Starting from 5,6-dihidroxyindole (DHI), synthesized according to standard procedures, an ad hoc protocol through the exposure of oxidizing selenium salts for the synthesis of previously uncharacterized trimeric and dimeric species was designed. The latter were characterized and identified, revealing that they consist of dimeric and trimeric species arising from a highly regioselective (C-2 position) pathway. Subsequent deposition on glass substrates (using spin-coating technology and exposure to ammonia vapors to polymerize the product) leads to the production of thin films with remarkable stability and reproducibility across a wide range of thicknesses. Current-voltage measurements show that the optimized product exhibits stable electrical conductivity over time and ohmic in nature. These observations, together with the high biocompatibility of the material, make these films promising candidates for the manufacture of biosensor devices.

Starting from 5,6-dihydroxyindole (DHI), synthesized according to standard procedures, an ad hoc protocol through the exposure of oxidizing selenium salts for the synthesis of previously uncharacterized trimeric and dimeric species was designed. The latter were characterized and identified, revealing that they consist of dimeric and trimeric species arising from a highly regioselective (oxidizing position) pathway. Subsequent deposition on glass substrates (using spin-coating technology and exposure to ammonia vapors to polymerize the product) leads to the production of thin films with remarkable stability and reproducibility across a wide range of thicknesses. Current-voltage measurements show that the optimized product exhibits stable electrical conductivity over time and ohmic in nature. These observations, together with the high biocompatibility of the material, make these films promising candidates for the manufacture of biosensor devices.

Galectins, β-galactoside-binding soluble proteins, are involved in a multitude of biological functions and several diseases, so numerous galectin inhibitors, from small molecules to multivalent glycoconjugates, have been developed and investigated as tools for therapeutic applications. Notably, multivalent ligands on a biocompatible backbone offer a promising perspective for creating high-performance selective inhibitors with nanomolar affinity. Leveraging the oxidative polymerization of 5,6-dihydroxyindole (DHI), a key intermediate of eumelanin pigments, here, we present the synthesis and complete nuclear magnetic resonance characterization of a submicromolar multivalent ligand of galectin-3 based on a naturally biocompatible eumelanin backbone. The integration of several complementary techniques, namely, UV-vis spectrometry, dynamic light scattering, isothermal titration calorimetry, and biolayer interferometry, in the investigation of galectin-3-eumelanin-related ligand interactions, allowed us to calculate the KD and to propose a model for the protein-polymer interaction.

Oligo- and polysaccharides encode biological information through the “sugar code” interpreted by galectins (Gals), a family of β-galactoside–binding lectins involved in key physiological and pathological processes, making them attractive therapeutic targets. This study aimed to elucidate the molecular basis of galectin–ligand recognition to support the rational design of selective inhibitors. Using high-resolution X-ray crystallography, isothermal titration calorimetry, and site-directed mutagenesis, we investigated the interactions of novel carbohydrate-based ligands with human Gal-1, Gal-3, and Gal-7. Guided by the reference structure of benzyl 3,3′-seleno-digalactoside bound to Gal-3, we designed and synthesized a new series of monosaccharide-based ligands that exploit a Gal-3–specific cation–π interaction between the ligand aromatic moiety and residue R144 within the carbohydrate-binding site. While R144 has previously been recognized as an important determinant of ligand affinity, our results demonstrate that it also plays a crucial role in conferring selectivity over other galectins. Consistently, mutation of R144 to serine significantly reduced ligand affinity and abolished the selectivity advantage, highlighting the dual contribution of this residue to both binding and selectivity. These findings identify R144 as a key structural determinant for the rational design of selective Gal-3 inhibitors.

Human galectin-3 (Gal-3), a β-galactoside binding lectin through its Carbohydrate Recognition Domain (CRD), is implicated in a wide range of cellular functions and is involved in critical biological processes including pathogen recognition, immune response, inflammation and fibrosis. Recently, Gal-3 has gained increasing attention for its role in pathological conditions such as cancer, where it influences cancer growth and progression, inflammatory processes and oxidative stress, opening the search for potential inhibitors. In this context, several naturally derived molecules have attracted particular interest, some of them being used in clinical trials. Here, we used the seeds of the legume Phaseolus coccineus as a green resource for bioactive compounds. The peptide-rich crude extracts were chemically characterized for their peptide and polyphenol contents, as well as their in vitro antioxidant activity, and the powerful obtained extract was tested for biological activities such as cytotoxicity and antioxidant and anti-inflammatory effects on cellular models. Furthermore, the interaction between the crude extract and the CRD of recombinant Gal-3 was verified with the aim of associating its biological effects with the inhibition of Gal-3 activity.

