Culture medium design

Abstract

This disclosure provides a technique for finding the optimal culture medium components and culture conditions, a method for efficiently finding the optimal culture medium and culture conditions, and a technique for utilizing this technique. [Solution] This disclosure provides a method for quality control of cells, which includes adding a culture medium containing a component to cells and culturing them, obtaining a coefficient related to the component, and determining whether the culture medium containing the component is correlated based on the coefficient.

Description

[Technical Field]

[0001] This disclosure relates to the design, quality control, and techniques for using culture media suitable for cells. This disclosure also relates to the optimization of culture medium components and cell culture conditions in cell culture conditions, in particular to methods for optimizing culture medium components and culture conditions to promote cell proliferation and differentiation and improve cell quality. [Background technology]

[0002] Because stem cells play a crucial role in regenerative medicine and cell therapy, there is a demand for a supply of high-quality stem cells. The quality of stem cells largely depends on the culture conditions, and it is known that the components of the culture medium, in particular, affect the characteristics of the cells.

[0003] Cell culture conditions are a crucial technology in many fields, including biotechnology, regenerative medicine, and drug screening. The composition of the culture medium and the culture conditions play a vital role in optimizing cell proliferation and differentiation. [Overview of the Initiative] [Means for solving the problem]

[0004] As a result of diligent research, this disclosure provides a technique for finding the optimal culture medium components and culture conditions, a method for efficiently finding the optimal culture medium and culture conditions, and a technique for utilizing this technique.

[0005] <Quality standards> (Item 1) A method for quality control of cells, comprising: adding a culture medium containing a component to cells and culturing them; obtaining a coefficient related to the component; and determining the correlation between the culture medium containing the component and the cells based on the coefficient. (Item 2) The method described in the above item, wherein the determination includes determining whether the target cell is the target cell. (Item 3) The method according to any one of the above items, wherein the determination includes determining whether the culture medium or the component is the culture medium or the component of the interest. (Item 4) The component is serum, various vitamins, various antibiotics, various hormones, various growth factors, and / or the coefficient is any coefficient described herein, the method of any one of the above items, for example, (1) Ingredients: Pyruvate, Cystine, Serine Coefficient: Mitochondrial size (2) Components: PTEN inhibitor, p53 inhibitor, p38 inhibitor, Wnt sigma A component selected from the group consisting of a naru activator and a ROCK inhibitor, preferably the ROCK inhibitor Y-27632. Coefficient: Expression level of Ror2 or Fzd5 The method described in any one of the above items, which may include the above. (Item 5) The method according to any one of the above items, wherein the determination includes referring to a database that stores information relating to the correlation between the component and the coefficient. (Item 6) The above determination is, Controlling the behavior of one or more specific protein molecules related to the quality of the aforementioned cells, To manage the expression behavior of related gene groups that are characteristic of the quality of the aforementioned cells, and The aforementioned specific protein molecule is a cell surface antigen. The characteristic gene groups associated with the aforementioned cells include immune-related gene groups, inflammation-related gene groups, angiogenesis-related gene groups, nerve regeneration-related gene groups, cartilage formation-related gene groups, bone formation-related gene groups, and lipids. It includes at least one selected from the group consisting of lipidogenesis-related genes, fibrosis-related genes, immunogenicity-related genes, migration-related genes, adhesion-related genes, aging-related genes, mesenchymal stem cell marker-related genes, surface marker-related genes, growth factor-related genes, and chemokine or cytokine-related genes, In managing the expression behavior of the gene group characteristic of the aforementioned cells, if the correlation between the expression pattern of the gene group obtained by measuring the expression state of the gene group characteristic of the aforementioned cells and a standard expression pattern is above a certain level, the component is determined to be suitable for quality control. The correlation is above a certain level when the correlation coefficient or the coefficient of determination, or the correlation coefficient and the coefficient of determination, are evaluated and the correlation coefficient or the coefficient of determination is above a certain level. The method described in any one of the above items. (Item 6A) When evaluating the correlation coefficient or the coefficient of determination, (i) Substitute missing values ​​for the lower limit of measurement for gene expression levels that cannot be measured. (ii) Substitute missing values ​​for which gene expression levels cannot be measured with the lowest value among the measured values, or (iii) Process missing values ​​for which gene expression levels cannot be measured using the following formula: Read count for each gene + (1 × gene length / average gene length) The correlation coefficient or the coefficient of determination is calculated by performing one of the following: The method described in any one of the above items. (Item 7) The method according to any one of the above items, wherein the behavior of the specific protein molecule is at the expression level. (Item 8) The method according to any one of the above items, wherein the cells are at least one type of cell selected from the group consisting of somatic stem cells, induced pluripotent stem cells, embryonic stem cells, differentiated cells derived from somatic stem cells, differentiated cells derived from induced pluripotent stem cells, and differentiated cells derived from embryonic stem cells. (Item 9) The method according to any one of the above items, wherein the cells are mesenchymal stem cells. (Item 10) The method according to any one of the above items, wherein the cells are mesenchymal stem cells for drug discovery. (Item 11) Selecting a culture medium and, if necessary, cells that meet the criteria from a cell population including a culture medium and, if necessary, candidate cells for therapeutic or drug discovery, by means of the method described in any one of the above items, and The selected cells are grown in the selected culture medium. A method for producing culture media or cells for therapeutic or drug discovery purposes, including the above. (Item 12) Adding a culture medium containing the component to the cells and culturing them, obtaining a coefficient related to the component, and determining whether the culture medium containing the component is correlated based on the coefficient, A method for producing a quality-controllable culture medium, which includes, if the aforementioned correlation is found, manufacturing a culture medium containing that component as a controllable culture medium. (Item 13) The component is serum, various vitamins, various antibiotics, various hormones, various growth factors, and / or the coefficient is any coefficient described herein, the method described in any one of the above items, for example, (1) Ingredients: Pyruvate, Cystine, Serine Coefficient: Mitochondrial size (2) Components: Components selected from the group consisting of PTEN inhibitors, p53 inhibitors, p38 inhibitors, Wnt signaling activators, and ROCK inhibitors, preferably the ROCK inhibitor Y-27632 Coefficient: The method described in any one of the above items, which may be the expression level of Ror2 or Fzd5, etc. (Item 14) The method according to any one of the above items, wherein the determination includes referring to a database that stores information relating to the correlation between the component and the coefficient. (Item 15) The above determination is, Controlling the behavior of one or more specific protein molecules related to the quality of the aforementioned cells, To manage the expression behavior of related gene groups that are characteristic of the quality of the aforementioned cells, and The aforementioned specific protein molecule is a cell surface antigen. The characteristic gene group for the aforementioned cells includes at least one selected from the group consisting of immune-related gene groups, inflammation-related gene groups, angiogenesis-related gene groups, nerve regeneration-related gene groups, chondrogenesis-related gene groups, bone formation-related gene groups, adipogenesis-related gene groups, fibrosis-related gene groups, immunogenicity-related gene groups, migration-related gene groups, adhesion-related gene groups, aging-related gene groups, mesenchymal stem cell marker-related gene groups, surface marker-related gene groups, growth factor-related gene groups, and chemokine or cytokine-related genes. In managing the expression behavior of the gene group characteristic of the aforementioned cells, if the correlation between the expression pattern of the gene group obtained by measuring the expression state of the gene group characteristic of the aforementioned cells and a standard expression pattern is above a certain level, the component is determined to be suitable for quality control. The correlation is above a certain level when the correlation coefficient or the coefficient of determination, or the correlation coefficient and the coefficient of determination, are evaluated and the correlation coefficient or the coefficient of determination is above a certain level. The method described in any one of the above items. (Item 16) The method according to any one of the above items, wherein the behavior of the specific protein molecule is at the expression level. (Item 17) The method according to any one of the above items, wherein the cells are at least one type of cell selected from the group consisting of somatic stem cells, induced pluripotent stem cells, embryonic stem cells, differentiated cells derived from somatic stem cells, differentiated cells derived from induced pluripotent stem cells, and differentiated cells derived from embryonic stem cells. (Item 18) The method according to any one of the above items, wherein the cells are mesenchymal stem cells. (Item 19) The method according to any one of the above items, wherein the cells are mesenchymal stem cells for drug discovery. (Item 20) Selecting a culture medium and, if necessary, cells that meet the criteria from a cell population including a culture medium and, if necessary, candidate cells for therapeutic or drug discovery, by means of the method described in any one of the above items, and The selected cells are grown in the selected culture medium. A method for producing culture media or cells for therapeutic or drug discovery purposes, including the above. (Item 21) A method for quality control of cells, A method comprising: confirming whether the culture medium used for culturing the cells contains components necessary for quality control of the cells; and, if the necessary components are not present, adding the necessary components to the culture medium. (Item 22) The component is serum, various vitamins, various antibiotics, various hormones, various growth factors, and / or the coefficient is any coefficient described herein, the method of any one of the above items, for example, (1) Ingredients: Pyruvate, Cystine, Serine Coefficient: Mitochondrial size (2) Components: Components selected from the group consisting of PTEN inhibitors, p53 inhibitors, p38 inhibitors, Wnt signaling activators, and ROCK inhibitors. Preferably, the ROCK inhibitor Y-27632 Coefficient: The method described in any one of the above items, which may be the expression level of Ror2 or Fzd5, etc. (Item 23) The method according to any one of the above items, wherein the verification includes referring to a database that stores information relating to the correlation between the component and coefficients related to the component. (Item 24) The above determination is, Controlling the behavior of one or more specific protein molecules related to the quality of the aforementioned cells, To manage the expression behavior of related gene groups that are characteristic of the quality of the aforementioned cells, and The aforementioned specific protein molecule is a cell surface antigen. The aforementioned cell-specific gene groups include immune-related gene groups, inflammation-related gene groups, and angiogenesis. It includes at least one selected from the group consisting of related gene groups, nerve regeneration related gene groups, chondrogenesis related gene groups, bone formation related gene groups, adipogenesis related gene groups, fibrosis related gene groups, immunogenicity related gene groups, migration related gene groups, adhesion related gene groups, aging related gene groups, mesenchymal stem cell marker related gene groups, surface marker related gene groups, growth factor related gene groups, and chemokine or cytokine related genes, In managing the expression behavior of the gene group characteristic of the aforementioned cells, if the correlation between the expression pattern of the gene group obtained by measuring the expression state of the gene group characteristic of the aforementioned cells and a standard expression pattern is above a certain level, the component is determined to be suitable for quality control. The correlation is above a certain level when the correlation coefficient or the coefficient of determination, or the correlation coefficient and the coefficient of determination, are evaluated and the correlation coefficient or the coefficient of determination is above a certain level. The method described in any one of the above items. (Item 25) The method according to any one of the above items, wherein the behavior of the specific protein molecule is at the expression level. (Item 26) The method according to any one of the above items, wherein the cells are at least one type of cell selected from the group consisting of somatic stem cells, induced pluripotent stem cells, embryonic stem cells, differentiated cells derived from somatic stem cells, differentiated cells derived from induced pluripotent stem cells, and differentiated cells derived from embryonic stem cells. (Item 27) The method according to any one of the above items, wherein the cells are mesenchymal stem cells. (Item 28) The method according to any one of the above items, wherein the cells are mesenchymal stem cells for drug discovery. (Item 29) Selecting a culture medium and, if necessary, cells that meet the criteria from a cell population including a culture medium and, if necessary, candidate cells for therapeutic or drug discovery, by means of the method described in any one of the above items, and The selected cells are grown in the selected culture medium. A method for producing culture media or cells for therapeutic or drug discovery purposes, including the above.

[0006] <Design Methods (Screening), Optimization, Business> (In silico) (Item 30) A method for causing a computer to perform the selection of appropriate culture medium components and / or cell culture conditions for cells provided by a user, the method being: The process of inputting cell information provided by the user into the computer, The process involves having the computer, if necessary, consult a database to derive culture medium components and culture conditions related to the cells, as well as optimal conditions if necessary. The process of having the computer calculate the optimal culture medium components and cell culture conditions for the cells based on the culture medium components and the culture conditions, If necessary, the process involves displaying the results of the calculation on a display. A method of including. (Item 30A1) A method for causing a computer to select appropriate culture medium components and / or cell culture conditions for cells provided by a user, the method being: The process of inputting cell information provided by the user into the computer, The process of inputting data on the culture results under specific culture conditions for the cells into the computer, The process of inputting desired output conditions for the cells to the computer, The process involves having the computer, if necessary, consult a database to derive culture medium components and culture conditions related to the cells, as well as optimal conditions if necessary. The process of having the computer calculate the optimal culture medium components and cell culture conditions for the cells based on the culture medium components and the culture conditions, If necessary, the process includes displaying the results of the calculation on a display, If necessary, the process includes documenting the results of the calculations, If necessary, the process includes outputting the documented document, A method of including. (Item 30A2) The documentation is the method described in any one of the above items, as documented in accordance with the standard. (Item 30A3) The above standard includes the method described in any one of the above items, including the standard for the Manufacturing Order Record (MBR) or the Standard Operating Procedure (SOP). (Item 30A4) The method according to any one of the above items, wherein the optimal culture medium components and cell culture conditions for the cells are components and conditions suitable for both small scale and commercial scale or either one, or include culture conditions in a stepwise manner from small scale to commercial scale. (Item 30A5) The method according to any one of the above items, further comprising the steps of extracting predicted risks with respect to the optimal culture medium components and cell culture conditions for the cells, formulating countermeasures for the risks, and providing information on the risks and countermeasures. (Item 30A6) The method according to any one of the above items, wherein the database includes at least one selected from the group consisting of cell information, specifications of an automated culture device, the number of people required depending on the culture conditions, the layout of the manufacturing plant, information on the wiring during culture, and information on the reagents held. (Item 30A7) The optimal culture medium components and cell culture conditions for the cells are the method described in any one of the above items, including components and culture conditions using an automated culture device. (Item 30A8) The cell culture conditions are those described in any one of the above items, including information on manufacturing costs. (Item 30A9) The method according to any one of the above items, wherein the risks include at least one selected from the group consisting of risks of changes in the state and / or quantity of cultured cells during scale-up, risks of non-compliance with GMP, risks of malfunction in reagent preparation during culture, risks in preparing culture medium components including powder preparation and on-demand preparation, risks of quality and lot variability in culture medium, and regional risks. (Item 30A10) The countermeasure is the method described in any one of the above items, which is output based on the information in the database. (Item 30A11) The method according to any one of the above items, wherein the countermeasures include at least one selected from the group consisting of suggesting compatible culture medium factors, suggesting methods for managing and supplying culture medium components, automatically generating risk reduction documents, suggesting routine responses that tolerate biogroups, and alternative culture methods. (Item 30A12) The method described in any one of the above items, including a proposed initial culture method up to commercial production. (Item 30A13) The method according to any one of the above items, wherein the document includes providing information on cell production volume, production efficiency, cost of culture medium components, and composition of culture medium components, as well as at least one selected from the group consisting of proposals for changes to culture medium components, proposals for culture conditions, proposals for alternative methods, provision of MBR proposals, provision of SOP proposals, provision of visualized manufacturing site information, and provision of a roadmap proposal. (Item 31) The above selection is (A) The step of selecting target cells; (B) The step of selecting K basic cellular functions of the cells; (C) A step of determining a set of N culture medium factors that determine the environment containing the components of the culture medium; (D) Using information stored in a database related to the cells, construct a functional map representing the intensity of activation or suppression of each of the K basic cellular functions by each of the N culture medium factors; (E) Using the functional map, the step of optimizing the composition of the cell culture medium so as to activate or suppress one or more of the basic cellular functions; The method described in any one of the above items, including, where K and N are independent integers. (Item 32) The above database contains cytokines, growth factors, and other substances useful for cell culture conditions. A library comprising proteins, small molecules, and physiologically active substances, prepared for use in the composition of culture media, as described in any one of the above items. (Item 33) The method according to any one of the above items, wherein the database is a library containing information on cell culture conditions that can be used to construct the culture medium. (Wet) (Item 34) A method for performing calculations to select appropriate culture medium components and / or cell culture conditions for cells provided by a user, the method is: A step of analyzing cells provided by the user using candidate culture medium components and / or cell culture conditions, The process involves deriving the culture medium components and culture conditions (optimal requirements as needed) related to the cells from the analysis results, A step of calculating appropriate culture medium components and cell culture conditions for the cells based on the culture medium components and culture conditions. A method of including. (Item 35) The above selection is (A) The step of selecting target cells; (B) The step of selecting K basic cellular functions of the cells; (C) A step of determining a set of N culture medium factors that determine the environment containing the components of the culture medium; (D) Using experimental data of novel culture medium samples and / or the supernatant of existing culture medium samples using the components of the culture medium and the cells, a functional map is constructed that represents the intensity of activation or inhibition of each of the K basic cellular functions by each of the N culture medium factors. (E) Using the functional map, the step of optimizing the composition of the cell culture medium so as to activate or suppress one or more of the basic cellular functions; The method described in any one of the above items, including: (Item 35A) The method described in any one of the above items, wherein the candidate culture component is selected from the library of culture components. (Item 36) The candidate cell culture conditions are selected from the library of cell culture conditions, according to the method described in any one of the above items. (Item 37) The method according to any one of the above items, wherein the analysis is performed based on the results of an actual experiment using the culture medium components. (Item 38) The method according to any one of the above items, wherein the analysis is performed based on the results of an actual experiment using the culture medium components and calculations based on those results. (Item 39) The method according to any one of the above items, wherein the analysis comprises applying a library of the culture medium components to cells in a multiwell plate and performing high-throughput screening using an automated instrument. (Item 40) The method according to any one of the above items, wherein the calculation is performed using a library parameterized for the performance and / or cost of the culture component to the cells. (Item 41) The method according to any one of the above items, wherein optimization of the culture medium is achieved to suit the purpose of seed cells or cell lines, cost, and growth performance. (Item 42) The construction of the functional map in step d) above is A step of performing a first run of a culture experiment, comprising a number of cultures equal to (N+1) or more obtained by adding 1 to the number of culture medium factors, wherein each culture is performed with a different culture medium component, and combinations of low-level and high-level culture medium factors are screened; For each culture experiment performed, the steps include obtaining initial and final biomass, product, and exometabolome data, or partial exometabolome data; Relative weighting coefficient λ j A subset of basal cellular functions with a value higher than zero is defined below. The steps include determining the exometabolome data, or the exometabolome data derived from the culture medium component data, using a linear model, and

[0007]

number

[0008]

number

[0009] (where v is a vector representing the rate of change of the component of the exometabolome from which the element was measured, I i,j This is the activation intensity parameter of basic cellular function j by culture medium factor i, determined by regression analysis. A step of performing a second run of a culture experiment, the number of cultures being greater than or equal to the number of activated cell functions plus 1, wherein each culture is performed with a different culture medium component, the medium component having a predetermined intensity parameter value I such that low and high values ​​of the weighted coefficient of activated basic cell function are identified as subsets of activated cell functions controlled by the medium factor. i,j The steps set up to screen using; In all of the above steps, a functional map is constructed from the data collected by linear regression analysis using equations (3a) and (3b), and the data is organized into the form of the functional map, wherein the intensity value I determined by the first run of the experiment is i,j However, this is corrected by the data from the second run of the experiment, resulting in an N×K data array (functional map = {I i,j The step of making it into the shape of}); The method described in any one of the above items, including: (Item 43) The optimization of the culture medium components in step e) is A step of forming basic cell function special culture medium components using the matrix of functional data, wherein the change in the value of the culture medium factor Δ(FAC j )

[0010]

number

[0011] The steps include: determining the change in the relative weights Δλi of basic cellular functions according to; The steps include: forming culture medium components to enhance or suppress a single basic cellular function j using formula (4) applied to the j-th column of the functional data array; The steps include: forming the culture medium components and manipulating cellular metabolism by increasing or suppressing a critical set of basic cellular functions using formula (4), which is simultaneously applied to multiple columns of the functional data array; The method described in any one of the above items, including: (Item 44) The method according to any one of the above items, wherein the target biological structure is a cell tissue, a whole cell, an organelle, or a coherent set of biochemical transformations exhibiting a predetermined cellular function. (Item 45) The method according to any one of the above items, wherein the target biological structure is genetically modified, and the modification includes a gene modification directed toward the activation or suppression of the basic cellular function. . (Item 46) A method characterized in that the medium factor is a physicochemical property of a mixture of solid and / or liquid and / or gas of essential nutrients and / or micronutrients and / or biological function molecules, the release rate of the compound, or the ingestion rate of the compound. (Item 47) The method according to any one of the above items, wherein the physicochemical property is temperature, and / or pressure, and / or pH, and / or ionic strength, and / or concentration, and / or activity, and / or osmolarity, and / or molality, and / or related properties. (Item 48) Essential nutrients and / or micronutrients and / or biological function molecules are salts, and / or vitamins, and / or cofactors of metabolism, and / or antibiotics, and / or carbohydrates, and / or lipid substances, and / or proteinaceous substances, and / or nucleotide substances, and / or signal transduction proteins, and / or molecules that suppress the activity of enzymes, and / or active molecules of proteins, and / or gene transformation modulators, and / or interfering ribonucleic acids, and / or complex mixtures of the above substances with known or unknown compositions containing serum, pure hydrogelate, or complex organic substances, which are inorganic substances and / or organic substances, and the method according to any one of the above items. (Item 49) The formula of the culture medium is Formula of culture medium = {FAC j}, j = 1, ···, N (where FAC j is the value of the medium factor j.), and the method according to any one of the above items, which is determined by the values of N medium factors. (Item 50) The target biological structure is Target biological structure = {e i}, i = 1, ···, K (where e i is a vector of q elements, and the values of the elements represent the weighted coefficients of each biochemical reaction in the basic cell function i.), and the method according to any one of the above items, which is determined by q biochemical reactions and K basic cell functions. (Item 51) The analysis includes classification and / or The above derivation includes a reference to a database (including the learning results). The method described in any one of the above items. (Item 52) A method for providing appropriate culture medium components and / or cell culture conditions for cells provided by a user using a computer, the method being: The process of inputting cell information provided by the user into the computer, The process involves having the computer refer to a database containing information about the cells and derive appropriate culture medium components and culture conditions for the cells. If necessary, the process involves displaying the results of the calculation on a display. A method of including. (Item 53) Includes recording the derived appropriate culture medium components and culture conditions in the database, Furthermore, when a query for appropriate culture medium components and / or cell culture conditions for the cells is received, the appropriate culture medium components and / or cell culture conditions recorded in the database are retrieved. If necessary, the process involves displaying the result of the aforementioned call on the display. The method described in any one of the above items, including: (Item 54) A program that codes a method for causing a computer to perform the selection of appropriate culture medium components and / or cell culture conditions for cells provided by a user, the method being: The process of inputting cell information provided by the user into the computer, The process involves having the computer, as necessary, consult a database to derive culture medium components and culture conditions related to the cells, and, if necessary, optimal conditions. The process of having the computer calculate the optimal culture medium components and cell culture conditions for the cells based on the culture medium components and the culture conditions, If necessary, the process involves displaying the results of the calculation on a display. A program that encompasses all of these. (Item 54A1) A program that codes a method for causing a computer to perform the selection of appropriate culture medium components and / or cell culture conditions for cells provided by a user, the method being: The process of inputting cell information provided by the user into the computer, The process of inputting data on the culture results under specific culture conditions for the cells into the computer, The process of inputting desired output conditions for the cells to the computer, The process involves having the computer, if necessary, consult a database to derive culture medium components and culture conditions related to the cells, as well as optimal conditions if necessary. The process of having the computer calculate the optimal culture medium components and cell culture conditions for the cells based on the culture medium components and the culture conditions, If necessary, the process includes displaying the results of the calculation on a display, If necessary, the process includes documenting the results of the calculations, If necessary, the process of outputting the documented document and A program that includes this. (Item 54A2) The documentation is the program described in any one of the above items, which is documented in accordance with the standard. (Item 54A3) The above standard includes the program described in any one of the above items, including the standard for the Original Manufacturing Record (MBR) or the Standard Operating Procedure (SOP). (Item 54A4) The optimal culture medium components and cell culture conditions for the cells are components and conditions suitable for both small-scale and commercial-scale, or either one of them, or a program described in any one of the above items that includes stepwise culture conditions from small-scale to commercial-scale. (Item 54A5) A program according to any one of the above items, further comprising the steps of extracting predicted risks with respect to the optimal culture medium components and cell culture conditions for the cells, formulating countermeasures for the risks, and providing information on the risks and countermeasures. (Item 54A6) The program described in any one of the above items, wherein the database includes at least one selected from the group consisting of cell information, specifications of the automated culture device, the number of people required depending on the culture conditions, the layout of the manufacturing plant, information on the wiring during culture, and information on the reagents held. (Item 54A7) The optimal culture medium components and cell culture conditions for the cells are the program described in any one of the above items, including components and culture conditions using an automated culture device. (Item 54A8) The cell culture conditions are the program described in any one of the above items, including information on manufacturing costs. (Item 54A9) The program described in any one of the above items, wherein the risks include at least one selected from the group consisting of risks of changes in the state and / or quantity of cultured cells during scale-up, risks of non-compliance with GMP, risks of malfunction in reagent preparation during culture, risks in the preparation of culture medium components including powder preparation and on-demand preparation, risks of quality and lot variability in culture medium, and regional risks. (Item 54A10) The countermeasure is a program described in any one of the above items, which is output based on the information in the database. (Item 54A11) The measures described above include at least one selected from the group consisting of suggesting compatible culture medium factors, suggesting methods for managing and supplying culture medium components, automatically generating risk reduction documents, suggesting routine responses that tolerate biogroups, and alternative culture methods, as described in any one of the above items. (Item 54A12) The document is a program described in any one of the above items, including a proposed initial culture method up to commercial production. (Item 54A13) The program described in any one of the above items, wherein the document includes providing information on cell production volume, production efficiency, cost of culture medium components, and composition of culture medium components, and providing at least one selected from the group consisting of proposals for changes to culture medium components, proposals for culture conditions, proposals for alternative methods, provision of MBR proposals, provision of SOP proposals, provision of visualized manufacturing site information, and provision of a roadmap. (Item 55) A recording medium storing a program that codes a method for causing a computer to perform the selection of appropriate culture medium components and / or cell culture conditions for cells provided by a user, wherein the method is The process of inputting cell information provided by the user into the computer, The process involves having the computer, if necessary, consult a database to derive culture medium components and culture conditions related to the cells, and, if necessary, optimal conditions. The process of having the computer calculate the optimal culture medium components and cell culture conditions for the cells based on the culture medium components and the culture conditions, If necessary, the process involves displaying the results of the calculation on a display. A recording medium that includes this. (Item 55A1) A recording medium storing a program that codes a method for causing a computer to perform the selection of appropriate culture medium components and / or cell culture conditions for cells provided by a user, wherein the method is The process of inputting cell information provided by the user into the computer, The process of inputting data on the culture results under specific culture conditions for the cells into the computer, The process of inputting desired output conditions for the cells to the computer, The process involves having the computer, if necessary, consult a database to derive culture medium components and culture conditions related to the cells, as well as optimal conditions if necessary. The process of having the computer calculate the optimal culture medium components and cell culture conditions for the cells based on the culture medium components and the culture conditions, If necessary, the process includes displaying the results of the calculation on a display, If necessary, the process includes documenting the results of the calculations, If necessary, the process of outputting the documented document and A recording medium that includes this. (Item 55A2) The documentation is a recording medium described in any one of the above items, which is documented in accordance with the standard. (Item 55A3) The above standard includes the recording medium specified in any one of the above items, including the standard for the Manufacturing Order Record (MBR) or the Standard Operating Procedure (SOP). (Item 55A4) The recording medium described in any one of the above items, wherein the optimal culture medium components and cell culture conditions for the cells include components and conditions suitable for both small scale and commercial scale or either one of them, or include culture conditions in a stepwise manner from small scale to commercial scale. (Item 55A5) A recording medium according to any one of the above items, further comprising the steps of extracting predicted risks with respect to the optimal culture medium components and cell culture conditions for the cells, formulating countermeasures against the risks, and providing information on the risks and countermeasures. (Item 55A6) The recording medium described in any one of the above items, wherein the database includes at least one selected from the group consisting of cell information, specifications of the automated culture device, the number of people required depending on the culture conditions, the layout of the manufacturing plant, information on the wiring during culture, and information on the reagents held. (Item 55A7) The recording medium described in any one of the above items, which includes the optimal culture medium components and cell culture conditions for the cells, and the components and culture conditions using an automated culture device. (Item 55A8) The cell culture conditions are recorded on a recording medium as described in any one of the above items, including information on the manufacturing cost. (Item 55A9) The recording medium described in any one of the above items, wherein the risks include at least one selected from the group consisting of risks of changes in the state and / or quantity of cultured cells during scale-up, risks of non-compliance with GMP, risks of malfunction in reagent preparation during culture, risks in preparing culture medium components including powder preparation and on-demand preparation, risks of quality and lot variability in culture medium, and regional risks. (Item 55A10) The countermeasure is a recording medium described in any one of the above items, which is output based on the information in the database. (Item 55A11) The recording medium described in any one of the above items, wherein the countermeasures include at least one selected from the group consisting of the proposal of compatible culture medium factors, the proposal of methods for managing and supplying culture medium components, the automatic generation of risk reduction documents, the proposal of routine responses that allow for biogroups, and alternative culture methods. (Item 55A12) The recording medium described in any one of the above items, including a proposed initial culture method up to commercial production. (Item 55A13) The recording medium described in any one of the above items, wherein the document includes at least one selected from the group consisting of providing information on cell production volume, production efficiency, cost of culture medium components, and composition of culture medium components, as well as proposals for changes to culture medium components, proposals for culture conditions, proposals for alternative methods, proposals for MBR, proposals for SOP, provision of visualized manufacturing site information, and proposals for roadmap provision. (Item 56) A system for selecting appropriate culture medium components and / or cell culture conditions for cells provided by a user, wherein the system: A cell information input unit that inputs cell information provided by the user, A culture medium component / culture condition deriving unit that, when necessary, consults a database to derive culture medium components and culture conditions related to the cells, and, if necessary, optimal conditions, An optimal calculation unit calculates the optimal culture medium components and cell culture conditions for the cells based on the culture medium components and the culture conditions, (If necessary) A display unit that displays the results of the calculation on a display. A system that encompasses all of these. (Item 56A1) A system that performs the selection of appropriate culture medium components and / or cell culture conditions for cells provided by a user, the system is: A cell information input unit that inputs cell information provided by the user, A culture result input unit for inputting culture result data under specific culture conditions for the aforementioned cells, An output condition input unit for inputting desired output conditions for the aforementioned cells, A culture medium component / culture condition deriving unit that, if necessary, consults a database to derive culture medium components and culture conditions related to the cells, and, if necessary, optimal conditions, An optimal calculation unit that calculates the optimal culture medium components and cell culture conditions for the cells based on the culture medium components and the culture conditions, A display unit that, if necessary, displays the results of the calculation on a display, A documentation unit that documents the results of the calculations as needed, If necessary, an output unit to output the documented document and A system that encompasses all of these. (Item 56A2) The documentation is a system described in any one of the above items, which is documented in accordance with the standard. (Item 56A3) The above standard includes the system described in any one of the above items, including the standard for the Original Manufacturing Record (MBR) or the Standard Operating Procedure (SOP). (Item 56A4) The optimal culture medium components and cell culture conditions for the cells are the system described in any one of the above items, which includes components and conditions suitable for both small-scale and commercial-scale, or for either one, or includes culture conditions in a stepwise manner from small-scale to commercial-scale. (Item 56A5) The system according to any one of the above items, further comprising the steps of extracting predicted risks with respect to the optimal culture medium components and cell culture conditions for the cells, formulating countermeasures for the risks, and providing information on the risks and countermeasures. (Item 56A6) The system described in any one of the above items, wherein the database includes at least one selected from the group consisting of cell information, specifications of the automated culture device, the number of people required depending on the culture conditions, the layout of the manufacturing plant, information on the wiring during culture, and information on the reagents held. (Item 56A7) The optimal culture medium components and cell culture conditions for the cells are the system described in any one of the above items, including components and culture conditions using an automated culture device. (Item 56A8) The cell culture conditions are the system described in any one of the above items, including information on manufacturing costs. (Item 56A9) The system described in any one of the above items, wherein the risks include at least one selected from the group consisting of risks of changes in the state and / or quantity of cultured cells during scale-up, risks of non-compliance with GMP, risks of malfunction in reagent preparation during culture, risks in the preparation of culture medium components including powder preparation and on-demand preparation, risks of quality and lot variability in the culture medium, and regional risks. (Item 56A10) The countermeasures described above are the systems described in any one of the above items, which are output based on the information in the database. (Item 56A11) The system described in any one of the above items, wherein the countermeasures include at least one selected from the group consisting of the suggestion of compatible culture medium factors, the suggestion of methods for managing and supplying culture medium components, the automatic generation of risk reduction documents, the suggestion of routine responses that tolerate biogroups, and alternative culture methods. (Item 56A12) The document is a system described in any one of the above items, including a proposed initial culture method up to commercial production. (Item 56A13) The system described in any one of the above items, wherein the document includes providing information on cell production volume, production efficiency, cost of culture medium components, and composition of culture medium components, as well as providing at least one selected from the group consisting of proposals for changes to culture medium components, proposals for culture conditions, proposals for alternative methods, proposals for MBR, proposals for SOP, provision of visualized production site information, and proposals for roadmap provision.

