Biopharmaceutical compositions and related methods

Abstract

The present application relates to biopharmaceutical compositions and related methods. The invention described herein provides compositions comprising an anti-BCMA antigen binding protein and related methods for treating a BCMA mediated disease or disorder.

Description

[0001] This application is a divisional application of a Chinese patent application with an application date of July 31, 2020, an application number of 202080069015.7, and an invention title of "Biopharmaceutical Compositions and Related Methods".

[0002] Sequence Listing

[0003] This application contains a sequence listing that has been electronically submitted in ASCII format and is hereby incorporated by reference in its entirety. Field of the Invention

[0004] The invention described herein provides compositions comprising an anti-BCMA antigen-binding protein and related methods for treating BCMA-mediated diseases or disorders. Background of the Invention

[0005] BCMA (CD269 or TNFRSF17) is a member of the TNF receptor superfamily. It is a non-glycosylated integral membrane receptor for the ligands BAFF and APRIL. The ligands of BCMA can also bind to other receptors: TACI (transmembrane activator and calcium modulator and cyclophilin ligand interactor), which binds APRIL and BAFF; and BAFF-R (BAFF receptor or BR3), which shows restricted but high affinity for BAFF. Collectively, these receptors and their corresponding ligands jointly regulate different aspects of humoral immunity, B cell development, and homeostasis.

[0006] The expression of BCMA is generally restricted to the B cell lineage and has been reported to increase terminal B cell differentiation. BCMA is expressed by human plasmablasts, plasma cells from tonsils, spleen, and bone marrow, but also by tonsillar memory B cells and germinal center B cells, which have a low TACI-BAFFR phenotype (Darce et al., 2007). BCMA is almost absent in naive and memory B-25 cells (Novak et al., 2004a and b). The BCMA antigen is expressed on the cell surface and is thus accessible to antibodies, but is also expressed in the Golgi apparatus. As its expression profile indicates, BCMA signaling, which is generally associated with B cell survival and proliferation, is important in the late stages of B cell differentiation as well as in the survival of long-lived bone marrow plasma cells (O'Connor et al., 2004) and plasmablasts (Avery et al., 2003). In addition, since BCMA binds APRIL with high affinity, indicating that the BCMA-APRIL signaling axis dominates in the later stages of B cell differentiation, this may be the most physiologically relevant interaction.

[0007] It has been reported that BCMA expression (transcripts and proteins) is associated with disease progression in various B cell disorders, including B cell cancers such as multiple myeloma (MM). MM is a clonal B cell malignancy that occurs at multiple sites within the bone marrow and then spreads to the circulation; it can arise de novo or from monoclonal gammopathy of undetermined significance (MGUS). It is typically characterized by increased paraprotein and osteoclast activity, as well as hypercalcemia, cytopenia, renal insufficiency, hyperviscosity, and peripheral neuropathy. Reduced normal antibody levels and neutrophil counts are also common, leading to susceptibility to life-threatening infections. BCMA is associated with the growth and survival of myeloma cell lines in vitro (Novak et al., 2004 and Moreaux et al., 2004). SUMMARY OF THE INVENTION

[0008] A composition comprising an isomerized variant of an anti-BCMA antibody, wherein the isomerized variant comprises a heavy chain amino acid sequence comprising CDRH1 of SEQ ID NO: 1, CDRH2 of SEQ ID NO: 2, and CDRH3 of SEQ ID NO: 3, and a light chain amino acid sequence comprising CDRL1 of SEQ ID NO: 4, CDRL2 of SEQ ID NO: 5, and CDRL3 of SEQ ID NO: 6; wherein the composition comprises ≤25% of the isomerized variant.

[0009] A composition comprising an oxidized variant of an anti-BCMA antibody, wherein the oxidized variant comprises a heavy chain amino acid sequence comprising CDRH1 of SEQ ID NO: 1, CDRH2 of SEQ ID NO: 2, and CDRH3 of SEQ ID NO: 3, and a light chain amino acid sequence comprising CDRL1 of SEQ ID NO: 4, CDRL2 of SEQ ID NO: 5, and CDRL3 of SEQ ID NO: 6; wherein the composition comprises ≤40% of the oxidized variant.

[0010] A composition comprising an anti-BCMA antibody, the antibody comprising CDRH1 having the amino acid sequence set forth in SEQ ID NO: 1, CDRH2 having the amino acid sequence set forth in SEQ ID NO: 2, CDRH3 having the amino acid sequence set forth in SEQ ID NO: 3, CDRL1 having the amino acid sequence set forth in SEQ ID NO: 4, CDRL2 having the amino acid sequence set forth in SEQ ID NO: 5, and CDRL3 having the amino acid sequence set forth in SEQ ID NO: 6; wherein the composition comprises 0.1-25% isomerization at D103 of CDRH3.

[0011] A composition comprising an anti-BCMA antibody, the antibody comprising a CDRH1 having the amino acid sequence set forth in SEQ ID NO: 1, a CDRH2 having the amino acid sequence set forth in SEQ ID NO: 2, a CDRH3 having the amino acid sequence set forth in SEQ ID NO: 3, a CDRL1 having the amino acid sequence set forth in SEQ ID NO: 4, a CDRL2 having the amino acid sequence set forth in SEQ ID NO: 5, and a CDRL3 having the amino acid sequence set forth in SEQ ID NO: 6; wherein the composition comprises 0.1-40% oxidation at M34 of CDRH1.

[0012] A composition comprising an anti-BCMA antibody that is at least about 90% identical to the heavy chain amino acid sequence of SEQ ID NO: 9 and the light chain amino acid sequence of SEQ ID NO: 10, wherein the composition comprises 0.1-25% isomerization at D103 of CDRH3.

[0013] A composition comprising an anti-BCMA antibody that is at least about 90% identical to the heavy chain amino acid sequence of SEQ ID NO: 9 and the light chain amino acid sequence of SEQ ID NO: 10, wherein the composition comprises 0.1-40% oxidation at M34 of CDRH1.

[0014] A composition comprising an anti-BCMA antibody-drug-conjugate (ADC), wherein the DL2 percentage is at least about 30%, about 15% to about 27% or about 15% to about 32%; the DL4a percentage is at least about 30%, about 35% to about 38% or about 30% to about 40%; the DL4b percentage is at least about 5%, about 7% to about 9% or about 5% to about 10%; the DL6 percentage is at least about 10%, about 14% to about 20% or about 10% to about 20%; and / or the DL8 is at least about 1%, about 6.0% to about 12.0% or about 4% to about 15%.

[0015] A composition comprising an anti-BCMA antibody-drug-conjugate (ADC), wherein the DL0 percentage is less than or equal to about 10% or about 5%. Brief Description of the Drawings

[0016] Figure 1 A schematic representation depicting a heterogeneous mixture of DL species within an ADC composition.

[0017] Figure 2 A representative HIC peak characterization for determining the DAR distribution in an ADC composition.

[0018] Figure 3 Shows the effect of the average DAR of an ADC composition on tumor volume in a xenograft model.

[0019] Figure 4 Depicts a representative cIEF electrophoretogram of belantamab. DETAILED DESCRIPTION OF THE INVENTION

[0020] The invention described herein provides compositions comprising an anti-BCMA antigen-binding protein and related methods for treating BCMA-mediated diseases or disorders. It should be understood that the compositions comprising an anti-BCMA antibody as described herein may also be referred to as an anti-BCMA antibody population as described herein: these phrases are interchangeable.

[0021] Anti-BCMA antigen-binding protein

[0022] The anti-BCMA antigen-binding proteins in the compositions described herein can be used to treat or prevent various BCMA-mediated diseases, including, for example, B cell-mediated cancers such as lymphoma and multiple myeloma. The anti-BCMA antigen-binding proteins described herein can bind to human BCMA, for example, human BCMA having the amino acid sequence of GenBank accession number Q02223.2, or a gene encoding human BCMA having at least 90% homology or at least 90% identity thereto.

[0023] As used herein, the term "antigen-binding protein" refers to antibodies, antibody fragments, and other protein constructs capable of binding BCMA, such as human BCMA. The antigen-binding proteins of the present invention can comprise the heavy chain variable region and the light chain variable region of the present invention, which can be formatted into the structure of a natural antibody or a functional fragment or equivalent thereof. Thus, when paired with an appropriate light chain, the antigen-binding proteins of the present invention can comprise the V H regions of the present invention formatted as a full-length antibody, (Fab')2 fragment, Fab fragment, or an equivalent thereof (e.g., scFV, diabody, triabody, or tetrabody, Tandab, etc.). The antibody can be IgG1, IgG2, IgG3, or IgG4; or IgM; IgA, IgE, or IgD, or a modified variant thereof. The constant domain of the antibody heavy chain can be selected accordingly. The light chain constant domain can be a κ or λ constant domain. In addition, the antigen-binding proteins can comprise all classes of modifications, for example, IgG dimers, Fc mutants that no longer bind Fc receptors or mediate Clq binding. The antigen-binding proteins can also be chimeric antibodies of the type described in WO86 / 01533, which comprise an antigen-binding region and a non-immunoglobulin region.

[0024] In another aspect of the present invention, the antigen-binding protein can be a dAb, Fab, Fab', F(ab')2, Fv, diabody, triabody, tetrabody, minibody or microbody. In one aspect of the present invention, the antigen-binding protein can be a fully human, humanized or chimeric antibody. In a further aspect, the antigen-binding protein is a humanized antibody. In one aspect of the present invention, the antigen-binding protein is a monoclonal antibody.

[0025] Chimeric antigen receptors (CARs) have been developed as artificial T cell receptors to generate new specificities in T cells without the need to bind to MHC antigen peptide complexes. These synthetic receptors can contain a target-binding domain that associates with one or more signaling domains through a flexible linker in a single fusion molecule. The target-binding domain can be used to target T cells to specific targets on the surface of pathological cells, and the signaling domain contains the molecular machinery for T cell activation and proliferation. A flexible linker that traverses the T cell membrane (i.e., forms a transmembrane domain) can allow cell membrane display of the target-binding domain of the CAR. CARs can successfully allow T cells to be redirected against antigens expressed on the surface of tumor cells from various malignancies, including lymphomas and solid tumors (Jena et al., 2010, Blood, 116(7):1035-44). In one aspect of the present invention, the anti-BCMA antigen-binding protein can comprise a chimeric antigen receptor. In a further aspect, the CAR can comprise a binding domain, a transmembrane domain, and an intracellular effector domain.

[0026] Exemplary anti-BCMA antigen-binding proteins and methods for their preparation are disclosed in International Publication No. WO2012 / 163805, which is incorporated herein by reference in its entirety. Other exemplary anti-BCMA antigen-binding proteins include those described in WO2016 / 014789, WO2016 / 090320, WO2016 / 090327, WO2016 / 020332, WO2016 / 079177, WO2014 / 122143, WO2014 / 122144, WO2017 / 021450, WO2016 / 014565, WO2014 / 068079, WO2015 / 166649, WO2015 / 158671, WO2015 / 052536, WO2014 / 140248, WO2013 / 072415, WO2013 / 072406, WO2014 / 089335, US2017 / 165373, WO2013 / 154760, and WO2017 / 051068, each of which is incorporated herein by reference in its entirety.

[0027] In another embodiment, the anti-BCMA antigen-binding proteins described herein can inhibit the binding of BAFF and / or APRIL to the BCMA receptor. In another embodiment, the anti-BCMA antigen-binding proteins described herein may be capable of binding to FcγRIIIA or may be capable of having FcγRIIIA-mediated effector functions.

[0028] In one embodiment, the anti-BCMA antigen-binding protein comprises an antibody (“anti-BCMA antibody”). In another embodiment, the anti-BCMA antigen-binding protein comprises a monoclonal antibody. As used herein, the term “antibody” refers to a molecule having an immunoglobulin-like domain (e.g., IgG, IgM, IgA, IgD, or IgE), and can include monoclonal, recombinant, polyclonal, chimeric, human, and humanized molecules of that type. Monoclonal antibodies can be cloned from eukaryotic cells expressing the antibody or from prokaryotic closed cells. Monoclonal antibodies can also be produced from eukaryotic cell lines that can recombinantly express the heavy and light chains of the antibody by introducing nucleic acid sequences encoding the heavy and light chains into the cells. Exemplary methods for producing antibodies from different eukaryotic cell lines such as Chinese hamster ovary cells, hybridomas, or immortalized antibody cells derived from animals (e.g., humans) are well known to those skilled in the art.

[0029] Antibodies can be derived from, for example, rats, mice, primates (e.g., cynomolgus monkeys, Old World monkeys, or great apes), humans, or other sources such as nucleic acids encoding antibody molecules generated using molecular biology techniques known to those skilled in the art.

[0030] Antibodies can comprise a constant region, which can be any isotype or subclass. The constant region can be of the IgG isotype, such as IgG1, IgG2, IgG3, IgG4, or variants thereof.

[0031] The antigen-binding protein can comprise one or more modifications, including, for example, a mutated constant domain such that when the antigen-binding protein is an antibody, the antibody has enhanced effector function / ADCC and / or complement activation.

[0032] In one embodiment, the anti-BCMA antibody has enhanced antibody-dependent cell-mediated cytotoxicity (ADCC) effector function. As used herein, the term "effector function" means one or more of antibody-dependent cell-mediated cytotoxicity (ADCC), complement-dependent cytotoxicity (CDC)-mediated responses, Fc-mediated phagocytosis, and / or antibody recycling via the FcRn receptor. For IgG antibodies, effector functions can include ADCC and ADCP can be mediated by the interaction of the heavy chain constant region with members of the Fcγ receptor family present on the surface of immune cells. In humans, these may include FcγRI (CD64), FcγRII (CD32), and FcγRIII (CD16). The interaction between the antigen-binding protein bound to the antigen and the formation of the Fc / Fcγ complex can induce a series of effects, including cytotoxicity, immune cell activation, phagocytosis, and / or the release of inflammatory cytokines.

[0033] In another embodiment, the anti-BCMA antibody can inhibit the binding of BAFF and / or APRIL to the BCMA receptor. In another embodiment, the anti-BCMA antibody may be able to bind to FcγRIIIA or may have FcγRIIIA-mediated effector function.

[0034] In one embodiment, the composition comprises an anti-BCMA antibody comprising two immunoglobulin (Ig) heavy chains ("HC") and two Ig light chains ("LC"). The basic antibody structural unit can include, for example, a tetramer of subunits. Each tetramer can include two pairs of polypeptide chains, each pair having one "light" chain (about 25 kDa) and one "heavy" chain (about 50-70 kDa). The amino-terminal portion of each chain can include a variable region of about 100 to 110 or more amino acids that is primarily responsible for antigen recognition. The variable region can initially be expressed linked to a cleavable signal peptide. The variable region without the signal peptide can be referred to as the mature variable region. Thus, in one example, the light chain mature variable region can comprise the light chain variable region without the light chain signal peptide. The carboxyl-terminal portion of each chain can define the constant region. The heavy chain constant region is primarily responsible for effector function.

[0035] The mature variable regions of each light / heavy chain pair can form an antibody binding site (also referred to as an antigen binding site). An "antigen binding site" refers to the site on the antibody that is capable of specifically binding an antigen, which can be a single variable domain, or it can be the paired V H / V LDomain. Thus, a complete antibody can have, for example, two binding sites. Except in bispecific or bifunctional antibodies, the two binding sites can be the same. Each of these chains can exhibit the same general structure of relatively conserved framework regions (FRs) joined by three hypervariable regions (also called complementarity-determining regions or "CDRs"). The CDRs from two chains of each pair can be aligned by the framework regions and be capable of binding to a specific epitope. Thus, in one example, from the N-terminus to the C-terminus, both the light chain and the heavy chain contain the domains FR1, CDR1, FR2, CDR2, FR3, CDR3, and FR4.

[0036] Define "CDR" as the amino acid sequence of the complementarity-determining region of an antibody. These are the hypervariable regions of the immunoglobulin heavy and light chains. There are three heavy chain and three light chain CDRs (or CDR regions) in the variable portion of the immunoglobulin. Thus, as used herein, "CDR" refers to all three heavy chain CDRs, all three light chain CDRs, all heavy and light chain CDRs, or at least two CDRs. In one embodiment, the composition comprises an anti-BCMA antibody that comprises one or more CDRs according to the present invention as described herein, or one or both of the heavy chain or light chain variable domains according to the present invention as described herein.

[0037] The terms "variant", "antibody variant", "CDR variant", and "post-translational modification variant" refer to at least one amino acid change in an antibody sequence. Variants can be the result of post-translational modifications, chemical changes, or sequence changes by at least one deletion, substitution, or addition. Some post-translational modifications result in chemical changes that do not alter the sequence (e.g., Met and oxidized Met; or Asp and isomerization / isoAsp; or aggregation), while others result in sequence changes, such as the conversion of one amino acid residue to another (e.g., conversion of Asn to Asp by deamidation; or deletion of lysine). Further post-translational modification variants are described below. Variant antibody sequences containing sequence changes can be the result of designed sequence changes or post-translational modifications. Amino acid sequence changes can be deletions, substitutions, or additions.

[0038] In one such embodiment, the substitution is a conservative substitution. In an alternative embodiment, the antibody variant comprises at least one substitution while retaining the canonical of the antigen-binding protein. In one embodiment, the antibody variant is an antibody that is at least about 80%, about 85%, about 90%, or about 95% identical (i.e., has sequence identity) to the antibody primary sequence. In another embodiment, the antibody variant includes an antibody that comprises a heavy chain amino acid sequence that is at least about 80%, about 85%, about 90%, or about 95% identical to the amino acid sequence of SEQ ID NO: 9, and / or a heavy chain amino acid sequence that is at least about 80%, about 85%, about 90%, or about 95% identical to the amino acid sequence of SEQ ID NO: 10.

[0039] The antigen-binding proteins of the present invention may have amino acid modifications that increase the affinity of the constant domain or a fragment thereof for FcRn. Increasing the half-life (i.e., serum half-life) of therapeutic and diagnostic IgG antibodies and other bioactive molecules has many benefits, including reducing the amount and / or frequency of administration of these molecules. In one embodiment, the antigen-binding proteins of the present invention comprise all or part (the FcRn-binding portion) of an IgG constant domain having one or more of the following amino acid modifications.

[0040] For example, with respect to IgG1, M252Y / S254T / T256E (commonly referred to as the "YTE" mutation) and M428L / N434S (commonly referred to as the "LS" mutation) increase FcRn binding at pH 6.0 (Wang et al. 2018).

[0041] The T250Q / M428L, V259I / V308F / M428L, N434A, and T307A / E380A / N434A mutations (referring to IgG1 and Kabat numbering) can also increase the half-life (Monnet et al.).

[0042] The half-life and FcRn binding can also be extended by introducing the H433K and N434F mutations (commonly referred to as the "HN" or "NHance" mutations) (with respect to IgG1) (WO2006 / 130834).

[0043] WO00 / 42072 discloses polypeptides comprising a variant Fc region having an altered FcRn binding affinity, the polypeptide comprising an amino acid modification at any one or more of amino acid positions 238, 252, 253, 254, 255, 256, 265, 272, 286, 288, 303, 305, 307, 309, 311, 312, 317, 340, 356, 360, 362, 376, 378, 380, 386, 388, 400, 413, 415, 424, 433, 434, 435, 436, 439, and 447 in the Fc region (EU index numbering).

[0044] WO02 / 060919 discloses modified IgG comprising an IgG constant domain having one or more amino acid modifications relative to the wild-type IgG constant domain, wherein the half-life of the modified IgG is increased relative to the half-life of an IgG having a wild-type IgG constant domain, and wherein said one or more amino acid modifications are located at one or more positions in 251, 253, 255, 285 - 290, 308 - 314, 385 - 389, and 428 - 435.

[0045] Shields et al. (2001, J Biol Chem; 276:6591-604) used alanine-scanning mutagenesis to alter residues in the Fc region of a human IgG1 antibody and then evaluated binding to human FcRn. Positions that effectively eliminated binding to FcRn when changed to alanine included I253, S254, H435, and Y436. Other positions showed less obvious binding reduction as follows: E233-G236, R255, K288, L309, S415, and H433. When changed to alanine, several amino acid positions exhibited improved FcRn binding; notably among these were P238, T256, E272, V305, T307, Q311, D312, K317, D376, E380, E382, S424, and N434. Many other amino acid positions exhibited a slight improvement in binding to FcRn (D265, N286, V303, K360, Q362, and A378) or no change (S239, K246, K248, D249, M252, E258, T260, S267, H268, S269, D270, K274, N276, Y278, D280, V282, E283, H285, T289, K290, R292, E293, E294, Q295, Y296, N297, S298, R301, N315, E318, K320, K322, S324, K326, A327, P329, P331, E333, K334, T335, S337, K338, K340, Q342, R344, E345, Q345, Q347, R356, M358, T359, K360, N361, Y373, S375, S383, N384, Q386, E388, N389, N390, K392, L398, S400, D401, K414, R416, Q418, Q419, N421, V422, E430, T437, K439, S440, S442, S444, and K447).

[0046] For combinatorial variants, the most dramatic effects on improved FcRn binding were found. At pH 6.0, the E380A / N434A variant bound to FcRn more than 8-fold better than native IgG1, 2-fold better than E380A, and 3.5-fold better than N434A. Adding T307A to it resulted in a 12-fold increase in binding relative to native IgG1. In one embodiment, the antigen-binding protein of the present invention comprises the E380A / N434A mutation and has increased binding to FcRn.

[0047] Dall’Acqua et al. (2002, J Immunol.;169:5171-80) described the random mutagenesis and screening of a human IgG1 hinge-Fc fragment phage display library against murine FcRn. They reported random mutagenesis at positions 251, 252, 254-256, 308, 309, 311, 312, 314, 385-387, 389, 428, 433, 434, and 436. The major improvement in IgG1-human FcRn complex stability occurred upon substitution of residues located in the band across the Fc-FcRn interface (M252, S254, T256, H433, N434, and Y436), and to a lesser extent of residues in the periphery, such as V308, L309, Q311, G385, Q386, P387, and N389. The variant with the highest affinity for human FcRn was obtained by combining the M252Y / S254T / T256E (“YTE”) and H433K / N434F / Y436H mutations and showed a 57-fold increase in affinity compared to wild-type IgG1. The in vivo behavior of such mutated human IgG1 showed an almost 4-fold increase in serum half-life in cynomolgus monkeys compared to wild-type IgG1.

[0048] Accordingly, the present invention provides antigen-binding proteins with optimized binding to FcRn. In a preferred embodiment, the antigen-binding protein comprises at least one amino acid modification in the Fc region of the antigen-binding protein, wherein the modification is at an amino acid position selected from the group consisting of 226, 227, 228, 230, 231, 233, 234, 239, 241, 243, 246, 250, 252, 256, 259, 264, 265, 267, 269, 270, 276, 284, 285, 288, 289, 290, 291, 292, 294, 297, 298, 299, 301, 302, 303, 305, 307, 308, 309, 311, 315, 317, 320, 322, 325, 327, 330, 332, 334, 335, 338, 340, 342, 343, 345, 347, 350, 352, 354, 355, 356, 359, 360, 361, 362, 369, 370, 371, 375, 378, 380, 382, 384, 385, 386, 387, 389, 390, 392, 393, 394, 395, 396, 397, 398, 399, 400, 401, 403, 404, 408, 411, 412, 414, 415, 416, 418, 419, 420, 421, 422, 424, 426, 428, 433, 434, 438, 439, 440, 443, 444, 445, 446 and 447 of the Fc region.

[0049] In addition, various publications describe methods for obtaining bioactive molecules with altered half-lives by introducing polypeptides that bind FcRn into the molecule (WO97 / 43316, US5869046, US5747035, WO96 / 32478 and WO91 / 14438), or by fusing the molecule to an antibody whose FcRn binding affinity is retained but whose affinity for other Fc receptors has been greatly reduced (WO99 / 43713), or by fusing to the FcRn binding domain of an antibody (WO00 / 09560, US4703039).

[0050] Fc variants with enhanced FcRn affinity were identified in screening at pH 6.0 to improve the cytotoxicity and half-life of antibodies. The selected IgG variants can be produced as low-fucosylated molecules. The resulting variants showed increased serum persistence in hFcRn mice, as well as conserved enhanced ADCC (Monnet et al.). Exemplary variants include (with respect to IgG1 and Kabat numbering):

[0051] P230T / V303A / K322R / N389T / F404L / N434S;

[0052] P228R / N434S;

[0053] Q311R / K334R / Q342E / N434Y;

[0054] C226G / Q386R / N434Y;

[0055] T307P / N389T / N434Y;

[0056] P230S / N434S;

[0057] P230T / V305A / T307A / A378V / L398P / N434S;

[0058] P23OT / P387S / N434S;

[0059] P230Q / E269D / N434S;

[0060] N276S / A378V / N434S;

[0061] T307A / N315D / A330V / 382V / N389T / N434Y;

[0062] T256N / A378V / S383N / N434Y;

[0063] N315D / A330V / N361D / A387V / N434Y;

[0064] V259I / N315D / M428L / N434Y;

[0065] P230S / N315D / M428L / N434Y;

[0066] F241L / V264E / T307P / A378V / H433R;

[0067] T250A / N389K / N434Y;

[0068] V305A / N315D / A330V / P395A / N434Y;

[0069] V264E / Q386R / P396L / N434S / K439R;

[0070] E294del / T307P / N434Y (where "del" indicates deletion).

[0071] The present invention also provides a method for producing an antigen-binding protein according to the present invention, which comprises the following steps: a) culturing a recombinant host cell comprising an expression vector containing the isolated nucleic acid as described herein, wherein the FUT8 gene encoding α-1,6-fucosyltransferase has been inactivated in the recombinant host cell; and b) recovering the antigen-binding protein. For example, such a method for producing an antigen-binding protein can be carried out using the POTELLIGENT technology system available from BioWa, Inc. (Princeton, NJ), wherein CHOK1SV cells lacking a functional copy of the FUT8 gene produce antibodies with enhanced monoclonal antibody-dependent cell-mediated cytotoxicity (ADCC) activity, which is increased relative to the same monoclonal antibody produced in cells having a functional FUT8 gene. Aspects of the POTELLIGENT technology system are described in US7214775, US6946292, WO0061739 and WO0231240, all of which are incorporated herein by reference. Those of ordinary skill in the art will also recognize other suitable systems and methods for generating antigen-binding proteins such as antibodies.

[0072] Antibodies can be recovered and purified by conventional protein purification procedures. For example, antibodies can be harvested directly from the culture medium. The cell culture medium can be harvested by clarification, for example, by centrifugation and / or depth filtration. The antibody is recovered and then purified to ensure sufficient purity. Thus, in one aspect, a cell culture medium containing the antibody as described herein is provided. In one embodiment, the cell culture medium contains CHO cells.

[0073] Subsequently, the antibody can be purified from the cell culture medium. This can include harvesting the cell culture supernatant, contacting the cell culture supernatant with a purification medium (e.g., protein A resin or protein G resin to bind the antibody molecule), and eluting the antibody molecule from the purification medium to produce an eluate. Thus, in one aspect, an eluate containing the antibody as described herein is provided.

[0074] One or more chromatographic steps can be used for purification, such as one or more chromatographic resins; and / or one or more filtration steps. For example, affinity chromatography using resins such as protein A, G or L can be used to purify the composition. Alternatively or in addition, ion exchange resins such as cation exchange resins can be used to purify the composition.

[0075] Alternatively, the purification step comprises: an affinity chromatography resin step followed by a cation exchange resin step.

[0076] In one embodiment, the anti-BCMA antibody comprises a heavy chain variable region CDR1 ("CDRH1") that comprises an amino acid sequence having at least about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the amino acid sequence set forth in SEQ ID NO: 1. In one embodiment, the heavy chain variable region CDR1 ("CDRH1") comprises an amino acid sequence having one amino acid variation ("variant") from the amino acid sequence set forth in SEQ ID NO: 1.

[0077] In one embodiment, the anti-BCMA antibody comprises a heavy chain variable region CDR2 ("CDRH2") that comprises an amino acid sequence having at least about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the amino acid sequence set forth in SEQ ID NO: 2. In one embodiment, the heavy chain variable region CDR2 ("CDRH2") comprises an amino acid sequence having one amino acid variation ("variant") from the amino acid sequence set forth in SEQ ID NO: 2.

[0078] In one embodiment, the anti-BCMA antibody comprises a heavy chain variable region CDR3 ("CDRH3") that comprises an amino acid sequence having at least about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the amino acid sequence set forth in SEQ ID NO: 3. In one embodiment, the heavy chain variable region CDR3 ("CDRH3") comprises an amino acid sequence having one amino acid variation ("variant") from the amino acid sequence set forth in SEQ ID NO: 3.

[0079] In one embodiment, the anti-BCMA antibody comprises a light chain variable region CDR1 ("CDRL1") that comprises an amino acid sequence having at least about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the amino acid sequence set forth in SEQ ID NO: 4. In one embodiment, the light chain variable region CDL1 ("CDR1") comprises an amino acid sequence having one amino acid variation ("variant") from the amino acid sequence set forth in SEQ ID NO: 4.

[0080] In one embodiment, the anti-BCMA antibody comprises a light chain variable region CDR2 ("CDRL2") that comprises an amino acid sequence having at least about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the amino acid sequence set forth in SEQ ID NO: 5. In one embodiment, the light chain variable region CDL2 ("CDR2") comprises an amino acid sequence having one amino acid variation ("variant") from the amino acid sequence set forth in SEQ ID NO: 5.

[0081] In one embodiment, the anti-BCMA antibody comprises a light chain variable region CDR3 ("CDRL3") that comprises an amino acid sequence having at least about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the amino acid sequence set forth in SEQ ID NO: 6. In one embodiment, the light chain variable region CDL3 ("CDR3") comprises an amino acid sequence having one amino acid variation ("variant") from the amino acid sequence set forth in SEQ ID NO: 6.

[0082] In one embodiment, the anti-BCMA antibody comprises CDRH1 that comprises an amino acid sequence that is at least about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence set forth in SEQ ID NO: 1; CDRH2 that comprises an amino acid sequence that is at least about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence set forth in SEQ ID NO: 2; CDRH3 that comprises an amino acid sequence that is at least about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence set forth in SEQ ID NO: 3; CDRL1 that comprises an amino acid sequence that is at least about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence set forth in SEQ ID NO: 4; CDRL2 that comprises an amino acid sequence that is at least about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence set forth in SEQ ID NO: 5; and / or CDRL3 that comprises an amino acid sequence that is at least about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence set forth in SEQ ID NO: 6.

[0083] In one embodiment, the anti-BCMA antibody comprises a heavy chain variable region (“V H ”), which comprises an amino acid sequence having at least about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the amino acid sequence set forth in SEQ ID NO: 7.

[0084] In one embodiment, the anti-BCMA antibody comprises a light chain variable region (“V L ”), which comprises an amino acid sequence having at least about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the amino acid sequence set forth in SEQ ID NO: 8.

[0085] In one embodiment, the anti-BCMA antibody comprises V H , which comprises an amino acid sequence having at least about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the amino acid sequence set forth in SEQ ID NO: 7; and V L , which comprises an amino acid sequence having at least about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the amino acid sequence set forth in SEQ ID NO: 8.

[0086] In one embodiment, the anti-BCMA antibody comprises a heavy chain variable region (“HC”), which comprises an amino acid sequence having at least about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the amino acid sequence set forth in SEQ ID NO: 9.

[0087] In one embodiment, the anti-BCMA antibody comprises a light chain variable region (“LC”), which comprises an amino acid sequence having at least about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the amino acid sequence set forth in SEQ ID NO: 10.

