WO2020043173A1 - 作为受体相互作用蛋白1(rip1)激酶抑制剂的杂环化合物 - Google Patents

作为受体相互作用蛋白1(rip1)激酶抑制剂的杂环化合物 Download PDF

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WO2020043173A1
WO2020043173A1 PCT/CN2019/103367 CN2019103367W WO2020043173A1 WO 2020043173 A1 WO2020043173 A1 WO 2020043173A1 CN 2019103367 W CN2019103367 W CN 2019103367W WO 2020043173 A1 WO2020043173 A1 WO 2020043173A1
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王能辉
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宁波文达医药科技有限公司
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Definitions

  • the invention belongs to the field of medicinal chemistry, and particularly relates to a heterocyclic compound as a receptor interaction protein 1 (RIP1) kinase inhibitor and a pharmaceutically acceptable salt thereof.
  • RIP1 receptor interaction protein 1
  • Receptor-interacting protein 1 (RIP1) kinases are a class of serine / threonine protein kinases that can regulate the activity of nuclear factor kB, are the intersections that determine cell survival and death, and are key in the programmed necroptosis signaling pathway. Regulatory factors. Programmed cell death plays an important role in many aspects such as individual development, body homeostasis, and pathological processes. Cell death mainly includes apoptosis, necrosis and autophagy. Among them, necrosis has long been considered a passive and unregulated process. However, recent years have shown that cell necrosis is also precisely regulated.
  • cell necrosis Unlike apoptosis, cell necrosis will activate the body's immune response, and local cell necrosis of the body will cause a global physiological and pathological response. Therefore, it is involved in many pathological processes, such as immune activation of viral infection, ischemic necrotic injury (ischemic injury), and the occurrence and development of neurodegenerative diseases. Compared with the more mature research on apoptosis, cell necrosis is a new field.
  • Programmed cell necrosis is a form of cell death triggered by the activation of kinases. Activation of death receptors (such as TNFR1) can eventually induce programmed cell necrosis, and the signaling that initiates cell necrosis mainly depends on the regulation of the kinases RIP1 and RIP3. After cell necrosis occurred, RIP1 combined with RIP3 and activated the kinase activity of RIP3, and then RIP3 autophosphorylated, allowing it to specifically bind to the substrate MLKL, which was then phosphorylated by RIP3. At this time, RIP1 / RIP3 / MLKL forms an active cell necrosis complex, which transmits the death signal downstream, so that programmed cell necrosis can finally occur. Programmed cell necrosis cells release their contents to the surroundings. These contents serve as DAMPs (damage-associated molecular patterns), which can stimulate the surrounding cells to undergo an inflammatory response and activate the body's immune response.
  • DAMPs damage-associated molecular patterns
  • Programmed cell necrosis has important pathophysiological effects, such as myocardial infarction, pancreatitis, ischemia-reperfusion injury, sepsis, stroke, coronary heart disease, inflammatory bowel disease, retinitis, alcoholic fatty liver, non-alcoholic fat Liver, Multiple Sclerosis, Dermatitis, Psoriasis, Chronic Kidney Disease, Acute Kidney Disease, Autoimmune Hepatitis, Hepatitis B, Hepatitis C, Acute Hepatic Insufficiency, Neurodegenerative Disease, Frozen Human Disease, Parkinson, Dementia , Osteoporosis, arthritis, bacterial infections, cancer, atherosclerosis, heart failure, chronic obstructive pulmonary disease, diabetes, blood glucose regulation, protection during tube transplantation, anti-aging, obesity, and many others and RIP1 Kinase-related diseases.
  • pathophysiological effects such as myocardial infarction, pancreatitis, ischemia-reperfusion injury, sep
  • Necrostatin 1 (Nec-1) was discovered, many different structures of RIP1 kinase inhibitors were discovered.
  • the object of the present invention is to provide an effective, safe and / or highly selective small molecule inhibitor of RIP1 kinase. Therefore, the present invention also provides an effective method for preventing and / or treating diseases associated with RIP1 kinase.
  • R 1 represents 1-3 substituents selected from the group consisting of H, halogen, -OH, substituted or unsubstituted C1-C6 alkyl, substituted or unsubstituted C2-C6 alkenyl, substituted or unsubstituted C2-C6 alkynyl, substituted or unsubstituted C3-C10 cycloalkyl, substituted or unsubstituted C3-C10 heterocycloalkyl, substituted or unsubstituted C6-C20 aryl, substituted or unsubstituted C3- C10 heteroaryl, C1-C4 carboxyl, -CN, NO 2 ;
  • R 2 is a substituted or unsubstituted C6-C20 aryl group, a substituted or unsubstituted C3-C10 heteroaryl group or a substituted or unsubstituted 5-8 membered heterocyclic ring;
  • the substitution means that one or more hydrogen atoms are each independently substituted by a substituent selected from the group consisting of H, halogen, -CN, NO 2 , -OH, -NRaRb, C1-C4 alkyl, C1 -C4 haloalkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C6 cycloalkyl, phenyl, benzyl, C6-C10 aryl, C3-C10 heteroaryl,
  • Ra and Rb are each independently H, C1-C3 alkyl, C3-C6 cycloalkyl.
  • R 1 is selected from the group consisting of H, halogen, C1-C6 alkyl, and halogenated C1-C6 alkyl.
  • R 1 is selected from the group consisting of H, halogen, and C1-C6 alkyl.
  • R 1 is selected from the group consisting of H, F, Cl, methyl, ethyl, n-propyl, and isopropyl.
  • R 1 is H or F.
  • R 1 is H.
  • R 2 is a substituted or unsubstituted phenyl group or a 5- to 6-membered heteroaryl group.
  • the 5-8 membered heterocyclic ring contains 1-3 heteroatoms selected from N, O, or S.
  • the 5-8 membered heterocyclic ring is saturated, unsaturated, or partially unsaturated.
  • R 1 represents 2 substituents.
  • R 1 represents R 8 and R 9 ; and R 8 and R 9 are each independently selected from the group consisting of H, halogen, -OH, substituted or unsubstituted C1-C6 alkyl, substituted or Unsubstituted C2-C6 alkenyl, substituted or unsubstituted C2-C6 alkynyl, substituted or unsubstituted C3-C10 cycloalkyl, substituted or unsubstituted C3-C10 heterocycloalkyl, substituted or unsubstituted C6-C20 aryl, substituted or unsubstituted C3-C10 heteroaryl, C1-C4 carboxyl, -CN, NO 2 .
  • R 8 and R 9 are as described above.
  • the compound is represented by Formula Ia
  • R 2 , R 8 and R 9 are as described above.
  • R 8 and R 9 are each independently selected from the group consisting of H, halogen, C1-C6 alkyl, and halogenated C1-C6 alkyl.
  • R 8 and R 9 are each independently selected from the group consisting of H and F.
  • R 8 and R 9 are each independently located at the 3 or 5 position of the benzene ring.
  • the compound is represented by Formula Ib
  • R 2 , R 8 and R 9 are as defined above.
  • the compound is represented by Formula II
  • M, Q, V, Y, and Z are each independently selected from the group: C, N, S, or O;
  • R 3 , R 4 , R 5 , R 6 and R 7 are each independently selected from the group consisting of H, halogen, -CN, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C10 Cycloalkyl, C3-C10 heterocycloalkyl, C3-C10 aryl, C3-C10 heteroaryl, -CN, NO 2 ;
  • any one or more (such as 1 to 3) of M, Q, V, Y, and Z are each independently N, S, or O, and the rest are C.
  • M, Q, V, Y, and Z are each independently C or N.
  • V and Z are N, and M, Q and Y are C.
  • any one or more (such as 1 to 3) of R 3 , R 4 , R 5 , R 6 and R 7 are each independently selected from the group consisting of: halogen, -CN , C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C10 cycloalkyl, C3-C10 heterocycloalkyl, C3-C10 aryl, C3-C10 heteroaryl, -CN, NO 2 ; and the rest is H.
  • R 6 is selected from the group consisting of: halogen, -CN, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C10 cycloalkyl, C3- C10 heterocycloalkyl, C3-C10 aryl, C3-C10 heteroaryl group, -CN, NO 2; and R 3, R 4, R 5 and R 7 is H.
  • R 6 is -CN.
  • R c is each independently selected from: H, halogen, -CN, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C10 cycloalkyl, C3-C10 heterocycloalkyl ( Preferably, C3-C5 cycloalkyl), C3-C10 aryl, C3-C10 heteroaryl, NO 2 (preferably, R c are each independently selected from: H, halogen, -CN, C1-C6 Alkyl, C3-C10 cycloalkyl (preferably C3-C5 cycloalkyl), NO 2 ; more preferably, selected from the group consisting of -CN, C1-C6 alkyl).
  • the compound is represented by Formula III
  • Q, V, Y, and Z are each independently selected from the group consisting of: C, N, S, or O;
  • R 3 , R 4 , R 5 and R 6 are each independently selected from the group consisting of: none, H, halogen, -CN, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C10 ring Alkyl, C3-C10 heterocycloalkyl, C3-C10 aryl, C3-C10 heteroaryl, -CN, NO 2 ;
  • V is C; Y and Z are N; and M is oxygen.
  • any one or more (such as 1 or 2) of R 3 , R 4 , R 5 and R 6 are each independently selected from the group consisting of: halogen, -CN, C1- C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C10 cycloalkyl, C3-C10 heterocycloalkyl, C3-C10 aryl, C3-C10 heteroaryl, -CN, NO 2 ; And the rest are H or None.
  • R 3 , R 4 and R 6 are none; and R 5 is selected from the group consisting of C1-C6 alkyl, C3-C10 heterocycloalkyl, and -CN.
  • R c is each independently selected from: H, halogen, -CN, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C10 cycloalkyl, C3-C10 heterocycloalkyl ( Preferably, C3-C5 cycloalkyl), C3-C10 aryl, C3-C10 heteroaryl, NO 2 (preferably, R c are each independently selected from: H, halogen, -CN, C1-C6 Alkyl, C3-C10 cycloalkyl (preferably C3-C5 cycloalkyl), NO 2 ; more preferably, selected from the group consisting of -CN, C1-C6 alkyl).
  • R 2 is selected from the following group:
  • the compound is selected from the group consisting of:
  • a pharmaceutical composition in a second aspect of the present invention, includes:
  • the heterocyclic compound according to the first aspect or a pharmaceutically acceptable salt thereof, or a pharmaceutically acceptable solvate thereof, which are used by (i) for Preparation of protein kinase inhibitors, (ii) preparation of drugs for treating protein kinase-related diseases, (iii) preparation of drugs for treating TNF-related diseases, and / or (iv) preparation of treatment for programmed cell necrosis-related diseases Drug.
  • the protein kinase is a serine / threonine protein kinase.
  • the protein kinase is a receptor-interacting protein 1 (RIP1) kinase.
  • RIP1 receptor-interacting protein 1
  • the protein kinase-related diseases include: myocardial infarction, pancreatitis, ischemia-reperfusion injury, sepsis, stroke, coronary heart disease, inflammatory bowel disease, retinitis, alcoholic fatty liver, Non-alcoholic fatty liver, multiple sclerosis, dermatitis, psoriasis, chronic kidney disease, acute kidney disease, autoimmune hepatitis, hepatitis B, hepatitis C, acute liver insufficiency, neurodegenerative disease, gradually frozen human disease, par Kingson, Alzheimer's, osteoporosis, arthritis, bacterial infections, cancer, atherosclerosis, heart failure, chronic obstructive pulmonary disease, diabetes, blood glucose regulation, protection during tube transplantation, anti-aging, and / or obesity disease.
  • the preparation method is method one; and method one includes the steps of reacting a compound of formula 8 with R 2 -X to form a compound of formula I;
  • the preparation method is method two; and method two includes the step of reacting a compound of formula 9 with a compound of formula 5 to form a compound of formula I;
  • R 1 and R 2 are as described above.
  • the first method includes steps:
  • R 1 and R 2 are the same as described above;
  • R 10 is a protecting group.
  • X is Cl or Br.
  • R 10 is -BOC (tert-butoxycarbonyl).
  • X is Cl
  • compound 9 is prepared by the following steps:
  • the method for preparing a compound of formula 5 includes steps:
  • a method for non-therapeutic inhibition of a protein kinase in vitro wherein the method includes the step of removing the heterocyclic compound according to the first aspect, or a pharmaceutically acceptable salt thereof, Or a pharmaceutically acceptable solvate thereof is contacted with the protein kinase to inhibit the activity of the protein kinase.
