CN103420981A

CN103420981A - Substituted-pyrrolidinyl-contained thiomorpholine compounds

Info

Publication number: CN103420981A
Application number: CN2012101596323A
Authority: CN
Inventors: 黄海洪; 申竹芳; 吴琪; 刘景龙; 环奕; 林紫云; 李鹏
Original assignee: Institute of Materia Medica of CAMS
Current assignee: Institute of Materia Medica of CAMS
Priority date: 2012-05-21
Filing date: 2012-05-21
Publication date: 2013-12-04

Abstract

The invention discloses substituted-pyrrolidinyl-contained thiomorpholine compounds, a preparation method thereof and applications thereof. In concrete, the invention relates to compounds having a general formula (I), stereoisomers of the compounds and pharmaceutical acceptable salts of the compounds, wherein R is shown in the specification. The invention also relates to pharmaceutical compositions comprising the compounds, and the applications of the compounds in preparing medicines used for treating and/or preventing a disease or a symptom which are related to over-high activity of DPP-IV or over-expression of the DPP-IV, and also relates to a method of using the compounds for treating related diseases. The compounds have activity for effectively inhibiting the DPP-IV.

Description

Substituted pyrrolidinyl-containing thiomorpholine compounds

Technical Field

The invention belongs to the technical field of medicines. Relates to a thiomorpholine compound containing substituted pyrrolidinyl as shown in general formula (I), pharmaceutically acceptable salt and isomer thereof, preparation of the compound, a pharmaceutical composition containing the compound and application of the compound in preventing and/or treating diabetes and non-insulin-dependent diabetes, in particular to application in inhibiting DPP-IV.

Background

Diabetes is a chronic metabolic disease, and the main manifestation of patients is hyperglycemia. Persistent hyperglycemia can lead to a number of complications, such as retinal, renal, nervous, and microvascular complications. Currently, about 2.46 million people worldwide have type 2 diabetes, and the number of patients is expected to reach 3.8 million within 20 years [ xu Si Sheng, research progress of novel antidiabetic drugs, proceedings of Chinese university of pharmacy 2011,42(2):97-106 ]. Diabetes has become a number 3 killer threatening human health following cardiovascular disease and cancer, with type 2 diabetic patients accounting for over 90% of the total number of diabetic patients. The commonly used clinical antidiabetic drugs comprise insulin, biguanides, sulfonylureas, glycosidase inhibitors, thiazolidinediones and the like, but the treatment effect is limited and has adverse reactions of different degrees: such as insulin, can increase body weight and increase the risk of hypoglycemia, while daily administration of injections can lead to poor patient compliance; biguanides, although first-line drugs for type 2 diabetes, have gastrointestinal side effects and may cause lactic acidosis in renal incompetent patients [ Duez H, DPP-4inhibitors in the treatment of type 2 diabetes [ J ]. Biochem Pharm,2012,83:823-832 ].

Glucagon-like peptide 1 (GLP-1) and glucose-dependent insulinotropic polypeptide (GIP) can promote insulin release and inhibit glucagon secretion when blood sugar is increased, and can stimulate insulin gene expression, promote insulin synthesis and beta cell proliferation, and play important roles in the in vivo blood sugar regulation process. Endogenous GLP-1 and GIP can be rapidly degraded and inactivated by dipeptidyl peptidase IV (DPP-IV) in vivo, and the half-life is very short. The DPP-IV inhibitor can increase the concentration of endogenous GIP and GLP-1 in blood, thereby effectively promoting insulin secretion, reducing blood sugar, protecting beta cell function, and not causing the side effects of hypoglycemia and weight gain.

With the intensive research on the DPP family, especially DPP8 and DPP9, since DPP8, DPP9 and DPP-IV have certain similarities in protein structure and substrate type, when DPP-IV inhibitor is less selective, simultaneous action on DPP8/9 can produce serious side effects such as alopecia, thrombocytopenia and anemia [ Havale SH, medical chemistry apoptosis to the inhibition of dipeptidyl peptidase-4 for the treatment of type 2 Diabetes, bioorg Med Chem Lett,2009,17: 1783-. Therefore, the DPP-IV inhibitor has to have high selectivity to a single DPP-IV target so as not to influence the normal physiological functions of other DPPs, which is also a difficult point and a key point for developing novel selective DPP-IV inhibitors.

Therefore, there is still a need in the art for new and potent selective DPP-IV inhibitors to meet the needs of clinical therapy.

Disclosure of Invention

The technical problem to be solved by the invention is to provide a selective DPP-IV inhibitor with a novel structure and strong activity. The inventor finds that the thiomorpholine compound containing the substituted pyrrolidinyl has stronger inhibitory activity and selectivity on DPP-IV, thereby providing a selective DPP-IV inhibitor with novel structure and strong activity, which can be used for preventing and treating diabetes. The present invention has been completed based on the above findings.

Summary of The Invention

To this end, the present invention provides, in a first aspect, compounds of formula (I) and stereoisomers thereof, and pharmaceutically acceptable salts thereof,

wherein,

r is selected from hydrogen and C_1-6Alkyl radical, C_3-8Cycloalkyl, aryl-C_1-3Alkyl, aryl-C_3-8Cycloalkyl, heteroaryl-C_1-3Alkyl, heteroaryl-C_3-8Cycloalkyl radicals containing from 3 to 8 (e.g. 3 to 7, 3 to 6, 3 to 5, or 3 to 4) carbon atoms and from 1 to 3 (e.g. 1 to 2, 1, 2, or 3) atoms selected from nitrogen, oxygenHetero-atomic heterocycloalkyl of sulfur, aryl-C_3-8Heterocycloalkyl, heteroaryl-C_3-8Heterocycloalkyl, benzyl-C_3-8Heterocycloalkyl radical, C_1-6Alkyl formyl radical, C_3-8Cycloalkyl-formyl, aryl-formyl, benzyl-formyl, heteroaryl-formyl, C_1-6Alkylsulfonyl radical, C_3-8Cycloalkylsulfonyl, arylsulfonyl, heteroarylsulfonyl, benzylsulfonyl; wherein said alkyl, cycloalkyl, aryl, heteroaryl, heterocycloalkyl, and benzyl are optionally substituted with 1 to 4 (e.g., 1 to 3, 1 to 2, 1, 2, or 3) groups selected from: H. hydroxy, halogen, cyano, amino, nitro, trifluoromethyl, C_1-6Alkyl radical, C_1-6Group, and NR₁R₂And R is₁And R₂Independently selected from H, C_1-6Alkyl, etc.;

a compound according to any one of the first aspect of the invention which is a compound of formula (IA) and stereoisomers thereof, pharmaceutically acceptable salts thereof,

wherein,

r is selected from hydrogen and C_1-6Alkyl radical, C_3-8Cycloalkyl, aryl-C_1-3Alkyl, aryl-C_3-8Cycloalkyl, heteroaryl-C_1-3Alkyl, heteroaryl-C_3-8Cycloalkyl, heterocycloalkyl containing from 3 to 8 (e.g. 3 to 7, 3 to 6, 3 to 5, or 3 to 4) carbon atoms and from 1 to 3 (e.g. 1 to 2, 1, 2, or 3) heteroatoms selected from nitrogen, oxygen, sulfur, aryl-C_3-8Heterocycloalkyl, heteroaryl-C_3-8Heterocycloalkyl, benzyl-C_3-8Heterocycloalkyl radical, C_1-6Alkyl formyl radical, C_3-8Cycloalkyl-formyl, aryl-formyl, benzyl-formyl, heteroaryl-formyl, C_1-6Alkylsulfonyl radical, C_3-8Cycloalkylsulfonyl, arylsulfonyl, heteroarylsulfonyl, benzylSulfonyl, wherein said alkyl, cycloalkyl, aryl, heteroaryl, heterocycloalkyl, and benzyl are optionally substituted with 1 to 4 (e.g., 1 to 3, 1 to 2, 1, 2, or 3) groups selected from: H. hydroxy, halogen, cyano, amino, nitro, trifluoromethyl, C_1-6Alkyl radical, C_1-6Alkyloxy, and NR₁R₂And R is₁And R₂Independently selected from H, C_1-6Alkyl, etc.;

a compound according to any one of the first aspect of the invention which is a compound of formula (IA) and stereoisomers thereof, pharmaceutically acceptable salts thereof, wherein

R is selected from C_1-6Alkyl, aryl, heteroaryl, C_3-8Nitrogen-containing heterocycloalkyl, aryl-C_1-3Alkyl, heteroaryl-C_1-3Alkyl, arylsulfonyl, heteroaryl-C_3-8Nitrogen-containing heterocycloalkyl, benzyl-C_3-8Nitrogen-containing heterocycloalkyl, etc.; wherein said alkyl, cycloalkyl, aryl, heteroaryl, heterocycloalkyl, and benzyl are optionally substituted with 1 to 4 (e.g., 1 to 3, 1 to 2, 1, 2, or 3) groups selected from: H. hydroxy, halogen, cyano, amino, nitro, trifluoromethyl, C_1-6Alkyl radical, C_1-6Alkyloxy, and NR₁R₂And R is₁And R₂Independently selected from H, C_1-6An alkyl group.

A compound according to any one of the first aspect of the present invention, and stereoisomers thereof, and pharmaceutically acceptable salts thereof, wherein,

r is C_1-6Alkyl, substituted or unsubstituted piperidinyl, substituted or unsubstituted phenyl, substituted or unsubstituted pyridinyl, substituted or unsubstituted quinolinyl, substituted or unsubstituted phenyl-C_1-3Alkyl, substituted or unsubstituted pyridyl-C_1-3Alkyl, substituted or unsubstituted pyridyl-piperidyl, substituted or unsubstituted benzyl-piperidyl, substituted or unsubstituted benzenesulfonyl, and the like; wherein said alkyl, piperidinyl, phenyl, pyridinyl, quinolinyl, and benzyl are optionally substituted with 1-4 (e.g., 1-3, 1-2, 1, 2, or 3)Substituted from the following groups: H. hydroxy, halogen, cyano, amino, nitro, trifluoromethyl, C_1-6Alkyl radical, C_1-6Alkyloxy, and NR₁R₂And R is₁And R₂Independently selected from H, C_1-6An alkyl group.

R is preferably 2, 2-dimethyl-3-hydroxypropyl, piperidinyl, phenyl, 4-methylphenyl, 4-cyanophenyl, 4-trifluoromethylphenyl, 3-nitrophenyl, pyridin-2-yl, 5-cyano-pyridin-2-yl, 3-cyano-pyridin-2-yl, quinolin-2-yl, 3, 4-dimethoxybenzyl, 4-cyanobenzyl, pyridin-2-yl-methyl, (5-cyanopyridin-2-yl) -piperidin-4-yl, 4-cyanobenzylpiperidin-4-yl, p-nitrobenzenesulfonyl, p-methylbenzenesulfonyl or the like.

The pharmaceutically acceptable salts described in the present invention are salts of the compounds of the present invention with an acid selected from the group consisting of: hydrochloric acid, p-toluenesulfonic acid, tartaric acid, maleic acid, lactic acid, methanesulfonic acid, sulfuric acid, phosphoric acid, citric acid, acetic acid or trifluoroacetic acid. Preferably hydrochloric acid, p-toluenesulfonic acid or trifluoroacetic acid.

A compound according to any one of the first aspect of the present invention, which is the subject compound of the present invention prepared in the examples (represented by structural formula or described by systematic name) and stereoisomers thereof, and pharmaceutically acceptable salts thereof.

A compound according to any one of the first aspect of the invention, which is a compound selected from:

in a second aspect, the present invention provides a process for the preparation of a compound according to any one of the first aspect of the invention, comprising the steps of:

reacting a compound of formula 1a with (Boc) in a solvent (e.g., water, dioxane, or a mixture thereof) in the presence of a base (e.g., an inorganic base such as sodium hydroxide or an organic base such as triethylamine) at a temperature of-10 ℃ to 40 ℃ (e.g., -10 ℃ to 35 ℃, -5 ℃ to 30 ℃, -5 ℃ to 25 ℃, or 0 ℃ to 25 ℃)₂And O reacts for 2 to 12 hours to obtain the compound shown as the formula 2 a.

