Method for killing brain tumor cells in vitro

Abstract

The invention discloses a method for killing brain tumor cells in vitro. According to the embodiment of the invention, the method comprises the following step: carrying out contact treatment on ABBV-075 and separated brain tumor cells so as to kill the brain tumor cells. According to the method provided by the embodiment of the invention, the efficiency of killing the brain tumor cells in vitro can be improved.

Description

Technical Field

[0001] The present invention relates to the field of medicine, and in particular, to a method for killing brain tumor cells in vitro. Further, the present invention relates to a method for targeting brain tumor cells in vitro, the use of a reagent in preparing a kit, the use of a reagent in preparing a drug, a pharmaceutical composition, a method for inhibiting CREB5 expression, and a method for promoting the differentiation of brain tumor cells into astrocytes. Background Art

[0002] Brain tumors are the most common malignant tumors in the central nervous system, with high invasiveness, easy recurrence and treatment resistance. Diffuse Intrinsic Pontine Glioma (DIPG) is a particularly challenging type of brain tumor that occurs in the brainstem, usually affecting children and adolescents, and has a very poor prognosis. From a classification point of view, DIPG belongs to H3K27M mutant diffuse midline glioma (Diffuse Midline Glioma-H3K27M, DMG-H3K27M). DMG is a new brain tumor added to the 2016 edition of the WHO classification of malignant tumors of the central nervous system (CNS). DMG occurs in the spinal cord, brainstem, pineal region and thalamus; clinical behavior is invasive. H3.3K27M mutation is a common genetic variation in DIPG, which is closely related to the aggressiveness and treatment resistance of the tumor.

[0003] Although some progress has been made in killing brain tumor cells in vitro in recent years, there are still many limitations to the current methods of killing brain tumor cells. The current methods of killing brain tumor cells are often accompanied by serious side effects, and the killing effect is limited and the specificity of targeting brain tumor cells is poor.

[0004] Therefore, it is urgent to develop a new method for killing brain tumor cells in vitro to improve the efficiency of killing brain tumor cells. Summary of the invention

[0005] The present invention aims to solve at least one of the technical problems existing in the prior art to a certain extent. To this end, the present invention provides a method for killing brain tumor cells in vitro.

[0006] The present invention is accomplished based on the following findings of the inventors:

[0007] Current methods for killing brain tumor cells often have serious side effects, low killing efficiency, and poor specificity in targeting brain tumor cells. In order to overcome this problem, the inventors proposed a method for killing brain tumor cells in vitro, which improves the efficiency of killing brain tumor cells in vitro and improves the specificity of targeting brain tumor cells.

[0008] In a first aspect of the present invention, the present invention provides a method for killing brain tumor cells in vitro. According to an embodiment of the present invention, the method comprises: contacting ABBV-075 with isolated brain tumor cells to kill the brain tumor cells. The method according to an embodiment of the present invention can improve the efficiency of killing brain tumor cells in vitro.

[0009] In a second aspect of the present invention, the present invention provides a method for targeting brain tumor cells in vitro. According to an embodiment of the present invention, the method comprises: contacting ABBV-075 with a separated cell population, wherein the cell population comprises brain tumor cells. According to the method of the embodiment of the present invention, brain tumor cells can be specifically targeted in vitro.

[0010] In the third aspect of the present invention, the present invention provides the use of a reagent in preparing a kit. According to an embodiment of the present invention, the reagent includes ABBV-075, and the kit is used to inhibit the expression of CREB5 and / or ID1.

[0011] In a fourth aspect of the present invention, the present invention provides a use of an agent in preparing a drug. According to an embodiment of the present invention, the agent includes ABBV-075, and the drug is used for treating or preventing glioma.

[0012] In a fifth aspect of the present invention, the present invention provides a pharmaceutical composition for treating or preventing glioma. According to an embodiment of the present invention, the pharmaceutical composition comprises: ABBV-075. The pharmaceutical composition according to an embodiment of the present invention can improve the efficiency of killing brain tumor cells in vitro and improve the specificity of targeting brain tumor cells.