[0012] <Cost Calculation> (Item 57) A method for causing a computer to perform a calculation of the cost of culture medium components and / or cell culture conditions for cells provided by a user, the method being: The process of inputting cell information provided by the user into the computer, The process involves having the computer, if necessary, consult a database and calculate the culture medium components and culture conditions related to the cells, as well as the optimal conditions if necessary. The process of having the computer calculate the cost of the culture medium components and the cell culture conditions based on the culture medium components and the culture conditions, If necessary, the process of displaying the aforementioned costs on a display and A method of including. (Item 58) The above selection is (A) The step of selecting target cells; (B) The step of selecting K basic cellular functions of the cells; (C) A step of determining a set of N culture medium factors that determine the environment containing the components of the culture medium; (D) Using information stored in a database related to the cells, construct a functional map representing the intensity of activation or suppression of each of the K basic cellular functions by each of the N culture medium factors; (E) Using the functional map, the step of optimizing the composition of the cell culture medium so as to activate or suppress one or more of the basic cellular functions; The method described in any one of the above items, including: (Item 59) The method of any one of the above items, further comprising the features of any one of the above items.

[0013] (Wet) (Item 60) For cells provided by the user, media components and / or cell culture strips. A method for performing the calculation of the expenses in question, the method being: The process of analyzing cells provided by the user, The process involves deriving the culture medium components and culture conditions related to the cells from the aforementioned analysis results, A step of calculating the cost of the culture medium components and the cell culture conditions based on the culture medium components and the culture conditions. A method of including. (Item 61) The above selection is (A) The step of selecting target cells; (B) The step of selecting K basic cellular functions of the cells; (C) A step of determining a set of N culture medium factors that determine the environment containing the components of the culture medium; (D) Using experimental data of novel culture medium samples and / or the supernatant of existing culture medium samples using the components of the culture medium and the cells, a functional map is constructed that represents the intensity of activation or inhibition of each of the K basic cellular functions by each of the N culture medium factors. (E) Using the functional map, the step of optimizing the composition of the cell culture medium so as to activate or suppress one or more of the basic cellular functions; The method described in any one of the above items, including: (Item 62) The method according to any one of the above items, wherein the computer calculates the cost of the culture medium components and the cell culture conditions based on the culture medium components and the culture conditions, taking into consideration a database storing information relating to the costs and benefits of the culture medium and equipment used for culture.

[0014] <Other> (Item 63) A method for managing culture medium components and / or cell production with respect to cells provided by a user, 1) A step of providing information regarding appropriate culture medium components for the cells provided by the user, 2) A step of assigning identifiable labels to the cells and the culture medium components, 3) A step of checking whether the cells and the culture medium components are compatible by referring to the label. A method of including. (Item 64) The method according to any one of the above items, wherein the information relating to the culture medium components further comprises the characteristics described in any one or more of the above items. (Item 65) A method for causing a computer to perform the selection of appropriate culture medium components and / or cell culture conditions for cells provided by a user, the method being: The process of inputting cell information provided by the user into the computer, The process involves having the computer, if necessary, consult a database to derive culture medium components and culture conditions related to the cells, as well as optimal conditions if necessary. The process involves having the computer calculate the optimal culture medium components and cell culture conditions for the cells based on the culture medium components and the culture conditions. If necessary, the process involves displaying the results of the calculation on a display. A method of any one of the above items that further includes all or part of a method that includes a method. (Item 66) The above selection is (A) The step of selecting target cells; (B) The step of selecting K basic cellular functions of the cells; (C) A step of determining a set of N culture medium factors that determine the environment containing the components of the culture medium; (D) Using the information stored in the database related to the cells, the N culture factors The steps include constructing a functional map representing the intensity of activation or inhibition of each of the K basic cellular functions by each of the children; (E) Using the functional map, the step of optimizing the composition of the cell culture medium so as to activate or suppress one or more of the basic cellular functions; The method described in any one of the above items, including: (Item 67) The method according to any one of the above items, wherein the database is a library of cytokines, growth factors, proteins, small molecules, and bioactive substances useful for cell culture conditions, and can be used in the composition of the culture medium. (Item 68) The method according to any one of the above items, wherein the database is a library containing information on cell culture conditions that can be used to construct the culture medium. (Item 69) A method for performing calculations to select appropriate culture medium components and / or cell culture conditions for cells provided by a user, the method being: A step of analyzing cells provided by the user using candidate culture medium components and / or cell culture conditions, The process involves deriving the culture medium components and culture conditions (optimal requirements as needed) related to the cells from the analysis results, A step of calculating appropriate culture medium components and cell culture conditions for the cells based on the culture medium components and culture conditions. A method of any one of the above items that further includes all or part of a method that includes a method. (Item 70) The above selection is (A) The step of selecting target cells; (B) A step of selecting the basic cellular functions (K) of the cells; (C) A step of determining a set of medium factors (N) that determine the environment containing the components of the medium; (D) Using experimental data of novel culture medium samples and / or the supernatant of existing culture medium samples using the components of the culture medium and the cells, a functional map is constructed that represents the intensity of activation or inhibition of each of the K basic cellular functions by each of the N culture medium factors. (E) Using the functional map, the step of optimizing the composition of the cell culture medium so as to activate or suppress one or more of the basic cellular functions; The method described in any one of the above items, including: (Item 71) The method described in any one of the above items, wherein the candidate culture component is selected from the library of culture components. (Item 72) The candidate cell culture conditions are selected from the library of cell culture conditions, according to the method described in any one of the above items. (Item 73) The method according to any one of the above items, wherein the analysis is performed based on the results of an actual experiment using the culture medium components. (Item 74) The method according to any one of the above items, wherein the analysis is performed based on the results of an actual experiment using the culture medium components and calculations based on those results. (Item 75) The method according to any one of the above items, wherein the analysis comprises applying a library of the culture medium components to cells in a multi-well plate and performing high-throughput screening using an automated instrument. (Item 76) The method according to any one of the above items, wherein the calculation is performed using a library parameterized for the performance and / or cost of the culture component to the cells. (Item 77) The method according to any one of the above items, wherein optimization of the culture medium is achieved to suit the purpose of seed cells or cell lines, cost, and growth performance. (Item 78) The construction of the functional map in step d) above is The first culture experiment includes a number of cultures equal to (N+1) or more, which is the number of culture factors plus 1. The steps include: performing a process in which each culture is carried out with different culture medium components, and screening for combinations of low-level and high-level media factors; For each culture experiment performed, the steps include obtaining initial and final biomass, product, and exometabolome data, or partial exometabolome data; Relative weighting coefficient λ j The steps include determining a subset of basal cellular functions with a value greater than zero by regression analysis of the exometabolome data or exometabolome data induced against culture medium component data using the following linear model, and

[0015]

number

[0016]

number

[0017] (where v is a vector representing the rate of change of the component of the exometabolome from which the element was measured, I i,j This is the activation intensity parameter of basic cellular function j by culture medium factor i, determined by regression analysis. A step of performing a second run of a culture experiment, the number of cultures being greater than or equal to the number of activated cell functions plus 1, wherein each culture is performed with a different culture medium component, the medium component having a predetermined intensity parameter value I such that low and high values ​​of the weighted coefficient of activated basic cell function are identified as subsets of activated cell functions controlled by the medium factor. i,j The steps set up to screen using; In all of the above steps, a functional map is constructed from the data collected by linear regression analysis using equations (3a) and (3b), and the data is organized into the form of the functional map, wherein the intensity value I determined by the first run of the experiment is i,j However, this is corrected by the data from the second run of the experiment, resulting in an N×K data array (functional map = {I i,j The step of making it into the shape of}); The method described in any one of the above items, including:

[0018] (Item 79) The optimization of the culture medium components in step e) above is A step of forming basic cell function special culture medium components using the matrix of functional data, wherein the change in the value of the culture medium factor Δ(FAC j )

[0019]

number

[0020] The steps include: determining the change in the relative weights Δλi of basic cellular functions according to; The steps include: forming culture medium components to enhance or suppress a single basic cellular function j using formula (4) applied to the j-th column of the functional data array; By forming the culture medium components and using formula (4), which is simultaneously applied to multiple columns of the functional data array, the critical set of basic cellular functions can be enhanced or suppressed. and steps that manipulate cellular metabolism; The method described in any one of the above items, including: (Item 80) The method according to any one of the above items, wherein the target biological structure is a cell tissue, a whole cell, an organelle, or a coherent set of biochemical transformations exhibiting a predetermined cellular function. (Item 81) The method according to any one of the above items, wherein the target biological structure is genetically modified, and the modification includes a gene modification directed toward the activation or suppression of the basic cellular function. (Item 82) A method characterized in that the culture medium factor is the physicochemical properties of a mixture of solids and / or liquids and / or gases of essential nutrients and / or micronutrients and / or biologically functional molecules, the release rate of the compound, or the feeding rate of the compound. (Item 83) The method according to any one of the above items, wherein the physicochemical property is temperature and / or pressure and / or pH and / or ionic strength and / or concentration and / or activity and / or volume molar osmotic concentration and / or gravimetric osmotic concentration and / or related properties. (Item 84) Essential nutrients and / or micronutrients and / or biologically functional molecules are The method according to any one of the above items, wherein the inorganic and / or organic substance is a salt, and / or vitamin, and / or metabolic cofactor, and / or antibiotic, and / or carbohydrate, and / or lipid substance, and / or protein substance, and / or nucleotide substance, and / or signaling protein, and / or molecule that inhibits enzyme activity, and / or protein active molecule, and / or gene transformation modulator, and / or interfering ribonucleic acid, and / or a complex mixture of the above substance and a known or unknown composition comprising serum, pure hydrosilate, or complex organic substance. (Item 85) The formula for the culture medium is: Culture medium formula = {FAC j},j=1,···,N(however, FAC jThe method according to any one of the above items, determined by the values ​​of N culture factors, using the value of the culture factor j. (Item 86) The target biological structure is, target biological structure={e i},i=1,···,K(where e i The method according to any one of the above items, wherein is a vector of q elements, and the values ​​of the elements represent weighted coefficients for each biochemical reaction in basic cellular function i. (Item 87) The analysis includes classification and / or The above derivation includes a reference to a database (including the learning results). The method described in any one of the above items. (Item 88) A method for providing appropriate culture medium components and / or cell culture conditions for cells provided by a user using a computer, the method being: The process of inputting cell information provided by the user into the computer, The process involves having the computer refer to a database containing information about the cells and derive appropriate culture medium components and culture conditions for the cells. If necessary, the process involves displaying the results of the calculation on a display. The method described in any one of the above items, including all or part of the method of including. (Item 89) Includes recording the derived appropriate culture medium components and culture conditions in the database, Furthermore, when a query for appropriate culture medium components and / or cell culture conditions for the cells is received, the appropriate culture medium components and / or cell culture conditions recorded in the database are retrieved. If necessary, the process involves displaying the result of the aforementioned call on the display. The method described in any one of the above items, including the method described in the above items. (Item 90) A program that codes a method for causing a computer to perform the selection of appropriate culture medium components and / or cell culture conditions for cells provided by a user, the method being: The process of inputting cell information provided by the user into the computer, The process involves having the computer, if necessary, consult a database to derive culture medium components and culture conditions related to the cells, as well as optimal conditions if necessary. The process involves having the computer calculate the optimal culture medium components and cell culture conditions for the cells based on the culture medium components and the culture conditions. If necessary, the process involves displaying the results of the calculation on a display. A program that encompasses all of these. (Item 91) A recording medium storing a program that codes a method for causing a computer to perform the selection of appropriate culture medium components and / or cell culture conditions for cells provided by a user, wherein the method is The process of inputting cell information provided by the user into the computer, The process involves having the computer, if necessary, consult a database to derive culture medium components and culture conditions related to the cells, as well as optimal conditions if necessary. The process involves having the computer calculate the optimal culture medium components and cell culture conditions for the cells based on the culture medium components and the culture conditions. If necessary, the process involves displaying the results of the calculation on a display. A recording medium that includes this. (Item 92) A system for selecting appropriate culture medium components and / or cell culture conditions for cells provided by a user, the system is A cell information input unit that inputs cell information provided by the user, A culture medium component / culture condition deriving unit that, when necessary, consults a database to derive culture medium components and culture conditions related to the cells, and, if necessary, optimal conditions, An optimal calculation unit calculates the optimal culture medium components and cell culture conditions for the cells based on the culture medium components and the culture conditions. If necessary, a display unit that displays the results of the calculation on a display. A system that encompasses all of these. (Item 93) A method for causing a computer to perform a calculation of the cost of culture medium components and / or cell culture conditions for cells provided by a user, the method being: The process of inputting cell information provided by the user into the computer, The process involves having the computer, if necessary, consult a database and calculate the culture medium components and culture conditions related to the cells, as well as the optimal conditions if necessary. A step of causing the computer to calculate the cost of the culture medium components and the cell culture conditions based on the culture medium components and the culture conditions. If necessary, the process of displaying the aforementioned costs on a display and A method of including. (Item 94) The above selection is (A) The step of selecting target cells; (B) The step of selecting K basic cellular functions of the cells; (C) A step of determining a set of N culture medium factors that determine the environment containing the components of the culture medium; (D) Using information stored in a database related to the cells, construct a functional map representing the intensity of activation or suppression of each of the K basic cellular functions by each of the N culture medium factors; (E) Using the functional map, the step of optimizing the composition of the cell culture medium so as to activate or suppress one or more of the basic cellular functions; The method described in any one of the above items, including: (Item 95) The method of any one of the above items, further comprising the features of any one of the above items.

[0021] (Wet) (Item 96) A method for performing a calculation of the cost of culture medium components and / or cell culture conditions for cells provided by a user, the method is: The process of analyzing cells provided by the user, The process involves deriving the culture medium components and culture conditions related to the cells from the aforementioned analysis results, A step of calculating the cost of the culture medium components and the cell culture conditions based on the culture medium components and the culture conditions. A method of including. (Item 97) The above selection is (A) The step of selecting target cells; (B) The step of selecting K basic cellular functions of the cells; (C) A step of determining a set of N culture medium factors that determine the environment containing the components of the culture medium; (D) Using experimental data of novel culture medium samples and / or the supernatant of existing culture medium samples using the components of the culture medium and the cells, a functional map is constructed that represents the intensity of activation or inhibition of each of the K basic cellular functions by each of the N culture medium factors. (E) Using the functional map, the step of optimizing the composition of the cell culture medium so as to activate or suppress one or more of the basic cellular functions; The method described in any one of the above items, including: (Item 98) The method according to any one of the above items, wherein the computer calculates the cost of the culture medium components and the cell culture conditions based on the culture medium components and the culture conditions, taking into consideration a database storing information relating to the costs and benefits of the culture medium and equipment used for culture. (Item 99) A method for causing a computer to manage culture medium components and / or cell production for cells provided by a user, 1) A step of providing information regarding appropriate culture medium components for the cells provided by the user, 2) A step of assigning identifiable labels to the cells and the culture medium components, 3) A step of checking whether the cells and the culture medium components are compatible by referring to the label. A method of including. (Item 100) The method according to any one of the above items, characterized in that the label is contained in the culture medium. (Item 101) The method according to any one of the above items, characterized in that the label is contained outside the culture medium (for example, in a container containing the culture medium). (Item 102) A method for causing a computer to perform the selection of appropriate culture medium components and / or cell culture conditions for cells provided by a user, the method being: The process of inputting cell information provided by the user into the computer, The process involves having the computer, if necessary, consult a database to derive culture medium components and culture conditions related to the cells, as well as optimal conditions if necessary. The process involves having the computer calculate the optimal culture medium components and cell culture conditions for the cells based on the culture medium components and the culture conditions. If necessary, the process involves displaying the results of the calculation on a display. A method of any one of the above items that further includes all or part of a method that includes a method. (Item 103) The above selection is (A) The step of selecting target cells; (B) The step of selecting K basic cellular functions of the cells; (C) A step of determining a set of N culture medium factors that determine the environment containing the components of the culture medium; (D) Using information stored in a database related to the cells, construct a functional map representing the intensity of activation or suppression of each of the K basic cellular functions by each of the N culture medium factors; (E) Using the functional map, the step of optimizing the composition of the cell culture medium so as to activate or suppress one or more of the basic cellular functions; The method described in any one of the above items, including: (Item 104) The method according to any one of the above items, wherein the database is a library of cytokines, growth factors, proteins, small molecules, and physiologically active substances useful for cell culture conditions, and can be used in the composition of the culture medium. (Item 105) The method according to any one of the above items, wherein the database is a library containing information on cell culture conditions that can be used to compose the culture medium. (Item 106) A method for performing calculations to select appropriate culture medium components and / or cell culture conditions for cells provided by a user, the method being: A step of analyzing cells provided by the user using candidate culture medium components and / or cell culture conditions, The process involves deriving the culture medium components and culture conditions (optimal requirements as needed) related to the cells from the analysis results, A step of calculating appropriate culture medium components and cell culture conditions for the cells based on the culture medium components and culture conditions. A method of any one of the above items that further includes all or part of a method that includes a method. (Item 107) The above selection is (A) The step of selecting target cells; (B) The step of selecting K basic cellular functions of the cells; (C) A step of determining a set of N culture medium factors that determine the environment containing the components of the culture medium; (D) Using experimental data of novel culture medium samples and / or the supernatant of existing culture medium samples using the components of the culture medium and the cells, a functional map is constructed that represents the intensity of activation or inhibition of each of the K basic cellular functions by each of the N culture medium factors. (E) Using the functional map, the step of optimizing the composition of the cell culture medium so as to activate or suppress one or more of the basic cellular functions; The method described in any one of the above items, including: (Item 108) The method described in any one of the above items, wherein the candidate culture component is selected from the library of culture components. (Item 109) The candidate cell culture conditions are selected from the library of cell culture conditions, according to the method described in any one of the above items. (Item 110) The method according to any one of the above items, wherein the analysis is performed based on the results of an actual experiment using the culture medium components. (Item 111) The method according to any one of the above items, wherein the analysis is performed based on the results of an actual experiment using the culture medium components and calculations based on those results. (Item 112) The method according to any one of the above items, wherein the analysis comprises applying a library of the culture medium components to cells in a multiwell plate and performing high-throughput screening using an automated instrument. (Item 113) The calculations described above include any one of the above items, which include calculating using a library parameterized for the performance and / or cost of the culture components on cells. The method described in section [section number]. (Item 114) The method according to any one of the above items, wherein optimization of the culture medium is achieved to suit the purpose of seed cells or cell lines, cost, and growth performance. (Item 115) The construction of the functional map in step d) above is A step of performing a first run of a culture experiment, comprising a number of cultures equal to (N+1) or more obtained by adding 1 to the number of culture medium factors, wherein each culture is performed with a different culture medium component, and combinations of low-level and high-level culture medium factors are screened; For each culture experiment performed, the steps include obtaining initial and final biomass, product, and exometabolome data, or partial exometabolome data; Relative weighting coefficient λ j The steps include determining a subset of basal cellular functions with a value greater than zero by regression analysis of the exometabolome data or exometabolome data induced against culture medium component data using the following linear model, and

[0022]

number

[0023]

number

[0024] (where v is a vector representing the rate of change of the component of the exometabolome from which the element was measured, I i,j This is the activation intensity parameter of basic cellular function j by culture medium factor i, determined by regression analysis. A step of performing a second run of a culture experiment, the number of cultures being greater than or equal to the number of activated cell functions plus 1, wherein each culture is performed with a different culture medium component, the medium component having a predetermined intensity parameter value I such that low and high values ​​of the weighted coefficient of activated basic cell function are identified as subsets of activated cell functions controlled by the medium factor. i,j The steps set up to screen using; In all of the above steps, a functional map is constructed from the data collected by linear regression analysis using equations (3a) and (3b), and the data is organized into the form of the functional map, wherein the intensity value I determined by the first run of the experiment is i,j However, this is corrected by the data from the second run of the experiment, resulting in an N×K data array (functional map = {I i,j The step of making it into the shape of}); The method described in any one of the above items, including: (Item 116) The optimization of the culture medium components in step e) above is A step of forming basic cell function special culture medium components using the matrix of functional data, wherein the change in the value of the culture medium factor Δ(FAC j )

[0025]

number

[0026] The steps include: determining the change in the relative weights Δλi of basic cellular functions according to; The steps include: forming culture medium components to enhance or suppress a single basic cellular function using formula (4) applied to the j-th column of the functional data array; The steps include: forming the culture medium components and manipulating cellular metabolism by increasing or suppressing a critical set of basic cellular functions using formula (4), which is simultaneously applied to multiple columns of the functional data array; The method described in any one of the above items, including: (Item 117) The method according to any one of the above items, wherein the target biological structure is a cell tissue, a whole cell, an organelle, or a coherent set of biochemical transformations exhibiting a predetermined cellular function. (Item 118) The method according to any one of the above items, wherein the target biological structure is genetically modified, and the modification includes a gene modification directed toward the activation or suppression of the basic cellular function. (Item 119) A method characterized in that the culture medium factor is the physicochemical properties of a mixture of solid and / or liquid and / or gaseous essential nutrients and / or micronutrients and / or biologically functional molecules, the release rate of the compound, or the feeding rate of the compound. (Item 120) The method according to any one of the above items, wherein the physicochemical properties are temperature, and / or pressure, and / or pH, and / or ionic strength, and / or concentration, and / or activity, and / or volume molar osmotic concentration, and / or gravimetric osmotic concentration, and / or related properties. (Item 121) Essential nutrients and / or micronutrients and / or biologically functional molecules are The method according to any one of the above items, wherein the inorganic and / or organic substance is a salt, and / or vitamin, and / or metabolic cofactor, and / or antibiotic, and / or carbohydrate, and / or lipid substance, and / or protein substance, and / or nucleotide substance, and / or signaling protein, and / or molecule that inhibits enzyme activity, and / or protein active molecule, and / or gene transformation modulator, and / or interfering ribonucleic acid, and / or a complex mixture of the above substance and a known or unknown composition comprising serum, pure hydrosilate, or complex organic substance. (Item 122) The formula for the culture medium is, Culture medium formula = {FAC j},j=1,···,N(however, FAC j The method according to any one of the above items, determined by the values ​​of N culture factors, using the value of the culture factor j. (Item 123) The target biological structure is, target biological structure={e i},i=1,···,K(where e i The method according to any one of the above items, wherein is a vector of q elements, and the values ​​of the elements represent weighted coefficients for each biochemical reaction in basic cellular function i. (Item 124) The analysis includes classification, and / or the derivation includes a reference to a database (including the learning results). The method described in any one of the above items. (Item 125) A method for providing appropriate culture medium components and / or cell culture conditions for cells provided by a user using a computer, the method being: The process of inputting cell information provided by the user into the computer, The process involves having the computer refer to a database containing information about the cells and derive appropriate culture medium components and culture conditions for the cells. If necessary, the process involves displaying the results of the calculation on a display. The method described in any one of the above items, including all or part of the method of including. (Item 126) Record the derived appropriate culture medium components and culture conditions in the database. This includes doing Furthermore, when a query for appropriate culture medium components and / or cell culture conditions for the cells is received, the appropriate culture medium components and / or cell culture conditions recorded in the database are retrieved. If necessary, the process involves displaying the result of the aforementioned call on the display. The method described in any one of the above items, including the method described in the above items. (Item 127) A program that codes a method for causing a computer to perform the selection of appropriate culture medium components and / or cell culture conditions for cells provided by a user, the method being: The process of inputting cell information provided by the user into the computer, The process involves having the computer, if necessary, consult a database to derive culture medium components and culture conditions related to the cells, as well as optimal conditions if necessary. The process involves having the computer calculate the optimal culture medium components and cell culture conditions for the cells based on the culture medium components and the culture conditions. If necessary, the process involves displaying the results of the calculation on a display. A program that encompasses all of these. (Item 128) A recording medium storing a program that codes a method for causing a computer to perform the selection of appropriate culture medium components and / or cell culture conditions for cells provided by a user, wherein the method is The process of inputting cell information provided by the user into the computer, The process involves having the computer, if necessary, consult a database to derive culture medium components and culture conditions related to the cells, as well as optimal conditions if necessary. The process involves having the computer calculate the optimal culture medium components and cell culture conditions for the cells based on the culture medium components and the culture conditions. If necessary, the process involves displaying the results of the calculation on a display. A recording medium that includes this. (Item 129) A system for selecting appropriate culture medium components and / or cell culture conditions for cells provided by a user, wherein the system: A cell information input unit that inputs cell information provided by the user, A culture medium component / culture condition deriving unit that, when necessary, consults a database to derive culture medium components and culture conditions related to the cells, and, if necessary, optimal conditions, An optimal calculation unit calculates the optimal culture medium components and cell culture conditions for the cells based on the culture medium components and the culture conditions. If necessary, a display unit that displays the results of the calculation on a display. A system that encompasses all of these. (Item 130) A method described in any one of the above items, The aforementioned basic cellular functions are obtained from the biological network of the target biological structure. The biological network is divided into K functional subnetworks, which include a subset of biological trait transformations. A method characterized in that the subnetwork is obtained manually and / or automatically. (Item 131) A method described in any one of the above items, The aforementioned basic cellular functions are obtained from genome-scale reconstruction of the biological network of the target biological structure. A method characterized in that a working set of K basic cellular functions is pre-reduced using transcriptome data and / or proteome data and / or endo-metabolome data and / or thermodynamic data, when such data is available. (Item 132) A method described in any one of the above items, The aforementioned functional map includes shaking flasks, T-flasks, reactors, microplates, and more. A method characterized by determination by continuous and / or parallel culture experiments performed in an microbioreactor or phenotypic microarray. (Item 133) A method described in any one of the above items, The aforementioned functional map is, Chromatographic methods such as liquid chromatography (LC) and gas chromatography (GC), 1 H-NMR, 13 A method characterized by determination by an exometabolome assay that includes analysis of the supernatant of a novel culture medium sample or a used culture medium sample, using an NMR method such as 13C-NMR, mass spectrometry (MS), or a chromatography method combined with mass spectrometry such as GC-MS or LC-MS, or by a method combining the aforementioned measurement methods. (Item 134) A method described in any one of the above items, The reduced set of the aforementioned activated basic cellular functions was identified by linear regression analysis or nonlinear regression analysis. The variance or covariance of the exometabolome data or the derived exometabolome data is maximized. The correlation between exometabolome data or induced exometabolome data and the values ​​of culture medium factors is maximized. A method characterized in that the basic cellular functions are ranked according to their correlation with or sensitivity to the values ​​of the culture medium factors. (Item 135) A method described in any one of the above items, The aforementioned functional map is determined by a high-throughput automated system. And so, A method characterized by the connection of a culture device, an analytical exometabolome device, and a computer algorithm to a physical device, thereby obtaining a high-throughput functional map. (Item 136) A method described in any one of the above items, The target basic cellular function i associated with product quantity and / or product quality is relative to the weight Δ(λ) iA method characterized by increasing ) by 60% to 100%. (Item 137) A chemically defined culture medium formulation obtained by any of the methods described in any of the above items, a) an aqueous solution of any trace component; b) an aqueous solution a) and a mixture of any complementary basic aqueous solution, or other complementary basic aqueous solutions; A culture medium formulation characterized by being composed of the following. (Item 138) The aforementioned trace components are serum, various vitamins, various antibiotics, various hormones, various growth factors, and / or The aforementioned basic aqueous solution is, for example, (1) Trace components: Components selected from the group consisting of PTEN inhibitors, p53 inhibitors, p38 inhibitors, Wnt signaling activators, and ROCK inhibitors. Preferably, in order: VO-OH Pic, Pifithrin-a, SB203580, Lici, Y-27632. Basic aqueous solution: Basic aqueous solution containing DMEM / F-12 (2) Trace components: IL-1 and / or TNF-α Basic aqueous solution: Basic aqueous solution containing αMEM A culture medium formulation according to any one of the above items, characterized in that it is the same as the above item. (Item 138A) Using the method described in any one of the above items Tissues, and / or cells, and / or viruses, and / or cellular components, and / or substances related to proteins, and / or carbohydrates, and / or nucleotide substances, and / or lipid substances, and / or primary metabolites, and / or secondary metabolites or a method for increasing the quantity and / or quality of a mixture of products in a biological manufacturing process such as the production of biofuels, vaccines, drugs, biopolymers, or precursors thereof. (Item 139) Using the method described in any one of the above items, A method for optimizing the composition of cell culture media for cell lines of the plant kingdom or animal kingdom, or the composition of other eukaryotic unicellular or multicellular organisms such as yeast or fungi. (Item 140) Using the method described in any one of the above items, A method for optimizing the composition of cell culture media for prokaryotic organisms. (Item 141) Using the method described in any one of the above items, A method for optimizing cell culture conditions and / or the composition of the culture medium. (Item 142) Using the method described in any one of the above items, A method for identifying biomarkers specific to cell function or culture medium components. (Item 143) Using the method described in any one of the above items, A method for designing drugs or optimizing drug mixtures that are aimed at altering cellular functions associated with a disease state. (Item 144) A program that includes code that causes a computer to perform the identification of a biomarker using any of the methods described in any of the above items. (Item 145) A culture medium design system that implements the method described in any one of the above items.

