Use of scutellarein in the preparation of a drug for preventing immune checkpoint inhibitor-associated myocarditis

Abstract

The application relates to application of wild scutellarein in preparation of a medicine for preventing ICI-associated-myocarditis (ICIAM). The wild scutellarein can be combined with TIMP1 protein in a targeted manner, competitively inhibit the combination of TIMP1 and its receptor CD74, thereby blocking the activation of the NF-kappa B signal path in macrophages, and then in an ICIAM mouse model, the heart function can be improved in a dose-dependent manner, the myocardial injury marker level is reduced, the myocardial tissue pathological injury is reduced, the transcription level of a local inflammatory factor of the heart is inhibited, and thus the ICI-associated-myocarditis (ICIAM) is effectively prevented.

Description

Technical Field

[0001] This invention belongs to the field of new pharmaceutical uses of baicalin, specifically involving the application of baicalin in the preparation of drugs for the prevention of immune checkpoint inhibitor-associated myocarditis (ICI-Associated Myocarditis, ICIAM). Background Technology

[0002] Immune checkpoint inhibitors (ICIs) represent a major breakthrough in tumor immunotherapy, but the immune-related adverse events (irAEs) caused by their overactivation of the immune system, especially ICIAM, have become a key factor limiting their clinical application and affecting patient survival. ICIAM has an insidious onset, rapid progression, and a mortality rate as high as 39.7%-50%, making it one of the most lethal irAEs.

[0003] Currently, clinical treatment strategies for ICIAM are extremely limited. High-dose glucocorticoids are the first-line treatment, but nearly half of patients still do not respond well; the timing, dosage, and long-term safety of second-line immunosuppressants remain unclear. Therefore, finding safe and effective therapeutic targets and drugs is a pressing clinical challenge in the field of cardio-oncology.

[0004] Existing research indicates that the core pathological component of ICIAM is a cytokine storm triggered by excessive immune activation. Macrophages, as key innate immune cells, are abnormally activated under ICI stimulation, secreting large amounts of pro-inflammatory cytokines such as IL-6 and IL-23. These inflammatory factors directly damage cardiomyocytes and further amplify the inflammatory cascade. However, specific intervention strategies targeting the upstream activation switches of macrophages are still lacking.

[0005] Our team, through in-depth preliminary research, discovered that matrix metalloproteinase tissue inhibitor 1 (TIMP1), a novel inflammatory cytokine, is highly expressed in cardiac-resident macrophages and serves as a key upstream switch in initiating the inflammatory cascade of intracardiac myocardial infarction (ICIAM). TIMP1, by binding to its receptor CD74, activates the macrophage NF-κB signaling pathway, thereby promoting the secretion of IL-6 and IL-23 and driving the formation of a local inflammatory storm in the heart. Therefore, targeting and blocking the TIMP1-CD74 interaction holds promise for interrupting the inflammatory response upstream, potentially representing a breakthrough strategy for treating ICIAM.

[0006] Scutellarin is a flavonoid compound extracted and isolated from the whole herb of *Scutellaria barbata* D. Don, a plant in the Lamiaceae family. Currently, scutellarin (a preparation of *Scutellaria barbata*) has been approved for clinical use in my country for ischemic cardiovascular and cerebrovascular diseases (such as sequelae of stroke, coronary heart disease, and angina pectoris). It has been used as an adjunct therapy for cardiovascular and cerebrovascular diseases for many years with good safety. However, there are no reports to date on whether scutellarin can be used to treat ischemic cardiovascular and cerebrovascular diseases (ICIAM). Summary of the Invention

[0007] Through experimental research, our team has discovered for the first time that baicalin can target and bind to the TIMP1 protein, competitively inhibiting the binding of TIMP1 to its receptor CD74, thereby blocking the activation of the NF-κB signaling pathway in macrophages. Consequently, in an ICIAM mouse model, it can dependently improve cardiac function, reduce the level of myocardial injury markers, alleviate myocardial tissue pathological damage, and inhibit the transcription level of local inflammatory factors in the heart, thus effectively preventing immune checkpoint inhibitor-associated myocarditis (ICIAM). This invention was thus completed.

[0008] This invention provides, in one aspect, the use of baicalin in the preparation of drugs for the prevention of immune checkpoint inhibitor-associated myocarditis (ICIAM).

