Stable antibody formulations
阅读说明：本技术 稳定的抗体制剂 (Stable antibody formulations ) 是由 Q·胡 D·刘 于 2018-03-23 设计创作，主要内容包括：本发明提供稳定的药物制剂,其包含特异性结合人程序性死亡-1蛋白(PD-1)的人抗体。在某些实施方案中,除抗PD-1抗体外,所述制剂还含有缓冲剂、氨基酸、非离子表面活性剂和糖。本发明的药物制剂在应激和储存后表现出相当程度的抗体稳定性。(The present invention provides a stable pharmaceutical formulation comprising a human antibody that specifically binds to human programmed death-1 protein (PD-1). In certain embodiments, the formulation contains, in addition to the anti-PD-1 antibody, a buffer, an amino acid, a nonionic surfactant, and a sugar. The pharmaceutical formulations of the present invention exhibit a considerable degree of antibody stability after stress and storage.)
1. A liquid pharmaceutical formulation comprising:
(a) an antibody that specifically binds human programmed death-1 (PD-1), wherein the antibody comprises SEQ ID NO: 1 (HCDR1, HCDR2 and HCDR3) and the three heavy chain Complementarity Determining Regions (CDRs) contained in the Heavy Chain Variable Region (HCVR) of SEQ ID NO: 2 (LCDR1, LCDR2, and LCDR 3);
(b) a buffer comprising histidine;
(c) an organic solvent comprising polysorbate;
(d) a stabilizer comprising a sugar; and
(e) a viscosity modifier comprising an amino acid;
wherein the formulation has a pH of 6.0 + -0.3.
2. The pharmaceutical formulation of claim 1, wherein the antibody concentration is 5mg/mL ± 0.75mg/mL to 250mg/mL ± 37.5 mg/mL.
3. The pharmaceutical formulation of claim 2, wherein the antibody concentration is 25mg/mL ± 3.75 mg/mL.
4. The pharmaceutical formulation of claim 2, wherein the antibody concentration is 50mg/mL ± 7.5 mg/mL.
5. The pharmaceutical formulation of claim 2, wherein the antibody concentration is 150mg/mL ± 22.5 mg/mL.
6. The pharmaceutical formulation of claim 2, wherein the antibody concentration is 175mg/mL ± 26.25 mg/mL.
7. The pharmaceutical formulation of any one of claims 1-6, wherein histidine buffer concentration is 5mM ± 1mM to 20mM ± 4 mM.
8. The pharmaceutical formulation of claim 7, wherein histidine buffer concentration is 10mM ± 2 mM.
9. The pharmaceutical formulation of claim 7 or 8, wherein histidine buffer comprises L-histidine and L-histidine monohydrochloride monohydrate.
10. The pharmaceutical formulation of claim 9, wherein the concentration of L-histidine is 4.8mM ± 0.96mM and the concentration of L-histidine monohydrochloride monohydrate is 5.2mM ± 1.04 mM.
11. The pharmaceutical formulation according to any one of claims 1 to 10, wherein the polysorbate concentration is 0.01% ± 0.005% to 0.5% ± 0.25% w/v.
12. The pharmaceutical formulation of claim 11, wherein the polysorbate concentration is 0.1% ± 0.05% w/v.
13. The pharmaceutical formulation of claim 11, wherein the polysorbate concentration is 0.2% ± 0.1% w/v.
14. The pharmaceutical formulation of any one of claims 11-13, wherein the organic solvent is polysorbate 80.
15. The pharmaceutical formulation of any one of claims 1-14, wherein the stabilizer is sucrose and the sucrose concentration is 0% to 20% ± 4% w/v.
16. The pharmaceutical formulation according to claim 15, wherein the sucrose concentration is 1% ± 0.2% to 10% ± 2% w/v.
17. The pharmaceutical formulation of claim 16, wherein the sucrose concentration is 5% ± 1% w/v.
18. The pharmaceutical formulation of claim 17, wherein the viscosity modifier is proline.
19. The pharmaceutical formulation of claim 18, wherein the proline concentration is 0 to 5% ± 1% w/v.
20. The pharmaceutical formulation of claim 19, wherein the proline concentration is 1.5% ± 0.3% w/v.
21. The pharmaceutical formulation of claim 18, comprising:
(a)175 mg/mL. + -. 26.25mg/mL antibody,
(b)5mM + -1 mM to 20mM + -4 mM histidine buffer,
(c) 0.1% + -0.05% to 0.5% + -0.25% w/v polysorbate,
(d) 1% + -0.2% to 10% + -2% w/v sucrose, and
(e) 1% ± 0.2% to 5% ± 1% w/v proline;
the pH was 6.0. + -. 0.3.
22. The pharmaceutical formulation of claim 21, comprising:
(a)175 mg/mL. + -. 26.25mg/mL antibody,
(b)10 mM. + -. 2mM histidine buffer,
(c) 0.2% + -0.1% w/v polysorbate,
(d) 5% +/-1% w/v sucrose, and
(e) 1.5% ± 0.3% w/v proline;
the pH was 6.0. + -. 0.3.
23. The pharmaceutical formulation of claim 22, comprising:
(a)175 mg/mL. + -. 26.25mg/mL antibody,
(b)4.8 mM. + -. 0.96mM L-histidine,
(c)5.2 mM. + -. 1.04mM L-histidine monohydrochloride monohydrate,
(d) 0.2% + -0.1% w/v polysorbate,
(e) 5% +/-1% w/v sucrose, and
(f) 1.5% ± 0.3% w/v proline;
the pH was 6.0. + -. 0.3.
24. The pharmaceutical formulation of claim 18, comprising:
(a)150 mg/mL. + -. 22.5mg/mL antibody,
(b)5mM + -1 mM to 20mM + -4 mM histidine buffer,
(c) 0.1% + -0.05% to 0.5% + -0.25% w/v polysorbate,
(d) 1% + -0.2% to 10% + -2% w/v sucrose, and
(e) 1% ± 0.2% to 5% ± 1% w/v proline;
the pH was 6.0. + -. 0.3.
25. The pharmaceutical formulation of claim 24, comprising:
(a)150 mg/mL. + -. 22.5mg/mL antibody,
(b)10 mM. + -. 2mM histidine buffer,
(c) 0.2% + -0.1% w/v polysorbate,
(d) 5% +/-1% w/v sucrose, and
(e) 1.5% + -0.3% w/v proline,
the pH was 6.0. + -. 0.3.
26. The pharmaceutical formulation of claim 25, comprising:
(a)150 mg/mL. + -. 22.5mg/mL antibody,
(b)4.8 mM. + -. 0.96mM L-histidine,
(c)5.2 mM. + -. 1.04mM L-histidine monohydrochloride monohydrate,
(d) 0.2% + -0.1% w/v polysorbate,
(e) 5% +/-1% w/v sucrose, and
(f) 1.5% ± 0.3% w/v proline;
the pH was 6.0. + -. 0.3.
27. The pharmaceutical formulation of claim 18, comprising:
(a)50 mg/mL. + -. 7.5mg/mL antibody,
(b)5mM + -1 mM to 20mM + -4 mM histidine buffer,
(c) 0.1% + -0.05% to 0.5% + -0.25% w/v polysorbate,
(d) 1% + -0.2% to 10% + -2% w/v sucrose, and
(e) 1% + -0.2% to 5% + -1% w/v proline,
the pH was 6.0. + -. 0.3.
28. The pharmaceutical formulation of claim 27, comprising:
(a)50 mg/mL. + -. 7.5mg/mL antibody,
(b)10 mM. + -. 2mM histidine buffer,
(c) 0.2% + -0.1% w/v polysorbate,
(d) 5% +/-1% w/v sucrose, and
(e) 1.5% + -0.3% w/v proline,
the pH was 6.0. + -. 0.3.
29. The pharmaceutical formulation of claim 28, comprising
(a)50 mg/mL. + -. 7.5mg/mL antibody,
(b)4.8 mM. + -. 0.96mM L-histidine,
(c)5.2 mM. + -. 1.04mM L-histidine monohydrochloride monohydrate,
(d) 0.2% + -0.1% w/v polysorbate,
(e) 5% +/-1% w/v sucrose, and
(f) 1.5% ± 0.3% w/v proline;
the pH was 6.0. + -. 0.3.
30. The pharmaceutical formulation of claim 18, comprising:
(a)25 mg/mL. + -. 3.75mg/mL antibody,
(b)5mM + -1 mM to 20mM + -4 mM histidine buffer,
(c) 0.1% + -0.05% to 0.5% + -0.25% w/v polysorbate,
(d) 1% + -0.2% to 10% + -2% w/v sucrose, and
(e) 1% + -0.2% to 5% + -1% w/v proline,
the pH was 6.0. + -. 0.3.
31. The pharmaceutical formulation of claim 30, comprising:
(a)25 mg/mL. + -. 3.75mg/mL antibody,
(b)10 mM. + -. 2mM histidine buffer,
(c) 0.2% + -0.1% w/v polysorbate,
(d) 5% +/-1% w/v sucrose, and
(e) 1.5% + -0.3% w/v proline,
the pH was 6.0. + -. 0.3.
32. The pharmaceutical formulation of claim 31, comprising:
(a)25 mg/mL. + -. 3.75mg/mL antibody,
(b)4.8 mM. + -. 0.96mM L-histidine,
(c)5.2 mM. + -. 1.04mM L-histidine monohydrochloride monohydrate,
(d) 0.2% + -0.1% w/v polysorbate,
(e) 5% +/-1% w/v sucrose, and
(f) 1.5% ± 0.3% w/v proline;
the pH was 6.0. + -. 0.3.
33. The pharmaceutical formulation of any one of claims 1-32, wherein the formulation has a viscosity of less than 20 cP.
34. The pharmaceutical formulation of any one of claims 1-33, wherein at least 90% of the antibody has native conformation after 28 days at 45 ℃.
35. The pharmaceutical formulation of any one of claims 1-34, wherein at least 35% of the antibody is the primary charge variant of the antibody after 28 days at 45 ℃.
36. The pharmaceutical formulation of any one of claims 1-35, wherein at least 94% of the antibody has native conformation after three months at 25 ℃.
37. The pharmaceutical formulation of any one of claims 1-36, wherein at least 44% of the antibodies are primary charge variants of the antibodies after three months at 25 ℃.
38. The pharmaceutical formulation of any one of claims 1-37, wherein at least 96% of the antibody has native conformation after 12 months at 5 ℃.
39. The pharmaceutical formulation of any one of claims 1-38, wherein at least 45% of the antibody is the primary charge variant of the antibody after 12 months at 5 ℃.
40. The pharmaceutical formulation of any one of claims 1-39, wherein at least 96% of the antibody has native conformation after 12 months at-20 ℃, -30 ℃ and/or-80 ℃.
41. A pharmaceutical formulation as claimed in any one of claims 1 to 40 wherein at least 40% of the antibody is the primary charge variant of the antibody after 12 months at-20 ℃, -30 ℃ and/or-80 ℃.