[0013] In a sixth aspect of the present invention, the present invention provides a method for inhibiting the expression of CREB5. According to an embodiment of the present invention, the method contacts ABBV-075 with isolated cells expressing CREB5 to inhibit the expression of CREB5. According to the method of an embodiment of the present invention, the expression of CREB5 can be inhibited.

[0014] In a seventh aspect of the present invention, the present invention provides a method for promoting the differentiation of brain tumor cells into astrocytes. According to an embodiment of the present invention, the method comprises contacting ABBV-075 with isolated brain tumor cells to promote the differentiation of the brain tumor cells into normal glial cells. The method according to an embodiment of the present invention can promote the differentiation of brain tumor cells into astrocytes.

[0015] Additional aspects and advantages of the present invention will be given in part in the following description and in part will be obvious from the following description, or will be learned through practice of the present invention. BRIEF DESCRIPTION OF THE DRAWINGS

[0016] The above and / or additional aspects and advantages of the present invention will become apparent and easily understood from the description of the embodiments in conjunction with the following drawings, in which:

[0017] Figure 1 This is a diagram of RNA-seq data analysis of DIPG treated with the BRD4 inhibitor JQ1 in Example 1.

[0018] Figure 2 This is a graph showing the qPCR results of detecting CREB5 and stemness-related genes after JQ1 treatment in Example 1 through qPCR experiments.

[0019] Figure 3 This is a graph showing the qPCR experimental results of 18 BRD4 small molecule inhibitors in Example 2.

[0020] Figure 4 This is a graph showing the cell killing effects of the five BRD4 small molecule inhibitors in Example 2 on DIPG cells and PPC cells.

[0021] Figure 5 This is a diagram showing the molecular docking experiment results of ABBV-075 and JQ1 in Example 3.

[0022] Figure 6 This is a Western Blot diagram of protein immunoblotting experiments (Western Blot) in Example 3 following ABBV-075 treatment for different time periods.

[0023] Figure 7 This is a Western Blot diagram of protein immunoblotting experiments (Western Blot) in Example 3 following treatment with different concentrations of ABBV-075.

[0024] Figure 8 This is a diagram showing the experimental results of down-regulating the levels of OLIG2 and ID1 proteins in Example 3 to induce neurite-like differentiation phenotype in DIPG cells.

[0025] Fig. 9 This is a graph showing the experimental results of the dual fluorescence reporter system in Example 3, which shows that the super enhancer of CREB5 can significantly promote the promoter activity of CREB5.

[0026] Fig.10 This is a graph showing the experimental results of the dual fluorescence reporter system in Example 3, which shows that ABBV-075 can inhibit the expression of CREB5 by inhibiting the activities of the super enhancer and promoter of CREB5.

[0027] Fig.11 This is a graph showing the experimental results of the dual fluorescence reporter system in Example 3, which shows that 20 nM ABBV-075 can significantly inhibit the activity of the CREB5 enhancer.

[0028] Fig.12 The graph shows the growth curve and spheroidization test results of ABBV-075 on different DIPG cells in Example 4.

[0029] Fig.13 This is the in vivo fluorescence imaging image in Example 4.

[0030] Fig.14 This is a data analysis diagram of the combined therapeutic effect of ABBV-075 and BRM014 in treating DIPG17 and TT150714 cells in Example 4. DETAILED DESCRIPTION

[0031] The embodiments of the present invention are described in detail below. The embodiments described below are exemplary and are only used to explain the present invention, and should not be understood as limiting the present invention.

[0032] It should be noted that the terms "first" and "second" are used for descriptive purposes only and should not be understood as indicating or implying relative importance or implicitly indicating the number of the indicated technical features. Therefore, the features defined as "first" and "second" may explicitly or implicitly include one or more of the features. Further, in the description of the present invention, unless otherwise specified, the meaning of "plurality" is two or more.