Reacting the compound of formula 2a with thiomorpholine in an organic solvent (e.g., N-dimethylformamide) at a temperature of 10 ℃ to 40 ℃ (e.g., 10 ℃ to 35 ℃, 15 ℃ to 30 ℃,20 ℃ to 30 ℃, or 20 ℃ to 25 ℃) in the presence of 1-2 equivalents of a condensing agent (e.g., EDCI, HOBT, or a mixture of both) and a base (e.g., triethylamine) for 4-8 hours to provide the compound of formula 3 a.

The compound represented by formula 3a is reacted with sulfonyl chloride (e.g., methanesulfonyl chloride, benzenesulfonyl chloride, p-toluenesulfonyl chloride, or the like) in a solvent (e.g., dichloromethane) at a temperature of-10 ℃ to 10 ℃ (e.g., -5 ℃ to 10 ℃,0 ℃ to 10 ℃, or 5 ℃ to 10 ℃) in the presence of a base (e.g., triethylamine) for 1-5 hours to give the compound represented by formula 4 a.

Reacting the compound represented by formula 4a with sodium azide at a temperature of 60 ℃ to 90 ℃ (e.g., 60 ℃ to 70 ℃, 70 ℃ to 80 ℃, or 80 ℃ to 90 ℃) in a solvent (e.g., N-dimethylformamide) for 4-6 hours to obtain the compound represented by formula 5 a.

The compound of formula 5a is reduced by hydrogenation over a catalyst (e.g., palladium on charcoal) in a solvent (e.g., methanol or ethanol) at room temperature (e.g., 15 ℃ to 25 ℃) for 5 to 8 hours to provide the compound of formula 6 a.

Subjecting the compound of formula 6a to a substitution reaction with a halide RX (X is halogen) in a base (e.g. potassium carbonate) and a suitable solvent (e.g. N, N-dimethylformamide) at a temperature of 60 ℃ to 90 ℃ (e.g. 60 ℃ to 70 ℃, 70 ℃ to 80 ℃ or 80 ℃ to 90 ℃) for about 4 to 12 hours to give a compound of formula b; or,

reacting a compound of formula 6a with an aldehyde or ketone containing an R group under acidic conditions (e.g., acetic acid) at a temperature of-10 ℃ to 40 ℃ (e.g., -10 ℃ to 35 ℃, -5 ℃ to 30 ℃, -5 ℃ to 25 ℃, or 0 ℃ to 25 ℃) for about 2-6 hours to provide a compound of formula b; or,

reacting the compound of formula 6a with a sulfonyl chloride containing an R group in a base (e.g., triethylamine) and a suitable solvent (e.g., dichloromethane) at a temperature of-20 ℃ to 0 ℃ (e.g., -20 ℃ to 0 ℃, -15 ℃ to 0 ℃, or-5 ℃ to 0 ℃) for about 2-6 hours to provide a compound of formula b; or,

the compound of formula 6a is reacted with the halogenated aromatic compound containing the R group using a suitable ligand such as Davep and a catalyst such as palladium acetate in a base such as sodium t-butoxide and a suitable solvent such as toluene at a temperature of 90 c to 150 c (e.g., 100 c to 150 c, 110 c to 150 c or 130 c to 150 c) for about 8 to 16 hours to provide the compound of formula b.

The Boc protecting group is removed from the compound of formula b in ethyl acetate hydrochloride solution at a temperature of 10 ℃ to 40 ℃ (e.g., 10 ℃ to 35 ℃, 15 ℃ to 30 ℃,20 ℃ to 30 ℃ or 20 ℃ to 25 ℃) to provide the compound of the invention of formula IA. If racemic 1a is used as the raw material, the compound of the invention shown in the formula I is obtained according to the steps and the method.

Wherein R is as defined in the first aspect of the invention.

In a third aspect, the present invention provides a pharmaceutical composition comprising a therapeutically and/or prophylactically effective amount of a compound according to any one of the first aspect of the present invention and stereoisomers thereof, pharmaceutically acceptable salts thereof, and optionally one or more pharmaceutically acceptable carriers or excipients.

The fourth aspect of the present invention provides a compound according to any one of the first aspect of the present invention and stereoisomers, pharmaceutically acceptable salts thereof, or a pharmaceutical composition according to any one of the third aspect of the present invention, for use in the manufacture of a medicament for the treatment and/or prevention of diseases or conditions associated with DPP-IV hyperactivity or DPP-IV overexpression. In one embodiment, the disease or condition associated with DPP-IV hyperactivity or DPP-IV overexpression is a disease or condition selected from: diabetes, hyperglycemia, non-insulin dependent diabetes, type 2 diabetes, and the like.

The fourth aspect of the present invention also provides a compound according to any one of the first aspect of the present invention and stereoisomers, pharmaceutically acceptable salts thereof, or a pharmaceutical composition according to any one of the third aspect of the present invention, for use in the manufacture of a medicament for the treatment and/or prophylaxis of diabetes, hyperglycemia, non-insulin dependent diabetes, type 2 diabetes and the like.

The fourth aspect of the present invention also provides a compound according to any one of the first aspect of the present invention and stereoisomers, pharmaceutically acceptable salts thereof, or a pharmaceutical composition according to any one of the third aspect of the present invention for use in the manufacture of a medicament for use as a DPP-IV inhibitor.

In a fifth aspect, the present invention provides a method for treating and/or preventing diseases or disorders associated with DPP-IV hyperactivity or DPP-IV overexpression in a subject in need thereof, the method comprising administering to the subject in need thereof a therapeutically and/or prophylactically effective amount of a compound according to any one of the first aspect of the present invention, a stereoisomer thereof, a pharmaceutically acceptable salt thereof, or a pharmaceutical composition according to any one of the third aspect of the present invention. The method according to the fifth aspect of the present invention, wherein the disease or disorder associated with DPP-IV hyperactivity or DPP-IV overexpression is selected from diabetes, hyperglycemia, non-insulin dependent diabetes mellitus, type 2 diabetes and the like.

The fifth aspect of the present invention also provides a method for the treatment and/or prophylaxis of diabetes, hyperglycemia, non-insulin dependent diabetes mellitus, type 2 diabetes in a subject in need thereof, which comprises administering to the subject in need thereof a therapeutically and/or prophylactically effective amount of a compound according to any one of the first aspect of the present invention and stereoisomers thereof, pharmaceutically acceptable salts thereof, or a pharmaceutical composition according to any one of the third aspect of the present invention.

In a sixth aspect, the present invention provides a compound according to any one of the first aspect of the present invention and stereoisomers thereof, and pharmaceutically acceptable salts thereof, for use in the treatment and/or prevention of diseases or conditions associated with DPP-IV hyperactivity or DPP-IV overexpression. A compound according to a sixth aspect of the present invention, wherein said disease or condition associated with DPP-IV hyperactivity or DPP-IV overexpression is selected from: diabetes, hyperglycemia, non-insulin dependent diabetes, type 2 diabetes, and the like.

The sixth aspect of the present invention also provides a compound according to any one of the first aspect of the present invention and stereoisomers thereof, and pharmaceutically acceptable salts thereof, for use in the treatment and/or prevention of diabetes, hyperglycemia, non-insulin dependent diabetes, type 2 diabetes and the like.

Any aspect of the invention or any one of the aspects having features is equally applicable to any other aspect or any one of the other aspects as long as they are not mutually inconsistent, although appropriate modifications to the respective features may be made as necessary when applicable to each other. In the present invention, for example, reference to "any of the first aspects of the invention" means any sub-aspect of the first aspects of the invention, and in other respects similarly referred to, has similar meaning.

Detailed description of the invention:

various aspects and features of the disclosure are described further below.

All documents cited herein are incorporated by reference in their entirety and to the extent such documents do not conform to the meaning of the present invention, the present invention shall control. Further, the various terms and phrases used herein have the ordinary meaning as is known to those skilled in the art, and even though such terms and phrases are intended to be described or explained in greater detail herein, reference is made to the term and phrase as being inconsistent with the known meaning and meaning as is accorded to such meaning throughout this disclosure. The following are definitions of various terms used herein, which apply to the terms used throughout the specification of the present application unless otherwise specified in specific instances.

The term "alkoxy" refers to alkyl-O-, wherein alkyl is as described herein.

As used herein, the terms "halo", "halogen atom", "halo", and the like, denote fluorine, chlorine, bromine, or iodine, and particularly denote fluorine, chlorine, or bromine.

As used herein, the term "alkyl" refers to an alkyl group having the specified number of carbon atoms, which is a straight or branched chain alkyl group, and which may include sub-groups thereof, such as the instant "C" group_1-6When "alkyl", it may also include C_1-5Alkyl radical, C_1-4Alkyl radical, C_2-6Alkyl radical, C_2-4Alkyl, etc., and specific groups such as methyl, ethyl, n-propyl, isopropyl, n-butyl, sec-butyl, tert-butyl, pentyl, hexyl.

The term "aryl" as described herein, alone or in combination, is defined herein as a monocyclic or bicyclic aromatic group. Examples of aryl groups include, but are not limited to, phenyl, naphthyl, and the like. Similarly, the term "aryloxy-" refers to an aryl group that is attached to the rest of the compound through an oxygen.

As used herein, the term "heteroaryl" refers herein to aryl groups having from 1 to 3 heteroatoms as ring atoms, the remaining ring atoms being carbon, the heteroatoms including oxygen, sulfur and nitrogen. Examples of heteroaryl groups include, but are not limited to, pyridyl, pyrimidinyl, imidazolyl, furyl, thienyl, pyrazinyl, quinolinyl, isoquinolinyl, tetrahydroisoquinolinyl, and the like. In one embodiment, the heteroaryl is pyridyl, quinolyl.

The term "aryl-C" as described herein_1-3Alkyl- "means an aryl group, which is bonded through C_1-3Alkyl groups are attached to the rest of the compound.

The term "heteroaryl-C" as described herein_1-3Alkyl- "means a heteroaryl group, which is bonded through C_1-3Alkylation withThe other part of the compound is attached.

As used herein, the term "cycloalkyl" refers to a cyclic alkyl group having the number of ring carbon atoms specified, and which may include a sub-group thereof, such as, for example, the time "C_3-8When cycloalkyl "is present, it may also include C_3-7Cycloalkyl radical, C_3-6Cycloalkyl radical, C_3-5Cycloalkyl radical, C_4-7Cycloalkyl, and the like, as well as specific groups such as cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, and the like.

As used herein, the term "heterocycloalkyl" refers to a cyclic heteroalkyl group having the indicated number of ring atoms, including monocyclic or fused ring groups, having from 5 to 9 ring atoms in the ring, wherein one or two ring atoms are heteroatoms selected from nitrogen, oxygen, or sulfur, and the remaining ring atoms are carbon. The rings may also have one or more double bonds, however, the rings do not have a completely conjugated n-electron system. Heterocycloalkyl groups include, but are not limited to, pyrrolidinyl, piperidinyl, piperazinyl, morpholinyl, thiomorpholinyl, and the like.

As used herein, the term "effective amount" refers to a dose that achieves treatment and/or prevention of a disease or disorder described herein in a subject.

As used herein, the term "pharmaceutical composition" may also refer to a "composition" that may be used to effect treatment and/or prevention of a disease or disorder described herein in a subject, particularly a mammal.

As used herein, the term "subject" can refer to a patient or other animal, particularly a mammal, e.g., a human, dog, monkey, cow, horse, etc., that receives a compound of formula I of the invention or a pharmaceutical composition thereof for the treatment and/or prevention of a disease or disorder described herein.

As used herein, the term "disease and/or disorder" refers to a physical condition of the subject that is associated with the disease and/or disorder of the present invention. For example, the disease and/or condition of the present invention may refer to a physical condition, such as a physical condition exhibiting elevated blood glucose levels, or a disease condition, such as a disease condition characterized by hyperglycemia, diabetes, and the like. The body state and the disease state are not distinguished herein or may be referred to one another, e.g., "hyperglycemia" may be used interchangeably with "hyperglycemia".

As used herein, "%" refers to weight/weight percentages, particularly where solid matter is described, as well as where not specifically indicated. Of course, in describing liquid materials, the "%" may refer to weight/volume percentages (for the case of solids dissolved in liquids) or may refer to volume/volume percentages (for the case of liquids dissolved in liquids).

