JP6524597B2 - α−ガラクトースを除去する方法 - Google Patents
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- A61L27/00—Materials for grafts or prostheses or for coating grafts or prostheses
- A61L27/36—Materials for grafts or prostheses or for coating grafts or prostheses containing ingredients of undetermined constitution or reaction products thereof, e.g. transplant tissue, natural bone, extracellular matrix
- A61L27/3683—Materials for grafts or prostheses or for coating grafts or prostheses containing ingredients of undetermined constitution or reaction products thereof, e.g. transplant tissue, natural bone, extracellular matrix subjected to a specific treatment prior to implantation, e.g. decellularising, demineralising, grinding, cellular disruption/non-collagenous protein removal, anti-calcification, crosslinking, supercritical fluid extraction, enzyme treatment
- A61L27/3687—Materials for grafts or prostheses or for coating grafts or prostheses containing ingredients of undetermined constitution or reaction products thereof, e.g. transplant tissue, natural bone, extracellular matrix subjected to a specific treatment prior to implantation, e.g. decellularising, demineralising, grinding, cellular disruption/non-collagenous protein removal, anti-calcification, crosslinking, supercritical fluid extraction, enzyme treatment characterised by the use of chemical agents in the treatment, e.g. specific enzymes, detergents, capping agents, crosslinkers, anticalcification agents
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- A61L27/00—Materials for grafts or prostheses or for coating grafts or prostheses
- A61L27/14—Macromolecular materials
- A61L27/22—Polypeptides or derivatives thereof, e.g. degradation products
- A61L27/24—Collagen
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- A61L27/00—Materials for grafts or prostheses or for coating grafts or prostheses
- A61L27/36—Materials for grafts or prostheses or for coating grafts or prostheses containing ingredients of undetermined constitution or reaction products thereof, e.g. transplant tissue, natural bone, extracellular matrix
- A61L27/3683—Materials for grafts or prostheses or for coating grafts or prostheses containing ingredients of undetermined constitution or reaction products thereof, e.g. transplant tissue, natural bone, extracellular matrix subjected to a specific treatment prior to implantation, e.g. decellularising, demineralising, grinding, cellular disruption/non-collagenous protein removal, anti-calcification, crosslinking, supercritical fluid extraction, enzyme treatment
- A61L27/3695—Materials for grafts or prostheses or for coating grafts or prostheses containing ingredients of undetermined constitution or reaction products thereof, e.g. transplant tissue, natural bone, extracellular matrix subjected to a specific treatment prior to implantation, e.g. decellularising, demineralising, grinding, cellular disruption/non-collagenous protein removal, anti-calcification, crosslinking, supercritical fluid extraction, enzyme treatment characterised by the function or physical properties of the final product, where no specific conditions are defined to achieve this
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- A61L27/00—Materials for grafts or prostheses or for coating grafts or prostheses
- A61L27/36—Materials for grafts or prostheses or for coating grafts or prostheses containing ingredients of undetermined constitution or reaction products thereof, e.g. transplant tissue, natural bone, extracellular matrix
