Use of a small molecule compound in the preparation of a medicament for treating a tumor

Abstract

This invention discloses the application of a small molecule compound in the preparation of drugs for treating tumors, belonging to the field of biomedical technology. The small molecule compound described in this invention, 4-ethoxyphenol, is an endogenous metabolite and a natural product derived from star anise. Experiments have demonstrated that 4-ethoxyphenol can significantly upregulate CD8. + This invention reveals a novel application of 4-ethoxyphenol as an immunomodulator, providing a new candidate small molecule drug for tumor immunotherapy. It enhances the expression of anti-tumor effector molecules on T cells and allows for the direct inhibition of tumor cell activity.

Description

Technical Field

[0001] This invention belongs to the field of biomedicine, specifically relating to the novel application of a small molecule compound, 4-ethoxyphenol, in the preparation of tumor immunomodulatory drugs and drugs that directly inhibit tumor cells. Background Technology

[0002] Malignant tumors are a global health challenge, and their prevention and treatment have become one of the core tasks of public health systems. Traditional tumor treatments such as surgical resection, chemotherapy, and radiotherapy, while able to eliminate tumor cells to some extent, have limitations such as poor targeting, high recurrence rates, and significant damage to the body, making it difficult to achieve long-term effective tumor control. In recent years, anti-tumor immunotherapy has emerged, its core logic being to awaken the body's own immune system and eliminate tumor cells through specific recognition. However, existing immunotherapies still face problems such as low response rates and severe immune-related adverse reactions, necessitating the development of novel, highly effective, and low-toxicity immunomodulators.

[0003] As the core effector cells of the anti-tumor immune response, the integrity of T cells' activation, proliferation, and killing functions directly determines the effectiveness of anti-tumor immunity. Under physiological conditions, the body can promptly identify and eliminate mutated tumor cells through the immune surveillance function of the immune system, preventing the occurrence and development of tumors. However, when tumor cells escape the immune system, such as by downregulating MHC molecule expression, secreting immunosuppressive factors, or inducing immunosuppressive cell infiltration, T cell activation is inhibited and their function is exhausted, rendering them unable to effectively exert their anti-tumor effects, ultimately leading to tumor progression, metastasis, and recurrence.

[0004] Currently, the anti-tumor immunotherapies used in clinical practice are mainly based on immune checkpoint inhibitors, and also include some cytokine preparations. Their mechanisms of action mostly revolve around activating the anti-tumor function of T cells, but all of them have certain side effects. In addition, clinical trials are exploring novel immunotherapies such as CAR-T cell therapy and tumor vaccines. However, CAR-T cell therapy has side effects such as complex preparation, extremely high cost, off-target effects, and cytokine release syndrome, while tumor vaccines have limited efficacy and a narrow applicable population, and have not yet achieved widespread clinical application.

[0005] 4-Ethoxyphenol, also known as p-ethoxyphenol, is an aromatic organic compound with CAS number 622-62-8 and molecular formula C8H2O. 10O2, with a molecular weight of 138.16, is a beige crystalline powder or lumps at room temperature, possessing a mild, sweet vanilla odor. It is slightly soluble in water and readily soluble in organic solvents such as ethanol and ether. Its melting point is 64-67℃, boiling point is 131℃ (9 mmHg), pKa is approximately 10.44±0.13, and LogP is 1.87. This compound is a versatile synthetic intermediate used in the synthesis of complex molecules such as liquid crystals and pharmaceuticals. It also has applications in cleaning products, cosmetics, fragrances, and food and beverages. Its phenolic hydroxyl and ether bond structures provide sites for further chemical modification, giving it the potential to be developed into a bioactive molecule. Furthermore, 4-ethoxyphenol, as an endogenous metabolite, possesses certain biological activities. Studies have shown that it exhibits anti-larval activity, and its structure is similar to that of phenolic compounds with immunomodulatory and antitumor activities (such as paeonol and resveratrol), providing a theoretical basis for exploring its antitumor immune-related effects.

[0006] Compared to currently used immunotherapy drugs, 4-ethoxyphenol has a wide range of sources, a relatively simple synthesis process, and a low cost. Furthermore, no significant serious toxicity has been found in studies to date, indicating its promising application potential. Summary of the Invention

[0007] To address the shortcomings of existing technologies, this invention provides the application of the small molecule compound 4-ethoxyphenol in the preparation of drugs for treating tumors. Experiments have shown that 4-ethoxyphenol promotes the production of cytokines TNF-α and IFN-γ and the protein Granzyme B in vitro, and inhibits tumor cell growth and metabolic activity, thus showing promise as a novel immunomodulator.

[0008] The 4-ethoxyphenol described in this invention has the molecular formula C8H. 10 O2, with a molecular weight of 138.16, has the following structural formula:

[0009] To achieve the above objectives, the present invention provides the following technical solution: This invention provides a novel use of 4-ethoxyphenol in the preparation of medicaments for treating tumors.

