Claims
- 1. An antibody that is capable of binding CTLA-4, comprising a heavy chain variable region amino acid sequence that comprises a contiguous amino acid sequence from within an FR1 sequence through an FR3 sequence that is encoded by a human VH3-33 family gene and that comprises at least one of the amino acid substitutions in the CDR1 sequences, CDR2 sequences, or framework sequences shown in FIG. 2.
- 2. The antibody of claim 1, wherein the amino acid sequence comprises a sequence selected from the group consisting of SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:5, SEQ ID NO:6, SEQ ID NO:8, SEQ ID NO:9, SEQ ID NO:10, SEQ ID NO:11, SEQ ID NO:12, SEQ ID NO:13, SEQ ID NO:63, SEQ ID NO:64, SEQ ID NO:66, SEQ ID NO:68, and SEQ ID NO:70.
- 3. The antibody of claim 1, further comprising a light chain variable region amino acid sequence comprising a sequence selected from the group consisting of a sequence comprising SEQ ID NO:14, SEQ ID NO:15, SEQ ID NO:16, SEQ ID NO:17, SEQ ID NO:18, SEQ ID NO:19, SEQ ID NO:20, SEQ ID NO:21, SEQ ID NO:22, SEQ ID NO:23, SEQ ID NO:24, SEQ ID NO:25, SEQ ID NO:26, SEQ ID NO:65, SEQ ID NO:67, SEQ ID NO:69, and SEQ ID NO:71.
- 4. The antibody of claim 2, further comprising a light chain variable region amino acid sequence comprising a sequence selected from the group consisting of a sequence comprising SEQ ID NO:14, SEQ ID NO:15, SEQ ID NO:16, SEQ ID NO:17, SEQ ID NO:18, SEQ ID NO:19, SEQ ID NO:20, SEQ ID NO:21, SEQ ID NO:22, SEQ ID NO:23, SEQ ID NO:24, SEQ ID NO:25, SEQ ID NO:26, SEQ ID NO:65, SEQ ID NO:67, SEQ ID NO:69, and SEQ ID NO:71.
- 5. An antibody comprising a heavy chain amino acid sequence comprising SEQ ID NO:1 and a light chain variable amino acid sequence comprising SEQ ID NO:14.
- 6. An antibody comprising a heavy chain amino acid sequence comprising SEQ ID NO:2 and a light chain variable amino acid sequence comprising SEQ ID NO:15.
- 7. An antibody comprising a heavy chain amino acid sequence comprising SEQ ID NO:4 and a light chain variable amino acid sequence comprising SEQ ID NO:17.
- 8. A isolated human monoclonal antibody that is capable of binding to CTLA-4.
- 9. The antibody of claim 8, wherein the antibody is capable of competing for binding with CTLA-4 with an antibody selected from the group consisting of 3.1.1, 4.1.1, 4.8.1, 4.10.2, 4.13.1, 4.14.3, 6.1.1, 11.2.1, 11.6.1, 11.7.1, 12.3.1.1, and 12.9.1.1.
- 10. The antibody of claim 8, wherein the antibody possesses a substantially similar binding specificity to CTLA-4 as an antibody selected from the group consisting of 3.1.1, 4.1.1, 4.8.1, 4.10.2, 4.13.1, 4.14.3, 6.1.1, 11.2.1, 11.6.1, 11.7.1, 12.3.1.1, and 12.9.1.1.
- 11. The antibody of claim 8, wherein the antibody is selected from the group consisting of 3.1.1, 4.1.1, 4.8.1, 4.10.2, 4.13.1, 4.14.3, 6.1.1, 11.2.1, 11.6.1, 11.7.1, 12.3.1.1, and 12.9.1.1.
- 12. The antibody of claim 8, wherein the antibody is not cross reactive with CTLA-4 from lower mammalian species.
- 13. The antibody of claim 12, wherein the lower mammalian species comprises mouse, rat, and rabbit.
- 14. The antibody of claim 8, wherein the antibody is cross reactive with CTLA-4 from primates.
- 15. The antibody of claim 14, wherein the primates comprise cynomolgous and rhesus monkeys.
- 16. The antibody of claim 8, wherein the antibody possesses a selectivity for CTLA-4 over CD28, B7-2, CD44, and hIgG1 of greater than about 100:1.