42. A pharmaceutical formulation, comprising:
(a) an antibody that specifically binds PD-1 at 175mg/mL ± 26.25mg/mL, wherein the antibody comprises SEQ ID NO: 1 and the HCVR of SEQ ID NO: the LCVR of 2 is used for determining the position of the target,
(b)10 mM. + -. 2mM histidine buffer, pH 6.0. + -. 0.3,
(c) 0.2% + -0.1% w/v polysorbate 80,
(d) 5% ± 1% w/v sucrose; and
(e) 1.5% ± 0.3% w/v proline; wherein:
(i) more than or equal to 90 percent of the antibody has the molecular weight of 143kDa +/-1 kDa;
(ii) the pharmaceutical formulation has a viscosity of less than 20 cP; and
(iii) at least 97% or more of the antibody has native conformation after 6 months of storage at-80 ℃, -30 ℃ and/or-20 ℃.
43. The pharmaceutical formulation of claim 42, consisting of an aqueous solution of: (a)175 mg/mL. + -. 26.25mg/mL antibody, (b)4.8 mM. + -. 0.96mM L-histidine, (c)5.2 mM. + -. 1.04mM L-histidine monohydrochloride monohydrate, (d) 0.2%. + -. 0.1% w/v polysorbate 80, (e) 5%. + -. 1% w/v sucrose, and (f) 1.5%. + -. 0.3% w/v proline, pH 6.0. + -. 0.3.
44. A pharmaceutical formulation, comprising:
(a) an antibody that specifically binds PD-1 at 150mg/mL ± 22.5mg/mL, wherein the antibody comprises SEQ ID NO: 1 and the HCVR of SEQ ID NO: the LCVR of 2 is used for determining the position of the target,
(b)10 mM. + -. 2mM histidine buffer, pH6. + -. 0.3,
(c) 0.2% + -0.1% w/v polysorbate 80,
(d) 5% ± 1% w/v sucrose; and
(e) 1.5% ± 0.3% w/v proline; wherein:
(i) more than or equal to 90 percent of the antibody has the molecular weight of 143kDa +/-1 kDa;
(ii) the pharmaceutical formulation has a viscosity of less than 15 cP; and
(iii) at least 97% or more of the antibody has native conformation after 6 months of storage at-80 ℃, -30 ℃ or-20 ℃.
45. The pharmaceutical formulation of claim 44, consisting of aqueous solutions of: (a)150 mg/mL. + -. 22.5mg/mL antibody, (b)4.8 mM. + -. 0.96mM L-histidine, (c)5.2 mM. + -. 1.04mM L-histidine monohydrochloride monohydrate, (d) 0.2%. + -. 0.1% w/v polysorbate 80, (e) 5%. + -. 1% w/v sucrose, and (f) 1.5%. + -. 0.3% w/v proline, pH 6.0. + -. 0.3.
46. A pharmaceutical formulation, comprising:
(a) an antibody that specifically binds PD-1 at 50mg/mL ± 7.5mg/mL, wherein the antibody comprises the amino acid sequence of SEQ ID NO: 1 and the HCVR of SEQ ID NO: the LCVR of 2 is used for determining the position of the target,
(b)10 mM. + -. 2mM histidine buffer, pH6. + -. 0.3,
(c) 0.2% + -0.1% w/v polysorbate 80,
(d) 1.5% + -0.3% w/v proline, and
(e) 5% ± 1% w/v sucrose; wherein:
(i) more than or equal to 90 percent of the antibody has the molecular weight of 143kDa +/-1 kDa;
(ii) after 12 months of storage at 5 ℃, more than 96% of the antibody has native conformation; and
(iii) the pharmaceutical formulation has a viscosity of less than 15 cP.
47. The pharmaceutical formulation of claim 46, consisting of aqueous solutions of: (a)50 mg/mL. + -. 7.5mg/mL antibody, (b)4.8 mM. + -. 0.96mM L-histidine, (c)5.2 mM. + -. 1.04mM L-histidine monohydrochloride monohydrate, (d) 0.2%. + -. 0.1% w/v polysorbate 80, (e) 5%. + -. 1% w/v sucrose, and (f) 1.5%. + -. 0.3% w/v proline, pH 6.0. + -. 0.3.
48. A pharmaceutical formulation, comprising:
(a) an antibody that specifically binds PD-1 at 25mg/mL ± 3.75mg/mL, wherein the antibody comprises SEQ ID NO: 1 and the HCVR of SEQ ID NO: the LCVR of 2 is used for determining the position of the target,
(b)10 mM. + -. 2mM histidine buffer, pH 6.0. + -. 0.3,
(c) 0.2% + -0.1% w/v polysorbate 80,
(d) 5% ± 1% w/v sucrose; and
(e) 1.5% ± 0.3% w/v proline; wherein:
(a) more than or equal to 90 percent of the antibody has the molecular weight of 143kDa +/-1 kDa;
(b) the pharmaceutical formulation has a viscosity of less than 10 cP; and
(c) after 12 months of storage at 5 ℃, at least 96% of the antibody has the native conformation.
49. The pharmaceutical formulation of claim 48, consisting of aqueous solutions of: (a)25mg/mL ± 3.75mg/mL antibody, (b)4.8mM ± 0.96mM L-histidine, (c)5.2mM ± 1.04mM L-histidine monohydrochloride monohydrate, (d) 0.2% w/v ± 0.1% polysorbate 80, (e) 5% w/v ± 1% sucrose, and (f) 1.5% w/v ± 0.3% proline, ph6.0 ± 0.3.
50. The pharmaceutical formulation of any one of claims 1-49, wherein the antibody comprises the amino acid sequence of SEQ ID NO: 3, HCDR1 of SEQ ID NO: 4 HCDR2, SEQ ID NO: 5 HCDR3, SEQ ID NO: LCDR1 of SEQ ID NO: LCDR2 of 7 and SEQ ID NO: LCDR3 of 8.
51. The pharmaceutical formulation of claim 50, wherein the antibody comprises the amino acid sequence of SEQ ID NO: 1 and SEQ id no: LCVR of 2.
52. The pharmaceutical formulation of any one of claims 1-51, wherein the antibody comprises an amino acid sequence identical to SEQ ID NO: 1 HCVR with 90% sequence identity.
53. The pharmaceutical formulation of any one of claims 1-51, wherein the antibody comprises an amino acid sequence identical to SEQ ID NO: 2 LCVR with 90% sequence identity.
54. The pharmaceutical formulation of any one of claims 1-51, wherein the antibody comprises an amino acid sequence identical to SEQ ID NO: 1 HCVR having 90% sequence identity and a sequence identical to SEQ ID NO: 2 LCVR with 90% sequence identity.
55. The pharmaceutical formulation of any one of claims 1-51, wherein the antibody comprises a heavy chain and a light chain, wherein the heavy chain comprises a heavy chain sequence selected from the group consisting of SEQ ID NOs: 9 and 11.
56. The pharmaceutical formulation of any one of claims 1-51, wherein the antibody comprises a heavy chain and a light chain, wherein the heavy chain comprises the amino acid sequence of SEQ ID NO: 9.
57. The pharmaceutical formulation of any one of claims 1-51, wherein the antibody comprises a heavy chain and a light chain, wherein the heavy chain comprises the amino acid sequence of SEQ ID NO: 11.
58. The pharmaceutical formulation of any one of claims 1-51, wherein the antibody comprises a heavy chain and a light chain, wherein the light chain comprises the amino acid sequence of SEQ ID NO: 10.
59. The pharmaceutical formulation of any one of claims 1-51, wherein the antibody comprises a heavy/light chain comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 9/10 and 11/10.
60. The pharmaceutical formulation of any one of claims 1-51, wherein the antibody comprises a heavy/light chain comprising the amino acid sequence of SEQ ID NO: 9/10 in a pharmaceutically acceptable carrier.
61. The pharmaceutical formulation of any one of claims 1-51, wherein the antibody comprises a heavy/light chain comprising the amino acid sequence of SEQ ID NO: 11/10 in a pharmaceutically acceptable carrier.
62. The pharmaceutical formulation of any one of claims 1-61, wherein the formulation is contained in a container.
63. The pharmaceutical formulation of claim 62, wherein the container is a vial.
64. The pharmaceutical formulation of claim 63, wherein the vial is a 10mL type 1 clear glass vial.
65. The pharmaceutical formulation of claim 62, wherein the container is a syringe.
66. The pharmaceutical formulation of claim 65, wherein the syringe is low tungsten glass.
67. The pharmaceutical formulation of claim 62, wherein the container is a prefilled syringe.
68. The pharmaceutical formulation of claim 62, contained in an autoinjector.
69. A kit comprising the pharmaceutical formulation of any one of claims 1-61, a container, and instructions.
70. The kit of claim 69, wherein the container is a glass vial.
71. The kit of claim 69, wherein the container is a pre-filled syringe.
72. The kit of claim 69, wherein the container is an auto-injector.
Technical Field
The present invention relates to the field of therapeutic antibody formulations. More specifically, the present invention relates to the field of pharmaceutical formulations comprising human antibodies that specifically bind to the human programmed death-1 (PD-1) protein.
Background
Therapeutic macromolecules (e.g., antibodies) must be formulated in such a way that not only is the molecule suitable for administration to a patient, but also retains its stability during storage and subsequent use. For example, therapeutic antibodies in liquid solutions are susceptible to degradation, aggregation, or unwanted chemical modification unless the solution is properly formulated. The stability of an antibody in a liquid formulation depends not only on the kind of excipients used in the formulation, but also on the amount and ratio of excipients relative to each other. In addition, other factors besides stability must be considered in preparing liquid antibody formulations. Examples of such additional considerations include the viscosity of the solution and the concentration of antibody that a given formulation can accommodate, as well as the visual quality or appeal of the formulation. Thus, when formulating therapeutic antibodies, great care must be taken to obtain a formulation that is stable, contains sufficient antibody concentration, and has the appropriate viscosity and other properties that enable the formulation to be conveniently administered to a patient.
Antibodies to the human programmed death-1 protein (PD-1) are one example of a therapeutically relevant macromolecule that requires appropriate formulation. anti-PD-1 antibodies are clinically useful for the treatment of cancer (e.g., lung, melanoma, and brain cancer) and viral infections and autoimmune diseases. Exemplary anti-PD-1 antibodies are described, inter alia, in U.S. patent/publication nos. 7101550, 7595048, 7488802, 7563869, 8008449, 8168757, 8216996, 20110008369, 20130017199, 20130022595, and WO2006121168, WO2009114335, WO2012145493, WO2013014668, WO2009101611, EP2262837, and EP 2504028. US20140234296 describes lyophilized formulations of anti-PD-1 antibodies.
Although anti-PD-1 antibodies are known, there is still a need in the art for novel pharmaceutical formulations comprising anti-PD-1 antibodies that are sufficiently stable and suitable for administration to patients.
Disclosure of Invention
The present invention meets the above-described need by providing a stable pharmaceutical formulation comprising a human antibody that specifically binds to human programmed death-1 protein (PD-1).
In one aspect, a stable liquid pharmaceutical formulation of low viscosity is provided comprising: (i) a human antibody that specifically binds to human programmed death protein (PD-1); (ii) a buffering agent; (iii) an organic co-solvent; (iv) a stabilizer; (v) a viscosity modifier.
