Login| Sign Up| Help| Contact|

Patent Searching and Data


Title:
HYDROXY-2,3-DIARYLXANTHONES AS THERAPEUTIC AGENTS OF PATHOLOGIES DUE TO REACTIVE OXYGEN SPECIES
Document Type and Number:
WIPO Patent Application WO/2011/117690
Kind Code:
A1
Abstract:
The present invention relates to the use of hydroxyl-2, 3- diarylxanthones (DAX) as therapeutic agent of pathologies due to reactive oxygen species. Certain polyhydroxy-2, 3-diaryl-9H-xanthen-9-ones present antioxidant properties much higher than that of the flavonol quercetin (Q), which is considered as one of the most efficient antioxidant of the polyphenolic family of flavonoids. At the same concentration DAX has a protector power three times higher than that of Q relatively to the copper (II) induced oxidation of human LDL. Concerning the tissue oxidant stress, polyhydroxy-2, 3- diaryl-9H-xanthen-9-ones have a protective power four times higher than that of Q relatively to the cytotoxicity induced by tert-butyl hydroperoxide (t-BuOOH) in cultured human keranocytes. The referred efficacy of DAX against the oxidant stress can be explained by their reactivity with reactive oxygen species (ROS), which is illustrated by their reactions with superoxide radical anions, peroxotrimethyl and Trp radicals selectively produced by pulse radiolysis.

