Paolo Ruzza

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 Alzheimers 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.

Glutathione Transferases as Targets for Cancer Therapy

Besides catalyzing the inactivation of various electrophile-producing anticancer agents via conjugation to the tripeptide glutathione, some cytosolic proteins belonging to the glutathione transferase (formerly glutatione-S-transferase; GST) superfamily are emerging as negative modulators of stress/drug-induced cell apoptosis through the interaction with specific signaling kinases. In addition, several data link the overexpression of some GSTs, in particular GSTP1-1, to both natural and acquired resistance to various structurally unrelated anticancer drugs. Tumor overexpression of these proteins has provided a rationale for the search of GST inhibitors and GST-activated cytotoxic prodrugs. In the present review we discuss the current structural and pharmacological knowledge of both types of GST-targeting compounds.

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. 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.

Therapeutic prospect of Syk inhibitors

Background: The non-receptor spleen tyrosine kinase (Syk; EC 2.7.10.2) is involved in signal transduction in a variety of cell types. In particular, it is a key mediator of immune receptors signaling in host inflammatory cells (B cells, mast cells, macrophages and neutrophils), important for both allergic and antibody-mediated autoimmune diseases. Deregulated Syk kinase activity also allows growth factor-independent proliferation and transforms bone marrow-derived pre-B cells that are able to induce leukemia. Consequently, the development of Syk kinase inhibitors could conceivably treat these disorders and so they have became a major focus in the pharmaceutical and biotech industry. Objective: In this review, we analyze the structure and role of Syk kinase, the use of small molecules, interacting with ATP-binding site, as inhibitors of kinase activity and finally the potential of using inhibitors of Syk kinase expression to attenuate pathological conditions. Conclusion: Syk kinase inhibition is suggested as a powerful tool for the therapy of different pathologies.

Curcumin and Curcumin-like Molecules: From Spice to Drugs

Curcumin is the major yellow pigment extracted from turmeric, a commonly used spice in Asian cuisine and extensively employed in ayurvedic herbal remedies. A number of studies have shown that curcumin can be a prevention and a chemotherapeutic agent for colon, skin, oral and intestinal cancers. Curcumin is also well known for its antiinflammatory and antioxidant properties, showing high reactivity towards peroxyl radicals, and thus acting as a free radical scavenger. Recently, experimental studies have demonstrated that curcumin might be used in the prevention and the cure of Alzheimers disease. Indeed, curcumin injected peripherally in vivo into aged Tg mice crossed the blood-brain barrier and bound to amyloid plaques, reducing amyloid levels and plaque formation decisively. The present review will resume the most recent developments in the medicinal chemistry of curcumin and curcumin-like molecules.

Antamanide, a Derivative of Amanita phalloides, Is a Novel Inhibitor of the Mitochondrial Permeability Transition Pore

Antamanide is a cyclic decapeptide derived from the fungus Amanita phalloides. Here we show that antamanide inhibits the mitochondrial permeability transition pore, a central effector of cell death induction, by targeting the pore regulator cyclophilin D. Indeed, (i) permeability transition pore inhibition by antamanide is not additive with the cyclophilin D-binding drug cyclosporin A, (ii) the inhibitory action of antamanide on the pore requires phosphate, as previously shown for cyclosporin A; (iii) antamanide is ineffective in mitochondria or cells derived from cyclophilin D null animals, and (iv) abolishes CyP-D peptidyl-prolyl cis-trans isomerase activity. Permeability transition pore inhibition by antamanide needs two critical residues in the peptide ring, Phe6 and Phe9, and is additive with ubiquinone 0, which acts on the pore in a cyclophilin D-independent fashion. Antamanide also abrogates mitochondrial depolarization and the ensuing cell death caused by two well-characterized pore inducers, clotrimazole and a hexokinase II N-terminal peptide. Our findings have implications for the comprehension of cyclophilin D activity on the permeability transition pore and for the development of novel pore-targeting drugs exploitable as cell death inhibitors.

