CN115554301A

CN115554301A - Use of an HDAC inhibitor and ibrutinib for the preparation of a medicament for the prevention or treatment of mantle cell lymphoma

Info

Publication number: CN115554301A
Application number: CN202211306941.9A
Authority: CN
Inventors: 吴文涛; 徐英霖
Original assignee: Xunuo Pharmaceutical Nanjing Co ltd
Current assignee: Xunuo Pharmaceutical Nanjing Co ltd
Priority date: 2022-10-24
Filing date: 2022-10-24
Publication date: 2023-01-03

Abstract

The invention provides an application of a pharmaceutical composition in preparing a medicine for preventing or treating mantle cell lymphoma, wherein the pharmaceutical composition comprises ibrutinib and an HDAC (HDAC) inhibitor, the HDAC inhibitor is selected from erbesat or salts thereof, and the mass ratio of the HDAC inhibitor to the ibrutinib is 1-3:1-10, experiments show that the pharmaceutical composition can significantly reduce the tumor volume and the relative tumor volume of mantle cell lymphoma, plays a synergistic effect, and has a good inhibitory effect on human mantle cell lymphoma Jeko-1.

Description

Use of an HDAC inhibitor and ibrutinib for the preparation of a medicament for the prevention or treatment of mantle cell lymphoma

Technical Field

The invention relates to the field of biomedical research, in particular to application of an HDAC (Histone deacetylase) inhibitor and ibrutinib in preparing a medicament for preventing or treating mantle cell lymphoma.

Background

Mantle Cell Lymphoma (MCL) is a special lymphoma subtype, and has the characteristics of refractory and invasive characteristics of indolent lymphoma, most of the patients are old, the median disease age is 58 years old, the proportion of men to women is about 2.

The clinical therapies for mantle cell lymphoma mainly include: anti-CD 20 monoclonal antibodies in combination with chemotherapy, hematopoietic stem cell transplantation, proteasome inhibitor Bortezomib, histone Deacetylase (HDAC) inhibitors, TOR inhibitors, and the like. N-hydroxy-4- {2- [3- (N, N-dimethylaminomethyl) benzofuran-2-ylcarbonylamino ] ethoxy } -benzamide (abeta) belongs to HDAC inhibitors and can act on HDAC1, 2, 3, 6 and 10 subtypes, and experiments show that the inhibitors can generate inhibition effects on tumors through various ways. However, for mantle cell lymphoma, no effective cure method exists so far, and particularly, the refractory relapsed patients lack effective treatment means, and the search for a new effective treatment method is particularly important.

Disclosure of Invention

Therefore, the present invention aims to provide a novel application of the HDAC inhibitor and ibrutinib, namely, the application of the HDAC inhibitor and ibrutinib in preparing a medicament for preventing or treating mantle cell lymphoma.

In order to solve the technical problems, the invention is specifically realized by the following technical scheme:

the invention provides an application of a pharmaceutical composition in preparing a medicine for preventing or treating mantle cell lymphoma, wherein the pharmaceutical composition comprises ibutinib and an HDAC (Histone citrate) inhibitor, the HDAC inhibitor is selected from erbestat or salt thereof, and the mass ratio of the HDAC inhibitor to the ibutinib is 1-3:1-10.

Further, the salt of ebactase is selected from the group consisting of a p-toluenesulfonate, a hydrochloride, a naphthalene-1, 5-disulfonate, a naphthalene-2-sulfonate, an oxalate, a benzenesulfonate, a sulfate or a phosphate of ebactase; and/or the mass ratio of the HDAC inhibitor to ibrutinib is 2.

Wherein, ebestat, the name Abexinostat of English has the following structural formula:

ibrutinib, the english name Ibrutinib, the structural formula of which is as follows:

further, the HDAC inhibitor and ibutinib are in the same formulation unit, or the HDAC inhibitor and ibutinib are in different formulation units, respectively.

Further, the HDAC inhibitor and a pharmaceutically acceptable carrier form a first formulation; the ibrutinib and a pharmaceutically acceptable carrier form a second formulation.

