Medium composition for producing cancer organoids

Abstract

The present invention relates to a medium composition for producing cancer organoids, comprising a translationally controlled tumor protein (TCTP). In addition, the present invention relates to a medium composition for promoting production of cancer organoids, a medium additive for preparing cancer organoids, a medium additive for promoting production of cancer organoids, a method for producing cancer organoids with a medium comprising the medium composition or the medium additive, cancer organoids produced by the method, a method for evaluating efficacy or toxicity of a cancer therapeutic substance by using the cancer organoids, and a screening method. In the production of cancer organoids, using the medium composition for producing cancer organoids of the present invention, a larger amount of cancer organoids can be obtained in a shorter period of time as compared to using a medium composition comprising R-spondin, and thus the medium composition for producing cancer organoids of the present invention can be used economically for mass production of cancer organoids. Therefore, in the production of a cancer organoid, the evaluation of efficacy or toxicity and screening using the cancer organoid, and the like, the medium composition for producing cancer organoids of the present invention may be used as a medium that is available at a more competitive price.

Description

Culture medium composition for the production of cancer organoids

[0001] This application claims the benefit of priority based on Korean Patent Application No. 10-2025-0033422 filed on March 14, 2025, and all contents disclosed in the document of said Korean patent application are incorporated herein as part of this specification.

[0002] The present invention relates to a culture medium composition for producing cancer organoids comprising TCTP (translationally controlled tumor protein). Furthermore, the present invention relates to a culture medium composition for promoting the production of cancer organoids, a culture medium additive for producing cancer organoids, a culture medium additive for promoting the production of cancer organoids, a method for producing cancer organoids using a culture medium comprising said culture medium composition or culture medium additive, cancer organoids produced by said method, and a method for evaluating the efficacy or toxicity of a cancer therapeutic substance using said cancer organoids and a screening method.

[0003] Due to their unique multipotency and self-renewal characteristics, research is continuously being conducted to utilize stem cells in various ways, including the treatment of incurable diseases, disease modeling, tissue or organ transplantation, and drug efficacy evaluation. In particular, it has been discovered that when stem cells are cultured in an appropriate three-dimensional in vitro environment, cell masses with structures and tissue characteristics similar to in vivo organs are formed; such cell masses are named organoids. Since organoid fabrication technology can theoretically produce almost any type of organ using only stem cells, it is expected to be utilized for evaluating the efficacy or screening of various therapeutic agents, as well as for transplantation into tissues or organs. Organoids may be more effective than two-dimensional cell tissues for testing the safety and efficacy of new drugs, and are believed to be useful for improving the condition of damaged or underdeveloped organs by transplantation. Accordingly, organoid culture, designed to mimic the human body environment, is a crucial technology in cancer research and is considered a highly useful research model for studying the development, growth, metastasis, and treatment of cancer. Cancer organoids are produced by culturing cancer cells isolated from patient-derived tissues or bodily fluids, and it is known that the success rate of culture varies depending on the components used. One of the components primarily used in conventional organoid culture is R-spondin, a representative substance of Wnt signaling regulators.

[0004] Recently, the industry has been conducting various studies to find components that can replace R-spondin. In this regard, substances used in organoid culture to replace R-spondin include, for example, RS-246204 disclosed in Registered Patent Publication No. 10-1966523 and escin disclosed in Published Patent Publication No. 10-2024-0041595.

[0005] Accordingly, the inventors conducted various studies to find a substance that can be used to efficiently produce cancer organoids of homogeneous quality by replacing R-spondin. As a result, they identified a culture medium component capable of efficiently proliferating cancer organoids and completed the present invention by experimentally proving its effects.

[0006] Each description and embodiment disclosed in the present invention may be applied to each other description and embodiment. That is, all combinations of the various elements disclosed in the present invention fall within the scope of the present invention. Furthermore, the scope of the present invention should not be considered limited by the specific descriptions provided below.

[0007] Furthermore, terms not specifically defined in this specification should be understood to have the meanings commonly used in the technical field to which the present invention pertains. Additionally, unless specifically defined in the context, the singular includes the plural, and the plural includes the singular.

[0008]

[0009] One aspect of the present invention provides a culture medium composition for producing cancer organoids, comprising TCTP (translationally controlled tumor protein).

[0010] The culture medium composition of the present invention can promote the proliferation of cancer organoids to form a large number of cancer organoids. Accordingly, the culture medium composition of the present invention can be usefully utilized in the production of cancer organoids applicable in various fields, such as drug efficacy evaluation or drug screening.

[0011] As used herein, the term "TCTP (translationally controlled tumor protein)" refers to a protein widely present in various eukaryotic species. The amino acid sequence of TCTP and the base sequence encoding it are known in existing databases. For example, it may be NCBI Reference Sequence: NP_003286.1, NM_003295.4 or GenBank: CUK27657.1, etc. The source of the TCTP is not particularly limited. For example, it may be prepared using commercially available materials or by using gene (protein) recombination technology according to methods known in the art. In the art, the term "TCTP (translationally controlled tumor protein)" may also be referred to as "fortilin," "histamine releasing factor (HRF)," "p02," "P23," or "Q23."

[0012] As used in this specification, the term "organoid" refers to a mass of cells having a three-dimensional structure. It is defined as a scaled-down and simplified version of an organ produced through an artificial culture process without being collected, acquired, or harvested from animals or the like. Organoids have the advantages of being capable of long-term culture and cryopreservation, as well as being easy to manipulate and observe. At the same time, they are experimental models that allow for the study of physiological phenomena at a higher level than cells by reproducing the hierarchical and histological structures of cells that could only be observed in vivo, as they do not require immortalization and thus maintain the original characteristics of the cells.

[0013] As used in this specification, the term "cancer organoid" refers to an organoid having the same or similar structure, cellular composition, and function as cancer in vivo, and may be produced by culturing cells derived from cancer tissue or cancer cells. The term "cancer organoid" may be used interchangeably with the terms "tumor organoid" or "tumoroid."

[0014] The above cancers may include, but are not limited to, liver cancer, lung cancer, non-small cell lung cancer, stomach cancer, colorectal cancer, pancreatic cancer, biliary tract cancer, colon cancer, small intestine cancer, brain cancer, bone cancer, melanoma, breast cancer, sclerosing adenoma, uterine cancer, endometrial cancer, cervical cancer, vaginal cancer, vulvar cancer, head cancer, cervical cancer, esophageal cancer, thyroid cancer, parathyroid cancer, endocrine gland cancer, kidney cancer, sarcoma, prostate cancer, urethral cancer, penile cancer, bladder cancer, ureteral cancer, hematological cancer, central nervous system tumor, spinal cord tumor, glioblastoma, brainstem glioma, pituitary adenoma, leukemia, lymphoma, fibroadenoma, etc.

