Norbornane derivative and pharmaceutical use thereof

Abstract

The present disclosure relates to a norbornene derivative and a pharmaceutical use thereof. Specifically, the present disclosure relates to a compound represented by formula I or a pharmaceutically acceptable salt thereof, and a use thereof in the preparation of a drug for treating cardiovascular and cerebrovascular diseases.

Description

norbornene derivatives and their pharmaceutical uses Technical Field

[0001] This disclosure pertains to the pharmaceutical field and relates to norbornene derivatives and their pharmaceutical uses. Background Technology

[0002] Stroke is a disease caused by a sudden interruption or severe reduction in blood supply to the brain. It can be divided into two main types: ischemic stroke and hemorrhagic stroke. Common types of ischemic stroke include embolic infarction and cerebral thrombosis, mainly caused by the blockage of arteries supplying blood to the brain, leading to cerebral ischemia and hypoxia. Common types of hemorrhagic stroke include cerebral hemorrhage and subarachnoid hemorrhage, mainly caused by the rupture of cerebral blood vessels leading to intracranial hemorrhage.

[0003] Borneol is a norbornene derivative that selectively activates α2-GABA. A The receptor has a clear anti-cerebral ischemia effect and a good neuroprotective effect in the cerebral ischemia-reperfusion model (Journal of Biomedical Research, 2017, 31(4): 306-314). In addition, it also has anti-inflammatory, analgesic, anticoagulant, anti-myocardial ischemia, tumor drug resistance reversal and arousal and cognitive function improvement effect in rats that have been working continuously for a long time.

[0004] As a lipid-soluble drug, dexborneol has a clear effect on promoting drug absorption and crossing the blood-brain barrier, and promoting drug distribution in the brain. However, it also has the disadvantages of poor water solubility and low oral bioavailability. Summary of the Invention

[0005] This disclosure provides compounds of the following formula or pharmaceutically acceptable salts thereof.

[0006] In some embodiments, the compound or its pharmaceutically acceptable salt is

[0007] On the other hand, this disclosure also provides a compound or a pharmaceutically acceptable salt thereof.

[0008] In some embodiments, the compound or its pharmaceutically acceptable salt is

[0009] This disclosure also provides isotopic substitutes of the aforementioned compounds or their pharmaceutically acceptable salts. In some embodiments, the isotopic substitutes are deuterated derivatives.

[0010] In some embodiments, the unit dose of the pharmaceutical composition is 0.001 mg to 1000 mg.

[0011] In certain embodiments, the pharmaceutical composition contains 0.01-99.99% of the aforementioned compound or its pharmaceutically acceptable salt or its isotopic substitution, based on the total weight of the composition. In certain embodiments, the pharmaceutical composition contains 0.1-99.9% of the aforementioned compound or its pharmaceutically acceptable salt or its isotopic substitution. In certain embodiments, the pharmaceutical composition contains 0.5%-99.5% of the aforementioned compound or its pharmaceutically acceptable salt or its isotopic substitution. In certain embodiments, the pharmaceutical composition contains 1%-99% of the aforementioned compound or its pharmaceutically acceptable salt or its isotopic substitution. In certain embodiments, the pharmaceutical composition contains 2%-98% of the aforementioned compound or its pharmaceutically acceptable salt or its isotopic substitution.

[0012] In some embodiments, the pharmaceutical composition contains 0.01% to 99.99% pharmaceutically acceptable excipients based on the total weight of the composition. In some embodiments, the pharmaceutical composition contains 0.1% to 99.9% pharmaceutically acceptable excipients. In some embodiments, the pharmaceutical composition contains 0.5% to 99.5% pharmaceutically acceptable excipients. In some embodiments, the pharmaceutical composition contains 1% to 99% pharmaceutically acceptable excipients. In some embodiments, the pharmaceutical composition contains 2% to 98% pharmaceutically acceptable excipients.

[0013] This disclosure also provides a method for treating cardiovascular and cerebrovascular diseases by administering to a patient a therapeutically effective amount of the aforementioned compound or its pharmaceutically acceptable salt or isotopic substitute, or the aforementioned pharmaceutical composition.

[0014] This disclosure also provides the use of the aforementioned compound or its pharmaceutically acceptable salt or the aforementioned pharmaceutical composition in the preparation of a medicament for the treatment of cardiovascular and cerebrovascular diseases.

[0015] The cardiovascular and cerebrovascular diseases described in this disclosure include, but are not limited to: ischemic cerebrovascular disease, cerebral infarction (stroke or cerebrovascular accident), myocardial infarction, and coronary heart disease.

[0016] The pharmaceutically acceptable salts of the compounds described in this disclosure may be selected from inorganic or organic salts. The inorganic salts are selected from hydrochloride, hydrobromide, phosphate, or sulfate; the organic salts are selected from acetate, trifluoroacetate, methanesulfonate, p-toluenesulfonate, citrate, maleate, tartrate, fumarate, citrate, or lactate.

