmon resonance (LSPR) responses and concomitant tonality transitions. Benefiting from cascaded signal amplification capacity of HRP-Ls and the intriguing optical properties of Au NBPs, an impressive sensitivity toward telomerase was obtained with detection limits equivalent to 1 HeLa cell for LSPR peak measurement and a visual detection limit of 20 HeLa cells. Furthermore, a facile and portable kit was fabricated for visual evaluation of telomerase activity in different cell lines.
关键词： Plasmonic colorimetry,Au nanobipyramids,Telomerase,Liposome-based amplification,Multicolor visual detection
更新于2025-11-14 17:04:02
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A Fluorescence Light-up Silver Nanocluster Beacon Modulated by Metal Ions and Its Application in Telomerase Activity Detection
摘要： DNA-templated silver nanoclusters (DNA-AgNCs) have been extensively studied in recent years. The enhancement of fluorescence emission from the DNA-AgNCs is still being explored. Herein, we report a new study on the fluorescence enhancement of DNA-AgNCs induced by metal ions. The enhancement is greatly dependent on the primary sequence and secondary structure of DNA strands. Thus, a label-free AgNCs-based molecular beacon (MB) is explored for the detection of telomerase activity. Nonfluorescent MB-AgNCs in phosphate buffer emit a dramatic red fluorescence when Mg2+ is introduced, whereas Mg2+ has a limited effect on the weak fluorescence of DNA-AgNCs when the hairpin structure of MB is opened. Telomerase primer (TP) can be elongated by telomerase, resulting in the unfolding of MB via strand displacement reaction. Based on the different brightnesses of AgNCs produced by the two DNA templates, telomerase activity is detected. MB-AgNCs sensing platform provides a simple and low-costing method to detect telomerase activity and shows a great potential in the construction of cost-effective probes for biomolecular detection.
关键词： telomerase detection,DNA,fluorescence enhancement,silver nanocluster,metal ions
更新于2025-09-23 15:23:52
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Dual-mode Au nanoprobe based on surface enhancement Raman scattering and colorimetry for sensitive determination of telomerase activity both in cell extracts and the urine of patients
摘要： Telomerase is a valuable biomarker, which is highly correlated to cancer diseases. However, single-mode probe for telomerase detection cannot satisfy the challenge of detection of telomerase activity rapidly, simply with high selectivity, sensitivity and accuracy both in preliminary diagnosis and point of care (POC) testing. Therefore, there is an urgent need to develop a robust approach with controllable assembly and high accuracy to consider both the simplification of preliminary diagnosis and POC testing and the quantification requirement for early clinical diagnosis and treatment. Herein, a novel dual-mode Au NPs probe was developed for determination of telomerase activity with controllable assembly and aggregation statement based on surface enhancement Raman scattering (SERS) and colorimetry. In this strategy, an Au dimer-based probe with high uniformity was assembled successfully, telomerase activity was reflected according to the color variations of solution and the Raman intensity of Raman reporter. Taking advantage of the uniformity of Au dimers and the combination of colorimetry and SERS techniques, our strategy determined the telomerase activity with high accuracy, sensitivity and wide range. The established probe possessed of high selectivity, sensitivity and accuracy, which was approved as a reliable, intuitional and convenient approach for detecting telomerase activity.
