Combination-therapy drug and composition of cholesterolylated TLR7 liposome and TLR9 agonist and use thereof

Abstract

The present invention belongs to the technical field of cancer immunotherapy and specifically relates to a combination-therapy drug and composition of a cholesterolylated TLR7 liposome and a TLR9 agonist and use thereof. Toll-like receptor agonists have poor targeting capability and significant side effects in anti-tumor treatment, and, as monotherapy in anti-tumor treatment, have limited efficacy. To address the described issues, the present invention provides the cholesterolylated TLR7 liposome, which is prepared from the cholesterol-modified 1V209 molecule 1V209-Cho, a lipid component, and cholesterol, and uses the liposome in combination with the TLR9 agonist to treat tumors. Animal results show that the combination of the liposome and the TLR9 agonist, when administered as nasal drops or by means of intramuscular injection, can significantly inhibit pulmonary metastasis of cervical cancer cells and melanoma cells, indicating that using the cholesterolylated liposome 1V209-Cho-Lip in combination with the TLR9 agonist to treat tumors is a strategy with great potential.

Description

Drugs, compositions and applications of combination therapy of cholesterol-modified TLR7 liposomes and TLR9 agonists Technical Field

[0001] This invention belongs to the field of cancer immunotherapy technology, specifically relating to drugs, compositions and their applications involving the combined use of cholesterol-modified TLR7 liposomes and TLR9 agonists. Background Technology

[0002] Cancer immunotherapy aims to trigger a host immune response to fight tumor cells. Several immunotherapies have been developed to stimulate both innate and adaptive immunity within the tumor microenvironment. Among these, cytotoxic CD8... + T lymphocytes (CTLs) are considered the most effective killer cells, eliminating tumor cells by secreting cytokines and releasing cytotoxic granules. Meanwhile, CTL activation primarily depends on antigen-presenting cells (APCs), such as macrophages and dendritic cells (DCs), which present tumor-associated antigens (TAAs) to immature T cells in a process known as cross-presentation.

[0003] Dendritic cells (DCs) are key active clinical cells (APCs) required to activate naive T cells, and their ability to activate T cells largely depends on their maturity state. DC activation in tumor tissue can trigger the secretion of chemokines, leading to antitumor effects. However, in most cases, dendritic cells maintain an immunosuppressive phenotype due to the lack of a tumor microenvironment that stimulates a set of pattern recognition receptors (PRRs) within them. Therefore, PRR-targeting agonists can be used to reverse the immunosuppressive state of DCs and effectively elicit antitumor T cell responses. However, preclinical and clinical studies of PRR agonists as antitumor agents have revealed their limitations, with few receiving FDA approval for the treatment of malignant diseases. The main reason is the serious adverse reactions caused by most PRR agonists, such as imidazoquinolines (TLR7 / 8 agonists) and cyclic di-nuocleotides (STING agonists), likely due to their systemic distribution, leading to the induction of pro-inflammatory cascades and severe immune-related toxicities.

[0004] Toll-like receptors (TLRs) belong to the PRR family and recognize conserved pamps expressed on foreign pathogens. These pamps are selectively expressed on immune cells, including dendritic cells (DCs), macrophages, and effector B / T cell populations. TLR7 is localized to the inner body membrane of innate immune cells and can be activated by viral single-stranded RNA rich in guanosine or uridine, generating a strong Th-1 biased immune response by activating plasma cell and myeloid cell DC subsets. This makes TLR7 on dendritic cells capable of inducing CD8+ in tumor immunotherapy. +T-cell expansion and the induction of adaptive anti-tumor immune responses make TLR7 a promising target. However, previous reports have shown that sustained systemic stimulation of TLR7 can lead to adverse side effects such as lymphopenia, splenomegaly, elevated pro-inflammatory cytokines, and lymphocyte subset contraction, significantly limiting the clinical translation of TLR7 agonists. Although studies have shown that nano-drug delivery systems can reduce the toxic side effects of TLR7 agonists to some extent, most nano-drug delivery systems suffer from complex synthesis, low biocompatibility, and difficulty in clinical translation. Therefore, there is an urgent need to develop a TLR7 agonist drug delivery system that is simple to synthesize, can significantly reduce its toxic side effects, and is easier to translate clinically. Currently, monotherapy in cancer treatment is often ineffective; combination therapy can not only enhance efficacy and reduce drug resistance but also reduce drug toxicity. Therefore, it is also necessary to develop combination therapies of TLR7 agonists with other anti-tumor drugs to achieve the best anti-tumor effect. Summary of the Invention