[0033] In this document, the terms “contain”, “include” or “comprise” are open expressions, that is, they include the contents specified in the present invention but do not exclude other contents.

[0034] As used herein, the terms "optionally", "optional" or "optionally" generally mean that the subsequently described event or circumstance may but need not occur, and that the description includes instances where the event or circumstance occurs and instances where it does not.

[0035] In this article, the term "H3.3K27M mutant diffuse intrinsic pontine glioma" is a malignant tumor that occurs in the brainstem, specifically in the pons. This tumor mainly affects children and usually has a very poor prognosis. The H3.3K27M mutation occurs on the third isoform of histone H3 (H3.3), specifically on lysine (K) at position 27. The normal lysine is mutated to methionine (M). This mutation affects the function of histone H3, leading to changes in chromatin structure and gene expression. DIPG mainly occurs in the pons of the brainstem, which controls many basic physiological functions, such as breathing and heartbeat.

[0036] The present invention provides an in vitro method for killing brain tumor cells, an in vitro method for targeting brain tumor cells, use of a reagent in preparing a kit, use of a reagent in preparing a drug, a pharmaceutical composition, a method for inhibiting CREB5 expression, and a method for promoting the differentiation of brain tumor cells into astrocytes, which will be described in detail below.

[0037] A method for killing brain tumor cells in vitro

[0038] In a first aspect of the present invention, the present invention provides a method for killing brain tumor cells in vitro. According to an embodiment of the present invention, the method comprises: contacting ABBV-075 with isolated brain tumor cells to kill the brain tumor cells. The method according to an embodiment of the present invention can improve the efficiency of killing brain tumor cells in vitro.

[0039] According to an embodiment of the present invention, the structure of ABBV-075 is as follows:

[0040]

[0041] According to an embodiment of the present invention, the concentration of ABBV-075 is 1nM to 1000nM, for example, it can be 1nM, 10nM, 100nM, 200nM, 300nM, 400nM, 500nM, 600nM, 700nM, 800nM, 900nM, 1000nM or a range of 10nM to 1000nM, 100nM to 1000nM. A too low concentration of ABBV-075 reduces the tumor killing effect of the drug, and a too high concentration causes the drug to have toxic side effects on normal tissues. The method according to an embodiment of the present invention can improve the efficiency of killing brain tumor cells in vitro.

[0042] According to an embodiment of the present invention, the brain tumor occurs in at least one of the spinal cord, brain stem, pineal region and thalamus.

[0043] According to an embodiment of the present invention, the brain tumor cells include H3.3K27M mutant diffuse intrinsic pontine glioma cells. According to an embodiment of the present invention, H3.3K27M mutant diffuse intrinsic pontine glioma (DIPG) is a malignant tumor occurring in the pons of the brainstem of children, characterized by aggressive growth, difficulty in surgical resection, lack of effective treatment methods, and extremely poor prognosis. H3.3K27M mutation is a common histone mutation in DIPG, which is associated with poor prognosis. This mutation leads to a decrease in genome-wide H3K27me3 and abnormal transcription, and is considered to be the main driving factor for the onset of DIPG. Treatment of DIPG usually includes radiotherapy, but the median survival period is still short, about 9 to 12 months, the 2-year survival rate is about 10% to 25%, and the 5-year survival rate is less than 1%. DIPG patients with H3.3K27M mutations usually have a worse prognosis, and there is currently no particularly effective treatment for this type of tumor. According to an embodiment of the present invention, ABBV-075 is contacted with isolated H3.3K27M mutant diffuse intrinsic pontine glioma cells to kill the H3.3K27M mutant diffuse intrinsic pontine glioma cells.

[0044] According to an embodiment of the present invention, the method further comprises contacting the isolated brain tumor cells with the BRG1 inhibitor.

[0045] According to an embodiment of the present invention, the BRG1 inhibitor includes BRM014.

