DE102008029072A1 - Substance, which inhibits serum and glucocorticoid dependent kinase 3, useful for the prophylaxis and/or treatment or diagnosis of age-related diseases e.g. arteriosclerosis, skin atrophy, myasthenia, infertility, stroke and kyphosis - Google Patents

Abstract

Substance, for the prophylaxis and/or treatment or diagnosis of age-related diseases, is claimed, where the substance: inhibits the expression and/or activity of serum and glucocorticoid dependent kinase 3 (sgK3); and is useful for detecting the expression, activity and/or gene modification of sgk3. Independent claims are included for: (1) a diagnosis kit, for diagnosis of age-related diseases, comprising at least one substance for detecting the expression, activity and/or gene modification of sgk3; and (2) a method for diagnosis of age-related diseases, where the expression, activity and/or gene modification of sgk3 for detecting in body sample of a patient with antibody against sgk3, oligonucleotide (with which specific DNA-fragments of sgk3 are amplified in PCR) and/or polynucleotide (that are hybridized with DNA or mRNA of sgk3 under stringent conditions) is detected. ACTIVITY : Antiarteriosclerotic; Respiratory-Gen.; Dermatological; Muscular-Gen.; Neuroprotective; Immunosuppressive; Immunostimulant; Antiinfertility; Metabolic; Cardiant; Cerebroprotective; Vasotropic; Nootropic; Antidiabetic; Ophthalmological; Cytostatic; Osteopathic; Antiparkinsonian; Antiallergic; Antiinflammatory; Antiarrhythmic; Uropathic. MECHANISM OF ACTION : Kinase inhibitor; Sgk3 inhibitor.

Description

The The present invention relates to substances for prophylaxis and / or Treatment and diagnosis of geriatric diseases, a diagnostic procedure as well as a related diagnostic kit.

A variety of external signals to which a cell is exposed in its environment lead to intracellular phosphorylation or dephosphorylation cascades to allow rapid and reversible transfer of these signals from the plasma membrane and its receptors into the cytoplasm and nucleus. The regulation of individual proteins involved in these cascades first allows the high specificity and flexibility of the cells, which allow them to respond very quickly to extracellular signals. In particular, kinases are involved in these regulatory processes. Serum and glucocorticoid-dependent kinase (sgk) was originally cloned from rat breast carcinoma cells ( Webster MK, Goya L, Firestone GL. J. Biol. Chem. 268 (16): 11482-11485, 1993 ; Webster MK, Goya L, Ge Y, Maiyar AC, Firestone GL. Mol. Cell. Biol. 13 (4): 2031-2040, 1993 ). The human kinase hsgk was cloned as a cell volume-regulated gene from liver cells ( Waldegger S, Barth P, Raber G, Lang F. Proc. Natl. Acad. Sci. USA 94: 4440-4445, 1997 ). The rat kinase was shown to stimulate the epithelial Na channel (EnaC) ( Chen SY, Bhargava A, Mastroberardino L, Meijer OC, Wang J, Buse P, Firestone GL, Verrey F, Pearce D. Proc. Natl. Acad. Sci. USA 96: 2514-2519, 1999 ; Naray-Fejes-Toth A, Canessa C, Cleaveland ES, Aldrich G, Fejes-Toth GJ Biol. Chem. 274: 16973-16978, 1999 ). Furthermore, it could be shown that an increased activity of ENaC is associated with hypertension ( Warnock DG. Kidney Ind. 53 (1): 1824, 1998 ).

The hsgk is also expressed in the brain ( Waldegger S, Barth P, Raber G, Lang F. Proc. Natl. Acad. Sci. USA 94: 4440-4445, 1997 ), where it regulates the voltage-dependent K ⁺ channels Kv1.3. It has been shown that these K ⁺ channels of the Kv1.3 type are involved in the regulation of neuronal excitability ( Pongs O. Physiol. Rev. 72: 69-88, 1992 ), regulation of cell proliferation ( Cahalan MD and Chandy KG. Cur. Opin. Biotech. 8 (6): 749-756, 1997 ) as well as the regulation of apoptotic cell death are involved ( Szabo I, Gulbins E, Apple H, Zhan X, Barth P, Busch AE, Schlottmann K, Pongs O, Lang FJ Biol. Chem. 271: 20465-20469, 1999 ; Lang F, Szabo I, Lepple-Wienhues A, Siemen D, Gulbins E. Newes Physiol. Sci. 14: 194-200, 1999 ). The Kv1.3 channel is also important in the regulation of lymphocyte proliferation and function ( Cahalan MD and Chandy KG, Cur. Opin. Biotech. 8 (6): 749-756, 1997 ).

Two other members of the sgk family have been cloned, the sgk2 and sgk3 ( Kobayashi T, Deak M, Morrice N, Cohen P. Biochem. J. 344: 189-197, 1999 ). It turned out that the sgk members form a serine-threonine protein kinase family that can be transcriptionally and post-transcriptionally regulated. Like sgk1, sgk3 is also ubiquitously expressed and activated by insulin and growth factors via phosphatidylinositol 3-kinase and PDK (3-phosphoinositide dependent kinase 1).

With regard to sgk3 it could be shown that a genetic elimination of this protein leads to delayed hair growth ( McCormick et al., 2004 ), a defect attributed to apoptosis of keratinocytes ( Alonso L, Okada H, Pasolli HA, Wakeham A, You T, Mak TW, Fox E: Sgk3 left growth factor signaling to progenitor cells in the hair follicle. J Cell Biol 2005; 170: 559-570 ). In addition, decreased intestinal glucose transport in sgk3 knockout mice ( Sandu C, Rexhepaj R, Grahammer F, McCormick JA, Henke G, Palmada M, Nammi S, Lang U, Butcher M, Just L, Skutella T, Dawson K, Wang J, Pearce D, Lang F: Decreased intestinal glucose transport in the sgk3-knockout mouse Pflugers Arch 2005; 451: 437-444 ) and a slight change in movement ( Lang UE, Wolfer DP, Grahammer F, Strutz-Seebohm N, Seebohm G, Lipp HP, McCormick JA, Hellweg R, Dawson K, Wang J, Pearce D, Lang F: Reduced locomotion in the serum and glucocorticoid inducible kinase 3 knockout mouse , Behav Brain Res 2006b; 167: 75-86 ).