[0005] To address the issues of poor targeting, significant side effects, and suboptimal efficacy of Toll-like receptor agonists as monotherapy in antitumor treatment, this invention provides a cholesterol-modified TLR7 liposome, prepared from a cholesterol-modified 1V209 molecule (1V209-Cho) combined with lipid components and cholesterol. This liposome is then used in combination with or in combination with a TLR9 agonist for tumor treatment, achieving optimal antitumor efficacy while reducing toxic side effects.

[0006] To achieve the above-mentioned objectives, the technical solution adopted in this application is as follows:

[0007] In a first aspect, the present invention provides a combination antitumor drug comprising cholesterol-modified TLR7 liposomes and TLR9 agonists administered separately or simultaneously.

[0008] The cholesterol-modified TLR7 liposome is a 1V209-Cho-Lip liposome, comprising: a cholesterol-modified 1V209 molecule 1V209-Cho, a lipid component, and cholesterol; the structural formula of the 1V209-Cho is shown in Formula II.

[0009] Formula II:

[0010] In a second aspect, the present invention provides an antitumor pharmaceutical composition comprising cholesterol-modified TLR7 liposomes and a TLR9 agonist as active ingredients, and comprising pharmaceutically acceptable auxiliary ingredients.

[0011] The cholesterol-modified TLR7 liposome is a 1V209-Cho-Lip liposome, comprising: a cholesterol-modified 1V209 molecule 1V209-Cho, a lipid component, and cholesterol; the structural formula of the 1V209-Cho is shown in Formula II.

[0012] Formula II:

[0013] Furthermore, in the above-mentioned pharmaceutical composition, the auxiliary component is at least one of the following: filler, disintegrant, wetting agent, antioxidant, chelating agent, surfactant, flavoring agent, chelating agent, pH adjuster, or colorant.

[0014] Furthermore, in the above-mentioned combined drug or drug composition, the 1V209 has the structural formula shown in Formula I.

[0015] Formula I:

[0016] Furthermore, in the above-mentioned combined drug or drug composition, the lipid component is at least one of lecithin, hydrogenated lecithin, or synthetic phospholipid.

[0017] The lipid component is preferably at least one of soybean lecithin or hydrogenated soybean lecithin.

[0018] The preferred lipid component is hydrogenated soybean lecithin (HSPC).

[0019] Furthermore, in the above-mentioned combined drug or drug composition, the ratio of lipid component, cholesterol and 1V209-Cho is 60-70:27-37:1-8 molar ratio.

[0020] The preferred ratio of lipid components, cholesterol, and 1V209-Cho in the liposomes is 63-68:30-35:2-4.

[0021] The optimal ratio of lipid components, cholesterol, and 1V209-Cho in the liposomes is 65:32:3.

[0022] Furthermore, in the above-mentioned combination drug or drug composition, the 1V209-Cho-Lip liposome is prepared by rotary evaporation, ethanol injection or microfluidic method.

[0023] The rotary evaporation method includes the following steps: dissolving lipid components, cholesterol, and 1V209-Cho in chloroform and methanol, forming a lipid film under rotary evaporation at 37-40°C, hydrating, and sonicating to form liposomes 1V209-Cho-Lip.

[0024] Preferably, the hydration in the rotary evaporation method is performed by hydration in PBS at 37–40°C for 1–1.5 hours.

[0025] Preferably, the ultrasonic treatment in the rotary evaporation method lasts for 30 to 45 minutes; then, the ultrasonic treatment is performed for 2 to 3 minutes using an 80W probe ultrasonic instrument, with an ultrasonic frequency of 5 seconds on and 5 seconds off.

[0026] The ethanol injection method includes the following steps: dissolving lipid components, cholesterol, and 1V209-Cho in ethanol and dimethyl sulfoxide, then adding the solution dropwise at 50-65°C to remove ethanol and dimethyl sulfoxide, thereby obtaining liposomes 1V209-Cho-Lip.

[0027] Preferably, the temperature at which the ethanol is added dropwise by injection is 55–60°C.

