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In recent years, glycomics have shown how pervasive the role of carbohydrates in biological systems is and how chemical tools are essential to investigate glycan function and modulate carbohydrate-mediated processes. Biomimetic receptors for carbohydrates can carry out this task but, although significant affinities and selectivities toward simple saccharides have been achieved, targeting complex glycoconjugates remains a goal yet unattained. In this work we report the unprecedented recognition of a complex biantennary sialylglycopeptide (SGP) by a tweezers-shaped biomimetic receptor, which selectively binds to the core GlcNAc2 disaccharide of the N-glycan with an affinity of 170 μM. Because of the simple structure and the remarkable binding ability, this biomimetic receptor can represent a versatile tool for glycoscience, opening the way to useful applications. We thank MIUR-Italy “Progetto Dipartimenti di Eccellenza 2018–2022” allocated to Department of Chemistry Ugo Schiff, COST Action (CA18132), MIUR-Italy PRIN2017 (2017XZ2ZBK) for granting a fellowship to F.M. and Ente Cassa di Risparmio di Firenze (Italy) is acknowledged for granting an ITC nanocalorimeter and a high-field NMR spectrometer. Open Access funding provided by Università degli Studi di Firenze within the CRUI-CARE Agreement.

Fully synthetic tumor-associated carbohydrate antigen (TACA)-based vaccines are a promising strategy to treat cancer. To overcome the intrinsic low immunogenicity of TACAs, the choice of the antigens' analogues and multivalent presentation have been proved to be successful. Here, we present the preparation, characterization, and

Hyaluronic acid (HA) is a naturally occurring polysaccharide with many molecular functions, including maintaining the structure and physiology of the tissues, tissue remodeling, and inflammation. HA is found naturally in physiological tear fluid, possesses excellent mucus-layer-adhesive properties, and is successfully employed in the treatment of dry eye syndrome (DES). However, HA has as major drawback: its rapidiin vivo/idegradation by hyaluronidase. We report on a unique material, namely, HA-b3/b, obtained by the functionalization of HA with the metalloproteinase inhibitorb3/b(MMPI). This material is characterized by an increased resistance to hyaluronidase degradation, associated with MMP inhibition properties. The ability of HA-b3/bto prevent dehydration of human corneal epithelial cellsiin vitro/iandiin vivo/imay accelerate the development of more efficient DES treatment and broaden the application of HA in human diseases.

GlcNAc2 is the core disaccharide fragment present in N‐glycans exposed on the surface of enveloped viruses of high health concern, such as coronaviruses. Because N‐glycans are directly involved in the docking of viruses to host cells, recognition of GlcNAc2 by a biomimetic receptor may be a convenient alternative to the use of lectins to interfere with viral entry and infection. Herein, we describe a simple biomimetic receptor recognizing the methyl‐β‐glycoside of GlcNAc2 in water with an unprecedented affinity of 160 μM, exceeding that of more structurally complex receptors reported in the literature. The tweezers‐shaped acyclic structure exhibits marked selectivity among structurally related disaccharides, and complete discrimination between mono‐ and disaccharides. Molecular modelling calculations supported by NOE data provided a three‐dimensional description of the binding mode, shedding light on the origin of the affinities and selectivities exhibited by the receptor., Think simple. In the development of biomimetic receptors for carbohydrates, an approach that focuses on simple acyclic architectures can be successfully exploited for disaccharide recognition, provided that the correct binding geometry can be achieved. Here we present a simple tweezers‐shaped receptor that selectively recognizes in water GlcNAc2, the core fragment of N‐glycans exposed on the surface of enveloped viruses, such as coronaviruses.

The "pancarcinoma" Tn antigen (αGalNAc-O-Ser/Thr) is a tumor-associated carbohydrate antigen (TACA) overexpressed on the surface of cancer cells and suitable target for anticancer vaccines. However, TACAs commonly show weak immunogenicity, low in vivo stability, and poor bioavailability. To address these issues, the development of physiologically stable TACA synthetic mimetics and novel nanocarriers for multivalent display are object of intense research. Nanomaterials represent suitable scaffolds to multimerize antigens, but absence of toxicity, easy functionalization and capability to incorporate biomolecules are compulsory characteristics for vaccine nanocarriers. Here, we report on the conjugation of a synthetic Tn-antigen mimetic to biocompatible and water-dispersible dextran-based single-chain nanoparticles (DXT-SCPNs). In vitro stimulation of PBMCs and analysis of interleukins production indicated a specific innate immune modulation mediated by the multivalent presentation of the Tn mimetic at the nanoparticle surface. These preliminary results pave the way for the development of Tn-mimetic clusters on biocompatible DXT-SCPN for TACA-based vaccines.

