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Calixarenes are displaying great potential for the development of new drug delivery systems, diagnostic imaging, biosensing devices and inhibitors of biological processes. In particular, calixarene derivatives are able to interact with many different enzymes and function as inhibitors. By screening of the potential drug target database (PDTD) with a reverse docking procedure, we identify and discuss a selection of 100 proteins that interact strongly with calix[4]arene. We also discover that leucine (23.5 %), isoleucine (11.3 %), phenylalanines (11.3 %) and valine (9.5 %) are the most frequent binding residues followed by hydrophobic cysteines and methionines and aromatic histidines, tyrosines and tryptophanes. Top binders are peroxisome proliferator-activated receptors that already are targeted by commercial drugs, demonstrating the practical interest in calix[4]arene. Nuclear receptors, potassium channel, several carrier proteins, a variety of cancer-related proteins and viral proteins are prominent in the list. It is concluded that calix[4]arene, which is characterized by facile access, well-defined conformational characteristics, and ease of functionalization at both the lower and higher rims, could be a potential lead compound for the development of enzyme inhibitors and theranostic platforms.
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Calixarenos , Simulación del Acoplamiento Molecular , Fenoles , Calixarenos/química , Fenoles/química , Fenoles/farmacología , Humanos , Sitios de Unión , Unión Proteica , Proteínas/química , Proteínas/metabolismoRESUMEN
Au(III) is highly reactive. At odds with its reduced counterpart, Au(I), it is hardly present in structural databases. And yet, it is the starting reactant to form gold nanoclusters (AuNCs) and the constitutive component of a new class of drugs. Its reactivity is a world apart from that of the iso-electronic Pt(II) species. Rather than DNA, it targets proteins. Its interaction with amino acid residues is manifold. It can strongly interact with the residue backbones, amino acid side chains and protein ends, it can form appropriate complexes whose stabilization energy reaches up to more than 40 kcal mol-1, it can affect the pKa of amino acid residues, and it can promote charge transfer from the residues to the amount that it is reduced. Here, quantum chemical calculations provide quantitative information on all the processes where Au(III) can be involved. A myriad of structural arrangements are examined in order to determine the strongest interactions and quantify the amount of charge transfer between protonated and deprotonated residues and Au(III). The calculated interaction energies of the amino acid side chains with Au(III) quantitatively reproduce the experimental tendency of Au(III) to interact with selenocysteine, cysteine and histidine and negatively charged amino acids such as Glu and Asp. Also, aromatic residues such as tyrosine and tryptophan strongly interact with Au(III). In proteins, basic pH plays a role in the deprotonation of cysteine, lysine and tyrosine and strongly increases the binding affinity of Au(III) toward these amino acids. The amino acid residues in the protein can also trigger the reduction of Au(III) ions. Sulfur-containing amino acids (cysteine and methionine) and selenocysteine provide almost one electron to Au(III) upon binding. Tyrosine also shows a considerable tendency to act as a reductant. Other amino acids, commonly identified in Au-protein adducts, such as Ser, Trp, Thr, Gln, Glu, Asn, Asp, Lys, Arg and His, possess a notable reducing power toward Au(III). These results and their discussion form a vade mecum that can find application in medicinal chemistry and nanotech applications of Au(III).
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Aminoácidos , Oro , Oro/química , Aminoácidos/química , Nanotecnología/métodos , Nanopartículas del Metal/química , Química FarmacéuticaRESUMEN
Filters made of graphene oxide (GO) are promising for purification of water and selective sieving of specific ions; while some results indicate the ionic radius as the discriminating factor in the sieving efficiency, the exact mechanism of sieving is still under debate. Furthermore, most of the reported GO filters are planar coatings with a simple geometry and an area much smaller than commercial water filters. Here, we show selective transport of different ions across GO coatings deposited on standard hollow fiber filters with an area >10 times larger than typical filters reported. Thanks to the fabrication procedure, we obtained a uniform coating on such complex geometry with no cracks or holes. Monovalent ions like Na+ and K+ can be transported through these filters by applying a low electric voltage, while divalent ions are blocked. By combining transport and adsorption measurements with molecular dynamics simulations and spectroscopic characterization, we unravel the ion sieving mechanism and demonstrate that it is mainly due to the interactions of the ions with the carboxylate groups present on the GO surface at neutral pH.
