Unveiling the Role of Capping Groups in Naphthalene N-Capped Dehydrodipeptide Hydrogels.

Self-assembled peptide-based hydrogels are archetypical nanostructured materials with a plethora of foreseeable applications in nanomedicine and as biomaterials. N-protected di- and tri-peptides are effective minimalist (molecular) hydrogelators. Independent variation of the capping group, peptide sequence and side chain modifications allows a wide chemical space to be explored and hydrogel properties to be tuned. In this work, we report the synthesis of a focused library of dehydrodipeptides N-protected with 1-naphthoyl and 2-naphthylacetyl groups. The 2-naphthylacetyl group was extensively reported for preparation of peptide-based self-assembled hydrogels, whereas the 1-naphthaloyl group was largely overlooked, owing presumably to the lack of a methylene linker between the naphthalene aromatic ring and the peptide backbone. Interestingly, dehydrodipeptides N-capped with the 1-naphthyl moiety afford stronger gels, at lower concentrations, than the 2-naphthylacetyl-capped dehydrodipeptides. Fluorescence and circular dichroism spectroscopy showed that the self-assembly of the dehydrodipeptides is driven by intermolecular aromatic π-π stacking interactions. Molecular dynamics simulations revealed that the 1-naphthoyl group allows higher order aromatic π-π stacking of the peptide molecules than the 2-naphthylacetyl group, together with hydrogen bonding of the peptide scaffold. The nanostructure of the gel networks was studied by TEM and STEM microscopy and was found to correlate well with the elasticity of the gels. This study contributes to understanding the interplay between peptide and capping group structure on the formation of self-assembled low-molecular-weight peptide hydrogels. Moreover, the results presented here add the 1-naphthoyl group to the palette of capping groups available for the preparation of efficacious low-molecular-weight peptide-based hydrogels.

Fisetin derivatives exhibit enhanced anti-inflammatory activity and modulation of endoplasmic reticulum stress.

Inflammation and endoplasmic reticulum (ER) stress are often hand in hand in the context of chronic disease. Both are activated upon perceived disturbances in homeostasis, being deleterious when intensely or chronically activated. Fisetin (FST) is a dietary flavonol that is known to possess multiple relevant bioactivities, raising the question of its potential health benefits and even its use in novel pharmacological approaches against ER stress and inflammation. To attain this prospect, some limitations to this molecule, namely its poor bioavailability and solubility, must be addressed. In an attempt to improve the biological properties of the parent molecule, we have synthesized a set of FST derivatives. These new molecules were tested along with the original compound for their ability to mitigate the activation of the signaling pathways underlying inflammation and ER stress. By reducing LPS-induced nuclear factor-kappa B (NF-κB) activation, cytokine release, inflammasome activation and reactive oxygen species (ROS) generation, FST has proven to be effective against the onset of inflammation. The molecule also decreases the activation of the unfolded protein response (UPR), as evidenced by the reduced expression of relevant UPR-related genes upon ER stress induction. Some of the tested derivatives are novel inhibitors of targets associated to inflammation and ER stress signaling, in some cases more potent than the parent compound. Furthermore, the reduced cytotoxicity of some of these molecules enabled the use of higher concentrations than that of FST, resulting in the observation of enhanced bioactivities.

Peptide-Based Supramolecular Hydrogels as Drug Delivery Agents: Recent Advances.

Supramolecular peptide hydrogels have many important applications in biomedicine, including drug delivery applications for the sustained release of therapeutic molecules. Targeted and selective drug administration is often preferential to systemic drug delivery, as it can allow reduced doses and can avoid the toxicity and side-effects caused by off-target binding. New discoveries are continually being reported in this rapidly developing field. In this review, we report the latest developments in supramolecular peptide-based hydrogels for drug delivery, focusing primarily on discoveries that have been reported in the last four years (2018-present). We address clinical points, such as peptide self-assembly and drug release, mechanical properties in drug delivery, peptide functionalization, bioadhesive properties and drug delivery enhancement strategies, drug release profiles, and different hydrogel matrices for anticancer drug loading and release.

Aryl-Capped Lysine-Dehydroamino Acid Dipeptide Supergelators as Potential Drug Release Systems.

