Application of ibrutinib as platinum drug sensitizer in gastric cancer treatment

Abstract

The invention relates to the technical field of medicines, and provides application of ibrutinib as a platinum drug sensitizer in gastric cancer treatment. Furthermore, the invention also provides an application of ibrutinib combined with platinum drugs in preparation of drugs for treating CREB1 high-expression gastric cancer, and further provides a pharmaceutical composition for treating gastric cancer. A mouse in-vivo experiment shows that the size and the growth speed of a stomach tumor overexpressed with CREB1 can be obviously reduced by combined use of ibrutinib and oxaliplatin, so that the platinum drug sensitivity of CREB1 high-expression stomach cancer cells is enhanced; through a gastric cancer organ model and an in-vitro cytology experiment, the ibrutinib can reduce the activity of CREB1 by inhibiting phosphorylation of various proteins of an EGFR-PI3K-AKT-CREB1 pathway, so that proliferation of gastric cancer cells is inhibited, the ferroptosis level of the gastric cancer cells is increased, and IC50 of the ibrutinib can be remarkably reduced by combined use of the ibrutinib and platinum drugs. The invention provides a new basis for the ibrutinib or the derivative thereof to inhibit the growth of CREB1 high-expression gastric cancer cells and enhance the sensitivity of platinum drugs.

Description

Technical Field

[0001] The present invention belongs to the field of biomedicine, and specifically relates to the application of ibrutinib as a platinum drug sensitizer, and also relates to the application of ibrutinib and platinum drugs in the preparation of gastric cancer therapeutic drugs, especially a new application in the treatment of gastric cancer with high CREB1 expression. Background Art

[0002] Gastric cancer is one of the most common digestive tract malignancies in the world, with the highest incidence in East Asia, posing a serious threat to human health. However, due to primary or secondary drug resistance of the tumor, the total effective rate of chemotherapy is between 20% and 58%, and the median survival time of patients with advanced gastric cancer is between 12 and 15 months. Tumor drug resistance is the main obstacle for gastric cancer patients to achieve the best therapeutic effect, and it is also one of the problems that need to be solved urgently in the clinic.

[0003] Although the treatment strategies for advanced gastric cancer vary in different countries and regions, the treatment regimen of fluorouracil combined with platinum has become the first-line chemotherapy regimen in most countries. Platinum drugs are non-specific drugs for the cell cycle. They enter the body and act on cell DNA, coordinating with DNA to form adducts of platinum (Pt) and DNA, thereby inhibiting DNA synthesis. Platinum drugs, including cisplatin, carboplatin, oxaliplatin, nedaplatin and lobaplatin, are widely used in chemotherapy regimens (see the literature: Zhou J, Kang Y, Chen L, et al. The Drug-Resistance Mechanisms of Five Platinum-Based Antitumor Agents. Front Pharmacol. 2020; 11: 343.). Primary or acquired drug resistance severely limits the clinical application of platinum therapy. The mechanism of platinum resistance is very complex, involving changes in drug transport and metabolism, increased drug efflux, enhanced DNA repair, changes in cell cycle regulation, inhibition of apoptosis, increased clearance of protein damage, increased protein glycosylation, enhanced glycolysis, dysregulation of the oxidative phosphorylation pathway, increased autophagy, etc. Solving the problem of tumor resistance mechanism is a long and arduous task, and exploring new and reliable molecular targets is a good breakthrough.

[0004] Ibrutinib (CAS registration number 936563-96-1), also known as PCI-32765, is a small molecule Bruton's tyrosine kinase (BTK) inhibitor that can covalently bind to the cysteine ​​residue in the active center of BTK, thereby inhibiting its activity. BTK is a key participant in B-cell receptor (BCR) signal transduction, transmitting signals in the BCR signaling pathway and cytokine receptor signaling pathway, mediating the migration, chemotaxis, and adhesion of B cells. In recent years, oral ibrutinib has become a breakthrough therapy for hematological malignancies such as chronic lymphocytic leukemia (CLL) (see references: Pal Singh S, Dammeijer F, Hendriks RW. Role of Bruton's tyrosine kinase in B cells and malignancies. Mol Cancer.

