N-acetylcarnosine lubricant eyedrops possess all-in-one universal antioxidant protective effects of L-carnosine in aqueous and lipid membrane environments, aldehyde scavenging, and transglycation activities inherent to cataracts: a clinical study of the new vision-saving drug N-acetylcarnosine eyedrop therapy in a database population of over 50,500 patients.

The antioxidant activity of L-carnosine (beta-alanyl-L-histidine, bioactivated in ocular tissues) versus N-acetylcarnosine (N-acetyl-beta-alanyl-L-histidine, ocular-targeted small dipeptide molecules) was studied in aqueous solution and in a lipid environment, employing liposomes as a model of lipid membranes. Reactive oxygen species (ROS) were generated by an iron/ascorbate promoter system for induction of lipid peroxidation (LPO). L-carnosine, which is stabilized from enzymatic hydrolysis, operates as a universal aldehyde and ROS scavenger in both aqueous and lipid environments and is effective at preventing ROS-induced damage to biomolecules.

N-Acetylcarnosine and histidyl-hydrazide are potent agents for multitargeted ophthalmic therapy of senile cataracts and diabetic ocular complications.

Aims: In human diabetes, the deleterious effects of chronic hyperglycemia are the result of excessive nonenzymatic modification of proteins and phospholipids by glucose and its by-products leading to the formation of irreversible oxidized, aromatic, and fluorescent ligands known as advanced glycation end products. This glycation process has been associated with deleterious health effects. The present invention provides the potent inhibitors of protein glycation and AGEs formation, which are particularly advantageous for eyedrop delivery in the prevention and treatment of diabetes- and age-related pathologies.

Mechanistic studies of amide bond scission during acidolytic deprotection of Pip containing peptide.

Unusual TFA catalyzed cleavage reaction is reported for peptide containing pipecolic acid residues. Although the use of TFA under standard cleavage conditions is sufficiently mild to prevent degradation of the desired products, the amide bond between consecutive pipecolic acid residues is unexpectedly hydrolyzed by standard TFA treatment. The hydrolysis is proposed to proceed via an oxazolinium ion intermediate.

Malondialdehyde scavenging and aldose-derived Schiff bases' transglycation properties of synthetic histidyl-hydrazide carnosine analogs.

Second-generation carnosine analogs bearing the histidyl-hydrazide moiety have been synthesized and tested for their efficiency in scavenging malondialdehyde (MDA) derived from lipid peroxidation and for their ability to reverse the glycation process in the glucose-ethylamine Schiff base model. The data obtained indicate that this class of compounds maintains the activity profile of carnosine and is a suitable candidate for the treatment of disorders caused by oxidative stress.

Neuroprotective actions of a histidine analogue in models of ischemic stroke.

Histidine is a naturally occurring amino acid with antioxidant properties, which is present in low amounts in tissues throughout the body. We recently synthesized and characterized histidine analogues related to the natural dipeptide carnosine, which selectively scavenge the toxic lipid peroxidation product 4-hydroxynonenal (HNE). We now report that the histidine analogue histidyl hydrazide is effective in reducing brain damage and improving functional outcome in a mouse model of focal ischemic stroke when administered intravenously at a dose of 20 mg/kg, either 30 min before or 60 min and 3 h after the onset of middle cerebral artery occlusion.

Spatial conformation and topography of the tyrosine aromatic ring in substrate recognition by protein tyrosine kinases.

The side chain orientation of the tyrosine residue included in a peptide, which is an excellent substrate of Syk tyrosine kinase, was fixed in different conformations by either incorporating the tyrosine in cyclic structures (6-OH-Tic, 5-OH-Aic, and Hat derivatives) or adding a sterically bulky substituent in the tyrosine side chain moiety (beta-MeTyr). Synthetic peptides containing tyrosine analogues displaying different side chain orientations were analyzed by NMR techniques and tested as potential substrates of the nonreceptor tyrosine kinases Syk, Csk, Lyn, and Fyn. The "rotamer scan" of the phosphorylatable residue generated optimal substrates in terms of both phosphorylation efficiency and selectivity for Syk tyrosine kinase, while the peptidomimetics were not recognized by the other tyrosine kinases.

4-Fluoroproline derivative peptides: effect on PPII conformation and SH3 affinity.

Eukaryotic signal transduction involves the assembly of transient protein-protein complexes mediated by modular interaction domains. Specific Pro-rich sequences with the consensus core motif PxxP adopt the PPII helix conformation upon binding to SH3 domains. For short Pro-rich peptides, little or no ordered secondary structure is usually observed before binding interactions.

Fluorescent, internally quenched, peptides for exploring the pH-dependent substrate specificity of cathepsin B.

Cathepsin B is a cysteine protease that in tumor tissues is localized in both acidic lysosomes and extracellular spaces. It can catalyze the cleavage of peptide bonds by two mechanisms: endoproteolytic attack with a pH optimum around 7.4, and attack from the C-terminus with a pH optimum at 4.

Carnosine and carnosine-related antioxidants: a review.

First isolated and characterized in 1900 by Gulewitsch, carnosine (beta-alanyl-L-hystidine) is a dipeptide commonly present in mammalian tissue, and in particular in skeletal muscle cells; it is responsible for a variety of activities related to the detoxification of the body from free radical species and the by-products of membrane lipids peroxidation, but recent studies have shown that this small molecule also has membrane-protecting activity, proton buffering capacity, formation of complexes with transition metals, and regulation of macrophage function. It has been proposed that carnosine could act as a natural scavenger of dangerous reactive aldehydes from the degradative oxidative pathway of endogenous molecules such as sugars, polyunsaturated fatty acids (PUFAs) and proteins. In particular, it has been recently demonstrated that carnosine is a potent and selective scavenger of alpha,beta-unsaturated aldehydes, typical by-products of membrane lipids peroxidation and considered second messengers of the oxidative stress, and inhibits aldehyde-induced protein-protein and DNA-protein cross-linking in neurodegenerative disorders such as Alzheimer's disease, in cardiovascular ischemic damage, in inflammatory diseases.

