A fangchinoline compound, a preparation method thereof, and use thereof in preparing anti-Parkinson's drugs

Abstract

The present invention discloses a fangchinoline compound, a preparation method thereof, and an application thereof in the preparation of an anti-Parkinson's drug, and relates to the technical field of anti-Parkinson's drugs. The present invention discloses a fangchinoline compound, a preparation method thereof, and an application thereof in the preparation of an anti-Parkinson's drug, and provides a new compound—a fangchinoline compound and a preparation method thereof. CCK8 experiments have found that the compound is non-toxic to SH-SY5Y cells at a concentration below 40 μM and has no toxicity to MPP. + The induced SH-SY5Y cells exhibited cytoprotective activity far stronger than that of the lead compound, and had important application value in the preparation of anti-Parkinson's drugs. The compounds disclosed in the present invention have the advantages of simple reaction, mild conditions and cheap and readily available reagents in their preparation methods.

Description

Technical Field

[0001] The present invention relates to the technical field of anti-Parkinson's disease drugs, and in particular to a fangchinoline compound, a preparation method thereof, and an application thereof in the preparation of anti-Parkinson's disease drugs. Background Art

[0002] Parkinson's disease (PD), also known as parkinsonism, is the second most common neurodegenerative disease after Alzheimer's disease. It is highly prevalent among middle-aged and elderly individuals. Its pathological characteristics are the loss of dopaminergic neurons in the substantia nigra and the presence of Lewy bodies composed of α-synuclein. Clinical symptoms of PD include motor symptoms such as bradykinesia, postural instability, and resting tremor. Non-motor symptoms include impairments in autonomic nervous system function, sleep, and mental function.

[0003] Because PD progresses slowly and its initial symptoms are subtle, its onset is often difficult to detect. The etiology of PD is relatively complex, and there is currently a lack of safe and effective monotherapy drugs. Furthermore, most drugs have significant side effects, necessitating continued research and development of treatments for PD. Summary of the Invention

[0004] To solve the above problems, the present invention provides a fangchinoline alkaloid compound, a preparation method thereof, and use thereof in the preparation of anti-Parkinson's drugs. The compound exhibits anti-Parkinson's disease effects in vitro and can be used in the preparation of anti-Parkinson's disease drugs and pharmaceutical compositions containing the same.

[0005] To achieve the above object, the present invention provides a fangchinoline compound, the chemical structural formula of which is:

[0006]

[0007] The above-mentioned fangchinoline compounds are named as follows:

[0008] 7-(2-(((1 1 S,3 1 S)-1 6 ,3 6 ,5 4 -trimethoxy-1 2 ,3 2 -dimethyl-1 1 ,1 2 ,1 3 ,1 4 ,3 1 ,3 2 ,3 3 ,3 4-octa hydro-2,6-dioxa-1(7,1),3(8,1)-diisoquinolina-5(1,3),7(1,4)-dibenzenacyclooctapha ne-3 7 -yl)ox-y)ethoxy)-2H-chromen-2-one; referred to as G2, the molecular formula is: C 48 H 48 N2O9; molecular weight: 796.33.

[0009] A method for preparing the above-mentioned fangchinoline compound comprises the following steps:

[0010] S1. Place 7-hydroxycoumarin in a round-bottom flask, add acetonitrile, K2CO3, and 1,2-dibromoethane, and stir evenly at 61-65°C until the reaction is complete. After the reaction is complete, add water and filter to obtain intermediate a.

[0011] S2. Dissolve 1 times the amount of fangchinoline and 1.2-1.5 times the amount of intermediate a in N,N-dimethylformamide, then add 3 times the amount of anhydrous cesium carbonate and react in an ice bath for 3-4 hours;

[0012] After the reaction of fangchinoline is complete, the target compound G2 is obtained by extraction, drying and purification.

[0013] Preferably, the ratio of the added amounts of 7-hydroxycoumarin:acetonitrile:K2CO3:1,2-dibromoethane is 10 mmol:10 mL:30 mmol:12 mmol.

