DE10356076A1 - Pharmaceutical preparation, for the treatment of obesity and type II diabetes, uses associated proteins to inhibit or encourage the deposit of lipid molecules in adipocytes - Google Patents

Abstract

A pharmaceutical preparation, for the prevention and/or treatment of obesity, using a polypeptide with a sequence identity no. 1,3,5,7,9,11,13,15,17,19,21, or 23; or a heterotrimer of alpha and beta lymphotoxin; or a nucleic acid coded for a selected polypeptide; or one or more nucleic acids coded for a heterotrimer of alpha and beta lymphotoxin; and a functional variant of their molecules, is new. ACTIVITY : Anorectic; Antidiabetic; Immunomodulator. No biological data is given. MECHANISM OF ACTION : None given.

Description

The The invention relates to proteins associated with obesity their use in the context of pharmaceutical compositions or diagnostics. Furthermore, the present invention relates to Use of inhibitors of these proteins for the treatment of type II diabetes.

obesity (Obesity) and Type II diabetes are two of the most important Metabolic diseases just increasing in the Western world pose a serious problem. Furthermore, especially in type II diabetes meanwhile observe tendencies to a pandemic.

obesity is a condition caused by excessive accumulation of fatty tissue in the body is marked. Essential for the formation of fatty tissue and thus the expression of an obese phenotype the adipocytes, i. those cells which are the fatty tissue of the body form. After recent understanding Obesity is a chronic health disorder that affects both on one Polygenetic predisposition is also due to environmental factors is conditional and accompanied by a high degree of concomitant and sequential morbidity. Therefore, for Obesity a long-term treatment and care concept essential.

The known in the art treatment methods for obesity restrict essentially based on a strict diet, coupled with increased exercise. A known therapy is so far only about the norepinephrine serotonin receptor tophak inhibitor "sibutramine" (BASF, Ludwigshafen) or the lipase inhibitor "Xenical" (Roche, Basel, Switzerland) possible. The field of application of these drugs, however, is due to the occurring Side effects / complications restricted. For example, sibutramine is not in hypertension - one typical complication of obesity - can be used. In addition, the Disease can only be treated symptomatically.

At the Diabetes mellitus (also commonly known as "diabetes mellitus") acts it is a chronic metabolic disease caused by a increased Blood sugar level is marked. Different causes the disease and also require different disease forms the distinction between two types, type I and type II diabetes.

Of the Type I diabetes (formerly: Juvenile diabetes) usually begins in adolescence and arises through an immunological destruction the islet cells of the pancreas (= pancreas). These islet cells produce the hormone insulin that works for the recovery of glucose from food is responsible. By the destruction the islet cells there is an absolute insulin deficiency. The glucose from food can no longer be broken down, and blood sugar levels increases. The treatment of type I diabetes is by administration of insulin.

Of the Type II diabetes (formerly: Adult or adult onset diabetes) usually develops in the higher Age. He is characterized in that the body cells, where insulin is supposed to work, not enough insulin react. This is due to a resistance of adipocytes to insulin due. Interestingly, are mostly the older, Mature adipocytes resistant while younger Adipocytes usually have no insulin resistance.

A such insulin resistance is due to persistently high blood sugar and insulin levels as e.g. to watch in obese people are. The therapy of type II diabetes takes place gradually: first is having a diet trying to lower the blood sugar level in general. Are the dietary measures to Treatment inadequate will lower blood sugar in the episode Medication and in the advanced stage also insulin administered.

The However, administration of insulin may be common in patients with type II diabetes due to insulin resistance in the target tissues to hyperinsulinemia (increased insulin levels in plasma, serum). Therapy is then no longer possible with insulin (Curr Diab Rep. 2003 Oct; 3 (5): 378-85).

There are attempts to significantly delay or even prevent the necessary administration of insulin by administering immunosuppressive therapy as long as the islet cells still produce their own insulin. Also, administration of nicotinamide in conjunction with intensified insulin therapy may maintain beta cell function for up to two years after diagnosis. However, such therapeutic approaches are all still in the experimental stage. The transplantation of islets or the ge whole pancreas is not yet very successful.

Also the therapeutic use of TNF-alpha, a known differentiation inhibitor for fat cells, can not. Although could for TNF-alpha demonstrated an improvement in insulin resistance in the animal model An effect in the human model could be in clinical trials however, can not be proved. In addition, TNF-alpha has toxic Side effects on (Int J Exp Diabesity Res. 2003 Apr-Jun; 4 (2)): 66-71).

The previous therapeutic approaches done only symptomatically either via altered diet and / or for example the administration of PPAR-gamma agonists (as ligands for PPAR-gamma These activate the PPAR-gamma transcription factor, which differentiates of adipocytes or activation of lipid metabolism in mature Adipocytes can cause).

One therapeutic use of transcription factors such as PPAR-gamma for therapy is not possible since the factors are introduced directly into the target cell or the nucleus Need to become. In addition, the binding to the DNA can only by mediation of other cofactors respectively.

One Significant disadvantage of symptomatic treatment is also the occurrence of sequelae, such as cardiovascular complications, diabetic nephropathies, neuropathies or retinopathies, the additionally need to be treated.

In order to are not promising methods of treatment in the prior art as well as Obesity as well as for Known type II diabetes.

task Thus, it is the present invention to provide means for the treatment of To provide obesity or type II diabetes.

According to the invention Task first solved by the use of a) a polypeptide having a sequence according to SEQ ID No: 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21 or 23, b) encoding a nucleic acid for a Polypeptide according to a), or c) a functional variant of the molecules according to a) or b), for the production a pharmaceutical composition for the treatment of obesity.

in the Within the scope of the invention, it has surprisingly been found that said polypeptides or nucleic acids encoding them have a essential role in the expression to play an obesity. More precisely, it has surprisingly found that the said proteins or nucleic acids in capable of differentiating or functioning of adipocytes to inhibit sustainably. this makes possible For the first time a treatment of adiposity at the molecular level.

The SEQ ID numbers listed in claim 1 relate to the following proteins and nucleic acids:

TNFSF-12 (Tweak) is a cell surface associated Type II membrane protein, originally as a member of the Tumor Necrosis Factor (TNF) superfamily has been. (a more detailed Review can be found in Wiley et al., 2003, Cytokine & Growth Factor Reviews, 14: 241-249).

One small biologically active part can be as soluble protein with a length of about 157 amino acids in the extracellular Millieu be split off.

The TNFSF-12 mRNA is present in a variety of tissues and cell types, but mainly expressed in leukocytes. Striking is the difference between mRNA expression and protein expression at the cell surface. Expression of TNFSF-12 at the cell surface could only be confirmed on monocytes after IFN-gamma treatment.