[0027] <database> (In silico) (Item 146) A method for causing a computer to create a database of appropriate culture medium components and / or cell culture conditions for cells provided by a user, the method being: 1) The process of inputting cell information provided by the user into the computer, 2) A step of having the computer, taking into account the database as necessary, calculate the culture medium components and culture conditions related to the cells, and, if necessary, the optimal conditions. 3) A step of causing the computer to calculate the optimal culture medium components and cell culture conditions for the cells based on the culture medium components and the culture conditions. 4) A step of recording the results of the above calculation in a database. 5) A process that repeats steps 1) to 4) A method of including. (Item 147) The method of (Item 146), wherein the analysis is based on the results of experiments using novel culture medium samples and / or existing culture medium samples. (Item 148) The method according to any one of the above items, comprising determining whether the subject is a mesenchymal stem cell. (Item 149) The method of any one of the above items, further comprising the features of any one or more of the above items. (Item 150) A method for causing a computer to perform the selection of appropriate culture medium components and / or cell culture conditions for cells provided by a user, the method being: The process of inputting cell information provided by the user into the computer, The process involves having the computer, if necessary, consult a database to derive culture medium components and culture conditions related to the cells, as well as optimal conditions if necessary. The process involves having the computer calculate the optimal culture medium components and cell culture conditions for the cells based on the culture medium components and the culture conditions. If necessary, the process involves displaying the results of the calculation on a display. The method described in any one of the above items, further encompassing all or part of the method described above. (Item 151) The above selection is (A) The step of selecting target cells; (B) The step of selecting K basic cellular functions of the cells; (C) A step of determining a set of N culture medium factors that determine the environment containing the components of the culture medium; (D) Using information stored in a database related to the cells, construct a functional map representing the intensity of activation or suppression of each of the K basic cellular functions by each of the N culture medium factors; (E) Using the functional map, the step of optimizing the composition of the cell culture medium so as to activate or suppress one or more of the basic cellular functions; The method described in any one of the above items, including: (Item 152) The method according to any one of the above items, wherein the database is a library of cytokines, growth factors, proteins, small molecules, and bioactive substances useful for cell culture conditions, and can be used in the composition of the culture medium. (Item 153) The method according to any one of the above items, wherein the database is a library containing information on cell culture conditions that can be used to construct the culture medium. (Item 154) A method for performing calculations to select appropriate culture medium components and / or cell culture conditions for cells provided by a user, the method being: A step of analyzing cells provided by the user using candidate culture medium components and / or cell culture conditions, The process involves deriving the culture medium components and culture conditions (optimal requirements as needed) related to the cells from the analysis results, A step of calculating appropriate culture medium components and cell culture conditions for the cells based on the culture medium components and culture conditions. A method of any one of the above items that further includes all or part of a method that includes a method. (Item 155) The above selection is (A) The step of selecting target cells; (B) A step of selecting the basic cellular functions (K) of the cells; (C) A step of determining a set of medium factors (N) that determine the environment containing the components of the medium; (D) Using experimental data of novel culture medium samples and / or the supernatant of existing culture medium samples using the components of the culture medium and the cells, a functional map is constructed that represents the intensity of activation or inhibition of each of the K basic cellular functions by each of the N culture medium factors. (E) Using the functional map, the step of optimizing the composition of the cell culture medium so as to activate or suppress one or more of the basic cellular functions; The method described in any one of the above items, including: (Item 156) The method described in any one of the above items, wherein the candidate culture component is selected from the library of culture components. (Item 157) The candidate cell culture conditions are selected from the library of cell culture conditions, according to the method described in any one of the above items. (Item 158) The method according to any one of the above items, wherein the analysis is performed based on the results of an actual experiment using the culture medium components. (Item 159) The method according to any one of the above items, wherein the analysis is performed based on the results of an actual experiment using the culture medium components and calculations based on those results. (Item 160) The method according to any one of the above items, wherein the analysis comprises applying a library of the culture medium components to cells in a multiwell plate and performing high-throughput screening using an automated instrument. (Item 161) The method according to any one of the above items, wherein the calculation is performed using a library parameterized for the performance and / or cost of the culture component to the cells. (Item 162) The method according to any one of the above items, wherein optimization of the culture medium is achieved to suit the purpose of seed cells or cell lines, cost, and growth performance. (Item 163) The construction of the functional map in step d) above is A step of performing a first run of a culture experiment, comprising a number of cultures equal to (N+1) or more obtained by adding 1 to the number of culture medium factors, wherein each culture is performed with a different culture medium component, and combinations of low-level and high-level culture medium factors are screened; For each culture experiment performed, the steps include obtaining initial and final biomass, product, and exometabolome data, or partial exometabolome data; Relative weighting coefficient λj The steps include determining a subset of basal cellular functions with a value greater than zero by regression analysis of the exometabolome data or exometabolome data induced against culture medium component data using the following linear model, and

[0028]

number

[0029]

number

[0030] (where v is a vector representing the rate of change of the component of the exometabolome from which the element was measured, I i,j This is the activation intensity parameter of basic cellular function j by culture medium factor i, determined by regression analysis. A step of performing a second run of a culture experiment, the number of cultures being greater than or equal to the number of activated cell functions plus 1, wherein each culture is performed with a different culture medium component, the medium component having a predetermined intensity parameter value I such that low and high values ​​of the weighted coefficient of activated basic cell function are identified as subsets of activated cell functions controlled by the medium factor. i,j The steps set up to screen using; In all of the above steps, a functional map is constructed from the data collected by linear regression analysis using equations (3a) and (3b), and the data is organized into the form of the functional map, wherein the intensity value I determined by the first run of the experiment is i,j However, this is corrected by the data from the second run of the experiment, resulting in an N×K data array (functional map = {I i,j The step of making it into the shape of}); The method described in any one of the above items, including: (Item 164) The optimization of the culture medium components in step e) above is A step of forming basic cell function special culture medium components using the matrix of functional data, wherein the change in the value of the culture medium factor Δ(FAC j )

[0031]

number

[0032] The steps include: determining the change in the relative weights Δλi of basic cellular functions according to; Using formula (4) applied to the j-th column of the aforementioned functional data array, a single basic detail The steps include forming culture medium components to enhance or inhibit cellular function; The steps include: forming the culture medium components and manipulating cellular metabolism by increasing or suppressing a critical set of basic cellular functions using formula (4), which is simultaneously applied to multiple columns of the functional data array; The method described in any one of the above items, including: (Item 165) The method according to any one of the above items, wherein the target biological structure is a cell tissue, a whole cell, an organelle, or a coherent set of biochemical transformations exhibiting a predetermined cellular function. (Item 166) The method according to any one of the above items, wherein the target biological structure is genetically modified, and the modification includes a gene modification directed toward the activation or suppression of the basic cellular function. (Item 167) A method characterized in that the culture medium factor is the physicochemical properties of a mixture of solid and / or liquid and / or gaseous essential nutrients and / or micronutrients and / or biologically functional molecules, the release rate of the compound, or the feeding rate of the compound. (Item 168) The method according to any one of the above items, wherein the physicochemical properties are temperature, and / or pressure, and / or pH, and / or ionic strength, and / or concentration, and / or activity, and / or volume molar osmotic concentration, and / or gravimetric osmotic concentration, and / or related properties. (Item 169) Essential nutrients and / or micronutrients and / or biologically functional molecules are The method according to any one of the above items, wherein the inorganic and / or organic substance is a salt, and / or vitamin, and / or metabolic cofactor, and / or antibiotic, and / or carbohydrate, and / or lipid substance, and / or protein substance, and / or nucleotide substance, and / or signaling protein, and / or molecule that inhibits enzyme activity, and / or protein active molecule, and / or gene transformation modulator, and / or interfering ribonucleic acid, and / or a complex mixture of the above substance and a known or unknown composition comprising serum, pure hydrosilate, or complex organic substance. (Item 170) The formula for the culture medium is: Culture medium formula = {FAC j},j=1,···,N(however, FAC j The method according to any one of the above items, determined by the values ​​of N culture factors, using the value of the culture factor j. (Item 171) The target biological structure is target biological structure={e i},i=1,···,K(where e i The method according to any one of the above items, wherein is a vector of q elements, and the values ​​of the elements represent weighted coefficients for each biochemical reaction in basic cellular function i. (Item 172) The analysis includes classification and / or The above derivation includes a reference to a database (including the learning results). The method described in any one of the above items. (Item 173) A method for providing appropriate culture medium components and / or cell culture conditions for cells provided by a user using a computer, the method being: The process of inputting cell information provided by the user into the computer, The process involves having the computer refer to a database containing information about the cells and derive appropriate culture medium components and culture conditions for the cells. If necessary, the process involves displaying the results of the calculation on a display. The method described in any one of the above items, including all or part of the method of including. (Item 174) Includes recording the derived appropriate culture medium components and culture conditions in the database, Furthermore, when a query for appropriate culture medium components and / or cell culture conditions for the cells is received, the appropriate culture medium components and / or cell culture conditions recorded in the database are retrieved. If necessary, the process involves displaying the result of the aforementioned call on the display. The method described in any one of the above items, including the method described in the above items. (Item 175) A program that codes a method for causing a computer to perform the selection of appropriate culture medium components and / or cell culture conditions for cells provided by a user, the method being: The process of inputting cell information provided by the user into the computer, The process involves having the computer, if necessary, consult a database to derive culture medium components and culture conditions related to the cells, as well as optimal conditions if necessary. The process involves having the computer calculate the optimal culture medium components and cell culture conditions for the cells based on the culture medium components and the culture conditions. If necessary, the process involves displaying the results of the calculation on a display. A program that encompasses all of these. (Item 176) A recording medium storing a program that codes a method for causing a computer to perform the selection of appropriate culture medium components and / or cell culture conditions for cells provided by a user, wherein the method is The process of inputting cell information provided by the user into the computer, A step of causing the computer to refer to a database as necessary, derive medium components and culture conditions related to the cells, and derive optimal conditions as necessary; A step of causing the computer to calculate optimal medium components and cell culture conditions for the cells based on the medium components and the culture conditions; A step of causing the result of the calculation to be displayed on a display as necessary; A recording medium including the above. (Item 177) A system for selecting appropriate medium components and / or cell culture conditions for cells provided by a user, the system comprising: A cell information input unit for inputting information on the cells provided by the user; A medium component / culture condition derivation unit for referring to a database as necessary, deriving medium components and culture conditions related to the cells, and deriving optimal conditions as necessary; An optimal calculation unit for calculating optimal medium components and cell culture conditions for the cells based on the medium components and the culture conditions; A display unit for causing the result of the calculation to be displayed on a display as necessary; A system including the above. (Item 177A) A method for selecting appropriate medium components and / or cell culture conditions for cells provided by a user, the method comprising: A step of inputting information on the cells provided by the user; A step of referring to a database as necessary, deriving medium components and culture conditions related to the cells, and deriving optimal conditions as necessary; A step of calculating optimal medium components and cell culture conditions for the cells based on the medium components and the culture conditions; A step of causing the result of the calculation to be displayed on a display as necessary; A method including the above. (Item 177B) A program for causing a computer to execute a method for selecting appropriate medium components and / or cell culture conditions for cells provided by a user, the method comprising: The process of inputting cell information provided by the user, The process involves, if necessary, consulting a database to derive the culture medium components and culture conditions related to the cells, and, if necessary, the optimal conditions. Based on the culture medium components and culture conditions, the optimal culture medium components and cells for the cells The process of calculating the culture conditions and If necessary, the process involves displaying the results of the calculation on a display. A program that encompasses all of these. (Item 177C) A recording medium that stores a program causing a computer to perform a method for selecting appropriate culture medium components and / or cell culture conditions for cells provided by a user, wherein the method is: The process of inputting cell information provided by the user, The process involves, if necessary, consulting a database to derive the culture medium components and culture conditions related to the cells, and, if necessary, the optimal conditions. A step of calculating the optimal culture medium components and cell culture conditions for the cells based on the culture medium components and the culture conditions. If necessary, the process involves displaying the results of the calculation on a display. A recording medium that includes this.

[0033] <Automation> (Item 178) A system for automating culture, 1) A cell donor unit that provides cells intended for culture, 2) A cell information providing unit that provides information about the cells as needed. 3) A culture medium information providing unit that provides information about the culture medium or culture medium components as needed. 4) A medium matching section for matching cells and culture medium as needed, 5) A culture medium supply unit that provides culture medium, 6) A cell culture section for culturing cells, 7) A cell culture condition adjustment unit for adjusting cell conditions, 8) Cell culture control unit for controlling and automating cell culture A system comprising the above, wherein the culture medium is optimized for the cells. (Item 179) The selection is made by the method described in any one of the above items, (Item 178) the system described in any one of the above items. (Item 180) A system for automating culture, 1) A cell donor unit that provides cells intended for culture, 2) A culture medium compatibility information storage unit that stores information regarding the compatibility of the cells and the culture medium, 5) A culture medium supply unit that provides culture medium, 6) A cell culture section for culturing cells, 7) A cell culture condition adjustment unit for adjusting cell conditions, 8) Cell culture control unit for controlling and automating cell culture A system comprising the above, wherein the culture medium is optimized for the cells. (Item 181) The system according to any one of the above items, further comprising a monitoring unit for monitoring the cells and / or components in the culture medium. (Item 181A1) The monitoring unit is a system according to any one of the above items, including a sensor that is in contact with or not in contact with the culture vessel or culture channel. (Item 181A2) The sensor is a system as described in any one of the above items, including an optical sensor, an electrical sensor, a magnetic sensor, a chemical sensor, an acoustic sensor, a microfluidic sensor, or a biosensor. (Item 181A3) Furthermore, the system according to any one of the above items further includes a sensor data storage unit for storing data obtained by the sensor, and at least one, preferably both, of an analysis and extraction unit for analyzing the data and extracting stable data and unstable data. (Item 181A4) The analysis and extraction unit is configured to continuously monitor unstable data, as described in any one of the above items. (Item 182) The system according to any one of the above items, wherein the cell culture condition adjustment unit can add or change the components or the culture medium based on the information obtained by the monitoring unit. (Item 183) The system according to any one of the above items, characterized in that the culture medium is selected from a plurality of suitable culture media and the optimal culture medium is selected or used by monitoring the state of the cells with the monitoring unit. (Item 184) The system according to any one of the above items, wherein the culture medium supply unit is configured to put multiple main components of the culture medium into multiple containers of the automated culture system. (Item 185) The monitoring unit is configured to monitor the culture state and instantly calculate the optimal composition in the culture medium compatibility unit, as described in any one of the above items. Tem. (Item 186) The system according to any one of the above items, wherein the culture medium supply unit is configured to blend the main components of the culture medium to achieve an optimal composition and to continue the culture. (Item 186A) The system according to any one of the above items, wherein the cell information providing unit is configured to specify the cell type and preferred parameters (e.g., cell number) and input them into a computer. (Item 186B) The system described in any one of the above items, wherein the culture medium information providing unit is configured to obtain culture media and culture medium components corresponding to cell types from a database and / or to input candidate culture media and components. (Item 186C) The system according to any one of the above items, wherein the culture medium matching unit is configured to calculate a matching coefficient C1 from information about cells and information about the culture medium or culture medium components, and, if necessary, to calculate a parameter P2' of the culture medium component such that C1 is equal to or greater than a predetermined value. (Item 186D) The system described in any one of the above items, wherein the culture medium information providing unit is configured to perform wet experiments (cell culture experiments) and / or in silico calculations (calculations of cell proliferation rates based on a pre-established algorithm) for each combination of culture medium and components. (Item 186E) The culture performance corresponding to the parameter to be prioritized (for example, cell growth rate (increase in the number of cells)) is obtained, for example, by comparing the culture performances of each candidate condition and determining the medium component and culture condition with the best culture performance as the optimal condition, in the system according to any one of the above items. (Item 186F) The cell culture condition adjustment unit is configured to provide a medium corresponding to the optimal condition, culture cells, and / or adjust the culture conditions according to the progress of the culture when culturing cells with the provided medium, in the system according to any one of the above items. (Item 186G) The cell providing unit, cell information providing unit, medium information providing unit, medium compatibility unit, the medium providing unit, and the cell culturing unit constitute a culture device in a form where a plurality of culture components are stored in a bottle during culture, and are configured to appropriately monitor the culture performance (for example, cell growth rate), and / or The cell culture condition adjustment unit is configured to optimize the combination and content of the culture components in real time by in-silico calculation according to the performance, in the system according to any one of the above items. (Item 187) A method for performing automated cell culture using the system according to any one of the above items. (Item 187A) The provision of the cell information includes specifying the cell type and the parameter to be prioritized (for example, the number of cells) and inputting them into a computer, in the method according to any one of the above items. (Item 187B) The provision of the medium information includes obtaining the medium and medium components corresponding to the cell type from a database and / or inputting candidate media and components, in the method according to any one of the above items. (Item 187C) The compatibility of the medium includes calculating the compatibility coefficient C1 from the information about the cells and the information about the medium or medium components, and optionally calculating the parameter P2' of the medium component such that C1 is not less than a predetermined value, in the method according to any one of the above items. (Item 187D) The method described in any one of the above items, which includes providing the culture medium information by performing wet experiments (cell culture experiments) and / or in silico calculations (calculations of cell proliferation rates based on a pre-established algorithm) for each combination of culture medium and components. (Item 187E) The culture performance corresponding to the priority parameter (e.g., cell proliferation rate (increase in cell number)) is obtained, for example, by comparing the culture performance of each candidate condition and determining the culture medium component and culture condition that yields the best culture performance as the optimal condition, as described in any one of the above items. (Item 187F) The adjustment of the cell culture conditions provides a medium corresponding to the optimal conditions for cells. The method according to any one of the above items, comprising culturing and / or culturing cells in a provided medium, and adjusting the culture conditions in accordance with the course of the culture. (Item 187G) The method according to any one of the above items, wherein the culture apparatus is configured such that multiple culture components are stored in bottles during culture, the culture results (e.g., cell proliferation rate) are monitored as appropriate, and / or the adjustment of the cell culture conditions is optimized in real time by in silico calculation according to the results. (Item 188) A program for performing automated cell culture in any of the systems described in any of the above items. (Item 189) A system according to any one of the above items, the method according to any one of the above items, or a program according to any one of the above items, wherein the cells are mesenchymal stem cells. (Item 189A) A computer-readable recording medium that stores a program described in any one of the above items. Logistics (Item 190) A system for delivering and transporting appropriate culture media to cells from a user, 1) Selection means for selecting appropriate culture medium components and culture and / or storage conditions for the cells, 2) A transport means including means for transporting the culture medium components, 3) Condition calculation means for calculating conditions appropriate for transporting the culture medium components, 4) Control unit and A system comprising at least one selected from the following. (Item 190A) The condition calculation means for calculating the conditions appropriate for the transport includes wet experiments (i.e., cell culture experiments under conditions corresponding to the transport conditions) or in silico calculations (using a pre-constructed database and / or algorithms) for each combination of culture medium / component and transport conditions. A system described in any one of the above items that performs cell viability calculations based on the formula and obtains culture (transport) results (e.g., cell viability). (Item 190B) A system according to any one of the above items, wherein the condition calculation means for calculating conditions suitable for the transport further, or alternatively, determines whether the cell viability of the cells is equal to or greater than a desired standard (e.g., 95%, 90%, 85%, 80%), and if it is less than the standard, changes the transport conditions and obtains the cell viability again, and repeats the process of obtaining the cell viability until a cell viability equal to or greater than the desired standard is obtained. (Item 190C) The condition calculation means for calculating the conditions suitable for the transport may further, or alternatively, specify the transport / storage conditions to some extent beforehand, and then determine the culture medium, components, and conditions that are suitable for those conditions. Select and optimize the system described in any one of the above items. (Item 190D) The condition calculation means for calculating appropriate conditions for transport further, or alternatively, if transport constraints stipulate that transport of cell A is required at 20°C for 8 hours, obtain candidate culture media and components for cell A from a database, or perform a wet experiment or in silico calculation equivalent to transport at 20°C for 8 hours, obtain the cell viability rate, and adjust and optimize the culture media and components until a viability rate above the desired standard is obtained, as described in any one of the above items. (Item 191) A system for delivering and transporting a combination of cells and a suitable culture medium from a user, 1) Selection means for selecting appropriate culture medium components and culture and / or storage conditions for the cells, 2) A transport means including means for transporting the combination of the culture medium components and the cells, 3) Condition calculation means for calculating conditions appropriate for transporting the combination of culture medium components and cells, 4) A control unit that controls the delivery and transport of the combination of the culture medium components and the cells, A system comprising at least one selected from the following. (Item 191A) The condition calculation means for calculating the conditions appropriate for the transport includes wet experiments (i.e., cell culture experiments under conditions corresponding to the transport conditions) for each combination of culture medium / component and transport conditions. A system described in any one of the above items that performs either a trial or in silico calculation (calculation of cell viability based on a pre-established database and / or algorithm) and obtains culture (transport) results (e.g., cell viability). (Item 191B) A system according to any one of the above items, wherein the condition calculation means for calculating conditions suitable for the transport further, or alternatively, determines whether the cell viability of the cells is equal to or greater than a desired standard (e.g., 95%, 90%, 85%, 80%), and if it is less than the standard, changes the transport conditions and obtains the cell viability again, and repeats the process of obtaining the cell viability until a cell viability equal to or greater than the desired standard is obtained. (Item 191C) The condition calculation means for calculating the conditions suitable for the transport may further, or alternatively, specify the transport / storage conditions to some extent beforehand, and then determine the culture medium, components, and conditions that are suitable for those conditions. Select and optimize the system described in any one of the above items. (Item 191D) The condition calculation means for calculating appropriate conditions for transport further, or alternatively, if transport constraints dictate that transport of cell A requires transport at 20°C for 8 hours, obtain candidate culture media and components for cell A from a database, or perform a wet experiment or in silico calculation equivalent to transport at 20°C for 8 hours, obtain cell viability, and adjust and optimize the culture media and components until a viability rate above a desired standard is obtained, as described in any one of the above items. (Item 192) The selection is the system described in any one of the above items, selected by the method described in any one of the above items. (Item 193) A culture medium suitable for transporting cells as described in the specification, containing the components described in the specification. (Item 194) A method for transporting the desired cells using a culture medium suitable for transporting the cells described in the specification, which contains the components described in the specification. (Item 195) The system according to any one of the above items, wherein the cells are mesenchymal stem cells, the culture medium according to any one of the above items, and the method according to any one of the above items. (Item 195A) The method according to any one of the above items, wherein calculating the conditions appropriate for the transport includes performing wet experiments (i.e., cell culture experiments under conditions equivalent to the transport conditions) or in silico calculations (calculation of cell viability based on a pre-established database and / or algorithm) for each combination of culture medium / component and transport conditions, and obtaining culture (transport) results (e.g., cell viability). (Item 195B) The method of any one of the above items, wherein calculating conditions suitable for the transport further, or alternatively, includes determining whether the cell viability of the cells is above a desired standard (e.g., 95%, 90%, 85%, 80%), changing the transport conditions and obtaining the cell viability again if it is below the standard, and repeating the process of obtaining the cell viability until a cell viability of above the desired standard is obtained. (Item 195C) Calculating the appropriate conditions for the aforementioned transport may be, or alternatively, done by first specifying the transport / storage conditions to some extent and then selecting the culture medium, components, and conditions that are suitable for those conditions. A method of any one of the above items, which includes optimization. (Item 195D) The method of any one of the above items, further or alternatively, if, due to transport constraints, it is determined that Cell A requires transport at 20°C for 8 hours, obtain candidate culture media and components for Cell A from a database, or perform a wet experiment or in silico calculation equivalent to transport at 20°C for 8 hours, obtain cell viability, and adjust and optimize the culture media and components until a viability above the desired standard is obtained. (Item 195E) A program causing a computer to perform a method of delivering and transporting cells from a user in combination with a suitable culture medium, wherein the method is the method described in any one of the above items. (Item 195F) A computer-readable recording medium that stores a program described in any one of the above items. <Monitoring> (Item 196) A method for managing culture medium components and / or cell production with respect to cells provided by a user, 1) A step of providing information regarding appropriate culture medium components for the cells provided by the user, 2) A step of assigning identifiable labels to the cells and the culture medium components, 3) A step of checking whether the cells and the culture medium components are compatible by referring to the label. A method of including. (Item 197) The control thereof refers to the method of any one of the above items, wherein the control refers to the information used in the method of any one of the above items. (Item 198) A method for causing a computer to manage culture medium components and / or cell production for cells provided by a user, 1) A step of providing information regarding appropriate culture medium components for the cells provided by the user, 2) A step of assigning identifiable labels to the cells and the culture medium components, 3) A step of checking whether the cells and the culture medium components are compatible by referring to the label. A method of including. (Item 198A) The method according to any one of the above items, wherein step 2) provides a living cell staining dye (e.g., Calcein-AM) as a label to the culture medium. (Item 198B) The method according to any one of the above items, wherein step 3) of providing a label (e.g., Calcein-AM) over time is measured and culture results (e.g., cell count) are monitored in real time from the signal. (Item 199) A program causing a computer to perform a method for managing culture medium components and / or cell production for cells provided by a user, wherein the method 1) A step of providing information regarding appropriate culture medium components for the cells provided by the user, 2) A step of assigning identifiable labels to the cells and the culture medium components, 3) A step of checking whether the cells and the culture medium components are compatible by referring to the label. A program that includes this. (Item 199A) A computer-readable recording medium that stores a program described in any one of the above items.

[0034] <Personalization> (Item 200) A method for providing the most appropriate healthcare and medical treatment to a subject, The process of providing medical information about the subject, A step of providing appropriate cells based on the aforementioned medical information, A step of providing optimal culture medium components or combinations based on the aforementioned cells and the aforementioned medical information. If necessary, the step of providing the cells cultured with the culture medium components or a combination thereof to the target. A method of including. (Item 200A) The method according to any one of the above items, wherein the step of providing the medical information includes calculating the health issues that need to be addressed most for the individual in question and information on side effects that should be considered, using an algorithm based on the input medical information, and selecting from a database candidate cells that have an effect that contributes to solving the health issues and do not have any concerns about side effects that should be considered. (Item 200B) The step of providing the optimal culture medium components or combinations based on the cells and the medical information involves selecting the culture medium components that maximize the effect on the selected candidate cells. The method described in any one of the above items, which includes selecting. (Item 200C) The method according to any one of the above items, wherein the step of providing an optimal culture medium component or combination thereof based on the cells and the medical information comprises selecting the combination that has the greatest effect among each candidate cell × culture medium component combination. (Item 201) The method described in any one of the above items, wherein the provision of the optimal culture medium components or combinations thereof is made by the method described in any one of the above items. (Item 202) A system that provides optimal healthcare and medical treatment to the target, Means of providing medical information about the subject, A means for providing appropriate cells based on the aforementioned medical information, Means for providing optimal culture medium components or combinations based on the aforementioned cells and the aforementioned medical information. Means for providing the cells cultured with the culture medium components or combinations thereof to the target, if necessary. A system that encompasses all of these. (Item 202A) The means for providing the medical information is configured to use an algorithm based on the input medical information to calculate the health issues that need to be addressed most for the individual concerned and information on side effects that should be considered, and to select from a database candidate cells that have an effect that contributes to solving the health issues and do not have any concerns about side effects that should be considered, as described in any one of the above items. (Item 202B) A means for providing an optimal culture medium component or combination based on the cells and the medical information, the system according to any one of the above items, configured to select the culture medium component that maximizes the effect on selected candidate cells. (Item 202C) A means for providing an optimal culture medium component or combination based on the cells and the medical information is a system according to any one of the above items, configured to select the combination that has the greatest effect among each candidate cell × culture medium component combination. (Item 203) A program for a computer to implement a method for providing an optimal healthcare or medical treatment to a subject, the method The process of providing medical information about the subject, A step of providing appropriate cells based on the aforementioned medical information, A step of providing optimal culture medium components or combinations based on the aforementioned cells and the aforementioned medical information. If necessary, the step of providing the cells cultured with the culture medium components or a combination thereof to the target. A program that encompasses all of these. (Item 204) The system according to any one of the above items, the method according to any one of the above items, or the program according to any one of the above items, wherein the cells are microbial cells or animal or plant cells. (Item 205) A system according to any one of the above items, a method according to any one of the above items, or a program according to any one of the above items, wherein the cells are mesenchymal stem cells. (Item 205A) A computer-readable recording medium that stores a program described in any one of the above items. (Item 206) A method for generating a process development method, 1) A step of inputting information about cells or proteins related to said cells into a database, 2) A step of inputting information on the modification of the cells or the modification of the proteins into a database, 3) A step of outputting from the database information regarding scaffold materials that readily adhere to adherent cells present in the cell culture, and information regarding equipment materials that do not readily adhere to adherent cells. 4) A step of outputting information from the database regarding adherent cells, suspension cells, and / or materials in which components eluted from cells are less likely to condense, 5) A step of outputting a culture vessel suitable for cell culture from the database, 6) A step of outputting information from the database regarding sensors that can sense cells, culture medium components, and culture vessels, 7) A step of optimizing the cell culture conditions and the culture medium components, and obtaining data for optimizing the process in accordance with the modification of the cells or the modification of the protein, 8)7) Analysis process to sort the data obtained in 7) into stable data and unstable data, 9) A method comprising the step of outputting a suggestion to monitor the unstable data more frequently than the stable data. (Item 207) A system for developing processes, 1) A cell protein input unit that inputs information about cells or proteins related to said cells into a database, 2) A modification input unit that inputs information on the modification of the cells or the modification of the proteins into a database, 3) An adhesion-related output unit that outputs from the database information regarding scaffold materials that readily adhere to adherent cells present in the cell culture and information regarding equipment materials that do not readily adhere to adherent cells, 4) A material-related output unit that outputs information from the database regarding adherent cells, suspension cells, and / or materials from which components eluted from cells are less likely to condense, 5) A container-related output unit that outputs a culture vessel suitable for cell culture from the database, 6) A sensor-related output unit that outputs information from the database about sensors that can sense cells, culture medium components, and culture vessels, 7) A data acquisition unit that acquires data for optimizing the cell culture conditions and the culture medium components, and for optimizing the process in accordance with the modification of the cells or the modification of the proteins, 8)7) An analysis unit that sorts the data obtained in 7) into stable data and unstable data, 9) A system including a proposal output unit that outputs a proposal to monitor the unstable data more frequently than the stable data.

