EP1225883A1

EP1225883A1 - Anti-ischemic compounds

Info

Publication number: EP1225883A1
Application number: EP00969499A
Authority: EP
Inventors: Carine Michiels; Martine Redon; José REMACLE
Original assignee: Societe de Conseils de Recherches et dApplications Scientifiques SCRAS SAS
Current assignee: Ipsen Pharma SAS
Priority date: 1999-10-15
Filing date: 2000-10-11
Publication date: 2002-07-31
Also published as: AU783713B2; US6780887B1; CA2387164A1; AU7919200A; WO2001028549A1; EP1092432A1

Abstract

The invention concerns a compound of formula (I), an optional salt or a prodrug thereof corresponding to formula (I) and its use for preventing and/or treating partial or total ischemia or pathologies associated with ischemia or associated with mitochondrial deficiencies.

Description

AJMTI-ISCHEMIC COMPOUNDS

Subject of the invention

The present invention relates to new therapeutic agents having a protective effect on the protemic complexes of the internal mitochondπal membrane, and which can be preferably used for the prevention and / or treatment of partial or total ischemia, pathologies associated with ischemia or associated with mitochondrial deficiencies, or apoptosis.

Technological background underlying the invention

Ischemia is an interruption of the blood supply to an organ or tissue by obliteration, compression or spasm of an artery. The consequences depend on the nature of the tissue deprived of oxygenated blood and the duration of the phenomenon. Total ischemia causes tissue death in a variable time depending on its oxygen requirements. Partial ischemia causes either a decrease in the organ's capacities (mental slowing down, insufficiency of the hepatic or renal functions, etc.), or the inability to modify its work at any stimulation, thus showing signs of suffering (crises angina pectoris, cramps of the artery). Ischemia is very harmful to the nervous tissue. Initially, it results in

confirmation copy reversible metabolic and functional disturbances which may give rise to abnormal nervous activity, for example an epileptic fit or certain ischemic damage to the cerebral cortex. After a few minutes of ischemia, irreversible lesions of the nervous tissue settle, with swelling then necrosis of the cell bodies. It is cerebral infarction or softening of the brain. It is known to use agonist molecules of alpha-adrenergic receptors and agonists of alpha-presynaptic receptors for the treatment of ischemia. These compounds are used at the heart level for the treatment of angina, which is a clinical expression of acute myocardial ischemia, and which is the result of a momentary imbalance between the oxygen demand of the myocardium and the oxygen supply. by circulation, which can lead in severe cases to a myocardial infarction.

State of the art

The international patent application WO98 / 51291 describes the use of different plant extracts for the treatment and / or prevention of ischemia or of pathologies associated with ischemia or with an energy deficit. This document also describes the mechanism of action of these molecules which allow inhibition of the activation cascade of endothelial cells induced by hypoxia as well as the protective effect of these products on certain mitochondπal deficiencies, in particular by a protective effect. of complexes I and III of the mitochondπes. This document proposes a biochemical model making it possible to obtain a protective effect of anti-ischemic compounds. The publication of Ukramets I. et al.

(Tetrahedron Let ters Vol. 50 No. 34 pp. 10331-10338 (1994)) describes derivatives and synthesis of malonic acid, in particular an ethylonic malonic acid and symmetrical diamylides.

International patent application O87 / 05898 describes derivatives of malonic acid and their use to retard the growth of plants. The publication of Venuti et al. (J. of Med. Chem. Vol. 31 pp. 2145-2152 (1988)) described as an intermediate compound, a derivative of malonic acid involved in the synthesis of compounds used as prodrugs. The publication of Oumar-Mahamat et al. (Tetrahedron Letters Vol. 30 No. 3 pp. 331-332 (1989)) describes cyclic derivatives of malonic acid.

Patent application EP-A-0 099 091 describes hexahydrodioxopyrimidme derivatives, their process for obtaining and their use as antiviral, antibacterial and antitumour compounds.

Patent application FR-2,539,412 describes 5-fluorouracyl derivatives and their therapeutic use as carcostatic agents. The publication of Holwood et al. (J ^" . Of Med. Chem. Vol. 10 No. 5 pp. 863-867 (1967)) described as local anesthetics, various derivatives, in particular 2-ethoxycarbonyl aceto-2 ', 6' -xylidide or its corresponding amide.

The publication of McLaughlm et al. (J. of Neurochem. Vol. 70 pp. 2406-2415 (1998)) describes the effect of dopamma on mitochondrial inhibition compared to that of methylmalonate.

Patent application WO97 / 16184 describes methods of treating ischemia consisting of: administration of an acyl inhibitor co-enzyme A cholesterol O acyltransferase (ACAT) such as N (2, 6-dιιsopropyl -phenyl) -2 -phenyl-malonamic acid dodecyl ester and an inhibitor of HMG-co- enzyme A reductase.

Aims of the invention

The present invention aims to provide new compounds intended to induce a protective effect on the protemic complexes of the internal mitochondπal membrane and to be used in particular in the prevention or treatment of partial or total ischemia, of related pathologies ischemia or pathologies related to mitochondπal deficiencies.

A particular object of the present invention is to provide compounds having improved activity and / or not having the side effects of molecules of the prior art.

Characteristic Elements of the Invention The present invention relates in particular to new compounds of formula I, their salts or prodrugs of such compounds:

⁽ D ⁿ

in which: - R ¹ is CH2, NH, or a ligand, preferably a carbon atom, forming with R3 and / or R ⁴ an aromatic or non-aromatic ring of 5 or 6 carbon atoms (n = 1 or 2) , possibly including on each cycle a heteroatom, preferably a nitrogen, oxygen or sulfur atom; R ³ and / or R ⁴ represent:

-an amine of formula -NR ^, or R ⁶

a group of formula -Z-R ^ in which:

- Z represents O or S,

- R5 represents H or an alkyl of 1 to 6 carbon atoms (preferably, R ^ represents a methyl or an ethyl),

R ^ represents a hydrophobic group, preferably chosen from the group consisting of a terbutyl, an allyl or a ring (aromatic or non-aromatic) of 5, 6 or 7 carbon atoms optionally comprising one or more heteroatoms (preferably N, O or S) and in which the carbon atoms are optionally substituted by:

an alkyl group R ⁷ of 1 to 6 carbon atoms (preferably a methyl or an ethyl), a ketone group,

- a hydroxyl group,

- an ester group,

- a halogen element (F, Cl, Br or I),

- a trifluoromethyl (CF3), or - a group of formula

- in which: - W represents CH or a nitrogen atom,

- Y represents S or O, and - R8 represents an alkyl group of 1 to 6 carbon atoms (preferably methyl or ethyl);

- with the additional condition that if R ³ forms with said R ¹ said cycle above or that if R ³ is either an amine of formula

-N-R5 OR the group of formula -Z-R ^

mentioned above, R ⁴ represents an alkyl (preferably propene) of 1 to 10 (preferably 1 to 6) carbon atoms, saturated or unsaturated, optionally comprising one or more heteroatoms (preferably N, S or O). In addition, in said alkyl, one or more carbons may optionally be substituted by a halogen element (fluorine, chromium, bromine or iodine) or a trifluoromethyl (CF3).

In the compounds of formula I, R ³ and R ⁴ preferably represent an amine of formula -NR ^ or

\

Rδ a group of formula -ZR ⁶ in which R ⁵ represents H or methyl, Z represents O or S (preferably O), and R ^ represents a cyclopentene, a cyclohexane or a benzene group.