[0046] According to an embodiment of the present invention, the structure of BRM014 is as follows:

[0047]

[0048] A method for targeting brain tumor cells in vitro

[0049] In a second aspect of the present invention, the present invention provides a method for targeting brain tumor cells in vitro. According to an embodiment of the present invention, the method comprises: contacting ABBV-075 with a separated cell population, wherein the cell population comprises brain tumor cells. According to the method of the embodiment of the present invention, brain tumor cells can be specifically targeted in vitro.

[0050] According to an embodiment of the present invention, the concentration of ABBV-075 is 1nM to 1000nM, for example, it can be 1nM, 10nM, 100nM, 200nM, 300nM, 400nM, 500nM, 600nM, 700nM, 800nM, 900nM, 1000nM or a range of 10nM to 1000nM, 100nM to 1000nM. A too low concentration of ABBV-075 reduces the tumor killing effect of the drug, and a too high concentration causes the drug to have toxic side effects on normal tissues. According to the method of the embodiment of the present invention, brain tumor cells can be specifically targeted in vitro.

[0051] According to an embodiment of the present invention, the brain tumor occurs in at least one of the spinal cord, brain stem, pineal region and thalamus.

[0052] According to an embodiment of the present invention, the brain tumor cells include H3.3K27M mutant diffuse intrinsic pontine glioma cells.

[0053] According to an embodiment of the present invention, the cell population further includes normal brain cells.

[0054] According to an embodiment of the present invention, the normal brain cells include at least one of glial cells and neuronal cells.

[0055] Use of reagents in preparing kits

[0056] In the third aspect of the present invention, the present invention provides the use of a reagent in preparing a kit. According to an embodiment of the present invention, the reagent includes ABBV-075, and the kit is used to inhibit the expression of CREB5 and / or ID1.

[0057] Use of reagents in the preparation of drugs

[0058] In a fourth aspect of the present invention, the present invention provides a use of an agent in preparing a drug. According to an embodiment of the present invention, the agent includes ABBV-075, and the drug is used for treating or preventing glioma.

[0059] According to an embodiment of the present invention, the reagent further comprises a BRG1 inhibitor. According to an embodiment of the present invention, ABBV-075 and the BRG1 inhibitor exert a synergistic effect and can improve the efficiency of killing gliomas.

[0060] According to an embodiment of the present invention, the BRG1 inhibitor includes BRM014. According to an embodiment of the present invention, ABBV-075 and BRM014 play a synergistic role and can improve the efficiency of killing glioma.

[0061] According to an embodiment of the present invention, the glioma occurs in at least one of the spinal cord, brain stem, pineal region and thalamus.

[0062] According to an embodiment of the present invention, the glioma includes H3.3K27M mutant diffuse intrinsic pontine glioma.

[0063] Pharmaceutical composition

[0064] In a fifth aspect of the present invention, the present invention provides a pharmaceutical composition for treating glioma. According to an embodiment of the present invention, the pharmaceutical composition comprises: ABBV-075. The pharmaceutical composition according to an embodiment of the present invention can improve the efficiency of killing brain tumor cells in vitro and improve the specificity of targeting brain tumor cells.

[0065] According to an embodiment of the present invention, the pharmaceutical composition further comprises: a BRG1 inhibitor. According to an embodiment of the present invention, ABBV-075 and a BRG1 inhibitor play a synergistic role, which can improve the efficiency of killing brain tumor cells in vitro and improve the specificity of targeting brain tumor cells.

[0066] According to an embodiment of the present invention, the BRG1 inhibitor includes BRM014. According to an embodiment of the present invention, ABBV-075 and BRM014 play a synergistic role, which can improve the efficiency of killing brain tumor cells in vitro and improve the specificity of targeting brain tumor cells.

[0067] According to an embodiment of the present invention, the ABBV-075 and the BRG1 inhibitor are suitable for simultaneous or consecutive administration.

[0068] According to an embodiment of the present invention, the glioma occurs in at least one of the spinal cord, brain stem, pineal region and thalamus.

