Artery- and vein-specific proteins and uses therefor
AssigneeCalifornia Institute of Technology
InventorsHai U. Wang, Zhoufeng Chen, David J. Anderson
Life of this patent
Estimating the term…
Abstract
Arterial and venous endothelial cells are molecularly distinct from the earliest stages of angiogenesis. This distinction is revealed by expression on arterial cells of a transmembrane ligand, called EphrinB2 whose receptor EphB4 is expressed on venous cells. Targeted disruption of the EphrinB2 gene prevents the remodeling of veins from a capillary plexus into properly branched structures. Moreover, it also disrupts the remodeling of arteries, suggesting that reciprocal interactions between pre-specified arterial and venous endothelial cells are necessary for angiogenesis.
Claims108
A method for altering angiogenesis in a mammal, comprising administering to the mammal, in a therapeutically effective quantity, a drug which alters binding or interaction of an artery-specific cell surface molecule with a vein-specific cell surface molecule in a therapeutically effective quantity.
A method for selectively delivering a drug to arteries in a mammal, comprising administering to the mammal a complex comprising:
- a) the drug and
- b) a component which binds an artery-specific cell surface molecule, under conditions appropriate for the component of (b) to bind the artery-specific cell surface molecule, whereby the drug is delivered to arteries.
A method for selectively delivering a drug to veins in a mammal, comprising administering to the mammal a complex comprising:
- a) the drug, and
- b) a component which binds a vein-specific cell surface molecule, under conditions appropriate for the component of (b) to bind the vein-specific cell surface molecule, whereby the drug is delivered to veins.
A transgenic mouse having an indicator gene which is detectably expressed in cells of arteries but not cells of veins.
A transgenic mouse of genotype EphrinB2+/−.
A transgenic mouse in which EphrinB2 genes comprise an insertion that marks all arteries but not veins.
A transgenic mouse of genotype EphrinB2taulacZ/+.
A method for identifying artery cells in a mouse having an indicator gene inserted in one or more alleles of EphrinB2, comprising staining a section of the mouse with a substance appropriate for detection of expression of the indicator gene.
A transgenic mouse having an indicator gene which is expressed in venous endothelial cells but not in arterial endothelial cells.
A transgenic mouse of genotype EphB4+/−.
A method for identifying vein cells in a mouse having an indicator gene inserted in one or more alleles of EphB4, comprising staining a section of the mouse with a substance appropriate for detection of expression of the indicator gene.
A method for testing an effect of a drug on growth of arteries, comprising administering the drug to a mouse having an indicator gene inserted in a gene specifically expressed in arteries, observing the effect of the drug, and comparing the effect to that produced in a suitable control mouse.
A method for testing an effect of a drug on growth of veins, comprising administering the drug to a mouse having an indicator gene inserted in a gene specifically expressed in veins, observing the effect of the drug, and comparing the effect to that produced in a suitable control mouse.
A method for identifying arterial endothelial cells in a tissue sample, comprising contacting the tissue sample with a molecule which binds to EphrinB2, wherein said molecule is linked to a label, and detecting the label, wherein if label is detected on a cell, the cell is an arterial endothelial cell.
A method for identifying venous endothelial cells in a tissue sample, comprising contacting the tissue sample with a molecule which binds to EphB4, wherein said molecule is linked to a label, and detecting the label, wherein if label is detected on a cell, the cell is a venous endothelial cell.
A method for directing a substance to arteries in a mammal, comprising administering to the mammal a complex which comprises the substance linked to a moiety which binds EphrinB2.
A method for directing a substance to veins in a mammal, comprising administering to the mammal a complex which comprises the substance linked to a moiety which binds EphB4.
A method for altering development of blood vessels in a mammal, comprising administering to the mammal a soluble polypeptide comprising the extracellular domain of an artery-specific cell surface protein or a soluble polypeptide comprising the extracellular domain of a vein-specific cell surface protein.