Inventors:
SANTUS RENE (FR)
SOARES DA SILVA ARTUR MANUEL (PT)
MOREIRA DOS SANTOS CLEMENTINA MARIA (PT)
DA SILVA CAVALEIRO JOSE ABRUNHEIRO (PT)
LEAL FILIPE PAULO MANUEL (PT)
MAZIERE JEAN-CLAUDE (FR)
MORLIERE PATRICE (PT)
Application Number:
PCT/IB2010/055685
Publication Date:
September 29, 2011
Filing Date:
December 09, 2010
Export Citation:
Click for automatic bibliography generation   Help
Assignee:
UNIV AVEIRO (PT)
SANTUS RENE (FR)
SOARES DA SILVA ARTUR MANUEL (PT)
MOREIRA DOS SANTOS CLEMENTINA MARIA (PT)
DA SILVA CAVALEIRO JOSE ABRUNHEIRO (PT)
LEAL FILIPE PAULO MANUEL (PT)
MAZIERE JEAN-CLAUDE (FR)
MORLIERE PATRICE (PT)
International Classes:
A61K31/352; A61P39/06
Foreign References:
US6730333B12004-05-04
Other References:
SANTOS C M M ET AL: "A novel and efficient route for the synthesis of hydroxylated 2,3-diarylxanthones", SYNLETT 20071217 DE LNKD- DOI:10.1055/S-2007-990900, no. 20, 17 December 2007 (2007-12-17), pages 3113 - 3116, XP002634033, ISSN: 0936-5214
SANTOS C M M ET AL: "2,3-Diarylxanthones as strong scavengers of reactive oxygen and nitrogen species: A structure-activity relationship study", BIOORGANIC & MEDICINAL CHEMISTRY, PERGAMON, GB, vol. 18, no. 18, 15 September 2010 (2010-09-15), pages 6776 - 6784, XP027252781, ISSN: 0968-0896, [retrieved on 20100725]
A. J. N. SELLES; H. T. V. CASTRO; J. AGUERO-AGUERO; J. GONZÁLEZ-GONZÁLEZ; F. NADDEO; F. DE SIMONE; L. RASTRELLI: "Isolation and quantitative analysis of phenolic antioxidants, free sugars, and polyols from Mango (Mangifera indica L.) stem bark aqueous decoction used in Cuba as a nutritional supplement", J. AGRIC. FOOD CHEM., vol. 50, 2002, pages 762 - 766, XP009049806, DOI: doi:10.1021/jf011064b
C. M. M. SANTOS; A. M. S. SILVA; J. A. S. CAVALEIRO: "A novel and efficient route for the synthesis of hydroxylated 2,3- diarylxanthones", EUR. J. ORG. CHEM., 2009, pages 2642 - 2660
D. K. DAS; N. MAULIK: "Resveratrol in cardioprotection: a therapeutic promise of alternative medicine", MOL. INTERV., vol. 6, 2006, pages 36 - 47
T. WATANABE; M. TAHARA; S. TODO.: "The novel antioxidant edavarone: From bench to bedside", CARDIOVASCULAR THERAPEUTICS, vol. 26, 2008, pages 101 - 114, XP055090100, DOI: doi:10.1111/j.1527-3466.2008.00041.x
H. YOSHIDA; K. SASAKI; Y. NAMIKI; N. SATO; N. TADA: "Edavarone, a novel radical scavenger, inhibits oxidative modification of low-density lipoproteins (LDL) and reverses oxidized LDL-mediated reduction in the expression of endothelial nitric oxide synthase", ATHEROSCLEROSIS, vol. 179, 2005, pages 97 - 102
R. A. FLOYD; R. D. KOPKE; C. H. CHOI; S. B. FOSTE; S. DOBLAS; R. A. TOWNER: "Nitrones as therapeutics", FREE RADIC. BIOL. MED., vol. 45, 2008, pages 1361 - 1374
K. R. LEES; J. A. ZIVIN; T. ASHWOOD; A. DAVALOS; S. M. DAVIS; H. C. DIENER; J. GROTTA; P. LYDEN; A. SHUAIB; H. G. HARDEMARK: "Stroke-acute ischemic NXY treatment (SAINT I) trial investigators", N. ENGL. J. MED., vol. 354, 2006, pages 588 - 600
H. YOSHIDA; K. SASAKI; Y. NAMIKI; N. SATO; N. TADA: "Edavarone, a novel radical scavenger, inhibits oxidative modification of low-density lipoprotein (LDL) and reverses oxidized LDL-mediated reduction in the expression of endothelial nitric oxide ' synthase", ATHEROSCLEROSIS, vol. 179, 2005, pages 97 - 102
L. BELGUENDOUZ; L. FREMONT; A. LINARD.: "Resveratrol inhibits metal ion-dependent and independent peroxidation of porcine low-density lipoproteins", BIOCHEM. PHARMACOL., vol. 53, 1997, pages 1347 - 1355
J. N. SELLES; H. T. V. CASTRO; J. AGÜERO-AGÜERO; J. GONZALEZ-GONZALEZ; F. NADDEO; F. DE SIMONE; L. RASTRELLI.: "Isolation and quantitative analysis of phenolic antioxidants, free sugars, and polyols from Mango (Mangifera indica L.) stem bark aqueous decoction used in Cuba as a nutritional supplement", J. AGRIC. FOOD CHEM., vol. 50, 2002, pages 762 - 766, XP009049806, DOI: doi:10.1021/jf011064b
C. M. M. SANTOS; A. M. S. SILVA; J. A. S. CAVALEIRO: "A novel and efficient route for the synthesis of hydroxylated 2,3-diarylxanthones", EUR. J. ORG. CHEM., 2009, pages 2642 - 2660
P. FILIPE; A. M. S. SILVA; R. S. G. R. SEIXAS; D. C. G. A. PINTO; A. SANTOS; L. K. PATTERSON; J. N. SILVA; J. A. S. CAVALEIRO; J.: "The alkyl chain length of 3-alkyl-3',4',5,7-tetrahydroxyflavones modulates effective inhibition of oxidative damage in biological systems: Illustration with LDL, red blood cells and human skin keratinocytes", BIOCHEM. PHARMACOL., vol. 77, 2009, pages 957 - 964, XP025990020, DOI: doi:10.1016/j.bcp.2008.11.023
H. ESTERBAUER; G. STRIEGL; H. PUHL; M. ROTHENEDER: "Continuous monitoring of in vivo oxidation of human low density lipoprotein", FREE RAD. RES. COMMUN., vol. 6, 1989, pages 67 - 75
D. R. BICKERS; M. ATHAR.: "Oxidative stress in the pathogenesis of skin disease", J INVEST. DERMATOL., vol. 126, 2006, pages 2565 - 2575, XP002524710, DOI: doi:10.1038/sj.jid.5700340
A. M. S. SILVA; P. FILIPE; R. S. G. R. SEIXAS; D. C. G. A. PINTO; L. K. PATTERSON; G. L. HUG, J. A. S. CAVALEIRO; J.-C. MAZIÈRE;: "One-electron reduction of superoxide radical-anions by 3-alkylpolyhydroxyflavones in micelles. Effect of antioxidant alkyl chain length on micellar structure and reactivity", J. PHYS. CHEM. B, vol. 112, 2008, pages 11456 - 11461
A. M. S. SILVA; P. FILIPE; R. S. G. R. SEIXAS; D. C. G. A. PINTO; L. K. PATTERSON; G. L. HUG; J. A. S. CAVALEIRO; J.-C. MAZIÈRE;: "One-electron reduction of superoxide radical-anions by 3-alkylpolyhydroxyflavones in micelles. Effect of antioxidant alkyl chain length on micellar structure and reactivity", J. PHYS. CHEM. B, vol. 112, 2008, pages 11456 - 11461
A. B. GIESSAUF; B. VAN WICKERN; T. SIMAT; H. STEINHART; H. ESTERBAUER: "Formation of N-formylkynurenine suggests the involvement of apolipoprotein B-100 centered tryptophan radicals in the initiation of lipid peroxidation", FEBS LETT., vol. 389, 1996, pages 136 - 140
A. M. S. SILVA; P. FILIPE; R. S. G. R. SEIXAS; D. C. G. A. PINTO; L. K. PATTERSON; G. L. HUG; J. A. S. CAVALEIRO; J.-C. MAZIÈRE;: "One-electron reduction of superoxide radical-anions by 3-alkylpolyhydroxyflavones in micelles. Effect of antioxidant alkyl chain length on micellar structure and reactivity", J. PHYS. CHEM. B, vol. 112, 2008, pages 11456 - 11461
A. M. S. SILVA; P. FILIPE; R. S. G. R. SEIXAS; D. C. G. A. PINTO; L. K. PATTERSON; G. L. HUG; J. A. S. CAVALEIRO; J.-C. MAZIÈRE;: "One-electron reduction of superoxide radical-anions by 3- alkylpolyhydroxyflavones in micelles. Effect of antioxidant alkyl chain length on micellar structure and reactivity", J. PHYS. CHEM. B, vol. 112, 2008, pages 11456 - 11461
A. J. N. SELLÉS; H. T. V. CASTRO; J. AGÜERO-AGÜERO; J. GONZÁLEZ-GONZÁLEZ; F. NADDEO; F. DE SIMONE; L. RASTRELLI: "Isolation and quantitative analysis of phenolic antioxidants, free sugars, and polyols from Mango (Mangifera indica L.) stem bark aqueous decoction used in Cuba as a nutritional supplement", J. AGRIC. FOOD CHEM., vol. 50, 2002, pages 762 - 766, XP009049806, DOI: doi:10.1021/jf011064b
P. FILIPE; A. M. S. SILVA; R. S. G. R. SEIXAS; D. C. G. A. PINTO; A. SANTOS; L. K. PATTERSON; J. N. SILVA; J. A. S. CAVALEIRO; J.: "The alkyl chain length of 3-alkyl-3',4',5,7-tetrahydroxyflavones modulates effective inhibition of oxidative damage in biological systems: Illustration with LDL, red blood cells and human skin keratinocytes", BIOCHEM. PHARMACOL., vol. 77, 2009, pages 957 - 964, XP025990020, DOI: doi:10.1016/j.bcp.2008.11.023
H. ESTERBAUER; G. STRIEGL; H. PUHL; M. ROTHENEDER.: "Continuous monitoring of in vivo oxidation of human low density lipoprotein", FREE RAD. RES. COMMUN., vol. 6, 1989, pages 67 - 75
D. K. DAS; N. MAULIK.: "Resveratrol in cardioprotection: a therapeutic promise of alternative medicine", MOL. INTERV., vol. 6, 2006, pages 36 - 47
H. YOSHIDA; K. SASAKI; Y. NAMIKI; N. SATO; N. TADA.: "Edavarone, a novel radical scavenger, inhibits oxidative modification of low-density lipoprotein (LDL) and reverses oxidized LDL-mediated reduction in the expression of endothelial nitric oxide synthase", ATHEROSCLEROSIS, vol. 179, 2005, pages 97 - 102
Attorney, Agent or Firm:
VIEIRA PEREIRA FERREIRA, Maria Silvina (Modet & Co.Rua Castilh, 50-9º -163 Lisboa, PT)
Download PDF:
Claims:
Claims