[0069] According to an embodiment of the present invention, the glioma includes H3.3K27M mutant diffuse intrinsic pontine glioma.

[0070] Methods for inhibiting CREB5 expression

[0071] In a sixth aspect of the present invention, the present invention provides a method for inhibiting the expression of CREB5. According to an embodiment of the present invention, the method contacts ABBV-075 with isolated cells expressing CREB5 to inhibit the expression of CREB5. According to the method of an embodiment of the present invention, the expression of CREB5 can be inhibited.

[0072] According to an embodiment of the present invention, the cells expressing CREB5 include brain tumor cells.

[0073] According to an embodiment of the present invention, the brain tumor occurs in at least one of the spinal cord, brain stem, pineal region and thalamus.

[0074] According to an embodiment of the present invention, the brain tumor cells include H3.3K27M mutant diffuse intrinsic pontine glioma cells.

[0075] According to an embodiment of the present invention, inhibiting the expression of CREB5 is performed in at least one of the following ways: the activity of CREB5 super enhancer is inhibited; the level of H3K27ac histone modification is reduced.

[0076] Method for promoting differentiation of brain tumor cells into astrocytes

[0077] In a seventh aspect of the present invention, the present invention provides a method for promoting the differentiation of brain tumor cells into astrocytes. According to an embodiment of the present invention, the method comprises contacting ABBV-075 with isolated brain tumor cells to promote the differentiation of the brain tumor cells into astrocytes. The method according to an embodiment of the present invention can promote the differentiation of brain tumor cells into astrocytes.

[0078] According to an embodiment of the present invention, the brain tumor cells include H3.3K27M mutant diffuse intrinsic pontine glioma cells.

[0079] According to an embodiment of the present invention, the brain tumor occurs in at least one of the spinal cord, brain stem, pineal region and thalamus.

[0080] The scheme of the present invention will be explained below in conjunction with the embodiments. It will be appreciated by those skilled in the art that the following embodiments are only used to illustrate the present invention and should not be considered as limiting the scope of the present invention. Where specific techniques or conditions are not indicated in the embodiments, the techniques or conditions described in the literature in this area or the product specifications are used. The reagents or instruments used are not indicated by the manufacturer and are all conventional products that can be obtained commercially.

[0081] Embodiment 1:

[0082] CREB5 is a malignant oncogenic factor that is specifically highly expressed in patients with diffuse intrinsic pontine glioma. Knockdown of this factor can significantly inhibit the proliferation of DIPG cells, promote their differentiation, and thus inhibit the growth of tumors in vivo. In addition, as an upstream regulatory protein of the tumor stemness regulatory factor ID1 in the H3.3K27M subtype, CREB5 can also collaborate with the chromatin remodeling complex SWI / SNF to regulate the expression of a series of genes associated with poor prognosis in tumor patients. Therefore, further targeting CREB5 is expected to provide good therapeutic effects for DIPG tumor patients.

[0083] In this example, the transcriptional expression profile of CREB5 knockdown was combined with Connectivity Map (L1000) analysis to find that the small molecule inhibitor of BRD4, JQ1 (HY-13030, MCE Company), had similar effects to CREB5 knockdown.

[0084] First, this example analyzes the ChIP-seq data of H3.3K27M and H3.1K27M subtypes (NCBI GEO database, https: / / www.ncbi.nlm.nih.gov / geo / query / acc.cgi?acc=GSE78801), and finds that the chromatin reader proteins BRD2 and BRD4, histone active modification marks H3K27ac and cancer histone H3.3K27M are enriched in the super enhancer region of CREB5. However, these phenomena do not exist in DIPG cells of the H3.1K27M subtype, which is consistent with the phenomenon that the expression level of CREB5 in DIPG cells of the H3.1K27M subtype is lower than that in DIPG cells of the H3.3K27M subtype.

