CN101565408A

CN101565408A - Receptor signal transduction positive modulator, preparation method and purpose thereof

Info

Publication number: CN101565408A
Application number: CNA2008100366101A
Authority: CN
Inventors: 王明伟; 苑芸芸; 周玲; 安德鲁·A·杨; 张翱; 高林东; 苏昊然; 吴茜茜; 汪佳; 王菊
Original assignee: NATIONAL CENTER FOR DRUG SCREENING; Shanghai Institute of Materia Medica of CAS
Current assignee: NATIONAL CENTER FOR DRUG SCREENING; Shanghai Institute of Materia Medica of CAS
Priority date: 2008-04-25
Filing date: 2008-04-25
Publication date: 2009-10-28
Also published as: WO2009129696A1

Abstract

The invention provides a substitute five membered heterocyclic compound represented by the following formula, and salt, ester, solvate and metal composition which are all accepted by pharmacy or a prodrug having the same biological activity. The invention also provides a purpose of the compound as a glucagon-like peptide-1 receptor (GLP-1R) signal transduction positive modulator and an application for preventing and/or curing metabolic diseases (including but not being limited to type-2 diabetes, insulin resistance and adiposity, and the like), angiocardiopathy and neurodegenerative diseases (such as Alzheimer's disease, AD) also called presenile dementia, and the like.

Description

One receptoroid signal transduction synergistic agent, Preparation Method And The Use

Technical field

The present invention relates to substitutive five membered heterocyclic compound and as pancreas hyperglycemia sample peptide-1 receptor (Glucagon-like peptide-1receptor, GLP-1R) characteristic of signal transduction synergistic agent (Positivemodulator), this compounds can strengthen the biological activity and/or the reaction of GLP-1R agonist and render a service.The present invention relates to this compounds and preventing and/or treating metabolic disease (including, but not limited to diabetes B, insulin resistant and obesity etc.), cardiovascular disorder and nerve degenerative diseases [as " alzheimer's disease " (Alzheimer ' s Disease, AD), claim presenile dementia (Alzheimer ' s dementia) again] etc. in medical usage.

Background technology

Carbohydrate metabolism disturbance, particularly diabetes have become the principal disease of modern society's serious threat human health and life.It is predicted that whole world diabetic subject just with the speed increase in every year 6%, has 3.2 hundred million patients (China is 6,000 ten thousand people, occupies second) to the year ends 2006.Diabetes are one group of clinical syndromes that caused by the h and E factor interaction, mainly be divided into 1 type and 2 types, wherein the basic pathology physiology of type 1 diabetes is the absoluteness hypoinsulinism, and clinical treatment is based on supplementation with insulin, so be called insulin-dependent diabetes mellitus again.Diabetes B accounts for more than 95% of ill colony, clinical study finds that most diabetes B patients can synthesize normal even excessive Regular Insulin, but because of the susceptibility reduction (also claim " insulin resistant ") of target cell to Regular Insulin, cause the Regular Insulin relative deficiency, be called non insulin dependent diabetes again.Insulin resistant is the key factor in diabetes B generation and the evolution.The medicine of diabetes B comprises sulfourea, biguanides, euglycemic agent and assist measure etc.After the receptors bind of sulfonylureas drugs for diabetes thing and pancreatic beta cell film, close potassium-channel, the blocking-up efflux of K+ ions causes the cytolemma depolarize, impels Ca ²⁺Channel opener causes the outer flow of calcium ions of born of the same parents, after intracellular free calcium level increases, triggers the release of Regular Insulin.Be divided into for two generations by its priority of coming out, the first-generation such as toluene sulphur third urea, the s-generation comprises Glyburide (glyburide), gliclazide (diamicron), Glipizide (minidiab) and gliquidone (Glurenor) etc.Biguanide antidiabetic medicament energy depress appetite, increase Regular Insulin combines with acceptor, promotes the anaerobic glycolysis of target cell to glucose, suppresses tissue respiration, suppresses the liver glycogen heteroplasia.Mainly contain N1,N1-Dimethylbiguanide, phenformin and buformin etc.Other antidiabetic drugs comprise that mainly thiazolidinediones (Thiazolidinediones) medicine (for example troglitazone, rosiglitazone, pioglitazone etc.), β 3-adrenoceptor conditioning agent, glucagon receptor antagonist, fatty acid metabolism disturb medicine, alpha-glycosidase Depressant (for example acarbose, voglibose, miglitol etc.) and aldose reductase inhibitor etc.Yet these medicines all have various side effects as causing hypoglycemia, put on weight etc., do not have a kind of medicine can stop the depletion of islet function in addition.

Glucagon-like peptide-1 receptor (GLP-1R) belong to the category-B type g protein coupled receptor (Gprotein-coupled receptor, GPCR).When body is taken in nutritive substance, intestines peptide hormone-glucagon-like-peptide-1 (Glucagon like peptide-1 that enteroendocrine cell discharges, GLP-1), by with the GLP-1R high degree of specificity combine and make its activation, stimulate insulin secretion, the generation of glucagon suppression makes the postprandial blood sugar reduction and maintains constant level.Some is by increasing insulin sensitivity for the hypoglycemic activity of GLP-1, improves that insulin resistant realizes.In addition, GLP-1 can also postpone stomach emptying, reduces appetite, causes feeling of repletion.Because the effect that GLP-1 stimulates insulin secretion depends on blood sugar concentration, can hypoglycemia not take place because of continuous release.Other there are some researches show that GLP-1 has proliferation and differentiation that promotes beta Cell of islet and the effect that suppresses apoptosis.External, and the class β cell that GLP-1 can impel embryonic stem cell differentiation to become to have insulin secretion function (JEndocrinol 2005,186:343-52), and the function of performance protection β cell.Because the The above physiological characteristic of GLP-1, be the research focus of many in the world new drug developments mechanism at the antidiabetic medicine exploitation of GLP-1R.

Up to now, the GLP-1R agonist that has gone on the market and carried out clinical study mainly is GLP-1 and polypeptide analog thereof, comprises the GLP-1 derivative L iraglutide of Denmark Novo Nordisk company and the GLP-1 analogue Exenatide of U.S. Amylin pharmaceuticals etc.Yet because polypeptide drugs inconvenience is oral, the effort of seeking non-peptide class GLP-1R agonist is continuing always.Still there is not at present the report that any relevant non-peptide micromolecular GLP-1R agonist successfully is used for clinical treatment.The inventor had had been found that before that the non-peptide micromolecular GLP-1R agonist (PCT/CN2006/001410) of class replacement quaternary ring compound not only had definite external biological activity, and the diabetes B mouse demonstrated good therapeutic action (Proc Natl Acad Sci USA, 2007,104:943-948).

Except that full agonist, the small molecules allosteric type conditioning agent of seeking GPCR also is the new direction in drug research field in recent years.Allosteric type modulator effect another site outside associated receptor endogenic ligand binding site, the activity and/or the maximum effectiveness that can improve endogenic ligand as this conditioning agent then are called positive allosteric type conditioning agent.With full agonist mutually this, positive allosteric type conditioning agent has a lot of advantages.At first, the allosteric type conditioning agent only could produce stirring effect under the situation that endogenic ligand exists and itself is inoperative, and the pharmacologically active that causes thus is more near physiological status; Secondly because its effect depends on the existence of endogenic ligand promptly corresponding to the latter's concentration, therefore reaction has saturability, even also can not cause overstimulation when high dosage, thereby has increased the security of using.The action site of allosteric type agonist has improved the discovery probability of receptor subtype specific compound not at the conservative region of acceptor.In addition, the allosteric type conditioning agent can optionally be strengthened the signal transduction usefulness that endogenic ligand causes, and makes the regulation and control of signal meticulousr, the synergism of performance signal transduction.

Summary of the invention

The object of the present invention is to provide a class by the substituted five-membered heterogeneous ring compound of following general formula (formula 1) expression and pharmaceutically acceptable salt, ester, solvate, metal complexes or have an identical bioactive prodrug;

Having another object of the present invention is to provide a kind of contains by the compound of following general formula (formula 1) expression or its pharmaceutical composition of acceptable salt, ester, solvate or metal complexes pharmaceutically;

Another purpose of the present invention is to provide that pharmaceutically acceptable salt, ester, solvate or metal complexes are as the purposes of GLP-1R signal transduction synergistic agent (including, but not limited to positive allosteric modulators) by the compound of following general formula (formula 1) expression or its, and the reaction that its role is to strengthen the GLP-1R agonist is renderd a service (as still can produce identical exciting effect when reducing agonist concentration) and/or increased maximum effect (still can improve range of reaction as reach saturation concentration at agonist when it occupies all receptor binding sites).

A further object of the present invention is to provide that pharmaceutically acceptable salt, ester, solvate or metal complexes are preventing and/or treating metabolic disease (including, but not limited to diabetes B, insulin resistant and obesity etc.), cardiovascular disorder and nerve degenerative diseases [as " alzheimer's disease " (Alzheimer ' s Disease as GLP-1R signal transduction synergistic agent by the compound of following general formula (formula 1) expression or its, AD), claim presenile dementia (Alzheimer ' s dementia) again] etc. in medical usage.

The invention provides glucagon-like peptide-1 receptor signal transduction synergistic agent, increased and prevented and/or treated metabolic disease (including, but not limited to diabetes B, insulin resistant and obesity etc.), cardiovascular disorder and nerve degenerative diseases [as " alzheimer's disease " (Alzheimer ' sDisease, AD), claim presenile dementia (Alzheimer ' s dementia) again] etc. the member of medicine.The present invention relates to substituted five-membered heterogeneous ring compound by following general formula (formula 1) expression, or acceptable salt, ester, solvate, metal complexes or have identical bioactive prodrug on its pharmacology:

[formula 1]

Described compound comprises the geometrical isomer that they are all.

