Tricyclic fused heterocycle PDE3 / 4 dual inhibitor as well as preparation method and application thereof

Abstract

The invention provides a tricyclic fused heterocyclic PDE3 / 4 dual inhibitor as well as a preparation method and application thereof, and particularly provides a compound with a structural formula I or a pharmaceutically acceptable form thereof, and each substituent group is defined in the specification. The compound or the pharmaceutically acceptable form of the compound can be used as a phosphodiesterase inhibitor and has relatively high activity. # imgabs0 #

Description

Technical Field

[0001] The present invention relates to a tricyclic fused heterocyclic compound and a preparation method and application thereof, and in particular, to a tricyclic fused heterocyclic compound and a preparation method thereof and application thereof as a phosphodiesterase PDE3 / 4 dual inhibitor. Background Art

[0002] Phosphodiesterase (PDE) belongs to a superfamily enzyme system, including at least 11 families and 22 subtypes, and is involved in intracellular and extracellular information transmission and functional regulation. PDE can catalyze the hydrolysis of intracellular second messengers cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP) to generate AMP and GMP, respectively.

[0003] The PDE3 family consists of two genes, namely PDE3A and PDE3B. The activity of PDE3 in the respiratory system is mainly concentrated in alveolar macrophages, endothelial cells, and platelets. PDE3 is involved in the regulation of various physiological activities in the body, such as vasodilation of vascular smooth muscle, anti-platelet aggregation, anti-thrombosis, cardiotonic and anti-cell proliferation. Excessive use of PDE3 inhibitors can cause adverse reactions such as hypotension and tachycardia, which greatly limits its clinical application.

[0004] PDE4 is an enzyme that specifically hydrolyzes cAMP. Its family consists of four subtypes: PDE4A, PDE4B, PDE4C, and PDE4D. Each subtype has a corresponding gene encoding and has different cell distribution and functions. PDE4 is mainly distributed in airway smooth muscle cells, inflammatory cells, and immune cells, regulating intracellular cAMP levels. Currently, most clinical PDE4 inhibitors have a certain degree of side effects, such as gastrointestinal reactions such as nausea and vomiting, and even depression.

[0005] Given the limitations of using either PDE3 or PDE4 inhibitors alone and the side effects of PDE inhibitors, dual inhibition of inhaled PDE3 / 4 appears to be a more attractive approach to target the key pathological features of COPD and asthma. Evidence has shown that inhaled dual-target PDE3 / 4 inhibitors have synergistic inhibitory effects, including synergistic anti-inflammatory and bronchodilatory effects.

[0006] CN100415743C discloses a pyrimido[6,1a]isoquinolin-4-one derivative:

[0007]

[0008] The compound of the general formula is used as a PDE inhibitor for the treatment of respiratory diseases such as asthma, has a longer duration of action than troquinein, and does not have the very bitter taste of troquinein.

[0009] CN112368281A discloses a class of tricyclic compounds as dual PDE3 / PDE4 inhibitors:

[0010]

[0011] The compounds of this general formula can be used to prepare drugs related to PDE3 / PDE4, especially drugs for chronic obstructive pulmonary disease (COPD).

[0012] There is still an urgent need in the art for new PDE3 / PDE4 inhibitors, especially PDE3 / PDE4 inhibitors with high activity and good drug-likeness. SUMMARY OF THE INVENTION

[0013] An object of the present invention is to provide a new compound as a PDE inhibitor.

[0014] Another object of the present invention is to provide a preparation method of the compound.

[0015] Another object of the present invention is to provide an application of the compound.

[0016] Another object of the present invention is to provide a pharmaceutical composition containing the compound and its application.

[0017] Another object of the present invention is to provide an intermediate for preparing the compound.

[0018] Another object of the present invention is to provide a preparation method of the intermediate.

[0019] <First aspect>

[0020] The present invention provides a compound having the structural formula I or a pharmaceutically acceptable form thereof,

[0021]

[0022] Wherein:

[0023] R 1 、R 2 Each independently selected from H, C 1-6 linear alkyl, C 3-6 branched alkyl and C 3-6 cycloalkyl; the linear alkyl, branched alkyl or cycloalkyl is optionally further substituted by 0 to 4 substituents selected from H, F, Cl, Br, I, OH, =O, NH 2 、CN, COOH, C 1-4 alkyl, C 1-4 alkoxy;

[0024] R 3 、R 4 、R5 Each independently selected from H, halogen, CN, C 1-6 alkoxy, C 1-6 linear alkyl, C 3-6 branched alkyl, and C 3-6 cycloalkyl; the alkoxy, linear alkyl, branched alkyl or cycloalkyl is optionally further substituted by 0 to 4 substituents selected from H, F, Cl, Br, I, OH, ═O, NH 2 , CN, COOH, C 1-4 alkyl, C 1-4 alkoxy;

[0025] R 6 is selected from The H in the said R 6 is optionally further substituted by 0 to 4 substituents selected from F, Cl, Br, I, OH, ═O, NH 2 , CN, COOH, C 1-4 alkyl, C 1-4 alkoxy;

[0026] R 7 is selected from H, ═O, NH 2 , CN, C 1-6 linear alkyl, C 3-6 branched alkyl, C 3-6 cycloalkyl and The H in the said R 7 is optionally further substituted by 0 to 4 substituents selected from F, Cl, Br, I, OH, ═O, NH 2 , CN, COOH, C 1-4 alkyl, C 1-4 alkoxy;

[0027] L is selected from n is 0, 1 or 2; k is 0, 1 or 2; the H in the said L is optionally further substituted by 0 to 4 substituents selected from F, Cl, Br, I, OH, ═O, NH 2 , CN, COOH, C 1-4 alkyl, C 1-4 alkoxy;

[0028] X is N;

[0029] Y is C 1-3 alkylene or absent; the alkylene is optionally further substituted by 0 to 4 substituents selected from F, Cl, Br, I, OH, ═O, NH 2 , CN, COOH, C 1-4 alkyl, C 1-4 alkoxy;

[0030] Optionally, R1 and R 2 are linked to the connected O to form a 5- or 6-membered ring; the H in the 5- or 6-membered ring is optionally further substituted by 0 to 4 substituents selected from F, Cl, Br, I, OH, =O, NH 2 , CN, COOH, C 1-4 alkyl, C 1-4 alkoxy;

[0031] Optionally, X and L are linked through C 1-2 alkylene to form a 5- or 6-membered ring; the H in the 5- or 6-membered ring is optionally further substituted by 0 to 4 substituents selected from F, Cl, Br, I, OH, =O, NH 2 , CN, COOH, C 1-4 alkyl, C 1-4 alkoxy;

[0032] Optionally, R 6 and L together form a 5- or 6-membered ring; the H in the 5- or 6-membered ring is optionally further substituted by 0 to 4 substituents selected from F, Cl, Br, I, OH, =O, NH 2 , CN, COOH, C 1-4 alkyl, C 1-4 alkoxy;

[0033] Optionally, the H of the compound of formula I is optionally further substituted by 0 to 6 D;

[0034] And, the compound of formula I is not:

[0035]

[0036] According to some specific embodiments of the present invention, in the compound of formula I of the present invention or its pharmaceutically acceptable form, R 1 , R 2 are each independently selected from H, CH 3 , CHF 2 , CD 3 .

[0037] According to some specific embodiments of the present invention, in the compound of formula I of the present invention or its pharmaceutically acceptable form, R 1 , R 2 correspond to the groups or values shown in any one of Compounds 1-31, respectively.

[0038] According to some specific embodiments of the present invention, in the compound of formula I of the present invention or its pharmaceutically acceptable form, R 3 , R 4 , R 5 are each independently selected from H, CH3 , i-Pr, OMe, CN, CD 3 or halogen.

[0039] In some specific embodiments of the present invention, in the compound of formula I or its pharmaceutically acceptable form of the present invention, R 3 , R 4 , R 5 correspond to the groups or values shown in any one of Compounds 1 - 31, respectively.

[0040] In some specific embodiments of the present invention, in the compound of formula I or its pharmaceutically acceptable form of the present invention, R 6 is

[0041] In some specific embodiments of the present invention, in the compound of formula I or its pharmaceutically acceptable form of the present invention, R 7 is H, =O or

[0042] In some specific embodiments of the present invention, in the compound of formula I or its pharmaceutically acceptable form of the present invention, L is selected from n is 0, 1 or 2; k is 1 or 2.

[0043] In some specific embodiments of the present invention, in the compound of formula I or its pharmaceutically acceptable form of the present invention, L corresponds to the group or value shown in any one of Compounds 1 - 31.

[0044] In some specific embodiments of the present invention, in the compound of formula I or its pharmaceutically acceptable form of the present invention, Y is absent.

[0045] In some specific embodiments of the present invention, in the compound of formula I or its pharmaceutically acceptable form of the present invention, R 1 , R 2 , R 3 , R 4 , R 5 , R 6 , R 7 , L, X, Y correspond to the groups or values shown in any one of Compounds 1 - 31, respectively.

[0046] In some specific embodiments of the present invention, in the compound of formula I or its pharmaceutically acceptable form of the present invention, the compound is selected from one or more of the compounds shown in Table 1:

[0047] Table 1

[0048]

[0049]

[0050]

[0051] According to some specific embodiments of the present invention, in the compound of formula I or its pharmaceutically acceptable form of the present invention, the pharmaceutically acceptable form is selected from pharmaceutically acceptable salts or co-crystals, stereoisomers, tautomers, deuterated compounds, solvates, chelates, non-covalent complexes or prodrugs.

[0052] <Second aspect>

[0053] The present invention also provides an intermediate compound having the structure shown in formula II:

[0054]

[0055] Wherein, R 1 , R 2 , R 3 , R 4 , R 5 , R 7 , X, Y are defined as in any one of the compounds of formula I or their pharmaceutically acceptable forms according to the <first aspect> of the present invention;

[0056] L 1 is selected from n is 0, 1 or 2; k is 0, 1 or 2; the H in the L 1 is optionally further substituted by 0 to 4 substituents selected from F, Cl, Br, I, OH, =O, NH 2 , CN, COOH, C 1-4 alkyl, C 1-4 alkoxy.

[0057] According to some specific embodiments of the present invention, the intermediate compound of the present invention has the following structure:

[0058]

[0059] <Third aspect>

[0060] The present invention also provides a method for preparing the compound or its pharmaceutically acceptable form described in the <first aspect> of the present invention. The synthetic route can be designed and synthesized according to the chemical structure of the compound of formula I or its pharmaceutically acceptable form with reference to the known methods in the art to prepare the compound of formula I or its pharmaceutically acceptable form.

[0061] According to some specific embodiments of the present invention, the method for preparing the compound or its pharmaceutically acceptable form described in the <first aspect> of the present invention includes:

[0062] Perform a modification reaction on the carboxyl terminus shown in the intermediate compound of Formula II according to the <second aspect> of the present invention to prepare a compound having Structural Formula I.

[0063] According to some specific embodiments of the present invention, the method for preparing the compound or its pharmaceutically acceptable form according to the <first aspect> of the present invention further includes a process for preparing the intermediate compound according to the <second aspect> of the present invention.

[0064] According to some specific embodiments of the present invention, the method for preparing the compound or its pharmaceutically acceptable form according to the <first aspect> of the present invention includes the steps shown in any one of the reaction routes in Examples 1 - 18.

[0065] According to some specific embodiments of the present invention, the method for preparing the compound or its pharmaceutically acceptable form according to the <first aspect> of the present invention includes:

[0066]

[0067] <Fourth aspect>

[0068] The present invention also provides a pharmaceutical composition, which includes: the compound or its pharmaceutically acceptable form (preferably a pharmaceutically acceptable salt) according to the <first aspect> of the present invention, and a pharmaceutically acceptable carrier, excipient, and / or one or more other therapeutic agents.

[0069] <Fifth aspect>

[0070] The present invention also provides the use of the compound or its pharmaceutically acceptable form (preferably a pharmaceutically acceptable salt) according to the <first aspect> of the present invention or the pharmaceutical composition according to the <fourth aspect> of the present invention in the preparation of a preparation for inhibiting phosphodiesterase. Preferably, the phosphodiesterase includes PDE3 and / or PDE4.

[0071] <Sixth aspect>

[0072] The present invention also provides the use of the compound or its pharmaceutically acceptable form (preferably a pharmaceutically acceptable salt) according to the <first aspect> of the present invention or the pharmaceutical composition according to the <fourth aspect> of the present invention in the preparation of a drug for treating phosphodiesterase-related diseases.

[0073] The present invention also provides a method for treating phosphodiesterase-related diseases, which includes administering to a subject an effective amount of the compound or its pharmaceutically acceptable form (preferably a pharmaceutically acceptable salt) according to the <first aspect> of the present invention or the pharmaceutical composition according to the <fourth aspect> of the present invention.

[0074] According to some specific embodiments of the present invention, the phosphodiesterase includes PDE3 and / or PDE4.

[0075] According to some specific embodiments of the present invention, the phosphodiesterase-related diseases include respiratory diseases such as asthma.

[0076] According to some specific embodiments of the present invention, the subject is a mammal or a human, preferably, the subject is a human.

[0077] The compound of the present invention having the structural formula I or its pharmaceutically acceptable form can be used as a phosphodiesterase inhibitor, has high inhibitory activity against phosphodiesterase, especially PDE3 and / or PDE4, and has practical value.

[0078] Definitions and Explanations

[0079] Unless otherwise specified, the following terms and phrases used herein are intended to have the following meanings. A particular term or phrase should not be considered indeterminate or unclear without a specific definition, but should be understood in accordance with its ordinary meaning. When a trade name appears herein, it is intended to refer to the corresponding product or its active ingredient.

[0080] Unless otherwise specified, a solid wedge bond and a dashed wedge bond represent the absolute configuration of a stereocenter, and a solid straight bond and a dashed straight bond represent the relative configuration of a stereocenter. A wavy line represents a solid wedge bond or a dashed wedge bond or a wavy line represents a solid straight bond and a dashed straight bond

[0081] The term "pharmaceutically acceptable" refers to those compounds, materials, compositions and / or dosage forms that are within the scope of sound medical judgment, suitable for contact with human and animal tissues, without excessive toxicity, irritation, allergic reaction or other problems or complications, and commensurate with a reasonable benefit / risk ratio.

[0082] The term "pharmaceutically acceptable salt" or "its pharmaceutically acceptable salt" refers to salts of the compounds of the present invention, prepared from compounds having specific substituents found in the present invention with relatively non-toxic acids or bases. When a compound of the present invention contains a relatively acidic functional group, a base addition salt can be obtained by contacting the neutral form of such a compound with a sufficient amount of a base in a pure solution or a suitable inert solvent. Pharmaceutically acceptable base addition salts include sodium, potassium, calcium, ammonium, organic amine or magnesium salts or similar salts. When a compound of the present invention contains a relatively basic functional group, an acid addition salt can be obtained by contacting the neutral form of such a compound with a sufficient amount of an acid in a pure solution or a suitable inert solvent. Examples of pharmaceutically acceptable acid addition salts include inorganic acid salts, where the inorganic acids include, for example, hydrochloric acid, hydrobromic acid, nitric acid, carbonic acid, bicarbonate, phosphoric acid, hydrogen phosphate, dihydrogen phosphate, sulfuric acid, hydrogen sulfate, hydroiodic acid, phosphorous acid, etc.; and organic acid salts, where the organic acids include, for example, acetic acid, propionic acid, isobutyric acid, maleic acid, malonic acid, benzoic acid, succinic acid, suberic acid, fumaric acid, lactic acid, mandelic acid, phthalic acid, benzenesulfonic acid, p-toluenesulfonic acid, citric acid, tartaric acid, methanesulfonic acid and similar acids; also salts of amino acids (such as arginine, etc.), and salts of organic acids such as glucuronic acid. Certain specific compounds of the present invention contain both basic and acidic functional groups and can thus be converted into either base or acid addition salts.

