José A. Martins

Supramolecular Assembly of an Amphiphilic GdIII Chelate: Tuning the Reorientational Correlation Time and the Water Exchange Rate

We report the synthesis and characterization of the novel ligand H(5)EPTPA-C(16) ((hydroxymethylhexadecanoyl ester)ethylenepropylenetriaminepentaacetic acid). This ligand was designed to chelate the Gd(III) ion in a kinetically and thermodynamically stable way while ensuring an increased water exchange rate (kappa(ex)) on the Gd(III) complex owing to steric compression around the water-binding site. The attachment of a palmitic ester unit to the pendant hydroxymethyl group on the ethylenediamine bridge yields an amphiphilic conjugate that forms micelles with a long tumbling time (tau(R)) in aqueous solution. The critical micelle concentration (cmc = 0.34 mM) of the amphiphilic [Gd(eptpa-C(16))(H(2)O)](2-) chelate was determined by variable-concentration proton relaxivity measurements. A global analysis of the data obtained in variable-temperature and multiple-field (17)O NMR and (1)H NMRD measurements allowed for the determination of parameters governing relaxivity for [Gd(eptpa-C(16))(H(2)O)](2-); this is the first time that paramagnetic micelles with optimized water exchange have been investigated. The water exchange rate was found to be kappa(298)(ex) = 1.7 x 10(8) s(-1), very similar to that previously reported for the nitrobenzyl derivative [Gd(eptpa-bz-NO(2))(H(2)O)](2-) kappa(298)(ex) = 1.5 x 10(8) s(-1)). The rotational dynamics of the micelles were analysed by using the Lipari-Szabo approach. The micelles formed in aqueous solution show considerable flexibility, with a local rotational correlation time of tau(298)(l0) = 330 ps for the Gd(III) segments, which is much shorter than the global rotational correlation time of the supramolecular aggregates, tau(298)(g0) = 2100 ps. This internal flexibility of the micelles is responsible for the limited increase of the proton relaxivity observed on micelle formation (r(1) = 22.59 mM(-1) s(-1) for the micelles versus 9.11 mM(-1) s(-1) for the monomer chelate (20 MHz; 25 degrees C)).

Gold nanoparticles functionalised with stable, fast water exchanging Gd3+ chelates as high relaxivity contrast agents for MRI

Gold nanoparticles functionalized with Gd(3+) chelates displaying fast water exchange, superb pH stability and inertness towards transmetalation with Zn(2+) have been prepared and characterized as a new high relaxivity (29 mM(-1) s(-1), 30 MHz, 25 °C) contrast agent potentially safe for in vivo MRI applications. The Lipari-Szabo treatment for internal rotation was used to evaluate the effect of linker flexibility on the relaxivity of the gold nanoparticles. The effect of fast water exchange on the relaxivity of gold nanoparticles functionalized with Gd(3+) chelates is also addressed in this communication.

Immobilisation on polystyrene of diazirine derivatives of mono- and disaccharides: biological activities of modified surfaces.

The potential of surface glycoengineering for biomaterials and biosensors originates from the importance of carbohydrate-protein interactions in biological systems. The strategy employed here utilises carbene generated by illumination of diazirine to achieve covalent bonding of carbohydrates. Here, we describe the synthesis of an aryl diazirine containing a disaccharide (lactose). Surface analysis techniques [X-ray photoelectron spectroscopy (XPS) and time of flight secondary ion mass spectroscopy (ToF-SIMS)] demonstrate its successful surface immobilisation on polystyrene (PS). Results are compared to those previously obtained with an aryl diazirine containing a monosaccharide (galactose). The biological activity of galactose- or lactose-modified PS samples is studied using rat hepatocytes, Allo A lectin and solid-phase semi-synthesis with alpha-2,6-sialyltransferase. Allo A shows some binding to galactose-modified PS but none to lactose-modified surfaces. Similar results are obtained with rat hepatocytes. In contrast, sialylation of lactose-modified PS is achieved but not with galactose-modified surfaces. The different responses indicate that the biological activity depends not only on the carbohydrate per se but also on the structure and length of the spacer.

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.

New self-assembled supramolecular hydrogels based on dehydropeptides

Supramolecular hydrogels rely on small molecules that self-assemble in water as a result of the cooperative effect of several relatively weak intermolecular interactions. Peptide-based low molecular weight hydrogelators have attracted enormous interest owing to the simplicity of small molecules combined with the versatility and biocompatibility of peptides. In this work, naproxen, a well known non-steroidal anti-inflammatory drug, was N-conjugated with various dehydrodipeptides to give aromatic peptide amphiphiles that resist proteolysis. Molecular dynamics simulations were used to obtain insight into the underlying molecular mechanism of self-assembly and to rationalize the design of this type of hydrogelators. The results obtained were in excellent agreement with the experimental observations. Only dehydrodipeptides having at least one aromatic amino acid gave hydrogels. The characterization of the hydrogels was carried out using transmission electron microscopy (TEM), circular dichroism (CD), fluorescence spectroscopy and also rheological assays.

