Selective molecular probes for G-quadruplex and G-quadruplex-duplex junctions: Cyanine dye interactions and structural insight.

There is growing interest in hybrid DNA structures, particularly G-quadruplex-duplex junctions, as potential ligand binding sites. In this work, we investigate the interaction of two cyanine dyes (R9 and 3b), which differ in hydrophilicity, with various DNA structures, including duplex DNA, parallel and antiparallel G-quadruplexes, and a G-quadruplex-duplex junction. We employed molecular spectroscopic techniques (UV-visible absorption, circular dichroism, fluorescence), nuclear magnetic resonance (NMR) spectroscopy, multivariate analysis, and molecular docking studies.

Modified Bis-pyrimidine Clamps for Triplex Formation and Their Use in SARS-CoV‑2 Detection.

The formation of nucleic acid triple helices ("triplexes") is an area of great interest due to their potential role in the natural and artificial regulation of gene expression or for use in analytical, diagnostic, or synthetic methods. During the coronavirus pandemic, a large search for novel methods for the detection of SARS-CoV-2 was undertaken. Based on triplex affinity capture and using polypurine reverse-Hoogsteen hairpins, a method known as Triplex Enhanced Nucleic Acid Detection Assay (TENADA) was developed for the rapid detection of SARS-CoV-2 without the need for polymerase chain reaction (PCR) amplification.

Interaction process behind the strong stabilization of G-quadruplexes by alkaloid fagaronine.

Benzo[c]phenanthridine alkaloids are known for their stabilizing effects on non-canonical DNA structures, particularly G-quadruplexes (G4s). In this study, the interaction of fagaronine, a rare benzo[c]phenanthridine alkaloid, with several DNA structures (including B-DNA, parallel, antiparallel and hybrid G4s) is studied using molecular fluorescence and circular dichroism (CD) spectroscopy. It has been found that fagaronine significantly enhances the stability of all tested G4 conformations.

Bioaggregachromism of Asymmetric Monomethine Cyanine Dyes as Noncovalent Binders for Nucleic Acids.

Two new asymmetric monomethine cyanine dyes, featuring dimethoxy quinolinium or methyl quinolinium end groups and benzothiazole or methyl benzothiazole end groups were synthesized. The chemical structures of the two dyes-()-6,7-dimethoxy-1-methyl-4-((3-methylbenzo[d]thiazol-2(3H)-ylidene)methyl)quinolin-1-ium iodide () and ()-4-((3,5-dimethylbenzo[d]thiazol-2(3H)-ylidene)methyl)-1,2-dimethylquinolin-1-ium iodide ()-were confirmed through NMR spectroscopy and MALDI-TOF mass spectrometry. A new methodology was developed to study monocationic dyes in the absence of a matrix and cationizing compounds in MALDI-TOF mass experiments.

Resolution of complex mixtures of duplex and antiparallel triplex DNA structures by capillary electrophoresis and multivariate analysis.

Triplex DNA structures, which are formed by the addition of an extra strand to a target B-DNA duplex, have attracted increasing interest due to their analytical and therapeutic applications. These structures are classified into parallel and antiparallel, depending on the orientation of the Triplex-Forming Oligonucleotide (TFO) relative to the B-DNA duplex. Whereas the formation of parallel triplexes is easily detected by monitoring spectral changes in the UV region, the formation of antiparallel triplexes produces small or even no spectral variations, which makes their detection difficult and uncertain.

Nanoconfined Chlorine-Substituted Monomethine Cyanine Dye with a Propionamide Function Based on the Thiazole Orange Scaffold-Use of a Fluorogenic Probe for Cell Staining and Nucleic Acid Visualization.

The development of fluorescence-based methods for bioassays and medical diagnostics requires the design and synthesis of specific markers to target biological microobjects. However, biomolecular recognition in real cellular systems is not always as selective as desired. A new concept for creating fluorescent biomolecular probes, utilizing a fluorogenic dye and biodegradable, biocompatible nanomaterials, is demonstrated.

Assessment of methodologies based on the formation of antiparallel triplex DNA structures and fluorescent silver nanoclusters for the detection of pyrimidine-rich sequences.

In this work, strategies for the detection of pyrimidine-rich DNA target sequences based on the formation of duplex and antiparallel triplex structures are studied. The presence of the target is detected from the changes in fluorescence of silver nanoclusters stabilized by the corresponding complementary DNA probes. In all cases, the formation of intermolecular structures has been assessed by means of melting experiments and multivariate analysis.