[0009] The specific information about scutellarin is as follows: Chinese names: Baicalin, Erigeron breviscapine, Erigeron breviscapine, Baicalin; English name: Scutellarin; Chemical name: Scutellarein-7-O-β-D-glucuronide; CAS No.: 27740-01-8; Molecular formula: C 21 H 18 O 12 ; Molecular weight: 462.366; Structural formula: Flavonoid compound, aglycone is baicalin, with a glucuronic acid group attached at position 7. The specific structure is as follows: .

[0010] Baicalin can be obtained from Scutellaria barbata, a plant in the Lamiaceae family (Laminaria japonica). Scutellaria barbataThe baicalin was extracted and isolated from the whole herb of *Scutellaria barbata* (D. Don). *Scutellaria barbata* is widely distributed and abundant, making it an important plant source of baicalin. For example, the dried whole herb of *Scutellaria barbata* was pulverized and extracted by reflux with 70% ethanol. The extract was concentrated and then extracted successively with petroleum ether and ethyl acetate. The ethyl acetate phase was purified by silica gel column chromatography and Sephadex LH-20 gel column chromatography to obtain crude baicalin, which was then purified by preparative HPLC and freeze-dried to obtain the final product. Baicalin can also be obtained from *Erigeron breviscapus* (a plant in the Asteraceae family) and *Scutellaria baicalensis* (a plant in the Asteraceae family). Scutellaria baicalensis Extracted from plants such as stems and leaves of Georgi.

[0011] Furthermore, the immune checkpoint inhibitors include, but are not limited to, anti-PD-1 antibodies, anti-PD-L1 antibodies, anti-CTLA-4 antibodies, or any combination thereof. ICIAM refers to myocardial inflammatory disease induced by treatment with the aforementioned immune checkpoint inhibitors.

[0012] Anti-PD-1 antibodies can be, for example, pembrolizumab, nivolumab, cemiplimab, toripalimab, sintilimab, tislelizumab, and camrelizumab, but are not limited to these.

[0013] Anti-PD-L1 antibodies can be, for example, durvalumab, atezolizumab, and avelumab, but are not limited to these.

[0014] Anti-CTLA-4 antibodies can be, for example, ipilimumab and tremelimumab, but are not limited to these.

[0015] Furthermore, the mechanism of action of the drug is as follows: baicalin targets and binds to the TIMP1 protein, competitively inhibiting the binding of TIMP1 to its receptor CD74, thereby blocking the activation of the NF-κB signaling pathway in macrophages, specifically manifested as follows: (a) Inhibit macrophages from secreting pro-inflammatory cytokines IL-6 and IL-23; (b) Reduce the levels of inflammatory factors (IL-6, IL-23, IL-1β, TNF-α, etc.) in serum and cardiac tissue; (c) Improve cardiac function and reduce the levels of myocardial injury markers (CTnI, NT-proBNP); (d) Reduces cardiac inflammatory cell infiltration and myocardial fibrosis.

[0016] Figure 1 This invention presents a mechanistic hypothesis diagram, which illustrates the pathological process by which ICIs lead to high expression of TIMP1 in cardiac macrophages, TIMP1 binding to CD74 to activate NF-κB, promote IL-6 / IL-23 secretion, thereby triggering the ICIAM inflammatory storm, and the targeted blocking of TIMP1-CD74 binding upstream of this pathway by baicalin.

[0017] Furthermore, the drug comprises a therapeutically effective amount of baicalin and a pharmaceutically acceptable carrier or excipient. The dosage form of the drug is any suitable for systemic administration (e.g., intravenous injection, intramuscular injection, oral administration), including but not limited to injections, lyophilized powder for injection, tablets, capsules, granules, etc.

[0018] Furthermore, in the aforementioned drug, the daily dosage range of baicalin is 10-200 mg / kg body weight, preferably 30-90 mg / kg body weight. Based on clinical experience, the recommended human dosage is as follows: oral administration of 40 mg three times daily, or intravenous injection / infusion of 20-50 mg daily.

[0019] In another aspect, the present invention provides a pharmaceutical composition comprising therapeutically effective amounts of the following active pharmaceutical ingredients: (1) baicalin; (2) immune checkpoint inhibitors.

[0020] The pharmaceutical composition may also contain a pharmaceutically acceptable carrier or excipient. The dosage form of the pharmaceutical composition may be any dosage form suitable for systemic administration (such as intravenous injection, intramuscular injection, or oral administration), including but not limited to injections, lyophilized powder for injection, tablets, capsules, granules, etc.

