TW202110433A - Chidamide-containing pharmaceutical composition and use thereof - Google Patents

Abstract

Provided are a chidamide-containing pharmaceutical composition and a use thereof. The pharmaceutical composition contains chidamide and a P-glycoprotein inhibitor, and the reasonable use of the two can improve the bioavailability of chidamide and the efficacy of chidamide. The combined use of chidamide and the P-glycoprotein inhibitor with an immune checkpoint inhibitor can further effectively improve the anti-tumor efficacy of chidamide.

Description

A chidamide pharmaceutical composition and its application

This application is based on the application with the CN application number 201910804149.8 and the application date on August 28, 2019, and the application with the CN application number 201911102897.8 and the application date on November 12, 2019, and claims its priority. The disclosure content of the CN application is hereby incorporated into this application as a whole.

The invention relates to the technical field of medicines, in particular to a chidamide pharmaceutical composition and its application.

Chidamide (English name: Chidamide) is a new type of anti-cancer drug with a brand-new chemical structure and global intellectual property rights independently designed and synthesized by Shenzhen Microchip Biotechnology Co., Ltd. Chidamide has histone deacetylase inhibitory activity and belongs to the class of epigenetic regulators, which can be used for the treatment of various diseases, such as cancer, viral diseases, autoimmune diseases, metabolic diseases, etc. The results of clinical studies that have been completed show that Chidamide has definite curative effects on various lymphomas including peripheral T-cell lymphoma, and it also has certain effects on some other solid tumors such as breast cancer, lung cancer, kidney cancer, colorectal cancer, and intrauterine cancer. Membrane cancer has the effect of clinical benefit, and its comprehensive indicators (safety, preliminary efficacy, pharmacokinetic characteristics) are better than the results of the same period of international studies of drugs with similar mechanisms of action.

After oral solid preparations enter the body, they all need to undergo a dissolution process before they can be absorbed by the body through the biofilm. The dissolution rate of poorly soluble drugs is limited by solubility, which affects drug absorption, so the effect is slow and the bioavailability is low.

Chidamide has extremely low solubility in water and low bioavailability, which leads to clinically poor drug absorption, high doses, and high gastrointestinal toxicity. Therefore, improving the bioavailability of chidamide is of great significance in reducing drug consumption, reducing drug production costs, and reducing gastrointestinal toxicity.

In view of this, the purpose of the present invention is to provide a chidamide pharmaceutical composition, which can significantly improve the bioavailability of chidamide, and can be applied to the fields of medicine preparation and disease treatment related to chidamide.

In order to achieve the above-mentioned purpose of the invention, the present invention provides the following technical solutions: A chidamide pharmaceutical composition includes chidamide and a P-glycoprotein inhibitor, and further, it also includes an immune checkpoint inhibitor.

式（1）The chidamide of the present invention has the structure shown in formula (1), and its chemical name is N-(2-amino-4-fluorophenyl)-4-[N-[(E)-3-(3- (Pyridine)acryloyl]aminomethyl]benzamide, in its structural formula, the configuration of 3-pyridylacryloyl is E type.

Formula 1)

The research of the present invention found that the reason for the low oral bioavailability of Chidamide is not only its poor water solubility, but Chidamide as a substrate of intestinal P-gp is another important reason for reducing its oral bioavailability. The effect of the P-gp efflux pump in the digestive tract reduces the absorption of chidamide into the blood. At the same time, the P-gp efflux pump also exists on the surface of tumor cells, which can lead to reduced drug efficacy and tumor cells to drug resistance. Therefore, the present invention starts from the discovery of the root cause of the problem and provides the combined application of Chidamide and P-gp inhibitor, which can significantly increase the bioavailability of Chidamide and improve the efficacy of Chidamide.

In some embodiments, the pharmaceutical composition includes chidamide and a P-glycoprotein inhibitor. The P-glycoprotein inhibitor of the present invention is selected from artificially synthesized P-glycoprotein inhibitors, natural P-glycoprotein inhibitors and pharmaceutical excipients P-glycoprotein inhibitors. In some embodiments, the P-glycoprotein inhibitor includes, but is not limited to, verapamil, dexverapamil, cyclosporin, dex-nicodipine, transfluoxetine, tamoxifen, valapamil Spoda (PSC833), Tariquidar, TPGS, β cyclodextrin, PEG, piperine, VX-710, tetrandrine, FG020326, S-9788, GF-120918, LY-335979, DMDCK and MMDCK, optional Any one or two or more of them; in the specific embodiment of the present invention, the P-glycoprotein inhibitor is preferably verapamil or Tariquidar.

