A class of enteric-lytic co-drugs and their preparation and uses

Abstract

The present invention relates to a class of enterolytic co-drugs and their preparation and uses. Specifically, the present invention provides a co-drug compound represented by Formula I. The present invention also provides methods for treating gastrointestinal autoimmune diseases, inflammatory diseases and cancers using such compounds; as well as methods and intermediates for preparing such compounds.

Description

Technical Field

[0001] The present invention relates to a class of gut-lytic co-drug compounds. The present invention also relates to a pharmaceutical composition comprising such compounds; a method for treating gastrointestinal autoimmune diseases, inflammatory diseases and cancers using such compounds; and a method and intermediate for preparing such compounds. Background Art

[0002] JAK family (JAK1, JAK2, JAK3, TYK2) inhibitors such as tofacitinib have been approved for the treatment of certain patients with moderate to severe active rheumatoid arthritis (RA) and moderate to severe ulcerative colitis (UC). In many clinical trials of JAK family inhibitors, a large number of adverse events mediated by systemic drug exposure have been reported, including severe infections, opportunistic infections and laboratory abnormalities, such as lymphopenia, neutropenia, elevated liver enzymes, elevated lipids and elevated serum creatinine. Currently marketed JAK inhibitors carry a black box warning of safety risks, including severe infections, malignancies and thrombosis risks. Therefore, developing a new generation of safer JAK family inhibitor drugs will require limiting their systemic exposure when treating local inflammatory diseases. For example, when treating UC, increasing the distribution of JAK family inhibitors in the gastrointestinal tract while minimizing the systemic exposure of the drug.

[0003] Berberine, also known as berberine hydrochloride, is an isoquinoline alkaloid extracted from plants such as Coptis chinensis. Berberine is a very safe drug and has a history of more than a thousand years in the application of traditional Chinese medicine. Its bioavailability is very low. Clinically, it is mainly used to treat gastrointestinal diseases such as diarrhea and intestinal infections. In recent years, studies have also found that berberine has certain therapeutic prospects in cardiovascular diseases and the regulation of glucose and lipid metabolism. Berberrubine is the main metabolite of berberine in vivo. Animal model studies have found that berberrubine has a therapeutic effect similar to that of berberine on ulcerative colitis. However, up to now, there is still a lack of effective ways to improve the therapeutic effect of berberine or its analogs in this field.

[0004] Chronic enteritis mainly includes two types: ulcerative colitis and Crohn's disease. These chronic intestinal inflammatory diseases have a long course, often recur, and long-term inflammation is prone to canceration. In recent years, the incidence of chronic enteritis has shown an upward trend. It is currently believed that the onset of chronic enteritis may be related to genetics, environment, immunity, and microorganisms, but its exact mechanism is not clear. Clinically, the treatment mainly uses aminosalicylate drugs, adrenal glucocorticoid drugs, and immunosuppressants, but they all have certain adverse reactions, such as gastrointestinal discomfort, allergic reactions, etc. We found in our previous work that the combination of JAK inhibitors and berberine analogs can act synergistically to achieve a more superior therapeutic effect on gastrointestinal inflammatory diseases. In the present invention, a class of enteric-lytic berberine analogs and JAK inhibitor co-drugs are designed, so that the two drug molecules are released and enriched in the gastrointestinal tract, while the systemic exposure of the compound is restricted, bringing better safety. At the same time, the two drug molecules act synergistically to achieve a better therapeutic effect. Summary of the Invention

[0005] In one aspect, the present invention provides a co-drug composed of a first therapeutic agent berberine analog (preferably berberrubine) and a second therapeutic agent JAK family inhibitor (preferably tofacitinib, upadacitinib, SHR0302) through a cleavable covalent linkage. The co-drug is designed to be able to release the JAK family inhibitor and the first therapeutic agent in the gastrointestinal tract directionally, thereby increasing the content of the JAK family inhibitor and the first therapeutic agent at the gastrointestinal inflammatory site and minimizing its systemic exposure.

[0006] In the first aspect of the present invention, a co-drug compound represented by the following formula I is provided. The co-drug compound is formed by coupling a first drug molecule, a second drug molecule, and a linker precursor:

[0007] D1-linker-D2;

[0008] I

[0009] Wherein,

[0010] D1 is a first drug group; the first drug group is a structural fragment in the first drug molecule that can be linked to the linker (that is, after the first drug molecule is coupled or condensed with the precursor of the linker and the reactive functional group is removed, the formed fragment, which does not include the linker part);

[0011] D2 is the second drug group; the second drug group is the structural fragment in the second drug molecule that can be linked to the linker; and the first drug molecule and the second drug molecule are drug molecules with synergistic effects (that is, the fragment formed after the second drug molecule is coupled or condensed with the precursor of the linker and then the reactive functional group is removed, and this fragment does not include the linker part);

[0012] Among them, the connection can be the formation of a covalent bond by losing a hydrogen atom, or covalently linked to the linker in other ways, such as the covalent bond formed by the condensation reaction of reactive groups such as hydroxyl, carboxyl, and amino groups;

[0013] And the linker has a structure selected from the following group (a), (b) or (c). In each formula, J1 is linked to the first drug group, and J2 is linked to the second drug group;

[0014]

[0015] The Glu has a structure selected from the following group:

[0016]

[0017] Among them, the A ring is selected from the following group: C6-C10 aryl, 5-10 membered heteroaryl, 3-12 membered heterocyclic group;

[0018]

[0019] Among them, the R4 is selected from the following group: H, C 1-6 alkyl, C 1～6 alkoxy-C 1～4 alkylene-, C 3～12 cycloalkyl, C 3～12 cycloalkyl-C 1～4 alkylene-;

[0020] (c)

[0021]

[0022] Among them, the B ring and the C ring are each independently selected from the following group: C6-C10 aryl, 5-10 membered heteroaryl, 3-12 membered heterocyclic group;

[0023] In the above formulas (a), (b) and (c), the J1 and J2 are each independently -(Y) z-, and the said Y is selected from the following groups: -NH-, -C(O)-, -C(O)O-, -NHC(O)NH-, -CH=CH-, -NH(CH2)-, -NHC(O)-, -CH2-, -OCH2CH2O-, -O-, -S-, -P(O)2O-, -S(O)2-, -S(O)-, -C(O)NH-, -N=N-; the said Y can be substituted by one or more R, provided that each Y together forms a chemically stable structure;

[0024] Each of L1, L2, L3, L4, L5, L6 and L7 is independently selected from the following groups: C1-C8 alkylene, C 1～6 alkylene-O-C 1～4 alkylene (-CH2-O-CH2-), C 2～6 alkenyl, C 2～6 alkynyl, C 3～6 cycloalkyl, C6-C10 arylene, heteroarylene of 5-10 atoms, heterocyclic group composed of 3-12 atoms, or a group selected from the following groups: -NH-, -C(O)-, -CH=CH-, -NH(CH2)-, -NHC(O)-, -CH2-, -OCH2CH2O-, -O-, -S-, -P(O)2O-, -S(O)2-, -S(O)-, -C(O)NH-, -C(O)O-, -NHC(O)NH-, -N=N-, -C(O)NH(CH2) (1-4) -NHC(O)-; provided that each of L1, L2, L3, L4, L5, L6 and L7 forms a stable divalent group;

[0025] and the said Y, L1, L2, L3, L4, L5, L6 and L7 are optionally substituted by one or more R, and the said R is selected from the following groups: H, -OH, C1-C4 alkyl, halogen, cyano, nitro, -OR4, C 1～6 haloalkyl, sulfonic acid group, C0-C4 alkyl-S(O)2-C1-C4 alkyl, formyl, carboxyl, -COOR4; provided that each of Y, L1, L2, L3, L4, L5, L6 and L7 together forms a chemically stable structure;

[0026] m, n, p, q, r, s and t are each independently selected from 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15 or 16;

[0027] z is selected from the following groups: 0, 1, 2, 3, 4, 5, 6; preferably, z is selected from the following groups: 1, 2 or 3.

[0028] In another preferred example, the said connection includes: the loss of a structural fragment in the drug molecule to form a connection site, or connection through a coordination bond.

[0029] In another preferred embodiment, in the above formulas (a), (b) and (c), J1 and J2 are each independently -(Y) z -, and Y is selected from the group consisting of: -NH-, -C(O)-, -C(O)O-, -NHC(O)NH-, -CH=CH-, -NH(CH2)-, -NHC(O)-, -CH2-, -OCH2CH2O-, -O-, -S-, -P(O)2O-, -S(O)2-, -S(O)-, -C(O)NH-, -N=N-; Y may be substituted by one or more R, provided that each Y together forms a chemically stable structure;

[0030] Each of L1, L2, L3, L4, L5, L6 and L7 is independently selected from the group consisting of: C1-C8 alkylene, C 1～6 alkylene-O-C 1～4 alkylene (-CH2-O-CH2-), C 2～6 alkenyl, C 2～6 alkynyl, C 3～6 cycloalkyl, C6-C10 arylene, 5-10 membered heteroarylene, 3-12 membered heterocycloalkylene, or a group selected from the group consisting of: -NH-, -C(O)-, -CH=CH-, -NH(CH2)-, -NHC(O)-, -CH2-, -OCH2CH2O-, -O-, -S-, -P(O)2O-, -S(O)2-, -S(O)-, -C(O)NH-, -C(O)O-, -NHC(O)NH-, -N=N-, -C(O)NH(CH2) (1-4) -NHC(O)-; provided that each of L1, L2, L3, L4, L5, L6 and L7 forms a stable divalent group;

[0031] And Y, L1, L2, L3, L4, L5, L6 and L7 are optionally substituted by one or more R, and R is selected from the group consisting of: H, -OH, C1-C4 alkyl, halogen, cyano, nitro, -OR4, C 1～6 haloalkyl, sulfonic acid group, C0-C4 alkyl-S(O)2-C1-C4 alkyl, formyl, carboxyl, -COOR4; provided that each of Y, L1, L2, L3, L4, L5, L6 and L7 together forms a chemically stable structure;

[0032] And m, n, p, q, r, s and t are each independently selected from 0, 1, 2, 3, 4, 5, 6, 7 or 8.

[0033] z is selected from the group consisting of: 0, 1, 2, 3, 4, 5, 6; preferably, z is selected from the group consisting of: 1, 2 or 3.

[0034] In another preferred example, in the above formulas (a), (b) and (c), J1 and J2 are each independently selected from the following group: -NH-, -C(O)-, -C(O)O-, -NHC(O)NH-, -CH=CH-, -NH(CH2)-, -NHC(O)-, -CH2-, -OCH2CH2O-, -O-, -S-, -P(O)2O-, -S(O)2-, -S(O)-, -C(O)NH-, -N=N-.

[0035] In another preferred example, J1 and J2 are each independently selected from the following group: methylene

[0036] In another preferred example, J1 is selected from the following group: methylene

[0037] In another preferred example, J2 is selected from the following group: methylene

[0038] In another preferred example, the first drug molecule is berberine, berberrubine and their analogs.

[0039] In another preferred example, the second drug molecule is a JAK family inhibitor and its analogs.

[0040] In another preferred example, the linker has the following structure:

[0041]

[0042] In another preferred example, the first drug molecule is a drug molecule of the following formula II, formula III or formula IV:

[0043]

[0044] wherein

[0045] Ro, Rp, Rq, Rr, Rs and Rt are each independently selected from the following group: H, substituted or unsubstituted C1-C4 alkyl, substituted or unsubstituted C1-C4 alkoxy; or Ro, Rp, Rq, Rr, Rs and Rt located on two adjacent atoms together with the atoms to which they are attached form a 5-7 membered heterocycle; wherein, the substitution means that the H atom on the group is substituted by one or more substituents selected from the following group: halogen, C1-C4 alkyl, phenyl.

[0046] In another preferred embodiment, the JAK family inhibitors and their analogs are selected from the group consisting of: Tofacitnib, Ruxolitinib, Oclacitinib, Baricitinib, Peficitinib, Abrocitinib, Filgotinib, Upadacitinib, Delgocitinib, Itacitinib, Fedratinib, Decernotinib, SHR-0302, AZD-4205, ASN-002, BMS-986165, PF-06700841, PF-06651600, R-348, INCB-52793, ATI-501, ATI-502, NS-018, KL-130008, or deuterated derivatives of the above molecules.

[0047] In another preferred embodiment, the first drug group is selected from the group consisting of:

[0048]

[0049] or the first drug group is a group formed by removing one hydrogen atom from a drug molecule selected from the group consisting of:

[0050]

[0051]

[0052] In another preferred embodiment, the first drug group has the structure shown in the following formula:

[0053]

[0054] In another preferred embodiment, the second drug group is selected from the group consisting of:

[0055]

[0056]

[0057] In another preferred embodiment, the second drug group is selected from the group consisting of:

[0058]

[0059] In another preferred embodiment, the first drug group is and the second drug group is

[0060]

[0061] In another preferred example, the A-(L7) p -J2- has the structure shown in the following formula:

[0062]

[0063] In another preferred example, the -A(Glu)-(L7) p -J2- has a structure selected from the following group:

[0064]

[0065] In another preferred example, the -(L1) m - and -(L2) n - each independently have a structure selected from the following group:

[0066]

[0067] In the above formulas,

[0068] Ra, Rb, and Rc are each independently a group formed by removing a hydrogen atom from an amino acid selected from the following group: Glycine, Alanine, Valine, Leucine, Isoleucine, Phenylalanine, Tryptophan, Tyrosine, Aspartate, Histidine, Asparagine, Glutamate, Lysine, Glutamine, Methionine, Arginine, Serine, Threonine, Cysteine, Proline.

[0069] In another preferred example, the linker is selected from the following group (A), (B), or (C):

[0070] Group (A) has the structure of -L a -L-, where the L a has a structure selected from the following group:

[0071]

[0072] and the L has the structure shown below, where * is the connection between L and L aLinking sites:

[0073]

[0074] (B) group:

[0075]

[0076] (C) group:

[0077]

[0078]

[0079] In another preferred embodiment, the compound is selected from the group consisting of:

[0080]

[0081]

[0082]

[0083]

[0084]

[0085]

[0086]

[0087] In another preferred embodiment, the compound is selected from the group consisting of:

[0088]

[0089]

[0090]

[0091]

[0092]

[0093] In a second aspect of the present invention, there is provided a pharmaceutical composition comprising a therapeutically effective amount of the compound or its stereoisomer or racemate or its pharmaceutically acceptable salt according to the first aspect of the present invention, and a pharmaceutically acceptable excipient.