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Photodynamic therapy (PDT) represents an emerging strategy to treat various malignancies, including colorectal cancer (CC), the third most common cancer type. This work presents an engineered M13 phage retargeted towards CC cells through pentavalent display of a disulfide-constrained peptide nonamer. The M13CC nanovector was conjugated with the photosensitizer Rose Bengal (RB), and the photodynamic anticancer effects of the resulting M13CC-RB bioconjugate were investigated on CC cells. We show that upon irradiation M13CC-RB is able to impair CC cell viability, and that this effect depends on i) photosensitizer concentration and ii) targeting efficiency towards CC cell lines, proving the specificity of the vector compared to unmodified M13 phage. We also demonstrate that M13CC-RB enhances generation and intracellular accumulation of reactive oxygen species (ROS) triggering CC cell death. To further investigate the anticancer potential of M13CC-RB, we performed PDT experiments on 3D CC spheroids, proving, for the first time, the ability of engineered M13 phage conjugates to deeply penetrate multicellular spheroids. Moreover, significant photodynamic effects, including spheroid disruption and cytotoxicity, were readily triggered at picomolar concentrations of the phage vector. Taken together, our results promote engineered M13 phages as promising nanovector platform for targeted photosensitization, paving the way to novel adjuvant approaches to fight CC malignancies.
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Bacteriófagos , Neoplasias del Colon , Fotoquimioterapia , Humanos , Fármacos Fotosensibilizantes/farmacología , Fármacos Fotosensibilizantes/uso terapéutico , Fotoquimioterapia/métodos , Muerte Celular , Rosa Bengala/farmacología , Rosa Bengala/química , Neoplasias del Colon/terapiaRESUMEN
The "on-demand" capture and utilization of CO2 is effectively realized with a readily accessible dual function organic composite. The covalent and controlled derivatization of graphene oxide (GO) surface with naturally occurring arginine led to a "smart" material capable of capturing (chemisorption) CO2 from high-purity flue-gas as well as low-concentration streams (i. e. direct air capture) and concomitant chemical activation toward the incorporation into cyclic carbonates. The overall integrated CO2 capture and conversion (ICCC) strategy has been fully elucidated mechanistically via dedicated computational, spectroscopic and thermal analyses.
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Carboranes have emerged as one of the most promising boron agents in boron neutron capture therapy (BNCT). In this context, in vivo studies are particularly relevant, since they provide qualitative and quantitative information about the biodistribution of these molecules, which is of the utmost importance to determine the efficacy of BNCT, defining their localization and (bio)accumulation, as well as their pharmacokinetics and pharmacodynamics. First, we gathered a detailed list of the carboranes used for in vivo studies, considering the synthesis of carborane derivatives or the use of delivery system such as liposomes, micelles and nanoparticles. Then, the formulation employed and the cancer model used in each of these studies were identified. Finally, we examined the analytical aspects concerning carborane detection, identifying the main methodologies applied in the literature for ex vivo and in vivo analysis. The present work aims to identify the current strengths and weakness of the use of carboranes in BNCT, establishing the bottlenecks and the best strategies for future applications.
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Growing antibiotic resistance has encouraged the revival of phage-inspired antimicrobial approaches. On the other hand, photodynamic therapy (PDT) is considered a very promising research domain for the protection against infectious diseases. Yet, very few efforts have been made to combine the advantages of both approaches in a modular, retargetable platform. Here, we foster the M13 bacteriophage as a multifunctional scaffold, enabling the selective photodynamic killing of bacteria. We took advantage of the well-defined molecular biology of M13 to functionalize its capsid with hundreds of photo-activable Rose Bengal sensitizers and contemporarily target this light-triggerable nanobot to specific bacterial species by phage display of peptide targeting moieties fused to the minor coat protein pIII of the phage. Upon light irradiation of the specimen, the targeted killing of diverse Gram(-) pathogens occurred at subnanomolar concentrations of the phage vector. Our findings contribute to the development of antimicrobials based on targeted and triggerable phage-based nanobiotherapeutics.
