Composition for treating solid malignant tumor, and kit for treating solid malignant tumor

Abstract

Provided is a new therapeutic composition. This composition for treating a solid malignant tumor is used so as to be administered to a subject having malignant tumor cells that produce at least one inflammatory cytokine among TNFα, IL-1β, IL-5, IL-6, IL-8, IL-17, and IL-23, in combination with at least one of immature dendritic cells, and cytotoxic lymphocytes induced by dendritic cells, and contains at least one antibody that inhibits the action of the inflammatory cytokine.

Description

Composition for treating solid malignant tumors and kit for treating solid malignant tumors

[0001] The present invention relates to a composition for treating solid malignant tumors and a kit for treating solid malignant tumors.

[0002] Cancer vaccine therapy using dendritic cells (hereinafter referred to as "dendritic cell vaccine therapy") is known as a cancer treatment method. In a typical dendritic cell vaccine therapy, immature dendritic cells collected from a patient are stimulated with an antigen in vitro, and the dendritic cells that have recognized the antigen are administered subcutaneously or into the lymph nodes of the patient to induce cytotoxic T cells (CTLs). Dendritic cell vaccine therapy is usually used in combination with radiation therapy (e.g., Non-Patent Documents 1 to 4).

[0003] Jesslca A Cintolo et. al., Dendritic cell-based vaccines: barriers and opportunities, Future Oncol. 2012 Oct;8(10): p. 1273-1299Jifeng Yu et. al., Resaerch progress on dendritic cell vaccines in cancer immunotherapy, Exp Hematol Oncol. 2022 Jan 24;11(1):3Ivan Y. Filin et. al., Recent Advances in Experimental Dendritic Cell Vaccines for Cancer, Front Oncol. 2021 Sep 23;11Beatris Mastelic-Gavillet et. al., Personalized Dendritic Cell Vaccines-Recent Breakthroughs and Encouraging Clinical Results, Front Immunol. 2019 Apr 11;10:766

[0004] However, known dendritic cell vaccine therapies are sometimes insufficient to achieve sufficient effects in regressing, reducing, or eliminating cancer, and there is a need for the development of novel cancer treatment methods.

[0005] The present invention has been made in view of the above problems, and an object of the present invention is to provide a novel therapeutic composition for treating solid malignant tumors.

[0006] In order to solve the above problems, the present inventors conducted extensive research and found that administering at least one type of immature dendritic cells and cytotoxic lymphocytes induced by dendritic cells to a subject having malignant tumor cells that produce a specific inflammatory cytokine, in combination with administering at least one type of antibody that inhibits the action of the inflammatory cytokine produced by the malignant tumor cells, is effective in treating solid malignant tumors, leading to the completion of the present invention.

[0007] In order to achieve the above object, a composition for treating solid malignant tumors according to one embodiment of the present invention is used to be administered to a subject having malignant tumor cells that produce at least one inflammatory cytokine selected from tumor necrosis factor α, interleukin-1β, interleukin-5, interleukin-6, interleukin-8, interleukin-17, and interleukin-23, in combination with at least one of immature dendritic cells and cytotoxic lymphocytes induced by dendritic cells, and contains at least one antibody that inhibits the action of the inflammatory cytokine.

[0008] Furthermore, one aspect of the present invention provides a kit for treating solid malignant tumors, which is used to be administered to a subject having malignant tumor cells that produce at least one inflammatory cytokine selected from tumor necrosis factor α, interleukin-1β, interleukin-5, interleukin-6, interleukin-8, interleukin-17, and interleukin-23, in combination with at least one of immature dendritic cells and cytotoxic lymphocytes induced by dendritic cells, and which comprises at least one antibody that inhibits the action of the inflammatory cytokine.

[0009] According to one aspect of the present invention, a novel therapeutic composition for treating solid malignant tumors can be provided.

[0010] FIG. 1 is a diagram showing Example 1 of administration of a composition for treating solid malignant tumors according to one embodiment of the present invention. FIG. 2 is a diagram showing Example 2 of administration of a composition for treating solid malignant tumors according to one embodiment of the present invention. FIG. 3 is a diagram showing Example 3 of administration of a composition for treating solid malignant tumors according to one embodiment of the present invention. It is a CT image showing the treatment results of Example 1. It is a CT image showing the treatment results of Example 2. It is a CT image showing the treatment results of Example 3. It is a CT image (left image) and an MRI image (right image) showing the treatment results of Example 4. It is a CT image showing the treatment results of Example 5. It is a PET image showing the treatment results of Example 2.

[0011] [1. Composition for Treating Solid Malignant Tumors] (Features) A ​​composition for treating solid malignant tumors according to one embodiment of the present invention (hereinafter also simply referred to as "therapeutic composition") is used to be administered to a subject having malignant tumor cells that produce at least one inflammatory cytokine selected from tumor necrosis factor α, interleukin-1β, interleukin-5, interleukin-6, interleukin-8, interleukin-17, and interleukin-23 in combination with at least one type of immature dendritic cells and cytotoxic lymphocytes induced by dendritic cells, and contains at least one antibody that inhibits the action of the inflammatory cytokine. In this specification, "immature dendritic cells" may be abbreviated as "iDCs," "dendritic cells" as "DCs," and "cytotoxic T cells" as "CTLs."

[0012] A therapeutic composition according to one embodiment of the present invention is expected to have the effect of regressing, reducing, or eliminating tumor cells in solid malignant tumor tissues by administering it in combination with at least one of iDCs and CTLs induced by DCs according to the above-mentioned usage method.

[0013] Furthermore, a therapeutic composition according to one embodiment of the present invention, when administered in combination with at least one of iDCs and CTLs induced by DCs according to the above-described method, is expected to have the effect of regressing, reducing, or eliminating tumor cells in solid malignant tumor tissues without the need for concomitant radiation therapy. It also has the advantage of having few side effects. Therefore, by using a therapeutic composition according to one embodiment of the present invention, a new cancer treatment method can be provided that is applicable to subjects who were previously ineligible for radiation therapy / chemotherapy due to reasons such as age, number of tumors, or tumor size. Therefore, treatment of solid malignant tumors can be provided to a wider range of subjects than previously possible. Furthermore, a therapeutic composition according to one embodiment of the present invention contains at least one inflammatory cytokine-inhibiting antibody corresponding to the type of inflammatory cytokine produced by the malignant tumor cells of the subject. Therefore, a cancer treatment method tailored to the subject (individualized) can be provided. Such effects can contribute, for example, to Goal 3 of the Sustainable Development Goals (SDGs) advocated by the United Nations, "Good Health and Well-Being."

[0014] However, the therapeutic composition according to one aspect of the present invention may be used in combination with other treatment methods useful for treating the target solid malignant tumor, if necessary, including, but not limited to, radiation therapy using X-rays, gamma rays, etc., particle beam therapy, surgical treatment such as surgery, chemotherapy, and molecular targeted therapy.

[0015] As used herein, "treatment" includes causing tumor cells in solid malignant tumor tissue to regress or decrease compared to before administration of the therapeutic composition in a subject to which the therapeutic composition according to one aspect of the present invention is administered, eliminating (eliminating) tumor cells in solid malignant tumor tissue, or preventing the progression of a solid malignant tumor.

[0016] A therapeutic composition according to one embodiment of the present invention contains at least one antibody that inhibits the action of at least one inflammatory cytokine produced by tumor cells, selected from tumor necrosis factor α, interleukin-1β, interleukin-5, interleukin-6, interleukin-8, interleukin-17, and interleukin-23. Therefore, the antibody inhibits the action of the inflammatory cytokine produced by tumor cells, thereby preventing or reducing inflammation in tumors that produce the inflammatory cytokine. Since inflammation in tumor tissue is thought to be a factor in inducing tumor cell mutation and promoting tumor progression, eliminating, reducing, or preventing inflammation in tumor tissue with a therapeutic composition according to one embodiment of the present invention is thought to bring about the therapeutic effect described above.

[0017] Additionally, the therapeutic composition according to one aspect of the present invention can provide regression, reduction or elimination of tumor cells in tumor tissue, which can be visually detected by MRI and / or CT and / or echo scanning.

[0018] As used herein, "solid malignant tumor" refers to any malignant tumor excluding blood cancer. In other words, solid malignant tumor refers to any solid cancer and malignant tumor of the brain. Examples of solid malignant tumors include lung cancer, rectal cancer, uterine cancer, gastric cancer, and pancreatic cancer. From the viewpoint of highly safe treatment, the therapeutic composition according to one embodiment of the present invention is preferably used for treating elderly people and cancers for which standard treatment is not applicable. Solid malignant tumors include both early-stage cancers and advanced cancers. Solid malignant tumors also include tumors formed by metastasis. As used herein, "malignant tumor" refers to solid malignant tumors.

[0019] (Antibody) A therapeutic composition according to one embodiment of the present invention contains at least one antibody that inhibits the action of at least one inflammatory cytokine selected from tumor necrosis factor α (TNFα), interleukin-1β (IL-1β), interleukin-5 (IL-5), interleukin-6 (IL-6), interleukin-8 (IL-8), interleukin-17 (IL-17), and interleukin-23 (IL-23). ​​As used herein, "inflammatory cytokine" refers to a cytokine that causes inflammatory symptoms in the body. Furthermore, as used herein, an antibody that inhibits the action of an inflammatory cytokine may be referred to as an "inflammatory cytokine-inhibiting antibody," and an antibody that inhibits the action of a specific inflammatory cytokine may be referred to as, for example, an "IL-6 inhibitory antibody," an "IL-5 inhibitory antibody," etc.

[0020] The type of inflammatory cytokine-inhibitory antibody is not particularly limited, as long as it specifically inhibits the action of inflammatory cytokines produced by malignant tumor cells in a subject. For example, the inflammatory cytokine-inhibitory antibody may be a polyclonal antibody, a monoclonal antibody (e.g., IgG, IgM, IgE, IgA, IgD, etc.), a modified antibody (e.g., a chimeric antibody, a humanized antibody, or a fully human antibody), an antibody fragment (e.g., Fab, Fab', F(ab')), or a soluble antibody (e.g., soluble antibody). ２ The present invention includes any antibody that can be used for pharmaceutical purposes, such as antibody fragments (e.g., scFv, etc.).

