HK1236935B - Substituted amino six-membered saturated heterocyclic fat used as long-acting dpp-iv inhibitor - Google Patents

Description

Substituted amino six-membered saturated heteroalicyclics as long-acting DPP-IV inhibitors

Technical Field

The present application relates to novel amino six-membered saturated heteroalicyclic derivatives having long-lasting dipeptidyl peptidase-IV (DPP-IV) inhibitory activity, methods for their preparation, pharmaceutical compositions thereof, and their use in treating diseases and conditions that benefit from DPP-IV inhibition.

Background Art

Dipeptidyl peptidase IV (DPP-IV) is a serine protease that rapidly cleaves proteins with proline or alanine residues at the N-terminus of their peptide chains. It is responsible for the metabolic cleavage of certain endogenous peptides in the body, such as GLP-1 and GIP. It has also been shown to have proteolytic activity against a variety of other peptides in vitro, such as GHRH, NPY, GLP-2, and VIP. Due to DPP-IV degradation, GLP-1 and GIP are rapidly inactivated in the body. Therefore, inhibiting DPP-IV activity can significantly prolong the duration of GLP-1 and GIP physiological activity in the body, thereby indirectly regulating insulin secretion and ultimately controlling blood sugar.

As a new type of diabetes treatment, DPP-IV inhibitors have the effect of stimulating insulin secretion in a glucose-dependent manner. They are less likely to cause the side effect of hypoglycemia during the process of controlling blood sugar and also have the characteristics of preserving pancreatic beta cell function. They have fewer gastrointestinal reactions and good tolerance. They can be taken orally without injection and have an efficacy comparable to existing oral hypoglycemic drugs.

Based on the above characteristics, DPP-IV inhibitors can be used to prevent and/or treat diseases and conditions mediated by DPP-IV, such as diabetes and obesity, especially type II diabetes.

Currently, eight DPP-IV inhibitors, including sitagliptin, vildagliptin, saxagliptin, linagliptin, alogliptin, anagliptin, gemigliptin, and teneligliptin, have been successfully launched on the market. Five DPP-IV inhibitors are in Phase II/III clinical research, and more DPP-IV inhibitors are in Phase I clinical and preclinical research stages.

Because diabetes is a chronic disease requiring lifelong medication, the convenience of medication has a direct impact on whether patients adhere to treatment. Therefore, new DPP-IV inhibitors, particularly long-acting DPP-IV inhibitors, are urgently needed.

Summary of the Invention

In one aspect, the present application provides a compound represented by formula I or a pharmaceutically acceptable salt thereof:

in,

Ring A is selected from a 6-membered aryl group or a 5-6-membered heteroaryl group containing 1-2 atoms selected from N, O or S;

X is selected from O or CH ₂ , Y is selected from N or CH, and when X is CH ₂ , Y is not CH;

R ₁ , R ₂ and R ₃ are each independently selected from H, C _1-3 alkyl, -NH ₂ or -OH;

Each R ₄ is independently selected from halogen, -NH ₂ , -OH, C _1-6 alkyl, C _1-6 alkoxy, phenoxy or benzyloxy;

Each R ₅ is independently selected from C _1-6 alkyl, halogen, -CN, -OH, -COOR ₆ , -NHR ₇ or -SO ₂ R ₈ , or two R ₅ groups together with the A ring atom to which they are attached form a 5-7 membered ring;

_R6 is selected from H or _C1-6 alkyl;

R ₇ is selected from H, C _1-6 alkyl or -SO ₂ R ₈ ;

Each R ₈ is independently selected from -OH, -NH ₂ , C _1-6 alkyl or C _3-6 cycloalkyl;

o and p are independently selected from 1, 2 or 3;

m and n are each independently selected from 1 or 2.

On the other hand, the present application provides a pharmaceutical composition containing a compound represented by Formula I or a pharmaceutically acceptable salt, solvate, polymorph, or metabolite thereof as an active ingredient, and one or more pharmaceutically acceptable carriers.

In another aspect, the present application provides the use of a compound of Formula I or a pharmaceutically acceptable salt, solvate, polymorph, metabolite or pharmaceutical composition thereof in the preparation of a medicament for treating diseases and conditions that benefit from DPP-IV inhibition.

In another aspect, the present application provides a method for treating diseases and conditions that benefit from DPP-IV inhibition, comprising administering to an individual in need thereof a compound of Formula I or a pharmaceutically acceptable salt, solvate, polymorph, metabolite, or pharmaceutical composition thereof.

In another aspect, the present application provides a compound of formula I or a pharmaceutically acceptable salt, solvate, polymorph, metabolite or pharmaceutical composition thereof for treating diseases and conditions that benefit from DPP-IV inhibition.

BRIEF DESCRIPTION OF THE DRAWINGS

Figure 1 shows the results of an in vitro study of the compound of Example 4 inhibiting DPP-IV activity in rat plasma. At an oral dose of 5 mg/kg body weight, the compound of Example 4 still achieved nearly 50% inhibition of DPP-IV activity in rat plasma after 120 hours, meeting the requirement for a long-acting inhibitor.

FIG2 shows the inhibitory effects of the compound of Example 4 and Omarigliptin at different doses on serum DPP-IV activity in ob/ob mice after single administration.

Detailed Description of the Invention

In the following description, certain specific details are included to provide a thorough understanding of the various disclosed embodiments. However, one skilled in the relevant art will recognize that the embodiments can be implemented without one or more of these specific details and with other methods, components, materials, etc.

Unless otherwise required in this application, throughout the specification and the claims that follow, the word "comprise" and its English variations such as "comprises" and "comprising" should be interpreted as having an open, inclusive meaning, that is, "including but not limited to".

Reference throughout this specification to "one embodiment" or "an embodiment" or "in another embodiment" or "in certain embodiments" means that at least one embodiment includes the specific referenced elements, structures, or features described in connection with that embodiment. Thus, appearances of the phrases "in one embodiment" or "in an embodiment" or "in another embodiment" or "in certain embodiments" in various places throughout this specification are not necessarily all referring to the same embodiment. Furthermore, the specific elements, structures, or features may be combined in any suitable manner in one or more embodiments.

It should be understood that as used in this specification and the appended claims, the singular article "a," "an," and "the" includes plural referents unless the context clearly dictates otherwise. Thus, for example, a reference to a reaction including "a catalyst" includes one catalyst, or two or more catalysts. It should also be understood that the term "or" is generally used in its sense including "and/or" unless the context clearly dictates otherwise.

Chemical terms and definitions

As used herein, the term "compound" includes all stereoisomeric, geometric isomeric, tautomeric, and isotopic forms of the compound.

The compounds described herein may be asymmetric, for example, having one or more stereoisomers. Unless otherwise indicated, all stereoisomers, such as enantiomers and diastereomers, are included within the scope of this application. The compounds containing asymmetric carbon atoms herein can be isolated in optically pure forms or racemic forms. Optically pure forms can be isolated from racemic mixtures by resolution or synthesized using chiral starting materials or chiral reagents.

The compounds of the present application also include tautomeric forms, which are derived from the exchange of a single bond with an adjacent double bond accompanied by the migration of a proton.

This application also includes all isotopes of atoms, whether in intermediates or final compounds. Isotopes of atoms include atoms with the same atomic number but different mass numbers. For example, isotopes of hydrogen include tritium and deuterium.

The term "halogen" refers to fluorine, chlorine, bromine or iodine.

The term "hydroxy" refers to -OH.

The term "carboxyl" refers to -COOH.

The term "cyano" refers to -CN.

As used herein, the term "sulfonyl" refers to _-SO2 -alkyl, _-SO2 -cycloalkyl, and _-SO2 -aryl.

The term "amino" as used herein refers to _-NH2 , -NH(alkyl) and -N(alkyl) ₂ . Specific examples of amino include, but are not limited to, _-NH2 , _-NHCH3 , -NHCH( _CH3 ) ₂ , _-N ( _{CH3 )2 ,} -NHC2H5, -N( _CH3 ) _C2H5 and _the like.

As used herein, the term "alkyl" refers to a straight-chain or branched saturated aliphatic hydrocarbon group consisting solely of carbon and hydrogen atoms, such as methyl, ethyl, propyl, butyl, pentyl, hexyl, heptyl, octyl, nonyl, decyl, and the _like . The specific alkyl group includes all isomeric forms thereof, for example, propyl includes _-CH2CH2CH3 and -CH( _CH3 ) _{2 ,} and butyl includes _{-CH2CH2CH2CH3 , -CH} (CH3 ₎ ( _CH2CH3 ), -C( _{CH3 ) 3} , and _-CH2CH ( _CH3 ) ₂ . _The term " _C1-6alkyl " refers to an alkyl group having 1 to 6 carbon atoms. The term " _C1-4alkyl " refers to an alkyl group having 1 to 4 carbon atoms. The term " _C1-3alkyl " refers to an alkyl group having 1 to 3 carbon atoms. The "alkyl", "C _1-8 alkyl", "C _1-6 alkyl" or "C _1-3 alkyl" may be unsubstituted or substituted by one or more substituents independently selected from hydroxy, halogen or amino.

The term "cycloalkyl" as used herein refers to a fully cyclic, saturated hydrocarbon group consisting solely of carbon and hydrogen atoms, such as a _C3-20 cycloalkyl group, preferably a _C3-6 cycloalkyl group, such as cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, etc. The cycloalkyl group may be unsubstituted or independently substituted with one or more substituents, including but not limited to alkyl, alkoxy, cyano, carboxyl, aryl, heteroaryl, amino, halogen, sulfonyl, sulfinyl, phosphoryl, and hydroxyl.

The term "aryl" as used herein refers to an all-carbon monocyclic or fused ring having a completely conjugated π electron system, having 6-14 carbon atoms, preferably 6-12 carbon atoms, and most preferably 6 carbon atoms. Aryl can be unsubstituted or independently substituted with one or more substituents, examples of which include but are not limited to alkyl, alkoxy, aryl, aralkyl, amino, halogen, hydroxyl, sulfonyl, sulfinyl, phosphoryl, and heteroalicyclic groups. Non-limiting examples of aryl include but are not limited to phenyl, naphthyl, and anthracenyl.

The term "arylalkyl" as used herein refers to an alkyl group substituted by an aryl group as defined above, preferably a C _1-6 alkyl group substituted by an aryl group. Non-limiting examples of arylalkyl groups include, but are not limited to, -CH ₂ -phenyl, -(CH ₂ ) ₂ -phenyl, -(CH ₂ ) ₃ -phenyl, -CH(CH ₃ )-phenyl, -CH ₂ -CH(CH ₃ )-phenyl, -(CH ₂ ) ₄ -phenyl, -CH ₂ -CH(CH ₃ )-CH ₂ -phenyl, -CH ₂ -CH ₂ -CH(XH ₃ )-phenyl, and the like.

As used herein, the term "heteroaryl" refers to a monocyclic or fused ring of 5-12 ring atoms, having 5, 6, 7, 8, 9, 10, 11, or 12 ring atoms, of which 1, 2, 3, or 4 are independently selected from N, O, or S, with the remaining ring atoms being C, and having a completely conjugated π-electron system. A heteroaryl group may be unsubstituted or independently substituted with one or more substituents including, but not limited to, alkyl, alkoxy, aryl, aralkyl, amino, halogen, hydroxy, cyano, nitro, carbonyl, and heteroalicyclic groups. Non-limiting examples of heteroaryl groups include, but are not limited to, pyrrolyl, furanyl, thienyl, imidazolyl, oxazolyl, pyrazolyl, pyridinyl, pyrimidinyl, pyrazinyl, quinolyl, isoquinolyl, tetrazolyl, and triazinyl.

The term "heteroarylalkyl" as used herein refers to an alkyl group substituted by a heteroaryl group as defined above, preferably a C _1-6 alkyl group substituted by a heteroaryl group. Non-limiting examples of heteroarylalkyl groups include, but are not limited to, -CH ₂ -pyrazolyl, -(CH ₂ ) ₂ -pyridinyl, -(CH ₂ ) ₃ -thienyl, -CH(CH ₃ )-pyrazinyl, -CH ₂ -CH(CH ₃ )-furyl, and the like.