Wherein Y is O or S;

R ₁, R ₂Be independently of one another H, ethanoyl, phenyl or

Benzyl or

The alkyl of styroyl, diphenyl-methyl, naphthyl or replacement or unsubstituted C1-C10; X wherein ₁, X ₂, X ₃Be N or CH independently of one another; R ₆, R ₇, R ₈, R ₉Be H, F, Cl, Br, CF independently of one another ₃, methyl-formiate base, phenyl, the alkyl of C1-C6, alkylamino radical or the alkoxyl group of C1-C6;

R ₃, R ₄, R ₅Be H, F, C1, Br, NO independently of one another ₂, acetamido, NH ₂Or NR ₁₀R ₁₁, OH or OR ₁₂, SR ₁₃, or the alkyl of replacement or unsubstituted C1-C10; R wherein ₁₀, R ₁₁, R ₁₂, R ₁₃Independently of one another for replacing or unsubstituted C1-C10 alkyl or C2-C6 alkyloyl or C2-C6 carboxylic acid group, amide group or ester group.

Wherein, the alkyl of described unsubstituted C1-C10 is methyl, ethyl, propyl group, sec.-propyl, normal-butyl, n-pentyl, cyclopropyl methyl, cyclopentyl-methyl, cyclohexyl methyl; The alkyl substituent of the C1-C10 that replaces comprises the substituted radical of the alkylamino radical of one or more F of being selected from, cyano group, C1-C10 or alkoxyl group, methyl-formiate base, group-4 ethyl formate.

Preferably, this compounds or its acceptable salt on pharmacology is the form with pharmaceutical composition, or separately, or with pharmacology on acceptable carrier or vehicle unite and provide.The present invention also provides the medicine that comprises above-claimed cpd, be used to prevent and/or treat metabolic disease (including, but not limited to diabetes B, insulin resistant and obesity etc.), cardiovascular disorder and nerve degenerative diseases [as " alzheimer's disease " (Alzheimer ' s Disease, AD), claim presenile dementia (Alzheimer ' s dementia) again] etc.

Again on the one hand, the present invention relates to prevent and/or treat metabolic disease (including, but not limited to diabetes B, insulin resistant and obesity etc.), cardiovascular disorder and nerve degenerative diseases [as " alzheimer's disease " (Alzheimer ' s Disease, AD), claim again presenile dementia (Alzheimer ' sdementia)] etc. method.This method comprises needs or is ready and the object of receiving treatment or preventing give acceptable salt on compound significant quantity, that can optionally strengthen GLP-1R signal transduction effect or its pharmacology, with prevention or treat above-mentioned disease or symptom.Preferably, suffers from above-mentioned metabolic disease (including, but not limited to diabetes B, insulin resistant and obesity etc.), cardiovascular disorder and nerve degenerative diseases are [as " alzheimer's disease " (Alzheimer ' s Disease, AD), claim presenile dementia (Alzheimer ' s dementia) again] etc. any object by giving the compound by following general formula (formula 1) expression of significant quantity with substituted five-membered heterocycle structure, or acceptable salt on its pharmacology, ester, solvate, metal complexes or have identical bioactive prodrug and prevent or treat:

[formula 1]

Wherein Y is O or S;

R ₁, R ₂Be independently of one another H, ethanoyl, phenyl or

Benzyl or

The alkyl of styroyl, diphenyl-methyl, naphthyl or replacement or unsubstituted C1-C10; X wherein ₁, X ₂, X ₃Be N or CH independently of one another; R ₆, R ₇, R ₈, R ₉Be H, F, C1, Br, CF independently of one another ₃, methyl-formiate base, phenyl, the alkyl of C1-C6, alkylamino radical or the alkoxyl group of C1-C6;

R ₃, R ₄, R ₅Be H, F, Cl, Br, NO independently of one another ₂, acetamido, NH ₂Or NR ₁₀R ₁₁, OH or OR ₁₂, SR ₁₃, or the alkyl of replacement or unsubstituted C1-C10; R wherein ₁₀, R ₁₁, R ₁₂, R ₁₃Independently of one another for replacing or unsubstituted C1-C10 alkyl or C2-C6 alkyloyl or C2-C6 carboxylic acid group, amide group or ester group.

On the other hand, the present invention relates to combined preparation, this combined preparation comprises a kind of selectivity enhancing GLP-1R signal transduction effect that has, especially the compound of endogenic ligand agonist activity, or acceptable salt on its pharmacology, or separately, or with pharmacology on acceptable carrier or excipient composition exist.This compound has the structure of following general formula (formula 1):

[formula 1]

Wherein Y is O or S;

R ₁, R ₂Be independently of one another H, ethanoyl, phenyl or

Benzyl or

The invention provides the medicine box that comprises above-mentioned combined preparation.The present invention also further provides the above-mentioned combined preparation of application to be used to prevent and/or treat metabolic disease (including, but not limited to diabetes B, insulin resistant and obesity etc.), cardiovascular disorder and nerve degenerative diseases [as " alzheimer's disease " (Alzheimer ' s Disease, AD), claim presenile dementia (Alzheimer ' s dementia) again] etc., produce the effect that optionally strengthens GLP-1R signal transduction effect, improve patient's clinical symptom and quality of life.

In order to illustrate summary of the invention and not limited to by it, the present invention is divided into following trifle is described in detail.

The A definition

Unless otherwise defined, technology that the present invention is used and scientific term have same meaning with the general understanding of the current techique in field under the present invention.All patents that derive from gene pool and other databases that this place mentions, application, the application of announcement and other publications and sequence are quoted as a reference by comprehensive income.If all patents that derive from gene pool and other databases of definition that this section is illustrated and this patent ginseng usefulness, application, the definition that the application of announcing and other publications and sequence are taken in and quoted is set forth opposite, or when inconsistent, the definition of illustrating with this section is as the criterion.

Used herein, " one " or " one " refers to " at least one " or " one or more ".

Used herein, " positive allosteric modulators " mean a class help the agonist effect, by with acceptor on another site outside the endogenic ligand binding site (Orthosteric site) combine the receptor modulators that plays a role." help the agonist effect " and mean reaction effectiveness (as when reducing agonist concentration, still producing identical exciting effect) that strengthens agonist and/or the binding mode that increases maximum effect (when it occupies all receptor binding sites, still can improve range of reaction) as reach saturation concentration at agonist.

Used herein, " signal transduction synergistic agent " mean a class can strengthen the agonist biological activity, by or not by with acceptor on the endogenic ligand binding site interact and to produce the receptor modulators of pharmacological effect." strengthen the agonist biological activity " including, but not limited to reaction effectiveness (as when reducing agonist concentration, still producing identical exciting effect) that strengthens agonist and/or the binding mode that increases maximum effect (when it occupies all receptor binding sites, still can improve range of reaction) as reach saturation concentration at agonist.

Used herein, " metabolic disease " means metabolism such as sugar, fat or the protein imbalance that is caused by a variety of causes and related symptoms and/or the disease that causes.

Used herein, " diabetes " refer to a kind of metabolic disease of multi-pathogenesis, and characteristics are chronic hyperglycemias, follow because of insulin secretion and/or effect the defective sugar, fat and the protein metabolism disorder that cause.Along with the fall ill prolongation of time of diabetes, the intravital metabolism disorder of body is controlled well as can not get, the chronic complicating diseases that can cause organs such as tissue such as eye, kidney, nerve, blood vessel and heart, so that final take place blind, lower limb are gangrenous, uremia, cerebral apoplexy or myocardial infarction, even threat to life.

Used herein, " insulin resistant " is meant that surrounding tissue is to the susceptibility reduction of Regular Insulin in the body, and target tissues such as muscle, fat promote the effect of glucose uptake that opposing has taken place to Regular Insulin.Insulin resistant is prevalent in the diabetes B, almost accounts for more than 90%, is one of morbidity principal element of diabetes B.

Used herein, " obesity " is meant that the amount of body fat is too much, and 25% or the woman's body weight that man's body weight surpasses ideal body weight surpasses 30% phenomenon of ideal body weight.Inherited genetic factors, hypothalamus sufferer, endocrine regulation, hyperalimentation and activity all are the reason that produces obesity very little.

Used herein, " alzheimer's disease " [Alzheimer ' s Disease, AD claims presenile dementia (Alzheimer ' s dementia) again] be a kind of neural carrying out property transformation disease, the chronic weakening and the chronic of memory that show as intellectual level are clinically lost.

Used herein, " cardiovascular disorder " comprises heart trouble, pulmonary heart disease, hypertension and hyperlipidaemia etc.Characteristics with " sickness rate height, mortality ratio height, disability rate height, recurrence rate height " and " complication is many ".

" significant quantity " that is used for the treatment of the compound of a certain specified disease used herein refers to enough improve or alleviate to a certain extent the amount of the sick symptom that accompanies therewith.This dosage can the single dose administration, also can be according to the treatment plan administration.But this dosage cure diseases, but be typically administration in order to improve this symptom.May need for improving the symptom repeat administration.

Used herein, " acceptable salt, ester or other derivative on the pharmacology " comprises any salt, ester or derivative that those skilled in the art are easy to prepare with currently known methods.The compound of deriving like this and generating can not have toxic action to animal and human's administration.This compound or have pharmaceutical activity, or prodrug.

Used herein, " treatment " refers to that disease and symptom are improved in any way, or other useful changes.Treatment also comprises the application (pharmaceutical applications confuse in methods of treatment) of The compounds of this invention in object.

Used herein, the symptom that gives a certain specified disease of a certain certain drug composition " improvement " is meant any alleviating, and is no matter permanent, interim, over a long time, of short duration, can both owing to this pharmaceutical composition use or relevant with using of this pharmaceutical composition.

Used herein, " pure basically " is meant enough even, can not survey impurity by those skilled in the art for the standard method of analysis of estimating purity and using, described standard method of analysis is just like thin layer chromatography (TLC), gel electrophoresis and high performance liquid chromatography (HPLC).Even perhaps enough pure also refer to be further purified can not change the observable physicochemical property of this material, for example enzymic activity and biological activity.Being used for purifying compounds and making the method for chemical pure compound basically, is known in those skilled in the art.Yet chemical pure basically compound can be the mixture of steric isomer or isomers.In this case, be further purified the specific activity that perhaps can increase compound.

Used herein, " prodrug " or " prodrug " is meant a kind of compound of vivo medicine-feeding, and this compound can be by metabolism, or be converted into biologically, on the pharmacology or the activity form on the therapeutics.In order to make prodrug, pharmaceutical active compounds will be modified, and this active compound is produced by metabolic process again.Prodrug can be designed to change its metabolic stability, or the precursor of transportation characterization, to cover its side effect or toxicity, improves the sense of taste of medicine, or changes other characteristics.Rely on the knowledge of pharmacokinetics and medicine internal metabolism, in case active compound is known on the pharmacology, those skilled in the art just can design the prodrug of this compound.[referring to Medicinal Chemistry ABiochemical Approach, Oxford University Press, New York, 1985, pages388-392].

