IP Library › Granted Patent US 12,196,661
Granted Patent B2
US 12,196,661 · App. 18/735,500 · Granted Jan 14, 2025

Size-tunable synthetic particles with tunable optical properties and methods for using the same for immune cell activation

Inventors: Daixuan Zhang (Fremont, CA); Solomon Stonebloom (Emeryville, CA); Juan Armas (Emeryville, CA); Harini Kethar (Danville, CA); Sunil Thomas (San Bruno, CA); Martina De Geus (Berkeley, CA)
Assignee: Slingshot Biosciences, Inc.
G01N15/0893G01N15/1012G01N15/1468G01N33/563G01N2015/1006
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Quick Facts
Patent No.
US 12,196,661
App. No.
18/735,500
Filed
Jun 6, 2024
Granted
Jan 14, 2025
Kind
B2
Examiner
WEST, PAUL M
Art Unit
2855
USPC
73/38
Abstract

A hydrogel particle, comprising a matrix comprising a polymerized monomer, said matrix comprising a plurality of micropores and a plurality of macropores, and one or more immunostimulatory biomolecules selected from the group consisting of an anti-CD3 antibody or antigen-binding fragment thereof, an anti-CD28 antibody or antigen-binding fragment thereof, and combinations thereof.

Claims (133)

1. A method of inducing an immune cell response, comprising:

contacting an immune cell with a particle comprising:

a matrix comprising a polymerized monomer having a plurality of micropores and a plurality of macropores; and

one or more immunostimulatory or co-stimulatory biomolecules,

wherein the immune cell response comprises activation and/or expansion of the immune cell, and wherein an average diameter of the plurality of macropores is between about 200 nm and about 2 μm.

2. The method of claim 1 , wherein an average diameter of the plurality of macropores is larger than an average diameter of the plurality of micropores.

3. The method of claim 1 , wherein an average diameter of the plurality of micropores is between about 1 nm and about 20 nm.

4. The method of claim 1 , wherein the average diameter of the plurality of micropores is between about 2 nm and about 4 nm.

5. The method of claim 1 , wherein the immune cell response from contacting the immune cell with the particle is higher than the immune cell response from a control immune cell contacted with an otherwise identical particle lacking the macropores.

6. The method of claim 5 , wherein the immune cell response is determined by IL-2 secretion from the immune cell.

7. The method of claim 5 , wherein the immune cell response is determined by CD25 expression from the immune cell.

8. The method of claim 5 , wherein the immune cell response is determined by CD69 expression from the immune cell.

9. The method of claim 1 , wherein the immune cell is a T cell.

10. The method of claim 1 , wherein the immune cell is a cytotoxic T cell.

11. The method of claim 1 , wherein the immune cell is a chimeric antigen receptor (CAR) T cell.

12. The method of claim 1 , wherein the particle has a porosity of about 5% to about 95% of a volume of the particle.

13. The method of claim 1 , wherein the one or more immunostimulatory or co-stimulatory biomolecules are attached to a surface of the matrix via a linker.

14. The method of claim 13 , wherein the linker comprises streptavidin.

15. The method of claim 1 , wherein the one or more immunostimulatory or co-stimulatory biomolecules are biotinylated.

16. The method of claim 1 , wherein the matrix comprises a bifunctional monomer and wherein the one or more immunostimulatory or co-stimulatory biomolecules are attached to the bifunctional monomer.

17. The method of claim 1 , wherein the one or more immunostimulatory or co-stimulatory biomolecules are selected from the group consisting of an anti-CD3 antibody or antigen-binding fragment thereof, an anti-CD28 antibody or antigen-binding fragment thereof, an anti-CD19 antibody or antigen-binding fragment, an anti-41BBL antibody or antigen-binding fragment, an anti-OX40L antibody or antigen-binding fragment, an anti-CD2 antibody or antigen-binding fragment, an anti-CD335 antibody or antigen-binding fragment, an anti-CD16 antibody or antigen-binding fragment, an anti-CD56 antibody or antigen-binding fragment, an anti-CD20 antibody or antigen-binding fragment, an anti-CD80 antibody or antigen-binding fragment, an anti-CD86 antibody or antigen-binding fragment, an anti-CD69 antibody or antigen-binding fragment, an anti-CD154 antibody or antigen-binding fragment, an anti-CD137 antibody or antigen-binding fragment, an IgM antibody or antigen-binding fragment, an IgG antibody or antigen-binding fragment, an IgE antibody or antigen-binding fragment, an IgA antibody or antigen-binding fragment, an IgD antibody or antigen-binding fragment, a toll-like receptor, and any combination thereof.