As used herein, the term "heteroalicyclic ring" refers to a monocyclic or fused ring having 3-12 ring atoms, having 3, 4, 5, 6, 7, 8, 9, 10, 11 or 12 ring atoms, wherein 1 or 2 ring atoms are heteroatoms independently selected from N, O, S(O) _n (wherein n is 0, 1 or 2), and the remaining ring atoms are C. Such rings may be saturated or unsaturated (e.g., having one or more double bonds), but do not have a completely conjugated π-electron system. Examples of 3-membered saturated heteroalicyclic rings include, but are not limited to, examples of 4-membered saturated heteroalicyclic rings include, but are not limited to, examples of 5-membered saturated heteroalicyclic rings include, but are not limited to, examples of 6-membered saturated heteroalicyclic rings include, but are not limited to, examples of 7-membered saturated heteroalicyclic rings include, but are not limited to, examples of 5-membered unsaturated heteroalicyclic rings include, but are not limited to, examples of 6-membered unsaturated heteroalicyclic rings include, but are not limited to.

The term "heteroalicyclic group" as used herein refers to the group remaining after removing one hydrogen atom from a "heteroalicyclic" molecule. The heteroalicyclic group may be unsubstituted or the hydrogen atoms therein may be independently substituted by one or more substituents, the substituents including but not limited to alkyl, alkoxy, =O, aryl, aralkyl, -COOH, -CN, amino, halogen or hydroxyl.

As used herein, the term "pharmaceutically acceptable salt" refers to salts that retain the biological effects and properties of the DPP-IV inhibitors of the present application and are not biologically or otherwise unacceptable. For example, pharmaceutically acceptable salts do not interfere with the beneficial effects of the agents of the present application in inhibiting DPP-IV, and include "pharmaceutically acceptable acid addition salts" and "pharmaceutically acceptable base addition salts."

As used herein, the term "pharmaceutically acceptable acid addition salts" refers to those salts which retain the biological effectiveness and properties of the free bases and which are biologically or otherwise suitable and which are formed using inorganic or organic acids.

As used herein, the term "pharmaceutically acceptable base addition salts" refers to salts which retain the biological effectiveness and properties of the free acids, which base addition salts are biologically or otherwise suitable. These salts are prepared by adding inorganic or organic bases to the free acids.

The term "pharmaceutical composition" as used herein refers to a preparation comprising one or more compounds of the present application or their salts and a carrier generally accepted in the art for delivering the biologically active compound to an organism (e.g., a human). The purpose of a pharmaceutical composition is to facilitate administration of the compound of the present application to an organism.

The term "pharmaceutically acceptable carrier" as used herein refers to carriers that have no significant irritating effect on organisms (e.g., humans) and do not impair the biological activity and properties of the active compound. "Pharmaceutically acceptable carrier" may also refer to an inert substance that is administered together with the active ingredient and facilitates the administration of the active ingredient, including but not limited to any glidant, sweetener, diluent, preservative, dye/colorant, flavor enhancer, surfactant, wetting agent, dispersant, disintegrant, suspending agent, stabilizer, isotonic agent, solvent or emulsifier approved by the U.S. Food and Drug Administration as acceptable for use in humans or animals (e.g., livestock). Non-limiting examples of such carriers include calcium carbonate, calcium phosphate, various sugars and various types of starch, cellulose derivatives, gelatin, vegetable oils and polyethylene glycol.

Compounds of formula I

In one aspect, the present application provides a compound represented by formula I or a pharmaceutically acceptable salt thereof:

in,

Ring A is selected from a 6-membered aryl group or a 5-6-membered heteroaryl group containing 1-2 atoms selected from N, O or S;

X is selected from O or CH ₂ , Y is selected from N or CH, and when X is CH ₂ , Y is not CH;

R ₁ , R ₂ and R ₃ are each independently selected from H, C _1-3 alkyl, -NH ₂ or -OH;

Each R ₄ is independently selected from halogen, -NH ₂ , -OH, C _1-6 alkyl, C _1-6 alkoxy, phenoxy or benzyloxy;

Each R ₅ is independently selected from C _1-6 alkyl, halogen, -CN, -OH, -COOR ₆ , -NHR ₇ or -SO ₂ R ₈ , or two R ₅ groups together with the A ring atom to which they are attached form a 5-7 membered ring;

_R6 is selected from H or _C1-6 alkyl;

R ₇ is selected from H, C _1-6 alkyl or -SO ₂ R ₈ ;

Each R ₈ is independently selected from -OH, -NH ₂ , C _1-6 alkyl or C _3-6 cycloalkyl;

o and p are independently selected from 1, 2 or 3;

m and n are each independently selected from 1 or 2.

On the other hand, the present application provides a compound represented by Formula II or a pharmaceutically acceptable salt thereof:

in,

X is selected from O;

Y is selected from N or CH;

_R1 is selected from _-NH2 or -OH;

_R2 and _R3 are both H;

R _4a and R _4b are each independently selected from F, Cl, Br, I, -NH ₂ or -OH;

Each R ₅ is independently selected from F, Cl, Br, I, -CN, -COOR ₆ , -NHR ₇ or -SO ₂ R ₈ ;

p is selected from 1 or 2;

_R6 is selected from H, methyl, ethyl, propyl or butyl;

R ₇ is selected from H, methyl, ethyl, propyl, butyl or -SO ₂ R ₈ ;

Each R ₈ is independently selected from -OH, -NH ₂ , methyl, ethyl, propyl, butyl, C ₃ cycloalkyl, C ₄ cycloalkyl, or C ₅ cycloalkyl.

On the other hand, the present application provides a compound represented by formula III or a pharmaceutically acceptable salt thereof:

in,

X is selected from O;

Y is selected from N or CH;

_R1 is selected from _-NH2 or -OH;

_R2 and _R3 are both H;

R _4a and R _4b are each independently selected from F, Cl, Br, -NH ₂ or -OH;

R _5a and R _5b are each independently selected from F, Cl, Br, I, -CN, -COOR ₆ , -NHR ₇ or -SO ₂ R ₈ ;

_R6 is selected from H, methyl, ethyl, propyl or butyl;

R ₇ is selected from H, methyl, ethyl, propyl, butyl or -SO ₂ R ₈ ;

Each R ₈ is independently selected from -OH, -NH ₂ , methyl, ethyl, propyl, butyl, C ₃ cycloalkyl, C ₄ cycloalkyl, or C ₅ cycloalkyl.

On the other hand, the present application provides a compound represented by formula IV or a pharmaceutically acceptable salt thereof:

in,

X is selected from O;

Y is selected from N or CH;

_R1 is selected from _-NH2 or -OH;

_R2 and _R3 are H;

R _4a and R _4b are each independently selected from F, Cl, Br, -NH ₂ , -OH;

R _5c and R _5d together with the pyrazole ring atoms to which they are attached form a 5-, 6- or 7-membered non-aromatic ring, said 5-, 6- or 7-membered non-aromatic ring preferably containing 1, 2 or 3 heteroatoms selected from N, O or S, and more preferably containing a -SO ₂ - group.

On the other hand, the present application provides a compound represented by Formula V or a pharmaceutically acceptable salt thereof:

in,

X is selected from O;

Y is selected from N or CH;

R ₁ , R ₂ and R ₃ are each independently selected from H, C _1-3 alkyl, -NH ₂ or -OH;

Each R ₄ is independently selected from halogen, -NH ₂ , -OH, C _1-6 alkyl, C _1-6 alkoxy, phenoxy or benzyloxy;

Each R ₅ is independently selected from C _1-6 alkyl, halogen, -CN, -OH, -COOR ₆ , -NHR ₇ or -SO ₂ R ₈ ;

_R6 is selected from H or _C1-6 alkyl;

R ₇ is selected from H, C _1-6 alkyl or -SO ₂ R ₈ ;

Each R ₈ is independently selected from -OH, -NH ₂ , C _1-6 alkyl or C _3-6 cycloalkyl;

o and p are independently selected from 1, 2 or 3.

In one embodiment, in the compound represented by formula V, R ₁ is preferably -NH ₂ or -OH.

In another embodiment, in the compound of formula V, R ₂ and R ₃ are preferably H.

In another embodiment, in the compound represented by formula V, R ₄ is preferably F, Cl, Br, I, -NH ₂ or -OH.

In another embodiment, in the compound represented by formula V, R ₅ is preferably F, Cl, Br, I, -COOR ₆ , -NHR ₇ or -SO ₂ R ₈ .

In another embodiment, in the compound represented by formula V, _R6 is preferably H, methyl, ethyl, propyl or butyl.

In another embodiment, in the compound represented by formula V, R ₇ is preferably H, methyl, ethyl, propyl, butyl or -SO ₂ R ₈ .

In another embodiment, in the compound represented by formula V, R ₈ is preferably -OH, -NH ₂ , methyl, ethyl, propyl, butyl, C ₃ cycloalkyl, C ₄ cycloalkyl or C ₅ cycloalkyl.

In another embodiment, in the compound represented by formula V, o is preferably 2.

In another embodiment, in the compound represented by formula V, p is preferably 1 or 2.

In another embodiment, in the compound represented by formula V, the substitution position of R ₄ is preferably the position of R _4a and R _4b in the structure represented by formula VI below:

wherein R _4a and R _4b are each independently selected from F, Cl, Br, I, -NH ₂ or -OH, and the other groups are the same as defined in the aforementioned formula V.

In another embodiment, in the compound represented by formula V, p is preferably 2, and the substitution position of R ₅ is preferably the position of R _5a and R _5b in the structure represented by formula VII below:

Wherein, R _5a and R _5b are each independently selected from F, Cl, Br, I, -COOR ₆ , -NHR ₇ or -SO ₂ R ₈ , and other substituents are the same as defined in the aforementioned formula V.

In another embodiment, in the compound represented by formula V, p is preferably 1, and the substitution position of R ₅ is preferably the position of R _5a in the structure represented by formula VIII below:

Wherein, R _5a is selected from F, Cl, Br, I, -COOR ₆ , -NHR ₇ or -SO ₂ R ₈ , and other substituents are the same as defined in the aforementioned formula V.

The present application preferably comprises the following compounds or pharmaceutically acceptable salts thereof;

Preparation method of compound of general formula I

In another aspect, the present application also provides a method for preparing the compound represented by Formula 1, which includes the following synthesis scheme:

Synthesis Scheme 1

Compound 1-6 can be synthesized using Synthesis Scheme 1. Halogenated phenol 1-7 reacts with an alkyl halide, benzyl halide, or aryl halide (metal catalysis required) in the presence of a base to obtain a halogenated aryl ether intermediate 1-1. Intermediate 1-1 is metallated with a Grignard reagent and reacted with a Weinreb amide to obtain a ketone intermediate 1-2. Intermediate 1-2 is selectively reduced with a chiral metal catalyst to obtain compound 1-3. Intermediate 1-3 is cyclized with a metal catalyst to obtain compound 1-4. The double bond of intermediate 1-4 is sequentially hydroborated and oxidized to obtain alcohol compound 1-5. The alcoholic hydroxyl group of intermediate 1-5 is catalytically oxidized to obtain compound 1-6.

Synthesis Scheme 2

Compound 2-4 can be synthesized using Synthesis Scheme 2 (q is selected from 0, 1, or 2). The reaction of intermediate 2-5 with N-iodosuccinimide yields iodine-substituted intermediate 2-1, which is then reacted with chloroalkylsulfonyl chloride to yield intermediate 2-2. Intermediate 2-2 is first metallated with zinc powder and then subjected to metal-catalyzed ring closure to yield intermediate 2-3. Intermediate 2-3 can be deprotected by acid to yield compound 2-4.

Synthesis Scheme 3

Compound 3-4 can be synthesized using Synthesis Scheme 3. 4-Bromophthalimide 3-5 is reduced with borane to give intermediate 3-1, which is protected with Boc anhydride and a base to give compound 3-2. Intermediate 3-2 reacts with sodium alkylsulfinate in the presence of cuprous ions and proline to give sulfone intermediate 3-3, which can be deprotected with acid to give compound 3-4.

Synthesis Scheme 4

Compound 4-2 can be synthesized according to Synthesis Scheme 4. 5-Amino-1-oxoisoindoline 4-3 reacts with alkylsulfonyl chloride to obtain intermediate 4-1, which is then reduced with borane to obtain intermediate 4-2.

Synthesis Scheme 5

Compound 5-4 can be synthesized according to Synthesis Scheme 5. 5-Amino-1-oxoisoindoline 4-3 reacts with N-chlorosuccinimide to obtain intermediate 5-1. Intermediate 5-1 is first diazotized and then reacted with potassium iodide to obtain intermediate 5-2. Intermediate 5-2 is then reacted with sodium alkylsulfinate in the presence of cuprous ions and proline to obtain sulfone intermediate 5-3. Intermediate 5-3 is reduced with borane to obtain compound 5-4.

