A sesquiterpene derivative, and a method of preparing and use thereof

Abstract

The application discloses a sesquiterpene derivative, a preparation method and application thereof. The application provides a novel sesquiterpene derivative which is obtained by modifying atractylodin III. Activity research shows that the sesquiterpene derivative has stronger inhibiting effect on IgE or histamine secretion than atractylodin III. It is known to those skilled in the art that allergy is mediated by IgE, and histamine secretion under the stimulation of IgE is an important cause of various allergic symptoms. Therefore, the sesquiterpene derivative provided by the application has the prospect of being developed into a medicine for preventing and / or treating diseases which are prevented and / or treated by inhibiting IgE or histamine secretion, such as allergic diseases like food allergy.

Description

Technical Field

[0001] This invention belongs to the field of medicinal chemistry and relates to the modification, preparation and application of natural products, specifically to a sesquiterpene derivative and its preparation method and uses. Background Technology

[0002] Food allergy is a type I hypersensitivity reaction mediated by immunoglobulin E (IgE). Its core mechanism involves the binding of specific food proteins (such as peanuts, milk, and eggs) to IgE antibodies, triggering the degranulation of mast cells and basophils, releasing inflammatory mediators such as histamine and leukotrienes. This leads to symptoms affecting multiple systems, including the skin, respiratory tract, and digestive tract, and in severe cases, can cause anaphylactic shock and even death. With the rising global incidence of allergic diseases, especially the significant increase in food allergies among children, this disease has become a major threat to public health, severely limiting patients' dietary freedom and quality of life, and incurring long-term psychological burdens and social costs. Current clinical treatment primarily focuses on strict avoidance of allergens, supplemented by antihistamines, corticosteroids to relieve acute symptoms, and adrenaline for emergency resuscitation. However, these methods only temporarily control allergic reactions and cannot block the IgE-mediated immune cascade reaction. Furthermore, long-term medication may lead to drug resistance, metabolic disorders, and other side effects. While desensitization strategies such as oral immunotherapy can induce tolerance through gradual exposure, their efficacy is unstable, the treatment period is long, and there is a risk of inducing severe allergic reactions. Therefore, the development of novel drugs that target the IgE pathway and precisely regulate abnormal immune responses has become a key direction for overcoming the bottleneck in food allergy treatment.

[0003] Atractylodes lactone III is a natural product. Studies have shown that it may have some inhibitory activity against IgE, but its activity has not reached a significant level, and its application in the treatment of food allergies still faces many challenges. Summary of the Invention

[0004] This invention addresses the technical deficiency of insufficient IgE inhibitory activity in existing atractylodes lactone compounds by providing a sesquiterpene derivative, a method for preparing the compound, and its use in preparing anti-allergy drugs.

[0005] The above-mentioned objective of this invention is achieved through the following technical solution:

[0006] A compound with the structure shown in formula (I) or a pharmaceutically acceptable salt or solvate thereof:

[0007]

[0008] A preparation method of the compound shown in the above formula (I) comprises the following steps: dissolving atractylenolide III and p-formylbenzoic acid in dichloromethane, adding 1-ethyl-(3-dimethylaminopropyl) carbodiimide hydrochloride and triethylamine, stirring to react, removing dichloromethane by distillation under reduced pressure after the reaction is completed, adding ethyl acetate to the residue, stirring, washing with saturated sodium bicarbonate aqueous solution, concentrating the ethyl acetate solution, and then loading on a normal phase silica gel column for chromatography, eluting with an eluent, detecting by TLC, collecting the eluent corresponding to the target compound, and concentrating and drying to obtain the target compound.

[0009] Preferably, the molar ratio of atractylenolide III to p-formylbenzoic acid is 1:(1-2).

[0010] Preferably, the molar ratio of 1-ethyl-(3-dimethylaminopropyl) carbodiimide hydrochloride to triethylamine is (1-2):2.

[0011] The use of the above compound or a pharmaceutically acceptable salt or solvate thereof for preparing a medicament for preventing and / or treating a disease that is prevented and / or treated by inhibiting IgE or histamine secretion.

[0012] Preferably, the disease that is prevented and / or treated by inhibiting IgE or histamine secretion is an allergic disease.

[0013] More preferably, the allergic disease is food allergy.

[0014] More preferably, the medicament has the above compound or a pharmaceutically acceptable salt or solvate thereof as an active ingredient, and is prepared into a pharmaceutically acceptable dosage form with a pharmaceutically acceptable excipient.

[0015] More preferably, the excipient is solid, liquid or semi-solid.

[0016] More preferably, the dosage form is a capsule, a tablet, an injection, a granule, a pill or a powder.

