CN1697661A - Methods of treating diabetes using PDE 11A inhibitors - Google Patents

Abstract

Methods of the invention relate to treatment of diabetes, particularly type 2 diabetes, and related disorders by administration of a PDE11A inhibitor. Such PDE11A inhibitors may be administered in conjunction with alpha-glucosidase inhibitors, insulin sensitizers, insulin secretagogues, hepatic glucose output lowering compounds, beta3 agonist or insulin. Such PDE11A inhibitors may also be administered in conjunction with body weight reducing agents. Further methods of the invention relate to stimulating insulin release from pancreatic cells, particularly in response to an elevation in blood glucose concentration, by administration of a PDE11A inhibitor.

Description

Methods of treating diabetes with PDE11A inhibitors

technical field

The present invention relates to methods of treating diabetes and related diseases by administering compounds that inhibit PDE11A.

Background technique

Diabetes mellitus is characterized by impaired glucose metabolism, particularly manifested in diabetic patients by elevated blood glucose levels. Diabetes is mainly divided into two types according to the underlying etiology: type 1 and type 2. Type 1 diabetes, or insulin-dependent diabetes mellitus (IDDM), occurs due to a lack of insulin-producing beta cells in the patient's pancreas. Type 2 diabetes, or non-insulin-dependent diabetes mellitus (NIDDM), occurs as a result of impaired beta-cell function in the patient as well as altered insulin function.

The current treatment for patients with type 1 diabetes is insulin injections, while most patients with type 2 diabetes are treated with agents that stimulate β-cell function or agents that enhance the patient's tissue sensitivity to insulin. Drugs currently used to treat type 2 diabetes include alpha-glucosidase inhibitors, insulin sensitizers, insulin secretagogues, metformin, and insulin.

Over time, more than a third of people with type 2 diabetes lose their response to oral medications. Insulin therapy is initiated when blood sugar is not adequately controlled by diet, exercise, and oral therapy. Drawbacks of insulin therapy are the need for drug injections, possible hypoglycemia, and weight gain.

Another strategy for the treatment of diabetes is based on the signaling mechanism of cyclic adenosine monophosphate (cAMP) and its effect on insulin secretion. Glucose metabolism promotes ATP-dependent K ⁺ channel closure, which leads to cellular depolarization and subsequent opening of Ca ⁺⁺ channels. This in turn leads to exocytosis of insulin granules. cAMP is the master regulator of glucose-stimulated insulin secretion. However, the effect of cAMP is glucose-dependent, that is, at low glucose concentrations, cAMP has little if any effect on insulin secretion (Weinhaus, A., et al., Diabetes 47:1426-1435 (1998) ). The effect of cAMP on insulin secretion is thought to be regulated through the protein kinase A pathway.

Endogenous secretagogues use the cAMP system to control insulin secretion in a glucose-dependent manner (Komatsu, M. et al., Diabetes 46:1928-1938 (1997)). Examples of such endogenous secretagogues include pituitary adenylate cyclase activating peptide (PACAP), vasoactive intestinal polypeptide (VIP), and glucagon-like peptide-1 (GLP-1).

PACAP is a potent enhancer of glucose-dependent insulin secretion from pancreatic β-cells. Three distinct PACAP receptor types (R1, R2 and R3) have been described (Harmar, A. et al., Pharmacol. Reviews 50:265-270 (1998)). The insulinotropic activity of PACAP is regulated by the GTP-binding protein Gs. The accumulation of intracellular cAMP can activate non-selective cation channels in β-cells, increase [Ca ⁺⁺ ], and promote exocytosis of insulin-containing secretory granules.

Vasoactive intestinal peptide (VIP) is a 28 amino acid peptide that was first isolated from the upper small intestine of pigs (Said and Mutt, Science 169:1217-1218, 1970; US Patent 3,879,371). The biological effects of VIP are regulated by the activation of membrane-bound receptor proteins coupled to the intracellular cAMP signaling system.

Postprandial GLP-1 is released from intestinal L-cells and acts as an endocrine hormone (ie, it promotes glucose-induced insulin release from pancreatic β-cells). GLP-1 is a 37-amino acid peptide that is differentially expressed by the glucagon gene according to tissue type. Clinical data supporting the beneficial effect of increasing cAMP levels in β-cells was collected with GLP-1. Infusion of GLP-1 in patients with poorly controlled type 2 diabetes normalized fasting blood glucose levels (Gutniak, M., et al., New Eng. J. Med. 326:1316-1322, (1992)), and The longer the infusion time, the more the function of β-cells in normal patients can be improved (Rachman, J., et al., Diabetes 45:1524-1530, (1996)). A recent report indicated that GLP-1 enhanced the ability of β-cells to respond to glucose in patients with impaired glucose tolerance (Byrne M., et al., Diabetes 47:1259-1265 (1998)).

