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1.
Am J Ther ; 16(6): 517-33, 2009.
Article in English | MEDLINE | ID: mdl-19487926

ABSTRACT

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. Second-generation carnosine analogs bearing the histidyl-hydrazide moiety were synthesized and tested versus L-carnosine for their ability to reverse the glycation process, also known as the Maillard reaction, and reverse the stable intermolecular cross-links, monitored in the glucose-ethylamine Schiff base model, ultimately resulting in the formation of the advanced glycation end products (AGEs) from nonenzymatic glycation, accumulating in numerous body tissues and fluids. The obtained data demonstrate the transglycation properties of the ophthalmically stabilized L-carnosine and L-carnosine histidyl-hydrazide derivatives tested and can be used to decrease or predict the occurrence of long-term complications of AGE formation and improve therapeutically the quality of vision and length of life for diabetes mellitus patients and survivors with early aging. Scientists at Innovative Vision Products, Inc. (IVP), developed lubricant eyedrops designed as a sustained-release 1% N-acetylcarnosine prodrug of L-carnosine. The eyedrops contain a mucoadhesive cellulose-based compound combined with corneal absorption promoters and glycerine in a drug-delivery system. Anti-aging therapeutics with the ophthalmic drug eyedrop formula including N-acetylcarnosine showed efficacy in the nonsurgical treatment of age-related cataracts for enrolled participants in the prospective, randomized, double-masked, placebo-controlled crossover clinical trial after controlling for age, gender, and daily activities. In a cohort in excess of 50,500 various patients seeking cutting-edge medical care, the N-acetylcarnosine topical eyedrops target therapy was demonstrated to have significant efficacy, safety, and good tolerability for the prevention and treatment of visual impairment in this older population with relatively stable patterns of causes for blindness and visual impairment. Overall, accumulated study data demonstrate that the IVP-designed new vision-saving drugs, including N-acetylcarnosine eyedrops, promote health vision and prevent vision disability from senile cataracts, primary open-angle glaucoma, age-related macular degeneration, diabetic retinopathy, and aging. N-acetylcarnosine eyedrop therapy is the crown jewel of the anti-aging medical movement and revolutionizes early detection, treatment, and rejuvenation of aging-related eye-disabling disorders. N-acetylcarnosine, as an innovative medical science tool and component of the home medicine and alternative medicine approaches, has the potential to alleviate visual impairment and its associated social, economic, and political woes for an aging population.


Subject(s)
Antioxidants/therapeutic use , Carnosine/analogs & derivatives , Cataract/drug therapy , Prodrugs/therapeutic use , Administration, Topical , Aged , Aged, 80 and over , Antioxidants/administration & dosage , Carnosine/administration & dosage , Carnosine/therapeutic use , Cross-Over Studies , Delayed-Action Preparations , Double-Blind Method , Female , Glycation End Products, Advanced/antagonists & inhibitors , Humans , Lipid Peroxidation/drug effects , Liposomes , Male , Middle Aged , Ophthalmic Solutions , Oxidative Stress/drug effects , Prodrugs/administration & dosage , Prospective Studies
4.
J Drug Target ; 17(1): 36-63, 2009 Jan.
Article in English | MEDLINE | ID: mdl-19037813

ABSTRACT

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. MAIN METHODS AND KEY FINDINGS: We proposed a deglycation system involving removal, by transglycation of sugar or aldehyde moieties from the Schiff bases by ophthalmic aldehyde scavenger L-carnosine derived from its ocular bioactivating sustained release prodrug 1% N-acetylcarnosine (NAC) lubricant eyedrops containing a mucoadhesive cellulose compound combined with corneal absorption promoters in drug delivery system. Carnosine analogs bearing the histidyl-hydrazide moiety were synthesized and patented in ophthalmic formulations with NAC bioactivating prodrug to moderate the enzymatic hydrolysis of a dipeptide by carnosinase (inhibited by a nonhydrolyzable substrate analog so that this keeps steadier levels of the drug active principle in the aqueous humor). Leucyl-histidylhydrazide peptidomimetic demonstrated the transglycation activity more pronounced than L-carnosine accounting for the ability of either molecule to reverse pre-existing, glycation-induced, cross-linking, and checking the nonenzymatic glycation cascade in the ophthalmic pathologies. The ophthalmic drug N-acetylcarnosine eye drop formulation with sustained time- release and increased absorption of L-carnosine in the aqueous humor (a prolonged effective dose) showed follow-up treatment efficacy for age-related cataracts for enrolled patients into the randomized double blind placebo controlled crossover clinical trial, and in over 50250 various cohort patients, was demonstrated to have an efficacy, safety and good tolerability for prevention and treatment of visual impairment in the older population data base. SIGNIFICANCE: The bioactivating antioxidant NAC and histidyl-hydrazide are potent agents with the pleiotropic effects for ophthalmic therapy of senile cataracts and diabetic ocular complications.