[0035] In this disclosure, the one or more of the above features are intended to be provided in combinations other than those explicitly stated. Further embodiments and advantages of this disclosure will be apparent to those skilled in the art, by reading and understanding the detailed description below as necessary. [Effects of the Invention]

[0036] According to this disclosure, the desired effect can be ensured by identifying and controlling the optimal culture medium for cells. Furthermore, according to this disclosure, quality control that clearly indicates the equivalence and homogeneity of optimal cells and culture media becomes possible. [Brief explanation of the drawing]

[0037] [Figure 1] Figure 1 is a flowchart illustrating the cell quality control method of this disclosure. [Figure 2] Figure 2 is a flowchart illustrating a method for producing a quality-controllable culture medium according to the present disclosure. [Figure 3] Figure 3 is a flowchart illustrating a specific example of the cell quality control method described herein. [Figure 4] Figure 4 is a flowchart illustrating a specific example of the correlation analysis method described herein. [Figure 5] Figure 5 is a flowchart illustrating a specific example of the cell quality control method described herein. [Figure 6] Figure 6 is a flowchart illustrating another specific example of the cell quality control method of this disclosure. [Figure 7] Figure 7 is a flowchart illustrating the cell quality control method of this disclosure. [Figure 8] Figure 8 is a flowchart illustrating the method for selecting the optimal culture conditions according to this disclosure. [Figure 9] Figure 9 is a flowchart of the culture optimization process described herein. [Figure 10] Figure 10 is a flowchart illustrating the method for selecting the optimal culture conditions according to this disclosure. [Figure 11] Figure 11 is a flowchart illustrating the provision of the optimal culture medium and conditions according to this disclosure. [Figure 12] Figure 12 is a flowchart illustrating the method for selecting the optimal culture medium and conditions according to this disclosure. [Figure 13] Figure 13 is a flowchart illustrating the provision of appropriate culture media and culture conditions according to this disclosure. [Figure 14] Figure 14 is a flowchart of the optimization process described herein. [Figure 15] Figure 15 is a flowchart of the cost calculation method of this disclosure. [Figure 16] Figure 16 is a flowchart illustrating another example of the cost calculation method of this disclosure. [Figure 17] Figure 17 is a flowchart illustrating an example of cost calculation in this disclosure. [Figure 18] Figure 18 is a flowchart illustrating another example of cost calculation in this disclosure. [Figure 19] Figure 19 is a flowchart of the production management process described herein. [Figure 20] Figure 20 is a flowchart illustrating another example of cost calculation in this disclosure. [Figure 21] Figure 21 is a flowchart illustrating another example of cost calculation in this disclosure. [Figure 22] Figure 22 is a flowchart illustrating an example of the cost reduction method described herein. [Figure 23] Figure 23 is a flowchart illustrating an example of database creation for the culture medium components and / or cell culture conditions of this disclosure. [Figure 24] Figure 24 is a flowchart illustrating another example of database creation for culture medium components and / or cell culture conditions according to this disclosure. [Figure 25] Figure 25 is a flowchart illustrating another example of database creation for culture medium components and / or cell culture conditions according to this disclosure. [Figure 26] Figure 26 is a flowchart illustrating an example of the culture automation described herein. [Figure 27] Figure 27 is a flowchart illustrating another example of culture automation as described herein. [Figure 28] Figure 28 is a flowchart illustrating another example of culture automation as described herein. [Figure 29] Figure 29 is a flowchart illustrating an example of the culture medium delivery and transport described herein. [Figure 30] Figure 30 is a flowchart illustrating another example of the culture medium delivery and transport described herein. [Figure 31A] Figure 31A is a flowchart illustrating another example of the culture medium delivery and transport described herein. [Figure 31B] Figure 31B is a flowchart illustrating another example of the culture medium delivery and transport described herein. [Figure 32] Figure 32 is a flowchart illustrating an example of a system for delivering and transporting appropriate culture medium and cell combinations according to this disclosure. [Figure 33] Figure 33 is a flowchart of another example of a system for delivering and transporting suitable culture medium and cell combinations according to this disclosure. [Figure 34A] Figure 34A is a flowchart of another example of a system for delivering and transporting suitable culture medium and cell combinations according to this disclosure. [Figure 34B] Figure 34B is a flowchart of another example of a system for delivering and transporting suitable culture medium and cell combinations according to this disclosure. [Figure 35] Figure 35 is a flowchart illustrating an example of the control of culture medium components and / or cell production according to this disclosure. [Figure 36] Figure 36 is a flowchart illustrating another example of the control of culture medium components and / or cell production according to this disclosure. [Figure 37] Figure 37 is a flowchart illustrating another example of the control of culture medium components and / or cell production according to this disclosure. [Figure 38] Figure 38 is a flowchart illustrating an example of healthcare and medical procedure optimization as described herein. [Figure 39] Figure 39 is a flowchart illustrating another example of healthcare and medical procedure optimization as described herein. [Figure 40] Figure 40 is a flowchart illustrating another example of healthcare and medical procedure optimization as described herein. [Figure 41] Figure 41 shows a graph illustrating the correlation between the concentration of the components in Examples 1-2 and the cell quality indicators. The x-axis represents the component concentration (μg / ml), and the y-axis represents the cell number (×10⁵ cells). [Modes for carrying out the invention]

[0038] Throughout this specification, singular expressions should be understood to include the concept of their plural form unless otherwise specified. Therefore, singular articles (e.g., "a," "an," "the" in English) should be understood to include the concept of their plural form unless otherwise specified. Furthermore, terms used herein should be understood to have the meaning commonly used in the art unless otherwise specified. Accordingly, unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by those skilled in the art to which this disclosure pertains. In case of any conflict, this specification (including definitions) shall prevail.

[0039] (definition) First, we will explain the terms and general technologies used in this disclosure.

[0040] In this specification, “culture medium” refers to a substrate or liquid that provides the nutrients and environment necessary for the growth and maintenance of microorganisms, cells, or plant tissues. Culture media are designed to meet the specific growth conditions of the target organism and may contain a variety of components. These components may include carbon sources, nitrogen sources, inorganic salts, vitamins, amino acids, serum, growth factors, etc. There are various appropriate culture media for animals, microorganisms, plants, etc., and examples of animal culture media include Dulbecco's Modified Eagle Medium (DMEM). In a more specific embodiment, the medium for culturing mammalian cells is preferably a buffered medium (preferably pH about 7.0, pH=7.3 to 6.6, pH=7.0) containing minimal essential nutrients and components, such as vitamins, trace elements, salts, bulk salts, amino acids, lipids, and carbohydrates. Non-limiting examples of such cell culture media include commercially available media such as RPMI, DMEM:F12, DMEM, HAM / F12, and patented media from various sources (e.g., medium 6.2). The cell culture medium may be a basal cell culture medium. The cell culture medium may also be a basal cell culture medium to which feed media and / or additives have been added. The cell culture medium may also be called a fermentation broth when the cells are cultured in a fermenter or bioreactor.

[0041] As used herein, the terms "basic medium," "basic culture medium," "basic cell culture medium," or "basic cell culture medium" refer to the specific details of mammalian cells, etc., as defined below. This is a cell culture medium for culturing cells. It refers to a medium in which cells are cultured from the start of the cell culture conditions execution and is not used as an additive to another medium, although various components may be added to the medium. The basal medium serves as a base to which further additives or feed medium may be added during culture, i.e., during the execution of the cell culture conditions. The basal cell culture medium is provided from the start of the cell culture conditions process. Generally, the basal cell culture medium provides nutrients such as carbon sources, amino acids, vitamins, bulk salts (e.g., sodium chloride or potassium chloride), various trace elements (e.g., manganese sulfate), pH buffer, lipids, and glucose. Major bulk salts are usually provided only in the basal medium, and it may be preferable that the final molar osmotic concentration in the cell culture conditions solution does not exceed about 280-350 mOsmo / kg so that the cell culture conditions solution can grow and proliferate under reasonable osmotic stress. The medium components of this disclosure may be components added to such a basal medium.

[0042] In this specification, the terms “feed” or “feed medium” refer to concentrated nutrient solutions / concentrated nutrient compositions used as feed in a culture medium for mammalian cells. It is provided as a “concentrated feed medium” to avoid dilution of the cell culture conditions solution, and typically, the feed medium is provided at 10–50 ml / L / day, preferably 15–45 ml / L / day, more preferably 20–40 ml / L / day, and even more preferably 30 ml / L / day, based on the culture start volume in the container (meaning the start volume on day 0). The feed rate is understood as the average feed rate over the feed period. The feed medium typically contains most, though not all, of the components of the basal cell culture medium at higher concentrations. Generally, the feed medium substitutes nutrients consumed during the cell culture conditions, such as amino acids and carbohydrates, but salts and buffers are less important and are generally provided together with the basal medium. The feed medium is typically added to the (basal) cell culture medium / fermentation broth in a fed-batch manner. However, the feed may be added in various ways, such as through continuous addition, bolus addition, or perfusion-related techniques (chemostat or hybrid-perfusion system). Preferably, the feed medium is added once a day, but it may be added more frequently, such as twice a day, or less frequently, such as every other day. Nutrient addition is generally carried out during culture (i.e., from day 0 onward). In contrast to the basal medium, the feed consists of a highly concentrated nutrient solution (e.g., more than 6 times) that provides all components similar to the basal medium, except for "compounds with very high molar osmotic concentrations," such as major bulk salts (e.g., NaCl, KCl, NaHCO3, MgSO4, Ca(NO3)2).Typically, more than six times or more of a basal medium having or lacking reduced bulk salts maintains the molar osmotic concentration in the cell culture condition solution at approximately 270–550 mOsmo / kg, preferably approximately 280–450 mOsmo / kg, more preferably approximately 280–350 mOsmo / kg, by maintaining good solubility of the compounds and sufficiently low molar osmotic concentrations (e.g., 270–1500 mOsmo / kg, preferably 310–800 mOsmo / kg; the molar osmotic concentration of medium 6.2 feed is approximately 1500 mOsmo / kg due to high glucose, salt, and optimized amino acids).

[0043] Cell culture media, i.e., both basal media and / or feed media, may be serum-free, have a chemically defined composition, or be chemically defined and protein-free. As used herein, “serum-free medium” refers to cell culture media for in vitro cell culture conditions that do not contain animal-derived serum. This is preferred because serum may contain contaminants of the animal, such as viruses, and because serum has an unclear composition and varies from batch to batch. The basal media and feed media according to this disclosure are serum-free.

[0044] In this specification, "chemically defined culture medium" refers to a cell culture medium in which all components are known and suitable for in vitro cell culture conditions. More specifically, animal It does not contain any supplements such as serum or hydrolysates of plants, yeast, or animals. It may contain hydrolysates only if all components have been analyzed, their exact composition is known, and it can be prepared reproducibly. The basal and feed media described herein are preferably chemically defined in composition. Therefore, many serum-free media fall under the category of "chemically defined media."

[0045] While not intended to be limiting, examples of serum-free media for use in the production of target cells include: a serum-free medium containing at least one culture factor selected from growth factors and differentiation-inducing factors, human serum albumin, a Src inhibitor, a PKC activator, methylcellulose, Essential 8 medium, recombinant human serum albumin, and non-human cells. It may also contain at least one additive selected from the group consisting of animal serum albumin. The culture factor is at least one differentiation-inducing factor, and the target cells are differentiated cells. The differentiation-inducing factor has the effect of promoting the differentiation of raw cells. The culture factor is at least one growth factor, and the target cells are proliferating cells. The growth factor has the effect of promoting the proliferation of raw cells.

[0046] The example serum-free medium is used in the production of target cells and can be used for various purposes. Specifically, the serum-free medium can be used in the production of proliferating cells or differentiated cells.

[0047] An exemplary serum-free medium is characterized by comprising, as a serum substitute, a combination of (a) human serum albumin and (b) at least one additive selected from the group consisting of a Src inhibitor, a PKC activator, methylcellulose, Essential 8 medium, recombinant human serum albumin, and non-human animal serum albumin.

[0048] In one embodiment, the exemplary serum-free medium is a medium in which the serum contained in a known cell differentiation induction medium containing differentiation-inducing factors is replaced with the serum substitute described above. In this embodiment, the serum-free medium comprises a basic medium (e.g., MEM, DMEM, IMDM, Ham's F-12, DMEM / F12, RPMI1640, etc.), differentiation-inducing factors, and the serum substitute described above. Alternatively, the serum-free medium is a medium in which the serum contained in a known cell growth medium containing growth factors is replaced with the serum substitute described above. In this embodiment, the serum-free medium comprises a basic medium containing growth factors (e.g., MEM, DMEM, IMDM, Ham's F-12, DMEM / F12, RPMI1640, etc.) and the serum substitute described above. In another embodiment, in a serum-free medium for use in the production of target cells, the target cells are megakaryocytes, and the culture factors are megakaryocyte differentiation-inducing factors. That is, in this particular embodiment, the serum-free medium comprises a basic medium (e.g., IMDM, etc.), megakaryocyte differentiation-inducing factors, and the serum substitute. Megakaryocyte differentiation-inducing factors include, for example, SCF, TA-316, KP-457, GNF-351, and Y39983.

[0049] The exemplary serum-free medium, while containing a serum substitute, is superior in that it can maintain the cell culture efficiency (specifically, differentiation efficiency and proliferation efficiency) that is achieved when using the corresponding serum-containing medium. Furthermore, the aforementioned serum-free medium can solve the problems of cost and the stability of cultured cells that arise when using serum by utilizing a serum substitute.

[0050] In one embodiment, the present disclosure includes at least one culture factor selected from growth factors and differentiation-inducing factors, human serum albumin, a Src inhibitor, a PKC activator, methylcellulose, Essential 8 medium, recombinant human serum albumin, and non-human animal serum albumin. A serum-free medium for use in the production of the product by target cells may be used, comprising at least one additive selected from the group consisting of lubumin. An example serum-free medium may be used in the production of the product by target cells.

[0051] An exemplary serum-free medium is characterized by comprising, as a serum substitute, a combination of (a) human serum albumin and (b) at least one additive selected from the group consisting of a Src inhibitor, a PKC activator, methylcellulose, Essential 8 medium, recombinant human serum albumin, and non-human animal serum albumin.

[0052] In one embodiment, a serum-free medium for use in the production of a product by target cells, wherein the production of the product by target cells is platelet production by megakaryocytes, and the culture factor is a megakaryocyte differentiation-inducing factor. That is, in this particular embodiment, the serum-free medium comprises a basic medium (e.g., IMDM), a megakaryocyte differentiation-inducing factor, and the serum substitute described above. The megakaryocyte differentiation-inducing factor may be, for example, SCF, TA-316, KP-457, GNF-351, and Y39983.

[0053] As mentioned above, KP-457 is known to function as a platelet function-preserving agent, meaning it is a substance that maintains the function of platelets produced by megakaryocytes (see WO2012 / 157586). In addition, GNF-351 and Y-39983 are known to function as platelet production promoters, meaning they are substances that promote platelet production by megakaryocytes (see WO2016 / 204256).

[0054] The exemplary serum-free medium, while containing a serum substitute, is superior in that it can maintain the cell culture efficiency and the efficiency of cell product production that would be achieved using a corresponding serum-containing medium. Furthermore, the aforementioned serum-free medium can solve the problems of cost and the stability of cultured cells that arise when using serum by utilizing a serum substitute.

[0055] As used herein, “protein-free medium” refers to a cell culture medium for in vitro cell culture conditions that contains no proteins whatsoever, except for proteins produced by the cells to be cultured, and wherein proteins refer to polypeptides of any length, but exclude single amino acids, dipeptides, or tripeptides. Specifically, growth factors such as insulin and insulin-like growth factor (IGF) are not present in the medium. Preferably, the basal and feed media described herein are chemically defined in composition and protein-free.

[0056] As used herein, the term “commercial media / culture system” refers to commercially available cell culture media having a fully known composition. These media serve as a reference for the media described herein due to the requirement of precise nutritional composition. Examples of commercial media include DMEM:F12(1:1), DMEM, Ham F12, and RPMI. The feed media of the commercial media used herein are prepared as a 12-fold concentrate of a basal medium without bulk salts. The term “commercial culture system” refers to a system comprising a commercially available basal cell culture medium, such as DMEM:F12(1:1), DMEM, Ham F12, and RPMI, and a feed medium which is a concentrated (e.g., 12-fold concentrated) basal medium having or without reduced bulk salts.

[0057] In this specification, “coefficient” refers to a constant multiplied by a variable in mathematics or physics, which in this disclosure may be directly or indirectly related to components and / or cells. Coefficients are used to adjust the influence of variables in formulas and equations and are expressed as numerical values ​​or constants.

[0058] In this specification, “quality control” refers to a set of activities and methods undertaken to ensure that products and services meet certain quality standards. Quality control encompasses the entire process from the design to the manufacture, supply, and use of products and services, and aims to maintain quality and Cell quality control is carried out with the aim of improving quality. Cell quality control refers to a series of processes and methods carried out to ensure that cultured cells and cell products meet specified quality standards. This includes checking the purity, activity, characteristics, and presence or absence of contamination of cells. Cell quality control is important for various purposes such as research, therapy, and product development. In the field of stem cell therapy, a rigorous quality control process is required to ensure the quality of stem cells transplanted into patients, by testing the purity, differentiation ability, and sterility of cells. Cell culture facilities maintain cell quality by regularly monitoring cell growth curves and optimizing culture conditions (temperature, pH, nutrients, etc.). In the manufacture of biopharmaceuticals, the quality control department regularly tests cells to confirm the characteristics of cell lines (genetic stability, protein production capacity, etc.). In laboratory cell culture, strict aseptic techniques and regular microbiological testing are carried out to prevent contamination. In the field of regenerative medicine, quality control methods such as cell surface marker analysis and functional assays are introduced to guarantee the quality of cell products.

[0059] In this specification, "correlated" means that two or more events or variables are related to each other, and that a change in one affects the other. Correlation is particularly important in statistics and data analysis, and is used to understand patterns and trends in data. In biological experiments, the expression of specific genes and the proliferation state of cells are often correlated, and new culture methods are developed based on this correlation.

[0060] In this specification, "judgment" refers to determining the outcome or state of something based on certain criteria or conditions and drawing a conclusion. Judgment is used in various fields and plays an important role in making objective evaluations and decisions. In cell quality control, "judgment" refers to the process of evaluating whether the characteristics and quality of cells meet predetermined standards or specifications and determining whether they are pass or fail or suitable. This judgment is based on various parameters such as cell purity, activity, morphology, genetic stability, and sterility. In the case of cell morphology judgment, the morphology and structure of cultured cells are determined to be normal by microscopic observation. If abnormal morphology is observed, the lot is judged to be unsuitable. In the case of cell purity judgment, the purity of the target cell population is measured using flow cytometry and it is determined whether it meets the predetermined purity standard. If the standard is not met, it is judged to be unsuitable. In the case of genetic stability judgment, genetic analysis of cells that have been cultured for a long period of time is performed to confirm that there are no genetic mutations. If mutations are detected, the cells are judged to be unsuitable. In the case of cell activity judgment, the proliferation rate of cells and specific functions (e.g., differentiation ability, protein production ability) are measured and it is determined whether they meet the standard values. If activity levels below the standard are detected, the cells are deemed unacceptable.

[0061] In this disclosure, determining whether the target cells are the target cells may be important. Such determination may be carried out by a process of confirming whether the cultured or collected cells have predetermined characteristics or markers and determining whether they are suitable for research or therapeutic purposes. This may include morphological observation, expression of specific cell markers, and confirmation of genetic characteristics. In the case of morphological observation, the morphology of the cells is observed using a microscope to confirm whether they have the morphology specific to the target cells. For example, the presence or absence of long axons and dendrites in nerve cells is checked. In the case of flow cytometry, flow cytometry is used to measure whether the target cells express specific cell surface markers (e.g., CD34, CD45). If they are target stem cells, it is confirmed that the specific markers are expressed at a certain rate. In the case of immunohistochemistry, epidemiological staining is used to confirm whether the target cells express specific proteins. For example, in the case of muscle cells, the presence of muscle-specific proteins (e.g., myosin) is confirmed. In the case of genetic analysis, the gene sequence of the target cells is analyzed using PCR or sequencing to confirm gene mutations and expression patterns specific to the target cells. When using functional assays, the target cells are tested to determine if they possess a specific function (e.g., insulin secretion, oxygen transport capacity). For example, in the case of β cells, insulin secretion capacity is tested. The cells are measured to determine if they are the target cells.

[0062] In this specification, "protein molecular behavior" refers to how protein molecules move, change shape, interact, and exert their functions both inside and outside cells. This includes various phenomena such as protein folding, dynamics, binding, denaturation, and activity. Protein behavior is crucial for cellular function and the life activities of organisms. Regarding protein folding, a newly synthesized polypeptide chain folds into a specific tertiary structure to become a functional protein. This folding process is assisted by accessory proteins called chaperones. Regarding protein dynamics, in intracellular signaling pathways, protein molecules are activated or inactivated through a series of phosphorylation and dephosphorylation reactions. This allows cells to respond to external stimuli. Regarding protein binding, specific binding between enzymes and substrates occurs at the enzyme's active site and catalyzes chemical reactions. The specificity and strength of this binding are important for enzyme function. Protein denaturation occurs when a protein molecule loses its normal tertiary structure and becomes unable to function due to extreme changes in temperature or pH. This is called protein denaturation. To explain protein activity, allosteric enzymes have their activity regulated by the binding of specific molecules to other sites on the enzyme. Such regulatory mechanisms are important for controlling metabolic pathways within cells.

[0063] In this specification, "expression behavior of related gene groups" refers to how multiple genes involved in specific functions or processes are expressed, how their expression is regulated, and how they interact with each other. This includes gene transcription, mRNA production, translation, and even post-transcriptional and post-translational modifications. Studying expression behavior is important for understanding gene function and elucidating disease mechanisms. The expression behavior of related gene groups in stress response is the phenomenon in which multiple genes encoding heat shock proteins (HSPs) are simultaneously expressed when cells are subjected to heat shock. This expression behavior is important for protecting cells and recovering from stress. The expression behavior of related gene groups involved in carcinogenesis is the phenomenon in cancer cells where the expression patterns of oncogenes that promote proliferation and tumor suppressor genes that inhibit it become abnormal. By analyzing the expression behavior of these gene groups, the mechanisms of cancer progression can be elucidated. The expression behavior of related gene groups in immune response is the phenomenon in which the expression of genes encoding cytokines and chemokines is induced when immune cells encounter pathogens. This expression behavior is important for regulating immune responses and eliminating pathogens. The expression behavior of related gene groups during development is a phenomenon in which homeotic genes are expressed in specific patterns at certain stages of embryonic development, determining the body axis and organ positions. This expression behavior is essential for normal development and morphogenesis. The expression behavior of related gene groups in metabolic pathways is a phenomenon in which the expression of gene groups encoding enzymes involved in glucose metabolism is regulated in response to fluctuations in blood glucose levels. This expression behavior is important for maintaining the balance of energy metabolism. Related gene groups include immune-related genes, inflammation-related genes, angiogenesis-related genes, nerve regeneration-related genes, chondrogenesis-related genes, osteogenesis-related genes, adipogenesis-related genes, fibrosis-related genes, immunogenicity-related genes, migration-related genes, adhesion-related genes, aging-related genes, mesenchymal stem cell marker-related genes, surface marker-related genes, growth factor-related genes, and chemokine or cytokine-related genes.

[0064] In this specification, "cell surface antigen" refers to specific molecular structures present on the surface of cells, primarily consisting of proteins and glycoproteins. These antigens can be recognized as foreign substances by the immune system and play a role in triggering an immune response. Cell surface antigens are involved in cell recognition, regulation of immune responses, and intercellular interactions.

[0065] In this specification, "expression pattern of a group of genes" refers to multiple expression patterns under specific conditions or time periods. This shows how genes are expressed and to what extent each gene is activated. This pattern includes the process of gene transcription and mRNA production, and the process of mRNA translation to produce proteins. Expression patterns vary depending on cell type, developmental stage, environmental conditions, pathological state, etc.

[0066] In this specification, the terms “cell cultivation” or “cell culture” are interchangeable terms and include cell culture conditions and fermentation processes at all scales (e.g., from microtita plates to large industrial bioreactors, i.e., from sub-mL to over 10,000L), in all different process modes (e.g., batch culture, fed-batch culture, perfusion culture, continuous culture), in all process control modes (e.g., uncontrolled systems, fully automated and controlled systems, with respect to pH, temperature, and oxygen content control), and in all types of fermentation systems (e.g., single-use systems, stainless steel systems, glassware systems). In preferred embodiments of this disclosure, the cell culture conditions are mammalian cell culture conditions and are either batch culture or fed-batch culture.

[0067] As used herein, the term “fed-batch” refers to cell culture conditions in which cells are fed a feed medium containing nutrients continuously or periodically. Feeding may begin immediately after the cell culture conditions are started on day 0, or typically one, two, or three days after the start of culture. Feeding may follow a pre-set schedule, such as daily, every two days, or every three days. Alternatively, the culture medium may be monitored for cell growth, nutrients, or toxic byproducts, and the feed may be adjusted accordingly. Common methods for monitoring animal cell culture conditions are described in the following experimental section. Generally, the following parameters are often determined on a daily basis, encompassing the concentration of viable cells, the concentration of products, and several metabolites, such as glucose or lactate (acidic waste metabolites that lower pH and originate from the conversion of glucose by cells), pH, molar osmotic pressure (an indicator of salt content), and ammonium (a growth inhibitor that adversely affects growth rate and reduces viable biomass). Compared to batch culture (culture without feeding), the fed-batch method can achieve higher titers of the product. Typically, the fed-batch culture is stopped at several point points, the cells and / or proteins of interest in the culture medium are collected, and optionally purified.

[0068] In this specification, the term “viability” is used synonymously with “vitality” and refers to the percentage of viable cells under cell culture conditions, as determined by methods known in the art, such as trypan blue exclusion using a Cedex instrument (Innovatis AG, Bielefeld) based on automated microscopy cell counting. However, many other methods exist for determining viability, such as fluorescence assays (e.g., based on propidium iodide), calorimetry, or enzymatic assays used to reflect the energy metabolism of viable cells, such as LDH lactate dehydrogenase or methods using specific tetrazolium salts, such as Alamar blue, MTT (3-(4,5-dimethylthiazole-2-yl-2,5-diphenyltetrazolium bromide) or TTC (tetrazolium chloride).

[0069] As used herein, the term “amino acid” refers to the 20 natural amino acids encoded by a common genetic code, typically L-type amino acids (i.e., L-alanine, L-arginine, L-asparagine, L-aspartic acid, L-cysteine, L-glutamic acid, L-glutamine, L-glycine, L-histidine, L-isoleucine, L-leucine, L-lysine, L-methionine, L-phenylalanine, L-proline, L-serine, L-threonine, L-tryptophan, L-tyrosine, and L-valine). The amino acids (e.g., glutamine and / or tyrosine) are preferably extended with L-alanine (L-ala-x) or L-glycine (L-gly-x), such as glycyl-glutamine and alanyl-glutamine, which have stability and / or It may be provided as a dipeptide with improved solubility. Furthermore, cysteine ​​may also be provided as L-cystine. As used herein, the term “amino acid” encompasses all of its different salts, for example (but not limited to) L-arginine monohydrochloride, L-asparagine monohydrate, L-cysteine ​​hydrochloride monohydrate, L-cystine dihydrochloride, L-histidine monohydrochloride dihydrate, L-lysine monohydrochloride, and hydroxyl L-proline, L-tyrosine disodium dehydrate. The exact form of the amino acid is not important to this disclosure as long as its characteristics, such as solubility, molar osmotic concentration, stability, and purity, are not impaired. Typically and preferably, L-arginine is used as L-arginine × HCl, L-asparagine as L-asparagine × H2O, L-cysteine ​​as L-cysteine ​​× HCl × H2O, L-cystine as L-cystine × 2HCl, L-histidine as L-histidine × HCl × H2O, and L-tyrosine as L-tyrosine × 2Na × 2H2O, with each preferred amino acid form being selected independently of others, together, or in any combination thereof. Dipeptides containing one or two related amino acids are also included. For example, L-glutamine is often added to cell culture media in the form of a dipeptide such as L-alanyl-L-glutamine to improve stability during storage or long-term culture and to reduce ammonium generation. This is also true for L-glycine-containing dipeptides or other L-alanine-containing dipeptides, which are taken into consideration for the calculation of amino acid ratios.

[0070] In this specification, the terms “all amino acids in the culture medium” or “total amino acid content” refer to the sum of “amino acids” as defined above in mM. In dipeptides, each amino acid is counted separately, so 1 mM alanyl glutamine is 1 mM L-alanine and 1 mM L-glutamine (molar ratio 1:1) are counted. Similarly, in L-cystine, each cysteine ​​is counted separately, so 1 mM L-cystine is counted as 2 mM It is counted as L-cysteine ​​(molar ratio 1:2). Typically, the total amino acid content in the concentrated feed medium is about 5 to 20 times, preferably about 7 to 15 times, and more preferably about 10 times or more, compared to the basal cell culture medium. The total amino acid content of the basal medium described herein is about 25 to 150 mM, preferably about 30 to 130 mM, more preferably about 35 to 120 mM, and even more preferably about 40 to 100 mM. The total amino acid content of the feed medium may be about 100 to 1000 mM, preferably about 200 to 900 mM, more preferably about 300 to 800 mM, and even more preferably about 400 to 700 mM. Other amino acids not directly encoded by the common genetic code, such as L-ornithine, hydroxyl-L-proline, or their metabolites, such as taurine, may also be present in the basal cell culture medium or feed medium, but these are not counted in the total amino acid content.

[0071] As used herein, the term “amino acid ratio” refers to the ratio of the molar concentration of each amino acid to the molar concentration of a reference amino acid. The molar ratio of all amino acids to the reference amino acid is calculated (using the unit [mM / mM]). To calculate the amino acid ratios in accordance with this disclosure, L-isoleucine is used as the reference amino acid (theoretically, other amino acids such as phenylalanine or methionine may be used as the reference amino acid). This may further be referred to as the molar ratio (mM / mM) to isoleucine. Typically, the reference amino acid can be readily measured using statistically low levels of variation and is provided in commonly used culture media at similar concentration ranges.

[0072] The term "adjustment of amino acid ratio in used culture medium" means that amino acids are adjusted based solely on the analysis of the used culture medium, without considering cellular and metabolic needs, or specific intracellular or extracellular velocities. Therefore, amino acid analysis is performed on samples taken from the cell culture supernatant on various days, and amino acids below a certain threshold are supplemented in the basal and feed media.

