Formulations of human Anti-PD-l1 antibodies

Abstract

To provide anti-PD-L1 antibody buffer formulations that retain liquid drug substance and lyophilized drug product stability.SOLUTION: An antibody formulation comprises 0 mg / mL to 50 mg / mL of anti-PD-L1 antibody, 15 mM to 35 mM of buffer, 255 mM to 275 mM of disaccharide, and 0.01%(w / v) to 0.05%(w / v) of surfactant, and a pH of the formulation is about pH 5.5 to about pH 7.2.SELECTED DRAWING: Figure 2

Description

Technical Field

[0001] The present disclosure relates to formulations and compositions of antibodies targeting human anti-PD-L1 such as durvalumab.

Background Art

[0002] Programmed death ligand 1 (PD-L1), also known as B7H1, is a 40 kDa transmembrane protein that poses a major obstacle to anti-cancer immunity. PD-L1 that binds to the programmed death receptor (PD-1) inactivates T cells, protects tumor cells, suppresses detection by the immune system, and allows unchecked growth of cancer cells. PD-L1 also binds to the costimulatory molecule CD80.

[0003] A wide range of tumorigenic and activated immune cell types, including antigen-presenting cells, macrophages, monocytes, B cells, T cells, and non-hematopoietic cells, naturally express PD-L1. In addition, inflammatory cytokines such as interferon gamma (IFNγ) may induce the expression of PD-L1. For example, activated T cells produce IFNγ, which is the most potent inducer of PD-L1. The expression of PD-L1 induced by IFNγ promotes tumor protection, which is a mechanism known as adaptive immune tolerance.

[0004] One strategy for suppressing adaptive immune tolerance and the lethality of PD-L1 is to use anti-PD-L1 antibodies. Consistent with this approach, the 149 kDa anti-PD-L1 antibody durvalumab, an affinity-optimized anti-PD-L1 monoclonal IgG1 triple mutant (TM) that interferes with the binding of PD-L1 to PD-1, can be employed to abrogate the immunosuppressive effect of PD-L1 on cytotoxic T cells. The result is a reduction in negative inhibitory signals that promote tumor growth, an enhancement of anti-tumor immunity, and a response that enhances the killing of tumor cells by the immune system.

[0005] Importantly, buffer formulations of anti-PD-L1 antibodies that maintain the stability of liquid drug substances and lyophilized drug products are essential for the effectiveness of anti-PD-L1 antibodies.

SUMMARY OF THE INVENTION

MEANS FOR SOLVING THE PROBLEM

[0006] In one aspect, the present disclosure provides an antibody formulation comprising an anti-PD-L1 antibody at 40 mg / mL to 50 mg / mL, a buffer at 15 mM to 35 mM, a disaccharide at 255 mM to 275 mM, and a surfactant at 0.01% (w / v) to 0.05% (w / v), wherein the pH of the formulation is from about pH 5.5 to about pH 7.2.

[0007] In one aspect, the present disclosure provides an antibody formulation comprising a human anti-PD-L1 antibody at 50 mg / mL, a histidine / histidine-HCl buffer at 26 mM, trehalose anhydrous at 275 mM, and polysorbate 80 at 0.02% (w / v), wherein the pH of the formulation is pH 6.0.

[0008] In one aspect, the present disclosure provides an antibody formulation comprising a human anti-PD-L1 antibody at 50 mg / mL, a histidine / histidine-HCl buffer at 25 mM, trehalose anhydrous at 265 mM, and polysorbate 80 at 0.02% (w / v), wherein the pH of the formulation is about pH 5.5.

[0009] In one aspect, the present disclosure provides an antibody formulation comprising a human anti-PD-L1 antibody at 50 mg / mL, a histidine / histidine-HCl buffer at 25 mM, trehalose anhydrous at 265 mM, and polysorbate 80 at 0.02% (w / v), wherein the pH of the formulation is about pH 6.5.

[0010] In one aspect, the present disclosure provides an antibody formulation comprising a human anti-PD-L1 antibody at 50 mg / mL, a histidine / histidine-HCl buffer at 25 mM, trehalose anhydrous at 265 mM, and polysorbate 80 at 0.04% (w / v), wherein the pH of the formulation is about pH 6.0.

[0011] In one aspect, the present disclosure provides an antibody formulation comprising a human anti-PD-L1 antibody at 50 mg / mL, 25 mM histidine / histidine-HCl buffer, 265 mM sucrose, and 0.02% (w / v) polysorbate 80, wherein the pH of the formulation is about pH 6.0.

[0012] In one aspect, the present disclosure provides an anti-PD-L1 antibody comprising a light chain having an amino acid sequence with at least 90% identity to the amino acid sequence of SEQ ID NO: 1 and a heavy chain having an amino acid sequence with at least 90% identity to the amino acid sequence of SEQ ID NO: 2, and a composition comprising a major form of the antibody containing 45% or more of the protein in the composition as measured using capillary isoelectric focusing (cIEF) of the composition.

[0013] In one aspect, the present disclosure provides an anti-PD-L1 antibody comprising a light chain having an amino acid sequence with at least 90% identity to the amino acid sequence of SEQ ID NO: 1 and a heavy chain having an amino acid sequence with at least 90% identity to the amino acid sequence of SEQ ID NO: 2, a major form of the antibody containing 45% or more of the protein in the composition as measured using cIEF of the composition, and an acidic form of the antibody containing 45% - 50% of the protein in the composition as measured using cIEF of the composition.

[0014] In one aspect, the present disclosure provides an anti-PD-L1 antibody comprising a light chain having an amino acid sequence with at least 90% identity to the amino acid sequence of SEQ ID NO: 1 and a heavy chain having an amino acid sequence with at least 90% identity to the amino acid sequence of SEQ ID NO: 2, a major form of the antibody containing 45% or more of the protein in the composition as measured using cIEF of the composition, and a basic form of the antibody containing 18% - 23% of the protein in the composition as measured using cIEF of the composition.

[0015] In one aspect, the present disclosure provides a composition comprising a light chain having an amino acid sequence with at least 90% identity to the amino acid sequence of SEQ ID NO: 1, and a heavy chain having an amino acid sequence with at least 90% identity to the amino acid sequence of SEQ ID NO: 2, a major form of the antibody comprising at least 45% of the protein in the composition as measured using cIEF of the composition, an acidic form of the antibody comprising 45% - 50% of the protein in the composition as measured using cIEF of the composition, and a basic form of the antibody comprising 18% - 23% of the protein in the composition as measured using cIEF of the composition.

[0016] In one aspect, the present disclosure provides a composition comprising an anti-PD-L1 antibody comprising a light chain having an amino acid sequence with at least 90% identity to the amino acid sequence of SEQ ID NO: 1, and a heavy chain having an amino acid sequence with at least 90% identity to the amino acid sequence of SEQ ID NO: 2, wherein the glycan structure of the anti-PD-L1 antibody comprises G0f, G1f, G2f, and G0 glycoforms.