In various embodiments, the antibody is provided at a concentration of about 5 ± 0.75mg/mL to about 250 ± 37.5 mg/mL. In one embodiment, the antibody is provided at a concentration of 12.5mg/mL ± 1.85mg/mL, or about 12.5 mg/mL. In one embodiment, the antibody is provided at a concentration of 25mg/mL ± 3.75mg/mL, or about 25 mg/mL. In another embodiment, the antibody is provided at a concentration of 50mg/mL ± 7.5mg/mL, or about 50 mg/mL. In another embodiment, the antibody is provided at a concentration of 100mg/mL ± 15mg/mL, or about 100 mg/mL. In one embodiment, the antibody is provided at a concentration of 150mg/mL ± 22.5mg/mL, or about 150 mg/mL. In another embodiment, the antibody is provided at a concentration of 175mg/mL ± 26.25mg/mL, or about 175 mg/mL. In another embodiment, the antibody is provided at a concentration of 200mg/mL ± 30mg/mL, or about 200 mg/mL.
In certain embodiments, the formulation comprises U.S. patent application publication nos.: 20150203579, which is incorporated herein in its entirety. In certain embodiments, the anti-PD-1 antibody comprises (a) a Heavy Chain Variable Region (HCVR) comprising heavy chain
In one embodiment, the pH of the liquid formulation is pH6.0 + -0.5, pH6.0 + -0.4, pH6.0 + -0.3, pH6.0 + -0.2, pH6.0 + -0.1, pH6.0 + -0.05, pH6.0 + -0.01, or pH 6.0. In one embodiment, the pH of the liquid formulation is about pH6.0 ± 0.3.
In one embodiment, the buffering agent comprises histidine. In certain embodiments, the concentration of histidine buffer is from 5mM + -1 mM to 50mM + -10 mM, preferably from 5mM + -1 mM to 25mM + -5 mM. In one embodiment, the concentration of histidine buffer is 10mM ± 2mM or about 10 mM. In one embodiment, the concentration of histidine buffer is 20mM ± 4mM or about 20 mM. In one embodiment, the concentration of histidine buffer is 40 nM. + -. 8mM or about 40 nM. In certain embodiments, the histidine buffer comprises L-histidine and L-histidine monohydrochloride monohydrate. In one embodiment, the concentration of L-histidine is between 2 mM. + -. 0.4mM and 25 mM. + -. 5mM, preferably between 4 mM. + -. 0.8mM and 20 mM. + -. 4 mM. In one embodiment, the concentration of L-histidine monohydrochloride monohydrate is from 2 mM. + -. 0.4mM to 25 mM. + -. 5mM, preferably from 4 mM. + -. 0.8mM to 20 mM. + -. 4 mM. In one embodiment, the buffer comprises L-histidine at a concentration of 4.8 mM. + -. 0.96mM and L-histidine monohydrochloride monohydrate at a concentration of 5.2 mM. + -. 1.04 mM. In one embodiment, the buffer comprises histidine at a concentration of 10mM + -2 mM, wherein histidine comprises L-histidine at a concentration of 4.8mM + -0.96 mM and L-histidine monohydrochloride monohydrate at a concentration of 5.2mM + -1.04 mM.
In certain embodiments, the organic co-solvent is a nonionic polymer containing polyoxyethylene moieties. In one embodiment, the organic solvent is a surfactant. In some embodiments, the organic co-solvent is any one or more of polysorbate, poloxamer 188 and polyethylene glycol 3350. In one embodiment, the organic co-solvent is
In one embodiment, the concentration of organic co-solvent is from about 0.01% ± 0.005% to about 1% ± 0.5% "weight/volume" or "w/v", where, for example, 0.1 g/ml-10%, 0.01 g/ml-1%. In certain embodiments, the organic solvent is polysorbate at a concentration of 0.05% ± 0.025% to 0.5% ± 0.25% (w/v). In one embodiment, the organic co-solvent is
In certain embodiments, the stabilizing agent is a sugar. In one embodiment, the sugar is sucrose. In various embodiments, the concentration of the stabilizing agent is from 1% ± 0.2% w/v to 20% ± 4% w/v, from 5% ± 1% w/v to 15% ± 3% w/v, or from 1% ± 0.2% to 10% ± 2% w/v. In one embodiment, the stabilizing agent is sucrose at a concentration of 5% + -1% w/v or about 5% w/v. In another embodiment, the stabilizing agent is sucrose at a concentration of 9% + -1.8% w/v or about 9% w/v. In another embodiment, the stabilizing agent is sucrose at a concentration of 10% + -2% w/v or about 10% w/v.
In one embodiment, the viscosity modifier is an amino acid. In one embodiment, the viscosity modifier is L-proline. In certain embodiments, the concentration of viscosity modifier is 1% + -0.2% to 5% + -1% w/v. In one embodiment, the viscosity modifier is proline at a concentration of 1.5% ± 0.3% or about 1.5%. In one embodiment, the viscosity modifier is proline at a concentration of 3% ± 0.6%, or about 3%.
In certain embodiments, the liquid pharmaceutical formulation has a viscosity of less than or equal to about 15cPoise ± 10% at 25 ℃. In certain embodiments, the viscosity at 25 ℃ is from 1.0cPoise 10% to 20cPoise 10%. In certain embodiments, the liquid pharmaceutical formulation has a viscosity of 15cPoise or less. In certain embodiments, the liquid pharmaceutical formulation has a viscosity of 20cPoise or less. In certain embodiments, the liquid pharmaceutical formulation has a viscosity of 10cPoise or less. In certain embodiments, the viscosity at 25 ℃ is 5cPoise + -10%, 6.0cPoise + -10%, 7.0cPoise + -10%, 7.1cPoise + -10%, 7.2cPoise + -10%, 7.9cPoise + -10%, 8.3cPoise + -10%, 9.0cPoise + -10%, 9.6cPoise + -10%, 10.0cPoise + -10%, 10.6cPoise + -10%, 11.4cPoise + -10%, 11.6cPoise + -10%, 11.8cPoise + -10%, 12.0cPoise + -10%, 13.0cPoise + -10%, 14.0cPoise + -10%, 15.0cPoise + -10%, or 16cPoise + -10%.
In one aspect, a stable liquid pharmaceutical formulation of low viscosity is provided comprising: (i) a human antibody that specifically binds human PD-1 at 5. + -. 0.75mg/ml to 250. + -. 37.5 mg/ml; (ii)0mM to 40 ± 8mM histidine buffer; (iii) 0% to 0.5% ± 0.25% (w/v)
In certain embodiments, a stable, low viscosity liquid pharmaceutical formulation is provided comprising: (i) a human antibody that specifically binds human PD-1 at 5. + -. 0.75mg/ml to 250. + -. 37.5 mg/ml; (ii)0mM to 40 ± 8mM histidine buffer; (iii) 0% to 0.5% ± 0.25% (w/v)
In certain embodiments, a stable, low viscosity liquid pharmaceutical formulation is provided comprising: (i) a human antibody that specifically binds human PD-1 at 5. + -. 0.75mg/ml to 250. + -. 37.5 mg/ml; (ii)0mM to 40 ± 8mM histidine buffer; (iii) 0% to 0.5% ± 0.25% (w/v)
In certain embodiments, the formulation of any of the preceding aspects has a property selected from the group consisting of: (i) the formulations are stable for long term storage at 25 ℃,5 ℃, -20 ℃, -30 ℃ and-80 ℃ as described herein; (ii) as described herein, the formulations are stable to agitation stress; (iii) the formulation is of low viscosity (viscosity less than 20cPoise, preferably less than 15 cPoise); (iii) as described herein, the formulations are stable even if the formulation excipient concentrations vary up to ± 50%; (iv) the formulation is isotonic to physiological conditions; (v) the formulation is stable and compatible with intravenous delivery devices and procedures; and (vi) the formulation is stable for long term storage in glass vials or pre-filled syringes.
In certain embodiments of this aspect, there is provided a stable liquid formulation comprising: (i) a human antibody that specifically binds human PD-1 at 5. + -. 0.75mg/ml to 250. + -. 37.5 mg/ml; (ii)5mM + -1 mM to 20 + -4 mM histidine buffer; (iii) 0.05% ± 0.025% to 0.3% ± 0.15% (w/v)
In one embodiment of this aspect, the stable liquid formulation comprises (i)25 ± 3.75mg/mL of an anti-PD-1 antibody; (ii)10 ± 2mM histidine buffer; (iii) 0.2% ± 0.1% (w/v)
In one embodiment of this aspect, the stable liquid formulation comprises (i)25 ± 3.75mg/mL of an anti-PD-1 antibody; (ii)4.8 mM. + -. 0.96mM L-histidine; (iii)5.2 mM. + -. 1.04mM L-histidine monohydrochloride monohydrate; (iv) 0.2% ± 0.1% (w/v)
In one embodiment of this aspect, the stable liquid formulation comprises (i)50 ± 7.5mg/mL of an anti-PD-1 antibody; (ii)10 ± 2mM histidine buffer; (iii) 0.2% ± 0.1% (w/v)
In one embodiment of this aspect, the stable liquid formulation comprises (i)50 ± 7.5mg/mL of an anti-PD-1 antibody; (ii)4.8 mM. + -. 0.96mM L-histidine; (iii)5.2 mM. + -. 1.04mM L-histidine monohydrochloride monohydrate; (iv) 0.2% ± 0.1% (w/v)
In one embodiment, a stable liquid formulation comprises (i)100 ± 15mg/mL of an anti-PD-1 antibody; (ii)10 ± 2mM histidine buffer; (iii) 0.2% ± 0.1% (w/v) (w/v)
In one embodiment of this aspect, the stable liquid formulation comprises (i)100 ± 15mg/mL of an anti-PD-1 antibody; (ii)4.8 mM. + -. 0.96mM L-histidine; (iii)5.2 mM. + -. 1.04mM L-histidine monohydrochloride monohydrate; (iv) 0.2% ± 0.1% (w/v)
In one embodiment, a stable liquid formulation comprises (i)150 ± 22.5mg/mL of an anti-PD-1 antibody; (ii)10 ± 2mM histidine buffer; (iii) 0.2% ± 0.1% (w/v)
In one embodiment of this aspect, the stable liquid formulation comprises (i)150 ± 22.5mg/mL of an anti-PD-1 antibody; (ii)4.8 mM. + -. 0.96mM L-histidine; (iii)5.2 mM. + -. 1.04mM L-histidine monohydrochloride monohydrate; (iv) 0.2% ± 0.1% (w/v)
In one embodiment of this aspect, the stable liquid formulation comprises (i)175 ± 26.25mg/mL of an anti-PD-1 antibody; (ii)10 ± 2mM histidine buffer; (iii) 0.2% ± 0.1% (w/v)
In one embodiment of this aspect, the stable liquid formulation comprises (i)175 ± 26.25mg/mL of an anti-PD-1 antibody; (ii)4.8 mM. + -. 0.96mM L-histidine; (iii)5.2 mM. + -. 1.04mM L-histidine monohydrochloride monohydrate; (iv) 0.2% ± 0.1% (w/v)
In one embodiment of this aspect, the stable liquid formulation comprises (i)200 ± 30.00mg/mL of the anti-PD-1 antibody; (ii)10 ± 2mM histidine buffer; (iii) 0.2% ± 0.1% (w/v)
In one embodiment of this aspect, the stable liquid formulation comprises (i)200 ± 30.00mg/mL of the anti-PD-1 antibody; (ii)4.8 mM. + -. 0.96mM L-histidine; (iii)5.2 mM. + -. 1.04mM L-histidine monohydrochloride monohydrate; (iv) 0.2% ± 0.1% (w/v)
In one embodiment, after 28 days of storage of the formulation at 45 ℃, > 90% of the antibody is native and > 35% of the antibody is in the form of a main charge. In one embodiment, > 94% of the antibody is native and ≧ 44% of the antibody is the primary charged form after storage of the formulation at 25 ℃ for three months. In one embodiment, > 96% of the antibody is native and > 50% of the antibody is in the primary charged form after storage of the formulation at 5 ℃ for 12 months. In one embodiment, > 96% of the antibodies are native and > 40% of the antibodies are in the primary charged form after storage of the formulation at-20 ℃ for 12 months. In one embodiment, > 96% of the antibodies are native and > 40% of the antibodies are in the primary charged form after storage of the formulation at-30 ℃ for 12 months. In one embodiment, > 96% of the antibodies are native and > 40% of the antibodies are in the primary charged form after storage of the formulation at-80 ℃ for 12 months. In one embodiment, more than 96% of the antibody has native conformation after 12 months of storage at 5 ℃. In one embodiment, at least 97% or more of the antibody has native conformation after 6 months of storage at-80 ℃, -30 ℃ and/or-20 ℃.