[0094] In another preferred embodiment, the pharmaceutical composition is an enteric-coated preparation.

[0095] In another preferred embodiment, the pharmaceutical composition is used for treating diseases selected from the group consisting of gastrointestinal inflammatory diseases (such as ulcerative colitis, Crohn's disease, colitis associated with immune checkpoint inhibitor therapy, collagenous colitis, lymphocytic colitis, pouchitis, acute / chronic gastritis, acute / chronic appendicitis), gastroenteritis caused by radiotherapy or chemotherapy, autoimmune diseases of the gastrointestinal tract (such as graft-versus-host disease, celiac disease, autoimmune enteropathy), peptic ulcer, irritable bowel syndrome, gastric cancer, esophageal cancer, and colon cancer.

[0096] In a third aspect of the present invention, there is provided the use of a precursor compound as described in the first aspect of the present invention, or a pharmaceutically acceptable salt thereof, or the pharmaceutical composition as described in the second aspect of the present invention, for preventing and treating gastrointestinal functional diseases.

[0097] In another preferred embodiment, the gastrointestinal functional disease is a gastrointestinal inflammatory disease.

[0098] In another preferred embodiment, the gastrointestinal inflammatory disease is selected from the group consisting of ulcerative colitis, Crohn's disease, and colitis associated with immune checkpoint inhibitor therapy.

[0099] It should be understood that within the scope of the present invention, the above technical features of the present invention and the technical features specifically described below (such as in the examples) can be combined with each other to form new or preferred technical solutions. Due to space limitations, they will not be elaborated one by one here. BRIEF DESCRIPTION OF THE DRAWINGS

[0100] Figure 1 Shows the curves of the concentration of the compound of Example 8 in different tissues over time after oral administration of the compound of Example 8 to mice;

[0101] Figure 2 Shows the curves of the concentration of berberrubine in different tissues over time after oral administration of the compound of Example 8 to mice;

[0102] Figure 3 Shows the curves of the concentration of tofacitinib in different tissues over time after oral administration of the compound of Example 8 to mice;

[0103] Figure 4 Shows the AUC of the content of berberrubine in different tissues after oral administration of the compounds of Example 1 and 8 to mice 0-24h ;

[0104] Figure 5 Shows the AUC of the content of tofacitinib in different tissues after oral administration of the compounds of Example 1 and 8 to mice 0-24h ;

[0105] Figure 6The graph shows the changes in the disease index of mice after administration of the compound of Example 8 in the oxazolone enema model. DETAILED DESCRIPTION

[0106] After long-term and in-depth research, the inventors found that preparing JAK family inhibitors and berberine analogs into a co-drug form for administration can achieve a better therapeutic effect for the treatment of gastrointestinal diseases than a single drug at the same dose, and the design of the co-drug molecule can achieve a targeted release of the drug. Based on the above findings, the inventors completed the present invention.

[0107] the term

[0108] Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs.

[0109] As used herein, the term "comprising" or "including (comprising)" may be open, semi-closed and closed. In other words, the term also includes "consisting essentially of" or "consisting of".

[0110] In the present application, as a group or part of other groups, the term "alkyl" refers to a fully saturated straight or branched hydrocarbon chain group, consisting only of carbon atoms and hydrogen atoms, having, for example, 1 to 12 (preferably 1 to 8, more preferably 1 to 6) carbon atoms, and connected to the rest of the molecule by a single bond, for example, including but not limited to methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, n-pentyl, 2-methylbutyl, 2,2-dimethylpropyl, n-hexyl, heptyl, 2-methylhexyl, 3-methylhexyl, octyl, nonyl and decyl, etc. For the purpose of the present invention, the term "C1-C6 alkyl" refers to an alkyl group containing 1 to 6 carbon atoms.

[0111] In the present application, as a group or part of other groups, the term "6-10 membered aromatic ring" means an aromatic ring having 6-10 ring atoms, wherein the ring atoms are carbon atoms. The aromatic ring can be a monocyclic or bicyclic ring. For example, a benzene ring, a naphthalene ring and the like.

[0112] In the present application, as a group or part of other groups, the term "5-10 membered heteroaromatic ring" means a heteroaromatic ring having 5-10 ring atoms, wherein at least one (or 1, 2 or 3) of the ring atoms is a heteroatom selected from nitrogen, oxygen and sulfur. The heteroaromatic ring may be monocyclic or bicyclic. For example, pyrimidopyrazole ring, pyrazinoimidazole ring, pyridopyrazole ring, pyridoimidazole ring, pyridopyrimidine ring, pyridopyridine ring.

[0113] In the present application, as a group or as part of other groups, the term "heterocyclic group" means a stable 3- to 20-membered non-aromatic cyclic group composed of 3 to 14 carbon atoms and 1 to 6 heteroatoms selected from nitrogen, phosphorus, oxygen, and sulfur. Unless otherwise specifically specified in this specification, the heterocyclic group can be a monocyclic, bicyclic, tricyclic, or more-ring ring system, which can include a fused-ring system, a bridged-ring system, or a spiro-ring system; the nitrogen, carbon, or sulfur atoms in the heterocyclic group can be optionally oxidized; the nitrogen atom can be optionally quaternized; and the heterocyclic group can be partially or fully saturated. The heterocyclic group can be connected to the rest of the molecule via a carbon atom or a heteroatom by a single bond. In a heterocyclic group containing fused rings, one or more rings can be an aryl or heteroaryl group as defined below, provided that the connection point to the rest of the molecule is a non-aromatic ring atom. For the purposes of the present invention, the heterocyclic group is preferably a stable 4- to 11-membered non-aromatic monocyclic group containing 1 to 3 heteroatoms selected from nitrogen, oxygen, and sulfur.

[0114] Co-drug

[0115] In this context, "co-drug", "co-drug", "co-loaded drug", or "interactive drug" can be used interchangeably, and all refer to a drug molecule that can be metabolized in vivo to form two drug molecules with different pharmacological effects. In this context, a typical co-drug is the compound shown in Formula I.

[0116] The co-drug described above can be metabolized in vivo to form a variety of different therapeutic agents. In the present invention, a preferred first therapeutic agent is berberine or its analogs, and the second therapeutic agent is a JAK family inhibitor.

[0117] First drug molecule

[0118] In this context, "first therapeutic agent" and "first drug molecule" can be used interchangeably, and both refer to the first drug molecule of the co-drug used in the present invention. After losing any active functional group on the leaving group, the first drug molecule can form a first drug group and connect to the connection site of the co-drug molecule.

[0119] In the present invention, berberine and its analogs can be used as the first therapeutic agent of the co-drug. Preferred first drug molecules are shown in the following Formula II, III, or IV:

[0120]

[0121] Second drug molecule

[0122] In this context, "second therapeutic agent" and "second drug molecule" can be used interchangeably, and both refer to the second drug molecule of the co-drug used in the present invention. After losing any hydrogen atom on the leaving group, the second drug molecule can form a second drug group and connect to the connection site of the co-drug molecule.

[0123] A preferred class of second drug molecules are JAK family inhibitors, which can be used clinically for intestinal diseases, such as the treatment of intestinal inflammatory diseases. The JAK family inhibitors can be JAK inhibitors known in the art, or compounds that have not been verified to have JAK inhibitory activity.

[0124] Exemplary second drug groups formed by JAK inhibitors are selected from the group consisting of:

[0125]

[0126]

[0127] The compounds of the present invention

[0128] The compounds of the present invention are compounds of formula I or their stereoisomers or racemates or their pharmaceutically acceptable salts.

[0129] The compounds of the present invention may contain one or more chiral carbon atoms, and thus enantiomers, diastereomers and other stereoisomeric forms may be generated. Each chiral carbon atom can be defined as (R)- or (S)- based on stereochemistry. The present invention is intended to include all possible isomers, as well as their racemates and optically pure forms. The compounds of the present invention can be prepared by selecting racemates, diastereomers or enantiomers as starting materials or intermediates. Optically active isomers can be prepared using chiral synthons or chiral reagents, or resolved using conventional techniques, such as crystallization and chiral chromatography.

[0130] Conventional techniques for the preparation / separation of individual isomers include chiral synthesis from suitable optically pure precursors or the resolution of racemates (or racemates of salts or derivatives) using, for example, chiral high-performance liquid chromatography. See, for example, Gerald Gübitz and Martin G. Schmid (Eds.), Chiral Separations, Methods and Protocols, Methods in Molecular Biology, Vol. 243, 2004; A. M. Stalcup, Chiral Separations, Annu. Rev. Anal. Chem. 3:341-63, 2010; Fumiss et al. (eds.), VOGEL’S ENCYCLOPEDIA OF PRACTICAL ORGANIC CHEMISTRY 5.sup.TH ED., Longman Scientific and Technical Ltd., Essex, 1991, 809-816; Heller, Acc. Chem. Res. 1990, 23, 128.

[0131] The term "pharmaceutically acceptable salts" includes pharmaceutically acceptable acid addition salts and pharmaceutically acceptable base addition salts.

[0132] "Pharmaceutically acceptable acid addition salts" refer to salts formed with inorganic or organic acids that are able to retain the biological effectiveness of the free base without other side effects. Inorganic acid salts include, but are not limited to, hydrochloride, hydrobromide, sulfate, nitrate, phosphate, etc.; organic acid salts include, but are not limited to, formate, acetate, 2,2-dichloroacetate, trifluoroacetate, propionate, caproate, caprylate, caprate, undecylenate, glycolate, gluconate, lactate, sebacate, adipate, glutarate, malonate, oxalate, maleate, succinate, fumarate, tartrate, citrate, palmitate, stearate, oleate, cinnamate, laurate, malate, glutamate, pyroglutamate, aspartate, benzoate, methanesulfonate, benzenesulfonate, p-toluenesulfonate, alginate, ascorbate, salicylate, 4-aminosalicylate, naphthalenedisulfonate, etc. These salts can be prepared by methods known in the art.

[0133] "Pharmaceutically acceptable base addition salts" refer to salts formed with inorganic or organic bases that can maintain the biological effectiveness of the free acid without other side effects. Salts derived from inorganic bases include, but are not limited to, sodium salts, potassium salts, lithium salts, ammonium salts, calcium salts, magnesium salts, iron salts, zinc salts, copper salts, manganese salts, aluminum salts, etc. Preferred inorganic salts are ammonium salts, sodium salts, potassium salts, calcium salts, and magnesium salts. Salts derived from organic bases include, but are not limited to, the following salts: primary amines, secondary amines, and tertiary amines, substituted amines, including naturally occurring substituted amines, cyclic amines, and basic ion exchange resins, such as ammonia, isopropylamine, trimethylamine, diethylamine, triethylamine, tripropylamine, ethanolamine, diethanolamine, triethanolamine, dimethylethanolamine, 2-dimethylaminoethanol, 2-diethylaminoethanol, dicyclohexylamine, lysine, arginine, histidine, caffeine, procaine, choline, betaine, ethylenediamine, glucosamine, methylglucosamine, theobromine, purine, piperazine, piperidine, N-ethylpiperidine, polyamine resins, etc. Preferred organic bases include isopropylamine, diethylamine, ethanolamine, trimethylamine, dicyclohexylamine, choline, and caffeine. These salts can be prepared by methods known in the art.

[0134] Preparation Method

[0135] The following reaction schemes exemplify methods for preparing the compounds of formula I or their stereoisomers or racemates or their pharmaceutically acceptable salts, wherein each group is as described above. It should be understood that in the following reaction schemes, combinations of substituents and / or variables in the general formula are only permissible when such combinations result in stable compounds. It should also be understood that other general formulas can be prepared by those skilled in the art of organic chemistry by the methods disclosed herein (by using appropriately substituted starting materials and modifying synthetic parameters as needed using methods well known to those skilled in the art) or by known methods.

[0136] In various aspects and embodiments, the present invention relates to glucuronide co-drugs of tofacitinib or their pharmaceutically acceptable salts; pharmaceutical compositions containing such compounds; methods of using such compounds to treat gastrointestinal inflammatory diseases; and methods and intermediates for preparing such compounds.

[0137] The compounds described herein may contain one or more chiral centers. In such cases, the depiction or naming of a particular stereoisomer means that the indicated stereogenic center has the specified stereochemistry, where it should be understood that, unless otherwise indicated, small amounts of other stereoisomers may also be present, provided that the utility of the depicted or named compound is not eliminated by the presence of another stereoisomer.

[0138] Additionally, as used herein, unless otherwise indicated, "compounds of the present invention" and "compounds of formula I" (or similar terms) are intended to include pharmaceutically acceptable salts.

[0139] Application

[0140] Since the co-drug compound of the present invention has excellent intestinal targeted release effect, the compounds of the present invention and their various crystal forms, pharmaceutically acceptable inorganic or organic salts, hydrates or solvates, and pharmaceutical compositions containing the compounds of the present invention as the main active ingredient can be used for preventing and / or treating intestinal functional diseases, preferably gastrointestinal inflammatory diseases.

[0141] In the present application, the term "pharmaceutical composition" refers to a preparation of the compound of the present invention and a medium commonly accepted in the art for delivering a bioactive compound to a mammal (such as a human). This medium includes a pharmaceutically acceptable carrier. The purpose of the pharmaceutical composition is to facilitate the administration to an organism, facilitate the absorption of the active ingredient and thus exert its biological activity.

[0142] In the present application, the term "pharmaceutically acceptable" refers to a substance (such as a carrier or diluent) that does not affect the biological activity or properties of the compound of the present invention and is relatively non-toxic, that is, the substance can be administered to an individual without causing adverse biological reactions or interacting with any component contained in the composition in an adverse manner.

[0143] In the present application, the term "pharmaceutically acceptable excipient" includes, but is not limited to, any adjuvant, carrier, excipient, glidant, sweetening agent, diluent, preservative, dye / colorant, flavoring agent, surfactant, wetting agent, dispersing agent, suspending agent, stabilizer, isotonic agent, solvent or emulsifying agent that is permitted by the relevant government regulatory authorities for use in humans or livestock.

