WO2009083929A1 - Caspase-1 inhibitors and uses thereof - Google Patents

Abstract

The invention is selective new caspase-1 inhibitors that recognize caspase-1 and block its activity based on SEQ1.

Description

CASP ASE l INHIBITORS AND USES THEREOF

The invention is in the field of medicinal biology and chemistry and relates to novel derivatives having inhibitory properties with respect to caspase-1 and their biological applications particularly to treat diseases and injuries where caspase-1 activity is implicated.

Researches of the inventors have lead them to develop new derivatives having caspase-1 inhitory properties of great interest.

The invention thus relates to new caspase-1 inhibitors.

It also relates to the use of said derivatives as active principles of drugs.

The derivatives of the invention have both peptide and organic entities, wherein the peptide entity is a tetrapeptide YVAD of sequence SEQ ID N° 1 ((S)-Υyr-(S)-Val-(S)-Aia.-(R,S)-Asp), the Asp residue being optionally substituted by O-methyl group (OMe) and has terminal organic end units respectively substituted by quinolinylcarbonyl and 2,6-difluorophenyl ester backbones.

More particularly ,the invention relates to a derivative of formula (I) 5-(2,6-Difluoro-phenoxy)-3(R,S)-[2(S)-(2(S)-{3-(4-hydroxy-phenyl)-2(S)-[(quinoline-2- carbonyl)-amino] -propionylamino } -3 -methyl-butyrylamino)-propionylamino] -4-oxo- pentanoic acid methyl ester, or Quinoline-2-carbonyl-fS>Tyr-fS>Val-fS>Ala-^^-Asp(OMe)-CH2OC6H3(2,6-F₂).

In a preferred embodiment, the above derivatives are under salt form such as chlorhydrate. I

The invention also relates to derivatives of formula (II) derived from those of formula (I),

P4 P3 P2 Pl

(H) wherein (i) the absolute configuration of each amino acid is either L or D ;

(ϋ) R², R³ ,R⁴ and R⁵ are natural or non natural amino acid side chains; (iii) P4, P3, P2 and Pl, are preferentially and respectively, Tyrosine, Valine, Alanine and Aspartate;the aspartate moiety being optionally methylated;

(iv) R¹ and R² are a hydrogen atom, deuterium atom, Ci_2o aliphatic, substituted or unsubstituted aryl, cycloalkyl, naphthyl, substituted naphthyl, 2-benzoxazolyl, substituted 2-oxazolyl, (CH₂)_ncycloalkyl, (CH₂)_nphenyl, (CH₂)_nsubstituted phenyl,

(CH₂)_n(l- or 2- naphthyl), (CH₂)_nheteroaryl, CH₂N₂, CH₂Y, OH, OR, NH₂, NHR,

NR₂, SR, COR, CO₂R, CONR₂, CH₂OCOR, CH₂O-CO aryl, CH₂O-C(O) substituted aryl (ex: 2,6-dimethylbenzoyloxymethyl), CH₂O-C(O) substituted aryl, CH₂O-C(O) heteroaryl, CH₂O-C(O) substituted heteroaryl or CH₂OPOR₂ ;

(v) R³, R⁴, R⁵, R⁶ and R⁷ are Ci_₂o aliphatic, cycloalkyl, naphthyl, substituted naphthyl,

2-benzoxazolyl, substituted 2-oxazolyl, (CH₂)_ncycloalkyl, (CH₂)_nphenyl,

(CH₂)_nsubstituted phenyl, (CH₂)_n(l- or 2- naphthyl), (CH₂)_nheteroaryl, CH₂N₂,

(CH₂)_nZ, CH₂OCOR, CH₂OCO (aryl), CH₂OCO (substituted aryl), CH₂OCO (heteroaryl), CH₂OCO (substituted heteroaryl) or CH₂OPOR₂ ;

(vi) R⁶ and R⁷ are malonyl, methylmalonyl, 2-quinolinylcarbonyl, succinyl, methylsuccinyl, acetyl, 2-quinolinylmalonyl group;

(vii) R⁶ is a hydrogen atom and R⁷ is R, U, C0(CH₂)_nNH(U), CO(CH₂)_nS(U);

