JPWO2022144489A5 - - Google Patents
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- JPWO2022144489A5 JPWO2022144489A5 JP2023540580A JP2023540580A JPWO2022144489A5 JP WO2022144489 A5 JPWO2022144489 A5 JP WO2022144489A5 JP 2023540580 A JP2023540580 A JP 2023540580A JP 2023540580 A JP2023540580 A JP 2023540580A JP WO2022144489 A5 JPWO2022144489 A5 JP WO2022144489A5
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- 229920002678 cellulose Polymers 0.000 claims description 104
- 239000001913 cellulose Substances 0.000 claims description 104
- 210000004027 cell Anatomy 0.000 claims description 57
- 238000000034 method Methods 0.000 claims description 25
- 239000006143 cell culture medium Substances 0.000 claims description 18
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 8
- 239000012736 aqueous medium Substances 0.000 claims description 7
- 239000011148 porous material Substances 0.000 claims description 7
- 238000000926 separation method Methods 0.000 claims description 7
- 210000004748 cultured cell Anatomy 0.000 claims description 6
- 239000000017 hydrogel Substances 0.000 claims description 6
- 239000011324 bead Substances 0.000 claims description 4
- 239000000499 gel Substances 0.000 claims description 4
- 238000004113 cell culture Methods 0.000 claims description 3
- 210000000130 stem cell Anatomy 0.000 claims description 3
- 229920001046 Nanocellulose Polymers 0.000 claims description 2
- 210000000577 adipose tissue Anatomy 0.000 claims description 2
- 239000011247 coating layer Substances 0.000 claims description 2
- 238000011437 continuous method Methods 0.000 claims description 2
- 238000007599 discharging Methods 0.000 claims description 2
- 239000012510 hollow fiber Substances 0.000 claims description 2
- 239000002609 medium Substances 0.000 claims description 2
- 238000003306 harvesting Methods 0.000 claims 1
- 210000001808 exosome Anatomy 0.000 description 7
- 239000012528 membrane Substances 0.000 description 2
- 230000028327 secretion Effects 0.000 description 2