[0017] In one aspect, the present disclosure provides a composition comprising an anti-PD-L1 antibody comprising a light chain having an amino acid sequence with at least 90% identity to the amino acid sequence of SEQ ID NO: 1, and a heavy chain having an amino acid sequence with at least 90% identity to the amino acid sequence of SEQ ID NO: 2, wherein 1.5% - 2.5% of the anti-PD-L1 antibody forms aggregates as determined by high performance size exclusion chromatography (HP-SPEC), and 97% - 98% of the anti-PD-L1 antibody exists as monomers as measured by HP-SEC. BRIEF DESCRIPTION OF THE DRAWINGS

[0018]

Figure 1

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Figure 5C

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Figure 12 - 1

Figure 12 - 2

Figure 12 - 3

Figure 12 - 4

Mode for Carrying Out the Invention

[0019] The present disclosure relates to formulations and compositions of antibodies targeting anti-PD-L1 such as durvalumab.

[0020] As used in accordance with the present disclosure, the following terms are understood to have the following meanings unless otherwise indicated. Unless the context requires otherwise, singular terms shall include the plural, and plural terms shall include the singular.

[0021] As used herein, the term "antibody" refers to a protein that recognizes, and in particular has the ability to bind to, an antigen. A normal or conventional mammalian antibody comprises a tetramer, which typically consists of two identical pairs of polypeptide chains, each pair consisting of one "light" chain (typically having a molecular weight of about 25 kDa) and one "heavy" chain (typically having a molecular weight of about 50-70 kDa). As used herein, the terms "heavy chain" and "light chain" refer to any immunoglobulin polypeptide having a variable domain sequence sufficient to confer specificity for a target antigen. The amino-terminal portion of each light and heavy chain typically contains a variable domain of about 100-110 or more amino acids that typically serves the role of antigen recognition. The carboxyl-terminal portion of each chain typically defines a constant domain that typically serves the role of effector function. Thus, in a naturally-occurring antibody, a full-length heavy-chain immunoglobulin polypeptide comprises a variable domain (VH) and three constant domains (CH1, CH2, and CH3), as well as a hinge region between CH1 and CH2, where the VH domain is at the amino terminus of the polypeptide and the CH3 domain is at the carboxyl terminus, and a full-length light-chain immunoglobulin polypeptide comprises a variable domain (VL) and a constant domain (CL), where the VL domain is at the amino terminus of the polypeptide and the CL domain is at the carboxyl terminus.

[0022] Within the full-length light and heavy chains, the variable and constant domains are typically linked by a "J" region of about 12 or more amino acids, and the heavy chain further includes a "D" region of more than about 10 amino acids. The variable regions of each light / heavy chain pair typically form the antigen-binding site. The variable domains of naturally occurring antibodies typically exhibit the same general structure of relatively conserved framework regions (FRs) that are linked by three hypervariable regions, also typically referred to as complementarity-determining regions or CDRs. The CDRs of the two chains of each pair typically align by the framework regions and may enable binding to a specific epitope. From the amino terminus to the carboxyl terminus, the variable domains of both the light and heavy chains typically include domains FR1, CDR1, FR2, CDR2, FR3, CDR3, and FR4.

[0023] The antibody formulations disclosed herein include an anti-PD-L1 antibody. In certain embodiments, the formulation includes an anti-PD-L1 antibody at 30 mg / mL, 35 mg / mL, 40 mg / mL, 45 mg / mL, 50 mg / mL, 55 mg / mL, or 60 mg / mL. In other embodiments, the formulation includes an anti-PD-L1 antibody at 40 mg / mL to 50 mg / mL. In other embodiments, the formulation includes an anti-PD-L1 antibody at 50 mg / mL. In certain embodiments, the anti-PD-L1 antibody is of human origin.

[0024] The antibody formulations disclosed herein include one or more buffers. As used herein, "buffer" refers to an excipient for maintaining the pH of the formulation. In certain embodiments, the buffer is a histidine / histidine-HCl buffer. The buffer is present at a concentration of about 15 mM, 20 mM, 25 mM, or 30 mM. In certain embodiments, the concentration of the buffer is 26 mM.

[0025] In certain embodiments, the antibody formulation comprises a disaccharide. In certain embodiments, the disaccharide is trehalose dihydrate or sucrose. The disaccharide is present at a concentration of about 250 mM, 255 mM, 260 mM, 265 mM, 270 mM, 275 mM, or 280 mM. In certain embodiments, the concentration of the disaccharide is 265 mM. In other embodiments, the concentration of the disaccharide is 275 mM.

[0026] In certain embodiments, the antibody formulation comprises a surfactant. As used herein, the term "surfactant" refers to an organic substance having an amphiphilic structure; that is, such a substance is composed of groups with opposing solubility tendencies, typically an oil-soluble hydrocarbon chain and a water-soluble ionic group. Surfactants can be classified into anionic, cationic, and nonionic surfactants according to the charge of the surface-active moiety. Surfactants are frequently used as wetting, emulsifying, solubilizing, and dispersing agents for various pharmaceutical formulations and preparations of biological substances. In certain embodiments, the surfactant is polysorbate 80. The surfactant is present at a concentration of about 0.001% to about 0.5% (v / v).

[0027] In certain embodiments, an antibody formulation is disclosed herein that comprises an anti-PD-L1 antibody at 40 mg / mL to 50 mg / mL, a buffer at 15 mM to 35 mM, a disaccharide at 255 mM to 275 mM, and a surfactant at 0.01% (w / v) to 0.05% (w / v), and the pH of the formulation is from about pH 5.5 to about pH 7.2. In certain embodiments, the antibody formulation comprises 50 mg / mL of a human anti-PD-L1 antibody, 26 mM of a histidine / histidine-HCl buffer, 275 mM of trehalose anhydrous, and 0.02% (w / v) of polysorbate 80, wherein the pH of the formulation is about 6.0. In other embodiments, the antibody formulation comprises 50 mg / mL of a human anti-PD-L1 antibody, 25 mM of a histidine / histidine-HCl buffer, 265 mM of trehalose anhydrous, and 0.02% (w / v) of polysorbate 80, wherein the pH of the formulation is about 5.5. In other embodiments, the antibody formulation comprises 50 mg / mL of a human anti-PD-L1 antibody, 25 mM of a histidine / histidine-HCl buffer, 265 mM of trehalose anhydrous, and 0.02% (w / v) of polysorbate 80, wherein the pH of the formulation is about 6.5. In other embodiments, the antibody formulation comprises 50 mg / mL of a human anti-PD-L1 antibody, 25 mM of a histidine / histidine-HCl buffer, 265 mM of trehalose anhydrous, and 0.04% (w / v) of polysorbate 80, wherein the pH of the formulation is about 6.0. In other embodiments, the antibody formulation comprises 50 mg / mL of a human anti-PD-L1 antibody, 25 mM of a histidine / histidine-HCl buffer, 265 mM of sucrose, and 0.02% (w / v) of polysorbate 80, wherein the pH of the formulation is about 6.0. In other embodiments, the antibody formulation comprises 50 mg / mL of a human anti-PD-L1 antibody, 25 mM of a histidine / histidine-HCl buffer, 265 mM of sucrose, and 0.02% (w / v) of polysorbate 80, wherein the pH of the formulation is about 6.0.

