Compositions and methods for enhancing phagocytosis or phagocyte activity
The present invention provides a system for enhancing clearance or destruction of undesirable cells or noncellular molecular entities by tagging such cells or noncellular molecular entities with a marker that targets the cells or noncellular molecular entities for phagocytosis (phagocytic marker). The target cells can be, for example, endothelial cells, tumor cells, leukocytes, or virus-infected cells. In certain embodiments of the invention the tagging is accomplished by administering a composition comprising an antibody or ligand linked to the phagcytotic marker, wherein the antibody or ligand binds to a cell type specific marker present on or in the cell surface of a target cell. In preferred embodiments of the invention, the phagocytic marker comprises phosphatidylserine or a group derived from phosphatidylserine, thrombospondin-1, annexin I, or a derivative of any of these.
1. A method of selecting a compound that enhances phagocytosis of target cells, comprising steps of:
contacting a population of target cells with a candidate compound not found in nature, the candidate compound comprising (i) a cell-binding moiety and (ii) a phagocytic marker, wherein the cell-binding moiety and phagocytic marker are linked;
contacting the target cells with a population of phagocytic cells;
comparing the degree of phagocytosis of the target cells by the phagocytic cells with the degree of phagocytosis of a comparable population of target cells that were not exposed to the candidate compound but were also exposed to phagocytic cells under similar conditions; and
selecting the candidate compound as a compound that enhances phagocytosis of target cells if the degree of phagocytosis of target cells that were contacted with the candidate compound is greater than the degree of phagocytosis of target cells that were not contacted with the candidate compound, wherein the cell-binding moiety is an antibody and the phagocytic marker is linked to the Fc domain of the antibody.
2. The method of claim 1 , wherein the phagocytic marker comprises a phosphatidylserine head group.
3. The method of claim 1 , wherein the population of phagocytic cells comprises macrophages.
4. The method of claim 1 , wherein the cell-binding moiety binds to a tumor marker.
5. The method of claim 1 , wherein the cell-binding moiety binds to an endothelial cell marker.
6. The method of claim 1 , wherein the target cells comprise endothelial cells.
7. The method of claim 1 , wherein the target cells comprise tumor cells.
8. The method of claim 1 , wherein the target cells comprise cells infected with a pathogen.
9. The method of claim 1 , wherein the target cells comprise eukaryotic cells.
10. The method of claim 1 , wherein the phagocytic marker comprises a moiety that is naturally displayed by apoptotic or pre-apoptotic cells and enhances recognition of apoptotic or pre-apoptotic cells by phagocytic cells.
11. The method of claim 1 , wherein the phagocytic marker and the cell-binding moiety are covalently linked.
12. The method of claim 1 , wherein the phagocytic marker and the cell-binding moiety are linked via a linking moiety.
13. The method of claim 1 , wherein the phagocytic marker and the cell-binding moiety are covalently linked via a bifunctional crosslinking reagent.
14. The method of claim 1 , wherein the phagocytic marker and the cell-binding moiety are linked to first and second linking moieties, and the first and second linking moieties are linked to each other.
15. The method of claim 14 , wherein the first and second linking moieties are linked by a biotin-avidin or biotin-streptavidin interaction.
16. The method of claim 1 , wherein the cell-binding moiety binds to an integrin or to a Tie receptor.
17. The method of claim 1 , wherein the cell-binding moiety binds to integrin alpha(v)beta(3).
18. The method of claim 1 , wherein the compound comprises a lipid-containing vesicle comprising lipids, a cell-binding moiety, and a phagocytic marker.
19. The method of claim 1 , further comprising comparing the ability of a compound comprising a naturally occurring phagocytic marker to enhance phagocytosis with the ability of a compound comprising a variant or fragment of a naturally occurring phagocytic marker to enhance phagocytosis.
20. The method of claim 1 , further comprising comparing the degree of phagocytosis of the target cells by the phagocytic cells with the degree of phagocytosis of a population of target cells that have been induced to undergo apoptosis and exposed to phagocytic cells under similar conditions.
21. The method of claim 1 , further comprising administering the compound to an animal model of disease.
22. A method of selecting a compound that enhances phagocytosis of target cells, comprising steps of:
contacting a population of target cells with a candidate compound not found in nature, the candidate compound comprising (i) a cell-binding moiety and (ii) a phagocytic marker, wherein the cell-binding moiety and phagocytic marker are linked;
contacting the target cells with a population of phagocytic cells;
comparing the degree of phagocytosis of the target cells by the phagocytic cells with the degree of phagocytosis of a comparable population of target cells that were not exposed to the candidate compound but were also exposed to phagocytic cells under similar conditions; and
selecting the candidate compound as a compound that enhances phagocytosis of target cells if the degree of phagocytosis of target cells that were contacted with the candidate compound is greater than the degree of phagocytosis of target cells that were not contacted with the candidate compound, wherein the compound is a fusion protein comprising a cell-binding moiety and a phagocytic marker, wherein if the cell-binding moiety comprises an antibody, the antibody is a single-chain antibody.
23. The method of claim 22 , wherein the compound is a fusion protein comprising a cell-binding moiety and a phagocytic marker, wherein the cell-binding moiety is a single-chain antibody.