A method for identifying a drug that inhibits interaction of an arterial endothelial cell-specific surface molecule with a venous endothelial cell-specific surface molecule, comprising:
- a) combining:
- 1) the arterial endothelial cell-specific surface molecule;
- 2) the venous endothelial cell-specific surface molecule;
- and
- 3) a drug to be assessed for its ability to inhibit interaction between the molecule of (1) and the molecule of (2), under conditions appropriate for interaction between the molecule of (1) and the molecule of (2);
- b) determining the extent to which the molecule of (1) and the molecule of (2) interact;
- and
- c) comparing the extent determined in (b) with the extent to which interaction of the molecule of (1) and the molecule of (2) occurs in the absence of the drug to be assessed and under the same conditions appropriate for interaction of the molecule of (1) with the molecule of (2);
- wherein if the extent to which interaction of the molecule of (1) and the molecule of (2) is less in the presence of the drug than in the absence of the drug, the drug is one which inhibits interaction of the arterial endothelial cell-specific molecule of (1) with the venous endothelial cell-specific molecule of (2).
A method for identifying a drug that enhances interaction of an arterial endothelial cell-specific surface molecule with a venous endothelial cell-specific surface molecule, comprising:
- a) combining:
- 1) the arterial endothelial cell-specific surface molecule;
- 2) the venous endothelial cell-specific surface molecule;
- and
- 3) a drug to be assessed for its ability to inhibit interaction between the molecule of (1) and the molecule of (2), under conditions appropriate for interaction between the molecule of (1) and the molecule of (2);
- b) determining the extent to which the molecule of (1) and the molecule of (2) interact;
- and
- c) comparing the extent determined in (b) with the extent to which interaction of the molecule of (1) and the molecule of (2) occurs in the absence of the drug to be assessed and under the same conditions appropriate for interaction of the molecule of (1) with the molecule of (2);
- wherein if the extent to which interaction of the molecule of (1) and the molecule of (2) is greater in the presence of the drug than in the absence of the drug, the drug is one which enhances interaction of the arterial endothelial cell-specific molecule of (1) with the venous endothelial cell-specific molecule of (2).
A method for isolating arterial endothelial cells, comprising dissociating cells of a tissue sample comprising arterial endothelial cells, contacting the dissociated cells with a substance which binds to a cell-surface protein expressed specifically on arterial endothelial cells, and separating the cells which have bound the substance from the cells which have not bound the substance.
A method for isolating arterial endothelial cells, comprising dissociating cells of a tissue sample comprising arterial endothelial cells, contacting the cells with a substance which binds to a cell-surface protein expressed specifically on arterial endothelial cells, wherein said substance is bound to a solid support, removing cells which do not bind to the substance, and releasing the arterial endothelial cells from the solid support.
A method for isolating venous endothelial cells, comprising dissociating cells of a tissue sample comprising venous endothelial cells, contacting the dissociated cells with a substance which binds to a cell-surface protein expressed specifically on venous endothelial cells, and separating the cells which have bound the substance from cells which have not bound the substance.
A method for isolating venous endothelial cells, comprising dissociating cells of a tissue sample comprising venous endothelial cells, contacting the cells of the tissue sample with a substance which binds to a cell-surface protein expressed specifically on venous endothelial cells, wherein said substance is bound to a solid support, removing cells which do not bind to the substance, and releasing the venous endothelial cells from the solid support.
Isolated arterial endothelial cells.
Isolated venous endothelial cells.
A method for assessing an effect of one or more drugs on arteries, comprising adding one or more drugs to isolated arterial endothelial cells, and observing the cells for the effect.
A method for assessing an effect of one or more drugs on veins, comprising adding one or more drugs to isolated venous endothelial cells, and observing the cells for the effect.
A cell line derived from arterial endothelial cells.
A cell line derived from venous endothelial cells.
A cell line which produces a protein that is detectably produced in arteries, but is not detectably produced in veins.
A cell line which produces a protein that is detectably produced in veins, but is not detectably produced in arteries.
A cDNA library produced from isolated arterial endothelial cells.
A cDNA library produced from isolated venous endothelial cells.