In some embodiments, the mass ratio of chidamide and P-glycoprotein inhibitor is 10:1 to 1:10; in specific embodiments of the present invention, the mass ratio is 10:1, 9:1, 8:1, 7:1, 6:1, 5:1, 4:1, 3:1, 2:1, 1:1, 1:2, 1:3, 1:4, 1:5, 1: 6, 1:7, 1:8, 1:9 or 1:10. In some embodiments, the mass ratio of chidamide and P-glycoprotein inhibitor is 5:1 to 1:5, for example, 5:1, 4.5:1, 4:1, 3.5:1, 3: 1, 2.5:1, 2:1, 1.9:1, 1.8:1, 1.7:1, 1.6:1, 1.5:1, 1.4:1, 1.3:1, 1.2:1, 1.1:1, 1:1, 1:1.1, 1:1.2, 1:1.3, 1:1.4, 1:1.5, 1:1.6, 1:1.7, 1:1.8, 1:1.9, 1:2, 1:2.5, 1:3, 1: 3.5, 1:4, 1:4.5 or 1:5.

In some embodiments, the pharmaceutical composition includes chidamide, and verapamil or Tariquidar, wherein the mass ratio of chidamide to verapamil or Tariquidar is 5:1 to 1:5, for example 5:1, 4.5:1, 4:1, 3.5:1, 3:1, 2.5:1, 2:1, 1.9:1, 1.8:1, 1.7:1, 1.6:1, 1.5:1, 1.4: 1, 1.3:1, 1.2:1, 1.1:1, 1:1, 1:1.1, 1:1.2, 1:1.3, 1:1.4, 1:1.5, 1:1.6, 1:1.7, 1:1.8, 1:1.9, 1:2, 1:2.5, 1:3, 1:3.5, 1:4, 1:4.5 or 1:5. In some embodiments, the pharmaceutical composition includes chidamide, a P-glycoprotein inhibitor, and an immune checkpoint inhibitor.

The immune checkpoint inhibitor of the present invention is selected from one or more of anti-CTLA-4 antibody, anti-PD-1/PD-L1 antibody, anti-LAG-3 antibody, anti-TIM-3 antibody and anti-TIGIT antibody. Among them, anti-CTLA-4 antibodies include but are not limited to Ipilimumab and Tremelimumab. Anti-PD-1/PD-L1 antibodies include but are not limited to RMP 1-14, pembrolizumab, nivolumab, ztezolizumab, avelumab, durvalumab, tislelizumab, carrelizumab, tereprizumab, sindizumab, and Nosumab, KN035, GLS-010 and CS1001 can be selected from any one or two or more of them. Anti-LAG-3 antibodies include but are not limited to MGD013, FS118, Relatlimab, GSK2831781, LAG525, REGN3767 and TSR033. Anti-TIM-3 antibodies include but are not limited to MBG453, TSR022, LY3321367, RO7121661, Sym023, INCAGN2390 and SHR1702. Anti-TIGIT antibodies include but are not limited to tiragolumab, BGB-A1217, MK-7684, BMS-986207, AB154, ASP8374, IBI-939 and OMP-313M32.

In some embodiments, the mass ratio of the chidamide, the P-glycoprotein inhibitor, and the immune checkpoint inhibitor is (1-10): (1-10): (1-10); wherein the western Dalbenamide, P-glycoprotein inhibitor and immune checkpoint inhibitor can be independently selected from 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 within the mass ratio range of 1-10. . In a specific embodiment of the present invention, the present invention provides a set of chidamide, P-glycoprotein inhibitors and immune checkpoint inhibitors with a mass ratio of 5:2:2 to illustrate the drug of the present invention as an example The composition has an anti-tumor effect.

In the pharmacokinetic test provided by the present invention, the AUC (area under the drug-time curve) of the pharmaceutical composition of the present invention is significantly improved compared with that of chidamide, and the combined multiple is more than 1 times, and the maximum can reach 5 At the same time, the maximum concentration of the drug is also significantly increased, the combined multiple is more than 1.5 times, and the highest can be more than 6 times; these results verify that chidamide, as a substrate of intestinal P-gp, reduces its oral bioavailability Another important reason for the degree.