Term " basically " is identical even or similar, can change to some extent in context the understanding of correlation technique according to those skilled in the art, and be generally at least 70%, is preferably at least 80%, and more preferably at least 90%, most preferably be at least 95% identical.

Here used " composition " refers to any mixture.Can be solution, suspension, liquid, powder, ointment, water-based, nonaqueous or their any combination.

Here used " associating " refers to any associating between two or more.

Term used herein " object " comprises humans and animals, for example, and dog, cat, ox, pig, rodent etc.Experienced implementer should understand object for being suitable for and being ready to metabolic disease (including, but not limited to diabetes B, insulin resistant and obesity etc.), cardiovascular disorder and nerve degenerative diseases [as " alzheimer's disease " (Alzheimer ' s Disease, AD), claim presenile dementia (Alzheimer ' s dementia) again] etc. treat and prevent.

Any protectiveness group used herein, the abbreviation of amino acid and other compounds, consistent with the biochemistry name of their abbreviations general, that generally acknowledge or the promulgation of the IUPAC-IUB council, unless stated otherwise.

B glucagon-like peptide-1 receptor signal transduction synergistic agent

The invention provides the plain sample peptide-1 receptor of glucagon signal transduction synergistic agent, increased and prevented and/or treated metabolic disease (including, but not limited to diabetes B, insulin resistant and obesity etc.), cardiovascular disorder and nerve degenerative diseases [as " alzheimer's disease " (Alzheimer ' sDisease, AD), claim presenile dementia (Alzheimer ' s dementia) again] etc. the member of medicine.The present invention relates to compound with substituted five-membered heterocycle structure by following general formula (formula 1) expression, or acceptable salt, ester, solvate, metal complexes or have identical bioactive prodrug on its pharmacology:

[formula 1]

Described compound comprises the geometrical isomer that they are all.

Wherein Y is O or S;

R ₁, R ₂Be independently of one another H, ethanoyl, phenyl or

Benzyl or

Compound of the present invention can prepare or synthesize according to any suitable method.Preferably, prepare this compound with the synthesis method of quoting as proof in the following F joint.

In addition preferably, acceptable salt provides with the form of pharmaceutical composition on this compound or its pharmacology, and is perhaps independent, perhaps combines with acceptable carrier or vehicle on a kind of pharmacology.

Compound of the present invention can prepare with the form of any suitable acid with acceptable salt on its pharmacology.For example, mineral acid example hydrochloric acid, Hydrogen bromide, nitric acid, sulfuric acid, phosphoric acid etc.; Organic acid such as formic acid, acetate, propionic acid, phenylformic acid, toxilic acid, fumaric acid, succsinic acid, tartrate, citric acid etc.; Alkylsulphonic acid such as methylsulphonic acid, ethylsulfonic acid etc.; Aryl sulfonic acid such as Phenylsulfonic acid, tosic acid etc. all can use.

C treatment and prevention method

The present invention relates to be used to prevent and/or treat metabolic disease (including, but not limited to diabetes B, insulin resistant and obesity etc.), cardiovascular disorder and nerve degenerative diseases [as " alzheimer's disease " (Alzheimer ' s Disease, AD), claim presenile dementia (Alzheimer ' s dementia) again] etc. method.This method comprises needs or is ready the object of receiving treatment or preventing, give significant quantity, optionally strengthen on the compound of GLP-1R signal transduction effect or its pharmacology acceptable salt and treat or prevent above-mentioned disease or symptom.

Preferably, above-mentioned disease by give significant quantity by acceptable salt, ester, solvate, metal complexes on the compound with substituted five-membered heterocycle structure of following general formula (formula 1) expression or its pharmacology or have identical bioactive prodrug and treat or prevent:

[formula 1]

Described compound comprises the geometrical isomer that they are all.

Wherein Y is O or S;

R ₁, R ₂Be independently of one another H, ethanoyl, phenyl or Benzyl or

Can prevent and treat any object with present method, preferred mammal, more preferably people.

Present method can be used to prevent and treat metabolic disease (including, but not limited to diabetes B, insulin resistant and obesity etc.), cardiovascular disorder and nerve degenerative diseases [as " alzheimer's disease " (Alzheimer ' s Disease, AD), claim presenile dementia (Alzheimer ' s dementia) again] etc.Preferred disease or symptom are any disease or symptoms that is caused or followed by insulin secretion and/or dysfunction.

When preventing and/or treating metabolic disease (including, but not limited to diabetes B, insulin resistant and obesity etc.), can use separately or comprise GLP-1R agonist, GLP-1 analogue with other Remedies for diabetes that maybe will go on the market of having gone on the market and/or act on and increase the active medicine of endogenous GLP-1 and unite and use compound of the present invention.The GLP-1R agonist comprises GLP-1 polypeptide and non-polypeptide analog etc. in a specific embodiment.In another specific embodiment, comprise all kinds of pepxs (Dipeptidyl peptidase-IV, DPP-IV) inhibitor, including, but not limited to PCT patent application WO 2005/075426,2006/011035,2006/040625 and United States Patent (USP) 7,205,323 and 7, the compound of indication in 230,002 grades.Any suitable metabolic disease (including, but not limited to diabetes B, insulin resistant and obesity etc.) medicine all can be united use with compound of the present invention.

In a preferred embodiment of the invention, compound of the present invention can comprise that the DPP-IV inhibitor unites use with the medicine that the GLP-1 agonist maybe can increase endogenous GLP-1 level.More preferably, the medicine that maybe can increase endogenous GLP-1 level with compound of the present invention and the above-mentioned GLP-1 agonist that maybe will go on the market of having gone on the market comprises that the DPP-IV inhibitor is united and uses and then reduce because of the too high toxic side effects that causes of the former dosage or treatment and use disease or the symptom that the former can not effectively control separately.

In another preferred embodiment, give above-mentioned euglycemic agent when using The compounds of this invention.More preferably, adopt compounds for treating of the present invention or prevention because of using the above-mentioned Remedies for diabetes (comprising euglycemic agent) that maybe will go on the market that gone on the market to develop immunity to drugs or caused disease of toxic side effects or symptom.

Can be by any suitable method separately with compound administration of the present invention, or with the GLP-1R agonist, or comprise the DPP-IV inhibitor, or comprise that with other suitable Remedies for diabetes euglycemic agent unites use with the medicine that can increase endogenous GLP-1 level.For example, can pass through intracavitary administration, subcutaneous injection, intravenous injection, intramuscularly, the intradermal injection, oral or local with compound administration of the present invention, or with acceptable salt administration on its pharmacology.

In specific embodiments, present method further comprises the disease of administration object or symptom is diagnosed and prognosis evaluation.Can use any suitable method to be used for diagnosis and assessment relative disease or symptom and prognosis thereof.Diagnosis and prognosis can be based on the substance in vivo that detects and/or identify any or all, for example glycolated hemoglobin, enzyme, antigen, antibody, nucleic acid or other pathologic and clinical marker thing etc. and related symptoms.For example, the diagnosis or the method for prognosis that can use international monopoly WO 01/44815 and United States Patent (USP) 5,571,674 to disclose.

The D combined preparation, the method for medicine box and drug combination

On the other hand, the present invention also relates to combined preparation, this associating comprises that a kind of selectivity strengthens the compound of GLP-1R signal transduction effect, or acceptable salt and one or more metabolic disease medicines comprise euglycemic agent on its pharmacology.

Preferably, this drug combination comprises that acceptable salt comprises the GLP-1R agonist with one or more metabolic disease medicines on The compounds of this invention or its pharmacology, the medicine that maybe can increase endogenous GLP-1 level comprises the DPP-IV inhibitor, and this compound is represented by following general formula (formula 1):

[formula 1]

Described compound comprises the geometrical isomer that they are all.

Wherein Y is O or S;

R ₁, R ₂Be independently of one another H, ethanoyl, phenyl or

Benzyl or

In combined preparation of the present invention, can use any suitable Remedies for diabetes to comprise the GLP-1R agonist, maybe can increase the medicine such as the DPP-IV inhibitor of endogenous GLP-1 level.In a particular, be used for combined preparation of the present invention and can comprise one or more of above-mentioned Remedies for diabetes.

In another particular, the invention provides that a kind of treatment or prevention are caused by insulin secretion and/or dysfunction or the disease followed or the method for symptom, this method comprises needs and is ready to receive treatment or the object that prevents gives the above-mentioned combined preparation of significant quantity, or acceptable salt on its pharmacology, thereby treat or prevent above-mentioned disease or symptom.

In another particular, the invention provides a medicine box, comprising acceptable salt on compound of the present invention or its pharmacology and use above-claimed cpd or its pharmacology on acceptable salt prevent and treat by insulin secretion and/or dysfunction and cause or the disease followed or the operation instruction of symptom.

In a further embodiment, the invention provides a medicine box, comprising above-mentioned combined preparation and use treatment of described combined preparation or prevention to cause or the disease followed or the operation instruction of symptom by insulin secretion and/or dysfunction.

E prescription and dosage

According to the present invention, compound of the present invention, separately or with other medicament, carrier or vehicle associating, for any suitable route of administration is formulated preparation, for example intracavitary administration, subcutaneous injection, intravenous injection, intramuscularly, intradermal are injected, oral or local application.Present method can be used injecting and administering preparations, with the form of single dose at ampoule, or in the multi-dose container with the buffer reagent drug administration by injection that adds.Preparation can be taked following form such as suspension, solution or the emulsion in oiliness or aqueous media.Preparation can contain prescription reagent such as suspensoid, stablizer and/or dispersion agent.In addition, before the use, activeconstituents can powder type and suitable carriers, aseptic no heat source water or other solvents formation formulation.Local application of the present invention can adopt foam, gel, ointment, ointment, transdermal patch, or paste.

Operable medicinal compositions and the method that is used for administration includes, but are not limited among the present invention, United States Patent (USP) 5,736,154,6,197,801B1,5,741,511,5,886,039,5,941,868,6,258,374B1 and 5,686,102 contents of being set forth.