Synthesis and evaluation of neuroprotective alpha,beta-unsaturated aldehyde scavenger histidyl-containing analogues of carnosine.

The synthesis, scavenging activity, and cytoprotective profiles of histidyl-containing carnosine analogues bearing hydrazide or 1,2-diol moieties is reported. Some compounds have demonstrated higher aldehyde-sequestering efficiency than carnosine and were also efficient in protecting SH-SY5Y neuroblastoma cells and rat hippocampal neurons from 4-hydroxy-trans-2,3-nonenal (HNE)-mediated death. The cytoprotective efficacy of these compounds suggests their potential use as therapeutic agents for disorders that involve excessive membrane lipids peroxidation and HNE-mediated neuronal toxicity.

Anchimeric assistance effect on regioselective hydrolysis of branched PEGs: a mechanistic investigation.

Branched poly(ethylene glycols) (PEG2) are nowadays widely used for protein and peptides bioconjugation, for their favourable properties (such as the ability to protect the protein surface in an 'umbrella like' fashion). The discovery that mPEG(2)-LysMetbeta AlaOEt lost one mPEG chain during standard base-catalysed ester hydrolysis conditions prompted us to investigate the hydrolytic stability of such systems and the mechanism involved in the PEG chain loss. A series of branched PEGs, substituted with different aminoacids and dipeptides, have been prepared to test the influence of steric hindrance, chain lengths, ramification and Lys-AA amide substitution on hydrolysis.

Tat cell-penetrating peptide has the characteristics of a poly(proline) II helix in aqueous solution and in SDS micelles.

Tat cell-penetrating peptide (GRKKRRQRRRPPQG) is able to translocate and carry molecules across cell membranes. Using CD spectroscopy the conformation of this synthetic peptide was studied in aqueous and membrane-mimicking, micellar SDS solutions at different temperatures. The CD spectrum of the Tat cell-penetrating peptide in SDS micellar solution was virtually unchanged from that in aqueous solution, and at low temperature it was close to that of a poly(proline) II helix.

Efficient and chemoselective N-acylation of 10-amino-7-ethyl camptothecin with poly(ethylene glycol).

A new poly(ethylene glycol) (PEG) conjugate of 10-amino-7-ethyl camptothecin, a potent antitumor analogue of camptothecin, has been synthesized and preliminary in vivo tests have been performed. Successful chemoselective N-acylation of 10-amino-7-ethyl camptothecin was accomplished using phenyl dichlorophosphate, a coupling reagent used in esterification of alcohols, while other coupling methods failed, due to the low nucleophilicity of the amino group in position 10. The conjugate was tested against P388 murine leukemia cell lines and resulted equipotent to CPT-11, a camptothecin analogue already in clinical use.

Synthesis, characterization, and preliminary in vivo tests of new poly(ethylene glycol) conjugates of the antitumor agent 10-amino-7-ethylcamptothecin.

Despite the high antitumor activity of camptothecins, few derivatives have been developed and tested for human treatment of solid tumors, due to unpredictable toxicity mainly connected to their poor water solubility. We report the conjugation of the antitumor agent 10-amino-7-hydroxy camptothecin (SN-392) to linear or branched poly(ethylene glycol)s (PEGs) of different loading capacity through a tri- or tetrapeptide spacer selectively cleaved by lysosomal enzymes (cathepsins). A synthetic strategy based on the chemoselective acylation of the aromatic amino group in the presence of the unprotected C20 tertiary alcohol allowed high overall yields.

Conformational constraints of tyrosine in protein tyrosine kinase substrates: Information about preferred bioactive side-chain orientation.

The side-chain orientation of a tyrosine residue located in a peptide, which is an excellent substrate of Syk tyrosine kinase (A. M. Brunati, A.

Synthesis, characterization and preliminary cytotoxicity assays of poly(ethylene glycol)-malonato-Pt-DACH conjugates.

Oxalate 1,2-diaminocyclohexane platinum (oxaliplatin(R)), a successfully employed platinum compound belonging to the family of Pt-DACH complexes, has been conjugated to different molecular weight poly(ethylene glycols) (PEG) by means of peptide spacers and a malonic acid bidentate residue. Tri- and tetrapeptidic substrates of lysosomal enzymes were used in order to increase the release of Pt-DACH complex inside the cell following endocytosis and enzymatic degradation of the peptide spacer. Other aminoacids (e.

PEGylation of the antimicrobial peptide nisin A: problems and perspectives.

Nisin is a natural antimicrobial peptide produced by Lactococcus lactis and widely employed as food preservative. Its low solubility in neutral aqueous solutions, its instability at physiological pH and its rapid breakdown by proteolytic enzymes has limited its use for processed foods (processed cheese, milk and derivatives, canned vegetables). The conjugation to poly(ethylene glycol) (PEG) could improve its solubility and protect it towards enzymes present in non optimally processed food.

An improved procedure for the synthesis of branched polyethylene glycols (PEGs) with the reporter dipeptide Met-betaAla for protein conjugation.

A new and more efficient route to the synthesis of branched PEG for protein conjugation, bearing a reporter dipeptide Met-betaAla, is described, which allows better purification of the final product by ion exchange chromatography. The product has the combined advantages of an 'umbrella-like' branched structure, which allows a better coverage of the protein surface, and the presence of the dipeptide Met-betaAla which has been used to detect the position of PEGylation within the peptide sequence.