18. The method of claim 1 , wherein the one or more immunostimulatory or co-stimulatory biomolecules are selected from the group consisting of anti-CD19, anti-41BBL, anti-OX40L, anti-CD2, anti-CD335, anti-CD16, anti-CD56, anti-CD20, anti-CD80, anti-CD86, anti-CD69, anti-CD154, anti-CD137, and any combination thereof.

19. The method of claim 1 , wherein the one or more immunostimulatory or co-stimulatory biomolecules comprise:

an anti-CD3 antibody or an antigen-binding fragment thereof; and/or

an anti-CD28 antibody or an antigen-binding fragment thereof.

20. The method of claim 1 , wherein the plurality of micropores are formed during monomer polymerization.

21. The method of claim 1 , wherein the plurality of macropores comprise between about 2% and about 30% of a total number of pores of the particle, the total number of pores of the particle being a combination of the plurality of micropores and the plurality of macropores.

22. The method of claim 1 , wherein the particle comprises the plurality of macropores at a concentration of at least 2.25% v/v, at least 3.4% v/v, and/or at least 4.5% v/v.

23. The method of claim 1 , wherein the particle has a diameter of between about 1 μm and about 25 μm.

24. The method of claim 23 , wherein the particle has a diameter of between about 2 μm and about 5 μm.

25. A method for calibrating a cytometric device for analysis of a target cell, comprising,

inserting into the device a hydrogel particle comprising (i) a matrix comprising a polymerized monomer having a plurality of micropores and a plurality of macropores; and (ii) one or more immunostimulatory or co-stimulatory biomolecules, wherein the particle has at least one optical or morphological property substantially similar to the target cell;

measuring the at least one optical or morphological property of the particle using the cytometric device; and

calibrating the cytometric device based on the optical or morphological property measurement of the particle.

26. A method for detecting a target cell in a sample, comprising,

inserting into a cytometric device a hydrogel particle comprising (i) a matrix comprising a polymerized monomer having a plurality of micropores and a plurality of macropores; and (ii) one or more immunostimulatory or co-stimulatory biomolecules, wherein the particle has at least one optical or morphological property substantially similar to the target cell;

measuring the at least one optical or morphological property of the particle using the cytometric device;

inserting a sample in the cytometric device comprising a plurality of cells;

measuring the at least one optical or morphological property of individual cells of the plurality; and

determining, based on the optical or morphological property measurement, whether the target cell or plurality thereof is present in the sample.

27. A method of inducing an immune cell response, comprising:

contacting an immune cell with a particle comprising:

a matrix comprising a polymerized monomer having a plurality of micropores and a plurality of macropores; and

one or more immunostimulatory or co-stimulatory biomolecules,

wherein the immune cell response comprises activation and/or expansion of the immune cell, and

wherein the immune cell response from contacting the immune cell with the particle is higher than the immune cell response from a control immune cell contacted with an otherwise identical particle lacking the macropores.

28. The method of claim 27 , wherein an average diameter of the plurality of macropores is larger than an average diameter of the plurality of micropores.

29. The method of claim 27 , wherein an average diameter of the plurality of macropores is between about 200 nm and about 2 μm.

30. The method of claim 27 , wherein an average diameter of the plurality of micropores is between about 1 nm and about 20 nm.

31. The method of claim 27 , wherein the average diameter of the plurality of micropores is between about 2 nm and about 4 nm.

32. The method of claim 27 , wherein the immune cell response is determined by IL-2 secretion from the immune cell.

33. The method of claim 27 , wherein the immune cell response is determined by CD25 expression from the immune cell.

34. The method of claim 27 , wherein the immune cell response is determined by CD69 expression from the immune cell.

35. The method of claim 27 , wherein the immune cell is a T cell.

36. The method of claim 27 , wherein the immune cell is a cytotoxic T cell.

37. The method of claim 27 , wherein the immune cell is a chimeric antigen receptor (CAR) T cell.

38. The method of claim 27 , wherein the particle has a porosity of about 5% to about 95% of a volume of the particle.

39. The method of claim 27 , wherein the one or more immunostimulatory or co-stimulatory biomolecules are attached to a surface of the matrix via a linker.

