Use of 4-hydroxyphenylacetic acid in combination with irinotecan as a cancer chemotherapeutic

Abstract

The application relates to the field of medicine, in particular to the application of 4-hydroxyphenylacetic acid and irinotecan as cancer chemotherapy drugs. In view of the technical defects that the existing irinotecan chemotherapy hepatoprotective drugs have single action and may interfere with the chemotherapy effect, the application provides the application of 4-hydroxyphenylacetic acid and irinotecan in the preparation of cancer chemotherapy drugs. The application has the following advantages: 1. clear hepatoprotective effect: 4HPAA can significantly alleviate the liver fatty degeneration induced by irinotecan; 2. significant synergistic anti-tumor effect: the tumor inhibition effect of the combination of 4HPAA and irinotecan is significantly better than that of the single-drug group; 3. high safety; and 4. great clinical transformation potential.

Description

Technical Field

[0001] This invention relates to the pharmaceutical field, and more particularly to the application of 4-hydroxyphenylacetic acid and irinotecan in combination as a chemotherapy drug for cancer. Background Technology

[0002] Irinotecan, a specific DNA topoisomerase I inhibitor, is a first-line chemotherapy drug for metastatic colorectal cancer, pancreatic cancer, and other solid tumors. However, the clinical application of irinotecan is limited by severe chemotherapy-related toxic side effects, among which chemotherapy-related hepatic steatosis is one of the most challenging problems. Clinical data show that the incidence of hepatic steatosis in patients receiving irinotecan chemotherapy reaches 20%-30%, and the incidence is even higher in patients with metabolic syndrome.

[0003] Existing technology clearly demonstrates that the active metabolite of irinotecan, SN-38, accumulates in the liver, triggering a cascade reaction of "mitochondrial dysfunction - reactive oxygen species (ROS) burst - inflammation activation - lipid metabolism imbalance," leading to lipid deposition in hepatocytes and impaired liver function. This pathological process not only forces clinicians to reduce chemotherapy dosages or discontinue treatment, directly weakening the anti-tumor efficacy, but also increases the risk of perioperative bleeding and liver failure in patients with liver metastases.

[0004] Currently used hepatoprotective drugs have two major drawbacks: First, their targets are limited to the liver, focusing mainly on direct antioxidant or lipid synthesis inhibition, making it difficult to block the multiple pathological processes induced by SN-38, resulting in limited hepatoprotective effects; second, some drugs may interfere with the liver's metabolism of chemotherapy drugs, posing a risk of "reducing toxicity but reducing efficacy," and failing to meet the core clinical needs for adjuvant chemotherapy drugs.

[0005] Existing intervention strategies do not fully utilize the gut-liver axis immune regulation pathway and lack an integrated solution that can both protect liver function and synergistically enhance anti-tumor immunity. There is an urgent clinical need for safe and effective irinotecan chemotherapy companion agents. Summary of the Invention

[0006] To address the technical shortcomings of existing hepatoprotective drugs like irinotecan in chemotherapy, which have limited efficacy and may interfere with the treatment's effectiveness, this invention provides the application of 4-hydroxyphenylacetic acid and irinotecan in combination as a cancer chemotherapy drug.

[0007] This invention is implemented as follows: This invention provides the application of 4-hydroxyphenylacetic acid and irinotecan in combination in the preparation of cancer chemotherapy drugs.

[0008] Furthermore, the cancer chemotherapy drugs include chemotherapy drugs for colon cancer.

[0009] Furthermore, the cancer chemotherapy drug is irinotecan, a chemotherapy-associated liver injury protective drug.

[0010] Furthermore, the cancer chemotherapy drug is a drug that enhances the anti-tumor efficacy of irinotecan.

[0011] Furthermore, the combination therapy is a simultaneous administration of 4-hydroxyphenylacetic acid (4HPAA) to the subject receiving irinotecan chemotherapy, and the administration continues until the end of chemotherapy.

[0012] The present invention has the following advantages: 1. Clear hepatoprotective effect: Based on preliminary experimental data, 4HPAA can significantly alleviate irinotecan-induced hepatic steatosis. In mice treated with irinotecan, the deposition of lipid droplets in liver tissue was significantly reduced after intervention with 4HPAA, providing direct evidence to avoid reducing or interrupting chemotherapy dosage.

[0013] 2. Significantly enhanced anti-tumor efficacy: Preliminary experiments have confirmed that the combination of 4HPAA and irinotecan has a significantly better anti-tumor effect than the single-drug group, and can enhance the IFN-γ secretion function of CD8⁺T cells, providing a new path for improving the efficacy of chemotherapy.