[0083] The pharmaceutically acceptable salts of the present invention can be synthesized by conventional chemical methods from parent compounds containing acid or base moieties. In general, the preparation method of such salts is to react these compounds in the form of free acids or bases with a stoichiometric amount of an appropriate base or acid in water or an organic solvent or a mixture of both.

[0084] The term "eutectic" refers to a crystalline material that contains two or more distinct solids at room temperature, each solid having different physical properties, such as structure, melting point and heat of fusion.

[0085] The term "stereoisomer" (or "optical isomer") refers to a stable isomer that has a perpendicular plane of asymmetry due to the presence of at least one chiral element (including chiral center, chiral axis, chiral plane, etc.), and thus can rotate plane-polarized light. Due to the presence of asymmetric centers and other chemical structures in the compounds of the present invention that may lead to stereoisomerism, the present invention also includes these stereoisomers and their mixtures. Since the compounds and salts thereof of the present invention may include asymmetric carbon atoms, they can exist in the form of a single stereoisomer, racemate, enantiomers, and a mixture of diastereomers. Generally, these compounds can be prepared in the form of a racemic mixture. However, if desired, such compounds can be prepared or separated to obtain pure stereoisomers, i.e., single enantiomers or diastereomers, or a mixture enriched in a single stereoisomer (purity ≥ 98%, purity ≥ 95%, ≥ 93%, ≥ 90%, ≥ 88%, ≥ 85% or ≥ 80%). The single stereoisomers of the compounds are synthesized from optically active starting materials containing the required chiral centers, or are prepared by separating or resolving the mixture of enantiomeric products obtained, such as converting to a mixture of diastereomers and then separating or recrystallizing, chromatographing, using chiral resolving agents, or directly separating the enantiomers on a chiral chromatographic column. The starting compounds with specific stereochemistry can be obtained commercially or prepared according to the methods described herein and then resolved by methods well known in the art. Unless otherwise indicated, all stereoisomeric forms of the compounds of the present invention are within the scope of the compounds of the present invention.

[0086] The term "tautomer" (or "tautomeric form") refers to structural isomers with different energies that can be interconverted through a low energy barrier. If tautomerism is possible (such as in solution), a chemical equilibrium of tautomers can be achieved. For example, proton tautomers (or proton-transfer tautomers) include (but are not limited to) interconversions through proton migration, such as keto-enol isomerization, imine-enamine isomerization, amide-imidol isomerization, etc. Unless otherwise indicated, all tautomeric forms of the compounds of the present invention are within the scope of the compounds of the present invention.

[0087] Unless otherwise indicated, the compounds represented by the structural formulas of the present invention can be in the form of a purified single stereoisomer or tautomer, or in the form of a mixture containing multiple stereoisomers or tautomers.

[0088] The term "solvate" refers to a substance formed by the binding of a compound of the present invention or its pharmaceutically acceptable salt with at least one solvent molecule through non-covalent intermolecular forces. Common solvates include (but are not limited to) hydrates, ethanolates, acetonates, etc.

[0089] The term "chelate" refers to a complex with a cyclic structure, which is obtained by the chelation of two or more ligands with the same metal ion to form a chelate ring.

[0090] The term "non-covalent complex" is formed by the interaction of a compound with another molecule, where no covalent bond is formed between the compound and the molecule. For example, complexation can occur through van der Waals interactions, hydrogen bonding, and electrostatic interactions (also known as ionic bonding).

[0091] The term "prodrug" refers to a derivative compound that can directly or indirectly provide the compound of the present invention after being administered to a patient. Particularly preferred derivative compounds or prodrugs are those that can improve the bioavailability of the compound of the present invention (e.g., are more readily absorbed into the bloodstream) when administered to a patient, or those that facilitate the delivery of the parent compound to the site of action (e.g., the lymphatic system). Unless otherwise indicated, all prodrug forms of the compounds of the present invention are within the scope of the present invention, and various prodrug forms are well known in the art.

[0092] The term "each independently" means that at least two groups (or ring systems) with the same or similar value ranges present in a structure can have the same or different meanings in a particular situation. For example, if substituent X and substituent Y are each independently hydrogen, halogen, hydroxyl, cyano, alkyl, or aryl, then when substituent X is hydrogen, substituent Y can be either hydrogen or halogen, hydroxyl, cyano, alkyl, or aryl; similarly, when substituent Y is hydrogen, substituent X can be either hydrogen or halogen, hydroxyl, cyano, alkyl, or aryl.

[0093] The terms "comprising" and "including" are used in their open, non-limiting sense.

[0094] "Optionally" or "optionally" means that the subsequently described event or condition may but need not occur, and the description includes both the case where the described event or condition occurs and the case where the described event or condition does not occur.

[0095] The term "substituted" means that any one or more hydrogen atoms on a particular atom are replaced by a substituent, which can include deuterium and variants of hydrogen, provided that the valence state of the particular atom is normal and the resulting compound is stable. When the substituent is oxygen (i.e., =O), it means that two hydrogen atoms are replaced. Oxygen substitution does not occur on aromatic groups. The term "optionally substituted" means that it can be substituted or not substituted, and unless otherwise specified, the type and number of substituents can be arbitrary based on what is chemically achievable.

[0096] When any variable (e.g., R) appears 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's, the group may optionally be substituted with up to two R's, and each R has independent options in each case. In addition, combinations of substituents and / or their variants are permitted only if such combinations result in stable compounds.

[0097] When a variable selected from among those in a structural formula is missing, it means its absence. For example, when R in C - R is selected as missing, it means the structure is actually C.

[0098] When a variable connecting two groups in a structural formula is selected from among a bond or absence, it means the two groups it connects are directly linked. For example, when L in A - L - Z represents a bond or absence, it means the structure is actually A - Z.

[0099] When it is not specified which atom of an enumerated substituent is bonded to the group being substituted, such a substituent may be bonded through any of its atoms. For example, a phenyl group as a substituent may be bonded to the group being substituted through any carbon atom on the benzene ring.

[0100] Unless otherwise specified, the term "alkyl" is used to denote a straight - chain or branched - chain saturated hydrocarbon group, which may be mono - substituted (e.g., - CH 2 F) or poly - substituted (e.g., - CF 3 ), and may be monovalent (e.g., methyl), divalent (e.g., methylene), or polyvalent (e.g., methine). Examples of alkyl groups include methyl (Me), ethyl (Et), propyl (e.g., n - propyl and isopropyl), butyl (e.g., n - butyl, isobutyl, s - butyl, t - butyl), pentyl (e.g., n - pentyl, isopentyl, neopentyl), etc.

[0101] Unless otherwise specified, the term "alkylene" refers to a divalent straight - chain or branched - chain alkane group consisting only of carbon and hydrogen atoms, having no unsaturation, and connected to other moieties through two single bonds, including (but not limited to) methylene, 1,1 - ethylene, and 1,2 - ethylene, etc. For example, "C 1-3 alkylene" refers to a saturated divalent straight - chain or branched - chain alkyl group containing 1 to 3 carbon atoms.

[0102] Unless otherwise specified, "cycloalkyl" includes any stable cyclic or polycyclic hydrocarbon group, any carbon atom of which is saturated, and which may be mono - substituted or poly - substituted, and may be monovalent, divalent, or polyvalent. Examples of these cycloalkyl groups include, but are not limited to, cyclopropyl, norbornyl, [2.2.2] bicyclooctane, [4.4.0] bicyclodecane, etc.

[0103] Unless otherwise specified, the term "alkoxy" means an alkyl group linked to the remainder of the molecule through an oxygen atom, where the alkyl group has the meaning as described in the present invention. Unless otherwise specified, C 1-5 Alkoxy groups include C1, C2, C3, C4, and C5 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. The alkoxy group may optionally be substituted by one or more substituents described in the present invention.

[0104] Unless otherwise specified, the term "3- to 6-membered ring" means a saturated or unsaturated monocyclic ring containing or not containing heteroatoms, having 3, 4, 5, or 6 C, O, S, or N atoms in the ring; the "3- to 6-membered ring" can be linked to the remainder of the structural formula through any one carbon atom, or, if present, a nitrogen atom.

[0105] Unless otherwise specified, the term "halogen" by itself or as part of another substituent means a fluorine (F), chlorine (Cl), bromine (Br), or iodine (I) atom. The term "hydroxyl" refers to -OH. The term "cyano" refers to -CN. The term "amino" refers to -NH 2 。

[0106] Unless otherwise specified, the term "pharmaceutical composition" means a mixture formed by one or more compounds described in the present invention or pharmaceutically acceptable forms and other chemical components, where the "other chemical components" refer to pharmaceutically acceptable carriers, excipients, and / or one or more other therapeutic agents. A "carrier" is a material that does not cause significant irritation to the organism and does not eliminate the biological activity and properties of the administered compound. An "excipient" is an inert substance added to the pharmaceutical composition to facilitate the administration of the compound. Non-limiting examples include calcium carbonate, calcium phosphate, sugars, starches, cellulose derivatives (including microcrystalline cellulose), gelatin, vegetable oils, polyethylene glycols, diluents, granulating agents, lubricants, binders, and disintegrants.

[0107] The compounds of the present invention have good PDE inhibitory activity and have potential application value in the treatment of diseases related to PDE, especially those related to PDE3 / PDE4. Detailed Description of the Invention

[0108] The embodiments of the present invention will be described in detail below in conjunction with the examples. However, those skilled in the art will understand that the following examples are only for illustrating the present invention and should not be construed as limiting the scope of the present invention. For those not specified in the examples, they are carried out under conventional conditions or conditions recommended by the manufacturer. For reagents or instruments not specified for the manufacturer, they are all conventional products that can be obtained commercially.

[0109] The structure of the compound was determined by nuclear magnetic resonance or mass spectrometry. Nuclear magnetic resonance was measured using a BRUKER 400M nuclear magnetic resonance instrument, and the measuring solvent was deuterated dimethyl sulfoxide (DMSO-d 6 ), or deuterated chloroform (CDCl 3 ). The internal standard was tetramethylsilane (TMS), and the chemical shift (δ) was given in units of 10 -6 (ppm). Mass spectrometry was measured using a Waters ACQUITY Arc / ACQUITY QDa or a Thermo U3000-ISQ EC liquid chromatography-mass spectrometry instrument.

[0110] High performance liquid chromatography analysis was performed using a Thermo U3000 high pressure liquid chromatography instrument. High performance liquid chromatography preparation was performed using a Hanbang DAC-50 or a Shimadzu LC-20AP preparative chromatograph.

[0111] Reaction monitoring was performed using thin layer chromatography or liquid chromatography-mass spectrometry. The developing agent systems used in thin layer chromatography were: dichloromethane and methanol system, petroleum ether and ethyl acetate system. The volume ratio of the solvents was adjusted according to the polarity of the compound, or a small amount of triethylamine was added for adjustment. Liquid chromatography-mass spectrometry was performed using a Waters ACQUITY Arc / ACQUITY QDa or a Thermo U3000-ISQ EC liquid chromatography-mass spectrometry instrument.

[0112] Column chromatography generally used silica gel with 200 - 300 mesh as the carrier. The eluent systems included: dichloromethane and methanol system, petroleum ether and ethyl acetate system. The volume ratio of the solvents was adjusted according to the polarity of the compound, or a small amount of triethylamine was added for adjustment.

[0113] Unless otherwise specified in the following examples, the reaction temperature was room temperature (20°C - 30°C), and the solvents were all dried and purified according to standard methods.

[0114] Example 1, Compound 1

[0115]

[0116] Under air protection, at 0 °C, isocyano trimethylsilane (37 mg, 0.3 mmol, 1.5 equiv) was added to a solution of (2E)-3-(3-aminocyclobutyl)-9,10-dimethoxy-2-[(2,4,6-trimethylphenyl)imino]-6H,7H-pyrimido[4,3-a]isoquinolin-4-one (100 mg, 0.2 mmol, 1 equiv) and TEA (0.1 mL, 0.6 mmol, 3 equiv) in DCM (2 mL). Under nitrogen protection, the resulting residue was stirred at room temperature for 1.5 h. The resulting residue was concentrated under reduced pressure. The crude product was purified by high performance liquid chromatography to obtain the product under the following conditions (column type: CHIRALPAK IG 2*25 cm, 5 μm; mobile phase A: ETOH:DCM = 1:1, mobile phase B: MtBE(0.1% DEA)-HPLC; flow rate: 20 mL / min; gradient: isocratic; wavelength: 220 nm; RT1(min): 11.5; RT2(min): 14.5; injection solvent: methanol; injection volume: 0.6 mL; number of runs: 4).

[0117] Two isomeric compounds were obtained: Compound 1-1 and Compound 1-2.

[0118] Compound 1-1 (isomer 1, RT1(min): 11.5, 10.36 mg, 9% yield)

[0119] LCMS(ESI): [M+H] + = 504.30

[0120] 1 H NMR(400 MHz, Methanol-d 4 ) δ 7.13(s, 2H), 6.93(s, 1H), 6.68(s, 1H), 5.45(s, 1H), 5.25–5.16(m, 1H), 4.14–4.07(m, 2H), 3.98–3.93(m, 1H), 3.89(s, 3H), 3.68(s, 3H), 2.96(t, J = 6.5 Hz, 2H), 2.38(s, 3H), 2.36–2.27(m, 4H), 2.16(s, 6H).

[0121] Compound 1-2 (isomer 2, RT2(min): 14.5, 40.48 mg, 36% yield)

[0122] LCMS(ESI): [M+H] + = 504.30

[0123] 1 H NMR(400 MHz, Methanol-d4 )δ 7.13 (s, 2H), 6.93 (s, 1H), 6.68 (s, 1H), 5.45 (s, 1H), 5.25–5.16 (m, 1H), 4.14–4.07 (m, 2H), 3.98–3.93 (m, 1H), 3.89 (s, 3H), 3.68 (s, 3H), 2.96 (t, J = 6.5 Hz, 2H), 2.38 (s, 3H), 2.36–2.27 (m, 4H), 2.16 (s, 6H).

[0124] Example 2, Compound 2

[0125]

[0126] Step 1.

[0127]

[0128] In a three-necked flask, potassium tert-butoxide (22.8 g, 203.4 mmol, 1.0 equiv) and n-pentane (200 mL) were added successively. Under nitrogen protection, the temperature was lowered to -30 °C, and at this temperature, 1,3-butadiene (toluene solution, 11.0 g, 203.4 mmol, 20%, 1.0 equiv) and tribromomethane (51.4 g, 203.4 mmol, 1.0 equiv) were added dropwise in sequence. After the addition was completed, the reaction was carried out at this temperature for 1.5 h, and then gradually returned to room temperature and reacted overnight. Water (30 mL) was added dropwise under an ice bath, and the mixture was extracted with n-pentane (3 x 100 mL). The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated under reduced pressure at a temperature not higher than 30 °C to obtain the crude product 1,1-dibromo-2-vinylcyclopropane (70.0 g, crude).

[0129] Step 2.