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Chlorin e6 (Ce6) and fullerene (C60) are among the most used photosensitizers (PSs) for photodynamic therapy (PDT). Through the combination of the chemical and photophysical properties of Ce6 and C60, in principle, we can obtain an "ideal" photosensitizer that is able to bypass the limitations of the two molecules alone, i.e., the low cellular uptake of Ce6 and the scarce solubility and absorption in the red region of the C60. Here, we synthesized and characterized a Ce6-C60 dyad. The UV-Vis spectrum of the dyad showed the typical absorption bands of both fullerene and Ce6, while a quenching of Ce6 fluorescence was observed. This behavior is typical in the formation of a fullerene-antenna system and is due to the intramolecular energy, or electron transfer from the antenna (Ce6) to the fullerene. Consequently, the Ce6-C60 dyad showed an enhancement in the generation of reactive oxygen species (ROS). Flow cytometry measurements demonstrated how the uptake of the Ce6 was strongly improved by the conjugation with C60. The Ce6-C60 dyad exhibited in A431 cancer cells low dark toxicity and a higher PDT efficacy than Ce6 alone, due to the enhancement of the uptake and the improvement of ROS generation.
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PURPOSE: Lumican is a major extracellular matrix (ECM) component in the cornea that is upregulated after injury and promotes corneal wound healing. We have recently shown that peptides designed based on the 13 C-terminal amino acids of lumican (LumC13 and LumC13C-A) are able to recapitulate the effects of lumican on promoting corneal wound healing. Herein we used computational chemistry to develop peptide mimetics derived from LumC13C-A with increased stability and half-life that are biologically active and non-toxic, thereby promoting corneal wound healing with increased pharmacological potential. METHODS: Different peptides staples were rationalized using LumC13C-A sequence by computational chemistry, docked to TGFßRI and the interface binding energies compared. Lowest scoring peptides were synthesized, and the toxicity of peptides tested using CCK8-based cell viability assay. The efficacy of the stapled peptides at promoting corneal wound healing was tested using a proliferation assay, an in vitro scratch assay using human corneal epithelial cells and an in vivo murine corneal debridement wound healing model. RESULTS: Binding free energies were calculated using MMGBSA algorithm, and peptides LumC13C and LumC13S5 displayed superior binding to ALK5 compared to the non-stapled peptide LumC13C-A. The presence of the hydrocarbon staple in LumC13C enhances the stability of the α-helical conformation, thereby facilitating more optimal interactions with the ALK5 receptor. The stapled peptides do not present cytotoxic effects on human corneal epithelial cells at a 300 nM concentration. Similar to lumican and LumC13C-A, both C13C and LumC13S5 significantly promote corneal wound healing both in vitro and in vivo. CONCLUSIONS: Highly stable and non-toxic stapled peptides designed based on LumC13, significantly promote corneal wound healing. As a proof of principle, our data shows that more stable and pharmacologically relevant peptides can be designed based on endogenous peptide sequences for treating various corneal pathologies.
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Lesiones de la Cornea , Epitelio Corneal , Humanos , Animales , Ratones , Lumican/metabolismo , Lumican/farmacología , Córnea/patología , Lesiones de la Cornea/metabolismo , Cicatrización de Heridas , Péptidos/farmacología , Péptidos/metabolismo , Epitelio Corneal/metabolismoRESUMEN
Alkylation of one of the inner-core nitrogen atoms is one possible approach to obtain dianionic corrole ligands, suitable for the coordination of divalent metal ions, such as PdII . Inner-core N-methylation can be obtained by treating the corrole with CH3 I, but the reaction conditions should be optimized to limit the formation of the dimethylated derivative. Two regioisomers, the N-21 and the N-22 methyl derivatives are obtained from the reaction, with the first product achieved in a higher amount. Structural characterization of the reaction products evidenced the distortion induced by the introduction of the methyl groups; the N-methylcorroles are chiral compounds, and the enantiomers were separated by chromatography, with their absolute configuration assigned by ECD computation. Palladium insertion was achieved in the case of monosubstituted corroles, but not with the dimethylated macrocycle; X-ray characterization of the complexes showed the distortion of the macrocycles. The Pd complexes do not show luminescence emission, but are able to produce singlet oxygen upon irradiation. The PdII complexes were also inserted in human serum albumin (HSA) and dispersed in water; in this case, the protein protects the corroles from photobleaching, and a switch from the type II to the type I mechanism in reactive oxygen species (ROS) production is observed.
