Application of flurbiprofen in preparation of medicine for enhancing colorectal cancer treatment effect of PD-L1 monoclonal antibody
By combining flurbiprofen with PD-L1 monoclonal antibody in the treatment of colorectal cancer, the problems of low response rate and drug resistance of PD-L1 monoclonal antibody in the treatment of colorectal cancer have been solved. This has achieved a safe and effective enhancement of the efficacy of PD-L1 monoclonal antibody, inhibiting the growth of colorectal cancer cells and enhancing the effect of immunotherapy.
Patent Information
- Application Number
- CN202511163772.1
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2025-08-20
- Publication Date
- 2025-11-11
AI Technical Summary
Current PD-L1 monoclonal antibodies have low response rates and exhibit drug resistance in the treatment of colorectal cancer, resulting in limited treatment efficacy. A safe, effective, and cost-efficient combination therapy strategy is needed to enhance the therapeutic effect.
Flurbiprofen was used in combination with PD-L1 monoclonal antibody at a concentration of 25-100 nM in vitro or 10-20 mg/kg in vivo. The ratio of flurbiprofen to PD-L1 monoclonal antibody in the combination was 10 mg/kg body weight: 50 μg/animal. This combination inhibited the growth of colorectal cancer cells and enhanced the efficacy of PD-L1 monoclonal antibody.
Flurbiprofen significantly inhibits the proliferation and migration of colorectal cancer cells, enhances the therapeutic effect of PD-L1 monoclonal antibodies, significantly slows tumor growth and improves the efficacy of immunotherapy, without increasing adverse reactions or treatment costs.
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Figure CN120919099A_ABST
Abstract
Description
Technical Field
[0001] This invention relates to the field of pharmaceutical technology, specifically to the use of flurbiprofen in the preparation of drugs for enhancing the efficacy of PD-L1 monoclonal antibody therapy for colorectal cancer. Background Technology
[0002] Colorectal cancer is one of the most common malignant tumors worldwide, with high incidence and mortality rates. Despite advancements in traditional treatments such as surgery, chemotherapy, and radiotherapy, the prognosis for patients with colorectal cancer remains unsatisfactory, especially for those with advanced or metastatic colorectal cancer. In recent years, the rise of immunotherapy has brought new hope to the treatment of colorectal cancer. PD-L1 monoclonal antibodies, as an important immune checkpoint inhibitor, have shown some therapeutic efficacy in colorectal cancer patients.
[0003] PD-L1 monoclonal antibodies work by blocking the binding of PD-L1 on the surface of tumor cells to PD-1 receptors on the surface of immune cells, thereby relieving the inhibitory effect of tumor cells on immune cells and activating the immune system to attack tumor cells. However, the application of PD-L1 monoclonal antibodies in the treatment of colorectal cancer still faces some challenges. First, the response rate of PD-L1 monoclonal antibodies in colorectal cancer patients is low, which is related to the complexity of the tumor microenvironment in colorectal cancer. Second, even in some patients, drug resistance exists, leading to limited treatment efficacy. Therefore, finding combination therapy strategies that can enhance the efficacy of PD-L1 monoclonal antibodies has become one of the current research hotspots.
[0004] Flurbiprofen is a nonsteroidal anti-inflammatory drug (NSAID) with anti-inflammatory and analgesic effects. In recent years, studies have revealed that flurbiprofen possesses potential anti-cancer properties in addition to its anti-inflammatory and analgesic effects, showing particular promise in the treatment of colorectal cancer. Flurbiprofen can influence the biological behaviors of cancer cells, such as proliferation and apoptosis, by regulating specific signaling pathways or targets, such as Wnt / β-catenin and NF-κB. Furthermore, flurbiprofen may synergize with PD-L1 monoclonal antibodies by modulating the tumor microenvironment and enhancing the activity of immune cells, thereby improving therapeutic efficacy. For example, flurbiprofen may reduce the inhibitory effect of tumor cells on the immune system by inhibiting the expression of certain immunosuppressive factors, or enhance the attack ability of PD-L1 monoclonal antibodies on tumor cells by promoting the infiltration and activation of immune cells.