is claimed is:

A composition comprising polyhydroxy-2 , 3-diaryl-9H- xanthen-9-ones represented by one of the following formulas (1), (2), (3), (4) and (5)

2. Composition accordingly with claim 1 wherein the polyhydroxy-2 , 3-diaryl-9H-xanthen-9-ones are antioxidant agents.

3. Use of a composition according to previous claims wherein the polyhydroxy-2 , 3-diaryl-9H-xanthen-9-ones represented by one of the following formulas (1), (2), (3), (4) and (5) act as therapeutic agents.

4. Use according to claims 1-3, wherein polyhydroxy-2 , 3- diaryl-9H-xanthen-9-ones react with reactive oxygen species, organic radicals and peroxyl radicals.

5. Use of a composition according to any of claims 1-4, in the production of a drug to protect against cutaneous aging and cancers induced by the oxidative stress in human skin.

6. Use of a composition according to any of the claims 1-4 in the production of a drug to prevent LDL from oxidation involved in atherosclerosis and more generally in all related cardiovascular diseases.

7. Use of a composition according to any of the claims 1-4 in the production of a drug for detoxification of liver injury to prevent damage from halogenated organic peroxyl radicals induced by polyhalogenated anaesthetics and other environmental polyhalogenated chemicals .

8. Use of a composition according to any of the claims 1-4 in the production of a drug for any pathology in which the antioxidant therapy can have beneficial effects such as carcinogenesis, diabetes-related vascular disorders, acute inflammatory and ischemia- reperfusion phases of pathologies related to neurology, cardiology and rheumatology and in chronic preventive therapy in :ancer and inherited and acquired neurodegenerative diseases .

Description:
DESCRIPTION

HYDROXY-2, 3-DIARYLXANTHONES AS THERAPEUTIC AGENTS OF PATHOLOGIES DUE TO REACTIVE OXYGEN SPECIES"

Technical field

The present invention relates to the use of hydroxyl-2 , 3- diarylxanthones (DAX) as therapeutic agents of pathologies due to reactive oxygen species.