[0017] The compounds of the present disclosure may exist in specific geometric or stereoisomeric forms. The present disclosure contemplates all such compounds, including cis and trans isomers, (-)- and (+)-enantiomers, (R)- and (S)-enantiomers, diastereomers, (D)-isomers, (L)-isomers, and racemic mixtures and other mixtures thereof, such as enantiomerically or diastereomerically enriched mixtures, all of which are within the scope of the present disclosure. Additional asymmetric carbon atoms may be present in substituents such as alkyl groups. All of these isomers and their mixtures are included within the scope of the present disclosure. The compounds of the present disclosure containing asymmetric carbon atoms can be isolated in optically pure form or in racemic form. Optically pure forms can be resolved from racemic mixtures or synthesized by using chiral starting materials or chiral reagents.

[0018] Optically active (R)- and (S)- isomers, as well as D- and L- isomers, can be prepared by chiral synthesis, chiral reagents, or other conventional techniques. To obtain an enantiomer of a compound disclosed herein, it can be prepared by asymmetric synthesis or derivatization with a chiral auxiliary, wherein the resulting diastereomeric mixture is separated and the auxiliary group is cleaved to provide the desired enantiomer in pure form. Alternatively, when the molecule contains a basic functional group (such as an amino group) or an acidic functional group (such as a carboxyl group), a salt of the diastereomeric isomer is formed with a suitable optically active acid or base, followed by diastereomeric resolution using conventional methods known in the art, and then the pure enantiomer is recovered. Furthermore, the separation of enantiomers and diastereomeric isomers is typically accomplished by using chromatography employing a chiral stationary phase and optionally combined with chemical derivatization (e.g., from amines to carbamates).

[0019] In the chemical structure of the compounds described in this disclosure, the bonds... This indicates that the configuration is not specified; that is, if chiral isomers exist in the chemical structure, the bond... It can be or Or simultaneously include and Two configurations.

[0020] In the chemical structure of the compounds described in this disclosure, the bonds... No configuration is specified, meaning it can be Z configuration, E configuration, or both configurations.

[0021] The compounds and intermediates disclosed herein may also exist in different tautomer forms, and all such forms are included within the scope of this disclosure. The terms "tautomer" or "tautomer form" refer to structural isomers of different energies that can interconvert via a low energy barrier. For example, proton tautomers (also known as proton transfer tautomers) include interconversion via proton transfer, such as the compounds of this disclosure comprising tautomer changes between formulas A and B as shown below.

[0022] All tautomers are within the scope of this disclosure. The nomenclature of compounds does not exclude any tautomers.

[0023] This disclosure also includes compounds identical to those described herein, but in which one or more atoms are labeled with isotopes whose atomic weights or mass numbers differ from those commonly found in nature. Examples of isotopes that can be incorporated into compounds of this disclosure include isotopes of hydrogen, carbon, nitrogen, oxygen, phosphorus, sulfur, fluorine, iodine, and chlorine, such as... 2 H, 3 H, 11 C 13 C 14 C 13 N、 15 N、 15 O、 17 O、 18 O、 31 P, 32 P, 35 S, 18 F, 123 I, 125 I and 36 Cl, etc.

[0024] Unless otherwise stated, when a position is specifically designated as deuterium (D), the position is understood to have at least 1000 times the abundance of deuterium greater than the natural abundance of deuterium (which is 0.015%) (i.e., at least 10% deuterium incorporation). In the example, the compound has a natural abundance greater than deuterium that can be at least 1000 times the abundance of deuterium, at least 2000 times the abundance of deuterium, at least 3000 times the abundance of deuterium, at least 4000 times the abundance of deuterium, at least 5000 times the abundance of deuterium, at least 6000 times the abundance of deuterium or more abundant deuterium. The disclosure also includes various deuterated forms of formula (I) compounds. Each available hydrogen atom connected to a carbon atom can be independently replaced by a deuterium atom. Those skilled in the art can synthesize deuterated forms of formula (I) compounds with reference to relevant literature. Commercially available deuterated starting materials may be used in the preparation of deuterated forms of the compounds of formula (I), or they may be synthesized using conventional techniques using deuterated reagents, including but not limited to deuterated borane, trideuterated borane in tetrahydrofuran, deuterated lithium aluminum hydride, deuterated iodoethane, deuterated iodomethane, and the like.

[0025] "Optional" or "optional" means that the event or situation subsequently described may, but does not have to, occur; the description includes the possibility or possibility that the event or situation may or may not occur. For example, "optionally halogenated or cyano-substituted C..." 1-6 "Alkyl" means that halogens or cyano groups may or may not be present. This description includes cases where alkyl groups are substituted by halogens or cyano groups and cases where alkyl groups are not substituted by halogens or cyano groups.

[0026] Terminology Explanation:

[0027] "Pharmaceutical composition" means a mixture containing one or more of the compounds described herein or their physiologically pharmaceutically acceptable salts or prodrugs, along with other chemical components, such as physiologically pharmaceutically acceptable carriers and excipients. The purpose of a pharmaceutical composition is to facilitate administration to a living organism, thereby promoting the absorption of the active ingredient and its biological activity.