[0014] 3. High safety: 4HPAA is an endogenous metabolite. When used as a monotherapy, it does not cause toxic reactions such as weight loss. When used in combination with irinotecan, it does not increase additional toxic side effects. It avoids the risk of traditional hepatoprotective drugs interfering with chemotherapy and is well tolerated.

[0015] 4. Great potential for clinical translation: Based on the completed preliminary experimental data, the 4HPAA application regimen provided by this invention has a clear composition and definite effect, and can be directly used for subsequent preclinical research and formulation development, providing a safe and effective adjuvant treatment option for patients undergoing irinotecan chemotherapy. Attached Figure Description

[0016] The present invention will be further described below with reference to the accompanying drawings and embodiments.

[0017] Figure 1 Oil Red O staining of liver tissue at week 5 of the liver injury experiment; a: control group; b: irinotecan group; c: irinotecan + 4HPAA group.

[0018] Figure 2 Tumor growth curves (left) and tumor images (right) of different treatment groups in the CT26 subcutaneous colon cancer model.

[0019] Figure 3 These are the results of flow cytometry.

[0020] Control was the control group, IRI was the irinotecan group, and IRI+4HPAA was the irinotecan + 4HPAA combination group. Detailed Implementation

[0021] The technical solution of the present invention will now be clearly and completely described in conjunction with the accompanying drawings and specific embodiments. Unless otherwise specified in the embodiments, conditions are performed according to conventional conditions or conditions recommended by the manufacturer. Reagents or instruments used, unless otherwise specified, are all commercially available conventional products.

[0022] I. Source of Raw Materials 4HPAA (4-hydroxyphenylacetic acid): Sigma-Aldrich, H50004.

[0023] Irinotecan hydrochloride: Source leaf, S25911.

[0024] The C57BL / 6 and BALB / c mice used in the experiment were obtained from Xingyi (Fuzhou) Biotechnology Co., Ltd. Specific implementation steps

[0025] 1.4 Verification of the protective effect of HPAA against irinotecan-induced hepatic steatosis (1) Experimental model: C57BL / 6 mice (16g, male) were randomly divided into 3 groups (n=6-8): control group, irinotecan group (60mg / kg), and irinotecan + 4HPAA group.

[0026] (2) Administration method: Irinotecan was administered intraperitoneally every two days; 4HPAA was dissolved in the daily drinking water of mice at a concentration of 1.5 mg / mL and allowed to drink freely for 5 weeks.

[0027] (3) Detection indicators: During the experiment, the mice's body weight and food and water intake were monitored every two days. At the end of the experiment, liver tissue was dissected and lipid deposition was analyzed by Oil Red O staining (frozen sections) to determine the hepatoprotective effect of 4HPAA.

[0028] (4) Results: like Figure 1 As shown, significant lipid droplet deposition was observed in the liver tissue of the irinotecan group, while lipid droplet deposition was significantly reduced in the group treated with 4HPAA, confirming that 4HPAA has a mitigating effect on irinotecan-induced hepatic steatosis.

[0029] 2.4 Safety and antitumor efficacy verification of HPAA combined with irinotecan (1) Experimental model: BALB / c mice (16g, male) were subcutaneously injected with 1×10⁻⁶ dredges on the right back. 6 CT26 colon cancer cells were used to construct a tumor-bearing model until the tumor volume reached 50 mm. 3Subsequently, the patients were randomly divided into 4 groups (n=6-8): control group, irinotecan group (60mg / kg), 4HPAA monotherapy group (1.5mg / mL drinking water), and irinotecan + 4HPAA combination group.

[0030] (2) Administration method: Irinotecan was administered intraperitoneally every two days, and 4HPAA was continuously dissolved in drinking water and administered until the experimental endpoint (the average tumor volume in the control group mice was 1500 mm). 3 The size is approximately 2000mm. 3 ).

[0031] (3) Detection indicators: Measure tumor volume every 2 days (volume = length × width) ² / 2) and mouse body weight, and plot the tumor growth curve.

[0032] (4) Results: like Figure 2 As shown, the 4HPAA monotherapy group had no significant effect on tumor growth; the tumor-suppressing effect of the irinotecan + 4HPAA combination group was significantly better than that of the irinotecan monotherapy group; in terms of safety, the weight gain of mice in the irinotecan monotherapy group was limited, while the combination group with 4HPAA did not show further weight loss or additional toxicity deterioration, confirming the safety and tolerability of the combination.