[0015] As used in this specification, the term "manufacturing" may be used interchangeably with the terms "culture" or "proliferation." Accordingly, the medium composition for manufacturing cancer organoids may be a medium composition for culturing cancer organoids, may also be a medium composition for forming cancer organoids, and may also be a medium composition for expanding cancer organoids. In this case, "forming" means forming cancer organoids from cancer-derived cells and means exhibiting organ or tissue characteristics to perform the inherent functions of cancer. Furthermore, "expanding" means increasing the number of formed cancer organoids to obtain a sufficient amount for use in the intended purpose.

[0016] In the present invention, the cancer organoid may be cultured for 0 or more passages, 3 or more passages, 5 or more passages, or 10 or more passages.

[0017] As used herein, the term "passage" refers to replacing the culture vessel or dividing the cell group for culture in a method of continuously culturing generations of cells to keep cells or organoids in a healthy state for a long period. As the number of passages increases, a larger number of organoids can be obtained. One passage is replacing the culture vessel once or dividing the cell group once, and twenty passages is replacing the culture vessel twenty times or dividing the cell group twenty times. The first passage may be performed for 1 to 15 days. In this specification, the term "passage" may be used interchangeably with the term "generation."

[0018] As used herein, the term "medium" refers to a mixture of nutrient substances that enables the growth, survival, expansion, or differentiation of cells or organoids in vitro, and includes all appropriate conventional media used in the art. Depending on the type of cell or organoid, the type of medium and culture conditions may be selected at the technical level of the art.

[0019] The content of TCTP in the culture medium composition can be easily selected at a technical level in the art depending on the type of cancer, cell line, or organoid. For example, TCTP may be 0.001 to 100 ng / mL, 0.01 to 50 ng / mL, or 0.1 to 20 ng / mL, but is not limited thereto.

[0020] The culture medium composition for producing cancer organoids according to the present invention may additionally include basal media.

[0021] As used herein, the term “basal media” may be a cell culture medium containing a carbon source, a nitrogen source, and trace elements. Specific examples may include, but are not limited to, DMEM (dulbecco's modified eagle's medium), MEM (minimal essential medium), BME (basal medium eagle), RPMI1640, F-10, F-12, α-MEM (α-Minimal essential Medium), GMEM (glasgow's minimal essential medium), IMDM (iscove's modified dulbecco's medium), McCoy's 5A, Neurobasal medium, DMEM / F12 (dulbecco's modified eagle medium / nutrient mixture F-12), or advanced DMEM / F12.

[0022] Organoids have typically been prepared using two types of culture media. For example, an organoid formation medium is used to form and grow organoids from cells or tissues collected or isolated from a subject, and then, after removing the formation medium, an organoid expansion medium is used to increase the number of organoids. When the organoid formation medium and the expansion medium are composed of different components, the procedure has the disadvantage of being cumbersome and complex, and requiring a lot of cost and time.

[0023] Furthermore, organoids are characterized by the fact that the results of their production vary significantly depending on the culture conditions. As examples of culture conditions, factors such as the culture period, culture temperature, culture medium, and culture substrate are important factors determining the characteristics of organoids; among these, the culture medium condition plays the most significant role in growing organoids to possess characteristics similar to the desired tissue or organ by regulating various signaling pathways in stem cells. Since organoids cultured under different culture medium conditions exhibit very different characteristics in terms of similarity to actual tissue or viability, the present invention establishes optimal culture medium conditions to obtain a sufficient number of cancer organoids that are morphologically and functionally very similar to actual tissues and have increased proliferative capacity, making them suitable for use in evaluating the efficacy or toxicity of anticancer drugs or anticancer drug candidates, or for screening. Furthermore, it has been confirmed that when using the culture medium of the present invention, the formation, growth, or expansion of cells or organoids can be successfully induced, or the rate thereof can be increased, even when using only one type of culture medium, without using two or more types of culture media.

[0024] The culture medium composition for preparing cancer organoids according to the present invention may additionally include components that are commonly used in the art for cell or organoid culture media, such as antibiotics, antioxidants, FBS (fetal bovine serum), GlutaMAX, N2, 2-mercaptoethanol (β-mercaptoethanol), and B27. The antibiotic may be, for example, penicillin, streptomycin, gentamicin, primocine, or normosin, but is not limited thereto. The antioxidant may be, for example, nicotinamide, 1-thioglycerol, valproic acid, or N-acetyl-L-cysteine, but is not limited thereto.

[0025]

[0026] Another aspect of the present invention provides a culture medium composition for promoting the production of cancer organoids, comprising TCTP.

[0027] In the culture medium composition for promoting the production of cancer organoids according to the present invention, each term has the same meaning as described in the use of the culture medium composition for producing cancer organoids, unless specifically stated otherwise.

[0028] The above-mentioned culture medium composition for promoting cancer organoid production may be a culture medium composition for increasing the rate of cancer organoid production, and may be a culture medium composition for promoting cancer organoid formation, growth, or expansion.

[0029] The promotion of cancer organoid production described above may include not only the expansion or increase in the number of cells or cancer organoids, but also an increase in the initial rate of formation, growth, or expansion of the cancer organoids. When cancer organoids are produced using the culture medium composition of the present invention, more cancer organoids can be obtained in a short period of time, thereby enabling the cancer organoids to be utilized more rapidly for evaluating the efficacy of anticancer drugs or for screening anticancer drugs or anticancer drug candidates.

[0030]

[0031] Another aspect of the present invention provides a culture medium additive for producing cancer organoids comprising TCTP.

[0032] In the culture medium additive for producing cancer organoids according to the present invention, each term has the same meaning as described in the culture medium composition for promoting cancer organoid production, unless specifically stated otherwise.

[0033] The culture medium additive of the present invention can successfully induce the formation, growth, or expansion of cells or organoids, or increase the rate thereof. Accordingly, the culture medium additive of the present invention may be used in any basic culture medium as long as its efficacy is not offset or lost. For example, it may be used by adding it to a conventional or novel basic culture medium for the production of cancer organoids, or by adding it to a culture medium composition customized for a specific type of cancer organoid, but is not limited thereto.

[0034] The culture medium additive for producing cancer organoids according to the present invention may additionally include components that are commonly used in the industry for cell or organoid culture media, such as antibiotics, antioxidants, FBS, GlutaMAX, N2, 2-mercaptoethanol, and B27.

[0035]

[0036] Another aspect of the present invention provides a medium additive for promoting the production of cancer organoids, comprising TCTP.