[0028] "Pharmaceutical excipients" include, but are not limited to, any adjuvants, carriers, excipients, flow aids, sweeteners, diluents, preservatives, dyes / colorants, flavorings, surfactants, wetting agents, dispersants, suspending agents, stabilizers, isotonic agents, solvents, or emulsifiers that have been approved by the U.S. Food and Drug Administration for use in humans or livestock.

[0029] The term "effective amount" or "effective therapeutic amount" as used in this disclosure includes an amount sufficient to improve or prevent symptoms or conditions of a medical condition. An effective amount also means an amount sufficient to allow or facilitate diagnosis. The effective amount for a particular patient or veterinary subject may vary depending on factors such as the condition to be treated, the patient's overall health, the route and dosage of administration, and the severity of side effects. An effective amount may be the maximum dose or administration regimen that avoids significant side effects or toxicity.

[0030] The term "THF" refers to tetrahydrofuran.

[0031] The term "LiHMDS" refers to lithium di(trimethylsilyl)amino.

[0032] The term "DCM" refers to dichloromethane.

[0033] The term "MTBE" refers to methyl tert-butyl ether.

[0034] The term "DIPEA" refers to N,N-diisopropylethylamine.

[0035] The term "TFA" refers to trifluoroacetic acid.

[0036] The term "DMAP" refers to 4-dimethylaminopyridine.

[0037] The term "DMF" refers to N,N-dimethylformamide.

[0038] The term "DCC" refers to N,N'-dicyclohexylcarbodiimide.

[0039] The term "TMSOTf" refers to trimethylsilyl trifluoromethanesulfonate.

[0040] The term "Selectfluor" refers to a selective fluorine reagent.

[0041] The term "TBAF" refers to tetrabutylammonium fluoride.

[0042] The term "LDA" refers to lithium diisopropylaminodimethylamine.

[0043] The term "NaHMDS" refers to sodium bis(trimethylsilyl)amino. Attached Figure Description

[0044] Figure 1. Effects of the compound on neurological function scores in rats with permanent ischemic stroke (Mean ± SD, n = 6).

[0045] Figure 2. Effect of the compound on the infarct area in rats with permanent ischemic stroke (Mean ± SD, n = 6). Detailed Implementation

[0046] The present disclosure is further described below with reference to embodiments, but these embodiments are not intended to limit the scope of the present disclosure.

[0047] Experimental methods in the examples of this disclosure that do not specify specific conditions are generally based on conventional conditions or the conditions recommended by the raw material or product manufacturers. Reagents without specific sources are conventional reagents purchased from the market.

[0048] The structure of the compound was determined by nuclear magnetic resonance (NMR) and / or mass spectrometry (MS). NMR shifts (δ) were expressed in 10⁻¹⁰ ohms. -6 The unit (ppm) is given. NMR measurements were performed using a Bruker AVANCE-400 NMR spectrometer. The solvents used were deuterated dimethyl sulfoxide (DMSO-d6), deuterated chloroform (CDCl3), and deuterated methanol (Methanol-d4). The internal standard was tetramethylsilane (TMS).

[0049] HPLC determination was performed using an Agilent 1100 high-performance liquid chromatograph, a GAS15B DAD UV detector, and a Water Vbridge C18 150*4.6mm 5um column.

[0050] MS measurements were performed using an Agilent 6120 triple quadrupole mass spectrometer with a G1315D DAD detector and a Waters Xbridge C18 4.6*50mm, 5µm column, in positive / negative ion mode, with a mass scan range of 80–1200.

[0051] The silica gel plates used for thin-layer chromatography are Yantai Huanghai HSGF254 silica gel plates. The silica gel plates used in thin-layer chromatography (TLC) have a size of 0.2mm ± 0.03mm, and the size used for thin-layer chromatography separation and purification of products is 0.4mm-0.5mm.

[0052] Rapid column purification systems use either the Combiflash Rf150 (TELEDYNE ISCO) or Isolara One (Biotage).

[0053] Normal column chromatography generally uses Yantai Huanghai silica gel 200-300 mesh or 300-400 mesh as the carrier, or Changzhou Santai pre-filled ultrapure normal phase silica gel column (40-63μm, 60g, 24g, 40g, 120g or other specifications).

[0054] The known starting materials disclosed herein can be synthesized using or in accordance with methods known in the art, or can be purchased from companies such as Shanghai Titan Technology, ABCR GmbH & Co. KG, Acros Organics, Aldrich Chemical Company, Accela ChemBio Inc, and Bid Pharmaceuticals.

[0055] Unless otherwise specified in the examples, all reactions can be carried out under a nitrogen atmosphere.

[0056] A nitrogen atmosphere refers to a reaction flask connected to a nitrogen balloon with a volume of approximately 1L.

[0057] A hydrogen atmosphere refers to a reaction flask connected to a hydrogen balloon with a volume of approximately 1L.

[0058] Hydrogen was produced by the QPH-1L hydrogen generator from Shanghai Quanpu Scientific Instruments Co., Ltd.

[0059] Nitrogen or hydrogen atmospheres are typically evacuated and then filled with nitrogen or hydrogen gas, and this process is repeated three times.