[0028] In certain embodiments, the human anti-PD-L1 antibody comprises a light chain variable domain comprising the amino acid sequence of SEQ ID NO: 1 and a heavy chain variable domain comprising the amino acid sequence of SEQ ID NO: 2. In other embodiments, the human anti-PD-L1 antibody comprises a VH CDR1 having the amino acid sequence of SEQ ID NO: 3, a VH CDR2 having the amino acid sequence of SEQ ID NO: 4, a VH CDR3 having the amino acid sequence of SEQ ID NO: 5, a VL CDR1 having the amino acid sequence of SEQ ID NO: 6, a VL CDR2 having the amino acid sequence of SEQ ID NO: 7, and a VL CDR3 having the amino acid sequence of SEQ ID NO: 8.

[0029] In certain embodiments, the human anti-PD-L1 antibody comprises a light chain variable domain comprising an amino acid sequence having less than 100% identity to the amino acid sequence of SEQ ID NO: 1 and a heavy chain variable domain comprising an amino acid sequence having less than 100% identity to the amino acid sequence of SEQ ID NO: 2. In other embodiments, the human anti-PD-L1 antibody comprises a light chain variable domain comprising an amino acid sequence having 90% sequence identity, 91% sequence identity, 92% sequence identity, 93% sequence identity, 94% sequence identity, 95% sequence identity, 96% sequence identity, 97% sequence identity, 98% sequence identity, or 99% sequence identity to the amino acid sequence of SEQ ID NO: 1, and a heavy chain variable domain comprising an amino acid sequence having 90% sequence identity, 91% sequence identity, 92% sequence identity, 93% sequence identity, 94% sequence identity, 95% sequence identity, 96% sequence identity, 97% sequence identity, 98% sequence identity, or 99% sequence identity to the amino acid sequence of SEQ ID NO: 2.

[0030] When used herein, the terms "MEDI4736" and "durvalumab" refer to antibodies that selectively bind to human anti-PD-L1 and interfere with the binding of PD-L1 to the PD-1 and CD80 receptors, as disclosed in U.S. Patent Nos. 8,779,108 and 9,493,565, each of which is incorporated herein by reference in its entirety. The fragment crystallizable (Fc) domain of durvalumab contains a triple mutation in the constant region of the IgG1 heavy chain that reduces binding to complement component C1q and Fcγ receptors involved in mediating antibody-dependent cell-mediated cytotoxicity (ADCC). Durvalumab can reduce PD-L1-mediated suppression of human T cell activation in vitro and inhibit tumor growth in xenograft models via T cell-dependent mechanisms.

[0031] As used herein, the phrases "pharmaceutical formulation", "formulation", and "antibody formulation" are used interchangeably and refer to a composition comprising an anti-PD-L1 antibody and one or more suitable buffers and / or excipients. Appropriately, the pharmaceutical formulations described herein are "pharmaceutically acceptable" and thus will meet the required approval requirements set forth by federal or state government regulatory authorities or as set forth in the United States Pharmacopeia, the European Pharmacopeia, or other generally recognized pharmacopeias for use in animals and particularly in humans.

[0032] The antibody formulations disclosed herein can be formulated as liquid formulations, frozen formulations, lyophilized formulations, or reconstituted formulations.

[0033] Lyophilization can be effected by drying in an oven, vacuum centrifugation, or other means known to those of skill in the art. Lyophilized durvalumab retains the activity of the anti-PD-L1 antibody when reconstituted.

[0034] The terms "stability" and "stable", as used herein in the context of a formulation comprising an anti-PD-L1 antibody, refer to the resistance of the antibody in the formulation to aggregation, degradation, or fragmentation under given manufacturing, preparation, transportation, and storage conditions. A "stable" formulation retains its biological activity under given manufacturing, preparation, transportation, and storage conditions. The stability of an antibody can be evaluated by the degree of aggregation, degradation, or fragmentation as measured by, for example, high pressure size exclusion chromatography (HP-SEC), static light scattering (SLS), Fourier transform infrared spectroscopy (FTIR), circular dichroism (CD), urea unfolding techniques, intrinsic tryptophan fluorescence, differential scanning calorimetry, and / or ANS binding techniques, compared to a reference formulation. The overall stability of a formulation comprising a human anti-PD-L1 antibody can be evaluated by various immunological assays including, for example, ELISA and radioimmunoassay using isolated antigen molecules.

[0035] In certain embodiments, less than about 1% of the anti-PD-L1 antibody forms aggregates upon storage at about 40 °C for about 1 month, as determined by HP-SEC. In other embodiments, at least about 97% of the human anti-PD-L1 antibody is present as a monomer after storage at about 40 °C for about 1 month, as measured by HP-SPEC. In other embodiments, at least about 99% of the human anti-PD-L1 antibody is present as a monomer after storage at about 40 °C for about 1 month, as measured by HP-SPEC. In other embodiments, at least about 98% of the human anti-PD-L1 antibody is present as a monomer after storage at about 5 °C for about 1 month, as measured by HP-SPEC. In certain embodiments, the antibody formulation maintains its stability after at least three freeze / thaw cycles.

[0036] In certain embodiments, the compositions disclosed herein comprise an anti-PD-L1 antibody comprising a light chain having an amino acid sequence with at least 90% identity to the amino acid sequence of SEQ ID NO: 1 and a heavy chain having an amino acid sequence with at least 90% identity to the amino acid sequence of SEQ ID NO: 2; wherein the major form of the antibody comprises at least 45% of the protein in the composition as measured using capillary isoelectric focusing (cIEF) of the composition. In other embodiments, the compositions disclosed herein comprise an anti-PD-L1 antibody comprising a light chain having an amino acid sequence with at least 90% identity to the amino acid sequence of SEQ ID NO: 1 and a heavy chain having an amino acid sequence with at least 90% identity to the amino acid sequence of SEQ ID NO: 2; wherein the major form of the antibody comprises at least 45% of the protein in the composition as measured using cIEF of the composition; wherein the acidic form of the antibody comprises 45% - 50% of the protein in the composition as measured using cIEF of the composition. In other embodiments, the compositions disclosed herein comprise an anti-PD-L1 antibody comprising a light chain having an amino acid sequence with at least 90% identity to the amino acid sequence of SEQ ID NO: 1 and a heavy chain having an amino acid sequence with at least 90% identity to the amino acid sequence of SEQ ID NO: 2; wherein the major form of the antibody comprises at least 45% of the protein in the composition as measured using cIEF of the composition; wherein the basic form of the antibody comprises 18% - 23% of the protein in the composition as measured using cIEF of the composition. In other embodiments, the compositions disclosed herein comprise an anti-PD-L1 antibody comprising a light chain having an amino acid sequence with at least 90% identity to the amino acid sequence of SEQ ID NO: 1 and a heavy chain having an amino acid sequence with at least 90% identity to the amino acid sequence of SEQ ID NO: 2; wherein the major form of the antibody comprises at least 45% of the protein in the composition as measured using cIEF of the composition; wherein the acidic form of the antibody comprises 45% - 50% of the protein in the composition as measured using cIEF of the composition; wherein the basic form of the antibody comprises 18% - 23% of the protein in the composition as measured using cIEF of the composition.