[0037] In the medium additive for promoting the production of cancer organoids according to the present invention, each term has the same meaning as described in the medium additive for producing cancer organoids unless specifically stated otherwise.

[0038]

[0039] Another aspect of the present invention provides a method for producing a cancer organoid, comprising the step of culturing cancer cells isolated from an individual in a medium comprising the medium composition of the present invention or the medium additive of the present invention.

[0040] In the method for producing a cancer organoid according to the present invention, each term has the same meaning as described in the medium additive for promoting cancer organoid production, unless specifically stated otherwise.

[0041] As used herein, the term "individual" includes all individuals that have never developed cancer, individuals that are likely to develop cancer, individuals that have developed cancer, or individuals that have been cured after developing cancer, and may include humans or any non-human animals without limitation. The non-human animals may be vertebrates, such as primates, dogs, cattle, horses, pigs, rodents, such as mice, rats, hamsters, guinea pigs, etc. In this specification, the term "individual" may be used interchangeably with the terms "subject" or "patient."

[0042] The cancer cells isolated from the above organism may include stem cells. Additionally, the isolated cancer cells may be isolated through a process of cutting the cancer tissue and an enzymatic degradation process. Specifically, the cutting may include both physical cutting and mechanical cutting, and may be performed using general tissue cutting methods known in the art. Additionally, the enzymatic degradation may be performed under general enzymatic degradation conditions known in the art, and may be performed, for example, using one or more enzymes selected from the group consisting of dispase II, DNA degrading enzyme (DNase) I, and collagenase II.

[0043] In addition, culture conditions such as the culture period, culture temperature, and culture substrate can be performed under conditions commonly used in the industry for the culture of organoids, and a person skilled in the art will be able to use a method suitable for the purpose of the present invention.

[0044]

[0045] Another aspect of the present invention provides a cancer organoid produced by the cancer organoid production method of the present invention.

[0046] In the cancer organoid according to the present invention, each term has the same meaning as described in the cancer organoid manufacturing method above, unless specifically stated otherwise.

[0047] The cancer organoid according to the present invention can reproduce the microenvironment of an actual tumor and thus possesses histological composition and function very similar to human cancer, making it useful for evaluating the efficacy (drug efficacy evaluation), toxicity assessment, or screening of anticancer drugs. In particular, when the cancer organoid is manufactured using patient-derived cells or tissues, it has the advantage of maintaining the patient's unique genotype and phenotype well; therefore, it can be used to identify patient-specific anticancer drug resistance, tolerance, or refractoryness in advance. Since most cancer patients have a short remaining lifespan, it is necessary to find and receive treatment with the most effective anticancer drug for them as soon as possible. To this end, personalized medicine and precision medicine have recently been developed, and the cancer organoid derived from a specific patient's cancer tissue according to the present invention can be very usefully utilized in such personalized medicine and precision medicine. Furthermore, by using the cancer organoid to observe gene expression regulation following drug treatment, it can contribute to research on molecular mechanisms for the purpose of cancer treatment.

[0048] As used herein, the term "anticancer agent" refers to a substance that exhibits an effect of treating, improving, or preventing cancer. Specifically, any substance that inhibits or improves the occurrence, growth, migration, or metastasis of cancer cells or tumors, or kills or increases the killing of cancer cells or tumors, may be used without limitation. It includes all substances capable of treating, improving, or preventing cancer or tumors, such as compounds, nucleic acids, proteins, compound-protein complexes, drug-protein complexes, antibodies, compound-antibody complexes, drug-antibody complexes, amino acids, peptides, viruses, carbohydrates, lipids, extracts, and fractions.

[0049] As used in this specification, the term "anticancer drug resistance" means that an individual resists the effects of an anticancer drug, resulting in the drug being ineffective from the beginning of treatment.

[0050] As used herein, the term "anticancer drug resistance" means that while an anticancer drug is effective during the initial stages of treatment, a larger amount of the drug is required to achieve the same effect as the individual becomes accustomed to the drug through regular use.

[0051] As used herein, the term "anticancer drug refractory" means that, despite a sufficient number of cycles of anticancer drug treatment, cancer or a tumor remains unresponsive to the effects of the anticancer drug to the desired degree.

[0052]

[0053] Another aspect of the present invention provides a method for evaluating the efficacy or toxicity of an anticancer agent, comprising the steps of: producing a cancer organoid by culturing cancer cells isolated from an individual in a culture medium composition for producing a cancer organoid according to the present invention; and treating the cancer organoid with an anticancer agent or a candidate anticancer agent.

[0054] In the method for evaluating the efficacy or toxicity of an anticancer agent according to the present invention, each term has the same meaning as described in the above cancer organoids unless specifically stated otherwise.

[0055] As used herein, the term "anticancer drug candidate" refers to a substance expected to be able to treat, improve, or prevent cancer. Specifically, any substance expected to inhibit or improve the occurrence, growth, migration, or metastasis of cancer cells or tumors, or to increase the death of cancer cells, may be used without limitation and includes all substances expected to treat, improve, or prevent cancer or tumors, such as compounds, nucleic acids, proteins, compound-protein complexes, drug-protein complexes, antibodies, compound-antibody complexes, drug-antibody complexes, amino acids, peptides, viruses, carbohydrates, lipids, extracts, fractions, etc.

[0056] As used in this specification, the term "treatment" may be used interchangeably with the term "administration."

[0057] The treatment of the above anticancer agent or anticancer agent candidate may be performed using methods known in the art. As a specific example, the above anticancer agent or anticancer agent candidate may be treated with the above cancer organoid and co-cultured, but is not limited thereto, and a person skilled in the art may use a method suitable for the purpose of the present invention.

[0058] The method for evaluating efficacy or toxicity of the present invention may further include the step of analyzing the anticancer activity of a cancer organoid treated with an anticancer agent or an anticancer agent candidate.

[0059] The step of analyzing the above anticancer activity may be a step of analyzing the inhibition of growth, increased death, reduction in size, reduction in area, reduction in weight, etc. of cancer organoids. Any method known to those skilled in the art may be used for the above analysis. Specific examples include Western blot, Co-Immunoprecipitation assay, ELISA (Enzyme Linked Immunosorbent Assay), tissue immunostaining, FACS (Fluorescence activated cell sorter), tissue biopsy analysis, HCS (High-Content Screening), and flow cytometry, but are not limited thereto, and those skilled in the art may use a method suitable for the purpose of the present invention.

[0060] In addition, the method for evaluating efficacy or toxicity of the present invention may further include a step of comparing the anticancer activity of a cancer organoid treated with an anticancer drug or anticancer drug candidate substance to be evaluated with the anticancer activity of a control group.

