A MEIS1-derived peptide and its use in preparing a drug for treating colorectal cancer

Abstract

This invention provides a MEIS1-derived peptide and its application in the preparation of drugs for treating colorectal cancer, belonging to the field of biomedical technology. The amino acid sequence of the MEIS1-derived peptide is SEQ ID NO: 1. In vitro experiments (CCK8 and plate cloning experiments) and in vivo animal experiments demonstrated that the MEIS1-derived peptide can significantly inhibit the proliferation of colorectal cancer cells, providing a theoretical basis and experimental evidence for the preparation of drugs for treating colorectal cancer patients and the development of new and effective targeted drug therapies.

Description

Technical Field

[0001] This invention relates to the field of biomedical technology, and in particular to a MEIS1-derived peptide and its application in the preparation of drugs for treating colorectal cancer. Background Art

[0002] Colorectal cancer (CRC) is a common malignant tumor of the digestive system, attracting widespread attention globally. In my country, the incidence of CRC continues to rise rapidly, ranking fifth in mortality among malignant tumors. Due to the lack of effective diagnostic methods, prognostic markers, and early recurrence, the prognosis for patients with advanced CRC remains far from satisfactory. Currently, treatment for CRC mainly includes surgery, radiotherapy, and chemotherapy. However, some patients are diagnosed at an advanced stage, limiting the possibility of surgical resection. Furthermore, traditional treatments have limited effectiveness for metastatic CRC. While immunotherapy has emerged as a new treatment approach, it is still in the research stage, and its long-term efficacy and safety in CRC treatment require further validation. Despite continuous efforts in research, clinical diagnosis, and personalized treatment over a considerable period, persistent tumor proliferation remains the most common cause of cancer-related death worldwide. Therefore, further screening of genes affecting CRC growth and the identification and development of specific targeted drugs for key genes are crucial for improving the survival of CRC patients and reducing the overall mortality rate of CRC.

[0003] Peptide drugs are short-chain proteins composed of amino acids, possessing a wide range of biological activities and pharmacological effects. Due to their ability to precisely bind to target proteins, peptide drugs exhibit broad application potential in the medical field. With the continuous development of science and technology, the application areas of peptide drugs are constantly expanding. In cancer treatment, some peptide drugs have shown good efficacy through mechanisms such as inhibiting tumor growth, promoting apoptosis, or blocking angiogenesis. Furthermore, peptide drugs are also used in areas such as immunomodulation, treatment of metabolic diseases, and neurological disorders. Due to their targeting and biocompatibility, peptide drugs have become one of the important tools for treating various diseases. However, peptide drugs face some challenges, such as poor biological stability, limitations in administration routes, and high preparation costs. Despite these difficulties, researchers are constantly striving to overcome them, improving the properties of peptide drugs through chemical synthesis, bioprocessing, and other methods, promoting their development in clinical applications, and providing new directions for innovative drug development.

[0004] Based on this, the present invention develops a MEIS1-derived peptide and its application in the preparation of drugs for treating colorectal cancer. Summary of the Invention

[0005] This invention provides a MEIS1-derived peptide and its application in the preparation of drugs for treating colorectal cancer, with the aim of solving the aforementioned problems existing in the background art.

[0006] To achieve the above objectives, embodiments of the present invention provide a MEIS1-derived peptide and its application in the preparation of drugs for treating colorectal cancer. In vitro (CCK8 and plate cloning experiments) and in vivo animal experiments have shown that the MEIS1-derived peptide can significantly inhibit the proliferation of colorectal cancer cells. The MEIS1-derived peptide provides a theoretical basis and experimental evidence for the preparation of drugs for treating colorectal cancer patients and the development of new and effective targeted therapies.

[0007] An embodiment of the present invention provides a MEIS1-derived peptide, the amino acid sequence of which is SEQ ID NO: 1.

[0008] Based on a general inventive concept, embodiments of the present invention provide the application of the above-described MEIS1 in the preparation of a medicament for treating colorectal cancer.

[0009] The embodiments of the present invention also provide the application of the above-mentioned MEIS1-derived peptide in the preparation of a medicament for treating colorectal cancer.

[0010] The above-described solution of the present invention has the following beneficial effects:

[0011] This invention demonstrates through in vitro experiments (CCK8 and plate cloning experiments) and in vivo animal experiments that MEIS1-derived peptides can significantly inhibit the proliferation of colorectal cancer cells, providing a theoretical basis and experimental evidence for the preparation of therapeutic drugs for colorectal cancer patients and the development of new and effective targeted drug therapies. Attached Figure Description

[0012] To more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the drawings used in the embodiments will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0013] Figure 1 This is a diagram showing the in vivo experimental results of the effect of the unmodified MEIS1 of the present invention on the growth of colorectal cancer;

[0014] Figure 2 This is a CCK8 assay result showing the effect of unmodified MEIS1 expression on MDM2-induced cell proliferation.

