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WO2025133344A1 - Procédés de culture de cellules souches pluripotentes par perfusion - Google Patents

Procédés de culture de cellules souches pluripotentes par perfusion Download PDF

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Publication number
WO2025133344A1
WO2025133344A1 PCT/EP2024/088240 EP2024088240W WO2025133344A1 WO 2025133344 A1 WO2025133344 A1 WO 2025133344A1 EP 2024088240 W EP2024088240 W EP 2024088240W WO 2025133344 A1 WO2025133344 A1 WO 2025133344A1
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cell
cells
cultured
sec
human
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Antony James DAVIES
Caspar Marinus QUAKKELAAR
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Meatable BV
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Meatable BV
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    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N5/00—Undifferentiated human, animal or plant cells, e.g. cell lines; Tissues; Cultivation or maintenance thereof; Culture media therefor
    • C12N5/06—Animal cells or tissues; Human cells or tissues
    • C12N5/0602—Vertebrate cells
    • C12N5/0603—Embryonic cells ; Embryoid bodies
    • C12N5/0606—Pluripotent embryonic cells, e.g. embryonic stem cells [ES]
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23L—FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES, NOT OTHERWISE PROVIDED FOR; PREPARATION OR TREATMENT THEREOF
    • A23L13/00—Meat products; Meat meal; Preparation or treatment thereof
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2527/00—Culture process characterised by the use of mechanical forces, e.g. strain, vibration
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N5/00—Undifferentiated human, animal or plant cells, e.g. cell lines; Tissues; Cultivation or maintenance thereof; Culture media therefor
    • C12N5/06—Animal cells or tissues; Human cells or tissues
    • C12N5/0602—Vertebrate cells
    • C12N5/0652—Cells of skeletal and connective tissues; Mesenchyme
    • C12N5/0653—Adipocytes; Adipose tissue
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N5/00—Undifferentiated human, animal or plant cells, e.g. cell lines; Tissues; Cultivation or maintenance thereof; Culture media therefor
    • C12N5/06—Animal cells or tissues; Human cells or tissues
    • C12N5/0602—Vertebrate cells
    • C12N5/0652—Cells of skeletal and connective tissues; Mesenchyme
    • C12N5/0658—Skeletal muscle cells, e.g. myocytes, myotubes, myoblasts
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N5/00—Undifferentiated human, animal or plant cells, e.g. cell lines; Tissues; Cultivation or maintenance thereof; Culture media therefor
    • C12N5/06—Animal cells or tissues; Human cells or tissues
    • C12N5/0602—Vertebrate cells
    • C12N5/0696—Artificially induced pluripotent stem cells, e.g. iPS

Definitions

  • the present invention relates to a method of culturing pluripotent stem cells by perfusion.
  • the invention relates to a method for reducing the size of cell aggregates, for example pluripotent stem cell aggregates, during cell culture the method comprising culturing the pluripotent cells by perfusion, such as for example continuous perfusion, in a cell culture medium and applying shear to the medium.
  • the shear rate is applied in the range from about 90 sec 1 to 2829 sec 1 , preferably about 1100 sec-1 to 2200 sec-1 , or more preferably about 1500 sec 1 to 2000 sec 1 . In one embodiment, during culture shear stress is applied in the range from about T10 -2 Pascal to about 1 -10 ° Pascal.
  • the cell is selected from the group consisting of embryonic stem cells, induced pluripotent stem cells, embryonic cell lines, somatic cell lines and immortalized cell lines.
  • the cells produced by the method are suitable for human and nonhuman dietary consumption.
  • the method is forthe proliferation ofthe pluripotent cell.
  • the method as described herein further 10 further comprises the step of: - incorporating the cultured cell into a food product for animal, preferably human, consumption.
  • the invention provides for a cell obtainable by or obtained by the method as described herein.
  • the invention provides for a cultured meat product for animal, preferably human, consumption, comprising at least one cell obtained by the method as described herein, optionally further comprising (cultured) mammalian myocytes.