In the literature so far only one receptor for TNFSF-12 is known, which binds tweak with physiological activity. It is described as "TweakR" or "fibroblast growth factor inducible 14" (FN14). This receptor is the smallest member of the group of TNF receptors. There are no other ligands known which bind to TweakR, nor is it known that Tweak itself binds to other known TNF receptors.

For TNFSF-12 are so far in the literature multiple biological activities such as Induction of angiogenesis, inflammatory cytokines and under special conditions (e.g., costimulation with IFN-gamma) stimulation described by apoptosis. In the context of the present invention, surprisingly an inhibition of adipocyte differentiation or inhibition of lipid storage in adipocytes.

TNFSF-12V1 differs from TNFSF-12V1 * by the absence of an amino acid in the memebrane part of the protein (see Example 1).

TNFSF-14 ("Light") is an integral Class II membrane protein of tumor necrosis factor (TNF) - ligand family (a more detailed Review can be found in Granger and Ricert, 2003, Cytokine & Growth Factor Reviews, 14; 289-296). It has a length of 240 amino acids up and contains an N-terminal cytosolic domain, a type II transmembrane domain as well a C-terminal receptor binding domain. The soluble protein is 167 amino acids long.

The Protein is a ligand for TNFRSSFI4, a member of the TNF receptor superfamily, also known as "herpes virus entry mediator "(HVEM) is known. Other receptors are lymphotoxin-beta receptor and DcR3.

So far has been implicated in immunological processes (T-cell proliferation, Cytokine secretion, arteriosclerosis, "graft versus host disease "as well shown an association with apoptotic processes: the protein stimulates T cell proliferation and triggers apoptosis of different tumor cells. Also known is prevention of TNF-alpha mediated apoptosis in hepatocytes.

expression TNFSF-14 is found on activated T cells, NK and dendritic Cells.

A Inhibition of adipocyte differentiation or inhibition of lipid storage in adipocytes was first in the context of the present invention to be shown.

For TNFSF-12 / TNFSF-14 it could be shown that the inhibitory effect is reversible. This could be demonstrated by removing the protein after day four. this makes possible the 3T3-L1 cells apparently re-enter the cell cycle or in the differentiation process. It was also found that the Effect is not cytotoxic, since the reversibility of the inhibitory Effects shows that the cells are not affected by the preceding inhibition Get damaged. This is also done by microscopic measurement of Living cell count confirmed.

The Secreted Cytokine Cardiotrophin has no canonical signal peptide on. It is mainly used in Heart, brain, lungs and liver are expressed and is called "survival factor "known. A connection with adipocyte differentiation or inhibition of Lipid storage in adipocytes is not shown in the prior art.

Calsyntenin-1 is a type I membrane protein. It consists of a large extracellular fragment, the two "cadherin-like repeats "and a contains small cytoplasmic segment that is capable of calcium ions to bind. The protein is proteolytically cleaved in the extracellular domain. In the literature is about a possible Involvement of the cytoplasmic part in the regulation of Calcium signals in the CNS speculated. The extracellular part has not been an independent one yet Assigned function. An inhibition of adipocyte differentiation or inhibition of lipid storage in adipocytes could stand the technique is not shown.

FGF-16 is a member of the "fibroblast growth factor family. These proteins have broad mitogenic and cell survival activity in a variety of biological processes, e.g. Embryo development, Cell growth, morphogenesis, tissue regeneration, tumor growth as well invasion involved. Although the protein is secreted, no Signal peptide known. FGF-16 was used as a growth factor for brown Adipocytes are described, but not as an inhibitor of differentiation of preadipocytes.

GH-2 ( "Growth Hormone 2 ") is a member of the somatrotropin / prolactin family of hormones that play an important role in growth control. An inhibition Adipocyte differentiation or inhibition of lipid storage in adipocytes by GH-2 could not be demonstrated in the prior art become.

at BM045 is an unspecified bone marrow protein. A function is not known yet.

GK001 has a transmembrane domain and a signal peptide. A function is not known yet. Due to the predicted transmembrane domain, it is possible for a secreted Protein fragment by extracellular / juxtamembrane proteolytic Fission is generated.

Endothelin-2 is a member of the endothelin family (ET-1, ET-2, ET-3), which from larger preproproteins about 21 amino acids generate long peptides by proteolytic cleavage. The extracellular protein contains two Endothelin domains and becomes mainly expressed in the kidney. Well-known functions are vasoconstriction and chemotaxis. An inhibition of adipocyte differentiation or Inhibition of lipid storage in adipocytes by endothelin-2 could not shown in the prior art.

As can be seen from Examples 4 and 5 and 1 - 5 results, said proteins are able to inhibit lipid storage in adipocytes.

As stated above The invention also relates to the use of functional variants those listed under a) or b) above molecules for the preparation of a pharmaceutical composition for treatment of obesity.

"Functional Variant "one Polypeptide according to the invention relates to a polypeptide which is substantially the biological function or functions of the corresponding protein Has. In the case of the present proteins this may be the ability act, the lipid storage in suitable cells, in particular 3T3 cells, to inhibit. A test to determine this activity is in Examples 2-5 shown.

Under a "functional Variant "one nucleic acid is understood according to the invention a nucleic acid, which is also an inventively used Polypeptide encoded.

Of the Expression "functional Variant "one Polypeptide or a nucleic acid preferably refers to polypeptides or nucleic acids a sequence homology, in particular a sequence identity of about at least 25%, preferably about 40%, especially about 60%, especially preferably about 70%, most preferably about 80%, in particular about 90% and most preferably 98% with the polypeptide. Such Variants are, for example, homologous polypeptides derived from others Come from organisms. Other examples of variants are polypeptides, which are encoded by different alleles of a gene, polypeptides from different individuals, from different organs of an organism or of different stages of development. Close functional variants preferably also naturally occurring Mutations, in particular mutations, which in quantitative Way the activity the peptides change, which are encoded by these sequences. Furthermore, such Variants preferably from a differential splicing of the coding Genes result.

In a particularly preferred embodiment includes the term "functional Variant "Derivatives with single nucleotide polymorphism (SNP) polymorphisms.

"Sequence identity" refers to the degree of identity (% Identity) two sequences, which in the case of polypeptides, for example by BLASTP 2.2.5 and in the case of nucleic acids for example by BLASTN 2.2.6 can be determined, with the low complexity filter (low complexity filter) is turned on and BLOSUM 62 is (Altschul et al., 1997, Nucleic Acids Res., 25: 3389-3402).

"Sequence homology" refers to the similarity (% Positive) of two nucleotide or polypeptide sequences for example by means of BLASTN 2.2.6 or BLASTP 2.0.1, wherein the Filter is turned on and BLOSUM 62 (BlastP) is (Altschul et al., 1997, Nucleic Acids Res., 25: 3389-3402).