[0015] Figure 3This is a fluorescence colorimetric experiment result of the lipid solubility of MEIS1-derived peptides with different contents of the present invention.

[0016] Figure 4 This is a CCK8 experimental result diagram showing the effect of the MEIS1-derived peptide of the present invention on the proliferation ability of colorectal cancer cells;

[0017] Figure 5 This is a graph showing the results of a plate colony experiment on the effect of the MEIS1-derived peptide of the present invention on the colony-forming ability of colorectal cancer cells.

[0018] Figure 6 This is a diagram showing the comparative results of a subcutaneous tumorigenesis experiment in nude mice on the effect of the MEIS1-derived peptide of this invention on the growth of colorectal cancer tumors. Detailed Implementation

[0019] To make the technical problems, solutions, and advantages of this invention clearer, a detailed description will be provided below with reference to the accompanying drawings and specific embodiments. Unless otherwise defined, all technical terms used herein have the same meaning as commonly understood by those skilled in the art. The technical terms used herein are for the purpose of describing specific embodiments only and are not intended to limit the scope of protection of this invention. Unless otherwise specifically stated, all raw materials, reagents, instruments, and equipment used in this invention are commercially available or can be prepared by existing methods.

[0020] This invention addresses existing problems by providing a MEIS1-derived peptide, the amino acid sequence of which is SEQ ID NO: 1. The invention also relates to the application of MEIS1 and its derivative peptide in the preparation of medicaments for treating colorectal cancer.

[0021] This invention demonstrates through the following in vitro and in vivo experiments that MEIS1-derived peptides can significantly inhibit the proliferation of colorectal cancer cells, providing a theoretical basis and experimental evidence for the preparation of therapeutic drugs for colorectal cancer patients and the development of new and effective targeted drug therapies.

[0022] The application will be explained below through specific embodiments.

[0023] Example 1

[0024] This application's embodiments include in vivo experiments investigating the effect of MEIS1 on colorectal cancer growth.

[0025] The specific process of this application embodiment includes the following steps:

[0026] First, male nude mice that were 4 weeks old were selected as the experimental animals.

[0027] After digesting the NC group (blank control group) and HCT8 cells stably overexpressing MEIS1 in culture flasks into dispersed cells using trypsin, the cells were counted at a rate of 3 × 10⁻⁶. 6 The cells were collected by centrifugation and placed in 100 μL of RPMI 1640 cell culture medium.

[0028] The collected cells were subcutaneously injected into the axillae of mice to treat tumors.

[0029] After cell inoculation, tumor volume was measured every 2 days (calculated as: tumor length × width). 2 The tumor weight (× 0.5) and the survival status of nude mice were measured. The tumor was removed after day 28, and the results were as follows: Figure 1 As shown, the tumor growth rate in the MEIS1 overexpression group was significantly slower, and correspondingly, the tumor volume and weight were significantly smaller than those in the control group. In summary, the in vivo experimental results indicate that MEIS1 can inhibit the growth of colorectal cancer HCT8 cells.

[0030] Example 2

[0031] This application's embodiments investigate the effect of MEIS1 expression on MDM2-induced cell proliferation using a CCK8 assay.

[0032] The specific process of this application embodiment includes the following steps:

[0033] Human colorectal cancer adherent cells HCT8 and HCT116 (transfection groups: ①si NC; ② siMDM2; ③ si MDM2+ si MEIS1) transfected for 24 h, as well as human colorectal cancer adherent cells RKO and LOVO (transfection groups: ①oe NC; ②oe MDM2; ③ oe MDM2+oe MEIS1), were digested and seeded at a density of 5000 cells / well in 96-well plates (oe represents overexpression; si represents knockdown).

[0034] After 24 hours of cell adhesion, remove the culture medium and place the cells in an incubator.

[0035] After 24 hours of continuous culture, 10 μl of CCK8 solution was added to each well. After incubation in an incubator for 2 hours, the absorbance at 450 nm was measured using a microplate reader. The OD values ​​of each cell under different treatments relative to the control group were calculated based on the absorbance. The results are as follows: Figure 2 As shown, compared with the control group, the MDM2 overexpression group promoted the proliferation of RKO and LOVO cells, while the MDM2 overexpression and MEIS1 overexpression group inhibited the ability of MDM2 to promote cell proliferation. Similarly, MDM2 knockdown inhibited the proliferation of HCT8 and HCT116, while the MDM2 knockdown and MEIS1 knockdown group reversed this inhibitory effect.

