Stable antibody formulations comprising finotumab

Abstract

The present invention provides stable pharmaceutical formulations comprising a human antibody that specifically binds to human Lymphocyte-Activation Gene-3 (LAG-3). In certain embodiments, the formulations comprise, in addition to the anti-LAG-3 antibody, a buffer, an amino acid, a non-ionic surfactant, and a sugar. The pharmaceutical formulations of the present invention exhibit a considerable degree of antibody stability upon stress and storage.

Description

Technical Field

[0001] This disclosure relates to the field of therapeutic antibody formulations. More specifically, this disclosure relates to the field of pharmaceutical formulations comprising human antibodies that specifically bind to the human lymphocyte activation gene-3 (LAG-3) protein.

[0002] sequence list An official copy of the sequence list was filed with the Patent Center along with this specification. The contents of the electronic sequence list (11396WO01_Sequence_Listing_ST26.xml; size: 12288 bytes; and creation date: October 29, 2024) are part of this specification and are incorporated herein by reference in their entirety. Background Technology

[0003] Therapeutic macromolecules ( For example Antibodies must be formulated in a way that not only makes the molecules suitable for administration to patients but also maintains their stability during storage and subsequent use. For example, unless the solution is properly formulated, therapeutic antibodies in liquid solutions are prone to degradation, aggregation, or undesirable chemical modifications. The stability of antibodies in liquid formulations depends not only on the type of excipients used in the formulation but also on the amount and proportion of the excipients relative to each other. Furthermore, other factors besides stability must be considered when preparing liquid antibody formulations. Examples of such additional considerations include the viscosity of the solution and the antibody concentration that a given formulation can accommodate, as well as the visual quality or attractiveness of the formulation. Therefore, when formulating therapeutic antibodies, great care must be taken to obtain a formulation that remains stable, contains a sufficient concentration of antibody, and has suitable viscosity and other properties, allowing for convenient administration to patients.

[0004] Antibodies against human lymphocyte activation gene-3 (LAG-3) are an example of therapeutically relevant macromolecules that require appropriate formulation. Anti-LAG-3 antibodies can be used clinically to stimulate or enhance immune responses and / or to treat subjects with cancer or chronic viral infections. Exemplary anti-LAG3 antibodies available in this article are described below. especiallyThis includes LAG525 (and other LAG3 antibodies disclosed in US 20100233183), relatlimab (and other LAG3 antibodies disclosed in US 20110150892), GSK2831781 (and other LAG3 antibodies disclosed in US 20140286935), MGD013 (and other LAG3 antibodies disclosed in WO2015200119), as well as US 20160222116, US 20170022273, US20170097333, US 20170137517, US 20170267759, and US LAG3 antibodies disclosed in 20170290914, US20170334995, WO2017062888, WO2016126858, WO2016200782, WO2017087589, WO2017087901, WO2017106129, WO2017149143, WO2017198741, WO2017219995 and WO2017220569.

[0005] Although anti-LAG-3 antibodies are known, there is still a need in the art for novel pharmaceutical formulations that contain sufficiently stable anti-LAG-3 antibodies suitable for administration to patients. Summary of the Invention

[0006] This disclosure addresses the aforementioned needs by providing a stable pharmaceutical formulation comprising a human antibody that specifically binds to human lymphocyte activation gene-3 (LAG-3).

[0007] In one aspect, a low-viscosity, stable liquid pharmaceutical formulation is provided, comprising: (i) a human antibody that specifically binds to human lymphocyte activation gene-3 protein (LAG-3); (ii) a buffer; (iii) an organic solubilizer; and (iv) a stabilizer.

[0008] In various embodiments, the antibody is provided at a concentration of about 5 ± 0.75 mg / mL to about 250 ± 45 mg / mL. In one embodiment, the antibody is provided at 12.5 mg / mL. It is provided at a concentration of 1.85 mg / mL or about 12.5 mg / mL. In one embodiment, the antibody is provided at 25 mg / mL. It is provided at a concentration of 3.75 mg / mL or about 25 mg / mL. In another embodiment, the antibody is provided at 50 mg / mL. It is provided at a concentration of 7.5 mg / mL or about 50 mg / mL. In another embodiment, the antibody is provided at 100 mg / mL. It is provided at a concentration of 15 mg / mL or about 100 mg / mL. In one embodiment, the antibody is provided at 150 mg / mL. It is provided at a concentration of 22.5 mg / mL or approximately 150 mg / mL. In another embodiment, the antibody is provided at 175 mg / mL. It is provided at a concentration of 26.25 mg / mL or approximately 175 mg / mL. In another embodiment, the antibody is provided at 200 mg / mL. It is available at a concentration of 30 mg / mL or approximately 200 mg / mL.

[0009] In some embodiments, the formulation comprises any of the anti-LAG-3 antibodies disclosed in US 20170101472, which is incorporated herein by reference in its entirety. In some embodiments, the anti-LAG-3 antibody comprises (a) a heavy chain variable region (HCVR) comprising heavy chain complementarity-determining regions 1, 2, and 3 (HCDR1-HCDR2-HCDR3), each heavy chain complementarity-determining region comprising the sequences of SEQ ID NO:3, SEQ ID NO:4, and SEQ ID NO:5, respectively; and (b) a light chain variable region (LCVR) comprising light chain complementarity-determining regions 1, 2, and 3 (LCDR1-LCDR2-LCDR3), each light chain complementarity-determining region comprising the sequences of SEQ ID NO:6, SEQ ID NO:7, and SEQ ID NO:8, respectively. In one embodiment, the antibody comprises an HCVR containing the amino acid sequence of SEQ ID NO:1 and an LCVR containing the amino acid sequence of SEQ ID NO:2. In one embodiment, the antibody comprises a heavy chain containing the amino acid sequence of SEQ ID NO: 9 and a light chain containing the amino acid sequence of SEQ ID NO: 10. In one embodiment, the antibody comprises HCVR having 90% sequence identity with SEQ ID NO: 1. In one embodiment, the antibody comprises LCVR having 90% sequence identity with SEQ ID NO: 2. In one embodiment, the antibody comprises HCVR having 90% sequence identity with SEQ ID NO: 1 and LCVR having 90% sequence identity with SEQ ID NO: 2. In one embodiment, the antibody comprises a heavy chain and a light chain, wherein the heavy chain comprises the amino acid sequence of amino acids 1-448 of SEQ ID NO: 9. In one embodiment, the antibody comprises a heavy chain and a light chain, wherein the light chain comprises the amino acid sequence of SEQ ID NO: 10. In one embodiment, the antibody comprises a heavy chain / light chain containing the amino acid sequence of SEQ ID NO: 9 / 10. In one embodiment, the antibody comprises a heavy chain containing the amino acid sequence of amino acids 1-448 of SEQ ID NO: 9 and a light chain containing the amino acid sequence of SEQ ID NO: 10.

[0010] In one embodiment, the pH of the liquid formulation is H 6.0. 0.5, pH 6.0 The pH values ​​are 0.4, 6.0 ± 0.3, 6.0 ± 0.2, 6.0 ± 0.1, 6.0 ± 0.05, 6.0 ± 0.01, or 6.0. In one embodiment, the pH of the liquid formulation is approximately pH 6.0. 0.3.

[0011] In one embodiment, the buffer contains histidine. In some embodiments, the concentration of the histidine buffer is from 5 mM ± 1 mM to 50 mM ± 10 mM, for example, from 5 mM ± 1 mM to 25 mM ± 5 mM. In one embodiment, the concentration of the histidine buffer is 10 mM ± 2 mM or about 10 mM. In one embodiment, the concentration of the histidine buffer is 20 mM ± 4 mM or about 20 mM. In one embodiment, the concentration of the histidine buffer is 40 nM ± 8 mM or about 40 nM. In some embodiments, the histidine buffer contains L-histidine and L-histidine hydrochloride monohydrate. In one embodiment, the concentration of L-histidine is from 2 mM ± 0.4 mM to 25 mM ± 5 mM, for example, from 4 mM ± 0.8 mM to 20 mM ± 4 mM. In one embodiment, the concentration of L-histidine hydrochloride monohydrate is from 2 mM ± 0.4 mM to 25 mM ± 5 mM, for example, from 4 mM ± 0.8 mM to 20 mM ± 4 mM. In one embodiment, the buffer contains 5.0 mM ± 1.0 mM of L-histidine and 5.0 mM ± 1.0 mM of L-histidine hydrochloride monohydrate. In one embodiment, the buffer contains 10 mM ± 2 mM of histidine, wherein the histidine contains 5.0 mM ± 1.0 mM of L-histidine and 5.0 mM ± 1.0 mM of L-histidine hydrochloride monohydrate.

[0012] In some embodiments, the organic co-solvent is a nonionic polymer containing a polyoxyethylene moiety. 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 polysorbate 80. In one embodiment, the organic co-solvent is polysorbate 20. In one embodiment, the organic co-solvent is polyethylene glycol, such as PEG3350.

[0013] In one embodiment, the concentration of the organic co-solvent is approximately 0.01%. 0.005% to approximately 1% 0.5% "weight-to-volume ratio" or "w / v", of which For example 0.1 g / ml = 10% and 0.01 g / ml = 1%. In some embodiments, the organic solvent is at a concentration of 0.05%. 0.025% to 0.5% 0.25% (w / v) polysorbate. In one embodiment, the organic co-solvent is at a concentration of 0.2%. 0.02% w / v or about 0.2% polysorbate 80. In another embodiment, the organic co-solvent is at a concentration of 0.1%. 0.05% w / v or about 0.1% w / v of polysorbate 80. In one embodiment, the organic co-solvent is at a concentration of 0.2%. 0.1% w / v or about 0.2% polysorbate 20. In another embodiment, the organic co-solvent is at a concentration of 0.1%. 0.05% w / v or about 0.1% w / v of polysorbate 20. In another embodiment, the organic co-solvent is at a concentration of 1.0%. PEG3350 at 0.5% w / v or about 1.0% w / v.

[0014] In some embodiments, the stabilizer is a sugar. In one embodiment, the sugar is sucrose. In various embodiments, the concentration of the stabilizer is 1%. 0.2% w / v to 20% 4% w / v, 5% 1% w / v to 15% 3% w / v or 1% 0.2% to 10% 2% w / v. In one implementation, the stabilizer is at a concentration of 5%. 1% w / v or about 5% w / v sucrose. In another embodiment, the stabilizer is at a concentration of 9%. 1.8% w / v or about 9% w / v sucrose. In another embodiment, the stabilizer is at a concentration of 10%. 2% w / v or about 10% w / v of sucrose.

[0015] In some embodiments, the stabilizer is an amino acid. In one embodiment, the stabilizer is an amino acid, such as arginine hydrochloride or proline. In another embodiment, the stabilizer is arginine hydrochloride. In some embodiments, the stabilizer is arginine hydrochloride, and the concentration is 1 mM. 2 mM to 100 mM 20 mM. In one embodiment, the stabilizer is at a concentration of 20 mM. 4 mM or about 20 mM arginine hydrochloride. In one embodiment, the stabilizer is at a concentration of 80 mM. 16 mM or about 80 mM of arginine hydrochloride.

[0016] In some embodiments, the stabilizer includes sucrose and arginine. In another embodiment, the stabilizer is at a concentration of 10%. 2% w / v or about 10% w / v sucrose and a concentration of 20 mM 4 mM or about 20 mM of arginine hydrochloride.

[0017] In one aspect, a stable liquid pharmaceutical formulation is provided, comprising: (i) 5 ± 0.75 mg / ml to 250 ± 37.5 mg / ml of a human antibody specifically binding to human LAG-3; (ii) 0 Up to 40 (iii) 8 mM histidine buffer; (iv) 0% to 0.5% ± 0.25% (w / v) polysorbate 80; 3% (w / v) sucrose; and (v) 1 mM 0.2 mM to 100 mM The 20 mM arginine hydrochloride has a pH of about 5.3 to about 6.7; wherein the anti-LAG-3 antibody comprises a heavy chain variable region (HCVR) and a light chain variable region (LCVR), such that the HCVR / LCVR combination comprises heavy chain and light chain complementarity-determining regions (HCDR1-HCDR2-HCDR3 / LCDR1-LCDR2-LCDR3) comprising the amino acid sequences of SEQ ID NO: 3–4–5 / SEQ ID NO: 6–7–8, respectively. In one embodiment, the anti-LAG-3 antibody comprises a heavy chain variable region (HCVR) and a light chain variable region (LCVR) comprising the amino acid sequences of SEQ ID NO: 1 and SEQ ID NO: 2, respectively. In some embodiments, the anti-LAG-3 antibody comprises an Fc region selected from the group consisting of human IgG1, IgG2, IgG3, and IgG4 isotypes. In one embodiment, the antibody comprises a human IgG4 isotype. In one embodiment, the antibody comprises a heavy chain containing the amino acid sequence of SEQ ID NO: 9 and a light chain containing the amino acid sequence of SEQ ID NO: 10. In one embodiment, the antibody has a molecular weight of 145 kDa. 5 kDa.

[0018] In some embodiments, a stable liquid pharmaceutical formulation is provided comprising: (i) 5 ± 0.75 mg / ml to 250 ± 37.5 mg / ml of a human antibody specifically binding to human LAG-3; (ii) 0 Up to 40 (iii) 8 mM histidine buffer; (iv) 0% to 0.5% ± 0.25% (w / v) polysorbate 80; 3% (w / v) sucrose; and (v) 1 mM 0.2 mM to 100 mM The 20 mM arginine hydrochloride, pH about 5.3 to about 6.7; wherein the anti-LAG-3 antibody comprises HCVR and LCVR, wherein the HCVR has 90% sequence identity with SEQ ID NO: 1, and / or the LCVR has 90% sequence identity with SEQ ID NO: 2. In one embodiment, the anti-LAG-3 antibody comprises HCVR containing the amino acid sequence of SEQ ID NO: 1 and LCVR containing the amino acid sequence of SEQ ID NO: 2. In one embodiment, the anti-LAG-3 antibody comprises a heavy chain containing the amino acid sequence of SEQ ID NO: 9 and a light chain containing the amino acid sequence of SEQ ID NO: 10.

[0019] In some embodiments, a stable liquid pharmaceutical formulation is provided comprising: (i) 5 ± 0.75 mg / ml to 250 ± 37.5 mg / ml of a human antibody specifically binding to human LAG-3; (ii) 0 Up to 40 (iii) 8 mM histidine buffer; (iv) 0% to 0.5% ± 0.25% (w / v) polysorbate 80; 3% (w / v) sucrose; and (v) 1 mM 0.2 mM to 100 mM The 20 mM arginine hydrochloride has a pH of about 5.3 to about 6.7; the anti-LAG-3 antibody comprises HCVR and LCVR, wherein the HCVR comprises the amino acid sequence of SEQ ID NO: 1 having no more than five amino acid substitutions, and wherein the LCVR comprises the amino acid sequence of SEQ ID NO: 2 having no more than two amino acid substitutions. In one embodiment, the anti-LAG-3 antibody comprises HCVR containing the amino acid sequence of SEQ ID NO: 1 and LCVR containing the amino acid sequence of SEQ ID NO: 2. In one embodiment, the anti-LAG-3 antibody comprises a heavy chain containing the amino acid sequence of SEQ ID NO: 9 and a light chain containing the amino acid sequence of SEQ ID NO: 10.

[0020] In some embodiments, the formulation of any of the foregoing aspects has properties selected from the group consisting of: (i) as described herein, the formulation is stable at long-term storage at 5°C; (ii) as described herein, the formulation is stable to agitation stress; (iii) as described herein, the formulation is stable even with a variation of up to 50% in the concentration of excipients; (iv) the formulation is stable to and compatible with intravenous delivery devices and procedures; and (v) the formulation is stable at long-term storage in glass vials.