[0017] Advantages:

[0018] The present application provides a novel sesquiterpene derivative modified from atractylenolide III. Activity studies show that the sesquiterpene derivative has a stronger inhibitory effect on IgE or histamine secretion than atractylenolide III. It is known to those skilled in the art that allergy is mediated by IgE, and histamine secretion under the stimulation of IgE is an important cause of various allergic symptoms. Therefore, the sesquiterpene derivative provided by the present application has the prospect of being developed into a medicament for preventing and / or treating a disease that is prevented and / or treated by inhibiting IgE or histamine secretion, such as an allergic disease like food allergy. BRIEF DESCRIPTION OF DRAWINGS

[0019] Figure 1 Safety evaluation of different concentrations of atractylodes lactone III and the compound of formula (I) on U266 cells;

[0020] Figure 2 Inhibition of IgE release by different concentrations of atractylodes lactone III and the compound of formula (I) on U266 cells;

[0021] Figure 3 Safety evaluation of different concentrations of atractylodes lactone III and the compound of formula (I) on RBL-2H3 cells;

[0022] Figure 4 Inhibition rate of histamine release under IgE stimulation by different concentrations of atractylodes lactone III and the compound of formula (I). DETAILED DESCRIPTION

[0023] The essential content of the present application will be specifically described below in conjunction with examples, but the protection scope of the present application is not limited by this.

[0024] Example 1: Preparation of the compound of formula (I)

[0025]

[0026] Atractylodes lactone III (1 mmol) and p-formylbenzoic acid (1 mmol) were dissolved in 50 mL of dichloromethane, 1-ethyl-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDCI, 1.5 mmol) and triethylamine (TEA, 2 mmol) were added, and the reaction system was stirred at 25°C for 24 h. After the reaction was completed, dichloromethane was removed by reduced pressure distillation, 30 mL of ethyl acetate was added to the residue, stirred, washed with 20 mL of saturated sodium bicarbonate aqueous solution for 3 times, and the ethyl acetate solution was concentrated and then loaded on a normal phase silica gel column for chromatography, eluted with a mixture of petroleum ether / ethyl acetate (3:1 by volume) at a constant rate, detected by TLC, collected the eluent corresponding to the target compound, and concentrated and dried to obtain the compound of formula (I) with a HPLC purity of 98%.

[0027] Spectroscopic data of the compound of formula (I): 1 H NMR (500 MHz, CDCl3) δ 10.10 (s, 1H), 8.14 (m, 2H), 7.96 (m, 2H), 4.87 (s, 1H), 4.60 (s, 1H), 3.70 (s, 1H), 2.92 (d, J = 14.3 Hz, 1H), 2.77 (m, 1H), 2.38 (m, 2H), 2.04 (m, 1H), 1.95 (m, 1H), 1.94 (s, 3H), 1.82 (m, 1H), 1.56 (m, 1H), 1.34-1.25 (m, 3H), 0.98 (s, 3H). 13C NMR (125 MHz, CDC13) δ 191.46, 171.16, 162.76, 157.96, 147.97, 139.49, 134.64, 130.37, 129.63, 124.35, 107.06, 104.65, 51.29, 49.92, 40.78, 36.67, 35.90, 24.62, 22.19, 16.66, 8.47. ESI-MS: Calcd. for C 23 H 24 O5, [M + Na] + m / z: 403.15, found: 403.14.

[0028] Example 2: Safety evaluation of atractylodes lactone III and the compound of formula (I) on U266 cells and its inhibition rate on IgE

[0029] 1. Experimental materials and instruments

[0030] Phosphate buffer solution (PBS), fetal bovine serum (FBS), RPMI1640 medium, trypan blue, dimethyl sulfoxide (DMSO) and the like were purchased from Beijing Solabio Technology Co., Ltd. U266 cell strains were from the laboratory of Henan University of Chinese Medicine, and IgE detection kit was purchased from mabtech Technology Co., Ltd. Carbon dioxide incubator (Thermo Fisher Scientific); DK-8D constant temperature water bath (Shanghai Jinghong Experimental Equipment Co., Ltd.); centrifuge 5702 (Germany eppendorf); 1510 type microplate reader (Thermo Fisher Scientific); high pressure sterilization pot (Japan); double person single surface purification workbench (Suzhou Purification Equipment Co., Ltd.). Atractylodes lactone III was purchased from Shanghai Aladdin Biochemical Technology Co., Ltd., and the compound of formula (I) was prepared by example 1.

