Anti-complement C1s antibody preparation

Abstract

Provided herein is a composition that can be stably stored for a long time and contains a humanized antibody (anti-C1s antibody) that specifically binds to complement component C1s. In addition to the humanized antibody, this composition may contain arginine or a salt thereof.

Description

Technical Field

[0001] Related Applications This application claims the benefit of U.S. Provisional Patent Application No. 63 / 352,475, filed on June 15, 2022, under 35 U.S.C. § 119(e), the entire content of which is incorporated herein by reference.

[0002] Reference to Electronic Sequence Listing The content of the electronic sequence listing (B155370017WO00-SEQ-JRV.xml; size: 21,726 bytes; created on June 9, 2023) is incorporated herein by reference in its entirety.

Background Art

[0003] The complement system is well known as an effector mechanism of the immune response and not only provides protection against pathogens and other harmful substances but also contributes to the recovery from injury. The classical complement pathway is initiated by the activation of the first component of complement, called the C1 complex, which includes the C1q, C1r, and C1s proteins. When C1 binds to immune complexes, a diisopropylfluorophosphate (DFP)-sensitive serine protease, which is a component of C1s, cleaves the complement components C4 and C2, initiating the classical complement pathway. The classical complement pathway appears to affect many diseases and disorders, and there is a need for stable formulations of antibodies that target this pathway.

Summary of the Invention

Means for Solving the Problems

[0004] The present disclosure provides a composition comprising a humanized antibody that specifically binds to complement C1s protein (i.e., a humanized anti-complement C1s antibody, also referred to herein as a "humanized anti-C1s antibody", a "humanized C1s antibody", or a "subject antibody"). The present disclosure also provides a method of treating a complement-mediated disease or disorder, the method comprising administering a composition of the present disclosure.

[0005] The target antibody has a higher tendency to self-associate and also has a tendency for the aspartic acid residue at position 32 of the light chain variable domain sequence located in the first complementarity-determining region (CDR1) of the light chain to isomerize (LC D32 isomerization). A series of comprehensive tests were conducted to understand the influence of the formulation composition and processing conditions on the biophysical and chemical stability of the target antibody. The tests described herein show that the presence of arginine or its salt reduces the D32 isomerization rate. In sample formulations using various batches of the target antibody, when the compositions of the present disclosure were used, it was found that they showed resistance to isomerization under refrigerated, room temperature, and accelerated storage conditions. The compositions of the present disclosure maintain the stability of the antibody during long-term storage, protect it from physicochemical stress, and at the same time prevent LC D32 isomerization, antibody aggregation, and deamidation. Furthermore, the compositions of the present disclosure also enable the target antibody to be formulated as a liquid.

[0006] In one aspect, the present disclosure provides a composition comprising: (a) a humanized antibody that specifically binds to complement component C1s, the humanized antibody comprising a light chain (LC) complementarity-determining region (CDR) 1 comprising the amino acid sequence of SEQ ID NO: 1, an LC CDR2 comprising the amino acid sequence of SEQ ID NO: 2, an LC CDR3 comprising the amino acid sequence of SEQ ID NO: 3, a heavy chain (HC) CDR1 comprising the amino acid sequence of SEQ ID NO: 4, an HC CDR2 comprising the amino acid sequence of SEQ ID NO: 5, and an HC CDR3 comprising the amino acid sequence of SEQ ID NO: 6; and (b) about 50 mM to about 200 mM of arginine or a salt thereof.

[0007] In some embodiments, the composition comprises about 100 mM to about 200 mM or about 100 mM to about 150 mM of arginine or a salt thereof. In some embodiments, the arginine salt is arginine hydrochloride, arginine citrate, arginine oxalate, arginine phosphate, arginine succinate, or arginine tartrate. In some embodiments, the arginine salt is arginine hydrochloride. In some embodiments, the composition comprises about 150 mM of arginine hydrochloride. In some embodiments, the arginine hydrochloride is L-arginine hydrochloride.

[0008] In some embodiments, the composition comprises a buffer. In some embodiments, the composition comprises the buffer at a concentration of about 1 mM to about 50 mM, about 5 mM to about 25 mM, or about 10 mM to about 20 mM. In some embodiments, the buffer is histidine, acetate, citrate, oxalate, phosphate, succinate, or tartrate. In some embodiments, the buffer is histidine. In some embodiments, the composition comprises about 10 mM of histidine.

[0009] In some embodiments, the composition further comprises a stabilizer. In some embodiments, the composition further comprises the stabilizer at a concentration of about 1% to about 8% (w / v) or about 1% to about 5% (w / v). In some embodiments, the stabilizer is sucrose, sorbitol, or trehalose. In some embodiments, the stabilizer is sucrose. In some embodiments, the composition comprises about 3% (w / v) of sucrose.

[0010] In some embodiments, the composition further comprises a chelating agent. In some embodiments, the composition comprises the chelating agent at a concentration of about 1 μM to about 50 μM, about 5 μM to about 25 μM, or about 10 μM to about 25 μM. In some embodiments, the chelating agent is diethylenetriaminepentaacetic acid (DTPA), ethylenediaminetetraacetic acid (EDTA). In some embodiments, the composition comprises about 10 μM of diethylenetriaminepentaacetic acid (DTPA). In some embodiments, the chelating agent is methionine. In some embodiments, the composition comprises about 5 mM to about 10 mM of methionine.

[0011] In some embodiments, the composition further comprises a surfactant. In some embodiments, the composition comprises the surfactant at a concentration of about 0.01% to about 0.1% (w / v) or about 0.03% to about 0.06% (w / v). In some embodiments, the surfactant is polysorbate 80 (PS80) or poloxamer 188 (P188). In some embodiments, the surfactant is PS80, and optionally, the composition comprises about 0.06% (w / v) of polysorbate 80.

[0012] In some embodiments, the composition has a pH of about 6 to about 7.5, about 6 to about 7, about 6.5 to about 7.5, or about 6.5 to about 7.1. In some embodiments, the composition has a pH of about 6.8.

[0013] In some embodiments, the composition comprises an anti-C1s antibody at about 50 mg / mL to about 250 mg / mL or about 100 mg / mL to about 200 mg / mL. In some embodiments, the composition comprises an anti-C1s antibody at about 150 mg / mL.

[0014] In some embodiments, the composition comprises: (a) an antibody at about 50 mg / mL to about 250 mg / mL, (b) arginine HCl at about 50 mM to about 200 mM, (c) histidine at about 1 mM to about 50 mM, (d) sucrose at about 1% to about 8% (w / v), (e) diethylenetriaminepentaacetic acid (DTPA) at about 1 μM to about 50 μM, and (f) polysorbate 80 (PS80) at about 0.01% to about 0.1% (w / v), and the composition has a pH of about 6 to about 7.5. In some embodiments, the arginine hydrochloride is L-arginine hydrochloride.

[0015] In some embodiments, the composition comprises: (a) an antibody at about 150 mg / mL, (b) arginine HCl at about 150 mM, (c) histidine at about 10 mM, (d) sucrose at about 3% (w / v), (e) DTPA at about 10 μM, and (f) PS80 at about 0.06% (w / v), and the composition has a pH of about 6.5 to about 7.1 or about 6.8. In some embodiments, the arginine hydrochloride is L-arginine hydrochloride.

[0016] In some embodiments, the antibody comprises a light chain variable region (VL) comprising the amino acid sequence of SEQ ID NO: 7 and a heavy chain variable region (VH) comprising the amino acid sequence of SEQ ID NO: 8. In some embodiments, the antibody is a Fab fragment, F(ab’)2 fragment, scFv or Fv. In some embodiments, the antibody comprises a heavy chain constant region of isotype IgG4. In some embodiments, the IgG4 constant region comprises proline, glutamic acid, leucine and serine substitutions at amino acid residues 108, 115, 308 and 31, respectively, relative to the IgG4 constant region sequence of SEQ ID NO: 11. In some embodiments, the antibody comprises a heavy chain constant region comprising the amino acid sequence of SEQ ID NO: 13. In some embodiments, the antibody comprises a light chain comprising the amino acid sequence of SEQ ID NO: 9 and a heavy chain comprising the amino acid sequence of SEQ ID NO: 10.

[0017] In some embodiments, the antibody has a lower isomerization rate at D32 of the light chain located in CDR1 compared to the antibody in the corresponding formulation without arginine or its salt, and optionally, the isomerization of D32 is determined by whole peptide map analysis. In some embodiments, the antibody has an isomerization rate that is at least about 2-3% lower per week at 40 °C, at least about 1-3% lower per month at 25 °C or at least about 0.4-0.6% lower per month at 5 °C compared to the antibody in the corresponding formulation without arginine or its salt, or (ii) has an isomerization rate that is about 10% lower after storage at 25 °C for 12 weeks compared to the antibody in the corresponding formulation without arginine or its salt.

[0018] In some embodiments, the antibody has an isomerization rate of less than 7.5% after storage at 5 °C for 12 weeks, less than 20% after storage at 25 °C for 12 weeks or less than 35% after storage at 40 °C for 12 weeks.

[0019] In some embodiments, the composition is in liquid form, lyophilized form or reconstituted lyophilized form. In some embodiments, the composition is in liquid form.

[0020] In one aspect, the present disclosure provides a container for containing the compositions of the present disclosure. In some embodiments, the container is a vial or a syringe. In some embodiments, the container is a syringe. In some embodiments, the syringe is a prefilled syringe.

[0021] In one aspect, the present disclosure provides a kit or an article of manufacture comprising the container of the present disclosure.

[0022] In one aspect, the present disclosure provides a pharmaceutical unit dosage form suitable for parenteral administration to a human, comprising the composition of the present disclosure within a container.

[0023] In one aspect, the present disclosure provides a method comprising administering the composition of the present disclosure to a human.

[0024] In one aspect, the present disclosure provides a method for reducing the level of complement component cleavage products in a human, comprising administering the composition of the present disclosure to the human.

[0025] In one aspect, the present disclosure provides a method for inhibiting C1s-mediated cleavage of complement components in a human, comprising administering the composition of the present disclosure to the human.

[0026] In some embodiments, the human has cold agglutinin disease (CAD), immune thrombocytopenic purpura (ITP), chronic inflammatory demyelinating polyneuropathy (CIDP) or antibody-mediated rejection (AMR).

[0027] In one aspect, the present disclosure provides a method for treating a complement-mediated disease in a human in need thereof, comprising administering the composition of the present disclosure to the human. In some embodiments, the complement-mediated disease is cold agglutinin disease (CAD), immune thrombocytopenic purpura (ITP), chronic inflammatory demyelinating polyneuropathy (CIDP) or antibody-mediated rejection (AMR).

[0028] In some embodiments, the administration is intravenous or subcutaneous.

[0029] In one aspect, the present disclosure provides a drug delivery device including a primary container containing the composition of the present disclosure, the drug delivery device being a sleeve-activated self-injector for manually inserting a needle.

[0030] Each of WO 2014 / 066744 pamphlet titled Anti-Complement C1s Antibodies and Uses Thereof filed on October 25, 2013, WO 2016 / 164358 pamphlet titled Humanized Anti-C1s Antibodies and Methods of Use Thereof filed on April 5, 2016, and WO 2018 / 071676 pamphlet titled Anti-C1s Antibodies and Methods of Use Thereof filed on October 12, 2017 is hereby incorporated by reference in its entirety.

Brief Description of the Drawings

[0031]

Figure 1

Figures 2A - 2D

Figures 3A - 3E

Figures 4A - 4E

Figures 5A - 5D

Figures 6A - 6F

Figures 7A - 7F

Figures 8A - 8E

Figure 9

Figures 10A - 10E

Figures 11A - 11E

Figures 12A - 12E

Figures 13A - 13E

Figures 14A - 14C

Figures 15A - 15E

Figures 16A - 16C

Figures 17A - 17G

Figures 18A - 18E

Figures 19A - 19G

Figures 20A - 20C

Figures 21A - 21G

Figures 22A - 22C

Figures 23A - 23F

Figures 24A - 24E

Figures 25A - 25G

Figures 26A - 26G

Figures 27A - 27E

Figures 28A - 28G

Figures 29A - 29E

Figures 30A - 30G

Figures 31A - 31E

Figures 32A - 32E

Figures 33A - 33F

Figures 34A - 34I

Mode for Carrying Out the Invention

[0032] Humanized anti-C1s antibody The compositions of the present disclosure include humanized anti-C1s antibodies. The humanized anti-C1s antibodies of the present disclosure bind to and inhibit active C1s within the classical pathway (CP). The complement system is a component of the innate immune system that mediates humoral immunity. The mechanism of this antibody is specific to the CP and does not affect the functions of the lectin pathway and the alternative pathway.

[0033] In some embodiments, the humanized anti-C1s antibodies of the present disclosure inhibit C1s-mediated cleavage of complement component C4, for example, by inhibiting the enzymatic activity of the serine protease domain of C1s. In some embodiments, the humanized anti-C1s antibodies of the present disclosure inhibit C1s-mediated cleavage of complement component C2. In some embodiments, the humanized anti-C1s antibodies of the present disclosure inhibit C1s-mediated cleavage of C4 and C2.

[0034] In some embodiments, the humanized anti-C1s antibodies of the present disclosure bind to a complement C1s protein having the amino acid sequence represented by SEQ ID NO: 15. The amino acid sequence of SEQ ID NO: 15 shown below represents the complement C1s protein of Homo sapiens.

Chemical Formula

[0035] In some embodiments, the humanized anti-C1s antibody of the present disclosure binds to human complement C1s protein with a dissociation constant (KD) of 2.5 nM or less. In some embodiments, the humanized anti-C1s antibody of the present disclosure binds to human complement C1s protein with a KD of 2 nM or less. In some embodiments, the humanized anti-C1s antibody of the present disclosure binds to human complement C1s protein with a KD of 1 nM or less. In some embodiments, the humanized anti-C1s antibody of the present disclosure binds to human complement C1s protein with a KD of 0.9 nM or less, 0.8 nM or less, 0.7 nM or less, 0.6 nM or less, 0.5 nM or less, 0.4 nM or less, 0.3 nM or less, 0.2 nM or less, or 0.1 nM or less. In some embodiments, the humanized anti-C1s antibody of the present disclosure binds to human complement C1s protein with a KD of 0.3 nM or less. In some embodiments, the humanized anti-C1s antibody of the present disclosure binds to human complement C1s protein with a KD of 0.2 nM or less. In some embodiments, the humanized anti-C1s antibody of the present disclosure binds to human complement C1s protein with a KD of 0.1 nM or less. Methods for measuring the binding of an antibody to human complement C1s protein can be determined by those skilled in the art.

[0036] In some embodiments, the humanized anti-C1s antibody of the present disclosure binds to human complement C1s protein with a KD of 90 pM or less, 80 pM or less, 70 pM or less, 60 pM or less, 50 pM or less, 40 pM or less, 30 pM or less, 20 pM or less, 10 pM or less, 9 pM or less, 8 pM or less, 7 pM or less, 6 pM or less, 5 pM or less, 4 pM or less, 3 pM or less, 2 pM or less, or 1 pM or less.

[0037] In some embodiments, the humanized anti-C1s antibody of the present disclosure inhibits the classical complement pathway at an inhibitory concentration (IC -8 of 10 -9 M or less, 5×10 -9 M or less, or 10 50 M or less.

[0038] "Antibody" includes antibodies or immunoglobulins of all isotypes, including, but not limited to, humanized antibodies and chimeric antibodies. Antibodies can be single-chain antibodies (scAbs) or single-domain antibodies (dAbs) (e.g., single-domain heavy-chain antibodies or single-domain light-chain antibodies; see Holt et al. (2003) Trends Biotechnol. 21:484). The term "antibody" also includes fragments of antibodies (antibody fragments) that retain specific binding to an antigen. "Antibody" includes proteins of the variable regions of the heavy chain (V H ) and light chain (V L ) of an antibody fused by connecting them with a short linker peptide, as well as single-chain variable fragments (scFvs) and non-covalent dimers of scFv fragments containing V H and V L linked by a small linker peptide (Zapata et al., Protein Eng. 8(10):1057-1062 (1995)). Other fusion proteins containing the antigen-binding portion of an antibody and a non-antibody protein are also included in the term "antibody".

[0039] "Antibody fragment" includes a part of an intact antibody, such as the antigen-binding region or variable region of an intact antibody. Examples of antibody fragments include antigen-binding fragments (Fab), Fab’, F(ab’)2, variable domain Fv fragments (Fv), Fd fragments, and antigen-binding fragments of chimeric antigen receptors.

[0040] Digestion of an antibody with papain generates two identical antigen-binding fragments, called "Fab" fragments, each having a single antigen-binding site, and the remainder, called the "Fc" fragment, which has a name indicating its ability to be easily crystallized. Treatment with pepsin generates F(ab’)2 fragments, which have two antigen-binding sites and can still cross-link antigens.

[0041] "Fv" is the smallest antibody fragment that contains the complete antigen recognition site and antigen binding site. This region contains a dimer of one heavy chain variable domain and one light chain variable domain that are non-covalently and tightly associated. The antigen binding site on the surface of the V-V dimer is defined by this structure. In summary, the six CDRs confer antigen binding specificity to the antibody. However, even a single variable domain (or half of the Fv containing only three CDRs specific for the antigen) has the ability to recognize and bind the antigen, although with lower affinity than the entire binding site. H -V L The antigen binding site on the surface of the dimer is defined by this structure. In summary, the six CDRs confer antigen binding specificity to the antibody. However, even a single variable domain (or half of the Fv containing only three CDRs specific for the antigen) has the ability to recognize and bind the antigen, although with lower affinity than the entire binding site.

[0042] The "Fab" fragment contains the constant domain of the light chain and the first constant domain of the heavy chain (CH1). The Fab' fragment differs from the Fab fragment in that it additionally has several residues containing at least one cysteine derived from the hinge region of the antibody at the carboxyl terminus of the heavy chain CH1 domain. Fab'-SH herein is the name for Fab' in which the cysteine residue of the constant domain has a free thiol group. The F(ab')2 antibody fragment was originally produced as a pair of Fab' fragments having a hinge cysteine between them. Chemically conjugated antibody fragments are also known.

[0043] The "scFv" antibody fragment contains the V H and V L regions, which are present in a single polypeptide chain. In some embodiments, the Fv polypeptide further includes a polypeptide linker between the V H region and the V L region, whereby the scFv can form a structure desirable for binding to the antigen. For an overview of scFv, see Pluckthun in The Pharmacology of Monoclonal Antibodies, vol. 113, Rosenberg and Moore eds., Springer-Verlag, New York, pp. 269-315 (1994).