[0060] Unless otherwise specified in the examples, "solution" refers to an aqueous solution.

[0061] Unless otherwise specified in the examples, the reaction temperature is room temperature, 20°C to 30°C.

[0062] The reaction progress in the examples was monitored by thin layer chromatography (TLC). The developing solvent used in the reaction, the column chromatography eluent system used to purify the compound, and the developing solvent system for thin layer chromatography, the volume ratio of the solvent were adjusted according to the polarity of the compound, and a small amount of alkaline or acidic reagents such as triethylamine and acetic acid could be added for adjustment.

[0063] Example 1: Preparation of Compound 1

[0064] Step 1:

[0065] Compound 1a (10.0 g, 65.69 mmol, purchased from Shaoyuan) was added to a flask, and 100 mL of toluene was added and stirred to dissolve. Under nitrogen protection, triethylamine (15.95 g, 157.65 mmol) and TMSOTf (29.20 g, 131.38 mmol) were added, and the mixture was heated to 125 °C and refluxed until the reaction was complete. At room temperature, 100 mL of n-hexane was added, and the mixture was washed twice with 50 mL of saturated sodium bicarbonate aqueous solution each time. The solution was dried over anhydrous sodium sulfate, filtered, and concentrated to give 13.7 g of compound 1b (yield: 92.94%).

[0066] 1H NMR (400MHz, CDCl3): δ4.45 (d, 1H, J = 3.6Hz), 1.99~2.00 (m, 1H), 1.51~1.58 (m, 1H), 1.26~1.32(m,1H),0.83~0.96(m,2H),0.68(s,6H),0.58(s,3H),0.00~0.01(s,9H).

[0067] Step 2:

[0068] Compound 1b (12.0 g, 53.47 mmol) was added to a flask and dissolved in 100 mL of anhydrous DMF. Under nitrogen protection, Selectfluor (18.94 g, 53.47 mmol) and a tetrahydrofuran solution of TBAF (1.0 M, 53.47 mL) were added at room temperature. Then, 200 mL of water was added and the mixture was stirred until the reaction was complete. The mixture was extracted three times with 100 mL of hexane each time. The organic phases were combined, dried over sodium sulfate, filtered, concentrated, and purified by column chromatography (PE:EA = 0-10%) to give 8.1 g of compound 1c (yield: 88.99%).

[0069] 1 H NMR (400MHz, CDCl3): δ4.36~4.96(m,1H), 1.25~2.30(m,4H), 0.84~1.12(m,10H).

[0070] Step 3:

[0071] Compound 1c (3.0 g, 17.62 mmol) was added to a flask, and 30 mL of anhydrous tetrahydrofuran was added and stirred to dissolve. The system was protected with nitrogen and cooled to -78 °C. LDA (1.0 M, 35.25 mL) was added dropwise, and the reaction was allowed to proceed until complete. The reaction was quenched with 30 mL of saturated ammonium chloride solution, and extracted three times with 30 mL of MTBE each time. The organic phases were combined, dried over sodium sulfate, filtered, concentrated, and purified by column chromatography (PE:EA = 50:1-20:1) to give 1.5 g of compound 1d (yield: 50%).

[0072] 1 H NMR (400MHz, CDCl3): δ4.83~4.97(m,1H), 2.40~2.42(m,1H), 1.74~1.88(m,3H), 1.26~1.58(m,1H), 0.84~1.12(m,9H).

[0073] Step 4:

[0074] Compound 1d (300 mg, 1.76 mmol) was added to a flask, and 10 mL of anhydrous tetrahydrofuran was added and stirred to dissolve. The system was protected under nitrogen and cooled to -78 °C. Lithium aluminum hydride (1.0 M, 2.29 mL) was added dropwise, and the reaction was allowed to proceed until complete. The mixture was quenched with 20 mL of saturated ammonium chloride solution, extracted three times with 30 mL of MTBE each time, and the organic phases were combined, dried over sodium sulfate, filtered, concentrated, and purified by column chromatography (PE:EA = 50:1-20:1) to give 120 mg of compound 1 (yield: 40%).

[0075] 1 H NMR (400MHz, CDCl3): δ4.92~5.10(m,1H), 3.83~3.89(m,1H), 1.98~2.06(m,2H) ), 1.68~1.82(m,2H), 1.49~1.54(m,1H), 1.23~1.30(m,1H), 0.84~0.93(m,9H).

[0076] Example 2 Preparation of Compound 2

[0077] Compound 1d (620 mg, 3.64 mmol) was added to a flask, dissolved in 10 mL of anhydrous isopropanol by stirring, and then aluminum isopropoxide (1.86 g, 9.11 mmol) was added. The system was protected under nitrogen atmosphere, and the reaction was carried out at 90 °C overnight. The mixture was quenched by adding 20 mL of protective sodium bicarbonate aqueous solution, extracted three times with 20 mL of MTBE each time, dried over sodium sulfate, filtered, concentrated, and purified by column chromatography (PE:EA = 50:1) to give 410 mg of compound 2 (yield: 65.3%).