Treatment of type 2 diabetes with these endogenous secretagogues also has several drawbacks. For example, the peptidic nature of these compounds necessitates injectable administration. Furthermore, the effect of endogenous secretagogues is short-lived because of the short shelf-life of the peptides.

Due to problems with current treatments, new treatments for type 2 diabetes are also needed. In particular, new therapeutic approaches that preserve normal (glucose-dependent) insulin secretion are needed. Such new drugs should have the following characteristics: 1) dependence on glucose when promoting insulin secretion, that is, a compound that can promote insulin secretion only when the blood sugar concentration increases, so that the possibility of hypoglycemia is very small; 2 ) low first and second failure rates; and 3) preservation of islet cell function. The present invention meets the above needs by inhibiting phosphodiesterase 11A (PDE11A) and thereby controlling the cAMP signaling system.

Phosphodiesterases (PDEs) are a collection of cAMP and/or cGMP-hydrolases that cleave 3',5'-cyclic nucleotide monophosphates into 5'-nucleotide monophosphates. PDEs are known to be involved in the regulation of the cAMP system. Specifically, PDE11A is a phosphodiesterase that hydrolyzes cAMP and cGMP, and its K _m value is about 1-5 μ M (Fawcett et al., Proc Natl Acad Sci USA , 2000 Mar 28; 97 (7): 3702-7 (2000); Hetman et al., Proc Natl Acad Sci USA , 2000 Nov 7;97(23):12891-5(2000); Yuasa et al., Eur J Biochem , 2001 Aug;268(16):4440-8(2001)). At least four splice variants of PDE11A have been described that share the C-terminal catalytic domain and differ in the size of the N-terminal portion of the molecule. Yuasa et al., Eur J Biochem. , (2001), 268(16), 4440-4448; Yuasa et al., J. Bio. Chem. (2000), 275(40), 31469-31479.

Brief description of the invention

The present invention relates to a method of treating diabetes, especially type 2 diabetes, in a mammal by administering an effective amount of a PDE11A inhibitor. Other methods of the invention involve the treatment of other diseases associated with diabetes, such as syndrome X, impaired glucose tolerance, and impaired fasting glucose, by administering PDE11A inhibitors. The present invention also relates to methods of stimulating insulin release from pancreatic cells in mammals by administering an effective amount of a PDE11A inhibitor. The method of insulin release is responsive to an increase in blood glucose concentration in a mammal. In the method of the present invention, PDE11A inhibitors can also be used together with other diabetes treatment drugs, such as α-glucosidase inhibitors (such as acarbose), insulin sensitizers (such as thiazolidinediones), reducing Glycogen producing compounds (eg metformin), insulin secretagogues (eg sulfonylureas), beta3-agonists and insulin. Furthermore, PDE11A inhibitors may also be used in combination with one or more weight reducing agents, such as xenical, sibutramine, β3-agonists or CB-1 antagonists. Finally, in another embodiment, the method of the invention provides for the use of a PDE11A inhibitor in combination with an HMGCoA reductase inhibitor, nicotine acid, bile acid sequestrants, fibricacid derivatives or antihypertensive drugs.

In other methods of the invention, PDE11A inhibitors may be used to treat dementia or urogenital disorders. Genitourinary disorders include, but are not limited to, incontinence, stress incontinence, benign prostatic hyperplasia, erectile dysfunction, female sexual dysfunction, and enlarged prostate. In other methods of the invention, PDE11A inhibitors are useful in the treatment of cardiovascular diseases such as hypertension, ischemic heart disease, myocardial infarction, stable and unstable angina, peripheral occlusive disease, and ischemic shock.

The present invention thus provides methods of treating diabetes by inhibiting PDE11A by administering a PDE11A inhibitor. These and other aspects of the invention will become more apparent from the following drawings, description and claims.

Description of drawings

Figures 1A-1C show expression of PDE11A in islet cells.

Figure 1A shows PCR products generated using the F2/R1 primer composition using islet cDNA as a template.

Figure 1B shows PCR products generated with the For3/R2 primer composition using islet cDNA as a template.

Figure 1C shows PCR products generated using the F4/Rev2 primer composition using islet cDNA as a template.