(viii) U is unsubstituted or substituted (2-, 3-, 4-, 5-, 6-, 7- or 8-) quinolinyl, Ci_₂o straight chain or branched alkyl, (CH₂)_ncycloalkyl, (CH₂)_nphenyl, (CH2)_nsubstituted phenyl, (CH₂)I₁(I- or 2- naphthyl), (CH2)_nheteroaryl, biotin, dinitrophenyl (DNP), iodoacetamides, DTNB, COR (ex. 2-quinolinylcarbonyl), COOR, C0(CH₂)_nNH(Z), Acridine derivatives (Red, yellow, orange ....), Fluorescein derivatives (fluorescein, FITC, FAM (carboxyfluorescein), 5-(and-6)- carboxynaphtho fluorescein, carboxyfluorescein, BCECF, naptofluorescein ),

Oregon Green® (2',7'-difluorofluorescein) dyes (Oregon Green® 488, Oregon Green® 514...), BODIPY® (4,4-difiuoro-5,7-dimethyl-4-bora-3a,4a- diaza-s- indacene-3 -propionic acid) dyes (BODIPY FL, BODIPY TMR, BODIPY TR, BODIPY 630/650, BODIPY 630/665 ), Bimane, Coumarin derivatives (amino methylcoumarin (AMC), AMCA, aminocoumarin, diethylamino coumarin hydroxymethylcoumarin; hydroxycoumarin, methoxycoumarin, AFC, ...), Cyanin derivatives (phycocyanin, allophycocyanin (APC), CY3.18, CY5.18, Cy2, Cy3, Cy3.5, Cy5, Cy5.5, Cy7...), Erythrin/Phycoerythrin derivatives (R-Phycoerythrin (PE), B-Phycoerythrin ...), APC/PE-Cy conjugates (PE-Cy5 conjugates, PE-Cy7 conjugates, APC-Cy7 conjugates....), Calcein derivatives (calcein, SNAFL calcein....), DANS, DANSA, Cascade Blue, Lucifer yellow, NBD, Red 613, FluorX, Rhodamine derivatives(Rhodamine 123, Rhodamine 110, Rhodamine B, Rhodamine 6A, Rhodamine 6G, TRITC, X-Rhodamine, sulphorhodamin, Rhodamine Red-X, Lissamine™ rhodamine B, DHR, Rhodamine Green....), PerCP, Texas Red, TruRed, Alexa Fluo ® (Alexa Fluor 350, Alexa Fluor 430,

Alexa Fluor 488, Alexa Fluor 532, Alexa Fluor 546, Alexa Fluor 555, Alexa Fluor 568, Alexa Fluor 594, Alexa Fluor 633, Alexa Fluor 647, Alexa Fluor 660, Alexa Fluor 680, Alexa Fluor 700, Alexa Fluor 750....), Q-DOTs ™ derivatives (655,605, 585,525,...), SNARF, Zenon™ derivatives (Zenon™ Alexa Fluor® 350, Zenon™ Alexa Fluor® 488, Zenon™ Alexa Fluor® 555, Zenon™ Alexa

Fluor® 594, Zenon™ Alexa Fluor® 647, Zenon™ Allophycocyanin, Zenon™

Biotin-XX, Zenon™ R-Phycoerythrin, ); NBD, Texas Red ®, QS Y® dyes

(QSY® 7, QSY® 9, QSY® 35, QSY® 21), Hoechst (33342, 33258), DAPI, Chromomycin A3, Mithramycin, , SYTOX (Blue, Green, Orange), Ethdium, Ethidium Bromide, 7-AAD, TOTO dyes, YOYO dyes, TO-PRO dyes, BOBO dyes, JO-PRO dyes, LO-PRO dyes, PO-PRO dyes, YO-PRO dyes, Thiazole Orange, Propidium Iodide (PI), LDS 751, Indo® dyes (Indo-1...), Fluo® dyes (Fluo-3...), DCFH, pNA, SYBR green II, SyPro (Orange, Ruby), EDANS, IR800, DiI, DiO, DiD, SNARF® derivatives, Fura dyes, QUIN dyes, NANOGOLD particules, NANOGOLD maleimide, AlexaFluor FluoroNANOGOLD, AlexaFluor FluoroNANOGOLD streptavidin, malachite green, Dabcyl, Dabsyl, Cascade yellow, dansyl, Dapoxyl, PyMPO, Pyrene, benzoxadiazole derivatives, strepavidin-/neutravidin-)biotin-labeled fluorescent microspheres, CMNB-caged fluorescein conjugate of streptavidin, calcofluor white, nile red, Y66F, Y66H,