Quadruplex-duplex junction in LTR-III: A molecular insight into the complexes with BMH-21, namitecan and doxorubicin.

Quadruplex-Duplex (Q-D) junctions are unique structural motifs garnering increasing interest as drug targets, due to their frequent occurrence in genomic sequences. The viral HIV LTR-III sequence was chosen as a Q-D junction model to study the affinity of the selected compounds BMH-21, namitecan (ST-1968), and doxorubicin (DOXO), all containing a planar polycyclic aromatic moiety, linked to either one short aminoalkyl or an aminoglycosyl group. A multidisciplinary approach that combines NMR spectroscopy, molecular modelling, circular dichroism (CD) and fluorescence spectroscopy was employed.

Biophysical evaluation of antiparallel triplexes for biosensing and biomedical applications.

Polypyrimidine sequences can be targeted by antiparallel clamps forming triplex structures either for biosensing or therapeutic purposes. Despite its successful implementation, their biophysical properties remain to be elusive. In this work, PAGE, circular dichroism and multivariate analysis were used to evaluate the properties of PPRHs directed to SARS-CoV-2 genome.

Rapid and Highly Efficient Separation of i-Motif DNA Species by CE-UV and Multivariate Curve Resolution.

The i-motif is a class of nonstandard DNA structure with potential biological implications. A novel capillary electrophoresis with an ultraviolet absorption spectrophotometric detection (CE-UV) method has been developed for the rapid analysis of the i-motif folding equilibrium as a function of pH and temperature. The electrophoretic analyses are performed in reverse polarity of the separation voltage with 32 cm long fused silica capillaries permanently coated with hydroxypropyl cellulose (HPC), after an appropriate conditioning procedure was used to achieve good repeatability.

Aggregation induced nucleic acids recognition by homodimeric asymmetric monomethyne cyanine fluorochromes in mesenchymal stem cells.

In the light of recent retrovirus pandemics, the issue of discovering new and diverse RNA-specific fluorochromes for research and diagnostics became of acute importance. The great majority of nucleic acid-specific probes either do not stain RNA or cannot distinguish between DNA and RNA. The versatility of polymethine dyes makes them suitable as stains for visualization, analysis, and detection of nucleic acids, proteins, and other biomolecules.

Exploring the stabilizing effect on the i-motif of neighboring structural motifs and drugs.

Cytosine-rich DNA sequences may fold into a structure known as i-motif, with potential in vivo modulation of gene expression. The stability of the i-motif is residual at neutral pH values. To increase it, the addition of neighboring moieties, such as Watson-Crick stabilized loops, tetrads, or non-canonical base pairs have been proposed.

Detection of pyrimidine-rich DNA sequences based on the formation of parallel and antiparallel triplex DNA and fluorescent silver nanoclusters.

In this work, the use of DNA-stabilized fluorescent silver nanoclusters for the detection of target pyrimidine-rich DNA sequences by formation of parallel and antiparallel triplex structures is studied by molecular fluorescence spectroscopy. In the case of parallel triplexes, the probe DNA fragments are Watson-Crick stabilized hairpins, and whereas in the case of antiparallel triplexes, the probe fragments are reverse-Hoogsteen clamps. In all cases, the formation of the triplex structures has been assessed by means of polyacrylamide gel electrophoresis, circular dichroism, and molecular fluorescence spectroscopies, as well as multivariate data analysis methods.

Correction: Clua et al. Properties of Parallel Tetramolecular G-Quadruplex Carrying N-Acetylgalactosamine as Potential Enhancer for Oligonucleotide Delivery to Hepatocytes. 2022, , 3944.

In the original article [...

Detection of SARS-CoV-2 Virus by Triplex Enhanced Nucleic Acid Detection Assay (TENADA).

SARS-CoV-2, a positive-strand RNA virus has caused devastating effects. The standard method for COVID diagnosis is based on polymerase chain reaction (PCR). The method needs expensive reagents and equipment and well-trained personnel and takes a few hours to be completed.

Exploring the Interaction of G-quadruplex Binders with a (3 + 1) Hybrid G-quadruplex Forming Sequence within the PARP1 Gene Promoter Region.

The enzyme PARP1 is an attractive target for cancer therapy, as it is involved in DNA repair processes. Several PARP1 inhibitors have been approved for clinical treatments. However, the rapid outbreak of resistance is seriously threatening the efficacy of these compounds, and alternative strategies are required to selectively regulate PARP1 activity.