- 17. The antibody of claim 16, wherein the selectivity is about 500:1 or greater.
- 18. The antibody of claim 8, wherein a binding affinity of the antibody is about 10−9 M or greater.
- 19. The antibody of claim 18, wherein a binding affinity of the antibody is about 10−10 M or greater.
- 20. The antibody of claim 8, wherein the antibody inhibits binding between CTLA-4 and B7-2 with an IC50 of lower than about 100 nM.
- 21. The antibody of claim 20, wherein the antibody inhibits binding between CTLA-4 and B7-2 with an IC50 of lower than about 0.38 nM.
- 22. The antibody of claim 8, wherein the antibody inhibits binding between CTLA-4 and B7-1 with an IC50 of lower than about 100 nM or greater.
- 23. The antibody of claim 22, wherein the antibody inhibits binding between CTLA-4 and B7-1 with an IC50 of lower than about 0.50 nM.
- 24. The antibody of claim 8, wherein the antibody enhances IL-2 production in a T cell blast/Raji assay by about 500 pg/ml or greater.
- 25. The antibody of claim 24, wherein the antibody enhances IL-2 production in a T cell blast/Raji assay by about 3846 pg/ml or greater.
- 26. The antibody of claim 8, wherein the antibody enhances IFN-γ production in a T cell blast/Raji assay by about 500 pg/ml or greater.
- 27. The antibody of claim 26, wherein the antibody enhances IFN-γ production in a T cell blast/Raji assay by about 1233 pg/ml or greater.
- 28. The antibody of claim 8, wherein the antibody induces IL-2 production in a hPBMC or whole blood superantigen assay by about 500 pg/ml or greater.
- 29. The antibody of claim 26, wherein the antibody induces IL-2 production in a hPBMC or whole blood superantigen assay by about 1500 pg/ml or greater
- 30. The antibody of claim 26, wherein the antibody induces IL-2 production in a hPBMC or whole blood superantigen assay by greater than about 30% relative to control.
- 31. The antibody of claim 26, wherein the antibody induces IL-2 production in a hPBMC or whole blood superantigen assay by greater than about 50% relative to control.
- 32. A humanized antibody that possesses a substantially similar binding specificity to CTLA-4 as an antibody selected from the group consisting of 3.1.1, 4.1.1, 4.8.1, 4.10.2, 4.13.1, 4.14.3, 6.1.1, 11.2.1, 11.6.1, 11.7.1, 12.3.1.1, and 12.9.1.1.
- 33. The antibody of claim 32, wherein the antibody is not cross reactive with CTLA-4 from lower mammalian species.
- 34. The antibody of claim 33, wherein the lower mammalian species comprises mouse, rat, and rabbit.
- 35. The antibody of claim 32, wherein the antibody is cross reactive with CTLA-4 from primates.
- 36. The antibody of claim 35, wherein the primates comprise cynomolgous and rhesus monkeys.
- 37. The antibody of claim 32, wherein the antibody possesses a selectivity for CTLA-4 over CD28, B7-2, CD44, and hIgG1 of greater than about 100:1.
- 38. The antibody of claim 37, wherein the selectivity is about 500:1 or greater.
- 39. The antibody of claim 32, wherein a binding affinity of the antibody is about 10−9 M or greater.
- 40. The antibody of claim 39, wherein a binding affinity of the antibody is about 10−10 M or greater.
- 41. The antibody of claim 32, wherein the antibody inhibits binding between CTLA-4 and B7-2 with an IC50 of lower than about 100 nM.
- 42. The antibody of claim 41, wherein the antibody inhibits binding between CTLA-4 and B7-2 with an IC50 of lower than about 0.38 nM.
- 43. The antibody of claim 32, wherein the antibody inhibits binding between CTLA-4 and B7-1 with an IC50 of lower than about 100 nM or greater.
- 44. The antibody of claim 43, wherein the antibody inhibits binding between CTLA-4 and B7-1 with an IC50 of lower than about 0.50 nM.
- 45. The antibody of claim 32, wherein the antibody enhances IL-2 production in a T cell blast/Raji assay by about 500 pg/ml or greater.
- 46. The antibody of claim 45, wherein the antibody enhances IL-2 production in a T cell blast/Raji assay by about 3846 pg/ml or greater.