[0130]

[0131] 1,1-Dibromo-2-vinylcyclopropane (74.0 g, 327.6 mmol, 1.0 equiv), cetrimonium chloride (3.1 g, 9.8 mmol, 0.03 equiv), DCM (150 mL) and water (150 mL) were successively added to a single-necked flask. Concentrated sulfuric acid (106.0 g, 1080.9 mmol, 3.3 equiv) was added dropwise under an ice bath. After completion, potassium permanganate (155.3 g, 982.7 mmol, 3.0 equiv) was slowly added at a temperature below 5 °C. The mixture was gradually warmed to room temperature and reacted for 12 h. 50% aqueous sulfuric acid solution and anhydrous sodium sulfate (100.0 g) were added dropwise under an ice bath. The mixture was gradually warmed to room temperature and reacted for 10 minutes. The aqueous phase was extracted with DCM (3 x 500 mL). The combined organic phases were dried over anhydrous sodium sulfate and concentrated below 30 °C to obtain the crude product 2,2-dibromocyclopropane-1-carboxylic acid (40.0 g, crude).

[0132] Step 3.

[0133]

[0134] 2,2-Dibromocyclopropane-1-carboxylic acid (20.0 g, 81.3 mmol, 1.0 equiv) and diethyl ether (100 mL) were successively added to a three-necked flask. Methyllithium (60 mL, 90.3 mmol, 1.1 equiv, 1 M in THF) was added dropwise under an ice bath. The mixture was gradually warmed to room temperature and reacted for 2 h. Water (100 mL) was added dropwise under an ice bath. After liquid separation, the aqueous phase was adjusted to acidic pH = 6 with dilute hydrochloric acid (2 M) and extracted with DCM (3 x 100 mL). The combined organic phases were dried over anhydrous sodium sulfate and concentrated below 30 °C to obtain the crude product 2-bromocyclopropane-1-carboxylic acid (10.0 g, crude).

[0135] LCMS (ESI, m / z): [M+H] - = 162.9.

[0136] Step 4.

[0137]

[0138] 2-Bromocyclopropane-1-carboxylic acid (10.0 g, crude), potassium carbonate (7.5 g, 5.5 mmol, 3.0 equiv), DMF (20 mL) and iodomethane (6.0 g, 36.3 mmol, 1.0 equiv) were successively added to a single-necked flask. The reaction was carried out at room temperature for 5 h. Water (100 mL) was added. The mixture was extracted with DCM (3 x 50 mL). The combined organic phases were dried over anhydrous sodium sulfate and concentrated below 30 °C to obtain the crude product, 2-bromocyclopropane-1-carboxylate (20.0 g, crude).

[0139] Step 5.

[0140]

[0141] In a single-necked flask, 2-bromocyclopropane-1-carboxylate (200 mg, 1.1 mmol, 1.0 equiv), (2E)-9,10-dimethoxy-2-[(2,4,6-trimethylphenyl)imino]-3H,6H,7H-pyrimido[4,3-a]isoquinolin-4-one (437 mg, 1.1 mmol, 1.0 equiv), cesium carbonate (1.1 g, 3.4 mmol, 3.0 equiv), Xantphos (106.5 mg, 0.2 mmol, 0.2 equiv) and 1,4-dioxane (80 mL) were successively added. Pd 2 (dba) 3 (64.2 mg, 0.1 mmol, 0.1 equiv) was added under nitrogen protection, and the reaction was carried out overnight at 100 °C. The solvent was evaporated to dryness, and the crude product was purified by normal phase using MeOH / DCM (1 / 10) as the mobile phase. After reverse (column type (YMC Triart C18 ExRs 5 μm, 30 mm * 150 mm; mobile phase A: water (10 mmol / L NH 4 HCO 3 ), mobile phase B: acetonitrile; flow rate: 60 mL / min; gradient: 39% B to 62% B in 10 min; wavelength: 254 nm / 220 nm; RT1 (min): 27)), methyl 2-[(2E)-9,10-dimethoxy-4-oxo-2-[(2,4,6-trimethylphenyl)imino]-6H,7H-pyrimidino[4,3-a]isoquinolin-3-yl]cyclopropane-1-carboxylate (100 mg, 18.3% yield) was further purified.

[0142] LCMS (ESI, m / z): [M + H] + = 490.2.

[0143] Step 6.

[0144]

[0145] 2-[(2E)-9,10-dimethoxy-4-oxo-2-[(2,4,6-trimethylphenyl)imino]-6H,7H-pyrimido[4,3-a]isoquinolin-3-yl]cyclopropane-1-carboxylic acid methyl ester (500 mg, 1.0 mmol, 1.0 equiv), methanol (10 mL), and an aqueous solution (2 mL) of lithium hydroxide (85 mg, 2.0 mmol, 2.0 equiv) were successively added to a single-necked flask, and the reaction was carried out at room temperature for 2 h. The solvent was evaporated, the pH was adjusted to 6 with 4 M hydrochloric acid solution, and the mixture was extracted with DCM (3 x 50 mL). The combined organic phases were dried over anhydrous sodium sulfate, and the organic phase was evaporated to obtain 2-[(2E)-9,10-dimethoxy-4-oxo-2-[(2,4,6-trimethylphenyl)imino]-6H,7H-pyrimido[4,3-a]isoquinolin-3-yl]cyclopropane-1-carboxylic acid (450 mg, 92.7% yield).

[0146] LCMS (ESI, m / z): [M+H] + = 476.2.

[0147] Step 7.

[0148]

[0149] 2-[(2E)-9,10-dimethoxy-4-oxo-2-[(2,4,6-trimethylphenyl)imino]-6H,7H-pyrimido[4,3-a]isoquinolin-3-yl]cyclopropane-1-carboxylic acid (500 mg, 1.1 mmol, 1.0 equiv), 1,4-dioxane (156 mL), triethylamine (319 mg, 3.2 mmol, 3.0 equiv), and DPPA (347.2 mg, 1.3 mmol, 1.2 equiv) were successively added to a three-necked flask under nitrogen protection, and the reaction was carried out at room temperature for 2 h. Then, the temperature was raised to 100 °C and the reaction was continued for 1 h. After cooling to room temperature, a tetrahydrofuran solution of ammonia (1.4 M) (100 mL) was added, and the reaction was carried out at room temperature for 2 h. The solvent was evaporated, and HPLC (column type: YMC Triart C18 ExRs 5 μm, 30 mm * 150 mm; mobile phase A: water (10 mmol / L NH 4 HCO 3 ), mobile phase B: acetonitrile; flow rate: 60 mL / min; gradient: 39% B to 62% B in 10 min; wavelength: 254 nm / 220 nm; RT1 (min): 27) was used for purification to obtain 2-[(2E)-9,10-dimethoxy-4-oxo-2-[(2,4,6-trimethylphenyl)imino]-6H,7H-pyrimido[4,3-a]isoquinolin-3-yl]cyclopropylurea (120 mg, 20.0% yield).

[0150] LCMS (ESI, m / z): [M+H] + = 490.2

[0151] Step 8.

[0152]

[0153] The crude product 2-[(2E)-9,10-dimethoxy-4-oxo-2-[(2,4,6-trimethylphenyl)imino]-6H,7H-pyrimido[4,3-a]isoquinolin-3-yl]cyclopropylurea (120 mg) was resolved by Chiral-HPLC (Column type: CHIRAL ARTCellulose-SB, 2 * 25 cm, 5 μm; Mobile phase A: DCM-HPLC, Mobile phase B: IPA (0.2% DEA); Flow rate: 16 mL / min; Gradient: isocratic 50; Wavelength: 220 nm; RT1 (min): 4.9; RT2 (min): 7.0; Sample Solvent: MeOH:DCM = 1:1-HPLC; Injection volume: 0.6 mL; Number Of Runs: 5) to obtain two isomeric compounds of the product: Compound 2-1, Compound 2-2.

[0154] Compound 2-1 (isomer 1, RT1 (min): 4.9, 52.0 mg, 47.6% yield):

[0155] LCMS (ESI, m / z): [M+H] + = 490.45

[0156] 1 1H NMR (400 MHz, Methanol-d 4 ) δ 7.12 (s, 1H), 7.07 (s, 1H), 6.94 (s, 1H), 6.68 (s, 1H), 5.47 (s, 1H), 4.19–4.13 (m, 1H), 4.10–4.05 (m, 1H), 3.89 (s, 3H), 3.67 (s, 3H), 2.99–2.93 (m, 3H), 2.84 (s, 1H), 2.37 (s, 3H), 2.22 (s, 3H), 2.02 (s, 3H), 0.99–0.92 (m, 1H), 0.75–0.72 (m, 1H).

[0157] Compound 2-2 (isomer 2, RT2 (min): 7.0, 51.7 mg, 47.6% yield):

[0158] LCMS (ESI, m / z): [M+H] += 490.45

[0159] 1 H NMR(400 MHz, Methanol-d 4 ) δ 7.12 (s, 1H), 7.07 (s, 1H), 6.94 (s, 1H), 6.68 (s, 1H), 5.47 (s, 1H), 4.19–4.13 (m, 1H), 4.10–4.05 (m, 1H), 3.89 (s, 3H), 3.67 (s, 3H), 2.99–2.93 (m, 3H), 2.84 (s, 1H), 2.37 (s, 3H), 2.22 (s, 3H), 2.02 (s, 3H), 0.99–0.92 (m, 1H), 0.75–0.72 (m, 1H).

[0160] Example 3, Compound 3

[0161]

[0162] Step 1.

[0163]

[0164] In a single-necked flask, N-(4-hydroxycyclohexyl)carbamic acid tert-butyl ester (3.0 g, 10.9 mmol, 1.0 equivalent), DCM (80 mL), p-toluenesulfonyl chloride (5.0 g, 13.2 mmol, 1.2 equivalents), triethylamine (3.3 g, 32.9 mmol, 3.0 equivalents) and DMAP (134 mg, 1.1 mmol, 0.1 equivalent) were added successively, and the reaction was carried out overnight at room temperature. The solvent was evaporated to dryness, EA (300 mL) was added, washed with saturated sodium bicarbonate solution (3 x 100 mL), and the organic phase was dried over anhydrous sodium sulfate. The organic phase was evaporated to dryness to obtain (N-{4-[(4-methylbenzenesulfonyl)oxy]cyclohexyl}carbamic acid tert-butyl ester) (5.0 g, 61.7%)

[0165] LCMS (ESI, m / z): [M+H] + = 370.2

[0166] Step 2.

[0167]

[0168] In a single-necked flask, (2E)-9,10-dimethoxy-2-[(2,4,6-trimethylphenyl)imino]-3H,6H,7H-pyrimido[4,3-a]isoquinolin-4-one (1.0 g, 2.6 mmol, 1.0 equiv), dioxane (100 mL), cesium carbonate (1.7 g, 5.1 mmol, 2.0 equiv), and tert-butyl (N-{4-[(4-methylbenzenesulfonyl)oxy]cyclohexyl}carbamate) (1.4 g, 3.8 mmol, 1.5 equiv) were added successively, and the reaction was carried out overnight at 100 °C. The reaction mixture was filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by reverse-phase column chromatography under the following conditions: column specification C18, mobile phase, water and acetonitrile, 10% to 1000% gradient for 30 minutes, UV detector at 220 nm. tert-Butyl (N-{4-[(2E)-9,10-dimethoxy-4-oxo-2-[(2,4,6-trimethylphenyl)imino]-6H,7H-pyrimido[4,3-a]isoquinolin-3-yl]cyclohexyl}carbamate) (110 mg, 7.3%) was obtained.

[0169] LCMS(ESI,m / z): [M+H] + = 589.3

[0170] Step 3.

[0171]

[0172] In a single-necked flask, tert-butyl (N-{4-[(2E)-9,10-dimethoxy-4-oxo-2-[(2,4,6-trimethylphenyl)imino]-6H,7H-pyrimido[4,3-a]isoquinolin-3-yl]cyclohexyl}carbamate) (100 mg, 0.2 mmol, 1.0 equiv) and a 1,4-dioxane solution of HCl (4 M, 15 mL) were added successively, and the reaction was carried out at room temperature for 2 h. The solvent was concentrated under reduced pressure to obtain (2E)-3-(4-aminocyclohexyl)-9,10-dimethoxy-2-[(2,4,6-trimethylphenyl)imino]-6H,7H-pyrimido[4,3-a]isoquinolin-4-one (80 mg, 96.4%).

[0173] LCMS(ESI,m / z): [M+H] + = 489.3

[0174] Step 4.

[0175]

[0176] (2E)-3-(4-Aminocyclohexyl)-9,10-dimethoxy-2-[(2,4,6-trimethylphenyl)imino]-6H,7H-pyrimido[4,3-a]isoquinolin-4-one (140 mg, 0.3 mmol, 1.0 equiv) was dissolved in dichloromethane (8 mL) as the solvent. Triethylamine (145 mg, 1.5 mmol, 5.0 equiv) and trimethylsilyl isocyanate (40 mg, 0.4 mmol, 1.2 equiv) were added to the above mixture at room temperature, and the mixture was stirred at room temperature for 3 h. The crude product was purified by Prep-HPLC under the following conditions (column: Xbridge BEH Shield RP18, 5 μm, 19 * 250 mm; mobile phase A: water (10 mmol / L ammonium bicarbonate), mobile phase B: acetonitrile; flow rate: 25 mL / min; gradient: from 50% B to 57% B within 10 minutes; wavelength: 254 nm / 220 nm; RT1 (min): 8.93), to obtain 4-[(2E)-9,10-dimethoxy-4-oxo-2-[(2,4,6-trimethylphenyl)imino]-6H,7H-pyrimido[4,3-a]isoquinolin-3-yl]cyclohexylurea (32.52 mg, yield 21.24%).

[0177] LCMS (ESI): [M+H] + = 532.30

[0178] 1 1H NMR (400 MHz, Methanol-d 4 ) δ 6.93 (s, 2H), 6.88 (s, 1H), 6.74 (s, 1H), 5.50 (s, 1H), 5.36 (s, 1H), 4.03–3.96 (m, 2H), 3.93 (s, 1H), 3.87 (s, 3H), 3.69 (s, 3H), 2.95 (t, J = 6.2 Hz, 2H), 2.79 (s, 2H), 2.29 (s, 3H), 2.05 (s, 6H), 1.98-1.90 (m, 2H), 1.80–1.70 (m, 2H), 1.68–1.62 (m, 2H).

[0179] Example 4, Compound 4

[0180]

[0181] Step 1.

[0182]

[0183] In a single-necked flask, sequentially add tert-butyl 4-hydroxypiperidine-1-carboxylate (3.0 g, 14.9 mmol, 1.0 equiv), p-toluenesulfonyl chloride (3.4 g, 17.9 mmol, 1.2 equiv), dichloromethane (100 mL), triethylamine (4.5 g, 44.7 mmol, 3.0 equiv) and 4-dimethylaminopyridine (182 mg, 1.5 mmol, 0.1 equiv), and react overnight at room temperature. Concentrate the solvent, add ethyl acetate (300 mL), wash with saturated sodium bicarbonate solution (3 x 100 mL), dry the organic phase with anhydrous sodium sulfate, and concentrate the organic phase to obtain tert-butyl 4-[(4-methylbenzenesulfonyl)oxy]piperidine-1-carboxylate (4.3 g, yield 81.2%).

[0184] LCMS(ESI, m / z): [M+H] + = 356.1

[0185] Step 2.