  • the protein kinase is a receptor-interacting protein 1 kinase.
  • a method for inhibiting therapeutic or non-therapeutic inhibition of protein kinases in vivo includes the steps:
  • An effective amount of a heterocyclic compound according to the first aspect, or a pharmaceutically acceptable salt thereof, or a pharmaceutically acceptable solvate thereof, or a pharmaceutical composition according to the second aspect, is administered to a subject. , Thereby inhibiting the activity of the protein kinase.
  • the subject is an animal; preferably a mammal; more preferably, a human.
  • a seventh aspect of the present invention there is provided a method for treating a protein kinase-related disease, a TNF-related disease, and / or a programmed cell necrosis-related disease, the method comprising the steps:
  • An effective amount of a heterocyclic compound according to the first aspect, or a pharmaceutically acceptable salt thereof, or a pharmaceutically acceptable solvate thereof, or a pharmaceutical composition according to the second aspect is administered to a target.
  • the target is an animal; preferably a mammal; more preferably, a human.
  • a preparation method comprising the steps:
  • R 1 is defined as in the first aspect.
  • step (a) the inert solvent is selected from methylene chloride.
  • the reaction temperature of the reaction in step (a) is 0 to 100 ° C; preferably, 0 to 50 ° C; more preferably, 10 to 30 ° C.
  • the reaction time of the reaction in step (a) is 0.1 to 20 hours; preferably, 0.5 to 10 hours; more preferably, 1 to 5 hours; most preferably. 1 to 3 hours.
  • the method further includes the steps:
  • R 1 and R 2 are the same as those in the first aspect.
  • the method further includes the steps:
  • R 1 and R 2 are as defined in the first aspect; R 10 is a protecting group; X is Cl or Br.
  • Figure 1 shows that Compound 1 (NHWD-1062) in one embodiment of the present invention (ie, Example 1) is effective in treating TNF-induced systemic inflammatory response syndrome.
  • alkyl refers to a saturated linear or branched alkyl group, such as methyl, ethyl, isopropyl.
  • C1-C6 alkyl refers to an alkyl group containing 1 to 6 carbon atoms.
  • alkynyl refers to a straight or branched chain hydrocarbon moiety containing at least one triple bond, such as -C ⁇ C-CH3.
  • cycloalkyl refers to a saturated or unsaturated cyclic hydrocarbyl moiety, such as cyclohexyl.
  • heterocycloalkyl refers to a saturated cyclic moiety containing at least one ring heteroatom (eg, N, O, or S), such as 4-tetrahydropyranyl.
  • aryl refers to a hydrocarbyl moiety containing one or more aromatic rings.
  • aryl moieties include, but are not limited to, phenyl (Ph), naphthyl, fluorenyl, anthracenyl, and phenanthryl.
  • heteroaryl refers to a monocyclic, bicyclic, or fused ring aromatic group having a specific number of ring-forming carbon atoms (for example, C 4-10 has 4-10 Ring carbon atoms), and includes at least one heteroatom selected from N, O or S which is the same or different.
  • the atoms in each ring can be arbitrarily substituted.
  • the heteroaryl group may be a 5- to 15-membered aromatic ring group having 1-5 heteroatoms each independently selected from N, O or S. Examples of heteroaryl groups include, but are not limited to, pyridine, pyrimidine, pyrrole, indazole, indole, furan, benzofuran, thiophene, or similar groups.
  • substitution refers to substitution with one or more substituents.
  • substituted refers to the replacement of one or more hydrogen atoms on a particular group with a particular substituent.
  • the specific substituent is the substituent described correspondingly in the foregoing, or the substituent appearing in each embodiment.
  • an optionally substituted group may have a substituent selected from a specific group at any substitutable position of the group, and the substituents may be the same or different at each position.
  • a cyclic substituent, such as a heterocycloalkyl group may be connected to another ring, such as a cycloalkyl group, to form a spirobicyclic ring system, for example, two rings having a common carbon atom.
  • a pharmaceutically acceptable salt of a compound of the invention refers to a salt suitable for contact with the tissue of a subject (eg, a human) without causing unpleasant side effects.
  • a pharmaceutically acceptable salt of a compound of the invention includes a salt of a compound of the invention (e.g., potassium, sodium, magnesium, calcium) having an acidic group or is basic A salt of a compound of the invention (eg, sulfate, hydrochloride, phosphate, nitrate, carbonate).
  • compound of the present invention or “active ingredient of the present invention” are used interchangeably and refer to a heterocyclic compound represented by formula I in the first aspect of the present invention, or a pharmaceutically acceptable salt thereof, or a solvent Hydrate, hydrate, racemate, stereoisomer, or prodrug.
  • the term "pharmaceutically acceptable salt” refers to a salt formed by a compound of the present invention with pharmaceutically acceptable inorganic and organic acids, wherein preferred inorganic acids include (but are not limited to): hydrochloric acid, hydrogen Bromic acid, phosphoric acid, nitric acid, sulfuric acid; preferred organic acids include (but are not limited to): formic acid, acetic acid, propionic acid, succinic acid, naphthalenedisulfonic acid (1,5), asiatic acid, oxalic acid, tartaric acid, lactic acid , Salicylic acid, benzoic acid, valeric acid, diethylacetic acid, malonic acid, succinic acid, fumaric acid, pimelic acid, adipic acid, maleic acid, malic acid, sulfamic acid, phenylpropionic acid, Gluconic acid, ascorbic acid, niacin, isonicotinic acid, methanesulfonic acid,
  • the term "pharmaceutically acceptable solvate” refers to a compound of the invention that forms a solvate with a pharmaceutically acceptable solvent, wherein the pharmaceutically acceptable solvent includes (but is not limited to): water , Ethanol, methanol, isopropanol, tetrahydrofuran, dichloromethane.
  • the pharmaceutically acceptable solvent includes (but is not limited to): water , Ethanol, methanol, isopropanol, tetrahydrofuran, dichloromethane.
  • Certain compounds of the invention can exist in unsolvated as well as solvated forms, including hydrated forms. Solvated forms are generally equivalent to unsolvated forms and should be included within the scope of the invention.
  • Certain compounds of the invention may exist in polymorphic or amorphous forms. Generally, all physical forms are equivalent for the applications considered by the present invention and should be included within the scope of the present invention.
  • a compound in a prodrug form in addition to a salt form, is provided.
  • Prodrugs of the compounds described herein are those compounds that are susceptible to chemical changes under physiological conditions to provide the compounds provided herein.
  • prodrugs can be converted to compounds provided herein in an ex vivo environment by chemical or biochemical methods.
  • a prodrug can be slowly converted to a compound provided herein when it is placed in a transdermal patch depot with a suitable enzyme or chemical agent.
  • the prodrug is a compound of formula I in the form of an ester.
  • Certain compounds of the invention possess asymmetric carbon atoms (optical centers) or double bonds; racemates, diastereomers, geometric isomers, regioisomers, and individual isomers (e.g., isolated enantiomers Are all included in the scope of the present invention.
  • the compounds provided herein have a defined stereochemistry (represented as R or S, or have a dashed or wedge-shaped bond)
  • those skilled in the art will understand that those compounds are substantially free of other isomers (e.g., at least 80% 90%, 95%, 98%, 99% and up to 100% free of other isomers).
  • the compounds of the invention may also contain unnatural proportions of atomic isotopes at one or more isotopic atoms constituting such compounds.
  • the unnatural proportion of an isotope can be defined as the amount of the atom in question to 100% of that atom.
  • compounds can incorporate radioactive isotopes such as tritium ( 3 H), iodine-125 ( 125 I) or carbon-14 ( 14 C), or non-radioactive isotopes such as deuterium ( 2 H) or carbon-13 ( 13 C ).
  • radioactive isotopes such as tritium ( 3 H), iodine-125 ( 125 I) or carbon-14 ( 14 C), or non-radioactive isotopes such as deuterium ( 2 H) or carbon-13 ( 13 C ).
  • isotope variants may provide additional uses.
  • isotopic variants of the compounds of the invention may have additional uses, including but not limited to as diagnostic and / or imaging agents, or as cytotoxic / radiotoxic therapeutic agents.
  • isotopic variants of the compounds of the invention may have altered pharmacokinetic and pharmacodynamic characteristics, thereby helping to increase safety, tolerability, or efficacy during treatment. All isotopic variations of the compounds of the invention, whether or not they are radioactive, should be included within the scope of the invention.
  • the compounds of formula I according to the present invention can be prepared by various methods well known to those skilled in the art of organic synthetic chemistry.
  • the compounds described in the present invention can be synthesized using the methods described below, together with known synthetic methods in the field of organic chemistry, or variations thereon as understood by those skilled in the art.
  • the method for preparing the compound of the formula I according to the present invention can use the following general methods and procedures from readily available starting materials to prepare the compound of the present invention. It will be understood that when typical or preferred process operating conditions are given (ie, reaction temperature, time, moles of reactants, solvents, pressure, etc.), other process operating conditions may also be used, unless stated otherwise. Optimal reaction conditions may vary with the particular reactants or solvents used, but these conditions can be determined by one skilled in the art through routine optimal procedures.
  • the methods of the compounds of formula I described herein according to the invention can be monitored according to any suitable method known in the art.
  • nuclear magnetic resonance, mass spectrometry, HPLC, and thin-layer chromatography are used to monitor product formation.
  • the preparation of compounds can involve the protection and deprotection of multiple chemical groups. The need for protection and deprotection, as well as the selection of an appropriate protecting group, can be easily determined by those skilled in the art.
  • R 1 , R 2 , R 10 and X are defined as described above.
  • the compounds of the present invention can be prepared using the reaction routes and processes described above, but are not limited to reagents and solvents in the reaction conditions.
  • the present invention also provides a preparation method with high yield and high ee value; the method includes the steps:
  • R 1 is as defined above.
  • the present invention also provides a pharmaceutical composition having excellent and highly selective inhibition of protein kinases, the composition comprising: (i) a compound of formula I as an active ingredient, or a pharmaceutically acceptable salt thereof, or A pharmaceutically acceptable solvate thereof, and (ii) one or more pharmaceutically acceptable carriers.
  • the compound itself or a pharmaceutically acceptable salt thereof or a pharmaceutically acceptable solvate thereof, and a mixture of pharmaceutically acceptable excipients, diluents and the like may be in the form of tablets, capsules, granules, powders or syrups Orally, or parenterally in the form of injections.
  • the pharmaceutical composition preferably contains the compound of formula I or a pharmaceutically acceptable salt thereof or a pharmaceutically acceptable solvate thereof as an active ingredient in a weight ratio of 0.01% to 99%, and more preferably a weight ratio of 0.1. % -90% active ingredient.
  • compositions can be prepared by conventional pharmaceutical methods.
  • useful pharmaceutical adjuvants include excipients (e.g. sugar derivatives such as lactose, sucrose, glucose, mannitol and sorbitol; starch derivatives such as corn starch, potato starch, dextrin and carboxymethyl starch; Cellulose derivatives such as crystalline cellulose, hydroxypropyl cellulose, carboxymethyl cellulose, calcium carboxymethyl cellulose, sodium carboxymethyl cellulose; acacia gum; dextran; silicate derivatives such as magnesium metasilicate Aluminum; phosphate derivatives such as calcium phosphate; carbonate derivatives such as calcium carbonate; sulfate derivatives such as calcium sulfate, etc.), binders (such as gelatin, polyvinylpyrrolidone, and polyethylene glycol), disintegrants ( For example, cellulose derivatives such as sodium carboxymethyl cellulose, polyvinylpyrrolidone), lubricants (such as talc, calcium stea
  • the amount of the compound of the present invention, a pharmaceutically acceptable salt thereof, or a pharmaceutically acceptable solvate thereof, or a pharmaceutical composition thereof varies depending on the age, sex, race, condition, etc. of the patient.
  • the compound of formula I of the present invention has excellent inhibitory activity on protein kinases (receptor interacting protein 1, RIP1), the compound of the present invention and its various crystal forms, pharmaceutically acceptable inorganic or organic salts, hydrates or The solvate and the pharmaceutical composition containing the compound of the present invention as a main active ingredient can be used to treat, prevent, and alleviate diseases related to the activity or expression level of protein kinases (receptor interaction protein 1, RIP1).
