Fasudil derivative compound and application thereof

Abstract

The invention discloses a fasudil derivative compound and a pharmaceutically acceptable salt of the fasudil derivative compound. The fasudil derivative compound disclosed by the invention can play a therapeutic role in protecting cerebral ischemia injury, has an important potential therapeutic value, and can be used for preparing medicines for treating cerebral apoplexy diseases.

Description

Technical Field

[0001] The present invention belongs to the field of pharmaceuticals, and provides a class of fasudil-derived compounds, their preparation methods and pharmaceutical uses. This class of compounds can play a therapeutic role in protecting against cerebral ischemic injury and can be used to prepare drugs for neuroprotection and the treatment of stroke diseases. Background Art

[0002] Stroke, also known as apoplexy, is the second most common cause of death globally. Stroke has brought a heavy economic and mental burden to society and families, and this burden will increase significantly in the next 20 years with the aging of the population. According to the pathological characteristics of stroke, stroke is divided into ischemic stroke and hemorrhagic stroke. Hemorrhagic stroke is mainly divided into intracerebral hemorrhage and subarachnoid hemorrhage due to different bleeding locations. Ischemic stroke is mainly caused by arterial occlusion and accounts for 71% of the total number of strokes.

[0003] Cerebral artery occlusion leads to severe oxygen and glucose deprivation (OGD), triggering a series of cellular and molecular events, which results in irreversible brain damage. Neurons are more vulnerable to hypoxia and rapidly malfunction or die after ischemic stroke. After ischemia, OGD causes mitochondrial dysfunction, leading to ATP depletion and the generation of a large amount of reactive oxidative species (ROS). Compared with other brain cells, neurons have a higher energy demand but insufficient energy reserves. After ATP depletion, a local ischemic cascade reaction is triggered, including ion pump failure, the influx of sodium ions, chloride ions and water molecules, the efflux of intracellular potassium ions and membrane depolarization. Neurons are unable to maintain their normal transmembrane ion gradients and homeostasis after ischemic stroke, thereby triggering a variety of pathological processes, including excitotoxicity, mitochondrial dysfunction, oxidative and nitrative stress, acidosis, Ca2+ overload, protein misfolding, inflammation, DNA damage, apoptosis, etc. These pathophysiological processes have harmful effects on neurons, glial cells and endothelial cells and activate each other through positive feedback loops, thus leading to nerve cell death.

[0004] According to the pathological characteristics of ischemic stroke, the most effective treatment method for ischemic stroke is to restore the blood flow of the cerebral artery blocked by thrombus or embolism. At present, the clinical treatment strategies for ischemic stroke mainly focus on two aspects: one is to improve blood supply, and through treatment strategies such as thrombolysis, thrombectomy, anticoagulation, antiplatelet aggregation, vasodilation, vascular remodeling, establishment of collateral circulation and regulation of blood state, restore and promote blood supply to the cerebral ischemic area; the other is to protect the structure and function of nerve cells, and through drugs to block the cascade reaction of nerve cell death, reduce nerve cell damage caused by ischemia, and improve the functional recovery of damaged nerve cells. However, in actual clinical applications, the drugs developed around the above two treatment strategies are very limited.

[0005] A class of fasudil-derived compounds of the present invention can play a good neuroprotective role and have broad medicinal prospects. This class of drugs can play a therapeutic role in protecting against cerebral ischemic injury. Summary of the Invention

[0006] Technical problems to be solved: The present invention provides a class of fasudil-derived compounds and their preparation methods. The most prominent feature of this class of compounds is their protective effect on cerebral ischemia-reperfusion injury, and they can be used to prepare drugs for treating stroke.

[0007] Technical solution: A class of fasudil-derived compounds shown in Formula I or their pharmaceutically acceptable salts, hydrates or solvates:

[0008]

[0009] Wherein, R1 is selected from C1-C4 alkyl or C1-C4 haloalkyl; R2 is selected from hydrogen or C1-C2 alkyl.

[0010] Preferably, R1 is selected from C1-C3 alkyl or C1-C3 haloalkyl;

[0011] R2 is selected from hydrogen, methyl or ethyl.

[0012] Preferably, R1 is selected from methyl, ethyl, isopropyl or dichloromethyl; R2 is selected from methyl.

[0013] More preferably, the compound is:

[0014]

[0015] Compound 1: As shown in S1;

[0016]

[0017] Compound 2: As shown in S2.

[0018] The compounds provided by the present invention also include pharmaceutically acceptable equivalents of the compounds or mixtures of two or more thereof.

[0019] Preferably, the compounds provided by the present invention may include one or a mixture of two or more of pharmaceutically acceptable salts, hydrates, and solvates.