[0021] An inflammatory cytokine-inhibitory antibody is also called an “inflammatory cytokine-neutralizing antibody.” An inflammatory cytokine-inhibitory antibody may be, for example, an antibody that specifically binds to a target inflammatory cytokine (ligand) and thereby inhibits the action of the inflammatory cytokine to which the antibody binds, or an antibody that specifically binds to a receptor of the target inflammatory cytokine and thereby inhibits the action of the target inflammatory cytokine.

[0022] The proinflammatory cytokine inhibitory antibody may be a commercially available antibody, such as tocilizumab, a humanized anti-IL-6 receptor antibody, or mepolizumab, a humanized anti-IL-5 receptor antibody.

[0023] A therapeutic composition according to one embodiment of the present invention may contain at least one antibody that inhibits the action of an inflammatory cytokine produced by solid malignant tumor cells, selected from the group of inflammatory cytokines consisting of TNFα, IL-1β, IL-5, IL-6, IL-8, IL-17, and IL-23. When the therapeutic composition is administered to a subject having solid malignant tumor cells that produce multiple inflammatory cytokines selected from the group of inflammatory cytokines consisting of TNFα, IL-1β, IL-5, IL-6, IL-8, IL-17, and IL-23, the therapeutic composition may contain multiple antibodies that inhibit the action of each inflammatory cytokine. A method for screening for inflammatory cytokines produced by solid malignant tumor cells will be described below in [3. Method for treating solid malignant tumors].

[0024] When administered to a subject having solid malignant tumor cells producing one or more inflammatory cytokines, including, for example, IL-6, from the group of inflammatory cytokines consisting of TNFα, IL-1β, IL-5, IL-6, IL-8, IL-17, and IL-23, a therapeutic composition according to one aspect of the present invention preferably comprises at least an IL-6 inhibitory antibody as an antibody that inhibits the action of the inflammatory cytokines. A therapeutic composition according to one aspect of the present invention often comprises at least an IL-6 inhibitory antibody and an IL-5 inhibitory antibody. This is because IL-6 and IL-5 are highly expressed in tumors and contribute particularly significantly to inflammation. Therefore, a therapeutic composition comprising at least an IL-6 inhibitory antibody and an IL-5 inhibitory antibody can effectively eliminate, reduce, or prevent inflammation caused by an immune response in malignant tumor tissue, thereby enabling regression, reduction, or elimination of tumor cells in solid malignant tumor tissue. However, when the therapeutic composition according to one aspect of the present invention is to be administered to a subject having solid malignant tumor cells that do not produce IL-6 or IL-5, the therapeutic composition according to one aspect of the present invention does not need to contain an IL-6 inhibitory antibody or an IL-5 inhibitory antibody as an antibody that inhibits the action of inflammatory cytokines.

[0025] Furthermore, when administered to a subject having solid malignant tumor cells producing two or more inflammatory cytokines, including IL-6 and IL-5, from the group of inflammatory cytokines consisting of TNFα, IL-1β, IL-5, IL-6, IL-8, IL-17, and IL-23, the therapeutic composition according to one aspect of the present invention preferably comprises at least an IL-5 inhibitory antibody and an IL-6 inhibitory antibody as antibodies that inhibit the action of the inflammatory cytokines. As described above, IL-6 and IL-5 are highly expressed in tumors and contribute significantly to inflammation. A therapeutic composition comprising an IL-5 inhibitory antibody and an IL-6 inhibitory antibody can suppress inflammation caused by both IL-5 and IL-6, thereby more effectively eliminating, reducing, or preventing inflammation in malignant tumor tissue, and as a result, can cause regression, reduction, or elimination of tumor cells in solid malignant tumor tissue.

[0026] When the therapeutic composition according to one aspect of the present invention is to be administered to a subject for whom it is difficult to identify the type of cytokine being produced by solid malignant tumor cells, and when it is predicted that the solid malignant tumor cells are producing at least one inflammatory cytokine selected from the group of inflammatory cytokines consisting of TNFα, IL-1β, IL-5, IL-6, IL-8, IL-17, and IL-23, even if the type of cytokine being produced by the solid malignant tumor cells is unknown, the therapeutic composition according to one aspect of the present invention preferably comprises at least an IL-5 inhibitory antibody and an IL-6 inhibitory antibody as antibodies that inhibit the action of inflammatory cytokines.

[0027] The present inventors have found that the expression rates of inflammatory cytokines in solid malignant tumor cells are approximately 90% for TNFα and IL-1β, and that the positive rates tend to decrease in the following order: IL-6, IL-5, IL-8, IL-23, and IL-17. However, when a therapeutic composition according to one aspect of the present invention is administered to a subject having solid malignant tumor cells that are found to have a tendency to express such inflammatory cytokines, the therapeutic composition may contain at least one antibody that inhibits the action of an inflammatory cytokine produced by solid malignant tumor cells from the inflammatory cytokine group consisting of IL-6, IL-5, IL-8, IL-23, and IL-17. Alternatively, the therapeutic composition may be administered in combination with an antibody that inhibits the action of an inflammatory cytokine other than TNFα, without administering an antibody that inhibits the action of TNFα.

[0028] A therapeutic composition according to one embodiment of the present invention comprises a proinflammatory cytokine-inhibitory antibody as an active ingredient. The therapeutic composition according to one embodiment of the present invention may also comprise an active ingredient other than the proinflammatory cytokine-inhibitory antibody. The content of the active ingredient in the therapeutic composition according to one embodiment of the present invention is not particularly limited, and may be, for example, 0.001% to 100% by weight, 0.01% to 100% by weight, 0.1% to 100% by weight, 0.1% to 95% by weight, 0.1% to 90% by weight, 0.1% to 80% by weight, 0.1% to 70% by weight, 0.1% to 60% by weight, 0.1% to 50% by weight, 0.1% to 40% by weight, 0.1% to 30% by weight, 0.1% to 20% by weight, or 0.1% to 10% by weight, relative to the total weight of the therapeutic composition according to one embodiment of the present invention.

[0029] (Other Components) The therapeutic composition according to one embodiment of the present invention may contain components other than the above-described inflammatory cytokine-inhibitory antibody, as necessary. The other components may be any pharmaceutically acceptable components, such as buffers, pH adjusters, isotonicity agents, preservatives, antioxidants, high-molecular-weight polymers, excipients, and solvents. These components may be substances typically contained in therapeutic compositions, and the types thereof are not particularly limited.

[0030] The therapeutic composition according to one aspect of the present invention may contain, as the other component, a medicinal ingredient having a desired effect, such as reducing side effects and helping to suppress inflammation.

[0031] The content of other components in a therapeutic composition according to one embodiment of the present invention is not particularly limited, and may be, for example, 0 wt % to 99.999 wt %, 0 wt % to 99.99 wt %, 0 wt % to 99.9 wt %, 5 wt % to 99.9 wt %, 10 wt % to 99.9 wt %, 20 wt % to 99.9 wt %, 30 wt % to 99.9 wt %, 40 wt % to 99.9 wt %, 50 wt % to 99.9 wt %, 60 wt % to 99.9 wt %, 70 wt % to 99.9 wt %, 80 wt % to 99.9 wt %, or 90 wt % to 99.9 wt %, relative to the total weight of the therapeutic composition according to one embodiment of the present invention.

[0032] (Formulations and Dosage Forms) The therapeutic composition according to one embodiment of the present invention can be formulated by known methods using the active ingredient, an inflammatory cytokine-inhibitory antibody, and other ingredients as raw materials.

[0033] The dosage form of the therapeutic composition according to one embodiment of the present invention is not particularly limited, but from the viewpoint of ease of administration into the tumor or blood vessel of a subject, a liquid pharmaceutical formulation is preferred, for example, an injectable formulation.

[0034] (Method of Use) A therapeutic composition according to one embodiment of the present invention is used to be administered to a subject having malignant tumor cells that produce at least one inflammatory cytokine selected from TNFα, IL-1β, IL-5, IL-6, IL-8, IL-17, and IL-23, in combination with iDCs and at least one CTL induced by DCs. When administered to a subject in combination with iDCs and at least one CTL induced by DCs, the therapeutic composition according to one embodiment of the present invention can eliminate, reduce, or prevent inflammation caused by immune responses in malignant tumors, resulting in the regression, reduction, or elimination of tumor cells in solid malignant tumor tissues.

[0035] Herein, the timing of administering at least one of iDCs and CTLs induced by DCs in combination with a therapeutic composition is not particularly limited. For example, an inflammatory cytokine-inhibitory antibody and at least one of iDCs and CTLs induced by DCs may be administered simultaneously, or an inflammatory cytokine-inhibitory antibody may be administered at a predetermined interval before or after administration of at least one of iDCs and CTLs induced by DCs, or these may be combined. From the viewpoint of effectively inhibiting inflammation caused by CTLs induced by DCs, it is preferable to administer a therapeutic composition according to one aspect of the present invention simultaneously with iDCs and at least one of CTLs induced by DCs.

[0036] When at least one of iDCs and CTLs induced by DCs and the therapeutic composition are administered at intervals, it is preferable to administer the therapeutic composition within 90 days after administration of at least one of iDCs and CTLs induced by DCs, from the viewpoint of suppressing transformation into an inflammatory tumor.

[0037] As used herein, "immature dendritic cells" or "iDCs" refer to dendritic cells that have not been stimulated with an antigen and have the ability to phagocytose antigens. Immature dendritic cells include a dendritic cell population that is positive for the myeloid markers CD11c and CD14, positive for the costimulatory markers CD14, CD86, and HLA-DR, and negative for the dendritic cell maturation marker CD83. The origin of immature dendritic cells is not particularly limited, but autologous immature dendritic cells obtained from the subject can be preferably used from the viewpoint of preventing rejection reactions. Autologous immature dendritic cells can be prepared by a method of culturing a monocytic cell fraction of peripheral blood mononuclear cells (PBMCs) collected from the subject, a method of obtaining iDCs from hematopoietic stem cells collected from the subject, a method of obtaining iDCs by apheresis from the subject, or the like.