Subject(s)
Carnosine/analogs & derivatives , Cataract/complications , Cataract/drug therapy , Diabetes Complications/diagnosis , Histidine/analogs & derivatives , Histidine/administration & dosage , Hydrazines/administration & dosage , Ophthalmic Solutions/administration & dosage , Aged , Aged, 80 and over , Aldehydes/chemistry , Aminooxyacetic Acid/administration & dosage , Aminooxyacetic Acid/analysis , Aminooxyacetic Acid/chemistry , Aminooxyacetic Acid/metabolism , Animals , Biological Availability , Carnosine/administration & dosage , Carnosine/chemical synthesis , Carnosine/chemistry , Carnosine/metabolism , Cataract/diagnosis , Cataract/physiopathology , Cornea/drug effects , Cornea/metabolism , Cross-Over Studies , Diabetes Complications/physiopathology , Disease Models, Animal , Drug Administration Schedule , Drug Delivery Systems , Drug Synergism , Female , Glycosylation/drug effects , Histidine/chemistry , Histidine/metabolism , Humans , Hydrazines/chemistry , Hydrazines/metabolism , Lubricants/administration & dosage , Lubricants/analysis , Lubricants/chemistry , Lubricants/metabolism , Male , Middle Aged , Ophthalmic Solutions/analysis , Ophthalmic Solutions/chemistry , Ophthalmic Solutions/metabolism , Ophthalmologic Surgical Procedures , Ophthalmoscopy , Rabbits
5.
J Pept Sci ; 14(8): 989-97, 2008 Aug.
Article in English | MEDLINE | ID: mdl-18407579

ABSTRACT

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. This mechanism is supported by H/D exchange as observed by ESI-MS and NMR experiments.


Subject(s)
Amides/chemistry , Peptides/chemistry , Pipecolic Acids/chemistry , Trifluoroacetic Acid/chemistry , Binding Sites , Chemistry, Physical , Chromatography, High Pressure Liquid/methods , Deuterium Exchange Measurement , Hydrolysis , Magnetic Resonance Spectroscopy/methods , Magnetic Resonance Spectroscopy/standards , Molecular Conformation , Peptides/chemical synthesis , Peptides/isolation & purification , Reference Standards , Spectrometry, Mass, Electrospray Ionization/methods , Stereoisomerism
6.
Bioorg Med Chem ; 15(18): 6158-63, 2007 Sep 15.
Article in English | MEDLINE | ID: mdl-17604632

ABSTRACT

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.


Subject(s)
Carnosine/analogs & derivatives , Histidine/chemistry , Hydrazines/chemistry , Malondialdehyde/chemistry , Schiff Bases , Ascorbic Acid/metabolism , Carnosine/chemical synthesis , Carnosine/pharmacology , Glycosylation , Iron/metabolism , Lipid Peroxidation , Liposomes , Magnetic Resonance Spectroscopy , Malondialdehyde/metabolism , Oxidative Stress , Thiobarbituric Acid Reactive Substances/metabolism
7.
J Neurochem ; 101(3): 729-36, 2007 May.
Article in English | MEDLINE | ID: mdl-17254011

ABSTRACT

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. The histidine analogue also protected cultured rat primary neurons against death induced by HNE, chemical hypoxia, glucose deprivation, and combined oxygen and glucose deprivation. The histidine analogue prevented neuronal apoptosis as indicated by decreased production of cleaved caspase-3 protein. These findings suggest a therapeutic potential for HNE-scavenging histidine analogues in the treatment of stroke and related neurodegenerative conditions.