[0020] Preferably, the compounds provided by the present invention include acid salts or base salts of the compounds provided by the present invention. The pharmaceutically acceptable salts have the pharmaceutical activity of the compounds and meet the requirements both biologically and in practical applications.

[0021] The present invention provides a fasudil-derived compound or a pharmaceutically acceptable salt thereof for treating stroke.

[0022] Beneficial effects:

[0023] A class of fasudil-derived compounds described in the present application, the most prominent feature of this class of drugs is that they have a protective effect on cerebral ischemia-reperfusion injury and can be used to prepare drugs for treating stroke. Detailed implementation manners

[0024] The present invention discloses fasudil-derived compounds and their uses. Those skilled in the art can draw on the content of this article and appropriately improve process parameters to achieve them. The methods and applications of the present invention have been described through preferred embodiments. Relevant personnel can obviously make changes or appropriate modifications and combinations to the methods and applications described in this article without departing from the content, spirit, and scope of the present invention to implement and apply the technology of the present invention.

[0025] In order to enable those skilled in the art to better understand the technical solutions of the present invention, the present invention will be further described in detail below with reference to specific embodiments.

[0026] Example 1: Synthesis of compound S1

[0027]

[0028] Synthesis route:

[0029]

[0030] Synthesis process:

[0031] The first step:

[0032] The raw material fasudil hydrochloride (1 g, 3.06 mmol) was dissolved in DCM (100 ml), 4A molecular sieve (6 g) was added, and the mixture was stirred at room temperature for 5 min. Then N-methylmorpholine (0.8 g, 7.95 mmol) was added, the temperature of the system was lowered to 0 - 5 °C, and chloroethyl chloroformate (0.66 g, 4.59 mmol) was added dropwise. After the addition, the reaction was carried out at 5 °C for 5 h. After the reaction, it was diluted with DCM (100 ml), the molecular sieve was filtered out, the organic phase was washed with water 3 times, dried and concentrated. The crude product was directly used for the next step.

[0033] Step 2:

[0034] The raw material isobutyric acid (0.11 g, 1.25 mmol) was dissolved in DCM (3 ml), TEA (0.134 g, 1.328 mmol) was added, and the mixture was stirred for 5 min. The crude product from the previous step (0.33 g) was added, and finally tetrabutylammonium iodide (0.15 g, 0.42 mmol) was added. The reaction was carried out at room temperature for 24 h. After completion, the system was diluted with DCM (60 ml), washed with water 3 times, dried and concentrated. It was purified by silica gel column chromatography with a PE:EA system to obtain 0.2 g of pure product (transparent oil).

[0035] ESI-MS: 450.1 [M+H] +

[0036] 1 H NMR (400 MHz, CDCl3) δ 9.38 (s, 1H), 8.73 (d, J = 5.7 Hz, 1H), 8.45–8.19 (m, 3H), 7.72 (t, J = 7.4 Hz, 1H), 6.77 (d, J = 2.9 Hz, 1H), 3.83–3.24 (m, 8H), 2.59–2.45 (m, 1H), 2.10–1.95 (m, 2H), 1.49 (d, J = 3.1 Hz, 3H), 1.12 (dt, J = 20.8, 10.3 Hz, 6H).

[0037] Example 2: Synthesis of Compound S2

[0038]

[0039] Synthesis route:

[0040]

[0041] Synthesis process:

[0042] Step 1:

[0043] The raw material fasudil hydrochloride (1 g, 3.06 mmol) was dissolved in DCM (100 ml), 4A molecular sieve (6 g) was added, and the mixture was stirred at room temperature for 5 min. Then N-methylmorpholine (0.8 g, 7.95 mmol) was added, the temperature of the system was lowered to 0 - 5 °C, chloroethyl chloroformate (0.66 g, 4.59 mmol) was added dropwise, and after addition, the reaction was carried out at 5 °C for 5 h. After the reaction, it was diluted with DCM (100 ml), the molecular sieve was filtered out, the organic phase was washed with water 3 times, dried and concentrated. The crude product was directly used for the next step.

[0044] Step 2:

[0045] The raw material dichloroacetic acid (0.24 g, 1.88 mmol) was dissolved in DMF (3 ml), sodium bicarbonate (0.168 g, 2 mmol) was added, and the mixture was stirred for 5 min. The crude product from the previous step (0.5 g) and 18-crown-6 (0.17 g, 0.63 mmol) were added, and finally potassium iodide (0.1 g, 0.63 mmol) was added, and the reaction was carried out at room temperature for 24 h. After completion, the system was diluted with EA (80 ml), washed with water 3 times, dried and concentrated. It was purified by silica gel column chromatography with a PE:EA system to obtain 0.3 g of pure product (pale yellow oil).