In the figure, arrows indicate PCR products with the expected size. The electrophoresis lane marks are as follows: 1Kb = 1Kb DNA standard marker, islet = islet cDNA template, -DNA = negative DNA control, PDE11A = positive control using a plasmid containing PDE11A as a template, Neg = using a plasmid containing an irrelevant gene Negative control as template.

Detailed description of the invention

The present invention provides methods of treating diabetes and its related diseases, especially type 2 diabetes and/or stimulating insulin release from pancreatic cells by administering PDE11A inhibitors. This approach treats any disease in which blood glucose levels rise on an empty stomach or after a meal by administering a PDE11A inhibitor. PDE11A has been identified in islet cells of pancreatic islets. PDE11A hydrolyzes cAMP to AMP, thereby reducing the intracellular cAMP concentration. By inhibiting the activity of PDE11A, intracellular cAMP levels are increased, thereby increasing the release of insulin-containing secretory granules and thus insulin secretion. Compounds that inhibit PDE11A activity in the islet cell assay also stimulate insulin secretion, as described in the text. Also as described herein, inhibitors of PDE11A are useful in the treatment of dementia, cardiovascular disease, or genitourinary tract disease.

treatment method

The methods of the invention are useful in the treatment of diseases such as diabetes, including type 1 diabetes and type 2 diabetes. These approaches can also delay the onset of diabetes and its complications. Other diseases and conditions treatable or preventable by the methods of the present invention include: Maturity-Onset Diabetes of the Young (MODY) (Herman et al., Diabetes 43:40 (1994)), adult occult Autoimmune diabetes (LatentAutoimmune Diabetes Adult, LADA) (Zimmet et al., Diabetes Med. 11:299 (1994)), impaired glucose tolerance (IGT) (Expert Committee on Diabetes Classification, Diabetes Care 22 (Supp.1) S5 (1999 )), Impaired Fasting Glucose (IFG) (Charles et al., Diabetes 40:796 (1991)), Gestational Diabetes Mellitus (Metzger, Diabetes 40:197 (1991)), and Metabolic Syndrome X.

The methods of the present invention can also be used to treat secondary causes of diabetes (Expert Committee on Classification of Diabetes, Diabetes Care 22 (Supp. 1) S5 (1999)). Such secondary causes include glucocorticoid excess, growth hormone excess, paraganglioma, and drug-induced diabetes. Drugs that may induce diabetes include, but are not limited to, pyriminil, nicotine acid, glucocorticoids, phenytoin, thyroxine, beta-adrenaline agents, alpha-interferon, and drugs for the treatment of HIV infection.

The release of cAMP-mediated insulin also depends on the presence of a stimulating glucose concentration. One method of the invention involves stimulating insulin release from islet cells by administering a PDE11A inhibitor. Glucose-dependent stimulation of insulin secretion with non-peptide compounds therefore lowers blood glucose concentrations without causing hypoglycemia.

The methods of the present invention may be used alone or in combination with other therapies and/or other compounds known to those skilled in the art for the treatment of diabetes and related diseases. Alternatively, in combination therapy, a PDE11A inhibitor may be used partially or completely.

PDE11A inhibitors may also be used in combination with other drugs known to treat diabetes, including PPAR agonists, sulfonylureas, nonsulfonylurea secretagogues, alpha-glucosidase inhibitors, insulin sensitizers , insulin secretagogues, compounds that reduce glycogen production, and insulin. These drugs may be administered before, simultaneously with, or after the administration of the PDE11A inhibitor. Insulin includes long- and short-acting forms as well as preparations of insulin. PPAR agonists may include agonists of any PPAR subtype or combinations thereof. For example, a PPAR agonist may include an agonist of PPAR-alpha, PPAR-gamma, PPAR-delta, or a combination of any two or three PPAR subunits. PPAR agonists include, for example, rosiglitazone and pioglitazone. Sulfonylureas include, for example, glibenclamide, glimepiride, chlorpropamide, and glipizide. Alpha-glucosidase inhibitors that can also be used when PDE11A inhibitors are used to treat diabetes include acarbose, miglitol, and voglibose. Insulin sensitizers that can be used concurrently with PDE11A inhibitors to treat diabetes include thiazolidinedione and non-thiazolidinedione. Glycogen production-lowering compounds that may also be used when PDE11A inhibitors are used to treat diabetes include metformin, such as metformin and metformin XR. Insulin secretagogues that can be used concurrently with PDE11A inhibitors for diabetes include sulfonylureas and non-sulfonylureas: GLP-1, GIP, PAC/VPAC receptor agonists, secretin, napaglidium Nai, meglitinib, repaglinide, glibenclamide, glimepiride, chlorpropamide, glipizide. GLP-1 includes GLP-1 derivatives with a longer half-life than GLP-1 itself, such as fatty acid derivatives and exendins of GLP-1. In one embodiment of the invention, a PDE11A inhibitor is used in combination with an insulin secretagogue to increase the sensitivity of pancreatic beta cells to the insulin secretagogue.