Nucleic acids that functional variants can be coded by using appropriate gene sequences be isolated to identify homologs, using procedures which are known to the person skilled in the art, e.g. PCR amplification or hybridization under stringent conditions (e.g., 60 ° C in 2.5x SSC buffer, followed by various washing steps at room temperature) with suitable samples, for example, from the corresponding gene sequences descended.

In a preferred embodiment, the functional variant is a soluble protein. Further Preferably, the functional variant is a soluble protein that lacks the transmembrane domain. Preferably, this means that the fragment containing the transmembrane domain has been cleaved off. In the case of type II proteins (TNFSF-12, TNFSF-14), the N-terminal fragment contains the transmembrane domain. In type I proteins (all other proteins used according to the invention), the C-terminal fragment contains the transmembrane domain. In a further preferred embodiment, a soluble extracellular fragment which is N-terminal (type II proteins) or C-terminal (type I proteins) shortened by further amino acids is used. More preferably, an N-terminal fragment (type II proteins) or a C-terminal fragment (type I proteins), for example a signal sequence, may also be cleaved off.

In a particularly preferred embodiment the proteins are protein fragments according to Table 1.

According to one preferred embodiment obesity is an uncomplicated obesity. An uncomplicated Obesity is characterized by the lack of insulin resistance pronounced is.

According to one another preferred embodiment act the invention used Polypeptides or nucleic acids in that it prevents the formation of terminally differentiated adipocytes inhibit.

there the formation of terminally differentiated adipocytes can be inhibited through promotion the proliferation of preadipocytes, by inhibiting differentiation into terminally differentiated adipocytes, in particular by inhibiting the transcription signal pathway for differentiation, or by promotion of cell death, especially apoptosis in preadipocytes.

The For example, proliferation can be by cell count, cell cycle analysis in the FACS or determination of the metabolic activity of the cells done, which measured as enzymatic turnover of mitochondria (e.g., by Alamar Blue detection).

The inhibition of differentiation to terminally differentiated adipocytes can be done by measuring the lipid incorporation (eg by staining with NileRed). With reporter gene assays such as PPAR-gamma, the inhibition of the transcription signal pathway can be measured. Apoptosis is determined, for example, by CDD + Assay (Cell Death Detection ELISA PLUS, Roche Diagnostics GmbH, Mannheim, Germany), DiOC6 Assay, AnnexinV Assay or CaspaTag ^™ Assay (CaspaTag ^™ Caspase-3 (DEVD) Activity Kit, Intergen Company, Purchase, NY, USA).

alternative can the invention used Polypeptides or nucleic acids by acting as a function of the terminal differentiated Inhibit adipocytes. This can be done, for example, by inhibiting the Lipogenesis or through a promotion the lipolysis are effected.

By coloring with lipophilic dyes such as e.g. Nile Red allows lipid incorporation into mature Prove adipocytes.

As already stated above, is for the molecules BM045 and GK001 so far described no function in the prior art Service. Thus, the invention also relates to a pharmaceutical composition containing a) a polypeptide having a sequence according to SEQ ID NO: 21 or 23, b) a nucleic acid coding for a polypeptide according to a) or c) a functional variant of the molecules according to a) or b).

The Invention further relates to a method for treating a patient, in which the patient has an effective amount of one or more the molecules defined above is administered. Preferably, the patient suffers from obesity. For this inventive method all apply above for the use according to the invention listed Embodiments.

As already described above the invention found Factors besides inhibition of differentiation of preadipocytes to adipocytes an inhibition of the function of mature or maturing Adipocytes by inhibiting lipogenesis or promotion lipolysis. In inhibition of this inhibitor of differentiation thus form by differentiation or modulation of the function of mature differentiated adipocytes, new young adipocytes that have no insulin resistance. The ratio of insulin-resistant adipocytes is thus shifted in favor of insulin-sensitive adipocytes.

Thereby is the first time a causal therapy approach of type II diabetes directly possible in fatty tissue, the then prevent or delay the development of secondary diseases can alleviate.

In order to The invention also relates to the use of an inhibitor of a Polypeptide having a sequence according to SEQ ID NO: 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21 or 23 or a functional variant of which for the preparation of a pharmaceutical composition for Treatment of diabetes type II, diabetes type II related diseases cardiovascular complications, arteriosclerosis, dyslipidemia, diabetic nephropathy, neuropathy, retinopathy, hyperglycemia, insulin resistance, Hyperinsulinemia or High blood pressure), cachexie or HARS.

cachexia is characterized by a decrease in fatty tissue or by an inhibition the formation of new fatty tissue. On the one hand, HARS ("HIV-associated-adipose-syndrome") occurs to a decrease of subcutaneous fat tissue deposits and on the other hand to an enlargement more specific smaller adipose tissue depots ("bullskin"). These changes are at an elevated level Expression or activity the invention used Attributed proteins. By The inhibition of these proteins may further degrade the fat tissue deposits prevents or build up these depots are achieved. At HARS a buildup of subcutaneous fat tissue deposits can be achieved.

According to one preferred embodiment the inhibitor is selected from the group consisting of proteins, peptides, antibodies, decoy receptors, Anticalins, Affilins, Low Molecular Weight Substances (LMWs), Antisense RNA, siRNA, aptamers and proteases.

By "inhibiting" or "inhibiting" according to the invention the complete or partially suppress understood a biological function.

Corresponding In the present invention, the term "inhibitor" refers to a biochemical or biochemical chemical compound, preferably the action of the invention described Proteins, i. for example, the inhibition of fat storage itself inhibited or reduced. This can be done, for example Suppression of the expression of the corresponding gene or the activity of the respective Protein can be achieved. Expression of the gene can be by RT-PCR or Western blot analysis. According to another preferred Ausfürungsform the inhibitor used causes inhibition of expression or activity increased proteins of the invention.

According to the invention is a Activator connect with the opposite function.

Examples such inhibitors are binding proteins or binding peptides, the against the molecule directed, in particular against the active site of the molecule, and nucleic acids, which are directed against the corresponding gene.

"Decoy receptors" are non-membranous receptors without transmembrane domain to understand that bind to ligands, but due to missing intracellular domains do not allow signal forwarding.

LMWs are molecules, no proteins, peptide antibodies or nucleic acids are, and which has a molecular weight of less than 5000 Da, preferably less than 2000 Da, more preferably less than 1000 Da, on most preferably less than 500 Da. Such LMWs can be used in identified as "high throughput" method which are carried out starting from according to molecular banks. Such methods are known in the art.

Of the The term "binding protein" or "binding peptide" refers to a class of proteins or peptides that bind the molecule or inhibit, including, without restriction, polyclonal or monoclonal antibodies, antibody fragments and protein scaffolds directed against these proteins.