[0005] In the exploration of immunotherapy for colorectal cancer, combination therapy strategies have gradually become a research focus. For example, some studies have attempted to combine PD-L1 monoclonal antibodies with other drugs (such as anti-angiogenic drugs, chemotherapy drugs, and targeted drugs) to overcome immunotherapy resistance and improve efficacy. These combination therapy regimens have improved patient prognosis to some extent, but some problems still exist, such as increased adverse reactions and higher treatment costs. Therefore, finding a safe, effective, and economical combination therapy regimen is of significant clinical importance. Summary of the Invention
[0006] To address the aforementioned technical problems, this invention provides a novel application of flurbiprofen. This invention discovers that flurbiprofen can inhibit the growth of colorectal cancer cells and enhance the efficacy of PD-L1 monoclonal antibodies in colorectal cancer. These results provide a potential therapeutic strategy for immunotherapy of colorectal cancer.
[0007] Specifically:
[0008] The use of flurbiprofen in the preparation of a drug for enhancing the efficacy of PD-L1 monoclonal antibody therapy for colorectal cancer. The dosage concentration of flurbiprofen is: 25-100 nM in vitro or 10-20 mg / kg in vivo.
[0009] The present invention also provides a pharmaceutical composition for treating colorectal cancer, comprising flurbiprofen, PD-L1 monoclonal antibody and a pharmaceutically acceptable carrier, wherein the ratio of flurbiprofen to PD-L1 monoclonal antibody is: flurbiprofen 10 mg / kg body weight and PD-L1 monoclonal antibody 50 μg / animal.
[0010] Pharmacological experiments have demonstrated that flurbiprofen can effectively inhibit the growth of colorectal cancer, significantly suppress the proliferation and migration of colorectal cancer cell lines, and enhance the efficacy of PD-L1 monoclonal antibodies, showing good potential as an adjuvant therapy for colorectal cancer. Currently, there are no research reports or patent documents regarding the use of flurbiprofen as an adjuvant therapy for PD-L1 monoclonal antibody immunotherapy in colorectal cancer. Attached Figure Description
[0011] Figure 1 This invention presents experimental results on the inhibition of colorectal cancer cell line proliferation by flurbiprofen.
[0012] Figure 2 This invention presents experimental results regarding the ability of flurbiprofen to inhibit the in vitro invasion of colorectal cancer cells SW480.
[0013] Figure 3 This invention presents experimental results regarding the ability of flurbiprofen to inhibit the in vitro invasion of Caco2 colorectal cancer cells.
[0014] Figure 4This invention presents experimental results showing that flurbiprofen inhibits tumor growth in C57 mouse MC38 cells.
[0015] Figure 5 This is the experimental result of the present invention showing that flurbiprofen enhances the inhibitory effect of PD-L1 monoclonal antibody on tumor-bearing MC38 cells of C57 mice. Detailed Implementation
[0016] To make the objectives, technical solutions, and advantages of the embodiments of the present invention clearer, the technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. The components of the embodiments of the present invention described and shown in the accompanying drawings can generally be arranged and designed in various different configurations.
[0017] Example:
[0018] This embodiment provides evidence that flurbiprofen inhibits colorectal cancer growth and enhances the efficacy of PD-L1 monoclonal antibodies. The specific method is as follows:
[0019] I. Experimental Materials and Methods
[0020] 1. Cell culture and treatment
[0021] Human colon cancer cell lines SW480 and Caco2 were purchased from the Cell Bank of the Chinese Academy of Sciences Type Culture Collection Committee. SW480 and Caco2 cells were cultured in DMEM high-glucose medium (gibco) containing 10% fetal bovine serum and 1% penicillin-streptomycin, respectively. Cells were cultured in a humidified incubator at 37°C and 5% CO2.
