WO2017071606A1 - Medical use of hydroxyl purine compound - Google Patents

Abstract

Disclosed in the present invention is an application of a series of hydroxyl purine compounds in the preparation of a drug for treating or preventing liver disease, and particularly disclosed is an application of a compound shown in formula (I), or a tautomer or pharmaceutically acceptable salt thereof in the preparation of a drug for treating liver disease.

Description

Medical use of hydroxy steroids Technical field

The present invention relates to the use of a series of hydroxy steroids in the preparation of a medicament for treating liver diseases, in particular to the use of a compound of the formula (I), a tautomer thereof or a pharmaceutically acceptable salt thereof for the preparation of a medicament for treating liver diseases .

technical background

Phosphodiesterase (PDE) catalyzes the hydrolysis of cyclized nucleotides cGMP and cAMP, and regulates various physiological responses by controlling the intramolecular concentrations of these two important secondary signaling factors. Cyclization of nucleotides cGMP and cAMP intramolecular Abnormal regulation is the cause of many diseases. Several drugs have been used to improve and treat diseases by inhibiting PDE activity, such as PDE5 inhibitors for pulmonary hypertension and PDE4 inhibitors for arthritis caused by psoriasis. There are eleven major classes of phosphodiesterase genes, each of which can express several subtypes. There are more than 100 PDE subtypes in total. Different subtypes have different structures, different tissue distributions, and circularized nuclei. The activities of cGMP and cAMP of glucosinolates are also very different, and the physiological functions of regulation are also very different.

PDE2 phosphodiesterase can catalyze the hydrolysis of cyclized nucleotides cGMP and cAMP, while cAMP activity is regulated by cGMP, which plays a key role in the balance of cGMP and cAMP functions in cells. PDE2 is widely expressed in human tissues and is mainly distributed in the heart, central nervous system, liver, adrenal gland, endothelial cells, and platelets. PDE2 is involved in the regulation of various physiological activities, such as central learning, memory and cognition, maintaining the basic rhythm of the heart, smooth muscle and endothelial cells, permeability of endothelial cells, and regulating inflammation. PDE2 knockout mice directly cause embryonic death. It can be used in a variety of central, cardiovascular, and inflammatory responses by inhibiting PDE2 activity.

Non-selective PDE inhibitory activities of various natural and synthetic terpenoids have long been discovered, such as caffeine, theophylline, pentoxifylline and the like. Pentoxifylline (PDE2 activity) is clinically approved for lower limb paralysis caused by peripheral vascular occlusion. The main role is to reduce blood viscosity, increase red blood cell deformation, and inhibit platelet aggregation. Novel highly selective PDE2 inhibitors have also been reported to control endothelial cell division and angiogenesis, and to improve central cognitive impairment. However, the development and application of the new novel selective PDE2 inhibitors are still very limited, and the discovery and application of new PDE2 inhibitors have broad prospects.

Tumor necrosis factor alpha (TNF-α) is a cytokine with many biological activities, which has an important impact on the occurrence, development and treatment of various diseases, especially immune and inflammation related diseases. TNF-α is mainly produced by monocytes and macrophage cell lines, and participates in the body's immune regulation and cytokine network coordination. Under normal circumstances, TNF-α plays an important role in immune defense and immune surveillance, but in some cases it has an adverse effect. Studies have shown that overexpression of TNF-α can induce the expression of proinflammatory cytokines such as interleukon-l (IL-1), IL-6, increase endothelial cell permeability, up-regulate adhesion molecule expression, and activate neutrality. Leukocytes and eosinophils, and induce the secretion of acute phase substances and tissue degrading enzymes by bone synovial cells and chondrocytes to promote the occurrence of inflammation. These pathological reactions play a very important role in the development of many immune-mediated inflammatory diseases (IMID), such as rheumatoid arthritis (RA), psoriatic arthritis (psoriatic arthritis). , PsA), ankylosing spondylitis (AS), inflammatory bowel disease (IBD), juvenile chronic arthritis (JCA), and vasculitis. Studies have shown that TNF-α is an ideal target for many of the above IMIDs, and for some diseases caused by chronic chronic inflammatory damage, such as steatohepatitis, chronic obstructive pneumonia, etc., using TNF-α inhibitors (TNF-α inhibitors) And excess TNF-α is also an effective prevention and treatment route. TNF-α monoclonal antibody has been clinically proven to inhibit TNF-α as a very effective means of treating the aforementioned inflammation-related diseases. PDE2 can be mechanically By regulating the expression of TNF-α, it is possible to control the level of TNF-α by regulating PDE2 activity, thereby achieving control of the inflammatory response.

Summary of the invention

The present invention provides a compound of the formula (I), a tautomer thereof or a pharmaceutically acceptable salt thereof for use in the preparation of a medicament for treating or preventing liver disease,

among them,

替换为

具体替换为

Structural unit

Replace with

Specifically replaced with

L _{11 is} selected from the group consisting of empty, C(R)(R');

R, R' are each independently selected from H, halogen, OH, NH ₂ , CN, optionally substituted 1 to 6 alkyl or heteroalkyl;

Optionally, R, R' may be cyclized to a 3-6 membered cycloalkyl or heterocycloalkyl group;

A is empty or is selected from optionally substituted cycloalkyl, heterocycloalkyl, aryl, heteroaryl;

L _{12 is} selected from an optionally substituted 1 to 6 membered alkyl or heteroalkyl group;

R _{1 is} selected from an optionally substituted 1 to 6 membered alkyl group, a 3 to 6 membered cycloalkyl group or a heteroalkyl group;

"Hetero" means N, O, S, C(=O), S(=O), S(=O) ₂ , and the number of heteroatoms on each group is selected from 1, 2, 3 or 4.

In some aspects of the invention, the liver disease is selected from the group consisting of nonalcoholic fatty liver and liver fibrosis.

In some embodiments of the invention, the substituents in the above R, R', A, L ₁₂ and R ₁ are each independently selected from the group consisting of halogen, OH, NH ₂ , CN, optionally substituted 1 to 6 alkyl, 3 A 6-membered cycloalkyl or heteroalkyl group, the number of each group substituent being independently selected from 1, 2 or 3.

In some embodiments of the invention, the substituents in the above R, R', A, L ₁₂ and R ₁ are each independently selected from the group consisting of halogen, CF ₃ , CN, OH, Me, Et, n-propyl, isopropyl, and ring. Propyl,

In some aspects of the invention, R and R' are each independently selected from the group consisting of H, Me, CF ₃ , and Et.

In some embodiments of the invention, the above L _{11 is} selected from

In some embodiments of the invention, the above A is selected from the group consisting of: a 3 to 12 membered cycloalkyl or heterocycloalkyl group, a 5 to 12 membered aryl group or a heteroaryl group.

In some embodiments of the invention, the above A is selected from the group consisting of: optionally substituted: cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, pentylpentyl, Phenyl, pyridyl, pyrazinyl, oxazolyl, isoxazolyl, thiazolyl, bicyclo[1.1.1]pentane, or a bicyclic ring or spiro ring selected from any two of the above groups Base or a cyclized group.

In some aspects of the invention, the above A is selected from the group consisting of:

In some aspects of the invention, the above A is selected from the group consisting of

In some embodiments of the invention, the above L _{12 is} selected from the group consisting of methylene,

环丙基、

In some embodiments of the invention, the above R _{1 is} selected from the group consisting of Me, CHF ₂ , CF ₃ , Et, CH ₂ CF ₃ , isopropyl,

Cyclopropyl,

The invention is selected from the group consisting of the following compounds in the preparation of a medicament for treating or preventing liver diseases:

Further, the present invention is selected from the group consisting of the following compounds in the preparation of a medicament for treating or preventing liver diseases:

Related definition

Unless otherwise stated, the following terms and phrases as used herein are intended to have the following meanings. A particular term or phrase should not be considered undefined or unclear without a particular definition, but should be understood in the ordinary sense. When a trade name appears in this document, it is intended to refer to its corresponding commodity or its active ingredient.

The term "pharmaceutically acceptable" as used herein is intended to mean that those compounds, materials, compositions and/or dosage forms are within the scope of sound medical judgment and are suitable for use in contact with human and animal tissues. Without excessive toxicity, irritation, allergic reactions or other problems or complications, commensurate with a reasonable benefit/risk ratio.

The term "pharmaceutically acceptable salt" refers to a salt of a compound of the invention prepared from a compound having a particular substituent found in the present invention and a relatively non-toxic acid or base. When a relatively acidic functional group is contained in the compound of the present invention, a base addition salt can be obtained by contacting a neutral amount of such a compound with a sufficient amount of a base in a neat solution or a suitable inert solvent. Pharmaceutically acceptable base addition salts include sodium, potassium, calcium, ammonium, organic ammonia or magnesium salts or similar salts. When a relatively basic functional group is contained in the compound of the present invention, an acid addition salt can be obtained by contacting a neutral form of such a compound with a sufficient amount of an acid in a neat solution or a suitable inert solvent. Examples of pharmaceutically acceptable acid addition salts include inorganic acid salts including, for example, hydrochloric acid, hydrobromic acid, nitric acid, carbonic acid, hydrogencarbonate, phosphoric acid, monohydrogen phosphate, dihydrogen phosphate, sulfuric acid, Hydrogen sulfate, hydroiodic acid, phosphorous acid, etc.; and an organic acid salt, such as acetic acid, propionic acid, isobutyric acid, maleic acid, malonic acid, benzoic acid, succinic acid, suberic acid, Similar acids such as fumaric acid, lactic acid, mandelic acid, phthalic acid, benzenesulfonic acid, p-toluenesulfonic acid, citric acid, tartaric acid, and methanesulfonic acid; and salts of amino acids (such as arginine, etc.) And salts of organic acids such as glucuronic acid (see Berge et al., "Pharmaceutical Salts", Journal of Pharmaceutical Science 66: 1-19 (1977)). Certain specific compounds of the invention contain both basic and acidic functional groups which can be converted to any base or acid addition salt.

Preferably, the salt is contacted with a base or acid in a conventional manner, and the parent compound is separated, thereby regenerating the neutral form of the compound. The parent form of the compound differs from the form of its various salts by certain physical properties, such as differences in solubility in polar solvents.

As used herein, a "pharmaceutically acceptable salt" is a derivative of a compound of the invention wherein the parent compound is modified by salt formation with an acid or with a base. Examples of pharmaceutically acceptable salts include, but are not limited to, inorganic or organic acid salts of bases such as amines, alkali metal or organic salts of acid groups such as carboxylic acids, and the like. Pharmaceutically acceptable salts include the conventional non-toxic salts or quaternary ammonium salts of the parent compound, for example salts formed from non-toxic inorganic or organic acids. Conventional non-toxic salts include, but are not limited to, those derived from inorganic acids and organic acids selected from the group consisting of 2-acetoxybenzoic acid, 2-hydroxyethanesulfonic acid, acetic acid, ascorbic acid, Benzenesulfonic acid, benzoic acid, Bicarbonate, carbonic acid, citric acid, edetic acid, ethane disulfonic acid, ethanesulfonic acid, fumaric acid, glucoheptose, gluconic acid, glutamic acid, glycolic acid, hydrobromic acid, hydrochloric acid, hydrogen Iodate, hydroxyl, hydroxynaphthalene, isethionate, lactic acid, lactose, dodecylsulfonic acid, maleic acid, malic acid, mandelic acid, methanesulfonic acid, nitric acid, oxalic acid, pamoic acid, pantothenic acid, Phenylacetic acid, phosphoric acid, polygalacturonan, propionic acid, salicylic acid, stearic acid, acrylic acid, succinic acid, sulfamic acid, p-aminobenzenesulfonic acid, sulfuric acid, tannin, tartaric acid and p-toluenesulfonic acid.

The pharmaceutically acceptable salts of the present invention can be synthesized from the parent compound containing an acid group or a base by conventional chemical methods. In general, such salts are prepared by reacting these compounds in water or an organic solvent or a mixture of the two via a free acid or base form with a stoichiometric amount of a suitable base or acid. Generally, a nonaqueous medium such as ether, ethyl acetate, ethanol, isopropanol or acetonitrile is preferred.

In addition to the form of the salt, the compounds provided herein also exist in the form of prodrugs. Prodrugs of the compounds described herein are readily chemically altered under physiological conditions to convert to the compounds of the invention. Furthermore, prodrugs can be converted to the compounds of the invention by chemical or biochemical methods in an in vivo setting.

Certain compounds of the invention may exist in unsolvated or solvated forms, including hydrated forms. In general, the solvated forms are equivalent to the unsolvated forms and are included within the scope of the invention.

Certain compounds of the invention may have asymmetric carbon atoms (optical centers) or double bonds. Racemates, diastereomers, geometric isomers and individual isomers are included within the scope of the invention.

表示一个立体中心的绝对构型，用

表示一个立体中心的相对构型。当本文所述化合物含有烯属双键或其它几何不对称中心，除非另有规定，它们包括E、Z几何异构体。同样地，所有的互变异构形式均包括在本发明的范围之内。Wedge and dashed keys unless otherwise stated

Represents the absolute configuration of a stereocenter,

Represents the relative configuration of a stereocenter. When the compounds described herein contain olefinic double bonds or other centers of geometric asymmetry, they include the E and Z geometric isomers unless otherwise specified. Likewise, all tautomeric forms are included within the scope of the invention.

The compounds of the invention may exist in specific geometric or stereoisomeric forms. All such compounds are contemplated by the present invention, including cis and trans isomers, (-)- and (+)-enantiomers, (R)- and (S)-enantiomers, diastereoisomers , (D)-isomer, (L)-isomer, and racemic mixtures thereof and other mixtures, such as enantiomerically or diastereomeric enriched mixtures, all of which belong to the present Within the scope of the invention. Additional asymmetric carbon atoms may be present in the substituents such as alkyl groups. All such isomers, as well as mixtures thereof, are included within the scope of the invention.

The optically active (R)- and (S)-isomers as well as the D and L isomers can be prepared by chiral synthesis or chiral reagents or other conventional techniques. If an enantiomer of a compound of the invention is desired, it can be prepared by asymmetric synthesis or by derivatization with a chiral auxiliary wherein the resulting mixture of diastereomers is separated and the auxiliary group cleaved to provide pure The desired enantiomer. Alternatively, when a molecule contains a basic functional group (e.g., an amino group) or an acidic functional group (e.g., a carboxyl group), a salt of a diastereomer is formed with a suitable optically active acid or base, and then by a conventional method known in the art. The diastereomers are resolved and the pure enantiomer is recovered. Furthermore, the separation of enantiomers and diastereomers is generally accomplished by the use of chromatography using a chiral stationary phase, optionally in combination with chemical derivatization (eg, formation of an amino group from an amine). Formate).

The compounds of the present invention may contain unnatural proportions of atomic isotopes on one or more of the atoms that make up the compound. For example, radiolabeled compounds can be used, such as tritium ⁽³ H), iodine -125 ⁽¹²⁵ I) or ^C-14 (14 C). Alterations of all isotopic compositions of the compounds of the invention, whether radioactive or not, are included within the scope of the invention.

The term "pharmaceutically acceptable carrier" refers to any formulation or carrier medium that is capable of delivering an effective amount of an active substance of the present invention, does not interfere with the biological activity of the active substance, and has no toxic side effects to the host or patient, including water, oil, Vegetables and minerals, cream base, wash Agent matrix, ointment base, and the like. These bases include suspending agents, tackifiers, transdermal enhancers and the like. Their formulations are well known to those skilled in the cosmetic or topical pharmaceutical arts. For additional information on vectors, reference is made to Remington: The Science and Practice of Pharmacy, 21st Ed., Lippincott, Williams & Wilkins (2005), the contents of which are hereby incorporated by reference.

The term "excipient" generally refers to the carrier, diluent and/or vehicle required to formulate an effective pharmaceutical composition.

The term "effective amount" or "therapeutically effective amount" with respect to a pharmaceutical or pharmacologically active agent refers to a sufficient amount of a drug or agent that is non-toxic but that achieves the desired effect. For oral dosage forms in the present invention, an "effective amount" of an active substance in a composition refers to the amount required to achieve the desired effect when used in combination with another active substance in the composition. The determination of the effective amount will vary from person to person, depending on the age and general condition of the recipient, and also on the particular active substance, and a suitable effective amount in a case can be determined by one skilled in the art based on routine experimentation.

The term "active ingredient", "therapeutic agent", "active substance" or "active agent" refers to a chemical entity that is effective in treating a target disorder, disease or condition.

"Optional" or "optionally" means that the subsequently described event or condition may, but is not necessarily, to occur, and that the description includes instances in which the event or condition occurs and instances in which the event or condition does not occur.

The term "substituted" means that any one or more hydrogen atoms on a particular atom are replaced by a substituent, including variants of heavy hydrogen and hydrogen, as long as the valence of the particular atom is normal and the substituted compound is stable. . When the substituent is a keto group (ie, =0), it means that two hydrogen atoms are substituted. Ketone substitution does not occur on the aryl group. The term "optionally substituted" means that it may or may not be substituted, and unless otherwise specified, the kind and number of substituents may be arbitrary on the basis of chemically achievable.

When any variable (eg, R) occurs more than once in the composition or structure of a compound, its definition in each case is independent. Thus, for example, if a group is substituted with 0-2 R, the group may optionally be substituted with at most two R, and each case has an independent option. Furthermore, combinations of substituents and/or variants thereof are permissible only if such combinations result in stable compounds.

When the number of one linking group is 0, such as -(CRR) ₀ -, it indicates that the linking group is a single bond.

When one of the variables is selected from a single bond, it means that the two groups to which it is attached are directly linked. For example, when L represents a single bond in A-L-Z, the structure is actually A-Z.

When a substituent is vacant, it means that the substituent is absent. For example, when X is vacant in A-X, the structure is actually A.

或

表示其可在环己基或者环己二烯上的任意一个位置发生取代。When a bond of a substituent can be cross-linked to two atoms on a ring, the substituent can be bonded to any atom on the ring. When the recited substituents do not indicate which atom is attached to a compound included in the chemical structural formula including but not specifically mentioned, such a substituent may be bonded through any atomic phase thereof. Combinations of substituents and/or variants thereof are permissible only if such combinations result in stable compounds. For example, a structural unit

or

It is indicated that it can be substituted at any position on the cyclohexyl or cyclohexadiene.

Unless otherwise specified, the term "halo" or "halogen", by itself or as part of another substituent, denotes a fluorine, chlorine, bromine or iodine atom. Further, the term "haloalkyl" is intended to include both monohaloalkyl and polyhaloalkyl. For example, the term "halo(C ₁ -C ₄ )alkyl" is intended to include, but is not limited to, trifluoromethyl, 2,2,2-trifluoroethyl, 4-chlorobutyl, 3-bromopropyl, and the like. Wait.

Examples of haloalkyl groups include, but are not limited to, trifluoromethyl, trichloromethyl, pentafluoroethyl, and pentachloroethyl. "Alkoxy" represents the above alkyl group having a specified number of carbon atoms attached through an oxygen bridge. The C _1-6 alkoxy group includes a C ₁ , C ₂ , C ₃ , C ₄ , C ₅ and C ₆ alkoxy groups. Examples of alkoxy groups include, but are not limited to, methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, sec-butoxy, tert-butoxy, n-pentyloxy and S- Pentyloxy. "Cycloalkyl" includes saturated cyclic groups such as cyclopropyl, cyclobutyl or cyclopentyl. The 3-7 cycloalkyl group includes C ₃ , C ₄ , C ₅ , C ₆ and C ₇ cycloalkyl groups. "Alkenyl" includes hydrocarbon chains in a straight or branched configuration wherein one or more carbon-carbon double bonds, such as vinyl and propylene groups, are present at any stable site on the chain.

The term "halo" or "halogen" refers to fluoro, chloro, bromo and iodo.

Unless otherwise specified, the term "hetero" denotes a hetero atom or a hetero atomic group (ie, a radical containing a hetero atom), including atoms other than carbon (C) and hydrogen (H), and radicals containing such heteroatoms, including, for example, oxygen (O). ), nitrogen (N), sulfur (S), silicon (Si), germanium (Ge), aluminum (Al), boron (B), -O-, -S-, =O, =S, -C (= O) O-, -C(=O)-, -C(=S)-, -S(=O), -S(=O) ₂ -, and optionally substituted -C(=O)N (H)-, -N(H)-, -C(=NH)-, -S(=O) ₂ N(H)- or -S(=O)N(H)-.

Unless otherwise specified, "ring" means substituted or unsubstituted cycloalkyl, heterocycloalkyl, cycloalkenyl, heterocycloalkenyl, cycloalkynyl, heterocycloalkynyl, aryl or heteroaryl. So-called rings include single rings, interlocking rings, spiral rings, parallel rings or bridge rings. The number of atoms on the ring is usually defined as the number of elements of the ring. For example, "5 to 7-membered ring" means 5 to 7 atoms arranged in a circle. Unless otherwise specified, the ring optionally contains from 1 to 3 heteroatoms. Thus, "5- to 7-membered ring" includes, for example, phenylpyridine and piperidinyl; on the other hand, the term "5- to 7-membered heterocycloalkyl ring" includes pyridyl and piperidinyl, but does not include phenyl. The term "ring" also includes ring systems containing at least one ring, each of which "ring" independently conforms to the above definition.

Unless otherwise specified, the term "heterocycle" or "heterocyclyl" means a stable monocyclic, bicyclic or tricyclic ring containing a hetero atom or a heteroatom group which may be saturated, partially unsaturated or unsaturated ( Aromatic) which comprise a carbon atom and 1, 2, 3 or 4 ring heteroatoms independently selected from N, O and S, wherein any of the above heterocycles may be fused to a phenyl ring to form a bicyclic ring. The nitrogen and sulfur heteroatoms can be optionally oxidized (i.e., NO and S(O)p, p is 1 or 2). The nitrogen atom can be substituted or unsubstituted (i.e., N or NR, wherein R is H or other substituents as already defined herein). The heterocyclic ring can be attached to the side groups of any hetero atom or carbon atom to form a stable structure. If the resulting compound is stable, the heterocycles described herein can undergo substitutions at the carbon or nitrogen sites. The nitrogen atom in the heterocycle is optionally quaternized. A preferred embodiment is that when the total number of S and O atoms in the heterocycle exceeds 1, these heteroatoms are not adjacent to each other. Another preferred embodiment is that the total number of S and O atoms in the heterocycle does not exceed one. The term "aromatic heterocyclic group" or "heteroaryl" as used herein means a stable 5, 6, or 7 membered monocyclic or bicyclic or aromatic ring of a 7, 8, 9 or 10 membered bicyclic heterocyclic group, It contains carbon atoms and 1, 2, 3 or 4 ring heteroatoms independently selected from N, O and S. The nitrogen atom can be substituted or unsubstituted (i.e., N or NR, wherein R is H or other substituents as already defined herein). The nitrogen and sulfur heteroatoms can be optionally oxidized (i.e., NO and S(O)p, p is 1 or 2). It is worth noting that the total number of S and O atoms on the aromatic heterocycle does not exceed one. Bridged rings are also included in the definition of heterocycles. A bridged ring is formed when one or more atoms (ie, C, O, N, or S) join two non-adjacent carbon or nitrogen atoms. Preferred bridged rings include, but are not limited to, one carbon atom, two carbon atoms, one nitrogen atom, two nitrogen atoms, and one carbon-nitrogen group. It is worth noting that a bridge always converts a single ring into a three ring. In the bridged ring, a substituent on the ring can also be present on the bridge.

Examples of heterocyclic compounds include, but are not limited to, acridinyl, octanoyl, benzimidazolyl, benzofuranyl, benzofuranylfuranyl, benzindenylphenyl, benzoxazolyl, benzimidin Oxazolinyl, benzothiazolyl, benzotriazolyl, benzotetrazolyl, benzisoxazolyl, benzisothiazolyl, benzimidazolyl, oxazolyl, 4aH-carbazolyl, Porphyrin, chroman, chromene, porphyrin-decahydroquinolinyl, 2H, 6H-1,5,2-dithiazinyl, dihydrofuro[2,3-b] Tetrahydrofuranyl, furyl, furfuryl, imidazolidinyl, imidazolinyl, imidazolyl, 1H-carbazolyl, nonenyl, indanyl, mesoindolyl, fluorenyl, 3H-indole Sulfhydryl, isatino, isobenzofuranyl, isodecyl, isoindoline, isoquinolyl, isothiazolyl, isoxazolyl, methylenedioxyphenyl, morpholinyl, Naphthyridinyl, octahydroisoquinolinyl, oxadiazolyl, 1,2,3-oxadiazolyl, 1,2,4-oxadiazolyl, 1,2,5-oxadiazolyl, 1 , 3,4-oxadiazolyl, oxazolidine Base, oxazolyl, hydroxymethyl, pyrimidinyl, phenanthryl, phenanthroline, phenazine, phenothiazine, benzoxanthyl, phenoloxazinyl, pyridazinyl, piperazinyl, piperazine Pyridyl, piperidinone, 4-piperidinone, piperonyl, pteridinyl, fluorenyl, pyranyl, pyrazinyl, pyrazolidinyl, pyrazolinyl, pyrazolyl, pyridazinyl , pyridooxazole, pyridoimidazole, pyridylthiazole, pyridyl, pyrrolidinyl, pyrrolinyl, 2H-pyrrolyl, pyrrolyl, quinazolinyl, quinolyl, 4H-quinazinyl, quinacrid Polinyl, quinuclidinyl, tetrahydrofuranyl, tetrahydroisoquinolinyl, tetrahydroquinolyl, tetrazolyl, 6H-1,2,5-thiadiazinyl, 1,2,3-thiadiazole 1,1,4-thiadiazolyl, 1,2,5-thiadiazolyl, 1,3,4-thiadiazolyl, thioxyl, thiazolyl, isothiazolylthiophenyl, thienyl , thienooxazolyl, thienothiazolyl, thienoimidazolyl, triazinyl, 1,2,3-triazolyl, 1,2,4-triazolyl, 1,2,5-triazolyl 1,3,4-triazolyl and xanthene. Also included are fused ring and spiro compounds.

Unless otherwise specified, the term "hydrocarbyl" or its subordinate concept (such as alkyl, alkenyl, alkynyl, aryl, etc.), by itself or as part of another substituent, is meant to be straight-chain, branched or cyclic. The hydrocarbon atom group or a combination thereof may be fully saturated (such as an alkyl group), a unit or a polyunsaturated (such as an alkenyl group, an alkynyl group, an aryl group), may be monosubstituted or polysubstituted, and may be monovalent (such as Methyl), divalent (such as methylene) or polyvalent (such as methine), may include divalent or polyvalent radicals with a specified number of carbon atoms (eg, C ₁ -C ₁₂ represents 1 to 12 carbons) , C _{1-12 is} selected from C ₁ , C ₂ , C ₃ , C ₄ , C ₅ , C ₆ , C ₇ , C ₈ , C ₉ , C ₁₀ , C ₁₁ and C ₁₂ ; C _{3-12 is} selected from C ₃ , C ₄ , C ₅ , C ₆ , C ₇ , C ₈ , C ₉ , C ₁₀ , C ₁₁ and C ₁₂ .). "Hydrocarbyl" includes, but is not limited to, aliphatic hydrocarbyl groups including chain and cyclic, including but not limited to alkyl, alkenyl, alkynyl groups including, but not limited to, 6-12 members. An aromatic hydrocarbon group such as benzene, naphthalene or the like. In some embodiments, the term "hydrocarbyl" means a straight or branched chain radical or a combination thereof, which may be fully saturated, unitary or polyunsaturated, and may include divalent and multivalent radicals. Examples of saturated hydrocarbon radicals include, but are not limited to, methyl, ethyl, n-propyl, isopropyl, n-butyl, t-butyl, isobutyl, sec-butyl, isobutyl, cyclohexyl, (cyclohexyl). A homolog or isomer of a methyl group, a cyclopropylmethyl group, and an atomic group such as n-pentyl, n-hexyl, n-heptyl, n-octyl. The unsaturated hydrocarbon group has one or more double or triple bonds, and examples thereof include, but are not limited to, a vinyl group, a 2-propenyl group, a butenyl group, a crotyl group, a 2-isopentenyl group, and a 2-(butadienyl group). , 2,4-pentadienyl, 3-(1,4-pentadienyl), ethynyl, 1- and 3-propynyl, 3-butynyl, and higher homologs and isomers body.

Unless otherwise specified, the term "heterohydrocarbyl" or its subordinate concept (such as heteroalkyl, heteroalkenyl, heteroalkynyl, heteroaryl, etc.), by itself or in combination with another term, means a stable straight chain, branched chain. Or a cyclic hydrocarbon radical or a combination thereof having a number of carbon atoms and at least one heteroatom. In some embodiments, the term "heterohydrocarbyl" by itself or in conjunction with another term denotes a stable straight chain, branched hydrocarbon radical or combination thereof, having a number of carbon atoms and at least one heteroatom. In a typical embodiment, the heteroatoms are selected from the group consisting of B, O, N, and S, wherein the nitrogen and sulfur atoms are optionally oxidized, and the nitrogen heteroatoms are optionally quaternized. The heteroatoms B, O, N and S can be located at any internal position of the heterohydrocarbyl group (including where the hydrocarbyl group is attached to the rest of the molecule). Examples include, but are not limited to, -CH ₂ -CH ₂ -O-CH ₃ , -CH ₂ -CH ₂ -NH-CH ₃ , -CH ₂ -CH ₂ -N(CH ₃ )-CH ₃ , -CH ₂ -S -CH ₂ -CH ₃ , -CH ₂ -CH ₂ , -S(O)-CH ₃ , -CH ₂ -CH ₂ -S(O) ₂ -CH ₃ , -CH=CH-O-CH ₃ ,- CH ₂ -CH=N-OCH ₃ and -CH=CH-N(CH ₃ )-CH ₃ . Up to two heteroatoms may be consecutive, for example, -CH ₂ -NH-OCH _3.

The terms "alkoxy", "alkylamino" and "alkylthio" (or thioalkoxy) are customary expressions and refer to those alkane which are attached to the remainder of the molecule through an oxygen atom, an amino group or a sulfur atom, respectively. Base group.

Unless otherwise specified, the term "alkyl" is used to denote a straight or branched saturated hydrocarbon group, which may be monosubstituted (eg, -CH ₂ F) or polysubstituted (eg, -CF ₃ ), and may be monovalent (eg, Methyl), divalent (such as methylene) or polyvalent (such as methine). Examples of the alkyl group include methyl (Me), ethyl (Et), propyl (e.g., n-propyl and isopropyl), butyl (e.g., n-butyl, isobutyl, s-butyl). , t-butyl), pentyl (eg, n-pentyl, isopentyl, neopentyl) and the like.

Unless otherwise specified, the term "halo" or "halogen", by itself or as part of another substituent, denotes a fluorine, chlorine, bromine or iodine atom. Further, the term "haloalkyl" is intended to include both monohaloalkyl and polyhaloalkyl. For example, the term "halo(C ₁ -C ₄ )alkyl" is intended to include, but is not limited to, trifluoromethyl, 2,2,2-trifluoroethyl, 4-chlorobutyl, 3-bromopropyl, and the like. Wait. Unless otherwise specified, examples of haloalkyl include, but are not limited to, trifluoromethyl, trichloromethyl, pentafluoroethyl, and pentachloroethyl.

The above-described alkyl groups having the specified number of carbon atoms, "alkoxy" represents attached through an oxygen bridge, unless otherwise specified, C _1-6 alkoxy groups include _{C 1, C 2, C 3} , C 4, C 5 , and C ₆ alkoxy groups. Examples of alkoxy groups include, but are not limited to, methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, sec-butoxy, tert-butoxy, n-pentyloxy and S- Pentyloxy.

Unless otherwise specified, the term "cycloalkyl", "heterocycloalkyl" or its subordinate concept (such as aryl, heteroaryl, cycloalkyl, heterocycloalkyl, cycloalkenyl, heterocycloalkenyl, cycloalkynyl) A heterocyclic alkynyl group, etc., by itself or in combination with other terms, denotes a cyclized "hydrocarbyl group" or "heterohydrocarbyl group", respectively. Further, in the case of a heterohydrocarbyl group or a heterocycloalkyl group (such as a heteroalkyl group or a heterocycloalkyl group), a hetero atom may occupy a position at which the hetero ring is attached to the rest of the molecule. Examples of cycloalkyl groups include, but are not limited to, cyclopentyl, cyclohexyl, 1-cyclohexenyl, 3-cyclohexenyl, cycloheptyl, and the like. Non-limiting examples of heterocyclic groups include 1-(1,2,5,6-tetrahydropyridyl), 1-piperidinyl, 2-piperidinyl, 3-piperidinyl, 4-morpholinyl, 3-morpholinyl, tetrahydrofuran-2-yl, tetrahydrofuran-3-yl, tetrahydrothiophen-2-yl, tetrahydrothiophen-3-yl, 1-piperazinyl and 2-piperazinyl.

Unless otherwise specified, the term "aryl" denotes a polyunsaturated, aromatic hydrocarbon substituent which may be monosubstituted, disubstituted or polysubstituted, may be monovalent, divalent or polyvalent, it may be monocyclic or Polycyclic (preferably 1 to 3 rings) which are fused together or covalently linked. The term "heteroaryl" refers to an aryl (or ring) containing one to four heteroatoms. In an illustrative example, the heteroatoms are selected from the group consisting of B, N, O, and S, wherein the nitrogen and sulfur atoms are optionally oxidized, and the nitrogen atom is optionally quaternized. A heteroaryl group can be attached to the remainder of the molecule through a heteroatom. Non-limiting examples of aryl or heteroaryl groups include phenyl, 1-naphthyl, 2-naphthyl, 4-biphenyl, 1-pyrrolyl, 2-pyrrolyl, 3-pyrrolyl, 3-pyridyl Azyl, 2-imidazolyl, 4-imidazolyl, pyrazinyl, 2-oxazolyl, 4-oxazolyl, 2-phenyl-4-oxazolyl, 5-oxazolyl, 3-isoxan Azyl, 4-isoxazolyl, 5-isoxazolyl, 2-thiazolyl, 4-thiazolyl, 5-thiazolyl, 2-furyl, 3-furyl, 2-thienyl, 3-thiophene , 2-pyridyl, 3-pyridyl, 4-pyridyl, 2-pyrimidinyl, 4-pyrimidinyl, 5-benzothiazolyl, indolyl, 2-benzimidazolyl, 5-indenyl, 1-isoquinolyl, 5-isoquinolinyl, 2-quinoxalinyl, 5-quinoxalinyl, 3-quinolyl and 6-quinolinyl. The substituents of any of the above aryl and heteroaryl ring systems are selected from the group of acceptable substituents described below.

For simplicity, aryl groups, when used in conjunction with other terms (eg, aryloxy, arylthio, aralkyl), include aryl and heteroaryl rings as defined above. Thus, the term "aralkyl" is intended to include those radicals to which an aryl group is attached to an alkyl group (eg, benzyl, phenethyl, pyridylmethyl, and the like), including wherein the carbon atom (eg, methylene) has been, for example, oxygen. Those alkyl groups substituted by an atom, such as phenoxymethyl, 2-pyridyloxymethyl 3-(1-naphthyloxy)propyl and the like.

The term "leaving group" refers to a functional group or atom which may be substituted by another functional group or atom by a substitution reaction (for example, an affinity substitution reaction). For example, representative leaving groups include triflate; chlorine, bromine, iodine; sulfonate groups such as mesylate, tosylate, p-bromobenzenesulfonate, p-toluenesulfonic acid Esters and the like; acyloxy groups such as acetoxy, trifluoroacetoxy and the like.

The term "protecting group" includes, but is not limited to, "amino protecting group", "hydroxy protecting group" or "thiol protecting group". The term "amino protecting group" refers to a protecting group suitable for preventing side reactions at the amino nitrogen position. Representative amino protecting groups include, but are not limited to, formyl; acyl, such as alkanoyl (e.g., acetyl, trichloroacetyl or trifluoroacetyl); alkoxycarbonyl, e.g., tert-butoxycarbonyl (Boc) Arylmethoxycarbonyl, such as benzyloxycarbonyl (Cbz) and 9-fluorenylmethoxycarbonyl (Fmoc); arylmethyl, such as benzyl (Bn), trityl (Tr), 1, 1-di -(4'-methoxyphenyl)methyl; silyl groups such as trimethylsilyl (TMS) and tert-butyldimethylsilyl (TBS) and the like. The term "hydroxy protecting group" refers to a protecting group suitable for use in preventing hydroxy side reactions. Representative hydroxy protecting groups include, but are not limited to, alkyl groups such as methyl groups, Ethyl and tert-butyl; acyl, such as alkanoyl (such as acetyl); arylmethyl, such as benzyl (Bn), p-methoxybenzyl (PMB), 9-fluorenylmethyl (Fm) And diphenylmethyl (diphenylmethyl, DPM); silyl groups such as trimethylsilyl (TMS) and tert-butyldimethylsilyl (TBS) and the like.

The compounds of the present invention can be prepared by a variety of synthetic methods well known to those skilled in the art, including the specific embodiments set forth below, combinations thereof with other chemical synthetic methods, and those well known to those skilled in the art. Equivalent alternatives, preferred embodiments include, but are not limited to, embodiments of the invention.

All solvents used in the present invention are commercially available and can be used without further purification. The present invention employs the following abbreviations: Compound 1 is the isomer 2 of Example 51; Pen. is pentoxifylline; INT-747 is 6-ethyl chenodeoxycholic acid; aq stands for water; HATU stands for O- (7-azabenzotriazol-1-yl)-N,N,N',N'-tetramethyluronium hexafluorophosphate; EDC stands for N-(3-dimethylaminopropyl)-N '-Ethylcarbodiimide hydrochloride; m-CPBA stands for 3-chloroperoxybenzoic acid; eq stands for equivalent, equivalent; CDI stands for carbonyldiimidazole; DCM stands for dichloromethane; PE stands for petroleum ether; DIAD stands for Diisopropyl azodicarboxylate; DMF stands for N,N-dimethylformamide; DMSO stands for dimethyl sulfoxide; EtOAc stands for ethyl acetate; EtOH stands for ethanol; MeOH stands for methanol; CBz stands for benzyloxycarbonyl, An amine protecting group; BOC represents tert-butylcarbonyl is an amine protecting group; HOAc represents acetic acid; NaCNBH ₃ represents sodium cyanoborohydride; rt represents room temperature; O/N represents overnight; THF represents tetrahydrofuran Boc ₂ O represents di-tert-butyl dicarbonate; TFA represents trifluoroacetic acid; DIPEA represents diisopropylethylamine; SOCl ₂ represents thionyl chloride; CS ₂ represents carbon disulfide; TsOH represents p-toluenesulfonic acid; NFSI stands for N-fluoro-N-(phenylsulfonyl)benzenesulfonamide; NCS stands for 1-chloropyrrolidine-2,5-dione; n-Bu ₄ NF stands for tetrabutylammonium fluoride; iPrOH stands for 2-propyl Alcohol; mp represents melting point; LDA represents lithium diisopropylamide; TMSCF ₃ represents trifluoromethyltrimethylsilane; Ti(Oi-Pr) ₄ represents tetraisopropyl titanate; MSCl represents methanesulfonyl chloride; DMAP Represents N,N-dimethyl-4-aminopyridine; TEA stands for triethylamine; BnBr stands for benzyl bromide; DIEA stands for diisopropylethylamine; BH ₃ DMS stands for borane dimethyl sulfide; DMP stands for Des Martin Transiodane; TBAF stands for tetrabutylammonium fluoride; HOBT stands for 1-hydroxybenzotriazole; AIBN stands for azobisisobutyronitrile; NBS stands for N-bromosuccinimide; RT buffer stands for reverse transcription Buffer; dNTP stands for deoxyribonucleoside triphosphate; PBS stands for phosphate buffer.

软件命名，市售化合物采用供应商目录名称。Compound by hand or

Software naming, commercially available compounds using the supplier catalog name.

DRAWINGS

Figure 1: Liver HE staining of MCD mice;

Figure 2: Liver histology scores of MCD mice, data expressed as mean ± standard error, n = 8, ####p<0.0001, ###p<0.001 vs. MCD control diet + vehicle control group, *** p<0.001 vs. MCD diet + vehicle control group, one-way repeated variance analysis and Tukey's group comparison;

Figure 3: Effect of Compound 1 on liver TG levels in MCD mice, data are expressed as mean ± standard error, n = 7-8. ####p<0.0001vs.MCD control diet + vehicle control group. Single factor repeated analysis of variance and pairwise comparison between Tukey’s groups, the overflow value (>mean + 2 × standard deviation, or < mean - 2 × standard deviation) has been removed;

Figure 4: Effect of Compound 1 on liver TNF-α mRNA levels in MCD mice. Data are expressed as mean ± standard error, n = 7-8. #p<0.05vs. MCD control diet + vehicle control group. Single factor repeated analysis of variance and pairwise comparison between Tukey’s groups, the overflow value (>mean + 2 × standard deviation, or < mean - 2 × standard deviation) has been removed;

Figure 5: Compound 1 inhibits liver IL-1β levels in MCD mice, and the data are expressed as mean ± standard error, n = 7-8. ###p<0.001vs.MCD control diet + vehicle control group, *p<0.001 vs. MCD diet + vehicle control group. One-way repeated analysis of variance and two between Tukey’s groups For the two comparisons, the overflow value (>mean + 2 × standard deviation, or < average - 2 × standard deviation) has been removed;

Figure 6: Compound 1 inhibits plasma TNF-α and IL-1β levels in MCD mice, and the data are expressed as mean ± standard error, n = 7-8. ####p<0.0001vs.MCD control diet + vehicle control group. *p<0.05, **p<0.01 vs. MCD diet + vehicle control group, one-way repeated analysis of variance and pairwise comparison between Uncorrected Fisher's LSD group, overflow value (>mean + 2 × standard deviation, or < mean - 2 × standard deviation) has been removed;

Figure 7: Effect of Compound 1 on plasma ALT and AST levels in MCD mice, data are expressed as mean ± standard error, n = 7-8. ####p<0.0001vs.MCD control diet + vehicle control group. Single factor repeated analysis of variance and pairwise comparison between Tukey’s groups, the overflow value (>mean + 2 × standard deviation, or < mean - 2 × standard deviation) has been removed;

Figure 8: Liver HE staining of nSTZ+HFD mice

A. Control group (STZ solvent control + normal diet) B. Model group (STZ + high fat diet) + vehicle control (t.i.d.)

C. Model group (STZ + high fat diet) + pentoxifylline (100 mg / kg, p.o, t.i.d.)

D. Model group (STZ + high fat diet) + Compound 1 (30 mg / kg, p.o, b.i.d.);

Figure 9: Liver histology NAS scores of nSTZ+HFD mice. Data are expressed as mean ± standard error, n = 12 to 17. ####p<0.0001,##p<0.01vs.Control group. One-way repeated analysis of variance and a pairwise comparison between Uncorrected Fish’s LSD groups;

Figure 10: Live picture of Sirius red staining in STZ+HFD mice;

Figure 11: Liver fibrosis scores in nSTZ+HFD mice. Data are expressed as mean ± standard error, n = 3, ##p < 0.01 vs. control group, *p < 0.05 vs. model group + vehicle control. One-way repeated analysis of variance and a pairwise comparison between Uncorrected Fish’s LSD groups;

Figure 12: Effect of Compound 1 on liver biomarker mRNA levels in nSTZ + HFD mice. Data are expressed as mean ± standard error, n = 4 to 20. #p<0.05, ##p<0.01vs. Control group, *p<0.05 vs. model group + vehicle control. One-way repeated analysis of variance and a pairwise comparison between Uncorrected Fish’s LSD groups, the overflow value (>mean + 2 × standard deviation, or < mean - 2 × standard deviation) has been removed;

Figure 13: Effect of Compound 1 on serum biochemical parameters of nSTZ+HFD mice. Data are expressed as mean ± standard error, n = 15 to 17. ###p<0.001,####p<0.0001vs.Control group. One-way repeated analysis of variance and a pairwise comparison between Uncorrected Fish’s LSD groups, the overflow value (>mean + 2 × standard deviation, or < mean - 2 × standard deviation) has been removed;

Figure 14: Liver HE staining of a model of cholestasis-induced liver fibrosis, 100-fold magnification.