PDE11A inhibitors may be used in combination with weight loss drugs. Weight loss drugs include beta-3 agonists, CB-1 antagonists, appetite suppressants such as sibutramine (Sibutramine), and lipase inhibitors such as orlistat (Xenical).

PDE11A inhibitors can also be used in combination with drugs commonly used to treat lipid metabolism disorders in diabetics. Such drugs include, but are not limited to, HMG-CoA reductase inhibitors, nicotine acid, bile acid sequestrants, and fibric acid derivatives. The methods of the invention may also be used in combination with antihypertensive drugs, such as beta-blockers and ACE inhibitors.

This combination therapy can optionally be combined with two or more drugs (for example, a PDE11A inhibitor combined with an insulin sensitizer and a weight loss drug). Such combination therapy can be administered in the form of pharmaceutical compositions, as described above.

Other methods of the invention relate to the treatment of dementia with inhibitors of PDE11A. Shimamoto et al., Mechanisms of Aging Development (1976), 5(4):241-250; Nicholson et al., Trends Pharmacological Sciences (1991), 12(1):19-27.

Another method of the invention involves the treatment of urogenital disorders with PDE11A inhibitors. The urogenital disorders include, but are not limited to, incontinence, stress urinary incontinence, benign prostatic hyperplasia, erectile dysfunction, female sexual dysfunction (including female sexual arousal disorder), and prostatic hypertrophy. Ballard et al., J Urology 159(6):2164-2171 (1998); EP1211313A2 (on the role of PDE11A on spermatogenesis).

Another method of the invention involves the use of PDE11A inhibitors in the treatment of cardiovascular diseases such as hypertension, ischemic heart disease, myocardial infarction, stable and unstable angina, peripheral occlusive disease, and ischemic shock. The PDE11 family includes enzymes responsible for cAMP and cGMP degradation in various tissues (Fawcett et al., PNAS (2000) 97, 3702-3707). Furthermore, the expression of PDE11 can be detected in the heart (Yuasa et al., Eur. J. Biochem. (2001) 268, 4440-4448). Cyclic GMP (cGMP) and cyclic AMP (cAMP) are important second messengers involved in the regulation of vascular smooth muscle tone. Activation of soluble, membrane-bound guanylate cyclase increases intracellular cGMP levels and induces vasodilation. Activation of various G protein-coupled receptors (GPCRs) expressed in vascular smooth muscle cells (eg adrenomedullin and CGRP receptors) leads to activation of adenylyl cyclase, production of intracellular cAMP and vasodilation. 3'5'-cyclic nucleotide phosphodiesterases (PDEs) catalyze the hydrolysis of 3'5'-cyclic nucleotides to their respective nucleotide 5'-monophosphates. For all of the above reasons, it is likely that PDE11A plays a role in the cardiovascular system.

pharmaceutical composition

The PDE11A inhibitors used in the methods of the invention may be administered as the compound itself. Alternatively, the PDE11A inhibitor can be administered in the form of a pharmaceutical composition together with a pharmaceutically acceptable carrier. A pharmaceutically acceptable carrier must be compatible with the other ingredients of the composition and must not be intolerably deleterious to the user. The carrier can be a solid or liquid, or a mixture of both, and is preferably formulated with the compound as a unit dosage composition, such as a tablet, which may contain from about 0.05% to about 95% by weight of the active compound, based on the total weight of the dosage form. Other pharmacologically active substances may also be present, including other compounds useful in the treatment of diabetes.

The PDE11A inhibitor used in the method of the present invention can be administered by any appropriate route, preferably in the form of a pharmaceutical composition suitable for this route of administration, and has a therapeutically effective dose required for treatment. PDE11A inhibitors can be administered, for example, orally, sublingually, nasally, pulmonary, mucosally, parenterally, intravenously, intraperitoneally, subcutaneously, intramuscularly administered or topically. Unit dosage formulations, particularly orally administrable unit dosage formulations such as tablets or capsules, generally contain, for example, from about 0.001 mg to about 500 mg, preferably from about 0.005 mg to about 100 mg, more preferably from about 0.01 mg to about 50 mg, of the active ingredient. For pharmaceutically acceptable salts, the weight of the above active ingredient refers to the weight of the pharmaceutically active ion obtained from the salt.