In one aspect, the present invention provides a stable liquid formulation comprising: (i) up to 100mg/mL of anti-PD-1 antibody; (ii)2mM ± 0.4mM to 20mM ± 4mM histidine buffer; (iii) up to 20% ± 4% (w/v) sucrose; and (iv) up to 0.2% + -0.1% w/v polysorbate at a pH of 6.0 + -0.3. In one embodiment, the stable liquid formulation comprises 25mg/mL of the anti-PD-1 antibody. In one embodiment, the stable liquid formulation comprises 50mg/mL of the anti-PD-1 antibody. In one embodiment, the stable liquid formulation comprises 75mg/mL of the anti-PD-1 antibody. In one embodiment, the stable liquid formulation comprises 10mM ± 2mM histidine buffer. In one embodiment, the stable liquid formulation comprises 5% sucrose. In one embodiment, the stable liquid formulation comprises 6% sucrose. In one embodiment, the stable liquid formulation comprises 9% sucrose. In one embodiment, the stable liquid formulation comprises 10% sucrose. In one embodiment, the stable liquid formulation comprises 0.1% polysorbate. In one embodiment, the polysorbate is
In one aspect, a stable liquid pharmaceutical formulation of any of the preceding aspects is provided in a container. In one embodiment, the container is a polycarbonate vial. In one embodiment, the container is a glass vial. In one embodiment, the glass vial is a
In one aspect, a kit is provided comprising the stable pharmaceutical composition of any one of the preceding aspects, a container, and instructions. In one embodiment, the container is a glass vial. In one embodiment, the container is a prefilled syringe. In one embodiment, the syringe is a 1mL or 2.25mL long glass syringe equipped with a 27-G thin walled needle, an 4023/50 rubber stopper coated with FLUOTEC, and an FM27 rubber tip cap. In one embodiment, the syringe is a 1mL, 2mL, 3mL, 5mL or 10mL plastic syringe fitted with a needle.
In certain embodiments, the present invention provides a prefilled syringe comprising a stable liquid pharmaceutical formulation comprising: (i) a human antibody that specifically binds human PD-1 at 5. + -. 0.75mg/ml to 250. + -. 37.5 mg/ml; (ii)5mM + -1 mM to 20 + -4 mM histidine buffer; (iii) 0.05% ± 0.025% to 0.3% ± 0.15% (w/v) polysorbate 80; (iv) 1% ± 0.2% to 10% ± 2% (w/v) sucrose; and (v) 1% ± 0.2% to 5% ± 1% proline at pH6.0 ± 0.3, wherein the antibody comprises HCVR/LCVR comprising SEQ ID NO: 1/2; wherein the formulation has an attribute selected from the group consisting of: (i) after 12 months of storage at 5 ℃, not less than 98% of the antibody is in native form; (ii) after 12 months of storage at 5 ℃, not less than 53% of the antibody is a primary charge variant; (iii) after 6 months of storage at 25 ℃, up to 97% of the antibody is in native form; (iv) the formulation is stable to agitation stress, wherein after 120 minutes of agitation stress in a pre-filled syringe, not less than 98% of the antibody is in native form; (v) more than 90% of the antibodies have a molecular weight of 143kDa ± 1 kDa; (vi) the pharmaceutical formulation has a viscosity of less than 20cP, less than 15cP, or less than 10 cP; (vii) after 12 months of storage at 5 ℃, more than 96% of the antibody has native conformation; and (viii) at least 97% or more of the antibody has native conformation after 6 months of storage at-80 ℃, -30 ℃ and/or-20 ℃.
In certain embodiments, the present invention provides a glass vial comprising a stable liquid pharmaceutical formulation comprising: (i) a human antibody that specifically binds human PD-1 at 5. + -. 0.75mg/ml to 250. + -. 37.5 mg/ml; (ii)5mM + -1 mM to 20 + -4 mM histidine buffer; (iii) 0.05% ± 0.025% to 0.3% ± 0.15% (w/v) polysorbate 80; (iv) 1% ± 0.2% to 10% ± 2% (w/v) sucrose; and (v) 1% ± 0.2% to 5% ± 1% proline at pH6.0 ± 0.3, wherein the antibody comprises HCVR/LCVR comprising SEQ ID NO: 1/2; wherein the formulation has an attribute selected from the group consisting of: (i) the formulation is stable to storage and stress in glass vials; (ii) the formulation is stable and compatible for IV delivery devices; (iii) for dilution with standard diluents known in the art (e.g., 0.9% sodium chloride or 5% dextrose), the formulations are chemically and physically stable; (iv) the formulation is stable to IV bags made of glass or polymeric plastics (e.g., polyvinyl chloride, phthalate, polyolefin, or polypropylene); (v) the formulation is compatible with standard infusion pumps (e.g., peristaltic pumps, fluid displacement pumps); (vi) more than or equal to 90 percent of the antibody has the molecular weight of 143kDa +/-1 kDa; (vii) the pharmaceutical formulation has a viscosity of less than 20cP, less than 15cP, or less than 10 cP; (viii) after 12 months of storage at 5 ℃, more than 96% of the antibody has native conformation; and (ix) at least 97% or more of the antibody has native conformation after 6 months of storage at-80 ℃, -30 ℃ and/or-20 ℃.
Other embodiments will become apparent upon reading the following detailed description.
Drawings
FIG. 1 is a table showing the effect of pH on the stability of 150mg/mL mAb1 incubated at 45 ℃ for 28 days.aSE-UPLC and CEX-UPLC "starting materials" results are the average of all formulation starting materials.
Figure 2 shows the storage stability of three formulations F1, F2, and F3, wherein F1 comprises 210mg/mL mAb1, 10mM histidine, and 3% proline, pH 6.0; f2 contained 210mg/mL mAb1, 10mM histidine and 3% sucrose, pH 6.0; f3 contained mAb1 at 210mg/mL, 10mM histidine and 5% sucrose, pH 6.0. Storage stability was measured by the% high molecular weight species (HMW) produced after storage at-80 ℃ (a), -30 ℃ (B) and-20 ℃ (C) for up to 9 months and analyzed by Size Exclusion Chromatography (SEC).
Figure 3 shows the storage stability of three formulations F1, F2, and F3, wherein F1 comprises 150mg/mL mAb1, 10mM histidine, 9% sucrose, and 0.2% polysorbate 80(PS80), ph 6.0; f2 contained 175mg/mL mAb1, 10mM histidine, 3% proline and 0.2% PS80, pH 6.0; f3 contained 175mg/mL mAb1, 10mM histidine, 5% sucrose, 1.5% proline and 0.2% PS80, pH 6.0. Storage stability was measured by the% high molecular weight species (HMW) produced after storage at-80 ℃ (a), -30 ℃ (B) and-20 ℃ (C) for up to 6 months and analyzed by Size Exclusion Chromatography (SEC).
Figure 4 is a table showing the viscosity of 150mg/mL mAb1 with the addition of excipients and viscosity modifiers.
FIG. 5 is a table showing the effect of viscosity modifiers on the stability of 175mg/mL mAb1 incubated at 45 ℃ for 14 days.aThe pH, SE-UPLC and CEX-UPLC "starting materials" results are the average of the starting materials for all formulations.
Detailed Description
Before the present methods are described, it is to be understood that this invention is not limited to the particular methodology and experimental conditions described, as such methods and conditions may vary. It is also to be understood that the terminology used herein is for the purpose of describing particular embodiments only, and is not intended to be limiting, since the scope of the present invention will be limited only by the appended claims.
Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. As used herein, the term "about," when used in reference to a specifically recited value or range of values, means that the value may differ from the recited value by no more than 1%. For example, as used herein, the expression "about 100" includes 99 and 101 and all values therebetween (e.g., 99.1, 99.2, 99.3, 99.4, etc.). Although any methods and materials similar or equivalent to those described herein can be used in the practice or testing of the present invention, the preferred methods and materials are now described. All publications mentioned herein are incorporated by reference in their entirety.
As used herein, the expression "pharmaceutical formulation" refers to a combination of at least one active ingredient (e.g., a small molecule, a macromolecule, a compound, etc., which is capable of exerting a biological effect in a human or non-human animal) and at least one inactive ingredient which, when combined with the active ingredient or one or more additional inactive ingredients, is suitable for therapeutic administration to a human or non-human animal. The term "formulation" as used herein means "pharmaceutical formulation" unless otherwise specifically indicated. The present invention provides pharmaceutical formulations comprising at least one therapeutic polypeptide. According to certain embodiments of the invention, the therapeutic polypeptide is an antibody, or antigen-binding fragment thereof, that specifically binds to a human programmed death-1 (PD-1) protein. More specifically, the invention includes pharmaceutical formulations comprising: (i) a human antibody that specifically binds human PD-1; (ii) a histidine buffer; (iii) an organic co-solvent as a non-ionic surfactant; (iv) a stabilizer for carbohydrates; and optionally, (v) a viscosity modifier that is an amino acid. Specific exemplary ingredients and formulations encompassed by the present invention are described in detail below.
Antibodies that specifically bind to PD-1
The pharmaceutical formulation of the invention may comprise a human antibody, or antigen-binding fragment thereof, that specifically binds human PD-1. As used herein, the term "PD-1" means a human programmed death-1 protein. Antibodies to human PD-1 are described in, for example, U.S. patent/publication nos. 8008449, 8168757, 20110008369, 20130017199, 20130022595, 20150203579, and WO2006121168, WO2009114335, WO2012145493, WO2013014668, WO2009101611, WO 1122015800, EP2262837, and EP 2504028.