[0044] "Diabody" refers to a small antibody fragment having two antigen-binding sites, which fragment contains a V L linked to a V H in the same polypeptide chain (V H -V L ). By using a linker that is too short for these two domains within the same chain to pair with each other, these domains are forced to pair with complementary domains on the other chain, creating two antigen-binding sites. Diabodies are described in more detail, for example, by Hollinger et al. Proc. Natl. Acad. Sci. USA 90:6444-6448 (1993).

[0045] Antibodies can be monovalent or divalent. Antibodies can be Ig monomers, which are "Y-shaped" molecules composed of four polypeptide chains: two heavy chains and two light chains linked by disulfide bonds.

[0046] Antibodies can be detectably labeled, for example, using radioisotopes, enzymes that produce detectable products, and / or fluorescent proteins. Furthermore, antibodies can be conjugated to other moieties, such as components of specifically binding pairs, such as the biotin component of a specifically binding biotin-avidin pair. Antibodies can also be bound to solid supports, such as, but not limited to, polystyrene plates and / or beads.

[0047] An "isolated" antibody is one that has been identified and separated and / or recovered from components in its natural environment (i.e., it is not naturally occurring). Contaminants in its natural environment are substances that would interfere with the use of the antibody (e.g., for diagnostic or therapeutic uses), and can include enzymes, hormones, and other proteinaceous or non-proteinaceous solutes. In some embodiments, the antibody is purified until (1) the weight of the antibody, as determined, for example, by the Lowry method, is greater than 90%, greater than 95%, greater than 98% or greater than 99%; (2) sufficient to obtain at least 15 residues of the N-terminal or internal amino acid sequence using a spinning cup sequenator; or (3) homogeneity is shown by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) using Coomassie blue or silver staining under reducing or non-reducing conditions. Isolated antibodies include antibodies present in recombinant cells, since in that case at least one of the components of the natural environment will not be present. In some embodiments, the isolated antibody is prepared by at least one purification step.

[0048] A "monoclonal antibody" is an antibody produced by the same population of cells that are produced by repeated replication of a single cell. That is, the clone of cells produces only a single species of antibody. Monoclonal antibodies can be produced using hybridoma production techniques, but other production methods known to those of skill in the art can also be used (e.g., antibodies obtained from antibody phage display libraries).

[0049] "Complementary determining region (CDR)" refers to the non - contiguous antigen - binding sites found within the variable regions of both the heavy and light chain polypeptides. The CDRs are described in Lefranc et al. (2003) Developmental and Comparative Immunology 27:55; Kabat et al., J. Biol. Chem. 252:6609 - 6616 (1977); Kabat et al., U.S. Dept. of Health and Human Services, "Sequences of proteins of immunological interest" (1991); Chothia et al., J. Mol. Biol. 196:901 - 917 (1987); and MacCallum et al., J. Mol. Biol. 262:732 - 745 (1996), and when compared to each other, their definitions include overlapping or subsets of amino acid residues. Nevertheless, applying any of the Kabat, Lefranc, Chothia, or MacCallum definitions (also called numbering systems) to reference the CDRs of an antibody or a grafted antibody or variants thereof is intended to be within the scope of the terms defined and used herein.

[0050] The terms "LC CDR1", "LC CDR2", and "LC CDR3" refer to the first, second, and third CDRs of the light - chain variable region, respectively. The terms "HC CDR1", "HC CDR2", and "HC CDR3" as used herein refer to the first, second, and third CDRs of the heavy - chain variable region, respectively. The terms "CDR1", "CDR2", and "CDR3" as used herein refer to the first, second, and third CDRs of the variable region of either chain, respectively.

[0051] As used with respect to an antibody variable region, "framework" includes all amino acid residues outside of the CDR regions within the variable region of the antibody. The framework of the variable region is generally a discontinuous amino acid sequence containing only the amino acids outside of the CDRs. The "framework region" includes each domain of the framework separated by the CDRs.

[0052] A "humanized antibody" is an antibody that contains a portion of antibodies of different origins and contains an amino acid sequence derived from a human in at least a part thereof. For example, a humanized antibody may contain a portion derived from an antibody of non-human origin, such as a mouse, having the required specificity, and a portion derived from a human-derived antibody sequence (e.g., a chimeric immunoglobulin), which is chemically bonded by a conventional technique (e.g., synthesis) or produced as a continuous polypeptide using genetic engineering techniques (e.g., a continuous polypeptide chain can be produced by expressing DNA encoding the protein portion of a chimeric antibody). Another example of a humanized antibody is an antibody that contains at least one chain containing a CDR derived from an antibody of non-human origin and a framework region of a human-derived light chain and / or heavy chain (e.g., a CDR-grafted antibody with or without a modified framework). Chimeric or CDR-grafted single-chain antibodies are also included in the term "humanized immunoglobulin". See, for example, U.S. Patent No. 4,816,567 to Cabilly et al.; European Patent No. 0,125,023B1 to Cabilly et al.; U.S. Patent No. 4,816,397 to Boss et al.; European Patent No. 0,120,694B1 to Boss et al.; International Publication No. 86 / 01533 pamphlet to Neuberger, M.S. et al.; European Patent No. 0,194,276B1 to Neuberger, M.S. et al.; U.S. Patent No. 5,225,539 to Winter; European Patent No. 0,239,400B1 to Winter; European Patent Application Publication No. 0,519,596A1 to Padlan, E.A. et al. For single-chain antibodies, also see U.S. Patent No. 4,946,778 to Ladner et al.; U.S. Patent No. 5,476,786 to Huston; and Bird, R.E. et al., Science, 242:423-426 (1988).

[0053] In some embodiments, the humanized antibody is made using synthetic and / or recombinant nucleic acids to create a gene (e.g., cDNA) encoding the desired humanized chain. For example, a nucleic acid (e.g., DNA) sequence encoding a humanized variable region can be constructed using PCR mutagenesis methods that modify a DNA sequence encoding a human or humanized chain, e.g., a DNA template from a previously humanized variable region (e.g., see Kamman, M., et al., Nucl. Acids Res., 17:5404 (1989)); Sato, K., et al., Cancer Research, 53:851-856 (1993); Daugherty, B.L. et al., Nucleic Acids Res., 19(9):2471-2476 (1991); and Lewis, A.P. and J.S.Crowe, Gene, 101:297-302 (1991)). Variants can also be readily made by using these or other suitable methods. For example, mutations can be induced in the cloned variable region and sequences encoding variants with the desired specificity can be selected (e.g., from a phage library; e.g., see U.S. Patent No. 5,514,548 to Krebber et al.; see also International Publication No. WO 93 / 06213 by Hoogenboom et al., published April 1, 1993).

[0054] In some embodiments, the humanized anti-C1s antibodies described herein are full-length IgG, Ig monomer, Fab fragment, F(ab’)2 fragment, Fd fragment, scFv, scAb, or Fv. In some embodiments, the humanized anti-C1s antibodies described herein are full-length IgG. In some embodiments, the heavy chain of any of the humanized anti-C1s antibodies described herein includes a heavy chain constant region (CH) or a portion thereof (e.g., CH1, CH2, CH3, or combinations thereof). The heavy chain constant region can be from any suitable origin, e.g., human, mouse, rat, or rabbit. In some embodiments, the heavy chain constant region is derived from human IgG (gamma heavy chain), e.g., IgG1, IgG2, or IgG4.

[0055] In some embodiments, one or more mutations (e.g., amino acid substitutions) can be introduced into any one of the heavy chain constant regions of the humanized anti-C1s antibodies described herein. In some embodiments, one, two, or more mutations are introduced into the heavy chain constant region (e.g., CH2 domain (residue numbers 231-340 of human IgG1) and / or CH3 domain (residue numbers 341-447 of human IgG1) and / or hinge region (numbering follows the Kabat numbering system (e.g., Kabat's EU index))) to increase or decrease the affinity of the antibody for Fc receptors on effector cells (e.g., activated Fc receptors). Techniques for mutating the Fc region of an antibody to decrease or increase its affinity for an Fc receptor and for introducing such mutations into an Fc receptor or a fragment thereof are known to those skilled in the art. Examples of mutations of the Fc receptor of an antibody that can alter its affinity for the Fc receptor are described, for example, in Smith P et al., (2012) PNAS 109:6181-6186, U.S. Patent No. 6,737,056, and International Publication Pamphlets International Publication No. 02 / 060919; International Publication No. 98 / 23289; and International Publication No. 97 / 34631, which are hereby incorporated by reference.

[0056] In some embodiments, one, two, or more mutations (e.g., amino acid substitutions) are introduced into the hinge region of the heavy chain constant region (CH1 domain) such that the number of cysteine residues in the hinge region is altered (e.g., increased or decreased), as described, for example, in U.S. Patent No. 5,677,425. The number of cysteine residues in the hinge region of the CH1 domain can be altered, for example, to facilitate the association of the light and heavy chains, or to alter (e.g., increase or decrease) the stability of the antibody, or to facilitate the attachment of a linker.

[0057] In some embodiments, one, two, or more mutations (i.e., substitutions, insertions, or deletions) are introduced into the IgG constant domain or an FcRn-binding fragment thereof to alter (e.g., shorten or extend) the in vivo half-life of the antibody. In some embodiments, the one or more mutations are introduced into the Fc or hinge-Fc domain fragment. For examples of mutations that would alter (e.g., shorten or extend) the in vivo half-life of an antibody, see, for example, International Publication No. WO 02 / 060919; International Publication No. WO 98 / 23289; and International Publication No. WO 97 / 34631; as well as U.S. Patent No. 5,869,046; U.S. Patent No. 6,121,022; U.S. Patent No. 6,277,375; and U.S. Patent No. 6,165,745.

[0058] In some embodiments, the constant region of the antibodies described herein is the constant region of IgG1, numbered according to the EU index by Kabat, and includes the substitution of tyrosine (Y) for methionine (M) at position 252, the substitution of threonine (T) for serine (S) at position 254, and the substitution of glutamic acid (E) for threonine (T) at position 256. See U.S. Patent No. 7,658,921, which is incorporated herein by reference. This type of IgG mutant, designated the "YTE mutant," has been shown to have a four-fold longer half-life compared to the wild-type form of the same antibody (see Dall’Acqua W F et al., (2006) J Biol Chem 281:23514-24). In some embodiments, the antibody includes an IgG constant domain that includes one, two, three, or more amino acid substitutions at amino acid residues 251-257, 285-290, 308-314, 385-389, and 428-436, numbered according to the EU index by Kabat. Additional mutations that can be introduced into the heavy chain constant region to extend the half-life of the antibody, such as the M428L / N434S (EU numbering; M459L / N466S in Kabat numbering) mutation described in Zalevsky et al., Nat Biotechnol. 2010 Feb;28(2):157-159, are known in the art.

[0059] In some embodiments, one, two, or more amino acid substitutions are introduced into the Fc region of the constant domain of an IgG to alter the effector function of the antibody. Effector binding sites that vary in affinity can be, for example, Fc receptors or the C1 component of complement. This approach is described in further detail in U.S. Patent No. 5,624,821 and U.S. Patent No. 5,648,260. In some embodiments, deletion or inactivation (via point mutation or other means) of a domain of the constant region can reduce binding of the circulating antibody to the Fc receptor, thereby enhancing localization to the tumor. See, for example, U.S. Patent No. 5,585,097 and U.S. Patent No. 8,591,886 for descriptions of mutations that delete or inactivate the constant domain and thereby enhance localization to the tumor. In some embodiments, at least one amino acid substitution is introduced into the Fc region of the antibodies described herein to remove potential glycosylation sites on the Fc region, thereby reducing binding to the Fc receptor (see, for example, Shields R L et al., (2001) J Biol Chem 276:6591-604).

[0060] In some embodiments, at least one amino acid of the constant region can be replaced with a different amino acid residue such that the C1q binding property of the antibody is altered and / or the complement-dependent cytotoxicity (CDC) activity is reduced or eliminated. This approach is described in further detail in U.S. Patent No. 6,194,551 (Idusogie et al.). In some embodiments, at least one amino acid residue in the N-terminal region of the CH2 domain of the antibodies described herein is altered, thereby altering the ability of the antibody to fix complement. This approach is described in further detail in International Publication No. WO 94 / 29351. In some embodiments, the Fc region of the antibodies described herein is modified to enhance the ability of the antibody to mediate antibody-dependent cell cytotoxicity (ADCC) activity and / or to increase the affinity of the antibody for the Fcγ receptor. This approach is described in further detail in International Publication No. WO 00 / 42072.

[0061] In some embodiments, to avoid potential problems resulting from Fab arm exchange, which is known to occur with native IgG4 mAbs, the antibodies provided herein are stabilized by converting serine 228 (EU numbering; residue 241 in Kabat numbering) to proline, resulting in an IgG1-like hinge sequence, and can include the “Adair” mutation (Angal S., et al., “A single amino acid substitution abolishes the heterogeneity of chimeric mouse / human (IgG4) antibody,” Mol Immunol 30, 105-108; 1993). In some embodiments, to reduce antibody-dependent cellular cytotoxicity activity, an L235E (corresponding to L248E in EU numbering and Kabat numbering) mutation is introduced into the heavy chain constant region, as described, for example, in Benhnia et al., J. Virology, Dec. 2009, p. 12355-12367.

[0062] In some embodiments, the humanized anti-C1s antibody includes a light chain complementarity determining region 1 (LC CDR1) having the amino acid sequence of KASQSVDYDGDSYMN (SEQ ID NO: 1). In some embodiments, the humanized anti-C1s antibody includes a light chain complementarity determining region 2 (LC CDR2) having the amino acid sequence of DASNLES (SEQ ID NO: 2). In some embodiments, the humanized anti-C1s antibody includes a light chain complementarity determining region 3 (LC CDR3) having the amino acid sequence of QQSNEDPWT (SEQ ID NO: 3). In some embodiments, the humanized anti-C1s antibody includes an LC CDR1 having the amino acid sequence of SEQ ID NO: 1, an LC CDR2 having the amino acid sequence of SEQ ID NO: 2, and an LC CDR3 having the amino acid sequence of SEQ ID NO: 3.

[0063] In some embodiments, the humanized anti-C1s antibody comprises a heavy chain complementarity determining region 1 (HC CDR1) comprising the amino acid sequence of DDYIH (SEQ ID NO: 4). In some embodiments, the humanized anti-C1s antibody comprises a heavy chain complementarity determining region 2 (HC CDR2) comprising the amino acid sequence of RIDPADGHTKYAPKFQV (SEQ ID NO: 5). In some embodiments, the humanized anti-C1s antibody comprises a heavy chain complementarity determining region 3 (HC CDR3) comprising the amino acid sequence of YGYGREVFDY (SEQ ID NO: 6). In some embodiments, the humanized anti-C1s antibody comprises an HC CDR1 comprising the amino acid sequence of SEQ ID NO: 4, an HC CDR2 comprising the amino acid sequence of SEQ ID NO: 5, and an HC CDR3 comprising the amino acid sequence of SEQ ID NO: 6.

[0064] In some embodiments, the humanized anti-C1s antibody comprises a light chain variable region (VL) comprising the amino acid sequence of DIVLTQSPDSLAVSLGERATISCKASQSVDYDGDSYMNWYQQKPGQPPKILIYDASNLESGIPARFSGSGSGTDFTLTISSLEPEDFAIYYCQQSNEDPWTFGGGTKVEIK (SEQ ID NO: 7), an LC CDR1 comprising the amino acid sequence of SEQ ID NO: 1, an LC CDR2 comprising the amino acid sequence of SEQ ID NO: 2, an LC CDR3 comprising the amino acid sequence of SEQ ID NO: 3, an HC CDR1 comprising the amino acid sequence of SEQ ID NO: 4, an HC CDR2 comprising the amino acid sequence of SEQ ID NO: 5, and an HC CDR3 comprising the amino acid sequence of SEQ ID NO: 6.

[0065] In some embodiments, the humanized anti-C1s antibody comprises a light chain variable region (VL) comprising the amino acid sequence of DIVLTQSPDSLAVSLGERATISCKASQSVDYDGDSYMNWYQQKPGQPPKILIYDASNLESGIPARFSGSGSGTDFTLTISSLEPEDFAIYYCQQSNEDPWTFGGGTKVEIK (SEQ ID NO: 7). In some embodiments, the humanized anti-C1s antibody comprises an LC CDR1, an LC CDR2, and an LC CDR3 of the VL comprising the amino acid sequence of SEQ ID NO: 7.

[0066] In some embodiments, the humanized anti-C1s antibody is

Chemical Structure

[0067] In some embodiments, the humanized anti-C1s antibody includes a VL containing the amino acid sequence of SEQ ID NO: 7 and a VH containing the amino acid sequence of SEQ ID NO: 8.

[0068] In some embodiments, the humanized anti-C1s antibody includes LC CDR1, LC CDR2, and LC CDR3 of VL containing the amino acid sequence of SEQ ID NO: 7 and HC CDR1, HC CDR2, and HC CDR3 of VH containing the amino acid sequence of SEQ ID NO: 8.

[0069] In some embodiments, the humanized anti-C1s antibody

Chemical formula

[0070] In some embodiments, the humanized anti-C1s antibody

Chemical formula

[0071] In some embodiments, the humanized anti-C1s antibody includes an LC containing the amino acid sequence of SEQ ID NO: 9 and an HC containing the amino acid sequence of SEQ ID NO: 10.

[0072] In some embodiments, the humanized anti-C1s antibody comprises an LC CDR1 comprising an amino acid sequence having three or fewer amino acid variations (e.g., three, two, or one or fewer amino acid variations) relative to the LC CDR1 amino acid sequence of SEQ ID NO: 1. In some embodiments, the humanized anti-C1s antibody comprises an LC CDR2 comprising an amino acid sequence having three or fewer amino acid variations (e.g., three, two, or one or fewer amino acid variations) relative to the LC CDR2 amino acid sequence of SEQ ID NO: 2. In some embodiments, the humanized anti-C1s antibody comprises an LC CDR3 comprising an amino acid sequence having three or fewer amino acid variations (e.g., three, two, or one or fewer amino acid variations) relative to the LC CDR3 amino acid sequence of SEQ ID NO: 3.

[0073] In some embodiments, the humanized anti-C1s antibody comprises an HC CDR1 comprising an amino acid sequence having three or fewer amino acid variations (e.g., three, two, or one or fewer amino acid variations) relative to the HC CDR1 amino acid sequence of SEQ ID NO: 4. In some embodiments, the humanized anti-C1s antibody comprises an HC CDR2 comprising an amino acid sequence having three or fewer amino acid variations (e.g., three, two, or one or fewer amino acid variations) relative to the HC CDR2 amino acid sequence of SEQ ID NO: 5. In some embodiments, the humanized anti-C1s antibody comprises an HC CDR3 comprising an amino acid sequence having three or fewer amino acid variations (e.g., three, two, or one or fewer amino acid variations) relative to the HC CDR3 amino acid sequence of SEQ ID NO: 6.