As used herein, the term "pharmaceutically acceptable" when describing a "pharmaceutically acceptable salt" means not only that the salt is physiologically acceptable to the subject, but may also refer to a synthetic substance of pharmaceutical value, e.g., a salt formed as an intermediate in order to effect chiral resolution, which salt may play a role in achieving the end product of the invention, although such intermediate salt may not be directly administered to the subject.

In a further aspect, the invention relates to pharmaceutical compositions comprising as active ingredient a compound of the invention. The pharmaceutical composition may be prepared according to methods well known in the art. The compounds of the invention may be formulated into any dosage form suitable for human or animal use by combining them with one or more pharmaceutically acceptable solid or liquid excipients and/or adjuvants. The compounds of the present invention are generally present in the pharmaceutical compositions in an amount of from 0.1 to 95% by weight.

The compounds of the present invention or pharmaceutical compositions containing them may be administered in unit dosage form by enteral or parenteral routes, such as oral, intravenous, intramuscular, subcutaneous, nasal, oromucosal, ophthalmic, pulmonary and respiratory, dermal, vaginal, rectal, and the like.

The dosage form for administration may be a liquid dosage form, a solid dosage form, or a semi-solid dosage form. The liquid dosage forms can be solution (including true solution and colloidal solution), emulsion (including o/w type, w/o type and multiple emulsion), suspension, injection (including water injection, powder injection and infusion), eye drop, nose drop, lotion, liniment, etc.; the solid dosage form can be tablet (including common tablet, enteric coated tablet, buccal tablet, dispersible tablet, chewable tablet, effervescent tablet, orally disintegrating tablet), capsule (including hard capsule, soft capsule, and enteric coated capsule), granule, powder, pellet, dripping pill, suppository, pellicle, patch, aerosol (powder), spray, etc.; semisolid dosage forms can be ointments, gels, pastes, and the like.

The compound can be prepared into common preparations, sustained release preparations, controlled release preparations, targeting preparations and various particle drug delivery systems.

For tableting the compound of the present invention, a wide variety of excipients known in the art may be used, including diluents, binders, wetting agents, disintegrants, lubricants, and solubilizers. The diluent can be starch, dextrin, sucrose, glucose, lactose, mannitol, sorbitol, xylitol, microcrystalline cellulose, calcium sulfate, calcium hydrogen phosphate, calcium carbonate, etc.; the humectant can be water, ethanol, isopropanol, etc.; the binder can be starch slurry, dextrin, syrup, Mel, glucose solution, microcrystalline cellulose, acacia slurry, gelatin slurry, sodium carboxymethylcellulose, methylcellulose, hydroxypropyl methylcellulose, ethyl cellulose, acrylic resin, carbomer, polyvinylpyrrolidone, polyethylene glycol, etc.; the disintegrant may be dry starch, microcrystalline cellulose, low-substituted hydroxypropyl cellulose, crosslinked polyvinylpyrrolidone, crosslinked sodium carboxymethylcellulose, sodium carboxymethyl starch, sodium bicarbonate and citric acid, polyoxyethylene sorbitol fatty acid ester, sodium dodecyl sulfate, etc.; the lubricant and cosolvent may be talc, silica, stearate, tartaric acid, liquid paraffin, polyethylene glycol, etc.

The tablets may be further formulated into coated tablets, such as sugar-coated tablets, film-coated tablets, enteric-coated tablets, or double-layer and multi-layer tablets.

To encapsulate the administration unit, the active ingredient of the compound of the present invention may be mixed with a diluent and a cosolvent, and the mixture may be directly placed in a hard capsule or soft capsule. Or the effective component of the compound of the invention can be prepared into granules or pellets with diluent, adhesive and disintegrating agent, and then placed into hard capsules or soft capsules. The diluents, binders, wetting agents, disintegrants, and cosolvents used to prepare the compound tablets of the present invention can also be used to prepare capsules of the compounds of the present invention.

For preparing the compound of the present invention into injection, water, ethanol, isopropanol, propylene glycol or their mixture can be used as solvent, and appropriate amount of solubilizer, cosolvent, pH regulator, and osmotic pressure regulator commonly used in the art can be added. The solubilizer or cosolvent can be poloxamer, lecithin, hydroxypropyl-beta-cyclodextrin, etc.; the pH regulator can be phosphate, acetate, hydrochloric acid, sodium hydroxide, etc.; the osmotic pressure regulator can be sodium chloride, mannitol, glucose, phosphate, acetate, etc. For example, mannitol and glucose can be added as proppant for preparing lyophilized powder for injection.

In addition, colorants, preservatives, flavors, or other additives may also be added to the pharmaceutical preparation, if desired.

For the purpose of administration and enhancing the therapeutic effect, the drug or pharmaceutical composition of the present invention can be administered by any known administration method.

The dosage of the pharmaceutical composition of the compound of the present invention to be administered may vary widely depending on the nature and severity of the disease to be prevented or treated, the individual condition of the patient or animal, the route and dosage form of administration, and the like. Generally, a suitable daily dosage range for a compound of the invention is from 0.001 to 150mg/Kg body weight, preferably from 0.1 to 100mg/Kg body weight, more preferably from 1 to 60mg/Kg body weight, and most preferably from 2 to 30mg/Kg body weight. The above-described dosage may be administered in one dosage unit or divided into several dosage units, depending on the clinical experience of the physician and the dosage regimen including the use of other therapeutic means.

The compounds or compositions of the present invention may be administered alone or in combination with other therapeutic or symptomatic agents. When the compound of the present invention is used in a synergistic manner with other therapeutic agents, the dosage thereof should be adjusted according to the actual circumstances.

Advantageous technical effects

The inventor finds that the in vitro DPP-IV inhibitory activity of most of thiomorpholine compounds containing substituted pyrrolidine reaches more than 50%, wherein the IC of 9 compounds₅₀The DPP-IV inhibitor reaches micromolar level, has low inhibition rate to DPP8 and DPP9, shows good DPP-IV selectivity, and thus, provides a selective DPP-IV inhibitor with novel structure and strong activity, and can be used for preventing and treating diabetes.

Detailed Description

The present invention will be further described by the following examples, however, the scope of the present invention is not limited to the following examples. It will be understood by those skilled in the art that various changes and modifications may be made to the invention without departing from the spirit and scope of the invention. The present invention has been described generally and/or specifically with respect to materials used in testing and testing methods. Although many materials and methods of operation are known in the art for the purpose of carrying out the invention, the invention is nevertheless described herein in as detail as possible.

For all of the following examples, standard procedures and purification methods known to those skilled in the art may be used. Unless otherwise indicated, all temperatures are expressed in degrees centigrade (degrees celsius). The structure of the compounds is determined by nuclear magnetic resonance spectroscopy (NMR) and/or Mass Spectrometry (MS). m.p. is the melting point given at ℃, temperature uncorrected.

Preparation examples section

The structure of the compound is as followsNuclear magnetic resonance hydrogen spectrum (¹H NMR) and Mass Spectrometry (MS). The hydrogen and carbon spectral shifts (δ) for nuclear magnetic resonance are given in parts per million (ppm). NMR spectra were obtained using a Mercury-300 or Mercury-400 NMR spectrometer, deuterated chloroform (CDCl)₃) Or heavy water (D)₂O) or deuterated dimethyl sulfoxide (DMSO-d)₆) As solvent Tetramethylsilane (TMS) was used as internal standard.

The melting point was measured using a Yanaco model M.P-500D melting point tester, Japan, and the temperature was not corrected.

The high resolution mass spectrum was measured using an Agilent 1100series LC/MSD trap mass spectrometer.

The specific optical rotation was measured using a Perkin-Elmer model 241MC polarimeter under sodium light at 20 ℃.

The electronic balance used was an electronic balance model Yanaco LY-300, Japan.

The column chromatography generally uses 200-300 mesh silica gel as a carrier.

The anhydrous solvents were all processed by standard methods. Other reagents were all commercially available analytical grade.

Wherein,

(Boc)₂o is Di-tert-butyl dicarbanate, namely Di-tert-butyl dicarbonate.

AcOH is an Acetic Acid, i.e., Acetic Acid.

DMF is N, N-Dimethylformamide, i.e., N-Dimethylformamide.

EDCI is 1- (3- (dimethylamino) propyl) -3-ethylcarbodiimide hydrochloride, i.e., 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride.

HOBT is 3-hydroxy-benzotriazole hydrate, namely 1-hydroxybenzotriazole.

Davep is 2-dicyclohexylphosphino-2 '- (N, N-dimethyllamino) biphenyl, i.e., 2-dicyclohexylphosphino-2' - (N, N-dimethylamino) biphenyl.

Preparation example

Synthetic route of Scheme 1 intermediate 6a

First step preparation of (2S, 4R) -N-tert-Butoxycarbonyl-4-hydroxypyrrolidine-2-carboxylic acid 2a

4-OH-L-proline (3.95g,30.1mmol) was dissolved in 50mL of 1, 4-dioxane and 50mL of 1 mol. L^-1Adding (Boc) to a mixed solution of sodium hydroxide solution at 0 deg.C₂O (7.85g, 36.0mmol), stirred at low temperature for 0.5h, then warmed to room temperature and stirred overnight. Concentrating under reduced pressure, steaming to near dryness, dissolving in 50mL of water, and dissolving with 1 mol/L^-1Adjusting the pH value of the HCl solution to 2-3, extracting with ethyl acetate, combining organic layers, washing the organic layers with saturated saline solution, drying with anhydrous sodium sulfate, filtering, and concentrating under reduced pressure to obtain an intermediate 2a which is 6.68g of white solid with the yield of about 96.3%.¹H NMR(D₂O,300MHz)δ:4.53(br,1H,CH-OH in pyrrolidine),4.42-4.36(m,1H,CHC=O),3.60-3.47(m,2H,CH₂N in pyrrolidine),2.42-2.35&2.19-2.10(m,2H,CH₂CHC=O in pyrrolidine),1.43(s,9H,Boc).

Second step preparation of 4- { [ (2S, 4R) -N-tert-Butoxycarbonyl-4-hydroxy-pyrrol-2-yl ] -formyl } thiomorpholine 3a

(2S, 4R) -N-tert-Butoxycarbonyl-4-hydroxypyrrolidine-2-carboxylic acid 2a (5.77g,25.0mmol) was dissolved in 50mL of anhydrous DMF and EDCI (5.82g,30.3mmol) and HOBT (4.06g,30.1mmol) were added and after complete dissolution thiomorpholine (2.83g,27.5mmol) and triethylamine (5.06g,50.1mmol) were added and stirred at room temperature overnight to completion. 50mL of ice water was added to the reaction system and stirred for 30 min. Extracting with ethyl acetate, combining organic layers, washing the organic layers with water and saturated sodium chloride water respectively, drying with anhydrous sodium sulfate, filtering, concentrating under reduced pressure, separating the obtained crude product by silica gel (200-300 meshes) column chromatography, and taking dichloromethane-methanol (V: V = 50: 1) mixed solution as an eluent to obtain an intermediate 3a, 5.94g of white solid, and the yield is 75.2%.

¹H NMR(CDCl₃,300MHz)δ:4.83-4.69(m,1H,CHC=O),4.55-4.50(br,1H,CH-NH₂ in pyrrolidine),3.97-3.79(m,4H,CH₂NCH₂ in thiomorpholine),3.70-3.48(m,2H,CH₂N in pyrrolidine),2.68-2.58(m,4H,CH₂SCH₂ in thiomorpholine),2.28-2.03(m 2H CH ₂CHC=O in pyrrolidine)1.45(s 9H Boc)HR-MS(ESI-TOF⁺):C₁₄H₂₅N₂O₄S calculated value 317.1530, found [ M + H]⁺317.1502.

Third step preparation of 4- { [ (2S, 4R) -N-tert-Butoxycarbonyl-4-methanesulfonyl-pyrrol-2-yl ] -formyl } thiomorpholine 4a

4- { [ (2S, 4R) -N-tert-Butoxycarbonyl-4-hydroxy-pyrrol-2-yl ] -formyl } thiomorpholine 3a (5.37g,17.0mmol) was dissolved in 50mL of anhydrous dichloromethane, triethylamine (2.58g,25.5mmol) was added, the reaction system was cooled to below 0 deg.C, methanesulfonyl chloride (1.9mL,24.5mmol) was slowly added dropwise, and the reaction was allowed to proceed at low temperature for 2 hours until completion (TLC trace). The reaction solution was warmed to room temperature, and the organic phase was washed successively with distilled water and saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was chromatographed on silica gel (200-300 mesh) using dichloromethane-methanol (V: V = 50: 1) as eluent to give intermediate 4a as a white solid (6.53 g, 97.5% yield).

