Screening assays for identifying differentiation-inducing agents and production of differentiated cells for cell therapy
AssigneeAdvanced Cell Technology, Inc.
InventorsMichael D. West, Raymond Page, Hans Scholer, Karen Chapman
Life of this patent
Estimating the term…
Abstract
The invention relates to assays for screening growth factors, adhesion molecules, immunostimulatory molecules, extracellular matrix components and other materials, alone or in combination, simultaneously or temporally, for the ability to induce directed differentiation of pluripotent and multipotent stem cells.
Claims86
A method for evaluating the differentiation of totipotent, nearly totipotent, or pluripotent stem cells, or cells therefrom, in response to one or more chemical or biological agents or physical conditions, comprising:
- (a) separating individual totipotent, nearly totipotent, or pluripotent stem cells, or cells therefrom, or groups of such cells, in culture medium into one or a plurality of separate wells which may be open or closed, which wells may be in the same or different apparatus;
- (b) exposing said separate wells of cells to one or more putative differentiation-inducing conditions simultaneously or sequentially;
- and
- (c) screening said individual cells or groups of cells to detect markers of differentiation of said individual cells or groups of cells.
A method for evaluating the differentiation of a totipotent, nearly totipotent, or pluripotent stem cell, or cells therefrom, or a group of such cells, in response to different compounds or combinations of compounds, comprising:
- (a) separating individual totipotent, nearly totipotent, or pluripotent stem cells, or cells therefrom, or groups of such cells, into one or a plurality of separate vessels which may be open or closed, which vessels may be in the same or different apparatus;
- (b) exposing said separate vessels of cells to a panel of different putative differentiation-inducing compounds or combinations thereof simultaneously or sequentially;
- and
- (c) comparing said individual cells or groups of cells to a reference panel of differentiated or partially differentiated cells in order to evaluate the differentiation of said individual totipotent, nearly totipotent, or pluripotent stem cells, or cells therefrom, or groups of such cells.
A method for evaluating the differentiation of a totipotent, nearly totipotent, or pluripotent stem cell, or cells therefrom, or a group of such cells, in response to different compounds or combinations of compounds, comprising:
- (a) isolating a transfected totipotent, nearly totipotent, or pluripotent stem cell, or cells therefrom, wherein said cell is transfected with at least one reporter gene, the expression of which is operably linked to a promoter that is activated when the cell is induced to differentiate or partially differentiate;
- (b) expanding said transfected cell in culture;
- (c) separating individual transfected cells or individual groups of cells into one or a plurality of separate vessels which may be open or closed, which vessels may be in the same or different apparatus;
- (d) systematically exposing said separate vessels of transfected cells simultaneously or sequentially to a panel of different putative differentiation-inducing compounds or combinations thereof;
- and
- (e) analyzing said individual transfected cells or groups of cells in order to detect expression of said at least one reporter gene.
A method for evaluating the differentiation of a totipotent, nearly totipotent, or pluripotent stem cells, or cells therefrom, or a group of such cells in response to one or more compounds, comprising:
- (a) obtaining a library of transfected totipotent, nearly totipotent, or pluripotent stem cells, or cells therefrom, each transfected with at least one reporter gene, the expression of which is linked to a promoter that is activated when the cell is induced to differentiate or partially differentiate;
- (b) separating individual members of said library into one or a plurality of separate vessels which may be open or closed, which vessels may be in the same or different apparatus;
- (c) exposing said separate vessels of transfected cells simultaneously or sequentially to the same one or more putative differentiation-inducing compounds;
- and
- (d) analyzing said individual members of said library in order to detect expression of said at least one reporter gene.
A library of two or more gene trap stem cell lines used simultaneously together to screen to detect agents or conditions that affect differentiation, survival, or proliferation of the stem cells.
A method for inducing differentiation of a stem cell to form cells of mesodermal lineage, comprising exposing the stem cells to Flt-3.
A method for inducing differentiation of a stem cell to form cells of mesodermal and neural lineage, comprising exposing the stem cells to TGF-beta-1.
A method for inducing differentiation of a stem cell to form cells selected from the group consisting of cells of endothelial lineage, and cells of endodermal lineage or appearance, comprising exposing the stem cells to tenascin.
A method for inducing differentiation of a stem cell comprising exposing the stem cells to Tie-1.
A method for inducing differentiation of a stem cell to form fibroblasts and/or other cells of connective tissue comprising exposing the stem cells to BMP-2.
A method for inducing differentiation of a stem cell to form myocardial cells comprising exposing the stem cells to endothelial inducer cells.
A method for inducing differentiation of a stem cell to form cells of mesodermal lineage comprising exposing the stem cells to fibroblast inducer cells.
The method of claim 84, further comprising exposing the stem cells to FGF-4 and TGF-beta-1.
Dependencies are inferred from the claim text.