[0074] In some embodiments, the humanized anti-C1s antibody comprises a VL comprising an amino acid sequence having twenty or fewer amino acid variations (e.g., twenty, nineteen, eighteen, seventeen, sixteen, fifteen, fourteen, thirteen, twelve, eleven, ten, nine, eight, seven, six, five, four, three, two, or one or fewer amino acid variations) relative to the VL amino acid sequence of SEQ ID NO: 7.

[0075] In some embodiments, the humanized anti-C1s antibody comprises a VH comprising an amino acid sequence having 20 or fewer amino acid variations (e.g., 20, 19, 18, 17, 16, 15, 14, 13, 12, 11, 10, 9, 8, 7, 6, 5, 4, 3, 2, or 1 or fewer amino acid variations) relative to the VH amino acid sequence of SEQ ID NO: 8.

[0076] In some embodiments, the humanized anti-C1s antibody comprises a VL comprising an LC CDR1 comprising the amino acid sequence of SEQ ID NO: 1, an LC CDR2 comprising the amino acid sequence of SEQ ID NO: 2, and an LC CDR3 comprising the amino acid sequence of SEQ ID NO: 3, and comprises a framework region having 20 or fewer amino acid variations (e.g., 20, 19, 18, 17, 16, 15, 14, 13, 12, 11, 10, 9, 8, 7, 6, 5, 4, 3, 2, or 1 or fewer amino acid variations) relative to the VL sequence of SEQ ID NO: 7.

[0077] In some embodiments, the humanized anti-C1s antibody comprises a VH comprising an HC CDR1 comprising the amino acid sequence of SEQ ID NO: 4, an HC CDR2 comprising the amino acid sequence of SEQ ID NO: 5, and an HC CDR3 comprising the amino acid sequence of SEQ ID NO: 6, and comprises a framework region having 20 or fewer amino acid variations (e.g., 20, 19, 18, 17, 16, 15, 14, 13, 12, 11, 10, 9, 8, 7, 6, 5, 4, 3, 2, or 1 or fewer amino acid variations) relative to the VH sequence of SEQ ID NO: 8.

[0078] In some embodiments, the humanized anti-C1s antibody comprises a VL comprising an LC CDR1 comprising the amino acid sequence of SEQ ID NO: 1, an LC CDR2 comprising the amino acid sequence of SEQ ID NO: 2, and an LC CDR3 comprising the amino acid sequence of SEQ ID NO: 3, and a framework region comprising 20 or fewer amino acid variations (e.g., 20, 19, 18, 17, 16, 15, 14, 13, 12, 11, 10, 9, 8, 7, 6, 5, 4, 3, 2, or 1 or fewer amino acid variations) relative to the VL sequence of SEQ ID NO: 7, and (b) a VH comprising an HC CDR1 comprising the amino acid sequence of SEQ ID NO: 4, an HC CDR2 comprising the amino acid sequence of SEQ ID NO: 5, and an HC CDR3 comprising the amino acid sequence of SEQ ID NO: 6, and a framework region comprising 20 or fewer amino acid variations (e.g., 20, 19, 18, 17, 16, 15, 14, 13, 12, 11, 10, 9, 8, 7, 6, 5, 4, 3, 2, or 1 or fewer amino acid variations) relative to the VH sequence of SEQ ID NO: 8.

[0079] In some embodiments, the humanized anti-C1s antibody comprises a VL comprising an amino acid sequence having at least 80% (e.g., 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%) identity to the VL amino acid sequence of SEQ ID NO: 7.

[0080] In some embodiments, the humanized anti-C1s antibody comprises a VH comprising an amino acid sequence having at least 80% (e.g., 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%) identity to the VH amino acid sequence of SEQ ID NO: 8.

[0081] In some embodiments, the humanized anti-C1s antibody comprises a VL comprising an LC CDR1 comprising the amino acid sequence of SEQ ID NO: 1, an LC CDR2 comprising the amino acid sequence of SEQ ID NO: 2, and an LC CDR3 comprising the amino acid sequence of SEQ ID NO: 3, and a framework region having at least 80% (e.g., 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%) identity to the framework region of the VL sequence of SEQ ID NO: 7.

[0082] In some embodiments, the humanized anti-C1s antibody comprises a VH comprising an HC CDR1 comprising the amino acid sequence of SEQ ID NO: 4, an HC CDR2 comprising the amino acid sequence of SEQ ID NO: 5, and an HC CDR3 comprising the amino acid sequence of SEQ ID NO: 6, and a framework region having at least 80% (e.g., 80%, 85%, 90%, 95%, 96%, 97%, 98% or 99%) identity to the framework region of the VH sequence of SEQ ID NO: 8.

[0083] In some embodiments, the humanized anti-C1s antibody comprises: (a) a VL comprising an LC CDR1 comprising the amino acid sequence of SEQ ID NO: 1, an LC CDR2 comprising the amino acid sequence of SEQ ID NO: 2, and an LC CDR3 comprising the amino acid sequence of SEQ ID NO: 3, and a framework region having at least 80% (e.g., 80%, 85%, 90%, 95%, 96%, 97%, 98% or 99%) identity to the framework region of the VL sequence of SEQ ID NO: 7; and (b) a VH comprising an HC CDR1 comprising the amino acid sequence of SEQ ID NO: 4, an HC CDR2 comprising the amino acid sequence of SEQ ID NO: 5, and an HC CDR3 comprising the amino acid sequence of SEQ ID NO: 6, and a framework region having at least 80% (e.g., 80%, 85%, 90%, 95%, 96%, 97%, 98% or 99%) identity to the framework region of the VH sequence of SEQ ID NO: 8.

[0084] In some embodiments, the heavy chain constant region in any one of the humanized anti-C1s antibodies described herein is an IgG4 constant region or a variant thereof. Examples of IgG4 constant regions and variants are shown in Table 1.

[0085] [Table 1]

[0086] In some embodiments, any one of the light chains of the humanized anti-C1s antibodies described herein can further comprise a light chain constant region (C L ). In some examples, C L is a kappa light chain. In other examples, C Lis a lambda light chain. In some embodiments, C L is a kappa light chain, and its sequence is as follows: RTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC (SEQ ID NO: 14).

[0087] Other antibody heavy and light chain constant regions are well known in the art and are provided, for example, in the IMGT database (imgt.org) or vbase2.org / vbstat.php, both of which are incorporated herein by reference.

[0088] Compositions The present disclosure provides compositions such as pharmaceutical compositions or formulations comprising a humanized anti-C1s antibody. The humanized anti-C1s antibody of the present disclosure can be formulated into a pharmaceutical composition by combining it with a suitable pharmaceutically acceptable carrier, pharmaceutically acceptable diluent, or other pharmaceutically acceptable pharmaceutical additive.

[0089] Exemplary antibody concentrations in the compositions of the present disclosure can range from about 50 mg / mL to about 250 mg / mL, from about 75 mg / mL to about 225 mg / mL, from about 100 mg / mL to about 200 mg / mL, from about 100 mg / mL to about 150 mg / mL, from about 150 mg / mL to about 200 mg / mL, from about 125 mg / mL to about 175 mg / mL, or from about 140 mg / mL to about 160 mg / mL. In some embodiments, the antibody concentration in the compositions of the present disclosure is about 50 mg / mL, about 75 mg / mL, about 100 mg / mL, about 125 mg / mL, about 130 mg / mL, about 140 mg / mL, about 150 mg / mL, about 160 mg / mL, about 170 mg / mL, about 175 mg / mL, about 200 mg / mL, or about 250 mg / mL. In some embodiments, the compositions of the present disclosure contain about 150 mg / mL of the humanized anti-C1s antibody.

[0090] In some embodiments, the compositions of the present disclosure contain arginine or a salt thereof. Arginine can function as a stabilizer. As described herein, the presence of arginine reduces the isomerization rate of the aspartic acid residue located at position 32 of the light chain variable domain sequence of the target antibody (D32 isomerization rate). Exemplary concentrations of arginine or a salt thereof in the composition can range from about 50 mM to about 200 mM, from about 75 mM to about 175 mM, from about 100 mM to about 150 mM, from about 125 mM to about 175 mM, or from about 125 mM to about 150 mM. In some embodiments, the compositions of the present disclosure contain arginine or a salt thereof at about 50 mM, about 55 mM, about 60 mM, about 65 mM, about 70 mM, about 75 mM, about 80 mM, about 85 mM, about 90 mM, about 95 mM, about 100 mM, about 105 mM, about 110 mM, about 115 mM, about 120 mM, about 125 mM, about 130 mM, about 135 mM, about 140 mM, about 145 mM, about 150 mM, about 155 mM, about 160 mM, about 165 mM, about 170 mM, about 175 mM, about 180 mM, about 185 mM, about 190 mM, or about 200 mM. Exemplary salts of arginine include arginine citrate, arginine hydrochloride, arginine oxalate, arginine phosphate, arginine succinate, or arginine tartrate. Other arginine salts can also be used. In some embodiments, the compositions of the present disclosure contain arginine hydrochloride. In some embodiments, the compositions of the present disclosure contain arginine hydrochloride at about 50 mM to about 200 mM, about 75 mM to about 175 mM, about 100 mM to about 150 mM, about 125 mM to about 175 mM, or about 125 mM to about 150 mM. In some embodiments, the compositions of the present disclosure contain arginine hydrochloride at about 50 mM, about 55 mM, about 60 mM, about 65 mM, about 70 mM, about 75 mM, about 80 mM, about 85 mM, about 90 mM, about 95 mM, about 100 mM, about 105 mM, about 110 mM, about 115 mM, about 120 mM, about 125 mM, about 130 mM, about 135 mM, about 140 mM, about 145 mM, about 150 mM, about 155 mM, about 160 mM, about 165 mM, about 170 mM, about 175 mM, about 180 mM, about 185 mM, about 190 mM, or about 200 mM. In some embodiments, the compositions of the present disclosure contain arginine hydrochloride at about 150 mM.In some embodiments, the arginine hydrochloride is L-arginine hydrochloride.

[0091] In some embodiments, the compositions of the present disclosure include a buffer. Exemplary concentrations of the buffer in the composition can range from about 1 mM to about 50 mM, from about 10 mM to about 40 mM, from about 10 mM to about 30 mM, from about 10 mM to about 25 mM, from about 10 mM to about 25 mM, from about 5 mM to about 25 mM, from about 5 mM to about 20 mM, or from about 5 mM to about 10 mM. In some embodiments, the compositions of the present disclosure include about 1 mM, about 2 mM, about 3 mM, about 4 mM, about 5 mM, about 6 mM, about 7 mM, about 8 mM, about 9 mM, about 10 mM, about 11 mM, about 12 mM, about 13 mM, about 14 mM, about 15 mM, about 16 mM, about 17 mM, about 18 mM, about 19 mM, or about 20 mM of the buffer. Exemplary buffers include acetate, citrate, histidine, oxalate, phosphate, succinate, and tartrate. Other buffers can also be used. In some embodiments, the buffer is histidine. In some embodiments, the compositions of the present disclosure include about 1 mM to about 50 mM of histidine, about 10 mM to about 40 mM, about 10 mM to about 30 mM, about 10 mM to about 25 mM, about 10 mM to about 25 mM, about 5 mM to about 25 mM, about 5 mM to about 20 mM, or about 5 mM to about 10 mM. In some embodiments, the compositions of the present disclosure include about 1 mM, about 2 mM, about 3 mM, about 4 mM, about 5 mM, about 6 mM, about 7 mM, about 8 mM, about 9 mM, about 10 mM, about 11 mM, about 12 mM, about 13 mM, about 14 mM, about 15 mM, about 16 mM, about 17 mM, about 18 mM, about 19 mM, or about 20 mM of histidine. In some embodiments, the compositions of the present disclosure include about 10 mM of histidine. The histidine buffer can include histidine and histidine salts. The concentration of histidine refers to the concentration of histidine and any optional histidine salts present. The contribution to the concentration of each component depends on the target pH. In some embodiments, the histidine buffer includes histidine and histidine hydrochloride. In some embodiments, the concentration of histidine refers to the concentration of both histidine and histidine hydrochloride.

[0092] In some embodiments, the compositions of the present disclosure also include a stabilizer in addition to arginine. Exemplary concentrations of the stabilizer in the composition can range from about 1% to about 8% (w / v), from about 1% to about 5% (w / v), from about 1% to about 3% (w / v), or from about 3% to about 5% (w / v). In some embodiments, the compositions of the present disclosure include the stabilizer at about 1% (w / v), about 2% (w / v), about 3% (w / v), about 4% (w / v), about 5% (w / v), about 6% (w / v), about 7% (w / v), or about 8% (w / v). In some embodiments, the stabilizer is a sugar, a sugar alcohol, or an amino sugar. Exemplary sugars, sugar alcohols, and amino sugars include fructose, galactose, glucose, lactose, maltose, mannose, raffinose, sorbitol, sorbose, sucrose, galactosamine, glucosamine, N-methylglucosamine, and neuraminic acid. Other stabilizers can also be used. In some embodiments, the compositions of the present disclosure include sucrose, trehalose, or sorbitol. In some embodiments, the compositions of the present disclosure include sucrose. In some embodiments, the compositions of the present disclosure include sucrose at about 1% to about 8% (w / v), about 1% to about 5% (w / v), about 1% to about 3% (w / v), or about 3% to about 5% (w / v). In some embodiments, the compositions of the present disclosure include sucrose at about 1% (w / v), about 2% (w / v), about 3% (w / v), about 4% (w / v), about 5% (w / v), about 6% (w / v), about 7% (w / v), or about 8% (w / v). In some embodiments, the compositions of the present disclosure include sucrose at about 3% (w / v). Sucrose can also function as a cryoprotectant. In some embodiments, the composition can include another cryoprotectant selected from ethylene glycol, dimethyl sulfoxide (DMSO), glycerol, trehalose, and propylene glycol. Other cryoprotectants can also be used. In some embodiments, the concentration of the cryoprotectant in the composition can range from about 1% to about 10% (w / v), from about 1% to about 5% (w / v), from about 1% to about 3% (w / v), or from about 3% to about 5% (w / v).In some embodiments, the compositions of the present disclosure comprise a cryoprotectant at about 1% (w / v), about 2% (w / v), about 3% (w / v), about 4% (w / v), about 5% (w / v), about 6% (w / v), about 7% (w / v), about 8% (w / v), about 9% (w / v), or about 10% (w / v).

[0093] In some embodiments, the compositions of the present disclosure include a chelating agent. Exemplary concentrations of the chelating agent in the composition can range from about 1 μM to about 50 μM, from about 5 μM to about 40 μM, from about 5 μM to about 30 μM, from about 5 μM to about 25 μM, from about 5 μM to about 20 μM, from about 5 μM to about 15 μM, from about 10 μM to about 20 μM, from about 10 μM to about 25 μM, or from about 10 μM to 30 μM. In some embodiments, the compositions of the present disclosure include the chelating agent at a concentration of about 1 μM, about 2 μM, about 3 μM, about 4 μM, about 5 μM, about 6 μM, about 7 μM, about 8 μM, about 9 μM, about 10 μM, about 11 μM, about 12 μM, about 13 μM, about 14 μM, about 15 μM, about 16 μM, about 17 μM, about 18 μM, about 19 μM, or about 20 μM. Exemplary chelating agents include ethylenediaminetetraacetic acid (EDTA), diethylenetriaminepentaacetic acid (DTPA), dihydroxyethylglycine, citric acid, tartaric acid, and methionine. Other chelating agents can also be used. In some embodiments, the chelating agent is methionine. In some embodiments, the compositions of the present disclosure include methionine at a concentration of about 5 mM to about 10 mM, about 5 mM to about 7.5 mM, about 7.5 mM to about 10 mM, about 6 mM to about 8 mM, or about 8 mM to about 10 mM. In some embodiments, the compositions of the present disclosure include methionine at a concentration of about 5 mM, about 5.5 mM, about 6 mM, about 6.5 mM, about 7 mM, about 7.5 mM, about 8 mM, about 8.5 mM, about 9 mM, about 9.5 mM, or about 10 mM. One of ordinary skill in the art will recognize that the concentration of the chelating agent can vary depending on the chelating agent used. In some embodiments, the chelating agent is DTPA. In some embodiments, the compositions of the present disclosure include from about 1 μM to about 50 μM of DTPA, from about 5 μM to about 40 μM, from about 5 μM to about 30 μM, from about 5 μM to about 25 μM, from about 5 μM to about 20 μM, from about 5 μM to about 15 μM, from about 10 μM to about 20 μM, from about 10 μM to about 25 μM, or from about 10 μM to 30 μM. In some embodiments, the compositions of the present disclosure include DTPA at a concentration of about 1 μM, about 2 μM, about 3 μM, about 4 μM, about 5 μM, about 6 μM, about 7 μM, about 8 μM, about 9 μM, about 10 μM, about 11 μM, about 12 μM, about 13 μM, about 14 μM, about 15 μM, about 16 μM, about 17 μM, about 18 μM, about 19 μM, or about 20 μM. In some embodiments, the compositions of the present disclosure include about 10 μM of DTPA.

[0094] In some embodiments, the compositions of the present disclosure include a surfactant. Exemplary concentrations of the surfactant in the composition are from about 0.01% to about 0.1% (w / v), from about 0.03% to about 0.6% (w / v), from about 0.03% to about 0.08% (w / v), from about 0.03% to about 0.1% (w / v), from about 0.05% to about 0.1% (w / v), or from about 0.06% to about 0.1% (w / v). In some embodiments, the compositions of the present disclosure include the surfactant at a concentration of about 0.01% (w / v), about 0.02% (w / v), about 0.03% (w / v), about 0.04% (w / v), about 0.05% (w / v), about 0.06% (w / v), about 0.07% (w / v), about 0.08% (w / v), about 0.09% (w / v), or about 0.1% (w / v). Exemplary surfactants include polysorbates (e.g., polysorbate 20 (PS20), polysorbate 40 (PS40), polysorbate 60 (PS60), and polysorbate 80 (PS80)), dicarboxylic acids, oxalic acid, succinic acid, fumaric acid, phthalic acid, polyoxyethylene sorbitan monooleate, poloxamers, poloxamers (e.g., P188), and polyethylene glycol. Other surfactants can also be used. In some embodiments, the surfactant is P188. In some embodiments, the surfactant is PS80. In some embodiments, the compositions of the present disclosure include PS80 from about 0.01% to about 0.1% (w / v), from about 0.03% to about 0.6% (w / v), from about 0.03% to about 0.08% (w / v), from about 0.03% to about 0.1% (w / v), from about 0.05% to about 0.1% (w / v), or from about 0.06% to about 0.1% (w / v). In some embodiments, the compositions of the present disclosure include PS80 at a concentration of about 0.01% (w / v), about 0.02% (w / v), about 0.03% (w / v), about 0.04% (w / v), about 0.05% (w / v), about 0.06% (w / v), about 0.07% (w / v), about 0.08% (w / v), about 0.09% (w / v), or about 0.1% (w / v). In some embodiments, the compositions of the present disclosure include PS80 at a concentration of about 0.06% (w / v).