[0186]

[0187] In a single-necked flask, sequentially add (2E)-9,10-dimethoxy-2-[(2,4,6-trimethylphenyl)imino]-3H,6H,7H-pyrimido[4,3-a]isoquinolin-4-one (1.0 g, 2.6 mmol, 1.0 equiv), 1,4-dioxane (100 mL), cesium carbonate (2.5 g, 7.7 mmol, 3.0 equiv) and tert-butyl 4-[(4-methylbenzenesulfonyl)oxy]piperidine-1-carboxylate (1.0 g, 2.8 mmol, 1.1 equiv), and react overnight at 100 °C. Filter, concentrate the filtrate, and purify the obtained residue by reverse-phase column chromatography under the following conditions: mobile phase, water and acetonitrile, 10% to 100% gradient for 30 minutes, UV detector at 220 nm. Obtain tert-butyl 4-[(2E)-9,10-dimethoxy-4-oxo-2-[(2,4,6-trimethylphenyl)imino]-6H,7H-pyrimido[4,3-a]isoquinolin-3-yl]piperidine-1-carboxylate (120 mg, yield 8.17%).

[0188] LCMS(ESI, m / z): [M+H] + = 575.3

[0189] Step 3.

[0190]

[0191] In a single-necked flask, tert-butyl 4-[(2E)-9,10-dimethoxy-4-oxo-2-[(2,4,6-trimethylphenyl)imino]-6H,7H-pyrimido[4,3-a]isoquinolin-3-yl]piperidine-1-carboxylate (100 mg, 0.2 mmol, 1.0 equiv) and a 1,4-dioxane solution of hydrogen chloride (2 N, 17 mL) were added successively, and the reaction was carried out at room temperature for 2 h. The solvent was concentrated to obtain (2E)-9,10-dimethoxy-3-(piperidin-4-yl)-2-[(2,4,6-trimethylphenyl)imino]-6H,7H-pyrimido[4,3-a]isoquinolin-4-one (80 mg, yield 96.9%).

[0192] LCMS(ESI,m / z): [M+H] + = 475.3

[0193] Step 4.

[0194]

[0195] To a solution of (2E)-9,10-dimethoxy-3-(piperidin-4-yl)-2-[(2,4,6-trimethylphenyl)imino]-6H,7H-pyrimido[4,3-a]isoquinolin-4-one (78 mg, 0.2 mmol, 1.0 equiv) in dichloromethane (2 mL) was added triethylamine (50 mg, 0.5 mmol, 3.0 equiv) and trimethylsilyl isocyanate (28 mg, 0.3 mmol, 1.5 equiv). The mixture was stirred at room temperature for 2 h. After the mixture was concentrated by rotary evaporation under reduced pressure, the resulting residue was purified by reverse-phase column chromatography under the following conditions: column: XBridge BEH Shield RP18, 5 μm, 19*250 mm; mobile phase A: water (10 mmol / L ammonium bicarbonate), mobile phase B: acetonitrile; flow rate: 25 mL / min; gradient: 46% B to 55% B in 10 min; wavelength: 254 nm / 220 nm; RT1(min): 8.57, to obtain 4-[(2E)-9,10-dimethoxy-4-oxo-2-[(2,4,6-trimethylphenyl)imino]-6H,7H-pyrimido[4,3-a]isoquinolin-3-yl]piperidine-1-carboxamide (34.40 mg, yield 40.36%).

[0196] LCMS(ESI): [M+H] + = 518.30

[0197] 1 H NMR(400MHz,Methanol-d 4)δ 6.89 (s, 2H), 6.85 (s, 1H), 6.72 (s, 1H), 5.50 (s, 1H), 5.46 (s, 1H), 4.19–4.13 (m, 2H), 3.94 (t, J = 6.1 Hz, 2H), 3.85 (s, 3H), 3.66 (s, 3H), 2.99–2.69 (m, 6H), 2.26 (s, 3H), 2.02 (s, 6H), 1.75–1.68 (m, 2H).

[0198] Example 5, Compound 5

[0199]

[0200] Step 1.

[0201]

[0202] (2E)-9,10-Dimethoxy-2-[(2,4,6-trimethylphenyl)imino]-3H,6H,7H-pyrimido[4,3-a]isoquinolin-4-one (5 g, 12.7 mmol, 1 equiv) and tert-butyl 3-bromoazetidine-1-carboxylate (3.62 g, 15.3 mmol, 1.2 equiv) were stirred in DMF (50 mL) and stirred overnight under a nitrogen atmosphere at 100 °C. The resulting mixture was concentrated under reduced pressure. It was purified by silica gel column chromatography and eluted with CH 2 Cl 2 / MeOH (10 / 1) to give tert-butyl 3-[(2E)-9,10-dimethoxy-4-oxo-2-[(2,4,6-trimethylphenyl)imino]-6H,7H-pyrimido[4,3-a]isoquinolin-3-yl]azetidine-1-carboxylate (1.3 g, 18.6%).

[0203] Step 2.

[0204]

[0205] tert-Butyl 3-[(2E)-9,10-dimethoxy-4-oxo-2-[(2,4,6-trimethylphenyl)imino]-6H,7H-pyrimido[4,3-a]isoquinolin-3-yl]azetidine-1-carboxylate (1.0 g, 1.8 mmol, 1.0 equiv) was stirred in 1,4-dioxane (20 mL) under a nitrogen atmosphere at room temperature for 1 h. The resulting mixture was concentrated under reduced pressure. As a result, (2E)-3-(azetidin-3-yl)-9,10-dimethoxy-2-[(2,4,6-trimethylphenyl)imino]-6H,7H-pyrimido[4,3-a]isoquinolin-4-one (0.7 g, 85.6%) was obtained. The crude product was directly used in the next step without further purification.

[0206] LCMS(ESI): [M+H] + = 447.25

[0207] Step 3.

[0208]

[0209] (2E)-3-(azetidin-3-yl)-9,10-dimethoxy-2-[(2,4,6-trimethylphenyl)imino]-6H,7H-pyrimido[4,3-a]isoquinolin-4-one (50 mg, 0.112 mmol, 1.0 eq) and trimethylsilyl isocyanate (14.19 mg, 0.12 mmol, 1.1 eq) were stirred in DCM (6 mL) overnight under nitrogen atmosphere at room temperature. The resulting mixture was concentrated under vacuum. Column type: Xbridge BEH Shield RP18, 5 μm, 19*250 mm; Mobile phase A: water (10 mmol / L NH 4 HCO 3 ), Mobile phase B: ACN; Flow rate: 25 mL / min; Gradient: 40% B to 48% B, 10 min; Wavelength: 254 nm / 220 nm; RT1 (min): 9.15 to give 3-[(2E)-9,10-dimethoxy-4-oxo-2-[(2,4,6-trimethylphenyl)imino]-6H,7H-pyrimido[4,3-a]isoquinolin-3-yl]azetidine-1-carboxamide (14.69 mg, 25.59%).

[0210] LCMS(ESI): [M+H] + = 490.25

[0211] 1 1H-NMR (400 MHz, Methanol-d 4 ) δ 7.23 (s, 1H), 7.13 (d, J = 3.9 Hz, 2H), 7.03 (s, 1H), 6.00 (s, 1H), 5.15–5.08 (m, 1H), 4.39–4.32 (m, 1H), 4.28–4.20 (m, 2H), 4.22–4.19 (m, 1H), 4.19–4.15 (m, 1H), 3.95 (s, 3H), 3.79 (s, 3H), 3.62–3.56 (m, 1H), 3.18–3.05 (m, 2H), 2.38 (s, 3H), 2.35 (s, 3H), 2.27 (s, 3H).

[0212] Example 6, Compound 6

[0213]

[0214] Step 1.

[0215]

[0216] To a stirred solution of (2E)-9,10-dimethoxy-2-[(2,4,6-trimethylphenyl)imino]-3H,6H,7H-pyrimido[4,3-a]isoquinolin-4-one (1.4 g, 3.5 mmol, 1 equiv) and Cs 2 CO 3 (2.3 g, 7.1 mmol, 2 equiv) in 1,4-dioxane (15 mL) was added tert-butyl N-(3-bromocyclobutyl)carbamate (1.3 g, 5.3 mmol, 1.5 equiv). The resulting mixture was stirred at 100 °C under nitrogen overnight. The filtrate was concentrated under reduced pressure and the residue was purified by reverse-phase flash chromatography under the following conditions: column, C18 silica gel; mobile phase, acetonitrile-water (10 mmol / L NH 4 HCO 3 ), 50%-58% gradient, 10 min; detector, UV 254 nm. tert-Butyl N-{3-[(2E)-9,10-dimethoxy-4-oxo-2-[(2,4,6-trimethylphenyl)imino]-6H,7H-pyrimido[4,3-a]isoquinolin-3-yl]cyclobutyl}carbamate (700 mg, 32% yield) was obtained.

[0217] LCMS (ESI): [M+H] + = 561.2

[0218] Step 2.

[0219]

[0220] tert-Butyl N-{3-[(2E)-9,10-dimethoxy-4-oxo-2-[(2,4,6-trimethylphenyl)imino]-6H,7H-pyrimido[4,3-a]isoquinolin-3-yl]cyclobutyl}carbamate ((700 mg, 1.2 mmol, 1 equiv) was added to a 1,4-dioxane solution of hydrochloric acid (10 ml, 4 M) and the resulting mixture was stirred at room temperature under nitrogen for 1.5 h. The resulting mixture was concentrated under reduced pressure. (2E)-3-(3-Aminocyclobutyl)-9,10-dimethoxy-2-[(2,4,6-trimethylphenyl)imino]-6H,7H-pyrimido[4,3-a]isoquinolin-4-one (800 mg, crude) was obtained.

[0221] LCMS (ESI): [M+H] + = 461.2

[0222] Step 3.

[0223]

[0224] Under a nitrogen atmosphere at 0 °C, isocyano trimethylsilane (300 mg, 2.6 mmol, 1.5 equiv) was added to a solution of (2E)-3-(3-aminocyclobutyl)-9,10-dimethoxy-2-[(2,4,6-trimethylphenyl)imino]-6H,7H-pyrimido[4,3-a]isoquinolin-4-one (800 mg, 1.7 mmol, 1 equiv) and TEA (0.7 mL, 5.2 mmol, 3 equiv) in DCM (10 mL). The resulting mixture was stirred overnight at room temperature under a nitrogen atmosphere. The residue was purified by silica gel column chromatography, eluting with dichloromethane / methanol (10 / 1) to give 3-[(2E)-9,10-dimethoxy-4-oxo-2-[(2,4,6-trimethylphenyl)imino]-6H,7H-pyrimido[4,3-a]isoquinolin-3-yl]cyclobutylurea (400 mg, 42% yield).

[0225] LCMS(ESI): [M+H] + = 504.2

[0226] Step 4.

[0227]

[0228] Under air protection at room temperature, aqueous HBr solution (3 mL) was added to a solution of 3-[(2E)-9,10-dimethoxy-4-oxo-2-[(2,4,6-trimethylphenyl)imino]-6H,7H-pyrimido[4,3-a]isoquinolin-3-yl]cyclobutylurea (400 mg, 0.7 mmol, 1 equiv) and acetic anhydride (3 mL) in acetic acid (3 mL). Under nitrogen protection, the resulting residue was stirred at 120 °C for 2 h. The reaction mixture was quenched with water at room temperature. The reaction mixture was extracted with dichloromethane / methanol (10 / 1) (2 X 30 mL). The combined organic phases were washed with saturated brine (2 X 100 mL), dried over anhydrous sodium sulfate. After filtration of the resulting mixture, the filtrate was concentrated under reduced pressure. 3-[(2E)-9,10-Dihydroxy-4-oxo-2-[(2,4,6-trimethylphenyl)imino]-6H,7H-pyrimido[4,3-a]isoquinolin-3-yl]cyclobutylurea (120 mg, crude) was obtained.

[0229] LCMS(ESI): [M+H] + = 476.2

[0230] Step 5.

[0231]

[0232] Under air protection, at room temperature, to a solution of 3-[(2E)-9,10-dihydroxy-4-oxo-2-[(2,4,6-trimethylphenyl)imino]-6H,7H-pyrimido[4,3-a]isoquinolin-3-yl]cyclobutylurea (100 mg, 0.2 mmol, 1 equiv) and potassium carbonate (87 mg, 0.6 mmol, 3 equiv) in DMF (1 mL), CD 3 I (76 mg, 0.5 mmol, 2.5 equiv) was added. Under nitrogen protection, at room temperature, the resulting residue was stirred overnight. The reaction mixture was quenched with water at room temperature. The reaction mixture was extracted with dichloromethane (2 X 30 mL). The combined organic phases were backwashed with saturated brine (2 X 50 mL) and dried over sodium sulfate. After the resulting mixture was filtered, the filtrate was concentrated under reduced pressure. The crude product was purified by chiral-SFC to obtain the product under the following conditions (column type: CHIRALPAK IH 2*25 cm, 5 μm; mobile phase A: MeOH:DCM = 1:1 - HPLC, mobile phase B: MtBE (0.1% DEA)-HPLC--; flow rate: 20 mL / min; gradient: isocratic; wavelength: 220 nm; RT1 (min): 13.9; RT2 (min): 17.2; solvent: methanol; injection volume: 0.5 mL; number of runs: 4).

[0233] Two isomeric compounds were obtained: Compound 6-1 and Compound 6-2.

[0234] Compound 6-1 (isomer 1, RT1 (min): 13.9, 25.91 mg, 24% yield):

[0235] LCMS (ESI): [M+H] + = 510.30

[0236] 1 H NMR (400 MHz, Methanol-d 4 ) δ 7.12 (s, 2H), 6.92 (s, 1H), 6.66 (s, 1H), 5.43 (s, 1H), 4.66–4.60 (m, 1H), 4.09 (t, J = 6.4 Hz, 2H), 3.89 (d, J = 5.5 Hz, 1H), 2.96 (t, J = 6.5 Hz, 2H), 2.82–2.72 (m, 2H), 2.37 (s, 3H), 2.16 (s, 6H), 1.96–1.90 (m, 2H).

[0237] Compound 6-2 (isomer 2, RT2 (min): 17.2, 9.41 mg, 8.5% yield):

[0238] LCMS(ESI): [M+H] + = 510.2

[0239] 1 H NMR(400 MHz, Methanol-d 4 ) δ 7.12 (s, 2H), 6.92 (s, 1H), 6.66 (s, 1H), 5.43 (s, 1H), 4.66–4.60 (m, 1H), 4.09 (t, J = 6.4 Hz, 2H), 3.89 (d, J = 5.5 Hz, 1H), 2.96 (t, J = 6.5 Hz, 2H), 2.82–2.72 (m, 2H), 2.37 (s, 3H), 2.16 (s, 6H), 1.96–1.88 (m, 2H).