[0037] In certain embodiments, the compositions disclosed herein include an anti-PD-L1 antibody comprising a light chain having an amino acid sequence with at least 90% identity to the amino acid sequence of SEQ ID NO: 1, and a heavy chain having an amino acid sequence with at least 90% identity to the amino acid sequence of SEQ ID NO: 2; wherein the glycan structure of the anti-PD-L1 antibody comprises G0f, G1f, G2f, and the G0 glycoform. In other embodiments, the glycan structure of the anti-PD-L1 antibody has a content higher than about 90% with respect to G0f, G1f, G2f, and the G0 form. In some embodiments, the composition of the anti-PD-L1 antibody comprises a G0f content of about 65 - 75%, a G1f content of 13 - 23%, a G2f content of 0 - 3%, and a G0 content of 0 - 4%. In other embodiments, the composition of the anti-PD-L1 antibody comprises a G0f content of about 71.9%, a G1f content of 18.4%, a G2f content of 1.5%, and a G0 content of 1.9%.

[0038] In certain embodiments, the compositions disclosed herein include an anti-PD-L1 antibody comprising a light chain having an amino acid sequence with at least 90% identity to the amino acid sequence of SEQ ID NO: 1, and a heavy chain having an amino acid sequence with at least 90% identity to the amino acid sequence of SEQ ID NO: 2; wherein, as determined by high pressure size exclusion chromatography (HP-SPEC), 1.5% - 2.5% of the anti-PD-L1 antibody forms aggregates, and wherein, as measured by HP-SEC, 97% - 98% of the anti-PD-L1 antibody exists as monomers.

[0039] Without limiting the present disclosure, numerous embodiments of the present disclosure are described below for illustrative purposes. The following examples illustrate specific embodiments of the invention and their various uses. They are described for illustrative purposes only and are not considered to limit the invention.

Examples

[0040] Example 1 Materials and Methods 1. Determination of Protein Concentration The Dulvalbumin protein concentration was determined by measuring the absorbance at 280 nm with an Aligent UV-V spectrophotometer. Dilutions were made in the formulation buffer. The protein concentration was calculated by using a theoretical extinction coefficient of 1.55 (mg / mL)-1cm-1. In part of the determination, an experimental coefficient of 1.52 (mg / mL)-1cm-1 was used in the calculation.

[0041] 2. High Performance Size Exclusion Chromatography (HP-SEC) Analysis Samples for HP-SEC analysis were first eluted at a flow rate of 1.0 mL / min at a uniform concentration using 0.1 M disodium phosphate pH 6.8 containing 0.1 M sodium sulfate. The eluted protein was detected at an absorbance of 280 nm. The results were reported as the area percentage of the product monomer peak compared to all other peaks. Buffer-related peaks observed at approximately 12 minutes were excluded from the reported results. Any peak eluting earlier than the monomer peak was recorded as the aggregation percentage. Peaks eluting after the monomer peak were recorded as the fragment percentage.

[0042] 3. Visual Analysis Visual inspection of the samples was performed by testing the glass vials for color, clarity, and the presence of particulates and fibrous matter using a light box equipped with both a bright background and a dark background. Visible particles and clarity were evaluated.

[0043] 4. Visible Particle Analysis by High-Intensity (HIAC) Liquid Particle Counter Samples were diluted to 5 mg / mL using filtered formulation buffer and then allowed to stand for 30 minutes before analysis. Samples were prepared in Falcon tubes rinsed three times with ultrapure water. Samples with a protein concentration of < 5 mg / mL were analyzed as is. Six readings were taken for each sample using a Pacific Standard HIAC Royco 3000A and an 8-channel particle counter 8000A. The average of the final three readings was recorded as the final sub-visible particle count. The cumulative particle count > 10 - 25 μm was recorded.

[0044] 5. Osmotic Pressure and pH The osmotic pressure was measured using a Gonotec Osmomat 030-D osmometer. The pH of the solution was measured using a PHM220 Lab pH meter.

[0045] 6. Karl Fischer Analysis The presence of residual moisture in the lyophilized formulation was measured using Karl Fischer titration (Mettler Toledo). The lyophilized material was reconstituted using dry methanol. The residual moisture was determined based on the amount of water in dry methanol and the weight of the total solids.

[0046] 7. Capillary Isoelectric Focusing (cIEF) The sample was adjusted to 0.25 mg / mL using HPLC grade water. The sample was digested with carboxypeptidase B (CBP) for 10 minutes at 37 °C and then diluted with 1% methylcellulose solution, Pharmalyte pH 3 - 10, pI marker 9.46, and pI marker 5.85. The sample was loaded onto an iCE280 Analyzer and focused at 1500 V for 1 minute and then at 3000 V for 7 minutes. The resulting electropherogram was analyzed using EZChrom software and compared to reference standards.

[0047] 8. Reducing and Non-Reducing Gel Electrophoresis The Agilent 2100 BioAnalyzer using Protein 230 LabChip technology (Agilent) was used to analyze durvalumab by reducing and non-reducing gel electrophoresis. The flow path of the LabChip enabled separation, staining, destaining, and detection. Samples and standards were adjusted to 4 mg / mL in PBS and mixed 1:1 with SDS that denatured the sample buffer in the presence of 60 mM N-ethylmaleimide (non-reducing) or 60 mM dithiothreitol (reducing). Then, the samples were heated, centrifuged, diluted with water, and loaded into the wells on the LabChip. In the first dimension, the proteins were separated with a resolution comparable to a 4-20% gradient gel. The proteins were separated in the second dimension by molecular weight. The fluorescent dye present in the sample buffer was excited at 633 nm.

[0048] 9. Differential Scanning Calorimetry Differential scanning calorimetry (DSC) was used to determine the melting temperature (Tm1).

[0049] 10. Viscosity The viscosities of the samples and buffers were measured using an Anton Paar AMVn Viscometer. The measurements were performed at the target concentrations.