[0095] The compositions of the present disclosure can have a pH of from about 6 to about 7.5, from about 6 to about 7, from about 6.5 to about 7.5, or from about 6.5 to about 7.1. In some embodiments, the pH of the compositions of the present disclosure is about 6.0, about 6.1, about 6.2, about 6.3, about 6.4, about 6.5, about 6.6, about 6.7, about 6.8, about 6.9, about 7.0, about 7.1, about 7.2, about 7.3, about 7.4, or about 7.5. In some embodiments, the pH of the compositions of the present disclosure is from about 6.5 to 7.1. In some embodiments, the pH of the compositions of the present disclosure is about 6.8. From the tests described herein, it has been shown that adjusting the pH of the compositions containing arginine is beneficial for reducing the D32 isomerization of the target antibody. The pH of the composition can be measured by any means known to those skilled in the art. The means for measuring pH is to use a pH meter equipped with a microelectrode. The pH of the composition can be adjusted by any means known in the art (e.g., by adding an acid, a base, or a buffer or by adjusting its concentration).

[0096] In some embodiments, the present disclosure provides a composition comprising a humanized anti-C1s antibody, arginine or a salt thereof, and one or more of a buffer, a stabilizer or a cryoprotectant, a chelating agent, and a surfactant.

[0097] In some embodiments, the present disclosure provides a composition comprising (a) a humanized anti-C1s antibody at about 50 mg / mL to about 250 mg / mL, (b) arginine HCl at about 50 mM to about 200 mM, (c) histidine at about 1 mM to about 50 mM, (d) sucrose at about 1% to about 10% (w / v), (e) DTPA at about 1 μM to about 50 μM, and (f) polysorbate 80 (PS80) at about 0.01% to about 0.1% (w / v), wherein the pH is from about 6 to about 7. In some embodiments, the arginine hydrochloride is L-arginine hydrochloride.

[0098] In some embodiments, the present disclosure provides a composition comprising (a) a humanized anti-C1s antibody at about 50 mg / mL to about 250 mg / mL, (b) arginine citrate at about 50 mM to about 200 mM, (c) histidine at about 1 mM to about 50 mM, (d) sucrose at about 1% to about 10% (w / v), (e) DTPA at about 1 μM to about 50 μM, and (f) polysorbate 80 (PS80) at about 0.01% to about 0.1% (w / v), wherein the pH is about 6 to about 7.

[0099] In some embodiments, the present disclosure provides a composition comprising (a) a humanized anti-C1s antibody at about 50 mg / mL to about 250 mg / mL, (b) arginine oxalate at about 50 mM to about 200 mM, (c) histidine at about 1 mM to about 50 mM, (d) sucrose at about 1% to about 10% (w / v), (e) DTPA at about 1 μM to about 50 μM, and (f) polysorbate 80 (PS80) at about 0.01% to about 0.1% (w / v), wherein the pH is about 6 to about 7.

[0100] In some embodiments, the present disclosure provides a composition comprising (a) a humanized anti-C1s antibody at about 50 mg / mL to about 250 mg / mL, (b) arginine phosphate at about 50 mM to about 200 mM, (c) histidine at about 1 mM to about 50 mM, (d) sucrose at about 1% to about 10% (w / v), (e) DTPA at about 1 μM to about 50 μM, and (f) polysorbate 80 (PS80) at about 0.01% to about 0.1% (w / v), wherein the pH is about 6 to about 7.

[0101] In some embodiments, the present disclosure provides a composition comprising (a) a humanized anti-C1s antibody at about 50 mg / mL to about 250 mg / mL, (b) arginine succinate at about 50 mM to about 200 mM, (c) histidine at about 1 mM to about 50 mM, (d) sucrose at about 1% to about 10% (w / v), (e) DTPA at about 1 μM to about 50 μM, and (f) polysorbate 80 (PS80) at about 0.01% to about 0.1% (w / v), wherein the pH is about 6 to about 7.

[0102] In some embodiments, the present disclosure provides a composition comprising (a) a humanized anti-C1s antibody at about 50 mg / mL to about 250 mg / mL, (b) arginine tartrate at about 50 mM to about 200 mM, (c) histidine at about 1 mM to about 50 mM, (d) sucrose at about 1% to about 10% (w / v), (e) DTPA at about 1 μM to about 50 μM, and (f) polysorbate 80 (PS80) at about 0.01% to about 0.1% (w / v), wherein the pH is about 6 to about 7.

[0103] In some embodiments, the present disclosure provides a composition comprising (a) a humanized anti-C1s antibody at about 50 mg / mL to about 250 mg / mL, (b) arginine hydrochloride at about 50 mM to about 200 mM, (c) acetate at about 1 mM to about 50 mM, (d) sucrose at about 1% to about 10% (w / v), (e) DTPA at about 1 μM to about 50 μM, and (f) polysorbate 80 (PS80) at about 0.01% to about 0.1% (w / v), wherein the pH is about 6 to about 7. In some embodiments, the arginine hydrochloride is L-arginine hydrochloride.

[0104] In some embodiments, the present disclosure provides a composition comprising (a) a humanized anti-C1s antibody at about 50 mg / mL to about 250 mg / mL, (b) arginine hydrochloride at about 50 mM to about 200 mM, (c) citrate at about 1 mM to about 50 mM, (d) sucrose at about 1% to about 10% (w / v), (e) DTPA at about 1 μM to about 50 μM, and (f) polysorbate 80 (PS80) at about 0.01% to about 0.1% (w / v), wherein the pH is about 6 to about 7. In some embodiments, the arginine hydrochloride is L-arginine hydrochloride.

[0105] In some embodiments, the present disclosure provides a composition comprising (a) a humanized anti-C1s antibody at about 50 mg / mL to about 250 mg / mL, (b) arginine hydrochloride at about 50 mM to about 200 mM, (c) oxalate at about 1 mM to about 50 mM, (d) sucrose at about 1% to about 10% (w / v), (e) DTPA at about 1 μM to about 50 μM, and (f) polysorbate 80 (PS80) at about 0.01% to about 0.1% (w / v), wherein the pH is about 6 to about 7. In some embodiments, the arginine hydrochloride is L-arginine hydrochloride.

[0106] In some embodiments, the present disclosure provides a composition comprising (a) a humanized anti-C1s antibody at about 50 mg / mL to about 250 mg / mL, (b) arginine hydrochloride at about 50 mM to about 200 mM, (c) phosphate at about 1 mM to about 50 mM, (d) sucrose at about 1% to about 10% (w / v), (e) DTPA at about 1 μM to about 50 μM, and (f) polysorbate 80 (PS80) at about 0.01% to about 0.1% (w / v), wherein the pH is about 6 to about 7. In some embodiments, the arginine hydrochloride is L-arginine hydrochloride.

[0107] In some embodiments, the present disclosure provides a composition comprising (a) a humanized anti-C1s antibody at about 50 mg / mL to about 250 mg / mL, (b) arginine hydrochloride at about 50 mM to about 200 mM, (c) succinate at about 1 mM to about 50 mM, (d) sucrose at about 1% to about 10% (w / v), (e) DTPA at about 1 μM to about 50 μM, and (f) polysorbate 80 (PS80) at about 0.01% to about 0.1% (w / v), wherein the pH is about 6 to about 7. In some embodiments, the arginine hydrochloride is L-arginine hydrochloride.

[0108] In some embodiments, the present disclosure provides a composition comprising (a) a humanized anti-C1s antibody at about 50 mg / mL to about 250 mg / mL, (b) arginine hydrochloride at about 50 mM to about 200 mM, (c) tartrate at about 1 mM to about 50 mM, (d) sucrose at about 1% to about 10% (w / v), (e) DTPA at about 1 μM to about 50 μM, and (f) polysorbate 80 (PS80) at about 0.01% to about 0.1% (w / v), wherein the pH is about 6 to about 7. In some embodiments, the arginine hydrochloride is L-arginine hydrochloride.

[0109] In some embodiments, the present disclosure provides a composition comprising (a) a humanized anti-C1s antibody at about 50 mg / mL to about 250 mg / mL, (b) arginine hydrochloride at about 50 mM to about 200 mM, (c) histidine at about 1 mM to about 50 mM, (d) sorbitol at about 1% to about 10% (w / v), (e) DTPA at about 1 μM to about 50 μM, and (f) polysorbate 80 (PS80) at about 0.01% to about 0.1% (w / v), wherein the pH is about 6 to about 7. In some embodiments, the arginine hydrochloride is L-arginine hydrochloride.

[0110] In some embodiments, the present disclosure provides a composition comprising (a) a humanized anti-C1s antibody at about 50 mg / mL to about 250 mg / mL, (b) arginine hydrochloride at about 50 mM to about 200 mM, (c) histidine at about 1 mM to about 50 mM, (d) trehalose at about 1% to about 10% (w / v), (e) DTPA at about 1 μM to about 50 μM, and (f) polysorbate 80 (PS80) at about 0.01% to about 0.1% (w / v), wherein the pH is about 6 to about 7. In some embodiments, the arginine hydrochloride is L-arginine hydrochloride.

[0111] In some embodiments, the present disclosure provides a composition comprising (a) a humanized anti-C1s antibody at about 50 mg / mL to about 250 mg / mL, (b) arginine hydrochloride at about 50 mM to about 200 mM, (c) histidine at about 1 mM to about 50 mM, (d) sucrose at about 1% to about 10% (w / v), (e) EDTA at about 1 μM to about 50 μM, and (f) polysorbate 80 (PS80) at about 0.01% to about 0.1% (w / v), wherein the pH is about 6 to about 7. In some embodiments, the arginine hydrochloride is L-arginine hydrochloride.

[0112] In some embodiments, the present disclosure provides a composition comprising (a) a humanized anti-C1s antibody at about 50 mg / mL to about 250 mg / mL, (b) arginine hydrochloride at about 50 mM to about 200 mM, (c) histidine at about 1 mM to about 50 mM, (d) sucrose at about 1% to about 10% (w / v), (e) methionine at about 5 mM to about 10 mM, and (f) polysorbate 80 (PS80) at about 0.01% to about 0.1% (w / v), wherein the pH is about 6 to about 7. In some embodiments, the arginine hydrochloride is L-arginine hydrochloride.

[0113] In some embodiments, the present disclosure provides a composition comprising (a) a humanized anti-C1s antibody at about 50 mg / mL to about 250 mg / mL, (b) arginine hydrochloride at about 50 mM to about 200 mM, (c) histidine at about 1 mM to about 50 mM, (d) sucrose at about 1% to about 10% (w / v), (e) DTPA at about 1 μM to about 50 μM, and (f) poloxamer 188 (P188) at about 0.01% to about 0.1% (w / v), wherein the pH is about 6 to about 7. In some embodiments, the arginine hydrochloride is L-arginine hydrochloride.

[0114] In some embodiments, the present disclosure provides a composition comprising (a) an antibody at about 150 mg / mL, (b) arginine HCl at about 150 mM, (c) histidine at about 10 mM, (d) sucrose at about 3% (w / v), (e) DTPA at about 10 μM, and (f) PS80 at about 0.06% (w / v), wherein the pH is about 6.5 to about 7.1 or about 6.8. In some embodiments, the composition comprises (a) an antibody at about 150 mg / mL, (b) arginine HCl at about 150 mM, (c) citrate at about 20 mM, and (f) PS80 at about 0.02% (w / v), and the pH of the composition is about 6.5. In some embodiments, the arginine hydrochloride is L-arginine hydrochloride.

[0115] The compositions of the present disclosure can also include other pharmaceutical additives such as, but not limited to, water for injection, diluents, solubilizing agents, soothing agents, additional buffering agents, inorganic or organic salts, antioxidants, etc. In some embodiments, the compositions of the present disclosure do not include other pharmaceutical additives except those described above. Other pharmaceutically acceptable carriers or pharmaceutical additives such as those described in Remington’s Pharmaceutical Sciences 16th edition, Osol, A. Ed. (1980) can also be included in the formulations as long as they do not adversely affect the desired properties of the formulation. In some embodiments, preservatives can be added. In some embodiments, the composition is substantially free of preservatives. Cryoprotectants or lyoprotectants can be included in the lyophilized formulations.

[0116] The compositions of the present disclosure can be in liquid form, lyophilized form, or liquid form reconstituted from lyophilized form, and the lyophilized formulations need to be reconstituted with a sterile solution before administration. Standard procedures for reconstituting lyophilized compositions involve adding back a certain volume of pure water (typically equal to the volume removed during lyophilization), although aqueous solutions containing antibacterial agents can also be used in the manufacture of pharmaceutical compositions for parenteral administration. See also Chen (1992) Drug Dev Ind Pharm 18, 1311 - 54. In some embodiments, the compositions of the present disclosure are in liquid form. The liquid formulations can be injectable as is or can be diluted prior to injection.

[0117] In some embodiments, the present disclosure provides a stable liquid antibody formulation comprising a humanized anti - C1s antibody, arginine or a salt thereof, and one or more of a buffering agent, a stabilizing agent or cryoprotectant, a chelating agent, and a surfactant.

[0118] In some embodiments, the present disclosure provides a stable liquid antibody formulation comprising: (a) a humanized anti - C1s antibody at about 50 mg / mL to about 250 mg / mL; (b) arginine HCl at about 50 mM to about 200 mM; (c) histidine at about 1 mM to about 50 mM; (d) sucrose at about 1% to about 10% (w / v); (e) DTPA at about 1 μM to about 50 μM; and (f) polysorbate 80 (PS80) at about 0.01% to about 0.1% (w / v), wherein the pH of the composition is about 6 to about 7. In some embodiments, the arginine hydrochloride is L - arginine hydrochloride.

[0119] In some embodiments, the present disclosure provides a stable liquid antibody formulation comprising: (a) an antibody at about 150 mg / mL; (b) arginine HCl at about 150 mM; (c) histidine at about 10 mM; (d) sucrose at about 3% (w / v); (e) DTPA at about 10 μM; and (f) PS80 at about 0.06% (w / v), wherein the pH of the composition is about 6.5 to about 7.1 or about 6.8. In some embodiments, the arginine hydrochloride is L - arginine hydrochloride.

[0120] The contemplated composition is stable at 5°C for at least about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 or 12 months or more. In some embodiments, the composition is stable at 5°C for at least about 12, 18, 24 or 30 months or more. In some embodiments, these are stable at 5°C for at least about 6 months or more. In some embodiments, these are stable at 5°C for at least about 9 months. In some embodiments, these are stable at 5°C for at least about 1 year or more or more than about 2 years.

[0121] The compositions of the present disclosure exhibit a high level of stability. As used herein with respect to a composition, the term "stable" means that the antibody in the composition after storage for a defined length of time retains an acceptable level of structure, and / or function, and / or biological activity. A composition can be stable even if the antibody contained therein does not maintain 100% of its structure, and / or function, and / or biological activity after storage for a defined length of time. Under certain circumstances, if after storage for a defined length of time, the structure, and / or function, and / or biological activity of the antibody is maintained at about 80%, about 85%, about 90%, about 95%, about 96%, about 97%, about 98% or about 99%, it can be considered "stable".

[0122] In some embodiments, stability is measured by determining the percentage of the original antibody remaining in the formulation after storage for a defined length of time at a given temperature. The percentage of the original antibody can be measured, for example, by size exclusion chromatography (e.g., size exclusion high performance liquid chromatography [SE-HPLC]) or by any other method known in the art for assessing the % monomer, % high molecular weight species, and % low molecular weight species. In some embodiments, stability is measured by performing differential scanning calorimetry (DSC) to determine thermal stability. In some embodiments, stability is measured by determining mechanical stability (e.g., by performing controlled agitation). In some embodiments, stability is measured by determining the turbidity of the solution (e.g., by measuring the optical density (OD) at 340 nm to 360 nm). In some embodiments, stability is determined by measuring subvisible particles (e.g., by light obscuration using a particle counter in solution or by microflow imaging (MFI)). In some embodiments, stability is determined by measuring chemical degradation such as isomerization, oxidation, and deamidation using tryptic peptide mapping. In some embodiments, stability is determined by measuring the opalescence of the solution using micro-nephelometry.

[0123] In some embodiments, stability can be indirectly evaluated by measuring the concentration of PS80. In some embodiments, the concentration of PS80 is measured by high performance liquid chromatography with a charged aerosol detector (HPLC-CAD).

[0124] Stability can also be evaluated by measuring the biological activity of the antibody and / or its binding affinity for its target. For example, the formulation of the present invention, after being stored at, for example, 5°C, 25°C, 45°C, etc. for a specified length of time (e.g., 1 to 12 months), if the humanized anti-C1s antibody contained in the composition binds to human C1s with an affinity of at least 50%, 60%, 70%, 80%, 90%, 95% or more than that of the antibody before the storage, it can be regarded as stable.

[0125] The compositions of the present disclosure inhibit antibody isomerization. In some embodiments, the isomerization is the isomerization from aspartic acid to isoaspartic acid. In some embodiments, the isomerization is the isomerization of D32 of the light chain located in CDR1 of the antibody. As long as the function of the antibody is not impaired, a certain degree of isomerization can be tolerated. Therefore, after the composition is stored for a specified length of time, even if the antibody contained therein does not maintain 100% of its structure, and / or function, and / or biological activity, the degree of isomerization can be tolerated. Under certain circumstances, after being stored for a specified length of time, if the structure, and / or function, and / or biological activity of the antibody is maintained at about 80%, about 85%, about 90%, about 95%, about 96%, about 97%, about 98% or about 99%, it can be regarded as an acceptable degree of isomerization. Isomerization can be determined by any suitable method known in the art. In some embodiments, isomerization is determined by performing whole peptide mapping analysis. In some embodiments, isomerization is measured by determining the isomerization rate in the formulation after storage at a given temperature for a specified length of time. In some embodiments, the isomerization rate of the antibody after storage at 5°C for 12 weeks is less than about 10%, less than about 9%, less than about 8% or less than about 7.5%. In some embodiments, the isomerization rate of the antibody after storage at 25°C for 12 weeks is less than about 25%, less than about 24%, less than about 23%, less than about 21% or less than about 20%. In some embodiments, the isomerization rate of the antibody after storage at 40°C for 12 weeks is less than about 40%, less than about 39%, less than about 38%, less than about 37%, less than about 36% or less than about 35%.