[0021] In some embodiments of this invention, a stable liquid formulation is provided comprising: (i) 5 ± 0.75 mg / ml to 250 ± 37.5 mg / ml of a human antibody specifically binding to human LAG-3; (ii) 5 1 mM to 20 (iii) 0.05% histidine buffer (4 mM); 0.025% to 0.3% ± 0.15% (w / v) of polysorbate 80; (iv) 1% 0.2% to 15% 3% (w / v) sucrose; and (v) 10 mM 2 mM to 30 mM The antibody comprises 6 mM arginine hydrochloride at a pH of approximately 6.0, wherein the antibody contains HCVR / LCVR containing the amino acid sequence pair of SEQ ID NO: 1 / 2.

[0022] In one embodiment of this, the stable liquid formulation comprises (i) 25 ± 3.75 mg / mL of anti-LAG-3 antibody; (ii) 10 (iii) 2 mM histidine buffer; (iv) 0.1% ± 0.05% (w / v) polysorbate 80; 4 mM arginine hydrochloride; and (v) 5% 1% (w / v) sucrose, pH 6.0 ± 0.3, wherein the antibody comprises HCVR / LCVR containing the amino acid sequence pair of SEQ ID NO: 1 / 2.

[0023] In one embodiment of this, the stable liquid formulation comprises (i) 50 ± 7.5 mg / mL of anti-LAG-3 antibody; (ii) 10 2 (iii) 0.1% ± 0.5% (w / v) polysorbate 80; (iv) 20 mM histidine buffer. 4 mM arginine hydrochloride; and (v) 10% The antibody comprises 2% (w / v) sucrose at pH 6.0 ± 0.3, and contains an HCVR / LCVR with the amino acid sequence pair of SEQ ID NO: 1 / 2. In one embodiment, more than 98% of the antibody retains its purity after being stored at 5°C for 12 months.

[0024] In one embodiment of this, the stable liquid formulation comprises (i) 50 ± 7.5 mg / mL of anti-LAG-3 antibody; (ii) 10 2 (iii) 0.1% ± 0.5% (w / v) polysorbate 80; and (iv) 5% 1% (w / v) sucrose, pH 6.0 ± 0.3, wherein the antibody comprises HCVR / LCVR containing the amino acid sequence pair of SEQ ID NO: 1 / 2.

[0025] In one embodiment, the stable liquid formulation comprises (i) 100 ± 15 mg / mL of anti-LAG-3 antibody; (ii) 10 (iii) 2 mM histidine buffer; (iv) 0.1% ± 0.05% (w / v) polysorbate 80; 2 mM to 100 mM 20% arginine hydrochloride; and (v) 1% 0.2% (w / v) sucrose, pH 6.0 ± 0.3, wherein the antibody comprises HCVR / LCVR containing the amino acid sequence pair of SEQ ID NO: 1 / 2.

[0026] In one embodiment, the stable liquid formulation comprises (i) 150 ± 22.5 mg / mL of anti-LAG-3 antibody; (ii) 10 (iii) 2 mM histidine buffer; (iv) 0.1% ± 0.05% (w / v) polysorbate 80; 0.2% to 10% 2% (w / v) sucrose; and (v) 1 mM 0.2 mM to 100 mM 20 mM arginine hydrochloride, pH 6.0 ± 0.3, wherein the antibody comprises HCVR / LCVR containing the amino acid sequence pair of SEQ ID NO: 1 / 2.

[0027] In one embodiment of this aspect, the stable liquid formulation comprises (i) 175 ± 26.25 mg / mL of anti-LAG-3 antibody; (ii) 10 ± 2 mM of histidine buffer; (iii) 0.1% ± 0.05% (w / v) of polysorbate 80; (iv) 1% ± 0.2% to 10% ± 2% (w / v) of sucrose; and (v) 10 mM ± 2 mM to 100 mM ± 20 mM of arginine hydrochloride at pH 6.0 ± 0.3, wherein the antibody comprises HCVR / LCVR containing the amino acid sequence pair of SEQ ID NO: 1 / 2.

[0028] In one embodiment of this, the stable liquid formulation comprises (i) 200 ± 30.00 mg / mL of anti-LAG-3 antibody; (ii) 10 (iii) 2 mM histidine buffer; (iv) 0.1% ± 0.05% (w / v) polysorbate 80; 2% (w / v) sucrose; and (v) 80 mM 16 mM arginine hydrochloride, pH 6.0 ± 0.3, wherein the antibody comprises HCVR / LCVR containing the amino acid sequence pair of SEQ ID NO: 1 / 2.

[0029] In one embodiment, the stable liquid formulation comprises (a) 5-250 mg / ml of an antibody that specifically binds to LAG-3, said antibody comprising HCVR of SEQ ID NO: 1 and LCVR of SEQ ID NO: 2; (b) 10 mM 2 mM histidine buffer, pH 6.0 0.3; (c) 0.1% 0.05% w / v polysorbate 80; and (d) 5% 1% w / v sucrose. In some cases, the stable liquid formulation further contains 10 mM sucrose. 2 mM to 100 mM 20 mM arginine hydrochloride.

[0030] In one embodiment, after storage at 40°C for 28 days, ≥ 95% of the antibodies are native, and ≥ 45% of the antibodies are in the major charge form. In one embodiment, after storage at 25°C for three months, > 98% of the antibodies are native, and ≥ 52% of the antibodies are in the major charge form. In one embodiment, after storage at 5°C for 12 months, > 98% of the antibodies are native, and > 50% of the antibodies are in the major charge form. In one embodiment, more than 98% of the antibodies have a native conformation after storage at 5°C for 24 months. In one embodiment, at least 98% or more of the antibodies have a native conformation after stirring for 120 minutes.

[0031] In one aspect, this disclosure provides a stable liquid formulation comprising: (i) up to 100 mg / mL of anti-LAG-3 antibody; (ii) 2 mM 0.4 mM to 20 mM (iii) up to 20% histidine buffer; (iv) 4% (w / v) sucrose; and (iv) up to 0.2% 0.1% w / v polysorbate, pH 6.0 0.3. In one embodiment, the stable liquid formulation comprises 25 mg / mL anti-LAG-3 antibody. In one embodiment, the stable liquid formulation comprises 50 mg / mL anti-LAG-3 antibody. In one embodiment, the stable liquid formulation comprises 75 mg / mL anti-LAG-3 antibody. In one embodiment, the stable liquid formulation comprises 10 mM 2 mM 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 stable liquid formulation comprises 0.2% polysorbate. In one embodiment, the polysorbate is polysorbate 80 or polysorbate 20. In one embodiment, the anti-LAG-3 antibody comprises HCVR / LCVR of SEQ ID NO: 1 / 2.

[0032] In one aspect, a stable liquid pharmaceutical formulation of any of the foregoing 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 vial is a 2 ml, 5 ml, 10 mL, or 20 ml Type 1 clear glass vial. In one embodiment, the glass vial is a Type 1 borosilicate glass vial with a fluorocarbon-coated butyl rubber stopper. In one embodiment, the container is a microinjector. In one embodiment, the container is a syringe. In one embodiment, the container is a pre-filled syringe. In one embodiment, the syringe includes a plunger coated with a fluorocarbon. In some embodiments, the syringe is a 1 mL or 2.25 mL long glass syringe containing less than about 500 parts per billion of tungsten, equipped with a 27-G needle, a fluorocarbon-coated butyl rubber stopper, and a latex-free, non-cytotoxic rubber end cap. In one embodiment, the syringe is a 1 mL long glass syringe equipped with a 27-G thin-walled needle, a FLUROTEC-coated 4023 / 50 rubber stopper, and an FM 27 rubber end cap. In one embodiment, the syringe is a 1 mL, 2 mL, 3 mL, 5 mL, or 10 mL plastic syringe equipped with a needle.

[0033] In one aspect, a kit is provided comprising a stable pharmaceutical composition, container, and instructions for use of any of the foregoing aspects. In one embodiment, the container is a glass vial. In one embodiment, the container is a pre-filled syringe. In one embodiment, the syringe is a 1 mL or 2.25 mL long glass syringe equipped with a 27-G thin-walled needle, a FLUROTEC-coated 4023 / 50 rubber stopper, and an FM 27 rubber end cap. In one embodiment, the syringe is a 1 mL, 2 mL, 3 mL, 5 mL, or 10 mL plastic syringe with a needle.

[0034] In some embodiments, this disclosure provides a pre-filled syringe comprising a stable liquid pharmaceutical formulation comprising: (i) 5 ± 0.75 mg / ml to 250 ± 37.5 mg / ml of a human antibody specifically binding to human LAG-3; (ii) 5 1 mM to 20 (iii) 0.05% histidine buffer (4 mM); 0.025% to 0.3% ± 0.15% (w / v) of polysorbate 80; (iv) 1% 0.2% to 15% 3% (w / v) sucrose; and (v) 0 to 100 mM 20 mM arginine hydrochloride, pH 6.0 0.3, wherein the antibody comprises an HCVR / LCVR containing the amino acid sequence pair of SEQ ID NO: 1 / 2; wherein the formulation has the property selected from the group consisting of: (i) after storage at 5°C for 24 months, ≥ 98% of the antibody is in the natural form; (ii) after storage at 5°C for 24 months, ≥ 50% of the antibody is a major charge variant; (iii) after storage at 25°C for 6 months, ≥ 98% of the antibody is in the natural form; (iv) after storage at 25°C for 6 months, ≥ 45% of the antibody is a major charge variant; (v) more than 90% of the antibody has a mass of 145 kDa. (vi) A molecular weight of 1 kDa; (vii) More than 98% of the antibodies retain the native conformation after 12 months of storage at 5°C; and (vii) At least 97% or more of the antibodies retain the native conformation after 28 days of storage at 40°C.

[0035] In some embodiments, this disclosure provides a glass vial containing a stable liquid pharmaceutical formulation comprising: (i) 5 ± 0.75 mg / ml to 250 ± 37.5 mg / ml of a human antibody specifically binding to human LAG-3; (ii) 5 1 mM to 20 (iii) 0.05% histidine buffer (4 mM); 0.025% to 0.3% ± 0.15% (w / v) of polysorbate 80; (iv) 1% 0.2% to 15% 3% (w / v) sucrose; and (v) 0 to 100 mM 20 mM arginine hydrochloride, pH 6.0 0.3, wherein the antibody comprises an HCVR / LCVR containing the amino acid sequence pair of SEQ ID NO: 1 / 2; wherein the formulation has properties selected from the group consisting of: (i) the formulation is stable in glass vials and under stress; (ii) the formulation is stable for IV delivery devices and suitable for use in said devices; (iii) the formulation is stable with standard diluents known in the art ( For example (iv) The formulation is chemically and physically stable when diluted with 0.9% sodium chloride or 5% glucose; For example IV bags made of polyvinyl chloride, phthalates, polyolefins or polypropylene are stable; (v) formulations with standard infusion pumps ( For example (vi) Compatible with peristaltic pumps and fluid displacement pumps; ≥ 90% of antibodies have a strength of 145 kDa. The molecular weight is 1 kDa; (vii) after 24 months of storage at 5°C, more than 98% of the antibodies retain the native conformation; and (viii) after 6 months of storage at 25°C, at least 97% or more of the antibodies retain the native conformation.

[0036] Other implementation methods will become apparent upon referring to the following detailed description. Attached Figure Description

[0037] Figure 1 The effect of polysorbate 80 concentration on the stability of 50 mg / mL mAb1 after stirring is shown (orbital oscillator at 250 RPM for 48 hours).

[0038] Figure 2 The effects on quality properties of fourteen DP formulations (50 mg / mL; 800 mg DP) after storage at 2–8 °C for 12 months were described.

[0039] Figure 3 The effects on quality properties of fourteen DP formulations (50 mg / mL; 800 mg DP) after a 6-month storage period at 25°C / 60% RH were depicted.

[0040] Figure 4 The effect of antibody concentration (mg / mL protein) on the viscosity of a formulation (pH 6) containing 10 mM histidine, 5% sucrose, 70 mM arginine HCl and 0.1% PS80 was depicted. Detailed Implementation

[0041] Before describing the method of the present invention, it should be understood that the invention is not limited to the specific methods and experimental conditions described, as such methods and conditions can vary. It should also be understood that the terminology used herein is for the purpose of describing particular embodiments only and is not intended to be limiting, as the scope of the invention will be limited only by the appended claims.

[0042] Unless otherwise defined, 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 pertains. The term "about," as used herein, when referring to a particular enumerated numerical value or range, means that the value may differ from the enumerated value by no more than 1%. For example, as used herein, the expression "about 100" includes 99 and 101 and all values ​​between them (e.g., 99.1, 99.2, 99.3, 99.4, etc.). Although any methods and materials similar to or equivalent to those described herein may be used in the practice or testing of this invention, preferred methods and materials are described here. All publications mentioned herein are incorporated herein by reference in their entirety.

[0043] As used herein, the term "pharmaceutical preparation" means at least one active ingredient ( For example This disclosure provides a combination of a small molecule, a large molecule, a compound, etc., capable of exerting a biological effect in humans or non-human animals, and at least one inactive ingredient, which, when combined with an active ingredient or one or more other inactive ingredients, is suitable for therapeutic administration to humans or non-human animals. Unless otherwise expressly stated, as used herein, the term "formulation" means "pharmaceutical formulation." This disclosure provides pharmaceutical formulations comprising at least one therapeutic polypeptide. According to certain embodiments of this disclosure, the therapeutic polypeptide is an antibody or antigen-binding fragment thereof that specifically binds to the human lymphocyte activation gene-3 (LAG-3) protein. More specifically, this disclosure includes pharmaceutical formulations comprising: (i) a human antibody that specifically binds to human LAG-3; (ii) a histidine buffer; (iii) an organic cosolvent that is a nonionic surfactant; and (iv) a stabilizer that is a carbohydrate and / or an amino acid. Specific exemplary components and formulations included in this invention are described in detail below.

[0044] Antibodies that specifically bind to LAG-3 The pharmaceutical formulation disclosed herein may comprise a human antibody or antigen-binding fragment thereof that specifically binds to human LAG-3. As used herein, the term "LAG-3" refers to human lymphocyte activation gene-3. Antibodies against human LAG-3 are described, for example, in US20100233183, US 20110150892, US 20140286935, WO2015200119, US 20160222116, US 20170022273, US 20170097333, US 20170137517, US 20170267759, US20170290914, US... 20170334995, WO2017062888, WO2016126858, WO2016200782, WO2017087589, WO2017087901, WO2017106129, WO2017149143, WO2017198741, WO2017219995 and WO2017220569.

[0045] As used herein, the term "antibody" is generally intended to refer to an immunoglobulin molecule comprising four polypeptide chains—two heavy chains (H chains) and two light chains (L chains) linked together by disulfide bonds—as well as its multimers ( For example IgM); however, only the heavy chain ( Right nowImmunoglobulin molecules composed of heavy chains (without light chains) are also included in the definition of the term "antibody". Each heavy chain contains a heavy chain variable region (abbreviated as HCVR or V in this document). H The heavy chain constant region contains three domains: CH1, CH2, and CH3. Each light chain contains a light chain variable region (abbreviated as LCVR or V in this paper). L The light chain constant region contains a structural domain (CL1). H District and V L The region can be further subdivided into highly variable regions known as complementary determinant regions (CDRs), which are interspersed with more conservative regions known as frame regions (FRs). Each V H and V L It consists of three CDRs and four FRs arranged in the following order from the amino terminus to the carboxyl terminus: FR1, CDR1, FR2, CDR2, FR3, CDR3, FR4.