The dosage size of treatment or prevention is the seriousness of feelings and route of administration and change to some extent due to illness.Dosage can react different because of age, body weight, healthy state and individual patient with the medication frequency.It is to be noted (the diagnosis and treatment doctor also should know), according to toxicity and side reaction, the termination of must taking the necessary measures, interruption or reduction therapeutic dose.On the contrary, if clinical response not obvious (getting rid of toxicity and side reaction), the doctor should suitably adjust treatment plan, improves dosage.

Any suitable route of administration all may be utilized.Formulation comprises tablet, lozenge, beans shape capsule, dispersion agent, suspension agent, solution, capsule, patch and analogue etc.

In actual applications, compound of the present invention is united separately or with other preparations, can be according to general pharmacology hybrid technology and pharmaceutical carrier or vehicle, and for example beta-cyclodextrin and 2-hydroxypropyl-beta-cyclodextrin are closely mixed.According to the needs of dispensing, can adopt the special carrier of universal support, part or parenteral route.Prepare non-parenteral dosage forms, for example intravenous injection or the composition inculcated can adopt similar drug media (carrier), water as known to those skilled in the art, ethylene glycol, oil, buffer reagent, sugar, sanitas, liposome etc.The example of this non-enteron aisle composition comprises, but is not restricted to dextrose, physiological saline or other solution of 5%W/V.The total dose of compound of the present invention, separately or and other preparation Combined Preparation, the administration of available bottle intravenous fluid, volume is approximately from 1 milliliter to 2000 milliliters.According to the total dose of administration, the dilution liquid measure also can be different.

The present invention also provides the medicine box of realizing treatment plan.This medicine box is united the The compounds of this invention of effective dose separately or with other reagent with acceptable form on the pharmacology, is included in one or more containers.Preferably medicament forms is and Sterile Saline, dextrose solution, and buffered soln, or other drug is learned upward, and acceptable sterile liquid share.Perhaps, composition can be by freeze-drying or drying; In this case, medicine box is randomly further with acceptable solution on a kind of pharmacology, and preferred aseptic solution is included in the container, is formed for injecting the solution of purpose to reformulate mixture.Acceptable solution is physiological saline and dextrose solution on the typical pharmacology.

In another embodiment, medicine box of the present invention further comprises and is used for the preferred with the pin of sterile form packing or the alcohol pads of syringe and/or packing of injectable composition.Can randomly comprise specification sheets for doctor or patient's use.

The F preparation method

GLP-1R signal transduction synergistic agent of the present invention can make by following reactions steps:

Step 1: by chemical equation 1 preparation thiocarbamide.

[chemical equation 1]

With ethanol or 1, the 4-dioxane is a solvent, reflux, and after the reaction solution cooling, target product 3 is separated out;

Step 2: by chemical equation 2 preparation thiazolidones.

[chemical equation 2]

With ethanol is solvent, and under TEBA catalysis reflux, the filtered while hot reaction solution carries out recrystallization after filtrate concentrates, target product 4;

Step 3: carry out aldol condensation by chemical equation 3.

[chemical equation 3]

Under the piperidines effect, make

compound

4 and 5 back flow reaction in dehydrated alcohol, reaction solution carries out recrystallization or column chromatography after cooling off, concentrating, and gets final product 6.

Step 4: undertaken thiocarbonyl groupization by chemical equation 4.

[chemical equation 4]

Compound 6 and Lawesson ' s reagent reacting by heating in toluene.Reaction solution cooling, concentrated is carried out column chromatography and is got target product 7.

In above-mentioned four-step reaction:

R wherein ₁, R ₂Be independently of one another H, ethanoyl, phenyl or

Benzyl or

Usually follow the tracks of the degree of carrying out of detection reaction with TLC (thin layer chromatography).After reaction finished, the general post-treating method that adopts comprised cooling, concentrating under reduced pressure, extraction, recrystallization, column chromatography for separation etc.Final product is identified with HPLC, ESI-MS and NMR.

Description of drawings

Fig. 1 a. reporter gene method detects the synergism of The compounds of this invention 6Cu to the GLP-1 agitated reaction.The concentration gradient of GLP-1 is 100,10,1,0.1,0.01,0.001 and 0.0001nM, is 100% with 100nM institute inductive luciferase activity, and the concentration gradient of compound 6Cu is 0.3,1,3 and 10 μ M.The result shows that the excitement of GLP-1 is renderd a service and maximum agonist activity all is significantly improved but the activating effect of the plain enzyme of HEK293 ICF of compound 6Cu dose-dependently ground enhancing GLP-1 inductive, expression humanized GLP-1R shows as.The Dark grey zone is that the signal of 6Cu amplifies effect with top.First area (light gray) is the dosage range of antidiabetic effect; Second area (black) is the dosage range of anti-obesity effect; The 3rd zone (redness) is single with all inaccessiable effect scope of any dosage GLP-1.

Fig. 1 b. reporter gene method detects the synergism of The compounds of this invention 6Cu to the Exendin-4 agitated reaction.The concentration gradient of Exendin-4 (GLP-1 polypeptide analog) is 100,10,1,0.1,0.01,0.001 and 0.0001nM, is 100% with 100nM GLP-1 institute inductive luciferase activity, and the concentration gradient of compound 6Cu is 0.3,1,3 and 10 μ M.The result shows, but compound 6Cu dose-dependently ground strengthens the activating effect of the plain enzyme of HEK293 ICF of Exendin-4 inductive, expression humanized GLP-1R, and the excitement effectiveness and the maximum agonist activity that show as Exendin-4 all are significantly improved.The Dark grey zone is that the signal of 6Cu amplifies effect with top.First area (light gray) is the dosage range of antidiabetic effect; Second area (black) is the dosage range of anti-obesity effect; The 3rd zone (redness) is single with any dosage GLP-1 and all inaccessiable effect scope of Exendin-4.

Fig. 1 c. reporter gene method detects the synergism of The compounds of this invention 6Cu to the Boc5 agitated reaction.The concentration gradient of Boc5 (non-peptide micromolecular GLP-1R agonist) is 100,30,10,3,1,0.3 and 0.1M, is 100% with the GLP-1 institute inductive luciferase activity of 100nM, and the 6Cu concentration gradient is 0.3,1,3 and 10 μ M.The result shows, but compound 6Cu dose-dependently ground strengthens the activating effect of the plain enzyme of HEK293 ICF of Boc5 inductive, expression humanized GLP-1R, and the excitement effectiveness and the maximum agonist activity that show as Boc5 all are significantly improved.The Dark grey zone is that the signal of 6Cu amplifies effect with top.First area (light gray) is the dosage range of antidiabetic effect; Second area (black) is the dosage range of anti-obesity effect; The 3rd zone (redness) is single with any dosage Boc5, GLP-1 and all inaccessiable effect scope of Exendin-4.

Fig. 2. receptor competition is in conjunction with the avidity of test method detection compound 6Cu to GLP-1R.The concentration gradient of compound 6Cu is 75,25,8.3,2.77,0.92,0.31,0.102 and 0 μ M.The result shows, 6Cu can be specifically with [ ¹²⁵I] GLP-1 of mark _(7-36)Competitive in conjunction with GLP-1R (left figure), its IC ₅₀Value is 3.52 μ M.When maximum concentration was 75 μ M, it was 36.12% to GLP-1's in conjunction with inhibiting rate.6Cu can also be specifically with [ ¹²⁵I] Exendin of mark _(9-39)(GLP-1R peptide agonist) is competitive in conjunction with GLP-1R (right figure), its IC ₅₀Value is 15.96 μ M.When maximum concentration was 75 μ M, it was to Exendin _(9-39)Be 43.2% in conjunction with inhibiting rate.

Fig. 3 .C57BL/6J chmice acute is ingested and is suppressed the influence of experiment detection compound 6Cu to appetite.Abdominal injection solvent carrier or 0.1,0.3,1 or the 6Cu of 3mg is tried behind the Foodball of mouse (fasting overnight) known weight to continue observation 2 hours every 15 minutes weighing food weights and record.The result shows, compound 6Cu can dose-dependently ground reduces the food ration of mouse, and wherein 3mg dosage group is compared with the solvent carrier group and had significant significant difference, the inhibition of the calculating gained median effective dose (ED that ingests ₅₀) be 0.9mg.

Embodiment

Laboratory apparatus and reagent

The HP1100HPLC system is equipped with binary gradient pump, online vacuum degassing machine, automatic sampler, column oven and photodiode array detector.Chromatographic column is ZORBAX SB-C18 (2.1 * 150mm, 3.5 μ m), and moving phase is acetonitrile: water is 65: 35, and flow velocity is 0.2 milliliter of a per minute, and the detection wavelength is 254nm.NMR is recorded by Varian Mercury-300 type nuclear magnetic resonance analyser that (solvent is CDCl ₃, CD ₃OD or DMSO-d ₆); ESI-MS is recorded by AB Mariner type mass spectrograph.Raw materials usedly in synthetic be commercially available prod (available from Shanghai reagent company or Alfa Aesa Tianjin company) except that specializing the source, product is through recrystallization or column chromatography purification, silica gel (200-300 order, Haiyang Chemical Plant, Qingdao).

The present invention is further elaborated for following specific embodiment, but do not limit the present invention.

The preparation of embodiment 1. compound 6Aa

[chemical equation 5]

According to top chemical reaction:

The first step: in the 100mL there-necked flask, add 1.46g (0.01mol) CH ₃NCS, 2.5g (0.0098mol) p-Chlorobenzoic acid amide and 20mL dehydrated alcohol are stirred to the solid dissolving, reflux, the reaction of TLC tracking monitor.After reaction finishes, reaction solution is placed 4 ℃ of refrigerator cooling crystallizations, suction filtration, drying gets 3.4g white solid 3a (yield 87%).

Second step: in the 100mL there-necked flask, add 3.4g (0.0169mol) compound 3a, 4.8g (0.0508mol) Mono Chloro Acetic Acid, 4.17g (0.0508mol) sodium-acetate, a small amount of TEBA and 20mL dehydrated alcohol.Reflux, the reaction of TLC tracking monitor.After reaction finishes, filtered while hot, filtrate decompression is concentrated into smaller size smaller, in 4 ℃ of refrigerator cooling crystallizations, suction filtration, drying gets 4.32g white solid 4a (yield 96%).

