Antibodies binding to C3bBb

Abstract

The present invention relates to anti-C3bBb antibodies and methods of use thereof.

Description

Technical Field

[0001] This invention relates to anti-C3bBb antibodies and methods of using them. Background Technology

[0002] The complement system is closely associated with the disease biology of age-related macular degeneration (AMD) (Cell Mol LifeSci 2021 May; 78(10): 4487-4505). The alternative pathway C3 convertase (C3bBb) is a key amplification factor in the complement cascade, formed by the binding of factor B (FB) to the C3b fragment of complement component C3, followed by cleavage by plasma serine protease factor D (FD) upon fragment Ba release. The remaining Bb fragment bound to C3b forms the functional C3 convertase C3bBb. C3bBb is relatively unstable, with a half-life of several minutes, making it unsuitable as a target for antibody production.

[0003] Given the central role of the bypass pathway in various pathologies, there remains a need to develop potent inhibitors that act at various complement components within this proteolytic cascade. Summary of the Invention

[0004] This invention relates to antibodies that specifically bind to human C3bBb.

[0005] This invention also relates to an antibody that binds to human C3bBb, wherein the antibody:

[0006] - Combined with wild-type C3bBb, and / or

[0007] - Binds to recombinant human C3bBb, which contains the Bb subunit of the recombinant factor B (FB) protein, which contains the D279G mutation and optionally the mutations K350N and / or M458I; and / or

[0008] - Suppress bypass pathways; and / or

[0009] - An agonist or antagonist of C3bBb activity; and / or

[0010] -Specifically binds to cynomolgus monkey C3bBb and human C3bBb, and / or

[0011] -Specifically binds to African green monkey C3bBb and human C3bBb, and / or

[0012] -Binding with an affinity of ≤ 10 nM at 37°C, as measured by SPR, and / or

[0013] -Does not compete with the prepared solution for C3bBb binding, and / or

[0014] - Do not stabilize C3 invertase.

[0015] In one aspect, the present invention relates to antibodies that specifically bind to wild-type human C3bBb.

[0016] In another aspect, the present invention relates to an antibody that specifically binds to recombinant human C3bBb, wherein C3bBb comprises the Bb subunit of a recombinant FB protein comprising D279G and optionally further mutants K350N and / or M458I. In one embodiment, the antibody specifically binds to human C3bBb comprising the Bb subunit of the recombinant FB protein according to SEQ ID NO: 492. In one embodiment, the antibody specifically binds to human C3bBb comprising the Bb subunit of the recombinant FB protein according to SEQ ID NO: 493.

[0017] In another aspect, the present invention relates to an antibody that specifically binds to an epitope of human C3bBb, the epitope comprising amino acid residues of a C3b subunit and an amino acid residue of a Bb subunit of human FB. In one embodiment, the antibody of the present invention specifically binds to human C3bBb, wherein the antibody does not specifically bind to the (isolated) C3b subunit of C3bBb, and wherein the antibody does not specifically bind to the (isolated) Bb subunit of C3bBb.

[0018] In another aspect, the present invention relates to an antibody that binds to human C3bBb, wherein the antibody comprises a heavy chain CDR and a light chain CDR selected from antibodies in Table D1. In one embodiment, the antibody comprises a heavy chain variable domain and a light chain variable domain selected from antibodies in Table D1.

[0019] In one embodiment, the antibody comprises a heavy chain variable domain (VH) comprising: (a) CDR-H1 comprising the amino acid sequence of SEQ ID NO:449; (b) CDR-H2 comprising the amino acid sequence of SEQ ID NO:450; and (c) CDR-H3 comprising the amino acid sequence of SEQ ID NO:451; and a light chain variable domain (VL) comprising: (d) CDR-L1 comprising the amino acid sequence of SEQ ID NO:452; (e) CDR-L2 comprising the amino acid sequence of SEQ ID NO:453; and (f) CDR-L3 comprising the amino acid sequence of SEQ ID NO:454 (corresponding to antibody #1, P1AG9426); or a heavy chain variable domain (VH) comprising: (a) CDR-H1 comprising the amino acid sequence of SEQ ID NO:455; (b) CDR-H2 comprising the amino acid sequence of SEQ ID NO:456; and (c) The CDR-H3 comprising the amino acid sequence of SEQ ID NO:457; and a light chain variable domain (VL) comprising: (d) a CDR-L1 comprising the amino acid sequence of SEQ ID NO:458; (e) a CDR-L2 comprising the amino acid sequence of SEQ ID NO:459; and (f) a CDR-L3 comprising the amino acid sequence of SEQ ID NO:460 (corresponding to antibody #2, P1AG9376); or a heavy chain variable domain (VH) comprising: (a) a CDR-H1 comprising the amino acid sequence of SEQ ID NO:461; (b) a CDR-H2 comprising the amino acid sequence of SEQ ID NO:462; and (c) a CDR-H3 comprising the amino acid sequence of SEQ ID NO:463; and a light chain variable domain (VL) comprising: (d) a CDR-L1 comprising the amino acid sequence of SEQ ID NO:464; (e) a CDR-L3 comprising the amino acid sequence of SEQ ID NO:464; (f) a CDR-L3 comprising the amino acid sequence of SEQ ID NO:465; (g) a CDR-L3 comprising the amino acid sequence of SEQ ID NO:465; (h ... CDR-L2 containing the amino acid sequence of SEQ ID NO:465; and (f) CDR-L3 containing the amino acid sequence of SEQ ID NO:466 (corresponding to antibody #3, P1AG9372); or heavy chain variable domain (VH) comprising: (a) CDR-H1 containing the amino acid sequence of SEQ ID NO:467; (b) CDR-H2 containing the amino acid sequence of SEQ ID NO:468; and (c) CDR-H3 containing the amino acid sequence of SEQ ID NO:469;and a light chain variable domain (VL) comprising: (d) a CDR-L1 comprising the amino acid sequence of SEQ ID NO:470; (e) a CDR-L2 comprising the amino acid sequence of SEQ ID NO:471; and (f) a CDR-L3 comprising the amino acid sequence of SEQ ID NO:472 (corresponding to antibody #4, P1AG9420); or a heavy chain variable domain (VH) comprising: (a) a CDR-H1 comprising the amino acid sequence of SEQ ID NO:473; (b) a CDR-H2 comprising the amino acid sequence of SEQ ID NO:474; and (c) a CDR-H3 comprising the amino acid sequence of SEQ ID NO:475; and a light chain variable domain (VL) comprising: (d) a CDR-L1 comprising the amino acid sequence of SEQ ID NO:476; (e) a CDR-L2 comprising the amino acid sequence of SEQ ID NO:477; and (f) The CDR-L3 comprising the amino acid sequence of SEQ ID NO:478 (corresponding to antibody #5, P1AG9391); or the heavy chain variable domain (VH) comprising: (a) the amino acid sequence of SEQ ID NO:479; (b) the amino acid sequence of SEQ ID NO:480; and (c) the amino acid sequence of SEQ ID NO:481; and the light chain variable domain (VL) comprising: (d) the amino acid sequence of SEQ ID NO:482; (e) the amino acid sequence of SEQ ID NO:483; and (f) the amino acid sequence of SEQ ID NO:484 (corresponding to antibody #6, P1AH1205); or the heavy chain variable domain (VH) comprising: (a) the amino acid sequence of SEQ ID NO:485; (b) the amino acid sequence of SEQ ID NO:482; (c) the amino acid sequence of SEQ ID NO:483; (d) the amino acid sequence of SEQ ID NO:484; (e) the amino acid sequence of SEQ ID NO:485; (f) the amino acid sequence of SEQ ID NO:484; (c) the amino acid sequence of SEQ ID NO:485; (d) the amino acid sequence of SEQ ID NO:484; (e) the amino acid sequence of SEQ ID NO:485; (f) the amino acid sequence of SEQ ID NO:484; (g) the amino acid sequence of SEQ ID NO:485; (g ... (a) CDR-H1 containing the amino acid sequence of SEQ ID NO:486; (b) CDR-H2 containing the amino acid sequence of SEQ ID NO:487; and (c) CDR-H3 containing the amino acid sequence of SEQ ID NO:487; and a light chain variable domain (VL) comprising: (d) CDR-L1 containing the amino acid sequence of SEQ ID NO:488; (e) CDR-L2 containing the amino acid sequence of SEQ ID NO:489; and (f) CDR-L3 containing the amino acid sequence of SEQ ID NO:490 (corresponding to antibody #7, P1AH1199).

[0020] In one embodiment, the antibody comprises a VH domain containing SEQ ID NO:1 and a VL domain containing SEQ ID NO:2 (corresponding to antibody #1, P1AG9426); or a VH domain containing SEQ ID NO:3 and a VL domain containing SEQ ID NO:4 (corresponding to antibody #2, P1AG9376); or a VH domain containing SEQ ID NO:5 and a VL domain containing SEQ ID NO:6 (corresponding to antibody #3, P1AG9372); or a VH domain containing SEQ ID NO:7 and a VL domain containing SEQ ID NO:8 (corresponding to antibody #4, P1AG9420); or a VH domain containing SEQ ID NO:9 and a VL domain containing SEQ ID NO:10 (corresponding to antibody #5, P1AG9391); or a VH domain containing SEQ ID NO:11 and a VL domain containing SEQ ID NO:12. The domain (corresponding to antibody #6, P1AH1205); or containing the VH domain of SEQ ID NO:13 and the VL domain of SEQ ID NO:14 (corresponding to antibody #7, P1AH1199).

