A combined pharmaceutical composition and use thereof in the preparation of a medicament for preventing, ameliorating or treating TP53-mutated double hit lymphoma

Abstract

The present application relates to a kind of combined pharmaceutical composition and its application in the preparation of preventing, improving or treating TP53 mutant double-hit lymphoma, the active component of the described pharmaceutical composition is composed of first active component and second active component;The first active component is selected from any one or combination of at least two of cedabanum or its pharmaceutically acceptable salt, isomer, solvate, metabolite;The second active component is selected from any one or combination of at least two of anlotinib or its pharmaceutically acceptable salt, isomer, solvate, metabolite.The combined pharmaceutical composition has excellent TP53 mutant double-hit lymphoma prevention, improvement or treatment effect, when two active components are used in combination, not only can the dosage be reduced, safety is improved, but also has more significant inhibitory effect than single active component, reaches the effect of synergistic promotion.

Description

Technical Field

[0001] This invention belongs to the field of biomedical technology and relates to a novel combination drug composition and its application in the preparation of drugs for the prevention, improvement or treatment of TP53-mutant double-hit lymphoma. Background Technology

[0002] Double-hit lymphoma (DHL) is a heterogeneous disease between diffuse large B-cell lymphoma (DLBCL) and Burkitt lymphoma (BL). It often presents as a fusion pattern of BL and DLBCL and is a high-grade B-cell lymphoma with MYC gene translocation, BCL2 or BCL6 rearrangement.

[0003] Double-hit lymphoma (DHL) is difficult to treat and is resistant to standard treatment regimens. Current first-line immunotherapy and chemotherapy regimens have high failure rates. Furthermore, DHL with TP53 mutations can lead to complex karyotypes, further complicating treatment. Therefore, there is an urgent need to develop new treatment strategies to improve the efficacy of DHL treatment.

[0004] Chidamide is a novel epigenetic regulator that reactivates the tumor-suppressive immune microenvironment and can be used alone or in combination with other drugs to address serious diseases such as malignant tumors that severely threaten human health. Studies have shown that chidamide has demonstrated some therapeutic efficacy in the clinical treatment of diffuse large B-cell lymphoma, and it also shows some efficacy in relapsed / refractory DLBCL, even TP53-mutant rituximab-treated relapsed DLBCL and triple-hit lymphoma, although some patients still do not respond well to it. Anlotinib, a Class 1.1 anti-tumor drug independently developed by Chia Tai Tianqing Pharmaceutical Group, can effectively inhibit signaling pathways such as VEGFR, PDGFR, FGFR, and c-Kit, and has anti-tumor angiogenesis and tumor growth-inhibiting effects.

[0005] Whether the combination of chidamide and anlotinib can be used to treat TP53-mutant double-hit lymphoma, and the mechanism of action of chidamide in TP53-mutant double-hit lymphoma, remains unclear. Summary of the Invention

[0006] In view of the shortcomings of the prior art, the purpose of this invention is to provide a new combination pharmaceutical composition and its application in the preparation of a medicament for the prevention, improvement or treatment of TP53-mutant double-hit lymphoma.

[0007] To achieve this objective, the present invention employs the following technical solution:

[0008] In a first aspect, the present invention provides a combination pharmaceutical composition, wherein the active component of the pharmaceutical composition is composed of a first active component and a second active component;

[0009] The first active component is selected from any one or a combination of at least two of the following: chidamide or its pharmaceutically acceptable salts, isomers, solvates, and metabolites;

[0010] The second active ingredient is selected from any one or a combination of at least two of anlotinib or its pharmaceutically acceptable salts, isomers, solvates, and metabolites.

[0011] This invention develops a novel drug combination therapy, which combines chidamide (or its pharmaceutically acceptable salts, isomers, solvates, or metabolites) and anlotinib (or its pharmaceutically acceptable salts, isomers, solvates, or metabolites). This combined drug composition exhibits excellent improvement or therapeutic effects in TP53-mutant double-hit lymphoma. The invention's research found that the two active components, when inhibiting TP53-mutant double-hit lymphoma, not only reduce the dosage of chidamide (or its pharmaceutically acceptable salts, isomers, solvates, or metabolites) or anlotinib (or its pharmaceutically acceptable salts, isomers, solvates, or metabolites), improving drug safety, but also have a more significant inhibitory effect on TP53-mutant double-hit lymphoma than using a single active component, achieving a synergistic effect.