Employing amino acids and peptides as molecular building blocks provides unique opportunities for generating supramolecular hydrogels, owing to their inherent biological origin, bioactivity, biocompatibility, and biodegradability. However, they can suffer from proteolytic degradation. Short peptides (<8 amino acids) attached to an aromatic capping group are particularly attractive alternatives for minimalistic low molecular weight hydrogelators. Peptides with low critical gelation concentrations (CGCs) are especially desirable, as the low weight percentage required for gelation makes them more cost-effective and reduces toxicity. In this work, three dehydrodipeptides were studied for their self-assembly properties. The results showed that all three dehydrodipeptides can form self-standing hydrogels with very low critical gelation concentrations (0.05−0.20 wt%) using a pH trigger. Hydrogels of all three dehydrodipeptides were characterised by scanning tunnelling emission microscopy (STEM), rheology, fluorescence spectroscopy, and circular dichroism (CD) spectroscopy. Molecular modelling was performed to probe the structural patterns and interactions. The cytotoxicity of the new compounds was tested using human keratinocytes (HaCaT cell line). In general, the results suggest that all three compounds are non-cytotoxic, although one of the peptides shows a small impact on cell viability. In sustained release assays, the effect of the charge of the model drug compounds on the rate of cargo release from the hydrogel network was evaluated. The hydrogels provide a sustained release of methyl orange (anionic) and ciprofloxacin (neutral), while methylene blue (cationic) was retained by the network.

An injectable, naproxen-conjugated, supramolecular hydrogel with ultra-low critical gelation concentration-prepared from a known folate receptor ligand.

Short peptides capped on the N-terminus with aromatic groups are often able to form supramolecular hydrogels-self-assembled networks of fibrils able to trap water molecules. Typically, these hydrogelators can form stiff gels at concentrations of 0.1 to 1.0 wt%-i.e. they consist of mainly water. The properties of these soft materials mimic those of the extracellular matrix (ECM) of biological tissue and therefore they have found many biomedical uses in tissue engineering, wound healing, drug delivery, biosensing and bioprinting applications. In drug delivery strategies related to cancer therapy, injectable hydrogels can serve as a depot formulation, where a sustained release of the chemotherapeutic from near the tumour site allows reduced doses and, therefore, decreased side effects. To further target cancer cells, folic acid-conjugated hydrogels and nanostructures are often sought, to exploit the overexpression of folate receptors on cancer cells-an approach which can allow the selective cellular uptake of an encapsulated drug. In this present study, two known dipeptide folate receptor ligands (1 and 2) recently identified from a screen of a DNA-encoded compound library, were synthesised and investigated for their hydrogelation ability and cytotoxicity. Compound 1, containing a naproxen capping group, rapidly forms hydrogels at concentrations as low as 0.03 wt%-one of the lowest critical gelation concentrations (CGCs) known for a supramolecular hydrogelator. In contrast, compound 2, which contains a 3-indolepropionic acid capping group, was unable to form hydrogels under a range of conditions and concentrations, instead forming nanospheres with diameters of 0.5 µm. Hydrogels of 1 were characterised by STEM microscopy, rheology, fluorescence spectroscopy and circular dichroism. Both compounds 1 and 2 had no impact on the proliferation of kerotinocytes (HaCaT cells) at concentrations up to 100 µM. Compound 1, containing the NSAID, was tested for anti-inflammatory activity in a human cyclooxygenase-1/2 model. The rate of the release of model drug compounds from within hydrogels of 1 was also investigated.

Initial Screening of Poly(ethylene glycol) Amino Ligands for Affinity Purification of Plasmid DNA in Aqueous Two-Phase Systems.

Gene therapy and DNA vaccination are among the most expected biotechnological and medical advances for the coming years. However, the lack of cost-effective large-scale production and purification of pharmaceutical-grade plasmid DNA (pDNA) still hampers their wide application. Downstream processing, which is mainly chromatography-based, of pDNA remains the key manufacturing step. Despite its high resolution, the scaling-up of chromatography is usually difficult and presents low capacity, resulting in low yields. Alternative methods that are based on aqueous two-phase systems (ATPSs) have been studied. Although higher yields may be obtained, its selectivity is often low. In this work, modified polymers based on poly(ethylene glycol) (PEG) derivatisation with amino groups (PEG-amine) or conjugation with positively charged amino acids (PEG-lysine, PEG-arginine, and PEG-histidine) were studied to increase the selectivity of PEG-dextran systems towards the partition of a model plasmid. A two-step strategy was employed to obtain suitable pure formulations of pDNA. In the first step, a PEG-dextran system with the addition of the affinity ligand was used with the recovery of the pDNA in the PEG-rich phase. Then, the pDNA was re-extracted to an ammonium-sulphate-rich phase in the second step. After removing the salt, this method yielded a purified preparation of pDNA without RNA and protein contamination.