- 47. The antibody of claim 32, wherein the antibody enhances IFN-γ production in a T cell blast/Raji assay by about 500 pg/ml or greater.
- 48. The antibody of claim 47, wherein the antibody enhances IFN-γ production in a T cell blast/Raji assay by about 1233 pg/ml or greater.
- 49. The antibody of claim 32, wherein the antibody enhances IL-2 production in a hPBMC or whole blood superantigen assay by about 500 pg/ml or greater.
- 50. The antibody of claim 49, wherein the antibody enhances IL-2 production in a hPBMC or whole blood superantigen assay by about 1500 pg/ml or greater
- 51. The antibody of claim 32, wherein the antibody enhances IL-2 production in a hPBMC or whole blood superantigen assay by greater than about 30% relative to control.
- 52. The antibody of claim 50, wherein the antibody enhances IL-2 production in a hPBMC or whole blood superantigen assay by greater than about 50% relative to control.
- 53. An antibody that binds to CTLA-4, comprising a heavy chain amino acid sequence comprising human FR1, FR2, and FR3 sequences encoded by a human VH 3-33 gene family operably linked in frame with a CDR1, a CDR2, and a CDR3 sequence, the CDR1, CDR2, and CDR3 sequences being independently selected from the CDR1, CDR2, and CDR3 sequences illustrated in FIG. 2.
- 54. The antibody of claim 32, further comprising any of the somatic mutations to the FR1, FR2, and FR3 sequences as illustrated in FIG. 2.
- 55. An antibody that binds to CTLA-4, comprising a heavy chain amino acid sequence comprising human FR1, FR2, and FR3 sequences encoded by a human VH 3-33 gene family operably linked in frame with a CDR1, a CDR2, and a CDR3 sequence, which antibody has the following properties:
a binding affinity for CTLA-4 of about 10−9 or greater; inhibits binding between CTLA-4 and B7-1 with an IC50 of about 100 nM or lower; inhibits binding between CTLA-4 and B7-2 with an IC50 of about 100 nM or lower; and enhances cytokine production in an assay of human T cells by 500 pg/ml or greater.
- 56. An antibody that binds to CTLA-4, comprising a heavy chain amino acid sequence comprising FR1, FR2, and FR3 sequences operably linked in frame with a CDR1, a CDR2, and a CDR3 sequence independently selected from the CDR1, CDR2, and CDR3 sequences illustrated in FIGS. 2 and 3, which antibody has the following properties:
a binding affinity for CTLA-4 of about 10−9 or greater; inhibits binding between CTLA-4 and B7-1 with an IC50 of about 100 nM or lower; inhibits binding between CTLA-4 and B7-2 with an IC50 of about 100 nM or lower; and enhances cytokine production in an assay of human T cells by 500 pg/ml or greater.
- 57. A cell culture system for assaying T cell stimulation, comprising a culture of human T cell blasts co-cultured with a Raji cell line.
- 58. The cell culture system of claim 57, wherein the T cell blasts are washed prior to culture with the Raji cell line.
- 59. An assay for measuring T cell stimulation, comprising:
providing a culture of human T cell blasts and a Raji cell line; contacting the culture with an agent; and measuring cytokine production by the culture.
- 60. The assay of claim 59, wherein the T cell blasts are washed prior to culture with the Raji cell line.
- 61. The assay of claim 59, wherein the cytokine is IL-2.
- 62. The assay of claim 59, wherein the cytokine is IFN-γ.
- 63. The assay of claim 59, wherein cytokine production is measured in supernatant isolated from the culture.
- 64. The assay of claim 59, wherein the agent is an antibody.
- 65. The assay of claim 59, wherein the antibody binds to CTLA-4.
- 66. A functional assay for screening a moiety for T cell stimulatory function, comprising:
providing a culture of human T cell blasts and a Raji cell line; contacting the culture with the moiety; and assessing cytokine production by the culture.
- 67. The assay of claim 66, wherein the T cell blasts are washed prior to culture with the Raji cell line.
- 68. The assay of claim 66, wherein the cytokine is IL-2.
- 69. The assay of claim 66, wherein the cytokine is IFN-γ.
- 70. The assay of claim 66, wherein cytokine production is assessed in supernatant isolated from the culture.