[0038] Immature dendritic cells are administered intratumorally, whereby they are sensitized within the tumor, for example, at the tumor site, and present a comprehensive range of tumor antigens, including both known and unknown tumor antigens, on their surface. Dendritic cells that present tumor antigens activate T cells and induce tumor antigen-specific CTLs.

[0039] The iDCs administered in combination with a therapeutic composition according to one aspect of the present invention may be added with an adjuvant prior to administration to induce their maturation. Examples of adjuvants include, but are not limited to, lipid-based, protein-based, and polysaccharide-based adjuvants such as lymphocyte culture medium, marignase, agaricus, OK432, BCG, lentinan (shiitake mushroom), reishi mushroom, polyporus mushroom, TNF-Meshimakobu, incomplete or complete Freund's adjuvant, LPS, fatty acids, TW80, phospholipids, cytokines, or viruses. In certain embodiments, the adjuvant may be a leukocyte culture medium (LCM) adjuvant. The LCM adjuvant can be at least three cytokines selected from the group consisting of eotaxin, FGF, G-CSF, GM-CSF, IFNγ, IP10, IL-1β, IL-1ra, IL-2, IL-4, IL-5, IL-6, IL-7, IL-8, IL-9, IL-10, IL-12, IL-13, IL-15, IL-17, MCP1, MIP1α, MIP1β, PDGFbb, RANTES, TNFα, and VEGF.

[0040] As used herein, "cytotoxic lymphocytes" or "CTLs" refer to cytotoxic lymphocytes induced by dendritic cells. CTLs include CTLs induced by dendritic cells that are sensitized and matured within a tumor after administration of immature dendritic cells to a subject. Autologous CTLs obtained from a subject can be preferably used as CTLs. Autologous CTLs can be prepared by culturing a T cell-enriched fraction of peripheral blood mononuclear cells (PBMCs) collected from a subject. Alternatively, CTLs may be induced and prepared by artificially sensitizing T cells in vitro to a limited, known antigen. In this case, the CTLs are effective against malignant tumors that respond to the limited antigen.

[0041] CTLs are a key component of cell-mediated immunity. They play a crucial role in controlling many infectious diseases and cancers. These T cells are responsible for "hunting down" and destroying other cells in the body that are infected with a virus or contain cancer. For example, when a virus or cancer uses a cell to replicate, the cell displays several viral proteins or cancer components on its surface. Cytotoxic T cells recognize these proteins or components and pursue them, destroying the infected or cancer-containing cells before they can release new infections or cancers into the bloodstream. Many vaccines are effective, at least in part, by stimulating the activation or response of this type of T cell. CTLs can also produce chemicals known as cytokines, which help regulate how the immune system fights disease.

[0042] Furthermore, a therapeutic composition according to one aspect of the present invention is preferably administered in combination with activated T cells (AT). AT comprises a cell population that is positive for the lymphocyte markers CD3 and CD4 and the T cell activation markers CD25 and CD154 (CD40L). Autologous AT cells obtained from a subject can be suitably used as AT. Autologous AT can be prepared by culturing a T cell-enriched fraction of peripheral blood mononuclear cells (PBMCs) collected from a subject. Administration of AT in combination efficiently produces CTLs, thereby enhancing the effect of shrinking solid malignant tumors. Furthermore, AT artificially expressing CD154 functions as pre-CTLs. After receiving antigen information from DCs, AT artificially expressing CD154 transforms into mature CTLs and forms immunological memory. Therefore, administration of AT in combination efficiently produces CTLs.

[0043] Furthermore, the therapeutic composition according to one embodiment of the present invention is preferably administered in combination with an anti-inflammatory agent other than a proinflammatory cytokine-inhibitory antibody. The anti-inflammatory agent is not particularly limited, and any anti-inflammatory agent available for pharmaceutical use can be used. Examples of such anti-inflammatory agents include corticosteroids and nonsteroidal anti-inflammatory agents. Examples of corticosteroids include dexamethasone, prednisolone, clopetazole propionate, betamethasone propionate, and hydrocortisone butyrate. Because the therapeutic composition according to one embodiment of the present invention is expected to have not only an anti-inflammatory effect but also an appetite-stimulating effect and an antiemetic effect, it is preferably administered to a subject in combination with dexamethasone. Furthermore, when the antibody is to be administered to a subject having malignant tumor cells that produce at least one inflammatory cytokine selected from the group of inflammatory cytokines consisting of TNFα, IL-1β, IL-5, IL-6, IL-8, IL-17, and IL-23, a corticosteroid may be administered to the subject in place of administering at least one of an IL-17 inhibitory antibody and an IL-23 inhibitory antibody, from the viewpoint of cost.

[0044] Dexamethasone may be in the form of a "pharmaceutically acceptable salt." That is, as used herein, the term "dexamethasone" encompasses pharmaceutically acceptable salts. As used herein, the term "pharmaceutically acceptable salt" refers to a salt that is physiologically acceptable for administration to a subject as a pharmaceutical, and specific examples thereof are not limited thereto. Examples of salts include alkali metal salts (potassium salt, etc.), alkaline earth metal salts (calcium salt, magnesium salt, etc.), ammonium salts, organic base salts (trimethylamine salt, triethylamine salt, pyridine salt, picoline salt, dicyclohexylamine salt, N,N'-dibenzylethylenediamine salt, etc.), organic acid salts (acetate, maleate, tartrate, methanesulfonate, benzenesulfonate, formate, toluenesulfonate, trifluoroacetate, etc.), and inorganic acid salts (hydrochloride, hydrobromide, sulfate, phosphate, etc.).

[0045] Furthermore, the therapeutic composition according to one aspect of the present invention may be administered in combination with a drug other than an antibody that inhibits the action of TNFα.

[0046] The therapeutic composition according to one embodiment of the present invention can be administered to a subject via any administration route. Examples of administration routes include intra-arterial administration, intravenous administration, intramuscular administration, intraperitoneal administration, intratumoral administration, intrathoracic administration, and subcutaneous administration. From the viewpoint of more easily obtaining an effect, the therapeutic composition according to one embodiment of the present invention is preferably administered into a malignant tumor of the subject.

[0047] As used herein, the terms "intravenous" and "through the blood vessels," as well as related terms using "through the blood vessels," refer to treatments that involve the administration of components and combinations thereof that can be administered to a subject into channels, such as veins or arteries, that carry fluids within a patient's body.

[0048] As used herein, the term "intratumoral therapy" and related terms using "intratumoral" refer to treatments that involve administering subject-administered components and combinations thereof directly to the tumor tissue of a patient.

[0049] Examples of methods of using the therapeutic composition will be described with reference to Figures 1 to 3. In Administration Examples 1 to 3 described below, a therapeutic composition containing an IL-6 inhibitory antibody and an IL-5 inhibitory antibody as inflammatory cytokine-inhibitory antibodies is administered to a subject. However, the inflammatory cytokine-inhibitory antibodies that can be used in the therapeutic composition according to one aspect of the present invention are not limited to the inflammatory cytokine-inhibitory antibodies used in Administration Examples 1 to 3; for example, only an IL-6 inhibitory antibody may be used.

[0050] FIG. 1 shows Example 1 of administration of a therapeutic composition. First, the therapeutic composition is administered to a subject simultaneously with iDCs and dexamethasone. Then, AT is administered within 24 hours or more and within 72 hours. In the therapeutic protocol shown in FIG. 1, administration of iDCs can induce comprehensive tumor antigen-specific CTLs, including both known and unknown tumor antigens. In the therapeutic protocol shown in FIG. 1, the therapeutic composition is administered simultaneously with administration of iDCs and dexamethasone. This method of administration can eliminate, reduce, or prevent inflammation in solid malignant tumor tissues caused by an immune response by CTLs induced by administration of iDCs, resulting in effective regression, reduction, or elimination of tumor cells in solid malignant tumor tissues.

[0051] FIG. 2 shows Example 2 of administration of a therapeutic composition. First, the therapeutic composition is administered to a subject simultaneously with iDCs and dexamethasone, followed by administration of AT within 24 to 72 hours. After a 2- to 6-week CTL production period, CTLs are isolated from peripheral blood mononuclear cells and cultured for 2 to 6 weeks. The therapeutic composition is then administered intratumorally together with the induced CTLs and dexamethasone. In the treatment protocol shown in FIG. 2, CTLs induced in the subject's body by administration of iDCs are collected from the subject, cultured in vitro, and reintroduced into the subject. This allows for the introduction of a sufficient amount of CTLs (e.g., a therapeutically effective amount) into the subject to effectively regress, reduce, or eliminate tumor cells in solid malignant tumor tissue.

[0052] In the treatment protocol shown in Figure 2, the therapeutic composition is administered simultaneously with the administration of iDCs and dexamethasone, and simultaneously with the administration of induced CTLs and dexamethasone. By using such a method, the therapeutic composition can eliminate, reduce, or prevent inflammation in solid malignant tumor tissues caused by an immune response by induced CTLs, thereby effectively regressing, reducing, or eliminating tumor cells in solid malignant tumor tissues.

[0053] Figure 3 shows Example 3 of the administration of a therapeutic composition. First, the therapeutic composition is administered to a subject simultaneously with iDCs and dexamethasone, followed by AT. After CTL induction, CTLs are isolated from peripheral blood mononuclear cells and cultured. The induced CTLs, iDCs, dexamethasone, and the therapeutic composition are then administered intratumorally. In the treatment protocol shown in Figure 3, iDCs can take up tumor cells damaged by the induced CTLs. Furthermore, administration of iDCs, induced CTLs, and dexamethasone can improve the CTL version. As a result, it is possible to treat malignant tumor cells that could not be regressed, reduced, or eliminated by the initial administration, or tumor cells in tumor tissue newly formed by metastasis.

[0054] In the treatment protocol shown in Figure 3, the therapeutic composition is administered simultaneously with the administration of iDCs and dexamethasone, and simultaneously with the administration of induced CTLs and dexamethasone. By using this method, the therapeutic composition can eliminate, reduce, or prevent inflammation in solid malignant tumor tissues caused by an immune response by induced CTLs, thereby effectively regressing, reducing, or eliminating tumor cells in solid malignant tumor tissues.