Complexes of Bifunctional DO3A-N-(α-amino)propinate Ligands with Mg(II), Ca(II), Cu(II), Zn(II), and Lanthanide(III) Ions: Thermodynamic Stability, Formation and Dissociation Kinetics, and Solution Dynamic NMR Studies.

The thermodynamic, kinetic, and structural properties of Ln3+ complexes with the bifunctional DO3A-ACE4- ligand and its amide derivative DO3A-BACE4- (modelling the case where DO3A-ACE4- ligand binds to vector molecules) have been studied in order to confirm the usefulness of the corresponding Gd3+ complexes as relaxation labels of targeted MRI contrast agents. The stability constants of the Mg2+ and Ca2+ complexes of DO3A-ACE4- and DO3A-BACE4- complexes are lower than for DOTA4- and DO3A3-, while the Zn2+ and Cu2+ complexes have similar and higher stability than for DOTA4- and DO3A3- complexes. The stability constants of the Ln(DO3A-BACE)- complexes increase from Ce3+ to Gd3+ but remain practically constant for the late Ln3+ ions (represented by Yb3+). The stability constants of the Ln(DO3A-ACE)4- and Ln(DO3A-BACE)4- complexes are several orders of magnitude lower than those of the corresponding DOTA4- and DO3A3- complexes. The formation rate of Eu(DO3A-ACE)- is one order of magnitude slower than for Eu(DOTA)-, due to the presence of the protonated amine group, which destabilizes the protonated intermediate complex. This protonated group causes the Ln(DO3A-ACE)- complexes to dissociate several orders of magnitude faster than Ln(DOTA)- and its absence in the Ln(DO3A-BACE)- complexes results in inertness similar to Ln(DOTA)- (as judged by the rate constants of acid assisted dissociation). The 1H NMR spectra of the diamagnetic Y(DO3A-ACE)- and Y(DO3A-BACE)- reflect the slow dynamics at low temperatures of the intramolecular isomerization process between the SA pair of enantiomers, R-Λ(λλλλ) and S-Δ(δδδδ). The conformation of the Cα-substituted pendant arm is different in the two complexes, where the bulky substituent is further away from the macrocyclic ring in Y(DO3A-BACE)- than the amino group in Y(DO3A-ACE)- to minimize steric hindrance. The temperature dependence of the spectra reflects slower ring motions than pendant arms rearrangements in both complexes. Although losing some thermodynamic stability relative to Gd(DOTA)-, Gd(DO3A-BACE)- is still quite inert, indicating the usefulness of the bifunctional DO3A-ACE4- in the design of GBCAs and Ln3+-based tags for protein structural NMR analysis.

Bolaamphiphilic Bis-Dehydropeptide Hydrogels as Potential Drug Release Systems.

The self-assembly of nanometric structures from molecular building blocks is an effective way to make new functional materials for biological and technological applications. In this work, four symmetrical bolaamphiphiles based on dehydrodipeptides (phenylalanyldehydrophenylalanine and tyrosyldehydrophenylalanine) linked through phenyl or naphthyl linkers (terephthalic acid and 2,6-naphthalenedicarboxylic acid) were prepared, and their self-assembly properties were studied. The results showed that all compounds, with the exception of the bolaamphiphile of tyrosyldehydrophenylalanine and 2,6-naphthalene dicarboxylic acid, gave self-standing hydrogels with critical gelation concentrations of 0.3 wt % and 0.4 wt %, using a pH trigger. The self-assembly of these hydrogelators was investigated using STEM microscopy, which revealed a network of entangled fibers. According to rheology, the dehydrodipeptide bolaamphiphilic hydrogelators are viscoelastic materials with an elastic modulus G' that falls in the range of native tissue (0.37 kPa brain-4.5 kPa cartilage). In viability and proliferation studies, it was found that these compounds were non-toxic toward the human keratinocyte cell line, HaCaT. In sustained release assays, we studied the effects of the charge present on model drug compounds on the rate of cargo release from the hydrogel networks. Methylene blue (MB), methyl orange (MO), and ciprofloxacin were chosen as cationic, anionic, and overall neutral cargo, respectively. These studies have shown that the hydrogels provide a sustained release of methyl orange and ciprofloxacin, while methylene blue is retained by the hydrogel network.