Background art

Xanthones are natural oxygen heterocyclic compounds found in higher plants, lichens and fungi. Members of this class of compounds have, among other, antiallergic, anti ¬ inflammatory, antitumor activities.

There are currently on the market two formulations with a complex composition of oxygenated and prenylated xanthones extracted from mango A. J. N. Selles, H. T. V. Castro, J. Aguero-Aguero, J. Gonzalez-Gonzalez, F. Naddeo, F. De Simone, L. Rastrelli. Isolation and quantitative analysis of phenolic antioxidants, free sugars, and polyols from Mango (Mangifera indica L.) stem bark aqueous decoction used in Cuba as a nutritional supplement, J. Agric. Food Chem. 2002, 50, 762-766] [A. R. Garrity, G. A. Morton, J. C. Morton. Nutraceutical mangosteen composition, US patent 6730333, 2004]. The strong antioxidant activity of these formulations observed in vivo in animals and humans prompted us to synthesize this family of polyphenolic compounds with a well-defined chemical structure [C. M. M. Santos, A. M. S. Silva, J. A. S. Cavaleiro. A novel and efficient route for the synthesis of hydroxylated 2,3- diarylxanthones , Eur. J. Org. Chem. 2009, 2642-2660] and to assess their antioxidant potential using as reference the flavonol quercetin (Q) one of the most effective antioxidant agents of the flavonoid family.

The literature identifies two classes of antioxidants used in beauty care or clinical applications. The polyphenolic derivatives, where emerges resveratrol, present in many cosmetic preparations and promise future in therapeutic use due to their action in cardiovascular diseases [D. K. Das, N. Maulik. Resveratrol in cardioprotection : a therapeutic promise of alternative medicine, Mol. Interv. 2006, 6, 36- 47]. Edavarone ( 3-methyl-l-phenyl-2-pyrazoline-5-one ) is active in its phenolic tautomeric form [T. Watanabe, M. Tahara, S. Todo . The novel antioxidant edavarone: From bench to bedside; Cardiovascular Therapeutics 2008, 26, 101-114] [H. Yoshida, K. Sasaki, Y. Namiki, N. Sato, N. Tada. Edavarone, a novel radical scavenger, inhibits oxidative modification of low-density lipoproteins (LDL) and reverses oxidized LDL-mediated reduction in the expression of endothelial nitric oxide synthase, Atherosclerosis, 2005, 179, 97-102]. This drug has been lately authorized in Japan under the name Radicut (Radicut®, registered trademark) is used together with nitrones, as the NXY-059 developed by AstraZeneca in Europe, in the treatment of the AVC with contestable results [R. A. Floyd, R. D. Kopke, C. H. Choi, S. B. Foste, S. Doblas, R. A. Towner. Nitrones as therapeutics. Free Radic. Biol. Med. 2008, 45, 1361-1374] [K. R. Lees, J. A. Zivin, T. Ashwood, A. Davalos, S. M. Davis, H. C. Diener, J. Grotta, P. Lyden, A. Shuaib, H. G. Hardemark, W. W. Wasiewski. Stroke-acute ischemic NXY treatment (SAINT I) trial investigators, N. Engl. J. Med. 2006, 354, 588-600]. Therefore these publications encourage the study of the antioxidant capacity or the radical scavenging activity of these molecules as important factors of their activity. We can illustrate the superiority of our derivatives DAX over edavarone because the evaluation of their antioxidant capacity involves the use of the copper (II) induced low density lipoprotein (LDL) oxidation model. It must be noted that the antioxidant index of DAX measured as the lag time for the beginning of LDL peroxidation for an equivalent ratio of [DAX] / [LDL] (Fig. 1) [H. Yoshida, K. Sasaki, Y. Namiki, N. Sato, N. Tada. Edavarone, a novel radical scavenger, inhibits oxidative modification of low-density lipoprotein (LDL) and reverses oxidized LDL-mediated reduction in the expression of endothelial nitric oxide synthase, Atherosclerosis, 2005, 179, 97-102] is approximately forty times superior with that of edavarone for the CS9, the "least" powerful of DAX. We can estimate from data on the oxidation of porcine LDL [L. Belguendouz, L. Fremont, A. Linard. Resveratrol inhibits metal ion- dependent and independent peroxidation of porcine low- density lipoproteins, Biochem. Pharmacol. 1997, 53, 1347- 1355] that for the same ratio [DAX] / [LDL] , the CS9 is at least equivalent to trans-resveratrol , itself higher than Trolox, the water-soluble derivative of vitamin E.

Summary

The present invention relates to a composition comprising polyhydroxy-2 , 3-diaryl-9H-xanthen-9-ones that are antioxidant agents and acting as therapeutic agents.