- KIUKXJAPPMFGSW-DNGZLQJQSA-N (2S,3S,4S,5R,6R)-6-[(2S,3R,4R,5S,6R)-3-Acetamido-2-[(2S,3S,4R,5R,6R)-6-[(2R,3R,4R,5S,6R)-3-acetamido-2,5-dihydroxy-6-(hydroxymethyl)oxan-4-yl]oxy-2-carboxy-4,5-dihydroxyoxan-3-yl]oxy-5-hydroxy-6-(hydroxymethyl)oxan-4-yl]oxy-3,4,5-trihydroxyoxane-2-carboxylic acid Chemical compound CC(=O)N[C@H]1[C@H](O)O[C@H](CO)[C@@H](O)[C@@H]1O[C@H]1[C@H](O)[C@@H](O)[C@H](O[C@H]2[C@@H]([C@@H](O[C@H]3[C@@H]([C@@H](O)[C@H](O)[C@H](O3)C(O)=O)O)[C@H](O)[C@@H](CO)O2)NC(C)=O)[C@@H](C(O)=O)O1 KIUKXJAPPMFGSW-DNGZLQJQSA-N 0.000 description 1
- 229920000936 Agarose Polymers 0.000 description 1
- 102000008186 Collagen Human genes 0.000 description 1
- 108010035532 Collagen Proteins 0.000 description 1
- 229920001436 collagen Polymers 0.000 description 1
- 210000005260 human cell Anatomy 0.000 description 1
- 229920002674 hyaluronan Polymers 0.000 description 1
- 229960003160 hyaluronic acid Drugs 0.000 description 1
- 210000004962 mammalian cell Anatomy 0.000 description 1
- 210000002901 mesenchymal stem cell Anatomy 0.000 description 1
- 230000010412 perfusion Effects 0.000 description 1
- 210000000229 preadipocyte Anatomy 0.000 description 1
- 230000004936 stimulating effect Effects 0.000 description 1
- 238000002626 targeted therapy Methods 0.000 description 1
- 238000011287 therapeutic dose Methods 0.000 description 1
Description
細胞培養フラスコフラスコと組み合わせたＮＦＣは、インビボ様エキソソーム分泌を増強した。測定されたサイズ分布がさらに狭いことから、Ｄｄ中のエキソソームの形態は、２Ｄエキソソームよりも同一であった。この結果は、均質性が高い産物の方が良好な標的化治療をもたらすと仮定することができるため、さらに機能的な又は「インビボ様」エキソソームに関する結論を裏付けている。インビボ様エキソソーム分泌の刺激におけるＮＦＣの使用は、臨床応用のためのエキソソーム治療投与量の送達における新しい枠組みをさらに効率的に提供する。
＜付記＞
本開示は以下の態様を含む。
＜項１＞
容器（１２）と、
細胞培養培地を前記容器に投入するための入口（１４）と、
細胞由来産物を含む細胞培養培地を前記容器から排出するための出口（１６）と、
を備え、
前記容器（１２）が、細胞を受容するように構成されたナノ構造セルロースを含む区画（１８）を備えるか、又はそれに接続され、前記区画が、前記ナノ構造セルロースを前記出口から分離し、細胞由来産物を含む細胞培養培地が第１の分離表面（２０）を通過することを可能にする第１の分離表面（２０）を備え、ガス交換のためのさらなる分離表面（３０）を任意に備える、培養細胞から細胞由来産物を抽出するためのバイオリアクタ（１０）。
＜項２＞
前記第１の分離表面（２０）が、前記細胞由来産物を含む細胞培養培地に対して透過性であり、細胞及びナノ構造セルロースに対して不透過性である膜若しくはメッシュなどの構造、例えば、０．４～３．０μｍの範囲内の平均細孔径を有する膜を備えるか、又はその形態である、＜項１＞に記載のバイオリアクタ。
＜項３＞
前記区画（１８）内の前記ナノ構造セルロースと細胞とを混合するための手段（２４）を備え、例えば、混合するための前記手段が、ミキサ、培地の流れを提供するための手段、及び／又は前記容器若しくは区画の容積を調整するための手段を備える、＜項１＞又は＜項２＞に記載のバイオリアクタ。
＜項４＞