[0126] In some embodiments, the isomerization rate of the antibody (e.g., D32 of light chain CDR1) is lower compared to the antibody in the corresponding formulation without arginine or its salt. In some embodiments, the isomerization rate of the antibody is at least about 2% or at least about 3% per week at 40°C. In some embodiments, the isomerization rate of the antibody is at least about 1-3% lower per month at 25°C or at least about 0.4%, 0.5% or 0.6% lower per month at 5°C compared to the antibody in the corresponding formulation without arginine or its salt. In some embodiments, the isomerization rate of the antibody after storage at 25°C for 12 weeks is about 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9% or 10% lower compared to the antibody in the corresponding formulation without arginine or its salt.

[0127] Route of administration The antibodies of the present disclosure can be administered to a subject using any available conventional methods and routes suitable for delivering conventional drugs, including systemic or local routes. Generally, the routes of administration contemplated by the present disclosure include, but are not necessarily limited to, enteral, parenteral or inhalation routes.

[0128] Parenteral routes of administration other than inhalation include, but are not necessarily limited to, local, transdermal, subcutaneous, intramuscular, intraorbital, intracapsular, intraspinal, intrasternal, intrathecal and intravenous routes, i.e., any route of administration other than via the gastrointestinal tract. Parenteral administration can be performed to deliver the subject antibody systemically or locally. When systemic delivery is performed, administration typically includes invasive or systemic absorption of the pharmaceutical formulation by local or mucosal administration.

[0129] The routes of administration can be combined as needed or adjusted according to the desired effect. The composition can be administered as a single dose or multiple doses. In some embodiments, the composition of the present disclosure is administered intravenously. In some embodiments, the composition of the present disclosure is administered subcutaneously.

[0130] Dosage and dosage form Suitable dosages can be determined by the attending physician or other qualified medical personnel based on a variety of clinical factors. As is well known in the medical arts, the dosage for any one patient will vary depending on many factors, such as the patient's size, body surface area, age, the specific compound being administered, the patient's sex, the time and route of administration, general health, and other drugs being administered concomitantly.

[0131] One of ordinary skill in the art will readily appreciate that dosages and administration schedules can vary depending on the particular antibody, the severity of the symptoms, and the subject's susceptibility to side effects. Preferred dosages and administration schedules for a given compound can be determined by those of ordinary skill in the art using a variety of means.

[0132] The present disclosure also provides a pharmaceutical unit dosage form comprising a therapeutically effective amount of the composition of the present disclosure, for treating one or more complement-mediated diseases in a subject by administering the dosage form to the subject. In some embodiments, the subject is a human. The term "pharmaceutical unit dosage form" refers to a physically discrete unit suitable as a unitary dosage for the subject to be treated, each unit containing a predetermined amount of active compound calculated to obtain a desired therapeutic / prophylactic effect.

[0133] The unit dosage form can be a container that contains the formulation. Suitable containers include, but are not limited to, sealed ampoules, vials (e.g., glass vials), bottles, syringes and test tubes. The container can be made of various materials such as glass or plastic, and can have a sterile connection port (e.g., the container can be a vial with a stopper that can be punctured by a hypodermic needle). In some embodiments, the container is a vial. In some embodiments, the container is a pre-filled syringe. Generally, the container needs to maintain the sterility and stability of the formulation.

[0134] In some embodiments, the composition is packaged in a Type I glass vial (10R), closed with a stopper (chlorobutyl coated with flurotec), and sealed with a flip-off cap with a flange (aluminum). In some embodiments, the vial is filled with 8 mL of the composition.

[0135] In some embodiments, the composition is packaged in a glass prefilled syringe (OMPI) and closed with a rubber stopper (Novapure). In some embodiments, the prefilled syringe is filled with 2 mL of the composition.

[0136] Exemplary drug delivery devices can include a needle-based injection system as described in Table 1 of Section 5.2 of ISO 11608-1:2014(E). As described in ISO 11608-1:2014(E), needle-based injection systems can be broadly classified into a multi-dose container system and a single-dose (partial or full discharge) container system. The container can be a replaceable container or an integrated non-replaceable container.

[0137] As further described in ISO 11608-1:2014(E), the multi-dose container system can include an infusion device that uses a replaceable container with a needle. In such a system, each container holds a plurality of doses, the amount of which can be fixed or variable (preset by the user). Other multi-dose container systems can include an infusion device that uses a needle integrated with a non-replaceable container. In such a system, each container holds a plurality of doses, the amount of which can be fixed or variable (preset by the user).

[0138] As further described in ISO 11608-1:2014(E), a single-dose container system can include an injection device that uses a needle with a replaceable container. In one example of such a system, each container holds a single-dose amount, such that the entire administrable amount is discharged (full discharge). In a further example, each container holds a single-dose amount, such that a portion of the administrable amount is discharged (partial discharge). Also as described in ISO 11608-1:2014(E), a single-dose container system can include an injection device that uses a needle with a non-replaceable container. In one example of such a system, each container holds a single-dose amount, such that the entire administrable amount is discharged (full discharge). In a further example, each container holds a single-dose amount, such that a portion of the administrable amount is discharged (partial discharge).

[0139] An exemplary sleeve-activated autoinjector with manually inserted needle is further described in WO 2015 / 004052 pamphlet. Examples of feedback mechanisms that can provide an audible confirmation of the end of administration are described in WO 2016 / 193346 pamphlet and WO 2016 / 193348 pamphlet. Examples of mechanisms for safely retaining the needle after use of the autoinjector are described in WO 2016 / 193352 pamphlet. Examples of needle cover removal mechanisms for syringe-type autoinjectors are described in WO 2016 / 193353 pamphlet. Examples of support mechanisms for supporting the axial position of the syringe are described in WO 2016 / 193355 pamphlet.

[0140] Kits and Manufactured Articles The present disclosure provides a kit or an article of manufacture comprising the composition of the present disclosure. The kit or article of manufacture comprises a container containing the composition of the present disclosure. The kit or article of manufacture may further comprise one or more containers containing pharmaceutically acceptable pharmaceutical additives, and may include other materials such as filters, needles, and syringes that are desirable from a commercial and user perspective. The kit may be accompanied by a package insert containing information regarding the use of such a therapeutic, prophylactic, or diagnostic product, such as indications, usage, dosage, manufacture, administration, contraindications, and / or warnings, which are typically included in the commercial packaging of the therapeutic, prophylactic, or diagnostic product. The kit can also be associated with a label which can be any kind of data carrier containing information (e.g., leaflet, seal, chip, printing, or barcode). In certain embodiments, the above-described package insert can be included inside or on the label. The kit may further comprise an instrument for administering the formulation and in particular an instrument for containing the composition, i.e., a prefilled device such as, but not limited to, a prefilled syringe or a prefilled autoinjector. The kit may also include a container containing the composition, i.e., a prefilled container such as a prefilled vial, cartridge, bag, or ampule.

[0141] Complement-mediated diseases The composition of the present disclosure is useful for the treatment of complement-mediated diseases. Thus, in one aspect, the present disclosure provides a method for treating a complement-mediated disease. The method generally comprises administering an effective amount of the composition of the present disclosure to a subject in need thereof. Optionally, by administering the composition of the present disclosure, the activity of complement C1s in the cells, tissues, body fluids, or organs of an individual is regulated, and a complement-mediated disease or disorder is treated. The present disclosure provides a method for inhibiting the activation of complement component C4 in an individual, the method comprising administering to the subject an effective amount of the composition of the present disclosure. The present disclosure provides a method for inhibiting complement C1s activity in a subject, the method comprising administering to the subject an effective amount of the composition of the present disclosure. The present disclosure provides a method for reducing the level of complement component cleavage products in a subject (e.g., in the body fluids, tissues, or organs of an individual), the method comprising administering to the subject an effective amount of the composition of the present disclosure.

[0142] Optionally, an "effective amount" of the composition of the present disclosure, when administered to a subject in need thereof in one or more doses, is an amount that reduces the level of complement component cleavage products in the subject (e.g., in the body fluid, tissue, or organ of an individual). Optionally, an "effective amount" of the composition of the present disclosure, when administered to a subject in need thereof in one or more doses, is an amount that reduces the level of complement component cleavage products in the subject (e.g., in the body fluid, tissue, or organ of an individual) by at least 10%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, or 100% compared to the level of complement component cleavage products in a body fluid, tissue, or organ that has not been treated with the composition, e.g., before treatment with the composition. In some embodiments, the complement component cleavage product is a C4 cleavage product (e.g., C4b). In some embodiments, the complement component cleavage product is a C2 cleavage product (e.g., C2a). In some embodiments, the complement component cleavage product is a C3 cleavage product.

[0143] In some cases, an “effective amount” of the composition of the present disclosure is an amount that, when administered to a subject in need thereof in one or more doses, reduces the activity of the classical complement pathway in the subject (e.g., in the body fluid, tissue, or organ of an individual). In some cases, an “effective amount” of the composition of the present disclosure is an amount that, when administered to a subject in need thereof in one or more doses, reduces the activity of the classical complement pathway in the subject (e.g., in the body fluid, tissue, or organ of an individual) within about 48 hours, within about 24 hours, within about 12 hours, within about 8 hours, or within about 4 hours from the administration of the humanized anti-C1s antibody, by at least 10%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, or 100% as compared to the activity of the classical complement pathway in a body fluid, tissue, or organ that has not been treated with the composition, e.g., before treatment with the composition. The activity level of the classical complement pathway can be determined using any of a variety of methods. As a non-limiting example, the activity of the classical complement pathway can be determined ex vivo by determining the activity level of the classical complement pathway in a blood, serum, or plasma sample obtained from an individual, for example. For example, the classical complement pathway in a blood, serum, or plasma sample can be activated ex vivo, and the amount of complement component cleavage products (such as C5b-9) produced by such activation can be determined.

[0144] In some embodiments, the "effective amount" of the compositions of the present disclosure, when administered to a subject in need thereof in one or more doses, is an amount that reduces and maintains the activity level of the classical complement pathway in the subject (e.g., in the body fluid, tissue, or organ of an individual) by at least 10%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, or 100% as compared to the activity level of the classical complement pathway in the body fluid, tissue, or organ that has not been treated with the composition, e.g., before treatment with the composition, and this reduction is maintained for a period of about 4 hours to about 30 days (e.g., 4 hours to 8 hours, 8 hours to 24 hours, 2 days to 4 days, 4 days to 7 days, 7 days to 14 days, 14 days to 21 days, or 21 days to 30 days).

[0145] In some embodiments, the "effective amount" of the compositions of the present disclosure, when administered to a subject in need thereof in one or more doses, is an amount that reduces and maintains the level of complement component cleavage products in the subject (e.g., in the body fluid, tissue, or organ of an individual) by at least 10%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, or 100% as compared to the level of complement component cleavage products in the body fluid, tissue, or organ that has not been treated with the composition, e.g., before treatment with the composition, and this reduction is maintained for a period of about 4 hours to about 30 days (e.g., 4 hours to 8 hours, 8 hours to 24 hours, 2 days to 4 days, 4 days to 7 days, 7 days to 14 days, 14 days to 21 days, or 21 days to 30 days).

[0146] In some cases, the "effective amount" of the compositions of the present disclosure, when administered to a subject in need thereof in one or more doses, is at least 10%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95% or 100% less production of C4b2a (i.e., the complex of complement C4b and C2a; also known as "C3 convertase") in the subject (e.g., in the body fluid, tissue or organ of an individual) compared to the amount of C4b2a produced in a subject not treated with the present composition, e.g., a subject or body fluid, tissue or organ prior to treatment with the present composition.

[0147] The present disclosure provides a method of modulating complement activation. In some embodiments, the method inhibits complement activation, e.g., to reduce the production of C4b2a. In some embodiments, the present disclosure provides a method of modulating complement activation in an individual having a complement-mediated disease, the method comprising administering to the individual a composition of the present disclosure. In some embodiments, such method inhibits complement activation.

[0148] In some embodiments, a complement-mediated disease is characterized by an increased amount (more than normal) of C1s present in cells, tissues or body fluids or an elevated complement C1s activity level. For example, in some embodiments, a complement-mediated disease is characterized by an increased amount of C1s present in brain tissue and / or cerebrospinal fluid and / or an elevated C1s activity. The amount of C1s in cells, tissues or body fluids being "more than normal" means that the amount of C1s in cells, tissues or body fluids is more than the normal control level, for example more than the normal control level of individuals or a population of individuals of the same age group. The C1s activity level in cells, tissues or body fluids being "higher than normal" means that proteolytic cleavage caused by C1s in cells, tissues or body fluids is higher than the normal control level, for example higher than the normal control level of individuals or a population of individuals of the same age group. In some embodiments, an individual having a complement-mediated disease additionally exhibits one or more symptoms of such a disease. The term "disease" should be understood to encompass "disorder". These two terms can be used interchangeably. In some embodiments, the complement-mediated disease is a classical complement-mediated disease.

[0149] In some embodiments, a complement-mediated disease is characterized by an amount of C1s present in cells, tissues or body fluids that is less than the normal amount or a decreased complement C1s activity level. For example, in some embodiments, a complement-mediated disease is characterized by a decreased amount of C1s present in brain tissue and / or cerebrospinal fluid and / or a decreased C1s activity. The amount of C1s in cells, tissues or body fluids being "less than normal" means that the amount of C1s in cells, tissues or body fluids is less than the normal control level, for example less than the normal control level of individuals or a population of individuals of the same age group. The C1s activity level in cells, tissues or body fluids being "lower than normal" means that proteolytic cleavage caused by C1s in cells, tissues or body fluids is lower than the normal control level, for example lower than the normal control level of individuals or a population of individuals of the same age group. In some embodiments, an individual having a complement-mediated disease additionally exhibits one or more symptoms of such a disease.

[0150] Complement-mediated diseases are diseases in which the amount or activity of complement C1s is an amount or activity sufficient to cause the disease in an individual. In some embodiments, the complement-mediated disease is selected from the group consisting of alloimmune diseases, autoimmune diseases, cancer, blood diseases, infectious diseases, inflammatory diseases, ischemia-reperfusion injury, neurodegenerative diseases, neurodegenerative disorders, eye diseases, kidney diseases, transplant rejection, vascular diseases, and vasculitis. In some embodiments, the complement-mediated disease is an autoimmune disease. In some embodiments, the complement-mediated disease is an alloimmune disease. In some embodiments, the complement-mediated disease is cancer. In some embodiments, the complement-mediated disease is an infectious disease. In some embodiments, the complement-mediated disease is an inflammatory disease. In some embodiments, the complement-mediated disease is a blood disease. In some embodiments, the complement-mediated disease is ischemia-reperfusion injury. In some embodiments, the complement-mediated disease is an eye disease. In some embodiments, the complement-mediated disease is a kidney disease. In some embodiments, the complement-mediated disease is transplant rejection. In some embodiments, the complement-mediated disease is antibody-mediated transplant rejection. In some embodiments, the complement-mediated disease is a vascular disease. In some embodiments, the complement-mediated disease is vasculitis. In some embodiments, the complement-mediated disease is a neurodegenerative disease.

[0151] Examples of complement-mediated diseases or disorders include, but are not limited to, age-related macular degeneration, Alzheimer's disease, amyotrophic lateral sclerosis, anaphylaxis, antibody-mediated rejection (AMR), argyrophilic grain dementia, arthritis (e.g., rheumatoid arthritis), asthma, atherosclerosis, atypical hemolytic uremic syndrome, autoimmune diseases (e.g., autoimmune hemolytic anemia (AIHA); warm-type AIHA; mixed-type AIHA, etc.), Barraquer-Simons syndrome, Behçet's disease, British-type amyloid angiopathy, bullous pemphigoid, Burger's disease, C1q nephropathy, cancer, catastrophic antiphospholipid antibody syndrome, cerebral amyloid angiopathy, chronic inflammatory demyelinating polyneuropathy (CIDP), cold agglutinin disease (CAD), corticobasal degeneration, Creutzfeldt-Jakob disease, Crohn's disease, cryoglobulinemic vasculitis, boxer dementia, dementia with Lewy bodies (DLB), diffuse neurofibrillary tangles with calcification, discoid lupus erythematosus, Down syndrome, Evans syndrome, focal segmental glomerulosclerosis, formal thought disorder, frontotemporal dementia (FTD), frontotemporal dementia linked to chromosome 17 with parkinsonism, frontotemporal lobar degeneration, Gerstmann-Straussler-Scheinker disease, Guillain-Barré syndrome, Hallervorden-Spatz syndrome, hemolytic uremic syndrome, hereditary angioedema, hypophosphatasia, idiopathic pulmonary inflammatory syndrome, immune complex diseases, inclusion body myositis, infectious diseases (e.g., diseases caused by bacteria (e.g., meningococcus or streptococcus), viruses (e.g., human immunodeficiency virus (HIV)) or other infectious pathogens), inflammatory diseases, ischemia-reperfusion injury, mild cognitive impairment, immune thrombocytopenic purpura (ITP), molybdenum cofactor deficiency (MoCD) type A, membranoproliferative glomerulonephritis (MPGN) type I, membranoproliferative glomerulonephritis (MPGN) type II (dense deposit disease), membranous nephropathy, multi-infarct dementia, lupus (e.g., systemic lupus erythematosus (SLE)), glomerulonephritis, Kawasaki disease, multifocal motor neuropathy, multiple sclerosis, multiple system atrophy, myasthenia gravis, myocardial infarction, myotonic dystrophy, neuromyelitis optica, Niemann-Pick disease type C, non-Guam type motor neuron disease with neurofibrillary tangles, Parkinson's disease, Parkinson's disease with dementia, paroxysmal nocturnal hemoglobinuria, pemphigus vulgaris, Pick's disease, post-encephalitic parkinsonism, polymyositis,Brain amyloid angiopathy, progressive subcortical gliosis, progressive supranuclear palsy, psoriasis, sepsis, Shiga toxin-producing Escherichia coli (STEC)-associated hemolytic uremic syndrome, spinal muscular atrophy, stroke, subacute sclerosing panencephalitis, neurofibrillary type senile dementia, transplant rejection, vasculitis (e.g., ANCA-associated vasculitis), Wegener's granulomatosis, sickle cell disease, cryoglobulinemia, mixed cryoglobulinemia, essential mixed cryoglobulinemia, type II mixed cryoglobulinemia, type III mixed cryoglobulinemia, nephritis, drug-induced thrombocytopenia, lupus nephritis, acquired epidermolysis bullosa, delayed hemolytic transfusion reaction, hypocomplementemic urticarial vasculitis syndrome, pseudophakic bullous keratopathy, and platelet transfusion refractoriness are included.

