Sara Pellegrino

1H‐Azepine‐4‐amino‐4‐carboxylic Acid: A New α,α‐Disubstituted Ornithine Analogue Capable of Inducing Helix Conformations in Short Ala‐Aib Pentapeptides

A very efficient synthesis of orthogonally protected 1H-azepine-4-amino-4-carboxylic acid, abbreviated as Azn, a conformationally restricted analogue of ornithine, was realized. It was obtained on a gram scale in good overall yield in five steps, three of which did not require isolation of the intermediates, starting from the readily available 1-amino-4-oxo-cyclohexane-4-carboxylic acid. Both enantiomers were used for the preparation of pentapeptide models containing Ala, Aib, and Azn. Conformational studies using both spectroscopic techniques (NMR, CD) and molecular dynamics on model 5-mer peptides showed that the (R)-Azn isomer possesses a marked helicogenic effect.

Chemistry of Biologically Active Isothiazoles

The isothiazole ring as well as the corresponding benzo- and heterocondensed rings are present in many chemically interesting compounds. The isothiazole ring can be a substituent of a bioactive scaffold or the pharmacophore of bioactive molecules. New compounds have been designed, synthesised and tested towards different biological targets and, in many cases, they display interesting pharmaceutical activities. Different SAR studies are reported starting from already known isothiazole derivatives or from new compounds characterised by a particular substitution pattern aiming to improve the biological activity. Agrochemical applications are also reported.

Mechanism of stabilization of helix secondary structure by constrained Cα-tetrasubstituted α-amino acids

The theoretical basis behind the ability of constrained Cα-tetrasubstituted amino acids (CTAAs) to induce stable helical conformations has been studied through Replica Exchange Molecular Dynamics Potential of Mean Force Quantum Theory of Atoms In Molecules calculations on Ac-l-Ala-CTAA-l-Ala-Aib-l-Ala-NHMe peptide models. We found that the origin of helix stabilization by CTAAs can be ascribed to at least two complementary mechanisms limiting the backbone conformational freedom: steric hindrance predominantly in the (+x,+y,-z) sector of a right-handed 3D Cartesian space, where the z axis coincides with the helical axis and the Cα of the CTAA lies on the +y axis (0,+y,0), and the establishment of additional and relatively strong C-H···O interactions involving the CTAA.

Chemotactic effect of prorenin on human aortic smooth muscle cells: a novel function of the (pro)renin receptor

AIMS The discovery of a specific prorenin receptor (PRR) suggests a biological function of prorenin that is independent of angiotensin I production. In the present study, we investigated the role of PRR on smooth muscle cell (SMC) migration. METHODS AND RESULTS PRR was expressed in human mammary arteries and in cultured human aortic SMCs. Prorenin induced SMC migration in a dose-dependent manner, as assessed by Boyden chamber chemotaxis assay, and increased SMC random motility, as determined by video microscopy. The prorenin decoy peptide inhibited SMC migration in response to prorenin, and knockdown of PRR by small interfering RNA completely inhibited the migratory response to prorenin, demonstrating that the chemotactic action of prorenin is mediated by the PRR. Prorenin induced cytoskeleton reorganization and lamellipodia formation and increased the intracellular levels of both RhoA-GTP and Rac1-GTP through PRR. These effects were required for SMC migration, because the suppression by small interfering RNA of either Rac1 or RhoA GTP-bound forms completely blocked the PRR-mediated chemotactic effect. Prorenin also induced the formation of larger focal adhesions and cleavage of the focal adhesion kinase (pp125(FAK)) into two main fragments with molecular weights of 50 and 90 kDa. The generation of these two fragments of pp125(FAK) was reduced by the calpain inhibitor ALLN, which also inhibited SMC migration in response to prorenin. CONCLUSIONS These results demonstrate that prorenin is a chemotactic factor for human aortic SMCs expressing PRR. This effect is elicited through reorganization of the cytoskeleton and focal adhesion, activation of RhoA and Rac1, and calpain-mediated cleavage of pp125(FAK).

A highly diastereoselective synthesis of α-hydroxy-β-amino acid derivatives via a Lewis acid catalyzed three-component condensation reaction.