[0088] In one embodiment, the anti-BCMA antibody comprises an HC that comprises an amino acid sequence having at least about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the amino acid sequence set forth in SEQ ID NO: 9; and an LC that comprises an amino acid sequence having at least about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the amino acid sequence set forth in SEQ ID NO: 10.

[0089] The "percent identity" between a query amino acid sequence and a subject amino acid sequence is the "identity" value, expressed as a percentage, calculated by the BLASTP algorithm after a pairwise BLASTP alignment when the subject amino acid sequence has 100% query coverage with the query amino acid. Such a pairwise BLASTP alignment between the query amino acid sequence and the subject amino acid sequence is performed using the default settings of the BLASTP algorithm available on the National Center for Biotechnology Information website, with the filter for low complexity regions turned off. Importantly, the query sequence can be described by the amino acid sequence determined in one or more claims herein.

[0090] In one embodiment, the anti-BCMA antibody comprises a CDRH1 having the amino acid sequence set forth in SEQ ID NO: 1; a CDRH2 having the amino acid sequence set forth in SEQ ID NO: 2; a CDRH3 having the amino acid sequence set forth in SEQ ID NO: 3; a CDRL1 having the amino acid sequence set forth in SEQ ID NO: 4; a CDRL2 having the amino acid sequence set forth in SEQ ID NO: 5; and a CDRL3 having the amino acid sequence set forth in SEQ ID NO: 6.

[0091] In one embodiment, the anti-BCMA antibody comprises a V having the amino acid sequence set forth in SEQ ID NO: 7 H ; and a V having the amino acid sequence set forth in SEQ ID NO: 8 L .

[0092] In one embodiment, the anti-BCMA antibody comprises belantamab, which comprises an HC having the amino acid sequence set forth in SEQ ID NO: 9 and an LC having the amino acid sequence set forth in SEQ ID NO: 10.

[0093] The sequences of antibodies can be determined by the Kabat numbering system (Kabat et al. Sequences of proteins of Immunological Interest NIH, 1987). Alternatively, they can be determined using the Chothia numbering system (Al-Lazikani et al., (1997) JMB 273, 927-948), the contact definition method (MacCallum R.M., and Martin A.C.R. and Thornton J.M, (1996), Journal of Molecular Biology, 262(5), 732-745) or any other established method known to those skilled in the art for numbering residues in antibodies and determining CDRs. Other numbering conventions for antibody sequences available to those skilled in the art include the "AbM" (University of Bath) and "Contact" (University College London) methods. Finally, antibody sequences can be numbered sequentially.

[0094] When making numerical references to the amino acids described herein, the sequences can be numbered according to the Kabat method or the sequential numbering method. Unless otherwise expressly stated, numerical references to specific amino acid numbers are described herein using the sequential numbering system. Throughout the specification, the terms "CDR", "CDRL1", "CDRL2", "CDRL3", "CDRH1", "CDRH2", "CDRH3" follow the Kabat numbering. Amino acid residues in the variable region sequences and full-length antibody sequences are numbered sequentially to denote any antibody sequence variant positions or post-translational modification variant positions, such as isomerization variants (e.g., D103), deamidation variants (e.g., N388) or oxidation variants (e.g., M34).

[0095] References to positions in the CDRs (e.g., M34 or D103) provide the position numbering (sequential numbering) relative to the entire antibody sequence. Thus, it should be understood that M34 of CDRH1 refers to the fourth residue of SEQ ID NO: 1, i.e., underlined as: NYW M H (SEQ ID NO: 1). Similarly, D103 of CDRH3 refers to the fifth residue of SEQ ID NO: 3, i.e., underlined as: GAIY D GYDV L DN (SEQ ID NO: 3).

[0096] In one aspect, the composition comprises an antibody variant that comprises a change in one or more amino acids in the primary sequence. In one embodiment, the composition comprises an antibody that is at least about 90% identical to the heavy chain amino acid sequence of SEQ ID NO: 9 and / or the light chain sequence of SEQ ID NO: 10, with an amino acid change from aspartic acid (D) to asparagine (N), such as D103N (i.e., D99N in Kabat numbering) in CDRH3.

[0097] In another embodiment, the composition comprises an antibody that comprises CDRH1 having the amino acid sequence set forth in SEQ ID NO: 1, CDRH2 having the amino acid sequence set forth in SEQ ID NO: 2, CDRH3 having the amino acid sequence set forth in SEQ ID NO: 3, CDRL1 having the amino acid sequence set forth in SEQ ID NO: 4, CDRL2 having the amino acid sequence set forth in SEQ ID NO: 5, CDRL3 having the amino acid sequence set forth in SEQ ID NO: 6, and comprises an amino acid change from aspartic acid (D) to asparagine (N), such as D103N in CDRH3.

[0098] In another embodiment, the anti-BCMA antibody comprises belantamab and comprises an amino acid change from aspartic acid (D) to asparagine (N), such as D103N in CDRH3.

[0099] In one embodiment, the composition comprises a mixture of antibodies that is at least about 90% identical to the heavy chain amino acid sequence of SEQ ID NO: 9 and / or the light chain sequence of SEQ ID NO: 10, wherein about ≥5%, ≥10%, ≥15%, ≥20%, ≥25%, ≥50%, ≥75% or ≥90% of the antibodies comprise D103N in CDRH3.

[0100] In one embodiment, the composition comprises a mixture of antibodies that comprises CDRH1 having the amino acid sequence set forth in SEQ ID NO: 1, CDRH2 having the amino acid sequence set forth in SEQ ID NO: 2, CDRH3 having the amino acid sequence set forth in SEQ ID NO: 3, CDRL1 having the amino acid sequence set forth in SEQ ID NO: 4, CDRL2 having the amino acid sequence set forth in SEQ ID NO: 5, CDRL3 having the amino acid sequence set forth in SEQ ID NO: 6, wherein about ≥5%, ≥10%, ≥15%, ≥20%, ≥25%, ≥50%, ≥75% or ≥90% of the antibodies in the mixture comprise D103N in CDRH3.

[0101] In one embodiment, the composition comprises belantamab, wherein about ≥5%, ≥10%, ≥15%, ≥20%, ≥25%, ≥50%, ≥75% or ≥90% of the belantamab comprises D103N in CDRH3.

[0102] In one embodiment, the composition comprises belantamab, which comprises at least one antibody variant selected from the group consisting of: G27Y, S30T, A93T, A24G, K73T, M48I, V67A, F71Y, D99N, M4L and K45E using the Kabat numbering system.

[0103] Post-translational modification product

[0104] The "post-translational modification product" of the antibody described herein is an antibody composition, wherein all or part of the composition comprises "post-translational modification". Post-translational modification is a chemical change of the antibody, which may be due to the production of the antibody in a host cell, upstream and downstream preparation and / or storage (e.g., exposure to light, temperature, pH, water or by reaction with excipients and / or direct container closure systems). Thus, the compositions of the present invention can be formed by the preparation or storage of the antibody. Exemplary post-translational modifications include antibody sequence changes ("antibody variants", as described above), cleavage of certain leader sequences, addition of various sugar moieties in various glycosylation patterns, non-enzymatic glycosylation, deamidation, oxidation, disulfide scrambling and other cysteine variants, such as free thiols, racemic disulfides, thioethers and trisulfides, isomerization, C-terminal lysine cleavage and / or N-terminal glutamine cyclization.

[0105] In one example, the post-translational modification product comprises "product-related impurities", which comprise chemical changes that result in a decrease in function and / or activity. In another example, the post-translational modification product comprises "product-related substances", which comprise chemical changes that do not result in a decrease in function and / or activity. Product-related impurities of the antibody described herein include isomerization variants and oxidation variants. Product-related substances of the antibody described herein include deamidation variants, glycosylation variants, C-terminal cleavage variants and N-terminal pyroglutamate variants.

[0106] In one embodiment, the composition comprises the heavy chain sequence of SEQ ID NO: 9 and the light chain sequence of SEQ ID NO: 10, which comprise one or more functional post-translational modifications. In another embodiment, the composition comprises the heavy chain sequence of SEQ ID NO: 11, SEQ ID NO: 12, SEQ ID NO: 13 or SEQ ID NO: 14, and the light chain of SEQ ID NO: 10, which comprise one or more functional post-translational modifications.

[0107] The percentage of variants provided herein is expressed as a percentage of the total amount of antibody in the composition (e.g., the antibody “pool”). For example, 40% or less oxidized variants means that, in a composition where the total amount of antibody is 100%, 40% or less is oxidized. For example, 25% or less isomerized variants means that, in a composition where the total amount of antibody is 100%, 25% or less is isomerized.

[0108] Glycation is a post-translational modification that involves a non-enzymatic chemical reaction between a reducing sugar (such as glucose) and a free amino group in a protein, and is typically observed at the ε-amino of a lysine side chain or the N-terminus of the protein. In the presence of a reducing sugar, glycation can occur during production and / or storage.

[0109] Deamidation, which can occur, for example, during production and / or storage, can be an enzymatic or chemical reaction. Deamidation can occur by intramolecular cyclization through a simple chemical reaction in which the amide nitrogen of the next amino acid in the chain nucleophilically attacks the amide (N+1 attacks N); a succinimide intermediate is formed. Deamidation can convert asparagine (N) to isoaspartic acid (isoaspartate) and aspartic acid (aspartate) (D) mainly at a ratio of 3:1. Thus, this deamidation reaction may be related to the isomerization of aspartate (D) to isoaspartate. Both deamidation of asparagine and isomerization of aspartate may involve the intermediate succinimide. Glutamine residues can be deamidated in a similar manner, but to a much lesser extent. Deamidation can occur in the CDR, Fab (non-CDR region), or Fc region. Isomerization is the conversion of aspartate (D) to isoaspartate, which involves the intermediate succinimide.

[0110] Oxidation can occur during production and / or storage (i.e., in the presence of oxidative conditions) and results in covalent modification of the protein directly induced by reactive oxygen species or indirectly induced by reaction with secondary by-products of oxidative stress. Oxidation may occur mainly on methionine residues, but can also occur on tryptophan and free cysteine residues. Oxidation can occur in the CDR, Fab (non-CDR) region, or Fc region.

[0111] Disulfide scrambling can occur under production and / or storage conditions. In some cases, disulfide bonds may break or misform, resulting in unpaired cysteine residues (-SH). These free (unpaired) thiol groups (-SH) may facilitate shuffling.

[0112] Via β -elimination of the disulfide bridge back to cysteine residues via dehydroalanine and persulfide intermediates, formation of thioethers and racemization of disulfide bonds can occur under alkaline conditions during production or storage. Subsequent crosslinking of dehydroalanine and cysteine may result in the formation of thioether bonds, or free cysteine residues may reform disulfide bonds with a mixture of D - and L -cysteine.

[0113] Trisulfides may result from the insertion of a sulfur atom into a disulfide bond (Cys - S - S - S - Cys) and may form due to the presence of hydrogen sulfide in the production cell culture.

[0114] The N - terminal glutamine (Q) and glutamate (glutamic acid) (E) in the heavy and / or light chains can cyclize to form pyroglutamic acid (pGlu). pGlu formation can occur in the production bioreactor, but can also form non - enzymatically, for example, depending on the pH and temperature of the processing and storage conditions. Cyclization of the N - terminal Q or E is commonly observed in native human antibodies.

[0115] C - terminal lysine cleavage is an enzymatic reaction catalyzed by carboxypeptidase and is commonly observed in recombinant and native human antibodies. Variants of this processing include removal of lysine from one or both heavy chains due to cellular enzymes from the recombinant host cell. Administration to human subjects / patients may result in the removal of any remaining C - terminal lysine.

[0116] The present invention encompasses antibodies that may have undergone or experienced one or more of the post - translational modifications described herein. Exemplary compositions can comprise a mixture or blend of antibodies that: 1) have and do not have post - translational modification(s), or 2) have more than one type of post - translational modification described herein.

[0117] The composition can comprise a mixture of antibody variants and post - translational modification variants. For example, an antibody composition can comprise one or more, such as two or more, oxidized variants, deamidated variants, isomerized variants, N - terminal pyroglutamic acid variants, and C - terminal lysine cleavage variants.

[0118] For example, in one embodiment, the composition can comprise a mixture of antibodies, where 10% of the antibodies in the mixture comprise the amino acid sequences of SEQ ID NOs 9 and 10, and 90% of the antibodies in the mixture comprise the amino acid sequences of SEQ ID NOs 9 and 10 with C - terminal lysine cleavage.

[0119] In another exemplary embodiment, the composition can comprise a mixture of antibodies, wherein 10% of the antibodies in the mixture comprise the amino acid sequences of SEQ ID NOs 9 and 10, 90% of the antibodies in the mixture comprise the amino acid sequences of SEQ ID NOs 9 and 10 with C-terminal lysine cleavage, and in the 100% total antibody mixture, up to 100% of the N-terminal glutamine is cyclized to pyroglutamic acid.

[0120] In another exemplary embodiment, the composition can comprise a mixture of antibodies, wherein 10% of the antibodies in the mixture comprise the amino acid sequences of SEQ ID NOs 9 and 10, 90% of the antibodies in the mixture comprise the amino acid sequences of SEQ ID NOs 9 and 10 with C-terminal lysine cleavage, and in the 100% total antibody mixture, up to 100% is N-terminal pyroglutamic acid and up to 23% is isomerized at D103 of CDRH3.

[0121] In yet another exemplary embodiment, the composition comprises a mixture of antibodies, wherein 20% of the antibodies in the mixture comprise the amino acid sequences of SEQ ID NOs 9 and 10, 80% of the antibodies in the mixture comprise the amino acid sequences of SEQ ID NOs 9 and 10 having variant N103 in CDRH3, and in the 100% total antibody mixture, up to 37% of the antibodies are oxidized at amino acid M34 in CDRH1.

[0122] In one embodiment, the post-translational modifications described herein do not result in significant changes in antigen-binding affinity, biological activity, pharmacokinetics (PK) / pharmacodynamics (PD), aggregation, immunogenicity, and / or binding to Fc receptors, unless specified and described as impurities associated with the product.

[0123] As described herein, "function" or "activity" is defined as one or more of the following: 1) binding to BCMA, 2) binding to FcγRIIIa, and / or 3) binding to FcRn. In one embodiment, "reduced function" or "reduced activity" means that the binding to BCMA, the binding to FcγRIIIa, or the binding to FcRn is reduced by a percentage compared to a reference standard and is significant in the assay variability. For example, a reduction in function or activity can be described as a reduction of ≥5%, ≥10%, ≥15%, ≥20%, ≥25%, ≥30%, ≥35%, ≥40%, ≥45%, or ≥50%.

[0124] In one embodiment, the anti-BCMA antibody comprises an antibody that is at least about 90% identical to the amino acid sequences of SEQ ID NO: 9 and SEQ ID NO: 10 and includes all post-translational modifications (if any) of the antibody.

[0125] In another embodiment, the anti-BCMA antibody comprises belantamab and all post-translational modifications, if any.

[0126] Antibody variants are commonly observed when analyzing the composition of an antibody by charge-based separation techniques such as isoelectric focusing (IEF) gel electrophoresis, capillary isoelectric focusing (cIEF) gel electrophoresis, cation exchange chromatography (CEX), and anion exchange chromatography (AEX).

[0127] Post-translational modifications can result in an increase or decrease in the net charge of the antibody and a decrease or increase in the pI value, resulting in acidic and basic variants (collectively referred to as "charged variants") relative to the major isoform. The major isoform is the population of antibodies that elute as the main peak on a chromatogram. When analyzing an antibody using an IEF-based method, acidic species are variants with a lower apparent pI, and basic species are variants with a higher apparent pI. When analyzed by a chromatography-based method, acidic and basic species are defined based on their retention time relative to the main peak. Acidic species are variants that elute earlier than the CEX main peak or later than the AEX main peak, while basic species are variants that elute later than the CEX main peak or earlier than the AEX main peak. These methods separate the major isoform of the antibody from acidic isoforms (acidic variants) and basic isoforms (basic variants). Charged variants can be detected by various methods such as ion exchange chromatography, e.g., WCX-10 HPLC (weak cation exchange chromatography) or IEF (isoelectric focusing). The percentage of charged species can be determined using capillary isoelectric focusing (cIEF). Capillary isoelectric focusing (cIEF) is used to measure the pI of dostarlimab and the separated charge variants (see Figure 1 ). This method can be used to quantify the percentage of acidic and basic species in the total peak area. The terms "species", "isoform", "form", and "peak" are used interchangeably to refer to the major isoform and charged variants (acidic variants and basic variants).

[0128] In one embodiment, the composition comprises an acidic variant of the antibody, wherein the acidic variant comprises CDRH1 of SEQ ID NO:1, CDRH2 of SEQ ID NO:2, and CDRH3 of SEQ ID NO:3, and a light chain amino acid sequence comprising CDRL1 of SEQ ID NO:4, CDRL2 of SEQ ID NO:5, and CDRL3 of SEQ ID NO:6; wherein the composition comprises 1-70% of the acidic variant.

[0129] On the one hand, the composition comprises ≤70% of the acidic variant. In one embodiment, the composition comprises ≤60%, ≤50%, ≤40%, ≤35% or ≤30% of the acidic variant. Alternatively, the composition comprises 10 - 70%, 10 - 60%, 10 - 50%, 10 - 40%, 10 - 35% or 10 - 30% of the acidic variant. Alternatively, the composition comprises 20 - 70%, 20 - 60%, 20 - 50%, 20 - 40%, 20 - 35% or 20 - 30% of the acidic variant. Alternatively, the composition comprises approximately 60%, approximately 50%, approximately 40%, approximately 35%, approximately 30%, approximately 25% or approximately 20% of the acidic variant.

[0130] On the one hand, the composition comprises a basic variant of the antibody, wherein the basic variant comprises a heavy chain amino acid sequence comprising CDRH1 of SEQ ID NO: 1, CDRH2 of SEQ ID NO: 2 and CDRH3 of SEQ ID NO: 3, and a light chain amino acid sequence comprising CDRL1 of SEQ ID NO: 4, CDRL2 of SEQ ID NO: 5 and CDRL3 of SEQ ID NO: 6; wherein the composition comprises 1 - 30% of the basic variant.

[0131] On the one hand, the composition comprises ≤30% of the basic variant. In one embodiment, the composition comprises ≤25%, ≤20%, ≤15%, ≤10%, ≤7.5% or ≤5% of the basic variant. In one embodiment, the composition comprises 1 - 30%, 1 - 25%, 1 - 20%, 1 - 15%, 1 - 10% or 1 - 5% of the basic variant. Alternatively, the composition comprises approximately 15%, approximately 10% or approximately 5% of the basic variant.

[0132] In one aspect, the composition comprises the major isotype of the antibody, wherein the major isotype comprises a heavy chain amino acid sequence comprising CDRH1 of SEQ ID NO: 1, CDRH2 of SEQ ID NO: 2 and CDRH3 of SEQ ID NO: 3, and a light chain amino acid sequence comprising CDRL1 of SEQ ID NO: 4, CDRL2 of SEQ ID NO: 5 and CDRL3 of SEQ ID NO: 6; wherein the composition comprises 1 - 90% of the major isomer.

[0133] In one aspect, the composition comprises ≥1% of the major isoform. In one embodiment, the composition comprises ≥5%, ≥10%, ≥20%, ≥30%, ≥40%, ≥50%, ≥55%, ≥60%, ≥65%, ≥70%, ≥75%, ≥80% or ≥90% of the major isoform. In one embodiment, the composition comprises 10 - 90%, 20 - 90%, 30 - 90%, 40 - 90%, 50 - 90% or 60 - 90% of the major isoform. In one embodiment, the composition comprises 10 - 80%, 20 - 80%, 30 - 80%, 40 - 80%, 50 - 80% or 60 - 80% of the major isoform. Alternatively, the composition comprises approximately 80%, approximately 75%, approximately 70%, approximately 65%, approximately 60%, approximately 50% or approximately 55% of the major isoform.

[0134] Capillary isoelectric focusing (cIEF) can be used to determine the percentage of acidic variants, the percentage of basic variants, and the percentage of the major isomer. It should be understood that these isoform / charge variant embodiments can be combined with any one or combination of the antibody variants described herein.

[0135] In one aspect, the composition comprises a charge variant of an antibody, which comprises a heavy chain amino acid sequence comprising CDRH1 of SEQ ID NO: 1, CDRH2 of SEQ ID NO: 2, and CDRH3 of SEQ ID NO: 3, and a light chain amino acid sequence comprising CDRL1 of SEQ ID NO: 4, CDRL2 of SEQ ID NO: 5, and CDRL3 of SEQ ID NO: 6; wherein the composition comprises: ≤70% of acidic variants; and / or ≤30% of basic variants; and / or ≥1% of the major isomer.

[0136] In one aspect, the composition comprises an antibody that comprises an isomerized post-translational modification (“isomerized” or “isomerization”) or “isomerized variant”. The variant may comprise isomerized amino acid residues in the heavy chain sequence and / or the light chain sequence, such as in the CDRs of the heavy chain sequence and / or the CDRs of the light chain sequence. The isomerized variant may be present in one or both of the heavy chain and the light chain. The isomerized post-translational modification results in isoaspartate and / or succinimide-aspartate. In one embodiment, aspartate (Asp) isomerization can be determined using tryptic peptide mapping tandem mass spectrometry (peptide mapping LC-MS / MS) as described herein. It will be understood that these isomerized variant embodiments can be combined with the antibody characteristics described herein.

[0137] In one embodiment, the composition comprises an isomerization variant of an anti-BCMA antibody, wherein the isomerization variant comprises a heavy chain amino acid sequence comprising CDRH1 of SEQ ID NO: 1, CDRH2 of SEQ ID NO: 2, and CDRH3 of SEQ ID NO: 3, and a light chain amino acid sequence comprising CDRL1 of SEQ ID NO: 4, CDRL2 of SEQ ID NO: 5, and CDRL3 of SEQ ID NO: 6; wherein the composition comprises ≤25% of the isomerization variant.

[0138] In one aspect, the composition comprises a population of anti-BCMA antibodies, which comprises:

[0139] an antibody, which comprises a heavy chain amino acid sequence comprising SEQ ID NO: 1 (CDRH1), SEQ ID NO: 2 (CDRH2), and SEQ ID NO: 3 (CDRH3) and a light chain amino acid sequence comprising SEQ ID NO: 4 (CDRL1), SEQ ID NO: 5 (CDRL2), and SEQ ID NO: 6 (CDRL3), and

[0140] its isomerization variant, wherein ≤25% of the population of antibodies consists of the isomerization variant.

[0141] In another embodiment, the composition comprises an isomerization variant of an anti-BCMA antibody, wherein the isomerization variant comprises a heavy chain amino acid sequence comprising CDRH1 of SEQ ID NO: 1, CDRH2 of SEQ ID NO: 2, and CDRH3 of SEQ ID NO: 3, and a light chain amino acid sequence comprising CDRL1 of SEQ ID NO: 4, CDRL2 of SEQ ID NO: 5, and CDRL3 of SEQ ID NO: 6; wherein the composition comprises ≤25% of the isomerization variant at amino acid D103 in CDRH3.

[0142] In one embodiment, the composition comprises an isomerization variant of an anti-BCMA antibody, wherein the isomerization variant comprises the heavy chain sequence of SEQ ID NO: 9 and the light chain sequence of SEQ ID NO: 10; wherein the composition comprises ≤25% of the isomerization variant.

[0143] Alternatively, the isomerization variant comprises the heavy chain sequence of SEQ ID NO: 11, 12, 13, or 14.

[0144] In one embodiment, the composition comprises an antibody that is at least about 90% identical to the heavy chain amino acid sequence of SEQ ID NO: 9 and / or the light chain sequence of SEQ ID NO: 10, and comprises isomerization in either the heavy chain sequence or the light chain sequence, such as isomerization at amino acid D103 in CDRH3.

[0145] In another embodiment, the composition comprises an antibody that comprises CDRH1 having the amino acid sequence set forth in SEQ ID NO: 1, CDRH2 having the amino acid sequence set forth in SEQ ID NO: 2, CDRH3 having the amino acid sequence set forth in the group of SEQ ID NO: 3, CDRL1 having the amino acid sequence set forth in SEQ ID NO: 4, CDRL2 having the amino acid sequence set forth in SEQ ID NO: 5, CDRL3 having the amino acid sequence set forth in SEQ ID NO: 6, and isomerization in at least one of the six CDR regions, such as isomerization at amino acid D103 in CDRH3.

[0146] In another embodiment, the anti-BCMA antibody comprises belantamab and comprises isomerization in either the heavy chain sequence or the light chain sequence, such as isomerization at amino acid D103 in CDRH3.

[0147] In one embodiment, the composition comprises a mixture of antibodies that are at least about 90% identical to the heavy chain amino acid sequence of SEQ ID NO: 9 and / or the light chain sequence of SEQ ID NO: 10, wherein about ≤25%, ≤23%, ≤20%, ≤15%, ≤10%, ≤8%, ≤7%, 0.1 - 25%, 0.1 - 20%, 0.1 - 15%, 0.1 - 10%, 0.1 - 8%, 0.1 - 7%, 1 - 6%, 2 - 6%, 3 - 6%, about 4%, about 5% or about 6% of the antibodies in the mixture are isomerized at amino acid D103 in CDRH3. In one embodiment, a composition comprising ≤25% or ≤23% isomerization at D103 in CDRH3 retains ≥70% of the BCMA-specific antigen binding.

[0148] In one embodiment, the composition comprises a mixture of antibodies, which comprises CDRH1 having the amino acid sequence set forth in SEQ ID NO: 1, CDRH2 having the amino acid sequence set forth in SEQ ID NO: 2, a group of CDRH3 having the amino acid sequence set forth in SEQ ID NO: 3, CDRL1 having the amino acid sequence set forth in SEQ ID NO: 4, CDRL2 having the amino acid sequence set forth in SEQ ID NO: 5, CDRL3 having the amino acid sequence set forth in SEQ ID NO: 6, wherein about ≤25%, ≤23%, ≤20%, ≤15%, ≤10%, ≤8%, ≤7%, 0.1-25%, 0.1-20%, 0.1-15%, 0.1-10%, 0.1-8%, 0.1-7%, 1-6%, 2-6%, 3-6%, about 4%, about 5% or about 6% of the antibodies in the mixture are isomerized at amino acid D103 of CDRH3. In one embodiment, the composition comprising ≤25% or ≤23% isomerization at D103 of CDRH3 retains ≥70% of the BCMA-specific antigen binding.

[0149] In another embodiment, the composition comprises belantamab, wherein about ≤25%, ≤23%, ≤20%, ≤15%, ≤10%, ≤8%, ≤7%, 0.1-25%, 0.1-20%, 0.1-15%, 0.1-10%, 0.1-8%, 0.1-7%, 1-6%, 2-6%, 3-6%, about 4%, about 5% or about 6% of belantamab is isomerized at amino acid D103 of CDRH3. In one embodiment, belantamab comprising ≤25% or ≤23% isomerization at D103 of CDRH3 retains ≥70% of the BCMA-specific antigen binding.

[0150] In one embodiment, the composition comprises an isomerized variant of an anti-BCMA antibody, wherein the isomerized variant comprises the heavy chain sequence of SEQ ID NO: 9 and the light chain sequence of SEQ ID NO: 10; wherein the composition comprises ≤25% of the isomerized variant.

[0151] In one example, aspartic acid (Asp) isomerization can be determined using tryptic peptide mapping tandem mass spectrometry (peptide mapping LC-MS / MS). In one example, a sample containing the composition described herein can be denatured, for example, in 6 M guanidine hydrochloride to a concentration of, for example, 4.2 μg / μL. Disulfide bonds can then be reduced, for example, with 50 mM DTT for 20 minutes at room temperature. Iodoacetate can then be added, for example, at 100 mM and reacted with free cysteine residues, for example, for 30 minutes at room temperature in the dark. Before digestion with 0.5% trypsin at 37°C for 15 minutes, the sample can be buffer exchanged, for example, using a BioRad spin column (part number 7326221). The resulting peptides can then be loaded onto a reverse-phase ultra-high performance liquid chromatography (UPLC) column, and the peptides can be eluted using a gradient of water and acetonitrile in, for example, 0.1% trifluoroacetic acid using UPLC. The peptides can then be detected with a UV detector and a mass spectrometer (e.g., Thermo Scientific LTQ Orbitrap XL). The extracted ion chromatograms of unmodified and modified peptides can be used to calculate the level of isomerization by dividing the area under the curve of the modified peptide by the total area under the curves of both modified and unmodified peptides.

[0152] In one aspect, the composition comprises an antibody that comprises an oxidative post-translational modification (“oxidation” or “oxidized”) or an “oxidized variant”. The variant can comprise oxidized amino acid residues in the heavy chain sequence and / or the light chain sequence, such as in the CDRs of the heavy chain sequence and / or the CDRs of the light chain sequence. The oxidized variant can be present in one or both of the heavy and light chains. It will be understood that these oxidized variant embodiments can be combined with the antibody features described herein.

[0153] In one embodiment, the composition comprises an oxidized variant of an anti-BCMA antibody, wherein the oxidized variant comprises a heavy chain amino acid sequence comprising CDRH1 of SEQ ID NO: 1, CDRH2 of SEQ ID NO: 2, and CDRH3 of SEQ ID NO: 3, and a light chain amino acid sequence comprising CDRL1 of SEQ ID NO: 4, CDRL2 of SEQ ID NO: 5, and CDRL3 of SEQ ID NO: 6; wherein the composition comprises ≤40% oxidized variant.

[0154] In one aspect, the composition comprises a population of anti-BCMA antibodies that includes:

[0155] An antibody that comprises a heavy chain amino acid sequence comprising SEQ ID NO: 1 (CDRH1), SEQ ID NO: 2 (CDRH2), and SEQ ID NO: 3 (CDRH3), and a light chain amino acid sequence comprising SEQ ID NO: 4 (CDRL1), SEQ ID NO: 5 (CDRL2), and SEQ ID NO: 6 (CDRL3), and

[0156] its oxidized variants, wherein ≤25% of the antibody population consists of oxidized variants.

[0157] In one embodiment, the oxidized variants include oxidation in one or more of the CDRs. In a further embodiment, the oxidized variants include oxidation at methionine and / or tryptophan residues in any of SEQ ID NOs: 1-6.

[0158] In another embodiment, the composition comprises an oxidized variant of an anti-BCMA antibody, wherein the oxidized variant comprises a heavy chain amino acid sequence comprising CDRH1 of SEQ ID NO: 1, CDRH2 of SEQ ID NO: 2, and CDRH3 of SEQ ID NO: 3, and a light chain amino acid sequence comprising CDRL1 of SEQ ID NO: 4, CDRL2 of SEQ ID NO: 5, and CDRL3 of SEQ ID NO: 6; wherein the composition comprises ≤40% of the oxidized variant at amino acid M34 in CDRH1.

[0159] In one embodiment, the composition comprises an oxidized variant of belantamab, wherein the oxidized variant comprises the heavy chain sequence of SEQ ID NO: 9 and the light chain sequence of SEQ ID NO: 10; wherein the composition comprises ≤40% of the oxidized variant.

[0160] In one embodiment, the composition comprises an antibody that is at least about 90% identical to the heavy chain amino acid sequence of SEQ ID NO: 9 and / or the light chain sequence of SEQ ID NO: 10, and comprises oxidation in the heavy chain amino acid sequence, such as oxidation at amino acids M34 (CDRH1), M256, and / or M432.