Evaluation of a Model Photo-Caged Dehydropeptide as a Stimuli-Responsive Supramolecular Hydrogel.

Short peptides capped on the N-terminus with aromatic groups are often able to form supramolecular hydrogels, via self-assembly, in aqueous media. The rheological properties of these readily tunable hydrogels resemble those of the extracellular matrix (ECM) and therefore have potential for various biological applications, such as tissue engineering, biosensors, 3D bioprinting, drug delivery systems and wound dressings. We herein report a new photo-responsive supramolecular hydrogel based on a "caged" dehydropeptide (CNB-Phe-ΔPhe-OH 2), containing a photo-cleavable carboxy-2-nitrobenzyl (CNB) group. We have characterized this hydrogel using a range of techniques. Irradiation with UV light cleaves the pendant aromatic capping group, to liberate the corresponding uncaged model dehydropeptide (H-Phe-ΔPhe-OH 3), a process which was investigated by 1H NMR and HPLC studies. Crucially, this cleavage of the capping group is accompanied by dissolution of the hydrogel (studied visually and by fluorescence spectroscopy), as the delicate balance of intramolecular interactions within the hydrogel structure is disrupted. Hydrogels which can be disassembled non-invasively with temporal and spatial control have great potential for specialized on-demand drug release systems, wound dressing materials and various topical treatments. Both 2 and 3 were found to be non-cytotoxic to the human keratinocyte cell line, HaCaT. The UV-responsive hydrogel system reported here is complementary to previously reported related UV-responsive systems, which are generally composed of peptides formed from canonical amino acids, which are susceptible to enzymatic proteolysis in vivo. This system is based on a dehydrodipeptide structure which is known to confer proteolytic resistance. We have investigated the ability of the photo-activated system to accelerate the release of the antibiotic, ciprofloxacin, as well as some other small model drug compounds. We have also conducted some initial studies towards skin-related applications. Moreover, this model system could potentially be adapted for on-demand "self-delivery", through the uncaging of known biologically active dehydrodipeptides.

Dehydropeptide Supramolecular Hydrogels and Nanostructures as Potential Peptidomimetic Biomedical Materials.

Supramolecular peptide hydrogels are gaining increased attention, owing to their potential in a variety of biomedical applications. Their physical properties are similar to those of the extracellular matrix (ECM), which is key to their applications in the cell culture of specialized cells, tissue engineering, skin regeneration, and wound healing. The structure of these hydrogels usually consists of a di- or tripeptide capped on the N-terminus with a hydrophobic aromatic group, such as Fmoc or naphthalene. Although these peptide conjugates can offer advantages over other types of gelators such as cross-linked polymers, they usually possess the limitation of being particularly sensitive to proteolysis by endogenous proteases. One of the strategies reported that can overcome this barrier is to use a peptidomimetic strategy, in which natural amino acids are switched for non-proteinogenic analogues, such as D-amino acids, ß-amino acids, or dehydroamino acids. Such peptides usually possess much greater resistance to enzymatic hydrolysis. Peptides containing dehydroamino acids, i.e., dehydropeptides, are particularly interesting, as the presence of the double bond also introduces a conformational restraint to the peptide backbone, resulting in (often predictable) changes to the secondary structure of the peptide. This review focuses on peptide hydrogels and related nanostructures, where α,ß-didehydro-α-amino acids have been successfully incorporated into the structure of peptide hydrogelators, and the resulting properties are discussed in terms of their potential biomedical applications. Where appropriate, their properties are compared with those of the corresponding peptide hydrogelator composed of canonical amino acids. In a wider context, we consider the presence of dehydroamino acids in natural compounds and medicinally important compounds as well as their limitations, and we consider some of the synthetic strategies for obtaining dehydropeptides. Finally, we consider the future direction for this research area.