Curcumin is recognized for its diverse biological activities, including the ability to induce apoptosis and ferroptosis. Therefore, it represents a promising candidate for the development of new compounds with neuroprotective and anticancer properties. In order to synthesize mimics with improved pharmacokinetic properties (better solubility and stability than curcumin) here, we present the design and synthesis of novel curcumin analogues named Ethylphosphonate-based curcumin mimics (EPs), which preserve the pharmacophoric features of curcumin. New EP mimics were synthesized by tyrosol- and melatonin-based building blocks using an orthogonal protection approach of the different precursors’ OH functions with good yields and in a few steps. Comparative screenings of the cytotoxic and cytoprotective properties (curcumin was used as a reference compound) were carried out on all new mimics in different cell lines (HeLa, A375, WM266, MDA-MB-231, LX2, and HDF). Assays with inhibitors of ferroptosis (Ferrostatin-1, Fer-1) and apoptosis (Quinoline-Val-Asp-difluorophenoxymethyl ketone, Q-VD), in combination with curcumin, suggested the specific cell death pathway (apoptotic or ferroptotic) of EPs, depending on the aromatic moieties contained in them. Interestingly, EP4 exhibited substantial cytotoxic effects against various human cancer cell lines (HeLa, A375, WM266) while sparing normal cells (HDFs). EP4 displayed a five-times-higher toxicity in triple-negative MDA-MB-231 and LX2 stellate cells than curcumin. The cytotoxicity exerted by EP4 involves only an apoptotic mechanism, contrary to curcumin, which exerts both apoptotic and ferroptotic effects. Additionally, EP4 was also found to be a very potent inhibitor of the ubiquitin-activating enzyme E1, reinforcing the anticancer potential of this compound. Furthermore, EP2 possesses high antioxidant properties, efficiently protects against cell death by ferroptosis, and inhibits the amyloid aggregation involved in AD.

Chimera-type galectin-3 (Gal-3) is a β-galactoside-binding protein containing a single conserved carbohydrate-recognition domain, crucial in fibrosis and carcinogenesis. Selenium-based Gal-3 inhibitors have emerged as promising therapeutic agents, particularly for treating neoplastic diseases. Among them, a seleno-digalactoside (SeDG) substituted with a benzyl group at position 3 of both saccharide residues (benzyl 3,3'-seleno-digalactoside, SeDG-Bn), attracted considerable attention for its selectivity and potent inhibitory efficacy against Gal-3. NMR spectroscopy and molecular dynamics simulations were combined to investigate the binding of SeDG-Bn to Gal-3 at the molecular level. This approach revealed the recognized epitope, the binding mode within Gal-3 binding pocket and enabled the generation of a 3D model of the complex. Our findings show that the presence of a single benzyl group establishes hydrophobic contacts with amino acids in Gal-3 β-sheets S2 and S3, crucially enhancing the binding affinity compared to unmodified SeDG. The digalactose backbone orientation in Gal-3 binding site is partially modified by the benzyl group with respect to complexes with lactosamine and SeDG. These results provide valuable insights into the design of more potent and selective inhibitors for Gal-3, potentially contributing to new therapeutic strategies for conditions such as cancer and fibrosis.

Agri-food waste represents a rich source of bioactive compounds with potential applications in the food, pharmaceutical, and cosmetic industries. This study investigates sustainable strategies for extracting bioactive compounds, with a specific focus on enzyme-assisted extraction (EAE) as an alternative to conventional chemical methods. Spent coffee grounds (SCGs), sunflower residues, and citrus residues were analyzed. Key parameters, including total phenolic content, antioxidant capacity, reducing sugar content, antimicrobial effects, and antiproliferative activity on human cells, were evaluated. SCG and sunflower extracts demonstrated the highest antioxidant content, albeit with minimal variations between enzyme treatments. Notably, enzyme-treated citrus extracts exhibited increased reducing sugar content, significant antimicrobial activity, and inhibition of microbial biofilms. Moreover, citrus extracts were non-toxic to healthy cells while showing pronounced effects against metastatic melanoma cells. These findings underscore the potential of citrus waste as a valuable source of bioactive compounds and highlight EAE as an effective and sustainable extraction technique.