[0021] In another aspect, the present invention provides the use of the above-described pharmaceutical composition in the preparation of a medicament for treating tumors while simultaneously preventing immune checkpoint inhibitor-associated myocarditis (ICIAM).

[0022] When used to treat tumors, the pharmaceutical composition according to the present invention can effectively prevent immune checkpoint inhibitor-associated myocarditis (ICIAM) caused by the overactivation of the immune system by immune checkpoint inhibitors.

[0023] The description of immune checkpoint inhibitors is as previously stated.

[0024] The tumors can be melanoma, non-small cell lung cancer (NSCLC), gastrointestinal tumors (esophageal cancer, gastric cancer, hepatocellular carcinoma (in combination with anti-angiogenic drugs), colorectal cancer (MSI-H / dMMR only)), urinary system tumors (renal cell carcinoma, urothelial carcinoma), and classical Hodgkin lymphoma.

[0025] Beneficial effects Compared with the prior art, this application has the following outstanding beneficial effects and significant technological advancements: (1) First discovery and identification of a novel therapeutic target: This invention establishes the TIMP1-CD74 signaling axis as a key upstream target for the treatment of ICIAM for the first time, surpassing the existing strategy of targeting only downstream inflammatory factors. As the first small molecule inhibitor targeting the interaction of TIMP1-CD74, baicalin provides the core material basis for the development of a first-in-class drug for ICIAM with independent intellectual property rights.

[0026] (2) Clear targeting and high affinity: Surface plasmon resonance (SPR) experiments showed that the equilibrium dissociation constant KD value of baicalin and TIMP1 protein was as low as 10.11 nM, which has extremely high binding affinity, providing a structural basis for its competitive blocking of TIMP1 binding to CD74.

[0027] (3) Significant in vitro efficacy: Baicalin (10 μM) can significantly inhibit the secretion of IL-6 and IL-23 by LPS-stimulated macrophages; at the same time, it inhibits the activation of the NF-κB signaling pathway in macrophages induced by TIMP1 (p65 phosphorylation).

[0028] (4) Clear in vivo efficacy and prominent cardioprotective effect: In the ICIAM mouse model, treatment with baicalin (30, 60, 90 mg / kg / d) can improve cardiac function (increase EF, FS) in a dose-dependent manner, reduce the level of myocardial injury markers (NT-proBNP, CTnI), alleviate myocardial tissue pathological damage (inflammatory infiltration and fibrosis), and inhibit the transcription level of local cardiac inflammatory factors (IL-6, IL-23).

[0029] (5) Good safety profile and no impact on antitumor efficacy: Preliminary cytotoxicity experiments showed that baicalin had no significant toxicity to macrophages and cardiomyocytes within the effective concentration range. The 4T1 tumor cell killing experiment further verified that baicalin exerts cardioprotective effects while also having antitumor effects, and has the potential for clinical combination therapy.

[0030] (6) The source is clear and conforms to the theory of traditional Chinese medicine: This study classifies ICIAM into the category of "heart jaundice" in traditional Chinese medicine, which is a severe heart disease caused by heat and toxic evils lodging in the heart and damaging the myocardium and blood vessels. Wild baicalin is derived from the traditional Chinese medicine Scutellaria barbata, which clears heat and detoxifies. Its application conforms to the pathogenesis of ICIAM in traditional Chinese medicine, and reflects the innovative path of "using ancient medicine in new ways" and "modernizing traditional Chinese medicine". It has a good foundation for clinical translation. Attached Figure Description

[0031] Figure 1This invention presents a hypothetical mechanism diagram. It illustrates the pathological process by which ICIs lead to high expression of TIMP1 in cardiac macrophages, TIMP1 binding to CD74 to activate NF-κB, promote IL-6 / IL-23 secretion, thereby triggering the ICIAM inflammatory storm, and the targeted blocking of TIMP1-CD74 binding upstream of this pathway by baicalin.

[0032] Figure 2 SPR sensing map (A) and dose-response curve (B) of baicalin binding to TIMP1 protein.

[0033] Figure 3 Baicalin inhibits the secretion of IL-6 and IL-23 by macrophages.