[0073] In this specification, the terms “polypeptide,” “protein,” “product,” “protein product,” and “amino acid residue sequence” are used synonymously. These terms refer to polymers of amino acids of any length. These terms also include proteins that are post-translationally modified through reactions including, but not limited to, glycosylation, glycation, acetylation, phosphorylation, oxidation, amidation, or protein processing. Modifications and alterations, such as fusion with other proteins, substitution, deletion, or insertion of amino acid sequences, can occur within the structure of a polypeptide while the molecule maintains its biological functional activity. For example, substitution of a particular amino acid sequence can occur within a polypeptide or its underlying nucleic acid coding sequence to obtain a protein having similar or modified properties. Amino acid modifications can be prepared, for example, by performing site-directed mutagenesis or polymerase chain reaction-mediated mutagenesis on the underlying nucleic acid sequence. Therefore, the terms “polypeptide,” “protein,” “product,” and “protein product” also include, for example, fusion proteins consisting of immunoglobulin components (e.g., Fc components) and growth factors (e.g., interleukins), antibodies, or molecular formats or antibody fragments derived from any antibody.

[0074] In this specification, "culture medium components" is also simply referred to as "components," and refers to the various components contained in the culture medium that provide the nutrients and environment necessary for the growth and maintenance of microorganisms, cells, or plant tissues. These components support cell proliferation and metabolic activity and play an important role in the target research and product development. Basic components include: carbon sources: glucose, galactose, etc.; nitrogen sources that serve as energy sources for cells: amino acids, peptones, yeast extract, etc.; inorganic salts necessary for protein synthesis and metabolism, such as calcium salts: CaCl2 (calcium chloride); magnesium salts involved in cell wall stabilization and signal transduction, such as MgSO4 (magnesium sulfate); and components that act as cofactors for enzymes. Vitamin B 12Examples include cyanocobalamin and vitamin B6 (pyridoxine). Vitamins that aid in cell metabolism and growth may also be added. By customizing the culture medium components according to specific cell lines and experimental conditions, the characteristics and functions of cells can be optimized. For example, specific vitamins or amino acids required by certain cell lines may be added. Some cells and microorganisms may require cultivation under special environmental conditions, such as hypoxic conditions or specific pH conditions. It is important to appropriately adjust the culture medium components to adapt to these conditions.

[0075] In this specification, “cellular information” refers to all biological data that a cell holds and transmits. This information includes genetic information (DNA and RNA sequences), intracellular signaling pathways, protein expression patterns, cell morphology, function, and metabolic state. Cellular information is essential for understanding how cells function and respond to the environment. At the core of cellular information is genetic information stored in the cell's DNA. This information determines cell growth, division, differentiation, and function. Signaling information includes intracellular signaling pathways that are activated when receptors on the cell surface bind to specific molecules (e.g., hormones, growth factors), thereby regulating the cell's response. Signaling information in this process modulates cell behavior. Gene expression information includes data on which genes are activated and which proteins are synthesized under specific conditions. This allows us to understand the functional state of the cell.

[0076] In this specification, “basic cellular functions” refer to the biochemical and physical processes essential for the survival, growth, differentiation, proliferation, and adaptation of cells in an organism. These include energy production and utilization, retention and expression of genetic information, transport and exchange of substances inside and outside the cell, signal transduction, and regulation of the cell cycle. Regarding energy production and utilization, this includes ATP production (oxidative phosphorylation) in mitochondria, where mitochondria react nutrients obtained from food with oxygen to produce ATP, an energy currency. This ATP is used to meet various energy demands within the cell. Regarding the retention and expression of genetic information, for example, DNA replication and Transcription is one such function. During cell division, cells accurately replicate their own DNA and transmit it to daughter cells. Also, specific sections of DNA are transcribed into RNA, which is used for protein synthesis. Regarding the transport and exchange of substances, there is ion transport across the cell membrane, where transport proteins such as the sodium-potassium pump regulate the ion concentration inside and outside the cell, maintaining cellular homeostasis. Regarding signal transduction, for example, there is hormonal signaling, where hormones such as insulin circulate in the blood and bind to receptors on target cells, thereby regulating the metabolic activity of the cell. Regarding the regulation of the cell cycle, for example, there are cell cycle checkpoints, where cells have checkpoints that detect DNA damage and other abnormalities and halt the progression of the cell cycle until repair is complete. This is an important function to prevent abnormal cell division. These basic cellular functions are essential for cells to function normally and maintain the health of tissues and organs, and therefore may be the subject of investigation.

[0077] In this specification, "culture medium factors" refer to substances and conditions necessary to promote the growth and proliferation of microorganisms and cells under specific environmental and cellular conditions. These factors are appropriately selected depending on the type and purpose of the organism being cultured. Examples include: nutrients (including carbon sources, nitrogen sources, vitamins, minerals, etc. For example, glucose is a major carbon source for many microorganisms and cells); growth factors (proteins and peptides that promote cell proliferation and differentiation. For example, epidermal growth factor (EGF) promotes the growth of epithelial cells); Serum (commonly used in animal cell culture; fetal bovine serum (FBS) is a typical example.) It contains many growth factors and hormones, hormones (which control the growth and function of specific cells; for example, insulin promotes glucose uptake by cells), amino acids (essential elements necessary for protein synthesis in cells; for example, L-glutamine is added to many culture media), vitamins (essential micronutrients for cell metabolism and growth; for example, vitamin B12 is important for cell division), and inorganic salts (necessary to maintain the osmotic balance of cells). Yes, for example, sodium salts and potassium salts are used. Physical conditions (such as temperature, pH, and oxygen concentration, which are also physical conditions of the culture environment) are also included as culture medium factors. For example, E. coli grows best at 37°C. By optimizing these culture medium factors, it is possible to effectively culture the target microorganism or cells.

[0078] In this specification, “activation” refers to the initiation or enhancement of biochemical and physical processes necessary for a cell to effectively perform its basic functions, such as survival, growth, differentiation, proliferation, and adaptation. Activation of basic cellular functions refers to the enhancement of cellular activity, where basic cellular functions (such as energy production, retention and expression of genetic information, substance transport, signal transduction, and regulation of the cell cycle) are promoted by specific stimuli or conditions.

[0079] In this specification, "suppression" refers to a state in which the basic functions of a cell, such as survival, growth, differentiation, proliferation, and adaptation, are suppressed, resulting in decreased cellular activity. It refers to the process in which the basic functions of a cell (energy production, retention and expression of genetic information, substance transport, signaling, regulation of the cell cycle, etc.) are reduced or stopped by specific stimuli or conditions. This reduces cellular activity and, in some cases, can lead to cell death. Suppression of basic cellular functions plays an important role as a regulatory mechanism in living organisms. For example, it is useful for suppressing the proliferation of unwanted or harmful cells or for eliminating damaged cells. It is also used as a drug or treatment to suppress the proliferation of cancer cells. However, excessive suppression can cause dysfunction of tissues and organs, so an appropriate balance is important.

[0080] In this specification, "sample" refers to a portion selected from a whole for a specific study, analysis, investigation, or evaluation. A sample is representative of the whole (population) and is used to understand its characteristics and behavior. Specific examples of "samples" in various fields include: in the case of cells, a portion of cells cultured in a laboratory is taken and used to examine their characteristics and reactions; or when inspecting a portion of a product, production... A randomly selected sample of product from the production line is taken and its quality is tested to estimate the overall quality. In the case of blood samples, a portion of blood collected from a patient is tested and used to assess their health. By appropriately selecting and analyzing samples, it is possible to obtain useful information in various fields, such as scientific research, statistical surveys, quality control, and medical diagnosis.

[0081] In this specification, a "functional map" refers to a diagram or other representation that visually depicts the functions of various structures and organelles within a cell, as well as their interactions and arrangement. By appropriately selecting and analyzing samples, it becomes possible to obtain useful information in various fields, such as scientific research, statistical surveys, quality control, and medical diagnostics. In the case of intracellular organelle arrangement and roles, this includes diagrams showing the roles and interactions of organelles such as the nucleus, mitochondria, ribosomes, Golgi apparatus, endoplasmic reticulum, and lysosomes. For example, mitochondria are involved in energy production (ATP synthesis), ribosomes in protein synthesis, and the Golgi apparatus in protein modification and transport. In the case of signaling pathway maps, they show how signal molecules are transmitted within the cell, starting from receptors on the cell membrane, and ultimately regulating gene expression and metabolic activity. For example, they can show details of specific signaling pathways such as the MAPK pathway and the PI3K / AKT pathway. In the case of metabolic maps, these are diagrams showing processes that generate energy within the cell, such as glucose breakdown (glycolysis), the citric acid cycle (TCA cycle), and the electron transport chain. This allows us to understand the interrelationships of each metabolic pathway and the flow of energy production. For cytoplasmic proteins, the diagram shows where specific proteins are localized within the cell and what functions they perform. For example, it shows that transcription factors are localized in the nucleus and play a role in regulating gene transcription. In the case of cytoskeleton structure and function, it shows the components of the cytoskeleton, such as actin filaments, microtubules, and intermediate filaments, along with their arrangement and function. This visualizes their roles in maintaining cell shape, movement, and substance transport. Functional maps of cells are a powerful tool for understanding the complex processes and structures within cells.

[0082] In this specification, "library" refers to a systematic collection, organization, and storage of information, resources, and data related to cells, or to a physical entity. Such libraries are crucial for researchers to efficiently access necessary data and resources and advance their research and experiments. They can include various types, such as protein libraries and small molecule libraries. Libraries related to cells play a vital role in streamlining research and experiments, sharing and reusing data, and facilitating scientific discovery.

[0083] In this specification, “environment” refers to the surrounding conditions and influencing factors necessary for a cell to survive, grow, and function. This is a complex concept that includes physical, chemical, and biological factors. It refers to the conditions and factors surrounding a cell in which it exists, and these directly or indirectly affect the cell’s physiological activity, growth, differentiation, and response. Physical environment includes temperature, and the optimal operating temperature for a cell varies depending on the species. For example, human cells typically function optimally at 37°C. Pressure is also important; some cells function optimally in specific pressure environments. For example, cells of deep-sea organisms are adapted to high pressure. Light is important as an energy source or signaling source. Plant cells and some algal cells use light to generate energy. Chemical environment includes pH; the pH inside and outside the cell has a significant impact on the cell’s enzyme activity and metabolic processes. For example, human cells typically function optimally at around pH 7.4. Nutrients, such as glucose, amino acids, vitamins, and minerals, are essential for cell growth and metabolism. Oxygen and carbon dioxide are also important; oxygen is an important respiratory substrate for many cells, and the concentration of carbon dioxide can also affect the cell’s metabolic activity. In terms of the biological environment, interactions with neighboring cells (adhesion, signal transduction) influence cell function and behavior. For example, cells that make up a tissue exchange signals with each other and function cooperatively. The extracellular matrix (ECM) provides the supporting structure for cells and is an important element that influences cell growth, migration, and differentiation. Looking at hormones and growth factors, cell division... These are important signaling molecules that regulate growth and differentiation. Looking at the immune environment, immune system cells, as immune cells, protect cells from infection and damage. These include macrophages and lymphocytes. Cytokines are signaling molecules that regulate the immune response and induce inflammation and immune reactions. Considering these factors, the cellular environment can have a significant impact on the physiological state and function of cells. For example, in terms of impact on growth and division, under optimal environmental conditions, cells grow and divide normally. Inappropriate environmental conditions can cause delayed or stopped growth, and even cell death. Regarding differentiation and adaptation, environmental factors promote the differentiation of stem cells and the adaptation of cells to specific environments. For example, a hypoxic environment induces gene expression that allows cells to adapt to oxygen deficiency. As a stress response, harmful environmental conditions (e.g., extreme temperature or the presence of toxins) trigger a stress response in cells, inducing repair mechanisms and apoptosis. Understanding and controlling the cellular environment is extremely important in cell biology and medicine, and is particularly important in fields such as cell culture, tissue engineering, and regenerative medicine.

[0084] (Basic culture medium composition) Culture medium and amino acid ratio The 20 standard amino acids encoded by a general genetic code (L-alanine, L-arginine, L-asparagine, L-aspartic acid, L-cysteine, L-glutamic acid, L-glutamine, L-glycine, L-histidine, L-isoleucine, L-leucine, L-lysine, L-methionine, L-phenylalanine, L-proline, L-serine, L-threonine, L-tryptophan, L-tyrosine, and L-valine) play essential roles in protein synthesis because they provide building blocks for both intracellular proteins and proteins of interest (e.g., monoclonal antibodies). Therefore, amino acids interact in diverse ways in cellular metabolism. They are extracted in specific amounts from cell culture media, interconverted in intracellular metabolism, directed either to host cell proteins or protein products, secreted by cells as byproducts, and linked in various ways to cellular metabolic catabolism and anabolism, for example, between amino acid metabolism and the citric acid cycle. In both basal and feed media, the optimal composition, concentration, and ratio of amino acids must be provided for optimal nutrient delivery throughout the life cycle of cell culture conditions (inoculation, lag phase, exponential growth phase, transition phase, quiescent phase, and death phase characterized by a significant decrease in cell viability). However, the amino acid ratio appears to be more important than the actual precise concentration of each individual amino acid.

[0085] Accordingly, in one embodiment of the present disclosure, a basal cell culture medium for culturing mammalian cells is provided, comprising L-leucine with the following molar ratios (mM / mM) of L-leucine / L-isoleucine: L-leucine with a ratio of approximately 1.2 to 2.2, L-phenylalanine with a ratio of approximately 0.5 to 0.9, L-tyrosine with a ratio of approximately 1.5 to 2.7, L-threonine with a ratio of approximately 1.0 to 1.9, and L-valine with a ratio of approximately 1.0 to 1.9, wherein the basal cell culture medium has a total amino acid content of approximately 25 to 150 mM of the amino acids. In one embodiment, the molar ratio (mM / mM) to isoleucine is: L-leucine / L-isoleucine of about 1.2 to 2.1, preferably about 1.3 to 1.8, more preferably about 1.5 to 1.8, and even more preferably about 1.7; L-phenylalanine / L-isoleucine of about 0.5 to 0.9, preferably 0.6 to 0.9, more preferably about 0.6 to 0.8, and even more preferably about 0.7; and about 1.6 to 2.6, preferably L-tyrosine / L-isoleucine in approximately 1.7-2.5, more preferably 1.9-2.3, even more preferably 2.1; L-threonine / L-isoleucine in approximately 1.1-1.8, preferably 1.2-1.8, more preferably 1.3-1.6, even more preferably 1.5; and L-valine / This is L-isoleucine. In certain embodiments, the culture medium of the present disclosure further comprises L-lysine in a molar ratio of about 1.6 to 2.9, preferably about 1.7 to 2.8, more preferably about 1.8 to 2.7, more preferably about 2.0 to 2.5, and even more preferably about 2.2 relative to isoleucine. In certain embodiments, the basal medium of the present disclosure further comprises L-tryptophan in a molar ratio of about 0.3 to 0.5, preferably about 0.3 to 0.5, more preferably about 0.3 to 0.4, even more preferably about 0.3 to 0.4, relative to isoleucine; or L-proline in a molar ratio of about 1.6 to 3.0, preferably about 1.7 to 2.8, more preferably about 1.8 to 2.7, more preferably about 2.0 to 2.5, even more preferably about 2.3, relative to isoleucine; or L-methionine in a molar ratio of about 0.4 to 0.7, preferably about 0.4 to 0.6, more preferably about 0.4 to 0.6, even more preferably about 0.5 to 0.6, even more preferably about 0.5, relative to isoleucine. In certain embodiments, the molar ratios of L-tryptophan, L-proline, and L-methionine to L-isoleucine are as defined above. The total amino acid content in the basal cell culture medium may be about 25 to 150 mM, preferably 30 to 130 mM, more preferably 35 to 120 mM, and even more preferably about 40 to 100 mM.

[0086] In one example, the amino acid ratios of L-leucine, L-phenylalanine, L-threonine, L-valine, and L-tyrosine, and optionally further L-lysine, L-tryptophan, L-proline, and / or L-methionine, to L-isoleucine are within 30%, 25%, 20%, or 10% of the ratios provided for basal medium 6.2 in Table 2a.

[0087] In one example, a more specific and exemplary amino acid ratio of a basal cell culture medium (basal medium 6.2) of the present disclosure is provided below as an amino acid ratio in a selected commercially available basal cell culture medium.

[0088] Embodiments of the present disclosure provide a feed medium for culturing mammalian cells comprising the following amino acids in the following molar ratios (mM / mM) to isoleucine: L-leucine with a L-leucine / L-isoleucine ratio of about 2.3 to 4.2, L-phenylalanine with a L-phenylalanine / L-isoleucine ratio of about 0.6 to 1.1, L-threonine with a L-threonine / L-isoleucine ratio of about 1.3 to 2.4, and L-valine with a L-valine / L-isoleucine ratio of about 1.1 to 2.0, wherein the feed medium has a total amino acid content of about 100 to 1000 mM. In one embodiment, the molar ratio (mM / mM) to isoleucine is: L-leucine / L-isoleucine of about 2.4 to 4.0, preferably about 2.6 to 3.9, more preferably about 2.9 to 3.5, and even more preferably about 3.2; L-phenylalanine / L-isoleucine of about 0.6 to 1.1, preferably about 0.7 to 1.0, more preferably about 0.8 to 0.9, and even more preferably about 0.9; L-threonine / L-isoleucine of about 1.4 to 2.3, more preferably about 1.5 to 2.2, more preferably about 1.7 to 2.0, and even more preferably about 1.8; and L-valine / L-isoleucine of about 1.2 to 2.0, preferably about 1.3 to 1.9, more preferably 1.4 to 1.7, and even more preferably about 1.6.

[0089] In one embodiment, the feed medium further contains L-tyrosine in a molar ratio of about 0.6 to 1.1 relative to isoleucine and / or L-lysine in a molar ratio of about 1.1 to 2.1 relative to isoleucine. Preferably, tyrosine is present in the feed medium in a ratio of about 0.6 to 1.0, preferably about 0.7 to 1.0, more preferably about 0.7 to 0.9, and even more preferably about 0.8. Preferably, lysine is present in the feed medium in a ratio of about 1.2 to 2.0, preferably about 1.3 to 1.9, more preferably about 1.4 to 1.8, and even more preferably about 1.6. Preferably, the molar ratios of L-tyrosine and L-lysine are as defined above. In certain embodiments, the feed medium of the present disclosure further contains isoleucine. The formulation comprises L-tryptophan in a molar ratio of about 0.3 to 0.6, preferably about 0.3 to 0.6, more preferably about 0.4 to 0.5, even more preferably about 0.5 relative to isoleucine; or L-proline in a molar ratio of about 0.9 to 1.8, preferably about 1.0 to 1.7, more preferably about 1.1 to 1.6, more preferably about 1.2 to 1.5, even more preferably about 1.4 relative to isoleucine; or L-methionine in a molar ratio of about 0.4 to 0.8, preferably about 0.4 to 0.7, more preferably about 0.5 to 0.7, more preferably about 0.5 to 0.6, even more preferably about 0.6 relative to isoleucine. In certain embodiments, the molar ratios of L-tryptophan, L-proline, and L-methionine relative to L-isoleucine are as defined above. The total amino acid content in the basal cell culture medium may be about 100 to 1000 mM, preferably about 200 to 900 mM, more preferably about 300 to 800 mM, and even more preferably about 400 to 700 mM.

[0090] In one embodiment, the amino acid ratios of L-leucine, L-phenylalanine, L-threonine, and L-valine, and optionally further L-tyrosine, L-lysine, L-tryptophan, L-proline, and / or L-methionine, to L-isoleucine are within 30%, 25%, 20%, or 10% of the ratios provided for feed medium 6.2 in Table 6.

[0091] The feed medium is added to the culture medium as concentrated feed medium. For example, the feed medium may be added at a rate of about 10-50 ml / L / day, preferably about 15-45 ml / L / day, more preferably about 20-40 ml / L / day, and even more preferably about 30 ml / L / day, based on the initial culture volume. The rate of addition of feed medium to the cell culture condition solution (mL / L / day) (volume / day) (volume added per liter of initial culture volume in the container per day) is understood as the average rate over the feeding period, and the volume added may be changed for each individual addition during the feeding period. Feeding may also be stopped about 1 to 3 days before the end of culture and / or collection. It is preferable to add small amounts to avoid dilution of other nutrients in the cell culture condition solution and to maintain the culture volume as constant as possible. The feed medium may be added continuously, several times a day, once a day, or every other day. Preferably, the feed medium is added daily or every other day, starting on day 0, day 1, or day 2.

[0092] The basal cell culture media and / or feed media of the present disclosure are serum-free, preferably have a chemically defined composition, or have a chemically defined composition and are protein-free. The media of the present disclosure are suitable for culturing all types of mammalian cells, such as rodent or human cells, where rodent cells are preferred. More preferably, the mammalian cells are Chinese hamster ovary cells (CHO), such as CHO-K1 cells, CHO-DG44 cells, DuxB11 cells, or CHO GS-deficient cells, most preferably CHO-DG44 cells or CHO GS-deficient cells.

[0093] In one embodiment, trace elements may also be considered. For example, iron is required as a trace element. In vivo, iron is mainly bound to ferritin and transferrin in serum. A typical iron source in cell culture media is transferrin. In advanced serum-free or even protein-free mammalian cell culture media, several iron-related situations need to be addressed, such as the identification of appropriate iron carriers, the low bioavailability of iron, the identification of appropriate physiological concentration ranges (due to the presence of harmful free radicals in vitro regarding the potential toxicity of iron compounds, e.g., having minimal / no effect on cell viability), complex binding behavior (iron can bind to multiple substances in the culture medium formulation, thereby potentially making it easily bioavailable for cell culture conditions), oxidation state, and performance of optimal cell culture conditions (e.g., titer).

[0094] The culture medium of this disclosure is serum-free, preferably has a chemically defined composition, or is chemically defined and protein-free. The culture medium of this disclosure is suitable for culturing all types of mammalian cells, such as rodent or human cells, where rodent cells are preferred. More preferably, the mammalian cells are Chinese hamster ovary cells (CHO), e.g., CHO-K1 cells, CHO-DG44 cells, DuxB11 cells or CHO These are GS-deficient cells, most preferably CHO-DG44 cells or CHO GS-deficient cells.

[0095] Culture medium and other components Cell culture media for culturing mammalian cells may further contain essential nutrients and components, such as vitamins, trace elements, salts, bulk salts, lipids or lipid precursors, and carbohydrates, preferably in a buffered medium.

[0096] Non-restrictive examples of suitable vitamins include biotin (B7), calcium pantothenate, cyanocobalamin (B12), folic acid, myo-inositol, niacinamide (B3), pyridoxal hydrochloride, pyridoxine hydrochloride, riboflavin (B2), and / or thiamine (B1). Non-restrictive examples of trace elements include ammonium molybdate, ammonium vanadate, copper sulfate, nickel sulfate, sodium selenite, sodium silicate, and zinc sulfate, and / or zinc chloride. Non-restrictive examples of lipid precursors include choline chloride, ethanolamine, glycerol, inositol, linoleic acid, fatty acids, phospholipids, or cholesterol-related compounds.

[0097] Furthermore, the salts may be, but are not limited to, calcium chloride, calcium nitrate, magnesium chloride, magnesium sulfate, potassium chloride, and / or sodium chloride. One function of the salts is to adjust the molar osmotic concentration in the culture medium.

[0098] Preferably, the feed medium of the Disclosure in any of its embodiments has a reduced or low salt content. A reduced or low salt content means, for example, a total salt concentration of about 100 mM or less, preferably about 50 mM or less (e.g., a feed medium that does not contain sodium chloride, and a reduced concentration of potassium chloride).

[0099] The most important contributors to molar osmotic concentration are sodium ions, chloride ions, and bicarbonates, as well as glucose and other carbon sources, such as amino acids. For culture medium developers, the challenge is to produce highly concentrated nutrient mixtures and powder formulations for manufacture that meet the following requirements (preferably x times the concentration of basal medium components (positive impact in terms of supply chain management and regulation)), provide essential nutrients and nutrients that cannot be synthesized in appropriate amounts by the cells themselves (preferably as a reasonably balanced composition), overcome solubility issues for feed concentrates, remove bulk salts for molar osmotic concentration reasons, avoid toxic ranges, and design powder formulations that require carbon carriers for galenic reasons. Furthermore, in typical fed-batch processes, feed media need to be concentrated to minimize culture volume during the culture period. Bioreactor size can indeed impose constraints on feed, where only a total feed volume of about 30% (25-35%) of the initial culture volume is possible.

[0100] Carbohydrates may include, but are not limited to, glucose, mannose, galactose, fructose, sucrose, or glucosamine. These carbohydrates may be added directly to the culture medium or separately to the cell culture conditions. Other energy sources include, but are not limited to, sodium pyruvate.

[0101] Mammalian cells should be cultured at a neutral pH, for example, about pH 6.5 to about pH 7.5, preferably about pH 6.6 to about pH 7.3, and more preferably about pH 7. Therefore, a buffer should be added to the basal cell culture medium. For feed medium, the pH may be slightly outside this range, as the addition of the feed medium does not cause the pH of the cell culture solution to fall outside this range, because the feed medium is added as a concentrate. The preferred pH range for feed medium is about 6 to about 8. Suitable buffers include, but are not limited to, Hepes, phosphate buffers (e.g., monobasic and dibasic potassium phosphate and / or dibasic sodium phosphate anhydrous and monobasic sodium phosphate), phenol red, sodium bicarbonate, and / or sodium bicarbonate.

[0102] Generally, feed media contain nutrients consumed during cell culture conditions, such as amino acids and carbohydrates, but salts and buffers are less important. Therefore, some salts may be completely excluded from feed media.

[0103] Results of cell culture conditions The media of this disclosure improve the performance of cell culture conditions. As used herein, “improved performance of cell culture conditions” includes, for example, significantly improved product titer, improved cell proliferation (e.g., viable cell count, cell viability), and preferred phenotypic behavior of the cell culture condition process, such as reduced overflow metabolism of undesirable toxic byproducts (e.g., reduced lactate formation). It also contributes to a reduction in molar osmotic concentration levels in the cell culture condition process.

[0104] The media of this disclosure meet the cell-specific requirements and metabolic needs of mammalian cell culture conditions over time. In other words, it meets (iii) the life cycle of culture execution (which is approximately 10-20 days), (ii) the cell culture conditions system, and (i) the cell-specific needs of mammalian cells. Mammalian cells in a culture medium have different nutritional requirements at different stages of the cell culture conditions process. However, ideally, just one optimal basal medium and just one (or very few) optimal feed mediums / media should be designed to enable the design of a robust, safe, and efficient bioprocess. The media provided herein meet this need.

[0105] The culture media of this disclosure exhibit improved cell culture performance. Non-limiting examples of improved cell culture performance include increased product titer, improved viable cell concentration, and / or cell viability. Cell proliferation may also be improved, which is necessary for a series of inoculations in scaled-up procedures. For example, the scale of culture can be gradually increased from cell bank thawing (mL scale) to production scale (scale exceeding 10,000 L). The better the proliferation at each Nx stage (where N stage means the final production scale, and Nx usually means the cell proliferation stage before the final production stage in a batch system), the faster and better each transition to the next stage can be. Specifically, better cell proliferation and higher viable cell concentration allow Nx cultures to be performed in a shorter run time (and therefore faster). Also, better cell proliferation and higher viable cell concentration improve the transition, resulting in improved overall performance. For example, if a certain Nx stage should be inoculated at a certain inoculation cell density and the viable cell concentration is high, a relatively small volume of cell culture medium needs to be transferred from one stage to the next (the transfer of inoculation volume per culture starting volume is defined as the passage ratio, which is typically 1:5 to 1:20). This also means that only the reduced volume of "used" cell culture medium is transferred from one stage to the next, and the maximum volume of "new" medium can be added to the next stage (a constant total culture volume). This also improves the overall cell culture condition performance in the final N stage (e.g., increased product titer). Using the novel media provided in this disclosure, these All stages are improved. The positive effects of novel iron carriers, such as iron choline citrate and / or novel amino acid ratios, are not limited to basal and feed media in the final production stage. It is also shown that the positive effects of the medium platform, particularly the amino acid ratios, apply to the Nx stage as well. These positive effects are also maintained in the case of medium modifications in the Nx stage. For example, to maintain selective pressure in mammalian cell culture conditions using recombinant cell lines such as CHO cell lines, preferably CHO-DG44 cell lines, MTX (methotrexate) is typically provided in the early stages of a series of inoculations. In such examples, the basal media of this disclosure significantly improve the viable cell concentration.

[0106] Cell culture / feed medium In contrast to typical batch fermentation, where concentrated feed medium is not added to the culture medium at all during the entire culture period, standard fed-batch applications require the addition of a nutrient concentrate, which is referred to as "feed medium." In contrast to batch applications, it is well known that in fed-batch processes, cell culture conditions, such as maximum viable cell count, final product titer, and metabolic waste accumulation, are significantly improved by the supplementation of nutrients, vitamins, salts, and other components. Typically, the maximum amount of feed solution added to the culture medium during the culture time depends on technical circumstances, but also on metabolically driven conditions: the maximum capacity of the bioreactor constrains the total volume of feed to be added, while non-technical feed doses are applied to meet the actual cellular nutritional needs at any point during culture. Furthermore, depending on the cell line and process method, feed addition can be done continuously, for example, in small batches of 2-80 L (with minimal effort), at a constant feed rate of, for example, 5-60 ml feed / L / day, or with minimal risk of contamination. To achieve this, a discontinuous (large-scale production, requiring more effort) approach is used, allowing for large-scale additions, for example, in volumes of 2,000 to 10,000 L. For example, the typical feed addition interval during an 11-day fed-batch culture can vary between several times a day, once a day, or once every 2 to 4 days, often depending on the actual nutrient levels, growth stage, culture conditions, such as pH, or the nutrient needs of the culture medium.

[0107] Lactate / Carbon dioxide / Glucose The less-than-ideal nutrient burning of major carbon in most cell culture media can be determined by overflow metabolism. This means that glucose, the primary carbon source, is used inefficiently, leading to an increase in organic acids such as lactic acid. This elevated level of lactic acid causes the pH to drop below 6.65, which negatively impacts the buffering capacity of the culture medium and therefore can negatively affect viability in the culture medium. For these reasons, the CO2 concentration in the culture atmosphere is reduced at the start of the exponential-logarithmic growth phase to minimize acidity levels in the culture medium.

[0108] Cell lines and cell cultures The media of this disclosure can be applied to all mammalian cell lines. However, the media of this disclosure may also be suitable for other eukaryotic cells, such as yeast cells, plant cells, or insect cells. Mammalian cells according to this disclosure may be oocytes, embryonic stem cells, hematopoietic stem cells, or any type of differentiated cell. Preferably, mammalian cells are human, monkey, mouse, goat, cattle, sheep, pig cells, or rodent cell lines, such as rat, rabbit, or hamster. Mammalian cells may be isolated primary cells or cell lines. Preferred cell lines or "host cells" for the production of recombinant biopharmaceuticals are human, monkey, or rodent cell lines (mouse, rat, or hamster). Preferred human cells are PER.C6 or HEK293 cells.