[0152] In some embodiments, the complement-mediated disease is CAD. In some embodiments, the complement-mediated disease is ITP. In some embodiments, the complement-mediated disease is CIDP. In some embodiments, the complement-mediated disease is AMR.

[0153] In some embodiments, the compositions of the present disclosure prevent or delay the onset of at least one symptom of a complement-mediated disease in a subject. In some embodiments, the compositions of the present disclosure reduce or eliminate at least one symptom of a complement-mediated disease in a subject. Examples of symptoms include, but are not limited to, symptoms associated with autoimmune diseases, cancer, blood diseases, infectious diseases, inflammatory diseases, ischemia-reperfusion injury, neurodegenerative diseases, neurodegenerative disorders, kidney diseases, transplant rejection, eye diseases, vascular diseases, or vasculitis. The symptoms can be neurological symptoms, such as cognitive dysfunction, memory impairment, decreased motor function, etc. The symptoms can also be the activity of C1s protein in the cells, tissues, or body fluids of an individual. The symptoms can also be the degree of complement activation in the cells, tissues, or body fluids of an individual.

[0154] "Treatment" means at least improving the symptoms associated with the condition afflicting the host, and improvement is used in a broad sense to refer to at least reducing a parameter associated with the condition being treated, such as a complement-mediated disease, for example the severity of the symptoms. In such cases, treatment also includes situations where the condition or at least the symptoms associated with it are completely prevented such that the host no longer suffers from the condition or at least the symptoms characterizing the condition, for example the occurrence is prevented or stopped, for example it ceases.

[0155] In accordance with the methods of the present subject matter, various hosts (the term "host" is used interchangeably herein with the terms "subject", "individual" and "patient") can be treated. Generally, such hosts are "mammals" or "mammalian", and these terms are widely used to represent organisms belonging to the class Mammalia, including carnivores (e.g., cats), herbivores (e.g., cows, horses and sheep), omnivores (e.g., dogs, goats and pigs), rodents (e.g., mice, guinea pigs and rats) and primates (e.g., humans, chimpanzees and monkeys). In some embodiments, the host is an individual having a complement system, such as a mammal, fish or invertebrate. In some embodiments, the host is a companion animal, agricultural animal, working animal, zoo animal or laboratory animal of a mammal, fish or invertebrate that includes a complement system. In some embodiments, the host is a human.

Examples

[0156] Example 1. Design of formulation development tests SAR445088 is a humanized immunoglobulin G (IgG) subclass 4 (IgG4) monoclonal antibody (mAb) produced in Chinese hamster ovary (CHO) cells that binds to and inhibits the activated form of C1s within the classical pathway (CP). The mechanism of SAR445088 is specific to the CP and does not affect the function of the lectin and alternative pathways. The sequences of the LC CDR1, LC CDR2, LC CDR3, HC CDR1, HC CDR2, HC CDR3, VH, VL, and the heavy and light chains of SAR445088 are described elsewhere in this application as SEQ ID NOs: 1-10. This sequence is also described in U.S. Patent No. 9,512,233, U.S. Patent No. 10,729,767, and U.S. Patent Application Publication No. 2020 / 0048332, which are hereby incorporated by reference in their entireties.

[0157] In early-stage development testing, this antibody was shown to have a higher tendency to self-associate (kD -22.7 mL / g in 10 mM histidine, pH 6.0 and -12.4 mL / g in 10 mM histidine, pH 6.0 + 150 mM NaCl) and a tendency for the aspartic acid in the light chain CDR1 to isomerize (LC D32 isomerization). To evaluate the feasibility and responsibilities associated with an optimized histidine-based liquid formulation (DP), formulation development studies were conducted. A series of comprehensive studies were performed to understand the impact of formulation composition and processing conditions on the biophysical and chemical stability of SAR445088. To optimally develop the SAR445088 liquid DP, the effects of buffer type and pH, concentration of polysorbate 80, and metal chelating agents were all critically evaluated. The specific evaluations and their objectives are as follows.

[0158] Management of D32 Isomerization The objectives are to identify incompatibility boundaries to prevent D32 isomerization and to evaluate the impact of pH and stabilizers on D32 isomerization.

[0159] Screening of Polysorbate 80 The aim is to evaluate the optimal concentration of polysorbate 80 (PS80) required to stabilize the SAR445088 liquid formulation when subjected to stress induced by agitation and light exposure. Additionally, when the SAR445088 sample was diluted 50-fold with 0.9% saline to 3 mg / mL, the optimal PS80 concentration in the liquid DP was determined by testing the ability of various concentrations of PS80 to mitigate the increase in turbidity and / or the formation of subvisible particles.

[0160] Metal Contamination and Chelating Agent Comparative Test The aim is to investigate the impact of transition metals that may leach into the drug substance (DS) during manufacturing on the chemical and physical stability of SAR445088 and PS80. The ability of the chelating agents EDTA and DTPA to protect proteins from degradation in a series of stability experiments under worst-case conditions was also evaluated.

[0161] pH / Buffering Agent Screening and DP Stability The aim is to evaluate the influence of buffer type and pH on the physical and chemical stability of the SAR445088 liquid formulation under refrigerated storage, room temperature storage, and accelerated storage conditions. The histidine and phosphate buffer systems were selected for this test. For the proposed target formulation matrix, their ability to stabilize high-concentration DP solutions during frozen storage and liquid storage was tested. The effect of adding 3% sucrose on the stability of the DP was also evaluated.

[0162] DP Robustness Test The aim is to confirm that the final composition of the proposed target formulation has sufficient robustness to withstand stress induced by agitation, freeze-thaw cycles, and light exposure without compromising the stability of the formulation during storage or manufacturing.

[0163] Example 2. Materials and Methods Materials Active pharmaceutical ingredient. The formulated drug substance (FDS) of SAR445088 was prepared at a concentration of >150 mg / mL. Specific lots and batches of the FDS were used for a given test conducted in formulation development. The FDS was formulated at the target protein and pharmaceutical additive concentrations and then the formulation was passed through a 0.22-μm filter in a laminar flow and aseptically filled to prepare a liquid formulation (DP).

[0164] Pharmaceutical additives. All pharmaceutical additives used in the formulation development tests were of grades compliant with ACS, United States Pharmacopeia (USP), and European Pharmacopeia (Eur). The pharmaceutical additives used in this test included L-histidine, L-histidine monohydrochloride, L-arginine monohydrochloride, sucrose, PS80, monosodium phosphate, disodium phosphate heptahydrate, EDTA, and DTPA. The pharmaceutical additives were all obtained from J.T. Baker except for DTPA which was obtained from Sigma Aldrich.

[0165] Analytical methods pH. The pH of the buffer solution and the formulated antibody solution was measured using a pH probe and pH meter from Mettler-Toledo. The results were considered equivalent if the difference between repeated measurements was within 0.1 pH unit.

[0166] Lunatic for determining protein concentration. The total protein concentration was determined by measuring the UV absorbance at 280 nm on a microfluidic chip using a Big Lunatic spectrometer (Unchained Labs). The measurements were performed in duplicate with a sample volume of 2 - 5 μL (ε1%: 14.6 (g / 100 mL)-1cm-1).

[0167] UV plate reader for determining turbidity and optical density. Turbidity of the samples was quantified by measuring the optical density (OD) at 340 nm to 360 nm using a SpectraMax i3 Microplate Reader (Molecular Devices). 200 μL of each sample was loaded into a UV-Vis transparent 96-well plate (Corning). OD was determined as the average of the absorbance values at 340 nm, 345 nm, 350 nm, 355 nm, and 360 nm.

[0168] Size exclusion HPLC for determining high molecular weight (HMW) species. Aggregation analysis was performed by size exclusion chromatography (SEC). On a 1260 series HPLC (Agilent, Santa Clara, CA) equipped with a TSK-GEL G3000SWXL (Tosoh Bioscience, Tokyo, Japan) analytical column and a corresponding guard column, using 20 mM sodium phosphate, 300 mM sodium chloride, with pH 6.5 ± 0.1 and a flow rate of 0.5 mL / min, the samples were separated over 30 minutes. Each sample was injected twice. Detection was performed by UV absorbance at 280 nm, and the peaks of the chromatogram were integrated to determine the relative proportions of each eluted species.

[0169] Size exclusion ultra-high pressure liquid chromatography (UPLC) for determining HMW and low molecular weight (LMW) species. Aggregation analysis was performed by size exclusion chromatography (SEC). On a Waters UPLC (Waters Corporation, Milford, MA) equipped with an Acquity BEH200 UPLC (Waters Corporation, Milford, MA) analytical column, using 50 mM sodium phosphate, 300 mM sodium chloride, with pH 7.0 and a flow rate of 0.3 mL / min, the samples were separated over 20 minutes. Each sample was injected three times. Detection was performed by UV absorbance at 280 nm, and the peaks of the chromatogram were integrated to determine the relative proportions of each eluted species.

[0170] Subvisible particles. The measurement of subvisible particles by light obscuration method was also carried out using a high-precision in-liquid particle counter (HIAC) Model 9703+ (Beckman Coulter). Degassed MilliQ water was passed through a 0.22 μm filter and flowed into the counter until the particle count reached ≤ 1 particle / mL at 10 μm. To confirm the accuracy of the particle count, standard particles of 2 μm, 10 μm, and 25 μm were measured, and then the counter was extensively washed to remove the background signal. Using the procedure for 1 mL, a 0.2 mL sample was injected and analyzed in four portions. The measured value of the first sample was ignored, and the average of the subsequent three measured values was taken.

[0171] cIEF for determining charge variants. The charge heterogeneity of the protein was measured by capillary isoelectric focusing (cIEF) using a Maurice instrument (Protein Simple) and detecting at 280 nm UV absorbance. The sample (1 mL) and the standard solution were first diluted to 15 mg / mL with the formulation buffer and then further diluted to 5 mg / mL with water. On-board mixing was used to mix the sample with the MasterMix before analysis. The isoelectric focusing of the sample was assumed to consist of a pre-focusing time of 3 minutes at 1500 V followed by a focusing time of 11 minutes at 3000 V. Results were considered equivalent if the difference between samples was 10% or less.

[0172] Measurement of PS80 by Charged Aerosol Detection (CAD). PS80 is a nonionic surfactant mainly composed of a hydrophilic sorbitan polyoxyethylene group linked to a fatty acid by an ester group. PS80 was separated from other sample components such as proteins and formulation components by mixed-mode HPLC using an Oasis MAX column (2.1×20 mm, particle size 30 μm, part number 186002052) (Waters) and Charged Aerosol Detection (CAD) (Agilent Technologies, CA). The column resin contains a hydrophobic polymer backbone that supports a positively charged quaternary amine. As a result, this column has both reversed-phase and ion-exchange capabilities. Thereby, it becomes possible to retain hydrophobic compounds such as PS80, and an electrostatic repulsion occurs against the positive charges on the protein / pharmaceutical additive. Samples were eluted using a gradient mobile phase of 2% formic acid in water (A) and 2% formic acid in isopropyl alcohol (B). Each sample was passed through the column for 8 minutes after injection, and the detection time range of CAD was set to 2.5 - 8 minutes.

[0173] Quantification of post-translational modification (PTM) by LC-MS. After diluting the protein sample to 1 mg / mL, reduction, alkylation, and digestion with a trypsin and Lys-C mixture were performed using a fully automated method with a STAR liquid handling system (Hamilton). Peptide mapping analysis was carried out using a Q Exactive HF LC-MS system. The acquired LC-MS and LC-MS / MS data were processed using BioPharma Finder 4.1 for the identification and relative quantification of modifications.

[0174] Pharmacological effect. C1s was directly immobilized on the surface of a Biacore SPR sensor chip. Without changing the buffer solution, a calibration curve for the binding of C1s by the antibody was created using the original sample of the formulated antibody. After diluting 1000-fold with the measurement buffer, all samples were tested. All samples were tested in triplicate using three different dilution methods (100×10, 50×20, and 25×40). The control sample was the original sample diluted to 1 μg / mL. The control samples were run every 6 samples.

[0175] Example 3. Control of D32 isomerization The D32 isomerization in the light chain (LC) CDR1 region of SAR445088 was identified as the main chemical degradation mechanism that directly affects the Biacore binding of the SAR445088 molecule. As can be seen from Figure 1, with the D32 isomerization of CDR1, the Biacore binding was affected in the decreasing direction. When the isomerization rate exceeded 40%, the Biacore binding decreased significantly. According to the reference investigation, the D32 isomerization at the time when the expected end of shelf life (EOS) level was reached was estimated to be around 10 - 15%.

[0176] Next, a systematic test was conducted to evaluate the effect of pH and the presence or absence of stabilizers such as arginine hydrochloride on the isomerization rate of SAR445088. As can be seen from Figures 2A - 2D, the D32 isomerization rate decreased as the pH increased from 6.2 to 6.8 at all test temperatures. Furthermore, when arginine hydrochloride was added as a stabilizer, the decrease in the isomerization rate was further promoted compared to the control without arginine hydrochloride. As expected, the isomerization of the lyophilized formulation was negligible at all test temperatures. Based on these results, increasing the pH (pH 6.8) and containing arginine hydrochloride seemed to be beneficial for further antibody formulation development. Since increasing the pH of the liquid formulation may enable degradation based on deamidation, the SAR445088 formulation preferably has an optimal pH and stabilizer concentration to minimize both isomerization and deamidation.

[0177] Example 4. Validation test for PS80 The optimal amount of PS80 in the SAR445088 formulation was determined by observing the resistance of a test formulation containing 0 - 0.1% PS80 (150 mg / mL SAR445088, 150 mM arginine HCl, 3% sucrose, 10 μM EDTA, 10 mM histidine, pH 7.0) to various stresses described in Table 2. The purpose of this test was to determine the appropriate amount of PS80 to reduce the effects of shear during manufacturing, shipping, and handling using the test conditions described below.

[0178]

Table 2

[0179] Orbital shaking test As can be seen from Figure 3, even in the presence of PS80, orbital shaking had no effect on the formation of high molecular weight (HMW) species of the protein (Figure 3A). Samples without PS80 showed an increase in optical density / turbidity over time due to orbital shaking. On the other hand, samples containing as little as 0.01% PS80 showed much higher stability (Figure 3B). Finally, samples without PS80 showed an increase in sub - visible particles of 2 μm, 10 μm, and 25 μm over time due to orbital shaking. On the other hand, samples containing more than 0.01% PS80 showed little or no increase in the formation of sub - visible particles (Figures 3C - E). Therefore, by using PS80 at a concentration above 0.01%, the degradation of SAR445088 due to shaking can be suppressed.

[0180] Restoration action shaking As can be seen from Figure 4, the samples without PS80 had the highest tendency to form protein aggregates (HMW species) upon shaking. Samples containing more than 0.03% PS80 showed good resistance to the formation of HMW species (Figure 4A). Samples without PS80 showed an increase in turbidity as early as 1 hour after the restoration reaction shaking. 0.01% PS80 was sufficient to prevent the increase in turbidity (Figure 4B). Samples without PS80 had the maximum number of subvisible particles of 2 μm, 10 μm, and 25 μm as early as 1 hour after shaking. Thereafter, white, insoluble precipitates appeared, so the subvisible particles of samples without PS80 were not evaluated thereafter. Samples containing more than 0.03% PS80 showed resistance to subvisible particle formation (Figures 4C - E).

[0181] Restoration reaction shaking + silicone incorporation As can be seen from Figure 5, samples without PS80 showed an increase in turbidity as early as 1 hour after the restoration reaction shaking. 0.01% PS80 was sufficient to prevent the increase in turbidity. White, insoluble precipitates were formed in samples without PS80, so these samples were not evaluated after T0 (Figure 5A). In samples containing more than 0.01% PS80, the presence of silicone oil increased the number of subvisible particles of 2 μm. Samples containing more than 0.03% PS80 showed resistance to the formation of 10 μm and 25 μm particles even when subjected to restoration reaction shaking in the presence of silicone (Figures 5B - D). SEC analysis was not performed to prevent damage to the column by silicone oil.

[0182] IV bag dilution test As can be seen from Figure 6, PS80 had no effect on the formation of protein aggregates in the sample diluted and held in the IV bag (Figure 6A). PS80 at any concentration had a minimal effect on the turbidity in the test IV bags over 24 hours (Figure 6B). All IV bags had the target protein concentration of 3 mg / mL (Figure 6C). In samples containing less than 0.01% PS80, subvisible particles of 2 μm and 10 μm increased within 24 hours in the IV bag. Samples containing more than 0.03% PS80 showed resistance to subvisible particle formation (Figures 6D - F).

[0183] Short - term photostability test As can be seen from Figure 7, PS80 had no effect on the ability of the protein exposed to ambient light to form HMW species, and the impact on HMW species formation was comparable at all PS80 concentrations tested (Figure 7A). It was clear that the turbidity increased when all samples were exposed to ambient light for 3 days (Figure 7B). No degradation of PS80 was observed over the entire period of the test under any test conditions (Figure 7C). In samples containing less than 0.01% PS80, particles of 2 μm, 10 μm, and 25 μm increased over time. Samples containing more than 0.01% PS80 showed resistance to particle formation (Figures 7D - F).

[0184] Summary Considering all the mechanical, physical, and chemical stresses that the formulation is expected to encounter during its shelf life and that are reflected in the stress tests conducted in this study, the PS80 required to obtain a minimum acceptable protection against stress - induced formation of HMW species and subvisible particles appears to be at least 0.03%. However, when PS80 was set at 0.06% (600 ppm), the antibody formulation was more fully protected from all the stresses tested.

[0185] Example 5. Validity evaluation test for chelating agents The types and concentrations of chelating agents suitable for preventing the oxidation of PS80 in the SAR445088 formulation were determined. Chelating agents (EDTA or DTPA) and metal ions (iron (Fe), copper (Cu), and tungsten (W)) were incorporated into different batches of FDS at pH 7.0 containing 150 mg / mL of SAR445088, 150 mM of arginine HCl, 3% sucrose, 0.06% PS80, and 10 mM histidine as described in Table 3 and subjected to refrigerated, room temperature, and accelerated (40 °C) stability tests. These two chelating agents were selected based on their use in approved biopharmaceuticals. The metal ions were selected based on metals found in the water used in manufacturing, stainless steel containers or parts that may come into contact, and tungsten used in the formation of the needles of glass prefilled syringes.