In the mouse tumor efficacy test of chidamide combined with P-glycoprotein inhibitor and immune checkpoint inhibitor of the present invention, compared with each single-drug test group, the chidamide combined with Tariquidar group showed good anti-tumor effects Drug efficacy, and the most significant anti-tumor efficacy is Chidamide combined with Tariquidar and Anti-PD-1 antibody group, indicating that Chidamide combined with P-gp inhibitors and immune checkpoint inhibitors can be further effectively improved Chidamide's anti-tumor efficacy.

In the present invention of Chidamide combined with P-gp inhibitors and immune checkpoint inhibitors to enhance the cytokine TNF-α and IL-6 gene expression test, compared with each single-drug test group, Chidamide combined with Tariquidar The group showed a certain effect of up-regulating the expression of IL-6 and TNF-α genes; while the Chidamide combined with Tariquidar and Anti-PD-1 antibody group showed a more significant up-regulation of the gene expression of these two cytokines, indicating Chidamide, P-gp inhibitors and immune checkpoint inhibitors are used in combination to further promote the expression of immune cytokines to exert anti-tumor efficacy.

Based on the above significant technical advantages, the present invention proposes the use of the pharmaceutical composition in the preparation of chidamide preparations and/or drugs for the treatment of diseases related to the action mechanism of histone deacetylase.

In some embodiments, the disease related to the action mechanism of histone deacetylase includes, but is not limited to, cancer, viral disease, autoimmune disease, and blood system disease. In some embodiments, the disease related to the mechanism of histone deacetylase action is cancer, such as lymphoma, breast cancer, lung cancer, kidney cancer, colorectal cancer, or endometrial cancer. In some embodiments, the disease related to the mechanism of histone deacetylase action is peripheral T-cell lymphoma.

Based on the above application, the present invention also provides a chidamide preparation comprising the chidamide pharmaceutical composition of the present invention, more specifically, the chidamide pharmaceutical composition and pharmaceutically acceptable excipients. Among them, the dosage form of the preparation includes, but is not limited to, tablets, capsules, pills, oral liquid preparations (such as syrups), granules, powders, ointments and dripping pills. All preparations can be prepared as sustained-release medicaments and controlled-release medicaments. And so on any form of release agent.

The pharmaceutically acceptable excipients can be adjusted and selected according to the dosage form to be prepared. In the present invention, the auxiliary material can be selected from one or more of fillers, disintegrants, binders, and lubricants.

Based on the above, the present invention further provides a drug combination, which includes chidamide and a P-glycoprotein inhibitor. In some embodiments, the P-glycoprotein inhibitor is selected from the group consisting of artificially synthesized P-glycoprotein inhibitors, natural P-glycoprotein inhibitors, and pharmaceutical excipients P-glycoprotein inhibitors. In some embodiments, the P-glycoprotein inhibitor includes, but is not limited to, verapamil, dexverapamil, cyclosporin, dex-nicodipine, transfluoxetine, tamoxifen, valapamil Any of SPC833, Tariquidar, TPGS, β cyclodextrin, PEG, piperine, VX-710, tetrandrine, FG020326, S-9788, GF-120918, LY-335979, DMDCK and MMDCK One or more than two. In a specific embodiment of the present invention, the P-glycoprotein inhibitor is preferably verapamil or Tariquidar.

In some embodiments, the mass ratio of chidamide and P-glycoprotein inhibitor is 10:1 to 1:10; in specific embodiments of the present invention, the mass ratio is 10:1, 9:1, 8:1, 7:1, 6:1, 5:1, 4:1, 3:1, 2:1, 1:1, 1:2, 1:3, 1:4, 1:5, 1: 6, 1:7, 1:8, 1:9 or 1:10. In some embodiments, the mass ratio of chidamide and P-glycoprotein inhibitor is 5:1 to 1:5, for example, 5:1, 4.5:1, 4:1, 3.5:1, 3: 1, 2.5:1, 2:1, 1.9:1, 1.8:1, 1.7:1, 1.6:1, 1.5:1, 1.4:1, 1.3:1, 1.2:1, 1.1:1, 1:1, 1:1.1, 1:1.2, 1:1.3, 1:1.4, 1:1.5, 1:1.6, 1:1.7, 1:1.8, 1:1.9, 1:2, 1:2.5, 1:3, 1: 3.5, 1:4, 1:4.5 or 1:5.