[0046] Unless otherwise specified, the term “antibody” as used herein should be understood to include the complete antibody molecule as well as its antigen-binding fragment. As used herein, the term “antigen-binding portion” or “antigen-binding fragment” (or simply “antibody portion” or “antibody fragment”) of an antibody refers to one or more fragments of an antibody that retain the ability to specifically bind to human LAG-3 or its epitopes.

[0047] As used herein, "isolated antibody" is intended to refer to an antibody that is substantially free of other antibodies with different antigen specificities. For example The isolated antibody that specifically binds to human LAG-3 basically does not contain antibodies that specifically bind to antigens other than human LAG-3.

[0048] The term "specific binding" refers to the formation of a relatively stable complex between an antibody or its antigen-binding fragment and the antigen under physiological conditions. Specific binding is characterized by a minimum of approximately 1 x 10-1 -8 Or a larger dissociation constant. Methods for determining whether two molecules specifically bind are well known in the art and include, for example, equilibrium dialysis, surface plasmon resonance, etc. However, isolated antibodies that specifically bind to human LAG-3 may be cross-reactive with other antigens, such as LAG-3 molecules from other species (orthologs). In the context of this disclosure, multispecificity (MPI) binding to human LAG-3 and one or more additional antigens is discussed. For example Bispecific antibodies are believed to "specifically bind" to human LAG-3. Furthermore, isolated antibodies may be substantially free of other cellular material or chemicals.

[0049] Exemplary anti-human LAG-3 antibodies that may be included in pharmaceutical formulations disclosed herein are set forth in patent applications US20170101472 and WO2017062888, the disclosures of which are incorporated herein by reference in their entirety.

[0050] According to certain embodiments of this disclosure, the anti-human LAG-3 antibody or its antigen-binding fragment comprises the heavy chain complementarity-determining region (HCDR) 1 of SEQ ID NO: 3, HCDR2 of SEQ ID NO: 4, and HCDR3 of SEQ ID NO: 5. In some embodiments, the anti-human LAG-3 antibody or its antigen-binding fragment comprises the HCVR of SEQ ID NO: 1.

[0051] According to certain embodiments of this disclosure, the anti-human LAG-3 antibody or its antigen-binding fragment comprises the light chain complementarity-determining region (LCDR) 1 of SEQ ID NO: 6, the LCDR2 of SEQ ID NO: 7, and the LCDR3 of SEQ ID NO: 8. In some embodiments, the anti-human LAG-3 antibody or its antigen-binding fragment comprises the LCDR of SEQ ID NO: 2.

[0052] According to certain embodiments of this disclosure, anti-human LAG-3 or its antigen-binding fragment comprises HCVR having 90%, 95%, 98% or 99% sequence identity with SEQ ID NO: 1.

[0053] According to certain embodiments of this disclosure, the anti-human LAG-3 or its antigen-binding fragment comprises an LCVR having 90%, 95%, 98%, or 99% sequence identity with SEQ ID NO: 2.

[0054] According to certain embodiments of this disclosure, anti-human LAG-3 or its antigen-binding fragment comprises an HCVR containing an amino acid sequence of SEQ ID NO: 1 having no more than 5 amino acid substitutions.

[0055] According to certain embodiments of this disclosure, anti-human LAG-3 or its antigen-binding fragment comprises an LCVR containing an amino acid sequence having no more than two amino acid substitutions in SEQ ID NO: 2.

[0056] Sequence identity can be determined by any method known in the art ( For example GAP, BESTFIT, and BLAST are used for measurement.

[0057] This disclosure also includes formulations comprising anti-LAG-3 antibodies, wherein the anti-LAG-3 antibodies comprise variants of any HCVR, LCVR, and / or CDR amino acid sequences disclosed herein having one or more conserved amino acid substitutions. For example, this disclosure includes formulations comprising anti-LAG-3 antibodies having HCVR, LCVR, and / or CDR amino acid sequences having, relative to any of the HCVR, LCVR, and / or CDR amino acid sequences disclosed herein, conserved amino acid substitutions such as 10 or fewer, 8 or fewer, 6 or fewer, 4 or fewer, etc.

[0058] In some implementations, the anti-LAG-3 antibody includes an Fc region selected from a group consisting of human IgG1, IgG2, IgG3 and IgG4 isotypes.

[0059] The non-limiting exemplary antibody used in the embodiments herein is referred to as “mAb1”. This antibody is also referred to as H4sH15482P in US20170101472, and is also referred to as “REGN3767” or “Fianlimab”. mAb1 (H4sH15482P) comprises the HCVR / LCVR amino acid sequence pair having SEQ ID NO: 1 / 2, and the HCDR1-HCDR2-HCDR3 / LCDR1-LCDR2-LCDR3 domain represented by SEQ ID NO: 3–4–5 / SEQ ID NO: 6–7–8.

[0060] According to certain embodiments of this disclosure, anti-human LAG-3 or its antigen-binding fragment comprises the heavy chain of SEQ ID NO: 9 and the light chain of SEQ ID NO: 10.

[0061] As is well known in the art, the terminal cleavage of amino acids can occur during antibody production (see, for example, Wang). et al. (2007, J. Pharma. Sci. 96: 1-26). Therefore, in some embodiments, the anti-LAG-3 antibody comprises a heavy chain and a light chain, wherein the heavy chain comprises the amino acid sequence of SEQ ID NO: 9, wherein a C-terminal lysine is not present in the amino acid sequence of SEQ ID NO: 9. In some embodiments, the formulation of this disclosure comprises about 50%, about 60%, about 70%, about 80%, about 90%, about 95%, about 98% or more of the anti-LAG-3 antibody, wherein a C-terminal lysine is absent.

[0062] The amount of antibody or its antigen-binding fragment contained in the pharmaceutical formulation disclosed herein can vary depending on the specific properties required by the formulation and the specific circumstances and purpose for which the formulation is intended to be used. In some embodiments, the pharmaceutical formulation is a liquid formulation that may contain 5 0.75 mg / mL to 250 37.5 mg / mL 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, the formulation disclosed herein may contain 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 140 mg / mL; about 145 mg / mL; about 150 mg / mL; about 155 mg / mL; about 160 mg / mL; about 140 mg / mL; about 145 mg / mL; about 150 mg / mL; about 155 mg / mL; about 160 mg / mL; about 140 mg / mL; about 155 mg / mL; about 160 mg / mL; about 140 mg / mL; about 155 mg / mL; about 150 mg / mL; about 160 mg / mL; about 140 mg / mL; about 155 mg / mL; about 150 mg / mL; about 155 mg / mL; about 160 mg / mL; about 140 mg / mL; about 155 mg / mL; about 150 mg / mL; about 155 mg / mL; mg / mL; approximately 165 mg / mL; approximately 170 mg / mL; approximately 175 mg / mL; approximately 180 mg / mL; approximately 185 mg / mL; approximately 190 mg / mL; approximately 195 mg / mL; approximately 200 mg / mL; approximately 205 mg / mL; approximately 210 mg / mL; approximately 215 mg / mL; approximately 220 mg / mL; approximately 225 mg / mL; approximately 230 mg / mL; approximately 235 mg / mL; approximately 240 mg / mL; approximately 245 mg / mL; or approximately 250 mg / mL of an antibody or antigen-binding fragment specifically binding to human LAG-3.

[0063] Excipients and pH The pharmaceutical formulations disclosed herein comprise one or more excipients. As used herein, the term "excipient" refers to any non-therapeutic agent added to the formulation to provide a desired consistency, viscosity, or stabilizing effect.

[0064] In some embodiments, the pharmaceutical formulation of the present invention comprises at least one organic cosolvent, the type and amount of which are determined during rough processing or stirring. For example The human LAG-3 antibody is stabilized under vortex conditions. In some embodiments, "stabilized" means preventing the formation of more than 3% (in molars) of aggregated antibody during the coarse treatment. In some embodiments, the coarse treatment involves vortexing a solution containing the antibody and an organic cosolvent for about 60 minutes or about 120 minutes.

[0065] In some 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 disclosed herein include... For examplePolysorbates, such as polysorbate 20, polysorbate 28, polysorbate 40, polysorbate 60, polysorbate 65, polysorbate 80, polysorbate 81, and polysorbate 85; poloxamers, such as poloxamer 181, poloxamer 188, and poloxamer 407; or polyethylene glycol (PEG, e.g., PEG3350). Polysorbate 20 is also known as TWEEN 20, dehydrated sorbitan monolaurate, and polyoxyethylene dehydrated sorbitan monolaurate. Poloxamer 188 is also known as Pronnick F68.

[0066] The amount of nonionic surfactant contained in the pharmaceutical formulations disclosed herein can vary depending on the specific properties required by the formulation and the specific circumstances and purpose for which the formulation is intended to be used. In some embodiments, the formulation may contain 0.01% 0.005% to 0.5% 0.25% surfactant. For example, the formulations disclosed herein 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% 0.21%; approximately 0.22%; approximately 0.23%; approximately 0.24%; approximately 0.25%; approximately 0.26%; approximately 0.27%; approximately 0.28%; approximately 0.29%; approximately 0.30%; approximately 0.35%; approximately 0.40%; approximately 0.45%; approximately 0.46%; approximately 0.47%; approximately 0.48%; approximately 0.49%; approximately 0.50%; approximately 0.55%; or approximately 0.60% of polysorbate 20, polysorbate 80, or PEG3350.

[0067] The pharmaceutical formulation disclosed herein may also contain one or more stabilizers, the type and amount of which stabilize the human LAG-3 antibody under heat stress or stirring stress conditions. In some embodiments, "stable" means when the solution containing the antibody and the heat stabilizer is at about 45°C. The antibody retains more than 91% of its native conformation when held for up to approximately 28 days. In some embodiments, "stable" means that the solution containing the antibody and heat stabilizer is kept at approximately 45°C. At a maximum of approximately 28 days, less than 6% of the antibody aggregated. As used herein, “natural” refers to the predominant form of the antibody by size exclusion, which is typically the intact monomer of the antibody. The term “natural” also refers to the non-aggregated and non-degradable form of the antibody.

[0068] In some embodiments, the heat stabilizer is a sugar, such as sucrose, and the amount of stabilizer contained in the formulation can vary depending on the specific circumstances and the intended purpose of the formulation. In some embodiments, the formulation may contain about 1% to about 15% sugar; about 2% to about 14% sugar; about 3% to about 13% sugar; about 4% to about 12% sugar; about 5% to about 12% sugar; about 6% to about 11% sugar; about 7% to about 10% sugar; about 8% to about 11% sugar; or about 9% to about 11% sugar. For example, the pharmaceutical formulation of this disclosure may contain 4% sugar. 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%; approximately 14% 2.8% sugar; or approximately 15% 3.0% sugar ( For example sucrose).

[0069] In some embodiments, the stabilizer is an amino acid, such as arginine. In some embodiments, the formulation may contain 1-20% w / v arginine. In some embodiments, the amino acid in the formulation may also be used as a viscosity reducer, for example in high-concentration formulations, such as when the antibody concentration is 100 mg / ml or greater. In some embodiments, when the antibody concentration is 175 mg / mL, arginine hydrochloride is added at approximately 1 mM. 0.2 mM to approximately 100 mM A concentration of 20 mM is present in the composition and acts as a viscosity reducer. In some embodiments, the pharmaceutical formulation comprises 175 mg / mL anti-LAG3 antibody and 20 mM arginine hydrochloride.

[0070] The pharmaceutical formulations disclosed herein may also contain buffers or buffer systems for maintaining a stable pH and helping to stabilize human LAG-3 antibodies. As used herein, the term "buffer" means a pharmaceutically acceptable buffer that maintains a stable pH or resists changes in the pH of a solution. In some aspects, the buffer contains phosphates. In some aspects, the buffer contains histidine.

[0071] In the context of this disclosure, "histidine buffer" or "histidine-containing buffer" is a buffer containing the amino acid histidine. Examples of histidine buffers include histidine chloride, histidine acetate, histidine phosphate, and histidine sulfate. In one embodiment, the histidine buffer is prepared by dissolving L-histidine and L-histidine hydrochloride in a defined amount and ratio. For example The histidine buffer is prepared by titrating L-histidine (free base, solid) with dilute hydrochloric acid. Throughout this disclosure, the term "histidine" is used interchangeably with "histidine buffer." In some embodiments, "stable" means that when the solution containing the antibody and buffer is kept at about 40°C for up to about 28 days, the antibody content is less than about 4.5%. 0.5% antibody aggregation.

[0072] In some implementations, "stable" means that when the solution containing antibodies and buffers is kept at about 40°C for up to about 28 days, the viscosity is less than 2%. 0.5% or less than 1% 0.5% antibody aggregation. In some implementations, "stable" means that when the solution containing antibodies and buffers is stored at approximately 40°C for up to approximately 28 days, it maintains at least 97% stability, as determined by size exclusion chromatography. 0.5% or at least 98% 0.5% of the antibody is in its native conformation. "Natural" or "native conformation" refers to antibody moieties that are neither aggregated nor degraded. This is typically determined by a assay measuring the relative size of the antibody entity, such as size exclusion chromatography. Unaggregated and undegraded antibodies elute in fractions equivalent to those of the natural antibody and are usually the dominant elution fraction. Aggregated antibodies elute in fractions larger than those of the natural antibody. Degraded antibodies elute in fractions indicating a smaller size than the natural antibody.

[0073] In some implementations, "stable" means that when the solution containing antibodies and buffers is stored at approximately 40°C for up to approximately 28 days, at least 35% of the required stability, as determined by cation exchange chromatography. 0.5% The antibody exists in its predominant charge form. In some embodiments, "stable" means that when the solution containing the antibody and buffer is stored at approximately 40°C for up to approximately 28 days, it maintains at least 46% stability, as determined by cation exchange chromatography. 0.5% The antibody is in its dominant charge form. "Dominant charge" or "dominant charge form" refers to the antibody fraction eluted from the ion exchange resin in the main peak. Typically, there are more "basic" peaks on one side of the main peak and more "acidic" peaks on the other side.

[0074] The pharmaceutical formulations disclosed herein can have a pH of about 5.2 to about 6.4. For example, the formulations disclosed herein can 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 about 6.5. In some embodiments, the pH is 6.0. 0.4; 6.0 0.3; 6.0 0.2; 6.0 0.1; approximately 6.0; or 6.0.

[0075] In some embodiments, the buffer or buffering system comprises at least one buffer whose buffering range completely or partially overlaps with the pH range of 5.5-7.4. In some embodiments, the buffer comprises a histidine buffer. In some embodiments, the histidine buffer is in a 5 mM solution. 1 mM to 15 mM 3 mM; 6 mM 1.2 mM to 14 mM 2.8 mM; 7 mM 1.4 mM to 13 mM 2.6 mM; 8 mM 1.6 mM to 12 mM 2.4 mM; 9 1.8 mM to 11 mM 2.2 mM; 10 mM The system is present at a concentration of 2 mM; or approximately 10 mM. In some embodiments, the buffer system contains 10 mM. 2 mM histidine at pH 6.0 0.3. In some embodiments, the histidine buffer comprises L-histidine and L-histidine hydrochloride hydrate. In one embodiment, the histidine buffer comprises 5.0 mM... In one embodiment, the histidine buffer contains 5.0 mM L-histidine. 1.0 mM L-histidine hydrochloride monohydrate. In one embodiment, the histidine buffer contains 5.0 mM. 1.0 mM L-histidine and 5.0 mM 1.0 mM L-histidine hydrochloride hydrate.