前記入口（１４）及び／又は前記出口（１６）を介して細胞培養培地の流れを提供するための手段、好ましくはナノ構造セルロースを含む前記区画（１８）を通る前記手段、例えば灌流バイオリアクタを得るための前記手段、を備える、＜項１＞～＜項３＞のいずれか一項に記載のバイオリアクタ。
＜項５＞
ナノ構造セルロースを含む前記区画（１８）が、１又は複数の表面上に、細胞培養ゲル、例えば、アガロース、コラーゲン若しくはヒアルロン酸又はそれらの誘導体を含むゲル、例えば、０．３～１．０％（ｗ／ｗ）の濃度を有するゲル、のコーティング層を備える、＜項１＞～＜項４＞のいずれか一項に記載のバイオリアクタ。
＜項６＞
ナノ構造セルロースを含む前記区画（１８）が、前記容器（１２）とは別個のユニット（３６）、例えば、固定床リアクタである、＜項１＞～＜項５＞のいずれか一項に記載のバイオリアクタ。
＜項７＞
前記第１の表面が、中空糸の形態である、＜項１＞～＜項６＞のいずれか一項に記載のバイオリアクタ。
＜項８＞
前記容器が、フラスコ又はボトルを備える、＜項１＞～＜項７＞のいずれか一項に記載のバイオリアクタ。
＜項９＞
前記第１の分離表面が、前記ナノ構造セルロースの表面によって形成される、＜項１＞～＜項８＞のいずれか一項に記載のバイオリアクタ。
＜項１０＞
前記ナノ構造セルロースが、ヒドロゲルビーズ及び／又は多孔質ビーズなどの別個の実体の形態である、＜項１＞～＜項９＞のいずれか一項に記載のバイオリアクタ。
＜項１１＞
前記ナノ構造セルロースが、２００ｎｍ以下の数平均直径を有するセルロースフィブリル及び／若しくはフィブリル束を含むナノフィブリル状セルロースを含み、並びに／又は前記ナノフィブリル状セルロースが、水に分散した場合、２２±１℃の水性培地中で０．５重量％の稠度で回転レオメータにより決定される、１００～５００００Ｐａ・ｓの範囲内、例えば、３００～８０００Ｐａ・ｓの範囲内のゼロ剪断粘度と、１～５０Ｐａの範囲内、例えば、２～１５Ｐａの範囲内の降伏応力とを提供する、＜項１＞～＜項１０＞のいずれか一項に記載のバイオリアクタ。
＜項１２＞
前記ナノフィブリル状セルロースが、アニオン的に改質されたナノフィブリル状セルロース又はカチオン的に改質されたナノフィブリル状セルロースなどの化学的に改質されたナノフィブリル状セルロースである、＜項１１＞に記載のバイオリアクタ。
＜項１３＞
前記ナノフィブリル状セルロースが、化学的に改質されていないナノフィブリル状セルロース、好ましくはまた、酵素的に改質されていないナノフィブリル状セルロースである、＜項１１＞に記載のバイオリアクタ。
＜項１４＞
前記ナノフィブリル状セルロースが、２～５０ｎｍ、例えば、２～２０ｎｍの範囲内の数平均直径を有するセルロースフィブリル及び／又はフィブリル束を含み、前記ナノフィブリル状セルロースが、水に分散した場合、２２±１℃の水性培地中で０．５重量％の稠度で回転レオメータにより決定される、１～５０Ｐａの範囲内、例えば、２～１５Ｐａの範囲内の降伏応力を提供し、好ましくは、前記ナノフィブリル状セルロースが、化学的に改質されていないナノフィブリル状セルロースである、＜項１１＞～＜項１３＞のいずれか一項に記載のバイオリアクタ。
＜項１５＞
前記ナノ構造セルロースが、２～４０ｎｍ、例えば、２～２０ｎｍの範囲内の数平均フィブリル直径と、１００ｎｍ以上、例えば、１００～４００ｎｍの範囲内の数平均フィブリル長とを有するナノ結晶セルロースを含む、＜項１＞～＜項１０＞のいずれか一項に記載のバイオリアクタ。
＜項１６＞
前記ナノ構造セルロースが、０．１～１０μｍ
２
、例えば、０．３～２．０μｍ
２
の範囲内の中心細孔径を有する、＜項１＞～＜項１５＞のいずれか一項に記載のバイオリアクタ。
＜項１７＞
前記細胞由来産物が、１０００ｎｍ以下の数平均直径を有する、＜項１＞～＜項１６＞のいずれか一項に記載のバイオリアクタ。
＜項１８＞
＜項１＞～＜項１７＞のいずれか一項に記載のバイオリアクタ（１０）を提供すること、
細胞培養培地、好ましくは無血清、動物起源不含、フィーダー不含及び／又は異種物不含の細胞培養培地を提供すること、
細胞を提供すること、
ナノ構造セルロースを含む区画内で前記細胞をインキュベートして細胞由来産物を形成し、任意に前記ナノ構造セルロースと前記細胞とを混合すること、
細胞由来産物が前記細胞から前記細胞培養培地中に拡散することを可能にすること、
好ましくは前記細胞由来産物を含む前記細胞培養培地を抽出するための出口を介して、前記細胞由来産物を含む前記細胞培養培地を前記バイオリアクタから採取して、前記インキュベートされた細胞から前記細胞由来産物を分離すること、
を含む、培養細胞から細胞由来産物を抽出する方法。
＜項１９＞
前記方法が連続的な方法であり、好ましくは、前記方法が、細胞由来産物を含む前記細胞培養培地が前記バイオリアクタから採取される速度と等しい速度で新しい細胞培養培地を前記バイオリアクタに添加することを含む、＜項１８＞に記載の方法。