[0021] Furthermore, this invention relates to an antibody that competes with the antibody of this invention for binding to wild-type human C3bBb.

[0022] The present invention also relates to an antibody that competes with the antibody of the present invention for binding to recombinant human C3bBb, the recombinant human C3bBb comprising the Bb subunit of a recombinant FB protein comprising D279G and optionally further mutants K350N and / or M458I.

[0023] Another aspect of the present invention is an isolated nucleic acid encoding the antibody of the present invention.

[0024] The present invention also relates to a host cell comprising the nucleic acid of the present invention.

[0025] Furthermore, the present invention relates to a method for generating an antibody that binds to human C3bBb, the method comprising culturing the host cells of the present invention under conditions suitable for antibody expression.

[0026] In another aspect, the present invention relates to a pharmaceutical composition comprising an antibody according to the present invention.

[0027] In another aspect, the present invention relates to the antibodies of the present invention used as pharmaceuticals.

[0028] The antibodies of the present invention are suitable for inhibiting the bypass pathway by binding to C3bBb. The antibodies of the present invention can inhibit the bypass pathway upon binding without stabilizing C3bBb, thereby avoiding the accumulation of C3bBb. Furthermore, the antibodies of the present invention are specific for the C3bBb complex but do not exhibit significant antigenic binding to any of its individual subunits.

[0029] The antibodies disclosed in this article can be applied to therapies, particularly for the treatment of ocular vascular diseases.

[0030] The antibodies of this invention offer several valuable properties that allow for their therapeutic applications, such as high affinity and high stability that is beneficial for long-term use. Furthermore, the antibodies of this invention are advantageously available in high-concentration liquid formulations with a viscosity suitable for ocular applications. Attached Figure Description

[0031] Figure 1: The left panel shows the trimer complex formed between the P1AF8499 Fab fragment and C3bBb. The epitope bound by the P1AF8499 Fab fragment contains amino acids from both components, C3b and Bb (right panel).

[0032] Figure 2: Binding of antibody P1AG9426 (#1) to wild-type human C3bBb complex. Sensing map generated as described in Example 14. Detailed Implementation

[0033] 1. definition

[0034] For the purposes of this document, a “recipient human frame” is a frame containing an amino acid sequence derived from a light chain variable domain (VL) frame or a heavy chain variable domain (VH) frame of a human immunoglobulin frame or a human common frame as defined below. A recipient human frame “derived from” a human immunoglobulin frame or a human common frame may contain the same amino acid sequence as that human immunoglobulin frame or human common frame, or it may contain amino acid sequence variations. In some aspects, the number of amino acid variations is 10 or fewer, 9 or fewer, 8 or fewer, 7 or fewer, 6 or fewer, 5 or fewer, 4 or fewer, 3 or fewer, or 2 or fewer. In some aspects, the VL recipient human frame is sequenceally identical to the VL human immunoglobulin frame sequence or the human common frame sequence.

[0035] “Affinity” refers to the strength of the sum of non-covalent interactions between a single binding site of a molecule (e.g., an antibody) and its binding partner (e.g., an antigen). Unless otherwise stated, as used herein, “binding affinity” refers to the intrinsic binding affinity that reflects a 1:1 interaction between members of a binding pair (e.g., antibody and antigen). The affinity of molecule X for its partner Y can generally be represented by the dissociation constant (KD). Affinity can be measured by conventional methods known in the art, including those described herein. Specific illustrative and exemplary methods for measuring binding affinity are described below. The dissociation constant (KD) of an antibody binding to human C3bBb is... D The nM values ​​are ≤ 1 μM, ≤ 100 nM, ≤ 10 nM, ≤ 1 nM, ≤ 0.1 nM, ≤ 0.01 nM, or ≤ 0.001 nM, and in one embodiment, 10 nM. -6 M or smaller, in one embodiment 10 -7 M or smaller, in one embodiment 10 -8 M or smaller, in one embodiment 10 -8 M to 10 -13 M, in one embodiment, is 10 -9 M to 10 -13 M. When the KD of an antibody is 1 μM or less, the antibody is said to "specifically bind" to human C3bBb. In some respects, anti-human C3bBb antibodies bind to epitopes of human C3bBb that are conserved in human C3bBb from different sources.

[0036] Components of the innate immune system, namely the different elements from the classical and alternative pathways, are referred to herein according to their standard meaning in the art, and the terminology used herein includes recombinant variants of the corresponding natural components of the innate immune system that maintain function, and may include variants containing labels and / or tags.

[0037] In short, as used herein, the term "C3bBb" refers to the bypass pathway C3 convertase, which is formed by the binding of a C3 fragment ("C3b") to factor B ("FB") and subsequent cleavage by plasma serine protease factor D ("FD") upon release of fragment Ba. The term "C3b" as used herein refers to the larger of two elements produced by bypass cleavage of complement component 3 (C3), the other being C3a. The term "C3 convertase" refers to a serine protease of the complement system and may refer to either the bypass pathway C3 convertase (C3bBb) or the classical and lectin pathway C3 convertase (C4bC2b). The terms "human factor B" or "FB" refer to complement factor B encoded by the CFB gene in humans. The term "human factor D" refers to complement factor D encoded by the CFD gene in humans. The term "C5 convertase" as used herein refers to the bypass pathway C5 convertase (C3bBbC3b).

[0038] The term "antibody screening" refers to a technique used to identify antibodies that specifically bind to a target antigen. Various techniques for identifying antibodies are known in the art, such as administering immunogenic target antigens to transgenic animals, hybridoma-based methods, or isolating variable domain sequences selected from phage display libraries. The term "phage library panning" refers to an affinity selection technique used to identify phage display variants that possess the desired binding properties against a target antigen.

[0039] The term “antibody” is used in the broadest sense and includes a variety of antibody structures, including but not limited to monoclonal antibodies, polyclonal antibodies, multispecific antibodies (e.g., bispecific antibodies), and antibody fragments, as long as they exhibit the desired antigen-binding activity.

[0040] "Antibody fragment" refers to a molecule other than a complete antibody that contains a portion of the complete antibody and binds to the antigen bound by the complete antibody. Examples of antibody fragments include, but are not limited to, Fv, Fab, Fab', Fab'-SH, F(ab')2; bisomatic antibodies; linear antibodies; single-chain antibody molecules (e.g., scFv and scFab); single-domain antibodies (dAb); and multispecific antibodies formed from antibody fragments. For a review of some antibody fragments, see Holliger and Hudson, Nature Biotechnology 23:1126-1136 (2005).

[0041] The term "epitope" refers to a site on a protein or non-protein antigen that binds to the anti-human C3bBb antibody. The epitopes bound by the antibodies of this invention contain discontinuous amino acids and are therefore conformational epitopes.

[0042] Screening for antibodies that bind to specific epitopes (i.e., those antibodies that bind to the same epitope) can be performed using methods conventional in the art, such as, but not limited to, alanine scanning, Western blotting (see Meth. Mol. Biol. 248 (2004) 443-463), peptide cleavage analysis, epitope excision, epitope extraction, chemical modification of antigens (see Prot. Sci. 9 (2000) 487-496), and cross-blocking (see “Antibodies”, Harlow and Lane (Cold Spring Harbor Press, NY)). In some embodiments used herein, the epitopes of the antibodies are identified by cryo-electron microscopy.

[0043] Furthermore, competitive binding can be used to readily determine whether an antibody competes with the reference anti-human C3bBb antibody of the present invention for binding to human C3bBb. For example, "antibody that competes with the reference anti-human C3bBb antibody for binding" refers to an antibody that blocks the binding of the reference anti-human C3bBb antibody to its antigen by 50% or more in a competitive assay, and conversely, the reference antibody that blocks the binding of the antibody to its antigen by 50% or more in a competitive assay.

[0044] In some respects, such as as measured in competitive binding assays, if one antibody inhibits the binding of another antibody by 1, 5, 10, 20, or 100 times more than the other, the two antibodies are considered to be competitively binding to human C3bBb (see, for example, Junghans et al., Cancer Res. 50 (1990) 1495-1502).

[0045] An antibody's "class" refers to the type of constant domain or constant region possessed by its heavy chain. There are five major classes of antibodies: IgA, IgD, IgE, IgG, and IgM, and some of them can be further subdivided into subclasses (isotypes), such as IgG1, IgG2, IgG3, IgG4, IgA1, and IgA2. In some respects, antibodies are IgG1 isotypes. In some respects, antibodies are IgG1 isotypes with P329G, L234A, and L235A mutations that reduce Fc region effector function. In other respects, antibodies are IgG2 isotypes. In some respects, antibodies are IgG4 isotypes, containing an S228P mutation in the hinge region to improve the stability of IgG4 antibodies. The constant domains of the heavy chain corresponding to different classes of immunoglobulins are called α, δ, ε, γ, and μ, respectively. The light chain of an antibody can be assigned to one of two types based on the amino acid sequence of its constant domain, referred to as kappa (κ) and lambda (λ). As used in this application, the term "constant region derived from human origin" or "human constant region" refers to the constant heavy chain region and / or constant light chain κ or λ region of a human antibody of subclass IgG1, IgG2, IgG3, or IgG4. Such constant regions are well known in the art and are described, for example, by the following: Kabat, EA et al., Sequences of Proteins of Immunological Interest, 5th edition, Public Health Service, National Institutes of Health, Bethesda, MD (1991) (see also, for example, Johnson, G. and Wu, TT, Nucleic Acids Res. 28 (2000) 214-218; Kabat, EA et al., Proc. Natl. Acad. Sci. USA 72 (1975) 2785-2788). Unless otherwise specified herein, the amino acid residues in the constant region are numbered according to the EU numbering system, also known as Kabat's EU index, as described in Kabat, EA et al., Sequences of Proteins of Immunological Interest, 5th edition, Public Health Service, National Institutes of Health, Bethesda, MD (1991), NIH Publication 91-3242.