Characterization of potential elastase inhibitor-peptides regulated by a molecular switch for wound dressings applications

Elastase plays an important role in wound healing process, degrading damaged tissue and allowing complete tissue recovery. The levels of human neutrophil elastase (HNE) are usually controlled by endogenous inhibitors. However, in the presence of high levels of elastase, like the ones present in chronic wounds, the inhibitors cannot overcome this overproduction and the enzyme starts to degrade the surrounding healthy tissue. In this work we report the development of a molecular switch to control the elastase activity in the exudate of non-healing chronic wounds. A peptide library was generated and screened in a microarray format for protein kinase-mediated phosphorylation. Two peptides were identified as casein kinase Iδ (CKI) substrates: KRCCPDTCGIKCL and its analogous peptide KRMMPDTMGIKML, with cysteine residues replaced by methionine residues. These peptides were studied in solution, both in the phosphorylated and non-phosphorylated forms as potential inhibitors for elastase. The obtained results show that the reversible process of phosphorylation/dephosphorylation results in differential inhibitory activity of the peptides. Thus the reversible process of phosphorylation/dephosphorylation can be used as a kind of molecular switch to control elastase activity. Degradation studies reveal that both the inhibitor-peptides and CKI are degraded by elastase. These results envisage the safe utilisation of these inhibitor-peptides together with CKI in the formulation of wound dressings.

A spatially stochastic epidemic model with partial immunization shows in mean field approximation the reinfection threshold

Recently, the notion of a reinfection threshold in epidemiological models of only partial immunity has been debated in the literature. We present a rigorous analysis of a model of reinfection which shows a clear threshold behaviour at the parameter point where the reinfection threshold was originally described. Furthermore, we demonstrate that this threshold is the mean field version of a transition in corresponding spatial models of immunization. The reinfection threshold corresponds to the transition between annular growth of an epidemics spreading into a susceptible area leaving recovered behind and compact growth of a susceptible-infected-susceptible region growing into a susceptible area. This transition between annular growth and compact growth was described in the physics literature long before the reinfection threshold debate broke out in the theoretical biology literature.

A scaling analysis in the SIRI epidemiological model.

For the spatial stochastic epidemic reinfection model SIRI, where susceptibles S can become infected I, then recover and remain only partial immune against reinfection R, we determine the phase transition lines using pair approximation for the moments derived from the master equation. We introduce a scaling argument that allows us to determine analytically an explicit formula for these phase transition lines and prove rigorously the heuristic results obtained previously.

Advances in the synthesis of homochiral (-)-1-azafagomine and (+)-5-epi-1-azafagomine. 1-N-phenyl carboxamide derivatives of both enantiomers of 1-azafagomine: Leads for the synthesis of active α-glycosidase inhibitors.

A new expeditious preparation of homochiral (-)-1-azafagomine and (+)-5-epi-1-azafagomine has been devised. Stoodleys diastereoselective cycloaddition of dienes bearing a 2,3,4,6-tetraacetyl glucosyl chiral auxiliary to 4-phenyl-1,2,4-triazole-3,5-dione was merged with Bolss protocol for functionalizing alkenes into molecules bearing a glucosyl framework. Homochiral (+)-5-epi-1-azafagomine was synthetized for the first time. Partial reductive cleavage of the phenyltriazolidinone moiety afforded new homochiral 1-N-phenyl carboxamide derivatives of 1-azafagomine. Both enantiomers of these derivatives were synthetized and tested, displaying a very good enzymatic inhibition toward bakers yeast α-glucosidase. The molecular recognition mechanism of the 1-N-phenyl carboxamide derivative of 1-azafagomine by α-glucosidase from bakers yeast was studied by molecular modeling. The efficient packing of the aromatic ring of the 1-N-phenyl carboxamide moiety into a hydrophobic subsite (pocket) in the enzymes active site seems to be responsible for the improved binding affinity in relation to underivatized (-)-1-azafagomine and (+)-1-azafagomine.

Amide conjugates of the DO3A-N-(α-amino)propionate ligand: leads for stable, high relaxivity contrast agents for MRI?

A novel synthetic methodology for preparing amide conjugates of the DO3A-N-(α-amino)propionate chelator is described, using the synthesis of the DO3A-N-(α-benzoylamido)propionate chelator as an illustrative example. The model Gd[DO3A-N-(α-benzoylamido)propionate] chelate displays accelerated water exchange, stability in a wide pH range and inertness towards transmetallation by Zn(2+). The Gd[DO3A-N-(α-benzoylamido)propionate] complex is mainly excreted via the kidneys, producing a significant increase in the kidney medulla/cortex enhancement ratio in MR images of Wistar rats, reflecting probably its higher lipophilicity compared with Gd(DTPA). The results presented suggest that Gd[DO3A-N-(α-amido)propionate] chelates can be valuable leads for preparing potentially safe high relaxivity MRI contrast agents.