[0010] Furthermore, the 4-ethoxyphenol comprises its pharmaceutically acceptable salts, enantiomers, diastereomers, tautomers, and other derivatives.

[0011] Furthermore, the 4-ethoxyphenol can be administered orally, by injection, or by infusion after in vitro activation of T cells.

[0012] Furthermore, the 4-ethoxyphenol enhances the anti-tumor immunity of T lymphocytes by upregulating the expression of anti-tumor effector molecules TNF-α, IFN-γ, and Granzyme B on T lymphocytes.

[0013] Furthermore, the 4-ethoxyphenol exerts its anti-tumor function by inhibiting tumor cell activity.

[0014] Furthermore, the drug uses 4-ethoxyphenol as its active ingredient and further comprises pharmaceutically acceptable excipients.

[0015] Compared with the prior art, the beneficial effects of the present invention are as follows: This invention demonstrates for the first time the dual-mechanism antitumor activity of 4-ethoxyphenol, breaking through the application boundaries of its food additives and daily chemical products. In vitro culture experiments confirm that this compound can significantly upregulate CD8. + The core effector molecules that enable T cells to exert their anti-tumor killing effects have been elucidated, revealing that they work by enhancing the body's CD8+. + The mechanism of action of T cell-mediated anti-tumor immune response in assisting tumor suppression. Building upon this, the present invention further confirms that 4-ethoxyphenol can significantly inhibit the growth of melanoma and lymphoma in vivo, and further verifies that it can directly inhibit the activity of colon cancer cells, indicating that it has the ability to directly target tumor cells for tumor suppression.

[0016] The advantages of this invention are that it not only clarifies the novel biological function of 4-ethoxyphenol, filling a research gap in its application in tumor treatment, but also demonstrates a broad-spectrum anti-tumor effect through validation in various tumor models such as melanoma, lymphoma, and colon cancer cells. Furthermore, this compound exhibits low toxicity, good biocompatibility, wide availability of raw materials, and low preparation cost. Compared to the strong cytotoxicity of traditional chemotherapy drugs and the scarcity, complex preparation, or safety concerns of existing novel immunomodulators, the active ingredient of this invention significantly reduces the safety risks and costs of clinical application while ensuring the enhancement of T-cell anti-tumor immune effects. Attached Figure Description

[0017] Figure 1 The 4-ethoxyphenol in Example 1 of this invention significantly promotes CD8 in vitro. + Upregulation of T-cell anti-tumor immune effector molecules; Figure 2 The 4-ethoxyphenol in Example 2 of this invention can significantly inhibit the growth of melanoma and lymphoma; Figure 3 The 4-ethoxyphenol in Example 3 of this invention can significantly inhibit the cell activity of MC38 tumor cells. Detailed Implementation

[0018] To make the objectives, technical solutions, and advantages of the present invention clearer, the present invention will be further described in detail below with reference to specific embodiments and accompanying drawings.

[0019] Unless otherwise specified, the experimental methods used in the following examples are conventional methods, performed in accordance with the techniques described in the literature or the product instructions. Unless otherwise specified, the materials and reagents used in the following examples are commercially available.

[0020] The following examples use GraphPad 8.0.2 statistical software to process the data. The experimental results are expressed as mean ± standard deviation. Paired samples t-tests were used for pairwise comparisons. P < 0.05 (*) indicates a significant difference, P < 0.01 (**) indicates a highly significant difference, P < 0.001 (***) indicates a highly significant difference, P < 0.0001 (****) indicates a highly significant difference, and NS indicates no significant difference.

[0021] In the examples below, the wild-type mouse strain involved was C57BL / 6, purchased from Beijing Vital River Laboratory Animal Technology Co., Ltd. All mice used in the experiments were 6-8 weeks old.

[0022] Example 1: 4-Ethoxyphenol can significantly promote CD8 in vitro. + Upregulation of T-cell anti-tumor effector molecules; 1. Original CD8 + In vitro culture of T cells: Mouse spleen and lymph nodes were extracted and infiltrated into MACS solution. The cells were then milled into a cell suspension through a 0.44 μm filter membrane and incubated at 4°C, 1800 rpm for 4 min. The supernatant was discarded, and the cells were resuspended in MACS solution. CD88 was then added. + Magnetic beads were thoroughly mixed and incubated at 4°C in the dark for 20 minutes. CD8 was then enriched using the immunomagnetic bead method. + T cells were then added to RPMI 1640 complete medium and treated with compounds: 4-ethoxyphenol in the intervention group and DMSO in the non-intervention group. Finally, the cells were seeded into cell culture plates coated with anti-CD3 and CD28 and cultured in a 37°C CO2 incubator for 2 days.