[0046] "Effective functions" refer to those biological activities attributable to the Fc region of an antibody that vary with antibody isotype. Examples of antibody effector functions include: C1q binding and complement-dependent cytotoxicity (CDC); Fc receptor binding; antibody-dependent cell-mediated cytotoxicity (ADCC); phagocytosis; downregulation of cell surface receptors (e.g., B cell receptors); and B cell activation.

[0047] The “effective amount” of a pharmaceutical agent (e.g., a pharmaceutical composition) refers to the amount that is sufficient to effectively achieve the desired therapeutic or preventative outcome at the necessary dose for the necessary period of time.

[0048] The term "Fc region" used herein is used to define the C-terminal region of an immunoglobulin heavy chain that comprises at least a portion of the constant region. This term includes both native sequence Fc regions and variant Fc regions. In one aspect, the human IgG heavy chain Fc region extends from Cys226 or Pro230 to the C-terminus of the heavy chain. However, antibodies produced by host cells can undergo post-translational cleavage of one or more (particularly one or two) amino acids from the C-terminus of the heavy chain. Therefore, antibodies produced by host cells by expressing a specific nucleic acid molecule encoding the full-length heavy chain can comprise the full-length heavy chain, or the antibody can comprise a cleaved variant of the full-length heavy chain. This could be the case where the last two C-terminal amino acids of the heavy chain are glycine (G446) and lysine (K447, EU numbering system). Therefore, the C-terminal lysine (Lys447) or the C-terminal glycine (Gly446) and lysine (Lys447) of the Fc region may or may not be present. Unless otherwise specified, the amino acid sequence of the heavy chain containing the Fc region is represented herein as lacking a C-terminal glycine-lysine dipeptide. In one aspect, a heavy chain comprising the Fc region as specified herein is included in an antibody according to the invention, the heavy chain comprising additional C-terminal glycine-lysine dipeptides (G446 and K447, EU numbering system). In another aspect, a heavy chain comprising the Fc region as specified herein is included in an antibody according to the invention, the heavy chain comprising additional C-terminal glycine residues (G446, according to EU index number). Unless otherwise specified herein, the numbering of amino acid residues in the Fc region or constant region is according to the EU numbering system, also known as the EU index, as described in Kabat et al., Sequences of Proteins of Immunological Interest, 5th edition, Public Health Service, National Institutes of Health, Bethesda, MD, 1991.

[0049] "Frame" or "FR" refers to the variable domain residues other than the complementarity-determining region (CDR). A variable domain FR typically consists of four FR domains: FR1, FR2, FR3, and FR4. Therefore, the CDR and FR sequences usually appear in the VH (or VL) as follows: FR1-CDR-H1(CDR-L1)-FR2-CDR-H2(CDR-L2)-FR3-CDR-H3(CDR-L3)-FR4.

[0050] The terms “full-length antibody,” “intact antibody,” and “all antibody” are used interchangeably herein to refer to antibodies having a structure substantially similar to that of natural antibodies or having a heavy chain containing an Fc region as defined herein.

[0051] The terms “host cell,” “host cell line,” and “host cell culture” are used interchangeably and refer to cells in which exogenous nucleic acids have been introduced, including progeny cells. Host cells include “transformations” and “transformed cells,” which include primary transformed cells and progeny derived from those primary transformed cells, regardless of passage number. Progeny cells may not have completely identical nucleic acid contents to the parent cells and may contain mutations. This article includes mutant progeny with the same function or biological activity as those screened or selected from the original transformed cells.

[0052] A "human antibody" is an antibody whose amino acid sequence corresponds to that of an antibody produced by a human or human cell, or to a non-human antibody derived from a complete library of human antibodies or other antibody-encoding sequences. This definition of a human antibody specifically excludes humanized antibodies containing non-human antigen-binding residues.

[0053] The "human common framework" is a framework that represents the most frequently occurring amino acid residues in the selection of the human immunoglobulin VL or VH framework sequence. Generally, the selection of the human immunoglobulin VL or VH sequence is derived from a subgroup of variable domain sequences. Generally, the sequence subgroups are those listed in Kabat et al., Sequences of Proteins of Immunological Interest, 5th Edition, NIH Publication 91-3242, Bethesda MD (1991), Volumes 1-3. In one aspect, for VL, this subgroup is subgroup κ I, as listed above by Kabat et al. In another aspect, for VH, this subgroup is subgroup III, as listed above by Kabat et al.

[0054] As used in this article, the term "hypervariant region" or "HVR" refers to the regions in the antibody variable domain that are hypervariable in sequence and determine antigen binding specificity, such as the "complementarity-determining region" ("CDR").

[0055] Typically, an antibody contains six CDRs; three in the VH region (CDR-H1, CDR-H2, CDR-H3) and three in the VL region (CDR-L1, CDR-L2, CDR-L3). Exemplary CDRs in this document include:

[0056] a) Hyperchromatic rings present at amino acid residues 26-32 (L1), 50-52 (L2), 91-96 (L3), 26-32 (H1), 53-55 (H2), and 96-101 (H3) (Chothia and Lesk, J. Mol. Biol. 196:901-917(1987));

[0057] b) CDRs located at amino acid residues 24-34 (L1), 50-56 (L2), 89-97 (L3), 31-35b (H1), 50-65 (H2), and 95-102 (H3) (Kabat et al., Sequences of Proteins of Immunological Interest, 5th ed., Public Health Service, National Institutes of Health, Bethesda, MD (1991)); and

[0058] c) Antigen contact sites located at amino acid residues 27c-36 (L1), 46-55 (L2), 89-96 (L3), 30-35b (H1), 47-58 (H2), and 93-101 (H3) (MacCallum et al., J. Mol. Biol. 262: 732-745 (1996)).

[0059] Unless otherwise stated, the CDR is determined according to the method described by Kabat et al. (ibid.). Those skilled in the art will understand that the CDR name can also be determined according to the methods described by Chothia (ibid.), McCallum (ibid.), or any other scientifically accepted naming system.

[0060] The “individual” or “subject” is a mammal. Mammals include, but are not limited to, domesticated animals (such as cows, sheep, cats, dogs, and horses), primates (such as humans and non-human primates, such as monkeys), rabbits, and rodents (such as mice and rats). In some respects, the individual or subject is a human.

[0061] "Isolated" antibodies are antibodies that have been separated from components of their natural environment. In some respects, antibodies are purified to a purity greater than 95% or 99%, as determined by methods such as electrophoresis (e.g., SDS-PAGE, isoelectric focusing (IEF), capillary electrophoresis) or chromatography (e.g., ion exchange or reversed-phase HPLC). For a review of methods for assessing antibody purity, see, for example, Flatman et al., J. Chromatogr. B 848:79-87 (2007).

[0062] The term "nucleic acid molecule" or "polynucleotide" includes any compound and / or substance comprising a nucleotide polymer. Each nucleotide consists of a base, specifically a purine or pyrimidine base (i.e., cytosine (C), guanine (G), adenine (A), thymine (T), or uracil (U)), a sugar (i.e., deoxyribose or ribose), and a phosphate ester group. Typically, nucleic acid molecules are described by a base sequence, where the bases represent the primary structure (linear structure) of the nucleic acid molecule. Base sequences are typically represented from 5' to 3'. In this document, the term nucleic acid molecule encompasses deoxyribonucleic acid (DNA) (including, for example, complementary DNA (cDNA) and genomic DNA), ribonucleic acid (RNA) (particularly messenger RNA (mRNA)), synthetic forms of DNA or RNA, and mixed polymers comprising two or more of these molecules. Nucleic acid molecules can be linear or circular. Furthermore, the term nucleic acid molecule includes both sense and antisense strands, as well as single-stranded and double-stranded forms. Furthermore, the nucleic acid molecules described herein may contain naturally occurring or non-naturally occurring nucleotides. Examples of non-naturally occurring nucleotides include modified nucleotide bases having derivatized sugar or phosphate backbone bonds or chemically modified residues. Nucleic acid molecules also encompass DNA and RNA molecules suitable as vectors for direct in vitro and / or in vivo (e.g., in a host or patient) expression of antibodies used in this invention. Such DNA (e.g., cDNA) or RNA (e.g., mRNA) vectors may be unmodified or modified. For example, mRNA may be chemically modified to enhance the stability of the RNA vector and / or the expression of the encoding molecule, enabling the mRNA to be injected into a subject to generate antibodies in vivo (see, for example, Stadler et al., Nature Medicine 2017, published online June 12, 2017, doi:10.1038 / nm.4356 or EP 2 101 823 B1).

[0063] "Isolated" nucleic acids refer to nucleic acid molecules that have been separated from components of their natural environment. Isolated nucleic acids include nucleic acid molecules that are contained in cells that normally contain nucleic acid molecules, but which are located outside the chromosome or at a chromosomal location different from their natural chromosomal location.