[0144] In the present application, the term "tumor" includes, but is not limited to, diseases such as glioma, sarcoma, melanoma, articular chondroma, cholangioma, leukemia, gastrointestinal stromal tumor, histiocytic lymphoma, non-small cell lung cancer, small cell lung cancer, pancreatic cancer, squamous cell carcinoma of the lung, adenocarcinoma of the lung, breast cancer, prostate cancer, liver cancer, skin cancer, epithelial cell carcinoma, cervical cancer, ovarian cancer, intestinal cancer, nasopharyngeal cancer, brain cancer, bone cancer, esophageal cancer, melanoma, renal cancer, oral cancer, etc.

[0145] In the present application, the terms "preventive", "prevention" and "prevention of" include reducing the likelihood of the occurrence or worsening of a disease or disorder in a patient.

[0146] In the present application, the terms "treatment" and other similar synonyms include the following meanings:

[0147] (i) Preventing the occurrence of a disease or disorder in a mammal, especially when such a mammal is susceptible to the disease or disorder but has not been diagnosed as having the disease or disorder;

[0148] (ii) Inhibiting a disease or disorder, that is, curbing its development;

[0149] (iii) alleviating a disease or disorder, i.e., causing the state of the disease or disorder to regress; or

[0150] (iv) reducing the symptoms caused by the disease or disorder.

[0151] In the present application, the terms "effective amount", "therapeutically effective amount" or "pharmaceutically effective amount" refer to the amount of at least one agent or compound that, when administered, is sufficient to alleviate to some extent one or more symptoms of the disease or disorder being treated. The result can be the reduction and / or alleviation of signs, symptoms or causative factors, or any other desired change in a biological system. For example, an "effective amount" for treatment is the amount of a composition comprising a compound disclosed herein that is required to provide a significant alleviation of the disorder clinically. Techniques such as dose escalation trials can be used to determine the effective amount suitable for any individual case.

[0152] In the present application, the terms "administering", "applying", "dosing", etc. refer to methods capable of delivering a compound or composition to the desired site for biological action. These methods include, but are not limited to, oral route, duodenal route, parenteral injection (including intravenous, subcutaneous, intraperitoneal, intramuscular, intraarterial injection or infusion), topical administration and rectal administration. Those skilled in the art are familiar with the administration techniques applicable to the compounds and methods described herein, such as those discussed in Goodman and Gilman, The Pharmacological Basis of Therapeutics, current ed.; Pergamon; and Remington’s, Pharmaceutical Sciences (current edition), Mack Publishing Co., Easton, Pa. In a preferred embodiment, the compounds and compositions discussed herein are administered orally.

[0153] In the present application, the terms "drug combination", "drug combination therapy", "combination therapy", "administering additional therapy", "administering additional therapeutic agent", etc. refer to a pharmaceutical treatment obtained by mixing or combining more than one active ingredient, which includes fixed and non-fixed combinations of active ingredients. The term "fixed combination" refers to the simultaneous administration to a patient of at least one compound described herein and at least one synergistic agent in the form of a single entity or a single dosage form. The term "non-fixed combination" refers to the simultaneous administration, co-administration or sequential administration at variable intervals to a patient of at least one compound described herein and at least one synergistic agent in the form of separate entities. These also apply to cocktail therapies, such as the administration of three or more active ingredients.

[0154] Compared with the prior art, the main advantages of the present invention are:

[0155] 1. The co-drug compound of the present invention itself cannot be effectively absorbed. It can release two active pharmaceutical ingredients directionally in the intestine, so it can cause the enrichment of the active pharmaceutical ingredients at the gastrointestinal treatment site and reduce the systemic drug exposure.

[0156] 2. The compound of the present invention can effectively release a JAK inhibitor (such as tofacitinib) and berberrubine or its analogues in the intestine for synergistically treating gastrointestinal autoimmune inflammatory diseases.

[0157] The present invention will be further described below in conjunction with specific embodiments. It should be understood that these embodiments are only used to illustrate the present invention and not to limit the scope of the present invention. The experimental methods without specific conditions noted in the following embodiments are usually carried out under conventional conditions or according to the conditions recommended by the manufacturer. Unless otherwise specified, percentages and parts are calculated by weight.

[0158] In each of the embodiments:

[0159] Analysis method I

[0160] LCMS instrument: waters Acquity UPLC-MS, UV detector: Acquity UPLC

[0161] Chromatographic column: Acquity UPLC HSS T3 1.8uM, column temperature 40°C

[0162] Mobile phase: A: H2O (0.1% TFA), B: acetonitrile, gradient elution

[0163] Intermediate A: (10-Methoxy-9-((methyl(2-(methyl(((10-oxo-10-((5-(4,7,10,10-tetrameth yl-3,8-dioxo-2,9-dioxo-4,7-diazaundecyl)-2-(((2S,3R,4S,5S,6S)-3,4,5-triethyl acetoxy-6-(methylester <methoxycarbonyl>)tetrahydro-2H-pyran-2-yl)oxy)phenyl)amino)decyl)oxy)carbonyl)amino yl)ethyl)carbamoyl)oxy)-5,6-dihydro-[1,3]dioxolo[4,5-g]isoquinolino[3,2-a]isoquinolin-7- ium) is prepared according to the steps shown in the following formula:

[0164]

[0165] Intermediate A-1: (2S,3R,4S,5S,6S)-2-(4-Formyl-2-nitrophenoxy)-6-(methylester <methoxycarb onyl>)tetrahydro-2H-pyran-3,4,5-triyl triacetate

[0166]

[0167] Under light - avoiding conditions, the reactants (2R,3R,4S,5S,6S)-2 - bromo - 6-(methylester group <methoxycarbonyl>) tetrahydro - 2H - pyran - 3,4,5 - triyl triacetate (300 g, 755 mmol), the reactant 4 - hydroxy - 3 - nitrobenzaldehyde (214.6 g, 1284 mmol) and silver oxide (788 g, 3400 mmol) were added to 4 L of acetonitrile and stirred at 25 - 30 °C for 5 hours. LCMS monitored the complete conversion of the raw materials. The reaction solution was filtered, and the filtrate was concentrated under reduced pressure to obtain the crude product. The crude product was diluted with ethyl acetate, filtered, and the filtrate was washed with saturated sodium bicarbonate solution and saturated brine respectively, separated, and the organic phase was dried over anhydrous sodium sulfate. The organic phase was concentrated under reduced pressure to obtain the title compound (295 g, 81%), which was a yellow solid.

[0168] MS(ESI): m / z = 506.1[M + Na] + .

[0169] Intermediate A-2: (2S,3R,4S,5S,6S)-2-(4-(Hydroxymethyl)-2-nitrophenoxy)-6-(methylester <methoxy carbonyl>)tetrahydro-2H-pyran-3,4,5-triyl triacetate

[0170]

[0171] Intermediate A - 1 (46.5 g, 96 mmol) and 19 g of silica gel were added to 450 mL of dichloromethane and 90 mL of isopropanol. The reaction was cooled to 0 °C, and 5.5 g of sodium borohydride was slowly added. The reaction solution was stirred at 0 °C for 2 hours. LCMS monitored the complete conversion of the raw materials. The reaction solution was filtered, saturated ammonium chloride solution (200 mL) was added to the filtrate, after separation, the organic phase was washed twice with saturated brine (300 mL), dried over anhydrous sodium sulfate, and concentrated under reduced pressure to obtain the crude product. The crude product was slurried with methyl tert - butyl ether to obtain the title compound (340 g, 72.9%), which was a white solid.

[0172] MS(ESI): m / z = 508.1[M + Na] + .

[0173] Intermediate A-3: (2S,3S,4S,5R,6S)-2-(methylester <methoxycarbonyl>)-6-(2-nitro-4-(4,7,10, 10-tetramethyl-3,8-dioxo-2,9-dioxo-4,7-diazaundecyl)phenoxy)tetrahydro-2H-pyran-3,4,5- triyl triacetate

[0174]

[0175] Intermediate A-2 (150 g, 310 mmol) and triethylamine (62.4 g, 620 mmol) were added to 1.5 L of dichloromethane. 4-Nitrophenyl chloroformate (71.6 g, 350 mmol) was dissolved in 300 mL of dichloromethane and added dropwise to the reaction solution under nitrogen protection at 0 °C. After the addition was complete, the reaction solution was stirred at 25 °C for 6 hours. LCMS monitored that the raw materials were completely converted. tert-Butyl methyl(2-(methylamino)ethyl)carbamate (75.8 g, 400 mmol) was added dropwise to the reaction solution of the previous step at 0 °C. After the addition was complete, the reaction solution was stirred at 25 °C for 16 hours. LCMS monitored that the raw materials were completely converted. The reaction solution was cooled to 0 °C, 1 L of saturated sodium bicarbonate solution was added, and the organic phase was separated and collected. The organic phase was washed with saturated sodium bicarbonate solution (800 mL * 8), saturated brine (800 mL), dried over anhydrous sodium sulfate, and filtered. The organic phase was concentrated under reduced pressure to obtain the target compound (200 g, 92%).

[0176] MS(ESI): m / z = 722.2 [M+Na] + .

[0177] Intermediate A-4: (2S,3R,4S,5S,6S)-2-(2-Amino-4-(4,7,10,10-tetramethyl-3,8-dioxo- 2,9-dioxo-4,7-diazaundecyl)phenoxy)-6-(methylester <methoxycarbonyl>)tetrahydro-2H-pyran-3,4,5- triyl triacetate

[0178]

[0179] Intermediate A-3 (200 g, 285.8 mmol) was dissolved in 2 L of methanol and 550 mL of water, and iron powder (80 g, 1429.1 mmol) and ammonium chloride (153 g, 2858.1 mmol) were slowly added. The reaction solution was stirred at 70 °C under nitrogen protection for 5 hours. LCMS monitored that the raw materials were completely converted. The reaction solution was filtered, and the filter cake was washed with 2 L of ethyl acetate. The organic phase was concentrated under reduced pressure to obtain the crude product. 2 L of ethyl acetate and 1.5 L of water were added to the crude product, and liquid separation was performed. The organic phase was washed with saturated brine three times (500 mL each time). After liquid separation, the organic phase was dried over anhydrous sodium sulfate and concentrated under reduced pressure to obtain the crude product. The crude product was purified by column chromatography (dichloromethane:methanol = 40:1) to obtain the target compound (100 g, 52%), which was a yellow oil.

[0180] MS(ESI): m / z = 670.2 [M+H] + .

[0181] 1HNMR(400MHz, DMSO-d6) δ 6.83 (d, J = 8.2 Hz, 1H), 6.66 (s, 1H), 6.50 (d, J = 8.2 Hz, 1H), 5.52–5.43 (m, 2H), 5.12–5.03 (m, 2H), 4.86 (s, 2H), 4.68 (d, J = 10.0 Hz, 3H), 3.64 (s, 3H), 3.31–3.25 (m, 3H), 2.79 (dd, J = 38.7, 13.2 Hz, 7H), 2.02 (d, J = 12.9 Hz, 9H), 1.36 (s, 9H).

[0182] Intermediate A-5: (2S,3R,4S,5S,6S)-2-(2-(10-Hydroxydecanoylamino)-4-(4,7,10,10-tetram ethyl-3,8-dioxo-2,9-dioxo-4,7-diazaundecyl)phenoxy)-6-(methylester <methoxycarbonyl>)tetrahydro- 2H-pyran-3,4,5-triyl triacetate

[0183]

[0184] Dissolve intermediate A-4 (100 g, 150 mmol) in N,N-dimethylformamide (600 mL). Add triethylamine (51.8 mL, 0.37 mmol) and reactant 10-hydroxydecanoic acid (39.3 g, 210 mmol) to it, and then slowly add O-(7-azabenzotriazol-1-yl)-N,N,N,N-tetramethyluronium hexafluorophosphate (79.5 g, 210 mmol). After adding the materials, under nitrogen protection, stir the reaction solution at 50 °C for 16 hours. Stop the reaction, and dilute the reactant with 2 L of ethyl acetate. Wash the diluted reaction solution with water (1.5 L × 8) and brine (1.5 L × 2), dry over anhydrous sodium sulfate, and concentrate under reduced pressure to obtain the crude product. Purify the crude product by column chromatography (dichloromethane:methanol = 20:1) to obtain the title compound (47.0 g, 37%).

[0185] MS(ESI): m / z = 862.2 [M+Na] + .

[0186] Intermediate A-6: (2S,3S,4S,5R,6S)-2-(methylester <methoxycarbonyl>)-6-(2-(10-((methyl(2-(meth ylamino)ethyl)carbamoyl)oxy)decanoylamino)-4-(4,7,10,10-tetramethyl-3,8-dioxo-2,9-dioxo -4,7-diazaundecyl)phenoxy)tetrahydro-2H-pyran-3,4,5-triyl triacetate

[0187]

[0188] Intermediate A-5 (46.0 g, 57.7 mmol) and triethylamine (15.2 mL, 109.8 mmol) were dissolved in 400 mL of dichloromethane. Under 0 °C and nitrogen protection, a solution of 4-nitrophenyl chloroformate (14.3 g, 71.2 mmol) in dichloromethane (60 mL) was added dropwise to the reaction solution. After the addition, the reaction solution was stirred at 25 - 30 °C for 16 hours. LCMS monitored the completion of raw material conversion. Triethylamine (22.8 mL, 164.4 mmol) was added to the reaction solution of the previous step. Then N1,N2-dimethylethane-1,2-diamine (14.5 g, 164.4 mmol) was added dropwise to the reaction solution of the previous step under 0 °C and nitrogen protection. After the addition, it was stirred at 25 - 30 °C for 4 hours. LCMS monitored the completion of the reaction, and the reaction solution was diluted with dichloromethane (800 mL). The diluted organic phase was washed with saturated sodium bicarbonate solution (600 mL * 3) and brine (700 mL), dried over anhydrous sodium sulfate, and concentrated under reduced pressure to obtain the crude product. The crude product was purified by normal-phase column chromatography (dichloromethane:methanol = 10:1) to obtain the title compound (36.0 g, 68%), which was a yellow oil.

[0189] MS(ESI): m / z = 954.5 [M+H] + .