[0109] More preferably are rodent cells, such as hamster cells, preferably BHK21, BHK TK-, Chinese hamster ovary cells (CHO), CHO-K1, and CHO-D XB11 (also known as CHO-DUKX or DuxB11), CHO-DUKX B 1. CHO-S, CHO-DG44, and CHO glutamine synthase (GS) deficient cells or derivatives / offspring of any of these cell lines. Particularly preferred are CHO-DG44, CHO-DUKX, CHO-K1, CHO-S, CHO-DG44GS deficient cell lines and BHK21, and even more preferred are CHO-DG44 cells, CHO These are GS-deficient cells (e.g., CHO-K1 GS-deficient cells) and CHO-DUKX cells. Furthermore, mouse myeloma cells, preferably NS0 and Sp2 / 0 cells, or derivatives / progeny of any such cell lines are also known as production cell lines for biopharmaceutical proteins.

[0110] All cells and cell lines can be used under all types of cell culture conditions, ranging from, for example, plastic microtita plates (on a scale of nL to mL) to industrial-scale stainless steel bioreactors (on a scale of L to kL), and also include all types of disposable systems and all types of process control strategies, ranging from uncontrolled systems to fully controlled systems, including, for example, advanced online monitoring and advanced control strategies. Suitable culture conditions for mammalian cells are known in the art. Mammalian cells may be cultured, for example, in suspension or attached to a solid-phase surface.

[0111] (Preferred embodiment) The following describes preferred embodiments, but it should be understood that these embodiments are illustrative of the Disclosure and the scope of the Disclosure is not limited to such preferred embodiments. Those skilled in the art will also understand that modifications, changes, etc., within the scope of the Disclosure can be easily made by referring to the following preferred embodiments. Therefore, it is clear that those skilled in the art can make appropriate modifications within the scope of the Disclosure by referring to the descriptions herein. Furthermore, it should be understood that the following embodiments of the Disclosure can be used individually or in combination.

[0112] (Quality standards) This disclosure provides a stem cell quality control method that includes determining whether a culture medium containing a component (e.g., X1) has a coefficient (e.g., C1) that correlates with it. In this disclosure, the flowcharts shown in Figures 1-7 can be referenced as non-limiting examples for quality standards and controls.

[0113] In one embodiment, the method of the present disclosure includes culturing cells in a medium containing a component, obtaining a coefficient associated with the component, and determining whether the medium containing the component is correlated based on the coefficient. Here, for example, for components: pyruvate, cystine, serine, the coefficient: mitochondrial size may be used; for example, for a component selected from the group consisting of PTEN inhibitors, p53 inhibitors, p38 inhibitors, Wnt signaling activators, and ROCK inhibitors (e.g., ROCK inhibitor Y-27632), the coefficient: Ror2 or Fzd5 expression level may be used as the coefficient.

[0114] In one embodiment, the determination includes determining whether the target cell is the cell of interest. Such determination can utilize any method known in the art.

[0115] In one embodiment, the components include any components described elsewhere in this specification or any components described in the referenced literature.

[0116] In one embodiment, the coefficient includes any coefficient described elsewhere in this specification or any coefficient described in any cited literature.

[0117] In one embodiment, the determination in this disclosure relates to the correlation between the component and the coefficient. This includes referencing databases that store information.

[0118] Such databases may include PubChem and ChEMBL. Alternatively, you may create your own custom database.

[0119] In this disclosure, as a specific embodiment, the determination in this disclosure is Controlling the behavior of one or more specific protein molecules related to the quality of the aforementioned cells, This includes controlling the expression behavior of a group of genes characteristic of the quality of the aforementioned cells, The aforementioned specific protein molecule is a cell surface antigen. The characteristic gene group for the aforementioned cells includes at least one selected from the group consisting of immune-related gene groups, inflammation-related gene groups, angiogenesis-related gene groups, nerve regeneration-related gene groups, chondrogenesis-related gene groups, bone formation-related gene groups, adipogenesis-related gene groups, fibrosis-related gene groups, immunogenicity-related gene groups, migration-related gene groups, adhesion-related gene groups, aging-related gene groups, mesenchymal stem cell marker-related gene groups, surface marker-related gene groups, growth factor-related gene groups, and chemokine or cytokine-related genes. In managing the expression behavior of the gene group characteristic of the aforementioned cells, if the correlation between the expression pattern of the gene group obtained by measuring the expression state of the gene group characteristic of the aforementioned cells and a standard expression pattern is above a certain level, the component is determined to be suitable for quality control. The correlation is considered to be above a certain level when the correlation coefficient or the coefficient of determination, or the correlation coefficient and the coefficient of determination, are evaluated and the correlation coefficient or the coefficient of determination is found to be above a certain level.

[0120] In one embodiment, when evaluating the correlation coefficient or coefficient of determination in this disclosure, (i) a missing value for which the gene expression level cannot be measured is replaced with a lower limit of measurement, (ii) a missing value for which the gene expression level cannot be measured is replaced with the lowest value among the measured values, or (iii) a missing value for which the gene expression level cannot be measured is replaced with the following formula: Read count for each gene + (1 × gene length / average gene length) The correlation coefficient or the coefficient of determination can be calculated by performing either of the following actions.

[0121] In one embodiment, the behavior of the specific protein molecule is at the expression level.

[0122] In one embodiment, the cells may be cell lines such as HeLa cells or A549 cells, or primary cells directly taken from living organisms (for example, keratinocytes taken from the skin or lungs). Collected alveolar epithelial cells), self-renewing and pluripotent stem cells (e.g., embryonic stem cells (ESCs), mesenchymal stem cells (MSCs), neural stem cells) may also be used. In addition, immune cells (T cells, B cells, natural killer cells (NK cells), etc.), hepatocytes, cardiomyocytes, fibroblasts, nerve cells, epithelial cells, and tumor cells, which are widely used in drug metabolism research and toxicity testing, may also be used. In one embodiment, the cells may be at least one type of cell selected from the group consisting of somatic stem cells, induced pluripotent stem cells, embryonic stem cells, tissue stem cells, differentiated cells derived from somatic stem cells, differentiated cells derived from induced pluripotent stem cells, differentiated cells derived from embryonic stem cells, and differentiated cells derived from tissue stem cells. In one preferred embodiment, the cells are mesenchymal stem cells.

[0123] While not particularly limited, the cells used are preferably at least one type of cell selected from the group consisting of somatic stem cells, induced pluripotent stem cells (iPS cells), embryonic stem cells (ES cells), differentiated cells derived from somatic stem cells, differentiated cells derived from induced pluripotent stem cells, and differentiated cells derived from embryonic stem cells.

[0124] As somatic stem cells, those present in bone marrow, blood, skin (epidermis, dermis, subcutaneous tissue), fat, hair follicles, brain, nerves, liver, pancreas, kidneys, muscles, and other tissues can be used. Preferably, mesenchymal stem cells can be used as somatic stem cells.

[0125] Mesenchymal stem cells are known to exist in bone marrow, synovial membrane, periosteum, adipose tissue, and muscle tissue, and are known to have the ability to differentiate into osteoblasts, chondrocytes, adipocytes, and muscle cells. Among the above mesenchymal stem cells, synovial-derived mesenchymal stem cells are known to have high cartilage formation ability.

[0126] Induced pluripotent stem cells (iPS cells) are cells that possess pluripotency (multipotency) and proliferative capacity, created by reprogramming somatic cells through methods such as the introduction of reprogramming factors. The somatic cells used to create iPS cells are not particularly limited; differentiated somatic cells or undifferentiated stem cells may be used.

[0127] In one embodiment, the target cells may be those used for drug discovery.

[0128] In one embodiment, the present disclosure provides a method for producing a culture medium or cells for therapeutic or drug discovery, comprising selecting a culture medium and, optionally, candidate cells from a population of cells including a culture medium and optionally, candidate cells, that meet certain criteria, and growing the selected cells in the selected culture medium.

[0129] The origin of the cells is not particularly limited; for example, cells from mammals such as humans, monkeys, mice, rats, guinea pigs, rabbits, cats, dogs, horses, cows, sheep, goats, and pigs can be used.

[0130] When cells controlled in the method of this disclosure are used for therapeutic purposes, the cells may be autologous or allogeneic, but are preferably autologous.

[0131] In this disclosure, differentiated cells derived from somatic stem cells, differentiated cells derived from induced pluripotent stem cells, or differentiated cells derived from embryonic stem cells may be used.

[0132] The conditions and procedures for culturing cells to induce differentiation in order to obtain differentiated cells are well known in the art.

[0133] The culture medium used for inducing the differentiation of stem cells is a basic medium containing components necessary for the survival and proliferation of stem cells (inorganic salts, carbohydrates, hormones, essential amino acids, non-essential amino acids, vitamins, and fatty acids), such as Dulbeco's Modified Eagle Medium (D-MEM), Minimum Essential Medium (MEM), RPMI1640, Basal Medium Eagle (BME), Dulbeco's Modified Eagle Medium:Nutrient Mixture F-12 (D-MEM / F-12), Glasgow Minimum Essential Medium (Glasgow MEM), or Hank's balanced salt solution, to which at least one differentiation-inducing factor according to the target cells is added. Examples of adipocyte differentiation-inducing factors include dexamethasone (DEX), 3-isobutyl-1-methylxanthine (IBMX), indomethacin (IDM), insulin (Ins), troglitazone, and biotin. Examples of osteoblast differentiation-inducing factors include dexamethasone (DEX), hydrocortisone, β-glycerophosphate, ascorbic acid, BMP4, and BMP2. Examples of chondrocyte differentiation-inducing factors include TGF-β3, dexamethasone (DEX), and ascorbic acid diphosphate. In addition, the above culture medium contains ingredients that increase the rate of cell proliferation. To increase the volume, growth factors such as basic fibroblast growth factor (bFGF) and epidermal growth factor (EGF), tumor necrosis factor (TNF), vitamins, interleukins, insulin, transferrin, heparin, heparan sulfate, collagen, bovine serum albumin (BSA), fibronectin, progesterone, selenite, B27 supplement, N2 supplement, ITS supplement, etc. may be added as needed, and antibiotics (penicillin, streptomycin, etc.) may also be added. Each component of the culture medium should be sterilized by an appropriate method before use. In addition to the above, it is preferable that the culture medium contains serum (for example, 10% fetal bovine serum (FBS)) at a concentration of 1-20%.

[0134] Differentiation-inducing media containing differentiation-inducing factors for target cells are commercially available, and these commercially available media may be used.

[0135] The culture temperature for inducing the proliferation and differentiation of stem cells varies depending on the cell origin, but for human-derived cells, for example, 30°C to 40°C is preferred, and 36°C to 38°C is more preferred. Furthermore, the CO2 gas concentration is preferably about 1% to 10%, and more preferably about 2% to 5%.

[0136] Confirmation of differentiation induction into the target cell type can be performed using known methods depending on the type of differentiated cell. For example, differentiation induction into adipocytes can be confirmed by staining cells with oil red O, measuring the amount of triglycerides in the cells, or measuring the expression level of the peroxisome proliferator-activated receptor-γ (PPARγ) gene in the cells. Differentiation induction into osteoblasts can be confirmed by quantifying the amount of calcium deposited in cells, staining cells with alkaline phosphatase, measuring the alkaline phosphatase activity in cells, or measuring the expression level of Osterix in cells. Differentiation induction into chondrocytes can be confirmed by quantifying the amount of glycosaminoglycans in cells, staining cells with Alcian blue, or measuring the expression level of Collagen type II in cells. This can be done by methods such as measuring the expression level.

[0137] In this disclosure, the behavior of one or more specific protein molecules can be controlled by any method. Specifically, controlling the behavior of specific protein molecules means controlling the expression level of those specific protein molecules.

[0138] The expression level of a specific protein molecule refers to the amount of expression of the gene for a specific protein, or the amount of expression of the specific protein itself. The expression level of a specific protein molecule can be calculated as an absolute value or a relative value (such as a ratio or difference from a comparison control or reference expression level).

[0139] The expression level of a specific protein molecule can be measured by any method known to those skilled in the art. The procedure can be carried out according to the standard method. As a way to measure the expression level of a specific protein molecule, the amount of mRNA, which is the transcript of the gene, may be measured. The method for measuring mRNA is not particularly limited as long as it can measure the desired amount of mRNA, and can be appropriately selected from known methods. For example, a gene amplification method using oligonucleotides that hybridize to the gene encoding the specific protein molecule as primers, or a hybridization method using oligo(poly)nucleotides that hybridize to the gene encoding the specific protein molecule as probes can be used. Specifically, examples include RT-PCR (reverse transcription polymerase chain reaction), real-time RT-PCR, DNA microarray, cell array, Northern blotting, dot blotting, and RNase protection assay.

[0140] The primers and probes used in the above measurement method can be labeled, and the amount of mRNA can be measured by examining the signal intensity of the label. Real-time RT-PCR can use RNA directly in the sample and optically measure the gene amplification process, which is necessary for amplification. This method is preferable because it allows for gene quantification based on the number of temperature cycles. Furthermore, as a control, the expression levels of housekeeping genes such as glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and beta-actin mRNA can be used to standardize the expression levels of genes encoding specific protein molecules. The primers and probes used in the above measurement method can be appropriately designed and prepared by those skilled in the art based on the nucleotide sequence information of the genes encoding specific protein molecules.

[0141] Protein expression levels can be measured immunologically, for example, using antibodies or antibody fragments against specific proteins. Specifically, this can include flow cytometry, Western blotting, enzyme-linked immunosorbent assay (ELISA), radioimmunoassay (RIA), immunofluorescence assay, and cell array assay. These measurement methods can also be performed using standard protocols or protocols that have been appropriately modified or changed from standard protocols.

[0142] For example, when measuring the expression level of a protein in a cell by flow cytometry, if the positive rate of a specific protein is preferably 70% or higher, more preferably 80% or higher, and even more preferably 90% or higher, the cell quality can be evaluated as high and the compatibility of the culture medium or its components is high.

[0143] In one embodiment, quality control can be performed, for example, by comparing the expression level of a specific protein molecule in cells measured by the method described above with a predetermined reference expression level. The reference expression level may be, for example, the expression level of a specific protein molecule in cells that have already been confirmed to have a certain quality (positive control), or the expression level of cells that have already been confirmed not to have a certain quality (negative control).

[0144] By comparing the expression level of a specific protein molecule in a cell with its reference expression level, if the expression level of the specific protein molecule in the cell is equal to or higher than the expression level of the positive control, the cell quality can be evaluated as high. Conversely, if the expression level of the specific protein molecule in the cell is equal to or lower than the expression level of the negative control, the cell quality can be evaluated as low.

[0145] Alternatively, a cutoff value for the expression level of a specific protein molecule may be set in advance, and the expression level of the specific protein molecule measured for the cells may be compared with the cutoff value. The cutoff value can be, for example, the expression level of the specific protein molecule that gives the desired effect, such as the quality of the cells or component, based on a regression line that shows the correlation between the expression level of the specific protein molecule and the desired effect, such as the quality of the cells or component. For example, if the expression level of the specific protein molecule in the cells is above the cutoff value, it can be evaluated that the compatibility between the culture medium or its components and the target cells is good, or that the quality of the cells is high. If it is below the cutoff value, it can be evaluated that the compatibility between the culture medium or its components and the target cells is poor, or that the quality of the cells is low.

[0146] Controlling the expression behavior of cell-specific gene groups for the purpose of quality control of the culture medium or its components can be carried out by any method.

[0147] The cell-specific group of related genes preferably consists of 10 to 5000 genes. More preferably, it consists of 10 to 2000 genes. The lower limit of the number of genes may be 15 or more, 20 or more, 30 or more, 40 or more, 50 or more, 100 or more, 200 or more, 300 or more, or 500 or more. The upper limit of the number of genes may be 5000 or less, 4000 or less, 3000 or less, 2000 or less, or 1500 or less.

[0148] The characteristic gene groups for cells are not particularly limited and can be appropriately selected depending on the type of cell. For example, if the cells are mesenchymal stem cells, then at least one of the following can be selected (for example, gene groups 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, or 16 of the gene groups listed above).

[0149] The various gene sets described above can be collected, for example, using commercially available panels, literature information, microarray data, and publicly known databases (e.g., the Ontology database (https: / / www.informatics.jax.org / vocab / gene_ontology / )). From a set of genes, genes that appear frequently can be selected and used.

[0150] In quality control, the usability of cells can be determined by the correlation between the expression pattern of a gene group characteristic of the cells, obtained by measuring its expression status, and a reference expression pattern. Specifically, if the correlation between the expression pattern of a gene group characteristic of the cells, obtained by measuring its expression status, and a reference expression pattern is above a certain level, it can be determined that the compatibility between the culture medium or its components and the target cells is high, the quality is maintained, or the cells are usable. For example, by evaluating the correlation coefficient or coefficient of determination, or both, between the expression pattern of a gene group characteristic of the cells, obtained by measuring its expression status, and a reference expression pattern, if the correlation coefficient or coefficient of determination is above a certain level, it can be determined that the compatibility between the culture medium or its components and the target cells is high, the quality is maintained, or the cells are usable.

[0151] The method for measuring expression status is not particularly limited, but the amount of mRNA can be measured. The method for measuring the amount of mRNA is not particularly limited as long as it can measure the desired amount of mRNA, and can be appropriately selected from known methods. For example, RNA sequencing (RNA-Seq), RT-PCR, and real-time RT-PCR can be used.

[0152] There are various methods for calculating the coefficient of determination, and it is not particularly limited. For example, one method can be used with the RSQ function included in Microsoft Excel. The RSQ function returns the value of r squared (coefficient of determination) for a regression line using (x, y) pairs of the total gene population, where y is the expression level of a certain gene in the first target sample and x is the expression level of the same gene in the second target sample. This value represents the coefficient of determination between two target samples. There are various methods that can be used to calculate the coefficient, and there are no particular limitations. For example, one method can be used that utilizes the CORREL function used in Microsoft Excel (registered trademark). The CORREL function returns the correlation coefficient value for (x, y) pairs of the total gene population, consisting of the expression level y of a certain gene in the first target sample and the expression level x of the same gene in the second target sample, thereby providing the correlation coefficient between the two target samples.

[0153] When evaluating the correlation coefficient or the coefficient of determination, (i) Substitute missing values ​​for the lower limit of measurement for gene expression levels that cannot be measured. (ii) Substitute missing values ​​for which gene expression levels cannot be measured with the lowest value among the measured values, or (iii) Missing values ​​for which gene expression levels cannot be measured are handled using the following formula: read count for each gene + (1 × gene length / average gene length) It is preferable to calculate the correlation coefficient or the coefficient of determination by performing one of the following actions.

[0154] RNA sequencing (RNA-Seq) is a method for quantifying gene expression levels using a sequencer. The analysis process involves (1) performing quality control on the reads output from the sequencer, (2) mapping the reads to a reference sequence, and (3) counting how many reads were mapped to each gene region. The number of reads counted for each gene region in this way can be considered as the relative expression level of that gene. In other words, the read count for each gene is a result that is generally obtained from gene expression analysis and is a value that is counted for each gene region.

[0155] The threshold for the coefficient of determination or correlation coefficient can be selected from, for example, 0.70 or higher, 0.71 or higher, 0.72 or higher, 0.73 or higher, 0.74 or higher, 0.75 or higher, 0.76 or higher, 0.77 or higher, 0.78 or higher, 0.79 or higher, 0.80 or higher, 0.81 or higher, 0.82 or higher, 0.83 or higher, 0.84 or higher, 0.85 or higher, 0.86 or higher, 0.87 or higher, 0.88 or higher, 0.89 or higher, etc., depending on the cell types being distinguished.

[0156] This disclosure provides a cell quality control method that, in one example, involves determining a coefficient showing the correlation between a culture medium containing a component and the characteristics of stem cells. This method allows for the quantitative evaluation of the impact of a component on stem cell quality and the establishment of optimal culture conditions. Therefore, cells whose quality has been confirmed by this disclosure can be used for regenerative medicine or basic research (e.g., drug discovery). For example, cells whose quality has been confirmed by this disclosure can be used as transplant material for the treatment of tissue damage or impairment, or for cosmetic purposes. In one example of this disclosure, the cells are used as a drug for treating arthritis. For example, the cells may be used as a drug for treating meniscus defects or osteoarthritis.

[0157] In another example of this disclosure, cells can be used for drug discovery. For example, cells quality-controlled by the methods of this disclosure can be used to evaluate the efficacy or toxicity of candidate drugs and screen for candidate drugs. For instance, the efficacy or toxicity of a candidate drug can be evaluated by comparing the properties of cells when they are in contact with the candidate drug with those when they are not.

[0158] (Design Methods (Screening) · Business) In one aspect, this disclosure provides a method for optimizing culture medium components and cell culture conditions. It concerns the optimization of culture medium components and cell culture conditions in cell culture conditions, particularly contributing to methods for optimizing culture medium components and conditions to promote cell proliferation and differentiation and improve cell quality. Cell culture conditions are a crucial technology in many fields, including biotechnology, regenerative medicine, and drug screening. The composition of the culture medium and culture conditions play a vital role in optimizing cell proliferation and differentiation. This disclosure provides a method for efficiently finding the optimal culture medium components and conditions. This disclosure concerns a method for optimizing culture medium components and cell culture conditions, specifically providing procedures for adjusting the components of the culture medium and optimizing the culture conditions. According to this disclosure, it is possible to improve cell proliferation rate, differentiation potential, and quality. In this disclosure, flowcharts shown in Figures 8-14 can be referenced as non-limiting examples for optimization, etc.

[0159] <Wet> In one aspect, this disclosure relates to a computer receiving cells provided by a user. The present invention provides a method for performing the selection of appropriate culture medium components and / or cell culture conditions. This method includes the steps of: inputting cell information provided by the user into the computer; causing the computer to derive culture medium components and culture conditions related to the cell, and optimal conditions, by consulting a database as necessary; causing the computer to calculate the optimal culture medium components and cell culture conditions for the cell based on the culture medium components and culture conditions; and (if necessary) displaying the results of the calculation on a display. In this disclosure, the flowcharts shown in Figures 8-14 can be used as non-limiting examples for selecting and optimizing appropriate culture medium components and / or cell culture conditions.

[0160] In one embodiment, the step of inputting cell information provided by the user into the computer can be carried out by any method known in the art. For example, this could involve inputting data obtained in a wet state using an input device to the computer, or reading from a recording medium in which already measured results are stored and inputting them into the target computer.

[0161] In one embodiment, the step of having the computer, if necessary, consult a database to derive culture medium components and culture conditions related to the cells, as well as optimal conditions if necessary, can be carried out by any method known in the art. More specifically, any method for obtaining any culture medium components, culture conditions, and optimal conditions related to the cells can be used. For example, this could involve searching using a query related to the cells and outputting information regarding culture medium components and culture requirements from the search results.

[0162] In one embodiment, the step of having the computer calculate the optimal culture medium components and cell culture conditions for the cells based on the culture medium components and culture conditions can be carried out by any method known in the art. More specifically, this includes having the computer retrieve input cell information, culture medium components and culture conditions for the cells, and, if necessary, optimal conditions, and, if necessary, calculate appropriate parameters. A database may be consulted as appropriate. The database may be a public database or a closed database developed by the user.

[0163] In one embodiment, the database is a library containing cytokines, growth factors, proteins, small molecules, and bioactive substances useful for cell culture conditions, which can be used to compose the culture medium.

[0164] In one embodiment, the database is a library that collects various information on cell culture conditions and makes it available for use in configuring the culture medium.

[0165] In one embodiment, the step of displaying the calculation results on a display can be carried out by any method known in the art. The display can be in any format, but examples include displaying a single optimal condition derived from the calculation results, displaying multiple candidate optimal conditions, displaying multiple candidate optimal conditions along with a recommended experimental plan, or displaying the calculation results along with a form that allows the user to input results obtained from wet experiments conducted based on those conditions and update the calculation results.

[0166] In one embodiment, the process includes storing the results of the calculation and information including the culture results, The process of selecting information stored in the storage unit, A step of calculating culture conditions based on the selected information in the storage unit, A process of documenting the culture conditions calculated by the calculation unit according to the standards as needed, This can include a process for generating documented results.

[0167] CDMO operators typically culture cells on a small scale and provide them to their customers. Cell culture conditions are optimized based on information from the customer. Small-scale cell culture conditions may differ from commercial-scale cell culture conditions, and CDMO operators may either optimize small-scale cell culture methods or develop commercial-scale cell culture conditions based on the customer's development status.

[0168] CDMO operators may develop culture conditions that can be used for commercial-scale cell culture based on small-scale cell culture conditions provided by customers, optimize the culture conditions if necessary, culture cells using this method, and provide customers with information on the optimized culture conditions. CDMO operators may also identify foreseeable risks related to the culture conditions, formulate countermeasures for those risks, and provide customers with information on the risks and countermeasures. CDMO operators may also document the culture conditions in consideration of standards, formulate and print draft Manufacturing Records (MBRs) and draft Standard Operating Procedures (SOPs), and provide them to customers.

[0169] In one embodiment, the database includes information on cells provided by the customer, as well as specifications of automated culture equipment owned by the CDMO operator, the number of people required depending on the culture conditions, the layout of the manufacturing plant, information on the flow of movement during culture, and information on the reagents owned. The CDMO operator outputs the optimal culture conditions (including the culture medium to be used) for cell culture from the database. The culture conditions may be those optimized for small-scale cell culture, those optimized for commercially scale cultured cells, or the culture conditions may be formulated in stages from small scale to commercial scale.

[0170] The culture components and / or culture conditions proposed to the customer may be those used with an automated culture system. The culture components may be the same as or different from those used in the small-scale case. The culture conditions may include information such as the area of ​​the manufacturing facility, the configuration and arrangement of the automated culture system, and the movement of people and objects (flow paths) during cultivation. The automated culture system may be a system that automates cultivation, which is one aspect of the present invention. Furthermore, the culture conditions may include information on manufacturing costs obtained from a cost calculation system.

[0171] Risks that CDMO operators may provide to customers include the risk of changes in the state or quantity of cultured cells during scale-up, the risk of non-compliance with GMP, and the risk of unsuccessful preparation of reagents during culture. Risk prediction may include visualization of risks when preparing culture medium components, including powder preparation and on-demand preparation, prediction of risks related to the quality and lot variations of the culture medium, and regional risks, based on database information. Risk mitigation measures may be output based on database information. Output information may include suggestions for compatible culture medium factors, suggestions for managing and supplying culture medium components, automatic generation of risk reduction documents, and suggestions for routine responses that tolerate biogroups. Given concerns that the supply of culture medium components may be suddenly reduced or stopped, it is preferable to propose alternative culture methods and consider culture conditions.

[0172] CDMO operators may develop and propose a roadmap to commercialization based on information from their customers. This may include proposing culture conditions for commercial-scale cell culture, suggesting equipment and materials to be used for culture, proposing alternative methods or modifications to culture conditions as risk mitigation measures, and providing draft MBRs and SOPs.

[0173] In one embodiment, the customer receives an initial culture method proposal from the CDMO operator, which covers the entire process up to commercial production. The proposal includes information on cell production volume, production efficiency, culture medium component costs, and culture medium component composition, as well as suggestions for changes to the culture medium components, suggested culture conditions, alternative methods, proposed MBRs, proposed SOPs, visualized production site information, and a roadmap.

[0174] In one embodiment, the selection is (A) A step of selecting target cells; (B) A step of selecting the basic cellular functions (K) of the said cells; (C) A step of determining a set of medium factors (N) that determine the environment containing the components of the medium; (D) Using information stored in a database related to the cells, construct a functional map representing the intensity of activation or suppression of each of the K basic cellular functions by each of the N culture medium factors; (E) Using the functional map, the step of optimizing the composition of the cell culture medium so as to activate or suppress one or more of the basic cellular functions; This includes the expression, where K and N are arbitrary integers that are independent of each other.

[0175] In another aspect, the present disclosure provides a method for performing calculations to select appropriate culture medium components and / or cell culture conditions for cells provided by a user, the method comprising the steps of analyzing the cells provided by the user using candidate culture medium components and / or cell culture conditions, The present invention provides a method comprising the steps of: deriving culture medium components and culture conditions (optimal requirements as necessary) related to the cells from the analysis results; and calculating appropriate culture medium components and cell culture conditions for the cells based on the culture medium components and culture conditions. In this disclosure, the flowcharts shown in Figures 8-14 can be used as non-limiting examples for calculating and optimizing the selection of appropriate culture medium components and / or cell culture conditions.

[0176] In one embodiment, the step of analyzing cells provided by the user using candidate culture medium components and / or cell culture conditions can be carried out by any method known in the art. For example, it is conceivable that culture experiments could be performed using the actual provided cells with candidate culture medium components and / or cell culture conditions, and the data collected and analyzed. Such culture experiments may be performed by applying the cells to multi-well plates and conducting high-throughput screening using an automated system. Candidate culture medium components and / or cell culture conditions may be provided as a library.

[0177] In one embodiment, the step of deriving the culture medium components and culture conditions (optimal requirements as needed) related to the cells from the analysis results can be carried out by any method known in the art. Such derivation may involve scoring and ranking the analysis results according to the desired purpose, but is not limited to these methods.

[0178] In one embodiment, the process of calculating appropriate culture medium components and cell culture conditions for the cells based on the culture medium components and culture conditions can be carried out by any method known in the art. From the derived results, it is conceivable that the culture medium components and culture conditions (optimal requirements as needed) that rank highly can be selected according to the desired objective. Desired objectives include, but are not limited to, selecting a culture medium that provides excellent growth regardless of cost, a culture medium that provides good growth within a certain cost, or a culture medium that provides good growth per unit of cost by calculating the cost-benefit ratio.

[0179] In one embodiment, the selection is (A) The step of selecting target cells; (B) A step of selecting the basic cellular functions (K) of the cells; (C) A step of determining a set of medium factors (N) that determine the environment containing the components of the medium; (D) Using experimental data of novel culture medium samples and / or the supernatant of existing culture medium samples using the components of the culture medium and the cells, a functional map is constructed that represents the intensity of activation or inhibition of each of the K basic cellular functions by each of the N culture medium factors. (E) Using the functional map, the step of optimizing the composition of the cell culture medium so as to activate or suppress one or more of the basic cellular functions; Includes.

[0180] In one embodiment, the candidate culture component is selected from a library of the culture component. As used herein, the term “library” is intended to mean a collection containing several heterogeneous units. Units within a collection may differ in structure and / or function. For example, a collection may contain nucleic acids having different nucleotide sequences, or a collection may contain proteins having different primary (i.e., amino acid sequences), secondary, tertiary, or quaternary structures. However, it is understood that some redundancy of units may exist within a library. For example, even in a library containing a wide variety of heterogeneous nucleic acids or proteins, multiple copies of a particular nucleic acid or protein may exist. Exemplary types of units that may be present in a library include those described herein with respect to candidate drugs or screening agents. Libraries in this disclosure include libraries of small molecules, libraries containing peptides or proteins, and libraries containing nucleic acids, or combinations thereof. Other examples include libraries of peptide-nucleic acid complexes, cyclic peptide compounds, or cyclic peptide-nucleic acid complexes, and libraries of cyclic peptide-mRNA complexes. Display libraries are preferred as libraries. Examples of display libraries include libraries that utilize displays, among others, mRNA display libraries, DNA display libraries, and ribosome display libraries. The term "low molecular weight" as used herein is intended to mean compounds having a molecular weight of less than approximately 1000 daltons. In certain embodiments, low molecular weights are nonpolymerizable. However, in other embodiments, low molecular weights may be dimers or trimers. It will also be understood that low molecular weights may be monomers that can be incorporated into polymers. Particularly useful low molecular weights are organic compounds. Useful low molecular weights may have molecular weights of less than 900, 800, 600, 400, 200, or 100 daltons.