  • the compounds of the present invention can be used to treat the following diseases (but not limited to): myocardial infarction, pancreatitis, ischemia-reperfusion injury, sepsis, stroke, coronary heart disease, inflammatory bowel disease, retinitis, alcohol Fatty liver, non-alcoholic fatty liver, multiple sclerosis, dermatitis, psoriasis, chronic kidney disease, acute kidney disease, autoimmune hepatitis, hepatitis B, hepatitis C, acute liver insufficiency, neurodegenerative disease, gradual freezing Human disease, Parkinson's disease, Alzheimer's disease, osteoporosis, arthritis, bacterial infections, cancer, atherosclerosis, heart failure, chronic obstructive pulmonary disease, diabetes, blood glucose regulation, protection during tube transplantation, anti-aging, Obesity, and other diseases associated with excessive RIP1 kinase activity.
  • diseases but not limited to): myocardial infarction, pancreatitis, ischemia-reper
  • the pharmaceutical composition of the present invention comprises the compound of the present invention or a pharmacologically acceptable salt thereof and a pharmacologically acceptable excipient or carrier within a safe and effective amount.
  • safe and effective amount is meant that the amount of the compound is sufficient to significantly improve the condition without causing serious side effects.
  • a pharmaceutical composition contains from 1 to 2000 mg of a compound / agent of the invention, and more preferably from 5 to 200 mg of a compound / agent of the invention.
  • the "one dose" is a capsule or tablet.
  • “Pharmaceutically acceptable carrier” refers to one or more compatible solid or liquid fillers or gel substances that are suitable for human use and must have sufficient purity and low enough toxicity. "Compatibility” herein means that each component of the composition can blend with the compound of the present invention and each other without significantly reducing the pharmacological effect of the compound.
  • pharmaceutically acceptable carriers are cellulose and its derivatives (such as sodium carboxymethyl cellulose, sodium ethyl cellulose, cellulose acetate, etc.), gelatin, talc, and solid lubricants (such as stearic acid).
  • Magnesium stearate calcium sulfate, vegetable oils (such as soybean oil, sesame oil, peanut oil, olive oil, etc.), polyols (such as propylene glycol, glycerin, mannitol, sorbitol, etc.), emulsifiers (such as Tween ), Wetting agents (such as sodium lauryl sulfate), colorants, flavoring agents, stabilizers, antioxidants, preservatives, pyrogen-free water, etc.
  • vegetable oils such as soybean oil, sesame oil, peanut oil, olive oil, etc.
  • polyols such as propylene glycol, glycerin, mannitol, sorbitol, etc.
  • emulsifiers such as Tween
  • Wetting agents such as sodium lauryl sulfate
  • the administration method of the compound or pharmaceutical composition of the present invention is not particularly limited, and representative administration methods include (but are not limited to): oral, intratumoral, rectal, parenteral (intravenous, intramuscular, or subcutaneous), and topical administration .
  • Solid dosage forms for oral administration include capsules, tablets, pills, powders and granules.
  • the active compound is mixed with at least one conventional inert excipient (or carrier), such as sodium citrate or dicalcium phosphate, or with the following ingredients: (a) a filler or compatibilizer, for example, Starch, lactose, sucrose, glucose, mannitol and silicic acid; (b) binders, such as hydroxymethyl cellulose, alginates, gelatin, polyvinylpyrrolidone, sucrose and gum arabic; (c) humectants, For example, glycerol; (d) disintegrating agents, such as agar, calcium carbonate, potato starch or tapioca starch, alginic acid, certain complex silicates, and sodium carbonate; (e) slow solvents, such as paraffin; (f) Absorption accelerators, such as quaternary amine compounds; (g) wetting agents, such as cetyl alcohol and g
  • Solid dosage forms such as tablets, dragees, capsules, pills and granules can be prepared using coatings and shell materials, such as casings and other materials known in the art. They may contain opaque agents and the release of the active compound or compounds in such a composition may be released in a certain part of the digestive tract in a delayed manner. Examples of embedding components that can be used are polymeric substances and waxes. If necessary, the active compound may also be microencapsulated with one or more of the aforementioned excipients.
  • Liquid dosage forms for oral administration include pharmaceutically acceptable emulsions, solutions, suspensions, syrups or elixirs.
  • liquid dosage forms may include inert diluents conventionally used in the art, such as water or other solvents, solubilizers, and emulsifiers.
  • inert diluents conventionally used in the art, such as water or other solvents, solubilizers, and emulsifiers.
  • ethanol isopropanol
  • ethyl carbonate ethyl acetate
  • propylene glycol 1
  • 3-butanediol dimethylformamide
  • oils especially cottonseed oil, peanut oil, corn germ oil, olive oil, castor oil, and sesame oil, or mixtures thereof.
  • composition may also contain adjuvants such as wetting agents, emulsifying and suspending agents, sweeteners, flavoring agents, and perfumes.
  • adjuvants such as wetting agents, emulsifying and suspending agents, sweeteners, flavoring agents, and perfumes.
  • the suspension may contain suspending agents, for example, ethoxylated isostearyl alcohol, polyoxyethylene sorbitol and sorbitan esters, microcrystalline cellulose, aluminum methoxide and agar, or mixtures of these, and the like.
  • suspending agents for example, ethoxylated isostearyl alcohol, polyoxyethylene sorbitol and sorbitan esters, microcrystalline cellulose, aluminum methoxide and agar, or mixtures of these, and the like.
  • compositions for parenteral injection may include physiologically acceptable sterile aqueous or anhydrous solutions, dispersions, suspensions or emulsions, and sterile powders for reconstitution into sterile injectable solutions or dispersions.
  • Suitable aqueous and non-aqueous vehicles, diluents, solvents or excipients include water, ethanol, polyols and suitable mixtures thereof.
  • Dosage forms of the compounds of the invention for topical administration include ointments, powders, patches, sprays and inhalants.
  • the active ingredient is mixed under sterile conditions with a physiologically acceptable carrier and any preservatives, buffers, or propellants that may be required if necessary.
  • the compounds of the invention may be administered alone or in combination with other pharmaceutically acceptable compounds.
  • a safe and effective amount of a compound of the present invention is applied to a mammal (such as a human) in need of treatment, wherein the dose when administered is a pharmaceutically considered effective dose.
  • a mammal such as a human
  • the dosage is usually 1 to 2000 mg, preferably 5 to 500 mg.
  • the specific dosage should also consider factors such as the route of administration, the patient's health, etc., which are all within the skill of a skilled physician.
  • the compounds of the present invention have excellent in vitro and in vivo inhibitory activity on RIP1.
  • the method of the present invention for preparing a compound of Formula 5 from a compound of Formula 4 has a high yield and a high ee value.
  • N-tert-butoxycarbonylhydroxylamine 600 mg, 4.50 mmol was added to DMF (10 ml), the ice bath was cooled to 0 ° C, and then NaH (180 mg, 4.5 mmol, 60%) was added, and kept at 0 after the addition. The mixture was reacted for half an hour at °C, and then the mixed solution was slowly added dropwise to a DMF (5 ml) solution containing compound 1a (700 mg, 4 mmol). The obtained reaction solution was reacted at 0 ° C for half an hour, and then at room temperature for 24 hours Saturated ammonium chloride solution was added, and the obtained aqueous phase was extracted with ethyl acetate.
  • Compound 1c can also be prepared as follows:
  • CisbiokinEASESTK kit and PromegaADP-Glo kinase kit was used to detect the inhibitory activity of compounds on RIP1.
  • the test compound was dissolved in DMSO to obtain a 10 mM stock solution.
  • Compound I was further diluted to 100 times the concentration to be measured with DMSO.
  • Compound 1 to be tested was diluted with kinase buffer (HEPES 250 mM, NaN 3 0.1%, BSA 0.05%, Orthovanadate 0.5 mM, pH 7.0) at a ratio of 1:40. Add 2 ⁇ L of the diluted compound to a 384-well plate.
  • kinase buffer HPES 250 mM, NaN 3 0.1%, BSA 0.05%, Orthovanadate 0.5 mM, pH 7.0
  • Human RIP1, STK substrate S3, and ATP were diluted with kinase buffer to a concentration of 5 ng / ⁇ L, 10 ⁇ M, and 250 ⁇ M, respectively. Then, to a 348-well plate to which Compound I has been added, diluted 1 ⁇ L of human RIP1, 1 ⁇ L of STK substrate S3, and 1 ⁇ L of ATP were sequentially added. The final concentrations of human RIP1, STK substrate S3, and ATP in the reaction wells were 1 ng / ⁇ L, 2 ⁇ M, and 50 ⁇ M, respectively.
  • Compound I human RIP1, substrate and ATP were incubated together at room temperature for 3 hours. After 3 hours, 5 ⁇ L of Promega ADP-Glo reagent was added. Stop the reaction and remove the remaining ATP. After stopping the reaction, the mixture was incubated at room temperature for 40 minutes.
  • Human monocyte leukemia U937 cells were used to test the in vitro activity of RIP1 kinase inhibitors in a viability assay to rescue TNF-alpha / Z-VAD-FMK-induced necrotic cells.
  • U937 cell culture was performed using RMPI 1640 medium containing 10% fetal bovine serum, 100 units / mL penicillin, and 100 ug / mL streptomycin.
  • test compound was dissolved in DMSO to obtain a 10 mM stock solution of the test compound. Dilute with DMSO to 100 times the final reaction concentration. Dissolve TNF ⁇ to 100 ⁇ g / mL with phosphate buffered saline containing 0.1% bovine serum albumin, and freeze it for future use. Z-VAD-FMK was dissolved in DMSO to obtain a 50 mM stock solution for later use.
  • test compound was diluted with the medium at a ratio of 1:25, and TNF-alpha and Z-VAD-FMK were diluted with the medium to 400 ng / ml and 400 uM.
  • TNF-alpha and Z-VAD-FMK were 100 ng / mL and 100 ⁇ M, respectively.
  • the cells were incubated in a 37 ° C, 5% CO 2 incubator for 24 hours. After 24hr, add 100ul Cell Titer Glo to each well, and place it at room temperature in the dark for 10min after shaking. Luminescence signals were read with a PerkinElmer EnVision microplate reader.
  • Mouse-derived mTNF- ⁇ was purchased from Nanjing Kingsray Biological Company and Z-VAD-FMK was purchased from Selleck Company. 30ug mTNF was dissolved in 200ul each time. -free PBS and Z-VAD-FMK were dissolved in 5ul DMSO per 0.25mg, and then diluted with 195ul endotoxin-free PBS.
  • TNF-induced systemic inflammatory response mouse model Male C57Bl / 6J mice were selected for 8-10 weeks. Shave (upper abdomen), grouped (5 per group), and measure the average 3 times with a thermometer (OMRON Infrared Electronic Thermometer MC-872).
  • TNF-induced systemic inflammatory response syndrome model establishment process This experiment was divided into four groups: (1) blank control group, first 100 ⁇ l corn oil (100ul / mouse, gavage) 20 minutes later, PBS (200ul / small Rat) Tail vein injection was used to measure body temperature every 30 minutes: (2) In the TNF-induced group, 100 ⁇ l of corn oil (100 ul / mouse) was administered to the stomach for 20 minutes, and mTNF- ⁇ (30 ⁇ g / mouse, 200 ul / mouse) was administered to the tail.