A: group-1, healthy control; B: group-2, model control; C: group-3INT-747-20 mg/kg, QD; D: group-4, pentoxifylline-100 mg/kg, TID; E : Group-5, compound 1-1 mg/kg, BID; F: Group-6, compound 1-3 mg/kg, BID; G: group-7, compound 1-10 mg/kg, BID.

Figure 15: Liver pathological inflammation score in cholestasis-type liver fibrosis model rats. Data are indicated as mean ± SEM, n = 12. ***p<0.001 vs model group;

Figure 16: Liver Sirius red staining of a model of cholestasis-induced liver fibrosis in rats, 50-fold magnification.

A: group-1, healthy control; B: group-2, model control; C: group-3INT-747-20 mg/kg, QD; D: group-4, pentoxifylline-100 mg/kg, TID; E : Group-5, compound 1-1 mg/kg, BID; F: Group-6, compound 1-3 mg/kg, BID; G: group-7, compound 1-10 mg/kg, BID.

Figure 17: Liver fibrosis area in a rat model of cholestasis liver fibrosis. Data were expressed as mean ± SEM, n = 12, compared with the Vechile control group, *p < 0.05, *** p < 0.001;

Figure 18: Liver HE staining picture of mice with liver fibrosis induced by CCl ₄ , 100-fold magnification.

A: group-1, healthy control; B: group-2, model control; C: group -3 INT-747-20 mg/kg, QD; D: group-4, pentoxifylline - 100 mg/kg, TID; E: Group-5, compound 1-1 mg/kg, BID; F: Group-6, compound 1-3 mg/kg, BID; G: Group-7, compound 1-10 mg/kg, BID.

Figure 19: Liver pathological inflammation score in mice with liver fibrosis induced by CCl ₄ . Data are indicated as mean ± SEM, n = 15. Compared with the model group, ***p<0.001, **p<0.01, *p<0.05;

Figure 20: Liver balloon-like variation scores in mice with liver fibrosis induced by CCl ₄ . Data were expressed as mean ± SEM, n = 15, compared with the model group, *** p < 0.001, ** p < 0.01, * p <0.05;

Figure 21: Liver water-like degeneration scores in mice with liver fibrosis induced by CCl ₄ . Data were expressed as mean ± SEM, n = 15, compared with the model group, *** p < 0.001, ** p < 0.01, * p <0.05;

FIG 22: CCl ₄ induced liver fibrosis model mouse liver necrosis score. Data were expressed as mean ± SEM, n = 15, compared with the model group, *** p < 0.001, ** p < 0.01, * p <0.05;

Figure 23: Comprehensive scores of liver damage in mice with liver fibrosis induced by CCl ₄ . Data were expressed as mean ± SEM, n = 15, compared with the model group, *** p < 0.001, ** p < 0.01, * p <0.05;

FIG 24: CCl ₄ liver fibrosis model mice induced by Sirius red staining images in liver of mice, 50-fold magnification.

A: group-1, healthy control; B: group-2, model control; C: group-3INT-747-20 mg/kg, QD; D: group-4, pentoxifylline-100 mg/kg, TID; E : Group-5, compound 1-1 mg/kg, BID; F: Group-6, compound 1-3 mg/kg, BID; G: group-7, compound 1-10 mg/kg, BID.

Figure 25: Liver fibrosis area of mouse liver fibrosis model mice induced by CCl ₄ . Data are presented as mean ± SEM, n = 15, compared to the model group, *p < 0.05, *** p < 0.001.

Detailed ways

The invention is described in detail below by the examples, but is not intended to limit the invention.

Example 1

3,7-Dimethyl-1-(6,6,6-trifluoro-5-hydroxy-5-methylhexyl)-1H-indole-2,6(3H,7H)-dione

first step

3,7-Dimethyl-1-(6,6,6-trifluoro-5-methyl-5-(trimethylsiloxy)hexyl)-1H-indole-2,6(3H,7H) -dione

3,7-Dimethyl-1-(5-oxohexyl)-1H-indole-2,6(3H,7H)-dione (200 mg, 0.719 mmol), cesium fluoride (10.9 mg, 0.0719 mmol) Dissolved in tetrahydrofuran (2 mL), trifluoromethyltrimethylsilane (153 mg, 1.08 mmol) was added dropwise at 0 °C. The reaction mixture was stirred at 20 ° C for 2 hr. The organic layer was washed with brine (100 mL×3) Drying in vacuo gave 3,7-dimethyl-1-(6,6,6-trifluoro-5-methyl-5-(trimethylsiloxy)hexyl)-1H-indole-2,6 (3H , 7H)-dione (200 mg, white solid), yield: 66%. MS-ESI calcd [M+H] ⁺ 422.

Second step

3,7-Dimethyl-1-(6,6,6-trifluoro-5-hydroxy-5-methylhexyl)-1H-indole-2,6(3H,7H)-dione

3,7-Dimethyl-1-(6,6,6-trifluoro-5-methyl-5-(trimethylsiloxy)hexyl)-1H-indole-2,6(3H,7H The diketone (200 mg, 0.476 mmol) was dissolved in tetrahydrofuran (2 mL), and 1M hydrochloric acid (0.5 mL) was added dropwise at 0 ° C and then stirred at 20 ° C for 1 hour. The mixture was cooled to 0 ° C and quenched with sodium bicarbonate (30 mL). The mixture was extracted with ethyl acetate (100 mL x 3). The organic layer was washed with brine (100 mL EtOAc) Separation and purification by preparative high performance liquid chromatography gave 3,7-dimethyl-1-(6,6,6-trifluoro-5-hydroxy-5-methylhexyl)-1H-indole-2,6 (3H, 7H)-Dione (50.0 mg, white solid), yield: 30%. ^{1 H NMR: (400MHz, Methonal} -d 4) δ7.85 (s, 1H), 4.02-3.98 (m, 2H), 3.96 (s, 3H), 3.52 (s, 3H), 1.69-1.64 (m, 4H), 1.52-1.48 (m, 2H), 1.28 (s, 3H). MS-ESI calcd for [M+H] ⁺ 349.

Example 2

1-(5-hydroxy-5-methylheptyl)-3,7-dimethyl-1H-indole-2,6(3H,7H)-dione

first step

Ethyl 5-(3,7-dimethyl-2,6-dioxo-2,3,6,7-tetrahydro-1H-indol-1-yl)pentanoic acid ethyl ester

3,7-Dimethyl-1H-indole-2,6(3H,7H)-dione (5.00 g, 28.0 mmol), ethyl bromopentanoate (7.51 g, 33.4 mmol), potassium carbonate (7.73 g) , 56.0 mmol) and potassium iodide (500 mg, 2.80 mmol) were dissolved in N,N-dimethylformamide (62 mL). The reaction solution was heated to 110 ° C and stirred for two hours. The reaction was poured into water and extracted with ethyl acetate (20 mL x 3). The organic phases were combined, dried over anhydrous sodium sulfate and filtered, and then filtered, ethyldiethyldiethyldiethyldiethyl嘌呤-1-yl)ethyl valerate (5.00 g, yellow solid), yield: 50%. ¹ H NMR: (400MHz, CDCl ₃ ) δ 7.51 (s, 1H), 4.14 - 4.09 (m, 2H), 4.04-4.01 (m, 2H), 3.97 (s, 3H), 3.57 (s, 3H) , 2.37-2.33 (m, 2H), 1.72-1.69 (m, 4H), 1.25 (t, J = 7.2 Hz, 3H).

Second step

1-(5-ethyl-5-hydroxyheptyl)-3,7-dimethyl-1H-indole-2,6(3H,7H)-dione

Ethyl 5-(3,7-dimethyl-2,6-dioxo-2,3,6,7-tetrahydro-1H-indol-1-yl)pentanoic acid ethyl ester (0.500 g, 1.62) Ethyl acetate was dissolved in anhydrous tetrahydrofuran (5 mL). N.sub.3. The reaction solution was stirred at -78 ° C for 0.5 hour, then slowly raised to 0 ° C and reacted for 0.5 hour. The reaction solution was poured into water and extracted with ethyl acetate (30 mL, 3). The organic phase was combined, dried over anhydrous sodium sulfate, filtered and evaporated. 7H) - dione (0.300 g, colorless oil). ^{yield: 57% 1 H NMR: (} 400MHz, CDCl 3) δ7.50 (s, 1H), 4.05-4.01 (m, 2H), 3.99. (s, 3H), 3.57 (s, 3H), 1.70-1.37 (m, 10H), 0.86 (t, J = 7.6 Hz, 6H). MS-ESI calculated [M+H] ⁺ 323, found 323 .

Example 3

1-(4-(1-hydroxycyclopropyl)butyl)-3,7-dimethyl-1H-indole-2,6(3H,7H)-dione

Ethylmagnesium bromide (3M ether solvent, 1.1 mL, 3.24 mmol) was added to ethyl 5-(3,7-dimethyl-2,6-dioxo-2,3 at -35 °C under nitrogen. ,6,7-Tetrahydro-1H-indol-1-yl)pentanoic acid ethyl ester (500 mg, 1.62 mmol) and tetraisopropyl titanate (461 mg, 1.62 mmol) in tetrahydrofuran (10 mL). The reaction solution was slowly heated to 25 ° C and stirred for 2 hours. The reaction was quenched by water (10 mL). The organic phase was combined, dried over anhydrous sodium sulfate, filtered, and then filtered and evaporated. -1H-indole-2,6(3H,7H)-dione (90.0 mg, white solid), yield: 19%. ¹ H NMR: (400 MHz, Methonal-d ₄ ) δ 7.86 (s, 1H), 4.03 - 3.90 (m, 5H), 3.51 (s, 3H), 1.72-1.53 (m, 6H), 0.68-0.59 ( m, 2H), 0.46-0.38 (m, 2H). MS-ESI calcd for [M ⁺ H] + 293, found 293.

Example 4

3,7-Dimethyl-1-((1-(1,1,1-trifluoro-2-hydroxypropan-2-yl)cyclopropyl)methyl)-1H-indole-2,6-( 3H,7H)-dione

first step

Ethyl 1-acetylcyclopropane

Ethyl-3-oxobutanoic acid (10.0 g, 76.8 mmol) and 1,2-dibromoethane (21.7 g, 115 mmol) were dissolved in dimethyl sulfoxide (300 mL) under nitrogen and then Potassium carbonate (42.5 g, 307 mmol) was added portionwise. The reaction solution was stirred at 25 ° C for 24 hours. Water (500 mL) was added, and the mixture was evaporated. mjjjjjjjjjjjjjjjjj Ethyl acetate, Rf = 0.4) gave ethyl 1- acetylcyclopropane (6.00 g, white oil), yield: 50%. ¹ H NMR: (400 MHz, Methonal-d ₄ ) δ 4.25 - 4.20 (m, 2H), 2.44 (s, 3H), 1.47-1.42 (m, 4H), 1.32-1.28 (m, 3H).

Second step

1-(1,1,1-trifluoro-2-hydroxypropan-2-yl)cyclopropanecarboxylic acid

Ethyl 1-acetylcyclopropane (2.00 g, 12.8 mmol), cesium fluoride (195 mg, 1.28 mmol) was dissolved in tetrahydrofuran (30 mL), then trifluoromethyltrimethylsilane (3.64 g, 25.6 mmol). The reaction solution was reacted under nitrogen for 20 hours at 20 °C. Then 4N dilute hydrochloric acid (7 mL) was added. The mixture was reacted under nitrogen for 6 hours at room temperature. The reaction was quenched with EtOAc EtOAc (EtOAc (EtOAc) Petroleum ether/ethyl acetate, Rf = 0.4) gave 1-(1,1,1-trifluoro-2-hydroxypropan-2-yl)cyclopropanecarboxylic acid (1.70 g, white oil). %. ^{1 H NMR: (400MHz, Methonal} -d 4) δ4.14-4.10 (m, 2H), 1.64 (s, 3H), 1.29-1.24 (m, 3H), 1.23-1.22 (m, 2H), 0.92- 0.90 (m, 2H).

third step

1,1,1-trifluoro-2-(1-(hydroxymethyl)cyclopropyl)propan-2-ol

1-(1,1,1-Trifluoro-2-hydroxypropan-2-yl)cyclopropanecarboxylic acid (400 mg, 1.77 mmol) was dissolved in anhydrous tetrahydrofuran (10 mL), and lithium tetrahydroaluminum was added at 0 °C. (81.0 mg, 2.12 mmol). The reaction solution was warmed to 25 ° C and stirred for 1 hour. Quenched with water (10 mL), EtOAc (EtOAc)EtOAc. 0.2), 1,1,1-Trifluoro-2-(1-(hydroxymethyl)cyclopropyl)propan-2-ol (200 mg, yellow oil) was obtained. ¹ H NMR: (400 MHz, DMSO-d ₆ ) δ 5.64 (s, 1H), 4.63-4.60 (m, 1H), 3.64 - 3.60 (m, 1H), 3.23 - 3.17 (m, 1H), 1.36 ( s, 3H), 0.83-0.91 (m, 1H), 0.56-0.55 (m, 1H), 0.39-0.35 (m, 2H).

the fourth step

(1-(1,1,1-trifluoro-2-hydroxypropan-2-yl)cyclopropyl)methyl methanesulfonate

Dissolve 1,1,1-trifluoro-2-(1-(hydroxymethyl)cyclopropyl)propan-2-ol (100 mg, 0.543 mmol) in dichloromethane (5 mL) and add at 0 °C Triethylamine (110 mg, 1.08 mmol) and methanesulfonyl chloride (62.2 mg, 0.543 mmol). The reaction solution was reacted at 0 ° C for 2 hours. The organic layer was extracted with a saturated aqueous solution of sodium chloride (10 mL×3), washed with a saturated sodium chloride solution (10 mL×3), dried over anhydrous sodium sulfate Concentration by pressure gave (1-(1,1,1-trifluoro-2-hydroxypropan-2-yl)cyclopropyl)methyl methanesulfonate (80.0 mg, yellow oil), yield: 56% .

the fifth step

3,7-Dimethyl-1-((1-(1,1,1-trifluoro-2-hydroxypropan-2-yl)cyclopropyl)methyl)-1H-indole-2,6-( 3H,7H)-dione

(1-(1,1,1-Trifluoro-2-hydroxypropan-2-yl)cyclopropyl)methyl methanesulfonate (80.0 mg, 0.305 mmol), 3,7-dimethyl-1H- Indole-2,6-(3H,7H)-dione (54.9 mg, 0.305 mmol), potassium iodide (5.10 mg, 0.0305 mmol) and potassium carbonate (126 mg, 0.915 mmol) dissolved in anhydrous N,N-dimethyl Formamide (5 mL). The reaction solution was heated to 120 ° C and allowed to react for 2 hours. The reaction solution was cooled to 20 ° C, filtered, and purified by preparative HPLC to give 3,7-dimethyl-1-((1-(1,1,1-trifluoro-2-hydroxypropan-2-yl) Cyclopropyl)methyl)-1H-indole-2,6-(3H,7H)-dione (40.0 mg, white solid), yield: 38%. ¹ H NMR: (400 MHz, Methonal-d ₄ ) δ 7.88 (s, 1H), 4.45 (d, J = 6.8 Hz, 1H), 4.24 (d, J = 6.8 Hz, 1H), 3.97 (s, 3H) ), 3.53 (s, 3H), 1.53 (s, 3H), 0.92-0.88 (m, 1H), 0.64-0.63 (m, 1H), 0.41-0.38 (m, 1H), 0.15-0.12 (m, 1H) ).

MS-ESI calcd for [M ⁺ H] + 347, found 347.

Example 5

1-((3-hydroxy-3-(trifluoromethyl)cyclobutyl)methyl)-3,7-dimethyl-1H-indole-2,6(3H,7H)-dione

First step

Methyl 3-oxocyclobutanecarboxylate

3-Oxocyclobutanecarboxylic acid (25.0 g, 0.220 mmol), methanol (14 mL) and N,N-dimethyl-4-aminopyridine (3.00 g, 353 mmol) After stirring at 25 ° C, 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (64.0 g, 340 mmol) was slowly added dropwise and stirred overnight. The reaction solution was washed with aqueous hydrochloric acid (1.5N, EtOAc), water (150 mL), and brine (75mL). The organic phase was dried over anhydrous sodium sulfate and evaporated, evaporated]]]]]]

Second step

Methyl 5,8-dioxaspiro[3,4]octane-2-carboxylate

Methyl-3-oxocyclobutanecarboxylic acid (25.0 g, 195 mmol), ethylene glycol (35.0 g, 564 mmol) and p-toluenesulfonic acid (3.50 g, 20.0 mmol) were dissolved in toluene (250 mL). The water separator was heated to reflux overnight. The reaction solution was cooled to 25 ° C, washed sequentially with water (300 mL×2) and saturated sodium hydrogen carbonate (500 mL×2). The organic phase was dried over anhydrous magnesium sulfate, filtered, and then filtered and evaporated tolululululululululululululululululululululu Rate: 90%.

third step

5,8-Dioxaspiro[3,4]octane-2-methanol

Lithium tetrahydroaluminum (5.20 g, 136 mmol) was slowly dissolved in tetrahydrofuran (240 mL) under nitrogen at 0 ° C, then 5,8-dioxaspiro[3,4] dissolved in tetrahydrofuran (60 mL) Methyl octane-2-carboxylate (19.5 g, 113 mmol). The reaction was slowly raised to 25 ° C and stirred for 3.5 hours. The reaction solution was cooled to 0 ° C, and water (5.20 g, 289 mmol), 15% sodium hydroxide (5.20 g, 19.5 mmol) and water (15.6 g, 867 mmol) were added slowly. Filtration, the filter cake was washed with tetrahydrofuran (10 mL×3), and the filtrate was concentrated under reduced pressure to silica gel column chromatography (1:1 petroleum ether/ethyl acetate, Rf=0.4) to give the product 5,8-dioxaspiro[3 4]octane-2-methanol (10.0 g, yellow liquid), yield: 62%. ^{1 H NMR: (400MHz, CDCl} 3) δ3.90-3.87 (m, 4H), 3.67 (d, J = 6.4Hz, 2H), 2.45-2.40 (m, 2H), 2.38-2.26 (m, 1H) , 2.13 - 2.08 (m, 2H).

the fourth step

5,8-Dioxaspiro[3,4]octane-2-ylmethylmethanesulfonate

5,8-Dioxaspiro[3,4]octane-2-methanol (500 mg, 53.1 mmol) and triethylamine (896 mg, 6.90 mmol) were dissolved in dichloromethane (23 mL) and slowly added at 0 °C Methanesulfonyl chloride (1.40 g, 12.6 mmol). The reaction solution was raised to 25 ° C and stirred overnight. The reaction was quenched with water (50 mL)EtOAcEtOAc The organic phases were combined, dried over anhydrous sodium sulfate and filtered and evaporated ).

MS-ESI calcd for [M ⁺ H] + 223, found 223.

the fifth step

(1-(5,8-Dioxaspiro[3,4]octane-2-ylmethyl)-3,7-dimethyl-1H-indole-2,6(3H,7H)-dione

5,8-Dioxaspiro[3,4]octane-2-ylmethyl methanesulfonate (1.00 g, 4.50 mmol), 3,7-dimethyl-1H-indole-2,6 ( 3H,7H)-Diketone (810 mg, 4.50 mmol), potassium carbonate (1.20 g, 13.5 mmol) and potassium iodide (75.0 mg, 0.45 mmol) were dissolved in N,N-dimethylformamide (20 mL). The reaction was heated to 130 ° C and stirred for 3.5 hours. The reaction solution was filtered, and the filtrate was concentrated under reduced pressure to give 1-(5,8-dioxaspiro[3,4]octane-2-ylmethyl)-3,7-dimethyl-1H-indole-2. 6(3H,7H)-dione (1.50 g, brown liquid), yield: 93%. MS-ESI calcd for [M+H] ⁺ 303.

Step 6

3,7-Dimethyl-1-((4-oxocyclohexyl)methyl)-1H-indole-2,6(3H,7H)-dione

1-(5,8-Dioxaspiro[3,4]octane-2-ylmethyl)-3,7-dimethyl-1H-indole-2,6(3H,7H)-dione (1.50 g, 5.00 mmol) was dissolved in EtOAc (EtOAc) (EtOAc) The reaction was heated to 30 ° C and stirred overnight. It was diluted with water, and the mixture was adjusted to EtOAc EtOAc (EtOAc) The organic phase was combined, dried over anhydrous sodium sulfate, filtered, evaporated, evaporated, evaporated, evaporated. 1-((4-oxocyclohexyl)methyl)-1H-indole-2,6(3H,7H)-dione (180 mg, white solid), yield: 14%.

^{1 H NMR: (400MHz, CDCl} 3) δ7.49 (s, 1H), 4.25 (d, J = 7.6Hz, 2H), 3.95 (s, 3H), 3.55 (s, 3H), 3.13-2.96 (m , 4H), 2.95-2.84 (m, 1H). MS-ESI calcd [M+H] ⁺ 262.

Seventh step

1-((3-hydroxy-3-(trifluoromethyl)cyclopentyl)methyl)-3,7-dimethyl-1H-indole-2,6(3H,7H)-dione

3,7-Dimethyl-1-((3-oxocyclopentyl)methyl)-1H-indole-2,6(3H,7H)-dione (100 mg, 0.382 mmol) and cesium fluoride (11.5 mg, 0.0763 mmol) was dissolved in anhydrous tetrahydrofuran (3 mL) and trifluoromethyltrimethylsilane (95.0 mg, 0.640 mmol). The reaction solution was slowly heated to 30 ° C and stirred for 12 hours. Aqueous hydrochloric acid (1 N, 5 mL) was then added to the reaction and stirring was continued for 0.5 h. The reaction mixture was diluted with water (50 mL), EtOAc (EtOAc) Trifluoromethyl)cyclopentyl)methyl)-3,7-dimethyl-1H-indole-2,6(3H,7H)-dione (80.0 mg, white solid). 1H NMR: (400MHz, Mehonal-d ₄ ) δ 8.54 (s, 1H), 4.13-4.07 (m, 5H), 3.56 (s, 3H), 2.58-2.48 (m, 3H), 2.14-2.10 (m) , 2H). MS-ESI calcd [M+H] ⁺ 333.

Example 6

1-((3,7-Dimethyl-2,6-dioxo-2,3,6,7-tetrahydro-1H-indol-1-yl)methyl)-3-hydroxycyclobutyronitrile

first step

(((1,3-dibromopropan-2-yl)oxy)methyl)benzene

2-(Bromomethyl)oxirane (8.40 g, 61.3 mmol) was added to benzyl bromide (8.74 g, 51.1 mmol) dissolved in cuprous chloride (6.87 g, 51.1 mmol) at room temperature. The reaction was stirred at 150 ° C for 11 hours. The reaction solution was cooled to room temperature, water (100 mL) was slowly added, and ethyl acetate (100 mL x 3). The organic phase was combined, dried over anhydrous sodium sulfate, filtered and evaporated, evaporated, evaporated, Alkyloxy)methyl)benzene (8.60 g, yellow oil), yield: 44%. ¹ H NMR: (400 MHz, CDCl ₃ ) δ 7.39 - 7.31 (m, 5H), 4.67 (s, 2H), 3.82-3.78 (m, 1H), 3.58 (d, J = 5.2 Hz, 4H).

Second step

Ethyl ethyl 3-(benzyloxy)-1-cyanocyclobutanecarboxylate

Ethyl cyanoacetate (2.76 g, 24.3 mmol) was slowly added to the solution (((1,3-dibromopropyl-2-yl)oxy)methyl)benzene (7.00 g, 18.2 mmol) and carbonic acid at room temperature. Potassium (10.0 g, 72.7 mmol) in N,N-dimethylformamide (35 mL). The reaction was stirred at 90 ° C for 4 hours. It was cooled to room temperature, filtered, and the~~~~ The organic phase obtained was washed with a saturated aqueous solution of ammonium chloride (20 mL x 3). The organic phase was dried over anhydrous sodium sulfate (MgSO4), filtered,jjjjjjjjjjjjjjjjj Ethyl-1-cyanocyclobutanecarboxylate (3.80 g, colorless oil), yield: 81%. ^{1 H NMR: (400MHz, Methonal} -d 4) δ7.40-7.28 (m, 5H), 4.48-4.44 (m, 2H), 4.37-4.31 (m, 1H), 4.30-4.24 (m, 2H), 2.97-2.80 (m, 2H), 2.73-2.65 (m, 2H), 1.37-1.30 (m, 3H).

third step

3-(benzyloxy)-1-(hydroxymethyl)cyclobutyronitrile

Sodium borohydride (1.39 g, 36.6 mmol) was dissolved in tetrahydrofuran and water (20 mL: 2 mL), and ethyl 3-(benzyloxy)-1-l was slowly added dropwise at 0 °C over 20 min. A solution of ethyl cyanocyclobutanecarboxylate (3.80 g, 14.6 mmol) in tetrahydrofuran (22 mL). The reaction was stirred at room temperature for 2 hours. The organic layer was washed with water (30 mL) and brine (30 mL). -(Hydroxymethyl)cyclobutyronitrile (3.70 g, colorless oil). ^{1 H NMR: (400MHz, DMSO} -d 6) δ7.38-7.25 (m, 5H), 5.57-5.52 (m, 1H), 4.39-4.36 (m, 2H), 4.13-4.04 (m, 1H), 3.57-3.51 (m, 2H), 2.58-2.51 (m, 1H), 2.49-2.45 (m, 1H), 2.31-2.09 (m, 2H).

the fourth step

(3-(benzyloxy)-1-cyanocyclobutyl)methyl methanesulfonate

3-(Benzyloxy)-1-(hydroxymethyl)cyclobutanenitrile (3.70 g, 15.3 mmol), triethylamine (3.10 g, 30.6 mmol) dissolved in dichloromethane (35 mL) Methanesulfonyl chloride (3.29 g, 28.7 mmol) was slowly added. The reaction mixture was stirred at room temperature for 4 hr. EtOAc (EtOAc) Oxy)-1-cyanocyclobutyl)methyl methanesulfonate (4.56 g, dark brown oil). ^{1 H NMR: (400MHz, Methonal} -d 4) δ7.36-7.26 (m, 5H), 4.47-4.45 (m, 2H), 4.44-4.38 (m, 2H), 3.21-3.18 (m, 1H), 3.17-3.14 (m, 3H), 2.81-2.60 (m, 2H), 2.53-2.26 (m, 2H).

the fifth step

3-(Benzyloxy)-1-((3,7-dimethyl-2,6-dioxo-2,3,6,7-tetrahydro-1H-indol-1-yl)methyl) Cyclobutyronitrile

(3-(Benzyloxy)-1-cyanocyclobutyl)methyl methanesulfonate (4.50 g, 15.2 mmol), 3,7-dimethyl-1H-indole-2,6-(3H , 7H)-dione (2.75 g, 15.2 mmol) and potassium iodide (1.26 g, 7.62 mmol) were dissolved in N,N-dimethylformamide (100 mL), and potassium carbonate (6.32 g, 45.7 mmol) was added. The mixture was heated to reflux at 120 ° C for 4 hours. The reaction solution was cooled to room temperature, filtered, and evaporated, evaporated, evaporated. The organic phase was combined, dried over anhydrous sodium sulfate, filtered and evaporated , 3,6,7-Tetrahydro-1H-indol-1-yl)methyl)cyclobutyronitrile (4.60 g, yellow solid). MS-ESI calcd [M+H] ⁺ 380.

Step 6

1-((3,7-Dimethyl-2,6-dioxo-2,3,6,7-tetrahydro-1H-indol-1-yl)methyl)-3-hydroxycyclobutyronitrile

3-(Benzyloxy)-1-((3,7-dimethyl-2,6-dioxo-2,3,6,7-tetrahydro-1H-indol-1-yl)methyl Cyclobutyronitrile (100 mg, 0.263 mmol) was dissolved in dichloromethane (10 mL), and then ferric chloride (128 mg, 0.790 mmol). The reaction was stirred at room temperature for 12 hours. Water (10 mL) was added and extracted with dichloromethane (40 mL×3). The organic phase was combined and dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure, and the obtained product was purified by preparative high-performance liquid chromatography to give the product 1-((3,7-dimethyl-2,6-dioxo- 2,3,6,7-Tetrahydro-1H-indol-1-yl)methyl)-3-hydroxycyclobutanenitrile (12.0 mg, yellow solid), yield: 16%. ^{1 H NMR: (400MHz, CDCl} 3) δ7.56 (s, 1H), 4.66-4.49 (m, 1H), 4.45-4.37 (m, 2H), 4.01 (s, 3H), 3.62 (s, 3H) , 2.96-2.85 (m, 2H), 2.60-2.49 (m, 2H). MS-ESI calcd for [M+H] ⁺ 290.

Example 7

first step

Methyl 3-(hydroxymethyl)bicyclo[1.1.1]pentan-1-carboxylate

3-(Methoxycarbonyl)bicyclo[1.1.1]pentan-1-carboxylic acid (100 mg, 0.587 mmol) and triethylamine (71.0 mg, 0.705 mmol) were dissolved in tetrahydrofuran (20 mL) at -10 °C Methyl chloroformate (56.0 mg, 0.587 mmol) was slowly added dropwise. The reaction solution was stirred at 0 ° C for half an hour, then sodium borohydride (33.0 mg, 0.881 mmol) was added and the reaction was continued for 2 hr. Water (10 mL) and ethyl acetate (10 mL×3) were evaporated, evaporated, evaporated, evaporated, evaporated, evaporated Methyl (hydroxymethyl)bicyclo[1.1.1]pentane-1-carboxylate (80.0 mg, colorless oil), yield: 87%. ¹ H NMR: (400 MHz, CDCl ₃ ) δ 3.65 (s, 3H), 3.60 (s, 2H), 2.00 (s, 6H).

Second step

Methyl 3-((((methylsulfonyl)oxy)methyl)bicyclo[1.1.1]pentan-1-carboxylate

Methyl 3-(hydroxymethyl)bicyclo[1.1.1]pentane-1-carboxylate (40.0 mg, 0.256 mmol) and triethylamine (39.0 mg, 0.384 mmol) were dissolved in dichloromethane (15 mL) Methanesulfonyl chloride (35.0 mg, 0.307 mmol) was slowly added dropwise at 0 °C. The reaction mixture was stirred at 0 ° C for 2 hours. The mixture was diluted with methylene chloride (10 mL). Methyl (((methylsulfonyl)oxy)methyl)-bicyclo[1.1.1]pentan-1-carboxylate (50.0 mg, yellow oil).

third step

Methyl 3-((3,7-dimethyl-2,6-dioxo-2,3,6,7-tetrahydro-1H-indol-1-yl)methyl)bicyclo[1.1.1]penta Methyl alkane-1-carboxylate

Methyl 3-((((methylsulfonyl)oxy)methyl)bicyclo[1.1.1]pentan-1-carboxylate (100 mg, 0.426 mmol) and 3,7-dimethyl-1H - 嘌呤-2,6-(3H,7H)-dione (77.0 mg, 0.427 mmol) was dissolved in N,N-dimethylformamide (20 mL), and potassium carbonate (88.0 mg, 0.640 mmol) And potassium iodide (8.00 mg, 0.0430 mmol). The reaction mixture was stirred at 100 ° C for 2 hr. The mixture was evaporated, evaporated, evaporated, evaporated, evaporated. Separation and purification by silica gel column chromatography (1:1 petroleum ether / ethyl acetate, Rf = 0.2) to give methyl 3-((3,7-dimethyl-2,6-dioxo-2,3,6) Methyl 7-tetrahydro-1H-indol-1-yl)methyl)bicyclo[1.1.1]pentane-1-carboxylate (100 mg, yellow solid), yield: 73%. ^{1 H NMR: (400MHz, CDCl} 3) δ7.50 (s, 1H), 4.12 (s, 2H), 3.95 (s, 3H), 3.60 (s, 3H), 3.53 (s, 3H), 1.95 (s , 6H). MS-ESI calcd for [M+H] ⁺ 319.

the fourth step

3-((3,7-Dimethyl-2,6-dioxo-2,3,6,7-tetrahydro-1H-indol-1-yl)methyl)bicyclo[1.1.1]pentane- 1-carboxylic acid

Methyl 3-((3,7-dimethyl-2,6-dioxo-2,3,6,7-tetrahydro-1H-indol-1-yl)methyl)bicyclo[1.1.1] Methyl pentane-l-carboxylate (100 mg, 0.314 mmol) was dissolved in tetrahydrofuran (15 mL) and water (5 mL). After stirring at room temperature for 2 hours, the reaction mixture was diluted with EtOAc EtOAc EtOAc (EtOAc) Dry, filter, and concentrate the filtrate under reduced pressure to 3-((3,7-dimethyl-2,6-dioxo-2,3,6,7-tetrahydro-1H-indol-1-yl)methyl)bicyclo [1.1.1] Pentane-1-carboxylic acid (90.0 mg, white solid), yield: 94%.

MS-ESI calcd for [M+H] ⁺ 303.

the fifth step

3-((3,7-Dimethyl-2,6-dioxo-2,3,6,7-tetrahydro-1H-indol-1-yl)methyl)-N-methoxy-N- Methylbicyclo[1.1.1]pentan-1-amide

3-((3,7-Dimethyl-2,6-dioxo-2,3,6,7-tetrahydro-1H-indol-1-yl)methyl)bicyclo[1.1.1]pentane 1-carboxylic acid (30.0 mg, 0.0986 mmol) and N,O-dimethylhydroxylamine (10.0 mg, 0.0986 mmol) were dissolved in dichloromethane (20 mL), and 2-(7-azobenzene) was added at room temperature. Triazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (75.0 mg, 0.197 mmol) and diisopropylethylamine (19.0 mg, 0.148 mmol). After stirring at room temperature for 12 hours, the reaction mixture was evaporated w~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~ Dry over anhydrous sodium sulfate, filter, and concentrate the filtrate under reduced pressure. Separation and purification by silica gel column chromatography (1:2 petroleum ether / ethyl acetate, Rf = 0.2) to give 3-((3,7-dimethyl-2,6-dioxo-2,3,6,7) -tetrahydro-1H-indol-1-yl)methyl)-N-methoxy-N-methylbicyclo[1.1.1]pentan-1-amide (30.0 mg, white solid), yield: 88 %.

MS-ESI calcd for [M ⁺ H] + 348, found 348.

Step 6

1-((3-acetylbicyclo[1.1.1]pentan-1-yl)methyl)-3,7-dimethyl-1H-indole-2,6-(3H,7H)-dione

3-((3,7-Dimethyl-2,6-dioxo-2,3,6,7-tetrahydro-1H-indol-1-yl)methyl)-N-methoxy-N -Methylbicyclo[1.1.1]pentan-1-amide (20.0 mg, 0.0575 mmol) was dissolved in tetrahydrofuran (20 mL). The reaction mixture was added to methyl magnesium bromide (3M diethyl ether solution, 0.040) at -78 °C. mL, 0.120 mmol), stirring was continued for 30 minutes and then allowed to react to room temperature for 4 hours. The reaction mixture was stirred at 0 ° C, EtOAc (EtOAc (EtOAc) The filtrate was concentrated under reduced pressure. Separation and purification by silica gel column chromatography (1:1 petroleum ether / ethyl acetate, Rf = 0.5) to give 1-((3-acetylbicyclo[1.1.1]pentan-1-yl)methyl)-3 , 7-Dimethyl-1H-indole-2,6-(3H,7H)-dione (15.0 mg, colorless oil), yield: 86%. ^{1 H NMR: (400MHz, CDCl} 3) δ7.55 (s, 1H), 4.17 (s, 2H), 3.99 (s, 3H), 3.59 (s, 3H), 2.07 (s, 3H), 1.97 (s , 6H). MS-ESI calcd [M+H] ⁺ 303, Found 303.

Seventh step

3,7-Dimethyl-1-((3-(1,1,1-trifluoro-2-hydroxypropan-2-yl)bicyclo[1.1.1]pentan-1-yl)methyl)- 1H-嘌呤-2,6-(3H,7H)-dione

1-((3-Acetylbicyclo[1.1.1]pentan-1-yl)methyl)-3,7-dimethyl-1H-indole-2,6-(3H,7H)-dione (20.0 mg, 0.0660 mmol) and trifluoromethyltrimethylchlorosilane (19.0 mg, 0.132 mmol) were dissolved in tetrahydrofuran (15 mL), and the reaction mixture was added cesium fluoride (10.0 mg, 00660 mmol) at room temperature, room temperature. Continue to react for 12 hours. 2N dilute hydrochloric acid (10 mL) was added to the reaction mixture, and the mixture was stirred for 30 minutes, and ethyl acetate (20 mL×2) was evaporated. Concentration by pressure and purification by high performance liquid chromatography to give 3,7-dimethyl-1-((3-(1,1,1-trifluoro-2-hydroxypropan-2-yl)bicyclo[1.1.1]penta Alkyl-1-yl)methyl)-1H-indole-2,6-(3H,7H)-dione (5.00 mg, colorless oil), yield: 20%. ^{1 H NMR: (400MHz, CDCl} 3) δ7.59 (s, 1H), 4.20 (s, 2H), 4.02 (s, 3H), 3.59 (s, 3H), 1.97 (s, 6H), 1.79 (s , 3H). MS-ESI calcd for [M ⁺ H] + 373, found 373.

Example 8

first step

3,7-Dimethyl-1-[[3-[2,2,2-trifluoro-1-hydroxy-1-(trifluoromethyl)ethyl]cyclobutyl]methyl]indole-2, 6-diketone

3,7-Dimethyl-1-[[3-(2,2,2-trifluoro-1,1-dihydroxy-ethyl)cyclobutyl]methyl]indole-2,6-dione (60.0 mg, 0.165 mmol), cesium fluoride (25.2 mg, 0.165 mmol) was dissolved in tetrahydrofuran (10 mL), trifluoromethyltrimethylchlorosilane (70.6 mg, 0.496 mmol) was added at room temperature and stirred for 12 hours. The reaction was quenched by the addition of water (20 mL). Extract with ethyl acetate (10 mL x 3). The combined organic layers were washed with brine, dried over anhydrous sodium sulfate Purification by preparative high performance liquid chromatography gave product 1 (8.00 mg, yellow solid) (isomer 1 , first peak), yield: 12%. ^{1 H NMR: (400MHz, Methonal} -d 4) δ8.01 (s, 1H), 4.22-4.17 (m, 2H), 4.01 (s, 3H), 3.54 (s, 3H), 3.55-3.19 (m, 1H), 2.63-2.56 (m, 1H), 2.50-2.42 (m, 2H), 1.82-1.78 (m, 2H). MS-ESI calcd for [M+H] ⁺ 415.

And product 2 (isomer 2, second peak), yield: 6%. ¹ H NMR: (400 MHz, Methonal-d ₄ ) δ 8.20 (s, 1H), 4.04-4.00 (m, 5H), 3.55 (s, 3H) 2.70-2.65 (m, 1H), 2.55-2.53 (m) , 1H), 2.17-2.12 (m, 2H), 2.02-1.98 (m, 2H). MS-ESI calcd for [M+H] ⁺ 415.

Example 9

first step

3-methylenecyclobutanecarboxylic acid

3-Methylenecyclobutyronitrile (10.0 g, 107 mmol) and potassium hydroxide (18.1 g, 322 mmol) were dissolved in ethanol (100 mL) and water (50 mL) and reacted at 100 ° C for 2 hours. 1N Hydrochloric acid (120 mL) was added. Dichloromethane (30 mL x 3), dried over anhydrous sodium sulfate, filtered, evaporated. ¹ H NMR: (400 MHz, Methonal-d ₄ ) δ 4.83-4.76 (m, 2H), 3.15-2.96 (m, 1H), 2.95 - 2.92 (m, 4H).

Second step

Methyl-3-methylenecyclobutanecarboxylic acid

3-Methylenecyclobutanecarboxylic acid (11.0 g, 98.1 mmol) and potassium carbonate (27.1 g, 196 mmol) were dissolved in acetone (100 mL), and dimethyl sulfate (14.8 g, 117 mmol) was added at 25 ° C. After reacting at 70 ° C for 12 hours. The reaction was quenched with water (20 mL) EtOAc (EtOAc md. Oily), yield: 97%. ^{1 H NMR: (400MHz, Methonal} -d 4) δ4.83-4.79 (m, 2H), 3.96 (s, 2H), 3.68 (s, 3H), 3.17-3.15 (m, 1H), 2.95-2.92 ( m, 2H).

third step

Methyl 3-(hydroxymethyl)cyclobutanecarboxylate

Methyl-3-methylenecyclobutanecarboxylic acid (2.00 g, 15.8 mmol) was dissolved in tetrahydrofuran (30 mL), and borane dimethyl sulfide (3.61 g, 47.5 mmol) was added dropwise at -10 ° C, then -10 ° C reaction for 3 hours, adding 3N aqueous sodium hydroxide solution (10 mL) and hydrogen peroxide (5 mL), the reaction was continued for 1 hour, the reaction solution was added to a saturated aqueous solution of sodium thiosulfate (30 mL) to quench the reaction, dichloromethane extraction (10 mL x 3) Drying with anhydrous sodium sulfate, filtration, and the filtrate was concentrated under reduced pressure to give ethyl 3-(hydroxymethyl)cyclobutanecarboxylate (2.00 g, yellow oil). ¹ H NMR: (400 MHz, Methonal-d ₄ ) δ 3.70 (s, 3H), 3.58 (d, J = 6.8 Hz, 1H), 3.49 (d, J = 6.8 Hz, 1H), 3.10-3.05 (m) , 1H), 2.32-2.26 (m, 3H), 2.03-1.98 (m, 2H).

the fourth step

Ethyl 3-(methylsulfonyloxymethyl)cyclobutanecarboxylate

Ethyl 3-(hydroxymethyl)cyclobutanecarboxylate (1.00 g, 6.94 mmol) and triethylamine (2.11 g, 20.8 mmol) were dissolved in dichloromethane (20 mL) and methanesulfonyl chloride was added at 0 °C (1.59 g, 13.9 mmol). The reaction solution was slowly warmed to room temperature and stirred for 2 hours. The reaction was quenched by the addition of aqueous sodium bicarbonate (50 mL). Extract with dichloromethane (10 mL x 3). The organic phase was combined, washed with brine, dried over anhydrous sodium sulfate sulfatessssssssssssssssssssssss , Yield: 91%. ¹ H NMR: (400 MHz, Methonal-d ₄ ) δ 4.28 (d, J = 6.8 Hz, 1H), 4.19 (d, J = 6.8 Hz, 1H), 3.70 (s, 3H), 3.20-3.08 (m) , 4H), 2.40-2.34 (m, 3H), 2.13 - 2.09 (m, 2H). MS-ESI calcd for [M ⁺ H] + 223, found 223.

the fifth step

Ethyl 3-[(3,7-dimethyl-2,6-dioxo-indol-1-yl)methyl]cyclobutanecarboxylate

Ethyl 3-(methylsulfonyloxymethyl)cyclobutanecarboxylate (1.40 g, 6.30 mmol), 3,7-dimethyl-1H-indole-2,6-(3H,7H)- Diketone (1.13 g, 6.30 mmol), potassium iodide (209 mg, 1.26 mmol) and potassium carbonate (2.61 g, 18.90 mmol) were dissolved in N,N-dimethylformamide (100 mL). The reaction solution was warmed to 120 ° C and stirred for 3 hours. It was cooled to room temperature, filtered, and water (100 mL) was evaporated. The combined organic layers were washed with brine, dried over anhydrous sodium sulfate Ethyl methyl]cyclobutanecarboxylate (1.50 g, yellow solid), yield: 78%.