[0055] In the treatment protocols shown in Figures 2 and 3, iDCs, induced CTLs, dexamethasone, and the therapeutic composition may be administered into the tumor where the therapeutic composition, iDCs, and dexamethasone were initially administered and into another tumor (e.g., a new lesion) six months or more after the initial administration of the therapeutic composition, iDCs, and dexamethasone. By administering iDCs, induced CTLs, and dexamethasone to new lesions in cases where tumor cell mutation has progressed, CTL version-up can be achieved. The therapeutic composition is administered simultaneously with the administration of iDCs, induced CTLs, and dexamethasone. This method of administration can eliminate, reduce, or prevent inflammation in solid malignant tumor tissue caused by an immune response by induced CTLs, resulting in effective regression, reduction, or elimination of tumor cells in solid malignant tumor tissue.

[0056] As an example of such a treatment protocol, for example, iDCs, a therapeutic composition, and dexamethasone can be administered, followed (e.g., 2 to 4 months later, preferably 3 months later) by CTLs, a therapeutic composition, and dexamethasone, and then (e.g., 2 to 4 months later, preferably 3 months later) by iDCs, CTLs, a therapeutic composition, and dexamethasone can be administered to new lesions. In this case, the degree of treatment achieved at each administration step can be about 50% after the first administration, about 80% after the second administration, and about 100% (complete remission) after the third administration.

[0057] 3, the therapeutic composition, iDCs, and dexamethasone may be repeatedly administered after a certain period of time has elapsed since the initial administration of the therapeutic composition, iDCs, and dexamethasone. By repeating the administration, malignant tumor cells that could not be regressed, reduced, or eliminated by the initial administration or tumor cells in tumor tissue newly formed by metastasis can be repeatedly treated.

[0058] (Dosage) When a therapeutic composition according to one embodiment of the present invention is administered to a subject, there is no limitation on the amount administered to the subject as long as the desired effect is achieved. For example, a therapeutic composition according to one embodiment of the present invention may be administered so that the dose of the proinflammatory cytokine-inhibitory antibody is 0.1 mg to 1000.0 mg / kg body weight, 0.1 mg to 500.0 mg / kg body weight, 1.0 mg to 500.0 mg / kg body weight, 1.0 mg to 300.0 mg / kg body weight, 1.0 mg to 100.0 mg / kg body weight, 1.0 mg to 50.0 mg / kg body weight, 1.0 mg to 10.0 mg / kg body weight, 1.0 mg to 10.0 mg / kg body weight, or 1.0 to 5.0 mg / kg body weight.

[0059] (Subject) The subject has malignant tumor cells that produce at least one inflammatory cytokine selected from TNFα, IL-1β, IL-5, IL-6, IL-8, IL-17, and IL-23.

[0060] As used herein, a "subject" includes mammals, including humans. As used herein, a "subject" may also be referred to as a "patient." The subject is preferably a mammal, more preferably a human. Examples of non-human mammals also include companion animals such as dogs and cats.

[0061] 2. Kit for Treating Solid Malignant Tumors (Features) A ​​kit for treating solid malignant tumors according to one embodiment of the present invention (hereinafter also simply referred to as a "treatment kit") is used to be administered to a subject having malignant tumor cells that produce at least one inflammatory cytokine selected from tumor necrosis factor α, interleukin-1β, interleukin-5, interleukin-6, interleukin-8, interleukin-17, and interleukin-23, in combination with at least one type of immature dendritic cells and cytotoxic lymphocytes induced by dendritic cells, and comprises at least one antibody that inhibits the action of the inflammatory cytokine.

[0062] The effects thereof are the same as those explained for the composition for treating solid malignant tumors of the present invention, and therefore will not be repeated here.

[0063] A therapeutic kit according to one embodiment of the present invention includes at least one inflammatory cytokine-inhibitory antibody. The user may administer a mixture of the inhibitory antibodies to a subject, or may administer each inhibitory antibody to a subject at different times. The inflammatory cytokine-inhibitory antibody and the dosage and administration of the inhibitory antibody are the same as those described in [1. Composition for treating solid malignant tumors].

[0064] The treatment kit according to one aspect of the present invention may include components other than the inflammatory cytokine-inhibitory antibody, as needed, such as reagents other than the inflammatory cytokine-inhibitory antibody, instruments, and instructions for use of the treatment kit.

[0065] The components other than the inflammatory cytokine-inhibitory antibody are the same as those described in [1. Composition for treating solid malignant tumors].

[0066] Examples of the device include a device for preparing a reagent and a device for administering the reagent to a subject.

[0067] The instruction manual for the treatment kit may include, for example, the dosage and administration described in [1. Composition for treating solid malignant tumors], or may include an example of a specific procedure for the treatment method described in [3. Method for treating solid malignant tumors] described below.

[0068] [3. Method for Treating Solid Malignant Tumors] (Features) A ​​method for treating solid malignant tumors according to one embodiment of the present invention (hereinafter simply referred to as the "treatment method") comprises the following steps (1) and at least one of (2) and (3) to (5): (1) screening for inflammatory cytokines produced by malignant tumor cells possessed by a subject from the group consisting of tumor necrosis factor α, interleukin-1β, interleukin-5, interleukin-6, interleukin-8, interleukin-17, and interleukin-23; (2) administering to the subject immature dendritic cells and at least one antibody that inhibits the action of the inflammatory cytokine screened in step (1); (3) collecting peripheral blood mononuclear cells from the subject; (4) culturing the collected peripheral blood mononuclear cells to form dendritic cell-induced cytotoxic lymphocytes; and (5) A step of administering the formed cytotoxic lymphocytes and at least one antibody that inhibits the action of the inflammatory cytokine screened in the step (1) to the subject.

[0069] According to one aspect of the present invention, a therapeutic method can be expected to have the effect of regressing, reducing, or eliminating tumor cells in solid malignant tumor tissue without the need for concomitant radiation therapy. Another advantage is that it has few side effects. Side effects of radiation therapy include, for example, potentially induced mutations and unknown mutations that occur when malignant tumor cells are lysed by irradiation. Therefore, the therapeutic method according to one aspect of the present invention can provide a new cancer treatment method that can be applied to subjects who were previously ineligible for radiation therapy / chemotherapy due to factors such as age, number of tumors, or tumor size. Therefore, treatment of solid malignant tumors can be provided to a wider range of subjects than previously possible.

[0070] However, the therapeutic method according to one aspect of the present invention may be used in combination with other treatment methods useful for treating the target solid malignant tumor, if necessary, including, but not limited to, radiation therapy using X-rays, gamma rays, etc., particle beam therapy, surgical treatment such as surgery, chemotherapy, and molecular targeted therapy.

[0071] A therapeutic method according to one aspect of the present invention comprises at least one antibody that inhibits the action of at least one inflammatory cytokine produced by tumor cells, selected from the group consisting of TNFα, IL-1β, IL-5, IL-6, IL-8, IL-17, and IL-23, and thus the antibody inhibits, neutralizes, or blocks the action of the inflammatory cytokine produced by tumor cells, thereby preventing or reducing inflammation in tumors that produce the inflammatory cytokine. Since inflammation in tumor tissue is thought to be one of the factors that induce mutation in tumor cells and promote tumor progression, it is thought that eliminating, reducing, or preventing inflammation in tumor tissue by a therapeutic method according to one aspect of the present invention will bring about the above-mentioned therapeutic effect.

[0072] Additionally, the treatment method according to one aspect of the present invention can provide regression, reduction or elimination of tumor cells in tumor tissue, which can be visually detected by MRI and / or CT and / or echo scanning.

[0073] The subject to which the treatment method according to one embodiment of the present invention is applied is the same as the subject described in [1. Composition for treating solid malignant tumors].

[0074] In the treatment method according to one aspect of the present invention, administration to a subject can be carried out via any administration route. Examples of administration routes are the same as those described in [1. Composition for treating solid malignant tumors]. In the treatment method according to one aspect of the present invention, the administration route is preferably administration into the malignant tumor of the subject, from the viewpoint of more easily obtaining an effect.

[0075] The treatment method according to one aspect of the present invention may include step (1) and step (2), or may include step (1) and steps (3) to (5), or may include steps (1) to (5).

[0076] When the treatment method according to one embodiment of the present invention comprises steps (1) and (2), iDCs and at least one antibody that inhibits the action of an inflammatory cytokine produced by malignant tumor cells of the subject, selected from the group of inflammatory cytokines consisting of TNFα, IL-1β, IL-5, IL-6, IL-8, IL-17, and IL-23, may be administered to the subject.

[0077] When the treatment method according to one embodiment of the present invention comprises step (1) and steps (3) to (5), CTLs induced by DCs and at least one antibody that inhibits the action of an inflammatory cytokine produced by malignant tumor cells of the subject, selected from the group of inflammatory cytokines consisting of TNFα, IL-1β, IL-5, IL-6, IL-8, IL-17, and IL-23, may be administered to the subject.

[0078] When the treatment method according to one embodiment of the present invention comprises steps (1) to (5), the following may be administered to a subject: iDCs; at least one antibody that inhibits the action of a pro-inflammatory cytokine produced by malignant tumor cells of the subject, selected from the group of pro-inflammatory cytokines consisting of TNFα, IL-1β, IL-5, IL-6, IL-8, IL-17, and IL-23; and CTLs induced by the DCs.

[0079] When a therapeutic method according to one embodiment of the present invention comprises steps (1) to (5), steps (3) to (5) may be performed after step (2), or steps (2) and (5) may be performed simultaneously. When steps (3) to (5) are performed after step (2), the resulting CTLs may include CTLs induced by DCs that are matured from the administered iDCs. In other words, by administering patient-derived iDCs to tumor tissue in which all known and unknown antigens (neoantigens) coexist, the present invention can produce, for example, significantly potent and comprehensive CTLs that can respond to a greater number of tumors using natural human immune functions.

[0080] As used herein, "generic" CTLs refer to both known and unknown antigen-driven CTLs, eg, each CTL has independence / specificity for each antigen.