[0014] Preferably, in step S2, extraction is carried out sequentially with ethyl acetate and saturated brine, drying is carried out using anhydrous sodium sulfate, and separation and purification is carried out using a mixed solvent of petroleum ether and ethyl acetate in a certain proportion according to the principle of column chromatography separation to obtain a pure product.

[0015] Preferably, in step S2, the volume ratio of intermediate a to N,N-dimethylformamide is 1:1.5.

[0016] The present invention provides a fangchinoline compound, a preparation method thereof, and use thereof in the preparation of anti-Parkinson's drugs, which have the following beneficial effects:

[0017] (1) A new compound, a fangchinoline compound and its preparation method are provided. CCK8 experiments show that the compound has no toxicity to SH-SY5Y cells below 40 μM and has no toxicity to MPP. + The induced SH-SY5Y cells exhibited cytoprotective activity far stronger than that of the lead compound, and have important application value in the preparation of anti-Parkinson's drugs;

[0018] (2) The compounds disclosed in the present invention have the advantages of simple reaction, mild conditions and cheap and readily available reagents in terms of preparation method.

[0019] The technical solution of the present invention is further described in detail below through the accompanying drawings and embodiments. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] Figure 1 is the structural formula of compound G2 of the present invention;

[0021] Figure 2 The effect of the compound G2 of the present invention on the viability of SH-SY5Y cells at concentrations of 2.5 μM to 40 μM;

[0022] Figure 3 Compound G2 of the present invention and fangchinoline (30 μM) inhibited MPP + Neuroprotective effect of SH-SY5Y cells induced by cytotoxicity. DETAILED DESCRIPTION

[0023] In order to make the purpose, technical solutions and advantages disclosed in the embodiments of the present invention more clear, the embodiments of the present invention are further described in detail below with reference to the accompanying drawings and examples. It should be understood that the specific embodiments described herein are only used to explain the embodiments of the present invention and are not intended to limit the embodiments of the present invention. Based on the embodiments in this application, all other embodiments obtained by ordinary technicians in this field without making creative work are within the scope of protection of this application.

[0024] Example 1

[0025] A method for preparing a fangchinoline compound comprises the following steps:

[0026] S1. Take 7-hydroxycoumarin (10 mmol) in a round-bottom flask, add 10 mL of acetonitrile, K2CO3 (4.41 g, 30 mmol) and 1,2-dibromoethane (2.25 g, 12 mmol) respectively and stir evenly at 65 °C until the reaction of the raw materials is completed.

[0027] S2. After the reaction is completed, water is added and filtered to obtain intermediate a.

[0028] S3. Dissolve 1 times the amount of fangchinoline and 1.2-1.5 times the amount of intermediate a in N,N-dimethylformamide (2 mL). Add 3 times the amount of anhydrous cesium carbonate and react in an ice bath for 4 h.

[0029] S4. After the reaction of fangchinoline is complete, extract with ethyl acetate and saturated brine in sequence, dry with anhydrous sodium sulfate, and then purify with column chromatography to obtain the target compound G2.

[0030] The obtained compound G2 was named:

[0031] 7-(2-(((1 1 S,3 1 S)-1 6 ,3 6 ,5 4 -trimethoxy-1 2 ,3 2 -dimethyl-1 1 ,1 2 ,1 3 ,1 4 ,3 1 ,3 2 ,3 3 ,3 4 -octa hydro-2,6-dioxa-1(7,1),3(8,1)-diisoquinolina-5(1,3),7(1,4)-dibenzenacyclooctapha ne-3 7 -yl)ox-y)ethoxy)-2H-chromen-2-one;

[0032] The structural formula of the obtained compound G2 is as follows Figure 1 As shown, the molecular formula is: C 48 H 48 N2O9; molecular weight: 796.33.