  • the invention provides for a cultured fat product or cultured muscle product for animal, preferably human, consumption, comprising at least one cell obtainable by a method as described herein.
  • the invention provides for the use of a cell obtained according to the method as described herein or use of the method as described for tissue engineering, optionally for the production of cultured meat.
  • At least a particular value means that particular value or more.
  • at least 2 is understood to be the same as “2 or more” i.e., 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 , 12, 13, 14, 15, ... ,etc.
  • the word “about” or “approximately” when used in association with a numerical value preferably means that the value may be the given value (of 10) more or less 0.1 % of the value.
  • “Expansion” refers to the proliferation of a cell with or without differentiation and may include no passaging, one passage or more than one passage and/or serial passages.
  • a “medium” or “cell culture medium” refers to an aqueous based solution that provides for the growth, viability, or storage of cells.
  • a medium as contemplated herein can be supplemented with one or more nutrients to promote the desired cellular activity, such as cell viability, growth, proliferation, differentiation of the cells cultured in the medium.
  • a medium, as used herein, includes a serum replacement, a medium supplement, a complete medium or a cryopreservation medium.
  • the pH of a culture medium should be suitable to the microorganisms that will be grown. Most bacteria grow in pH 6.5-7.0 while most animal cells thrive in pH 7.2-7 .4.
  • “Aggregate” refers to an association of cells in which the association is caused by cell-cell interaction rather than adherence to a substrate.
  • two or more cells associate with each other by biologic attachments to one another. This can be through surface proteins, such as extracellular matrix proteins.
  • cells spontaneously associate in suspension to form cell-cell attachments independent of any adherence to a surface.
  • cells can be initially grown on a substrate where some cells associate with (adhere to) the substrate but further growth forms cell-cell associations (aggregation) that do not depend on association (adherence) of the further-grown cells with the substrate.
  • a cellular feeder layer is also considered a substrate. So attachment of cells to a feeder layer is also a form of adherent culture (not an aggregate) since attachment of the cells is not to each other but to the cells in the feeder layer.
  • cultured and “cultivated”, in the context of “cultured meat” and “cultivated meat”, “cultured fat/muscle”, cultivated fat/muscle” may be used interchangeably herein.
  • perfusion refers to the process of keeping culture cells alive by continuously feeding the cells with fresh media and removing spent media while keeping cells in culture. Perfusion is typically a continuous process. Perfusion culturing includes, but is not limited to continuous flow and semi-continuous flow, for example step-wise flow or staggered flow. In one embodiment of the method as described herein, during the expansions and/or proliferation of the cells, the average diameter of each expanded cell aggregate is no more than about 250 micron in size. In one embodiment of the method as described herein, during the expansions and/or proliferation of the cells, the average diameter of each expanded cell aggregate is no more than about 200 micron in size.
  • the average diameter of each expanded cell aggregate is no more than about 150 micron in size. In one embodiment of the method as described herein, during the expansions and/or proliferation of the cells, the average diameter of each expanded cell aggregate is no more than about 100 micron in size. In one embodiment of the method as described herein, during the expansions and/or proliferation of the cells, the average diameter of each expanded cell aggregate is no more than about 50 micron in size. It will be appreciated that where size is indicated throughout the document, the size refers to an average size in a population of aggregates.
  • shear forces are a common phenomenon.
  • a shear force is understood to exist when there is a first force acting on (part of) a composition in a first direction, and a second force acting on (part of) the composition that is stationary or moving in a second non-aligned direction.
  • Shear rate is the rate at which a progressive shearing deformation is applied. For a simple case, this can be the gradient of velocity in a flowing material. Shear rate is expressed in “reciprocal seconds, i.e. in “sec-1".
  • the a shear rate applied to the cultured pluripotent cells in the range of about 990 sec-1 to 2829 sec-1 , about 1100 sec-1 to 2200 sec-1 , or about 1500 sec-1 to 2000 sec- 1 . In preferred embodiments, the shear rate is about 1800 sec-1 , such as about 1839 sec-1 .
  • T the average shear stress
  • A the cross-sectional area of material with area parallel to the applied force vector.