[0081] (Step (1)) Step (1) is a step of screening for inflammatory cytokines produced by the malignant tumor cells present in a subject from the group consisting of TNFα, IL-1β, IL-5, IL-6, IL-8, IL-17, and IL-23. The screening method is not particularly limited, and can be performed by any known method capable of screening inflammatory cytokines. For example, a qualitative screening method includes a method in which malignant tumor cells are immunostained using an antibody that specifically recognizes each cytokine, and the presence or absence of expression of an inflammatory cytokine protein in the tumor cells is confirmed. Instead of confirming the presence or absence of expression of an inflammatory cytokine, the presence or absence of expression of an inflammatory cytokine receptor protein in the tumor cells may be confirmed. Furthermore, a quantitative screening method includes a method in which the amount of inflammatory cytokine in the blood secreted from tumor cells is measured.

[0082] When confirming the presence or absence of expression of inflammatory cytokines in malignant tumors, any malignant tumor among the malignant tumors to be treated may be screened. It is preferable to collect cells from at least two sites of any malignant tumor and screen them.

[0083] Furthermore, screening may be performed for at least one of the above-mentioned inflammatory cytokines, or multiple types may be selected for screening. As described above, the expression rates of inflammatory cytokines in solid malignant tumor cells are approximately 90% for TNFα and IL-1β, and the positive rates tend to decrease in the following order: IL-6, IL-5, IL-8, IL-23, and IL-17. Therefore, from the perspective of improving screening efficiency, cytokines with higher positive rates may be preferentially confirmed for expression. IL-6 and IL-5 have high expression rates in tumors and contribute particularly significantly to inflammation. Therefore, it is preferable to first screen for at least one of IL-6 and IL-5.

[0084] Step (1) may be performed at any timing before administering to a subject an antibody that inhibits the action of a screened inflammatory cytokine. Furthermore, when an antibody that inhibits the action of a screened inflammatory cytokine is administered to a subject multiple times, the screening only needs to be performed once, and the screening may be performed each time before administering the antibody. For example, inflammatory cytokines produced by any malignant tumor may be screened, and then steps (2) or (5) may be performed, followed by screening for inflammatory cytokines produced by malignant tumors that have not regressed, decreased, or been eliminated, or by metastatic malignant tumors.

[0085] (Step (2)) Step (2) is a step of administering iDCs and at least one antibody that inhibits the action of the inflammatory cytokine screened in step (1) to a subject. According to step (2), by administering iDCs to a subject, the iDCs are sensitized within the tumor, for example, at the tumor site, and present comprehensive tumor antigens including both known and unknown tumor antigens on their cell surface. Dendritic cells that present tumor antigens activate T cells and induce tumor antigen-specific CTLs. Therefore, administration of iDCs can actively promote an immune response to induce comprehensive CTLs. The iDCs are the same as those described in [1. Composition for treating solid malignant tumors].

[0086] In step (2), the iDC and the inflammatory cytokine-inhibitory antibody may be administered simultaneously, or the other of the iDC and the inflammatory cytokine-inhibitory antibody that has not been administered may be administered at a predetermined interval before or after the administration of at least one of the iDC and the inflammatory cytokine-inhibitory antibody.

[0087] Alternatively, the iDCs and the pro-inflammatory cytokine inhibitory antibody may be combined to form a composition, and the formed composition may be administered intratumorally to a patient.

[0088] The dose of iDCs administered may be a therapeutically effective amount for solid malignant tumors. As used herein, the term "therapeutically effective amount" refers to the amount of a component effective in treating solid malignant tumors required to produce a desired effect in a human or other mammal. As used herein, "a component effective in treating solid malignant tumors" refers to iDCs, ATs, comprehensive CTLs, inflammatory cytokine-inhibitory antibodies, dexamethasone, anti-inflammatory agents, adjuvants, or combinations thereof. In all cases, at its most basic level, the desired effect is the regression, reduction, or elimination of tumor cells in the patient's tumor tissue compared to the tumor cells in the patient's tumor tissue prior to the use of the treatments and methods of the present invention.

[0089] iDCs are administered to a subject once, for example, at a dose of 5×10 ６ pieces ~ 1×10 ７ The dose of iDCs can be adjusted appropriately depending on the size of the tumor, and it is preferable to increase the dose as the tumor size increases.

[0090] Step (2) may include a step of collecting iDCs from the subject. Methods for collecting iDCs from a subject include, for example, collecting monocytes from the subject and culturing the collected monocytes to form iDCs, obtaining iDCs from hematopoietic stem cells, and obtaining iDCs by apheresis. In the former case, collection and culture of monocytes can be performed by known methods. For example, monocytes can be obtained by collecting and isolating peripheral blood mononuclear cells from the subject. Subsequently, activated T cells can be prepared using a T cell-enriched fraction of peripheral blood mononuclear cells depleted of monocytes. The culture medium may vary and can be selected from those known in the art. Non-limiting examples include IL-4, GM-CFS, and mixtures thereof. In the latter case, suitable methods include methods conventionally known in the art. Furthermore, when the number of tumors in a subject is small, there is no need to prepare a large amount of CTLs. Therefore, from the viewpoint of ease of CTL preparation and reduction of the physical burden on the subject, it is preferable to collect 200 to 400 mL of peripheral blood from the subject and induce CTLs.

[0091] Furthermore, without intending to be bound by any particular theory, it is believed that iDCs formed by culturing monocyte cells collected from a patient, and induced comprehensive CTLs generated by and collected from a patient, have improved therapeutic efficacy when administered to the same patient, compared to iDCs and CTLs generated and obtained by other means. Induced comprehensive CTLs collected, cultured, and re-administered to the same patient, and iDCs formed from the patient's own collected monocyte cells, provide improved coupling or interaction with other cells within the patient's body.

[0092] The antibody administered to the subject may be at least one antibody that inhibits the action of the inflammatory cytokine screened in step (1). When multiple inflammatory cytokines are screened in step (1), a physician can appropriately select which inflammatory cytokine to administer to the subject with an antibody that inhibits the action of that cytokine. When multiple inflammatory cytokines including at least one of IL-6 and IL-5 are screened, it is preferable to administer at least either an IL-6 inhibitory antibody or an anti-IL-5 inhibitory antibody as the inflammatory cytokine inhibitory antibody, from the viewpoint of the magnitude of their contribution to inflammation. The inhibitory antibody is the same as that described for the inhibitory antibody in [1. Composition for treating solid malignant tumors].

[0093] The dose of an antibody that inhibits the action of inflammatory cytokines can be a therapeutically effective amount. The inhibitory antibody can be administered to a subject once or multiple times. When administering multiple times, it is preferable to administer the next administration 30 days after the previous administration. The dose of the inhibitory antibody can be, for example, one-tenth of the determined systemic dose of each inhibitory antibody.

[0094] Step (2) may comprise administering iDCs and an inflammatory cytokine-inhibitory antibody to a subject, and then administering AT to the subject. AT may be administered immediately after or a short time after the administration of iDCs and an inflammatory cytokine-inhibitory antibody. Alternatively, autologous AT may be administered within about 24 hours to about 72 hours after the administration of iDCs and an inflammatory cytokine-inhibitory antibody. The dose of AT is, for example, 1 x 10 per administration. ８ pieces ~ 2×10 ９ The dose of AT can be adjusted appropriately depending on the size of the tumor, and it is preferable to increase the dose as the tumor size increases.

[0095] AT can be collected from a subject, cultured, and then administered back to the subject. The culture medium can vary and can be selected from those known in the art. Non-limiting examples include, but are not limited to, IL-2, CD3, and mixtures thereof. AT can also be obtained from a subject by apheresis. Suitable methods of apheresis include those conventionally known in the art.

[0096] The iDCs administered in step (2) may be those to which an adjuvant has been added prior to administration to induce maturation. From the viewpoint of suitability for human-derived therapeutic vaccines (HIV / AIDS), the adjuvant preferably contains leukocyte culture medium (LCM). Examples of adjuvants that can be used in the present invention are the same as those described for adjuvants in [1. Composition for treating solid malignant tumors].

[0097] (Step (3)) Step (3) is a step of collecting peripheral blood mononuclear cells (PBMCs) from a subject. PBMCs can be obtained by collecting blood from a subject and isolating the PBMCs.

[0098] Generally, PBMCs are collected from a patient (who has received autologous immature dendritic cells and proinflammatory cytokine-inhibitory antibodies) for culture to form induced comprehensive CTLs if present in sufficient numbers in the patient's immune system.

[0099] In a specific embodiment in which step (3) is performed after step (2), it is preferable to collect PBMCs 2 to 6 weeks after administration of immature dendritic cells to the subject, in order to allow a period for induction of cytotoxic lymphocytes.

[0100] In addition, in a specific embodiment in which AT is administered to a subject in step (2), a comprehensive cytotoxic T lymphocyte (CTL) induction period of about 2 to 6 weeks is provided after the administration of AT. Following the comprehensive CTL induction period, peripheral blood mononuclear cells (PBMCs) can be collected from the patient (who has been administered iDCs and a proinflammatory cytokine-inhibitory antibody).

[0101] (Step (4)) Step (4) is a step of culturing the collected PBMCs to form dendritic cell-induced CTLs. The PBMCs can be cultured by known methods. The culture medium may vary and can be selected from those known in the art. Non-limiting examples include, but are not limited to, IL-2, CD3, and a mixture thereof.

[0102] (Step (5)) Step (5) is a step of administering to a subject the formed CTLs and at least one antibody that inhibits the action of the inflammatory cytokine screened in step (1). The CTLs are the same as those described in [1. Composition for treating solid malignant tumors].

[0103] According to the present invention, a sufficient quantity and quality of induced comprehensive CTLs are administered to the patient's body, particularly to the tumor site, to shrink, reduce, or eliminate tumor cells. It is contemplated that the quantity and quality of induced comprehensive CTLs administered in the first administration (e.g., intratumoral injection) may be sufficient (e.g., a therapeutically effective amount) to achieve complete remission. However, the quantity and quality of induced comprehensive CTLs may be insufficient, and therefore, one or more additional administrations (e.g., a second, third, etc.) may be required to achieve complete remission.

[0104] The doses of the CTLs and the proinflammatory cytokine-inhibitory antibody may be therapeutically effective. When the CTLs and the proinflammatory cytokine-inhibitory antibody are administered at intervals, it is preferable to administer one of them within 90 days of the other. The dose of the CTLs is, for example, 5 x 10 per administration. ８ pieces ~ 5×10 ９ The dose of CTLs can be adjusted appropriately depending on the size of the tumor, and it is preferable to increase the dose as the tumor size increases.

