Icaritin carbamate prodrug as well as preparation method and application thereof

Abstract

The invention discloses an icaritin carbamate prodrug as well as a preparation method and application thereof, belongs to the field of pharmacy, and particularly relates to an icaritin carbamate prodrug as shown in a formula 3-N or 7-N and pharmaceutically acceptable salts thereof as well as a preparation method and application of the icaritin carbamate prodrug. Specifically, a main metabolic site (3-site or 7-site hydroxyl) of a natural flavone compound icaritin is subjected to structural modification through a chemical synthesis method, and an appropriate group is introduced into the site to form the icaritin carbamate prodrug. Amines adopted by the carbamate are aliphatic amines. Due to existence of a carbamate structure, II-phase metabolism of icaritin in SD rats is reduced, compared with icaritin raw material medicine, the 3-N-01 prodrug has the advantages that oral bioavailability is remarkably improved, the 3-N-01 prodrug is the optimal prodrug, clinical medication is expected to be capable of reducing administration dosage and improving patient compliance, and the 3-N-01 prodrug can be applied to preparation of the SD rats. And an effective strategy is expected to be provided for improving the oral bioavailability of phenolic drugs. # imgabs0 #

Description

Technical Field

[0001] The invention belongs to the pharmaceutical field, and specifically relates to carbamate prodrugs of icariin and pharmaceutically acceptable salts thereof, as well as preparation and application thereof. Background Art

[0002] Icaritin (ICT), also known as epimedin, is a natural 8-isopentenyl flavonoid compound and the main active ingredient in epimedium. The domestically marketed dosage form is a soft capsule for the first-line treatment of advanced liver cancer. ICT also has physiological activities such as anti-breast cancer, anti-osteoporosis, antioxidant, and anti-inflammatory.

[0003] ICT has poor water solubility. The 3- or 7-hydroxyl groups of ICT are very likely to undergo phase II metabolic reactions in the intestine and liver, so ICT has a very strong first-pass effect, resulting in a very low bioavailability of ICT (absolute bioavailability is about 4.33%), which limits its clinical application. The structure of ICT is as follows:

[0004]

[0005] Most existing technologies use formulation methods to improve the solubility of ICT. Although the bioavailability of ICT has been improved to a certain extent, there are still certain limitations, such as the use of a large number of excipients, complex preparation methods, and low drug loading. Therefore, combining the structural characteristics of ICT, developing a method to improve the metabolic stability of ICT to improve its bioavailability is a reliable and effective research strategy. Based on the above, we modified the main metabolic sites of ICT (hydroxyl groups at the 3rd or 7th positions) with alkyl carbamates and synthesized carbamate prodrugs of ICT and their salts, aiming to reduce the phase II metabolism of ICT and reduce the first-pass effect. This can not only achieve the purpose of improving the oral bioavailability of ICT, but also has a simple modified group and high drugability. Summary of the invention

[0006] The first object of the present invention is to provide an ICT carbamate prodrug or a pharmaceutically acceptable salt thereof.

[0007] The ICT carbamate prodrug compound of the present invention has the following general structural formula:

[0008]

[0009] Wherein R1 and R2 each independently represent H or C 1-5 Alkyl, preferably H or C 1-2 alkyl.

[0010] The acid radical of the pharmaceutically acceptable salt is a pharmaceutically acceptable inorganic acid or organic acid.

[0011] The following provides definitions of various groups in the compounds of the present invention, which are used uniformly in the specification unless otherwise defined.

[0012] “C 1-5 "Alkyl" includes methyl, ethyl, propyl, butyl and pentyl.

[0013] The inorganic acid is hydrochloric acid, nitric acid, phosphoric acid, sulfuric acid, hydrobromic acid or hydroiodic acid;

[0014] The organic acid is formic acid, acetic acid, propionic acid, butyric acid, hydroxybutyric acid, malic acid, tartaric acid, amino acids, methanesulfonic acid, ethanesulfonic acid, benzenesulfonic acid, naphthalenesulfonic acid, trienesulfonic acid, camphorsulfonic acid, taurine, citric acid, fumaric acid, maleic acid, citric acid, succinic acid, cholic acid or deoxycholic acid.

[0015] The second object of the present invention is to provide a method for preparing ICT carbamate prodrugs or their salts.