¹ H NMR: (400 MHz, Methonal-d ₄ ) δ 7.51 (s, 1H), 4.18 - 4.10 (m, 2H), 3.99 (s, 3H), 3.67 (s, 3H), 3.55 (s, 3H) , 3.26-2.65 (m, 2H), 2.29-2.13 (m, 4H). MS-ESI calcd for [M+H] ⁺ 303.

Step 6

3-[(3,7-Dimethyl-2,6-dioxo-indol-1-yl)methyl]cyclobutanecarboxylic acid

Ethyl 3-[(3,7-dimethyl-2,6-dioxo-indol-1-yl)methyl]cyclobutanecarboxylate (1.00 g, 3.26 mmol), potassium hydroxide (548 mg) , 9.78 mmol) was dissolved in methanol (10 mL) and water (5 mL). The reaction solution was warmed to 90 ° C and stirred for 3 hours. After cooling to room temperature, it was neutralized by adding 1N hydrochloric acid (20 mL), and extracted with dichloromethane (10 mL x 3). The combined organic layers were washed with brine, dried over anhydrous sodium sulfate Methyl]cyclobutanecarboxylic acid (800.00 mg, yellow solid), yield: 84%. MS-ESI calcd for [M ⁺ H] + 293, found 293.

Seventh step

3-[(3,7-Dimethyl-2,6-dioxo-indol-1-yl)methyl]-N-methoxy-N-methylcyclobutanecarboxamide

3-[(3,7-Dimethyl-2,6-dioxo-indol-1-yl)methyl]cyclobutanecarboxylic acid (300 mg, 1.03 mmol), N,O-dimethylhydroxylamine Hydrochloride (200 mg, 2.05 mmol), 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride (394 mg, 2.06 mmol), 1-hydroxybenzotriazole (27.8 mg, 0.206 mmol) and triethylamine (312 mg, 3.09 mmol) were dissolved in dichloromethane (10 mL). Stir at 25 ° C for 12 hours. The reaction solution was concentrated under reduced pressure and purified (yield ethyl acetate, Rf value = 0.3) to afford 3-[(3,7-dimethyl-2,6-dioxo-indol-1-yl). Base: -N-methoxy-N-methylcyclobutanecarboxamide (200 mg, yellow solid), yield: 58%. MS-ESI calculated [M+H] ^{+ s} .

Eighth step

1-[(3-acetylcyclobutyl)methyl]-3,7-dimethylindole-2,6-dione

3-[(3,7-Dimethyl-2,6-dioxo-indol-1-yl)methyl]-N-methoxy-N-methylcyclobutanecarboxamide (300 mg, 0.894 Methyl) was dissolved in tetrahydrofuran (10 mL). Methylmagnesium bromide (3M in diethyl ether, 1.49 mL, 4.47 mmol) was added dropwise at 0 ° C for 3 hours. The reaction mixture was quenched with EtOAc EtOAc (EtOAc) The organic phase was combined, washed with brine, dried over anhydrous sodium sulfate, filtered, evaporated, evaporated, evaporated, evaporated Methyl]-3,7-dimethylindole-2,6-dione (200 mg, yellow solid), yield: 77%.

MS-ESI calcd for [M+H] ⁺ 291.

Step 9

1-[(3-Acetylcyclobutyl)methyl]-3,7-dimethylindole-2,6-dione (250 mg, 0.861 mmol), cesium fluoride (130 mg, 0.861 mmol) Trimethyl-trifluoromethyl-silane (244 mg, 1.72 mmol) was added to tetrahydrofuran (10 mL) at room temperature and stirred for 12 hours. The reaction was quenched by the addition of water (20 mL). Extract with ethyl acetate (10 mL x 3). The combined organic layers were washed with brine, dried over anhydrous sodium sulfate Purification by preparative high performance liquid chromatography gave product 1 (65.0 mg, yellow solid) (isomer, first peak), yield: 20%. ^{1 H NMR: (400MHz, Methonal} -d 4) δ8.21 (s, 1H), 4.23 (d, J = 7.6Hz, 2H), 4.03 (s, 3H), 3.55 (s, 3H), 3.26-3.19 (m, 2H), 2.63-2.56 (m, 2H), 2.55-2.42 (m, 2H), 1.82-1.78 (m, 3H). MS-ESI calcd [M+H] ⁺ 361.

Product 2 (isomer 2, second peak), yield: 22%. ^{1 H NMR: (400MHz, Methonal} -d 4) δ8.05 (s, 1H), 4.01-3.99 (m, 5H), 4.03 (s, 3H), 3.54 (s, 3H), 2.71-2.66 (m, 1H), 2.55-2.54 (m, 1H), 2.17-2.12 (m, 2H), 2.02-1.98 (m, 2H). MS-ESI calcd [M+H] ⁺ 361.

Example 10

first step

Methyl 1,4-dioxaspiro[4.4]decane-7-carboxylate

Methyl 3-oxo-cyclopentanoate (16.0 g, 110 mmol), p-toluenesulfonic acid (14.0 g, 220 mmol) and ethylene glycol (969 mg, 5.60 mmol) were dissolved in anhydrous toluene (160 mL). The water separator was heated and refluxed for 4 hours. The reaction was quenched with water (2OmL)EtOAc. The combined organic layers were washed with EtOAc EtOAc EtOAc. The filtrate was concentrated under reduced pressure and purified by silica gel column chromatography (5:1 petroleum ether/ethyl acetate, Rf=0.3) 1,4-dioxaspiro[4.4]decane-7-carboxylic acid methyl ester (6.20) g, yellow oil), yield: 29%. ¹ H NMR: (400 MHz, CDCl ₃ ) δ 3.93-3.89 (m, 4H), 3.69 (s, 3H), 2.91-2.89 (m, 1H), 2.11-1.82 (m, 6H). MS-ESI calcd for [M+H] ⁺ 187.

Second step

(1,4-Dioxaspiro[4.4]decane-7-yl)-methanol

Methyl 1,4-dioxaspiro[4.4]nonane-7-carboxylate (1.00 g, 10.7 mmol) was dissolved in anhydrous tetrahydrofuran (30 mL), and the mixture was stirred under nitrogen. (531 mg, 13.9 mmol). The reaction solution was slowly warmed to 25 ° C and stirred for 3 hours. Water (0.5 mL), 15% sodium hydroxide solution (0.5 mL), and water (1.5 mL) were sequentially added to the mixture. The insoluble material was removed by filtration, and the filtrate was concentrated under reduced pressure to give (4-dioxaspiro[4.4]decane-7-yl)-methanol (m. ¹ H NMR: (400 MHz, CDCl ₃ ) δ 3.94 - 3.89 (m, 4H), 3.58 - 3.57 (m, 2H), 2.31-1.48 (m, 7H). MS-ESI calcd [M+H] ⁺ 159.

third step

1,4-Dioxaspiro[4.4]decane-7-ylmethyl methanesulfonate

(1,4-Dioxaspiro[4.4]decane-7-yl)-methanol (500 mg, 53.1 mmol) and triethylamine (800 mg, 7.92 mmol) were dissolved in anhydrous dichloromethane (5 mL). Under nitrogen, methanesulfonyl chloride (433 mg, 3.80 mmol) was slowly added at 0 °C. The reaction solution was raised to 25 ° C and stirred for 2 hours. The reaction was quenched with water (40 mL) andEtOAc. The combined organic phases were washed with water (20 mL×2), EtOAc (EtOAc) 4.4] decane-7-ylmethyl ester (800 mg, yellow oil). MS-ESI calcd for [M ⁺ H] + 237, found 237.

the fourth step

(1,4-Dioxaspiro[4.4]decane-7-ylmethyl)-3,7-dimethyl 1H-indole-2,6(3H,7H)-dione

The 1,4-dioxaspiro[4.4]decane-7-ylmethyl methanesulfonate (300 mg, 1.27 mmol) was dissolved in anhydrous N,N-dimethylformamide (10 mL). Potassium carbonate (350 mg, 2.54 mmol), potassium iodide (21.0 mg, 0.130 mmol), 2,6-hydroxy-3,7-dimethylindole (275 mg, 1.52 mmol) were added at 25 °C. The reaction solution was heated to 130 ° C and stirred for 3 hours. The reaction mixture was cooled to 25 ° C, then quenched with water (40 mL). The organic phase was combined, washed with a saturated sodium chloride solution (100 mL×2), dried over anhydrous magnesium sulfate, filtered, and the filtrate was concentrated under vacuo to give (1,4-dioxaspiro[4.4]decane-7-yl. 3,7-Dimethyl 1H-indole-2,6(3H,7H)-dione (200 mg, white solid), yield: 45%. MS-ESI calculated [M+H] ⁺ 353.

the fifth step

3,7-Dimethyl-1-(3-oxo-cyclopentylmethyl)-1H-indole-2,6(3H,7H)-dione

(1,4-Dioxaspiro[4.4]decane-7-ylmethyl)-3,7-dimethyl 1H-indole-2,6(3H,7H)-dione (200 mg, 0.620 mmol) Dissolved in anhydrous tetrahydrofuran (5 mL), protected with nitrogen, and concentrated hydrochloric acid (3 mL) at 25 °C. The reaction solution was stirred at 25 ° C for 1 hour. It was diluted with water (60 mL), and the mixture was extracted with ethyl acetate (20 mL). The organic phase was combined, washed with aq. EtOAc EtOAc (EtOAc m. 0.3), 3,7-dimethyl-1-(3-oxo-cyclopentylmethyl)-1H-indole-2,6(3H,7H)-dione (100 mg, yellow oil) was obtained. Rate: 57%. MS-ESI calcd for [M+H] ⁺ 277.

Step 6

1,3 trans-1-((3-hydroxy-3-(trifluoromethyl)cyclopentyl)methyl)-3,7-dimethyl-1H-indole-2,6(3H,7H) -dione 1,3 cis-1-((3-hydroxy-3-(trifluoromethyl)cyclopentyl)methyl)-3,7-dimethyl-1H-indole-2,6 (3H ,7H)-dione

3,7-Dimethyl-1-((3-oxocyclopentyl)methyl)-1H-indole-2,6(3H,7H)-dione (100 mg, 0.362 mmol) and cesium fluoride (11.0 mg, 0.0725 mmol) was dissolved in anhydrous tetrahydrofuran (3 mL) and trifluoromethyltrimethylsilane (95.0 mg, 0.640 mmol). The reaction solution was slowly heated to 30 ° C and stirred for 12 hours. Aqueous hydrochloric acid (1 N, 5 mL) was added to the mixture and the mixture was stirred at 30 ° C for 0.5 hour. Water (50 mL) was added to the mixture and the mixture was evaporated. The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and evaporated.

Product 1 (isomer 1, first peak) (40.0 mg, white solid), yield: 32%. ¹ H NMR: (400 MHz, Methonal-d ₄ ) δ 7.68 (s, 1H), 4.13-4.08 (m, 2H), 4.05 (s, 3H), 3.61 (s, 3H), 2.80-2.78 (m, 1H), 2.40-2.24 (m, 1H), 2.04-2.03 (m, 1H), 2.01-1.87 (m, 2H), 1.84-1.76 (m, 1H), 1.62-1.60 (m, 1H). MS-ESI calcd for [M ⁺ H] + 347, found 347.

Product 2 (isomer 2, second peak) (20.0 mg, white solid), yield: 16%. ^{1 H NMR: (400MHz, Methonal} -d 4) δ7.62 (s, 1H), 4.22-4.18 (m, 1H), 4.05-4.04 (m, 1H), 4.00 (s, 3H), 3.63 (s, 3H), 2.65-2.63 (m, 1H), 2.09-2.01 (m, 4H), 1.70-1.68 (m, 1H), 1.67-1.65 (m, 1H). MS-ESI calcd for [M ⁺ H] + 347, found 347.

Example 11

1-((4-Hydroxy-4-(trifluoromethyl)cyclohexyl)methyl)-3,7-dimethyl-1H-indole-2,6(3H,7H)-dione

first step

Ethyl 1,4-dioxaspiro[4,5]decane-8-carboxylate

Ethyl 4-oxocyclohexanecarboxylic acid (30.0 g, 176 mmol), ethylene glycol (22.0 g, 353 mmol) and p-toluenesulfonic acid (304 mg, 1.70 mmol) were dissolved in toluene (315 mL). The mixture was heated to reflux overnight. The reaction mixture was cooled to 25 ° C, and washed with water (300 mL EtOAc) 1:1 petroleum ether / ethyl acetate, Rf = 0.3) gave the product ethyl 1,4-dioxaspiro[4,5]nonane-8-carboxylic acid ethyl ester (37.2 g, yellow liquid), yield :99%. MS-ESI calcd for [M ⁺ H] + 215, found 215.

Second step

1,4-Dioxaspiro[4,5]decane-8-ylmethanol

Lithium tetrahydroaluminum (2.30 g, 61.0 mmol) was slowly added to tetrahydrofuran (60 mL) under nitrogen at 0 ° C, and ethyl 1,4-dioxaspiro[4,5]nonane-8-carboxyl was added dropwise. A solution of ethyl acetate (10.0 g, 42.0 mmol) in tetrahydrofuran (40 mL). The reaction was slowly raised to 25 ° C and stirred for 3.5 hours. The reaction solution was cooled to 0 ° C, and water (2.3 g, 127 mmol), 15% sodium hydroxide (2.3 g, 8.60 mmol) and water (6.9 g, 383 Filtration, the filter cake was washed with tetrahydrofuran (50 mL×3), the organic phase was combined, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to give the product 1,4-dioxaspiro[4,5]nonane-8- Methanol (6.22 g, yellow liquid), yield: 89%. MS-ESI calcd for [M+H] ⁺ 173.

third step

1,4-Dioxaspiro[4,5]decane-8-ylmethyl methanesulfonate

1,4-Dioxaspiro[4,5]decane-8-ylmethanol (2.00 g, 12.0 mmol) and diisopropylethylamine (3.10 g, 24.0 mmol) were dissolved in dichloromethane (40 mL) Methanesulfonyl chloride (3.90 g, 30.0 mmol) was slowly added at 0 °C. The reaction solution was raised to 25 ° C and stirred overnight. The reaction was quenched with EtOAc (EtOAc)EtOAcEtOAc The organic phase was combined, dried over anhydrous magnesium sulfate, filtered, evaporated, evaporated, evaporated, evaporated. [4,5]decane-8-ylmethyl methanesulfonate (1.80 g, yellow liquid), yield: 60%. MS-ESI calcd for [M ⁺ H] + 251, found 251.

the fourth step

1-(1,4-Dioxaspiro[4,5]decane-8-ylmethyl)-3,7-dimethyl-1H-indole-2,6(3H,7H)-dione

1,4-Dioxaspiro[4,5]decane-8-ylmethyl methanesulfonate (1.50 g, 6.00 mmol), 3,7-dimethyl-1H-indole-2,6 ( 3H,7H)-dione (1.00 g, 6.00 mmol) and potassium carbonate (2.50 g, 18.0 mmol), potassium iodide (100 mg, 0.600 mmol) dissolved in N,N-dimethylformamide (20 mL) Heat to 130 ° C and stir for 3 hours. The reaction solution was cooled to 25 ° C, then EtOAc (EtOAc) The organic phase was combined, dried over anhydrous magnesium sulfate, filtered, evaporated, evaporated, evaporated, evaporated. Oxaspiro[4,5]decane-8-ylmethyl)-3,7-dimethyl-1H-indole-2,6(3H,7H)-dione (1.75 g, white solid) Rate: 63%. MS-ESI calcd for [M ⁺ H] + 335, found 335.

the fifth step

3,7-Dimethyl-1-((4-oxocyclohexyl)methyl)-1H-indole-2,6(3H,7H)-dione

1-(1,4-Dioxaspiro[4,5]decane-8-ylmethyl)-3,7-dimethyl-1H-indole-2,6(3H,7H)-dione (1.50 g, 4.50 mmol) was dissolved in EtOAc (15 mL). The reaction was stirred at rt EtOAc (EtOAc)EtOAc. The organic phase was combined, dried over anhydrous sodium sulfate, filtered, evaporated,jjjjjjjjjjjjjjjjj ((4-Oxocyclohexyl)methyl)-1H-indole-2,6(3H,7H)-dione (1.02 g, white solid). MS-ESI calcd for [M+H] ⁺ 291.

Step 6

1-((4-Hydroxy-4-(trifluoromethyl)cyclohexyl)methyl)-3,7-dimethyl-1H-indole-2,6(3H,7H)-dione

3,7-Dimethyl-1-((4-oxocyclohexyl)methyl)-1H-indole-2,6(3H,7H)-dione (100 mg, 0.330 mmol) and cesium fluoride (60.0) The mg, 0.350 mmol) was dissolved in tetrahydrofuran (5 mL) and trifluoromethyltrimethylsilane (75.0 mg, 0.500 mmol) was slowly added under nitrogen. The reaction solution was stirred at 30 ° C for 3 hours. After cooling to 25 ° C, aqueous hydrochloric acid (4N, 3 mL) was added and stirred at 25 ° C for half an hour, pH was adjusted to 7, diluted with water and extracted with ethyl acetate (20 mL x 3). The organic phase was combined, dried over anhydrous sodium sulfate, filtered, evaporated, evaporated , 7-Dimethyl-1H-indole-2,6(3H,7H)-dione (24.0 mg, white solid), yield: 39%.

^{1 H NMR: (400MHz, Methonal} -d 4) δ7.86 (s, 1H), 4.04 (d, J = 7.2Hz, 1H), 3.97 (s, 3H), 3.89 (d, J = 7.6Hz, 1H ), 3.53 (s, 3H), 2.06-1.97 (m, 2H), 1.88-1.77 (m, 3H), 1.62-1.43 (m, 4H). MS-ESI calcd [M+H] ⁺ 361.

Example 12

first step

Ethyl 4-hydroxy-4-(trifluoromethyl)cyclohexanecarboxylate

Ethyl 4-oxocyclohexanecarboxylate (10.0 g, 58.7 mmol) was dissolved in tetrahydrofuran (100 mL), trifluoromethyltrimethylsilane (12.5 g, 88.1 mmol) and cesium fluoride ( 8.92 g, 58.7 mmol). The reaction mixture was stirred at room temperature for 12 hrs, then EtOAc (EtOAc, EtOAc (EtOAc) Drying with anhydrous sodium sulfate, concentrating by filtration and purification by silica gel column chromatography (10:1 petroleum ether / ethyl acetate, Rf = 0.5) to give 4-hydroxy-4-(trifluoromethyl)cyclohexanecarboxylic acid Ethyl ester (12.0 g, colorless oil), yield: 85%. ¹ H NMR: (400 MHz, Methanol-d ₄ ) δ 4.20 - 4.12 (m, 2H), 2.03-1.86 (m, 9H), 1.29 - 1.25 (m, 3H).

MS-ESI calcd for [M+H] ⁺ 241.

Second step

4-(hydroxymethyl)-1-(trifluoromethyl)cyclohexanol

Ethyl 4-hydroxy-4-(trifluoromethyl)cyclohexanecarboxylate (12.00 g, 49.9 mmol) was dissolved in tetrahydrofuran (20 mL). Reaction for 2 hours. The reaction was quenched by the addition of water (30 mL). It was extracted with ethyl acetate (50 mL×3), dried over anhydrous sodium sulfate, filtered, evaporated, evaporated, evaporated. -(Hydroxymethyl)-1-(trifluoromethyl)cyclohexanol (9.00 g, colorless oil), yield: 91%. ¹ H NMR: (400 MHz, Methanol-d ₄ ) δ 3.58- 3.40 (m, 2H), 1.90-1.40 (m, 9H).

third step

(4-hydroxy-4-(trifluoromethyl)cyclohexyl)methyl methanesulfonate

(4-(Hydroxymethyl)-1-(trifluoromethyl)cyclohexanol (11.0 g, 55.5 mmol) and triethylamine (1.18 g, 11.6 mmol) were dissolved in dichloromethane (80 mL) Methanesulfonyl chloride (14.4 g, 125 mmol) was added at rt. The reaction mixture was stirred at room temperature for 2 hr, then diluted with methylene chloride (60 mL). Filtration, the filtrate was concentrated under reduced pressure and purified by silica gel column chromatography (4:1 petroleum ether / ethyl acetate, Rf = 0.5) to give (4-hydroxy-4-(trifluoromethyl)cyclohexyl)methylmethyl Sulfonic acid ester (13.00 g, colorless oil), yield: 85%. ¹ H NMR: (400 MHz, Methanol-d ₄ ) δ 4.25-4.01 (m, 2H), 3.10-3.07 (m, 3H) , 2.03-1.24 (m, 9H).

the fourth step

1-(((1S,4S)-4-hydroxy-4-(trifluoromethyl)cyclohexyl)methyl)-3,7-dimethyl-1H-indole-2,6(3H,7H)- Diketone

(4-Hydroxy-4-(trifluoromethyl)cyclohexyl)methyl methanesulfonate (10.0 g, 36.2 mmol) was dissolved in N,N-dimethylformamide (100 mL). 3,7-Dimethyl-1H-indole-2,6(3H,7H)-dione (6.52 g, 36.2 mmol), potassium carbonate (7.50 g, 54.3 mmol) and potassium iodide (184 mg, 1.11 mmol) were added. ). The reaction mixture was heated to 100 ° C, and the reaction was concentrated for 5 hours. The reaction mixture was evaporated. EtOAcjjjjjjjjjjjjjj Separation by preparative SFC, separation conditions: column: Chiralpak AD-3 150x4.6mmI.D., 3um mobile phase: ethanol (0.05% diethylamine) in CO ₂ from 5% to 40%at 2.5mL/ Min wavelength: 220 nm gave product 1 (2.5 g, white solid) (isomer 1, first peak), yield: 19%. ¹ H NMR: (400MHz, Methanol-d ₄ ) δ 7.88 (s, 1H), 4.02 (d, J = 7.6 Hz, 2H), 3.98 (s, 3H), 3.53 (s, 3H), 2.16-2.02 (m, 1H), 1.99-1.98 (m, 2H), 1.87-1.80 (m, 2H), 1.60-1.49 (m, 2H), 1.48-1.46 (m, 2H). MS-ESI calcd [M+H] ⁺ 361.

Product 2 (2.40 g, white solid) (isomer 2, second peak), yield: 19%. ¹ H NMR: (400 MHz, CDCl ₃ ) δ 7.88 (s, 1H), 3.99 (s, 3H), 3.90 (d, J = 7.6 Hz, 2H), 3.54 (s, 3H), 1.84-1.81 (m) , 3H), 1.58-1.46 (m, 6H). MS-ESI calcd [M+H] ⁺ 361.

Example 13

1-((4-Hydroxy-4-methylcyclohexyl)methyl)-3,7-dimethyl-1H-indole-2,6(3H,7H)-dione

3,7-Dimethyl-1-((4-oxocyclohexyl)methyl)-1H-indole-2,6(3H,7H)-dione (50.0 mg, 0.170 mmol) was dissolved in tetrahydrofuran (2 mL) )in. Under a nitrogen atmosphere, methyl Grignard reagent (3M diethyl ether solvent, 0.4 mL, 1.20 mmol) was slowly added at -78 °C. The reaction solution was stirred at -78 ° C for 0.5 hours, slowly warmed to 0 ° C and stirred for 0.5 hours. It was quenched by the addition of a saturated solution of ammonium chloride, and the pH was adjusted to 7. It was extracted with ethyl acetate, dried over anhydrous sodium sulfate and filtered. The filtrate was concentrated under reduced pressure and purified by preparative high-performance liquid chromatography to give 1-((4-hydroxy-4-methylcyclohexyl)methyl)-3,7-dimethyl-1H-indole-2,6 (3H, 7H)-Dione (8.0 mg, white solid), yield: 16%. ^{1 H NMR: (400MHz, Methonal} -d 4) δ7.86 (s, 1H), 3.97 (s, 3H), 3.88 (d, J = 7.6Hz, 2H), 3.52 (s, 3H), 1.85-1.78 (m, 1H), 1.73-1.57 (m, 3H), 1.46-1.33 (m, 2H), 1.32-1.15 (m, 6H). MS-ESI calcd for ^{_{[M + HH 2 O] +}} 289, found 289.

Example 14

1-((4-ethyl-4-hydroxycyclohexyl)methyl)-3,7-dimethyl-1H-indole-2,6(3H,7H)-dione

3,7-Dimethyl-1-((4-oxocyclohexyl)methyl)-1H-indole-2,6(3H,7H)-dione (50.0 mg, 0.170 mmol) was dissolved in tetrahydrofuran (2 mL) )in. Under a nitrogen atmosphere, ethyl Grignard reagent (3M in ether solvent, 0.4 mL, 1.20 mmol) was slowly added at -78 °C. The reaction solution was stirred at -78 ° C for 0.5 hours, slowly warmed to 0 ° C and stirred for 0.5 hours. It was quenched by the addition of a saturated solution of ammonium chloride, and the pH was adjusted to 7. It was extracted with ethyl acetate, dried over anhydrous sodium sulfate and filtered. The filtrate was concentrated under reduced pressure and purified by preparative high-performance liquid chromatography to give 1-((4-ethyl-4-hydroxycyclohexyl)methyl)-3,7-dimethyl-1H-indole-2,6 (3H, 7H)-Dione (42.0 mg, white solid), yield: 77%. ^{1 H NMR: (400MHz, Methonal} -d 4) δ7.86 (s, 1H), 3.97 (s, 3H), 3.88 (d, J = 7.6Hz, 2H), 3.54-3.50 (m, 3H), 1.93 -1.80 (m, 1H), 1.76-1.72 (m, 2H), 1.66-1.51 (m, 3H), 1.38-1.28 (m, 3H), 1.27-1.13 (m, 2H), 0.89 (t, J = 7.2 Hz, 3H). MS-ESI calcd for ^{_{[M + HH 2 O] +}} 303, found 303.

Example 15

first step

Methyl 2-(1,4-dioxaspiro[4.5]decane-8-ylidene)acetate

Triethyl phosphonoacetate (12.2 g, 54.4 mmol) was dissolved in tetrahydrofuran (100 mL), and sodium hydride (1.92 g, 48.0 mmol) was added portionwise at 0 ° C, and the reaction was stirred for 30 min under nitrogen atmosphere. A solution of 1,4-cyclohexanedione monoethylene ketal (5.00 g, 32.0 mmol) dissolved in tetrahydrofuran (15 mL) was added dropwise to the reaction solution at 0 ° C, and the reaction solution was stirred at 25 ° C for 3 hours. . The reaction was quenched with water (25 mL) andEtOAcEtOAc. The organic phase was combined, washed with brine (20 mL), dried over anhydrous sodium sulfate sulfatessssssssssssssssssssssssss Methyl 2-(1,4-dioxaspiro[4.5]decane-8-ylidene)acetate (6.30 g, colorless oil), yield: 93%. ^{1 H NMR: (400MHz, CDCl} 3) δ5.67 (s, 1H), 4.15 (q, J = 7.2Hz, 2H), 3.98 (s, 4H), 3.00 (t, J = 6.4Hz, 2H), 2.38 (t, J = 6.4 Hz, 2H), 1.84-1.68 (m, 4H), 1.28 (t, J = 7.2 Hz, 3H). MS-ESI calcd for [M ⁺ H] + 227, found 227.

Second step

Ethyl 2-(1,4-dioxaspiro[4.5]decane-8-yl)ethyl acetate

Methyl 2-(1,4-dioxaspiro[4.5]decane-8-ylidene)acetate (3.80 g, 17.9 mmol) was dissolved in methanol (50 mL) and dry palladium carbon (palladium 10%) , water 1%, 400 mg), the reaction solution was reacted under hydrogen (50 psi) for 18 hours at room temperature. The reaction mixture was filtered, and the filtrate was evaporated tolulujjjjjjjjjjjj .

¹ H NMR: (400MHz, CDCl ₃ ) δ 4.12 (q, J = 7.2 Hz, 2H), 3.93 (s, 4H), 2.22 (d, J = 7.2 Hz, 2H), 1.90-1.64 (m, 5H) ), 1.63-1.48 (m, 2H), 1.40-1.16 (m, 5H). MS-ESI calcd for [M+H] ⁺ 229.

third step

2-(1,4-Dioxaspiro[4.5]decane-8-yl)ethanol

Ethyl 2-(1,4-dioxaspiro[4.5]decane-8-yl)acetate (1.00 g, 4.38 mmol) was dissolved in tetrahydrofuran (20 mL), and tetrahydrolithium was added portionwise at 0 °C. Aluminum (216 mg, 5.69 mmol) was stirred under nitrogen for 18 hours. The reaction solution was cooled to 0 ° C, and water (0.2 mL), 15% aqueous sodium hydroxide (0.2 mL) and water (0.6 mL) were added slowly. Filtration and concentration of the filtrate under reduced pressure afforded 2-(4-dioxaspiro[4.5]decane-8-yl)ethanol (780 mg,yield of yellow oil).

^{1 H NMR: (400MHz, CDCl} 3) δ3.94 (s, 4H), 3.69 (t, J = 6.4Hz, 2H), 1.79-1.65 (m, 4H), 1.59-1.38 (m, 5H), 1.34 -1.17 (m, 2H). MS-ESI calcd for [M+H] ⁺ 187.

the fourth step

2-(1,4-Dioxaspiro[4.5]decane-8-yl)ethyl methanesulfonate

2-(1,4-Dioxaspiro[4.5]decane-8-yl)ethanol (400 mg, 2.15 mmol) and triethylamine (435 mg, 4.30 mmol) were dissolved in dichloromethane (10 mL) Methanesulfonyl chloride (369 mg, 3.23 mmol) was slowly added at 0 °C. The reaction solution was stirred at 0 ° C for 4 hours. The reaction was quenched with water (10 mL)EtOAcEtOAc The organic phase was combined, washed with aq. NaHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHH Ethyl mesylate (500 mg crude, yellow oil).

¹ H NMR: (400 MHz, CDCl ₃ ) δ 4.28 (t, J = 6.4 Hz, 2H), 3.94 (s, 4H), 3.01 (s, 3H), 1.76-1.63 (m, 6H), 1.60-1.43 (m, 3H), 1.37-1.21 (m, 2H). MS-ESI calcd [M+H] ⁺ 265.

the fifth step

1-(2-(1,4-Dioxaspiro[4.5]decane-8-yl)ethyl)-3,7-dimethyl-1H-indole-2,6-(3H,7H)- Diketone

3,7-Dimethyl-1H-indole-2,6-(3H,7H)-dione (204 mg, 1.13 mmol) was dissolved in N,N-dimethylformamide (15 mL). 1,4-Dioxaspiro[4.5]decane-8-yl)ethyl methanesulfonate (300 mg, 1.13 mmol), potassium carbonate (312 mg, 2.26 mmol) and potassium iodide (225 mg, 1.36 mmol). The reaction solution was heated to 120 ° C and stirred for 3 hours. Concentration under reduced pressure, the residue was purified mjjjjjjjjjjjjj -yl)ethyl]-3,7-dimethylindole-2,6-dione (190 mg, white solid), yield: 48%.

^{1 H NMR: (400MHz, CDCl} 3) δ7.50 (s, 1H), 4.09-4.03 (m, 2H), 4.02 (s, 3H), 3.99 (s, 4H), 3.57 (s, 1H), 1.90 -1.70 (m, 5H), 1.68-1.47 (m, 6H), 1.45-1.31 (m, 2H). MS-ESI calcd for [M+H] ⁺ 349.

Step 6

3,7-Dimethyl-1-(2-(4-oxocyclohexyl)ethyl)-1H-indole-2,6-(3H,7H)-dione

Dissolve 1-[2-(1,4-dioxaspiro[4.5]decane-8-yl)ethyl]-3,7-dimethylindole-2,6-dione (190 mg, 545 umol) Concentrated hydrochloric acid (1 mL) was added to tetrahydrofuran (3 mL). The reaction solution was stirred at room temperature for 18 hours. The reaction mixture was concentrated under reduced pressure. EtOAcjjjjjjjjjjjjjjjjjjjjjjjjjjjjjjjjjjjjjjjjjjjjjjjjjjjjjjjjjjjjjjjjj Filtration, concentrating under reduced pressure, EtOAc (EtOAc m. 1H-indole-2,6-(3H,7H)-dione (150 mg, colorless oil), yield: 90%.

MS-ESI calcd for [M+H] ⁺ 303.

Seventh step

3,7-Dimethyl-1-(2-(4-(trifluoromethyl)-4-((trimethylsilyl)oxy)cyclohexyl)ethyl)-1-indole-2, 6-(3H,7H)-dione

Dissolving 3,7-dimethyl-1-[2-(4-oxocyclohexyl)ethyl]indole-2,6-dione (145 mg, 0.476 mmol) and cesium fluoride (7.2 mg, 0.0476 mmol) Trifluoromethyltrimethylsilane (203 mg, 1.43 mmol) was slowly added in tetrahydrofuran (10 mL) under nitrogen. The reaction solution was stirred at 25 ° C for 18 hours. The mixture was diluted with water (20 mL), EtOAc (EtOAc) -Dimethyl-1-(2-(4-(trifluoromethyl)-4-((trimethylsilyl)oxy)cyclohexyl)ethyl)-1-in-2,6-( 3H,7H)-dione (170 mg, colorless liquid), yield: 80%.

MS-ESI calcd for [M+H] ⁺ 447.

Eighth step

3,7-Dimethyl-1-[2-[4-(trifluoromethyl)-4-trimethylsiloxy-cyclohexyl]ethyl]indole-2,6-dione (160 mg, 0.358 mmol) was dissolved in tetrahydrofuran (3 mL) and concentrated hydrochloric acid (12M, 0.107 mL). The reaction solution was stirred at 25 ° C for 18 hours. Diluted with water, aq. sodium hydrogen carbonate (10 mL) was adjusted to pH 7 and extracted with ethyl acetate (10 mL x 2). The organic phase was combined, dried over anhydrous sodium sulfate, filtered, evaporated, evaporated,jjjjjjjjjjjjjjjj %. ^{1 H NMR: (400MHz, CDCl} 3) δ8.01 (s, 1H), 4.09-3.94 (m, 5H), 3.53 (s, 3H), 1.97-1.79 (m, 4H), 1.76-1.62 (m, 3H), 1.61-1.45 (m, 4H). MS-ESI calcd for [M+H] ⁺ 372. And product 2 (15.0 mg, white solid) (isomer 2, second peak), yield: 10%. ^{1 H NMR: (400MHz, CDCl} 3) δ8.01 (s, 1H), 4.09-3.95 (m, 5H), 3.53 (s, 3H), 1.87-1.68 (m, 4H), 1.64-1.48 (m, 4H), 1.46-1.25 (m, 3H). MS-ESI calcd [M+H]+ 375.

Example 16

1-((4-Hydroxy-1-methyl-4-(trifluoromethyl)cyclohexyl)methyl)-3,7-dimethyl-1H-indole-2,6-(3H,7H)- Diketone

first step

Ethyl-8-methyl-1,4-dioxaspiro[4.5]decane-8-carboxylic acid tert-butyl ester

Ethyl ethyl 1,4-dioxaspiro[4.5]decane-8-carboxylate (5.00 g, 23.3 mmol) was dissolved in anhydrous tetrahydrofuran (100 mL). A lithium diisopropylamide solution (2M tetrahydrofuran solution, 14.0 mL, 28.0 mmol) was added, and the mixture was stirred at -78 ° C for one hour. Methyl iodide (6.62 g, 46.7 mmol) was slowly added and stirring was continued for 1 hour. The reaction was quenched by the addition of water (100 mL). The reaction mixture was extracted with EtOAc (EtOAc (EtOAc)EtOAc. 0.4) Obtained ethyl 8-methyl-1,4-dioxaspiro[4.5]decane-8-carboxylic acid tert-butyl ester (5.00 g, yellow oil). ¹ H NMR: (400 MHz, Methonal-d ₄ ) δ 4.16-4.10 (m, 2H), 3.93-3.86 (m, 4H), 2.13-2.06 (m, 2H), 1.61-1.48 (m, 6H), 1.25-1.22 (m, 3H), 1.15 (s, 3H).

Second step

Ethyl-1-methyl-4-oxocyclohexanecarboxylic acid

Ethyl 8-methyl-1,4-dioxaspiro[4.5]decane-8-carboxylic acid tert-butyl ester (5.00 g, 21.9 mmol) was dissolved in tetrahydrofuran (50 mL) and 1N hydrochloric acid was added dropwise at 0 °C. The aqueous solution (20 mL) was stirred at 20 ° C for 1 hour. The mixture was cooled to 0 ° C and quenched with sodium bicarbonate (50 mL). The mixture was extracted with ethyl acetate (100 mL x 3). The organic layer was washed with brine (100 mL EtOAc) Purification by silica gel column chromatography (10:1 petroleum ether / ethyl acetate, Rf = 0.4) afforded ethyl-1-methyl-4-oxocyclohexanecarboxylic acid (3.00 g, colorless oil) Rate: 74%. ¹ H NMR: (400 MHz, Methonal-d ₄ ) δ 4.26-4.11 (m, 2H), 2.46-2.29 (m, 5H), 1.74-1.55 (m, 3H), 1.33-1.26 (m, 6H).

third step

Ethyl 4-hydroxy-1-methyl-4-(trifluoromethyl)cyclohexanecarboxylic acid

Ethyl-1-methyl-4-oxocyclohexanecarboxylic acid (3.00 g, 16.3 mmol), cesium fluoride (247 mg, 1.63 mmol) was dissolved in tetrahydrofuran (50 mL), then trimethyl group was added at 0 °C. Silicotrifluoromethyl (4.63 g, 35.3 mmol). The reaction solution was reacted under nitrogen for 20 hours at 20 °C. Then 4N aqueous hydrochloric acid (4 mL) was added. The mixture was reacted under nitrogen for 6 hours at room temperature. The reaction was quenched with EtOAc EtOAc (EtOAc (EtOAc) Petroleum ether / ethyl acetate, Rf = 0.3) gave ethyl 4-hydroxy-1-methyl-4-(trifluoromethyl)cyclohexanecarboxylic acid (3.00 g, colourless oil). %. ¹ H NMR: (400 MHz, Methanol-d ₄ ) δ 4.20 - 4.12 (m, 2H), 2.03-1.31 (m, 8H), 1.29-1.23 (m, 6H).

the fourth step

4-(hydroxymethyl)-4-methyl-1-(trifluoromethyl)cyclohexanol

Ethyl 4-hydroxy-1-methyl-4-(trifluoromethyl)cyclohexanecarboxylic acid (3.00 g, 11.8 mmol) was dissolved in anhydrous tetrahydrofuran (50 mL), and lithium tetrahydroaluminum was added at 0 ° C. (896 mg, 23.6 mmol). The reaction solution was warmed to 25 ° C and stirred for 1 hour. Quenched with water (20 mL), EtOAc (EtOAc m. 0.2), 4-(hydroxymethyl)-4-methyl-1-(trifluoromethyl)cyclohexanol (2.00 g, mp. ¹ H NMR: (400 MHz, Methanol-d ₄ ) δ 3.25 (s, 2H), 1.76-1.64 (m, 6H), 1.29-1.26 (m, 2H), 0.93-0.91 (m, 3H).

the fifth step

(4-hydroxy-1-methyl-4-(trifluoromethyl)cyclohexyl)methyl methanesulfonate

4-(Hydroxymethyl)-4-methyl-1-(trifluoromethyl)cyclohexanol (2.00 g, 9.42 mmol) was dissolved in dichloromethane (30 mL) and triethylamine was added at 0 °C (953 mg, 9.42 mmol) and methanesulfonyl chloride (1.08 g, 9.42 mmol). The reaction solution was reacted at 0 ° C for 2 hours. The organic layer was extracted with a saturated aqueous solution of sodium chloride (50 mL×3), washed with a saturated sodium chloride solution (50 mL×3), dried over anhydrous sodium sulfate Concentration by pressure gave (4-hydroxy-1-methyl-4-(trifluoromethyl)cyclohexyl)methyl methanesulfonate (2.00 g, m.

Step 6

1-((4-Hydroxy-1-methyl-4-(trifluoromethyl)cyclohexyl)methyl)-3,7-dimethyl-1H-indole-2,6-(3H,7H)- Diketone

(4-Hydroxy-1-methyl-4-(trifluoromethyl)cyclohexyl)methyl methanesulfonate (100 mg, 0.344 mmol), 3,7-dimethyl-1H-indole-2,6- (3H,7H)-dione (62.1 mg, 0.344 mmol), potassium iodide (5.70 mg, 0.0344 mmol) and potassium carbonate (47.6 mg, 0.344 mmol) dissolved in anhydrous N,N-dimethylformamide (5 mL) in. The reaction solution was heated to 150 ° C in the microwave for 4 hours. The reaction solution was cooled to 20 ° C, filtered, and purified by preparative HPLC to give 1-((4-hydroxy-1-methyl-4-(trifluoromethyl)cyclohexyl)methyl)-3,7- Dimethyl-1H-indole-2,6-(3H,7H)-dione (3.0 mg, white solid), yield: 2%. ^{1 H NMR: (400MHz, Methonal} -d 4) δ7.88 (s, 1H), 3.98 (s, 3H), 3.96 (s, 2H), 3.54 (s, 3H), 1.81-1.64 (m, 6H) , 1.63-1.34 (m, 2H), 1.00 (s, 3H). MS-ESI calcd for [M+H] ⁺ 372.

Example 17

1-((4-Hydroxy-1-methyl-4-(trifluoromethyl)cyclohexyl)methyl)-3,7-dimethyl-1H-indole-2,6-(3H,7H)- Diketone

first step

Ethyl 8-(methoxymethyl)-1,4-dioxaspiro[4.5]decane-8-carboxylic acid ethyl ester

Ethyl ethyl 1,4-dioxaspiro[4.5]decane-8-carboxylate (5.00 g, 23.3 mmol) was dissolved in anhydrous tetrahydrofuran (100 mL). A lithium diisopropylamide solution (2M-hexane solution, 14.0 mL, 28.0 mmol) was added, and the mixture was stirred at -78 ° C for one hour. Methoxybromomethane (5.83 g, 46.7 mmol) was slowly added and stirring was continued for 1 hour. The reaction was quenched by the addition of water (100 mL). The reaction mixture was extracted with EtOAc (EtOAc (EtOAc)EtOAc. 0.3) Ethyl ethyl 8-(methoxymethyl)-1,4-dioxaspiro[4.5]decane-8-carboxylate (5.00 g, yellow oil). ¹ H NMR: (400MHz, Methanol-d ₄ ) δ 4.18 (q, J = 6.8 Hz, 2H), 3.94 (s, 4H), 3.55 (s, 2H), 3.33 (s, 3H), 2.14-2.12 (m, 2H), 1.65-1.57 (m, 6H), 1.26 (t, J = 6.8 Hz, 3H).