[0064] "Isolated nucleic acid encoding anti-human C3bBb antibody" refers to one or more nucleic acid molecules encoding the heavy and light chains (or fragments thereof) of anti-human C3bBb antibody, including such nucleic acid molecules in a single vector or in separate vectors, and such nucleic acid molecules present at one or more locations in the host cell.

[0065] As used herein, the term "monoclonal antibody" refers to an antibody obtained from a substantially homogeneous group of antibodies, meaning that, apart from possible variant antibodies (e.g., those containing naturally occurring mutations or produced during the production of a monoclonal antibody formulation, such variants are typically present in small quantities), the individual antibodies constituting this group are identical and / or bind to the same epitopes. In contrast to polyclonal antibody formulations, which typically comprise different antibodies targeting different determinants (epitaxes), each monoclonal antibody in a monoclonal antibody formulation targets a single determinant on the antigen. Therefore, the modifier "monoclonal" indicates that the antibody is characterized by being obtained from a substantially homogeneous group of antibodies and should not be construed as requiring the antibody to be produced by any particular method. For example, monoclonal antibodies according to the invention can be prepared by a variety of techniques, including but not limited to hybridoma methods, recombinant DNA methods, phage display methods, and methods utilizing transgenic animals containing all or part of the human immunoglobulin loci, such methods and other exemplary methods for preparing monoclonal antibodies are described herein.

[0066] The term "packaging insert" is used to refer to the instruction leaflet typically included in the commercial packaging of a therapeutic product, which contains information concerning the indications, usage, dosage, administration, combination therapy, contraindications, and / or warnings related to the use of such therapeutic products.

[0067] The "percentage of amino acid sequence identity (%)" relative to a reference polypeptide sequence is defined as the percentage of amino acid residues in the candidate sequence that are identical to those in the reference polypeptide sequence after aligning the candidate sequence with the reference polypeptide sequence and introducing vacancies (if necessary) to achieve the maximum percentage of sequence identity, and for alignment purposes without considering any conserved substitutions as part of sequence identity. Alignment used to determine the percentage of amino acid sequence identity can be performed in various ways within the scope of the art, such as using publicly available computer software, such as BLAST, BLAST-2, Clustal W, Megalign (DNASTAR) software, or the FASTA package. Those skilled in the art can determine appropriate parameters for aligning sequences, including any algorithms required to achieve maximum alignment across the full length of the sequences being compared. Alternatively, the sequence comparison computer program ALIGN-2 can be used to generate the percentage of identity values. The ALIGN-2 sequence comparison computer program was written by Genentech, and the source code has been submitted with the user documentation to the US Copyright Office, Washington DC, 20559, registered under US Copyright Registry No. TXU510087 and described in WO 2001 / 007611.

[0068] Unless otherwise indicated, for the purposes of this article, the ggsearch program of FASTA package version 36.3.8c or later was used to generate the percentage of amino acid sequence identity using the BLOSUM50 comparison matrix. The FASTA package was created by WR Pearson and DJ Lipman (1988), “Improved Tools for Biological Sequence Analysis”, PNAS 85:2444-2448; WR Pearson (1996) “Effective proteins sequence comparison” Meth. Enzymol. 266:227-258; and Pearson et al. (1997) Genomics 46:24-36 and is publicly available at www.fasta.bioch.virginia.edu / fasta_www2 / fasta_down.shtml or www.ebi.ac.uk / Tools / sss / fasta. Alternatively, sequences can be compared using a public server accessible at fasta.bioch.virginia.edu / fasta_www2 / index.cgi, using the ggsearch (global protein: protein) program with default options (BLOSUM50; open: -10; ext: -2; Ktup=2) to ensure a global rather than local alignment. The percentage of amino acid identity is given in the output alignment header.

[0069] The terms “pharmaceutical composition” or “pharmaceutical formulation” refer to a formulation in which the active ingredient contained therein is in a biologically effective form and does not contain any additional components that would have unacceptable toxicity to a subject to whom the pharmaceutical composition will be administered.

[0070] "Pharmaceutically acceptable carriers" refer to components in a pharmaceutical composition or formulation other than the active ingredient that are non-toxic to the subject. Pharmaceutically acceptable carriers include, but are not limited to, buffer solutions, excipients, stabilizers, or preservatives.

[0071] As used herein, “treatment” (and its grammatical variations such as treat or treating) refers to an attempt to alter the natural course of a disease in the treated individual and can be performed for prevention or may be performed during a clinicopathological process. The desired effects of treatment include, but are not limited to, preventing the onset or recurrence of disease, alleviating symptoms, attenuating any direct or indirect pathological consequences of the disease, preventing metastasis, slowing the rate of disease progression, improving or mitigating the disease state, and alleviating or improving prognosis. In some aspects, the antibodies of this invention are used to delay the development of disease or slow its progression.

[0072] The term "variable region" or "variable domain" refers to a domain of the antibody heavy or light chain involved in antibody-antigen binding. The variable domains (VH and VL, respectively) of the heavy and light chains of natural antibodies generally have similar structures, with each domain containing four conserved frame regions (FRs) and three complementarity-determining regions (CDRs). (See, for example, Kindt et al., Kuby Immunology, 6th ed., WH Freeman and Co., p. 91 (2007)). A single VH or VL domain may be sufficient to confer antigen-binding specificity. Furthermore, antibodies binding to a specific antigen can be isolated using either the VH or VL domain from the antibody binding to that antigen to screen libraries of complementary VL or VH domains. See, for example, Portolano et al., J. Immunol. 150:880-887 (1993); Clarkson et al., Nature 352:624-628 (1991).

[0073] As used herein, the term "vector" refers to a nucleic acid molecule capable of carrying another nucleic acid linked to it. This term includes vectors that function as self-replicating nucleic acid structures, as well as vectors incorporated into the genome of a host cell into which they have been introduced. Some vectors are capable of directing the expression of the nucleic acid to which they are operatively linked. Such vectors are referred to herein as "expression vectors."

[0074] 2. Detailed embodiments of the present invention

[0075] In one aspect, the present invention is based in part on providing antibodies that specifically bind to human C3bBb, obtained when screened using a stable C3bBb complex. In other aspects, antibodies that bind to human C3bBb are provided. The antibodies of the present invention can be used, for example, to diagnose, prevent, or treat diseases such as eye diseases, such as age-related macular degeneration.

[0076] A. Exemplary anti-C3bBb antibody

[0077] In one aspect, the present invention provides an antibody that specifically binds to human C3bBb. In another aspect, the present invention provides an antibody that specifically binds to human C3bBb, which comprises a Bb subunit of factor B (FB) having a D279G mutation. In one embodiment, the antibody binds to human C3bBb, which comprises a Bb subunit of factor B (FB) having the amino acid sequence SEQ ID NO: 492.

[0078] In another aspect, the present invention provides an antibody that specifically binds to human C3bBb, which comprises a Bb subunit of factor B (FB) having mutations of D279G, K350N, and M458I. In one embodiment, the antibody binds to human C3bBb, which comprises a Bb subunit of factor B (FB) having the amino acid sequence of SEQ ID NO: 493.

[0079] In another aspect, the present invention provides an antibody that binds to human C3bBb, wherein the antibody:

[0080] - Combined with wild-type C3bBb, and / or

[0081] - Binds to recombinant human C3bBb, which contains the Bb subunit of the recombinant factor B (FB) protein, which contains the D279G mutation and optionally the mutations K350N and / or M458I; and / or

[0082] - Suppress bypass pathways; and / or

[0083] - An agonist or antagonist of C3bBb activity; and / or

[0084] -Specifically binds to cynomolgus monkey C3bBb and human C3bBb, and / or

[0085] -Specifically binds to African green monkey C3bBb and human C3bBb, and / or

[0086] -Binding with an affinity of ≤ 10 nM at 37°C, as measured by SPR.

[0087] The antibodies of this invention can be provided by screening using stable C3bBb, which is provided using recombinant FB protein having D279G and optionally further mutants K350N and / or M458I. Wild-type C3bBb is a rapidly decaying molecule and needs to be stable for antibody screening purposes. The inventors of this invention have been able to increase the half-life of the C3bBb complex from about 5 minutes to 11 hours, which allows for antibody generation using stable complexes in phage library panning.

[0088] Therefore, the antibody of the present invention specifically binds to stable C3bBb, particularly to C3bBb comprising a Bb subunit of factor B (FB) having a D279G mutation and optionally a mutation K350N and / or M458I. In one embodiment, the antibody of the present invention specifically binds to stable C3bBb comprising a Bb subunit of factor B (FB) having mutations of D279G, K350N, and M458I (preferably FB having the amino acid sequence of SEQ ID NO: 493).

[0089] Therefore, one aspect of the present invention is an antibody that binds to human C3bBb, which comprises a Bb subunit of recombinant factor B (FB) (preferably having the amino acid sequence of SEQ ID NO: 493) having mutations of D279G, K350N and M458I, which inhibits the bypass pathway.

[0090] Furthermore, the antibody of the present invention specifically binds to wild-type human C3bBb (Example 14) and is able to inhibit the bypass pathway (Example 9).

[0091] The antibodies of this invention bind to an epitope on human C3bBb, which comprises amino acid residues of both the C3b and Bb subunits of human C3bBb. This can be achieved by screening antibodies using stable C3bBb as a target antigen, for example, by the methods described herein.