  • a shear stress is applied in the range from about 1x10 -1 , about 10 -1 5 - about 1 -10’ 2 Pascal to about 1 10° Pascal.
  • shear stress is applied by use of an Alternating Tangential Flow (ATF).
  • ATF Alternating Tangential Flow
  • the ATF system uses hollow fibres over which the shear is homogeneously distributed. Due to slow alternating flows (every ⁇ 10 seconds), the change in pressure is relatively low which also results in lower shear stress peaks and an overall lower stress rate (990 - 2829 sec- 1).
  • a cell used in the invention may be a “pluripotent stem cell”.
  • the term "pluripotent stem cells” includes embryonic stem cells, embryo-derived stem cells, epiblast- derived stem cells, induced pluripotent stem cells and somatic cells, regardless of the method by which the pluripotent stem cells are derived.
  • the pluripotent stem cell is selected from the group consisting of embryonic stem cells, embryo-derived stem cell lines, epiblast-derived stem cell lines, induced pluripotent stem cells, embryonic cell lines, somatic cell lines and and immortalized cell lines.
  • the pluripotent stem cells are epiblast-derived stem cells (EpiSC).
  • EpiSC epiblast-derived stem cells
  • pluripotent stem cells express one or more markers selected from the group consisting of: OCT-4, Sox2, Klf4, c-MYC, Nanog, Lin28, alkaline phosphatase, SSEA-3, SSEA-4, TRA-1-60, and TRA-1-81.
  • Exemplary pluripotent stem cells can be generated using, methods known in the art. "Induced pluripotent stem cells" (iPS cells or iPSC) can be produced by protein transduction of reprogramming factors in a somatic cell.
  • the pluripotent stem cell according to the invention can be from any species. Embryonic stem cells have been successfully derived in, for example, mice, multiple species of non-human primates, and humans, and embryonic stem-like cells have been generated from numerous additional species. Thus, one of skill in the art can generate pluripotent stem cells, such as embryonic stem cells, embryo-derived stem cells, epiblast-derived stem cells (EpiSC), from any species, including but not limited to human, non-human primates, rodents (mice, rats), ungulates (cows, sheep, etc.), dogs (domestic and wild dogs), cats (domestic and wild cats such as lions, tigers, cheetahs), rabbits, hamsters, gerbils, squirrel, guinea pig, goats, elephants, panda (including giant panda), pigs, raccoon, horse, zebra, marine mammals (dolphin, whales, etc.) and the like.
  • iPS cells can be from any species. These iPS cells have been successfully generated using mouse and human cells. Furthermore, iPS cells have been successfully generated using embryonic, fetal, newborn, and adult tissue. Accordingly, one can readily generate iPS cells using a donor cell from any species.
  • the pluripotent stem cell according to the invention, or for use in the invention is an animal cell. In certain embodiments the pluripotent stem cell according to the invention, or for use in the invention if from an edible animal species.
  • the cell is from a mammal, preferably a non-human mammal.
  • the cell is an animal cell.
  • cell according to the invention is from an edible non-human animal species.
  • the cells produced by the method as described herein are suitable for human and non-human dietary consumption.
  • the pluripotent stem cells are of a livestock or poultry species.
  • Poultry species include but are not limited to domestic chicken, turkeys, ducks, geese and pigeons.
  • the cells originate from common game species such as wild deer, gallinaceous fowl, waterfowl and hare.
  • Livestock species include but are not limited to domestic cattle, pigs, sheep, goats, lamb, camels, water buffalo and rabbits.
  • the cell is a cell that is naturally capable of lipid accumulation such as a fat cells (adipocytes) or precursor cells thereof (such as fibro-adipogenic progenitor cells, lipoblasts, adipoblasts and, pre-adipocytes) or liver cells (hepatocyte) or precursor cells thereof (such as pre-adipocytes, hepatoblasts and pre-heptatocytes).
  • adipocytes a fat cells
  • precursor cells thereof such as fibro-adipogenic progenitor cells, lipoblasts, adipoblasts and, pre-adipocytes
  • liver cells hepatocyte
  • precursor cells thereof such as pre-adipocytes, hepatoblasts and pre-heptatocytes.