Abnormal glycoconjugates have distinctly been recognized as potential biomarkers for cancer diagnosis. A great deal of attention has been focused on Tn antigen, an oversimplified mucin-1 O-glycan, over-expressed in different cancers. Herein, we investigate the possibility to replace the use of anti-Tn monoclonal antibodies with an innovative class of catecholamine-based Molecularly Imprinted Polymers (MIPs), emerging in recent years as promising tools for bioanalytical applications. MIPs are synthetic receptors characterized by high sensitivity and specificity towards the imprinted target. Here, original polynorepinephrine-based MIPs coupled to Surface Plasmon Resonance biosensing for Tn antigen recognition are reported. We have verified the imprinting and binding capacity of these MIPs towards very small antigenic entities, represented by the natural Tn antigen and the TnThr mimetic 1 (conjugated to BSA or linked to a MUC1 hexapeptide analogue), and compared the biosensor performances with an anti-Tn monoclonal antibody. The results clearly display the effectiveness of the pursued imprinting strategies.

The selective recognition of caffeine in water among structurally related xanthines and purine or pyrimidine bases was achieved by a simple tweezer-shaped receptor featuring sulfonate hydrosolubilizing groups. The remarkable affinity for caffeine, among the highest reported thus far in the literature and larger than that shown by adenosine receptors of all subtypes, stems from a synergistic combination of hydrogen bonding, CH-pi, and pi-stacking interactions.

Cellulose is the most abundant natural biopolymer and owing to its compatibility with biological tissues, it is considered a versatile starting material for developing new and sustainable materials from renewable resources. With the advent of drug-resistance among pathogenic microorganisms, recent strategies have focused on the development of novel treatment options and alternative antimicrobial therapies, such as antimicrobial photodynamic therapy (aPDT). This approach encompasses the combination of photoactive dyes and harmless visible light, in the presence of dioxygen, to produce reactive oxygen species that can selectively kill microorganisms. Photosensitizers for aPDT can be adsorbed, entrapped, or linked to cellulose-like supports, providing an increase in the surface area, with improved mechanical strength, barrier, and antimicrobial properties, paving the way to new applications, such as wound disinfection, sterilization of medical materials and surfaces in different contexts (industrial, household and hospital), or prevention of microbial contamination in packaged food. This review will report the development of porphyrinic photosensitizers supported on cellulose/cellulose derivative materials to achieve effective photoinactivation. A brief overview of the efficiency of cellulose based photoactive dyes for cancer, using photodynamic therapy (PDT), will be also discussed. Particular attention will be devoted to the synthetic routes behind the preparation of the photosensitizer-cellulose functional materials.

Here, two conformationally constrained sialyl analogues were synthesized and characterized in their interaction with the inhibitory Siglec, human CD22 (h-CD22). An orthogonal approach, including biophysical assays (SPR and fluorescence), ligand-based NMR techniques, and molecular modelling, was employed to disentangle the interaction mechanisms at a molecular level. The results showed that the Sialyl-TnThr antigen analogue represents a promising scaffold for the design of novel h-CD22 inhibitors. Our findings also suggested that the introduction of a biphenyl moiety at position 9 of the sialic acid hampers the canonical accommodation of the ligand in the protein binding pocket, even though the affinity with respect to the natural ligand is increased. Our outcomes address the search of novel modifications of the Neu5Ac-a(2-6)-Gal epitope, outlining new hints for the design and synthesis of high-affinity h-CD22 ligands, offering new prospects for therapeutic intervention to prevent autoimmune diseases and B-cells malignancies.

Pairing glycans with tissue lectins controls multiple effector pathways in (patho)physiology. A clinically relevant example is the prodegradative activity of galectins-1 and -3 (Gal-1 and -3) in the progression of osteoarthritis (OA) via matrix metalloproteinases (MMPs), especially MMP-13. The design of heterobifunctional inhibitors that can block galectin binding and MMPs both directly and by preventing their galectin-dependent induction selectively offers a perspective to dissect the roles of lectins and proteolytic enzymes. We describe the synthesis of such a reagent with a bivalent galectin ligand connected to an MMP inhibitor and of two tetravalent glycoclusters with a subtle change in headgroup presentation for further elucidation of influence on ligand binding. Testing was performed on clinical material with mixtures of galectins as occurring in vivo, using sections of fixed tissue. Two-colour fluorescence microscopy monitored binding to the cellular glycome after optimization of experimental parameters. In the presence of the inhibitor, galectin binding to OA specimens was significantly reduced. These results open the perspective to examine the inhibitory capacity of custom-made ditopic compounds on binding of lectins in mixtures using sections of clinical material with known impact of galectins and MMPs on disease progression.

