公开(公告)号：US20160002595A1

公开(公告)日：2016-01-07

申请号：US14768409

申请日：2014-02-18

Applicant: UNIVERSITY HEALTH NETWORK ,  THE HOSPITAL FOR SICK CHILDREN

Inventor： Gordon KELLER ,  Shinichiro OGAWA ,  Anand GHANEKAR ,  Christine BEAR ,  Binita M. KAMATH ,  Mina OGAWA ,  James SURAPISITCHAT

IPC: C12N5/071

CPC classification number: C12N5/067 ,  A61K35/407 ,  C12N5/0679 ,  C12N2501/01 ,  C12N2501/115 ,  C12N2501/119 ,  C12N2501/12 ,  C12N2501/155 ,  C12N2501/16 ,  C12N2501/237 ,  C12N2501/415 ,  C12N2501/42 ,  C12N2501/724 ,  C12N2501/727 ,  C12N2501/999 ,  C12N2502/00 ,  C12N2502/14 ,  C12N2503/02 ,  C12N2506/02 ,  C12N2506/45

Abstract: Methods for producing hepatocyte and/or cholangiocyte lineage cells from pluripotent stem cells, the method comprising (a) specifying the extended nodal agonist treated induced endodermal cell population to obtain a cell population comprising hepatocyte and/or cholangiocyte progenitors by contacting the extended nodal agonist treated induced endodermal cell population with specification media comprising a FGF agonist and a BMP4 agonist and/or active conjugates and/or fragments thereof; (b) inducing maturation, and optionally further lineage specification and/or expansion of the hepatocyte and/or cholangiocyte progenitors of the cell population to obtain a population comprising hepatocyte lineage cells such as hepatoblasts, hepatocytes and/or cholangiocytes, the inducing maturation step comprising generating aggregates of the cell population. Optionally, the method also comprises activating the cAMP pathway within the aggregates and forming co-aggregates.

Abstract translation: 用于从多能干细胞产生肝细胞和/或胆管细胞谱系细胞的方法，所述方法包括（a）指定延长的淋巴结激动剂治疗的诱导的内胚层细胞群，以通过使经延长的淋巴结激动剂治疗以获得包含肝细胞和/或胆管细胞祖细胞的细胞群 具有包含FGF激动剂和BMP4激动剂和/或其活性缀合物和/或其片段的规范培养基的诱导内胚层细胞群; （b）诱导成熟，以及任选地进一步谱系说明和/或扩增细胞群体的肝细胞和/或胆管细胞祖细胞以获得包含肝细胞谱系细胞如肝母细胞，肝细胞和/或胆管细胞的群体，诱导成熟步骤包括 产生细胞群体的聚集体。 任选地，该方法还包括激活聚集体内的cAMP途径并形成共聚集体。

公开(公告)号：US20160038544A1

公开(公告)日：2016-02-11

申请号：US14782070

申请日：2014-04-02

Applicant: UNIVERSITY HEALTH NETWORK

Inventor： Gordon KELLER ,  April M. CRAFT

IPC: A61K35/32 ,  C12N5/077 ,  G01N33/50

CPC classification number: A61K35/32 ,  C12N5/0655 ,  C12N2501/115 ,  C12N2501/155 ,  C12N2501/16 ,  C12N2501/415 ,  C12N2501/999 ,  C12N2503/02 ,  C12N2506/02 ,  C12N2506/03 ,  G01N33/5044