[0028] The microfluidic method includes the following steps: dissolving lipid components, cholesterol, and 1V209-Cho in ethanol and dimethyl sulfoxide as the organic phase, and physiological saline as the aqueous phase, with the volume ratio of the aqueous phase to the organic phase being 3:1, and preparing liposomes 1V209-Cho-Lip using microfluidic technology.

[0029] Furthermore, in the above-mentioned combination drug or drug composition, the TLR9 agonist is at least one of CpG 1826, OND 2216 or NOD1018.

[0030] Preferably, the TLR9 agonist is CpG 1826.

[0031] Furthermore, in the above-mentioned combined drug or drug composition, the mass ratio of TLR9 agonist to 1V209-Cho-Lip liposome is 0.8–1.2:3.7–4.2.

[0032] The preferred mass ratio of the TLR9 agonist CpG 1826 to 1V209-Cho-Lip is 1:4.

[0033] Thirdly, the present invention provides the use of the above-mentioned combined drugs or drug compositions in the preparation of antitumor drugs.

[0034] Furthermore, in the above applications, the drug is a pharmaceutical preparation administered by injection, oral administration, nasal mucosa, lung, rectum, oral mucosa, or skin.

[0035] Furthermore, the drug is an injectable preparation, and the injection route is at least one of intramuscular injection, intravenous injection, subcutaneous injection, intradermal injection, intramyocardial injection, or intraperitoneal injection.

[0036] Furthermore, the drug is administered via nasal spray.

[0037] Furthermore, in the above applications, the tumor is at least one of melanoma, cervical cancer, colorectal cancer, breast cancer, pancreatic ductal carcinoma, liver cancer, stomach cancer, uterine cancer, ovarian cancer, testicular cancer, basal cell carcinoma, or lung cancer.

[0038] Beneficial Effects: This invention utilizes cholesterol to modify the TLR7 agonist 1V209, obtaining the liposome 1V209-Cho-Lip, and further combines this liposome with a TLR9 agonist for tumor treatment. Animal experiments showed that in a mouse TC1 cervical cancer lung metastasis model, administration of 1V209-Cho-Lip in combination with CpG 1826 via nasal spray significantly reduced lung metastatic tumor nodules compared to the control group. Furthermore, the combination of 1V209-Cho-Lip and CpG 1826 in treating B16F10 mouse melanoma lung metastasis demonstrated that nasal spray administration reduced the number of lung metastatic nodules, while intramuscular injection in the B16F10 mouse melanoma lung metastasis model resulted in even better anti-tumor effects. In summary, the cholesterol-modified TLR7 agonist 1V209-Cho-Lip obtained in this invention, when used in combination with CpG 1826, exhibits good anti-tumor effects, providing a better combination therapy approach for cancer immunotherapy. Attached Figure Description

[0039] Figure 1 is a schematic diagram of the administration procedure of CpG 1826 and 1V209-Cho-Lip alone in the B16F10 melanoma lung metastasis model in Example 1A); BC) statistical graphs of the lungs and lung nodules of mice in each group after treatment.

[0040] Figure 2 shows a schematic diagram of the combined administration procedure of 1V209-Cho-Lip and CpG 1826 in the TC1 cervical cancer lung metastasis model in Example 2A); BC) lung dissection and weight of mice in each group after treatment; DF) total number of lung nodules in each group (D), total number of lung nodules larger than 3 mm (E), and total number of lung nodules smaller than 3 mm. Data are expressed as mean ± SEM (*p<0.05, **p<0.01, ***p<0.001).

[0041] Figure 3 shows a schematic diagram of the combined administration procedure of 1V209-Cho-Lip and CpG 1826 in the B16F10 melanoma lung metastasis model in Example 3A); Figures BC show the lungs and their weights of mice after dissection in each group after treatment. Data are expressed as mean ± SEM (*p<0.05, **p<0.01, ***p<0.001).

[0042] Figure 4 shows the lung H&E images of mice after combined treatment with 1V209-Cho-Lip and CpG 1826 in the B16F10 melanoma lung metastasis model of Example 4B16F10. Detailed Implementation

[0043] To make the technical problems, solutions, and beneficial effects of this application clearer, the following detailed description is provided in conjunction with the embodiments. Unless otherwise defined, all technical terms used herein have the same meaning as understood by one of ordinary skill in the art.