Publsher's version (útgefin grein), Therapeutic vaccination offers great promise as an intervention for a diversity of infectious and non-infectious conditions. Given that most chronic health conditions are thought to have an immune component, vaccination can at least in principle be proposed as a therapeutic strategy. Understanding the nature of protective immunity is of vital importance, and the progress made in recent years in defining the nature of pathological and protective immunity for a range of diseases has provided an impetus to devise strategies to promote such responses in a targeted manner. However, in many cases, limited progress has been made in clinical adoption of such approaches. This in part results from a lack of safe and effective vaccine adjuvants that can be used to promote protective immunity and/or reduce deleterious immune responses. Although somewhat simplistic, it is possible to divide therapeutic vaccine approaches into those targeting conditions where antibody responses can mediate protection and those where the principal focus is the promotion of effector and memory cellular immunity or the reduction of damaging cellular immune responses as in the case of autoimmune diseases. Clearly, in all cases of antigen-specific immunotherapy, the identification of protective antigens is a vital first step. There are many challenges to developing therapeutic vaccines beyond those associated with prophylactic diseases including the ongoing immune responses in patients, patient heterogeneity, and diversity in the type and stage of disease. If reproducible biomarkers can be defined, these could allow earlier diagnosis and intervention and likely increase therapeutic vaccine efficacy. Current immunomodulatory approaches related to adoptive cell transfers or passive antibody therapy are showing great promise, but these are outside the scope of this review which will focus on the potential for adjuvanted therapeutic active vaccination strategies., This article/publication is based upon work from COST Action CA16231 ENOVA (European Network of Vaccine Adjuvants), supported by COST (European Cooperation in Science and Technology—www.cost.eu).

The fully characterization of tumor associated antigens (TAAs) and of tumor associated carbohydrate antigens (TACAs) have opened the avenue of cancer immunotherapy. The intrinsic poor immunogenicity of TACAs, however, spotlighted the importance of multivalent presentation of the antigen(s) to trigger an immune response. Nanoparticles are excellent scaffolds for this purpose. Here we reported on the easy glycosylation of iron-based and biocompatible dextran-based nanoparticles with 1, a mimetic of the TnThr antigen. The multivalent presentation of 1 induced the induction of TNF-α and IL-6/IL10, respectively. The multivalent glycosylation of silica nanoparticles (GSiNPs) was also performed and saccharide loading qualitative assessed by solid state NMR. Our results offer the proof of concept that biomolecules coating can also be investigated on solid system by NMR.

The present paper describes the functionalization of sodium hyaluronate (NaHA) with a small molecule (2-((N-(6-aminohexyl)-4-methoxyphenyl)sulfonamido)-N-hydroxyacetamide) (MMPI) having proven inhibitory activity against membrane metalloproteins involved in inflammatory processes (i.e. MMP12). The obtained derivative (HA-MMPI) demonstrated an increased resistance to the in-vitro degradation by hyaluronidase, viscoelastic properties close to those of healthy human synovial fluid, cytocompatibility towards human chondrocytes and nanomolar affinity towards MMP 12. Thus, HA-MMPI can be considered a good candidate as viscosupplement in the treatment of knee osteoarticular disease.

When facing the dilemma of following a preorganized or adaptive design approach in conceiving the architecture of new biomimetic receptors for carbohydrates, shape‐persistent macrocyclic structures were most often chosen to achieve effective recognition of neutral saccharides in water. In contrast, acyclic architectures have seldom been explored, even though potentially simpler and more easily accessible. In this work, comparison of the binding properties of two structurally related diaminocarbazolic receptors, featuring a macrocyclic and an acyclic tweezer‐shaped architecture, highlighted the advantages provided by the acyclic receptor in terms of selectivity in the recognition of 1,4‐disaccharides of biological interest. Selective recognition of GlcNAc2, the core fragment of N‐glycans exposed on the surface of enveloped viruses, stands as an emblematic example. NMR spectroscopic data and molecular modeling calculations were used to ascertain the differences in binding mode and to shed light on the origin of recognition efficacy and selectivity., Adaptive to be selective: Comparison of the binding properties of two structurally related biomimetic receptors toward disaccharides sheds light on the differences between a rigid macrocyclic architecture and a flexible tweezer‐shaped structure showing, counterintuitively, that the adaptive structure can be advantageous in terms of selectivity. NMR spectroscopy and molecular modeling calculations were used to provide a 3D description of the receptor‐disaccharide complexes in solution and explaining the origin of the observed selectivity.