As used herein, the term "antibody" generally means an immunoglobulin molecule comprising four polypeptide chains in which two heavy (H) chains and two light (L) chains are interconnected by disulfide bonds, as well as multimers thereof (e.g., IgM); however, immunoglobulin molecules consisting of only heavy chains (i.e., lacking light chains) are also included within the definition of the term "antibody". Each heavy chain comprises a heavy chain variable region (abbreviated herein as HCVR or V)H) And a heavy chain constant region. The heavy chain constant region comprises three domains, CH1, CH2, and
The term "antibody" as used herein is understood to include intact antibody molecules and antigen-binding fragments thereof, unless specifically stated otherwise. As used herein, the term "antigen-binding portion" or "antigen-binding fragment" of an antibody (or simply "antibody portion" or "antibody fragment") refers to one or more fragments of an antibody that retain the ability to specifically bind to human PD-1 or an epitope thereof.
As used herein, "isolated antibody" means an antibody that is substantially free of other antibodies having different antigen specificities (e.g., an isolated antibody that specifically binds human PD-1 is substantially free of antibodies that specifically bind antigens other than human PD-1).
The term "specifically binds" or the like refers to the formation of a relatively stable complex of an antibody or antigen-binding fragment thereof with an antigen under physiological conditions. Specific binding may be through at least about 1X 10-8M or greater. Methods for determining whether two molecules specifically bind are well known in the art and include, for example, equilibrium dialysis, surface plasmon resonance, and the like. However, an isolated antibody that specifically binds human PD-1 may be cross-reactive with other antigens (e.g., PD-1 molecules from other species) (orthologs). In the context of the present invention, a multispecific (e.g., bispecific) antibody that binds human PD-1 and one or more other antigens is considered to "specifically bind" human PD-1. Furthermore, the isolated antibody may be substantially free of other cellular material or chemicals.
Exemplary anti-human PD-1 antibodies that can be included in the pharmaceutical formulations of the present invention are set forth in patent application publications US20150203579 and WO2015112800, the disclosures of which are incorporated by reference in their entirety.
According to certain embodiments of the invention, the anti-human PD-1 antibody or antigen-binding fragment thereof comprises SEQ ID NO: 3, heavy chain complementarity determining region (HCDR)1 of SEQ ID NO: 4 and HCDR2 of SEQ ID NO: HCDR3 of 5. In certain embodiments, the anti-human PD-1 antibody or antigen-binding fragment thereof comprises SEQ ID NO: 1 HCVR.
According to certain embodiments of the invention, anti-human PD-1 or an antigen-binding fragment thereof comprises SEQ ID NO: 6 light chain complementarity determining region (LCDR)1, SEQ ID NO: LCDR2 of 7 and SEQ ID NO: LCDR3 of 8. In certain embodiments, the anti-human PD-1 antibody or antigen-binding fragment thereof comprises SEQ ID NO: LCVR of 2.
According to certain embodiments of the invention, anti-human PD-1 or an antigen-binding fragment thereof comprises a sequence identical to SEQ ID NO: 1 HCVR with 90%, 95%, 98% or 99% sequence identity.
According to certain embodiments of the invention, anti-human PD-1 or an antigen-binding fragment thereof comprises a sequence identical to SEQ ID NO: 2 LCVR with 90%, 95%, 98% or 99% sequence identity.
According to certain embodiments of the invention, anti-human PD-1 or an antigen-binding fragment thereof comprises a HCVR comprising the amino acid sequence of SEQ ID NO: 1.
According to certain embodiments of the invention, anti-human PD-1 or an antigen-binding fragment thereof comprises an LCVR comprising the amino acid sequence of SEQ ID NO: 2.
Sequence identity can be measured by any method known in the art (e.g., GAP, BESTFIT, and BLAST).
The invention also includes formulations comprising an anti-PD-1 antibody, wherein the anti-PD-1 antibody comprises a variant of any of the HCVR, LCVR, and/or CDR amino acid sequences disclosed herein, with one or more conservative amino acid substitutions. For example, the invention includes formulations comprising an anti-PD-1 antibody having a HCVR, LCVR, and/or CDR amino acid sequence with, e.g., 10 or fewer, 8 or fewer, 6 or fewer, 4 or fewer, etc., conservative amino acid substitutions relative to any HCVR, LCVR, and/or CDR amino acid sequence disclosed herein.
In certain embodiments, the anti-PD 1 antibody comprises an Fc region selected from the group consisting of human IgG1, IgG2, IgG3, and IgG4 isotypes.
A non-limiting exemplary antibody used in the examples herein is referred to as "
According to certain embodiments of the invention, anti-human PD-1 or an antigen-binding fragment thereof comprises SEQ ID NO: 9 and SEQ ID NO: 10, light chain.
It is well known in the art that terminal cleavage of amino acids can occur during antibody production (see, e.g., Wang et al 2007, J.Pharma.Sci.96: 1-26). Thus, in certain embodiments, an anti-PD-1 antibody comprises a heavy chain and a light chain, wherein the heavy chain comprises SEQ ID NO: 11. SEQ ID NO: 11 comprises a heavy chain amino acid sequence wherein the C-terminal lysine is substituted with respect to SEQ ID NO: 9 is deleted. In certain embodiments, the formulations of the invention contain about 50%, about 60%, about 70%, about 80%, about 90%, about 95%, about 98%, or more of the anti-PD-1 antibody, wherein the C-terminal lysine is absent.
The amount of antibody or antigen-binding fragment thereof included in a pharmaceutical formulation of the invention can vary depending on the particular properties of the formulation desired and the particular circumstances and purposes for which the formulation is intended to be used. In certain embodiments, the pharmaceutical formulation is a liquid formulation, which may contain from 5 ± 0.75mg/mL to 250 ± 37.5mg/mL of the antibody; 10 + -1.5 mg/mL to 240 + -36 mg/mL antibody; 20 + -3.0 mg/mL to 230 + -34.5 mg/mL antibody; 25 + -3.75 mg/mL to 240 + -36 mg/mL antibody; 50 + -7.5 mg/mL to 230 + -34.5 mg/mL antibody; 60 + -9 mg/mL to 240 + -36 mg/mL antibody; 70 + -10.5 mg/mL to 230 + -34.5 mg/mL antibody; 80 + -12 mg/mL to 220 + -33 mg/mL antibody; 90 + -13.5 mg/mL to 210 + -31.5 mg/mL antibody; 100 + -15 mg/mL to 200 + -30 mg/mL antibody; 110 + -16.5 mg/mL to 190 + -28.5 mg/mL antibody; 120 + -18 mg/mL to 180 + -27 mg/mL antibody; 130 + -19.5 mg/mL to 170 + -25.5 mg/mL antibody; 140 + -21 mg/mL to 160 + -24 mg/mL antibody; 150 + -22.5 mg/mL antibody; or 175. + -. 26.25 mg/mL. For example, a formulation of the invention may comprise about 5 mg/mL; about 10 mg/mL; about 15 mg/mL; about 20 mg/mL; about 25 mg/mL; about 30 mg/mL; about 35 mg/mL; about 40 mg/mL; about 45 mg/mL; about 50 mg/mL; about 55 mg/mL; about 60 mg/mL; about 65 mg/mL; about 70 mg/mL; about 75 mg/mL; about 80 mg/mL; about 85 mg/mL; about 90 mg/mL; about 95 mg/mL; about 100 mg/mL; about 105 mg/mL; about 110 mg/mL; about 115 mg/mL; about 120 mg/mL; about 125 mg/mL; about 130 mg/mL; about 135 mg/mL; about 140 mg/mL; about 145 mg/mL; about 150 mg/mL; about 155 mg/mL; about 160 mg/mL; about 165 mg/mL; about 170 mg/mL; about 175 mg/mL; about 180 mg/mL; about 185 mg/mL; about 190 mg/mL; about 195 mg/mL; about 200 mg/mL; about 205 mg/mL; about 210 mg/mL; about 215 mg/mL; about 220 mg/mL; about 225 mg/mL; about 230 mg/mL; about 235 mg/mL; about 240 mg/mL; about 245 mg/mL; or about 250mg/mL of an antibody or antigen-binding fragment thereof that specifically binds human PD-1.
Excipients and pH
The pharmaceutical formulations of the present invention comprise one or more excipients. As used herein, the term "excipient" refers to any non-therapeutic agent added to a formulation to provide a desired consistency, viscosity, or stabilizing effect.
In certain embodiments, the pharmaceutical formulations of the invention comprise at least one organic co-solvent of a type and amount that stabilizes human PD-1 antibody under crude handling or agitation conditions (e.g., vortexing). In some embodiments, "stable" refers to preventing the formation of aggregated antibodies (on a molar basis) that exceed 3% of the total amount of antibodies during crude treatment. In some embodiments, the coarse rate treatment is vortexing the solution containing the antibody and organic co-solvent for about 60 minutes or about 120 minutes.
In certain embodiments, the organic co-solvent is a nonionic surfactant, such as an alkyl poly (ethylene oxide). Specific nonionic surfactants that may be included in the formulations of the present invention include, for example, polysorbates, such as
The amount of nonionic surfactant included in the pharmaceutical formulations of the present invention can vary depending on the particular properties desired for the formulation and the particular circumstances and purposes for which the formulation is intended to be used. In certain embodiments, the formulation may contain from 0.01% ± 0.005% to 0.5% ± 0.25% surfactant. For example, a formulation of the invention may contain about 0.005%; about 0.01%; about 0.02%; about 0.03%; about 0.04%; about 0.05%; about 0.06%; about 0.07%; about 0.08%; about 0.09%; about 0.1%; about 0.11%; about 0.12%; about 0.13%; about 0.14%; about 0.15%; about 0.16%; about 0.17%; about 0.18%; about 0.19%; about 0.20%; about 0.21%; about 0.22%; about 0.23%; about 0.24%; about 0.25%; about 0.26%; about 0.27%; about 0.28%; about 0.29%; about 0.30%; about 0.35%; about 0.40%; about 0.45%; about 0.46%; about 0.47%; about 0.48%; about 0.49%; about 0.50%; about 0.55%; or about 0.575
The pharmaceutical formulation of the present invention may further comprise one or more stabilizing agents, of a type and in an amount that stabilizes human PD-1 antibody under conditions of heat stress. In some embodiments, "stable" refers to greater than about 91% of the antibody maintaining native conformation when a solution containing the antibody and thermal stabilizer is held at about 45 ℃ for up to about 28 days. In some embodiments, "stable" refers to solutions containing the antibody and thermal stabilizer that aggregate less than about 6% of the antibody when maintained at about 45 ℃ for up to about 28 days. As used herein, "native" refers to the predominant form of an antibody, which is typically an intact monomer of the antibody, by size exclusion. The term "native" also refers to non-aggregated and non-degraded forms of the antibody.
In certain embodiments, where the thermal stabilizer is a sugar, such as sucrose, the amount of sugar included in the formulation may vary depending on the particular situation in which the formulation is used and the intended purpose. In certain embodiments, the formulation may contain from about 1% to about 15% sugar; from about 2% to about 14% sugar; from about 3% to about 13% sugar; from about 4% to about 12% sugar; from about 5% to about 12% sugar; from about 6% to about 11% sugar; from about 7% to about 10% sugar; from about 8% to about 11% sugar; or from about 9% to about 11% sugar. For example, a pharmaceutical formulation of the present invention may comprise 4% ± 0.8%; 5% +/-1%; 6% +/-1.2%; 7% +/-1.4%; 8% +/-1.6%; 9% +/-1.8%; 10% +/-2%; 11% +/-2.2%; 12% ± 2.4%; 13% ± 2.6%; or about 14% ± 2.8% sugar (e.g., sucrose).