[0092] The epitopes of the antibodies of the present invention can be identified by methods known in the art, such as cryo-electron microscopy. The P1AF8499 of the present invention (#20, see Examples 2 and 4) was analyzed by cryo-electron microscopy of the antibody-C3bBb trimer complex (Example 4). As shown in Figure 1, the epitopes are conformational epitopes containing amino acid residues of the C3b subunit and the Bb subunit of human C3bBb, which results in specific binding of the antibody to C3bBb, while no associated binding activity is shown for its subunits C3b and Bb. Cryo-electron microscopy was used to identify the amino acids bound by the antibodies of the present invention (Table E5 of Example 4).

[0093] In one aspect of the invention, the antibody binds to an epitope containing the following amino acid residues on wild-type human C3bBb, as detected by cryo-electron microscopy (preferably using the method according to Example 4):

[0094] -C3b subunits: Arg444, Lys534, Gly539, Ser540, Val524, Lys544, Gly546, Gln547, Ser548, Arg551, Gln557, Gln558, Thr560, Lys562, Glu564, Glu758, Pro759, Lys761, Asn762, Ile764, Leu768, Asn770, Asp797, and

[0095] -Bb subunits: Tyr465, Lys473, Ile474, Ser475, Ile477, Gly482, His483, Lys513, Val514, Ser515, Lys520, Arg521, Asp522, Glu610, and Lys613.

[0096] In one embodiment, the antibody binds to the same or overlapping epitopes as an antibody having the VH domain of SEQ ID NO: 39 and the VL domain of SEQ ID NO: 30.

[0097] The screening of antibodies binding to human C3bBb was accomplished using phage library panning and further engineering of directly generated antibodies, resulting in a variety of antibodies identified as binding to stable C3bBb. The amino acid sequences of the VH and VL domains of the antibodies are shown in Table D1.

[0098] Table D1: Amino acid sequence numbers of the variable heavy chain domain and variable light chain domain of the anti-C3bBb antibody

[0099]

[0100] Antibodies derived from antibody screening

[0101] Each of the 217 antibodies disclosed herein is disclosed as an invention, defined by six CDRs derived from the amino acid sequences of its VH domain and VL domain as shown in Table D1. Alternatively, the antibodies of the present invention are characterized in that the VH domain has the VH amino acid sequence shown in Table D1 and the VL domain has the VL amino acid sequence shown in Table D1.

[0102] One aspect of the invention is an antibody that specifically binds to human C3bBb, the antibody comprising a set of six CDRs of any antibody in Table D1. In one embodiment, the antibody comprises a variable light chain amino acid sequence and a variable heavy chain amino acid sequence of any antibody in Table D1.

[0103] In one aspect, the present invention relates to antibodies selected from antibodies #1 to #217 as shown in the first column of Table D1, wherein the antibody comprises a set of six CDRs derived from the amino acid sequences of their respective VH domain amino acid sequences and their respective VL amino acid sequences as shown in Table D1. In one embodiment, the antibody comprises the variable light chain amino acid sequence and the variable heavy chain amino acid sequence of any of the antibodies in Table D1.

[0104] In one aspect, the present invention relates to antibodies selected from antibodies #1 to #122 as shown in the first column of Table D1, wherein the antibody comprises a set of six CDRs derived from the amino acid sequences of their respective VH domain amino acid sequences and their respective VL amino acid sequences as shown in Table D1. In one embodiment, the antibody comprises the variable light chain amino acid sequence and the variable heavy chain amino acid sequence of any of the antibodies in Table D1.

[0105] In one aspect, the present invention relates to antibodies selected from antibodies #1 to #7 and #22 to #217 as shown in the first column of Table D1, wherein the antibody comprises a set of six CDRs derived from the amino acid sequences of their respective VH domain amino acid sequences and their respective VL amino acid sequences as shown in Table D1. In one embodiment, the antibody comprises the variable light chain amino acid sequence and the variable heavy chain amino acid sequence of any of the antibodies in Table D1.

[0106] In one aspect, the present invention relates to antibodies selected from antibodies #1 to #7 and #22 to #122 as shown in the first column of Table D1, wherein the antibody comprises a set of six CDRs derived from the amino acid sequences of their respective VH domain amino acid sequences and their respective VL amino acid sequences as shown in Table D1. In one embodiment, the antibody comprises a variable light chain amino acid sequence and a variable heavy chain amino acid sequence of any of the antibodies in Table D1.

[0107] In one aspect, the present invention relates to antibodies selected from antibodies #1 to #7 as shown in the first column of Table D1, wherein the antibody comprises a set of six CDRs derived from the amino acid sequences of their respective VH domain amino acid sequences and their respective VL amino acid sequences as shown in Table D1. In one embodiment, the antibody comprises a variable light chain amino acid sequence and a variable heavy chain amino acid sequence of any of the antibodies in Table D1.

[0108] In one aspect, the present invention relates to antibodies derived from antibody #22 as shown in the first column of Table D1.

[0109] Another aspect of the invention is an antibody that specifically binds to human C3bBb, the antibody comprising a set of VH and VL domains of any of the antibodies in Table D1.

[0110] In one embodiment, the antibody comprises a set of six CDRs of any antibody in Table E4. In another embodiment, the antibody comprises a set of VH and VL domains of any antibody in Table E4.

[0111] Single antibody

[0112] Antibody P1AG9426

[0113] In another aspect, the present invention provides an antibody comprising: (a) CDR-H1, comprising the amino acid sequence of SEQ ID NO:449; (b) CDR-H2, comprising the amino acid sequence of SEQ ID NO:450; (c) CDR-H3, comprising the amino acid sequence of SEQ ID NO:451; (d) CDR-L1, comprising the amino acid sequence of SEQ ID NO:452; (e) CDR-L2, comprising the amino acid sequence of SEQ ID NO:453; and (f) CDR-L3, comprising the amino acid sequence of SEQ ID NO:454 (corresponding to antibody #1, P1AG9426).

[0114] In another embodiment, the anti-C3bBb antibody comprises one or more CDR sequences of VH of SEQ ID NO:1. In another embodiment, the anti-C3bBb antibody comprises one or more CDR sequences of VL of SEQ ID NO:2. In yet another embodiment, the anti-C3bBb antibody comprises the CDR sequence of VH of SEQ ID NO:1 and the CDR sequence of VL of SEQ ID NO:2.

[0115] On the other hand, the anti-C3bBb antibody comprises the amino acid sequences of CDR-H1, CDR-H2 and CDR-H3 of the VH domain of SEQ ID NO:1 and the amino acid sequences of CDR-L1, CDR-L2 and CDR-L3 of the VL domain of SEQ ID NO:2.

[0116] In one aspect, the anti-C3bBb antibody comprises one or more heavy chain CDR amino acid sequences of the VH domain of SEQ ID NO:1 and a frame amino acid sequence having at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, and 99% sequence identity with the frame amino acid sequence of the VH domain of SEQ ID NO:1. In another aspect, the anti-C3bBb antibody comprises three heavy chain CDR amino acid sequences of the VH domain of SEQ ID NO:1 and a frame amino acid sequence having at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, and 99% sequence identity with the frame amino acid sequence of the VH domain of SEQ ID NO:1. On one hand, the anti-C3bBb antibody comprises the three heavy chain CDR amino acid sequences of the VH domain of SEQ ID NO:1 and a frame with at least 95% sequence identity to the frame amino acid sequence of the VH domain of SEQ ID NO:1. On the other hand, the anti-C3bBb antibody comprises the three heavy chain CDR amino acid sequences of the VH domain of SEQ ID NO:1 and a frame with at least 98% sequence identity to the frame amino acid sequence of the VH domain of SEQ ID NO:1.

[0117] In one aspect, the anti-C3bBb antibody comprises one or more light chain CDR amino acid sequences of the VL domain of SEQ ID NO:2 and a frame amino acid sequence having at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, and 99% sequence identity with the frame amino acid sequence of the VL domain of SEQ ID NO:2. In another aspect, the anti-C3bBb antibody comprises three light chain CDR amino acid sequences of the VL domain of SEQ ID NO:2 and a frame amino acid sequence having at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, and 99% sequence identity with the frame amino acid sequence of the VL domain of SEQ ID NO:2. On one hand, the anti-C3bBb antibody comprises the three light chain CDR amino acid sequences of the VL domain of SEQ ID NO:2 and a framework amino acid sequence having at least 95% sequence identity with the VL domain of SEQ ID NO:2. On the other hand, the anti-C3bBb antibody comprises the three light chain CDR amino acid sequences of the VL domain of SEQ ID NO:2 and a framework amino acid sequence having at least (particularly at least) 98% sequence identity with the VH domain of SEQ ID NO:2.

[0118] In one aspect, the anti-C3bBb antibody comprises: (a) CDR-H1 comprising the amino acid sequence of SEQ ID NO:449; (b) CDR-H2 comprising the amino acid sequence of SEQ ID NO:450; (c) CDR-H3 comprising the amino acid sequence of SEQ ID NO:451; (d) CDR-L1 comprising the amino acid sequence of SEQ ID NO:452; (e) CDR-L2 comprising the amino acid sequence of SEQ ID NO:453; and (f) CDR-L3 comprising the amino acid sequence of SEQ ID NO:454; and a VH domain having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity with the amino acid sequence of SEQ ID NO:1 and a VH domain having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity with the amino acid sequence of SEQ ID NO:2. The amino acid sequence of the VL domain has at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity. In one aspect, the VH domain has at least 95% sequence identity with the amino acid sequence of SEQ ID NO:1. In another aspect, the VL domain has at least 95% sequence identity with the amino acid sequence of SEQ ID NO:2.