Understanding how the Severe Acute Respiratory Syndrome CoronaVirus 2 (SARS-CoV-2) highjacks epithelial cells and infiltrates organs and tissues is essential for developing strategies against this highly contagious virus. Currently, SARS-CoV-2 is thought to evade innate antiviral immunity, undergo endocytosis, and fuse with the host cell membrane by exploiting ACE2 receptors. Both, virus and ACE2, are highly glycosylated and the role of glycans in the above steps seems to be essential. Indeed, exploiting O-linked and N-linked glycans exposed on the membrane of SARS-CoV-2 as binding sites for ACE2 represents a virus strategy for infecting the human host. We unprecedentedly found that, taking advantage of small abiotic molecules capable of binding to glycans of the spike protein, binding of the latter to hACE2 can be inhibited and the viral infection can be largely neutralized. Results obtained in this work with a lead structure may open the way to the development of SARS-CoV-2 therapeutic candidates. Funding Information: Ministero dell’Universita e della Ricerca (Miur) of Italy (project n. FISR2020IP_01095). Declaration of Interests: The authors declare no competing interests.

International audience; The monosaccharide Tn and the disaccharide STn are tumor antigens with similar structures and common biosynthetic pathways. Both are always over-expressed simultaneously on tumor cell surfaces. We report herein the efficient synthesis of the STnThr antigen analogue 2, featuring the immunogenic TnThr mimetic 1 aglycon. Analogously to the native STn, 2 is recognized by the influenza N1 neuraminidase. A model of the N1·2 complex showed the sialyl moiety of 2 well nested in the active site pocket, with docking unaffected by the rigid aglycon. The analogue 2 is, therefore, in association with mimetic 1, a good determinant for the design of new multiantigen cancer vaccines.

Two orthogonal, metal free click reactions, enabled to glycosylate ubiquitin and its mutant A28C forming two protein scaffolds with high affinity for BambL, a lectin from the human pathogen Burkholderia ambifaria. A new fucoside analogue, with high affinity with BambL, firstly synthetized and co-crystallized with the protein target, provided the insights for sugar determinants grafting onto ubiquitin. Three ubiquitin-based glycosides were thus assembled. Fuc-Ub, presented several copies of the fucoside analogue, with proper geometry for multivalent effect; Rha-A28C, displayed one thio-rhamnose, known for its ability to tuning the immunological response; finally, Fuc-Rha-A28C, included both multiple fucoside analogs and the rhamnose residue. Fuc-Ub and Fuc-Rha-A28C ligands proved high affinity for BambL and unprecedented immune modulatory properties towards macrophages activation., Metal free click reactions used to glycosylate ubiquitin and its mutant A28C afforded two protein scaffolds with high affinity for Burkholderia ambifaria lectin (BambL).

Coupling of sulfonyl fluorides, fluorosulfates and iminosulfur oxydifluorides with proteins gave the corresponding sulfonamide, sulfamate and sulfamide adducts under mild conditions.

GM3-ganglioside is known to be involved in melanoma proliferation. In order to modulate metastatic-related events, we have functionalized multi-walled carbon nanotubes (MWCNTs) with multiple copies of a GM3-lactone mimetic. The MWCNTs proved to guarantee the appropriate spatial arrangement of the mimetic allowing a stronger inhibition of migration and invasiveness of human melanoma (A375) cells compared to other multivalent constructs reported before. In addition, the effect of the multivalent tubular conjugate on the inhibition of specific tyrosine kinases, which are associated with the ganglioside complexes within the membrane domains, was demonstrated. Finally, the short-term fate of the conjugate was assessed, for the first time, by means of the 1H NMR relaxometry technique by exploiting the signal arising from the CNTs.