Those skilled in the art can select suitable excipients for the drugs to which the present application relates based on general knowledge in the pharmaceutical field. Such as, but not limited to, at least one of a pharmaceutically acceptable solvent, solubilizer, cosolvent, emulsifier, colorant, binder, disintegrant, filler, lubricant, wetting agent, osmotic pressure modifier, stabilizer, glidant, flavoring agent, preservative, suspending agent, coating material, anti-adherent, integrating agent, permeation enhancer, pH modifier, buffer, plasticizer, surfactant, thickener, encapsulating agent, humectant, absorbent, diluent, flocculant, deflocculant, filter aid, release retardant, polymeric matrix material, and film-forming material.

By "prevention" in this application is meant preventing the reoccurrence of the cured cancer or associated symptoms.

Further, the first preparation is an injection and the second preparation is an oral administration preparation. The dosage form of the present application may be selected from a variety of dosage forms known and studied in the art, including but not limited to oral dosage forms such as tablets, capsules, oral liquid formulations, pills, granules, powders; injection such as water injection, oil injection, emulsion injection, and powder for injection; and other dosage forms, such as topical dosage forms, e.g., ointments, patches, sprays, and the like; the methods and adjuvants required for the preparation of these dosage forms are available in the relevant books and literature of pharmaceutics.

Further, the solvent of the first preparation is an aqueous solution containing 15-25wt% of hydroxypropyl betacyclodextrin, and the solvent of the second preparation is an aqueous solution containing 0.3-0.8wt% of methylcellulose.

Further, the mantle cell lymphoma is human mantle cell lymphoma Jeko-1.

The pharmaceutical compositions and methods of the present application may be used in combination with other known and studied drugs or methods of treatment or adjuvant treatment of cancer, including, but not limited to, chemotherapy and corresponding chemotherapeutic agents, radiotherapy, immunotherapy methods and corresponding drugs, herbal medicines, and the like.

The compositions of the present application can be used in a variety of animals including, but not limited to, primate mammals, murine, bovine, equine, canine, chicken, duck, goose species, and the like, with primate mammals being preferred, and human families being particularly preferred.

The invention also provides a medicine for preventing or treating mantle cell lymphoma, which comprises the following components in a mass ratio of 1-3:1-10 and ibrutinib, said HDAC inhibitor selected from abemostat or a salt thereof.

Further, the salt of ibesistat is selected from the group consisting of ibesistat tosylate, hydrochloride, naphthalene-1, 5-disulfonate, naphthalene-2-sulfonate, oxalate, benzenesulfonate, sulfate, or phosphate; and/or the mass ratio of the HDAC inhibitor to ibrutinib is 2.

Further, the HDAC inhibitor is ebesistat p-toluenesulfonate.

Further, the HDAC inhibitor and a pharmaceutically acceptable carrier form a first formulation; the HDAC inhibitor and a pharmaceutically acceptable carrier form a second formulation.

The technical scheme of the invention has the following advantages:

the invention combines an HDAC inhibitor selected from erbestat or salt thereof and ibrutinib, and controls the mass ratio of the HDAC inhibitor to the ibrutinib to be 1-3:1-10, can obviously reduce the tumor volume and the relative tumor volume of mantle cell lymphoma, plays a synergistic effect and has good inhibition effect on human mantle cell lymphoma Jeko-1.

Drawings

In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings used in the description of the embodiments or the prior art will be briefly described below, and it is obvious that the drawings in the following description are some embodiments of the present invention, and other drawings can be obtained by those skilled in the art without creative efforts.

FIG. 1 is a graph showing the average body weight change of each group in example 1;

FIG. 2 is a graph showing the change in tumor volume in each group in example 1;

FIG. 3 is a graph showing the relative change in tumor volume of each group in example 1;

FIG. 4 shows the tumor weights of the groups in example 1;

FIG. 5 is a graph showing the change in tumor volume in each group in example 2;

FIG. 6 is a graph showing the relative tumor volume changes of the groups in example 2;

FIG. 7 shows the weights of the tumors in each group in example 2.