According to the invention, are excluded as compounds de] to described in the prior art, the compounds of formula I in which R ¹ is CH2; R ³ is a compound of formula ZR ⁶ in which Z represents O, R ^ represents an ethyl, R ³ is an amine of formula NH-R ⁶ 'in which R ^' represents a cyclic compound (optionally substituted in position 2 by a hydroxyl group) and the compounds of formula I in which R ^ is CH2; R ³ and R ⁴ are amines of formula NH-R ⁶ in which R ⁶ represents a ring with 6 carbon atoms substituted in position 2 by a hydroxyl group. Are also excluded from the invention as compounds described in the compounds described in Tables 1, 2 and A of application O87 / 05898, N- (2,6- dusopropyl-phenyl) -2 - phenyl-malonamic acid dodecyl ester and the compound of formula I in which Ri is CH2, R ⁴ is a group of formula ZR ^ (Z represents oxygen and R ⁶ represents alkyl) and in which R ³ is an amme of formula II : NO alkyl

S alkyl

II o

in which X represents O, S, CH2 or C = 0 and the derivatives of formula

in which R ¹ represents a C1-C6 alkyl group, A represents a saturated or msaturated ring containing 3 to 7 carbon atoms; each of the symbols R ² , R ³ and R ⁴ independently represents a hydrogen or halogen atom or a group such as a C1-C6 alkoxy group, C1-C6 alkyl, C1-C6 alkylthio, C1-C6 alkoxycarbonyl, carboxyl, carboxamido, sulfonic acid, sulfonamido, acylamino, sulfonylammo, C1-C6 alkylsulfonyde, nitπle, nitro, acyloxy, phenyl or methylene -dioxy and each of the symbols m and n represents an integer ranging from 0 to

AT

in which

RA represents an alkyl, R ² a hydrogen or an alkyl

in which Z = CO Methyl, COO Methyl, or COO Alkyl O

Z = C0 ₂ CH ₃ , C0 ₂ C ₂ H ₃ or C0 ₂ H

N H

According to the invention, the aromatic or non-aromatic rings optionally comprise one or more heteroatoms, preferably one or more oxygen or sulfur atoms, and in which the atoms of carbon are substituted by ketone, hydroxyl or ester groups or by a halogen element as described above.

The preferred compounds of the invention are chosen from the group consisting of dιcyclopenten-2 -yle malonate, 2-propenyl 3 -oxa-2 -oxobicyclo

[3.1.0] hexane-1-carboxylate, cyclopenten-2-yl and ethyl malonate, cyclopenten-2-methyl and methyl malonate, N, N '-diphenyl malonamide, N-phenylamido ethanoate ethyl, 2, 6-methyl-N-phenylamιdo ethyl ethanoate, ethyl N-methyl -N-cyclohexylamido ethanoate and dιcyclopentene-2 -yl malonate, optionally their salts or prodrugs thereof this. The present invention also relates to the pharmaceutically acceptable non-toxic acid addition salts of the compounds of the invention comprising an amine. As examples of pharmaceutically acceptable acids, mention may be made of mineral acids such as hydrochloric, hydrobromic, sulphuric, nitric, phosphoric acids, etc. and organic acids such as acetic, citric, tartaric, benzoic, salicylic, maleic, etc.

The present invention also relates to the first application of the compounds of formula I as a medicament, as well as their salts and the prodrugs of said compounds, with the exception of the following known compounds: N- (2, 6-dιιsopropyl- phenyl) -2 -phenyl-malonamic acid dodecyl ester and the compounds of formula I in which R ^ is

CH ₂ , R ⁴ is a group of formula ZR ⁶ in which Z represents an oxygen and R ° represents an alkyl and in which R ³ is a cord of formula II: NO alkyl

\ He

S X alkyl

II O

in which the X represents O, S, CH2 or C = 0 and the derivatives of form

wherein R -'- represents a C1-C6 alkyl group, A represents a saturated or unsaturated ring containing 3 to 7 carbon atoms; each of the symbols R ² , R ³ and R ⁴ independently represents a hydrogen or halogen atom or a group such as a C1-C6 alkoxy group, C1-C6 alkyl, C1-C6 alkylthio, C1-C6 alkoxycarbonyl, carboxyl, carboxamido, sulfonic acid, sulfonamido, acylamino, sulfonylamino, C1-C6 alkylsulfonate, nitrile, nitro, acyloxy, phenyl or methylene-dioxy and each of the symbols m and n represents an integer ranging from 0 to 4.

₂₂

in which R ¹ represents an alkyl, R ² a hydrogen or an alkyl. According to the invention, the term "prodrugs" means functional derivatives of the compounds of formula I which can be converted, preferably in vivo, in the patient according to the required form of formula I. Such prodrugs of these compounds can be obtained by methods well known to those skilled in the art such as those described in the document "Design and Prodrugs", Ed. H. Bundgaard, Elsevier (1985). The compounds of the invention relate to the two enantiomeric forms, the different separate isomers or a mixture of them.

The new compounds of the invention are advantageously used to obtain a protective effect on the protemic complexes of the internal mitochondrial membrane of the cells of a mammal, preferably a human. The compounds of the present invention find particular application in the prevention and / or treatment of partial or total ischemia, of pathologies associated with ischemia or of pathologies associated with mitochondrial deficiencies. The term “ischemia (partial or total), pathologies associated with ischemia and pathologies associated with mitochondπal deficiencies” means the diseases, preferably vascular, chosen from the group consisting of myocardial infarction, cerebral ischemia, chronic venous insufficiency, arteriopathies, i.e. lesions due to atherosclerosis affecting the arteries of patients, in particular those of the lower limbs, Raynaud's phenomenon linked to vasospasms leading to vasoconstriction of arteries, ulcers, gangrene, impaired capillary permeability, fragile capillaries, scarring, skin changes, retinal defects of ischemic origin, reduced hearing acuity of ischemic origin , the troubles associated with stays at high altitude, angina caused by short moments of coronary obstruction, pulmonary hypertension, hepatic ischemia, Parkmson's disease, myopathies and syndromes associated with vascular problems such as diabetes, where hypertension and impaired blood flow appear in the lower limbs. These diseases and pathologies linked to ischemia are well known to clinicians and physicians, who can adapt the use of the pharmaceutical composition for the treatment and / or prevention of symptoms and dysfunctions of the human or animal body associated with the abovementioned diseases and / or to prevent or decrease the possibility of having it. The inventors have unexpectedly discovered that these different syndromes or diseases can be treated with the compounds of the invention, their possible salts and prodrugs thereof, and that these compounds act according to the same mode of action. biochemical. Another aspect of the present invention relates to a pharmaceutical composition comprising a suitable pharmaceutical vehicle or excipient and a sufficient amount of one or more of the compounds of the invention, that is to say an amount sufficient for at least ameliorate or prevent the above symptoms in a mammal, especially a human. This sufficient amount may vary depending on certain factors such as the condition of the animal to be treated, the mode of administration, the severity of the side effects, the stability of the compound in the serum or the circulating blood, etc. . Preferably, the sufficient amount of compound used in the pharmaceutical composition of the invention is between 0.1 and 200 mg / patient, preferably between 1 and 50 mg / patient, preferably between 20 and 30 mg / patient, this quantity being able to be adapted in particular according to the administration doses required and the patient's weight, as can be extrapolated from the examples of application in vivo below

The pharmaceutical composition comprises a suitable pharmaceutical vehicle which can vary according to the mode of administration and can optionally be combined with an adjuvant so as to improve the therapeutic properties of the compound of the invention or to reduce its possible side effects. Such suitable pharmaceutical vehicles or adjuvants are well known to those skilled in the art and can be prepared according to the procedures generally applied by pharmacists and can include any non-toxic, solid (including powdered), liquid pharmaceutical vehicle.