- A61L27/38—Materials for grafts or prostheses or for coating grafts or prostheses containing ingredients of undetermined constitution or reaction products thereof, e.g. transplant tissue, natural bone, extracellular matrix containing added animal cells
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L27/00—Materials for grafts or prostheses or for coating grafts or prostheses
- A61L27/50—Materials characterised by their function or physical properties, e.g. injectable or lubricating compositions, shape-memory materials, surface modified materials
- A61L27/54—Biologically active materials, e.g. therapeutic substances
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- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L27/00—Materials for grafts or prostheses or for coating grafts or prostheses
- A61L27/50—Materials characterised by their function or physical properties, e.g. injectable or lubricating compositions, shape-memory materials, surface modified materials
- A61L27/58—Materials at least partially resorbable by the body
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- G—PHYSICS
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- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/573—Immunoassay; Biospecific binding assay; Materials therefor for enzymes or isoenzymes
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- A—HUMAN NECESSITIES
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- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L2300/00—Biologically active materials used in bandages, wound dressings, absorbent pads or medical devices
- A61L2300/40—Biologically active materials used in bandages, wound dressings, absorbent pads or medical devices characterised by a specific therapeutic activity or mode of action
- A61L2300/412—Tissue-regenerating or healing or proliferative agents
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- A61L2430/00—Materials or treatment for tissue regeneration
- A61L2430/34—Materials or treatment for tissue regeneration for soft tissue reconstruction
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L2430/00—Materials or treatment for tissue regeneration
- A61L2430/40—Preparation and treatment of biological tissue for implantation, e.g. decellularisation, cross-linking
Description
様々な実施形態では、α−galエピトープは、所望の量のα−galエピトープを除去するのに十分な濃度で十分な時間にわたり、組織製品をアルカラーゼ、ブロメライン、トリプシン、またはディスパーゼなどのプロテアーゼに曝露させることで、移植に先だって組織製品から除去され得る。いくつかの実施形態では、プロテアーゼ酵素は、組織の細胞外マトリックスに対する障害を最小化させながら、α−galを除去するその能力について選択される。いくつかの実施形態では、本明細書の方法を使用して、組織の細胞外マトリックスを損傷することなく、α−galが除去される。
様々な実施形態では、組織製品は、組織をアルカラーゼ、ブロメライン、トリプシン、および/またはディスパーゼなどの1つまたは複数のプロテアーゼ酵素に曝露させることで、α−galエピトープが除去された、α−galエピトープを発現する非霊長類哺乳類からの無処理または脱細胞化組織を含んでなり得る。いくつかの実施形態では、組織は、部分的にまたは完全に脱細胞化され得るが、移植組織製品周囲の組織から天然細胞がその中に移動して増殖し得る、細胞外マトリックスの少なくともいくらかの構成要素を保持し、それによって天然組織の修復、再生、治癒、または治療の速度または全体的なレベルを高める。脱細胞化は、組織をアルカラーゼ、ブロメライン、トリプシン、および/またはディスパーゼなどの1つまたは複数のプロテアーゼ酵素に曝露する前に、それと同時に、および/または後に行い得る。一実施形態では、脱細胞化は、1つまたは複数のプロテアーゼへの曝露後に実施される。
屠殺場からブタ皮膚が収集され、表皮および皮下脂肪を物理的に除去することで、開裂された。残る皮膚組織は、抗生物質溶液を使用して汚染除去された。汚染除去に続いて、組織は無菌条件下で処理された。
α−galの存在は、一連の抗体および比色分析(colormetric)検出試薬を使用して検出され得る。組織は、最初にスクロース溶液中で保存され、次に包埋剤(OCT)中に包埋されて、液体窒素中で冷凍された。組織を含有する凍結ブロックは、次に、クリオスタットミクロトームを使用して薄い(ミクロン)切片に切断されて、顕微鏡スライド上に配置された。組織を含有するスライドは、非特異的結合を妨げる溶液でブロックされ、次に、α−gal残基に特異的に結合するビオチン化抗体である一次抗体と共に、インキュベートされた。第1の抗体が洗い落とされた後、第2の抗体(西洋ワサビペルオキシダーゼ共役ストレプトアビジン)が、スライドに添加された。第2の抗体中のストレプトアビジンは、一次抗体中のビオチンと結合する。インキュベーション時間後、第2の抗体もまた洗い落とされて、その後、検出試薬(DAB)がスライドに添加された。DABは、西洋ワサビペルオキシダーゼの存在下で、スライド上に褐色色素を沈着させた。
α−Gal含有量の定量的評価は、阻害ELISAアッセイを使用して判定された。様々な無細胞皮膚マトリックス(ADM)は、刻まれて、抗α−Gal抗体と共にインキュベートされた。インキュベーション時間後、あらゆる非結合抗体を含有する上清は、α−Gal被覆96ウェルプレートに移された。引き続いてウェルプレートと結合する、上清中に存在する非結合抗体の量は、アルカリホスファターゼ共役二次抗体と、それに続くp−ニトロフェニルリン酸塩検出基質を使用して検出された。