Previous studies proved the effectiveness of an intravenous PEGylated liposomal formulation of opiorphin (1mg/mL) in protecting the drug from enzymatic degradation, and improving intensity and duration of its painkilling effect. Therefore, considering the advantages of nasal administration, the aim of this work was the development of a liposomal mucoadhesive thermo-sensitive in situ gel for the extended nasal delivery of opiorphin. With this purpose, the potential of a series of combinations of different polymers (i.e. chitosan, hydroxypropylmethylcellulose, Poloxamer, Carbopol) in forming solutions able to rapidly gel at the nasal cavity temperature (34 °C) has been investigated. The best formulations were further characterized for gel strength and mucoadhesion properties. The selected formulation, composed by Poloxamer 407 (26.5%) and Carbopol 934P (1%), showed short gelation time at 34 °C (10s) and suitable mucoadhesion duration (5.5h) and strength (27g/cm2). Due to the low volume administrable via the nasal route, a concentrated liposomal formulation of the peptide (16.5mg/mL) was developed and loaded in the selected in situ gel formulation. Ex-vivo permeation studies, by excised nasal porcine mucosa, showed that the liposomal hydrogel formulation enabled a sustained and controlled delivery of opiorphin over more than 5h, and highlighted the role of the liposomal carrier in enhancing up to 6 times permeability coefficient and permeation rate of the peptide through the lipophilic nasal mucosa compared to a free peptide-loaded gel.

We report on the electrochemistry of a peptide molecular wire as a conductive support for ligand immobilization in biosensing assays. The helical 9-mer peptide, tagged at Nterminus with Methylene Blue (MB) and thiol-functionalized at Cterminus was anchored on a gold surface via gold/thiol chemistry. The helical peptide acts as molecular wire, mediating the two-step electron transfer (ET) from MB to the gold surface. The forward and backward square wave voltametric (SWV) components recorded in the presence of peptide wire were used to estimate the kinetic parameters of the electrode reaction. The simulated data matched the experimental ones for the two-step sequential surface mechanism (EE), with the rate constant of the first step ksur,1 = 20 s−1 and the cathodic ET coefficient α1 = 0.55. The kinetic parameters of the second step were ksur,2 > 1000 s−1, and α2 = 0.5. Small ligands for high-molecular weight targets can be grafted on the peptide wire between the MB tag and the surface through covalent bonding. The binding of the target to the peptide-anchored ligand hampers the ET transfer from MB to the electrode surface, causing a decrease of the peak current. These findings were used further to develop an electrochemical peptide-based biosensor with signaling-off interrogation. The biosensor was tested against two relevant targets for medical diagnosis: the anti-tumor-associated carbohydrate antigen (α-Tn) antibody and the growth hormone secretagogue receptor (GHS-R1a), after ligand grafting. Both targets were detected in the nanomolar range with an overall assay time of 10 min.

Oxaliplatin (OXA) is a valuable and largely used cancer drug which induces a serious and intractable neuropathy. The lipoyl-homotaurine derivative (ADM_12) reverts in vivo OXA-induced neuropathy, and it is an effective antagonist of the nociceptive sensor channel TRPA1. Unprecedentedly, this safe analgesic showed a synergy with OXA in vitro and proved to inhibit CA IX, a relevant therapeutic target, clearly interfering with pancreatic cancer cells' aggressiveness.

[Image: see text] The HIV-1 nucleocapsid protein 7 (NC) is a potential target for effective antiretroviral therapy due to its central role in virus replication, mainly linked to nucleic acid (NA) chaperone activity, and low susceptibility to drug resistance. By screening a compounds library, we identified the aminopyrrolic compound CN14_17, a known carbohydrate binding agent, that inhibits the NC chaperone activity in the low micromolar range. Different from most of available NC inhibitors, CN14_17 fully prevents the NC-induced annealing of complementary NA sequences. Using fluorescence assays and isothermal titration calorimetry, we found that CN14_17 competes with NC for the binding to NAs, preferentially targeting single-stranded sequences. Molecular dynamics simulations confirmed that binding to cTAR occurs preferably within the guanosine-rich single stranded sequence. Finally, CN14_17 exhibited antiretroviral activity in the low micromolar range, although with a moderate therapeutic index. Overall, CN14_17 might be the progenitor of a new promising class of NC inhibitors.

The transient receptor potential akyrin type-1 (TRPA1) is a non-selective cation channel playing a pivotal role in pain sensation and neurogenic inflammation. TRPA1 channels expressed in the central nervous system (CNS) have a critical role in the modulation of cortical spreading depression (CSD), which is a key pathophysiological basis of migraine pain. ADM_09 is a recently developed lipoic acid-based TRPA1 antagonist that is able to revert oxaliplatin-induced neuropathic pain and inflammatory trigeminal allodynia. In this context, aiming at developing drugs that are able to target TRPA1 channels in the CNS and promote an antioxidant effect, permeability across the blood&ndash, brain barrier (BBB) represents a central issue. Niosomes are nanovesicles that can be functionalized with specific ligands selectively recognized by transporters expressed on the BBB. In this work, the activity of ADM_09 on neocortex cultures was studied, and an efficient formulation to cross the BBB was developed with the aim of increasing the concentration of ADM_09 into the brain and selectively delivering it to the CNS rapidly after parenteral administration.