In some embodiments, the drug combination includes chidamide, and verapamil or Tariquidar, wherein the mass ratio of chidamide to verapamil or Tariquidar is 5:1 to 1:5, such as 5 :1, 4.5:1, 4:1, 3.5:1, 3:1, 2.5:1, 2:1, 1.9:1, 1.8:1, 1.7:1, 1.6:1, 1.5:1, 1.4:1 , 1.3:1, 1.2:1, 1.1:1, 1:1, 1:1.1, 1:1.2, 1:1.3, 1:1.4, 1:1.5, 1:1.6, 1:1.7, 1:1.8, 1 :1.9, 1:2, 1:2.5, 1:3, 1:3.5, 1:4, 1:4.5 or 1:5.

In some embodiments, the drug combination includes chidamide, P-glycoprotein inhibitor, and immune checkpoint inhibitor. In some embodiments, the immune checkpoint inhibitor is selected from one or both of anti-CTLA-4 antibodies, anti-PD-1/PD-L1 antibodies, anti-LAG-3 antibodies, anti-TIM-3 antibodies, and anti-TIGIT antibodies. More than species. In some embodiments, the anti-CTLA-4 antibodies include but are not limited to Ipilimumab and Tremelimumab. In some embodiments, the anti-PD-1/PD-L1 antibody includes, but is not limited to, RMP 1-14, pembrolizumab, nivolumab, ztezolizumab, avelumab, durvalumab, tislelizumab, carrelizumab, tereprizumab , Sindizumab, Genozumab, KN035, GLS-010 and CS1001, can be selected from any one or two or more of them. In some embodiments, the anti-LAG-3 antibodies include but are not limited to MGD013, FS118, Relatlimab, GSK2831781, LAG525, REGN3767, and TSR033. Anti-TIM-3 antibodies include but are not limited to MBG453, TSR022, LY3321367, RO7121661, Sym023, INCAGN2390 and SHR1702. In some embodiments, the anti-TIGIT antibodies include, but are not limited to, tiragolumab, BGB-A1217, MK-7684, BMS-986207, AB154, ASP8374, IBI-939, and OMP-313M32.

In some embodiments, the mass ratio of the chidamide, the P-glycoprotein inhibitor, and the immune checkpoint inhibitor is (1-10): (1-10): (1-10); wherein the western Dalbenamide, P-glycoprotein inhibitor and immune checkpoint inhibitor can be independently selected from 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 within the mass ratio range of 1-10. .

In some embodiments, the chidamide, P-glycoprotein inhibitor, and immune checkpoint inhibitor are located in the same or separate formulation units.

In another aspect, the present invention provides a method for inhibiting HDAC, which comprises administering to a subject in need thereof an effective amount of the chidamide pharmaceutical composition, chidamide preparation or drug combination described herein step.

In some embodiments, the drugs in the pharmaceutical composition may be administered simultaneously, separately, or sequentially.

In another aspect, the present invention provides a method for treating diseases related to the mechanism of histone deacetylase action, which comprises administering an effective amount of the chidamide pharmaceutical composition described herein to a subject in need thereof, Steps of chidamide preparation or drug combination.

In some embodiments, the disease related to the action mechanism of histone deacetylase is selected from cancer, viral disease, autoimmune disease and blood system disease. In some embodiments, the disease related to the action mechanism of histone deacetylase is cancer, such as lymphoma, breast cancer, lung cancer, kidney cancer, colorectal cancer, or endometrial cancer. In some embodiments, the disease related to the mechanism of action of histone deacetylase is peripheral T-cell lymphoma.

In some embodiments, the drugs in the pharmaceutical composition may be administered simultaneously, separately, or sequentially.

It can be seen from the above technical solutions that one of the reasons for the poor bioavailability of chidamide in the present invention is that it is a substrate of intestinal P-gp. Therefore, the present invention provides a substance containing chidamide and P-sugar. The drug composition of protein inhibitor, reasonable application of the two can significantly improve the bioavailability of chidamide and improve the efficacy of chidamide. The present invention also found through research that Chidamide combined with P-glycoprotein inhibitors showed good anti-tumor efficacy, and the most significant anti-tumor efficacy is Chidamide combined with P-glycoprotein inhibitors and immune checkpoints. Inhibitors, indicating that the combined application of Chidamide, P-gp inhibitors and immune checkpoint inhibitors can further effectively improve the anti-tumor efficacy of Chidamide.