Detailed Description

The following examples are provided to further understand the present invention, not to limit the scope of the present invention, but to provide the best mode, not to limit the content and the protection scope of the present invention, and any product similar or similar to the present invention, which is obtained by combining the present invention with other prior art features, falls within the protection scope of the present invention. The examples do not indicate specific experimental procedures or conditions, and can be performed according to the procedures or conditions of the conventional experimental procedures described in the literature in the field. The reagents or instruments used are not indicated by manufacturers, and are all conventional reagent products which can be obtained commercially.

1. Experimental drugs, main reagents:

experimental drug XP101 was provided by yono pharmaceutical (tokyo) ltd, powder, lot number: 877721/001, the basic information is as follows:

the molecular formula is as follows: c ₂₁ H ₂₃ N ₃ O ₅ ,C ₇ H ₈ O ₃ S

Molecular weight: 569.6 (397.431+172.205)

Chiral/stereochemistry: non-asymmetric carbon

Base conversion factor: 0.698.

experimental drug ibutinib was purchased from MCE, lot No.: HY-10997.

Other reagents:

reagent	Company(s)	Goods number
			MEM medium	Corning cellgro	10-010-CV
Fetal Bovine Serum (FBS)	GIBCO	10099-141
			Green streptomycin (P.S.)	GIBCO	15140-122
Trypsin-ethylenediamine tetraacetic acid (EDTA)Acid(s)	GIBCO	25200-072
			PBS	Corning cellgro	21-040-CVR
Matrigel	Corning cellgro	354234

2. The main apparatus is as follows:

instrument for measuring the position of a moving object	Company(s)	Model number
			Centrifugal machine	Saimer Feishale	Thermo Fisher legend MACH 1.6/
Inverted microscope	Nikang medicine	TS100
			Biological safety cabinet (cell culture)	Saimer Feishale	1379
CO ₂ Culture box	Saimer Feishale	Thermo HERA cell 150
			Water bath pot	Shanghai essence macro	DK-8B
Liquid nitrogen tank	Saimer Feishale	Thermo LOCATOR 4
			Pipettor (electric) (cell culture)	Capp	PA-100
Pipettor (1 ml) (cell culture)	Aibende	L20989H
			Pipettor (100. Mu.l) (cell culture)	Aibende	L38752H
Pipettor (20. Mu.l) (cell culture)	Aibende	N22990H
			Anesthesia machine	MATRX	MATRX VIP3000
Biological safety cabinet (cell inoculation)	Saimei FeishiEr (Chinese character)	Thermo MSC-Advantage
			Biological safety cabinet (dispensing)	Saimei Feishire	Thermo MSC-Advantage
Balance (animal weight)	Denver, USA	DENVER INSTRUMENT TP-602
			Vernier caliper	Japan Sanfeng	mitutoyo CD-15cX

Example 1

1. Preparation of test drugs

The solvent for ibrutinib was an aqueous solution containing 0.5wt% of methylcellulose (hereinafter simply referred to as "0.5wt% mc solution").

Table 1 formulation of the test drugs

2. Animal grouping, modeling and administration

(1) Animal(s) production

BALB/c nude mice, no history of administration, female, 6-7 weeks old, 32 out of them, 8 were ready for use, purchased from shanghai jihui experimental animals feeding limited, acclimation period 3-7 days, room in SPF area, indoor temperature 20-26 ℃, relative humidity 40-70%, fluorescent lamp lighting, 12 hour lighting (08-00) and 12 hour no lighting, 2-6 per cage (same administration group), unlimited access to feed (sterilized by irradiation, jiangsu province cooperative medical biotechnology limited, china), unlimited access to drinking water (tap water treated by reverse osmosis or autoclaving).

(2) Cell culture

Human mantle cell lymphoma Jeko-1 (purchased from ATCC) was cultured in MEM medium supplemented with 10% FBS in a medium containing 5% CO ₂ 37 ℃ incubator. Approximately 5X 10 cells were cultured for ten successive passages ⁶ Jeko-1 cells were resuspended in PBS, mixed with an equal volume of Matrigel, and then subcutaneously inoculated into 32 BALB/c nude mice in a volume of 200. Mu.L.

(3) Grouping and administration.

