Application of human beta-defensin-1 combined alphaPD-1 antibody in preparation of immunomodulatory medicine for treating or preventing colon cancer

Abstract

The invention provides application of a human beta-defensin-1 combined alphaPD-1 antibody in preparation of immunomodulatory drugs for treating or preventing colon cancer, and relates to the technical field of biological medicines, the regulation effect of the human beta-defensin-1 combined alphaPD-1 antibody on immune cells is observed in in-vivo experiments, TNF-alpha + CD8 + T and IL-2 + CD8 + T can be reduced, INF-gamma + CD8 + T and alphaPD-1 + CD8 + T cells are obviously improved, and the immunomodulatory effect of the human beta-defensin-1 combined alphaPD-1 antibody on the colon cancer is improved. The invention provides a new thought and approach for the application of the human beta defensin-1 combined alphaPD-1 antibody in anti-tumor immunity, and particularly improves the immune tolerance generated by single alphaPD-1 antibody treatment and promotes the infiltration of CD8 + T into tumors.

Description

Technical Field

[0001] The present invention relates to the field of biomedicine technology, and specifically to the use of human β-defensin-1 combined with αPD-1 antibody in the preparation of immunomodulatory drugs for treating or preventing colon cancer. Background Art

[0002] Colon cancer is one of the most common digestive tract malignancies with the highest morbidity and mortality rates worldwide, posing a serious threat to human health. Colon cancer is often the result of a complex interaction between multiple factors, including the intestinal microbiome, mucosal barrier, environment, and immune system, making its prevention and treatment a significant challenge. Over the past few years, immune checkpoint inhibitors (ICPIs), initially approved for the clinical treatment of metastatic melanoma, have achieved tremendous success. These therapeutic agents have been subsequently used to treat many other cancer types, including colorectal cancer.

[0003] Immune checkpoint blockade, which blocks inhibitory checkpoints and restores cancer immune responses, has significantly improved outcomes for patients with several cancer types, such as melanoma, colorectal cancer, and triple-negative breast cancer. However, immunotherapy remains ineffective in the majority of patients. The response rate to immunotherapy in melanoma patients is 20-30%. In other tumors, such as breast cancer, colon cancer, and prostate cancer, the response rate is 13%-38%. Many patients also respond to treatment initially but develop immune tolerance later in the course of treatment. One of the main reasons for immune tolerance is the inability of CD8+ T cells to infiltrate the tumor. Therefore, addressing immunotherapy resistance and improving response rates have become urgent issues.

[0004] Therefore, there is an urgent need for an immunomodulatory drug to prevent colon cancer. Summary of the Invention

[0005] To solve the above technical problems, the present invention provides the use of human β-defensin-1 combined with αPD-1 antibody in the preparation of immunomodulatory drugs for treating or preventing colon cancer, as follows: The invention relates to the use of human β-defensin-1 combined with αPD-1 antibody in the preparation of an immunomodulatory drug for treating or preventing colon cancer. The drug comprises human β-defensin-1 (hBD-1) and αPD-1 antibody, and is used to alleviate the symptoms of colon cancer and to have an immunomodulatory effect on colon cancer. The drug is used to alleviate the symptoms of colon cancer and to have an immunomodulatory effect on colon cancer.

[0006] Moreover, the drug exerts its therapeutic effect by regulating CD8+ T cell subsets in the tumor microenvironment.

[0007] Furthermore, the regulation of CD8+ T cell subsets in the tumor microenvironment includes: Reduce the ratio of TNF-α+CD8+T cells and IL-2+CD8+T cells; Increase the ratio of INF-γ+CD8+T cells and αPD-1+CD8+T cells.

[0008] Furthermore, the human β-defensin-1 is administered by enema.

[0009] Furthermore, the αPD-1 antibody is administered by intramuscular injection.

[0010] Furthermore, the medicament is used for immunomodulatory treatment of colon cancer.

[0011] Moreover, the drug is used for immunomodulatory treatment of colon cancer in the following ways: improving immune tolerance produced by single αPD-1 antibody treatment and promoting CD8+ T cell infiltration into the tumor.

[0012] Furthermore, the drug is used for patients with colon cancer who do not respond to or develop resistance to single αPD-1 antibody treatment.