Synthetic Fangcha 6

Compound 6-3 can be synthesized according to Synthesis Scheme 6. 5-Amino-1-oxoisoindoline 4-3 is first diazotized and then reacted with sulfur dioxide and cuprous chloride to obtain sulfonyl chloride intermediate 6-1. Intermediate 6-1 reacts with amine to obtain intermediate 6-2, which is then reduced with borane to obtain compound 6-3.

Synthesis Scheme 7

Compound 7-3 can be synthesized according to Synthesis Scheme 7. Ketone 7-4 and amine 7-5 undergo reductive amination to give intermediate 7-1, which is then reacted with alkylsulfonyl chloride to give intermediate 7-2. Intermediate 7-2 is deprotected under acidic conditions to give the final compound 7-3.

Synthesis Scheme 8

Compound 8-2 can be synthesized according to Synthesis Scheme 8. Ketone 8-3 and amine 8-4 undergo reductive amination to give intermediate 8-1, and intermediate 8-1 is deprotected under acidic conditions to give final compound 8-2.

The above synthesis scheme is only an illustrative example of the preparation method of some compounds in this application. Ordinary technicians in this field can also synthesize the compounds in this application using similar methods based on the above synthesis scheme.

Pharmaceutical composition

The compound of the present application or its salt can be administered alone as an active substance, but is preferably administered in the form of a pharmaceutical composition.

On the other hand, the present application provides a pharmaceutical composition containing a compound represented by Formula I or a pharmaceutically acceptable salt, solvate, polymorph, or metabolite thereof as an active ingredient, and one or more pharmaceutically acceptable carriers.

The administration of the present application compound or its pharmaceutically acceptable salt can be carried out in pure form or in the form of a suitable pharmaceutical composition by providing any acceptable mode of administration of a medicine of similar use. The pharmaceutical composition of the present application can be prepared by combining the present application compound with a suitable pharmaceutically acceptable carrier, diluent, medium or excipient. The pharmaceutical composition of the present application can be formulated into solid, semi-solid, liquid or gaseous preparations, such as tablets, pills, capsules, powders, granules, ointments, emulsions, suspensions, solutions, suppositories, injections, inhalants, gels, microspheres and aerosols, etc.

Typical routes of administration of the compounds of the present application, or pharmaceutically acceptable salts thereof, or pharmaceutical compositions thereof include, but are not limited to, oral, rectal, transmucosal, enteral administration, or topical, transdermal, inhalation, parenteral, sublingual, vaginal, intranasal, intraocular, intraperitoneal, intramuscular, subcutaneous, intravenous administration, etc. The preferred route of administration is oral administration.

The pharmaceutical composition of the present application can be manufactured by methods known to those skilled in the art, such as conventional mixing methods, dissolution methods, granulation methods, sugar-coated pill making methods, grinding methods, emulsification methods, freeze-drying methods, etc.

In a preferred embodiment, the pharmaceutical composition is in oral form. For oral administration, the pharmaceutical composition can be formulated by mixing the active compound with a pharmaceutically acceptable carrier well known in the art. These carriers enable the compounds of the present invention to be formulated into tablets, pills, lozenges, dragees, capsules, liquids, gels, slurries, suspensions, and the like for oral administration to a patient.

Solid oral pharmaceutical compositions can be prepared by conventional mixing, filling, or tableting methods. For example, they can be obtained by mixing the active compound with a solid excipient, optionally grinding the resulting mixture, adding other suitable excipients as needed, and then processing the mixture into granules to obtain a tablet or dragee core. Suitable excipients include, but are not limited to, binders, diluents, disintegrants, lubricants, glidants, sweeteners, or flavoring agents. Examples include microcrystalline cellulose, glucose solution, acacia mucilage, gelatin solution, sucrose, and starch paste; talc, starch, magnesium stearate, calcium stearate, or stearic acid; lactose, sucrose, starch, mannitol, sorbitol, or dicalcium phosphate; silicon dioxide; cross-linked sodium carboxymethylcellulose, pregelatinized starch, sodium starch glycolate, alginic acid, corn starch, potato starch, methylcellulose, agar, carboxymethylcellulose, cross-linked polyvinyl pyrrolidone, and the like. The dragee core can optionally be coated according to methods known in conventional pharmaceutical practice, particularly with an enteric coating.

The pharmaceutical composition of the present application may also be suitable for parenteral administration, such as sterile solutions, suspensions or lyophilized products in suitable unit dosage forms. Suitable excipients, such as fillers, buffers or surfactants, can be used.

Therapeutic uses

In one aspect, the present application provides the use of a compound of Formula I or a pharmaceutically acceptable salt, solvate, polymorph, metabolite or pharmaceutical composition thereof in the preparation of a medicament for treating or preventing diseases and conditions that benefit from DPP-IV inhibition.

In another aspect, the present application provides a method for treating diseases and conditions that benefit from DPP-IV inhibition, comprising administering to an individual in need thereof a compound of Formula I or a pharmaceutically acceptable salt, solvate, polymorph, metabolite, or pharmaceutical composition thereof.

In another aspect, the present application provides a compound of formula I or a pharmaceutically acceptable salt, solvate, polymorph, metabolite or pharmaceutical composition thereof for treating diseases and conditions that benefit from DPP-IV inhibition.

The disease or condition that benefits from DPP-IV inhibition is selected from insulin resistance, hyperglycemia, type II diabetes, diabetic dyslipidemia, impaired glucose tolerance (IGT), low fasting plasma glucose (IFG), metabolic acidosis, ketosis, appetite regulation, obesity, various cancers, nervous system disorders or immune system disorders, etc., preferably including type II diabetes or obesity.

The substituted amino six-membered heteroalicyclic compounds provided in the present application have very good DPP-IV inhibitory activity, which is equivalent to or better than that of Omarigliptin, and have very good in vivo metabolism levels and very long in vivo half-life, and are long-acting DPP-IV inhibitor drugs.

Example

The following specific examples are provided for the purpose of enabling those skilled in the art to more clearly understand and implement the present invention. They should not be considered as limiting the scope of the present invention, but are merely illustrative and representative of the present invention. Those skilled in the art will appreciate that there are other synthetic pathways for forming the compounds of the present application, and the following are non-limiting examples.

All operations involving raw materials that are easily oxidized or hydrolyzed were performed under nitrogen protection. Unless otherwise specified, the raw materials used in this application were purchased directly from the market and used directly without further purification.

Column chromatography was performed using silica gel (200-300 mesh) manufactured by Qingdao Chemical Co., Ltd. Thin-layer chromatography was performed using precast plates (silica gel 60PF ₂₅₄ , 0.25 mm) manufactured by E. Merck. Chiral compound separation and enantiomeric excess (ee) determination were performed using an Agilent LC 1200 series (column: CHIRALPAK AD-H, 5 μm, 30°C). Nuclear magnetic resonance (NMR) chromatography was performed using a Varian VNMRS-400 NMR spectrometer; liquid chromatography-mass spectrometry (LC/MS) was performed using a FINNIGAN Thermo LCQ Advantage MAX, an Agilent LC 1200 series (column: Waters Symmetry C18, 5 μm, 35°C) in ESI (+) ion mode.

Intermediate 1: tert-Butyl (2R,3S)-2-(2,5-difluorophenyl)-5-oxotetrahydro-2H-pyran-3-ylcarbamate

Step 1: Methyl 2-(benzhydrylimino)acetate

Glycine methyl ester hydrochloride (39.4 g, 0.314 mol) was dissolved in 300 mL of dichloromethane. Benzophenone imine (50.0 g, 0.276 mol) was added all at once with stirring. The reaction mixture was stirred at room temperature for 1 day. The resulting solid was removed by filtration, and the filtrate was washed with water, sodium carbonate solution, and saturated brine, respectively. The oily product, methyl 2-(benzhydrylimino)acetate (64.2 g), solidified upon cooling and was used directly in the next step. Yield: 92%. ¹ H-NMR (400 MHz, CDCl ₃ ): δ = 7.66 (2H, m), 7.45 (4H, m), 7.35 (2H, m), 7.17 (2H, m), 4.22 (2H, s), 3.74 (3H, s).

Step 2: Methyl 2-(benzhydrylimino)pent-4-ynoate

To a stirred solution of methyl 2-(benzhydrylimino)acetate (64.2 g, 0.254 mol), propargyl bromide (27.8 mL, 0.322 mol), and tetra-n-butylamine bromide (8.66 g, 26.9 mmol) in methyl tert-butyl ether (600 mL) was added cesium carbonate (175.4 g, 0.538 mol). The mixture was stirred at 50°C for 2 days. The solid was filtered off, the filter cake was washed with a small amount of methyl tert-butyl ether (MTBE), and the filtrate was concentrated to 300 mL and used directly in the next step. ¹ H-NMR (400MHz, CDCl ₃ ): δ = 7.66 (2H, m), 7.45 (4H, m), 7.35 (2H, m), 7.17 (2H, m), 4.32 (1H, m), 3.73 (3H, s), 2.83 (2H, m), 1.95 (1H, s).

Step 3: 2-(tert-Butoxycarbonylamino)pent-4-ynoic acid

To the concentrated solution obtained in the previous step, 280 mL of 1N hydrochloric acid was added and the mixture was stirred at room temperature until TLC indicated the disappearance of methyl 2-(benzhydrylimino)pent-4-ynoate, which took approximately 12 hours. The organic phase was separated and the aqueous phase was extracted with methyl tert-butyl ether and the organic phase was discarded. 50% sodium hydroxide solution (18.75 N, 0.712 mol) was added to the aqueous phase and stirred for 2 hours. 50 mL of water was added, followed by a solution of Boc anhydride (61.0 g, 0.28 mol) in methyl tert-butyl ether (200 mL). The mixture was stirred at room temperature for 6 hours and then acidified with 10% hydrochloric acid to a pH of 3 while cooling in an ice bath. The organic phase was separated and the aqueous phase was extracted with methyl tert-butyl ether. The combined organic phases were dried and concentrated to give the product, 2-(tert-butoxycarbonylamino)pent-4-ynoic acid (38.8 g), with a two-step yield of 72%. ¹ H-NMR (400MHz, CDCl ₃ ): δ=7.65 (1H, brs), 5.36 (1H, d, J=8.0Hz), 4.52 (1H, m), 2.77 (2H, m), 2.08 (1H, s), 1.46 (9H, s).

Step 4: 2-(tert-Butoxycarbonylamino)pent-4-ynoyl-(N-methoxy-N-methyl)amine

2-(tert-Butoxycarbonylamino)pent-4-ynoic acid (20.22 g, 94.9 mmol) was dissolved in acetonitrile (200 mL), and 2-(7-azobenzotriazole)-N,N,N′,N′-tetramethyluronium hexafluorophosphate (HATU) (43.27 g, 113.9 mmol), N,O-dimethylhydroxylamine hydrochloride (11.11 g, 113.9 mmol) and triethylamine (46.2 mL, 332.2 mmol) were added respectively, and the mixture was stirred at room temperature for 2 hours. The reaction solution was poured into 1500 mL of water and extracted three times with ethyl acetate. The organic phases were combined and washed sequentially with 1N hydrochloric acid, water, saturated sodium bicarbonate solution, and saturated brine. The product was dried, concentrated, and purified by silica gel column chromatography (petroleum ether/ethyl acetate, 5:1-4:1) to obtain 2-(tert-butoxycarbonylamino)pent-4-ynyl-(N-methoxy-N-methyl)amine (19.6 g) in a yield of 81%. ¹ H-NMR (400 MHz, CDCl ₃ ): δ = 5.45 (1H, d, J = 8.0 Hz), 4.82 (1H, m), 3.77 (3H, s), 3.24 (3H, s), 2.66 (2H, m), 2.04 (1H, s), 1.45 (9H, s).

Step 5: tert-Butyl 1-(2,5-difluorophenyl)-1-oxopent-4-yn-2-ylcarbamate

2,5-Difluorobromobenzene (11.58 g, 60 mmol) was dissolved in 25 mL of toluene and cooled to -10°C to -5°C. Lithium bromide (2.61 g, 30 mmol) was added, and a solution of isopropylmagnesium fluoride in tetrahydrofuran (2M, 33 mL, 66 mmol) was added dropwise over 1.5 hours. The mixture was stirred at low temperature for 1 hour. 2-(tert-Butoxycarbonylamino)pent-4-ynoyl-(N-methoxy-N-methyl)amine (7.68 g, 30 mmol) was dissolved in 35 mL of tetrahydrofuran and added dropwise over 1 hour. The temperature was slowly warmed to room temperature, and then stirred at room temperature for 1 hour. 22 mL of 3N hydrochloric acid was added dropwise to the reaction solution to quench the reaction. The organic phase was washed sequentially with water, saturated sodium bicarbonate solution, and saturated brine, dried, concentrated, and then purified by silica gel column chromatography (petroleum ether/ethyl acetate, 10:1) to give tert-butyl 1-(2,5-difluorophenyl)-1-oxopent-4-yn-2-ylcarbamate (6.07 g) in a yield of 65%. ¹ H-NMR (400 MHz, CDCl ₃ ): δ = 7.56 (1H, m), 7.26 (1H, m), 7.15 (1H, m), 5.68 (1H, d, J = 7.6 Hz), 5.24 (1H, m), 2.91 (1H, m), 2.68 (1H, m), 1.99 (1H, s), 1.45 (9H, s).