[0061] In addition, the method for evaluating efficacy or toxicity according to the present invention may further include a step of determining the efficacy or toxicity of an anticancer drug or an anticancer drug candidate. The determination of efficacy may, for example, determine that the efficacy of the anticancer drug or anticancer drug candidate is superior if the anticancer activity in the cancer organoid treated with the anticancer drug or anticancer drug candidate increases compared to a negative control, or if the anticancer activity increases or is similar compared to a positive control. Furthermore, the determination of toxicity may determine the level or degree of toxicity of the treated anticancer drug or anticancer drug candidate based on the results of comparing the activity of the cancer organoid and the control.

[0062] In the present invention, the control group may be a negative control group and / or a positive control group. The negative control group refers to a cancer organoid that has not been treated with an anticancer agent or an anticancer agent candidate. The positive control group refers to a cancer organoid treated with a substance known in the art to be effective in the treatment, improvement, or prevention of cancer, or a drug used as an anticancer agent, in addition to the anticancer agent or anticancer agent candidate whose efficacy or toxicity is to be evaluated. The negative control group and the positive control group may be used to compare the results of anticancer activity with the cancer organoid treated with the anticancer agent or anticancer agent candidate subject to the efficacy or toxicity evaluation.

[0063] The efficacy or toxicity of an anticancer drug or anticancer drug candidate can be evaluated using the cancer organoid of the present invention, which replicates the tumor microenvironment in which cancer cells exist in vivo, by using the method for evaluating the efficacy or toxicity of an anticancer drug according to the present invention. Therefore, by using the method for evaluating the efficacy or toxicity of an anticancer drug according to the present invention, the efficacy or toxicity of the anticancer drug or anticancer drug candidate when administered in vivo can be accurately predicted.

[0064]

[0065] Another aspect of the present invention provides a system for evaluating anticancer efficacy or toxicity comprising a cancer organoid of the present invention.

[0066] In addition, another aspect of the present invention provides a system for evaluating anticancer drug efficacy or toxicity, comprising a method for evaluating anticancer drug efficacy or toxicity according to the present invention.

[0067] In the anticancer drug efficacy or toxicity evaluation system according to the present invention, each term has the same meaning as described in the anticancer drug efficacy or toxicity evaluation method above, unless specifically stated otherwise.

[0068]

[0069] Another aspect of the present invention provides a method for screening anticancer agents, comprising the steps of: producing a cancer organoid by culturing cancer cells isolated from an individual in a culture medium composition for producing a cancer organoid according to the present invention; and treating the cancer organoid with an anticancer agent or a candidate anticancer agent.

[0070] In the screening method for anticancer drugs according to the present invention, each term has the same meaning as described in the efficacy or toxicity evaluation system for anticancer drugs unless specifically stated otherwise.

[0071] In the screening method for anticancer agents of the present invention, the screening of anticancer agents may involve finding a substance that can be used as an anticancer agent among anticancer agent candidate substances, or finding an anticancer agent that is most specifically effective for a patient among drugs currently used as anticancer agents in the industry.

[0072] The screening method for anticancer agents according to the present invention may further include a step of analyzing the anticancer activity of a cancer organoid treated with an anticancer agent or an anticancer agent candidate substance. Additionally, the screening method for anticancer agents according to the present invention may further include a step of comparing the anticancer activity of a cancer organoid treated with an anticancer agent or an anticancer agent candidate substance with the anticancer activity of a control group. The control group may be a cancer organoid not treated with an anticancer agent or an anticancer agent candidate substance (negative control group), or a cancer organoid treated with an anticancer agent having anticancer activity in the subject (positive control group).

[0073] In addition, the method may further include a step of determining that the anticancer agent or anticancer agent candidate can be used as an anticancer agent if the anticancer activity in the cancer organoid treated with the anticancer agent or anticancer agent candidate increases compared to a negative control, or if the anticancer activity increases or is similar compared to a positive control.

[0074] By using the anticancer agent screening method according to the present invention, anticancer agents or anticancer agent candidate substances capable of exhibiting excellent efficacy when administered in vivo can be screened through the cancer organoid of the present invention, which replicates the tumor microenvironment in which cancer cells exist in vivo. Therefore, the anticancer agent screening method of the present invention can be very usefully utilized for personalized medicine and precision medicine for patients.

[0075]

[0076] Another aspect of the present invention provides an anticancer drug screening system comprising a cancer organoid of the present invention.

[0077] In addition, another aspect of the present invention provides an anticancer drug screening system comprising a screening method for anticancer drugs according to the present invention.

[0078] In the anticancer drug screening system according to the present invention, each term has the same meaning as described in the anticancer drug screening method unless specifically stated otherwise.

[0079]

[0080] Another aspect of the present invention provides the use of a culture medium composition or culture medium additive comprising TCTP for the production of cancer organoids.

[0081] Another aspect of the present invention provides the use of a medium composition or medium additive comprising TCTP for promoting the production of cancer organoids.

[0082] In the use of a culture medium composition or culture medium additive containing TCTP according to the present invention, each term has the same meaning as described in the screening system of the anticancer agent, unless specifically stated otherwise.

[0083] In the production of cancer organoids, using a medium composition containing the translationally controlled tumor protein (TCTP) of the present invention allows for obtaining a larger amount of cancer organoids in a shorter period; therefore, the medium composition for producing cancer organoids of the present invention can be economically used for the mass production of cancer organoids. Accordingly, in the production of cancer organoids, the evaluation of the efficacy or toxicity of anticancer drugs or anticancer drug candidates using cancer organoids, and screening, the medium composition for producing cancer organoids of the present invention can be utilized as a medium that can be provided at a more competitive price.

[0084] Figure 1 is an image of organoids taken with an optical microscope on the 14th day after culture of lung cancer organoids #1 and #2 derived from pleural fluid and gastric cancer organoids derived from ascites. #1 and #2 refer to one of the lung cancer organoids prepared in Example 1, control group 1 refers to a medium containing R-spondin, and experimental group 3 refers to a medium containing TCTP.

[0085] Figure 2a is a graph showing the results of a cell viability assay measuring the ATP (adenosine triphosphate) concentration (μM) of lung cancer organoid #1 derived from pleural fluid. #1 refers to the type of lung cancer organoid prepared in Example 1, control group 1 refers to a medium containing R-spondin, experimental group 1 refers to a medium containing 0.625 ng / mL of TCTP, experimental group 2 refers to a medium containing 1.25 ng / mL of TCTP, experimental group 3 refers to a medium containing 2.5 ng / mL of TCTP, experimental group 4 refers to a medium containing 5 ng / mL of TCTP, and experimental group 5 refers to a medium containing 10 ng / mL of TCTP.