Mycalin A (MA) is a polybrominated C-15 acetogenin isolated from the marine sponge Mycale rotalis. Since this substance displays a strong antiproliferative bioactivity towards some tumour cells, we have now directed our studies towards the elucidation of the MA interactome through functional proteomic approaches, (DARTS and t-LIP-MS). DARTS experiments were performed on Hela cell lysates with the purpose of identifying MA main target protein(s); t-LiP-MS was then applied for an in-depth investigation of the MA–target protein interaction. Both these techniques exploit limited proteolysis coupled with MS analysis. To corroborate LiP data, molecular docking studies were performed on the complexes. Finally, biological and SPR analysis were conducted to explore the effect of the binding. Mortalin (GRP75) was identified as the MA’s main interactor. This protein belongs to the Hsp70 family and has garnered significant attention due to its involvement in certain forms of cancer. Specifically, its overexpression in cancer cells appears to hinder the pro-apoptotic function of p53, one of its client proteins, because it becomes sequestered in the cytoplasm. Our research, therefore, has been focused on the possibility that MA might prevent this sequestration, promoting the re-localization of p53 to the nucleus and facilitating the apoptosis of tumor cells.

Given the significant involvement of galectins in the development of numerous diseases, the aim of the following work is to further study the interaction between galectin-3 (Gal3) and the LPS from Pseudomonas aeruginosa. This manuscript focused on the study of the interaction of the carbohydrate recognition domain of Gal3 with the LPS from Pseudomonas aeruginosa by means of different complementary methodologies, such as circular dichroism; spectrofluorimetry; dynamic and static light scattering and evaluation of the impact of Gal3 on the redox potential membranes of Escherichia coli and P. aeruginosa cells, as well as ITC and NMR studies. This thorough investigation reinforces the hypothesis of an interaction between Gal3 and LPS, unraveling the structural details and providing valuable insights into the formation of these intricate molecular complexes. Taken together, these achievements could potentially prompt the design of therapeutic drugs useful for the development of agonists and/or antagonists for LPS receptors such as galectins as adjunctive therapy for P. aeruginosa.

Galectins, β-galactoside-binding proteins, play relevant roles in different biological processes; therefore, they are becoming emerging targets for diagnostic and therapeutic approaches [...]

Galectins are an ancient family of lectins characterized by the specific binding of b-galactosides through evolutionarily conserved sequence elements of the carbohydrate recognition domain. Interest in this protein family is growing due to the crucial role of galectins not only as therapeutic agents but also as biomarkers of the inflammatory stage occurring in several diseases, including cancer, cardiovascular disease, type 2 diabetes, musculoskeletal disorders, and neurodegenerative diseases. For this reason, the biosensing of galectin becomes crucial for the evaluation of a pathological state as well as for the followup of a therapeutic treatment. The design of biosensors for galectin detection is becoming a reality in recent years, as complementary analytical tools to be exploited at the point of need to support laboratory setup methodologies. This review reports the latest trends in biosensing systems for galectins based on different natural and artificial bioreceptors, integrated into different transduction systems and exploiting nanomaterials to improve analytical performance.

1,2-trans methyl glycosides can be readily obtained from peracetylated sugars through their initial conversion into glycosyl iodide donors and subsequent exposure of these latter to a slight excess of sodium methoxide in methanol. Under these conditions a varied set of mono- and disaccharide precursors afforded the corresponding 1,2-trans glycosides with concomitant de-O-acetylation in satisfying yields (in the range 59-81%). A similar approach also proved effective when using GlcNAc glycosyl chloride as the donor.