(solutions, suspensions, emulsions, etc.) or gaseous. The percentage of active pharmaceutical compound (generally between 5 and 70% by weight) can vary depending on the frequency of administration and possible side effects on animals, including humans.

To prepare such pharmaceutical compositions in the form of tablets, granules, capsules or cachets, suspensions, etc., it is possible to incorporate elements such as corn starch, lactose, sucrose, sorbitol , talc, stearic acid, magnesium stearate, gums or diluents comprising a variable percentage of water or a solvent such as a syrup, oily or aqueous suspensions, perfumed emulsions, etc. Dispersible agents used in aqueous compositions can include gums, algae, dextrans, carboxymethyl cellulose derivatives, etc. The present invention also relates to a method of therapeutic or preventive treatment of ischemia or pathologies linked to ischemia as mentioned above comprising the administration of a sufficient amount of the compound of formula I or of the pharmaceutical composition of the invention to said patient, preferably a human, so as to prevent, reduce or eliminate the symptoms of ischemia, pathologies linked to ischemia or associated with mitochondrial deficiencies, or apoptosis, with the additional condition that the compound is not N- (2, 6-dιιsopropyl -phenyl) -2 -phenyl - malonamic acid dodecyl ester nor a compound of formula I in which R ¹ is CH2, R ⁴ is a group of formula

ZR ^ in which Z represents an oxygen and R ^ represents an alkyl and in which R ³ is an amme of formula II:

alkyl N O

S alkyl

II o

in which X represents O, S, CH2, C = 0.

A final aspect of the present invention relates to the use of these compounds or of the pharmaceutical composition of the invention for the preparation of a medicament intended for the treatment or prevention of ischemia, pathologies linked to the 'ischemia or associated with mitochondπale deficiencies as mentioned above or apoptosis with the additional condition mentioned in the previous paragraph. The present invention also relates to the process for the preparation of the compounds of the invention of formula I, comprising the following steps: - esterification of an intermediate compound R ³ -H by a group of formula

COCI

(

C0 ₂ ~ R ⁴

in the presence of dicyclohexyl carbodnmide (DCC),

- optionally followed by one or more cyclisacion reactions in the presence of an oxidizing agent such as manganese acetate Mn (III) or copper acetate

Cu (II) monohydrate and optionally sodium acetate.

The definitions of the radicals mentioned are those given in formula I for the compounds of the invention.

The various aspects of the present invention are described in detail in the nonlimiting examples presented below.

Examples

Example 1: Synthesis of dicyclopenten -2-malonate Synthesis of 2 -cyclopentenol

This synthesis is carried out according to the method described by A.L. Gemal and J.L. Luche (J. Am. Chem. Soc. 103, p. 5454 (1981)).

To a solution of 2 -cyclopentenone (5.00 g; 60.90 mmol) in methanol (commercial, 99%) is added cerium chloride heptahydrate (CeCl3.7H2θ) (22.69 g;

60.90 mmol) and the reaction mixture is stirred at room temperature for 40 minutes to homogenization. The mixture is then cooled to 0 ° C. and sodium borohydride (NaBH ^ (2.30 g; 60.90 mmol) is slowly added. After complete addition, the mixture is stirred at 0 ° C. for 3 hours. hydrochloric acid is then added to adjust the pH to 7. The phases are separated and the aqueous phase is extracted several times with ether (5 × 50 ml) The combined organic phases are dried over anhydrous MgSU4 and concentrated to give 2-cyclopentenol ( 3.81 g, 74%).

Synthesis of dιcyclopentèn-2 -yle malonate

To a solution of 2 -cyclopentenol (1.00 g;

11.89 mmol) in 50 ml of dichloromethane are added successively malonic acid (1.12 g; 10.81 mmol) and 4-dιméthylamιno pyπdme (DMAP)

(0.13 g; 1.08 mmol). The reaction mixture is stirred under an inert atmosphere and cooled to 0 ° C. At this temperature, dicyclohexyl carbodumide (DCC) (2.34 g; 11.35 mmol) is added in a single portion. A milky white mixture is obtained which is stirred at 0 ° C for 1 hour then at room temperature until complete reaction (control of the progress of the reaction by TLC, eluent Pentane / ethyl acetate: 8/2). The precipitate is removed by filtration on celite and washed with dichloromethane. The filtrate is washed with a saturated aqueous solution of NaHCC> 3 (10 ml). The phases are separated and the aqueous phase is extracted with dichloromethane

(3x10 ml). The combined organic phases are washed successively with a saturated aqueous NaCl solution and with water, then dried over anhydrous MgSC <4 and concentrated. The precipitate is again removed by filtration on celite and washed with pentane. The purification of the reaction crude is carried out by chromatography on a silica column (Pentane / ethyl acetate elution: 8/2) to give MRC2P119 (2.40 g; 94%) in the form of a colorless oil.

Characterization of the product ^λ U NMR (CDC1 ₃ , 400 MHz) δ: 1.81-2.55 (m, 8H), 3.32 (s,

2H), 5.76-5.84 (m, 4H), 6.12 (m, 2H); ¹³ C NMR (CDCI3,

175 MHz) δ: 29.05, 30.53, 41.36, 80.97, 128.33, 137.44,

165.92; IR (film): 3450, 3062, 2947, 2858, 1727, 1618, 1454, 1411, 1348, 1319, 1270, 1151, 1029, 973, 917, 878,

783, 735 cm ^-1 ; PM: 236.27; MS m / z 171 (M ⁺ - - 65); Calculated Elemental Analysis C (66.09%), H (6.82%); found C (66.09%), H (6.90%).

Example 2: Synthesis of 2 -propenyl 3 -oxa -2 -oxobicyclo [3. 1. 0] hexan - 1 - carboxylate

Diallyl malonate synthesis

To a solution of malonic acid (5.00 g; 48.05 mmol) in 330 ml of dichloromethane are added successively allyl alcohol (10.51 g; 124.93 mmol) and DMAP (1.17 g; 9.61 mmol). The reaction mixture is stirred under an inert atmosphere and cooled to

0 ° C. At this temperature, the DCC (20.82 g; 100.90 mmol) is added in a single portion. A milky white mixture is obtained which is stirred at 0 ° C for 1 hour then at room temperature until complete reaction (control of the progress of the reaction by TLC; eluent Pentane / ethyl acetate: 9/1). The precipitate is removed by filtration on celite and washed with dichloromethane. The filtrate is washed with a saturated aqueous solution of NaHCO ₃ (20 ml). The phases are separated and the aqueous phase is extracted with dichloromethane (3x20 ml). The combined organic phases are washed successively with a saturated aqueous NaCl solution and with water, then dried over anhydrous MgSC and concentrated. The precipitate is again filtered through celite and washed with pentane. The purification is carried out by chromatography on a silica column (elution Pentane / ethyl acetate: 9/1) to give the compound of the invention (MRC2PSS) (8.09 g; 92%) in the form of a colorless oil.