[0031] 2. Experimental method

[0032] Atractylodes lactone III and compound (I) were weighed and dissolved in DMSO to prepare a 40 mg / mL solution. U266 cryopreserved cell lines were revived in a 37°C water bath, DMSO was removed by centrifugation with 10 mL PBS, and the cells were resuspended in RPMI 1640 medium containing 10% FBS and cultured in a 37°C, 5% CO2 incubator. After the cells recovered to a growth state, the U266 cell density was counted using a cell counting chamber, and the cells were diluted to 1×10⁵ cells / mL using pre-prepared RPMI 1640 complete medium (without antibiotics) containing 10% fetal bovine serum. The 96-well plates were divided into control, model, and experimental groups with different concentrations. Each group had three replicates. The control group was added with 100 μL of complete medium, while the model and experimental groups were added with 100 μL of cell suspension at a concentration of 1×10⁵ cells / mL. Different treatments were then applied. The blank and model groups were added with 100 μL of complete culture medium, while the experimental groups were added with 100 μL of complete culture medium containing either the tested atractylodes lactone III or compound (I), respectively, to achieve final drug concentrations of 1.25 μg / mL, 2.5 μg / mL, 5 μg / mL, 10 μg / mL, and 20 μg / mL in the 96-well plates. The culture plates were incubated at 37°C for 72 h. After incubation, U266 cells were in suspension. 50 μL of the mixture from each 96-well plate was removed, and trypan blue staining was used to determine cell viability after treatment with different drug concentrations. The culture plates were then centrifuged at 1000 rpm for 5 min, and the supernatant was collected and the total IgE level was determined using an ELISA kit.

[0033] IgE experimental data are expressed as inhibition rate (I), where E 实验 E represents the detected value of IgE in each experimental group. 模型 E represents the average IgE level in the U266 cells of the model group. 空白 The formula representing the blank group detection value is as follows:

[0034] Inhibition rate Ic for each group = 100% - (E 实验 -E 空白 ) / (E 模型 -E 空白 )×100%.

[0035] Statistical analysis was performed on the survival rate and IgE inhibition rate of U266 cells by different concentrations of atractylodes lactone III and compound (I), and bar charts were plotted for comparison. ****P<0.0001.

[0036] 3. Results

[0037] The results are as follows Figure 1As shown, the compound of formula (I) prepared in the application has high safety in the concentration range of 1.25-20 μg / mL, and has no obvious inhibitory effect on cells. As shown in Table 1 and Figure 2 As shown, in the concentration range of 1.25-20 μg / mL, the inhibitory effect of the compound of formula (I) on IgE is stronger than that of the same concentration of atractylenolide. The above results show that the compound of formula (I) provided in the application has obvious inhibitory effect on IgE, and the effect is better than that of atractylenolide III. Those skilled in the art know that allergy is mediated by IgE, so the compound of formula (I) has more excellent anti-allergic effect than atractylenolide III.

[0038] Table 1 Inhibition rate of different concentrations of compounds on IgE

[0039]

[0040] Example 3: Inhibition rate of atractylenolide III and the compound of formula (I) on histamine release of RBL-2H3 cells under IgE stimulation

[0041] 1. Experimental materials and instruments

[0042] Anti-dinitrophenol (DNP), IgE monoclonal antibody (anti-DNP IgE) were purchased from Sigma Company of the United States, bovine serum albumin (DNP-BSA), PBS buffer, cck8 kit were purchased from Beijing Solabio Technology Co., Ltd. RBL-2H3 cell strain was from the laboratory of Henan University of Chinese Medicine, and a histamine kit was purchased from mabtech Technology Company. Carbon dioxide incubator (Thermo Fisher Scientific); DK-8D constant temperature water bath (Shanghai Jinghong Experimental Equipment Co., Ltd.); centrifuge 5702 (Germany eppendorf); 1510 type enzyme label instrument (Thermo Fisher Scientific); high-pressure sterilization pot (Japan); double-person single-face purification workbench (Suzhou Purification Equipment Co., Ltd.). Atractylenolide III was purchased from Shanghai Aladdin Biochemical Technology Co., Ltd., and the compound of formula (I) was prepared by Example 1.

[0043] 2. Experimental method

[0044] Atractylenolide III and the compound of formula (I) were weighed and dissolved in DMSO to prepare an 80 mg / mL solution. First, the RBL-2H3 cell line in the logarithmic growth phase was collected, and the cell density was adjusted to 1×10 5The cells were inoculated into 96-well culture plates at a concentration of 1 x 105 cells / mL with a volume of 100 μL per well, and blank, control, model and different concentration experimental groups were set up, with three replicate wells in each group. The inoculated culture plates were transferred to a 37°C, 5% CO2 incubator for incubation, so that the cells could be fully adherent for 24 h, and then the culture medium was replaced. The 96-well culture plates were grouped and treated, and blank, model and different concentration experimental groups were set up, with three replicate wells in each group. The blank group was added with 100 μL complete culture medium, and the model and experimental groups were added with 100 μL cell suspension at a concentration of 1 x 105 cells / mL. Then, the cells were treated, and the blank, model and experimental groups were added with 100 μL complete culture medium containing anti-DNP IgE for sensitization, and the cell supernatant was aspirated after 24 h of incubation. The blank and model groups were added with 100 μL complete culture medium, and the experimental groups were added with 100 μL complete culture medium containing the tested atractylodes lactone III or compound of formula (I), so that the final concentration reached 10 μg / mL and 20 μg / mL, respectively. After 30 min of incubation, the cell supernatant was aspirated, and 100 μL DNP-BSA at a concentration of 1 μg / mL was added to the blank, model and experimental groups for activation of the cells, and the reaction was terminated by ice bath for 10 min. The culture plates were centrifuged at 1000 rpm for 5 min, and the supernatant was collected. The histamine level was determined by using an ELISA kit. In addition, 100 μL of a mixture of culture medium and CCK-8 detection reagent (volume ratio of 9:1) was added to each well, and the absorbance (OD value) of each well at a wavelength of 450 nm was determined by using an enzyme-labeled instrument after 4 h of incubation at 37°C in the dark. The relative cell viability of each group was calculated according to the formula:

