Application of indole-3-carbinol in preparation of thyroid cancer treatment medicine

Abstract

The invention belongs to the technical field of medicines, and particularly relates to application of indole-3-carbinol in preparation of thyroid cancer immunotherapy drugs. Through cell and animal experiments, the indole-3-methanol can be used for remarkably enhancing the curative effects of thyroid cancer immunotherapy and targeted therapy by inhibiting the activity of WWP1 ubiquitin ligase, stabilizing STAT6 protein expression, inhibiting TDO2-mediated tryptophan metabolism and remodeling an immune microenvironment. Indole-3-A is clear in action mechanism and high in safety, and has important clinical application value. In a mouse model, single use of indole-3-carbinol can significantly inhibit thyroid cancer growth and enhance CD8 + T cell infiltration and granzyme B activity, and the effect of indole-3-carbinol is equivalent to that of a PD-1 monoclonal antibody; the combination of indole-3-methanol and PD-1mAb can synergistically enhance the anti-tumor immune response and significantly inhibit tumor growth. The pharmaceutical composition is especially suitable for immunotherapy of drug-resistant or advanced refractory cases, and shows a remarkable synergistic effect compared with the existing therapy.

Description

Technical Field

[0001] The present invention belongs to the field of medical technology, and particularly relates to the application of indole-3-methanol in the preparation of thyroid cancer immunotherapy drugs. Background Art

[0002] Thyroid cancer is the most common malignant tumor of the endocrine system. Among them, differentiated thyroid cancer (such as papillary carcinoma) has a good prognosis, but advanced and undifferentiated thyroid cancer still faces severe challenges of lack of treatment options and low survival rate. Although targeted therapy (such as BRAF / MEK inhibitors) and immune checkpoint blockade therapy (such as PD-1 / PD-L1 inhibitors) bring hope to some patients, their objective response rate is less than 30%, and the problem of drug resistance is prominent. In addition, tryptophan metabolic reprogramming in the thyroid cancer microenvironment significantly weakens CD8 by producing the immunosuppressive metabolite kynic acid. + T cell anti-tumor activity exacerbates immune escape. Therefore, the development of new therapies targeting the metabolic-immune regulatory network, especially intervention strategies for immunotherapy resistance, has become a clinical challenge that needs to be solved urgently.

[0003] Indole-3-methanol (I3C) is a naturally occurring plant secondary metabolite widely found in cruciferous plants (such as broccoli and cabbage). Studies have shown that I3C and its active metabolites (such as 3,3'-diindolylmethane (DIM)) exhibit multi-target anti-tumor effects in various tumor models, including inducing apoptosis, inhibiting the cell cycle, regulating hormone levels, and suppressing tumor invasion and metastasis. Currently, I3C has demonstrated promising anti-tumor activity in a variety of solid tumors, including breast cancer and colorectal cancer. For example, in breast cancer, I3C can inhibit tumor cell proliferation by inhibiting estrogen receptor-related signaling pathways. Furthermore, recent studies have revealed that I3C may also modulate the tumor microenvironment and activate immune responses (e.g., enhancing the efficacy of PD-1 immune checkpoint inhibitors), further expanding its potential in immuno-oncology therapy. Although I3C holds broad application prospects in the anti-tumor field, its low bioavailability, poor stability, and complex in vivo metabolism continue to limit its clinical translation. Furthermore, its specific mechanism of action has not yet been fully elucidated, and currently lacks systematic development and application approaches. Therefore, developing I3C derivatives with modified structures or better delivery methods, or combination drug strategies to enhance its therapeutic efficacy and safety, has become a key technical issue in current research and application. Summary of the Invention

[0004] The purpose of the present invention is to provide an application of indole-3-methanol in the preparation of a thyroid cancer treatment drug and to provide a new thyroid cancer treatment drug with good efficacy.