Abstract: A method for generating chondrocytes and/or cartilage, optionally articular like non-hypertrophic chondrocyte cells and/or cartilage like tissue and/or hypertrophic chondrocyte like cells and/or cartilage like tissue, the method comprising: a. culturing a primitive streak-like mesoderm population, optionally a CD56+, PDGFRalpha+ KDR-primitive streak-like mesoderm population, with a paraxial mesoderm specifying cocktail comprising: i. a FGF agonist; ii. a BMP inhibitor; optionally Noggin, LDN-193189, Dorsomorphin; and iii. optionally one or more of a TGFbeta inhibitor, optionally SB431524; and a Wnt inhibitor, optionally DKK1, IWP2, or XAV939; to specify a paraxial mesoderm population expressing cell surface CD73, CD105 and/or PDGFR-beta; b. generating a chondrocyte precursor population comprising: i. culturing the paraxial mesoderm population expressing CD73, CD105 and/or PDGFR-beta at a high cell density optionally in serum free or serum containing media; ii. culturing the high cell density CD73+, CD105+ and/or PDGFRbeta+ paraxial mesoderm population with a TGFbeta3 agonist in serum free media to produce a high cell density Sox9+, collagen 2+ chondrocyte precursor population; and c. either i. culturing the high cell density Sox9+, collagen 2+ chondrocyte precursor population with the TGFbeta3 agonist for an extended period of time to produce an articular like non-hypertrophic chondrocyte cells and/or cartilage like tissue; or ii. culturing the high cell density Sox9+ collagen2+ chondrocyte precursor population with a BMP4 agonist for an extended period of time to produce a hypertrophic chondrocyte like cells and/or cartilage like tissue.

Abstract translation: 一种用于产生软骨细胞和/或软骨的方法，任选地关节像非增生性软骨细胞和/或软骨样组织和/或肥大软骨细胞样细胞和/或软骨样组织，所述方法包括：a。 培养原始条纹样中胚层群体，任选地是CD56 +，PDGFRalpha + KDR-原始条纹状中胚层群体，其中傍轴中胚层指定混合物，包括：i。 FGF激动剂; ii。 BMP抑制剂; 任选地，Noggin，LDN-193189，Dorsomorphin; 和iii。 任选地一种或多种TGFβ抑制剂，任选的SB431524; 和Wnt抑制剂，任选地DKK1，IWP2或XAV939; 指定表达细胞表面CD73，CD105和/或PDGFR-β的近轴中胚层群; b。 产生软骨细胞前体群体，包括：i。 以高细胞密度任选地在无血清或含血清培养基中培养表达CD73，CD105和/或PDGFR-β的近轴中胚层群; ii。 在血清游离培养基中培养具有TGFbeta3激动剂的高细胞密度CD73 +，CD105 +和/或PDGFRβ+近轴中胚层群体以产生高细胞密度Sox9 +，胶原蛋白+2软骨细胞前体种群; 和c。 我是 用TGFbeta3激动剂长时间培养高细胞密度Sox9 +，胶原蛋白+软骨细胞前体种群，以产生关节性非肥大软骨细胞和/或软骨样组织; 或ii。 用BMP4激动剂长时间培养高细胞密度的Sox9 +胶原蛋白2 +软骨细胞前体群体，以产生肥大的软骨细胞样细胞和/或软骨样组织。

公开(公告)号：US20190111086A1

公开(公告)日：2019-04-18

申请号：US15977536

申请日：2018-05-11

Applicant: UNIVERSITY HEALTH NETWORK

Inventor： Gordon KELLER ,  April M. CRAFT

IPC: A61K35/32 ,  C12N5/077 ,  G01N33/50

Abstract: A method for generating chondrocytes and/or cartilage, optionally articular like non-hypertrophic chondrocyte cells and/or cartilage like tissue and/or hypertrophic chondrocyte like cells and/or cartilage like tissue, the method comprising: a. culturing a primitive streak-like mesoderm population, optionally a CD56+, PDGFRalpha+ KDR− primitive streak-like mesoderm population, with a paraxial mesoderm specifying cocktail comprising: i. a FGF agonist; ii. a BMP inhibitor; optionally Noggin, LDN-193189, Dorsomorphin; and iii. optionally one or more of a TGFbeta inhibitor, optionally SB431524; and a Wnt inhibitor, optionally DKK1, IWP2, or XAV939; to specify a paraxial mesoderm population expressing cell surface CD73, CD105 and/or PDGFR-beta; b. generating a chondrocyte precursor population comprising: i. culturing the paraxial mesoderm population expressing CD73, CD105 and/or PDGFR-beta at a high cell density optionally in serum free or serum containing media; ii. culturing the high cell density CD73+, CD105+ and/or PDGFRbeta+ paraxial mesoderm population with a TGFbeta3 agonist in serum free media to produce a high cell density Sox9+, collagen 2+ chondrocyte precursor population; and c. either i. culturing the high cell density Sox9+, collagen 2+ chondrocyte precursor population with the TGFbeta3 agonist for an extended period of time to produce an articular like non-hypertrophic chondrocyte cells and/or cartilage like tissue; or ii. culturing the high cell density Sox9+ collagen2+ chondrocyte precursor population with a BMP4 agonist for an extended period of time to produce a hypertrophic chondrocyte like cells and/or cartilage like tissue.