[0105] Step (5) may be performed only once or multiple times. If the subject does not achieve complete remission after performing step (5) once due to reasons such as the tumor size being too large, it is preferable to perform step (5) multiple times to achieve complete remission. When performing step (5) multiple times, it is preferable to repeatedly perform steps (3) and (4) to form CTLs in order to induce powerful comprehensive CTLs. In performing step (5) for the second or subsequent times, it is preferable to administer CTLs and an inflammatory cytokine inhibitory antibody to malignant tumors that have partially regressed, decreased, or eliminated and newly occurring metastases.

[0106] In the treatment method according to one aspect of the present invention, an anti-inflammatory agent other than an inflammatory cytokine-inhibitory antibody may be administered to the subject in at least one of steps (2) and (5).

[0107] Suitable anti-inflammatory agents may include those known in the art. The anti-inflammatory agent may be administered simultaneously with at least one of iDCs, a pro-inflammatory cytokine-inhibitory antibody, and CTLs, or may be administered at a predetermined interval from the administration thereof. Typically, the iDCs, the pro-inflammatory cytokine-inhibitory antibody, and the CTLs and the anti-inflammatory agent are administered simultaneously or substantially simultaneously, or the time lapse between the administrations is relatively short.

[0108] Examples of anti-inflammatory agents are the same as those described for anti-inflammatory agents in [1. Composition for treating solid malignant tumors]. Dexamethasone is preferably administered to the subject because it is expected to have not only an anti-inflammatory effect but also an appetite-stimulating effect and an antiemetic effect. Dexamethasone is the same as that described for dexamethasone in [1. Composition for treating solid malignant tumors].

[0109] Dexamethasone may be administered simultaneously with at least one of iDCs, an inflammatory cytokine-inhibitory antibody, and CTLs, or may be administered at a predetermined interval before or after the administration of at least one of iDCs, an inflammatory cytokine-inhibitory antibody, and CTLs. Dexamethasone may be combined with at least one of iDCs, an inflammatory cytokine-inhibitory antibody, and CTLs to form a composition, and the formed composition may be administered into a patient's tumor.

[0110] The dose of dexamethasone may be a therapeutically effective amount. Dexamethasone may be administered to a subject once or multiple times. When multiple doses are administered, it is preferable to administer the next dose 30 days after the previous dose. The dose of dexamethasone may be, for example, one-tenth to one-fourth of the prescribed systemic dose of dexamethasone.

[0111] [Summary] The composition for treating solid malignant tumors according to Aspect 1 of the present invention is used to be administered to a subject having malignant tumor cells that produce at least one inflammatory cytokine selected from tumor necrosis factor α, interleukin-1β, interleukin-5, interleukin-6, interleukin-8, interleukin-17, and interleukin-23, together with at least one type of immature dendritic cells and cytotoxic lymphocytes induced by dendritic cells, and contains at least one antibody that inhibits the action of the inflammatory cytokine.

[0112] A composition for treating solid malignant tumors according to Aspect 2 of the present invention is preferably the same as that according to Aspect 1, wherein the antibody is an antibody that inhibits the action of interleukin-6.

[0113] A composition for treating solid malignant tumors according to Aspect 3 of the present invention is preferably the same as that according to Aspect 1, wherein the antibodies are an antibody that inhibits the action of interleukin-5 and an antibody that inhibits the action of interleukin-6.

[0114] The composition for treating solid malignant tumors according to Aspect 4 of the present invention, in any one of Aspects 1 to 3, is preferably administered intraarterially, intravenously, intramuscularly, intraperitoneally, intratumorally, intrathoracically, or subcutaneously.

[0115] The composition for treating solid malignant tumors according to Aspect 5 of the present invention is preferably used in any one of Aspects 1 to 4 so as to be administered to the subject together with dexamethasone.

[0116] A kit for treating solid malignant tumors according to Aspect 6 of the present invention is used to be administered to a subject having malignant tumor cells that produce at least one inflammatory cytokine selected from tumor necrosis factor α, interleukin-1β, interleukin-5, interleukin-6, interleukin-8, interleukin-17, and interleukin-23, together with at least one type of immature dendritic cells and cytotoxic lymphocytes induced by dendritic cells, and the kit comprises at least one antibody that inhibits the action of the inflammatory cytokine.

[0117] A method for treating solid malignant tumors according to a seventh aspect of the present invention comprises the following steps (1) and at least one of (2) and (3) to (5): (1) screening for inflammatory cytokines produced by malignant tumor cells in a subject from the group consisting of tumor necrosis factor-α, interleukin-1β, interleukin-5, interleukin-6, interleukin-8, interleukin-17, and interleukin-23; (2) administering to the subject immature dendritic cells and at least one antibody that inhibits the action of the inflammatory cytokine screened in the step (1); (3) collecting peripheral blood mononuclear cells from the subject; (4) culturing the collected peripheral blood mononuclear cells to form dendritic cell-induced cytotoxic lymphocytes; and (5) administering to the subject the formed cytotoxic lymphocytes and at least one antibody that inhibits the action of the inflammatory cytokine screened in the step (1).

[0118] A method for treating a solid malignant tumor according to an eighth aspect of the present invention is preferably the method of seventh aspect, in which the steps (3) to (5) are carried out after the step (2).

[0119] In the method for treating a solid malignant tumor according to Aspect 9 of the present invention, in Aspect 1 or 2, it is preferable that dexamethasone is further administered to the subject in at least one of the steps (2) and (5).

[0120] A method for treating a solid malignant tumor according to Aspect 10 of the present invention is preferably administered into the malignant tumor of the subject in any one of Aspects 1 to 3.

[0121] A method for regressing, reducing, or eliminating tumor cells in tumor tissue of a patient according to aspect 11 of the present invention comprises the following steps (a) and (b): (a) administering therapeutically effective amounts of autologous immature dendritic cells, anti-interleukin-6 antibody, anti-interleukin-5 antibody, and dexamethasone intratumorally to the patient; and (b) following step (a), administering a therapeutically effective amount of autologous activated T cells intravenously to the patient.

[0122] The method according to aspect 12 of the present invention is preferably the same as in aspect 11, further comprising the following steps (c) to (e): (c) collecting peripheral blood mononuclear cells from the patient following steps (a) and (b); (d) culturing the collected peripheral blood mononuclear cells following step (c) to form induced comprehensive cytotoxic T lymphocytes; and (e) administering the induced comprehensive cytotoxic T lymphocytes, an anti-interleukin-6 antibody or an anti-interleukin-5 antibody, and dexamethasone intratumorally following step (d).

[0123] The method according to Aspect 13 of the present invention is the method according to Aspect 12, further comprising the following step (f): (f) It is preferable to repeat the step (e).

[0124] In the method according to Aspect 14 of the present invention, in any one of Aspects 11 to 13, tumor cells in the tumor tissue are preferably regressed, reduced, or eliminated without using radiation therapy.

[0125] A method according to Aspect 15 of the present invention is preferably the method according to any one of Aspects 11 to 13, wherein the patient is in remission after the treatment step.

[0126] In the method according to aspect 16 of the present invention, in aspect 12, it is preferable that steps (c), (d), and (e) are carried out if steps (a) and (b) do not result in complete remission of the patient.

[0127] In the method according to aspect 17 of the present invention, in aspect 12, steps (c), (d), and (e) are preferably performed on partially regressed tumor cells and / or newly developed metastases to achieve complete remission.

[0128] In the method according to aspect 18 of the present invention, in aspect 13, it is preferable that step (f) is carried out if steps (a), (b), (c), (d), and (e) do not result in complete remission of the patient.

[0129] In the method according to aspect 19 of the present invention, in aspect 13, step (f) is preferably performed on partially regressed tumor cells and / or newly developed metastases to achieve complete remission.

[0130] Aspect 20 of the present invention is a method according to aspect 11, wherein the administration of the anti-interleukin-6 antibody, the interleukin-5 antibody, and the dexamethasone is preferably carried out simultaneously with the administration of the autologous immature dendritic cells.

[0131] Aspect 21 of the present invention is preferably a method according to aspect 20, wherein the autologous immature dendritic cells, the anti-interleukin-6 antibody and the interleukin-5 antibody, and the dexamethasone form a composition, and the composition is administered into the tumor of the patient.

[0132] Aspect 22 of the present invention is the method of aspect 11, wherein step (b) is preferably carried out immediately after or a short time after administration in step (a).

[0133] Aspect 23 of the present invention is the method of aspect 22, wherein step (b) is preferably carried out about 24 to 72 hours after step (a).

[0134] Aspect 24 of the present invention is the method of aspect 12, wherein steps (c), (d), and (e) are preferably carried out within about 2 to 6 hours after steps (a) and (b).

[0135] Aspect 25 of the present invention is preferably the method of aspect 12, wherein an induction period for the cytotoxic T lymphocytes is provided between steps (b) and (c).

[0136] Aspect 26 of the present invention is a method according to aspect 12, wherein the cytotoxic T lymphocytes are preferably cultured for about 2 to 6 weeks during step (d).

[0137] Aspect 27 of the present invention is a method according to aspect 12, wherein the administration of the anti-interleukin-6 antibody, the anti-interleukin-5 antibody, and the dexamethasone is preferably carried out simultaneously with the administration of the induced cytotoxic T cells.

[0138] Aspect 28 of the present invention is preferably a method according to aspect 12, wherein the induced cytotoxic T lymphocytes, anti-interleukin-6 antibody, anti-interleukin-5 antibody, and dexamethasone form a composition, and the composition is administered into the tumor of the patient.

[0139] Aspect 29 of the present invention is preferably the method of aspect 12, wherein the CTLs are cultured in a culture medium selected from the group consisting of IL-2, CD3, and a mixture thereof.

[0140] In the method according to Aspect 30 of the present invention, in Aspect 11, the tumor cells are preferably present in metastatic tumor tissue.

[0141] In the method according to Aspect 31 of the present invention, in Aspect 11, the patient is preferably a human or non-human mammal.

[0142] Aspect 32 of the present invention is a method according to aspect 11, wherein the autologous immature dendritic cells are preferably administered in combination with an adjuvant.