[0086] Figure 2b is a graph showing the results of a cell viability test measuring the ATP concentration (μM) of ascites-derived gastric cancer organoids. The terms control group 1 and experimental groups 1 to 5 have the same meaning as defined in Figure 2a.

[0087] Figure 2c is a graph showing the results of a cell viability test measuring the ATP concentration (μM) of tissue-derived liver cancer organoids. The terms control group 1, experimental group 1, experimental group 3, and experimental group 5 have the same meaning as defined in Figure 2a.

[0088] Figure 3 is a graph showing the cell RLU (relative light units) % measured on the 7th day after culture of tissue-derived endometrial cancer organoids. Experimental group 1 refers to a medium containing 0.625 ng / mL of TCTP, experimental group 2 refers to a medium containing 1.25 ng / mL of TCTP, and experimental group 3 refers to a medium containing 2.5 ng / mL of TCTP.

[0089] Figure 4a is an image of an organoid taken with an optical microscope on the 14th day after culture of a tissue-derived endometrial cancer organoid. Control group 2 refers to a medium containing R-spondin, and experimental group 6 refers to a medium containing TCTP.

[0090] Figure 4b is a graph showing the RLU % measured on day 14 after culture of tissue-derived endometrial cancer organoids. Control group 2 refers to a medium containing R-spondin, experimental group 6 refers to a medium containing TCTP, and NS means not significant.

[0091] Figure 5 is a graph showing ATP concentrations measured on days 1, 4, 7, and 14 after culture of tissue-derived colorectal cancer organoids. Control group 3 refers to a medium containing noggin and growth factors, experimental group 7 refers to a medium containing TCTP and growth factors, and experimental group 8 refers to a medium containing TCTP and noggin.

[0092] Figure 6a is an image of an organoid taken with an optical microscope on the 11th day after culture of a gastric cancer organoid derived from ascites. Control group 4 refers to a medium containing R-spondin, noggin, and growth factors, experimental group 9 refers to a medium containing TCTP, noggin, and growth factors, experimental group 10 refers to a medium containing TCTP and growth factors, and experimental group 11 refers to a medium containing TCTP and noggin.

[0093] Figure 6b is a graph showing the number of organoids measured on day 11 after culture of ascites-derived gastric cancer organoids. Control group 4 refers to a medium containing R-spondin, noggin, and growth factors, experimental group 9 refers to a medium containing TCTP, noggin, and growth factors, experimental group 10 refers to a medium containing TCTP and growth factors, experimental group 11 refers to a medium containing TCTP and noggin, and NS means not significant.

[0094] Figure 7a is an image of the organoid taken with an optical microscope on the 14th day after culture of lung cancer organoid #2 derived from pleural fluid. #2 refers to the type of lung cancer organoid prepared in Example 1, control group 5 refers to a medium containing R-spondin, noggin, and growth factors, experimental group 12 refers to a medium containing TCTP, noggin, and growth factors, experimental group 13 refers to a medium containing TCTP and growth factors, and experimental group 14 refers to a medium containing TCTP and noggin.

[0095] Figure 7b is a graph showing the RLU % measured on day 14 after culture of lung cancer organoid #2 derived from pleural fluid. #2 refers to the type of lung cancer organoid prepared in Example 1, control group 5 refers to a medium containing R-spondin, noggin, and growth factors, experimental group 12 refers to a medium containing TCTP, noggin, and growth factors, experimental group 13 refers to a medium containing TCTP and growth factors, experimental group 14 refers to a medium containing TCTP and noggin, and NS means not significant.

[0096] Figure 8a shows images of the organoid taken with an optical microscope on days 7 and 11 after culturing lung cancer organoid #1 derived from pleural fluid. #1 refers to the type of lung cancer organoid prepared in Example 1, control group 5 refers to a medium containing R-spondin, noggin, and growth factors, experimental group 12 refers to a medium containing TCTP, noggin, and growth factors, experimental group 13 refers to a medium containing TCTP and growth factors, and experimental group 14 refers to a medium containing TCTP and noggin.

[0097] Figure 8b is a graph showing the RLU % measured on day 7 after culture of pleural fluid-derived lung cancer organoid #1. #1 refers to the type of lung cancer organoid prepared in Example 1, control group 5 refers to a medium containing R-spondin, noggin, and growth factors, experimental group 12 refers to a medium containing TCTP, noggin, and growth factors, experimental group 13 refers to a medium containing TCTP and growth factors, experimental group 14 refers to a medium containing TCTP and noggin, * indicates P < 0.05, ** indicates P < 0.01, and NS indicates not significant.

[0098] Figure 8c is a graph showing the RLU % measured on day 11 after culture of pleural fluid-derived lung cancer organoid #1. #1 refers to the type of lung cancer organoid prepared in Example 1, control group 5 refers to a medium containing R-spondin, noggin, and growth factors, experimental group 12 refers to a medium containing TCTP, noggin, and growth factors, experimental group 13 refers to a medium containing TCTP and growth factors, experimental group 14 refers to a medium containing TCTP and noggin, and NS means not significant.

[0099] Figure 9a is an image showing the results of a comparative sequencing analysis regarding the correlation of the genomes of gastric cancer tissue, gastric cancer organoids derived from ascites, and lung cancer organoids #1 and #2 derived from pleural effusion. #1 and #2 refer to the types of lung cancer organoids prepared in Example 1, control group 1 refers to a medium containing R-spondin, and experimental group 3 refers to a medium containing 2.5 ng / ml of TCTP.

[0100] Figure 9b is an image showing the oncoplot of the top 27 variant genes. Each term has the same meaning as defined in Figure 9a.

[0101] The present invention will be explained in more detail below by way of examples. However, these examples are intended to illustrate the invention and the scope of the invention is not limited by these examples.

[0102]

[0103] Example 1. Preparation of cancer organoids according to culture medium components

[0104] After counting cells derived from pleural effusion, tissue, or ascites, the cells were mixed with 70% Matrigel to a concentration of 4,000 to 6,000 cells / µl. The mixture was seeded in a dome shape at a dose of 5 µl per well of a 96-well plate, then the plate was inverted and stored in a 37°C incubator to gel. Once gelation was complete, 100 µl of culture medium was dispensed into each well of the plate and cultured for 14 days to produce pleural effusion-derived lung cancer organoids #1 and #2, ascites-derived gastric cancer organoids, tissue-derived colorectal cancer organoids, tissue-derived endometrial cancer organoids, and tissue-derived liver cancer organoids. Different cell lines were used for lung cancer organoids #1 and #2, respectively. Advanced DMEM / F12 was used as the basic medium for the above culture medium, and the components listed in Table 1 were added to prepare control groups 1 and 2 and experimental groups 1 to 6. Except for the components listed in Table 1, the conditions of the control and experimental groups were identical. Here, R-spondin is well known as a component mainly used in organoid culture.