[0161] In another embodiment, the composition comprises an antibody that comprises a CDRH1 having the amino acid sequence set forth in SEQ ID NO: 1, a CDRH2 having the amino acid sequence set forth in SEQ ID NO: 2, a CDRH3 having the amino acid sequence set forth in SEQ ID NO: 3, a CDRL1 having the amino acid sequence set forth in SEQ ID NO: 4, a CDRL2 having the amino acid sequence set forth in SEQ ID NO: 5, a CDRL3 having the amino acid sequence set forth in SEQ ID NO: 6, and includes oxidation in the heavy chain sequence, such as oxidation at amino acids M34 (CDRH1), M256, and / or M432.

[0162] In another embodiment, the anti-BCMA antibody comprises belantamab and includes oxidation in the heavy chain sequence, such as oxidation at amino acids M34 (CDRH1), M256, and / or M432.

[0163] In one embodiment, the composition comprises a mixture of antibodies that are at least about 90% identical to the heavy chain amino acid sequence of SEQ ID NO: 9 and / or the light chain sequence of SEQ ID NO: 10, wherein about ≤40%, ≤35%, 30%, ≤25%, ≤20%, ≤15%, ≤10%, ≤7.5%, ≤5%, ≤2.5%, ≤2%, 0.1 - 40%, 0.1 - 35%, 0.1 - 30%, 0.1 - 25%, 0.1 - 20%, 0.1 - 15%, 0.1 - 10%, 0.1 - 7.5%, 0.1 - 5%, 0.1 - 2.5%, 0.1 - 2%, about 0.5%, about 1%, about 2%, or about 5% of the antibodies are oxidized at amino acid M34. In one embodiment, a composition comprising ≤40% oxidation at heavy chain M34 retains ≥70% of BCMA-specific antigen binding. In another embodiment, a composition comprising ≤37% oxidation at heavy chain M34 retains ≥70% of BCMA-specific antigen binding.

[0164] In one embodiment, the composition comprises an antibody mixture that includes a CDRH1 having the amino acid sequence set forth in SEQ ID NO: 1, a CDRH2 having the amino acid sequence set forth in SEQ ID NO: 2, a group of CDRH3 having the amino acid sequence set forth in SEQ ID NO: 3, a CDRL1 having the amino acid sequence set forth in SEQ ID NO: 4, a CDRL2 having the amino acid sequence set forth in SEQ ID NO: 5, and a CDRL3 having the amino acid sequence set forth in SEQ ID NO: 6, wherein about ≤40%, ≤35%, 30%, ≤25%, ≤20%, ≤15%, ≤10%, ≤7.5%, ≤5%, ≤2.5%, ≤2%, 0.1 - 40%, 0.1 - 35%, 0.1 - 30%, 0.1 - 25%, 0.1 - 20%, 0.1 - 15%, 0.1 - 10%, 0.1 - 7.5%, 0.1 - 5%, 0.1 - 2.5%, 0.1 - 2%, about 0.5%, about 1%, about 2%, or about 5% of the antibodies in the mixture are oxidized at amino acid M34. In one embodiment, a composition comprising ≤40% oxidation at heavy chain M34 retains ≥70% of BCMA - specific antigen binding. In another embodiment, a composition comprising ≤37% oxidation at heavy chain M34 retains ≥70% of BCMA - specific antigen binding.

[0165] In another embodiment, the composition comprises belantamab, wherein about ≤40%, ≤35%, 30%, ≤25%, ≤20%, ≤15%, ≤10%, ≤7.5%, ≤5%, ≤2.5%, ≤2%, 0.1 - 40%, 0.1 - 35%, 0.1 - 30%, 0.1 - 25%, 0.1 - 20%, 0.1 - 15%, 0.1 - 10%, 0.1 - 7.5%, 0.1 - 5%, 0.1 - 2.5%, 0.1 - 2%, about 0.5%, about 1%, about 2%, or about 5% of the belantamab is oxidized at amino acid M34. In one embodiment, belantamab comprising ≤40% oxidation at heavy chain M34 retains ≥70% of BCMA - specific antigen binding. In another embodiment, belantamab comprising ≤37% oxidation at heavy chain M34 retains ≥70% of BCMA - specific antigen binding.

[0166] In one embodiment, the composition comprises an oxidized variant of an anti-BCMA antibody, wherein the oxidized variant comprises a heavy chain amino acid sequence comprising CDRH1 of SEQ ID NO: 1, CDRH2 of SEQ ID NO: 2, and CDRH3 of SEQ ID NO: 3, and a light chain amino acid sequence comprising CDRL1 of SEQ ID NO: 4, CDRL2 of SEQ ID NO: 5, and CDRL3 of SEQ ID NO: 6; wherein the composition comprises ≤90% of the oxidized variant in the Fc region.

[0167] In one embodiment, the antibody comprises oxidation at methionine and / or tryptophan residues in the Fc region of the heavy chain sequence and / or the Fc region of the light chain sequence. In some embodiments, the oxidized variant comprises one or a combination of oxidations at the following positions: M256 and / or M432 in the Fc region of the heavy chain sequence.

[0168] In another embodiment, the composition comprises an oxidized variant of an anti-BCMA antibody, wherein the oxidized variant comprises a heavy chain amino acid sequence comprising CDRH1 of SEQ ID NO: 1, CDRH2 of SEQ ID NO: 2, and CDRH3 of SEQ ID NO: 3, and a light chain amino acid sequence comprising CDRL1 of SEQ ID NO: 4, CDRL2 of SEQ ID NO: 5, and CDRL3 of SEQ ID NO: 6; wherein the composition comprises ≤90% of the oxidized M256 and / or M432 variant.

[0169] In one embodiment, the composition comprises an oxidized variant of belantamab, wherein the oxidized variant comprises the heavy chain sequence of SEQID NO: 9 and the light chain sequence of SEQ ID NO: 10; wherein the composition comprises ≤90% of the oxidized variant in the Fc region.

[0170] Alternatively, the oxidized variant comprises the heavy chain sequence of SEQ ID NO: 11, 12, 13 or 14.

[0171] In one embodiment, the composition comprises a mixture of antibodies having at least about 90% identity to the heavy chain amino acid sequence of SEQ ID NO: 9 and / or the light chain sequence of SEQ ID NO: 10, wherein about ≤90%, ≤80%, ≤70%, ≤65%, ≤50%, ≤40%, ≤30%, ≤20%, ≤10%, ≤7.5%, ≤5%, 0.1 - 90%, 0.1 - 80%, 0.1 - 70%, 0.1 - 65%, 0.1 - 50%, 0.1 - 40%, 0.1 - 30%, 0.1 - 20%, 0.1 - 10%, 1 - 10%, 1 - 5%, 2 - 10%, 2 - 4%, about 2%, about 3% or about 4% of the antibodies in the mixture are oxidized at amino acid M256. In one embodiment, a composition comprising ≤90% or ≤89% oxidation at heavy chain M256 retains ≥70% FcγRIIIA binding. In another embodiment, a composition comprising ≤65% or ≤64% oxidation at heavy chain M256 retains ≥70% FcRn binding.

[0172] In one embodiment, the composition comprises a mixture of antibodies comprising CDRH1 having the amino acid sequence set forth in SEQ ID NO: 1, CDRH2 having the amino acid sequence set forth in SEQ ID NO: 2, a group of CDRH3 having the amino acid sequence set forth in SEQ ID NO: 3, CDRL1 having the amino acid sequence set forth in SEQ ID NO: 4, CDRL2 having the amino acid sequence set forth in SEQ ID NO: 5, CDRL3 having the amino acid sequence set forth in SEQ ID NO: 6, wherein about ≤90%, ≤80%, ≤70%, ≤65%, ≤50%, ≤40%, ≤30%, ≤20%, ≤10%, ≤7.5%, ≤5%, 0.1 - 90%, 0.1 - 80%, 0.1 - 70%, 0.1 - 65%, 0.1 - 50%, 0.1 - 40%, 0.1 - 30%, 0.1 - 20%, 0.1 - 10%, 1 - 10%, 1 - 5%, 2 - 10%, 2 - 4%, about 2%, about 3% or about 4% of the antibodies in the mixture are oxidized at amino acid M256. In one embodiment, a composition comprising ≤90% or ≤89% oxidation at heavy chain M256 retains ≥70% FcγRIIIA binding. In another embodiment, a composition comprising ≤65% or ≤64% oxidation at heavy chain M256 retains ≥70% FcRn binding.

[0173] In another embodiment, the composition comprises belantamab, wherein about ≤90%, ≤80%, ≤70%, ≤65%, ≤50%, ≤40%, ≤30%, ≤20%, ≤10%, ≤7.5%, ≤5%, 0.1-90%, 0.1-80%, 0.1-70%, 0.1-65%, 0.1-50%, 0.1-40%, 0.1-30%, 0.1-20%, 0.1-10%, 1-10%, 1-5%, 2-10%, 2-4%, about 2%, about 3%, or about 4% of the belantamab is oxidized at amino acid M256. In one embodiment, belantamab comprising ≤90% or ≤89% oxidation at heavy chain M256 retains ≥70% FcγRIIIA binding. In another embodiment, belantamab comprising ≤65% or ≤64% oxidation at heavy chain M256 retains ≥70% FcRn binding.

[0174] In one embodiment, the composition comprises a mixture of antibodies that is at least about 90% identical to the heavy chain amino acid sequence of SEQ ID NO: 9 and / or the light chain sequence of SEQ ID NO: 10, wherein about ≤86%, ≤70%, ≤60%, ≤50%, ≤40%, ≤30%, ≤20%, ≤10%, ≤7.5%, ≤5%, ≤2.5%, ≤2%, 0.1-86%, 0.1-70%, 0.1-60%, 0.1-50%, 0.1-40%, 0.1-30%, 0.1-20%, 0.1-10%, 0.1-5%, 0.1-3%, about 0.5%, about 1%, about 2%, or about 3% of the antibodies in the mixture are oxidized at amino acid M432. In one embodiment, the composition comprising ≤86% oxidation at heavy chain M432 retains ≥70% FcγRIIA binding. In another embodiment, the composition comprising ≤60% oxidation at heavy chain M432 retains ≥70% FcRn binding.

[0175] In one embodiment, the composition comprises a mixture of antibodies, the mixture comprising a CDRH1 having the amino acid sequence set forth in SEQ ID NO: 1, a CDRH2 having the amino acid sequence set forth in SEQ ID NO: 2, a group of CDRH3 having the amino acid sequence set forth in SEQ ID NO: 3, a CDRL1 having the amino acid sequence set forth in SEQ ID NO: 4, a CDRL2 having the amino acid sequence set forth in SEQ ID NO: 5, and a CDRL3 having the amino acid sequence set forth in SEQ ID NO: 6, wherein about ≤86%, ≤70%, ≤60%, ≤50%, ≤40%, ≤30%, ≤20%, ≤10%, ≤7.5%, ≤5%, ≤2.5%, ≤2%, 0.1-86%, 0.1-70%, 0.1-60%, 0.1-50%, 0.1-40%, 0.1-30%, 0.1-20%, 0.1-10%, 0.1-5%, 0.1-3%, about 0.5%, about 1%, about 2%, or about 3% of the antibodies are oxidized at amino acid M432. A composition comprising ≤86% oxidation at heavy chain M432 retains ≥70% FcγRIIIA binding. In another embodiment, a composition comprising ≤60% oxidation at heavy chain M432 retains ≥70% FcRn binding.

[0176] In another embodiment, the composition comprises belantamab, wherein about ≤86%, ≤70%, ≤60%, ≤50%, ≤40%, ≤30%, ≤20%, ≤10%, ≤7.5%, ≤5%, ≤2.5%, ≤2%, 0.1-86%, 0.1-70%, 0.1-60%, 0.1-50%, 0.1-40%, 0.1-30%, 0.1-20%, 0.1-10%, 0.1-5%, 0.1-3%, about 0.5%, about 1%, about 2%, or about 3% of the belantamab is oxidized at amino acid M432. In one embodiment, belantamab comprising ≤86% oxidation at heavy chain M432 retains ≥70% FcγRIIIa binding. In another embodiment, belantamab comprising ≤60% oxidation at heavy chain M432 retains ≥70% FcRn binding.

[0177] In one instance, oxidation can be determined using tryptic peptide mapping tandem mass spectrometry (peptide mapping LC-MS / MS). In one instance, a sample containing the compositions described herein can be denatured, for example, in 6 M guanidine hydrochloride to a concentration of, for example, 4.2 μg / μL. Disulfide bonds can then be reduced, for example, with 50 mM DTT for 20 minutes at room temperature. Iodoacetate can then be added, for example, at 100 mM and reacted with free cysteine residues, for example, for 30 minutes at room temperature in the dark. The sample can then be buffer exchanged, for example, using a BioRad spin column (part number 7326221) before digestion with 0.5% trypsin at 37 °C for 15 minutes. The resulting peptides can then be loaded onto a reversed-phase ultra-performance liquid chromatography (UPLC) column and eluted using a gradient of water and acetonitrile in, for example, 0.1% trifluoroacetic acid using UPLC. The peptides can then be detected with a UV detector and a mass spectrometer (e.g., Thermo Scientific LTQ Orbitrap XL). Extracted ion chromatograms of unmodified and modified peptides are used to calculate the level of oxidation by dividing the area under the curve of the modified peptide by the total area under the curves of both modified and unmodified peptides.

[0178] In one aspect, the composition comprises an antibody that comprises a deamidated post-translational modification (“deamidation” or “deamidated”) or “deamidation variant”. In one embodiment, the antibody comprises deamidation of asparagine residues in the CDR of the heavy chain sequence and / or the CDR of the light chain sequence. In a further embodiment, the antibody comprises deamidation of asparagine residues in the CDR of the heavy chain sequence. In one embodiment, the antibody comprises deamidation of asparagine residues in the Fc region of the heavy chain sequence and / or the Fc region of the light chain sequence. The deamidation variants can be present in one or both of the heavy or light chains. It will be understood that these deamidation variant embodiments can be combined with the antibody features described herein. In some embodiments, the deamidation variant comprises one or a combination of deamidations at positions N388 and / or N393 of the Fc region of the heavy chain sequence.

[0179] In one embodiment, the deamidation variant comprises a deamidated residue selected from: an aspartic acid residue, a succinimide-aspartic acid residue, or an isoaspartic acid residue.

[0180] In one embodiment, the composition comprises an antibody that is at least about 90% identical to the heavy chain amino acid sequence of SEQ ID NO: 9 and / or the light chain sequence of SEQ ID NO: 10 and comprises deamidation in the heavy chain amino acid sequence, for example, deamidation at amino acids N388 and / or N393.

[0181] In one embodiment, the composition comprises a deamidated variant of an anti-BCMA antibody, wherein the deamidated variant comprises a heavy chain amino acid sequence comprising CDRH1 of SEQ ID NO: 1, CDRH2 of SEQ ID NO: 2, and CDRH3 of SEQ ID NO: 3, and a light chain amino acid sequence comprising CDRL1 of SEQ ID NO: 4, CDRL2 of SEQ ID NO: 5, and CDRL3 of SEQ ID NO: 6; wherein the composition comprises up to 100% of the deamidated variant.

[0182] In another embodiment, the composition comprises a deamidated variant of an anti-BCMA antibody, wherein the oxidized variant comprises a heavy chain amino acid sequence comprising CDRH1 of SEQ ID NO: 1, CDRH2 of SEQ ID NO: 2, and CDRH3 of SEQ ID NO: 3, and a light chain amino acid sequence comprising CDRL1 of SEQ ID NO: 4, CDRL2 of SEQ ID NO: 5, and CDRL3 of SEQ ID NO: 6; wherein the composition comprises up to 100% of the N388 and / or N393 deamidated variant.

[0183] In one embodiment, the composition comprises a deamidated variant of belantamab, wherein the deamidated variant comprises the heavy chain sequence of SEQ ID NO: 9 and the light chain sequence of SEQ ID NO: 10; wherein the composition comprises up to 100% of the deamidated variant. In another embodiment, the composition comprises a deamidated variant that comprises the heavy chain sequence of SEQ ID NO: 12, SEQ ID NO: 13, or SEQ ID NO: 14 and the light chain of SEQ ID NO: 10.

[0184] In another embodiment, the composition comprises an antibody that comprises CDRH1 having the amino acid sequence set forth in SEQ ID NO: 1, CDRH2 having the amino acid sequence set forth in SEQ ID NO: 2, CDRH3 having the amino acid sequence set forth in SEQ ID NO: 3, CDRL1 having the amino acid sequence set forth in SEQ ID NO: 4, CDRL2 having the amino acid sequence set forth in SEQ ID NO: 5, CDRL3 having the amino acid sequence set forth in SEQ ID NO: 6, and comprises deamidation in the heavy chain sequence, such as deamidation at amino acids N388 and / or N393.

[0185] In another embodiment, the anti-BCMA antibody comprises belantamab and comprises deamidation in the heavy chain sequence, such as deamidation at amino acids N388 and / or N393.

[0186] In one embodiment, the composition comprises a mixture of antibodies that is at least about 90% identical to the heavy chain amino acid sequence of SEQ ID NO: 9 and / or the light chain sequence of SEQ ID NO: 10, wherein about ≤100%, ≤75%, ≤60%, ≤50%, ≤40%, ≤30%, ≤25%, ≤20%, ≤15%, ≤10%, ≤5%, ≤2%, 0.1 - 100%, 0.1 - 75%, 0.1 - 50%, 0.1 - 40%, 0.1 - 30%, 0.1 - 20%, or 0.1 - 10%, 0.1 - 5%, 0.1 - 3%, about 0.5%, about 1%, about 2%, about 5%, or about 10% of the antibodies in the mixture are deamidated at amino acid N388.

[0187] In one embodiment, the composition comprises a mixture of antibodies that comprises CDRH1 having the amino acid sequence set forth in SEQ ID NO: 1, CDRH2 having the amino acid sequence set forth in SEQ ID NO: 2, CDRH3 having the amino acid sequence set forth in SEQ ID NO: 3, CDRL1 having the amino acid sequence set forth in SEQ ID NO: 4, CDRL2 having the amino acid sequence set forth in SEQ ID NO: 5, and CDRL3 having the amino acid sequence set forth in SEQ ID NO: 6, wherein about ≤100%, ≤75%, ≤60%, ≤50%, ≤40%, ≤30%, ≤25%, ≤20%, ≤15%, ≤10%, ≤5%, ≤2%, 0.1 - 100%, 0.1 - 75%, 0.1 - 50%, 0.1 - 40%, 0.1 - 30%, 0.1 - 20%, or 0.1 - 10%, 0.1 - 5%, 0.1 - 3%, about 0.5%, about 1%, about 2%, about 5%, or about 10% of the antibodies in the mixture are deamidated at amino acid N388.

[0188] In another embodiment, the composition comprises belantamab, wherein about ≤100%, ≤75%, ≤60%, ≤50%, ≤40%, ≤30%, ≤25%, ≤20%, ≤15%, ≤10%, ≤5%, ≤2%, 0.1 - 100%, 0.1 - 75%, 0.1 - 50%, 0.1 - 40%, 0.1 - 30%, 0.1 - 20%, or 0.1 - 10%, 0.1 - 5%, 0.1 - 3%, about 0.5%, about 0.5%, about 1%, about 2%, about 5%, or about 10% of the belantamab is deamidated at amino acid N388.

[0189] In one embodiment, the composition comprises a mixture of antibodies that is at least about 90% identical to the heavy chain amino acid sequence of SEQ ID NO: 9 and / or the light chain sequence of SEQ ID NO: 10, wherein about ≤100%, ≤85%, ≤70%, ≤60%, ≤50%, ≤40%, ≤30%, ≤20%, ≤15%, ≤10%, ≤5%, ≤2%, 0.1 - 100%, 0.1 - 75%, 0.1 - 50%, 0.1 - 40%, 0.1 - 30%, 0.1 - 20%, or 0.1 - 10%, 0.1 - 5%, 0.1 - 3%, about 0.5%, about 1%, about 2%, about 5%, or about 10% of the antibodies in the mixture are deamidated at amino acid N393.

[0190] In one embodiment, the composition comprises a mixture of antibodies that comprises CDRH1 having the amino acid sequence set forth in SEQ ID NO: 1, CDRH2 having the amino acid sequence set forth in SEQ ID NO: 2, a group of CDRH3 having the amino acid sequence set forth in SEQ ID NO: 3, CDRL1 having the amino acid sequence set forth in SEQ ID NO: 4, CDRL2 having the amino acid sequence set forth in SEQ ID NO: 5, and CDRL3 having the amino acid sequence set forth in SEQ ID NO: 6, wherein about ≤100%, ≤85%, ≤70%, ≤60%, ≤50%, ≤40%, ≤30%, ≤20%, ≤15%, ≤10%, ≤5%, ≤2%, 0.1 - 100%, 0.1 - 75%, 0.1 - 50%, 0.1 - 40%, 0.1 - 30%, 0.1 - 20%, or 0.1 - 10%, 0.1 - 5%, 0.1 - 3%, about 0.5%, about 1%, about 2%, about 5%, or about 10% of the antibodies in the mixture are deamidated at amino acid N393.

[0191] In another embodiment, the composition comprises belantamab, wherein about ≤100%, ≤85%, ≤70%, ≤60%, ≤50%, ≤40%, ≤30%, ≤20%, ≤15%, ≤10%, ≤5%, ≤2%, 0.1 - 100%, 0.1 - 75%, 0.1 - 50%, 0.1 - 40%, 0.1 - 30%, 0.1 - 20%, or 0.1 - 10%, 0.1 - 5%, 0.1 - 3%, about 0.5%, about 1%, about 2%, about 5%, or about 10% of the belantamab is deamidated at amino acid N393.

[0192] In one example, deamidation can be determined using tryptic peptide mapping tandem mass spectrometry (peptide mapping LC-MS / MS). In one example, a sample containing the compositions described herein can be denatured, for example, in 6 M guanidine hydrochloride to a concentration of, for example, 4.2 μg / μL. The disulfide bonds can then be reduced, for example, with 50 mM DTT for 20 minutes at room temperature. Iodoacetate can then be added, for example, at 100 mM and reacted with free cysteine residues, for example, for 30 minutes at room temperature in the dark. Before digestion with 0.5% trypsin at 37°C for 15 minutes, the sample can be buffer exchanged, for example, using a BioRad spin column (part number 7326221). The resulting peptides can then be loaded onto a reversed-phase ultra-performance liquid chromatography (UPLC) column, and eluted using UPLC with a gradient of water and acetonitrile in, for example, 0.1% trifluoroacetic acid. The peptides can then be detected using a UV detector and a mass spectrometer (e.g., Thermo Scientific LTQ Orbitrap XL). The extracted ion chromatograms of the unmodified and modified peptides are used to calculate the level of deamidation by dividing the area under the curve of the modified peptide by the total area under the curves of both the modified and unmodified peptides.

[0193] In one embodiment, the post-translational modification is an antibody sequence variant. Exemplary post-translational modification antibody sequence variants include asparagine (N) to aspartic acid (D) switches, N-terminal pyroglutamic acid, and / or C-terminal lysine cleavage.

[0194] In one example, antibody variants, such as N103D at CDRH3, can be determined using tryptic peptide mapping tandem mass spectrometry (peptide mapping LC-MS / MS). In one example, a sample containing the compositions described herein can be denatured, for example, in 6 M guanidine hydrochloride to a concentration of, for example, 4.2 μg / μL. The disulfide bonds can then be reduced, for example, with 50 mM DTT for 20 minutes at room temperature. Iodoacetate can then be added, for example, at 100 mM and reacted with free cysteine residues, for example, for 30 minutes at room temperature in the dark. Before digestion with 0.5% trypsin at 37°C for 15 minutes, the sample can be buffer exchanged, for example, using a BioRad spin column (part number 7326221). The resulting peptides can then be loaded onto a reversed-phase ultra-performance liquid chromatography (UPLC) column, and eluted using UPLC with a gradient of water and acetonitrile in, for example, 0.1% trifluoroacetic acid. The peptides can then be detected using a UV detector and a mass spectrometer (e.g., Thermo Scientific LTQ Orbitrap XL). The extracted ion chromatograms of the unmodified and modified peptides are used to calculate the level of the antibody variant, such as N103D at CDRH3, by dividing the area under the curve of the modified peptide by the total area under the curves of both the modified and unmodified peptides.

[0195] In one aspect, the composition comprises an antibody that comprises an N-terminal pyroglutamic acid (“pyroGlu”) post-translational modification in the heavy chain amino acid sequence. In one embodiment, the composition comprises an antibody that is at least about 90% identical to the heavy chain amino acid sequence of SEQ ID NO: 9 and / or the light chain sequence of SEQ ID NO: 10, and that comprises pyroGlu at the N-terminus of the heavy chain.

[0196] In another embodiment, the composition comprises an antibody that comprises CDRH1 having the amino acid sequence set forth in SEQ ID NO: 1, CDRH2 having the amino acid sequence set forth in SEQ ID NO: 2, CDRH3 having the amino acid sequence set forth in the group of SEQ ID NO: 3, CDRL1 having the amino acid sequence set forth in SEQ ID NO: 4, CDRL2 having the amino acid sequence set forth in SEQ ID NO: 5, CDRL3 having the amino acid sequence set forth in SEQ ID NO: 6, and that comprises pyroGlu at the N-terminus of the heavy chain.

[0197] In another embodiment, the anti-BCMA antibody comprises belantamab and comprises pyroGlu at the N-terminus of the heavy chain.

[0198] In one embodiment, the composition comprises a mixture of antibodies that is at least about 90% identical to the heavy chain amino acid sequence of SEQ ID NO: 9 and / or the light chain sequence of SEQ ID NO: 10, wherein about ≥25%, ≥50%, ≥75%, ≥80%, ≥85%, ≥90%, ≥95%, 100% or less, 95% or less, 90% or less, 85% or less, 80% or less, 75% or less, or 50% or less of the antibodies in the mixture comprise an N-terminal pyroglutamic acid in the heavy chain amino acid sequence.

[0199] In one embodiment, the composition comprises a mixture of antibodies that comprises CDRH1 having the amino acid sequence set forth in SEQ ID NO: 1, CDRH2 having the amino acid sequence set forth in SEQ ID NO: 2, CDRH3 having the amino acid sequence set forth in the group of SEQ ID NO: 3, CDRL1 having the amino acid sequence set forth in SEQ ID NO: 4, CDRL2 having the amino acid sequence set forth in SEQ ID NO: 5, CDRL3 having the amino acid sequence set forth in SEQ ID NO: 6, wherein about ≥25%, ≥50%, ≥75%, ≥80%, ≥85%, ≥90%, ≥95%, 100% or less, 95% or less, 90% or less, 85% or less, 80% or less, 75% or less, or 50% or less of the antibodies in the mixture comprise an N-terminal pyroglutamic acid in the heavy chain amino acid sequence.

[0200] In one embodiment, the composition comprises belantamab, wherein about ≥25%, ≥50%, ≥75%, ≥80%, ≥85%, ≥90%, ≥95%, 100% or less, 95% or less, 90% or less, 85% or less, 80% or less, 75% or less, or 50% or less of the belantamab comprises an N-terminal pyroglutamic acid in the heavy chain amino acid sequence.

[0201] In one example, the N-terminal pyroglutamic acid can be determined using tryptic peptide mapping tandem mass spectrometry (peptide mapping LC-MS / MS). In one example, a sample comprising the composition described herein can be denatured, for example, in 6 M guanidine hydrochloride to a concentration of, for example, 4.2 μg / μL. The disulfide bonds can then be reduced, for example, with 50 mM DTT for 20 minutes at room temperature. Iodoacetate can then be added, for example, at 100 mM and reacted with the free cysteine residues, for example, for 30 minutes at room temperature in the dark. The sample can then be buffer exchanged, for example, using a BioRad spin column (part number 7326221) before digestion with 0.5% trypsin at 37 °C for 15 minutes. The resulting peptides can then be loaded onto a reversed-phase ultra-performance liquid chromatography (UPLC) column and eluted using a gradient of water and acetonitrile in, for example, 0.1% trifluoroacetic acid using UPLC. The peptides can then be detected with a UV detector and a mass spectrometer (e.g., Thermo Scientific LTQ Orbitrap XL). The extracted ion chromatograms of the unmodified and modified peptides are used to calculate the level of pyroglutamic acid by dividing the area under the curve of the modified peptide by the total area under the curves of both the modified and unmodified peptides.

[0202] In one aspect, the composition comprises an antibody that comprises a C-terminal lysine cleavage post-translational modification in the heavy chain amino acid sequence. In one embodiment, the composition comprises an antibody that is at least about 90% identical to the heavy chain amino acid sequence of SEQ ID NO: 9 and / or the light chain sequence of SEQ ID NO: 10 and comprises a C-terminal lysine cleavage of the heavy chain.

[0203] In another embodiment, the composition comprises an antibody that comprises a CDRH1 having the amino acid sequence set forth in SEQ ID NO: 1, a CDRH2 having the amino acid sequence set forth in SEQ ID NO: 2, a CDRH3 having the amino acid sequence set forth in the group of SEQ ID NO: 3, a CDRL1 having the amino acid sequence set forth in SEQ ID NO: 4, a CDRL2 having the amino acid sequence set forth in SEQ ID NO: 5, a CDRL3 having the amino acid sequence set forth in SEQ ID NO: 6, and comprises a C-terminal lysine cleavage of the heavy chain.

[0204] In another embodiment, the anti-BCMA antibody comprises belantamab and comprises C-terminal lysine cleavage of the heavy chain.

[0205] In one embodiment, the composition comprises a mixture of antibodies that is at least about 90% identical to the heavy chain amino acid sequence of SEQ ID NO: 9 and / or the light chain sequence of SEQ ID NO: 10, wherein about ≥25%, ≥50%, ≥75%, ≥80%, ≥85%, ≥90%, ≥95%, 100% or less, 95% or less, 90% or less, 85% or less, 80% or less, 75% or less, or 50% or less of the antibodies in the mixture comprise C-terminal lysine cleavage.

[0206] In one embodiment, the composition comprises a mixture of antibodies that comprises CDRH1 having the amino acid sequence set forth in SEQ ID NO: 1, CDRH2 having the amino acid sequence set forth in SEQ ID NO: 2, CDRH3 having the amino acid sequence set forth in the group of SEQ ID NO: 3, CDRL1 having the amino acid sequence set forth in SEQ ID NO: 4, CDRL2 having the amino acid sequence set forth in SEQ ID NO: 5, CDRL3 having the amino acid sequence set forth in SEQ ID NO: 6, wherein about ≥25%, ≥50%, ≥75%, ≥80%, ≥85%, ≥90%, ≥95%, 100% or less, 95% or less, 90% or less, 85% or less, 80% or less, 75% or less, or 50% or less of the antibodies in the mixture comprise C-terminal lysine cleavage of the heavy chain.

[0207] In one embodiment, the composition comprises belantamab, wherein about ≥25%, ≥50%, ≥75%, ≥80%, ≥85%, ≥90%, ≥95%, 100% or less, 95% or less, 90% or less, 85% or less, 80% or less, 75% or less, or 50% or less of the belantamab comprises C-terminal lysine cleavage of the heavy chain.