[0050] 11. Amino Acid Sequence Analysis for Hot Spots The amino acid sequence of durvalumab was analyzed using BLAZE software to identify amino acid residues for hot spots or potential modification sites. The reactivity of the hot spots was assigned a high, medium, or low risk score that met the sequence liability criteria based on experience with other monoclonal antibodies.

[0051] 12. Identification of N-Linked Oligosaccharides in Durvalumab The N-linked oligosaccharides in durvalumab, detected as significant peaks in ultra-high performance liquid chromatography (UPLC), were identified using a Waters UPLC system with an FLR detector. A durvalumab reference standard (10.2 mg / mL; 100 μg), formulated in 26 mM histidine / histidine-HCl, 275 mM trehalose dihydrate, 0.02% (w / v) polysorbate 80, pH 6.0, was reconstituted to 0.5 mg / mL in 50 mM Tris buffer (pH 7.8). Samples were digested with 2 μL of PNGase F (Promega), labeled with the fluorescent tag 2-aminobenzamide (2-AB; Sigma Aldrich), washed with a HILIC SPE cartridge, and eluted into water for UPLC profiling. The 2-AB-labeled oligosaccharides were further digested with various exo-glycosidases including fucosidase, sialidase A, mannosidase, β-galactosidase, and β-N-acetylhexosaminidase for peak identification.

[0052] Samples were injected onto an Acquity UPLC® BEH Glycan column (1.7 μM, 2.1 × 150 mm) on a Waters UPLC system using 50 mM ammonium formate, pH 4.4 as mobile phase A and 100% acetonitrile as mobile phase B. Data were collected and the profiles of the glycosidase-digested samples were carefully compared with those of the undigested samples to identify the linkages of the glycans and their oligosaccharides.

[0053] Example 2 Developmental studies The durvalumab formulation was determined after developmental studies including hot spot analysis, melting temperature, isoelectric point (pI), and stability determination (Figure 1).

[0054] 1. Hot spot analysis The stability of durvalumab during storage is essential for the efficacy of the monoclonal antibody. The risk of antibody storage is the loss of activity through the primary degradation pathway via aggregation. In addition, modification of amino acid residues during long-term storage may affect the stability and activity of the antibody.

[0055] The loss of activity of durvalumab was evaluated by homogeneous time-resolved fluorescence assay (HTRF). No loss of activity was observed after incubation at 40 °C for 1 month in both high and low pH buffers. Only a minor level of Asp isomerization at D54(G) was detected by peptide mapping after 1 month in pH 6.0 buffer at 40 °C.

[0056] 2. Determination of melting temperature, isoelectric point (pI), and stability The melting point of the antibody was determined using differential scanning calorimetry (DSC) of 3 mg / mL durvalumab in formulation buffer. The pI and stability of durvalumab were determined using capillary isoelectric focusing and high-pressure size exclusion chromatography (HPSEC), respectively.

[0057] The DSC profile of durvalumab was shown as Tm of 64.5 °C 1 and Tm of 73.04 °C 2 (Figure 2). Four peaks in the range of 8.3 - 8.8 were detected in the capillary isoelectric focusing (cIEF) profile of durvalumab. The major peak had an isoelectric point of 8.6 (Figure 3). The study of liquid stability showed that Clone 1 and Clone 2 were stable in the default formulation after incubation at either 5 °C or 40 °C for 1 month (Figure 4). The results of the development study are summarized in Table 1.

[0058]

Table 1

[0059] Example 3 Lyophilized formulation Five formulations were screened to evaluate the stability of the liquid drug substance and the lyophilized drug product. The screened formulations are shown in Table 2.

[0060] An initial level of P80 (i.e., 0.02% w / v) was used. During subsequent formulation screening for the lyophilized formulation, if particles were observed upon reconstitution of the formulation containing 0.02% (w / v) P80, 0.04% (w / v) P80 (Formulation 4) was also included. Based on the data from this study, 0.02% (w / v) P80 was found to be the optimal surfactant level for durvalumab.

[0061]

Table 2

[0062] The formulations were lyophilized for the formulation screening study. The formulations were subjected to freezing, annealing, primary drying, and secondary drying in a Virtis Genesis 25EL Freeze-Dryer. The parameters of the lyophilization cycle are provided in Table 3.

[0063]

Table 3

[0064] 1. Stability After 1 month at 40°C, the monomer loss rate for all lyophilized formulations was <0.3% per month (Table 4A). At 5°C, the percentage of monomer loss per month for all formulations was <0.1% per month. All formulations were equivalent from the perspective of aggregation after 4 weeks at 40°C (Table 4B). Aggregation was not present in any of the formulations after 4 weeks at 40°C (Table 4C).

[0065] Freeze-thaw studies were performed on all five formulations. The formulations were subjected to 0, 1, and 3 freeze-thaw cycles consisting of freezing at -70°C and thawing at ambient temperature. Samples were visually inspected and analyzed by HP-SEC at the end of the cycles. The HP-SEC results showed no significant change in purity by HP-SEC in any of the samples. All samples contained substantially no visible particles after 3 cycles.

[0066]

Table 4

[0067]

Table 5

[0068]

Table 6

[0069] 2. Stability after Reconstitution All five lyophilized formulations were stored at 5 °C and 25 °C for up to 24 hours and then reconstituted and tested for stability. None of the five formulations showed a change in the number of visible particles or a significant decrease in monomer purity when evaluated by SEC-HPLC. The results at T0 are shown in Table 5A, the results at 5 °C for 24 hours are shown in Table 5B, and the results at 25 °C for 24 hours are shown in Table 5C.

[0070]

Table 7

[0071]

Table 8

[0072]

Table 9

[0073] 3. Freeze-Thaw Studies of Durvalumab in a formulation of 3.26 mM Histidine / Histidine-HCl, 275 mM Trehalose Anhydrous, 0.02% (w / v) Polysorbate 80, pH 6.0 The stability of durvalumab was investigated by evaluating the degradation of the antibody structure with repeated freeze / thaw cycles. Consistent with this, the stability of durvalumab was tested using freeze-thaw studies.

[0074] Freeze-thaw studies of the unformulated drug substance (UDS) of durvalumab were conducted against the formulation (26 mM histidine / histidine-HCl, 275 mM trehalose anhydrous, 0.02% (w / v) polysorbate 80, pH 6.0). Durvalumab was subjected to 0, 1, and 3 uncontrolled freeze-thaw cycles in 8 mL Nalgene bottles filled with 5.6 mL or in 30 mL Celsius Pak bags filled with 20 mL. The Nalgene bottles were frozen at -80 °C and the Celsius Pak bags were frozen at -40 °C, and both were thawed at ambient temperature. The samples were then analyzed for purity by high-pressure size exclusion chromatography (HP-SEC) analysis, protein concentration determined by A280, and appearance evaluated after each freeze-thaw cycle.

[0075] Durvalumab was found to be stable after 3 freeze-thaw cycles as shown in Table 6A (Nalgene bottles) and 6B (Celsius Pak) below.