24. The method of claim 22 , wherein the phagocytic marker comprises a phosphatidylserine head group.
25. The method of claim 22 , wherein the population of phagocytic cells comprises macrophages.
26. The method of claim 22 , wherein the cell-binding moiety binds to a tumor marker.
27. The method of claim 22 , wherein the cell-binding moiety binds to an endothelial cell marker.
28. The method of claim 22 , wherein the target cells comprise endothelial cells.
29. The method of claim 22 , wherein the target cells comprise tumor cells.
30. The method of claim 22 , wherein the target cells comprise cells infected with a pathogen.
31. The method of claim 22 , wherein the target cells comprise eukaryotic cells.
32. The method of claim 22 , wherein the phagocytic marker comprises a moiety that is naturally displayed by apoptotic or pre-apoptotic cells and enhances recognition of apoptotic or pre-apoptotic cells by phagocytic cells.
33. The method of claim 22 , wherein the phagocytic marker and the cell-binding moiety are covalently linked.
34. The method of claim 22 , wherein the phagocytic marker and the cell-binding moiety are linked via a linking moiety.
35. The method of claim 22 , wherein the phagocytic marker and the cell-binding moiety are covalently linked via a bifunctional crosslinking reagent.
36. The method of claim 22 , wherein the phagocytic marker and the cell-binding moiety are linked to first and second linking moieties, and the first and second linking moieties are linked to each other.
37. The method of claim 36 , wherein the first and second linking moieties are linked by a biotin-avidin or biotin-streptavidin interaction.
38. The method of claim 22 , wherein the cell-binding moiety binds to an integrin or to a Tie receptor.
39. The method of claim 22 , wherein the cell-binding moiety binds to integrin alpha(v)beta(3).
40. The method of claim 22 , wherein the compound comprises a lipid-containing vesicle comprising lipids, a cell-binding moiety, and a phagocytic marker.
41. The method of claim 22 , further comprising comparing the ability of a compound comprising a naturally occurring phagocytic marker to enhance phagocytosis with the ability of a compound comprising a variant or fragment of a naturally occurring phagocytic marker to enhance phagocytosis.
42. The method of claim 22 , further comprising comparing the degree of phagocytosis of the target cells by the phagocytic cells with the degree of phagocytosis of a population of target cells that have been induced to undergo apoptosis and exposed to phagocytic cells under similar conditions.
43. The method of claim 22 , further comprising administering the compound to an animal model of disease.
44. A method of selecting a compound that enhances phagocytosis of target cells, comprising steps of:
contacting a population of target cells with a candidate compound not found in nature, the candidate compound comprising (i) a cell-binding moiety and (ii) a phagocytic marker, wherein the cell-binding moiety and phagocytic marker are linked;
contacting the target cells with a population of phagocytic cells;
comparing the degree of phagocytosis of the target cells by the phagocytic cells with the degree of phagocytosis of a comparable population of target cells that were not exposed to the candidate compound but were also exposed to phagocytic cells under similar conditions; and
selecting the candidate compound as a compound that enhances phagocytosis of target cells if the degree of phagocytosis of target cells that were contacted with the candidate compound is greater than the degree of phagocytosis of target cells that were not contacted with the candidate compound, wherein the cell-binding moiety comprises an antibody fragment lacking an Fc domain.
45. The method of claim 44 , wherein the phagocytic marker comprises a phosphatidylserine head group.
46. The method of claim 44 , wherein the population of phagocytic cells comprises macrophages.
47. The method of claim 44 , wherein the target cells comprise endothelial cells.
48. The method of claim 44 , wherein the target cells comprise tumor cells.
49. The method of claim 44 , wherein the target cells comprise eukaryotic cells.
50. The method of claim 44 , wherein the phagocytic marker comprises a moiety that is naturally displayed by apoptotic or pre-apoptotic cells and enhances recognition of apoptotic or pre-apoptotic cells by phagocytic cells.
51. The method of claim 44 , wherein the phagocytic marker and the cell-binding moiety are covalently linked.
52. The method of claim 44 , wherein the phagocytic marker and the cell-binding moiety are linked via a linking moiety.
53. The method of claim 44 , wherein the phagocytic marker and the cell-binding moiety are covalently linked via a bifunctional crosslinking reagent.
54. The method of claim 44 , wherein the phagocytic marker and the cell-binding moiety are linked to first and second linking moieties, and the first and second linking moieties are linked to each other.
55. The method of claim 44 , wherein the first and second linking moieties are linked by a biotin-avidin or biotin-streptavidin interaction.
56. The method of claim 44 , wherein the compound comprises a lipid-containing vesicle comprising lipids, a cell-binding moiety, and a phagocytic marker.
57. The method of claim 44 , further comprising comparing the ability of a compound comprising a naturally occurring phagocytic marker to enhance phagocytosis with the ability of a compound comprising a variant or fragment of a naturally occurring phagocytic marker to enhance phagocytosis.
58. The method of claim 44 , further comprising comparing the degree of phagocytosis of the target cells by the phagocytic cells with the degree of phagocytosis of a population of target cells that have been induced to undergo apoptosis and exposed to phagocytic cells under similar conditions.
59. The method of claim 44 , further comprising administering the compound to an animal model of disease.