[0208] In one example, C-terminal lysine cleavage can be determined using tryptic peptide mapping tandem mass spectrometry (peptide mapping LC-MS / MS). In one example, a sample containing the composition described herein can be denatured, for example, in 6 M guanidine hydrochloride to a concentration of, for example, 4.2 μg / μL. Disulfide bonds can then be reduced, for example, with 50 mM DTT for 20 minutes at room temperature. Iodoacetate can then be added, for example, at 100 mM and reacted with free cysteine residues, for example, for 30 minutes at room temperature in the dark. Before digestion with 0.5% trypsin at 37°C for 15 minutes, the sample can be buffer exchanged, for example, using a BioRad spin column (part number 7326221). The resulting peptides can then be loaded onto a reversed-phase ultra-high performance liquid chromatography (UPLC) column, and the peptides can be eluted using a gradient of water and acetonitrile in, for example, 0.1% trifluoroacetic acid using UPLC. The peptides can then be detected using a UV detector and a mass spectrometer (e.g., Thermo Scientific LTQ Orbitrap XL). The extracted ion chromatograms of the unmodified and modified peptides are used to calculate the level of C-terminal lysine cleavage by dividing the area under the curve of the modified peptide by the total area under the curves of both the modified and unmodified peptides.

[0209] In one aspect, the composition comprises an antibody that comprises a post-translational glycosylation modification (“glycosylation modification”) or a glycosylation variant. Exemplary glycosylation modifications include changes in the expression of G0, G1, G0-GlcNac, G2, and sialylation on the antibody. In one embodiment, the composition comprises an antibody that is at least about 90% identical to the heavy chain amino acid sequence of SEQ ID NO: 9 and / or the light chain sequence of SEQ ID NO: 10 and comprises a glycosylation modification.

[0210] In another embodiment, the composition comprises an antibody that comprises CDRH1 having the amino acid sequence set forth in SEQ ID NO: 1, CDRH2 having the amino acid sequence set forth in SEQ ID NO: 2, CDRH3 having the amino acid sequence set forth in SEQ ID NO: 3, CDRL1 having the amino acid sequence set forth in SEQ ID NO: 4, CDRL2 having the amino acid sequence set forth in SEQ ID NO: 5, CDRL3 having the amino acid sequence set forth in SEQ ID NO: 6, and comprises a glycosylation variant.

[0211] In another embodiment, the anti-BCMA antibody comprises belantamab and comprises a glycosylation variant.

[0212] In one embodiment, the composition comprises a mixture of antibodies that is at least about 90% identical to the heavy chain amino acid sequence of SEQ ID NO: 9 and / or the light chain sequence of SEQ ID NO: 10, wherein the composition comprises about ≥25%, ≥30%, ≥35%, ≥40%, ≥45%, ≥50%, ≥55%, ≥60%, 0-100%, 1-100%, 30-100%, 40-90%, 50-80%, or 55-80% levels of G0; about ≥2.5%, ≥5%, ≥10%, ≥15%, ≥20%, ≥25%, ≥30%, ≥50%, 0-100%, 1-100%, 0-50%, 1-50%, 1-40%, 1-35%, or 8-31% levels of G1; about ≤5%, ≤7.5%, ≤10%, ≤15%, ≤20%, ≤25%, ≤30%, ≤40%, ≤50%, ≤75%, 0-100%, 0.5-100%, 0-50%, 0.5-50%, 0.5-25%, 0.5-10%, 0.5-7.5%, or 0.9-5.3% levels of G0-GlcNac; 0-100%, 1-100%, or 39-92% levels of G2; and / or 0-100%, 1-100%, or 38-88% levels of G0-2GlcNac.

[0213] In one embodiment, the composition comprises an antibody mixture that includes a CDRH1 having the amino acid sequence set forth in SEQ ID NO: 1, a CDRH2 having the amino acid sequence set forth in SEQ ID NO: 2, a CDRH3 having the amino acid sequence set forth in the group of SEQ ID NO: 3, a CDRL1 having the amino acid sequence set forth in SEQ ID NO: 4, a CDRL2 having the amino acid sequence set forth in SEQ ID NO: 5, and a CDRL3 having the amino acid sequence set forth in SEQ ID NO: 6, wherein the composition comprises a level of G0 of about ≥25%, ≥30%, ≥35%, ≥40%, ≥45%, ≥50%, ≥55%, ≥60%, 0 - 100%, 1 - 100%, 30 - 100%, 40 - 90%, 50 - 80%, or 55 - 80%; a level of G1 of about ≥2.5%, ≥5%, ≥10%, ≥15%, ≥20%, ≥25%, ≥30%, ≥50%, 0 - 100%, 1 - 100%, 0 - 50%, 1 - 50%, 1 - 40%, 1 - 35%, or 8 - 31%; a level of G0-GlcNac of about ≤5%, ≤7.5%, ≤10%, ≤15%, ≤20%, ≤25%, ≤30%, ≤40%, ≤50%, ≤75%, 0 - 100%, 0.5 - 100%, 0 - 50%, 0.5 - 50%, 0.5 - 25%, 0.5 - 10%, 0.5 - 7.5%, or 0.9 - 5.3%; a level of G2 of 0 - 100%, 1 - 100%, or 39 - 92%; and / or a level of G0-2GlcNac of 0 - 100%, 1 - 100%, or 38 - 88%.

[0214] In one embodiment, the composition comprises belantamab, wherein the composition comprises a level of G0 of about ≥25%, ≥30%, ≥35%, ≥40%, ≥45%, ≥50%, ≥55%, ≥60%, 0 - 100%, 1 - 100%, 30 - 100%, 40 - 90%, 50 - 80%, or 55 - 80%; a level of G1 of about ≥2.5%, ≥5%, ≥10%, ≥15%, ≥20%, ≥25%, ≥30%, ≥50%, 0 - 100%, 1 - 100%, 0 - 50%, 1 - 50%, 1 - 40%, 1 - 35%, or 8 - 31%; a level of G0-GlcNac of about ≤5%, ≤7.5%, ≤10%, ≤15%, ≤20%, ≤25%, ≤30%, ≤40%, ≤50%, ≤75%, 0 - 100%, 0.5 - 100%, 0 - 50%, 0.5 - 50%, 0.5 - 25%, 0.5 - 10%, 0.5 - 7.5%, or 0.9 - 5.3%; a level of G2 of 0 - 100%, 1 - 100%, or 39 - 92%; and / or a level of G0-2GlcNac of 0 - 100%, 1 - 100%, or 38 - 88%.

[0215] In one embodiment, the composition comprises a mixture of antibodies, where 100% are afucosylated. In another embodiment, the composition comprises a mixture of antibodies, where 0% are fucosylated.

[0216] In one example, glycosylation modifications and the resulting profiles can be determined using ultra - performance liquid chromatography (UPLC) with hydrophilic interaction liquid chromatography (HILIC) separation and fluorescence detection. In one example, a composition described herein, for example, a composition comprising belantamab, can be diluted with water to a concentration of 10 μg / μL, and then the glycans can be released from the composition comprising belantamab, for example, by enzymatic digestion with PNGaseF using a PNGaseF kit (Cat #P0705L) from New England BioLabs. The glycans can be released by PNGaseF and labeled with o - aminobenzamide (Sigma - Aldrich, Cat #A89804). The labeled glycans can then be purified using an HILIC column step to remove excess labeling solution; the glycans can be loaded and washed with water and eluted with acetonitrile. The labeled glycans can then be separated using a Waters Glycan BEH Amide column (part number 186004742) on a Waters Acquity UPLC with an ammonium formate / formic acid and acetonitrile gradient. The glycans can then be detected, for example, by fluorescence detection using excitation at 365 nm and emission at 438 nm. Quantification of the glycans can be achieved, for example, by dividing the area under the curve of the glycans by the total area under the curve of all detected glycans.

[0217] In one aspect, the composition comprises antibodies that are aggregated antibodies (high - molecular - weight (HMW) species), also referred to herein as "aggregation variants". Aggregated antibodies can comprise dimers or higher - order structures formed from antibody monomers and their subunits. Aggregation variants can be, for example, covalent or non - covalent, reducible or non - reducible, and visible or only sub - visibly visible aggregates of the antibodies disclosed herein. Aggregated or fragmented variants can be characterized and distinguished from the antibodies based on their size. For example, size - exclusion chromatography (SEC), such as SE - HPLC, can be used to detect the size distribution of the antibody composition.

[0218] In one aspect, the composition comprises an aggregated variant of an antibody, wherein the aggregated variant comprises a heavy chain sequence comprising CDRH1 of SEQ ID NO:1, CDRH2 of SEQ ID NO: 2, and CDRH3 of SEQ ID NO: 3, and a light chain sequence comprising CDRL1 of SEQ IDNO: 4, CDRL2 of SEQ ID NO: 5, and CDRL3 of SEQ ID NO: 6; wherein the composition comprises ≤10% of the aggregated variant.

[0219] The antibody composition may comprise ≤10% of the aggregated variant, such as ≤7.5%, ≤5%, ≤4%, ≤3%, ≤2% or ≤1% of the aggregated variant. In another embodiment, the composition may comprise 1-10%, 1-5%, 1-4%, 1-3% or 1-2% of the aggregated variant. Alternatively, the composition comprises more than 1% and less than 10% of the aggregated variant. Alternatively, the composition may comprise about 7.5%, about 5%, about 4%, about 3%, about 2% or about 1% of the aggregated variant.

[0220] A fragmented variant (“fragment variant”) is a variant that comprises a portion of a full-length antibody. For example, such fragments include Fab, Fab’, F(ab’)2 and Fv fragments, diabodies, linear antibodies, single-chain antibody molecules, and individual variable domains of immunoglobulins.

[0221] In one aspect, the composition comprises a fragment variant of an antibody, wherein the fragment variant comprises a heavy chain sequence comprising CDRH1 of SEQ ID NO:1, CDRH2 of SEQ ID NO: 2, and CDRH3 of SEQ ID NO: 3, and a light chain sequence comprising CDRL1 of SEQ IDNO: 4, CDRL2 of SEQ ID NO: 5, and CDRL3 of SEQ ID NO: 6; wherein the composition comprises ≤10% of the fragment variant.

[0222] The antibody composition may comprise ≤10% of the fragmented antibody, such as ≤5%, ≤4%, ≤3%, ≤2% or ≤1% of the fragmented antibody. In another embodiment, the composition may comprise 0.5-10%, 0.5-5%, 0.5-4%, 0.5-3%, 0.5-2%, 0.5-1.5% or 0.5-1% of the fragmented antibody. Alternatively, the composition may comprise about 5%, about 4%, about 3%, about 2%, about 1% or about 0.5% of the fragmented antibody.

[0223] The composition may comprise acidic, basic, isomerized, oxidized, deamidated, N-terminal pyroglutamic acid, C-terminal lysine cleavage variants, and / or any percentage of glycosylation modification variants, and / or any one or combination of aggregated and / or fragmented variants as described herein.

[0224] In one embodiment, the composition has ≥70% BCMA-specific antigen binding, ≥70% FcγRIIIa binding, and / or ≥70% FcRn binding.

[0225] In another embodiment, the composition has about ≥75%, ≥80%, ≥85%, ≥90%, or ≥95% BCMA-specific antigen binding. In another embodiment, the composition has about ≥75%, ≥80%, ≥85%, ≥90%, ≥95% FcγRIIIa binding. In another embodiment, the composition has about ≥75%, ≥80%, ≥85%, ≥90%, ≥95% FcRn binding.

[0226] In another embodiment, the composition has specific antigen binding in the range of about 70% to 130%, FcγRIIIa binding in the range of about 70% to 130%, and / or FcRn binding in the range of about 70% to 130%.

[0227] In some embodiments, the composition has specific antigen binding in the range of about 75% to about 125%, about 80% to about 120%, about 90% to about 110%, about 70%, about 80%, about 90%, or 100%, about 110%, about 120%, or about 130%. In some embodiments, the composition has FcγRIIIa binding in the range of about 75% to about 125%, about 80% to about 120%, about 90% to about 110%, about 90%, about 95%, about 100%, about 105%, or about 110%. In some embodiments, the composition has FcRn binding in the range of about 75% to about 125%, about 80% to about 120%, about 90% to about 110%, about 90%, about 95%, about 100%, about 105%, about 110%.

[0228] In another embodiment, the composition comprising the variant has at least 70%, at least 75%, at least 80%, at least 85%, at least 90% or at least 95% of the activity of belantamab with 100% activity. In one aspect, the composition comprises a variant that includes a heavy chain amino acid sequence comprising CDRH1 of SEQ ID NO: 1, CDRH2 of SEQ ID NO: 2, and CDRH3 of SEQ ID NO: 3, and a light chain amino acid sequence comprising CDRL1 of SEQ ID NO: 4, CDRL2 of SEQ ID NO: 5, and CDRL3 of SEQ ID NO: 6; wherein the composition has at least 70% of the potency of a composition comprising: a heavy chain sequence of SEQ ID NO: 9, 11, 12, 13 or 14 and a light chain sequence of SEQ ID NO: 10 and any one or combination of the following (i) up to 23% isomerization at D103, and / or (ii) up to 37% oxidation at M34.

[0229] In another aspect, the composition comprises a variant comprising a heavy chain amino acid sequence comprising CDRH1 of SEQ ID NO: 1, CDRH2 of SEQ ID NO: 2, and CDRH3 of SEQ ID NO: 3, and a light chain amino acid sequence comprising CDRL1 of SEQ ID NO: 4, CDRL2 of SEQ ID NO: 5, and CDRL3 of SEQ ID NO: 6; wherein the composition has at least 70% of the potency of a composition comprising: a heavy chain sequence of SEQ ID NO: 9, 11, 12, 13 or 14 and a light chain sequence of SEQ ID NO: 10 and any one or combination of the following (i) up to 23% isomerization at D103, (ii) up to 37% oxidation at M34, (iii) up to 64% oxidation at M256, (iv) up to 61% oxidation at M432, (v) up to 100% deamidation at N388, and / or (vi) up to 100% deamidation at N393. In yet another aspect, the composition comprises a variant comprising a heavy chain amino acid sequence comprising CDRH1 of SEQ ID NO: 1, CDRH2 of SEQ ID NO: 2, and CDRH3 of SEQ ID NO: 3, and a light chain amino acid sequence comprising CDRL1 of SEQ ID NO: 4, CDRL2 of SEQ ID NO: 5, and CDRL3 of SEQ ID NO: 6; wherein the composition has at least 70% of the potency of a composition comprising: a heavy chain sequence of SEQ ID NO: 9, 11, 12, 13 or 14 and a light chain sequence of SEQ ID NO: 10 and any one or combination of the following (i) up to 23% isomerization at D103, (ii) up to 37% oxidation at M34, (iii) up to 64% oxidation at M256, (iv) up to 61% oxidation at M432, (v) up to 100% deamidation at N388, (vi) up to 100% deamidation at N393, (vii) up to 100% HC C-terminal lysine cleavage, and / or (viii) up to 100% HC N-terminal pyroglutamic acid.

[0230] In one example, surface plasmon resonance (SPR) is used to measure the binding of belantamab mafodotin to BCMA and FcγRIIIa. Belantamab mafodotin can be diluted to 10 μg / mL with PBST, injected, and captured by Protein A immobilized on a CM5 sensor chip. Then BCMA can be injected and bound to the captured belantamab mafodotin. Next, FcγRIIIa can be injected and bound to the captured belantamab mafodotin. The functional concentrations of belantamab mafodotin binding to BCMA and FcγRIIIa can be calculated from a reference standard curve and reported separately as the BCMA or FcγRIIIa binding concentration. The total belantamab mafodotin concentration of the sample is pre-determined by the absorbance at 280 nm. The specific binding activity (%) can be calculated by dividing the BCMA or FcγRIIIa binding concentration by the absorbance concentration at 280 nm.

[0231] The binding of the neonatal Fc receptor (FcRn) to an anti-BCMA antigen-binding protein (such as belantamab) can be measured using surface plasmon resonance (SPR). Belantamab can be captured by FcRn immobilized on a nitrilotriacetic acid (NTA) sensor chip. The FcRn binding concentration of the sample can be determined by interpolating the binding response on a calibration curve. The specific binding activity (%) is calculated by dividing the FcRn binding concentration by the total protein concentration.

[0232] In other methods known to those skilled in the art, surface plasmon resonance (SPR) can be used to measure the binding of an anti-BCMA antigen-binding protein (such as belantamab mafodotin) to BCMA and FcγRIIIa. In one example, belantamab mafodotin is injected and captured by Protein A immobilized on a CM5 sensor chip. Then BCMA is injected and bound to the captured belantamab mafodotin. Next, FcγRIIIa is injected and bound to the captured belantamab mafodotin. The functional concentrations of belantamab mafodotin binding to BCMA and FcγRIIIa can be calculated from a reference standard curve and reported separately as the BCMA or FcγRIIIa binding concentration. The total belantamab mafodotin concentration of the sample can be pre-determined by the absorbance at 280 nm. The specific binding activity (%) can be calculated by dividing the BCMA or FcγRIIIa binding concentration by, for example, the absorbance concentration at 280 nm.

[0233] In certain embodiments, the average DAR or DL percentage affects the binding to FcRn. In another embodiment, the average DAR or DL percentage does not affect the binding to FcRn. In yet another embodiment, the composition comprises belantamab mafodotin, and the average DAR or DL percentage affects the binding to FcRn. In yet another embodiment, the composition comprises belantamab mafodotin, and the average DAR or DL percentage does not affect the binding to FcRn. In one embodiment, the average DAR or DL percentage may weaken the binding to FcRn.

[0234] Surface plasmon resonance (SPR) measurements can be used to measure the binding of the neonatal Fc receptor (FcRn) to an anti-BCMA antigen-binding protein (such as belantamab mafodotin). Belantamab mafodotin can be captured by FcRn immobilized on a nitrilotriacetic acid (NTA) sensor chip. The FcRn-binding concentration of a sample can be determined by interpolating the binding response on a calibration curve. The specific binding activity (%) is calculated by dividing the FcRn-binding concentration by the total protein concentration.

[0235] When the anti-BCMA antigen-binding protein comprises belantamab mafodotin, the SPR methods described herein for specific antigen binding, FcγRIIIa, and FcRn binding can use a reference standard of belantamab or belantamab mafodotin. The belantamab or belantamab mafodotin reference standard can be used to obtain system suitability and sample comparability data to ensure that the method is performed correctly. The reference standard can allow the establishment of a calibration curve and the interpolation of the concentration of the sample from the curve. For example, the reference standard can be a composition comprising the heavy chain amino acid sequence of SEQ ID NO: 9 and the light chain amino acid sequence of SEQ ID NO: 10

[0236] Antibody compositions comprising the above antibodies and antibody variants retain specific antigen binding and / or FcRn binding and / or FcγRIIIa binding and / or potency. For example, an antibody composition comprising the above antibodies and antibody variants and post-translational modification variants has >0.70 BCMA-specific antigen binding; and / or >70% FcRn binding and / or 70% FcγRIIIa binding and / or >70% potency. Thus, these levels (%) of variants can be tolerated in the antibody composition without significantly affecting function (i.e., not resulting in decreased activity). In one embodiment, "decreased function" or "decreased activity" means that the binding to BCMA or the binding to FcRn or the binding to FcγRIIIa or the potency is decreased by a percentage compared to a reference standard and is significant in assay variability. For example, a decrease in function or activity or potency can be described as a decrease of ≥5%, ≥10%, ≥15%, ≥20%, ≥25%, ≥30%, ≥35%, ≥40%, ≥45%, or ≥50%.

[0237] In another embodiment, the reference sample standard is a composition comprising the heavy chain amino acid sequence of SEQ ID NO: 9 and the light chain amino acid sequence of SEQ ID NO: 10, wherein the composition comprises 80% or more heavy chain C-terminal lysine cleavage and 100% or less heavy chain N-terminal pyroglutamic acid. In a further embodiment, the reference sample standard is a composition comprising the heavy chain amino acid sequence of SEQ ID NO: 9 and the light chain amino acid sequence of SEQ ID NO: 10, wherein the composition comprises 80% or more heavy chain C-terminal lysine cleavage and 100% or less heavy chain N-terminal pyroglutamic acid, and 7% or less isomerization at amino acid D103 in CDRH3. In a further embodiment, the reference sample standard is a composition comprising the heavy chain amino acid sequence of SEQ ID NO: 9 and the light chain amino acid sequence of SEQ ID NO: 10, wherein the composition comprises 80% or more heavy chain C-terminal lysine cleavage and 100% or less heavy chain N-terminal pyroglutamic acid, 7% or less isomerization at amino acid D103 in CDRH3, and 5% or less oxidation at amino acids M34, M256, and / or M432. In a further embodiment, the reference sample standard is a composition comprising the heavy chain amino acid sequence of SEQ ID NO: 9 and the light chain amino acid sequence of SEQ ID NO: 10, wherein the composition comprises 80% or more heavy chain C-terminal lysine cleavage and 100% or less heavy chain N-terminal pyroglutamic acid, 7% or less isomerization at amino acid D103 in CDRH3, 5% or less oxidation at amino acids M34, M256, and / or M432, and 2% or less deamidation at amino acids N388 and / or N393. In a further embodiment, the reference sample standard is a composition comprising the heavy chain amino acid sequence of SEQ ID NO: 9 and the light chain amino acid sequence of SEQ ID NO: 10, wherein the composition comprises 80% or more heavy chain C-terminal lysine cleavage and 100% heavy chain N-terminal pyroglutamic acid, 7% or less isomerization at amino acid D103 in CDRH3, 5% or less oxidation at M256, 2% or less oxidation at M34, M256, and M432, and 2% or less deamidation at amino acids N388 and N393.

[0238] Antibody-drug conjugate (ADC)

[0239] Antibody-drug conjugates (ADCs) are a class of emerging potent anti-cancer agents that have recently shown significant clinical benefit. An ADC consists of a cytotoxic agent chemically conjugated to an antibody via a linker. Presumably, through a series of events including antigen binding at the cell surface, endocytosis, trafficking to the lysosome, ADC degradation, release of the payload, disruption of cellular processes (e.g., mitosis), and apoptosis, an ADC may destroy cancer cells with overexpression of cell surface proteins. ADCs combine the antigen-driven targeting properties of monoclonal antibodies and the potent anti-tumor effects of cytotoxic agents. For example, in 2011 ADCETRIS® (anti-CD30 antibody-MMAE ADC) received regulatory approval for the treatment of refractory Hodgkin lymphoma and systemic anaplastic large cell lymphoma.

[0240] ADCs have been used for the local delivery of cytotoxic agents, i.e., drugs that kill or inhibit cell growth or proliferation, in cancer therapy (Lambert, J. (2005) Curr. Opinion in Pharmacology 5:543-549; Wu et al. (2005) Nature Biotechnology 23(9):1137-1146; Payne, G. (2003) i 3:207-212; Syrigos and Epenetos (1999) Anticancer Research 19:605-614; Niculescu-Duvaz and Springer (1997) Adv. Drug Deliv. Rev. 26:151-172; U.S. Patent No. 4,975,278). ADCs allow for the targeted delivery of the drug moiety to tumors and intracellular accumulation therein, where systemic administration of the unconjugated drug may result in unacceptable levels of toxicity to normal cells as well as the tumor cells sought to be eliminated (Baldwin et al., Lancet (Mar. 15, 1986) pp. 603-05; Thorpe (1985) "Antibody Carriers Of Cytotoxic Agents In Cancer Therapy: A Review," in Monoclonal Antibodies '84: Biological And Clinical Applications (A. Pinchera et al., eds) pp. 475-506). Both polyclonal and monoclonal antibodies have been reported to be useful in these strategies (Rowland et al., (1986) Cancer Immunol. Immunother. 21:183-87).Toxins for use in antibody-toxin conjugates include bacterial toxins such as diphtheria toxin, plant toxins such as ricin, small molecule toxins such as geldanamycin (Mandler et al (2000) J. Nat. Cancer Inst. 92(19):1573-1581; Mandler et al.(2000) Bioorganic & Med. Chem. Letters 10:1025-1028; Mandler et al (2002)Bioconjugate Chem. 13:786-791), maytansinoids (EP 1391213; Liu et al.(1996) Proc. Natl. Acad. Sci. USA 93:8618-8623) and calicheamicin (Lode et al (1998) Cancer Res. 58:2928; Hinman et al. (1993) Cancer Res. 53:3336-3342).

[0241] In one embodiment, the anti-BCMA antigen-binding protein is an antibody-drug conjugate (“anti-BCMA ADC”) that comprises an antibody or antibody fragment conjugated to one or more cytotoxic agents such as chemotherapeutic agents, drugs, growth inhibitors, toxins (e.g., protein toxins, enzymatic active toxins or fragments thereof from bacteria, fungi, plants or animals) or radioisotopes (i.e., radio-conjugates).

[0242] In one embodiment, the anti-BCMA ADC has the following general structure:

[0243] ABP-((linker) n -Ctx) m

[0244] Wherein:

[0245] ABP is an antigen-binding protein, antibody or antibody fragment;

[0246] The linker is absent or is any cleavable or non-cleavable linker;

[0247] Ctx is any cytotoxic agent as described herein;

[0248] n is 0, 1, 2 or 3; and,

[0249] m is 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10.

[0250] In exemplary embodiments, enzyme active toxins and fragments thereof that can be used include diphtheria A chain, the non-binding active fragments of diphtheria toxin, exotoxin A chain (from Pseudomonas aeruginosa), ricin A chain, abrin A chain, modeccin A chain, α-sarcin, Aleurites fordii protein, dianthin protein, Phytolaca americana proteins (PAPI, PAPII, and PAP-S), momordica charantia inhibitor, jatrophin, croton toxin, sapaonaria officinalis inhibitor, gelonin, mitogellin, restrictocin, phenomycin, enomycin, and trichothecenes. See, for example, WO93 / 21232 published October 28, 1993. A variety of radionuclides can be used to produce radiolabeled conjugated antibodies, including, for example 211 At, 212 Bi, 131 I, 131 In, 90 Y or 186 Re.

[0251] The anti-BCMA antibody or fragment thereof of the present invention can also be conjugated with one or more cytotoxic agents, including but not limited to calicheamicin, maytansinoids, dolastatins, auristatins, trichothecenes, and CC1065, or derivatives of these toxins with toxin activity. Suitable cytotoxic agents include, for example, auristatins, including methylvaline-valine-dolaisoleucine-dolaproline-phenylalanine (MMAF) and monomethyl auristatin E (MMAE) and ester forms of MMAE, DNA minor groove binders, DNA minor groove alkylating agents, enediynes, lexitropsin, duocarmycin, taxanes, including paclitaxel and docetaxel, puromycin, dolastatin, maytansine, and vinca alkaloids. Specific cytotoxic agents include topotecan, morpholino-doxorubicin, rhizoxin, cyanomorpholino-doxorubicin, dolastatin-10, echinosporin, combretastatin, chalicheamicin, maytansine, DM-1, DM-4, netropsin. Other suitable cytotoxic agents include antimicrotubule agents, such as auristatins, vinca alkaloids, podophyllotoxin, taxanes, baccatin derivatives, cryptophycins, maytansine, combretastatin, or dolastatin. Antimicrotubule agents include dimethylvaline-valine-dolaisoleucine-dolaproline-phenylalanine-p-phenylenediamine (AFP), MMAF, MMAE, auristatin E, vincristine, vinblastine, vindesine, vinorelbine, VP-16, camptothecin, paclitaxel, docetaxel, epothilone A, epothilone B, nocodazole, colchicine, colchicine amide, estramustine, zimadotin, discodermolide, maytansine, DM-1, DM-4, or eleutherobin.

[0252] In one embodiment, the anti-BCMA ADC comprises an anti-BCMA antibody conjugated with MMAE or MMAF.

[0253]

[0254]

[0255] Exemplary linkers include cleavable and non-cleavable linkers. Cleavable linkers may be susceptible to cleavage under intracellular conditions. Suitable cleavable linkers include, for example, peptide linkers that can be cleaved by intracellular proteases such as lysosomal or endosomal proteases. In an exemplary embodiment, the linker can be a dipeptide linker such as a valine-citrulline (val-cit) or phenylalanine-lysine (phe-lys) linker. Other suitable linkers include, for example, linkers that are hydrolysable at a pH of less than 5.5 such as hydrazone linkers. Other suitable cleavable linkers include, for example, disulfide linkers. Exemplary linkers include 6-maleimidocaproyl (MC), maleimidopropionyl (MP), valine-citrulline (val-cit), alanine-phenylalanine (ala-phe), p-aminobenzyloxycarbonyl (PAB), N-succinimidyl 4-(2-pyridylthio)pentanoate (SPP), N-succinimidyl 4-(N-maleimidomethyl)cyclohexane-1-carboxylate (SMCC), and N-succinimidyl (4-iodo-acetyl)aminobenzoate (SIAB).

[0256] In one embodiment, the linker can comprise a thiol-reactive maleimide, a hexanoyl spacer, the dipeptide valine-5-citrulline, p-aminobenzyloxycarbonyl, a self-immolative fragment group, or a protease-resistant maleimidocaproyl.

[0257] In another embodiment, the anti-BCMA ADC comprises an anti-BCMA antibody linked to MMAE or MMAF via an MC linker as shown in the following structure:

[0258]

[0259]

[0260] The anti-BCMA ADCs described herein can comprise any anti-BCMA antibody described herein and any cytotoxic agent described herein.

[0261] In one embodiment, the anti-BCMA ADC comprises an anti-BCMA antibody that comprises CDRH1 comprising an amino acid sequence having at least about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the amino acid sequence set forth in SEQ ID NO: 1; CDRH2 comprising an amino acid sequence having at least about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the amino acid sequence set forth in SEQ ID NO: 2; CDRH3 comprising an amino acid sequence having at least about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the amino acid sequence set forth in SEQ ID NO: 3; CDRL1 comprising an amino acid sequence having at least about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the amino acid sequence set forth in SEQ ID NO: 4; CDRL2 comprising an amino acid sequence having at least about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the amino acid sequence set forth in SEQ ID NO: 5; and / or CDRL3 comprising an amino acid sequence having at least about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the amino acid sequence set forth in SEQ ID NO: 6; and is conjugated to MMAE or MMAF.

[0262] In yet another embodiment, the anti-BCMA ADC comprises an anti-BCMA antibody that comprises CDRH1 having the amino acid sequence set forth in SEQ ID NO: 1; CDRH2 having the amino acid sequence set forth in SEQ ID NO: 2; CDRH3 having the amino acid sequence set forth in SEQ ID NO: 3; CDRL1 having the amino acid sequence set forth in SEQ ID NO: 4; CDRL2 having the amino acid sequence set forth in SEQ ID NO: 5; and CDRL3 having the amino acid sequence set forth in SEQ ID NO: 6; and is conjugated to MMAF or MMAE.

[0263] In one embodiment, the anti-BCMA ADC comprises an anti-BCMA antibody that comprises V H, which comprises an amino acid sequence having at least about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the amino acid sequence set forth in SEQ ID NO: 7; and / or V L , which comprises an amino acid sequence having at least about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the amino acid sequence set forth in SEQ ID NO: 8; and is conjugated to MMAE or MMAF.

[0264] In yet another embodiment, the anti-BCMA ADC comprises an anti-BCMA antibody that comprises a V H having the amino acid sequence set forth in SEQ ID NO: 7; and a V L having the amino acid sequence set forth in SEQ ID NO: 8; and is conjugated to MMAF or MMAE.

[0265] In one embodiment, the anti-BCMA ADC comprises an anti-BCMA antibody that comprises an HC that comprises an amino acid sequence having at least about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the amino acid sequence set forth in SEQ ID NO: 9; and / or an LC that comprises an amino acid sequence having at least about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to the amino acid sequence set forth in SEQ ID NO: 10; and is conjugated to MMAF or MMAE.

[0266] In yet another embodiment, the anti-BCMA ADC is belantamab mafodotin comprising an anti-BCMA antibody that comprises an HC having the amino acid sequence set forth in SEQ ID NO: 9 and an LC having the amino acid sequence set forth in SEQ ID NO: 10; and is conjugated to MMAF.