[0012] The TP53 mentioned refers to the TP53 gene, a tumor suppressor gene that encodes a 53 kDa protein. This protein is expressed at low levels in normal cells but at high levels in malignant tumors. The P53 protein, translated from the TP53 gene, is an important regulator of cell growth, proliferation, and damage repair. When cellular DNA is damaged, the P53 protein can arrest the cell in the G1 / S phase to repair the damage; if repair fails, it promotes apoptosis.

[0013] This invention first demonstrates, using TP53-mutant double-hit lymphoma cell lines (TMD8 and Toledo cells), that compared to the single-agent groups of chidamide and anlotinib, the combined drug composition significantly inhibits the proliferation of TP53-mutant double-hit lymphoma cells, induces apoptosis, and triggers cell cycle arrest. Western blot experiments show that the combined drug composition inhibits the proliferation of TP53-mutant double-hit lymphoma cell lines and promotes apoptosis and cell cycle arrest by inducing downregulation of PI3K / AKT signaling pathway molecules in TP53-mutant DHL cells. Furthermore, by constructing a cell-derived xenograft (CDX) mouse model, this combined drug composition reduces the tumor burden in CDX mice. This invention provides an effective drug combination strategy for the prevention, improvement, or treatment of TP53-mutant double-hit lymphoma, and is of significant importance.

[0014] In this invention, the combined pharmaceutical composition is a single compound preparation or a combination of two separate preparations.

[0015] Preferably, the combined pharmaceutical composition is a combination of two separate formulations, which are administered simultaneously or sequentially.

[0016] The combined drug composition can be a single compound preparation or a combination of two separate preparations; when it is a combination of two separate preparations, it can be administered simultaneously, alternately, or sequentially.

[0017] In this invention, the formulation is any pharmaceutically acceptable dosage form.

[0018] Preferably, the combined pharmaceutical composition further contains pharmaceutically acceptable excipients.

[0019] Preferably, the pharmaceutically acceptable excipients include any one or a combination of at least two of the following: carrier, diluent, binder, wetting agent, disintegrant, emulsifier, cosolvent, solubilizer, osmotic pressure regulator, surfactant, coating material, colorant, pH adjuster, antioxidant, antibacterial agent, or buffer.

[0020] In a second aspect, the present invention provides the use of the combined pharmaceutical composition according to the first aspect in the preparation of a medicament for the prevention, improvement or treatment of TP53-mutant double-hit lymphoma.

[0021] This invention develops a novel approach for the prevention, improvement, or treatment of TP53-mutant double-hit lymphoma, which involves the combined use of chidamide (or its pharmaceutically acceptable salts, isomers, solvates, or metabolites) and anlotinib (or its pharmaceutically acceptable salts, isomers, solvates, or metabolites). This combined drug composition exhibits excellent efficacy in improving or treating TP53-mutant double-hit lymphoma. The invention's research found that the two active components, when inhibiting TP53-mutant double-hit lymphoma, not only reduce the dosage of chidamide (or its pharmaceutically acceptable salts, isomers, solvates, or metabolites) or anlotinib (or its pharmaceutically acceptable salts, isomers, solvates, or metabolites), improving drug safety, but also significantly inhibit TP53-mutant double-hit lymphoma compared to using a single active component, achieving a synergistic effect.

[0022] Thirdly, the present invention provides the use of the combined pharmaceutical composition according to the first aspect in the preparation of a TP53-mutant double-hit lymphoma cell proliferation inhibitor.

[0023] According to the research results of this invention, the combined pharmaceutical composition significantly inhibits the proliferation of TP53-mutant double-hit lymphoma cells. Therefore, this result indicates that the combined pharmaceutical composition can be used as an in vitro experimental reagent in scientific research, such as to study the growth, apoptosis, and metabolic mechanisms or behavior of TP53-mutant double-hit lymphoma cells, and to screen therapeutic drugs. The proliferation inhibitor claimed in this invention is not intended to eliminate the cause or lesion; that is, it is an application in the preparation of a TP53-mutant double-hit lymphoma cell proliferation inhibitor for a non-therapeutic purpose.