The choice between adaptive and preorganized architectures, or of the most effective hydrogen bonding groups to be selected, are dilemmas that supramolecular chemists must address in designing synthetic receptors for such a challenging guest as carbohydrates. In this paper, structurally related architectures featuring two alternative hydrogen bonding motifs were compared to ascertain the structural and functional origin of their binding differences and the advantages that can be expected in monosaccharide recognition. A set of structurally related macrocyclic receptors were prepared, and their binding properties were measured by NMR and ITC techniques in chloroform vs a common saccharidic target, namely, the β-octyl glycoside of D-glucose. Results showed that the diaminocarbazolic motif, recently reported as the constituting unit of highly effective receptors for saccharides in water, is a superior hydrogen bonding motif compared to the previously described diaminopyrrolic motif, which was successfully employed in molecular recognition of carbohydrates in polar organic solvents, due to intrinsic structural and functional factors, rather than to hydrophobic contributions. In addition, the occurrence of a rare example of a thermodynamic template effect exerted by the beta-glucoside has been ascertained, enhancing the synthesis outcome of the otherwise low yielding preparation of the described macrocyclic receptors.

In the last two decades, the paramount importance of Tumor Associated Carbohydrate Antigens (TACAs) as targets for anticancer vaccine development has been firmly assessed. The Tn antigen is an ideal target for immunotherapy, in that it is masked on normal cells, but exposed on cancer cells. However, it is difficult to elicit an effective and long-lasting response against Tn antigen and other TACAs. Here we report on the Tn antigen analogues developed to boost the latent Tn immune response. Hopefully, the results reported herein will be of help for the rational design of effective TACA-based immunostimulants.

Caffeine is a competitive inhibitor of adenosine receptors and possesses wide pharmacological activity. Artificial receptors recognizing caffeine potentially have a wide range of biomedical and industrial applications. Herein, we describe two structurally related and readily available artificial receptors: 1) a macrocyclic receptor, which binds caffeine with the unprecedented affinity of 9.3 microM, though with poor selectivity; and 2) a tweezers-like structure, showing an affinity of 26 microM and a 4.5-fold and 6-fold selectivity toward theophylline and theobromine, respectively. Binding affinities were measured by 1H NMR titrations and were confirmed by isothermal titration calorimetry. The X-ray structure of the complex between caffeine and the acyclic receptor revealed the origin of the recognition, explained the selectivity, and shed light on the role of hydrogen bonding and CH-pi/pi-pi interactions.

The design and synthesis of the Ln3+ complexes of a DOTA-containing (DOTA=1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid) inhibitor of matrix metalloproteinases are reported. The tight binding of the sulfonamide scaffold to the catalytic domain of the investigated matrix metalloproteinase is not impaired by the presence of the Ln3+ -DOTA moiety. The paramagnetic properties of the Ln3+ complex are exploited to obtain insights into the structural features of the ligand-protein interactions and to evaluate the influence of the linker length on the quality of the paramagnetic restraints.

Transient receptor potential ankyrin type-1 (TRPA1) channels are known to actively participate in different pain conditions, including trigeminal neuropathic pain, whose clinical treatment is still unsatisfactory. The aim of this study was to evaluate the involvement of TRPA1 channels by means of the antagonist ADM_12 in trigeminal neuropathic pain, in order to identify possible therapeutic targets. A single treatment of ADM_12 in rats 4 weeks after the chronic constriction injury of the infraorbital nerve (IoN-CCI) significantly reduced the mechanical allodynia induced in the IoN-CCI rats. Additionally, ADM_12 was able to abolish the increased levels of TRPA1, calcitonin gene-related peptide (CGRP), substance P (SP), and cytokines gene expression in trigeminal ganglia, cervical spinal cord, and medulla induced in the IoN-CCI rats. By contrast, no significant differences between groups were seen as regards CGRP and SP protein expression in the pars caudalis of the spinal nucleus of the trigeminal nerve. ADM_12 also reduced TRP vanilloid type-1 (TRPV1) gene expression in the same areas after IoN-CCI. Our findings show the involvement of both TRPA1 and TRPV1 channels in trigeminal neuropathic pain, and in particular, in trigeminal mechanical allodynia. Furthermore, they provide grounds for the use of ADM_12 in the treatment of trigeminal neuropathic pain.