[0022] SW480 and CACO2 cells were treated with different concentrations (25 nM, 50 nM, and 100 nM) of flurbiprofen (purchased from GLPBIO). Cells were then collected for subsequent experiments.
[0023] 2. CCK-8 cell viability assay
[0024] SW480 and CACO2 cell viability was detected using CCK-8 assay.
[0025] Take a 96-well plate and add 100 μL to each well to adjust the concentration to 1 × 10⁻⁶. 5 Cell suspensions of 10 cells / mL were prepared, with 20 replicates for each intervention condition (25 nM, 50 nM, and 100 nM). Cells were gently agitated to ensure even distribution, and the 96-well plates were placed in a CO2 incubator at 37°C with 5% CO2. After cell attachment, the medium was replaced with 10% CCK8 and incubated for 1–1.5 h. The absorbance was then measured at 450 nm using a microplate reader (PerkinElmer, USA).
[0026] 3. Scratches on the flat surface
[0027] Take a 6-well plate, mark the bottom with a streak, add 2 mL of cell suspension to each well, and set up 3 replicates for each treatment group.
[0028] When the cell density in the 6-well plate reaches more than 90%, a 200 μL sterile pipette tip is used to streak the cells vertically along the marked line. The cells are then washed three times with PBS to remove any floating cells. Serum-free medium containing different concentrations (25 nM, 50 nM, and 100 nM) of flurbiprofen is added to observe and record the changes in cell scratches at 0 h and 48 h. The changes are photographed, recorded, and analyzed.
[0029] 4. Animal model establishment and treatment
[0030] Mouse colon cancer cell line MC38 cell suspension was cultured according to protocol 1.
[0031] Wash with an appropriate amount of PBS to remove residual culture medium. Gently pipette the resulting clean cell pellet with PBS, count the cells, and then dilute to an appropriate concentration to ensure a cell concentration of 5 × 10⁻⁶ cells / mL. 6 Cells / mL. Each mouse was inoculated with 100 μL of cell suspension, i.e., 5 × 10⁶ cells / mL. 5 Cells. Mice were prepared by shaving the left axilla one day prior to inoculation. The injection site was disinfected with an alcohol swab. After inserting the needle approximately 1 cm, 0.1 mL of cell suspension was slowly injected, followed by slow withdrawal to prevent leakage. Tumor formation was observed 3 days after injection. Once tumor formation was successful, drug intervention was initiated via daily gavage. Tumor volume changes were recorded throughout the experiment. Tumor volume was calculated as: V = (longest diameter of tumor × shortest diameter of tumor²) / 2. Intervention was discontinued after 2 weeks of drug administration, or when the tumor size approached 2 cm. Mice were euthanized, tumor tissue was collected, weighed, recorded, and photographed for documentation.
[0032] First, the effective concentration of flurbiprofen in mice needed to be determined. Mice with successfully developed tumors were randomly divided into three groups (n=5): non-nuclear (NC), low-concentration flurbiprofen (10 mg / kg), and high-concentration flurbiprofen (20 mg / kg). After two weeks of continuous administration, mice were sacrificed under isoflurane anesthesia, and subcutaneous tumors were collected, photographed, and weighed. After determining the effective concentration of flurbiprofen, the efficacy of combining flurbiprofen with PD-L1 monoclonal antibody was observed. At this point, mice with successfully developed tumors were randomly divided into four groups (n=6): NC, flurbiprofen (10 mg / kg), PD-L1 monoclonal antibody (50 μg / mouse, STARTER), and PD-L1 monoclonal antibody combined with flurbiprofen. After two weeks of continuous administration, mice were sacrificed under isoflurane anesthesia, and subcutaneous tumors were collected, photographed, and weighed.
[0033] 5. Statistical Analysis
[0034] All experimental data were expressed as mean ± standard deviation and analyzed using one-way ANOVA with GraphpadPrism 10.0 software. A p-value < 0.05 was considered statistically significant.
[0035] II. Experimental Results
[0036] 1. Flurbiprofen inhibits the proliferation of colorectal cancer cell lines.