The present invention discloses a chidamide pharmaceutical composition and its preparation method and application. Those skilled in the art can learn from the content of this article and appropriately improve the process parameters to achieve it. In particular, it should be pointed out that all similar replacements and modifications are obvious to those skilled in the art, and they are all deemed to be included in the present invention. The pharmaceutical composition and its application of the present invention have been described by the preferred embodiments. The relevant personnel can obviously modify or appropriately modify the pharmaceutical composition and its application herein without departing from the content, spirit and scope of the present invention. In combination with, to realize and apply the technology of the present invention.

The following is a further description of a chidamide pharmaceutical composition and its application provided by the present invention. Example 1: In vitro culture of Caco-2 cell absorption experiment

Cell culture: Coca-2 cells were seeded on the polyethylene membrane (PET) of the 96-well plate insertion system, 2 x 10 ⁵ cells per square centimeter, until 21-28 days to form a monolayer of confluent cells. Change the medium every 3-4 days.

Experimental method: Dilute the test compound with transport buffer (HBSS+1%BSA) in a 10 mM stock solution to a concentration of 10μm, and smear it on the top or basolateral side of the cell monolayer. Under the conditions of 37°C and 5% carbon dioxide and 95% relative humidity, during the 120-minute incubation, the permeation of the test compound from A to B or B to A was measured repeatedly. In addition, the elution ratio of each compound was also determined. According to the peak area ratio of analyte/Is, the test compound and the reference compound were quantified by LC-MS/MS analysis. The experimental results are as follows in Table 1: Table 1 Summary Mean Papp (10 ^-6 cm/s) Efflux Ratio Rank Compound ID A to B B to A Papp P-gp Atenolol 1.73 1.33 0.77 Low Propranolol 22.10 11.16 0.51 High Digoxin 0.48 8.06 16.64 Low Substrate Digoxin+GF120918A 3.83 3.16 0.82 Moderate CS055 0.96 3.24 3.38 Low Substrate CS055+GF120918A 1.45 1.47 1.01 Low CS02 0.84 0.94 1.12 Low CS02+GF120918A 0.81 0.69 0.85 Low CS03 8.22 4.85 0.59 Moderate CS03+GF120918A 7.89 3.01 0.38 Moderate CS04 2.91 5.16 1.78 Moderate CS04+GF120918A 5.11 3.69 0.72 Moderate In the above table, CS055 is Chidamide. The Efflux ratio is calculated by Papp(B to A)/Papp(A to B). When the ratio is greater than 2, it means that it is P-gp-mediated efflux ratio. The platoon mechanism works. The efflux rate of chidamide reached 3.38, indicating that chidamide is a substrate of P-gp. The ratio of other drugs except Digoxin is less than 2, indicating that not all drugs are substrates of P-glycoprotein. Example 2: Rat pharmacokinetic test of chidamide combined with verapamil

Twelve SD rats were selected and randomly divided into 2 groups, each with 6 rats, half male and half female. During the experiment, SD rats were fasted overnight before administration. Each rat in group 1 was given a suspension of chidamide (20mg/kg) alone, and each rat in group 2 was given a mixture of chidamide (20mg/kg) and verapamil (25mg/kg) at the same time. Suspension. Collect blood samples before administration and 15min, 0.5h, 1h, 2h, 4h, 6h, 8h after administration, each sample is collected about 0.2ml, heparin sodium is anticoagulated, placed on ice after collection, and centrifuged within 1 hour The plasma was separated, the drug was extracted with acetonitrile, and the concentration of the drug in the plasma was detected by LC-MS/MS. The test results are shown in Table 2 and Figure 1. Table 2 Comparison of pharmacokinetic parameters of the two groups of animals Pharmacokinetic parameters unit Chidamide Chidamide + Verapamil Combined multiple AUC _(0-t) h·ng/mL 1688.1 4789.4 2.83 C _max ng/mL 703.6 2,765.6 3.93 The results of pharmacokinetic experiments in rats showed that when chidamide was used in combination with the first-generation P-gp inhibitor verapamil, the blood exposure AUC increased by 2.83 times, and the C _max increased by 3.93 times. The above experiments further confirmed that Chidamide is a substrate of P-gp, and P-gp inhibitors can significantly increase the bioavailability of Chidamide in vivo. Example 3: Pharmacokinetic test of Chidamide combined with Tariquidar in rats