[0034] Figure 4 Evaluation of the efficacy of baicalin in alleviating cardiac function decline in ICIAM mice. (A, B) Echocardiographic results showed that baicalin dose-dependently increased EF and FS in ICIAM mice. (C, D) ELISA results showed that baicalin reduced serum NT-proBNP and CTnI levels. Figure 5 Evaluation of the efficacy of baicalin in reducing myocardial injury in ICIAM mice. (A) Hematoxylin and eosin (H&E) staining of myocardial tissue showed that baicalin reduced inflammatory infiltration in a dose-dependent manner. (B) Masson staining of myocardial tissue showed that baicalin reduced fibrosis levels in a dose-dependent manner. (C) Ratio of inflammatory cell infiltration rate to fibrosis area derived from A and B.

[0035] Figure 6 Effects of baicalin on inflammatory factors in the cardiac tissue of ICIAM mice.

[0036] Figure 7 Baicalin dose-dependently inhibited the phosphorylation levels of TIMP1 and NF-κB p65 in the heart of a mouse model.

[0037] Figure 8 Baicalin can inhibit the proliferation of 4T1 tumor cells.

[0038] Figure 4-7 In this study, IgG served as the normal control group, aPD-1+aCTLA-4 served as the model control group, Scu-L / M / H served as the low / medium / high dose groups of baicalin, and Dex served as the dexamethasone group. Detailed Implementation

[0039] To more clearly illustrate the present invention, specific embodiments are provided below for further explanation. These embodiments are for illustrative purposes only and do not constitute a limitation on the scope of protection of the present invention.

[0040] raw material The baicalin used in the examples was an analytical standard purchased from Shanghai Yuanye Biotechnology Co., Ltd. (product number B21478). It is a pale yellow crystalline powder, and its purity is ≥98% as identified by HPLC-MS and NMR.

[0041] 0.5% sodium carboxymethyl cellulose (CMC-Na) solution: Dissolve 0.5 mg CMC-Na in 100 ml ddH2O.

[0042] Preparation method For cell experiments, accurately weigh an appropriate amount of baicalin, dissolve it in DMSO to prepare a 100 mM stock solution, aliquot it, and store it at -20°C protected from light. During experiments, dilute with cell culture medium or physiological saline to the required concentration (final DMSO concentration <0.1%). For animal experiments, prepare a suspension of the appropriate concentration using 0.5% sodium carboxymethyl cellulose (CMC-Na) solution.

[0043] Example 1: Determination of the binding affinity between baicalin and TIMP1 protein 1.1 Experimental Methods Surface plasmon resonance (SPR) technology was employed. Recombinant human TIMP1 protein (purchased from Sinocare Biotechnology Co., Ltd.) was immobilized on a CM5 chip (purchased from Cytiva) via amino-coupling. Different concentrations (0.625 nM–320 nM) of baicalin solutions (in PBS-P buffer containing 5% v / v DMSO (purchased from Cytiva)) were flowed through the chip surface at a flow rate of 30 μL / min, with a binding time of 120 seconds and a dissociation time of 300 seconds. Using a Biacore T200 system and analysis software, a 1:1 Langmuir binding model was used to fit the sensor map, and the equilibrium dissociation constant KD was calculated.

[0044] 1.2 Experimental Results Figure 2 The SPR sensing map (A) and dose-response curve (B) show the binding of baicalin to TIMP1 protein.

[0045] like Figure 2 As shown, the KD value of baicalin binding to TIMP1 protein is 10.11 nM, indicating that the two have extremely high binding affinity, providing a structural basis for baicalin's competitive blocking of TIMP1 binding to CD74.

[0046] Example 2: In vitro experiment on the inhibition of macrophage inflammatory factor secretion by baicalin 2.1 Experimental Methods RAW264.7 macrophages were cultured in DMEM complete medium (Thermo Fisher Scientific, 11995065). A blank control group, a model group (stimulated with lipopolysaccharide (LPS) 500 ng / mL for 12 h), and a group with the optimal concentration of baicalin (10 μM baicalin added 2 h before LPS stimulation) were set up and cultured together for 24 h. Cell supernatants were collected, and IL-6 and IL-23 levels were detected using an ELISA kit.

[0047] 2.2 Experimental Results Figure 3 The study showed that baicalin inhibited the secretion of IL-6 (A) and IL-23 (B) by macrophages. Ctrl was the blank control group, Model was the model group, and Scu was the optimal concentration of baicalin.