[0105] R-spondin 3 R-spondin 1 TCTP Control Group 1,250 ng / ㎖ -- Control Group 2 -- 1,000 ng / ㎖ -- Experimental Group 1 -- 0.625 ng / ㎖ Experimental Group 2 -- 1.25 ng / ㎖ Experimental Group 3 -- 2.5 ng / ㎖ Experimental Group 4 -- 5 ng / ㎖ Experimental Group 5 -- 10 ng / ㎖ Experimental Group 6 -- 0.156 ng / ㎖

[0106]

[0107] In addition, control groups 3 to 5 and experimental groups 7 to 14 were prepared by adding the components listed in Table 2 to the basic medium above. Except for the components listed in Table 2, the conditions of the control and experimental groups are identical. Here, R-spondin, noggin, and growth factors are well known as components mainly used in organoid culture.

[0108] R-spondin 3TCTPnogginEGFFGF7FGF10Control 3--100 ng / mL50 ng / mL--Control 4250 ng / mL-100 ng / mL50 ng / mL-200 ng / mLControl 5250 ng / mL-100 ng / mL50 ng / mL5 ng / mL20 ng / mL Experimental group 7-2.5 ng / mL-50 ng / mL--Experimental group 8-2.5 ng / mL100 ng / mL---Experimental group 9-2.5 ng / mL100 ng / mL50 ng / mL-200 ng / mL experimental group 10-2.5 ng / mL-50 ng / mL-200 ng / mL experimental group 11-2.5 ng / mL100 ng / mL---experimental group 12-2.5 ng / mL100 ng / mL50 ng / mL5 ng / mL20 ng / mL Experimental group 13-2.5 ng / mL-50 ng / mL5 ng / mL20 ng / mL Experimental group 14-2.5 ng / mL100 ng / mL---

[0109]

[0110] Example 2. Confirmation of cancer organoid growth

[0111] Example 2-1.

[0112] To compare the growth of cancer organoids prepared using a medium containing TCTP and cancer organoids prepared using a medium containing R-spondin, the number of cancer organoids prepared in Example 1 was compared. The number of organoids refers to the number of organoids grown from a single cell, and through the number of organoids, it can be confirmed that the colony-forming ability and proliferation-promoting ability of cancer organoids differ depending on the medium components.

[0113] Specifically, images of lung cancer organoids #1, #2 and gastric cancer organoids prepared in Example 1 were taken with an optical microscope on the 14th day after culture, and the results are shown in Figure 1. In addition, the number of organoids was automatically counted using the ImageJ program, and the results are shown in Table 3.

[0114] Organoid Type | Culture Medium | Organoid | Number | Lung Cancer Organoid #1 | Control Group 1796 | Experimental Group 32816 | Lung Cancer Organoid #2 | Control Group 12174 | Experimental Group 32227 | Gastric Cancer Organoid | Control Group 1720 | Experimental Group 34220

[0115] As a result, as can be seen in Figure 1 and Table 3, the number of cancer organoids produced in a medium containing 2.5 ng / ml of TCTP (experimental group 3) was higher or similar to the number of cancer organoids produced in a medium containing R-spondin (control group 1). These results suggest that a medium containing TCTP promotes the proliferation of cancer organoids, allowing for a larger number of cancer organoids to be produced, and that the ability of the medium containing TCTP to promote cancer organoid proliferation is similar to or significantly higher than that of the medium containing R-spondin.

[0116]

[0117] Example 3. Confirmation of cell activity of cancer organoids

[0118] An ATP (adenosine triphosphate) assay was performed to confirm the cell activity of cancer organoids prepared using a medium containing TCTP of the present invention and cancer organoids prepared using a medium containing R-spondin.

[0119] Specifically, for the ATP assay, relative light units (RLU) were measured using the celltiter glo 3D kit, and the ATP concentration and organoid number were calculated based on the measured RLU %. As ATP serves as a marker of cell viability, serving as an energy source for living cells and participating in various biochemical reactions, cell viability can be verified through ATP measurement (Yoshihiko Maehara et al. (1987), The ATP assay is more sensitive than the succinate dehydrogenase inhibition test for predicting cell viability, European Journal of cancer and clinical oncology, volume 23, issue 3, pages 273-276; and Russell D. Petty, et al. (1995), Comparison of MTT and ATP-based assays for the measurement of viable cell number, Journal of Bioluminescence and Chemiluminescence, volume 10, issue 1, pages 29-34). Therefore, a higher ATP concentration means that there are more living cells.

[0120]

[0121] Example 3-1.

[0122] The ATP concentration (μM) of the lung cancer organoid #1 and gastric cancer organoid prepared in Example 1 was measured on days 1, 3, 7, and 14 after culture, and the results are shown in Figures 2a and 2b.

[0123] In addition, the ATP concentration (μM) was measured on the 7th day after culture of the liver cancer organoid prepared in Example 1, and the results are shown in Figure 2c.

[0124] As a result, as can be seen in Figures 2a to 2c, the ATP concentration of cancer organoids prepared with a medium containing TCTP (experimental groups 1 to 5) was found to be higher or similar to the ATP concentration of cancer organoids prepared with a medium containing R-spondin (control group 1).

[0125] The above results suggest that a medium containing TCTP promotes the proliferation of cancer organoids, allowing for a larger number of cancer organoids to be obtained, and that the ability of a medium containing TCTP to promote cancer organoid proliferation is similar to or significantly higher than that of R-spondin.

[0126]

[0127] Example 3-2.

[0128] In Example 1, the RLU % of endometrial cancer organoids prepared in media containing TCTP at concentrations of 0.625 ng / mL, 1.25 ng / mL, and 2.5 ng / mL (in order: experimental group 1, experimental group 2, and experimental group 3) was measured on the 7th day after culture, and the results are shown in Figure 3.

[0129] As a result, as can be seen in Figure 3, the RLU % increased as the concentration of TCTP in the medium increased.

[0130] The above results suggest that the ability of a medium containing TCTP to promote cancer organoid proliferation tends to be dependent on the concentration of TCTP.

[0131]

[0132] Example 3-3.

[0133] In Example 1, images were taken of endometrial cancer organoids prepared in a medium not containing TCTP (control group 2) or a medium containing TCTP (experimental group 6) on the 14th day after culture and RLU % was measured, and the results are shown in Figures 4a and 4b, respectively.