The pharmaceutical formulations of the present invention may also contain a buffer or buffer system for maintaining a stable pH and helping to stabilize human PD-1 antibodies. The term "buffer" as used herein means a pharmaceutically acceptable buffer that maintains a stable pH of a solution or resists changes in the pH of a solution. In a preferred embodiment, the buffering agent comprises histidine. In the context of the present disclosure, a "histidine buffer" or "buffer comprising histidine" is a buffer comprising the amino acid histidine. Examples of histidine buffers include histidine chloride, histidine acetate, histidine phosphate and histidine sulfate. In a preferred embodiment, the histidine buffer is prepared by dissolving L-histidine and L-histidine hydrochloride (e.g., as a monohydrate) in defined amounts and ratios. In one embodiment, the histidine buffer is prepared by titration of L-histidine (free base, solid) with dilute hydrochloric acid. Throughout the present disclosure, the term "histidine" is used interchangeably with "histidine buffer". In some embodiments, "stable" refers to less than 4.5% ± 0.5% aggregation of the antibody when a solution containing the antibody and buffer is maintained at about 45 ℃ for up to about 28 days. In some embodiments, "stable" refers to less than 3% ± 0.5% or less than 2.5% ± 0.5% aggregation of the antibody when a solution containing the antibody and buffer is maintained at about 37 ℃ for up to about 28 days. In some embodiments, "stable" refers to a solution containing an antibody and a buffer in which at least 93% ± 0.5% or at least 94% ± 0.5% of the antibody is in its native conformation, as determined by size exclusion chromatography, when the solution is maintained at about 45 ℃ for up to about 28 days. In some embodiments, "stable" refers to a solution containing an antibody and a buffer in which at least 94% ± 0.5% or at least 95% ± 0.5% of the antibody is in its native conformation, as determined by size exclusion chromatography, when the solution is maintained at about 37 ℃ for up to about 28 days. "native" or "native conformation" refers to the fraction of antibody that is not aggregated or degraded. This is typically determined by assays that measure the relative size of the antibody entities, such as size exclusion chromatography assays. The non-aggregated and non-degraded antibodies elute in a fraction equivalent to the native antibodies, and are usually the major elution fraction. Aggregated antibodies elute in a fraction of larger size than native antibodies. The degraded antibody is eluted in a fraction of smaller size than the native antibody.
In some embodiments, "stable" refers to a solution containing an antibody and a buffer in which at least 35% ± 0.5% of the antibody is in its primary charge form when maintained at about 45 ℃ for up to about 28 days, as determined by cation exchange chromatography. In some embodiments, "stable" refers to a solution containing an antibody and a buffer in which at least 46% ± 0.5% or at least 39% ± 0.5% of the antibody is in its primary charge form when maintained at about 37 ℃ for up to about 28 days, as determined by cation exchange chromatography. "major charge" or "major charge form" refers to the fraction of antibody that elutes from the ion exchange resin into a major peak, typically a more "basic" peak on one side of the major peak and a more "acidic" peak on the other side.
The pharmaceutical formulations of the present invention may have a pH of about 5.2 to about 6.4. For example, a formulation of the present invention may have a ph of about 5.5; about 5.6; about 5.7; about 5.8; about 5.9; about 6.0; about 6.1; about 6.2; about 6.3; about 6.4; or a pH of about 6.5. In some embodiments, the pH is 6.0 ± 0.4; 6.0 plus or minus 0.3; 6.0 plus or minus 0.2; 6.0 plus or minus 0.1; about 6.0; or 6.0.
In some embodiments, the buffer or buffer system comprises at least one buffer having a buffering range that completely or partially overlaps with the range of ph 5.5-7.4. In certain embodiments, the buffer comprises histidine buffer. In certain embodiments, histidine buffer is present at a concentration of 5mM ± 1mM to 15mM ± 3 mM; 6mM + -1.2 mM to 14mM + -2.8 mM; 7mM + -1.4 mM to 13mM + -2.6 mM; 8mM + -1.6 mM to 12mM + -2.4 mM; 9 mM ± 1.8mM to 11mM ± 2.2 mM; 10mM +/-2 mM; or about 10 mM. In certain embodiments, the buffer system comprises 10 m. + -. 2mM histidine, pH 6.0. + -. 0.3. In a preferred embodiment, the histidine buffer comprises L-histidine and L-histidine monohydrochloride monohydrate. In one embodiment, the histidine buffer comprises L-histidine at a concentration of 4.8 mM. + -. 0.96 mM. In one embodiment, the histidine buffer comprises L-histidine monohydrochloride monohydrate at a concentration of 5.2mM ± 1.04 mM. In one embodiment, the histidine buffer comprises L-histidine at a concentration of 4.8 mM. + -. 0.96mM and L-histidine monohydrochloride monohydrate at a concentration of 5.2 mM. + -. 1.04 mM.
The pharmaceutical formulations of the present invention may also contain one or more excipients for maintaining a reduced viscosity or reducing the viscosity of formulations containing high concentrations of anti-PD-1 antibody drug substance (e.g., typically ≧ 150mg/ml antibody). In certain embodiments, the viscosity modifier is an amino acid. In one embodiment, the amino acid is proline. In one embodiment, the pharmaceutical preparation of the invention contains proline, preferably L-proline, at a concentration of 1%, 1.5%, 2%, 2.5%, 3%, 3.5%, 4%, 4.5% or 5%. Throughout this disclosure, the term "proline" is used interchangeably with "L-proline". In some embodiments, the formulation comprises proline in an amount sufficient to maintain the viscosity of the liquid formulation at less than 20 ± 3cPoise, less than 15 ± 2.25cPoise, or less than 11 ± 1.65 cPoise. In some embodiments, the formulation comprises proline in an amount sufficient to maintain a viscosity at or below 15 ± 2.25 cPoise. In certain embodiments, the formulation may contain from about 1% to about 5% proline; about 2% to about 4% proline; or about 3% proline. For example, a pharmaceutical formulation of the present invention may comprise 1% ± 0.2%; 1.5% ± 0.3%; 2% ± 0.4%; 2.5% ± 0.5%; 3% +/-0.6%; 3.5% + -0.7%; 4% +/-0.8%; 4.5% ± 0.9%; or about 5% ± 1% proline.
During antibody purification, it may be necessary or necessary to exchange one buffer for another to obtain the appropriate excipient concentration, antibody concentration, pH, etc. Buffer exchange can be accomplished, for example, by ultrafiltration/diafiltration (UF/DF), e.g., using a semi-permeable tangential flow filter membrane. However, it is possible to induce the Gibbs-Donnan effect using these techniques [ Bolton et al, 2011, Biotechnol. prog.27(1): 140-. The enhancement of positive charge on the product side of the membrane during protein concentration is electrically offset by the preferential movement of cations to the opposite side of the membrane. A potential consequence of this phenomenon is that the final concentration of certain components (e.g. histidine, L-proline, etc.) may be lower than the expected target concentration of these components due to the electrostatic repulsion of the positively charged antibody protein by the positively charged diafiltration buffer excipients in the UF/DF step. Thus, the present invention includes formulations wherein, for example, the concentration of histidine and/or L-proline differs from the amounts or ranges described herein due to the Gibbs-Donnan effect.
Volume exclusion describes the behavior of a high concentration sample, in which a significant portion of the total volume of the solution is occupied by a solute, particularly large molecules such as proteins, from which solvent is removed. This then reduces the total volume of available solvent in which other solutes are to be dissolved, which can result in unequal partitioning across the ultrafiltration membrane. Thus, the invention includes formulations wherein, for example, the concentration of histidine and/or L-proline may differ from the amounts or ranges described herein due to volume exclusion effects.
During the process of preparing the formulation of the present invention, changes in formulation composition may occur. Such changes may include the concentration of the active ingredient, the concentration of excipients, and/or the pH of the formulation. Since any change in these parameters may potentially affect the stability or efficacy of a pharmaceutical product, a validated acceptable range (PAR) study was conducted to assess whether a change in the composition within a defined range would affect the stability or efficacy of the antibody. Thus, the invention includes formulations containing anti-PD-1 antibodies that are stable and maintain potency over excipient concentration variations of up to 50%. For example, included herein are anti-PD-1 antibody formulations wherein the stability and efficacy of the formulation is not affected by ± 10%, ± 20%, ± 30%, ± 40% or ± 50% variation in antibody, sucrose, histidine buffer and/or polysorbate concentration.
Stability and viscosity of pharmaceutical formulations
The pharmaceutical formulations of the present invention generally exhibit a high level of stability. As used herein, the term "stable" with respect to a pharmaceutical formulation refers to an antibody in a pharmaceutical formulation that retains an acceptable degree of chemical structure or biological function after storage under defined conditions. The formulation may be stable even if the antibody contained therein does not maintain 100% of its chemical structure or biological function after storage for a defined period of time. In certain instances, an antibody structure or function that maintains about 90%, about 95%, about 96%, about 97%, about 98%, or about 99% after storage over a defined period of time may be considered "stable".
Stability can be measured inter alia by determining the percentage of native antibody retained in the formulation after storage at a defined temperature for a defined time. The percentage of native antibody can be determined, inter alia, by size exclusion chromatography (e.g., size exclusion ultra high performance liquid chromatography [ SE-UPLC ]), where native means non-aggregated and non-degraded. The phrase "acceptable stability" as used herein means that at least 90% of the antibody in its native form can be detected in the formulation after storage at a given temperature for a defined time. In certain embodiments, at least about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% of the antibody in its native form can be detected in the formulation after storage at a defined temperature for a defined time. The defined time after which stability is measured can be at least 14 days, at least 28 days, at least 1 month, at least 2 months, at least 3 months, at least 4 months, at least 5 months, at least 6 months, at least 7 months, at least 8 months, at least 9 months, at least 10 months, at least 11 months, at least 12 months, at least 18 months, at least 24 months, or longer. The pharmaceutical formulation may be stored at a defined temperature when evaluating stability, which may be any temperature from about-80 ℃ to about 45 ℃, e.g., at about-80 ℃, about-30 ℃, about-20 ℃, about 0 ℃, about 4 ℃ to 8 ℃, about 5 ℃, about 25 ℃, about 35 ℃, about 37 ℃, or about 45 ℃. For example, a pharmaceutical formulation may be considered stable if, after 6 months of storage at 5 ℃, SE-UPLC detects greater than about 95%, 96%, 97%, or 98% of the native antibody. A pharmaceutical formulation may also be considered stable if more than about 95%, 96%, 97% or 98% of the native antibody is detected by SE-UPLC after 6 months of storage at 25 ℃. A pharmaceutical formulation may also be considered stable if more than about 89%, 90%, 91%, 92%, 93%, 94%, 95% or 96% of the native antibody is detected by SE-UPLC after 28 days of storage at 45 ℃. The pharmaceutical formulation may also be considered stable if more than about 96%, 97% or 98% of the native antibody is detected by SE-UPLC after 12 months of storage at-20 ℃. The pharmaceutical formulation may also be considered stable if more than about 96%, 97% or 98% of the native antibody is detected by SE-UPLC after 12 months of storage at-30 ℃. The pharmaceutical formulation may also be considered stable if more than about 96%, 97% or 98% of the native antibody is detected by SE-UPLC after 12 months of storage at-80 ℃.