[0181] In one embodiment, the candidate cell culture conditions are selected from a library of cell culture conditions. In one embodiment, the analysis is performed based on the results of actual experiments using the culture medium components, or the analysis is performed based on the results of actual experiments using the culture medium components and calculations based on those results.

[0182] In one embodiment, the analysis involves applying the library of culture medium components to cells in a multi-well plate and performing high-throughput screening using an automated system.

[0183] The term "high-throughput screening" or "HTS" refers to methods that enable rapid, highly simultaneous biological research and drug discovery using various techniques and methods, such as optics, chemistry, biology, or image analysis. HTS methods are well known in the art and are generally carried out in multiwell plates equipped with automated liquid handling and detection devices, but the methods of the present invention may also be carried out on microarrays or in microfluidic systems.

[0184] In one embodiment, the calculations in this disclosure include performing calculations using a library parameterized for the performance and / or cost of the culture components on cells.

[0185] In one embodiment, the method described above achieves optimization of the culture medium to suit the purpose of seed cells or cell lines, cost, and growth performance.

[0186] In one embodiment, when performing wet optimization, step D) constructing a functional map representing the intensity of activation or inhibition of each of the K basic cellular functions by each of the N culture factors, using experimental data of the supernatant of a novel culture medium sample and / or an existing culture medium sample using the components of the medium and the cells, is a step of performing a first run of culture experiments including a number of cultures of (N+1) or more, where each culture is performed with different culture medium components and combinations of low-level and high-level culture factor components are screened; For each culture experiment performed, the steps include obtaining initial and final biomass, product, and exometabolome data, or partial exometabolome data; Relative weighting coefficient λ j The steps include determining a subset of basal cellular functions with a value greater than zero by regression analysis of the exometabolome data or exometabolome data induced against culture medium component data using the following linear model, and

[0187]

number

[0188]

number

[0189] (where v is a vector representing the rate of change of the component of the exometabolome from which the element was measured, I i,j This is the activation intensity parameter of basic cellular function j by culture medium factor i, determined by regression analysis.

[0190] A step of performing a second run of a culture experiment, the number of cultures being greater than or equal to the number of activated cell functions plus 1, wherein each culture is performed with a different culture medium component, the medium component having a predetermined intensity parameter value I such that low and high values ​​of the weighted coefficient of activated basic cell function are identified as subsets of activated cell functions controlled by the medium factor. i,j The steps set up to screen using;

[0191] In all of the above steps, a functional map is constructed from the data collected by linear regression analysis using equations (3a) and (3b), and the data is organized into the form of the functional map, wherein the intensity value I determined by the first run of the experiment is i,j However, this is corrected by the data from the second run of the experiment, resulting in an N×K data array (functional map = {I i,j This includes the step of shaping it into a form.

[0192] The optimization of the culture medium components in step e) is A step of forming basic cell function special culture medium components using the matrix of functional data, wherein the change in the value of the culture medium factor Δ(FAC j )

[0193]

number

[0194] The steps include: determining the change in the relative weights Δλi of basic cellular functions according to;

[0195] The steps include: forming culture medium components to enhance or suppress a single basic cellular function j using formula (4) applied to the j-th column of the functional data array; The steps include: forming the culture medium components and manipulating cellular metabolism by increasing or suppressing a critical set of basic cellular functions using formula (4), which is simultaneously applied to multiple columns of the functional data array; Includes.

[0196] In certain embodiments, the target biological structure is a cell tissue, a whole cell, an organelle, or a coherent set of biochemical transformations exhibiting a predetermined cellular function.

[0197] In certain embodiments, the target biological structure is genetically modified, including gene modifications directed towards the activation or suppression of the basic cellular function.

[0198] In certain embodiments, the culture medium factor is the physicochemical properties of a mixture of solids and / or liquids and / or gases of essential nutrients and / or micronutrients and / or biologically functional molecules, the release rate of the compound, or the feeding rate of the compound.

[0199] In certain embodiments, the physicochemical properties are temperature, and / or pressure, and / or pH, and / or ionic strength, and / or concentration, and / or activity, and / or volume molar osmotic concentration, and / or gravimetric osmotic concentration, and / or related properties.

[0200] In certain embodiments, essential nutrients and / or micronutrients and / or biologically functional molecules are inorganic and / or organic substances, including salts and / or vitamins, and / or metabolic cofactors and / or antibiotics and / or carbohydrates and / or lipid substances and / or protein substances and / or nucleotide substances and / or signaling proteins and / or molecules that inhibit enzyme activity and / or protein-active molecules and / or gene transformation modulators and / or interfering ribonucleic acids and / or complex mixtures of said substances with known or unknown compositions including serum, pure hydrosilates, or complex organic substances.

[0201] In a particular embodiment, the formula for the culture medium is: Formula for culture medium = {FAC j},j=1,···,N(however, FAC j This is the value of the culture medium factor j. ) is used to determine the value of N culture medium factors.

[0202] In a particular embodiment, the target biological structure is defined as target biological structure = {e i},i=1 ,···,K(however, e i This is a vector with q elements, and the values ​​of the elements are basic cells. This represents the weighted coefficients for each biochemical reaction in function i. It is determined by q biochemical reactions and K basic cellular functions.

[0203] In certain embodiments, the analysis includes classification, and / or the derivation includes a reference to a database (including the learning results).

[0204] In another aspect, a method for providing appropriate culture medium components and / or cell culture conditions for cells provided by a user using a computer, the method comprising: inputting information about the cells provided by the user into the computer; causing the computer to derive appropriate culture medium components and culture conditions for the cells by referring to a database containing the cell information; and, if necessary, displaying the results of the calculation on a display. Provides a method for including this. In this disclosure, the flowcharts shown in Figures 8-14 can be referenced as non-limiting examples for providing and optimizing appropriate culture medium components and / or cell culture conditions.

[0205] In a particular embodiment, the process includes recording the derived appropriate culture medium components and culture conditions in the database, and further, when a query for appropriate culture medium components and / or cell culture conditions for the cells is received, retrieving the appropriate culture medium components and / or cell culture conditions recorded in the database, and, if necessary, displaying the results of the retrieval on a display.

[0206] In a particular context, a program that codes a method for causing a computer to perform the selection of appropriate culture medium components and / or cell culture conditions for cells provided by a user, the method comprising the step of inputting information about the cells provided by the user into the computer, The present invention provides a program that includes the steps of: causing the computer to, if necessary, refer to a database to derive culture medium components and culture conditions related to the cells, as well as optimal conditions if necessary; causing the computer to calculate the optimal culture medium components and cell culture conditions for the cells based on the culture medium components and culture conditions; and, if necessary, displaying the results of the calculation on a display. In this disclosure, the flowcharts shown in Figures 8-14 can be referenced as non-limiting examples for optimization and the like.

[0207] A particular embodiment may be a system, method, or computer program product, or a combination thereof, at any possible level of technically detailed integration. The computer program product may include a recording medium (or more mediums) having computer-readable program instructions thereon for a processor to perform one or more aspects of the embodiment. The computer-readable recording medium may be a tangible device capable of holding and storing multiple instructions for use by an instruction execution device. The computer-readable medium may be, for example, but not limited to, an electronic recording device, a magnetic recording device, an optical recording device, an electromagnetic recording device, a semiconductor recording device, or any preferred combination thereof. Not limited embodiments of computer-readable recording media include portable computer disks, hard disks, random access memory (RAM), read-only memory (ROM), erasable programmable read-only memory (EPROM or flash memory®), static random access memory (SRAM), portable compact disk read-only memory (CD-ROM), digital multipurpose disk (DVD), memory stick, floppy disk®, punch cards or mechanically encoded devices having a structure protruding into grooves on which instructions are recorded, and any preferred combination thereof. As used herein, computer-readable recording media are not interpreted as transient signals themselves, such as radio waves or other freely propagating electromagnetic waves, waveguides or other communication media (e.g., light pulses passing through optical fiber cables), or electrical signals transmitted through wires. In this regard, various embodiments used herein Such computer-readable recording media may include tangible, non-transient computer-readable recording media.

[0208] The computer programs described herein can be downloaded from a computer-readable recording medium to a computing / processing device, or downloaded to an external computer or external recording device via a network, such as the Internet, a local area network, a wide area network, or a wireless network, or a combination thereof. The network may include copper communication cables, optical fiber, wireless communications, routers, firewalls, switches, gateway computers, and edge servers, or a combination thereof. A network adapter card or network interface in each computing / processing device receives computer-readable program instructions from the network and transfers these computer-readable program instructions to a computer-readable recording medium within the computing / processing device for storage. Computer-readable program instructions for performing operations of one or more embodiments may be assembler instructions, instruction set architecture (ISA) instructions, machine language instructions, machine-dependent instructions, microcode, firmware instructions, state setting data, configuration data for integrated circuits, or source code or object code written in any combination of programming languages, including object-oriented programming languages ​​such as Smalltalk®, C++, the “C” programming language, or similar programming languages. Computer-readable program instructions may be executed entirely on a user computer, partially on a user computer as a standalone software package, partially on a user computer and partially on a remote computer, or entirely on a remote computer or server.In the latter scenario, the remote computer can connect to the user computer through any type of network, including a local area network (LAN) or a wide area network (WAN), or the connection can be made to an external computer (e.g., through an Internet service provider). In some embodiments, electronic circuits, including, for example, a programmable logic circuit, a field-programmable gate array (FPGA), or a programmable logic array (PLA), can be personalized to execute computer-readable program instructions using computer-readable program instruction state information to perform one or more aspects of the present embodiment.

[0209] One or more aspects described herein have been described in accordance with embodiments of the present invention, with reference to flowchart instructions and block diagrams of methods, or both, devices (systems), and computer-readable recording media and computer programs. It is understood that any combination of flowchart illustrations and block diagrams, or both, and blocks and block diagrams, or both, in flowchart illustrations can be implemented by computer-readable program instructions. These computer-readable program instructions can be provided to a general-purpose computer, a specific-purpose computer, or other programmable data processing device for generating other processors or machines, and execution by the computer processor or other programmable data processing device generates means for implementing functions / operations specified by blocks or multiple blocks or combinations thereof in flowcharts and block diagrams. These computer-readable program instructions that instruct computers, programmable data processing devices and other devices or combinations thereof to function in a particular way can also be stored in computer-readable recording media, and computer-readable recording media containing the instructions can be used to store blocks or combinations thereof in flowcharts and block diagrams. A manufactured product comprises instructions that implement the functional / operational features specified by multiple blocks or combinations thereof. Computer-readable program instructions are also loaded onto a computer, other programmable data processing device, or other device, causing a computer implementation process for a series of operation steps on the computer, other programmable device, or other device, thereby implementing the functional / operational features specified by blocks or multiple blocks or combinations thereof in a flowchart and block diagram on the computer, other programmable device, or other device.

[0210] In another aspect, the Disclosure provides a recording medium storing a program that codes for a method of causing a computer to perform the selection of appropriate culture medium components and / or cell culture conditions for cells provided by a user. The method performed by the program stored on the recording medium includes the steps of: inputting information about the cells provided by the user into the computer; causing the computer to derive culture medium components and culture conditions related to the cells, and optionally optimal conditions, by consulting a database as necessary; causing the computer to calculate the optimal culture medium components and cell culture conditions for the cells based on the culture medium components and culture conditions; and (if necessary) displaying the results of the calculation on a display. In this disclosure, the flowcharts shown in Figures 8-14 can be referenced as non-limiting examples for optimization and the like.

[0211] In another aspect, the Disclosure provides a system for selecting appropriate culture medium components and / or cell culture conditions for cells provided by a user. The system includes a cell information input unit for inputting information about the cells provided by the user; a culture medium component / culture condition deriving unit which, if necessary, consults a database to derive culture medium components and culture conditions related to the cells, and if necessary, optimal conditions; an optimal calculation unit which calculates the optimal culture medium components and cell culture conditions for the cells based on the culture medium components and culture conditions; and (if necessary) a display unit which displays the results of the calculation on a display. In this disclosure, the flowcharts shown in Figures 8-14 can be referenced as non-limiting examples for optimization and the like.

[0212] For example, in one embodiment, there is a storage unit that stores information including the calculation results and culture results, and a selection unit that selects the information stored in the storage unit. A calculation unit that calculates culture conditions based on the selected information in the storage unit, A calculation unit that documents the culture conditions calculated by the calculation unit according to the standards as needed, The system may include a generation unit that generates the results documented by the calculation unit. In this system, the calculation unit is responsible for generating the calculation results as a document, and the generation unit is responsible for printing, displaying, or recording the documented results onto a recording medium such as a CD-R.

[0213] In one embodiment, a cell information input unit that inputs cell information provided by the user can be constructed using techniques known in the art.

[0214] In one embodiment, a culture medium component / culture condition deriving unit, which consults a database as needed to derive culture medium components and culture conditions related to the cells, as well as optimal conditions as needed, can be constructed using techniques known in the art.

[0215] In one embodiment, an optimal calculation unit that calculates the optimal culture medium components and cell culture conditions for the cells based on the culture medium components and culture conditions can be constructed using techniques known in the art.

[0216] In one embodiment, a display unit for displaying the results of the calculation on a screen (if necessary) can be constructed using art known in the art.

[0217] <Cost Calculation> In another aspect, the Disclosure provides a method for causing a computer to perform a calculation of the cost of culture medium components and / or cell culture conditions for cells provided by a user, the method comprising: inputting information of the cells provided by the user into the computer; and, if necessary, consulting a database, the computer to calculate the cost of the culture medium components related to the cells. The present invention provides a method comprising the steps of: causing the computer to calculate culture conditions and, if necessary, optimal conditions; causing the computer to calculate the cost of the culture medium components and the cell culture conditions based on the culture medium components and the culture conditions; and (if necessary) displaying the cost on a display. This method can be implemented in combination with one or more features of other embodiments described elsewhere in this specification. In this disclosure, the flowcharts shown in Figures 15-22 can be used as non-limiting examples for calculating costs and expenses.

[0218] In one embodiment, the selection is (A) The step of selecting target cells; (B) A step of selecting the basic cellular functions (K) of the cells; (C) A step of determining a set of medium factors (N) that determine the environment containing the components of the medium; (D) Using information stored in a database related to the cells, construct a functional map representing the intensity of activation or suppression of each of the K basic cellular functions by each of the N culture medium factors; (E) Using the functional map, the step of optimizing the composition of the cell culture medium so as to activate or suppress one or more of the basic cellular functions; Includes.

[0219] In one aspect, the Disclosure provides a method for performing a calculation of the cost of culture medium components and / or cell culture conditions for cells provided by a user. The method includes the steps of: analyzing the cells provided by the user; deriving culture medium components and culture conditions related to the cells from the results of the analysis; and calculating the cost of the culture medium components and cell culture conditions based on the culture medium components and culture conditions. In this disclosure, the flowcharts shown in Figures 15-22 can be used as non-limiting examples for calculating costs and expenses.

[0220] In one embodiment, the selection is (A) The step of selecting target cells; (B) A step of selecting the basic cellular functions (K) of the cells; (C) A step of determining a set of medium factors (N) that determine the environment containing the components of the medium; (D) Using experimental data of novel culture medium samples and / or the supernatant of existing culture medium samples using the components of the culture medium and the cells, a functional map is constructed that represents the intensity of activation or inhibition of each of the K basic cellular functions by each of the N culture medium factors. (E) Using the functional map, the step of optimizing the composition of the cell culture medium so as to activate or suppress one or more of the basic cellular functions; Includes.

[0221] In one embodiment, the computer calculates the cost of the culture medium components and the cell culture conditions based on the culture medium components and the culture conditions, taking into account a database that stores information on the costs and benefits of the culture medium and the equipment used for cultivation.

[0222] In another aspect, the present disclosure provides a method for managing culture medium components and / or cell production for cells provided by a user, comprising: 1) providing information on appropriate culture medium components for the cells provided by the user; 2) assigning identifiable labels to the cells and the culture medium components; and 3) referring to the labels, the cells and the culture medium components The present invention provides a method that includes a process for managing whether the conformity is maintained. In this disclosure, the flowcharts shown in Figures 1 to 7 can be referenced as non-limiting examples regarding management, etc.

[0223] In another aspect, the present invention provides a method for causing a computer to manage culture medium components and / or cell production for cells provided by a user, the method comprising: 1) providing information on appropriate culture medium components for the cells provided by the user; 2) assigning identifiable labels to the cells and the culture medium components; and 3) managing whether the cells and the culture medium components are compatible by referring to the labels. In this disclosure, the flowcharts shown in Figures 1 to 7 can be referenced as non-limiting examples regarding management, etc.

[0224] In this specification, "labeling" refers to the presence (e.g., substance, energy, electromagnetic wave, etc.) that distinguishes a target molecule or substance from others. Examples of such labeling methods include radioisotope (RI) methods, fluorescence methods, biotin methods, and chemiluminescence methods. In this specification, when multiple (two or more) markers or factors or means that capture them are labeled by fluorescence, the labeling is performed using fluorescent substances with different fluorescence emission maximum wavelengths. The difference in fluorescence emission maximum wavelengths is preferably 10 nm or more. When labeling ligands, any substance that does not affect their function can be used, but examples of fluorescent substances include Alexa™Fluor BODIPY, ATTO, quantum dots (QDot), and fluorescent proteins (GFP, YFP, mCherry, etc.). Alexa™Fluor is a water-soluble fluorescent dye obtained by modifying coumarin, rhodamine, fluorescein, cyanine, etc., and is a series that corresponds to a wide range of fluorescence wavelengths. Compared to other fluorescent dyes of the same wavelength, it is very stable, bright, and less pH sensitive. Examples of fluorescent dye combinations with a fluorescence maximum wavelength of 10 nm or more include the combination of Alexa™ 555 and Alexa™ 633, and the combination of Alexa™ 488 and Alexa™ 555. When labeling nucleic acids, any substance that can bind to the base portion can be used, but it is preferable to use cyanine dyes (e.g., Cy3 and Cy5 of the CyDye™ series), rhodamine 6G reagent, 2-acetylaminofluorene (AAF), AAIF (iodine derivative of AAF), etc. Examples of fluorescent substances with a fluorescence emission maximum wavelength difference of 10 nm or more include the combination of Cy5 and rhodamine 6G reagent, the combination of Cy3 and fluorescein, and the combination of rhodamine 6G reagent and fluorescein. In this disclosure, such labels can be used to modify the target object so that it can be detected by the detection means used. Such modifications are known in the art, and those skilled in the art can carry out such methods as appropriate depending on the label and the target object.

[0225] In one embodiment, the basic cellular function is derived from a biological network of the target biological structure, the biological network is divided into K functional subnetworks, the subnetworks being obtained manually and / or automatically, the subnetworks being obtained manually and / or automatically.

[0226] In one embodiment, the basic cellular functions are obtained from genome-scale reconstruction of the biological network of the target biological structure, and a working set of K basic cellular functions is pre-reduced using transcriptome data and / or proteome data and / or endo-metabolome data and / or thermodynamic data, when such data is available.

[0227] In one embodiment, the functional map is performed in a shaking flask, T-flask, reactor, microplate, microbioreactor, or phenotypic microarray. This is determined by continuous culture experiments and / or parallel culture experiments.

[0228] In one embodiment, the functional map is determined by an exometabolome assay that includes analysis of the supernatant of a novel culture medium sample or a used culture medium sample, using a chromatography method such as liquid chromatography (LC) or gas chromatography (GC), an NMR method such as 1H-NMR or 13C-NMR, mass spectrometry (MS), or a chromatography method combined with mass spectrometry such as GC-MS or LC-MS, or by a method combining the aforementioned measurement methods.

[0229] In one embodiment, the reduced set of activated basic cellular functions is identified by linear or nonlinear regression analysis, the variance or covariance of the exometabolome data or induced exometabolome data is maximized, the correlation between the exometabolome data or induced exometabolome data and the values ​​of the culture medium factors is maximized, and the basic cellular functions are ranked according to their correlation with or sensitivity to the values ​​of the culture medium factors.

[0230] In one embodiment, the functional map is determined by a high-throughput automated system, where a culture device, an analytical exometabolome device, and a computer algorithm are connected to a physical device to obtain a high-throughput functional map.

[0231] In one embodiment, the target basic cellular function i associated with product quantity and / or product quality is relative to the weight Δ(λ i Increase ) by 60% to 100%.

[0232] In one embodiment, the culture medium formulation of the present disclosure is characterized by enhancing heterologous protein expression function by 60% to 100%, and is a chemically defined culture medium formulation obtained by the method of the present disclosure for the culture of animal cells, plant cells, Gram-positive bacteria, Gram-negative bacteria, or yeast, comprising: a) an aqueous solution of trace components selected from the group consisting of methanol, various vitamins, various metal ions, and various amino acids; and b) a mixture of aqueous solution a) and a complementary basic aqueous solution, or other complementary basic aqueous solution, composed of components selected from the group consisting of MHH medium, BMMH medium, BMMY medium, and BMGY medium (but not limited to these). In one embodiment, the trace components are components selected from the group consisting of PTEN inhibitors, p53 inhibitors, p38 inhibitors, Wnt signaling activators, and ROCK inhibitors, preferably including, in order, VO-OH Pic, Pifithrin-a, SB203580, Lici, Y-27632, in which case the basic aqueous solution may be a basic aqueous solution containing DMEM / F-12. Alternatively, in another embodiment, the trace component may be IL-1 and / or TNF-α, and the basal aqueous solution may be a basal aqueous solution containing αMEM.

[0233] In one embodiment, the method of the present disclosure is used to increase the quantity and / or quality of tissues, and / or cells, and / or viruses, and / or cellular components, and / or protein-related substances, and / or carbohydrates, and / or nucleotide substances, and / or lipid substances, and / or primary metabolites, and / or secondary metabolites, or mixtures of products in biological manufacturing processes such as the production of biofuels, vaccines, drugs, biopolymers, or precursors thereof.

[0234] In one embodiment, the method disclosed herein is used to optimize the composition of a cell culture medium for a plant or animal kingdom cell line, or the composition of other eukaryotic unicellular or multicellular organisms such as yeast or fungi.

[0235] In one embodiment, the method disclosed herein optimizes the composition of the cell culture medium for a prokaryotic organism. It is used for this purpose.

[0236] In one embodiment, the method of the present disclosure is used to optimize the conditions and / or the composition of the culture medium for cell culture.

[0237] In one embodiment, the method of the present disclosure is used to identify biomarkers specific to cell function or culture medium components.

[0238] In one embodiment, the method of the present disclosure is used to design a drug or optimize a drug mixture that is directed to alter cellular functions associated with a disease state.

[0239] In one aspect, the Disclosure provides a program that includes code causing a computer to perform the identification of a biomarker using the method of the Disclosure.

[0240] In one embodiment, this disclosure provides a culture medium design system for carrying out the method of the present invention.

[0241] <database> In another aspect, the present disclosure provides a method for causing a computer to create a database of appropriate culture medium components and / or cell culture conditions for cells provided by a user, the method comprising: 1) inputting information of the cells provided by the user into the computer; 2) causing the computer to calculate culture medium components and culture conditions related to the cells, and optimal conditions, taking into account the database as necessary; 3) causing the computer to calculate optimal culture medium components and cell culture conditions for the cells based on the culture medium components and culture conditions; 4) recording the results of the calculation in the database; and 5) repeating steps 1) to 4). In this disclosure, the flowcharts shown in Figures 23-25 ​​can be used as non-limiting examples for database creation and other related processes.

[0242] In one embodiment, the analysis is based on the results of experiments using novel culture medium samples and / or existing culture medium samples.

[0243] In one embodiment, the method includes determining whether the target is a mesenchymal stem cell.

[0244] (Specific example) (1) Culture medium components In one embodiment, the following procedure is performed with the culture medium components of the present disclosure. Select a basic culture medium (e.g., DMEM, RPMI-1640, etc.). Add the necessary components (e.g., amino acids, vitamins, inorganic salts, growth factors, hormones) to the basic culture medium. To determine the optimal concentration of each component, experiments are conducted at multiple concentration ranges. The culture medium is mixed uniformly, filtered using a sterile filter, and kept sterile. (2)Cell culture conditions After the culture medium components are added, the cell culture conditions are implemented according to the following procedure. Select a cell line (e.g., stem cells, cancer cells, progenitor cells, etc.). Seed the cells using the prepared culture medium and transfer them to a suitable culture vessel. To promote cell proliferation and differentiation, adjust culture conditions such as temperature, CO2 concentration, and humidity. The culture medium is changed regularly, and the condition of the cells is observed. (3) Optimization process To optimize the culture medium and culture conditions, perform the following steps: Test multiple culture medium components and culture conditions to evaluate cell proliferation rate, differentiation potential, and quality. Statistically analyze the results of each experiment to determine the optimal culture medium components and conditions. Use the optimized conditions to establish reproducible, high-quality cell culture conditions.

[0245] The specific explanation is as follows: In one embodiment, the disclosure relates to a method for optimizing culture medium components and cell culture conditions, specifically providing a procedure for adjusting the components of a culture medium and optimizing culture conditions. According to the disclosure, it is possible to improve the proliferation rate, differentiation potential, and quality of cells.

[0246] (method, etc.) (1) Culture medium components The following procedure is performed for the culture medium components. Select a basic culture medium (e.g., DMEM, RPMI-1640, etc.). Add the necessary components (e.g., amino acids, vitamins, inorganic salts, growth factors, hormones) to the basic culture medium. To determine the optimal concentration of each component, experiments are conducted at multiple concentration ranges. The culture medium is mixed uniformly, filtered using a sterile filter, and kept sterile. (2)Cell culture conditions After the culture medium components are added, the cell culture conditions are implemented according to the following procedure. Select a cell line (e.g., stem cells, cancer cells, progenitor cells, etc.). Seed the cells using the prepared culture medium and transfer them to a suitable culture vessel. To promote cell proliferation and differentiation, adjust culture conditions such as temperature, CO2 concentration, and humidity. The culture medium is changed regularly, and the condition of the cells is observed. (3) Optimization process To optimize the culture medium and culture conditions, perform the following steps. Multiple culture medium components and culture conditions are tested to evaluate cell proliferation rate, differentiation potential, and quality. The results of each experiment will be statistically analyzed to determine the optimal culture medium components and conditions. Optimized conditions are used to establish reproducible, high-quality cell culture conditions.

[0247] (Production optimization) The development of mammalian cell culture conditions and processes for the large-scale industrial production of therapeutic proteins (e.g., monoclonal antibodies) began in the latter half of the 20th century. Efficient bioprocesses for biopharmaceutical production require optimal technical bioprocesses characterized primarily by (i) highly productive, stable, and regulatory-approved (typically mammalian) cell lines, (ii) optimal cell culture media to support cell proliferation and production (e.g., at various scales and such as batch processes, fed-batch processes, and perfusion processes) in various (typically mammalian) host cells and various culture systems and process schemes, and (iii) optimal oxygen supply through appropriate stirrer placement and gas supply, automated control of all relevant process parameters to ensure consistent product quality, or process design that can scale up from small-scale process development (mL to L scale) to large-scale production (over 2,000 L) without compromising performance and product quality.

[0248] This disclosure shows that, in this context, cell culture media play a crucial role and that the complex nutritional needs of mammalian cells cultured in suspensions within a technological system, as opposed to those of natural origin, are met. Certain conditions must be met. For example, the cell lines most widely used for biopharmaceutical production were originally derived from Chinese hamster ovary (CHO) cells.

[0249] In the past, serum was used as a culture medium additive to provide nutrients or carrier proteins not typically present in cell culture media, such as cholesterol and transferrin, or factors for cell-to-substrate adhesion (e.g., fibronectin), other hormones, and growth factors. However, it was also used to protect certain essential nutrients and bind toxic components in the culture medium. However, in cell culture media used for the production of therapeutic drugs, serum can potentially introduce animal viruses, and because the source of raw materials is unclear, it can introduce other undesirable contaminants (e.g., antibiotics or proteases) into the cell culture conditions and processes. These compounds have been successfully replaced, and serum-free cell culture media have become industry practice for biopharmaceutical process development and recombinant production.

[0250] Another group of culture medium components commonly used in the production of biopharmaceuticals are hydrolysates, which are either of animal or plant origin. Due to safety risks, hydrolysates of animal origin are removed from the process whenever possible. Hydrolysates typically contain mixtures of amino acids, small peptides, inorganic ions, trace elements, carbohydrates, and vitamins, and are widely used to enrich culture media with various (essential) nutrients to improve overall growth and productivity. Another drawback, aside from safety concerns, is that the chemical composition of hydrolysates is not clear, and therefore the exact composition can vary from lot to lot (lot variability), which can negatively impact process reproducibility. Because hydrolysates contain many compounds and have complex (not fully known) compositions, they cannot be easily replaced without affecting the performance of cell culture conditions (e.g., product yield). Selecting such unclear raw materials and substituting them with components whose composition is chemically defined, while simultaneously maintaining consistent product quality and high product potency, remains an unresolved challenge in bioproce...

Claims

1. A method for controlling the quality of cells, comprising: culturing cells with a culture medium containing a component; obtaining a coefficient related to the component; and determining the correlation between the culture medium containing the component and the cells based on the coefficient.

2. The method according to claim 1, wherein the determination includes determining whether the target cell is a target cell.

3. The method according to claim 1, wherein the determination includes determining whether the culture medium or the component is the target culture medium or the target component.

4. The method according to any one of claims 1 to 3, wherein the determination includes referring to a database storing information regarding the correlation between the component and the coefficient.

5. Adding a culture medium containing the component to the cells and culturing them, obtaining a coefficient related to the component, and determining whether the culture medium containing the component can be correlated based on the coefficient, A method for producing a culture medium with controllable quality, which includes, if the aforementioned correlation is found, manufacturing a culture medium containing that component as a controllable culture medium.

6. The method according to claim 5, wherein the determination includes referring to a database that stores information regarding the correlation between the component and the coefficient.

7. A method for quality control of cells, A method comprising: confirming whether the culture medium used for culturing the cells contains components necessary for quality control of the cells; and, if the necessary components are not present, adding the necessary components to the culture medium.

8. The method according to claim 7, wherein the verification includes referring to a database that stores information regarding the correlation between the component and coefficients related to the component.