[0005] 2018;17(1):57.).

[0006] There is no literature reporting the role of ibrutinib as a platinum-based drug sensitizer in the treatment of gastric cancer. Summary of the invention

[0007] The purpose of the present invention is to provide a new medical use of ibrutinib, provide the use of ibrutinib as a platinum drug sensitizer in the treatment of gastric cancer, and also provide the use of ibrutinib and a platinum drug in combination in the preparation of a gastric cancer treatment drug, especially a new use in the treatment of gastric cancer with high CREB1 expression.

[0008] The first aspect of the present invention provides the use of ibrutinib in the preparation of a platinum-based drug sensitizer for treating gastric cancer.

[0009] Preferably, the gastric cancer is CREB1 highly expressed gastric cancer.

[0010] More preferably, the sensitizer is a drug that reduces platinum efflux in gastric cancer cells. Specifically, the drug that reduces platinum efflux is a drug that inhibits the protein phosphorylation process of the EGFR-PI3K-AKT-CREB1 pathway, or a drug that inhibits the CREB1-GPX4-ferroptosis axis in gastric cancer.

[0011] Studies have found that the EGFR-PI3K-AKT-CREB1 pathway is involved in the platinum resistance of gastric cancer cells, and cisplatin mainly affects the phosphorylation level of pathway proteins, while ibrutinib can inhibit the protein phosphorylation process of this pathway, thereby increasing the sensitivity of gastric cancer cells to platinum drugs. Ibrutinib can increase the level of cell ferroptosis, and this effect is more obvious in cells with high CREB1 expression, which can offset the effect of CREB1 high-expressing cells resisting ferroptosis. Ferroptosis is involved in the Pt efflux process; combined use of ibrutinib inhibits the CREB1-GPX4-ferroptosis axis, reduces the cellular GSH content, and promotes tumor cell death.

[0012] In a second aspect, the present invention provides the use of ibrutinib in the preparation of a drug for treating gastric cancer with high CREB1 expression. The present invention compares the therapeutic effects of an animal model control group (DMSO treatment), an oxaliplatin group, an ibrutinib group, and an oxaliplatin + ibrutinib group. The results show that the sensitivity of tumor tissues with high CREB1 expression to oxaliplatin is lower than that of tissues with low CREB1 expression; after treatment with the three treatment groups, the volume of tumors with high CREB1 expression was reduced, the therapeutic effects of the oxaliplatin group, the ibrutinib group, and the oxaliplatin + ibrutinib group gradually increased, and the growth rate and tumor size of the tumors in the CREB1 overexpression group and the control group after receiving the combined administration of oxaliplatin and ibrutinib were significantly reduced compared with the administration of the two drugs alone.

[0013] Therefore, the third aspect of the present invention provides the use of ibrutinib combined with platinum drugs in the preparation of a drug for treating gastric cancer with high CREB1 expression.

[0014] Preferably, the platinum drugs include cisplatin, carboplatin, oxaliplatin, nedaplatin and lobaplatin.

[0015] In a fourth aspect, the present invention provides a drug combination for treating gastric cancer, which is composed of ibrutinib and a platinum drug. Preferably, the gastric cancer is CREB1-overexpressing gastric cancer.

[0016] Functions and Effects of the Invention

[0017] In vivo experiments in mice showed that the combination of ibrutinib and oxaliplatin can significantly reduce the size and growth rate of gastric tumors overexpressing CREB1, thereby enhancing the sensitivity of gastric cancer cells with high CREB1 expression to platinum drugs. In gastric cancer organoid models and in vitro cytological experiments, ibrutinib can reduce CREB1 activity by inhibiting the phosphorylation of proteins in the EGFR-PI3K-AKT-CREB1 pathway, thereby inhibiting the proliferation of gastric cancer cells and increasing the level of ferroptosis of gastric cancer cells. In addition, its combination with platinum drugs can significantly reduce its IC 50Therefore, the present invention provides a new basis for ibrutinib or its derivatives to inhibit the growth of gastric cancer cells with high CREB1 expression and enhance their sensitivity to platinum drugs.