The 3rd step: in the 10mL there-necked flask, add 60mg (0.25mmol) compound 4a, 66mg (0.3mmol) 5a, 0.1mL piperidines and 3mL dehydrated alcohol, reflux, the reaction of TLC tracking monitor.After reaction finishes, cooling crystallization.Suction filtration, filter cake washs successively with ethanol, ethyl acetate, and drying gets 58.9mg safran solid 6Aa (yield 53.2%).

Compound 6Aa: ¹H NMR (300MHz, CDCl ₃) δ 7.817 (s, 1H); 7.648 (s, 1H); 7.444 (d, 1H, J=9.0Hz); 7.360 (d, 2H, J=7.2Hz); 6.959 (d, 2H, J=7.2Hz); 6.910 (d, 1H, J=9.0Hz); 3.437 (s, 3H); 3.261 (m, 4H); 2.004 (m, 4H).

The preparation of embodiment 2. compound 6Ba

[chemical equation 6]

According to top chemical reaction:

The first step: replace respectively among the embodiment 1 preparing compound 3b with the method that first step is identical among the embodiment 1 outside the 1a in the first step, the 2a divided by raw material 1b, 2b.

Second step: replace respectively among the embodiment 1 preparing compound 4b with the method that second step is identical among the embodiment 1 outside the 3a in second step divided by raw material 3b.

The 3rd step: replace respectively among the embodiment 1 preparing compound 6Ba with the method that third step is identical among the embodiment 1 outside the 4a in the third step, the 5a divided by raw material 4b, 5b.

Compound 6Ba: ¹H NMR (300MHz, DMSO-d ₆) δ 0.930 (t, 3H, J=7.35Hz), 1.358 (sex, 2H, J=7.5Hz), 1.691 (qu, 2H, J=7.35Hz), 2.283 (s, 6H), 3.892 (t, 2H, J=6.9Hz), 7.055 (d, 2H, J=8.4Hz), 7.393 (d, 1H, J=8.4Hz), 7.457 (m, 4H), 7.761 (s, 1H).

The preparation of embodiment 3. compound 6Ca

[chemical equation 7]

According to top chemical reaction:

The first step: in the 50mL there-necked flask, add 1.7g (22mmol) NH ₄SCN and 10mL acetone are stirred to the solid dissolving under the room temperature, slowly drip 2.82g (20mmol) Benzoyl chloride, white solid occurs, solution turned yellow, and thickening is thick.After dripping end, reflux half an hour, in reaction system, slowly drip acetone (10mL) solution of 2.551g (20mmol) p-Chlorobenzoic acid amide 2a again.After dripping, continue to reflux the reaction of TLC tracking monitor.After reaction finished, cooling was poured reaction solution in the 150mL water into, stirred, and separated out yellow solid.Suction filtration is dissolved in solid in the 3g NaOH aqueous solution (27mL), boiled 5 minutes, and cooling, suction filtration gained solid gets target product 3c 2.98g (yield 80%).

Second step: replace respectively among the embodiment 1 preparing compound 4c with the method that second step is identical among the embodiment 1 outside the 3a in second step divided by raw material 3c.

The 3rd step: replace respectively among the embodiment 1 preparing compound 6Ca with the method that third step is identical among the embodiment 1 outside the 4a in the third step, the 5a divided by raw material 4c, 5b.

Compound 6Ca: ¹H NMR (300MHz, CD ₃OD-d ₄) δ 7.76 (s, 1H); 7.64 (d, 1H, J=6.9Hz); 7.38 (m, 5H); 7.02 (d, 1H, J=8.1Hz); 2.33 (m, 6H)

The preparation of embodiment 4. compound 6Ab

Divided by with the corresponding aldehyde of compound 6Ab be outside the raw material, prepare compound 6Ab with the method identical with embodiment 1.

The structure of compound 6Ab:

Compound 6Ab: ¹H NMR (300MHz, CDCl ₃) δ 7.737 (s, 1H); 7.547 (d, 2H, J=6Hz); 7.143 (m, 4H); 6.953 (d, 2H, J=6Hz); 3.446 (s, 3H); 2.304 (s, 3H).

The preparation of embodiment 5. compound 6Ac

Divided by with the corresponding aldehyde of compound 6Ac be outside the raw material, prepare compound 6Ac with the method identical with embodiment 1.

The structure of compound 6Ac:

Compound 6Ac: ¹H NMR (300MHz, DMSO-d ₆) δ 9.770 (s, 1H); 7.674 (s, 1H); 7.447 (d, 2H, J=9.0Hz); 7.164 (s, 1H); 7.046 (d, 2H, J=8.4Hz); 6.899 (m, 2H); 4.037 (q, 2H, J=6.9Hz); 3.301 (s, 3H); 1.301 (t, 3H, J=6.9Hz).

The preparation of embodiment 6. compound 6Ad

Divided by with the corresponding aldehyde of compound 6Ad be outside the raw material, prepare compound 6Ad with the method identical with embodiment 1.

The structure of compound 6Ad:

Compound 6Ad: ¹H NMR (300MHz, CDCl ₃) δ 7.708 (s, 1H); 7.330 (m, 3H); 6.943 (m, 5H); 3.450 (s, 3H).

The preparation of embodiment 7. compound 6Ae

Divided by with the corresponding aldehyde of compound 6Ae be outside the raw material, prepare compound 6Ae with the method identical with embodiment 1.

The structure of compound 6Ae:

Compound 6Ae: ¹H NMR (300MHz, CDCl ₃) δ 7.672 (s, 1H); 7.350 (d, 2H, J=8.4Hz); 6.960 (m, 5H); 3.918 (s, 3H); 3.434 (s, 3H).

The preparation of embodiment 8. compound 6Af

Divided by with the corresponding aldehyde of compound 6Af be outside the raw material, prepare compound 6Af with the method identical with embodiment 1.

The structure of compound 6Af:

Compound 6Af: ¹H NMR (300MHz, CDCl ₃) δ 10.388 (s, 1H); 7.717 (d, 2H, J=8.7Hz); 7.673 (s, 1H); 7.463 (d, 2H, J=8.4Hz); 7.451 (d, 2H, J=8.4Hz); 7.050 (d, 2H, J=9.0Hz); 3.365 (s, 3H); 2.059 (s, 3H).

The preparation of embodiment 9. compound 6Ag

Divided by with the corresponding aldehyde of compound 6Ag be outside the raw material, prepare compound 6Ag with the method identical with embodiment 1.

The structure of compound 6Ag:

Compound 6Ag: ¹H NMR (300MHz, CDCl ₃) δ 7.705 (s, 1H); 7.353 (d, 2H, J=8.4Hz); 7.345 (d, 2H, J=8.4Hz); 7.272 (d, 2H, J=8.1Hz); 6.957 (d, 2H, J=8.4Hz); 3.442 (s, 3H); 2.978 (q, 2H, J=7.5Hz); 1.343 (t, 3H, J=7.5Hz).

The preparation of embodiment 10. compound 6Ah

Divided by with the corresponding aldehyde of compound 6Ah be outside the raw material, prepare compound 6Ah with the method identical with embodiment 1.

The structure of compound 6Ah:

Compound 6Ah: ¹H NMR (300MHz, CDCl ₃) δ 7.770 (s, 1H); 7.666 (d, 2H, J=8.4Hz); 7.545 (d, 2H, J=8.4Hz); 7.363 (d, 2H, J=8.1Hz); 6.949 (d, 2H, J=8.4Hz); 3.495 (s, 3H).

The preparation of embodiment 11. compound 6Ai

Divided by with the corresponding aldehyde of compound 6Ai be outside the raw material, prepare compound 6Ai with the method identical with embodiment 1.

The structure of compound 6Ai:

Compound 6Ai: ¹H NMR (300MHz, DMSO-d ₆) δ 9.687 (brs, 1H); 9.408 (brs, 1H); 7.571 (s, 1H); 7.458 (d, 2H, J=8.7Hz); 7.049 (d, 2H, J=2.1Hz); 6.960 (m, 2H); 6.814 (m, 1H); 3.297 (s, 3H).

The preparation of embodiment 12. compound 6Aj

Divided by with the corresponding aldehyde of compound 6Aj be outside the raw material, prepare compound 6Aj with the method identical with embodiment 1.

The structure of compound 6Aj:

Compound 6AJ: ¹H NMR (300MHz, DMSO-d ₆) δ 10.324 (s, 1H); 7.647 (s, 1H); 7.463 (d, 2H, J=8.4Hz); 7.184 (d, 1H, J=8.4Hz); 7.095 (m, 1H); 7.054 (m, 4H); 3.330 (s, 3H).

The preparation of embodiment 13. compound 6Ak

Divided by with the corresponding aldehyde of compound 6Ak be outside the raw material, prepare compound 6Ak with the method identical with embodiment 1.

The structure of compound 6Ak:

Compound 6Ak: ¹H NMR (300MHz, CDCl ₃) δ 7.716 (s, 1H); 7.487 (m, 5H); 7.044 (m, 3H); 4.468 (s, 2H); 3.309 (s, 3H).

The preparation of embodiment 14. compound 6Al

Divided by with the corresponding aldehyde of compound 6Al be outside the raw material, prepare compound 6Al with the method identical with embodiment 1.

The structure of compound 6Al:

Compound 6Al: ¹H NMR (300MHz, CDCl ₃) δ 8.018 (d, 1H, J=2.4Hz); 7.686 (s, 1H); 7.500 (d, 1H, J=2.1Hz); 7.377 (d, 2H, J=8.7Hz); 6.949 (d, 2H, J=8.7Hz); 3.469 (s, 3H); 2.372 (s, 3H); 2.355 (s, 3H).

The preparation of embodiment 15. compound 6Am

Divided by with the corresponding aldehyde of compound 6Am be outside the raw material, prepare compound 6Am with the method identical with embodiment 1.

The structure of compound 6Am:

Compound 6Am: ¹H NMR (300MHz, DMSO-d ₆) δ 7.487 (s, 1H); 7.435 (d, 2H, J=8.4Hz); 7.048 (d, 2H, J=8.4Hz); 7.009 (d, 2H, J=2.4Hz); 6.744 (d, 2H, J=2.4Hz); 3.628 (s, 3H); 3.262 (s, 3H).