[0037] The experimental design included three concentration gradients (25 nM, 50 nM, 100 nM) and three time gradients (24 h, 48 h, and 72 h).
[0038] The experimental results showed that, compared with the control group, the proliferation capacity of SW480 cells treated with flurbiprofen was significantly reduced at 24h (100 nM), 48h (50 nM), 100 nM, and 72h. Figure 1 A).
[0039] The proliferation capacity of Caco2 cells was significantly reduced at three time-gradient treatments with a concentration of 100 nM. Figure 1 B). Both cell lines showed the strongest inhibitory effect on colon cancer cells at a flurbiprofen concentration of 100 nM. The differences were statistically significant (P < 0.05).
[0040] 2. Flurbiprofen's ability to inhibit the in vitro invasion of colorectal cancer cells.
[0041] Two types of human colon cancer cells, SW480 and Caco2, were selected for migration and scratch assays.
[0042] Three concentrations of flurbiprofen were used in the treatment group (25 nM, 50 nM, and 100 nM). SW480 migration assays showed a significant decrease in cell migration in the flurbiprofen-treated group. Figure 2 The scratch test results for groups A and B showed that the scratch healing rate in the flurbiprofen-treated group was significantly lower than that in the control group. Figure 2 C, D). Results of Caco2 cell migration experiments ( Figure 3 A, B) and scratch test results ( Figure 3 C and D) were consistent with SW480. All results were statistically significant (P < 0.05).
[0043] 3. Flurbiprofen inhibits tumor growth in C57 mouse MC38 cells.
[0044] The experiment was divided into three groups: a control group, and treatment groups with flurbiprofen (10 mg / kg and 20 mg / kg). After successful tumor bearing, the mice were administered the medication via gavage. Tumor volume changes in the tumor-bearing mice were recorded every other day during the administration period, and the results were plotted as a line graph. Figure 4 B), the results showed that compared with the control group, the tumor growth rate of mice treated with flurbiprofen was significantly slower, and there was no significant difference between the two treatment groups with different concentrations. Figure 4 C).
[0045] 4. Flurbiprofen enhances the inhibitory effect of PD-L1 monoclonal antibody on tumor-bearing MC38 cells in C57 mice.
[0046] The experiment was divided into three groups: a control group, a flurbiprofen group, a PD-L1 monoclonal antibody group, and a PD-L1 monoclonal antibody combined with flurbiprofen group. After successful tumor implantation, the drugs were administered via gavage. Tumor volume and weight were measured at the end of the treatment period. Figure 5 The results showed that the tumor volume and weight in the flurbiprofen treatment group were smaller than those in the control group. Simultaneously, the tumor volume and weight in the PD-L1 monoclonal antibody combined with flurbiprofen group were smaller than those in the PD-L1 monoclonal antibody group, indicating that flurbiprofen has an adjuvant effect in PD-L1 monoclonal antibody treatment of colorectal cancer. All results were statistically significant (P < 0.05).
[0047] The present invention and its embodiments have been described above. This description is not restrictive, and the accompanying drawings are only one embodiment of the present invention; the actual structure is not limited thereto. In conclusion, if those skilled in the art are inspired by this description and design similar structures and embodiments without departing from the spirit of the invention, such designs should fall within the protection scope of the present invention.
Claims
1. Application of flurbiprofen in the preparation of drugs for enhancing the efficacy of PD-L1 monoclonal antibody therapy for colorectal cancer.
2. The application according to claim 1, characterized in that: The recommended concentration of flurbiprofen is 25-100 nM in vitro or 10-20 mg / kg in vivo.
3. A pharmaceutical composition for treating colorectal cancer, characterized in that, It contains flurbiprofen, PD-L1 monoclonal antibodies, and pharmaceutically acceptable carriers.
4. The pharmaceutical composition according to claim 3, characterized in that: The ratio of flurbiprofen to PD-L1 monoclonal antibody is as follows: flurbiprofen 10 mg / kg body weight, PD-L1 monoclonal antibody 50 μg / animal.