  • Intravenous injection and Z-VAD-FMK (0.25mg / only, 200ul / only) intraperitoneally measured body temperature every 30min;
  • TNF Compound 1 treatment group compound 1 (10mg / kg) mixed with corn oil 100 ⁇ l (100ul / only small) (Rat) 20 minutes after gavage, body temperature was measured every 30 minutes after mTNF- ⁇ (30 ⁇ g / piece, 200 ul / mouse) tail vein injection and Z-VAD-FMK (0.25 mg / piece, 200 ul / mouse) intraperitoneal injection;
  • TNF + GSK-2982772 existing RIP1 inhibitor
  • first GSK-2982772 (10mg / kg) mixed with corn oil 100 ⁇ l (100ul / mouse) was administered orally for 20 minutes, and mTNF- ⁇ (30 ⁇ g / mouse, 200ul / mouse) tail vein injection and Z-VAD-FMK (0.25mg / mouse, 200ul / mouse) intraperitoneal injection to measure body

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Abstract

本发明提供了作为受体相互作用蛋白1(RIP1)激酶抑制剂的杂环化合物。具体地，本发明提供了如式I所示的化合物、或其药学上可接受的盐、或其药学上可接受的溶剂合物。式I化合物对蛋白激酶(如RIP1)具有高效、高选择的抑制作用。

Description

作为受体相互作用蛋白1(RIP1)激酶抑制剂的杂环化合物 技术领域
本发明属于一种医药化学领域，具体涉及一种作为受体相互作用蛋白1(RIP1)激酶抑制剂的杂环化合物及其药学上可接受的盐。
背景技术
受体相互作用蛋白1(RIP1)激酶是一类丝氨酸/苏氨酸蛋白激酶，可以调节核因子kB的活性，是决定细胞生存和死亡的交叉点，是程序性坏死(necroptosis)信号通路中关键的调控因子。程序性细胞死亡对于个体发育，机体稳态维持及病理过程等诸多方面都有着重要作用。细胞死亡主要包括凋亡、坏死和自噬等方式。其中，细胞坏死(necrosis)长期以来都被认为是一种被动且不可调控的过程。然而近几年研究表明细胞坏死也是受到精密调控的。不同于细胞凋亡(apoptosis)，细胞坏死会激活机体的免疫应答，机体局部的细胞坏死会引起全局性的生理病理反应。因此它参与了众多病理过程，例如病毒感染(viral infection)的免疫激活、缺血性坏死损伤(ischemic injuries)、以及神经退行性疾病(neurodegenerative disease)发生及发展等等。相对于研究较为成熟的细胞凋亡，细胞坏死还是一个崭新的领域。
程序性细胞坏死是一种由激酶的激活而引发的细胞死亡方式。死亡受体(例如TNFR1)的激活最终可以诱发程序性细胞坏死，而起动细胞坏死的信号传导主要依赖于激酶RIP1和RIP3的调控。细胞坏死发生之后，RIP1与RIP3相结合并激活了RIP3的激酶活性，然后RIP3发生自身磷酸化，使其可以特异性地和底物MLKL相结合，MLKL进而被RIP3磷酸化。这时的RIP1/RIP3/MLKL形成一个有活性的细胞坏死复合体，传递死亡信号给下游，使程序性细胞坏死得以最终发生。程序性细胞坏死的细胞会向周围释放其内容物，这些内容物作为DAMPs(damage-associated molecular patterns)，可刺激周围细胞发生炎症反应，激活机体免疫应答。
程序性细胞坏死有着重要的病理生理作用，如心肌梗死、胰腺炎、缺血再灌注损伤、脓血症、中风、冠心病、炎症性肠病、视网膜炎、酒精性脂肪肝、非酒精性脂肪肝、多发性硬化、皮炎、银屑病、慢性肾病、急性肾病、自身免疫性肝炎、乙型肝炎、丙型肝炎、急性肝功能不全、神经变性疾病、渐冻人症、帕金森、老年痴呆、骨质疏松、关节炎、细菌感染、癌症、动脉粥样硬化、心衰、慢性阻塞性肺病、糖尿病、血糖调控、器管移植过程中的保护、抗衰老、肥胖症、以及其它许多和RIP1激酶有关联的疾病。考虑到RIP1激酶靶向治疗这一研究发现的重要性，避免其有可能会对机体的其他细胞造成不必要的副作用，为了改善和程序性细胞坏死有关联的疾病，因此急需寻找一种有效的，高选择性的RIP1激酶抑制剂。
第一个RIP1激酶抑制剂Necrostatin 1(Nec-1)被发现后，又有许多不同结构的RIP1 激酶抑制剂被发现。
然而，目前已有的RIP1激酶抑制剂的活性、安全性尚难以令人满意，因此本领域迫切需要开发一种有效的、安全和/或高选择性的RIP1激酶小分子抑制剂。
发明内容
本发明的目的就是提供一种有效的、安全和/或高选择性的RIP1激酶小分子抑制剂。因此，本发明还提供了和RIP1激酶相关联的疾病的有效的预防和/或治疗的方法。
在本发明的第一方面，提供了一种杂环化合物、或其药学上可接受的盐、或其药学上可接受的溶剂合物，其中，所述化合物如式I所示：
Figure PCTCN2019103367-appb-000001
其中，R 1表示1-3个选自下组的取代基：H、卤素、-OH、取代或未取代的C1-C6烷基、取代或未取代的C2-C6烯基、取代或未取代的C2-C6炔基、取代或未取代的C3-C10环烷基、取代或未取代的C3-C10杂环烷基、取代或未取代的C6-C20芳基、取代或未取代的C3-C10杂芳基、C1-C4羧基、-CN、NO 2；
R 2为取代或未取代的C6-C20芳基、取代或未取代的C3-C10杂芳基或取代或未取代的5-8元杂环；
其中，所述取代是指一个或多个氢原子各自独立地被选自下组的取代基所取代：H、卤素、-CN、NO 2、-OH、-NRaRb、C1-C4烷基、C1-C4卤代烷基、C2-C6烯基、C2-C6炔基、C3-C6环烷基、苯基、苄基、C6-C10芳基、C3-C10杂芳基，
其中Ra和Rb各自独立地为H、C1-C3烷基、C3-C6环烷基。
在另一优选例中，R 1选自下组：H、卤素、C1-C6烷基、卤代C1-C6烷基。
在另一优选例中，R 1选自下组：H、卤素、C1-C6烷基。
在另一优选例中，R 1选自下组：H、F、Cl、甲基、乙基、正丙基、异丙基。
在另一优选例中，R 1为H或F。
在另一优选例中，R 1为H。
在另一优选例中，R 2为取代或未取代的苯基或5-6元杂芳基。
在另一优选例中，所述的5-8元杂环含有1-3个选自N、O、或S的杂原子。
在另一优选例中，所述的5-8元杂环是饱和的、不饱和的、或部分不饱和的。
在另一优选例中，R 1表示2个取代基。
在另一优选例中，R 1表示R 8和R 9；且R 8和R 9各自独立地选自下组：H、卤素、-OH、取代或未取代的C1-C6烷基、取代或未取代的C2-C6烯基、取代或未取代的C2-C6炔基、取代或未取代的C3-C10环烷基、取代或未取代的C3-C10杂环烷基、取代或未取代的C6-C20芳基、取代或未取代的C3-C10杂芳基、C1-C4羧基、-CN、NO 2。
在另一优选例中，
Figure PCTCN2019103367-appb-000002
为
Figure PCTCN2019103367-appb-000003
其中，R 8和R 9的定义如前所述。
在另一优选例中，所述化合物如式Ia所示
Figure PCTCN2019103367-appb-000004
其中，R 2、R 8和R 9的定义如前所述。
在另一优选例中，R 8和R 9各自独立地选自下组：H、卤素、C1-C6烷基、卤代C1-C6烷基。
在另一优选例中，R 8和R 9各自独立地选自下组：H、F。
在另一优选例中，R 8和R 9各自独立地位于苯环的3位或5位。
在另一优选例中，
Figure PCTCN2019103367-appb-000005
为
Figure PCTCN2019103367-appb-000006
在另一优选例中，所述化合物如式Ib所示
Figure PCTCN2019103367-appb-000007
其中，R 2、R 8和R 9的定义如前。
在另一优选例中，所述化合物如式II所示
Figure PCTCN2019103367-appb-000008
其中，
M、Q、V、Y及Z各自独立地选自下组：C、N、S、或O；
R 3、R 4、R 5、R 6及R 7各自独立地选自下组：H、卤素、-CN、C1-C6烷基、C2-C6 烯基、C2-C6炔基、C3-C10环烷基、C3-C10杂环烷基、C3-C10芳基、C3-C10杂芳基、-CN、NO 2；
Figure PCTCN2019103367-appb-000009
表示单键或双键。
在另一优选例中，式II中，M、Q、V、Y及Z中任意一个或多个(如1～3个)各自独立地为N、S或O，以及其余的为C。
在另一优选例中，式II中，M、Q、V、Y及Z各自独立地为C或N。
在另一优选例中，式II中，V及Z为N，以及M、Q及Y为C。
在另一优选例中，式II中，R 3、R 4、R 5、R 6及R 7中任意一个或多个(如1～3个)各自独立地选自下组：卤素、-CN、C1-C6烷基、C2-C6烯基、C2-C6炔基、C3-C10环烷基、C3-C10杂环烷基、C3-C10芳基、C3-C10杂芳基、-CN、NO 2；以及其余为H。
在另一优选例中，式II中，R 6选自下组：卤素、-CN、C1-C6烷基、C2-C6烯基、C2-C6炔基、C3-C10环烷基、C3-C10杂环烷基、C3-C10芳基、C3-C10杂芳基、-CN、NO 2；以及R 3、R 4、R 5及R 7为H。
在另一优选例中，R 6为-CN。
在另一优选例中，Z-R 3、Y-R 4、V-R 5、M-R 6和Q-R 7各自独立地选自：-O-、-S-、-N＝、-CR c＝、＝N-、＝CR c-；
其中，R c各自独立地选自：H、卤素、-CN、C1-C6烷基、C2-C6烯基、C2-C6炔基、C3-C10环烷基、C3-C10杂环烷基(较佳地，C3-C5环烷基)、C3-C10芳基、C3-C10杂芳基、NO 2(较佳地，R c各自独立地选自：H、卤素、-CN、C1-C6烷基、C3-C10环烷基(较佳地，C3-C5环烷基)、NO 2；更佳地，选自下组：-CN、C1-C6烷基)。
在另一优选例中，所述化合物如式III所示
Figure PCTCN2019103367-appb-000010
其中，
Q、V、Y及Z各自独立地选自下组：C、N、S、或O；
R 3、R 4、R 5及R 6各自独立地选自下组：无、H、卤素、-CN、C1-C6烷基、C2-C6烯基、C2-C6炔基、C3-C10环烷基、C3-C10杂环烷基、C3-C10芳基、C3-C10杂芳基、-CN、NO 2；
Figure PCTCN2019103367-appb-000011
表示单键或双键。
在另一优选例中，式III中，V为C；Y和Z为N；以及M为氧。
在另一优选例中，式III中，R 3、R 4、R 5及R 6中任意一个或多个(如1或2个)各自独立地选自下组：卤素、-CN、C1-C6烷基、C2-C6烯基、C2-C6炔基、C3-C10环烷基、 C3-C10杂环烷基、C3-C10芳基、C3-C10杂芳基、-CN、NO 2；以及其余为H或无。
在另一优选例中，式III中，R 3、R 4及R 6为无；且R 5选自下组：C1-C6烷基、C3-C10杂环烷基、-CN。
在另一优选例中，Z-R 3、Y-R 4、V-R 5、M-R 6和Q-R 7各自独立地选自：-O-、-S-、-N＝、-CR c＝、＝N-、＝CR c-；
其中，R c各自独立地选自：H、卤素、-CN、C1-C6烷基、C2-C6烯基、C2-C6炔基、C3-C10环烷基、C3-C10杂环烷基(较佳地，C3-C5环烷基)、C3-C10芳基、C3-C10杂芳基、NO 2(较佳地，R c各自独立地选自：H、卤素、-CN、C1-C6烷基、C3-C10环烷基(较佳地，C3-C5环烷基)、NO 2；更佳地，选自下组：-CN、C1-C6烷基)。
在另一优选例中，R 2选自下组：
Figure PCTCN2019103367-appb-000012
在另一优选例中，所述化合物选自下组：
Figure PCTCN2019103367-appb-000013
在本发明的第二方面，提供了一种药物组合物，所述的药物组合物包括：