- 71. The assay of claim 66, wherein the moiety is an antibody.
- 72. The assay of claim 66, wherein the antibody binds to CTLA-4.
- 73. A T cell stimulatory assay for CTLA-4 inhibitory function, comprising contacting a culture comprising human T cell blasts and a Raji cell line with an agent and assessing cytokine production by the culture.
- 74. The assay of claim 73, wherein the T cell blasts are washed prior to culture with the Raji cell line.
- 75. The assay of claim 73, wherein the cytokine is IL-2.
- 76. The assay of claim 73, wherein the cytokine is IFN-γ.
- 77. The assay of claim 73, wherein cytokine production is assessed in supernatant isolated from the culture.
- 78. The assay of claim 73, wherein the agent is an antibody.
- 79. The assay of claim 73, wherein the antibody binds to CTLA-4.
- 80. A method for screening an agent for T cell stimulatory activity, comprising:
contacting the agent with a cell culture comprising human T cell blasts and a Raji cell line; and assessing cytokine production by the culture.
- 81. The method of claim 80, wherein the T cell blasts are washed prior to culture with the Raji cell line.
- 82. The method of claim 80, wherein the cytokine is IL-2.
- 83. The method of claim 80, wherein the cytokine is IFN-γ.
- 84. The method of claim 80, wherein cytokine production is assessed in supernatant isolated from the culture.
- 85. The method of claim 80, wherein the agent is an antibody that binds to CTLA-4.
- 86. An assay for measuring T cell stimulation, comprising:
providing a population of human peripheral blood mononuclear cells or human whole blood stimulated with staphylococcus enterotoxin A; contacting the culture with an agent; and measuring cytokine production by the cell population.
- 87. The assay of claim 86, wherein the cytokine is IL-2.
- 88. The assay of claim 86, wherein cytokine production is measured in supernatant isolated from the cell population.
- 89. The assay of claim 86, wherein the agent is an antibody.
- 90. The assay of claim 86, wherein the antibody binds to CTLA-4.
- 91. A functional assay for screening a moiety for T cell stimulatory function, comprising:
providing a population of human peripheral blood mononuclear cells or human whole blood stimulated with staphylococcus enterotoxin A; contacting the culture with the moiety; and assessing cytokine production by the cell population.
- 92. The assay of claim 91, wherein the cytokine is IL-2.
- 93. The assay of claim 91, wherein cytokine production is assessed in supernatant isolated from the cell population.
- 94. The assay of claim 91, wherein the moiety is an antibody.
- 95. The assay of claim 91, wherein the antibody binds to CTLA-4.
- 96. A T cell stimulatory assay for CTLA-4 inhibitory function, comprising contacting a population of human peripheral blood mononuclear cells or human whole blood stimulated with staphylococcus enterotoxin A with an agent and assessing cytokine production by the cell population.
- 97. The assay of claim 96, wherein the cytokine is IL-2.
- 98. The assay of claim 96, wherein cytokine production is assessed in supernatant isolated from the cell population.
- 99. The assay of claim 96, wherein the agent is an antibody.
- 100. The assay of claim 96, wherein the antibody binds to CTLA-4.
- 101. A method for screening an agent for T cell stimulatory activity, comprising:
contacting the agent with a population of human peripheral blood mononuclear cells or human whole blood stimulated with staphylococcus enterotoxin A; and assessing cytokine production by the cell population.
- 102. The method of claim 101, wherein the cytokine is IL-2.
- 103. The method of claim 101, wherein cytokine production is assessed in supernatant isolated from the cell population.
- 104. The method of claim 101, wherein the agent is an antibody that binds to CTLA-4.
CROSS-REFERENCE TO RELATED APPLICATIONS
[0001] The present application is a divisional application of U.S. patent application Ser. No. 09/472,087, filed Dec. 23, 1999, which claims priority to U.S. Provisional Patent Application Ser. No. 60/113,647, filed Dec. 23, 1998, the disclosures of which are hereby incorporated in their entirety herein.
Provisional Applications (1)
|
Number |
Date |
Country |
|
60113647 |
Dec 1998 |
US |
Divisions (1)
|
Number |
Date |
Country |
Parent |
09472087 |
Dec 1999 |
US |
Child |
10612497 |
Jul 2003 |
US |