When the mean tumor volume reached about 150mm ³ In the meantime, 24 tumor-bearing mice were randomly divided into 4 groups of 6 mice each according to tumor volume and body weight, and the daily divided groups were defined as Day 0, and administration was started on the respective days, and the specific administration schedule was as shown in the following table:

TABLE 2 dosing regimen

Wherein, the administration volume is calculated according to the volume of the experimental animal by 10 mul/g, QD 21Days represents once daily administration for 21 Days.

3. Detecting the index

(1) Animal Body weight (BW, body weight)

Mice body weights were measured and recorded twice weekly after grouping.

(2) Tumor volume (TV, tumor volume)

Tumor volumes were measured twice weekly for 4 consecutive weeks after grouping. Tumor Volume (TV) was calculated as follows: TV = (length × width) ² )/2。

(3) Tumor inhibition rate (TGI, tumor growth inhibition)

TGI% = (1-T/C) × 100%. Where T and C are the Relative Tumor Volumes (RTV) at a particular time point for the test group (G2, G3 or G4 group) and vehicle control group (G1 group), respectively.

4. Standard of experimental withdrawal

During the course of the experiment, mice were discontinued following the following principle: when the weight loss is more than or equal to 10 percent, the administration is stopped, and the drug stopping period is long enough to recover the weight of the mouse. No discontinuation occurred in the mice in this experiment.

5. End point of sidewalk

During the experiment, the animals should be euthanized if any one or more of the following occurs:

1) When the average tumor volume in the group exceeds 2000mm ³ ；

2) Tumors develop ulcerations, necrosis or infection;

3) Abnormal movement or paralysis of the animal;

4) The animal's body weight decreased by more than 20% of the body weight at the time of starting the drug treatment for three consecutive days.

5) Other disease symptoms

No mice in this experiment were euthanized for the reasons described above. No animals died midway or were euthanized in this experiment.

6. Statistical analysis

The results will be presented as mean ± s.e.m. The comparison between the two groups will be checked with Dunnett's multi-contrast test. A statistically significant difference is considered if p < 0.05.

7. Results of the experiment

(1) Body weight

TABLE 3 mouse body weight (g)

The body weight of the G1 group continued to increase during the experiment. Compared with the group G1, the body weight of the whole experiment of the group G2 has no obvious difference with the body weight of the group G1; the weight of the G3 group is significantly less than that of G1 at Day 3, day 8-10, day 17, day 23-28; the body weight of the G4 group was significantly less than that of G1 at Day 8-10 and Day 23-28.

(2) Tumor Volume (TV) and Relative Tumor Volume (RTV)

The tumor volume of G1 is continuously increased in the experimental process, so that the subcutaneous transplantation tumor model of human mantle cell lymphoma Jeko-1 on BALB/c nude mice can be successfully established. The tumor volume of G1 was 148.62. + -. 1.62mm at Day 0 ³ 2223.87. + -. 224.52mm at Day28 ³ (ii) a The relative tumor volumes were 1.58. + -. 0.11 at Day 3 and 15.01. + -. 1.55 at Day 28.

Compared to the G1 group, the tumor volumes and relative tumor volumes of G2, G3, and G4 were significantly reduced during Day 3-28.

The tumor volumes of the groups are shown in table 4 and fig. 2, and the relative tumor volumes are shown in table 5 and fig. 3.

TABLE 4 tumor volume (mm) ³ )

TABLE 5 relative tumor volume

(3) Tumor weight

At the end of Day28 in vivo experiments, tumors were removed from euthanized animals and weighed, and G2, G3, and G4 tumors were significantly reduced in weight compared to the G1 control group. The tumor weight statistics for each group are shown in table 6 and figure 4.

TABLE 6 tumor weight (g)

(4) Relative tumor inhibition (TGI)

TABLE 7 relative tumor inhibition (%)

The animals have no obvious toxic effect and weight change under different dosages of the ibrutinib, 10mg/kg, 30mg/kg and 90mg/kg, and the tumor inhibition effect is in a dose-dependent relationship.

Example 2

1. Preparation of test drugs

The solvent for XP101 was a 20wt% aqueous solution of hydroxypropyl betacyclodextrin (HP- β -CD) (hereinafter referred to as "20wt% HP- β -CD solution").