Twelve SD rats were selected and randomly divided into 2 groups, each with 6 rats, half male and half female. During the experiment, SD rats were fasted overnight before administration. Each rat in group 1 was given a suspension of chidamide (20mg/kg) alone, and each rat in group 2 was given a suspension of chidamide (20mg/kg) and Tariquidar (12mg/kg) at the same time. Collect blood samples before administration and 15min, 0.5h, 1h, 2h, 4h, 6h, 8h after administration, each sample is collected about 0.2ml, heparin sodium is anticoagulated, placed on ice after collection, and centrifuged within 1 hour The plasma was separated, the drug was extracted with acetonitrile, and the concentration of the drug in the plasma was detected by LC-MS/MS. The test results are shown in Table 3 and Figure 2. Table 3 Comparison of pharmacokinetic parameters between the two groups of animals Pharmacokinetic parameters unit Chidamide Chidamide + Tariquidar Combined multiple AUC _(0-t) h·ng/mL 2,757.1 14517.6 5.26 C _max ng/mL 970.0 6,139.6 6.33 The results of pharmacokinetic experiments in rats showed that when Chidamide was used in combination with the third-generation P-gp inhibitor Tariquidar, the blood exposure AUC increased significantly to 5.26 times, and C _max increased to 6.33 times. The above experiments further confirmed that Chidamide is a substrate of P-gp, and P-gp inhibitors can significantly increase the bioavailability of Chidamide in vivo. Example 4: Rat pharmacokinetic test of chidamide combined with vitamin E polyethylene glycol succinate (TPGS)

Twelve SD rats were selected and randomly divided into 2 groups, each with 6 rats, half male and half female. During the experiment, SD rats were fasted overnight before administration. Each rat in group 1 was given a suspension of chidamide (20mg/kg) alone, and each rat in group 2 was given a suspension of chidamide (20mg/kg) and TPGS (3.3mg/kg) at the same time . Collect blood samples before administration and 15min, 0.5h, 1h, 2h, 4h, 6h, 8h after administration, each sample is collected about 0.2ml, heparin sodium is anticoagulated, placed on ice after collection, and centrifuged within 1 hour The plasma was separated, the drug was extracted with acetonitrile, and the concentration of the drug in the plasma was detected by LC-MS/MS. The test results are shown in Table 4 and Figure 3. Table 4 Comparison of pharmacokinetic parameters of the two groups of animals Pharmacokinetic parameters unit Chidamide Chidamide+TPGS Combined multiple AUC _(0-t) h·ng/mL 2,757.1 2,782.4 1 C _max ng/mL 970.0 1685.1 1.73 The results of pharmacokinetic experiments in rats showed that when Chidamide was used in combination with the pharmaceutical excipient P-gp inhibitor TPGS, the blood exposure AUC did not change much, but the C _max could be increased to 1.73 times that of the single drug. . It shows that the pharmaceutical excipient P-gp inhibitor can also promote the absorption of chidamide. However, the effect is insufficient compared with P-gp inhibitors such as verapamil and Tariquidar. Example 5: Rat pharmacokinetic test of chidamide combined with hydroxypropyl β cyclodextrin

Twelve SD rats were selected and randomly divided into 2 groups, each with 6 rats, half male and half female. During the experiment, SD rats were fasted overnight before administration. Each rat in group 1 was given a suspension of chidamide at 20 mg/kg alone, and each rat in group 2 was given a mixture of chidamide (20 mg/kg) containing 40% hydroxypropyl β-cyclodextrin. Suspension. Collect blood samples before administration and 15min, 0.5h, 1h, 2h, 4h, 6h, 8h after administration, each sample is collected about 0.2ml, heparin sodium is anticoagulated, placed on ice after collection, and centrifuged within 1 hour The plasma was separated, the drug was extracted with acetonitrile, and the concentration of the drug in the plasma was detected by LC-MS/MS. The test results are shown in Table 5 and Figure 4. Table 5 Comparison of pharmacokinetic parameters between the two groups of animals Pharmacokinetic parameters unit Chidamide Chidamide + hydroxypropyl β cyclodextrin Combined multiple AUC _(0-t) h·ng/mL 2,757.1 3020.6 1.1 C _max ng/mL 970.0 1593.0 1.64 The results of pharmacokinetic experiments in rats showed that when chidamide was used in combination with the pharmaceutical excipient P-gp inhibitor hydroxypropyl β cyclodextrin, the blood drug exposure AUC increased to 1.1 times, and the C _max increased to 1.64 times that of the drug alone. It shows that pharmaceutical excipients P-gp inhibitors can also promote the absorption of chidamide, but the effect is insufficient compared with P-gp inhibitors such as verapamil and Tariquidar. Example 6: Tumor efficacy test of chidamide combined with P-gp inhibitor in mice