Second step

Ethyl ethyl-1-(methoxymethyl)-4-oxocyclohexanecarboxylate

Ethyl ethyl 8-(methoxymethyl)-1,4-dioxaspiro[4.5]decane-8-carboxylate (5.00 g, 19.4 mmol) was dissolved in tetrahydrofuran (50 mL) at 0 ° C After 1N diluted hydrochloric acid (10 mL) was added dropwise, the mixture was stirred at 20 ° C for 1 hour. The mixture was cooled to 0 ° C and quenched with sodium bicarbonate (50 mL). The mixture was extracted with ethyl acetate (100 mL x 3). The organic layer was washed with brine (100 mL EtOAc) Purification by silica gel column chromatography (10:1 petroleum ether / ethyl acetate, Rf = 0.4) afforded ethyl ethyl 1-(methoxymethyl)-4-oxocyclohexanecarboxylate (3.00 g, White oil), Yield: 73%. ¹ H NMR: (400MHz, Methanol-d ₄ ) δ 4.25 (q, J = 6.8 Hz, 2H), 3.52 (s, 2H), 3.34 (s, 3H), 2.52-2.30 (m, 6H), 1.82 -1.78 (m, 2H), 1.30 (t, J = 6.8 Hz, 3H).

third step

Ethyl 4-hydroxy-1-(methoxymethyl)-4-(trifluoromethyl)cyclohexanecarboxylate

Ethyl ethyl 1-(methoxymethyl)-4-oxocyclohexanecarboxylate (3.00 g, 14.0 mmol), EtOAc (EtOAc (EtOAc) Trimethylsilyltrifluoromethyl (3.98 g, 28.0 mmol) was then added at 0 °C. The reaction solution was reacted under nitrogen for 20 hours at 20 °C. Then 4N dilute hydrochloric acid (7 mL) was added. The mixture was reacted under nitrogen for 6 hours at room temperature. The reaction was quenched with EtOAc EtOAc (EtOAc (EtOAc) Petroleum ether / ethyl acetate, Rf = 0.4) to give ethyl 4-hydroxy-1-(methoxymethyl)-4-(trifluoromethyl)cyclohexanecarboxylate (1.7 g, colorless oil) , Yield: 43%. ^{1 H NMR: (400MHz, Methonal} -d 4) 4.18-4.09 (m, 2H), 3.61 (s, 2H), 3.33 (s, 3H), 1.84-1.71 (m, 8H), 1.28-1.25 (m, 3H).

the fourth step

4-(hydroxymethyl)-4-(methoxymethyl)-1-(trifluoromethyl)cyclohexanol

Ethyl 4-hydroxy-1-(methoxymethyl)-4-(trifluoromethyl)cyclohexanecarboxylate (1.50 g, 5.28 mmol) was dissolved in anhydrous tetrahydrofuran (50 mL). Lithium tetrahydroaluminum (220 mg, 5.81 mmol) was added. The reaction solution was warmed to 25 ° C and stirred for 1 hour. Quenched with water (20 mL), EtOAc (EtOAc)EtOAc. 0.2), 4-(hydroxymethyl)-4-(methoxymethyl)-1-(trifluoromethyl)cyclohexanol (1.20 g, m. ¹ H NMR: (400 MHz, Methanol-d ₄ ) δ 3.33 - 3.32 (m, 7H), 1.67-1.63 (m, 4H), 1.52-1.48 (m, 4H).

the fifth step

(4-hydroxy-1-methyl-4-(trifluoromethyl)cyclohexyl)methyl methanesulfonate

4-(Hydroxymethyl)-4-(methoxymethyl)-1-(trifluoromethyl)cyclohexanol (1.20 g, 4.95 mmol) was dissolved in dichloromethane (20 mL) at 0 ° C Triethylamine (851 mg, 9.91 mmol) and methanesulfonyl chloride (851 mg, 7.43 mmol) were added. The reaction solution was reacted at 0 ° C for 2 hours. The organic layer was extracted with a saturated aqueous solution of sodium chloride (50 mL×3), washed with a saturated sodium chloride solution (50 mL×3), dried over anhydrous sodium sulfate Concentration by pressure gave (4-hydroxy-1-methyl-4-(trifluoromethyl)cyclohexyl)methyl methanesulfonate (1.30 g, m.

Step 6

1-((4-Hydroxy-1-methyl-4-(trifluoromethyl)cyclohexyl)methyl)-3,7-dimethyl-1H-indole-2,6-(3H,7H)- Diketone

(4-Hydroxy-1-methyl-4-(trifluoromethyl)cyclohexyl)methyl methanesulfonate (300 mg, 1.05 mmol), 3,7-dimethyl-1H-indole-2,6- (3H,7H)-Dione (189 mg, 1.05 mmol), potassium iodide (17.4 mg, 0.105 mmol) and potassium carbonate (435 mg, 3.15 mmol) were dissolved in anhydrous N,N-dimethylformamide (5 mL). The reaction solution was heated to 150 ° C in the microwave for 2 hours. The reaction solution was cooled to 20 ° C, filtered, and purified by preparative HPLC to give 1-((4-hydroxy-1-methyl-4-(trifluoromethyl)cyclohexyl)methyl)-3,7- Dimethyl-1H-indole-2,6-(3H,7H)-dione (12.0 mg, white solid), yield: 3%. ^{1 H NMR: (400MHz, Methonal} -d 4) δ7.87 (s, 1H), 4.06 (s, 2H), 3.83 (s, 3H), 3.98 (s, 3H), 3.53 (s, 2H), 3.42 (s, 3H), 1.69-1.58 (m, 8H). MS-ESI calcd for [M+H] ⁺ 405.

Example 18

1-((4-(3-hydroxypenta-3-yl)-cyclohexyl)methyl)-3,7-dimethyl 1H-indole-2,6(3H,7H)-dione

first step

Methyl 4-hydroxymethylcyclohexanecarboxylate

Methyl 1,4-cyclohexanecarboxylate (1.20 g, 6.45 mmol) was dissolved in anhydrous tetrahydrofuran (20 mL), and then filtered under nitrogen, and borane dimethyl sulfide (10 M, 1.0 mL) was slowly added dropwise at 0 ° C. 10.3 mmol), the reaction solution was stirred at 0 ° C for 0.5 hour, slowly raised to 25 ° C, and stirring was continued for 1 hour. The reaction was quenched by the addition of water (40 mL). The organic phase was combined, washed with EtOAc EtOAc (EtOAc m. : 91%. ¹ H NMR: (400MHz, CDCl ₃ ) δ 3.67 (s, 3H), 3.48-3.46 (m, 2H), 2.26-2.25 (m, 1H), 2.05-2.01 (m, 2H), 1.89-1.85 ( m, 2H), 1.47-1.43 (m, 2H), 1.31 (s, 1H), 1.01 - 0.98 (m, 2H). MS-ESI calcd for [M+H] ⁺ 173.

Second step

Methyl 4-methanesulfonyloxymethyl-cyclohexanecarboxylate

Methyl 4-hydroxymethylcyclohexanecarboxylate (900 mg, 5.20 mmol) and triethylamine (1.58 g, 15.6 mmol) were dissolved in anhydrous dichloromethane (5 mL). Sulfonyl chloride (720 mg, 6.30 mmol). The reaction solution was raised to 25 ° C and stirred for 2 hours. The reaction was quenched by the addition of water (60 mL). The organic phase was combined, washed sequentially with water, EtOAc EtOAc EtOAc (HHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHHH Product 4-Methylsulfonyloxymethyl-cyclohexanecarboxylic acid methyl ester (1.00 g, white solid), yield: 91%. ¹ H NMR: (400MHz, CDCl ₃ ) δ 3.67 (s, 3H), 3.48-3.46 (m, 2H), 3.01 (s, 3H), 2.26-2.25 (m, 1H), 2.05-2.01 (m, 2H), 1.89-1.85 (m, 2H), 1.47-1.43 (m, 2H), 1.31 (s, 1H), 1.01 - 0.98 (m, 2H). MS-ESI calcd for [M ⁺ H] + 251, found 251.

third step

Methyl 4-((3,7-dimethyl-2,6-dioxo-2,3,6,7-tetrahydro-1H-indol-1-yl)methyl)-cyclohexanecarboxylate

Methyl 4-methanesulfonyloxymethyl-cyclohexanecarboxylate (580 mg, 2.32 mmol) was dissolved in 5 mL of anhydrous N,N-dimethylformamide, and potassium carbonate was added at 25 ° C under nitrogen atmosphere ( 640 mg, 4.64 mmol), potassium iodide (38.0 mg, 0.230 mmol), 2,6-hydroxy-3,7-dimethylindole (501 mg, 2.80 mmol). The reaction solution was stirred at 130 ° C for 3 hours. The mixture was extracted with ethyl acetate. ((3,7-Dimethyl-2,6-dioxo-2,3,6,7-tetrahydro-1H-indol-1-yl)methyl)-cyclohexanecarboxylic acid methyl ester (400 mg , white solid), yield: 52%. MS-ESI calcd for [M ⁺ H] + 335, found 335.

the fourth step

1-((4-(3-hydroxypenta-3-yl)-cyclohexyl)methyl)-3,7-dimethyl 1H-indole-2,6(3H,7H)-dione

Methyl 4-((3,7-dimethyl-2,6-dioxo-2,3,6,7-tetrahydro-1H-indol-1-yl)-cyclohexanecarboxylic acid (100 mg , 0.30 mmol) was dissolved in 5 mL of anhydrous tetrahydrofuran. Ethyl magnesium bromide solution (3M diethyl ether solution, 1 mL, 3.00 mmol) was slowly added dropwise at -65 ° C under nitrogen atmosphere, and the reaction mixture was stirred at -65 ° C for 2 hours. The reaction mixture was poured with water (40 mL), EtOAc (EtOAc) Purification afforded the product 1-((4-(3-hydroxypenta-3-yl)-cyclohexyl)methyl)-3,7-dimethyl 1H-indole-2,6(3H,7H)-dione (20.0 Mg, white solid), Yield: 19%. ¹ H NMR: (400 MHz, CDCl ₃ ) δ 7.52 (s, 1H), 4.00 (s, 3H), 3.90-3.88 (m, 2H), 3.59 (s , 3H), 1.80- 1.74 (m, 6H), 1.50-1.45 (m, 4H), 1.11-1.10 (m, 4H), 0.86-0.82 (m, 6H). MS ESI calculated [M+H] ⁺ 363, found 363.

Example 19

3,7-Dimethyl-1-[[trans-4-(2,2,2-trifluoro-1-hydroxy-1-methyl-ethyl)cyclohexyl]methyl]indole-2,6 -dione

first step

Trans-4-methylmethylcyclohexanecarboxylic acid methyl ester

The trans-cyclohexane-1,4-dicarboxylic acid monomethyl ester (5.00 g, 26.8 mmol) was dissolved in tetrahydrofuran (100 mL), and borane dimethyl sulfide (3.06 g, 40.3 mmol) was added at 0 °C. The reaction was carried out for 2 hours at room temperature. The reaction was quenched by the addition of saturated MeOH (50 mL). After concentrating, water (50 mL) was evaporated (EtOAc m. Oily), yield: 87%. ^{1 H NMR: (400MHz, Methonal} -d 4) δ3.67 (s, 3H), 3.43-3.38 (m, 2H), 2.31-2.54 (m, 1H), 2.03-1.98 (m, 2H), 1.90- 1.82 (m, 2H), 1.45-1.38 (m, 3H), 1.03-0.99 (m, 2H). MS-ESI calcd for [M+H] ⁺ 173.

Second step

Trans-4-methylsulfonyloxymethyl-cyclohexanecarboxylic acid methyl ester

Methyl trans-4-hydroxymethylcyclohexanecarboxylate (4.00 g, 23.2 mmol) and triethylamine (7.05 g, 69.6 mmol) were dissolved in dichloromethane (50 mL). Acid chloride (7.98 g, 69.6 mmol). The reaction solution was slowly warmed to room temperature and stirred for 2 hours. The reaction was quenched by the addition of aqueous sodium bicarbonate (50 mL). Extract with dichloromethane (20 mL x 3). The organic layer was combined, washed with EtOAc EtOAc m. Yellow oil), yield: 99%. ¹ H NMR: (400 MHz, Methonal-d ₄ ) δ 4.10-4.03 (m, 2H), 3.65 (s, 3H), 3.07 (s, 3H), 2.42-2.31 (m, 1H), 2.10-2.03 ( m, 2H), 1.90-1.82 (m, 2H), 1.75-1.66 (m, 1H), 1.48-1.42 (m, 2H), 1.21-1.10 (m, 2H). MS-ESI calcd for [M ⁺ H] + 251, found 251.

third step

Trans-methyl 4-[(3,7-dimethyl-2,6-dioxo-indol-1-yl)methyl]cyclohexanecarboxylic acid

Methyl trans-4-methanesulfonyloxymethyl-cyclohexanecarboxylate (1.00 g, 4.00 mmol), 3,7-dimethyl-1H-indole-2,6(3H,7H)- Diketone (719 mg, 4.00 mmol), potassium iodide (66.0 mg, 0.397 mmol) and potassium carbonate (1.10 g, 7.96 mmol) were dissolved in N,N-dimethylformamide (10 mL). The reaction solution was warmed to 120 ° C and stirred for 3 hours. It was cooled to room temperature, filtered, and the filtrate was concentrated. Separation and purification by silica gel column chromatography (ethyl acetate, Rf value = 0.1) gave trans-methyl 4-[(3,7-dimethyl-2,6-dioxo-indol-1-yl) Methyl]cyclohexanecarboxylic acid (800 mg, yellow solid), yield: 60%. ^{1 H NMR: (400MHz, Methonal} -d 4) δ7.88 (s, 1H), 3.98 (s, 3H), 3.90-3.82 (m, 2H), 3.72 (s, 3H), 3.51 (s, 3H) , 2.33 - 2.25 (m, 1H), 2.03-1.98 (m, 2H), 1.80-1.74 (m, 3H), 1.42-1.36 (m, 2H), 1.21-1.10 (m, 2H). MS-ESI calcd for [M ⁺ H] + 335, found 335.

the fourth step

1-(trans-4-acetylcyclohexylmethyl)-3,7-dimethyl-3,7-dihydro-indole-2,6-dione

Trans-methyl 4-[(3,7-dimethyl-2,6-dioxo-indol-1-yl)methyl]cyclohexanecarboxylic acid (300 mg, 0.897 mmol) and O, N -Methylhydroxylamine hydrochloride (114 mg, 1.17 mmol) was dissolved in THF (25 mL). The reaction solution was slowly warmed to room temperature and stirred for 12 hours. The reaction was quenched by the addition of saturated aqueous ammonium chloride (10 mL). Extract with ethyl acetate (10 mL x 3). The combined organic layers were washed with EtOAc EtOAc. Purification by preparative TLC (ethyl acetate, Rf = 0.4) gave 1-(trans-4-acetylcyclohexylmethyl)-3,7-dimethyl-3,7-dihydro-indole-2 , 6-diketone (80.0 mg, yellow oil), yield: 29%. ¹ H NMR: (400 MHz, Methonal-d ₄ ) δ 7.88 (s, 1H), 3.98 (s, 3H), 3.92-3.84 (m, 2H), 3.55 (s, 3H), 2.42-2.33 (m, 1H), 2.15 (s, 3H), 1.98-1.88 (m, 2H), 1.85-1.75 (m, 3H), 1.32-1.10 (m, 4H). MS-ESI calcd for [M+H] ⁺ 319.

the fifth step

3,7-Dimethyl-1-[[trans-4-(2,2,2-trifluoro-1-hydroxy-1-methyl-ethyl)cyclohexyl]methyl]indole-2,6 -dione

Fluorinated 1-(trans-4-acetylcyclohexylmethyl)-3,7-dimethyl-3,7-dihydro-indole-2,6-dione (80.0 mg, 0.251 mmol) The hydrazine (11.5 mg, 0.753 mmol) was dissolved in tetrahydrofuran (10 mL), and trimethyl-trifluoromethyl-silane (71.6 mg, 0.502 mmol) was added at room temperature and stirred for 12 hours. After 1 N hydrochloric acid (10 mL) was added, the mixture was stirred at room temperature for 1 hour, and then the mixture was stirred and evaporated. Extract with ethyl acetate (10 mL x 3). The combined organic layers were washed with EtOAc EtOAc. Purification by preparative high performance liquid chromatography to give 3,7-dimethyl-1-[[trans-4-(2,2,2-trifluoro-1-hydroxy-1-methyl-ethyl) Cyclohexyl]methyl]anthracene-2,6-dione (35.0 mg, yellow solid), yield: 70%. ¹ H NMR: (400 MHz, Methonal-d ₄ ) δ 7.88 (s, 1H), 3.98 (s, 3H), 3.88 (d, J = 6.8 Hz, 2H), 3.53 (s, 3H), 1.96-1.67 (m, 6H), 1.22 (s, 3H), 1.15 - 1.06 (m, 4H). MS-ESI calcd for [M ⁺ H] + 389, found 389.

Example 20

3,7-Dimethyl-1-[trans-4-(2,2,2-trifluoro-1-hydroxy-1-trifluoromethyl-ethyl)-cyclohexylmethyl]-3,7 -dihydro-indole-2,6-dione

first step

3,7-Dimethyl-1-[trans-4-(2,2,2-trifluoro-1-hydroxy-1-trifluoromethyl-ethyl)-cyclohexylmethyl]-3,7 -dihydro-indole-2,6-dione

Trans-methyl 4-[(3,7-dimethyl-2,6-dioxo-indol-1-yl)methyl]cyclohexanecarboxylic acid (200 mg, 0.598 mmol), cesium fluoride (45.4 mg, 0.299 mmol) was dissolved in tetrahydrofuran (10 mL), and trimethyl-trifluoromethyl-silane (340 mg, 2.39 mmol) was added at room temperature and stirred for 12 hours. After adding 1 N hydrochloric acid (10 mL), the mixture was stirred at room temperature for 1 hour, and then the mixture was stirred and evaporated. Extract with ethyl acetate (10 mL x 3). The combined organic layers were washed with EtOAc EtOAc. Purification by preparative high performance liquid chromatography to give 3,7-dimethyl-1-[trans-4-(2,2,2-trifluoro-1-hydroxy-1-trifluoromethyl-ethyl -cyclohexylmethyl]-3,7-dihydro-indole-2,6-dione (35.0 mg, yellow solid), yield: 41%.

^{1 H NMR: (400MHz, Methonal} -d 4) δ7.88 (s, 1H), 3.98 (s, 3H), 3.88 (d, J = 6.8Hz, 2H), 3.53 (s, 3H), 2.08-1.79 (m, 6H), 1.30-1.24 (m, 2H), 1.11-1.08 (m, 2H). MS-ESI calcd for [M+H] ⁺ 443.

Example 21

1-[[trans-4-(1-hydroxycyclopropyl)cyclohexyl]methyl]-3,7-dimethylindole-2,6-dione

first step

1-[[trans-4-(1-hydroxycyclopropyl)cyclohexyl]methyl]-3,7-dimethylindole-2,6-dione

Trans-methyl 4-[(3,7-dimethyl-2,6-dioxo-indol-1-yl)methyl]cyclohexanecarboxylic acid (200 mg, 0.598 mmol), tetraisopropyl The base titanium oxide (340 mg, 1.20 mmol) was dissolved in tetrahydrofuran (10 mL), ethyl acetate (3M diethyl ether, 0.39 mL, 1. The reaction was quenched by the addition of saturated aqueous ammonium chloride (50 mL). Extract with ethyl acetate (10 mL x 3). The combined organic layers were washed with EtOAc EtOAc. The residue was purified by preparative high performance liquid chromatography to give 1-[[trans-4-(1-hydroxycyclopropyl)cyclohexyl]methyl]-3,7-dimethylindole-2,6-di Ketone (70.0 mg, yellow solid), yield: 35%.

¹ H NMR: (400 MHz, Methonal-d ₄ ) δ 7.87 (s, 1H), 3.98 (s, 3H), 3.88 (d, J = 6.8 Hz, 2H), 3.53 (s, 3H), 1.79-1.71 (m, 5H), 1.29-1.07 (m, 5H), 0.60-0.57 (m, 2H), 0.42-0.39 (m, 2H). MS-ESI calcd [M+H] ⁺ 333.

Example 22

1-(2-(3-ethyl-3-hydroxycyclohexyl)ethyl)-3,7-dimethyl-1H-indole-2,6(3H,7H)-dione

first step

2-(3-ethyl-3-hydroxycyclohexyl)ethyl methanesulfonate

1-Ethyl-3-(2-hydroxyethyl)cyclohexanol (450 mg, 2.61 mmol) and diisopropylethylamine (500 mg, 3.92 mmol) were dissolved in dichloromethane (10 mL) Methanesulfonyl chloride (600 mg, 5.40 mmol) was slowly added at °C. The reaction solution was stirred at 0 ° C for 0.5 hours. The reaction was quenched with EtOAc (EtOAc)EtOAc The organic phase was combined, dried over anhydrous sodium sulfate, filtered, filtered,jjjjjjjjjjjjjj Hydroxycyclohexyl)ethyl methanesulfonate (450 mg, yellow oil), yield: 69%. MS-ESI calcd for [M ⁺ H] + 251, found 251.

Second step

1-(2-(3-ethyl-3-hydroxycyclohexyl)ethyl)-3,7-dimethyl-1H-indole-2,6(3H,7H)-dione

2-(3-Ethyl-3-hydroxycyclohexyl)ethyl methanesulfonate (200 mg, 0.790 mmol), 3,7-dimethyl-1H-indole-2,6(3H,7H)-di Ketone (144 mg, 0.790 mmol) and potassium carbonate (220 mg, 1.60 mmol), potassium iodide (13.1 mg, 0.0790 mmol) were dissolved in N,N-dimethylformamide (3 mL). The reaction solution was heated to 130 ° C and stirred for 3 hours. The reaction solution was cooled to 25 ° C, and brine (EtOAc) The organic phase was combined, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure and purified by preparative high-performance liquid chromatography to give the product 1-(2-(3-ethyl-3-hydroxycyclohexyl)ethyl)-3 , 7-Dimethyl-1H-indole-2,6(3H,7H)-dione (70.0 mg, white solid), yield: 26%. ¹ H NMR: (400 MHz, Methonal-d ₄ ) δ 7.86 (s, 1H), 4.09-3.94 (m, 5H), 3.52 (s, 3H), 1.88-1.84 (m, 1H), 1.80-1.40 ( m, 10H), 1.26-1.16 (m, 1H), 1.02-0.95 (m, 1H), 0.91 (t, J = 7.2 Hz, 3H). MS-ESI calcd for ^{[M + H-18] +} 317, found 317.

Example 23

first step

3-trifluoromethyl-3-trimethylsiloxy-cyclohexanecarboxylic acid ethyl ester

Ethyl-3-oxocyclohexanecarboxylic acid (1.00 g, 5.88 mmol), cesium fluoride (446 mg, 2.94 mmol) was dissolved in tetrahydrofuran (30 mL) and trimethyl-trifluoromethyl- Silane (1.67 g, 11.7 mmol) was stirred for 12 hours. The reaction was quenched by the addition of water (20 mL). Extract with ethyl acetate (20 mL x 3). The organic layer was combined, washed with brine brine (lilulu g, yellow oil), yield: 76%. MS-ESI calcd for [M+H] ⁺ 313.

Second step

(3-trifluoromethyl-3-trimethylsiloxycyclohexyl)methanol

Ethyl 3-trifluoromethyl-3-trimethylsiloxy-cyclohexanecarboxylate (1.00 g, 3.20 mmol) was dissolved in tetrahydrofuran (10 mL). 6.40 mmol), react for 1 hour. The reaction was quenched by the addition of water (10 mL). Extracted with ethyl acetate (20 mL×3), dried over anhydrous sodium sulfate ), yield: 92%. MS-ESI calcd [M+H] ⁺ 271.

third step

[3-(Trifluoromethyl)-3-trimethylsiloxycyclohexyl]methyl methanesulfonate

3-Trifluoromethyl-3-trimethylsiloxycyclohexyl)methanol (850 mg, 3.14 mmol) and triethylamine (953 mg, 9.42 mmol) were dissolved in dichloromethane (15 mL). Methanesulfonyl chloride (719 mg, 6.28 mmol). The reaction solution was slowly warmed to room temperature and stirred for 2 hours. The reaction was quenched by the addition of aqueous sodium bicarbonate (10 mL). Extract with dichloromethane (20 mL x 3). The organic layer was combined, washed with brine (20 mL) Sulfonate (900 mg, yellow oil), yield: 82%. ¹ H NMR: (400 MHz, Methonal-d ₄ ) δ 4.36-4.32 (m, 1H), 4.17 - 4.13 (m, 1H), 3.08 (s, 3H), 2.12-1.60 (m, 9H), 0.16 ( s, 9H). MS-ESI calcd for [M+H] ⁺ 349.

the fourth step

3,7-Dimethyl-1-[[3-(trifluoromethyl)-3-trimethylsiloxy-cyclohexyl]methyl]indole-2,6-dione

[3-(Trifluoromethyl)-3-trimethylsiloxycyclohexyl]methyl methanesulfonate (200 mg, 0.573 mmol), 3,7-dimethyl-1H-indole-2,6 (3H,7H)-Dione (103 mg, 0.574 mmol), potassium iodide (28.6 mg, 0.172 mmol) and potassium carbonate (374 mg, 1.15 mmol) were dissolved in N,N-dimethylformamide (30 mL). The reaction solution was warmed to 120 ° C and stirred for 3 hours. After cooling to room temperature, filtration, the filtrate was concentrated under reduced pressure and purified by preparative high-purity liquid chromatography to give 3,7-dimethyl-1-[[3-(trifluoromethyl)-3-trimethylsiloxy- Cyclohexyl]methyl]anthracene-2,6-dione (150 mg, yellow solid), yield: 60%. MS-ESI calcd for [M+H] ⁺ 433.

the fifth step

3,7-Dimethyl-1-[[3-(trifluoromethyl)-3-trimethylsiloxy-cyclohexyl]methyl]indole-2,6-dione (200 mg, 0.462 mmol) Dissolved in tetrahydrofuran (10 mL), added 1N hydrochloric acid (10 mL) and stirred for 1 hour at room temperature and then quenched with saturated sodium hydrogen carbonate (50 mL). Extract with ethyl acetate (20 mL x 3). The combined organic layers were washed with brine, dried over anhydrous sodium sulfate Purification by preparative high performance liquid chromatography gave product 1 (10.0 mg, yellow solid) (isomer 1, first peak), yield: 6%. ¹ H NMR: (400 MHz, Methonal-d ₄ ) δ 7.87 (s, 1H), 3.97 (s, 3H), 3.89-3.83 (m, 2H), 3.52 (s, 3H), 2.23-2.22 (m, 1H), 1.76-1.07 (m, 8H). MS-ESI calcd [M+H] ⁺ 361. Product 2 (85.0 mg of yellow solid) (isomer 2, second peak), yield: 51%. ¹ H NMR: (400 MHz, Methonal-d ₄ ) δ 7.87 (s, 1H), 4.31-4.26 (m, 1H), 3.99-3.95 (m, 4H), 3.55 (s, 3H), 2.26-1.88 ( m, 3H), 1.79-1.47 (m, 6H). MS-ESI calcd [M+H] ⁺ 361.

Example 24

1-((5-Hydroxy-5-(trifluoromethyl)tetrahydro-2H-pyran-2-yl)methyl)-3,7-dimethyl-1H-indole-2,6-(3H ,7H)-dione

first step

(3,4-dihydro-2H-pyran-2-yl)methanol

3,4-Dihydro-2H-pyran-2-carbaldehyde (3.00 g, 26.7 mmol) was dissolved in methanol (20 mL), and sodium borohydride (2.02 g, 53.5 mmol) was added at 0 ° C for 2 hours. The reaction was quenched by the addition of saturated aqueous ammonium chloride (30 mL). Extracted with dichloromethane (20 mL x 3), dried over anhydrous sodium sulfate. Yield: 49%. ¹ H NMR: (400 MHz, Methonal-d ₄ ) δ 6.40 (d, J = 6.0 Hz, 1H), 4.71-4.68 (m, 1H), 3.86-3.83 (m, 1H), 3.82-3.61 (m, 2H), 2.13-2.12 (m, 1H), 2.10-2.08 (m, 1H), 2.02-2.01 (m, 1H), 1.68-1.63 (m, 1H).

Second step

(3,4-dihydro-2H-pyran-2-yl)methylmethanesulfonate

(3,4-Dihydro-2H-pyran-2-yl)methanol (1.50 g, 13.1 mmol) and triethylamine (2.66 g, 26.3 mmol) were dissolved in dichloromethane (20 mL) Methanesulfonyl chloride (3.01 g, 26.3 mmol) was added. The reaction solution was slowly warmed to room temperature and stirred for 2 hours. The reaction was quenched by the addition of aqueous sodium bicarbonate (10 mL). Extract with dichloromethane (20 mL x 3). The organic layer was combined, washed with brine, dried over anhydrous sodium sulfate Yellow oil), Yield: 67%. MS-ESI calcd for [M+H] ⁺ 193.

third step

1-((3,4-Dihydro-2H-pyran-2-yl)methyl)-3,7-dimethyl-1H-indole-2,6-(3H,7H)-dione

(3,4-Dihydro-2H-pyran-2-yl)methyl methanesulfonate (1.70 g, 8.84 mmol), 3,7-dimethyl-1H-indole-2,6-(3H , 7H)-dione (1.59 g, 8.84 mmol), potassium iodide (146 mg, 0.884 mmol) and potassium carbonate (2.44 g, 17.7 mmol) were dissolved in N,N-dimethylformamide (50 mL). The reaction solution was warmed to 120 ° C and stirred for 3 hours. The mixture was cooled to room temperature, filtered, and the filtrate was evaporated,jjjjjjjjjj -3,7-Dimethyl-1H-indole-2,6-(3H,7H)-dione (1.30 g, yellow solid), yield: 53%. MS-ESI calcd for [M+H] ⁺ 277.

the fourth step

1-((5-Hydroxytetrahydro-2H-pyran-2-yl)methyl)-3,7-dimethyl-1H-indole-2,6-(3H,7H)-dione

1-(3,4-Dihydro-2H-pyran-2-ylmethyl)-3,7-dimethylindole-2,6-dione (600 mg, 2.17 mmol) was dissolved in tetrahydrofuran (30 mL) Borane dimethyl sulfide (825 mg, 10.7 mmol) was added at 0 °C. The reaction solution was slowly warmed to room temperature and stirred for 12 hours. A 3 N aqueous sodium hydroxide solution (30 mL) and hydrogen peroxide (10 mL) were added and the reaction was continued for one hour. The reaction was quenched with EtOAc (EtOAc) (EtOAc)EtOAc. The combined organic layers were washed with brine, dried over anhydrous sodium sulfate Separation and purification by preparative TLC plate (20:1 dichloromethane/methanol, Rf=0.3) affords 1-((5-hydroxytetrahydro-2H-pyran-2-yl)methyl)-3,7-dimethyl Base-1H-indole-2,6-(3H,7H)-dione (130 mg, yellow oil), yield: 20%. ¹ H NMR: (400 MHz, Methonal-d ₄ ) δ 7.88 (s, 1H), 4.25-4.23 (m, 1H), 4.20 (s, 3H), 3.98-3.67 (m, 5H), 3.54 (s, 3H), 2.10 - 1.77 (m, 2H), 1.49-1.31 (m, 2H). MS-ESI calcd for [M+H] ⁺ 295.

the fifth step

3,7-Dimethyl-1-((5-oxotetrahydro-2H-pyran-2-yl)methyl)-1H-indole-2,6-(3H,7H)-dione

1-((5-Hydroxytetrahydro-2H-pyran-2-yl)methyl)-3,7-dimethyl-1H-indole-2,6-(3H,7H)-dione (130 mg , 0.441 mmol) was dissolved in dichloromethane (10 mL). EtOAc &lt;RTI ID=0.0&gt;&gt; The reaction was quenched with EtOAc EtOAc (EtOAc)EtOAc. Separation and purification by preparative TLC (20:1 dichloromethane/methanol, Rf=0.4) afforded 3,7-dimethyl-1-((5-oxotetrahydro-2H-pyran-2-yl) Methyl)-1H-indole-2,6-(3H,7H)-dione (60.0 mg, yellow solid), yield: 47%. MS-ESI calcd for [M ⁺ H] + 293, found 293.

Step 6

1-((5-Hydroxy-5-(trifluoromethyl)tetrahydro-2H-pyran-2-yl)methyl)-3,7-dimethyl-1H-indole-2,6-(3H ,7H)-dione

3,7-Dimethyl-1-((5-oxotetrahydro-2H-pyran-2-yl)methyl)-1H-indole-2,6-(3H,7H)-dione ( 60.0 mg, 0.205 mmol), cesium fluoride (6.24 mg, 0.0411 mmol) was dissolved in tetrahydrofuran (10 mL), and trimethyltrifluoromethylsilane (87.5 mg, 0.615 mmol) was added at room temperature and stirred for 5 hours. After adding 1 N hydrochloric acid (10 mL), the mixture was stirred at room temperature for 1 hour, and then the mixture was stirred and evaporated. Extract with ethyl acetate (10 mL x 3). The combined organic layers were washed with brine, dried over anhydrous sodium sulfate Purification by preparative high performance liquid chromatography to give 1-((5-hydroxy-5-(trifluoromethyl)tetrahydro-2H-pyran-2-yl)methyl)-3,7-dimethyl- 1H-indole-2,6-(3H,7H)-dione (15.0 mg, yellow solid), yield: 30%. ^{1 H NMR: (400MHz, Methonal} -d 4) δ8.28 (s, 1H), 4.39-4.10 (m, 2H), 4.05 (s, 3H), 3.93-3.89 (m, 2H), 3.55 (s, 3H), 3.32-3.27 (m, 1H), 1.89-1.65 (m, 4H). MS-ESI calcd [M+H] ⁺

Example 25

1-(4-(3-Hydroxypentan-3-yl)benzyl)-3,7-dimethyl-1H-indole-2,6(3H,7H)-dione

first step

Methyl 4-((3,7-dimethyl-2,6-oxo-2,3,6,7-tetrahydro-1H-indol-1-yl)methyl)benzoate

Nitrogen protection, 3,7-dimethyl-1H-indole-2,6(3H,7H)-dione (180 mg, 1.00 mmol), methyl 4-(bromomethyl)benzoate at 25 ° C (251 mg, 1.10 mmol), potassium iodide (55.0 mg, 0.33 mmol) and potassium carbonate (179 mg, 1.30 mmol) were dissolved in anhydrous N,N-dimethylformamide (4 mL) and heated to 110 ° C, stirring 3 hour. After cooling to 25 ° C, it was diluted with water and extracted with ethyl acetate (30 mL EtOAc). The organic phase was combined, dried over anhydrous sodium sulfate, filtered, evaporated, evaporated, m. Methyl 2,6-oxo-2,3,6,7-tetrahydro-1H-indol-1-yl)methyl)benzoate (300 mg, white solid), yield: 91%. MS-ESI calcd for [M+H] ⁺ 329.

Second step

1-(4-(3-Hydroxypentan-3-yl)benzyl)-3,7-dimethyl-1H-indole-2,6(3H,7H)-dione

Methyl 4-((3,7-dimethyl-2,6-oxo-2,3,6,7-tetrahydro-1H-indol-1-yl)methyl)benzoate (200 mg, 0.610 Methyl) was dissolved in dry tetrahydrofuran (3 mL). Under nitrogen atmosphere, ethylmagnesium bromide (3M in diethyl ether, 1.2 mL, 3.60 mmol) was added dropwise at -78 °C. The reaction solution was stirred at this temperature for 0.5 hour, and naturally raised to 25 ° C to continue the reaction for 1 hour. It was quenched with saturated aqueous ammonium chloride (5 mL) andEtOAcEtOAc The organic phase was combined, dried over anhydrous sodium sulfate, filtered, evaporated, evaporated, evaporated. 3-yl)benzyl)-3,7-dimethyl-1H-indole-2,6(3H,7H)-dione (190 mg, white solid). ¹ H NMR: (400 MHz, Methonal-d ₄ ) δ 7.85 (s, 1H), 7.32 (d, J = 8.0 Hz, 2H), 7.28 (d, J = 8.0 Hz, 2H), 5.25 (s, 2H) ), 3.96 (s, 3H), 3.52 (s, 3H), 1.82-1.72 (m, 4H), 0.69 (t, J = 7.2 Hz, 6H). MS-ESI calcd [M+H] ⁺

Example 26

3,7-Dimethyl-1-(3-(1,1,1-trifluoro-2-hydroxypropan-2-yl)benzyl)-1H-indole-2,6-(3H,7H)- Diketone

first step

Ethyl 3-acetylbenzoate

3-Acetylbenzoic acid (500 mg, 3.05 mmol) was dissolved in N,N-dimethylformamide (20 mL), ethyl iodoethane (475 mg, 3.05 mmol) and potassium carbonate (632 mg, 4.57 mmol) After stirring at room temperature for 2 hours, the reaction mixture was evaporated. EtOAcjjjjjjjjjjjjjjj Isolation and purification (5:1 petroleum ether / ethyl acetate, Rf = 0.5) gave ethyl 3-ethyl benzoate (530 mg, white solid). ^{1 H NMR: (400MHz, CDCl} 3) δ8.60 (s, 1H), 8.24 (d, J = 7.6Hz, 1H), 8.15 (d, J = 7.6Hz, 1H), 7.56 (t, J = 7.6 Hz, 1H), 4.41 (q, J = 7.2 Hz, 2H), 2.66 (s, 3H), 1.42 (t, J = 7.2 Hz, 3H). MS-ESI calcd for [M+H] ⁺ 193.

Second step

Ethyl ethyl 3-(1,1,1-trifluoro-2-hydroxypropan-2-yl)benzoate

Ethyl 3-acetylbenzoate (500 mg, 2.60 mmol) was dissolved in tetrahydrofuran (20 mL), and trifluoromethyltrimethylsilane (370 mg, 2.60 mmol) and cesium fluoride (79.0 mg, 0.520) were added at room temperature. Mm). After stirring at room temperature for 12 hours, the reaction mixture was diluted with ethyl acetate (30 mL). Ethyl ethyl 3-(1,1,1-trifluoro-2-hydroxypropan-2-yl)benzoic acid ethyl ester (600 mg, yellow solid). 88%. ^{1 H NMR: (400MHz, CDCl} 3) δ8.26 (s, 1H), 8.05 (d, J = 7.6Hz, 1H), 7.80 (d, J = 7.6Hz, 1H), 7.48 (t, J = 7.6 Hz, 1H), 4.39 (q, J = 7.2 Hz, 2H), 1.82 (s, 3H), 1.40 (t, J = 7.2 Hz, 3H). MS-ESI calcd [M+H] ⁺ 262.

third step

1,1,1-trifluoro-2-(3-hydroxymethyl)phenyl)propan-2-ol

Ethyl ethyl 3-(1,1,1-trifluoro-2-hydroxypropan-2-yl)benzoate (500 mg, 1.91 mmol) was dissolved in tetrahydrofuran (20 mL). Lithium aluminum hydride (108 mg, 2.87 mmol) was stirred at room temperature for 2 hours, and water (0.1 mL), 15% sodium hydroxide (0.1 mL) and water (0.3 mL) were added to the mixture and stirred for 20 min. The reaction mixture was diluted with ethyl acetate (30 mL). EtOAc. Methyl)phenyl)propan-2-ol (400 mg, yellow solid), yield: 95%.

¹ H NMR: (400MHz, CDCl ₃ ) δ 7.62 (s, 1H), 8.05 (d, J = 7.6 Hz, 1H), 7.41-7.37 (m, 2H), 4.73 (s, 2H), 1.80 (s) , 3H). MS-ESI calcd for [M ⁺ H] + 221, found 221.

the fourth step

3-(1,1,1-trifluoro-2-hydroxypropan-2-yl)benzyl methanesulfonate

1,1,1-Trifluoro-2-(3-hydroxymethyl)phenyl)propan-2-ol (400 mg, 1.82 mmol) and triethylamine (275 mg, 2.72 mmol) were dissolved in dichloromethane (20 mL) The reaction mixture was added with methanesulfonyl chloride (250 mg, 2.18 mmol) at 0 ° C, and stirred for 2 hr, diluted with dichloromethane (30 mL). Filtration and concentration of the filtrate under reduced pressure gave 3-(1,1,1-trifluoro-2-hydroxypropan-2-yl)benzyl methanesulfonate (500 mg, m. MS-ESI calcd [M+H] ⁺ 299.

the fifth step

3,7-Dimethyl-1-(3-(1,1,1-trifluoro-2-hydroxypropan-2-yl)benzyl)-1H-indole-2,6-(3H,7H)- Diketone

3-(1,1,1-Trifluoro-2-hydroxypropan-2-yl)benzyl mesylate (100 mg, 0.335 mmol) and 3,7-dimethyl-1H-indole-2,6 -(3H,7H)-dione (60.0 mg, 0.335 mmol) was dissolved in N,N-dimethylformamide (20 mL), and potassium carbonate (70.0 mg, 0.502 mmol) and potassium iodide (6.00 mg) The mixture was stirred at rt. Purification by high performance liquid chromatography gave 3,7-dimethyl-1-(3-(1,1,1-trifluoro-2-hydroxypropan-2-yl)benzyl)-1H-indole-2,6 -(3H,7H)-dione (30.0 mg, white solid), yield: 23%. ^{1 H NMR: (400MHz, CDCl} 3) δ7.90 (s, 1H), 7.70 (s, 1H), 7.49 (d, J = 7.6Hz, 1H), 7.38-7.32 (m, 2H), 5.21 (s , 2H), 4.00 (s, 3H), 3.55 (s, 3H), 1.71 (s, 3H). MS-ESI calcd for [M+H] ⁺ 383.

Example 27

3,7-Dimethyl-1-(4-(1,1,1-trifluoro-2-hydroxypropyl-2-yl)phenyl)-1H-indole-2,6(3H,7H)- Diketone

first step

Methyl 4-(1,1,1-trifluoro-2-hydroxypropyl-2-yl)benzoate

Nitrogen-protected, methyl 4-acetylbenzoate (10.0 g, 56.1 mmol) and trimethyl(trifluoromethyl)silane (16.0 g, 112 mmol) were dissolved in anhydrous tetrahydrofuran (150 mL) at 0 ° C. Tetrabutylammonium fluoride (22.0 g, 84.2 mmol) was added dropwise. The reaction was slowly warmed to room temperature and stirred overnight. The reaction was quenched by the addition of water (50 mL). Extract with ethyl acetate (50 mL x 3). The organic phase was combined, washed with aq. Methyl 2-benzoate (7.00 g, yellow liquid), yield: 50%. ^{1 H NMR: (400MHz, CDCl} 3) δ8.04 (d, J = 8.0Hz, 2H), 7.68 (d, J = 8.0Hz, 2H), 3.92 (s, 3H), 3.27 (s, 1H), 1.80 (s, 3H). MS-ESI calcd [M+H] ⁺ 495.