The use of sgk3 as a therapeutic and diagnostic target for diseases associated with impaired activity of ion channels is known from US Pat WO 02/17893 A2 known.

The Invention sets itself the task of further therapeutic and diagnostic Applications using sgk3 as the target molecule or Target provide.

These The object is achieved by the subject matter of independent claims 1, 9 and FIGS. 12 to 15. Preferred embodiments are disclosed in FIGS called dependent claims. The content of all this Claims are hereby incorporated by reference Description made.

According to the invention a substance for the prophylaxis and / or treatment or therapy of Age-related diseases are provided, the substance being expression and / or the activity of sgk3 (serum and glucocorticoid dependent kinase 3) inhibits or inhibits.

Surprisingly, animal experiments have shown that switching off or knocking out sgk3 leads to a delay in biological aging and thus to a prolongation in the relevant experimental animals. To elucidate the influence of sgk isoforms, in particular of sgk3, on the occurrence of age-related diseases and thus on the general lifespan or life expectancy, klotho-deficient mice (klotho ^{- / -} ) with sgk1-deficient mice (sgk1 ^{- / -} ) and sgk3-deficient mice (sgk3 ^{- / -} ) were crossed. Generally, the Klotho gene encodes a membrane protein which is expressed mainly in the kidney, parathyroid glands and choroid plexus ( Takeshita K, Fujimori T, Kurotaki Y, Honjo H, Tsujikawa H, Yasui K, Lee JK, Kamiya K, Kitaichi K, Yamamoto K, Ito M, Kondo T, lino S, Inden Y, Hirai M, Murohara T, Kodama I , Nabeshima Y: Sinoatrial node dysfunction and early death of mice with a defect of clot gene expression. Circulation 2004; 109: 1776-1782 . Tsujikawa H, Kurotaki Y, Fujimori T, Fukuda K, Nabeshima Y: Klotho, a gene related to a syndrome resembling human premature aging, functions in a negative regulatory circuit of vitamin D endocrine system. Mol Endocrinol 2003; 17: 2393-2403 ). The extracellular domain of the Klotho protein can be cleaved and released into the cerebrospinal fluid and blood ( Imura A, Ivano A, Tohyama O, Tsuji Y, Nozaki K, Hashimoto N, Fujimori T, Nabeshima Y: Secreted Klotho protein in sera and CSF: implication for post-translational cleavage in the release of Klotho protein from cell membrane. FEBS Lett 2004; 565: 143-147 ). It is known that the genetic elimination of the Klotho gene in mice leads to a dramatically shortened lifespan and premature onset of age-related disorders such as atherosclerosis, pulmonary emphysema, skin atrophy, muscle weakness, immunodeficiency, infertility, kyphosis and impaired CaPO ₄ metabolism (calcium Phosphate metabolism) with osteoporosis and extensive calcification of connective tissue ( Kuro-o M, Matsumura Y, Aizawa H, Kawaguchi H, Suga T, Utsugi T, Ohyama Y, Kurabayashi M, Kaname T, Kume E; Iwasaki H, lida A, Shiraki-lida T, Nishikawa S, Nagai R, Nabeshima YI: mutation of mouse klotho gene leads to a syndrome resembling aging. Nature 1997; 390: 45-51 ). In contrast, an overexpression of the Klotho gene causes a significantly extended lifespan ( Kurosu H, Yamamoto M, Clark JD, Pastor JV, Nandi A, Gurnani P, Mc-Guinness OP, Chikuda H, Yamaguchi M, Kawaguchi H, Shimomura I, Takayama Y, Heart J, Kahn CR, Rose Leaf KP, Kuro-o M: Suppression of aging in mice by the hormone Klotho. Science 2005; 309: 1829-1833 ). In addition, it was demonstrated that the shortened lifespan in the case of Klotho-deficient mice is associated with an accelerated occurrence of age-related diseases ( Aizawa H, Saito Y, Nakamura T, Inoue M, Imanari T, Ohyama Y, Matsumura Y, Masuda H, Oba S, Mise N, Kimura K, Hasegawa A, Kurabayashi M, Kuro-o M, Nabeshima Y, Nagai R: Downregulation of the Klotho gene in the kidney under sustained circulatory stress in rats. Biochem Biophys Res Commun 1998; 249: 865-871 ; Kuro-o M: Klotho as a regulator of oxidative stress and senescence. Biol Chem 2008; 389: 233-241 ; Lanske B, Razzaque MS: Premature aging in klotho mutant mice: cause or consequence? Aging Res Rev 2007; 6: 73-79 ; Saito Y, Yamagishi T, Nakamura T, Ohyama Y, Aizawa H, Suga T, Matsumura Y, Masuda H, Kurabayashi M, Kuro-o M, Nabeshima Y, Nagai R: Klotho protein pro tects against endothelial dysfunction. Biochem Biophys Res Commun 1998; 248: 324-329 ; Suga T, Kurabayashi M, Sando Y, Ohyama Y, Maeno T, Maeno Y, Aizawa H, Matsumura Y, Kuwaki T, Kuro O, Nabeshima Y, Nagai R: Disruption of the Klotho gene causes pulmonary emphysema in mice. Defect in maintenance of pulmonary integrity during postnatal life. On J Respir Cell Mol Biol 2000; 22: 26-33 ). A connection between the expression of variants of the Klotho gene and longevity on the one hand and the absence of the Klotho gene and the manifestation of arteriosclerosis on the other hand has already been demonstrated in humans ( Arking DE, Krebsova A, Macek M, Sr., Macek M, Jr., Arking A, Mian IS, Fried L, Hamosh A, Dey S, Mclntosh I, Dietz HC: Association of human aging with a functional variant of klotho. Proc Natl Acad Sci USA 2002; 99: 856-861 ; Arking DE, Becker DM, Yanek LR, Fallin D, Judge DP, Moy TF, Becker LC, Dietz HC: KLOTHO allele status and the risk of early-onset occult coronary artery disease. At the J Hum Genet 2003; 72: 1154-1161 ; Freathy RM, Weedon MN, Melzer D, Shields B, Hitman GA, Walker M, McCarthy MI, Hattersly AT, Frayling TM: The functional "KL-VS" variant of KLOTHO is not associated with type 2 diabetes in 5028 UK Caucasians. BMC Med Genet 2006; 7:51 ; Imamura A, Okumura K, Ogawa Y, Murakami R, Torigoe M, Numaguchi Y, Murohara T: Klotho gene polymorphism may be a genetic risk factor for atherosclerotic coronary artery disease but not vasospastic angina in Japanese. Clin Chim Acta 2006; 371: 66-70 ; Low AF, O'Donnell CJ, Kathiresan S, Everett B, Chae CU, Shaw SY, Ellinor PT, Macrae CA: Aging syndrome genes and premature coronary artery disease. BMC Med Genet 2005; 6:38 ). The Klotho gene is therefore a gene in both humans and mice which can delay the onset of age-related diseases. Therefore, mouse knockout mice or knockout knockout mice are ideal model systems for the study of biological aging.