[0048] Figure 3 The results showed that, compared with the model group, baicalin at a concentration of 10 μM significantly inhibited LPS-induced IL-6 and IL-23 secretion (*p<0.01). This result confirms that the optimal concentration of baicalin (10 μM) inhibits the levels of IL-6 and IL-23 in the culture supernatant of LPS-stimulated macrophages, and that baicalin can effectively inhibit the inflammatory activation of macrophages.

[0049] Example 3: In vivo efficacy evaluation of baicalin in an ICIAM mouse model 3.1 Model Construction Six- to eight-week-old C57BL / 6J mice were selected and injected intraperitoneally with anti-mouse PD-1 antibody (25 mg / kg) and anti-CTLA-4 antibody (25 mg / kg) every three days for a total of five injections to induce the ICIAM model.

[0050] A normal control group was set up, and an equal amount of IgG isotype control (50 mg / kg) was injected.

[0051] The PD-1 antibody was Anti-mouse PD-1 (CD279)-InVivo, purchased from Selleck, clone number: RMP1-14, A2122; the CTLA-4 antibody was Anti-mouse CTLA-4 (CD152)-InVivo, purchased from Selleck, clone number: 9H10, A2103; and the IgG was Rat IgG2a isotype control-InVivo, purchased from Selleck, clone number: 2A3, A2123. All three monoclonal antibodies were diluted to a working solution of 5 mg / ml with sterile PBS (pH 7.4).

[0052] 3.2 Grouping and Dosing The model mice were randomly divided into a model control group (administered by gavage with 0.2 ml of 0.5% sodium carboxymethyl cellulose solution), low-, medium-, and high-dose groups of baicalin (30, 60, and 90 mg / kg / day, administered by gavage for 14 consecutive days), and a positive control group (administered by intraperitoneal injection of dexamethasone, 1.5 / 0.75 mg / kg / day, 1.5 mg / kg / day for the first 3 days and 0.75 mg / kg / day for the next 4 days, for 7 consecutive days). There were 10 mice in each group.

[0053] 3.3 Detection Indicators and Results 3.3.1 Cardiac function Echocardiography was performed on mice after the last administration. Results are shown below. Figure 4 In the A and B groups, IgG was the normal control group, aPD-1+aCTLA-4 was the model control group, Scu-L / M / H were the low / medium / high dose groups of baicalin, and Dex was the dexamethasone group.

[0054] The results showed that, compared with the model control group, the left ventricular ejection fraction (EF%) and fractional shortening (FS%) of mice in the high-dose baicalin group were significantly increased (*p<0.01), approaching the normal level.

[0055] 3.3.2 Myocardial Injury Markers Blood was collected and serum was separated to detect NT-proBNP and CTnI. Results are shown below. Figure 4 C and D, where IgG is the normal control group, aPD-1+aCTLA-4 is the model control group, Scu-L / M / H are the low / medium / high dose groups of baicalin, and Dex is the dexamethasone group.

[0056] The results showed that, compared with the model control group, the serum NT-proBNP and CTnI levels in the treatment group mice were significantly reduced (*p<0.01), indicating that baicalin can improve cardiac function and reduce the levels of myocardial injury markers.

[0057] 3.3.3 Cardiac Histopathology Heart tissue was taken for H&E and Masson staining. Results are shown below. Figure 5 A (H&E staining map), B (Masson staining map), and C (ratio of inflammatory cell infiltration rate and fibrosis area obtained from A and B), where IgG is the normal control group, aPD-1+aCTLA-4 is the model control group, Scu-L / M / H are the low / medium / high dose groups of baicalin, respectively, and Dex is the dexamethasone group.

[0058] The results showed that the myocardial tissue in the model control group exhibited extensive inflammatory cell infiltration and significant fibrosis. The baicalin treatment group showed improved myocardial structure and a significant reduction in inflammatory infiltration and fibrosis area, indicating that baicalin can alleviate cardiac inflammatory cell infiltration and myocardial fibrosis.

[0059] 3.3.4 Levels of myocardial inflammatory factors Heart tissue homogenate was collected, and the levels of IL-6, IL-1β, and IL-23 were detected by qPCR. Results are shown below. Figure 6 Among them, IgG was the normal control group, aPD-1+aCTLA-4 was the model control group, and Scu-L / H were the low-dose and high-dose groups of baicalin, respectively.