[0044] 1V209 is a synthetic small-molecule TLR7 agonist with antitumor activity, but it also exhibits strong systemic immunotoxicity. Combination therapy or drug combinations can not only enhance antitumor efficacy and reduce drug resistance, but also mitigate drug toxicity. TLR9, as an important component of the innate immune response, plays a crucial role in recruiting T cells to tumor regions. Therefore, combining or integrating TLR7 agonists with TLR9 agonists for cancer treatment may yield optimal antitumor effects.

[0045] To reduce the systemic immunotoxicity of the TLR7 agonist 1V209, this invention creatively provides a cholesterol-modified TLR7 liposome, 1V209-Cho-Lip, which is prepared by conventional methods, such as rotary evaporation, ethanol injection, or microfluidic methods, using cholesterol-modified 1V209 molecules (1V209-Cho), lipid components, and cholesterol. This liposome is then used in combination with TLR9 agonists such as CpG 1826, OND 2216, or NOD1018 for tumor treatment.

[0046] The lipid component used is lecithin, hydrogenated lecithin, or synthetic phospholipid. Further, the lipid component can be selected from soybean lecithin or hydrogenated soybean lecithin. In this embodiment of the invention, the lipid component used is hydrogenated soybean lecithin (HSPC).

[0047] In one specific embodiment of the present invention, experimental results show that, compared with CpG 1826 and 1V209-Cho-Lip monotherapy, the combination of cholesterol-modified TLR7 liposome 1V209-Cho-Lip and TLR9 agonist CpG 1826 administered via nasal spray or intramuscular injection can significantly inhibit TC1 cervical cancer and B16F10 melanoma lung metastases, demonstrating excellent anti-tumor effects in both tumor models.

[0048] The following specific embodiments will be provided to explain the solution of the present invention. Those skilled in the art will understand that the following embodiments are for illustrative purposes only and should not be considered as limiting the scope of the invention. Where specific techniques or conditions are not specified in the embodiments, they are performed according to the techniques or conditions described in the literature in the field or according to the product instructions. Reagents or instruments whose manufacturers are not specified are all conventional products that can be obtained commercially.

[0049] The main materials used in the following embodiments are as follows:

[0050] 1V209 cells were purchased from Selleck. The mouse TC1 cervical cancer cell line and B16F10 melanoma cell line were obtained from the Cell Bank of the Chinese Academy of Sciences (Shanghai, China). Cells were cultured in RPMI 1640 or DMEM medium (containing 10% fetal bovine serum (Gibco), 100 U / mL). -1 Penicillin G and 100 UmL -1 Animals were cultured with streptomycin sulfate and incubated at 37°C and 5% CO2. Female BALB / c mice and C57BL / 6 mice (6–8 weeks old, 18–20 g) were purchased from Huafukang Biotechnology Co., Ltd. (Beijing, China). All animal experiments were conducted in accordance with the guidelines assessed and approved by the Ethics Committee of Sichuan University.

[0051] Preparation of TLR7 agonist drug delivery system 1V209-Cho-Lip liposomes

[0052] (1) 1V209-Cho was prepared by the following synthetic method:

[0053] Step a: Cholesterol, EDCI, and DMAP were dissolved in 10 mL of DCM at a ratio of 1 eq: 2 eq: 0.1 eq. Then, 1.2 eq of BOC-aminobutyric acid was added. After reacting at room temperature for 24 hours, 25 mL of DCM was added to extract the organic layer. The organic layer was washed with brine, dried over anhydrous sodium sulfate, and concentrated under vacuum to obtain the crude product. The crude product was purified by silica gel chromatography using petroleum ether / ethyl acetate to obtain product 1.

[0054] Product 1CHO-boc:

[0055] Step b: Product 1 was dissolved in 5 mL of DCM and 10% TFA was added. The mixture was stirred at room temperature for 1 h, and 25 mL of DCM was added. The organic layer was extracted, washed with brine, dried with anhydrous sodium sulfate, and concentrated under vacuum to obtain product 2.

[0056] Product 2CHO-NH2:

[0057] In step c, 1V209, HATU, and TEA were dissolved in 5 mL of DMF at a ratio of 1 eq: 1.2 eq: 2 eq. Then, product 2 dissolved in DMF was added at a volume of 1.2 eq. After reacting at room temperature for 48 hours, the solvent DMF was removed by rotary evaporation to obtain the crude product. The crude product was then purified by silica gel column chromatography using dichloromethane / methanol to obtain 1V209-Cho.

