A polypeptide that targets and blocks the binding of TRIM25 to BRD7 protein and its application

Abstract

The present invention belongs to the field of biomedicine and specifically relates to a polypeptide that targets and blocks the binding of TRIM25 to the BRD7 protein and its application. Based on the PRYSPRY domain (439 aa-630 aa), the minimal binding region of TRIM25 and BRD7 proteins, the present invention screened and obtained a blocking peptide TB16 that can specifically block the binding of TRIM25 and BRD7 proteins, thereby inhibiting the ubiquitination and degradation of BRD7 and increasing the stability of the BRD7 tumor suppressor protein. The polypeptide drug TB16 described in the present invention can effectively inhibit the proliferation of breast cancer, nasopharyngeal cancer, ovarian cancer, lung cancer, and liver cancer cells, as well as the growth of breast cancer tumors in vivo. It has excellent activity against solid tumors and is expected to become a new targeted anti-tumor drug.

Description

Technical Field

[0001] The present invention belongs to the field of biomedicine, and specifically relates to a polypeptide that targets and blocks the binding of TRIM25 to BRD7 protein, and its application in anti-tumor and BRD7 protein stabilization. Background Art

[0002] Commonly used anti-tumor drugs in clinical practice suffer from shortcomings such as weak targeting, significant adverse reactions, and the development of drug resistance. Compared to traditional anti-tumor drugs, peptides offer advantages such as smaller molecular weight, stronger targeting, higher activity, lower toxicity, and easier transmembrane absorption. They can act directly or indirectly on tumor cells, regulating biological functions such as tumor growth and apoptosis. Furthermore, solid-phase chemical synthesis of peptides reduces production costs compared to other biopharmaceuticals, facilitating large-scale production. Peptide-based therapies are not only highly effective and low-toxic, but can also increase tumor sensitivity to other treatments. They hold significant value in the clinical treatment of tumors and are currently a hot topic of research.

[0003] BRD7 is a lowly expressed gene in nasopharyngeal carcinoma (NPC) isolated and cloned through methods such as cDNA representational differential analysis and library screening. BRD7 has been shown to be downregulated in various tumors and functions as a tumor suppressor gene in cancers such as NPC, breast cancer, ovarian cancer, lung cancer, liver cancer, and osteosarcoma, and is associated with poor patient prognosis. As a tumor suppressor, BRD7 plays a key role in various biological processes, including cell proliferation, cell cycle progression, apoptosis, invasion, and migration, by regulating gene expression.

[0004] The present invention discovered a potential interaction between TRIM25 and BRD7, suggesting that TRIM25 may contribute to the malignant progression of various tumors by reducing BRD7 protein stability. Therefore, developing a peptide drug that blocks the TRIM25 / BRD7 interaction could stabilize BRD7 protein in tumor cells, thereby exerting its tumor suppressor function, and is a promising clinical treatment strategy. The development of this peptide drug is expected to fill the gap in the field of peptide-based tumor drugs targeting this mechanism and holds great promise for future applications. Summary of the Invention

[0005] In view of this, the present invention aims to provide a polypeptide that blocks the interaction between TRIM25 and BRD7 and its use. This polypeptide can target and stabilize the BRD7 protein to achieve anti-tumor effects, representing an effective and feasible strategy for treating solid tumors. It is capable of treating a variety of solid tumors, including breast cancer, nasopharyngeal cancer, ovarian cancer, lung cancer, and liver cancer.

[0006] In a first aspect of the present invention, an anti-tumor polypeptide is provided that targets and blocks the TRIM25 / BRD7 interaction, thereby stabilizing the BRD7 protein. The amino acid sequence of the polypeptide is: KVLETFLAKSRPELLE.

[0007] Furthermore, it includes the polypeptide and a pharmaceutically acceptable modification type, wherein the polypeptide modification type includes at least one of C-terminal modification, N-terminal modification, intermediate residue modification and cyclization modification.

[0008] In the present invention, based on the TRIM25 and BRD7 interaction domain PRYSPRY, a polypeptide that targets and blocks the binding of TRIM25 to BRD7 protein and can stabilize the BRD7 protein is screened and prepared as an anti-tumor drug.