[0024] Preferably, the TP53-mutated double-hit lymphoma cells include the TMD8 cell line and / or the Toledo cell line.

[0025] Fourthly, the present invention provides the use of the combined pharmaceutical composition according to the first aspect in the preparation of an apoptosis promoter for TP53-mutant double-hit lymphoma cells.

[0026] According to the research results of this invention, the combined pharmaceutical composition significantly induces apoptosis in TP53-mutant double-hit lymphoma cells. Therefore, this result indicates that the combined pharmaceutical composition can be used as an in vitro experimental reagent in scientific research, such as to study the growth, apoptosis, and metabolic mechanisms or behaviors of TP53-mutant double-hit lymphoma cells, and to screen therapeutic drugs. The apoptosis promoter claimed in this invention is not intended to eliminate the cause or lesion; that is, it is an application in the preparation of an apoptosis promoter for TP53-mutant double-hit lymphoma cells for a non-therapeutic purpose.

[0027] Preferably, the TP53-mutated double-hit lymphoma cells include the TMD8 cell line and / or the Toledo cell line.

[0028] Fifthly, the present invention provides the use of the combined pharmaceutical composition according to the first aspect in the preparation of a cell cycle arrestor for TP53-mutant double-hit lymphoma.

[0029] According to the research results of this invention, the combined pharmaceutical composition has a significant effect in inducing cell cycle arrest in TP53-mutant double-hit lymphoma cells. Therefore, this result indicates that the combined pharmaceutical composition can be used as an in vitro experimental reagent in scientific research, such as to study the growth, apoptosis, and metabolic mechanisms or behaviors of TP53-mutant double-hit lymphoma cells, and to screen therapeutic drugs. The cell cycle arrestor claimed in this invention is not intended to eliminate the cause or lesion; that is, it is an application in the preparation of a TP53-mutant double-hit lymphoma cell cycle arrestor for a non-therapeutic purpose.

[0030] Preferably, the TP53-mutated double-hit lymphoma cells include the TMD8 cell line and / or the Toledo cell line.

[0031] In a sixth aspect, the present invention provides the use of the combined pharmaceutical composition according to the first aspect in the preparation of an inhibitor of the PI3K / AKT signaling pathway in TP53-mutant double-hit lymphoma cells.

[0032] According to the research results of the present invention, the combined drug composition has a significant inhibitory effect on the PI3K / AKT signaling pathway in TP53-mutant double-hit lymphoma cells. Therefore, this result indicates that the combined drug composition can be used as an in vitro experimental reagent in the field of scientific research, such as to study the growth, apoptosis and metabolic mechanisms or behavior of TP53-mutant double-hit lymphoma cells, and to screen drugs for the treatment of TP53-mutant double-hit lymphoma.

[0033] Compared with the prior art, the present invention has the following beneficial effects:

[0034] This invention develops a novel drug combination therapy, which combines chidamide (or its pharmaceutically acceptable salts, isomers, solvates, or metabolites) and anlotinib (or its pharmaceutically acceptable salts, isomers, solvates, or metabolites). This combined drug composition exhibits excellent improvement or therapeutic effects in TP53-mutant double-hit lymphoma. The invention's research found that when inhibiting TP53-mutant double-hit lymphoma, the two active components not only reduce the dosage of chidamide (or its pharmaceutically acceptable salts, isomers, solvates, or metabolites) or anlotinib (or its pharmaceutically acceptable salts, isomers, solvates, or metabolites), improving drug safety, but also have a more significant inhibitory effect on TP53-mutant double-hit lymphoma than using a single active component, achieving a synergistic effect. Attached Figure Description

[0035] Figure 1A This is a graph showing the statistical results of cell proliferation inhibition rate after 24 hours of drug treatment of TMD8 cells in Example 1;

[0036] Figure 1B This is a graph showing the statistical results of the cell proliferation inhibition rate of Toledo cells after 24 hours of drug treatment in Example 1;