Impact of Citrate and Lipid-Functionalized Magnetic Nanoparticles in Dehydropeptide Supramolecular Magnetogels: Properties, Design and Drug Release.

Currently, the nanoparticle functionalization effect on supramolecular peptide-based hydrogels remains undescribed, but is expected to affect the hydrogels' self-assembly and final magnetic gel properties. Herein, two different functionalized nanoparticles: citrate-stabilized (14.4 ± 2.6 nm) and lipid-coated (8.9 ± 2.1 nm) magnetic nanoparticles, were used for the formation of dehydropeptide-based supramolecular magnetogels consisting of the ultra-short hydrogelator Cbz-L-Met-Z-ΔPhe-OH, with an assessment of their effect over gel properties. The lipid-coated nanoparticles were distributed along the hydrogel fibers, while citrate-stabilized nanoparticles were aggregated upon gelation, which resulted into a heating efficiency improvement and decrease, respectively. Further, the lipid-coated nanoparticles did not affect drug encapsulation and displayed improved drug release reproducibility compared to citrate-stabilized nanoparticles, despite the latter attaining a stronger AMF-trigger. This report points out that adsorption of nanoparticles to hydrogel fibers, which display domains that improve or do not affect drug encapsulation, can be explored as a means to optimize the development of supramolecular magnetogels to advance theranostic applications.

Exploring the properties and potential biomedical applications of NSAID-capped peptide hydrogels.

The development of strategies to minimise the adverse side-effects of non-steroidal anti-inflammatory drugs (NSAIDs) remains a challenge for medicinal chemists. One such strategy is the development of NSAID-peptide prodrug conjugates and this conjugation to a peptide often confers the additional property of hydrogelation. This review summarises the work published by our research group, alongside other research groups, on supramolecular hydrogels consisting of short peptides conjugated to NSAIDs. Generally, supramolecular low molecular weight hydrogels (LMWHs) are composed of amphiteric molecules, usually consisting of short peptides attached to an aromatic capping group. When the aromatic capping group is switched for an NSAID to afford hybrid gelators, some conjugates exhibit retained or improved anti-inflammatory properties of the parent drug, and sometimes new and unexpected biological activities are observed. Conjugation to peptides often provides selective COX-2 inhibition over COX-1 inhibtion, which is key to retaining the anti-inflammatory benefits of NSAIDs whilst minimising gastric side-effects. Naproxen is the most commonly employed NSAID capping group, partly due to its similarity in structure to commonly employed naphthalene capping groups. Biomimetic approaches, where canonical amino acids are switched for non-natural amino acids such as d-amino acids or dehydroamino acids, are often employed, to tune the stability. The future direction for this area of research is discussed.

Biological Evaluation of Naproxen-Dehydrodipeptide Conjugates with Self-Hydrogelation Capacity as Dual LOX/COX Inhibitors.

The use of peptide-drug conjugates is emerging as a powerful strategy for targeted drug delivery. Previously, we have found that peptides conjugated to a non-steroidal anti-inflammatory drug (NSAID), more specifically naproxen-dehydrodipeptide conjugates, readily form nanostructured fibrilar supramolecular hydrogels. These hydrogels were revealed as efficacious nano-carriers for drug delivery applications. Moreover, the incorporation of superparamagnetic iron oxide nanoparticles (SPIONs) rendered the hydrogels responsive to external magnetic fields, undergoing gel-to-solution phase transition upon remote magnetic excitation. Thus, magnetic dehydrodipeptide-based hydrogels may find interesting applications as responsive Magnetic Resonance Imaging (MRI) contrast agents and for magnetic hyperthermia-triggered drug-release applications. Supramolecular hydrogels where the hydrogelator molecule is endowed with intrinsic pharmacological properties can potentially fulfill a dual function in drug delivery systems as (passive) nanocariers for incorporated drugs and as active drugs themselves. In this present study, we investigated the pharmacological activities of a panel of naproxen-dehydrodipeptide conjugates, previously studied for their hydrogelation ability and as nanocarriers for drug-delivery applications. A focused library of dehydrodipeptides, containing N-terminal canonical amino acids (Phe, Tyr, Trp, Ala, Asp, Lys, Met) N-capped with naproxen and linked to a C-terminal dehydroaminoacid (ΔPhe, ΔAbu), were evaluated for their anti-inflammatory and anti-cancer activities, as well as for their cytotoxicity to non-cancer cells, using a variety of enzymatic and cellular assays. All compounds except one were able to significantly inhibit lipoxygenase (LOX) enzyme at a similar level to naproxen. One of the compounds 4 was able to inhibit the cyclooxygenase-2 (COX-2) to a greater extent than naproxen, without inhibiting cyclooxygenase-1 (COX-1), and therefore is a potential lead in the search for selective COX-2 inhibitors. This hydrogelator is a potential candidate for dual COX/LOX inhibition as an optimised strategy for treating inflammatory conditions.