In the context of the invention it has now been demonstrated that the negative consequences in mice a genetic elimination of the Klotho gene, namely the accelerated occurrence of age-related diseases, by the simultaneous elimination of the gene for sgk3 can be significantly delayed. Namely, the mice exhibiting both a knockout of the klotho gene and a knockout of the sgk3 gene showed a significantly longer life expectancy than mice having only a knockout of the klotho gene (cf. 2 ).

Under Age disorders in the sense of the present invention are said to be diseases or diseases that are due to the biological Aging process in humans and / or animals can occur.

Under Activity within the meaning of the present invention is intended to be Enzyme activity, in particular the kinase activity, to be understood by sgk3.

In a preferred embodiment is in the Substance around a substance directed against sgk3. Preferably said substance is an anti-sgk3 antibody. The production of anti-sgk3 antibodies, which are monoclonal or polyclonal antibodies can act, the skilled person is well known. Appropriate For example, antibodies can be made thereby that animals, such as horses, cattle, pigs, Sheep or rabbits, protein fragments of sgk3 for immunization purposes be administered. After administration of the sgk3 fragments produce the animals specifically against sgk3-directed antibodies, which is isolated and purified by standard procedures can be.

In In another embodiment, the substance is against activators, Inhibitors, regulators, biological precursors, variants, especially isoforms, directed by sgk3. The ones listed above For example, compounds can be up and / or downstream lying members of the sgk3 signal transduction cascade, transcription factors, which is responsible for the level of expression of sgk3 are, and / or proteases, which are for the proteolytic Degradation of activators, inhibitors, regulators, biological precursors or variants of sgk3 are responsible. Suitable activators are, for example, the phosphatidylinositol-3-kinase mentioned in the introduction and PDK1. As an inhibitor, for example, the ubiquitin ligase Nedd4-2 into consideration. The transcription factors may be for example, by at least one factor from the group Forkhead transcription factor FKHRL1, β-catenin and NFκB (nuclear factor kappa B).

Furthermore, the substance may be a polynucleotide which codes for a peptide, preferably a polypeptide, which inhibits the expression and / or activity of sgk3. For example, the substance which can be used according to the invention may be a so-called knase deficient mutant. Further examples of how the expression and / or activity of sgk3 can be influenced by genetically modified variants of sgk3 are familiar to the person skilled in the art and can be found in a number of textbooks and instructions for laboratory work ( Maniatis T, Fritsch EF, Sambruk J. Coldspring Harbor, NY: Coldspring Harbor Labrator 1996 ; Leonard D, Davis PhD, Michael W, Kuehl Md, James F, Battey MD. McCraw-Hill Professional Publishing, 1995 ).

Prefers it is the inventively usable Substance around a transdominant negative kinase mutant of sgk3. In general, genetically modified variants can be used of sgk3 by transfection or transduction into cells. In this case, genetically modified sgk3 variants in the As part of a chemical transfection into cells are introduced. In this case, the genes of the sgk3 variants together with a electrically charged compound, preferably calcium phosphate, to cells added. The electrically charged connection interferes doing the structure of the cell membrane, causing the DNA of the sgk3 variants can get into the cell interior. According to the invention it is equally possible that the gene variants of sgk3 get in the way a physical transfection into cells are introduced. As physical transfection methods are basically Microinjection, electroporation and loaded with sgk3 gene variants Particles, in particular gold particles, into consideration. According to the invention it is particularly preferred that gene variants of sgk3 by transduction, in particular by a viral vector, into cells to be treated or tissue are introduced. The viral vectors are based usually on so-called retroviruses. By the above Thus, it is possible to use the sequence of sgk3 itself to use in the appropriate cells or Tissues to achieve a therapeutic effect.

In a further embodiment, the substance is a low-molecular-weight compound ("small molecular compound"), preferably having a molecular weight (MW) <1000. According to the invention, it may be provided that the substance is a kinase inhibitor is. Before The substance is selected from the group consisting of staurosporine, chelerythrine, SB 203580 (MW 377.4), SB 202190 (MW 331.3) and derivatives derived therefrom. The kinase inhibitors SB 203580 and SB 202190 are imidazole derivatives which are described, for example, by Cal Biochem. be sold commercially.