[0058] (2) The synthesis of 1V209-Cho-Lip liposomes was performed using a microfluidic method: The lipid components HSPC, cholesterol, and 1V209-Cho were dissolved in anhydrous ethanol and dimethyl sulfoxide at a molar ratio of 65:32:3 to form the organic phase (anhydrous ethanol to dimethyl sulfoxide volume ratio of 10:1). The concentration of 1V209-Cho in the organic phase was 0.8 μg / μL. Physiological saline was used as the aqueous phase. The aqueous phase and organic phase were rapidly passed through a microfluidic chip at a volume ratio of 3:1 to obtain 1V209-Cho-Lip.

[0059] Example 1: Anticancer efficacy of CpG 1826 and 1V209-Cho-Lip alone in a B16F10 lung metastasis model.

[0060] Eight-week-old C57BL / 6 mice were randomly divided into three groups: a PBS control group, a CpG 1826 group, and a 1V209-Cho-Lip group. The CpG 1826 dosage was 250 μg / kg, and the 1V209-Cho-Lip dosage was 1 mg / kg. Six mice were administered the medication via nasal spray on days -7, 0, and 7, and via tail vein injection on day 1. 5 A melanoma lung metastasis model was established using B16F10 cells. Mice were sacrificed on day 13, and their lungs were dissected, photographed, and the number of lung nodules was counted (Figure 1A). Compared with the PBS control group, the CpG 1826 group and the 1V209-Cho-Lip group, administered intranasally alone, did not significantly inhibit B16F10 melanoma lung metastasis (Figures 1B and 1C). There was no significant difference in the number of lung nodules in the two single-administered groups compared with the PBS control group, indicating that intranasal administration of CpG 1826 and 1V209-Cho-Lip alone cannot effectively inhibit tumor lung metastasis.

[0061] Example 2: Anticancer effect of 1V209-Cho-Lip combined with CpG 1826 in a TC1 cervical cancer lung metastasis model.

[0062] Eight-week-old C57BL / 6 mice were randomly divided into two groups: a PBS control group and a 1V209-Cho-Lip + CpG 1826 combined administration group. The CpG 1826 dosage was 250 μg / kg, and the 1V209-Cho-Lip dosage was 1 mg / kg. Nine mice were administered the medication via nasal spray on days -7, 0, and 7, and 2 × 10⁻⁶ mg / kg via tail vein injection on day 1. 5 A TC1 cell line was used to establish a TC1 cervical cancer lung metastasis model. Mice were sacrificed on day 13, and their lungs were dissected, weighed, and photographed to count the number of lung nodules (Figure 2A). Compared with the PBS control group, the combination of 1V209-Cho-Lip and CpG 1826 administered intranasally significantly inhibited TC1 cervical cancer lung metastasis (Figures 2B and 2C), and the number of lung nodules in the treated group was significantly less than that in the PBS control group (Figures 2D-F).

[0063] Example 3: Anticancer effect of 1V209-Cho-Lip combined with CpG 1826 in a B16F10 melanoma lung metastasis model.

[0064] Eight-week-old C57BL / 6 mice were randomly divided into two groups: a PBS control group and a 1V209-Cho-Lip + CpG 1826 combined administration group. The CpG 1826 dosage was 250 μg / kg, and the 1V209-Cho-Lip dosage was 1 mg / kg. Nine mice were administered the drugs via nasal spray or intramuscular injection on days -7, 0, and 7, respectively. On day 1, 2 × 10⁻⁶ CpG 1826 was administered via tail vein injection. 5 A B16F10 melanoma lung metastasis model was established using B16F10 cells. Mice were sacrificed on day 13, and their lungs were dissected, weighed, and photographed (Figure 3A). The combination of 1V209-Cho-Lip and CpG 1826, whether administered intranasally or intramuscularly, significantly inhibited B16F10 melanoma lung metastasis (Figures 3B and 3C).

[0065] Example 4: H&E staining experiment of mouse lungs in a B16F10 melanoma lung metastasis model treated with 1V209-Cho-Lip combined with CpG 1826.

