Use of phenolic compounds and / or pharmaceutically acceptable salts thereof for the manufacture of a medicament for the treatment of breast cancer, pharmaceutical composition

Abstract

The application provides application of phenolic compounds and / or pharmaceutically acceptable salts thereof in preparation of drugs for treating breast cancer, a pharmaceutical composition, and relates to the technical field of biological medicines.The application of phenolic compounds and / or pharmaceutically acceptable salts thereof in preparation of drugs for treating breast cancer, wherein the phenolic compounds include compound 1 and / or compound 2.The compound 1 (arnebidin) and the compound 2 ((+)-(R)-de-O-methyllasiodiplodin) used in the application have good inhibiting effects on human triple-negative breast cancer MDA-MB-231 cell strains, the IC 50 value of the compound 1 is 1.16 muM, the IC 50 value of the compound 2 is 0.68 muM, which provides a new choice for developing a therapeutic drug for triple-negative breast cancer, and has a good application prospect in preparation of drugs for treating breast cancer, in particular, drugs for treating triple-negative breast cancer.

Description

Technical Field

[0001] This invention relates to the field of biomedical technology, specifically to the use of phenolic compounds and / or their pharmaceutically acceptable salts in the preparation of medicaments for treating breast cancer, and pharmaceutical compositions thereof. Background Technology

[0002] Malignant tumors are common and prevalent diseases that seriously threaten human health and life. Among them, breast cancer, as the malignant tumor with the highest incidence and mortality rate among women worldwide, has always received widespread attention. Triple-negative breast cancer is a special type of breast carcinoid, accounting for approximately 15-20% of all breast cancers. Compared with other breast cancer subtypes, triple-negative breast cancer lacks immunohistochemical expression of estrogen receptor, progesterone receptor, and human epidermal growth factor receptor-2, making it more aggressive, rapidly developing, with a high recurrence rate and poor prognosis, and belonging to the most malignant type of breast cancer. There is a lack of suitable therapeutic targets for triple-negative breast cancer; commonly used endocrine therapy and targeted therapy are relatively ineffective, and available treatment methods are relatively limited. Currently, there is an urgent need to research and develop drugs that can effectively inhibit the activity of triple-negative breast cancer. Summary of the Invention

[0003] In view of this, the object of the present invention is to provide the use of phenolic compounds and / or their pharmaceutically acceptable salts in the preparation of medicaments for treating breast cancer, and pharmaceutical compositions thereof. Compounds 1 and 2 used in the present invention exhibit high inhibitory activity against triple-negative breast cancer.

[0004] To achieve the above-mentioned objectives, the present invention provides the following technical solution:

[0005] This invention provides the use of phenolic compounds and / or their pharmaceutically acceptable salts in the preparation of medicaments for treating breast cancer, said phenolic compounds comprising compound 1 and / or compound 2:

[0006]

[0007] Preferably, the breast cancer includes triple-negative breast cancer.

[0008] Preferably, the pharmaceutically acceptable salt includes sodium or potassium salts.

[0009] Preferably, the method for preparing the phenolic compound includes the following steps:

[0010] The root of Lithospermum erythrorhizon was extracted using a low-grade alcohol aqueous solution to obtain an alcohol extract.

[0011] The alcohol extract was subjected to a first silica gel column chromatography, and the third and fourth fractions were collected and designated Fr.3 and Fr.4, respectively. The elution method of the first silica gel column chromatography was gradient elution, and the eluents used in the gradient elution included petroleum ether-ethyl acetate and ethyl acetate-methanol, respectively. The volume ratio of petroleum ether to ethyl acetate in the petroleum ether-ethyl acetate mixture was 98:2, 95:5, 90:10, 80:20, 70:30, 60:40, 50:50, 25:75, and 0:100. The volume ratio of ethyl acetate to methanol in the ethyl acetate-methanol mixture was 90:10, 80:20, 50:50, and 0:100, respectively.

[0012] The preparation method of compound 1 includes the following steps:

[0013] The first fraction, Fr.3, was collected by second silica gel column chromatography and designated as Fr.3.1. The eluent used in the second silica gel column chromatography was petroleum ether-ethyl acetate. The elution method for the second silica gel column chromatography was gradient elution, and the volume ratio of petroleum ether to ethyl acetate in the petroleum ether-ethyl acetate mixture was 80:1, 60:1, 30:1, 15:1, and 0:100, respectively.

