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Alpha-synuclein, encoded by the SNCA gene, is a pivotal protein implicated in the pathogenesis of synucleinopathies, including Parkinson's disease. Current approaches for modulating alpha-synuclein levels involve antisense nucleotides, siRNAs, and small molecules targeting SNCA's 5'-UTR mRNA. Here, we propose a groundbreaking strategy targeting G-quadruplex structures to effectively modulate SNCA gene expression and lowering alpha-synuclein amount. Novel G-quadruplex sequences, identified on the SNCA gene's transcription starting site and 5'-UTR of SNCA mRNAs, were experimentally confirmed for their stability through biophysical assays and in vitro experiments on human genomic DNA. Biological validation in differentiated SH-SY5Y cells revealed that well-known G-quadruplex ligands remarkably stabilized these structures, inducing the modulation of SNCA mRNAs expression, and the effective decrease in alpha-synuclein amount. Besides, a novel peptide nucleic acid conjugate, designed to selectively disrupt of G-quadruplex within the SNCA gene promoter, caused a promising lowering of both SNCA mRNA and alpha-synuclein protein. Altogether our findings highlight G-quadruplexes' key role as intriguing biological targets in achieving a notable and successful reduction in alpha-synuclein expression, pointing to a novel approach against synucleinopathies., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

To develop efficient anticancer theranostic systems, we studied the interaction between a cyanine dye, analogue of thiazole orange (named CyOH), and two G-quadruplex-forming aptamers, V7t1 and 3R02, recognizing the Vascular Endothelial Growth Factor 165 (VEGF 165 ) - an angiogenic protein overexpressed in cancer cells, responsible for the rapid growth and metastases of solid tumours. We demonstrated, by exploiting different biophysical techniques - i.e. gel electrophoresis, circular dichroism (CD), UV-vis and fluorescence spectroscopy - that this cyanine interacted with both aptamers giving a marked fluorescence light-up only when bound to their dimeric forms. Interestingly, both oligonucleotides recognized VEGF 165 with higher affinity when adopting dimeric G-quadruplexes, largely prevalent over their monomeric forms in pseudo-physiological conditions. Notably, the fluorescence light-up produced by the probe was maintained when the dimeric aptamer-CyOH complexes bound to the target protein. These complexes, tested on MCF-7 cancer cells using non-tumorigenic MCF-10A cells as control, were effectively internalized in cells and colocalized with a fluorescently-labelled anti-VEGF-A antibody, allowing both recognition and detection of the target. Our experiments showed that the studied systems are promising tools for anticancer theranostic strategies, combining the therapeutic potential of the G4-forming anti-VEGF aptamers with the diagnostic efficacy of the cyanine selective fluorescence light-up., Competing Interests: Declaration of competing interest Daniela Montesarchio reports financial support was provided by Italian Association for Cancer Research. No other relationship or activity that may be interpreted as a conflict of interest by the reader., (Copyright © 2022. Published by Elsevier B.V.)

In human cells, nucleic acids adopt several non-canonical structures that regulate key cellular processes. Among them, G-quadruplexes (G4s) are stable structures that form in guanine-rich regions in vitro and in cells. G4 folded/unfolded state shapes numerous cellular processes, including genome replication, transcription, and translation. Moreover, G4 folding is involved in genomic instability. G4s have been described to multimerize, forming high-order structures in both DNA and/or RNA strands. Multimeric G4s can be formed by adjacent intramolecular G4s joined by stacking interactions or connected by short loops. Multimeric G4s can also originate from the assembly of guanines embedded on independent DNA or RNA strands. Notably, crucial regions of the human genome, such as the 3'-terminal overhang of the telomeric DNA as well as the open reading frame of genes involved in the preservation of neuron viability in the human central and peripheral nervous system are prone to form multimeric G4s. The biological importance of such structures has been recently described, with multimeric G4s playing potentially protective or deleterious effects in the pathogenic cascade of various diseases. Here, we portray the multifaceted scenario of multimeric G4s, in terms of structural properties, biological roles, and targeting strategies., (Copyright © 2022 The Authors. Published by Elsevier B.V. All rights reserved.)

To selectively target telomeric G-quadruplex (G4) DNA, monomeric and dimeric naphthalene diimides (NDIs) were investigated as binders of multimeric G4 structures able to discriminate duplex DNA. These NDIs were analysed by the affinity chromatography-based screening G4-CPG (G-quadruplex on Controlled Pore Glass), using the sequence d[AGGG(TTAGGG) 7 ] (tel46), folding into two consecutive G4s, as model of the human telomeric G4 multimer. In parallel, a telomeric G4 monomer (tel26) and a duplex structure (ds27) were used as controls. According to G4-CPG screening, NDI-5 proved to be the best ligand in terms of dimeric G4 vs. duplex DNA selectivity and was analysed by circular dichroism (CD), gel electrophoresis, isothermal titration calorimetry (ITC) and fluorescence spectroscopy in its interactions with tel46. NDI-5 strongly binds and stabilizes tel46 G4, favouring a hybrid folding in K + -containing buffer. Under these conditions, the binding process comprises a first event involving three molecules of NDI-5 and a second one in which other six molecules bind to the DNA. In a metal cation-free system, NDI-5 induces tel46 G4 folding, as indicated by CD and PAGE, favouring an antiparallel structuring. Docking simulations showed that NDI-5 can effectively bind to the pocket between two G4 units, representing a promising ligand for multimeric G4s., (Copyright © 2020 Elsevier B.V. All rights reserved.)