Subject(s)
Histidine/analogs & derivatives , Infarction, Middle Cerebral Artery/prevention & control , Neuroprotective Agents/therapeutic use , Animals , Animals, Newborn , Brain Infarction/drug therapy , Brain Infarction/etiology , Carnosine/analogs & derivatives , Carnosine/therapeutic use , Caspase 3/metabolism , Cell Death/drug effects , Cells, Cultured , Chromatography, High Pressure Liquid/methods , Disease Models, Animal , Glucose/deficiency , Histidine/metabolism , Hypoxia/drug therapy , Infarction, Middle Cerebral Artery/complications , Male , Mass Spectrometry/methods , Mice , Mice, Inbred C57BL , Neurons/drug effects , Neuroprotective Agents/metabolism , Rats , Rats, Sprague-Dawley , Time Factors
8.
J Med Chem ; 49(6): 1916-24, 2006 Mar 23.
Article in English | MEDLINE | ID: mdl-16539378

ABSTRACT

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. In particular, l-beta-MeTyr and d-Hat containing peptides resulted to be both suitable substrates for the specific monitoring of Syk and consensus sequence scaffolds for the design of potential inhibitors highly selective for this tyrosine kinase.


Subject(s)
Oligopeptides/chemistry , Protein-Tyrosine Kinases/chemistry , Tyrosine/chemistry , Kinetics , Magnetic Resonance Spectroscopy , Models, Molecular , Molecular Conformation , Molecular Mimicry , Oligopeptides/chemical synthesis , Phosphorylation , Protein-Tyrosine Kinases/antagonists & inhibitors , Stereoisomerism , Structure-Activity Relationship , Substrate Specificity
9.
J Pept Sci ; 12(7): 462-71, 2006 Jul.
Article in English | MEDLINE | ID: mdl-16506148

ABSTRACT

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. The association of a Pro-rich peptide with the SH3 domain involves unfavorable binding entropy due to the loss of rotational freedom on forming the PPII helix. With the aim of stabilizing the PPII helix conformation in the Pro-rich HPK1 decapeptide PPPLPPKPKF (P2), a series of P2 analogues was prepared, in which specific Pro positions were alternatively occupied by 4(S)- or 4(R)-4-fluoro-L-proline. The interactions of these peptides with the SH3 domain of the HPK1-binding partner HS1 were quantitatively analyzed by the NILIA-CD approach. A CD thermal analysis of the P2 analogues was performed to assess their propensity to adopt the PPII helix conformation. Contrary to our expectations, the K(d) values of the analogues were lower than that of the parent peptide P2. These results clearly show that the induction of a stable PPII helix conformation in short Pro-rich peptides is not sufficient to increase their affinity toward the SH3 domain and that the effect of 4-fluoroproline strongly depends on the position of this residue in the sequence and the chirality of the substituent in the pyrrolidine ring.


Subject(s)
Oligopeptides/chemistry , Peptides/chemistry , Proline/analogs & derivatives , Adaptor Proteins, Signal Transducing/chemistry , Adaptor Proteins, Signal Transducing/genetics , Adaptor Proteins, Signal Transducing/metabolism , Amino Acid Sequence , Animals , Circular Dichroism , DNA-Binding Proteins/chemistry , DNA-Binding Proteins/genetics , DNA-Binding Proteins/metabolism , Drug Design , In Vitro Techniques , Kinetics , Mice , Models, Molecular , Multiprotein Complexes , Proline/chemistry , Protein Binding , Protein Conformation , Protein Serine-Threonine Kinases/chemistry , Protein Serine-Threonine Kinases/genetics , Protein Serine-Threonine Kinases/metabolism , Protein Structure, Secondary , Recombinant Fusion Proteins/chemistry , Recombinant Fusion Proteins/genetics , Recombinant Fusion Proteins/metabolism , Solutions , Water , src Homology Domains
10.
J Pept Sci ; 12(7): 455-61, 2006 Jul.
Article in English | MEDLINE | ID: mdl-16485313

ABSTRACT

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 4.5-5.5. In this work, seven fluorescent, internally quenched, decapeptides have been synthesized using the prototypical cathepsin B selective substrate Z-Phe-Arg-AMC as a lead, and used to identify the structural factors determining the susceptibility of peptides to hydrolysis at acidic and neutral pH values. Each peptide differs from the others in one amino acid (residue 6) and contains a highly fluorescent Nma group linked to the alpha-amino function of the N-terminal Orn residue and a Dnp group linked to the side chain of the Lys(8) residue acting as a quencher. Proteolytic cleavage was monitored by measuring the increase of fluorescence at 440 nm upon excitation at 340 nm, and the cleavage sites were determined by HPLC followed by ESI-MS analysis. Peptides containing Ala or Phe at position 6 are good substrates for the enzyme at both pH 5.0 and 7.4. By contrast, those containing Glu, Asp, Lys or Val are not cleaved at all by cathepsin B at pH 7.4, and are poorly hydrolyzed at pH 5.0. These findings provide new information for the rational design of cathepsin B-activated peptide-containing anticancer drugs.