＜項２０＞
前記細胞由来産物が、エキソソームなどの細胞外小胞を含み、好ましくは、前記細胞が、ヒト細胞などの哺乳動物細胞であり、例えば、前記細胞が、幹細胞、例えば、間葉系幹細胞などの初代細胞であり、及び／又は前記細胞が、スフェロイドなどの凝集体として存在する、＜項１８＞又は＜項１９＞に記載の方法。
＜項２１＞
前記細胞由来産物が、１０００ｎｍ以下の数平均直径を有する、＜項１８＞～＜項２０＞のいずれか一項に記載の方法。
＜項２２＞
前記細胞が、マウス胚由来脂肪前駆細胞（ＡＰＣ）などの脂肪組織から得られた幹細胞である、＜項２０＞又は＜項２１＞に記載の方法。
＜項２３＞
第１の分離表面が、前記ナノ構造セルロースの表面によって形成される、＜項１８＞～＜項２２＞のいずれか一項に記載の方法。
＜項２４＞
前記ナノ構造セルロースが、２～５０ｎｍ、例えば、２～２０ｎｍの範囲内の数平均直径を有するセルロースフィブリル及び／又はフィブリル束を含むナノフィブリル状セルロースであり、前記ナノフィブリル状セルロースが、水に分散した場合、２２±１℃の水性培地中で０．５重量％の稠度で回転レオメータにより決定される、１～５０Ｐａの範囲内、例えば、２～１５Ｐａの範囲内の降伏応力を提供し、好ましくは、前記ナノフィブリル状セルロースが、化学的に改質されていないナノフィブリル状セルロースである、＜項１８＞～＜項２３＞のいずれか一項に記載の方法。
＜項２５＞
前記ナノ構造セルロースが、０．１～１０μｍ
２
、例えば、０．３～２．０μｍ
２
の範囲内の中心細孔径を有するナノ構造セルロースヒドロゲルを含む、＜項１８＞～＜項２４＞のいずれか一項に記載の方法。
＜項２６＞
培養細胞から細胞由来産物を抽出するためのナノ構造セルロース品であって、０．１～１０μｍ
２
、例えば、０．３～２．０μｍ
２
の範囲内の中心細孔径を有するナノ構造セルロースヒドロゲルを含むナノ構造セルロース品。
＜項２７＞
ナノ構造セルロースが、２～５０ｎｍ、例えば、２～２０ｎｍの範囲内の数平均直径を有するセルロースフィブリル及び／又はフィブリル束を含むナノフィブリル状セルロースであり、前記ナノフィブリル状セルロースが、水に分散した場合、２２±１℃の水性培地中で０．５重量％の稠度で回転レオメータにより決定される、１～５０Ｐａの範囲内、例えば、２～１５Ｐａの範囲内の降伏応力を提供する、＜項２６＞に記載のナノ構造セルロース品。
＜項２８＞
前記ナノフィブリル状セルロースが、アニオン的に改質されたナノフィブリル状セルロース又はカチオン的に改質されたナノフィブリル状セルロースなどの化学的に改質されたナノフィブリル状セルロースである、＜項２６＞又は＜項２７＞に記載のナノ構造セルロース品。
＜項２９＞
前記ナノフィブリル状セルロースが、化学的に改質されていないナノフィブリル状セルロース、好ましくはまた、酵素的に改質されていないナノフィブリル状セルロースである、＜項２６＞又は＜項２７＞に記載のナノ構造セルロース品。
＜項３０＞
好ましくは＜項１８＞～＜項２５＞のいずれか一項に記載の方法を用いて、インキュベートされた細胞から細胞由来産物を抽出するための、＜項２６＞～＜項２９＞のいずれか一項に記載のナノ構造セルロース品の使用。
＜項３１＞
例えば、小胞の数平均直径が、２００ｎｍ以下、さらに具体的には１５０ｎｍ以下、又は１３０ｎｍ以下、例えば、９０～２００ｎｍ、９０～１５０ｎｍ、９０～１３０ｎｍ、１００～２００ｎｍ、１００～１５０ｎｍ、若しくは１００～１３０ｎｍの範囲内である、＜項１８＞～＜項２５＞のいずれか一項に記載の方法を用いて得られた細胞外小胞。
NFC in combination with cell culture flask enhanced in vivo-like exosome secretion. The morphology of exosomes in Dd was more uniform than in 2D exosomes, as the measured size distribution was narrower. This result supports the conclusion regarding more functional or "in vivo-like" exosomes, as it can be hypothesized that products with higher homogeneity will yield better targeted therapy. The use of NFC in stimulating in vivo-like exosome secretion provides a new framework for more efficient delivery of exosome therapeutic doses for clinical applications.