[0046] ESI-MS: 490.0 [M+H] +

[0047] 1 H NMR (400 MHz, CDCl3) δ 9.38 (s, 1H), 8.73 (d, J = 5.7 Hz, 1H), 8.45–8.19 (m, 3H), 7.72 (t, J = 7.4 Hz, 1H), 6.77 (d, J = 2.9 Hz, 1H), 6.3 (s, 1H), 3.83–3.24 (m, 8H), 2.10–1.95 (m, 2H), 1.49 (d, J = 3.1 Hz, 3H).

[0048] Example 3: Effect of S1 on focal cerebral ischemia-reperfusion injury

[0049] 1 Materials and methods

[0050] 1.1 Experimental animals

[0051] Sprague-Dawley (SD) rats, male, body weight: 250 - 280 g, SPF grade

[0052] 1.2 Tested drugs

[0053] Compound S1, dissolved in 2% propylene glycol. Edaravone injection, specification: 5 mL:10 mg, produced by Nanjing Simcere Dongyuan Pharmaceutical Co., Ltd., diluted with normal saline before use.

[0054] 1.3 Experimental methods

[0055] 1.3.1 Preparation of focal cerebral ischemia-reperfusion model. Main steps: Rats were anesthetized by intraperitoneal injection of 10% chloral hydrate (350 mg / kg). The right external carotid artery was isolated, ligated and transected. A nylon suture with a swollen head was slowly inserted about 18 mm along the common carotid artery and internal carotid artery from the stump of the external carotid artery to block the entrance of the middle cerebral artery and cause ischemia. After 2 h of ischemia, the suture was removed for 24 h of reperfusion; in the sham operation group, only the blood vessels were dissected, and the other steps were the same as those in the model group. The successful preparation of the model was indicated by the appearance of Homer's sign and contralateral body motor dysfunction after the animals woke up.

[0056] 1.3.2 Animal grouping and drug administration Experimental animals were randomly divided into 5 groups, namely the model group, the positive control edaravone group (6 mg / kg), and the S13 groups (10 mg / kg, 20 mg / kg, 40 mg / kg respectively), with 10 - 12 animals in each group. Each group of S1 test substances and the edaravone injection group were given a single intravenous injection via the tail vein immediately after reperfusion, for a total of 1 injection.

[0057] The drug administration volume for each group was 0.6 mL / 100 g.

[0058] 1.3.3 Determination of neurological deficit score and cerebral infarction volume

[0059] The modified Bederson 5-point system was used to evaluate neurological deficit symptoms.

[0060] For the determination of cerebral infarction volume, after the last neurological function deficit score of the animals, the heads were severed to remove the brains. The olfactory brain, lower brainstem and cerebellum were removed. The remaining part was immediately weighed wet. The brain was cut into 5 slices of basically the same thickness along the coronal plane on ice and incubated in 2,3,5-triphenyltetrazolium chloride dye at 37 °C for 30 min. Normal brain tissue was rose-red, and the infarcted area was white. Then the brain slices were fixed in 10% formaldehyde, and the white tissue was carefully removed and weighed. The percentage of the infarcted tissue weight to the total brain weight was used as the determination index of the infarct volume.

[0061] 2 Results

[0062] 2.1 Effects on cerebral infarction volume and neurological deficit score after ischemia-reperfusion

[0063] Compared with the model group, the drug groups could significantly reduce the infarct volume of rats with cerebral ischemia-reperfusion (P < 0.01); regarding the effect on neurological deficit symptoms, they significantly improved the neurological deficit symptoms of rats. The results are shown in Table 1.

[0064] Table 1 Effects on cerebral infarction volume and neurological deficit score after ischemia-reperfusion

[0065] Group Cerebral infarction volume (%) Neurological deficit score (points) Model group 36.25±3.12 3.2±1.15 <![CDATA[Edaravone group (6 mg·kg -1 )]]> 23.43±1.66* 2.6±1.12 <![CDATA[S1(10mg·kg -1 )]]> 26.21±2.56* 2.1±0.96 <![CDATA[S1(20mg·kg -1 )]]> 23.11±3.44* 1.6±0.67* <![CDATA[S1(40mg·kg -1 )]]> 20.73±2.38** 0.9±0.63*

[0066] *P < 0.05, **P < 0.01, compared with the model group.