Second step

1,1,1-trifluoro-2-(4-(hydroxymethyl)phenyl)propyl-2-ol

Lithium aluminum hydride (1.61 g, 42.3 mmol) was slowly added to methyl 4-(1,1,1-trifluoro-2-hydroxypropyl-2-yl)benzoate (7.00 g) under nitrogen at 0 °C. , 28.2 mmol) in tetrahydrofuran (150 mL). The reaction solution was stirred at 0 ° C for 3 hours. At 0 ° C, water (1.60 mL), 15% sodium hydroxide solution (1.60 mL) and water (4.80 mL) were added slowly. Filtration and concentration of the filtrate under reduced pressure gave the product 1,1,1-trifluoro-2-(4-(hydroxymethyl)phenyl)propyl-2-ol (2.40 g, yellow liquid), yield: 93%. ^{1 H NMR: (400MHz, CDCl} 3) δ7.55 (d, J = 8.0Hz, 2H), 7.34 (d, J = 8.0Hz, 2H), 4.66 (s, 2H), 3.37 (s, 1H), 2.39 (s, 1H), 1.75 (s, 3H). MS-ESI calcd for [M ⁺ H] + 221, found 221.

third step

4-(1,1,1-trifluoro-2-hydroxypropyl-2-yl)phenyl methanesulfonate

1,1,1-Trifluoro-2-(4-(hydroxymethyl)phenyl)propyl-2-ol (5.80 g, 26.3 mmol) and diisopropylethylamine (10.2 g, 79.0 mmol) Dissolved in dichloromethane (80 mL) and slowly added methanesulfonyl chloride (4.53 g, 39.5 mmol) at 0 °C. The reaction solution was stirred at 0 ° C for 0.5 hour. The reaction was quenched with aqueous EtOAc (EtOAc)EtOAc The combined organic layers were washed with EtOAc EtOAc EtOAc (HHHHHHHHHHHHHHHHHHHHHHH The product 4-(1,1,1-trifluoro-2-hydroxypropyl-2-yl)phenyl methanesulfonate (3.45 g, m. ^{1 H NMR: (400MHz, CDCl} 3) δ7.66 (d, J = 8.0Hz, 2H), 7.46 (d, J = 8.0Hz, 2H), 5.26 (s, 2H), 2.96 (s, 3H), 2.84 (s, 1H), 1.80 (s, 3H). MS-ESI calcd [M+H] ⁺ 299.

the fourth step

3,7-Dimethyl-1-(4-(1,1,1-trifluoro-2-hydroxypropyl-2-yl)phenyl)-1H-indole-2,6(3H,7H)- Diketone

4-(1,1,1-Trifluoro-2-hydroxypropyl-2-yl)phenyl methanesulfonate (1.95 g, 10.8 mmol), 3,7-dimethyl-1H-indole-2 6(3H,7H)-dione (652 mg, 3.62 mmol) and potassium carbonate (2.99 g, 21.6 mmol) and potassium iodide (180 mg, 1.08 mmol) were dissolved in N,N-dimethylformamide (30 mL). The reaction solution was heated to 130 ° C and stirred for 3 hours. The reaction solution was cooled to room temperature and brine (20 mL) The organic phase was combined, dried over anhydrous sodium sulfate, filtered, evaporated, evaporated, evaporated. 1-(4-(1,1,1-Trifluoro-2-hydroxypropyl-2-yl)phenyl)-1H-indole-2,6(3H,7H)-dione (1.27 g, white solid ), yield: 31%. ^{1 H NMR: (400MHz, CDCl} 3) δ7.57-7.55 (m, 5H), 5.20 (s, 2H), 3.99 (s, 3H), 3.58 (s, 3H), 2.60 (s, 1H), 1.74 (s, 3H). MS-ESI calcd for [M+H] ⁺ 383.

Example 28

first step

6-bromonicotinic acid methyl ester

6-Bromonicotinic acid (1.00 g, 4.95 mmol) was dissolved in N,N-dimethylformamide (30 mL), EtOAc (EtOAc (EtOAc) The reaction solution was stirred at 20 ° C for 12 hours. The reaction mixture was diluted with water (100 mL), EtOAc (EtOAc (EtOAc) /ethyl acetate, Rf = 0.5) gave 6-bromonicotinic acid methyl ester (1.00 g, white solid), yield: 94%. MS-ESI calculated [M+H] ⁺ 216 and 218, found 216 and 218.

Second step

(6-bromopyridin-3-yl)methanol

Methyl 6-bromoinotinic acid (1.00 g, 4.63 mmol) was dissolved in tetrahydrofuran (20 mL), and then evaporated. The reaction was quenched by the addition of water (10 mL). It was extracted with ethyl acetate (20 mL×3), dried over anhydrous sodium sulfate, filtered, evaporated, evaporated, evaporated. -Bromopyridin-3-yl)methanol (600 mg, yellow oil), yield: 69%. MS-ESI calculated [M+H] ⁺ 188 and 190, found 188 and 190.

third step

(6-bromopyridin-3-yl)methyl methanesulfonate

(6-Bromopyridin-3-yl)methanol (1.00 g, 5.32 mmol) and triethylamine (1.18 g, 11.6 mmol) were dissolved in dichloromethane (20 mL), and methanesulfonyl chloride (1.38) was added at 0 °C. g, 12.0 mmol). After the reaction mixture was stirred at room temperature for 2 hr, EtOAc (EtOAc m. Purification (4:1 petroleum ether / ethyl acetate, Rf = 0.5) afforded (6-bromopyridin-3-yl)methyl methanesulfonate (1.20 g, colourless oil). MS-ESI calculated [M+H] ⁺ 266 and 268, found 266 and 268.

the fourth step

1-((6-bromopyridin-3-yl)methyl)-3,7-dimethyl-1H-indole-2,6(3H,7H)-dione

(6-Bromopyridin-3-yl)methyl methanesulfonate (500 mg, 1.88 mmol) was dissolved in N,N-dimethylformamide (20 mL). Dimethyl-1H-indole-2,6(3H,7H)-dione (338 mg, 1.88 mmol), potassium carbonate (389 mg, 2.82 mmol) and potassium iodide (184 mg, 1.11 mmol). The reaction mixture was heated to 100 ° C, and the reaction was stirred for 2 hr. EtOAc (EtOAc) (EtOAc) Chromatography separation and purification (1:1 petroleum ether / ethyl acetate, Rf = 0.3) to give 1-((6-bromopyridin-3-yl)methyl)-3,7-dimethyl-1H-indole- 2,6(3H,7H)-dione (300 mg, yellow solid), yield: 46%. MS-ESI calculated [M+H] ⁺ 350 and 352, found 350 and 352.

the fifth step

1-((6-Acetylpyridin-3-yl)methyl)-3,7-dimethyl-1H-indole-2,6(3H,7H)-dione

Dissolving 1-((6-bromopyridin-3-yl)methyl)-3,7-dimethyl-1H-indole-2,6(3H,7H)-dione (2.00 g, 5.71 mmol) In 1,4-dioxane (50 mL), the reaction solution was added tributyl(1-ethoxyvinyl)stannane (8.25 g, 22.8 mmol) and tetrakistriphenylphosphine palladium (329 mg) at room temperature. , 0.285 mmol). The reaction mixture was heated to 120 ° C for 2 hours. The reaction mixture was cooled to EtOAc EtOAc EtOAc (EtOAc) Separation and purification by silica gel column chromatography (3:1 petroleum ether / ethyl acetate, Rf = 0.3) affords 1-((6-acetylpyridin-3-yl)methyl)-3,7-dimethyl- 1H-indole-2,6(3H,7H)-dione (1.00 g, yellow solid), yield: 56%. ^{1 H NMR: (400MHz, CDCl} 3) δ8.83 (s, 1H), 8.00-7.98 (m, 1H), 7.95-7.93 (m, 1H), 7.54 (s, 1H), 5.27 (s, 2H) , 4.01 (s, 3H), 3.59 (s, 3H), 2.71 (s, 3H). MS-ESI calcd for [M ⁺ H] + 314, found 314.

Step 6

3,7-Dimethyl-1-((6-(1,1,1-trifluoro-2-hydroxypropan-2-yl)pyridin-3-yl)methyl)-1H-indole-2,6 (3H,7H)-dione

Dissolving 1-((6-acetylpyridin-3-yl)methyl)-3,7-dimethyl-1H-indole-2,6(3H,7H)-dione (150 mg, 0.478 mmol) Trifluoromethyl trimethylsilane (102 mg, 0.718 mmol) and cesium fluoride (73.0 mg, 0.478 mmol) were added to tetrahydrofuran (30 mL) at room temperature. The reaction mixture was stirred at room temperature for 12 hr. EtOAc (EtOAc) (EtOAc) Drying with anhydrous sodium sulfate, filtering, and concentrating the filtrate under reduced pressure by high-performance liquid chromatography to obtain 3,7-dimethyl-1-((6-(1,1,1-trifluoro-2-hydroxypropane-2) -yl)pyridin-3-yl)methyl)-1H-indole-2,6(3H,7H)-dione (50 mg, white solid), yield: 27%. ^{1 H NMR: (400MHz, CDCl} 3) δ8.76 (s, 1H), 7.99 (d, J = 8.0Hz, 1H), 7.53 (s, 1H), 7.44 (d, J = 8.0Hz, 1H), 5.23 (s, 2H), 3.99 (s, 3H), 3.58 (s, 3H), 1.68 (s, 3H). MS-ESI calcd for [M+H] ⁺ 384.

Example 29

3,7-Dimethyl-1-[[5-(2,2,2-trifluoro-1-hydroxy-1-methyl-ethyl)-2-pyridyl]methyl]indole-2,6 -dione

first step

1-[6-(bromomethyl)-3-pyridyl]ethanone

1-(6-Methyl-3-pyridyl)ethanone (500 mg, 3.70 mmol), N-bromosuccinimide (658 mg, 3.70 mmol), azobisisobutyronitrile (182 mg, 1.11 mmol) Dissolved in carbon tetrachloride (20 mL) and reacted at 90 ° C for 12 hours. The reaction was quenched by the addition of saturated sodium thiosulfate solution (30 mL). The mixture was extracted with EtOAc (EtOAc m. Yield: 16%. MS-ESI calcd for [M ⁺ H] + 214,216, 214, 216 found.

Second step

1-[(5-acetyl-2-pyridyl)methyl]-3,7-dimethylindole-2,6-dione

1-[6-(Bromomethyl)-3-pyridyl]ethanone (100 mg, 0.467 mmol), 3,7-dimethylindole-2,6-dione (84.2 mg, 0.467 mmol), potassium iodide (7.70 mg, 0.0467 mmol) and potassium carbonate (194 mg, 1.40 mmol) were dissolved in N,N-dimethylformamide (10 mL). The reaction solution was warmed to 120 ° C and stirred for 3 hours. It was cooled to room temperature, filtered, and the filtrate was concentrated under reduced pressure and purified by preparative TLC (ethyl acetate, Rf value = 0.3) to give 1-[(5-acetyl-2-pyridyl)methyl]-3,7- Methyl hydrazine-2,6-dione (50.0 mg, yellow solid), yield: 34%. MS-ESI calcd for [M ⁺ H] + 314, found 314.

third step

3,7-Dimethyl-1-[[5-(2,2,2-trifluoro-1-hydroxy-1-methyl-ethyl)-2-pyridyl]methyl]indole-2,6 -dione

1-[(5-Acetyl-2-pyridyl)methyl]-3,7-dimethylindole-2,6-dione (50.0 mg, 0.159 mmol), cesium fluoride (24.2 mg, 0.159) Methyl acetate was dissolved in tetrahydrofuran (10 mL), trimethyl-trifluoromethyl-silane (113 mg, 0.798 mmol) was added at room temperature and stirred for 12 hours. The reaction was quenched by the addition of water (20 mL). Extract with ethyl acetate (10 mL x 3). The combined organic layers were washed with brine, dried over anhydrous sodium sulfate Purification by preparative high performance liquid chromatography to give 3,7-dimethyl-1-[[5-(2,2,2-trifluoro-1-hydroxy-1-methyl-ethyl)-2-pyridine Methyl] fluorene-2,6-dione (10.0 mg, yellow solid), yield: 16%. ^{1 H NMR: (400MHz, Methonal} -d 4) δ8.96 (s, 1H), 8.75 (d, J = 8.0Hz, 1H), 8.06 (d, J = 8.0Hz, 1H), 7.97 (s, 1H ), 5.55 (s, 2H), 4.00 (s, 3H), 3.57 (s, 3H), 1.86 (s, 3H). MS-ESI calcd for [M+H] ⁺ 384.

Example 30

3,7-Dimethyl-1-((5(1,1,1-trifluoro-2-hydroxypropan-2-yl)pyrazin-2-yl)methyl)-indole-2,6(3H ,7H)-dione

first step

N-methoxy-N,5-dimethylpyrazine-2-carboxamide

5-Methylpyrazine-2-carboxylic acid (2.00 g, 14.5 mmol), 1-hydroxybenzotriazole (391 mg, 2.90 mmol), 1-(3-dimethylaminopropyl)-3-ethyl The carbodiimide hydrochloride (1.69 g, 17.4 mmol) was dissolved in anhydrous dichloromethane (10 mL) and trichloromethane (30 mL). EtOAc (3. (mmol), the reaction solution was stirred at 25 ° C for 12 hours. The reaction was quenched by the addition of water (50 mL). The reaction mixture was extracted with EtOAc (EtOAc)EtOAc. Rf = 0.1) gave N-methoxy-N,5-dimethylpyrazine-2-carboxamide (2.00 g,yield of yellow oil). ¹ H NMR: (400 MHz, CDCl ₃ ) δ 8.80 (s, 1H), 8.45 (s, 1H), 3.73 (s, 3H), 3.40 (s, 3H), 2.61 (s, 3H).

Second step

1-(5-methylpyrazin-2-yl)ethanone

N-Methoxy-N,5-dimethylpyrazine-2-carboxamide (1.50 g, 8.28 mmol) was dissolved in tetrahydrofuran (30 mL), and methylmagnesium bromide (3M diethyl ether solution) was added dropwise at 0 °C. 13.3 mL, 39.9 mmol), then stirred at 25 ° C for 1 hour. The mixture was cooled to 0 ° C and quenched by water (10 mL). The mixture was extracted with EtOAc (30 mLEtOAc) The residue was purified by EtOAc EtOAcjjjjjjjjj 62%. MS-ESI calcd for [M ⁺ H] + 137, found 137.

third step

1-(5(bromomethyl)pyrazin-2-yl)ethanone

1-(5-Methylpyrazin-2-yl)ethyl (700 mg, 5.14 mmol) was dissolved in carbon tetrachloride (20 mL), then azobisisobutyronitrile (169 mg, 1.03 mmol) and N- Bromosuccinimide (1.14 g, 6.43 mmol). The reaction solution was reacted under nitrogen at 100 ° C for 5 hours. The reaction mixture was directly filtrated and concentrated under reduced pressure. EtOAcjjjjjjjjjj (300 mg, yellow oil), Yield: 27%. MS-ESI calculated [M+H] ⁺ 215 and 217, found 215 and 217.

the fourth step

1-((5-Acetylpyrazin-2-yl)methyl)-3,7-dimethyl-1H-indole-2,6(3H,7H)-dione

1-(5(Bromomethyl)pyrazin-2-yl)ethyl (300 mg, 1.40 mmol), 3,7-dimethyl-1H-indole-2,6(3H,7H)-dione (251 mg) , 1.40 mmol), potassium iodide (23.2 mg, 0.140 mmol) and potassium carbonate (578 mg, 4.19 mmol) were dissolved in anhydrous N,N-dimethylformamide (20 mL). The reaction solution was heated to 120 ° C and allowed to react for 3 hours. The reaction solution was cooled to 20 ° C, filtered, and the filtrate was evaporated, evaporated,jjjjjjjjjjjjjjjjjj -3,7-Dimethyl-1H-indole-2,6(3H,7H)-dione (300 mg, yellow solid), yield: 68%. MS ESI calcd for [M+H] ⁺ 315.

the fifth step

3,7-Dimethyl-1-((5(1,1,1-trifluoro-2-hydroxypropan-2-yl)pyrazin-2-yl)methyl)-indole-2,6(3H ,7H)-dione

1-((5-Acetylpyrazin-2-yl)methyl)-3,7-dimethyl-1H-indole-2,6(3H,7H)-dione (300 mg, 0.954 mmol), The cesium fluoride (14.5 mg, 0.0954 mmol) was dissolved in anhydrous tetrahydrofuran (10 mL) then trimethylsilyltrifluoromethyl (407 mg, 2.86 mmol). The reaction solution was reacted under nitrogen at 25 ° C for 2 hours. Then hydrochloric acid (4N, 4 mL) was added. The mixture was reacted for 1 hour under a nitrogen atmosphere at room temperature. The reaction was quenched with EtOAc EtOAc (EtOAc (EtOAc) 1 petroleum ether / ethyl acetate, Rf = 0.3) to give 3,7-dimethyl-1-((5(1,1,1-trifluoro-2-hydroxypropan-2-yl)pyrazine-2- Methyl)-indole-2,6(3H,7H)-dione (100 mg, white solid), yield: 40%. ¹ H NMR: (400 MHz, Methonal-d ₄ ) δ 8.85 (s, 1H), 8.65 (s, 1H), 7.92 (s, 1H), 5.40 (s, 2H), 3.99 (s, 3H), 3.56 (s, 3H), 1.78 (s, 3H). MS ESI calcd for [M+H] ⁺ 385.

Example 31

1-((3-(1,1,1,3,3,3-hexafluoro-2-hydroxypropan-2-yl)isoxazole-5-yl)methyl)-3,7-dimethyl -1H-嘌呤-2,6-(3H,7H)-dione

first step

Methyl 5-(bromomethyl)isoxazole-3-carboxylic acid ethyl ester

Ethyl methyl 5-methylisoxazole-3-carboxylate (5.00 g, 35.4 mmol), N-bromosuccinimide (6.31 g, 35.4 mmol), benzoyl peroxide (858 mg, 3.54 mmol) was dissolved in carbon tetrachloride (20 mL) and reacted at 80 ° C for 12 hours. The reaction was quenched by the addition of a saturated sodium thiosulfate solution (30 mL). Extracted with dichloromethane (20 mL x 3), dried over anhydrous sodium sulfate, filtered, evaporated, evaporated, evaporated, evaporated Ethyl 5-(bromomethyl)isoxazol-3-carboxylate (2.00 g, yellow oil), yield: 26%. ¹ H NMR: (400 MHz, Methonal-d ₄ ) δ 6.88 (s, 1H), 4.73 (s, 2H), 3.97 (s, 3H). MS-ESI calcd for [M+H] ⁺ 220, 222.

Second step

Methyl 5-((3,7-dimethyl-2,6-dioxo-2,3,6,7-tetrahydro-1H-indol-1-yl)methyl)isoxazole-3- Ethyl carboxylate

Ethyl 5-(bromomethyl)isoxazol-3-carboxylate (2.00 g, 9.09 mmol), 3,7-dimethyl-1H-indole-2,6-(3H,7H)-dione (1.64 g, 9.09 mmol), potassium iodide (151 mg, 0.909 mmol) and potassium carbonate (2.51 g, 18.2 mmol) were dissolved in N,N-dimethylformamide (50 mL). The reaction solution was warmed to 120 ° C and stirred for 3 hours. The mixture was cooled to room temperature, filtered, and the filtrate was concentrated and evaporated to silica gel column chromatography (ethyl acetate, Rf value = 0.4) to give methyl 5-((3,7-dimethyl-2,6-dioxo-) Ethyl 2,3,6,7-tetrahydro-1H-indol-1-yl)methyl)isoxazole-3-carboxylate (1.70 g, yellow solid), yield: 59%. ¹ H NMR: (400 MHz, Methonal-d ₄ ) δ 8.06 (s, 1H), 6.82 (s, 1H), 5.22 (s, 2H), 3.87 (s, 3H), 3.83 (s, 3H), 3.45 (s, 3H). MS-ESI calc. [M+H] ⁺ 320, Found 320.

third step

1-((3-(1,1,1,3,3,3-hexafluoro-2-hydroxypropan-2-yl)isoxazole-5-yl)methyl)-3,7-dimethyl -1H-嘌呤-2,6-(3H,7H)-dione

Methyl 5-((3,7-dimethyl-2,6-dioxo-2,3,6,7-tetrahydro-1H-indol-1-yl)methyl)isoxazole-3 Ethyl carboxylate (200 mg, 0.626 mmol), cesium fluoride (95.0 mg, 0.626 mmol) dissolved in tetrahydrofuran (10 mL). Trimethyltrifluoromethylsilane (445 mg, 3.13 mmol) was added at room temperature and stirred 12 hour. After adding 1 N hydrochloric acid (10 mL), the mixture was stirred at room temperature for 1 hour, and then the mixture was stirred and evaporated. Extract with ethyl acetate (10 mL x 3). The combined organic layers were washed with EtOAc EtOAc. Purification by preparative high performance liquid chromatography to give 1-((3-(1,1,1,3,3,3-hexafluoro-2-hydroxypropan-2-yl)isoxazole-5-yl) -3,7-Dimethyl-1H-indole-2,6-(3H,7H)-dione (10.0 mg, yellow solid), yield: 4%. ¹ H NMR: (400 MHz, Methonal-d ₄ ) δ 7.95 (s, 1H), 6.52 (s, 1H), 5.37 (s, 2H), 4.00 (s, 3H), 3.57 (s, 3H). MS-ESI calcd for [M ⁺ H] + 428, found 428.

Example 32

3,7-Dimethyl-1-((3-(1,1,1-trifluoro-2-hydroxypropan-2-yl)isoxazol-5-yl)methyl)-1H-indole-2 ,6-(3H,7H)-dione

first step

1-(5-methylisoxazol-3-yl)ethanone

Ethyl methyl 5-methylisoxazole-3-carboxylate (5.00 g, 35.4 mmol) and triethylamine (21.5 g, 213 mmol) were dissolved in tetrahydrofuran (80 mL), and methyl bromide was added at 0 °C. Magnesium (3M in diethyl ether, 35 mL, 105 mmol) was reacted for three hours. The reaction was quenched by the addition of saturated aqueous ammonium chloride (30 mL). It was extracted with ethyl acetate (30 mL×3), dried over anhydrous sodium sulfate, filtered, evaporated, evaporated, evaporated. (5-Methylisoxazol-3-yl)ethanone (1.00 g, yellow oil), yield: 23%. ¹ H NMR: (400 MHz, Methonal-d ₄ ) δ 6.39 (s, 1H), 2.58 (s, 3H), 2.49 (s, 3H). MS-ESI calcd for [M+H] ⁺ 126.

Second step

1-(5-(bromomethyl)isoxazol-3-yl)ethanone

1-(5-Methylisoxazol-3-yl)ethanone (100 mg, 0.799 mmol), N-bromosuccinimide (142 mg, 0.799 mmol), benzoyl peroxide (19.3 mg, 0.0800 mmol) was dissolved in carbon tetrachloride (10 mL) and reacted at 90 ° C for 12 hours. The reaction was quenched by the addition of saturated sodium thiosulfate solution (30 mL). Extracted with dichloromethane (10 mL×3), dried over anhydrous sodium sulfate , Yield: 93%. MS-ESI calculated [M+H] ⁺ 204 and 206, found 204 and 206.

third step

1-((3-acetylisoxazole-5-yl)methyl)-3,7-dimethyl-1H-indole-2,6-(3H,7H)-dione

1-(5-(Bromomethyl)isoxazol-3-yl)ethanone (150 mg, 0.735 mmol), 3,7-dimethyl-1H-indole-2,6-(3H,7H)- Diketone (132 mg, 0.735 mmol), potassium iodide (61.0 mg, 0.367 mmol) and potassium carbonate (305 mg, 2.21 mmol) were dissolved in N,N-dimethylformamide (10 mL). The reaction solution was warmed to 120 ° C and stirred for 3 hours. It was cooled to room temperature, filtered, and the filtrate was concentrated under reduced pressure and purified by preparative TLC (ethyl acetate, Rf value = 0.3) to give 1-((3-acetylisoxazole-5-yl)methyl)-3 , 7-Dimethyl-1H-indole-2,6-(3H,7H)-dione (50.0 mg, yellow solid), yield: 22%. MS-ESI calcd for [M+H] ⁺ 303.

the fourth step

3,7-Dimethyl-1-((3-(1,1,1-trifluoro-2-hydroxypropan-2-yl)isoxazol-5-yl)methyl)-1H-indole-2 ,6-(3H,7H)-dione

1-[(3-Acetyloxazol-5-yl)methyl]-3,7-dimethylindole-2,6-dione (50.0 mg, 0.164 mmol), cesium fluoride (25.0 mg) , 0.164 mmol) was dissolved in tetrahydrofuran (10 mL), and trimethyltrifluoromethylsilane (70.3 mg, 0.494 mmol) was added at room temperature and stirred for 12 hours. After adding 1 N hydrochloric acid (10 mL), the mixture was stirred at room temperature for 1 hour, and then the mixture was stirred and evaporated. Extract with ethyl acetate (10 mL x 3). The combined organic layers were washed with brine, dried over anhydrous sodium sulfate Purification by preparative high performance liquid chromatography to give 3,7-dimethyl-1-((3-(1,1,1-trifluoro-2-hydroxypropan-2-yl)isoxazole-5-yl Methyl)-1H-indole-2,6-(3H,7H)-dione (22.0 mg, yellow solid), yield: 36%. ^{1 H NMR: (400MHz, Methonal} -d 4) δ7.98 (s, 1H), 6.48 (s, 1H), 5.33 (s, 2H), 4.01 (s, 3H), 3.57 (s, 3H), 1.71 (s, 3H). MS-ESI calcd [M+H] ⁺ 372.

Example 33

3,7-Dimethyl-1-((2-(1,1,1-trifluoro-2-hydroxypropan-2-yl)thiazol-4-yl)methyl)-1H-indole-2,6 -(3H,7H)-dione

first step

1-(4-(bromomethyl)thiazol-2-yl)ethanone

1-(4-Methylthiazol-2-yl)ethanone (200 mg, 1.42 mmol), N-bromosuccinimide (252 mg, 1.42 mmol), azobisisobutyronitrile (46.6 mg, 0.284) Methyl) was dissolved in carbon tetrachloride (20 mL) and reacted at 80 ° C for 12 hours. The reaction was quenched by the addition of saturated sodium thiosulfate solution (30 mL). Extracted with dichloromethane (10 mL x 3), dried over anhydrous sodium sulfate. Yield: 64%. ¹ H NMR: (400 MHz, Methonal-d ₄ ) δ 7.97 (s, 1H), 4.73 (s, 2H), 2.66 (s, 3H). MS-ESI calcd for [M+H] ⁺ 220, 222.

Second step

1-((2-Acetylthiazol-4-yl)methyl)-3,7-dimethyl-1H-indole-2,6-(3H,7H)-dione

1-(4-(Bromomethyl)thiazol-2-yl)ethanone (100 mg, 0.454 mmol), 3,7-dimethyl-1H-indole-2,6-(3H,7H)-dione (81.9 mg, 0.454 mmol), potassium iodide (7.50 mg, 0.0454 mmol) and potassium carbonate (125 mg, 0.908 mmol) were dissolved in N,N-dimethylformamide (10 mL). The reaction solution was warmed to 120 ° C and stirred for 3 hours. It was cooled to room temperature, filtered, and the filtrate was concentrated under reduced pressure and purified by preparative TLC (ethyl acetate, Rf value = 0.3) to give 1-((2-acetylthiazol-4-yl)methyl)-3, 7- Methyl-1H-indole-2,6-(3H,7H)-dione (80.0 mg, yellow solid), yield: 55%. ^{1 H NMR: (400MHz, Methonal} -d 4) δ7.92 (s, 1H), 7.73 (s, 1H), 5.38 (s, 2H), 4.00 (s, 3H), 3.57 (s, 3H), 2.64 (s, 3H). MS-ESI calc. [M+H] ⁺ 320, Found 320.

third step

3,7-Dimethyl-1-((2-(1,1,1-trifluoro-2-hydroxypropan-2-yl)thiazol-4-yl)methyl)-1H-indole-2,6 -(3H,7H)-dione

1-((2-Acetylthiazol-4-yl)methyl)-3,7-dimethyl-1H-indole-2,6-(3H,7H)-dione (200 mg, 0.626 mmol), The ruthenium fluoride (95.0 mg, 0.626 mmol) was dissolved in tetrahydrofuran (10 mL), and trimethyl-trifluoromethyl-silane (267 mg, 1.88 mmol) was added at room temperature and stirred for 12 hours. The reaction was quenched by the addition of water (20 mL). Extract with ethyl acetate (10 mL x 3). The combined organic layers were washed with brine, dried over anhydrous sodium sulfate Purification by preparative high performance liquid chromatography to give 3,7-dimethyl-1-((2-(1,1,1-trifluoro-2-hydroxypropan-2-yl)thiazol-4-yl)- -1H-indole-2,6-(3H,7H)-dione (100 mg, yellow solid), yield: 41%. ¹ H NMR: (400 MHz, Methonal-d ₄ ) δ 8.10 (s, 1H), 7.33 (s, 1H), 5.32 (s, 2H), 4.03 (s, 3H), 3.57 (s, 3H), 1.80 (s, 3H). MS-ESI calcd for [M+H] ⁺ 390.

Example 34

3,7-Dimethyl-1-((5-methyl-2-(1,1,1-trifluoro-2-hydroxypropan-2-yl)thiazol-4-yl)methyl)-1H-嘌呤-2,6-(3H,7H)-dione

first step

1-(4-(bromomethyl)-5-methylthiazol-2-yl)ethanone

1-(4,5-Dimethylpyridin-2-yl)ethanone (200 mg, 1.29 mmol), N-bromosuccinimide (229 mg, 1.29 mmol), azobisisobutyronitrile (21.1 Mg, 0.129 mmol) was dissolved in carbon tetrachloride (10 mL) and reacted at 80 ° C for 12 hours. The reaction was quenched by the addition of saturated sodium thiosulfate solution (30 mL). It was extracted with dichloromethane (10 mL x 3), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to give 1-(4-(bromomethyl)-5-methylthiazol-2-yl)ethanone (200 mg) , yellow oily), yield: 66%. ¹ H NMR: (400 MHz, Methonal-d ₄ ) δ 4.88 (s, 2H), 2.65 (s, 3H), 2.47 (s, 3H). MS-ESI calcd for [M+H] ⁺ 234, 236.

Second step

1-((2-Acetyl-5-methylthiazol-4-yl)methyl)-3,7-dimethyl-1H-indole-2,6-(3H,7H)-dione

1-(4-(Bromomethyl)-5-methylthiazol-2-yl)ethanone (200 mg, 0.854 mmol), 3,7-dimethyl-1H-indole-2,6-(3H, 7H)-Dione (154 mg, 0.854 mmol), potassium iodide (14.0 mg, 0.0854 mmol) and potassium carbonate (354 mg, 2.56 mmol) were dissolved in N,N-dimethylformamide (10 mL). The reaction solution was warmed to 120 ° C and stirred for 3 hours. It was cooled to room temperature, filtered, and the filtrate was concentrated under reduced pressure and purified (yield ethyl acetate, Rf value = 0.3) to give 1-((2-acetyl-5-methylthiazol-4-yl)methyl)- 3,7-Dimethyl-1H-indole-2,6-(3H,7H)-dione (200 mg, yellow solid), yield: 70%. ^{1 H NMR: (400MHz, Methonal} -d 4) δ7.90 (s, 1H), 5.35 (s, 2H), 4.00 (s, 3H), 3.55 (s, 3H), 2.66 (s, 3H), 2.61 (s, 3H). MS-ESI calcd for [M ⁺ H] + 334, found 334.

third step

3,7-Dimethyl-1-((5-methyl-2-(1,1,1-trifluoro-2-hydroxypropan-2-yl)thiazol-4-yl)methyl)-1H-嘌呤-2,6-(3H,7H)-dione

1-((2-Acetyl-5-methylthiazol-4-yl)methyl)-3,7-dimethyl-1H-indole-2,6-(3H,7H)-dione (80.0 Mg, 0.240 mmol), cesium fluoride (18.2 mg, 0.120 mmol) was dissolved in tetrahydrofuran (10 mL), and trimethyl-trifluoromethyl-silane (102 mg, 0.720 mmol) was added at room temperature and stirred for 12 hours. The reaction was quenched by the addition of water (20 mL). Extract with ethyl acetate (10 mL x 3). The combined organic layers were washed with brine, dried over anhydrous sodium sulfate Purification by preparative high performance liquid chromatography to give 3,7-dimethyl-1-((5-methyl-2-(1,1,1-trifluoro-2-hydroxypropan-2-yl)thiazole- 4-yl)methyl)-1H-indole-2,6-(3H,7H)-dione (35.0 mg, yellow solid), yield: 36%. ¹ H NMR: (400 MHz, Methonal-d ₄ ) δ 8.27 (s, 1H), 5.36 (s, 2H), 4.06 (s, 3H), 3.57 (s, 3H), 2.73 (s, 3H), 1.90 (s, 3H). MS-ESI calculated [M+H] ⁺ 404. Found 404.

Example 35

3,7-Dimethyl-1-((2-(1,1,1-trifluoro-2-hydroxypropan-2-yl)thiazol-5-yl)methyl)-1H-indole-2,6 -(3H,7H)-dione

first step

1-(5-methylthiazol-2-yl)ethylcyclohexanone

5-Methylthiazole (2.00 g, 20.2 mmol) was dissolved in tetrahydrofuran (50 mL), and n-butyllithium (2.5 M tetrahydrofuran solution, 9.68 mL, 24.2 mmol) was slowly added dropwise under nitrogen. The reaction mixture was stirred at -78 ° C for 0.5 hr, and N-methoxy-N-methylacetamide (2.50 g, 24.2 mmol) dissolved in THF (1 mL). The reaction solution was heated to 0 ° C and stirred for 1.5 hours. Water (10 mL) was slowly added to the reaction mixture at 0 ° C, and extracted with ethyl acetate (30 mL x 3). The organic phase was combined, dried over anhydrous sodium sulfate, filtered and evaporated and evaporated tolulujjjjjjjjjjjjjjjjjjjjj 2-yl)ethylcyclohexanone (1.45 g, yellow solid), yield: 51%. ¹ H NMR: (400 MHz, Methonal-d ₄ ) δ 7.73 (s, 1H), 2.61 (s, 3H), 2.57 (s, 3H). MS-ESI calcd for [M ⁺ H] + 142, found 142.

Second step

1-(5-(bromomethyl)thiazol-2-yl)ethylcyclohexanone

1-(5-Methylthiazol-2-yl)ethylcyclohexanone (200 mg, 1.42 mmol) and azoisobutylidene (2.33 mg, 0.0142 mmol) were dissolved in chloroform (5 mL). Desuccinimide (252 mg, 1.42 mmol). The reaction solution was heated to 78 ° C and stirred for 16 hours. The reaction solution was cooled to room temperature, water (30 mL) was slowly added and extracted with chloroform (30 mL x 3). The organic phase was combined, dried over anhydrous sodium MS-ESI calculated [M+H] ⁺ 220 and 222, found 220 and 222.

third step

1-((2-acetylthiazole-5-yl)methyl)-3,7-dimethyl-1H-indole-2,6-(3H,7H)-dione

1-(5-(Bromomethyl)thiazol-2-yl)ethylcyclohexanone (290 mg, 1.05 mmol), 3,7-dimethyl-1H-indole-2,6-(3H,7H) The diketone (284 mg, 1.58 mmol) and potassium iodide (17.5 mg, 0.105 mmol) were dissolved in N,N-dimethylformamide (5 mL), and potassium carbonate (437 mg, 3.16 mmol) was added and reacted at 130 ° C for 2.5 hours. The reaction solution was cooled to room temperature, filtered, and the filtrate was evaporated, evaporated, mjjjjjjjjjjjjjjjjjjjjjjjjjjjjjjjj 3,7-Dimethyl-1H-indole-2,6-(3H,7H)-dione (292 mg, yellow solid), yield: 87%. MS-ESI calc. [M+H] ⁺ 320, Found 320.

the fourth step

3,7-Dimethyl-1-((2-(1,1,1-trifluoro-2-hydroxypropan-2-yl)thiazol-5-yl)methyl)-1H-indole-2,6 -(3H,7H)-dione

1-((2-Acetylthiazol-5-yl)methyl)-3,7-dimethyl-1H-indole-2,6-(3H,7H)-dione (280 mg, 0.438 mmol) Cesium fluoride (6.66 mg, 0.0438 mmol) was dissolved in tetrahydrofuran (6 mL) and trifluoromethyltrimethylsilane (75.0 mg, 0.500 mmol) was slowly added under nitrogen. The reaction was stirred at 25 ° C for 1.5 hours. After adding 4N aqueous hydrochloric acid (0.2 mL) and stirring at room temperature for half an hour, the pH was adjusted to 7 with saturated sodium hydrogen carbonate solution (10 mL), water (20 mL) and ethyl acetate (50 mL x 3) Drying over sodium sulfate and concentrating under reduced pressure afforded crude product purified by preparative high-purity chromatography to give the product 3,7-dimethyl-1-((2-(1,1,1-trifluoro-2-hydroxypropane-2) -yl)thiazol-5-yl)methyl)-1H-indole-2,6-(3H,7H)-dione (32.0 mg, white solid), yield: 19%. ^{1 H NMR: (400MHz, Methonal} -d 4) δ7.89 (s, 1H), 7.82 (s, 1H), 5.35 (s, 2H), 4.00 (s, 3H), 3.56 (s, 3H), 1.76 (s, 3H). MS-ESI calcd for [M+H] ⁺ 390.

Example 36

3,7-Dimethyl-1-(2-(4-methyl-2-(1,1,1-trifluoro-2-hydroxypropan-2-yl)thiazol-5-yl)ethyl)- 1H-嘌呤-2,6-(3H,7H)-dione

first step

1-(5-(2-hydroxyethyl)-4-methylthiazol-2-yl)ethanone

2-(4-Methylthiazol-5-yl)ethanol (500 mg, 3.49 mmol) was dissolved in tetrahydrofuran (100 mL), and n-butyl lithium (3M-hexanes, 2.33 mL, 6. After half an hour of reaction, N-methoxy-N-methyl-acetamide (432 mg, 4.19 mmol) was added and stirring was continued for 3 hours. The reaction was quenched by the addition of saturated aqueous ammonium chloride (50 mL). Ethyl acetate extraction (10 mL x 3), dried over anhydrous sodium sulfate, filtered, filtered, evaporated, evaporated, evaporated. (5-(2-Hydroxyethyl)-4-methylthiazol-2-yl)ethanone (200 mg, yellow oil), yield: 31%. ¹ H NMR: (400 MHz, CDCl ₃ ) δ 3.78 (t, J = 6.4 Hz, 2H), 3.18 (t, J = 6.4 Hz, 2H), 2.67 (s, 3H), 2.47 (s, 3H). MS-ESI calcd for [M+H] ⁺ 186.

Second step

2-(2-acetyl-4-methyl-thiazol-5-yl)ethyl methanesulfonate

1-(5-(2-Hydroxyethyl)-4-methylthiazol-2-yl)ethanone (120 mg, 0.647 mmol) and triethylamine (196 mg, 1.94 mmol) were dissolved in dichloromethane (10 mL) Methanesulfonyl chloride (148 mg, 1.30 mmol) was added at 0 °C. The reaction solution was slowly warmed to room temperature and stirred for 2 hours. The reaction was quenched by the addition of aqueous sodium bicarbonate (50 mL). Extract with dichloromethane (10 mL x 3). The organic phase was combined, washed with brine, dried over anhydrous sodium sulfate, filtered, evaporated, evaporated, evaporated. -Acetyl-4-methyl-thiazol-5-yl)ethyl methanesulfonate (150 mg, yellow oil), yield: 88%. ^{1 H NMR: (400MHz, CDCl3} ) δ4.41 (t, J = 6.4Hz, 2H), 3.27 (t, J = 6.4Hz, 2H), 3.01 (s, 3H), 2.67 (s, 3H), 2.46 (s, 3H). MS-ESI calcd [M+H] ⁺ 264.

third step

1-(2-(2-acetyl-4-methylthiazol-5-yl)ethyl)-3,7-dimethyl-1H-indole-2,6-(3H,7H)-dione

2-(2-Acetyl-4-methyl-thiazol-5-yl)ethyl methanesulfonate (150 mg, 0.569 mmol), 3,7-dimethyl-1H-indole-2,6-( 3H,7H)-Dione (102 mg, 0.569 mmol), potassium iodide (18.9 mg, 0.114 mmol) and potassium carbonate (236 mg, 1.71 mmol) were dissolved in N,N-dimethylformamide (10 mL). The reaction solution was warmed to 120 ° C and stirred for 3 hours. It was cooled to room temperature, filtered, and the filtrate was concentrated. Separation and purification by preparative TLC (ethyl acetate, Rf value = 0.5) gave 1-(2-(2-acetyl-4-methylthiazol-5-yl)ethyl)-3,7-dimethyl -1H-indole-2,6-(3H,7H)-dione (30.0 mg, yellow solid), yield: 15%. MS-ESI calcd for [M ⁺ H] + 348, found 348.

the fourth step

3,7-Dimethyl-1-(2-(4-methyl-2-(1,1,1-trifluoro-2-hydroxypropan-2-yl)thiazol-5-yl)ethyl)- 1H-嘌呤-2,6-(3H,7H)-dione

1-(2-(2-Acetyl-4-methylthiazol-5-yl)ethyl)-3,7-dimethyl-1H-indole-2,6-(3H,7H)-dione (40.0 mg, 0.115 mmol), cesium fluoride (17.5 mg, 0.115 mmol) was dissolved in tetrahydrofuran (10 mL), and trimethyltrifluoromethylsilane (49.0 mg, 0.345 mmol) was added at room temperature and stirred for 12 hours. The reaction was quenched by the addition of water (20 mL). Extract with ethyl acetate (10 mL x 3). The combined organic layers were washed with brine, dried over anhydrous sodium sulfate Purification by preparative high performance liquid chromatography to give 3,7-dimethyl-1-(2-(4-methyl-2-(1,1,1-trifluoro-2-hydroxypropan-2-yl)) Thiazol-5-yl)ethyl)-1H-indole-2,6-(3H,7H)-dione (15.0 mg, yellow solid), yield: 31%. ¹ H NMR: (400 MHz, Methonal-d ₄ ) δ 8.37 (s, 1H), 4.25 (t, J = 6.4 Hz, 2H), 4.01 (s, 3H), 3.54 (s, 3H), 3.26 (t) , J = 6.4 Hz, 2H), 2.50 (s, 3H), 1.90 (s, 3H). MS-ESI calcd for [M ⁺ H] + 418, found 418.

Example 37

1-(3-hydroxy-2-(hydroxymethyl)-2-methylpropyl)-3,7-dimethyl-1H-indole-2,6-(3H,7H)-dione

3,7-Dimethyl-1-[(3-methyloxetan-3-yl)methyl]indole-2,6-dione (20.0 mg, 0.0757 mmol) was dissolved in 0.16% hydrochloric acid (0.5 mL), the reaction solution was stirred at room temperature for 6 hours, and the pH was adjusted to 7 with a saturated aqueous sodium hydrogen carbonate aqueous solution and purified by high-performance liquid chromatography to give 1-(3-hydroxy-2-(hydroxymethyl)- 2-Methylpropyl)-3,7-dimethyl-1H-indole-2,6-(3H,7H)-dione (12.0 mg, white solid), yield: 56%. ¹ H NMR: (400 MHz, CDCl ₃ ) δ 7.58 (s, 1H), 4.25-3.94 (m, 7H), 3.62 (s, 3H), 3.35-3.26 (m, 2H), 3.25-3.14 (m, 2H), 1.01 (s, 3H). MS-ESI calcd for [M+H] ⁺ 283.

Example 38

1-(2-(2-hydroxy-2-methylcyclopropyl)ethyl)-3,7-dimethyl-1H-indole-2,6(3H,7H)-dione

first step

2-(2-(3,7-Dimethyl-2,6-dioxo-2,3,6,7-tetrahydro-1H-indolyl)ethoxy)acetate

To a solution of sodium hydride (21.0 mg, 0.890 mmol) in N,N-dimethylformamide (10 mL), 1-(2-hydroxyethyl)-3,7-dimethyl-1H-嘌呤-2,6(3H,7H)-dione (100 mg, 0.446 mmol), and the mixture was stirred at 25 ° C for 1 hour. Additional ethyl 2-bromoacetate (149 mg, 0.890 mmol) was added. The reaction solution was stirred for further 16 hours. The insoluble matter was removed by filtration, and the filtrate was concentrated under reduced pressure, and purified by preparative high-performance liquid chromatography to give 2-(2-(3,7-dimethyl-2,6-dioxo-2,3,6,7- Hydrogen-1H-indenyl)ethoxy)acetate (60.0 mg, white solid), yield: 43%. MS-ESI calcd [M+H] ⁺ 311, Found 311.