Subject(s)
Cathepsin B/chemistry , Cathepsin B/metabolism , Oligopeptides/chemistry , Oligopeptides/metabolism , Animals , Cattle , Fluorescent Dyes , Hydrogen-Ion Concentration , In Vitro Techniques , Kinetics , Oligopeptides/chemical synthesis , Protein Conformation , Spectrometry, Fluorescence , Substrate Specificity
11.
Curr Med Chem ; 12(20): 2293-315, 2005.
Article in English | MEDLINE | ID: mdl-16181134

ABSTRACT

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. The research for new and more potent scavengers for HNE and other alpha,beta-unsaturated aldehydes has produced a consistent variety of carnosine analogs, and the present review will resume, through the scientific literature and the international patents, the most recent developments in this field.


Subject(s)
Antioxidants , Carnosine , Aldehydes/antagonists & inhibitors , Animals , Antioxidants/chemistry , Antioxidants/metabolism , Carnosine/analogs & derivatives , Carnosine/chemistry , Carnosine/metabolism , Dipeptidases/metabolism , Glycation End Products, Advanced/antagonists & inhibitors , Humans , Structure-Activity Relationship
12.
J Med Chem ; 48(19): 6156-61, 2005 Sep 22.
Article in English | MEDLINE | ID: mdl-16162015

ABSTRACT

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.


Subject(s)
Aldehydes/toxicity , Carnosine/analogs & derivatives , Carnosine/chemical synthesis , Histidine/chemistry , Neuroprotective Agents/chemical synthesis , Aldehydes/metabolism , Animals , Carnosine/chemistry , Carnosine/pharmacology , Cell Death/drug effects , Cell Line, Tumor , Cells, Cultured , Cytoprotection , Hippocampus/cytology , Hippocampus/drug effects , Humans , Neurons/cytology , Neurons/drug effects , Neuroprotective Agents/chemistry , Neuroprotective Agents/pharmacology , Rats , Structure-Activity Relationship
13.
Bioorg Med Chem ; 12(19): 5031-7, 2004 Oct 01.
Article in English | MEDLINE | ID: mdl-15351387

ABSTRACT

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. Unexpected results reveal an anchimeric assistance of the Lys-AaA amide proton to the hydrolysis of the carbamoyl moiety joining mPEG to the alpha-amino group of lysine through the formation of an hydantoin system.


Subject(s)
Lysine/chemistry , Polyethylene Glycols/chemistry , Amino Acids/chemistry , Dipeptides/chemistry , Drug Stability , Hydrolysis
14.
J Pept Sci ; 10(7): 423-6, 2004 Jul.
Article in English | MEDLINE | ID: mdl-15298177

ABSTRACT

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.


Subject(s)
Gene Products, tat/chemistry , Gene Products, tat/metabolism , Micelles , Peptides/chemistry , Sodium Dodecyl Sulfate/chemistry , Water/chemistry , Circular Dichroism , Peptides/metabolism , Protein Folding , Protein Structure, Secondary , Solutions/chemistry
15.
Bioorg Med Chem Lett ; 14(7): 1803-5, 2004 Apr 05.
Article in English | MEDLINE | ID: mdl-15026076

ABSTRACT

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.


Subject(s)
Camptothecin/analogs & derivatives , Camptothecin/chemistry , Camptothecin/metabolism , Polyethylene Glycols/chemistry , Polyethylene Glycols/metabolism , Acylation , Stereoisomerism
16.
J Med Chem ; 47(5): 1280-9, 2004 Feb 26.
Article in English | MEDLINE | ID: mdl-14971908

ABSTRACT

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. Two conjugates demonstrated good stability at physiological pH and in mouse plasma (nonspecific proteases) but slowly released the drug payload in the presence of the lysosomal enzyme cathepsin B1. Compound 3, selected for in vivo experiments, was very active against P388, P388/ADM leukaemia, and Meth A fibrosarcoma cell lines, scoring T/C% values comparable with the camptothecin derivative CPT-11. Pharmacokinetic studies indicated that 3 acts as a reservoir of 10-amino-7-ethylcamptothecin, as the mean residence time (MRT) is about 3-fold higher than that of the free drug.