(i)安全有效量的如第一方面所述的杂环化合物、或其药学上可接受的盐、或其药学上可接受的溶剂合物；以及
(ii)药学上可用的载体。
在本发明的第三方面，提供了如第一方面所述的杂环化合物、或其药学上可接受的盐、或其药学上可接受的溶剂合物的用途，它们被(i)用于制备蛋白激酶抑制剂，(ii)用于制备治疗蛋白激酶相关的疾病的药物，(iii)用于制备治疗TNF相关疾病的药物，和/或(iv)用于制备治疗程序性细胞坏死相关疾病的药物。
在另一优选例中，所述蛋白激酶是丝氨酸/苏氨酸蛋白激酶。
在另一优选例中，所述蛋白激酶是受体相互作用蛋白1(RIP1)激酶。
在另一优选例中，所述蛋白激酶相关的疾病包括：心肌梗死、胰腺炎、缺血再灌注损伤、脓血症、中风、冠心病、炎症性肠病、视网膜炎、酒精性脂肪肝、非酒精性脂肪肝、多发性硬化、皮炎、银屑病、慢性肾病、急性肾病、自身免疫性肝炎、乙型肝炎、丙型肝炎、急性肝功能不全、神经变性疾病、渐冻人症、帕金森、老年痴呆、骨质疏松、关节炎、细菌感染、癌症、动脉粥样硬化、心衰、慢性阻塞性肺病、糖尿病、血糖调控、器管移植过程中的保护、抗衰老，和/或肥胖症。
在本发明的第四方面，提供了一种如第一方面所述的杂环化合物、或其药学上可接受的盐、或其药学上可接受的溶剂合物的制备方法，
(i)所述制备方法为方法一；且方法一包括步骤：使式8化合物与R 2-X反应从而生成式I化合物；
Figure PCTCN2019103367-appb-000014
其中，X为Cl或Br；R 1和R 2的定义如前所述；
或者，
(ii)所述制备方法为方法二；且方法二包括步骤：使式9化合物与式5化合物反应从而生成式I化合物；
Figure PCTCN2019103367-appb-000015
其中，R 1和R 2的定义如前所述。
在另一优选例中，所述方法一包括步骤：
(1)将式6化合物与式5化合物进行反应，从而得到式7化合物；
Figure PCTCN2019103367-appb-000016
(2)对所述式7化合物进行脱保护基团，从而得到式8化合物；以及
Figure PCTCN2019103367-appb-000017
(3)将所述式8化合物与R 2-X进行反应，从而得到式I化合物；
Figure PCTCN2019103367-appb-000018
其中，R 1、R 2的定义同前所述；
R 10为保护基团。
X为Cl或Br。
在另一优选例中，R 10为-BOC(叔丁氧羰基)。
在另一优选例中，X为Cl。
在另一优选例中，方法二中，化合物9通过下述步骤制备：
在惰性溶剂中，在碱性条件下，使化合物10反应生成化合物9
Figure PCTCN2019103367-appb-000019
在另一优选例中，式5化合物的制备方法包括步骤：
(1.1)将式1化合物与式2化合物进行反应，从而得到式3化合物
Figure PCTCN2019103367-appb-000020
(1.2)将式3化合物与甲基磺酰氯进行反应，从而得到式4化合物
Figure PCTCN2019103367-appb-000021
(1.3)在惰性溶剂中，在三氟乙酸(TFA)存在下，使式4化合物进行反应，从而得到式5化合物
Figure PCTCN2019103367-appb-000022
在本发明的第五方面，提供了一种体外非治疗性抑制蛋白激酶的方法，其中，所述方法包括步骤将如第一方面所述的杂环化合物、或其药学上可接受的盐、或其药学上可接受的溶剂合物与所述蛋白激酶接触从而抑制所述蛋白激酶的活性。
在另一优选例中，所述的蛋白激酶为受体相互作用蛋白1激酶。
在本发明的第六方面，提供了一种体内抑制治疗或非治疗性抑制蛋白激酶的方法，其中，所述方法包括步骤：
向对象(subject)施用有效量的如第一方面所述的杂环化合物、或其药学上可接受的盐、或其药学上可接受的溶剂合物或者如第二方面所述的药物组合物，从而抑制所述蛋白激酶的活性。
在另一优选例中，所述的对象为动物；优选地为哺乳动物；更优选地，为人类。
在本发明的第七方面，提供了一种治疗蛋白激酶相关的疾病、TNF相关疾病、和/或程序性细胞坏死相关疾病的方法，所述方法包括步骤：
向目标施用有效量的如第一方面所述的杂环化合物、或其药学上可接受的盐、或其药学上可接受的溶剂合物或者如第二方面所述的药物组合物。
在另一优选例中，所述的目标为动物；优选地为哺乳动物；更优选地，为人类。
在本发明的第八方面，提供了一种制备方法，所述方法包括步骤：
(a)在惰性溶剂中，任选地在三氟乙酸(TFA)存在下，使式4化合物进行反应，从而得到式5化合物
Figure PCTCN2019103367-appb-000023
式中，R 1的定义同第一方面中定义。
在另一优选例中，步骤(a)中，所述的惰性溶剂选自为二氯甲烷。
在另一优选例中，步骤(a)的反应的反应温度为0～100℃；较佳地，0～50℃；更佳地，10～30℃。
在另一优选例中，步骤(a)的反应的反应时间为0.1～20小时；较佳地，0.5～10小时；更佳地，1～5小时；最佳地。1～3小时。
在另一优选例中，所述方法还包括步骤：
(b1)将式9化合物与式5化合物进行反应，从而得到式I化合物；
Figure PCTCN2019103367-appb-000024
式中，R 1、R 2的定义同第一方面中定义。
在另一优选例中，所述方法还包括步骤：
(c1)将式6化合物与式5化合物进行反应，从而得到式7化合物；
Figure PCTCN2019103367-appb-000025
(c2)对所述式7化合物进行脱保护基团，从而得到式8化合物；以及
Figure PCTCN2019103367-appb-000026
(c3)将所述式8化合物与R 2-X进行反应，从而得到式I化合物；
Figure PCTCN2019103367-appb-000027
式中，R 1、R 2的定义同第一方面中定义；R 10为保护基团；X为Cl或Br。
应理解，在本发明范围内中，本发明的上述各技术特征和在下文(如实施例)中具体描述的各技术特征之间都可以互相组合，从而构成新的或优选的技术方案。限于篇幅，在此不再一一累述。
附图说明
图1显示了本发明一个实施例(即实施例1)中的化合物1(NHWD-1062)可有效治疗TNF诱导的全身炎症反应综合征。
具体实施方式
本发明人经过广泛而深入的研究，首次开发了一类具有新颖结构、抑制活性显著、且安全性高的的杂环化合物。该类化合物对受体相互作用蛋白1(RIP1)激酶有优异的、选择性高的抑制作用，可用作RIP1激酶抑制剂。在此基础上完成了本发明。
术语
如本文所用，术语“烷基”是指饱和的直链或支链烷基，如甲基、乙基、异丙基。此外，C1-C6烷基指含有1-6个碳原子的烷基。
如本文所用，术语“烯基”是指包含至少一个双键的直链或支链烃基部分，例如-CH＝CH-CH3。
如本文所用，术语“炔基”是指包含至少一个三键的直链或支链烃基部分，例如-C≡C-CH3。
如本文所用，术语“环烷基”是指饱和或不饱和的环状烃基部分，例如环己基。
如本文所用，术语“杂环烷基”是指包含至少一个环杂原子(例如N，O或S)的饱和的环状部分，例如4-四氢吡喃基。
如本文所用，术语“芳基”是指包含一个或多个芳环的烃基部分。芳基部分的例子包括但不限于苯基(Ph)、萘基、芘基、蒽基和菲基。
如本文所用，术语“杂芳基”是指单环，二环或稠环的芳香族基团，所述基团具有特定的成环碳原子数(例如，C 4-10即具有4-10个成环碳原子)，且包括至少一个相同或不同的选自N、O或S的杂原子。每个环上原子可以被任意取代。所述的杂芳基可以是5-至15-元的，具有1-5个各自独立地选自N、O或S的杂原子的芳香环基。杂芳基的例子例如(但并不限于)：吡啶、嘧啶、吡咯、吲唑、吲哚、呋喃、苯并呋喃、噻吩，或类 似基团。
如本文所用，取代是指被一个或多个取代基取代。
如本文所用，术语“取代”(在有或无“任意地”修饰时)指特定的基团上的一个或多个氢原子被特定的取代基所取代。特定的取代基为在前文中相应描述的取代基，或各实施例中所出现的取代基。除非特别说明，某个任意取代的基团可以在该基团的任何可取代的位点上具有一个选自特定组的取代基，所述的取代基在各个位置上可以是相同或不同的。环状取代基，例如杂环烷基，可以与另一个环相连，例如环烷基，从而形成螺二环系，例如，两个环具有一个共用碳原子。本领域技术人员应理解，本发明所预期的取代基的组合是那些稳定的或化学上可实现的组合。
如本文所用，除非特别说明，术语“药学上可接受的盐”指适合与对象(例如，人)的组织接触，而不会产生不适度的副作用的盐。在一些实施例中，本发明的某一化合物的药学上可接受的盐包括具有酸性基团的本发明的化合物的盐(例如，钾盐、钠盐、镁盐、钙盐)或具有碱性基团的本发明的化合物的盐(例如，硫酸盐、盐酸盐、磷酸盐、硝酸盐、碳酸盐)。
应当理解，在本文中，一个原子所连接的基团数量超过其稳定价态时可连接的最大基团数量时，其连接的基团定义中的H是指无。例如，式II中，当V或Z为N时，与V或Z相连的R 5和R 3基团是指无。
活性成分
如本文所用，术语“本发明化合物”或“本发明活性成分”可互换使用，指本发明第一方面中的式I所示的杂环化合物、或其药学上可接受的盐、或溶剂合物、水合物、消旋体、立体异构体、或前药。
如本文所用，术语“药学上可接受的盐”指本发明化合物与药学上可接受的无机酸和有机酸所形成的盐，其中，优选的无机酸包括(但并不限于)：盐酸、氢溴酸、磷酸、硝酸、硫酸；优选的有机酸包括(但并不限于)：甲酸、乙酸、丙酸、丁二酸、萘二磺酸(1,5)、亚细亚酸、草酸、酒石酸、乳酸、水杨酸、苯甲酸、戊酸、二乙基乙酸、丙二酸、琥珀酸、富马酸、庚二酸、己二酸、马来酸、苹果酸、氨基磺酸、苯丙酸、葡糖酸、抗坏血酸、烟酸、异烟酸、甲磺酸、对甲苯磺酸、柠檬酸，以及氨基酸。
如本文所用，术语“药学上可接受的溶剂合物”指本发明化合物与药学上可接受的溶剂形成溶剂合物，其中，所述药学上可接受的溶剂包括(但并不限于)：水、乙醇、甲醇、异丙醇、四氢呋喃、二氯甲烷。本发明的某些化合物可以非溶剂化形式以及溶剂化形式存在，包括水化形式。溶剂化形式通常与非溶剂化形式等价，应包括在本发明范围内。本发明的某些化合物可以多晶型或无定形形式存在。通常，就本发明所考虑的应用而言，所有物理形式是等价的，应包括在本发明范围内。
在本发明中，除了盐形式之外，还提供了呈前药形式的化合物。本文所述化合物的 前药是在生理条件下易于发生化学变化以提供本文提供的化合物的那些化合物。此外，前药可以通过化学或生物化学方法在离体环境中转化为本文提供的化合物。例如，当将前药置于具有合适的酶或化学试剂的透皮贴剂储库中时，可以将其缓慢转化为本文提供的化合物。优选地，所述的前药为酯形式的式I化合物。
本发明的某些化合物拥有不对称碳原子(光学中心)或双键；消旋体、非对映体、几何异构体、区域异构体和单独的异构体(例如，分离的对映体)均应包括在本发明范围内。当本文提供的化合物具有确定的立体化学(表示为R或S，或具有虚线或楔形键指明)时，被本领域技术人员将理解那些化合物为基本上不含其他异构体(例如至少80％，90％，95％，98％，99％和至多100％不含其他异构体)。
本发明化合物还可在构成此类化合物的一个或多个同位素原子处含有非天然比例的原子同位素。某同位素的非天然比例可以定义为从所讨论原子的天然发现的量到100％该原子的量。例如，化合物可以掺入放射性同位素，例如氚( 3H)、碘-125( 125I)或碳-14( 14C)，或非放射性同位素，例如氘( 2H)或碳-13( 13C)。除了本申请所述的那些用途，此类同位素变体可提供额外的用途。例如，本发明化合物的同位素变体可以有额外的用途，包括但不限于作为诊断的和/或成像试剂，或作为细胞毒性/放射毒性治疗剂。另外，本发明化合物的同位素变体可具有改变的药代动力学和药效学特征，从而有助于增加治疗期间的安全性、耐受性或疗效。无论是否有放射性，本发明化合物的所有同位素变体均应包括在本发明范围内。
制备方法
对于根据本发明的式I化合物，可以用有机合成化学领域及技术人员熟知的各种方法制备。可以使用下文中所描述的方法，与有机化学领域己知的合成方法一起或本领域技术人员所理解的在其上的变化来合成本发明化合物。
本发明所述式I化合物的制备方法可以从易于获得的起始原料使用以下的一般性方法和过程来制备本发明的化合物。将理解的是当给出典型的或优选的工艺操作条件(即反应温度、时间、反应物的摩尔、溶剂、压力等)时，还可以使用其他工艺操作条件，除非另有说明。最佳反应条件可以随所用的具体反应物或溶剂而变化，但这些条件可以由本领域技术人员通过常规最佳过程加以确定。
在此所描述的本发明所述式I化合物的方法可以根据本领域己知的任何适宜的方法加以监控。例如，核磁共振，质谱分析，HPLC，薄层色谱来监控产物的生成。化合物的制备可以涉及多个化学基团的保护和脱保护。对于保护和脱保护的需要，以及对适当的保护基的选择可以由本领域技术人员容易的加以确定.