Ceftriaxone Blocks the Polymerization of α-Synuclein and Exerts Neuroprotective Effects in Vitro

The β-lactam antibiotic ceftriaxone was suggested as a therapeutic agent in several neurodegenerative disorders, either for its ability to counteract glutamate-mediated toxicity, as in cerebral ischemia, or for its ability to enhance the degradation of misfolded proteins, as in Alexanders disease. Recently, the efficacy of ceftriaxone in neuroprotection of dopaminergic neurons in a rat model of Parkinsons disease was documented. However, which characteristics of ceftriaxone mediate its therapeutic effects remains unclear. Since, at the molecular level, neuronal α-synuclein inclusions and pathological α-synuclein transmission play a leading role in initiation of Parkinson-like neurodegeneration, we thought of investigating, by circular dichroism spectroscopy, the capability of ceftriaxone to interact with α-synuclein. We found that ceftriaxone binds with good affinity to α-synuclein and blocks its in vitro polymerization. Considering this finding, we also documented that ceftriaxone exerts neuroprotective action in an in vitro model of Parkinsons disease. Our data, in addition to the findings on neuroprotective activity of ceftriaxone on Parkinson-like neurodegeneration in vivo, indicates ceftriaxone as a potential agent in treatment of Parkinsons disease.

A synthetic hexapeptide designed to resemble a proteinaceous p-loop nest is shown to bind inorganic phosphate

The hexapeptide Ser‐Gly‐Ala‐Gly‐Lys‐Thr has been synthesized and characterized. It was designed as a minimal soluble peptide that would be likely to have the phosphate‐binding properties observed in the P‐loops of proteins that bind the β‐phosphate of GTP or ATP. The β‐phosphate in such proteins is bound by a combination of the side chain ε‐amino group of the lysine residue plus the concavity formed by successive main chain peptide NH groups called a nest, which is favored by the glycines. The hexapeptide is shown to bind HPO42− strongly at neutral pH. The affinities of the various ionized species of phosphate and hexapeptide are analyzed, showing that they increase with pH. It is likely the main chain NH groups of the hexapeptide bind phosphate in much the same way as the corresponding P‐loop atoms bind the phosphate ligand in proteins. Most proteinaceous P‐loops are situated at the N‐termini of α‐helices, and this observation has frequently been considered a key aspect of these binding sites. Such a hexapeptide in isolation seems unlikely to form an α‐helix, an expectation in accord with the CD spectra examined; this suggests that being at the N‐terminus of an α‐helix is not essential for phosphate binding. An unexpected finding about the hexapeptide‐HPO42− complex is that the side chain ε‐amino group of the lysine occurs in its deprotonated form, which appears to bind HPO42− via an N···H‐O‐P hydrogen bond. Proteins 2012.

Small molecules interacting with α-synuclein: antiaggregating and cytoprotective properties.

Curcumin, a dietary polyphenol, has shown a potential to act on the symptoms of neurodegenerative disorders, including Alzheimer’s and Parkinson’s diseases, as a consequence of its antioxidant, anti-inflammatory and anti-protein aggregation properties. Unfortunately, curcumin undergoes rapid degradation at physiological pH into ferulic acid, vanillin and dehydrozingerone, making it an unlikely drug candidate. Here, we evaluated the ability of some curcumin by-products: dehydrozingerone (1), its O-methyl derivative (2), zingerone (3), and their biphenyl analogues (4–6) to interact with α-synuclein (AS), using CD and fluorescence spectroscopy. In addition, the antioxidant properties and the cytoprotective effects in rat pheochromocytoma (PC12) cells prior to intoxication with H2O2, MPP+ and MnCl2 were examined while the Congo red assay was used to evaluate the ability of these compounds to prevent aggregation of AS. We found that the biphenyl zingerone analogue (6) interacts with high affinity with AS and also displays the best antioxidant properties while the biphenyl analogues of dehydrozingerone (4) and of O-methyl-dehydrozingerone (5) are able to partially inhibit the aggregation process of AS, suggesting the potential role of a hydroxylated biphenyl scaffold in the design of AS aggregation inhibitors.

Glutathione Transferase (GST)-Activated Prodrugs

Glutathione transferase (formerly GST) catalyzes the inactivation of various electrophile-producing anticancer agents via conjugation to the tripeptide glutathione. Moreover, several data link the overexpression of some GSTs, in particular GSTP1-1, to both natural and acquired resistance to various structurally unrelated anticancer drugs. Tumor overexpression of these proteins has provided a rationale for the search of GST inhibitors and GST activated cytotoxic prodrugs. In the present review we discuss the current structural and pharmacological knowledge of GST-activated cytotoxic compounds.