[0033] The NMR data of the obtained compound G2 are as follows:

[0034] mp118-119℃;yield 56%; 1 H NMR (300MHz, CDCl3) δ7.65 (d, J=9.5Hz,

[0035] 1H),7.35(s,1H),7.31(s,1H),7.12(d,J=8.1Hz,1H),6.89(s,2H),6.83(s,1H),6.81(s,1H),6.79(s,1H),6.54(d,J=5.4Hz,2H) ,6.32(d,J=6.5Hz,2H),6.28(d,J=9.4Hz,1H),5.92(s,1H),3.95(s,3H),3.93–3.87(m,1H),3.78(s,2H),3.74(s,3H),3.73–3.7 0(m,1H),3.65(s,1H),3.64(s,1H),3.55(s,1H),3.51(s,1H),3.40(s,3H),3.29–3.18(m,1H),2.97(s,1H),2.96(s,1H),2.92(s ,1H),2.91–2.85(m,1H),2.82(s,1H),2.78(s,1H),2.69(s,1H),2.62(s,3H),2.52(d,J=13.9Hz,1H),2.43(s,1H),2.35(s,3H); 13 C NMR (75MHz, CDCl3) δ162.10,161.24,155.88,153.68,151.17,149.39,148.71,148.23,147.05,143. 53,143.40,136.34,135.29,134.80,132.68,130.03,128.70,128.55,128.50,128.34,122.98,122.7 7,121.99,121.89,119.89,116.10,113.36,113.10,112.82,112.48,111.57,105.79,101.49,70.51,67.32,64.03,61.40,56.16,55.89,55.84,45.27,44.06,42.73,42.32,41.91,37.48,25.69,22.05.

[0036] Example 2

[0037] Drug combination group: 1000 tablets, each containing 100 mg of active ingredient. Formulation: Compound G2 (100 g) in Example 1, hydroxypropyl methylcellulose (2 g), wheat starch (10 g), sucrose (100 g), and magnesium stearate (6 g). The dosage is based on the patient's age, weight, severity of the disease, etc., with a daily dose range of 0.1 mg-1.0 g, which can be administered once or several times.

[0038] The medical uses of the compounds of the present invention are further demonstrated by the following experiments:

[0039] Test Example 1

[0040] CCK8 assay to determine the cytotoxic effects of compounds on SH-SY5Y cells

[0041] Take 100 μL of the solution with a density of 1×10 4 A suspension of SH-SY5Y cells at a concentration of 100 μL / mL was inoculated into a 96-well plate, and 100 μL of culture medium was added to the edge wells and cultured for 24 hours. The culture medium was gently aspirated, and 100 μL of DMEM culture medium containing a 2.5 μM-40 μM fangchinoline derivative G2 was added to the experimental group, and an equal volume of DMEM culture medium was added to the blank group. The cells were then cultured in a sterile incubator for 24 hours. After 24 hours, 10 μL of CCK-8 was added to each well in the dark, and then cultured for another 1 hour. The absorbance value (A value) at 450 nm was detected using a microplate reader to calculate the cell survival rate.

[0042] Cell survival rate (%) = (average A value of the experimental group) / (average A value of the blank group) × 100%.

[0043] The results are as follows Figure 2 As shown, the CCK-8 assay results showed that the degree of damage to SH-SY5Y cells by compound G2 at five concentrations was determined, and the results showed that compound G2 had no toxicity to SH-SY5Y cells at a concentration of 40 μM.

[0044] Test Example 2

[0045] CCK8 assay to determine the protective effect of compounds on SH-SY5Y cells

[0046] Take 100 μL of the solution with a density of 1×10 4 SH-SY5Y cell suspension with a volume of 100 μL / mL was inoculated into a 96-well plate, and 100 μL of culture medium was added to the edge wells. The plate was cultured in a sterile incubator for 24 hours. After 24 hours, the culture medium was gently removed, and 200 μL of DMEM culture medium was added to the blank group. + The volume of the group added was 200 μL and the concentration was 2 mM MPP + DMEM medium was added to the experimental group, and 100 μL of 20 μM compound G2 and 100 μL of 4 mM MPP were slowly added. + DMEM medium (MPP + The final concentration was 2 mM; the final concentration of compound G2 was 10 μM), and the culture was continued for 24 hours. After 24 hours, the absorbance value (A value) at 450 nm was measured using a microplate reader.