具体地，本发明式I化合物的合成路线如下所示：
路线一
Figure PCTCN2019103367-appb-000028
式中，R 1、R 2、R 10和X的定义如上所述。
或者，路线二
Figure PCTCN2019103367-appb-000029
Figure PCTCN2019103367-appb-000030
其中，式5化合物的合成方法参照路线一所示。
通常，可以采用如上所述的反应路线及工艺制备本发明化合物，但并不限于反应条件中的试剂和溶剂。
此外，本发明还提供了一种高收率和高ee值的制备方法；该方法包括步骤：
(a)在惰性溶剂中，在三氟乙酸存在下，使式4化合物进行反应，从而得到式5化合物
Figure PCTCN2019103367-appb-000031
式中，R 1如前定义。
药物组合物
本发明还提供了一种药物组合物，其具有优秀的、高选择的蛋白激酶抑制作用，该组合物包括：(i)作为活性成分的式I化合物、或其药学上可接受的盐、或其药学上可接受的溶剂合物，以及(ii)一种或多种药学上可用的载体。
可将化合物本身或其药学上可接受的盐或其药学上可接受的溶剂合物与可药用赋形剂、稀释剂等的混合物以片剂、胶囊、颗粒剂、散剂或糖浆剂的形式口服给药，或以注射剂的形式非口服给药。该药物组合物优选含有重量比为0.01％-99％的本发明的式I化合物或其药学上可接受的盐或其药学上可接受的溶剂合物作为活性成分，更优选含有重量比为0.1％-90％的活性成分。
上述制剂可通过常规制药方法制备。可用的药用辅剂的例子包括赋形剂(例如糖类衍生物如乳糖、蔗糖、葡萄糖、甘露糖醇和山梨糖醇；淀粉衍生物如玉米淀粉、土豆淀粉、糊精和羧甲基淀粉；纤维素衍生物如结晶纤维素、羟丙基纤维素、羧甲基纤维素、羧甲基纤维素钙、羧甲基纤维素钠；阿拉伯胶；右旋糖酐；硅酸盐衍生物如偏硅酸镁铝；磷酸盐衍生物如磷酸钙；碳酸盐衍生物如碳酸钙；硫酸盐衍生物如硫酸钙等)、粘合剂(例如明胶、聚乙烯吡咯烷酮和聚乙二醇)、崩解剂(例如纤维素衍生物如羧甲基纤维素钠、聚乙烯吡咯烷酮)、润滑剂(例如滑石、硬脂酸钙、硬脂酸镁、鲸蜡、硼酸、苯甲酸钠、亮氨酸)、稳定剂(对羟基苯甲酸甲酯、对羟基苯甲酸丙酯等)、矫味剂(例如常用的甜味剂、酸味剂和香料等)、稀释剂和注射液用溶剂(例如水、乙醇和甘油等)。
本发明的化合物、其药学上可接受的盐或或其药学上可接受的溶剂合物，或其药物组合物的给药量随患者的年龄、性别、种族、病情等的不同而不同。
药物组合物和施用方法
由于本发明式I化合物具有优异的对蛋白激酶(受体相互作用蛋白1，RIP1)的抑制活性，因此本发明化合物及其各种晶型，药学上可接受的无机或有机盐，水合物或溶剂合物，以及含有本发明化合物为主要活性成分的药物组合物可用于治疗、预防以及缓解由与蛋白激酶(受体相互作用蛋白1，RIP1)活性或表达量相关的疾病。
在本发明中，本发明化合物可用于治疗以下疾病(但并不限于)：心肌梗死、胰腺炎、缺血再灌注损伤、脓血症、中风、冠心病、炎症性肠病、视网膜炎、酒精性脂肪肝、非酒精性脂肪肝、多发性硬化、皮炎、银屑病、慢性肾病、急性肾病、自身免疫性肝炎、乙型肝炎、丙型肝炎、急性肝功能不全、神经变性疾病、渐冻人症、帕金森、老年痴呆、骨质疏松、关节炎、细菌感染、癌症、动脉粥样硬化、心衰、慢性阻塞性肺病、糖尿病、血糖调控、器管移植过程中的保护、抗衰老、肥胖症，以及其它与RIP1激酶活性过高相关的疾病。
本发明的药物组合物包含安全有效量范围内的本发明化合物或其药理上可接受的盐及药理上可以接受的赋形剂或载体。其中“安全有效量”指的是：化合物的量足以明显改善病情，而不至于产生严重的副作用。通常，药物组合物含有1-2000mg本发明化合物/剂，更佳地，含有5-200mg本发明化合物/剂。较佳地，所述的“一剂”为一个胶囊或药片。
“药学上可以接受的载体”指的是：一种或多种相容性固体或液体填料或凝胶物质，它们适合于人使用，而且必须有足够的纯度和足够低的毒性。“相容性”在此指的是组合物中各组份能和本发明的化合物以及它们之间相互掺和，而不明显降低化合物的药效。药学上可以接受的载体部分例子有纤维素及其衍生物(如羧甲基纤维素钠、乙基纤维素钠、纤维素乙酸酯等)、明胶、滑石、固体润滑剂(如硬脂酸、硬脂酸镁)、硫酸钙、植物油(如豆油、芝麻油、花生油、橄榄油等)、多元醇(如丙二醇、甘油、甘露醇、山梨醇等)、乳化剂(如吐温
Figure PCTCN2019103367-appb-000032
)、润湿剂(如十二烷基硫酸钠)、着色剂、调味剂、稳定剂、抗氧化剂、防腐剂、无热原水等。
本发明化合物或药物组合物的施用方式没有特别限制，代表性的施用方式包括(但并不限于)：口服、瘤内、直肠、肠胃外(静脉内、肌肉内或皮下)、和局部给药。
用于口服给药的固体剂型包括胶囊剂、片剂、丸剂、散剂和颗粒剂。在这些固体剂型中，活性化合物与至少一种常规惰性赋形剂(或载体)混合，如柠檬酸钠或磷酸二钙，或与下述成分混合：(a)填料或增容剂，例如，淀粉、乳糖、蔗糖、葡萄糖、甘露醇和硅酸；(b)粘合剂，例如，羟甲基纤维素、藻酸盐、明胶、聚乙烯基吡咯烷酮、蔗糖和阿拉伯胶；(c)保湿剂，例如，甘油；(d)崩解剂，例如，琼脂、碳酸钙、马铃薯淀粉或木薯淀粉、藻酸、某些复合硅酸盐、和碳酸钠；(e)缓溶剂，例如石蜡；(f)吸收加速剂，例如，季胺化合物；(g)润湿剂，例如鲸蜡醇和单硬脂酸甘油酯；(h)吸附剂，例如，高岭土；和(i)润滑剂，例如，滑石、硬脂酸钙、硬脂酸镁、固体聚乙二醇、十二烷基 硫酸钠，或其混合物。胶囊剂、片剂和丸剂中，剂型也可包含缓冲剂。
固体剂型如片剂、糖丸、胶囊剂、丸剂和颗粒剂可采用包衣和壳材制备，如肠衣和其它本领域公知的材料。它们可包含不透明剂，并且，这种组合物中活性化合物或化合物的释放可以延迟的方式在消化道内的某一部分中释放。可采用的包埋组分的实例是聚合物质和蜡类物质。必要时，活性化合物也可与上述赋形剂中的一种或多种形成微胶囊形式。
用于口服给药的液体剂型包括药学上可接受的乳液、溶液、悬浮液、糖浆或酊剂。除了活性化合物外，液体剂型可包含本领域中常规采用的惰性稀释剂，如水或其它溶剂，增溶剂和乳化剂，例知，乙醇、异丙醇、碳酸乙酯、乙酸乙酯、丙二醇、1,3-丁二醇、二甲基甲酰胺以及油，特别是棉籽油、花生油、玉米胚油、橄榄油、蓖麻油和芝麻油或这些物质的混合物等。
除了这些惰性稀释剂外，组合物也可包含助剂，如润湿剂、乳化剂和悬浮剂、甜味剂、矫味剂和香料。
除了活性化合物外，悬浮液可包含悬浮剂，例如，乙氧基化异十八烷醇、聚氧乙烯山梨醇和脱水山梨醇酯、微晶纤维素、甲醇铝和琼脂或这些物质的混合物等。
用于肠胃外注射的组合物可包含生理上可接受的无菌含水或无水溶液、分散液、悬浮液或乳液，和用于重新溶解成无菌的可注射溶液或分散液的无菌粉末。适宜的含水和非水载体、稀释剂、溶剂或赋形剂包括水、乙醇、多元醇及其适宜的混合物。
用于局部给药的本发明化合物的剂型包括软膏剂、散剂、贴剂、喷射剂和吸入剂。活性成分在无菌条件下与生理上可接受的载体及任何防腐剂、缓冲剂，或必要时可能需要的推进剂一起混合。
本发明化合物可以单独给药，或者与其他药学上可接受的化合物联合给药。
使用药物组合物时，是将安全有效量的本发明化合物适用于需要治疗的哺乳动物(如人)，其中施用时剂量为药学上认为的有效给药剂量，对于60kg体重的人而言，日给药剂量通常为1～2000mg，优选5～500mg。当然，具体剂量还应考虑给药途径、病人健康状况等因素，这些都是熟练医师技能范围之内的。
本发明的主要优点包括：
(a)本发明化合物对RIP1有优异的体外和体内抑制活性。
(b)本发明化合物具有良好的安全性和选择性。
(c)本发明化合物的制法简便，产率高。
(d)本发明由式4化合物制备式5化合物的方法(成环方法)具有高收率和高ee值。
下面结合具体实施例，进一步阐述本发明。应理解，这些实施例仅用于说明本发明而不用于限制本发明的范围。下列实施例中未注明具体条件的实验方法，通常按照常规 条件，或按照制造厂商所建议的条件。除非另外说明，否则百分比和份数是重量百分比和重量份数。
实施例1化合物1(NHWD-1062)的制备
Figure PCTCN2019103367-appb-000033
步骤A
Figure PCTCN2019103367-appb-000034
将化合物1b,N-叔丁氧羰基羟胺(600mg,4.50mmol)加入到DMF(10ml)后，冰浴冷却到0℃，然后加入NaH(180mg,4.5mmol,60％)，加完后保持0℃反应半小时，然后再将这个混合液缓慢地滴加到含有化合物1a(700mg,4mmol)的DMF(5ml)溶液中，所得到的反应液先在0℃反应半小时，然后室温反应24小时，加入饱和氯化铵溶液，所得到的水相用乙酸乙酯萃取，合并有机相，无水硫酸钠干燥后，旋干溶液后过硅胶柱(流动相用PE：EA＝10:1-1:1)得到无色油状化合物1c，970mg，收率89％，直接用于下一步。
还可以按以下方法制备化合物1c：
将化合物1b(N-叔丁氧羰基羟胺)(542mg，4.10mmol)加入到DMF(10ml)后，冰浴冷却到0℃，然后加入氢钠(NaH)(180mg,4.5mmol,60％)，加完后保持0℃反应半小时，再缓慢滴加化合物1a,((R)-(+)-3-氯-1-苯基-1-丙醇)(350mg，2mmol)的DMF(5ml)溶液，所得到的反应液先在0℃反应半小时，然后室温反应3天，TLC显示反应结束，加入饱和氯化铵溶液，所得到的水相用乙酸乙酯萃取，合并有机相，无水硫酸钠干燥后，旋干溶液后过硅胶柱(流动相用PE：EA＝10:1-1:1)得到无色油状化合物1c。
步骤B
Figure PCTCN2019103367-appb-000035
将化合物3(4.4g,16.45mmol)加入到二氯甲烷(200ml)溶液中，然后加入三乙胺(4.5mmol,33mmol)，反应液用冰浴冷却到0℃，然后滴加甲基磺酰氯(1.4ml,19mmol)，加完后继续在0℃反应2小时，TLC显示反应结束，加入400ml水，分出有机相，水相再 用二氯甲烷萃取，合并有机相，饱和氯化钠洗涤后，无水硫酸钠干燥后旋干得到化合物1d,4.9g,收率87％,直接用于步骤C中，或者将旋干所得残余物过硅胶柱(流动相用PE：EA＝10:1-1:1)得到化合物1d。
步骤C
Figure PCTCN2019103367-appb-000036
将化合物1d(3.4g,10mmol)溶解到二氯甲烷(100ml)中，然后加入TFA(10ml)，室温搅拌2小时，TLC显示反应结束，将反应液加入到饱和碳酸氢钠中，分出有机相，用二氯甲烷萃取，合并有机相后，再用饱和碳酸氢钠溶液和饱和氯化钠洗涤，有机相用无水硫酸钠干燥后，旋干得到化合物1e，0.98g,收率66％,ee 92％，无进一步纯化直接用于下一步。
步骤D
Figure PCTCN2019103367-appb-000037
将化合物1f(1-叔丁氧羰酰哌啶-4-羧酸)(2.7g,12mmol)加入到DMF(40ml)溶液中，然后加入HATU(4.6g,12mmol)，室温搅拌1小时，然后将化合物1e(1.5g,10mml)加入到反应液中，室温搅拌过夜，TLC显示反应结束，加入100ml水，用乙酸乙酯萃取三次，合并有机相，水洗，饱和氯化钠洗，无水硫酸钠干燥后，旋干所得残余物过硅胶柱(流动相为PE：EA＝3:1，V:V)，得到目标产物为淡黄色固体化合物1g,3.2g,收率89％。MS(ESI)m/z:361.2(M+H+).1HNMR(400MHz，CDCl3)7.1-7.3(m，5H)，5.3(m，1H)，4.2(m，1H)，4.1(m，2H)，3.8(m,1H)，2.7-2.8(m,4H)，2.2-2.3(m，1H)，1.8(m，1H)，1.6(m，3H)，1.37(s，9H).
