Search Results (182)

Cancer is one of the leading causes of human mortality worldwide, and aberrant expression of the c-myb oncogene is closely associated with the development of numerous malignancies. The c-myb promoter region contains G-rich and C-rich sequences capable of forming G-quadruplex (G4) and i-motif (IM or C-quadruplex) structures, respectively. These secondary structures function as “molecular switches” for gene transcriptional regulation and represent promising targets for novel anti-tumor therapeutics. Through extensive screening, we identified carbazole derivative G51 as a unique dual-targeting ligand that simultaneously destabilized the i-motif and stabilized the G-quadruplex, consequently suppressing c-myb expression efficiently. In comparison, the single-targeting ligand G50, which could specifically bind to and unfold the G-quadruplex only, exhibited significantly weaker anti-tumor activity than G51. Notably, G51 showed potent anti-tumor efficacy in a human colorectal cancer xenograft model without significant toxicity to vital organs. G51, as a dual-targeting ligand, had specific binding to c-myb promoter quadruplexes, with destabilization of the i-motif and concurrent stabilization of the G-quadruplex. This opposing effect could provide a good opportunity for specific gene regulation, with great potential for further development of a precise therapeutic agent. This study provides a novel example for a practical therapeutic approach through coordinated gene quadruplex modulations, which sets up a good foundation for developing high-efficacy anti-tumor drugs without significant side effects. Full article

Background: Based on our previously reported series of novel 1,3,5-tris[(4-(substituted-aminomethyl)phenyl)methyl]benzene and 1,3,5-tris[(4-(substituted-aminomethyl)phenoxy)methyl]benzene derivatives, we have now designed, synthesized, and tested a new series of novel restricted and simplified structural analogues of these compounds against Plasmodium falciparum in vitro; i.e., the 1,3-bis[(4-(substituted-aminomethyl)phenyl)methyl]benzene and 1,3-bis[(4-(substituted-aminomethyl)phenoxy)methyl]benzene compounds. Methods & Results: The pharmacological results revealed significant antimalarial activity, with IC₅₀ values in the submicromolar to micromolar range. Additionally, the in vitro cytotoxicity of these new nitrogen-containing polyphenyl- or -phenoxymethylbenzene compounds was evaluated on human HepG2 cells. The compound 1f, the 1,3-bis[(4-(3-(morpholin-1-yl)propyl)aminomethyl)phenoxy)methyl]benzene derivative, emerged as one of the most potent and promising antimalarial candidates, demonstrating a cytotoxicity/antiprotozoal activity ratio of 594 against the chloroquine-sensitive Plasmodium falciparum 3D7 strain. Additionally, the 1,3-bis[((substituted aminomethyl)phenyl)methyl]benzene compound 1j and the 1,3-bis[((substituted aminomethyl)phenoxy)methyl]benzenes 2p and 2q also showed strong antimalarial potential, with selectivity indexes (SI) of over 303, 280, and 217, respectively, against the 3D7 strain, which has mefloquine-reduced sensitivity. Furthermore, the 1,3-bis[(4-(pyridin-2-ylethylaminomethyl)phenyl)methyl]benzene 2k was identified as the most noteworthy antimalarial compound, exhibiting a selectivity index (SI) that was superior to 178 against the chloroquine-resistant Plasmodium falciparum W2 strain. It has previously been suggested that the telomeres of P. falciparum may serve as potential targets for these polyaromatic compounds; thus, we assessed the ability of our novel derivatives to stabilize parasitic telomeric G-quadruplexes using a FRET melting assay. Conclusions: However, regarding the stabilization of the protozoan G-quadruplex, it was noted that the few substituted derivatives, which showed interesting stabilization profiles, were not necessarily the most effective antimalarial compounds against both Plasmodium strains. Moreover, these new compounds did not show promising stabilizing effects on the different G4 sequences. Therefore, no correlation arises between their antimalarial activity and the selectivity of their binding to G-quadruplexes. Full article