[0134] As a result, as can be seen in Figures 4a and 4b, the RLU % of endometrial cancer organoids prepared in a medium containing TCTP (experimental group 6) was found to be not significantly different from the RLU % of endometrial cancer organoids prepared in a medium containing R-spondin (control group 2) (NS, not significant).

[0135] The above results suggest that the ability of a medium containing TCTP to promote cancer organoid proliferation is similar to that of a medium containing R-spondin.

[0136]

[0137] Examples 3-4.

[0138] In Example 1, the ATP concentration (μM) was measured on days 1, 4, 7, and 14 after culturing colorectal cancer organoids prepared in a medium not containing TCTP (control group 3) or a medium containing TCTP (experimental group 7 or experimental group 8), and the results are shown in Figure 5.

[0139] As a result, as shown in Figure 5, although the medium without TCTP (control group 3) contained both noggin and growth factors, and the medium containing TCTP (experimental groups 7 and 8) did not contain noggin or growth factors, the ATP concentration of colorectal cancer organoids prepared with the medium containing TCTP (experimental groups 7 and 8) was found to be higher or similar to that of colorectal cancer organoids prepared with the medium without TCTP (control group 3). In particular, on the 14th day after culture, the ATP concentration of experimental group 7 was approximately twice as high as that of the control group, and the ATP concentration of experimental group 8 was approximately five times higher than that of the control group.

[0140] The above results suggest that the ability of a medium containing TCTP to promote cancer organoid proliferation is similar to or significantly higher than that of a medium containing R-spondin. Furthermore, it suggests that a medium containing TCTP can reduce dependence on the medium components conventionally used for cancer organoid production.

[0141]

[0142] Examples 3-5.

[0143] In Example 1, images were taken on the 11th day after culturing gastric cancer organoids prepared in a medium containing R-spondin (control group 4) or a medium containing TCTP (experimental groups 9 to 11), and the number of organoids was calculated, and the results are shown in Figures 6a and 6b, respectively.

[0144] As a result, as can be seen in Figures 6a and 6b, the number of gastric cancer organoids produced in a medium containing TCTP (experimental groups 9 to 11) was found to be not significantly different from the number of gastric cancer organoids produced in a medium containing R-spondin (control group 4) (NS, not significant). In particular, while the medium containing R-spondin (control group 4) contains both noggin and growth factors, among the TCTP-containing media, experimental group 10 does not contain noggin and experimental group 11 does not contain growth factors; however, the number of gastric cancer organoids produced in experimental group 10 or experimental group 11 was found to be not significantly different from the number of gastric cancer organoids produced in a medium containing R-spondin (control group 4).

[0145] The above results suggest that a medium containing TCTP promotes the proliferation of cancer organoids, allowing for a larger number of cancer organoids to be obtained, and that the ability of the medium containing TCTP to promote cancer organoid proliferation is similar to that of R-spondin.

[0146]

[0147] Examples 3-6.

[0148] In Example 1, lung cancer organoid #2 prepared in a medium containing R-spondin (control group 5) or a medium containing TCTP (experimental groups 12 to 13) was imaged on the 14th day after culture and RLU % was measured, and the results are shown in Figures 7a and 7b, respectively.

[0149] As a result, as can be seen in Figures 7a and 7b, the RLU % of lung cancer organoids prepared in a medium containing TCTP (experimental groups 12 to 13) was found to be not significantly different from the RLU % of lung cancer organoids prepared in a medium containing R-spondin (control group 5) (NS, not significant). In particular, while the medium containing R-spondin (control group 5) contains noggin, EGF, and FGF, experimental group 13 among the TCTP-containing media does not contain noggin and experimental group 14 does not contain growth factors, the RLU % of lung cancer organoids prepared in experimental group 13 or experimental group 14 was found to be not significantly different from the RLU % of lung cancer organoids prepared in a medium containing R-spondin (control group 5).

[0150] The above results suggest that a medium containing TCTP promotes the proliferation of cancer organoids, enabling the production of a larger number of cancer organoids, and that the ability of the medium containing TCTP to promote cancer organoid proliferation is similar to that of R-spondin. Furthermore, the results suggest that a medium containing TCTP can reduce dependence on the medium components conventionally used for cancer organoid production.

[0151]

[0152] Examples 3-7.

[0153] In Example 1, lung cancer organoid #1 prepared in a medium containing R-spondin (control group 5) or a medium containing TCTP (experimental groups 12 to 14) was imaged on days 7 and 11 after culture and RLU % was measured, and the results are shown in Figures 8a to 8c.

[0154] As a result, as can be seen in Figures 8a to 8c, on the 11th day after culture, the ATP concentration (uM) of lung cancer organoids prepared in a medium containing TCTP (experimental groups 12 to 14) was found to be not significantly different from the ATP concentration of lung cancer organoids prepared in a medium containing R-spondin (control group 5) (NS, not significant). In addition, while the medium containing R-spondin (control group 5) contains both noggin and growth factors, experimental group 13 among the TCTP-containing media does not contain noggin and experimental group 14 does not contain growth factors, the RLU % of lung cancer organoids prepared in experimental group 13 or experimental group 14 was found to be not significantly different from the RLU % of lung cancer organoids prepared in a medium containing R-spondin (control group 5).

[0155] In particular, on the 7th day after culturing lung cancer organoids, the RLU % of lung cancer organoids produced in a medium containing TCTP (experimental groups 12 to 14) was found to be similar to or significantly higher than the RLU % of lung cancer organoids produced in a medium containing R-spondin (control group 5), confirming that the medium containing TCTP (experimental groups 12 to 14) produced a similar or greater number of organoids than the medium containing R-spondin (control group 5) in a short period of time.

[0156] The above results suggest that the ability of a medium containing TCTP to promote cancer organoid proliferation is similar to or significantly higher than that of a medium containing R-spondin. Furthermore, the medium containing TCTP suggests that more cancer organoids can be produced in a shorter time compared to a medium containing R-spondin, and that dependence on the medium components conventionally used for cancer organoid production can be reduced.

[0157]

[0158] Example 4. Comparative Genome Analysis of Cancer Organoids

[0159] Genomic similarity was analyzed to compare the genomic variation characteristics of a cancer organoid prepared using a medium containing TCTP of the present invention and a cancer organoid prepared using a medium containing R-spondin.