[0240] Example 7, Compound 7

[0241]

[0242] Under air protection, at room temperature, to a solution of (E)-1-(3-(9,10-dihydroxy-2-(methyltrimethylimine)-4-oxo-6,7-dihydro-2H-pyrimido[6,1-a]isoquinolin-3(4H)-yl)cyclobutyl)urea (100 mg, 0.2 mmol, 1 equiv) and K 2 CO 3 (87 mg, 0.6 mmol, 3 equiv) in DMSO (2 mL), sodium 2-chloro-2,2-difluoroacetate (80 mg, 0.5 mmol, 2.5 equiv) was added. Under nitrogen protection, at room temperature, the resulting residue was stirred overnight. The reaction mixture was quenched with 50 mL of water at room temperature. The reaction mixture was extracted with dichloromethane (2X 30 ml). The combined organic phases were backwashed with saturated brine (2X 50 ml) and dried over sodium sulfate. After filtration of the resulting mixture, the filtrate was concentrated under reduced pressure. The crude product was purified by high performance liquid chromatography to give 3-[(2E)-9,10-bis(difluoromethoxy)-4-oxo-2-[(2,4,6-trimethylphenyl)imino]-6H,7H-pyrimido[4,3-a]isoquinolin-3-yl]cyclobutylurea (6.77 mg, 5.3% yield) under the following conditions (column type: CHIRALPAK IH 2*25 cm, 5 μm; mobile phase A: MeOH:DCM = 1:1 - HPLC, mobile phase B: MtBE (0.1% DEA)-HPLC--; flow rate: 20 mL / min; gradient: isocratic; wavelength: 220 nm; RT1 (min): 13.9; RT2 (min): 17.2; solvent: methanol; injection volume: 0.5 mL; number of runs: 4)

[0243] LCMS(ESI): [M+H]+ = 576.25

[0244] 1 H NMR (400 MHz, Methanol-d 4 ) δ 7.31 (s, 1H), 7.13 (d, J = 2.3 Hz, 3H), 6.95–6.55 (m, 2H), 5.51 (d, J = 7.1 Hz, 1H), 4.66–4.60 (m, 1H), 4.13 (t, J = 6.4 Hz, 2H), 4.02–3.88 (m, 1H), 3.04 (t, J = 6.4 Hz, 2H), 2.85–2.70 (m, 1H), 2.50–2.15 (m, 5H), 2.16 (d, J = 2.5 Hz, 6H), 1.96–1.90 (m, 1H).

[0245] Example 8, Compound 8

[0246]

[0247] Step 1.

[0248]

[0249] 2-[(2,6-Diisopropyl-4-methylphenyl)amino]-9,10-dimethoxy-6H,7H-pyrimido[4,3-a]isoquinolin-4-one (500 mg, 1.1 mmol, 1 equiv), cesium carbonate (1.3 g, 3.3 mmol, 3 equiv) and tert-butyl (3-bromocyclobutyl)carbamate (560 mg, 2.3 mmol, 1.5 equiv) were dissolved in 1,4-dioxane (10 mL), protected by nitrogen, and stirred at 100 °C overnight. Cooled to room temperature, dissolved in 20 mL of water, extracted with ethyl acetate (3 × 10 mL), the organic layer was washed with 20 mL of saturated sodium chloride, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by reverse-phase column chromatography with the following conditions (column: C18; mobile phase A: water (10 mmol / L ammonium bicarbonate), mobile phase B: acetonitrile; flow rate: 60 mL / min; gradient: 50% B to 80% B in 20 min; wavelength: 254 nm;) to obtain tert-butyl (E)-(3-(2-((2,6-diisopropyl-4-methylphenyl)imino)-9,10-dimethoxy-4-oxo-6,7-dihydro-2H-pyrimido[6,1-a]isoquinolin-3(4H)-yl)cyclobutyl)carbamate (300 mg, yield 43.5%).

[0250] LCMS (ESI): [M+H] + = 617.55

[0251] Step 2.

[0252]

[0253] Dissolve tert-butyl (E)-(3-(2-((2,6-diisopropyl-4-methylphenyl)imino)-9,10-dimethoxy-4-oxo-6,7-dihydro-2H-pyrimido[6,1-a]isoquinolin-3(4H)-yl)cyclobutyl)carbamate (300 mg, 0.49 mmol, 1 equiv) in a 1,4-dioxane solution of 10 mL of hydrochloric acid and stir at room temperature for 1 hour. Concentrate under reduced pressure, dissolve in 10 mL of water, extract with ethyl acetate (3 × 10 mL), wash the organic layer with 20 mL of saturated sodium chloride, and anhydrous Na 2 SO 4 Dry, filter, and concentrate under reduced pressure to obtain 2-((1-aminopropan-2-yl)(2,6-diisopropyl-4-methylphenyl)amino)-9,10-dimethoxy-6,7-dihydro-4H-pyrimido[6,1-a]isoquinolin-4-one (300 mg, crude).

[0254] LCMS(ESI): [M+H] + = 517.55

[0255] Step 3.

[0256]

[0257] Dissolve 2-((1-aminopropan-2-yl)(2,6-diisopropyl-4-methylphenyl)amino)-9,10-dimethoxy-6,7-dihydro-4H-pyrimido[6,1-a]isoquinolin-4-one (300 mg, 0.59 mmol, 1.0 equiv.), triethylamine (180 mg, 1.8 mmol, 3 equiv) and trimethylsilyl isocyanate (103 mg, 1.2 mmol, 2 equiv.) in 10 mL of DCM and stir at room temperature for 2 hours. Concentrate under reduced pressure, dissolve in 10 mL of water, extract with ethyl acetate (3 × 10 mL), wash the organic layer with 20 mL of saturated sodium chloride, and anhydrous Na 2 SO 4Dry, filter, and concentrate under reduced pressure. The crude product (150 mg) was purified by Chiral-SFC under the following conditions (column: CHIRALPAK IH, 2 * 25 cm, 5 μm; mobile phase A: methanol:dichloromethane = 1:1 - HPLC, mobile phase B: methyl tert-butyl ether (0.1% ethylenediamine) - HPLC; flow rate: 20 mL / min; gradient: isocratic; wavelength: 220 nm; RT1 (min): 8; RT2 (min): 13.0; sample solvent: methanol - HPLC; injection volume: 1 mL) to obtain (E)-1-(3-(2-((2,6-diisopropyl-4-methylphenyl)imino)-9,10-dimethoxy-4-oxo-6,7-dihydro-2H-pyrimido[6,1-a]isoquinolin-3(4H)-yl)cyclobutyl)urea (33.18 mg, yield 14.96%).

[0258] LCMS(ESI): [M + H] + = 560.30

[0259] 1 H NMR(400 MHz, Methanol-d 4 ) δ 7.21 (s, 2H), 6.93 (s, 1H), 6.64 (s, 1H), 5.46 (s, 1H), 4.49–4.44 (m, 1H), 4.14–4.06 (m, 2H), 3.97–3.88 (m, 1H), 3.89 (s, 3H), 3.64 (s, 3H), 2.94 - 2.90 (m, 4H), 2.74 - 2.78 (m, 2H), 2.44 (s, 3H), 2.20–2.08 (m, 2H), 1.30 (d, J = 6.8 Hz, 7H), 1.11 (d, J = 6.8 Hz, 6H).

[0260] Example 9, Compound 9

[0261]

[0262] Step 1.

[0263]

[0264] (2E)-2-[(2,6-Dimethoxy-4-methylphenyl)imino]-9,10-dimethoxy-3H,6H,7H-pyrimido[4,3-a]isoquinolin-4-one (500 mg, 1.2 mmol, 1 equiv) and tert-butyl N-(3-bromocyclobutyl)carbamate (354 mg, 1.4 mmol, 1.2 equiv) were added to DMF (1 mL), and cesium carbonate ((769 mg, 2.3 mmol, 2 equiv) was added. The resulting mixture was stirred overnight under nitrogen at 100 °C. The filter cake was filtered and washed with ethyl acetate (2 x 100 mL). The filtrate was concentrated under reduced pressure, and the residue was purified by reverse-phase flash chromatography under the following conditions: column, C18 silica gel; mobile phase, acetonitrile in water (10 mmol / L NH 4 HCO 3 ), 45%-56% gradient, 10 min; detector, UV 254 nm. As a result, tert-butyl N-{3-[(2E)-2-[(2,6-dimethoxy-4-methylphenyl)imino]-9,10-dimethoxy-4-oxo-6H,7H-pyrimido[4,3-a]isoquinolin-3-yl]cyclobutyl}carbamate (400 mg, 45% yield) was obtained.

[0265] LCMS (ESI): [M+H] + = 593.2

[0266] Step 2.

[0267]

[0268] Tert-butyl N-{3-[(2E)-2-[(2,6-dimethoxy-4-methylphenyl)imino]-9,10-dimethoxy-4-oxo-6H,7H-pyrimido[4,3-a]isoquinolin-3-yl]cyclobutyl}carbamate (400 mg, 0.7 mmol, 1 equiv) was added to a solution of hydrogen chloride gas in 1,4-dioxane (4 mL, 4 M). The resulting mixture was stirred for 1.5 h under nitrogen at room temperature. The resulting mixture was concentrated under reduced pressure. The residue was purified by Prep-TLC (CH 2 Cl 2 / MeOH = 10 / 1) to give (2E)-3-(3-aminocyclobutyl)-2-[(2,6-dimethoxy-4-methylphenyl)imino]-9,10-dimethoxy-6H,7H-pyrimido[4,3-a]isoquinolin-4-one (130 mg, 37% yield).

[0269] LCMS (ESI): [M+H] + = 493.2

[0270] Step 3.

[0271]

[0272] Under a nitrogen atmosphere at 0 °C, (2E)-3-(3-aminocyclobutyl)-2-[(2,6-dimethoxy-4-methylphenyl)imino]-9,10-dimethoxy-6H,7H-pyrimido[4,3-a]isoquinolin-4-one (100 mg, 0.2 mmol, 1 equiv) and triethylamine (0.1 mL, 0.6 mmol, 3 equiv) were added to a solution of DCM (2 mL), and then isocyano trimethylsilane (35 mg, 0.3 mmol, 1.5 equiv) was added. The resulting mixture was stirred overnight at room temperature under a nitrogen atmosphere. The crude product was purified by high performance liquid chromatography under the following conditions (column: XBridge BEH C18 5.0 mmol: XBridge BEH C18 5 μm, 30 * 150 mm; mobile phase A: water (10 mmol / L NH 4 HCO 3 ), mobile phase B: acetonitrile; flow rate: 60 mL / min; column temperature: 25 °C: flow rate: 60 mL / min; gradient: from 26% B to 37% B in 7 minutes; wavelength: 254 nm / 220 nm; RT1 (min): 6.37; RT2 (min): 7.05, to obtain two isomeric compounds: Compound 9-1 and Compound 9-2.

[0273] Compound 9-1 (isomer 1, RT1 (min): 6.37, 1.18 mg, 0.8% yield).

[0274] LCMS (ESI): [M+H] + = 536.25

[0275] 1 H NMR (400 MHz, Methanol-d 4 ) δ 6.92 (s, 1H), 6.73–6.69 (m, 3H), 5.57 (s, 1H), 5.31–5.25 (m, 1H), 4.10–4.05 (m, 2H), 3.89 (s, 3H), 3.81 (d, J = 1.6 Hz, 6H), 3.70 (s, 3H), 2.95 (t, J = 6.4 Hz, 2H), 2.48 (s, 3H), 2.26–2.20 (m, 4H).

[0276] Compound 9-2 (isomer 2, RT2 (min): 7.05, 34.31 mg, 22% yield)

[0277] LCMS (ESI): [M+H] + = 536.30

[0278] 1 1H NMR (400 MHz, Methanol-d 4 ) δ 6.92 (s, 1H), 6.73–6.69 (m, 3H), 5.57 (s, 1H), 5.31–5.25 (m, 1H), 4.10–4.05 (m, 2H), 3.89 (s, 3H), 3.81 (d, J = 1.6 Hz, 6H), 3.70 (s, 3H), 2.95 (t, J = 6.4 Hz, 2H), 2.48 (s, 3H), 2.26–2.20 (m, 4H).

[0279] Example 10, Compound 10

[0280]

[0281] Step 1.

[0282]

[0283] Under nitrogen protection, 2-chloro-9,10-dimethoxy-2H,3H,6H,7H-pyrimido[4,3-a]isoquinolin-4-one (1 g, 3.4 mmol, 1 equiv) and 2,6-difluoro-4-methylaniline (582 mg, 4.0 mmol, 1.2 equiv), solvent isopropanol (15 mL), reacted at 80 °C overnight. Concentrated under reduced pressure to obtain (E)-2-((2,6-difluoro-4-methylphenyl)imino)-9,10-dimethoxy-2,3,6,7-tetrahydro-4H-pyrimido[6,1-a]isoquinolin-4-one (1 g, 73.80%).

[0284] LCMS (ESI): [M+H] + = 400.14

[0285] Step 2.

[0286]

[0287] Under nitrogen protection, (E)-2-((2,6-difluoro-4-methylphenyl)imino)-9,10-dimethoxy-2,3,6,7-tetrahydro-4H-pyrimido[6,1-a]isoquinolin-4-one (1 g, 2.5 mmol, 1 equiv), tert-butyl (3-bromocyclobutyl)carbamate (625 mg, 2.5 mmol, 1 equiv), cesium carbonate (2.4 g, 7.5 mmol, 3 equiv), and the solvent 1,4-dioxane solution (20 mL) were stirred at 100 °C overnight. The reaction mixture was diluted with water (100 mL), extracted with ethyl acetate (3 × 100 mL), the combined organic phases were backwashed with saturated sodium chloride (1 × 100 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by reverse-phase column chromatography under the following conditions: C18 column, mobile phase, water and acetonitrile, 10% to 50% gradient for 20 minutes, UV detector at 254 nm, to give tert-butyl (E)-(3-(2-((2,6-difluoro-4-methylphenyl)imino)-9,10-dimethoxy-4-oxo-6,7-dihydro-2H-pyrimido[6,1-a]isoquinolin-3(4H)-yl)cyclobutyl)carbamate (400 mg, 28.10%).

[0288] LCMS(ESI): [M+H] + = 569.25

[0289] Step 3.

[0290]

[0291] tert-Butyl (E)-(3-(2-((2,6-difluoro-4-methylphenyl)imino)-9,10-dimethoxy-4-oxo-6,7-dihydro-2H-pyrimido[6,1-a]isoquinolin-3(4H)-yl)cyclobutyl)carbamate (400 mg, 0.52 mmol, 1 equiv) was dissolved in 10 mL of 1,4-dioxane solution of hydrochloric acid (4 M) and stirred at room temperature for 1 hour. It was concentrated under reduced pressure, dissolved in 10 mL of water, extracted with ethyl acetate (3 × 10 mL), the organic layer was washed with 20 mL of saturated sodium chloride, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give (E)-3-(3-aminocyclobutyl)-2-((2,6-difluoro-4-methylphenyl)imino)-9,10-dimethoxy-2,3,6,7-tetrahydro-4H-pyrimido[6,1-a]isoquinolin-4-one (400 mg, crude).

[0292] LCMS(ESI): [M+H] + = 469.20

[0293] Step 4.

[0294]

[0295] Under nitrogen protection, at room temperature, a solution of (E)-3-(3-aminocyclobutyl)-2-((2,6-difluoro-4-methylphenyl)imino)-9,10-dimethoxy-2,3,6,7-tetrahydro-4H-pyrimido[6,1-a]isoquinolin-4-one (100 mg, 0.2 mmol, 1 equiv), trimethylsilyl isocyanate (36 mg, 0.3 mmol, 1.5 equiv) and triethylamine (64 mg, 0.6 mmol, 3 equiv) in dichloromethane (2 mL) was stirred for 2 h. Concentrated under reduced pressure. The crude product was purified by high performance liquid chromatography under the following conditions (column: YMC Triart C18 ExRs 5 μm, 20 mm × 250 mm; mobile phase A: water (10 mmol / L ammonium bicarbonate), mobile phase B: acetonitrile; flow rate: 25 mL / min; gradient: 30% B to 38% B in 10 min; wavelength: 254 nm / 220 nm; RT1 (min): 9.28 / 10.10) to obtain two isomeric compounds: Compound 10-1 and Compound 10-2.