Magnetic Dehydrodipeptide-Based Self-Assembled Hydrogels for Theragnostic Applications.

Self-assembled peptide hydrogels have emerged in recent years as the new paradigm in biomaterials research. We have contributed to this field the development of hydrogels based on dehydrodipeptides N-capped with naproxen. The dehydrodipeptide hydrogels can be loaded with drugs, thus being potential nanocarriers for drug delivery. In this work novel dehydrodipeptides containing tyrosine and aspartic acid amino acid residues N-capped with naproxen and C-terminal dehydrophenylalanine were prepared and characterized. Superparamagnetic iron oxide nanoparticles (SPIONs) were incorporated into the dehydrodipeptide-based hydrogels and their effect on the self-assembly, structure and rheological and magnetic properties of the hydrogels was studied. Magnetic hydrogels, with incorporated SPIONs, displayed concentration-dependent T2-MRI contrast enhancement. Moreover, upon magnetic excitation (alternating magnetic field -AMF-) the SPIONs were able to generate a significant amount of heat. Hence, magnetic hyperthermia can be used as a remote trigger for release of drug cargos and SPIONs incorporated into the self-assembled dehydrodipeptide hydrogels.

Fluorescence-Lifetime Imaging and Super-Resolution Microscopies Shed Light on the Directed- and Self-Assembly of Functional Porphyrins onto Carbon Nanotubes and Flat Surfaces.

Functional porphyrins have attracted intense attention due to their remarkably high extinction coefficients in the visible region and potential for optical and energy-related applications. Two new routes to functionalised SWNTs have been established using a bulky ZnII -porphyrin featuring thiolate groups at the periphery. We probed the optical properties of this zinc(II)-substituted, bulky aryl porphyrin and those of the corresponding new nano-composites with single walled carbon nanotube (SWNTs) and coronene, as a model for graphene. We report hereby on: i) the supramolecular interactions between the pristine SWNTs and ZnII -porphyrin by virtue of π-π stacking, and ii) a novel covalent binding strategy based on the Bingel reaction. The functional porphyrins used acted as dispersing agent for the SWNTs and the resulting nanohybrids showed improved dispersibility in common organic solvents. The synthesized hybrid materials were probed by various characterisation techniques, leading to the prediction that supramolecular polymerisation and host-guest functionalities control the fluorescence emission intensity and fluorescence lifetime properties. For the first time, XPS studies highlighted the differences in covalent versus non-covalent attachments of functional metalloporphyrins to SWNTs. Gas-phase DFT calculations indicated that the ZnII -porphyrin interacts non-covalently with SWNTs to form a donor-acceptor complex. The covalent attachment of the porphyrin chromophore to the surface of SWNTs affects the absorption and emission properties of the hybrid system to a greater extent than in the case of the supramolecular functionalisation of the SWNTs. This represents a synthetic challenge as well as an opportunity in the design of functional nanohybrids for future sensing and optoelectronic applications.

Self-assembled RGD dehydropeptide hydrogels for drug delivery applications.