The method for producing an antibody or antibody fragment is carried out by methods known to those skilled in the art, for example by immunizing a mammal, for example a rabbit, with the corresponding peptide, where appropriate adjuvants, for example Freund's adjuvant and / or aluminum hydroxide gels, can be used (see, for example, Diamond, BA et al. (1981) The New England Journal of Medicine: 1344-1349). The polyclonal antibodies that are formed in the animal as a result of an immunological reaction can subsequently be isolated from the blood using well-known methods and then purified, for example, by column chromatography. Monoclonal antibodies may be prepared, for example, in accordance with the known method of Winter & Milstein (Winter, G. & Milstein, C. (1991) Nature, 349, 293-299).

According to the present invention, the term antibody and antibody fragment is also understood to include antibodies or antigen-binding portions thereof which have been recombinantly produced and modified as necessary, for example, chimeric antibodies, humanized antibodies, multifunctional antibodies, bispecific or oligospecific antibodies, single stranded antibodies and F (ab) or F (ab) ₂ fragments (see for example EP-B1 368 684, US 4,816,567 . US 4,816,397 WO 88/01649, WO 93/06213 or WO 98/24884).

When Alternative to classic antibodies it is also possible protein scaffolds (protein scaffolds) to use against the respective molecule, for example Anticalins based on lipocalin (Beste et al. (1999) Proc. Natl. Acad. Sci. USA, 96, 1898-1903). The natural ones Ligand binding sites of lipocalins, such as the retinol-binding protein or the bilin-binding protein can be altered, for example using a combinatorial Protein design "approach, in such a way that they bind selected haptens, (Skerra, 2000, Biochim. Biophys. Acta, 1482, 337-50). From other known protein scaffolds are known to be alternatives to antibodies (Skerra (2000) J. Mol. Recognit, 13, 167-287).

Of the Expression "Nucleic acids directed against the corresponding gene "refers on double-stranded or single-stranded DNA or RNA, for example the expression of the respective gene or the activity of the respective molecules inhibits and closes without restriction Antisense nucleic acids, Aptamers, siRNAs (small interfering RNAs) and ribozymes.

The nucleic acids, e.g. the antisense nucleic acids or siRNAs be chemically synthesized, for example according to the Phosphotriesterverfahren (see, for example, Uhlmann, E. & Peyman, A. (1990) Chemical Reviews, 90, 543-584).

Aptamers are nucleic acids that bind to a polypeptide with high affinity. Aptamers can be obtained by selection methods such as SELEX (see, eg, Jayasena (1999) Clin. Chem., 45, 1628-50; Klug and Famulok (1994) M. Mol. Biol. Rep., 20, 97-107; US 5,582,981 ) are isolated from a large pool of different single-stranded RNA molecules. Aptamers may also be synthesized and selected in their mirror form, for example as the L-ribonucleotide (Nolte et al., 1996) (Nat. Biotechnol., 14, 1116-9; Klussmann et al. (1996) Nat. Biotechnol., 14 , 1112-5). Forms isolated in this manner have the advantage that they are not degraded by naturally occurring ribonucleases and therefore have greater stability.

Nucleic acids can pass through Endonucleases or exonucleases are degraded, in particular by DNases and RNases that can be found in the cell. It is therefore advantageous to nucleic acids too modify it to face it Stabilize degradation, thereby ensuring that a high Concentration of nucleic acid in the cell above a long period of time (Beigelman et al. Nucleic Acids Res., 23: 3989-94; WO 95/11910; WO 98/37240; WO 97/29116). Typically, one can such stabilization by introducing one or more internucleotide phosphorus groups or by insertion one or more non-phosphorus internucleotide analogs become.

suitable modified internucleotides are described in Uhlmann and Peyman (1990), see above, (see also Beigelman et al., (1995) Nucleic Acids Res., 23: 3989-94; WO 95/11910; WO 98/37240; WO 97/29116). Modified internucleotide phosphorus groups, shut down for example, methylphosphonates, phosphorothioates, phosphoramidates, Phosphorodithioates and / or phosphate esters while the non-phosphorus internucleotide analogs for example siloxane bridges, carbonate bridges, Carboxymethyl ester, acetamidate bridges and / or thioether bridges contain. It is also intended that these modifications be the To improve the lifetime of a pharmaceutical composition, used according to the uses of the present invention can be.

The Use of suitable antisense nucleic acids is further disclosed in Zheng and Kemeny (1995) Clin. Exp. Immunol., 100, 380-2; Nellen and Lichtenstein (1993) Trends Biochem. Sci., 18, 419-23; Stein (1992) Leukemia, 6. 697-74; or Yacyshyn, B.R. et al. (1998) Gastroenterology, 114, 1142).

The production and use of siRNAs as an aid to RNA interference in the process of down-regulating or shutting down gene expression is described, for example, in Elbashir, SM et al. (2001) Genes Dev., 15, 188, or Elbashir, SM et al. (2001) Nature, 411, 494. Preferably have siRNAs have a length of less than 30 nucleotides, wherein the identity region of the sense strand of the siRNA preferably comprises at least 19 nucleotides.

ribozymes are also suitable tools for the translation of nucleic acids inhibit because they are able to specifically bind the mRNAs and to cut. They are described, for example, in Amarzguioui et al. (1998) Cell. Mol. Life Sci., 54, 1175-202; Vaish et al. (1998) Nucleic Acids Res., 26, 5237-42; Persidis (1997) Nat. Biotechnol., 15, 921-2; or couture and stinchcomb (1996) Trends Gene., 12, 510-5. A single-stranded DNA or RNA is for the use as antisense oligonucleotide or ribozyme preferred.

Therefore can the nucleic acids described herein are used to obtain the Expression of the respective genes in the cells both in vivo and inhibit or reduce in vitro. That's why they can act as inhibitors in the context of the present invention.

According to one preferred embodiment The inhibitor used acts by the fact that the pharmaceutical Composition the formation of terminal differentiated adipocytes promotes. This can be done, for example, by inhibiting proliferation of preadipocytes, through promotion differentiation to terminally differentiated adipocytes, in particular through promotion the transcription signal pathway of differentiation or by inhibition of cell death, preferably apoptosis in preadipocytes.

there the respective effect can be measured as indicated above.

According to one another preferred embodiment acts according to the invention used Inhibitor in that the pharmaceutical composition the Promotes function of terminal differentiated adipocytes. This For example, by a promotion lipogenesis or by inhibiting lipolysis.

These Effects can measured as described above.

Corresponding a further preferred embodiment The inhibitor used acts by the fact that the pharmaceutical Composition a sensitization of other insulin-dependent non-adipocyte cell types (such as skeletal muscle cells, hepatocytes, islet cells, endothelial cells) causes. Here, the negative regulation of insulin-dependent signaling, those by the proteins according to the invention caused by the inhibitor used.

The Invention further relates to a pharmaceutical composition, containing an inhibitor of a polypeptide having a sequence according to SEQ ID No: 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21 or 23, or a functional one Variant of it.