[0296] Compound 10-1 (isomer 1, RT (min): 9.28, 16.86 mg, yield 15.38%)

[0297] LCMS (ESI): [M+H] + = 512.25

[0298] 1 H NMR (400 MHz, CDCl 3 ) δ 7.26 - 6.72 (m, 5H), 5.49 (s, 1H), 4.65 - 4.15 (m, 3H), 4.14 - 4.00 (m, 2H), 3.92 (s, 1H), 3.74 (s, 3H), 3.65 (s, 3H), 2.96 (m, 4H), 2.63 (s, 2H), 2.42 (s, 5H).

[0299] Compound 10-2 (isomer 2, RT (min): 10.10, 4.74 mg, yield 4.19%)

[0300] LCMS (ESI): [M+H] + = 512.25

[0301] 1 H NMR (400 MHz, CD 3OD) δ 7.13 - 6.82 (m, 4H), 5.65–5.35 (m, 2H), 4.21–4.11 (m, 2H), 4.10–3.73 (m, 5H), 3.61 (s, 2H), 3.16 - 2.88 (m, 2H), 2.67 (s, 3H), 2.48–2.19 (m, 4H).

[0302] Example 11, Compound 11

[0303]

[0304] Step 1.

[0305]

[0306] tert-Butyl (E)-(2-(2-(methylsulfonimidoyl)-9,10-dimethoxy-4-oxo-6,7-dihydro-2H-pyrimido[6,1-a]isoquinolin-3(4H)-yl)propyl)carbamate (500 mg, 0.911 mmol) was stirred in dioxane hydrochloride (10 mL, 4 M) at room temperature for 0.5 h. The resulting mixture was concentrated under reduced pressure. The product was obtained (300 mg, yield 73.39%).

[0307] LCMS(ESI): [M + H] + = 449.16

[0308] Step 2.

[0309]

[0310] (2E)-3-(1-Aminopropan-2-yl)-9,10-dimethoxy-2-[(2,4,6-trimethylphenyl)imino]-6H,7H-pyrimido[4,3-a]isoquinolin-4-one (200 mg, 0.45 mmol), triethylamine (135 mg, 1.338 mmol) and trimethylsilyl isocyanate (102.73 mg, 0.9 mmol) were stirred in DCM (10 mL) at room temperature for 2 h. The resulting mixture was concentrated under reduced pressure. The resulting mixture was extracted with EtOAc (3 x 50 mL). The combined organic layers were washed with brine (3 x 50 mL) and dried over anhydrous Na 2 SO 4Dry. After filtration, the filtrate was concentrated under reduced pressure. The crude product was purified by reversed-phase flash chromatography under the following conditions (column: Xselect CSH C18 5 μm, 30 mm × 150 mm; mobile phase A: water (0.1% FA), mobile phase B: acetonitrile; flow rate: 60 mL / min; gradient: from 28% B to 38% B in 8 minutes; wavelength: 254 nm / 220 nm; RT1 (min): 5.85 / 7.65), to obtain the product (100 mg, yield 45.62%)

[0311] LCMS(ESI): [M+H] + = 492.35

[0312] Step 3.

[0313]

[0314] The product (100 mg) was purified by CHIRAI-HPLC under the following conditions (column: CHIRALPAK IE-3, 4.6 * 50 mm, 3 μm; mobile phase A: DCM - HPLC, mobile phase B: ETOH:DCM = 1:1; flow rate: 18 mL / min; gradient: isocratic 20; wavelength: 220 nm; sample solvent: MeOH - HPLC injection volume: 0.45 mL; number of runs: 5), to obtain two isomeric compounds of the product: Compound 11-1, Compound 11-2.

[0315] Compound 11-1 (isomer 1, RT (min) = 17.4 min, 6.9 mg, yield 6.28%)

[0316] LCMS(ESI): [M+H] + = 492.35

[0317] 1 H NMR(400MHz, Methanol-d 4 ) δ 7.14–7.06 (m, 2H), 6.92 (s, 1H), 6.67 (s, 1H), 5.50 (s, 1H), 4.58–4.50 (m, 1H), 4.20–4.10 (m, 1H), 4.11–4.00 (m, 1H), 3.95–3.87 (m, 1H), 3.87 (s, 3H), 3.66 (s, 3H), 3.28–3.19 (m, 1H), 2.97 (t, J = 6.5 Hz, 2H), 2.35 (s, 3H), 2.17 (d, J = 8.1 Hz, 6H), 1.15 (d, J = 6.3 Hz, 3H).

[0318] Compound 11-2 (isomer 2, RT (min) = 21.8 min, 17.1 mg, yield 37.07)

[0319] LCMS (ESI): [M+H] + = 492.35

[0320] 1 H NMR (400 MHz, Methanol-d 4 ) δ 7.14–7.06 (m, 2H), 6.92 (s, 1H), 6.67 (s, 1H), 5.50 (s, 1H), 4.58–4.50 (m, 1H), 4.20–4.10 (m, 1H), 4.11–4.00 (m, 1H), 3.95–3.87 (m, 1H), 3.87 (s, 3H), 3.66 (s, 3H), 3.28–3.19 (m, 1H), 2.97 (t, J = 6.5 Hz, 2H), 2.35 (s, 3H), 2.17 (d, J = 8.1 Hz, 6H), 1.15 (d, J = 6.3 Hz, 3H).

[0321] Example 12, Compound 12

[0322]

[0323] Step 1.

[0324]

[0325] To a stirred solution of 2-chloro-9,10-dimethoxy-6H,7H-pyrimido[4,3-a]isoquinolin-4-one (2.0 g, 6.8 mmol) in DCM (20 mL) was added dropwise boron tribromide (8.5 g, 34.1 mmol) at 0 °C. The resulting mixture was stirred at room temperature for 2 h. The reaction was quenched with water / ice at room temperature. The residue was purified by reverse-phase flash chromatography with the following conditions: column, C18 silica gel; mobile phase, acetonitrile water (10 mmol / L NH 4 HCO 3 ) with a gradient from 10% to 50% in 10 min; detector, UV 254 nm. 2-Chloro-9,10-dihydroxy-6H,7H-pyrimido[4,3-a]isoquinolin-4-one (1.8 g, yield 69.6%) was obtained,

[0326] LCMS (ESI): [M+H] + = 265.0

[0327] Step 2.

[0328]

[0329] A solution of 2-chloro-9,10-dihydroxy-6H,7H-pyrimido[4,3-a]isoquinolin-4-one (1.8 g, 6.8 mmol) in DMF (20 mL) was treated with CD 3 I (2.4 g, 17.0 mmol) overnight at room temperature. The resulting mixture was diluted with water (50 mL). The aqueous layer was extracted with EtOAc (3 x 50 mL). The residue was purified by reverse-phase flash chromatography under the following conditions: column, C18 silica gel; mobile phase, MeCN in water (10 mmol / L NH 4 HCO 3 ), gradient from 10% to 50% in 10 minutes; detector, UV 254 nm. 2-Chloro-9,10-bis(2H3)methoxy-6H,7H-pyrimido[4,3-a]isoquinolin-4-one (700 mg, yield 34.4%) was obtained.

[0330] LCMS (ESI): [M+H] + = 299.1

[0331] Step 3.

[0332]

[0333] 2-Chloro-9,10-bis(2H3)methoxy-6H,7H-pyrimido[4,3-a]isoquinolin-4-one (700 mg, 2.3 mmol) in 10 mL of isopropanol (IPA) was reacted with 2,4,6-trimethylaniline (1.5 g, 11.7 mmol) overnight under a nitrogen atmosphere at 90 °C. The resulting mixture was concentrated under reduced pressure. The residue was washed with EtOAc (20 mL). (2E)-9,10-Bis(2H3)methoxy-2-[(2,4,6-trimethylphenyl)imino]-3H,6H,7H-pyrimido[4,3-a]isoquinolin-4-one (900 mg, yield 77.3%) was obtained..

[0334] LCMS (ESI): [M+H] + = 398.2

[0335] Step 4.

[0336]

[0337] (2E)-9,10-Bis(2H3)methoxy-2-[(2,4,6-trimethylphenyl)imino]-3H,6H,7H-pyrimido[4,3-a]isoquinolin-4-one (900 mg, 2.2 mmol) in 1,4-dioxane (10 mL) was reacted with tert-butyl N-(2-bromoethyl)carbamate (761 mg, 3.3 mmol), Pd2 (dba) 3 (207 mg, 0.2 mmol) and cesium carbonate (2.2 g, 6.7 mmol) were heated overnight at 100 °C under nitrogen. The resulting mixture was diluted with water (20 mL). The aqueous layer was extracted with EtOAc (3 x 50 mL). The residue was purified by reverse-phase flash chromatography under the following conditions: column, C18 silica gel; mobile phase, MeCN-water (10 mmol / L NH 4 HCO 3 ), gradient from 10% to 50% in 10 minutes; detector, UV 254 nm. N-{2-[(2E)-9,10-Bis(2H3)methoxy-4-oxo-2-[(2,4,6-trimethylphenyl)imino]-6H,7H-pyrimido[4,3-a]isoquinolin-3-yl]ethyl}carbamic acid tert-butyl ester (350 mg, yield 28.5%) was finally obtained.

[0338] LCMS (ESI): [M+H] + = 541.3

[0339] Step 5.

[0340]

[0341] At room temperature, a mixture of N-{2-[(2E)-9,10-bis(2H3)methoxy-4-oxo-2-[(2,4,6-trimethylphenyl)imino]-6H,7H-pyrimido[4,3-a]isoquinolin-3-yl]ethyl}carbamic acid tert-butyl ester (350 mg, 0.6 mmol) and 1,4-dioxane (10 mL, 4 M) of HCl was stirred for 1 hour. The resulting mixture was concentrated under reduced pressure. 350 mg of the crude product was obtained. The crude product was used directly in the next step without further purification.

[0342] LCMS (ESI): [M+H] + = 441.3

[0343] Step 6.

[0344]

[0345] (2E)-3-(2-Aminoethyl)-9,10-bis(2H3)methoxy-2-[(2,4,6-trimethylphenyl)imino]-6H,7H-pyrimido[4,3-a]isoquinolin-4-one (350 mg, 0.8 mmol) was treated with trimethylsilyl isocyanide (137.2 mg, 1.2 mmol) in DCM (5 mL) overnight at room temperature. The resulting mixture was concentrated under reduced pressure. The resulting mixture was diluted with water (10 mL). The aqueous layer was extracted with EtOAc (3 x 50 mL). The residue was purified by reverse-phase flash chromatography under the following conditions: column, C18 silica gel; mobile phase, MeCN-water (10 mmol / L NH 4 HCO 3 ), gradient from 10% to 50% in 10 minutes; detector, UV 254 nm. 2-[(2E)-9,10-Bis(2H3)methoxy-4-oxo-2-[(2,4,6-trimethylphenyl)imino]-6H,7H-pyrimido[4,3-a]isoquinolin-3-yl]ethylurea (30.6 mg, yield 7.9%) was obtained.

[0346] LCMS (ESI): [M+H] + = 484.40

[0347] 1 H NMR (400 MHz, Methanol-d 4 ) δ 7.11 (s, 2H), 6.94 (d, J = 6.6 Hz, 1H), 6.70 (s, 1H), 5.51 (s, 1H), 4.16–4.09 (m, 2H), 3.95 (t, J = 6.8 Hz, 2H), 3.44 (t, J = 6.8 Hz, 2H), 2.98 (t, J = 6.4 Hz, 2H), 2.36 (s, 3H), 2.18 (s, 6H).

[0348] Example 13, Compound 13

[0349]

[0350] Step 1.

[0351]

[0352] tert-Butyl carbamate (0.8 g, 6.8 mmol) was reacted with NaH (0.4 g, 17.1 mmol) in THF (20 mL) at 0 °C for 30 minutes under a nitrogen atmosphere, and then stirred at 0 °C for 30 minutes under a nitrogen atmosphere. Then 2-chloro-9,10-dimethoxy-6H,7H-pyrimido[4,3-a]isoquinolin-4-one (1 g, 3.4 mmol, 1.0 equiv) was added portionwise at 0 °C. The resulting mixture was stirred at room temperature under a nitrogen atmosphere for 2 hours. The resulting mixture was extracted with EtOAc (3 x 50 mL). The combined organic layers were washed with brine (2 x 50 mL) and dried over anhydrous Na 2 SO 4 After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography, eluting with CH 2 Cl 2 / MeOH to give benzyl N-[(2E)-9,10-dimethoxy-4-oxo-3H,6H,7H-pyrimido[4,3-a]isoquinolin-2-ylidene]carbamate (750 mg, yield 53.9%).

[0353] LCMS (ESI, m / z): [M+H] + = 374.2

[0354] Step 2.

[0355]

[0356] To a stirred solution of tert-butyl N-[(2E)-9,10-dimethoxy-4-oxo-3H,6H,7H-pyrimido[4,3-a]isoquinolin-2-ylidene]carbamate (850 mg, 2.3 mmol) and tert-butyl N-(2-bromoethyl)carbamate (612.1 mg, 2.7 mmol) in dioxane (3 mL) at room temperature under a nitrogen atmosphere, Cs 2 CO 3 (1483.3 mg, 4.6 mmol, 2.0 equiv) and Pd 2 (dba) 3 (208.5 mg, 0.2 mmol) were added portionwise. The resulting mixture was stirred at 100 °C under a nitrogen atmosphere overnight. The resulting mixture was extracted with EtOAc (3 x 50 mL). The combined organic layers were washed with brine (2 x 50 mL) and dried over anhydrous Na 2 SO 4Dry. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by reverse-phase flash chromatography under the following conditions: column, C18 silica gel; mobile phase, aqueous MeCN, gradient from 10% to 50% in 10 minutes; detector, UV 254 nm. N-[(2E)-3-(2-[(tert-butoxycarbonyl)amino]ethyl-9,10-dimethoxy-4-oxo-6H,7H-pyrimido[4,3-a]isoquinolin-2-ylidene]carbamic acid tert-butyl ester (450 mg, yield 38.3%) was obtained.

[0357] LCMS(ESI,m / z):[M+H] + =517.3

[0358] Step 3.

[0359]

[0360] A mixture of N-[(2E)-3-(2-[(tert-butoxycarbonyl)amino]ethyl-9,10-dimethoxy-4-oxo-6H,7H-pyrimido[4,3-a]isoquinolin-2-ylidene]carbamic acid tert-butyl ester (450 mg, 0.9 mmol) and 1,4-dioxane (10 mL) of HCl was stirred for 4 h at room temperature under a nitrogen atmosphere. The resulting mixture was concentrated under reduced pressure. The residue was purified by reverse-phase flash chromatography under the following conditions: column, C18 silica gel; mobile phase, MeCN in water (0.1% FA), 10%-90% gradient, 30 min; detector, UV 254 nm. 3-(2-Aminoethyl)-2-imino-9,10-dimethoxy-6H,7H-pyrimido[4,3-a]isoquinolin-4-one (200 mg, yield 72.6%) was obtained.

[0361] LCMS(ESI,m / z):[M+H] + =317.2

[0362] Step 4.