¹H NMR(CDCl₃,300MHz)δ:5.32-5.24(br,1H,CH-OSO₂CH₃ in pyrrolidine),4.88-4.71(m,1H,CHC=O in pyrrolidine),4.00-3.97(m,2H,CH₂N in pyrrolidine),3.90-3.77(m,4H,CH₂NCH₂ in thiomorpholine),3.05(s,3H,CH₃),2.89-2.21(m,6H,CH₂SCH₂ in thiomorpholine,CH ₂CHC=O in pyrrolidine),1.25(s,9H,Boc).HR-MS(ESI-TOF⁺):C₁₅H₂₇N₂O₆S₂Calculated value 395.1305, found [ M + H]⁺395.1272.

Step four preparation of 4- { [ (2S,4S) -4-azido-N-tert-butoxycarbonyl-pyrrol-2-yl ] -formyl } thiomorpholine 5a

4- { [ (2S, 4R) -N-tert-butoxycarbonyl-4-methanesulfonyl-pyrrol-2-yl ] -formyl } thiomorpholine 4a (6.30g,16.0mmol) was dissolved in 30mL of anhydrous DMF, sodium azide (1.56g,24.0mmol) was added, reaction was carried out at 80 ℃ for 5 hours until completion, 30mL of ice water was added to the reaction system, stirring was carried out for 30 minutes, extraction was carried out with ethyl acetate, the organic layers were combined, washed with water and saturated brine respectively, dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and the resulting crude product was subjected to column chromatography on silica gel (200-300 mesh) using a petroleum ether-ethyl acetate (V: V = 1: 1) mixture as an eluent, to give intermediate 5a, 4.53g of a white solid, and yield of 82.9%.

¹H NMR(CDCl₃,300MHz)δ:4.69-4.51(m,1H,CHC=O),4.12-4.03(m,1H,CH-N₃ in pyrrolidine),3.95-3.85(m,4H,CH₂NCH₂ in thiomorpholine),3.79-3.27(m,2H,CH₂N in pyrrolidine),2.68-2.48(m,4H,CH₂SCH₂ in thiomorpholine),2.91-1.88(m,2H,CH ₂CHC=O in pyrrolidine),1.51(s,9H,Boc).HR-MS(ESI-TOF⁺):C₁₄H₂₄N₅O₃S calculated value 342.1594, found [ M + H]⁺342.1654.

Fifth step preparation of 4- { [ (2S,4S) -4-amino-N-tert-butoxycarbonyl-pyrrol-2-yl ] -formyl } thiomorpholine 6a

4- { [ (2S,4S) -4-azido-N-t-butoxycarbonyl-pyrrol-2-yl ] -formyl } thiomorpholine 5a (4.42g,13.0mmol) was dissolved in 20mL of methanol, 10% by mass of Pd/C (0.66g, 15% by mass) was added, and the reaction was hydrogenated at medium pressure for 4 hours until completion. The solid in the reaction solution was filtered off, and the crude product was separated by column chromatography on silica gel (200-mesh 300-mesh) using dichloromethane-methanol (V: V = 50: 3, 1mL ammonia per 200 mL) as eluent to give intermediate 6a as a white solid 3.05g with a yield of 74.5%.

¹H NMR(CDCl₃,300MHz)δ:4.68-4.51(m,1H,CHC=O),4.10-3.93(m,1H,CH-NH₂ in pyrrolidine),3.85-3.71(m,4H,CH₂NCH₂ in thiomorpholine),3.56-3.34(m,2H,CH₂N in pyrrolidine),2.83-2.60(m,4H,CH₂SCH₂ in thiomorpholine),2.44-1.63(m,2H,CH ₂CHC=O in pyrrolidine),1.45(s,9H,Boc).HR-MS(ESI-TOF⁺):C₁₄H₂₆N₃O₃S calculated value 316.1689, found [ M + H]⁺316.1680.

Examples

Example 1

The compound 14- { [ (2S,4S) -4- (piperidin-4-yl-amino) -pyrrol-2-yl ] -formyl } thiomorpholine trihydrochloride

First step preparation of 4- { [ (2S, S) -N-tert-Butoxycarbonyl-4- (piperidin-4-yl) amino-pyrrol-2-yl ] -formyl } thiomorpholine 1b

4- { [ (2S,4S) -4-amino-N-tert-butoxycarbonyl-pyrrol-2-yl ] -formyl } thiomorpholine 6a (0.15g,0.48mmol) and N-Boc-4-piperidone (0.098g,0.49mmol) were dissolved in 10mL of dichloromethane, and then a small amount of AcOH was added to adjust the pH of the reaction system to 3-5, and the mixture was stirred at room temperature for 3 hours, the reaction system was cooled to below 0 deg.C, sodium triacetoxyborohydride (0.317g,1.50mmol) was added, and the mixture was warmed to room temperature and stirred for 6 hours until the reaction was complete. To the reaction solution was added 20mL of water, followed by extraction with dichloromethane, and the organic layers were combined, washed with a saturated sodium bicarbonate solution and a saturated brine, respectively, and the organic layer was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was chromatographed on a silica gel (200-300 mesh) column using dichloromethane-methanol (V: V = 50: 1) as eluent. Intermediate 1b was obtained as a pale yellow oil 167mg, yield 69.7%.

Second step preparation of 4- { [ (2S,4S) -4- (piperidin-4-yl-amino) -pyrrol-2-yl ] -formyl } thiomorpholine trihydrochloride (Compound 1)

Coupling 4- { [ (2S,4S) -N-tert-butoxycarbonyl-4- (piperidin-4-yl) amino-pyrrol-2-yl]-formyl } thiomorpholine 1b (167mg,0.33mmol) and 2mL of ethyl acetate hydrochloride solution (7 mol. L)_-1) Mixing, stirring at room temperature for 1 hour, precipitating white solid from clear reaction liquid, and washing the obtained white solid with n-hexane solvent. Compound 1 was obtained as an off-white solid, 108mg, yield 79.2, m.p.>250℃，

¹H NMR(DMSO-d₆,300MHz)δ:4.65(m,1H,CHC=O in pyrrolidine),3.96(br,1H,CHNH in pyrrolidine),3.78-3.71(m,4H,CH₂NCH₂ in thiomorpholine),3.59(m,1H,CH in piperidine),2.86(m,4H CH ₂NHCH ₂ in piperidine),2.71(m,2H,CH ₂CHC=O in pyrrolidine),2.62(m,4H,CH₂SCH₂ in thiomorpholine),2.19-1.88(m,6H,CH ₂CHCH ₂ in piperidine&NHCHCH ₂NH in pyrrolidine).HR-MS(ESI-TOF⁺):C₁₄H₂₆N₄OS calculated 299.1900, found [ M-3HCl + H]⁺299.1894。

Example 2

The compound 24- { [ (2S,4S) -4- (3-hydroxy-2, 2-dimethyl-propylamino) -pyrrol-2-yl ] -formyl } thiomorpholine dihydrochloride

First step preparation of 4- { [ (2S,4S) -N-tert-Butoxycarbonyl-4- (3-hydroxy-2, 2-dimethylpropano) -pyrrol-2-yl ] -formyl } thiomorpholine 2b

4- { [ (2S,4S) -4-amino-N-tert-butoxycarbonyl-pyrrol-2-yl ] -formyl } thiomorpholine 6a (0.15g,0.48mmol) and 2, 2-dimethyl-3-hydroxypropanal (0.051g,0.50mmol) were dissolved in 10mL of dichloromethane, and after complete dissolution, a small amount of acetic acid was added to the reaction pH =3-5, and stirred at room temperature for 3 hours, the reaction was cooled to below 0 deg.C, sodium triacetoxyborohydride (0.317g,1.50mmol) was added, and the reaction was warmed to room temperature and stirred for 6 hours until completion. To the reaction solution was added 20mL of water, and the combined organic layers were extracted with dichloromethane, washed with a saturated sodium bicarbonate solution and a saturated brine, respectively, and the organic layer was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. Separating by column chromatography, and eluting with dichloromethane-methanol (V: V = 100: 3) mixture. Intermediate 2b was obtained as a pale yellow oil 124mg, yield 64.4%.

Second step preparation of 4- { [ (2S,4S) -4- (3-hydroxy-2, 2-dimethyl-propylamino) -pyrrol-2-yl ] -formyl } thiomorpholine dihydrochloride (Compound 2)

Using 2b (124mg,0.31mmol) as a starting material, by a procedure similar to the second step in example 1, Compound 2 was obtained in a yield of 79.4% as an off-white solid. m.p.207-209 ℃,

¹H NMR(DMSO-d₆,300MHz)δ:4.66-4.64(m,1H,CHC=O),3.79-3.61(m,4H,CH ₂NH,CH ₂OH),3.35(b r,4H,CH₂NCH₂ in thiomorpholine),2.83-2.61(m,9H,CH₂SCH₂ in thiomorpholine,5H in pyrrolidine),0.93(s,6H,CH₃CCH₃).HR-MS(ESI-TOF⁺):C₁₄H₂₈N₃O₂s calculated 302.1897, found [ M-2HCl + H]⁺302.1893.

Example 3

The compound 34- { [ (2S,4S) -4- (4-pyridin-2-yl-methylamino) -pyrrol-2-yl ] -formyl } thiomorpholine trihydrochloride

First step preparation of 4- { [ (2S,4S) -N-tert-Butoxycarbonyl-4- (pyridin-2-yl-methylamino) -pyrrol-2-yl ] -formyl } thiomorpholine 3b

4- { [ (2S,4S) -4-amino N-tert-butoxycarbonyl-pyrrol-2-yl ] -formyl } thiomorpholine 6a (0.15g,0.48mmol) and 2-pyridinecarboxaldehyde (0.054g,0.50mmol) were dissolved in 10mL of dichloromethane, and after complete dissolution, a small amount of AcOH was added until the reaction pH =3-5, and the mixture was stirred at room temperature for 3 hours, the reaction was cooled to below 0 deg.C, sodium triacetoxyborohydride (0.315g,1.49mmol) was added, and the mixture was warmed to room temperature and stirred for 5 hours until the reaction was complete. Water was added to the reaction solution, followed by extraction with dichloromethane, and the organic layers were combined, washed with a saturated sodium bicarbonate solution and a saturated brine, respectively, and the organic layer was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. Separating by column chromatography, and eluting with dichloromethane-methanol (V: V = 100: 3) mixture. Intermediate 3b was obtained as 119mg of a yellow oil, yield 61.1%.

Second step preparation of 4- { [ (2S,4S) -4- (4-pyridin-2-yl-methylamino) -pyrrol-2-yl ] -formyl } thiomorpholine trihydrochloride (Compound 3)

Using 3b (119mg,0.29mmol) as a starting material, by a procedure similar to the second step in example 1, Compound 3 was obtained in the form of an off-white solid (88 mg, yield 73.1%). m.p.97-99 ℃,

¹H NMR(DMSO-d₆,300MHz)δ:8.65(1H,d,J=4.2Hz,in pyridine),7.96(t,1H,J=7.8Hz,in pyridine),7.64(d,1H,J=7.5Hz,in pyridine),7.49(t,1H,J=6.6Hz,inpyridine),4.70(s,1H,CHC=O,in pyrrolidine),4.37(s,2H,CH₂),4.06-4.04(m,1H,CHNH in pyrrolidine),3.85-3.62(m,6H CH₂NCH₂ in thiomorpholine,CH ₂NH inpyrrolidine),2.69-2.49(m,4H,CH₂SCH₂ in thiomorpholine),2.88-2.12(m,2H,CH ₂CHC=O in pyrrolidine);HR-MS(ESI-TOF⁺):C₁₅H₂₃N₄OS calculated 307.1587, found [ M-3HCl + H]⁺307.1583.