[0013] Moreover, the human β-defensin-1 is used in combination with the αPD-1 antibody, the human β-defensin-1 is administered by enema, and the αPD-1 antibody is administered by intramuscular injection to achieve its immunomodulation on colon cancer.

[0014] Compared with the prior art, the present invention has the following beneficial effects: 1. The present invention has developed a human β-defensin-1 combined with αPD-1 antibody that can alleviate the symptoms of colon cancer and has an immunomodulatory effect on colon cancer. In vivo experiments have observed its regulatory effect on immune cells. It can reduce TNF-α+CD8+T and IL-2+CD8+T, and significantly increase INF-γ+CD8+T and αPD-1+CD8+T cells. These four types of T cells are extremely important in tumor immunotherapy (TNF-α+CD8+T cells: T cells that secrete TNF-α, overactivation may aggravate inflammatory damage; IL-2+CD8 +T cells: T cells that secrete IL-2, which are associated with T cell exhaustion or ineffective proliferation; INF-γ+CD8+T cells: T cells that secrete interferon γ (INF-γ), which are the core effector cells of anti-tumor immunity, can directly kill tumor cells and activate other immune cells; αPD-1+CD8+T cells: T cells that express PD-1. Combined therapy may reactivate these originally suppressed T cells by blocking PD-1 signals). This provides new ideas and approaches for the application of human β-defensin-1 combined with αPD-1 antibodies in anti-tumor immunity.

[0015] 2. The present invention provides an application of human β-defensin-1 combined with αPD-1 antibody in the preparation of immunomodulatory drugs for the treatment or prevention of colon cancer. hBD-1 regulates innate immunity and the tumor microenvironment, while αPD-1 antibody relieves T cell inhibition, and the two synergistically activate a stronger anti-tumor immune response.

[0016] 3. The present invention provides an application of human β-defensin-1 combined with αPD-1 antibody in the preparation of immunomodulatory drugs for the treatment or prevention of colon cancer. The combined treatment of human β-defensin-1 and αPD-1 antibody significantly increases tumor-infiltrating CD8+T cells, especially INF-γ+ cells, converting immune "cold tumors" into "hot tumors" sensitive to immunotherapy, thereby alleviating the symptoms of colon cancer and exerting an immunomodulatory effect on colon cancer.

[0017] 4. The use of human β-defensin-1 combined with αPD-1 antibody provided by the present invention in the preparation of immunomodulatory drugs for treating or preventing colon cancer can regulate T cell subsets, specifically: (1) Inhibit harmful subpopulations: Reduce TNF-α+CD8+T and reduce inflammatory damage; Reduce IL-2+CD8+T to avoid ineffective exhaustion; (2) Activation of key effector cells: Enhance INF-γ+CD8+T cells: core effector cells that directly kill tumor cells; Enhance αPD-1+CD8+T cells: restore the function of PD-1 positive exhausted T cells.

[0018] 5. In the treatment of colon cancer with colon cancer drugs, a single αPD-1 antibody has a poor response to colon cancer. The present invention provides an application of human β-defensin-1 combined with αPD-1 antibody in the preparation of immunomodulatory drugs for the treatment or prevention of colon cancer. The combination of human β-defensin-1 and αPD-1 antibody can improve insufficient T cell infiltration (cold tumor problem) and reverse the immunosuppressive microenvironment (reduce regulatory T cells (Treg), myeloid-derived suppressor cells (MDSC) and other suppressive cells).

[0019] 6. The present invention uses a mouse colon cancer model to demonstrate that the human β-defensin-1 combined with the αPD-1 antibody provided by the present invention can significantly inhibit tumor growth, showing a synergistic effect. Compared with the use of human β-defensin-1 or the use of αPD-1 antibody alone, the human β-defensin-1 combined with the αPD-1 antibody of the present invention has an enhanced anti-colon cancer tumor effect, showing great potential in the field of tumor treatment. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] Figure 1 Treatment with human β-defensin-1 inhibits colon cancer progression in CAC mice. Figure 1 A: Flowchart for constructing mouse models in each group; Figure 1 B: Colon length and tumor number growth of mice in each group; Figure 1 C: Body weight of mice in each group; Figure 1 D: The number of tumors in each group of mice; Figure 1 E: Colon length of mice in each group; Figure 1 F: HE staining results of colon tissue; Figure 1 G: Ki67 immunohistochemistry results of colon tissue.