Product characterization

! H NMR (CDC1 ₃ , 400 MHz) δ: 3.44 (s, 2H), 4.64 (d, J = 5.9 Hz, 4H), 5.24 (d, J = 10.3 Hz, 2H), 5.34 (d, J = 17.1 Hz, 2H), 5.91 (ddt, J = 17.1-10.3 -5.9 Hz, 2H); ¹³ C NMR (CDCl ₃ ,

175 MHz) δ: 40.99, 65.57, 118.20, 131.26, 165.68; IR (film): 3653, 3089, 2989, 2951, 1739, 1650, 1450, 1414,

1368, 1330, 1274, 1150, 995, 935, 631 cm ^"1 ; PM: 184.19; MS m / z 184 (M + -); Calculated Elemental Analysis C (58.69%), H (6.57%); found C (58.66 %), H (6.57%).

Synthesis of 2-propenyl 3 -oxa-2 -oxobicyclo [3.1.0] hexane-1- carboxylate

To a solution of diallyl malonate (MRC2P55) (2.00 g; 10.86 mmol) in 55 ml of glacial acetic acid are added successively manganese acetate (III) dihydrate (Mn (OAc) 3.2H ₂ 0) (11.64 g;

43.43 mmol), Copper (II) acetate monohydrate (Cu (OAc) ₂ -H ₂ 0) (4.34 g; 21.72 mmol) and sodium acetate (AcONa) (1.78 g; 21.72 mmol). The brown solution obtained is heated at 80 ° C for 20 hours (this corresponds to the disappearance of the brown coloration and the appearance of a turquoise blue coloration). The progress of the reaction is monitored by TLC (eluent Pentane / Ethyl acetate: 9/1). The precipitate is removed by filtration on celite and washed with dichloromethane. The filtrate is washed with water (30 ml). The aqueous phase is extracted with dichloromethane (2x20 ml). The combined organic phases are washed successively with water (2x20 ml) and with a saturated aqueous solution of NaHCO3

(2x20 ml) then dried over anhydrous MgSO4 and concentrated. The crude reaction product is purified by chromatography on a silica column (Pentane / ethyl acetate elution: 1/1) to give the compound of the invention (MCR2PS7) (0.49 g; 25%) in the form of a colorless oil.

Characterization of the product -H NMR (CDC1 ₃ , 400 MHz) δ: 1.41 (dd, J = 5.4-4.9 Hz, 1H), 2.10 (dd, J = 8.0-4.6 Hz, 1H), 2.79 (m, 1H), 4.20 (d,

J = 9.3 Hz, 1H), 4.38 (dd, J = 9.3-4.9 Hz, 1H), 4.68 (d,

J = 5.4 Hz, 2H), 5.28 (dd, J = 10.2-1.5 Hz, 1H), 5.39 (dd,

J = 17.1-1.5 Hz, 1H), 5.94 (ddt, J = 17.1- 10.2 -5.4 Hz, 1H);

¹³ C NMR (CDCI3, 175 MHz) δ: 20.56, 27.82, 29.09, 65.97, 66.85, 118.60, 131.10, 166.14, 170.20; IR (film): 3653, 3094, 2989, 1777, 1725, 1650, 1447, 1393, 1314, 1271, 1183,

1115, 1090, 1047, 998, 939, 847, 793, 765, 703, 632 cm ^"1 ;

PM: 182.18; MS m / z 183 (MH ^{+ •} ); Calculated Elemental Analysis

C (59.34%), H (5.53%); found C (59.28%), H (5.68%).

EXAMPLE 3 Synthesis of Cyclopenten -2 -yle and Ethyl Malonate

Synthesis of ethyl malonic acid

Step 1 In a 250 ml tπcol flask, equipped with an addition funnel and a condenser, the diethyl malonate (20.00 g; 124.87 mmol) is dissolved in 80 ml ethanol (commercial). The colorless mixture obtained is stirred at room temperature and a KOH solution

(7.00 g; 124.76 mmol) in ethanol (80 ml) is slowly added (duration of the addition: 1 h). After complete addition, the mixture obtained is stirred at room temperature overnight (precipitation of potassium and ethyl malonate). The precipitation of the salt is advantageously accelerated by cooling the reaction flask in an ice bath. The potassium salt is recovered by filtration, washed with small amounts of ether and then dried under reduced pressure (17.77 g; 84%).

2nd step

In a 250 ml tπcol flask, equipped with a magnetic stirrer, an addition funnel and a thermometer, the potassium and ethyl malonate (17.77 g; 104.49 mmol) is dissolved in 18 ml of water. The reaction mixture is cooled via an ice bath and 8.2 ml of concentrated hydrochloric acid are slowly added. The reaction mixture is filtered and the KCl precipitate is washed with ether (3 × 50 ml). The filtrate is decanted and the aqueous phase is extracted with ether (3x50 ml). The combined ethereal phases are dried over anhydrous MgSO ₄ , filtered and concentrated to give ethyl malonic acid (13.45 g; 82% overall yield for the two stages) in the form of a colorless oil.

Synthesis of 2-cyclopenten-ethyl and ethyl malonate To a solution of 2-cyclopentenol (0.70 g; 8.32 mmol) in 30 ml of dichloromethane are successively added ethyl malonic acid (0.85 g; 6.40 mmol) and DMAP (0.08 g; 0.64 mmol). The reaction mixture is stirred under an inert atmosphere and cooled to 0 ° C. At this temperature, the DCC (1.39 g; 6.72 mmol) is added in a single portion. A milky white mixture is obtained which is stirred at 0 ° C for 1 hour then at room temperature until complete reaction (control of the progress of the reaction by TLC, eluent Pentane / ethyl acetate: 8/2). The precipitate is removed by filtration on celite and washed with dichloromethane. The filtrate is washed with a saturated aqueous solution of NaHCO3 (10 ml). The phases are separated and the aqueous phase is extracted with dichloromethane (3x10 ml). The combined organic phases are washed successively with a saturated aqueous NaCl solution and with water, then dried over anhydrous MgS04 and concentrated. The precipitate is again removed by filtration on celite and washed with pentane. The purification of the reaction crude is carried out by chromatography on a silica column (elution Pentane / Ethyl acetate: 8/2) to give the compound of the invention (MCR2P45) (1.39 g; 84%) in the form of an oil colorless.

