Discovery of inhibitors of carbonyl related degenerative disease : peptides and derivatives as detoxifying agents of cytotoxic aldehydes
Tesi di Dottorato
Data di Pubblicazione:
2007
Citazione:
Discovery of inhibitors of carbonyl related degenerative disease : peptides and derivatives as detoxifying agents of cytotoxic aldehydes / L. Gamberoni ; Tutor: G. Aldini. DIPARTIMENTO DI SCIENZE FARMACEUTICHE "PIETRO PRATESI", 2007 Feb 01. 20. ciclo, Anno Accademico 2004/2005.
Abstract:
Introduction
Reactive carbonyl species (RCS) are important cytotoxic mediators generated by lipid oxidation of PUFAs, leading to alteration of the cellular function and inducing irreversible structural modifications to biomolecules. RCS belong to different chemical classes such as alfa,beta-unsaturated aldehydes [4-hydroxy-trans-2-nonenal (HNE), acrolein (ACR)], dialdehydes [malondialdehyde (MDA), glyoxal (GO)], levuglandines and prostaglandin member of J2 series. RCS are electrophilic and reactive compounds capable to form covalent adducts with nucleophilic molecules in particular proteins and nucleic acids. RCS and the corresponding adducts with proteins (carbonylated proteins) are widely used as biomarkers of lipid peroxidation and, in general, of oxidative stress. Moreover, there are several convincing evidences supporting a pathogenic role for RCS, such as in the case of diabetic-related diseases, age-dependent tissue dysfunction, and metabolic distress syndrome. Consequently, RCS, in addition of being a predictive biomarker, also represents a biological target for drug discovery (1).
Aim of the work
The present work is devoted to better understand the molecular and cellular effects of RCS and to design novel compounds able to inhibit the RCS-derived cellular dysfunction, the RCS sequestering agents (RCS-SA). To reach these goals, the following steps have been conducted: (i) to study the effects of RCS on protein and cellular function; (ii) to design specific and efficient sequestering agents of RCS; (iii) to set-up an analytical method aimed to screen reactivity, selectivity and to elucidate the reaction mechanisms of RCS-SA; (iv) to identify a biomarker of protein carbonylation to monitor the efficacy of RCS-SA in in vivo studies.
RCS effect on protein and cellular function
The cause or effect role of RCS has been evaluated in cellular models by using 15d-PGJ2, a neurotoxic prostaglandin member of J2 series, as model of RCS. By using biotinylated 15d-PGJ2, it was found that actin is the main protein cellular target of 15d-PGJ2, which specifically binds through a Michael adduction to Cys374, leading to a protein conformational change and consequently disruption of the actin cytoskeleton, F-actin depolymerization, and impairment of G-actin polymerization (2). We further studied the effect of acrolein and HNE on actin function and the sites and mechanism of adduction fully characterized by MS analyses (3).
Discovery of specific and efficient sequestering agents of RCS
Taking RCS and carbonylation damage as drug-targets, different molecular strategies have been up to now considered in order to neutralize/reduce these pathogenetic factors and the most promising is that based on nucleophilic compounds capable to form covalent and unreactive adducts with RCS. Among the most studied RCS sequestering agent, the vitamer pyridoxamine (PYR), hydralazine (HY), and aminoguanidine (AG). Most of the RCS-SA above listed are characterized by a high nucleophilic amino group accompanied by a low basicity favoring the unprotonated form. This feature, if from one side permits a high reactivity towards a broad range of RCS, from another side greatly limits the specificity of the RCS sequestering agents, since it makes the compounds reactive toward biogenic and physiological aldehydes, such as pyridoxal phosphate (4). We recently found that carnosine (beta-alanyl-L-histidine, CAR), an endogenous dipeptide present in mM concentrations in some tissues such as skeletal muscle, is a specific quencher of alfa,beta-unsaturated aldehydes (5). The reaction mechanism involves both the Schiff base formation between the beta-alanine amino group and the aldehydic function, which then catalyses the Michael adduction between the C3 of the aldehyd
Tipologia IRIS:
13 - Tesi di dottorato discussa entro ottobre 2010
Elenco autori:
L. Gamberoni
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