[0037] Figure 2A This is a flow cytometry result of TMD8 cells treated with the drug for 24 hours in Example 2;

[0038] Figure 2B This is a graph showing the statistical results of the apoptosis rate of TMD8 cells treated with the drug for 24 hours in Example 2;

[0039] Figure 2C This is a flow cytometry result of Toledo cells treated with the drug for 24 hours in Example 2;

[0040] Figure 2D This is a graph showing the statistical results of the apoptosis rate of Toledo cells treated with the drug for 24 hours in Example 2;

[0041] Figure 3A This is a graph showing the percentage of cells in each cell cycle after 24 hours of drug treatment of TMD8 cells in Example 3.

[0042] Figure 3B This is a graph showing the percentage of cells in each cell cycle after 24 hours of drug treatment of Toledo cells in Example 3.

[0043] Figure 4 This is a Western blot result of the expression levels of PI3K / AKT pathway-related proteins in TMD8 or Toledo cells after 24 hours of drug treatment in Example 4.

[0044] Figure 5A This is a graph showing the changes in tumor volume after drug treatment in a tumor-bearing mouse model in Example 5;

[0045] Figure 5B This is a graph showing the changes in tumor weight after drug treatment in a tumor-bearing mouse model in Example 5;

[0046] Figure 5C This is an anatomical diagram of the tumor in the tumor-bearing mouse model after drug treatment in Example 5;

[0047] Figure 5D This is a Western blot result of the expression level of PI3K / AKT pathway-related proteins in tumor tissue after drug treatment of the tumor-bearing mouse model in Example 5. Detailed Implementation

[0048] The technical solution of the present invention will be further illustrated below through specific embodiments. Those skilled in the art should understand that the embodiments described are merely illustrative of the present invention and should not be construed as limiting the invention in any way.

[0049] The processes, conditions, reagents, and experimental methods used in implementing this invention, except as specifically mentioned below, are all common knowledge and general knowledge in the field, and this invention does not have any particular limitations. Experimental methods in the embodiments that do not specify specific conditions are generally performed under conventional conditions or as recommended by the manufacturer.

[0050] Unless otherwise stated, all technical terms and scientific terms used in this specification have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains. However, in the event of any conflict, the specification containing the definitions shall prevail.

[0051] The drug Chidamide involved in the following examples was provided by Shenzhen Chipscreen Biosciences Co., Ltd.; the drug Anlotinib was provided by Chia Tai Tianqing Pharmaceutical Group Co., Ltd.

[0052] The TP53-mutated double-hit lymphoma cell lines (TMD8 and Toledo cells) were provided by the Institute of Hematology, School of Medicine, Xiamen University.

[0053] CB17-SCID mice were purchased from Beijing Vital River Laboratory Animal Technology Co., Ltd. and bred by the Laboratory Animal Center of Xiamen University.

[0054] The experimental results were statistically analyzed using GraphPad Prism 8.0. * indicates p < 0.05, ** indicates p < 0.01, *** indicates p < 0.001, and **** indicates p < 0.0001.

[0055] Example 1

[0056] The inhibitory effect of the combined drug combination on the proliferation of TP53-mutant double-hit lymphoma cell lines:

[0057] The operation method is as follows: Take a quantity of 2 × 10 5 Log-growing TP53-mutated double-hit lymphoma cell lines (TMD8 and Toledo cell lines) were seeded in 96-well plates, with chidamide monotherapy group, anlotinib monotherapy group, chidamide and anlotinib combination group, and control group set up.

[0058] The concentrations of chidamide monotherapy were 0 μM, 1 μM, 2 μM, 4 μM, 8 μM, and 16 μM, respectively. The concentration of anlotinib monotherapy was 10 μM. The concentrations of chidamide and anlotinib combination therapy were 0 μM+10 μM, 1 μM+10 μM, 2 μM+10 μM, 4 μM+10 μM, 8 μM+10 μM, and 16 μM+10 μM, respectively (the former being chidamide and the latter anlotinib). The control group received the same volume of DMSO. DMSO or the drug was added to the cells in 96-well cell culture plates and gently shaken to mix. After culturing at 37°C for 24 h, cell proliferation was assessed using a CCK8 assay kit.