Subject(s)
Antineoplastic Agents/chemical synthesis , Camptothecin/chemical synthesis , Oligopeptides/chemical synthesis , Polyethylene Glycols/chemistry , Prodrugs/chemical synthesis , Animals , Antineoplastic Agents/pharmacokinetics , Antineoplastic Agents/pharmacology , Camptothecin/analogs & derivatives , Camptothecin/pharmacokinetics , Camptothecin/pharmacology , Cathepsin B/chemistry , Cell Line, Tumor , Drug Resistance, Neoplasm , Drug Screening Assays, Antitumor , Hydrolysis , Male , Mice , Oligopeptides/pharmacokinetics , Oligopeptides/pharmacology , Prodrugs/pharmacokinetics , Prodrugs/pharmacology , Rats , Rats, Sprague-Dawley , Structure-Activity Relationship
17.
Biopolymers ; 71(4): 478-88, 2003.
Article in English | MEDLINE | ID: mdl-14517899

ABSTRACT

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. Donella-Deana, M. Ruzzene, O. Marin, L. A. Pinna, FEBS Letters, 1995, Vol. 367, pp. 149-152), was fixed in the gauche (+) or gauche (-) conformation by using the 7-hydroxy-1,2,3,4-tetrahydro isoquinoline-3-carboxylic (Htc) structure. The tyrosine trans conformation was blocked by using an aminobenzazepine-type (Hba) structure. The proposed side-chain orientations were confirmed by the analysis of the (1)H-NMR parameters: chemical shifts, coupling constants, and nuclear Overhauser effects to the tyrosine constraints in the different analogs. This "rotamer scan" of the phosphorylatable residue allowed us to generate optimal substrates in terms of both phosphorylation efficiency and selectivity for Syk tyrosine kinase. In contrast, these conformationally restricted tyrosine analogs were not tolerated by the Src-related tyrosine kinases Lyn and c-Fgr.


Subject(s)
Protein-Tyrosine Kinases/chemistry , Amino Acid Sequence , Kinetics , Magnetic Resonance Spectroscopy , Models, Chemical , Molecular Sequence Data , Peptide Biosynthesis , Peptides/chemistry , Phosphorylation , Protein Binding , Protein Conformation , Protein-Tyrosine Kinases/metabolism , Substrate Specificity , Time Factors , Tyrosine/chemistry
18.
Eur J Med Chem ; 38(7-8): 739-49, 2003.
Article in English | MEDLINE | ID: mdl-12932905

ABSTRACT

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.g. norleucine) have been also employed. 1H-NMR of some conjugates was performed as characterisation of the product, while 195Pt-NMR analysis was carried out to detect the rearrangement of the platinum complex from the Pt(O,O) to the Pt(O,N) form. The compound PEG(5000)-Nle-malonato-Pt-DACH (4) has been tested against L1210-implanted mice and showed and appreciable increase in cytotoxicity as compared to the reference standard Cl(2)PtDACH.


Subject(s)
Antineoplastic Agents/chemical synthesis , Antineoplastic Agents/pharmacology , Cyclohexylamines/chemical synthesis , Cyclohexylamines/pharmacology , Organoplatinum Compounds/chemical synthesis , Organoplatinum Compounds/pharmacology , Animals , Drug Screening Assays, Antitumor , Drug Stability , Leukemia L1210/drug therapy , Leukemia L1210/metabolism , Magnetic Resonance Spectroscopy , Malonates/chemistry , Mice , Molecular Weight , Neoplasm Transplantation , Polyethylene Glycols/chemistry , Structure-Activity Relationship
19.
Farmaco ; 58(1): 45-50, 2003 Jan.
Article in English | MEDLINE | ID: mdl-12595036

ABSTRACT

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. We report the synthesis of a PEG-nisin conjugate, and the microbiology assays against some bacterial cell lines.


Subject(s)
Anti-Infective Agents/chemical synthesis , Nisin/chemical synthesis , Polyethylene Glycols/chemical synthesis , Anti-Bacterial Agents/chemical synthesis , Anti-Bacterial Agents/chemistry , Anti-Bacterial Agents/pharmacology , Anti-Infective Agents/chemistry , Anti-Infective Agents/pharmacology , Microbial Sensitivity Tests/statistics & numerical data , Nisin/chemistry , Nisin/pharmacology , Polyethylene Glycols/chemistry , Polyethylene Glycols/pharmacology
20.
Bioorg Med Chem Lett ; 12(2): 177-80, 2002 Jan 21.
Article in English | MEDLINE | ID: mdl-11755348

ABSTRACT

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.


Subject(s)
Alanine/chemistry , Dipeptides/chemistry , Methionine/chemistry , Polyethylene Glycols/chemical synthesis , Proteins/chemistry , Magnetic Resonance Spectroscopy , Polyethylene Glycols/chemistry , Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
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