步骤E
Figure PCTCN2019103367-appb-000038
将化合物1g(720mg,2mmol)加入到无水二氯甲烷中(15ml)，然后冰浴下加入TFA(3ml)，室温反应两小时，TLC显示反应结束，将反应液加入到50ml饱和碳酸氢钠 溶液中，然后分出有机相，水相再用二氯甲烷萃取，合并有机相，水洗，饱和氯化钠洗，无水硫酸钠干燥后，旋干得化合物1h，520mg,收率100％无进一步纯化直接用于下一步。
步骤F
Figure PCTCN2019103367-appb-000039
将化合物1h(800mg,粗品)加入到DMF(8ml)溶液中，然后加入DBU(912mg，6mmol)，再加入化合物1i(6-氯-4-嘧啶甲腈)(552mg,4mmol)，80℃反应6小时，TLC显示原料化合物1h大量剩余，然后补加化合物9(6-氯-4-嘧啶甲腈)(276mg,2mmol))，反应温度升到100℃反应12小时，将反应液倒入到水中，用二氯甲烷萃取，合并有机相，水洗，饱和氯化钠洗，无水硫酸钠干燥后，旋干所得残余物过硅胶柱(流动相为PE：EA＝3:1),得到粗品，化合物1 0.98g,收率88％,进一步经过高效液相制备得到化合物1。
MS(ESI)m/z:364.4(M+H+).1HNMR(400MHz，CDCl3)8.6(s，1H)，7.2-7.4(m，5H)，6.8(s，1H)，5.4(m，1H)，4.3(m，2H)，3.9(m,1H)，3.1-3.2(m，3H)，2.8(m,1H)，2.4(m,1H)，2.1(m，1H)，1.7-1.9(m，3H)，1.3(m，1H).
实施例2化合物2的制备
Figure PCTCN2019103367-appb-000040
步骤A
Figure PCTCN2019103367-appb-000041
三口瓶中加入THF 1.5L,Mg(28.4g,1.17mol,1.1eq),碘粒2粒,N2保护,内温60℃,滴加少量的化合物2a(215g,1.11mol,1eq)的500ml THF,2小时后反应引发.再缓慢地滴加全部的化合物2a(215g,1.11mol,1eq)的500ml THF(55℃<反应內温<65℃),滴毕后回流2小时,冷却,待用.室温,5L三口瓶中加入THF 1L,N2保护,3-氯丙酰氯(141g,1.11mol,1eq),无水氯化锂(2.8g,0.06eq),无水氯化锰(4.18g,0.03eq),氯化亚铜(3.3g, 0.03eq),搅拌1小时后,0℃滴加到上述制备的格氏试剂中,搅拌2小时后,用HCl(1M)溶液淬灭,加水稀释后,用MTBE萃取,有机相旋干得粗品120g,过硅胶柱后得化合物2b 86g，收率38％。.MS(ESI)m/z:204,206(M+H+).1HNMR(400MHz,CDCl3)7.49(s,2H),7.07(s,1H),3.9(t,2H),3.4(t,2H).
步骤B
Figure PCTCN2019103367-appb-000042
2L的三口瓶中加入THF 500ml,再加入(S)-(-)-α，α-二苯基脯氨醇(7.42g,0.1e),硼酸三甲酯(0.14g,0.0045eq),N2保护,室温,搅拌2小时后,加入硼烷二甲基硫醚络合物(2M四氢呋喃溶液)(147ml,0.29mol,1eq),-5℃缓慢滴加化合物2b(60g,0.29mol,1eq)的500ml THF溶液,约3小时后,自然升温到室温,且搅拌过夜.冷却在冰浴,用300ml甲醇淬灭反应,再滴加HCl(1M).旋干溶剂后,用MTBE打浆,过滤.滤液旋干后得无色液体化合物2c 40g.GC 94％,ee 88％，收率63％,MS(ESI)m/z:206,208(M+H+).1HNMR(400MHz,CDCl3)6.9(m,2H),6.8(m,1H),5.0(m,1H),3.8(m,1H),3.6(m,1H),2.4(m,1H),2.1(m,2H).
步骤C
Figure PCTCN2019103367-appb-000043
在冰浴,N2保护,向1L的三口瓶中加入DMF 400ml,N-Boc-羟胺(26g,0.2mol,1.05eq),和分批加入NaH(8.14g,60％,0.2mol,1.05eq),搅拌2小时后,缓慢地将100ml的化合物2c(40g,0.19mol)DMF溶液滴加进去.再在30℃搅拌过夜.反应用水淬灭，用乙酸乙酯萃取，合并有机相后，再用饱和碳酸氢钠溶液和饱和氯化钠洗涤，有机相用无水硫酸钠干燥后,旋干得到化合物2d,56g，收率95％，直接用于下一步.MS(ESI)m/z:302.1(M-H+).1HNMR(400MHz,CDCl3)7.4(s,1H),7.0(d,2H),6.7(t,1H),5.0(m,1H),4.5(m,1H),4.0(m,2H),2.0(m,1H),1.9(m,1H),1.5(s,9H).
步骤D
Figure PCTCN2019103367-appb-000044
在冰浴,N2保护下,向含有化合物2d(56g,0.18mol,1eq),二氯甲烷500ml,三乙胺(37.3g,0.37mol,2eq)的1L三口瓶中缓慢滴加甲磺酰氯(23.3g,0.2mol,1.1eq)，室温搅拌1小时，加入100ml水，用乙酸乙酯萃取三次，合并有机相，水洗，饱和氯化钠洗，无水硫酸钠干燥后，旋干得到化合物2e，56g,收率80％，直接用于下一步。
步骤E
Figure PCTCN2019103367-appb-000045
将化合物2e(0.18mol)加入到无水300ml二氯甲烷中,然后冰浴下加入TFA 300ml，室温反应过夜.用泵尽量抽干溶剂后,用300ml二氯甲烷将残留液稀释,再用碳酸氢钠溶液调pH大于7,分出有机相，饱和氯化钠洗，无水硫酸钠干燥后,旋干得残留物23g,ee80％，反应收率69％.19g残留物中,加入D-(+)-二苯甲酰酒石酸33g(0.9eq)的150ml甲醇溶液,在30℃搅拌2小时,冷却到室温,析出固体.过滤,MTBE洗涤,ee 92％.滤饼用甲醇加热溶解,加50ml MTBE析出固体,ee 99.6％.用碳酸氢钠溶液游离,MTBE萃取,得无色液体化合物2f,6.7g.MS(ESI)m/z:185(M+H+).1HNMR(400MHz,CDCl3)7.0(m,2H),6.7(m,1H),4.9(m,1H),4.5(m,1H),4.1(m,1H),4.0(m,1H),2.7(m,1H),2.3(m,1H).
步骤F
Figure PCTCN2019103367-appb-000046
向1L单口瓶中加入6-氯-4-嘧啶甲腈(22.6g,0.16mol,1eq),200ml乙腈,DIPEA(48g,0.37mol,2.3eq),4-哌啶甲酸甲酯(27g,0.19mol,1.15eq),反应回流过夜.旋干所得残余物过硅胶柱得化合物2i,28g，收率69％。MS(ESI)m/z:247.2(M+H+).1HNMR(400MHz,CDCl3)8.6(s,1H),6.9(s,1H),4.3(m,2H),3.7(s,3H),3.2(m,2H),2.7(m,1H),2.1(m,2H),1.9(m,2H)).
步骤G
Figure PCTCN2019103367-appb-000047
向500ml单口瓶中加入化合物2i,100ml THF,加入LiOH(6.3g,0.15mol,1.5eq)的100ml水溶液,搅拌2小时.加MTBE分液后,水相调pH大约4,析出白色固体,过滤,MTBE洗涤,烘干得化合物2j,22.5g，收率87％。MS(ESI)m/z:233(M+H+).1HNMR(400MHz,DMSO)12.4(s,1H),8.6(s,1H),7.6(s,1H),4.3(m,2H),3.2(m,2H),2.6(m,1H),1.9(m,2H),1.5(m,2H)).
步骤H
Figure PCTCN2019103367-appb-000048
向100ml单口瓶中加入化合物2j(1.25g,1eq),无水30ml二氯甲烷,CDI(0.9g,1eq),和化合物2f(1g,1eq).在30℃搅拌过夜,旋干所得残余物过硅胶柱得白色固体.再用MTBE打浆,过滤,烘干得化合物2,1.5g，收率76％.MS(ESI)m/z:399.9(M+H+).1HNMR(400MHz,CDCl3)8.6(s,1H),7.0(m,1H),6.9(m,2H),6.8(m,1H),5.4(m,1H),4.6(m,3H),4.0(m,1H),3.3-3.1(m,3H),2.9(m,1H),2.4(m,1H),2.1(m,1H),1.9(m,3H).
实施例3化合物3的制备
Figure PCTCN2019103367-appb-000049
步骤A
Figure PCTCN2019103367-appb-000050
Figure PCTCN2019103367-appb-000051
向冰浴中,含有4-哌啶甲酸乙酯(50g,0.31mol,1eq),500ml THF,DIPEA(123g,0.95mol,3eq)的混合液中,滴加三光气(47.3g,0.16mol,0.5eq),保温1小时后,室温搅拌过夜,过滤,滤液直接用于下一步.
向一个含有水合肼(38g,0.95mol,3eq)的三口瓶中,在5℃的条件下,缓慢滴加上述的滤液,再搅3小时.加水淬灭,用乙酸乙酯萃取三次,旋于得化合物3b,36g，收率53％。MS(ESI)m/z:216.1(M+H+).1HNMR(400MHz,CDCl3)6.2(s,1H),4.1(m,2H),3.9(m,2H),3.1(m,2H),3.0(m,2H),2.5(m,1H),1.9(m,2H),1.7-1.5(m,2H),1.2(m,3H).
步骤B
Figure PCTCN2019103367-appb-000052
向含有化合物3b(34g,0.16mol),500ml THF,乙酸酐(32.3g,0.32mol,2eq)的混合液中,滴加POCl3(48.5g,0.32mol,2eq),搅拌2小时后,倒入冰水中,用乙酸乙酯萃取三次,旋于所得残余物过硅胶柱得化合物3c,20g，收率53％.MS(ESI)m/z:240.1(M+H+).1HNMR(400MHz,CDCl3)4.2(m,2H),3.9(m,2H),3.1(m,2H),2.5(m,1H),2.4(s,3H),2.0(m,2H),1.9(m,2H),1.2(t,3H).
步骤C
Figure PCTCN2019103367-appb-000053
向500ml单口瓶中加入化合物3c,20g,100ml THF,加入含水LiOH(3.5g,0.08mol,1eq)的50ml水溶液,搅拌过夜.加MTBE分液后,水相调pH大约4,析出白色固体,过滤,MTBE洗涤,烘干得化合物3d,5g，收率27％。MS(ESI)m/z:210(M-H+).1HNMR(400MHz,DMSO)3.8(m,2H),3.2(m,2H),2.5(m,1H),2.4(s,3H),1.9(m,2H),1.6(m,2H).