In a preferable embodiment the polyhydroxy-2 , 3-diaryl-9H- xanthen-9-ones react with reactive oxygen species, organic radicals and peroxyl radicals.

In another preferable embodiment the composition is used to produce a drug: to protect against cutaneous aging and cancers induced by the oxidative stress in human skin.

to prevent LDL from oxidation involved in atherosclerosis and more generally in all related cardiovascular diseases.

for detoxification of liver injury to prevent damage from halogenated organic peroxyl radicals induced by polyhalogenated anaesthetics and other environmental polyhalogenated chemicals.

for any pathology in which the antioxidant therapy can have beneficial effects such as carcinogenesis, diabetes-related vascular disorders, acute inflammatory and ischemia-reperfusion phases of pathologies related to neurology, cardiology and rheumatology and in chronic preventive therapy in cancer and inherited and acquired neurodegenerative diseases .

Detailed disclosure of the invention

Xanthones are natural oxygen heterocyclic compounds found in higher plants, lichens and fungi. Members of this class of compounds have, among other, antiallergic, anti ¬ inflammatory, antitumor activities. One of the most interesting properties is their strong antioxidant activity due to the presence in some key positions of hydroxyl groups and / or a catechol moiety. There are currently on the market two formulations with a complex composition of oxygenated and prenylated xanthones extracted from mango [J. N. Selles, H. T. V. Castro, J. Aguero-Aguero, J. Gonzalez-Gonzalez, F. Naddeo, F. De Simone, L. Rastrelli. Isolation and quantitative analysis of phenolic antioxidants, free sugars, and polyols from Mango (Mangifera indica L.) stem bark aqueous decoction used in Cuba as a nutritional supplement, J. Agric. Food Chem. 2002, 50, 762-766] [A. R. Garrity, G. A. Morton, J. C. Morton. Nutraceutical mangosteen composition, US patent 6730333, 2004]. The strong antioxidant activity of these formulations observed in vivo in animals and humans prompted us to synthesize this family of polyphenolic compounds with a well-defined chemical structure [C. M. M. Santos, A. M. S. Silva, J. A. S. Cavaleiro. A novel and efficient route for the synthesis of hydroxylated 2,3- diarylxanthones , Eur. J. Org. Chem. 2009, 2642-2660] and to assess their antioxidant potential using as reference the flavonol quercetin, one of the most effective antioxidant agents of the flavonoid family. This work allows us to select five polyhydroxyxanthones based on the exceptional level of antioxidant protection obtained in vitro on two biological systems of medical interest easily reproducible and with increasing complexity: the copper ( I I ) -induced oxidation of low density lipoproteins (LDLs) from human blood and the oxidative stress induced by tert-butyl hydroperoxide (t-BuOOH) in cultures of proliferating human skin keratinocytes .

The chemical structures of the evaluated five more active synthetic compounds (CS3, CS6, CS7, CS8 and CS9) are represented by the following formulas (1), (2), (3), (4) e (5) . The novel and efficient route developed allows the synthesis of these polyhydroxydiarylxanthones (DAX) in good yields (40 to 60%) [C. M. M. Santos, A. M. S. Silva, J. A. S. Cavaleiro. A novel and efficient route for the synthesis of hydroxylated 2 , 3-diarylxanthones , Eur. J. Org. Chem. 2009, 2642-2660]. The methodology for their selection was based in comparison of their antioxidant activity compared to that of quercetin (Q) in the model Cu ( 11 ) -induced LDL oxidation and in the oxidative stress induced by t-BuOOH in cultures of proliferating human skin keratinocytes , already reported in our recent publications concerning another antioxidant family, the polyhydroxy-3-alkylflavones [P. Filipe, A. M . S. Silva, R. S. G. R. Seixas, D. C. G. A. Pinto, A. Santos, L. K. Patterson, J. N. Silva, J. A. S. Cavaleiro, J. P. Freitas, J.-C. Maziere, R. Santus, P. Morliere. The alkyl chain length of 3-alkyl-3 ' , 4 ' , 5 , 7- tetrahydroxyflavones modulates effective inhibition of oxidative damage in biological systems: Illustration with LDL, red blood cells and human skin keratinocytes , Biochem. Pharmacol. 2009, 77, 957-964].

The present invention is also related to the use of polyhydroxy-2 , 3-diaryl-9H-xanthen-9-ones as medicines against cancer and cutaneous ageing and for treatment of cardiovascular diseases and diabetes-related vascular disorders, acute inflammatory and ischemia-reperfusion phases of pathologies related to neurology, cardiology and rheumatology and chronic preventive therapy in cancer and inherited and acquired neurodegenerative diseases. Brief description of the figures

Fig. 1 shows the kinetics of conjugated dienes formation during LDL oxidation induced by 5 μΜ Cu 2+ . The LDL phosphate buffer solutions at pH 7.4 contain 0.12 mg protein/ml (240 nM) and were incubated during 15 min at 37°C in the presence or absence of 0.75 μΜ of antioxidant. The absorbance is read at 234 mm. The points are the average of 4 independent experiments. The error bars are not indicated for clarity of presentation of the results. The vertical axis denotes the absorbance and the horizontal axis the time in minutes.