Properties of Parallel Tetramolecular G-Quadruplex Carrying -Acetylgalactosamine as Potential Enhancer for Oligonucleotide Delivery to Hepatocytes.

The development of oligonucleotide conjugates for in vivo targeting is one of the most exciting areas for oligonucleotide therapeutics. A major breakthrough in this field was the development of multifunctional GalNAc-oligonucleotides with high affinity to asialoglycoprotein receptors (ASGPR) that directed therapeutic oligonucleotides to hepatocytes. In the present study, we explore the use of G-rich sequences functionalized with one unit of GalNAc at the 3'-end for the formation of tetrameric GalNAc nanostructures upon formation of a parallel G-quadruplex.

Studies on the interactions of Ag(i) with DNA and their implication on the DNA-templated synthesis of silver nanoclusters and on the interaction with complementary DNA and RNA sequences.

Silver nanoclusters (AgNCs) prepared by the reduction of silver ions in the presence of DNA oligonucleotides have attracted great interest as potential diagnostic tools for their tunable and high fluorescent properties. In this work, three DNA sequences that consist of a 12-nucleotide long probe sequence at the 5'-end linked to the complementary sequence to three miRNAs are studied. First, the interaction of these sequences with Ag(i) was characterized by means of circular dichroism spectroscopy.

Alkaloid Escholidine and Its Interaction with DNA Structures.

Berberine, the most known quaternary protoberberine alkaloid (QPA), has been reported to inhibit the SIK3 protein connected with breast cancer. Berberine also appears to reduce the bcl-2 and XIAP expression-proteins responsible for the inhibition of apoptosis. As some problems in the therapy with berberine arose, we studied the DNA binding properties of escholidine, another QPA alkaloid.

Study of the interaction of the palmatine alkaloid with hybrid G-quadruplex/duplex and i-motif/duplex DNA structures.

There is an increasing interest in the study of guanine or cytosine-rich sequences that may fold into G-quadruplex (G4) or i-motif (iM) structures showing a short hairpin (or stem-loop) stabilized by Watson-Crick base pairs. These hybrid spatial arrangements may be target of ligands that have been shown to interact strongly with B-DNA. In this work, the interaction of the palmatine alkaloid with several sequences forming different G4s, iMs, and hybrid structures has been studied by means of spectroscopic and separation techniques, as well as multivariate data analysis methods.

Investigation of the Complexes Formed between PARP1 Inhibitors and PARP1 G-Quadruplex at the Gene Promoter Region.

DNA repair inhibitors are one of the latest additions to cancer chemotherapy. In general, chemotherapy produces DNA damage but tumoral cells may become resistant if enzymes involved in DNA repair are overexpressed and are able to reverse DNA damage. One of the most successful drugs based on modulating DNA repair are the poly(ADP-ribose) polymerase 1 (PARP1) inhibitors.

Exploring the Interaction of Curaxin CBL0137 with G-Quadruplex DNA Oligomers.

Curaxins and especially the second-generation derivative curaxin CBL0137 have important antitumor activities in multiple cancers such as glioblastoma, melanoma and others. Although most of the authors suggest that their mechanism of action comes from the activation of p53 and inactivation of NF-kB by targeting FACT, there is evidence supporting the involvement of DNA binding in their antitumor activity. In this work, the DNA binding properties of curaxin CBL0137 with model quadruplex DNA oligomers were studied by H NMR, CD, fluorescence and molecular modeling.

Structural Effects of Incorporation of 2'-Deoxy-2'2'-Difluorodeoxycytidine (Gemcitabine) in A- and B-Form Duplexes.

We report the structural effect of 2'-deoxy-2',2'-difluorocytidine (dFdC) insertions in the DNA strand of a DNA : RNA hybrid duplex and in a self-complementary DNA : DNA duplex. In both cases, the modification slightly destabilizes the duplex and provokes minor local distortions that are more pronounced in the case of the DNA : RNA hybrid. Analysis of the solution structures determined by NMR methods show that dFdC is an adaptable derivative that adopts North type sugar conformation when inserted in pure DNA, or a South sugar conformation in the context of DNA : RNA hybrids.

G-quadruplex binding properties of a potent PARP-1 inhibitor derived from 7-azaindole-1-carboxamide.

Poly ADP-ribose polymerases (PARP) are key proteins involved in DNA repair, maintenance as well as regulation of programmed cell death. For this reason they are important therapeutic targets for cancer treatment. Recent studies have revealed a close interplay between PARP1 recruitment and G-quadruplex stabilization, showing that PARP enzymes are activated upon treatment with a G4 ligand.