40. The method of claim 39 , wherein the linker comprises streptavidin.

41. The method of claim 27 , wherein the one or more immunostimulatory or co-stimulatory biomolecules are biotinylated.

42. The method of claim 27 , wherein the matrix comprises a bifunctional monomer and wherein the one or more immunostimulatory or co-stimulatory biomolecules are attached to the bifunctional monomer.

43. The method of claim 27 , wherein the one or more immunostimulatory or co-stimulatory biomolecules are selected from the group consisting of an anti-CD3 antibody or antigen-binding fragment thereof, an anti-CD28 antibody or antigen-binding fragment thereof, an anti-CD19 antibody or antigen-binding fragment, an anti-41BBL antibody or antigen-binding fragment, an anti-OX40L antibody or antigen-binding fragment, an anti-CD2 antibody or antigen-binding fragment, an anti-CD335 antibody or antigen-binding fragment, an anti-CD16 antibody or antigen-binding fragment, an anti-CD56 antibody or antigen-binding fragment, an anti-CD20 antibody or antigen-binding fragment, an anti-CD80 antibody or antigen-binding fragment, an anti-CD86 antibody or antigen-binding fragment, an anti-CD69 antibody or antigen-binding fragment, an anti-CD154 antibody or antigen-binding fragment, an anti-CD137 antibody or antigen-binding fragment, an IgM antibody or antigen-binding fragment, an IgG antibody or antigen-binding fragment, an IgE antibody or antigen-binding fragment, an IgA antibody or antigen-binding fragment, an IgD antibody or antigen-binding fragment, a toll-like receptor, and any combination thereof.

44. The method of claim 27 , wherein the one or more immunostimulatory or co-stimulatory biomolecules are selected from the group consisting of anti-CD19, anti-41BBL, anti-OX40L, anti-CD2, anti-CD335, anti-CD16, anti-CD56, anti-CD20, anti-CD80, anti-CD86, anti-CD69, anti-CD154, anti-CD137, and any combination thereof.

45. The method of claim 27 , wherein the one or more immunostimulatory or co-stimulatory biomolecules comprise:

an anti-CD3 antibody or an antigen-binding fragment thereof; and/or

an anti-CD28 antibody or an antigen-binding fragment thereof.

46. The method of claim 27 , wherein the plurality of micropores are formed during monomer polymerization.

47. The method of claim 27 , wherein the plurality of macropores comprise between about 2% and about 30% of a total number of pores of the particle, the total number of pores of the particle being a combination of the plurality of micropores and the plurality of macropores.

48. The method of claim 27 , wherein the particle comprises the plurality of macropores at a concentration of at least 2.25% v/v, at least 3.4% v/v, and/or at least 4.5% v/v.

49. The method of claim 27 , wherein the particle has a diameter of between about 1 μm and about 25 μm.

50. The method of claim 49 , wherein the particle has a diameter of between about 2 μm and about 5 μm.

51. A method of inducing an immune cell response, comprising:

contacting an immune cell with a particle comprising:

a matrix comprising a polymerized monomer having a plurality of micropores and a plurality of macropores; and

one or more immunostimulatory or co-stimulatory biomolecules,

wherein the immune cell response comprises activation and/or expansion of the immune cell, and

wherein the plurality of micropores are formed during monomer polymerization.

52. The method of claim 51 , wherein an average diameter of the plurality of macropores is larger than an average diameter of the plurality of micropores.

53. The method of claim 51 , wherein an average diameter of the plurality of macropores is between about 200 nm and about 2 μm.

54. The method of claim 51 , wherein an average diameter of the plurality of micropores is between about 1 nm and about 20 nm.

55. The method of claim 51 , wherein the average diameter of the plurality of micropores is between about 2 nm and about 4 nm.

56. The method of claim 51 , wherein the immune cell response from contacting the immune cell with the particle is higher than the immune cell response from a control immune cell contacted with an otherwise identical particle lacking the macropores.