[0078] 1 H NMR (400MHz, CDCl3): δ4.99~5.15(m,1H), 3.55~3.63(m,1H), 2.00(t,1H), 1.89 (d,1H), 1.47~1.74(m,3H), 1.14~1.20(m,1H), 1.02(s,3H), 0.84~0.88(m,6H).

[0079] Example 3 Preparation of Compound 3

[0080] Step 1:

[0081] Compound 1c (4.45 g, 26.14 mmol) was added to a flask, and 90 mL of anhydrous tetrahydrofuran was added and stirred to dissolve. Under nitrogen protection, (R)-methyl-CBS-oxazolium borane (1.45 g, 5.23 mmol, purchased from B&D) was added. The mixture was cooled to 0 °C and a tetrahydrofuran solution of borane (1.0 M, 52.3 mL) was added. The reaction mixture was stirred until complete. 1 M dilute hydrochloric acid was added to the reaction solution to pH 3. The organic solvent was removed by concentration. The aqueous phase was extracted three times with 50 mL of methyl tert-butyl ether. The organic phases were combined and washed successively with 50 mL of water and 50 mL of saturated brine. The organic phase was dried over sodium sulfate, filtered, concentrated, and purified by column chromatography (EA:PE = 0-5%) to give 170 mg of compound 3 (yield: 4%).

[0082] 1 H NMR (400MHz, CDCl3): δ4.40-4.22(m,1H),4.20-4.08(m,1H),2.00-1.88(m,1H),1.84-1.65(m,2H) ,1.30-1.24(m,1H),1.22-1.18(m,1H),1.16-1.05(m,1H),1.00(s,3H),0.91(s,3H),0.87(s,3H).

[0083] Example 4: Preparation of Compound 4

[0084] Step 1:

[0085] Compound 1c (2.0 g, 11.75 mmol) was added to a flask, and 20 mL of anhydrous tetrahydrofuran was added and stirred to dissolve. The system was protected with nitrogen and cooled to -60 °C. A tetrahydrofuran solution of tri-sec-butylborohydride (1.0 M, 15.3 mL) was added, and the reaction was allowed to proceed to completion. The reaction was quenched by adding 20 mL of 1 M hydrochloric acid aqueous solution. The mixture was extracted three times with 50 mL of MTBE each time. The organic phases were combined, dried over sodium sulfate, filtered, concentrated, and purified by column chromatography (PE:MTBE = 100:15) to give 633 mg of compound 4 (yield: 30%).

[0086] 1 H NMR (400MHz, CDCl3): δ4.70-4.54(m,1H),3.69-3.63(m,1H),2.04-1.99(m,2H),1.73 -1.69(m,1H),1.50-1.44(m,1H),1.10-1.08(m,3H),0.97-0.91(m,5H),0.83(s,3H).

[0087] Example 5: Preparation of compounds 5 and 6

[0088] Step 1:

[0089] Compound 1a (2.0 g, 13.14 mmol) was added to a flask, followed by the addition of 20 mL of tetrahydrofuran. The mixture was stirred and dissolved under nitrogen protection. The mixture was cooled to -78 °C, and 22.99 mL of NaHMDS (1.0 M) was added. After the addition was complete, the mixture was heated to 0 °C and reacted until complete. The mixture was then cooled to -78 °C again, and 100 mL of a tetrahydrofuran solution of N-fluorobis(benzenesulfonamide) (0.53 M) was added dropwise. After the addition was complete, the mixture was reacted at room temperature until complete. The reaction was quenched with 100 mL of 1N HCl, extracted with 100 mL of ethyl acetate, washed once with 50 mL of saturated sodium bicarbonate, dried over sodium sulfate on the organic phase, filtered, concentrated, and slurried with 200 mL of n-hexane. The mixture was filtered, concentrated, and purified by column chromatography (PE:EA = 50:1-30:1) to give 1.3 g of compound 5a (yield: 53%).

[0090] 1 H NMR (400MHz, CDCl3): δ3.87~3.94(m,1H), 2.04~2.07(m,1H), 1.89~1.94(m,1H), 1.67~1.77(m,3H), 1.20~1.30(m,1H), 0.89~1.00(m,9H).

[0091] Step 2:

[0092] Compound 5a (1.08 g, 5.76 mmol) was added to a flask, and 15 mL of anhydrous isopropanol was added and stirred to dissolve. Aluminum isopropoxide (2.94 g, 14.39 mmol) was added, and the system was protected under nitrogen atmosphere. The reaction was heated until complete. The mixture was quenched with 30 mL of protective sodium bicarbonate aqueous solution, extracted with MTBE (20 mL × 3), dried over sodium sulfate, filtered, concentrated, and purified by column chromatography (PE:EA = 150:1-100-1) to give 38 mg of compound 5 (3% yield) and 560 mg of compound 6 (51% yield).

[0093] Compound 5: 1 H NMR (400MHz, CDCl3): δ3.88~3.94(m,1H), 2.04~2.07(m,1H), 1.90~1.93(m,1H), 1.62~1.75(m,3H), 1.24~1.30(m,1H), 0.90-1.00(m,9H).