Example 4

The compound 44- { [ (2S,4S) -4- (3, 4-dimethoxy-benzylamino) -pyrrol-2-yl ] -formyl } thiomorpholine dihydrochloride

First step preparation of 4- { [ (2S,4S) -N-tert-Butoxycarbonyl-4- (3, 4-dimethoxy-benzylamino) -pyrrol-2-yl ] -formyl } thiomorpholine 4b

4- { [ (2S,4S) -4-amino-N-tert-butoxycarbonyl-pyrrol-2-yl ] -formyl } thiomorpholine 6a (0.15g,0.48mmol) and 3, 4-dimethoxybenzaldehyde (0.083g,0.50mmol) were dissolved in 10mL of dichloromethane, and after complete dissolution, a small amount of AcOH was added until the reaction pH =3-5, and stirred at room temperature for 3 hours, the reaction was cooled to below 0 deg.C, sodium triacetoxyborohydride (0.315g,1.50mmol) was added, and stirred at room temperature for 5 hours until the reaction was complete. To the reaction solution was added 20mL of water, followed by extraction with dichloromethane, and the organic layers were combined, washed with a saturated sodium bicarbonate solution and a saturated brine, respectively, and the organic layer was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. Separating by column chromatography, and eluting with dichloromethane-methanol (V: V = 50: 1) mixture. Intermediate 4b was obtained as a pale yellow oil, 148mg, 66.7% yield.

Second step preparation of 4- { [ (2S,4S) -4- (3, 4-dimethoxy-benzylamino) -pyrrol-2-yl ] -formyl } thiomorpholine dihydrochloride (Compound 4)

Using 4b (148mg,0.32mmol) as a starting material, the procedure was carried out in a similar manner to the second step in example 1 to give compound 4 as an off-white solid (122 mg) with a yield of 87.1%. m.p.161-163 ℃ in,

¹H NMR(DMSO-d₆,300MHz)δ:7.32(s,1H,aromatic),7.08(d,J=8.7Hz,1H,aromatic),6.99(d,J=8.4Hz,1H,aromatic),4.70-4.64(m,1H,CHC=O),3.85(m,1H,CHNH in pyrrolidine),3.79(s,3H,OCH₃),3.77(s,3H,OCH₃),3.71(m,4H,CH₂NCH₂in thiomorpholine),3.55-3.57(m,2H,CH ₂NH in pyrrolidine),2.81-2.63(m,4H,CH₂SCH₂ in thiomorpholine),2.85-1.99(m,2H,CH ₂CHC=O in pyrrolidine).HR-MS(ESI-TOF⁺):C₁₅H₂₁N₄O₃s calculated 366.1846, found [ M-2HCl + H]⁺366.1843.

Example 5

The compound 54- { [ (2S,4S) -4- (4-cyano-benzylamino) -pyrrol-2-yl ] -formyl } thiomorpholine dihydrochloride

First step preparation of 4- { [ (2S,4S) -1-tert-Butoxycarbonyl-4- (4-cyano-benzylamino) -pyrrol-2-yl ] -formyl } thiomorpholine 5b

4- { [ (2S,4S) -4-amino-N-tert-butoxycarbonyl-pyrrol-2-yl ] -formyl } thiomorpholine 6a (0.15g,0.48mmol) and potassium carbonate (0.276g,2.0mmol) were dissolved in 15mL of anhydrous DMF, 4-cyanobenzyl chloride (0.076g,0.50mmol) was added, and the mixture was heated at 80 ℃ for reaction for 8 hours. 30mL of ice water was added to the reaction system, the mixture was stirred for 30min, extracted with ethyl acetate, and the organic layers were combined, washed with water and saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was chromatographed on silica gel (200-300 mesh) using dichloromethane-methanol (V: V = 50: 1) as eluent to give intermediate 5b as pale yellow crystals 176mg, 85.3% yield

Second step preparation of 4- { [ (2S,4S) -4- (4-cyano-benzylamino) -pyrrol-2-yl ] -formyl } thiomorpholine dihydrochloride (Compound 5)

Using 5b (176mg,0.41mmol) as a starting material, by a procedure similar to the second step in example 1, Compound 5 was obtained in the form of an off-white solid (134 mg, yield 81.1%). m.p.194-196 ℃,

¹H NMR(DMSO-d₆,300MHz)δ:7.93(d,J=8.1Hz,2H,aromatic),7.81(d,J=7.8Hz,2H,aromatic),4.71-4.65(m,1H,CHC=O),4.30(s,2H,CH₂),3.79-3.58(m,5H,CH₂NCH₂ in thiomorpholine&CHNH in pyrrolidine),3.57-3.55(m,2H,CH ₂CHC=Oin pyrrolidine),2.87-2.09(m,2H,CH ₂CHC=O in pyrrolidine),2.74-2.49(m,4H,CH₂SCH₂ in thiomorpholine).HR-MS(ESI-TOF⁺):C₁₇H₂₂N₄OS calculated 331.1586, found [ M-2HCl + H]⁺331.1587.

Example 6

The compound 64- { [ (2S,4S) -4- (phenylamino) -pyrrol-2-yl ] -formyl } thiomorpholine dihydrochloride

First step preparation of 4- { [ (2S,4S) -N-tert-Butoxycarbonyl-4- (phenylamino) -pyrrol-2-yl ] -formyl } thiomorpholine 6b

4- { [ (2S,4S) -4-amino-N-t-butoxycarbonyl-pyrrol-2-yl ] -formyl } thiomorpholine 6a (0.101g,0.32mmol) and bromobenzene (0.046g,0.29mmol) were dissolved in 15mL of anhydrous toluene, and sodium t-butoxide (0.058g,0.60mmol), Davep (3mg) and palladium acetate (10mg) were added, and the mixture was refluxed for 10 hours under argon atmosphere, cooled to room temperature, and insoluble matter in the reaction system was filtered off and concentrated under reduced pressure. The crude product was separated by column chromatography using petroleum ether/ethyl acetate (V: V = 1: 1) mixture as eluent to give intermediate 6b as off-white crystals 73mg in 64.4% yield.

Second step preparation of 4- { [ (2S,4S) -4- (phenylamino) -pyrrol-2-yl ] -formyl } thiomorpholine dihydrochloride (Compound 6)

Using 6b (73mg,0.19mmol) as a starting material, by a procedure similar to the second step in example 1, Compound 6 was obtained in the form of a white powder (64 mg) with a yield of 92.5%. m.p.165-167 deg.C,

¹H NMR(DMSO-d₆,300MHz)δ:6.68-6.53(m,3H,aromatic),7.15-7.10(m,2H,aromatic),5.09-5.05(m,1H,CHC=O),4.12(br,1HCHNH in pyrrolidine),3.76-3.72(m,4H,CH₂NCH₂ in thiomorpholine),3.12(m,2H,CH ₂NH in pyrrolidine),2.70-2.55(m,4H,CH₂SCH₂ in thiomorpholine),2.34-1.60(m,2H,CH ₂CHC=O in pyrrolidine).HR-MS(ESI-TOF⁺):C₁₅H₂₂N₃OS calculated 292.1478, found [ M-2HCl + H]⁺292.1475.

Example 7

The compound 74- { [ (2S,4S) -4- (4-cyanophenyl-amino) -pyrrol-2-yl ] -formyl } thiomorpholine trihydrochloride

First step preparation of 4- { [ (2S,4S) -N-tert-Butoxycarbonyl-4- (4-cyanophenyl-amino) -pyrrol-2-yl ] -formyl } thiomorpholine 7b

4- { [ (2S,4S) -4-amino-N-t-butoxycarbonyl-pyrrol-2-yl ] -formyl } thiomorpholine 6a (0.103g,0.33mmol) and 4-cyanochlorobenzene (0.045g,0.33mmol) were dissolved in 15mL of anhydrous toluene, and sodium t-butoxide (0.057g,0.59mmol), Davep (3mg) and palladium acetate (10mg) were added, and the mixture was refluxed for 10 hours under argon atmosphere, cooled to room temperature, and insoluble matter was filtered off from the reaction system and concentrated under reduced pressure. The crude product was separated by column chromatography using petroleum ether/ethyl acetate (V: V = 1: 1) mixture as eluent to give intermediate 7b as off-white crystals 77mg in 56.1% yield.

Second step preparation of 4- { [ (2S,4S) -4- (4-cyanophenyl-amino) -pyrrol-2-yl ] -formyl } thiomorpholine trihydrochloride (Compound 7)

Using 7b (77mg,0.19mmol) as a starting material, by a procedure similar to the second step in example 1, Compound 7 was obtained as a white powder in a yield of 57mg, 77.1%. m.p.171-173 ℃,

¹H NMR(DMSO-d₆,300MHz)δ:7.51(d,J=8.1Hz,2H,aromatic),6.69(d,J=8.4Hz,2H,aromatic),4.65(m,1H,CHC=O),4.24-4.20(m,1H,CHNH in pyrrolidine),3.74-3.72(br,4H CH₂NCH₂ in thiomorpholine),3.50-3.13(m,2H,CH ₂NH inpyrrolidine),2.81-1.58(m,2H,CH ₂CHC=O in pyrrolidine),2.59(br,4H,CH₂SCH₂ inthiomorpholine).HR-MS(ESI-TOF⁺):C₁₆H₂₁N₄OS calculated 317.1431, found [ M-2HCl + H]⁺317.1430.

Example 8

The compound 84- { [ (2S,4S) -4- (4-trifluoromethylphenyl-amino) -pyrrol-2-yl ] -formyl } thiomorpholine dihydrochloride

First step preparation of 4- { [ (2S,4S) -N-tert-Butoxycarbonyl-4- (4-trifluoromethylphenyl-amino) -pyrrol-2-yl ] -formyl } thiomorpholine 8b

4- { [ (2S,4S) -4-amino-N-t-butoxycarbonyl-pyrrol-2-yl ] -formyl } thiomorpholine 6a (0.104g,0.33mmol) and 4-trifluoromethylbromobenzene (0.074g,0.33mmol) were dissolved in 15mL of anhydrous toluene, and sodium t-butoxide (0.061g,0.63mmol), Davep (3mg) and palladium acetate (10mg) were added, and the mixture was refluxed under argon atmosphere for 10 hours, cooled to room temperature, and insoluble matter was filtered off and concentrated under reduced pressure. The crude product was isolated by column chromatography using petroleum ether/ethyl acetate (V: V = 1: 1) mixture as eluent to give intermediate 8b as a pale yellow oil 93mg with 61.4% yield.

Second step preparation of 4- { [ (2S,4S) -4- (4-trifluoromethylphenyl-amino) -pyrrol-2-yl ] -formyl } thiomorpholine dihydrochloride (Compound 8)

Using 8b (93mg,0.20mmol) as a starting material, by a procedure similar to the second step in example 1, Compound 8 was obtained as a white powder in an yield of 72mg, 83.3%. m.p.141-143 ℃,

¹H NMR(DMSO-d₆,300MHz)δ:7.44(d,J=8.7Hz,2H,aromatic),6.72(d,J=8.7Hz,2H,aromatic),4.66(br,1H,CHC=O),4.22-4.19(m,1H,CHNH in pyrrolidine),3.78-3.24(m,4H CH₂NCH₂ in thiomorpholine),3.35-3.14(br,2H,CH ₂NH inpyrrolidine),2.90-1.59(m,2H,CH ₂CHC=O in pyrrolidine),2.80-2.64(m,4H,CH₂SCH₂ in thiomorpholine).HR-MS(ESI-TOF⁺):C₁₆H₂₁N₃OSF₃calculated value 360.1353, found [ M-2HCl + H]⁺360.1357.

Example 9

The compound 94- { [ (2S,4S) -4- (4-methylphenyl-amino) -pyrrol-2-yl ] -formyl } thiomorpholine dihydrochloride

First step preparation of 4- { [ (2S,4S) -N-tert-Butoxycarbonyl-4- (4-methylphenyl-amino) -pyrrol-2-yl ] -formyl } thiomorpholine 9b

4- { [ (2S,4S) -4-amino-N-t-butoxycarbonyl-pyrrol-2-yl ] -formyl } thiomorpholine 6a (0.102g,0.32mmol) and 4-bromotoluene (0.056g,0.33mmol) were dissolved in 15mL of anhydrous toluene, and sodium t-butoxide (0.061g,0.64mmol), Davep (3mg) and palladium acetate (10mg) were added, and the mixture was refluxed under argon atmosphere for 16 hours, cooled to room temperature, and insoluble matter was filtered off from the reaction system and concentrated under reduced pressure. The crude product was separated by column chromatography using petroleum ether/ethyl acetate (V: V = 1: 1) mixture as eluent to give intermediate 9b as a pale yellow oil 78mg with a yield of 60.2%.