worin
R¹, R² jeweils unabhängig voneinander H, CHO oder Acetyl,
R³, R⁴, R⁵, R⁶, R⁷,
R⁸, R⁹, R¹⁰, R¹¹ jeweils unabhängig voneinander
H, A, OSO₂A, Hal, NO₂, OR¹², N(R¹²)₂, CN, O-COA, -[C(R¹²)₂]_nCOOR¹², O-[C(R¹²)₂]₀COOR¹², SO₃H, -[C(R¹²)₂]_nAr, -CO-Ar, O-[C(R¹²)₂]_nAr, -[C(R¹²)₂]_nHet, -[C(R¹²)₂]_nO≡CH, O-[C(R¹²)₂]_nC≡CH, -[C(R¹²)₂]_nCON(R¹²)₂, -[C(R¹²)₂]_nCONR¹²N(R¹²)₂, O-[C(R¹²)₂]_nCON(R¹²)₂, O-[C(R¹²)₂]_oCONR¹²N(R¹²)₂, NR¹²COA, NR¹²CON(R¹²)₂, NR¹²SO₂A, N(SO₂A)₂, COR¹², S(O)_mAr, SO₂NR¹² oder S(O)_mA,
R³ und R⁴ zusammen auch CH=CH-CH=CH,
R³ und R⁴, R⁷ und R⁸
oder R⁸ und R⁹ zusammen auch Alkylen mit 3, 4 oder 5 C-Atomen,
worin eine oder zwei CH₂-Gruppen durch Sauerstoff ersetzt sein können,
A unverzweigtes oder verzweigtes Alkyl mit 1-6 C-Atomen, worin 1-7 H-Atome durch F ersetzt sein können, oder cylisches Alkyl mit 3-7 C-Atomen,
Ar unsubstituiertes oder ein-, zwei- oder dreifach durch Hal, A, OR¹², N(R¹²)₂, NO₂, CN, Phenyl, CON(R¹²)₂, NR¹²COA, NR¹²CON(R¹²)₂, NR¹²SO₂A, COR¹², SO₂N(R¹²)₂, S(O)_mA, -[C(R¹²)₂]_n-COOR¹² und/oder -O[C(R¹²)₂]_o-COOR¹² substituiertes Phenyl, Naphthyl oder Biphenyl,
Het einen ein- oder zweikernigen gestättigten, ungesättigten oder aromatischen Heterocyclus mit 1 bis 4 N-, O- und/oder S-Atomen, der ein-, zwei- oder dreifach durch Hal, A, OR¹², N(R¹²)₂, NO₂, CN, COOR¹², CON(R¹²)₂, NR¹²COA, NR¹²SO₂A, COR¹², SO₂NR¹², S(O)_mA, =S, =NR¹² und/oder =O (Carbonylsauerstoff) substituiert sein kann,
R¹² H oder A,
Hal F, Cl, Br oder I,
m 0, 1 oder 2,
n 0, 1, 2 oder 3,
o 1, 2 oder 3
bedeuten,
sowie ihre pharmazeutisch verwendbaren Derivate, Salze, Solvate und Stereoisomere, einschließlich deren Mischungen in allen Verhältnissen.In a further embodiment, the substance is a mandelic acid hydrazide derivative. The substance is preferably a compound of the formula I.

wherein
R ¹ , R ^{2 are} each independently H, CHO or acetyl,
R ³ , R ⁴ , R ⁵ , R ⁶ , R ⁷ ,
R ⁸ , R ⁹ , R ¹⁰ , R ^{11 are} each independently
H, A, OSO ₂ A, Hal, NO ₂ , OR ¹² , N (R ¹² ) ₂ , CN, O-COA, - [C (R ¹² ) ₂ ] _n COOR ¹² , O- [C (R ¹² ) ₂ ] ₀ COOR ¹² , SO ₃ H, - [C (R ¹² ) ₂ ] _n Ar, -CO-Ar, O- [C (R ¹² ) ₂ ] _n Ar, - [C (R ¹² ) ₂ ] _n Het, - [C (R ¹² ) ₂ ] _n O≡CH, O- [C (R ¹² ) ₂ ] _n C≡CH, - [C (R ¹² ) ₂ ] _n CON (R ¹² ) ₂ , - [ C (R ¹² ) ₂ ] _n CONR ¹² N (R ¹² ) ₂ , O- [C (R ¹² ) ₂ ] _n CON (R ¹² ) ₂ , O- [C (R ¹² ) ₂ ] _o CONR ¹² N ( R ¹² ) ₂ , NR ¹² COA, NR ¹² CON (R ¹² ) ₂ , NR ¹² SO ₂ A, N (SO ₂ A) ₂ , COR ¹² , S (O) _m Ar, SO ₂ NR ¹² or S (O ) _m A,
R ³ and R ⁴ together also CH = CH-CH = CH,
R ³ and R ⁴ , R ⁷ and R ⁸
or R ⁸ and R ⁹ together also alkylene having 3, 4 or 5 C atoms,
wherein one or two CH ₂ groups may be replaced by oxygen,
A is unbranched or branched alkyl having 1-6 C atoms, in which 1-7 H atoms may be replaced by F, or cyclic alkyl having 3-7 C atoms,
Ar is unsubstituted or mono-, di- or trisubstituted by Hal, A, OR ¹² , N (R ¹² ) ₂ , NO ₂ , CN, phenyl, CON (R ¹² ) ₂ , NR ¹² COA, NR ¹² CON (R ¹² ) ₂ , NR ¹² SO ₂ A, COR ¹² , SO ₂ N (R ¹² ) ₂ , S (O) _m A, - [C (R ¹² ) ₂ ] _n -COOR ¹² and / or -O [C (R ¹² ) ₂ ] _o -COOR ¹² substituted phenyl, naphthyl or biphenyl,
Het a mono- or binuclear saturated, unsaturated or aromatic heterocycle having 1 to 4 N, O and / or S atoms, the one, two or three times by Hal, A, OR ¹² , N (R ¹² ). ₂ , NO ₂ , CN, COOR ¹² , CON (R ¹² ) ₂ , NR ¹² COA, NR ¹² SO ₂ A, COR ¹² , SO ₂ NR ¹² , S (O) _m A, = S, = NR ¹² and / or = O (carbonyl oxygen) may be substituted,
R ¹² H or A,
Hal F, Cl, Br or I,
m 0, 1 or 2,
n 0, 1, 2 or 3,
o 1, 2 or 3
mean,
and their pharmaceutically usable derivatives, salts, solvates and stereoisomers, including mixtures thereof in all ratios.