[0094] Compound 6: 1H NMR (400MHz, CDCl3): δ3.60~3.64(m,1H), 2.01~2.04(m,1H), 1.96(br s,1H), 1.56~1.66(m,3H), 1.24~1.27(m,1H), 1.12-1.13(d,3H), 0.94(s,3H), 0.88(s,3H).

[0095] Example 6 Preparation of Compound 7

[0096] Step 1:

[0097] Compound 1a (300 mg, 1.97 mmol) was added to a flask, and 6 mL of anhydrous tetrahydrofuran was added and stirred to dissolve. The system was protected under nitrogen atmosphere, and the temperature was lowered to below -60°C with dry ice and ethanol. A tetrahydrofuran solution of LDA (2.0 M, 1.2 mL) was added, and the reaction was stirred until complete. Methyl chlorosulfonate (309 mg, 2.36 mmol) was added, and the reaction was stirred until complete. 12 mL of 1 M dilute hydrochloric acid was added to the reaction solution, and the mixture was concentrated. The aqueous phase was extracted with petroleum ether (6 mL × 3). The organic phases were combined and washed successively with 3 mL of water, 3 mL of saturated sodium bicarbonate aqueous solution, and 3 mL of saturated brine. The organic phase was dried over sodium sulfate, filtered, concentrated, and purified by column chromatography (EA:PE = 0-5%) to give 175 mg of compound 7a (yield: 48%).

[0098] Step 2:

[0099] Compound 7a (100 mg, 0.54 mmol) was added to a flask, and 2 mL of anhydrous tetrahydrofuran was added and stirred to dissolve. The system was protected with nitrogen and cooled to -50 °C. A tetrahydrofuran solution of tri-sec-butylborohydride (1.0 M, 3.9 mL) was added dropwise until the reaction was complete. The reaction was quenched with 10 mL of 1 M hydrochloric acid aqueous solution, extracted with MTBE (10 mL × 2), the organic phases were combined, dried over sodium sulfate, filtered, concentrated, and purified by column chromatography (EA:PE = 0-5%) to give 92 mg of compound 7 (yield: 91%).

[0100] 1 H NMR (400MHz, CDCl3): δ4.70~4.66(m,1H), 3.89~3.85(m,1H), 2.06~2.05(m,1H) ), 1.97~1.95(m,1H), 1.85~1.77(m,2H), 1.37~1.20(m,2H), 0.95~0.90(m,9H).

[0101] Biological evaluation

[0102] The present disclosure is further described and explained below with reference to test examples, but these embodiments are not intended to limit the scope of the present disclosure.

[0103] Test Example 1: Neuroprotective effect of compound 1 on SD rats with cerebral ischemia-reperfusion injury

[0104] This study used a middle cerebral artery occlusion (MCAO) model in SD rats to evaluate the neuroprotective effect of the test compound on SD rats with cerebral ischemia-reperfusion injury by analyzing the infarct area and neurological function scores after intravenous injection of the test compound.

[0105] 1. Experimental materials

[0106] 1.1 Main Instruments

[0107] 1.2 Laboratory Animals

[0108] Male SD rats, weighing 240–260g.

[0109] 2. Experimental Methods

[0110] 2.1 Experimental Procedure

[0111] Step 1): Model preparation: The method for preparing the SD rat middle cerebral artery occlusion (MCAO) model was based on the 4th edition of "Pharmacological Experimental Methodology". Adult male SD rats underwent cerebral ischemia-reperfusion for 90 min using the suture method.

[0112] Step 2): Animal grouping: 60 minutes after the silk thread is inserted, based on the neurological function score (10-11 points), select successfully modeled rats and randomly group them into groups of 8.

[0113] Step 3): Administration method: A single tail vein bolus injection was administered immediately after reperfusion. The endpoint of the experiment was 24 hours after the end of administration.

[0114] Step 4): Detection indicators:

[0115] Neurological function assessment: The degree of neurological function impairment in animals was assessed using a blind method before and 24 hours after drug administration, with a total score of 16 points.

[0116] Infarct area measurement: 24 hours after administration, the whole brain was decapitated and the brain tissue sections were stained in 1% red tetrazolium (TTC) solution to calculate the infarct area.

[0117] 2.2 Data Analysis

[0118] Experimental data are expressed as mean ± standard deviation (mean ± SD), and data analysis was performed using IBM SPSS Statistics 25.0 software. One-way ANOVA was used for comparisons among multiple groups. First, a homogeneity of variance test was performed. If P ≤ 0.05, the data were considered to have unequal variances; if P > 0.05, the data were considered to have homogeneous variances. Multiple comparisons were performed using the LSD method to determine homogeneity of variances. If the variances were unequal, Dunnett's T3 test was used for statistical analysis, and P < 0.05 was considered statistically significant.

[0119] Test Example 2: Anti-inflammatory effect of compound on neuroinflammatory cell model

[0120] This study used mouse microglia (BV-2 cells) to construct an inflammatory cell model by combined stimulation with INF-γ and TNF-α. The in vitro anti-inflammatory effects of the compounds were evaluated by analyzing their inhibitory effects on the transcriptional levels of IL-1β and IL-6 mRNA.