Forty female BALB/c mice aged 6-7 weeks were selected and randomly divided into 4 groups with 10 mice in each group. At the beginning of the experiment, each mouse axillary was inoculated subcutaneously with mouse intestinal cancer cell CT26, ⁵ *10 5 cells/mouse. ^{The administration was started when the tumor grew to 50 mm 3} on 7 days after inoculation. The single-agent chidamide (25mg/kg), chidamide combined with Tariquida, chidamide combined with verapamil, and chidamide were administered respectively. Amine combined with TPGS was continuously administered until 21 days after vaccination, during which the tumor growth of each group was measured every two days. The dosage and test results of each group are shown in Figure 5.

It can be seen from Figure 5 that the anti-tumor efficacy of chidamide combined with Tariquidar group is the best in each experimental group, followed by chidamide combined with verapamil group and chidamide combined with TPGS group. The efficacy is better than that of the chidamide single-agent group, indicating that the combined application of chidamide and P-gp inhibitor can effectively improve the anti-tumor efficacy of chidamide. Example 7: Mice tumor efficacy test of chidamide combined with P-gp inhibitor and immune checkpoint inhibitor

Sixty female BALB/c mice aged 6-7 weeks were selected and randomly divided into 6 groups with 10 mice in each group. At the beginning of the experiment, each mouse axillary was inoculated subcutaneously with mouse intestinal cancer cell CT26, ⁵ *10 5 cells/mouse. The administration was started 7 days after the inoculation when the tumor grew to 50 mm ³ , and the solvent control was given respectively. Chidamide (25 mg/kg, gavage, once a day), Tariquidar (10 mg/kg, gavage, once a day). Once), single-drug Anti-PD-1 antibody (RMP 1-14, BioXcell, 10mg/kg, intraperitoneal injection, twice a week) and Chidamide (25mg/kg, gavage, once a day) combined with Tariquidar (10mg /kg, gavage, once a day), and Chidamide (25mg/kg, gavage, once a day) combined with Tariquidar (10mg/kg, gavage, once a day) and Anti-PD-1 antibody (RMP 1- 14. BioXcell, 10mg/kg, intraperitoneal injection, twice a week), continuously administered for 15 days, during which the tumor growth of each group was measured every two days. The dosage and test results of each group are shown in Figure 6.

It can be seen from Figure 6 that compared with the single-drug test groups, the Chidamide combined with Tariquidar group showed good anti-tumor efficacy; and the most significant anti-tumor efficacy is Chidamide combined with Tariquidar and Anti-PD-1 antibody Group, indicating that the combined application of chidamide, P-gp inhibitors and immune checkpoint inhibitors can further effectively improve the anti-tumor efficacy of chidamide. Example 8: Chidamide combined with P-gp inhibitors and immune checkpoint inhibitors enhance the gene expression of cytokine TNF-α and IL-6

At the end of the experiment of Example 7 above, the spleens of mice in each experimental group were taken. A single spleen cell suspension is obtained through steps such as separation and removal of red blood cells. Take 5*10 ⁶ spleen cells from each group, and extract total RNA with TRIzol reagent (Invitrogen). The cDNA was obtained by reverse transcription using the Transcriptor First Strand cDNA Synthesis Kit (Roche), and then real-time quantitative PCR was performed on the ABI Prism 7000 PCR instrument using Roche's SYBR Green Master Dye and other reagents. The primer sequences of mouse cytokine IL-6 and TNF-α genes are respectively, IL-6 Forward: 5'-GGAGCCCACCAAGAACGATAG-3', IL-6 Reverse: 5'-GTGAAGTAGGG AAGGCCGTG-3'; TNF-α Forward: 5'-TGATCGGTCCCCAAAGGGAT-3', TNF-α Reverse: 5'-GCTACGACGTGGGCTACAGG-3'. The primer sequence of the mouse internal reference β-actin is β-actin Forward: 5'-GTCGAGTCGCGTCCACC-3', β-actin Reverse: 5'-ACGATGGAGGGGAATACAGC-3'. Figure 7 shows the relative expression results of splenic cytokine IL-6 and TNF-α genes in each group.