[0005] Through cell and animal experiments, the present invention found that indole-3-carbinol significantly enhances the efficacy of thyroid cancer immunotherapy and targeted therapy by inhibiting WWP1 ubiquitin ligase activity, stabilizing STAT6 protein expression, inhibiting TDO2-mediated tryptophan metabolism, and reshaping the immune microenvironment. Indole-3-carbinol has a clear mechanism of action, high safety, and important clinical application value. Specifically:

[0006] Indole-3-carbinol alone significantly inhibited thyroid cancer growth and enhanced CD8 + The effects of indole-3-carbinol and PD-1 mAb were comparable to those of T cell infiltration and granzyme B activity. Combining indole-3-carbinol with PD-1 mAb synergistically enhanced the anti-tumor immune response and significantly inhibited tumor growth.

[0007] In vitro experiments have shown that indole-3-carbinol can promote T cells to kill thyroid cancer cells in vitro.

[0008] Accordingly:

[0009] The present invention provides application of indole-3-methanol in preparing medicine for treating thyroid cancer.

[0010] Wherein, the treatment of thyroid cancer includes immunotherapy and targeted therapy; accordingly, the drugs are immunotherapy drugs and targeted therapy drugs.

[0011] Furthermore, the drug, in addition to indole-3-carbinol, also includes PD-1 antibody, that is, indole-3-carbinol and PD-1 antibody are used in combination;

[0012] The present invention also provides a pharmaceutical composition for treating thyroid cancer; the pharmaceutical composition comprises indole-3-methanol, PD-1 antibody and a pharmaceutically acceptable carrier.

[0013] The pharmaceutical composition may be in the form of tablets, capsules, powders, granules or oral liquids.

[0014] Compared with the prior art, the present invention has the following advantages:

[0015] (1) Promote immunotherapy for thyroid cancer: In mice, oral administration of indole-3-methanol alone achieved the same therapeutic effect as PD-1 antibody treatment compared with the control group. Combining indole-3-methanol with PD-1 antibody significantly improved the effect of PD-1 antibody treatment. Figure 3 As shown in A, B, C, D, and E;

[0016] (2) Animal experiments have confirmed that the above therapeutic effects can be achieved through oral administration of indole-3-carbinol;

[0017] (3) The present invention demonstrates that the intervention strategy targeting the WWP1-STAT6-TDO2 axis has important application value in the preparation of therapeutic drugs for thyroid cancer, and is particularly suitable for immunotherapy-resistant or advanced refractory cases. Compared with existing therapies, it exhibits significant synergistic effects. BRIEF DESCRIPTION OF THE DRAWINGS

[0018] Figure 1 This is a diagram showing the effect of indole-3-carbinol upregulating STAT6 and downregulating TDO2 protein in thyroid cancer cells.

[0019] Figure 2 The figure shows the effect of indole-3-carbinol on promoting T cells to kill thyroid cancer cells in vitro. + T cells, B is crystal violet staining of surviving tumor cells, and C is the survival rate of tumor cells.

[0020] Figure 3 Indole-3-carbinol effectively inhibits thyroid cancer growth in mice; the combined use of indole-3-carbinol and PD-1 antibodies enhances the efficacy of PD-1 antibody immunotherapy. Figure A is a schematic diagram of a transplanted tumor, B is a mouse thyroid tumor, C is a mouse thyroid tumor growth curve, D is the mouse thyroid tumor weight, and E is the mouse thyroid tumor STAT6, TDO2, and GAPDH protein levels. DETAILED DESCRIPTION

[0021] The present invention is further described below by way of examples with reference to the accompanying drawings. Experimental methods in the following examples where specific conditions are not specified were performed according to conventional methods and conditions, or selected according to the product specifications.

[0022] The PD-1 antibody described in the examples is InVivoMAb anti-mouse PD-1 (CD279), a commonly used PD-1 antibody in the art. Those skilled in the art may also select other PD-1 antibodies for use in combination with indole-3-carbinol based on clinical or experimental needs.

[0023] Example 1, indole-3-carbinol upregulates STAT6 and downregulates TDO2 protein in thyroid cancer cells;

[0024] 100 μM indole-3-carbinol was added to the culture medium of human thyroid cancer TPC-1 cells, and the expression levels of STAT6 and TDO2 in the cells were detected by Western blot 48 hours later. Figure 1 shown.