EBFP, GFP wild type, QFP mutants H9/P4/P9/P11/W, GFPuv, ECFP, Y66W, S65A, S65C, S65L, S65T, EGFP, EYFP, ECFP, DsRedl, DsRed2, NANOGOLD^® particules, streptavidin-Nanogold^®, Monomaleido Nanogold®, Mono-Sulfo-NHS- Nanogold®, Monoamino Nanogold®, positively/negatively charged Nanogold® (NN, NHSN, NHSNA, NHSNS), Non-Functionalized Nanogold®, Monomaleido

Nanogold®, Mono-Sulfo-NHS-Nanogold®, Monoamino Nanogold®, Nonfunctional Nanogold®, Nanogold®-conjugates, Nanogold®-streptavidin, lipide- Nanogold (Palmitoyl Nanogold®, DPPE Nanogold®, Palmitoyl Undecagold, DPPE Undecagold), Ni-NTA-Nanogold®, Alexa Fluor® 488 FluoroNanogold, Alexa Fluor® 594 FluoroNanogold, Fluorescein FluoroNanogold, HRP substrate-

Nanogold,;

(ix) R⁷ and R⁶ are also taken together with the intervening nitrogen to form a heterocyclic ring such as substituted or unsubstituted tetrahydroquinoline, tetrahydroisoquinoline, dihydroacridine, benzazepine, pyrrolidine, morpholine, thiomorpholine, piperazine, piperidine, dihydropyridine, benzimidazole, imidazole, imidazoline, pyrrole, pyrrolidine, pyrroline, pyrazole, pyrazoline, pyrazolidine, triazole, piperidine, morpholine, thiomorpholine, piperazine, carbazole, phenothiazine, phenoxazine, dihydrophenazine, dihydrocinnoline, dihydroquinoxaline, dihydronaphthyridine, tetrahydronaphthyridine, dihydroacridine, indole, isoindole, dihydroindole, indoline, indazole, purine, 9,10- dihydrophenanthridine, 5H-dibenzo[b,fJazepine, 10,l l-dihydro-5H- dibenzo[b,f]azepine, β-carboline, pyrido[4,3-b]indole, 2,3,9-triazofluorene, 9-thia-

2,10-diazaanthracene, thieno[3,2b]pyrrole, dihydrophenanthrine;

(x) R is a hydrogen atom, Ci_2o aliphatic group, aryl, substituted aryl (ex: A- nitrophenyl or coumarine derivatives), hetetoaryl (ex. 2-pyridine), substituted heteroaryl, cycloalkyl, naphthyl, substituted naphthyl, (CH₂)_ncycloalkyl, (CH2)_nphenyl, (CH2)_nsubstituted phenyl (ex: 2,6-dihalophenyl), (CH₂)I₁(I- or 2- naphthyl), (CH₂)_nheteroaryl or (un)substituted (2-, 3-, A-, 5-, 6-, 7- or 8-) quinolinyl, fluorenmethyl ; (xi) Y is an electronegative leaving group including halogens such as F, Cl, Br or I, aryl or alkylsulfonyloxy groups, trifluoromethanesulfonyloxy, OR, SR, COOR, OP(O)R₂ wherein R is an aliphatic group, an aryl group, an alkyl group, a carbocyclic group, an alkyl carbocyclic group, or a heterocyclic group ; (xii) Z is a halogen (F, Cl, Br or I), CN, OH, OR, NH₂, NHR, NR₂, SR, COR, CO₂R,

CONR₂;