[Image: see text] The formation of amorphous protein aggregates containing human superoxide dismutase (hSOD1) is thought to be involved in amyotrophic lateral sclerosis onset. cis-Platin inhibits the oligomerization of apo hSOD1, but its toxicity precludes any possible use in therapy. Herein, we propose a less toxic platinum complex, namely oxa/cis-platin, as hSOD1 antiaggregation lead compound. Oxa/cis-platin is able to interact with hSOD1 in the disulfide oxidized apo form by binding cysteine 111 (Cys111). The mild neurotoxic phenomena induced in vitro and in vivo by oxa/cis-platin can be successfully reverted by using lypoyl derivatives, which do not interfere with the antiaggregation properties of the platin derivative.

Protein glycosylation is the most complex post-translational modification process. More than 50 % of human cells proteins are glycosylated, whereas bacteria such as E. coli do not have this modification machinery. Indeed, the carbohydrate residues in natural proteins affect their folding, immunogenicity, and stability toward proteases, besides controlling biological properties and activities. It is therefore important to introduce such structural modification in bioengineered proteins lacking the presence of carbohydrate residues. This is not trivial as it requires reagents and conditions compatible with the protein's stability and reactivity. This work reports on the introduction of lactose moieties in two natural proteins, namely ubiquitin (Ub) and l-asparaginase II (ANSII). The synthetic route employed is based on the sulfur(VI) fluoride exchange (SuFEx) coupling of a lactose tethered arylfluorosulfate (Lact-Ar-OSO2 F) with the ϵ-NH2 group of lysine residues of the proteins. This metal-free click SuFEx reaction relies on the properties of the fluorosulfate employed, which is easily prepared in multigram scale from available precursors and reacts chemoselectively with the ϵ-NH2 group of lysine residues under mild conditions. Thus, iterative couplings of Lact-Ar-OSO2 F to Ub and ANSII, afforded multiple glycosylations of these proteins so that up to three and four Lact-Ar-OSO2 groups were introduced in Ub and ANSII, respectively, via the formation of a sulfamoyl (OSO2 -NH) linkage.

Antibiotic resistance in the opportunistic pathogen Pseudomonas aeruginosa is partly caused by biofilms forming a physical barrier to antibiotic penetration. Here we focused on modifying tetravalent glycopeptide dendrimer ligands of P. aeruginosa lectins LecB or LecA to increase their biofilm inhibition activity. First heteroglycoclusters were investigated displaying one pair each of LecB specific fucosyl groups and LecA specific galactosyl groups and binding simultaneously to both lectins, one of which gave the first fully resolved crystal structure of a peptide dendrimer as LecB complex providing a structural model for dendrimer-lectin interactions (PDB ; 5D2A). Biofilm inhibition was increased by introducing additional cationic residues in these dendrimers but resulted in bactericidal effects similar to those of non-glycosylated polycationic antimicrobial peptide dendrimers. In a second approach dendrimers displaying four copies of the natural LecB ligand Lewisa were prepared leading to slightly stronger LecB binding and biofilm inhibition. Finally synergistic application of a LecB specific non-bactericidal antibiofilm dendrimer with the antibiotic tobramycin at sub-inhibitory concentrations of both compounds allowed effective biofilm inhibition and dispersal.

Cycloadduct 1 is a conformationally constrained mimetic of the tumor-associated Tn antigen. It maintains the 4C1 conformation and the α-O-glycosidic linkage of the natural epitopes. Due to its tricyclic structure, the anomeric linkage is constrained and its conformation “frozen”. Using saturation transfer difference NMR spectroscopy experiments, epitope mapping for binding by three lectins [i.e., from Erythrina cristagalli (ECL), human macrophages (MGL), and from Helix pomatia (HPA)] was carried out. Striking differences in the epitope viewed from the ligand's perspective were revealed by this comparison. Evidently, the structural change around the C-2 position has a major impact, if the contact pattern to the ligand is not mostly restricted to galacto configuration with the axial hydroxyl group at C-4, in combination with C-3/C-6. These measurements thus provide insights into actual contacts, which help predict applicability as an inhibitor for distinct lectins, and as an elicitor of specific antibodies.

The binding abilities of a set of structurally related aminopyrrolic synthetic receptors for mannosides, endowed with antimycotic activity against yeast and yeast-like pathogens bearing mannoproteins on their cell surface, have been investigated towards the highly mannosylated gp120 and gp41 glycoproteins of the HIV envelope. A pronounced binding interaction with both glycoproteins was observed by SPR for most of the investigated compounds. Comparison of their binding properties towards the glycoproteins with their binding affinities toward mannosides revealed a direct correlation, supporting their role as carbohydrate binding agents (CBAs). Cytostatic activity studies revealed antiproliferative activity dependent on the nature and the structure of compounds. Antiviral activity studies against a broad panel of DNA and RNA viruses showed inhibitory effect against HIV infection of the T-lymphocyte CEM cell line for two compounds, suggesting antiviral activity similar to other CBAs, such as the nonpeptidic pradimicin antibiotics.