Second step

1-(2-(2-hydroxy-2-methylcyclopropyl)ethyl)-3,7-dimethyl-1H-indole-2,6(3H,7H)-dione

2-(2-(3,7-Dimethyl-2,6-dioxo-2,3,6,7-tetrahydro-1H-indenyl)ethoxy)acetate at -78 °C A solution of (100 mg, 0.322 mmol) in tetrahydrofuran (5 mL) was slowly added dropwise a solution of methylmagnesium bromide (3M in tetrahydrofuran, 0.43 mL, 1.29 mmol). The reaction solution was stirred at -78 ° C for 2 hours. The reaction was quenched by the addition of saturated aqueous ammonium chloride (20 mL). The mixture was extracted with ethyl acetate (20 mL x 3). The organic phase was combined, concentrated under reduced pressure and purified by preparative HPLC to give 1-(2-(2-hydroxy-2-methylcyclopropyl)ethyl)-3,7-dimethyl-1H. - 嘌呤-2,6(3H,7H)-dione (40.0 mg, colorless oil). Yield: 42%. ¹ H NMR: (400 MHz, Methonal-d ₄ ) δ 7.88 (s, 1H), 4.23 (t, J = 5.8 Hz, 2H), 3.98 (s, 3H), 3.72 (t, J = 5.8 Hz, 2H) ), 3.53 (s, 3H), 3.32 (s, 2H), 1.13 (s, 6H). MS-ESI calcd for [M+H] ⁺ 296.

Example 39

1-(2-((1-hydroxycyclobutyl)methoxy)ethyl)-3,7-dimethyl-1H-indole-2,6(3H,7H)-dione

first step

1-(hydroxymethyl)cyclobutanol

A solution of 1-hydroxycyclobutyric acid (1.16 g, 10.0 mmol) in tetrahydrofuran (10 mL) was added dropwise to a solution of lithium tetrahydroaluminum (1.52 g, 40.0 mmol) in tetrahydrofuran (30 mL). The reaction solution was heated to reflux and allowed to react for 1 hour. The reaction mixture was cooled to 25 ° C, EtOAc (EtOAc)EtOAc. Butanol (0.800 g, colorless oil), yield: 80%.

Second step

2-(3,7-Dimethyl-2,6-dioxo-2,3,6,7-tetrahydro-1H-indol-1-yl)ethylmethanesulfonate

To a solution of 1-(2-hydroxyethyl)-3,7-dimethyl-1H-indole-2,6(3H,7H)-dione (448 mg, 2.00 mmol) in dichloromethane (25 mL) Triethylamine (600 mg, 6.00 mmol) and methanesulfonyl chloride (342 mg, 3.00 mmol) were added to the solution. The reaction solution was stirred at 0 ° C for 0.5 hours. The reaction was quenched with saturated aqueous NaHCO3 (30 mL). The organic phase was washed with brine (20 mL EtOAc) -tetrahydro-1H-indol-1-yl)ethylmethanesulfonate (650 mg, yellow oil), yield: 100%.

third step

1-(2-((1-hydroxycyclobutyl)methoxy)ethyl)-3,7-dimethyl-1H-indole-2,6(3H,7H)-dione

To the mixture 1-(hydroxymethyl)cyclobutanol (102 mg, 1.00 mmol) and 2-(3,7-dimethyl-2,6-dioxo-2,3,6,7-tetrahydro-1H- Potassium carbonate (414 mg, 3.00 mmol) and potassium iodide (16.0 mg, 0.100 mmol) were added to a solution of 嘌呤-1-yl)ethyl methanesulfonate (450 mg, 1.50 mmol) in N,N-dimethylformamide (5 mL). ). The reaction solution was heated to 60 ° C and stirred overnight. It was then slowly cooled to room temperature and quenched with water (20 mL). The mixture was extracted with EtOAc (20 mL EtOAc)EtOAc. Filtration, concentration of the filtrate under reduced pressure, and purification by preparative high performance liquid chromatography to give 1-(2-((1-hydroxycyclobutyl)methoxy)ethyl)-3,7-dimethyl-1H-indole -2,6(3H,7H)-dione (50.0 mg, white solid), yield: 16%. ¹ H NMR: (400 MHz, Methonal-d ₄ ) δ 7.88 (s, 1H), 4.57-4.59 (m, 2H), 4.21-4.24 (m, 2H), 3.98 (s, 3H), 3.80 (s, 2H), 3.54 (s, 3H), 2.07-1.95 (m, 4H), 1.52-1.54 (m, 2H). MS-ESI calcd [M+H] ⁺ 303.

Example 40

(S)-1-(2-((2-hydroxypropyl)amino)ethyl)-3,7-dimethyl-1H-indole-2,6(3H,7H)-dione

first step

1-(3-chloropropyl)-3,7-dimethyl-1H-indole-2,6(3H,7H)-dione

3,7-Dimethyl-1H-indole-2,6(3H,7H)-dione (1.00 g, 5.56 mmol) was dissolved in methanol (20 mL), 30% sodium methoxide (9.64 g, 49.9 mmol) The reaction solution was refluxed for 1 hour. Further, 1-bromo-2-chloroethane (47.2 g, 299 mmol) was added, and the mixture was stirred for 16 hr. The reaction was quenched with EtOAc (EtOAc (EtOAc)EtOAc. Separation and purification (1:2 petroleum ether / ethyl acetate) to give 1-(3-chloropropyl)-3,7-dimethyl-1H-indole-2,6(3H,7H)-dione (230 mg) , white solid), Yield: 17%. ^{1 H NMR: (400MHz, CDCl} 3) δ7.50 (s, 1H), 4.36 (t, J = 6.4Hz, 2H), 3.97 (s, 3H), 3.75 (t, J = 6.4Hz, 2H), 3.56(s,3H).

Second step

(S)-1-(2-((2-hydroxypropyl)amino)ethyl)-3,7-dimethyl-1H-indole-2,6(3H,7H)-dione

To the mixture 1-(3-chloropropyl)-3,7-dimethyl-1H-indole-2,6(3H,7H)-dione (62.4 mg, 0.826 mmol) and (S) at 25 °C Potassium carbonate (138 mg, 1.03 mmol) and potassium iodide (86.3 mg, 0.517 mmol) were added to a solution of 1-aminopropan-2-ol (50.0 mg, 0.207 mmol) in EtOAc (2 mL). The reaction solution was stirred at 90 ° C for 4 hours. The reaction was quenched with water (10 mL)EtOAcEtOAc The organic layer was washed with brine (20 mL EtOAc) Separation and purification by preparative high performance liquid chromatography gave (S)-1-(2-((2-hydroxypropyl)amino)ethyl)-3,7-dimethyl-1H-indole-2,6 (3H, 7H)-Dione (10.0 mg, white solid), yield: 17%. ^{1 H NMR: (400MHz, Methonal} -d 4) δ7.86 (s, 1H), 4.16 (t, J = 6.4Hz, 2H), 3.97 (s, 3H), 3.84 (m, 1H), 3.56 (s , 3H), 2.93 (m, 2H), 2.64 (m, 2H), 1.14 (d, J = 6.4 Hz, 3H). MS-ESI calcd for [M+H] ⁺ 282.

Example 41

first step

1-(1,4-Dioxaspiro[4.5]decane-8-ylmethyl)-7-(difluoromethyl)-3-methyl-1H-indole-2,6(3H,7H) -dione

1-(1,4-Dioxaspiro[4.5]decane-8-ylmethyl)-9-(difluoromethyl)-3-methyl-1H-indole-2,6(3H,7H) -dione

7-(Difluoromethyl)-3-methyl-1H-indole-2,6(3H,7H)-dione and 9-(difluoromethyl)-3-methyl-1H-indole-2 , a mixture of 6(3H,7H)-dione (200 mg, 0.930 mmol) was dissolved in N,N-dimethylformamide (20 mL), and the reaction mixture was added 1,4-dioxaspiro at room temperature [ 4.5] decane-8-ylmethyl methanesulfonate (245 mg, 1.10 mmol), potassium iodide (183 mg, 1.10 mmol) and potassium carbonate (303 mg, 2.20 mmol). The reaction solution was heated to 100 ° C and stirred for 2 hours. The reaction mixture was diluted with ethyl acetate (30 mL). EtOAc (EtOAc m. Methyl)-7-(difluoromethyl)-3-methyl-1H-indole-2,6(3H,7H)-dione and 1-(1,4-dioxaspiro[4.5]decane a mixture of -8-ylmethyl)-9-(difluoromethyl)-3-methyl-1H-indole-2,6(3H,7H)-dione (234 mg, yellow oil). 68%. MS-ESI calcd [M+H] ⁺ 372.

Second step

7-(Difluoromethyl)-3-methyl-1-((4-oxocyclohexyl)methyl)-1H-indole-2,6(3H,7H)-dione

9-(Difluoromethyl)-3-methyl-1-((4-oxocyclohexyl)methyl)-1H-indole-2,6(3H,7H)-dione

1-(1,4-Dioxaspiro[4.5]decane-8-ylmethyl)-7-(difluoromethyl)-3-methyl-1H-indole-2,6(3H,7H )-dione, 1-(1,4-dioxaspiro[4.5]decane-8-ylmethyl)-9-(difluoromethyl)-3-methyl-1H-indole-2,6 A mixture of (3H,7H)-dione (230 mg, 0.750 mmol) was dissolved in tetrahydrofuran (15 mL), and 10% hydrochloric acid (5 mL) was added at room temperature, and the mixture was heated to 50 ° C for 1 hour. The mixture was cooled to room temperature, diluted with EtOAc EtOAc (EtOAc)EtOAc. : 1 petroleum ether / ethyl acetate, Rf = 0.3) to give 7-(difluoromethyl)-3-methyl-1-((4-oxocyclohexyl)methyl)-1H-indole-2,6 ( 3H,7H)-dione,9-(difluoromethyl)-3-methyl-1-((4-oxocyclohexyl)methyl)-1H-indole-2,6(3H,7H)-di Mixture of ketone (200 mg, white solid), yield: 81%. MS-ESI calcd for [M+H] ⁺ 327.

third step

7-(Difluoromethyl)-1-(4-hydroxy-4-(trifluoromethyl)cyclohexyl)methyl)-3-methyl-1H-indole-2,6(3H,7H)-di ketone

9-(Difluoromethyl)-1-(4-hydroxy-4-(trifluoromethyl)cyclohexyl)methyl)-3-methyl-1H-indole-2,6(3H,7H)-di ketone

7-(Difluoromethyl)-3-methyl-1-((4-oxocyclohexyl)methyl)-1H-indole-2,6(3H,7H)-dione, 9-(difluoro A mixture of methyl)-3-methyl-1-((4-oxocyclohexyl)methyl)-1H-indole-2,6(3H,7H)-dione (168 mg, 0.515 mmol) was dissolved in tetrahydrofuran ( In 30 mL), trifluoromethyltrimethylsilane (109 mg, 0.773 mmol) and cesium fluoride (15.7 mg, 0.103 mmol) were added at room temperature. The reaction mixture was stirred at room temperature for 12 hr. EtOAc (EtOAc) (EtOAc) Drying with anhydrous sodium sulfate, filtering, and concentrating the filtrate under reduced pressure by high-performance liquid chromatography to give 7-(difluoromethyl)-1-(4-hydroxy-4-(trifluoromethyl)cyclohexyl)methyl) 3-methyl-1H-indole-2,6(3H,7H)-dione (54 mg, white solid), yield: 23%, ¹ H NMR: (400 MHz, Methanol-d ₄ ) δ 8.46 (s , 1H), 7.89-7.74 (m, 1H), 4.06 (d, J = 7.2 Hz, 2H), 3.59 (s, 3H), 2.19-2.17 (m, 1H), 2.05-1.99 (m, 2H), 1.88-1.81 (m, 2H), 1.61-1.58 (m, 2H), 1.51-1.47 (m, 2H). MS-ESI calcd for [M+H] ⁺ 397.

9-(Difluoromethyl)-1-(4-hydroxy-4-(trifluoromethyl)cyclohexyl)methyl)-3-methyl-1H-indole-2,6(3H,7H)-di Ketone (12 mg, white solid), yield: 10%. ¹ H NMR: (400 MHz, Methanol-d ₄ ) δ 8.49 (s, 1H), 7.93-7.89 (m, 1H), 4.07 (d, J = 7.2 Hz, 2H), 3.59 (s, 3H), 2.20 -2.19 (m, 1H), 2.05-1.99 (m, 2H), 1.88-1.85 (m, 2H), 1.61-1.58 (m, 2H), 1.51-1.48 (m, 2H). MS-ESI calcd for [M+H] ⁺ 397.

Example 42

7-Ethyl-1-(5-ethyl-5-hydroxyheptyl)-3-methyl-1H-indole-2,6(3H,7H)-dione

first step

7-Ethyl-3-methyl-1H-indole-2,6(3H,7H)-dione

3-Methyl-1H-indole-2,6(3H,7H)-dione (500 mg, 3.00 mmol), potassium carbonate (414 mg, 3.00 mmol) and potassium iodide (4.0 mg, 0.300 mmol) were dissolved in N, N - in dimethylformamide (15 mL). The reaction solution was heated to 80 ° C for half an hour. Iodoethane (470 mg, 4.50 mmol) was added. Continue the reaction for 5 hours. The reaction mixture was poured into aq. EtOAc (50 mL). The aqueous phase was adjusted to pH 7 with 1N dilute hydrochloric acid (10 mL), filtered, and dried to give 7-ethyl-3-methyl-1H-indole-2,6(3H,7H)-dione ( 500 mg, pale yellow solid), yield: 86%. ^{1 H NMR: (400MHz, DMSO} -d 6): δ8.05 (s, 1H), 4.25-4.19 (m, 2H), 3.34 (s, 3H), 1.37 (t, J = 7.2Hz, 3H). MS-ESI calculated [M+H] ⁺ 195, found 195.

Second step

Ethyl 5-(7-ethyl-3-methyl-2,6-dioxo-2,3,6,7-tetrahydro-1H-indol-1-yl)pentanoic acid ethyl ester

7-Ethyl-3-methyl-1H-indole-2,6(3H,7H)-dione (0.300 g, 1.55 mmol), ethyl bromopentanoate (480 mg, 2.32 mmol), potassium carbonate (430 mg) 3.10 mmol) and potassium iodide (26.0 mg, 0.155 mmol) were dissolved in N,N-dimethylformamide (4 mL). The reaction solution was heated to 110 ° C for 2 hours. The reaction was poured into water (20 mL). The combined organic layers were dried with anhydrous sodium sulfate, filtered and evaporated -1H-Indol-1-yl)ethyl valerate (320 mg, yellow solid), yield: 62%. MS-ESI calcd [M+H] ⁺ 323.

third step

7-Ethyl-1-(5-ethyl-5-hydroxyheptyl)-3-methyl-1H-indole-2,6(3H,7H)-dione

Ethyl 5-(7-ethyl-3-methyl-2,6-dioxo-2,3,6,7-tetrahydro-1H-indol-1-yl)pentanoic acid ethyl ester (0.100 g , 0.310 mmol) was dissolved in anhydrous tetrahydrofuran (10 mL), and ethyl magnesium bromide (3M tetrahydrofuran solution, 0.62 mL, 1.86 mmol) was slowly added dropwise at -78 °C. The reaction solution was reacted at -78 ° C for 0.5 hour, and slowly raised to 0 ° C for 0.5 hour. After the reaction was completed, the mixture was poured into water (20 mL) The organic phase was dried over anhydrous sodium sulfate, filtered, evaporated,lulululululululululululululululululululu 2,6(3H,7H)-dione (30.0 mg, colorless oil), yield: 30%.

^{1 H NMR: (400MHz, CDCl} 3): δ7.56 (s, 1H), 4.37-4.32 (m, 2H), 4.05 (t, J = 7.2Hz, 2H), 3.60 (s, 3H), 1.68- 1.37 (m, 13H), 0.86 (t, J = 7.2 Hz, 6H). MS-ESI calcd for [M ⁺ H] + 337, found 337.

Example 43

7-Ethyl-3-methyl-1-(6,6,6-trifluoro-5-hydroxy-5-methylhexyl)-1H-indole-2,6(3H,7H)-dione

First step

7-Ethyl-3-methyl-1-(5-oxohexyl)-1H-indole-2,6(3H,7H)-dione

7-Ethyl-3-methyl-1H-indole-2,6(3H,7H)-dione (0.100 g, 0.515 mmol), 6-chloro-2-pentanone (90.0 mg, 0.670 mmol), Potassium carbonate (140 mg, 1.03 mmol) and potassium iodide (8.5 mg, 0.0155 mmol) were dissolved in DMF (2 mL) and heated to 110 ° C for two hours. The reaction was poured into water and extracted with ethyl acetate (20 mL x 3). The organic phase was combined, dried, filtered, evaporated, evaporated, evaporated, evaporated 7H)-Dione (100 mg, white solid), Yield: 70%. MS-ESI calcd for [M ⁺ H] + 293, found 293.

Second step

7-Ethyl-3-methyl-1-(6,6,6-trifluoro-5-hydroxy-5-methylhexyl)-1H-indole-2,6(3H,7H)-dione

7-Ethyl-3-methyl-1-(5-oxohexyl)-1H-indole-2,6(3H,7H)-dione (100 mg, 0.340 mmol) was dissolved in 1 mL of THF. Trifluoromethyltrimethylsilane (53.0 mg, 0.370 mmol) and cesium fluoride (10.0 mg, 0.0340 mmol) were added in that order, and the mixture was reacted at 30 ° C for 3 hours. The reaction solution was poured into dilute hydrochloric acid (10%, 10 mL) and stirred for half an hour. Extract with ethyl acetate (20 mL x 3). The organic phase was combined, dried and concentrated, and the title compound was purified by preparative chromatography column 7-ethyl-3-methyl-1-(6,6,6-trifluoro-5-hydroxy-5-methylhexyl)- 1H-indole-2,6(3H,7H)-dione (20.0 mg, white solid), yield: 79%. ¹ H NMR: (400 MHz, CDCl ₃ ): δ 7.95 (s, 1H), 4.39-4.33 (m, 2H), 4.04-4.00 (m, 2H), 3.53 (s, 3H), 1.71-1.64 (m, 4H), 1.50-1.46 (m, 5H), 1.28 (s, 3H). MS-ESI calcd [M+H] ⁺

Example 44

1-((4-Hydroxy-4-(trifluoromethyl)cyclohexyl)methyl)-3-methyl-7-(2,2,2-trifluoroethyl)-1H-indole-2,6 (3H,7H)-dione

first step

1-(1,4-Dioxaspiro[4,5]decane-8-ylmethyl)-3-methyl-7-(2,2,2-trifluoroethyl)-1H-indole- 2,6(3H,7H)-dione

1,4-Dioxaspiro[4,5]decane-8-ylmethyl methanesulfonate (200 mg, 0.800 mmol), 3-methyl-7-(2,2,2-trifluoroethyl -1H-indole-2,6(3H,7H)-dione (200 g, 0.800 mmol) and potassium carbonate (334 mg, 2.42 mmol), potassium iodide (14.0 mg, 0.0800 mmol) dissolved in N,N-dimethyl In the formamide (3 mL), the reaction solution was heated to 130 ° C and stirred for 3.5 hours. The reaction solution was directly filtered, and the filtrate was concentrated under reduced pressure to give 1-(1,4-dioxaspiro[4,5]decane-8-ylmethyl)-3,7-dimethyl-1H-indole as a crude product. -2,6(3H,7H)-dione. MS-ESI calcd for [M+H] ⁺ 403.

Second step

3-methyl-1-((4-oxocyclohexyl)methyl)-7-(2,2,2-trifluoroethyl)-1H-indole-2,6(3H,7H)-dione

1-(1,4-Dioxaspiro[4,5]decane-8-ylmethyl)-3,7-dimethyl-1H-indole-2,6(3H,7H)-dione (2.50 g, 6.00 mmol) was dissolved in acetone (18 mL). The reaction was stirred at 30 ° C over EtOAc (EtOAc)EtOAc. The organic phase was combined, dried over anhydrous sodium sulfate, filtered, filtered,jjjjjjjjjjjjjj -oxocyclohexyl)methyl)-7-(2,2,2-trifluoroethyl)-1H-indole-2,6(3H,7H)-dione (220 mg, white solid) 11%. ^{1 H NMR: (400MHz, CDCl} 3) δ7.68 (s, 1H), 5.08-4.99 (m, 2H), 4.00 (d, J = 7.0Hz, 2H), 3.61 (s, 3H), 2.46-2.24 (m, 5H), 2.04-1.96 (m, 2H), 1.63-1.56 (m, 2H). MS-ESI calcd [M+H] ⁺ 359.

third step

1-((4-Hydroxy-4-(trifluoromethyl)cyclohexyl)methyl)-3-methyl-7-(2,2,2-trifluoroethyl)-1H-indole-2,6 (3H,7H)-dione

3-Methyl-1-((4-oxocyclohexyl)methyl)-7-(2,2,2-trifluoroethyl)-1H-indole-2,6(3H,7H)-di The ketone (128 mg, 0.360 mmol) and hydrazine fluoride (6.0 mg, 0.0360 mmol) were dissolved in tetrahydrofuran (3 mL) and trifluoromethyltrimethylsilane (77.0 mg, 0.540 mmol) was slowly added under nitrogen. The reaction solution was stirred at 30 ° C for 3 hours. After cooling to room temperature, 4N aqueous hydrochloric acid (2.5 mL) was added, and stirred at 25 ° C for half an hour, pH was adjusted to 7, diluted with water and extracted with ethyl acetate (20 mL x 3). The organic phase was combined, dried over anhydrous sodium sulfate, filtered, evaporated, evaporated Methyl-7-(2,2,2-trifluoroethyl)-1H-indole-2,6(3H,7H)-dione (14.0 mg, white solid), yield: 10%. ¹ H NMR: (400 MHz, CDCl ₃ ) δ 8.09 (s, 1H), 5.27-5.20 (m, 2H), 4.08-3.91 (m, 2H), 3.58 (s, 3H), 2.07-1.98 (m, 2H) ), 1.89-1.80 (m, 2H), 1.62-1.46 (m, 5H). MS-ESI calcd [M+H] ⁺ 422.

Example 45

1-((4-Hydroxy-4-(trifluoromethyl)cyclohexyl)methyl)-3-methyl-7-(2,2,2-trifluoroethyl)-1H-indole-2,6 (3H,7H)-dione (500 mg, 1.17 mmol) was isolated by preparative SFC to afford two isomers. Separation conditions: Column: AD 250 mm x 30 mm, 10 um mobile phase: A: supercritical carbon dioxide, B: ethanol (0.05% aqueous ammonia), A: B = 550:45 flow rate: 80 mL/min Wavelength: 220 nm.

Product 1 (isomer 1, first peak) (300 mg, white solid), yield: 90%. ^{1 H NMR: (400MHz, DMSO} -d 6) δ8.23 (s, 1H), 5.64 (s, 1H), 5.31-5.24 (m, 2H), 3.89 (d, J = 3.6Hz, 2H), 3.43 (s, 3H), 2.06-2.05 (m, 1H), 1.87-1.81 (m, 2H), 1.73-1.61 (m, 2H), 1.49-1.45 (m, 2H), 1.33-1.31 (m, 2H) .MS ESI calc'd.[M+H] ⁺ 429,found 429.

Product 2 (isomer 2, second peak) (150 mg, white solid), yield 90%. ^{1 H NMR: (400MHz, DMSO} -d 6) δ8.22 (s, 1H), 5.63 (s, 1H), 5.29-5.23 (m, 2H), 3.74 (d, J = 3.6Hz, 2H), 3.42 (s, 3H), 1.68-1.66 (m, 3H), 1.45-1.31 (m, 6H). MS ESI calc'd. [M+H] ⁺ 429, found 429.

Example 46

first step

1-(1,4-Dioxaspiro[4,5]decane-8-ylmethyl)-3-methyl-7-(2,2,2-trifluoroethyl)-1H-indole- 2,6-(3H,7H)-dione

1,4-Dioxaspiro[4,5]decane-8-ylmethyl methanesulfonate (603 mg, 2.41 mmol), 3-methyl-7-(2,2,2-trifluoroethyl -1H-indole-2,6-(3H,7H)-dione (500 mg, 2.01 mmol) and potassium iodide (33.3 mg, 0.201 mmol) were dissolved in N,N-dimethylformamide (8 mL). Potassium carbonate (555 mg, 4.02 mmol) was added, and the mixture was heated to reflux at 130 ° C for 4 hours. The reaction solution was cooled to room temperature, filtered, and the filtrate was evaporated evaporated. (2,2,2-Trifluoroethyl)-1H-indole-2,6-(3H,7H)-dione (980 mg, yellow oil). MS-ESI calcd for [M+H] ⁺ 403.

Second step

3-methyl-1-((4-oxocyclohexyl)methyl)-7-(2,2,2-trifluoroethyl)-1H-indole-2,6-(3H,7H)-di ketone

1-(1,4-Dioxaspiro[4,5]decane-8-ylmethyl)-3-methyl-7-(2,2,2-trifluoroethyl)-1H-indole -2,6-(3H,7H)-dione (980 mg, 1.51 mmol) was dissolved in acetone (8 mL). The reaction was stirred at rt EtOAc (EtOAc)EtOAc. 1 petroleum ether / ethyl acetate, Rf = 0.3) to give the product 3-methyl-1-((4-oxocyclohexyl)methyl)-7-(2,2,2-trifluoroethyl)-1H - 嘌呤-2,6-(3H,7H)-dione (78.0 mg, yellow solid), yield: 15%. MS-ESI calcd [M+H] ⁺ 359.

3-Methyl-1-((4-oxocyclohexyl)methyl)-7-(2,2,2-trifluoroethyl)-1H-indole-2,6-(3H,7H)- The diketone (66.0 mg, 0.184 mmol) was dissolved in tetrahydrofuran (2 mL), and the methyl Grignard reagent (3M diethyl ether solution, 0.184 mL, 0.552 mmol) was slowly added under a nitrogen atmosphere at -78 ° C, and stirred at -78 ° C for half an hour. The reaction was then carried out at 0 ° C for 2 hours. After adding water (10 mL), the mixture was extracted with ethyl acetate (30 mL×3), and the organic phase was dried over anhydrous sodium sulfate. White solid), Yield: 12%, ¹ H NMR: (400 MHz, Methonal-d ₄ ) δ 8.08 (s, 1H), 5.27-5.19 (m, 2H), 3.92 (d, J = 7.2 Hz, 2H ), 3.58 (s, 3H), 1.71-1.62 (m, 4H), 1.46-1.38 (m, 2H), 1.32-1.18 (m, 6H). MS-ESI calcd for ^{_{[M + HH 2 O] +}} 357, found 357.

Product 2 (12.0 mg, white solid) (isomer 2, second peak), yield: 17%. ^{1 H NMR: (400MHz, Methonal} -d4) δ8.08 (s, 1H), 5.27-5.21 (m, 2H), 3.91 (d, J = 7.2Hz, 2H), 3.57 (s, 3H), 1.69- 1.66 (m, 2H), 1.49-1.44 (m, 3H), 1.37-1.28 (m, 4H), 1.17 (s, 3H). MS-ESI calcd for ^{_{[M + HH 2 O] +}} 357, found 357.

Example 47

7-Cyclopropyl-3-methyl-1-(6,6,6-trifluoro-5-hydroxy-5-methylhexyl)-1H-indole-2,6(3H,7H)-dione

first step

6-Amino-5-bromo-1-methylpyrimidine-2,4(1H,3H)-dione

Mixture of 6-amino-1-methylpyrimidine-2,4(1H,3H)-dione (5.46 g, 40.0 mmol) and bromosuccinimide (7.56 g, 42.0 mmol) in acetonitrile (100 mL) Heated under nitrogen for 1.5 hours. The reaction solution was cooled to room temperature, filtered, and the solvent was evaporated, and then evaporated, evaporated, evaporated,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,, White solid), Yield: 98%. ^{1 H NMR: (400MHz, DMSO} -d6) δ10.90 (s, 1H), 7.04 (s, 2H), 3.28 (s, 3H).

Second step

6-Amino-5-(cyclopropylamine)-1-methylpyrimidine-2,4(1H,3H)-dione

6-Amino-5-bromo-1-methylpyrimidine-2,4(1H,3H)-dione (2.19 g, 10.0 mmol) was dissolved in a mixed solvent of cyclopropylamine (20 mL) and water (5 mL). The reaction solution was heated to reflux for 5 hours. The reaction solution was filtered to remove the solvent to give the crude product 6-amino-5-(cyclopropylamine)-1-methylpyrimidine-2,4(1H,3H)-dione.

third step

7-Cyclopropyl-3-methyl-1H-indole-2,6(3H,7H)-dione

6-Amino-5-(cyclopropylamine)-1-methylpyrimidine-2,4(1H,3H)-dione (1.96 g, 10.0 mmol), trimethyl orthoformate (2.12) under N2 g, 20.0 mmol) and p-toluenesulfonic acid (86.0 mg, 0.500 mmol) were dissolved in anhydrous N,N-dimethylformamide (20 mL). The reaction solution was heated to 100 ° C overnight. The reaction solution was filtered, and the solvent was evaporated to give the crude product 7-cyclopropyl-3-methyl-1H-indole-2, 6(3H,7H)-dione.

the fourth step

7-Cyclopropyl-3-methyl-1-(5-oxohexane)-1H-indole-2,6(3H,7H)-dione

7-Cyclopropyl-3-methyl-1H-indole-2,6(3H,7H)-dione (100 mg, 0.480 mmol), 6-chlorohexane-2-one (97.0 mg) under nitrogen atmosphere , 0.730 mmol) and potassium carbonate (132 mg, 0.960 mmol) in N,N-dimethylformamide (5 mL). The reaction solution was heated to 120 ° C for 3 hours. After the reaction was cooled to room temperature, it was diluted with water (20 mL) and ethyl acetate (10 mL), ethyl acetate (30 mL). 3-Methyl-1-(5-oxohexane)-1H-indole-2,6(3H,7H)-dione was used directly in the next step. MS-ESI calcd for [M+H] ⁺ 303.

the fifth step

7-Cyclopropyl-3-methyl-1-(6,6,6-trifluoro-5-hydroxy-5-methylhexyl)-1H-indole-2,6(3H,7H)-dione

7-Cyclopropyl-3-methyl-1-(5-oxohexane)-1H-indole-2,6(3H,7H)-dione (200 mg, 0.660 mmol) was dissolved in a nitrogen atmosphere. Trifluoromethyltrimethylsilane (0.2 mL, 0.990 mmol) and cesium fluoride (20.0 mg, 0.130 mmol) were added sequentially in anhydrous tetrahydrofuran (3 mL). The resulting reaction solution was reacted at 30 ° C for 2 hours. The reaction mixture was diluted with water (30 mL), EtOAc (EtOAc)EtOAc. , Rf = 0.3) gives 7-cyclopropyl-3-methyl-1-(6,6,6-trifluoro-5-hydroxy-5-methylhexyl)-1H-indole-2,6 (3H, 7H)-Dione (150 mg, white solid). Yield: 61%. ¹ H NMR: (400MHz, CDCl ₃ ) δ 7.55 (s, 1H), 4.3-4.02 (m, 2H), 3.63-3.61 (m, 1H), 3.55 (s, 3H), 2.96 (s, 1H) , 1.90- 1.68 (m, 2H), 1.67-1.64 (m, 2H), 1.47-1.45 (m, 2H), 1.28 (s, 3H), 1.18-1.16 (m, 2H), 1.06-1.04 (m, 2H). MS-ESI calcd for [M+H] ⁺ 372.

Example 48

7-(Cyclopropylmethyl)-1-((4-hydroxy-4-(trifluoromethyl)cyclohexyl)methyl)-3-methyl-1H-indole-2,6(3H,7H) -dione

first step

1-(1,4-Dioxaspiro[4.5]decane-8-ylmethyl)-7-isopropyl-3-methyl-1H-indole-2,6(3H,7H)-dione

1,4-Dioxaspiro[4.5]decane-8-ylmethyl methanesulfonate (250 mg, 1.00 mmol), 7-isopropyl-3-methyl-1H-indole-2,6 ( 3H,7H)-Dione (208 mg, 1.00 mmol), potassium iodide (15.8 mg, 0.100 mmol) and potassium carbonate (276 mg, 2.00 mmol) were dissolved in anhydrous N,N-dimethylformamide (8 mL). The reaction solution was heated to 120 ° C and stirred for 3 hours. The reaction was cooled to 20 ° C, and the mixture was filtered. -1H-indole-2,6(3H,7H)-dione (300 mg, white oil). MS-ESI calcd for [M+H] ⁺ 372.

Second step

7-Isopropyl-3-methyl-1-((4-oxocyclohexyl)methyl)-1H-indole-2,6(3H,7H)-dione

1-(1,4-Dioxaspiro[4.5]decane-8-ylmethyl)-7-isopropyl-3-methyl-1H-indole-2,6(3H,7H)-di The ketone (300 mg, 0.828 mmol) was dissolved in acetone (10 mL) and hydrochloric acid (0.5 mL). The reaction was stirred at room temperature for 30 minutes. Water was added to the reaction mixture, and the mixture was evaporated, evaporated, evaporated,,,,,,,,,,,,,, The organic layer was dried over anhydrous sodium sulfate, filtered, and evaporated. Hexyl)methyl)-1H-indole-2,6(3H,7H)-dione (180 mg, white oil), yield: 68%. MS-ESI calcd for [M+H] ⁺ 319.

third step

7-Isopropyl-3-methyl-1-((4-hydroxy-4-(trifluoromethyl)cyclohexyl)methyl)-1H-indole-2,6(3H,7H)-dione

7-Isopropyl-3-methyl-1-((4-oxocyclohexyl)methyl)-1H-indole-2,6(3H,7H)-dione (122 mg, 0.382 mmol) and fluorinated The hydrazine (11.5 mg, 0.0763 mmol) was dissolved in anhydrous tetrahydrofuran (3 mL) and trifluoromethyltrimethylsilane (9. The reaction solution was heated to 30 ° C and stirred for 12 hours. Then aqueous hydrochloric acid (1 N, 5 mL) was added and stirred for additional 30 min. Water was added to the reaction mixture, and the mixture was evaporated. EtOAcjjjjjjjjjjjjjjjjjj Dry over anhydrous sodium sulfate, filter, and concentrate the filtrate under reduced pressure and purified by preparative high-performance liquid chromatography to give 7-isopropyl-3-methyl-1-((4-hydroxy-4-(trifluoromethyl)) Cyclohexyl)methyl)-1H-indole-2,6(3H,7H)-dione (80.0 mg, white solid), yield: 53%. ^{1 H NMR: (400MHz, Methonal} -d 4) δ8.10 (s, 1H), 5.06-5.00 (m, 1H), 4.08-3.91 (m, 2H), 3.55 (s, 3H), 2.17-2.00 ( m, 2H), 1.88-1.84 (m, 2H), 1.61-1.40 (m, 6H), 1.59-1.57 (m, 5H). MS-ESI calcd for [M ⁺ H] + 389, found 389.

Example 49

7-(Cyclopropylmethyl)-1-((4-hydroxy-4-(trifluoromethyl)cyclohexyl)methyl)-3-methyl-1H-indole-2,6(3H,7H) -dione

first step

1-(1,4-Dioxaspiro[4.5]decane-8-ylmethyl)-7-(cyclopropylmethyl)-3-methyl-1H-indole 2,6(3H,7H) -dione

(cyclopropylmethyl)-3-methyl-1H-indole-2,6(3H,7H)-dione (220 mg, 1.00 mmol), 1,4-dioxaspiro[4.5]decane- 8-ylmethyl methanesulfonate (250 mg, 1.00 mmol), potassium iodide (15.8 mg, 0.100 mmol) and potassium carbonate (276 mg, 2.00 mmol) dissolved in anhydrous N,N-dimethylformamide (8 mL) The reaction solution was heated to 120 ° C and stirred for 3 hours. The reaction was cooled to 20 ° C, and the mixture was filtered and evaporated tolululululululululululululululululululu -Methyl-1H-indole-2,6(3H,7H)-dione (300 mg, white oil), yield: 80%. MS-ESI calcd for [M+H] ⁺ 372.

Second step

7-(Cyclopropylmethyl)-3-methyl-1-((4-oxocyclohexyl)methyl)-1H-indole-2,6(3H,7H)-dione

1-(1,4-Dioxaspiro[4.5]decane-8-ylmethyl)-3,7-dimethyl-1H-indole-2,6(3H,7H)-dione (300 mg , 0.802 mmol) was dissolved in acetone (10 mL), hydrochloric acid (0.5 mL) was added, and the mixture was stirred at room temperature for 30 min. Water (30 mL) was added to the mixture and the mixture was evaporated. The filtrate was concentrated under reduced pressure and purified by silica gel column chromatography (ethyl acetate, Rf=0.3) to afford 7-(cyclopropylmethyl)-3-methyl-1-((4-oxocyclohexyl)methyl -1H-indole-2,6(3H,7H)-dione (180 mg, white oil), yield: 75%. MS-ESI calcd for [M ⁺ H] + 331, found 331.

third step

7-(Cyclopropylmethyl)-1-((4-hydroxy-4-(trifluoromethyl)cyclohexyl)methyl)-3-methyl-1H-indole-2,6(3H,7H) -dione

7-(Cyclopropylmethyl)-3-methyl-1-((4-oxocyclohexyl)methyl)-1H-indole-2,6(3H,7H)-dione (126 mg, 0.382 mmol) And cesium fluoride (11.5 mg, 0.0763 mmol) were dissolved in anhydrous tetrahydrofuran (3 mL), and trifluoromethyltrimethylsilane (95.0 mg, 0.640 mmol) was added under nitrogen. The mixture was stirred at 30 ° C for 12 hours. Then 1N aqueous hydrochloric acid (5 mL) was added and stirring was continued for further 30 min. Water (30 mL) was added to the reaction mixture, EtOAc (EtOAc) (EtOAc) Washing, drying over anhydrous sodium sulfate, filtration, concentration of the filtrate under reduced pressure, and purification by preparative high-performance liquid chromatography to give 7-(cyclopropylmethyl)-1-((4-hydroxy-4-(trifluoro)) Cyclo)yl)methyl)-3-methyl-1H-indole-2,6(3H,7H)-dione (80.0 mg, white solid), yield: 53%. ¹ H NMR: (400 MHz, Methonal-d ₄ ) δ 8.16-8.15 (m, 1H), 4.24-4.22 (m, 2H), 4.08-3.91 (m, 2H), 3.57 (s, 3H), 2.18- 2.07 (m, 2H), 1.85-1.82 (m, 2H), 1.61-1.47 (m, 6H), 0.64-0.60 (m, 2H), 0.50-0.48 (m, 2H). MS-ESI calcd for [M+H] ⁺ 401.

Example 50

7-(Cyclopropylmethyl)-1-((4-hydroxy-4-(trifluoromethyl)cyclohexyl)methyl)-3-methyl-1H-indole-2,6(3H,7H) Di-one (900 mg, 2.25 mmol) was isolated by preparative SFC to afford two isomers. Separation conditions: column: AD 250 mm x 30 mm, 5 um mobile phase: A: supercritical carbon dioxide, B: methanol (0.05% aqueous ammonia), A: B = 55:45 flow rate: 40 mL/min wavelength: 220 nm. Product 1 (isomer 1, first peak) (600 mg, white solid), yield: 100%. ^{1 H NMR: (400MHz, DMSO} -d 6) δ8.11 (s, 1H), 5.62 (s, 1H), 4.08 (d, J = 7.6Hz, 2H), 3.88 (d, J = 7.6Hz, 2H ), 3.43 (s, 3H), 2.05-2.04 (m, 1H), 1.85-1.82 (m, 2H), 1.48-1.45 (m, 2H), 1.33-1.32 (m, 2H), 1.30-1.28 (m , 3H), 0.48-0.46 (m, 2H), 0.41-0.39 (m, 2H). MS-ESI calcd for [M+H] ⁺ 401.

Product 2 (isomer 2, second peak) (300 mg, white solid), yield 100%. ¹ H NMR: (400MHz, DMSO-d ₆ ) δ 8.11 (s, 1H), 5.62 (s, 1H), 4.09 (d, J = 7.6 Hz, 2H), 3.74 (d, J = 7.6 Hz, 2H) ), 3.42 (s, 3H), 1.69-1.45 (m, 3H), 1.45-1.29 (m, 7H), 0.48-0.46 (m, 2H), 0.41 - 0.39 (m, 2H). MS-ESI calcd for [M+H] ⁺ 401.

Example 51

first step

1-(1,4-Dioxaspiro[4,5]decane-8-ylmethyl-7-(cyclopropylmethyl)-3-methyl-1H-indole-2,6-(3H ,7H)-dione

1,4-Dioxaspiro[4,5]nonane-8-ylmethyl methanesulfonate (682 mg, 2.72 mmol), 7-(cyclopropylmethyl)-3-methyl-1H-嘌呤-2,6-(3H,7H)-dione (500 mg, 2.27 mmol) and potassium iodide (37.7 mg, 0.227 mmol) were dissolved in N,N-dimethylformamide (10 mL), and potassium carbonate (627 mg) , 4.54 mmol), and the reaction was heated to reflux at 130 ° C for 4 hours. The reaction solution was cooled to room temperature, filtered, and the filtrate was evaporated evaporated evaporated. ) -3-methyl -1H- purine -2,6- (3H, 7H) - dione (1.10g, yellow oil) .MS-ESI calcd for [M ⁺ H] + 375, found 375.

Second step

7-(Cyclopropylmethyl)-3-methyl-1-((4-oxocyclohexyl)methyl)-1H-indole-2,6-(3H,7H)-dione

1-(1,4-Dioxaspiro[4,5]decane-8-ylmethyl-7-(cyclopropylmethyl)-3-methyl-1H-indole-2,6-( 3H,7H)-dione (1.20 g, 2.09 mmol) was dissolved in EtOAc (EtOAc) (EtOAc) (EtOAc) The organic phase is dried over anhydrous sodium sulfate, filtered, and the filtrate is evaporated, evaporated, mjjjjjjjjjjjjjj Methyl)-3-methyl-1-((4-oxocyclohexyl)methyl)-1H-indole-2,6-(3H,7H)-dione (52.0 mg, yellow solid). 8% .MS-ESI calcd for [M ⁺ H] + 331, found 331.

third step

7-(Cyclopropylmethyl)-3-methyl-1-((4-oxocyclohexyl)methyl)-1H-indole-2,6-(3H,7H)-dione (100 mg, 0.303 Methyl) was dissolved in tetrahydrofuran (5 mL). Methyl Grignard reagent (3M diethyl ether solution, 0.600 mL, 1.81 mmol) was slowly added under nitrogen at -78 ° C, stirred at -78 ° C for half an hour, and then reacted at 0 ° C for 2 hours. Water (10 mL) was added, and the mixture was evaporated. White solid) (isomer 1, first peak), yield: 25%. ¹ H NMR: (400 MHz, Methonal-d ₄ ) δ 7.99 (s, 1H), 4.19 (d, J = 7.6 Hz, 2H), 3.90 (d, J = 7.6 Hz, 2H), 3.54 (s, 3H) ), 1.90- 1.79 (m, 1H), 1.70-1.61 (m, 4H), 1.45-1.36 (m, 3H), 1.27-1.16 (m, 5H), 0.65-0.55 (m, 2H), 0.49-0.42 (m, 2H). MS-ESI calcd for ^{_{[M + HH 2 O] +}} 329, found 329.

Product 2 (42.0 mg, white solid) (isomer 2, second peak), yield: 40%. ¹ H NMR: (400 MHz, Methonal-d ₄ ) δ 7.99 (s, 1H), 4.19 (d, J = 7.6 Hz, 2H), 3.89 (d, J = 7.6 Hz, 2H), 3.54 (s, 3H) ), 1.81-1.70 (m, 1H), 1.69-1.62 (m, 2H), 1.51-1.41 (m, 4H), 1.39-1.25 (m, 3H), 1.15 (s, 3H), 0.63-0.56 (m) , 2H), 0.48-0.42 (m, 2H). MS-ESI calcd for ^{_{[M + HH 2 O] +}} 329, found 329.