[0085] Secondly, combined with the common BRD4 inhibitor JQ1 to treat DIPG RNA-seq data analysis, the results are as follows Figure 1 As shown, this example found that the expression level of CREB5 in H3.3K27M subtype DIPG cells decreased with the extension of JQ1 treatment time, which further verified that the inhibitor of BRD4 had a similar expression profile as CREB5 knockdown.

[0086] Next, this example detected the expression of CREB5 and some stemness-related genes (OLIG2, SOX2, PDGFRA, PROM1) after JQ1 treatment by qPCR experiments. The results are as follows: Figure 2 As shown, the results showed that JQ1 treatment can indeed significantly downregulate the expression of CREB5 and some other stemness genes. Therefore, the above experimental data show that small molecule inhibitors of BRD4 can significantly inhibit the expression of CREB5, which is more important for further screening of more effective small molecule inhibitors of BRD4 for targeting CREB5 transcription factors.

[0087] Embodiment 2:

[0088] In order to find potential BRD4 small molecule inhibitors targeting CREB5 super enhancer activation, this example next screened the MCE active compound library. The following is the screening strategy: 1. This example finds small molecule inhibitors that can target BRD4 activity from the MCE active compound library; 2. The inhibition level of BRD4 small molecule inhibitors on CREB5 is evaluated in DIPG cells through qPCR experiments; 3. By comparing the effects of small molecule inhibitors on the cell viability of normal pons-derived progenitor cells (PPC cells) and DIPG cells, safe and reliable BRD4 small molecule inhibitors are screened out.

[0089] In this example, 18 BRD4 small molecule inhibitors (HY-13960, HY-111422, HY-112610, HY-13235, HY-100015, HY-13959, HY-112149, HY-101146, HY-111102, HY-112718, HY-16586, HY-16954, HY-50698, H Y-103633, HY-101838, HY-125232, HY-126428, HY-100653A, MCE Company), and then through qPCR experiments, it was found that 5 small molecule inhibitors (HY-13960, HY-111422, HY-112610, HY-13235, HY-100015, MCE Company) had a significant and effective inhibitory effect on the expression of CREB5. The results are as follows Figure 3 Finally, the cell killing effects of these five small molecule inhibitors on DIPG cells and PPC cells were tested, and the results were as follows Figure 4 As shown, it was found that only the fifth small molecule inhibitor (HY-100015, also known as ABBV-075) could significantly inhibit the growth of DIPG cells and had no obvious toxic side effects on PPC cells.

[0090] Embodiment 3:

[0091] In order to improve the clinical application potential of ABBV-075 in the treatment of DIPG, this example needs to further clarify the mechanism of action of ABBV-075 in inhibiting CREB5 expression by inhibiting CREB5 super enhancer activity. Figure 5 As shown, it was found that ABBV-075 and JQ1 have similar abilities to bind to the WPF domain of BRD4 protein, and it is predicted that ABBV-075 can inhibit the ability of BRD4 to recognize histone acetylation.

[0092] Therefore, in the following example, a protein immunoblot experiment (Western Blot) was performed to find that the H3K27ac histone modification level can be significantly downregulated with different time and concentration of ABBV-075 treatment. Figure 6 and Figure 7 In this process, this example found that ABBV-075 not only down-regulated the protein level of CREB5, but also down-regulated the protein levels of OLIG2 and ID1, inducing DIPG cells to develop a neurite-like differentiation phenotype. The results are shown in Figure 8 shown.

[0093] The above results demonstrate that ABBV-075 can serve as an effective small molecule inhibitor of BRD4, which can inhibit the expression of CREB5 by reducing the level of H3K27ac and further promote the differentiation of DIPG cells.