The preparation of embodiment 16. compound 6An

Divided by with the corresponding aldehyde of compound 6An be outside the raw material, prepare compound 6An with the method identical with embodiment 1.

The structure of compound 6An:

Compound 6An: ¹H NMR (300MHz, CDCl ₃) δ 7.629 (s, 1H); 7.339 (d, 2H, J=8.7Hz); 6.977 (m, 3H); 6.875 (d, 1H, J=2.1Hz); 6.626 (d, 1H, J=8.4Hz); 4.254 (t, 2H, J=4.35Hz); 3.415 (s, 3H); 3.359 (t, 2H, J=4.5Hz); 2.948 (s, 3H).

The preparation of embodiment 17. compound 6Ao

Divided by with the corresponding aldehyde of compound 6Ao be outside the raw material, prepare compound 6Ao with the method identical with embodiment 1.

The structure of compound 6Ao:

Compound 6Ao: ¹H NMR (300MHz, DMSO-d ₆) δ 7.729 (s, 1H); 7.451 (d, 2H, J=8.4Hz); 7.380 (brs, 1H); 7.333 (brs, 1H); 7.250 (m, 1H); 7.054 (d, 2H, J=8.4Hz); 7.019 (m, 1H); 6.969 (d, 1H, J=8.4Hz); 4.468 (s, 2H); 3.779 (s, 3H); 3.296 (s, 3H).

The preparation of embodiment 18. compound 6Bb

Divided by with the corresponding amine of compound 6Bb be outside the raw material, prepare compound 6Bb with the method identical with embodiment 2.

The structure of compound 6Bb:

Compound 6Bb: ¹H NMR (300MHz, DMSO-d ₆) δ 2.292 (s, 6H), 7.012 (d, 2H, J=8.4Hz), 7.471 (m, 5H), 7.625 (m, 4H), 7.832 (s, 1H).

The preparation of embodiment 19. compound 6Bc

Divided by with the corresponding amine of compound 6Bc be outside the raw material, prepare compound 6Bc with the method identical with embodiment 2.

The structure of compound 6Bc:

Compound 6Bc: ¹H NMR (300MHz, DMSO-d ₆) δ 1.507 (d, 6H, J=6.9Hz), 2.281 (s, 6H), 4.903 (sep, 1H, J=6.9Hz), 7.056 (d, 2H, J=8.4Hz), 7.377 (m, 5H), 7.713 (s, 1H).

The preparation of embodiment 20. compound 6Bd

Divided by with the corresponding amine of compound 6Bd be outside the raw material, prepare compound 6Bd with the method identical with embodiment 2.

The structure of compound 6Bd:

Compound 6Bd: ¹H NMR (300MHz, DMSO-d ₆) δ 1.095 (m, 5H), 1.660 (m, 6H), 2.283 (s, 6H), 3.754 (d, 2H, J=7.2Hz), 7.043 (d, 2H, J=8.7Hz), 7.440 (m, 5H), 7.759 (s, 1H).

The preparation of embodiment 21. compound 6Be

Divided by with the corresponding amine of compound 6Be be outside the raw material, prepare compound 6Be with the method identical with embodiment 2.

The structure of compound 6Be:

Compound 6Be: ¹H NMR (300MHz, DMSO-d ₆) δ 2.383 (s, 6H), 2.500 (s, 3H), 5.171 (s, 2H), 7.095 (d, 2H, J=8.7Hz), 7.269 (m, 4H), 7.550 (m, 5H), 7.916 (s, 1H).

The preparation of embodiment 22. compound 6Bf

Divided by with the corresponding amine of compound 6Bf be outside the raw material, prepare compound 6Bf with the method identical with embodiment 2.

The structure of compound 6Bf:

Compound 6Bf: ¹H NMR (300MHz, DMSO-d ₆) δ 0.420 (m, 2H), 0.523 (d, 2H, J=7.2Hz), 1.321 (m, 1H), 2.290 (s, 6H), 3.781 (d, 2H, J=6.9Hz), 7.078 (d, 2H, J=8.7Hz), 7.403 (d, 1H, J=7.5Hz), 7.473 (m, 4H), 7.788 (s, 1H).

The preparation of embodiment 23. compound 6Bg

Divided by with the corresponding amine of compound 6Bg be outside the raw material, prepare compound 6Bg with the method identical with embodiment 2.

The structure of compound 6Bg:

Compound 6Bg: ¹H NMR (300MHz, DMSO-d ₆) δ 1.940 (qu, 2H, J=6.45Hz), 2.284 (s, 6H), 3.222 (s, 3H), 3.412 (t, 2H, J=6.0Hz), 3.957 (t, 2H, J=6.9Hz), 7.058 (d, 2H, J=8.7Hz), 7.394 (d, 1H, J=8.7Hz), 7.473 (m, 4H), 7.762 (s, 1H).

The preparation of embodiment 24. compound 6Bh

Divided by with corresponding amine of compound 6Bh and aldehyde be outside the raw material, prepare compound 6Bh with the method identical with embodiment 2.

The structure of compound 6Bh:

Compound 6Bh: ¹H NMR (300MHz, DMSO-d ₆) δ 2.211 (s, 6H), 2.596 (s, 2H), 3.965 (s, 2H), 6.821 (d, 1H, J=8.4Hz), 6.922 (m, 2H), 7.051 (d, 2H, J=8.7Hz), 7.472 (d, 2H, J=8.4Hz), 7.575 (s, 1H), 9.648 (brs, 2H).

The preparation of embodiment 25. compound 6Bi

Divided by with the corresponding amine of compound 6Bi be outside the raw material, prepare compound 6Bi with the method identical with embodiment 2.

The structure of compound 6Bi:

Compound 6Bi: ¹H NMR (300MHz, DMSO-d ₆) δ 1.269 (t, 3H, J=7.05Hz), 2.288 (s, 6H), 3.936 (q, 2H, J=6.9Hz), 7.084 (d, 2H, J=8.7Hz), 7.401 (d, 1H, J=8.7Hz), 7.472 (m, 4H), 7.773 (s, 1H).

The preparation of embodiment 26. compound 6Bj

Divided by with corresponding amine of compound 6Bj and aldehyde be outside the raw material, prepare compound 6Bj with the method identical with embodiment 2.

The structure of compound 6Bj:

Compound 6Bj: ¹H NMR (300MHz, DMSO-d ₆) δ 3.728 (s, 6H), 3.786 (s, 3H), 4.993 (s, 2H), 6.939 (m, 3H), 7.047 (m, 5H), 7.475 (d, 2H, J=8.4Hz), 7.650 (s, 1H).

The preparation of embodiment 27. compound 6Bk

Divided by with the corresponding amine of compound 6Bk be outside the raw material, prepare compound 6Bk with the method identical with embodiment 2.

The structure of compound 6Bk:

Compound 6Bk: ¹H NMR (300MHz, DMSO-d ₆) δ 1.220 (m, 3H), 2.288 (s, 6H), 4.197 (q, 2H, J=6.3Hz), 4.692 (s, 2H), 7.016 (d, 2H, J=6.0Hz), 7.476 (m, 5H), 7.849 (s, 1H).

The preparation of embodiment 28. compound 6Bl

Divided by with corresponding amine of compound 6Bl and aldehyde be outside the raw material, prepare compound 6Bl with the method identical with embodiment 2.

The structure of compound 6Bl:

Compound 6Bl: ¹H NMR (300MHz, DMSO-d ₆) δ 5.101 (s, 2H), 6.812 (d, 1H, J=8.7Hz), 6.938 (m, 2H), 7.025 (d, 2H, J=8.4Hz), 7.353 (d, 2H, J=6.0Hz), 7.459 (d, 2H, J=9.0Hz), 7.632 (s, 1H), 8.547 (d, 2H, J=5.7Hz).

The preparation of embodiment 29. compound 6Cb

Divided by with corresponding amine of compound 6Cb and aldehyde be outside the raw material, prepare compound 6Cb with the method identical with embodiment 1.

The structure of compound 6Cb:

Compound 6Cb: ¹H NMR (300MHz, CDCl ₃) δ 7.70 (s, 1H); 7.35 (m, 3H); 7.25 (m, 2H); 6.95 (d, 2H, J=8.1Hz); 3.45 (s, 3H); 2.31 (s, 3H); 2.29 (s, 3H).

The preparation of embodiment 30. compound 6Cc

Divided by with corresponding amine of compound 6Cc and aldehyde be outside the raw material, prepare compound 6Cc with the method identical with embodiment 1.

The structure of compound 6Cc:

Compound 6Cc: ¹H NMR (300MHz, CDCl ₃) δ 7.69 (s, 1H); 7.34 (dd, 1H, J=8.4Hz, J=2.1Hz); 7.25 (m, 2H); 7.08 (m, 2H); 6.97 (m, 2H); 3.44 (s, 3H); 2.30 (s, 3H); 2.29 (s, 3H).

The preparation of embodiment 31. compound 6Cd

Divided by with the corresponding amine of compound 6Cd be outside the raw material, prepare compound 6Cd with the method identical with embodiment 1.

The structure of compound 6Cd:

Compound 6Cd: ¹H NMR (300MHz, CDCl ₃) δ 7.72 (s, 1H); 7.66 (d, 2H, J=8.7Hz); 7.34 (dd, 1H, J=8.4Hz, J=2.1Hz); 7.25 (m, 2H); 7.10 (d, J=8.4Hz, 2H); 3.46 (s, 3H); 2.30 (s, 3H); 2.29 (s, 3H).

The preparation of embodiment 32. compound 6Ce

Divided by with corresponding amine of compound 6Ce and aldehyde be outside the raw material, prepare compound 6Ce with the method identical with embodiment 1.

The structure of compound 6Ce:

Compound 6Ce: ¹H NMR (300MHz, CDCl ₃) δ 7.68 (s, 1H); 7.35 (d, 1H, J=8.4Hz); 7.22 (m, 4H); 6.92 (m, 2H); 3.46 (s, 3H); 2.38 (s, 3H); 2.30 (m, 6H).