[0059] Experimental results:

[0060] The cell proliferation levels of each group are as follows: Figure 1A (TMD8) and Figure 1B(Toledo) is shown. (Through) Figure 1A , Figure 1B The results showed that, compared with the chidamide monotherapy group and the anlotinib monotherapy group, the chidamide and anlotinib combination group exhibited a more significant inhibitory effect on the proliferation of TP53-mutant double-hit lymphoma cell lines TMD8 and Toledo, indicating the synergistic promoting effect of the two drugs.

[0061] Example 2

[0062] The apoptosis-inducing effect of the combination drug regimen on TP53-mutant double-hit lymphoma cell lines:

[0063] The operation method is as follows: Take a quantity of 2 × 10 5 Log-phase TP53-mutated double-hit lymphoma cell lines (TMD8 and Toledo) were seeded in 96-well plates, with chidamide monotherapy group, anlotinib monotherapy group, chidamide and anlotinib combination group, and control group set up.

[0064] In the TMD8 cell line, the administration concentrations of chidamide monotherapy were 0 nM, 375 nM, and 750 nM; the administration concentration of anlotinib monotherapy was 1.6 μM; and the administration concentrations of the combination of chidamide and anlotinib were 0 nM + 1.6 μM, 375 nM + 1.6 μM, and 750 nM + 1.6 μM (the former being chidamide and the latter anlotinib). In the Toledo cell line, the administration concentration of chidamide monotherapy was... The concentrations of the drugs were 0 μM, 3 μM, 6 μM, 12 μM, and 24 μM, respectively. The concentration of anlotinib monotherapy was 2.5 μM, and the concentrations of the combination of chidamide and anlotinib were 0 μM + 2.5 μM, 3 μM + 2.5 μM, 6 μM + 2.5 μM, 12 μM + 2.5 μM, and 24 μM + 2.5 μM, respectively (the former being chidamide and the latter being anlotinib). All cells in the control group were treated with the same volume of DMSO. After gently shaking and mixing the drug or DMSO with the cells in the 24-well cell culture plates, the cells were cultured in a cell culture incubator for 24 h. Cells were then collected by centrifugation at 300 g for 5 min at 4 °C, washed once with PBS, and the apoptosis level and rate were detected by Annexin V / PI flow cytometry.

[0065] Experimental results: The results of flow cytometry detection of apoptosis levels are as follows: Figure 2A (TMD8) Figure 2C (Toledo) shows the statistical results for the apoptosis rate as follows: Figure 2B (TMD8) Figure 2B(Toledo) The results show that in TP53-mutant double-hit lymphoma cells, the combination of chidamide and anlotinib promoted apoptosis better than either the chidamide monotherapy group or the anlotinib monotherapy group, and the apoptosis effect gradually increased with increasing drug concentration.

[0066] Example 3

[0067] The combination drug combination's effect on cell cycle arrest in TP53-mutant double-hit lymphoma cell lines:

[0068] The operation method is as follows: Take a quantity of 2 × 10 5 Log-phase TP53-mutated double-hit lymphoma cell lines (TMD8 and Toledo cell lines) were seeded in 24-well cell culture plates, with chidamide monotherapy group, anlotinib monotherapy group, chidamide and anlotinib combination group, and control group set up.

[0069] In the TMD8 cell line, the concentration of chidamide monotherapy was 2 μM; the concentration of anlotinib monotherapy was 1.6 μM; and the concentrations of the combination of chidamide and anlotinib were 2 μM + 1.6 μM (the former being chidamide and the latter being anlotinib). In the Toledo cell line, the concentration of chidamide monotherapy was 750 nM; the concentration of anlotinib monotherapy was 1.6 μM; and the concentrations of the combination of chidamide and anlotinib were 750 nM + 1.6 μM (the former being chidamide and the latter being anlotinib). The control group cells were all treated with the same volume of DMSO. After gently shaking and mixing the drug or DMSO with the cells in the 24-well cell culture plate, the cells were cultured in a cell culture incubator for 24 hours. The cells were then collected by centrifugation at 300g for 5 minutes at 4°C, washed once with PBS, fixed with 75% ethanol at 4°C for 4 hours, stained with PI for 15 minutes, and then analyzed by flow cytometry to detect cell cycle arrest.