[0143] Aspect 33 of the present invention is a method according to aspect 32, wherein the adjuvant is selected from the group consisting of lipid-based, protein-based, and polysaccharide-based adjuvants, and mixtures thereof.

[0144] A method according to Aspect 34 of the present invention is preferably the method according to Aspect 33, wherein the adjuvant is selected from the group consisting of lymphocyte culture medium, Marignase, Agaricus, OK432, BCG, Lentinan (Shiitake mushroom), Reishi mushroom, Polyporus polyporus, TNF Meshimakobu, incomplete or complete Freund's adjuvant, LPS, fatty acids, TW80, phospholipids, cytokines or viruses, and mixtures thereof.

[0145] Aspect 35 of the present invention relates to a method according to aspect 34, wherein the adjuvant preferably comprises leukocyte culture medium (LCM).

[0146] Aspect 36 of the present invention is a method according to aspect 11, wherein one or more of the autologous immature dendritic cells, the activated T cells, and the cytotoxic T lymphocytes are preferably obtained by apheresis from the patient.

[0147] Those skilled in the art will appreciate that changes could be made to the above-described embodiments without departing from the broad inventive concept thereof. It is understood, therefore, that the invention is not limited to the particular embodiments disclosed, but that it is intended to cover modifications within the spirit and scope of the invention as defined by the appended claims.

[0148] Reagents for which the manufacturer is not specified in the examples were those commonly used in the art. [Example 1] (Subject) Diagnosis: Right-sided progressive lung cancer and multiple bone metastases Pathology: Adenocarcinoma

[0149] (Method for preparing immature dendritic cells) Immature dendritic cells (iDCs) were obtained by the following method: Thawed monocyte nuclei (approximately 6 × 10 ８) was resuspended in 20 mL of AIM-V solution, and each was placed in a 44-T-75 cm ２ The cells were then distributed in 5 mL aliquots into polystyrene flasks. After 2 hours of incubation at 37°C, nonadherent cells were removed with a pipette, transferred to conical tubes, and stored for AT cell generation, as described below. 15 mL of DC growth solution (AIM-V solution (CellGenix, Germany) supplemented with 800 IU / mL GM-CSF + 500 mL IL4 (BD Pharmingen)) was added to each flask containing adherent cells. Flasks were incubated at 37°C in 5% carbon dioxide. The growth solution was refreshed on day 3, and DCs were harvested by pipetting on day 7. The harvested cells were counted and resuspended in AIM-V freezing solution containing 20% ​​autologous serum + 10% DMSO and frozen in BICELL containers (Nihon Freezer Co., Tokyo, Japan). The BICELL containers allow for stepwise cell freezing using a programmed freezing process (freezing rate of 1°C / min). Cells were stored at −80° C. until injection into patients (0.5 to 3 months).

[0150] (Preparation method of activated T cells) Activated T cells (AT) were prepared by the following method: non-adherent T cells (approximately 6-9 × 10 ) were harvested after monocyte adherence for DC generation. ８ The AT cell suspension (5 mL) and AT cell solution (35 mL) were washed and resuspended in 20 mL of AIM-V solution. Five mL of this cell suspension and 35 mL of the AT cell solution were each transferred to four T-225cm plates coated with anti-CD3 antibody. ２The flasks were then incubated at 37°C in a 5% CO atmosphere. Three hours before harvesting, 1 μg / mL ionomycin (Sigma, USA) was added to stimulate the T cells (Sato, T. et al., Cancer Immunol Immunother, 53:53-61, 2004). The AT cell solution consisted of AIM-V solution supplemented with IL-2 and autologous serum, resulting in a final concentration of 1000 IU / mL and 10% autologous serum in each flask. Anti-CD3 antibody coating was performed by adding 10 mL of 5 μg / mL anti-CD3 antibody (Orthoclone, OKT3 injection, Janssen Pharmaceutical, KK) in DPBS to the flasks and leaving them at room temperature for 2 hours. The flasks were then washed three times with 15 mL of DPBS before adding the cells. Harvested cells were cryopreserved and stored at −80° C. (0.5-3 months) before injection into patients.

[0151] (Treatment Method) First, before the start of treatment, the blood levels of inflammatory cytokines in the subjects were measured using an ELISA assay.

[0152] Next, the prepared autologous immature dendritic cells were added to 1 × 10 ７ The subjects received four puncture injections into their primary tumors at a dose of 1 x 10 per tumor (dates of administration: 1st injection: April 5, 2021, 2nd injection: April 13, 2021, 3rd injection: May 19, 2021, 4th injection: May 20). Immediately after the first injection (24 to 48 hours later), AT1 x 10 ８ One tumor per subject was administered intravenously.

[0153] Blood samples were then taken and blood concentrations were measured, including white blood cells (WBC), hemoglobin (Hb), platelets (plate), total protein (TP), serum lactate dehydrogenase (LDH), and alanine transferase (ALT). Blood inflammatory cytokine concentrations were also measured.

[0154] Next, based on the measurement results of the blood inflammatory cytokine concentrations, the therapeutic composition of Example 1 was prepared by the following method: 80 mg / 4 mL of tocilizumab (manufactured by Chugai Pharmaceutical Co., Ltd.), an anti-IL-6 receptor antibody, was used as the therapeutic composition.

[0155] Before administration of the therapeutic composition, a computed tomography (CT) scan was performed to obtain CT images of the lungs. On June 9, 2021, 21 days after administration of the immature dendritic cells, 80 mg of tocilizumab (liquid volume 4 ml) was administered as the therapeutic composition into the subject's primary tumor by puncture.

[0156] After administration of the therapeutic composition, CT scans and blood component measurements were again performed.

[0157] (Results) The results of measuring the concentrations of inflammatory cytokines in the blood before and after the administration of immature dendritic cells are shown in Table 1. The results shown in Table 1 confirmed that IL-6 and IL-8 increased after the administration of immature dendritic cells. In the subject of Example 1, no tumor shrinkage was observed after the administration of autologous immature dendritic cells, and the tumor rapidly grew, suggesting the possibility that these inflammatory cytokines are involved in the growth of the tumor.

[0158] The results of the treatment are shown in Figure 4. In Figure 4, the left image is a CT image taken after iDC administration but before administration of the therapeutic composition, and the right image is a CT image taken after administration of the therapeutic composition. In Figure 4, the treated solid malignant tumor is circled, and the cross inside the circle indicates the tumor diameter. As shown in Figure 4, the administration of an anti-IL-6 receptor antibody as the therapeutic composition reduced the size of the solid malignant lung tumor. These results confirmed that inflammatory cytokines are involved in tumor growth after administration of autologous immature dendritic cells, and that inhibiting the action of inflammatory cytokines by administering an inflammatory cytokine-inhibiting antibody can result in tumor reduction.

[0159] The amounts of components in the blood are shown in Table 2. As shown in Table 2, no changes in the amounts of components that would indicate side effects were observed before and after administration of the therapeutic composition. Therefore, no side effects due to administration of the therapeutic composition were confirmed.

[0160] [Example 2] (Subject) Diagnosis: Multiple abdominal lymph node metastases due to recurrent rectal cancer Pathology: Adenocarcinoma

[0161] (Treatment Method) Autologous immature dendritic cells obtained by the same method as in Example 1 were administered at a dose of 1 × 10 ７ The subject received two doses of the drug via puncture into the right pelvic lymph node at a dose of 1 / 1 tumor (first dose: December 20, 2021, second dose: December 21, 2021).

[0162] Inflammatory cytokines were screened in solid malignant tumors of subjects after administration of immature dendritic cells. Cells were collected from rectal malignant tumors, and the presence or absence of intracellular protein expression of TNFα, IL-1β, and IL-5ra was confirmed using antibodies that specifically recognize TNFα, IL-1β, IL-5 receptor α (IL-5ra), IL-6, IL-8, IL-17 receptor α (IL-17ra), and IL-23 receptor α (IL-23ra). Regarding IL-5, IL-17, and IL-23, the expression of their receptors, rather than the cytokines themselves, was confirmed. Cells in which receptor expression was confirmed can be said to also express cytokines, which are ligands for those receptors. After administration of immature dendritic cells, CT scans were performed to obtain abdominal CT images. Blood samples were also collected to measure the concentrations of blood components. The same components were measured as in Example 1, except that aspartate aminotransferase (AST) was measured instead of LDH.

[0163] Next, based on the results of the screening, a therapeutic composition of Example 2 was prepared. That is, mepolizumab, a humanized anti-IL-5 receptor antibody, was used as the therapeutic composition.

[0164] On April 26, 2022, 126 days after the administration of the immature dendritic cells, 25 mg of mepolizumab was administered as a therapeutic composition to the subject's right pelvic lymph node by puncture.

[0165] After administration of the therapeutic composition, CT scans and blood component measurements were again performed.

[0166] (Results) The results of the screening are shown in Table 3. A bar (-) in the table indicates that no protein expression was observed. The results of the screening revealed that the tumor cells were producing TNFα, IL-1β, and IL-5.

[0167] The results of the treatment are shown in Figure 5. In Figure 5, the left image is a CT image taken after iDC administration but before administration of the therapeutic composition, and the right image is a CT image taken after administration of the therapeutic composition. In Figure 5, the solid malignant tumor that was treated is circled, and the cross inside the circle indicates the tumor diameter. As shown in Figure 5, the solid malignant tumor in the right pelvic lymph node shrank as a result of the treatment. These results confirmed that a sufficient tumor shrinkage effect could be achieved by administering only a humanized anti-IL-5 receptor antibody as a therapeutic composition.

[0168] The amounts of components in the blood are shown in Table 2. As shown in Table 4, no change in the amounts of components that cause side effects was observed before and after administration of the therapeutic composition. Therefore, no side effects due to administration of the therapeutic composition were confirmed.

[0169] In this test example, the treated malignant solid tumor achieved complete remission (CR) 7 months after the start of treatment. In Figure 9, the left image shows a PET image of the subject's upper body before the start of treatment, and the right image shows a PET image of the subject's upper body 7 months after the start of treatment. As shown in Figure 9, the treated malignant solid tumors (the areas indicated by the arrows in the left image, i.e., malignant solid tumors in six para-aortic lymph nodes, two aortic lymph nodes, four left iliac lymph nodes, and two right iliac lymph nodes) were confirmed to have disappeared 7 months after the start of treatment. The arrows in the right image indicate the rectal anastomosis and physiological uptake of fluorodeoxyglucose (FDG) in urine.