(xiii) R¹, R², R³ ,R⁴ ,R⁵ ,R⁶ and R⁷ may include in their structure radio labelling isotopes including (but not restrictively) Chlorine-36, Carbon-14, Tritium , Americium- 241, Caesium-137, Silver-l lOm, Cobalt-60, Lanthanum- 140, Scandium-46, GoId- 198, Cobalt-60, Gold-198, Technetium-99m, Strontium-90, Krypton-85, Thallium-

204, Gallium 67, thallium 201, Zinc-65, Manganese-54, Iridium-192, Chromium- 57, Ytterbium- 169, Iridium-192, Selenium-75;

(xiv) R¹, R², R³ ,R⁴ ,R⁵ ,R⁶ and R⁷ may include (but not restrictively) "cold labeling" in their structure including ¹³C, ²H, ¹⁵N, ¹⁹F, ³¹P, ²³Na or 31 K ; (xv) (x) n is 0 to 20.

The term "aliphatic" herein means straight chained or branched Ci_₂o hydrocarbons which are completely saturated or which contain one or more units of unsaturation. The term "alkyl" used alone or as part of a larger moiety refers to both straight or branched chains containing one to twenty carbon atoms. The term "aryl" refers to mono cyclic or polycyclic aromatic ring groups having five to fourteen atoms, such as phenyl, naphthyl or anthryl. The term "heterocyclic group" refers to saturated or unsaturated polycyclic or monocyclic ring systems containing one or more heteroatoms and a ring size of three to nine such as furanyl, thienyl, pyrrolyl, pyrrolinyl, pyrrolidinyl, dioxolanyl, oxazolyl, thiazolyl, imidazolyl, imidazolinyl, imidazolidinyl, pyrazolyl, pyrazolinyl, pyrazolidinyl, isoxalolyl, isothiazolyl, oxadiazolyl, dioxanyl, morpholinyl, dithianyl, thiomorpholinyl, pyridazinyl, pyrimidinyl, pyrazinyl, piperazinyl, triazinyl, trithianyl, indolizinyl, indolyl, isoindolyl, indolinyl, benzofuranyl, benzothiophenyl, indazolyl, benzamidazolyl, benzthiazolyl, purinyl, quinolizinyl, quinolinyl, isoquinolinyl, cinnolinyl, phthalazinyl, quinazolinyl, quinoxalinyl, 1,8-naphthyridinyl, pteridinyl, quinuclidinyl, carbazolyl, acridinyl, phenazinyl, phenothiazinyl, or phenoxazinyl. "Heteroaryl" refers to a heterocyclic ring that is aromatic. The term "carbocyclic group" refers to unsaturated monocyclic or polycyclic carbon ring systems of three to fourteen carbons which may be fused to aryl or heterocyclic groups. An aliphatic, alkyl, aryl, heteroaryl (ex: quinoline), heterocyclyl, or carbocyclyl, used alone or as part of a larger moiety, refers to substituted or unsubstituted groups. When substituted, these groups may contain one or more substituents. These substituents can be halogen (F, Cl, Br, I), OH, U, CO(CH₂)_nNH(U), CO(CH₂)_nS(U), OR, SR, NH₂, NHR, NR₂, OCOR, OP(O)R₂ wherein R is an aliphatic group, an aryl or substituted group, an aralkyl group, a carbocyclic group, an alkyl carbocyclic group, a heterocyclic group or a radio-isotope (ex: I¹²⁵, H³, S³⁵, C¹⁴, P³³, P³², Cr⁵¹, Ca⁴⁵, Fe⁵⁹, Ni⁶³, Ba¹³³, Cs¹³⁷, Eu¹⁵², Ra ²²⁶, Xe¹³³, technetium 99, thallium 201). FITC stands for fluorescein isothiocyanate.

As shown by the results described hereinafter in the Examples, the above defined derivatives recognize caspase-1 and block its activity.

The invention thus relates to the use of said derivatives as inhibitors for preventing or blocking caspase-1 activity in in vitro or in vivo cell death.

Advantageously, said derivatives are used for In vivo inhibition of caspase-1 activity to provide cytoprotective effect.

According to another embodiment, they are used for //? vitro inhibition of caspase-1 activity.

The invention also relates to the use of said derivatives as drugs.