Step 6: tert-Butyl (1R,2S)-1-(2,5-difluorophenyl)-1-hydroxypent-4-yn-2-ylcarbamate

Tert-butyl 1-(2,5-difluorophenyl)-1-oxopentan-4-yn-2-ylcarbamate (6.07 g, 19.7 mmol) and 1,4-diazabicyclo[2.2.2]octane (DABCO) (6.61 g, 59 mmol) were dissolved in 70 mL of tetrahydrofuran and purged with nitrogen for 30 minutes. (p-cymene)((1R,2R)-N-p-toluenesulfonyl-1,2-diphenylethylenediamine)ruthenium chloride (1) (63 mg, 0.1 mmol) was added, and the mixture was purged with nitrogen/evacuated three times. Formic acid (4.53 g, 98.5 mmol) was added dropwise to the solution under a nitrogen atmosphere, and the mixture was stirred at 40°C for 2 days. Add 200 mL of dichloromethane, wash with 5% citric acid solution, saturated sodium bicarbonate solution and saturated brine in sequence, dry and concentrate, and then chromatograph on a silica gel column (petroleum ether/ethyl acetate, 25:1-8:1) to give the product (1R,2S)-1-(2,5-difluorophenyl)-1-hydroxypent-4-yn-2-ylcarbamic acid tert-butyl ester (6.11 g) in a yield of 100%.

Step 7: tert-Butyl (2R,3S)-2-(2,5-difluorophenyl)-3,4-dihydro-2H-pyran-3-ylcarbamate

(1R, 2S)-1-(2,5-difluorophenyl)-1-hydroxypent-4-yn-2-ylcarbamic acid tert-butyl ester (6.11g, 19.6mmol) is dissolved among the 60mL DMF, passes through nitrogen 30 minutes.Add tris(tris(3-fluorophenyl)phosphine)rhodium chloride catalyst (427mg, 0.393mmol), pass through nitrogen/vacuum 3 times, under nitrogen atmosphere, stir 16 hours at 80 ℃.Add water and saturated sodium bicarbonate solution each 150mL after cooling, extract with toluene, merge organic phase, wash several times with water, dry and concentrate the back and use silica gel column chromatography (petroleum ether/ethyl acetate, 30: 1-25: 1) to obtain product (2R, 3S)-2-(2,5-difluorophenyl)-3,4-dihydro-2H-pyrans-3-ylcarbamic acid tert-butyl ester (4.89g), yield 80%.

Step 8: tert-Butyl (2R,3S)-2-(2,5-difluorophenyl)-5-hydroxytetrahydro-2H-pyran-3-ylcarbamate

Tert-butyl (2R,3S)-2-(2,5-difluorophenyl)-3,4-dihydro-2H-pyran-3-ylcarbamate (2.81 g, 9.04 mmol) was dissolved in 50 mL of tetrahydrofuran and cooled to -10°C. Borane-dimethyl sulfide complex (2.26 mL, 22.6 mmol) was added dropwise. The mixture was stirred at low temperature for 2 hours and then warmed to 15°C. 1N sodium hydroxide solution (27.1 mL, 27.1 mmol) was added dropwise to the solution, followed by sodium perborate (4.17 g, 27.1 mmol) and stirring overnight. 100 mL of water was added, the organic phase was separated, and the aqueous phase was extracted with dichloromethane. The organic phases were combined, washed with water, dried, concentrated, and then purified by silica gel column chromatography (dichloromethane/methanol, 30:1-12:1) to give the product (2R,3S)-2-(2,5-difluorophenyl)-5-hydroxytetrahydro-2H-pyran-3-ylcarbamic acid tert-butyl ester (2.33 g) as a white solid in a yield of 78%.

Step 9: tert-Butyl (2R,3S)-2-(2,5-difluorophenyl)-5-oxotetrahydro-2H-pyran-3-ylcarbamate

Tert-butyl (2R,3S)-2-(2,5-difluorophenyl)-5-hydroxytetrahydro-2H-pyran-3-ylcarbamate (2.33 g, 7.08 mmol) was dissolved in a mixture of 24 mL of acetonitrile, 4 mL of water, and 4 mL of acetic acid. A 4 mL aqueous solution of hydrated ruthenium chloride (3.7 mg, 0.0142 mmol) was added. The mixture was cooled to 0°C and sodium bromate (535 mg, 3.54 mmol) was added. The mixture was stirred at low temperature for approximately 1.5 hours until the reaction of the raw materials was complete. 120 mL of water was added to the solution, and the mixture was stirred at 0°C overnight. The mixture was then extracted with dichloromethane. The organic phase was washed with water, dried, concentrated, and then purified by silica gel column chromatography (petroleum ether/ethyl acetate, 10:1) to afford the intermediate 1-tert-butyl (2R,3S)-2-(2,5-difluorophenyl)-5-oxotetrahydro-2H-pyran-3-ylcarbamate (1.71 g) as a white solid in a yield of 74%. ¹ H-NMR (400 MHz, CDCl ₃ ): δ = 7.22 (1H, m), 7.00 (1H, m), 4.82 (1H, m), 4.63 (1H, m), 4.29 (1H, d, J = 16.2 Hz), 4.11 (1H, d, J = 16.4 Hz), 4.05 (1H, m), 3.05 (1H, m), 2.85 (1H, m), 1.30 (9H, s).

Intermediate 2: 2,3,4,5,6,7-hexahydropyrrolo[3′,4′:3,4]pyrazolo[1,5-b][1,2]thiazine-1,1-dioxide p-toluenesulfonate

Step 1: tert-Butyl 3-iodo-4,6-dihydropyrrolo[3,4-c]pyrazole-5(2H)-carboxylate

Dissolve tert-butyl 4,6-dihydropyrrolo[3,4-c]pyrazole-5(2H)-carboxylate (9.41 g, 45 mmol) in 200 mL of 1,2-dichloroethane, add N-iodosuccinimide (13.16 g, 58.5 mmol), and reflux overnight. After evaporation of the solvent, the product was purified by silica gel column chromatography to afford tert-butyl 3-iodo-4,6-dihydropyrrolo[3,4-c]pyrazole-5(2H)-carboxylate (4.66 g) in a 31% yield. MS m/z [ESI]: 336.0 [M+1].

Step 2: tert-Butyl 2-(3-chloropropanesulfonyl)-3-iodo-4,6-dihydropyrrolo[3,4-c]pyrazole-5(2H)-carboxylate

Dissolve tert-butyl 3-iodo-4,6-dihydropyrrolo[3,4-c]pyrazole-5(2H)-carboxylate (2.80 g, 8.36 mmol) and triethylamine (1.69 g, 16.7 mmol) in tetrahydrofuran (80 mL), cool to -10°C, and add 3-chloropropanesulfonyl chloride (1.77 g, 10 mmol) dropwise. Stir overnight at low temperature. Add water (100 mL) and extract with dichloromethane. The organic phase is washed sequentially with citric acid solution, water, and brine, dried, evaporated, and concentrated, then purified by silica gel column chromatography to obtain tert-butyl 2-(3-chloropropanesulfonyl)-3-iodo-4,6-dihydropyrrolo[3,4-c]pyrazole-5(2H)-carboxylate (2.70 g) in a yield of 68%. ¹ H-NMR (400MHz, CDCl ₃ ): δ = 4.675 (2H, m), 4.35 (2H, m), 3.68 (4H, m), 2.26 (2H, m), 1.51 (9H, s).

Step 3: 6-tert-Butyloxycarbonyl-2,3,4,5,6,7-hexahydropyrrolo[3′,4′:3,4]pyrazolo[1,5-b][1,2]thiazine-1,1-dioxide

tert-Butyl 2-(3-chloropropanesulfonyl)-3-sulfo-4,6-dihydropyrrolo[3,4-c]pyrazole-5(2H)-carboxylate (230 mg, 0.48 mmol), zinc powder (126 mg, 1.93 mmol), a 0.5 M solution of zinc chloride in tetrahydrofuran (1.93 mL, 0.97 mmol), and tetrahydrofuran (20 mL) were added to a microwave reaction tube and nitrogen was introduced for 5 minutes. The reaction was heated in a microwave at 100°C under a nitrogen atmosphere for 1.5 hours. After cooling, tetrakis(triphenylphosphine)palladium (56 mg, 0.048 mmol) was added and nitrogen was introduced for 5 minutes. The reaction was heated in a microwave at 100°C under a nitrogen atmosphere for 2 hours. The product was filtered, and the filtrate was concentrated by evaporation and separated by silica gel column chromatography to give 6-tert-butyloxycarbonyl-2,3,4,5,6,7-hexahydropyrrolo[3′,4′:3,4]pyrazolo[1,5-b][1,2]thiazine-1,1-dioxide (23 mg) in a 15% yield. MS m/z [ESI]: 314.1 [M+1]. ¹ H-NMR (400 MHz, CDCl ₃ ): δ = 4.36 (2H, m), 4.11 (2H, m), 3.70 (2H, m), 3.35 (2H, m), 2.26 (2H, m), 1.47 (9H, s).

Step 4: 2,3,4,5,6,7-Hexahydropyrrolo[3′,4′:3,4]pyrazolo[1,5-b][1,2]thiazine-1,1-dioxide p-toluenesulfonate

6-tert-Butyloxycarbonyl-2,3,4,5,6,7-hexahydropyrrolo[3',4':3,4]pyrazolo[1,5-b][1,2]thiazine-1,1-dioxide (46 mg, 0.15 mmol) was dissolved in 1.5 mL of ethyl acetate, p-toluenesulfonic acid (47 mg, 0.30 mmol) was added, and the mixture was stirred at room temperature overnight. The resulting solid was collected by filtration, washed with ethyl acetate, and dried to give 2,3,4,5,6,7-hexahydropyrrolo[3',4':3,4]pyrazolo[1,5-b][1,2]thiazine-1,1-dioxide p-toluenesulfonate (48 mg) in an 83% yield. MS m/z [ESI]: 214.1 [M+1]. ¹ H-NMR (400MHz, DMSO-d ₆ ): δ = 9.40 (2H, brs), 7.48 (2H, d, J = 8.0Hz), 7.12 (2H, d, J = 8.0Hz), 4.27 (2H, s), 4. 08 (2H, s), 3.77 (2H, t, J = 6.6Hz), 3.45 (2H, t, J = 7.4Hz), 2.29 (3H, s), 2.16 (2H, m).

Intermediate 3: 3-methyl-2,4,5,6-tetrahydropyrrolo[3,4-c]pyrazole

Step 1: tert-Butyl 3-methyl-2-((2-(trimethylsilyl)ethoxy)methyl)-4,6-dihydropyrrolo[3,4-c]pyrazole-5(2H)-carboxylate

Dissolve tert-butyl 2-((2-(trimethylsilyl)ethoxy)methyl)-4,6-dihydropyrrolo[3,4-c]pyrazole-5(2H)-carboxylate (3.0 g, 8.85 mmol) in dry tetrahydrofuran (60 mL), cool to -78°C, and add dropwise a solution of butyllithium in tetrahydrofuran (2.4 M, 5.5 mL, 13.2 mmol). Stir at low temperature for 1.5 hours. Add dropwise iodomethane (1.90 g, 13.2 mmol), and stir at low temperature for 5 hours. Warm to room temperature, add water (50 mL), and extract with ethyl acetate. The organic phase is dried, evaporated, and concentrated, followed by silica gel column chromatography to yield tert-butyl 3-methyl-2-((2-(trimethylsilyl)ethoxy)methyl)-4,6-dihydropyrrolo[3,4-c]pyrazole-5(2H)-carboxylate (1.5 g) in a yield of 48%. MS m/z[ESI]：354.2[M+1].

