Targeting of G‐quadruplex‐forming DNA in the BCL2 gene promoter to inhibit the expression of anti‐apoptotic Bcl‐2 protein is an attractive approach to cancer treatment. So far, efforts made in the ...discovery of molecules that are able to target the BCL2 G‐quadruplex have succeeded mainly in the identification of ligands with poor drug‐like properties. Here, a small series of furo2,3‐dpyridazin‐4(5H)‐one derivatives were evaluated as a new class of drug‐like G‐quadruplex‐targeting compounds. Biophysical studies showed that two derivatives could effectively bind to BCL2 G‐quadruplex with good selectivity. Moreover, one such ligand was found to appreciably inhibit BCL2 gene transcription, with a substantial decrease in protein expression levels, and also showed significant cytotoxicity toward the Jurkat human T‐lymphoblastoid cell line.
Up with down‐regulation: G‐quadruplex DNA structures are found in gene promoter regions and are emerging therapeutic targets in oncology. In this study, some furopyridazinone derivatives were evaluated as a new class of drug‐like G‐quadruplex‐targeting compounds. Two derivatives were shown to selectively bind and stabilize a G‐quadruplex from the promoter of the BCL2 gene, and one of them inhibited gene transcription in the cell. These compounds will aid the development of more potent derivatives with anticancer potential.
QK peptide is a vascular endothelial growth factor (VEGF)‐mimetic molecule with significant proangiogenic activity. In particular, QK is able to bind and activate VEGF receptors (VEGFRs) to stimulate ...a functional response in endothelial cells. To characterize the peptide bioactivity and its molecular recognition properties, a detailed picture of the interaction between peptide QK and VEGF receptors is reported. By combining NMR spectroscopy studies in solution on the purified receptor and in the presence of intact endothelial cells, a molecular description of the binding interaction between peptide QK and VEGFR2 in the cellular context is obtained. These results reveal useful insights into the peptide biological mechanism, which opens the way to further optimization of this class of VEGF‐mimicking peptides.
Receptive cell surfaces: The molecular details of the interaction between a vascular endothelial growth factor (VEGF)‐mimetic peptide and receptor VEGFR2 were determined by combining in vitro and cellular NMR spectroscopy experiments (see figure).
Silibinin is the main biologically active component of silymarin extract and consists of a mixture 1:1 of two diastereoisomeric flavonolignans, namely silybin A (
) and silybin B (
), which we call ...here silybins. Despite the high interest in the activity of this flavonolignan, there are still few studies that give due attention to the role of its stereochemistry and, there is still today a strong need to investigate in this area. In this regard, here we report a study concerning the radical scavenger ability and the antiproliferative activity on different cell lines, both of silybins and phosphodiester-linked silybin dimers. An efficient synthetic strategy to obtain silybin dimers in an optical pure form (
,
and
) starting from a suitable building block of silybin A and silybin B, obtained by us from natural extract silibinin, was proposed. New dimers show strong antioxidant properties, determined through hydroxyl radical (HO
) scavenging ability, comparable to the value reported for known potent antioxidants such as quercetin. A preliminary screening was performed by treating cells with 10 and 50 μM concentrations for 48 h to identify the most sensitive cell lines. The results show that silibinin compounds were active on Jurkat, A375, WM266, and HeLa, but at the tested concentrations, they did not interfere with the growth of PANC, MCF-7, HDF or U87. In particular, both monomers (
and
) and dimers (
,
and
) present selective anti-proliferative activity towards leukemia cells in the mid-micromolar range and are poorly active on normal cells. They exhibit different mechanisms of action in fact all the cells treated with the
and
go completely into apoptosis, whereas only part of the cells treated with
and
were found to be in apoptosis.
The crucial role of integrin in pathological processes such as tumor progression and metastasis formation has inspired intense efforts to design novel pharmaceutical agents modulating integrin ...functions in order to provide new tools for potential therapies. In the past decade, we have investigated the biological proprieties of the chimeric peptide RGDechi, containing a cyclic RGD motif linked to an echistatin C-terminal fragment, able to specifically recognize αvβ3 without cross reacting with αvβ5 and αIIbβ3 integrin. Additionally, we have demonstrated using two RGDechi-derived peptides, called RGDechi1-14 and ψRGDechi, that chemical modifications introduced in the C-terminal part of the peptide alter or abolish the binding to the αvβ3 integrin. Here, to shed light on the structural and dynamical determinants involved in the integrin recognition mechanism, we investigate the effects of the chemical modifications by exploring the conformational space sampled by RGDechi1-14 and ψRGDechi using an integrated natural-abundance NMR/MD approach. Our data demonstrate that the flexibility of the RGD-containing cycle is driven by the echistatin C-terminal region of the RGDechi peptide through a coupling mechanism between the N- and C-terminal regions.
Certain G-quadruplex forming guanine-rich oligonucleotides (GROs), including AS1411, are endowed with cancer-selective antiproliferative activity. They are known to bind to nucleolin protein, ...resulting in the inhibition of nucleolin-mediated phenomena. However, multiple nucleolin-independent biological effects of GROs have also been reported, allowing them to be considered promising candidates for multi-targeted cancer therapy. Herein, with the aim of optimizing AS1411 structural features to find GROs with improved anticancer properties, we have studied a small library of AS1411 derivatives differing in the sequence length and base composition. The AS1411 derivatives were characterized by using circular dichroism and nuclear magnetic resonance spectroscopies and then investigated for their enzymatic resistance in serum and nuclear extract, as well as for their ability to bind nucleolin, inhibit topoisomerase I, and affect the viability of MCF-7 human breast adenocarcinoma cells. All derivatives showed higher thermal stability and inhibitory effect against topoisomerase I than AS1411. In addition, most of them showed an improved antiproliferative activity on MCF-7 cells compared to AS1411 despite a weaker binding to nucleolin. Our results support the hypothesis that the antiproliferative properties of GROs are due to multi-targeted effects.