[0009] Specifically, the polypeptide blocks the binding of the PRYSPRY domain of TRIM25 to BRD7, inhibits TRIM25-mediated ubiquitination and degradation of BRD7, and achieves stabilization of BRD7 protein in breast cancer, nasopharyngeal cancer, ovarian cancer, lung cancer and liver cancer, and is used as a tumor treatment drug.

[0010] The second aspect of the present invention provides a use of the above-mentioned polypeptide in the preparation of a drug or preparation for preventing and / or treating solid tumors.

[0011] The solid tumor is a solid tumor with low BRD7 expression, including one or more of breast cancer, nasopharyngeal cancer, ovarian cancer, lung cancer and liver cancer.

[0012] Breast cancer cells include at least one of MDA-MB-231 and MCF7; nasopharyngeal cancer cells include CNE2; ovarian cancer cells include A2780; lung cancer cells include PC9; and liver cancer cells include Hep3B.

[0013] Furthermore, the medicine also includes pharmaceutically acceptable excipients.

[0014] In a third aspect, the present invention provides a use of the above-mentioned polypeptide in the preparation of a BRD7 tumor suppressor protein stabilizing drug or preparation, wherein the polypeptide can increase the stability of the BRD7 protein by blocking the binding of TRIM25 to the BRD7 protein.

[0015] Furthermore, the drug further comprises pharmaceutically acceptable excipients, wherein the excipients include conventional diluents, fillers, adhesives, wetting agents, absorption enhancers, surfactants, lubricants and stabilizers in the pharmaceutical field.

[0016] In the applications described herein, the drug dosage form is an oral dosage form and / or an injectable dosage form. These include, but are not limited to, tablets, pills, capsules, sprays, granules, powder injections, and injection solutions. The dosage of the drug is a pharmaceutically acceptable dose. Administration methods include, but are not limited to, tumor injection, intravenous injection, or intraperitoneal injection, as well as oral administration such as pills and capsules.

[0017] Preferably, the drug is in a pharmaceutically acceptable dosage form.

[0018] The present invention uses the analysis of the α-helix in the TRIM25-PRYSPRY domain that binds to BRD7 to synthesize two possible blocking peptide sequences that have the ability to block the binding of TRIM25 to BRD7; the two blocking peptides are used in in vitro experiments to screen out polypeptides that have the ability to target and stabilize the BRD7 protein.

[0019] Beneficial effects of the present invention

[0020] The present invention provides an anti-tumor peptide, TB16, that specifically blocks the TRIM25 / BRD7 interaction. It can specifically block the interaction between TRIM25 and BRD7, reversing the elevated TRIM25-mediated BRD7 ubiquitination, thereby stabilizing the BRD7 protein. Biological phenotypes confirm the anti-tumor effects and biosafety of this peptide drug in breast cancer, nasopharyngeal carcinoma, ovarian cancer, lung cancer, and liver cancer. Therefore, TB16 can be used to prepare a potential anti-solid tumor drug for the treatment of multiple tumors, including breast cancer, nasopharyngeal carcinoma, ovarian cancer, lung cancer, and liver cancer, by targeting and stabilizing the BRD7 protein. BRIEF DESCRIPTION OF THE DRAWINGS

[0021] Figure 1 : TRIM25 negatively regulates BRD7 protein stability through the ubiquitin-proteasome pathway;

[0022] Figure 1 A: Silencing TRIM25 expression in breast cancer MDA-MB-231 and MCF7 cells increased BRD7 protein stability.

[0023] Figure 1 B: Treatment with MG132 (a proteasome inhibitor) can reverse the downregulation of BRD7 protein mediated by TRIM25 overexpression in breast cancer MDA-MB-231 and MCF7 cells;

[0024] Figure 1 C: Effect of TRIM25 silencing expression on the ubiquitination level of BRD7 in breast cancer MDA-MB-231 and MCF7 cells.