Second step preparation of 4- { [ (2S,4S) -4- (4-methylphenyl-amino) -pyrrol-2-yl ] -formyl } thiomorpholine dihydrochloride (Compound 9)

Using 9b (78mg,0.19mmol) as a starting material, by a procedure similar to the second step in example 1, Compound 9 was obtained as a white powder in a yield of 67mg and 93.3%. m.p.167-169 ℃ of,

¹H NMR(DMSO-d₆,300MHz)δ:7.30(d,J=7.8Hz,2H,aromatic),6.94(d,J=7.5Hz,2H,aromatic),4.95(br,1H,CHC=O),4.44(m,1H,CHNH in pyrrolidine),4.09-4.02(m,4H,CH₂NCH₂ in thiomorpholine),3.78-3.48(br,2H,CH ₂CHC=O inpyrrolidine),3.16-1.96(m,2H,CH ₂CHC=O in pyrrolidine),3.04-2.81(m,4H,CH₂SCH₂ in thiomorpholine).HR-MS(ESI-TOF⁺):C₁₆H₂₄N₃OS calculated 306.1627, found [ M-2HCl + H]⁺306.1635.

Example 10

The compound 104- { [ (2S,4S) -4- (3-nitrophenyl-amino) -pyrrol-2-yl ] -formyl } thiomorpholine dihydrochloride

First step preparation of 4- { [ (2S,4S) -N-tert-Butoxycarbonyl-4- (3-nitrophenyl-amino) -pyrrol-2-yl ] -formyl } thiomorpholine 10b

4- { [ (2S,4S) -4-amino-N-t-butoxycarbonyl-pyrrol-2-yl ] -formyl } thiomorpholine 6a (0.101g,0.32mmol) and 3-nitrochlorobenzene (0.053g,0.34mmol) were dissolved in 15mL of anhydrous toluene, and sodium t-butoxide (0.060g,0.63mmol), Davep (3mg) and palladium acetate (10mg) were added, and the mixture was refluxed for 10 hours under an argon atmosphere, cooled to room temperature, and insoluble matter in the reaction system was filtered off and concentrated under reduced pressure. The crude product was isolated by column chromatography using a mixture of petroleum ether and ethyl acetate (V: V = 1: 1) as eluent to give intermediate 10b as a yellow oil in a yield of 65.9% as 92 mg.

Second step preparation of 4- { [ (2S,4S) -4- (3-nitrophenyl-amino) -pyrrol-2-yl ] -formyl } thiomorpholine dihydrochloride (Compound 10)

Using 10b (92mg,0.21mmol) as a starting material, by a procedure similar to the second step in example 1, Compound 10 was obtained in the form of a yellow powder (62 mg) with a yield of 72.2%. m.p.237-239 ℃,

¹H NMR(DMSO-d₆,300MHz)δ:7.44-7.37(m,3H,aromatic),7.02(d,J=8.4Hz,1H,aromatic),4.66(m,1H,CHC=O),4.26-4.22(m,1H,CHNH in pyrrolidine),3.73-3.49(4H,CH₂NCH₂ in thiomorpholine),3.15-2.82(m,2H,CH ₂NH in pyrrolidine),2.72-1.65(m,2H,CH ₂CHC=O in pyrrolidine),2.60-2.56(m,4H,CH₂SCH₂ inthiomorpholine).HR-MS(ESI-TOF⁺):C₁₅H₂₁N₄O₃s calculated 337.1329, found [ M-2HCl + H]⁺337.1337.

Example 11

The compound 114- { [ (2S,4S) -4- (pyridin-2-yl-amino) -pyrrol-2-yl ] -formyl } thiomorpholine trihydrochloride

First step preparation of 4- { [ (2S,4S) -N-tert-Butoxycarbonyl-4- (pyridin-2-yl-amino) -pyrrol-2-yl ] -formyl } thiomorpholine 10b

4- { [ (2S,4S) -4-amino-N-t-butoxycarbonyl-pyrrol-2-yl ] -formyl } thiomorpholine 6a (0.101g,0.32mmol) and 2-chloropyridine (0.038g,0.34mmol) were dissolved in 15mL of anhydrous toluene, and sodium t-butoxide (0.060g,0.63mmol), Davep (3mg) and palladium acetate (10mg) were added, and the mixture was refluxed for 10 hours under an argon atmosphere, cooled to room temperature, and insoluble matter in the reaction system was filtered off and concentrated under reduced pressure. The crude product was isolated by column chromatography using a mixture of petroleum ether and ethyl acetate (V: V = 1: 1) as eluent to give intermediate 11b as a yellow oil in a yield of 61.4% as 77 mg.

Second step preparation of 4- { [ (2S,4S) -4- (pyridin-2-yl-amino) -pyrrol-2-yl ] -formyl } thiomorpholine trihydrochloride (Compound 11)

Using 11b (77mg,0.196mmol) as a starting material, the same procedures as in the second step of example 1 were carried out to give compound 11 as a pale yellow solid, 67mgThe yield thereof was found to be 85.2%. m.p.183-185 deg.c,

¹H NMR(DMSO-d₆,300MHz)δ:8.00(d,J=8.7Hz,1H in pyridine),7.91(d,J=6.0Hz,1H in pyridine),7.11(d,J=9.3Hz,1H in pyridine),7.00(t,J=6.6Hz,1H inpyridine),4.74-4.68(m,1H,CHC=O),3.90(m,1H,CHNH in pyrrolidine),3.83-3.81(m,4H,CH₂NCH₂ in thomorpholine),3.56-3.05(m,2H,CH ₂NH in pyrrolidine),2.69-2.61(m,4H,CH₂SCH₂ in thiomorpholine),2.4-1.96(m,2H,CH ₂CHC=O inpyrrolidine).HR-MS(ESI-TOF⁺):C₁₄H₂₁N₄OS calculated 293.1431, found [ M-3HCl + H]⁺293.1436.

Example 12

The compound 124- { [ (2S,4S) -4- (5-cyanopyridin-2-yl-amino) -pyrrol-2-yl ] -formyl } thiomorpholine trihydrochloride

First step preparation of 4- { [ (2S,4S) -N-tert-Butoxycarbonyl-4- (5-cyanopyridin-2-yl-amino) -pyrrol-2-yl ] -formyl } thiomorpholine 12b

4- { [ (2S,4S) -4-amino-N-t-butoxycarbonyl-pyrrol-2-yl ] -formyl } thiomorpholine 6a (0.102g,0.33mmol) and 2-chloro-5-cyanopyridine (0.046g,0.34mmol) were dissolved in 15mL of anhydrous toluene, and sodium t-butoxide (0.060g,0.63mmol), Davep (3mg) and palladium acetate (10mg) were added, and the mixture was refluxed under an argon atmosphere for 10 hours, cooled to room temperature, and insoluble matter was filtered off from the reaction system and concentrated under reduced pressure. The crude product was separated by column chromatography using a mixture of petroleum ether and ethyl acetate (V: V = 1: 1) as eluent to give the intermediate 12b as a yellow oil 69mg, 50.1% yield.

Second step preparation of 4- { [ (2S,4S) -4- (5-cyanopyridin-2-yl-amino) -pyrrol-2-yl ] -formyl } thiomorpholine trihydrochloride (Compound 12)

Using 12b (69mg,0.16mmol) as a starting material, by the similar procedure as in the second step in example 1, Compound 12 was obtained as a pale yellow solid, 51mg, in 74.8% yield. m.p.102-104 ℃,

¹H NMR(DMSO-d₆,300MHz)δ:8.46(s,1H in pyridine),7.77(d,J=8.7Hz,1Hin pyridine),6.61(d,J=9.0Hz,1H in pyridine),4.68(m,1H,CHC=O),3.78-3.72(m,4H,CH₂NCH₂ in thomorpholine),3.50(m,1H,CHNH in pyrrolidine),3.84-2.17(m,2H,CH ₂NH in pyrrolidine),2.73-1.80(m,2H,CH₂CHC=O in pyrrolidine),2.61-2.50(m,4H,CH₂SCH₂ in thiomorpholine).HR-MS(ESI-TOF⁺):C₁₅H_20N5OS calculated 318.1383, found [ M-3HCl + H]⁺318.1387.

Example 13

The compound 134- { [ (2S,4S) -4- (3-cyanopyridin-2-yl-amino) -pyrrol-2-yl ] -formyl } thiomorpholine trihydrochloride

First step preparation of 4- { [ (2S,4S) -N-tert-Butoxycarbonyl-4- (3-cyanopyridin-2-yl-amino) -pyrrol-2-yl ] -formyl } thiomorpholine 13b

4- { [ (2S,4S) -4-amino-N-t-butoxycarbonyl-pyrrol-2-yl ] -formyl } thiomorpholine 6a (0.102g,0.33mmol) and 2-chloro-5-cyanopyridine (0.046g,0.34mmol) were dissolved in 15mL of anhydrous toluene, and sodium t-butoxide (0.060g,0.66mmol), Davep (3mg) and palladium acetate (10mg) were added, and the mixture was refluxed under an argon atmosphere for 10 hours, cooled to room temperature, and insoluble matter was filtered off from the reaction system and concentrated under reduced pressure. The crude product was isolated by column chromatography using a mixture of petroleum ether and ethyl acetate (V: V = 1: 1) as eluent to give intermediate 13b as a yellow oil in 47.2% yield (65 mg).

Second step preparation of 4- { [ (2S,4S) -4- (3-cyanopyridin-2-yl-amino) -pyrrol-2-yl ] -formyl } thiomorpholine trihydrochloride (Compound 13)

Using 13b (65mg,0.156mmol) as a starting material, by the similar procedure as in the second step in example 1, Compound 13 was obtained as a pale yellow solid (43 mg, yield 64.7%). m.p.156-158 ℃,

¹H NMR(DMSO-d₆,300MHz)δ:8.32(d,J=4.8Hz,1H in pyridine),7.98(d,J=7.5Hz,1H in pyridine),7.21(d,J=7.2Hz,1H in pyridine),4.85-4.80(m,1H,CHC=O),4.65(m,1H,CHNH in pyrrolidine),3.83-3.70(m,4H,CH₂NCH₂ inthomorpholine),3.46-3.30(m,2H,CH ₂NH in pyrrolidine),2.84-1.86(m,2H,CH ₂CHC=O in pyrrolidine),2.72-2.60(m,4H,CH₂SCH₂ in thiomorpholine).HR-MS(ESI-TOF⁺):C₁₅H₂₀N₅OS calculated 318.1383, found [ M-3HCl + H]⁺318.1385.

Example 14

The compound 144- { [ (2S,4S) -4- (quinolin-2-yl-amino) -pyrrol-2-yl ] -formyl } thiomorpholine trihydrochloride

First step preparation of 4- { [ (2S,4S) -N-tert-Butoxycarbonyl-4- (quinolin-2-yl-amino) -pyrrol-2-yl ] -formyl } thiomorpholine 14b

4- { [ (2S,4S) -4-amino-N-t-butoxycarbonyl-pyrrol-2-yl ] -formyl } thiomorpholine 6a (0.102g,0.32mmol) and 2-chloroquinoline (0.054g,0.33mmol) were dissolved in 15mL of anhydrous toluene, and sodium t-butoxide (0.060g,0.63mmol), Davep (3mg) and palladium acetate (10mg) were added, and the mixture was refluxed for 10 hours under an argon atmosphere, cooled to room temperature, and insoluble matter in the reaction system was filtered off and concentrated under reduced pressure. The crude product was isolated by column chromatography using petroleum ether/ethyl acetate (V: V = 1: 1) as eluent to give intermediate 14b as yellow crystals 101mg in 71.4% yield.