The solvent for ibrutinib was an aqueous solution containing 0.5wt% of Methylcellulose (MC) (hereinafter referred to as "0.5wt% MC solution").

Table 8 formulation of the test drugs

2. Animal grouping, modeling and administration

(1) Animal(s) production

BALB/c nude mice, no drug history, female, 6-7 weeks old, 52 out of 12 for standby, purchased from Shanghai Jihui laboratory animal feeding Co., ltd, adaptation period 7 days, room in SPF district, indoor temperature 20-26 ℃, relative humidity 40-70%, fluorescent lamp lighting, 12 hour lighting (08 00-20) and 12 hour no lighting, 2-6 per cage (same drug group), and obtaining feed without limitation (sterilized by irradiation, jiangsu province cooperative medical bioengineering Co., ltd., china), and drinking water (tap water treated by reverse osmosis or autoclaving).

(2) Cell culture

Human mantle cell lymphoma Jeko-1 was cultured in MEM medium supplemented with 10% FBS in a medium containing 5% CO ₂ 37 ℃ incubator. Approximately 5X 10 cells were cultured for ten successive passages ⁶ Jeko-1 cells were resuspended in PBS, mixed with an equal volume of Matrigel, and then subcutaneously inoculated into 52 BALB/c nude mice in a volume of 200. Mu.L.

(3) Grouping and administration.

When the mean tumor volume reached about 148mm ³ In this case, 40 tumor-bearing mice were divided into 4 groups of 10 mice each at random according to the tumor volume and body weight, and the Day of each group was defined as Day 0, and administration was started on each Day, and the specific administration schedule was as shown in the following table:

TABLE 9 dosing regimen

Wherein the administration volume is calculated as 10. Mu.l/g based on the volume of the experimental animal. QD 28Days indicates once daily dosing for 28 consecutive Days. 4h BID 4days/wk means 2 doses per day (BID) with 4 hours interval between two doses, 4days per week. 4h BID 2days/wk means 2 doses per day (BID) with 4 hours intervals between two doses, 2days per week.

3. Detecting the index

(1) Animal Body weight (BW, body weight)

Mice body weights were measured and recorded twice a week after the grouping.

(2) Tumor volume (TV, tumor volume)

(3) Tumor inhibition rate (TGI, tumor growth inhibition)

TGI% = (1-T/C) × 100%. Where T and C are the Relative Tumor Volumes (RTV) of the test and vehicle control groups, respectively, at a particular time point.

4. Medicine for experiment

G3-1, G3-3, G3-6, G3-8, G4-1, G4-2, G4-4, G4-5, G4-7, G4-9 and G4-10 were discontinued at Day 8 due to a weight loss of 10% or more; g4-4 and G4-10 stopped the drug at Day24 due to a weight loss of 10% or more.

5. End point of sidewalk

1) When the average tumor volume in the group exceeds 2000mm ³ ；

2) Ulceration, necrosis or infection of the tumor;

3) Abnormal movement or paralysis of the animal;

5) Other disease symptoms

6. Statistical analysis

7. Results of the experiment

(1) Tumor Volume (TV) and Relative Tumor Volume (RTV)

The tumor volume of G1 continuously increased during the experiment, representing the successful model construction of human mantle cell lymphoma Jeko-1 in BALB/c nude mice. The tumor volume of G1 was 148.62. + -. 1.62mm at Day 0 ³ 2223.87. + -. 224.52mm at Day28 ³ (ii) a The relative tumor volume of G1 was 1.58. + -. 0.11 at Day 3 and 15.01. + -. 1.55 at Day 28.

Compared to the G1 group, the tumor volumes and relative tumor volumes of the G2, G3, and G4 groups were significantly reduced during Day 3-Day 28.

Compared to G2, the tumor volume and relative tumor volume of G4 decreased significantly between Day7-17 and Day 24-28.

TABLE 10 tumor volume (mm) ³ )

Wherein, P Value is G2, G3 or G4 VS G1, and P Value # is G2 VS G4.