International audience; Bacterial and fungal pathogens involved in lung infection in cystic fibrosis patients utilize a particular family of glycan-binding proteins, characterized by the presentation of six fucose-binding sites on a ring shape scaffold. These lectins are attractive targets for anti-infectious compounds that could interfere in the recognition of host tissues by pathogens. The design of a cyclopeptide-based hexavalent structure allowed for the presentation of six fucose residues. The synthetic hexavalent compound displays appropriate geometry resulting in high avidity binding by lectins from Aspergillus fumigatus and Burkholderia ambifaria. Replacing the fucose residue with a conformationally constrained fucomimetic does not alter the affinity and provide fine specificity, with no binding to other fucose-specific lectins.

Carbohydrate recognition in water by biomimetic receptors is an attractive, but very challenging goal. Despite advances achieved in glucose recognition, little or no success has been obtained in the recognition of other saccharidic epitopes of paramount importance in biological processes. Herein, the unprecedented recognition of fucose in water by an artificial receptor that shows affinities closely comparable to those of several lectins is reported. The receptor has been constructed by assembling a hydrogen-bonding element (carbazole), a hydrophobic aromatic moiety (anthracene), and a water-solubilising function (phosphonate) into a macrocyclic structure to provide the appropriate binding geometry. The described receptor binds fucose with sub-millimolar affinity in water at physiological pH; this shows that enthalpic binding can be ascribed to hydrogen bonding to saccharidic hydroxy groups and to CH-π interactions between the sugar backbone and aromatic moieties. Experimental NOE contacts coupled to conformational search calculations return a picture of a binding site in which fucose assumes a staggered orientation reminiscent of that shown by fucose when bound to the Ralstonia solanacearum lectin (RSL).

We describe a simple method to prepare water dispersible core-shell CdSe/ZnS quantum dots (QDs) 1 by capping QDs with a new thiol-containing heterobifunctional dicarboxylic ligand 4 (DHLA-EDADA). This ligand, obtained on a gram scale through a few synthetic steps, provides a compact layer on the QDs, whose hydrodynamic size in H2O is 15 nm ± 3 nm. The colloidal stability is dramatically enhanced with respect to the well-known (±) α-lipoic acid (DHLA). The ligand affinity towards QDs and the water dispersibility of nanocrystals 1 are addressed by the dithiol groups of DHLA, which chelate the zinc of the shell, and by the dicarboxylic groups of the ethylenediamine-N,N-diacetic acid (EDADA) residue, respectively. The effects of pH, buffer solutions, and biological medium on the stability of QDs 1 were assessed by monitoring the photoluminescence (PL) and hydrodynamic size over time. Highly fluorescent QD dispersions, stable over extended periods of time and over broad pH ranges and buffer types, were obtained. Furthermore, we show that the DHLA-EDADA ligand 4 also endows QDs with functional groups suitable for further conjugation and for metal ion detection. As a case study to illustrate the potential of our approach, we report the preparation and characterization of a highly luminescent orange light emitting polymer-QD 1 composite film.

Inflammation of the trigeminal nerve is considered one of the most painful conditions known to humankind. The diagnosis is often difficult; moreover, safe and effective pharmacological treatments are lacking. A new molecule, ADM_12, formed by a lipoic and omotaurine residues covalently linked, is here reported. In vitro and in vivo tests showed that ADM_12 is a very attractive original compound presenting (i) a remarkable safety profile; (ii) a high binding constant versus TRPA1; (iii) an intriguing behavior versus TRPV1; and (iv) the ability to significantly and persistently reduce mechanical facial allodynia in rats. Noteworthy, by testing ADM_12, we shed light on the unprecedented involvement of TRPA1 and TRPV1 channels in orofacial pain.

The ability of a rigid α-Tn mimetic (compound 1) to activate murine invariant natural killer T (iNKT) and human natural killer (NK) cells, two subsets of lymphocytes involved in cancer immunesurveillance, was investigated. For this purpose, the mimetic 1 was properly conjugated to a stearic acid containing glycerol-based phospholipid (compound 5) to be presented, in the context of the conserved non polymorphic major histocompatibility complex class I-like molecules (CD1d), to iNKT cells. On the contrary, the mimetic 1 was conjugated to a multivalent peptide-based scaffold (compound 6) to induce NK cell activation.