It is then an object of the invention to provide pharmaceutical compositions comprising an effective amount of at least one derivative such as above defined in association with a pharmaceutically acceptable carrier.

Said pharmaceutical compositions are useful for administration by oral, nasal, local (intracerebroventricular, intrahippocampal, other intracerebral delivery, intracerebral implantation of instrumentation for mechanical delivery) or systemic (for example: intraperitoneal, intravenous....) administration or intradermic delivery, intrathecal delivery or as aerosol to reduce in vivo liver cell death, but not restricted to these delivery systems. They are administered in amounts adapted to the patient and the disease to be prevented or treated.

The pharmaceutical compositions of the invention are useful to prevent and/or treat oxygen-induced organ damages linked to anoxia, hypoxia or hyperoxia to prevent and/or treat inflammation or inflammatory diseases, inflammatory bowel disease, sepsis and septic shock, or - to prevent and/or treat apoptosis during degenerative diseases including Alzheimer's disease, Huntingtons' disease, Parkinsons' disease, Multiple sclerosis, amyotrophic lateral sclerosis, spinobulbar atrophy, prion disease, dementia; brain hypoxia, anoxia, hyperoxia; ischemic multifocal lesions involving the cortical or lenticulo striate branches of the MCA, ischemic lesions in the territory of the middle cerebral artery (MCA) or left cerebral hemisphere, caused by haemodynamic differences from a patent ductus arteriosus, or a more direct route involving the left common carotid; focal arterial infarction, or to prevent and/or treat retinal pericyte apoptosis, retinal neurons apoptosis glaucoma, retinal damages resulting from local ischemia, diabetic retinopaty, or - to prevent and/or treat epilepsy, or to prevent and/or treat apoptosis during spinal cord injury, or to prevent and/or treat apoptosis resulting from traumatic brain injury, retinal ischemia, or to prevent and/or treat apoptosis during pathological situations of focal cerebral ischemia, or - to provide cerebroprotective effect, or to prevent and/or treat cytotoxic T cell and natural killer cell-mediated apoptosis associated with autoimmune disease and transplant rejection, or to prevent and/or treat cell death of cardiac cells including heart failure, cardiomyopathy, viral infection or bacterial infection of heart, myocardial ischemia, myocardial infarct, and myocardial ischemia, coronary artery by-pass graft, or to prevent and/or treat mitochondrial drug toxicity e.g. as a result of chemotherapy or HIV therapy, to prevent and/or treat cell death during viral infection or bacterial infection, or to prevent cell death from follicle to ovocyte stages, from ovocyte to mature egg stages and sperm (for example, methods of freezing and transplanting ovarian tissue, artificial fecondation), or to preserve fertility in women and men after chemotherapy, or to preserve fertility in females and males animals, or to prevent and/or treat, macular degenerescence and glaucoma, or to prevent and/or treat acute hepatitis, chronic active hepatitis, hepatitis-B, and hepatitis-C, or to prevent and/or treat hair loss, and said hair loss due-to male-pattern baldness, radiation, chemotherapy or emotional stress, or - to treat or ameliorate skin damage (due to exposure to high level of radiation, heat, burns, chemicals, sun, and autoimmune diseases), or to prevent cell death of bone marrow cells in myelodysplastic symdromes (MDS), or to prevent and/or treat pancreatisis, or to prevent and/or treat respiratory symdrome, or - to prevent and/or treat osteoarthitis, rheumatoid arthritis, psoriasis, glomerulonephritis, atheroscerosis, and graft versus host disease, or to prevent and/or treat disease states associated with an increase of apoptosis, or to prevent cell death in vegetals (for example: plants, flowers, thallophytes

(mushrooms, seaweed)...)

The term "treatment or treat" as used herein means an approach for obtaining beneficial or desired results, including clinical results. Beneficial or desired clinical results can include, but are not limited to, alleviation or amelioration of one or more symptoms or conditions, diminishment of extent of disease, stabilized (i.e. not worsening) state of disease, preventing spread of disease, delay or slowing of disease progression, amelioration or palliation of the disease state, and remission (whether partial or total), whether detectable or undetectable. "Treat" can also mean prolonging survival as compared to expected survival if not receiving treatment.