公开(公告)号：US20180028572A1

公开(公告)日：2018-02-01

申请号：US15550572

申请日：2016-02-17

Applicant: UNIVERSITY HEALTH NETWORK

Inventor： Gordon KELLER ,  Stephanie PROTZE

IPC: A61K35/34 ,  C12N5/10 ,  C12N5/077

CPC classification number: A61K35/34 ,  C12N5/0657 ,  C12N5/10 ,  C12N2501/115 ,  C12N2501/15 ,  C12N2501/155 ,  C12N2501/16 ,  C12N2501/165 ,  C12N2501/385 ,  C12N2501/727 ,  C12N2501/999 ,  C12N2506/02 ,  C12N2506/45

Abstract: Provided are methods for producing compositions comprising a population of cardiomyocytes enriched for or substantially devoid of sinoatrial node-like pacemaker cardiomyocytes (SANLCM) from human pluripotent stem cells (hPSCs), and methods of use thereof.

公开(公告)号：US20230330152A1

公开(公告)日：2023-10-19

申请号：US18336410

申请日：2023-06-16

Applicant: UNIVERSITY HEALTH NETWORK

Inventor： Gordon KELLER ,  April M. CRAFT

IPC: A61K35/32 ,  C12N5/077 ,  G01N33/50

CPC classification number: A61K35/32 ,  C12N5/0655 ,  G01N33/5044 ,  C12N2501/115 ,  C12N2501/155 ,  C12N2501/16 ,  C12N2501/415 ,  C12N2501/999 ,  C12N2503/02 ,  C12N2506/02 ,  C12N2506/03

Abstract: A method for generating chondrocytes and/or cartilage, optionally articular like non-hypertrophic chondrocyte cells and/or cartilage like tissue and/or hypertrophic chondrocyte like cells and/or cartilage like tissue, the method comprising:

a. culturing a primitive streak-like mesoderm population, optionally a CD56+, PDGFRalpha+ KDR- primitive streak-like mesoderm population, with a paraxial mesoderm specifying cocktail comprising:

i. a FGF agonist;
ii. a BMP inhibitor; optionally Noggin, LDN-193189, Dorsomorphin; and
iii. optionally one or more of a TGFbeta inhibitor, optionally SB431524; and a Wnt inhibitor, optionally DKK1, IWP2, or XAV939;

to specify a paraxial mesoderm population expressing cell surface CD73, CD105 and/or PDGFR-beta;
b. generating a chondrocyte precursor population comprising:

i. culturing the paraxial mesoderm population expressing CD73, CD105 and/or PDGFR-beta at a high cell density optionally in serum free or serum containing media;
ii. culturing the high cell density CD73+, CD105+ and/or PDGFRbeta+ paraxial mesoderm population with a TGFbeta3 agonist in serum free media to produce a high cell density Sox9+, collagen 2+ chondrocyte precursor population; and


c. either

i. culturing the high cell density Sox9+, collagen 2+ chondrocyte precursor population with the TGFbeta3 agonist for an extended period of time to produce an articular like non-hypertrophic chondrocyte cells and/or cartilage like tissue; or
ii. culturing the high cell density Sox9+ collagen2+ chondrocyte precursor population with a BMP4 agonist for an extended period of time to produce a hypertrophic chondrocyte like cells and/or cartilage like tissue.