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We report on the synthesis of ß-cyclodextrin (ßCD) modified graphene oxide (GO) nanosheets, having different sized alkyl linkers (GO-Cn -ßCD) and their exploitation as sorbent of per- and polyfluoroalkyl substances (PFAS) from drinking water. ßCD were functionalized with a pending amino group, and the resulting precursors grafted to GO nanosheets by epoxide ring opening reaction. Loading of ßCD units in the range 12 %-36 % was estimated by combined XPS and elemental analysis. Adsorption tests on perfluorobutanoic acid (PFBA), a particularly persistent PFAS selected as case study, revealed a strong influence of the alkyl linker length on the adsorption efficiency, with the hexyl linker derivative GO-C6 -ßCD outperforming both pristine GO and granular activated carbon (GAC), the standard sorbent benchmark. Molecular dynamic simulations ascribed this evidence to the favorable orientation of the ßCD unit on the surface of GO which enables a strong contaminant molecules retention.
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Semiconducting single walled carbon nanotubes (SWCNTs) are promising materials for biosensing applications with electrolyte-gated transistors (EGT). However, to be employed in EGT devices, SWCNTs often require lengthy solution-processing fabrication techniques. Here, we introduce a simple solution-based method that allows fabricating EGT devices from stable dispersions of SWCNTs/bovine serum albumin (BSA) hybrids in water. The dispersion is then deposited on a substrate allowing the formation of a SWCNTs random network as the semiconducting channel. We demonstrate that this methodology allows the fabrication of EGT devices with electric performances that allow their use in biosensing applications. We demonstrate their application for the detection of cortisol in solution, upon gate electrode functionalization with anti-cortisol antibodies. This is a robust and cost-effective methodology that sets the ground for a SWCNT/BSA-based biosensing platform that allows overcoming many limitations of standard SWCNTs biosensor fabrications.
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Técnicas Biosensibles , Nanotubos de Carbono , Albúmina Sérica Bovina , Técnicas Biosensibles/métodos , ElectrólitosRESUMEN
Carboranes are promising agents for applications in boron neutron capture therapy (BNCT), but their hydrophobicity prevents their use in physiological environments. Here, by using reverse docking and molecular dynamics (MD) simulations, we identified blood transport proteins as candidate carriers of carboranes. Hemoglobin showed a higher binding affinity for carboranes than transthyretin and human serum albumin (HSA), which are well-known carborane-binding proteins. Myoglobin, ceruloplasmin, sex hormone-binding protein, lactoferrin, plasma retinol-binding protein, thyroxine-binding globulin, corticosteroid-binding globulin and afamin have a binding affinity comparable to transthyretin/HSA. The carborane@protein complexes are stable in water and characterized by favorable binding energy. The driving force in the carborane binding is represented by the formation of hydrophobic interactions with aliphatic amino acids and BH-π and CH-π interactions with aromatic amino acids. Dihydrogen bonds, classical hydrogen bonds and surfactant-like interactions also assist the binding. These results (i) identify the plasma proteins responsible for binding carborane upon their intravenous administration, and (ii) suggest an innovative formulation for carboranes based on the formation of a carborane@protein complex prior to the administration.