[0190] Intermediate A-7: (2S,3R,4S,5S,6S)-2-(2-(10-(((2-((chlorocarbonyl)(methyl)amino)ethyl)(methyl ((4,7,10,10-tetramethyl-3,8-dioxo-2,9-dioxo-4,7-diazaundecyl)amino)carbonyl)oxo)decylamino)-4-(4,7,10,10-tetramethyl-3,8-dioxo-2,9-dioxo-4,7-diaza undecyl)phenoxy)-6-(methylester group <methoxycarbonyl>)tetrahydro-2H-pyran-3,4,5-triyltriacetate

[0191]

[0192] Triphosgene (11.2 g, 37.7 mmol) was dissolved in a three-necked flask with 100 mL of dichloromethane. Intermediate A-6 (36.0 g, 37.7 mmol) was dissolved in 300 mL of dichloromethane and added dropwise to the triphosgene dichloromethane solution under 0 °C and nitrogen protection. After the addition, the reaction solution was stirred at room temperature for 10 minutes, and then triethylamine (15.7 mL, 113.2 mmol) was added dropwise to the reaction solution under 0 °C and nitrogen protection. After the addition, the reaction solution was stirred at 25 - 30 °C for 3 hours. LCMS monitored the complete conversion of the raw material. The reaction solution was cooled to 0 °C, saturated sodium bicarbonate solution (300 mL) was added, and liquid separation was carried out. The organic phase was washed with saturated sodium bicarbonate solution (300 mL * 2) and saturated brine (200 mL). After drying over anhydrous sodium sulfate and concentrating under reduced pressure, the target compound (46.0 g of crude product) was obtained and directly used for the next reaction without further purification.

[0193] MS(ESI): m / z = 1038.3 [M+Na] + .

[0194] Intermediate A-8: 10-methoxy-5,6-dihydro-[1,3]dioxolo[4,5-g]isoquinolino[3,2-a]isoquinolin-7-ium-9-olate

[0195]

[0196] Berberine hydrochloride (35.0 g, 94.3 mmol) was placed in a round-bottom flask and heated to 180 °C under vacuum with an oil pump. After 4 hours, it was cooled to room temperature. The crude product was triturated with ethanol, filtered, and dried to obtain the title compound (23.0 g, 73%), which was a red solid.

[0197] MS(ESI): m / z = 322.1 [M] + .

[0198] Intermediate A: 10-methoxy-9-((methyl(2-(methyl(((10-oxo-10-((5-(4,7,10,10-tetra methyl-3,8-dioxo-2,9-dioxo-4,7-diazaundecyl)-2-(((2S,3R,4S,5S,6S)-3,4,5-trieth anoyloxy-6-(methylester group <methoxycarbonyl>)tetrahydro-2H-pyran-2-yl)oxo)phenyl)amino)decyl)oxo)carbonyl)am ((ethyl)carbamoyl)oxo)-5,6-dihydro-[1,3]dioxolo[4,5-g]isoquinolino[3,2-a]isoquinolin-7- ium

[0199]

[0200] Intermediate A-8 (9.6 g, 30.0 mmol) was dissolved in 100 mL of pyridine in a three-necked flask. Intermediate A-7 (46.0 g, 45.0 mmol) was dissolved in 300 mL of pyridine and added dropwise to the three-necked flask at 0 °C under nitrogen protection. The reaction mixture was stirred at 25 °C for 16 hours. LCMS was used to monitor the completion of the conversion of the starting materials. The reaction mixture was concentrated under reduced pressure, and the crude product was purified by column chromatography (dichloromethane:methanol = 10:1) to obtain a crude black solid product. The crude product was further purified by normal-phase column chromatography (dichloromethane:methanol = 3:2) to obtain the title compound (11.0 g, 22%).

[0201] MS(ESI): m / z = 601.6 (M - 100 + H / 2) + .

[0202] 1 HNMR(400 MHz, CDCl3) δ 11.24–10.55 (m, 1H), 8.45 (s, 2H), 7.83 (dd, J = 35.6, 27.7 Hz, 3H), 7.42 (d, J = 20.4 Hz, 1H), 7.21–6.74 (m, 3H), 6.27 (d, J = 4.1 Hz, 0H), 6.08 (s, 2H), 5.78 (s, 0H), 5.57–5.23 (m, 6H), 5.06 (d, J = 11.4 Hz, 2H), 4.23–3.95 (m, 5H), 3.78 (d, J = 18.4 Hz, 3H), 3.54–2.73 (m, 20H), 2.34 (t, J = 23.9 Hz, 2H), 2.09 (dt, J = 9.8, 4.0 Hz, 8H), 1.88–0.86 (m, 27H).

[0203] Intermediate B: 2-hydroxy-5-((4-(4,7,10,10-tetramethyl-3,8-dioxo-2,9-dioxo-4,7-diazacy clo)phenyl)diazenyl)benzoic acid

[0204]

[0205] Intermediate B-1: 2-hydroxy-5-((4-(hydroxymethyl)phenyl)diazenyl)benzoic acid

[0206]

[0207] A suspension of 4-aminobenzyl alcohol (2.0 g, 16.2 mmol) in water (30 mL) at 0 °C was treated with 3.4 mL of concentrated hydrochloric acid, and then an aqueous solution of ice-cold NaNO2 (1.2 g, 17.0 mmol, 8 mL) was added slowly. After stirring at 0 °C for 1 hour, the above reaction solution was added to an aqueous solution (25 mL) of sodium 2-hydroxybenzoate (2.72 g, 0.35 mmol) and potassium carbonate (3.2 g, 22.7 mmol). During the whole dropping process, the pH value of the reaction solution was maintained at 13 - 14 by dropping aqueous sodium hydroxide solution. The mixture was stirred at room temperature for 1 hour, the pH was adjusted to 4 - 5 with hydrochloric acid (2N), the product precipitated out, the precipitate was filtered and washed with water (50 mL), and dried in vacuo to obtain the title compound (4.0 g, 90%), as a red solid.

[0208] MS(ESI): m / z = 272.8 [M + H] + .

[0209] Intermediate B: 2-hydroxy-5-((4-(4,7,10,10-tetramethyl-3,8-dioxo-2,9-dioxo-4,7-diazacy clo)phenyl)diazenyl)benzoic acid

[0210]

[0211] Intermediate B-1 (200 mg, 0.73 mmol) and diisopropylethylamine (114 mg, 0.88 mmol) were dissolved in 5 mL of dichloromethane, and bis(4-nitrophenyl) carbonate (268 mg, 0.88 mmol) was added. The reaction solution was stirred at room temperature for 48 hours. tert-Butyl methyl(2-(methylamino)ethyl)carbamate (165 mg, 0.88 mmol) and diisopropylethylamine (114 mg, 0.88 mmol) were added dropwise to the above reaction solution at 0 °C. After the addition was completed, the reaction solution was stirred at 25 °C for 2 hours. LCMS monitored that the raw materials were completely converted. The reaction solution was concentrated under reduced pressure, and the target compound (195 mg, 55%) was obtained by reverse-phase column chromatography, as a red solid.

[0212] MS(ESI): m / z = 508.9 [M + Na] + .

[0213] 11H NMR (400 MHz, CDCl3) δ 8.29 (d, J = 2.4 Hz, 1H), 8.04 (dd, J1 = 2.4 Hz, J2 = 8.8 Hz, 1H), 7.82 (d, J = 8.4 Hz, 2H), 7.50 (d, J = 8.0 Hz, 2H), 7.11 (d, J = 9.2 Hz, 1H), 5.10 (s, 2H), 3.35 - 3.32 (m, 4H), 2.88 - 2.82 (m, 3H), 2.73 - 2.68 (m, 3H), 1.32 (s, 9H).

[0214] Intermediate C: 9-(((2-(13-carboxytridecanoylamino)ethyl)(methyl)carbamoyl)oxo)-10-meth oxy-5,6-dihydro-[1,3]dioxolo[4,5-g]isoquinolino[3,2-a]isoquinolin-7-ium

[0215]

[0216] Intermediate C-1: 9-(((2-((tert-butoxycarbonyl)amino)ethyl)(methyl)carbamoyl)oxo)-10-meth oxy-5,6-dihydro-[1,3]dioxolo[4,5-g]isoquinolino[3,2-a]isoquinolin-7-ium

[0217]

[0218] Dissolve tert-butyl (2-(methylamino)ethyl)carbamate (15.0 g, 86.0 mmol) in dichloromethane (200 mL). Slowly add triphosgene (25.6 g, 86.0 mmol) and pyridine (20.0 g, 258 mmol) successively under an ice bath. Stir at room temperature (15 °C) for 1 hour. Monitor the reaction by TLC until the raw materials are completely reacted. Wash the reaction solution with water (200 mL), and extract the aqueous phase with dichloromethane (100 mL × 2). Combine the organic phases, wash with saturated brine, dry the organic phase over anhydrous sodium sulfate, filter, and concentrate under reduced pressure to obtain the intermediate. Dissolve the intermediate in pyridine (20 mL), and add it to intermediate A-8 (27.7 g, 86.0 mmol) dissolved in pyridine (30 mL) under an ice bath. Raise the reaction temperature to room temperature (15 °C) and stir for 16 hours. Monitor the reaction by LCMS until the raw materials are completely reacted. Concentrate the reaction solution under reduced pressure and purify it by normal-phase column chromatography (dichloromethane:methanol = 10:1) to obtain a yellow solid product (6.5 g, 14%).

[0219] MS (ESI): m / z = 522.1 [M] + .

[0220] Intermediate C: 9-(((2-aminoethyl)(methyl)carbamoyl)oxo)-10-methoxy-5,6-dihydro-[1,3] dioxolo[4,5-g]isoquinolino[3,2-a]isoquinolin-7-ium hydrochloride

[0221]

[0222] Intermediate C-1 (4.5 g, 8.6 mmol) was mixed in hydrochloric acid methanol solution (2 mol / L, 100 mL) and stirred overnight at room temperature (15 °C). The reaction was monitored by LCMS until completion. The reaction mixture was concentrated under reduced pressure to give the title compound (3.6 g, 100%) as a brown solid.

[0223] MS(ESI): m / z = 422.1 [M] + .

[0224] Intermediate D: 9-((4-hydroxybenzyl)oxo)-10-methoxy-5,6-dihydro-[1,3]dioxolo[4,5- g]isoquinolino[3,2-a]isoquinolin-7-ium

[0225]

[0226] To a solution of intermediate A-8 (300 mg, 0.93 mmol) in acetonitrile (3 mL) was added 4-(chloromethyl)phenyl acetate (257 mg, 1.4 mmol) and potassium carbonate (257 mg, 1.86 mmol). The reaction mixture was heated to 80 °C and stirred for 16 h. The reaction mixture was diluted with dichloromethane (50 mL), filtered, and the filter cake was washed with water (50 mL). The filter cake was purified by normal phase column chromatography (dichloromethane:methanol = 10:1) to give the title compound (97 mg, 24%) as a dark red solid.

[0227] MS(ESI): m / z = 428.1 [M] + .

[0228] Preparation of co-drug compounds

[0229] Example 1: 9-(((2-((((10-((2-(((2S,3R,4S,5S,6S)-6-carboxy-3,4,5-trihydroxytetrahydro- (2H-Pyran-2-yl)oxy)-5-((((2-(4-(((3R,4R)-1-(2-cyanoacetyl)-4-methylpiperidin-3-yl)(methyl)amino)-N-methyl-7H-pyrrolo[2,3-d]pyrimidine-7-carboxamido<oxalamido>)ethyl)(methyl)amino (methyl)carbamoyl)oxy)methyl)phenyl)amino)-10-oxodecyl)oxy)carbonyl)(methyl)amino)ethyl)(methyl)amino (methyl)carbamoyl)oxy)-10-methoxy-5,6-dihydro-[1,3]dioxolo[4,5-g]isoquinolino[3,2-a]isoquinolin-7-yl cation (methyl)carbamoyl)oxy)-10-methoxy-5,6-dihydro-[1,3]dioxolo[4,5-g]isoquinolino[3,2-a]isoquinolin-7-yl cation (methyl)carbamoyl)oxy)-10-methoxy-5,6-dihydro-[1,3]dioxolo[4,5-g]isoquinolino[3,2-a]isoquinolin-7-yl cation

[0230]

[0231] (methyl)carbamoyl)oxy)-10-methoxy-5,6-dihydro-[1,3]dioxolo[4,5-g]isoquinolino[3,2-a]isoquinolin-7-yl cation (methyl)carbamoyl)oxy)-10-methoxy-5,6-dihydro-[1,3]dioxolo[4,5-g]isoquinolino[3,2-a]isoquinolin-7-yl cation

[0232]

[0233] Tofacitinib (8.9 g, 28.6 mmol) was dissolved in dichloromethane solution (140 mL), and an aqueous solution (48 mL) of sodium hydroxide (3.4 g, 85.6 mmol) and tetrabutylammonium bromide (920 mg, 2.86 mmol) was added. The above reaction solution was added dropwise to a dichloromethane (48 mL) solution of p-nitrophenyl chloroformate (11.5 g, 57.1 mmol). After the addition was completed, the reaction solution was stirred at room temperature for 4 h. LCMS monitored that the raw materials were completely converted. 500 mL of dichloromethane was added for dilution, washed with saturated ammonium chloride (200 mL), and the insoluble substances were filtered off with diatomaceous earth. The organic phase was separated from the filtrate and then washed with 200 mL of saturated brine. The organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was slurried with dichloromethane and petroleum ether for 5 times to obtain the title compound (16.3 g, 85%), a yellow foamy solid.

[0234] MS(ESI): m / z=478.1[M+H] + .

[0235] (methyl)carbamoyl)oxy)-10-methoxy-5,6-dihydro-[1,3]dioxolo[4,5-g]isoquinolino[3,2-a]isoquinolin-7-yl cation (methyl)carbamoyl)oxy)-10-methoxy-5,6-dihydro-[1,3]dioxolo[4,5-g]isoquinolino[3,2-a]isoquinolin-7-yl cation (methyl)carbamoyl)oxy)-10-methoxy-5,6-dihydro-[1,3]dioxolo[4,5-g]isoquinolino[3,2-a]isoquinolin-7-yl cation (methyl)carbamoyl)oxy)-10-methoxy-5,6-dihydro-[1,3]dioxolo[4,5-g]isoquinolino[3,2-a]isoquinolin-7-yl cation

[0236]

[0237] Trifluoroacetic acid (10 mL) was added to a dichloromethane (40 mL) solution of intermediate A (4.8 g, 3.7 mmol), and the mixture was stirred at room temperature for 1 h. LCMS monitored that the reaction was complete. The reaction solution was concentrated under reduced pressure, dissolved in dichloromethane (30 mL), concentrated under reduced pressure again to remove trifluoroacetic acid as much as possible, and dried with an oil pump to obtain the title compound (4.43 g, 100%), a yellow oil.