The development of molecules that interact with G-quadruplex (G4) sequences requires effective evaluation methods. Several techniques are currently available, including nuclear magnetic resonance (NMR) spectroscopy and X-ray crystallography, surface plasmon resonance (SPR), isothermal titration calorimetry (ITC) and mass spectrometry (MS), fluorescence using FRET-melting, G4-fluorescent intercalator displacement assay (G4-FID) and affinity chromatography. Among these, CD spectroscopy is gaining prominence due to its lower material requirements, faster experimentation and quicker data processing. However, conventional CD methods have limitations, such as higher sample volume required and the inability to handle high-throughput analysis efficiently. The use of synchrotron radiation in high-throughput analysis methods (HT-SRCD) has further advanced the investigation of small-molecule interactions with DNA G4 structures in the presence of various monovalent cations. HT-SRCD offers the capability to analyze multiple samples simultaneously, overcoming the limitations of conventional CD methods. To validate this approach, three biologically relevant G4 sequences—HTelo1, G3T3 and T95-2T—were investigated. Their interactions with a library of small tetrazole-based molecules, synthesized via a four-component Ugi reaction, and with a peptide sequence deriving from RHAU helicases (Rhau25), were evaluated. The results demonstrate that this method not only effectively discriminates between different ligands but also provides valuable insights into the selectivity and the modes of interaction of these ligands with the G4 sequences. Full article

The use of radiosensitizers is a beneficial approach in cancer radiotherapy treatment. However, the enhancement of radiation effects on cancer cells by radiosensitizers involves several different mechanisms, reflecting the chemical nature of the radiosensitizer. G-quadruplex (G4) DNA ligands have emerged in recent years as a potential new class of radiosensitizers binding to specific DNA sequences. Recently, we have shown that the Re(I) tricarbonyl complex PDF-Pz-Re and its pyrazolyl-diamine chelator PDF-Pz, carrying a N-methylated pyridostatin (PDF) derivative, act as G4 binders of various G4-forming DNA and RNA sequences. As described in this contribution, these features prompted us to evaluate PDF-Pz-Re and PDF-Pz as radiosensitizers of prostate cancer PC3 cells submitted to concomitant treatment with Co-60 radiation. The compound RHPS4 was also tested, as this G4 ligand was previously shown to exhibit strong radiosensitizing properties in other cancer cell lines. The assessment of the resulting radiobiological effects, namely through clonogenic cell survival, DNA damage, and ROS production assays, showed that PDF-Pz-Re and PDF-Pz were able to radiosensitize PC3 cells despite being less active than RHPS4. Our results corroborate that G4 DNA ligands are a class of compounds with potential interest as radiosensitizers, deserving further studies to optimize their radiosensitization activity and elucidate the mechanisms of action. Full article

Background/Objectives: Aberrant expression of c-MYC drives aggressive cancers. A guanine-rich promoter sequence (Pu27) folds into a transcriptionally repressive G-quadruplex (G4). Epigallocatechin gallate (EGCG), the main green tea polyphenol, displays anticancer activity, but clear, easily replicated evidence for direct binding to the c-MYC G4 is lacking. We therefore obtained the first biophysical confirmation of an EGCG–c-MYC G4 interaction using routine UV–visible spectroscopy. Methods: A pre-annealed Pu27 G4 (5 µM) in potassium-rich buffer was titrated with freshly prepared EGCG (0–20 µM) at 25 °C. Full-range UV–Vis spectra (220–400 nm) were recorded after each addition, and absorbance variations at the DNA (260 nm) and ligand (275 nm) maxima were quantified across three independent replicates. Results: EGCG induced pronounced, concentration-dependent hyperchromicity at 260 nm, reaching ~8–10% above baseline at a 4:1 ligand/DNA ratio and exhibiting saturable binding behaviour. Concurrently, the 275 nm band displayed relative hypochromicity coupled with a subtle bathochromic shift. These reciprocal perturbations—absent in buffer-only controls—constitute definitive evidence of a specific EGCG•G4 complex most consistent with external π-stacking or groove engagement rather than intercalation. Conclusions: This study delivers the first rigorous, quantitative UV–Vis confirmation that a readily consumed dietary polyphenol directly targets the c-MYC promoter G4. By marrying conceptual elegance with methodological accessibility, it provides a compelling molecular rationale for EGCG’s anti-oncogenic repertoire, inaugurates an expedient platform for screening G4-reactive nutraceuticals, and paves the way for structural and cellular investigations en route to next-generation c-MYC-directed therapies. Full article