[0267] Preparation and characterization of ADC

[0268] Some native IgG1 molecules contain 16 disulfide bonds (32 cysteine or thiol groups). In some aspects, an antibody can be reduced in such a way that only four inter-chain disulfide bonds are reduced and conjugated to a cytotoxic agent, allowing for up to eight attachment sites for the cytotoxic agent. In other words, the drug payload (“DL”), i.e., the number of cytotoxic agents per antibody molecule, can range from 0 to 8 and is described herein as DL0, DL2 (including DL2a and DL2b), DL4 (including DL4a, DL4b, and DL4c), DL6 (including DL6a and DL6b), and DL8.

[0269] For a given ADC composition, the conjugation process can result in heterogeneity in drug-antibody attachment, which can vary in 1) the number of drugs bound to each antibody molecule and 2) the location of the cytotoxic agent. This can result in an ADC composition having various DL species as Figure 1 illustrated. As used herein, the term “ADC composition” refers to a composition containing a heterogeneous mixture of antibody species that contain various drug loadings (“DL”). (See, e.g., Figure 2 ). The average drug-antibody ratio of the entire heterogeneous ADC composition is referred to herein as the “average DAR” or “DAR”. For example, an ADC composition may contain a mixture of antibody species, each with their own DL (some species in the mixture are DL2, some species in the mixture are DL4, some species in the mixture are DL6, some species in the mixture are DL8), and the average DAR of the entire composition may be about 4.

[0270] In another embodiment, the term “DL percentage” can be used to describe the percentage of a particular DL species in a heterogeneous ADC composition (e.g., the DL2 percentage is about 10% to about 30% of the total heterogeneous ADC composition).

[0271] In certain aspects of the present invention, a drug can be conjugated to an antibody via a thiol group on the antibody. The thiol group can be a thiol group on a cysteine side chain. The cysteine residue can be naturally present in the antibody (e.g., interchain disulfides) or introduced by other means, such as mutagenesis. Methods for conjugating a drug to a thiol group on an antibody are well known in the art (see, e.g., U.S. Patent Nos. 7,659,241, 7,498,298 and International Publication Nos. WO 2011 / 130613, WO 2014 / 152199, WO 2015 / 077605 and Bioconjugate Chem. 2005, 16, 1282-1290). The antibody is typically reduced prior to conjugation to make the thiol groups available for conjugation. Antibodies can be reduced using conditions known in the art. The reducing conditions are those that generally do not cause any substantial denaturation of the antibody and generally do not affect the antigen-binding affinity of the antibody.

[0272] In one aspect of the present invention, the reducing agent used in the reduction step is TCEP (tris(2-carboxyethyl)phosphine), and TCEP is added, for example, in excess for 30 minutes at room temperature. For example, at room temperature, a 10 mM solution (pH 7.4) of 250 µL of TCEP will readily reduce the interchain disulfides of 1 to 100 ug of antibody within 30 minutes. However, other reducing agents and conditions can be used. Examples of reaction conditions include temperatures from 5°C to 37°C in the pH range from 5 to 8.

[0273] There are a variety of methods and are known to those skilled in the art for calculating the percentage of DL species and / or the average DAR in an ADC composition. For example, the heterogeneity of a cysteine-linked ADC is typically measured by hydrophobic interaction chromatography (HIC), which separates DL species based on the amount of drug loaded. LC-MS assays have also been developed to evaluate the DL distribution. Exemplary methods for calculating the drug payload distribution in an ADC composition can be found, for example, in Journal of Chromatography B 1060 (2017) 182-18.

[0274] For example, DL0 has no drug payload on the antibody. For example, DL2 has a drug payload of 2. In one embodiment, the conjugation sites of DL2 are LC C214 and HC 224. For example, DL4 has a drug payload of 4. In one embodiment, the conjugation sites of DL4a are LC C214, HC 224, LC C214 and HC 224. In one embodiment, the conjugation sites of DL4b are LC C230, HC 233, LC C230 and HC 233. For example, DL6 has a drug payload of 6. In one embodiment, the conjugation sites of DL6 are LC C214, HC 224, LC C230, HC 233, LC C230 and HC 233. For example, DL8 has a drug payload of 8. In one embodiment, the conjugation sites of DL8 are LC C214, HC 224, LC C214, HC 224, LC C230, HC 233, LC C230 and HC 233.

[0275] In one embodiment, the percentage of a specific DL species (e.g., DL0 percentage, DL2 percentage, DL4a percentage, DL4b percentage, DL6 percentage, DL8 percentage) can be determined by separating individual DL species using hydrophobic interaction chromatography (HIC), calculating the area under the curve of each DL peak, and dividing each DL peak by the total area under the curve of all combined DL species. In one embodiment, the average DAR can be calculated from the area under the curve of each DL species using the following formula:

[0276]

[0277] In one embodiment, the percentage of a specific DAR subspecies (e.g., the percentage of DL2a in total DL2) is determined by collecting a specific DL species using a combination of analytical techniques that can include HIC, non-reducing separation methods, and mass spectrometry techniques.

[0278] In one embodiment, the anti-BCMA ADC composition has an average DAR of from about 2 to about 7, from about 2 to about 6, from about 2.1 to about 5.7, from about 2.1 to about 5.0, from about 2.1 to about 4.6, from about 2.1 to about 4.1, from about 2.1 to about 3.5, from about 2.1 to about 3.0, from about 3.0 to about 5.7, from about 3.0 to about 5.0, from about 3.0 to about 4.6, from about 3.0 to about 4.1, from about 3.0 to about 3.5, from about 3.5 to about 5.7, from about 3.5 to about 5.0, from about 3.5 to about 4.6, from about 3.5 to about 4.1, from about 3.8 to about 4.5, from about 4.1 to about 5.7, from about 4.1 to about 5.0, from about 4.1 to about 4.6, from about 4.6 to about 5.7, from about 4.6 to about 5.0, from about 5.0 to about 5.7, about 2.1, about 3.0, about 3.5, about 4.1, about 4.6, about 5.0, or about 5.7.

[0279] In another embodiment, the composition comprises an anti-BCMA ADC, wherein the average DAR is from about 2.1 to about 5.7, from about 3.4 to about 4.6, from about 3.8 to about 4.5, or about 4.

[0280] In one embodiment, the composition comprises an anti-BCMA ADC, wherein the antibody comprises a CDRH1 having the amino acid sequence set forth in SEQ ID NO: 1, a CDRH2 having the amino acid sequence set forth in SEQ ID NO: 2, a CDRH3 having the amino acid sequence set forth in SEQ ID NO: 3, a CDRL1 having the amino acid sequence set forth in SEQ ID NO: 4, a CDRL2 having the amino acid sequence set forth in SEQ ID NO: 5, and a CDRL3 having the amino acid sequence set forth in SEQ ID NO: 6; wherein the cytotoxic agent is MMAE or MMAF; and wherein the average DAR is from about 2 to about 6, from about 2.1 to about 5.7, from about 3.4 to about 4.6, or from about 3.8 to about 4.5.

[0281] In one embodiment, the composition comprises an anti-BCMA ADC, wherein the antibody comprises a V having the amino acid sequence set forth in SEQ ID NO: 7 H and a V having the amino acid sequence set forth in SEQ ID NO: 8 L ; wherein the cytotoxic agent is MMAE or MMAF; and wherein the average DAR is from about 2 to about 6, from about 2.1 to about 5.7, from about 3.4 to about 4.6, or from about 3.8 to about 4.5.

[0282] In one embodiment, the composition comprises belantamab mafodotin, wherein the average DAR is from about 2 to about 6, from about 2.1 to about 5.7, from about 3.4 to about 4.6, or from about 3.8 to about 4.5.

[0283] In one embodiment, the percentage of DL0 species in the anti-BCMA ADC composition is about 10% or less, about 5% or less, about 1% to about 10%, about 1% to about 5%, or about 2.8% to about 4.7%.

[0284] In one embodiment, the percentage of DL2 species in the anti-BCMA ADC composition is at least about 10%, at least about 15%, about 15.8% to about 26.3%, about 15% to about 27%, about 15% to about 32%, or about 10% to about 40%.

[0285] In one embodiment, the percentage of DL4a species in the anti-BCMA ADC composition is at least about 30%, at least about 35%, about 35.5% to about 37.9%, about 35% to about 38%, about 30% to about 40%, or about 20% to about 50%. In another embodiment, the percentage of DL4a species is the major species in the anti-BCMA ADC composition and comprises about ≥30%, ≥40%, ≥50%, ≥60%, ≥70%, ≥80%, or ≥90% of all species combinations.

[0286] In one embodiment, the percentage of DL4b species in the anti-BCMA ADC composition is at least about 5%, at least about 7%, about 7.1% to about 8.5%, about 7% to about 9%, about 5% to about 10%, or about 1% to about 15%.

[0287] In one embodiment, the percentage of DL6 species in the anti-BCMA ADC composition is at least about 10%, at least about 14%, about 14.0% to about 19.1%, about 14% to about 20%, about 10% to about 20%, or about 5% to about 30%.

[0288] In one embodiment, the percentage of DL8 species in the anti-BCMA ADC composition is at least about 1%, at least about 6%, about 6.0% to about 12.0%, about 4% to about 15%, or about 1% to about 20%.

[0289] In one embodiment, the composition comprises an anti-BCMA ADC, wherein the DL2 percentage is about 15% to about 27% or about 15% to about 32%, the DL4a percentage is about 35% to about 38% or about 30% to about 40%, the DL4b percentage is about 7% to about 9% or about 5% to about 10%, the DL6 percentage is about 14% to about 20% or about 10% to about 20%, and / or the DL8 is about 6.0% to about 12.0% or about 4% to about 15%.

[0290] In one embodiment, the composition comprises an anti-BCMA ADC, wherein the antibody comprises a CDRH1 having the amino acid sequence set forth in SEQ ID NO: 1, a CDRH2 having the amino acid sequence set forth in SEQ ID NO: 2, a CDRH3 having the amino acid sequence set forth in SEQ ID NO: 3, a CDRL1 having the amino acid sequence set forth in SEQ ID NO: 4, a CDRL2 having the amino acid sequence set forth in SEQ ID NO: 5, and a CDRL3 having the amino acid sequence set forth in SEQ ID NO: 6; wherein the cytotoxic agent is MMAE or MMAF; and wherein the DL2 percentage is from about 15% to about 27% or from about 15% to about 32%, the DL4a percentage is from about 35% to about 38% or from about 30% to about 40%, the DL4b percentage is from about 7% to about 9% or from about 5% to about 10%, the percentage DL6 is from about 14% to about 20% or from about 10% to about 20%, and / or the DL8 is from about 6.0% to about 12.0% or from about 4% to about 15%.

[0291] In one embodiment, the composition comprises an anti-BCMA ADC, wherein the antibody comprises a V having the amino acid sequence set forth in SEQ ID NO: 7 H and a V having the amino acid sequence set forth in SEQ ID NO: 8 L ; wherein the cytotoxic agent is MMAE or MMAF; and wherein the DL2 percentage is from about 15% to about 27% or from about 15% to about 32%, the DL4a percentage is from about 35% to about 38% or from about 30% to about 40%, the DL4b percentage is from about 7% to about 9% or from about 5% to about 10%, the percentage DL6 is from about 14% to about 20% or from about 10% to about 20%, and / or the DL8 is from about 6.0% to about 12.0% or from about 4% to about 15%.

[0292] In one embodiment, the composition comprises belantamab mafodotin, wherein the DL2 percentage is from about 15% to about 27% or from about 15% to about 32%, the DL4a percentage is from about 35% to about 38% or from about 30% to about 40%, the DL4b percentage is from about 7% to about 9% or from about 5% to about 10%, the DL6 percentage is from about 14% to about 20% or from about 10% to about 20%, and / or the DL8 is from about 6.0% to about 12.0% or from about 4% to about 15%.

[0293] As used herein, the term "undesired DAR species" refers to any DAR species that is not desired in the final composition and may have a negative impact on certain properties of the final therapeutic product (e.g., target binding, efficacy, safety, etc.). In one embodiment, the undesired DAR species is DL0, i.e., an antibody that does not bind to the cytotoxin after the conjugation process. In one embodiment, the percentage of DL0 in the ADC composition is less than or equal to about 15%, about 14%, about 13%, about 12%, about 11%, about 10%, about 9%, about 8%, about 7%, about 6%, about 5%, about 4%, about 3%, about 2%, about 1% or about 0.5%. In another embodiment, the percentage of DL0 in the ADC composition is from about 1% to about 10%, from about 2% to about 5% or from about 2.0% to about 4.8%.

[0294] In one embodiment, the composition comprises an anti-BCMA ADC, wherein the antibody comprises CDRH1 having the amino acid sequence set forth in SEQ ID NO: 1, CDRH2 having the amino acid sequence set forth in SEQ ID NO: 2, CDRH3 having the amino acid sequence set forth in SEQ ID NO: 3, CDRL1 having the amino acid sequence set forth in SEQ ID NO: 4, CDRL2 having the amino acid sequence set forth in SEQ ID NO: 5, and CDRL3 having the amino acid sequence set forth in SEQ ID NO: 6; wherein the cytotoxin is MMAE or MMAF; wherein the percentage of DL0 is less than or equal to about 10% or about 5%.

[0295] In one embodiment, the composition comprises an anti-BCMA ADC, wherein the antibody comprises V having the amino acid sequence set forth in SEQ ID NO: 7 H and V having the amino acid sequence set forth in SEQ ID NO: 8 L ; wherein the cytotoxin is MMAE or MMAF; wherein the percentage of DL0 is less than or equal to about 10% or about 5%.

[0296] In one embodiment, the composition comprises belantamab mafodotin, wherein the percentage of DL0 is less than or equal to about 10% or about 5%.

[0297] In one embodiment, the composition comprises belantamab mafodotin, wherein the percentage of DL0 is less than or equal to about 10% or about 5%, the percentage of DL2 is from about 15% to about 27% or from about 15% to about 32%, the percentage of DL4a is from about 35% to about 38% or from about 30% to about 40%, the percentage of DL4b is from about 7% to about 9% or from about 5% to about 10%, the percentage of DL6 is from about 14% to about 20% or from about 10% to about 20%, and / or the percentage of DL8 is from about 6.0% to about 12.0% or from about 4% to about 15%.

[0298] In certain embodiments, the average DAR or DL percentage affects cell growth inhibition and / or tumor volume. In certain embodiments, the average DAR does not affect cell growth inhibition and / or tumor volume. In another embodiment, as the average DAR or DL percentage increases, cell growth inhibition increases and / or tumor volume decreases. In yet another embodiment, the composition comprises belantamab mafodotin, and as the average DAR or DL percentage of the composition increases, cancer cell growth inhibition increases and / or tumor volume decreases.

[0299] The relative potency of cell growth inhibition can be determined by measuring the cell viability of a cell line (e.g., a multiple myeloma cell line) after incubation with a composition described herein (e.g., belantamab mafodotin). Cell viability can be measured using a cell viability assay known to those of skill in the art. A dose response (half maximal effective concentration or EC50) can be generated using a non-linear regression logistic model. The ratio of the EC50 of a reference standard to the EC50 of a sample containing the composition can be calculated to determine the relative potency.

[0300] In one embodiment, the composition has a relative potency of cell growth inhibition of from about 0.5 to about 1.3, or from about 0.8 to about 1.1. In another embodiment, the composition comprises an average DAR of from about 2.1 to about 5.7 and has a relative potency of cell growth inhibition of from about 0.5 to about 1.3. In another embodiment, the composition comprises an average DAR of from about 3.0 to about 5.0 or from about 3.5 to about 4.6 and has a relative potency of cell growth inhibition of from about 0.8 to about 1.1. In another embodiment, the composition comprises belantamab mafodotin having an average DAR of from about 3.0 to about 5.0 or from about 3.5 to about 4.6 and has a relative potency of cell growth inhibition of from about 0.8 to about 1.1.

[0301] In certain embodiments, the average DAR or DL percentage affects ADCC activity. In another embodiment, the average DAR or DL percentage does not affect ADCC activity. In yet another embodiment, the composition comprises belantamab mafodotin and the average DAR or DL percentage affects ADCC activity. In yet another embodiment, the composition comprises belantamab mafodotin and the average DAR or DL percentage does not affect ADCC activity.

[0302] For example, the relative potency of ADCC activity can be measured by incubating belantamab mafodotin, cells (such as multiple myeloma cells), and NK cells (effector cells). Without being bound by theory, belantamab mafodotin binds to BCMA expressed on the cell surface, and the Fc region of the antibody binds to FcγRIIIa on the effector cells through its FcγRIIIa receptor. The binding of these receptors to the effector cell surface results in the synthesis and secretion of cytokines (IFNg), as well as the release of granules (perforin and granzymes) that enter the cytoplasm of the target cells. Granzymes initiate signaling events within the target cells, leading to the death of these cells by apoptosis. The source of NK cells can be peripheral blood mononuclear cells (PBMCs) isolated from human whole blood. Then, BATDA (bis-(acetoxymethyl) 2,2’:6’,2’’-terpyridine-6,6’’-dicarboxylate) can be added to penetrate the target cell membrane to label the cells. After cell lysis, the ligand can associate with the DELFIA europium solution to form a highly fluorescent and stable chelate (EuTDA). The measured signal is directly related to the amount of lysed cells. The ADCC activity can then be reported as the ratio of the sample EC50 value to the reference standard value.

[0303] In one embodiment, the composition has a relative potency of ADCC activity of about 0.70 to about 1.30, or about 0.8 to about 1.1. In another embodiment, the composition has an average DAR of about 2.1 to about 5.7 and a relative potency of ADCC activity of about 0.5 to about 1.3. In another embodiment, the composition has an average DAR of about 3.0 to about 5.0 or about 3.5 to about 4.6 and a relative potency of ADCC activity of about 0.8 to about 1.1. In another embodiment, the composition comprises belantamab mafodotin having an average DAR of about 3.0 to about 5.0 or about 3.5 to about 4.6 and a relative potency of ADCC activity of about 0.8 to about 1.1.

[0304] In certain embodiments, the average DAR or DL percentage affects the binding to BCMA. In another embodiment, the average DAR or DL percentage does not affect the binding to BCMA. In yet another embodiment, the composition comprises belantamab mafodotin and the average DAR or DL percentage affects the binding to BCMA. In yet another embodiment, the composition comprises belantamab mafodotin and the average DAR or DL percentage does not affect the binding to BCMA. In one embodiment, the average DAR or DL percentage may weaken the binding to BCMA.

[0305] In one embodiment, the composition has a relative BCMA-specific antigen binding of ≥70%, ≥75%, ≥80%, ≥85%, ≥90%, ≥95%. In another embodiment, the composition comprises belantamab mafodotin and has a relative BCMA-specific antigen binding of greater than 85% or greater than 90%. In another embodiment, the composition comprises belantamab mafodotin, has an average DAR of about 2.1 to about 5.7 or about 3.0 to about 5.0 or about 3.5 to about 4.6, and has a relative BCMA-specific antigen binding of greater than 85% or greater than 90%.

[0306] In certain embodiments, the average DAR or DL percentage affects the binding to FcγRIIIa. In another embodiment, the average DAR or DL percentage does not affect the binding to FcγRIIIa. In another embodiment, the composition comprises belantamab mafodotin and the average DAR or DL percentage affects the binding to FcγRIIIa. In yet another embodiment, the composition comprises belantamab mafodotin and the average DAR or DL percentage does not affect the binding to FcγRIIIa. In one embodiment, the average DAR or DL percentage may weaken the binding to FcγRIIIa.

[0307] In one embodiment, the composition has a relative FcγRIIIa binding of ≥70%, ≥75%, ≥80%, ≥85%, ≥90%, ≥95%. In another embodiment, the composition comprises belantamab mafodotin and has a relative FcγRIIIa binding of greater than 85% or greater than 90%. In another embodiment, the composition comprises belantamab mafodotin, has an average DAR of about 2.1 to about 5.7 or about 3.0 to about 5.0 or about 3.5 to about 4.6, and has a relative FcγRIIIa binding of greater than 85% or greater than 90%.

[0308] Among other methods known to those skilled in the art, surface plasmon resonance (SPR) can be used to measure the binding of an anti-BCMA antigen-binding protein (such as belantamab mafodotin) to BCMA and FcγRIIIa. In one example, belantamab mafodotin is injected and captured by Protein A immobilized on a CM5 sensor chip. Then BCMA is injected and binds to the captured belantamab mafodotin. Next, FcγRIIIa is injected and binds to the captured belantamab mafodotin. The functional concentrations of belantamab mafodotin binding to BCMA and FcγRIIIa can be calculated from a reference standard curve and reported separately as the BCMA or FcγRIIIa binding concentration. The total belantamab mafodotin concentration of the sample can be pre-determined by absorbance at 280 nm. The specific binding activity (%) can be calculated by dividing the BCMA or FcγRIIIa binding concentration by, for example, the absorbance concentration at 280 nm.

[0309] In certain embodiments, the average DAR or DL percentage affects the binding to FcRn. In another embodiment, the average DAR or DL percentage does not affect the binding to FcRn. In yet another embodiment, the composition comprises belantamab mafodotin and the average DAR or DL percentage affects the binding to FcRn. In yet another embodiment, the composition comprises belantamab mafodotin and the average DAR or DL percentage does not affect the binding to FcRn. In one embodiment, the average DAR or DL percentage may weaken the binding to FcRn.

[0310] Surface plasmon resonance (SPR) can be used to measure the binding of the neonatal Fc receptor (FcRn) to an anti-BCMA antigen-binding protein, such as belantamab mafodotin. Belantamab mafodotin can be captured by FcRn immobilized on a nitrilotriacetic acid (NTA) sensor chip. The FcRn binding concentration of the sample can be determined by interpolating the binding response on a calibration curve. The specific binding activity (%) is calculated by dividing the FcRn binding concentration by the total protein concentration.

[0311] When the anti-BCMA antigen-binding protein comprises belantamab mafodotin, the SPR methods described herein for specific antigen binding, FcγRIIIa and FcRn binding can use a reference standard of belantamab or belantamab mafodotin. The belantamab or belantamab mafodotin reference standard can be used in the assay to obtain system suitability and sample comparability data to ensure that the method is performed correctly. The reference standard can allow the establishment of a calibration curve and the interpolation of the concentration of the sample from the curve. For example, the reference standard can be a composition comprising the heavy chain amino acid sequence of SEQ ID NO: 9 and the light chain amino acid sequence of SEQ ID NO: 10 and having a known DL and / or average DAR content.

[0312] Exemplary reference standards can include samples of belantamab mafodotin having known DL species components / amounts and / or average DAR.

[0313] Pharmaceutical composition

[0314] The compositions described herein can be in the form of pharmaceutical compositions. A "pharmaceutical composition" can contain the compositions described herein (i.e., the active ingredient) and one or more pharmaceutically acceptable excipients. The excipients must be acceptable in the sense of being compatible with the other ingredients of the formulation, capable of being made into a pharmaceutical formulation, not harmful to its recipient, and / or not interfering with the efficacy of the active ingredient.

[0315] As used herein, "pharmaceutically acceptable excipients" can include any and all solvents, diluents, carriers, dispersion media, coatings, antibacterial and antifungal agents, isotonic agents, and / or absorption delaying agents. Examples of pharmaceutically acceptable excipients include one or more of buffers, water, saline, phosphate buffered saline, dextrose, glycerol, ethanol, etc., and combinations thereof. In many cases, it is preferred to include an isotonic agent in the composition, such as a polyol, sugar, polyol such as mannitol, sorbitol, or sodium chloride. Preservatives; co-solvents; antioxidants, including ascorbic acid and methionine; chelating agents, such as EDTA; metal complexes (e.g., Zn2+-protein complexes); biodegradable polymers; and / or salt-forming counterions, such as sodium or potassium.

[0316] The exact nature of the excipient or other materials may depend on the route of administration, e.g., oral, rectal, nasal, topical (including buccal and sublingual), vaginal, parenteral (including subcutaneous, intramuscular, intravenous, intradermal), intrathecal, and epidural), and intratumoral. It should be understood that the preferred excipients can vary, e.g., with the condition of the recipient and the disease to be treated.

[0317] A mixture of excipients and their respective concentrations together form a "pharmaceutical formulation" (or "formulation"). The formulation can be in liquid form or lyophilized form. The composition in the liquid formulation can be filled into a container and frozen. In certain embodiments, aliquots of the frozen formulation containing the composition can be lyophilized. The lyophilized product can be reconstituted by adding water or other aqueous solutions to produce a reconstituted formulation containing the composition.

[0318] In some embodiments, the anti-BMCA antigen-binding protein is present in the formulation at a concentration of at least about 10 mg / mL or at least about 20 mg / mL. In some embodiments, the anti-BMCA antigen-binding protein is present in the formulation at a concentration of about 20 mg / mL to about 100 mg / mL or about 20 mg / mL to about 60 mg / mL. In certain embodiments, the concentration of the anti-BCMA antigen-binding protein in the formulation is about 20 mg / mL, about 25 mg / mL, about 50 mg / mL, about 60 mg / mL, or about 100 mg / mL. In one embodiment, the anti-BMCA antigen-binding protein is present in the liquid formulation at a concentration of about 20 mg / mL or about 25 mg / mL. In another embodiment, the anti-BMCA antigen-binding protein is present in the lyophilized formulation at a concentration of about 50 mg / mL or about 60 mg / mL. In yet another embodiment, the anti-BMCA antigen-binding protein is present in the reconstituted formulation at a concentration of about 50 mg / mL.

[0319] In certain embodiments, the buffer is a citrate buffer. The citrate buffer can be achieved, for example, by using a conjugate acid / conjugate base system (sodium citrate / citric acid) or by HCl titration of a sodium citrate solution. In certain embodiments, the concentration of the citrate buffer is about 10 mM to about 30 mM. In a preferred embodiment, the concentration of the citrate buffer is 25 mM. In some embodiments, the buffer is a histidine buffer at a concentration of about 5 mM to about 35 mM.

[0320] A buffer can be used to help maintain a preferred pH range. In certain embodiments, the pH of the formulation is about 5.5 to about 7 or about 5.9 to about 6.5, with a preferred pH of 6.2.

[0321] In some embodiments, the formulation contains a polyol. In some embodiments, the polyol is a sugar, preferably a non-reducing sugar. In some embodiments, the non-reducing sugar is trehalose. In some embodiments, the formulation contains trehalose in the range of about 120 mM to about 240 mM. In yet another embodiment, the formulation contains about 200 mM of trehalose.

[0322] In one embodiment, the formulation contains a chelating agent. In another embodiment, the chelating agent is EDTA. In certain embodiments, the formulation contains EDTA at a concentration of 0.01 mM to about 0.1 mM. In yet another embodiment, the formulation contains EDTA at a concentration of 0.05 mM.

[0323] In some embodiments, the formulation comprises a surfactant. A "surfactant" is a surface-active agent that, due to its chemical composition, contains both hydrophilic and hydrophobic groups and can act at the surface of solid-solid, solid-liquid, liquid-liquid, and liquid-gas interfaces. Surfactants can reduce the concentration of proteins in dilute solutions at the air-water and / or water-solid interfaces where the proteins can be adsorbed and potentially aggregated. Surfactants can bind to the hydrophobic interfaces in the protein formulation. Some pharmaceutically acceptable nonionic surfactants comprise polysorbates or polyether groups. Polysorbate 20 and 80 are suitable surfactant stabilizers in the formulations of the present invention. In some embodiments, the formulation comprises from about 0.01% to about 0.05% of polysorbate 20 or polysorbate 80. In yet another embodiment, the formulation comprises about 0.02% of polysorbate 20 or polysorbate 80. In a preferred embodiment, the formulation comprises about 0.02% of polysorbate 80.

[0324] One aspect of the present invention relates to a formulation comprising from about 20 mg / mL to about 100 mg / mL of an anti-BCMA antigen-binding protein, from about 10 mM to about 25 mM of a buffer, from about 120 mM to about 240 mM of a polyol, and a pH in the range of 5.5 to 6.5.

[0325] In one embodiment, the formulation comprises from about 20 mg / mL to about 60 mg / mL of an anti-BCMA antigen-binding protein, from about 10 mM to about 30 mM of a citrate buffer, from about 120 mM to about 240 mM of trehalose, from about 0.01 mM to about 0.1 mM of EDTA, from about 0.01% to about 0.05% of polysorbate 20 or polysorbate 80, and a pH of from about 5.9 to about 6.5.

[0326] In one embodiment, the composition comprises an antibody in a formulation, wherein the antibody comprises a CDRH1 having the amino acid sequence set forth in SEQ ID NO: 1, a CDRH2 having the amino acid sequence set forth in SEQ ID NO: 2, a CDRH3 having the amino acid sequence set forth in SEQ ID NO: 3, a CDRL1 having the amino acid sequence set forth in SEQ ID NO: 4, a CDRL2 having the amino acid sequence set forth in SEQ ID NO: 5, and a CDRL3 having the amino acid sequence set forth in SEQ ID NO: 6; and wherein the formulation comprises from about 20 mg / mL to about 60 mg / mL of the antibody, from about 10 mM to about 30 mM of a citrate buffer, from about 120 mM to about 240 mM of trehalose, from about 0.01 mM to about 0.1 mM of EDTA, from about 0.01% to about 0.05% of polysorbate 20 or polysorbate 80, and has a pH of from about 5.9 to about 6.5.

[0327] In one embodiment, the composition comprises an antibody in a formulation, wherein the antibody comprises a V having the amino acid sequence set forth in SEQ ID NO: 7 H and a V having the amino acid sequence set forth in SEQ ID NO: 8 L ; and wherein the formulation comprises from about 20 mg / mL to about 60 mg / mL of the antibody, from about 10 mM to about 30 mM of a citrate buffer, from about 120 mM to about 240 mM of trehalose, from about 0.01 mM to about 0.1 mM of EDTA, from about 0.01% to about 0.05% of polysorbate 20 or polysorbate 80, and has a pH of from about 5.9 to about 6.5.

[0328] In one embodiment, the composition comprises an antibody in a formulation, wherein the antibody is belantamab; and wherein the formulation comprises from about 20 mg / mL to about 60 mg / mL of belantamab, from about 10 mM to about 30 mM of a citrate buffer, from about 120 mM to about 240 mM of trehalose, from about 0.01 mM to about 0.1 mM of EDTA, from about 0.01% to about 0.05% of polysorbate 20 or polysorbate 80, and has a pH of from about 5.9 to about 6.5.

[0329] In one embodiment, the composition comprises an ADC in a formulation, wherein the antibody comprises a CDRH1 having the amino acid sequence set forth in SEQ ID NO: 1, a CDRH2 having the amino acid sequence set forth in SEQ ID NO: 2, a CDRH3 having the amino acid sequence set forth in SEQ ID NO: 3, a CDRL1 having the amino acid sequence set forth in SEQ ID NO: 4, a CDRL2 having the amino acid sequence set forth in SEQ ID NO: 5, and a CDRL3 having the amino acid sequence set forth in SEQ ID NO: 6; wherein the cytotoxin is MMAE or MMAF; and wherein the formulation comprises from about 20 mg / mL to about 60 mg / mL of the ADC, from about 10 mM to about 30 mM of a citrate buffer, from about 120 mM to about 240 mM of trehalose, from about 0.01 mM to about 0.1 mM of EDTA, from about 0.01% to about 0.05% of polysorbate 20 or polysorbate 80, and has a pH of from about 5.9 to about 6.5.