Second step preparation of 4- { [ (2S,4S) -4- (quinolin-2-yl-amino) -pyrrol-2-yl ] -formyl } thiomorpholine trihydrochloride (Compound 14)

Using 14b (101mg,0.23mmol) as a starting material, by a procedure similar to the second step in example 1, Compound 14 was obtained in 96mg as an off-white solid with a yield of92.5%。m.p.120-122℃,

¹H NMR(DMSO-d₆,300MHz)δ:8.37(d,J=8.7Hz,1H in quinoline),8.28(br,1H in quinoline),7.94(d,J=7.8Hz,1H in quinoline),7.81(t,J=7.5Hz,1H inquinoline),7.53(t,J=7.5Hz,1H in quinoline),7.17(d,J=8.7Hz,1H in quinoline),5.03(br,1H,CHC=O),4.73(m,1H,CHNH in pyrrolidine),3.82-3.70(m,4H,CH₂NCH₂ in thomorpholine),3.51-3.45(m,2H,CH ₂NH in pyrrolidine),3.15-2.71(m,2H,CH ₂CHC=O in pyrrolidine),2.65-2.51(m,4H,CH₂SCH₂ in thiomorpholine).HR-MS(ESI-TOF⁺):C₁₈H₂₃N₄OS calculated 343.1587, found [ M-3HCl + H]⁺343.1588.

Example 15

Compound 154- { [ (2S,4S) -4- (4-Methylbenzenesulfonyl-amino) -pyrrol-2-yl ] -formyl } thiomorpholine hydrochloride

First step preparation of 4- { [ (2S,4S) -N-tert-Butoxycarbonyl-4- (4-methylbenzenesulfonyl-amino) -pyrrol-2-yl ] -formyl } thiomorpholine 15b

4- { [ (2S,4S) -4-amino-N-tert-butoxycarbonyl-pyrrol-2-yl ] -formyl } thiomorpholine 6a (0.156g,0.50mmol) was dissolved in 15mL of anhydrous dichloromethane, triethylamine (0.15g,1.5mmol) was added, the reaction system was cooled to 0 ℃ or lower, p-toluenesulfonyl chloride (0.114g,0.60mmol) was slowly added dropwise, the reaction was continued at low temperature for 2 hours until completion, the reaction solution was warmed to room temperature, the organic phase was washed with water and saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was separated by column chromatography using petroleum ether/ethyl acetate (V: V = 1: 1) mixture as eluent to give intermediate 15b as a white crystal 176mg, yield 75.1%.

Second step preparation of 4- { [ (2S,4S) -4- (4-Methylbenzenesulfonyl-amino) -pyrrol-2-yl ] -formyl } thiomorpholine hydrochloride (Compound 15)

Using 15b (150mg,0.32mmol) as a starting material, by a procedure similar to the second step in example 1, compound 15 was obtained in the form of an off-white solid (118 mg) with a yield of 90.8%. m.p.209-211 ℃,

¹H NMR(DMSO-d₆,300MHz)δ:8.11(d,J=6.0Hz,1H,NHSO₂),7.73(d,J=7.8Hz,2H,aromatic),7.44(d,J=7.8Hz,2H,aromatic),4.50(t,J＝8.7Hz,3H,CHC=O),3.81-3.68(m,5H,CH₂NCH₂ in thiomorpholine,CHNH in pyrrolidine),3.63(s,3H,CH₃),3.04-3.21(m,2H,CH ₂NH in pyrrolidine);2.50(br,4H,CH₂SCH₂ inthiomorpholine),2.46-1.56(m,2H,CH ₂CHC=O in pyrrolidine).HR-MS(ESI-TOF⁺):C₁₆H₂₄N₃O₃S₂calculated value 370.1254, found [ M-HCl + H]⁺370.1251.

Example 16

The compound 164- { [ (2S,4S) -4- (4-Nitrobenzenesulfonyl-amino) -pyrrol-2-yl ] -formyl } thiomorpholine hydrochloride

First step preparation of 4- { [ (2S,4S) -N-tert-Butoxycarbonyl-4- (4-nitrobenzenesulfonyl-amino) -pyrrol-2-yl ] -formyl } thiomorpholine 16b

4- { [ (2S,4S) -4-amino-N-tert-butoxycarbonyl-pyrrol-2-yl ] -formyl } thiomorpholine 6a (0.156g,0.50mmol) was dissolved in 15mL of anhydrous dichloromethane, triethylamine (0.15g,1.50mmol) was added, the reaction system was cooled to below 0 ℃, p-nitrobenzenesulfonyl chloride (0.128g,0.58mmol) was slowly added dropwise, the reaction was continued at low temperature for 4 hours until completion, the reaction solution was warmed to room temperature, the organic phase was washed with water and saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was separated by column chromatography using petroleum ether/ethyl acetate (V: V = 1: 1) mixture as eluent to give intermediate 16b as a white crystal 170mg with a yield of 68.0%.

Second step preparation of 4- { [ (2S,4S) -4- (4-Nitrobenzenesulfonyl-amino) -pyrrol-2-yl ] -formyl } thiomorpholine hydrochloride (Compound 16)

Using 16b (150mg,0.30mmol) as a starting material, by a procedure similar to the second step in example 1, compound 16 was obtained in the form of an off-white solid (107 mg) with a yield of 81.8%. m.p.209-211 ℃,

¹H NMR(DMSO-d₆,300MHz)δ:8.61(br,1H,NH),8.45(d,J=8.7Hz,2H,aromatic),8.09(d,J=9.0Hz,2H,aromatic),4.50(t,J=9.3Hz,1H,CHC=O),3.86(br,1H,CHNH in pyrrolidine),3.74&3.61(s,2H)(s,2H,CH₂NCH₂ in thiomorpholine),3.26-3.01(m,2H,CH ₂NH in pyrrolidine),2.591.63(m,6HCH ₂CHC=O in pyrrolidine,CH₂SCH₂ in thiomorpholine).HR-MS(ESI-TOF⁺):C₁₅H₂₁N₄O₅S₂calculated value 401.0948, found [ M-HCl + H]⁺401.0953.

Example 17

The compound 174- { [ (2S,4S) -4- [ (5-cyanopyridin-2-yl) -piperidin-4-yl-amino ] -pyrrol-2-yl ] -formyl } thiomorpholine tetrah-ydrochloride

First step preparation of 5-cyanopyridin-2-yl-piperidin-4-one 7a

4-Piperidinone hydrochloride (0.272g,2.0mmol) and 5-cyano-2-chloropyridine (0.276g,2.0mmol) were dissolved in 20mL of anhydrous DMF, potassium carbonate (0.56g,4.1mmol) was added, the reaction was heated at 80 ℃ for 10 hours, 30mL of ice water was added to the reaction system and stirred for 30min, extraction was performed with ethyl acetate, the organic layers were combined, washed with water and saturated brine, respectively, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. Separating the crude product with column, eluting with petroleum ether/ethyl acetate (V: V = 1: 1) mixture to obtain intermediate 7a as pale yellow solid 280mg with 69.7% yield

Second step preparation of 4- { [ (2S,4S) -N-tert-Butoxycarbonyl-4- [ (5-cyanopyridin-2-yl) -piperidin-4-yl-amino) ] -pyrrol-2-yl ] -formyl } thiomorpholine 17b

4- { [ (2S,4S) -4-amino-N-tert-butoxycarbonyl-pyrrol-2-yl ] -formyl } thiomorpholine 6a (0.103g,0.33mmol) and 7a (0.066g,0.33mmol) were dissolved in 10mL of dichloromethane, and after complete dissolution, a small amount of AcOH was added to the reaction pH =3-5, stirred at room temperature for 3 hours, the reaction was cooled to below 0 deg.C, sodium triacetoxyborohydride (0.211g,1.0mmol) was added, warmed to room temperature, and stirred for 8 hours until the reaction was complete. To the reaction solution was added 20mL of water, followed by extraction with dichloromethane, and the organic layers were combined, washed with a saturated sodium bicarbonate solution and a saturated brine, respectively, and the organic layer was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product is separated by column chromatography, and a dichloromethane-methanol (V: V = 50: 1) mixed solution is used as an eluent. Intermediate 17b was obtained as a pale yellow solid 112mg, yield 67.7%.

Third step preparation of 4- { [ (2S,4S) -4- [ (5-cyanopyridin-2-yl) -piperidin-4-yl-amino ] -pyrrol-2-yl ] -formyl } thiomorpholine tetrahydrate hydrochloride (Compound 17)

Using 17b (112mg,0.22mmol) as a starting material, by a procedure similar to the second step in example 1, Compound 17 was obtained in the form of an off-white solid (105 mg) with a yield of 87.4%. m.p. > 250 ℃,

¹H NMR(DMSO-d₆,300MHz)δ:8.52(s,1H in pyridine),7.91(s,1H,inpyridine),7.04(m,1H,in pyridine),4.67(s,1H CHC=O),4.57(br,2H,CHNHCH),3.82-3.56(m,8H,CH₂NCH₂ in piperidine,CH₂NCH₂ in thiomorpholine),3.00-2.66(m,8H CH ₂CHCH ₂ in piperidine,CH₂SCH₂ in thiomorpholine),1.97-1.60(m,4H,CH ₂NH,CH ₂CH in pyrrolidine).HR-MS(ESI-TOF⁺):C₂₀H₃₀N₆OS calculated 401.2118, found [ M-4HCl + H]⁺401.2056.

Example 18

The compound 184- { [ (2S,4S) -4- [ (4-cyanobenzyl) -piperidin-4-yl-amino ] -pyrrol-2-yl ] -formyl } thiomorpholine trihydrochloride

First step preparation of 5-cyanobenzylpiperidin-4-one 8a

4-Piperidinone hydrochloride (0.272g,2.0mmol) and 5-cyanobenzyl chloride (0.301g,2.0mmol) were dissolved in 20mL of anhydrous DMF, potassium carbonate (0.56g,4.1mmol) was added, the reaction system was heated at 80 ℃ for 10 hours, 30mL of ice water was added to the reaction system and stirred for 30min, extraction was performed with ethyl acetate, the organic layers were combined, washed with water and saturated brine, respectively, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. Separating the crude product by silica gel column chromatography, eluting with petroleum ether/ethyl acetate (V: V = 1: 1) mixture to obtain intermediate 5-cyanobenzylpiperidine-4-one 8a as pale yellow solid 362mg with 84.6% yield

Second step preparation of 4- { [ (2S,4S) -N-tert-Butoxycarbonyl-4- [ (4-cyanobenzyl) -piperidin-4-yl-amino ] -pyrrol-2-yl ] -formyl } thiomorpholine 18b

4- { [ (2S,4S) -4-amino-N-tert-butoxycarbonyl-pyrrol-2-yl ] -formyl } thiomorpholine 6a (0.103g,0.33mmol) and 8a (0.068g,0.32mmol) were dissolved in 10mL of dichloromethane, and after complete dissolution, a small amount of AcOH was added to the reaction pH =3-5, stirred at room temperature for 3 hours, the reaction was cooled to below 0 deg.C, sodium triacetoxyborohydride (0.211g,1.0mmol) was added, warmed to room temperature, and stirred for 10 hours to complete the reaction. To the reaction solution was added 20mL of water, followed by extraction with dichloromethane, and the organic layers were combined, washed with a saturated sodium bicarbonate solution and a saturated brine, respectively, and the organic layer was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product is separated by column chromatography, and a dichloromethane-methanol (V: V = 50: 1) mixed solution is used as an eluent. Intermediate 18b was obtained as a pale yellow solid, 106mg, yield 62.5%.

Third step preparation of 4- { [ (2S,4S) -4- [ (4-cyanobenzyl) -piperidin-4-yl-amino ] -pyrrol-2-yl ] -formyl } thiomorpholine trihydrochloride (Compound 18)

Using 18b (106mg,0.21mmol) as a starting material, by a procedure similar to the second step in example 1, compound 18 was obtained in the form of an off-white solid (88 mg, yield 80.3%). m.p. > 250 ℃,

¹H NMR(DMSO-d₆,300MHz)δ:7.97(d,J=8.4Hz,2H,aromatic),7.83(d,J=8.4Hz,2H,aromatic),4.67(br,1H CHC=O),4.36(br,1H,NHCH in pyrrolidine),4.07-3.99(m,4H,CH₂NCH₂ in piperidine),3.78(br,1H,NHCH in piperidine),3.44-3.71(m,8H,Ar-CH₂,CH ₂NH in pyrrolidine,CH₂NCH₂ in thiomorpholine),2.86-2.24(m,8H,CH ₂CHCH ₂ in piperidine,CH₂NCH₂ in thiomorpholine),2.07(br,2H,CH ₂CH in pyrrolidine).HR-MS(ESI-TOF⁺):C₂₂H₃₂N₅OS calculated 414.2322, found [ M-3HCl + H]⁺414.2246.