The substance may in particular be selected from the group consisting of 2,4-dihydroxy-6-methylbenzoic acid N '- [2' (3,4-difluoro-phenyl) -2-hydroxy-acetyl] -hydrazide, 2,4-dihydroxy- 6-methylbenzoic acid N '- [2-hydroxy-2- (3-phenoxy-phenyl) -acetyl] hydrazide, 2,4-dihydroxy-6-methylbenzoic acid N' - (2-hydroxy-2 -phenyl-acetyl) hydrazide, 2,4-dihydroxy-6-methylbenzoic acid N '- [2-hydroxy-2- (3-chloro-phenyl) -acetyl] -hydrazide, 2,4-dihydroxy-6 Methyl-benzoic acid N '- [2-hydroxy-2- (3-trifluoromethyl-phenyl) -acetyl] -hydrazide, 2,4-Dihydroxy-6-methyl-benzoic acid-N' - [2-acetoxy-2- (3-chloro-phenyl) -acetyl] -hydrazide, 2,4-Dihydroxy-6-methylbenzoic acid N '- [2-hydroxy-2- (3-fluoro-phenyl) -acetyl] -hydrazide, 2, 4-Dihydroxy-6-methylbenzoic acid N '- [2-hydroxy-2- (3,5-difluoro-phenyl) -acetyl] hydrazide, 2,4-dihydroxy-6-methylbenzoic acid N' (2-hydroxy-2- (2,3,4,5,6-pentafluorophenyl) acetyl] hydrazide, 2,4-dihydroxy-6-methylbenzoic acid N '- (2-benzo [1, 3] dioxol-5-yl-2-hydroxy-acetyl) hydrazide, 2,4-dihydroxy-6-methylbenzoic acid N '- [2-hydroxy-2- (3-h ydroxy-4-methoxy-phenyl) -acetyl] -hydrazide, 2,4-dihydroxy-5-chloro-benzoic acid N '- [2-hydroxy-2- (3-chloro-phenyl) -acetyl] -hydrazide, 3 Chloro-2-ethyl-4-hydroxy-benzoic acid N '- [2-hydroxy-2- (3-chloro-phenyl) -acetyl] -hydrazide, 2,4-dihydroxy-6-methylbenzoic acid N' - [(R) -2- (3-chloro-phenyl) -2-hydroxy-acetyl] -hydrazide, 2-chloro-4,6-dihy hydroxy-benzoic acid N '- [2-hydroxy-2- (3-hydroxy-phenyl) -acetyl] -hydrazide, 2-methyl-4,6-dihydroxy-benzoic acid N' - [2-hydroxy-2- (2-hydroxy) 3-hydroxy-phenyl) -acetyl] -hydrazide, 2-ethyl-4,6-dihydroxybenzoic acid N '- [2-hydroxy-2- (3-hydroxy-phenyl) -acetyl] -hydrazide, 2-methyl 4,6-dihydroxybenzoic acid N '- (2-hydroxy-2-phenyl-acetyl) hydrazide, 2,4-dihydroxy-6-methylbenzoic acid N' - [2-hydroxy-2- (3 -methyl-phenyl) -acetyl] -hydrazide, 2,4-Dihydroxy-5-methylbenzoic acid N '- [2-hydroxy-2- (3,4-difluoro-phenyl) -acetyl] -hydrazide, 2, 4-Dihydroxy-6-methylbenzoic acid N '- [2-formyloxy-2- (3,4-difluoro-phenyl) -acetyl] hydrazide, 2,4-dihydroxy-6-ethylbenzoic acid N' [2-hydroxy-2- (3,4-difluoro-phenyl) -acetyl] -hydrazide and 4-hydroxy-2-ethyl-3-methylbenzoic acid N '- [2-hydroxy-2- (3,4 -difluorophenyl) acetyl] hydrazide.

The compounds mentioned in the previous section are in the WO 2007/093264 A1 listed as compounds "A1" to "A23". With regard to further features and details of these compounds and their preparation is therefore on the WO 2007/093264 A1 the content of which is incorporated herein by express reference.

The Invention further relates to a substance for the diagnosis of geriatric diseases, wherein the substance is for detection of expression, activity and / or a gene modification of sgk3 (serum and glucocorticoid dependent kinase 3) is used in eukaryotic cells. With For example, help with the substance can be a change activity of sgk3. Preferred is the substance is an anti-sgk3 substance, especially an anti-sgk3 antibody. Such an antibody may be within the skill of the art known detection methods, for example ELISA (enzyme-linked immunosorbant assay). In this immunoassay becomes specific to the antigen to be determined Antibodies (or homologous for antibody determinations Test antigens) to a carrier material, for example cellulose or polystyrene, bound. Form on the carrier material after incubation with a sample containing the antigen, Immune complexes. In a subsequent step, these immune complexes become a labeled antibody, which also against the However, antigen is judiciously addressed another site of the antigen binds than that on the substrate bound antibodies added. In this marked Antibody is usually an antibody-enzyme conjugate, the enzyme is usually alkaline phosphatase or horseradish peroxidase. By the addition of the marked Antibody ultimately forms ternary complexes from the antigen and the two each directed against the antigen Antibodies. These ternary complexes can be by adding chromogenic substrates, for example para-nitrophenol, to make visible. From this, the antigen concentration in the sample over a photometric determination of the immunocomplex-linked marker enzymes by comparison with standards of known enzyme activity be determined. Likewise it is possible to directed against sgk3 To use antibodies in ELISPOT procedures. These methods are sufficiently familiar to the person skilled in the art, so that is waived further comments on this. Both in question, gene modifications may be in particular Nucleotide polymorphisms and / or insertion mutations. at the samples are usually body samples of a patient containing eukaryotic cells. As samples come tissue samples as well as samples of body fluids, for example, blood, in question.

According to one Another embodiment is the substance to an oligo- or polynucleotide, which with sgk3 under stringent Conditions hybridized. With the help of such oligo- or polynucleotides For example, Southern or Northern blots may be performed to detect the DNA or mRNA content of sgk3. Corresponding methods are also sufficiently familiar to the person skilled in the art. In this way, for example, the transcription rate of sgk3 measured in diseased or diseased tissues and with the corresponding transcription rate of sgk3 in healthy Tissues are compared.

The Age disorders of the present invention are preferred the group arteriosclerosis, pulmonary emphysema, skin atrophy ("aging skin"), Muscle weakness, immunodeficiency, infertility, Kyphosis, connective tissue caecification, myocardial infarction, stroke, Osteoarthritis, dementia, diabetes mellitus, cataract, cancer, osteoporosis, Parkinson's disease, chronic senile rhinitis, atrial fibrillation and Incontinence selected.