[0238] MS(ESI): m / z=601.4[M / 2] + .

[0239] (methyl)carbamoyl)oxy)-10-methoxy-5,6-dihydro-[1,3]dioxolo[4,5-g]isoquinolino[3,2-a]isoquinolin-7-yl cation (methyl)carbamoyl)oxy)-10-methoxy-5,6-dihydro-[1,3]dioxolo[4,5-g]isoquinolino[3,2-a]isoquinolin-7-yl cation (methyl)carbamoyl)oxy)-10-methoxy-5,6-dihydro-[1,3]dioxolo[4,5-g]isoquinolino[3,2-a]isoquinolin-7-yl cation (methyl)carbamoyl)oxy)-10-methoxy-5,6-dihydro-[1,3]dioxolo[4,5-g]isoquinolino[3,2-a]isoquinolin-7-yl cation (methyl)carbamoyl)oxy)-10-methoxy-5,6-dihydro-[1,3]dioxolo[4,5-g]isoquinolino[3,2-a]isoquinolin-7-yl cation (methyl)carbamoyl)oxy)-10-methoxy-5,6-dihydro-[1,3]dioxolo[4,5-g]isoquinolino[3,2-a]isoquinolin-7-yl cation

[0240]

[0241] A solution of Example 1-10 (4.4 g, 3.7 mmol) in dichloromethane (100 mL) was cooled to 0 °C, N,N-diisopropylethylamine (1.56 g, 12 mmol) was added, and Example 1-11 (1.76 g, 3.69 mmol) was added. The mixture was stirred at room temperature for 1 hour. The reaction was monitored by LCMS and was found to be complete. The reaction mixture was diluted with 300 mL of dichloromethane, washed successively with water and saturated brine, and the washed organic phase was dried over anhydrous sodium sulfate and concentrated under reduced pressure. The crude product was separated by normal-phase column chromatography (eluted with dichloromethane containing 7%-9% methanol) to obtain the title compound (2.83 g, 53%), which was a yellow foamy solid.

[0242] MS(ESI): m / z = 770.7 [M / 2] + .

[0243] (methyl)carbamoyl)oxy)-10-methoxy-5,6-dihydro-[1,3]dioxolo[4,5-g]isoquinolino[3,2-a]isoquinolin-7-yl cation (methyl)carbamoyl)oxy)-10-methoxy-5,6-dihydro-[1,3]dioxolo[4,5-g]isoquinolino[3,2-a]isoquinolin-7-yl cation (methyl)carbamoyl)oxy)-10-methoxy-5,6-dihydro-[1,3]dioxolo[4,5-g]isoquinolino[3,2-a]isoquinolin-7-yl cation ((Methylcarbamoyl)oxy)methyl)phenyl)amino)-10-oxodecyl)oxo)carbonyl)(methyl)amino)ethyl)(methyl)amino (Formyl)oxy)-10-methoxy-5,6-dihydro-[1,3]dioxolo[4,5-g]isoquinolino[3,2-a]isoquinolin-7-ium Ion

[0244]

[0245] A solution of Example 1-12 (163 mg, 0.105 mmol) in methanol (4 mL) was cooled to 0 °C, and an aqueous potassium carbonate solution (1 mmol / mL, 1 mL) was added. The mixture was stirred at 0 °C for 2 hours. The reaction was monitored by LCMS and was found to be complete. The pH of the reaction mixture was adjusted to 5 with acetic acid and then concentrated under reduced pressure. The crude product was separated by Prep-HPLC (gradient elution with acetonitrile / water) to obtain the title compound (35.7 mg, 24%), which was a yellow solid.

[0246] MS(ESI): m / z = 1399.5 [M] + .

[0247] 1 1H NMR: (400 MHz, CD3OD) δ 10.06 - 9.59 (m, 1H), 8.71 - 8.61 (m, 1H), 8.15 - 7.98 (m, 4H), 7.60 - 7.50 (m, 1H), 7.26 - 7.17 (m, 1H), 6.95 - 6.72 (m, 3H), 6.72 - 6.66 (m, 1H), 6.05 (s, 2H), 5.05 - 4.90 (m, 4H), 4.78 - 4.59 (m, 2H), 4.21 - 4.08 (m, 2H), 4.04 (s, 3H), 3.96 - 3.35 (m, 18H), 3.21 - 2.95 (m, 13H), 2.29 - 1.84 (m, 3H), 1.84 - 1.02 (m, 20H).

[0248] The following compounds were obtained by using a method similar to that of Example 1 and replacing the corresponding raw materials.

[0249]

[0250] Example 4A: 9-((5-((E)-(4-((((2-(4-(((3R,4R)-1-(2-cyanoacetyl)-4-methylpiperidin-3-yl)(methyl)amino)-N-methyl-7H-pyrrolo[2,3-d]pyrimidine-7-carboxamido<oxalamido>)ethyl)(methyl)aminocarbonyl)oxy)methyl)phenyl)diazenyl)-2-hydroxybenzoyl)oxy)-10-methoxy-5,6- Dihydro-[1,3]dioxolo[4,5-g]isoquinolino[3,2-a]isoquinolin-7-ium ((Methylcarbamoyl)oxy)methyl)phenyl)diazenyl)-2-hydroxybenzoyl)oxy)-10-methoxy-5,6- Dihydro-[1,3]dioxolo[4,5-g]isoquinolino[3,2-a]isoquinolin-7-ium

[0251] Example 4B: 9-((5-((Z)-(4-((((2-(4-(((3R,4R)-1-(2-cyanoacetyl)-4-methylpiperidin-3-yl)(methyl)amino)-N-methyl-7H-pyrrolo[2,3-d]pyrimidine-7-carboxamido<oxalamido>)ethyl)(methyl)aminocarbonyl)oxy)methyl)phenyl)diazenyl)-2-hydroxybenzoyl)oxy)-10-methoxy-5,6- ((Methylcarbamoyl)oxy)methyl)phenyl)diazenyl)-2-hydroxybenzoyl)oxy)-10-methoxy-5,6- Dihydro-[1,3]dioxolo[4,5-g]isoquinolino[3,2-a]isoquinolin-7-ium Dihydro-[1,3]dioxolo[4,5-g]isoquinolino[3,2-a]isoquinolin-7-ium

[0252]

[0253] Example 4-1: (E)-9-((2-hydroxy-5-((4-(4,7,10,10-tetramethyl-3,8-dioxo-2,9-dioxa-4,7-diazoundecyl)phenyl)diazenyl)benzoyl)oxy)-10-methoxy-5,6-dihydro-[1,3]dioxo L[4,5-g]isoquinolino[3,2-a]isoquinolin-7-ium Dihydro-[1,3]dioxolo[4,5-g]isoquinolino[3,2-a]isoquinolin-7-ium

[0254]

[0255] Intermediate B (486 mg, 1.0 mmol), Intermediate A-8 (322 mg, 1.0 mmol) and dicyclohexylcarbodiimide (247 mg, 1.2 mmol) were placed in a single-necked flask, and dichloromethane (10 mL) was added. After stirring at room temperature for 1 hour and monitoring the reaction by LCMS until completion, the reaction solution was filtered, concentrated under reduced pressure, and the crude product was separated by reverse-phase column chromatography to obtain the title compound (156 mg, 19.7%), which was a brown oil.

[0256] MS (ESI): m / z = 790.1 [M] + .

[0257] Example 4A: 9-((5-((E)-(4-((((2-(4-(((3R,4R)-1-(2-cyanoacetyl)-4-methylpiperidin-3-yl)(methyl)amino)-N-methyl-7H-pyrrolo[2,3-d]pyrimidine-7-carboxamido<oxalamido>)ethyl)(methyl)aminocarbonyl)oxy)methyl)phenyl)diazenyl)-2-hydroxybenzoyl)oxy)-10-methoxy-5,6- ((Methylcarbamoyl)oxy)methyl)phenyl)diazenyl)-2-hydroxybenzoyl)oxy)-10-methoxy-5,6- Dihydro-[1,3]dioxolo[4,5-g]isoquinolino[3,2-a]isoquinolin-7-ium And

[0258] Example 4B: 9-((5-((Z)-(4-((((2-(4-(((3R,4R)-1-(2-cyanoacetyl)-4-methylpiperidin-3-yl)(methyl)amino)-N-methyl-7H-pyrrolo[2,3-d]pyrimidine-7-carboxamido<oxalamido>)ethyl)(methyl)aminocarbonyl)oxy)methyl)phenyl)diazenyl)-2-hydroxybenzoyl)oxy)-10-methoxy-5,6- ((Methylcarbamoyl)oxy)methyl)phenyl)diazenyl)-2-hydroxybenzoyl)oxy)-10-methoxy-5,6- Dihydro-[1,3]dioxolo[4,5-g]isoquinolino[3,2-a]isoquinolin-7-ium Dihydro-[1,3]dioxolo[4,5-g]isoquinolino[3,2-a]isoquinolin-7-ium

[0259]

[0260] To a solution of Example 4-1 (156 mg, 0.19 mmol) in dichloromethane (2 mL) was added trifluoroacetic acid (0.4 mL), and the mixture was stirred at room temperature for 20 minutes. The reaction was monitored by LCMS until completion. The reaction solution was concentrated under reduced pressure, dried by an oil pump, dissolved in dichloromethane (2 mL) solution, cooled to 0 °C, N,N-diisopropylethylamine (101 mg, 0.78 mmol) was added, Example 1-11 (94 mg, 0.19 mmol) was added, and the mixture was stirred at room temperature for 1 hour. The reaction was monitored by LCMS until completion. The reaction solution was concentrated under reduced pressure, and the crude product was separated by Prep-HPLC (gradient elution with acetonitrile / water (containing 0.1% trifluoroacetic acid)) to obtain the title compound 4A (6.0 mg, 3.0%), as a yellow solid; 4B (6.0 mg, 3.0%), as a yellow solid.

[0261] MS(ESI): m / z = 1028.4 [M] + .

[0262] Example 5: 9-(((2-(5-((E)-(4-((((2-(4-(((3R,4R)-1-(2-cyanoacetyl)-4-methyl piperidin-3-yl)(methyl)amino)-N-methyl-7H-pyrrolo[2,3-d]pyrimidine-7-carboxamido<oxalamido>) ethyl)(methyl)carbamoyl)oxo)methyl)phenyl)diazenyl)-2-hydroxybenzamido)ethyl)(methyl)amino carbamoyl)oxo)-10-methoxy-5,6-dihydro-[1,3]dioxolo[4,5-g]isoquinolino[3,2-a]isoquinolin-7- ium

[0263]

[0264] Example 5-1: (E)-9-(((2-(2-hydroxy-5-((4-(4,7,10,10-tetramethyl-3,8-dioxo-2,9- dioxa-4,7-diazoundecyl)phenyl)diazenyl)benzamido)ethyl)(methyl)carbamoyl)oxo)- 10-methoxy-5,6-dihydro-[1,3]dioxolo[4,5-g]isoquinolino[3,2-a]isoquinolin-7-ium

[0265]

[0266] Intermediate C (700 mg, 1.65 mmol), Intermediate B (782 mg, 1.65 mmol) and 1-(3-

[0267] dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (477 mg, 2.48 mmol) were placed in a single-necked flask, N,N-dimethylformamide (5 mL) was added, and 4-dimethylaminopyridine (50 mg, 0.41 mmol) was added. The mixture was stirred at room temperature for 1 hour, and diisopropylethylamine (427 mg, 3.31 mmol) was added. The reaction solution was stirred at room temperature for 5 hours.

[0268] After monitoring the reaction by LCMS until completion, 1N hydrochloric acid was added to quench the reaction, and the reaction solution was directly injected for separation by reversed-phase preparation to obtain the title compound (470 mg, 32%), as a yellow solid.

[0269] MS(ESI): m / z = 890.3 [M] + .

[0270] Example 5-2: ((E)-9-(((2-(2-hydroxy-5-((4-(((methyl(2-(methylamino)ethyl)amino carbamoyl)oxo)methyl)phenyl)diazenyl)benzamido)ethyl)(methyl)carbamoyl)oxo)-10-methoxy-5, 6-dihydro-[1,3]dioxolo[4,5-g]isoquinolino[3,2-a]isoquinolin-7-ium

[0271]

[0272] To a solution of Example 5-1 (470 mg, 0.53 mmol) in methanol (2 mL) was added ethyl acetate hydrochloride solution (4 mol / L, 2 mL), and the mixture was stirred at room temperature for 1 hour. The reaction was monitored by LCMS until completion. The reaction solution was concentrated under reduced pressure, dissolved in dichloromethane (10 mL), and concentrated under reduced pressure again. This process was repeated twice, and then dried under vacuum with an oil pump to obtain the title compound (390 mg, 93.5%) as a yellow solid.

[0273] MS(ESI): m / z = 790.2 [M+H] + .

[0274] Example 5: 9-(((2-(5-((E)-(4-((((2-(4-(((3R,4R)-1-(2-cyanoacetyl)-4-methyl piperidin-3-yl)(methyl)amino)-N-methyl-7H-pyrrolo[2,3-d]pyrimidine-7-carboxamido<oxalamido>) ethyl)(methyl)carbamoyl)oxo)methyl)phenyl)diazenyl)-2-hydroxybenzamido)ethyl)(methyl)amino carbamoyl)oxo)-10-methoxy-5,6-dihydro-[1,3]dioxolo[4,5-g]isoquinolino[3,2-a]isoquinolin-7- ium

[0275]

[0276] A solution of Example 5-2 (390 mg, 0.49 mmol) in dichloromethane (5 mL) was cooled to 0 °C, N,N-diisopropylethylamine (254 mg, 1.97 mmol) was added, and then Example 1-11 (235 mg, 0.49 mmol) was added. The mixture was stirred at room temperature for 1 hour. The reaction was monitored by LCMS until completion. The reaction solution was diluted with dichloromethane (50 mL), washed successively with water (50 mL) and saturated brine (50 mL), dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was purified by normal-phase column chromatography (using dichloromethane and methanol with 0.1% formic acid as the eluent) to obtain the title compound (114 mg, 20%) as a yellow solid.