G-quadruplex (G4) structures are non-canonical nucleic acid conformations that play crucial roles in gene regulation, DNA replication, and telomere maintenance. Recent studies have highlighted G4 ligands as promising anticancer agents due to their ability to modulate oncogene expression and induce DNA damage. By stabilizing G4 structures, these ligands affect tumor progression. Additionally, they have been implicated in tumor immunity modulation, particularly through the activation and immunogenic cell death induction of the cyclic GMP–AMP synthase (cGAS)–stimulator of interferon genes (STING) signaling pathway. Moreover, their disruption of telomere maintenance and regulation of key oncogenes, such as c-MYC and KRAS, position them as candidates for immune-based therapeutic interventions. Despite their therapeutic potential, challenges remain in optimizing their clinical applications, particularly in patient stratification and elucidating their immunomodulatory effects. This review provides a comprehensive overview of the mechanisms through which G4 ligands influence tumor progression and immune regulation, highlighting their potential role in future cancer immunotherapy strategies. Full article

G-quadruplex (G4) structures have emerged as critical regulatory elements in viral genomes and represent potential targets for antiviral intervention. In this study, we identified and characterized G4 structures in the unique long (UL) region of the Pseudorabies virus (PRV) genome, highlighting their role as novel antiviral targets. Bioinformatic analysis revealed two guanine-rich regions (R1 and R2) that form stable G4 structures, as confirmed by fluorescence assays, circular dichroism (CD) spectroscopy, and immunofluorescence staining. Notably, these G4 structures exhibit a tandem repeat arrangement, a previously unreported feature in the PRV genome. Epiberberine (EPI), a natural G4-stabilizing ligand, bound to and stabilized these structures, leading to the inhibition of Taq polymerase progression. Functional assays demonstrated that EPI effectively suppressed PRV replication in vitro while having no significant impact on viral entry or release. In vivo, EPI treatment significantly improved survival rates and reduced viral loads in multiple organs, including the brain, heart, lungs, and kidneys of infected mice. These findings provide new insights into the role of G4 structures in PRV replication and demonstrate that EPI exhibits potential antiviral activity by targeting G4 structures. Full article

MicroRNA-155-3p (miR-155-3p) is an important biomarker in various pathological conditions, including cancer, making the development of sensitive and specific detection methods crucial. Here, we present a molecular beacon (MB-G4) that underwent a conformational switch upon hybridization with miR-155-3p, enabling the formation of a G-quadruplex (G4) structure. This G4 was recognized by the fluorogenic ligand N-methyl mesoporphyrin IX (NMM), producing a fluorescence signal proportional to the target concentration, making it a new detection method. The conformational dynamics of MB-G4 were characterized through circular dichroism (CD) spectroscopy and native polyacrylamide gel electrophoresis (PAGE), confirming the transition from a hairpin structure to an RNA–DNA hybrid duplex that facilitated G4 formation. The optimization of the experimental conditions, including the potassium chloride (KCl) and NMM concentrations, ensured selective detection with minimal background signal. The detection limit (LOD) was determined to be 10.85 nM, using a linear fluorescence response curve, and the specificity studies demonstrated a clear distinction between miR-155-3p and miR-155-5p. Furthermore, MB-G4 was studied with total RNA extracted from the lung cancer cell line A549 to evaluate its detection in a more complex environment and was able to detect its target, validating its potential for biological sample analysis. Full article