[0014] The first gel column chromatography was performed on Fr.3.1, and the second fraction was collected and denoted as Fr.3.1.2. The second gel column chromatography was performed on Fr.3.1.2, and the second fraction was collected and denoted as Fr.3.1.2.2. The eluent used in the first gel column chromatography was dichloromethane and methanol, and the volume fraction of dichloromethane in the first gel column chromatography was 20-80%. The eluent used in the second gel column chromatography was methanol.

[0015] The Fr.3.1.2.2 compound was separated by a first reversed-phase chromatography column to obtain compound 1; the eluent used for the first reversed-phase chromatography column separation was methanol-water.

[0016] The preparation method of compound 2 includes the following steps:

[0017] The Fr.4 fraction was subjected to a third silica gel column chromatography to collect the second fraction, denoted as Fr.4.2. The eluent used in the third silica gel column chromatography was dichloromethane and methanol. The elution method of the third silica gel column chromatography was gradient elution, and the volume ratio of dichloromethane to methanol in the third silica gel column chromatography was 100:1, 80:1, 50:1, 25:1, 10:1 and 0:100, respectively.

[0018] The Fr.4.2 fraction was subjected to a third gel column chromatography, and the fourth fraction was collected and designated Fr.4.2.4. The Fr.4.2.4 fraction was then subjected to a fourth gel column chromatography, and the third fraction was collected and designated Fr.4.2.4.3. The eluent used in the third gel column chromatography was dichloromethane and methanol, and the volume fraction of dichloromethane during the third gel column chromatography was 20-80%. The eluent used in the fourth gel column chromatography was methanol.

[0019] The Fr.4.2.4.3 was separated by a second reversed-phase chromatographic column to obtain compound 2; the eluent used for the second reversed-phase chromatographic column separation was methanol-water.

[0020] Preferably, the volume fraction of the lower alcohol in the aqueous solution is 50-100%; the lower alcohol includes methanol and / or ethanol.

[0021] The dry weight of the Lithospermum root and the volume ratio of the lower alcohol aqueous solution used for a single extraction are 4-6 kg: 6-7 L.

[0022] The extraction temperature is 15–40℃, the number of extractions is 2–4, and the extraction time for each extraction is 24–120 hours.

[0023] Preferably, the gels used for the first, second, third, and fourth gel column chromatography include hydroxypropyl dextran gels.

[0024] Preferably, the volume fraction of methanol in the methanol-water separation column used in the first reversed-phase chromatography column is 85-95%;

[0025] The volume fraction of methanol in the methanol-water mixture used for separation on the second reversed-phase column is 86-90%.

[0026] The present invention also provides a pharmaceutical composition comprising an active ingredient and pharmaceutically acceptable excipients; wherein the active ingredient is a phenolic compound and / or a pharmaceutically acceptable salt thereof, and the phenolic compound is the phenolic compound used in the above-described technical solutions.

[0027] Preferably, the content of the active ingredient in the pharmaceutical composition is 0.1 to 99 wt%.

[0028] Preferably, the dosage form of the pharmaceutical composition includes an oral dosage form or an injectable dosage form.

[0029] The compounds 1 (arnebidin) and 2 ((+)-(R)-de-O-methyllasiodiplodin) used in this invention have good inhibitory effects on the human triple-negative breast cancer MDA-MB-231 cell line. The IC50 of compound 1 is [not specified]. 50 The value was 1.16 μM, and the IC50 of compound 2 was...50 With a value of 0.68 μM, it provides a new option for the development of therapeutic drugs for triple-negative breast cancer and has a promising application prospect in the preparation of drugs for treating breast cancer, especially triple-negative breast cancer.

[0030] This invention first extracts Lithospermum erythrorhizon root by cold maceration with a low-grade alcohol aqueous solution, then separates compounds 1 and 2 using silica gel column chromatography, gel chromatography, and reversed-phase chromatography. In this invention, silica gel column chromatography is simple to operate, provides significant separation, and has a large sample capacity; gel chromatography offers good separation, fast separation speed, and convenient operation; reversed-phase chromatography offers flexible sample handling, processing samples from micrograms to milligrams, with high separation efficiency, and yields compounds 1 and 2 with high purity. Compounds 1 and 2 used in this invention exhibit good inhibitory activity against human triple-negative breast cancer MDA-MB-231 cells. Detailed Implementation

[0031] This invention provides the use of phenolic compounds and / or their pharmaceutically acceptable salts in the preparation of medicaments for treating breast cancer, said phenolic compounds comprising compound 1 and / or compound 2:

[0032]

[0033] In this invention, the English name of compound 1 is arnebidin; the English name of compound 2 is (+)-(R)-de-O-methyllasiodiplodin.