Galectins, β-galactoside-binding proteins, play relevant roles in different biological processes; therefore, they are becoming emerging targets for diagnostic and therapeutic approaches ....
Our goal is to evaluate benzodifuran-based scaffolds for biomedical applications.
We here explored the anticancer and anti-amyloid activities of a novel compound (BZ4) in comparison with other known ...benzodifuran analogs, previously studied in our group, and we have explored its ability to interact with different DNA model systems.
BZ4 shows antiproliferative activity on different cancer cells; does not affect noncancerous control cells and alters the aggregation properties of β-amyloid, as ascertained by circular dichroism, fluorescence spectroscopy and scanning electron microscopy analysis. An overall, qualitative picture on the mechanistic aspects related to the biological activities is discussed in light of the dynamic light scattering, UV, circular dichroism and fluorescence data, as well as of the metal ion-binding properties of BZ4.
To further explore the properties and bioactivity of benzodifurans, three compounds in this class were synthesised and characterised using spectroscopic and spectrometric techniques, as well as SEM ...and thermogravimetric analysis. The synthesised compounds showed interesting antiproliferative activity on different human cancer cells (HeLa, Hep‐G2, WM266), whereas no relevant cytotoxic effect was observed on healthy control cells. To study the possible mechanism of action of the benzodifurans, their ability to bind various model DNA systems, their aggregation properties, and their ability to bind biologically relevant metal ions was evaluated. The molecules were found to form monodisperse nanoaggregates stable at physiological temperature, which are likely related to the observed bioactivity.
The more, the better! Three benzodifuran derivatives were synthesized, characterized and tested for antiproliferative activity (see figure). They showed interesting cytotoxic effects on human cancer cell lines, with no significant effect on healthy cells. In preliminary efforts to determine the mechanism responsible for their activity, the molecules were found to form monodisperse nanoaggregates stable at physiological temperature, which are likely related to the observed bioactivity.
Recent investigations have highlighted a key role of the proteins of the KCTD (K-potassium channel tetramerization domain containing proteins) family in several fundamental biological processes. ...Despite the growing importance of KCTDs, our current understanding of their biophysical and structural properties is very limited. Biochemical characterizations of these proteins have shown that most of them act as substrate adaptor in E3 ligases during protein ubiquitination. Here we present a characterization of the KCTD5-Cullin3 interactions which are mediated by the KCTD5 BTB domain. Isothermal titration calorimetry experiments reveal that KCTD5 avidly binds the Cullin3 (Cul3). The complex presents a 5:5 stoichiometry and a dissociation constant of 59nM. Molecular modeling and molecular dynamics simulations clearly indicate that the two proteins form a stable (KCTD5–Cul3)5 pinwheel-shaped heterodecamer in which two distinct KCTD5 subunits cooperate in the binding of each cullin chain. Molecular dynamics simulations indicate that different types of interactions contribute to the stability of the assembly. Interestingly, residues involved in Cul3 recognitions are conserved in the KCTD5 orthologs and paralogs implicated in important biological processes. These residues are also rather well preserved in most of the other KCTD proteins. By using molecular modeling techniques, the entire ubiquitination system including the E3 ligase, the E2 conjugating enzyme and ubiquitin was generated. The analysis of the molecular architecture of this complex machinery provides insights into the ubiquitination processes which involve E3 ligases with a high structural complexity.
Display omitted
•We report the first quantification of Cul3 binding to a protein of the KCTD family.•MD simulations highlight the regions involved in Cul3-KCTD interactions.•KCTD5-Cul3 recognition is characterized by a high level structural complexity.•Homology modeling indicates that INC interacts with Cul3 using a similar mechanism.•The architecture of the entire KCTD5-containing E3 ligase system has been analyzed.
The scalp-ear-nipple (SEN) syndrome is an autosomal-dominant disorder characterized by cutis aplasia of the scalp and malformations of breast, external ears, digits, and nails. Genetic analyses have ...shown that the disease is caused by missense mutations of the KCTD1 protein, although the functional/structural basis of SEN insurgence is hitherto unknown. With the aim of unravelling the molecular basis of the SEN syndrome associated with KCTD1 mutations we here expressed and characterized several disease causing mutants. A preliminary dissection of the protein provides insights into the role that individual domains play in KCTD1 stability. The characterization of SEN-causing mutants indicates that, although the mutation sites are located in distant regions of the BTB domain or of the pre-BTB region, all of them are unable to interact with the transcription factor AP-2α, a well-known KCTD1 biological partner. Notably, all mutations, including the one located in the pre-BTB region, produce a significant destabilization of the protein. The structural role of the pre-BTB region in KCTD1 and other proteins of the family is corroborated by its sequence conservation in orthologs and paralogs. Interestingly, SEN-causing mutations also favor the tendency of KCTD1 to adopt structural states that are characterized by the ability to bind the β-amyloid fluorescent dye thioflavin T. The formation of aggregation-prone species may have important implications for the disease etiology. Collectively, these findings provide an intriguing picture of the functional and structural alterations induced by KCTD1 mutations that ultimately lead to disease.
PDF