[0018] In addition, since ibrutinib is a mature targeted anticancer drug for the treatment of chronic lymphocytic leukemia, its trade names include Yike, Ibitiweika (Imbruvica), etc., its pharmacological action is clear, the overall effective rate is high, and it is well tolerated, which can benefit patients for a long time (see literature: BroccoliA, Argnani L, Morigi A, et al. Long-Term Efficacy and Safety of Ibrutinib in the Treatment of CLL Patients: A Real Life Experience. J Clin Med. 2021; 10 (24): 5845.). The new indications of ibrutinib provided by the present invention can be quickly transformed into clinical practice, and have great potential in the treatment of gastric cancer with high CREB1 expression. BRIEF DESCRIPTION OF THE DRAWINGS

[0019] Figure 1 These are the validation diagrams of the gastric cancer CDX model in different treatment groups, where A is the flow chart for constructing the gastric cancer CDX model, B is the general display diagram of the tumors in each group after drug treatment, C is the growth curve diagram of the tumors in each group, and D is the bar graph of the tumor weights in each group after treatment.

[0020] Figure 2 These are the IHC staining images of tumor tissues of gastric cancer CDX models in different treatment groups, where AC are the IHC staining images of p-CREB1 (A), CREB1 (B), and GPX4 (C) in each group of tumors, respectively.

[0021] Figure 3 Figure 1 is a validation diagram of the gastric cancer PDO model in different treatment groups, where A is a flow chart for the construction of the gastric cancer PDO model, BC is a bar graph of the mRNA expression levels of CREB1 (B) and GPX4 (C) in 5 PDO-Ts relative to their corresponding PDO-N, DE is a bar graph of the relative mRNA expression levels of CREB1 (D) and GPX4 (E) in 5 pairs of PDOs, FG is a bar graph of the mRNA expression levels of CREB1 (F) and GPX4 (G) after overexpression or knockdown of CREB1 in PDO-2T and PDO-3T, HI is a microscopic photograph after overexpression or knockdown of CREB1 in PDO-2T and PDO-3T and (H) a bar graph of PDO diameter (I), JK is the IC values ​​of PDO-2T (J) and PDO-3T (K) with overexpression or knockdown of CREB1 to different drugs 50 Graph.

[0022] Figure 4 This is the Western Blot result of each protein in the EGFR-PI3K-AKT-CREB1 pathway and its phosphorylation levels under the stimulation of ibrutinib and platinum drugs.

[0023] Figure 5 IC of ibrutinib for enhancing the sensitivity of CREB1-overexpressing gastric cancer cells to platinum-based drugs 50 Curve graph, where AB is the IC of cisplatin for platinum-resistant cells MKN-45-DR (A) and NCI-N87-DR (B) under the combination of ibrutinib and cisplatin 50 Graphs. CD are the IC of ibrutinib after overexpression or knockdown of CREB1 in MKN-74 (C) and HGC-27 (D). 50 Curve diagram. EF is the IC of MKN-74 (E) and HGC-27 (F) to oxaliplatin in combination with ibrutinib and oxaliplatin 50 Graph. GH is the IC of MKN-74 (G) and HGC-27 (H) to cisplatin under the combination of ibrutinib and cisplatin 50 Graph.

[0024] Figure 6 The figures are validation diagrams of ibrutinib inhibiting the ferroptosis level of gastric cancer cells with high CREB1 expression, where AB is the bar graph of MDA content of MKN-74 (A) and HGC-27 (B) under ibrutinib stimulation, CD is the bar graph of GSH to GSSG ratio of MKN-74 (A) and HGC-27 (B) under ibrutinib stimulation, EF is the bar graph of intracellular Lipid ROS level after ibrutinib stimulation in MKN-74 (E) and HGC-27 (F) transfected with CREB1-specific siRNA, and GH is the fluorescence images of LipidROS in the oxidized state (green) and reduced state (red) in MKN-74 (G) and HGC-27 (H) after ibrutinib stimulation.