[0119] In one aspect, the anti-C3bBb antibody comprises: (a) a CDR-H1 comprising the amino acid sequence of SEQ ID NO:449; (b) a CDR-H2 comprising the amino acid sequence of SEQ ID NO:450; (c) a CDR-H3 comprising the amino acid sequence of SEQ ID NO:451; (d) a CDR-L1 comprising the amino acid sequence of SEQ ID NO:452; (e) a CDR-L2 comprising the amino acid sequence of SEQ ID NO:453; and (f) a CDR-L3 comprising the amino acid sequence of SEQ ID NO:454; and a VH domain having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity with the amino acid sequence of SEQ ID NO:1; and a VH domain having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity with the amino acid sequence of SEQ ID NO:2 ... The antibody has a VL domain with at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity with the amino acid sequence of SEQ ID NO:1; wherein the antibody specifically binds to human C3bBb. In one aspect, the VH domain has at least 95% sequence identity with the amino acid sequence of SEQ ID NO:1. In another aspect, the VL domain has at least 95% sequence identity with the amino acid sequence of SEQ ID NO:2. In one aspect, the dissociation constant (KD) of the antibody binding to C3bBb is reduced by up to 10-fold or increased by up to 10-fold compared to the dissociation constant (KD) of the antibody comprising the VH sequence of SEQ ID NO:1 and the VL sequence of SEQ ID NO:2.

[0120] In another aspect, an anti-C3bBb antibody comprises a heavy chain variable domain (VH) sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity with the amino acid sequence of SEQ ID NO:1. In another aspect, the anti-C3bBb antibody comprises a heavy chain variable domain (VH) sequence having at least 95% sequence identity with the amino acid sequence of SEQ ID NO:2. In some aspects, the VH sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity contains substitutions (e.g., conserved substitutions), insertions, or deletions relative to a reference sequence, but the anti-C3bBb antibody containing this sequence retains its ability to bind to C3bBb. In some aspects, in SEQ ID NO:1, a total of 1 to 10 amino acids have been substituted, inserted, and / or deleted. In some respects, substitution, insertion, or deletion occurs in regions outside the CDR (i.e., in the FR). Optionally, the anti-C3bBb antibody comprises the VH sequence shown in SEQ ID NO:1, including post-translational modifications of that sequence. In one specific aspect, the VH comprises one, two, or three CDRs selected from: (a) CDR-H1 comprising the amino acid sequence of SEQ ID NO:449, (b) CDR-H2 comprising the amino acid sequence of SEQ ID NO:450, and (c) CDR-H3 comprising the amino acid sequence of SEQ ID NO:451. In another aspect, an anti-C3bBb antibody is provided, wherein the antibody comprises a light chain variable domain (VL) sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity with the amino acid sequence of SEQ ID NO:2. In one aspect, the anti-C3bBb antibody comprises a light chain variable domain (VL) sequence having at least 95% sequence identity with the amino acid sequence of SEQ ID NO:2. In some aspects, the VL sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity contains substitutions (e.g., conserved substitutions), insertions, or deletions relative to the reference sequence, but the anti-C3bBb antibody containing this sequence retains its ability to bind to C3bBb. In some aspects, in SEQ ID NO:2, a total of 1 to 10 amino acids have been substituted, inserted, and / or deleted. In some aspects, the substitution, insertion, or deletion occurs in a region outside the CDR (i.e., in the FR).Optionally, the anti-C3bBb antibody comprises the VL sequence shown in SEQ ID NO:2, including post-translational modifications of that sequence. In one specific aspect, the VL comprises one, two, or three CDRs selected from the following: (a) CDR-L1 comprising the amino acid sequence of SEQ ID NO:452, (b) CDR-L2 comprising the amino acid sequence of SEQ ID NO:453, and (c) CDR-L3 comprising the amino acid sequence of SEQ ID NO:454.

[0121] In another aspect, an anti-C3bBb antibody is provided, wherein the antibody comprises a VH sequence as provided in any of the aspects above and a VL sequence as provided in any of the aspects above. In one aspect, the antibody comprises the VH sequence and the VL sequence as in SEQ ID NO:1 and SEQ ID NO:2, respectively, including post-translational modifications of those sequences.

[0122] On the other hand, an anti-C3bBb antibody is provided, wherein the antibody comprises the heavy chain amino acid sequence of SEQ ID NO:435 and the light chain amino acid sequence of SEQ ID NO:436.

[0123] In one aspect, an antibody is provided comprising the VH domain as described above and the heavy chain constant domain amino acid sequence as included in SEQ ID NO:435, and the VL domain comprising the light chain constant domain amino acid sequence as described above and as included in SEQ ID NO:436.

[0124] Antibody P1AG9376

[0125] In another aspect, the present invention provides an antibody comprising: (a) CDR-H1, comprising the amino acid sequence of SEQ ID NO:455; (b) CDR-H2, comprising the amino acid sequence of SEQ ID NO:456; (c) CDR-H3, comprising the amino acid sequence of SEQ ID NO:457; (d) CDR-L1, comprising the amino acid sequence of SEQ ID NO:458; (e) CDR-L2, comprising the amino acid sequence of SEQ ID NO:459; and (f) CDR-L3, comprising the amino acid sequence of SEQ ID NO:460 (corresponding to antibody #2, P1AG9376).

[0126] In another embodiment, the anti-C3bBb antibody comprises one or more CDR sequences of VH of SEQ ID NO:3. In another embodiment, the anti-C3bBb antibody comprises one or more CDR sequences of VL of SEQ ID NO:4. In yet another embodiment, the anti-C3bBb antibody comprises the CDR sequence of VH of SEQ ID NO:3 and the CDR sequence of VL of SEQ ID NO:4.

[0127] On the other hand, the anti-C3bBb antibody comprises the amino acid sequences of CDR-H1, CDR-H2 and CDR-H3 of the VH domain of SEQ ID NO:3 and the amino acid sequences of CDR-L1, CDR-L2 and CDR-L3 of the VL domain of SEQ ID NO:4.

[0128] In one aspect, the anti-C3bBb antibody comprises one or more heavy chain CDR amino acid sequences of the VH domain of SEQ ID NO:3 and a frame amino acid sequence having at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, and 99% sequence identity with the frame amino acid sequence of the VH domain of SEQ ID NO:3. In another aspect, the anti-C3bBb antibody comprises three heavy chain CDR amino acid sequences of the VH domain of SEQ ID NO:3 and a frame amino acid sequence having at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, and 99% sequence identity with the frame amino acid sequence of the VH domain of SEQ ID NO:3. On one hand, the anti-C3bBb antibody comprises the three heavy chain CDR amino acid sequences of the VH domain of SEQ ID NO:3 and a frame with at least 95% sequence identity to the frame amino acid sequence of the VH domain of SEQ ID NO:3. On the other hand, the anti-C3bBb antibody comprises the three heavy chain CDR amino acid sequences of the VH domain of SEQ ID NO:3 and a frame with at least 98% sequence identity to the frame amino acid sequence of the VH domain of SEQ ID NO:3.

[0129] In one aspect, the anti-C3bBb antibody comprises one or more light chain CDR amino acid sequences of the VL domain of SEQ ID NO:4 and a frame amino acid sequence having at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, and 99% sequence identity with the frame amino acid sequence of the VL domain of SEQ ID NO:4. In another aspect, the anti-C3bBb antibody comprises three light chain CDR amino acid sequences of the VL domain of SEQ ID NO:4 and a frame amino acid sequence having at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, and 99% sequence identity with the frame amino acid sequence of the VL domain of SEQ ID NO:4. On one hand, the anti-C3bBb antibody comprises the three light chain CDR amino acid sequences of the VL domain of SEQ ID NO:4 and a framework amino acid sequence having at least 95% sequence identity with the VL domain of SEQ ID NO:4. On the other hand, the anti-C3bBb antibody comprises the three light chain CDR amino acid sequences of the VL domain of SEQ ID NO:4 and a framework amino acid sequence having at least (particularly at least) 98% sequence identity with the VH domain of SEQ ID NO:4.

[0130] In one aspect, the anti-C3bBb antibody comprises: (a) a CDR-H1 comprising the amino acid sequence of SEQ ID NO:455; (b) a CDR-H2 comprising the amino acid sequence of SEQ ID NO:456; (c) a CDR-H3 comprising the amino acid sequence of SEQ ID NO:457; (d) a CDR-L1 comprising the amino acid sequence of SEQ ID NO:458; (e) a CDR-L2 comprising the amino acid sequence of SEQ ID NO:459; and (f) a CDR-L3 comprising the amino acid sequence of SEQ ID NO:460; and a VH domain having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity with the amino acid sequence of SEQ ID NO:455 and a VH domain having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity with the amino acid sequence of SEQ ID NO:455. The amino acid sequence of the VL domain has at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity. In one aspect, the VH domain has at least 95% sequence identity with the amino acid sequence of SEQ ID NO:3. In another aspect, the VL domain has at least 95% sequence identity with the amino acid sequence of SEQ ID NO:4.