[0170] [Example 3] (Subject) Diagnosis: Left lung cancer with multiple mediastinal lymph node metastases Pathology: Adenocarcinoma

[0171] (Treatment Method) Inflammatory cytokines in solid malignant tumors of subjects were screened before administration of the therapeutic composition by the same method as in Example 2, except that antibodies specifically recognizing TNFα, IL-1β, IL-6, and IL-8, respectively, were used. In addition, CT scans were performed before administration of the therapeutic composition to obtain CT images of the mediastinal lymph nodes and aortic lymph nodes. Blood samples were also collected before administration of the therapeutic composition, and WBC, Hb, and plate counts were measured.

[0172] Next, based on the results of the screening, a therapeutic composition for Example 3 was prepared. Specifically, 20 mg / mL of tocilizumab (manufactured by Chugai Pharmaceutical Co., Ltd.), an anti-IL-6 receptor antibody, was used as the therapeutic composition. Immature dendritic cells were prepared in the same manner as in Example 1.

[0173] Autologous immature dendritic cells 1 x 10 ７ The tumors and 20 mg of tocilizumab (liquid volume 1 ml) as a therapeutic composition were simultaneously administered by puncture into the mediastinal lymph nodes of the subjects (date of administration: March 8, 2022).

[0174] After administration of the therapeutic composition, a CT scan and blood sampling were again performed to measure the concentrations of the components in the blood.

[0175] (Results) The results of the screening are shown in Table 5. In the table, a bar (-) indicates that no protein expression was observed, and a blank cell indicates that the protein was not measured. The results of the screening revealed that the tumor cells were producing TNFα, IL-1β, and IL-6.

[0176] The results of the treatment are shown in Figure 6. In Figure 6, the upper row shows CT images taken before administration of immature dendritic cells and the therapeutic composition, and the lower row shows CT images taken after administration of immature dendritic cells and the therapeutic composition. In Figure 6, the treated solid malignant tumors are circled, and the cross within the circle indicates the tumor diameter. As shown in Figure 6, the treatment resulted in the reduction of the size of two solid malignant tumors in the mediastinal lymph nodes and the solid malignant tumor in the aortic lymph node.

[0177] The amounts of components in the blood are shown in Table 6. As shown in Table 6, no change in the amounts of components that cause side effects was observed before and after administration of immature dendritic cells and the therapeutic composition. Therefore, no side effects due to administration of the therapeutic composition were confirmed.

[0178] [Example 4] (Subject) Diagnosis: Recurrent cervical cancer and multiple pelvic lymph node metastases Pathology:

[0179] (Treatment Method) 1 x 10 autologous immature dendritic cells prepared in the same manner as in Example 1 were used. ７ Each tumor was administered into five sites in the right pelvic lymph nodes of each subject.

[0180] Four weeks after the administration of autologous immature dendritic cells, blood was collected from the subjects and peripheral blood mononuclear cells were isolated. Subsequently, the peripheral blood mononuclear cells were cultured in RPMI medium under negative selection conditions in the presence of CD3 and IL-2 to obtain cytotoxic lymphocytes.

[0181] Next, inflammatory cytokines in the solid malignant tumors of the subjects were screened before administration of the therapeutic composition using the same method as in Example 2. Also, CT scans were performed before administration of the therapeutic composition to obtain CT images of the pelvic lymph nodes. Blood samples were also collected, and WBC, Hb, platelet count, TP, AST, and ALT were measured.

[0182] Based on the screening results, a therapeutic composition of Example 4 was prepared. That is, mepolizumab, the same as in Example 2, was used as the anti-IL-5 receptor antibody, and tocilizumab, the same as in Example 1, was used as the anti-IL-6 receptor antibody.

[0183] Dexamethasone 2 mg, therapeutic composition (mepolitumab 10 mg, tocilizumab 5 mg), and cytotoxic lymphocytes 5 × 10 ８ The individual doses were administered simultaneously via puncture into five sites in the right pelvic lymph nodes of each subject (date of administration: June 7, 2022).

[0184] After administration of the therapeutic composition, blood samples were taken again to measure the concentrations of the components in the blood, and MRI images of the pelvic lymph nodes were obtained using magnetic resonance imaging (MRI).

[0185] (Results) The results of the screening are shown in Table 7. A bar (-) in the table indicates that no protein expression was observed. The results of the screening revealed that the tumor cells were producing TNFα, IL-1β, IL-5, IL-6, IL-17, and IL-23.

[0186] The results of the treatment are shown in Figure 7. In Figure 7, the left image is a CT image taken after iDC administration and before administration of dexamethasone, CTLs, and the therapeutic composition, and the right image is an MRI image taken after administration of dexamethasone, CTLs, and the therapeutic composition. In Figure 7, the treated solid malignant tumor is circled, and the cross within the circle indicates the tumor diameter. As shown in Figure 7, the treatment reduced the size of the solid malignant tumor in the pelvic lymph node.

[0187] The amounts of components in the blood are shown in Table 2. As shown in Table 8, no amounts of components that would cause side effects were observed after administration of the therapeutic composition. Therefore, no side effects due to administration of the therapeutic composition were confirmed.

[0188] [Example 5] (Subject) Diagnosis: pancreatic cancer (inoperable) Pathology: adenocarcinoma

[0189] (Treatment Method) A therapeutic composition was prepared. Because a method for testing inflammatory cytokines produced by the tumor of the subject in this example could not be established before the start of treatment, screening of inflammatory cytokines produced by tumor cells could not be performed. However, because IL-5 and IL-6 were predicted to be expressed, mepolizumab, the same as in Example 2, was used as the anti-IL-5 receptor antibody, and tocilizumab, the same as in Example 1, was used as the anti-IL-6 receptor antibody, in the therapeutic composition of Example 5. In addition, immature dendritic cells and AT were prepared by the same method as in Example 1.

[0190] Three independent primary pancreatic tumors of each subject were treated with 1 x 10 autologous immature dendritic cells. ７ The tumors were then punctured and administered simultaneously with 220 mg of tocilizumab as a therapeutic composition. ８Each tumor was administered intravenously to the subject. In Example 5, these administrations are referred to as the "initial administration." Six months after the initial administration, blood was collected from the subject, and peripheral blood mononuclear cells were isolated. The peripheral blood mononuclear cells were then cultured in RPMI medium under negative selection conditions in the presence of CD3 and IL-2 to obtain cytotoxic lymphocytes.

[0191] After the first administration, a CT scan was performed to obtain a CT image of the pancreas. Blood samples were also taken to measure WBC, Hb, plate, TP, AST, and ALT.

[0192] On June 29, 2022, seven months after the initial administration, 4 mg of dexamethasone, 20 mg of mepolizumab and 20 mg of tocilizumab as therapeutic compositions, and 1 x 10 immature dendritic cells were administered to each of the three tumors where the initial administration was performed. ７ cells / 1 tumor and 5 x 10 cytotoxic lymphocytes ８ Each tumor was administered by puncture at the same time. In Example 5, these administrations are referred to as "second administrations."

[0193] After the second administration, CT scans and blood samples were taken again to measure the concentrations of components in the blood. In addition, since a method for testing inflammatory cytokines produced by the tumors of the subjects in this example after the second administration was established, inflammatory cytokines in the solid malignant tumors of the subjects after the second administration were screened using the same method as in Example 2 to confirm the inflammatory cytokines produced by tumor cells.

[0194] (Results) The results of the screening are shown in Table 9. A bar (-) in the table indicates that no protein expression was observed. The results of the screening revealed that the tumor cells were producing TNFα, IL-1β, IL-6, and IL-23.

[0195] The results of the treatment are shown in Figure 8. In Figure 8, the left image is a CT image taken after the first administration, and the right image is a CT image taken after the second administration. In Figure 8, the treated solid malignant tumor is circled, and the cross inside the circle indicates the tumor diameter. As shown in Figure 8, the treatment resulted in the reduction of the size of the pancreatic solid malignant tumor.

[0196] The amounts of components in the blood are shown in Table 10. As shown in Table 10, no amounts of components that would cause side effects were observed after administration of the therapeutic composition. Therefore, no side effects due to administration of the therapeutic composition were confirmed.

[0197] From the above, it has become clear that the therapeutic composition and therapeutic method according to one embodiment of the present invention can reduce the size of solid malignant tumors by suppressing inflammation within the tumors.

[0198] The present invention can be used in the treatment of solid malignant tumors.

Claims

1. The method is used to administer to a subject having solid malignant tumor cells that produce the following inflammatory cytokines (i) and (ii) in combination with at least one of immature dendritic cells and cytotoxic lymphocytes induced by dendritic cells: (i) at least one of interleukin-5 and interleukin-6; (ii) at least one of tumor necrosis factor α, interleukin-1β, interleukin-8, interleukin-17, and interleukin-23; A composition for treating solid malignant tumors, comprising an antibody that inhibits the action of interleukin-5.

2. A composition for treating solid malignant tumors as described in claim 1, further comprising an antibody that inhibits the action of interleukin-6.

3. A composition for treating solid malignant tumors described in claim 1 or 2, wherein the solid malignant tumor is adenocarcinoma.

4. The composition for treating solid malignant tumors according to claim 1 or 2, which is administered intraarterially, intravenously, intramuscularly, intraperitoneally, intratumorally, intrathoracically, or subcutaneously.

5. The composition for treating solid malignant tumors according to claim 1 or 2, which is administered to the subject in combination with dexamethasone.

6. The method is used to administer to a subject having solid malignant tumor cells that produce the following inflammatory cytokines (i) and (ii) in combination with at least one of immature dendritic cells and cytotoxic lymphocytes induced by dendritic cells: (i) at least one of interleukin-5 and interleukin-6; (ii) at least one of tumor necrosis factor α, interleukin-1β, interleukin-8, interleukin-17, and interleukin-23; A kit for treating solid malignant tumors, comprising an antibody that inhibits the action of interleukin-5.