[0025] Figure 7 The graphs show that ibrutinib increases the platinum content in gastric cancer cells with high CREB1 expression, wherein AD is the distribution curve of platinum content in cells of different treatment groups in MKN-74 cells with overexpression or knockdown of CREB1, and EH is the distribution curve of platinum content in cells of different treatment groups in HGC-27 cells with overexpression or knockdown of CREB1. DETAILED DESCRIPTION

[0026] The present invention is described in detail below in conjunction with the embodiments and the accompanying drawings. However, the following embodiments should not be considered as limiting the scope of the present invention.

[0027] Example 1: In vivo study in nude mice: Ibrutinib can reduce the size and growth rate of CREB1-overexpressing gastric tumors and enhance the sensitivity of platinum drugs

[0028] 1. Experimental Methods

[0029] Thirty-two 4-week-old male BALB / c-nu / nu nude mice were randomly divided into two groups (n=16) and given the same feeding conditions. HGC-27CREB1-vector (CREB1 low expression) and HGC-27CREB1-OE (CREB1 high expression) stable transfected cells in good growth state and logarithmic growth phase were collected and the cells were cultured at 5×10 7 / mL was resuspended in a solution prepared by mixing PBS and Matrigel at a ratio of 1:1 (placed on ice to prevent solidification); the two groups of tumor cells were implanted subcutaneously on the right back of each nude mouse (100 μL, 5×10 6 / mouse) to establish the nude mouse CDX model.

[0030] When the tumor volume reaches about 50 mm 3 At the same time, each group of nude mice was randomly divided into 4 groups (n=4) again, and the solution was prepared according to the following method and intraperitoneal injection was performed regularly:

[0031] 1) Control group (2% DMSO + 40% PEG 300 + 5% Tween 80 + 53% dd H2O)

[0032] 2) Oxaliplatin group (dd H2O dissolved, 5 mg kg -1 / every 3 days)

[0033] 3) Ibrutinib group (gradually dissolved in 2% DMSO + 40% PEG 300 + 5% Tween 80 + 53% ddH2O, 6 mg kg -1 / sky)

[0034] 4) Oxaliplatin + ibrutinib group

[0035] Tumor size was measured every 3 days using a vernier caliper, and tumor volume was calculated using the following formula (where L represents length and W represents width): tumor volume (mm 3 )=L×W 2 / 2.

[0036] After 21 days of treatment, the mice were killed by cervical dislocation, and the tumor tissues were removed, weighed, photographed, and subjected to IHC staining analysis.

[0037] 2. Experimental Results

[0038] The above experimental results are from Figure 1-2 show:

[0039] (1) Construct a gastric cancer cell nude mouse CDX model as shown in the figure ( Figure 1 A), it can be seen that the growth rate and tumor size of the tumor overexpressing CREB1 are greater than those of the control group, indicating that CREB1 can promote the growth and proliferation of gastric cancer cells in nude mice; compared with the control group, the tumor volume of the tumor overexpressing CREB1 after receiving oxaliplatin treatment is significantly increased, indicating that the sensitivity of tumor tissues with high CREB1 expression to oxaliplatin is lower than that of tissues with low CREB1 expression; compared with the DMSO-treated group, the reduction in the volume of the tumor overexpressing CREB1 in the ibrutinib-treated group is slightly higher than that in the control group; the growth rate and tumor size of the tumors in the CREB1 overexpression group and the control group after receiving the combination of oxaliplatin and ibrutinib are significantly reduced compared with the administration of the two drugs alone, and there is no statistical difference in the tumor size between the CREB1 overexpression group and the control group, which greatly weakens the pro-proliferation and platinum-resistant effects brought about by CREB1 overexpression ( Figure 1 BD).

[0040] (2) p-CREB1( Figure 2 A) CREB1( Figure 2 B) and GPX4( Figure 2 C) IHC staining of the three indicators. As shown in the figure, p-CREB1 and CREB1 are mainly expressed in the nucleus, while GPX4 is mainly expressed in the cytoplasm. After oxaliplatin treatment, the expression of p-CREB1 and CREB1 in the control group increased slightly, while the expression of p-CREB1 and CREB1 in the CREB1-OE group increased significantly. Ibrutinib treatment can significantly reduce the increase of p-CREB1 and CREB1 caused by oxaliplatin, indicating that ibrutinib can inhibit the CREB1-GPX4-ferroptosis axis in gastric cancer and increase the sensitivity to platinum drugs.