[0186]

Table 3

[0187] Chelating Agent Test for Batch 1 As can be seen from Figure 8, in all samples containing 10 μM EDTA and with or without metal ions, degradation of PS80 was observed as early as 2 weeks at 40°C. On the other hand, when 10 μM DTPA was used, degradation of PS80 was alleviated up to 3 months at 40°C even in the presence of metal ions (Figure 8A). When EDTA was 50 μM or DTPA was 50 μM, PS80 did not degrade (Figure 8B). Furthermore, from ester profile analysis, it was found that the root cause of PS80 degradation was oxidation, and oxidation of PS80 was prevented by the presence of 10 μM DTPA (Figure 8C). The tendency to form HMW species was highest when 10 μM EDTA was used. When 10 μM DTPA was used, the formation of HMW species was less than when 10 μM EDTA was used. When 50 μm EDTA and DTPA were used, the formation of HMW species was more greatly inhibited compared to the case of 10 μM EDTA, but the difference was not so significant compared to the case of 10 μM DTPA (Figure 8D). When 10 μM EDTA was used, coloring of the solution was also observed compared to other test groups (Figure 9). From Biacore analysis, it was found that EDTA and DTPA did not directly affect the efficacy of the molecule. At 40°C, a slight decrease in efficacy over time was observed (Figure 8E). The chelating agent had no significant effect on the tendency of the molecule to undergo LC D32 isomerization, heavy chain (HC) M251 oxidation, HC PENNYK deamidation, or HC VSNK deamidation.

[0188] Chelating Agent Test for Batch 2 As can be seen from Fig. 10, in all samples with or without ions containing 10 μM EDTA, the degradation of PS80 was observed as early as 2 weeks at 40°C. On the other hand, when 10 μM DTPA was used, the degradation of PS80 was alleviated up to 3 months at 40°C even in the presence of metal ions (Fig. 10A). When EDTA was 50 μM or DTPA was 50 μM, PS80 did not degrade (Fig. 10B). The formation of HMW species was maximized when EDTA was 10 μM. The formation of HMW species when using 10 μM DTPA was not as obvious as when using 10 μM EDTA. When using 50 μm EDTA and DTPA, the formation of HMW species was more strongly inhibited compared to when using 10 μM EDTA, but the inhibition of HMW species formation by 50 μm EDTA was comparable to that of 10 μM DTPA (Fig. 10C). Regarding the formation of LMW species, no significant trend according to the chelating agent was observed (Fig. 10D). When EDTA was 10 μM, coloring of the solution was also observed compared to other test groups. From Biacore analysis, no loss of drug efficacy was observed in the samples after 6 months at 25°C whether in the presence or absence of the chelating agent, so it was found that EDTA and DTPA did not directly affect the drug efficacy of the molecule. At 40°C, a slight decrease in drug efficacy over time was observed (Fig. 10E). The chelating agent did not have a significant effect on the tendency of the molecule to undergo LC D32 isomerization, HC M251 oxidation, HC PENNYK deamidation, or HC VSNK deamidation.

[0189] Chelating Agent Test for Batch 3 As can be seen from Fig. 11, in all samples, less degradation of PS80 was observed compared to the previous batch. In formulations without a chelating agent or containing 10 μM EDTA after storage at 25 °C for 6 months, some degradation of PS80 was observed compared to samples containing 10 μM DTPA (Figs. 11A - B). Regarding the formation of HMW species, both formulations without a chelating agent or containing 10 μM EDTA were higher. When using 10 μM DTPA, it was clear that less HMW species were formed compared to when using 10 μM EDTA (Fig. 11C). Regarding the formation of LMW species, no significant trend according to the chelating agent was observed. In all groups tested, an overall increase in the formation of LMW species according to storage time and temperature was observed (Fig. 11D). No coloring of the solution was observed in samples at 5 °C or 25 °C, but it was observed in samples at 40 °C. From Biacore analysis, it was found that EDTA and DTPA did not directly affect the efficacy of the molecule, and after 3 months at 25 °C, the efficacy was substantially retained regardless of the chelating agent used. A slight decrease in efficacy over time was observed at 40 °C (Fig. 11E). The chelating agent had no significant effect on the tendency of the molecule to undergo LC D32 isomerization, HC M251 oxidation, HC PENNYK deamidation, or HC VSNK deamidation.

[0190] Chelating Agent Test of Batch 4 As can be seen from Figure 12, in all samples, less degradation of PS80 was observed compared to the previous batch. EDTA and DTPA appeared to mitigate the degradation of PS80 to a similar extent at all concentrations (Figures 12A - B). Similar HMW species formation profiles were observed for the EDTA - containing formulation and the DTPA - containing formulation when held at 5°C and 25°C. Regarding the formation of HMW species at 40°C, the DTPA - containing formulation was shown to be less than the formulation without a chelating agent or containing EDTA (Figure 12C). The DTPA - containing formulation was shown to form fewer LMW species compared to the formulation without a chelating agent or containing EDTA. In all groups tested, an overall increase in LMW species formation was observed with increasing storage time and temperature (Figure 12D). At 5°C or 25°C, no coloring of the solution was observed in any sample. At 40°C, the coloring of the solution with DTPA as the chelating agent was not obvious. From the Biacore analysis, it was found that EDTA and DTPA did not directly affect the efficacy of the molecule, and the efficacy was substantially maintained even after one month at 25°C. A slight decrease in efficacy over time was observed at 40°C (Figure 12E). The chelating agent did not have a significant effect on the tendency of the molecule to undergo LC D32 isomerization, HC M251 oxidation, HC PENNYK deamidation, or HC VSNK deamidation.

[0191] Summary Based on the comparative stability study of SAR445088 formulation samples containing DTPA or EDTA, a concentration of 10 μM DTPA was determined to most appropriately mitigate the degradation of PS80 across different batches, prevent the formation of HMW species and solution coloring, and have the least impact on the molecule's ability to maintain efficacy.

[0192] Example 6. pH Screening / DP Stability Test By subjecting different batches of SAR445088 to refrigerated, room temperature, accelerated, and frozen storage stability tests, the optimal buffer system and target pH for the SAR445088 formulation were identified. This test was conducted using different primary containers, namely the vials and prefilled syringes (PFS) described in Table 4, and the stability of SAR445088 liquid DP in these containers was tested in parallel.

[0193]

Table 4

[0194] Formulation test of Batch 1 PFS stability test As shown in Figure 13, under accelerated conditions at 40°C, SAR445088 had a higher tendency to form HMW species in potassium phosphate buffer than in histidine buffer. Overall, at room temperature (25°C) and refrigerated storage (5°C), SAR445088 maintained substantial stability under all pH and buffer conditions tested. Regarding storage in BD neopak low, BD neopak high, and OMPI syringes, no significant difference was observed in the formation of HMW species. In histidine buffer at pH 7.2, a slight decrease in HMW species formation was observed at T0 and subsequent time points compared to histidine buffer at pH 6.7 or 6.8 (Figure 13A). None of the buffers, pH, or PFS had a significant effect on the antibody concentration (Figure 13B) or pH (Figure 13C) measured at the end of the stability test period. An overall increase in turbidity was seen depending on time and temperature for all buffers and target pH. On the other hand, no significant difference in turbidity was observed among the different pH, buffer, or syringe types tested (Figure 13D). Under storage conditions at 25°C and 40°C, PS80 deteriorated to some extent over time in all formulations. Histidine-containing formulations had a higher degradation of PS80 than potassium phosphate-containing formulations at all pH values (Figure 13E).

[0195] As shown in Figure 14, except for the potassium phosphate formulation stored at pH 7.0 (K7.0) at 40°C for 3 months, for all pH values and buffers tested, no significant increase in subvisible particle formation occurred upon storage at different temperatures. The formation of 2-μm particles in OMPI PFS was higher than that in its comparator BD (Figure 14A). The formation of subvisible particles with particle diameters of ≤10 μm and ≤25 μm was almost unchanged, although there were a few outliers due to the variability in the evaluation (Figures 14B - C).

[0196] Vial stability test As shown in Figure 15, under the accelerated conditions at 40°C, SAR445088 had a higher tendency to form HMW species in potassium phosphate buffer than in histidine buffer. Overall, at room temperature (25°C) and refrigerated storage (5°C), SAR445088 maintained substantial stability under all pH and buffer conditions. When using the buffer at H7.2, a slight decrease in the formation of HMW species was observed at T0 and subsequent time points compared to H6.7 and H6.8 (Figure 15A). Neither the buffer nor the pH had a significant effect on the antibody concentration (Figure 15B) or pH (Figure 15C) measured at the end of the test period. An overall increase in turbidity was observed depending on time and temperature for all buffers and target pH values. However, the increase in turbidity was more evident in the potassium phosphate buffer-containing formulation than in the histidine-containing formulation (Figure 15D). It was clear that PS80 deteriorated over time in all formulations under the storage conditions of 25°C and 40°C. The histidine-containing formulation had a higher degree of PS80 deterioration than the potassium phosphate-containing formulation at all pH values (Figure 15E). The deterioration of PS80 was higher when the same formulation was placed in vials than when placed in PFS, probably because the headspace in the vial was large.

[0197] As shown in Figure 16, for all pH values and buffers tested, no significant increase in subvisible particle formation was observed even when stored at different temperatures. The formation of subvisible particles with sizes of ≤2μm, ≤10μm, and ≤25μm remained almost unchanged, although there were a few outliers due to the variability in the evaluation (Figures 16A - C).

[0198] Freeze - storage stability test As shown in Figure 17, it was observed that freezing and storing at - 80°C and - 30°C slightly decreased the formation of HMW species (Figure 17A). Freeze - storage had no observable effect on the antibody concentration (Figure 17B), pH (Figure 17C), turbidity (Figure 17D), and subvisible particles (Figure 17E) measured at the end of the storage period. SAR445088 was stable up to 12 months under freeze - storage.

[0199] Peptide mapping and Biacore binding analysis As shown in Figure 18, for all buffers and pH values tested, an overall increase in the degree of modification of hotspots according to temperature and time was observed. Specifically, regarding D32 isomerization, in the formulation using histidine buffer, the degree of modification was approximately 6% per week when stored at pH 6.8 and 40°C for 3 months, while it was approximately 3% when stored at 5°C for 12 months. No significant difference in the degree of modification was observed among different pH values and buffer types (Figures 18A - D). For the oxidation of HC M251 stored in K7.0 buffer at 40°C for 2 months (2M - 40C), the degree was on the trend side but seemed to be due to operator error. Biacore analysis showed that SAR445088 retained its relative activity after storage at 25°C for 6 months and at 5°C for 12 months in the sample in histidine buffer at pH 6.8. A slight decrease in drug efficacy was observed in the sample exposed to 40°C for 3 months.

[0200] Summary The SAR445088 formulation was shown to have a higher tendency to form HMW species in potassium phosphate buffer than in histidine buffer. The degree of PS80 degradation was lower when using potassium phosphate buffer than when using histidine buffer. However, the degradation of PS80 in the presence of histidine was effectively alleviated by using 10 μM DTPA. Therefore, histidine was selected as the buffer mainly because of the tendency of the molecules to aggregate. No significant differences were found in the histidine buffer in the pH range of 6.7 - 7.2 for the critical quality attributes (CQAs) of the formulation. Except for the relatively increased formation of 2 μm particles in OMPI PFS, no significant differences in CQAs were found when comparing BD Neopak Low, BD Neopak High, and OMPI syringes. In the OMPI syringe, the number of subvisible particles ≤10 μm and ≤25 μm was also high, but this number was below the USP-specified limit values (≤6000 10-μm particles per small-volume container and ≤600 25-μm particles per large-volume container).

[0201] Formulation test of Batch 2 PFS stability test As shown in Fig. 19, it was observed that the HMW species increased as a whole over time and at 40°C. Regarding the degree of HMW species formation occurring at 5°C or 25°C, no significant difference was found at pH 6.4 - 7.1. At 5°C and 25°C, it was observed that at H7.1, the formation of HMW species was slightly less at T0 and subsequent time points compared to the formulations of H6.4 and H6.8 (Fig. 19A). Regarding the LMW species, it was observed that they increased as a whole over time and at 40°C. At 5°C and 25°C, no tendency of LMW species formation according to the pH of the formulation was recognized (Fig. 19B). Regarding the antibody concentration (Fig. 19C) and pH (Fig. 19D) at the end of the storage period, no influence by the pH of the formulation was seen. For all the formulations tested, an overall increase in turbidity according to time and temperature was recognized, but no significant difference between different pHs was recognized (Fig. 19E). The observed degradation of PS80 was very slight in any of the test formulations under all storage stability test conditions (Fig. 19F). From the charge variant analysis, it was shown that in all the formulations tested, the generation of acidic variants increased as a whole at 25°C and 40°C, and correspondingly the monomer and basic variants decreased. Due to the decreased peak resolution, the charge variant analysis was performed only up to those stored at 40°C for 1 month. At 5°C, for all the conditions tested, the charge variants remained relatively unchanged (Fig. 19G).

[0202] As can be seen from Fig. 20, in all the formulations tested, the 2 - μm sub - visible particles were more numerous than the larger particles, which is probably due to the presence of silicone oil. On the other hand, the generation of sub - visible particles was not significantly increased by storing at different temperatures. The appearance of sub - visible particles with particle diameters of ≤2 μm, ≤10 μm, and ≤25 μm had several outliers within the range of deviation, but it hardly changed among the formulation test samples (Fig. 20A - C).

[0203] Vial Stability Test As shown in Figure 21, it was observed that the HMW species increased over time and overall at 40°C. For HMW species formation occurring at 5°C or 25°C, no significant difference was found at pH 6.4 - 7.1. At 5°C and 25°C, it was observed that at H7.1, the formation of HMW species was slightly less at T0 and subsequent time points compared to the formulations of H6.4 and H6.8 (Figure 21A). For the LMW species, it was observed that they increased over time and overall at 40°C. At 5°C and 25°C, no tendency of LMW species formation according to the pH of the formulation was recognized (Figure 21B). Regarding the antibody concentration (Figure 21C) and pH (Figure 21D) at the end of the storage period, no influence by the pH of the formulation was seen. For all the formulations tested, an overall increase in turbidity according to time and temperature was recognized, but no significant difference between different pHs was recognized (Figure 21E). No significant degradation of PS80 was observed in any of the test formulations under all storage stability test conditions (Figure 21F). From the charge variant analysis by cIEF, it was shown that in all the formulations tested, the generation of acidic variants increased overall at 25°C and 40°C, and correspondingly the monomer and basic variants decreased. Due to the decreased peak resolution, the charge variant analysis was performed only up to those stored at 40°C for 1 month. At 5°C, for all the conditions tested, the charge variants remained relatively unchanged (Figure 21G).

[0204] As shown in Figure 22, for any of the formulations tested, the formation of subvisible particles did not clearly and significantly increase by storing at different temperatures. The appearance of subvisible particles with particle diameters of ≦2μm, ≦10μm, and ≦25μm was almost the same among the formulation test samples, although there were a few outliers due to the variation in the evaluation (Figure 22A - C).

[0205] Frozen storage stability test As shown in Figure 23, the formation of HMW species when stored frozen at -80 °C and -30 °C did not vary significantly between the different formulations tested (Figure 23A). The formation of LMW species when stored frozen at -80 °C and -30 °C remained at a similar level over time. The formation of LMW species in the formulation sample of pH 7.1 (H7.1) containing histidine buffer stored at -30 °C was trending outwards. This was due to variability in the evaluation (Figure 23B). Frozen storage did not have a significant impact on the antibody concentration (Figure 23C), pH (Figure 23D), turbidity (Figure 23E), and amount of subvisible particle formation (Figure 23F) measured at the end of the storage period.

[0206] Peptide mapping and Biacore binding analysis As shown in Figure 24, an overall increase in the degree of modification of hot spots as a function of temperature and time was observed in vials and in PFS for all buffers and pHs tested. Specifically, for D32 isomerization at 40 °C, a degree of modification of 6% per week was observed in the formulation (H6.8) in histidine buffer at pH 6.8. No significant differences in the degree of modification were observed between different pH values and buffer types (Figure 24A - D). From Biacore analysis, it was shown that SAR445088 retained relative activity after storage for 6 months at 25 °C and 5 °C in samples in histidine buffer at pH 6.8. A slight decrease in drug efficacy was observed in all samples exposed to 40 °C for 3 months (Figure 24E).

[0207] Summary For the SAR45088 test formulations with pH values of 6.4 - 7.1, no significant differences were seen in any of the CQAs investigated. Formulation samples subjected to frozen stability in PFS and in vials maintained substantial stability over the test period (12 months).

[0208] Formulation testing of Batch 3 PFS stability test As shown in Fig. 25, it was observed that the HMW species increased over time and as a whole at 40°C. Regarding the degree of HMW species formation observed at 5°C or 25°C, no significant difference was found between pH 6.4 and 7.1. At 5°C and 25°C, it was observed that the formation of HMW species at H7.1 was slightly less at T0 and subsequent time points compared to the formulations of H6.4 and H6.8 (Fig. 25A). No clear difference was recognized in the degree of LMW species formation among the pH values tested (Fig. 25B). The pH of the formulation had no observable effect on the concentration (Fig. 25C) or pH (Fig. 25D) measured at the end of the storage period. For all the formulations tested, an overall increase in turbidity was recognized depending on time and temperature, but no significant difference was found among different pH values (Fig. 25E). The observed degradation of PS80 was very slight in any of the test formulations under all the storage stability test conditions (Fig. 25F). Finally, in all the formulations tested, the number of sub-visible particles of 2 μm was larger than the number of particles of other particle sizes. However, when stored at 5°C and 25°C, no obvious increase in the concentration of sub-visible particles was seen. At 40°C, an increase over time in sub-visible particles was observed in all the formulations (Fig. 25G). No significant difference was found in the tendency of sub-visible particles among the pH values tested.

[0209] Vial stability test As shown in Fig. 26, it was observed that the HMW species increased over time and as a whole at 40°C. Regarding the degree of HMW species formation observed at 5°C or 25°C, no significant difference was found at pH 6.4 - 7.1. At 5°C and 25°C, it was observed that the formation of HMW species was slightly less at H7.1 compared to the formulations of H6.4 and H6.8 at T0 and subsequent time points (Fig. 26A). No clear difference was recognized in the degree of LMW species formation among the pH values tested (Fig. 26B). The pH of the formulation had no observable effect on the concentration (Fig. 26C) or pH (Fig. 26D) measured at the end of the storage period. For all formulations tested, an overall increase in turbidity according to time and temperature was recognized, but no significant difference was found among different pH values (Fig. 26E). The observed degradation of PS80 was negligible in any of the test formulations under all storage stability test conditions (Fig. 26F). Finally, it was found that the degree of subvisible particle formation clearly increased by storing any of the test formulations at different temperatures. The degree of subvisible particle formation with particle diameters of ≤2μm, ≤10μm, and ≤25μm was almost unchanged, although there were a few outliers due to operator error (Fig. 26G).