The The present invention further relates to a diagnostic kit for the diagnosis of geriatric diseases. This includes at least one Substance that is required for the detection of expression, activity and / or a gene modification of sgk3 is suitable. In terms of Other features and characteristics of the diagnostic kit will be on the previous description with reference.

Furthermore, the invention comprises a method, preferably an in vitro method, for the diagnosis of Age disorders, wherein the expression, activity and / or a gene modification of sgk3 by a preferably quantitative detection in a body sample of a patient with anti-sgk3 antibodies, with oligonucleotides, which, for example, in a polymerase chain reaction (PCR) certain DNA sections of sgk3 are amplified, and / or with polynucleotides which hybridize with DNA or mRNA of sgk3 under stringent conditions is detected. As already mentioned, tissue samples and / or blood samples may be considered as body samples. In this method, it is provided in particular to detect by means of the abovementioned substances also certain mutations, for example insertion mutations, or polymorphisms in sgk3.

Farther The present invention also relates to a method for prophylaxis and / or treatment of geriatric diseases, including administration a therapeutically effective amount of a substance which inhibits expression and / or activity of sgk3. In terms of the substances in question is based on the previous description directed.

After all The invention also relates to the use of a substance for the production a medicament for the prophylaxis and / or treatment of geriatric diseases, wherein the substance is the expression and / or activity of sgk3 inhibits or inhibits. Furthermore, the invention also relates the use of a substance for the preparation of a diagnostic agent for the diagnosis of geriatric diseases, where the substance is for detection expression, activity and / or gene modification used by sgk3. Regarding the candidate Substances are referred to the previous description.

According to the invention also several of the above-described substances for prophylaxis and / or Treatment and / or diagnosis of geriatric diseases become.

The existing and other features of the invention will become apparent the following description of preferred embodiments in conjunction with the subclaims and the figures. Here, the individual features for each be realized in one or more in combination with each other.

In show the pictures:

1 : the Kaplan-Meyer representation for the survival of klotho-deficient mice (klotho ^{- / -} ) as well as of mice in which in addition to the Klotho gene also the gene for sgk1 was switched off (klotho ^{- / -} / sgk1 ^{- / -} )

2 : the Kaplan-Meyer representation for the survival of klotho-deficient mice (klotho ^{- / -} ) as well as of mice, in which in addition to the Klotho gene the gene for sgk3 was switched off (klotho ^{- / -} / sgk3 ^{- / -} ) ,

figure description

In 1 is a Kaplan-Meyer estimate for the survival of klotho-deficient mice (klotho ^{- / -} ) as well as klotho and sgk1-deficient mice (klotho ^{- / -} / sgk1 ^{- / -} ) shown graphically. The ordinate of the graph shows the survival rate of the examined mice in percent. The abscissa shows the survival time or the age of the mice in days. In the mice either Klotho alone (klotho ^{- / -} ) or Klotho and sgk1 (klotho ^{- / -} / sgk1 ^{- / -} ) were genetically switched off. The solid line graph shows the survival rate of klotho-deficient mice (klotho ^{- / -} ) as a function of age. In contrast, the dashed line graph shows the survival rate of klotho and sgk1-deficient mice (klotho ^{- / -} / sgk1 ^{- / -} ), depending on their age. The graph clearly shows that the knockout of sgk1 does not delay the aging process and therefore does not prolong the life span of the mice studied.

In 2 the Kaplan-Meyer estimation for the survival of klotho-deficient mice (klotho ^{- / -} ) and of klotho and sgk3-deficient mice (klotho ^{- / -} / sgk3 ^{- / -} ) is shown graphically. The ordinate represents the survival rate of the examined mice in percent. The abscissa shows the survival time or the age of the mice in days. In the mice either Klotho alone (klotho ^{- / -} ) or Klotho and sgk1 (klotho ^{- / -} / sgk3 ^{- / -} ) were genetically switched off. The solid line graph shows the survival rate of klotho-deficient mice (klotho ^{- / -} ) as a function of age. In contrast, the dashed line graph shows the survival rate of klotho and sgk3-deficient mice (klotho ^{- / -} / sgk3 ^{- / -} ) as a function of age. The graph clearly shows that switching off or knock-out of sgk3 in klotho-deficient mice leads to a strong retardation of biological aging and thus to a delay life span leads. Thus, in the case of a predominant proportion of the experimental animals, a doubling of the life span or expectation was found.

QUOTES INCLUDE IN THE DESCRIPTION

This list The documents listed by the applicant have been automated generated and is solely for better information recorded by the reader. The list is not part of the German Patent or utility model application. The DPMA takes over no liability for any errors or omissions.

Cited patent literature

• WO 02/17893 A2 [0006]
• WO 2007/093264 A1 [0021, 0021]