Product characterization

1-H NMR (CDC1 ₃ , 400 MHz) δ: 1.28 (t, J = 7.3 Hz, 3H), 1.83

(m, 1H), 2.31 (m, 2H), 2.50 (m, 1H), 3.34 (s, 1H), 4.20 (q, J = 7.3 Hz, 2H), 5.76 (m, 1H), 5.83 (m, 1H), 6.13 (m, 1H);

¹³ C NMR (CDCI3, 175 MHz) δ: 13.41, 29.00, 30.47, 41.10, 60.64, 81.07, 128.46, 165.87, 165.94; IR (film): 3451, 2983, 1750, 1731, 1459, 1412, 1370, 1346, 1323, 1271, 1189,

1151, 1032, 971, 917, 879, 736 cm ^"1 ; PM: 198.22; MS m / z

170 (M ⁺ - - 28). Example 4 Synthesis of malona te of cyclopenten -2 -yle and methyl

Synthesis of methyl malonic acid Step 1 In a three-necked 100 ml flask equipped with an addition funnel and a condenser, dimethyl malonate (10.00 g; 75.69 mmol) is dissolved in 40 ml of methanol (commercial). The colorless mixture obtained is stirred at room temperature and a solution of KOH (4.25 g; 75.69 mmol) in methanol (40 ml) is slowly added (duration of the addition: 1 h). After complete addition, the mixture obtained is stirred at room temperature overnight (precipitation of potassium and methyl malonate). Salt precipitation can be accelerated by cooling the reaction flask with ice bam. The potassium salt is recovered by filtration, washed with small amounts of ether and then dried under reduced pressure (10.27 g; 87%). Step 2 In a 50 ml three-necked flask, equipped with a magnetic stirrer, an addition funnel and a thermometer, the potassium methyl malonate (10.27 g; 65.76 mmol) is dissolved in 10 ml of water. The reaction mixture is cooled using an ice-cold bath and 5.5 ml of concentrated hydrochloric acid are slowly added. The reaction mixture is filtered and the KCl precipitate is washed with ether (3x10 ml). The filtrate is decanted and the aqueous phase is extracted with ether (3x10 ml). The combined ethereal phases are dried over anhydrous MgSO4, filtered and concentrated to give methyl malonic acid (7.42 g; 96%) in the form of a colorless oil. Synthesis of 2-cyclopenten-methyl malonate and methyl [0046] To a solution of 2-cyclopentenol (1.00 g; 11.89 mmol) in 35 ml of dichloromethane are successively added methyl malonic acid (1.08 g; 9.15 mmol) and DMAP (0.11 g; 0.91 mmol). The reaction mixture is stirred under an inert atmosphere and cooled to 0 ° C. At this temperature, the DCC (1.98 g; 9.60 mmol) is added in a single portion. A milky white mixture is obtained which is stirred at 0 ° C for 1 hour then at room temperature until complete reaction (control of the progress of the reaction by TLC, eluent Pentane / ethyl acetate: 8/2). The precipitate is removed by filtration on celite and washed with dichloromethane. The filtrate is washed with a saturated aqueous solution of NaHCθ3 (10 ml). The phases are separated and the aqueous phase is extracted with dichloromethane (3x10 ml). The combined organic phases are washed successively with a saturated aqueous NaCl solution and with water, then dried over anhydrous MgSO4 and concentrated. The precipitate is again removed by filtration on celite and washed with pentane. The purification of the reaction crude is carried out by chromatography on a silica column (elution Pentane / ethyl acetate: 8/2) to give the compound of the invention (MCR2P85) (1.40 g; 83%) in the form of an oil colorless.

Characterization of the NMR product (CDC1 ₃ , 400 MHz) δ: 1.85 (m, 1H), 2.29 (m, 2H),

2.51 (m, 1H), 3.36 (s, 2H), 3.75 (s, 3H), 5.76 (m, 1H), 5.83 (m, 1H), 6.13 (m, 1H); ¹³ C NMR (CDCl ₃ , 175 MHz) δ:

29.27, 30.73, 41.16, 52.04, 81.43, 128.44, 137.83, 166.10, 166.74; IR (film): 2955, 2858, 1736, 1438, 1412, 1348, 1273, 1152, 1028, 968, 917, 878, 737 cm ^"1 ; PM: 184.19; MS m / z 156 (M ⁺ - - 28); Calculated Elemental Analysis C (58.69%), H (6.57%); found C (58.60%), H (6.59%).

Example 5 Synthesis of N, N '-diphenyl malonamide

To a solution of N-phenyl amme (0.75 ml; 8.22 mmol) in 25 ml of dichloromethane is added the triethylamme (0.72 ml; 5.14 mmol). The colorless mixture obtained is cooled to 0 ° C. by means of an ice-cold bath and the malonyl dichloride (0.5 ml; 5.14 mmol) is added slowly to the syringe. An intense red / orange solution accompanied by a precipitate is obtained. The reaction mixture is stirred at 0 ° C. for 2 hours (monitoring the progress of the reaction by TLC) (eluent Pentane / ethyl acetate: 1/9), then, approximately 4 ml of a HCl solution 10 % are added. The phases are separated and the organic phase is washed successively with water (10 ml) and with a saturated aqueous NaCl solution (10 ml). After drying over anhydrous MSO, filtration and concentration, an orange-colored solid is isolated. The purification is carried out by recrystallization from a Pentane / ethyl acetate mixture to provide the compound of the invention (MCR2P219) (0.33 g; 32%) in the form of an unbleached powder.

Characterization of the product ^λ E NMR (DMSO-dg, 400 MHz) δ: 3.47 (s, 2H), 7.06 (m, 2xlH),

7.31 (m, 2x2H), 7.60 (m, 2x2H), 10.17 (s, 2H); ¹³ C NMR

(DMSO-dg, 175 MHz) δ: 37.20, 110.30, 114.60, 120.00, 130.20, 156.60; IR (KBr patch): 3274, 3152, 3066, 1669, 1650, 1599, 1560, 1538, 1500, 1444, 1416, 1358, 1309, 1294, 1251, 1193, 1162, 979, 905, 857, 752, 693, 618 cm ^"1 ; mp = 220 ° C; MP: 254.29.

Example 6: Synthesis of N-phenylamido ethyl ethanoate Synthesis of ethyl chloroformyl ethanoate

In a three-necked flask of 50 ml, equipped with an addition funnel, a thermometer and a condenser, ethyl malonic acid (1.00 g; 7.57 mmol) is introduced into 8 ml of dry dichloromethane. The flask is cooled to 0 ° C by means of an ice bath. At this temperature, 1.5 equivalents (0.83 ml; 11.35 mmol) of thiony chloride diluted in 5 ml of dichloromethane are slowly added. After complete addition, the reaction mixture is stirred at 0 ° C for 10 minutes and then heated at reflux for 2 days. After concentration, ethyl chloroformyl ethanoate (0.99 g; 87%) is isolated, which is used without additional purification in the next step.

Product characterization

^X H NMR (CDC1 ₃ , 400 MHz) δ: 1.31 (t, J = 7.3 Hz, 3H), 3.87

(s, 2H), 4.26 (q, J = 7.3 Hz, 2H); IR (film): 2989, 1802, 1742, 1577, 1468, 1401, 1372, 1323, 1260, 1164, 1035, 982,

940, 859, 652, 612 cm ^"1 ; PM: 150.56.

Synthesis of ethyl N-phenylamido ethanoate To a solution of amino N-phenyl (0.25 g; 0.24 ml; 2.66 mmol) in 15 ml of dry dichloromethane are added 1.25 equivalents (0.46 ml; 3.32 mmol) of triethylamme. The solution obtained is placed at 0 ° C via an ice bath. At this temperature, 1.25 equivalents (0.43 ml; 3.32 mmol) of chloroformyl ethyl ethanoate are slowly added using a syringe. An intense orange solution is obtained which is stirred at 0 ° C. overnight (monitoring the progress of the reaction by TLC) (eluent Pentane / ethyl acetate: 1/9), then, approximately 4 ml of a 10% HCl solution are added. The two phases are separated and the organic phase is washed with water. The combined aqueous phases are extracted with dichloromethane. The organic phases are washed with a saturated aqueous NaCl solution, dried over anhydrous MgSO4, filtered and concentrated. The purification is carried out by chromatography on a silica column (eluent Pentane / ethyl acetate: 1/9) to quantitatively give the compound of the invention

(MCR2P231) (0.55g; 100%;

Characterization of the product 1 H NMR (CDC1 ₃ , 400 MHz) δ: 1.32 (t, J = 7.3 Hz, 3H), 3.47 (s, 2H), 4.26 (q, J = 7.3 Hz, 2H), 7.12 (t, J = 7.3 Hz, 1H), 7.33 (t, J = 7.8 Hz, 2H), 7.55 (d, J = 7.8 Hz, 2H); ¹³ C NMR (CDCI3, 175 MHz) δ: 13.67, 42.26, 61.38, 119.98, 124.25,

128.55, 137.37, 163.77, 168.70; IR (film): 3313, 3143, 2985, 1738, 1668, 1603, 1549, 1500, 1445, 1372, 1342, 1244,

1190, 1156, 1033, 906, 844, 757, 695 cm ^"1 ; PM: 207.23; MS m / z 207 (M ⁺ -).