[0060] The results showed that baicalin treatment significantly reduced the transcriptional levels of inflammatory factors IL-6, IL-1β, and IL-23 in cardiac tissue homogenate.

[0061] 3.3.5 Activation level of myocardial inflammatory pathways Heart tissue homogenate was collected, and Western blot was used to detect the phosphorylation levels of TIMP1 and NF-κB p65. Results are shown below. Figure 7 IgG was the normal control group, aPD-1+aCTLA-4 was the model control group, and Scu-L / M / H were the low / medium / high dose groups of baicalin, respectively.

[0062] The results showed that baicalin dose-dependently inhibited the phosphorylation levels of TIMP1 and NF-κB p65 in the heart of a mouse model, thus baicalin treatment could significantly inhibit the activation of inflammatory pathways.

[0063] Example 4: Evaluation of the efficacy of baicalin in inhibiting the proliferation of 4T1 tumor cells in vitro 4.1 Experimental Methods 4T1 mouse triple-negative breast cancer cells (purchased from the Cell Resource Center of the Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences and Peking Union Medical College), frozen at -80°C, were resuspended in DMEM medium (Thermo Fisher Scientific, 11995065) and cultured in a 37°C, 5% CO2 incubator. When the cells reached 80-90% confluence, they were digested by pipetting, centrifuged to collect the cells, resuspended, and counted. A blank control group and gradient groups of baicalin (5 / 10 / 20 / 40 μM) were set up, and the cell suspension density was adjusted to 5 × 10⁶ cells / mL. 3Cells were seeded at a density of 1 mL / well in 96-well plates, with 1 mL of cell suspension added to each well. The plates were incubated overnight (12 h) at 37°C with 5% CO2 to allow complete cell adhesion. The original culture medium in the 96-well plates was discarded, and the drug-treated groups were incubated with diluted drug-containing medium. After 24 h of incubation, the original medium was discarded, and basal medium containing 10% v / v CCK-8 (purchased from Meilun Biotechnology, MA0218) (purchased from Thermo Fisher Scientific, 11995065) was added. The plates were incubated at 37°C with 5% CO2 for 1 h, and the absorbance was measured at 450 nm using a microplate reader.

[0064] 4.2 Experimental Results Figure 8 This demonstrates the efficacy of baicalin in inhibiting the proliferation of 4T1 tumor cells.

[0065] Figure 8 The results showed that, compared with the blank control group, baicalin significantly inhibited the proliferation of 4T1 tumor cells in the concentration range of 5-40 μM.

Claims

1. Application of baicalin in the preparation of drugs for the prevention of immune checkpoint inhibitor-associated myocarditis.

2. The application according to claim 1, wherein, The immune checkpoint inhibitor is selected from one or more of anti-PD-1 antibody, anti-PD-L1 antibody, and anti-CTLA-4 antibody.

3. The application according to claim 1, wherein, Baicalin targets and binds to TIMP1 protein, inhibits the interaction between TIMP1 and CD74, and blocks the activation of the macrophage NF-κB signaling pathway.

4. The application according to claim 1, wherein, Baicalin exerts one or more of the following functions: (a) Inhibit macrophages from secreting pro-inflammatory cytokines IL-6 and IL-23; (b) Reduce the levels of inflammatory factors in serum and cardiac tissue; (c) Improves cardiac function and reduces the levels of myocardial injury markers; (d) Reduces inflammatory cell infiltration and fibrosis in cardiac tissue.

5. The application according to claim 1, wherein, The drug contains a therapeutically effective amount of baicalin and a pharmaceutically acceptable carrier or excipient.

6. The application according to claim 1, wherein, The dosage form of the drug is selected from injection, lyophilized powder for injection, tablets, capsules, and granules.

7. The application according to claim 1, wherein, In the aforementioned drug, the daily dosage range of baicalin is 10-200 mg / kg body weight.

8. The application according to claim 7, wherein, The daily dosage range for baicalin was selected as 30-90 mg / kg body weight.

9. A pharmaceutical composition comprising a therapeutically effective amount of the following active pharmaceutical ingredients: (1) baicalin; (2) an immune checkpoint inhibitor.

10. Use of the pharmaceutical composition according to claim 9 in the preparation of a medicament for treating tumors while simultaneously preventing immune checkpoint inhibitor-associated myocarditis.

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