57. The method of claim 56 , wherein the immune cell response is determined by IL-2 secretion from the immune cell.

58. The method of claim 56 , wherein the immune cell response is determined by CD25 expression from the immune cell.

59. The method of claim 56 , wherein the immune cell response is determined by CD69 expression from the immune cell.

60. The method of claim 51 , wherein the immune cell is a T cell.

61. The method of claim 51 , wherein the immune cell is a cytotoxic T cell.

62. The method of claim 51 , wherein the immune cell is a chimeric antigen receptor (CAR) T cell.

63. The method of claim 51 , wherein the particle has a porosity of about 5% to about 95% of a volume of the particle.

64. The method of claim 51 , wherein the one or more immunostimulatory or co-stimulatory biomolecules are attached to a surface of the matrix via a linker.

65. The method of claim 64 , wherein the linker comprises streptavidin.

66. The method of claim 51 , wherein the one or more immunostimulatory or co-stimulatory biomolecules are biotinylated.

67. The method of claim 51 , wherein the matrix comprises a bifunctional monomer and wherein the one or more immunostimulatory or co-stimulatory biomolecules are attached to the bifunctional monomer.

68. The method of claim 51 , wherein the one or more immunostimulatory or co-stimulatory biomolecules are selected from the group consisting of an anti-CD3 antibody or antigen-binding fragment thereof, an anti-CD28 antibody or antigen-binding fragment thereof, an anti-CD19 antibody or antigen-binding fragment, an anti-41BBL antibody or antigen-binding fragment, an anti-OX40L antibody or antigen-binding fragment, an anti-CD2 antibody or antigen-binding fragment, an anti-CD335 antibody or antigen-binding fragment, an anti-CD16 antibody or antigen-binding fragment, an anti-CD56 antibody or antigen-binding fragment, an anti-CD20 antibody or antigen-binding fragment, an anti-CD80 antibody or antigen-binding fragment, an anti-CD86 antibody or antigen-binding fragment, an anti-CD69 antibody or antigen-binding fragment, an anti-CD154 antibody or antigen-binding fragment, an anti-CD137 antibody or antigen-binding fragment, an IgM antibody or antigen-binding fragment, an IgG antibody or antigen-binding fragment, an IgE antibody or antigen-binding fragment, an IgA antibody or antigen-binding fragment, an IgD antibody or antigen-binding fragment, a toll-like receptor, and any combination thereof.

69. The method of claim 51 , wherein the one or more immunostimulatory or co-stimulatory biomolecules are selected from the group consisting of anti-CD19, anti-41BBL, anti-OX40L, anti-CD2, anti-CD335, anti-CD16, anti-CD56, anti-CD20, anti-CD80, anti-CD86, anti-CD69, anti-CD154, anti-CD137, and any combination thereof.

70. The method of claim 51 , wherein the one or more immunostimulatory or co-stimulatory biomolecules comprise:

an anti-CD3 antibody or an antigen-binding fragment thereof; and/or

an anti-CD28 antibody or an antigen-binding fragment thereof.

71. The method of claim 51 , wherein the plurality of macropores comprise between about 2% and about 30% of a total number of pores of the particle, the total number of pores of the particle being a combination of the plurality of micropores and the plurality of macropores.

72. The method of claim 51 , wherein the particle comprises the plurality of macropores at a concentration of at least 2.25% v/v, at least 3.4% v/v, and/or at least 4.5% v/v.

73. The method of claim 51 , wherein the particle has a diameter of between about 1 μm and about 25 μm.

74. The method of claim 73 , wherein the particle has a diameter of between about 2 μm and about 5 μm.

75. A method of inducing an immune cell response, comprising:

contacting an immune cell with a particle comprising:

a matrix comprising a polymerized monomer having a plurality of micropores and a plurality of macropores; and

one or more immunostimulatory or co-stimulatory biomolecules,

wherein the immune cell response comprises activation and/or expansion of the immune cell, and

wherein the particle has a diameter of between about 1 μm and about 25 μm.

76. The method of claim 75 , wherein an average diameter of the plurality of macropores is larger than an average diameter of the plurality of micropores.

77. The method of claim 75 , wherein an average diameter of the plurality of macropores is between about 200 nm and about 2 μm.

78. The method of claim 75 , wherein an average diameter of the plurality of micropores is between about 1 nm and about 20 nm.

79. The method of claim 75 , wherein the average diameter of the plurality of micropores is between about 2 nm and about 4 nm.

80. The method of claim 75 , wherein the immune cell response from contacting the immune cell with the particle is higher than the immune cell response from a control immune cell contacted with an otherwise identical particle lacking the macropores.