Example 52

1-((4-Hydroxy-1-methyl-4-(trifluoromethyl)cyclohexyl)methyl)-3,7-dimethyl-1H-indole-2,6-(3H,7H)- Diketone

first step

1-((4-Hydroxy-1-methyl-4-(trifluoromethyl)cyclohexyl)methyl)-3,7-dimethyl-1H-indole-2,6-(3H,7H)- Diketone

(4-Hydroxy-1-(methoxymethyl)-4-(trifluoromethyl)cyclohexyl)methyl methanesulfonate (100 mg, 0.349 mmol, 7-(cyclopropylmethyl)-3- Methyl-1H-indole-2,6-(3H,7H)-dione (76.9 mg, 0.349 mmol), potassium iodide (5.8 mg, 0.0349 mmol) and potassium carbonate (149 mg, 1.05 mmol) dissolved in anhydrous N. N-dimethylformamide (5 mL). The reaction solution was heated to 150 ° C in the microwave for 2 hours. The reaction solution was cooled to 20 ° C, filtered, and purified by preparative high-performance liquid chromatography to give 1-((4-hydroxy-) 1-Methyl-4-(trifluoromethyl)cyclohexyl)methyl)-3,7-dimethyl-1H-indole-2,6-(3H,7H)-dione (10.0 mg, white solid ), Yield: 6%. ¹ H NMR: (400MHz, DMSO-d ₆ ) δ 8.13 (s, 1H), 4.12 (s, 2H), 3.94 (s, 1H), 3.43-3.38 (m, 4H) ), 3.31 (s, 3H), 3.19 (s, 3H), 1.56-1.45 (m, 8H), 1.43-1.31 (m, 1H), 0.51-0.49 (m, 2H), 0.44-0.42 (m, 2H) ).

MS-ESI calcd for [M+H] ⁺ 445.

Example 53

7-(Cyclopropylmethyl)-1-((4-hydroxy-1-methyl-4-(trifluoromethyl)cyclohexyl)methyl)-3-methyl-1H-indole-2,6 -(3H,7H)-dione

first step

7-(Cyclopropylmethyl)-1-((4-hydroxy-1-methyl-4-(trifluoromethyl)cyclohexyl)methyl)-3-methyl-1H-indole-2,6 -(3H,7H)-dione

(4-Hydroxy-1-methyl-4-(trifluoromethyl)cyclohexyl)methyl methanesulfonate (100 mg, 0.344 mmol), 7-(cyclopropylmethyl)-3-methyl-1H -嘌呤-2,6-(3H,7H)-dione (75.9 mg, 0.344 mmol), potassium iodide (5.70 mg, 0.0344 mmol) and potassium carbonate (47.6 mg, 0.344 mmol) dissolved in anhydrous N,N- Methylformamide (5 mL). The reaction solution was heated to 150 ° C and microwaved for 4 hours. The reaction solution was cooled to 20 ° C, filtered, concentrated, and then purified by preparative HPLC to give 7-(cyclopropylmethyl)-1-((4-hydroxy-1-methyl-4-(trifluoro) Cyclo) hexyl)methyl)-3-methyl-1H-indole-2,6-(3H,7H)-dione (10.0 mg, white solid). ¹ H NMR: (400 MHz, DMSO-d ₆ ) δ 8.13 (s, 1H), 4.13-4.09 (m, 2H), 3.83 (s, 1H), 3.43 (s, 3H), 3.34 (s, 2H) , 1.67-1.53 (m, 6H), 1.23-1.20 (m, 3H), 0.88 (s, 3H), 0.50-0.42 (m, 4H). MS-ESI calcd for [M+H] ⁺ 415.

Example 54

7-(cyclopropylmethyl)-3-methyl-1-((5-methyl-2-(1,1,1-trifluoro-2-hydroxypropan-2-yl)thiazol-4-yl )methyl)-1H-嘌呤2,6-(3H,7H)-dione

first step

1-(4-(bromomethyl)-5-methylthiazol-2-yl)ethanone

1-(4,5-Dimethylpyridin-2-yl)ethanone (200 mg, 1.29 mmol), N-bromosuccinimide (229 mg, 1.29 mmol), azobisisobutyronitrile (21.2 Mg, 0.129 mmol) was dissolved in carbon tetrachloride (20 mL) and reacted at 80 ° C for 12 hours. The reaction was quenched by the addition of saturated sodium thiosulfate solution (30 mL). It was extracted with dichloromethane (10 mL x 3), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to give 1-(4-(bromomethyl)-5-methylthiazol-2-yl)ethanone (200 mg) , yellow oily), yield: 46%. MS-ESI calcd for [M+H] &^lt;

Second step

1-((2-acetyl-5-methylthiazol-4-yl)methyl)-7-(cyclopropylmethyl)-3-methyl-1H-indole-2,6-(3H,7H )-dione

1-(4-(Bromomethyl)-5-methylthiazol-2-yl)ethanone (200 mg, 0.598 mmol), 7-(cyclopropylmethyl)-3-methyl-1H-indole- 2,6-Dione (132 mg, 0.598 mmol), potassium iodide (19.8 mg, 0.119 mmol) and potassium carbonate (248 mg, 1.79 mmol) were dissolved in N,N-dimethylformamide (10 mL). The reaction solution was warmed to 120 ° C and stirred for 3 hours. It was cooled to room temperature, filtered, and the filtrate was concentrated under reduced pressure and purified by preparative TLC (1:1 petroleum ether / ethyl acetate, Rf = 0.4) to give 1-((2-acetyl-5-methylthiazole-4-) Methyl)-7-(cyclopropylmethyl)-3-methyl-1H-indole-2,6-(3H,7H)-dione (100 mg, yellow solid), yield: 45%. ^{1 H NMR: (400MHz, Methonal} -d 4) δ8.02 (s, 1H), 5.35 (s, 2H), 4.21 (d, J = 7.6Hz, 2H), 3.56 (s, 3H), 2.66 (s , 3H), 2.60 (s, 3H), 1.46-1.41 (m, 1H), 0.65-0.61 (m, 2H), 0.60-0.48 (m, 2H). MS-ESI calcd [M+H] ⁺ 372.

third step

7-(cyclopropylmethyl)-3-methyl-1-((5-methyl-2-(1,1,1-trifluoro-2-hydroxypropan-2-yl)thiazol-4-yl )methyl)-1H-嘌呤2,6-(3H,7H)-dione

1-((2-Acetyl-5-methylthiazol-4-yl)methyl)-7-(cyclopropylmethyl)-3-methyl-1H-indole-2,6-(3H, 7H)-Dione (100 mg, 0.267 mmol), cesium fluoride (40.6 mg, 0.267 mmol) was dissolved in tetrahydrofuran (10 mL), and trimethyl-trifluoromethyl-silane (114 mg, 0.803 mmol) was added at room temperature. Stir for 12 hours. The reaction was quenched by the addition of water (20 mL). Extract with ethyl acetate (10 mL x 3). The combined organic layers were washed with brine, dried over anhydrous sodium sulfate Purification by preparative high performance liquid chromatography to give 7-(cyclopropylmethyl)-3-methyl-1-((5-methyl-2-(1,1,1-trifluoro-2-hydroxypropane) 2-yl)thiazol-4-yl)methyl)-1H-indole 2,6-(3H,7H)-dione (50.0 mg, yellow solid), yield: 42%. ¹ H NMR: (400 MHz, Methonal-d ₄ ) δ 8.11 (s, 1H), 5.33 (s, 2H), 4.23 (d, J = 7.6 Hz, 2H), 3.57 (s, 3H), 2.64 (s) , 3H), 1.81 (s, 3H), 1.45-1.41 (m, 1H), 0.65-0.61 (m, 2H), 0.60-0.49 (m, 2H). MS-ESI calcd for [M+H] ⁺ 444.

Example 55

7-(Cyclopropylmethyl)-3-methyl-1-((6-(1,1,1-trifluoro-2-hydroxypropan-2-yl)pyridin-3-yl)methyl)- 1H-嘌呤2,6-(3H,7H)-dione

first step

1-[5-(bromomethyl)-2-pyridyl]ethanone

1-(5-Methyl-2-pyridyl)ethanone (500 mg, 3.70 mmol), N-bromosuccinimide (658 mg, 3.70 mmol), azobisisobutyronitrile (182 mg, 1.11 mmol) Dissolved in carbon tetrachloride (20 mL) and reacted at 90 ° C for 12 hours. The reaction was quenched by the addition of saturated sodium thiosulfate solution (30 mL). The mixture was extracted with EtOAc (EtOAc m. Yield: 16%. MS-ESI calculated [M+H] ⁺ 214 and 216, found 214 and 216.

Second step

1-[(6-acetyl-3-pyridyl)methyl]-7-(cyclopropylmethyl)-3-methyl-1H-indole-2,6-dione

1-[5-(Bromomethyl)-2-pyridyl]ethanone (100 mg, 0.467 mmol), 7-(cyclopropylmethyl)-3-methyl-1H-indole-2,6-di Ketone (103 mg, 0.467 mmol), potassium iodide (15.5 mg, 0.0934 mmol) and potassium carbonate (193 mg, 1.40 mmol) were dissolved in N,N-dimethylformamide (10 mL). The reaction solution was warmed to 120 ° C and stirred for 3 hours. After cooling to room temperature, filtration, the filtrate was concentrated under reduced pressure and purified by preparative TLC (ethyl acetate, Rf = 0.4) to give 1-[(6-acetyl-3-pyridyl)methyl]-7-(cyclopropyl) Methyl)-3-methyl-1H-indole-2,6-dione (50.0 mg, yellow solid), yield: 30%. MS-ESI calcd [M+H] ⁺ 355.

third step

7-(Cyclopropylmethyl)-3-methyl-1-((6-(1,1,1-trifluoro-2-hydroxypropan-2-yl)pyridin-3-yl)methyl)- 1H-嘌呤2,6- (3H,7H)-dione

1-[(6-Acetyl-3-pyridyl)methyl]-7-(cyclopropylmethyl)-3-methyl-1H-indole-2,6-dione (100 mg, 0.283 mmol) The cesium fluoride (43.0 mg, 0.283 mmol) was dissolved in tetrahydrofuran (10 mL), and trimethyl-trifluoromethyl-silane (60.4 mg, 0.424 mmol) was added at room temperature and stirred for 12 hours. The reaction was quenched by the addition of water (20 mL). Extract with ethyl acetate (10 mL x 3). The combined organic layers were washed with brine, dried over anhydrous sodium sulfate Purification by preparative high performance liquid chromatography to give 7-(cyclopropylmethyl)-3-methyl-1-((6-(1,1,1-trifluoro-2-hydroxypropan-2-yl)) Pyridin-3-yl)methyl)-1H-indole 2,6-(3H,7H)-dione (40.0 mg, yellow solid), yield: 32%. ^{1 H NMR: (400MHz, Methonal} -d 4) δ9.00 (s, 1H), 8.80-8.72 (m, 1H), 8.46 (s, 1H), 8.35-8.29 (m, 1H), 5.43 (s, 2H), 4.28 (d, J = 7.6 Hz, 2H), 3.58 (s, 3H), 1.95 (s, 3H), 1.50-1.46 (m, 1H), 0.68-0.64 (m, 2H), 0.53-0.51 (m, 2H). MS-ESI calcd for [M ⁺ H] + 424, found 424.

Example 56

7-(cyclopropylmethyl)-3-methyl-1-((5-methyl-2-(1,1,1-trifluoro-2-hydroxypropan-2-yl)thiazol-4-yl )methyl)-1H-嘌呤2,6-(3H,7H)-dione

first step

1-[(5-acetyl-2-pyridyl)methyl]-7-(cyclopropylmethyl)-3-methyl-1H-indole-2,6-dione

1-[6-(Bromomethyl)-3-pyridyl]ethanone (100 mg, 0.467 mmol), 7-(cyclopropylmethyl)-3-methyl-1H-indole-2,6-di Ketone (103 mg, 0.467 mmol), potassium iodide (15.5 mg, 0.0934 mmol) and potassium carbonate (193 mg, 1.40 mmol) were dissolved in N,N-dimethylformamide (10 mL). The reaction solution was warmed to 120 ° C and stirred for 3 hours. After cooling to room temperature, filtration, the filtrate was concentrated under reduced pressure and purified by preparative TLC (ethyl acetate, Rf value = 0.5) to give 1-[(5-acetyl-2-pyridyl)methyl]-7-(cyclopropyl) Methyl)-3-methyl-1H-indole-2,6-dione (50.0 mg, yellow solid), yield: 30%. MS-ESI calcd [M+H] ⁺ 355.

Second step

7-(cyclopropylmethyl)-3-methyl-1-((5-methyl-2-(1,1,1-trifluoro-2-hydroxypropan-2-yl)thiazol-4-yl )methyl)-1H-嘌呤 2,6-(3H,7H)-dione

1-[(5-Acetyl-2-pyridyl)methyl]-7-(cyclopropylmethyl)-3-methyl-1H-indole-2,6-dione (100 mg, 0.283 mmol) The cesium fluoride (43.0 mg, 0.283 mmol) was dissolved in tetrahydrofuran (10 mL), and trimethyl-trifluoromethyl-silane (60.4 mg, 0.424 mmol) was added at room temperature and stirred for 12 hours. The reaction was quenched by the addition of water (20 mL). Extract with ethyl acetate (10 mL x 3). The combined organic layers were washed with brine, dried over anhydrous sodium sulfate Purification by preparative high performance liquid chromatography to give 7-(cyclopropylmethyl)-3-methyl-1-((5-methyl-2-(1,1,1-trifluoro-2-hydroxypropane) 2-yl)thiazol-4-yl)methyl)-1H-indole 2,6-(3H,7H)-dione (10.0 mg, yellow solid). ¹ H NMR: (400MHz, Methonal-d ₄ ) δ 8.92 (s, 1H), 8.75 (d, J = 7.6 Hz, 1H), 8.09 (d, J = 7.6 Hz, 1H), 7.97 (s, 1H) ), 5.54 (s, 2H), 4.21 (d, J = 7.6 Hz, 2H), 3.58 (s, 3H), 1.85 (s, 3H), 1.45-1.42 (m, 1H), 0.64-0.59 (m, 2H), 0.50-0.46 (m, 2H). MS-ESI calcd for [M ⁺ H] + 424, found 424.

Example 57

1-((4-Hydroxy-4-(trifluoromethyl)cyclohexyl)methyl)-7-isobutyl-3-methyl-1H-indole-2,6(3H,7H)-dione

first step

1-(1,4-Dioxaspiro[4,5]decane-8-ylmethyl)-7-isobutyl-3-methyl-1H-indole-2,6(3H,7H)- Diketone

1,4-Dioxaspiro[4,5]nonane-8-ylmethyl methanesulfonate (1.07 g, 4.81 mmol), 7-isobutyl-3-methyl-1H-indole-2 6(3H,7H)-dione (1.00g, 4.01mmol) and potassium carbonate (647mg, 4.81mmol) were dissolved in N,N-dimethylformamide (14mL), added potassium iodide (66.5mg, 0.401mmol) The reaction was heated to reflux at 130 ° C for 3 hours. The reaction solution was directly filtered, and the filtrate was concentrated under reduced pressure to give the crude product 1-(1,4-dioxaspiro[4,5]decane-8-ylmethyl)-7-isobutyl-3-methyl- 1H-indole-2,6(3H,7H)-dione (crude 2.56 g, brown oil). MS-ESI calcd for [M+H] ⁺ 377.

Second step

7-Isobutyl-3-methyl-1-((4-oxocyclohexyl)methyl)-1H-indole-2,6(3H,7H)-dione

1-(1,4-Dioxaspiro[4,5]decane-8-ylmethyl)-7-isobutyl-3-methyl-1H-indole-2,6(3H,7H) The diketone (2.50 g, 6.00 mmol) was dissolved in acetone (12 mL) and 4N aqueous hydrochloric acid (2 mL). The reaction was stirred at 30 ° C overnight, and aq. Water (100 mL) was added to the reaction mixture, and the mixture was evaporated. Petroleum ether / ethyl acetate, Rf = 0.3) gave the product 7-isobutyl-3-methyl-1-((4-oxocyclohexyl)methyl)-1H-indole-2,6 (3H,7H) - Diketone (150 mg, white solid), Yield: 13%. ¹ H NMR: (400 MHz, Methonal-d ₄ ) δ 7.94 (s, 1H), 4.14 (d, J = 7.6 Hz, 2H), 3.99 (d, J = 7.6 Hz, 2H), 3.55 (s, 3H) ), 2.29-2.38 (m, 5H), 2.20-2.13 (m, 1H), 2.03-1.98 (m, 2H), 1.53-1.47 (m, 2H), 0.92 (d, J = 6.4 Hz, 6H). MS-ESI calcd [M+H] ⁺ 333.

third step

1-((4-Hydroxy-4-(trifluoromethyl)cyclohexyl)methyl)-7-isobutyl-3-methyl-1H-indole-2,6(3H,7H)-dione

7-Isobutyl-3-methyl-1-((4-oxocyclohexyl)methyl)-1H-indole-2,6(3H,7H)-dione (150 mg, 0.450 mmol) and fluorinated The hydrazine (8.0 mg, 0.0450 mmol) was dissolved in tetrahydrofuran (3 mL) and trifluoromethyltrimethylsilane (950 mg, 0.640 mmol) was slowly added under nitrogen. The reaction was stirred at 30 ° C for 16 hours, 4N aqueous HCl (3 mL) was added and stirred at 25 ° C for half an hour, then aq. The organic phase is dried over anhydrous sodium sulfate and concentrated under reduced pressure to give a crude product which is purified by preparative HPLC to give 1-((4-hydroxy-4-(trifluoromethyl)cyclohexyl)methyl)- 7-Isobutyl-3-methyl-1H-indole-2,6(3H,7H)-dione (86.0 mg, white solid). ¹ H NMR: (400 MHz, Methanol-d ₄ ) δ 7.93 (s, 1H), 4.15-4.04 (m, 2H), 3.89 (d, J = 7.6 Hz, 1H), 3.54 (s, 3H), 2.20 -1.98 (m, 3H), 1.86-1.79 (m, 2H), 1.61-1.42 (m, 5H), 0.92 (d, J = 6.4 Hz, 6H). MS-ESI calcd for [M+H] ⁺ 403.

Example 58

1-((4-Hydroxy-4-(trifluoromethyl)cyclohexyl)methyl)-7-isobutyl-3-methyl-1H-indole-2,6(3H,7H)-dione ( 900 mg, 2.24 mmol), two isomers were isolated by preparative SFC. Isomer 1 separation conditions: column: AD 250 mm x 30 mm, 5 um mobile phase: A: supercritical carbon dioxide, B: ethanol (0.05% aqueous ammonia), A: B = 80:20 flow rate: 50 mL/min Wavelength: 220 nm. Isomer 2 separation conditions: column: WEEK-1300mm x 25mm, 5um mobile phase: A: supercritical carbon dioxide, B: ethanol (0.05% ammonia), A: B = 60: 40, flow rate: 60mL / min, wavelength : 220 nm.

Product 1 (isomer 1, first peak) (216 mg, white solid), yield: 36%. ^{1 H NMR: (400MHz, DMSO} -d 6) δ8.09 (s, 1H), 5.65 (s, 1H), 4.07 (d, J = 7.2Hz, 2H), 3.90 (d, J = 7.2Hz, 2H ), 3.43 (s, 3H), 2.14 - 2.00 (m, 2H), 1.92-1.80 (m, 2H), 1.77-1.66 (m, 2H), 1.52-1.44 (m, 2H), 1.37-1.30 (m , 2H), 0.84 (d, J = 6.4 Hz, 6H). MS-ESI calcd for [M+H] ⁺ 403.

Product 2 (isomer 2, second peak) (101 mg, white solid), yield 37%. ^{1 H NMR: (400MHz, DMSO} -d 6) δ8.07 (s, 1H), 5.64 (s, 1H), 4.05 (d, J = 7.6Hz, 2H), 3.75 (d, J = 7.6Hz, 2H ), 3.42 (s, 3H), 2.16-2.03 (m, 1H), 1.71-1.66 (m, 3H), 1.48-1.30 (m, 6H), 0.83 (d, J = 6.4 Hz, 6H). MS-ESI calcd for [M+H] ⁺ 403.

Example 59

7-(2,3-Dihydroxypropyl)-1-(5-ethyl-5-hydroxyheptyl)-3-methyl-1H-indole-2,6(3H,7H)-dione

first step

7-((2,2-Dimethyl-1,3-dioxolan-4-yl)methyl)-3-methyl-1H-indole-2,6(3H,7H)-dione 3-Methyl-1H-indole-2,6(3H,7H)-dione (200 mg, 1.20 mmol), sodium carbonate (128 mg, 1.20 mmol), 4-(chloromethyl)-2,2-dimethyl The 1,3- dioxolan (217 mg, 1.44 mmol) and potassium iodide (20.0 mg, 0.120 mmol) were dissolved in N,N-dimethylformamide (10 mL). The reaction solution was heated to 110 ° C and reacted for 36 hours. The reaction was quenched with EtOAc (EtOAc) (EtOAc (EtOAc) , Rf = 0.5) gives 7-((2,2-dimethyl-1,3-dioxolan-4-yl)methyl)-3-methyl-1H-indole-2,6 (3H, 7H)-Dione (169 mg, yellow solid), yield: 50%. MS-ESI calcd for [M ⁺ H] + 281, found 281.

Second step

Ethyl 5-(7-((2,2-dimethyl-1,3-dioxolan-4-yl)methyl)-3-methyl-2,6-dioxo-2,3 ,6,7-tetrahydro-1H-decane-1-yl)pentanoic acid ethyl ester

7-((2,2-Dimethyl-1,3-dioxolan-4-yl)methyl)-3-methyl-1H-indole-2,6(3H,7H)-dione (169 mg, 0.604 mmol), ethyl bromopentanoate (178 mg, 0.906 mmol), potassium carbonate (167 mg, 1.21 mmol) and potassium iodide (20.0 mg, 0.0600 mmol) dissolved in N,N-dimethylformamide (10 mL) in. The reaction mixture was heated to 130 ° C, and reacted for 3 hours, filtered, concentrated, and purified by preparative TLC (ethyl acetate, Rf = 0.4) to give ethyl 5-(7-((2,2-dimethyl-1). 3-dioxolan-4-yl)methyl)-3-methyl-2,6-dioxo-2,3,6,7-tetrahydro-1H-decane-1-yl)pentanoic acid Ethyl ester (124 mg, yellow solid), yield: 50%. MS-ESI calcd for [M+H] ⁺ 409.

third step

7-((2,2-Dimethyl-1,3-dioxolan-4-yl)methyl)-1-(5-ethyl-5-hydroxyheptyl)-3-methyl-1H -嘌呤-2,6(3H,7H)dione

Ethyl 5-(7-((2,2-dimethyl-1,3-dioxolan-4-yl)methyl)-3-methyl-2,6-dioxo-2, 3,6,7-Tetrahydro-1H-decane-1-yl)pentanoic acid ethyl ester (30.0 mg, 0.0750 mmol) was dissolved in anhydrous tetrahydrofuran (1 mL) and slowly added dropwise at -65 °C. Magnesium bromide (3M in tetrahydrofuran, 0.15 mL, 0.450 mmol). The reaction was allowed to react at -65 ° C for 0.5 hours and then at 0 ° C for 0.5 hour. The reaction mixture was poured into water (5 mL) EtOAc (EtOAc m. To give 7-((2,2-dimethyl-1,3-dioxolan-4-yl)methyl)-1-(5-ethyl-5-hydroxyheptyl)-3-methyl -1H-indole-2,6(3H,7H)dione (20.0 mg, yellow oil), yield: 63%. MS-ESI calcd for [M ⁺ H] + 423, found 423.

the fourth step

7-(2,3-Dihydroxypropyl)-1-(5-ethyl-5-hydroxyheptyl)-3-methyl-1H-indole-2,6(3H,7H)-dione

7-((2,2-Dimethyl-1,3-dioxolan-4-yl)methyl)-1-(5-ethyl-5-hydroxyheptyl)-3-methyl- 1H-indole-2,6(3H,7H)dione (20.0 mg, 0.0470 mmol) was dissolved in anhydrous tetrahydrofuran (1 mL) and diluted hydrochloric acid (0.3 mL), and the mixture was reacted at 25 ° C for 36 hours. After the reaction was completed, it was concentrated under reduced pressure and purified by preparative TLC plate (8:1 ethyl acetate / methanol, Rf = 0.3) to give 7-(2,3-dihydroxypropyl)-1-(5-ethyl) -5-Hydroxyheptyl)-3-methyl-1H-indole-2,6(3H,7H)-dione (5.0 mg, white solid). ^{1 H NMR: (400MHz, Methonal} -d 4) δ7.90 (s, 1H), 4.57-4.53 (m, 1H), 4.26-4.22 (m, 1H), 4.03-3.96 (m, 3H), 3.58- 3.55 (m, 2H), 3.54 (s, 3H), 1.66-1.61 (m, 2H), 1.48-1.44 (m, 6H), 1.34-1.29 (m, 2H), 0.85 (t, J = 8.0 Hz, 6H). MS-ESI calcd for [M+H] ⁺ 383.

Example 60

1-(5-ethyl-5-hydroxyheptyl)-7-(2-hydroxyethyl)-3-methyl-1H-indole-2,6(3H,7H)-dione

first step

7-(2-hydroxyethyl)-3-methyl-1H-indole-2,6(3H,7H)-dione

3-Methyl-1H-indole-2,6(3H,7H)-dione (1.00 g, 6.00 mmol), potassium carbonate (830 mg, 6.00 mmol) was dissolved in N,N-dimethylformamide (10 mL) )in. The reaction solution was heated to 80 ° C for 0.5 hour, and 2-bromoethanol (900 mg, 7.20 mmol) was added. The reaction solution was heated to 130 ° C and allowed to react overnight. The reaction mixture was concentrated to give crude 7-(2-hydroxyethyl)-3-methyl-1H-indole-2, 6(3H,7H)-dione as directly.

Second step

Ethyl 5-(7-(2-hydroxyethyl)-3-methyl-2,6-dioxo-2,3,6,7-tetrahydro-1H-indol-1-yl)pentanoic acid ester

7-(2-Hydroxyethyl)-3-methyl-1H-indole-2,6(3H,7H)-dione (1.05 g, 5.00 mmol), ethyl 5-bromopentanoate (1.25 g, 6.00 Methyl acetate and potassium carbonate (1.66 g, 12.0 mmol) were dissolved in N,N-dimethylformamide (3 mL). The reaction solution was heated to 130 ° C for 3 hours. The reaction mixture was poured into water (20 mL). The organic phase was combined, dried over anhydrous sodium sulfate, filtered, evaporated, evaporated, evaporated, evaporated,,,,,,,,,,,,,,,,,,,,, Ethyl-2,3,6,7-tetrahydro-1H-indol-1-yl)pentanoate (700 mg, yellow oil), yield: 35%. MS-ESI calcd for [M ⁺ H] + 339, found 339.

third step

1-(5-ethyl-5-hydroxyheptyl)-7-(2-hydroxyethyl)-3-methyl-1H-indole-2,6(3H,7H)-dione

Ethyl 5-(7-(2-hydroxyethyl)-3-methyl-2,6-dioxo-2,3,6,7-tetrahydro-1H-indol-1-yl)pentanoic acid Ethyl acetate (700 mg, 2.07 mmol) was dissolved in anhydrous tetrahydrofuran (7 mL), and ethyl acetate (3M tetrahydrofuran solution, 7 mL, 2.10 mmol) was slowly added dropwise at -78 °C. The reaction solution was reacted at -78 ° C for 1 hour. The reaction mixture was poured into water (20 mL) and evaporated, The organic phase was dried over anhydrous sodium sulfate, filtered, evaporated, evaporated, evaporated,,,,,,,,,,,,,,,,,,,,,,, Methyl-1H-indole-2,6(3H,7H)-dione (110 mg, white solid), yield: 15%. ^{1 H NMR: (400MHz, Methonal} -d 4): δ7.90 (s, 1H), 4.41 (t, J = 5.0Hz, 2H), 4.00 (t, J = 7.6Hz, 2H), 3.87 (t, J=5.0 Hz, 2H), 3.54 (s, 3H), 1.68-1.59 (m, 2H), 1.50-1.42 (m, 5H), 1.39-1.29 (m, 3H), 0.95-0.77 (m, 6H) . MS-ESI calcd for ^{_{[M + HH 2 O] +}} 335, found 335.

Example 61

1-(5-ethyl-5-hydroxyheptanol)-7-(2-hydroxy-3-((2-hydroxyethyl)(methyl)amino)propyl)-3-methyl-1H-indole -2,6(3H,7H)-dione

first step

1-bromo-3-((2-hydroxyethyl)(methyl)amino)propan-2-ol

2-(Methylamino)ethanol (135 mg, 1.80 mmol) was dissolved in N,N-dimethylformamide (5 mL), and 2-(bromomethyl)oxirane (206 mg, 1.51 mmol) The reaction was carried out for 1.5 hours. After the reaction was completed, the reaction solution was used directly for the next reaction.

Second step

7-(2-Hydroxy-3-((2-hydroxyethyl)(methyl)amino)propyl)-3-methyl-1H-indole-2,6(3H,7H)-dione to 1- To a solution of bromo-3-((2-hydroxyethyl)(methyl)amino)propan-2-ol, 3-methyl-1H-indole-2,6(3H,7H)-dione (100 mg, 0.602 mmol), sodium carbonate (64.0 mg, 0.602 mmol) and potassium iodide (10.0 mg, 0.0600 mmol). The reaction solution was heated to 80 ° C and allowed to react for 10 hours. After the reaction was completed, it was filtered and concentrated under reduced pressure to give crude 7-(2-hydroxy-3-((2-hydroxyethyl)(methyl)amino)propyl)-3-methyl-1H-indole-2. 6(3H,7H)-dione was used directly in the next step. MS-ESI calcd [M+H] ⁺ 298.

third step

Ethyl 5-(7-(2-hydroxy-3-((2-hydroxyethyl))(methyl)amino)propyl)-3-methyl-2,6-dioxo-2,3,6 ,7-tetrahydro-1H-decane-1-yl)pentanoic acid ethyl ester

7-(2-Hydroxy-3-((2-hydroxyethyl)(methyl)amino)propyl)-3-methyl-1H-indole-2,6(3H,7H)-dione (170mg , 0.572 mmol), ethyl bromopentanoate (169 mg, 0.858 mmol), potassium carbonate (158 mg, 1.14 mmol) and potassium iodide (10.0 mg, 0.0570 mmol) were dissolved in N,N-dimethylformamide (5 mL). The reaction solution was heated to 130 ° C and reacted for 3 hours. The reaction mixture was poured into water (5 mL) and evaporated and evaporated. The organic phase was combined, dried over anhydrous sodium sulfate, filtered, evaporated, evaporated, evaporated, ((2-hydroxyethyl)(methyl)amino)propyl)-3-methyl-2,6-dioxo-2,3,6,7-tetrahydro-1H-decane-1-yl Ethyl valerate (118 mg, yellow oil), yield: 49%. MS-ESI calcd for [M ⁺ H] + 426, found 426.

the fourth step

1-(5-ethyl-5-hydroxyheptanol)-7-(2-hydroxy-3-((2-hydroxyethyl)(methyl)amino)propyl)-3-methyl-1H-indole -2,6(3H,7H)-dione

Ethyl 5-(7-(2-hydroxy-3-((2-hydroxyethyl))(methyl)amino)propyl)-3-methyl-2,6-dioxo-2,3, Ethyl 6,7-tetrahydro-1H-decane-1-yl)pentanoate (100 mg, 0.235 mmol) was dissolved in anhydrous tetrahydrofuran (4 mL), and then slowly dropped into ethyl bromide at -65 °C. Magnesium (3M in tetrahydrofuran, 0.47 mL, 1.41 mmol). The reaction was reacted at -65 ° C for 0.5 hour and then reacted at 0 ° C for 0.5 hours. After the reaction was completed, the reaction mixture was evaporated, evaporated, mjjjjjjjjjjjjjjjj Purification of ethyl acetate/methanol, Rf = 0.3) to give 1-(5-ethyl-5-hydroxyheptanol)-7-(2-hydroxy-3-((2-hydroxyethyl)(methyl)amino) Propyl)-3-methyl-1H-indole-2,6(3H,7H)-dione (3.0 mg, white solid). ^{1 H NMR: (400MHz, Methonal} -d 4) δ7.92 (s, 1H), 4.91-4.59 (m, 2H), 4.52-4.39 (m, 3H), 4.30-4.25 (m, 2H), 4.03- 3.99 (m, 2H), 3.88-3.84 (m, 2H), 3.55 (s, 3H), 2.90 (s, 3H), 1.68-1.59 (m, 4H), 1.48-1.45 (m, 6H), 0.88- 0.84 (m, 6H). MS-ESI calcd for [M+H] ⁺ 440.

Experimental Example 1: Evaluation of PDE2 phosphodiesterase inhibitory activity in vitro

EXPERIMENTAL OBJECTIVE: To detect the AMP/GMP concentration produced in the reaction system by detecting the AlexaFluor 633 fluorescent dye substituted on the AMP/GMP antibody by fluorescence polarization analysis, and calculate the IC ₅₀ value of the PDE2 phosphodiesterase inhibition of the test compound.

Experimental Materials:

Determination of buffer solution: 10 mM Tris buffer, pH 7.5, 5 mM magnesium chloride, 0.01% polyoxyethylene lauryl ether, 1 mM dithiothreitol and 1% DMSO.

Enzyme: Expression of recombinant full-length human PDE2A protein in insect Sf9 cells using baculovirus using N-terminal GST tag

Substrate: 1μM cGMP

Detection method:

AMP²/GMP²抗体，AMP2/GMP2AlexaFluor 633荧光染料

AMP ² / GMP ² antibody, AMP2 / GMP2AlexaFluor 633 fluorescent dye

Experimental operation:

AMP²/GMP²抗体，AMP2/GMP2AlexaFluor 633荧光染料)，室温下反应90分钟，然后使用Ex/Em 620/688检测荧光偏振。The freshly prepared buffer solution was placed in the reaction solution, and then added to the reaction well, and the DMSO solution of the test compound was added through an Echo 550 non-contact nano-sound sonic pipetting system, and then pre-incubated for 10 minutes at room temperature, and the substrate was added (1 μM). The reaction was initiated by cGMP) and allowed to react at room temperature for one hour. Then join the detection system (

AMP ² /GMP ² antibody, AMP2/GMP2 AlexaFluor 633 fluorescent dye), reacted at room temperature for 90 minutes, and then detected fluorescence polarization using Ex/Em 620/688.

Fluorescence polarization intensity was converted to nM concentration by AMP/GMP standard curve, then relative enzyme activity inhibition relative to DMSO blank was calculated, and IC50 values and curves were calculated using the Prism software package (GraphPad Software, San Diego California, USA).

Experimental results:

Table 1 PDE2 phosphodiesterase inhibitory activity test results

Test article (compound prepared in each example) PDE2 phosphodiesterase inhibitory activity Example 1 + Example 2 ++ Example 3 + Example 4 ++ Example 5 + Example 6 -- Example 7 -- Example 8 Isomer 1 / Isomer 2 ++/+ Example 9 Isomer 1 / Isomer 2 +/+ Example 10 Isomer 1 / Isomer 2 --/-- Example 11 + Example 12 Isomer 1 / Isomer 2 +/++ Example 13 -- Example 14 -- Example 15 Isomer 1 / Isomer 2 ++/++ Example 16 -- Example 17 -- Example 18 ++ Example 19 + Example 20 + Example 21 + Example 22 ++ Example 23 Isomer 1 / Isomer 2 --/-- Example 24 -- Example 25 ++ Example 26 + Example 27 ++

Example 28 ++ Example 29 -- Example 30 -- Example 31 + Example 32 -- Example 33 + Example 34 + Example 35 + Example 36 + Example 37 -- Example 38 -- Example 39 -- Example 40 -- Example 41 Isomer 1 / Isomer 2 --/-- Example 42 ++ Example 43 + Example 44 ++ Example 45/45' --/++ Example 46 Isomer 1 / Isomer 2 --/+ Example 47 + Example 48 + Example 49 ++ Example 50 Isomer 1 / Isomer 2 +/+++ Example 51 Isomer 1 / Isomer 2 +/++ Example 52 + Example 53 + Example 54 ++ Example 55 +++ Example 56 + Example 57 ++

Example 58 Isomer 1 / Isomer 2 --/-- Example 59 -- Example 60 -- Example 61 --

Note: 10 μM ≤ "+" < 50 μM; 1 μM ≤ "++" < 10 μM; "+++" < 1 μM; - N/A Conclusion: The compounds of the present invention have significant and unexpected PDE2A protease inhibitory activity.

Experimental Example 2: Evaluation of the effect of compounds on the TNF-α in the blood of rats induced by LPS in vitro

OBJECTIVE: To examine the effects of compounds on the TNF-α in the blood of rats induced by LPS in vitro, and to evaluate the inhibitory effect of the compounds on LPS-induced TNF-α in rat blood.

Experimental Materials:

Sprague Dawley rats (male, 210-260 g, 8-10 weeks old, Shanghai Slack)

Rat TNF-alpha Quantikine ELISA Kit (R&D, #SRTA00)

Experimental operation:

A test compound solution having a concentration of 1 mM was prepared, and 40 μL (the final concentration of the compound was 100 uM) was added to a 48-well cell culture plate, respectively. After the rats were anesthetized with isoflurane, blood was collected from the heart (heparin anticoagulation). Blood was added to a 48-well plate to which the test compound had been added, 320 μL per well. The 48-well plate was placed in a cell culture incubator, and after 30 minutes of incubation, it was taken out, 40 μL of LPS solution (100 ug/mL) was added, mixed, and placed in an incubator to continue incubation. After 5 hours, the 48-well plate was taken out, and the blood sample was transferred to a 1.5 mL centrifuge tube, centrifuged in a centrifuge (4,500 rpm, 4 ° C, 5 minutes), the upper layer was separated, and the plasma was separated, frozen, and stored in a -80 degree refrigerator. . The next day, according to the kit instructions, the R&D ELISA kit was used to detect TNF-α levels in plasma samples.

Experimental results:

Table 2 TNF-α inhibitory activity test results

Test article (compound prepared in each example) TNF-α inhibition ratio Example 1 + Example 11 + Example 12 Isomer 1 / Isomer 2 +/+ Example 27 + Example 40 + Example 44 ++ Example 49 ++ Example 50 Isomer 1 / Isomer 2 --/++ Example 51 Isomer 1 / Isomer 2 --/++ Example 55 ++

Note: 60% ≤ "+" <80%; 80% ≤ "++" <100%; - N/A

Conclusion: The compounds of the invention have significant and unexpected TNF-α inhibitory activity.

Experimental Example 3: Evaluation of Compound Pharmacokinetics

Objective: To test the pharmacokinetics of compounds in SD rats

Experimental Materials:

Sprague Dawley rats (male, 200-300 g, 7-9 weeks old, Shanghai Slack)

Experimental operation:

The rodent pharmacological characteristics of the compound after intravenous injection and oral administration were tested by a standard protocol. In the experiment, the candidate compound was formulated into a clear solution, and the rats were administered a single intravenous injection and oral administration. The intravenous and oral vehicles are a certain proportion of aqueous hydroxypropyl β-cyclodextrin or physiological saline solution. Collect whole blood samples within 24 hours, centrifuge at 3000g for 15 minutes, separate the supernatant to obtain plasma samples, add 4 times volume of acetonitrile solution containing internal standard to precipitate protein, centrifuge to remove the supernatant, add equal volume of water and centrifuge to remove the supernatant. The LC-MS/MS analysis method was used to quantitatively analyze the plasma concentration, and the pharmacokinetic parameters such as peak concentration, peak time, clearance rate, half-life, area under the curve of the drug, and bioavailability were calculated.

Experimental results:

Table 3 pharmacokinetic test results

Conclusion: The compounds of the present invention can significantly increase the single or partial index of rat pharmacokinetics.

Experimental Example 4: In vivo pharmacology study of MCD mouse model

main mission:

Evaluation of Compound 1 for nonalcoholic steatohepatitis model MCD mice liver histology, TG levels, TNF-α, mRNA levels and inflammatory factors (IL-1β levels; plasma ALT/AST levels, inflammatory factors (TNF-α, IL) -1β) level of influence.

The problem to be solved:

Test compound 1 on liver histology, TG level, TNF-α mRNA level and inflammatory factor (IL-1β) level in MCD mice; plasma ALT/AST level, inflammatory factor (TNF-α, IL-1β) level, evaluation Compounds protect liver function in mice with non-alcoholic steatohepatitis and reduce liver inflammation.

Experimental model: a model of nonalcoholic steatohepatitis induced by diet in mice with methionine and choline deficiency

(Methionine-choline-deficient diet treated mice, MCD mice)

Experimental animals: C57BL/6J mice (male, purchased at 4 weeks old, Shanghai Slack)

Experimental reagents:

1.MCD feed (Research Diets, #A02082002B)

2. MCD control feed (Research Diets, #A02082003B)

Mini RNA提取试剂盒(Qiagen，#74104)3.

Mini RNA Extraction Kit (Qiagen, #74104)

4.High Capacity cDNA Reverse Transcription Kit (ABI, #4368814)

Gene Expression Assay TNF-α(ABI，Assay ID：Mm00443258_m1)5.

Gene Expression Assay TNF-α (ABI, Assay ID: Mm00443258_m1)

Endogenous Control 18S(ABI，Assay ID：Mm03928990_g1)6.

Endogenous Control 18S (ABI, Assay ID: Mm03928990_g1)

Fast Universal PCR Mater Mix(ABI，#4366072)7.

Fast Universal PCR Mater Mix (ABI, #4366072)

8.One cOmplete, EDTA-free protease inhibitor tablet (Roche, #11873580001)

9.BCA Protein Quantification Kit (Pierce, #23227)

10.MSD inflammatory factor test kit (Meso Scale Discovery, #K15A0H-2)

11.Triglyceride Quantification Kit (BioVision, #K622-100)

12.Hematoxylin (Baso, #BA4097)

13.Eosin(Baso,#BA4099)

experimental method:

1. Experimental procedure

2. In vivo experimental methods

Four-week-old C57BL/6J mice were purchased and special feeds were started after 4 weeks of adaptation to the environment. Pre-adaptation of oral administration was started 1 week after the special feed was raised. Preconditioning method: 0.5% (w/v) hydroxypropyl methylcellulose according to 5 mL/kg oral vehicle control. After a week of preconditioning, randomization was performed and oral administration was performed. The grouping and administration information is as follows:

Two weeks after the administration, the mice were sacrificed by CO ₂ , and the hearts were bled 500 μL to 600 μL (K ₂ -EDTA anticoagulation). The blood samples were centrifuged in a centrifuge (4,500 rpm, 4 ° C, 5 minutes), and the upper layer was separated for plasma. After sub-packaging, freeze it and store it in a -80 °C refrigerator. Liver tissue was dissected for histological analysis and biomarker detection.

Frozen plasma samples were tested for ALT and AST the following day.