[0034] In this invention, the breast cancer includes triple-negative breast cancer.

[0035] In this invention, the pharmaceutically acceptable salt preferably includes a sodium or potassium salt. In this invention, the pharmaceutically acceptable salt is preferably prepared by a salt-forming reaction of a phenolic compound and a metal hydroxide; the salt-forming site of the phenolic compound is the phenolic hydroxyl group. In this invention, the metal hydroxide is preferably sodium hydroxide or potassium hydroxide. This invention does not impose any particular limitation on the conditions of the salt-forming reaction; conventional salt-forming reaction conditions in the art can be used.

[0036] In this invention, the method for preparing the phenolic compound includes the following steps:

[0037] The root of Lithospermum erythrorhizon was extracted using a low-grade alcohol aqueous solution to obtain an alcohol extract.

[0038] The alcohol extract was subjected to a first silica gel column chromatography, and the third and fourth fractions were collected and designated Fr.3 and Fr.4, respectively. The elution method of the first silica gel column chromatography was gradient elution, and the eluents used in the gradient elution included petroleum ether-ethyl acetate and ethyl acetate-methanol, respectively. The volume ratio of petroleum ether to ethyl acetate in the petroleum ether-ethyl acetate mixture was 98:2, 95:5, 90:10, 80:20, 70:30, 60:40, 50:50, 25:75, and 0:100. The volume ratio of ethyl acetate to methanol in the ethyl acetate-methanol mixture was 90:10, 80:20, 50:50, and 0:100, respectively.

[0039] The preparation method of compound 1 includes the following steps:

[0040] The first fraction, Fr.3, was collected by second silica gel column chromatography and designated as Fr.3.1. The eluent used in the second silica gel column chromatography was petroleum ether-ethyl acetate. The elution method for the second silica gel column chromatography was gradient elution, and the volume ratio of petroleum ether to ethyl acetate in the petroleum ether-ethyl acetate mixture was 80:1, 60:1, 30:1, 15:1, and 0:100, respectively.

[0041] The first gel column chromatography was performed on Fr.3.1, and the second fraction was collected and denoted as Fr.3.1.2. The second gel column chromatography was performed on Fr.3.1.2, and the second fraction was collected and denoted as Fr.3.1.2.2. The eluent used in the first gel column chromatography was dichloromethane and methanol, and the volume fraction of dichloromethane in the first gel column chromatography was 20-80%. The eluent used in the second gel column chromatography was methanol.

[0042] The Fr.3.1.2.2 compound was separated by a first reversed-phase chromatography column to obtain compound 1; the eluent used for the first reversed-phase chromatography column separation was methanol-water.

[0043] The preparation method of compound 2 includes the following steps:

[0044] The Fr.4 fraction was subjected to a third silica gel column chromatography to collect the second fraction, denoted as Fr.4.2. The eluent used in the third silica gel column chromatography was dichloromethane and methanol. The elution method of the third silica gel column chromatography was gradient elution, and the volume ratio of dichloromethane to methanol in the third silica gel column chromatography was 100:1, 80:1, 50:1, 25:1, 10:1 and 0:100, respectively.

[0045] The Fr.4.2 fraction was subjected to a third gel column chromatography, and the fourth fraction was collected and designated Fr.4.2.4. The Fr.4.2.4 fraction was then subjected to a fourth gel column chromatography, and the third fraction was collected and designated Fr.4.2.4.3. The eluent used in the third gel column chromatography was dichloromethane and methanol, and the volume fraction of dichloromethane during the third gel column chromatography was 20-80%. The eluent used in the fourth gel column chromatography was methanol.

[0046] The Fr.4.2.4.3 was separated by a second reversed-phase chromatographic column to obtain compound 2; the eluent used for the second reversed-phase chromatographic column separation was methanol-water.

[0047] Unless otherwise specified, the materials and equipment used in this invention are all commercially available products in the field.

[0048] This invention utilizes a low-grade alcohol aqueous solution to extract the root of Lithospermum erythrorhizon to obtain an alcohol extract.