A very efficient three-component synthesis of a series of syn α-hydroxy-β-amino esters, obtained in high diastereoselection and yield, was realized starting from an aldehyde, benzylamine, and the ketene silyl acetals derived from Leys lactones. The synthetic protocol was optimized and the above compounds were obtained without the isolation of intermediates. The origin of the observed diastereoselection was investigated through a computational model of the key reaction step.

Asymmetric modular synthesis of a semirigid dipeptide mimetic by cascade cycloaddition/ring rearrangement and borohydride reduction.

A new semirigid dipeptide mimetic was prepared on multigram scale, in good yield, and in a stereocontrolled way, starting from commercially available and unexpensive reagents, i.e., N-benzylpiperidone, tosyl azide, and proline methyl ester. The optimized multicomponent process consisted of a cascade click cycloaddition and a ring rearrangement reaction, followed by a reductive step. Theoretical calculations were performed to elucidate the reaction mechanism and support the stereochemical outcome of the reduction. Finally, the new scaffold was used for the preparation of model peptidomimetics, whose β turn conformation was confirmed by dynamic NMR experiments.

Expedient chemical synthesis of 75mer DNA binding domain of MafA: an insight on its binding to insulin enhancer

An expedient chemical synthesis of a 75mer peptide corresponding to the DNA binding domain (DBD, 227–301) of the human MafA leucine zipper transcription factor is reported. The application of microwave-assisted solid phase peptide synthesis (MW-SPPS) with a protocol modified respect to the standard one allowed obtaining the desired 75mer peptide in a short time with high quantity and optimal purity. MW-SPPS methodology was thus demonstrated as a valuable alternative to recombinant methods to obtain protein domains. Considering that recent findings suggest an involvement of MafA in the pathogenesis of diabetes mellitus, we also performed circular dichroism studies both on DBD folding and its interaction with MafA recognition element (MARE) on insulin enhancer. From our results, it was evicted that a disorder to order transition occurs after DBD interaction with insulin MARE which is mediated by specific structural elements on the N-terminus of the DBD.

Synthetic peptides containing a conserved sequence motif of the Id protein family modulate vascular smooth muscle cell phenotype

Modulation of smooth muscle cells to a proliferating and migrating phenotype with downregulated alpha-actin expression is observed upon vascular lesion formation. The Id proteins (inhibitors of cell differentiation) play a role in the development of this phenotype. In contrast, synthetic peptides based on a conserved 11-residue Id sequence trigger the switch to a contractile phenotype that shows reduced cell growth and migration, increased expression of alpha-actin and decreased Id protein levels.

Promising antiproliferative platinum(II) complexes based on imidazole moiety: Synthesis, evaluation in HCT-116 cancer cell line and interaction with Ctr-1 Met-rich domain

A series of imidazole based platinum(II) complexes were synthesised and evaluated for their cytotoxicity in HCT-116 cancer cell line, known for being partially resistant to cisplatin but sensitive to oxaliplatin. Lipophilicity was modulated by introducing differently long saturated and unsaturated chains at the N1 of the imidazole moiety. Pt-I displayed the higher cytotoxic effect achieving a IC50=38.0±14.1μM, comparable to the oxaliplatin value. The interaction between the imidazole platinum(II) complexes and the octapeptide called Mets7, the methionine-rich motif mimicking the N-terminal domain of the yCtr-1, was evaluated in order to have a major insight of the uptake and the eventual resistance mechanisms for the so-synthesised novel platinum compounds.

Dipeptide Nanotubes Containing Unnatural Fluorine-Substituted β2,3-Diarylamino Acid and l-Alanine as Candidates for Biomedical Applications

The synthesis and the structural characterization of dipeptides composed of unnatural fluorine-substituted β(2,3)-diarylamino acid and L-alanine are reported. Depending on the stereochemistry of the β amino acid, these dipeptides are able to self-assemble into proteolytic stable nanotubes. These architectures were able to enter the cell and locate in the cytoplasmic/perinuclear region and represent interesting candidates for biomedical applications.