For all pharmaceutical compositions according to the present invention, the following applies:
For the preparation of the pharmaceutical composition according to the invention, the corresponding molecules are usually formulated with suitable additives or excipients, such as physiological buffer solution, for example sodium chloride solution, demineralized water, stabilizers, such as protease or nuclease inhibitors, preferably aprotinin, ε-aminocaproic acid or pepstatin A or chelating agents such as EDTA, gel formulations such as white petrolatum, low viscosity paraffin or yellow wax, depending on the route of administration.

suitable additional Additives are, for example, detergents, such as Triton X-100 or sodium deoxycholate, but also polyols, such as Polyethylene glycol or glycerin, sugars, such as sucrose or glucose, zwitterionic compounds such as amino acids such as glycine or in particular taurine or betaine or a protein such as bovine or human serum albumin. Detergents, polyols and / or zwitterionic compounds are preferred.

The physiological buffer solution preferably has a pH of about 6.0-8.0, in particular a pH of about 6.8-7.8, in particular a pH of about 7.4, and / or an osmolarity of about 200-400 milliosmol / Liter, preferably from about 290-310 milliosmol / liter. The pH of the drug is generally adjusted using a suitable organic or inorganic buffer such as phosphate buffers, tris (tris (hydroxymethyl) aminomethane), HEPES buffer ([4- (2-hydroxyethyl) piperazino] ethanesulfonic acid) or MOPS buffer (3-morpholino-1-propanesulfonic acid). The choice of the appropriate buffer generally depends on the desired buffer molarity. Phos phat buffer is suitable, for example, for injection or infusion solutions.

Become injection solutions Generally used when only a small amount of a solution or Suspension, for example about 1 to about 20 ml should. infusion solutions are generally used when a larger amount of a solution or Suspension, for example one or more liters, are administered should. As opposed to the infusion solution only a few milliliters in the case of injection solutions are low pH differences or osmotic Pressure of the blood or tissue fluid compared to the injection solution itself not noticeable or play a big role. The dilution of the Formulation prior to use is therefore generally unnecessary. If relatively large Quantities should be administered, the inventive formulation diluted shortly before administration so that about an isotonic solution is obtained. An example an isotonic solution is a 0.9% saline solution. In the case of an infusion, the dilution may be, for example, below Use of sterile water can be achieved while the Administration can be performed for example by a so-called bypass can.

According to the invention is included that subjects corresponding to the present Be treated or diagnosed, preferably mammals and Include people, dead or living.

The Pharmaceutical according to the invention Composition can be administered in different ways, a suitable inhibition of lipid storage or symptoms to cause a type II diabetes.

Basically the effective dose of the weight and condition of each depend on the subject. It is assumed that the skilled person is aware of how a suitable dosage can be determined.

The pharmaceutical composition can be administered in a variety of ways, for example, intramuscularly, subcutaneously, intrathecally, in adipose tissue, percutaneously (dissolved in DMSO), intranasally (inhalation), intravenously or intraperitoneally, or by fusion or gels containing the subject medicament , It is also possible to administer the medicament topically and locally, for example in the form of liposome complexes. Further, the drug may be administered by a transdermal therapeutic system (TTS), which allows for timed delivery of the drug. For example, TTS are off EP 944,398 . EP 916,366 . EP 889 723 or EP 852 493 known. A slower release of the protein, peptide, or antibody is achieved by combination with polymers, a longer half-life by addition of PEG. A combination with chemical agents (eg appetite suppressants, lipase inhibitors) may increase the effect.

Preferably become the molecules at a dose of 1 pg / kg to 15 mg / kg, preferably between 5 pg / kg and 5 mg / kg, and more preferably between 0.2 and 2 mg / kg Patient weight administered. Continuous infusions can also be used, for example by using an osmotic minipump, as described in Heyman et al., Nat. Med., 1999, 5, 1135-152. In In this case, an infusion dose of between 5 and 20 μg / kg / minute, preferably be applied between 7 and 15 pg / kg / minute.

If nucleic acids be administered, so the gene transfer can either by means of naked DNA or done in a vector. Appropriate vectors are known in the art (Barnacal, P. (1998) SRIP's complete Guide to Gene Therapy. PJB Publication) Vectors that are subject to the present Invention can be used, for example, on adenovirus and / or replication-deficient retrovirus-based vectors. Corresponding Techniques are known to the person skilled in the art.

If required can adjustable vectors as described, for example, in Ozawa et al., Annu. Rev. Pharmacol. & Toxicol., 2000, 40, 295-317, be used.

According to a preferred embodiment, the pharmaceutical composition according to the present invention is in a form which allows the contact of the active ingredient with a cell-surface component (non-cellular) (Cardiovasc Pharmacol Ther. 2002 Jul; 7 (3): 171-80). Alternatively, the pharmaceutical composition may be in a form that allows transport of the drug into the cell interior (cell-like). This can be made possible, for example, by packaging the active substance with cell-permeable liposomes. Examples of corresponding liposomes can be found in Curr Med Chem. 2003 Jul; 10 (14): 1279-87 (Cationic liposomes as in vivo delivery vehicles, Templeton NS); Trends Pharmacol Sci. 2003 May; 24 (5): 213-5 (proteins); Green I, Christison R, Voyce CJ, Bundel KR, Lindsay MA. (Protein transduction domains: are they delivering?)

The Invention further relates to a method for treating a patient, in the patient, an effective amount of one or more of the above defined inhibitors is administered. Preferably, the patient suffers diabetes type II or diabetes type II related diseases. More preferably, the patient suffers from cachexia or HARS. For this inventive method all apply above for the use according to the invention of the inhibitor listed Embodiments.

The The invention further relates to the use of the polypeptides or nucleic acid such as defined above for the preparation of a diagnostic for diagnosis of obesity or type II diabetes.

The Detection of the protein or peptide in serum, fat, brain or muscle can be detected by Western Blot, the detection of nucleic acids PCR analysis done.

Of the Detection of the expressed protein or peptide serves as Markers for the phenotype illness. This is especially important in the diagnosis of risk groups Helpful to an increased or decreased expression of the marker exhibit. For example, this marker is in patients on Cachexia or HARS (HIV-associated-adipose redistribution syndrome ") increases expression level the marker can be used for the Determination of the therapy method and possibly for the dosage of the therapeutic agent. Based on the provision of the Protein level before, while and after the therapy is also a determination of the course of the disease possible.

The Presence of genetic variants SNPs ("small nuclear polymorphisms") on chromosomal Level can also have very far-reaching effects on predisposition of the patients have (sensitivity across from Obesity). By determining genetic variants of the proteins according to the invention or Peptides can be the course of therapies in the environment obesity diagnose and / or predict type II diabetes.

Further The present invention also relates to a diagnostic agent containing a) a polypeptide having a sequence according to SEQ ID NO: 21 or 23, b) a nucleic acid coding for a polypeptide according to a) or c) a functional variant of the molecules according to a) or b).