[0121] 1. Experimental materials

[0122] 1.1 Main Instruments

[0123] 1.2 Experimental Cells

[0124] BV-2 (mouse microglia).

[0125] 2. Experimental methods

[0126] 2.1 Experimental Procedure

[0127] Step 1): Cell Culture: Remove the old culture medium and wash once with PBS. Then add 0.25% trypsin and incubate at 37°C for 2-3 minutes. Add the appropriate culture medium to terminate digestion. Gently pipette the cell suspension to separate the aggregated cells. Transfer the cell suspension to a sterile centrifuge tube and centrifuge at 1000 rpm for 5 minutes to collect the cells. After centrifugation, discard the supernatant and seed the cells into 10 cm cell culture dishes, with a seeding density of 2.5 × 10⁶ cells per dish. 5 Each cell.

[0128] Step 2): Establishment of cell inflammation model: After cell plating, cell growth was observed under a microscope until it reached 70-80%. Different concentrations of compounds were added. After 3 hours, TNF-α and INF-γ solutions were added to a final concentration of 10 ng / mL to construct a TNF-α and INF-γ-induced cell inflammation model. Relevant tests were performed 24 hours later.

[0129] Step 3): Detection index: Lyse cells, extract RNA for subsequent experiments or store at -80 °C. Obtain cDNA template by reverse transcription, and detect the intracellular transcriptional levels of IL-1β and IL-6 mRNA using qPCR.

[0130] 2.2 Data analysis

[0131] After summarizing and statistically analyzing the data, use SPSS statistical software for analysis. According to the SPSS analysis results, use Graph Pad software to draw images.

[0132] Test Example 3 Pharmacokinetics Test in Beagle Dogs

[0133] 1. Sample preparation

[0134] Control preparation: Weigh the required amount of (+)-camphene (purchased from Jiangxi Xinsen Natural Vegetable Oil Co., Ltd.), add an appropriate volume of 5% DMSO + 95% normal saline, and use vortex or ultrasonic to dissolve it completely to obtain a clear administration solution. After preparation, filter it through a 0.22 μm microporous membrane for intravenous injection.

[0135] Test preparation: Weigh the required amount of Compound 1, add an appropriate volume of normal saline, and use vortex or ultrasonic to dissolve it completely to obtain a clear administration solution. After preparation, filter it through a 0.22 μm microporous membrane for intravenous injection.

[0136] 2. Test animals

[0137] Beagle dogs, male, weighing about 8 - 10 kg. The animals were purchased from Jiangsu Mas Biotechnology Co., Ltd., and the animal production license: SCXK (Su) 2021 - 0012.

[0138] 3. Test method

[0139] Administration method: Weigh before administration, calculate the dosage according to the weight, and administer by intravenous injection. Before administration and at 2 min, 5 min, 0.25 h, 0.5 h, 1 h, 2 h, 4 h, 6 h, and 8 h after administration, collect 0.6 - 1 mL of blood from the limb veins, place it in an EDTA-K2 anticoagulant test tube (pre-cooled in advance), and centrifuge within 30 min on wet ice (3500 rpm, 10 min, 2 - 8 °C). Separate the plasma and transfer it to a -80 °C refrigerator for freezing and storage. Pay attention to low-temperature operation during the whole process from collection to sub-packaging. Detect the concentrations of (+)-camphene and Compound 1 in the plasma respectively.

[0140] Table 1 Administration plan * Fast for more than 12 h during the test, drink water freely, and provide food 4 h after administration.

[0141] 4. Test results

[0142] The pharmacokinetic parameters of each group of animals are shown in Table 2.

[0143] Table 2 Pharmacokinetic data

[0144] Pharmacodynamic test in SD rats in Test Example 4

[0145] 1. Sample preparation

[0146] Weigh the required amount of Compound 1, Compound 5 or Compound 7, add an appropriate volume of normal saline, and use vortex or ultrasonic treatment to fully dissolve it to obtain a clear administration solution. After preparation, filter it through a 0.22 μm microporous filter membrane for intravenous injection.

[0147] 2. Test animals

[0148] SD rats, male, weighing about 240 - 260 g. The animals were purchased from Beijing Vital River Laboratory Animal Technology Co., Ltd., and the animal production license: SCXK (Beijing) 2021 - 0011.

[0149] 3. Test method

[0150] 3.1 Test steps:

[0151] (1) Model preparation: The preparation method of the middle cerebral artery occlusion model (MCAO) in SD rats was referred to the 4th edition of "Pharmacological Experiment Methodology", and a permanent cerebral ischemia model was prepared by the silk thread embolization method.

[0152] (2) Animal grouping: Neurological function scoring was performed 55 minutes after embolization (scores of 10 - 11 with relatively uniform injury degree, and those with excessive or mild injury were excluded), and laser speckle flow imaging was used to detect that the cerebral blood flow decreased by ≥50% compared with the ipsilateral baseline value. Rats meeting both criteria could be included in the group. The successfully modeled rats were randomly divided into 4 groups with 6 rats in each group, and another 6 rats were set as the sham operation group. The specific grouping information is as follows:

[0153] Table 3 Grouping and administration information table

[0154] (3) Administration method: 60 minutes after embolization, Compound 1, Compound 5 or Compound 7 solution was injected into the tail vein once respectively, and the sham operation and model control groups were injected with the vehicle normal saline once via the tail vein. The test endpoint was 24 hours after cerebral embolization.