[0023] 2. Flow cytometry analysis: Lymphocytes were isolated using lymphocyte separation medium, first stained with the membrane-labeled antibody CD8, and then intracellularly stained, including TNF-α, IFN-γ and Granzyme B, and detected by flow cytometry.

[0024] 3. Experimental Results: Depend on Figure 1 As shown, in wild-type mice CD8 + In T cell differentiation experiments, treatment with 4-ethoxyphenol resulted in CD8... + The levels of key effector molecules of T cell anti-tumor immunity, TNF-α, IFN-γ and Granzyme B, were also significantly upregulated.

[0025] 4. Discussion of Results: In vitro, this invention involved treating wild-type mouse CD8 cells with 4-ethoxyphenol. + T cell differentiation experiments revealed that 4-ethoxyphenol can enhance CD8+. + The role of T cells in anti-tumor killing function. To verify the effect of 4-ethoxyphenol on tumor growth, the inhibitory effect of 4-ethoxyphenol on tumor growth will be investigated next.

[0026] Example 2: 4-Ethoxyphenol can significantly inhibit the growth of melanoma and lymphoma; 1. Tumor modeling process: 4-Ethoxyphenol was dissolved in DMSO and diluted with physiological saline. Starting 7 days before tumor implantation, mice were injected intraperitoneally daily with 30 mg / kg of 4-ethoxyphenol, while the control group was injected with DMSO. On Day 0, each mouse was injected with an equal number of tumor cells; Observe and measure the size and condition of the mouse tumors daily, and calculate the tumor volume; 2. Experimental Results: like Figure 2 As shown, tumor growth in mice treated with 4-ethoxyphenol was significantly inhibited compared to the control group.

[0027] 3. Discussion of Results: Based on the experimental results of Example 2, the present invention discovered that 4-ethoxyphenol can significantly inhibit tumor growth in vivo. The present invention further demonstrates the direct inhibitory effect of 4-ethoxyphenol on tumor cells.

[0028] Example 3: 4-Ethoxyphenol can significantly inhibit the metabolic activity of tumor cells; 1,4-Ethoxyphenol treatment of MC38 tumor cells: MC38 cells were cultured in DMEM complete medium containing different concentrations of 4-ethoxyphenol for 24 h. 2. CCK8: Cells were treated with DMEM complete medium containing 10% CCK8 for 1 hour, followed by measurement of OD. 450 value; 3. Experimental Results: Depend on Figure 3 As shown, the metabolic activity of MC38 tumor cells was significantly reduced in the 4-ethoxyphenol treatment group in a concentration-dependent manner.

[0029] 4. Discussion of Results: In this embodiment, the effects of different concentrations of 4-ethoxyphenol on the metabolic activity of tumor cells were investigated by measuring OD450 values ​​using the CCK8 assay after treatment for 24 hours. The results suggest that 4-ethoxyphenol can directly inhibit the activity of MC38 tumor cells, thereby exerting a direct antitumor effect. This result echoes the in vitro immunomodulation and in vivo tumor suppression experiments in Example 2, further confirming that 4-ethoxyphenol possesses a dual antitumor mechanism of action—immunomodulation and direct tumor suppression—providing direct experimental evidence for its application as a novel antitumor drug.

[0030] The specific embodiments described above further illustrate the inventive content, research objective, technical solution, and potential pharmacological effects of this invention. It should be noted that the above are merely preferred embodiments of this invention and are not intended to limit the scope of implementation of this invention. All technical routes falling within the research concept of this invention are within the protection scope of this invention. In particular, any modifications, deletions, and equivalent substitutions made without departing from the spirit and scope of this invention should be within the protection scope of this invention.

[0031] Any aspects not covered in this invention are applicable to existing technologies.

Claims

1. The application of a small molecule compound in the preparation of a drug for treating tumors, characterized in that, The drug contains an effective dose of 4-ethoxyphenol, or a pharmaceutically acceptable salt or solvate thereof as the main active ingredient, and further contains pharmaceutically acceptable excipients.

2. The application according to claim 1, characterized in that, Drugs include pharmaceutically acceptable salts, enantiomers, diastereomers, tautomers and other derivatives; The molecular formula of 4-ethoxyphenol is C8H. 10 O2, with a molecular weight of 138.16, has the following structural formula: 。 3. The application according to claim 1, characterized in that, The drugs can be administered orally, by injection, or by infusion after in vitro activation of T cells.

4. The application according to claim 1 or 2, characterized in that, 4-Ethoxyphenol upregulates the expression of antitumor effector molecules on T lymphocytes, thereby enhancing the antitumor immunity of T lymphocytes.

5. The application according to claim 1 or 2, characterized in that, 4-Ethoxyphenol can exert its anti-tumor function by inhibiting the activity of tumor cells.

6. The application according to any one of claims 1-5, characterized in that, Tumors include, but are not limited to, melanoma, lymphoma, or colon cancer.

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