[0025] Figure 2: The results show that BRD7 binds to the PRYSPRY domain of TRIM25;

[0026] Figure 2 A: Schematic diagram of the Flag-TRIM25 domain deletion vector;

[0027] Figure 2 B: Schematic diagram of the co-immunoprecipitation (Co-IP) assay detecting the interaction between the TRIM25 PRYSPRY domain and BRD7 in breast cancer cells MCF7;

[0028] Figure 2 C: Co-IP experimental results showing that the TRIM25-PRYSPRY domain binds to BRD7 in breast cancer cells MCF7;

[0029] Figure 2 D: Schematic diagram of the binding between BRD7 and TRIM25-PRYSPRY verified by molecular docking using the HDOCK website (http: / / hdock.phys.hust.edu.cn).

[0030] Figure 3 : Schematic diagram of the design and synthesis of α-helical peptides based on the PRYSRY binding region of BRD7 and TRIM25;

[0031] Figure 3 A: Schematic diagram of the protein structure of TRIM25-PRYSPRY from the PDB website (https: / / www.rcsb.org);

[0032] Figure 3 B: Schematic diagram of analyzing the α-helix present in the TRIM25-PRYSPRY domain and designing and synthesizing two blocking peptides based on its amino acid sequence.

[0033] Figure 4 TB16, a peptide drug derived from the TRIM25 PRYSPRY domain, competitively inhibits the binding of TRIM25 to BRD7 protein and increases the stability of BRD7 protein.

[0034] Figure 4 A: Western Blot analysis of the effects of TB5 and TB16 on BRD7 protein expression in breast cancer cells MDA-MB-231.

[0035] Figure 4 B: Western Blot analysis of the effects of TB5 and TB16 on BRD7 protein expression in breast cancer cells MCF7.

[0036] Figure 4C: Co-IP detection of the effects of TB5 and TB16 on the binding ability of TRIM25 and BRD7 proteins in breast cancer cells MCF7;

[0037] Figure 4 D: Western Blot analysis of the effects of TB5 and TB16 on TRIM25-mediated ubiquitination of BRD7 protein in breast cancer cells MCF7.

[0038] Figure 5 : The results show that the TAT-modified peptide TAT-TB16 can increase the stability of BRD7 protein in nasopharyngeal carcinoma, ovarian cancer, liver cancer and lung cancer;

[0039] Figure 5 A: Western Blot analysis of the effect of TAT-TB16 on BRD7 protein expression in nasopharyngeal carcinoma cell lines CNE2.

[0040] Figure 5 B: Western Blot analysis of the effect of TAT-TB16 on BRD7 protein expression in ovarian cancer cells A2780;

[0041] Figure 5 C: Western Blot analysis of the effect of TAT-TB16 on BRD7 protein expression in lung cancer PC9 cells;

[0042] Figure 5 D: Western Blot detection of the effect of TAT-TB16 on BRD7 protein expression in liver cancer cells Hep3B.

[0043] Figure 6 : The results of the peptide drug TAT-TB16 exerting anti-tumor effects in breast cancer, nasopharyngeal cancer, ovarian cancer, liver cancer and lung cancer;

[0044] Figure 6 A: CCK8 assay to detect the effect of TAT-TB16 on the proliferation of breast cancer cells (MDA-MB-231, MCF7), nasopharyngeal carcinoma cells (CNE2), ovarian cancer cells (A2780), lung cancer cells (PC9), and liver cancer cells (Hep3B);

[0045] Figure 6 B: Effect of TAT-TB16 on the clone-forming ability of breast cancer cells MDA-MB-231 and MCF7.

[0046] Figure 7 : The peptide drug TAT-TB16 can significantly inhibit the growth of breast cancer transplanted tumors in vivo;

[0047] Figure 7A: Photos of tumor-bearing mice in the normal saline group, TAT group, and TAT-TB16 group;

[0048] Figure 7 B: After the mice were sacrificed, the transplanted tumors in the saline group, TAT group, and TAT-TB16 group were removed and compared;

[0049] Figure 7 C: The size of transplanted tumors was measured every 2 days, and the mouse tumor growth curve was drawn;

[0050] Figure 7 D: Comparison of transplanted tumor weights in each group;

[0051] Figure 7 E: Body weight changes of mice in the normal saline group, TAT group and TAT-TB16 group.