Claims (16)
- ガラクトースα−1,3−ガラクトース部分を含有する、少なくとも1つのコラーゲン含有組織マトリックスを選択するステップと;
前記組織からガラクトースα−1,3−ガラクトース部分を除去するのに十分な条件下で、前記少なくとも1つの組織マトリックスをアルカラーゼまたはブロメラインから選択される少なくとも1つのタンパク質分解酵素と接触させるステップと
を含んでなることを特徴とする、組織製品を調製する方法。 - 請求項1に記載の方法において、前記少なくとも1つのコラーゲン含有組織マトリックスが、細胞組織内に含有されており、方法が、前記細胞組織を脱細胞化するステップをさらに含んでなることを特徴とする方法。
- 請求項1または2に記載の方法において、前記組織マトリックスが、完全に脱細胞化されていることを特徴とする方法。
- 請求項1乃至3の何れか一項に記載の方法において、α−ガラクトースが、実質的にまたは完全に前記コラーゲン含有組織マトリックスから除去されることを特徴とする方法。
- 請求項1乃至4の何れか一項に記載の方法において、前記タンパク質分解酵素が、アルカラーゼであることを特徴とする方法。
- 請求項1乃至5の何れか一項に記載の方法において、前記タンパク質分解酵素が、少なくとも1つの組織マトリックスに損害を与えることなく、ガラクトースα−1,3−ガラクトース部分を除去することを特徴とする方法。
- 請求項1乃至6の何れか一項に記載の方法において、アッセイを実施して、ガラクトースα−1,3−ガラクトース部分が、前記少なくとも1つのコラーゲン含有組織マトリックスから除去されたかどうかを判定するステップをさらに含んでなることを特徴とする方法。
- 請求項1乃至7の何れか一項に記載の方法において、前記タンパク質分解酵素が、アルカラーゼであり、前記アルカラーゼが、約0.5時間から約24時間に及ぶ時間にわたり、約0.0001%から約0.1%に及ぶ濃度で使用されることを特徴とする方法。
- 請求項1乃至7の何れか一項に記載の方法において、前記タンパク質分解酵素が、ブロメラインであり、前記ブロメラインが、約1時間から約24時間に及ぶ時間にわたり、約10単位/リットルから約200単位/リットルに及ぶ濃度で使用されることを特徴とする方法。
- 請求項1乃至8の何れか一項に記載の方法において、前記組織をα−ガラクトシダーゼに接触させるステップをさらに含んでなることを特徴とする方法。
- 請求項1乃至10の何れか一項に記載の方法において、前記少なくとも1つのコラーゲン含有組織マトリックスが、骨、皮膚、脂肪組織真皮、腸管、膀胱、腱、靱帯、筋肉、筋膜、血管、神経性、血管、肝臓、心臓、肺、腎臓、および軟骨組織の少なくとも1つに由来する組織マトリックスを含んでなることを特徴とする方法。
- 請求項1乃至11の何れか一項に記載の方法において、前記少なくとも1つのコラーゲン含有組織マトリックスが、ブタ組織マトリックスであることを特徴とする方法。
- 請求項1に記載の方法において、前記少なくとも1つのコラーゲン含有組織マトリックスが、ブタ無細胞皮膚組織マトリックスからなることを特徴とする方法。
- 請求項1乃至13の何れか一項に記載の方法において、1つまたは複数の生存能力があり組織適合性の細胞を前記組織製品に添加するステップをさらに含んでなることを特徴とする方法。
- 必要とする対象を治療するための医薬品の製造における、請求項1乃至14の何れか一項に記載の方法によって調製された組織製品の使用。
- 組織欠陥を治療するための装置の製造における、請求項1乃至14の何れか一項に記載の方法によって調製された組織製品の使用。
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US201361899647P | 2013-11-04 | 2013-11-04 | |
US61/899,647 | 2013-11-04 | ||
PCT/US2014/063796 WO2015066668A1 (en) | 2013-11-04 | 2014-11-04 | Methods of removing alpha-galactose |
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JP2016536315A JP2016536315A (ja) | 2016-11-24 |
JP2016536315A5 JP2016536315A5 (ja) | 2017-12-28 |
JP6524597B2 true JP6524597B2 (ja) | 2019-06-05 |
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US (2) | US10307510B2 (ja) |
EP (2) | EP3065790B1 (ja) |
JP (1) | JP6524597B2 (ja) |
CN (1) | CN105682697B (ja) |
AU (2) | AU2014341866B2 (ja) |
BR (1) | BR112016009178B1 (ja) |
CA (1) | CA2925332C (ja) |
DK (1) | DK3065790T3 (ja) |
ES (1) | ES2846758T3 (ja) |
WO (1) | WO2015066668A1 (ja) |
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CN106860861B (zh) * | 2017-03-03 | 2020-11-03 | 北京博辉瑞进生物科技有限公司 | 一种免疫增强组合物、制备方法及其应用 |
JP6772390B2 (ja) | 2017-03-06 | 2020-10-21 | ティーイーアイ バイオサイエンシズ,インコーポレイテッド | 穿孔された組織移植片 |
SG11202011723XA (en) * | 2018-07-02 | 2020-12-30 | Gebauer Klopotek Patent Verwaltungs Ug | Stabilization of collagen scaffolds |
WO2021034928A1 (en) * | 2019-08-20 | 2021-02-25 | Lifecell Corporation | Method of manufacturing particulate tissue products |
CN111632196A (zh) * | 2020-06-01 | 2020-09-08 | 内蒙古自治区中医药研究所 | 一种去α-半乳糖基抗原脱细胞基质的制备方法 |
CN114276974A (zh) * | 2021-12-24 | 2022-04-05 | 上海理工大学 | 封装细胞的间质材料及其制备方法和应用 |
CN114377206B (zh) * | 2021-12-24 | 2022-09-30 | 杭州华迈医疗科技有限公司 | 一种脱细胞基质生物材料的制备方法 |
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US20190231931A1 (en) | 2019-08-01 |
CA2925332C (en) | 2022-07-12 |
AU2018236910A1 (en) | 2018-10-18 |
EP3065790A1 (en) | 2016-09-14 |
CN105682697B (zh) | 2022-10-14 |
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AU2014341866B2 (en) | 2018-07-05 |
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CN105682697A (zh) | 2016-06-15 |
BR112016009178A8 (pt) | 2020-03-24 |
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