Within the framework of rational drug design, this study introduces a novel approach to enhance the specificity of small molecules in targeting cancer cells. This approach starts from the use of dyads merging into a single entity, a naphthalene diimide (NDI) and core-extended NDI (ceNDI), both known as G-quadruplex (G4) ligands and fluorescent probes. The strategy aims to leverage the unique diagnostic strengths of the ceNDI moiety featuring red emission by improving its binding affinity and target selectivity through inclusion in dyads built with different linkers. The newly developed NDI-ceNDI dyads are promising probes, as they exhibit fluorescence turn-on upon DNA recognition and induced circular dichroism signals dependent on DNA conformation. Both dyads have an excellent affinity for hybrid G4, with two orders of magnitude higher binding constants than those for ds DNA. Their high cytotoxicity on cancer cell lines further demonstrates their potential as therapeutic agents, highlighting the role of the linker in target selectivity. Specifically, only the dyad with the rigid triazole linker exhibits selectively induced DNA damage in transformed cells, compared to normal cells primarily targeting telomeric regions. Our findings shed light on DIPAC’s potential as a promising theranostic agent, offering insights into future developments in precision medicine. Full article

G-quadruplex (G4), an important secondary structure of nucleic acids, is polymorphic in structure. G4 monomers can associate with each other to form multimers, which show better application performance than monomers in some aspects. G4 dimers, the simplest and most widespread multimeric structures, are often used as a representative for studying multimers. RHAU, a G4 ligand, has been reported to recognize G4 dimers. However, there are few reports focusing on interactions between RHAU and different G4 dimers. In this work, interactions between RHAU peptide and six G4 dimers were investigated by size-exclusion chromatography (SEC). It was revealed that compared to the hybrid G4 monomer, the hybrid tandem unstacked G4 dimer could form special binding sites, leading to a weak interaction with RHAU. It was also found that the steric hindrance at terminal G-tetrads of a special Z-G4 structure greatly weakened their interactions with RHAU. Additionally, RHAU exhibited stronger interactions with intermolecular stacked/interlocked parallel dimers than with intramolecular tandem stacked parallel dimers. This work enriches the understanding of interactions between RHAU and G4 dimers, which is conducive to the elucidation of G4 polymorphism, and provides a strong reference for studying G4 multimer–peptide interactions. Full article

G-quadruplexes (G4s) are non-canonical secondary structures that play a crucial role in the regulation of genetic expression. This study explores the interaction between G4s and a small family of oligostyrylbenzene (OSB) derivatives, characterized by tris(styryl)benzene and tetrastyrylbenzene backbones, functionalized with either trimethylammonium or 1-methylpyridinium groups. Initially identified as DNA ligands, these OSB derivatives have now been recognized as potent G4 binders, surpassing in binding affinity commercially available ligands such as pyridostatin and displaying good selectivity for G4s over duplex DNA. Furthermore, OSB derivatives 1 and 2 demonstrated significant antiparasitic activity against bloodstream forms of T. brucei and extracellular L. major, with high selectivity indices when compared to MRC-5 healthy control cells. Derivatives 1 and 2 exhibited moderate biocidal effects against a range of Gram-positive and Gram-negative bacterial strains. Notably, a synergistic antibacterial effect was observed when these compounds were combined with traditional antibiotics, particularly against Acinetobacter baumannii, highlighting their potential utility in addressing drug-resistant bacterial infections. The differences in bioactivity among the OSB derivatives can be attributed to variations in cellular uptake, as proved by flow cytometry analysis. This suggests that the degree of cellular internalization plays a pivotal role in the observed antiparasitic and antibacterial efficacy. Full article