TABLE 11 relative tumor volumes

Wherein, P Value is G2, G3 or G4 VS G1, and P Value # is G2 VS G4.

(2) Tumor weight

At the end of the Day28 in vivo experiment, the animals were euthanized and tumors were weighed. The tumor weights were significantly reduced in the G2, G3 and G4 groups compared to the G1 control group. The tumor weight was significantly reduced in the G4 group compared to G2.

TABLE 12 tumor weight (g)

Wherein, P Value is G2, G3 or G4 VS G1, and P Value # is G2 VS G4.

(3) Relative tumor inhibition Rate (TGI)

TABLE 13 relative tumor inhibition (%)

Wherein, the TGI is G2, G3 or G4 VS G1, and the TGI is G2 VS G4.

Compared with the group with the single ibutinib QD administration of 90mg/kg, the combined administration group has the advantage that the G4 relative tumor inhibition rate is obviously improved.

8. Conclusion of the experiment

Compared with the group with 90mg/kg QD single-dose ibrutinib, the combined-dose group G4 has significantly reduced average tumor volume and relative tumor volume at Day7-17 and Day24-28, and shows that the synergistic anti-tumor effect of ibrutinib and XP101 is generated.

It should be understood that the above examples are only for clarity of illustration and are not intended to limit the embodiments. Other variations and modifications will be apparent to persons skilled in the art in light of the above description. And are neither required nor exhaustive of all embodiments. And obvious variations or modifications therefrom are within the scope of the invention.

Claims

1. Use of a pharmaceutical composition in the preparation of a medicament for preventing or treating mantle cell lymphoma, wherein the pharmaceutical composition comprises ibutinib and an HDAC inhibitor, the HDAC inhibitor is selected from erbestat or a salt thereof, and the mass ratio of the HDAC inhibitor to the ibutinib is 1-3:1-10.

2. Use according to claim 1, characterized in that the salt of ebactase is selected from the group consisting of the p-toluenesulfonate, hydrochloride, naphthalene-1, 5-disulfonate, naphthalene-2-sulfonate, oxalate, benzenesulfonate, sulfate or phosphate salt of ebactase; and/or the mass ratio of the HDAC inhibitor to ibrutinib is 2.

3. The use according to claim 1, wherein the HDAC inhibitor and ibrutinib are in the same formulation unit or the HDAC inhibitor and ibrutinib are in separate formulation units.

4. The use according to claim 3, wherein the HDAC inhibitor and the pharmaceutically acceptable carrier form a first formulation; the ibrutinib and a pharmaceutically acceptable carrier form a second formulation.

5. The use according to claim 4, wherein the pharmaceutically acceptable carrier is selected from at least one of pharmaceutically acceptable solvents, solubilizers, emulsifiers, colorants, binders, disintegrants, fillers, lubricants, wetting agents, tonicity adjusting agents, stabilizers, glidants, flavoring agents, preservatives, suspending agents, coating materials, anti-adherents, integration agents, penetration enhancers, pH adjusting agents, buffers, plasticizers, surfactants, thickeners, encapsulation agents, humectants, absorbents, diluents, flocculants, deflocculants, filter aids, release retardants, polymeric matrix materials, and film-forming materials.

6. The use according to claim 4, wherein the first formulation is an injectable formulation and the second formulation is an orally administrable formulation.

7. The use according to claim 4, wherein the solvent of the first formulation is an aqueous solution containing 15-25wt% hydroxypropyl betacyclodextrin and the solvent of the second formulation is an aqueous solution containing 0.3-0.8wt% methylcellulose.

8. Use according to any one of claims 1 to 7, wherein the mantle cell lymphoma is human mantle cell lymphoma Jeko-1.

9. The medicine for preventing or treating mantle cell lymphoma is characterized by comprising the following components in a mass ratio of 1-3:1-10 and ibrutinib, said HDAC inhibitor selected from abemostat or a salt thereof.

10. The agent for the prevention or treatment of mantle cell lymphoma according to claim 9, wherein said HDAC inhibitor and a pharmaceutically acceptable carrier form a first formulation; the HDAC inhibitor and a pharmaceutically acceptable carrier form a second formulation.