[0049] In this invention, the Lithospermum root is preferably dried and then cut before use. The drying temperature is preferably 20–35°C, more preferably 25°C; the drying time is preferably 8–20 days, more preferably 10–15 days. The length of the cut Lithospermum root is preferably 0.5–2 cm, more preferably 1–1.5 cm.

[0050] In this invention, the volume fraction of the lower alcohol in the aqueous solution is preferably 50% to 100%. In specific embodiments, the volume fraction of the lower alcohol in the aqueous solution can be 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 100%. The lower alcohol preferably includes methanol and / or ethanol, and more preferably methanol.

[0051] In this invention, the preferred volume ratio of the dry weight of the Lithospermum root to the lower alcohol aqueous solution is 4-6 kg: 6-7 L. In specific embodiments, the volume ratio of the dry weight of the Lithospermum root to the lower alcohol aqueous solution for a single extraction can be 4 kg: 6 L, 4.5 kg: 6 L, 5 kg: 6 L, 5.5 kg: 6 L, 6 kg: 6 L, 4 kg: 7 L, 4.5 kg: 7 L, 5 kg: 7 L, 5.5 kg: 7 L, 6 kg: 7 L, 4 kg: 8 L, 4.5 kg: 8 L, 5 kg: 8 L, 5.5 kg: 8 L, or 6 kg: 8 L.

[0052] In this invention, the extraction temperature is preferably 15-40℃, more preferably 20-30℃; the number of extractions is preferably 2-4 times, more preferably 3 times; the extraction time for a single extraction is preferably 24-120 hours. In specific embodiments, the extraction time for a single extraction can be 24 hours, 30 hours, 40 hours, 50 hours, 60 hours, 70 hours, 80 hours, 90 hours, 100 hours, 110 hours, or 120 hours.

[0053] After the extraction is completed, the present invention preferably further includes combining the obtained extracts and concentrating them to obtain an alcohol extract. The present invention does not have any particular limitation on the concentration; any concentration method well known to those skilled in the art can be used until the volume no longer changes, such as vacuum concentration.

[0054] After obtaining the alcohol extract, the present invention performs a first silica gel column chromatography on the alcohol extract, collecting the third and fourth fractions, which are respectively designated as Fr.3 and Fr.4. The elution method for the first silica gel column chromatography is preferably gradient elution, and the eluents used in the gradient elution preferably include petroleum ether-ethyl acetate and ethyl acetate-methanol in sequence. The volume ratio of petroleum ether to ethyl acetate in the petroleum ether-ethyl acetate mixture is preferably 98:2, 95:5, 90:10, 80:20, 70:30, 60:40, 50:50, 25:75, and 0:100; the volume ratio of ethyl acetate to methanol in the ethyl acetate-methanol mixture is preferably 90:10, 80:20, 50:50, and 0:100 in sequence. In the present invention, the particle size of the silica gel used in the first silica gel column chromatography is preferably 200-300 mesh. In the present invention, the first silica gel column chromatography yields 12 fractions, designated as Fr.1 to Fr.12.

[0055] The preparation method of compound 1 is described below.

[0056] After obtaining Fr.3, the present invention performs a second silica gel column chromatography on Fr.3 to collect the first fraction, denoted as Fr.3.1. In the present invention, the eluent used in the second silica gel column chromatography is preferably petroleum ether-ethyl acetate; the elution method of the second silica gel column chromatography is preferably gradient elution, and the volume ratio of petroleum ether to ethyl acetate in the petroleum ether-ethyl acetate mixture during the second silica gel column chromatography is preferably 30:1, 10:1, 5:1, 2:1, and 0:100, respectively; the particle size of the silica gel used in the second silica gel column chromatography is preferably 200-300 mesh. In the present invention, the second silica gel column chromatography yields six fractions, denoted as Fr.3.1 to Fr.3.6.

[0057] After obtaining Fr.3.1, the present invention performs a first gel column chromatography on Fr.3.1 to collect the second component, denoted as Fr.3.1.2. In the present invention, the eluent used in the first gel column chromatography is dichloromethane and methanol, and the volume fraction of dichloromethane in the first gel column chromatography is 20% to 80%. In specific embodiments, the volume fraction of dichloromethane in the first gel column chromatography can be 20%, 30%, 40%, 50%, 60%, 70%, or 80%. The gel used for the first gel column chromatography preferably includes hydroxypropyl dextran gel (Sephadex LH-20). The elution method for the first gel column chromatography is preferably isocratic elution.