Example 7 Synthesis of Ethyl 2,6-dimethyl -N-phenylamido ethanoate

To a solution of 2,6-dιméthyl-N-phenyl amme

(1.31 ml; 10.63 mmol) in 60 ml of dry dichloromethane is added the triethylamme (1.85 ml; 13.28 mmol).

The colorless mixture obtained is cooled to 0 ° C. via an ice bam and added slowly syringe with ethyl chloroformyl ethanoate (1.3 ml; 10.15 mmol). An intense red / orange solution is obtained which is stirred at 0 ° C for 1 night (monitoring the progress of the reaction by TLC) (eluent Pentane / Ethyl acetate: 1/9), then, approximately 4 ml of a 10% HCl solution are added. The two phases are separated and the organic phase is washed with water (3x10 ml). The combined aqueous phases are extracted with dichloromethane. The organic phase is washed with a saturated aqueous solution of NaCl and then dried over anhydrous MgSO4. After filtration and concentration, the crude ester amide is obtained in the form of an orange solid. The purification is carried out by recrystallization from a Pentane / ethyl acetate mixture to provide the compound of the invention (MCR2P255) (1.67 g; 67%) in the form of an unbleached powder.

Product characterization

^X H NMR (CDC1 ₃ , 400 MHz) δ: 1.35 (t, J = 7.3 Hz, 3H), 2.20 (s, 6H), 3.50 (s, 2H), 4.25 (q, J = 7.3 Hz, 2H), 7.10 (m,

3H), 8.55 (extended s, 1H); ¹³ C NMR (CDCl ₃ , 175 MHz) δ:

13.98, 18.29, 41.20, 61.67, 127.23, 128.06, 133.50, 135.10, 163.38, 169.66; IR (KBr patch): 3226, 3031, 1731, 1666, 1642, 1595, 1536, 1477, 1411, 1397, 1370, 1351, 1264, 1218, 1204, 1166, 1030, 986, 880, 761, 723, 635 cm ^"1 ; mp =

103 ° C; PM: 235.28; MS m / z 235 (M + ^• ).

Example 8 Synthesis of N-methyl -N-cyclohexamido ethanoate of ethyl e To a solution of N-methyl N-cyclohexylamme (0.41 ml; 3.12 mmol) in 20 ml of dry dichloromethane is added the triethylamme (0.54 ml; 3.91 mmol). The colorless mixture obtained is cooled to 0 ° C. by means of an ice bam and ethyl chloroformyl ethanoate (0.5 ml; 3.91 mmol) is slowly added to it with a syringe. An intense red / orange solution is obtained which is stirred at 0 ° C for 4 hours (control of the reaction progress by TLC) (eluent Pentane / Ethyl acetate: 1/9), then, approximately 4 ml of a 10% HCl solution are added. The two phases are separated and the organic phase is washed with water (3x10 ml). The combined aqueous phases are extracted with dichloromethane. The organic phase is washed with a saturated aqueous solution of NaCl and then dried over anhydrous MgSO4. After filtration and concentration, the crude ester amide is obtained. The purification is carried out by chromatography on a silica column (eluent Pentane / ethyl acetate: 1/9) to provide the compound of the invention (MCR2P237) (0.71 g; 100%) in the form of an orange liquid.

Characterization of the product obtained 1 H NMR (CDC1 ₃ , 400 MHz) δ: 1.27-1.67 (m, 14H), 2.84 (s,

3H), 3.44 (s, 1H), 3.47 (s, 1H), 4.21 (q, J = 7.3 Hz, 2H),

4.40 (m, enol form contribution); ¹³ C NMR (CDC1 ₃ , 175 MHz) δ: 14.52, 24.99, 29.05, 30.14, 41.44, 52.09, 60.48, 65.10, 165.13, 167.06; IR (film): 2933, 2858, 1741, 1645, 1477,

1450, 1408, 1370, 1326, 1254, 1164, 1098, 1035, 947, 895,

847, 788, 674 cm ^"1 ; PM: 227.30. Example 9 Synthesis of 2-dicyclopenten-malona te, S, S enantiomer

Synthesis of 2 -Bromo-2 -cyclopentenone

In a three-necked 250 ml flask, equipped with an addition funnel and a condenser, 2 - cyclopentenone (7.24 g; 88.2 mmol) is dissolved in 60 ml of CCI4.

The solution is cooled to 0 ° C and a Br2 solution

(15.51 g; 97 mmol) in CCI ₄ is added dropwise. After complete addition, a solution of N-Et3 (13.39 g; 132 mmol) in CC1 ₄ (60 ml) is added at the same temperature dropwise. The mixture obtained is stirred at room temperature for 2 hours, then filtered. The filtrate is washed successively with 1M HCl, 10% NaHCO 3 and H2O. The organic phase is dried over anhydrous MgSO ₄ , filtered and concentrated. The solid thus obtained is re-installed (cyclohexane / Et2θ) to give 2-bromo-

2 -cyclopentenone (7.5 g; 53%) in the form of a white solid.

Synthesis of (±) -2-Bromo-2 -cyclopentenol To a solution of 2 -bromo-2 -cyclopentenone (8 g; 49.7 mmol) in methanol (150 ml) is added cerium chloride heptahydrate (CeCl3 .7H2θ) (20.36 g;

54.7 mmol) and the reaction mixture is stirred at room temperature for 40 minutes. The mixture is then cooled to 0 ° C. and sodium borohydride (NaBH ₄ )

(2.07 g; 54.7 mmol) is slowly added. After complete addition, the mixture is stirred at 0 ° C for 2 hours. An aqueous solution of NH ₄ CI is then added. The phases are separated and the aqueous phase is extracted several times with ether (5 x 50 ml). The combined organic phases are dried over anhydrous MgSO ₄ and concentrated. The crude product thus obtained is purified by chromatography on silica (Pentane / ethyl acetate elution: 9/1) to give the (±) -2 -bromo-2 -cyclopentenol (6.0 g, - 74%) in the form of a yellow oil.

Characterization of the product

¹ H NMR (CDC1 ₃ , 400 MHz) δ: 1.85-2.04 (m, 2H), 2.20-2.60 (m, 3H), 4.71 (b, 1H), 6.05-6.08 (m, 1H).