[0330] In one embodiment, the composition comprises an ADC in a formulation, wherein the antibody comprises a V having the amino acid sequence set forth in SEQ ID NO: 7 H and a V having the amino acid sequence set forth in SEQ ID NO: 8 L ; wherein the cytotoxin is MMAF or MMAE; and wherein the formulation comprises from about 20 mg / mL to about 60 mg / mL of the ADC, from about 10 mM to about 30 mM of a citrate buffer, from about 120 mM to about 240 mM of trehalose, from about 0.01 mM to about 0.1 mM of EDTA, from about 0.01% to about 0.05% of polysorbate 20 or polysorbate 80, and has a pH of from about 5.9 to about 6.5.

[0331] In one embodiment, the composition comprises an ADC in a formulation, wherein the ADC is belantamab mafodotin; and wherein the formulation comprises from about 20 mg / mL to about 60 mg / mL of belantamab mafodotin, from about 10 mM to about 30 mM of a citrate buffer, from about 120 mM to about 240 mM of trehalose, from about 0.01 mM to about 0.1 mM of EDTA, from about 0.01% to about 0.05% of polysorbate 20 or polysorbate 80, and has a pH of from about 5.9 to about 6.5.

[0332] In one embodiment, the composition comprises belantamab mafodotin in a formulation that comprises about 20 mg / mL, about 25 mg / mL, about 50 mg / mL or 60 mg / mL belantamab mafodotin, 25 mM citrate buffer, 200 mM trehalose, 0.05 mM disodium EDTA, 0.02% polysorbate 80 or Tween 80, and has a pH of from about 5.9 to about 6.5.

[0333] A “stable” formulation is one in which the protein substantially maintains its physical and / or chemical stability during manufacture, transport, storage, and administration. Stability can be measured at selected temperatures over selected time periods. For example, for a product stored at the recommended temperature of 2°C to 8°C, the formulation is stable at room temperature, about 30°C, or at 40°C for at least 1 month and / or stable at about 2 to 8°C for at least 1 year, preferably at least 2 years. For example, the degree of aggregation during storage can be used as an indicator of protein stability. Thus, a “stable” formulation can be one in which, for example, less than about 10%, preferably less than about 5%, of the protein is present as aggregates in the formulation. A variety of analytical techniques for measuring protein stability are available in the art and are reviewed, for example, in Peptide and Protein Drug Delivery, 247-301, Vincent Lee Ed., Marcel Dekker, Inc., New York, N.Y., Pubs. (1991) and Jones, A. Adv. Drug Delivery Rev. 10: 29-90 (1993).

[0334] In certain aspects of the invention, the formulation allows the composition to remain stable upon freezing, thawing, and / or mixing.

[0335] In yet another aspect, the invention relates to industrial products, such as kits, that comprise a container housing the composition in the formulation described herein. In one aspect, an injection device comprising the formulation is provided. The injection device can comprise a pen injector device or an autoinjector device. In one embodiment, the formulation is contained in a prefilled syringe.

[0336] Therapeutic methods and use of the composition

[0337] The object of the present invention is to provide a treatment method for treating B cell-related disorders or diseases, such as antibody-mediated or plasma cell-mediated diseases, or plasma cell malignancies (e.g., cancers, such as multiple myeloma), or other diseases that can be treated by anti-BCMA antigen-binding proteins. In particular, the object of the present invention is to provide a composition comprising an anti-BCMA antigen-binding protein, such as an anti-BCMA antibody, which specifically binds to BCMA (e.g., human BCMA) and regulates (i.e., inhibits or blocks) the interaction between BCMA and its ligands (such as BAFF and / or APRIL), for treating diseases and disorders responsive to the regulation of this interaction.

[0338] In another aspect of the present invention, there is provided a method for treating a subject (e.g., a human patient) suffering from a B cell-related disorder or disease, such as an antibody-mediated or plasma cell-mediated disease, or a plasma cell malignancy (e.g., cancer, such as multiple myeloma), the method comprising the step of administering to the subject a therapeutically effective amount of the anti-BCMA antigen-binding protein composition described herein.

[0339] In yet another embodiment, the present invention provides a method for treating a cancer patient, the method comprising the step of administering to the patient a therapeutically effective amount of the anti-BCMA antigen-binding protein composition described herein.

[0340] As used herein, the terms "cancer" and "tumor" are used interchangeably and, whether in the singular or plural, refer to cells that have undergone transformation, such as malignant transformation, which renders them pathological to the host organism. Primary cancer cells can be readily distinguished from non-cancerous cells by well-established techniques, particularly histological examination. As used herein, the definition of cancer cells includes not only primary cancer cells but also any cells derived from cancer cell progenitors. This includes metastatic cancer cells, as well as in vitro cultures and cell lines derived from cancer cells. When referring to cancer types that typically present as solid tumors, a "clinically detectable" tumor refers to a tumor detectable based on a tumor mass; e.g., by procedures such as computed tomography (CT) scan, magnetic resonance imaging (MRI), X-ray, ultrasound, or palpation during a physical examination, and / or detectable due to the expression of one or more cancer-specific antigens in a sample obtainable from the patient. A tumor can be a hematopoietic (or blood or hematological or blood-related) cancer, e.g., a cancer derived from blood cells or immune cells, which can be referred to as a "liquid tumor". Specific examples of clinical conditions based on blood tumors include leukemias, such as chronic myeloid leukemia, acute myeloid leukemia, chronic lymphocytic leukemia, and acute lymphocytic leukemia; plasma cell malignancies, such as multiple myeloma, MGUS, and Waldenström macroglobulinemia; lymphomas, such as non-Hodgkin lymphoma, Hodgkin lymphoma; and the like.

[0341] Cancer can be any cancer in which there is an abnormal number of primitive cells or unwanted cell proliferation or is diagnosed as a blood cancer, including lymphatic and bone marrow malignancies. Myeloid malignancies include, but are not limited to, acute myeloid (or myelocytic or myelogenous or myeloblast) leukemia (undifferentiated or differentiated), acute promyelocytic (or promyelocytic or promyelocyte or promyeloblast) leukemia, acute myelomonocytic (or myelomonoblast) leukemia, acute monocytic (or monoblast) leukemia, erythroleukemia, and megakaryocytic (or megakaryoblast) leukemia. These leukemias can be collectively referred to as acute myeloid (or myelocytic or myelogenous) leukemia (AML). Myeloid malignancies also include myeloproliferative disorders (MPD), including, but not limited to, chronic myeloid (or myelogenous) leukemia (CML), chronic myelomonocytic leukemia (CMML), essential thrombocythemia (or thrombocythemia), and polycythemia vera (PCV). Myeloid malignancies also include myelodysplasia (or myelodysplastic syndrome or MDS), which can be referred to as refractory anemia (RA), refractory anemia with excess blasts (RAEB), and refractory anemia with excess blasts in transformation (RAEBT); and myelofibrosis (MFS) with or without myeloid metaplasia of unknown origin, etc.

[0342] Hematopoietic system cancers also include lymphoid malignancies, which may affect lymph nodes, spleen, bone marrow, peripheral blood, and / or extranodal sites. Lymphoid cancers include B-cell malignancies, including but not limited to B-cell non-Hodgkin lymphoma (B-NHL). B-NHL may be indolent (or low-grade), intermediate (or aggressive), or high-grade (very aggressive). Indolent B-cell lymphomas include follicular lymphoma (FL); small lymphocytic lymphoma (SLL); marginal zone lymphoma (MZL) including nodal MZL, extranodal MZL, splenic MZL, and splenic MZL with villous lymphocytes; lymphoplasmacytic lymphoma (LPL); and mucosa-associated lymphoid tissue (MALT or extranodal marginal zone) lymphoma. Intermediate B-NHL includes mantle cell lymphoma (MCL) with or without leukemia, diffuse large cell lymphoma (DLBCL), follicular large cell (or grade 3 or 3B) lymphoma, and primary mediastinal lymphoma (PML). High-grade B-NHL includes Burkitt lymphoma (BL), Burkitt-like lymphoma, small non-cleaved cell lymphoma (SNCCL), and lymphoblastic lymphoma. Other B-NHL includes immunoblastic lymphoma (or immunocytoma), primary effusion lymphoma, HIV-associated (or AIDS-associated) lymphoma, and post-transplant lymphoproliferative disorder (PTLD) or lymphoma. B-cell malignancies also include but are not limited to chronic lymphocytic leukemia (CLL), prolymphocytic leukemia (PLL), Waldenstrom macroglobulinemia (WM), hairy cell leukemia (HCL), large granular lymphocyte (LGL) leukemia, acute lymph(oblastic) leukemia, and Castleman disease. NHL may also include T-cell non-Hodgkin lymphoma (T-NHL), including but not limited to T-cell non-Hodgkin lymphoma, not otherwise specified (NOS), peripheral T-cell lymphoma (PTCL), anaplastic large cell lymphoma (ALCL), angioimmunoblastic lymphoproliferative disorder (AILD), nasal natural killer (NK) / T-cell lymphoma, γ / δ lymphoma, cutaneous T-cell lymphoma, mycosis fungoides, and Sézary syndrome, among others.

[0343] Hematopoietic cancers also include Hodgkin lymphoma (or disease), including classical Hodgkin lymphoma, nodular sclerosing Hodgkin lymphoma, mixed cellularity Hodgkin lymphoma, lymphocyte predominance (LP) Hodgkin lymphoma, nodular LP Hodgkin lymphoma, and lymphocyte depletion Hodgkin lymphoma. Hematopoietic cancers also include plasma cell disorders or cancers, such as multiple myeloma (MM), including smoldering MM, monoclonal gammopathy of undetermined significance (MGUS), plasmacytoma (bone, extramedullary), lymphoplasmacytic lymphoma (LPL), Waldenstroem macroglobulinemia, plasma cell leukemia, and primary amyloidosis (AL). Hematopoietic cancers can also include other cancers of other hematopoietic cells, including polymorphonuclear leukocytes (or neutrophils), basophils, eosinophils, dendritic cells, platelets, red blood cells, and natural killer cells. Tissues of hematopoietic cells, including what is referred to herein as "hematopoietic cell tissue", include bone marrow; peripheral blood; thymus; and peripheral lymphoid tissues, such as the spleen, lymph nodes, mucosa-associated lymphoid tissue (e.g., gut-associated lymphoid tissue), tonsils, Peyer's patches, and appendix, and other mucosa-associated lymphoid tissues, such as the bronchial lining.

[0344] In one embodiment, the cancer is selected from colorectal cancer (CRC), gastric cancer, esophageal cancer, cervical cancer, bladder cancer, breast cancer, head and neck cancer, ovarian cancer, melanoma, renal cell carcinoma (RCC), EC squamous cell carcinoma, non-small cell lung cancer, mesothelioma, pancreatic cancer, and prostate cancer.

[0345] As used herein, the term "treatment" and its derivatives are intended to include therapeutic treatment. With respect to a particular condition, treatment means: (1) improving the condition or one or more biological manifestations of the condition; (2) interfering with (a) one or more points in the biological cascade that leads to or is responsible for the condition, or (b) one or more biological manifestations of the condition; (3) alleviating one or more symptoms, effects, or side effects associated with the condition or one or more symptoms, effects, or side effects associated with the condition or its treatment; (4) slowing the progression of the condition or one or more biological manifestations of the condition and / or (5) curing the condition or one or more biological manifestations of the condition by reducing one or more biological manifestations of the condition to an undetectable level for a period of time, which is considered a state of remission of the manifestation, without additional treatment during the remission period. Those skilled in the art will understand the duration considered to be remission of a particular disease or condition.

[0346] B cell disorders can be classified into B cell development / immunoglobulin production defects (e.g., immunodeficiencies) and excessive / uncontrolled proliferation (e.g., lymphoma, leukemia). As used herein, B cell disorders refer to two types of diseases, and methods for treating B cell disorders with the compositions described herein are provided.

[0347] In certain aspects, the disease or disorder is multiple myeloma (MM), chronic lymphocytic leukemia (CLL), solitary plasmacytoma (bone, extramedullary), amyloidosis (AL), smoldering multiple myeloma (SMM), solitary plasmacytoma (bone, extramedullary medulla), or Waldenström's macroglobulinemia.

[0348] Prophylactic therapies are also contemplated. Those skilled in the art will understand that "prevention" is not an absolute term. In medicine, "prevention" is understood to refer to the prophylactic administration of a drug to significantly reduce the likelihood or severity of a condition or its biological manifestations, or to delay the onset of such a condition or its biological manifestations. Prophylactic therapies are appropriate, for example, when a subject is considered to be at high risk of developing cancer, such as when the subject has a strong family history of cancer or when the subject has been exposed to a carcinogen.

[0349] "Subject" or "patient" are used interchangeably herein and are broadly defined to include any human in need of treatment, such as a human in need of cancer treatment. A subject can include a mammal. In one embodiment, the subject is a human patient. Subjects in need of cancer treatment can include patients at different stages, including newly diagnosed, relapsed, refractory, progressive disease, remission, and others. Subjects in need of cancer treatment can also include patients who have undergone stem cell transplantation or are considered ineligible for transplantation.

[0350] Subjects can be pre-screened to select for treatment with the compositions described herein. In one embodiment, BCMA expression in a sample from the subject is tested prior to treatment with the compositions described herein.

[0351] Prior to treatment with the compositions of the invention, a subject may have received at least one prior cancer therapy. In one embodiment, a subject has been treated with at least 1, at least 2, at least 3, at least 4, at least 5, at least 6, or at least 7 prior cancer therapies prior to treatment with the compositions of the invention.

[0352] In another embodiment, the subject has newly diagnosed cancer and has received 0 prior treatments prior to treatment with the compositions of the invention.

[0353] The compositions of the present invention can be administered by any suitable route. For some compositions, suitable routes include oral, rectal, nasal, topical (including buccal and sublingual), vaginal, parenteral (including subcutaneous, intramuscular, intravenous, intradermal, intrathecal and epidural), and intratumoral. It will be understood that the preferred route may vary depending on, for example, the condition of the recipient and the cancer to be treated.

[0354] In certain embodiments, the compositions of the present invention are administered as pharmaceutical compositions.

[0355] As used herein, the term "administer" refers to delivering the compositions described herein to achieve a therapeutic goal. The compositions can be administered at an administration interval sufficient to achieve a clinical benefit. The compositions can be administered to a subject in a manner that targets the treatment to a specific site.

[0356] In some embodiments, the compositions are administered by injection. Accordingly, in one aspect, there is provided an injection device comprising a composition, pharmaceutical composition or formulation of the present invention. The injection device can include a pen injector device or an autoinjector device.

[0357] As used herein, the term "therapeutically effective amount" or "therapeutically effective dose" of a composition refers to an amount effective in preventing or treating or alleviating a B cell-mediated disorder or symptoms of a disorder. The therapeutically effective amount and treatment regimen are typically determined empirically and may depend on factors such as the age, weight and health of the patient and the disease or disorder to be treated. These factors are within the purview of the attending physician.

[0358] One of ordinary skill in the art will readily determine an appropriate therapeutically effective dose of a composition comprising an anti-BCMA antigen-binding protein. Suitable doses of the compositions described herein can be calculated based on the weight of the patient, for example, a suitable dose can be in the range of about 0.1 mg / kg to about 20 mg / kg, such as about 1 mg / kg to about 20 mg / kg, such as about 10 mg / kg to about 20 mg / kg or such as about 1 mg / kg to about 15 mg / kg, such as about 10 mg / kg to about 15 mg / kg.

[0359] In one embodiment, the therapeutically effective dose of the composition comprising an anti-BCMA antigen-binding protein ranges from about 0.03 mg / kg to about 4.6 mg / kg. In yet another embodiment, the therapeutically effective dose of the composition comprising an anti-BCMA antigen-binding protein is 0.03 mg / kg, 0.06 mg / kg, 0.12 mg / kg, 0.24 mg / kg, 0.48 mg / kg, 0.96 mg / kg, 1.92 mg / kg, 3.4 mg / kg, or 4.6 mg / kg. In yet another embodiment, the therapeutically effective dose of the composition comprising an anti-BCMA antigen-binding protein is 1.9 mg / kg, 2.5 mg / kg, or 3.4 mg / kg.

[0360] In certain embodiments, the composition can be co-administered to a subject with one or more additional therapeutic agents. In another embodiment, the composition can be co-administered to a subject with one or more additional cancer therapeutic agents. The additional cancer therapeutic agents can include, but are not limited to, other immunomodulatory drugs, therapeutic antibodies (e.g., anti-CD38 antibodies, such as daratumumab), CAR-T therapeutic agents, BiTE, HDAC inhibitors, proteasome inhibitors (e.g., bortezomib), anti-inflammatory compounds, and immunomodulatory imide drugs (IMiD) (e.g., thalidomide and its analogs).

[0361] "Co-administer" means the administration of two or more different pharmaceutical compositions or treatments (e.g., radiotherapy) that are administered to a subject by combination in the same pharmaceutical composition or in separate pharmaceutical compositions. Thus, co-administration involves the simultaneous administration of a single pharmaceutical composition containing two or more agents or the administration of two or more different compositions to the same subject at the same or different times.

[0362] In one aspect of the invention, the invention provides a method of treating a B cell disease or disorder in a subject in need thereof by administering a therapeutically effective dose of any of the compositions comprising an anti-BCMA antigen-binding protein described herein.

[0363] In one embodiment, the invention provides a method of treating cancer in a subject in need thereof, which comprises administering a therapeutically effective dose of a composition comprising an anti-BCMA ADC, wherein the average DAR is from about 3.4 to about 4.6.

[0364] In another embodiment, the present invention provides a method of treating cancer in a subject in need thereof, comprising administering a therapeutically effective dose of a composition comprising an anti-BCMA ADC, wherein the antibody comprises a CDRH1 having the amino acid sequence set forth in SEQ ID NO: 1, a CDRH2 having the amino acid sequence set forth in SEQ ID NO: 2, a CDRH3 having the amino acid sequence set forth in SEQ ID NO: 3, a CDRL1 having the amino acid sequence set forth in SEQ ID NO: 4, a CDRL2 having the amino acid sequence set forth in SEQ ID NO: 5, and a CDRL3 having the amino acid sequence set forth in SEQ ID NO: 6; wherein the cytotoxic agent is MMAE or MMAF; wherein the average DAR is from about 3.4 to about 4.6.

[0365] In another embodiment, the present invention provides a method of treating cancer in a subject in need thereof, comprising administering a therapeutically effective dose of a composition comprising an anti-BCMA ADC, wherein the antibody comprises a V having the amino acid sequence set forth in SEQ ID NO: 7 H and a V having the amino acid sequence set forth in SEQ ID NO: 8 L ; wherein the cytotoxic agent is MMAE or MMAF; wherein the average DAR is from about 3.4 to about 4.6.

[0366] In yet another embodiment, the present invention provides a method of treating cancer in a subject in need thereof, comprising administering a therapeutically effective dose of a composition comprising belantamab mafodotin, wherein the average DAR is from 3.4 to about 4.6.

[0367] In yet another embodiment, the present invention provides a method of treating multiple myeloma in a subject in need thereof, comprising administering a therapeutically effective dose of a composition comprising belantamab mafodotin, wherein the average DAR is from 3.4 to about 4.6.

[0368] In one embodiment, the present invention provides a method of treating cancer in a subject in need thereof, comprising administering a therapeutically effective dose of a composition comprising an anti-BCMA ADC; wherein the DL0 percentage is less than or equal to about 10% or about 5%, the DL2 percentage is from about 15% to about 27% or from about 15% to about 32%, the DL4a percentage is from about 35% to about 38% or from about 30% to about 40%, the DL4b percentage is from about 7% to about 9% or from about 5% to about 10%, the DL6 percentage is from about 14% to about 20% or from about 10% to about 20%, and / or the DL8 is from about 6.0% to about 12.0% or from about 4% to about 15%.

[0369] In another embodiment, the present invention provides a method of treating cancer in a subject in need thereof, comprising administering a therapeutically effective dose of a composition comprising an anti-BCMA ADC, wherein the antibody comprises a CDRH1 having the amino acid sequence set forth in SEQ ID NO: 1, a CDRH2 having the amino acid sequence set forth in SEQ ID NO: 2, a CDRH3 having the amino acid sequence set forth in SEQ ID NO: 3, a CDRL1 having the amino acid sequence set forth in SEQ ID NO: 4, a CDRL2 having the amino acid sequence set forth in SEQ ID NO: 5, and a CDRL3 having the amino acid sequence set forth in SEQ ID NO: 6; wherein the cytotoxic agent is MMAE or MMAF; and wherein the DL0 percentage is less than or equal to about 10% or about 5%, the DL2 percentage is from about 15% to about 27% or from about 15% to about 32%, the DL4a percentage is from about 35% to about 38% or from about 30% to about 40%, the DL4b percentage is from about 7% to about 9% or from about 5% to about 10%, the DL6 percentage is from about 14% to about 20% or from about 10% to about 20%, and / or the DL8 is from about 6.0% to about 12.0% or from about 4% to about 15%.

[0370] In another embodiment, the present invention provides a method of treating cancer in a subject in need thereof, comprising administering a therapeutically effective dose of a composition comprising an anti-BCMA ADC, wherein the antibody comprises a V having the amino acid sequence set forth in SEQ ID NO: 7 H and a V having the amino acid sequence set forth in SEQ ID NO: 8 L ; wherein the cytotoxic agent is MMAE or MMAF; and wherein the DL0 percentage is less than or equal to about 10% or about 5%, the DL2 percentage is from about 15% to about 27% or from about 15% to about 32%, the DL4a percentage is from about 35% to about 38% or from about 30% to about 40%, the DL4b percentage is from about 7% to about 9% or from about 5% to about 10%, the DL6 percentage is from about 14% to about 20% or from about 10% to about 20%, and / or the DL8 is from about 6.0% to about 12.0% or from about 4% to about 15%.

[0371] In yet another embodiment, the present invention provides a method of treating cancer in a subject in need thereof, comprising administering a therapeutically effective dose of a composition comprising belantamab mafodotin, wherein the DL0 percentage is less than or equal to about 10% or about 5%, the DL2 percentage is from about 15% to about 27% or from about 15% to about 32%, the DL4a percentage is from about 35% to about 38% or from about 30% to about 40%, the DL4b percentage is from about 7% to about 9% or from about 5% to about 10%, the DL6 percentage is from about 14% to about 20% or from about 10% to about 20%, and / or the DL8 is from about 6.0% to about 12.0% or from about 4% to about 15%.

[0372] In yet another embodiment, the present invention provides a method of treating multiple myeloma in a subject in need thereof, comprising administering a therapeutically effective dose of a composition comprising belantamab mafodotin, wherein the DL0 percentage is less than or equal to about 10% or about 5%, the DL2 percentage is from about 15% to about 27% or from about 15% to about 32%, the DL4a percentage is from about 35% to about 38% or from about 30% to about 40%, the DL4b percentage is from about 7% to about 9% or from about 5% to about 10%, the DL6 percentage is from about 14% to about 20% or from about 10% to about 20%, and / or the DL8 is from about 6.0% to about 12.0% or from about 4% to about 15%.

[0373] In one aspect, the present invention provides a method of treating cancer in a subject in need thereof, comprising administering a therapeutically effective dose of a composition comprising an antibody, which comprises a CDRH1 having the amino acid sequence set forth in SEQ ID NO: 1, a CDRH2 having the amino acid sequence set forth in SEQ ID NO: 2, a CDRH3 having the amino acid sequence set forth in SEQ ID NO: 3, a CDRL1 having the amino acid sequence set forth in SEQ ID NO: 4, a CDRL2 having the amino acid sequence set forth in SEQ ID NO: 5, and a CDRL3 having the amino acid sequence set forth in SEQ ID NO: 6; wherein the composition comprises ≤25% isomerization at heavy chain D103 of CDRH3.

[0374] In one embodiment, the present invention provides a method for treating multiple myeloma in a subject in need thereof, comprising administering a therapeutically effective dose of a composition comprising an antibody, which comprises a CDRH1 having the amino acid sequence set forth in SEQ ID NO: 1, a CDRH2 having the amino acid sequence set forth in SEQ ID NO: 2, a CDRH3 having the amino acid sequence set forth in SEQ ID NO: 3, a CDRL1 having the amino acid sequence set forth in SEQ ID NO: 4, a CDRL2 having the amino acid sequence set forth in SEQ ID NO: 5, and a CDRL3 having the amino acid sequence set forth in SEQ ID NO: 6; wherein the composition comprises ≤25% isomerization at heavy chain D103 of CDRH3.

[0375] In one embodiment, the present invention provides a method for treating multiple myeloma in a subject in need thereof, comprising administering a therapeutically effective dose of a composition comprising belantamab; wherein the composition comprises ≤25% isomerization at heavy chain D103 of CDRH3.

[0376] In one aspect, the present invention provides a method for treating cancer in a subject in need thereof, comprising administering a therapeutically effective dose of a composition comprising an antibody, which comprises a CDRH1 having the amino acid sequence set forth in SEQ ID NO: 1, a CDRH2 having the amino acid sequence set forth in SEQ ID NO: 2, a CDRH3 having the amino acid sequence set forth in SEQ ID NO: 3, a CDRL1 having the amino acid sequence set forth in SEQ ID NO: 4, a CDRL2 having the amino acid sequence set forth in SEQ ID NO: 5, and a CDRL3 having the amino acid sequence set forth in SEQ ID NO: 6; wherein the composition comprises ≤40% oxidation at heavy chain M34 (CDRH1).

[0377] In an embodiment, the present invention provides a method for treating multiple myeloma in a subject in need thereof, comprising administering a therapeutically effective dose of a composition comprising an antibody, which comprises a CDRH1 having the amino acid sequence set forth in SEQ ID NO: 1, a CDRH2 having the amino acid sequence set forth in SEQ ID NO: 2, a CDRH3 having the amino acid sequence set forth in SEQ ID NO: 3, a CDRL1 having the amino acid sequence set forth in SEQ ID NO: 4, a CDRL2 having the amino acid sequence set forth in SEQ ID NO: 5, and a CDRL3 having the amino acid sequence set forth in SEQ ID NO: 6; wherein the composition comprises ≤40% oxidation at heavy chain M34 (CDRH1).

[0378] In an embodiment, the present invention provides a method for treating multiple myeloma in a subject in need thereof, comprising administering a therapeutically effective dose of a composition comprising belantamab; wherein the composition comprises ≤40% oxidation at heavy chain M34 (CDRH1).

[0379] In one aspect of the present invention, the present invention provides a composition comprising an anti-BCMA antigen-binding protein as described herein for treating B cell diseases or disorders.

[0380] In one embodiment, the present invention provides a composition comprising an anti-BCMA ADC as described herein for treating cancer, wherein the average DAR is from about 3.4 to about 4.6.

[0381] In another embodiment, the present invention provides a composition comprising an anti-BCMA ADC as described herein for treating cancer, wherein the antibody comprises a CDRH1 having the amino acid sequence set forth in SEQ ID NO: 1, a CDRH2 having the amino acid sequence set forth in SEQ ID NO: 2, a CDRH3 having the amino acid sequence set forth in SEQ ID NO: 3, a CDRL1 having the amino acid sequence set forth in SEQ ID NO: 4, a CDRL2 having the amino acid sequence set forth in SEQ ID NO: 5, and a CDRL3 having the amino acid sequence set forth in SEQ ID NO: 6; wherein the cytotoxic agent is MMAE or MMAF; wherein the average DAR is from about 3.4 to about 4.6.

[0382] In another embodiment, the present invention provides a composition comprising an anti-BCMA ADC as described herein for treating cancer, wherein the antibody comprises a V H having the amino acid sequence set forth in SEQ ID NO: 7 L and a V having the amino acid sequence set forth in SEQ ID NO: 8; wherein the cytotoxic agent is MMAE or MMAF; wherein the average DAR is from about 3.4 to about 4.6.

[0383] In another embodiment, the present invention provides a composition comprising belantamab mafodotin for treating cancer, wherein the average DAR is from 3.4 to about 4.6.

[0384] In yet another embodiment, the present invention provides a composition comprising belantamab mafodotin for treating multiple myeloma, wherein the average DAR is from 3.4 to about 4.6.

[0385] In one embodiment, the present invention provides a composition for treating cancer comprising an anti-BCMA ADC as described herein; wherein the percentage of DL0 is less than or equal to about 10% or about 5%, the percentage of DL2 is about 15% to about 27% or about 15% to about 32%, the percentage of DL4a is about 35% to about 38% or about 30% to about 40%, the percentage of DL4b is about 7% to about 9% or about 5% to about 10%, the percentage of DL6 is about 14% to about 20% or about 10% to about 20%, and / or DL8 is about 6.0% to about 12.0% or about 4% to about 15%.

[0386] In another embodiment, the present invention provides a composition for treating cancer comprising an anti-BCMA ADC as described herein, wherein the antibody comprises a CDRH1 having the amino acid sequence set forth in SEQ ID NO: 1, a CDRH2 having the amino acid sequence set forth in SEQ ID NO: 2, a CDRH3 having the amino acid sequence set forth in SEQ ID NO: 3, a CDRL1 having the amino acid sequence set forth in SEQ ID NO: 4, a CDRL2 having the amino acid sequence set forth in SEQ ID NO: 5, and a CDRL3 having the amino acid sequence set forth in SEQ ID NO: 6; wherein the cytotoxic agent is MMAE or MMAF; and wherein the percentage of DL0 is less than or equal to about 10% or about 5%, the percentage of DL2 is about 15% to about 27% or about 15% to about 32%, the percentage of DL4a is about 35% to about 38% or about 30% to about 40%, the percentage of DL4b is about 7% to about 9% or about 5% to about 10%, the percentage of DL6 is about 14% to about 20% or about 10% to about 20%, and / or DL8 is about 6.0% to about 12.0% or about 4% to about 15%.

[0387] In another embodiment, the present invention provides a composition for treating cancer comprising an anti-BCMA ADC as described herein, wherein the antibody comprises a V having the amino acid sequence set forth in SEQ ID NO: 7 H and a V having the amino acid sequence set forth in SEQ ID NO: 8 L ; wherein the cytotoxic agent is MMAE or MMAF; and wherein the percentage of DL0 is less than or equal to about 10% or about 5%, the percentage of DL2 is about 15% to about 27% or about 15% to about 32%, the percentage of DL4a is about 35% to about 38% or about 30% to about 40%, the percentage of DL4b is about 7% to about 9% or about 5% to about 10%, the percentage of DL6 is about 14% to about 20% or about 10% to about 20%, and / or DL8 is about 6.0% to about 12.0% or about 4% to about 15%.

[0388] In another embodiment, the present invention provides a composition for treating cancer comprising belantamab mafodotin, wherein the DL0 percentage is less than or equal to about 10% or about 5%, the DL2 percentage is from about 15% to about 26%, the DL4a percentage is from about 35% to about 38%, the DL4b is from about 7% to about 10%, the DL6 percentage is from about 14% to about 20%, and / or the DL8 percentage is from about 6% to about 12%.

[0389] In yet another embodiment, the present invention provides a composition for treating multiple myeloma comprising belantamab mafodotin, wherein the DL0 percentage is less than or equal to about 10% or about 5%, the DL2 percentage is from about 15% to about 27% or from about 15% to about 32%, the DL4a percentage is from about 35% to about 38% or from about 30% to about 40%, the DL4b percentage is from about 7% to about 9% or from about 5% to about 10%, the DL6 percentage is from about 14% to about 20% or from about 10% to about 20%, and / or the DL8 is from about 6.0% to about 12.0% or from about 4% to about 15%.