[0363]

[0364] To a solution of 3-(2-aminoethyl)-2-imino-9,10-dimethoxy-6H,7H-pyrimido[4,3-a]isoquinolin-4-one (200 mg, 0.6 mmol) and triethylamine (319.9 mg, 3.2 mmol) in DCM (3 mL) under nitrogen at room temperature, isocyanotrimethylsilane (145.7 mg, 1.3 mmol) was added portionwise. The resulting mixture was stirred overnight under nitrogen at room temperature. The resulting mixture was concentrated under reduced pressure. The residue was purified by reverse-phase flash chromatography under the following conditions: column, C18 silica gel; mobile phase, aqueous acetonitrile, 10%-50% gradient, 20 minutes; detector, UV at 254 nm. As a result, 2-(2-imino-9,10-dimethoxy-4-oxo-6H,7H-pyrimido[4,3-a]isoquinolin-3-ylethyl)urea (150 mg, yield 66.1%) was obtained.

[0365] LCMS(ESI,m / z):[M+H] + =360.2

[0366] Step 5.

[0367]

[0368] To a stirred solution of 2-(2-imino-9,10-dimethoxy-4-oxo-6H,7H-pyrimido[4,3-a]isoquinolin-3-ylethyl)urea (150 mg, 0.417 mmol) and K 2 CO 3 (115.4 mg, 0.8 mmol) in DMF (5 mL) under nitrogen at room temperature, 2-(bromomethyl)-1,3,5-trimethylbenzene (177.9 mg, 0.8 mmol) was added portionwise. The resulting mixture was stirred overnight at 70 °C under nitrogen. The resulting mixture was concentrated under reduced pressure. The residue was purified by reverse-phase flash chromatography under the following conditions: (column: YMC Triart C18 ExRs5m, 30 mm * 150 mm; mobile phase A: water (10 mmol / L ammonium bicarbonate), mobile phase B: acetonitrile; flow rate: 60 mL / min; gradient: from 43% B to 45% B in 10 minutes; wavelength: 254 nm / 220 nm; RT1(min): 9.38), and 2-[(2E)-9,10-dimethoxy-4-oxo-2-([(2,4,6-trimethylphenyl)methyl]imino-6H,7H-pyrimido[4,3-a]isoquinolin-3-yl]ethyl]urea (4.8 mg, 2.32%) was obtained.

[0369] LCMS(ESI,m / z):[M+H] + =492.50

[0370] 1 1H NMR (400 MHz, Methanol-d 4 ) δ 7.41 (s, 1H), 6.90 (s, 1H), 6.79 (s, 2H), 6.42 (s, 1H), 5.24 (s, 2H), 3.96 (t, J=6.2 Hz, 1H), 3.95 (s, 4H), 3.91 (s, 3H), 3.48 (d, J=6.2 Hz, 2H), 3.29 (t, J=6.2 Hz, 2H), 2.92 (t, J=6.2 Hz, 2H), 2.33 (s, 6H), 2.23 (s, 3H), 0.12 (d, J=2.2 Hz, 1H).

[0371] Example 14, Compound 14

[0372]

[0373] Step 1.

[0374]

[0375] Add H 2 O 2 (14.8 mL, 190.6 mmol, 5.0 eq, 30%) to a mixture of 2,6-dimethylbenzonitrile (5.0 g, 38.1 mmol) and KOH (4.28 g, 76.2 mmol, 2.0 eq) in MeOH (50 mL) and DMSO (5 mL), and stir under an air atmosphere at 0 °C. Stir the resulting mixture overnight at room temperature under a nitrogen atmosphere. Extract the resulting mixture with ethyl acetate (2 × 50 mL). Combine the organic layers and wash with brine (3 × 50 mL), and dry over anhydrous Na 2 SO 4 . Concentrate the filtered filtrate under reduced pressure. Purify by silica gel column chromatography, eluting with PE / EA (3 / 2) to obtain 2,6-dimethylbenzylamine (4.8 g, yield 80.19%).

[0376] LCMS (ESI): [M+H]+ = 150.1

[0377] Step 2.

[0378]

[0379] Add XantPhos (100 mg, 0.2 mmol, 0.1 eq) and Cs 2 CO 3(1.7 g) was added portionwise to a stirred solution of 2,6-dimethylbenzamide (250 mg, 1.7 mmol) and 2-chloro-9,10-dimethoxy-6H,7H-pyrimido[4,3-a]isoquinolin-4-one (500 mg, 1.7 mmol) in dioxane (5 mL). The resulting mixture was stirred at 105 °C under a nitrogen atmosphere for 6 h. The resulting mixture was filtered and the filter cake was washed with methanol (3 × 15 mL). The filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography, eluting with EA / MeOH (8 / 1) to give N-[(2E)-9,10-dimethoxy-4-oxo-3H,6H,7H-pyrimido[4,3-a]isoquinolin-2-yl]-2,6-dimethylbenzamide (450 mg, yield 64.98%).

[0380] LCMS(ESI): [M+H]+ = 406.4

[0381] Step 3.

[0382]

[0383] N-[(2E)-9,10-Dimethoxy-4-oxo-3H,6H,7H-pyrimido[4,3-a]isoquinolin-2-yl]-2,6-dimethylbenzylamine (200 mg, 0.5 mmol) was added to a solution of N-(2-bromoethyl)carbamic acid tert-butyl ester (120 mg, 0.5 mmol) and Cs 2 CO 3 (200 mg, 0.6 mmol) stirred in DMF (4 ml). The resulting mixture was stirred at room temperature under a nitrogen atmosphere for 14 h. The resulting mixture was filtered and the filter cake was washed with methanol (3 × 50 mL). The filtrate was concentrated under reduced pressure. Purification was carried out by reverse phase flash chromatography, conditions: the column was C18 silica gel; the mobile phase, acetonitrile in water (10 mmol / L NH 4 HCO 3 ), gradient 60% - 70%, 10 min; UV detector, 254 nm. As a result, tert-butyl N-{2-[(2E)-2-(2,6-dimethylbenzoyl)-9,10-dimethoxy-4-oxo-6H,7H-pyrimido[4,3-a]isoquinolin-3-yl]ethyl}carbamate (200 mg, yield 73.90%) was obtained.

[0384] LCMS(ESI): [M+H]+ = 549.3

[0385] Step 4.

[0386]

[0387] N-{2-[(2E)-2-(2,6-Dimethylbenzoyl)-9,10-dimethoxy-4-oxo-6H,7H-pyrimido[4,3-a]isoquinolin-3-yl]ethyl}carbamate (200 mg, 0.4 mmol) was stirred in 1,4-dioxane (4 mL, 4 M) under an air atmosphere at room temperature for 2 h. The resulting mixture was concentrated under reduced pressure. The obtained mixture was concentrated in vacuo. As a result, N-[(2E)-3-(2-aminoethyl)-9,10-dimethoxy-4-oxo-6H,7H-pyrimido[4,3-a]isoquinolin-2-yl]-2,6-dimethylbenzamide (120 mg, yield 73.39%) was obtained.

[0388] LCMS(ESI): [M+H] + = 449.2

[0389] Step 5.

[0390]

[0391] Trimethylsilyl isocyanate (33 mg, 0.3 mmol, 1.2 equiv.) was added dropwise to a stirred solution of N-[(2E)-3-(2-aminoethyl)-9,10-dimethoxy-4-oxo-6H,7H-pyrimido[4,3-a]isoquinolin-2-yl]-2,6-dimethylbenzylamine (110 mg, 0.2 mmol) and TEA (55 mg, 0.5 mmol) in DCM (2 mL) at room temperature. The resulting mixture was stirred under an air atmosphere at room temperature for 2 h. The resulting mixture was concentrated under reduced pressure. Preparative HPLC (column: XBridge BEH C18 OBD Prep column 130, 5 μm, 30 mm * 150 mm; mobile phase A: water (10 mmol / L NH4HCO3), mobile phase B: ACN; flow rate: 60 mL / min; gradient: 30% B to 48% B, 10 min; wavelength: 254 nm / 220 nm; RT1 (min): 9.6.) was used to obtain N-[(2E)-3-[2-(carbamoylamino)ethyl]-9,10-dimethoxy-4-oxo-6H,7H-pyrimido[4,3-a]isoquinolin-2-yl]-2,6-dimethylbenzamide (22.7 mg, yield 18.77%).

[0392] LCMS(ESI): [M+H] + = 492.35

[0393] 11H NMR (400 MHz, DMSO-d6) δ 7.17–7.09 (m, 3H), 7.06 (s, 1H), 7.03 (d, J = 7.5 Hz, 2H), 6.05 (t, J = 6.0 Hz, 1H), 5.34 (s, 2H), 4.10–4.02 (m, 4H), 3.87–3.83 (m, 6H), 3.24 (q, J = 6.4 Hz, 2H), 2.99 (t, J = 6.3 Hz, 2H), 2.30 (s, 6H).

[0394] Example 15, Compound 15

[0395]

[0396] Step 1.

[0397]

[0398] Add 2-[(2E)-9,10-dimethoxy-4-oxo-2-[(2,4,6-trimethylphenyl)imino]-6h,7h-pyrimido[4,3-a]isoquinolin-3-yl]ethylurea (100 mg, 0.2 mmol) and HBr (0.5 mL) into an 8 mL vial. The resulting mixture was stirred at 110 °C for 3 hours under a nitrogen atmosphere. Purification was carried out by reverse-phase flash chromatography with the following conditions: the column was C18 silica gel, the mobile phase was acetonitrile (0.1% FA) in water with a 10% - 50% gradient for 10 min; UV detector at 254 nm. As a result, 2-[(2E)-9,10-dihydroxy-4-oxo-2-[(2,4,6-trimethylphenyl)imino]-6h,7h-pyrimido[4,3-a]isoquinolin-3-yl]ethylurea (50 mg, yield 53.12%) was obtained.

[0399] LCMS (ESI): [M+H] + = 450.2

[0400] Step 2.

[0401]

[0402] 2-[(2E)-9,10-Dihydroxy-4-oxo-2-[(2,4,6-trimethylphenyl)imino]-6H,7H-pyrimido[4,3-a]isoquinolin-3-yl]ethyl urea (100 mg, 0.2 mmol) and ethyl 2-bromo-2,2-difluoroacetate (59 mg, 0.3 mmol) were stirred in DMF (0.5 mL), and K2CO3 (92 mg, 0.7 mmol) was added under a nitrogen atmosphere at 25 °C. The reaction mixture was stirred under a nitrogen atmosphere at 60 °C for 2 h. Purification was performed by reverse-phase flash chromatography with the following conditions: column, C18 silica gel; mobile phase, acetonitrile (0.1% FA) in water, 10% - 50% gradient, 10 min; UV detector, 254 nm. The result was 2-[(2E)-9-(difluoromethoxy)-10-hydroxy-4-oxo-2-[(2,4,6-trimethylphenyl)imino]-6H,7H-pyrimido[4,3-a]isoquinolin-3-yl]ethyl urea (12 mg, yield 10.80%).

[0403] LCMS(ESI): [M+H] + = 500.2

[0404] Step 3.

[0405]

[0406] 2-[(2E)-9-(Difluoromethoxy)-10-hydroxy-4-oxo-2-[(2,4,6-trimethylphenyl)imino]-6H,7H-pyrimido[4,3-a]isoquinolin-3-yl]ethyl urea (50 mg, 0.1 mmol) and (bromoethyl)cyclopropane (14 mg, 0.1 mmol) were stirred in DMF (0.5 mL), and K2CO3 (17 mg, 0.1 mmol) was added under a nitrogen atmosphere at 25 °C. The reaction mixture was stirred under a nitrogen atmosphere at 50 °C for 2 h. Purification was performed by reverse-phase flash chromatography with the following conditions: column, C18 silica gel; mobile phase, acetonitrile (0.1% FA) in water, gradient 50% - 80%, 10 min; UV detector, 254 nm. The result was 2-[(2E)-10-(cyclopropylmethoxy)-9-(difluoromethoxy)-4-oxo-2-[(2,4,6-trimethylphenyl)imino]-6H,7H-pyrimido[4,3-a]isoquinolin-3-yl]ethyl urea (4.7 mg, yield 8.37%).

[0407] LCMS(ESI): [M+H] + = 555.41

[0408] 1 H NMR(400MHz, Methanol-d 4)δ7.14(s,1H),7.11(s,2H),6.96–6.88(m,1H),6.86(s,1H),5.56(s,1H),4.59(s,1H),4.13(t,J=6.3Hz,2H),3.99–3.93(m,2H),3.78(d,J=6.8Hz,2H),3.46(t,J=6.8Hz,2H),2.97(t,J=6.3Hz,2H),2.37(s,3H),2.18(s,6H),2.14(s,1H),1.34–1.28(m,1H),1.18–1.05(m,1H),0.63–0.52(m,2H),0.28–0.22(m,2H).

[0409] Example 16, Compound 16

[0410]

[0411] Step 1.

[0412]

[0413] At room temperature, dissolve 4-bromo-2,6-dimethylaniline (1.0 g, 5.0 mmol) in DMF (15 ml), and successively add K 2 CO 3 (2.07 g, 15.0 mmol) and KI (2.07 g, 12.5 mmol). Under a nitrogen atmosphere, add dropwise BnBr (2.14 g, 12.5 mmol). After addition, raise the temperature to 130 °C and stir for 16 h. After completion of the reaction, add water (20 ml) to the mixture, and extract with ethyl acetate (30 ml × 3). Combine the organic phases, dry over anhydrous sodium sulfate, filter, remove the solvent, and concentrate under reduced pressure. The obtained residue is purified by flash silica gel column chromatography (eluent: n-heptane) to obtain the product (1.6 g, 84.2%).

[0414] LCMS(ESI,m / z):[M+H] + =380.0

[0415] Step 2.

[0416]

[0417] At room temperature, dissolve compound N,N-dibenzyl-4-bromo-2,6-dimethylaniline (1.5 g, 3.9 mmol, 1 eq) in THF (15 ml). Under a nitrogen atmosphere, cool the solution to -80 °C, add 1.6 M n-BuLi solution (3.75 ml, 5.85 mmol). After addition, stir the mixture at -80 °C for 16 h. After the reaction is complete, add saturated aqueous ammonium chloride solution (30 ml) to the mixture, and extract with ethyl acetate (60 ml × 2). Combine the organic phases, dry over anhydrous sodium sulfate, filter, remove the solvent, and concentrate under reduced pressure. The resulting residue is purified by flash silica gel column chromatography (eluent: n-heptane / ethyl acetate = 50:1 - 10:1) to obtain the product (0.78 g, 62.1%).

[0418] LCMS(ESI,m / z):[M+H] + =319.2

[0419] Step 3.

[0420]

[0421] At room temperature, dissolve compound N,N-dibenzyl-2,6-dimethyl-4-(methyl-d3)aniline (1.4 g, 4.4 mmol) in ethyl acetate (70 ml). Sequentially add Pd / C (0.14 g, 10% w / w) and Pd(OH) 2 (0.14 g, 10% w / w), pressurize with a hydrogen balloon, and stir the reaction mixture at 40 °C for 16 h. After the reaction is complete, filter the mixture, and concentrate the filtrate under reduced pressure to obtain the product (0.65 g, 62.1%).

[0422] LCMS(ESI,m / z):[M+H] + =139.12

[0423] Step 4.