Step 2: 3-Methyl-2,4,5,6-tetrahydropyrrolo[3,4-c]pyrazole

Dissolve tert-butyl 3-methyl-2-((2-(trimethylsilyl)ethoxy)methyl)-4,6-dihydropyrrolo[3,4-c]pyrazole-5(2H)-carboxylate (1.1 g, 3.12 mmol) in 20 mL of ethanol, add 1N hydrochloric acid (40 mL), and react at 90°C in a sealed tube for 3 hours. After cooling, adjust the pH to 13 with sodium hydroxide solution. After concentration, silica gel column chromatography was used to obtain the product, 3-methyl-2,4,5,6-tetrahydropyrrolo[3,4-c]pyrazole (280 mg), in a yield of 73%. MS m/z [ESI]: 124.1 [M+1].

Intermediate 4: 5-Methanesulfonylisoindoline hydrochloride

Step 1: 5-Bromoisoindoline

4-Bromophthalimide (22.6 g, 100 mmol) was added to dry tetrahydrofuran (250 mL), and borane-dimethyl sulfide complex (51 mL, 500 mmol) was added dropwise. The mixture was stirred at room temperature for 2 hours and then refluxed overnight. After cooling, methanol was carefully added dropwise to destroy the excess borane. The product was concentrated by evaporation and purified by silica gel column chromatography to afford 5-bromoisoindoline (10.36 g) in a yield of 52%. MS m/z [ESI]: 198.0 [M+1].

Step 2: 5-Bromo-2-tert-butyloxycarbonylisoindoline

Dissolve 5-bromoisoindoline (10.36 g, 52.3 mmol) in 80 mL of dichloromethane and cool in an ice bath. Add Boc anhydride (22.8 g, 104.6 mmol) dropwise, followed by sodium carbonate (16.6 g, 156.9 mmol) and water (150 mL). Stir in an ice bath for 4 hours. Separate the organic phase and wash with brine. After concentration, silica gel column chromatography affords the product, 5-bromo-2-tert-butyloxycarbonylisoindoline (13.3 g), in an 85% yield. MS m/z [ESI]: 298.0 [M+1]. ¹ H-NMR (400 MHz, CDCl ₃ ): δ = 7.37 (2H, m), 7.11 (1H, m), 4.62 (4H, m), 1.51 (9H, s).

Step 3: 5-Methanesulfonyl-2-tert-Butyloxycarbonylisoindoline

5-Bromo-2-tert-butoxycarbonylisoindoline (5.96 g, 20 mmol), sodium methanesulfinate (90%, 2.94 g, 26 mmol), cuprous iodide (762 mg, 4 mmol), and L-proline (920 mg, 8 mmol) were added to dimethyl sulfoxide (80 mL), purged with nitrogen, and stirred at 120°C for 2 days. After cooling, the mixture was poured into water and extracted with ethyl acetate. The organic phase was dried, evaporated, and concentrated, followed by silica gel column chromatography to afford 5-methanesulfonyl-2-tert-butoxycarbonylisoindoline (5.46 g) in a 92% yield. MS m/z [ESI]: 298.1 [M+1].

Step 4: 5-Methanesulfonylisoindoline hydrochloride

5-Methanesulfonyl 2-tert-butoxycarbonyl isoindoline (5.46 g, 18.4 mmol) was dissolved in methanol/dichloromethane (1:1, 80 mL). Hydrogen chloride gas was introduced to saturation and stirred at room temperature for 1 hour. The mixture was poured into 800 mL of diethyl ether, and the precipitate was collected by filtration, washed with diethyl ether, and dried to give 5-methanesulfonyl isoindoline hydrochloride (3.44 g) in an 80% yield. MS m/z [ESI]: 198.0 [M+1]. ¹ H-NMR (400 MHz, CDCl ₃ ): δ = 7.82 (1H, s), 7.81 (1H, d, J = 8.0 Hz), 7.43 (1H, d, J = 8.0 Hz), 4.31 (4H, s), 3.05 (3H, s), 2.30 (2H, brs).

Intermediate 5: 5-Methanesulfonamidoisoindoline

Step 1: 1-Oxo-5-methanesulfonamidoisoindoline

5-Aminoisoindolin-1-one (444 mg, 3 mmol) was dissolved in 15 mL of pyridine, and methylsulfonyl chloride (378 mg, 3.3 mmol) was added. The mixture was stirred at room temperature for 4 hours. After evaporation of the solvent, the mixture was purified by silica gel column chromatography to afford 1-oxo-5-methylsulfonamidoisoindoline (610 mg) in a 90% yield. MS m/z [ESI]: 227.0 [M+1].

Step 2: 5-Methanesulfonamidoisoindoline

Dissolve 1-oxo-5-methylsulfonylisoindoline (610 mg, 2.7 mmol) in tetrahydrofuran (10 mL). Add a 1 M solution of borane in tetrahydrofuran (8.1 mL, 8.1 mmol). Stir at room temperature for 2 hours and then reflux overnight. After cooling, methanol is carefully added dropwise to destroy excess borane. Evaporation and concentration followed by silica gel column chromatography afford 5-methylsulfonylisoindoline (315 mg) in a 55% yield. MS m/z [ESI]: 213.1 [M+1].

Intermediate 6: 4-chloro-5-methylsulfonylisoindoline

Step 1: 4-Chloro-5-aminoisoindolin-1-one

5-Aminoisoindolin-1-one (2.96 g, 20 mmol) was added to chloroform (50 mL), followed by N-chlorosuccinimide (2.67 g, 20 mmol). The mixture was stirred at reflux for 2 hours. Evaporation and concentration were followed by silica gel column chromatography to afford 4-chloro-5-aminoisoindolin-1-one (2.58 g) in a 70% yield. MS m/z [ESI]: 183.0 [M+1].

Step 2: 4-Chloro-5-iodoisoindolin-1-one

4-Chloro-5-aminoisoindolin-1-one (2.58 g, 14 mmol) was added to 15 mL of 2M sulfuric acid and cooled in an ice bath. A solution of sodium nitrite (0.97 g, 28 mmol) and water (1.5 mL) was added dropwise and stirred at low temperature for 30 minutes. Potassium iodide (11.62 g, 70 mmol) was then added and stirred in an ice bath for 2 hours and then at room temperature for 2 hours. The mixture was extracted with dichloromethane and the organic phase was washed with water and then with brine. After concentration, silica gel column chromatography was used to obtain the product, 4-chloro-5-iodoisoindolin-1-one (2.48 g), in a 60% yield. MS m/z [ESI]: 293.9 [M+1]. ¹ H-NMR (400MHz, CDCl ₃ ): δ=8.01 (1H, d, J=8.0Hz), 7.49 (1H, d, J=8.0Hz), 4.44 (4H, m).

Step 3: 4-Chloro-5-methylsulfonylisoindolin-1-one

4-Chloro-5-iodoisoindolin-1-one (1.76 g, 6 mmol), sodium methanesulfinate (90%, 0.884 g, 7.8 mmol), cuprous iodide (229 mg, 1.2 mmol), and L-proline (276 mg, 2.4 mmol) were added to dimethyl sulfoxide (25 mL), purged with nitrogen, and stirred at 110°C for 2 days. After cooling, the mixture was poured into water and extracted with ethyl acetate. The organic phase was dried, evaporated, and then purified by silica gel column chromatography to give 4-chloro-5-methylsulfonylisoindolin-1-one (1.03 g) in a 70% yield. MS m/z [ESI]: 246.0 [M+1].

Step 4: 4-Chloro-5-methylsulfonylisoindoline

4-Chloro-5-methylsulfonylisoindolin-1-one (249 mg, 1 mmol) was dissolved in tetrahydrofuran (10 mL). A 1 M solution of borane in tetrahydrofuran (4 mL, 4 mmol) was added. The mixture was stirred at room temperature for 2 hours and then refluxed overnight. After cooling, methanol was carefully added dropwise to destroy the excess borane. The residue was concentrated by evaporation and purified by silica gel column chromatography to afford 4-chloro-5-methylsulfonylisoindolin-1-one (170 mg) in a 73% yield. MS m/z [ESI]: 232.0 [M+1]. ¹ H-NMR (400 MHz, CDCl ₃ ): δ = 8.08 (1H, d, J = 8.0 Hz), 7.32 (1H, d, J = 8.0 Hz), 4.30 (4H, m), 3.32 (3H, s), 2.80 (1H, brs).

Intermediate 7: Isoindoline-5-sulfonamide

Step 1: 1-Oxoisoindoline-5-sulfonyl chloride

5-Aminoisoindolin-1-one (5.92 g, 40 mmol) was added to a mixed solvent of concentrated hydrochloric acid/glacial acetic acid (13.3 mL/4.0 mL) and cooled in an ice bath. A solution of sodium nitrite (3.04 g, 28 mmol) and water (4.4 mL) was added dropwise and stirred at low temperature for 30 minutes. Meanwhile, glacial acetic acid was added to a separate flask and sulfur dioxide was passed through until saturated. Cuprous chloride (0.99 g, 10 mmol) was added and sulfur dioxide was continued to be passed through with stirring until the solid was substantially dissolved. The diazonium salt solution prepared above was slowly added dropwise and stirred at low temperature for half an hour and then at room temperature for 1 hour. Extract with dichloromethane, wash with water, dry, and spin-dry to obtain 1-oxoisoindoline-5-sulfonyl chloride (8.33 g) in a 90% yield. MS m/z [ESI]: 232.0 [M+1].

Step 2: 1-Oxoisoindoline-5-sulfonamide

1-Oxoisoindoline-5-sulfonyl chloride (463 mg, 2 mmol) was added to 20 mL of acetonitrile. Concentrated aqueous ammonia (1 mL, 12 mmol) was added dropwise and stirred for 3 hours. The mixture was neutralized with 6N hydrochloric acid until neutral. The acetonitrile was removed by rotary evaporation, and the residue was filtered after adding water. The filter cake was washed with water and ethyl acetate, and dried to obtain 1-oxoisoindoline-5-sulfonamide (288 mg) in a yield of 68%. MS m/z [ESI]: 213.0 [M+1].

Step 3: Isoindoline-5-sulfonamide

Dissolve 1-oxoisoindoline-5-sulfonamide (288 mg, 1.36 mmol) in tetrahydrofuran (15 mL). Add borane in tetrahydrofuran (1 M, 6.8 mL, 6.8 mmol). Stir at room temperature for 2 hours and then reflux overnight. After cooling, carefully add methanol dropwise to destroy excess borane. Evaporation and concentration followed by silica gel column chromatography afford isoindoline-5-sulfonamide (162 mg) in a 60% yield. MS m/z [ESI]: 199.0 [M+1].

Intermediate 8: 5-cyclopropylsulfonylisoindoline hydrochloride

Step 1: 5-cyclopropylsulfonyl-2-tert-butoxycarbonylisoindoline

5-Bromo-2-tert-butoxycarbonylisoindoline (2.98 g, 10 mmol), sodium cyclopropylsulfinate (90%, 1.85 g, 13 mmol), cuprous iodide (381 mg, 2 mmol), and L-proline (460 mg, 4 mmol) were added to dimethyl sulfoxide (40 mL), purged with nitrogen, and stirred at 110°C for 2 days. After cooling, the mixture was poured into water and extracted with ethyl acetate. The organic phase was dried, evaporated, and then purified by silica gel column chromatography to obtain 5-cyclopropylsulfonyl-2-tert-butoxycarbonylisoindoline (2.30 g) in a yield of 71%. MS m/z [ESI]: 324.1 [M+1].

Step 4: 5-Cyclopropanesulfonylisoindoline hydrochloride

Dissolve 5-cyclopropylsulfonyl-2-tert-butoxycarbonylisoindoline (2.30 g, 7.1 mmol) in methanol/dichloromethane (1:1, 40 mL). Bubble hydrogen chloride gas into the mixture until saturated and stir at room temperature for 2 hours. Pour the mixture into 250 mL of diethyl ether, collect the precipitate by filtration, wash with diethyl ether, and dry to obtain 5-cyclopropylsulfonylisoindoline hydrochloride (1.85 g) in a 100% yield. MS m/z [ESI]: 224.1 [M+1].