Herein, we report the structure-based selection via molecular docking of four N-heterocyclic bis-carbene gold(I) complexes, whose potential as ligands for the hTel23 G-quadruplex structure has been investigated using circular dichroism (CD) spectroscopy, CD melting, and polyacrylamide gel electrophoresis (PAGE). The complex containing a bis(1,2,3,4,6,7,8,9-octahydro-11H-11λ³-pyridazino[1,2-a]indazol-11-yl) scaffold induces a transition from the hybrid (3 + 1) topology to a prevalent parallel G-quadruplex conformation, whereas the complex featuring a bis(2-(2-acetamidoethyl)-3λ³-imidazo[1,5-a]pyridin-3(2H)-yl) moiety disrupted the original G-quadruplex structure. These results deserve particular attention in light of the recent findings on the pathological involvements of G-quadruplexes in neurodegenerative diseases. Full article

The analysis of nucleic acid structures, topologies, nano-mechanics and interactions with ligands and other biomacromolecules (most notably proteins) at the single molecule level has become a fundamental topic in molecular biophysics over the last two decades. Techniques such as molecular tweezers, single-molecule fluorescence resonance energy transfer, and atomic force microscopy have enabled us to disclose an unprecedented insight into the mechanisms governing gene replication, transcription and regulation. In this minireview, we survey the main working principles and discuss technical caveats of the above techniques, using as a fil-rouge the history of their achievements in dissecting G-quadruplexes. The revised literature offers a clear example of the superior ability of single-molecule techniques with respect to ensemble techniques to unveil the structural and functional diversity of the several polymorphs corresponding to a single G-quadruplex folding sequence, thus shedding new light on the extreme complexity of these fascinating non-Watson–Crick structures. Full article

Guanine-quadruplex (G4) selective photosensitizers have huge potential for photodynamic therapy against various diseases correlated with G4 DNA and G4 RNAs; however, the types of photosensitizer skeletons available are limited. Herein, we investigated the ability of our original G4 ligands, tripodal quinone-cyanine dyes (tpQCy(s)), which were developed as fluorescent probes for G4, to act as photosensitizers for cancer-selective apoptosis inducers. The results indicated that the tpQCy skeleton has great potential for developing G4-targeted cancer-selective photosensitizers for photodynamic therapy. Among the two tpQCys, only QCy(BnBT)₃, which has greater G4 selectivity, exhibited photoinduced cytotoxicity in HeLa cell growth, suggesting that the direct oxidation of G4 DNA or RNA is crucial for photoinduced cytotoxicity. RNA-seq analysis using a next-generation sequencing technique revealed that apoptosis was clearly induced by photoirradiation after QCy(BnBT)₃ treatment. Full article

This article provides a comprehensive examination of non-canonical DNA structures, particularly focusing on G-quadruplexes (G4s) and i-motifs. G-quadruplexes, four-stranded structures formed by guanine-rich sequences, are stabilized by Hoogsteen hydrogen bonds and monovalent cations like potassium. These structures exhibit diverse topologies and are implicated in critical genomic regions such as telomeres and promoter regions of oncogenes, playing significant roles in gene expression regulation, genome stability, and cellular aging. I-motifs, formed by cytosine-rich sequences under acidic conditions and stabilized by hemiprotonated cytosine–cytosine (C:C+) base pairs, also contribute to gene regulation despite being less prevalent than G4s. This review highlights the factors influencing the stability and dynamics of these structures, including sequence composition, ionic conditions, and environmental pH. Molecular dynamics simulations and high-resolution structural techniques have been pivotal in advancing our understanding of their folding and unfolding mechanisms. Additionally, the article discusses the therapeutic potential of small molecules designed to selectively bind and stabilize G4s and i-motifs, with promising implications for cancer treatment. Furthermore, the structural properties of these DNA forms are explored for applications in nanotechnology and molecular devices. Despite significant progress, challenges remain in observing these structures in vivo and fully elucidating their biological functions. The review underscores the importance of continued research to uncover new insights into the genomic roles of G4s and i-motifs and their potential applications in medicine and technology. This ongoing research promises exciting developments in both basic science and applied fields, emphasizing the relevance and future prospects of these intriguing DNA structures. Full article