[0277] MS(ESI): m / z = 1128.4 [M+H] + .

[0278] 1 H NMR: (400 MHz, CD3OD) δ 9.79 - 9.59 (m, 1H), 8.59 - 7.87 (m, 7H), 7.65 - 7.40 (m, 4H), 7.03 - 6.83 (m, 3H), 6.70 - 6.44 (m, 1H), 6.08 (s, 2H), 5.32 - 4.94 (m, 3H), 3.91 - 3.72 (m, 6H), 3.62 - 3.43 (m, 9H), 3.35 (s, 3H), 3.22 - 2.66 (m, 13H), 2.41 - 2.26 (m, 1H), 1.84 - 1.52 (m, 2H), 1.36 - 1.26 (m, 4H), 1.03 - 0.85 (m, 3H).

[0279] Example 6: 9-(((2-(6-(5-((E)-(4-((((2-(4-(((3R,4R)-1-(2-cyanoacetyl)-4-meth ylpiperidin-3-yl)(methyl)amino)-N-methyl-7H-pyrrolo[2,3-d]pyrimidine-7-carboxamido<oxalamido >)ethyl)(methyl)carbamoyl)oxo)methyl)phenyl)diazenyl)-2-hydroxybenzamido)hexanamido)eth yl)(methyl)carbamoyl)oxo)-10-methoxy-5,6-dihydro-[1,3]dioxolo[4,5-g]isoquinolino[3,2- a]isoquinolin-7-ium

[0280]

[0281] Example 6-1: Methyl (E)-6-(2-hydroxy-5-((4-(4,7,10,10-tetramethyl-3,8-dioxo-2,9-dioxo a-4,7-diazoundecyl)phenyl)diazenyl)benzamido)hexanoate

[0282]

[0283] Dissolve intermediate B (1900 mg, 3.9 mmol), methyl 6-aminohexanoate hydrochloride (849 mg, 4.6 mmol) and diisopropylethylamine (2017 mg, 15.6 mmol) in N,N-dimethylformamide (19 mL), and add O-(7-azabenzotriazol-1-yl)-N,N,N',N'-tetramethyluronium hexafluorophosphate (2228 mg, 5.86 mmol). Stir at 30 °C for 3 hours. After monitoring the completion of the reaction by LCMS, dilute the reaction solution with ethyl acetate (150 mL), wash it 4 times with water (100 mL), wash it once with saturated brine (100 mL), dry it over anhydrous sodium sulfate, filter, and concentrate it under reduced pressure. The crude product was separated by normal-phase column chromatography (petroleum ether:ethyl acetate = 2:1) to obtain the title compound (870 mg,

[0284] 36.3%), as a yellow solid.

[0285] MS (ESI): m / z = 636.2 [M+Na] + .

[0286] Example 6-2: (E)-6-(2-hydroxy-5-((4-(4,7,10,10-tetramethyl-3,8-dioxo-2,9-dioxo a-4,7-diazoundecyl)phenyl)diazenyl)benzamido)hexanoic acid

[0287]

[0288] Dissolve Example 6-1 (800 mg, 1.3 mmol) in methanol (5 mL) and water (2 mL), and add lithium hydroxide monohydrate (247 mg, 6.5 mmol). Stir the reaction solution at 65 °C for 1 hour. After monitoring the completion of the reaction by LCMS, cool it to room temperature, neutralize it to pH 4-5 with dilute hydrochloric acid, extract it with 100 mL of ethyl acetate, wash the organic phase with saturated brine, dry it over anhydrous sodium sulfate, and concentrate it under reduced pressure to obtain the title compound (784 mg, 100%), as a yellow solid.

[0289] MS (ESI): m / z = 622.2 [M+Na] + .

[0290] Example 6-3: (E)-9-(((2-(6-(2-hydroxy-5-((4-(4,7,10,10-tetramethyl-3,8-dioxo-2,9-dioxo-4,7-diazoundecyl)phenyl)diazenyl)benzamido)hexanoylamino)ethyl)(methyl)amino formyl)oxy)-10-methoxy-5,6-dihydro-[1,3]dioxolo[4,5-g]isoquinolino[3,2-a]isoquinolin-7-ium ion

[0291]

[0292] To a mixture of Example 6-2 (820 mg, 1.37 mmol), Intermediate C (693 mg, 1.64 mmol) and DMF (1 mL), O-(7-azabenzotriazol)-N,N,N,N-tetramethyluronium hexafluorophosphate (780 mg, 2.05 mmol) and diisopropylethylamine (706 mg, 5.47 mmol) were added successively. After the addition was complete, the mixture was stirred at room temperature for 0.5 h. After monitoring the reaction to completion by LCMS, the reaction solution was diluted with dichloromethane (100 mL), washed with water (50 mL × 4) and brine (50 mL × 2), dried over anhydrous sodium sulfate, and concentrated under reduced pressure to obtain a crude product. The crude product was purified by silica gel column chromatography (methanol:dichloromethane = 1:12, (containing 0.1% formic acid)) to obtain the title compound (525 mg, 38%), a yellow solid.

[0293] MS(ESI): m / z = 1003.3 [M] + .

[0294] Example 6: 9-(((2-(6-(5-((E)-(4-((((2-(4-(((3R,4R)-1-(2-cyanoacetyl)-4-methylpiperidin- 3-yl)(methyl)amino)-N-methyl-7H-pyrrolo[2,3-d]pyrimidine-7-carboxamido<oxalamido >)ethyl)(methyl)amino)carbonyl)oxy)methyl)phenyl)diazenyl)-2-hydroxybenzamido)hexanoylamino)eth yl)(methyl)amino)carbonyl)oxy)-10-methoxy-5,6-dihydro-[1,3]dioxolo[4,5-g]isoquinolino[3,2- a]isoquinolin-7-ium ion

[0295]

[0296] To a solution of Example 6-3 (525 mg, 0.52 mmol) in dichloromethane (2 mL) was added trifluoroacetic acid (0.4 mL), and the mixture was stirred at room temperature for 1 h. The reaction was monitored to completion by LCMS. The reaction solution was concentrated under reduced pressure, dried under the oil pump, dissolved in dichloromethane (2 mL), cooled to 0 °C, N,N-diisopropylethylamine (270 mg, 2.09 mmol) was added, and Example 1-11 (249 mg, 0.52 mmol) was added. The mixture was stirred at room temperature for 0.5 h. The reaction was monitored to completion by LCMS. The reaction solution was concentrated under reduced pressure, and the crude product was separated by Prep-HPLC (gradient elution with acetonitrile / water (containing 0.1% trifluoroacetic acid)) to obtain the title compound (135 mg, 21%), a yellow solid.

[0297] MS(ESI): m / z = 1241.8 [M] + .

[0298] 11H NMR: (400 MHz, CD3OD) δ 9.89 - 9.59 (m, 1H), 8.59 - 8.47 (m, 2H), 8.37 - 8.31 (m, 1H), 8.12 - 7.93 (m, 2H), 7.70 - 7.61 (m, 2H), 7.51 - 7.43 (m, 2H), 6.75 - 6.73 (m, 1H), 6.05 (s, 2H), 5.17 - 4.90 (m, 4H), 3.98 (s, 3H), 3.79 - 3.25 (m, 14H), 3.23 - 3.06 (m, 9H), 2.32 - 2.27 (m, 3H), 1.72 - 1.21 (m, 11H), 0.99 - 0.87 (m, 4H).

[0299] Example 7: 9-(((2-(14-((4-((((2-(4-(((3R,4R)-1-(2-cyanoacetyl)-4-methylpiperidin- 3-yl)(methyl)amino)-N-methyl-7H-pyrrolo[2,3-d]pyrimidine-7-carboxamido<oxalamido>)eth yl)(methyl)amino)carbonyl)oxy)methyl)phenyl)amino)-14-oxotetradecanoylamino)ethyl)(methyl)aminocarb onyl)oxy)-10-methoxy-5,6-dihydro-[1,3]dioxolo[4,5-g]isoquinolino[3,2-a]isoquinolin-7-ium ion

[0300]

[0301] Example 7-1: 4-(((tert-butyldimethylsilyl)oxy)methyl)aniline

[0302]

[0303] To a solution of (4 - aminophenyl)methanol (5.0 g, 40.6 mmol) and imidazole (3.04 g, 44.66 mmol) in dichloromethane (70 mL) was added tert - butyldimethylchlorosilane (6.12 g, 40.6 mmol), and the reaction was carried out at room temperature for 1 hour. Ethyl acetate (200 mL) was added to the reaction mixture. The organic phase was washed with water (400 mL), dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The residue was separated and purified by normal - phase column chromatography (petroleum ether:ethyl acetate = 5:1) to give the title compound (9.3 g, 95%), which is a pale yellow liquid.

[0304] MS(ESI): m / z = 238.1 [M + H] + .

[0305] Example 7-2: 14-((4-(((tert-butyldimethylsilyl)oxy)methyl)phenyl)amino)-14-oxotetradecanoic acid Example 7-3: 9-(((2-(14-((4-(((tert-butyldimethylsilyl)oxy)methyl)phenyl)amino)-14-oxotetradecanoylamino)ethyl)(methyl)aminocarbonyl)oxy)-10-methoxy-5,6-dihydro-[1,3]dioxolo[4,5-g]isoquinolino[3,2-a]isoquinolin-7-ium ion

[0306]

[0307] To a solution of Example 7-1 (4280 mg, 18.06 mmol) and sebacic acid (6989 mg, 27.09 mmol) in dichloromethane (43 mL) was added N,N-diisopropylethylamine (4659 mg, 36.12 mmol). The temperature was lowered to 0 °C and stirred for 20 minutes. 2-(7-Azabenzotriazol-1-yl)-N,N,N',N'-tetramethyluronium hexafluorophosphate (10.3 g, 27.09 mmol) was added to the reaction solution. The reaction was warmed to room temperature and stirred for 16 hours. The reaction solution was diluted with dichloromethane (20 mL) and washed with water (40 mL). The organic phase was dried over anhydrous sodium sulfate and concentrated under reduced pressure. The residue was separated and purified by normal-phase column chromatography (petroleum ether:ethyl acetate = 1:1) to give the title compound (6.7 g, 79%), which was a pale yellow solid.

[0308] MS(ESI): m / z = 500.2 [M+Na] + .

[0309] ​ ​ ​

[0310]

[0311] A solution of Example 7-2 (1800 mg, 3.77 mmol) and Intermediate C (1592 mg, 3.77 mmol) in dichloromethane (20 mL) was placed in a single-necked flask. At 0 °C, N,N-diisopropylethylamine (1947 mg, 15.09 mmol) was added. After stirring for 20 minutes, 2-(7-azabenzotriazol-1-yl)-N,N,N',N'-tetramethyluronium hexafluorophosphate (2870 mg, 7.55 mmol) was added. The reaction solution was warmed to room temperature and stirred for 40 minutes. The reaction solution was washed with water (40 mL). The organic phase was dried over anhydrous sodium sulfate and concentrated under reduced pressure. The residue was separated and purified by normal-phase column chromatography (ethyl acetate:petroleum ether = 1:1) to give the target compound (2200 mg, 66.2%), which was a white solid.

[0312] MS(ESI): m / z = 881.4 [M] + .

[0313] Example 7-4: 9-(((2-(14-((4-(Hydroxymethyl)phenyl)amino)-14-oxotetradecanamido)ethyl) (Methyl)carbamoyloxy)-10-methoxy-5,6-dihydro-[1,3]dioxolo[4,5-g]isoquinolino[3,2-a]iso quinolin-7-ium

[0314]

[0315] To tetrahydrofuran (30 mL) of Example 7-3 (2100 mg, 2.38 mmol) was added pyridinium hydrofluoride (753 mg, 9.53 mmol). The reaction was stirred at room temperature for 16 h. After monitoring the completion of the reaction by LCMS, it was diluted with dichloromethane (30 mL), washed with water, dried over anhydrous sodium sulfate, and concentrated under reduced pressure to obtain the target compound (1360 mg, 74.3%), which was a white solid.

[0316] MS(ESI): m / z = 767.3 [M] + .

[0317] Example 7-5: 10-Methoxy-9-((methyl(2-(14-oxo-14-((4-(4,7,10,10-tetramethyl-3,8- dioxo-2,9-dioxa-4,7-diazoundecyl)phenyl)amino)tetradecanamido)ethyl)carbamoyloxy )-5,6-dihydro-[1,3]dioxolo[4,5-g]isoquinolino[3,2-a]isoquinolin-7-ium

[0318]

[0319] To dichloromethane (10 mL) of Example 7-4 (675 mg, 0.88 mmol) at 0 °C was slowly added triethylamine (267 mg, 2.64 mmol). A solution of 4-nitrobenzyl chloroformate (265 mg, 1.32 mmol) in dichloromethane (1 mL) was slowly added dropwise. The reaction was stirred at room temperature for 1 h. The reaction was cooled to 0 °C, and a solution of tert-butylmethyl(2-(methylamino)ethyl)carbamate (249 mg, 1.32 mmol) in dichloromethane (1.32 mL) was slowly added dropwise. The reaction was stirred at room temperature for 1 h. The reaction solution was washed with water (20 mL × 2), dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The residue was separated and purified by silica gel column chromatography (ethyl acetate: petroleum ether = 1:1) to obtain the target compound (240 mg, 28%), which was an orange-yellow oil.

[0320] MS(ESI): m / z = 981.5 [M] + .

[0321] Example 7: 9-(((2-(14-((4-((((2-(4-(((3R,4R)-1-(2-cyanoacetyl)-4-methylpiper idin-3-yl)(methyl)amino)-N-methyl-7H-pyrrolo[2,3-d]pyrimidine-7-carboxamido<oxalamido>)eth yl)(methyl)amino)carbamoyloxy)methyl)phenyl)amino)-14-oxotetradecanamido)ethyl)(methyl)carbam oyloxy)-10-methoxy-5,6-dihydro-[1,3]dioxolo[4,5-g]isoquinolino[3,2-a]isoquinolin-7-ium ion

[0322]

[0323] The solution of Example 7-5 (210 mg, 0.214 mmol) in dichloromethane (2 mL) was placed in a single-necked flask, trifluoroacetic acid (0.4 mL) was added, and the reaction solution was stirred at room temperature for 15 minutes. After monitoring the reaction by LCMS until completion, the reaction solution was concentrated under reduced pressure, dichloromethane (1 mL), N,N-diisopropylethylamine (0.1 mL, 0.64 mmol), and Example 1-11 (102 mg, 0.21 mmol) were added, and the reaction was carried out at room temperature for 15 minutes. The reaction solution was concentrated under reduced pressure, N,N-dimethylformamide (2 mL) was added, and then it was directly injected for separation by Prep-HPLC (gradient elution with acetonitrile / water (containing 0.1% trifluoroacetic acid)) to obtain the target compound (15.2 mg, 5.8%), which was a yellow solid.