81. The method of claim 80 , wherein the immune cell response is determined by IL-2 secretion from the immune cell.

82. The method of claim 80 , wherein the immune cell response is determined by CD25 expression from the immune cell.

83. The method of claim 80 , wherein the immune cell response is determined by CD69 expression from the immune cell.

84. The method of claim 75 , wherein the immune cell is a T cell.

85. The method of claim 75 , wherein the immune cell is a cytotoxic T cell.

86. The method of claim 75 , wherein the immune cell is a chimeric antigen receptor (CAR) T cell.

87. The method of claim 75 , wherein the particle has a porosity of about 5% to about 95% of a volume of the particle.

88. The method of claim 75 , wherein the one or more immunostimulatory or co-stimulatory biomolecules are attached to a surface of the matrix via a linker.

89. The method of claim 88 , wherein the linker comprises streptavidin.

90. The method of claim 75 , wherein the one or more immunostimulatory or co-stimulatory biomolecules are biotinylated.

91. The method of claim 75 , wherein the matrix comprises a bifunctional monomer and wherein the one or more immunostimulatory or co-stimulatory biomolecules are attached to the bifunctional monomer.

92. The method of claim 75 , wherein the one or more immunostimulatory or co-stimulatory biomolecules are selected from the group consisting of an anti-CD3 antibody or antigen-binding fragment thereof, an anti-CD28 antibody or antigen-binding fragment thereof, an anti-CD19 antibody or antigen-binding fragment, an anti-41BBL antibody or antigen-binding fragment, an anti-OX40L antibody or antigen-binding fragment, an anti-CD2 antibody or antigen-binding fragment, an anti-CD335 antibody or antigen-binding fragment, an anti-CD16 antibody or antigen-binding fragment, an anti-CD56 antibody or antigen-binding fragment, an anti-CD20 antibody or antigen-binding fragment, an anti-CD80 antibody or antigen-binding fragment, an anti-CD86 antibody or antigen-binding fragment, an anti-CD69 antibody or antigen-binding fragment, an anti-CD154 antibody or antigen-binding fragment, an anti-CD137 antibody or antigen-binding fragment, an IgM antibody or antigen-binding fragment, an IgG antibody or antigen-binding fragment, an IgE antibody or antigen-binding fragment, an IgA antibody or antigen-binding fragment, an IgD antibody or antigen-binding fragment, a toll-like receptor, and any combination thereof.

93. The method of claim 75 , wherein the one or more immunostimulatory or co-stimulatory biomolecules are selected from the group consisting of anti-CD19, anti-41BBL, anti-OX40L, anti-CD2, anti-CD335, anti-CD16, anti-CD56, anti-CD20, anti-CD80, anti-CD86, anti-CD69, anti-CD154, anti-CD137, and any combination thereof.

94. The method of claim 75 , wherein the one or more immunostimulatory or co-stimulatory biomolecules comprise:

an anti-CD3 antibody or an antigen-binding fragment thereof; and/or

an anti-CD28 antibody or an antigen-binding fragment thereof.

95. The method of claim 75 , wherein the plurality of micropores are formed during monomer polymerization.

96. The method of claim 75 , wherein the plurality of macropores comprise between about 2% and about 30% of a total number of pores of the particle, the total number of pores of the particle being a combination of the plurality of micropores and the plurality of macropores.

97. The method of claim 75 , wherein the particle comprises the plurality of macropores at a concentration of at least 2.25% v/v, at least 3.4% v/v, and/or at least 4.5% v/v.

98. The method of claim 75 , wherein the particle has a diameter of between about 2 μm and about 5 μm.

Assignments (1)
ASSIGNMENT OF ASSIGNOR'S INTEREST Recorded Jul 16, 2024
From: ZHANG, DAIXUAN; STONEBLOOM, SOLOMON; ARMAS, JUAN; KETHAR, HARINI; THOMAS, SUNIL; DE GEUS, MARTINA
To: SLINGSHOT BIOSCIENCES, INC.
Reel/Frame 067996/0507 →
Continuity (4)
Continuation PCTUS2023077961 · Oct 26, 2023
Provisional Application 63488949 · Mar 7, 2023
Provisional Application 63419580 · Oct 26, 2022
Related Publication 20240353305A1 · Oct 24, 2024
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