关键词： colorimetry,bladder cancer,surface enhanced Raman scattering,Au nanoparticles,telomerase activity
更新于2025-09-23 15:21:21
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Molecular imaging of telomerase and the enzyme activity-triggered drug release by using a conformation-switchable nanoprobe in cancerous cells
摘要： So far, the development of a unique strategy for specific biomolecules activity monitoring and precise drugs release in cancerous cells is still challenging. Here, we designed a conformation-switchable smart nanoprobe to monitor telomerase activity and to enable activity-triggered drug release in cancerous cells. The straightforward nanoprobe contained a gold nanoparticle (AuNP) core and a dense layer of 5-carboxyfluorescein (FAM)-labeled hairpin DNA shell. The 3′ region of hairpin DNA sequence could function as the telomerase primer to be elongated in the presence of telomerase, resulting in the conformational switch of hairpin DNA. As a result, the FAM fluorescence was activated and the anticancer drug doxorubicin (Dox) molecules which intercalated into the stem region of the hairpin DNA sequence were released into cancerous cells simultaneously. The smart method could specifically distinguish cancerous cells from normal cells based on telomerase activity. It also showed a good performance for monitoring telomerase activity in the cytoplasm by molecular imaging and precise release of Dox triggered by telomerase activity in cancerous cells. These advantages may offer a great potential of this method for monitoring telomerase activity in cancer progression and estimating therapeutic effect.
关键词： drug release,molecular imaging,nanoprobe,telomerase,cancerous cells
更新于2025-09-23 15:21:21
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A Conformational Switch-based Fluorescent Biosensor for Homogeneous Detection of Telomerase Activity
摘要： As a universal tumor biomarker, research on the activity and inhibition of telomerase is of great importance for cancer diagnosis and therapy. Herein, we demonstrate the conformational switch-based fluorescence detection of telomerase activity using a redesigned RNA aptamer Spinach. Briefly, the original Spinach aptamer was extended at its 5’ end and folded into an inactive conformation, where association with the small molecule fluorophore, 5-difluoro-4-hydroxybenzylidene imidazolinone (DFHBI) was prevented. Only in the presence of telomerase, (TTAGGG)n repeats were added to the 3’ end of the telomerase substrate primer, and the elongation products hybridized with inactive Spinach molecules, triggering its conformational switch and refolding it into the active, DFHBI-binding conformation. Moreover, the fluorescence signal was further amplified through a target recycling circuit, where Ribonuclease H (RNase H) specifically hydrolyzed the phosphodiester bonds of RNA in the DNA-RNA hybrid. The released telomere products could then hybridize to new inactive Spinach molecules and initiate multiple amplification cycles. The proposed fluorescent biosensor presented great performance for telomerase activity detection from 100 to 5 × 104 Hela cells with a detection limit of 100 cells. Besides, this new assay offers a good biosensing platform for differentiation of cancer cell lines from normal cell line and evaluation the inhibition efficiency of telomere-binding ligand, which is of great importance for telomerase-related cancer diagnosis and therapy.
关键词： Telomerase activity,Spinach,Ribonuclease H,Inhibitor,Fluorescent biosensors
更新于2025-09-19 17:15:36
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DNA detection by dye labeled oligonucleotides using surface enhanced Raman spectroscopy
摘要： Dye labeled oligonucleotides have been applied to detection of DNA with high sensitivity using surface enhanced Raman spectroscopy (SERS). Duplex formation of dye labeled oligonucleotides with DNA targets, followed by selective conjugation with silver nanoparticles, made it possible to determine the optimal distance between the SERS label and the silver surface, which was necessary for strong signal enhancement. For all Rh6G labeled oligonucleotides the limits of detection were 3 fmol dm–3, while for Cy3 labeled oligonucleotides the signal enhancement depended on the distance between Cy3 dye and silver nanoparticles.