3. Histological analysis method

3.1HE staining method

3.1.1 Organizational Processing

Using the Leica ASP200S automatic dewatering machine, the relevant dewatering steps are as follows:

a) 10% neutral buffer formalin 5 min - 6 h

b) 70% ethanol 45 minutes

c) 80% ethanol 45 minutes

d) 95% ethanol 45 minutes

e) 95% ethanol 45 minutes

f) 100% ethanol 60 minutes

g) 100% ethanol 60 minutes

h) 100% ethanol 60 minutes

i) xylene 60 minutes

j) xylene 60 minutes

k) paraffin wax 60 minutes

l) Paraffin wax 60 minutes

m) paraffin wax 60 minutes

3.1.2 Organizational embedding

a) Preparation before embedding

The composition and function of the Leica tissue embedding system

The Leica tissue embedding system consists of two parts: the embedding table (Leica EG1150H) and the freezing station (Leica EG1150C). The embedding station includes a left metal mold storage cylinder, a right side tissue cassette storage cylinder and a paraffin bath on the upper part of the instrument. There is a small cold stage at the front of the embedding station that can quickly cool a single embedding mold. The freezing station is mainly composed of a cold stage, and rapidly cools the paraffin embedded in the mold. After the cassette is cooled, the wax block is stored at room temperature.

b) Turn on the cold station.

c) Take out the cassette in the dehydrator and place it in the wax bath of the embedding machine.

d) embedded

The hot tweezers are taken out of the tissue cassette in the embedding machine wax bath and placed on the hot stage of the embedding machine. Place the embedding box corresponding to the tissue on the mold. The tissue is placed in the embedding mold with hot tweezers, and a little dissolved paraffin is added to the embedding mold, and the embedding mold is placed on the cold table until the paraffin becomes completely solid. Finally, the solid wax block is taken out of the embedding mold.

3.1.3 Preparation of tissue sections

a) Confirm the status before using the Leica RM2235 paraffin slicer

Before loading the wax block, make sure that the slice wheel handle is loosely locked, open the knife holder cover, and clean the wax on the knife holder.

Carefully insert an unused disposable knife to confirm that the blade is in place at home and lock the knife holder according to the manufacturer's instructions. Confirm the inclination of the blade, which should be set to 3-6 degrees.

b) Fix the tissue wax block to the desired position in the clipper slot.

Follow the manufacturer's instructions for roughing with the roughing wheel handles so that the wax block slowly advances until the desired tissue plane is exposed.

c) Start slicing, first set the slice thickness, start slicing, and rotate the slice runner forward until the flat wax sheet is formed.

Lock the slice roller handle and use a forceps or brush to move the tissue piece into the spreader (Leica HI1210) and flatten it onto the slide.

d) After roughing or slicing, lock the wheel handle and carefully remove the wax block.

3.1.4 HE staining

HE staining was performed using a fully automated staining workstation consisting of Leica ST5010, Leica TS5010 and Leica CV5030.

Press the <Stain> key in the main menu of the dyeing machine. Conventional HE staining is performed by the up and down arrow selection procedure 15 of the button area. Open the load compartment, place the slicer, and close the load compartment. Start the dye program after pressing the <Load> button. The procedure is as follows:

a) Oven 55 ° C 2 minutes

b) xylene I 10 minutes

c) xylene II 10 minutes

d) xylene III 10 minutes

e) absolute ethanol I 2 minutes

f) absolute ethanol II 1.5 minutes

g) 95% ethanol I 1 minute

h) 80% ethanol I 0.5 min

i) Washing for 3 minutes

j) Hematoxylin dye solution 3 minutes

k) Washing for 2 minutes

l) 1% hydrochloric acid-ethanol 5 seconds

m) Washing for 2 minutes

n) Washing for 20 minutes

o) 80% ethanol II 1 minute

p) Yihong 5 seconds

q) 95% ethanol II 30 seconds

r) absolute ethanol III 2 minutes

s) Anhydrous ethanol IV 2 minutes

t) absolute ethanol V 2 minutes

u) Anhydrous ethanol VI 2 minutes

v) xylene IV 3 minutes

w) xylene V 3 minutes

3.2 Histological non-alcoholic fatty liver disease activity score (NAS) related scoring criteria:

The histological diagnosis and clinical efficacy evaluation of nonalcoholic fatty liver disease (NAFLD) was performed in accordance with the NASH Clinical Research Network Pathology Working Group Guidelines of the National Institutes of Health, and the NAFLD activity score (NAS) was routinely performed.

NAS score (0 to 8 points):

a) Hepatocyte steatosis: 0 points (<5%); 1 point (5% to 33%); 2 points (33% to 66%); 3 points (>66%).

b) Intralobular inflammation (20-fold microscopic count necrosis): 0 points, none; 1 point (<2); 2 points (2 to 4); 3 points (> 4).

c) Hepatocyte balloon-like changes: 0 points, no; 1 point, rare; 2 points, more common.

4. Liver TG detection method

4.1 Sample preparation: 5% NP40 lysate was added to each liver tissue to a concentration of 100 mg/mL. The tissue was lysed on a tissue homogenizer. The sample was heated in an 80-100 degree water bath for 5 min and then cooled to room temperature. Repeat the heating once. Centrifuge at maximum speed for 2 min and take the supernatant. The supernatant was diluted 60-fold and 30 μL was added to each well of the assay plate.

4.2 Standard preparation: 1 mM triglyceride was diluted to 0.2 mM, and 0, 10, 20, 30, 40, 50 μL of 0.2 mM triglyceride standard was added to the standard well column, and the test solution was added to 50 μL.

4.3 Add 2 μL of lipase to each well and mix for 20 min at room temperature.

4.4 Proportionally arranged enzyme reaction solution: 46 μL of triglyceride detection solution + 2 μL of triglyceride probe + 2 μL of triglyceride reaction solution.

Add 50 μL of enzyme reaction solution to all wells, mix and incubate for 30-60 min at room temperature.

4.5 Detection: The absorbance at 570 nm was measured with a microplate reader.

5. Method for detecting TNF-α mRNA in liver tissue

Mini RNA提取试剂盒(Qiagen#74104)提取RNA，当天完成逆转录，逆转录体系配比(ABI#4368814)如下：According to the operation manual, use

RNA was extracted from Mini RNA extraction kit (Qiagen #74104), and reverse transcription was completed on the same day. The reverse transcription system ratio (ABI#4368814) was as follows:

Reverse transcription program:

temperature duration 25°C 10min 37 ° C 120min 85 ° C 5min 4 ° C ∞

The cDNA was stored in a refrigerator at -20 °C.

Fast Universal试剂盒(ABI，#4366072)说明书，进行Q-PCR检测，体系配比如下：according to

Fast Universal kit (ABI, #4366072) instructions for Q-PCR detection, the system is equipped with the following:

Q-PCR program:

6. Inflammatory factor detection method

6.1 Liver tissue homogenate and sample preparation

6.1.1 start pre-cooling centrifuge;

6.1.2 A piece of One cOmplete, EDTA-free protease inhibitor tablet was dissolved in 50 mL of phosphate buffer, and PBS was placed in wet ice for 1:10 (w/v).

6.1.3 Centrifuge 1000g, 10min, 4°C, transfer 220μL of the supernatant to the ultracentrifuge tube, and prepare two tubes for ultracentrifugation in each supernatant.

6.1.4 has a pre-cooled sample is placed in an ultracentrifuge Optima ^{TM L-80XP} (Beckman), ultracentrifugation 100,000g, 4 ℃, 50min.

6.1.5 Pre-cool the V-bottom 96-well plate on dry ice, and combine the two tube supernatants corresponding to the same sample into a 1.5 mL EP tube. After mixing with light bomb, take 120 μL and dispense into a 96-well plate at the bottom of the V. Dry ice is frozen. A total of three sample plates were prepared (one plate for BCA protein quantification, one plate for MSD detection, and one remaining spare).

Determination of protein concentration in tissue homogenate by 6.2BCA method

6.2.1 Preparation of BCA working solution: Reagents A and B were mixed according to reagent A: reagent B=50:1.

6.2.2 Preparation of BSA (bovine serum white egg) standard solution A ~ I:

6.2.3 Dilute the homogenate with PBS, 20 μL homogenate + 80 μL PBS.

6.2.4 Add 10 μL of BSA standard solutions A to I and diluted samples to be tested in a 96-well plate.

6.2.5 Add 200 μL of BCA working solution to each well to the standard solution and the sample to be tested, and mix on the platen;

6.2.6 Incubate for 30 minutes in a 37 ° C incubator;

6.2.7 remove the plate and let it cool to room temperature;

6.2.8 Measure the absorbance at 562 nm on a microplate reader.

6.3 MSD detection of inflammatory factors in samples

6.3.1 Remove the MSD plate from the refrigerator and store at room temperature.

6.3.2 Dilute the standard solution according to a 1:3 gradient.

6.3.3 Add 50 μL of standard and sample solution to each well, fix on a vibrating plate, and incubate at 300-1000 rpm for 2 hr at room temperature.

6.3.4 Add 300 μL Wash buffer to each well and wash the plate three times. Add 25 μL of 1× detection antibody solution to each well, fix it on the vibrating plate, and incubate at 300-1000 rpm for 2 hr at room temperature.

6.3.5 Add 300 μL Wash buffer to each well and wash the plate three times. Add 150 μL of 1× reading solution to each well and insert the signal value into the Sector Imager 6000 Model 1200.

Experimental results:

1. Histological results

1.1HE staining picture

The results of HE staining showed that the liver fat of the mice increased significantly and the inflammation of the hepatic lobule increased significantly.

Two doses of pentoxifylline (200mpk, po., tid) and INT-747 (30mpk, po., qd) were administered for two weeks. There was no significant improvement in liver lesions in MCD mice, while Compound 1 (30mpk, po., bid) The hepatic lobular inflammation of MCD mice was significantly improved after two weeks of administration. The experimental results are shown in Figure 1.

1.2NAS score

Liver histology of NAS and lobular inflammation in MCD mice was significantly increased. The three compounds tested in this experiment had no significant effect on NAS, but compound 1 significantly reduced liver lobular inflammation in MCD mice. The experimental results are shown in Figure 2.

2. Liver TG test results

The liver TG level of MCD mice was significantly increased. The three compounds tested in this experiment had no significant effect on the liver TG level of MCD mice. The experimental results are shown in Fig. 3.

3. Liver TNF-α mRNA level test results

Compound 1 (30mpk, po., b.i.d.) had a downward trend in liver TNF-α mRNA levels in MCD mice for two weeks, but there was no statistically significant difference. The experimental results are shown in Figure 4.

4. Liver inflammatory factor test results

After 4 weeks of MCD feeding, the level of IL-1β protein in the liver of mice was significantly increased, and pentoxifylline (200mpk, po., tid) and INT-747 (30mpk, po., qd) were administered for two weeks to the liver of MCD mice. The level of IL-1β protein was not significantly affected; the level of IL-1β protein in liver of MCD mice was significantly decreased in compound 1 (30 mpk, po., bid) for two weeks. The experimental results are shown in Fig. 5.

5. Plasma inflammatory factor test results

MCD mice plasma TNF-α and IL-1β protein levels were significantly increased, pentoxifylline (200mpk, po., tid) and INT-747 (30mpk, po., qd) for two weeks on MCD mouse plasma There was no significant effect of TNF-α and IL-1β protein levels. The levels of plasma TNF-α and IL-1β protein in MCD mice were significantly decreased in compound 1 (30 mpk, po., bid) for two weeks. The experimental results are shown in Fig. 6.

6. Plasma ALT and AST test results

Plasma ALT and AST levels were significantly increased in MCD mice. The three compounds tested in this experiment could not reduce plasma ALT and AST levels in MCD mice. The experimental results are shown in Figure 7.

to sum up:

1. MCD mice are widely used animal models of non-alcoholic steatohepatitis. The model mice have obvious liver steatosis and TG levels are significantly increased. Histological HE staining pictures show obvious lobular inflammation, and liver TNF-α mRNA is significantly elevated. High, liver inflammatory factors IL-1β, plasma TNF-α and IL-1β protein levels were significantly higher than normal; plasma ALT and AST levels were significantly higher than normal mice.

2. Compound 1 (30mpk, po., b.i.d.) After two weeks of administration, liver lobular inflammation was significantly improved in MCD mice, and liver inflammatory factors IL-1β, plasma TNF-α and IL-1β protein levels were significantly decreased.

3. Compound 1 (30mpk, po., b.i.d.) was administered for two weeks, and had no significant effect on liver TG level, liver TNF-α mRNA, plasma ALT and AST levels in MCD mice.

Experimental Example 5 In vivo pharmacology study of nSTZ+HFD mouse model

main mission:

The effects of Compound 1 on liver histology, TNF-α mRNA, TGF-β1 mRNA and collagen 1α1 mRNA levels, serum ALT, AST, TG and TC levels in non-alcoholic steatohepatitis model nSTZ+HFD mice were evaluated.

The problem to be solved:

The effects of test compound 1 on liver histology, TNF-α mRNA, TGF-β1 mRNA and Collagen 1α1 mRNA levels, serum ALT, AST, TG and TC levels in nSTZ+HFD mice were evaluated for compounds in nonalcoholic steatohepatitis model mice. Liver function protection reduces the effects of liver inflammation and fibrosis.

Experimental model: STZ injection of neonatal mice combined with high-fat diet induced non-alcoholic steatohepatitis model

(nSTZ+HFD mice)

Experimental animals: C57BL/6J mice (female pregnant mice, purchased 14-15 days pregnant, Shanghai Lingchang Biotechnology Co., Ltd.)

Experimental reagents:

14. Streptozotocin (STZ) (Sigma, #S0130-1G)

15. High-fat diet (Research Diets, #D12492i)

Mini RNA提取试剂盒(Qiagen，#74104)16.

Mini RNA Extraction Kit (Qiagen, #74104)

17.High Capacity cDNA Reverse Transcription Kit (ABI, #4368814)

Gene Expression Assay TNF-α(ABI，Assay ID：Mm00443258_m1)18.

Gene Expression Assay TNF-α (ABI, Assay ID: Mm00443258_m1)

Gene Expression Assay TGF-β1(ABI，Assay ID：Mm00441724_m1)19.

Gene Expression Assay TGF-β1 (ABI, Assay ID: Mm00441724_m1)

Gene Expression Assay Collagen 1α1(ABI，Assay ID：Mm00801666_g1)20.

Gene Expression Assay Collagen 1α1 (ABI, Assay ID: Mm00801666_g1)

Endogenous Control 18S(ABI，Assay ID：Mm03928990_g1)twenty one.

Endogenous Control 18S (ABI, Assay ID: Mm03928990_g1)

Fast Universal PCR Mater Mix(ABI，#4366072)twenty two.

Fast Universal PCR Mater Mix (ABI, #4366072)

23.Hematoxylin (Baso, #BA4097)

24.Eosin(Baso,#BA4099)

25.Sirius red (Bogoo, #PT036)

experimental method:

1. Experimental procedure

2. In vivo experimental methods

14 to 15 days of gestational age C57BL/6J mice were purchased, and newborn rats were born for 2 to 3 days for STZ injection (200 ug, s.c.). Oral diets are raised until 4 weeks of age to start a high-fat diet. Animals were randomized after 1 week of high fat feeding and started oral administration.

The grouping and drug administration information is as follows:

Fasting was continued overnight after 5 weeks of continuous administration. The mice were sacrificed by CO ₂ , and the hearts were taken for 2 hours at room temperature. The blood samples were centrifuged (2000 g, 4 ° C, 15 minutes) in a centrifuge, and the supernatant was separated, frozen, and stored in a -80 ° C refrigerator. Liver tissue was dissected for histological analysis and biomarker detection.

Frozen serum samples were tested for ALT, AST, TG and TC the following day.

3. Histological analysis method

3.1HE staining method

3.1.1 Organizational Processing

Using the LeicaASP200S automatic dewatering machine, the relevant dewatering steps are as follows:

n) 10% neutral buffered formalin for 5 minutes to 6 hours;

o) 70% ethanol 45 minutes

p) 80% ethanol 45 minutes

q) 95% ethanol 45 minutes

r) 95% ethanol 45 minutes

s) 100% ethanol 60 minutes

t) 100% ethanol 60 minutes

u) 100% ethanol 60 minutes

v) xylene 60 minutes

w) xylene 60 minutes

x) Paraffin wax 60 minutes

y) paraffin wax 60 minutes

z) Paraffin wax 60 minutes

3.1.2 Organizational embedding

e) Preparation before embedding

The composition and function of the Leica tissue embedding system

The Leica tissue embedding system consists of two parts: the embedding table (Leica EG1150H) and the freezing station (Leica EG1150C). The embedding station includes a left metal mold storage cylinder, a right side tissue cassette storage cylinder and a paraffin bath on the upper part of the instrument. There is a small cold stage at the front of the embedding station that can quickly cool a single embedding mold. The freezing station is mainly composed of a cold stage, and rapidly cools the paraffin embedded in the mold. After the cassette is cooled, the wax block is stored at room temperature.

f) Open the cold station

g) Take out the embedding box in the dehydrator and put it into the embedding machine wax bath

h) embedded

The hot tweezers are taken out of the tissue cassette in the embedding machine wax bath and placed on the hot stage of the embedding machine. Place the embedding box corresponding to the tissue on the mold. The tissue is placed in the embedding mold with hot tweezers, and a little dissolved paraffin is added to the embedding mold, and the embedding mold is placed on the cold table until the paraffin becomes completely solid. Finally, the solid wax block is taken out of the embedding mold.

3.1.3 Preparation of tissue sections

e) Confirm the status of the Leica RM2235 paraffin slicer

Before loading the wax block, make sure that the slice wheel handle is loosely locked, open the knife holder cover, and clean the wax on the knife holder.

Carefully insert an unused disposable knife to confirm that the blade is in place at home and lock the knife holder according to the manufacturer's instructions. Confirm the inclination of the blade, which should be set to 3-6 degrees.

f) Fix the tissue wax block to the desired position in the clipper slot.

Follow the manufacturer's instructions for roughing with the roughing wheel handles so that the wax block slowly advances until the desired tissue plane is exposed.

g) Start slicing, first set the slice thickness, start slicing, and rotate the slice runner forward until the flat wax sheet is formed.

Lock the slice roller handle and use a forceps or brush to move the tissue piece into the spreader (Leica HI1210) and flatten it onto the slide.

h) After roughing or slicing, lock the wheel handle and carefully remove the wax block.

3.1.4 HE staining

HE staining was performed using a fully automated staining workstation consisting of Leica ST5010, Leica TS5010 and Leica CV5030.

Press the <Stain> key in the main menu of the dyeing machine. Conventional HE staining is performed by the up and down arrow selection procedure 15 of the button area. Open the load compartment, place the slicer, and close the load compartment. Start the dye program after pressing the <Load> button. The procedure is as follows:

x) Oven 55 ° C 2 minutes

y) xylene I 10 minutes

z) xylene II 10 minutes

Aa) xylene III 10 minutes

Bb) absolute ethanol I 2 minutes

Cc) absolute ethanol II 1.5 minutes

Dd) 95% ethanol I 1 minute

Ee)80% ethanol I 0.5 min

Ff) Washing for 3 minutes

Gg) hematoxylin dye solution 3 minutes

Hh) washing for 2 minutes

Ii) 1% hydrochloric acid-ethanol 5 seconds

Jj) washing for 2 minutes

Kk) wash for 20 minutes

Ll) 80% ethanol II 1 minute

Mm) Eosin 5 seconds

Nn) 95% ethanol II 30 seconds

Oo) absolute ethanol III 2 minutes

Pp) absolute ethanol IV 2 minutes

Qq) absolute ethanol V 2 minutes

Rr) absolute ethanol VI 2 minutes

Ss) xylene IV 3 minutes

Tt) xylene V 3 minutes

Uu) xylene V 3 minutes

Vv)EXIT xylene 3 minutes

Ww)Leica CV5030 automatic sealing machine cover slice

3.2 Sirius red staining method

The tissue white sheets prepared in 3.1.3 were hand-made into Sirius red stained sections by the following steps:

a) xylene I 15 minutes

b) xylene II 15 minutes

c) absolute ethanol I 3 minutes

d) absolute ethanol II 3 minutes

e) 95% ethanol I 3 minutes

f) 95% ethanol II 3 minutes

g) 80% ethanol 3 minutes

h) 70% ethanol 3 minutes

i) distilled water I 3 minutes

j) distilled water II 3 minutes

k) distilled water III 3 minutes

l) Sirius red dye solution 1 hour

m) distilled water for 5 minutes

n) 95% ethanol III 30 seconds

o) Anhydrous ethanol III 2 minutes

p) absolute ethanol IV 2 minutes

q) Anhydrous ethanol V 2 minutes

r) absolute ethanol VI 2 minutes

s) xylene IV 15 minutes

t) xylene V 15 minutes

u) Neutral gum seal

3.3 Histological non-alcoholic fatty liver disease activity score (NAS) related scoring criteria:

The histological diagnosis and clinical efficacy evaluation of nonalcoholic fatty liver disease (NAFLD) was performed in accordance with the NASH Clinical Research Network Pathology Working Group Guidelines of the National Institutes of Health, and the NAFLD activity score (NAS) was routinely performed.

NAS score (0 to 8 points):

a) Hepatocyte steatosis: 0 points (<5%); 1 point (5% to 33%); 2 points (33% to 66%); 3 points (>66%).

b) Intralobular inflammation (20-fold microscopic count necrosis): 0 points, none; 1 point (<2); 2 points (2 to 4); 3 points (> 4).

c) Hepatocyte balloon-like changes: 0 points, no; 1 point, rare; 2 points, more common.

3.4 Fibrosis scoring criteria:

Stage of liver fibrosis (0~4):

a) 0 points: no fibrosis;

b) 1 point: 1A, mild sinus fibrosis in the 3rd area of the liver acinus; 1B, moderate sinus fibrosis in the 3rd area of the liver acinus; 1C, only fibrosis around the portal vein;

c) 2 points: sinus periplasmic fibrosis in the 3rd district of the liver acinus combined with fibrosis around the portal vein;

d) 3 points: bridging fibrosis;

e) 4: Highly suspicious or confirmed cirrhosis

4. Liver tissue biomarker mRNA detection method

Mini RNA提取试剂盒(Qiagen#74104)提取RNA，当天完成逆转录，逆转录体系配比(ABI#4368814)如下：According to the operation manual, use

RNA was extracted from Mini RNA extraction kit (Qiagen #74104), and reverse transcription was completed on the same day. The reverse transcription system ratio (ABI#4368814) was as follows:

Reverse transcription program:

temperature duration 25°C 10min 37 ° C 120min 85 ° C 5min 4 ° C ∞

The cDNA was stored in a refrigerator at -20 °C.

Fast Universal试剂盒(ABI，#4366072)说明书，进行Q-PCR检测，体系配比如下：according to

Fast Universal kit (ABI, #4366072) instructions for Q-PCR detection, the system is equipped with the following:

Q-PCR program:

Experimental results:

1. Histological results

1.1HE staining picture

HE staining results showed that liver steatosis was significantly increased in nSTZ+HFD mice, balloon-like changes in hepatocytes, and increased hepatic lobular inflammation, pentoxifylline (100mpk, po., tid) and compound 1 (30mpk, po., Bid) 5 weeks of treatment did not significantly improve the pathological changes in liver tissue, such as fatty degeneration, balloon degeneration and inflammatory aggregation in nSTZ+HFD mice. The experimental results are shown in Figure 8.

1.2NAS score

The liver histology of NAS in nSTZ+HFD mice was significantly increased, liver fatification was significantly increased, a small amount of hepatocyte ballooning, hepatic lobules The inflammation is significantly increased. Treatment with pentoxifylline (100 mpk, po., t.i.d.) and Compound 1 (30 mpk, po., b.i.d.) for 5 weeks did not significantly affect liver NAS in nSTZ+HFD mice. The experimental results are shown in Figure 9.

1.3 Sirius red stained picture

The results of Sirius red staining showed that the liver tissue of nSTZ+HFD mice showed more fibrotic changes around the sinus and 3 areas combined with portal vein. See also suspected bridging fibrosis in individual samples. The 5-week treatment group of pentoxifylline (100mpk, po., tid) and compound 1 (30mpk, po., bid) alleviated the degree of fibrosis in the above-mentioned liver tissue of STZ+HFD mice. 1 (30mpk, po., bid) 5 weeks treatment group significantly improved the morphological changes of liver tissue around the sinus periportal and portal vein fibrosis. The experimental results are shown in Figure 10.

1.4 Fibrosis score

The liver fibrosis score of nSTZ+HFD mice increased significantly. The 5-week treatment with pentoxifylline (100mpk, po., tid) reduced the liver fibrosis trend of nSTZ+HFD mice, but statistically, there was no significantity. Compound 5 (30 mpk, po., bid) treatment for 5 weeks significantly reduced liver fibrosis scores in nSTZ+HFD mice. The experimental results are shown in Figure 11.

2. Liver biomarker mRNA detection results

The TNF-α and TGF-β1 mRNA in the liver of nSTZ+HFD mice increased significantly, and the levels of TNF-α and TGF-β1 mRNA in the liver of nSTZ+HFD mice were not significantly treated with pentoxifylline (100mpk, po., tid) for 5 weeks. Sexual effects, compound 1 (30mpk, po., bid) administration 5 weeks nSTZ + HFD mice liver TNF-α and TGF-β1 mRNA levels were significantly reduced. The expression of Collagen 1α1 mRNA in nSTZ+HFD mice was increased. Compound 1 (30mpk, po., bid) was administered for 5 weeks. The expression of Collagen 1α1 mRNA in nSTZ+HFD mice decreased, but the statistical analysis was not significant. Further research was needed. Research and analysis. The experimental results are shown in Figure 12.

3. Serum biochemical indicators test results

The serum ALT and TC levels of nSTZ+HFD mice were significantly increased. Treatment with pentoxifylline (100mpk, po., tid) and compound 1 (30mpk, po., bid) for 5 weeks did not significantly affect the serum of nSTZ+HFD mice. ALT and TC levels. The experimental results are shown in Figure 13.

to sum up:

1. Newborn rats were injected with low-dose streptozotocin (STZ) and fed a high-fat diet for 1 week to form diabetic liver with fatty liver. A persistent high-fat diet results in liver fat deposition, increased hepatic lobular inflammation, and is accompanied by foam-like macrophage infiltration. The nSTZ+HFD mouse model mimics the pathological progression of human diabetes-nonalcoholic steatohepatitis-hepatocellular carcinoma, and is a widely used animal model of nonalcoholic steatohepatitis. Liver histology HE staining images of nSTZ+HFD mice showed significant steatosis, balloon-like changes and lobular inflammation, Sirius red staining showed significant fibrosis, liver inflammatory biomarker TNF-α mRNA was significantly elevated, fibrosis The biomarker TGF-β1 mRNA was significantly elevated, and serum ALT and TG levels were significantly higher than normal mice.

2. Compound 1 (30mpk, po., b.i.d.) After 5 weeks of administration, liver fibrosis was significantly improved in nSTZ+HFD mice, and liver TNF-α and TGF-β1 mRNA levels were significantly decreased.

3. Compound 1 (30mpk, po., b.i.d.) was administered for 5 weeks, and had no significant effect on liver NAS, serum ALT, AST, TG and TC levels in nSTZ+HFD mice.

Experimental Example 6 Study on Anti-rat Cholestatic Liver Fibrosis

main mission:

The effects of Compound 1 on liver histology, serum ALT, AST, TG and TC levels in a rat cholestasis liver fibrosis model were evaluated.

The problem to be solved:

The effect of test compound 1 on liver histology, serum ALT, AST, TG and TC levels in rat cholestasis-type liver fibrosis model, evaluation of compound liver function protection in rat cholestasis liver fibrosis model, reduction of liver inflammation and fiber The role of the role.

Experimental model: (ANIT)-induced rat model of cholestasis-induced liver fibrosis

Experimental animals: SD rats (male, Shanghai Slack Laboratory Animals Co., Ltd.)

Experimental reagents:

26.ANIT(1-NAPHTHYL ISOTHIOCYANATE, Aldrich-N4525)

27.HPMC (Hydroxypropyl methyl cellulose, Sigma-H7509)

experimental method:

1. Experimental procedure

2. In vivo experimental methods

90 rats were ordered, and the animals were grouped on the same day according to the principle of the smallest difference between the groups, with 12 rats in each group and 4 cages per cage. After the animals arrive at the facility, the adaptation period is 5 days. During the adaptation period, animals are provided with normal diet and drinking water, and the health of the animals is monitored daily. If any abnormalities or infections are found, the rats need to be removed from the test group.

On day 0, the drug was administered first. After 2 hours, the normal feed in all the cages except the first group was changed to the ANIT supplement, which was the normal three-day intake of the rats (this amount is based on the previous experimental results). The ANIT additive is updated every three days. The first group of rats consumed normal feed throughout the experiment.

Group Test compound Dosing schedule (dose | mode of administration | frequency | total duration) 1 1% HPMC (pure water configuration) B.i.d, p.o, days 0-7 2 1% HPMC (pure water configuration) B.i.d, p.o, days 0-7 3 INT-747 20mg/kg, p.o, q.d, days 0-7 4 Pentoxifylline 100mg/kg, p.o, t.i.d, days 0-7 5 Compound 1 1mg/kg, p.o, b.i.d, days 0-7 6 Compound 1 3mg/kg, p.o, b.i.d, days 0-7 7 Compound 1 10mg/kg, p.o, b.i.d, days 0-7

All rats were withdrawn from food on the morning of the 8th day until the end of the experiment, with a fasting time of more than 5 hours. Euthanasia method: pentobarbital sodium After drunk, the heart was killed by blood. (Collect as much blood as possible).

1) Collect 2 mL of blood; collect 750 μL of serum and dispense into 3 centrifuge tubes. (Serum collection method: 10000 rpm, centrifugation at 4 ° C for 10 minutes, serum was collected, placed on dry ice and transferred to -80 ° C for storage);

2) Collect the liver of the animal and rinse it with PBS;

3) Wipe dry with a sterilized paper towel, weigh and record;

4) photographing the entire isolated liver;

5) From the largest diameter of the left lobe of the liver, cut one, about 3 mm wide, and immerse 10% of the tissue volume of 10% formalin solution (pure water configuration, need to adjust the PH value to neutral);

6) The remaining left lobe of the liver, take two pieces of 100mg, cut into small pieces and put them into two cryotubes for liver TG/TC detection;

7) The middle lobe of the liver is divided into two, each part is placed in two cryotubes.

3. Histological analysis method

3.1 Organizational Processing

Dehydration of liver tissue, paraffin block production, paraffin section, HE stained paraffin piece thickness 3 μm, Sirius red stained paraffin piece thickness 4 μm. HE staining and Sirius red staining were performed according to the KCI pathological standard staining SOP. The HE stained films were scored by two pathologists (the criteria were found in the pathological report), and the mean values of the two groups were used to analyze the liver damage of each group. Degree; fibrosis stained sections were scanned on a Digital Pathscope 4S scanner and subjected to whole analysis.

3.2 rat cholestasis model inflammatory pathology scoring standard

Experimental results:

1. Histological results

1.1HE staining pictures and results

According to the HE staining results (Fig. 14), the liver inflammation levels were scored according to the inflammatory pathology scoring criteria (Fig. 15), and the results were subjected to t-test. The healthy control group was significantly lower than the model group, but the score of the positive control INT-747 administration group was significantly increased. No improvement in hepatic inflammation was observed in pentoxifylline-100 mg/kg and Compound 1.

1.2 Sirius red staining picture and fibrosis score

Liver tissue sections were stained by Sirius red (Fig. 16), and subjected to a full-scale scan of Digital pathscope 4S. The panoramic sections were analyzed to calculate the percentage of collagen fibers in the sections to determine the degree of fibrosis of each group of animals after compound treatment. Change (Figure 17). The results showed that the scores of the model group were significantly higher than those of the healthy control group, indicating that the liver fibrosis was successfully modeled. Compared with the model group, Compound 1 had an inhibitory effect on liver fibrosis (p=0.03) at the highest dose group of 10 mg/kg in this study, and the inhibition rate was 17%. The improvement in liver fibrosis was similar for INT-747 (p=0.008). However, pentoxifylline did not improve liver fibrosis at a dose of 100 mg/kg.

2. Results of serum biochemical indicators (Table 4)

Compared with the normal group, the ALT, AST, ALP, TBA, TB, DB and IB of the model group were significantly increased, indicating that the modeling was successful. Compared with the model group, the INT-747 group can reduce serum ALT and AST levels to 18% and 25%, respectively (mean comparison), but there is no statistical difference in this trial; Both ketomatole and Compound 1 did not significantly reduce the effects of AST and ALT. None of the drug-administered groups reduced the level of ALP in the serum.

The level of total bile acid (TBA) was significantly increased in the model group compared with the normal group, which was about 9 times higher. Compared with the model group, the TBA level of the INT-747, 20 mg/kg treatment group was significantly reduced by about 75%. The Compound 1 treatment group had a certain effect on reducing total bile acid, which was 37%, 26%, and 33%, respectively, but was not statistically significant.

The results of TB, DB, and IB showed that the values of the model group were significantly higher than those of the normal control group, which were 53-fold, 62-fold, and 36-fold higher, respectively, while the drug-treated group was positive compared with the model group. The control group INT-747 significantly reduced the effects of TB, DB, IB, respectively, by 53%, 54%, 49%; Compound 1 decreased the levels of TB, DB, IB under high dose, and can The corresponding dose dependence was seen, but there was no statistically significant experimental result compared to the model group.

to sum up:

1) In the ANIT-induced cholestasis-induced liver fibrosis rat model, the serum ALT/AST/ALP levels in the model group were significantly higher in the serum than in the healthy control group; TBA/TB/DB/IB in serum The results of the test were also significantly increased; in the inflammatory scores, the degree of fibrosis in the model group was significantly higher than that in the healthy control group; in the pathological fibrosis results, the fibrosis of the model group was compared with the healthy control group. The degree is obviously increased.

2) Compound 1 (10mpk, po., b.i.d.) treatment group showed significant anti-fibrotic effect on the degree of pathological fibrosis.

3) Compound 1 treatment groups had no significant effect on serum ALT, AST, TG and TC levels.

Experimental Example 7 Study on the effect of carbon tetrachloride-induced liver fibrosis in mice

main mission:

The effects of Compound 1 on liver histology in the liver fibrosis model induced by carbon tetrachloride in mice, as well as serum ALT, AST, TG and TC levels were evaluated.

The problem to be solved:

The effect of test compound 1 on liver histology, serum ALT, AST, TG and TC levels in mice induced by carbon tetrachloride in liver fibrosis model, and evaluation of compound liver function protection in mice induced by carbon tetrachloride-induced liver fibrosis To reduce the effects of liver damage and fibrosis.

Experimental model: carbon tetrachloride-induced liver fibrosis in mice

Experimental animals: C57BL/6 mice (male, Shanghai Lingchang Biotechnology Co., Ltd.)

Experimental reagents:

1.CCl ₄ (Jiangsu Qiangsheng Functional Chemical Co., Ltd.)

2. Olive oil (Aladdin Industries, 0108686-500ml)

3.HPMC (Hydroxypropyl methyl cellulose, Sigma-H7509)

experimental method:

1. Experimental procedure

2. In vivo experimental methods

108 mice were ordered, of which 3 were reserved. On the day the animals arrived, they were grouped according to the principle of the smallest difference between the groups, with 15 in each group. After the animals arrive at the facility, the adaptation period is 5 days. During the adaptation period, the health of the animals was monitored daily. If any abnormalities or infections are found, the mice will need to be removed from the test group. The 25% CCl ₄ solution is prepared in pure olive oil and is ready for use. Except for the animals of the first group, the injection time was on day 0, day 4, day 8, and day 12, and all other groups of mice were intraperitoneally injected, and administered according to the body weight, 2 mL/kg. The first group of mice was injected intraperitoneally with pure olive oil.

Group Test compound Dosing regimen (dose, frequency, mode of administration, total duration)

1 1% HPMC (pure water configuration) 1% HPMC (pure water configuration), b.i.d**, p.o., 0-1-3 days 2 1% HPMC (pure water configuration) 1% HPMC (pure water configuration), b.i.d**, p.o., 0-1-3 days 3 INT-747 INT-747, 20mg/kg, q.d, p.o., 0-1-3 days 4 Pentoxifylline Pentoxifylline, 100 mg/kg, t.i.d*, p.o., days 0-13 5 Compound 1 Compound 1, 1 mg/kg, b.i.d**, p.o., days 0-13 6 Compound 1 Compound 1, 3 mg/kg, b.i.d**, p.o., days 0-13 7 Compound 1 Compound 1, 10 mg/kg, b.i.d**, p.o., day 0-13

All mice were withdrawn from food on the morning of the 14th day until the end of the experiment, with a fasting time of more than 5 hours. Euthanasia method: After anesthesia, blood was collected from the posterior venous plexus, and the neck was removed to confirm death. (Collect as much blood as possible).

1) Collect as much blood as possible; operate to collect 250 ul of serum and dispense into the corresponding centrifuge tube. (Serum collection method: 10000 rpm, centrifugation at 4 degrees for 10 minutes, serum was collected, and placed on dry ice and transferred to -80 degrees for storage.)

2) Collect animal liver and rinse with PBS

3) Wipe dry with a sterilized paper towel, weigh and record;

4) taking pictures;

5) From the largest diameter of the left lobe of the liver, cut a piece, about 3mm wide, and immerse it in 10 times volume of 10% formalin solution (pure water configuration, need to adjust PH value in advance)), and fix it vertically;

6) The remaining left hepatic lobe, cut into small pieces and packed in two cryotubes for detecting liver TG/TC;

7) The middle lobe of the liver is divided into two and placed in two cryotubes as samples for protein analysis and mRNA detection. It should be noted that the samples of samples 6) and 7) need to be quickly immersed in liquid nitrogen after collection, and it is necessary to ensure that the cryotubes are completely immersed in liquid nitrogen after being thrown into liquid nitrogen.

3. Histological analysis method

3.1 Organizational Processing

Dehydration of liver tissue, paraffin block production, paraffin section, HE stained paraffin piece thickness 3 μm, Sirius red stained paraffin piece thickness 4 μm. HE staining and Sirius red staining were performed according to the KCI pathological standard staining SOP. HE-stained films were scored by two pathologists, and the mean values of the two groups were used to analyze the degree of liver damage in each group; fibrosis stained sections. After a full scan of the Digital Pathscope 4S scanner, a full analysis was performed.

3.2 Carbon tetrachloride induced liver injury animal model pathological scoring standard

Experimental results:

1. Histological results

1.1HE staining pictures and results

Liver pathological sections were scored by HE staining (Fig. 18) according to pathological scoring criteria for inflammation, ballooning, watery degeneration and necrosis. The results showed that compared with the healthy control group, the scores of the four indicators in the model group were extremely significantly increased, which proved that the modeling was successful (Fig. 19, Fig. 20, Fig. 21, Fig. 22).

Compared with the model group, the scores of watery degeneration and necrosis in the three dose groups of Compound 1 were lower than those in the model group (both statistically significant), and the effect showed a dose-dependent manner. The improvement of watery degeneration and necrosis by the compound 1-10 mg/kg group was comparable to that of the pentoxifylline-100 mg/kg group. Compared with the model group, the water-like degeneration and necrosis scores of the compound 1-10 mg/kg were decreased by 0.93 and 0.86, respectively, and the water-like degeneration and necrosis scores of the pentoxifylline-100 mg/kg group were decreased by 0.90 and 0.67, respectively.

The sum of the scores of the four indicators showed that each test compound had a significant improvement in the sum score. There was a significant improvement in liver damage in the three test doses of Compound 1, with a dose-related effect comparable to pentoxifylline - 100 mg/kg (Figure 23).

1.2 Sirius red staining picture and fibrosis score

The Sirius red staining results (Fig. 24) read the fibrosis area (Fig. 25). The scores of the model group were significantly higher than those of the healthy control group, indicating that the liver fibrosis was successfully modeled. Compared with the model group, the high, medium and low dose groups of Compound 1, pentoxifylline and INT-747 all significantly reduced fibrosis of liver tissue.

2. Results of serum biochemical indicators (Table 5)

Compared with the normal group, the ALT, AST, TG, and TC of the model group were significantly increased, indicating that the modeling was successful.

Compared with the model group, Compound 1 had a certain effect of lowering ALT and AST, and it was dose-dependent. Compound 1 reduced ALT by 24%, 52% and 56% at doses of 1, 3 and 10 mg/kg, respectively. AST decreased by 23%, 42%, and 52%, respectively. However, only the 1-10 mg/kg dose of the compound had a statistically significant change in AST (p=0.02). INT-747 significantly reduced serum ALT and AST levels by 75% and 64%, respectively, and INT-747 also decreased TG and TC by 11% and 12%, respectively. Compared with the model group, pentoxifylline did not improve ALT, AST, TG, and TC.

to sum up:

1) Three doses of Compound 1 can reduce ALT/AST in serum to a certain extent, and show a certain dose-dependent, especially the highest amount of 10 mg / kg can significantly reduce AST;

2) From the results of HE staining, Compound 1 can significantly improve liver damage, including inflammation level, degree of watery degeneration, and degree of necrosis. The improvement effect of these indicators on the dose of 10 mg/kg was the strongest among the three doses.

3) Compound 1 can significantly reduce the degree of liver fibrosis at three test doses.

Claims (8)

The use of a compound of the formula (I), a tautomer thereof or a pharmaceutically acceptable salt thereof for the preparation of a medicament for the treatment or prevention of liver disease,

among them, Structural unit

Replace with

Specifically replaced with

L _{11 is} selected from the group consisting of empty, C(R)(R'); R, R' are each independently selected from H, halogen, OH, NH ₂ , CN, optionally substituted 1 to 6 alkyl or heteroalkyl; Optionally, R, R' may be cyclized to a 3-6 membered cycloalkyl or heterocycloalkyl group; A is empty or is selected from optionally substituted cycloalkyl, heterocycloalkyl, aryl, heteroaryl; L _{12 is} selected from an optionally substituted 1 to 6 membered alkyl or heteroalkyl group; R _{1 is} selected from an optionally substituted 1 to 6 membered alkyl group, a 3 to 6 membered cycloalkyl group or a heteroalkyl group; "Heter" stands for N, O, S, C(=O), S(=O), S(=O) ₂ , and the number of heteroatoms per group is selected from 1, 2, 3 or 4; Specifically, the disease is selected from the group consisting of nonalcoholic steatohepatitis and liver fibrosis. The use according to claim 1, wherein the substituents in R, R', A, L ₁₂ and R ₁ are each independently selected from the group consisting of halogen, OH, NH ₂ , CN, optionally substituted 1 to 6-membered alkyl group. a 3-6 membered cycloalkyl or heteroalkyl group, the number of each group substituent being independently selected from 1, 2 or 3; specifically, a substituent in R, R', A, L ₁₂ , R ₁ Individually selected from the group consisting of halogen, CF ₃ , CN, OH, Me, Et, n-propyl, isopropyl, cyclopropyl,

The application according to claim 1 or 2, wherein R and R' are each independently selected from the group consisting of H, Me, CF ₃ and Et; Specifically, L _{11 is} selected from

The use according to claim 1 or 2, wherein A is selected from the group consisting of: a 4- to 12-membered cycloalkyl or heterocycloalkyl group, a 5- to 12-membered aryl group or a heteroaryl group; Specifically, A is selected from the group consisting of: optionally substituted: cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, pentylpentyl, phenyl, pyridyl, pyrazinyl, oxazolyl, isoxazolyl a thiazolyl group, a bicyclo[1.1.1]pentane, or a bicyclic group, a spiro group or a bicyclic group selected from any two of the above groups; Specifically, A is selected from the group consisting of:

Specifically, A is selected from

The use according to claim 1 or 2, wherein L _{12 is} selected from the group consisting of methylene groups,

The use according to claim 1 or 2, wherein R _{1 is} selected from the group consisting of Me, CHF ₂ , CF ₃ , Et, CH ₂ CF ₃ , isopropyl,

Cyclopropyl,

The use of the compound of the formula in the preparation of a medicament for treating or preventing liver disease:

The use of the compound of the formula in the preparation of a medicament for treating or preventing liver disease:

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