9. A method for causing a computer to select appropriate culture medium components and / or cell culture conditions for cells provided by a user, the method being: The process of inputting cell information provided by the user into the computer, The process involves having the computer, if necessary, consult a database to derive culture medium components and culture conditions related to the cells, as well as optimal conditions if necessary. The process involves having the computer calculate the optimal culture medium components and cell culture conditions for the cells based on the culture medium components and the culture conditions. If necessary, the process involves displaying the results of the calculation on a display. A method of including.

10. A method for causing a computer to select appropriate culture medium components and / or cell culture conditions for cells provided by a user, the method being: The process of inputting cell information provided by the user into the computer, The process of inputting data on the culture results under specific culture conditions for the cells into the computer, The process of inputting desired output conditions for the cells to the computer, The process involves having the computer, if necessary, consult a database to derive culture medium components and culture conditions related to the cells, as well as optimal conditions if necessary. The process of having the computer calculate the optimal culture medium components and cell culture conditions for the cells based on the culture medium components and the culture conditions, If necessary, the process includes displaying the results of the calculation on a display, If necessary, the process includes documenting the results of the calculations, If necessary, the process of outputting the documented document and A method of including.

11. The method according to claim 10, wherein the documentation is documented in accordance with the standard.

12. The method according to claim 11, wherein the standard includes a standard for a Manufacturing Order Record (MBR) or a Standard Operating Procedure (SOP).

13. The method according to claim 10, wherein the optimal culture medium components and cell culture conditions for the cells include components and conditions suitable for both small-scale and commercial-scale, or for either one, or include culture conditions progressively from small-scale to commercial-scale.

14. The method according to claim 10, further comprising the steps of extracting predicted risks with respect to the optimal culture medium components and cell culture conditions for the cells, formulating countermeasures against the risks, and providing information on the risks and countermeasures.

15. The method according to claim 10, wherein the database includes at least one selected from the group consisting of cell information, specifications of an automated culture device, the number of people required depending on the culture conditions, the layout of the manufacturing plant, information on the wiring during culture, and information on the reagents held.

16. The method according to claim 10, wherein the optimal culture medium components and cell culture conditions for the cells include components and culture conditions using an automated culture device.

17. The method according to claim 10, wherein the cell culture conditions include information on manufacturing costs.

18. The method according to claim 14, wherein the aforementioned risks include at least one selected from the group consisting of risks of changes in the state and / or quantity of cultured cells during scale-up, risks of non-compliance with GMP, risks of malfunctions in reagent preparation during culture, risks in preparing culture medium components including powder preparation and on-demand preparation, risks of variations in culture medium quality and lot, and regional risks.

19. The countermeasure is the method according to claim 14, which is output based on the information in the database.

20. The method according to claim 14, wherein the countermeasures include at least one selected from the group consisting of proposing compatible culture medium factors, proposing methods for managing and supplying culture medium components, automatically generating risk reduction documents, proposing routine responses that allow for biogroups, and alternative culture methods.

21. The method according to claim 10, wherein the document includes a proposed initial culture method up to commercial production.

22. The method according to claim 10, wherein the document includes providing information on cell production volume, production efficiency, cost of culture medium components, and composition of culture medium components, as well as providing at least one selected from the group consisting of suggestions for changes to culture medium components, suggestions for culture conditions, suggestions for alternative methods, MBR proposals, SOP proposals, visualization of the manufacturing site, and proposed roadmaps.

23. The above selection is (A) The step of selecting target cells; (B) The step of selecting K basic cellular functions of the cells; (C) A step of determining a set of N culture medium factors that determine the environment containing the components of the culture medium; (D) Using information stored in a database related to the cells, construct a functional map representing the intensity of activation or suppression of each of the K basic cellular functions by each of the N culture medium factors; (E) Using the functional map, the step of optimizing the composition of the cell culture medium so as to activate or suppress one or more of the basic cellular functions; The method according to claims 9 to 22, wherein K and N are independently arbitrary integers.

24. The method according to claims 9 to 23, wherein the database is a library containing cytokines, growth factors, proteins, small molecules, and physiologically active substances useful for cell culture conditions, and can be used to compose the culture medium.

25. The method according to any one of claims 9 to 24, wherein the database is a library containing information on cell culture conditions that can be used to compose the culture medium.

26. A method for performing calculations to select appropriate culture medium components and / or cell culture conditions for cells provided by a user, wherein the method is: A step of analyzing cells provided by the user using candidate culture medium components and / or cell culture conditions, The process involves deriving the culture medium components and culture conditions (optimal requirements as needed) related to the cells from the analysis results, A step of calculating appropriate culture medium components and cell culture conditions for the cells based on the culture medium components and culture conditions. A method of including.

27. The above selection is (A) The step of selecting target cells; (B) The step of selecting K basic cellular functions of the cells; (C) A step of determining a set of N culture medium factors that determine the environment containing the components of the culture medium; (D) Using experimental data of a novel culture medium sample and / or the supernatant of an existing culture medium sample using the components of the culture medium and the cells, a functional map is constructed that represents the intensity of activation or inhibition of each of the K basic cellular functions by each of the N culture medium factors. (E) Using the functional map, the step of optimizing the composition of the cell culture medium so as to activate or suppress one or more of the basic cellular functions; The method according to claim 26, including the method described in claim 26.

28. The method according to claim 26 or 27, wherein the candidate culture component is selected from the library of culture components.

29. The method according to any one of claims 26 to 28, wherein the candidate cell culture conditions are selected from the library of cell culture conditions.

30. The method according to any one of claims 26 to 29, wherein the analysis is performed based on the results of an actual experiment using the culture medium components.

31. The method according to any one of claims 26 to 30, wherein the calculation is performed using a library parameterized for the performance and / or cost of the culture component to cells.

32. The aforementioned analysis includes classification and / or The above derivation includes a reference to a database containing the learning results. The method according to any one of claims 26 to 31.

33. A method for providing appropriate culture medium components and / or cell culture conditions for cells provided by a user using a computer, wherein the method is: The process of inputting cell information provided by the user into the computer, The process involves having the computer refer to a database containing information about the cells and derive appropriate culture medium components and culture conditions for the cells. If necessary, the process involves displaying the results of the calculation on a display. A method of including.

34. This includes recording the derived appropriate culture medium components and culture conditions in the database. Furthermore, when a query for appropriate culture medium components and / or cell culture conditions for the aforementioned cells is received, the appropriate culture medium components and / or cell culture conditions recorded in the database are retrieved. If necessary, the process involves displaying the result of the aforementioned call on the display. The method according to claim 33, including the method described in claim 33.

35. A program that codes a method for causing a computer to perform the selection of appropriate culture medium components and / or cell culture conditions for cells provided by a user, wherein the method is: The process of inputting cell information provided by the user into the computer, The process involves having the computer, as necessary, consult a database to derive culture medium components and culture conditions related to the cells, and, if necessary, optimal conditions. The process involves having the computer calculate the optimal culture medium components and cell culture conditions for the cells based on the culture medium components and the culture conditions. If necessary, the process involves displaying the results of the calculation on a display. A program that encompasses all of these.

36. A program that codes a method for causing a computer to perform the selection of appropriate culture medium components and / or cell culture conditions for cells provided by a user, wherein the method is: The process of inputting cell information provided by the user into the computer, The process of inputting data on the culture results under specific culture conditions for the cells into the computer, The process of inputting desired output conditions for the cells to the computer, The process involves having the computer, if necessary, consult a database to derive culture medium components and culture conditions related to the cells, as well as optimal conditions if necessary. The process of having the computer calculate the optimal culture medium components and cell culture conditions for the cells based on the culture medium components and the culture conditions, If necessary, the process includes displaying the results of the calculation on a display, If necessary, the process includes documenting the results of the calculations, If necessary, the process of outputting the documented document and A program that includes this.

37. The program according to claim 36, wherein the documentation is documented in accordance with the standard.

38. The program according to claim 37, wherein the standard includes a standard for a Manufacturing Order Record (MBR) or a Standard Operating Procedure (SOP).

39. The program according to claim 36, wherein the optimal culture medium components and cell culture conditions for the cells include components and conditions suitable for both small-scale and commercial-scale, or for either one, or include culture conditions in a stepwise manner from small-scale to commercial-scale.

40. The program according to claim 36, further comprising the steps of extracting predicted risks with respect to the optimal culture medium components and cell culture conditions for the cells, formulating countermeasures for the risks, and providing information on the risks and countermeasures.

41. The program according to claim 36, wherein the database includes at least one selected from the group consisting of cell information, specifications of an automated culture device, the number of people required depending on the culture conditions, the layout of the manufacturing plant, information on the wiring during culture, and information on the reagents held.

42. The program according to claim 36, wherein the optimal culture medium components and cell culture conditions for the cells include components and culture conditions using an automated culture device.

43. The program according to claim 36, wherein the cell culture conditions include information on manufacturing costs.

44. The program according to claim 40, wherein the risks include at least one selected from the group consisting of risks of changes in the state and / or quantity of cultured cells during scale-up, risks of non-compliance with GMP, risks of malfunctions in reagent preparation during culture, risks in preparing culture medium components including powder preparation and on-demand preparation, risks of variations in culture medium quality and lot, and regional risks.

45. The countermeasure is the program according to claim 40, which is output based on the information in the database.

46. The program according to claim 40, wherein the countermeasures include at least one selected from the group consisting of suggesting compatible culture medium factors, suggesting methods for managing and supplying culture medium components, automatically generating risk reduction documents, suggesting routine responses that allow for biogroups, and alternative culture methods.

47. The program according to claim 36, wherein the document includes a proposed initial culture method up to commercial production.

48. The program according to claim 36, wherein the document includes providing information on cell production volume, production efficiency, cost of culture medium components, and composition of culture medium components, as well as providing suggestions for changes to culture medium components, suggestions for culture conditions, suggestions for alternative methods, MBR proposals, SOP proposals, visualization of the manufacturing site, and proposed roadmaps.

49. A recording medium storing a program that codes a method for causing a computer to perform the selection of appropriate culture medium components and / or cell culture conditions for cells provided by a user, wherein the method is The process of inputting cell information provided by the user into the computer, The process involves having the computer, if necessary, consult a database to derive culture medium components and culture conditions related to the cells, and, if necessary, optimal conditions. The process involves having the computer calculate the optimal culture medium components and cell culture conditions for the cells based on the culture medium components and the culture conditions. If necessary, the process involves displaying the results of the calculation on a display. A recording medium that includes this.

50. A recording medium storing a program that codes a method for causing a computer to perform the selection of appropriate culture medium components and / or cell culture conditions for cells provided by a user, wherein the method is The process of inputting cell information provided by the user into the computer, The process of inputting data on the culture results under specific culture conditions for the cells into the computer, The process of inputting desired output conditions for the cells to the computer, The process involves having the computer, if necessary, consult a database to derive culture medium components and culture conditions related to the cells, as well as optimal conditions if necessary. The process of having the computer calculate the optimal culture medium components and cell culture conditions for the cells based on the culture medium components and the culture conditions, If necessary, the process includes displaying the results of the calculation on a display, If necessary, the process includes documenting the results of the calculations, If necessary, the process of outputting the documented document and A recording medium that includes this.

51. The recording medium according to claim 50, wherein the aforementioned documentation is documented in accordance with the standard.

52. The recording medium according to claim 51, wherein the standard includes a standard for a Manufacturing Order Record (MBR) or a Standard Operating Procedure (SOP).

53. The recording medium according to claim 50, wherein the optimal culture medium components and cell culture conditions for the cells include components and conditions suitable for both small-scale and commercial-scale, or for either one, or include culture conditions in a stepwise manner from small-scale to commercial-scale.

54. The recording medium according to claim 50, further comprising the steps of extracting predicted risks with respect to the optimal culture medium components and cell culture conditions for the cells, formulating countermeasures against the risks, and providing information on the risks and countermeasures.

55. The recording medium according to claim 50, wherein the database includes at least one selected from the group consisting of cell information, specifications of an automated culture device, the number of people required depending on the culture conditions, the layout of the manufacturing plant, information on the wiring during culture, and information on the reagents held.

56. The recording medium according to claim 50, wherein the optimal culture medium components and cell culture conditions for the cells include components and culture conditions using an automated culture device.

57. The recording medium according to claim 50, wherein the cell culture conditions include information on manufacturing costs.

58. The recording medium according to claim 54, wherein the aforementioned risks include at least one selected from the group consisting of risks of changes in the state and / or quantity of cultured cells during scale-up, risks of non-compliance with GMP, risks of malfunctions in reagent preparation during culture, risks in preparing culture medium components including powder preparation and on-demand preparation, risks of variations in culture medium quality and lot, and regional risks.

59. The aforementioned countermeasure is the recording medium according to claim 54, which is output based on the information in the database.

60. The recording medium according to claim 54, wherein the countermeasures include at least one selected from the group consisting of proposing compatible culture medium factors, proposing methods for managing and supplying culture medium components, automatically generating risk reduction documents, proposing routine responses that allow for biogroups, and alternative culture methods.

61. The recording medium according to claim 50, wherein the document includes a proposed initial culture method up to commercial production.

62. The recording medium according to claim 50, wherein the document includes at least one selected from the group consisting of providing information on cell production volume, production efficiency, cost of culture medium components, and composition of culture medium components, as well as proposals for changes to culture medium components, proposals for culture conditions, proposals for alternative methods, proposals for MBR, proposals for SOP, provision of visualized production site information, and proposals for roadmap provision.

63. A system that performs the selection of appropriate culture medium components and / or cell culture conditions for cells provided by a user, wherein the system A cell information input unit that inputs cell information provided by the user, A culture medium component / culture condition deriving unit that, when necessary, consults a database to derive culture medium components and culture conditions related to the cells, and, if necessary, optimal conditions, An optimal calculation unit calculates the optimal culture medium components and cell culture conditions for the cells based on the culture medium components and the culture conditions. If necessary, a display unit that displays the results of the calculation on a display. A system that encompasses all of these.

64. A system that performs the selection of appropriate culture medium components and / or cell culture conditions for cells provided by a user, wherein the system A cell information input unit that inputs cell information provided by the user, A culture result input unit for inputting culture result data under specific culture conditions for the aforementioned cells, An output condition input unit for inputting desired output conditions for the aforementioned cells, A culture medium component / culture condition deriving unit that, if necessary, consults a database to derive culture medium components and culture conditions related to the cells, and, if necessary, optimal conditions, An optimal calculation unit that calculates the optimal culture medium components and cell culture conditions for the cells based on the culture medium components and the culture conditions, A display unit that, if necessary, displays the results of the calculation on a display, A documentation unit that documents the results of the calculations as needed, If necessary, an output unit to output the documented document and A system that encompasses all of these.

65. The system according to claim 64, wherein the documentation is documented in accordance with the standard.

66. The system according to claim 65, wherein the standard includes a standard for a Manufacturing Order Record (MBR) or a Standard Operating Procedure (SOP).

67. The system according to claim 64, wherein the optimal culture medium components and cell culture conditions for the cells include components and conditions suitable for both small-scale and commercial-scale, or for either one, or include culture conditions in a stepwise manner from small-scale to commercial-scale.

68. The system according to claim 64, further comprising the steps of extracting predicted risks with respect to the optimal culture medium components and cell culture conditions for the cells, formulating countermeasures against the risks, and providing information on the risks and countermeasures.

69. The system according to claim 64, wherein the database includes at least one selected from the group consisting of cell information, specifications of an automated culture device, the number of people required depending on the culture conditions, the layout of the manufacturing plant, information on the wiring during culture, and information on the reagents held.

70. The system according to claim 64, wherein the optimal culture medium components and cell culture conditions for the cells include components and culture conditions using an automated culture device.

71. The system according to claim 64, wherein the cell culture conditions include information on manufacturing costs.

72. The system according to claim 68, wherein the aforementioned risks include at least one selected from the group consisting of risks of changes in the state and / or quantity of cultured cells during scale-up, risks of non-compliance with GMP, risks of malfunctions in reagent preparation during culture, risks in preparing culture medium components including powder preparation and on-demand preparation, risks of variations in culture medium quality and lot, and regional risks.

73. The aforementioned countermeasure is output based on information in the database, according to the system described in claim 68.

74. The system according to claim 68, wherein the countermeasures include at least one selected from the group consisting of suggesting compatible culture medium factors, suggesting methods for managing and supplying culture medium components, automatically generating risk reduction documents, suggesting routine responses that tolerate biogroups, and alternative culture methods.

75. The system according to claim 64, wherein the aforementioned document includes a proposed initial culture method up to commercial production.

76. The system according to claim 64, wherein the document includes at least one selected from the group consisting of providing information on cell production volume, production efficiency, cost of culture medium components, and composition of culture medium components, as well as proposals for changes to culture medium components, proposals for culture conditions, proposals for alternative methods, proposals for MBR, proposals for SOP, provision of visualized production site information, and proposals for roadmap provision.

77. A method for causing a computer to perform a calculation of the cost of culture medium components and / or cell culture conditions for cells provided by a user, the method being: The process of inputting cell information provided by the user into the computer, The process involves having the computer, if necessary, consult a database and calculate the culture medium components and culture conditions related to the cells, as well as the optimal conditions if necessary. A step of causing the computer to calculate the cost of the culture medium components and the cell culture conditions based on the culture medium components and the culture conditions. If necessary, the process of displaying the aforementioned costs on a display and A method of including.

78. The above selection is (A) The step of selecting target cells; (B) The step of selecting K basic cellular functions of the cells; (C) A step of determining a set of N culture medium factors that determine the environment containing the components of the culture medium; (D) Using information stored in a database related to the cells, construct a functional map representing the intensity of activation or suppression of each of the K basic cellular functions by each of the N culture medium factors; (E) Using the functional map, the step of optimizing the composition of the cell culture medium so as to activate or suppress one or more of the basic cellular functions; The method according to claim 77, including the method described in claim 77.

79. The method according to claim 77 or 78, further comprising the features of any one or more of claims 9 to 76.

80. A method for performing a calculation of the cost of culture medium components and / or cell culture conditions for cells provided by a user, the method being: The process of analyzing cells provided by the user, The process involves deriving the culture medium components and culture conditions related to the cells from the aforementioned analysis results, A step of calculating the cost of the culture medium components and the cell culture conditions based on the culture medium components and the culture conditions. A method of including.

81. The above selection is (A) The step of selecting target cells; (B) The step of selecting K basic cellular functions of the cells; (C) A step of determining a set of N culture medium factors that determine the environment containing the components of the culture medium; (D) Using experimental data of a novel culture medium sample and / or the supernatant of an existing culture medium sample using the components of the culture medium and the cells, a functional map is constructed that represents the intensity of activation or inhibition of each of the K basic cellular functions by each of the N culture medium factors. (E) Using the functional map, the step of optimizing the composition of the cell culture medium so as to activate or suppress one or more of the basic cellular functions; The method according to claim 80, including the method described in claim 80.

82. The method according to any one of claims 77 to 81, wherein the computer calculates the cost of the culture medium components and the cell culture conditions based on the culture medium components and the culture conditions, taking into consideration a database storing information on the costs and benefits of the culture medium and equipment used for culture.

83. For cells provided by users, the person responsible for managing the culture medium components and / or cell production. It is a law, 1) A step of providing information regarding appropriate culture medium components for the cells provided by the user, 2) A step of assigning identifiable labels to the cells and the culture medium components, 3) A step of checking whether the cells and the culture medium components are compatible by referring to the label. A method of including.

84. The method according to claim 83, wherein the information relating to the culture medium components further comprises the features described in any one or more of claims 9 to 82.

85. A method for causing a computer to perform a calculation of the cost of culture medium components and / or cell culture conditions for cells provided by a user, the method being: The process of inputting cell information provided by the user into the computer, The process involves having the computer, if necessary, consult a database and calculate the culture medium components and culture conditions related to the cells, as well as the optimal conditions if necessary. A step of causing the computer to calculate the cost of the culture medium components and the cell culture conditions based on the culture medium components and the culture conditions. If necessary, the process of displaying the aforementioned costs on a display and A method of including.

86. The above selection is (A) The step of selecting target cells; (B) The step of selecting K basic cellular functions of the cells; (C) A step of determining a set of N culture medium factors that determine the environment containing the components of the culture medium; (D) Using information stored in a database related to the cells, construct a functional map representing the intensity of activation or suppression of each of the K basic cellular functions by each of the N culture medium factors; (E) Using the functional map, the step of optimizing the composition of the cell culture medium so as to activate or suppress one or more of the basic cellular functions; The method according to claim 85, including the method described in claim 85.

87. The method according to claim 85 or 86, further comprising the features of any one or more of claims 9 to 76.

88. A method for performing a calculation of the cost of culture medium components and / or cell culture conditions for cells provided by a user, the method being: The process of analyzing cells provided by the user, The process involves deriving the culture medium components and culture conditions related to the cells from the aforementioned analysis results, A step of calculating the cost of the culture medium components and the cell culture conditions based on the culture medium components and the culture conditions. A method of including.

89. The above selection is (A) The step of selecting target cells; (B) The step of selecting K basic cellular functions of the cells; (C) A step of determining a set of N culture medium factors that determine the environment containing the components of the culture medium; (D) Using experimental data of a novel culture medium sample and / or the supernatant of an existing culture medium sample using the components of the culture medium and the cells, a functional map is constructed that represents the intensity of activation or inhibition of each of the K basic cellular functions by each of the N culture medium factors. (E) Using the functional map, the step of optimizing the composition of the cell culture medium so as to activate or suppress one or more of the basic cellular functions; The method according to claim 88, including the method described in claim 88.

90. The method according to any one of claims 85 to 89, wherein the computer calculates the cost of the culture medium components and the cell culture conditions based on the culture medium components and the culture conditions, taking into consideration a database storing information on the costs and benefits of the culture medium and equipment used for culture.

91. A method for causing a computer to manage the culture medium components and / or cell production for cells provided by a user, 1) A step of providing information regarding appropriate culture medium components for the cells provided by the user, 2) A step of assigning identifiable labels to the cells and the culture medium components, 3) A step of checking whether the cells and the culture medium components are compatible by referring to the label. A method of including.

92. A method for causing a computer to create a database of appropriate culture medium components and / or cell culture conditions for cells provided by a user, the method being: 1) The process of inputting cell information provided by the user into the computer, 2) A step of having the computer calculate the culture medium components and culture conditions related to the cells, and the optimal conditions, taking into account the database as necessary, 3) A step of causing the computer to calculate the optimal culture medium components and cell culture conditions for the cells based on the culture medium components and the culture conditions. 4) A step of recording the results of the calculation in a database. 5) A process that repeats steps 1) to 4) and A method of including.

93. The method according to claim 92, wherein the analysis is based on the results of experiments using novel culture medium samples and / or existing culture medium samples.

94. The method according to claim 92 or 93, further comprising determining whether the subject is a mesenchymal stem cell.

95. The method according to any one of claims 92 to 94, further comprising the features of any one or more of claims 9 to 76.

96. A system for automating culture, 1) A cell donor unit that provides cells intended for culture, 2) A cell information providing unit that provides information about the cells as needed. 3) A culture medium information providing unit that provides information on the culture medium or culture medium components as needed. 4) A culture medium matching section for matching cells and culture medium as needed, 5) A culture medium supply unit that provides culture medium, 6) A cell culture section for culturing cells, 7) A cell culture condition adjustment unit for adjusting cell conditions, 8) Cell culture control unit for controlling and automating cell culture A system comprising the above, wherein the culture medium is optimized for the cells.

97. The system according to claim 96, wherein the selection is made by the method described in any one of claims 9 to 76.

98. A system for automating culture, 1) A cell donor unit that provides cells intended for culture, 2) A culture medium compatibility information storage unit that stores information regarding the compatibility of the cells and the culture medium, 5) A culture medium supply unit that provides culture medium, 6) A cell culture section for culturing cells, 7) A cell culture condition adjustment unit for adjusting cell conditions, 8) Cell culture control unit for controlling and automating cell culture A system comprising the above, wherein the culture medium is optimized for the cells.

99. The system according to any one of claims 96 to 98, further comprising a monitoring unit for monitoring the cells and / or components in the culture medium.

100. The system according to claim 99, wherein the monitoring unit includes a sensor that is in contact with or not in contact with the culture vessel or culture channel.

101. The system according to claim 100, wherein the sensor includes an optical sensor, an electrical sensor, a magnetic sensor, a chemical sensor, an acoustic sensor, a microfluidic sensor, or a biosensor.

102. Furthermore, the system according to claim 100, further comprising a sensor data storage unit for storing data obtained by the sensor, and at least one, preferably both, of an analysis and extraction unit for analyzing the data and extracting stable data and unstable data.

103. The system according to claim 102, wherein the analysis and extraction unit is configured to continuously monitor unstable data.

104. The system according to any one of claims 99 to 103, wherein the cell culture condition adjustment unit can add or change the components or the culture medium based on information obtained by the monitoring unit.

105. The system according to any one of claims 99 to 104, characterized in that the culture medium is selected from a plurality of suitable culture media and the optimal culture medium is selected or used by monitoring the state of the cells with the monitoring unit.

106. The system according to any one of claims 99 to 105, wherein the culture medium supply unit is configured to dispense multiple main components of a culture medium into multiple containers of the automated culture system.

107. The system according to any one of claims 99 to 106, wherein the monitoring unit is configured to monitor the culture state and to instantly calculate the optimal composition in the culture medium adaptation unit.

108. The system according to any one of claims 99 to 107, wherein the culture medium supply unit is configured to blend the main components of the culture medium to achieve an optimal composition and to continue the culture.

109. A method for performing automated cell culture using the system described in any one of claims 96 to 108.

110. A program for performing automated cell culture in the system according to any one of claims 96 to 108.

111. The system according to any one of claims 96 to 108, the method according to claim 109, and the program according to claim 110, wherein the cells are mesenchymal stem cells.

112. A system for delivering and transporting appropriate culture media to cells from a user, 1) Selection means for selecting appropriate culture medium components and culture and / or storage conditions for the cells, 2) A transport means including means for transporting the culture medium components, 3) Condition calculation means for calculating appropriate conditions for transporting the culture medium components, 4) A control unit that controls the delivery and transport of the culture medium and A system comprising at least one selected from the following.

113. A system for delivering and transporting a combination of cells and a suitable culture medium from a user, 1) Selection means for selecting appropriate culture medium components and culture and / or storage conditions for the cells, 2) A transport means including means for transporting the combination of the culture medium components and the cells, 3) Condition calculation means for calculating conditions appropriate for transporting the combination of culture medium components and cells, 4) A control unit that controls the delivery and transport of the combination of the culture medium components and the cells, A system comprising at least one selected from the following.

114. The system according to claim 112 or 113, wherein the selection is made by the method described in any one of claims 9 to 76.

115. A method for managing culture medium components and / or cell production for cells provided by a user, 1) A step of providing information regarding appropriate culture medium components for the cells provided by the user, 2) A step of assigning identifiable labels to the cells and the culture medium components, 3) A step of checking whether the cells and the culture medium components are compatible by referring to the label. A method of including.

116. The method of claim 115, wherein the management refers to information used in the method of any one of claims 9 to 76.

117. A method for causing a computer to manage the culture medium components and / or cell production for cells provided by a user, 0) A process of inputting information about cells provided by the user, 1) A step of providing information regarding appropriate culture medium components for the cells, 2) A step of assigning identifiable labels to the cells and the culture medium components, 3) A step of checking whether the cells and the culture medium components are compatible by referring to the label. A method of including.

118. A program that causes a computer to perform a method for managing culture medium components and / or cell production for cells provided by a user, wherein the method 0) A process of inputting information about cells provided by the user, 1) A step of providing information regarding appropriate culture medium components for the cells provided by the user, 2) A step of assigning identifiable labels to the cells and the culture medium components, 3) A step of checking whether the cells and the culture medium components are compatible by referring to the label. A program that includes this.

119. A method for providing the most appropriate healthcare and medical treatment to a subject, The process of providing medical information about the subject, A step of providing appropriate cells based on the aforementioned medical information, A step of providing optimal culture medium components or combinations based on the aforementioned cells and the aforementioned medical information. If necessary, the step of providing the cells cultured with the culture medium components or a combination thereof to the target. A method of including.

120. The method according to claim 119, wherein the provision of the optimal culture medium components or combination thereof is made by the method according to any one of claims 9 to 76.

121. A system that provides the most appropriate healthcare and medical treatment to the target, Means of providing medical information about the subject, A means for providing appropriate cells based on the aforementioned medical information, Means for providing optimal culture medium components or combinations based on the aforementioned cells and the aforementioned medical information. Means for providing the cells cultured with the culture medium components or combinations thereof to the target, if necessary. A system that encompasses all of these.

122. A program for implementing a method for providing a computer with the most appropriate healthcare and medical treatment for a subject, wherein the method The process of providing medical information about the subject, A step of providing appropriate cells based on the aforementioned medical information, A step of providing optimal culture medium components or combinations based on the aforementioned cells and the aforementioned medical information. If necessary, the step of providing the cells cultured with the culture medium components or a combination thereof to the target. A program that encompasses all of these.

123. The system of claim 64, the method according to claim 121, and the program according to claim 122, wherein the cells are microbial cells or animal or plant cells.

124. A method for generating process development methods, 1) A step of inputting information about cells or proteins related to said cells into a database, 2) A step of inputting information on the modification of the cells or the modification of the proteins into a database, 3) A step of outputting from the database information regarding scaffold materials that readily adhere to adherent cells present in the cell culture, and information regarding equipment materials that do not readily adhere to adherent cells. 4) A step of outputting information from the database regarding adherent cells, suspension cells, and / or materials in which components eluted from cells are less likely to condense, 5) A step of outputting a culture vessel suitable for cell culture from the database, 6) A step of outputting information from the database regarding sensors that can sense cells, culture medium components, and culture vessels, 7) A step of optimizing the cell culture conditions and the culture medium components, and obtaining data for optimizing the process in accordance with the modification of the cells or the modification of the protein, 8) An analysis process to separate the data obtained in 7) into stable data and unstable data, 9) A method comprising the step of outputting a suggestion to monitor the unstable data more frequently than the stable data.

125. A system for developing processes, 1) A cell protein input unit that inputs information about cells or proteins related to said cells into a database, 2) A modification input unit that inputs information on the modification of the cells or the modification of the proteins into a database, 3) An adhesion-related output unit that outputs from the database information regarding scaffold materials that readily adhere to adherent cells present in the cell culture and information regarding equipment materials that do not readily adhere to adherent cells, 4) A material-related output unit that outputs information from the database regarding adherent cells, suspension cells, and / or materials from which components eluted from cells are less likely to condense, 5) A container-related output unit that outputs a culture vessel suitable for cell culture from the database, 6) A sensor-related output unit that outputs information from the database regarding sensors that can sense cells, culture medium components, and culture vessels, 7) A data acquisition unit that acquires data for optimizing the cell culture conditions and the culture medium components, and for optimizing the process in accordance with the modification of the cells or the modification of the proteins, 8) An analysis unit that sorts the data obtained in 7) into stable data and unstable data, 9) A system including a suggestion output unit that outputs a suggestion to monitor the unstable data more frequently than the stable data.

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