[0076]

Table 10

[0077]

Table 11

[0078] These results indicate that durvalumab was stable after repeated freeze / thaw cycles in both Nalgene bottles and Celsius Pak.

[0079] Example 4: Formation and Detection of Pyroglutamic Acid Dulvalumab has N-terminal glutamic acid on the heavy and light chains. Over time, glutamic acid cyclizes to pyroglutamic acid. This conversion was detected in a cIEF assay as a peak at pI 8.8 after storage at 25 °C and 40 °C for 3 months. The peak also occurred after 24 months at 2 - 8 °C (Figure 4). Cyclization occurred in both lyophilized and liquid forms. A mutant dulvalumab was created that showed that cyclization did not affect the antibody's potency.

[0080] Example 5: Parameters and Stability of the Final Lyophilized Formulation The lyophilized formulation of 26 mM histidine / histidine-HCl, 275 mM trehalose anhydrous, 0.02% (w / v) polysorbate 80, pH 6.0 contained 50 mg / mL of dulvalumab. The parameters of the formulation of 26 mM histidine / histidine-HCl, 275 mM trehalose anhydrous, 0.02% polysorbate 80, pH 6.0 are shown in Table 7, and the stability of the lyophilized drug product in the final formulation is provided in Table 8. The solubility profile of the lyophilized formulation ingredients in the final formulation in polypropylene tubes is shown in Table 9.

[0081] [Table 12]

[0082] [Table 13]

[0083] [Table 14]

[0084] Example 6 Development of the Lyophilization Cycle The development of the lyophilization cycle resulted in the parameters shown in Table 10.

[0085]

Table 15

[0086] Using freeze-drying microscopy, the freeze-drying cycle temperatures at the onset of collapse (-33 °C) and complete collapse (-27.3 °C; Figure 6) were evaluated. The glass transition temperature (Tg’) was determined to be -27.14 °C by differential scanning calorimetry (DSC; Figure 7).

[0087] A design space model for the freeze-drying process was generated using Design Expert 7, which employs an approach of design of experiments (DoE). Using the DoE approach, a robust maintenance cycle of < 7 days was achieved. The selected operating conditions indicating the midpoint (approx. Tg’) and the four “corners” of the experimental space are described in Table 11.

[0088]

Table 16

[0089] All experimental cycles had the same ramp rate, freezing temperature, and secondary drying conditions as those described for the final cycle. The final Dulvalumab freeze-drying cycle conditions are shown in Table 12.

[0090]

Table 17

[0091] The main criteria in the development of the freeze-drying cycle are: (1) maintaining the product temperature above the collapse temperature (nearly equivalent to Tg’); (2) ensuring that the primary drying time does not exceed 115 hours to keep the cycle time under 6 days; and (3) providing sufficient robustness for scale-up (shelf temperature of 2 - 3 °C and chamber pressure of 40 mTorr).

[0092] The experimental runs of Table 11 led to the design space of the lyophilization process. The design space generated from five experimental runs is illustrated in Figure 8. Briefly, the mid-cycle point in the chamber, the robustness of the process, and the shelf temperature were determined. The data of the NMF Edwards lyophilizer showed the convergence of the Pirani gauge (Δ10 ubar) in the primary drying process of approximately 103 to 115 hours. This is equivalent to an approximately 10% safety margin (Figure 9). This is consistent with previous large-scale runs, and the Amsco lyophilizer showed the convergence of the product thermocouple within the allotted primary time (Figure 10).

[0093] Example 7 Development of Liquid Formulations 1. Shaking Studies to Optimize Surfactant Levels The suitability of liquid formulations of polysorbate 80 was evaluated via shaking studies in vials. Samples containing 0, 0.005%, 0.01%, 0.02%, 0.03%, 0.04%, and 0.05% (w / v) of polysorbate 80 were placed in tubes, and in the worst-case scenario, the air-to-liquid volume ratio was 6.5 mL. The tubes were vigorously stirred at 600 rpm for 4 hours at ambient temperature. The samples were then examined by visual analysis, A280, BioA, HP-SEC, and microflow imaging (MFI). Control unpenetrated tubes placed at 2 - 8 °C were also analyzed.

[0094] Observable changes were not noticed in percent purity as analyzed by BioA and HP-SEC. Changes in A280 protein concentration were not brought about by shaking. MFI analysis showed that all samples had low levels of >10 μm and >25 μm sub-visible particles (Figure 11). The sub-visible particle levels were overall low in all tubes, and 0.02% of polysorbate 80 was selected as the optimal surfactant level for the liquid formulation.

[0095] 2. Controlled Freeze-Thaw Studies The formulation of 26 mM histidine / histidine-HCl, 275 mM trehalose anhydrous, 0.02% polysorbate 80, pH 6.0 was subjected to 5 controlled freeze-thaw cycles, thawing at 5 °C, and freezing at -40 °C. At the end of the 5th freeze / thaw cycle, purity, appearance, subvisible particles, and protein concentration were determined. No significant change in the quality of the product after the 5th freeze-thaw cycle was observed (Table 13).

[0096]

Table 18

[0097] 3. Overview of formulation stability The stability profile of the lyophilized drug product in the final formulation is provided in Table 14. The results demonstrate that the lyophilized durvalumab in the final formulation remains stable after long-term storage.

[0098]

Table 19

[0099] Example 8 Identification of N-linked oligosaccharides The Fc region of durvalumab contains an N-linked oligosaccharide chain linked to a single site on the heavy chain at Asn-301. Structural characterization of the oligosaccharides on durvalumab is essential for understanding the structural microheterogeneity of the product. It is also important for quality control when the process changes.

[0100] Oligosaccharides cleaved from durvalumab present in the 2-AB labeled N-linked oligosaccharide profile were characterized by ultra-high performance liquid chromatography (UPLC) and contained a small number of glycoforms. Digestion of the 2-AB labeled oligosaccharides with exo-glycosidase was performed as described in Table 15 to demonstrate non-reducing terminal monosaccharide residues. Oligosaccharide profiling was completed as shown in Table 16. LC / MS analysis was used to demonstrate the molecular weight.

[0101]

Table 20

[0102]

Table 21

[0103] The N-linked oligosaccharides in dulvalumab detected as significant peaks in UPLC were identified, and the results are shown in Figure 12. The major glycoforms of dulvalumab are fucosylated biantennary complex oligosaccharides that do not have terminal galactose residues (G0f), or mono-galactosylated (G1f) and di-galactosylated (G2f) forms. Minor complex glycoforms were afucosylated G0 and G1, truncated G0f and G0 forms without N-acetylglucosamine (GlcNAc), sialylated G1f and G2f forms (G1f+NAc, G2f+NAc, or G2f+2NAc), and bisecting structure G0fb and G1fb. High mannose glycoforms (Man4, Man5, Man6, Man7, and Man8) were also present depending on the maturation of Chinese hamster ovary (CHO) cells when IgG was harvested.