[0070] Experimental results: The results obtained are as follows Figure 3A (TMD8) and Figure 3B As shown in (Toledo), the figure shows that:

[0071] In the TMD8 cell line experiments: compared with the DMSO group, the number of cells arrested in G2 phase was significantly lower in the chidamide monotherapy group; the number of cells arrested in G1 phase was significantly higher in the chidamide and anlotinib combination group, while the number of cells arrested in S phase and G2 phase was significantly lower; compared with the anlotinib monotherapy group, the number of cells arrested in G1 phase was significantly higher in the chidamide and anlotinib combination group, while the number of cells arrested in S phase was significantly lower in the chidamide and anlotinib combination group; compared with the chidamide monotherapy group, the number of cells arrested in S phase was significantly lower in the chidamide and anlotinib combination group.

[0072] In Toledo cell line experiments: compared with the DMSO group, the number of cells arrested in the G2 phase was significantly increased in the chidamide monotherapy group; compared with the chidamide monotherapy group or the anlotinib monotherapy group, the number of cells arrested in the S phase was significantly decreased in the chidamide combined with anlotinib group, and the number of cells arrested in the G2 phase was significantly increased.

[0073] The results above indicate that, compared with the chidamide monotherapy group and the anlotinib monotherapy group, the combination of chidamide and anlotinib can more significantly induce cell cycle arrest in TP53-mutant double-hit lymphoma cells.

[0074] Example 4

[0075] Inhibitory effect of the combined drug combination on the expression of PI3K / AKT pathway-related proteins:

[0076] The operation method is as follows: take the number of units in the logarithmic growth phase as 1×10. 6 TP53-mutated double-hit lymphoma cell lines (including TMD8 and Toledo) were seeded in 6 cm cell culture dishes and set up a chidamide monotherapy group, anlotinib monotherapy group, a chidamide and anlotinib combination group, and a control group.

[0077] In the TMD8 cell line, the concentration of chidamide monotherapy was 1.5 μM, the concentration of anlotinib monotherapy was 1.6 μM, and the concentration of the combination of chidamide and anlotinib was 1.5 μM + 1.6 μM (the former being chidamide and the latter being anlotinib). In the Toledo cell line, the concentration of chidamide monotherapy was 375 nM, the concentration of anlotinib monotherapy was 1.6 μM, and the concentration of the combination of chidamide and anlotinib was 375 nM + 1.6 μM (the former being chidamide and the latter being anlotinib). The control group cells were all treated with the same volume of DMSO.

[0078] After gently agitating and mixing the drug or DMSO with the cells in the 6cm cell culture dish, the cells were cultured in a cell culture incubator for 24 hours. The cells were then collected by centrifugation at 300g for 5 minutes at 4°C, mixed with 200μL of RIPA lysis buffer, and lysed on ice for 1 hour. After centrifugation at 12000g for 30 minutes, the supernatant was collected as the extracted total intracellular protein. Western blot experiments were performed using the extracted protein to detect any changes in the expression levels of PI3K / AKT pathway-related proteins.

[0079] Experimental results: The results are as follows Figure 4As shown in the results, compared to the chidamide monotherapy group or the anlotinib monotherapy group, the combination of chidamide and anlotinib significantly downregulated the protein levels of HDAC3, VEGFR2, PI3K p85α, p-PI3KP85α / P55γ / P85β-Y467 / Y199 / Y464, AKT1, p-AKT1(Ser473), MCL-1, BCL-XL, BCL-2, c-Myc, and Cyclin A2.

[0080] Example 5

[0081] Effects of combined drug combinations on in vivo tumorigenesis in TP53-mutant double-hit lymphoma cells:

[0082] The operation method is as follows:

[0083] (1) CB17-SCID mice (weighing 16.1-20.1g) were divided into four groups of 5 mice each: control group, chidamide monotherapy group, anlotinib monotherapy group, and chidamide and anlotinib combination group. Chidamide was prepared as a suspension with 0.5% sodium carboxymethyl cellulose, and anlotinib was first dissolved in 1% DMSO and then 99% 0.5% sodium carboxymethyl cellulose was added to prepare a suspension for gavage.