[0390] In one aspect, the present invention provides a composition for treating cancer comprising an antibody, wherein the antibody comprises a CDRH1 having the amino acid sequence set forth in SEQ ID NO: 1, a CDRH2 having the amino acid sequence set forth in SEQ ID NO: 2, a CDRH3 having the amino acid sequence set forth in SEQ ID NO: 3, a CDRL1 having the amino acid sequence set forth in SEQ ID NO: 4, a CDRL2 having the amino acid sequence set forth in SEQ ID NO: 5, and a CDRL3 having the amino acid sequence set forth in SEQ ID NO: 6; and wherein the composition comprises ≤25% isomerization at the heavy chain D103 of CDRH3.

[0391] In one aspect, the present invention provides a composition for treating multiple myeloma comprising an antibody, wherein the antibody comprises a CDRH1 having the amino acid sequence set forth in SEQ ID NO: 1, a CDRH2 having the amino acid sequence set forth in SEQ ID NO: 2, a CDRH3 having the amino acid sequence set forth in SEQ ID NO: 3, a CDRL1 having the amino acid sequence set forth in SEQ ID NO: 4, a CDRL2 having the amino acid sequence set forth in SEQ ID NO: 5, and a CDRL3 having the amino acid sequence set forth in SEQ ID NO: 6; and wherein the composition comprises ≤25% isomerization at the heavy chain D103 of CDRH3.

[0392] In yet another embodiment, the present invention provides a composition for treating multiple myeloma comprising belantamab mafodotin, wherein the composition comprises ≤25% isomerization at heavy chain D103 of CDRH3.

[0393] In one aspect, the present invention provides a composition for treating cancer comprising an antibody, wherein the antibody comprises CDRH1 having the amino acid sequence set forth in SEQ ID NO: 1, CDRH2 having the amino acid sequence set forth in SEQ ID NO: 2, CDRH3 having the amino acid sequence set forth in SEQ ID NO: 3, CDRL1 having the amino acid sequence set forth in SEQ ID NO: 4, CDRL2 having the amino acid sequence set forth in SEQ ID NO: 5, and CDRL3 having the amino acid sequence set forth in SEQ ID NO: 6; and wherein the composition comprises ≤40% oxidation at heavy chain M34 (CDRH1).

[0394] In one aspect, the present invention provides a composition for treating multiple myeloma comprising an antibody, wherein the antibody comprises CDRH1 having the amino acid sequence set forth in SEQ ID NO: 1, CDRH2 having the amino acid sequence set forth in SEQ ID NO: 2, CDRH3 having the amino acid sequence set forth in SEQ ID NO: 3, CDRL1 having the amino acid sequence set forth in SEQ ID NO: 4, CDRL2 having the amino acid sequence set forth in SEQ ID NO: 5, and CDRL3 having the amino acid sequence set forth in SEQ ID NO: 6; and wherein the composition comprises ≤40% oxidation at heavy chain M34 (CDRH1).

[0395] In yet another embodiment, the present invention provides a composition for treating multiple myeloma comprising belantamab mafodotin, wherein the composition comprises ≤40% oxidation at heavy chain M34 (CDRH1).

[0396] In one aspect of the present invention, there is provided the use of a composition in the manufacture of a medicament for treating B-cell diseases or disorders.

[0397] In one embodiment, there is provided the use of a composition comprising an anti-BCMA ADC in the manufacture of a medicament for treating cancer, wherein the average DAR is from about 3.4 to about 4.6.

[0398] In another embodiment, provided is the use of a composition comprising an anti-BCMA ADC in the preparation of a medicament for treating cancer, wherein the antibody comprises a CDRH1 having the amino acid sequence set forth in SEQ ID NO: 1, a CDRH2 having the amino acid sequence set forth in SEQ ID NO: 2, a CDRH3 having the amino acid sequence set forth in SEQ ID NO: 3, a CDRL1 having the amino acid sequence set forth in SEQ ID NO: 4, a CDRL2 having the amino acid sequence set forth in SEQ ID NO: 5, and a CDRL3 having the amino acid sequence set forth in SEQ ID NO: 6; wherein the cytotoxic agent is MMAE or MMAF; wherein the average DAR is from about 3.4 to about 4.6.

[0399] In another embodiment, provided is the use of a composition comprising an anti-BCMA ADC in the preparation of a medicament for treating cancer, wherein the antibody comprises a V having the amino acid sequence set forth in SEQ ID NO: 7 H and a V having the amino acid sequence set forth in SEQ ID NO: 8 L ; wherein the cytotoxic agent is MMAE or MMAF; wherein the average DAR is from about 3.4 to about 4.6.

[0400] In another embodiment, provided is the use of a composition comprising belantamab mafodotin in the preparation of a medicament for treating cancer, wherein the average DAR is from 3.4 to about 4.6.

[0401] In another embodiment, provided is the use of a composition comprising belantamab mafodotin in the preparation of a medicament for treating multiple myeloma, wherein the average DAR is from 3.4 to about 4.6.

[0402] In one embodiment, provided is the use of a composition comprising an anti-BCMA ADC in the preparation of a medicament for treating cancer; wherein the DL0 percentage is less than or equal to about 10% or about 5%, the DL2 percentage is from about 15% to about 27% or from about 15% to about 32%, the DL4a percentage is from about 35% to about 38% or from about 30% to about 40%, the DL4b percentage is from about 7% to about 9% or from about 5% to about 10%, the DL6 percentage is from about 14% to about 20% or from about 10% to about 20%, and / or the DL8 is from about 6.0% to about 12.0% or from about 4% to about 15%.

[0403] In another embodiment, provided is the use of a composition comprising an anti-BCMA ADC in the preparation of a medicament for treating cancer, wherein the antibody comprises CDRH1 having the amino acid sequence set forth in SEQ ID NO: 1, CDRH2 having the amino acid sequence set forth in SEQ ID NO: 2, CDRH3 having the amino acid sequence set forth in SEQ ID NO: 3, CDRL1 having the amino acid sequence set forth in SEQ ID NO: 4, CDRL2 having the amino acid sequence set forth in SEQ ID NO: 5, and CDRL3 having the amino acid sequence set forth in SEQ ID NO: 6; wherein the cytotoxic agent is MMAE or MMAF; and wherein the DL0 percentage is less than or equal to about 10% or about 5%, the DL2 percentage is about 15% to about 27% or about 15% to about 32%, the DL4a percentage is about 35% to about 38% or about 30% to about 40%, the DL4b percentage is about 7% to about 9% or about 5% to about 10%, the DL6 percentage is about 14% to about 20% or about 10% to about 20%, and / or the DL8 is about 6.0% to about 12.0% or about 4% to about 15%.

[0404] In another embodiment, provided is the use of a composition comprising an anti-BCMA ADC in the preparation of a medicament for treating cancer, wherein the antibody comprises V having the amino acid sequence set forth in SEQ ID NO: 7 H and V having the amino acid sequence set forth in SEQ ID NO: 8 L ; wherein the cytotoxic agent is MMAE or MMAF; and wherein the DL0 percentage is less than or equal to about 10% or about 5%, the DL2 percentage is about 15% to about 27% or about 15% to about 32%, the DL4a percentage is about 35% to about 38% or about 30% to about 40%, the DL4b percentage is about 7% to about 9% or about 5% to about 10%, the DL6 percentage is about 14% to about 20% or about 10% to about 20%, and / or the DL8 is about 6.0% to about 12.0% or about 4% to about 15%.

[0405] In another embodiment, provided is the use of a composition comprising an anti-BCMA ADC in the preparation of a medicament for treating cancer, wherein the DL0 percentage is less than or equal to about 10% or about 5%, the DL2 percentage is about 15% to about 27% or about 15% to about 32%, the DL4a percentage is about 35% to about 38% or about 30% to about 40%, the DL4b percentage is about 7% to about 9% or about 5% to about 10%, the DL6 percentage is about 14% to about 20% or about 10% to about 20%, and / or the DL8 is about 6.0% to about 12.0% or about 4% to about 15%.

[0406] In another embodiment, provided is the use of a composition comprising belantamab mafodotin in the preparation of a medicament for treating cancer, wherein the percentage of DL0 is less than or equal to about 10% or about 5%, the percentage of DL2 is from about 15% to about 27% or from about 15% to about 32%, the percentage of DL4a is from about 35% to about 38% or from about 30% to about 40%, the percentage of DL4b is from about 7% to about 9% or from about 5% to about 10%, the percentage of DL6 is from about 14% to about 20% or from about 10% to about 20%, and / or DL8 is from about 6.0% to about 12.0% or from about 4% to about 15%.

[0407] In yet another embodiment, provided is the use of a composition comprising belantamab mafodotin in the preparation of a medicament for treating multiple myeloma, wherein the percentage of DL0 is less than or equal to about 10% or about 5%, the percentage of DL2 is from about 15% to about 27% or from about 15% to about 32%, the percentage of DL4a is from about 35% to about 38% or from about 30% to about 40%, the percentage of DL4b is from about 7% to about 9% or from about 5% to about 10%, the percentage of DL6 is from about 14% to about 20% or from about 10% to about 20%, and / or DL8 is from about 6.0% to about 12.0% or from about 4% to about 15%.

[0408] In one aspect, provided is the use of a composition comprising an anti-BCMA antigen-binding protein in the preparation of a medicament for treating cancer, wherein the composition comprises an antibody that comprises a CDRH1 having the amino acid sequence set forth in SEQ ID NO: 1, a CDRH2 having the amino acid sequence set forth in SEQ ID NO: 2, a CDRH3 having the amino acid sequence set forth in SEQ ID NO: 3, a CDRL1 having the amino acid sequence set forth in SEQ ID NO: 4, a CDRL2 having the amino acid sequence set forth in SEQ ID NO: 5, and a CDRL3 having the amino acid sequence set forth in SEQ ID NO: 6; and wherein the composition comprises ≤25% isomerization at heavy chain D103 of CDRH3.

[0409] In one embodiment, provided is the use of a composition comprising an anti-BCMA antigen-binding protein in the preparation of a medicament for multiple myeloma, wherein the composition comprises an antibody comprising a CDRH1 having the amino acid sequence set forth in SEQ ID NO: 1, a CDRH2 having the amino acid sequence set forth in SEQ ID NO: 2, a CDRH3 having the amino acid sequence set forth in SEQ ID NO: 3, a CDRL1 having the amino acid sequence set forth in SEQ ID NO: 4, a CDRL2 having the amino acid sequence set forth in SEQ ID NO: 5, and a CDRL3 having the amino acid sequence set forth in SEQ ID NO: 6; and wherein the composition comprises ≤25% isomerization at heavy chain D103 of CDRH3.

[0410] In yet another embodiment, provided is the use of a composition comprising belantamab in the preparation of a medicament for treating multiple myeloma, wherein the composition comprises ≤25% isomerization at heavy chain D103 of CDRH3.

[0411] In one aspect, provided is the use of a composition comprising an anti-BCMA antigen-binding protein in the preparation of a medicament for treating cancer, wherein the composition comprises an antibody comprising a CDRH1 having the amino acid sequence set forth in SEQ ID NO: 1, a CDRH2 having the amino acid sequence set forth in SEQ ID NO: 2, a CDRH3 having the amino acid sequence set forth in SEQ ID NO: 3, a CDRL1 having the amino acid sequence set forth in SEQ ID NO: 4, a CDRL2 having the amino acid sequence set forth in SEQ ID NO: 5, and a CDRL3 having the amino acid sequence set forth in SEQ ID NO: 6; and wherein the composition comprises ≤40% oxidation at heavy chain M34 (CDRH1).

[0412] In one embodiment, provided is the use of a composition comprising an anti-BCMA antigen-binding protein in the preparation of a medicament for multiple myeloma, wherein the composition comprises an antibody comprising a CDRH1 having the amino acid sequence set forth in SEQ ID NO: 1, a CDRH2 having the amino acid sequence set forth in SEQ ID NO: 2, a CDRH3 having the amino acid sequence set forth in SEQ ID NO: 3, a CDRL1 having the amino acid sequence set forth in SEQ ID NO: 4, a CDRL2 having the amino acid sequence set forth in SEQ ID NO: 5, and a CDRL3 having the amino acid sequence set forth in SEQ ID NO: 6; and wherein the composition comprises ≤40% oxidation at heavy chain M34 (CDRH1).

[0413] In yet another embodiment, there is provided the use of a composition comprising belantamab mafodotin in the manufacture of a medicament for the treatment of multiple myeloma, wherein the composition comprises ≤40% oxidation at heavy chain M34 (CDRH1).

[0414] All patents and references disclosed herein are hereby incorporated by reference in their entirety and completely.

[0415] The invention described herein includes the following embodiments:

[0416] 1. A composition comprising an isomerization variant of an anti-BCMA antibody, wherein the isomerization variant comprises a heavy chain amino acid sequence comprising CDRH1 of SEQ ID NO: 1, CDRH2 of SEQ ID NO: 2, and CDRH3 of SEQ ID NO: 3, and a light chain amino acid sequence comprising CDRL1 of SEQ ID NO: 4, CDRL2 of SEQ ID NO: 5, and CDRL3 of SEQ ID NO: 6; wherein the composition comprises ≤25% of the isomerization variant.

[0417] 2. A composition comprising an oxidation variant of an anti-BCMA antibody, wherein the oxidation variant comprises a heavy chain amino acid sequence comprising CDRH1 of SEQ ID NO: 1, CDRH2 of SEQ ID NO: 2, and CDRH3 of SEQ ID NO: 3, and a light chain amino acid sequence comprising CDRL1 of SEQ ID NO: 4, CDRL2 of SEQ ID NO: 5, and CDRL3 of SEQ ID NO: 6; wherein the composition comprises ≤40% of the oxidation variant.

[0418] 3. A composition comprising an anti-BCMA antibody, the anti-BCMA antibody comprising CDRH1 having the amino acid sequence set forth in SEQ ID NO: 1, CDRH2 having the amino acid sequence set forth in SEQ ID NO: 2, CDRH3 having the amino acid sequence set forth in SEQ ID NO: 3, CDRL1 having the amino acid sequence set forth in SEQ ID NO: 4, CDRL2 having the amino acid sequence set forth in SEQ ID NO: 5, and CDRL3 having the amino acid sequence set forth in SEQ ID NO: 6; wherein the composition comprises 0.1-25% isomerization at D103 of CDRH3.

[0419] 4. A composition comprising an anti-BCMA antibody, wherein the anti-BCMA antibody comprises a CDRH1 having the amino acid sequence set forth in SEQ ID NO: 1, a CDRH2 having the amino acid sequence set forth in SEQ ID NO: 2, a CDRH3 having the amino acid sequence set forth in SEQ ID NO: 3, a CDRL1 having the amino acid sequence set forth in SEQ ID NO: 4, a CDRL2 having the amino acid sequence set forth in SEQ ID NO: 5, and a CDRL3 having the amino acid sequence set forth in SEQ ID NO: 6; wherein the composition comprises 0.1-40% oxidation at M34 of CDRH1.

[0420] 5. A composition comprising an anti-BCMA antibody that is at least about 90% identical to the heavy chain amino acid sequence of SEQ ID NO: 9 and the light chain amino acid sequence of SEQ ID NO: 10, wherein the composition comprises 0.1-25% isomerization at D103 of CDRH3.

[0421] 6. A composition comprising an anti-BCMA antibody that is at least about 90% identical to the heavy chain amino acid sequence of SEQ ID NO: 9 and the light chain amino acid sequence of SEQ ID NO: 10, wherein the composition comprises 0.1-40% oxidation at M34 of CDRH1.

[0422] 7. The composition according to any one of the preceding embodiments, wherein the composition comprises ≤65% oxidation at M256 of the heavy chain and / or ≤60% oxidation at M432 of the heavy chain.

[0423] 8. The composition according to any one of the preceding embodiments, wherein the composition comprises an antibody variant comprising at least one selected from the group consisting of: deamidation of the heavy chain at N388 and / or N393, D103 to N103 in CDRH3, C-terminal lysine cleavage, and conversion of N-terminal glutamine to pyroglutamic acid.

[0424] 9. The composition according to any one of the preceding embodiments, wherein the composition comprises at least one selected from the group consisting of: up to 100% deamidation at N388 and / or N393, up to 100% N103 in CDRH3, up to 100% C-terminal lysine cleavage, and up to 100% conversion of N-terminal glutamine to pyroglutamic acid.

[0425] 10. The composition according to any one of the preceding embodiments, wherein the composition comprises any percentage of glycoform G0, G1, G2, G0-GlcNac, or G0-2GlcNac.

[0426] 11. The composition according to any one of the foregoing embodiments, wherein the anti-BCMA antibody is belantamab.

[0427] 12. The composition according to any one of the foregoing embodiments, wherein the anti-BCMA antibody is conjugated to a cytotoxic agent to form an antibody-drug conjugate.

[0428] 13. The composition according to any one of the foregoing embodiments, wherein the anti-BCMA antibody is belantamab mafodotin.

[0429] 14. The composition according to embodiments 12-13, wherein the DL2 percentage is at least about 30%, about 15% to about 27% or about 15% to about 32%; the DL4a percentage is at least about 30%, about 35% to about 38% or about 30% to about 40%; the DL4b percentage is at least about 5%, about 7% to about 9% or about 5% to about 10%; the DL6 percentage is at least about 10%, about 14% to about 20% or about 10% to about 20%; and / or the DL8 is at least about 1%, about 6.0% to about 12.0% or about 4% to about 15%.

[0430] 15. The composition according to embodiments 12-14, wherein the average DAR is about 3.4 to about 4.6.

[0431] 16. The composition according to embodiments 12-15, wherein the DL0 percentage is less than or equal to about 10% or about 5%.

[0432] 17. A pharmaceutical composition comprising the composition according to any one of the foregoing embodiments and at least one pharmaceutically acceptable excipient.

[0433] 18. A formulation comprising the pharmaceutical composition according to embodiment 17, which comprises from about 20 mg / mL to about 60 mg / mL of an anti-BCMA antigen-binding protein, from about 10 mM to about 30 mM of a citrate buffer, from about 120 mM to about 240 mM of trehalose, from about 0.01 mM to about 0.1 mM of EDTA, from about 0.01% to about 0.05% of polysorbate 20 or polysorbate 80, and has a pH of about 5.9 to about 6.5.

[0434] 19. The formulation according to embodiment 18, which comprises about 20 mg / mL, about 25 mg / mL, about 50 mg / mL or about 60 mg / mL of belantamab mafodotin, 25 mM citrate buffer, 200 mM trehalose, 0.05 mM disodium EDTA, 0.02% polysorbate 20 or polysorbate 80, and has a pH of about 5.9 to about 6.5.

[0435] 20. A method of treating cancer, comprising administering to a subject in need thereof a therapeutically effective amount of the composition of embodiments 1-16.

[0436] 21. The composition of embodiments 1-14 for treating cancer.

[0437] 22. Use of the composition of embodiments 1-16 in the preparation of a medicament for treating cancer.

[0438] 23. A composition comprising an anti-BCMA antibody-drug-conjugate (ADC), wherein the percentage of DL2 is at least about 30%, about 15% to about 27% or about 15% to about 32%; the percentage of DL4a is at least about 30%, about 35% to about 38% or about 30% to about 40%; the percentage of DL4b is at least about 5%, about 7% to about 9% or about 5% to about 10%; the percentage of DL6 is at least about 10%, about 14% to about 20% or about 10% to about 20%; and / or DL8 is at least about 1%, about 6.0% to about 12.0% or about 4% to about 15%.

[0439] 24. The composition according to embodiment 23, wherein the antibody comprises CDRH1 having the amino acid sequence set forth in SEQ ID NO: 1, CDRH2 having the amino acid sequence set forth in SEQ ID NO: 2, CDRH3 having the amino acid sequence set forth in SEQ ID NO: 3, CDRL1 having the amino acid sequence set forth in SEQ ID NO: 4, CDRL2 having the amino acid sequence set forth in SEQ ID NO: 5, and CDRL3 having the amino acid sequence set forth in SEQ ID NO: 6.

[0440] 25. The composition according to embodiment 23 or 24, wherein the percentage of DL2 is about 15% to about 32%, the percentage of DL4a is about 30% to about 40%, the percentage of DL4b is about 5% to about 10%, the percentage of DL6 is about 10% to about 20%, and DL8 is about 4% to about 15%.

[0441] 26. The composition according to embodiment 23 or 24, wherein the percentage of DL2 is about 15% to about 27%, the percentage of DL4a is about 35% to about 38%, the percentage of DL4b is about 7% to about 9%, the percentage of DL6 is about 14% to about 20%, and DL8 is about 6.0% to about 12.0%.

[0442] 27. The composition according to embodiments 23-26, wherein the average drug-antibody ratio (DAR) is about 2.1 to about 5.7.

[0443] 28. The composition according to any one of embodiments 23-26, wherein the average DAR is from about 3.4 to about 4.6.

[0444] 29. The composition according to any one of embodiments 23-26, wherein the average DAR is from about 3.8 to about 4.5.

[0445] 30. A composition comprising an anti-BCMA antibody-drug conjugate (ADC), wherein the percentage of DL0 is less than or equal to about 10% or about 5%.

[0446] 31. The composition according to embodiment 30, wherein the antibody comprises CDRH1 having the amino acid sequence set forth in SEQ ID NO: 1, CDRH2 having the amino acid sequence set forth in SEQ ID NO: 2, CDRH3 having the amino acid sequence set forth in SEQ ID NO: 3, CDRL1 having the amino acid sequence set forth in SEQ ID NO: 4, CDRL2 having the amino acid sequence set forth in SEQ ID NO: 5, and CDRL3 having the amino acid sequence set forth in SEQ ID NO: 6.

[0447] 32. The composition according to embodiment 30 or 31, wherein the percentage of DL0 is less than or equal to about 5%.

[0448] 33. The composition according to any one of embodiments 30-32, wherein the percentage of DL2 is from about 15% to about 32%, the percentage of DL4a is from about 30% to about 40%, the percentage of DL4b is from about 5% to about 10%, the percentage of DL6 is from about 10% to about 20%, and DL8 is from about 4% to about 15%.

[0449] 34. The composition according to any one of embodiments 30-32, wherein the percentage of DL2 is from about 15% to about 27%, the percentage of DL4a is from about 35% to about 38%, the percentage of DL4b is from about 7% to about 9%, the percentage of DL6 is from about 14% to about 20%, and DL8 is from about 6.0% to about 12.0%.

[0450] 35. The composition according to any one of embodiments 30-34, wherein the average drug-antibody ratio (DAR) is from about 2.1 to about 5.7.

[0451] 36. The composition according to any one of embodiments 30-34, wherein the average DAR is from about 3.4 to about 4.6.

[0452] 37. The composition according to any one of embodiments 30-34, wherein the average DAR is from about 3.8 to about 4.5.

[0453] 38. The composition according to any one of embodiments 23 - 37, wherein the antibody comprises a V having the amino acid sequence set forth in SEQ ID NO: 7 H and a V having the amino acid sequence set forth in SEQ ID NO: 8 L .

[0454] 39. The composition according to any one of embodiments 23 - 38, wherein the antibody is belantamab.

[0455] 40. The composition according to any one of embodiments 23 - 37, wherein the cytotoxic agent is MMAE or MMAF.

[0456] 41. The composition according to any one of embodiments 21 - 40, wherein the anti - BCMA ADC is belantamab mafodotin.

[0457] 42. The composition according to any one of embodiments 23 - 41, wherein the DL percentage is determined by separating individual DL species using hydrophobic interaction chromatography (HIC), calculating the area under the curve for each DL peak, and dividing each DL peak by the total area under the curve of all combined DL species.

[0458] 43. The composition according to embodiment 42, wherein the average DAR is calculated from the area under the curve for each DL species using the following formula:

[0459] .

[0460] 44. A pharmaceutical composition comprising the composition according to any one of embodiments 23 - 43 and at least one pharmaceutically acceptable excipient.

[0461] 45. A formulation comprising the pharmaceutical composition according to embodiment 44, which comprises from about 20 mg / mL to about 60 mg / mL of an anti - BCMA antigen - binding protein, from about 10 mM to about 30 mM of a citrate buffer, from about 120 mM to about 240 mM of trehalose, from about 0.01 mM to about 0.1 mM of EDTA, from about 0.01% to about 0.05% of polysorbate 20 or polysorbate 80, and has a pH of from about 5.9 to about 6.5.

[0462] 46. The formulation according to embodiment 45, which comprises about 20 mg / mL, about 25 mg / mL, about 50 mg / mL or about 60 mg / mL of belantamab mafodotin, 25 mM citrate buffer, 200 mM trehalose, 0.05 mM disodium EDTA, 0.02% polysorbate 20 or polysorbate 80, and has a pH of from about 5.9 to about 6.5.

[0463] 47. A method of treating cancer, comprising administering to a subject in need thereof a therapeutically effective amount of the composition of embodiments 23-43.

[0464] 48. The composition of embodiments 23-43 for treating cancer.

[0465] 49. Use of the composition of embodiments 23-43 in the preparation of a medicament for treating cancer.

[0466] 50. A composition comprising an acidic variant of an antibody, wherein the acidic variant comprises CDRH1 of SEQ ID NO: 1, CDRH2 of SEQ ID NO: 2, and CDRH3 of SEQ ID NO: 3, and a light chain amino acid sequence comprising CDRL1 of SEQ ID NO: 4, CDRL2 of SEQ ID NO: 5, and CDRL3 of SEQ ID NO: 6; wherein the composition comprises 1-70% of the acidic variant.

[0467] 51. A composition comprising an acidic variant of an antibody, wherein the acidic variant comprises CDRH1 of SEQ ID NO: 1, CDRH2 of SEQ ID NO: 2, and CDRH3 of SEQ ID NO: 3, and a light chain amino acid sequence comprising CDRL1 of SEQ ID NO: 4, CDRL2 of SEQ ID NO: 5, and CDRL3 of SEQ ID NO: 6; wherein the composition comprises ≤70% of the acidic variant.

[0468] 52. A composition comprising a basic variant of an antibody, wherein the basic variant comprises a heavy chain amino acid sequence comprising CDRH1 of SEQ ID NO: 1, CDRH2 of SEQ ID NO: 2, and CDRH3 of SEQ ID NO: 3, and a light chain amino acid sequence comprising CDRL1 of SEQ ID NO: 4, CDRL2 of SEQ ID NO: 5, and CDRL3 of SEQ ID NO: 6; wherein the composition comprises 1-30% of the basic variant.

[0469] 53. A composition comprising a basic variant of an antibody, wherein the basic variant comprises a heavy chain amino acid sequence comprising CDRH1 of SEQ ID NO: 1, CDRH2 of SEQ ID NO: 2, and CDRH3 of SEQ ID NO: 3, and a light chain amino acid sequence comprising CDRL1 of SEQ ID NO: 4, CDRL2 of SEQ ID NO: 5, and CDRL3 of SEQ ID NO: 6; wherein the composition comprises ≤30% of the basic variant.

[0470] 54. A composition comprising a major isotype of an antibody, wherein the major isotype comprises a heavy chain amino acid sequence comprising CDRH1 of SEQ ID NO: 1, CDRH2 of SEQ ID NO: 2, and CDRH3 of SEQ ID NO: 3, and a light chain amino acid sequence comprising CDRL1 of SEQ ID NO: 4, CDRL2 of SEQ ID NO: 5, and CDRL3 of SEQ ID NO: 6; wherein the composition comprises 1 - 90% of the major isotype.

[0471] 55. A composition comprising a major isotype of an antibody, wherein the major isotype comprises a heavy chain amino acid sequence comprising CDRH1 of SEQ ID NO: 1, CDRH2 of SEQ ID NO: 2, and CDRH3 of SEQ ID NO: 3, and a light chain amino acid sequence comprising CDRL1 of SEQ ID NO: 4, CDRL2 of SEQ ID NO: 5, and CDRL3 of SEQ ID NO: 6; wherein the composition comprises ≥1% of the major isotype.

[0472] 56. A composition comprising a charged variant of an antibody, the charged variant of the antibody comprising a heavy chain amino acid sequence comprising CDRH1 of SEQ ID NO: 1, CDRH2 of SEQ ID NO: 2, and CDRH3 of SEQ ID NO: 3, and a light chain amino acid sequence comprising CDRL1 of SEQ ID NO: 4, CDRL2 of SEQ ID NO: 5, and CDRL3 of SEQ ID NO: 6; wherein the composition comprises: ≤70% of acidic variants; and / or ≤30% of basic variants; and / or ≥1% of the major isotype. Examples

[0473] Example 1: Determination of the percentage of DL species and the average DAR in an ADC composition.

[0474] For Examples 2 - 9, the DL percentage and the average DAR were calculated as follows:

[0475] The percentage of a specific DL species (e.g., DL0 percentage, DL2 percentage, DL4a percentage, DL4b percentage, DL6 percentage, and DL8 percentage) was determined by separating individual DL species using hydrophobic interaction chromatography (HIC) (as Figure 2 illustrated), calculating the area under the curve of each DL peak, and dividing each DL peak by the total area under the curve of all combined DL species.

[0476] The average DAR of each sample was calculated from the area under the curve of each DL species using the following formula:

[0477] 。

[0478] Reference standards used in Examples 2-8 and can also be used in other experiments, including the samples in Table 1.

[0479] Table 1

[0480]

[0481] Example 2: Effect of average DAR on cell growth inhibition.

[0482] The cell growth inhibitory effect of belantamab mafodotin was determined by measuring the cell viability of human multiple myeloma cell line NCI-H929 cells after incubation with belantamab mafodotin for 48 hours. Cell viability was measured using Promega's CellTiterGlo technology. An increase in the concentration of belantamab mafodotin corresponded proportionally to a decrease in the CellTiter Glo luminescence signal. A dose response (EC50) was generated using a 4-parameter non-linear regression logistic model by SoftMax Pro. The ratio of the reference standard #132371424 EC50 to the sample EC50 was calculated to determine the relative potency. The results are summarized in Table 2.

[0483] Table 2:

[0484]

[0485] Example 3: Effect of average DAR on ADCC activity.

[0486] Incubate belantamab mafodotin, multiple myeloma cells, and NK cells (effector cells). Without being bound by theory, belantamab mafodotin binds to BCMA expressed on the surface of multiple myeloma cells, and the Fc region of the antibody binds to FcγRIIIa on effector cells through its FcγRIIIa receptor. The binding of these receptors to the surface of effector cells leads to the synthesis and secretion of cytokines (IFNg) and the release of granules (perforin and granzyme) that enter the cytoplasm of target cells. Granzyme initiates signaling events within target cells, leading to the death of these cells by apoptosis. The source of NK cells is peripheral blood mononuclear cells (PBMCs) isolated from human whole blood. Then, 10 µL of the fluorescence-enhancing ligand BATDA (bis-(acetoxymethyl) 2,2’:6’,2’’-terpyridine-6,6’’-dicarboxylate) (Perkin-Elmer Cat #C136-100) is added to 1 mL of NCI-H929 cells (human multiple myeloma cell line), which penetrates the cell membrane. The ester bonds in BATDA are hydrolyzed to form a hydrophilic ligand (TDA) that can no longer pass through the cell membrane. The labeled cells are added to different amounts of belantamab mafodotin and effector cells (PBMCs). The cells lyse and release TDA. After cell lysis, the ligand associates with 200 µL of DELFIA europium solution (Perkin-Elmer Cat #C135-100) to form a highly fluorescent and stable chelate (EuTDA). When measured with a fluorescence plate reader, the fluorescence measured is directly related to the amount of lysed cells. The ADCC activity of belantamab mafodotin is reported as the ratio of the EC50 value of the sample to the EC50 value of reference standard #132371424. The results are summarized in Table 3.