[0094] At the same time, this example also constructs a dual fluorescence reporter system of CREB5 super enhancers and common enhancers and their promoters to evaluate the specific molecular mechanism of ABBV-075 on CREB5. Consistent with the phenomenon that CREB5 has common super enhancers and highly active transcription in DIPG cells, compared with common enhancers, CREB5 super enhancers can significantly promote CREB5 promoter activity. The results are as follows Fig. 9 After treatment with ABBV-075, the promotion effect of CREB5 super enhancer on its promoter activity was significantly reduced, indicating that ABBV-075 can inhibit the expression of CREB5 by inhibiting the activity of CREB5 super enhancer and promoter. Fig.10 Using the concentration inhibition curve, this example found that ABBV-075 can significantly inhibit the activity of the CREB5 enhancer at the level of 20 nM. The results are shown in Fig.11 As shown, ABBV-075 is a very potent and effective BRD4 small molecule inhibitor for the treatment of DIPG, further emphasizing its higher inhibitory concentration and toxic side effects compared to other BRD4 small molecule inhibitors.

[0095] Embodiment 4:

[0096] This example first evaluated the therapeutic effect of ABBV-075 on DIPG cells in vitro. This example used JQ1 as a positive control, and used growth curve and spheroidization experiments to detect the killing effect of ABBV-075 on different DIPG cells. The results of this example found that ABBV-075 had less toxic side effects on normal PPC cells than JQ1, but had a more effective killing effect on DIPG glioma cells and a lower IC50 value. The results are as follows: Fig.12This indicates that ABBV-075 has a highly selective killing ability against DIPG cells compared to JQ1, and has lower toxicity to PPC cells that are similar to DIPG cells.

[0097] Therefore, this example further evaluated the anti-tumor effect of ABBV-075 in vivo on PDX mice injected with TT150630 cells in situ with DIPG. In this example, TT150630 cells derived from DIPG patients were first injected into the pons of immunodeficient mice (NCG mice) (5 control groups, 5 drug-treated groups, a total of 10 mice; the mice were from Nanjing Jicui Yaokang, NOD / ShiLtJGpt-Prkdcem26Cd52II2rgem26Cd22 / Gpt (NCG mice) and allowed to form tumors for 14 days. Then, the mice were given the drug by oral gavage every three days, with a drug concentration of 1 mg / kg, for a total of 8 times, and the growth of tumor cells was detected by in vivo fluorescence imaging every week. Fig.13 As shown, the results of bioluminescence imaging showed that the growth of DIPG tumor-bearing mice in the ABBV-075-treated group was significantly inhibited compared with the saline control group, and their survival period was significantly longer. These results indicate that ABBV-075 can significantly inhibit the growth of DIPG tumors in vivo.

[0098] Considering that CREB5 in DIPG can coordinate with the catalytic subunit BRG1 of the SWI / SNF chromatin remodeling complex to promote cancer, and the inhibitor of BRG1 has been disclosed in the prior art to have anti-tumor effects on DIPG cells, this example also evaluates the drug treatment effects of ABBV-075 and BRG1 small molecule inhibitor (BRM014) on DIPG cells. By using the combined index model and the Bliss independent model, this example evaluates the combined therapeutic effect of ABBV-075 and BRM014 on DIPG17 and TT150714 cells. At lower concentrations, the combined treatment showed a very significant inhibitory effect in both DIPG cells. By using the compusyn software to calculate the therapeutic index of each combination, this example identified the synergistic treatment area in the combined treatment matrix. Further, by calculating the comprehensive synergistic treatment score of the two small molecule inhibitors, this example found that both DIPG17 (Bliss: 12.04) and TT150714 (Bliss: 16.01) cells showed very significant synergistic treatment effects, as shown in the following results. Fig.14Therefore, based on all the above results, this example found that ABBV-075 can exert its anti-tumor effect by inhibiting the activity of CREB5 super enhancer, and its toxicity to normal neural stem cells is low. In addition, by combining ABBV-075 and a small molecule inhibitor of BRG1, it was found that there was a significant combined therapeutic effect on H3.3K27M DIPG cells.