Fig. 2 shows the kinetics of carotenoids consumption under the same conditions as those of figure 1. The points are the average of 4 independent experiments. The error bars are not indicated for clarity of presentation of the results. The vertical axis denotes the percentage of carotenoids consumed and the horizontal axis the time in minutes .

Fig. 3 shows the effect of polyhydroxyxanthones and quercetin on the survival of human NCTC 2544 keratinocytes . The cells are incubated in absence or in presence of antioxidant at concentrations of 0.5 μΜ, 2 μΜ and 5 μΜ. The viability test to the Neutral Red was used according to the recommendations of the "Commission Directive 2000/33/EC" published in the Journal of the European Community of 8.6.2000: L136/90-L/07. The vertical axis shows all the samples tested with the concentration defined and the horizontal axis shows the percentage of human NCTC 2544 keratinocytes survival.

Fig.4 shows the transient absorbance spectra of radicals •CS3 and *CS8 observed in pH 7 aqueous solutions, saturated with oxygen containing 10 mM of CTAB, 0,1 M of formate ions and 200 μΜ of CS after production of 4,5 μΜ (CS3) and 3.2 μΜ (CS8) of superoxide anion radical ( ·02 ~ ) by a radiolytic pulse dose : 7 Gy for CS3 and 5 Gy for CS8) . Kinetics of •CS radicals formation under the same experimental conditions. The vertical axis shows the absorbance of the samples and the horizontal axis shows the wavelength in nanometers .

Examples

For an easier understanding of the invention, examples of preferred achievements of the invention are described below, which, however, are not intended to limit the object of this invention.

Example 1 - LDL protection

It is common knowledge that LDLs are natural carriers of important antioxidants such as vitamin E and carotenoids. Because antioxidants compete with LDL lipids for the propagation of the radical chain reactions, a lag time, due to the antioxidant consumption, is observed between the start of the oxidation by Cu(II) ions and the appearance of lipid peroxidation products. The duration of this induction period depends on the constitutive antioxidant content of the LDLs which may vary among blood donors [H. Esterbauer, G. Striegl, H. Puhl, M. Rotheneder . Continuous monitoring of in vivo oxidation of human low density lipoprotein, Free Rad. Res. Commun . 1989; 6, 67-75] . The LDL peroxidation results in the formation of conjugated dienes. The kinetics represented in Fig. 1 are performed at 37°C by measuring the absorbance of the LDL solutions at 234 nm, after the addition of Cu(II) . It is important to notice that for the native LDL without other additives than Cu(II) ions, the formation of conjugated dienes requires a latency time that increases according to a sigmoid curve as shown in Fig.l. Latency time translates the inhibition of lipid peroxidation by natural antioxidants (vitamin E and carotenoids) transported by native LDLs. Using the time interval between the addition of Cu(II) and the passage in the center of sigmoid curve as indicator of the antioxidant capacity of LDL solutions, the addition of 0.75 μΜ of CS3, CS6, CS7, CS8 and CS9, slows down considerably the formation of these dienes compared to that observed in the presence of 0.75 μΜ of quercetin, thus illustrating their strong antioxidant potential. It is observed, for example, that xanthone CS3, the most effective, introduced a delay three times longer than that of Q.

The consumption of antioxidants is highlighted by the disappearance of carotenoids, as shown in Fig.2, following the kinetics of the characteristic absorbance decreasing at 400-500 nm, after the addition of Cu(II) ions. The protective capacity of the antioxidants DAX with respect to the consumption of the carotenoids parallels that observed with the production of dienes. The error bars are not indicated for clarity of presentation of the results.

Example 2 - Inhibition of the cytotoxicity of tert-butyl hydroperoxide

The lipid membrane peroxidation is a key-process in many cutaneous lesions [D. R. Bickers, M. Athar . Oxidative stress in the pathogenesis of skin disease, J Invest. Dermatol. 2006, 126, 2565-2575] . At the molecular level, it leads not only to the modification of the lipid double- layers but the free radicals produced by the chain reactions of lipid peroxidation oxidize certain amino acids residues of proteins like Trp, His or Cys causing their inactivation . The oxidative stress of human NCTC 2544 keratinocytes culture was generated by 5 mM of t-BuOOH a membrane-permeant oxidant extensively used to induce oxidative stress in different systems. Its tert-butyl group confers lipid solubility leading to the formation of reactive oxygen species (ROS) comparable to those met during the propagation of the chain reactions of the lipid peroxidation. From Fig. 3 one can see that with a 0.5 μΜ concentration of CS3, CS9, CS8, CS7, CS6 or Q, only the diarylxanthones substantially protect human keratinocytes against the cytotoxic effect of t-BuOOH.