[0021] Figure 2 These are the in vivo experimental results of human β-defensin-1 combined with αPD-1 antibody in mice with colon cancer; Figure 2 A shows the changes of TNF-α+CD8+T lymphocytes; Figure 2 B shows the changes of INF-γ+CD8+T lymphocytes; Figure 2 C is the change of IL-2+CD8+T lymphocytes; Figure 2 D shows the changes in PD-1+CD8+T lymphocytes. DETAILED DESCRIPTION

[0022] Example 1 In vivo experiment in mice The AOM / DSS method (azoxymethane combined with dextran sulfate sodium) was used to induce an inflammation-associated colon cancer model in mice (CAC mouse colon cancer model). Sixty female C57BL / 6j mice (6 weeks old) were selected and maintained at 22±2°C and 30-70% relative humidity with alternating light and dark for 12 hours. The mice were allowed to acclimate to a free diet for one week and then randomly assigned to five groups (n=12 / group): 1. Control group: No AOM / DSS treatment or any other treatment was given.

[0023] 2. Model group: AOM / DSS processing only.

[0024] 3. αPD-1 treatment group: AOM / DSS treatment; Treatment: Intramuscular injection of αPD-1 antibody (100 μg / 200 μL / kg, twice a week).

[0025] 4. Human β-defensin-1 treatment group: AOM / DSS treatment; Treatment: Intraperitoneal injection of hBD-1 (0.2 mg / kg / time, 2 times / week) and enema of hBD-1 (0.2 mg / kg / time, 2 times / week) were performed simultaneously.

[0026] 5. Human β-defensin-1 + αPD-1 antibody combined treatment group: AOM / DSS treatment; Treatment: enema hBD-1 (0.2 mg / kg / time, 2 times / week), intramuscular injection of αPD-1 antibody (100 μg / 200 μL / / kg time, 2 times / week).

[0027] AOM / DSS treatment: 10 mg / kg AOM was injected intraperitoneally on the first day, and then 2.0% DSS was added to the drinking water every day for 1 week. Normal drinking water was restored for the next 2 weeks, and this cycle was repeated three times.

[0028] Experimental results: Human β-defensin-1 treatment inhibits colon cancer progression in CAC mice ( Figure 1 On the first day, 10 mg / kg AOM was injected intraperitoneally. After one week, 2.0% DSS was added to the drinking water every day for one week. Normal drinking water was restored for the next two weeks. This cycle was repeated three times, and the body weight of the mice was monitored weekly. In the eighth week of modeling, which was the last cycle of 2.0% DSS, the CAC mice in the treatment group received treatment with αPD-1, human β-defensin-1, and human β-defensin-1 + αPD-1 antibody until the end of the tenth week ( Figure 1 A). The weight (1C), colon length (1B) and number of tumors (1D) of mice in each group were counted. The three treatment groups of human β-defensin-1, αPD-1 and human β-defensin-1 + αPD-1 all slowed down the shortening of the colon (1B) and weight loss (1C) of CAC mice ( p < 0.05), and the number of colon tumors was significantly reduced compared with the Model group (1D) ( p <0.05). Among the three treatment groups, the αPD-1 treatment group, the human β-defensin-1 treatment group, and the human β-defensin-1 + αPD-1 antibody combination treatment group had the best treatment effect. The HE staining results of colon tissue showed ( Figure 1 F) The colon glandular epithelium of the model showed obvious abnormal proliferation. Some abnormal crypts were found in the αPD-1 treatment group and the human β-defensin-1 treatment group, while only mild inflammatory infiltration was found in the β-defensin-1 + αPD-1 antibody combined treatment group. The results of Ki67 immunohistochemistry of colon tissue showed ( Figure 1G) The expression of Ki67 in the colon of the Model was significantly higher than that of the Control, while the expression of Ki67 in the αPD-1 treatment group, the human β-defensin-1 treatment group, and the human β-defensin-1 + αPD-1 antibody combination treatment group was significantly decreased compared with the Model. Among them, there was no statistical difference between the αPD-1 treatment group and the human β-defensin-1 treatment group. The remission rate in the human β-defensin-1 + αPD-1 antibody combination treatment group was better than that of the αPD-1 treatment group or the human β-defensin-1 treatment group alone. In summary, human β-defensin-1 treatment inhibited the progression of colon cancer in CAC mice, and the effect was better when combined with αPD-1.