[0076] The pharmaceutical formulation disclosed herein may also comprise one or more excipients for maintaining the presence of an anti-LAG-3 antibody pharmaceutical substance (e.g., 50 mg / mL antibody), and in some respects, for containing a high concentration of the LAG-3 antibody pharmaceutical substance. For example The stabilizer is a substance that reduces or lowers the viscosity of a liquid formulation (typically ≥ 150 mg / mL antibody). In some embodiments, the stabilizer is an amino acid. In one embodiment, the amino acid is arginine hydrochloride. In one embodiment, the pharmaceutical formulation of this disclosure contains arginine hydrochloride at concentrations of 1 mM, 10 mM, 15 mM, 20 mM, 25 mM, 30 mM, 35 mM, 40 mM, 45 mM, 50 mM, 55 mM, 60 mM, 65 mM, 70 mM, 75 mM, 80 mM, 85 mM, or 90 mM. In some embodiments, the formulation contains sufficient amounts to maintain the viscosity of the liquid formulation at less than 20. 3 centipoise, less than 15 2.25 centipoise or less than 11 1.65 centipoise of arginine hydrochloride. In some embodiments, the formulation contains sufficient amounts to maintain a viscosity of 15. 2.25 centipoise or less of arginine hydrochloride. In some embodiments, the formulation may contain about 1 mM to about 80 mM; about 10 mM to about 30 mM of arginine hydrochloride; or about 20 mM of arginine hydrochloride. For example, a pharmaceutical formulation of this disclosure may contain 1 mM 0.2 mM, 10 mM 2 mM, 15 mM 3 mM, 20 mM 4 mM, 25 mM 5 mM, 30 mM 6mM, 35mM 7 mM, 40 mM 8 mM, 45 mM 9 mM, 50 mM 10 mM, 55 mM 11 mM, 60 mM 12 mM, 65 mM 13 mM, 70 mM 14 mM, 75 mM 15 mM, 80 mM 16 mM, 85 mM 17mM or 90mM 18 mM arginine hydrochloride.

[0077] During antibody purification, it may be necessary or appropriate to change one buffer to another to obtain suitable excipient concentration, antibody concentration, pH, etc. Buffer exchange can... For example By using For example This is accomplished using a semi-permeable tangential flow filtration membrane via ultrafiltration / percolation (UF / DF). However, the use of this technique may lead to the Gibbs-Donna effect [Bolton et al., 2011, Biotechnol. Prog. 27(1):140-152]. During protein concentration, the accumulation of positive charge on the product side of the membrane is offset by the preferential migration of cations to the opposite side of the membrane. A potential consequence of this phenomenon is that certain components ( example like The final concentrations of components such as arginine hydrochloride, histidine, and L-proline may be lower than the expected target concentrations due to the electrostatic repulsion of positively charged percolation buffer excipients to positively charged antibody proteins during the UF / DF step. Therefore, this disclosure includes formulations in which, due to the Gibbs-Donnan effect, For example The concentrations of histidine and / or arginine hydrochloride differ from the amounts or ranges described herein.

[0078] Size exclusion describes the behavior of highly concentrated samples, where a large portion of the total solution volume is occupied by the solute, particularly large molecules such as proteins, excluding the solvent in that space. This reduces the total volume of solvent available to dissolve other solutes, potentially leading to uneven distribution on the ultrafiltration membrane. Therefore, this disclosure includes formulations in which, due to the size exclusion effect, For example The concentrations of histidine and / or arginine hydrochloride may differ from the amounts or ranges described herein.

[0079] During the manufacturing process of the formulations disclosed herein, the composition of the formulation may change. These changes may include the concentration of the active ingredient, the concentration of the excipients, and / or the pH of the formulation. Because any change in these parameters can affect the stability or potency of the pharmaceutical product, a demonstrated acceptable range (PAR) study is conducted to assess whether compositional changes within a defined range would affect the stability or potency of the antibody. Therefore, this disclosure includes formulations comprising an anti-human LAG-3 antibody that is stable and retains potency even with excipient concentration changes of up to 50%. For example, this document includes anti-LAG-3 antibody formulations in which the stability and potency of the formulation are not affected by changes of ±10%, ±20%, ±30%, ±40%, or ±50% in the concentrations of the antibody, sucrose, histidine buffer, and / or polysorbate.

[0080] Stability and viscosity of pharmaceutical preparations The pharmaceutical formulations disclosed herein generally exhibit a high level of stability. As used herein with reference to pharmaceutical formulations, the term "stable" means that the antibody in the pharmaceutical formulation retains an acceptable degree of chemical structure or biological function after storage under defined conditions. The formulation may be considered stable even if the antibody contained in the formulation does not retain 100% of its chemical structure or biological function after storage for a defined time. In some cases, retention of approximately 90%, approximately 95%, approximately 96%, approximately 97%, approximately 98%, or approximately 99% of the antibody structure or function after storage for a defined time may be considered "stable."

[0081] stability especially The percentage of natural antibodies retained in the formulation after storage at a specified temperature for a specified time can be determined. especially Size exclusion chromatography (SOC) can be used to measure the size exclusion chromatography method. For example The stability is determined by size exclusion ultra-high performance liquid chromatography (SE-UPLC), where "natural" means non-aggregated and non-degradable. As used herein, the phrase "acceptable degree of stability" means that at least 90% of the naturally occurring antibody is detectable in the formulation after storage at a given temperature for a given period of time. In some embodiments, at least about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% of the naturally occurring antibody is detectable in the formulation after storage at a defined temperature for a defined period of time. The defined period for subsequently measuring stability 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. When assessing stability, the defined storage temperature for the pharmaceutical formulation can be any temperature from about -80°C to about 45°C. For example Store at approximately -80°C, approximately -30°C, approximately -20°C, approximately 0°C, or approximately 4°C-8°C. C. Approximately 5°C, approximately 25°C, approximately 35°C, approximately 37°C, or approximately 45°C. For example, if a drug formulation is found to contain greater than approximately 95%, 96%, 97%, or 98% of natural antibodies by SE-UPLC after 6 months of storage at 5°C, it is also considered stable. If a drug formulation is found to contain greater than approximately 95%, 96%, 97%, or 98% of natural antibodies by SE-UPLC after 6 months of storage at 25°C, it is also considered stable. If a drug formulation is found to contain greater than approximately 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, or 98% of natural antibodies by SE-UPLC after 28 days of storage at 40°C, it is also considered stable. If a drug formulation is found to contain greater than approximately 96%, 97%, or 98% of natural antibodies by SE-UPLC after 12 months of storage at -20°C, it is also considered stable. A pharmaceutical preparation can also be considered stable if, after 12 months of storage at -30°C, greater than approximately 96%, 97%, or 98% of natural antibodies are detected by SE-UPLC. Similarly, a pharmaceutical preparation can be considered stable if, after 12 months of storage at -80°C, greater than approximately 96%, 97%, or 98% of natural antibodies are detected by SE-UPLC.

[0082] stability especially Stability can be measured by determining the percentage of antibodies that form aggregates after the formulation has been stored at a specified temperature for a specified time; the stability is inversely proportional to the percentage of aggregated antibodies. (Percentage of aggregated antibodies) especially Size exclusion chromatography (SOC) can be used to measure the size exclusion chromatography method. For example The stability is determined by size exclusion ultra-high performance liquid chromatography (SE-UPLC). As used herein, the phrase "acceptable degree of stability" refers to the detection of up to 5% of the antibody in aggregate form (also known as high molecular weight - HMW form) in the formulation after storage at a given temperature for a specified time. In some embodiments, acceptable degree of stability means that after storage at a given temperature for a specified time, up to about 5%, 4%, 3%, 2%, 1%, 0.5%, or 0.1% of the antibody in aggregate form can be detected in the formulation. The specified time for subsequently measuring stability can be 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. When assessing stability, the storage temperature of the pharmaceutical formulation can be any temperature from about -80°C to about 45°C. For exampleStore at approximately -80°C, approximately -30°C, approximately -20°C, approximately 0°C, or approximately 4°C-8°C. C. Approximately 5°C, approximately 25°C, approximately 35°C, approximately 37°C, or approximately 45°C. For example, if after 12 months of storage at 5°C, less than approximately 2%, 1%, 0.5%, or 0.1% of antibodies are detected in aggregated form, the drug formulation is considered stable. If after three months of storage at 25°C, less than approximately 4%, 3%, 2%, 1%, 0.5%, or 0.1% of antibodies are detected in aggregated form, the drug formulation is also considered stable. If after 28 days of storage at 45°C, less than approximately 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, or 0.5% of antibodies are detected in aggregated form, the drug formulation is also considered stable. If after three months of storage at -20°C, -30°C, or -80°C, less than approximately 3%, 2%, 1%, 0.5%, or 0.1% of antibodies are detected in aggregated form, the drug formulation is also considered stable.

[0083] stability especially This can be measured by determining the percentage of antibody that migrates during ion exchange in a fraction (“acidic form”) that is more acidic than the antibody’s major fraction (“major charge form”), where stability is inversely proportional to the acidic form of the antibody fraction. While not wishing to be bound by theory, antibody deamidation can result in antibodies carrying a greater negative charge and thus being more acidic relative to undeamidated antibodies (see [link to relevant documentation]). For example Robinson, N., ProteinDeamidation, PNAS , April 16, 2002, 99(8):5283-5288). Percentage of “acidified” antibodies. especially It can be obtained through ion exchange chromatography ( For exampleThe stability is determined by cation exchange ultra-high performance liquid chromatography (CEX-UPLC). As used herein, the phrase "acceptable level of stability" means that, after storage at a defined temperature for a defined time, up to 45% of the antibody in a more acidic form is detected in the formulation. In some embodiments, acceptable level of stability means that, after storage at a given temperature for a defined time, up to about 40%, 35%, 30%, 25%, 20%, 15%, 10%, 5%, 4%, 3%, 2%, 1%, 0.5%, or 0.1% of the antibody in an acidic form is detectable in the formulation. In one embodiment, acceptable level of stability means that, after storage at a given temperature for a defined time, less than 30%, 25%, 20%, 15%, 10%, 5%, 4%, 3%, 2%, 1%, 0.5%, or 0.1% of the antibody in an acidic form is detectable in the formulation. The specified timeframe for subsequently measuring stability may be 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. When assessing stability, the drug formulation can be stored at any temperature from about -80°C to about 45°C. For example Store at approximately -80°C, approximately -30°C, approximately -20°C, approximately 0°C, or approximately 4°C-8°C. C. Approximately 5°C, approximately 25°C, or approximately 45°C. For example, if after three months of storage at -80°C, -30°C, or -20°C, less than approximately 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, the pharmaceutical preparation can be considered stable. If, after six months of storage at 5°C, less than approximately 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 in a more acidic form, the pharmaceutical preparation is also considered stable. If, after six months of storage at 25°C, less than approximately 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 in a more acidic form, the pharmaceutical preparation is also considered stable. If, after storage at 45°C for 28 days, less than approximately 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 antibodies are detected in a more acidic form, then the pharmaceutical preparation can also be considered stable.

[0084] Other methods can be used to assess the stability of the formulations disclosed herein. For example Thermal stability was determined using differential scanning calorimetry (DSC), mechanical stability was determined using controlled stirring, and solution turbidity was determined using absorbance at approximately 350 nm or approximately 405 nm. For example, the OD of the formulation after storage at approximately 5°C to approximately 25°C for 6 months or longer was also considered. 405 The change was less than the OD of the formulation at time zero. 405Approximately 0.05 ( For example If the concentration is 0.04, 0.03, 0.02, 0.01 or less, then the formulation disclosed herein can be considered stable.

[0085] Measuring the antibody's biological activity or binding affinity to its target can also be used to assess stability. For example, if in For example Storage time limits at 5℃, 25℃, 45℃, etc. For example After 1 to 12 months, the binding affinity of the anti-LAG-3 antibody contained in the formulation to LAG-3 is at least 90%, 95%, or more of the binding affinity of the antibody before storage. Binding affinity can be determined by... For example Determined by ELISA or surface plasmon resonance. Bioactivity can be measured by LAG-3 activity assay. For example This is determined by contacting LAG-3-expressing cells with a formulation containing an anti-LAG-3 antibody. The binding of the antibody to these cells can be measured directly. For example Analysis can be performed using FACS. Alternatively, the downstream activity of the LAG-3 system can be measured in the presence of the antibody and compared with the activity of the LAG-3 system in the absence of the antibody. In some embodiments, LAG-3 may be endogenous in cells. In other embodiments, LAG-3 may be ectopically expressed in cells.

[0086] Other methods for assessing antibody stability in formulations are described in the examples presented below.

[0087] In some embodiments, the liquid pharmaceutical formulations of this disclosure may exhibit low to moderate viscosity levels. As used herein, “viscosity” can be “kinematic viscosity” or “absolute viscosity.” “Kinematic viscosity” is a measure of the resistance to flow of a fluid under the influence of gravity. When two fluids of equal volume are placed in the same capillary viscometer and allowed to flow under gravity, the viscous fluid will take 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 takes 400 seconds, the viscosity of the second fluid is twice that of the first fluid on a kinematic viscosity scale. “Absolute viscosity,” sometimes called dynamic viscosity or simple viscosity, is the product of kinematic viscosity and fluid density (absolute viscosity = kinematic viscosity x density). The dimension of kinematic viscosity is L. 2 / T, where L is length and T is time. Kinematic viscosity is typically expressed in centistokees (cSt). The SI unit for kinematic viscosity is mm. 2 / s, or 1 cSt. Absolute viscosity is expressed in centipoise (cP). The SI unit for absolute viscosity is millipascal-second (mPa-s), where 1 cP = 1 mPa-s.

[0088] As used herein, a low viscosity level will exhibit an absolute viscosity of less than about 20 centipoise (cP) relative to the fluid formulations of this disclosure. For example, if a formulation exhibits an absolute viscosity of about 20 cP, about 19 cP, about 18 cP, about 15 cP, about 12 cP, about 10 cP, about 9 cP, about 8 cP, or lower when measured using standard viscosity measurement techniques, the fluid formulations of this invention will be considered to have “low viscosity”. As used herein, with reference to the fluid formulations of this disclosure, a medium viscosity level will exhibit an absolute viscosity between about 35 cP and about 20 cP. For example, if the formulation exhibits an absolute viscosity of about 34 cP, about 33 cP, about 32 cP, about 31 cP, about 30 cP, about 29 cP, about 28 cP, about 27 cP, about 26 cP, about 25 cP, about 24 cP, about 23 cP, about 22 cP, about 21 cP, about 20 cP, about 19 cP, 18 cP, about 17 cP, about 16 cP, or about 15 cP when measured using standard viscosity measurement techniques, then the fluid formulation of the present invention will be considered to have “medium viscosity”.

[0089] As shown in the following examples, the inventors have surprisingly discovered that a high concentration of anti-human LAG-3 antibody can be obtained by formulating the antibody with about 1 mM to about 100 mM of arginine hydrochloride and about 5% or about 10% sucrose. For example Stable liquid formulations (approximately 50 mg / mL to 250 mg / mL). These formulations are stress-stable during treatment and are also stable when stored at temperatures ranging from 5°C to 45°C (as shown in this article).

[0090] Exemplary formulation According to one aspect of this disclosure, the pharmaceutical preparation is a stable, generally physiologically isotonic liquid preparation comprising: (i) a concentration of up to 250 mg / mL 45 mg / mL of human antibody that specifically binds to human LAG-3 ( For example (ii) at approximately pH 6.0 (REGN3767); (iii) an organic cosolvent that provides sufficient buffering at 0.3; (iv) a heat stabilizer for the sugar; and optionally (v) an amino acid that is used to maintain stability and injection-manageable viscosity in a convenient volume, for example in high-concentration formulations suitable for subcutaneous administration.