[0210] Peptide Mapping and Biacore Binding Analysis As shown in Fig. 27, an overall increase in the degree of modification of hot spots according to temperature and time was observed for all buffers and pH values tested. Specifically, regarding D32 isomerization at 40°C, it was observed that the degree of modification was 6% per week in the formulation (H6.8) in histidine buffer at pH 6.8. No significant difference was recognized in the degree of modification among different pH values and types of buffers (Fig. 27A - D). From Biacore analysis, it was shown that SAR445088 retained relative activity after storage at 25°C and 5°C for 3 months in the sample in histidine buffer at pH 6.8. A slight decrease in drug efficacy was observed in all samples exposed to 40°C for 3 months (Fig. 27E).

[0211] Summary In the SAR445088 test formulation with a pH of 6.4 - 7.1, no significant differences were observed in all the investigated CQAs. The PFS and the formulation samples in the vials maintained substantially stable throughout the test period (6 months).

[0212] Formulation test of Batch 4 Vial stability test As shown in Figure 28, it was observed that the HMW species increased over time and overall at 40°C, but the degree was small compared to the previous batches. Regarding the degree of HMW species formation observed at 5°C or 25°C, no significant differences were found at pH 6.4 - 7.1. At 5°C and 25°C, it was observed that HMW species formation was slightly less at pH 7.1 than at pH 6.4 and pH 6.8 formulations at T0 and subsequent time points (Figure 28A). At 5°C and 25°C, no clear differences were recognized in the degree of LMW species formation among the tested pH values, but a slight increase over time was observed at 40°C (Figure 28B). The pH of the formulation had no observable effect on the concentration (Figure 28C) or pH (Figure 28D) measured at the end of the storage period. For all the formulations tested, an overall increase in turbidity was recognized depending on time and temperature, but no significant differences were found among different pH values (Figure 28E). The observed degradation of PS80 was negligible in any of the test formulations under all the storage stability test conditions (Figure 28F). Finally, no obvious increase in the concentration of subvisible particles was found in any of the test formulations by storing at different temperatures. The degree of subvisible particle formation with particle sizes of ≤2μm, ≤10μm, and ≤25μm was almost unchanged, although there were a few outliers due to the variation in the evaluation (Figure 28G).

[0213] Peptide mapping and Biacore binding analysis As shown in Figure 29, an overall increase in the degree of modification of the hot spots according to temperature and time was observed for all buffers and pH values tested. Specifically, for D32 isomerization at 40°C, a degree of modification of 10% per week was observed for the formulation (H6.8) in histidine buffer at pH 6.8. No significant difference in the degree of modification was observed between different pH values and buffer types (Figures 29A - D). From Biacore analysis, it was shown that SAR445088 retained relative activity after storage for one month in the sample in histidine buffer at pH 6.8 (Figure 29E).

[0214] Summary For the SAR445088 test formulations with pH values of 6.4 - 7.1, no significant differences were seen in all the investigated CQAs throughout the test period (3 months).

[0215] Formulation Test of Batch 5 As shown in Figure 30, it was observed that the degree of HMW species formation increased slightly over time and at 40°C, but the extent was small compared to the previous batches (Figure 30A). The formation of LMW species was observed to increase slightly over time at 40°C (Figure 30B). The pH of the formulation had no observable effect on the measured concentration (Figure 30C) or pH (Figure 30D) at the end of the storage period. An overall increase in turbidity according to time and temperature was seen in all test formulations (Figure 30E). The observed degradation of PS80 was negligible under any storage stability conditions (Figure 30F). Finally, the degree of subvisible particles did not increase significantly by storing at different temperatures. The concentrations of subvisible particles with particle sizes of ≤2μm, ≤10μm, and ≤25μm remained almost unchanged (Figure 30G).

[0216] Example 7. Robustness Study The SAR445088 formulations using different batches of SAR445088 were subjected to various physicochemical stresses as described in Table 5.

[0217] [Table 5]

[0218] Restore action oscillation (vial) As shown in Figure 31, although the starting values of HMW species in different batches were various, the restore action oscillation in the vial did not have a significant effect on the degree of HMW species formation (Figure 31A). The restore action oscillation did not have an observable effect on LMW species formation (Figure 31B), turbidity (Figure 31C), or charge variant formation (Figure 31D). From the subvisible particle analysis, although the subvisible particles at the start were different among batches, it was shown that there was no tendency for the formation of subvisible particles of ≤2 μm, ≤10 μm, and ≤25 μm to increase even with restore action oscillation in the vial (Figure 31E).

[0219] Restore action oscillation (PFS) As shown in Figure 32, although the starting values of HMW species in different batches were various, the restore action oscillation during PFS did not have a significant effect on HMW species formation (Figure 32A). The restore action oscillation did not have an observable effect on LMW species formation (Figure 32B), turbidity (Figure 32C), or charge variant formation (Figure 32D). From the subvisible particle analysis, the subvisible particles at the start were different among batches and the difference was larger compared to the vial, but it was shown that there was no tendency for the formation of subvisible particles of ≤2 μm, ≤10 μm, and ≤25 μm to increase even with restore action oscillation during PFS (Figure 32E).

[0220] Freeze - thaw cycle As shown in Figure 33, the starting values of HMW species in different batches were various, but freeze-thaw did not have a significant effect on the degree of HMW species formation (Figure 33A). Freeze-thaw did not have an observable effect on the LMW species formation (Figure 33B), turbidity (Figure 33C), concentration (Figure 33D), or pH (Figure 33E) of the formulation measured at the end of the storage period. Subvisible particle analysis showed a slight tendency for the formation of particles ≤2 μm, ≤10 μm, and ≤25 μm to increase by repeating the freeze-thaw cycle, but the number of particles per container remained well within the USP specified ranges (≤6000 10-μm particles per small-volume container and ≤600 25-μm particles per small-volume container) (Figure 33F).

[0221] Photo stability test As shown in Figure 34, it was observed that the formation of HMW species increased slightly with increasing room temperature and light exposure compared to the control samples covered with aluminum foil (Figure 34A). Light exposure did not have an observable effect on the LMW species formation (Figure 34B), turbidity (Figure 34C), concentration (Figure 34D), or pH (Figure 34E) of the formulation measured at the end of the storage period. Degradation of PS80 was not observed in any of the samples (Figure 34F). No significant change in charge variant formation was seen even after exposure to environmental light for up to 14 days (Figure 34G). Total peptide map analysis showed an overall increase in D32 isomerization, VSNK deamidation, and PENNYK deamidation, suggesting that these were not a direct result of light exposure as the control samples protected by the cover were also included. However, M251 oxidation was increased by light exposure compared to the covered control. This effect was expected (Figure 34H). Subvisible particle formation analysis showed that environmental light exposure had little effect on subvisible particles ≤2 μm, ≤10 μm, and ≤25 μm (Figure 34I).

[0222] Summary The SAR445088 formulation was shown to be robust to physicochemical stresses such as list action shaking, freeze-thaw cycles, and 14 days of environmental light exposure within vials and PFS. Robustness was observed across different batches and between pH values of 6.4 - 7.1.

[0223] Example 8. Exemplary Liquid Formulations Through the characterization test of the formulations described herein, the appropriate concentration of PS80 was identified, the optimal type and concentration of chelating agents were identified, the appropriate buffer and pH values centered around the optimal pH were identified to impart stability to DP and support the robustness of DP against physicochemical stresses, in addition to managing D32 isomerization, and the effect of adding 3% sucrose was evaluated.

[0224] Specifically, by increasing the pH to 6.8 and higher and including arginine hydrochloride in the formulation, the D32 isomerization rate in the LC CDR1 of SAR445088 decreased. Such optimized formulations conferred resistance to isomerization across different batches of SAR445088 tested under refrigerated, room temperature, and accelerated storage conditions.

[0225] The optimal PS80 concentration was determined by evaluating the resistance of formulations containing different concentrations of PS80 to mechanical stress, dilution simulating clinical use, and short-term light exposure. From these tests, 600 ppm (0.06%) of PS80 was identified as the optimal surfactant concentration to protect SAR445088 from the effects of shear, IV dilution, and light exposure.

[0226] The metal ion content of various API batches varies between batches, and formulations containing 10 μM of EDTA do not have the ability to reduce the degradation of PS80 across different batches. Therefore, 10 μM of DTPA was selected as the optimal chelating agent to reduce the risk of PS80 degradation. DTPA is more effective in preventing the degradation (oxidation) of PS80, reducing the formation of HMW species, and avoiding the coloring of the product solution, which was observed in formulations containing 10 μM of EDTA or formulations without a chelating agent (see Figures 8A, 8B, 8D, 9, 10A, 10B, and 10C).

[0227] In a direct comparison of the SAR445088 formulation in histidine buffer and phosphate buffer, the histidine buffer at pH 6.8 was shown to have the optimal ability to reduce the tendency to aggregate under accelerated stability test conditions.

[0228] From the stability tests, it was shown that the addition of sucrose had no adverse effect on the stability of the protein and conferred resistance to aggregation that may occur during repeated freeze-thaw cycles.

[0229] When different PFSs were compared, they showed similar product profiles. The DP stability tests in vials and PFS were carried out at pH values of 6.4 - 7.1 under refrigerated, ambient, accelerated, and frozen conditions. The PFS showed acceptable product profiles across different batches within this pH range. That is, the tendency of HMW species was overall controlled in each case, the oxidation of PS80 was reduced, and the molecular-specific D32 isomerization was controlled.

[0230] An exemplary SAR445088 liquid formulation shown to be optimal for the stabilization of not only 150 mg / mL of liquid DP but also 150 mg / mL of frozen FDS from the analysis described herein is 10 mM histidine, 150 mM arginine HCl, 3% (w / v) sucrose, 10 μM DTPA, 0.06% (w / v) PS80, and the pH is 6.8 ± 0.3.

[0231] Example 9. Components of the formulation SAR445088DP can be composed of SAR445088, L-histidine, L-histidine hydrochloride monohydrate, L-arginine hydrochloride, sucrose, DTPA and polysorbate 80. These pharmaceutical additives are known to have good tolerance after parenteral administration and are water-soluble. The details of the exemplary DP composition and the closures of the containers are summarized in Table 6 below.

[0232]

Table 6

[0233] All references, patents and patent applications disclosed in this specification are hereby incorporated by reference into this specification with respect to the subject matter to which each is cited, and in some cases the entire document may be included.

[0234] In this specification, the indefinite articles "a" and "an" used in this specification and the claims are to be understood to mean "at least one" unless clearly indicated otherwise.

[0235] In any method comprising two or more steps or acts claimed in this specification, the order of the steps or acts of the method is not necessarily limited to the order of the steps or acts of the method described unless clearly indicated otherwise.

[0236] In the claims and the above specification, all transitional phrases such as "comprising", "including", "possessing", "having", "containing", "accompanying", "holding" and "consisting of" are open-ended, i.e., they are to be understood to mean including but not limited to. Only the transitional phrases "consisting of" and "consisting essentially of" are to be considered closed or semi-closed transitional phrases as described in Section 2111.03 of the United States Patent and Trademark Office Patent Examination Procedure Manual.

[0237] The terms "about" and "substantially" preceding a numerical value mean ±10% of the recited numerical value.

[0238] Where a range of values is indicated, each value between the upper and lower boundary values, including the boundary values themselves, is specifically intended and recited therein.

Claims

1. (a) A humanized antibody that specifically binds to complement component C1s, comprising a light chain (LC) complementarity-determining region (CDR) 1 containing the amino acid sequence of SEQ ID NO: 1, LC CDR 2 containing the amino acid sequence of SEQ ID NO: 2, LC CDR 3 containing the amino acid sequence of SEQ ID NO: 3, and heavy chain (HC) CDR 1 containing the amino acid sequence of SEQ ID NO: 4, HC CDR 2 containing the amino acid sequence of SEQ ID NO: 5, and HC CDR 3 containing the amino acid sequence of SEQ ID NO: 6, (b) Arginine or salt thereof in a concentration of approximately 50 mM to approximately 200 mM A composition containing the following:

2. The composition according to claim 1, comprising approximately 100 mM to approximately 200 mM or approximately 100 mM to approximately 150 mM of arginine or a salt thereof.

3. The composition according to claim 1, wherein the arginine salt is arginine hydrochloride, arginine citrate, arginine oxalate, arginine phosphate, arginine succinate, or arginine tartrate.

4. The composition according to claim 3, wherein the arginine salt is arginine hydrochloride, and optionally the composition comprises about 150 mM arginine hydrochloride.

5. The composition according to claim 1, further comprising a buffering agent at a concentration of approximately 1 mM to approximately 50 mM, approximately 5 mM to approximately 25 mM, or approximately 10 mM to approximately 20 mM.

6. The buffer is histidine, acetate, citrate, oxalate, phosphate, succinate, or tartrate, and optionally the composition comprises about 10 mM histidine, according to claim 5.

7. The composition according to claim 1, further comprising a stabilizer at a concentration of about 1% to about 8% (w / v) or about 1% to about 5% (w / v).

8. The stabilizer is sucrose, sorbitol, or trehalose, and optionally, The composition according to claim 7, comprising about 3% (w / v) sucrose.

9. The composition according to claim 1, further comprising a chelating agent, wherein the chelating agent is optionally concentrated in a concentration of about 1 μM to about 50 μM, or about 5 μM to about 25 μM, or about 10 μM to about 25 μM.

10. The composition according to claim 9, wherein the chelating agent is diethylenetriaminepentaacetic acid (DTPA), ethylenediaminetetraacetic acid (EDTA), or methionine, and optionally the composition comprises about 10 μM of diethylenetriaminepentaacetic acid (DTPA).

11. The composition according to claim 1, further comprising a surfactant at a concentration of about 0.01% to about 0.1% (w / v) or about 0.03% to about 0.06% (w / v).

12. The composition according to claim 11, wherein the surfactant is polysorbate 80 (PS80) or poloxamer 188 (P188), and optionally the composition comprises about 0.06% (w / v) of polysorbate 80.

13. The composition according to claim 1, wherein the composition has a pH of about 6 to about 7.5, about 6 to about 7, about 6.5 to about 7.5, or about 6.5 to about 7.1, and optionally, the composition has a pH of about 6.

8.

14. The composition according to claim 1, wherein the composition comprises about 50 mg / mL to about 250 mg / mL or about 100 mg / mL to about 200 mg / mL of the anti-C1s antibody, and optionally comprises about 150 mg / mL of the anti-C1s antibody.

15. (a) The antibody in an amount of approximately 50 mg / mL to approximately 250 mg / mL, (b) Arginine HCl at approximately 50 mM to approximately 200 mM, (c) Histidine in concentrations of approximately 1 mM to 50 mM, (d) Sucrose in an amount of approximately 1% to approximately 8% (w / v), (e) Diethylenetriaminepentaacetic acid (DTPA) in a concentration of approximately 1 μM to approximately 50 μM, (f) Polysorbate 80 (PS80) in an amount of approximately 0.01% to approximately 0.1% (w / v) The composition according to claim 1, comprising and having a pH of about 6 to about 7.

5.

16. (a) The antibody in an amount of approximately 150 mg / mL, (b) Approximately 150 mM L-arginine HCl, (c) Approximately 10 mM histidine and (d) Approximately 3% (w / v) sucrose, (e) Approximately 10 μM of DTPA and (f) PS80 at approximately 0.06% (w / v) The composition according to claim 15, comprising and having a pH of about 6.5 to about 7.1 or about 6.

8.

17. The composition according to claim 1, wherein the antibody comprises a light chain variable region (VL) containing the amino acid sequence of SEQ ID NO: 7 and a heavy chain variable region (VH) containing the amino acid sequence of SEQ ID NO:

8.

18. The aforementioned antibody is a Fab fragment, F(ab') 2 The composition according to claim 1, wherein the composition is a fragment, scFv, or Fv.

19. The antibody comprises a heavy chain constant region of isotype IgG4, optionally comprising proline, glutamic acid, leucine, and serine substitutions at amino acid residues 108, 115, 308, and 314, respectively, relative to the IgG4 constant region sequence of SEQ ID NO: 11, and further optionally comprising a light chain comprising the amino acid sequence of SEQ ID NO:

9. The composition according to claim 1, comprising a heavy chain containing the amino acid sequence of SEQ ID NO:

10.

20. The antibody has a lower isomerization rate at D32 of the light chain CDR1 compared to the antibody in the corresponding formulation without arginine or its salt, and optionally, the isomerization at D32 is determined by whole peptide mapping analysis; and / or The aforementioned antibody is (i) Having an isomerization rate that is at least about 2-3% lower per week at 40°C, at least about 1-3% lower per month at 25°C, or at least about 0.4-0.6% lower per month at 5°C compared to the antibody in the corresponding formulation without arginine or its salt; or (ii) Having an isomerization rate about 10% lower after storage at 25°C for 12 weeks compared to the antibody in the corresponding preparation without arginine or its salt; or (iii) The composition according to claim 1, having an isomerization rate of less than 7.5% after storage at 5°C for 12 weeks, less than 20% after storage at 25°C for 12 weeks, or less than 35% after storage at 40°C for 12 weeks.

21. The composition according to any one of claims 1 to 20, wherein the composition is in liquid form, freeze-dried form, or reconstituted freeze-dried form, and optionally in liquid form.

22. A container for containing the composition according to any one of claims 1 to 20, wherein the container is optionally a vial or a syringe, and further optionally the syringe is a pre-filled syringe.

23. A kit or manufactured article comprising the container described in claim 22.

24. A pharmaceutical dosage form suitable for parenteral administration to humans, comprising a composition according to any one of claims 1 to 20 in a container.

25. A pharmaceutical composition comprising the composition according to any one of claims 1 to 20 for use in reducing the level of complement component cleavage products in humans, wherein the human optionally has chronic inflammatory demyelinating polyneuropathy (CIDP), cold agglutinin disease (CAD), immune thrombocytopenic purpura (ITP), or antibody-mediated rejection (AMR).

26. A pharmaceutical composition comprising the composition according to any one of claims 1 to 20 for use in inhibiting C1s-mediated cleavage of complement components in humans, wherein the human optionally has chronic inflammatory demyelinating polyneuropathy (CIDP), cold agglutinin disease (CAD), immune thrombocytopenic purpura (ITP), or antibody-mediated rejection (AMR).

27. A pharmaceutical composition comprising the composition according to any one of claims 1 to 20 for use in treating a complement-mediated disorder in a person in need thereof, wherein the complement-mediated disorder is optionally chronic inflammatory demyelinating polyneuropathy (CIDP), cold agglutinin disease (CAD), immune thrombocytopenic purpura (ITP), or antibody-mediated rejection (AMR).

28. The pharmaceutical composition according to claim 27, which is formulated for intravenous or subcutaneous use.

29. A drug delivery device comprising a primary container for containing the composition according to any one of claims 1 to 20, wherein the drug delivery device is a sleeve-activated self-injector for manually inserting a needle.

Images