The The invention further relates to a method for identifying a Modulator of a polypeptide having a sequence according to SEQ ID NO: 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, or 23, or a functional one Variant of this, where you have a suitable cell with a combination from a potential modulator and a corresponding polypeptide or a functional variant of it and then the change in biological activity certainly.

This process according to the invention can be carried out, for example, as follows:
Producer cells (eg HEK 293 cells) are transfected, for example, with cDNA expression plasmids which code for the corresponding proteins according to the invention. After transfection, the cells are cultured for a period of, for example, 2 days (expression of the protein in the cell supernatant).

To at the appropriate time (for example after 3 days) after sowing the recipient cells (recipient cell line or cell lines (e.g., 3T3-L1 cells) or also to be tested Primary cells) and preferably after induction of differentiation (addition of corresponding Differentiation stimuli), the addition of corresponding modulators (Inhibitors or activators) to these cells. The modulators can selected here are from collections / libraries of chemical substances (low molecular weight substances, LMWs), peptides, proteins, antibodies, decoy receptors, Anticalins, Affilins, Antisense RNAs, siRNA, aptamers or proteases. Thereafter, for example after an incubation period known to the person skilled in the art, the carryover takes place of the cell supernatant from the producer cells to the recipient cells. The addition of the Modulators do not necessarily have to carry over the cell supernatant respectively. Alternatively, the addition of the modulator is also simultaneous or delayed. This can be particularly beneficial if the complex formation from the protein according to the invention with cellular or soluble Receptors amplified the modulation. For example, in certain cases, a recognition of a protein by an antibody only possible after complex formation of protein and associated receptor.

The recipient cells are cultured for a suitable period of time (for example 5 days). Then the effect of the modulators on the cells is determined. This can be for example an increased lipid storage (inhibitors) or a reduction in lipid storage (activators). Both can be determined by Nile Red assay.

Appropriate Process steps for preparing and cultivating the cells and the NileRed assay are described in the Examples.

Instead of the expression of the proteins according to the invention in producer cells can also recombinantly produced protein (see Example 1) to the recipient cells be added. The concentration of added recombinant Protein then corresponds to the concentration of the secreted protein in the cell supernatant the producer cell.

alternative can Non-adipocyte cells can also be used as recipient cells. For this purpose, for example, the corresponding molecular protein of the invention Receptor recombinantly expressed in these cells and used for determination the lipid storage or the insulin-dependent signal transmission ("Insulin signaling") a corresponding Reporter gene-coupled assay used. Corresponding reporter gene assays are known in the art.

The Identification of the modulator can be done using high throughput screening (HTS) method, such as in WO 01/48239 or WO 03/014346 described, performed become.

The The invention will be explained in the following by examples and figures which are not intended in any way, the subject of the invention limit.

In The examples show in particular that the inventively used Polypeptides are capable of lipid storage in murine 3T3-L1 Inhibit cells. The inhibition could also be in primary human Adipocytes of omental origin are shown. The proof took place each by NileRed assay.

Further is shown to be soluble Variants of these proteins, which are recombinantly available, or commercially available Protein fragments are also capable of lipid storage to inhibit.

Description of the figures:

1 :
Shown is the percentage inhibition of lipid storage in 3T3-L1 cell cultures relative to the negative control (empty vector). 3T3-L1 cell cultures were incubated with supernatants from 293 cells transfected with the expression plasmids coding for the respective proteins. A low signal strength means low lipid storage in the 3T3-L1 cells, a high signal strength means a strong lipid storage.

2 :
Shown is the dose response curve showing the lipid content of 3T3-L1 cells as a function of the added amount of recombinant secreted or soluble protein fragment TNFSF-12.

The Measurement results are as relative fluorescence units (RFU) per day 5 indicated after addition of the protein.

3 :
Shown is the dose response curve showing the lipid content of 3T3-L1 cells as a function of the added amount of recombinant soluble or secreted or soluble protein fragment TNFSF-14.

The Measurement results are as relative fluorescence units (RFU) per day 5 indicated after addition of the protein.

4 :
Shown is the dose response curve showing the lipid content of 3T3-L1 cells as a function of the added amount of recombinant cardiotrophin-1 protein. The measurement results are given as relative fluorescence units (RFU) on day 5 after addition of the protein.

5 :
Shown is the dose response curve showing the lipid content of 3T3-L1 cells as a function of the added amount of recombinant secreted or soluble protein fragment FGF-16. The measurement results are given as relative fluorescence units (RFU) on day 5 after addition of the protein.

HEK-293 Cells were probed with cDNA expression plasmids appropriate for the respective Encode proteins, transfected. After transfection, the HEK-293 Cells over cultivated for a period of 2 days. Thereafter, the cell supernatant of the HEK-293 cells were removed and transferred to 3T3-L1 cells (3 days after their sowing). Immediately afterwards, there was an addition of differentiation stimuli to the 3T3-L1 cells. The 3T3-L1 cells were incubated with the supernatants of the HEK-293 cells for cultured for a further 5 days and the inhibition of lipid storage by means of NileRed assay determined.

Example 1: Production an active cell culture supernatant with the help of transfection of a producer cell line (HEK 293 cells):

HEK293 cells were cultured by standard methods in DMEM medium (10% FCS) and seeded for transfection in a cell density of 2.2 × 10 ⁴ cells / well of a 96 well plate in 100 μl of medium. The transfection of the 293 cells with the cDNA expression plasmid was carried out using standard transfection methods, such as, for example, calcium phosphate transfection by Roussel et al. (Mol. Cell Biol. 4 (1984), 1999-2009). After 48 hours, 50 μl of the medium supernatant were removed and applied to the appropriately prepared preadipocyte cells (see below).

The cDNAs coding for encoding the respective proteins were either from a "full-length" enriched normalized cDNA library (Tissue sources: visceral adipose tissue, hypothalamus, skeletal muscle, Liver, pancreas), by Invitrogen, Carlsbad, USA, by a proprietary Invitrogen technology was created from a "Human Full Length Clone Collection "; OriGene; Rockville MD, USA (human cDNA clones from human cell lines and tissues) or from an MGC clone collection (IRAK collection; "Mammalian Gene Collection "; RZPD, Berlin, which includes human cDNA clones from human cell lines and tissues and in Strausberg RL, Feingold EA, Klausner RD, Collins FS. The Mammalian Gene Collection. Science, 1999, 286, 455-457 is won.

The origin of the cDNA sequences is listed below:

NFSF 12 V1 comes from a "full-length" - enriched normalized cDNA library; Invitrogen, Carlsbad, USA. The inhibition of lipid storage was corresponding Example 3 in addition with a recombinant, secreted or soluble protein from R & D Systems GmbH, Wiesbaden; CatNo .: 1090-TW; Aminoterminal 6 × HIS-tagged extracellular domain of human TWEAK (Arg 93-His 249; see Chicheportiche, et al., 1997, J. Biol. Chem. 272: 32401-32410) certainly.