[0066] The lung tissue obtained in Example 3 was fixed in 4% paraformaldehyde. After 24 hours, it was embedded in paraffin and sectioned. The sections were then sequentially immersed in xylene I for 10 min, xylene II for 10 min, anhydrous ethanol I for 5 min, anhydrous ethanol II for 5 min, 95% ethanol for 5 min, 90% ethanol for 5 min, 80% ethanol for 5 min, and 70% ethanol for 5 min, washed with distilled water, and dewaxed. Next, the sections were stained with Harris hematoxylin for 3-8 min, washed with tap water, differentiated with 1% hydrochloric acid alcohol for a few seconds, rinsed with tap water, and then blued with 0.6% ammonia solution, and rinsed with running water. The sections were then stained with eosin for 1-3 min. After staining, the sections were sequentially immersed in 95% ethanol I for 5 min, 95% ethanol II for 5 min, anhydrous ethanol I for 5 min, anhydrous ethanol II for 5 min, xylene I for 5 min, and xylene II for 5 min to dehydrate and clear. The sections were removed from the xylene and slightly dried, then mounted with neutral resin. Microscopic examination and image analysis were performed. The results were consistent with those in Figure 3. H&E staining of mice showed that the number of lung nodules in the mice treated with the combination of 1V209-Cho-Lip and CpG 1826 was significantly less than that in the PBS control group (Figure 4).

Claims

1. A combination antitumor drug, characterized in that: The cholesterized TLR7 liposome and the TLR9 agonist are administered separately or simultaneously. The cholesterized TLR7 liposome is 1V209-Cho-Lip liposome, which comprises a cholesterized 1V209 molecule 1V209-Cho, a lipid component and cholesterol. Formula II:

2. A pharmaceutical composition against tumors, characterized by: The cholesterized TLR7 liposome is 1V209-Cho-Lip liposome, which comprises a cholesterized 1V209 molecule 1V209-Cho, a lipid component and cholesterol. The cholesterized TLR7 liposome is 1V209-Cho-Lip liposome, which comprises a cholesterized 1V209 molecule 1V209-Cho, a lipid component and cholesterol. Formula II:

3. The pharmaceutical composition of claim 2, wherein: The auxiliary component is at least one of a filler, a disintegrant, a wetting agent, an antioxidant, a chelating agent, a surfactant, a flavoring agent, a chelating agent, a pH regulator or a pigment.

4. The combination of claim 1 or the pharmaceutical composition of claim 2 or 3, wherein: The structural formula of 1V209 is shown as formula I, Formula I:

5. The combination of claim 1 or the pharmaceutical composition of claim 2 or 3, wherein: The lipid component is at least one of lecithin, hydrogenated lecithin or synthetic phospholipid; preferably, the lipid component is at least one of soybean lecithin or hydrogenated soybean lecithin; most preferably, the lipid component is HSPC.

6. The combination of claim 1 or the pharmaceutical composition of claim 2 or 3, wherein: The molar ratio of the lipid component, cholesterol and 1V209-Cho is 60-70:27-37:1-8; preferably, the molar ratio is 63-68:30-35:2-4; most preferably, the molar ratio is 65:32:

3.

7. The combination of claim 1 or the pharmaceutical composition of claim 2 or 3, wherein: The 1V209-Cho-Lip liposome is prepared by a rotary evaporation method, an ethanol injection method or a microfluidic method.

8. The combination of claim 1 or the pharmaceutical composition of claim 2 or 3, wherein: The TLR9 agonist is at least one of CpG 1826, OND 2216 or NOD 1018; preferably, the TLR9 agonist is CpG 1826.

9. The combination of claim 1 or the pharmaceutical composition of claim 2 or 3, wherein: The mass ratio of the TLR9 agonist and the 1V209-Cho-Lip liposome is 0.8-1.2:3.7-4.2; preferably, the mass ratio is 1:

4.

10. Use of the combination drug of any one of claims 1, 4-9 or the pharmaceutical composition of any one of claims 2-9 in the preparation of an antitumor drug.

11. Use according to claim 10, characterized in that: The drug is a pharmaceutical preparation for injection, oral administration, nasal mucosa, lung, rectum, oral mucosa or skin; further, the drug is an injection preparation, and the injection route is at least one of intramuscular injection, intravenous injection, subcutaneous injection, intradermal injection, intramyocardial injection or intraperitoneal injection; further, the drug is administered by nasal spray.

12. Use according to claim 10 or 11, characterized in that: The tumor is at least one of melanoma, cervical cancer, colorectal cancer, breast cancer, pancreatic ductal carcinoma, liver cancer, gastric cancer, uterine cancer, ovarian cancer, testicular cancer, basal cell carcinoma or lung cancer.

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