Example 1: (2R,3S,5R)-5-(2,3,4,5,6,7-hexahydropyrrolo[3′,4′:3,4]pyrazolo[1,5-b][1,2]thiazine-1,1-dioxide-6-yl)-2-(2,5-difluorophenyl)tetrahydro-2H-pyran-3-amine

Step 1: tert-Butyl (2R,3S,5R)-5-(2,3,4,5,6,7-hexahydropyrrolo[3′,4′:3,4]pyrazolo[1,5-b][1,2]thiazine-1,1-dioxide-6-yl)-2-(2,5-difluorophenyl)tetrahydro-2H-pyran-3-ylcarbamate

Tert-butyl (2R,3S)-2-(2,5-difluorophenyl)-5-oxotetrahydro-2H-pyran-3-ylcarbamate (48 mg, 0.146 mmol), 2,3,4,5,6,7-hexahydropyrrolo[3′,4′:3,4]pyrazolo[1,5-b][1,2]thiazine-1,1-dioxide p-toluenesulfonate (48 mg, 0.13 mmol), and triethylamine (10 mg, 0.1 mmol) were added to N,N-dimethylacetamide (2 mL) and stirred at room temperature for 3 hours. The mixture was cooled in an ice bath, and sodium triacetoxyborohydride (87 mg, 0.39 mmol) was added. The mixture was slowly warmed to room temperature and stirred overnight. Saturated sodium bicarbonate solution was added, and the mixture was extracted with dichloromethane, washed with saturated brine, dried, concentrated, and purified by column chromatography to give tert-butyl (2R,3S,5R)-5-(2,3,4,5,6,7-hexahydropyrrolo[3′,4′:3,4]pyrazolo[1,5-b][1,2]thiazine-1,1-dioxide-6-yl)-2-(2,5-difluorophenyl)tetrahydro-2H-pyran-3-ylcarbamate (48 mg). Yield: 71%. MS m/z [ESI]: 525.2 [M+1]. ¹ H NMR (400MHz, CDCl ₃ ): δ = 7.20 (1H, m), 6.98 (2H, m) 4.53 (1H, m), 4.16 (4H, m), 4.02 (2H, m), 3.64 (2H, m) ), 3.32 (3H, m), 2.80 (2H, m), 2.56 (2H, m), 2.39 (1H, m), 1.45 (1H, m), 1.26 (9H, s).

Step 2: (2R,3S,5R)-5-(2,3,4,5,6,7-hexahydropyrrolo[3′,4′:3,4]pyrazolo[1,5-b][1,2]thiazine-1,1-dioxide-6-yl)-2-(2,5-difluorophenyl)tetrahydro-2H-pyran-3-amine

Tert-butyl 5-(2,3,4,5,6,7-hexahydropyrrolo[3′,4′:3,4]pyrazolo[1,5-b][1,2]thiazine-1,1-dioxide-6-yl)-2-(2,5-difluorophenyl)tetrahydro-2H-pyran-3-ylcarbamate (45 mg, 0.086 mmol) and p-toluenesulfonic acid monohydrate (75 mg, 0.43 mmol) were added to dichloromethane (1 mL) and the mixture was stirred at room temperature. The mixture was stirred overnight. Saturated sodium bicarbonate solution was added, extracted with dichloromethane, the solvent was dried, and purified by column chromatography to obtain (2R, 3S, 5R)-5-(2,3,4,5,6,7-hexahydropyrrolo[3′,4′:3,4]pyrazolo[1,5-b][1,2]thiazine-1,1-dioxide-6-yl)-2-(2,5-difluorophenyl)tetrahydro-2H-pyran-3-amine (15 mg), with a yield of 42%, MS m/z[ESI]: 425.1[M+1]. ¹ H NMR (400MHz, CDCl ₃ ): δ=7.14(1H,m), 7.01(2H,m), 4.18(4H,m), 4.03(2H,m), 3.66(2H,m), 3.32 (3H,m), 2.80(2H,m), 2.56(2H,m), 2.35(1H,m), 1.38(1H,m), 1.26(2H,brs).

Example 2: 5-((3R,5S,6R)-5-amino-6-(2,5-difluorophenyl)tetrahydro-2H-pyran-3-yl)-3-methyl-1-methanesulfonyl-1,4,5,6-tetrahydropyrrolo[3,4-c]pyrazole

Step 1: 5-((3R,5S,6R)-5-tert-Butoxycarbonylamino-6-(2,5-difluorophenyl)tetrahydro-2H-pyran-3-yl)-3-methyl-1,4,5,6-tetrahydropyrrolo[3,4-c]pyrazole

Tert-butyl (2R,3S)-2-(2,5-difluorophenyl)-5-oxotetrahydro-2H-pyran-3-ylcarbamate (327 mg, 1 mmol), 3-methyl-2,4,5,6-tetrahydropyrrolo[3,4-c]pyrazole (123 mg, 1 mmol), and glacial acetic acid (30 mg, 0.5 mmol) were added to 1,2-dichloroethane (10 mL) and stirred at room temperature for 2 hours. The mixture was cooled in an ice bath, and sodium triacetoxyborohydride (672 mg, 3 mmol) was added. The mixture was slowly warmed to room temperature and stirred overnight. Saturated sodium bicarbonate solution was added, and the mixture was extracted with dichloromethane, washed with saturated brine, dried, concentrated, and purified by column chromatography to give 5-((3R,5S,6R)-5-tert-butoxycarbonylamino-6-(2,5-difluorophenyl)tetrahydro-2H-pyran-3-yl)-3-methyl-1,4,5,6-tetrahydropyrrolo[3,4-c]pyrazole (145 mg) in a 33% yield. MS m/z [ESI]: 435.2 [M+1].

Step 2: 5-((3R, 5S, 6R)-5-tert-butoxycarbonylamino-6-(2,5-difluorophenyl)tetrahydro-2H-pyran-3-yl)-3-methyl-1-methanesulfonyl-1,4,5,6-tetrahydropyrrolo[3,4-c]pyrazole

5-((3R,5S,6R)-5-tert-Butoxycarbonylamino-6-(2,5-difluorophenyl)tetrahydro-2H-pyran-3-yl)-3-methyl-1,4,5,6-tetrahydropyrrolo[3,4-c]pyrazole (145 mg, 0.33 mmol) and triethylamine (53 mg, 0.53 mmol) were added to tetrahydrofuran (10 mL), cooled in an ice bath, and methylsulfonyl chloride (49 mg, The reaction mixture was stirred for 2 hours. The mixture was poured into water and extracted with dichloromethane. The organic phase was dried and purified by column chromatography to obtain 5-((3R,5S,6R)-5-tert-butoxycarbonylamino-6-(2,5-difluorophenyl)tetrahydro-2H-pyran-3-yl)-3-methyl-1-methanesulfonyl-1,4,5,6-tetrahydropyrrolo[3,4-c]pyrazole (16 mg) in a yield of 9.5%. m/z[ESI]: 513.2[M+1]. ¹ H NMR (400MHz, CDCl ₃ ): δ = 7.21 (1H, m), 6.96 (2H, m), 4.52 (1H, m), 4.37-4.20 (2H, m), 4.08 (2H, m), 3.75 (3H, m), 3 .39(1H,m), 3.28(3H,s), 3.07(1H,m), 2.48(1H,m), 2.26(3H,s), 1.53(1H,m), 1.27(9H,s).

Step 3: 5-((3R,5S,6R)-5-amino-6-(2,5-difluorophenyl)tetrahydro-2H-pyran-3-yl)-3-methyl-1-methanesulfonyl-1,4,5,6-tetrahydropyrrolo[3,4-c]pyrazole

5-((3R,5S,6R)-5-tert-Butyloxycarbonylamino-6-(2,5-difluorophenyl)tetrahydro-2H-pyran-3-yl)-3-methyl-1-methylsulfonyl-1,4,5,6-tetrahydropyrrolo[3,4-c]pyrazole (16 mg, 0.031 mmol) and benzenesulfonic acid (20 mg, 0.125 mmol) were added to dichloromethane (0.7 mL) and stirred at room temperature overnight. Triethylamine (23 mg, 0.228 mmol) was added, the solvent was dried and purified by column chromatography to give 5-((3R,5S,6R)-5-amino-6-(2,5-difluorophenyl)tetrahydro-2H-pyran-3-yl)-3-methyl-1-methylsulfonyl-1,4,5,6-tetrahydropyrrolo[3,4-c]pyrazole (1.7 mg) in a yield of 13%. MS m/z[ESI]: 413.2[M+1]. ¹ H NMR (400MHz, CDCl ₃ ): δ = 7.16 (1H, m), 7.01 (2H, m), 4.30 (1H, m), 4.22 (1H, m), 4.10 (1H, m), 3.90-3.70 (3H, m), 3 .42(1H,m), 3.27(3H,s), 3.02(2H,m), 2.48(2H,m), 2.26(3H,s), 2.00(1H,m), 1.60(1H,m).

Example 3: 5-((3R,5S,6R)-5-amino-6-(2,5-difluorophenyl)tetrahydro-2H-pyran-3-yl)-5,6-dihydro-4H-thieno[3,2-c]pyrrole-2-carboxylic acid

Step 1: 5-((3R,5S,6R)-5-tert-Butoxycarbonylamino-6-(2,5-difluorophenyl)tetrahydro-2H-pyran-3-yl)-5,6-dihydro-4H-thieno[3,2-c]pyrrole-2-carboxylic acid

Following the procedure of Step 1 in Example 1, except that 2,3,4,5,6,7-hexahydropyrrolo[3′,4′:3,4]pyrazolo[1,5-b][1,2]thiazine-1,1-dioxide p-toluenesulfonate was replaced with 5,6-dihydro-4H-thieno[3,2-c]pyrrole-2-carboxylic acid hydrochloride (commercially available), the title compound was obtained in a 44% yield. MS m/z [ESI]: 481.1 [M+1]. ¹ H NMR (400MHz, CDCl ₃ ): δ = 9.22 (1H, s), 7.56 (1H, s), 7.11 (1H, m), 6.96 (2H, m), 4.55 (1H, m), 4.30 (2H, m), 4.12 (2H, m) ), 3.96 (2H, m), 3.80 (1H, m), 3.47 (1H, m), 3.05 (1H, m), 2.52 (1H, m), 1.58 (1H, m), 1.26 (9H, s).

Step 2: 5-((3R,5S,6R)-5-amino-6-(2,5-difluorophenyl)tetrahydro-2H-pyran-3-yl)-5,6-dihydro-4H-thieno[3,2-c]pyrrole-2-carboxylic acid

Referring to the method of Step 2 in Example 1, the title compound was obtained by replacing tert-butyl 5-(2,3,4,5,6,7-hexahydropyrrolo[3′,4′:3,4]pyrazolo[1,5-b][1,2]thiazine-1,1 dioxide-6-yl)-2-(2,5-difluorophenyl)tetrahydro-2H-pyran-3-ylcarbamate with 5-((3R,5S,6R)-5-tert-butoxycarbonylamino-6-(2,5-difluorophenyl)tetrahydro-2H-pyran-3-yl)-5,6-dihydro-4H-thieno[3,2-c]pyrrole-2-carboxylic acid in a yield of 61%. MS m/z[ESI]: 381.1[M+1]. ¹ H NMR (400MHz, DMSO-d ₆ ): δ=7.35-7.10(3H,m), 7.06(1H,s), 4.12(2H,m), 3.90(2H,m), 3.76(2H,m), 3.23(1H,m), 2.82(2H,m), 2.28(1H,m), 1.37(1H,m).

Example 4: (2R,3S,5R)-5-(5-methylsulfonylisoindolin-2-yl)-2-(2,5-difluorophenyl)tetrahydro-2H-pyran-3-amine

Step 1: tert-Butyl (2R,3S,5R)-5-(5-methylsulfonylisoindolin-2-yl)-2-(2,5-difluorophenyl)tetrahydro-2H-pyran-3-ylcarbamate

Following the procedure of Step 1 in Example 1, substituting 5-methanesulfonylisoindoline hydrochloride for 2,3,4,5,6,7-hexahydropyrrolo[3′,4′:3,4]pyrazolo[1,5-b][1,2]thiazine-1,1-dioxide p-toluenesulfonate, the title compound was obtained in a 54% yield. MS m/z [ESI]: 509.2 [M+1]. ¹ H NMR (400MHz, CDCl ₃ ): δ=7.83(1H,d,J=8.0Hz), 7.82(1H,s), 7.42(1H,d,J=8.0Hz), 7.24(1H,m), 6.97(2H,m), 4.50(1H,m), 4.32(2H,m), 4.08 (4H, m), 3.80 (1H, m), 3.43 (1H, t, J = 10.6Hz), 3.04 (3H, s), 2.95 (1H, m), 2.54 (1H, m), 1.54 (1H, m), 1.28 (9H, s).