Peptide-based self-assembled hydrogels have triggered remarkable research interest in recent years owing to their biocompatibility and biomimetic properties and responsiveness, which warrant many technological and biomedical applications. Dehydrodipeptides N-capped with naproxen emerged from our research as effective hydrogelators endowed with resistance to proteolysis. Dehydrodipeptide-based hydrogels are promising nanocarriers for drug delivery applications. In this work, we demonstrate that dehydrodipetide Npx-l-Ala-Z-ΔPhe-OH can be deployed as a minimalist hydrogelator module for synthesizing a gelating construct Npx-l-Ala-Z-ΔPhe-G-R-G-D-G-OH bearing a GRGDG adhesion motif. The self-assembly of the peptide construct and the drug delivery properties of the hydrogel were studied in this work. The peptide construct showed no toxicity towards a fibroblast cell line expressing the αvß3 integrin. Docking studies suggest that the hydrogelator block does not interfere with the recognition of the RGD motif by the integrin receptor. The self-assembly seems to be directed by intermolecular naphthalene π-π stacking interactions, with the peptide backbone assuming a random coil conformation both in solution and in the gel phase. TEM and STEM imaging revealed that the hydrogel is made of entangled bundles of long thin fibres (width circa 23 nm). The hydrogel exhibits viscoelastic properties, thermo-reversibility and recovery after mechanical fluidization. FRET studies showed that curcumin incorporated into the hydrogel interacts non-covalently with the hydrogel fibrils. Delivery of curcumin from the hydrogel into Nile red loaded model membranes (SUVs) was demonstrated by FRET. Naproxen N-capped dehydrodipeptides are efficacious minimalist hydrogelator modules for obtaining hydrogels functionalized with peptide ligands for cell receptors. These hydrogels are potential nanocarriers for drug delivery.

Booster dose after 10 years is recommended following 17DD-YF primary vaccination.

A single vaccination of Yellow Fever vaccines is believed to confer life-long protection. In this study, results of vaccinees who received a single dose of 17DD-YF immunization followed over 10 y challenge this premise. YF-neutralizing antibodies, subsets of memory T and B cells as well as cytokine-producing lymphocytes were evaluated in groups of adults before (NVday0) and after (PVday30-45, PVyear1-4, PVyear5-9, PVyear10-11, PVyear12-13) 17DD-YF primary vaccination. YF-neutralizing antibodies decrease significantly from PVyear1-4 to PVyear12-13 as compared to PVday30-45, and the seropositivity rates (PRNT≥2.9Log10mIU/mL) become critical (lower than 90%) beyond PVyear5-9. YF-specific memory phenotypes (effector T-cells and classical B-cells) significantly increase at PVday30-45 as compared to naïve baseline. Moreover, these phenotypes tend to decrease at PVyear10-11 as compared to PVday30-45. Decreasing levels of TNF-α(+) and IFN-γ(+) produced by CD4(+) and CD8(+) T-cells along with increasing levels of IL-10(+)CD4(+)T-cells were characteristic of anti-YF response over time. Systems biology profiling represented by hierarchic networks revealed that while the naïve baseline is characterized by independent micro-nets, primary vaccinees displayed an imbricate network with essential role of central and effector CD8(+) memory T-cell responses. Any putative limitations of this cross-sectional study will certainly be answered by the ongoing longitudinal population-based investigation. Overall, our data support the current Brazilian national immunization policy guidelines that recommend one booster dose 10 y after primary 17DD-YF vaccination.

Dehydrodipeptide Hydrogelators Containing Naproxen N-Capped Tryptophan: Self-Assembly, Hydrogel Characterization, and Evaluation as Potential Drug Nanocarriers.

In this work, we introduce dipeptides containing tryptophan N-capped with the nonsteroidal anti-inflammatory drug naproxen and C-terminal dehydroamino acids, dehydrophenylalanine (ΔPhe), dehydroaminobutyric acid (ΔAbu), and dehydroalanine (ΔAla) as efficacious protease resistant hydrogelators. Optimized conditions for gel formation are reported. Transmission electron microscopy experiments revealed that the hydrogels consist of networks of micro/nanosized fibers formed by peptide self-assembly. Fluorescence and circular dichroism spectroscopy indicate that the self-assembly process is driven by stacking interactions of the aromatic groups. The naphthalene groups of the naproxen moieties are highly organized in the fibers through chiral stacking. Rheological experiments demonstrated that the most hydrophobic peptide (containing C-terminal ΔPhe) formed more elastic gels at lower critical gelation concentrations. This gel revealed irreversible breakup, while the C-terminal ΔAbu and ΔAla gels, although less elastic, exhibited structural recovery and partial healing of the elastic properties. A potential antitumor thieno[3,2-b]pyridine derivative was incorporated (noncovalently) into the gel formed by the hydrogelator containing C-terminal ΔPhe residue. Fluorescence and Förster resonance energy transfer measurements indicate that the drug is located in a hydrophobic environment, near/associated with the peptide fibers, establishing this type of hydrogel as a good drug-nanocarrier candidate.