[0155] (The) detection indexes:

[0156] Neurological function scoring: The degree of neurological impairment of animals was scored blindly before modeling, 55 minutes after ischemia, and 24 hours after modeling, with a total score of 16 points;

[0157] Infarct area measurement: 24 hours after administration, the whole brain was decapitated and the brain tissue sections were stained in 1% red tetrazolium (TTC) solution to calculate the percentage of cerebral infarction area.

[0158] 3.2 Data Analysis

[0159] Experimental data are expressed as mean ± standard deviation (mean ± SD), and data analysis was performed using IBM SPSS Statistics 25.0 software. One-way ANOVA was used for comparisons among multiple groups. First, a homogeneity of variance test was performed. If P ≤ 0.05, the data were considered to have unequal variances; if P > 0.05, the data were considered to have homogeneous variances. Multiple comparisons were performed using the LSD method to determine homogeneity of variances. If the variances were unequal, Dunnett's T3 test was used for statistical analysis, and P < 0.05 was considered statistically significant.

[0160] 4. Test Results

[0161] The neurological function scores of rats in each group are shown in Figure 1 and Table 4. Compared with the sham-operated group, rats in the model control group showed significant behavioral impairment 24 hours after modeling, with a score of 9.0 (P < 0.001). Compared with the model control group, administration of 0.4 mg / kg compound 1 1 hour after embolization significantly improved neurological function scores, with an improvement rate of 24.4% (P < 0.01). Compounds 0.4 mg / kg and 0.4 mg / kg compound 7 showed a trend of improvement in neurological function scores, with improvement rates of 11.1% (P > 0.05) and 13.0% (P > 0.05), respectively.

[0162] Table 4. Effects of the compounds on neurological function scores in rats with permanent ischemic stroke (Mean ± SD, n = 6) Note 1. Compared with the sham surgery group: ### P<0.001; 2. Compared with the model control group: ** P<0.01

[0163] The percentage of infarct area in each group of rats is shown in Figure 2 and Table 5. Compared with the sham-operated group, the rats in the model control group showed significant cerebral infarction, with an average infarct area of ​​25.88 mm (P < 0.001). Compared with the model control group, administration of 0.4 mg / kg compound 1 1 h after embolization significantly improved the cerebral infarct area, with an improvement rate of 33.0% (P < 0.01). Compounds 0.4 mg / kg and 0.4 mg / kg of compound 5 and 0.4 mg / kg of compound 7 showed a trend of improvement in cerebral infarct area, with improvement rates of 8.5% (P > 0.05) and 9.6% (P > 0.05), respectively. Compared with equivalent doses of compound 5 or compound 7, 0.4 mg / kg compound 1 significantly improved the cerebral infarct area (33.0% vs 8.5% vs 9.6%, P < 0.05).

[0164] Table 5. Effects of compounds on infarct area in rats with permanent ischemic stroke (Mean ± SD, n = 6) Note 1. Compared with the sham surgery group: ### P<0.001; 2. Compared with the model control group: ** P<0.01; 3. Compared with compound 5 or compound 7: $ P<0.05

Claims

1. A compound or a medicinally usable salt thereof, 2. A compound or a pharmaceutically acceptable salt thereof, 3. A compound or a medicinally acceptable salt thereof, 4. A compound or a medicinally acceptable salt thereof, 5. The compound according to any one of claims 1 to 4, or an isotope-substituted product of the pharmaceutically acceptable salt thereof, preferably, the isotope-substituted product is a deuterated product.

6. A pharmaceutical composition comprising at least one therapeutically effective amount of the compound of any one of claims 1 to 4 or a pharmaceutically acceptable salt thereof, or an isotope substitute as described in claim 5, and a pharmaceutically acceptable excipient.

7. A method for treating cardiovascular and cerebrovascular diseases, comprising administering to the patient a therapeutically effective amount of a compound as claimed in any one of claims 1 to 4 or a pharmaceutically acceptable salt thereof, or an isotope substitute as claimed in claim 5, or a pharmaceutical composition as claimed in claim 6.

8. The method according to claim 7, characterized in that, The cardiovascular and cerebrovascular diseases mentioned are selected from ischemic cerebrovascular disease, cerebral infarction, myocardial infarction, and coronary heart disease.

9. Use of the compound of any one of claims 1 to 4 or a pharmaceutically acceptable salt thereof, or an isotope substitute as described in claim 5, or a pharmaceutical composition as described in claim 6, in the preparation of a medicament for treating cardiovascular and cerebrovascular diseases.

10. The use according to claim 9, characterized in that, The cardiovascular and cerebrovascular diseases mentioned are selected from ischemic cerebrovascular diseases, cerebral infarction, myocardial infarction, and coronary heart disease.

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