[0041] Example 2: Gastric cancer PDO model: Ibrutinib can reduce the size and growth rate of CREB1-overexpressing organoids and enhance their sensitivity to platinum drugs

[0042] 1. Experimental methods:

[0043] Organoids (PDO) refer to 3D miniature organs formed in vitro by a cluster of cells with self-renewal and self-organization capabilities, which are similar to real tissues and organs in the human body to a certain extent. Tumor tissues and normal gastric mucosal tissues of 5 gastric cancer patients were selected, and they were processed and cultured immediately after surgery to construct a gastric cancer PDO model. Appropriate amounts of PDO were selected for qRT-PCR experiments to determine the mRNA levels of CREB1 and GPX4. PDO-2T and PDO-3T were selected for lentiviral transfection of CREB1 overexpression, knockdown and their respective controls, and the transfection efficiency was verified using qRT-PCR. PDO-2T and PDO-3T transfected with CREB1 were digested into single cells and continued to be subcultured. Photos were taken on the 7th day, and the diameter of PDO in each group was measured and counted using Image J software.

[0044] IC50 determination of PDO: PDO with good growth status was digested and mixed with Matrigel in a 1:1 ratio and then resuspended in a 96-well plate (5 μL per well, 6000 cells). After culturing for 3-5 days, the cells were divided into experimental groups (oxaliplatin, Erastin, oxaliplatin + Erastin, oxaliplatin + Fer1 (iron perturbator), ibrutinib, oxaliplatin + ibrutinib), and drugs of different concentration gradients were added. The cells were cultured for another 5 days at 37°C and 5% CO2. The cell concentration was determined by CCK8 method, and the IC50 of PDO for each group of drugs was calculated. 50 And draw the picture.

[0045] 2. Experimental Results

[0046] The gastric cancer PDO model was constructed as shown in the figure ( Figure 3 A), the mRNA levels of CREB1 and GPX4 were measured, and it can be seen that the expression trends of the two are roughly the same ( Figure 3 BC); Since the expression of CREB1 and GPX4 in PDO-2T and PDO-3T were both high ( Figure 3 DE), so CREB1 overexpression and knockdown virus transfection were performed on them; it was found that the GPX4 expression of PDO after overexpression of CREB1 was also increased, and the mRNA level of GPX4 decreased accordingly after CREB1 was knocked down in PDO, indicating that the expression of CREB1 and GPX4 in PDO is consistent ( Figure 3 FG); As shown in Figure HI, the diameter of PDO overexpressing CREB1 is significantly larger than that of the control group, while the diameter of PDO after CREB1 knockdown is smaller than that of the NC group. These results confirm that CREB1 has a proliferative effect on gastric cancer cells at the PDO level; IC 50 The results showed that the IC values ​​of oxaliplatin for PDO-2T and PDO-3T overexpressing CREB1 were 50After knocking down CREB1, the IC 50 This indicates that CREB1 at the PDO level can reduce the sensitivity of gastric cancer cells to platinum drugs; after combined use of Erastin, the IC of PDO-2T and PDO-3T to oxaliplatin 50 The ferroptosis inhibitor Fer1 can rescue the killing effect of oxaliplatin on PDO, indicating that the ferroptosis pathway is closely related to CREB1 in promoting platinum resistance in gastric cancer. The IC of PDO with overexpression and knockdown of CREB1 on ibrutinib 50 There are differences but they are not very obvious. The combined use of oxaliplatin and ibrutinib can significantly weaken the effect of overexpression of CREB1 on reducing the sensitivity of gastric cancer to platinum drugs, indicating that ibrutinib can enhance the sensitivity of patients with high CREB1 expression to oxaliplatin ( Figure 3 JK).