[0131] In one aspect, the anti-C3bBb antibody comprises: (a) a CDR-H1 comprising the amino acid sequence of SEQ ID NO:455; (b) a CDR-H2 comprising the amino acid sequence of SEQ ID NO:456; (c) a CDR-H3 comprising the amino acid sequence of SEQ ID NO:457; (d) a CDR-L1 comprising the amino acid sequence of SEQ ID NO:458; (e) a CDR-L2 comprising the amino acid sequence of SEQ ID NO:459; and (f) a CDR-L3 comprising the amino acid sequence of SEQ ID NO:460; and a VH domain having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity with the amino acid sequence of SEQ ID NO:45 ... The antibody has a VL domain with at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity with the amino acid sequence of SEQ ID NO:3; wherein the antibody specifically binds to human C3bBb. In one aspect, the VH domain has at least 95% sequence identity with the amino acid sequence of SEQ ID NO:4. In one aspect, the VL domain has at least 95% sequence identity with the amino acid sequence of SEQ ID NO:4. In one aspect, the dissociation constant (KD) of the antibody binding to C3bBb is reduced by up to 10-fold or increased by up to 10-fold compared to the dissociation constant (KD) of the antibody comprising the VH sequence of SEQ ID NO:3 and the VL sequence of SEQ ID NO:4.

[0132] In another aspect, an anti-C3bBb antibody comprises a heavy chain variable domain (VH) sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity with the amino acid sequence of SEQ ID NO:3. In another aspect, the anti-C3bBb antibody comprises a heavy chain variable domain (VH) sequence having at least 95% sequence identity with the amino acid sequence of SEQ ID NO:4. In some aspects, the VH sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity contains substitutions (e.g., conserved substitutions), insertions, or deletions relative to a reference sequence, but the anti-C3bBb antibody containing this sequence retains its ability to bind to C3bBb. In some aspects, in SEQ ID NO:3, a total of 1 to 10 amino acids have been substituted, inserted, and / or deleted. In some respects, substitution, insertion, or deletion occurs in regions outside the CDR (i.e., in the FR). Optionally, the anti-C3bBb antibody comprises the VH sequence shown in SEQ ID NO:3, including post-translational modifications of that sequence. In one specific aspect, the VH comprises one, two, or three CDRs selected from: (a) CDR-H1 comprising the amino acid sequence of SEQ ID NO:455, (b) CDR-H2 comprising the amino acid sequence of SEQ ID NO:456, and (c) CDR-H3 comprising the amino acid sequence of SEQ ID NO:457. In another aspect, an anti-C3bBb antibody is provided, wherein the antibody comprises a light chain variable domain (VL) sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity with the amino acid sequence of SEQ ID NO:4. In one aspect, the anti-C3bBb antibody comprises a light chain variable domain (VL) sequence having at least 95% sequence identity with the amino acid sequence of SEQ ID NO:4. In some aspects, the VL sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity contains substitutions (e.g., conserved substitutions), insertions, or deletions relative to the reference sequence, but the anti-C3bBb antibody containing this sequence retains its ability to bind to C3bBb. In some aspects, in SEQ ID NO:4, a total of 1 to 10 amino acids have been substituted, inserted, and / or deleted. In some aspects, the substitution, insertion, or deletion occurs in a region outside the CDR (i.e., in the FR).Optionally, the anti-C3bBb antibody comprises the VL sequence shown in SEQ ID NO:4, including post-translational modifications of that sequence. In one specific aspect, the VL comprises one, two, or three CDRs selected from the following: (a) CDR-L1 comprising the amino acid sequence of SEQ ID NO:458, (b) CDR-L2 comprising the amino acid sequence of SEQ ID NO:459, and (c) CDR-L3 comprising the amino acid sequence of SEQ ID NO:460.

[0133] In another aspect, an anti-C3bBb antibody is provided, wherein the antibody comprises a VH sequence as provided in any of the aspects above and a VL sequence as provided in any of the aspects above. In one aspect, the antibody comprises the VH sequence and the VL sequence as in SEQ ID NO:3 and SEQ ID NO:4, respectively, including post-translational modifications of those sequences.

[0134] On the other hand, an anti-C3bBb antibody is provided, wherein the antibody comprises the heavy chain amino acid sequence of SEQ ID NO:437 and the light chain amino acid sequence of SEQ ID NO:438.

[0135] In one aspect, an antibody is provided comprising the VH domain as described above and the heavy chain constant domain amino acid sequence as included in SEQ ID NO:437, and the VL domain comprising the light chain constant domain amino acid sequence as described above and as included in SEQ ID NO:438.

[0136] Antibody P1AG9372

[0137] In another aspect, the present invention provides an antibody comprising: (a) CDR-H1, comprising the amino acid sequence of SEQ ID NO:461; (b) CDR-H2, comprising the amino acid sequence of SEQ ID NO:462; (c) CDR-H3, comprising the amino acid sequence of SEQ ID NO:463; (d) CDR-L1, comprising the amino acid sequence of SEQ ID NO:464; (e) CDR-L2, comprising the amino acid sequence of SEQ ID NO:465; and (f) CDR-L3, comprising the amino acid sequence of SEQ ID NO:466 (corresponding to antibody #3, P1AG9372).

[0138] In another embodiment, the anti-C3bBb antibody comprises one or more CDR sequences of VH of SEQ ID NO:5. In another embodiment, the anti-C3bBb antibody comprises one or more CDR sequences of VL of SEQ ID NO:6. In yet another embodiment, the anti-C3bBb antibody comprises the CDR sequence of VH of SEQ ID NO:5 and the CDR sequence of VL of SEQ ID NO:6.

[0139] On the other hand, the anti-C3bBb antibody comprises the amino acid sequences of CDR-H1, CDR-H2 and CDR-H3 of the VH domain of SEQ ID NO:5 and the amino acid sequences of CDR-L1, CDR-L2 and CDR-L3 of the VL domain of SEQ ID NO:6.

[0140] In one aspect, the anti-C3bBb antibody comprises one or more heavy chain CDR amino acid sequences of the VH domain of SEQ ID NO:5 and a frame amino acid sequence having at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, and 99% sequence identity with the frame amino acid sequence of the VH domain of SEQ ID NO:5. In another aspect, the anti-C3bBb antibody comprises three heavy chain CDR amino acid sequences of the VH domain of SEQ ID NO:5 and a frame amino acid sequence having at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, and 99% sequence identity with the frame amino acid sequence of the VH domain of SEQ ID NO:5. On one hand, the anti-C3bBb antibody comprises the three heavy chain CDR amino acid sequences of the VH domain of SEQ ID NO:5 and a frame with at least 95% sequence identity to the frame amino acid sequence of the VH domain of SEQ ID NO:5. On the other hand, the anti-C3bBb antibody comprises the three heavy chain CDR amino acid sequences of the VH domain of SEQ ID NO:5 and a frame with at least 98% sequence identity to the frame amino acid sequence of the VH domain of SEQ ID NO:5.

[0141] In one aspect, the anti-C3bBb antibody comprises one or more light chain CDR amino acid sequences of the VL domain of SEQ ID NO:6 and a frame amino acid sequence having at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, and 99% sequence identity with the frame amino acid sequence of the VL domain of SEQ ID NO:6. In another aspect, the anti-C3bBb antibody comprises three light chain CDR amino acid sequences of the VL domain of SEQ ID NO:6 and a frame amino acid sequence having at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, and 99% sequence identity with the frame amino acid sequence of the VL domain of SEQ ID NO:6. On one hand, the anti-C3bBb antibody comprises the three light chain CDR amino acid sequences of the VL domain of SEQ ID NO:6 and a framework amino acid sequence having at least 95% sequence identity with the VL domain of SEQ ID NO:6. On the other hand, the anti-C3bBb antibody comprises the three light chain CDR amino acid sequences of the VL domain of SEQ ID NO:6 and a framework amino acid sequence having at least (particularly at least) 98% sequence identity with the VH domain of SEQ ID NO:6.

[0142] In one aspect, the anti-C3bBb antibody comprises: (a) a CDR-H1 comprising the amino acid sequence of SEQ ID NO:461; (b) a CDR-H2 comprising the amino acid sequence of SEQ ID NO:462; (c) a CDR-H3 comprising the amino acid sequence of SEQ ID NO:463; (d) a CDR-L1 comprising the amino acid sequence of SEQ ID NO:464; (e) a CDR-L2 comprising the amino acid sequence of SEQ ID NO:465; and (f) a CDR-L3 comprising the amino acid sequence of SEQ ID NO:466; and a VH domain having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity with the amino acid sequence of SEQ ID NO:5 and a VH domain having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity with the amino acid sequence of SEQ ID NO:6. The amino acid sequence of the VL domain has at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity. In one aspect, the VH domain has at least 95% sequence identity with the amino acid sequence of SEQ ID NO:5. In another aspect, the VL domain has at least 95% sequence identity with the amino acid sequence of SEQ ID NO:6.