Example 3 - Mechanistic stage : Reactivity of CS with superoxide anion radical and peroxo-trichloromethyl radicals

The reactivity of DAX with superoxide anion radicals and peroxo-trichloromethyl radicals - a good model of organic peroxide oxidant implied in the carbon tetrachloride hepatotoxicity and prototypes of oxyradicals produced by chlorinated anesthesics - was measured by fast kinetic spectroscopy in period of time from microseconds (ps) to several milliseconds (ms) by pulse radiolysis. Superoxide anion radical can be selectively produced in some ps from primary radicals of water radiolysis and trapped stoichiometrically by positively charged micelles of cetyl trimethyl ammonium bromide in which DAX are soluble. One of our previous papers reports a parallel work on polyhydroxy- 3-alkylflavones and describes the experimental procedure and subjacent methodology [A. M. S. Silva, P. Filipe, R. S. G. R. Seixas, D. C. G. A. Pinto, L. K. Patterson, G. L. Hug, J. A. S. Cavaleiro, J.-C. Maziere, R. Santus, P. Morliere. One-electron reduction of superoxide radical- anions by 3-alkylpolyhydroxyflavones in micelles. Effect of antioxidant alkyl chain length on micellar structure and reactivity, J. Phys . Chem. B 2008, 112, 11456-11461]. As an example, (Fig.4) shows for CS3 and CS8, the kinetics of formation and the transient absorption spectra of neutral radical " DAX produced by one-electron oxidation reaction:

"( + DAX(H) → " DAX + H02 ~ [1]; where (H) represents the hydrogen atom of an hydroxyl group of DAX, the electron donor in that reaction. The radical " DAX is perfectly stable over at least 50 ms (not shown) and the protonation of H02 ~ leads to H 2 O 2 which can be eliminated by catalase. Kinetic treatment carried out according to that described in r7 led to reaction rate constant in the order of 106 NT 1 s _1 for reaction [1] . The ' CCI 3 O 2 radicals due to their higher redox potential than that of Ό2 ~ , oxidize DAX very quickly by a diffusion process according to the main reaction " CC1 3 0 2 + CS (H) → ' CS + CCI 3 OOH [2], whose bimolecular reaction rate constant is > 10 8 M _1 s _1 for CS3, CS6, CS 7 , CS8, CS9.

It is necessary to envisage the remarkable parallel between the reactivity of DAX with ROS and their biological activity. For example, compound CS5, an isomer of CS3 with two OH groups in positions 4' and 4", inactive in biological systems, does not react with " 02 and it is five times less reactive than CS3 with the ' CCI 3 O 2 radicals The 2 , 3-diphenyl-9H-xanthene-9-one (CS1) without hydroxyl groups is, as expected, completely inactive. Example 4 - Repair of * Trp radicals by CS, produced by oxidation of Trp

The tryptophan (Trp) residues are particularly sensitive targets of apolipoprotein-B (Apo-B) during copper (II) induced LDL oxidation [A. M. S. Silva, P. Filipe, R. S. G. R. Seixas, D. C. G. A. Pinto, L. K. Patterson, G. L. Hug, J. A. S. Cavaleiro, J.-C. Maziere, R. Santus, P. Morliere. One-electron reduction of superoxide radical-anions by 3- alkylpolyhydroxyflavones in micelles. Effect of antioxidant alkyl chain length on micellar structure and reactivity, J. Phys. Chem. B 2008, 112, 11456-11461] [A. B. Giessauf, B. van Wickern, T. Simat, H. Steinhart, H. Esterbauer. Formation of N-formylkynurenine suggests the involvement of apolipoprotein B-100 centered tryptophan radicals in the initiation of lipid peroxidation, FEBS Lett. 1996, 389, 136-140] and they are formed in the circulating proteins during the photo-oxidant stress induced on skin by UV or cutaneous photosensibilization reactions. We have studied if, like some flavonoids [A. M. S. Silva, P. Filipe, R. S. G. R. Seixas, D. C. G. A. Pinto, L. K. Patterson, G. L. Hug, J. A. S. Cavaleiro, J.-C. Maziere, R. Santus, P. Morliere. One-electron reduction of superoxide radical- anions by 3-alkylpolyhydroxyflavones in micelles. Effect of antioxidant alkyl chain length on micellar structure and reactivity, J. Phys. Chem. B 2008, 112, 11456-11461], DAX compounds could repair the " Trp radicals. We have used pulse radiolysis to selectively prepare these radicals in a few tens of ps [A. M. S. Silva, P. Filipe, R. S. G. R. Seixas, D. C. G. A. Pinto, L. K. Patterson, G. L. Hug, J. A. S. Cavaleiro, J.-C. Maziere, R. Santus, P. Morliere. One-electron reduction of superoxide radical-anions by 3- alkylpolyhydroxyflavones in micelles. Effect of antioxidant alkyl chain length on micellar structure and reactivity, J. Phys. Chem. B 2008, 112, 11456-11461]. Concerning the DAX derivatives, we observed that the reaction of repair " Trp + CS (H) —> " CS + Trp [3] is very effective and occurs with a reaction rate constant > 10 7 M^s -1 . These results showed that the repair reaction of the protein components must also be taken into account in the antioxidant potential of DAX.