Synthesis of (S) -2 -Bromo-2 -cyclopentenol To a solution of (±) -2 -oromo-2 -cyclopentenol (6.00g; 36.8 mmol) in a mixture of isopropenylacetate (24 ml) and cyclohexane (96 ml) is added the enzyme Lipase Novozym 435 (1.5 g). The mixture is stirred for 1 hour at room temperature and then filtered. The filtrate is evaporated and the residue thus obtained is purified by chromatography on silica (elution cyclohexane / AcOEt: 9/1) to give the (S) -2-bromo-2-cyclopentenol (1.5 g; 50%) as a solid White.

Product characterization

! H NMR (CDCI3, 400 MHz) δ: 1.85-2.04 (m, 2H), 2.20-2.60 (m, 3H), 4.71 (b, 1H), 6.05-6.08 (m, 1H). [α] ²² _D = -47.5 ° (c = 1.0, CHCI3).

Synthesis of (S) -2 -Cyclopentenol

In a 50 ml three-necked flask, equipped with an addition funnel, a thermometer and a refrigerant, tBuLi (1.7 M pentane sol; 23 ml; 39.1 mmol) and Et2θ are introduced. (10 ml). The mixture is then cooled to -78 ° C and (S) -2-bromo-2-cyclopentenol (1.90 g; 11.7 mmol) dissolved in Et2θ (15 ml) is slowly added. After complete addition, the mixture is left heat to 0 ° C and stirred at this temperature for 2 hours. A dilute solution of NH4CI in water is then added slowly. The organic phase is separated and the aqueous phase extracted several times with Et2θ. The combined organic phases are dried over anhydrous MgS0 and concentrated to give the (S) -2 -cyclopentenol in the form of an oil (600 g, - 61%).

Characterization of the product ^X H NMR (CDCI3, 400 MHz) δ: 1.65-1.73 (m, 2H), 2.22-2.32

(m, 2H), 2.48-2.56 (m, 1H), 4.87 (b, 1H), 5.81-5.89 (m, 1H), 5.90-6.10 (m, 1H).

[] ²² _D = -119.2 ° (c = 1.0, CHCI3)

Synthesis of di- (S) -2-Cyclopenten-2-yl malonate

The synthesis of dι- (S) -2- Cyclopentè-2 -yle malonate is carried out according to the same protocol as that described in Example 1.

Characterization of the H NMR product (CDCI3, 400 MHz) δ: 1.80-1.92 (m, 2H), 2.20-2.40

(m, 4H), 2.45-2.60 (m, 2H), 3.34 (s, 2H), 5.75-7.79 (m, 2H), 5.83-5.86 (m, 2H), 6.10-6.17 (m, 2H).

[α] ^{2 2} _D = 212. 8 ° (c = 0.97, CHCl ₃ )

Example 10: Tests in vi tro

Effect of the Products at the Level of M tochondrial Respiration (Effect at Stages III and IV and at the Level of RCR) The compounds of the invention are characterized by the inhibition of the decrease in ATP content, of the activation of phospholipase A2 and adhesion of polymorphonuclear neutrophils (PMN) on human umbilical vein endothelial cells in culture when these are incubated under hypoxic conditions. The inhibition by the compounds of the invention of the activation cascade of endothelial cells induced by hypoxia and the maintenance of the ATP content in endothelial cells under hypoxia are obtained by the compounds of the invention which preserve mitochondπale respiratory activity. This is confirmed by measuring the respiratory activity expressed by the respiratory control (RCR) of mitochondria of liver of rats treated per os. The mitochondria are isolated according to the method described by Nowicki et al. , J. of Cerebral Blood Flow and Metabolism 2, pp. 33-40 (1982). Mitochondrial respiration is measured by an oxygen electrode linked to a recorder. The RCR represents respiratory control, and represents the relationship between oxygen consumption in the presence of endogenous substrate (glutamate / malate) and consumption after phosphorylation of ADP to ATP. This technique has been described by Chance & William (Na ture 175, pp. 1120-1121 (1955)).

Different compounds of the invention have been tested at different concentrations on the respiratory activity of the purified mitochondπes and obtained from rat livers. The mitochondria were preincubated in the presence of the compounds so as to measure the slope of stages III and IV of respiration as described in the document WO98 / 51291. RCR was then calculated.

The results below show that the products of Example 1 and of Example 2 exert an action on the RCR with a maximum effect at 10 ^{~ 7} mol / 1 for the product of Example 2. This increase in CPR is mainly due to a decrease in stage IV breathing.

The compound of Example 1 significantly increases the RCR of mitochondπes, and this because this molecule greatly decreases stage IV of respiration. The effect is optimal at 10 ^" ^ mol / 1 and decreases with concentration to be negligible at 10 ^" ^ mol / 1. The table below summarizes the effect of other compounds of the invention with the optimal concentrations at which these effects have been observed.

Table I

(*): in% compared to control increased to 100%

The results show that the compounds of the invention are capable of reproducibly protecting the decrease in ATP content induced by hypoxia in endothelial cells. The maximum average protection of 92.5% is observed at a concentration by 10 ^-4 mol / 1 for the compound of Example 2. This protection decreases with the concentration of this compound to be negligible at 10 ^-8 mol / 1. The results also show that the compound of Example 1 is capable of reproducibly protecting the decrease in ATP content induced by hypoxia in endothelial cells. The average maximum protection of 92.1% is observed at concentrations of 10 ^-4 to 10- ^ mol / 1. This protection decreases with the concentration of this compound to be negligible at 10 ^-8 mol / 1.

This protection is reproducible insofar as the same changes in the curve are always found for all the experiments, and even if there is variability between experiments in terms of the effect of hypoxia on the content in ATP. This variability is inherent in the experimental model. Indeed, for each experiment, a different culture of cells isolated from a different umbilical cord is used, and there is a variability of behavior of the cells from one cord to another.

Example 11 Stability in Human Plasma The compounds of Examples 1 and 2 were subjected to stability tests in order to evaluate their behavior in contact with the plasma after an incubation of 3 and 24 hours at a temperature of 37 ° C. These tests were carried out by GC following the evolution of the chromatograms of the different esters compared to an internal standard as a function of time. The injection of the pure products in GC / MF was carried out under the following conditions:

- injection temperature: 250 ° C,

- initial temperature: 40 ° C, - final temperature: 250 ° C,

- temperature gradient: 10 ° C / mm,

- injection method: split,

- retention time: compound of Example 1: 13.39 compound of Example 2: 11.23

The different compounds were prepared as follows. 10 μl of pure compound were dissolved in 100 μl of plasma at a temperature of 37 ° C.

The solution obtained is stirred. After 3 hours or 24 hours, 5 μl of internal standard (N-benzylam) and 1 ml of ether are added to the solution maintained at

37 ° C. The sample is centrifuged for 10 mm at a speed of 13,000 rpm. 1 μl of the supernatant phase

(ethereal phase) is injected in GC. The sample is then analyzed by GC. Analysis of the chromatograms makes it possible to demonstrate that after 3 hours in contact with human plasma maintained at 37 ° C., the peak corresponding to the compounds of the invention is still observed. The compounds of the invention therefore undergo little or no significant degradation after 3 hours in the presence of plasma. No other peak which may correspond to metabolic products is observed. After 24 hours in contact with human plasma, the presence of the peak corresponding to the compound of Example 1 is still observed. However, the peaks corresponding to the compound of Example 2 are no longer found.