[0036] Example 3

[0037] This application's embodiments include a fluorescence colorimetric experiment to investigate the lipid solubility of MEIS1-derived peptides.

[0038] Based on the phenomenon that overexpression of MEIS1 can inhibit MDM2-induced cell proliferation, this application designs and synthesizes MEIS1-derived peptides according to the amino acid sequences of MDM2 and MEIS1. The sequence SEQ ID NO: 1 is NFCHRYISCLK, and a FITC-labeled peptide is added to the N-terminus.

[0039] The specific process of this application embodiment includes the following steps:

[0040] First, different concentrations of MEIS1-derived peptides (0 μM, 10 μM, 20 μM, and 30 μM) were added to HCT8 colorectal cancer cells and incubated for 48 hours. The lipid solubility of the MEIS1-derived peptides was then detected under a fluorescence microscope. The fluorescence results are shown below. Figure 3 The results showed that MEIS1-derived peptides could enter HCT8 colorectal cancer cells and had good lipid solubility.

[0041] Example 4

[0042] This application's embodiments investigate the effect of MEIS1-derived peptides on the proliferation ability of colorectal cancer cells using a CCK8 assay.

[0043] The specific process of this application embodiment includes the following steps:

[0044] The following adherent cells (colorectal cancer HCT116 and HCT8 cells) were digested and seeded in 96-well plates at a density of 5000 cells / well.

[0045] After 24 hours of cell adhesion, the culture medium was removed and replaced with the control group and the MEIS1-derived peptide group, and then placed in an incubator.

[0046] After 24 hours of continuous culture, 10 μl of CCK8 solution was added to each well. After incubation in an incubator for 2 hours, the absorbance at 450 nm was measured using a microplate reader, and the OD value of cells under different treatments relative to the control group was calculated based on the absorbance. The results are as follows: Figure 4 As shown, compared with the control group, MEIS1-derived peptides significantly inhibited the proliferation of colorectal cancer HCT116 and HCT8 cells.

[0047] Example 5

[0048] This application's embodiments investigate the effect of MEIS1-derived peptides on the colony-forming ability of colorectal cancer cells using a plate colony-forming experiment.

[0049] The specific process of this application embodiment includes the following steps:

[0050] After digesting the following adherent cells (colorectal cancer HCT116 and HCT8 cells), seed them in 12-well plates at a density of 500 cells per well. After the cells adhere, add PBS buffer or 30 uM MEIS1-derived peptide to the corresponding well.

[0051] Each group was set up with 3 replicates and cultured at 37℃ for 1-2 weeks.

[0052] Cells grew to a visible cell cluster, the culture medium was discarded, and the cells were washed three times with PBS buffer. Then, 4% paraformaldehyde was added to fix the cells for 30 min. After fixation, the 24-well plate was inverted onto filter paper and dried for 10 min. Crystal violet was then added to the wells for staining for 20 min, followed by washing three times with PBS buffer and drying at room temperature. Results are as follows: Figure 5 The results showed that MEIS1-derived peptides could significantly inhibit the clonogenic ability of colorectal cancer HCT116 and HCT8 cells.

[0053] Example 6

[0054] This application includes a subcutaneous tumorigenesis experiment in nude mice to investigate the effect of MEIS1-derived peptides on colorectal cancer tumor growth.

[0055] The specific process of this application embodiment includes the following steps:

[0056] The experimental animals were 4-week-old male nude mice.

[0057] HCT8 cells cultured in culture dishes were digested with trypsin to disperse the cells, and then counted at 3 × 10⁻⁶ cells. 6 Cells were collected by centrifugation in 100 μL RPMI 1640.

[0058] The collected HCT8 cells were subcutaneously injected into the tumor in the axilla of mice.

[0059] Xenograft tumors were allowed to grow for 7 days. Then, mice were intraperitoneally injected with MEIS1-derived peptide at a dose of 30 mg / kg once daily, and tumor size was measured (tumor volume = 0.5 × length × width). 2 The tumor was removed 18 days after the first day of drug administration, and the results were as follows: Figure 6 As shown, consistent with in vitro experimental results, MEIS1-derived peptide treatment significantly reduced tumor volume compared to the control group, indicating that MEIS1-derived peptides can effectively inhibit tumor growth.

[0060] The above description represents the preferred embodiments of the present invention. It should be noted that those skilled in the art can make various improvements and modifications without departing from the principles of the present invention, and these improvements and modifications should also be considered within the scope of protection of the present invention.

Claims

1. A MEIS1-derived peptide, characterized in that, The amino acid sequence of the MEIS1-derived peptide is SEQ ID NO:

1.

2. The use of the MEIS1-derived peptide as described in claim 1 in the preparation of a medicament for treating colorectal cancer.

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