[0058] After obtaining Fr.3.1.2, the present invention performs a second gel column chromatography on Fr.3.1.2 to collect a second component, denoted as Fr.3.1.2.2. In the present invention, the eluent used for the second gel column chromatography preferably includes methanol; the gel used for the second gel column chromatography preferably includes hydroxypropyl dextran gel.

[0059] After obtaining Fr.3.1.2.2, the present invention separates Fr.3.1.2.2 using a first reversed-phase chromatography column to obtain compound 1. In the present invention, the eluent used for the first reversed-phase chromatography column separation is preferably methanol-water, and the volume fraction of methanol in the methanol-water used for the first reversed-phase chromatography column separation is preferably 85-95%. In specific embodiments, the volume fraction of methanol in the methanol-water used for the first reversed-phase chromatography column separation can be 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, or 95%.

[0060] The preparation method of compound 2 is described in detail below.

[0061] After obtaining Fr.4, the present invention performs a third silica gel column chromatography on Fr.4 to collect the second fraction, denoted as Fr.4.2. In the present invention, the eluent used in the third silica gel column chromatography is preferably dichloromethane and methanol; the elution method of the third silica gel column chromatography is preferably gradient elution, and the volume ratio of dichloromethane to methanol in the third silica gel column chromatography is preferably 100:1, 80:1, 50:1, 25:1, 10:1, and 0:100, respectively; the particle size of the silica gel used in the third silica gel column chromatography is preferably 200-300 mesh. In the present invention, the third silica gel column chromatography yields five fractions, denoted as Fr.4.1 to Fr.4.5.

[0062] After obtaining Fr.4.2, the present invention performs a third gel column chromatography on Fr.4.2 to collect the fourth component, denoted as Fr.4.2.4. In the present invention, the eluent used in the third gel column chromatography is preferably dichloromethane and methanol, and the volume fraction of dichloromethane in the third gel column chromatography is preferably 20-80%. In specific embodiments, the volume fraction of dichloromethane in the third gel column chromatography can be 20%, 30%, 40%, 50%, 60%, 70%, or 80%. The gel used for the third gel column chromatography preferably includes hydroxypropyl dextran gel. The elution method of the third gel column chromatography is preferably isocratic elution.

[0063] After obtaining Fr.4.2.4, the present invention performs a fourth gel column chromatography on Fr.4.2.4 to collect the third fraction, denoted as Fr.4.2.4.3. In the present invention, the eluent used for the fourth gel column chromatography is preferably methanol; the gel used for the fourth gel column chromatography preferably includes hydroxypropyl dextran gel.

[0064] After obtaining Fr.4.2.4.3, the present invention separates Fr.4.2.4.3 using a second reversed-phase chromatography column to obtain compound 2. In the present invention, the eluent used for separation using the second reversed-phase chromatography column is preferably methanol-water; the volume fraction of methanol in the methanol-water used for separation using the second reversed-phase chromatography column is preferably 86-90%. In specific embodiments, the volume fraction of methanol in the methanol-water used for separation using the second reversed-phase chromatography column can be 86%, 86.5%, 87%, 87.5%, 88%, 88.5%, 89%, 89.5%, or 90%.

[0065] The present invention also provides a pharmaceutical composition comprising an active ingredient and pharmaceutically acceptable excipients; wherein the active ingredient is a phenolic compound and / or a pharmaceutically acceptable salt thereof, and the phenolic compound is the phenolic compound used in the above-described technical solutions.

[0066] In this invention, the content of the active ingredient in the pharmaceutical composition is preferably 0.1 to 99 wt%. In specific embodiments, the content of the active ingredient in the pharmaceutical composition can be 0.1 wt%, 0.5 wt%, 1 wt%, 5 wt%, 10 wt%, 15 wt%, 20 wt%, 25 wt%, 30 wt%, 35 wt%, 40 wt%, 45 wt%, 50 wt%, 55 wt%, 60 wt%, 65 wt%, 70 wt%, 75 wt%, 80 wt%, 85 wt%, 90 wt%, 95 wt%, or 99 wt%.

[0067] In this invention, the dosage form of the pharmaceutical composition preferably includes an oral dosage form or an injectable dosage form. In this invention, the oral dosage form preferably includes powder, tablet, sugar-coated tablet, capsule, drop pill, or solution syrup; the injectable dosage form preferably includes powder injection or solution injection.