Other characteristics and advantages of the invention are given in the following examples, wherein it is referred to Figures 1 and 2,

Figures 1 and 2 representing the caspase-1 activity (%) as a function of inhibitor concentration (μM).

In vitro caspase activity assays:

In vitro caspase activity assays were performed as described hereinafter to determine the inhibitory profile of the above derivative of the invention. 25U of human (caspase-1 to -10 and -14; Bio mo 1, Plymouth, Pennsylvania, USA) or rodent (mouse caspase-1, Biovision, Mountain View, USA).

Recombinant caspases were pre-incubated during 30 min at 37°C with the test compound (or not) in the assay buffer (50 mM HEPES, pH 7.4, 10OmM NaCl, 0.1% CHAPS, 10 mM DTT, ImM EDTA, 10% glycerol) and then mixed with 200 μM of the corresponding fluorogenic caspase substrates (Biomol , Sigma, St-Quentin Fallavier, France): Ac-YVAD-AMC (caspase- 1, Cl), Ac-VDVAD-AMC (Caspase-2, C2), Ac-DEVD-AMC (caspase-3/7, C3 and CJ), Ac- WEHD-AMC (caspase-4,-5,-11, -14; C4, C5, CI l, C14), Ac-VEID-AMC (caspase-6; C6), Ac-IETD-AMC (caspase-1; C8), Ac-LEHD-AMC (caspase-9; C9) or Ac-IETD-AMC (caspase- 10; ClO). Substrate cleavage was assessed at 2-3h by spectrofluorimetry (TECAN model Genios; λ_ex= 380 nm; λ_em= 465 nm). An IC50 value corresponding to the concentration that could inhibit 50% of caspase activity was determined from the dose-response sigmoid curve. Specific activity of each caspase was internally controlled with the corresponding specific inhibitors (1-2 μM, Biomol and MPQBiogene (Illkirsh, France). Calpain inhibitor (E64d, 1 μM; Sigma) was used as negative control for caspase inhibition.

As shown by the results given on Figure 1, the above defined derivative of formula (I) inhibits selectively human recombinant caspase-1 in vitro (with an ICso= 50 ± 15 nM, Figure 1) and also inhibits mouse caspase-1 (with an ICso= 50 ± 8 nM, Figure I)). Said derivative may poorly cross-react with others caspases (Figure 2).

Claims

1/ Selective new caspase-1 inhibitors that recognize caspase-1 and block its activity based on SEQl.

2/ Selective new caspase-1 inhibitors that recognize caspase-1 and block its activity based on a salt form of SEQl (preferentially but not exclusively a chlorhydrate form).

3/ Selective new caspase-1 inhibitors that recognize caspase-1 and block its activity based on demethylated form of SEQ 1.

4/ Selective new caspase-1 inhibitors that recognize caspase-1 and block its activity derived from the inhibitor according to anyone of claims 1 to 3 and based on structure II or SEQ2.

5/ Selective new caspase-1 inhibitors that recognize caspase-1 and block its activity derived from the inhibitor according to anyone of claims 1 to 3.

6/ Inhibitors according to anyone of claims 1 to 3, for preventing or blocking caspase-1 activity in in vitro or in vivo cell death.

11 Inhibitors according to claim 4, for preventing or blocking caspase-1 activity in in vitro or in vivo cell death.

8/ In vivo inhibition of caspase-1 activity with at least one molecule containing at least one sequence according to anyone of claims 1 to 3 to provide cytoprotective effect.

9/ In vivo inhibition of caspase-1 activity with at least one molecule containing at least one sequence according to claim 4 to provide cytoprotective effect.

12/ In vitro inhibition of caspase-1 activity with at least one molecule according to anyone of claims 1 to 4.

13/ Pharmaceutical compositions, comprising an effective amount of at least one compound as defined in anyone of claims 1 to 12. 14/ Pharmaceutical compositions, comprising a therapeutically effective amount of at least one caspase-1 inhibitor according to anyone of claims 1 to 13, in association or not with a pharmaceutically acceptable carrier.