Test examples section

Test example 1 in vitro screening method for DPP-IV inhibitor

Reagent:

1. reaction substrate: Gly-Pro-p-nitroanilide hydrochloride (Sigma, G0513) using ddH₂Dissolving O to obtain 0.026M stock solution, and storing at-20 deg.C in dark.

DPP-IV enzyme: recombinant human DPP-IV protein (Sigma, D4943) is preserved at-20 ℃.

3. A sample to be tested: DMSO is dissolved in 10^-2M, storing at 4 ℃.

4. Positive control drug: sitagliptin is dissolved in DMSO to 10^-2M, storing at 4 ℃.

5.2×Hepes buffer：1.6g NaCl，0.074g KCl，0.027g Na₂HPO₄·2H₂O, 0.2g Glucose, 1g Hepes in 90ml ddH₂Adjusting pH to 7.05 with NaOH, diluting to 100ml, filtering at 0.22 μm, and storing at 4 deg.C.

Tis-HCl buffer: 6.06g Tris in 1L distilled water, adjusted to pH 8.0 with HCl.

Detection method

ddH for sample and positive drug Sitagliptin₂Diluting O to 10%^-4M, DPP-IV was formulated with Tis/HCl buffer (pH = 8.0) at a concentration of 2 mU/ml. The substrate Gly-Pro-p-nitroanilide working solution was diluted with Hepes buffer (pH = 7.05) at a concentration of 0.26 mM. The experiment is provided with a negative control group, a positive control group and a sample group. The total volume of the reaction was 100. mu.l, in which ddH was added to the negative control group₂O10 mul, DPP-IV enzyme working solution 50 mul and substrate working solution 40 mul; adding Sitagliptin solution 10 μ l, DPP-IV enzyme working solution 50 μ l and substrate working solution 40 μ l into the positive control group; the sample set was loaded with 10. mu.l of sample solution, 50. mu.l of DPP-IV working solution and 40. mu.l of substrate working solution. The inhibition of DPP-IV by samples was evaluated by monitoring the change in absorbance of the reaction at 405nm at 37 ℃ for 60minThe application is as follows.

The inhibition rate of the sample on DPP-IV is calculated as follows:

inhibition (%) = (Δ OD)_60-0Negative control group-. DELTA.OD_60-0Sample set)/Δ OD_60-0Negative control group X100%

The positive control Sitagliptin is generally considered to be at 10^-5The inhibition rate of M is 90-100%, the reaction of the experiment is reliable, and the inhibition rate of the sample is more than 40%, so that the reaction is effective.

Positive Compound IC ₅₀ The calculation of (2):

1. for the primary screening (concentration of test compound 10)^-5M) active compounds (i.e. inhibitory activity greater than 50%), with different concentration gradients set i.e. 10^-8，10^-7，10^-6And 10^-5M DPP-IV inhibition experiments are carried out.

2. Drawing a concentration-reaction curve by using the reaction concentration and the inhibition rate of the compound, statistically processing the curve to obtain a fitting formula, wherein Y is the inhibition rate, X is the concentration of the compound, and when Y is 50 percent, namely the activity of 50 percent of the inhibition rate is reached, the corresponding concentration of the compound is the half effective Inhibition Concentration (IC) of the compound₅₀) The test results are shown in Table 1.

TABLE 1 results of enzyme level Activity assay of Compounds of examples of the invention

And (4) conclusion:

of the compounds tested, 12 examples had greater than 50% in vitro DPP-IV inhibitory activity, of which 9 examples had IC₅₀Reaching the level of μ M. Example 14 has the highest in vitro DPP-IV inhibitory activity.

Test example 2 DPP-IV inhibitor selectivity evaluation method (DPP 8/9 inhibitory Activity in vitro evaluation method)

1. Reaction substrate: the method is the same as the DPP-IV inhibitor in vitro screening method.

DPP8/9 enzyme: the recombinant human DPP8/9 protein is purified and freeze-dried and then stored at-20 ℃.

3. Sample preparation: the method is the same as the DPP-IV inhibitor in vitro screening method.

4. Positive control drug: the compound (S) -4- (2-amino- (R) -3-methylpentanoyl) isoindoline hydrochloride was dissolved in DMSO to give a solution of 10^-2M, storing at 4 ℃.

5.2 × Hepes buffer: the method is the same as the DPP-IV inhibitor in vitro screening method.

Tis/HCl buffer: the method is the same as the DPP-IV inhibitor in vitro screening method.

Evaluation method

Samples and Positive drugs ddH₂Diluting O to 10%_-4M, DPP8 enzyme was formulated with Tis/HCl buffer (pH = 8.0) at a concentration of 60ng/ml and DPP9 enzyme was formulated at a concentration of 40 ng/ml. The substrate Gly-Pro-p-nitroanilide working solution was diluted with Hepes buffer (pH = 7.05) at a concentration of 0.52 mM. The experiment is provided with a negative control group, a positive control group and a sample group. The total volume of the reaction was 100. mu.l, in which ddH was added to the negative control group₂O10 mul, DPP8/9 enzyme working solution 50 mul and substrate working solution 40 mul; adding 10 mul of positive control solution, 50 mul of DPP8/9 enzyme working solution and 40 mul of substrate working solution into a positive control group; the sample set was loaded with 10. mu.l of sample solution, 50. mu.l of DPP8/9 enzyme working solution and 40. mu.l of substrate working solution. The selectivity of the DPP-IV inhibitor, i.e. DPP8/9 inhibition, was evaluated by monitoring the change in absorbance of the reaction at 405nm at 37 ℃ for 60 min. The selectivity of the DPP-IV inhibitor (DPP 8/9 inhibitory activity) is calculated as follows:

The test results are shown in Table 2

The inhibition rate of the positive drug at the same concentration is 96.7%.

And (4) conclusion:

11 tested compounds with DPP-IV inhibitory activity have no obvious inhibitory activity on DPP8/9, which shows that the 11 compounds have high DPP-IV selectivity.

Claims

1. A compound represented by the general formula (I) and stereoisomers thereof, and pharmaceutically acceptable salts thereof,

wherein,

r is selected from hydrogen and C_1-6Alkyl radical, C_3-8Cycloalkyl, aryl-C_1-3Alkyl, aryl-C_3-8Cycloalkyl, heteroaryl-C_1-3Alkyl radicalheteroaryl-C_3-8Cycloalkyl, heterocycloalkyl containing from 3 to 8 (e.g. 3 to 7, 3 to 6, 3 to 5, or 3 to 4) carbon atoms and from 1 to 3 (e.g. 1 to 2, 1, 2, or 3) heteroatoms selected from nitrogen, oxygen, sulfur, aryl-C_3-8Heterocycloalkyl, heteroaryl-C_3-8Heterocycloalkyl, benzyl-C_3-8Heterocycloalkyl radical, C_1-6Alkyl formyl radical, C_3-8Cycloalkyl-formyl, aryl-formyl, benzyl-formyl, heteroaryl-formyl, C_1-6Alkylsulfonyl radical, C_3-8Cycloalkylsulfonyl, arylsulfonyl, heteroarylsulfonyl, benzylsulfonyl;

wherein said alkyl, cycloalkyl, aryl, heteroaryl, heterocycloalkyl, and benzyl are optionally substituted with 1 to 4 (e.g., 1 to 3, 1 to 2, 1, 2, or 3) groups selected from: H. hydroxy, halogen, cyano, amino, nitro, trifluoromethyl, C_1-6Alkyl radical, C_1-6Alkyloxy, and NR₁R₂And R is₁And R₂Independently selected from H, C_1-6An alkyl group.

2. A compound according to claim 1, said compound being represented by the general formula (IA), and stereoisomers and pharmaceutically acceptable salts thereof

Wherein,

r is selected from hydrogen and C_1-6Alkyl radical, C_3-8Cycloalkyl, aryl-C_1-3Alkyl, aryl-C_3-8Cycloalkyl, heteroaryl-C_1-3Alkyl, heteroaryl-C_3-8Cycloalkyl, heterocycloalkyl containing from 3 to 8 (e.g. 3 to 7, 3 to 6, 3 to 5, or 3 to 4) carbon atoms and from 1 to 3 (e.g. 1 to 2, 1, 2, or 3) heteroatoms selected from nitrogen, oxygen, sulfur, aryl-C_3-8Heterocycloalkyl, heteroaryl-C_3-8Heterocycloalkyl, benzyl-C_3-8Heterocycloalkyl radical, C_1-6Alkyl formyl radical, C_3-8Cycloalkylcarboxoyl radicalArylformyl, benzylformyl, heteroarylformyl, C_1-6Alkylsulfonyl radical, C_3-8Cycloalkylsulfonyl, arylsulfonyl, heteroarylsulfonyl, benzylsulfonyl;

3. A compound according to any one of claims 1 to 2 and stereoisomers thereof, pharmaceutically acceptable salts thereof, wherein

R is selected from C_1-6Alkyl, aryl, heteroaryl, C_3-8Nitrogen-containing heterocycloalkyl, aryl-C_1-3Alkyl, heteroaryl-C_1-3Alkyl, arylsulfonyl, heteroaryl-C_3-8Nitrogen-containing heterocycloalkyl, benzyl-C_3-8A nitrogen-containing heterocycloalkyl group;

4. A compound according to any one of claims 1 to 3 and stereoisomers thereof, pharmaceutically acceptable salts thereof, wherein

R is C_1-6Alkyl, substituted or unsubstituted piperidinyl, substituted or unsubstituted phenyl, substituted or unsubstituted pyridinyl, substituted or unsubstituted quinolinyl, substituted or unsubstituted phenyl-C_1-3Alkyl, substituted or unsubstituted pyridyl-C_1-3An alkyl group, a carboxyl group,substituted or unsubstituted pyridyl-piperidinyl, substituted or unsubstituted benzyl-piperidinyl, substituted or unsubstituted phenylsulfonyl.

Wherein said alkyl, piperidinyl, phenyl, pyridinyl, quinolinyl, and benzyl are optionally substituted with 1-4 (e.g., 1-3, 1-2, 1, 2, or 3) groups selected from: H. hydroxy, halogen, cyano, amino, nitro, trifluoromethyl, C_1-6Alkyl radical, C_1-6Alkyloxy, and NR₁R₂And R is₁And R₂Independently selected from H, C_1-6An alkyl group.

5. A compound according to any one of claims 1 to 4, and stereoisomers thereof, and pharmaceutically acceptable salts thereof, which is a compound selected from the group consisting of:

6. a process for the preparation of a compound according to any one of claims 1 to 5, comprising the steps of:

reacting the compound of formula 6a with an R group-containing halogenated aromatic compound using a suitable ligand (e.g., Davep) and a catalyst (e.g., palladium acetate) in a base (e.g., sodium t-butoxide) and a suitable solvent (e.g., toluene) at a temperature of 90 ℃ to 150 ℃ (e.g., 100 ℃ to 150 ℃, 110 ℃ to 150 ℃ or 130 ℃ to 150 ℃) for about 8-16 hours to provide the compound of formula b;

the Boc protecting group is removed from the compound of formula b in ethyl acetate hydrochloride solution at a temperature of 10 ℃ to 40 ℃ (e.g., 10 ℃ to 35 ℃, 15 ℃ to 30 ℃,20 ℃ to 30 ℃ or 20 ℃ to 25 ℃) to provide the compound of the invention of formula IA. If racemic 6a is used as the raw material, the compound of the invention shown in the formula I is obtained according to the steps and the method.

Wherein R is as defined in any one of claims 1 to 5.

7. A pharmaceutical composition comprising a therapeutically and/or prophylactically effective amount of a compound of any one of claims 1 to 5, and stereoisomers thereof, pharmaceutically acceptable salts thereof, and optionally one or more pharmaceutically acceptable carriers or excipients.

8. Use of a compound according to any one of claims 1 to 5, a stereoisomer thereof, a pharmaceutically acceptable salt thereof, or a pharmaceutical composition according to claim 7, for the manufacture of a medicament for use in a method for the treatment and/or prevention of diseases or conditions associated with DPP-IV hyperactivity or DPP-IV overexpression.

9. Use according to claim 8, wherein the disease or condition associated with DPP-IV hyperactivity or DPP-IV overexpression is a disease or condition selected from the group consisting of: diabetes, hyperglycemia, non-insulin dependent diabetes, type 2 diabetes.