[0052] Figure 8 : The peptide drug TAT-TB16 has no obvious toxic side effects on the important organs of mice;

[0053] The data are presented as mean ± standard error. , p<0.05; , p<0.01; , p<0.001;ns, no statistical difference. DETAILED DESCRIPTION

[0054] The technical solutions of the present invention will be further described in detail below with reference to the accompanying drawings and specific embodiments, but the present invention is not limited to the following technical solutions.

[0055] The MDA-MB-231, MCF7, CNE2, A2780, PC9, and Hep3B cell lines used in this study were all maintained in the Molecular Genetics Laboratory of the Cancer Research Institute of Central South University. MDA-MB-231, MCF7, A2780, and Hep3B cells were cultured in DMEM liquid medium supplemented with 10% fetal bovine serum (FBS) and 1% double-streptomycin (penicillin and streptomycin), allowing adherent growth in a constant-temperature incubator at 37°C and 5% CO2. CNE2 and PC9 cells were cultured in 1640 liquid medium supplemented with 10% fetal bovine serum (FBS) and 1% double-streptomycin (penicillin and streptomycin), allowing adherent growth in a constant-temperature incubator at 37°C and 5% CO2.

[0056] Example 1: TRIM25 negatively regulates BRD7 protein stability through the ubiquitin-proteasome pathway

[0057] 1.1 Experimental plan:

[0058] (1) siNC and siTRIM25 were transfected into MDA-MB-231 and MCF7 cells, respectively. The cells were then treated with CHX (50 μM). The cells were collected at different time points (0, 1, 2, and 4 h) and protein extraction was performed immediately. The protein expression level of BRD7 was detected by Western Blot technology.

[0059] (2) The blank vector and TRIM25 plasmid were transfected into MDA-MB-231 and MCF7 cells. 24 h after transfection, the cells were treated with the proteasome inhibitor MG132 (20 μM) for 4 h. The cells were collected and protein extraction was performed immediately. The protein expression level of BRD7 was detected by Western Blot technology.

[0060] (3) siNC, siTRIM25#1, and siTRIM25#2 were transfected into MDA-MB-231 and MCF7 cells, respectively. 48 h after transfection, cells were treated with 20 µM proteasome inhibitor MG132 for 4 h, then harvested and immediately subjected to protein extraction. Co-IP experiments were performed using a BRD7 antibody, and the ubiquitination level of BRD7 was detected by Western blotting.

[0061] 1.2 Experimental results:

[0062] The experimental results showed that silencing TRIM25 expression in two breast cancer cells could significantly prolong the half-life of BRD7 protein ( Figure 1 A). In addition, MG132 treatment reversed the inhibitory effect of TRIM25 overexpression on BRD7 protein levels, indicating that TRIM25 promotes BRD7 protein degradation through the proteasome pathway ( Figure 1 B). Ubiquitination assay results showed that TRIM25 silencing expression reduced the ubiquitination level of BRD7 in MDA-MB-231 and MCF7 cells ( Figure 1 C). The above results indicate that TRIM25 reduces the protein stability of BRD7 and promotes its ubiquitination degradation in a ubiquitin-proteasome-dependent manner.

[0063] Example 2, BRD7 binds to the PRYSPRY domain of TRIM25

[0064] 2.1 Experimental plan:

[0065] (1) Based on the protein domain structure of TRIM25, a series of Flag-TRIM25 domain deletion vectors were constructed, including Flag-TRIM25-△RING (RING domain deletion), Flag-TRIM25-△Coiled-Coil (Coiled-Coil domain deletion), and Flag-TRIM25-△PRYSPRY (PRYSPRY domain deletion). Flag-TRIM25 series truncations and HA-BRD7 were co-transfected into the breast cancer cell line MCF7. Co-IP experiments were performed using HA antibodies to explore the minimal domain for the interaction between TRIM25 and BRD7. At the same time, the binding of BRD7 and TRIM25-PRYSPRY was verified by molecular docking using the HDOCK website (http: / / hdock.phys.hust.edu.cn).