Example 12: In vi tro effect on the adhesion of neutrophils to endothelial cells incubated under hypoxia

Endothelial cells were incubated under hypoxia in the presence of different concentrations of the compounds of the invention. After incubation, the cells are co-incubated with a suspension of unstimulated human neutrophils. Under normoxic conditions, the adhesion of neutrophils to endothelial cells is low (5 to 10%), but it increases on the order of 3 times under hypoxia.

The effect of the compound of Example 1 on the adhesion of neutrophils to endothelial cells incubated under hypoxia is characterized by an inhibition of the adhesion induced by hypoxia, and this in a manner dependent on the concentration between 10 ^" ^ and 10 ^-8 mol / 1. An optimum activity should be obtained between 10 ^{~ 7} and 10 ^" 5 mol / 1. Similarly, the compound of Example 2 inhibits the increase in the adhesion of neutrophils induced by hypoxia between 10 ^" ^ and 10 ^-8 mol / 1 with a maximum of 71% at 10 ^-8 mol / 1 .

EXAMPLE 13 In Vivo Effect on the Liver After Treatment of Rats Per Orally for 5 Days The rats were treated per os either with solutions of increasing concentration of each of the compounds of the invention, or with batches containing DMSO at the same concentration as that used to dissolve the molecules (control rats) once a day for 5 days. Following the treatment, the rats are sacrificed. The mitochondπes are isolated from the liver and their respiratory activity is measured every 15 mm during the 75 mm following the isolation procedure.

The optimal pre-mcubation concentration for the compound of Example 1 is 10 ^~ mol / 1 (0.236 μg / 1). The dose used to treat rats is therefore 2.36 mg / kg, and doses 5 times less, 5 times higher and 20 times higher were also tested.

For the compound of Example 2, the optimal concentration in pre-mcubation is 10 ^" 'mol / 1 (0.0183 μg / ml). The doses tested are therefore 0.18 mg / kg Given that this dose was weakly active, doses 5 times higher and 20 times higher were also tested. The compound of example 1 increases in a dose-dependent and significant manner the RCR of the hepatic mitochondπes after a treatment of 5 days with a 25% increase to 2.36 mg / kg At a higher dose, the molecule becomes slightly toxic and the RCR drops slightly below the control values. This increase in RCR is due to a marked decrease in stage IV of breathing.

The compound of Example 2 increases dose-dependent and significantly the RCR of the mitochondria with a maximum of 17% at 3.6 mg / kg. This increase in CPR is also due to a decrease in stage IV breathing.

EXAMPLE 14 In Vivo Effect on the Heart After Treatment of Rats Per Orally for One Day The rats were treated per os with the concentrations of 2.36 mg / kg of compound of Example 1 or 3, 6 mg / kg of compound of Example 2, either with batches containing DMSO at the same concentration as that used to dissolve the molecules (control rats) at a rate of once.

The next day, the rats are sacrificed, and the mitochondπes are isolated from the heart. Their activity Respiratory is measured every 15 minutes for the 75 minutes following the isolation procedure. The compound of Example 1 increases the RCR of the cardiac mitochondria after a day's treatment with an increase of 41% at 2.36 mg / kg The compound of Example 2 increases the RCR of the mitochondria after a one day treatment with an increase of 56% to 3.6 mg / kg.

EXAMPLE 15 In Vitro Application for the Treatment of Apoptosis

Promyelocytic cells are incubated in the presence of agents inducing apoptosis such as etoposide, a type II topoisomerase inhibitor. After incubation, various parameters indicative of apoptosis are detectable: activation of caspases, release by the mitochondπes of cytochrome c, DNA fragmentation, etc. Unexpectedly, if the cells are incubated in the presence of etoposide and in the presence of the compound of Example 1, an inhibition of the induction of apoptosis is observed. This inhibition is dependent on the dose between 10 ^-5 and 10 ^"8 mol / 1. Thus, the activation of caspase 3, the release by the mitochondria of cytochrome c and the fragmentation of DNA are inhibited.

Claims

1. Compound of formula (I), a salt or a prodrug thereof corresponding to formula I:

(D ⁿ

0 0

in which :

- R ¹ is CH ₂ , NH, or a ligand, preferably a carbon atom, forming with R ³ and / or R ⁴ an aromatic or non-aromatic ring of 5 or 6 carbon atoms, optionally comprising on each ring a heteroatom , preferably a nitrogen, oxygen or sulfur atom;

- R ³ and / or R ⁴ represent: - an amme of formula -NR ^, or -a group of formula -ZR ^ in which:

- Z represents O or S,

- R ⁵ represents H or an alkyl of 1 to 6 carbon atoms (preferably, R ^ represents a methyl or an ethyl), - R6 represents a hydrophobic group, preferably a terbutyl, an allyl or an aromatic or non-aromatic ring of 5, 6 or 7 carbon atoms optionally comprising one or more heteroatoms and in which the carbon atoms are optionally substituted by:

- an alkyl group R ⁷ of 1 to 6 carbon atoms (preferably methyl or ethyl), - a ketone group,

- a hydroxyl group,

- an ester group,

- a halogen element (F, Cl, Br or I), i - a trifluoromethyl (CF ₃ ), or

- a grouping of formula

- in which :

- W represents CH or a nitrogen atom,

- Y represents S or O, and

- R ⁸ represents an alkyl group of 1 to 6 carbon atoms, preferably methyl or ethyl;

-NR ⁵ or the group of formula -ZR ^ R ⁶ mentioned above, R ⁴ represents an alkyl of 1 to 10 carbon atoms, saturated or unsaturated, optionally comprising one or more heteroatoms.

2. Compound according to claim 1, characterized in that R ⁵ , R ⁷ and R ⁸ represent H or methyl, and in that R ¹ is CH2.

3. Compound according to claim 1 or 2, characterized in that R ¹ is CH2 and in that R ³ and R ⁴ represent an amine of formula -NR ^ or a group R5 of formula -OR ^ in which R ^ represents a tert-butyl, a cyclopentene, a cyclohexane or a benzene whose carbon atoms are optionally substituted by a methyl group, a ketone group, a hydroxyl group, an ester group, a halogen element or a tπfluoromethyl group (CF3).

4. Compound according to any one of the preceding claims, characterized in that it is chosen from the group consisting of dιcyclopentèn-2-yle malonate, 2-propenyl 3 -oxa-2 -oxobicyclo

[3.1.0] hexane-1-carboxylate, cyclopenten-2-yl and ethyl malonate, di- (S) -cyclopentene-2 -yl malonate, cyclopenten-2-methyl and methyl malonate , N, N'- diphenyl malonamide, N-phenylamido ethyl ethanoate, 2, 6-methyl-N-phenylamido ethyl ethanoate and N- methyl -N-cyclohexylamido ethyl ethanoate, optionally their salts or prodrugs thereof.

5. Pharmaceutical composition comprising a suitable pharmaceutical vehicle and one or more compound (s) according to any one of the preceding claims.

6. Use of the compound according to any one of claims 1 to 4 or of the pharmaceutical composition according to claim 5 for the preparation of a medicament intended for the treatment and / or prevention of ischemia, pathologies linked to the 'ischemia or pathologies linked to mitochondπal deficiencies.

7. Process for the preparation of the compound according to claim 1, comprising the following steps: esterification of an intermediate compound of formula R -H with a group

COCI

C0 ₂ -R ⁴ in the presence of dicyclohexyl carbodiimide (DCC), optionally followed by one or more cyclization reactions in the presence of an oxidizing agent.