Cited non-patent literature

• - Webster MK, Goya L, Firestone GL. J. Biol. Chem. 268 (16): 11482-11485, 1993 [0002]
• - Webster MK, Goya L, Ge Y, Maiyar AC, Firestone GL. Mol. Cell. Biol. 13 (4): 2031-2040, 1993 [0002]
• - Waldegger S, Barth P, Raber G, Lang F. Proc. Natl. Acad. Sci. USA 94: 4440-4445, 1997 [0002]
• - Chen SY, Bhargava A, Mastroberardino L, Meijer OC, Wang J, Buse P, Firestone GL, Verrey F, Pearce D. Proc. Natl. Acad. Sci. USA 96: 2514-2519, 1999 [0002]
• - Naray-Fejes-Toth A, Canessa C, Cleaveland ES, Aldrich G, Fejes-Toth GJ Biol. Chem. 274: 16973-16978, 1999 [0002]
• - Warnock DG. Kidney Ind. 53 (1): 1824, 1998 [0002]
• - Waldegger S, Barth P, Raber G, Lang F. Proc. Natl. Acad. Sci. USA 94: 4440-4445, 1997 [0003]
• - Pongs O. Physiol. Rev. 72: 69-88, 1992 [0003]
• - Cahalan MD and Chandy KG. Cur. Opin. Biotech. 8 (6): 749-756, 1997 [0003]
• Szabo I, Gulbins E, Apple H, Zhan X, Barth P, Busch AE, Schlottmann K, Pongs O, Lang FJ Biol. Chem. 271: 20465-20469, 1999 [0003]
• Lang F, Szabo I, Lepple-Wienhues A, Siemen D, Gulbins E. Newes Physiol. Sci. 14: 194-200, 1999 [0003]
• - Cahalan MD and Chandy KG, Cur. Opin. Biotech. 8 (6): 749-756, 1997 [0003]
• Kobayashi T, Deak M, Morrice N, Cohen P. Biochem. J. 344: 189-197, 1999 [0004]
• McCormick et al., 2004 [0005]
• - Alonso L, Okada H, Pasolli HA, Wakeham A, You T, Mak TW, Fox E: Sgk3 left growth factor signaling to progenitor cells in the hair follicle. J Cell Biol 2005; 170: 559-570 [0005]
• - Sandu C, Rexhepaj R, Grahammer F, McCormick JA, Henke G, Palmada M, Nammi S, Lang U, Butcher M, Just L, Skutella T, Dawson K, Wang J, Pearce D, Lang F: Decreased intestinal glucose transport in the sgk3-knockout mouse Pflugers Arch 2005; 451: 437-444 [0005]
• Lang UE, Wolfer DP, Grahammer F, Strutz-Seebohm N, Seebohm G, Lipp HP, McCormick JA, Hellweg R, Dawson K, Wang J, Pearce D, Lang F: Reduced locomotion in the serum and glucocorticoid inducible kinase 3 knockout mouse. Behav Brain Res 2006b; 167: 75-86 [0005]
• - Takeshita K, Fujimori T, Kurotaki Y, Honjo H, Tsujikawa H, Yasui K, Lee JK, Kamiya K, Kitaichi K, Yamamoto K, Ito M, Kondo T, lino S, Inden Y, Hirai M, Murohara T, Kodama I, Nabeshima Y: Sinoatrial node dysfunction and early death of mice with a defect of clot gene expression. Circulation 2004; 109: 1776-1782 [0010]
• - Tsujikawa H, Kurotaki Y, Fujimori T, Fukuda K, Nabeshima Y: Klotho, a gene related to a syndrome resembling human premature aging, functions in a negative regulatory circuit of vitamin D endocrine system. Mol Endocrinol 2003; 17: 2393-2403 [0010]
• - Imura A, Ivano A, Tohyama O, Tsuji Y, Nozaki K, Hashimoto N, Fujimori T, Nabeshima Y: Secreted Klotho Protein in Sera and CSF: implication for post-translational cleavage in release of Klotho protein from cell membrane. FEBS Lett 2004; 565: 143-147 [0010]
• - Kuro-o M, Matsumura Y, Aizawa H, Kawaguchi H, Suga T, Utsugi T, Ohyama Y, Kurabayashi M, Kaname T, Kume E; Iwasaki H, lida A, Shiraki-lida T, Nishikawa S, Nagai R, Nabeshima YI: mutation of mouse klotho gene leads to a syndrome resembling aging. Nature 1997; 390: 45-51 [0010]
• - Kurosu H, Yamamoto M, Clark JD, Pastor JV, Nandi A, Gurnani P, Mc-Guinness OP, Chikuda H, Yamaguchi M, Kawaguchi H, Shimomura I, Takayama Y, Heart J, Kahn CR, Rose Leaf KP, Kuro o M: Suppression of aging in mice by the hormone Klotho. Science 2005; 309: 1829-1833 [0010]
• - Aizawa H, Saito Y, Nakamura T, Inoue M, Imanari T, Ohyama Y, Matsumura Y, Masuda H, Oba S, Mise N, Kimura K, Hasegawa A, Kurabayashi M, Kuro-o M, Nabeshima Y, Nagai R : Downregulation of the Klotho gene in the kidney under sustained circulatory stress in rats. Biochem Biophys Res Commun 1998; 249: 865-871 [0010]
• - Kuro-o M: Klotho as a regulator of oxidative stress and senescence. Biol Chem 2008; 389: 233-241 [0010]
• - Lanske B, Razzaque MS: Premature aging in klotho mutant mice: cause or consequence? Aging res Rev 2007; 6: 73-79 [0010]
• - Yamato Y, Yamagishi T, Nakamura T, Ohyama Y, Aizawa H, Suga T, Matsumura Y, Masuda H, Kurabayashi M, Kuro-o M, Nabeshima Y, Nagai R: Klotho protein protects against endothelial dysfunction. Biochem Biophys Res Commun 1998; 248: 324-329 [0010]
• - Suga T, Kurabayashi M, Sando Y, Ohyama Y, Maeno T, Maeno Y, Aizawa H, Matsumura Y, Kuwaki T, Kuro O, Nabeshima Y, Nagai R: Disruption of the Klotho gene causes pulmonary emphysema in mice. Defect in maintenance of pulmonary integrity during postnatal life. On J Respir Cell Mol Biol 2000; 22: 26-33 [0010]
• - Arking DE, Krebsova A, Macek M, Sr., Macek M, Jr., Arking A, Mian IS, Fried L, Hamosh A, Dey S, Mclntosh I, Dietz HC: Association of human aging with a functional variant of klotho , Proc Natl Acad Sci USA 2002; 99: 856-861 [0010]
• - Arking DE, Becker DM, Yanek LR, Fallin D, Judge DP, Moy TF, Becker LC, Dietz HC: KLOTHO allele status and the risk of early-onset occult coronary artery disease. At the J Hum Genet 2003; 72: 1154-1161 [0010]
• - Freathy RM, Weedon MN, Melzer D, Shields B, Hitman GA, Walker M, McCarthy MI, Hattersly AT, Frayling TM: The functional "KL-VS" variant of KLOTHO is not associated with type 2 diabetes in 5028 UK Caucasians. BMC Med Genet 2006; 7:51 [0010]
• - Imamura A, Okumura K, Ogawa Y, Murakami R, Torigoe M, Numaguchi Y, Murohara T: Klotho gene polymorphism may be a genetic risk factor for atherosclerotic coronary artery disease but not for vasospastic angina in Japanese. Clin Chim Acta 2006; 371: 66-70 [0010]
• Low AF, O'Donnell CJ, Kathiresan S, Everett B, Chae CU, Shaw SY, Ellinor PT, Macrae CA: Aging syndrome genes and premature coronary artery disease. BMC Med Genet 2005; 6:38 [0010]
• Maniatis T, Fritsch EF, Sambruk J. Coldspring Harbor, NY: Coldspring Harbor Labrator 1996 [0016]
• - Leonard D, Davis PhD, Michael W, Kuehl Md, James F, Battey MD. McCraw-Hill Professional Publishing, 1995 [0016]