The preparation of embodiment 33. compound 6Cf

Divided by with corresponding amine of compound 6Cf and aldehyde be outside the raw material, prepare compound 6Cf with the method identical with embodiment 1.

The structure of compound 6Cf:

Compound 6Cf: ¹H NMR (300MHz, CDCl ₃) δ 7.68 (s, 1H); 7.28 (m, 4H); 6.97 (m, 3H); 3.83 (s, 3H); 3.50 (s, 3H); 2.29 (m, 6H).

The preparation of embodiment 34. compound 6Cg

Divided by with the corresponding amine of compound 6Cg be outside the raw material, prepare compound 6Cg with the method identical with embodiment 1.

The structure of compound 6Cg:

Compound 6Cg: ¹H NMR (300MHz, CDCl ₃) δ 7.68 (s, 1H); 7.35 (dd, 1H, J=8.4Hz, J=2.1Hz); 7.25 (m, 2H); 6.89 (d, 1H, J=8.4Hz); 6.58 (m, 2H); 3.91 (s, 3H); 3.89 (s, 3H); 3.46 (s, 3H); 2.30 (s, 3H); 2.29 (s, 3H).

The preparation of embodiment 35. compound 6Ch

Divided by with corresponding amine of compound 6Ch and aldehyde be outside the raw material, prepare compound 6Ch with the method identical with embodiment 1.

The structure of compound 6Ch:

Compound 6Ch: ¹H NMR (300MHz, CDCl ₃) δ 7.67 (s, 1H); 7.28 (m, 3H); 6.96 (m, 4H); 3.89 (t, 4H, J=4.5Hz); 3.45 (s, 3H); 3.19 (t, 4H, J=4.5Hz); 2.31 (s, 3H); 2.29 (s, 3H).

The preparation of embodiment 36. compound 6Ci

Divided by with corresponding amine of compound 6Ci and aldehyde be outside the raw material, prepare compound 6Ci with the method identical with embodiment 1.

The structure of compound 6Ci:

Compound 6Ci: ¹H NMR (300MHz, CDCl ₃) δ 8.00 (d, 1H, J=7.5Hz); 7.88 (d, 1H, J=6.9Hz); 7.72 (d, 1H, J=8.4Hz); 7.71 (s, 1H); 7.51 (m, 3H); 7.29 (dd, 1H, J=8.4Hz, J=2.1Hz); 7.20 (m, 2H); 7.08 (d, 1H, J=8.4Hz); 3.62 (s, 3H); 2.27 (s, 6H).

The preparation of embodiment 37. compound 6Cj

Divided by with corresponding amine of compound 6Cj and aldehyde be outside the raw material, prepare compound 6Cj with the method identical with embodiment 1.

The structure of compound 6Cj:

Compound 6Cj: ¹H NMR (300MHz, CDCl ₃) δ 7.70 (s, 1H); 7.65 (m, 9H); 7.11 (m, 3H); 3.48 (s, 3H); 2.31 (m, 6H).

The preparation of embodiment 38. compound 6Ck

Divided by with corresponding amine of compound 6Ck and aldehyde be outside the raw material, prepare compound 6Ck with the method identical with embodiment 1.

The structure of compound 6Ck:

Compound 6Ck: ¹H NMR (300MHz, CDCl ₃) δ 8.08 (m, 2H); 7.70 (s, 1H); 7.32 (m, 1H); 7.24 (m, 2H); 7.06 (m, 2H); 3.92 (s, 3H); 3.45 (s, 3H); 2.29 (m, 6H).

The preparation of embodiment 39. compound 6Cl

Divided by with corresponding amine of compound 6Cl and aldehyde be outside the raw material, prepare compound 6Cl with the method identical with embodiment 3.

The structure of compound 6Cl:

Compound 6Cl: ¹H NMR (300MHz, CD ₃OD-d ₄) δ 7.26 (s, 1H); 7.00 (m, 5H); 6.69 (m, 3H); 3.57 (t, 2H, J=7.2Hz); 2.72 (t, 2H, J=7.3Hz).

The preparation of embodiment 40. compound 6Cm

Divided by with the corresponding amine of compound 6Cm, lsothiocyanates and aldehyde be outside the raw material, prepare compound 6Cm with the method identical with embodiment 1.

The structure of compound 6Cm:

Compound 6Cm: ¹H NMR (300MHz, CDCl ₃) δ 7.88 (s, 1H); 7.38 (m, 6H); 7.20 (d, 2H, J=7.2Hz); 5.51 (s, 2H); 2.40 (s, 3H); 2.35 (s, 3H); 2.32 (s, 3H).

The preparation of embodiment 41. compound 6Cn

Divided by with the corresponding amine of compound 6Cn be outside the raw material, prepare compound 6Cn with the method identical with embodiment 3.

The structure of compound 6Cn:

Compound 6Cn: ¹H NMR (300MHz, DMSO-d ₆) δ 7.61 (s, 1H); 7.25 (m, 13H); 6.59 (s, 1H); 2.25 (m, 6H).

The preparation of embodiment 42. compound 6Co

Divided by with corresponding amine of compound 6Co and aldehyde be outside the raw material, prepare compound 6Co with the method identical with embodiment 1.

The structure of compound 6Co:

Compound 6Co: ¹H NMR (300MHz, DMSO-d ₆) δ 9.491 (s, 1H); 7.566 (s, 1H); 7.366 (d, 1H, J=7.2Hz); 7.254 (dd, 1H, J=1.65Hz, J=7.88Hz); 7.058 (m, 4H); 6.924 (dd, 1H, J=1.2Hz, J=7.65Hz); 4.528 (s, 2H); 3.821 (s, 3H); 3.806 (s, 3H); 3.244 (s, 3H).

The preparation of embodiment 43. compound 6Cp

Divided by with corresponding amine of compound 6Cp and aldehyde be outside the raw material, prepare compound 6Cp with the method identical with embodiment 1.

The structure of compound 6Cp:

Compound 6Cp: ¹H NMR (300MHz, CDCl ₃) δ 7.657 (s, 1H); 7.321 (dd, 1H, J=3.7Hz); 7.195 (td, 1H, J=3.42Hz); 7.113 (d, 1H, J=2.1Hz); 6.975 (m, 4H); 3.841 (s, 6H); 3.492 (s, 3H); 2.315 (s, 3H).

The preparation of embodiment 44. compound 6Cq

Divided by with corresponding amine of compound 6Cq and aldehyde be outside the raw material, prepare compound 6Cq with the method identical with embodiment 1.

The structure of compound 6Cq:

Compound 6Cq: ¹H NMR (300MHz, DMSO-d ₆) δ 9.449 (brs, 1H); 7.742 (m, 1H); 7.487 (m, 1H); 7.230 (t, 2H, J=8.7Hz); 7.003 (m, 4H); 3.799 (s, 3H); 3.356 (s, 3H).

The preparation of embodiment 45. compound 6Cr

Divided by with corresponding amine of compound 6Cr and aldehyde be outside the raw material, prepare compound 6Cr with the method identical with embodiment 1.

The structure of compound 6Cr:

Compound 6Cr: ¹H NMR (300MHz, CDCl ₃) δ 7.652 (s, 1H); 7.183 (m, 1H); 6.970 (m, 5H); 6.857 (d, 1H, J=8.1Hz); 5.664 (s, 1H); 3.911 (s, 3H); 3.833 (s, 3H); 3.490 (s, 3H).

The preparation of embodiment 46. compound 6Cs

Divided by with corresponding amine of compound 6Cs and aldehyde be outside the raw material, prepare compound 6Cs with the method identical with embodiment 1.

The structure of compound 6Cs:

Compound 6Cs: ¹H NMR (300MHz, DMSO-d ₆) δ 7.658 (s, 1H), 7.373 (d, 2H, J=8.7Hz), 7.122 (m, 2H), 6.937 (m, 2H), 6.866 (d, 2H, J=9.6Hz), 4.114 (s, 2H), 3.741 (s, 3H), 3.338 (s, 3H).

The preparation of embodiment 47. compound 6Ct

Divided by with corresponding amine of compound 6Ct and aldehyde be outside the raw material, prepare compound 6Ct with the method identical with embodiment 1.

The structure of compound 6Ct:

Compound 6Ct: ¹H NMR (300MHz, DMSO-d ₆) δ 7.719 (s, 1H), 7.530 (d, 2H, J=8.7Hz), 7.155 (m, 3H), 7.084 (d, 1H, J=7.2Hz), 6.936 (m, 2H), 5.200 (s, 2H), 3.742 (s, 3H), 3.333 (s, 3H).

The preparation of embodiment 48. compound 6Cu

Divided by with the corresponding aldehyde of compound 6Cu be outside the raw material, prepare compound 6Cu with the method identical with embodiment 1.

The structure of compound 6Cu:

Compound 6Cu: ¹H NMR (300MHz, DMSO-d ₆) δ 7.77 (s, 1H); 7.47 (d, 2H, J=8.79Hz); 7.45 (m, 2H); 7.40 (d, 1H, J=8.79Hz); 7.06 (d, 2H, J=8.79Hz); 3.33 (s, 3H); 2.29 (s, 6H).

The preparation of embodiment 49. compound 6Cv

Divided by with corresponding amine of compound 6Cv and aldehyde be outside the raw material, prepare compound 6Cv with the method identical with embodiment 1.

The structure of compound 6Cv:

Compound 6Cv: ¹H NMR (300MHz, CDCl ₃) δ 7.692 (s, 1H), 7.396 (d, 2H, J=9Hz), 7.155 (m, 1H), 6.979 (m, 3H), 6.908 (d, 2H, J=9Hz), 4.649 (s, 2H), 3.832 (s, 3H), 3.802 (s, 3H), 3.493 (s, 3H).

The preparation of embodiment 50. compound 6Cw

Divided by with corresponding amine of compound 6Cw and aldehyde be outside the raw material, prepare compound 6Cw with the method identical with embodiment 1.

The structure of compound 6Cw:

Compound 6Cw: ¹H NMR (300MHz, DMSO-d ₆) δ 7.902 (d, 1H, J=7.5Hz), 7.723 (m, 3H), 7.563 (m, 1H), 7.487 (d, 2H, J=8.7Hz), 7.161 (m, 3H), 7.084 (m, 1H), 6.937 (m, 2H), 5.270 (s, 2H), 3.740 (s, 3H), 3.335 (s, 3H).