[0016] The present invention adopts the following method to synthesize the above-mentioned compound, and the synthesis route is as follows:

[0017] The preparation method of ICT carbamate prodrugs is to react ICT with carbamoyl halide in an aprotic solvent under alkaline conditions;

[0018]

[0019] The aprotic solvent is N,N-dimethylformamide, N,N-dimethylacetamide, tetrahydrofuran, ether, 1,4-dioxane, acetonitrile, dichloromethane, chloroform, benzene, toluene or a mixture of the above solvents; the base is triethylamine, N,N-diisopropylethylamine, pyridine, N,N-lutidine, sodium carbonate, potassium carbonate, sodium bicarbonate, potassium bicarbonate, sodium hydroxide, potassium hydroxide, sodium hydride, potassium hydride, sodium amide, potassium amide, alkyl lithium or hexamethylsilylamide lithium; the carbamoyl halide is carbamoyl chloride or carbamoyl bromide.

[0020] The aprotic solvent is preferably acetonitrile, tetrahydrofuran or N,N-dimethylformamide; and the carbamoyl halide is preferably carbamoyl chloride.

[0021] The ICT carbamate prodrug compound is salified with an inorganic acid or an organic acid in a corresponding solvent to obtain a salt form thereof, wherein the solvent is selected from methanol, ethanol, propanol, isopropanol, dichloromethane, ethyl acetate, tetrahydrofuran, ether, 1,2-dimethoxyethane, acetonitrile and 1,4-dioxane.

[0022] A pharmaceutical composition comprising the ICT carbamate prodrug or a pharmaceutically acceptable salt thereof.

[0023] The third object of the present invention is to provide the above-mentioned ICT carbamate prodrug or its pharmaceutically acceptable salt as an active ingredient for preparing anti-tumor drugs for intravenous injection or oral administration.

[0024] The poor water solubility of ICT is an important reason for its low bioavailability. Therefore, investigating the equilibrium solubility of prodrug compounds in water is helpful to predict the dissolution and absorption of prodrugs in vivo.

[0025] Phase II metabolism is very likely to occur in ICT rats. The present invention is intended to provide a prodrug that can significantly improve the pharmacokinetic properties of ICT in vivo, such as increasing oral bioavailability. Such prodrugs can reduce the glucuronide metabolism rate of ICT in vivo, ultimately achieving the purpose of improving bioavailability, reducing dosage, and reducing toxic side effects.

[0026] Table 1 lists some of the compounds synthesized by the present invention.

[0027] Table 1 Structures and structural identification information of some carbamate prodrugs

[0028]

[0029]

[0030] Beneficial effects of the present invention:

[0031] The present invention utilizes the hydroxyl group at position 3 or 7 in the ICT molecule, performs structural modification by chemical synthesis, and introduces a suitable group at this position to synthesize its novel carbamate prodrug. The amines used in such carbamates are all fatty amines, among which the monomethylamine fragment is easily metabolized in the body, which is convenient for the release of the original drug ICT. At the same time, the presence of the carbamate structure increases the solubility of such prodrugs, increases metabolic stability, and can improve its oral bioavailability.

[0032] The present invention is of great significance for increasing the solubility and metabolic stability of ICT, improving its oral bioavailability, reducing its administration dosage, reducing its toxic side effects, and further searching for new ICT anti-tumor drugs. DETAILED DESCRIPTION

[0033] The present invention is further illustrated below with specific examples, but is not limited thereto.

[0034] According to the general synthetic route of ICT carbamates, the following compounds were prepared respectively.

[0035] Example 1

[0036] In an ice bath, slowly dropwise add ICT (10 mmol) in N, N-dimethylformamide to a mixture of N, N-dimethylformamide and 60% sodium hydride (30 mmol), stir for 30 min, and then add methylaminoformyl chloride (12 mmol). After the addition, the reaction temperature is 5°C-10°C, the reaction is carried out for 2h-4h, and the reaction is monitored by thin layer chromatography.

[0037] The reaction mixture was slowly poured into ice water and dichloromethane for extraction, the organic layer was separated, and the aqueous layer was extracted with dichloromethane. The organic layers were combined, washed with saturated brine (×3), dried over anhydrous sodium sulfate, filtered, and concentrated to obtain an oily substance.

[0038] The oil was dissolved with dichloromethane, and then adjusted to pH 2 with HCl in dichloromethane, and stirred for 10 min. The solvent was evaporated under reduced pressure to obtain a crude product. The product was separated and purified by silica gel column chromatography to obtain compounds 3-N-01 and 7-N-01.