[0047] Cell viability (%) = (average A value of the experimental group - average A value of the blank group) / (average A value of the MPP+ group - average A value of the blank group) × 100%.

[0048] The results are shown in Table 1 and Figure 3 As shown:

[0049] Table 1 MPP of compound G2 + Induced effect on SH-SY5Y cell survival rate (10μM-30μM)

[0050] Cmd Cellviability (% Control) Blank group 100±0 <![CDATA[MPP + ]]> <![CDATA[63.67 ### ±0.69]]> G2 10 μM 69.39***±0.58 G2 20 μM 78.54***±2.22 G2 30 μM 81.90***±0.80 Fan 61.04±1.65

[0051] The results are the mean ± SD of three experiments. ### P<0.001 compared with the blank group, there was statistical significance; ***P<0.001 compared with MPP + The group comparisons were statistically significant.

[0052] It can be seen that MPP + The damage of SH-SY5Y cells induced by β-catenin was successfully established. Compound G2 reversed the effect of β-catenin on MPP at a concentration of 10 μM to 30 μM. + The protective activity of the compound was much stronger than that of the lead compound.

[0053] The present invention discloses a fangchinoline compound and its preparation method and its application in the preparation of anti-Parkinson's drugs. The CCK-8 test found that the compound has no toxicity to SH-SY5Y cells below 40 μM and has no toxicity to MPP. + The induced SH-SY5Y cells exhibited cell protective activity far stronger than that of the lead compound, and had important application value in the preparation of anti-Parkinson's drugs.

[0054] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention rather than to limit the same. Although the present invention has been described in detail with reference to the preferred embodiments, those skilled in the art should understand that they can still modify or replace the technical solutions of the present invention with equivalents, and these modifications or equivalent replacements cannot cause the modified technical solutions to deviate from the spirit and scope of the technical solutions of the present invention.

Claims

1. A fangchinoline compound, characterized in that: Its chemical structural formula is:

2. A method for preparing the fangchinoline compound according to claim 1, characterized in that: The steps include: S1. Place 7-hydroxycoumarin in a round-bottom flask, add acetonitrile, K2CO3, and 1,2-dibromoethane, and stir evenly at 61-65°C until the reaction is complete. After the reaction is complete, add water and filter to obtain intermediate a. S2. Dissolve 1 times the amount of fangchinoline and 1.2-1.5 times the amount of intermediate a in N,N-dimethylformamide, then add 3 times the amount of anhydrous cesium carbonate and react in an ice bath for 3-4 hours; After the reaction of fangchinoline is complete, the target compound G2 is obtained by extraction, drying and purification.

3. The method for preparing the fangchinoline compound according to claim 1, wherein: The ratio of the added amounts of 7-hydroxycoumarin: acetonitrile: K2CO3: 1,2-dibromoethane is 10 mmol: 10 mL: 30 mmol: 12 mmol.

4. The method for preparing the fangchinoline compound according to claim 1, wherein: In step S2, extraction is carried out sequentially with ethyl acetate and saturated brine, and drying is carried out using anhydrous sodium sulfate. According to the principle of column chromatography separation, a mixed solvent of petroleum ether and ethyl acetate in a certain proportion is used for separation and purification to obtain a pure product.

5. The method for preparing the fangchinoline compound according to claim 1, wherein: In step S2, the volume ratio of intermediate a to N,N-dimethylformamide is 1:1.

5.

6. Use of the fangchinoline compound according to claim 1 in the preparation of anti-Parkinson's disease drugs.

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