公开(公告)号：US20200157494A1

公开(公告)日：2020-05-21

申请号：US16109202

申请日：2018-08-22

Applicant: UNIVERSITY HEALTH NETWORK ,  THE HOSPITAL FOR SICK CHILDREN

Inventor： Gordon KELLER ,  Shinichiro OGAWA ,  Anand GHANEKAR ,  Christine BEAR ,  Binita M. KAMATH ,  Mina OGAWA ,  James SURAPISITCHAT

IPC: C12N5/071

Abstract: Methods for producing hepatocyte and/or cholangiocyte lineage cells from pluripotent stem cells, the method comprising (a) specifying the extended nodal agonist treated induced endodermal cell population to obtain a cell population comprising hepatocyte and/or cholangiocyte progenitors by contacting the extended nodal agonist treated induced endodermal cell population with specification media comprising a FGF agonist and a BMP4 agonist and/or active conjugates and/or fragments thereof; (b) inducing maturation, and optionally further lineage specification and/or expansion of the hepatocyte and/or cholangiocyte progenitors of the cell population to obtain a population comprising hepatocyte lineage cells such as hepatoblasts, hepatocytes and/or cholangiocytes, the inducing maturation step comprising generating aggregates of the cell population. Optionally, the method also comprises activating the cAMP pathway within the aggregates and forming co-aggregates.

公开(公告)号：US20160215263A1

公开(公告)日：2016-07-28

申请号：US14915992

申请日：2014-09-12

Applicant: UNIVERSITY HEALTH NETWORK

Inventor： Gordon KELLER ,  Alec Drake WITTY ,  Steven James KATTMAN

IPC: C12N5/077 ,  G01N33/50

Abstract: Provided are methods and products for obtaining cardiovascular lineage cells from hPSCs. The method for obtaining a cardiomyocyte lineage or an epicardial lineage cell population from human pluripotent stem cells (hPSCs) comprises one or more of the following steps: (a) contacting BMP component primed hPSCs with a cardiovascular mesoderm programming cocktail suitable for inducing the hPSCs to differentiate to a cardiovascular mesoderm cell population under conditions suitable for the programming cocktail to penetrate the hPSCs and culturing the contacted hPSCs for a period of time to generate a KDR+ and PDGFRalpha+ cardiovascular mesoderm cell population; (b) contacting the cardiovascular mesoderm cell population with a cardiovascular progenitor specification cocktail suitable to specify a NKX2-5+ or WT1+ cardiovascular progenitor cell population under conditions suitable for the specification cocktail to penetrate the cardiovascular mesoderm cell population and culturing the contacted cardiovascular mesoderm cell population for a period of time to generate a NKX2-5+ or WT1+ cardiovascular progenitor cell population; and (d) contacting the cardiovascular progenitor cell population with a maturation cocktail under conditions suitable for the maturation cocktail to penetrate the cardiovascular progenitor cell population and culturing the contacted cardiovascular progenitor population for a period of time to produce a cardiovascular population optionally cardiomyocyte lineage cells expressing cardiac troponin T (cTnT) and/or SIRPA and/or epicardial lineage cells expressing WT1.

Abstract translation: 提供从hPSC获得心血管谱系细胞的方法和产品。 用于从人多能干细胞（hPSC）获得心肌细胞谱系或心外膜谱系细胞群的方法包括一个或多个以下步骤：（a）使BMP组分引发的hPSC与适合诱导hPSC的心血管中胚层编程混合物接触 在适合编程混合物的条件下分化为心血管中胚层细胞群体以渗透hPSC并培养接触的hPSC一段时间以产生KDR +和PDGFRα+心血管中胚层细胞群; （b）使心血管中胚层细胞群体与适合于规范混合液以渗透心血管中胚层细胞群体并培养接触的心血管中胚层细胞的条件下，适用于指定NKX2-5 +或WT1 +心血管祖细胞群的心血管祖细胞规格混合物接触 人口一段时间以产生NKX2-5 +或WT1 +心血管祖细胞群体; 和（d）在适合于成熟混合物的条件下将心血管祖细胞群与成熟混合物接触以渗透心血管祖细胞群体并培养接触的心血管祖细胞群体一段时间以产生心血管群体，任选地表达心肌细胞谱系细胞 心肌肌钙蛋白T（cTnT）和/或表达WT1的SIRPA和/或心外膜谱系细胞。