The saponin composition of leaves and roots from Medicago marina L., sea medic, was investigated by a combination of chromatographic, spectroscopic and spectrometric (GC, LC, ESI-MS/MS, NMR) methods. Several compounds were detected and quantified by HPLC using the external standard method. Saponins from this plant species consist of a mixture of high molecular weight bidesmosidic derivatives of medicagenic and zanhic acid, containing up to six sugars in the molecules. Six of the detected saponins were previously isolated and reported as constituents of other Medicago spp.; one saponin was previously described in other plant species; four saponins are undescribed compounds in Medicago and never reported before in other plant species. These are: 3-O-β-D-glucopyranosyl-(1 → 2)-β-D-glucopyranosylzanhic acid 28-O-β-D-xylopyranosyl-(1 → 4)-[β-D-apiofuranosyl-(1 → 3)]-α-L-rhamnopyranosyl-(1 → 2)-α-L-arabinopyranosyl ester; 3-O-β-D-glucopyranosyl-(1 → 2)-β-D-glucopyranosylzanhic acid 28-O-β-D-xylopyranosyl-(1 → 4)-α-L-rhamnopyranosyl-(1 → 2)-α-L-arabinopyranosyl ester; 3-O-β-D-glucopyranosyl-(1 → 2)-β-D-glucopyranosylmedicagenic acid 28-O-β-D-xylopyranosyl-(1 → 4)-[α-L-arabinopyranosyl-(1 → 3)]-α-L-rhamnopyranosyl-(1 → 2)-α-L-arabinopyranosyl ester and 3-O-β-D-glucopyranosylmedicagenic acid 28-O-β-D-xylopyranosyl-(1 → 4)-[α-L-arabinopyranosyl-(1 → 3)]-α-L-rhamnopyranosyl-(1 → 2)-α-L-arabinopyranosyl ester. The specific saponins synthesized by M. marina may have a role in its tolerance to environment, representing a reservoir of osmolytic sugars., Competing Interests: Declaration of competing interests The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020. Published by Elsevier Ltd.)

Target selectivity is one of the main challenges in the search for small molecules able to act as effective and non-toxic anticancer and/or antiviral drugs. To achieve this goal, handy, rapid and reliable High Throughput Screening methodologies are needed. We here describe a novel functionalization for the solid phase synthesis of oligonucleotides on Controlled Pore Glass, including a flexible hexaethylene glycol spacer linking the first nucleoside through the nucleobase via a covalent bond stable to the final deprotection step. This allowed us preparing fully deprotected oligonucleotides still covalently attached to their supports. In detail, on this support we performed both the on-line synthesis of different secondary structure-forming oligonucleotides and the affinity chromatography-based screenings of conformation-selective G-quadruplex ligands. By using a fluorescent core-extended naphthalene diimide with different emitting response upon binding to sequences folding into G-quadruplexes of different topologies, we have been able to discriminate not only G-quadruplex vs. duplex DNA structures, but also different G-quadruplex conformations on the glass beads by confocal microscopy., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Background: Fluorescent sensing of G-quadruplex nucleic acids (G4s) is an effective strategy to elucidate their role in vitro and in vivo. Small molecule ligands have often been exploited, producing an emission light up upon binding. Naphthalene diimides (NDIs), although potent G4 binders exhibiting red-NIR fluorophores, have only been marginally exploited, as they are usually quenched upon binding. Contrary, aggregating core-extended naphthalene diimides (c ex -NDIs) proved to be effective probes., Methods: We prepared a library of eighteen c ex -NDIs by organic synthesis, characterising their aggregation-dependent absorption and emission properties. Absorption and emission titrations, fluorescent intercalator displacement assay (FID) and circular dichroism (CD) analysis were performed to elucidate their behavior as G4 fluorescent sensors, selectivity and binding mode., Results: c ex -NDIs aggregate under aqueous solvents and as a result, their fluorescence is mostly quenched under physiological conditions. Upon G4 binding, they disaggregate into binding monomers, producing a fluorescent light-up with anti-parallel and hybrid G4s. Contrary, with parallel G4s a light-off was recorded. For the formers a groove-like interaction was inferred by ICD signals, while for the latter an end-stacking interaction mode was hypothesized by G4-FID data., Conclusions: c ex -NDIs G4 sensing mechanism works via a induced disaggregation. The emission response depends on the G4 topology, which dictates the prevailing -groove or end-stacking- binding mode., General Significance: This study highlights the potential of c ex -NDIs as G4 fluorescent probes. Besides being readily synthesized and conveniently emitting above 600nm, they light-up upon binding to anti-parallel and hybrid G4, complementing a number of other probes' selectivity for the parallel topology. This article is part of a Special Issue entitled "G-quadruplex" Guest Editor: Dr. Concetta Giancola and Dr. Daniela Montesarchio., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Changes in the time to first bite (TFB) and the bloodfeeding behavior of adult female Aedes aegypti (L.) mosquitoes following dissemination of Sindbis virus (SINV) were observed after exposure to repellents with the active ingredients (AI) DEET, picaridin, 2-undecanone (2-U), and oil of lemon eucalyptus. Dissemination of SINV significantly decreased (P<0.0001) the TFB of DEET (15%) and picaridin (15%) by 46% and 37%, respectively. Significant (P<0.0001) changes in activation, probing, and engorgement times were observed in SINV infected mosquitoes after exposure to the four repellents compared to uninfected mosquitoes. Taken together, a decrease in TFB and time to complete the four bloodfeeding stages will lessen the prey-status, and enhance both the chances of mosquito survival and arbovirus transmission., (Published by Elsevier B.V.)