G-Quadruplexes and Their Ligands: Biophysical Methods to Unravel G-Quadruplex/Ligand Interactions

Santos, Tiago; Salgado, Gilmar F.; Cabrita, Eurico J.; Cruz, Carla

doi:10.3390/ph14080769

Cited by 83 publications

(57 citation statements)

References 242 publications

(338 reference statements)

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“…However, noncanonical forms of nucleic acids have recently come to the attention of researchers using high pressure techniques. Among them, the four-strand G-quadruplex (GQ) structures are the most important ones [ 132 , 133 ]. They are formed by the guanine-rich sequences of the genome; four guanines are arranged in a planar structure stabilized by Hoogsteen-type hydrogen bonds.…”

Section: Pressure Effect On Some Biomoleculesmentioning

confidence: 99%

Biomolecules under Pressure: Phase Diagrams, Volume Changes, and High Pressure Spectroscopic Techniques

Smeller

2022

IJMS

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Pressure is an equally important thermodynamical parameter as temperature. However, its importance is often overlooked in the biophysical and biochemical investigations of biomolecules and biological systems. This review focuses on the application of high pressure (>100 MPa = 1 kbar) in biology. Studies of high pressure can give insight into the volumetric aspects of various biological systems; this information cannot be obtained otherwise. High-pressure treatment is a potentially useful alternative method to heat-treatment in food science. Elevated pressure (up to 120 MPa) is present in the deep sea, which is a considerable part of the biosphere. From a basic scientific point of view, the application of the gamut of modern spectroscopic techniques provides information about the conformational changes of biomolecules, fluctuations, and flexibility. This paper reviews first the thermodynamic aspects of pressure science, the important parameters affecting the volume of a molecule. The technical aspects of high pressure production are briefly mentioned, and the most common high-pressure-compatible spectroscopic techniques are also discussed. The last part of this paper deals with the main biomolecules, lipids, proteins, and nucleic acids: how they are affected by pressure and what information can be gained about them using pressure. I I also briefly mention a few supramolecular structures such as viruses and bacteria. Finally, a subjective view of the most promising directions of high pressure bioscience is outlined.

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Section: Pressure Effect On Some Biomoleculesmentioning

confidence: 99%

Biomolecules under Pressure: Phase Diagrams, Volume Changes, and High Pressure Spectroscopic Techniques

Smeller

2022

IJMS

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“…In order to identify true hits, the 103 computationally selected small molecules were screened for their ability to thermally stabilize TERRA G4 by using circular dichroism (CD) melting assay. Although CD is not usually used for large-scale screening purposes, it is widely employed to select nucleic acid-interacting compounds with high reliability [ 57 , 58 ]. Indeed, CD experiments require unmodified oligonucleotides, so changes in CD melting curves should only be produced by the direct interaction of the putative ligand with the nucleic acid structure.…”

Section: Resultsmentioning

confidence: 99%

Targeting of Telomeric Repeat-Containing RNA G-Quadruplexes: From Screening to Biophysical and Biological Characterization of a New Hit Compound

Marzano

Pagano

Iaccarino

et al. 2021

IJMS

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DNA G-quadruplex (G4) structures, either within gene promoter sequences or at telomeres, have been extensively investigated as potential small-molecule therapeutic targets. However, although G4s forming at the telomeric DNA have been extensively investigated as anticancer targets, few studies focus on the telomeric repeat-containing RNA (TERRA), transcribed from telomeres, as potential pharmacological targets. Here, a virtual screening approach to identify a library of drug-like putative TERRA G4 binders, in tandem with circular dichroism melting assay to study their TERRA G4-stabilizing properties, led to the identification of a new hit compound. The affinity of this compound for TERRA RNA and some DNA G4s was analyzed through several biophysical techniques and its biological activity investigated in terms of antiproliferative effect, DNA damage response (DDR) activation, and TERRA RNA expression in high vs. low TERRA-expressing human cancer cells. The selected hit showed good affinity for TERRA G4 and no binding to double-stranded DNA. In addition, biological assays showed that this compound is endowed with a preferential cytotoxic effect on high TERRA-expressing cells, where it induces a DDR at telomeres, probably by displacing TERRA from telomeres. Our studies demonstrate that the identification of TERRA G4-targeting drugs with potential pharmacological effects is achievable, shedding light on new perspectives aimed at discovering new anticancer agents targeting these G4 structures.

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“…A major outcome of our screen is the identification of non-standard G4-interacting molecular scaffolds ( Fig 1 and S2 Fig ). Here “non-standard” means that very few of our initial hits share features with the common scaffolds derived from G4 virtual and actual screens or by design [ 14 , 80 , 81 ]. Common G4 scaffolds are flat, polycyclic, and highly conjugated molecules that often prefer interaction by stacking or end-pasting onto the terminal G-tetrads [ 80 ].…”

Section: Discussionmentioning

confidence: 99%

Drug discovery of small molecules targeting the higher-order hTERT promoter G-quadruplex

et al. 2022

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DNA G-quadruplexes (G4s) are now widely accepted as viable targets in the pursuit of anticancer therapeutics. To date, few small molecules have been identified that exhibit selectivity for G4s over alternative forms of DNA, such as the ubiquitous duplex. We posit that the lack of current ligand specificity arises for multiple reasons: G4 atomic models are often small, monomeric, single quadruplex structures with few or no druggable pockets; targeting G-tetrad faces frequently results in the enrichment of extended electron-deficient polyaromatic end-pasting scaffolds; and virtual drug discovery efforts often under-sample chemical search space. We show that by addressing these issues we can enrich for non-standard molecular templates that exhibit high selectivity towards G4s over other forms of DNA. We performed an extensive virtual screen against the higher-order hTERT core promoter G4 that we have previously characterized, targeting 12 of its unique loop and groove pockets using libraries containing 40 million drug-like compounds for each screen. Using our drug discovery funnel approach, which utilizes high-throughput fluorescence thermal shift assay (FTSA) screens, microscale thermophoresis (MST), and orthogonal biophysical methods, we have identified multiple unique G4 binding scaffolds. We subsequently used two rounds of catalogue-based SAR to increase the affinity of a disubstituted 2-aminoethyl-quinazoline that stabilizes the higher-order hTERT G-quadruplex by binding across its G4 junctional sites. We show selectivity of its binding affinity towards hTERT is virtually unaffected in the presence of near-physiological levels of duplex DNA, and that this molecule downregulates hTERT transcription in breast cancer cells.

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G-Quadruplexes and Their Ligands: Biophysical Methods to Unravel G-Quadruplex/Ligand Interactions

Cited by 83 publications

References 242 publications

Biomolecules under Pressure: Phase Diagrams, Volume Changes, and High Pressure Spectroscopic Techniques

Biomolecules under Pressure: Phase Diagrams, Volume Changes, and High Pressure Spectroscopic Techniques

Targeting of Telomeric Repeat-Containing RNA G-Quadruplexes: From Screening to Biophysical and Biological Characterization of a New Hit Compound

Drug discovery of small molecules targeting the higher-order hTERT promoter G-quadruplex

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