[0068] The pharmaceutically acceptable excipients of this invention preferably include pharmaceutical carriers and / or excipients. In this invention, the pharmaceutically acceptable excipients preferably include one or more of lactose, starch, magnesium stearate, and sodium chloride solution. In this invention, the pharmaceutically acceptable excipients are non-toxic to humans and animals and are inert excipients.

[0069] In this invention, the preferred method for preparing the tablets includes the following steps: mixing the active ingredient, lactose, and starch to obtain a mixture; wetting the mixture with water and then sieving it to obtain the sieve residue; mixing the sieve residue with magnesium stearate and then compressing it into tablets to obtain tablets. In this invention, the preferred mass ratio of the active ingredient to lactose is 1:17-20. In specific embodiments, the mass ratio of the active ingredient to lactose can be 1:17, 1:17.5, 1:18, 1:18.5, 1:19, 1:19.5, or 1:20. The preferred mass ratio of the active ingredient to starch is 1:5.3-5.8. In specific embodiments, the mass ratio of the active ingredient to starch can be 1:5.3, 1:5.35, 1:5.4, 1:5.45, 1:5.5, 1:5.55, 1:5.6, 1:5.65, 1:5.7, 1:5.75, or 1:5.8. This invention has no special requirements for the first mixing step, as long as it can be mixed evenly. In this invention, the preferred mass ratio of the active component to magnesium stearate is 1:0.4 to 0.7. In specific embodiments, the mass ratio of the active component to magnesium stearate can be 1:0.4, 1:0.45, 1:0.5, 1:0.55, 1:0.6, 1:0.65, or 1:0.7. In this invention, the water is preferably distilled water or sterile water, more preferably distilled water. In this invention, the preferred pore size of the sieve used for sieving is 75 to 85 mesh, more preferably 80 mesh. This invention has no special requirements for the second mixing step, as long as it can be mixed evenly. This invention has no special requirements for the tableting step; conventional methods in the art can be used.

[0070] In this invention, the tablets are preferably 240-260 mg / tablet, more preferably 250 mg / tablet, and the active ingredient content in the tablets is preferably 9-11 mg, more preferably 10 mg.

[0071] In this invention, the preferred method for preparing the capsules includes the following steps: mixing the active ingredients and starch and sieving them, then taking the sieved material and filling it into magnesium stearate capsule shells to obtain the capsules.

[0072] In this invention, the preferred mass ratio of the active component to starch is 0.8–1.2:1. In specific embodiments, the mass ratio of the active component to starch can be 0.8:1, 0.85:1, 0.9:1, 0.95:1, 1:1, 1.05:1, 1.1:1, 1.15:1, or 1.2:1. This invention does not have special requirements for the mixing process; as long as mixing is possible, it is acceptable. In this invention, the preferred aperture of the sieve used for sieving is 75–85 mesh, more preferably 80 mesh.

[0073] In this invention, the preferred method for preparing the injectable ampoule includes the following steps: dissolving the active ingredient and sodium chloride in water for injection, filtering, and sealing the filtrate in an ampoule bottle under aseptic conditions to obtain the injectable ampoule. In this invention, the mass ratio of the active ingredient to sodium chloride is preferably 1:8 to 11. In specific embodiments, the mass ratio of the active ingredient to sodium chloride can be 1:8, 1:8.5, 1:9, 1:9.5, 1:10, 1:10.5, or 1:11. In this invention, the preferred mass ratio of the active ingredient to the volume ratio of water for injection is 2 mg:90 to 110 mL. In specific embodiments, the mass ratio of the active ingredient to the volume ratio of water for injection can be 2 mg:90 mL, 2 mg:95 mL, 2 mg:100 mL, 2 mg:105 mL, or 2 mg:110 mL.

[0074] In this invention, the pharmaceutical composition is preferably used in the form of a dose per unit body weight, and the route of administration is preferably oral and / or injection, wherein the injection preferably includes intravenous injection or intramuscular injection.

[0075] To further illustrate the present invention, the following detailed description, in conjunction with embodiments, illustrates the application of phenolic compounds and / or their pharmaceutically acceptable salts in the preparation of medicaments for treating breast cancer, and the pharmaceutical compositions thereof, but these descriptions should not be construed as limiting the scope of protection of the present invention.