  • the cells are derived from an immortalized cell line.
  • exemplary cell lines include, but are not limited to, 3T3-L1 (mouse pre-adipocytes), buffalo rat liver cells (BRL 3A), chicken liver cells (LMH),
  • the cell is an adipocyte originating from a domestic pig or domestic cattle.
  • the cell is an hepatocyte originating from a duck or a goose.
  • the cell is a cell that is a muscle cell (myocyte), such as a skeletal muscle cell, or a or precursor cell thereof.
  • myocyte muscle cell
  • the cell is a muscle cell originating from a domestic pig or domestic cattle.
  • the cell is a pluripotent cell.
  • the pluripotent cell is selected from the group consisting of embryonic stem cells, induced pluripotent stem cells, embryonic cell lines, and somatic cell lines.
  • the pluripotent stem cell is a non-human animal cell, such as a porcine or a bovine pluripotent stem cell.
  • a porcine pluripotent stem cell is a non-human animal cell, such as a porcine or a bovine pluripotent stem cell.
  • a porcine pluripotent stem cell may be a fat cell or a precursor cell thereof.
  • Such a cell may be a muscle cell or a precursor cell thereof.
  • the stem cell according to the invention is a porcine epiblast stem cell (pEpiSCs) or a bovine epiblast stem cell (pEpiSCs).
  • pEpiSCs porcine epiblast stem cell
  • pEpiSCs bovine epiblast stem cell
  • Such a cell may be a fat cell or a precursor cell thereof.
  • Such a cell may be a muscle cell or a precursor cell thereof.
  • a pluripotent stem cell according to the invention is not a human cell.
  • the cells as used in the methods as described herein are described in patent application nos. W02024/084082, WO2024/170696, WO2024/170702 and W02024/252001 . Any cell described in any one of those patent applications may be useful in the methods of the invention for culturing cells.
  • the method according to the invention is for the proliferation (i.e. expansion) of the pluripotent cell.
  • the method as described herein can be used for differentiation the pluripotent cell.
  • the culture medium is a basal medium that is supplemented with said medium components.
  • a basal medium may be provided in liquid or powdered format.
  • a basal medium that is not supplemented with any compound may enable cellular growth, but supplementation may be required for growth depending on the cell type.
  • a basal medium may be supplemented with one or more components selected from the nonlimiting group consisting of amino acids, lipids, sugars, carbohydrates, anions, cations, buffering agents, colorants, vitamins, antioxidants, hormones, enzymes, proteins and trace elements.
  • the basal medium as disclosed herein is a commercially available basal medium such as DMEM (Dulbecco’s Modified Eagle Medium) and Ham’s F- 12.
  • the basal medium is the medium described in any of patent application nos. W02024/084082, WO2024/170696 and WO2024/170702 and W02024/252001 , which are incorporated by reference herein. Any of the medial described in any one of those patent applications may be used as basal medium in the methods of the invention.
  • the methods as described in the various embodiments of the invention further comprises the step of:
  • the invention provides for a cell obtainable by or obtained by the methods as described in the various embodiments as described herein.
  • cells obtained by the method as described herein are suitable for human and non-human dietary consumption.
  • the invention provides for a food product (also referred to as “foodstuff’) comprising the cells produced, obtained by the methods and/or cultured in any of the mediums as described herein.
  • a food product also referred to as “foodstuff’
  • the food product is for animal, preferably human, consumption.
  • a food product of the present invention may comprise one or more of minerals, synthetic substances, flavors, texture enhancers, nutritional additives, preservatives, and fats.
  • the flavors are selected from one or more of essential oils, oleoresin (ESO), enzymes (ENZ), natural substances and extractives (NAT), non-nutritive sweetener (NNS), nutritive sweetener (NUTRS), herbs, spices, natural seasonings & flavorings (SP), and synthetic flavors (SY/FL), fumigant (FUM), artificial sweeteners and yeast extract.