<Additional notes>
The present disclosure includes the following aspects.
<Section 1>
a container (12);
an inlet (14) for introducing cell culture medium into the container;
an outlet (16) for discharging cell culture medium containing cell-derived products from said container;
Equipped with
Said container (12) comprises or is connected to a compartment (18) comprising nanostructured cellulose configured to receive cells, said compartment separating said nanostructured cellulose from said outlet and containing cells. comprising a first separation surface (20) allowing cell culture medium containing derived products to pass through the first separation surface (20), optionally comprising a further separation surface (30) for gas exchange; , a bioreactor (10) for extracting cell-derived products from cultured cells.
<Section 2>
The first separation surface (20) is a structure such as a membrane or mesh that is permeable to the cell culture medium containing the cell-derived products and impermeable to cells and nanostructured cellulose, e.g. The bioreactor according to item 1, comprising or in the form of a membrane having an average pore size within the range of 0.4 to 3.0 μm.
<Section 3>
means (24) for mixing said nanostructured cellulose and cells in said compartment (18), for example said means for mixing comprising a mixer, means for providing a flow of medium, and/or Or the bioreactor according to <Item 1> or <Item 2>, comprising means for adjusting the volume of the container or compartment.
<Section 4>
means for providing a flow of cell culture medium through said inlet (14) and/or said outlet (16), preferably through said compartment (18) comprising nanostructured cellulose, e.g. a perfusion bioreactor; The bioreactor according to any one of <Section 1> to <Section 3>, comprising the means for obtaining the bioreactor.
<Section 5>
Said compartment (18) comprising nanostructured cellulose comprises on one or more surfaces a cell culture gel, for example a gel containing agarose, collagen or hyaluronic acid or derivatives thereof, for example 0.3-1.0%. The bioreactor according to any one of <Section 1> to <Section 4>, comprising a coating layer of a gel having a concentration of (w/w).
<Section 6>
According to any one of paragraphs 1 to 5, wherein the compartment (18) comprising nanostructured cellulose is a separate unit (36) from the container (12), for example a fixed bed reactor. bioreactor.
<Section 7>
The bioreactor according to any one of <Section 1> to <Section 6>, wherein the first surface is in the form of a hollow fiber.
<Section 8>
The bioreactor according to any one of <Item 1> to <Item 7>, wherein the container comprises a flask or a bottle.
<Section 9>
The bioreactor according to any one of <Item 1> to <Item 8>, wherein the first separation surface is formed by the surface of the nanostructured cellulose.
<Section 10>
The bioreactor according to any one of Items 1 to 9, wherein the nanostructured cellulose is in the form of separate entities such as hydrogel beads and/or porous beads.
<Section 11>
When the nanostructured cellulose comprises nanofibrillar cellulose comprising cellulose fibrils and/or fibril bundles having a number average diameter of 200 nm or less, and/or the nanofibrillar cellulose is dispersed in water, the temperature is 22±1°C. zero shear viscosity in the range 100 to 50 000 Pa s, e.g. The bioreactor according to any one of <Item 1> to <Item 10>, which provides a yield stress within the range of, for example, 2 to 15 Pa.
<Section 12>
<Item 11>, wherein the nanofibrillar cellulose is chemically modified nanofibrillar cellulose such as anionically modified nanofibrillar cellulose or cationically modified nanofibrillar cellulose. The bioreactor described in .
<Section 13>
The bioreactor according to <Item 11>, wherein the nanofibrillar cellulose is chemically unmodified nanofibrillar cellulose, preferably also enzymatically unmodified nanofibrillar cellulose.
<Section 14>
When the nanofibrillar cellulose comprises cellulose fibrils and/or fibril bundles having a number average diameter in the range of 2 to 50 nm, for example 2 to 20 nm, and the nanofibrillar cellulose is dispersed in water, 22± Preferably, said nanofibrils provide a yield stress in the range of 1 to 50 Pa, such as in the range of 2 to 15 Pa, determined by a rotational rheometer at a consistency of 0.5% by weight in an aqueous medium at 1°C. The bioreactor according to any one of <Item 11> to <Item 13>, wherein the cellulose is chemically unmodified nanofibrillar cellulose.