[0104] It should be understood that the foregoing disclosure emphasizes certain specific embodiments of the present invention, and that all equivalent variations or alternatives thereof are within the spirit and scope of the present invention as set forth in the appended claims.

[0105]

Table 22

[0106] [Sequence Listing] SEQUENCE LISTING <110> MEDIMMUNE LIMITED <120> FORMULATIONS OF HUMAN ANTI-PD-L1 ANTIBODIES <130> PA25-022 <150> US62 / 662,324 <151> 2018-04-25 <160> 8 <170> PatentIn version 3.5 <210> 1 <211> 108 <212> PRT <213> Homo sapiens <400> 1 Glu Ile Val Leu Thr Gln Ser Pro Gly Thr Leu Ser Leu Ser Pro Gly 1 5 10 15 Glu Arg Ala Thr Leu Ser Cys Arg Ala Ser Gln Arg Val Ser Ser Ser 20 25 30 Tyr Leu Ala Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Arg Leu Leu 35 40 45 Ile Tyr Asp Ala Ser Ser Arg Ala Thr Gly Ile Pro Asp Arg Phe Ser 50 55 60 Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Arg Leu Glu 65 70 75 80 Pro Glu Asp Phe Ala Val Tyr Tyr Cys Gln Gln Tyr Gly Ser Leu Pro 85 90 95 Trp Thr Phe Gly Gln Gly Thr Lys Val Glu Ile Lys 100 105 <210> 2 <211> 121 <212> PRT <213> Homo sapiens <400> 2 Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Arg Tyr 20 25 30 Trp Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ala Asn Ile Lys Gln Asp Gly Ser Glu Lys Tyr Tyr Val Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Ser Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Arg Glu Gly Gly Trp Phe Gly Glu Leu Ala Phe Asp Tyr Trp Gly 100 105 110 Gln Gly Thr Leu Val Thr Val Ser Ser 115 120 <210> 3 <211> 10 <212> PRT <213> Homo sapiens <400> 3 Gly Phe Thr Phe Ser Arg Tyr Trp Met Ser 1 5 10 <210> 4 <211> 17 <212> PRT <213> Homo sapiens <400> 4 Asn Ile Lys Gln Asp Gly Ser Glu Lys Tyr Tyr Val Asp Ser Val Lys 1 5 10 15 Gly <210> 5 <211> 12 <212> PRT <213> Homo sapiens <400> 5 Glu Gly Gly Trp Phe Gly Glu Leu Ala Phe Asp Tyr 1 5 10 <210> 6 <211> 12 <212> PRT <213> Homo sapiens <400> 6 Arg Ala Ser Gln Arg Val Ser Ser Ser Tyr Leu Ala 1 5 10 <210> 7 <211> 7 <212> PRT <213> Homo sapiens <400> 7 Asp Ala Ser Ser Arg Ala Thr 1 5 <210> 8 <211> 9 <212> PRT <213> Homo sapiens <400> 8 Gln Gln Tyr Gly Ser Leu Pro Trp Thr 1 5

Claims

1. (a) 40 mg / mL to 50 mg / mL of an anti-PD-L1 antibody; (b) 15 mM to 35 mM buffer; (c) 255 mM to 275 mM disaccharides; and (d) 0.01% (w / v) to 0.05% (w / v) of a surfactant; An antibody formulation comprising: wherein the pH of the formulation is between pH 5.5 and pH 7.

2.

2. The anti-PD-L1 antibody comprises: (a) a light chain variable domain comprising the amino acid sequence of SEQ ID NO: 1, and a heavy chain variable domain comprising the amino acid sequence of SEQ ID NO: 2; or (b) a VH CDR1 having the amino acid sequence of SEQ ID NO:3; VH CDR2 having the amino acid sequence of SEQ ID NO:4; a VH CDR3 having the amino acid sequence of SEQ ID NO:5; VL CDR1 having the amino acid sequence of SEQ ID NO:6; A VL CDR2 having the amino acid sequence of SEQ ID NO:7; and VL CDR3 having the amino acid sequence of SEQ ID NO:8, The antibody formulation of claim 1 , comprising:

3. The antibody formulation of claim 1, wherein the buffer is a histidine / histidine-HCl buffer.

4. 2. The antibody formulation of claim 1, wherein the disaccharide is trehalose dihydrate.

5. The antibody formulation of claim 1 , wherein the disaccharide is sucrose.

6. The antibody formulation of claim 1 , wherein the surfactant is polysorbate 80.

7. The antibody formulation of any one of claims 1 to 6, wherein the formulation is a liquid formulation, a frozen formulation, a lyophilized formulation, or a reconstituted formulation.

8. 2. The antibody formulation of claim 1, wherein less than about 1% of the anti-PD-L1 antibody forms aggregates upon storage at 40° C. for about 1 month as measured by high pressure size exclusion chromatography (HP-SPEC).

9. 2. The antibody formulation of claim 1, wherein at least 97% of the anti-PD-L1 antibody is present as a monomer after storage at about 40° C. for about 1 month as measured by high pressure size exclusion chromatography (HP-SEC).

10. 2. The antibody formulation of claim 1, wherein at least 99% of the anti-PD-L1 antibody is present as a monomer after storage at about 40° C. for about 1 month as measured by high pressure size exclusion chromatography (HP-SEC).

11. 2. The antibody formulation of claim 1, wherein at least 98% of the anti-PD-L1 antibody is present as a monomer after storage at about 5° C. for about 1 month as measured by high pressure size exclusion chromatography (HP-SEC).

12. 2. The antibody formulation of claim 1, wherein the antibody formulation remains stable after at least three freeze / thaw cycles.

13. (a) 50 mg / mL human anti-PD-L1 antibody; (b) 26 mM histidine / histidine-HCl buffer; (c) 275 mM trehalose dihydrate; and (d) 0.02% (w / v) polysorbate 80; An antibody formulation comprising: wherein the pH of the formulation is pH 6.

0.

14. (a) 50 mg / mL human anti-PD-L1 antibody; (b) 25 mM histidine / histidine-HCl buffer; (c) 265 mM trehalose dihydrate; and (d) 0.02% (w / v) polysorbate 80; An antibody formulation comprising: wherein the pH of the formulation is pH 5.

5.

15. (a) 50 mg / mL human anti-PD-L1 antibody; (b) 25 mM histidine / histidine-HCl buffer; (c) 265 mM trehalose dihydrate; and (d) 0.02% (w / v) polysorbate 80; An antibody formulation comprising: wherein the pH of the formulation is pH 6.

5.

16. (a) 50 mg / mL human anti-PD-L1 antibody; (b) 25 mM histidine / histidine-HCl buffer; (c) 265 mM trehalose dihydrate; and (d) 0.04% (w / v) polysorbate 80; An antibody formulation comprising: wherein the pH of the formulation is pH 6.

0.