A method for identifying a gene which shows differential expression in venous endothelial cells compared to arterial endothelial cells, comprising producing a transgenic mouse having an indicator insertion gene in a gene to be tested for differential expression, and observing expression of the indicator insertion gene, wherein a difference in expression of the indicator insertion gene in venous endothelial cells and arterial endothelial cells indicates a gene which shows differential expression.
A method for modifying arteries in a mammal, comprising genetically altering isolated arterial endothelial cells and introducing the altered cells into the mammal.
A method for modifying veins in a mammal, comprising genetically altering isolated veins endothelial cells and introducing the altered cells into the mammal.
A method for identifying differentially expressed genes comprising;
- a) dissociating cells of a tissue sample comprising arterial endothelial cells;
- b) contacting the dissociated cells with a first substance which binds to a cell-surface protein expressed specifically on arterial endothelial cells;
- c) separating the arterial endothelial cells which have bound said first substance from the cells which have not bound said first substance where the bound cells are arterial endothelial cells;
- d) dissociating cells of a tissue sample comprising venous endothelial cells;
- e) contacting the dissociated cells with a second substance which binds to a cell-surface protein expressed specifically on venous endothelial cells;
- f) separating the cells which have bound said second substance from cells which have not bound said second substance wherein cells which have bound are venous endothelial cells;
- g) subjecting the separated arterial endothelial cells from step (c) and the venous endothelial cells from step (f) to conditions for differential expression;
- and
- h) identifying differentially expressed genes.
A method for identifying differentially expressed genes comprising:
- a) dissociating cells of a tissue sample comprising arterial endothelial cells;
- b) contacting the cells with a first substance which binds to a cell-surface protein expressed specifically on arterial endothelial cells, wherein said first substance is bound to a first solid support;
- c) removing cells which do not bind to said first substance;
- d) releasing the arterial endothelial cells from said first solid support;
- e) dissociating cells of a tissue sample comprising venous endothelial cells;
- f) contacting the cells of the tissue sample with a second substance which binds to a cell-surface protein expressed specifically on venous endothelial cells, wherein said second substance is bound to a second solid support;
- g) removing cells which do not bind to said second substance;
- h) releasing the venous endothelial cells from said second solid support;
- i) subjecting the released arterial endothelial cells and the venous endothelial cells in steps (d) and (h) to conditions for differential expression;
- and
- j) identifying differentially expressed genes.
A method for identifying differentially expressed genes comprising;
- a) dissociating cells of a tissue sample comprising arterial endothelial cells;
- b) contacting the dissociated cells with a first substance which binds to an Ephrin family ligand expressed specifically on arterial endothelial cells;
- c) separating the arterial endothelial cells which have bound said first substance from the cells which have not bound said first substance where the bound cells are arterial endothelial cells;
- d) dissociating cells of a tissue sample comprising venous endothelial cells;
- e) contacting the dissociated cells with a second substance which binds to an Eph family receptor expressed specifically on venous endothelial cells
- f) separating the cells which have bound said second substance from cells which have not bound said second substance wherein cells which have bound are venous endothelial cells;
- g) subjecting the separated arterial endothelial cells from step (c) and the venous endothelial cells from step (f) to conditions for differential expression;
- and
- h) identifying differentially expressed genes.
A method for identifying differentially expressed genes comprising;
- a) dissociating cells of a tissue sample comprising arterial endothelial cells;
- b) contacting the dissociated cells with a first substance which binds to EphrinB2 expressed on arterial endothelial cells;
- c) separating the arterial endothelial cells which have bound said first substance from the cells which have not bound said first substance where the bound cells are arterial endothelial cells;
- d) dissociating cells of a tissue sample comprising venous endothelial cells;
- e) contacting the dissociated cells with a second substance which binds to EphB4 expressed on venous endothelial cells;
- f) separating the cells which have bound said second substance from cells which have not bound said second substance wherein cells which have bound are venous endothelial cells;
- g) subjecting the separated arterial endothelial cells from step (c) and the venous endothelial cells from step (f) to conditions for differential expression;
- and
- h) identifying differentially expressed genes.
Dependencies are inferred from the claim text.