[0487] Table 3:

[0488]

[0489] Example 4: Effect of average DAR on BCMA binding and FcγRIIIa binding.

[0490] Binding of belantamab mafodotin to BCMA and FcγRIIIa was measured using surface plasmon resonance (SPR). Belantamab mafodotin was diluted to 10 μg / mL with PBST, injected, and captured by Protein A immobilized on a CM5 sensor chip. Then BCMA was injected and bound to the captured belantamab mafodotin. Next, FcγRIIIa was injected and allowed to bind to the captured belantamab mafodotin. The functional concentrations of belantamab mafodotin binding to BCMA and FcγRIIIa were calculated from a reference standard curve (reference standard #132371424) and reported separately as the BCMA or FcγRIIIa binding concentration. The total belantamab mafodotin concentration of the sample was pre-determined by absorbance at 280 nm. Specific binding activity (%) was calculated by dividing the BCMA or FcγRIIIa binding concentration by the absorbance concentration at 280 nm. The results are summarized in Table 4.

[0491] Table 4:

[0492]

[0493] Example 5: Effect of average DAR on tumor volume.

[0494] Multiple myeloma cell lines were subcutaneously implanted into the flanks of severe combined immunodeficiency (SCID) mice. Starting around day 15, all tumors were measured three times a week using a caliper system, and the length and width of each mouse's tumor were recorded to calculate tumor volume (volume = length x (width2) x 0.5). When the average tumor volume reached approximately 200 mm3, the mice were randomly grouped and dosed twice a week for 2 weeks with one of the belantamab mafodotin with average DAR samples. All tumors were measured in this way, and individual mice were euthanized once their tumors reached an average tumor measurement of 2.0 mm3 or on day 60 (whichever came first). A summary of the study design is summarized in Table 5, and the results are depicted in Figure 3 as follows.

[0495] Table 5:

[0496]

[0497] Example 6: Effect of DL species on BCMA binding, FcγRIIIa binding, and FcRn binding.

[0498] Samples of belantamab mafodotin containing specific DL species were prepared by collecting individual peaks of the HIC chromatogram. Binding of belantamab mafodotin to BCMA and FcγRIIIa was measured using surface plasmon resonance (SPR) as described in Example 4.

[0499] The binding of neonatal Fc receptor (FcRn) to belantamab mafodotin was measured using surface plasmon resonance (SPR). Samples were diluted and belantamab mafodotin was captured by FcRn immobilized on a nitrilotriacetic acid (NTA) sensor chip. The FcRn-binding concentration of the sample was determined by interpolating the binding response on a calibration curve. The specific binding activity (%) was calculated by dividing the FcRn-binding concentration by the total protein concentration. The results are summarized in Table 6.

[0500] Reference standard #162397940 was used.

[0501] Table 6:

[0502]

[0503] Example 7: Effect of DL species on cell growth inhibition.

[0504] Specific DL species samples of belantamab mafodotin were prepared as in Example 7. Cell growth inhibition was determined as in Example 2. Reference standard #162397940 was used. The results are summarized in Table 7.

[0505] Table 7:

[0506]

[0507] Example 8: Effect of DL species on ADCC activity.

[0508] Belantamab mafodotin binds to BCMA expressed on the surface of multiple myeloma cells. The Fc region of belantamab mafodotin binds to FcγRIIIa (CD16a) on Jurkat T effector cells (Promega, Cat #G7102, BioCat #140011), which have been engineered to stably express 1) the high-affinity V158 variant of the human FcγRIIIa receptor and 2) a luciferase reporter gene fused to a promoter downstream of the NFAT activation sequence. When the antibody binds to both H929 and the effector cells simultaneously, activation of the NFAT pathway leads to gene transcription of the luciferase reporter gene and expression of firefly luciferase within the effector cells. After addition of the luminescence substrate (Bio-GloTM Luciferase Assay System, Promega, Cat #G7940) and cell lysis, luciferase produced due to NFAT activation was measured in relative light units (RLU) using a microplate reader. In this assay, belantamab mafodotin was added in a dose-dependent manner; therefore, a dose response (half-maximal effective concentration or EC50) was generated using a non-linear regression logistic model. The ratio of the reference standard EC50 to the sample EC50 was calculated to determine the relative potency. Reference standard #162397940 was used. The results are summarized in Table 8.

[0509] Table 8:

[0510]

[0511] Example 9: Average DAR and percentage of DL species of several batches of belantamab mafodotin

[0512] Several (19) batches of belantamab mafodotin were prepared. The average DAR and percentage of DL species of each batch were calculated as described in Example 1. The results are summarized in Tables 9 - 12.

[0513] Table 9:

[0514]

[0515] Table 10:

[0516]

[0517] Table 11:

[0518]

[0519] Table 12:

[0520]

[0521] Example 10: Design of the forced degradation study of belantamab mafodotin.

[0522] The summary of the study designs of Examples 11 - 18 is depicted in Table 13.

[0523] Table 13: Design of the forced degradation study of belantamab mafodotin

[0524]

[0525] 1. RH: Relative humidity

[0526] Example 11: Oxidation conditions

[0527] To create oxidative stress samples, belantamab mafodotin samples were diluted to 10 mg / mL and hydrogen peroxide was added such that the molar ratio of hydrogen peroxide to belantamab mafodotin was 500:1. The samples were quenched with methionine and buffer exchange was performed using a 3 kDa molecular weight cut-off filter (MWCO).

[0528] Deamidation and oxidation were determined using tryptic peptide mapping tandem mass spectrometry (peptide mapping LC-MS / MS). Samples were denatured in 6 M guanidine hydrochloride to a concentration of 4.2 µg / µL. Disulfide bonds were reduced with 50 mM DTT for 20 minutes at room temperature. 100 mM iodoacetate was added and reacted with free cysteine residues for 30 minutes at room temperature in the dark. Samples were buffer exchanged using a BioRad spin column (part number 7326221) before digestion with 0.5% trypsin from Worthington (part number TRTPCK) at 37 °C for 15 minutes. The resulting peptides were loaded onto a Waters reversed-phase ultra performance liquid chromatography (UPLC) column (part number 186003687) and eluted with a gradient of water and acetonitrile in 0.1% trifluoroacetic acid using a Waters Acquity UPLC. Peptides were detected using an ultraviolet detector and a mass spectrometer such as a Thermo Scientific LTQ Orbitrap XL. Deamidation or oxidation levels were calculated using the extracted ion chromatograms of unmodified and modified peptides by dividing the area under the curve of the modified peptide by the total area under the curves of both modified and unmodified peptides.

[0529] Binding of belantamab mafodotin to BCMA and FcγRIIIa was measured using surface plasmon resonance (SPR) as described in Example 4. Binding of the neonatal Fc receptor (FcRn) to belantamab mafodotin was measured using surface plasmon resonance (SPR) as described in Example 6.

[0530] Reference standard #182407660A was used. Summaries of the results are depicted in Tables 14 and 15.

[0531] Table 14: PTM Abundance in H2O2-Treated Belantamab Mafodotin

[0532]

[0533] Table 15: Binding Activity of H2O2-Treated Belantamab Mafodotin

[0534]

[0535] Extrapolating from the oxidation data of HC Met 34, up to 37% oxidation can result in at least 70% antigen-specific binding activity. This was calculated using the linear slopes of the time 0 (0.2 - 0.4%) and 24-hour (45.1 - 45.5%) M34 oxidized samples, which had 88 - 99% and 62 - 68% antigen-specific binding activity, respectively.

[0536] Extrapolation from the oxidation data of HC Met 256 shows that up to 89% oxidation can result in at least 70% FcγRIIIa - specific binding activity. This was calculated using the linear slopes of the time 0 (2.9 - 3.5%) and 24 - hour (98.3 - 98.7%) M256 oxidized samples, which had 89 - 99% and 66 - 70% FcγRIIIa - specific binding activity, respectively. Extrapolation from the oxidation data of HC Met 256 shows that up to 64% oxidation can result in at least 70% FcRn - specific binding activity. This was calculated using the linear slopes of the time 0 (2.9 - 3.5%) and 24 - hour (98.3 - 98.7%) M256 oxidized samples, which had 93 - 97% and 52 - 57% FcRn - specific binding activity, respectively.

[0537] Extrapolation from the oxidation data of HC Met 432 shows that up to 86% oxidation can result in at least 70% FcγRIIIa - specific binding activity. This was calculated using the linear slopes of the time 0 (0.4 - 0.6%) and 24 - hour (94.8 - 95.2%) M432 oxidized samples, which had 88 - 99% and 66 - 70% FcγRIIIa - specific binding activity, respectively. Extrapolation from the oxidation data of HC Met 432 shows that up to 61% oxidation can result in at least 70% FcRn - specific binding activity. This was calculated using the linear slopes of the time 0 (0.4 - 0.6%) and 24 - hour (94.8 - 95.2%) M432 oxidized samples, which had 93 - 97% and 52 - 57% FcRn - specific binding activity, respectively.

[0538] Example 12: Chemistry: High pH (alkali treatment)

[0539] To create high - pH samples, the belantamab mafodotin sample was diluted with 4 mM Tris buffer to adjust the pH to 9. The sample was further diluted to 10 mg / mL and incubated at 25°C / 50% RH for up to 21 days.

[0540] As described in Example 11, deamidation, isomerization, and oxidation were determined using tryptic peptide mapping by tandem mass spectrometry (peptide mapping LC - MS / MS). The extracted ion chromatograms of the unmodified and modified peptides were used to calculate the levels of deamidation, isomerization, or oxidation by dividing the area under the curve of the modified peptide by the total area under the curves of both the modified and unmodified peptides.

[0541] As described in Example 4, the binding of belantamab mafodotin to BCMA and FcYRIIIa was measured using surface plasmon resonance (SPR). As described in Example 6, the binding of the neonatal Fc receptor (FcRn) to belantamab mafodotin was measured using surface plasmon resonance (SPR).

[0542] Reference standard #182407660A was used. The results are summarized in Tables 16 and 17.

[0543] Table 16: PTM Abundance in Alkaline-Treated Belantamab Mafodotin

[0544]

[0545] Table 17: Binding Activity of Alkaline-Treated Belantamab Mafodotin

[0546]

[0547] Deamidation of HC Asn 388 and HC Asn 393 is expected to be higher than the reported levels of 10.3% and 14.5% respectively, with no effect on antigen-specific binding, FcγRIIIa-specific binding, and FcRn-specific binding.

[0548] Example 13: Chemistry: Low pH (Acid Treatment)

[0549] To create a low pH sample, the belantamab mafodotin sample was diluted with citrate buffer to adjust the pH to 5. The sample was further diluted to 10 mg / mL and incubated at 25°C / 50% RH for up to 21 days.

[0550] As described in Example 11, isomerization, deamidation, and oxidation were determined using tryptic peptide mapping tandem mass spectrometry (peptide mapping LC-MS / MS). The extracted ion chromatograms of unmodified and modified peptides were used to calculate the level of isomerization, deamidation, or oxidation by dividing the area under the curve of the modified peptide by the total area under the curves of both modified and unmodified peptides.

[0551] As described in Example 4, surface plasmon resonance (SPR) was used to measure the binding of belantamab mafodotin to BCMA and FcYRIIIa. As described in Example 6, surface plasmon resonance (SPR) was used to measure the binding of the neonatal Fc receptor (FcRn) to belantamab mafodotin.

[0552] Reference standard #182407660A was used. The results are summarized in Tables 18 and 19.

[0553] Table 18: PTM Abundance in Acid-Treated Belantamab Mafodotin

[0554]

[0555] Table 19: Binding Activity of Acid-Treated Belantamab Mafodotin

[0556]

[0557] Example 14: Heat: Elevate temperature

[0558] To create heat stressed samples, belantamab mafodotin samples were diluted to 10 mg / mL in formulation buffer and incubated at 40°C / 75% RH for up to 28 days.

[0559] As described in Example 11, isomerization, deamidation, and oxidation were determined using tryptic peptide mapping tandem mass spectrometry (peptide mapping LC-MS / MS). Extracted ion chromatograms of unmodified and modified peptides were used to calculate the level of isomerization, deamidation, or oxidation by dividing the area under the curve of the modified peptide by the total area under the curves of both modified and unmodified peptides.

[0560] As described in Example 4, the binding of belantamab mafodotin to BCMA and FcγRIIIa was measured using surface plasmon resonance (SPR). As described in Example 6, the binding of the neonatal Fc receptor (FcRn) to belantamab mafodotin was measured using surface plasmon resonance (SPR).

[0561] Reference standard #182407660A was used. The results are summarized in Tables 20 and 21.

[0562] Table 20: PTM abundance in heat-treated belantamab mafodotin

[0563]

[0564] Table 21: Binding activity of heat-treated belantamab mafodotin

[0565]

[0566] Extrapolating from the isomerization data of HC Asp 130, up to 23% isomerization can result in at least 70% antigen-specific binding activity. This was calculated using the linear slope of the time 0 (4.1 - 4.4%) and 28-day (28.7 - 29.3%) D103 isomerized samples, which had 88 - 99% and 55 - 62% antigen-specific binding activity, respectively.

[0567] Example 15: Light exposure

[0568] To create light-exposed samples, belantamab mafodotin samples were diluted to 10 mg / mL and loaded into glass vials, then transferred to a Caron light stability chamber at 25°C for varying degrees of light exposure as shown in Table 10.

[0569] As described in Example 11, oxidation, deamidation, and isomerization were determined using tryptic peptide mapping tandem mass spectrometry (peptide mapping LC-MS / MS). The extracted ion chromatograms of the unmodified and modified peptides were used to calculate the levels of oxidation, deamidation, or isomerization by dividing the area under the curve of the modified peptide by the total area under the curves of both the modified and unmodified peptides

[0570] As described in Example 4, surface plasmon resonance (SPR) was used to measure the binding of belantamab mafodotin to BCMA and FcγRIIIa. As described in Example 6, surface plasmon resonance (SPR) was used to measure the binding of the neonatal Fc receptor (FcRn) to belantamab mafodotin.

[0571] Reference standard #182407660A was used. The results are summarized in Tables 22 and 23.

[0572] Table 22: PTM Abundances in Photo-Treated Belantamab Mafodotin

[0573]

[0574] Table 23: Binding Activities of Photo-Treated Belantamab Mafodotin

[0575]

[0576] Example 16: C-Terminal Cleavage and N-Terminal Pyroglutamic Acid

[0577] As described in Example 11, tryptic peptide mapping tandem mass spectrometry (peptide mapping LC-MS / MS) was used to analyze the N-terminal pyroglutamic acid levels and C-terminal cleavage of several batches of belantamab. The extracted ion chromatograms of the unmodified and modified peptides were used to calculate the levels of C-terminal cleavage and N-terminal pyroglutamic acid by dividing the area under the curve of the modified peptide by the total area under the curves of both the modified and unmodified peptides

[0578] As described in Example 4, surface plasmon resonance (SPR) was used to measure the binding of belantamab to BCMA and FcγRIIIa. As described in Example 6, surface plasmon resonance (SPR) was used to measure the binding of the neonatal Fc receptor (FcRn) to belantamab.

[0579] The reference standards used were #122368059 and #172405900. The results are summarized in Tables 24 - 25.

[0580] Table 24: Abundances of Pyroglutamic Acid and Lysine Cleavage in Belantamab and the Corresponding Activities

[0581]

[0582] Table 25: Abundances of pyroglutamic acid and lysine cleavage in belantamab and corresponding activities

[0583]

[0584] Example 17: Glycosylation

[0585] The glycosylation patterns of several batches of belantamab were analyzed. Curves were determined using ultra-performance liquid chromatography (UPLC) with hydrophilic interaction liquid chromatography (HILIC) separation and fluorescence detection. Samples were diluted with water to a concentration of 10 μg / μL, and glycans were released from belantamab by enzymatic cleavage with PNGaseF using the PNGaseF kit from New England BioLabs (Cat #P0705L). The PNGaseF-released glycans were labeled with o-aminobenzamide (Sigma-Aldrich, Cat #A89804). The labeled glycans were then purified using an HILIC column step to remove excess labeling solution; the glycans were loaded and washed with water and eluted with acetonitrile. The labeled glycans were then separated using a Waters Glycan BEH Amide column (part number 186004742) on a Waters Acquity UPLC with an ammonium formate / formic acid and acetonitrile gradient. Glycans were detected using fluorescence detection with excitation at 365 nm and emission at 438 nm. Quantification of the glycans was achieved by dividing the area under the curve of the glycans by the total area under the curve of all detected glycans.

[0586] The reference standard used was #122368059. The results are summarized in Tables 26 - 27.

[0587] Table 26: Glycosylation patterns of belantamab

[0588]

[0589] Table 27: Glycosylation patterns of belantamab

[0590]

[0591] Example 18: Glycoengineering

[0592] The effects of a sugar-rich belantamab sample on ADCC activity and binding were determined. ADCC activity was measured as in Example 3. BCMA and FcγRIIIa binding was determined as in Example 4.

[0593] For galactosylation experiments, glycosylation was measured using reduced LC-MS. Samples were diluted to 1 mg / mL, 50 uL of 1 M DTT was added and reacted at 25 °C or 37 °C for 30 minutes, and then analyzed on a mass spectrometer that may include a Micromass Q-tof. Size-exclusion chromatography with an isocratic flow of water, acetonitrile, and trifluoroacetic acid was used to separate the heavy and light chains. Spectra for each heavy and light chain were summed and deconvoluted using MaxEnt software from Waters. The major glycoforms were detected and the relative amounts were estimated based on signal counts or area under the curve. The results are summarized in Tables 28 - 29.

[0594] Table 28: Glycan-rich samples of belantamab and corresponding activities

[0595]

[0596] Table 29: Abundances (%) of glycan-rich samples of belantamab

[0597]

[0598] Example 19: Tolerable range

[0599] The tolerable range (70 - 130% activity) was determined by using the data at the first and last time points in Tables 11 to 18 (day 21 or day 28, as appropriate). Binding data were plotted against the relative percentages of relevant post-translational modifications to determine the slope of the relationship. Using this information, predicted levels for each post-translational modification were calculated for at least 70% of the binding measurements. The results of this extrapolation are summarized in Table 30. The trends reported broadly reflect the observations seen with belantamab or belantamab mafodotin.

[0600] Table 30: Extrapolation of data for functional variants of belantamab and belantamab mafodotin

[0601]

[0602]

[0603] Example 20: Degradation product summary

[0604] The incidence of degradation products that have no effect on the activity of belantamab mafodotin observed under different forced degradation conditions is summarized below (Table 31).

[0605] Table 31: Degradation products that do not affect the potency of belantamab mafodotin

[0606]

[0607] Example 21

[0608] The study designs for the forced degradation of belantamab in Examples 21 to 26 are summarized and depicted in Table 32. The methods were generally similar to those for belantamab mafodotin described in Examples 11 - 18 above (unless otherwise stated).

[0609] Table 32: Study Designs for the Forced Degradation of Belantamab

[0610]

[0611] 1. RH: Relative Humidity

[0612] Example 22: Oxidation Conditions of Belantamab

[0613] After 24 hours, the oxidation of HC M256 increased from approximately 2% to approximately 98%, and the oxidation of M432 increased from approximately 1% to approximately 96%. The specific binding of SPR to FcγRIIIa decreased by 21 - 25%, and the binding to FcRn decreased by 10 - 18%. Oxidation in the Fc may alter the binding activity of belantamab to FcγRIIIa and FcRn. After 24 hours, the oxidation of HC M34 in CDR1 increased from approximately 0.3% to 47.7 - 48.5%, resulting in no change in antigen binding within the assay variability. The levels of cysteine and tryptophan oxidation were low throughout the study, and no other significant post - translational modifications were detected.

[0614] Example 23: Base Treatment of Belantamab

[0615] Isomerization was observed to increase from approximately 3.5% to approximately 6.5% in HC D103 after 28 days. After 28 days, deamidation increased from 0.1% to approximately 2.5% in HC N31; from approximately 2.0% to approximately 10% in HC N388, and from approximately 1.7% to approximately 18.5% in HC N393. In addition, an increase in the oxidation of HC M256 from approximately 2.2% to approximately 3.7% was also observed in belantamab stressed at pH 9 for 28 days. cIEF analysis showed an increase in acidic variants from approximately 25% to approximately 62%; and a decrease in basic variants from approximately 9% to approximately 4.5% (see Table 33 below). All the changes observed were within the assay variability of antigen, FcγRIIIa, and FcRn specific binding; thus, the binding of SPR was comparable to belantamab stressed at pH 9.0 for 28 days.

[0616] Table 33: cIEF Results of Base - Treated Belantamab

[0617]

[0618] Example 24: Acid Treatment of Belantamab

[0619] After 28 days, the fragment increased from 0.8% to 2.3 - 2.7%. After 28 days, the formation of succinimide at HC D103 was observed to increase from 0.2% to 4.0%, and the isomerization of HC D103 increased from approximately 3.5% to approximately 5.8%. In addition to aspartic acid isomerization, an increase in HC M256 oxidation from approximately 2.2% to approximately 3.7% was also observed in belantamab after 28 days of pH 3.5 stress. cIEF analysis showed an increase in acidic variants from approximately 25% to approximately 28%; and an increase in basic variants from approximately 9% to approximately 13% (see Table 34 below). All changes observed in antigen, FcγRIIIa, and FcRn specific binding were within the assay variability; thus, the binding of SPR was comparable to belantamab after 28 days of pH 3.5 stress.

[0620] Table 34: cIEF Results of Acid - Treated Belantamab

[0621]

[0622] Example 25: Heat Treatment of Belantamab

[0623] After 28 days, the fragment increased from 0.8% to 2.2 - 2.3%. The % polymerization did not change. After 28 days, the formation of succinimide at HC D103 was observed to increase from 0.2% to 2.2%, and the isomerization of HC D103 increased from approximately 3.5% to approximately 29%. In addition to aspartic acid isomerization, an increase in oxidation in HC M256 from approximately 2.2% to approximately 5.3% and in HC M432 from approximately 1% to 2% was also observed in belantamab after 28 days of heat stress. After 28 days, deamidation increased from approximately 0% to approximately 7% in HC N329, from approximately 2.0% to approximately 2.5% in HC N388, and from approximately 1.7% to approximately 2.6% in HC N393. Antigen specific binding decreased to 63 - 67% at 28 days, which was consistent with the increase in HC D103 isomerization, which has been shown to affect antigen binding. The changes observed in FcγRIIIa and FcRn specific binding were within the assay variability.

[0624] Example 26: Light Treatment of Belantamab

[0625] At 1.5X ICH, the fragmentation increased from 0.8% to 1.5%, and the aggregation increased from approximately 1% to 6.5 - 7.5%. Oxidation at 1.5X ICH increased from approximately 0.3% to 1.6 - 2.1% in HCM34, from approximately 2.2% to 18.4 - 25.1% in HCM256, and from approximately 1% to 13.7 - 19.5% in HCM432. cIEF analysis showed that the acidic variants increased from approximately 25% to approximately 34%; the basic variants did not change (see Table 35 below). All changes observed in antigen, FcγRIIIa, and FcRn specific binding were within the assay variability; thus, the binding of SPR was comparable to that of belantamab after pH 3.5 stress for 28 days.

[0626] Table 35: cIEF Results of Photo - Treated Belantamab

[0627]

[0628] The forced degradation study of belantamab showed results consistent with the above - mentioned forced degradation study of belantamab mofetil, with the only exception being that up to 48.5% oxidation at HC M34 resulted in no change in antigen binding, within the assay variability.

[0629] Notably, cIEF data for belantamab mofetil were not presented because the drug payload contributed to the charge profile, while for belantamab, cIEF effectively separated the acidic and basic variants from the major species (see Figure 4 ).

[0630] Sequence listing

[0631] SEQ. ID. NO. 1 – CDRH1

[0632] NYWMH

[0633] SEQ. ID. NO. 2: CDRH2

[0634] ATYRGHSDTYYNQKFKG

[0635] SEQ. ID. NO. 3: CDRH3

[0636] GAIYDGYDV L DN

[0637] SEQ. ID. NO. 4: CDRL1

[0638] SASQDISNYLN

[0639] SEQ. ID. NO. 5: CDRL2

[0640] YTSNLHS

[0641] SEQ. ID. NO. 6: CDRL3

[0642] QQYRKLPWT

[0643] SEQ. ID. NO. 7: Heavy chain variable region (CDRs underlined)

[0644] QVQLVQSGAEVKKPGSSVKVSCKASGGTFS NYWMH WVRQAPGQGLEWMG ATYRGHSDTYYNQKFKG RVTITADKSTSTAYMELSSLRSEDTAVYYCAR GAIYDGYDV L DN WGQGTLVTVSS

[0645] SEQ. ID. NO. 8: Light chain variable region (CDRs underlined)

[0646] DIQMTQSPSSLSASVGDRVTITC SASQDISNYLN WYQQKPGKAPKLLIY YTSNLHS GVPSRFSGSGSGTDFTLTISSLQPEDFATYYC QQYRKLPWT FGQGTKLEIKR

[0647] SEQ. ID. NO. 9: Heavy chain region (CDRs underlined)

[0648] QVQLVQSGAEVKKPGSSVKVSCKASGGTFS NYWMH WVRQAPGQGLEWMG ATYRGHSDTYYNQKFKG RVTITADKSTSTAYMELSSLRSEDTAVYYCAR GAIYDGYDV L DN WGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGV H TFPAV L QSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEV H NAKTKPREEQYNSTYRVVSV L TV LHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPV L DSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK

[0649] SEQ. ID. NO. 10: Light chain region (CDRs underlined)

[0650] DIQMTQSPSSLSASVGDRVTITC SASQDISNYLN WYQQKPGKAPKLLIY YTSNLHS GVPSRFSGSGSGTDFTLTISSLQPEDFATYYC QQYRKLPWT FGQGTKLEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC

[0651] SEQ. ID. NO. 11: Heavy chain region with D103N (CDRs underlined)

[0652] QVQLVQSGAEVKKPGSSVKVSCKASGGTFS NYWMH WVRQAPGQGLEWMG ATYRGHSDTYYNQKFKG RVTITADKSTSTAYMELSSLRSEDTAVYYCAR GAIYNGYDV L DN WGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGV H TFPAV L QSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEV H NAKTKPREEQYNSTYRVVSV L TV LHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPV L DSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK

[0653] SEQ. ID. NO. 12: Heavy chain region with N388D (CDRs underlined)

[0654] QVQLVQSGAEVKKPGSSVKVSCKASGGTFS NYWMH WVRQAPGQGLEWMG ATYRGHSDTYYNQKFKG RVTITADKSTSTAYMELSSLRSEDTAVYYCAR GAIYDGYDV L DN WGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGV H TFPAV L QSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEV H NAKTKPREEQYNSTYRVVSV L TV L HQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESDGQPENNYKTTPPV L DSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK

[0655] SEQ. ID. NO. 13: Heavy chain region with N393D (CDRs underlined)

[0656] QVQLVQSGAEVKKPGSSVKVSCKASGGTFS NYWMH WVRQAPGQGLEWMG ATYRGHSDTYYNQKFKG RVTITADKSTSTAYMELSSLRSEDTAVYYCAR GAIYDGYDV L DNWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGV H TFPAV L QSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEV H NAKTKPREEQYNSTYRVVSV L TV L HQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPEDNYKTTPPV L DSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK

[0657] SEQ. ID. NO. 14: Heavy chain region with N388D and N393D (CDRs underlined)

[0658] VQLVQSGAEVKKPGSSVKVSCKASGGTFS NYWMH WVRQAPGQGLEWMG ATYRGHSDTYYNQKFKG RVTITADKSTSTAYMELSSLRSEDTAVYYCAR GAIYDGYDV L DN WGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGV H TFPAV L QSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEV H NAKTKPREEQYNSTYRVVSV L TV L HQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESDGQPEDNYKTTPPV LDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK。

Claims

1. A composition comprising an isomerization variant of an anti-BCMA antibody, wherein the isomerization variant comprises a heavy chain amino acid sequence comprising CDRH1 of SEQ ID NO: 1, CDRH2 of SEQ ID NO: 2, and CDRH3 of SEQ ID NO: 3, and a light chain amino acid sequence comprising CDRL1 of SEQ ID NO: 4, CDRL2 of SEQ ID NO: 5, and CDRL3 of SEQ ID NO: 6; wherein the composition comprises ≤25% of the isomerization variant.

2. A composition comprising an oxidation variant of an anti-BCMA antibody, wherein the oxidation variant comprises a heavy chain amino acid sequence comprising CDRH1 of SEQ ID NO: 1, CDRH2 of SEQ ID NO: 2, and CDRH3 of SEQ ID NO: 3, and a light chain amino acid sequence comprising CDRL1 of SEQ ID NO: 4, CDRL2 of SEQ ID NO: 5, and CDRL3 of SEQ ID NO: 6; wherein the composition comprises ≤40% of the oxidation variant.

3. A composition comprising an anti-BCMA antibody, the anti-BCMA antibody comprising CDRH1 having the amino acid sequence set forth in SEQ ID NO: 1, CDRH2 having the amino acid sequence set forth in SEQ ID NO: 2, CDRH3 having the amino acid sequence set forth in SEQ ID NO: 3, CDRL1 having the amino acid sequence set forth in SEQ ID NO: 4, CDRL2 having the amino acid sequence set forth in SEQ ID NO: 5, and CDRL3 having the amino acid sequence set forth in SEQ ID NO: 6; wherein the composition comprises 0.1-25% isomerization at D103 of CDRH3.

4. A composition comprising an anti-BCMA antibody, the anti-BCMA antibody comprising CDRH1 having the amino acid sequence set forth in SEQ ID NO: 1, CDRH2 having the amino acid sequence set forth in SEQ ID NO: 2, CDRH3 having the amino acid sequence set forth in SEQ ID NO: 3, CDRL1 having the amino acid sequence set forth in SEQ ID NO: 4, CDRL2 having the amino acid sequence set forth in SEQ ID NO: 5, and CDRL3 having the amino acid sequence set forth in SEQ ID NO: 6; wherein the composition comprises 0.1-40% oxidation at M34 of CDRH1.

5. A composition comprising an anti-BCMA antibody that is at least about 90% identical to the heavy chain amino acid sequence of SEQ ID NO: 9 and the light chain amino acid sequence of SEQ ID NO: 10, wherein the composition comprises 0.1-25% isomerization at D103 of CDRH3.

6. A composition comprising an anti-BCMA antibody that is at least about 90% identical to the heavy chain amino acid sequence of SEQ ID NO: 9 and the light chain amino acid sequence of SEQ ID NO: 10, wherein the composition comprises 0.1-40% oxidation at M34 in CDRH1.

7. The composition according to any one of the preceding claims, wherein the composition comprises ≤65% oxidation at M256 in the heavy chain and / or ≤60% oxidation at M432 in the heavy chain.

8. The composition according to any one of the preceding claims, wherein the composition comprises an antibody variant comprising at least one selected from the group consisting of: deamidation of the heavy chain at N388 and / or N393, D103 to N103 in CDRH3, C-terminal lysine cleavage, and conversion of N-terminal glutamine to pyroglutamic acid.

9. The composition according to any one of the preceding claims, wherein the composition comprises at least one selected from the group consisting of: up to 100% deamidation at N388 and / or N393, up to 100% N103 in CDRH3, up to 100% C-terminal lysine cleavage, and up to 100% conversion of N-terminal glutamine to pyroglutamic acid.

10. The composition according to any one of the preceding claims, wherein the composition comprises any percentage of glycoform G0, G1, G2, G0-GlcNac or G0-2GlcNac.

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