[0047] Example 3: Ibrutinib can inhibit the protein phosphorylation process of the EGFR-PI3K-AKT-CREB1 pathway, thereby increasing the sensitivity of gastric cancer cells to platinum drugs

[0048] 1. Experimental methods:

[0049] As a classic BTK inhibitor, studies have shown that ibrutinib can inhibit multiple kinases (EGFR, HER2, etc.), and can be used as a radiosensitizer and chemosensitizer to play its therapeutic role in solid tumors. To further clarify its mechanism of action in inhibiting CREB1, drug stimulation (Blank, cisplatin, ibrutinib, and cisplatin + ibrutinib) was performed in wild-type HGC-27 cells, and Western Blot was used to analyze the changes in the proteins and their phosphorylation levels of the EGFR-PI3K-AKT-CREB1 pathway.

[0050] 2. Experimental Results

[0051] like Figure 4 As shown in the data, after the addition of cisplatin, the expression levels of p-EGFR, p-PI3K, p-AKT and p-CREB1 proteins increased significantly, CREB1 also increased, while EGFR, PI3K and AKT did not change significantly; after the addition of ibrutinib alone, the phosphorylation levels of each protein in the pathway decreased, and the total protein content did not change significantly; and the addition of ibrutinib at the same time as cisplatin can offset the increase in the phosphorylation levels of each protein brought by cisplatin to a certain extent, which indicates that the EGFR-PI3K-AKT-CREB1 pathway is involved in the platinum resistance of gastric cancer cells, and cisplatin mainly affects the phosphorylation level of pathway proteins, while ibrutinib can inhibit the protein phosphorylation process of this pathway, thereby increasing the sensitivity of gastric cancer cells to platinum drugs.

[0052] Example 4: Ibrutinib can enhance the sensitivity of CREB1-overexpressing gastric cancer cells to platinum-based drugs

[0053] 1. Experimental methods:

[0054] To verify whether ibrutinib can enhance the sensitivity of gastric cancer cells to platinum-based drugs, we first measured the IC values ​​of cisplatin and cisplatin + ibrutinib in WT and DR of MKN-45 and NCI-N87 cells. 50 The IC of ibrutinib was then measured in MKN-74 and HGC-27 CREB1-stable transfected cells. 50 Measured and tested by IC 50 The curve selects the maximum concentration of ibrutinib that can ensure that cells do not die as much as possible for subsequent combined administration (i.e., IC 50 The concentration corresponding to the inflection point of the curve platform) was 15 μM for MKN-74 and 4 μM for HGC-27. Finally, the IC value of CREB1 stable transfected cells to the combination of ibrutinib and platinum drugs (cisplatin, oxaliplatin) was determined. 50 To clarify the platinum-sensitizing effect of ibrutinib on gastric cancer cells.

[0055] 2. Experimental Results

[0056] The results showed that MKN-45-WT had an IC of 5.34% for cisplatin after combined use of ibrutinib. 50 The IC of MKN-45-DR against cisplatin decreased from 13.64 μM to 8.45 μM after combined use of ibrutinib. 50 The IC of NCI-N87-WT to cisplatin decreased from 33.32μM to 15.38μM after combined use of ibrutinib 50 The IC of NCI-N87-DR against cisplatin decreased from 11.50 μM to 10.12 μM after combined use of ibrutinib. 50 The decrease from 40.63 μM to 15.36 μM indicated that ibrutinib could effectively increase the sensitivity of platinum-resistant strains to platinum drugs ( Figure 5 AB); then the IC of ibrutinib was measured in MKN-74 and HGC-27 CREB1 stable transfected cells. 50 In the assay, it can be seen that interfering with CREB1 expression did not significantly change the sensitivity of gastric cancer cells to ibrutinib ( Figure 5 CD); Figure 5 EH showed that the combined use of ibrutinib significantly reduced the IC of CREB1 stable transfected cells to platinum drugs compared with cisplatin or oxaliplatin alone. 50 values, and this effect was more pronounced in cells overexpressing CREB1.

[0057] Example 5: Ibrutinib sensitizes CREB1-overexpressing cells to platinum drugs by inhibiting the ferroptosis pathway

[0058] 1. Experimental methods:

[0059] To determine whether ibrutinib can enhance the sensitivity of CREB1-overexpressing cells to platinum drugs by inhibiting the ferroptosis pathway, after adding ibrutinib to MKN-74 and HGC-27 cells with different CREB1 expression levels, the levels of MDA, GSH / GSSG, and Lipid ROS in each group of cells were measured to represent the degree of ferroptosis in gastric cancer cells. CREB1 stable transfectants were used in the MDA and GSH / GSSG assays, and wild-type MKN-74 and HGC-27 cells were transiently transfected with CREB1-specific siRNA for Lipid ROS assays.