[0424]

[0425] At room temperature, add compound 2,6-dimethyl-4-(methyl-d3)aniline (6.4 g, 5.0 mmol, 5 eq) and 2-chloro-9,10-dimethoxy-6,7-dihydro-4H-pyrimido[6,1-a]isoquinolin-4-one (3.0 g, 12.5 mmol) to IPA (30 ml). Under a nitrogen atmosphere, heat the mixture to 90 °C and stir for 18 h. After the reaction is complete, concentrate under reduced pressure to remove the solvent. The resulting residue is purified by flash silica gel column chromatography (eluent: n-heptane / ethyl acetate = 30:1 - 1:1) to obtain the product (3.4 g, 84.2%).

[0426] LCMS(ESI,m / z):[M+H] += 395.21

[0427] Step 5.

[0428]

[0429] At room temperature, (E)-2-((2,6-dimethyl-4-(methyl-d3)phenyl)imino)-9,10-dimethoxy-2,3,6,7-tetrahydro-4H-pyrimido[6,1-a]isoquinolin-4-one (3.4 g, 8.6 mmol) and Nphth-I (13.20 g, 51.6 mmol) were added to MIBK (120 ml), and K 2 CO 3 (10.77 g, 77.4 mmol) was added. Under a nitrogen atmosphere, the temperature was raised to 85 °C and the mixture was stirred for 64 h. After the reaction was completed, the solvent was removed by concentration under reduced pressure. The resulting residue was stirred with methanol (50 ml), filtered, and the product was obtained (2.26 g, 84.2%).

[0430] LCMS(ESI, m / z): [M+H] + = 568.2579

[0431] Step 6.

[0432]

[0433] At room temperature, (E)-2-(2-((2,6-dimethyl-4-(methyl-d3)phenyl)imino)-9,10-dimethoxy-4-oxo-6,7-dihydro-2H-pyrimido[6,1-a]isoquinolin-3(4H)-yl)ethyl)isoindoline-1,3-dione (2.2 g, 3.9 mmol) was dissolved in chloroform (22 ml) and EtOH (22 ml), and hydrazine hydrate (2.2 ml, 85%) was added. Under a nitrogen atmosphere, the mixture was stirred at 25 °C for 16 h. After the reaction was completed, it was filtered, and the solvent was removed by concentration under reduced pressure from the filtrate to obtain the product (1.56 g, 92%).

[0434] LCMS(ESI, m / z): [M+H] + = 438.2580

[0435] Step 7.

[0436]

[0437] At room temperature, dissolve (E)-3-(2-aminoethyl)-2-((2,6-dimethyl-4-(methyl-d3)phenyl)imino)-9,10-dimethoxy-2,3,6,7-tetrahydro-4H-pyrimido[6,1-a]isoquinolin-4-one (1.35 g, 3.1 mmol) in 1,2-dichloroethane (27 ml), add trimethylsilyl isocyanate (1.24 g, 3.1 mmol) dropwise, and under a nitrogen atmosphere, stir the mixture at 25 °C for 16 h. After the reaction is completed, concentrate the solvent under reduced pressure, and purify the obtained residue by flash silica gel column chromatography (eluent: dichloromethane / methanol = 100:1 to 10:1) to obtain the product (0.8 g, 92%).

[0438] LCMS(ESI,m / z):[M+H] + =481.10

[0439] 1 H NMR(400MHz,CDCl 3 )δ6.88(s,2H),6.69(s,1H),6.67(s,1H),5.44(s,1H),4.41-4.38(t,J=7.2Hz,2H),4.05-4.02(t,J=6.1Hz,2H),3.90(s,3H),3.76(s,3H),3.53-3.52(d,J=7.5Hz,2H),2.92–2.89(t,2H),2.05-2.04(s,6H).

[0440] Example 17, Compound 17

[0441]

[0442] Step 1.

[0443]

[0444] At room temperature, add (E)-3-(2-aminoethyl)-2-(methyltrimethylimine)-9,10-dimethoxy-2,3,6,7-tetrahydro-4H-pyrimido[6,1-a]isoquinolin-4-one (0.5 g, 1.2 mmol) and methanol (5 mL) to a 50 mL flask and stir. Add 3,4-dimethoxycyclobut-3-ene-1,2-one (0.17 g, 1.2 mmol), and stir at room temperature under nitrogen protection for 24 h. After the reaction is completed, cool the system to 0 °C and stir for 1 h. Filter by suction, wash the filter cake with 1 mL of methanol, and purify by silica gel column to obtain the product (0.4 g, 63.8%).

[0445] LCMS(ESI,m / z):[M+H] + =545.2

[0446] Step 2.

[0447]

[0448] At room temperature, (E)-3-((2-(2-(methyltrimethylimine)-9,10-dimethoxy-4-oxo-6,7-dihydro-2H-pyrimido[6,1-a]isoquinolin-3(4H)-yl)ethyl)amino)-4-methoxycyclobut-3-ene-1,2-dione (1 g, 1.84 mmol) and methanol (15 mL) were added to a 50 mL flask and stirred. 7M ammonia-methanol solution (0.39 mL) was added, and the mixture was stirred at room temperature under nitrogen protection for 24 h. After the reaction was completed, the system was evaporated to dryness under reduced pressure and purified by silica gel column chromatography to obtain the product (711 mg, 73.1%).

[0449] LCMS(ESI,m / z):[M+H] + =530.1

[0450] 1 H NMR(400MHz,DMSO-d 6 )δ7.90 - 7.15(broad,3H),6.95(s,1H),6.85(s,2H),6.65(s,1H),5.35(s,1H),4.35(m,2H),3.95 - 3.75(m,6H),3.62(s,3H),2.88(s,2),2.23(s,3H),1.96(s,6H).

[0451] Example 18, Compound 18

[0452]

[0453] Step 1.

[0454]

[0455] Under a nitrogen atmosphere, (E)-2-(methylimine)-9,10-dimethoxy-2,3,6,7-tetrahydro-4H-pyrimido[6,1-a]isoquinolin-4-one (20.61 g) and tert-butyl 3-(2-(toluenyloxy)ethyl)imidazolidine-1-carboxylate (3.5 g) were dissolved in DMF (70 ml). Sodium carbonate (11.13 g) and sodium iodide (8.17 g) were added successively, and the temperature was raised to 80 °C and the reaction was maintained for 72 h. After the reaction was completed, water (140 ml) and ethyl acetate (100 ml × 3) were added. The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The obtained residue was purified by flash silica gel column chromatography (eluent: n-heptane:ethyl acetate = 20:1 to 5:1) to obtain the product (1.4 g).

[0456] LCMS (ESI, m / z): [M+H] + = 604.31

[0457] Step 2.

[0458]

[0459] At room temperature, dissolve tert-butyl (E)-3-(2-(2-(methyltrimethyliminium)-9,10-dimethoxy-4-oxo-6,7-dihydro-2H-pyrimido[6,1-a]isoquinolin-3(4H)-yl)ethyl)-2-oxoimidazolidine-1-carboxylate (1.3 g) in ethyl acetate (13 ml), add dropwise a solution of hydrogen chloride in ethyl acetate (4 N, 5.4 ml), stir the reaction for 4 h, adjust the pH to about 8 - 9 by adding saturated aqueous sodium bicarbonate dropwise, separate the layers, wash with water (15 ml × 3), dry the organic phase over anhydrous sodium sulfate, filter off the desiccant, concentrate under reduced pressure, and subject the resulting residue to high-pressure preparation to obtain the product (30 mg, 2.8%).

[0460] LCMS (ESI, m / z): [M+H] + = 504.25

[0461] 1 H NMR (400 MHz, CDCl 3 ) δ 6.87 (s, 2H), 6.72 (s, 1H), 6.65 (s, 1H), 5.45 (s, 1H), 4.49 - 4.40 (m, 2H), 4.21 (s, 1H), 4.08 - 4.01 (m, 2H), 3.90 (s, 3H), 3.74 (s, 3H), 3.71 - 3.52 (m, 4H), 3.43 - 3.36 (m, 2H), 2.92 - 2.85 (m, 2H), 2.27 (s, 3H), 2.08 (s, 6H).

[0462] PDE enzyme activity inhibition experiment

[0463] Use the FP method to test the inhibition of PDE enzyme activity by the compounds of each example, and use the compound RPL-554 in Example 1 of CN100415743C as a positive control.

[0464]

[0465] Prepare the PDE enzyme and Substrate (FAM-cyclic adenosine monophosphate / cyclic adenosine monophosphate) solutions in the reaction buffer (1×IMAP Reaction Buffer containing 0.1% BSA supplemented with 1 mM DTT). The starting concentration of the positive control for PDE is 1 / 10 μM, diluted 3-fold, 10 + 0 dose. Deliver 0.05 μL of the compound in 100% DMSO to a 384-well plate (Corning 4514) by acoustic liquid delivery technology (Echo 655) and centrifuge at 1000 rpm for 1 minute; transfer 2.5 μL of the PDE enzyme solution to the 384-well reaction plate and centrifuge at 1000 rpm for 1 minute, incubate at 25 °C for 10 min; transfer 2.5 μL of the Sub solution to the 384-well reaction plate, centrifuge at 1000 rpm for 1 minute, and incubate at 25 °C for 60 min; transfer 15 μL of the binder mixture to the 384-well reaction plate and centrifuge at 1000 rpm for 1 minute, incubate at 25 °C for 60 min; finally, read the FP signal using BMG (PHERAstar FSX). Obtain the IC50 value and non-linear regression curve fitting using GraphPad Prism software.

[0466] Table 2 shows the IC 50 values of the enzyme activity experiments of the compounds in the comparative examples and examples. 50 The smaller the IC

[0467] Table 2

[0468]

[0469] The results show that the compounds of the present invention have high inhibitory activity against PDE3 / 4 and have practical value.

[0470] The above are only specific embodiments of the present invention. It should be noted that for those of ordinary skill in the art, without departing from the principle of the present invention, several improvements and refinements can be made, and these improvements and refinements should also be regarded as the protection scope of the present invention.

Claims

1. A compound having the structural formula I or a pharmaceutically acceptable form thereof, wherein the pharmaceutically acceptable form is selected from pharmaceutically acceptable salts or co-crystals, stereoisomers, tautomers, deuterated compounds, solvates, chelates, non-covalent complexes or prodrugs: Wherein: R 1 and R 2 are each independently selected from H, C 1-6 linear alkyl, C 3-6 branched alkyl, and C 3-6 cycloalkyl; the linear alkyl, branched alkyl, or cycloalkyl is optionally further substituted with 0 to 4 substituents selected from H, F, Cl, Br, I, OH, ═O, NH 2 , CN, COOH, C 1-4 alkyl, C 1-4 alkoxy; R 3 、R 4 、R 5 each independently selected from H, halogen, CN, C 1-6 alkoxy, C 1-6 linear alkyl, C 3-6 branched alkyl, and C 3-6 cycloalkyl; the alkoxy, linear alkyl, branched alkyl or cycloalkyl is optionally further substituted with 0 to 4 substituents selected from H, F, Cl, Br, I, OH, =O, NH 2 , CN, COOH, C 1-4 alkyl, C 1-4 alkoxy; R 6 selected from said R 6 in which H is optionally further substituted by 0 to 4 substituents selected from F, Cl, Br, I, OH, ═O, NH 2 , CN, COOH, C 1-4 alkyl, C 1-4 alkoxy; R 7 selected from H, =O, NH 2 , CN, C 1-6 linear alkyl, C 3-6 branched alkyl, C 3-6 cycloalkyl and said R 7 the H in which is optionally further substituted by 0 to 4 substituents selected from F, Cl, Br, I, OH, =O, NH 2 , CN, COOH, C 1-4 alkyl, C 1-4 alkoxy; L is selected from n is 0, 1 or 2; k is 0, 1 or 2; the H in said L is optionally further substituted by 0 to 4 substituents selected from F, Cl, Br, I, OH, ═O, NH 2 , CN, COOH, C 1-4 alkyl, C 1-4 alkoxy; X is N; Y is C 1-3 an alkylene group or is absent; the alkylene group is optionally further substituted with 0 to 4 substituents selected from F, Cl, Br, I, OH, =O, NH 2 , CN, COOH, C 1-4 alkyl, C 1-4 alkoxy; Optionally, R 1 and R 2 are connected together with the attached O to form a 5- or 6-membered ring; the H in the 5- or 6-membered ring is optionally further substituted by 0 to 4 substituents selected from F, Cl, Br, I, OH, ═O, NH 2 , CN, COOH, C 1-4 alkyl, C 1-4 alkoxy; Optionally, X and L are linked by C 1-2 to form a 5- or 6-membered ring; the H in the 5- or 6-membered ring is optionally further substituted by 0 to 4 substituents selected from F, Cl, Br, I, OH, =O, NH 2 , CN, COOH, C 1-4 alkyl, C 1-4 alkoxy; Optionally, R 6 forms a 5- or 6-membered ring together with L; the H in the 5- or 6-membered ring is optionally further substituted with 0 to 4 substituents selected from F, Cl, Br, I, OH, ═O, NH 2 , CN, COOH, C 1-4 alkyl, C 1-4 alkoxy; Optionally, the H of the compound of formula I is further optionally substituted by 0 to 6 D; And, the compound of formula I is not:

2. The compound or its pharmaceutically acceptable form according to claim 1, Wherein: R 1 、R 2 are each independently selected from H, CH 3 、CHF 2 、CD 3 。 3. The compound or its pharmaceutically acceptable form according to claim 1 or 2, Wherein: R 3 、R 4 、R 5 Each independently selected from H, CH 3 、i-Pr, OMe, CN, CD 3 or halogen.

4. The compound or its pharmaceutically acceptable form according to any one of claims 1-3, Wherein: R 6 for and / or R 7 is H, =O or 5. The compound or its pharmaceutically acceptable form according to any one of claims 1-4, Wherein: L is selected from n is 0, 1 or 2; k is 1 or 2.

6. The compound or its pharmaceutically acceptable form according to any one of claims 1-5, Wherein, Y is absent.

7. The compound or its pharmaceutically acceptable form according to claim 1, Wherein, The compound is selected from one or more of the following compounds:

8. An intermediate compound having the structure shown in formula II: Wherein, R 1 、R 2 、R 3 、R 4 、R 5 、R 7 、X, Y are defined as in any one of claims 1 - 7; L 1 Selected from n is 0, 1 or 2; k is 0, 1 or 2; the L 1 in which H is optionally further substituted by 0 to 4 substituents selected from F, Cl, Br, I, OH, ═O, NH 2 , CN, COOH, C 1-4 alkyl, C 1-4 alkoxy; Preferably, the intermediate compound has the following structure:

9. A method for preparing the compound or its pharmaceutically acceptable form according to any one of claims 1-7, the method Comprises: Performing a modification reaction on the carboxyl terminus shown in the intermediate compound of formula II of claim 8 to prepare a compound having the structural formula I.

10. The method according to claim 9, the method further comprises the process of preparing the intermediate compound of claim 8.

11. A pharmaceutical composition, which Comprises: The compound or its pharmaceutically acceptable form according to any one of claims 1-7, and a pharmaceutically acceptable carrier, excipient and / or one or more other therapeutic agents.

12. Use of the compound or its pharmaceutically acceptable form according to any one of claims 1-7 or the pharmaceutical composition according to claim 11 in the preparation of a preparation for inhibiting phosphodiesterase.

13. Use of the compound or its pharmaceutically acceptable form according to any one of claims 1-7 or the pharmaceutical composition according to claim 11 in the preparation of a drug for treating phosphodiesterase-related diseases; Preferably, wherein the phosphodiesterase-related diseases include respiratory diseases such as asthma.

Images