Claims (15)

Substance for the prophylaxis and / or treatment of geriatric diseases, characterized in that the substance inhibits the expression and / or activity of sgk3. Substance according to claim 1, characterized that the substance is an anti-sgk3 substance, preferably an antibody directed against sgk3, is. Substance according to claim 1 or 2, characterized that the substance is resistant to activators, inhibitors, regulators, biological Precursors, variants, especially isoforms, of sgk3 is directed. Substance according to one of the preceding claims, characterized in that the substance is a polynucleotide, which encodes a peptide, preferably a polypeptide, wherein this peptide the expression and / or activity of sgk3 inhibited. Substance according to one of the preceding claims, characterized in that the substance is a low molecular weight Compound, preferably having a molecular weight (MW) <1000, is. Substance according to one of the preceding claims, characterized in that the substance is a kinase inhibitor. Substance according to one of the preceding claims, characterized in that the substance from the group of congestion rosporin, Chelerythrine, SB 203580, SB 202190 and derivatives derived therefrom is selected. Substance according to one of the preceding claims, characterized in that the substance is a compound of the formula I:

wherein R ¹ , R ^{2 are} each independently H, CHO or acetyl, R ³ , R ⁴ , R ⁵ , R ⁶ , R ⁷ , R ⁸ , R ⁹ , R ¹⁰ , R ^{11 are} each independently H, A, OSO ₂ A, Hal, NO _2, OR ^12, N (R ¹²⁾ _2, CN, O-COA, - [C (R ¹²⁾ _{2] n} COOR ^12, O- [C (R ¹²⁾ _{2] 0} COOR ^12, SO ₃ H, - [C (R ¹² ) ₂ ] _n Ar, -CO-Ar, O- [C (R ¹² ) ₂ ] _n Ar, - [C (R ¹² ) ₂ ] _n Het, - [C ( R ¹² ) ₂ ] _n O≡CH, O- [C (R ¹² ) ₂ ] _n C≡CH, - [C (R ¹² ) ₂ ] _n CON (R ¹² ) ₂ , - [C (R ¹² ) ₂ ] _n CONR ¹² N (R ¹² ) ₂ , O- [C (R ¹² ) ₂ ] _n CON (R ¹² ) ₂ , O- [C (R ¹² ) ₂ ] _o CONR ¹² N (R ¹² ) ₂ , NR ¹² COA, NR ¹² CON (R ¹² ) ₂ , NR ¹² SO ₂ A, N (SO ₂ A) ₂ , COR ¹² , S (O) _m Ar, SO ₂ NR ¹² or S (O) _m A, R ³ and R ⁴ together also form CH = CH-CH = CH, R ³ and R ⁴ , R ⁷ and R ⁸ or R ⁸ and R ⁹ together also alkylene having 3, 4 or 5 C atoms, wherein one or two CH ₂ - A may be replaced by oxygen, A is unbranched or branched alkyl having 1-6 C atoms, wherein 1-7 H atoms may be replaced by F, or alkyl having 3-7 C atoms, Ar u n-substituted or mono-, di- or trisubstituted by Hal, A, OR ¹² , N (R ¹² ) ₂ , NO ₂ , CN, phenyl, CON (R ¹² ) ₂ , NR ¹² COA, NR ¹² CON (R ¹² ) ₂ , NR ¹² SO ₂ A, COR ¹² , SO ₂ N (R ¹² ) ₂ , S (O) _m A, - [C (R ¹² ) ₂ ] _n -COOR ¹² and / or -O [C (R ¹² ) ₂ ] _o -COOR ¹² substituted phenyl, naphthyl or biphenyl, Het a mono- or binuclear saturated, unsaturated or aromatic heterocycle having 1 to 4 N, O and / or S atoms, the one, two or three times by Hal, A, OR ¹² , N (R ¹² ) ₂ , NO ₂ , CN, COOR ¹² , CON (R ¹² ) ₂ , NR ¹² COA, NR ¹² SO ₂ A, COR ¹² , SO ₂ NR ¹² , S (O ) _m A, = S, = NR ¹² and / or = O (carbonyl oxygen) may be substituted, R ^{12 is} H or A, Hal is F, Cl, Br or I, m is 0, 1 or 2, n is 0, 1, 2 or 3, 0 is 1, 2 or 3, and their pharmaceutically usable derivatives, salts, solvates and stereoisomers , including mixtures thereof in all proportions. Substance for the diagnosis of geriatric diseases, thereby characterized in that the substance is for detection of expression, activity and / or a gene modification of sgk3. Substance according to claim 9, characterized that the substance is an anti-sgk3 substance, especially an antibody directed against sgk3, is. Substance according to one of the preceding claims, characterized in that the age diseases from the group Arteriosclerosis, pulmonary emphysema, skin atrophy, muscle weakness, Immune deficiency, infertility, kyphosis, calcification of connective tissue, heart attack, stroke, osteoarthritis, dementia, diabetes mellitus, cataract, cancer, osteoporosis, Parkinson's disease, chronic senile rhinitis, atrial fibrillation and incontinence are. Diagnosis kit comprising at least one substance for Detection of expression and / or activity and / or a Gene modification of sgk3, for the diagnosis of geriatric diseases. Method for the diagnosis of geriatric diseases, wherein the expression and / or activity and / or a gene modification of sgk3 by detection in a body sample of a patient with antibodies against sgk3, with oligonucleotides, with which in a polymerase chain creation (PCR) certain DNA sections of sgk3 are amplified and / or polynucleotides containing DNA or hybridize sgk3 mRNA under stringent conditions, is detected. Use of a substance according to one of the claims 1 to 8 or 11 for the manufacture of a medicament for prophylaxis and / or treatment of geriatric diseases. Use of a substance according to one of the claims 9 to 11 for the preparation of a diagnostic means for the diagnosis of Age-related diseases.