Stability can be measured, inter alia, by determining the percentage of antibody that forms aggregates in the formulation after storage for a defined time at a defined temperature, wherein stability is inversely proportional to the percentage of aggregates formed. The percentage of aggregated antibody can be determined, inter alia, by size exclusion chromatography (e.g., size exclusion ultra performance liquid chromatography [ SE-UPLC ]). The phrase "acceptable degree of stability" as used herein refers to the detection in a formulation that at most 5% of the antibody is in an aggregated form (also referred to as high molecular weight-HMW-form) after storage at a given temperature for a defined time. In certain embodiments, an acceptable degree of stability refers to detection in the formulation of at most about 5%, 4%, 3%, 2%, 1%, 0.5%, or 0.1% of the antibody in aggregated form after storage for a defined period of time at a given temperature. Stability is measured after a defined time, which may be measured after at least 2 weeks, at least 28 days, at least 1 month, at least 2 months, at least 3 months, at least 4 months, at least 5 months, at least 6 months, at least 7 months, at least 8 months, at least 9 months, at least 10 months, at least 11 months, at least 12 months, at least 18 months, at least 24 months or longer. The pharmaceutical formulation may be stored at any temperature from about-80 ℃ to about 45 ℃ when evaluating stability, for example, at about-80 ℃, about-30 ℃, about-20 ℃, about 0 ℃, about 4 ℃ to 8 ℃, about 5 ℃, about 25 ℃, about 35 ℃, about 37 ℃, or about 45 ℃. For example, a pharmaceutical formulation may be considered stable if less than about 2%, 1%, 0.5% or 0.1% of the antibody is detected in aggregated form after 12 months of storage at 5 ℃. A pharmaceutical formulation may also be considered stable if less than about 4%, 3%, 2%, 1%, 0.5% or 0.1% of the antibody is detected in aggregated form after storage for three months at 25 ℃. A pharmaceutical formulation may also be considered stable if less than about 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1% or 0.5% of the antibody is detected in aggregated form after 28 days of storage at 45 ℃. A pharmaceutical formulation may also be considered stable if after three months of storage at-20 ℃, -30 ℃ or-80 ℃ less than about 3%, 2%, 1%, 0.5% or 0.1% of the antibody is detected in aggregated form.
Stability can be measured, inter alia, by determining the percentage of antibody that migrates during ion exchange in a fraction that is more acidic ("acidic form") than the main fraction of antibody ("primary charged form"), where stability is inversely proportional to the acidic form of the antibody fraction. While not wishing to be bound by theory, Deamidation of an antibody may result in the antibody becoming more negatively charged and thus more acidic relative to non-deamidated antibodies (see, e.g., Robinson, n., Protein Deamidation, PNAS, 16.4.2002, 99(8): 5283-5288). The percentage of "acidified" antibody can be determined, inter alia, by ion exchange chromatography (e.g., cation exchange ultra performance liquid chromatography [ CEX-UPLC ]). The phrase "acceptable degree of stability" as used herein means that up to 45% of the antibody is detected in the formulation as being in a more acidic form after storage at a defined temperature for a defined time. In certain embodiments, an acceptable degree of stability refers to at most about 40%, 35%, 30%, 25%, 20%, 15%, 10%, 5%, 4%, 3%, 2%, 1%, 0.5%, or 0.1% of the antibody detected in the formulation after storage at a given temperature for a defined time is in an acidic form. In one embodiment, an acceptable degree of stability refers to less than 30%, 25%, 20%, 15%, 10%, 5%, 4%, 3%, 2%, 1%, 0.5%, or 0.1% of the antibody detected in the formulation after storage at a given temperature for a defined time is in an acidic form. Stability is measured after a defined time, which may be measured after at least 2 weeks, at least 28 days, at least 1 month, at least 2 months, at least 3 months, at least 4 months, at least 5 months, at least 6 months, at least 7 months, at least 8 months, at least 9 months, at least 10 months, at least 11 months, at least 12 months, at least 18 months, at least 24 months or longer. The pharmaceutical formulation may be stored at any temperature from about-80 ℃ to about 45 ℃ when evaluating stability, e.g., at about-80 ℃, about-30 ℃, about-20 ℃, about 0 ℃, about 4 ℃ to 8 ℃, about 5 ℃, about 25 ℃, or about 45 ℃. For example, a pharmaceutical formulation may be considered stable if less than about 30%, 29%, 28%, 27%, 26%, 25%, 24%, 23%, 22%, 21%, 20%, 19%, 18%, 17%, 16%, 15%, 14%, 13%, 12%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, 0.5%, or 0.1% of the antibody is in a more acidic form after storage at-80 ℃, -30 ℃, or-20 ℃ for three months. A pharmaceutical formulation may also be considered stable if less than about 32%, 31%, 30%, 29%, 28%, 27%, 26%, 25%, 24%, 23%, 22%, 21%, 20%, 19%, 18%, 17%, 16%, 15%, 14%, 13%, 12%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, 0.5% or 0.1% of the antibody is in a more acidic form after six months of storage at 5 ℃. A pharmaceutical formulation may also be considered stable if less than about 43%, 42%, 41%, 40%, 39%, 38%, 37%, 36%, 35%, 34%, 33%, 32%, 31%, 30%, 29%, 28%, 27%, 26%, 25%, 24%, 23%, 22%, 21%, 20%, 19%, 18%, 17%, 16%, 15%, 14%, 13%, 12%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, 0.5% or 0.1% of the antibody is in a more acidic form after six months of storage at 25 ℃. A pharmaceutical formulation may also be considered stable if after 28 days of storage at 45 ℃, less than about 49%, 48%, 47%, 46%, 45%, 44%, 43%, 42%, 41%, 40%, 39%, 38%, 37%, 36%, 35%, 34%, 33%, 32%, 31%, 30%, 29%, 28%, 27%, 26%, 25%, 24%, 23%, 22%, 21%, 20%, 19%, 18%, 17%, 16%, 15%, 14%, 13%, 12%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, 0.5% or 0.1% of the antibodies are detected as being in a more acidic form.
Other methods may be used to assess the stability of the formulations of the invention, for example using Differential Scanning Calorimetry (DSC) to determine thermal stability, controlled agitation to determine mechanical stability, and absorbance at about 350nm or about 405nm to determine solution turbidity. For example, the OD of a formulation when compared to 0 if stored at about 5 ℃ to about 25 ℃ for 6 months or longer405In contrast, OD of the preparation405A change of less than about 0.05 (e.g., 0.04, 0.03, 0.02, 0.01, or less) can be considered stable.
Stability can also be assessed using measurements of the biological activity or binding affinity of the antibody to its target. For example, a formulation of the invention may be considered stable if the anti-PD-1 antibody comprised in the formulation binds to PD-1 with an affinity that is at least 90%, 95% or more of the binding affinity of said antibody prior to storage after storage for a defined time (e.g., 1 to 12 months), e.g., at 5 ℃, 25 ℃, 45 ℃, etc. Binding affinity can be determined by, for example, ELISA or surface plasmon resonance. Biological activity can be determined by a PD-1 activity assay, e.g., contacting a cell expressing PD-1 with a formulation comprising an anti-PD-1 antibody. Binding of the antibody to such cells can be measured directly, for example, by FACS analysis. Alternatively, the downstream activity of the PD-1 system can be measured in the presence of an antibody and compared to the activity of the PD-1 system in the absence of the antibody. In some embodiments, PD-1 may be endogenous to the cell. In other embodiments, PD-1 may be ectopically expressed in the cell.
Additional methods for assessing antibody stability in a formulation are demonstrated in the examples set forth below.
In certain embodiments, the liquid pharmaceutical formulations of the present invention may exhibit low to moderate levels of viscosity.As used herein, "viscosity" may be "kinematic viscosity" or "absolute viscosity". "kinematic viscosity" is a measure of the resistance of a fluid to flow under the influence of gravity. When two fluids of the same volume are placed in the same capillary viscometer and allowed to flow by gravity, the viscous fluid takes longer to flow through the capillary than the less viscous fluid. For example, if one fluid takes 200 seconds to complete its flow and another fluid takes 400 seconds, the viscosity of the second fluid is twice that of the first on a kinematic viscosity scale. "absolute viscosity" is sometimes referred to as dynamic viscosity or simple viscosity, which is the product of kinematic viscosity and fluid density (absolute viscosity ═ kinematic viscosity x density). Kinematic viscosity dimension L2Where L is the length and T is the time. Typically, kinematic viscosities are expressed in centistokes (cSt). The SI unit of the kinematic viscosity is mm2And/s, i.e., 1 cSt. Absolute viscosity is expressed in units of centipoise (cP). The SI unit of absolute viscosity is milliPascal-seconds (mPa-s), where 1cP is 1 mPa-s.
As used herein, with respect to the fluid formulations of the present invention, a low level viscosity will exhibit an absolute viscosity of less than about 20cpoise (cp). For example, a fluid formulation of the present invention is considered to have a "low viscosity" if the formulation exhibits an absolute viscosity of about 20cP, about 19cP, about 18cP, about 15cP, about 12cP, about 10cP, about 9cP, about 8cP, or less, when measured using standard viscosity measurement techniques. As used herein, with respect to the fluid formulations of the present invention, a moderate level of viscosity will exhibit an absolute viscosity of between about 35cP to about 20 cP. For example, a fluid formulation of the present invention is considered to have a "medium viscosity" if the formulation exhibits an absolute viscosity of about 34cP, about 33cP, about 32cP, about 31cP, about 30cP, about 29cP, about 28cP, about 27cP, about 26cP, about 25cP, about 24cP, about 23cP, about 22cP, about 21cP, about 20cP, about 19cP, 18cP, about 17cP, about 16cP, or about 15.1cP, as measured using standard viscosity measurement techniques.
As shown in the examples below, the present inventors have surprisingly found that low viscosity liquid formulations comprising high concentrations of anti-human PD-1 antibody (e.g., from about 50mg/mL up to 250mg/mL) can be obtained by formulating the antibody with about 1% to about 5% proline and about 5% sucrose. These formulations are stable to stress during processing and storage at temperatures of 45 ℃ to-80 ℃ (shown herein) and have low viscosities (ranging from 7 to 15 cP).
Exemplary formulations
According to one aspect of the invention, the pharmaceutical formulation is a stable, low viscosity, generally physiologically isotonic liquid formulation comprising: (i) a human antibody that specifically binds human PD-1 (e.g., H4H7798N) at a concentration of up to 250mg/mL ± 45 mg/mL; (ii) a histidine buffer system which provides sufficient buffering at about ph6.0 ± 0.3; (iii) an organic cosolvent, which protects the structural integrity of the antibody; (iv) as a heat stabilizer for sugars; (iv) as a viscosity modifier for amino acids, for maintaining an easily injectable viscosity in a volume convenient for subcutaneous administration.