[0076] Example 1

[0077] S1: 50 kg of Lithospermum root was dried at 25℃ for 10 days and then cut into 1 cm long segments. The cut Lithospermum root segments were soaked in a mixed solvent of 95% methanol and water at 30℃ for extraction. The extraction was repeated 3 times. The amount of mixed solvent of methanol and water used in each soaking and extraction was 60 L and the soaking and extraction time was 72 h. The extracts were combined and then concentrated by vacuum concentration at 40℃ to obtain 3.5 kg of alcohol extract.

[0078] S2: The alcohol extract was subjected to first silica gel column chromatography to obtain 12 fractions, denoted as Fr.1 to Fr.12. The third fraction was collected and denoted as Fr.3; the fourth fraction was collected and denoted as Fr.4. The silica gel used for the first silica gel column chromatography had a particle size of 200 to 300 mesh. Gradient elution was performed using petroleum ether-ethyl acetate with volume ratios of 98:2, 95:5, 90:10, 80:20, 70:30, 60:40, 50:50, 25:75 and 0:100 and ethyl acetate-methanol with volume ratios of 90:10, 80:20, 50:50 and 0:100.

[0079] S3: Perform a second silica gel column chromatography on Fr.3 to obtain 6 components, denoted as Fr.3.1 to Fr.3.6. Collect the first component, denoted as Fr.3.1. The silica gel used for the second silica gel column chromatography has a particle size of 200-300 mesh. The second silica gel column chromatography uses gradient elution with volume ratios of 80:1, 60:1, 30:1, 15:1 and 0:100 petroleum ether-ethyl acetate.

[0080] S4: Perform a first gel column chromatography on Fr.3.1 and collect the second component, denoted as Fr.3.1.2; perform a second gel column chromatography on Fr.3.1.2 and collect the second component, denoted as Fr.3.1.2.2; the eluent used in the first gel column chromatography is dichloromethane-methanol with a volume ratio of 1:1, and the eluent used in the second gel column chromatography is methanol.

[0081] S5: Fr.3.1.2.2 was separated by a first reversed-phase column using 89 v / v% methanol-water as eluent to obtain compound 1 (arnebidin).

[0082] S6: Perform a third silica gel column chromatography on Fr.4 to obtain 5 fractions, denoted as Fr.4.1 to Fr.4.5, and collect the second fraction, denoted as Fr.4.2; the silica gel used in the third silica gel column chromatography has a particle size of 200 to 300 mesh, and gradient elution is performed using dichloromethane-methanol with volume ratios of 100:1, 80:1, 50:1, 25:1, 10:1 and 0:100 respectively.

[0083] S7: Perform a third gel column chromatography on Fr.4.2 to collect the fourth component, denoted as Fr.4.2.4; perform a fourth gel column chromatography on Fr.4.2.4 to collect the third component, denoted as Fr.4.2.4.3; the eluent used in the third gel column chromatography is dichloromethane-methanol with a volume ratio of 1:1, and the eluent used in the fourth gel column chromatography is methanol.

[0084] S8: Fr.4.2.4.3 was separated by a second reversed-phase column using 87 v / v% methanol-water as eluent to obtain compound 2 ((+)-(R)-de-O-methyllasiodiplodin).

[0085] The arnebidin prepared in Example 1 was subjected to NMR and mass spectrometry analysis, and the results are as follows:

[0086] Molecular formula: C 32 H 26 O8; ESIMS: m / z 561[M+Na] + ; 1 H NMR(500MHz,CDCl3)δ:12.49(1H,s,8-OH),12.39(1H,s,5′-OH),12.30(1H,s,5-OH),12.08(1H,s,8′-OH),7.30(1H,d,J=9.3Hz,1H,H-7),7. 28(1H,d,J=9.3Hz,H-6),7.26(1H,d,J=9.3Hz,H-6′),7.24(1H,d,J=9.3Hz,H-7′),4.62(1H,dt,J=9.0,1.4Hz,H-13′),4.38(1H,dt,J=9.0,1 .4Hz,H-13),3.88(1H,td,J=4.5,1.2Hz,H-11′),3.82(1H,dd,J=9.0,4.5Hz,H-12),3.35(1H,ddd,J=9.0,4.5,2.6Hz,H-12′),2.68(1H,dd,J =2.6,1.2Hz,H-2),1.83(3H,d,J=1.4Hz,H3-15),1.72(3H,d,J=1.4Hz,H3-16′),1.61(3H,d,J=1.4Hz,H3-15′),1.54(3H,d,J=1.4Hz,H3-16); 13C NMR(126MHz, CDCl3)δ:195.5(C-4),193.1(C-1),182.6(C-4′),182.2(C-1′),159.0(C-5′),158.5(C-8′),157.3(C-8),15 6.3(C-5),144.3(C-2′),140.0(C-3′),138.6(C-14),137.8(C-14′),129.9(C-7′),129.3(C-6′),129.1(C-6),128.9(C-7) The results show that the compound prepared in Example 1 is arnebidin. (C-13′, 117.4, 116.8, 112.7, 112.4, 111.7, 111.4, 49.2, 45.3, 40.6, 38.1, 37.6, 26.0, 25.8, 18.7, 18.6).