[0039] Example 2

[0040] In an ice bath, slowly add ICT (10 mmol) in N, N-dimethylformamide solution to a mixture of N, N-dimethylformamide and 60% sodium hydride (30 mmol), stir for 30 min, and then add ethylaminoformyl chloride (12 mmol). After the addition, the reaction temperature is 10°C-30°C, the reaction is carried out for 2h-4h, and the reaction is monitored by thin layer chromatography.

[0041] The reaction mixture was slowly poured into ice water and dichloromethane for extraction, the organic layer was separated, and the aqueous layer was extracted with dichloromethane. The organic layers were combined, washed with saturated brine (×3), dried over anhydrous sodium sulfate, filtered, and concentrated to obtain an oily substance.

[0042] The oil was dissolved with dichloromethane, and then adjusted to pH 2 with HCl in dichloromethane, and stirred for 10 min. The solvent was evaporated under reduced pressure to obtain a crude product. The crude product was separated and purified by silica gel column chromatography to obtain compounds 3-N-02 and 7-N-02.

[0043] Example 3

[0044] In an ice bath, slowly add ICT (10 mmol) in N, N-dimethylformamide solution to a mixture of N, N-dimethylformamide and 60% sodium hydride (30 mmol), stir for 30 min, and then add N, N-dimethylcarbamoyl chloride (12 mmol). After the addition, the reaction temperature is 10°C-25°C, the reaction time is 2h-4h, and the reaction is monitored by thin layer chromatography.

[0045] The reaction mixture was slowly poured into ice water and dichloromethane for extraction, the organic layer was separated, and the aqueous layer was extracted with dichloromethane. The organic layers were combined, washed with saturated brine (×3), dried over anhydrous sodium sulfate, filtered, and concentrated to obtain an oily substance.

[0046] The oil was dissolved with dichloromethane, and then adjusted to pH 2 with HCl in dichloromethane, and stirred for 10 min. The solvent was evaporated under reduced pressure to obtain a crude product. The product was purified by silica gel column chromatography to obtain compounds 3-N-03 and 7-N-03.

[0047] Example 4

[0048] In an ice bath, slowly add 10 mmol of N,N-dimethylformamide solution to a mixture of N,N-dimethylformamide and 60% sodium hydride (30 mmol), stir for 30 min, and then add N,N-diethylcarbamoyl chloride (12 mmol). After the addition, the reaction temperature is 10-25°C, the reaction time is 2-4 hours, and the reaction is monitored by thin layer chromatography.

[0049] The reaction mixture was slowly poured into ice water and dichloromethane for extraction, the organic layer was separated, and the aqueous layer was extracted with dichloromethane. The organic layers were combined, washed with saturated brine (×3), dried over anhydrous sodium sulfate, filtered, and concentrated to obtain an oily substance.

[0050] The oil was dissolved with dichloromethane, and then adjusted to pH 2 with HCl in dichloromethane, and stirred for 10 min. The solvent was evaporated under reduced pressure to obtain a crude product. The product was purified by silica gel column chromatography to obtain compounds 3-N-04 and 7-N-04.

[0051] Example 5

[0052] Equilibrium solubility of ICT and ICT carbamate prodrugs in water

[0053] The poor water solubility of ICT is an important reason that limits its bioavailability. Therefore, studying the equilibrium solubility of prodrugs in water is helpful to predict the dissolution and absorption of prodrugs in vivo. Take an excess of ICT API and ICT carbamate prodrug and add an appropriate amount of water, shake in an air constant temperature oscillator at 37°C until equilibrium, and obtain the equilibrium solubility of ICT and its ICT carbamate prodrug in water.

[0054] Table 2 Equilibrium solubility of ICT and its prodrugs in water

[0055]

[0056] As shown in Table 2, compared with ICT, the equilibrium solubility of each prodrug compound in water is improved, and compound 3-N-01 has the highest equilibrium solubility in water.

[0057] Example 6

[0058] In vitro metabolic stability of ICT and ICT carbamate prodrugs

[0059] ICT has a strong first-pass effect, which is an important reason for limiting its bioavailability. Modifying the metabolic site of ICT is beneficial to reduce its phase II metabolism, reduce the first-pass effect, and thus improve bioavailability. In order to determine whether the prodrug can improve its metabolic stability, we used an in vitro liver microsome incubation method to simulate in vivo metabolism. The metabolism of ICT and prodrug in rat liver microsomes was evaluated using the percentage of remaining ICT and its prodrug as an indicator. The metabolic half-life is shown in Table 3.