TNFSF-12 V1 * comes from a "full-length" enriched normalized cDNA library; Invitrogen, Carlsbad, USA (Accession Number (Acc No.) NM_003809). The secreted protein fragment is identical to that of TNFSF12 V1. TNFSF-12 V1 * differs from TNFSF-12 V1 by an additional amino acid in the membrane protein portion. The inhibition of lipid storage was in accordance with Example 3 in addition to a recombinant, secreted or soluble protein from R & D Systems GmbH, Wiesbaden; CatNo .: 1090-TW; Aminoterminal 6X HIS-tagged extracellular domain of human TWEAK (Arg 93-His 249; Chicheportiche, et al., 1997, J. Biol. Chem. 272: 32401-32410) certainly.

TNFSF-14 was obtained from a "Human Full Length Clone Collection", ORIGENE; Rockville MD, USA (BC1286 _- C05; Acc No. NM_003807) Inhibition of lipid incorporation was supplemented with recombinant, secreted, and soluble, respectively, as described in Example 3 Protein fragment protein from R & D Systems GmbH, Wiesbaden: CatNo .: 664-LI [CD33 signal (met1-17) / 10 × His / GGGSGGGSGGGSIEGR /] - Asp-74 - Val-240.

Cardiotrophin-1 comes from a "full-length" enriched normalized cDNA library; Invitrogen, Carlsbad, USA (Acc No. NM_001330). The inhibition of Lipideinlagerung was according to Example 3 in addition with a recombinant protein from R & D Systems GmbH, Wiesbaden: CatNo .: 612-CD (protein sequence from Pennica, D. et al., 1996, Cytokines 8: 183-189) certainly.

Endothelin-2 TNFSF-14 is derived from an MGC clone collection (IRAK collection; "Mammalian Gene Collection "; RZPD, Berlin (5185394; Acc No. NM_001956). FGF-16 is derived from a" full-length "enriched normalized cDNA library; Invitrogen, Carlsbad, USA (Acc No. NM 003868). The inhibition of lipid incorporation was carried out according to Example 3 additionally with a recombinant or soluble protein from R & D Systems GmbH, Wiesbaden: CatNo .: 1212-FG / CF (As 2-207 see Miyake, A. et al., 1998, Biochem. Biophys Res. Commun. 243: 148-152).

GH-2 comes from a "Human Full Length Clone Collection "; OriGene; Rockville MD, United States. (PL1040_A04; Acc No. NM_002059).

Calsyntenin-1 comes from a "Human Full Length Clone Collection "; OriGene; Rockville MD, United States. (FB2936_B05; Acc No. NM_014944).

BM045 comes from a "Human Full Length Clone Collection "; OriGene; Rockville MD, USA; (Acc No. NM_018459).

GK001 comes from a "full-length" enriched normalized cDNA library; Invitrogen, Carlsbad, USA (Acc No. NM 020198).

PDGF-D v1 comes from a "full-length" enriched normalized cDNA library; Invitrogen, Carlsbad, USA (Acc No. NM 025208).

PDGF-D V2 comes from a "Human Full Length Clone Collection "; OriGene; Rockville MD, United States. (FB1975_A05; Acc No. NM 033135). at PDGF-D V1 and V2 are two different splice variants from PDGF-D.

Example 2: Cultivation of preadipocyte cells (3T3-L1) and preparation for induction for differentiation:

3T3-L1 Cells are mouse fibroblasts of embryonic origin which under appropriate conditions to mature a terminal differentiation Perform fat cells (adipocytes) can. Therefore, the proliferative culture of 3T3-L1 also becomes as preadipocytes designated. They grow in mono layers and have about one Doubling time of 24 h. The 3T3-L1 cells used here were derived from the ATCC (CL-173). Because the 3T3-L1 cells spontaneously at confluency differentiate and thereby change the characteristics of the culture, the cells were not cultured to confluence (max 80%; s. Passage protocol example 3).

The passenger was carried out at 2-day intervals with no medium change in between. Except for the transfer of the supernatants, there are basically no media changes. Cells were screened only at passage 5 and only until passage 18. The maintenance culture of the 3T3-L1 cells was in T-75 culture flasks. For passage every 2 days, these were inoculated with 4.5 × 10 ⁵ cells / T-75 bottle. There was no medium change until the next passage.

The seeding of the 3T3-L1 cells for the experiment was carried out in 96 well microtiter plates: The 3T3-L1 cells were seeded in a cell density of 1.5 × 10 ⁴ cells / well in 200 μl of growth medium / well. The cells were then incubated without change of medium for 3 days and then induced for differentiation as described below and contacted with the HEK-293 supernatant or recombinant soluble or secreted protein fragment.

The 3T3-L1 growth medium (GM) (composition: DMEM, 10% FCS, 1% penicillin-streptomycin, 1% Glutamine, 1% Na pyruvate) was up to 293 cells with the medium an exception identical: it has twice the amount of glutamine (2% instead of 1%, see example 3). The differentiation medium (DM) became basic as a 2-fold concentrate directly after addition of the 293-cell supernatant added. It consists in this form of a by induction factors (200 nM insulin, 1 mM IBMX and 2 μM Dexamethasone) 3T3-L1 growth medium.

Example 3: induction for the differentiation and addition of the supernatant or protein to be tested:

Three days after seeding the 3T3-L1 cells, 140 μl of medium was removed from the 3T3-L1 cells and 50 μl of a cell culture supernatant obtained from HEK293 cells was added. Alternatively wur de 50 ul of growth medium were added, were dissolved in the appropriate amounts of recombinant protein (0-3 ug / ml protein). In addition, 100 μl of 2X differentiation medium was added to the 3T3-L1 cells followed by incubation of the 3T3-L1 cells in the incubator for 5 days. After this time, a determination of the intracellularly incorporated lipid was made.

Example 4: Measurement of the intracellularly embedded lipid:

to quantitative determination of the incorporation of cellular lipid NileRed reagent (Molecular Probes, Leiden, Netherlands; CAS Number 7385-67-3) (Nile Red staining solution: 4 μg / ml Nile Red in PBS / 40% DMSO). The fluorescence was at an excitation wavelength of 485 nm and an emission wavelength of 590 nm. The Required amount of Nile Red was added to the calculated amount of DMSO and mixed. Subsequently the calculated amount of PBS is added and the solution mixed.