[0087] The (+)-(R)-de-O-methyllasiodiplodin prepared in Example 1 was subjected to NMR analysis, mass spectrometry analysis, and specific rotation analysis. The results are as follows:

[0088] Molecular formula: C 16 H 22 O4; ESIMS: m / z 301[M+Na] + [α]20D=+20(c 0.2,MeOH), 1 HNMR(500MHz,CD3OD)δ:6.14(1H,d,J=2.5Hz,H-3),6.08(1H,d,J=2.5Hz,H-5),5.09(1H,m,H-13),3.19(1H,td,J=12.2,3.5Hz,H -6a),2.43(1H,m,H-6b),1.88(1H,m,H-12a),1.76(1H,m,H-12b),1.48(10H,m,H2-7,8,9,10,11,),1.29(3H,d,J=6.2Hz,H3-16); 13C NMR (126MHz, CD3OD) δ: 173.1 (C-14), 165.9 (C-1), 163.5 (C-3), 149.7 (C-5), 111.8 (C-4), 106.0 (C-15), 101.8 (C-2), 75.9 (C-13), 34.2 (C-6), 32.1 (C-12), 32.0 (C-7), 28.2 (C-8), 25.6 (C-11), 25.4 (C-10), 22.5 (C-9), 20.4 (C-16). Based on the above detection results, the compound prepared in Example 1 is (+)-(R)-de-O-methyllasiodiplodin.

[0089] Test Example 1

[0090] Viability assay of MDA-MB-231 cells against triple-negative breast cancer:

[0091] Triple-negative breast cancer MDA-MB-231 cells were placed in 96-well plates at a density of 5000 cells per well. Cells were incubated with different concentrations of test compounds for 48 h. After incubation, 10 μL of CCK-8 solution was added to each well and the plates were incubated at 37°C for 2 h. Absorbance was then measured at 450 nm using a microplate reader (Molecular Devices, USA). Relative cell viability was calculated, and IC50 was calculated using GraphPadPrism 9.0.0. 50 The values ​​are listed in Table 1. Paclitaxel and β,β-dimethylacrylshikonin were used as positive control drugs.

[0092] Table 1. IC50 inhibitory activity of MDA-MB-231 cells 50 value

[0093] Test compounds <![CDATA[IC 50 (μM)]]> arnebidin 1.16 (+)-(R)-de-O-methyllasiodiplodin 0.68 Paclitaxel 0.48 β,β-dimethylacrylshikonin 2.45

[0094] It is known that the compounds arnebidin and (+)-(R)-de-O-methyllasiodiplodin have good inhibitory activity against triple-negative breast cancer MDA-MB-231 cells.

[0095] The above description is only a preferred embodiment of the present invention. It should be noted that for those skilled in the art, several improvements and modifications can be made without departing from the principle of the present invention, and these improvements and modifications should also be considered within the scope of protection of the present invention.

Claims

1. The use of a phenolic compound or a pharmaceutically acceptable salt thereof in the preparation of a medicament for treating breast cancer, wherein the phenolic compound is compound 1 or compound 2; and the breast cancer is triple-negative breast cancer; Compound 1 Compound 2.

2. The use of phenolic compounds or pharmaceutically acceptable salts thereof in the preparation of medicaments for treating breast cancer, wherein the phenolic compounds are compounds 1 and 2; and the breast cancer is triple-negative breast cancer; Compound 1 Compound 2.

3. The application according to claim 1 or 2, characterized in that, The pharmaceutically acceptable salts include sodium or potassium salts.

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