15/ The pharmaceutical compositions according to claim 13 or 14 for administration by oral, nasal, local (intracerebroventricular, intrahippocampal, other intracerebral delivery, intracerebral implantation of instrumentation for mechanical delivery) or systemic (for example: intraperitoneal, intravenous....) administration or intradermic delivery, intrathecal delivery or as aerosol to reduce in vivo liver cell death, but not restricted to these delivery systems.

16/ Pharmaceutical compositions, comprising an effective amount of a combination of at least one compound according to anyone of claims 1 to 15, in association or not with a pharmaceutically acceptable carrier.

17/ The pharmaceutical compositions according to anyone of claims 13 to 16: to prevent and/or treat inflammation or inflammatory diseases, inflammatory bowel disease, sepsis and septic shock, or - to prevent and/or treat apoptosis during neurodegenerative diseases including

Alzheimer's disease, Huntingtons' disease, Parkinsons' disease, Multiple sclerosis, amyotrophic lateral sclerosis, spinobulbar atrophy, prion disease, dementia; ischemic multifocal lesions involving the cortical or lenticulo striate branches of the MCA, ischemic lesions in the territory of the middle cerebral artery (MCA) or left cerebral hemisphere, caused by haemodynamic differences from a patent ductus arteriosus, or a more direct route involving the left common carotid; focal arterial infarction; anoxia-, or hypoxia-, or hyperoxia brain damages, or to prevent and/or treat oxygen- induced organ damages, or to prevent and/or treat retinal pericyte apoptosis, retinal neurons apoptosis glaucoma, retinal damages resulting from local ischemia, diabetic retinopaty, or to prevent and/or treat epilepsy, or to prevent and/or treat apoptosis during spinal cord injury, or to prevent and/or treat apoptosis resulting from traumatic brain injury, retinal ischemia, or to prevent and/or treat apoptosis during pathological situations of focal cerebral ischemia, or to provide cerebroprotective effect, or to prevent and/or treat cytotoxic T cell and natural killer cell-mediated apoptosis associated with autoimmune disease and transplant rejection, or to prevent and/or treat cell death of cardiac cells including heart failure, cardiomyopathy, viral infection or bacterial infection of heart, myocardial ischemia, myocardial infarct, and myocardial ischemia, coronary artery by-pass graft, or to prevent and/or treat mitochondrial drug toxicity e.g. as a result of chemotherapy or HIV therapy, to prevent and/or treat cell death during viral infection or bacterial infection, or to prevent cell death from follicle to ovocyte stages, from ovocyte to mature egg stages and sperm (for example, methods of freezing and transplanting ovarian tissue, artificial fecondation), or - to preserve fertility in women and men after chemotherapy, or to preserve fertility in females and males animals, or to prevent and/or treat, macular degenerescence and glaucoma, or to prevent and/or treat acute hepatitis, chronic active hepatitis, hepatitis-B, and hepatitis-C, or - to prevent and/or treat hair loss, and said hair loss due-to male-pattern baldness, radiation, chemotherapy or emotional stress, or to treat or ameliorate skin damage (due to exposure to high level of radiation, heat, burns, chemicals, sun, and autoimmune diseases), or to prevent cell death of bone marrow cells in myelodysplastic symdromes (MDS), or - to prevent and/or treat pancreatisis, or to prevent and/or treat respiratory symdrome, or to prevent and/or treat osteoarthitis, rheumatoid arthritis, psoriasis, glomerulonephritis, atheroscerosis, and graft versus host disease, or to prevent and/or treat disease states associated with an increase of apoptosis, or - to prevent cell death in vegetals (for example: plants, flowers, thallophytes

(mushrooms, seaweed)...)

The term "treatment or treat" as used herein means an approach for obtaining beneficial or desired results, including clinical results. Beneficial or desired clinical results can include, but are not limited to, alleviation or amelioration of one or more symptoms or conditions, diminishment of extent of disease, stabilized (i.e. not worsening) state of disease, preventing spread of disease, delay or slowing of disease progression, amelioration or palliation of the disease state, and remission (whether partial or total), whether detectable or undetectable. "Treat" can also mean prolonging survival as compared to expected survival if not receiving treatment.