步骤D
Figure PCTCN2019103367-appb-000054
向100ml单口瓶中加入化合物3d(4.9g,1eq),无水30ml二氯甲烷,CDI(3.76g,1eq),和化合物2f(4.3g,1eq).在30℃搅拌过夜,旋干所得残余物过硅胶柱得白色固体.再用MTBE打浆,过滤,烘干得化合物3,3.6g，收率41％..MS(ESI)m/z:379(M+H+).1HNMR(400MHz,CDCl3)6.8(m,2H),6.7(m,1H),5.4(m,1H),4.3(m,1H),4.0(m,2H),3.9(m,1H),3.2(m,2H),3.0(m,1H),2.9(m,1H),2.4(s,3H),2.4-2.3(m,1H),2.1(m,1H),1.9(m,1H).
实施例4体外活性测试
(1)ADP-Glo活性测试.
联合应用Cisbio kinEASE STK试剂盒和Promega ADP-Glo激酶试剂盒检测化合物对RIP1抑制活性。用DMSO溶解待测化合物得到10mM储液。用DMSO进一步稀释化合物I至待测浓度的100倍。用激酶缓冲液(HEPES 250mM，NaN3 0.1％，BSA 0.05％，Orthovanadate 0.5mM，pH7.0)按照1:40的比例稀释待测化合物I。向384孔板中加入2μL稀释好的化合物。
用激酶缓冲液稀释人RIP1、STK底物S3和ATP至浓度分别为5ng/μL、10μM和250μM。然后向已经加入化合物I的348孔板中，依次加入稀释好的的1μL的人RIP1、1μL的STK底物S3和1μL的ATP。人RIP1、STK底物S3和ATP在反应孔的终浓度分别为1ng/μL、2μM和50μM。
化合物I、人RIP1、底物和ATP在室温共同孵育3小时.3小时后，加入5μL的Promega ADP-Glo试剂。中止反应并且去除剩余的ATP。中止反应后的混合物在室温孵育40分钟。
接着加入10μL的Promega ADP-Glo激酶检测试剂，使激酶反应产生的ADP转化成ATP。ATP进一步启动荧光素酶和荧光素的发光反应。反应30分钟后,用PerkinElmer EnVision Multilabel酶标仪检测发光信号。
通过全活性孔和背景信号孔计算出每个孔的抑制率，同时用专业的画图分析软件Prism 5.0对每个待测化合物进行半数抑制活性(IC50)的拟合，结果见表l。
(2)U937细胞测试
拯救TNF-alpha/Z-VAD-FMK诱导的坏死细胞的活力测定中使用人单核白血病U937细胞测试RIP1激酶抑制剂的体外活性。
使用含10％胎牛血清、100units/mL青霉素和100ug/mL链霉素的RMPI 1640培养基进行U937细胞培养。
用DMSO溶解待测化合物，得到10mM的待测化合物的母液。再用DMSO稀释至反应终浓度的100倍。用含有0.1％牛血清白蛋白的磷酸盐缓冲液溶解TNFα至100μg/mL，冻存备用。用DMSO溶解Z-VAD-FMK，得到50mM储液备用。
用培养基按照1:25的比例稀释待测化合物，用培养基稀释TNF-alpha和Z-VAD-FMK至400ng/ml和400uM。
取对数生长期的U937细胞，离心后用新鲜培养基重悬，按照每孔15000个细胞的密度铺人96孔细胞培养板，每孔50ul。同时加入稀释好的化合物的溶液25ul和TNF-alpha/Z-VAD-FMK 25ul。TNF-alpha和Z-VAD-FMK的终浓度分别为100ng/mL和100μM。
细胞置于37℃，5％CO 2培养箱中孵育24小时。24hr后，每孔加入100ul Cell Titer Glo，振荡后室温避光放置10min。用PerkinElmer EnVision酶标仪读取发光信号。
通过全活性孔和背景信号孔计算出每个孔的抑制率来测试化合物I拯救TNFα/QVD诱导的坏死细胞的效果，同时用专业的画图分析软件Prism 5.0对每个测试化合物I进行半数抑制活性(IC50)的拟合，结果见表l。
表l
Figure PCTCN2019103367-appb-000055
实施例5体内活性测试
TNF诱导的全身炎症反应综合征动物模型：
注射用鼠源mTNF-α和Z-VAD-FMK的配制：鼠源mTNF-α购买于南京金斯瑞生物公司和Z-VAD-FMK购买于Selleck公司，每次量取30ug mTNF溶于200ul endotoxin-free PBS和每0.25mg Z-VAD-FMK溶于5ul DMSO，再195ul endotoxin-free PBS稀释。
TNF诱导的全身炎症反应综合征小鼠模型准备：选取8-10周，雄性C57Bl/6J小鼠。剃毛(上腹部)，分组(每组5只)，利用温度计测量3次取平均值(欧姆龙红外线电子温度计MC-872)。
TNF诱导的全身炎症反应综合征模型建立过程：本实验分为四组：(1)空白对照组， 首先玉米油100μl(100ul/只小鼠，灌胃)20分钟后，PBS(200ul/只小鼠)尾静脉注射每30min测一次体温:(2)TNF诱导组，首先玉米油100μl(100ul/只小鼠)灌胃20分钟后，mTNF-α(30μg/只，200ul/只小鼠)尾静脉注射和Z-VAD-FMK(0.25mg/只，200ul/只)腹腔注射后每30min测一次体温；TNF化合物1治疗组，首先化合物1(10mg/kg)混合玉米油100μl(100ul/只小鼠)灌胃20分钟后，mTNF-α(30μg/只，200ul/只小鼠)尾静脉注射和Z-VAD-FMK(0.25mg/只，200ul/只)腹腔注射后每30min测一次体温；TNF+GSK-2982772(现有的RIP1抑制剂)治疗组，首先GSK-2982772(10mg/kg)混合玉米油100μl(100ul/只小鼠)灌胃20分钟后，mTNF-α(30μg/只，200ul/只小鼠)尾静脉注射和Z-VAD-FMK(0.25mg/只，200ul/只)腹腔注射后每30min测一次体温；所有上述小鼠体温下降7℃后，实验终止，结果如图1所示。
由图1中可以看出化合物I显著改善了TNF诱导的全身炎症反应综合征。
在本发明提及的所有文献都在本申请中引用作为参考，就如同每一篇文献被单独引用作为参考那样。此外应理解，在阅读了本发明的上述讲授内容之后，本领域技术人员可以对本发明作各种改动或修改，这些等价形式同样落于本申请所附权利要求书所限定的范围。

Claims (13)

  1. 一种杂环化合物、或其药学上可接受的盐、或其药学上可接受的溶剂合物，其特征在于，所述化合物如式I所示
    Figure PCTCN2019103367-appb-100001
    其中，R 1表示1-3个选自下组的取代基：H、卤素、-OH、取代或未取代的C1-C6烷基、取代或未取代的C2-C6烯基、取代或未取代的C2-C6炔基、取代或未取代的C3-C10环烷基、取代或未取代的C3-C10杂环烷基、取代或未取代的C6-C20芳基、取代或未取代的C3-C10杂芳基、C1-C4羧基、-CN、NO 2；
    R 2为取代或未取代的C6-C20芳基、取代或未取代的C3-C10杂芳基或取代或未取代的5-8元杂环；
    其中，所述取代是指一个或多个氢原子各自独立地被选自下组的取代基所取代：H、卤素、-CN、NO 2、-OH、-NRaRb、C1-C4烷基、C1-C4卤代烷基、C2-C6烯基、C2-C6炔基、C3-C6环烷基、苯基、苄基、C6-C10芳基、C3-C10杂芳基，
    其中Ra和Rb各自独立地为H、C1-C3烷基、C3-C6环烷基。
  2. 如权利要求1所述的杂环化合物、或其药学上可接受的盐、或其药学上可接受的溶剂合物，其特征在于，R 1选自下组：H、卤素、C1-C6烷基。
  3. 如权利要求1所述的杂环化合物、或其药学上可接受的盐、或其药学上可接受的溶剂合物，其特征在于，所述化合物如式II所示：
    Figure PCTCN2019103367-appb-100002
    其中，
    M、Q、V、Y及Z各自独立地选自下组：C、N、S、或O；
    R 3、R 4、R 5、R 6及R 7各自独立地选自下组：H、卤素、-CN、C1-C6烷基、C2-C6烯基、C2-C6炔基、C3-C10环烷基、C3-C10杂环烷基、C3-C10芳基、C3-C10杂芳基、-CN、NO 2；
    Figure PCTCN2019103367-appb-100003
    表示单键或双键。
  4. 如权利要求3所述的杂环化合物、或其药学上可接受的盐、或其药学上可接受的溶剂合物，其特征在于，V及Z为N，以及M、Q及Y为C。
  5. 如权利要求1所述的杂环化合物、或其药学上可接受的盐、或其药学上可接受的溶剂合物，其特征在于，所述式I化合物为
    Figure PCTCN2019103367-appb-100004
  6. 一种药物组合物，其特征在于，所述的药物组合物包括：
    (i)安全有效量的如权利要求1所述的杂环化合物、或其药学上可接受的盐、或其药学上可接受的溶剂合物；以及
    (ii)药学上可用的载体。
  7. 如权利要求1所述的杂环化合物、或其药学上可接受的盐、或其药学上可接受的溶剂合物的用途，其特征在于，(i)用于制备蛋白激酶抑制剂，(ii)用于制备治疗蛋白激酶相关的疾病的药物，(iii)用于制备治疗TNF相关疾病的药物，和/或(iv)用于制备治疗程序性细胞坏死相关疾病的药物。
  8. 一种如权利要求1所述杂环化合物或其药学上可接受的盐或其药学上可接受的溶剂合物的制备方法，其特征在于，所述方法包括步骤：
    (1)将式6化合物与式5化合物进行反应，从而得到式7化合物；
    Figure PCTCN2019103367-appb-100005
    (2)对所述式7化合物进行脱保护基团，从而得到式8化合物；以及
    Figure PCTCN2019103367-appb-100006
    (3)将所述式8化合物与R 2-X进行反应，从而得到式I化合物；
    Figure PCTCN2019103367-appb-100007
    其中，R 1、R 2的定义同权利要求1；
    R 10为保护基团；
    X为Cl或Br。
  9. 如权利要求8所述的制备方法，其特征在于，式5化合物的制备方法包括步骤：
    (1.1)将式1化合物与式2化合物进行反应，从而得到式3化合物
    Figure PCTCN2019103367-appb-100008
    (1.2)将式3化合物与甲基磺酰氯进行反应，从而得到式4化合物
    Figure PCTCN2019103367-appb-100009
    (1.3)在惰性溶剂中，在三氟乙酸存在下，使式4化合物进行反应，从而得到式5化合物
    Figure PCTCN2019103367-appb-100010
  10. 一种体外非治疗性抑制蛋白激酶的方法，其特征在于，所述方法包括步骤：将如权利要求1所述的杂环化合物、或其药学上可接受的盐、或其药学上可接受的溶剂合物与所述蛋白激酶接触从而抑制所述蛋白激酶的活性。
  11. 一种制备方法，其特征在于，所述方法包括步骤：
    (a)在惰性溶剂中，使式4化合物进行反应，从而得到式5化合物
    Figure PCTCN2019103367-appb-100011
    式中，R 1的定义同权利要求1。
  12. 如权利要求11所述的制备方法，其特征在于，步骤(a)在三氟乙酸存在下进行。
  13. 如权利要求11所述的制备方法，其特征在于，所述方法还包括步骤：
    (b1)将式9化合物与式5化合物进行反应，从而得到式I化合物；
    Figure PCTCN2019103367-appb-100012
    各式中，R 1、R 2的定义同权利要求1；
    或者，所述方法还包括步骤：
    (c1)将式6化合物与式5化合物进行反应，从而得到式7化合物；
    Figure PCTCN2019103367-appb-100013
    (c2)对所述式7化合物进行脱保护基团，从而得到式8化合物；以及
    Figure PCTCN2019103367-appb-100014
    (c3)将所述式8化合物与R 2-X进行反应，从而得到式I化合物；
    Figure PCTCN2019103367-appb-100015
    各式中，R 1、R 2的定义同权利要求1；R 10为保护基团；X为Cl或Br。
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WO2023040870A1 (zh) * 2021-09-16 2023-03-23 维泰瑞隆（北京）生物科技有限公司 Rip1激酶抑制剂及其用途
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