[0324] MS(ESI):m / z=1219.7[M] + .

[0325] 1 HNMR(400MHz,CD3OD)δ10.039-9.792(m,1H),8.76-8.73(m,1H),8.53(s,1H),8.15(m,3H),7.64-7.62(m,1H),7.51-7.50(m,1H),7.46-7.44(m,1H),7.29-7.27(m,1H),6.93-6.91(m,1H),6.78-6.66(m,2H),6.082(s,2H),5.00-4.99(m,5H),4.06(s,3H),3.96-3.87(m,4H),3.68(s,3H),3.57-3.55(m,2H),3.48-3.44(m,4H),3.23-3.21(m,3H),3.08(s,2H),2.42-2.40(m,1H),2.33-2.29(t,J=8.0Hz,3H),2.25-2.17(m,3H),1.71-1.54(m,7H),1.28-1.20(m,21H),1.09-0.99(m,5H).

[0326] Example 8: 9-((5-(5-((E)-(4-((((2-(4-(((3R,4R)-1-(2-cyanoacetyl)-4-methylpiper idin-3-yl)(methyl)amino)-N-methyl-7H-pyrrolo[2,3-d]pyrimidine-7-carboxamido<oxalamido>)eth yl)(methyl)amino)carbamoyloxy)methyl)phenyl)diazenyl)-2-hydroxybenzamido)valeryl)oxy)-10- methoxy-5,6-dihydro-[1,3]dioxolo[4,5-g]isoquinolino[3,2-a]isoquinolin-7-ium

[0327]

[0328] Example 8-1: Methyl (E)-5-(2-hydroxy-5-((4-(4,7,10,10-tetramethyl-3,8-dioxo-2,9-di oxa-4,7-diazoundecyl)phenyl)diazenyl)benzamido)valerate

[0329]

[0330] To a solution of intermediate B (3.0 g, 6.17 mmol), 1-hydroxybenzotriazole (1.001 g, 7.41 mmol), and dicyclohexylcarbodiimide (1.783 g, 8.64 mmol) in dichloromethane (20 mL) was stirred at 0 °C for 20 minutes. Methyl 5-aminopentanoate hydrochloride (1.212 g, 7.41 mmol) and N,N-diisopropylethylamine (1752 mg, 13.6 mmol) were added thereto, and the reaction was returned to room temperature and reacted for 2 hours. The reaction was monitored by LCMS to completion, and the reaction solution was filtered, and the filtrate was concentrated under reduced pressure. The residue was separated and purified by normal-phase column chromatography (petroleum ether:ethyl acetate = 1:1) to give the title compound (3.34 g, 91%), which was a red solid.

[0331] MS(ESI): m / z = 622.2 [M+Na] + .

[0332] Example 8-2: (E)-5-(2-hydroxy-5-((4-(4,7,10,10-tetramethyl-3,8-dioxo-2,9-dioxa -4,7-diazoundecyl)phenyl)diazenyl)benzamido)valeric acid

[0333]

[0334] To a solution of Example 8-1 (3300 mg, 5.51 mmol) in methanol (30 mL) and water (15 mL) was added lithium hydroxide monohydrate (1157 mg, 27.5 mmol), and the reaction was heated to reflux at 65 °C for 1 hour. After cooling, the reaction solution was adjusted to pH 5 with dilute hydrochloric acid. Extracted 3 times with ethyl acetate (100 mL), the organic phase was dried over anhydrous sodium sulfate, and concentrated under reduced pressure to give the title compound (3000 mg, 93.1%), which was an orange solid.

[0335] MS(ESI): m / z = 608.2 [M+Na] + .

[0336] Example 8-3: (E)-9-((5-(2-hydroxy-5-((4-(4,7,10,10-tetramethyl-3,8-dioxo-2,9-di oxa-4,7-diazoundecyl)phenyl)diazenyl)benzamido)valeryl)oxy)-10-methoxy-5,6- dihydro-[1,3]dioxolo[4,5-g]isoquinolino[3,2-a]isoquinolin-7-ium

[0337]

[0338] To a solution of Example 8-2 (1500 mg, 2.56 mmol) in dichloromethane (20 mL) placed in a single-necked flask was added dicyclohexylcarbodiimide (792 mg, 3.84 mmol) and intermediate A-8 (908 mg, 2.82 mmol). The reaction solution was stirred at room temperature for 2 hours. The reaction was monitored by LCMS to completion, and the reaction solution was washed 3 times with water (60 mL), the organic phase was dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The residue was separated and purified by normal-phase column chromatography (dichloromethane:methanol = 10:1) to give the title compound (367 mg, 16.1%), which was an orange oil.

[0339] MS(ESI): m / z = 889.4 [M] + .

[0340] Example 8: 9 - ((5 - (5 - ((E)-(4 - ((((2 - (4 - ((((3R,4R)-1-(2 - cyanoacetyl)-4 - methylpiperidin - 3 - yl)(methyl)amino)-N - methyl - 7H - pyrrolo[2,3 - d]pyrimidine - 7 - carbonylamino<oxalylamino>)ethyl)(methyl)carbamoyl)oxo)methyl)phenyl)diazenyl)-2 - hydroxybenzoylamino)valeryl)oxo)-10 - methoxy - 5,6 - dihydro - [1,3]dioxazolo[4,5 - g]isoquinolino[3,2 - a]isoquinolin - 7 - ylium Example 10: 9 - ((3 - (2 - ((E)-(4 - ((((2 - (4 - ((((3R,4R)-1-(2 - cyanoacetyl)-4 - methyl piperidin - 3 - yl)(methyl)amino)-N - methyl - 7H - pyrrolo[2,3 - d]pyrimidine - 7 - carbonylamino<oxalylamino>)ethyl)(methyl)carbamoyl)oxo)methyl)phenyl)diazenyl)phenyl)propanoyl)oxo)-10 - methoxy - 5,6 - di

[0341]

[0342] To the dichloromethane (2.5 mL) of Example 8-3 (300 mg, 0.34 mmol) was added trifluoroacetic acid (0.5 mL). The reaction mixture was stirred at room temperature for 15 minutes, and the solution was concentrated under reduced pressure. Then, N,N-dimethylformamide (2 mL), N,N-diisopropylethylamine (95.8 mg, 0.74 mmol), and Example 1-11 (193 mg, 0.4 mmol) were added. The reaction was carried out at room temperature for 15 minutes. After completion of the reaction detected by LCMS, 1N hydrochloric acid was added to quench the reaction. The reaction solution was directly injected, and the target compound (62.0 mg, 16%) was obtained by separation with Prep-HPLC (gradient elution with acetonitrile / water (containing 0.1% trifluoroacetic acid)) as a yellow solid.

[0343] MS(ESI): m / z = 1127.5 [M] + .

[0344] 1 H NMR (400 MHz, CD3OD) δ 9.63 - 9.60 (m, 1H), 8.70 - 8.62 (m, 1H), 8.39 - 8.35 (m, 1H), 8.11 - 8.02 (m, 3H), 7.91 - 7.89 (m, 1H), 7.68 - 7.66 (m, 2H), 7.59 - 7.56 (m, 2H), 7.46 - 7.40 (m, 2H), 7.01 - 6.85 (m, 3H), 6.09 (s, 2H), 5.18 - 5.16 (m, 2H), 4.03 - 4.02 (m, 3H), 3.92 - 3.91 (m, 3H), 3.81 - 3.73 (m, 3H), 3.65 - 3.59 (m, 2H), 3.56 - 3.52 (m, 4H), 3.46 - 3.39 (m, 2H), 3.24 - 3.18 (m, 5H), 3.17 - 3.16 (m, 3H), 2.94 - 2.91 (m, 4H), 2.36 - 2.28 (m, 1H), 1.99 - 1.94 (m, 2H), 1.91 - 1.86 (m, 2H), 1.68 - 1.60 (m, 5H), 0.94 - 0.92 (m, 2H).

[0345] The following compounds were obtained by using a method similar to that of Example 8 and replacing the corresponding raw materials.

[0346]

[0347] hydro - [1,3]dioxazolo[4,5 - g]isoquinolino[3,2 - a]isoquinolin - 7 - ylium Example 10 - 1: 1 - hydroxy - 3,4 - dihydroquinolin - 2(1H) - one Example 10 - 2: 3 - (2 - nitrophenyl)propanoic acid Example 10 - 3: (E)-3 - (2 - ((4 - (hydroxymethyl)phenyl)diazenyl)phenyl)propanoic acid

[0348]

[0349] Example 10 - 4: (E)-3 - (2 - ((4 - (4,7,10,10 - tetramethyl - 3,8 - dicarbonyl - 2,9 - dioxo - 4,7 -

[0350]

[0351] Hydrogen peroxide (35%, 22 mL) was added dropwise to a methanol solution (200 mL) of 1,2,3,4-tetrahydroquinoline (10.0 g, 75 mmol) and sodium tungstate dihydrate (1.9 g, 3.7 mmol). After the addition was complete, the mixture was stirred at room temperature overnight. The reaction solution was concentrated under reduced pressure, and the residue was dissolved in water (200 mL) and extracted with dichloromethane (100 mL × 2). The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to obtain the crude product. The crude product was slurried with dichloromethane / methanol (1:1, 100 mL) and filtered. The filter cake was dried to obtain the title compound (8.8 g, 72%) as a brownish-yellow solid.

[0352] MS (ESI): m / z = 164.1 [M+H] + .

[0353] 1 1H NMR (CDCl3, 400 MHz) δ 9.03 (br, 1H), 7.33 (d, J = 8.0 Hz, 1H), 7.28 (t, J = 8.0 Hz, 1H), 7.15 (d, J = 8.8 Hz, 1H), 7.05 (t, J = 7.6 Hz, 1H), 2.93 (t, J = 7.6 Hz, 2H), 2.76 (t, J = 8.0 Hz, 2H).

[0354] diazoundecyl)phenyl)diazenyl)phenyl)propanoic acid

[0355]

[0356] Under ice-water bath cooling, sodium periodate (23.1 g, 107 mmol) was added to a solution of Example 10-1 (8.8 g, 53 mmol) in tetrahydrofuran (120 mL) and water (30 mL). Stir at room temperature for 1 hour. The reaction solution was concentrated under reduced pressure to remove the organic solvent, and the residue was diluted with water (100 mL), and the pH value was adjusted to weakly acidic with hydrochloric acid (2M), and extracted with ethyl acetate (450 mL). The combined organic phases were dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The obtained crude product was slurried with ethyl acetate (100 mL) to obtain the title compound (4.5 g, 46%), which was a yellow solid.

[0357] MS(ESI): m / z = 171.2 [M+H] + .

[0358] Example 10 - 5: (E)-10 - methoxy - 9 - ((3 - (2 - ((4 - (4,7,10,10 - tetramethyl - 3,8 - dicarbonyl - 2,

[0359]

[0360] Example 10-2 (4.5 g, 25.1 mmol) and tetraaminobenzyl alcohol (3.1 g, 25.1 mmol) were dissolved in dichloromethane (150 mL), acetic acid (15 mL) was added, and the mixture was stirred at room temperature for 48 hours under nitrogen protection. LCMS monitored that the raw materials reacted completely. The reaction solution was diluted with dichloromethane (100 mL), washed successively with water (100 mL * 2) and saturated brine (100 mL), the organic phase was dried over anhydrous sodium sulfate, concentrated under reduced pressure, and the residue was purified by normal-phase column chromatography (dichloromethane: methanol = 94:6), and slurried with ethyl acetate (30 mL) to obtain the title compound (4.0 g, 56%), which was a red solid.

[0361] MS(ESI): m / z = 285.1 [M+H] + .

[0362] 9 - dioxo - 4,7 - diazoundecyl)phenyl)diazenyl)phenyl)propanoyl)oxo)-5,6 - dihydro - [1,3]dioxo zolo[4,5 - g]isoquinolino[3,2 - a]isoquinolin - 7 - ylium

[0363]

[0364] Example 10-3 (3.7 g, 13.0 mmol) and bis(p-nitrophenyl) carbonate (4.752 g, 15.6 mmol) were mixed in dichloromethane (50 mL), and diisopropylethylamine (3.361 g, 26.0 mmol) was added dropwise under cooling in an ice-water bath. The reaction mixture was stirred at room temperature for 30 minutes, and then tert-butyl methyl(2-(methylamino)ethyl)carbamate (3.184 g, 16.9 mmol) was added dropwise to the reaction mixture at 0 °C. After the addition was complete, the reaction mixture was reacted at room temperature for 1 hour and then heated to 40 °C and stirred for 2 hours. The conversion of the starting materials was monitored by LCMS to be complete. The reaction mixture was diluted with dichloromethane (200 mL), washed with water (100 mL), the organic phase was dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The residue was purified by normal-phase column chromatography (dichloromethane:methanol = 10:1) to give the target compound (1.1 g, 17%), as a yellow solid.

[0365] MS(ESI): m / z = 521.1 [M+Na] + .