关键词： surface enhanced Raman spectroscopy,promoter of human telomerase catalytic subunit gene,optical sensors,DNA detection,dye labeled oligonucleotides,specific sequence,silver nanoparticles
更新于2025-09-19 17:13:59
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Facile Synthesis of Fluorescent Tungsten Oxide Quantum Dots for Telomerase Detection Based on the Inner Filter Effect
摘要： The traditional detection of telomerase activity is mainly based on the polymerase chain reaction (PCR), which has the disadvantages of being time-consuming and susceptible to interference, and we here propose a facile method to fabricate fluorescent tungsten oxide quantum dots (WOx QDs) and employ them for telomerase activity sensing. It is found that the fluorescence of WOx QDs can be significantly quenched by hemin based on the inner filter effect (IFE). However, in the presence of telomerase, the primer-DNA can be extended to generate repeating unit of TTAGGG to form G-quadruplex, and thus hemin can be encapsulated to reduce its absorbance, resulting in decreased IFE and efficient fluorescence recovery of WOx QDs. Based on the fluorescence changes of IFE between hemin and WOx QDs, the telomerase activity with the range of 50-30000 HeLa cells can be detected and the lowest detection amount can reach 17 cells. The method exhibits good versatility that can be also applied to the telomerase detection from A549 and L929 cells. In addition, because of the good biocompatibility of the sensor, it can be used for real-time monitoring of telomerase activity in living cells, showing great potential in tumor diagnosis and inhibitor drug screening.
关键词： inner filter effect,G-quadruplex,hemin,telomerase activity detection,fluorescent tungsten oxide quantum dots
更新于2025-09-19 17:13:59
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Efficient Cytoplasmic Delivery of Antisense Probes Assisted by Cyclized Peptide-mediated Photo-induced Endosomal Escape
摘要： Intracellular delivery and endosomal release of antisense oligonucleotides remain a significant challenge in the development of gene-targeted therapeutics. Previously, non-covalently cyclized TAT peptide, accomplished by hybridization of two short complementary γPNA segments, has been proven more efficient than its linear analogues to enter cells. Since Cyc-TAT also readily accommodates a binding site, i.e., an overhanging γPNA sequence, for co-delivery of functional nucleic acid probes into cells, herein we demonstrated that the overhang-Cyc-TAT penetrated into A549 cells, carrying an anti-telomerase γPNA, which specifically reduced telomerase activity over 97%. In addition, we firstly reported that the cyclized TAT(FAM) can much more efficiently escape endosomes than the linear TAT(FAM) by LED illumination (490 nm). Based on this observation, the endosomal release of overhang-Cyc-TAT(FAM)/anti-telomerase γPNA complex can be greatly enhanced by photo-activation, shortening the cell treatment time from 60 h to 3 h with the same high efficiency in inhibiting telomerase activity inside A549 cells.
关键词： photo activation,cyclized TAT,cytoplasmic delivery,anti-telomerase γPNA
更新于2025-09-09 09:28:46
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Binding Study of the Fluorescent Carbazole Derivative with Human Telomeric G-Quadruplexes
摘要： The carbazole ligand 3 was synthesized, characterized and its binding interactions with human telomeric (22HT) G-quadruplex DNA in Na+ and K+-containing buffer were investigated by ultraviolet-visible (UV-Vis) spectrophotometry, fluorescence, circular dichroism (CD) spectroscopy, and DNA melting. The results showed that the studied carbazole ligand interacted and stabilized the intramolecular G-quadruplexes formed by the telomeric sequence in the presence of sodium and potassium ions. In the UV-Vis titration experiments a two-step complex formation between ligand and G-quadruplex was observed. Very low fluorescence intensity of the carbazole derivative in Tris HCl buffer in the presence of the NaCl or KCl increased significantly after addition of the 22HT G4 DNA. Binding stoichiometry of the ligand/G-quadruplex was investigated with absorbance-based Job plots. Carbazole ligand binds 22HT with about 2:1 stoichiometry in the presence of sodium and potassium ions. The binding mode appeared to be end-stacking with comparable binding constants of ~105 M?1 as determined from UV-Vis and fluorescence titrations data. The carbazole ligand is able to induce formation of G4 structure of 22HT in the absence of salt, which was proved by CD spectroscopy and melting studies. The derivative of carbazole 3 shows significantly higher cytotoxicity against breast cancer cells then for non-tumorigenic breast epithelial cells. The cytotoxic activity of ligand seems to be not associated with telomerase inhibition.
关键词： carbazole derivative,telomerase,biological activity,G-quadruplex,telomere,spectroscopy
更新于2025-09-09 09:28:46