According to one embodiment, the stable low viscosity pharmaceutical formulation comprises: (i) a human IgG4 antibody that specifically binds human PD-1 at a concentration of up to 200mg/mL ± 30mg/mL, comprising the amino acid sequence of SEQ ID NO: 3, HCDR1 of SEQ ID NO: 4 HCDR2, SEQ ID NO: 5 HCDR3, SEQ ID NO: LCDR1 of SEQ ID NO: LCDR2 of 7 and SEQ ID NO: LCDR3 of 8; (ii)10mM + -2 mM histidine buffer buffered at pH6.0 + -0.3; (iii) 0.2% w/v ± 0.1% w/
According to one embodiment, the stable low viscosity pharmaceutical formulation comprises: (i) a human IgG4 antibody that specifically binds to human PD-1 at a concentration of 175mg/mL ± 26.25mg/mL, comprising the amino acid sequence of SEQ ID NO: 3, HCDR1 of SEQ ID NO: 4 HCDR2, SEQ ID NO: 5 HCDR3, SEQ ID NO: LCDR1 of SEQ ID NO: LCDR2 of 7 and SEQ ID NO: LCDR3 of 8; (ii)10mM + -2 mM histidine buffer buffered at pH6.0 + -0.3; (iii) 0.2% w/v ± 0.1% w/
According to one embodiment, the stable low viscosity pharmaceutical formulation comprises: (i) a human IgG4 antibody that specifically binds to human PD-1 at a concentration of 150mg/mL ± 22.5mg/mL, comprising the amino acid sequence of SEQ ID NO: 3, HCDR1 of SEQ ID NO: 4 HCDR2, SEQ ID NO: 5 HCDR3, SEQ ID NO: LCDR1 of SEQ ID NO: LCDR2 of 7 and SEQ ID NO: LCDR3 of 8; (ii)10mM + -2 mM histidine buffer buffered at pH6.0 + -0.3; (iii) 0.2% w/v ± 0.1% w/
According to one embodiment, the stable low viscosity pharmaceutical formulation comprises: (i) a human IgG4 antibody that specifically binds to human PD-1 at a concentration of 100mg/mL ± 15mg/mL, comprising the amino acid sequence of SEQ ID NO: 3, HCDR1 of SEQ ID NO: 4 HCDR2, SEQ ID NO: 5 HCDR3, SEQ ID NO: LCDR1 of SEQ ID NO: LCDR2 of 7 and SEQ ID NO: LCDR3 of 8; (ii)10mM + -2 mM histidine buffer buffered at pH6.0 + -0.3; (iii) 5% w/v ± 1% w/v sucrose; (iv) 0.2% w/v ± 0.1% w/
According to one embodiment, the stable low viscosity pharmaceutical formulation comprises: (i) a human IgG4 antibody that specifically binds to human PD-1 at a concentration of 50mg/mL ± 7.5mg/mL, comprising the amino acid sequence of SEQ ID NO: 3, HCDR1 of SEQ ID NO: 4 HCDR2, SEQ ID NO: 5 HCDR3, SEQ ID NO: LCDR1 of SEQ ID NO: LCDR2 of 7 and SEQ ID NO: LCDR3 of 8; (ii)10mM + -2 mM histidine buffer buffered at pH6.0 + -0.3; (iii) 5% w/v ± 1% w/v sucrose; (iv) 0.2% w/v ± 0.1
According to one embodiment, the stable low viscosity pharmaceutical formulation comprises: (i) a human IgG4 antibody that specifically binds to human PD-1 at a concentration of 25mg/mL ± 3.75mg/mL, comprising the amino acid sequence of SEQ ID NO: 3, HCDR1 of SEQ ID NO: 4 HCDR2, SEQ ID NO: 5 HCDR3, SEQ ID NO: LCDR1 of SEQ ID NO: LCDR2 of 7 and SEQ ID NO: LCDR3 of 8; (ii)10mM + -2 mM histidine buffer buffered at pH6.0 + -0.3; (iii) 5% w/v ± 1% w/v sucrose; (iv) 0.2% w/v ± 0.1
Other non-limiting examples of pharmaceutical formulations encompassed by the present invention are set forth elsewhere herein, including the working examples given below.
Container and method of application
The pharmaceutical formulations of the present invention may be contained in any container suitable for storing pharmaceutical and other therapeutic compositions. For example, the pharmaceutical preparation may be contained in a sealed and sterilized plastic or glass container having a defined volume, such as a vial, ampoule, syringe, cartridge or bottle. Different types of vials can be used to contain the formulations of the present invention, including, for example, clear and opaque (e.g., amber) glass or plastic vials. Likewise, any type of syringe may be used to contain or administer the pharmaceutical formulations of the present invention.
The pharmaceutical formulations of the present invention may be contained in a "conventional tungsten" syringe or a "low tungsten" syringe. As understood by those of ordinary skill in the art, the process of making glass syringes typically involves the use of a hot tungsten rod which is used to pierce the glass, thereby forming a hole from which the liquid can be drawn and expelled from the syringe. This process results in the deposition of trace amounts of tungsten on the inner surface of the injector. Subsequent washing and other processing steps may be used to reduce the amount of tungsten in the syringe. As used herein, the term "conventional tungsten" means that the syringe contains greater than or equal to 500 parts per billion (500ppb) of tungsten. The term "low tungsten" means that the syringe contains less than 500ppb tungsten. For example, a low tungsten injector according to the present invention may contain less than about 490, 480, 470, 460, 450, 440, 430, 420, 410, 390, 350, 300, 250, 200, 150, 100, 90, 80, 70, 60, 50, 40, 30, 20, 10, or less ppb tungsten.
The rubber stopper used in the syringe and the rubber stopper used to close the opening of the vial may be coated to prevent contamination of the contents of the syringe or vial or to maintain stability thereof. Thus, according to certain embodiments, the pharmaceutical formulations of the present invention may be contained within a syringe comprising a coated stopper, or within a vial sealed with a coated rubber stopper. For example, the plug or bung may be coated with a fluorocarbon film. Examples of coated stoppers or stoppers suitable for vials and syringes containing the pharmaceutical formulations of the present invention are described in, for example, U.S. patent nos. 4,997,423; 5,908,686, respectively; 6,286,699, respectively; 6,645,635, respectively; and 7,226,554, the contents of which are incorporated herein by reference in their entirety. Can be used forCertain exemplary coated rubber stoppers and stoppers for use in the context of the present invention are commercially available from West Pharmaceutical Services, Inc. (Lionville, Pa.), under the trade name West Pharmaceutical Services, Inc
According to certain embodiments of the invention, the pharmaceutical formulation may be contained in a low tungsten syringe comprising a fluorocarbon-coated stopper.
The pharmaceutical formulations can be administered to the patient by parenteral routes, such as injection (e.g., subcutaneous, intravenous, intramuscular, intraperitoneal, etc.) or transdermal, mucosal, nasal, pulmonary, or oral administration. Many reusable pen or autoinjector delivery devices may be used to subcutaneously deliver the pharmaceutical formulations of the present invention. Examples include, but are not limited to, AUTOPENTM(Owen Mumford,Inc.,Woodstock,UK)、DISETRONICTMPen (Disetronic Medical Systems, Bergdorf, Switzerland),
The use of microinfusion devices to deliver the pharmaceutical formulations of the present invention is also contemplated herein. As used herein, the term "microinfusion device" refers to a subcutaneous delivery device designed to slowly administer a large volume (e.g., up to about 2.5mL or more) of a therapeutic formulation over an extended period of time (e.g., about 10, 15, 20, 25, 30 minutes, or more). See, e.g., U.S.6,629,949; US 6,659,982; and Meehan et al, J.controlled Release 46: 107-. Microinfusion devices are particularly useful for delivering high concentrations (e.g., about 100, 125, 150, 175, 200 or more mg/mL) or large doses of therapeutic proteins contained in viscous solutions.
In certain embodiments, the stable liquid pharmaceutical formulation of any preceding aspect is contained in a sterile glass vial and administered as an IV infusion.
In one embodiment, the container is a
In one embodiment, a liquid pharmaceutical formulation of the invention comprising about 25mg/mL or 50mg/mL mAb1 is administered intravenously, and may be contained in a glass vial.
In certain embodiments, the present invention provides an autoinjector comprising any of the liquid formulations described herein. In some embodiments, the present invention provides an autoinjector comprising a stable liquid formulation comprising about 50mg/mL, about 100mg/mL, about 150mg/mL, or about 175mg/mL mAb1, about 10mM histidine, about pH6.0, about 5% sucrose, about 1.5% proline and about 0.2
In certain embodiments, the present invention provides a prefilled syringe comprising any of the liquid formulations described herein. In some embodiments, the invention provides a prefilled syringe comprising a stable liquid formulation comprising about 50mg/mL, about 100mg/mL, about 150mg/mL, or about 175mg/mL mAb1, about 10mM histidine, about ph0.6, about 5% sucrose, about 1.5% proline, and about 0.2
In one embodiment, a liquid pharmaceutical formulation containing about 175mg/mL ± 26.25mg/mL mAb1 is administered in a pre-filled syringe in a volume of about up to 2 mL. In certain embodiments, the syringe is a 1mL or 2.25mL long glass syringe fitted with a 27 gauge thin-walled needle, a fluorocarbon coated rubber stopper, and a rubber needle shield. In one embodiment, the syringe is an OMPI 1mL long glass syringe fitted with a 27 gauge needle, FM27 rubber needle shield and coating
In one embodiment, a liquid pharmaceutical formulation containing about 150mg/mL ± 22.5mg/mL of an anti-PD-1 antibody is administered in a volume of about up to 2mL in a pre-filled syringe. In one embodiment, the syringe is a 1mL or 2.25mL long glass syringe fitted with a 27 gauge thin-walled needle, a fluorocarbon coated rubber stopper, and a rubber needle shield. In one embodiment, the syringe is an OMPI 1mL long glass syringe fitted with a 27 gauge needle, FM27 rubber needle shield and coating
Therapeutic use of pharmaceutical formulations
The pharmaceutical formulations of the present invention are particularly useful for treating, preventing or ameliorating any disease or condition associated with PD-1 activity, including diseases or conditions mediated by PD-1. Exemplary, non-limiting diseases and conditions that may be treated or prevented by administration of the pharmaceutical formulations of the present invention include viral infections, autoimmune diseases, and various cancers, such as brain cancer, lung cancer, prostate cancer, colorectal cancer, head and neck cancer, skin cancer, various blood cancers, and endometrial cancer.
Examples
The following examples are put forth so as to provide those of ordinary skill in the art with a complete disclosure and description of how to make and use the methods and compositions of the present invention, and are not intended to limit the scope of what the inventors regard as their invention. Efforts have been made to ensure accuracy with respect to numbers used (e.g., amounts, temperatures, etc.) but some experimental error and deviation should be accounted for. Unless otherwise indicated, parts are mole parts, molecular weight is average molecular weight, temperature is in degrees Celsius, and pressure is at or near atmospheric.
- 上一篇：一种医用注射器针头装配设备
- 下一篇：具有与吸入器一起使用的颗粒的容器