[0084] (2) Constructing a CDX mouse model

[0085] Mice were irradiated with 1.5 Gy, and 2 × 10⁻⁶ samples were collected 18 hours later. 7 TMD8 cells in the logarithmic growth phase were injected subcutaneously into the right abdomen of CB-17SCID mice to induce tumor formation and establish a CDX mouse model.

[0086] (3) Ten days after injection of TMD8 cells, the mice were administered the drug by gavage. The administration regimen is shown in Table 1. The drugs were administered for 7 consecutive days. The tumor progression in mice was detected by measuring the size of the subcutaneous tumor, and the first day was recorded in the drug administration diary.

[0087] (4) After the administration of the drug, the mice were euthanized and the tumors were collected for observation.

[0088] (5) After washing the tumor tissue with PBS, 300 μL of RIPA lysis buffer containing 1% protease and phosphatase inhibitors was added to the washed tissue. The tissue was homogenized and lysed on ice for 30 min. Then, the tissue was centrifuged at 13,000 rpm for 30 min at 4 °C to obtain tumor proteins. The extracted proteins were used for Western blot experiments to detect whether the expression levels of PI3K / AKT pathway-related proteins were altered.

[0089] Table 1

[0090]

[0091] Experimental results: The changes in tumor volume in each group of tumor-bearing mice are as follows: Figure 5A As shown; the changes in tumor weight in each group of tumor-bearing mice are as follows: Figure 5B As shown; the tumor anatomy diagrams of each group of tumor-bearing mice after the experiment are shown below. Figure 5C As shown; the results of Western blot analysis of proteins extracted from tumor tissues of tumor-bearing mice in each group are as follows. Figure 5D As shown above, the results indicate that the combination of chidamide and anlotinib can better reduce the burden of TP53-mutant double-hit lymphoma and effectively inhibit the tumorigenesis process in mice bearing TP53-mutant double-hit lymphoma compared to chidamide or anlotinib alone.

[0092] The applicant declares that the technical solution of this invention is illustrated by the above embodiments, but this invention is not limited to the above embodiments, that is, it does not mean that this invention must rely on the above embodiments to be implemented. Those skilled in the art should understand that any improvements to this invention, equivalent substitutions of raw materials for the products of this invention, addition of auxiliary components, selection of specific methods, etc., all fall within the protection scope and disclosure scope of this invention.

[0093] The preferred embodiments of the present invention have been described in detail above. However, the present invention is not limited to the specific details in the above embodiments. Within the scope of the technical concept of the present invention, various simple modifications can be made to the technical solution of the present invention, and these simple modifications all fall within the protection scope of the present invention.

[0094] It should also be noted that the various specific technical features described in the above specific embodiments can be combined in any suitable manner without contradiction. In order to avoid unnecessary repetition, the present invention will not describe the various possible combinations separately.

Claims

1. The use of a combination pharmaceutical composition in the preparation of a medicament for improving or treating TP53-mutant double-hit lymphoma; The active component of the pharmaceutical composition is composed of a first active component and a second active component; The first active ingredient is selected from chidamide or a pharmaceutically acceptable salt thereof; The second active ingredient is selected from anlotinib or a pharmaceutically acceptable salt thereof.

2. Use according to claim 1, characterized in that, The combined drug composition is a single compound preparation or a combination of two separate preparations.

3. Use according to claim 2, characterized in that, The combined drug composition is a combination of two separate formulations, which are administered simultaneously or sequentially.

4. The application according to claim 3, characterized in that, The formulation can be any pharmaceutically acceptable dosage form.

5. The application according to claim 1, characterized in that, The combined pharmaceutical composition also contains pharmaceutically acceptable excipients.

6. The application according to claim 5, characterized in that, The pharmaceutically acceptable excipients are selected from any one or a combination of at least two of the following: carriers, diluents, binders, wetting agents, disintegrants, emulsifiers, cosolvents, solubilizers, osmotic pressure regulators, coating materials, colorants, pH adjusters, antioxidants, and antibacterial agents.

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