Example 5: Implementation of the NileRed assay:

On day 5 after induction for differentiation, 140 μl of medium was removed from the 3T3-L1 cell cultures and 200 μl of PBS added. Thereafter, 150 .mu.l of liquid was removed and added to each 50 .mu.l NileRed staining solution. The incubation of the plates was carried out for ₄ h at 37 ° C in the CO ₂ incubator. The reading was carried out in a fluorescence reader at an excitation of 485 nm and an emission of 590 nm (200 msec reading time). The measurement results are known as Relative Fluorescence Units (RFU) in 1 (Cell supernatants from transfected HEK-293 cells or in 2 - 5 (recombinant protein).

Example 6: Reduction of weight or weight gain in the mouse model

In mice of the strain C57 / B16 / J is determined by the maximum tolerated Dose (MTD) a) Reducing weight gain in normal weight mice (without previous high calorie diet) and b) the net reduction of the obese Tissue in obese mice (under high calorie diet) after addition of the invention used Protein determined by weighing or determination of the BMI ("body mass index").

The Determination of the maximum tolerated dose is carried out with 3 mice (strain C57B16 / J wild type) by administration of 0.002; 0.02; 0.2; 2.0 and 10 mg Protein / kg as a single intravenous dose. A subsequent fine adjustment The dose is administered with 3 mice each (Strain C57B16 / J wild type) by a single dose of the protein. In one Dosage tolerance test is performed by repeatedly administering a selected dose over several Applications based on blood parameters and serum stability Application determined necessary dose.

in the Framework of the continuing Experiments will reduce the maximum dose found above to a minimum Reduces the dose that still has a therapeutic effect. In the subsequent Test to determine the change in weight is the thus determined Dose administered intraperitoneally.

to Determining the reduction of weight gain in normal weight mice become normal-weight mice under normal diet (20 kcal% fat diet) each without (untreated) and with the addition of the protein according to the invention (treated) held for 10-14 weeks. During and after the end of the test series The weight gain is determined by weighing and determining the BMI.

at treated mice, the under normal diet are kept in contrast to untreated mice one To observe weight reduction. In treated mice, the under high calorie diet is kept compared to untreated mice to observe less weight gain or a weight reduction.

to Identify the reduction in weight gain in obese mice become obese mice (previously under high calorie diet (45 kcal% fat diet) under normal diet (20 kcal% fat diet) each without (untreated) and with addition of the protein according to the invention (treated) held for 10-14 weeks. During and after the end of the test series The weight gain is determined by weighing and determining the BMI.

In treated mice kept under normal diet, a greater weight reduction can be shown than in untreated mice. In untreated mice under high calorie diet, a further increase in weight is observed. In treated mice, however, is a weight loss me or a reduced increase in weight determine.

Preferred Embodiments: Protein Fragments: TNFSF-12 v1 and v2 (AS 93-249 and 250, respectively)

TNFSF-14 (AS 74-240)

FGF-16 (AS 2-207)

Endothelin-2 (AS 49-69)

It follows a sequence listing according to WIPO St. 25. This can from the official publication platform downloaded from the DPMA.

Claims (22)

use of  a) a polypeptide having a Sequence according to SEQ ID No: 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, or 23,  b) one nucleic acid coding for a polypeptide according to a), or  c) a functional variant of the molecules according to a) or b)  for the production a pharmaceutical composition for the treatment of obesity. Use according to claim 1, characterized that obesity is an uncomplicated obesity. Use according to one of claims 1 or 2, characterized that the pharmaceutical composition makes the formation terminally differentiated Adipocytes inhibits. Use according to claim 3, characterized that the formation of terminally differentiated adipocytes is inhibited through promotion the proliferation of preadipocytes, by inhibition of differentiation to terminally differentiated adipocytes, in particular by inhibition of the transcriptional signaling pathway of differentiation or by promotion of cell death, preferably apoptosis in preadipocytes. Use according to one of claims 1 or 2, characterized that the pharmaceutical composition is the function of the terminal differentiated adipocytes inhibits. Use according to claim 5, wherein the inhibition is by inhibiting lipogenesis or promoting lipolysis becomes. Containing a pharmaceutical composition  a) a polypeptide having a sequence according to SEQ ID No: 21 or 23,  b) a nucleic acid coding for a polypeptide according to a), or  c) a functional variant of the molecules according to a) or b). Use of an inhibitor of a polypeptide with a sequence according to SEQ ID No: 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, or 23 or a functional one Variant thereof for the preparation of a pharmaceutical composition for the treatment of diabetes type II, diabetes type II related diseases, Cachexie or HARS. Use according to claim 8, wherein the inhibitor is selected from the group consisting of antibodies, decoy receptors, peptides, Proteins, Anticalins, Affilins, Aptamers, Low Molecular Substances (LMWs), antisense RNA, siRNA and proteases. Use according to one of claims 8 or 9, characterized that the pharmaceutical composition makes the formation terminally differentiated Promotes adipocytes. Use according to claim 10, characterized that the formation of terminally differentiated adipocytes is promoted by inhibiting the proliferation of preadipocytes, by promoting differentiation to terminally differentiated adipocytes, in particular through promotion the transcriptional signaling pathway of differentiation or by inhibition of cell death, preferably apoptosis in preadipocytes. Use according to one of claims 8 or 9, characterized that the pharmaceutical composition is the function of the terminal differentiated adipocytes promotes. Use according to claim 12, wherein the promotion through a promotion lipogenesis or by inhibiting lipolysis. Use according to one of claims 8 to 13, characterized that inhibition of insulin-dependent signal transmission in non-adipocyte cell types will be annulled. A pharmaceutical composition containing a Inhibitor of a polypeptide having a sequence according to SEQ ID No: 21 or 23 or a functional variant of it. Use or pharmaceutical composition according to one of the claims 1 to 15, characterized in that the pharmaceutical composition is present in a form which the contact of the active ingredient with a Component of the cell surface allows. Use or pharmaceutical composition according to one of the claims 1 to 15, characterized in that the pharmaceutical composition is present in a form which facilitates transport of the active ingredient into the Cell interior allows. Method for identifying a modulator of a Polypeptide having a sequence according to SEQ ID No: 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, or 23 or a functional variant of which you have a suitable cell with a combination of a potential modulator and a polypeptide having a sequence according to SEQ ID No: 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, or 23 or a functional one Variant of incubated and then the change in biological activity certainly. Method according to claim 18, characterized that change the biological activity of adipocytes, preferably 3T3-L1 cells. Method according to one of claims 18 or 19, characterized that change the biological activity in a reduction or increase in fat storage in cells, preferably adipocytes. use of  a) a polypeptide having a Sequence according to SEQ ID No: 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, or 23,  b) one nucleic acid coding for a polypeptide according to a), or  c) a functional variant of the molecules according to a) or b)  for the production a diagnostic for the diagnosis of obesity. Containing diagnostic agent  a) a polypeptide with a sequence according to SEQ ID No: 21 or 23,  b) a nucleic acid encoding a polypeptide according to a), or  c) a functional variant of the molecules according to a) or b).