Gold nanoparticles functionalised with fast water exchanging Gd3+ chelates: linker effects on the relaxivity.

The relaxivity displayed by Gd(3+) chelates immobilized onto gold nanoparticles is the result of the complex interplay between the nanoparticle size, the water exchange rate and the chelate structure. In this work we study the effect of the length of ω-thioalkyl linkers, anchoring fast water exchanging Gd(3+) chelates onto gold nanoparticles, on the relaxivity of the immobilized chelates. Gold nanoparticles functionalized with Gd(3+) chelates of mercaptoundecanoyl and lipoyl amide conjugates of the DO3A-N-(α-amino)propionate chelator were prepared and studied as potential CA for MRI. High relaxivities per chelate, of the order of magnitude 28-38 mM(-1) s(-1) (30 MHz, 25 °C), were attained thanks to simultaneous optimization of the rotational correlation time and of the water exchange rate. Fast local rotational motions of the immobilized chelates around connecting linkers (internal flexibility) still limit the attainable relaxivity. The degree of internal flexibility of the immobilized chelates seems not to be correlated with the length of the connecting linkers. Biodistribution and MRI studies in mice suggest that the in vivo behavior of the gold nanoparticles was determined mainly by size. Small nanoparticles (HD = 3.9 nm) undergo fast renal clearance and avoidance of the RES organs while larger nanoparticles (HD = 4.8 nm) undergo predominantly hepatobiliary excretion. High relaxivities, allied to chelate and nanoparticle stability and fast renal clearance in vivo suggest that functionalized gold nanoparticles hold great potential for further investigation as MRI contrast agents. This study contributes to a better understanding of the effect of linker length on the relaxivity of gold nanoparticles functionalized with Gd(3+) complexes. It is a relevant contribution towards "design rules" for nanostructures functionalized with Gd(3+) chelates as Contrast Agents for MRI and multimodal imaging.

Ga[NO2A-N-(α-amino)propionate] chelates: synthesis and evaluation as potential tracers for 68Ga PET.

The availability of commercial (68)Ge/(68)Ga cyclotron-independent (68)Ga(3+) generators is making Positron Emission Tomography (PET) accessible to most hospitals, which is generating a surge of interest in the design and synthesis of bi-functional chelators for Ga(3+). In this work we introduce the NO2A-N-(α-amino)propionic acid family of chelators based on the triazacyclononane scaffold. Complexation of the parent NO2A-N-(α-amino)propionic acid chelator and of a low molecular weight (model) amide conjugate with Ga(3+) was studied by (1)H and (71)Ga NMR. The Ga(3+) chelate of the amide conjugate shows pH-independent N3O3 coordination in the pH range 3-10 involving the carboxylate group of the pendant propionate arm in a 6 member chelate. For the Ga[NO2A-N-(α-amino)propionate] chelate, a reversible pH-triggered switch from Ga(3+) coordination to the carboxylate group to coordination to the amine group of the propionate arm was observed upon pH increase/decrease in the pH range 4-6. This phenomenon can conceivably constitute the basis of a physiological pH sensor. Both complexes are stable in the physiological range. The [(67)Ga][NO2A-N-(α-benzoylamido)propionate] chelate was found to be stable in human serum. Biodistribution studies of the (67)Ga(3+)-labeled pyrene butyric acid conjugate NO2A-N-(α-pyrenebutanamido)propionic acid revealed that, despite its high lipophilicity and concentration-dependent aggregation properties, the chelate follows mainly renal elimination with very low liver/spleen accumulation and no activity deposition in bones after 24 hours. Facile synthesis of amide conjugates of the NO2A-N-(α-amino)propionic acid chelator, serum stability of the Ga(3+) chelates and fast renal elimination warrant further evaluation of this novel class of chelators for PET applications.