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Introduction: Thanks to recent advances in synthetic methodology, water-soluble fullerene nanomaterials that interfere with biomolecules, especially DNA/RNA and selected proteins, have been found with tremendous potential for applications in nanomedicine. Herein, we describe the synthesis and evaluation of a water-soluble glycine-derived [60]fullerene hexakisadduct (HDGF) with T h symmetry, which is a first-in-class BTK protein inhibitor. Methods: We synthesized and characterized glycine derived [60]fullerene using NMR, ESI-MS, and ATR-FT-IR. DLS and zeta potential were measured and high-resolution transmission electron microscopy (HRTEM) observations were performed. The chemical composition of the water-soluble fullerene nanomaterial was examined by X-ray photoelectron spectrometry. To observe aggregate formation, the cryo-TEM analysis was carried out. The docking studies and molecular dynamic simulations were performed to determine interactions between HDGF and BTK. The in vitro cytotoxicity was evaluated on RAJI and K562 blood cancer cell lines. Subsequently, we examined the induction of cell death by autophagy and apoptosis by determining the expression levels of crucial genes and caspases. We investigated the direct association of HDGF on inhibition of the BTK signalling pathway by examining changes in the calcium levels in RAJI cells after treatment. The inhibitory potential of HDGF against non-receptor tyrosine kinases was evaluated. Finally, we assessed the effects of HDGF and ibrutinib on the expression of the BTK protein and downstream signal transduction in RAJI cells following anti-IgM stimulation. Results: Computational studies revealed that the inhibitory activity of the obtained [60]fullerene derivative is multifaceted: it hampers the BTK active site, interacting directly with the catalytic residues, rendering it inaccessible to phosphorylation, and binds to residues that form the ATP binding pocket. The anticancer activity of produced carbon nanomaterial revealed that it inhibited the BTK protein and its downstream pathways, including PLC and Akt proteins, at the cellular level. The mechanistic studies suggested the formation of autophagosomes (increased gene expression of LC3 and p62) and two caspases (caspase-3 and -9) were responsible for the activation and progression of apoptosis. Conclusion: These data illustrate the potential of fullerene-based BTK protein inhibitors as nanotherapeutics for blood cancer and provide helpful information to support the future development of fullerene nanomaterials as a novel class of enzyme inhibitors.
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Antineoplásicos , Fulerenos , Neoplasias Hematológicas , Neoplasias , Humanos , Fulerenos/farmacología , Fulerenos/química , Agammaglobulinemia Tirosina Quinasa/metabolismo , Espectroscopía Infrarroja por Transformada de Fourier , Agua , Antineoplásicos/química , Inhibidores de Proteínas Quinasas/farmacología , Inhibidores de Proteínas Quinasas/química , Caspasas , GlicinaRESUMEN
Chlorin e6 (Ce6) is among the most used sensitizers in photodynamic (PDT) and sonodynamic (SDT) therapy; its low solubility in water, however, hampers its clinical exploitation. Ce6 has a strong tendency to aggregate in physiological environments, reducing its performance as a photo/sono-sensitizer, as well as yielding poor pharmacokinetic and pharmacodynamic properties. The interaction of Ce6 with human serum albumin (HSA) (i) governs its biodistribution and (ii) can be used to improve its water solubility by encapsulation. Here, using ensemble docking and microsecond molecular dynamics simulations, we identified the two Ce6 binding pockets in HSA, i.e., the Sudlow I site and the heme binding pocket, providing an atomistic description of the binding. Comparing the photophysical and photosensitizing properties of Ce6@HSA with respect to the same properties regarding the free Ce6, it was observed that (i) a red-shift occurred in both the absorption and emission spectra, (ii) a maintaining of the fluorescence quantum yield and an increase of the excited state lifetime was detected, and (iii) a switch from the type II to the type I mechanism in a reactive oxygen species (ROS) production, upon irradiation, took place.
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Clorofilidas , Fotoquimioterapia , Porfirinas , Humanos , Albúmina Sérica Humana/metabolismo , Fármacos Fotosensibilizantes/química , Distribución Tisular , Porfirinas/química , Fotoquimioterapia/métodos , Línea Celular TumoralRESUMEN
Temoporfin (mTHPC) is one of the most promising photosensitizers used in photodynamic therapy (PDT). Despite its clinical use, the lipophilic character of mTHPC still hampers the full exploitation of its potential. Low solubility in water, high tendency to aggregate, and low biocompatibility are the main limitations because they cause poor stability in physiological environments, dark toxicity, and ultimately reduce the generation of reactive oxygen species (ROS). Applying a reverse docking approach, here, we identified a number of blood transport proteins able to bind and disperse monomolecularly mTHPC, namely apohemoglobin, apomyoglobin, hemopexin, and afamin. We validated the computational results synthesizing the mTHPC-apomyoglobin complex (mTHPC@apoMb) and demonstrated that the protein monodisperses mTHPC in a physiological environment. The mTHPC@apoMb complex preserves the imaging properties of the molecule and improves its ability to produce ROS via both type I and type II mechanisms. The effectiveness of photodynamic treatment using the mTHPC@apoMb complex was then demonstrated in vitro. Blood transport proteins can be used as molecular "Trojan horses" in cancer cells by conferring mTHPC (i) water solubility, (ii) monodispersity, and (iii) biocompatibility, ultimately bypassing the current limitations of mTHPC.