17. (a) 50 mg / mL human anti-PD-L1 antibody; (b) 25 mM histidine / histidine-HCl buffer; (c) 265 mM sucrose; and (d) 0.02% (w / v) polysorbate 80; An antibody formulation comprising: wherein the pH of the formulation is pH 6.

0.

18. The antibody formulation of any one of claims 13 to 17, wherein the human anti-PD-L1 antibody comprises a light chain having the amino acid sequence of SEQ ID NO:1 and a heavy chain having the amino acid sequence of SEQ ID NO:

2.

19. The human anti-PD-L1 antibody comprises: A VH CDR1 having the amino acid sequence of SEQ ID NO:3; and A VH CDR2 having the amino acid sequence of SEQ ID NO:4; and A VH CDR3 having the amino acid sequence of SEQ ID NO:5; and A VL CDR1 having the amino acid sequence of SEQ ID NO:6; and A VL CDR2 having the amino acid sequence of SEQ ID NO:7; and VL CDR3 having the amino acid sequence of SEQ ID NO:8, The antibody formulation of any one of claims 13 to 17, comprising:

20. The antibody formulation of any one of claims 13 to 17, wherein the formulation is a liquid formulation, a frozen formulation, a lyophilized formulation, or a reconstituted formulation.

21. 18. The antibody formulation of any one of claims 13-17, wherein less than about 1% of the human anti-PD-L1 antibody forms aggregates upon storage at 40°C for about 1 month as determined by high pressure size exclusion chromatography (HP-SPEC).

22. 18. The antibody formulation of any one of claims 13-17, wherein at least 97% of the human anti-PD-L1 antibody is present as a monomer after storage at about 40°C for about 1 month as measured by high pressure size exclusion chromatography (HP-SEC).

23. 18. The antibody formulation of any one of claims 13-17, wherein at least 99% of the human anti-PD-L1 antibody is present as a monomer after storage at about 40°C for about 1 month as measured by high pressure size exclusion chromatography (HP-SEC).

24. 18. The antibody formulation of any one of claims 13-17, wherein at least 98% of the human anti-PD-L1 antibody is present as a monomer after storage at about 5°C for about 1 month as measured by high pressure size exclusion chromatography (HPSEC).

25. The antibody formulation of any one of claims 13 to 17, wherein the antibody formulation remains stable after at least three freeze / thaw cycles.

26. 1. A composition comprising: (a) an anti-PD-L1 antibody comprising a light chain having an amino acid sequence that is at least 90% identical to the amino acid sequence of SEQ ID NO:1, and a heavy chain having an amino acid sequence that is at least 90% identical to the amino acid sequence of SEQ ID NO:2; and (b) a major form of the antibody that comprises greater than 45% of the protein in the composition as measured using capillary isoelectric focusing (cIEF) of the composition; A composition comprising:

27. 1. A composition comprising: (a) an anti-PD-L1 antibody comprising a light chain having an amino acid sequence that is at least 90% identical to the amino acid sequence of SEQ ID NO:1, and a heavy chain having an amino acid sequence that is at least 90% identical to the amino acid sequence of SEQ ID NO:2; (b) a major form of said antibody that comprises 45% or more of the protein in said composition as measured using cIEF of said composition; and (c) an acidic form of the antibody comprising 45% to 50% of the protein in said composition as measured using cIEF of said composition; A composition comprising:

28. 1. A composition comprising: (a) an anti-PD-L1 antibody comprising a light chain having an amino acid sequence that is at least 90% identical to the amino acid sequence of SEQ ID NO:1, and a heavy chain having an amino acid sequence that is at least 90% identical to the amino acid sequence of SEQ ID NO:2; (b) a major form of said antibody that comprises 45% or more of the protein in said composition as measured using cIEF of said composition; and (c) the basic form of the antibody comprises between 18% and 23% of the protein in the composition as measured using cIEF of the composition; A composition comprising:

29. 1. A composition comprising: (a) an anti-PD-L1 antibody comprising a light chain having an amino acid sequence that is at least 90% identical to the amino acid sequence of SEQ ID NO:1, and a heavy chain having an amino acid sequence that is at least 90% identical to the amino acid sequence of SEQ ID NO:2; (b) a major form of said antibody that comprises greater than 45% of the protein in said composition as measured using cIEF of said composition; (c) an acidic form of the antibody that comprises 45% to 50% of the protein in the composition as measured using cIEF of the composition; and (d) the basic form of the antibody comprises between 18% and 23% of the protein in the composition as measured using cIEF of the composition; A composition comprising:

30. 1. A composition comprising: (a) an anti-PD-L1 antibody comprising a light chain having an amino acid sequence that is at least 90% identical to the amino acid sequence of SEQ ID NO:1, and a heavy chain having an amino acid sequence that is at least 90% identical to the amino acid sequence of SEQ ID NO:2; and (b) the glycan structure of the anti-PD-L1 antibody comprises G0f, G1f, G2f, and G0 glycoforms.

31. 31. The composition of claim 30, wherein the glycan structure of the anti-PD-L1 antibody has a content of the G0f, G1f, G2f, and G0 glycoforms that is greater than 90%.

32. 31. The composition of claim 30, wherein the anti-PD-L1 antibody comprises a G0f content of 71.9%, a G1f content of 18.4%, a G2f content of 1.5%, and a G0 content of 1.9%.

33. 1. A composition comprising: (a) an anti-PD-L1 antibody comprising a light chain having an amino acid sequence that is at least 90% identical to the amino acid sequence of SEQ ID NO:1, and a heavy chain having an amino acid sequence that is at least 90% identical to the amino acid sequence of SEQ ID NO:2; (b) between 1.5% and 2.5% of the anti-PD-L1 antibody forms aggregates, as determined by high pressure size exclusion chromatography (HP-SPEC); and (c) the composition, wherein 97% to 98% of the anti-PD-L1 antibody is present as a monomer, as measured by HP-SEC.

34. The PD-L1 antibody comprises: A VH CDR1 having the amino acid sequence of SEQ ID NO:3; and A VH CDR2 having the amino acid sequence of SEQ ID NO:4; and A VH CDR3 having the amino acid sequence of SEQ ID NO:5; and A VL CDR1 having the amino acid sequence of SEQ ID NO:6; and A VL CDR2 having the amino acid sequence of SEQ ID NO:7; and VL CDR3 having the amino acid sequence of SEQ ID NO:8, The composition of any one of claims 26 to 33, comprising:

35. (a) 50 mg / mL human anti-PD-L1 antibody; (b) 26 mM histidine / histidine-HCl buffer; (c) 275 mM trehalose dihydrate; and (d) 0.02% (w / v) polysorbate 80; An antibody formulation comprising: wherein the pH of the formulation is pH 6.0, and wherein the human anti-PD-L1 antibody comprises a light chain having the amino acid sequence of SEQ ID NO:1, and a heavy chain having the amino acid sequence of SEQ ID NO:2.

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