For oral administration, the pharmaceutical compositions can be in the form of, for example, tablets, capsules, suspensions, emulsions, pastes, solutions, syrups or other liquids. Pharmaceutical compositions are preferably presented in dosage unit form containing specific quantities of the active ingredient. If administered orally, the compound may be formulated with, for example, lactose, sucrose, starch powder, cellulose alkanoates, cellulose alkyl esters, talc, stearic acid, magnesium stearate, magnesium oxide, sodium salts of phosphoric and sulfuric acids and Calcium salt, gelatin, gum arabic, sodium alginate, polyvinylpyrrolidone, and/or polyvinyl alcohol are mixed, and then compressed into tablets or filled into capsules for convenient administration.

Oral administration of PDE11A inhibitors may include dosage forms known in the art that release the drug immediately, or delayed or sustained, to the gastrointestinal tract through a variety of mechanisms. Dosage forms for immediate release include, but are not limited to, oral solutions, oral suspensions, fast-dissolving tablets or capsules, sublingual tablets, disintegrating tablets, and the like. Delayed or sustained release dosage forms include, but are not limited to, pH-sensitive release of the active ingredient from dosage forms based on changes in the pH of the small intestine, slow erosion of tablets or capsules, retention in the stomach based on physical properties of the formulation, effects of the dosage form on the gut. Bioadhesion of mucosal layers, or enzymatic release of active drug from dosage forms. The desired effect is to prolong the duration of action by controlling the dosage form so that the active drug molecule is delivered to the site of action. Thus, enteric and enteric controlled release formulations are useful in the methods of the invention. Suitable enteric coatings include cellulose acetate phthalate, polyvinyl acetate phthalate, hydroxypropylmethylcellulose phthalate, and methacrylic and methyl methacrylate anions polymer.

Pharmaceutical compositions suitable for oral administration may be presented as discrete units, such as capsules, cachets, lozenges or tablets, each containing a predetermined amount of at least one compound of the invention; in powder or granule form; aqueous or without Aqueous solutions or suspensions; oil-in-water or water-in-oil emulsions. As indicated herein, such compositions may be prepared by any suitable method in pharmacy, including bringing into association the inhibitor and the carrier (which may contain one or more excipients). In general, the compositions are prepared by uniformly and intimately bringing into association the inhibitor with liquid or finely divided solid carriers, or both, and then, if necessary, shaping the product. For example, a tablet may be obtained by compressing or molding the inhibitor powder or granules, optionally with one or more accessory ingredients. Compressed tablets may be obtained by compressing, in a suitable machine, the compound in a free-flowing state, such as a powder or granules, optionally with a binder, lubricant, inert diluent and/or surface active/dispersing agent. derived from a mixture. Molded tablets may be obtained, for example, by molding the powdered compound in a suitable machine.

Oral liquid dosage forms may include pharmaceutically acceptable emulsions, solutions, suspensions, syrups and elixirs containing inert diluents commonly used in the art, such as water. Such compositions may also contain adjuvants such as wetting agents, emulsifying and suspending agents, and sweetening, flavoring and perfuming agents.

Pharmaceutical compositions suitable for buccal (sublingual) administration include lozenges containing the PDE11A inhibitor in a flavored base, usually sucrose, acacia or tragacanth, and in an inert base such as Pastilles containing inhibitors in gelatin and glycerin or sucrose and acacia.

Preparations for parenteral administration may be in the form of, for example, aqueous or non-aqueous isotonic sterile injection solutions or suspensions. These solutions and suspensions can be prepared from sterile powders or granules with one or more of the carriers or diluents previously mentioned for use in the preparation of oral formulations. PDE11A inhibitors can be dissolved in water, polyethylene glycol, propylene glycol, ethanol, corn oil, cottonseed oil, peanut oil, sesame oil, benzyl alcohol, sodium chloride, and/or various buffers. Other adjuvants and modes of administration are well known in the art of pharmacy.

Pharmaceutically acceptable carriers include all of the foregoing and their analogs. Pharmaceutical compositions containing a PDE11A inhibitor for use in the methods of the invention can be prepared by any technique known in pharmacy, such as mixing the components. The above considerations with regard to effective dosage forms and methods of administration are well known in the art and are described in standard texts.

The dosage of the PDE11A inhibitor used in the method of the present invention is generally about 0.001 mg to about 10000 mg/day, preferably about 0.005 mg to about 1000 mg/day. Based on mg/kg daily dosage, or single or multiple administration, the dosage range is generally about 0.001/75mg/kg-about 10000/75mg/kg, preferably about 0.005/75mg/kg-about 1000/75mg /kg.