<Section 15>
The nanostructured cellulose comprises nanocrystalline cellulose having a number average fibril diameter in the range of 2 to 40 nm, such as 2 to 20 nm, and a number average fibril length in the range of 100 nm or more, such as 100 to 400 nm. The bioreactor according to any one of <Item 1> to <Item 10>.
<Section 16>
The nanostructured cellulose according to any one of <Item 1> to <Item 15>, wherein the nanostructured cellulose has a central pore diameter within the range of 0.1 to 10 μm 2 , for example, 0.3 to 2.0 μm 2 . bioreactor.
<Section 17>
The bioreactor according to any one of <Item 1> to <Item 16>, wherein the cell-derived product has a number average diameter of 1000 nm or less.
<Section 18>
Providing the bioreactor (10) according to any one of <Item 1> to <Item 17>;
providing a cell culture medium, preferably serum-free, animal origin-free, feeder-free and/or xenobiotic-free;
providing cells;
incubating said cells in a compartment comprising nanostructured cellulose to form a cell-derived product and optionally mixing said nanostructured cellulose and said cells;
allowing cell-derived products to diffuse from said cells into said cell culture medium;
The cell culture medium containing the cell-derived products is harvested from the bioreactor, preferably via an outlet for extracting the cell-culture medium containing the cell-derived products from the incubated cells. separating the products;
A method for extracting cell-derived products from cultured cells, including:
<Section 19>
Said method is a continuous method, preferably said method adds fresh cell culture medium to said bioreactor at a rate equal to the rate at which said cell culture medium containing cell-derived products is withdrawn from said bioreactor. The method according to <Section 18>, comprising:
<Section 20>
Said cell-derived product comprises extracellular vesicles such as exosomes, preferably said cell is a mammalian cell such as a human cell, for example said cell is a stem cell, for example a primary cell such as a mesenchymal stem cell. The method according to <Item 18> or <Item 19>, wherein the cell is a cell and/or the cell is present as an aggregate such as a spheroid.
<Section 21>
The method according to any one of <Section 18> to <Section 20>, wherein the cell-derived product has a number average diameter of 1000 nm or less.
<Section 22>
The method according to <Section 20> or <Section 21>, wherein the cells are stem cells obtained from adipose tissue such as mouse embryonic preadipocyte cells (APCs).
<Section 23>
The method according to any one of <Section 18> to <Section 22>, wherein the first separation surface is formed by the surface of the nanostructured cellulose.
<Section 24>
The nanostructured cellulose is nanofibrillar cellulose comprising cellulose fibrils and/or fibril bundles having a number average diameter in the range of 2 to 50 nm, for example 2 to 20 nm, and the nanofibrillar cellulose is dispersed in water. If so, it provides a yield stress in the range of 1 to 50 Pa, such as in the range of 2 to 15 Pa, as determined by a rotational rheometer at a consistency of 0.5% by weight in an aqueous medium at 22±1° C., preferably The method according to any one of <Item 18> to <Item 23>, wherein the nanofibrillar cellulose is chemically unmodified nanofibrillar cellulose.
<Section 25>
Any of <Item 18> to <Item 24>, wherein the nanostructured cellulose comprises a nanostructured cellulose hydrogel having a central pore diameter in the range of 0.1 to 10 μm 2 , for example, 0.3 to 2.0 μm 2 The method described in paragraph (1).
<Section 26>
A nanostructured cellulose product for extracting cell-derived products from cultured cells, the nanostructured cellulose hydrogel having a central pore size in the range of 0.1 to 10 μm 2 , such as 0.3 to 2.0 μm 2 . Nanostructured cellulose products containing.