[0060] 2. Experimental Results

[0061] The results showed that compared with the blank control group, the intracellular MDA content and Lipid ROS level increased significantly after the addition of ibrutinib, while the GSH / GSSG ratio decreased ( Figure 6 AH), indicating that ibrutinib can increase the level of cell ferroptosis, and this effect is more obvious in cells with high CREB1 expression, which can offset the effect of CREB1 high-expressing cells in resisting ferroptosis.

[0062] Example 6: Ibrutinib can reduce Pt efflux and increase platinum drug sensitivity by inhibiting CREB1 and its phosphorylation. 1. Experimental method:

[0063] The mechanism of cell resistance to platinum is very complex, involving multiple pathways such as increased drug efflux, changes in drug transport and metabolism, enhanced DNA repair, inhibition of apoptosis, changes in cell cycle regulation, increased clearance of protein damage, increased protein glycosylation, enhanced glycolysis, dysregulation of oxidative phosphorylation pathways, and increased autophagy. Single-cell inductively coupled plasma mass spectrometry was used to determine the intracellular Pt content in CREB1 stable transfected cells after drug stimulation (oxaliplatin, oxaliplatin + Erastin, oxaliplatin + Fer-1, and oxaliplatin + ibrutinib).

[0064] 2. Experimental Results

[0065] like Figure 7As shown in the figure, under the action of the same concentration of oxaliplatin, the Pt content of cells with high CREB1 expression decreased, indicating that CREB1 can promote Pt efflux; the Pt content of the oxaliplatin + Erastin treatment group increased, and the Pt content of the oxaliplatin + Fer-1 treatment group decreased. The ferroptosis perturbator affected the intracellular Pt content, indicating that ferroptosis was involved in the Pt efflux process; and the combined use of ibrutinib inhibited the CREB1-GPX4-ferroptosis axis and reduced the cellular GSH content, resulting in an increase in the Pt concentration in the oxaliplatin + ibrutinib treatment group, which promoted tumor cell death.

[0066] The basic principles, main features and significant advantages of the present invention have been described in detail above. The purpose of the description in the embodiments and the specification is only to further explain the operating principle of the present invention. Without violating the core idea of ​​the present invention and the statutory protection scope, the present invention allows and covers all kinds of reasonable changes and improvements. These changes and improvements will be deemed to be within the scope of protection requested by the present invention. Specifically, the scope of protection of the present invention will be clearly and strictly defined by the attached claims and their equivalents with equal legal effect.

Claims

1. Application of ibrutinib in the preparation of platinum-based drug sensitizers for the treatment of gastric cancer.

2. The use according to claim 1, characterized in that: The gastric cancer is CREB1 highly expressed gastric cancer.

3. The use according to claim 1, characterized in that: The sensitizer is a drug that reduces the efflux of platinum in gastric cancer cells.

4. The use according to claim 3, characterized in that: The drug for reducing platinum efflux is a drug for inhibiting the protein phosphorylation process of the EGFR-PI3K-AKT-CREB1 pathway, or a drug for inhibiting the CREB1-GPX4-ferroptosis axis in gastric cancer.

5. Application of ibrutinib in the preparation of drugs for the treatment of gastric cancer with high CREB1 expression.

6. Application of ibrutinib combined with platinum drugs in the preparation of drugs for the treatment of gastric cancer with high CREB1 expression.

7. The use according to claim 6, characterized in that: The platinum drugs include cisplatin, carboplatin, oxaliplatin, nedaplatin and lobaplatin.

8. A drug combination for treating gastric cancer, characterized in that: It consists of ibrutinib and a platinum drug.

9. The drug combination for treating gastric cancer according to claim 8, characterized in that: The gastric cancer is CREB1 highly expressed gastric cancer.

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