[0366] Example 10: 9 - ((3 - (2 - ((E)-(4 - ((((2 - (4 - ((((3R,4R)-1-(2 - cyanoacetyl)-4 - methyl piperidin - 3 - yl)(methyl)amino)-N - methyl - 7H - pyrrolo[2,3 - d]pyrimidine - 7 - carbonylamino<oxalylamino>)ethyl)(methyl)carbamoyl)oxo)methyl)phenyl)diazenyl)phenyl)propanoyl)oxo)-10 - methoxy - 5,6 - di hydro - [1,3]dioxazolo[4,5 - g]isoquinolino[3,2 - a]isoquinolin - 7 - ylium

[0367]

[0368] Example 10-4 (650 mg, 1.3 mmol) and pyridine (412 mg, 5.2 mmol) were mixed in dichloromethane (30 mL), and oxalyl chloride (486 mg, 1 mmol) was added dropwise at room temperature. The reaction mixture was stirred at room temperature for 30 minutes, and the conversion of the starting materials was monitored by LCMS. The reaction mixture was concentrated under reduced pressure, dried with an oil pump, and then dissolved in acetonitrile (5 mL), which was added dropwise to a solution of intermediate A-8 (420 mg, 1.3 mmol) and pyridine (412 mg, 5.2 mmol) in acetonitrile (30 mL). The reaction mixture was stirred at room temperature for 20 minutes, and the conversion of the starting materials was monitored by LCMS to be complete. The reaction mixture was concentrated under reduced pressure and diluted with dichloromethane (100 mL), washed with water (100 mL), dilute hydrochloric acid (1N, 100 mL), the organic phase was dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The residue was purified by normal-phase column chromatography (dichloromethane:methanol = 92:8, containing 0.1% trifluoroacetic acid) to give the target compound (217 mg, 21%), as a yellow oil.

[0369] MS(ESI): m / z = 803.2 [M+H] + .

[0370] Example 11: (E)-9 - ((5 - (5 - ((4 - ((((2 - (4 - (1 - (3 - (cyanomethyl)-1 - (ethylsulfonyl)azetidin - ​ ​ ​

[0371]

[0372] To a solution of Example 10-5 (217 mg, 0.27 mmol) in dichloromethane (2 mL) was added trifluoroacetic acid (0.4 mL), and the mixture was stirred at room temperature for 1 hour. The reaction was monitored by LCMS until completion. The reaction solution was concentrated under reduced pressure, dried with an oil pump, dissolved in N,N-dimethylformamide (1.5 mL) solution, cooled to 0 °C, N,N-diisopropylethylamine (139 mg, 1.1 mmol) was added, and Example 1-11 (258 mg, 0.54 mmol) was added. The mixture was stirred at room temperature for 0.5 hour. After quenching the reaction solution with 1N hydrochloric acid, it was directly injected and separated by Prep-HPLC (gradient elution with acetonitrile / water (containing 0.1% trifluoroacetic acid)) to obtain the title compound (63.0 mg, 22.4%), which was a yellow solid.

[0373] MS(ESI): m / z = 1140.4 [M] + .

[0374] 1 1H NMR (400 MHz, CD3OD) δ 9.60 - 9.47 (m, 1H), 8.72 - 8.67 (m, 1H), 8.19 - 8.07 (m, 3H), 7.87 - 7.74 (m, 2H), 7.63 - 7.39 (m, 6H), 6.99 - 6.51 (m, 4H), 6.09 (s, 2H), 5.21 - 5.05 (m, 1H), 4.97 - 4.86 (m, 3H), 4.00 - 3.71 (m, 8H), 3.66 - 3.37 (m, 8H), 3.24 - 3.16 (m, 9H), 3.05 - 2.81 (m, 6H), 2.40 - 2.26 (m, 1H), 1.90 - 1.77 (m, 1H).

[0375] ​ (pyridin-3-yl)-1H-pyrazol-4-yl)-N-methyl-7H-pyrrolo[2,3-d]pyrimidine-7-carboximidamido<oxalamidino >)ethyl)(methyl)carbamoyl)oxy)methyl)phenyl)diazenyl)-2-hydroxybenzamido)valeryl)oxy)- 10-methoxy-5,6-dihydro-[1,3]dioxolo[4,5-g]isoquinolino[3,2-a]isoquinolin-7-ium

[0376]

[0377] Example 8-3-A: (E)-9-((5-(2-hydroxy-5-((4-(((methyl(2-(methylamino)ethyl)amino carbamoyl)oxy)methyl)phenyl)diazenyl)benzamido)valeryl)oxy)-10-methoxy-5,6-dihydro-[1,3]di oxolo[4,5-g]isoquinolino[3,2-a]isoquinolin-7-ium hydrochloride

[0378]

[0379] At 0 °C, hydrochloric acid ethyl acetate solution (4 N, 70 mL) was added to a solution of Example 8-3 (7.7 g, 7.13 mmol) in ethyl acetate (35 mL), and the reaction was carried out at 25 °C for 1 hour. After the reaction was monitored by LCMS to completion, the reaction mixture was filtered, and the filter cake was washed with ethyl acetate to obtain the title compound (6.8 g, 110%), a red solid.

[0380] MS(ESI): m / z = 789.3 [M] + .

[0381] Example 1-A: 4-nitrophenyl 4-(1-(3-(cyanomethyl)-1-(ethylsulfonyl)azetidin-3-yl)-1H-py razol-4-yl)-7H-pyrrolo[2,3-d]pyrimidine-7-carboxylate

[0382]

[0383] At room temperature, p-nitrophenyl chloroformate (108 mg, 0.54 mmol) was added to a mixture of baricitinib (100 mg, 0.27 mmol), triethylamine (109 mg, 1.08 mmol) and dichloromethane (5 mL). The reaction mixture was stirred at room temperature for 1.5 hours. The reaction was monitored by LCMS until the starting materials were completely converted. The reaction mixture was diluted with dichloromethane (50 mL), washed with water (50 mL) and brine (50 mL) respectively, and the organic phase was dried over anhydrous sodium sulfate and concentrated under reduced pressure to obtain the crude product. The crude product was triturated with dichloromethane (5 mL), filtered, and the filter cake was dried to obtain the title compound (100 mg, 85%), a yellow solid.

[0384] MS(ESI): m / z = 537.0 [M+H] + .

[0385] Example 11: (E)-9-((5-(5-((4-((((2-(4-(1-(3-(cyanomethyl)-1-(ethylsulfonyl)azetid in-3-yl)-1H-pyrazol-4-yl)-N-methyl-7H-pyrrolo[2,3-d]pyrimidine-7-carboximidamido<oxalamidino >)ethyl)(methyl)carbamoyl)oxy)methyl)phenyl)diazenyl)-2-hydroxybenzamido)valeryl)oxy)- 10-methoxy-5,6-dihydro-[1,3]dioxolo[4,5-g]isoquinolino[3,2-a]isoquinolin-7-ium

[0386]

[0387] To a mixture of Example 8-3-A (118 mg, 0.15 mmol), N-methylmorpholine (19 mg, 0.18 mmol) and N,N-dimethylformamide (2 mL), Example 1-A (100 mg, 0.18 mmol) was added. The reaction mixture was stirred at room temperature for 2 hours. The reaction was monitored by LCMS until the starting materials were completely converted. The reaction mixture was quenched with hydrochloric acid (1 N). The reaction mixture was directly injected and separated by Prep-HPLC (gradient elution with acetonitrile / water (containing 0.1% trifluoroacetic acid)) to obtain the title compound (93 mg, 42%), a yellow solid.

[0388] MS(ESI): m / z = 1186.4 [M] + .

[0389] 1 1H NMR (400 MHz, CD3CN) δ 9.37 (s, 1H), 8.98 - 8.55 (m, 4H), 8.18 - 8.02 (m, 4H), 7.66 - 7.33 (m, 6H), 7.04 - 6.88 (m, 3H), 6.12 (s, 2H), 5.29 - 5.09 (m, 1H), 4.77 - 4.55 (m, 5H), 4.28 - 4.22 (m, 3H), 4.02 (s, 3H), 3.77 - 3.46 (m, 9H), 3.12 - 2.91 (m, 14H), 1.34 - 1.31 (m, 5H).

[0390] The following compounds can be obtained by using a method similar to that of Example 11 and replacing the corresponding raw materials. The reaction solvent for preparing Example 1 - A can be dichloromethane, N,N - dimethylformamide, etc., and the base can be triethylamine, 2,6 - dimethylpyridine, etc. The reaction solvent for preparing Example 1 can be N,N - dimethylformamide, N,N - dimethylacetamide, etc.

[0391]

[0392]

[0393]

[0394] Biological Test 1: Ex vivo release experiment of co - drug in the duodenum or colon contents of mice (Lumen content ex vivo assay).

[0395] Male C57 BL / 6 mice (6 - 8 weeks old) were sacrificed by carbon dioxide euthanasia and dissected. The duodenum and colon segments were removed and placed in 1.5 - milliliter centrifuge tubes, and PBS solution was added simultaneously. The intestinal segments were longitudinally incised, shaken to release the intestinal contents, and mixed by inverting. DMSO solutions of the compounds in the examples were respectively prepared, and 20 μL of each of the above DMSO solutions was placed in 1 mL of PBS solution of duodenal or colon contents. The mixture was repeatedly inverted and mixed, and then placed in a water bath at 37 °C. At the time points of 0 h, 1 h, 4 h, 16 h, and 20 h, the above solutions were taken, acetonitrile was added, the solution was vortexed, and centrifuged for 10 minutes. The supernatant was taken, and 10 μL of the internal standard compound (Intermediate D) was added. The amounts (ng) of the compounds in the examples, berberrubine, and tofacitinib were detected by liquid chromatography - mass spectrometry and quantified with a standard curve, and the results are shown in Table 1. The results show that the co - drug compounds of the present invention can be released in the intestine after administration.

[0396] Table 1 Release of berberrubine, tofacitinib, SHR0302, and upadacitinib over time in different intestinal environments in Examples 1, 8, 14, and 16

[0397]

[0398]

[0399] N.D. indicates not detected

[0400] Bioassay 2: Pharmacokinetics experiment of co-administered compounds in mice

[0401] CD-1 mice were orally administered (PO, 15 mg / kg) the test compounds, and blood samples and tissue samples from various segments of the gastrointestinal tract were collected at different time points. The concentrations of the co-administered compounds of the examples and the released tofacitinib and berberrubine in the plasma of mice were determined by LC-MS / MS. The animals were approximately 6 - 8 weeks old at the start of the dosing experiment. Sampling times for blood and tissue samples: 0.5, 1, 2, 4, 8, and 24 hours after dosing. A bioanalytical method for biological samples and a sample detection method were established.

[0402] The results are as Figure 1-5 shown. The results indicate that the co-administered compounds of the examples can release tofacitinib and berberrubine in the mouse intestine, and the co-administered compounds, tofacitinib, and berberrubine are mainly confined to the intestinal tissue, with only extremely low drug exposure in the plasma.

[0403] Bioassay 3: Efficacy experiment of co-administered compounds on an oxazolone-induced mouse colitis model

[0404] C57 BL / 6 mice were used to establish an oxazolone-induced colitis model according to the method of Heller et al. On the first day, the hair on the nape of the mice was shaved (2 cm × 2 cm), and 150 μl of 3% oxazolone solution (dissolved in a 4:1 mixed solution of acetone and olive oil) was applied for sensitization. On the 6th day after sensitization, the mice were randomly grouped. Subsequently, the corresponding compounds of the examples were administered by gavage (120 mg / kg), and the blank control group and the model group were given the solvent, with a gavage volume of 10 ml / kg body weight. Subsequently, on the next day, 50 μl of 1.2% oxazolone solution was administered by enema, and pure water was injected into the blank control group. Subsequent gavage administration continued for 4 days, and the Disease Activity Index (DAI) was recorded every day. The results are as Figure 6 shown. Compared with the model group, the co-administered compound group of the examples can significantly improve the disease activity index.

[0405] All documents mentioned in this invention are cited herein as references, as if each document was individually cited as a reference. In addition, it should be understood that after reading the above teachings of this invention, those skilled in the art can make various changes or modifications to this invention, and these equivalent forms also fall within the scope defined by the appended claims of this application.

Claims

1. A co-drug compound represented by the following formula I, which is formed by coupling a first drug molecule, a second drug molecule and a linker precursor: D1-linker-D2; I wherein, D1 is a first drug group; and has the following structure: The second drug group is selected from the following group: And the linker has a structure selected from group (a) or (c). In each formula, J1 is connected to the first drug group, and J2 is connected to the second drug group; (a)-L a -L-, wherein the L a has the structure shown below: and the L has the structure shown below, where * is the connection site of L and L a : (c) wherein, the B ring and the C ring are C6 aryl groups; The described J1 is -(Y) z -, and the described Y is selected from the group consisting of: -C(O)-, -C(O)NH-; the Y may be substituted by one or more Rs, provided that the respective Ys together form a chemically stable structure; z is 1; t = 0, J2- is R4 is H or C 1-6 alkyl; Each L5 is independently selected from the following group: C1-C8 alkylene, -NHC(O)-; And the L5 is optionally substituted by one or more R, provided that each L5 forms a chemically stable structure; And the R is selected from the following group: H, -OH, C1-C4 alkyl; s are independently selected from 0, 1, 2, 3 or 4; And when the first drug group has the structure shown by the following formula: the linker has the structure of (a); When the first drug group has the structure shown by the following formula: the linker has the structure of group (c).

2. The co-drug compound according to claim 1, wherein The second drug group is selected from the following group:

3. The co-drug compound according to claim 1, wherein The first drug group is: The linker is selected from group (C): (C) group:

4. The compound of formula I according to claim 1, characterized in that, The compound is selected from the following group:

5. A pharmaceutical composition comprising a therapeutically effective amount of the compound or racemate or a pharmaceutically acceptable salt thereof according to claim 1, and a pharmaceutically acceptable excipient.

6. The pharmaceutical composition according to claim 5, wherein The pharmaceutical composition is an enteric-coated preparation.

7. The pharmaceutical composition according to claim 5, characterized in that, The pharmaceutical composition is used for treating diseases selected from the following group: ulcerative colitis, Crohn's disease, collagenous colitis, lymphocytic colitis, acute / chronic gastritis, acute / chronic appendicitis.

8. The pharmaceutical composition according to claim 5, characterized in that, The pharmaceutical composition is used for treating colitis associated with immune checkpoint inhibitor therapy.

9. The pharmaceutical composition according to claim 5, characterized in that, The pharmaceutical composition is used for treating autoimmune enteropathy.

10. Use of the compound according to claim 1 or a pharmaceutically acceptable salt thereof or the pharmaceutical composition according to claim 5, characterized in that, For preparing a drug for preventing and treating gastrointestinal functional diseases, the gastrointestinal functional diseases are selected from the following group: ulcerative colitis, Crohn's disease.

11. Use of the compound according to claim 1 or a pharmaceutically acceptable salt thereof or the pharmaceutical composition according to claim 5, characterized in that, For preparing a drug for preventing and treating gastrointestinal functional diseases, the gastrointestinal functional disease is autoimmune enteropathy.

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