[0099] In the description of this specification, the description with reference to the terms "one embodiment", "some embodiments", "example", "specific example", or "some examples" etc. means that the specific features, structures, materials or characteristics described in conjunction with the embodiment or example are included in at least one embodiment or example of the present invention. In this specification, the schematic representations of the above terms do not necessarily refer to the same embodiment or example. Moreover, the specific features, structures, materials or characteristics described may be combined in any one or more embodiments or examples in a suitable manner. In addition, those skilled in the art may combine and combine the different embodiments or examples described in this specification and the features of the different embodiments or examples, without contradiction.

[0100] Although the embodiments of the present invention have been shown and described above, it is to be understood that the above embodiments are exemplary and are not to be construed as limitations of the present invention. A person skilled in the art may change, modify, replace and vary the above embodiments within the scope of the present invention.

Claims

1. A method for killing brain tumor cells in vitro, characterized in that: include: ABBV-075 is contacted with isolated brain tumor cells to kill the brain tumor cells.

2. A method for targeting brain tumor cells in vitro, characterized in that: include: ABBV-075 is contacted with an isolated cell population comprising brain tumor cells.

3. The method according to claim 1 or 2, characterized in that: include: The concentration of ABBV-075 is 1nM to 1000nM; Optionally, the brain tumor occurs in at least one of the spinal cord, brain stem, pineal region, and thalamus; Optionally, the brain tumor cells include H3.3K27M mutant diffuse intrinsic pontine glioma cells.

4. The method according to claim 2, characterized in that: The cell population further includes normal brain cells; Optionally, the normal brain cells include at least one of glial cells and neuronal cells.

5. The method according to claim 1, characterized in that The method further comprises contacting the isolated brain tumor cells with the BRG1 inhibitor; Optionally, the BRG1 inhibitor comprises BRM014.

6. Use of a reagent in preparing a kit, wherein the reagent comprises ABBV-075, and the kit is used to inhibit the expression of CREB5 and / or ID1.

7. Use of an agent in preparing a drug, wherein the agent includes ABBV-075, and the drug is used to treat or prevent glioma.

8. The use according to claim 7, characterized in that The reagent further comprises a BRG1 inhibitor; Optionally, the BRG1 inhibitor comprises BRM014; Optionally, the glioma occurs in at least one of the spinal cord, brain stem, pineal region, and thalamus; Optionally, the glioma comprises H3.3K27M mutant diffuse intrinsic pontine glioma.

9. A pharmaceutical composition for treating or preventing glioma, characterized in that: include: ABBV-075; Preferably, further comprising: a BRG1 inhibitor; Optionally, the BRG1 inhibitor comprises BRM014; Optionally, the ABBV-075 and the BRG1 inhibitor are suitable for simultaneous or sequential administration; Optionally, the glioma occurs in at least one of the spinal cord, brain stem, pineal region, and thalamus; Preferably, the glioma comprises H3.3K27M mutant diffuse intrinsic pontine glioma.

10. A method for inhibiting CREB5 expression, characterized in that: ABBV-075 is contacted with isolated cells expressing CREB5 to inhibit the expression of CREB5.

11. The method according to claim 10, characterized in that The cells expressing CREB5 include brain tumor cells; Optionally, the brain tumor occurs in at least one of the spinal cord, brain stem, pineal region, and thalamus; Optionally, the brain tumor cells include H3.3K27M mutant diffuse intrinsic pontine glioma cells.

12. The method according to claim 10, characterized in that The expression of CREB5 is inhibited by at least one of the following methods: The activity of the CREB5 super-enhancer is inhibited; The level of H3K27ac histone modification was reduced.

13. A method for promoting the differentiation of brain tumor cells into astrocytes, characterized in that: include: ABBV-075 is contacted with isolated brain tumor cells to promote the differentiation of the brain tumor cells into astrocytes.

14. The method according to claim 13, characterized in that The brain tumor cells include H3.3K27M mutant diffuse intrinsic pontine glioma cells; Optionally, the brain tumor occurs in at least one of the spinal cord, brain stem, pineal region and thalamus.

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