The total daily dose of each inhibitor may be administered to the patient in one dose or in subdoses. Generally, sub-doses may be administered 2-6 times a day, preferably 2-4 times a day, even more preferably 2-3 times a day. The agent may be in an immediate release or sustained release form sufficiently potent to effectively manage the diabetic condition.

Dosage regimens for preventing, treating, alleviating or alleviating diabetic conditions or diseases, or otherwise preventing or treating diabetes with the combinations and compositions of the present invention are selected based on a variety of factors. These factors include, but are not limited to, the patient's type of diabetes, age, weight, sex, diet and medical condition, severity of disease, route of administration, pharmacological factors such as the activity, potency, pharmacokinetics and The toxicological profile, whether a drug delivery system was used, and whether the inhibitor was administered with other active ingredients. Thus, the actual dosage regimen employed may vary widely and may therefore deviate from the preferred dosage regimens set forth above.

Pharmaceutically acceptable salts of compounds useful as PDE11A inhibitors in the methods of the invention include those commonly used to form alkali metal salts or to form addition salts with free acids or free bases. The nature of the salt itself is not critical as long as it is a pharmaceutically acceptable salt. Suitable pharmaceutically acceptable acid addition salts may be prepared from inorganic or organic acids. Examples of inorganic acids are hydrochloric acid, hydrobromic acid, hydroiodic acid, nitric acid, carbonic acid, sulfuric acid and phosphoric acid. Suitable organic acids may be selected from fatty acids, cyclic fatty acids, aromatic acids, heterocyclic acids, carboxylic acids and organic sulfonic acids. Examples of organic and organic sulfonic acids include, but are not limited to, formic acid, acetic acid, propionic acid, butyric acid, glycolic acid, gluconic acid, lactic acid, malic acid, tartaric acid, citric acid, ascorbic acid, glucuronic acid, maleic acid, Malic acid, pyruvic acid, aspartic acid, glutamic acid, benzoic acid, anthranilic acid, mesylic acid, salicylic acid, 4-hydroxybenzoic acid, phenylacetic acid, mandelic acid, embemic acid ( embonic, pamoic), methanesulfonic acid, ethanesulfonic acid, benzenesulfonic acid, pantothenic acid, 2-hydroxyethanesulfonic acid, p-toluenesulfonic acid, p-aminobenzenesulfonic acid, cyclamate, stearic acid, algenic acid, N-hydroxybutyric acid, salicylic acid, galactaric acid and galacturonic acid and combinations thereof.

The PDE11A inhibitors used in the present invention may also be administered in the form of pharmaceutically acceptable salts, protected acids, conjugate acids, tautomers, prodrugs or stereoisomers of compounds found to inhibit PDE11A activity . Tautomers include, for example, hydroxy tautomers. Protected acids include, but are not limited to, protected acids such as esters, hydroxylamine derivatives, amides and sulfonamides. The formation of prodrugs is well known in the art and is aimed at enhancing the properties of the parent compound; such properties include solubility, absorption, biostability and release time (see "Pharmaceutical Dosage Form and Drug Delivery Systems" (Sixth Edition) , edited by Ansel et al., Williams & Wilkins, pp. 27-29 (1995), incorporated herein by reference). Commonly used prodrugs designed to take advantage of major drug biotransformation reactions are also considered within the scope of the present invention. The main drug biotransformation reactions include N-dealkylation, O-dealkylation, aliphatic hydroxylation, aromatic hydroxylation, N-oxidation, S-oxidation, deamination, hydrolysis, glucuronidation, Sulfation and Acetylation (See Goodman and Gilman, The Pharmacological Basis of Therapeutics (Ninth Edition), eds. Molinoff et al., McGraw-Hill Publishing, pp. 11-13 (1996), incorporated herein by reference).

In addition to being used to treat human diseases, PDE11A inhibitors can also be used in the veterinary treatment of companion animals (eg, horses, dogs, cats, etc.), exotic animals, and farm animals. Although the present invention has been described in terms of human biology, those of ordinary skill in the art will appreciate that the present invention is also applicable to other mammals.

Expression pattern map

The expression of PDE11A in pancreatic islet cells was verified by PCR. PCR was performed with DNA templates from an insular cell cDNA library with the following primer combinations recognizing different regions of the PDE11A gene:

F2(5'-CATACCATGCAACATGTTCA-3')+R1(5'-CAGTTTCACGTTGACCTTCA-3'), which would give the expected product with 928 base pairs.