[0033] 3.4 Preliminary analysis of the immune mechanism of HPAA enhancement (1) Experimental design: Based on the above tumor-bearing mouse model, a control group, an irinotecan group, and an irinotecan + 4HPAA combination group were selected for preliminary exploration of the immune mechanism (n=5 / group).

[0034] (2) Detection methods: At the end of the experiment, tumor tissue was collected and cut into 1mm pieces using sterile scissors. 3 The cells were sized and added to a digestion solution containing 0.2 mg / ml hyaluronidase, 1 mg / ml type IV collagenase, and 0.02 mg / ml DNase I. The cells were digested in a shaker at 37°C for one hour, gently inverting and mixing once every 10 minutes. After digestion, the cells were filtered through a 70 μm cell sieve to obtain a single-cell suspension. The suspension was centrifuged at 4°C and 400 × g for 5 minutes, the supernatant was discarded, and the cells were resuspended in PBS. Red blood cell lysis buffer was added and the cells were incubated at room temperature for 5 minutes. The cells were then centrifuged and washed twice to obtain a purified suspension of tumor-infiltrating immune cells.

[0035] (3) The cell staining procedure shall be strictly performed in accordance with the following steps: ① Staining of live and dead cells: Take 1×10 6One cell was added to the fluorescent dye in the live / dead cell staining kit and incubated at room temperature in the dark for 30 minutes, followed by washing once with PBS. ② Surface antigen staining: Add fluorescently labeled anti-mouse CD45, CD3, and CD8a monoclonal antibodies to the cell suspension, incubate at room temperature in the dark for 60 minutes, and wash twice with PBS; ③ Fixation and permeabilization: Resuspend cells in 250 μL of pre-cooled Fixation / permeabilization solution, incubate at 4°C for 30 minutes, centrifuge to remove supernatant, add 500 μL of 1×BD perm / wash buffer (diluted with dd water), and centrifuge at 2000×g for 5 minutes; ④ Intracellular antigen staining: Add fluorescently labeled anti-mouse IFN-γ monoclonal antibody to the cells after membrane perforation, incubate at room temperature in the dark for 60 minutes, wash twice with perforation buffer, and resuspend the cells with an appropriate amount of PBS; ⑤ Flow cytometry detection.

[0036] (4) Results: like Figure 3 As shown, compared with the control group and the irinotecan monotherapy group, the irinotecan + 4HPAA combination group had significantly higher levels of intratumoral CD8+. + The proportion of T cells did not change significantly, but the intracellular IFN-γ positivity level showed an increasing trend, confirming that 4HPAA may enhance CD8+. + T-cell effector functions enhance anti-tumor efficacy.

[0037] III. Results and Analysis This invention verifies the hepatoprotective effect and antitumor synergistic effect of 4-hydroxyphenylacetic acid (4HPAA) on irinotecan chemotherapy, and preliminarily elucidates its immune regulation mechanism, providing direct preliminary experimental evidence for subsequent clinical translation.

[0038] This invention proposes an intervention strategy based on "gut-hepatic axis immune regulation" targeting 4-hydroxyphenylacetic acid (4HPAA), an endogenous aromatic amino acid metabolite: 1. Specifically improves irinotecan-induced hepatic steatosis; 2.4 The safety profile of HPAA combined with irinotecan does not diminish its single-agent antitumor efficacy; 3. By utilizing the ability of 4HPAA to activate key immune effector cells such as CD8⁺T cells, the tumor immunosuppressive microenvironment can be improved, thereby enhancing the anti-tumor effect of irinotecan.

[0039] While specific embodiments of the present invention have been described above, those skilled in the art should understand that the specific embodiments described are merely illustrative and not intended to limit the scope of the invention. Equivalent modifications and variations made by those skilled in the art in accordance with the spirit of the invention should be covered within the scope of protection of the claims of the present invention.

Claims

Application of 1,4-hydroxyphenylacetic acid and irinotecan in the preparation of cancer chemotherapy drugs.

2. The application according to claim 1, characterized in that: The cancer chemotherapy drugs mentioned include those for colon cancer.

3. The application according to claim 1, characterized in that: The cancer chemotherapy drug mentioned is irinotecan, a chemotherapy-associated liver injury protective drug.

4. The application according to claim 1, characterized in that: The cancer chemotherapy drug mentioned is a drug that enhances the anti-tumor efficacy of irinotecan.

5. The application according to claim 1, characterized in that: The combined use refers to simultaneous administration of medication.

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