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The combination of photodynamic therapy with chemotherapy (photochemotherapy, PCT) can lead to additive or synergistic antitumor effects. Usually, two different molecules, a photosensitizer (PS) and a chemotherapeutic drug are used in PCT. Doxorubicin is one of the most successful chemotherapy drugs. Despite its high efficacy, two factors limit its clinical use: severe side effects and the development of chemoresistance. Doxorubicin is a chromophore, able to absorb light in the visible range, making it a potential PS. Here, we exploited the intrinsic photosensitizing properties of doxorubicin to enhance its anticancer activity in leukemia, breast, and epidermoid carcinoma cells, upon irradiation. Light can selectively trigger the local generation of reactive oxygen species (ROS), following photophysical pathways. Doxorubicin showed a concentration-dependent ability to generate peroxides and singlet oxygen upon irradiation. The underlying mechanisms leading to the increase in its cytotoxic activity were intracellular ROS generation and the induction of necrotic cell death. The nuclear localization of doxorubicin represents an added value for its use as a PS. The use of doxorubicin in PCT, simultaneously acting as a chemotherapeutic agent and a PS, may allow (i) an increase in the anticancer effects of the drug, and (ii) a decrease in its dose, and thus, its dose-related adverse effects.
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Antineoplásicos , Fotoquimioterapia , Especies Reactivas de Oxígeno/metabolismo , Doxorrubicina/farmacología , Antineoplásicos/farmacología , Fármacos Fotosensibilizantes/farmacologíaRESUMEN
Fullerenes are considered excellent photosensitizers, being highly suitable for photodynamic therapy (PDT). A lack of water solubility and low biocompatibility are, in many instances, still hampering the full exploitation of their potential in nanomedicine. Here, we used human serum albumin (HSA) to disperse fullerenes by binding up to five fullerene cages inside the hydrophobic cavities. Albumin was bioconjugated with folic acid to specifically address the folate receptors that are usually overexpressed in several solid tumors. Concurrently, tetramethylrhodamine isothiocyanate, TRITC, a tag for imaging, was conjugated to C60@HSA in order to build an effective phototheranostic platform. The in vitro experiments demonstrated that: (i) HSA disperses C60 molecules in a physiological environment, (ii) HSA, upon C60 binding, maintains its biological identity and biocompatibility, (iii) the C60@HSA complex shows a significant visible-light-induced production of reactive oxygen species, and (iv) folate bioconjugation improves both the internalization and the PDT-induced phototoxicity of the C60@HSA complex in HeLa cells.
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The chemical functionalization of polysaccharides to obtain functional materials has been of great interest in the last decades. This traditional synthetic approach has drawbacks, such as changing the crystallinity of the material or altering its morphology or texture. These modifications are crucial when a biogenic matrix is exploited for its hierarchical structure. In this work, the use of lectins and carbohydrate-binding proteins as supramolecular linkers for polysaccharide functionalization is proposed. As proof of concept, a deproteinized squid pen, a hierarchically-organized ß-chitin matrix, was functionalized using a dye (FITC) labeled lectin; the lectin used was the wheat germ agglutinin (WGA). It has been observed that the binding of this functionalized protein homogenously introduces a new property (fluorescence) into the ß-chitin matrix without altering its crystallographic and hierarchical structure. The supramolecular functionalization of polysaccharides with protein/lectin molecules opens up new routes for the chemical modification of polysaccharides. This novel approach can be of interest in various scientific fields, overcoming the synthetic limits that have hitherto hindered the technological exploitation of polysaccharides-based materials.
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Lectinas , Polisacáridos , Quitina , Lectinas/metabolismo , Lectinas de Plantas , Aglutininas del Germen de Trigo/química , Aglutininas del Germen de Trigo/metabolismoRESUMEN
Lysine-covalently modified graphene oxide (GO-Lys) was prepared by an innovative procedure. Lysine brushes promote enhanced adsorption of bisphenol A, benzophenone-4 and carbamazepine contaminants from tap water, with a removal capacity beyond the state of the art.