[0045] Cell viability (%) = (OD 实验 - OD 空白 ) / (OD 对照 - OD 空白 ) x 100%.

[0046] The histamine experimental data were expressed by inhibition (I), wherein E 实验 represents the detection value of histamine in each experimental group, E 模型 represents the average value of histamine in the model group, and E 空白 represents the detection value of histamine in the blank group, and the formula is as follows:

[0047] The inhibition rate Ic of each experimental group = 100% - (E 实验 - E 空白 ) / (E 模型 - E 空白 ) x 100%.

[0048] The survival rate and histamine inhibition rate of RBL-2H3 cells by different concentrations of atractylodes lactone III and compound (I) were statistically analyzed and compared by plotting bar charts. ***P<0.001.

[0049] 3. Results

[0050] The results are as follows Figure 3 As shown, the compound of formula (I) prepared in this invention exhibits high safety within a concentration range of 10–20 μg / mL and has no significant effect on cell viability. Meanwhile, Table 2 and... Figure 4 The compound of formula (I) showed a higher inhibition rate of histamine release from IgE-stimulated mast cells at concentrations of 10–20 μg / mL than atractylodes lactone III. These results indicate that the compound of formula (I) provided by this invention can significantly inhibit histamine secretion under IgE stimulation, and its effect is significantly better than that of atractylodes lactone III. Those skilled in the art know that histamine secretion under IgE stimulation is an important cause of various allergic symptoms; therefore, the compound of formula (I) has a superior anti-allergic effect compared to atractylodes lactone III.

[0051] Table 2. Inhibition rate of histamine secretion under IgE stimulation by different concentrations of compounds.

[0052]

[0053] The above results indicate that the compound of formula (I) provided by the present invention can inhibit IgE and significantly inhibit histamine release under IgE stimulation, and has the potential to be developed into a drug for allergic diseases such as food allergies.

[0054] The purpose of the above embodiments is to specifically illustrate the substantive content of the present invention, but those skilled in the art should know that the scope of protection of the present invention should not be limited to the specific embodiments.

Claims

1. A compound of the following structure (I) or a pharmaceutically acceptable salt thereof: [Structure (I)] （I）。 2. A process for the preparation of a compound of formula (I) as claimed in claim 1, characterized in that, The method comprises the following steps: dissolving atractylenolide III and p-formylbenzoic acid in dichloromethane, adding 1-ethyl-(3-dimethylaminopropyl) carbodiimide hydrochloride and triethylamine, stirring the reaction, removing dichloromethane by distillation under reduced pressure after the reaction is completed, adding ethyl acetate to the residue, stirring, washing with saturated sodium bicarbonate aqueous solution, concentrating the ethyl acetate solution, and then loading the concentrated solution onto a normal phase silica gel column for chromatography, eluting with an eluent, detecting by TLC, collecting the eluent corresponding to the target compound, and concentrating and drying to obtain the target compound.

3. The method of claim 2, wherein: The molar ratio of atractylenolide III to p-formylbenzoic acid is 1: (1-2).

4. The method of claim 2, wherein: The molar ratio of 1-ethyl-(3-dimethylaminopropyl) carbodiimide hydrochloride to triethylamine is (1-2) :

2. 5.Use of the compound of claim 1 or a pharmaceutically acceptable salt thereof for preparing a drug for preventing and treating allergic diseases.

6. Use according to claim 5, characterized in that: The allergic disease is food allergy.

7. Use according to claim 5 or 6, characterized in that: The drug is prepared by taking the compound of claim 1 or a pharmaceutically acceptable salt thereof as an active ingredient, taking pharmaceutically acceptable excipients, and taking the active ingredient and the excipients to form a pharmaceutically acceptable dosage form.

8. Use according to claim 7, characterized in that: The excipients are solid, liquid or semi-solid.

9. Use according to claim 7, characterized in that: The dosage form is a capsule, a tablet, an injection, a granule, a pill or a powder.

Images