[0066] (2) A Flag-TRIM25 PRYSPRY vector was constructed and Flag-TRIM25 PRYSPRY and HA-BRD7 were co-transfected into the breast cancer cell line MCF7. HA antibody and Flag antibody were used for forward and reverse Co-IP experiments to explore the interaction between TRIM25 PRYSPRY and BRD7.

[0067] 2.2 Experimental results:

[0068] Co-IP and Western Blot results showed that BRD7 interacts with the PRYSPRY domain of TRIM25 ( Figure 2 AC). Molecular docking was used to further confirm the interaction between BRD7 and TRIM25-PRYSPRY using the HDOCK website (http: / / hdock.phys.hust.edu.cn). Figure 2 D). These results indicate that TRIM25-PRYSPRY interacts with BRD7, providing a structural basis for the development of peptide drugs based on the TRIM25 / BRD7 regulatory axis.

[0069] Example 3: Design and synthesis of α-helical peptides based on the PRYSRY binding region of BRD7 and TRIM25

[0070] 3.1 Experimental plan:

[0071] To further design peptides targeting the TRIM25 / BRD7 interaction, we focused on protein structure. The α-helix, a crucial component of protein secondary structure, imparts stability and morphology through stable hydrogen bonds and helical morphology, participating in molecular recognition and binding. Therefore, analyzing α-helices within protein domains and synthesizing α-helical peptides is an important approach and tool for designing peptide drugs that block protein-target interactions. Based on the crystal structure of TRIM25 PRYSPRY (PDB: 6FLM), we analyzed the α-helix within the TRIM25 PRYSPRY domain and further synthesized α-helical peptides.

[0072] 3.2 Experimental results:

[0073] We used the PDB website (https: / / www.rcsb.org) to identify the protein structure of TRIM25-PRYSPRY ( Figure 3 A), the α-helix present in the TRIM25-PRYSPRY domain was analyzed and two α-helices were found. The first α-helix is located at 439aa-454aa, with the amino acid sequence KVLETFLAKSRPELLE, and the second α-helix is located at 537aa-541aa, with the amino acid sequence GPESR ( Figure 3 B), we synthesized two peptides based on these two amino acid sequences and named them TB16 (439aa-454aa) and TB5 (537aa-541aa).

[0074] Example 4: TB16, a peptide drug derived from the TRIM25 PRYSPRY domain, competitively inhibits the binding of TRIM25 to BRD7 and increases BRD7 protein stability

[0075] 4.1 Experimental plan:

[0076] (1) Breast cancer cells MDA-MB-231 and MCF7 were treated with different concentrations of TB5 and TB16 (0 µM, 5 µM, 10 µM, 20 µM, and 40 µM), respectively. After 24 h of treatment, the cells were harvested and protein extraction was performed immediately. The protein expression level of BRD7 was detected by Western Blot.

[0077] (2) HA-BRD7 and Flag-TRIM25 plasmids were co-transfected into MCF7 breast cancer cells. 24 h after transfection, the cells were treated with TB5 (20 μM) and TB16 (20 μM), respectively. After a further 24 h of treatment, Co-IP experiments were performed using Flag antibodies to investigate the effects of TB5 and TB16 on the binding ability of TRIM25 to BRD7.

[0078] (3) HA-BRD7, Flag-TRIM25, and Ub plasmids were co-transfected into MCF7 breast cancer cells. 24 h after transfection, the cells were treated with TB5 (20 μM) and TB16 (20 μM) for 24 h, respectively. Subsequently, the cells were treated with 20 μM MG132 for 4 h. Co-IP experiments were performed using BRD7 antibody, and the ubiquitination level of BRD7 was detected by Western Blot technology.