Industrial Applicability

Hydroxy-2 , 3-diarylxanthones are excellent/promising therapeutic agents of pathologies due to reactive oxygen species especially in the treatment of cancer and cutaneous ageing in treatment of cardiovascular diseases and diabetes-related vascular disorders, acute inflammatory and ischemia-reperfusion phases of pathologies related to neurology, cardiology and rheumatology and chronic preventive therapy in cancer and inherited and acquired neurodegenerative diseases.

References cited in the description

Non-patent literature cited in the description rl. A. J. N. Selles, H. T. V. Castro, J. Aguero-Aguero, J.

Gonzalez-Gonzalez, F. Naddeo, F. De Simone, L. Rastrelli. Isolation and quantitative analysis of phenolic antioxidants, free sugars, and polyols from Mango (Mangifera indica L.) stem bark aqueous decoction used in Cuba as a nutritional supplement, J. Agric. Food Chem. 2002, 50, 762-766. r2. A. R. Garrity, G. A. Morton, J. C. Morton.

Nutraceutical mangosteen composition, US patent 6730333, 2004.

r3. C. M. M. Santos, A. M. S. Silva, J. A. S. Cavaleiro. A novel and efficient route for the synthesis of hydroxylated 2 , 3-diarylxanthones , Eur. J. Org. Chem. 2009, 2642-2660.

r4. P. Filipe, A. M. S. Silva, R. S. G. R. Seixas, D. C. G.

A. Pinto, A. Santos, L. K. Patterson, J. N. Silva, J. A. S. Cavaleiro, J. P. Freitas, J.-C. Maziere, R. Santus, P. Morliere. The alkyl chain length of 3-alkyl- 3 ', 4 ', 5, 7-tetrahydroxyflavones modulates effective inhibition of oxidative damage in biological systems: Illustration with LDL, red blood cells and human skin keratinocytes , Biochem. Pharmacol. 2009, 77, 957-964. r5. H. Esterbauer, G. Striegl, H. Puhl, M. Rotheneder .

Continuous monitoring of in vivo oxidation of human low density lipoprotein, Free Rad. Res. Commun. 1989; 6, 67-75.

r6. D. R. Bickers, M. Athar . Oxidative stress in the pathogenesis of skin disease, J Invest. Dermatol. 2006, 126, 2565-2575.

r7. A. M. S. Silva, P. Filipe, R. S. G. R. Seixas, D. C. G.

A. Pinto, L. K. Patterson, G. L. Hug, J. A. S. Cavaleiro, J.-C. Maziere, R. Santus, P. Morliere. One- electron reduction of superoxide radical-anions by 3- alkylpolyhydroxyflavones in micelles. Effect of antioxidant alkyl chain length on micellar structure and reactivity, J. Phys . Chem. B 2008, 112, 11456-11461 r8. A. B. Giessauf, B. van Wickern, T. Simat, H. Steinhart, H. Esterbauer. Formation of N-formylkynurenine suggests the involvement of apolipoprotein B-100 centered tryptophan radicals in the initiation of lipid peroxidation, FEBS Lett. 1996, 389, 136-140. D. K. Das, N. Maulik. Resveratrol in cardioprotection : a therapeutic promise of alternative medicine, Mol. Interv. 2006, 6, 36-47.

T. Watanabe, M. Tahara, S. Todo. The novel antioxidant edavarone: From bench to bedside; Cardiovascular Therapeutics 2008, 26, 101-114.

H. Yoshida, K. Sasaki, Y. Namiki, N. Sato, N. Tada. Edavarone, a novel radical scavenger, inhibits oxidative modification of low-density lipoprotein (LDL) and reverses oxidized LDL-mediated reduction in the expression of endothelial nitric oxide synthase, Atherosclerosis, 2005, 179, 97-102.

R. A. Floyd, R. D. Kopke, C. H. Choi, S. B. Foste, S. Doblas, R. A. Towner. Nitrones as therapeutics. Free Radic. Biol. Med. 2008, 45, 1361-1374.

K. R. Lees, J. A. Zivin, T. Ashwood, A. Davalos, S. M. Davis, H. C. Diener, J. Grotta, P. Lyden, A. Shuaib, H. G. Hardemark, W. W. Wasiewski. Stroke-acute ischemic NXY treatment (SAINT I) trial investigators, N. Engl. J. Med. 2006, 354, 588-600.

L. Belguendouz, L. Fremont, A. Linard. Resveratrol inhibits metal ion-dependent and independent peroxidation of porcine low-density lipoproteins, Biochem. Pharmacol. 1997, 53, 1347-1355.