Step 2: (2R,3S,5R)-5-(5-methylsulfonylisoindolin-2-yl)-2-(2,5-difluorophenyl)tetrahydro-2H-pyran-3-amine

Referring to the method of Step 2 in Example 1, the title compound was obtained by replacing tert-butyl 5-(2,3,4,5,6,7-hexahydropyrrolo[3′,4′:3,4]pyrazolo[1,5-b][1,2]thiazine-1,1 dioxide-6-yl)-2-(2,5-difluorophenyl)tetrahydro-2H-pyran-3-ylcarbamate with tert-butyl (2R,3S,5R)-5-(5-methylsulfonylisoindolin-2-yl)-2-(2,5-difluorophenyl)tetrahydro-2H-pyran-3-ylcarbamate. The yield was 68%. MS m/z[ESI]: 409.1[M+1]. ¹ H NMR (400MHz, CDCl ₃ ): δ = 7.83 (1H, d, J = 8.0Hz), 7.82 (1H, s), 7.42 (1H, d, J = 8.0Hz), 7.17 (1H, m), 7.02 (2H, m), 4.28 (1H, m), 4.24 (2 H, d, J=9.6Hz), 4.09 (4H, m), 3.45 (1H, m), 3.04 (3H, s), 2.92 (2H, m), 2.49 (1H, m), 1.47 (1H, m), 1.30 (2H, brs).

Example 5: (2R,3S,5R)-5-(5-methanesulfonamidoisoindolin-2-yl)-2-(2,5-difluorophenyl)tetrahydro-2H-pyran-3-amine

Step 1: tert-Butyl (2R,3S,5R)-5-(5-methanesulfonamidoisoindolin-2-yl)-2-(2,5-difluorophenyl)tetrahydro-2H-pyran-3-ylcarbamate

Referring to the method of Step 1 in Example 1, except that 5-methanesulfonamidoisoindoline was used instead of 2,3,4,5,6,7-hexahydropyrrolo[3′,4′:3,4]pyrazolo[1,5-b][1,2]thiazine-1,1-dioxide p-toluenesulfonate, the yield was 50%. MS m/z [ESI]: 524.2 [M+1].

Step 2: (2R,3S,5R)-5-(5-methanesulfonamidoisoindolin-2-yl)-2-(2,5-difluorophenyl)tetrahydro-2H-pyran-3-amine

Referring to the method of Step 2 in Example 1, the title compound was obtained in a yield of 57% by replacing tert-butyl 5-(2,3,4,5,6,7-hexahydropyrrolo[3′,4′:3,4]pyrazolo[1,5-b][1,2]thiazine-1,1 dioxide-6-yl)-2-(2,5-difluorophenyl)tetrahydro-2H-pyran-3-ylcarbamate with tert-butyl (2R,3S,5R)-5-(5-methanesulfonamidoisoindolin-2-yl)-2-(2,5-difluorophenyl)tetrahydro-2H-pyran-3-ylcarbamate. MS m/z[ESI]: 424.1[M+1]. ¹ H NMR (400MHz, CDCl ₃ ): δ = 7.24 (1H, m), 7.17 (1H, m), 7.07-6.90 (4H, m), 5.35 (1H, s), 4.35-4.20 (2H, m), 4.08 ( 4H, m), 3.65 (2H, m), 3.00 (3H, s), 2.82 (1H, m), 2.32 (1H, m), 1.47 (1H, m), 1.25 (2H, brs).

Example 6: (2R,3S,5R)-5-(5-bromoisoindolin-2-yl)-2-(2,5-difluorophenyl)tetrahydro-2H-pyran-3-amine

Step 1: tert-Butyl (2R,3S,5R)-5-(5-bromoisoindolin-2-yl)-2-(2,5-difluorophenyl)tetrahydro-2H-pyran-3-ylcarbamate

Following the procedure of Step 1 in Example 1, substituting 5-bromoisoindoline for 2,3,4,5,6,7-hexahydropyrrolo[3′,4′:3,4]pyrazolo[1,5-b][1,2]thiazine-1,1-dioxide p-toluenesulfonate, the title compound was obtained in a 62% yield. MS m/z [ESI]: 509.1 [M+1]. ¹ H NMR (400MHz, CDCl ₃ ): δ = 7.36 (1H, s), 7.34 (1H, d, J = 8.0Hz), 7.23 (1H, m), 7.09 (1H, d, J = 8.0Hz), 6.97 (2H, m), 4.49 (1H, m), 4.30 (2H, m) , 3.94 (4H, m), 3.79 (1H, m), 3.42 (1H, t, J = 10.8Hz), 2.92 (1H, m), 2.52 (1H, d, J = 10.8Hz), 1.54 (1H, m), 1.27 (9H, s).

Step 2: (2R,3S,5R)-5-(5-bromoisoindolin-2-yl-2-(2,5-difluorophenyl)tetrahydro-2H-pyran-3-amine

Referring to the method in Step 2 of Example 1, the title compound was obtained by replacing tert-butyl 5-(2,3,4,5,6,7-hexahydropyrrolo[3′,4′:3,4]pyrazolo[1,5-b][1,2]thiazine-1,1 dioxide-6-yl)-2-(2,5-difluorophenyl)tetrahydro-2H-pyran-3-ylcarbamate with tert-butyl (2R,3S,5R)-5-(5-bromoisoindolin-2-yl)-2-(2,5-difluorophenyl)tetrahydro-2H-pyran-3-ylcarbamate. The yield was 69%. MS m/z[ESI]: 409.1[M+1]. ¹ H NMR (400MHz, CDCl ₃ ): δ = 7.37 (1H, s), 7.34 (1H, d, J = 8.0Hz), 7.15 (1H, m), 7.10 (1H, d, J = 8.0Hz), 7.02 (2H, m), 4.27 (1H, m), 4.22 (1H, d, J = 9.2 Hz), 3.99 (4H, m), 3.79 (1H, m), 3.42 (1H, t, J = 10.8Hz), 2.87 (2H, m), 2.47 (1H, d, J = 10.8Hz), 1.48 (1H, m), 1.32 (2H, brs).

Example 7: (2R,3S,5R)-5-(4-chloro-5-methylsulfonylisoindolin-2-yl)-2-(2,5-difluorophenyl)tetrahydro-2H-pyran-3-amine

Step 1: tert-Butyl (2R,3S,5R)-5-(4-chloro-5-methanesulfonylisoindolin-2-yl)-2-(2,5-difluorophenyl)tetrahydro-2H-pyran-3-ylcarbamate

Following the procedure of Step 1 in Example 1, substituting 4-chloro-5-methylsulfonylisoindoline for 2,3,4,5,6,7-hexahydropyrrolo[3′,4′:3,4]pyrazolo[1,5-b][1,2]thiazine-1,1-dioxide p-toluenesulfonate, the title compound was obtained in a 56% yield. MS m/z [ESI]: 543.1 [M+1].

Step 2: (2R,3S,5R)-5-(4-chloro-5-methanesulfonylisoindolin-2-yl)-2-(2,5-difluorophenyl)tetrahydro-2H-pyran-3-amine

Referring to the method of Step 2 in Example 1, the title compound was obtained by replacing tert-butyl 5-(2,3,4,5,6,7-hexahydropyrrolo[3′,4′:3,4]pyrazolo[1,5-b][1,2]thiazine-1,1 dioxide-6-yl)-2-(2,5-difluorophenyl)tetrahydro-2H-pyran-3-ylcarbamate with tert-butyl (2R,3S,5R)-5-(4-chloro-5-methanesulfonylisoindolin-2-yl)-2-(2,5-difluorophenyl)tetrahydro-2H-pyran-3-ylcarbamate. The yield was 47%. MS m/z[ESI]: 443.1[M+1]. ¹ HNMR (400MHz, CDCl ₃ ): δ = 8.06 (1H, d, J = 7.6Hz), 7.33 (1H, d, J = 7.6Hz), 7.19 (1H, m), 6.99 (2H, m), 4.69 (1H, m), 4.22 (4H , m), 3.71 (1H, m), 3.33 (1H, m), 3.27 (3H, s), 2.91 (1H, m), 2.61 (2H, m), 1.52 (1H, m), 1.32 (2H, brs).

Example 8: 2-((3R,5S,6R)-5-amino-6-(2,5-difluorophenyl)tetrahydro-2H-pyran-3-yl)isoindoline-5-sulfonamide

Step 1: 2-((3R,5S,6R)-5-tert-Butoxycarbonylamino-6-(2,5-difluorophenyl)tetrahydro-2H-pyran-3-yl)isoindoline-5-sulfonamide

Following the procedure of Step 1 in Example 1, substituting isoindoline-5-sulfonamide hydrochloride for 2,3,4,5,6,7-hexahydropyrrolo[3′,4′:3,4]pyrazolo[1,5-b][1,2]thiazine-1,1-dioxide p-toluenesulfonate, the title compound was obtained in a 46% yield. MS m/z [ESI]: 510.2 [M+1].

Step 2: 2-((3R,5S,6R)-5-amino-6-(2,5-difluorophenyl)tetrahydro-2H-pyran-3-yl)isoindoline-5-sulfonamide

Referring to the method of Step 2 in Example 1, tert-butyl 5-(2,3,4,5,6,7-hexahydropyrrolo[3′,4′:3,4]pyrazolo[1,5-b][1,2]thiazine-1,1 dioxide-6-yl)-2-(2,5-difluorophenyl)tetrahydro-2H-pyran-3-ylcarbamate was replaced with 2-((3R,5S,6R)-5-tert-butoxycarbonylamino-6-(2,5-difluorophenyl)tetrahydro-2H-pyran-3-yl)isoindoline-5-sulfonamide to obtain the title compound in a yield of 37%. MS m/z[ESI]: 410.1[M+1]. ¹ H NMR (400MHz, CDCl ₃ ): δ=7.81 (1H, d, J=7.6Hz), 7.80 (1H, s), 7.37 (1H, d, J=7.6Hz), 7.17 (1H, m), 7.02 (2H, m), 4.79 (2H, brs), 4.28 (1H, d, J=9 .6Hz), 4.23 (1H, d, J = 9.6Hz), 4.07 (4H, m), 3.43 (1H, t, J = 10.8Hz), 2.90 (2H, m), 2.49 (1H, m), 1.49 (1H, m), 1.31 (2H, brs).

Example 9: (2R,3S,5R)-5-(5-cyclopropylsulfonylisoindolin-2-yl)-2-(2,5-difluorophenyl)tetrahydro-2H-pyran-3-amine

Step 1: tert-Butyl (2R,3S,5R)-5-(5-cyclopropylsulfonylisoindolin-2-yl)-2-(2,5-difluorophenyl)tetrahydro-2H-pyran-3-ylcarbamate

Following the procedure of Step 1 in Example 1, substituting 5-cyclopropylsulfonylisoindoline hydrochloride for 2,3,4,5,6,7-hexahydropyrrolo[3′,4′:3,4]pyrazolo[1,5-b][1,2]thiazine-1,1-dioxide p-toluenesulfonate, the title compound was obtained in a 53% yield. MS m/z [ESI]: 535.2 [M+1].

Step 2: (2R,3S,5R)-5-(5-cyclopropanesulfonylisoindolin-2-yl)-2-(2,5-difluorophenyl)tetrahydro-2H-pyran-3-amine

Referring to the method of Step 2 in Example 1, the title compound was obtained by replacing tert-butyl 5-(2,3,4,5,6,7-hexahydropyrrolo[3′,4′:3,4]pyrazolo[1,5-b][1,2]thiazine-1,1 dioxide-6-yl)-2-(2,5-difluorophenyl)tetrahydro-2H-pyran-3-ylcarbamate with tert-butyl (2R,3S,5R)-5-(5-cyclopropylsulfonylisoindolin-2-yl)-2-)2,5-difluorophenyl)tetrahydro-2H-pyran-3-ylcarbamate to obtain tert-butyl 5-(2,3,4,5,6,7-hexahydropyrrolo[3′,4′:3,4]pyrazolo[1,5-b][1,2]thiazine-1,1 dioxide-6-yl)-2-(2,5-difluorophenyl)tetrahydro-2H-pyran-3-ylcarbamate in an 80% yield. MS m/z[ESI]: 435.2[M+1]. ¹ H NMR (400MHz, CDCl ₃ ): δ = 7.77 (1H, d, J = 8.0Hz), 7.76 (1H, s), 7.42 (1H, d, J = 8.0Hz), 7.17 (1H, m), 7.02 (2H, m), 4.30 (2H, m), 4.10 (4H, m), 3 .54(1H,t,J=10.8Hz), 3.24(1H,m), 2.95(2H,m), 2.45(1H,m), 1.52(1H,m), 1.28(2H,brs), 1.05(2H,m), 0.87(2H,m).

Biological Experiment Example 1: Determination of DPP-IV Inhibitory Activity

The following method is used to determine the inhibitory activity of the compounds of the present application on DPP-IV in plasma. The _inhibitory activity is expressed as _IC50 value, which is the concentration of the compound when the activity of DPP-IV is inhibited by 50%.