  • the texture enhancers are selected from one or more of pureed plant material, guar gum, cellulose, hemicellulose, lignin, beta glucans, soy, wheat, maize and rice isolates and beet fiber, pea fiber, bamboo fiber, plant derived fiber, plant derived gluten, carrageenan, xanthan gum, lecithin, pectin, agar, alginate, natural polysaccharides, grain husk, calcium citrate, calcium phosphates, calcium sulfate, magnesium sulfate and salts.
  • the nutritional additives are selected from one or more of trace elements, bioactive compounds, endogenous antioxidants, A, B-complex, C, D, E vitamins, zinc, thiamin, riboflavin, selenium, iron, niacin, potassium, phosphorus, omega-3, omega-6, fatty acids, magnesium, protein, amino acids salt, creatine, taurine, carnitine, carnosine, ubiquinone, glutathione, choline, glutathione, lipoic acid, spermine, anserine, linoleic acid, pantothenic acid, cholesterol, Retinol, folic acid, dietary fiber and amino acids.
  • the fats are selected from one or more of saturated, monounsaturated, polyunsaturated fats, corn oil, canola oil, sunflower oil, safflower oil, olive oil, peanut oil, soybean, flax seed oil, sesame oil, canola oil, avocado oil, seed oils, nut oils, safflower and sunflower oils, palm oil, coconut oil, omega-3, fish oil, lard, butter, processed animal fat, adipose tissue, cellular agriculture derived fat essential oil and oleoresin.
  • the preservative and/or antioxidant is selected from one or more of: sodium salt, chloride salt, iodine salt. Nitrates, nitrosamines.
  • BHA butylated hydroxyanisole
  • BHT butylated hydroxytoluene
  • MSG monosodium glutamate
  • sulphur dioxide sulphites
  • antibiotics antibiotics. It is noted here that any one additive, flavor, texture enhancer, nutrient additive, fat/oil and/or preservative/antioxidant may supply more than one attribute to food product of the present invention.
  • the food product of the present invention may further comprise (cultured) myocytes.
  • the food products comprises the cultured myocytes as described in patent application nos. WO2024/170696 and WO2024/170702. Any cell described in any one of those patent applications may be useful in a method of the invention for culturing cells and in a food product of the invention.
  • the food product of the present invention may further comprise (cultured) adipocytes.
  • the food products comprises the cultured adipocytes as described in patent application no. W02024/084082. Any cell described in any one of those patent applications may be useful in a method of the invention for culturing cells.
  • a food product of the invention may comprise one or more types of cell, for example two or more types of cell. At least one of the cell types is a cultured cell.
  • a food product of the invention may comprise two or more types of cultured cell, one or two or more of which may be obtained using a method of the invention.
  • the food product is a cultured meat product, for example a cultured fat product or a culture muscle product.
  • the cultured meat product may be a structured product.
  • the product is structured with scaffolding.
  • the structured product is free of scaffolding and the structure is provided by the cells as obtained by the methods as described herein.
  • the invention provides for a cultured fat product or a cultured muscle product for animal, preferably human, consumption, comprising at least one cell obtainable by a method as described herein.
  • the invention provides for a use of the cell obtained or obtainable by the method as described herein for tissue engineering, optionally for the production of cultured meat.
  • Figure 1 Cell-aggregate staining. Nuclei were visualized with DAPI (blue), dead cells (green) are visualized with LIVE/DEADTM Fixable Green stain. Image was taken with a EVOS M7000 microscope.
  • Figure 2 Repeated fed-batch culture for 5 days.
  • A) shows the median cell aggregate diameter over time in pm.
  • Figure 3 Cell culture under perfusion with stainless steel filter. A) shows the median cell aggregate diameter over time in pm. B) Cell density over time.
  • FIG. 4 Cell culture with ATF.
  • A) shows the median cell aggregate diameter over time in pm.
  • Figure 5 Cell culture with peristatic pump recirculation cell viability after 30min of culture.