Starting from D-xylonolactone and D-ribonolactone, several five-membered bromolactones, related to the C1–C5 portion of mycalin A lactone, have been synthesized. The bromination of D-ribonolactone with HBr/AcOH, without a subsequent transesterification step, has been studied for the first time, giving us most of the acetylated lactones investigated in the present study. For each compound, where possible, both the C-3 alcohol and the corresponding acetate were prepared. Evaluation of their anti-tumor activity showed that all the acetates possess a good cytotoxicity towards human melanoma (A375), human cervical adenocarcinoma (HeLa) and human metastatic melanoma (WM266) cancer cells, comparable or even higher than that displayed by the original mycalin A lactone. Lactone acetates derived from D-ribonolactone showed the higher selectivity of action, exhibiting a strong cytotoxicity on all the tested tumor cells but only a limited toxicity on healthy human dermal fibroblast (HDF) cells, used as a control. Wound healing assays showed that two of these substances inhibit the migration of the WM266 cells.

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.

Galectins (Gals) are small cytosolic proteins that bind β-galactoside residues via their evolutionarily conserved carbohydrate recognition domain. Their dysregulation has been shown to be associated with many diseases. Consequently, targeting galectins for clinical applications has become increasingly relevant to develop tailored inhibitors selectively for one galectin. Accordingly, binding studies providing the molecular details of the interaction between galectin and inhibitor may be useful for the rational design of potent and selective antagonists. Gal-1 and Gal-3 are among the best-studied galectins, mainly for their roles in cancer progression; therefore, the molecular details of their interaction with inhibitors are demanded. This work gains more value by focusing on the interaction between Gal-1 and Gal-3 with the selenylated analogue of the Gal inhibitor thiodigalactose, characterized by a selenoglycoside bond (SeDG), and with unsymmetrical diglycosyl selenides (unsym(Se). Gal-1 and Gal-3 were produced heterologously and biophysically characterized. Interaction studies were performed by ITC, NMR spectroscopy, and MD simulation, and thermodynamic values were discussed and integrated with spectroscopic and computational results. The 3D complexes involving SeDG when interacting with Gal-1 and Gal-3 were depicted. Overall, the collected results will help identify hot spots for the design of new, better performing, and more specific Gal inhibitors.

The N-capping region of an α-helix is a short N-terminal amino acid stretch that contributes to nucleate and stabilize the helical structure. In the VEGF mimetic helical peptide QK, the N-capping region was previously demonstrated to be a key factor of QK helical folding. In this paper, we explored the effect of the chiral inversion of the N-capping sequence on QK folding, performing conformational analysis in solution by circular dichroism and NMR spectroscopy. The effect of such a modification on QK stability in serum and the proliferative effect were also evaluated.

Silibinin is the main biologically active component of silymarin extract and consists of a mixture 1:1 of two diastereoisomeric flavonolignans, namely silybin A (1a) and silybin B (1b), 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 (6aa, 6ab and 6bb) 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 (1a and 1b) and dimers (6aa, 6ab and 6bb) 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 1a and 1b go completely into apoptosis, whereas only part of the cells treated with 6aa and 6ab were found to be in apoptosis.

Galectins are soluble β-D-galactoside-binding proteins whose implication in cancer progression and disease outcome makes them prominent targets for therapeutic intervention. In this frame, the development of small inhibitors that block selectively the activity of galectins represents an important strategy for cancer therapy which is, however, still relatively underdeveloped. To this end, we designed here a rationally and efficiently novel diglycosylated compound, characterized by a selenoglycoside bond and the presence of a lipophilic benzyl group at both saccharide residues. The relatively high binding affinity of the new compound to the carbohydrate recognition domain of two galectins, galectin 3 and galectin 9, its good antiproliferative and anti-migration activity towards melanoma cells, as well as its anti-angiogenesis properties, pave the way for its further development as an anticancer agent.

Although the approved vaccines are proving to be of utmost importance in containing the Coronavirus disease 2019 (COVID-19) threat, they will hardly be resolutive as new severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2, a single-stranded RNA virus) variants might be insensitive to the immune response they induce. In this scenario, developing an effective therapy is still a dire need. Different targets for therapeutic antibodies and diagnostics have been identified, among which the SARS-CoV-2 spike (S) glycoprotein, particularly its receptor-binding domain, has been defined as crucial. In this context, we aim to focus attention also on the role played by the S N-terminal domain (S1-NTD) in the virus attachment, already recognized as a valuable target for neutralizing antibodies, in particular, building on a cavity mapping indicating the presence of two druggable pockets and on the recent literature hypothesizing the presence of a ganglioside-binding domain. In this perspective, we aim at proposing S1-NTD as a putative target for designing small molecules hopefully able to hamper the SARS-CoV-2 attachment to host cells.