The preparation of embodiment 51. compound 6Cx

Divided by with corresponding amine of compound 6Cx and aldehyde be outside the raw material, prepare compound 6Cx with the method identical with embodiment 1.

The structure of compound 6Cx:

Compound 6Cx: ¹H NMR (300MHz, CDCl ₃) δ 7.703 (s, 1H), 7.408 (d, 2H, J=8.7Hz), 7.190 (m, 2H), 6.979 (m, 4H), 4.708 (d, 2H, J=2.4Hz), 3.834 (s, 3H), 3.495 (s, 3H), 2.530 (s, 1H).

Embodiment 52. inside and outside pharmacodynamics tests

1. reporter gene expression detects

GLP-1R is a g protein coupled receptor, when GLP-1R with after agonist combines, the proteic G alpha subunit of G is activated, and stimulates adenylate cyclase, causes that the cAMP level raises in the cell.Because of there is the cAMP response element in the promoter region of proinsulin gene, cAMP starts the proinsulin gene transcription with after this response element combines, thus stimulate the expression of Regular Insulin and secretion (Diabetes, 2000,49:1156-1164).This experimental technique adopts stable transfection GLP-1R acceptor gene expression vector and is subjected to the human embryonic kidney cell line (HEK293) of the luciferase reporter gene expression vector of cAMP response element adjusting, detect its reaction (Cell Biology to test compound, 1992,89:8641-8645; Proc Natl Acad Sci USA, 1987,84:3434-3438).When compound is screened, but the sample of the plain enzyme reporter gene expression of induced fluorescence is considered as having the GLP-1R agonist activity.

1.1 test materials and instrument

The HEK293/GLP-1R+Luc cell strain (The National Center for Drug Screening is self-built) of cell strain: GLP-1R and luciferase stably express

Foetal calf serum (GIBCO company)

DMEM substratum (GIBCO company)

Steady-GloTM luciferase analytical system (Promega company)

GLP-1 standard substance (Sigma company)

G418 (Invitrogen company)

Forma CO2gas incubator (Forma company)

Envision plate reading machine (Wallac company)

1.2 test method

The HEK293/GLP1R+Luc cell inserts 96 well culture plates with 20,000/100 μ L/ holes, cultivates based on 37 ℃ of overnight incubation with the DMEM that contains 10% foetal calf serum and 500 μ g/mL G418.Compound 6Cu and GLP-1 standard substance etc. is diluted to the finite concentration gradient, adds earlier 6Cu with 1 μ L/ hole then, 37 ℃ add the GLP-1 standard substance to above-mentioned 96 hole microtest plates after hatching 15min.At 37 ℃, 5%CO ₂Cultivated 6 hours under the condition.Press Steady-Glo ^TMThe explanation of luciferase analytical system test kit detects uciferase activity, Victor ²Plate reading machine carries out reading.

1.3 test-results

Result of study shows (Fig. 1 a-c), but compound 6Cu dose-dependently ground strengthens GLP-1, Exendin-4 and Boc5 inductive, expresses the activating effect of the plain enzyme of HEK293 ICF of humanized GLP-1R, and the excitement effectiveness and the maximum agonist activity that show as the three all are significantly improved.The Dark grey zone is that the signal of 6Cu amplifies effect with top.First area (light gray) is the dosage range of antidiabetic effect; Second area (black) is the dosage range of anti-obesity effect; The 3rd zone (redness) is single with any dosage Boc5, GLP-1 and all inaccessiable effect scope of Exendin-4.

2, receptors bind vigor test

For determine active compound to acceptor in conjunction with vigor, the cell of preparation great expression GLP-1R, with [ ¹²⁵I] GLP-1 of mark _(7-36)Or [ ¹²⁵I] Exendin of mark _(9-39)As part, add compound to be detected simultaneously.When testing compound with ¹²⁵The being at war with property of part of I mark in conjunction with the time, isotope activity on the cytolemma reduces.Can assess in view of the above compound to acceptor avidity (J Mol Endocrinol, 2000,25:321-35; J Biomol Screen, 2000,5:377-84).

2.1 test materials and instrument:

HEK 293/GLP-1R+Luc cell strain (The National Center for Drug Screening is self-built)

Tagged compound: [ ¹²⁵I] GLP-1 of mark _(7-36)And [ ¹²⁵I] Exendin of mark _(9-39)(Amersham Biosciences company)

Wallac MicroBata workstation (PerkinElmer company)

TomTech cell harvestor (TomTec company)

Scintillation solution (Wallac company)

2.2 test method

Get 10 ⁵The HEK 293/GLP-1R+Luc cell of logarithmic phase, under 25 ℃ of conditions, in the 200 μ L assay buffer, with [ ¹²⁵I] GLP-1 of mark _(7-36)Or [ ¹²⁵I] Exendin of mark _(9-39)(final concentration 100pM) hatched 4 hours altogether, added positive peptide of non-marked or compound 6Cu etc. simultaneously.Use cell harvestor, use washing soln washed cell three times.Add scintillation solution, on the MicroBata counter, read every hole reading.

2.3 test-results

The result shows, 6Cu can be specifically with [ ¹²⁵I] GLP-1 of mark _(7-36)Competitive in conjunction with GLP-1R (Fig. 2 left side), its IC ₅₀Value is 3.52 μ M.When maximum concentration was 75 μ M, it was 36.12% to GLP-1's in conjunction with inhibiting rate.6Cu can also be specifically with [ ¹²⁵I] Exendin of mark _(9-39)(GLP-1R peptide agonist) is competitive in conjunction with GLP-1R (Fig. 2 right side), its IC ₅₀Value is 15.96 μ M.When maximum concentration was 75 μ M, it was to Exendin _(9-39)Be 59.7% in conjunction with inhibiting rate.

3.C57BL/6J chmice acute feeding experiment

When taking in nutritive substance, body stimulates small intestine L emiocytosis GLP-1 _7-36amide, GLP-1 is diffused into the capillary vessel stimulation from the intestines wall and imports Sensory neurone into, equally also acts on the pylic afferent neuron of liver, stimulates the neurone in the nucleus tractus solitaril, reflexes to hypothalamus again.Spread out of stimulus signal by efferent neuron and act on pancreas and gi tract, stimulate insulin secretion on the one hand, suppress stomach emptying on the one hand and suppress to ingest.Acute abdominal injection compound 6Cu is adopted in this experiment, observes the variation of mouse food-intake, is intended to observe the effect whether this compounds can strengthen endogenous GLP-1 in vivo, causes the minimizing of food ration.

3.1 test materials and instrument

Mouse: C57BL/6J mouse, 11 ages in week, 22-26g body weight.

3.2 test method

Overnightly divide the cage fasting with mouse, next day, abdominal injection solvent carrier or 0.1,0.3,1 or the 6Cu of 3mg was tried behind the Foodball of mouse known weight to continue observation 2 hours every 15 minutes weighing food weights and record.

3.3 test-results

Compound 6Cu can dose-dependently ground reduce the food ration of mouse, wherein this has significant significant difference to 3mg dosage group mutually with the solvent carrier group, the inhibition of the calculating gained median effective dose (ED that ingests ₅₀) be 0.9mg (Fig. 3)

External activity detected result (the EC of GLP-1 of table 1. part of compounds ₅₀Be 4.6nM)

NA does not record in conjunction with active.

Claims

1, have identical bioactive prodrug by acceptable salt on the substitutive five membered heterocyclic compound of following general formula (formula 1) expression or its pharmacology, ester, solvate, metal complexes or with it:

[formula 1]

Wherein Y is O or S;

R ₁, R ₂Be independently of one another H, ethanoyl, phenyl or

Benzyl or

2, have identical bioactive prodrug according to acceptable salt on the substitutive five membered heterocyclic compound of claim 1 or its pharmacology, ester, solvate, metal complexes or with it, it is characterized in that:

The alkyl of described unsubstituted C1-C10 is methyl, ethyl, propyl group, sec.-propyl, normal-butyl, n-pentyl, cyclopropyl methyl, cyclopentyl-methyl, cyclohexyl methyl; The alkyl substituent of the C1-C10 that replaces comprises the substituted radical of the alkylamino radical of one or more F of being selected from, cyano group, C1-C10 or alkoxyl group, methyl-formiate base, group-4 ethyl formate.

3, has identical bioactive prodrug according to acceptable salt on any described substitutive five membered heterocyclic compound of claim 1 to 2 or its pharmacology, ester, solvate, metal complexes or with it, it is characterized in that, when having geometrical isomer in the molecule, described compound is various independent geometrical isomers or its mixture.

4, according to acceptable salt, ester, solvate, metal complexes on any described substitutive five membered heterocyclic compound or its pharmacology in the claim 1 to 3 or have identical bioactive prodrug with it and be used to prepare the purposes that prevents and/or treats medicines such as metabolic disease, cardiovascular disorder and nerve degenerative diseases.

5, purposes according to claim 4, wherein said metabolic disease are diabetes B, insulin resistant and obesity etc., and described nerve degenerative diseases is an alzheimer's disease etc.

6, purposes according to claim 5 is characterized in that described disease can cause because of the medicine that has gone on the market is produced resistance or produces toxic side effects.

7, a kind of pharmaceutical composition that is used to prevent and/or treat metabolic disease, cardiovascular disorder and nerve degenerative diseases, said composition are the concatenator as acceptable salt and other molecule or carrier on each described compound or its pharmacology in the claim 1 to 3 of activeconstituents.

8, pharmaceutical composition according to claim 7 is characterized in that further containing acceptable carrier and/or vehicle on the pharmacology.

9, a kind of combined preparation comprises as acceptable salt and other treatment medicine on compound as described in the claim 1 to 3 any or its pharmacology.

10, a kind of medicine box, it comprises in the claim 1 to 3 acceptable salt on any described compound or its pharmacology, and uses acceptable salt on described compound or its pharmacology to prevent and/or treat the explanation of metabolic disease, cardiovascular disorder and nerve degenerative diseases etc.

11, a kind of medicine box, it comprises the described combined preparation of claim 9 and uses described combined preparation to prevent and/or treat the explanation of metabolic disease, cardiovascular disorder and nerve degenerative diseases etc.