[0091] According to one implementation scheme, the stable pharmaceutical formulation comprises: (i) a concentration of up to 250 mg / ml (ii) 45 mg / mL of human IgG4 antibody specifically binding to human LAG-3, comprising HCDR1 of SEQ ID NO: 3, HCDR2 of SEQ ID NO: 4, HCDR3 of SEQ ID NO: 5, LCDR1 of SEQ ID NO: 6, LCDR2 of SEQ ID NO: 7, and LCDR3 of SEQ ID NO: 8; 2 mM histidine buffer at pH 6.0 0.3% buffer; (iii) 0.1% 0.05% w / v to 0.2% w / v (iv) 0.1% w / v polysorbate 80; (v) 5% ± 1% w / v to 10% ± 2% w / v sucrose; and (v) 1 mM 0.2 mM to 100 mM 20 mM arginine hydrochloride.

[0092] According to one embodiment, the stable pharmaceutical formulation comprises: (i) a concentration of 175 mg / ml (ii) 26.25 mg / mL of human IgG4 antibody specifically binding to human LAG-3, comprising HCDR1 of SEQ ID NO: 3, HCDR2 of SEQ ID NO: 4, HCDR3 of SEQ ID NO: 5, LCDR1 of SEQ ID NO: 6, LCDR2 of SEQ ID NO: 7, and LCDR3 of SEQ ID NO: 8; 2 mM histidine buffer at pH 6.0 (iii) 0.3% w / v 0.05% w / v polysorbate 80; (iv) 10% ± 2% w / v sucrose; and (v) 1 mM 0.2 mM to 100 mM 20 mM arginine hydrochloride.

[0093] According to one embodiment, the stable pharmaceutical formulation comprises: (i) a concentration of 150 mg / ml (ii) 22.5 mg / mL of human IgG4 antibody specifically binding to human LAG-3, comprising HCDR1 of SEQ ID NO: 3, HCDR2 of SEQ ID NO: 4, HCDR3 of SEQ ID NO: 5, LCDR1 of SEQ ID NO: 6, LCDR2 of SEQ ID NO: 7, and LCDR3 of SEQ ID NO: 8; 2 mM histidine buffer at pH 6.0 (iii) 0.1% w / v 0.05% w / v polysorbate 80; (iv) 10% ± 2% w / v sucrose; and (v) 1 mM 0.2 mM to 100 mM 20 mM arginine hydrochloride.

[0094] According to one embodiment, the stable pharmaceutical formulation comprises: (i) a concentration of 100 mg / mL A 15 mg / mL human IgG4 antibody that specifically binds to human LAG-3, comprising HCDR1 of SEQ ID NO: 3, HCDR2 of SEQ ID NO: 4, HCDR3 of SEQ ID NO: 5, LCDR1 of SEQ ID NO: 6, LCDR2 of SEQ ID NO: 7, and LCDR3 of SEQ ID NO: 8; (ii) 10 2mM histidine buffer at pH 6.0 0.3 underbuffer; (iii) 10% w / v (iv) 2% w / v sucrose; 0.05% polysorbate 80; and (v) 1 mM 0.2 mM to 100 mM 20mM arginine hydrochloride.

[0095] According to one embodiment, the stable pharmaceutical formulation comprises: (i) a concentration of 50 mg / mL 7.5 mg / mL of human IgG4 antibody specifically binding to human LAG-3, comprising HCDR1 of SEQ ID NO: 3, HCDR2 of SEQ ID NO: 4, HCDR3 of SEQ ID NO: 5, LCDR1 of SEQ ID NO: 6, LCDR2 of SEQ ID NO: 7, and LCDR3 of SEQ ID NO: 8; (ii) 10 2mM histidine buffer at pH 6.0 0.3 underbuffer; (iii) 10% w / v (iv) 2% w / v sucrose; 0.05% polysorbate 80; and (v) 20 mM 4 mM arginine hydrochloride.

[0096] According to one embodiment, the stable pharmaceutical formulation comprises: (i) a concentration of 25 mg / mL 3.75 mg / mL of human IgG4 antibody specifically binding to human LAG-3, comprising HCDR1 of SEQ ID NO: 3, HCDR2 of SEQ ID NO: 4, HCDR3 of SEQ ID NO: 5, LCDR1 of SEQ ID NO: 6, LCDR2 of SEQ ID NO: 7, and LCDR3 of SEQ ID NO: 8; (ii) 10 2mM histidine buffer at pH 6.0 0.3 underbuffer; (iii) 10% w / v (iv) 2% w / v sucrose; 0.05% polysorbate 80; and (v) 20 mM 4 mM arginine hydrochloride.

[0097] Further non-limiting examples of pharmaceutical formulations included in this invention are given elsewhere herein, including the working examples given below.

[0098] Containers and application methods The pharmaceutical formulations disclosed herein can be contained in any container suitable for storing pharmaceuticals and other therapeutic compositions. For example, the pharmaceutical formulations can be contained in sealed and sterile plastic or glass containers of a defined volume, such as vials, ampoules, syringes, cartridges, or bottles. Different types of vials can be used to contain the formulations disclosed herein, including… For example Transparent and opaque For exampleAmber-colored glass or plastic vial. Similarly, any type of syringe can be used to contain or administer the pharmaceutical preparations disclosed herein.

[0099] The pharmaceutical formulations disclosed herein can be contained in either a "standard tungsten" syringe or a "low tungsten" syringe. As will be understood by those skilled in the art, methods of manufacturing glass syringes typically involve using a heated tungsten rod to puncture the glass, creating a hole through which liquid can be drawn and expelled from the syringe. This method results in trace amounts of tungsten deposited on the inner surface of the syringe. Subsequent cleaning and other treatment steps can be used to reduce the amount of tungsten in the syringe. As used herein, the term "standard tungsten" means that the syringe contains 500 or more parts per billion (ppb) of tungsten. The term "low tungsten" means that the syringe contains less than 500 ppb of tungsten. For example, according to this disclosure, a low-tungsten injector 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 ppb or less of tungsten.

[0100] The rubber plunger used in the syringe and the rubber stopper used to seal the vial opening can be coated to prevent contamination of the drug contents of the syringe or vial, or to maintain their stability. Therefore, according to some embodiments, the pharmaceutical formulations of this disclosure can be contained in a syringe including a coated plunger, or in a vial sealed with a coated rubber stopper. For example, the plunger or stopper can be coated with a fluorocarbon film. Examples of coated stoppers or plungers suitable for use in vials and syringes containing pharmaceutical formulations of this disclosure are provided in... For example Mentions in U.S. Patents 4,997,423; 5,908,686; 6,286,699; 6,645,635; and 7,226,554 are incorporated herein by reference in their entirety. Certain exemplary coated rubber stoppers and plungers that may be used in the context of this disclosure are commercially available under the trade name “FluroTec®” from West Pharmaceutical Services, Inc. (Lionville, PA). FluroTec® is an example of a fluorocarbon coating used to minimize or prevent pharmaceutical products from adhering to rubber surfaces.

[0101] According to certain embodiments of this disclosure, a pharmaceutical preparation may be contained in a low-tungsten syringe comprising a fluorocarbon-coated piston.

[0102] Drug preparations can be administered via parenteral routes such as injection. For exampleThe pharmaceutical preparations disclosed herein may be administered to the patient via subcutaneous, intravenous, intramuscular, intraperitoneal, or transdermal, mucosal, nasal, pulmonary, or oral administration. Many reusable pen or auto-injector delivery devices may be used for subcutaneous delivery of these pharmaceutical preparations. Examples include, but are not limited to, AUTOPEN™ (Owen Mumford, Inc., Woodstock, UK), DISETRONIC™ pen (Disetronic Medical Systems, Bergdorf, Switzerland), HUMALOG MIX 75 / 25™ pen, HUMALOG™ pen, HUMALIN 70 / 30™ pen (Eli Lillyand Co., Indianapolis, IN), NOVOPEN™ I, II and III (Novo Nordisk, Copenhagen, Denmark), NOVOPEN JUNIOR™ (Novo Nordisk, Copenhagen, Denmark), BD™ pen (Becton Dickinson, Franklin Lakes, NJ), OPTIPEN™, OPTIPEN PRO™, OPTIPEN STARLET™ and OPTICLIK™ (sanofi-aventis, Frankfurt, Germany). Examples of disposable pen-type or auto-injector-type delivery devices that can be used for subcutaneous delivery of the pharmaceutical compositions disclosed herein include, but are not limited to, the SOLOSTAR™ pen (sanofi-aventis), FLEXPEN™ (Novo Nordisk), KWIKPEN™ (Eli Lilly), and SURECLICK. TM Automatic injectors (Amgen, Thousand Oaks, CA), PENLET TM (Haselmeier, Stuttgart, Germany), EPIPEN (Dey, LP) and HUMIRA TM pen (Abbott Labs, Abbott Park, IL).

[0103] This document also considers the use of microinfusion devices for delivering the pharmaceutical formulations of this disclosure. As used herein, the term "microinfusion device" refers to a subcutaneous delivery device designed to slowly administer a large volume of ( ) over an extended period of time (e.g., approximately 10, 15, 20, 25, 30 minutes or longer). For example Therapeutic agents (up to approximately 2.5 mL or more). See, for example US 6,629,949; US 6,659,982; and Meehan et al. , J. Controlled Release 46 :107-116(1996). Microinfusion sets are particularly suitable for delivering high concentrations of ( For example High doses of therapeutic proteins contained in viscous solutions (approximately 100, 125, 150, 175, 200 or higher mg / mL).

[0104] In some embodiments, the stable liquid pharmaceutical preparation of any of the foregoing aspects is contained in a sterile glass vial and administered as an IV infusion.

[0105] In one embodiment, the container is a 20 mL Type 1 clear borosilicate glass vial. In some embodiments, the container is a 2 mL, 5 mL, or 10 mL Type 1 borosilicate glass vial with a chlorobutyl stopper and FluroTec. ® coating.

[0106] In one embodiment, the liquid pharmaceutical formulation of this disclosure, comprising about 25 mg / mL or 50 mg / mL mAb1, is administered intravenously and may be contained in a glass vial. In some aspects, the glass vial contains 400 mg mAb1. In some aspects, the glass vial contains 800 mg mAb1.

[0107] In one embodiment, the liquid pharmaceutical formulation of this disclosure, containing about 100 mg / mL mAb1, is administered subcutaneously and may be contained in a glass vial.

[0108] In some embodiments, this disclosure provides an autoinjector comprising any of the liquid formulations described herein. In some embodiments, this disclosure provides an autoinjector comprising a stable liquid formulation comprising about 50 mg / mL, about 100 mg / mL, about 150 mg / mL, or about 175 mg / mL mAb1, about 10 mM histidine (pH about 6.0), about 5% sucrose or about 10% sucrose, about 20 mM arginine hydrochloride, and about 0.1% or about 0.2% polysorbate 80. In some aspects, the autoinjector contains 400 mg mAb1. In some aspects, the autoinjector contains 800 mg mAb1. In some aspects, the autoinjector contains 1600 mg mAb1.

[0109] In some embodiments, this disclosure provides a pre-filled syringe comprising any of the liquid formulations described herein. In some embodiments, this disclosure provides a pre-filled syringe comprising a stable liquid formulation comprising about 50 mg / mL, about 100 mg / mL, about 150 mg / mL, or about 175 mg / mL mAb1, about 10 mM histidine (pH about 6.0), about 5% sucrose or about 10% sucrose, about 20 mM arginine hydrochloride, and about 0.1% or about 0.2% polysorbate 80. In some embodiments, the syringe is a 1 mL or 2.25 mL long glass syringe filled with a 27-gauge thin-walled needle, a fluorocarbon-coated rubber plunger, and a rubber needle guard. In some aspects, the pre-filled syringe contains 400 mg mAb1. In some aspects, the pre-filled syringe contains 800 mg mAb1. In some aspects, the auto-injector contains 1600 mg mAb1.

[0110] In one implementation, it contains approximately 50 mg / mL The 7.5 mg / mL mAb1 liquid formulation is administered in a pre-filled syringe in a volume of approximately 2 mL. In some embodiments, the syringe is a 1 mL or 2.25 mL long glass syringe filled with a 27-gauge thin-walled needle, a fluorocarbon-coated rubber plunger, and a rubber needle guard. In one embodiment, the syringe is an OMPI 1 mL long glass syringe fitted with a 27-gauge needle, an FM27 rubber needle guard, and... Coated 4023 / 50 rubber plunger.

[0111] In one embodiment, it contains approximately 100 mg / mL A liquid formulation of 15 mg / mL anti-LAG-3 antibody is administered in a pre-filled syringe in a volume of approximately 2 mL. In one embodiment, the syringe is a 1 mL or 2.25 mL long glass syringe filled with a 27-gauge thin-walled needle, a fluorocarbon-coated rubber plunger, and a rubber needle guard. In another embodiment, the syringe is an OMPI 1 mL long glass syringe equipped with a 27-gauge needle, an FM27 rubber needle guard, and... Coated 4023 / 50 rubber plunger.

[0112] Therapeutic uses of pharmaceutical preparations The pharmaceutical formulation disclosed herein especiallyThis medicine may be used to treat, prevent, or improve any disease or condition associated with LAG-3 activity, including diseases or conditions mediated by LAG-3. Exemplary non-limiting diseases and conditions that may be treated or prevented by administration of pharmaceutical formulations disclosed herein include viral infections, autoimmune diseases, and various cancers. For example Brain cancer, lung cancer, prostate cancer, colorectal cancer, head and neck cancer, skin cancer, various blood cancers, and endometrial cancer.

[0113] Example The following examples are provided to provide a complete disclosure and description of how to prepare and use the methods and compositions of the present invention to those skilled in the art, and are not intended to limit the scope of what the inventors consider to be their invention. Efforts have been made to ensure the accuracy of the figures used (e.g., amounts, temperatures, etc.), but some experimental errors and deviations should be taken into account. Unless otherwise specified, parts are molar parts, molecular weights are average molecular weights, temperatures are in degrees Celsius, and pressures are atmospheric pressure or close to atmospheric pressure.

[0114] Example 1: Development of anti-LAG-3 antibody formulation The goal of formulation activities is to develop formulations with the following properties: ● Liquid formulation with anti-LAG-3 antibody concentrations sufficient to deliver doses of 50 mg, 400 mg, 800 mg, 1600 mg or more; ● When using common diluents, For example A near-isotonic preparation that is stable when diluted with 0.9% sodium chloride injection or 5% glucose injection, for intravenous infusion; ● A formulation compatible with and stable within the type 1 clear glass vial and standard serum stopper used as packaging; and ● Sterile pharmaceutical (DP) solutions that support long-term stability; ○ Formulations that minimize the high molecular weight (HMW) of antibodies when subjected to treatment and heat stress; ○ A formulation that minimizes changes in the relative distribution of antibody-carrying substances during heat stress; and ○ Formulations that retain biological activity when subjected to treatment and heat stress.

[0115] Throughout the formulation development process, three main protein stress conditions (representing extreme handling conditions that the antibody drug product would not experience during processing, manufacturing, transportation, storage, and labeling) were employed to develop and optimize the antibody formulation and to evaluate the effects of potential real-world stress on the stability of the drug product. These stress conditions included: ● Stir (vortex) the protein solution at room temperature. Vortexing in the glass vial surpasses stirring in protein handling and manufacturing processes.