[0060] Table 3 Phase II metabolic half-life of ICT and ICT carbamate prodrugs (t 1 / 2 )

[0061]

[0062] The results showed that ICT was rapidly metabolized, while prodrugs were metabolized slowly, especially the 3-N series carbamate prodrugs with significantly increased metabolic stability. The results showed that the carbamate strategy could improve the phase II metabolic stability of ICT.

[0063] Example 7

[0064] Pharmacokinetic study in rats

[0065] Sprague-Dawley rats in the experimental group and the control group were gavaged with olive oil solutions of ICT and its carbamate prodrug (120 mg / kg for ICT) respectively. The plasma concentrations of ICT and its prodrug were measured, and the results are shown in Table 4.

[0066] Table 4 Pharmacokinetic parameters of rats after intragastric administration of ICT or ICT carbamate prodrug (n=4)

[0067]

[0068] The pharmacokinetic parameters of ICT and its carbamate prodrug after intragastric administration are shown in Table 4. Compared with ICT alone, the C max and AUC 0-t The C of ICT in the blood of rats after administration of other prodrugs was significantly increased. max and AUC 0-tThe increase was not significant or not significant. After administration of 3-N-01, the blood concentration of ICT can be significantly increased. Compared with the administration of ICT alone, the bioavailability of ICT and the sum of ICT and prodrug in rat blood after administration of 3-N-01 was 4.5 times and 13.9 times that of ICT alone. In vitro tests also showed that 3-N-01 can increase solubility and metabolic stability, and is the best prodrug.

Claims

1. An icariin carbamate prodrug or a pharmaceutically acceptable salt thereof, characterized in that: Its structural formula is as follows: Wherein R1 and R2 are independently selected from H, C 1-5 alkyl; The acid radical of the pharmaceutically acceptable salt is a pharmaceutically acceptable inorganic acid or organic acid.

2. The icariin carbamate prodrug or a pharmaceutically acceptable salt thereof according to claim 1, characterized in that: The C 1-5 The alkyl group includes methyl, ethyl, propyl, butyl and pentyl; the inorganic acid is hydrochloric acid, nitric acid, phosphoric acid, sulfuric acid, hydrobromic acid or hydroiodic acid, and the organic acid is formic acid, acetic acid, propionic acid, butyric acid, hydroxybutyric acid, malic acid, tartaric acid, amino acid, methanesulfonic acid, ethanesulfonic acid, benzenesulfonic acid, naphthalenesulfonic acid, trienesulfonic acid, camphorsulfonic acid, taurine, citric acid, fumaric acid, maleic acid, citric acid, succinic acid, cholic acid or deoxycholic acid.

3. The icariin carbamate prodrug or a pharmaceutically acceptable salt thereof according to claim 1 or 2, characterized in that: Selected from the following compounds:

4. The method for preparing the icariin carbamate prodrug or a pharmaceutically acceptable salt thereof according to claim 1, characterized in that: It is prepared by the reaction of icariin with carbamoyl halide in an aprotic solvent under alkaline conditions; The aprotic solvent is N,N-dimethylformamide, N,N-dimethylacetamide, tetrahydrofuran, ether, 1,4-dioxane, acetonitrile, dichloromethane, chloroform, benzene, toluene or a mixture of the above solvents; the base is triethylamine, N,N-diisopropylethylamine, pyridine, N,N-lutidine, sodium carbonate, potassium carbonate, sodium bicarbonate, potassium bicarbonate, sodium hydroxide, potassium hydroxide, sodium hydride, potassium hydride, sodium amide, potassium amide, alkyl lithium or hexamethylsilylamide lithium; the carbamoyl halide is carbamoyl chloride or carbamoyl bromide.

5. The method for preparing the icariin carbamate prodrug or a pharmaceutically acceptable salt thereof according to claim 4, characterized in that: The aprotic solvent is acetonitrile, tetrahydrofuran or N,N-dimethylformamide; and the carbamoyl halide is carbamoyl chloride.

6. A pharmaceutical composition, characterized in that The invention comprises the icariin carbamate prodrug or a pharmaceutically acceptable salt thereof according to any one of claims 1 to 3, and a pharmaceutically acceptable excipient.

7. Use of the icariin carbamate prodrug or a pharmaceutically acceptable salt thereof according to any one of claims 1 to 3 in the preparation of an anti-tumor drug.

8. Use of the pharmaceutical composition according to claim 6 in the preparation of anti-tumor drugs.

9. The use according to claim 7 or 8, characterized in that The prepared anti-tumor drug is administered by intravenous injection or oral administration.

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