Materials and methods:

Material:

a. White 384-well plate (Perkin Elmer, Catalog No. 607290/99)

b. HEPES buffer: Prepare 50 mL of 0.5 M HEPES buffer using 1 M HEPES buffer (Invitrogen, Catalog No. 15630-080): Take 25 mL of 1 M HEPES buffer, add appropriate amount of ddH ₂ O (redistilled water), adjust the pH to 7.8 with NaOH, and finally add ddH ₂ O to 50 mL.

c. Rat plasma: Blood was collected from the rat orbit, anticoagulated with heparin, and centrifuged at 4000 rpm for 10 minutes. The supernatant plasma was used as the enzyme source for DPP-IV.

d. The DPP-IV enzyme reaction substrate H-Gly-Pro-AMC (glycine-proline-7-amino-4-methylcoumarin) was synthesized by the inventors of the present application and dissolved in DMSO to form a 100 mM stock solution.

e.1M _MgCl2

f.1.5M NaCl

g.10% BAS

h.DMSO

i.ddH ₂ O

j. Test compounds: positive control compound Omarigliptin and some compounds of the present invention examples

Follow these steps in order:

1. Prepare DPP-IV enzyme reaction buffer: 50 mM HEPES (pH = 7.8), 80 mM MgCl ₂ , 150 mM NaCl, 1% BSA, and place on ice until ready to use:

2. Dilute the test compound from 10 mM to 1 mM (100 times the final concentration) with DMSO, then perform a 3-fold serial dilution on a 96-well plate for a total of 11 concentrations. Add DMSO to the 12th well as a blank control. Then dilute 25-fold with enzyme reaction buffer to 4 times the final concentration for later use.

3. Thaw the DPP-IV enzyme reaction substrate H-Gly-Pro-AMC, dilute it to 160 μM (4 times the final concentration) with enzyme reaction buffer, and place it on ice until used;

4. Thaw rat plasma, dilute 100-fold with enzyme reaction buffer (2 times the final concentration) and place on ice until ready for use;

5. Add 5 μL of the test compound (4 times the final concentration) to a 384-well plate, then add 10 μL of rat plasma (2 times the final concentration) and centrifuge to mix.

6. Add 5 μL of enzyme reaction substrate H-Gly-Pro-AMC (4 times the final concentration), centrifuge to mix, and seal the 384-well plate with sealing film;

7. Incubate in an incubator (22-23°C) for 1 hour;

8. Use a FlexStationl3 (Molecular devices) microplate reader to read the fluorescence signal of the reaction: excitation at 380 nm and emission spectrum at 460 nm;

9. Generation of _IC50 values for DPP-IV inhibition by compounds: _IC50 values of compounds were calculated using GraFit6 software.

Table 1 DPP-IV inhibitory activity of the example compounds

Biological Experiment Example 2: Determination of Pharmacokinetic Parameters

The pharmacokinetic parameters of the compounds of the present application were determined using the following methods.

The study used healthy male adult rats aged 7-9 weeks. Each group of animals (3 male rats) received a single oral gavage of 5 mg/kg. The animals in the oral gavage group fasted overnight before the experiment, and the fasting period was from 10 hours before to 4 hours after the administration.

Blood was collected at 0.25, 0.5, 1, 2, 4, 6, 8, and 24 hours after administration. After isoflurane anesthesia using a small animal anesthesia machine, 0.3 mL of whole blood was collected from the retinal vein and placed in a heparinized tube. The sample was centrifuged at 4000 rpm for 5 minutes at 4°C. The plasma was transferred to a centrifuge tube and stored at -80°C until analysis.

Plasma samples were analyzed using a validated liquid chromatography-tandem mass spectrometry method (LC-MS/MS). Plasma concentration-time data for individual animals were analyzed using WinNonlin (Professional Edition, Version 6.3; Pharsight). A non-compartmental model was used for concentration analysis. Pharmacokinetic parameters for the compounds were calculated and are shown in Table 2 below:

Table 2 Pharmacokinetic parameters

Biological Experiment Example 3: Determination of _IC50 for Inhibition of CYP Enzymes

The following method was used to determine the IC ₅₀ of the inhibition of the compounds of the present application on the CYP enzyme system.

The human liver microsomes stored in a -80°C refrigerator were placed on ice to thaw. Immediately after thawing, 100 μL was taken and incubated in a constant temperature shaking box at 60°C and 100 rpm (1 hour). The remaining liver microsomes were immediately placed in a -80°C refrigerator for freezing. After 1 hour, 100 μL of inactivated liver microsomes were taken out and 400 μL of phosphate buffer was added to mix into a 4 mg/mL inactivated liver microsome solution; separately, the human liver microsomes stored in a -80°C refrigerator were placed on ice to thaw. Immediately after thawing, 100 μL was taken out and 400 μL of phosphate buffer was added to mix into a 4 mg/mL liver microsome solution. The positive control, test article, and negative control incubation mixtures were prepared in parallel according to Table 3 below:

Table 3 Incubation mixture of positive control, test article and negative control group

The mixture was placed in a constant temperature shaking box at 37°C and 100 rpm and incubated for 5 minutes.

Take 2.5 μL of the test article or positive control working solution (the negative control group is added with the test article working solution), add 91.5 μL of the incubation mixture and 6 μL of the NADPH solution and vortex to start the reaction. Place the solution in a constant temperature shaking box at 37°C and 100 rpm and incubate for the incubation time shown in Table 4 below:

Table 4 Incubation time

After incubation for the corresponding time, 200 μL of internal standard solution (the internal standard solution for CYP2C19 is a 100 ng/mL chloramphenicol solution in acetonitrile, and the other internal standard solutions are 250 ng/mL warfarin and 500 ng/mL propranolol solutions in acetonitrile) was added to terminate the reaction. The terminated sample was centrifuged at 12000 rpm for 10 minutes, and the supernatant was taken for sampling and detection.

Process the data using Analyst 1.4.2 or equivalent software. Check the integration to ensure that all peaks are properly integrated and adjust the integration parameters if necessary.

Quantitation of the analyte was defined as the ratio of the peak area of the analyte to the peak area of the internal standard. Analysis was performed using liquid chromatography-tandem mass spectrometry (LC-MS/MS). Parameters such as IC50 were calculated using Graphpad Prism (version 5.03) software. The results are shown in Table 5 below:

Table 5 Inhibition IC ₅₀ (μM) of the compound of Example 4 on CYP enzyme system

.

Biological Experiment Example 4: Metabolic Stability of Liver Microsomes

The following method was used to determine the metabolic stability of the compounds of the present application in liver microsomes.

8 μL of human liver microsomes (20 mg/mL), 20 μL of NADPH, and 368 μL of 0.1 M phosphate buffer were mixed and preincubated at 37°C for 5 minutes. 4 μL of analytical working solution (test article or positive control) was added, and at 37°C preincubation time of 0, 10, 20, 30, 45, and 60 minutes, 50 μL of the incubation solution was added to 150 μL of acetonitrile containing the internal standard (0.25 M warfarin). 4 μL of rat liver microsomes (20 mg/mL), 10 μL of NADPH, and 184 μL of 0.1 M phosphate buffer were mixed and preincubated at 37°C for 5 minutes. 2 μL of analytical working solution (test product or positive control) was added respectively, and 20 μL of the incubation solution was added to 180 μL of acetonitrile containing internal standard (0.25 M warfarin) at 37°C for 0, 10, 20, 30, 45 and 60 minutes. All samples were vortexed and centrifuged at 4000 rpm for 15 minutes, and then 150 μL of the supernatant was added to a 96-well sample plate. 5 μL was taken for detection by LC/MS/MS system. The analytical column was C18 1.7 μm 2.1×50 mm (Waters). A triple quadrupole mass spectrometer (API4000, AB Company) was used for detection. The ratio of the peak area of CT-1225 to the peak area of the internal standard was detected in positive ion mode. The half-life value is the ratio of the peak area of the test product/internal standard to time. The result data are shown in Table 6 below:

Table 6 Metabolic stability of the compound of Example 4 and the reference compound in liver microsomes

Biological Experiment Example 5: Inhibitory Effect of a Single Dose on Serum DPP-IV Activity in Ob/ob Mice

Thirty-six female ob/ob mice were randomly divided into six groups of six mice each: a model control group, a 1 mg/kg group receiving the compound of Example 4, a 3 mg/kg group receiving the compound of Example 4, a 10 mg/kg group receiving the compound of Example 4, a 30 mg/kg group receiving the compound of Example 4, and a positive control group receiving 30 mg/kg omarigliptin. Each group of mice was orally administered with varying doses of the compound of Example 4 or omarigliptin. The model control group was orally administered with 0.25% CMC-Na. Blood was collected before administration and at 2, 4, 10, 24, 34, 48, 58, 72, and 96 hours after administration. Serum was isolated and assayed for serum DPP-IV activity.

Serum DPP-IV activity assay method: Take 5 μL of serum sample, add 45 μL of 80 mM _MgCl2 buffer, mix, and preincubate at room temperature for 5 minutes. Add 10 μL of 0.1 mM reaction substrate Gly-Pro-7-AMC and 40 μL of buffer, protect from light, mix, and perform fluorescence measurement every 3 minutes (excitation wave 380 nm/emission wave 460 nm) until 18 minutes, for a total of 6 measurements. Based on the measurement results, after subtracting the blank background, a time-fluorescence value curve is plotted, and the slope is the activity value. The serum DPP-IV activity value at 0 h before administration is 100%. The specific activity value of serum DPP-IV at each time point after administration is calculated according to the following formula: Specific activity value (%) = activity value after administration/activity value before administration × 100%.

Results: After single oral administration of various doses of Example 4 to ob/ob mice, serum DPP-IV activity was significantly inhibited in a dose- and time-dependent manner. Within 10 hours after administration of 1 mg/kg of Example 4, the inhibition rate for serum DPP-IV activity in mice was greater than 70%. Within 24 hours after administration of 3 mg/kg of Example 4, the inhibition rate was greater than 70%. Within 34 hours after administration of 10 mg/kg of Example 4, the inhibition rate was greater than 70%. After administration of 30 mg/kg of Example 4, the inhibition rate remained above 70% for 72 hours. In the positive control group, the inhibition rate for serum DPP-IV activity in mice was greater than 70% within 34 hours after administration of 30 mg/kg of Example 4.

Claims (8)

1. The compound represented by formula V or a pharmaceutically acceptable salt thereof: The substitution positions of _R5 are as shown in _R5a and _R5b in structural formula VII, or as shown in _R5a in structural _formula VIII: in, X is selected from O; Y is selected from CH; _R1 is selected from _-NH2 or -OH; _R2 and _R3 are selected from H; Each _R4 is independently selected from F, Cl, Br, _-NH2 , or -OH; R _5a is selected from -NHR ₇ or -SO ₂ R ₈ ; R _5b is selected from F, Cl, or Br; R ₇ is selected from -SO ₂ R ₈ ; Each _R8 is independently selected from -OH, _-NH2 , _C1-6 alkyl, or _C3-6 cycloalkyl; o can be selected from 1, 2, or 3; p is selected from 1 or 2. 2. The compound of claim 1 or a pharmaceutically acceptable salt thereof, wherein each _R8 is independently selected from -OH, _-NH2 , methyl, ethyl, propyl, butyl, _C3 cycloalkyl, _C4 cycloalkyl or _C5 cycloalkyl. 3. The compound of claim 1 or a pharmaceutically acceptable salt thereof, wherein o is 2. 4. The compound of claim 1 or a pharmaceutically acceptable salt thereof, wherein the substitution positions of _R4 are as shown by _R4a and _R4b in structural formula VI: _R4a and _R4b are each independently selected from F, Cl, Br, _-NH2 or -OH, and the definitions of other substituents are the same as those in claim 1. 5. The compound of claim 1 or a pharmaceutically acceptable salt thereof, wherein the compound is selected from: 6. A pharmaceutical composition comprising a therapeutically effective amount of the compound of any one of claims 1-5 or a pharmaceutically acceptable salt, tautomer, or pharmaceutically acceptable carrier thereof. 7. Use of the compound of any one of claims 1-5 or a pharmaceutically acceptable salt thereof or the pharmaceutical composition of claim 6 in the preparation of a medicament for treating and/or preventing diseases and conditions that benefit from DPP-IV inhibition. 8. The use according to claim 7, wherein the diseases and conditions that benefit from DPP-IV inhibition are selected from type 2 diabetes.