[0160] Specifically, the genomes of gastric cancer tissue, gastric cancer organoids prepared using the medium containing R-spondin in Example 1 (Control Group 1) or the medium containing TCTP 2.5 ng / mL (Experimental Group 3), and lung cancer organoids #1 and #2 were sequenced, and then the genomes and the oncoplots of the top 27 variant genes were compared and analyzed. The comparative genomic analysis was performed on the correlation, matching, and concordance of the genomes, and the results are shown in Table 4 and Figure 9a.

[0161] Sample A Sample B Correlation Matching Status Similarity Gastric Cancer Organoid (Control Group 1) Gastric Cancer Organoid (Experimental Group 3) 0.9925 Match 0.890 Lung Cancer Organoid #1 (Control Group 1) Lung Cancer Organoid #1 (Experimental Group 3) 0.9919 Match 0.879 Lung Cancer Organoid #2 (Control Group 1) Lung Cancer Organoid #2 (Experimental Group 3) 0.9896 Match 0.885

[0162]

[0163] In addition, tumor plot analysis of the top 27 variant genes was performed on missense mutations, nonsense mutations, frame shift mutations caused by insertion, in frame insertion mutations, and multi-variants, and the results are shown in Figure 9b. Missense mutations refer to mutations in which the amino acid codon is changed due to substitution of the base sequence, and missense mutations are classified into non-conservative mutations, in which the type of amino acid changes, and conservative mutations, in which the type of amino acid does not change. Nonsense mutations refer to mutations in which a stop codon is introduced due to a substitution of the base sequence, resulting in the production of a short protein; frameshift mutations due to base insertion refer to mutations in which a base is inserted, causing a shift in the reading frame of the codon; inframe insertion mutations refer to mutations in which a base is inserted but does not cause a shift in the reading frame; and multiple mutations refer to the occurrence of two or more mutations.

[0164] As a result, as shown in Table 4 and Figure 9a, a high correlation is observed when comparing the genomic characteristics of gastric cancer organoids prepared with a medium containing R-spondin (Control Group 1) and gastric cancer organoids prepared with a medium containing TCTP (Experimental Group 3) with the genomic characteristics of gastric cancer tissue. Furthermore, the genomic characteristics of the cancer organoids prepared with the medium containing TCTP (Experimental Group 3) show a similarity of approximately 88% or more with the genomic characteristics of the cancer organoids prepared with the medium containing R-spondin (Control Group 1).

[0165] In addition, as can be seen in Figure 9b, the cancer organoids prepared with a medium containing TCTP (experimental group 3) were similar to the cancer organoids prepared with a medium containing R-spondin (control group 1) in most of the top 27 gene variants.

[0166] The above results suggest that since the medium containing TCTP does not affect changes in the cancer organoid genome, TCTP can be used in the medium for producing cancer organoids as a replacement for R-spondin.

Claims

1. A culture medium composition for producing cancer organoids, comprising TCTP (translationally controlled tumor protein).

2. In Paragraph 1, A culture medium composition for preparing cancer organoids, wherein the above cancer is selected from the group consisting of liver cancer, lung cancer, non-small cell lung cancer, stomach cancer, colorectal cancer, pancreatic cancer, biliary tract cancer, colon cancer, small intestine cancer, brain cancer, bone cancer, melanoma, breast cancer, sclerosing adenoma, uterine cancer, endometrial cancer, cervical cancer, vaginal cancer, vulvar cancer, head cancer, cervical cancer, esophageal cancer, thyroid cancer, parathyroid cancer, endocrine gland cancer, kidney cancer, sarcoma, prostate cancer, urethral cancer, penile cancer, bladder cancer, ureteral cancer, hematological cancer, central nervous system tumor, spinal cord tumor, glioblastoma, brainstem glioma, pituitary adenoma, leukemia, lymphoma, and fibroadenoma.

3. In Paragraph 1, A culture medium composition for producing cancer organoids, wherein the culture medium composition additionally comprises basal media.

4. In Paragraph 3, A medium composition for preparing cancer organoids, wherein the basic medium is DMEM, MEM, BME, RPMI1640, F-10, F-12, α-MEM, GMEM, IMDM, McCoy's 5A, Neurobasal medium, DMEM / F12, or Advanced DMEM / F12.

5. A medium additive for the production of cancer organoids containing TCTP.

6. In Paragraph 5, A culture medium additive for preparing cancer organoids, wherein the above cancer is selected from the group consisting of liver cancer, lung cancer, non-small cell lung cancer, stomach cancer, colorectal cancer, pancreatic cancer, biliary tract cancer, colon cancer, small intestine cancer, brain cancer, bone cancer, melanoma, breast cancer, sclerosing adenoma, uterine cancer, endometrial cancer, cervical cancer, vaginal cancer, vulvar cancer, head cancer, cervical cancer, esophageal cancer, thyroid cancer, parathyroid cancer, endocrine gland cancer, kidney cancer, sarcoma, prostate cancer, urethral cancer, penile cancer, bladder cancer, ureteral cancer, hematological cancer, central nervous system tumor, spinal cord tumor, glioblastoma, brainstem glioma, pituitary adenoma, leukemia, lymphoma, and fibroadenoma.

7. In Paragraph 5, The above-mentioned medium is a medium additive for manufacturing cancer organoids, which additionally includes a basic medium.

8. In Paragraph 7, The above basic medium is a medium additive for the preparation of cancer organoids, which is DMEM, MEM, BME, RPMI1640, F-10, F-12, α-MEM, GMEM, IMDM, McCoy's 5A, Neurobasal medium, DMEM / F12, or Advanced DMEM / F12.

9. A method for producing a cancer organoid comprising the step of culturing cancer cells isolated from an individual in the culture medium composition of claim 1.

10. A step of producing a cancer organoid by culturing cancer cells isolated from an individual in the culture medium composition of claim 1; and A method for evaluating the efficacy or toxicity of an anticancer drug, comprising the step of treating the cancer organoid with an anticancer drug or a candidate anticancer drug.

11. In Paragraph 10, A method for evaluating the efficacy or toxicity of an anticancer drug, wherein the method further comprises the step of analyzing the anticancer activity of a cancer organoid treated with an anticancer drug or an anticancer drug candidate substance.

12. In Paragraph 11, A method for evaluating the efficacy or toxicity of an anticancer drug, wherein the above anticancer activity is inhibition of growth, increased death, reduction in size, reduction in area, or reduction in weight of cancer organoids.

13. A step of producing a cancer organoid by culturing cancer cells isolated from an individual in the culture medium composition of claim 1; and A screening method for anticancer drugs comprising the step of treating the cancer organoid with an anticancer drug or an anticancer drug candidate.

14. In Paragraph 13, The above screening method for anticancer drugs further comprises the step of analyzing the anticancer activity of a cancer organoid treated with an anticancer drug or an anticancer drug candidate.

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