[0067] Example 4: Effect of S2 on Focal Cerebral Ischemia-Reperfusion Injury

[0068] 1 Materials and Methods

[0069] 1.1 Experimental Animals

[0070] Sprague-Dawley (SD) rats, male, body weight: 250 - 280 g, SPF grade

[0071] 1.2 Tested Drugs

[0072] Compound S2, dissolved in 2% propylene glycol. Edaravone injection, specification: 5 mL: 10 mg, produced by Nanjing Simcere Dongyuan Pharmaceutical Co., Ltd., diluted with normal saline before use.

[0073] 1.3 Experimental Methods

[0074] 1.3.1 Preparation of focal cerebral ischemia-reperfusion model. Main steps: Rats were anesthetized by intraperitoneal injection of 10% chloral hydrate (350 mg / kg), the right external carotid artery was isolated, ligated and transected, and a nylon suture with a swollen head was slowly inserted about 18 mm along the common carotid artery and internal carotid artery from the stump of the external carotid artery to block the entrance of the middle cerebral artery and cause ischemia. After 2 h of ischemia, the suture was removed for 24 h of reperfusion; in the sham operation group, only the blood vessels were separated, and the other steps were the same as those in the model group. The successful model preparation was indicated by the appearance of Homer sign and contralateral body motor disorder after the animals woke up.

[0075] 1.3.2 Animal grouping and administration Experimental animals were randomly divided into 5 groups, namely the model group, the positive control edaravone group (6 mg / kg), and 3 S2 groups (10 mg / kg, 20 mg / kg, 40 mg / kg respectively), with 10 - 12 animals in each group. Each group of S2 test substances and the edaravone injection group were given a single intravenous injection via the tail vein immediately after reperfusion, for a total of 1 injection.

[0076] The administration volume for each group was 0.6 mL / 100 g.

[0077] 1.3.3 Determination of neurological deficit score and cerebral infarction volume

[0078] The neurological deficit symptoms were evaluated using the modified Bederson 5-point system.

[0079] Measurement of cerebral infarction volume: After the last neurological deficit score was obtained from the animals, the animals were decapitated to remove the brains. The rhinencephalon, lower brainstem, and cerebellum were removed, and the remaining part was immediately weighed wet. The brain was cut into 5 slices of basically the same thickness along the coronal plane on ice, and then incubated in 2,3,5-triphenyltetrazolium chloride dye at 37 °C for 30 min. Normal brain tissue appeared rose red, and the infarct area appeared white. Then the brain slices were fixed in 10% formaldehyde, and the white tissue was carefully dissected and weighed. The percentage of the infarct tissue weight to the total brain weight was used as the determination index of infarct volume.

[0080] 2 Results

[0081] 2.1 Effects on cerebral infarction volume and neurological deficit score after ischemia-reperfusion

[0082] Compared with the model group, the drug groups could significantly reduce the infarct volume of rats with cerebral ischemia-reperfusion (P < 0.01); regarding the effect on neurological deficit symptoms, they significantly improved the neurological deficit symptoms of rats. The results are shown in Table 2.

[0083] Table 2 Effects on cerebral infarction volume and neurological deficit score after ischemia-reperfusion

[0084] Group Cerebral infarction volume (%) Neurological deficit score (points) Model group 35.74±3.15 3.2±1.17 <![CDATA[Edaravone group (6 mg·kg -1 )]]> 23.16±1.76* 2.7±1.13 <![CDATA[S2 (10 mg·kg -1 )]]> 27.51±2.63* 2.2±0.98 <![CDATA[S2 (20 mg·kg -1 )]]> 24.12±3.54* 1.7±0.65* <![CDATA[S2(40mg·kg -1 )]]> 19.75±2.48** 1.1±0.65*

[0085] *P < 0.05, **P < 0.01, compared with the model group.

Claims

1. A fasudil-derived compound of formula I or a pharmaceutically acceptable salt, hydrate or solvate thereof: Wherein, R1 is selected from C1-C4 alkyl or C1-C4 haloalkyl; R2 is selected from hydrogen or C1-C2 alkyl.

2. Any compound according to claim 1, characterized in that Wherein, R1 is selected from C1-C3 alkyl or C1-C3 haloalkyl; R2 is selected from hydrogen, methyl or ethyl.

3. Any compound according to claim 1, characterized in that, Wherein, R1 is selected from methyl, ethyl, isopropyl or dichloromethyl; R2 is selected from methyl.

4. Any compound according to claim 1, characterized in that, The compounds are: Compound 1: as shown in S1; Compound 2: as shown in S2.

5. Use of the compound according to any one of claims 1 to 4 or a pharmaceutically acceptable salt thereof in the preparation of a medicament for treating stroke.

6. A pharmaceutical composition, characterized in that, Comprising the compound according to any one of claims 1 to 4 or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable carrier.

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