[0079] 4.2 Experimental Results

[0080] The results showed that in two breast cancer cell lines, MDA-MB-231 and MCF7, different concentrations of TB5 had no significant effect on the protein expression level of BRD7, while with the increase of TB16 concentration, the protein expression level of BRD7 gradually increased ( Figure 4 AB). The results of the CO-IP experiment showed that in MCF7 cells, the protein binding ability of TRIM25 and BRD7 was significantly weakened in the TB16-treated group compared with the TB5-treated group ( Figure 4 C). In addition, in MCF7 cells, TRIM25 overexpression can promote the ubiquitination level of BRD7, while TB16 can reverse the promoting effect of TRIM25 overexpression on the ubiquitination level of BRD7 ( Figure 4 D). These results indicate that TB16 can inhibit the protein interaction between TRIM25 and BRD7 in breast cancer cells, thereby suppressing the TRIM25-mediated increase in BRD7 ubiquitination levels and thereby targeting and stabilizing the BRD7 protein.

[0081] Example 5: TAT-TB16, a TAT-modified peptide of the peptide drug TB16, can increase the stability of BRD7 protein in nasopharyngeal carcinoma, ovarian cancer, liver cancer, and lung cancer.

[0082] In order to increase the cell membrane permeability of TB16, we introduced a cell-penetrating peptide TAT (amino acid sequence: GRKKRRQRRRPPQ) before the amino acid sequence of TB16 and further synthesized TAT-TB16.

[0083] 5.1 Experimental plan:

[0084] Nasopharyngeal carcinoma CNE2, ovarian cancer A2780, liver cancer PC9, and lung cancer Hep3B cells were treated with various concentrations of TAT-TB16 (0 µM, 10 µM, 20 µM, and 40 µM). Cells were harvested 24 hours after treatment and immediately subjected to protein extraction. BRD7 protein expression was assessed by Western blotting.

[0085] 5.2 Experimental Results

[0086] Western Blot results showed that in nasopharyngeal carcinoma cell CNE2, ovarian cancer cell A2780, liver cancer cell PC9 and lung cancer cell Hep3B, the protein expression level of BRD7 gradually increased with the increase of TAT-TB16 treatment concentration ( Figure 5 AD). The above results indicate that TAT-TB16 can target and stabilize BRD7 protein in various tumor cells.

[0087] Example 6: The peptide drug TAT-TB16 can exert anti-tumor effects in in vitro cultured cells of breast cancer, nasopharyngeal cancer, ovarian cancer, liver cancer and lung cancer.

[0088] 6.1 Experimental plan:

[0089] CCK8 experiment:

[0090] (1) Using a variety of in vitro cultured cells, including breast cancer cells (MDA-MB-231, MCF7), nasopharyngeal carcinoma cells (CNE2), ovarian cancer cells (A2780), liver cancer cells (PC9), and lung cancer cells (Hep3B), different tumor cells were seeded into 96-well plates at 1000 cells / well.

[0091] (2) After the cells were fully attached, MDA-MB-231, MCF7, CNE2, A2780, PC9 and Hep3B cells were treated with 100 μM TAT-TB16. The control group was the TAT-treated group.

[0092] (3) Set 6 time points: 0 d, 1 d, 2 d, 3 d, 4 d, and 5 d. Then add 10% CCK8 solution to each well and incubate in a 37°C incubator for 2 h. Use a microplate reader (wavelength 450 nm) to measure the absorbance value.

[0093] (4) Data analysis and plotting: Cell proliferation curves were plotted using GraphicPad Prism 9.5.0 software.

[0094] Colony formation assay:

[0095] (1) Breast cancer cells (MDA-MB-231, MCF7) with good growth status were counted and plated in 12-well plates at 500 cells / well. After small clones grew, the tumor cells were co-cultured with TAT (100 μM) or TAT-TB16 (100 μM) for 3 days, and then replaced with complete culture medium and continued to be cultured for 10-14 days.

[0096] (2) Discard the culture medium and wash the cells with 1×PBS three times for 5 minutes each time. Then add 4% paraformaldehyde and fix at room temperature for 1 hour.

[0097] (3) Aspirate the fixative and rinse three times with 1× PBS for 5 minutes each. Then add crystal violet stain to each well and stain at room temperature for 30 minutes.

[0098] (4) Aspirate the crystal violet staining solution and rinse with distilled water until no crystal violet residue remains. After drying, use a scanner to scan and record the image.

[0099] (5) Count the number of effective cell clones in each group and analyze the data.