[0116] ● At high temperatures (37°C, 40°C) relative to the recommended DP storage conditions (2°C-8°C). Incubate the protein solution at ℃ or 45℃.

[0117] ● The protein is subjected to multiple freeze-thaw cycles. Since the protein will undergo at least one freeze-thaw cycle during the manufacturing process of DP, the multiple freeze-thaw cycles simulate and exceed the actual stress that the protein is expected to experience.

[0118] Other properties of the formulation will become clear from the description in this article.

[0119] Anti-LAG-3 antibody: Anti-LAG-3 antibodies are described in US 10,358,495, which is incorporated herein in its entirety. The exemplary antibody used in the following examples is a fully human anti-LAG-3 antibody referred to as “REGN3767” or “fenolinumab”, comprising a heavy chain complementarity-determining region (HCDR) within the heavy chain variable region (HCVR) amino acid sequence of SEQ ID NO: 1 and a light chain complementarity-determining region (LCDR) within the light chain variable region (LCVR) amino acid sequence of SEQ ID NO: 2; or comprising the HCVR / LCVR amino acid sequence pair of SEQ ID NO: 1 / 2; or comprising the heavy chain and light chain CDR sequences of SEQ ID NO: 3-8; or the heavy chain CDR within the heavy chain amino acid sequence of SEQ ID NO: 9 and the light chain CDR within the light chain amino acid sequence of SEQ ID NO: 10; or the heavy chain amino acid sequence of SEQ ID NO: 9 and the light chain amino acid sequence of SEQ ID NO: 10; referred to herein as “mAb1”. The antibody has an approximate molecular weight of about 145 kDa, and the heavy chain has an IgG4 isotype constant region.

[0120] In the first human study, mAb1 was evaluated for intravenous (IV) and subcutaneous (SC) administration. Initially, a single dual-purpose lyophilized formulation was developed, enabling the lyophilized mAb1 drug product (DP) to be reconstituted with sterile water for injection to achieve concentrations of 50 mg / mL mAb1 for IV infusion or 100 mg / mL mAb1 for SC injection.

[0121] Formulation development activities included evaluating buffers, pH values, organic cosolvents, surfactants, and sucrose (as a heat stabilizer) to identify excipients that enhance protein stability. Buffers and pH were screened early in the formulation development process. Three buffers were tested: acetate (pH 4.5–5.5), histidine (pH 5.5–6.50), and phosphate (pH 6.0–7.0). 10 mM histidine at pH 6.0 showed the best stability under accelerated conditions and was selected for continued use. Three surfactants / cosolvents were tested: polysorbate 20, polysorbate 80, and PEG3350. PS80 was selected for the formulation. The results from these studies were used to develop a stable lyophilized formulation containing 50 mg / mL mAb1, 10 mM histidine (pH 6.0), 5% (w / v) sucrose, and 0.1% (w / v) polysorbate 80, suitable for reconstitution into liquid form prior to clinical use.

[0122] For later-stage clinical development, formulation development studies are conducted to develop a liquid DP formulation of mAb1. Information obtained during early-stage formulation development forms the basis for determining the buffer and pH in later-stage formulations.

[0123] Buffer and pH The effects of buffers and pH on thermal stability were examined in liquid formulations by incubating the 25 mg / mL mAb1 formulation at 45 °C for 28 days in a range of buffer systems with different pH ranges (Table 1). A 10 mM L-histidine buffer with pH 6.0 was chosen as the formulation buffer because it provides optimal overall protein stability in terms of the formation of low molecular weight (LMW) and high molecular weight (HMW) substances, as well as charge variants.

[0124] Table 1: Effect of pH on the stability of 25 mg / mL mAb1 after incubation at 45°C for 14 days

[0125] a If it is transparent to slightly milky white, contains virtually no visible particles, and is colorless to pale yellow, then it is acceptable. b The change in purity is relative to the starting material. In all formulations, the starting material (unincubated) contained ≥98.5% of the natural peak (by SE-UPLC) and ≥57.2% of the main peak (by CEXUPLC).

[0126] CEX, cation exchange; DS, active pharmaceutical ingredient; HMW, high molecular weight; LMW, low molecular weight; OD, optical density; Ph. Eur, European Pharmacopoeia; RP, reversed-phase; SE, size exclusion; UPLC, ultra-high performance liquid chromatography; USP, United States Pharmacopeia. Surfactant The effects of the selected surfactants / cosolvents (polysorbate 20, polysorbate 80, and PEG 3350) on the agitation stress of 25 mg / mL mAb1 were enhanced. All surfactants / cosolvents sufficiently protected the protein from agitation-induced instability, and no increase in HMW species was observed. Polysorbate 80 was selected as the surfactant for the DP formulation because it stabilized the protein against agitation stress and had proven safety for use in monoclonal antibody formulations (Table 2).

[0127] Table 2: Effects of surfactants on the stability of 25 mg / mL mAb1 after vortexing (120 minutes at 1000 RPM)

[0128] a It is qualified if it is transparent to slightly milky white, essentially free of visible particles, and colorless to light yellow.

[0129] b It is reported as the purity change relative to the starting material. In all formulations, the starting material (unincubated) contained ≥97.7% of the main peak (by SE-UPLC) and ≥60.5% of the main peak (by CEX UPLC).

[0130] CEX, cation exchange; DS, drug substance; HMW, high molecular weight; LMW, low molecular weight; OD, optical density; Ph. Eur, European Pharmacopoeia; RP, reverse phase; SE, size exclusion; UPLC, ultra performance liquid chromatography; USP, United States Pharmacopoeia To optimize the concentration of polysorbate 80, agitation studies were conducted using a range of polysorbate 80 concentrations (0.01% to 0.20% w / v). The results showed that mAb1 containing 0.10% (w / v) polysorbate 80 fell within the stable region of the characteristic range, with sufficient tolerance on either side to withstand changes in the polysorbate 80 concentration( Figure 1 ). The 0.10% (w / v) concentration of polysorbate 80 was consistent with the final concentration of polysorbate 80 in the reconstituted IV formulation used in the initial clinical trial studies.

[0131] Thermal stabilizer Stabilizers such as sucrose are usually added to antibody formulations to increase the thermal stability of the protein. The effect of sucrose was investigated by placing 25 mg / mL formulations with or without 5% (w / v) sucrose under accelerated heat stress conditions (Table 3). After incubation at 45 °C for 28 days, the HMW species increased by 2.0% and 1.5% in the control and sucrose formulations, respectively.

[0132] Table 3: Effect of Sucrose on the Stability of 25 mg / mL mAb1 Incubated at 45 °C for 28 Days

[0133] a It is qualified if it is transparent to slightly milky white, basically free of visible particles, and colorless to light yellow.

[0134] b Report the purity change relative to the starting material. In both formulations, the starting material (unincubated) contains ≥97.7% of the main peak (by SE-UPLC) and ≥60.5% of the main peak (by CEX UPLC).

[0135] CEX, cation exchange; DS, drug substance; HMW, high molecular weight; LMW, low molecular weight; OD, optical density; Ph. Eur, European Pharmacopoeia; RP, reverse phase; SE, size exclusion; UPLC, ultra performance liquid chromatography; USP, United States Pharmacopoeia Different stabilizers and concentrations were evaluated using mAb1 drug substance (DS) to develop a liquid formulation of 50 mg / mL mAb1. As part of this study, the following stabilizers and combinations were evaluated: 5% (w / v) sucrose, 5% (w / v) sucrose and 20 mM arginine hydrochloride, and 10% (w / v) sucrose and 20 mM arginine hydrochloride. A summary of the stability results is presented in Table 4. Based on SE-UPLC and CEX-UPLC analyses, all three formulations confer similar levels of protein stability to mAb1. Among the three formulations, 10% (w / v) sucrose and 20 mM arginine hydrochloride produce the best overall protein stability levels in terms of the formation of low molecular weight (LMW) species, high molecular weight (HMW) species, and charge variants. The addition of L-arginine hydrochloride has no negative impact on stability. L-arginine hydrochloride was included in the 50 mg / mL formulation because high concentrations of drug substance (DS) were targeted to support the development of IV formulations and subcutaneous (SC) formulations at higher mAb1 concentrations. To manufacture ≥175 mg / mL mAb1 DS, L-arginine hydrochloride was added as a viscosity reducer. Table 5 describes the effect of the stabilizer in the high-concentration formulation, and Figure 4 shows the viscosity changes at different antibody concentrations and storage temperatures. The anti-LAG3 antibody formulation containing 175 mg / mL antibody in 10 mM histidine, 5% sucrose, 70 mM arginine HCl, 0.1% PS80 (pH 6) and maintained at 20 °C has a viscosity not exceeding 25 cP.

[0136] Select a formulation containing 50 mg / mL mAb1, 10 mM histidine, 10% (w / v) sucrose, 20 mM arginine hydrochloride, and 0.1% polysorbate 80 (pH 6.0) as the mAb1 DP formulation.

[0137] Table 4: Effect of stabilizers on the stability of 50 mg / mL mAb1 incubated at 40 °C for 28 days

[0138] a Reported as the change in purity relative to the starting material. In all formulations, the starting material (non-incubated) contained ≥98.5% of the native peak (by SE-UPLC) and ≥57.3% of the main peak (by CEX-UPLC).

[0139] CEX, cation exchange; DS, drug substance; HMW, high molecular weight; LMW, low molecular weight; OD, optical density; RP, reverse phase; SE, size exclusion; UPLC, ultra-performance liquid chromatography Table 5: Effect of stabilizers on the stability of 175 mg / mL REGN3767 incubated at 2 - 8 °C for 36 months

[0140] a Reported as the change in purity relative to the starting material. In all formulations, the starting material (non-incubated) contained ≥98.0% of the native peak (by SE-UPLC) and ≥57.3% of the main peak (by CEX-UPLC).

[0141] b It is qualified if it is transparent to slightly milky white, substantially free of visible particles, and colorless to light yellow.

[0142] CEX, cation exchange; DS, drug substance; HMW, high molecular weight; LMW, low molecular weight; OD, optical density; SE, size exclusion; UPLC, ultra-performance liquid chromatography Example 2: Exemplary Formulations In certain embodiments, mAb1 is formulated into an aqueous buffer formulation containing 5 mg / ml 0.75 mg / ml to 250 mg / ml 45.0 mg / ml of mAb1, 10 mM 2mM of histidine buffer, 0.05% 0.05% to 0.2% 0.1% w / v of polysorbate, 1% 0.2% to 15% 3% w / v sucrose and 1 mM 0.2 mM to 100 mM 20 mM arginine hydrochloride, pH 6.0 0.3.

[0143] Exemplary formulations include: ● A stable pharmaceutical preparation comprising: 25 mg / ml 3.75 mg / mL mAb1, 10 ± 2 mM histidine buffer, 0.1% ± 0.05% w / v polysorbate 80, 5% ± 1% or 10% ± 2% w / v sucrose and 20 mM ± 4 mM arginine hydrochloride, pH 6.0 ± 0.3.

[0144] ● A stable pharmaceutical preparation comprising: 25 mg / ml 3.75 mg / mL mAb1, 10 ± 2 mM histidine buffer, 0.1% ± 0.05% w / v polysorbate 80, 5% ± 1% w / v sucrose, pH 6.0 ± 0.3.

[0145] ● A stable pharmaceutical preparation comprising: 50 mg / ml 7.5 mg / mL mAb1, 10 ± 2 mM histidine buffer, 0.1% ± 0.05% w / v polysorbate 80, 5% ± 1% or 10% ± 2% w / v sucrose and 20 mM ± 4 mM arginine hydrochloride, pH 6.0 ± 0.3.

[0146] ● A stable pharmaceutical preparation comprising: 50 mg / ml 7.5 mg / mL mAb1, 10 ± 2 mM histidine buffer, 0.1% ± 0.05% w / v polysorbate 80, 5% ± 1% w / v sucrose, pH 6.0 ± 0.3.

[0147] ● A stable pharmaceutical preparation comprising: 100 15 mg / mL mAb1, 10 ± 2 mM histidine buffer, 0.1% ± 0.05% w / v polysorbate 80, 10% ± 2% w / v sucrose and 10 mM... 2 mM to 100 mM 20 mM arginine hydrochloride, pH 6.0±0.3.

[0148] ● A stable pharmaceutical preparation comprising: 150 22.5 mg / mL mAb1, 10 ± 2 mM histidine buffer, 0.2% ± 0.1% w / v polysorbate 80, 10% ± 2% w / v sucrose and 10 mM... 2 mM to 100 mM 20 mM arginine hydrochloride, pH 6.0±0.3.

[0149] ● A stable pharmaceutical preparation comprising: 175 37.5 mg / mL mAb1, 10 ± 2 mM histidine buffer, 0.1% ± 0.05% w / v polysorbate 80, 10% ± 2% w / v sucrose and 10 mM... 2 mM to 100 mM 20 mM arginine hydrochloride, pH 6.0±0.3.

[0150] Example 3: A method for evaluating formulation stability The following assays are used to assess formulation stability: ● Visually inspect color and appearance ● pH ● Turbidity measured by increase in OD at 405 nm via Micro-Flow Imaging ™ Subvisible particle analysis performed by (MFI) and photoresist analysis via HIAC ● Protein concentration measurement using reversed-phase ultra-high performance liquid chromatography (RP-UPLC) ● The purity of DP was assessed using the following assays: ○ Size exclusion ultra-high performance liquid chromatography (SE-UPLC) ○ Reducing and non-reducing microchip capillary electrophoresis-sodium dodecyl sulfate (MCE-SDS) ● The following measurements were used to determine charge variant analysis: ○ Cation exchange UPLC (CEX-UPLC) ○ Imaging capillary isoelectric focusing (iCIEF) Charge variants are reported as percentages for regions 1, 2, and 3. Region 1 corresponds to acidic substances eluted before the main peak; region 2 corresponds to the main peak; and region 3 corresponds to basic substances eluted after the main peak.

[0151] ● Assessing potency through bioassays: Determine the relative potency of each sample through bioassays, and define it as: (IC) 50 Reference Sample / IC50 (Sample) × 100%. The potency of the sample for storage stability measurement must be within 50%-150% of the potency of the reference standard.

[0152] The physical stability of a formulation refers to properties such as color, appearance, pH, turbidity, and protein concentration. The presence of visible particles in the solution can be detected by visual inspection. If the solution is clear to slightly milky white, substantially free of visible particles, and colorless to pale yellow, the solution passes visual inspection. Additionally, turbidity measured by OD at 405 nm can also be used to detect particles in the solution. An increase in OD at 405 nm may indicate the presence of particles, an increase in milkiness, or a color change in the sample. MFI is used to measure sub-visible particles ≥2 µm in size. The protein concentration of mAb1 is measured by RP-UPLC assay and reported as a percentage of protein recovery relative to the starting material. In RP-UPLC assays, mAb1 elutes as a single peak from the RP column. The protein concentration is determined by the total peak area of ​​mAb1 by comparing the protein concentration to a calibration curve generated using mAb1 standards. The recovery percentage is calculated based on the percentage of the measured protein concentration relative to the starting protein concentration.

[0153] Chemical stability refers to the formation of covalently modified forms of proteins (e.g., covalent aggregates, cleavage products, or charge variants) and non-covalently modified forms (e.g., non-covalent aggregates). Higher and lower molecular weight degradation products can be separated from native mAb1 using SE-UPLC and MCE-SDS methods. The percentage of degraded mAb1 in SE-UPLC and MCE-SDS methods is calculated as the ratio of the area of ​​all non-native peaks to the total area of ​​all mAb1 peaks. Charge variants of mAb1 are resolved using CEX-UPLC and iCIEF. In CEX-UPLC, peaks with retention times earlier than the main peak are labeled "acidic" peaks; peaks with retention times later than the main peak are labeled "basic" peaks. In iCIEF, peaks with a pI lower than the main peak are labeled "acidic" peaks, while peaks with a pI higher than the main peak are labeled "basic" peaks.