For3 (5'-AAAAGCGGCCGCCCACCATGAGCCCAAAGTGCAGT GCTGA-3'; note that this primer includes additional sequences for cloning purposes)

+R2 (5'-CTGACAAGTTCAAAGAATTCA-3'), which would give the expected product of 1060 bp.

F4 (5'-CGCTGTACTTTGAGAGGAGA-3')+Rev2 (5'-AAAAAAGCTTGTTTAGTTCCCTGTCTTCCTT-3'; note that this primer includes additional sequences for cloning purposes) which would yield the expected product of 474 base pairs .

A 50 μl PCR reaction was assembled as follows:

5 μl 10x amplification buffer (supplied with polymerase)

5 μl 10x PCR enhancer (supplied with polymerase)

1 μl 50mM _MgSO4

8μl each of 1.25mM dATP, dCTP, dGTP, dTTP

1 μl 100 μM forward primer

1 μl 100 μM reverse primer

1 μl island cell cDNA

27 μl H ₂ O

1 μl PLATINUM Pfx DNA Polymerase (Life Technologies)

Reactions were cycled in a Perkin Elmer GeneAmp PCR System 9600 Thermal Cycler using the following parameters:

95°C for 2 minutes/35x(95°C for 30 seconds/55°C for 30 seconds/72°C for 2 minutes)/72°C for 10 minutes

1 μl of Taq DNA polymerase (Perkin Elmer) was added to the reaction and reacted at 72°C for another 10 minutes, then the reaction was cooled to 4°C.

The reaction was loaded onto a 1% agar TBE gel and electrophoresed at 150V for 20 minutes. PCR products were visualized by UV irradiation after ethidium bromide staining.

The PCR product of the F2/R1 reaction was selected for further characterization by DNA sequencing because this product corresponds to the part of the PDE11A gene encoding the catalytic domain. Briefly, PCR fragments were excised from the gel and purified using the Qiagen Gel extraction kit following the protocol provided with the kit. The purified PCR product was inserted into the pcDNA3.1/V5/His-TOPO matrix, and TOP10 active bacterial cells were subsequently transformed using the Eukaryotic TOPO TA cloning kit (Invitrogen) as described by the manufacturer. Pick five colonies into 2ml LB broth and culture with 100μg/ml carbenicillin at 37°C overnight. Miniprep DNA was prepared from overnight cultures, and the presence of the insert in the PCR product was verified by restriction enzyme analysis. The insertion was identified as positive for PDE11A by DNA sequencing.

Figures 1A-1C show PED11A PCR products generated using the primer compositions described above. As shown in the figure, PDE11A was found in islet cells. The expression of PDE11A in islet cells suggests that PDE11A may play a role in regulating insulin secretion/blood glucose concentration.

PDE11A inhibition assay

Compounds were added to human recombinant enzyme assay buffer and spotted on a 96-well whitewall/clear bottom isoplate (Wallac). Reactions were initiated by adding 3H-cAMP (Amersham) or 3H-cGMP (Amersham). After 45 minutes at room temperature, the reaction was quenched by adding SPA yttrium silicate beads (Amersham). After 30 minutes, the plate was read in SPA mode in Microbeta (Wallac) for 30 seconds. Data are expressed as percentage of control. Use PDE2, PDE3A, PDE4B and PDE11A, 3H-cAMP as substrates. With PDE5, 3H-cGMP as the substrate.

Compounds were identified as inhibiting the activity of PDE11A with _IC50 values of 1 μM or less. Compounds include the following:

These same compounds are used in methods for assaying inhibition of PDE2, PDE3A, PDE4B and PDE5. _In these methods, compounds were found to have _IC50 values 10-fold higher than those in the PDE11A inhibition method. These compounds are therefore selective for PDE11A.

These compounds can be administered in the methods of the invention. In addition, their stereoisomers, pharmaceutically acceptable salts, tautomers, protected acids and conjugate acids and/or prodrugs may be administered in the methods of the present invention.

Island Cell Assay

Isolation of pancreatic islet cells: Lean rats (Sprague-Dawley, male, 200-250 g) were anesthetized with sodium pentobarbital (60 mg/kg, i.p.), and the abdomen of the rats was opened to expose the liver and pancreas. The pancreas was inflated by injecting Hank's solution into the bile duct, and then the pancreas was cut out with scissors in Hank's solution and minced. After washing the tissue with buffer, the pancreas was digested with collagenase for 10 min, washed, and islet cells separated from debris by the Ficoll gradient method. The isolated islet cell fraction was washed with buffer and the islet cells were hand-selected under a microscope. Islet cells were pre-incubated in 3 mM glucose for 30 minutes, then transferred to medium containing appropriate conditions and incubated for an additional 30 minutes. The content of insulin in the medium was then determined with an ELISA kit (Alpco Diagnostics, Windham, NH).