[0025] Example 2: Indole-3-carbinol promotes T cells to kill thyroid cancer cells in vitro;

[0026] (a) In vitro culture and activation of CD8+ T cells. 10 ml of fresh blood was obtained from healthy adult volunteers, and human peripheral blood mononuclear cells (PBMCs) were isolated. PBMCs were cultured in ImmunoCult-XF T Cell Expansion Medium containing IL-2 (10 ng / mL). PBMCs were activated using ImmunoCult Human CD3 / CD28 / CD2 T Cell Activator (25 μg / ml) for the first 3 days of culture. Activated human CD8+ T cells were obtained after 7 days of culture. Figure 2 As shown in A, CD8+ T cells aggregate into clusters after activation.

[0027] (b) Indole-3-carbinol treatment of tumor cells. TPC-1 tumor cells were divided into two groups: the control group was cultured with standard DMEM medium, while the treated group was cultured with DMEM medium supplemented with 100 μM indole-3-carbinol. After 72 hours of culture, the cells in both groups were trypsinized, counted, and evenly plated into two wells of a 24-well plate.

[0028] (c)CD8 + Co-culture of T cells and tumor cells: The activated CD8+ T cells were added to a tumor cell culture plate at a ratio of 1:10 tumor cells to T cells, and co-cultured in DMEM / F12 medium containing CD3 antibody (100 ng / mL) and IL-2 (10 ng / mL).

[0029] (d) Detection of tumor cell survival rate. CD8 + After 48 hours of co-culture of T cells and tumor cells, the liquid in the culture plate was aspirated and cell debris was washed away with PBS. The surviving tumor cells in the cell culture plate were stained with crystal violet staining solution, then decolorized with 33% (m / v) acetic acid solution, and the absorbance of the decolorized solution was measured at 570 nm. The results are shown in Figure 2. Figure 2 As shown in Figure B, the combination of I3C and PD1 significantly enhanced the tumor killing effect of CD8+ T cells. Figure 2C is a statistical chart of the results.

[0030] Example 3: Indole-3-carbinol effectively inhibits the growth of thyroid cancer in mice; the combined use of indole-3-carbinol and PD-1 antibody promotes the effect of PD-1 antibody immunotherapy.

[0031] (a) Construction of mouse thyroid cancer model. 6 BPC mouse thyroid cancer cells were injected subcutaneously into female C57 mice, and solid thyroid tumors grew after about 6 days.

[0032] (b) Intraperitoneal injection of indole-3-carbinol: From day 6 to day 22 after tumor cell inoculation, mice were intraperitoneally injected with indole-3-carbinol at 20 mg / kg each time every three days.

[0033] (c) PD-1 antibody was administered by intraperitoneal injection. From day 6 to day 22 after tumor cell inoculation, mice were intraperitoneally injected with 200 μg of PD-1 antibody every three days.

[0034] (d) Tumor volume measurement. Starting from the inoculation of the tumor, every two days, the length and width of the subcutaneous tumor of the mouse were measured with a vernier caliper, and the volume was calculated. Figure 3 A is a schematic diagram of the transplanted tumor, B is a mouse thyroid tumor, C is the growth curve of the mouse thyroid tumor, D is the weight of the mouse thyroid tumor, and E is the expression of STAT6, TDO2, and GAPDH proteins in the mouse thyroid tumor.

Claims

1. The use of indole-3-carbinol in the preparation of drugs for treating thyroid cancer.

2. The use according to claim 1 is characterized in that the treatment of thyroid cancer includes immunotherapy and targeted therapy; accordingly, the drugs are immunotherapy drugs and targeted therapy drugs.

3. The use according to claim 2, characterized in that The drugs, in addition to indole-3-carbinol, also include PD-1 antibodies.

4. A pharmaceutical composition for treating thyroid cancer, characterized in that: The invention comprises indole-3-carbinol, PD-1 antibody and a pharmaceutically acceptable carrier.

5. The pharmaceutical composition according to claim 4, characterized in that The dosage form is tablet, capsule, powder, granule or oral liquid.

Images