It can be seen from Figure 7 that compared with the single drug test groups, the Chidamide combined with Tariquidar group showed a certain effect of up-regulating the expression of IL-6 and TNF-α genes; while Chidamide combined with Tariquidar and Anti-PD-1 antibodies The group showed more significant up-regulation of the gene expression of these two cytokines, indicating that the combination of chidamide, P-gp inhibitors and immune checkpoint inhibitors can further promote the expression of immune cytokines to exert anti-tumor drugs. effect.

The above are only the preferred embodiments of the present invention. It should be pointed out that for those of ordinary skill in the art, without departing from the principle of the present invention, several improvements and modifications can be made. These improvements and modifications It should also be regarded as the protection scope of the present invention.

no.

Figure 1 shows the results of rat pharmacokinetic experiment of chidamide combined with verapamil; Figure 2 shows the results of the pharmacokinetic experiment of Chidamide combined with Tariquidar in rats; Figure 3 shows the results of the pharmacokinetic experiment of chidamide combined with TPGS in rats; Figure 4 shows the results of rat pharmacokinetic experiment of chidamide combined with hydroxypropyl β-cyclodextrin; Figure 5 shows the mouse tumor efficacy test of chidamide combined with P-gp inhibitor; Figure 6 shows the mouse tumor efficacy test of chidamide combined with P-gp inhibitor and immune checkpoint inhibitor; Figure 7 shows that Chidamide combined with P-gp inhibitors and immune checkpoint inhibitors enhance the gene expression of cytokine TNF-α and IL-6.

Claims (12)

A chidamide pharmaceutical composition, which is characterized by comprising chidamide and a P-glycoprotein inhibitor. The pharmaceutical composition according to claim 1, wherein the P-glycoprotein inhibitor is selected from the group consisting of artificially synthesized P-glycoprotein inhibitors, natural P-glycoprotein inhibitors, and pharmaceutical excipients P-glycoprotein inhibitors One or two or more of them. The pharmaceutical composition according to claim 1 or claim 2, wherein the P-glycoprotein inhibitor is selected from the group consisting of verapamil, cyclosporine, dexnicodipine, transfluthixol, tamoxifen One of Fen, Valspoda, Tariquidar, TPGS, β-cyclodextrin, PEG, Piperine, VX-710, Tetrandrine, FG020326, S-9788, GF-120918, LY-335979, DMDCK and MMDCK Or two or more. The pharmaceutical composition according to any one of claims 1 to 3, wherein the mass ratio of chidamide and P-glycoprotein inhibitor is 10:1 to 1:10. The pharmaceutical composition according to any one of claim 1 to claim 4, which further includes an immune checkpoint inhibitor. The pharmaceutical composition according to claim 5, wherein the immune checkpoint inhibitor is selected from the group consisting of anti-CTLA-4 antibody, anti-PD-1/PD-L1 antibody, anti-LAG-3 antibody, anti-TIM-3 antibody and anti- One or two or more of TIGIT antibodies. The pharmaceutical composition according to claim 6, wherein the anti-PD-1/PD-L1 antibody is selected from RMP 1-14, pembrolizumab, nivolumab, ztezolizumab, avelumab, durvalumab, tislelizumab, carrelizumab, and One or more of Riprolizumab, Sindizumab, Genostizumab, KN035, GLS-010 and CS1001. The pharmaceutical composition according to any one of claims 5 to 7, wherein the mass ratio of chidamide, P-glycoprotein inhibitor and immune checkpoint inhibitor is (1-10): (1- 10): (1-10). Use of the pharmaceutical composition according to any one of claim 1 to claim 8 in the preparation of chidamide preparations and/or drugs for treating diseases related to the action mechanism of histone deacetylase. The use according to claim 9, wherein the diseases related to the action mechanism of histone deacetylase include cancer, viral diseases, autoimmune diseases and blood system diseases. A chidamide preparation, which is characterized by comprising the chidamide pharmaceutical composition according to any one of claim 1 to claim 8. The preparation according to claim 11, wherein the dosage form of the preparation includes a tablet, a capsule, a pill, an oral liquid preparation (syrup), a granule, a powder, an ointment, and a dripping pill.

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