  • Cells are inoculated as single cells in bioreactor with inoculation media. Over time, the bioreactor is filled with feed media and cells will form aggregates. Once the maximal bioreactor volume is reached, the cells are harvested and require a single celling procedure to be seeded into a new bioreactor.
  • Cells are inoculated as single cells or small aggregates (depending on the specific experiment) with inoculation media. Over time, the bioreactor is filled with feed media and cells will form aggregates. Once the desired working volume is reached, perfusion will be started. Here the flow rate of broth out of the bioreactor will be equal to that of the feed media flowing in. In order to remove very little cells, different cell retention devices can be used. This allows cell to be retained in the bioreactor while spent media can be removed. In these examples two different setups are compared: ATF1 filter (Repligen, USA) and a submerged stainless steel stationary dead-end filter are used.
  • Example 1 Repeated fed-batch - no shear applied
  • a repeated fed-batch experiment was set-up in a 500 ml single use Appliflex bioreactor. Cells were grown in a regular fed-batch phase for 5 days. After that, the bioreactor was partially harvested (50% of volume removed) and filled again over the next 24 hours to repeated the process for 4 days (process time in total is 9 days). During these 9 days, samples were taken to analyze aggregate size distribution, growth rate and to quantify the change in several pluripotency markers over time. Results
  • ATF uses 60 cm hollow fibres over which the shear is homogeneously distributed. Due to slow alternating flows (every ⁇ 10 seconds), the change in pressure is relatively low which also results in lower shear stress peaks and an overall lower stress rate (990 - 2829 sec 1 ). The process was very comparable to the one described in Experiment 2. However, perfusion was started on day 5 instead of day 2 and the run was continued for a total of 12 days.
  • peristaltic pump a D-SP086 with Masterflex 06402-25 Tygon tubing at ⁇ 150 ml/min.
  • peristaltic pump the pressure differences are significantly higher and more frequent (multiple times per sec) and therefore the cells also tend to experience very high peaks of shear stress and an overall higher shear rate of about 3000 sec 1 .
  • a method for reducing the size of stem cell aggregates during cell culture comprising culturing the pluripotent cells by perfusion in a cell culture medium and applying shear to the medium. 2. The method according to embodiment 1 , wherein a shear rate is applied in the range from about 90 sec-1 to 2829 sec-1 , preferably about 1100 sec-1 to 2200 sec-1 , or more preferably about 1500 sec-1 to 2000 sec-1.
  • the cell is selected from the group consisting of pluripotent stem cells, embryonic stem cells, induced pluripotent stem cells, embryonic cell lines, somatic cell lines, and immortalized cell lines.
  • a cell or cell culture obtainable by or obtained by the method according to any of the preceding embodiments.
  • a cultured meat product for animal, preferably human, consumption comprising at least one cell obtained by the method according to any one of embodiments 1-9, optionally further comprising a second cell type, such as (cultured) mammalian myocytes.

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Abstract

La présente invention concerne un procédé de culture de cellules souches pluripotentes par perfusion, le procédé comprenant la culture des cellules pluripotentes dans un milieu de culture cellulaire et l'application d'un cisaillement au milieu. L'invention concerne également des cellules obtenues par les procédés, ainsi que des produits, tels que des produits alimentaires, comprenant les cellules obtenues par le procédé.
PCT/EP2024/088240 2023-12-21 2024-12-20 Procédés de culture de cellules souches pluripotentes par perfusion Pending WO2025133344A1 (fr)

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US20220195359A1 (en) * 2019-05-02 2022-06-23 Aleph Farms Ltd. Cultivation systems and methods for large-scale production of cultured food
WO2024084082A1 (fr) 2022-10-21 2024-04-25 Meatable B.V. Maturation d'adipocytes
WO2024170696A1 (fr) 2023-02-15 2024-08-22 Meatable B.V. Maturation de cellules musculaires squelettiques
WO2024170702A1 (fr) 2023-02-15 2024-08-22 Meatable B.V. Maturation de cellules musculaires squelettiques
WO2024252001A1 (fr) 2023-06-08 2024-12-12 Meatable B.V. Procédés de culture de cellules

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