[0154] Example 4: Stability of the 50 mg / mL formulation when stored upside down A formulation containing 50 mg / mL mAb1, 10 mM L-histidine, 10% (w / v) sucrose, 20 mM L-arginine hydrochloride, and 0.1% polysorbate 80 (pH 6.0) was selected as the liquid mAb1 formulation for co-administration with cimiprilmab and for the manufacture of mAb1. As shown in Table 6, after storage at 5°C (2–8°C) for 42 months, no significant changes in physical or chemical stability were observed in any monitored properties, demonstrating the suitability of this formulation.

[0155] Table 6: Stability of mAb1 drug products at 50 mg / mL when stored upside down at 2-8°C

[0156]

[0157] a Data from release test A 280， Absorbance at 280 nm; C2P1, Cell Line 2 Process 1; CCI, Container Closure Integrity; EU, Endotoxin Unit; HMW, High Molecular Weight; iCIEF, Imaging Capillary Isoelectric Focusing; LEFVP, Liquid Essentially Free of Visible Particles; LMW, Low Molecular Weight; LO, Optical Obscuration; MCE, Microchip Capillary Electrophoresis; MFI, MP, main peak; NDD, dye not detected; NGHC, non-glycosylated heavy chain; Not > BY2; NR, not required; PGMP, partially glycosylated main peak; Ph.Eur., European Pharmacopoeia; SE, size exclusion; UPLC, ultra-high performance liquid chromatography; USP, United States Pharmacopeia The DP solution contained 50 mg / mL mAb1, 10 mM histidine (pH 6.0), 10% (w / v) sucrose, 20 mM arginine hydrochloride, and 0.1% (w / v) polysorbate 80. The following conclusions can be drawn from the stability study: mAb1 DP is stable when stored at 2°C to 8°C for at least 42 months.

[0158] The main degradation pathways identified are the formation of high molecular weight components and charge variants.

[0159] Overfeeding The formulation does not include overfilling; however, it does include slight overfilling to ensure the correct volume is drawn from the vial.

[0160] One vial of DP contains 16.5 mL of 50 mg / mL mAb1. Each vial contains 0.5 mL of excess filler, sufficient to accurately draw 16.0 mL (800 mg mAb1) of DP from the vial.

[0161] This overfill is not designed to compensate for losses during manufacturing, degradation during manufacturing, degradation during storage (shelf life), or to extend shelf life. This overfill is only intended to ensure accurate dispensing of 16.0 mL.

[0162] Example 5: Stability Study of 400 mg mAb1 Liquid Drug Product (DP) This report summarizes the stability studies of mAb1 liquid pharmaceutical product (DP) (50 mg / mL mAb1, 10 mM L-histidine, 20 mM L-arginine hydrochloride, 10% (w / v) sucrose, 0.1% (w / v) polysorbate 80, pH 6.0) in 10R 1 type borosilicate glass vials with a filling volume of 8.5 mL (and a draw volume of 8.0 mL).

[0163] DP was incubated under storage, accelerated, and stress conditions. Accelerated and stress conditions were selected to simulate conditions that DP would not experience during manufacturing and processing, and to elucidate the degradation pathway of mAb1 DP.

[0164] This study was conducted in an upright orientation under the conditions described below. The duration of the following stress / storage conditions and time points was examined in the mAb1 stability study.

[0165] ● Long-term storage conditions: 2-8℃, up to 60 months ● Thermal acceleration conditions: 25℃ / 60% RH, up to 6 months ● Heat stress conditions: 40℃ / 75% RH, up to 3 months ● Physical stress conditions: stirring by vortexing at 1000 RPM for 120 minutes. ● Physical stress conditions: freezing and thawing at -30°C for up to 4 cycles. The stability of mAb1 was analyzed using the following methods: ● Color and appearance ● pH ● Protein concentration obtained via Solo VPE ● Purity (SE-UPLC method) ● Charge heterogeneity (iCIEF method) ● Purity obtained via MCE (reduced and non-reduced) - all time points except T during stirring. 结束(Final time point) and F / T ● Particulate matter obtained via HIAC 0, 3, and 6 m at 5℃ and annual TP (time points) 3m and T at 25℃ / 60% RH and 40℃ / 75% RH 结束 Stirring T 结束 and F / T ● Particulate matter obtained through MFI 0, 6m and annual TP at 5℃ 3m and T at 25℃ / 60% RH and 40℃ / 75% RH 结束 Stirring T 结束 and F / T ● Polysorbate 80 concentration obtained by CAD-UPLC 0, 6m and annual TP at 5℃ T at 25℃ / 60% RH and 40℃ / 75% RH 结束 Stirring T 结束 and F / T ● Potency obtained through bioassays - annual TP at 0, 6m and 5°C Results of long-term storage stability studies Twelve months of stability data were available for mAb1 DP. mAb1 DP is physically and chemically stable when stored at storage conditions (2–8 °C) for at least 12 months. As shown in Table 7, no significant changes in stability were detected in any monitored property at 2–8 °C. These results indicate that mAb1 DP is stable for at least 12 months under storage conditions.

[0166] Results of thermal acceleration and stress stability studies Table 8 provides the analytical results of mAb1 DP after incubation under accelerated and stress heat conditions.

[0167] After incubation at 25°C / 60% RH for one month, no significant changes in physical or chemical stability were detected, indicating that mAb1 DP can be exposed at room temperature for up to one month. As shown in Table 9, the formation of region 3 was detected by iCIEF after incubation at 25°C / 60% RH for six months. See also Table 8.

[0168] After incubation at 40 °C / 75% RH for 3 months, the obvious formation of HMW substances was determined by SE-UPLC, the obvious formation of LMW substances was determined by MCE, and the obvious formation of charge variants was determined by iCIEF (as shown in Table 10). For details of charge variants under stress conditions, see Table 8. The incubation results under accelerated and stress conditions showed that the increased formation of HMW substances, LMW substances and charge variants was the main degradation pathway of mAb1 DP.

[0169] Table 7: Stability of mAb1 liquid drug product stored at 2 - 8 °C

[0170] a It is qualified if it is transparent to slightly milky white, essentially free of visible particles, and colorless to light yellow.

[0171] b The influence of bubbles was minimized by filtering out particles with an aspect ratio > 0.85.

[0172] HMW, high molecular weight; iCIEF, imaging capillary isoelectric focusing; LEFVP, liquid essentially free of visible particles; LMW, low molecular weight; LO, light obscuration method; MCE, microchip capillary electrophoresis; MFI, Micro-Flow Imaging™; MP, main peak; Not > BY2, color intensity not deeper than reference solution BY2; Not > IV, not more turbid than reference suspension IV; NR, not required for each protocol; Ph. Eur., European Pharmacopoeia; SE, size exclusion; UPLC, ultra-high performance liquid chromatography; USP, United States Pharmacopoeia Table 8: Stability of mAb1 liquid drug product incubated at 25 °C / 60% RH and 40 °C / 75% RH

[0173] a It is qualified if it is transparent to slightly milky white, essentially free of visible particles, and colorless to light yellow.

[0174] b The influence of bubbles was minimized by filtering out particles with an aspect ratio > 0.85.

[0175] HMW, High Molecular Weight; iCIEF, Imaging Capillary Isoelectric Focusing; LEFVP, Liquid Essentially Free from Visible Particulates; LMW, Low Molecular Weight; LO, Light Obscuration; MCE, Microchip Capillary Electrophoresis; MFI, Micro-Flow Imaging™; MP, Main Peak; Not > BY2, Not Deeper in Color Intensity than the Reference Solution BY2; Not > IV, Not More Turbid than the Reference Suspension IV; NR, Not Required for Each Protocol; Ph. Eur., European Pharmacopoeia; SE, Size Exclusion; UPLC, Ultra Performance Liquid Chromatography; USP, United States Pharmacopoeia Table 9: Summary of Stability Changes of mAb1 DP (400 mg) after Incubation at 25°C / 60% RH for 6 Months

[0176] Table 10: Summary of Stability Changes of mAb1 DP (400 mg) after Incubation at 40°C / 75% RH for 3 Months

[0177] Results of Stirring and Freezing / Thawing Stability Studies The analytical results of mAb1 DP after vortexing and freezing / thawing stress conditions are provided in Table 11. No significant changes in physical or chemical stability were detected after vortexing for 120 minutes and 4 cycles of freezing (-30°C) and thawing (room temperature). These results indicate that mAb1 DP (400 mg) is stable to stirring stress and up to 4 cycles of freezing and thawing.

[0178] Table 11: Stability of mAb1 Liquid Drug Product - Effects of Stirring and Freezing / Thawing

[0179] a It is acceptable if it is transparent to slightly milky white, essentially free from visible particulates, and colorless to light yellow.

[0180] b Minimize the effect of air bubbles by filtering out particles with an aspect ratio > 0.85.

[0181] HMW, High Molecular Weight; iCIEF, Imaging Capillary Isoelectric Focusing; LEFVP, Liquid with virtually no visible particles; LMW, Low Molecular Weight; LO, Optical Obscuration; MCE, Microchip Capillary Electrophoresis; MFI, Micro-Flow Imaging™; MP, Main Peak; Not > BY2, Color intensity is not deeper than reference solution BY2; Not > IV, Color intensity is not more turbid than reference suspension IV; NR, Not required for each protocol; Ph. Eur., European Pharmacopoeia; SE, Size Exclusion; UPLC, Ultra-High Performance Liquid Chromatography; USP, United States Pharmacopeia. in conclusion At the time of writing, 12 months of stability data were available from a stability study of mAb1 (400 mg strength). This stability study evaluated the stability of mAb1 DP under long-term, accelerated, and stress conditions. No significant changes in mAb1 DP stability were detected by any monitoring property during long-term storage at 2–8°C for up to 12 months. Furthermore, mAb1 DP was physically and chemically stable when subjected to vortexing and freeze-thaw stress.

[0182] The results of this study support the recommended long-term storage conditions for mAb1 DP at 2–8 °C. DP stability data support storage for up to one month under accelerated conditions at up to 25 °C / 60% RH.

[0183] Example 6: Stability Study of 50 mg / mL mAb1 Liquid Drug Product (DP) Formulation The following formulation stability study was conducted on mAb1 as a liquid pharmaceutical product (DP). Liquid DP contains 50 mg / mL mAb1 in 10 mM L-histidine, 10% (w / v) sucrose, 0.1% (w / v) polysorbate 80, and 20 mM L-arginine hydrochloride (pH 6.0), and is stored in 20 mL Type 1 borosilicate glass vials. Because any changes in these formulation factors could affect the stability of DP, a formulation stability study was conducted to assess whether changes in the DP formulation composition within the defined demonstrated acceptable range (PAR) would have a meaningful impact on the long-term stability of mAb1 DP.

[0184] All formulations were prepared and filled into 20 mL Type 1 glass vials with a filling volume of 16.5 mL, and their long-term storage stability at 2–8 °C was assessed. Table 12 summarizes the formulation parameters and their ranges examined in this study, and Table 13 shows the formulations examined in the stability studies. The storage conditions for this study are shown in Table 14.

[0185] Table 12: Summary of Formulation Parameters and Formulation Range

[0186] PAR, indicating acceptable range Table 13: Formulations tested in PAR studies

[0187] Table 14: Research Conditions

[0188] a 250 RPM on the orbital oscillator The analytical testing plan uses the following methods to assess the stability of the formulations in Table 13: ● Osmotic pressure obtained through vapor pressure ● Viscosity ● Color and appearance ● pH ● Protein content obtained through Solo VPE ● Purity content obtained through SEC ● Purity obtained through MCE (non-reducing and reducing forms) FDS : -30℃ and -20℃ at t=0, 6 and 12m DP : t=0, 6 and 12m at 5℃, 6m at 25℃ / 60% RH, 3m at 40℃ / 75% RH, and stirring at the last time point. ● Charge heterogeneity obtained through iCIEF FDS : -30℃ and -20℃ at t=0, 3, 6 and 12m DP : t=0, 3, 6 and 12m at 5℃, 1 and 6m at 25℃ / 60% RH, 3m at 40℃ / 75% RH, and stirring at the last time point. ● Polysorbate obtained by CAD-UPLC FDS : -30℃ and -20℃ at t=0, 3, 6 and 12m DP : t=0, 3, 6 and 12m at 5℃, 6m at 25℃ / 60% RH, 3m at 40℃ / 75% RH, and stirring at the last time point. ● Particulate matter obtained through MFI FDS : -30℃ and -20℃ at t=0, 3, 6 and 12m DP: t=0, 3, 6 and 12m at 5℃, 6m at 25℃ / 60% RH, 3m at 40℃ / 75% RH, and stirring at the last time point. ● Particulate matter obtained via HIAC DP : t=0, 3, 6 and 12m at 5℃, 6m at 25℃ / 60% RH, 3m at 40℃ / 75% RH, and stirring at the last time point. Long-term DP stability study results The mAb1 DP formulations listed in Table 15 were stored at 2–8°C. The effects of different protein concentrations, sucrose concentrations, pH, histidine concentrations, and polysorbate 80 concentrations on the long-term stability of mAb1 DP were examined. The quality attributes assessed were molecular weight variant formation as determined by SE-UPLC and MCE, charge variant formation as determined by iCIEF, appearance, pH, protein recovery, and subvisible particle levels as determined by HIAC and MFI. Figure 2 The results show the stability observed after storage at 2-8°C for more than 12 months.

[0189] These results indicate that after 12 months of storage at 2–8°C, altering the formulation composition within the test range does not have a meaningful impact on the stability or quality of mAb1.

[0190] ● No changes in color, sediment, or visible particles were detected during visual inspection.

[0191] ● No significant changes in protein recovery were observed using Solo VPE.

[0192] ● The pH of the formulation is stable.

[0193] ● No significant increase in subvisible particles was observed by HIAC.

[0194] All values ​​are below the acceptable limits set by USP <787>.

[0195] ● No significant changes were observed in HMW or LMW levels as determined by SE-UPLC or MCE.

[0196] ● As determined by iCIEF, no significant difference was observed in the levels of acidic (region 1) or basic (region 3) charge variants.

[0197] Data indicate that the 50 mg / mL mAb1 DP formulation (800 mg DP formulation) is robust across all assessed quality attributes within the tested formulation composition range.

[0198] Results of accelerated and stress (thermal) DP stability studies Figure 3 The results summarize the stability over time (accelerated and stress thermal stability studies) at 25°C / 60% RH.

[0199] No significant changes in stability were observed when the formulation was incubated at 25°C / 60%RH for one month, indicating that the 50 mg / mL C2P1 DP formulation (800 mg present) within the tested formulation composition range can be exposed to room temperature for a short period. Data not shown.

[0200] After incubation at 40°C / 75% RH for 3 months, significant formation of HMW, LMW, and charge variants was detected. Results under accelerated conditions indicate that the formation of HMW, LMW, and charge variants is a major degradation pathway for 50 mg / mL mAb1 DP within the tested formulation composition range. Data not shown.

[0201] Results of DP stability study by stirring Of the 14 formulations examined in the PAR stress (non-thermal) stability study, all were physically and chemically stable after stirring (orbiting at 250 RPM at ambient temperature) for 48 hours.