<Section 27>
The nanostructured cellulose is nanofibrillar cellulose comprising cellulose fibrils and/or fibril bundles having a number average diameter in the range of 2 to 50 nm, such as 2 to 20 nm, and the nanofibrillar cellulose is dispersed in water. <terms> providing a yield stress in the range of 1 to 50 Pa, such as in the range of 2 to 15 Pa, as determined by a rotational rheometer at a consistency of 0.5% by weight in an aqueous medium at 22±1° C. 26>.
<Section 28>
<Item 26>, wherein the nanofibrillar cellulose is chemically modified nanofibrillar cellulose such as anionically modified nanofibrillar cellulose or cationically modified nanofibrillar cellulose. Or the nanostructured cellulose product according to <Item 27>.
<Section 29>
According to <Item 26> or <Item 27>, wherein the nanofibrillar cellulose is chemically unmodified nanofibrillar cellulose, preferably also enzymatically unmodified nanofibrillar cellulose. nanostructured cellulose products.
<Section 30>
Any one of <Item 26> to <Item 29> for extracting a cell-derived product from the incubated cells, preferably using the method described in any one of <Item 18> to <Item 25>. Use of the nanostructured cellulose product according to paragraph 1.
<Section 31>
For example, the number average diameter of the vesicles is 200 nm or less, more specifically 150 nm or less, or 130 nm or less, such as 90 to 200 nm, 90 to 150 nm, 90 to 130 nm, 100 to 200 nm, 100 to 150 nm, or 100 to Extracellular vesicles obtained using the method according to any one of <Item 18> to <Item 25>, which have a wavelength within the range of 130 nm.
Claims (32)
細胞培養培地を前記容器に投入するための入口（１４）と、
細胞由来産物を含む細胞培養培地を前記容器から排出するための出口（１６）と、
を備え、
前記容器（１２）が、細胞を受容するように構成されたナノ構造セルロースを含む区画（１８）を備えるか、又はそれに接続され、前記区画が、前記ナノ構造セルロースを前記出口から分離し、細胞由来産物を含む細胞培養培地が第１の分離表面（２０）を通過することを可能にする第１の分離表面（２０）を備える、培養細胞から細胞由来産物を抽出するためのバイオリアクタ（１０）。 a container (12);
an inlet (14) for introducing cell culture medium into the container;
an outlet (16) for discharging cell culture medium containing cell-derived products from said container;
Equipped with
Said container (12) comprises or is connected to a compartment (18) comprising nanostructured cellulose configured to receive cells, said compartment separating said nanostructured cellulose from said outlet and containing cells. A bioreactor for extracting cell-derived products from cultured cells ( 10).
細胞培養培地を提供すること、
細胞を提供すること、
ナノ構造セルロースを含む区画内で前記細胞をインキュベートして細胞由来産物を形成すること、
細胞由来産物が前記細胞から前記細胞培養培地中に拡散することを可能にすること、
前記細胞由来産物を含む前記細胞培養培地を前記バイオリアクタから採取して、前記インキュベートされた細胞から前記細胞由来産物を分離すること、
を含む、培養細胞から細胞由来産物を抽出する方法。 providing a bioreactor (10) according to any one of claims 1 to 17;
providing a cell culture medium;
providing cells;
incubating the cells within a compartment comprising nanostructured cellulose to form a cell-derived product;
allowing cell-derived products to diffuse from said cells into said cell culture medium;
harvesting the cell culture medium containing the cell-derived products from the bioreactor and separating the cell-derived products from the incubated cells;
A method for extracting cell-derived products from cultured cells, including:
Extracellular vesicles obtained using the method according to any one of claims 18 to 26 , wherein the vesicles have a number average diameter of 200 nm or less .
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| EP20397519.8 | 2020-12-31 | ||
| EP20397519.8A EP4023744A1 (en) | 2020-12-31 | 2020-12-31 | A bioreactor and a method for separating cell-derived products from cultured cells and a nanostructured cellulose product |
| PCT/FI2021/050576 WO2022144489A1 (en) | 2020-12-31 | 2021-08-26 | A bioreactor and a method for extracting cell-derived products from cultured cells and a nanostructured cellulose product |
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| EP3581591A1 (en) * | 2018-06-13 | 2019-12-18 | UPM-Kymmene Corporation | A nanofibrillar cellulose product and a method for manufacturing thereof |
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