[0029] The changes of TNF-α+CD8+T lymphocytes are as follows Figure 2 As shown in Figure A. Specifically, the frequencies of TNF-α+CD8+T lymphocytes changed significantly in the human β-defensin-1 treatment group and the human β-defensin-1+αPD-1 combined treatment group (P<0.05).

[0030] Figure 2 Figure B shows the changes in INF-γ+CD8+T lymphocytes. The data showed that the frequency of INF-γ+CD8+T lymphocytes was significantly higher in the human β-defensin-1+αPD-1 combined treatment group than in the other groups (P<0.05).

[0031] Figure 2 C depicts the changing trend of IL-2+CD8+T lymphocytes. The results showed that the frequency of IL-2+CD8+T lymphocytes was the lowest in the human β-defensin-1+αPD-1 combined treatment group (P<0.05).

[0032] Figure 2 D shows the changes in αPD-1+CD8+ T lymphocytes. The data showed that the frequency of αPD-1+CD8+ T lymphocytes was the highest in the human β-defensin-1+αPD-1 combination treatment group (P<0.05).

[0033] In summary, human β-defensin-1 combined with αPD-1 antibody treatment can significantly regulate the number of different CD8+T lymphocyte subsets in colon cancer mice, thereby synergistically enhancing the anti-tumor ability of the immune system through multiple mechanisms.

Claims

1. Use of human β-defensin-1 combined with αPD-1 antibody in the preparation of immunomodulatory drugs for treating or preventing colon cancer, characterized in that: The drug includes human β-defensin-1 and αPD-1 antibody, is used to alleviate the symptoms of colon cancer, and has an effect on the immune regulation of colon cancer.

2. The use of human β-defensin-1 combined with αPD-1 antibody as claimed in claim 1 in the preparation of immunomodulatory drugs for treating or preventing colon cancer, characterized in that: The drug exerts its therapeutic effect by regulating CD8+ T cell subsets in the tumor microenvironment.

3. The use of human β-defensin-1 combined with αPD-1 antibody in the preparation of an immunomodulatory drug for treating or preventing colon cancer according to claim 2, characterized in that: The regulation of CD8+T cell subsets in the tumor microenvironment includes: Reduce the ratio of TNF-α+CD8+T cells and IL-2+CD8+T cells; Increase the ratio of INF-γ+CD8+T cells and αPD-1+CD8+T cells.

4. The use of human β-defensin-1 combined with αPD-1 antibody in the preparation of an immunomodulatory drug for treating or preventing colon cancer according to claim 1, characterized in that: The human β-defensin-1 is administered by enema.

5. The use of human β-defensin-1 combined with αPD-1 antibody in the preparation of an immunomodulatory drug for treating or preventing colon cancer according to claim 1, characterized in that: The αPD-1 antibody is administered by intramuscular injection.

6. The use of human β-defensin-1 combined with αPD-1 antibody in the preparation of an immunomodulatory drug for treating or preventing colon cancer according to claim 1, characterized in that: The medicine is used for immunomodulatory treatment of colon cancer.

7. The use of human β-defensin-1 combined with αPD-1 antibody in the preparation of an immunomodulatory drug for treating or preventing colon cancer according to claim 6, characterized in that: The drug is used for immunomodulatory treatment of colon cancer by improving immune tolerance produced by single αPD-1 antibody treatment and promoting CD8+T cell infiltration into the tumor.

8. The use of human β-defensin-1 combined with αPD-1 antibody in the preparation of an immunomodulatory drug for treating or preventing colon cancer according to claim 1, characterized in that: The drug is used to treat colon cancer patients who do not respond to or develop resistance to single αPD-1 antibody treatment.

9. The use of human β-defensin-1 combined with αPD-1 antibody in the preparation of an immunomodulatory drug for treating or preventing colon cancer according to claim 1, characterized in that: The human β-defensin-1 is used in combination with the αPD-1 antibody, wherein the human β-defensin-1 is administered by enema and the αPD-1 antibody is administered by intramuscular injection to achieve its immunomodulation on colon cancer.

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