Compounds identified as inhibitors of PDE11A in the PDE11A inhibition assay described above were tested in this islet cell assay. These compounds were also found to stimulate insulin release by at least 1.5-fold compared to basal insulin release.

in vivo test

Lean rats (Wistar, male, 250-300 g) were fasted overnight and divided into two groups: a matrix-treated group and a compound-treated group (8 rats each). The vehicle or compound was administered by oral gavage (1.5ml/mouse). Two hours later, a glucose solution (30%, 2 g/kg body weight) was injected intraperitoneally into the rats. Tail blood samples were collected at 0, 15, 30, and 60 minutes after the injection of the glucose solution, and blood glucose concentrations were determined with a glucose meter (Bayer Diagnostics, Mishawaka, IN).

Compounds identified in the PDE11A inhibition assay described above and tested in the islet cell assay described above are expected to have hypoglycemic effects when tested in the present method.

The present invention may be embodied in other specific forms without departing from its gist or essential characteristics. The foregoing examples are presented only to illustrate the invention. Accordingly, the scope of the invention is to be limited only by the appended claims.

Claims (17)

1. treatment or prevention are selected from the method for following disease or disease: maturity-onset diabetes (MODY), the invisible Autoimmune Diabetes (LADA) of being grown up, carbohydrate tolerance impaired (IGT), impaired fasting glucose (IFG) (IFG), gestational diabetes and the X metabolic syndrome of diabetes, young morbidity comprise the PDE11A inhibitor to the administration effective dose.

2. the process of claim 1 wherein that described diabetes are type 2 diabetes mellitus.

3. the method for claim 1 also comprises chemical compound, Alpha-glucosidase inhibitor or insulin and described PDE11A inhibitor drug combination with ppar agonist, euglycemic agent, sulfonylureas, insulin succagoga, reduction glycogen output.

4. the method for claim 3, wherein said ppar agonist is selected from rosiglitazone and pioglitazone.

5. the method for claim 3, wherein said sulfonylureas is selected from glibenclamide, glimepiride, chlorpropamide and glipizide.

6. the method for claim 3, wherein said insulin succagoga is selected from GLP-1, GIP, PAC/VPAC receptor stimulating agent, secretin, Nateglinide, meglitinide, repaglinide, glibenclamide, glimepiride, chlorpropamide and glipizide.

7. the method for claim 3, wherein said Alpha-glucosidase inhibitor is selected from acarbose, miglitol and voglibose.

8. the method for claim 3, the chemical compound of wherein said reduction glycogen output is a metformin.

9. the method for claim 1 also comprises HMG-CoA reductase inhibitor, Nicotinicum Acidum, cholic acid chelating agent, fiber acid derivative, antihypertensive drug or appetrol and described PDE11A inhibitor drug combination.

10. the method for claim 9, wherein said appetrol are selected from beta-3 agonist, CB-1 antagonist and lipase inhibitor.

11. the method for the secondary inducement of treatment or prevent diabetes, comprise PDE11A inhibitor, the diabetes that the secondary inducement of wherein said diabetes is selected from that glucocorticoid is excessive, growth hormone is excessive, pheochromocytoma and medicine cause to the administration effective dose.

12. strengthen the method for pancreatic beta cell, comprise PDE11A inhibitor to the administration effective dose to the sensitivity of insulin succagoga.

13. the method for claim 12, wherein said insulin succagoga is selected from GLP-1, GIP, PAC/VPAC receptor stimulating agent, secretin, Nateglinide, meglitinide, repaglinide, glibenclamide, glimepiride, chlorpropamide and glipizide.

14. treatment or the dull-witted method of prevention comprise the PDE11A inhibitor to the administration effective dose.

15. the method for treatment or angiocardiopathy preventing, comprise the PDE11A inhibitor to the administration effective dose, described cardiovascular disease is selected from hypertension, ischemic heart desease, myocardial infarction, stable and unsettled angor, periphery closure disease and ischemic shock.

16. the method for treatment or prevention of urogenital disease, comprise the PDE11A inhibitor to the administration effective dose, described urogenital tract disease is selected from incontinence, stress incontinence, benign prostatic hyperplasia, erection disturbance, female sexual disorder and prostate hyperplasia.

17. the method for claim 16, wherein said female sexual disorder are meant women's sexual excitation obstacle.

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