[0100] 6.2 Experimental Results

[0101] The results of CCK8 experiments showed that, compared with the TAT group, TAT-TB16 could inhibit the cell proliferation of breast cancer, nasopharyngeal cancer, ovarian cancer cells, liver cancer cells and lung cancer cells ( Figure 6 A); The results of the clone formation experiment showed that TAT-TB16 could significantly inhibit the clone formation ability of breast cancer cells compared with the TAT group ( Figure 6 B). The above results indicate that TAT-TB16 exerts anti-tumor effects in a variety of tumors, including breast cancer, nasopharyngeal cancer, ovarian cancer, liver cancer, and lung cancer.

[0102] Example 7: The peptide drug TAT-TB16 can significantly inhibit the growth of breast cancer transplanted tumors in vivo without obvious tissue and organ toxicity

[0103] 7.1 Experimental plan:

[0104] Twenty-one 4- to 6-week-old female BALB / C nude mice were purchased from Hunan Slake Jingda Laboratory Animal Co., Ltd. All nude mice passed quality inspection. Animal husbandry and related operations were performed under specific pathogen-free (SPF) conditions at the Animal Experiment Center of Hunan Cancer Hospital.

[0105] (1) Prepare breast cancer cells MCF7 in good growth condition, collect the cells and wash them three times with pre-cooled saline.

[0106] (2) Prepare 3×10 6150 μL of cell suspension containing 10 cells and 50 μL of matrix gel was inoculated into the subcutaneous area above the axilla of the right forelimb of nude mice.

[0107] (3) Starting from the fourth day after injection, the length (L) and width (W) of the subcutaneous tumor of the nude mouse were measured every two days with a vernier caliper. The results were recorded and the tumor volume was calculated using the following formula: Volume = L × W 2 / 2; the weight of mice was recorded at the same time.

[0108] (4) When the tumor volume of the mouse reaches 30-50 mm 3 Afterwards, the mice were randomly divided into three groups, with seven mice in each group. Each mouse in the experimental group received an intratumoral injection of 15 mg / kg of TAT or TAT-TB16, while the control group received a saline injection. The injections were given every two days for a total of six times.

[0109] (5) On the 18th day of feeding, the mice were euthanized by cervical dislocation, the tumors were removed, photographed, and the tumor volume and weight were recorded.

[0110] (6) Three mice were randomly selected from each group. The heart, liver, spleen, lung, and kidney were removed, dehydrated, fixed, paraffin-embedded, and sliced. HE staining was then performed to observe the tissue status of each organ.

[0111] 7.2 Experimental Results

[0112] The results of the in vivo nude mouse transplant tumor model showed that compared with the saline group and the TAT group, the tumor growth of the mice in the TAT-TB16 treatment group was significantly inhibited and the tumor weight was significantly reduced ( Figure 7 AD), indicating that TAT-TB16 can significantly inhibit tumor growth in vivo, thus showing an anti-tumor effect in vivo. In addition, the body weight of mice in each group was not affected during the treatment ( Figure 7 E). According to the results of HE staining, there was no obvious damage to the organs and tissues of the mice in the TAT-TB16 treatment group, indicating that TAT-TB16 has a certain degree of biosafety in vivo ( Figure 8 The above results show that TAT-TB16 can significantly inhibit the growth of breast cancer cells without obvious organ toxicity.

Claims

1. A polypeptide that targets and blocks the binding of TRIM25 to BRD7 protein and stabilizes BRD7 protein, characterized in that: The amino acid sequence of the polypeptide is: KVLETFLAKSRPELLE.

2. Use of the polypeptide according to claim 1 in the preparation of a drug for preventing and / or treating solid tumors; the solid tumor is one or more of breast cancer, nasopharyngeal cancer, ovarian cancer, lung cancer and liver cancer.

3. The use according to claim 2, characterized in that Breast cancer cells include at least one of MDA-MB-231 and MCF7; nasopharyngeal cancer cells include CNE2; ovarian cancer cells include A2780; lung cancer cells include PC9; and liver cancer cells include Hep3B.

4. The use according to claim 2, characterized in that The medicine also includes pharmaceutically acceptable excipients.

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