[0202] Osmotic pressure and viscosity results The osmotic pressure and viscosity of the PAR formulations described in Table 13 were measured and are summarized in Table 15. The viscosity range at 20°C was from about 2.0 CP to about 3.0 CP. The osmotic pressure ranged from about 330 to about 530 mOsm / kg.

[0203] Table 15: Osmotic pressure and viscosity of mAb1 formulation pharmaceutical products

[0204] FDS stability results Prior to DP filling, the bulk FDS used to generate the mAb1 PAR formulation DP was exposed to two freeze / thaw cycles (frozen at -30°C and thawed at room temperature). The bulk FDS was physically and chemically stable after undergoing two freeze / thaw cycles. Data not shown.

[0205] mAb1 PAR formulations stored in polycarbonate vials are physically and chemically stable when stored at -20°C or -30°C for up to 12 months. Data not shown.

[0206] in conclusion: Formulation stability studies have shown that, under recommended storage conditions (2–8 °C), compositional changes in the mAb1 formulation within the study range do not adversely affect the stability or quality of 50 mg / mL mAb1 DP. Therefore, the 50 mg / mL mAb1 DP formulation is considered stable at 20 mM L-arginine hydrochloride and a pH range of 5.7 to 6.3, for protein concentrations ranging from 45 mg / mL to 55 mg / mL, sucrose concentrations ranging from 8% to 12% (w / v), L-histidine concentrations ranging from 8 mM to 12 mM, and polysorbate 80 concentrations ranging from 0.05% to 0.2% (w / v).

[0207] The scope of this invention is not limited to the specific embodiments described herein. In fact, various modifications to the invention will become apparent to those skilled in the art from the foregoing description and drawings, in addition to those described herein. Such modifications are intended to fall within the scope of the appended claims.

[0208] Table 16: Informal Sequence List .

Claims

1. A liquid pharmaceutical preparation comprising: (a) An antibody that specifically binds to human lymphocyte activation gene-3 (LAG-3), wherein the antibody comprises three heavy chain complementarity-determining regions (CDRs) (HCDR1, HCDR2 and HCDR3) contained in the heavy chain variable region (HCVR) of SEQ ID NO: 1 and three light chain CDRs (LCDR1, LCDR2 and LCDR3) contained in the light chain variable region (LCVR) of SEQ ID NO: 2; (b) Buffer solutions containing histidine, acetate, or phosphate; (c) Organic cosolvents containing polysorbate or polyethylene glycol; and (d) Stabilizers containing sugars or amino acids; The pH of the formulation is 6.

0. 0.

3.

2. The pharmaceutical formulation of claim 1, wherein the antibody concentration is 5 mg / mL. 0.75 mg / mL to 250 mg / mL 37.5 mg / mL.

3. The pharmaceutical formulation of claim 2, wherein the antibody concentration is 50 mg / mL. 7.5 mg / mL.

4. The pharmaceutical formulation of claim 2, wherein the antibody concentration is 100 mg / mL. 15 mg / mL.

5. The pharmaceutical formulation of claim 2, wherein the antibody concentration is 150 mg / mL. 22.5 mg / mL.

6. The pharmaceutical formulation of claim 2, wherein the antibody concentration is 175 mg / mL. 26.25 mg / mL.

7. The pharmaceutical formulation according to any one of claims 1-6, wherein the buffer is histidine, and the concentration of the histidine buffer is 5 mM. 1 mM to 20 mM 4 mM.

8. The pharmaceutical formulation of claim 7, wherein the histidine buffer concentration is 10 mM. 2 mM.

9. The pharmaceutical formulation according to any one of claims 1-8, wherein the cosolvent is polysorbate, and the concentration of the polysorbate is 0.01%. 0.005% to 0.5% 0.25% w / v.

10. The pharmaceutical formulation of claim 9, wherein the cosolvent is polysorbate, and the concentration of the polysorbate is 0.05%. 0.025% w / v, 0.075 0.0375% w / v or 0.1% 0.05% w / v.

11. The pharmaceutical formulation of claim 9, wherein the cosolvent is polysorbate, and the concentration of the polysorbate is 0.2%. 0.1% w / v.

12. The pharmaceutical formulation according to any one of claims 9-11, wherein the cosolvent is polysorbate, and the cosolvent is polysorbate 80.

13. The pharmaceutical formulation according to any one of claims 1-12, wherein the stabilizer comprises sucrose, and the sucrose concentration is from 1% to 20%. 4% w / v.

14. The pharmaceutical preparation of claim 13, wherein the sucrose concentration is 5%. 1% w / v.

15. The pharmaceutical preparation of claim 14, wherein the sucrose concentration is 10%. 2% w / v.

16. The pharmaceutical formulation of any one of claims 1-15, wherein the stabilizer comprises sugar and amino acids.

17. The pharmaceutical formulation of claim 16, wherein the amino acid is present at a concentration of 0 to 80 mM. 16 mM arginine hydrochloride.

18. The pharmaceutical formulation of claim 17, wherein the arginine hydrochloride concentration is 20 mM. 4 mM.

19. The pharmaceutical preparation of claim 1, comprising: (a) 175 mg / mL 26.25 mg / mL of antibody, (b) 5 mM 1mM to 20mM 4mM histidine buffer. (c) 0.1% 0.05% to 0.5% 0.25% w / v polysorbate, (d) 1% 0.2% to 15% 3% w / v sucrose, and (e) 1 mM 0.2 mM to 100 mM 20 mM arginine hydrochloride, pH is 6.0 0.

3.

20. The pharmaceutical preparation of claim 1, comprising: (a) 150 mg / mL 22.5 mg / mL antibody, (b) 5 mM 1 mM to 20 mM 4mM 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 mM 0.2 mM to 100 mM 20 mM arginine hydrochloride; pH is 6.0 0.

3.

21. The pharmaceutical preparation of claim 1, comprising: (a) 100 mg / mL 15 mg / mL antibody, (b) 5 mM 1 mM to 20 mM 4mM 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 mM 0.2 mM to 100 mM 20 mM arginine hydrochloride; pH is 6.0 0.

3.

22. The pharmaceutical formulation of claim 21, comprising: (a) 100 mg / mL 15 mg / mL antibody, (b) 10 mM 2 mM histidine buffer, (c) 0.1% 0.05% w / v polysorbate, (d) 10% 2% w / v sucrose, and (e) 20 mM 4 mM arginine hydrochloride, pH is 6.0 0.

3.

23. The pharmaceutical preparation of claim 1, comprising: (a) 50 mg / mL 7.5 mg / mL antibody, (b) 5 mM 1mM to 20mM 4mM 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) 0 mM to 80 mM 16 mM arginine hydrochloride, pH is 6.0 0.

3.

24. The pharmaceutical preparation of claim 23, comprising: (a) 50 mg / mL 7.5 mg / mL antibody, (b) 10 mM 2 mM histidine buffer, (c) 0.1% 0.05% w / v polysorbate, (d) 5% 1% w / v sucrose, and (e) 20 mM 4 mM arginine hydrochloride, pH is 6.0 0.

3.

25. The pharmaceutical preparation of claim 23, comprising: (a) 50 mg / mL 7.5 mg / mL antibody, (b) 10 mM 2 mM histidine buffer, (c) 0.1% 0.05% w / v polysorbate, (d) 10% 2% w / v sucrose, and (e) 20 mM 4 mM arginine hydrochloride, pH is 6.0 0.

3.

26. The pharmaceutical formulation of claim 23, comprising: (a) 50 mg / mL 7.5 mg / mL antibody, (b) 10 mM 2 mM histidine buffer, (c) 0.1% 0.05% w / v polysorbate, and (d) 5% 1% w / v sucrose, and pH is 6.0 0.

3.

27. The pharmaceutical formulation of any one of claims 1-26, wherein at least 90% of the antibody has a natural conformation after 28 days at 40°C.

28. The pharmaceutical formulation according to any one of claims 1-27, wherein in 40 After 28 days at C, at least 45% of the antibodies were the major charge variants of the antibodies.

29. The pharmaceutical formulation according to any one of claims 1-28, wherein at least 98% of the antibody has a natural conformation after three months at 25°C.

30. The pharmaceutical formulation according to any one of claims 1-29, wherein in 25 Three months after C, at least 50% of the antibodies were the major charge variants of the antibodies.

31. The pharmaceutical formulation according to any one of claims 1-30, wherein in 2-8 After 12 months at C, at least 98% of the antibodies retained their native conformation.

32. The pharmaceutical formulation according to any one of claims 1-31, wherein in 2-8 After 12 months at C, at least 50% of the antibodies were the major charge variants of the antibodies.

33. The pharmaceutical formulation according to any one of claims 1-32, wherein in 2-8 After 24 months at C, at least 98% of the antibodies retained their native conformation.

34. The pharmaceutical formulation according to any one of claims 1-33, wherein in 2-8 After 24 months at C, at least 50% of the antibodies were the major charge variants of the antibodies.

35. A pharmaceutical preparation comprising: (a) 175 mg / mL A 26.25 mg / mL antibody specifically binding to LAG-3, wherein the antibody comprises HCVR of SEQ ID NO: 1 and LCVR of SEQ ID NO:

2. (b) 10 mM 2 mM histidine buffer, pH 6.0 0.3, (c) 0.1% 0.05% w / v polysorbate 80, (d) 1% 0.2% to 10% 2% w / v sucrose; and (e) 1 mM 0.2 mM to 100 mM 20 mM arginine hydrochloride.

36. The pharmaceutical formulation of claim 35, wherein: (i) ≥ 90% of the antibodies have a strength of 145 kDa Molecular weight of 1 kDa; as well as (ii) After being stored at 2-8°C for 6 months, at least 98% or more of the antibodies retain their native conformation.

37. A pharmaceutical preparation comprising: (a) 150 mg / mL A 22.5 mg / mL antibody that specifically binds to LAG-3, wherein the antibody comprises HCVR of SEQ ID NO: 1 and LCVR of SEQ ID NO:

2. (b) 10 mM 2 mM histidine buffer, pH 6 0.3, (c) 0.1% 0.05% w / v polysorbate 80, (d) 1% 0.2% to 10% 2% w / v sucrose; and (e) 1 mM 0.2 mM to 100 mM 20 mM arginine hydrochloride.

38. The pharmaceutical formulation of claim 37, wherein: (i) ≥ 90% of the antibodies have a strength of 145 kDa A molecular weight of 1 kDa; and (ii) After being stored at 2-8°C for 6 months, at least 98% or more of the antibodies retain their native conformation.

39. A pharmaceutical preparation comprising: (a) 100 mg / mL A 7.5 mg / mL antibody specifically binding to LAG-3, wherein the antibody comprises HCVR of SEQ ID NO: 1 and LCVR of SEQ ID NO:

2. (b) 10 mM 2 mM histidine buffer, pH 6.0 0.3, (c) 0.1% 0.05% w / v polysorbate 80, (d) 1% 0.2% to 10% 2% w / v sucrose; and (e) 1 mM 0.2 mM to 100 mM 20 mM arginine hydrochloride.

40. The pharmaceutical preparation of claim 39, wherein: (a) ≥ 90% of the antibodies have a strength of 145 kDa A molecular weight of 1 kDa; and (b) After being stored at 2–8°C for 6 months, at least 98% of the antibodies retain their native conformation.

41. A pharmaceutical preparation comprising: (a) 50 mg / mL A 7.5 mg / mL antibody specifically binding to LAG-3, wherein the antibody comprises HCVR of SEQ ID NO: 1 and LCVR of SEQ ID NO:

2. (b) 10 mM 2 mM histidine buffer, pH 6 0.3, (c) 0.1% 0.05% w / v polysorbate 80, (d) 10% 2% w / v sucrose, and (e) 20 mM 4 mM arginine hydrochloride.

42. The pharmaceutical preparation of claim 41, wherein: (i) ≥ 90% of the antibodies have a strength of 145 kDa A molecular weight of 1 kDa; and (ii) After being stored at 2-8°C for 12 months, more than 98% of the antibodies retain their native conformation.

43. The pharmaceutical formulation of claim 41, comprising: (a) 50 mg / mL (a) 7.5 mg / mL of the antibody, (b) 10.0 mM ± 2.0 mM histidine buffer, (c) 0.1% 0.05% w / v polysorbate 80, (d) 10% 2% w / v sucrose; and (e) 20 mM 4 mM arginine hydrochloride; the above substances have a pH of 6.0 in water. 0.

3.

44. A pharmaceutical preparation comprising: (a) 5-250 mg / ml of antibodies that specifically bind to LAG-3, wherein the antibodies comprise HCVR of SEQ ID NO: 1 and LCVR of SEQ ID NO:

2. (b) 10 mM 2 mM histidine buffer, pH 6.0 0.3, (c) 0.1% 0.05% w / v polysorbate 80, and (d) 5% 1% w / v sucrose.

45. The pharmaceutical formulation further comprises 1 mM 0.2 mM to 100 mM 20 mM arginine hydrochloride.

46. ​​The pharmaceutical formulation according to any one of claims 1-45, wherein the antibody comprises HCDR1 of SEQ ID NO: 3, HCDR2 of SEQ ID NO: 4, HCDR3 of SEQ ID NO: 5, LCDR1 of SEQ ID NO: 6, LCDR2 of SEQ ID NO: 7, and LCDR3 of SEQ ID NO:

8.

47. The pharmaceutical formulation of claim 46, wherein the antibody comprises HCVR of SEQ ID NO: 1 and LCVR of SEQ ID NO:

2.

48. The pharmaceutical formulation of any one of claims 1-46, wherein the antibody comprises HCVR having 90% sequence identity with SEQ ID NO:

1.

49. The pharmaceutical formulation of any one of claims 1-46, wherein the antibody comprises LCVR having 90% sequence identity with SEQ ID NO:

2.

50. The pharmaceutical formulation of any one of claims 1-46, wherein the antibody comprises HCVR having 90% sequence identity with SEQ ID NO: 1 and LCVR having 90% sequence identity with SEQ ID NO:

2.

51. The pharmaceutical formulation of any one of claims 1-46, 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.

52. The pharmaceutical formulation of any one of claims 1-46, wherein the antibody comprises a heavy chain and a light chain, wherein the heavy chain comprises an amino acid sequence containing amino acids 1-448 of SEQ ID NO:

9.

53. The pharmaceutical formulation of any one of claims 1-46, 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.

54. The pharmaceutical formulation of any one of claims 1-46, wherein the antibody comprises a heavy chain / light chain containing the amino acid sequence of SEQ ID NO: 9 / 10.

55. The pharmaceutical formulation of any one of claims 1-46, wherein the antibody comprises a heavy chain amino acid sequence containing amino acids 1-448 of SEQ ID NO: 9 and a light chain containing amino acid sequence SEQ ID NO:

10.

56. The pharmaceutical preparation of any one of claims 1-55, wherein the preparation is contained in a container.

57. The pharmaceutical preparation of claim 56, wherein the container is a vial.

58. The pharmaceutical preparation of claim 57, wherein the vial is a 2 ml, 5 ml, 10 mL or 20 mL Type I transparent glass vial.

59. The pharmaceutical preparation of claim 57, wherein the container is a syringe.

60. The pharmaceutical formulation of claim 57, wherein the syringe is made of low-tungsten glass.

61. The pharmaceutical formulation of claim 57, wherein the container is a pre-filled syringe.

62. The pharmaceutical preparation of claim 57, wherein it is contained in an autoinjector.

63. A kit comprising a pharmaceutical preparation, a container, and instructions for use as described in any one of claims 1-55.

64. The kit of claim 63, wherein the container is a glass vial.

65. The kit of claim 63, wherein the container is a pre-filled syringe.

66. The kit of claim 63, wherein the container is an autoinjector.

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