[0143] In one aspect, the anti-C3bBb antibody comprises: (a) a CDR-H1 comprising the amino acid sequence of SEQ ID NO:461; (b) a CDR-H2 comprising the amino acid sequence of SEQ ID NO:462; (c) a CDR-H3 comprising the amino acid sequence of SEQ ID NO:463; (d) a CDR-L1 comprising the amino acid sequence of SEQ ID NO:464; (e) a CDR-L2 comprising the amino acid sequence of SEQ ID NO:465; and (f) a CDR-L3 comprising the amino acid sequence of SEQ ID NO:466; and a VH domain having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity with the amino acid sequence of SEQ ID NO:5; and a VH domain having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity with the amino acid sequence of SEQ ID NO:6 ... The antibody has a VL domain with at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity with the amino acid sequence of SEQ ID NO:5; wherein the antibody specifically binds to human C3bBb. In one aspect, the VH domain has at least 95% sequence identity with the amino acid sequence of SEQ ID NO:5. In another aspect, the VL domain has at least 95% sequence identity with the amino acid sequence of SEQ ID NO:6. In one aspect, the dissociation constant (KD) of the antibody binding to C3bBb is reduced by up to 10-fold or increased by up to 10-fold compared to the dissociation constant (KD) of the antibody comprising the VH sequence of SEQ ID NO:5 and the VL sequence of SEQ ID NO:6.

[0144] In another aspect, an anti-C3bBb antibody comprises a heavy chain variable domain (VH) sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity with the amino acid sequence of SEQ ID NO:5. In another aspect, the anti-C3bBb antibody comprises a heavy chain variable domain (VH) sequence having at least 95% sequence identity with the amino acid sequence of SEQ ID NO:6. In some aspects, the VH sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity contains substitutions (e.g., conserved substitutions), insertions, or deletions relative to a reference sequence, but the anti-C3bBb antibody containing this sequence retains its ability to bind to C3bBb. In some aspects, in SEQ ID NO:5, a total of 1 to 10 amino acids have been substituted, inserted, and / or deleted. In some respects, substitution, insertion, or deletion occurs in regions outside the CDR (i.e., in the FR). Optionally, the anti-C3bBb antibody comprises the VH sequence shown in SEQ ID NO:5, including post-translational modifications of that sequence. In one specific aspect, the VH comprises one, two, or three CDRs selected from: (a) CDR-H1 comprising the amino acid sequence of SEQ ID NO:461, (b) CDR-H2 comprising the amino acid sequence of SEQ ID NO:462, and (c) CDR-H3 comprising the amino acid sequence of SEQ ID NO:463. In another aspect, an anti-C3bBb antibody is provided, wherein the antibody comprises a light chain variable domain (VL) sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity with the amino acid sequence of SEQ ID NO:6. In one aspect, the anti-C3bBb antibody comprises a light chain variable domain (VL) sequence having at least 95% sequence identity with the amino acid sequence of SEQ ID NO:6. In some aspects, the VL sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity contains substitutions (e.g., conserved substitutions), insertions, or deletions relative to the reference sequence, but the anti-C3bBb antibody containing this sequence retains its ability to bind to C3bBb. In some aspects, in SEQ ID NO:6, a total of 1 to 10 amino acids have been substituted, inserted, and / or deleted. In some aspects, the substitutions, insertions, or deletions occur in regions outside the CDR (i.e., in the FR).Optionally, the anti-C3bBb antibody comprises the VL sequence shown in SEQ ID NO:6, including post-translational modifications of that sequence. In one specific aspect, the VL comprises one, two, or three CDRs selected from the following: (a) CDR-L1 comprising the amino acid sequence of SEQ ID NO:464, (b) CDR-L2 comprising the amino acid sequence of SEQ ID NO:465, and (c) CDR-L3 comprising the amino acid sequence of SEQ ID NO:466.

[0145] In another aspect, an anti-C3bBb antibody is provided, wherein the antibody comprises a VH sequence as provided in any of the aspects above and a VL sequence as provided in any of the aspects above. In one aspect, the antibody comprises the VH sequence and the VL sequence as described in SEQ ID NO:5 and SEQ ID NO:6, respectively, including post-translational modifications of those sequences.

[0146] On the other hand, an anti-C3bBb antibody is provided, wherein the antibody comprises the heavy chain amino acid sequence of SEQ ID NO:439 and the light chain amino acid sequence of SEQ ID NO:440.

[0147] In one aspect, an antibody is provided comprising the VH domain as described above and the heavy chain constant domain amino acid sequence as included in SEQ ID NO:439, and the VL domain comprising the light chain constant domain amino acid sequence as described above and as included in SEQ ID NO:440.

[0148] Antibody P1AG9420

[0149] In another aspect, the present invention provides an antibody comprising: (a) CDR-H1, comprising the amino acid sequence of SEQ ID NO:467; (b) CDR-H2, comprising the amino acid sequence of SEQ ID NO:468; (c) CDR-H3, comprising the amino acid sequence of SEQ ID NO:469; (d) CDR-L1, comprising the amino acid sequence of SEQ ID NO:470; (e) CDR-L2, comprising the amino acid sequence of SEQ ID NO:471; and (f) CDR-L3, comprising the amino acid sequence of SEQ ID NO:472 (corresponding to antibody #4, P1AG9420).

[0150] In another embodiment, the anti-C3bBb antibody comprises one or more CDR sequences of VH of SEQ ID NO:7. In another embodiment, the anti-C3bBb antibody comprises one or more CDR sequences of VL of SEQ ID NO:8. In yet another embodiment, the anti-C3bBb antibody comprises the CDR sequence of VH of SEQ ID NO:7 and the CDR sequence of VL of SEQ ID NO:8.

[0151] On the other hand, the anti-C3bBb antibody comprises the amino acid sequences of CDR-H1, CDR-H2 and CDR-H3 of the VH domain of SEQ ID NO:7 and the amino acid sequences of CDR-L1, CDR-L2 and CDR-L3 of the VL domain of SEQ ID NO:8.

[0152] In one aspect, the anti-C3bBb antibody comprises one or more heavy chain CDR amino acid sequences of the VH domain of SEQ ID NO:7 and a frame amino acid sequence having at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, and 99% sequence identity with the frame amino acid sequence of the VH domain of SEQ ID NO:7. In another aspect, the anti-C3bBb antibody comprises three heavy chain CDR amino acid sequences of the VH domain of SEQ ID NO:7 and a frame amino acid sequence having at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, and 99% sequence identity with the frame amino acid sequence of the VH domain of SEQ ID NO:7. On one hand, the anti-C3bBb antibody comprises the three heavy chain CDR amino acid sequences of the VH domain of SEQ ID NO:7 and a frame with at least 95% sequence identity to the frame amino acid sequence of the VH domain of SEQ ID NO:7. On the other hand, the anti-C3bBb antibody comprises the three heavy chain CDR amino acid sequences of the VH domain of SEQ ID NO:7 and a frame with at least 98% sequence identity to the frame amino acid sequence of the VH domain of SEQ ID NO:7.

[0153] In one aspect, the anti-C3bBb antibody comprises one or more light chain CDR amino acid sequences of the VL domain of SEQ ID NO:8 and a frame amino acid sequence having at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, and 99% sequence identity with the frame amino acid sequence of the VL domain of SEQ ID NO:8. In another aspect, the anti-C3bBb antibody comprises three light chain CDR amino acid sequences of the VL domain of SEQ ID NO:8 and a frame amino acid sequence having at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, and 99% sequence identity with the frame amino acid sequence of the VL domain of SEQ ID NO:8. On one hand, the anti-C3bBb antibody comprises the three light chain CDR amino acid sequences of the VL domain of SEQ ID NO:8 and a framework amino acid sequence having at least 95% sequence identity with the VL domain of SEQ ID NO:8. On the other hand, the anti-C3bBb antibody comprises the three light chain CDR amino acid sequences of the VL domain of SEQ ID NO:8 and a framework amino acid sequence having at least (particularly at least) 98% sequence identity with the VH domain of SEQ ID NO:8.

[0154] In one aspect, the anti-C3bBb antibody comprises: (a) a CDR-H1 comprising the amino acid sequence of SEQ ID NO:467; (b) a CDR-H2 comprising the amino acid sequence of SEQ ID NO:468; (c) a CDR-H3 comprising the amino acid sequence of SEQ ID NO:469; (d) a CDR-L1 comprising the amino acid sequence of SEQ ID NO:470; (e) a CDR-L2 comprising the amino acid sequence of SEQ ID NO:471; and (f) a CDR-L3 comprising the amino acid sequence of SEQ ID NO:472; and a VH domain having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity with the amino acid sequence of SEQ ID NO:7 and a VH domain having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity with the amino acid sequence of SEQ ID NO:8. The amino acid sequence of the VL domain has at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity. In one aspect, the VH domain has at least 95% sequence identity with the amino acid sequence of SEQ ID NO:7. In another aspect, the VL domain has at least 95% sequence identity with the amino acid sequence of SEQ ID NO:8.

[0155] In one aspect, the anti-C3bBb antibody comprises: (a) a CDR-H1 comprising the amino acid sequence of SEQ ID NO:467; (b) a CDR-H2 comprising the amino acid sequence of SEQ ID NO:468; (c) a CDR-H3 comprising the amino acid sequence of SEQ ID NO:469; (d) a CDR-L1 comprising the amino acid sequence of SEQ ID NO:470; (e) a CDR-L2 comprising the amino acid sequence of SEQ ID NO:471; and (f) a CDR-L3 comprising the amino acid sequence of SEQ ID NO:472; and a VH domain having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity with the amino acid sequence of SEQ ID NO:7; and a VH domain having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity with the amino acid sequence of SEQ ID NO:8 ... The antibody has a VL domain with at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity with the amino acid sequence of SEQ ID NO:7; wherein the antibody specifically binds to human C3bBb. In one aspect, the VH domain has at least 95% sequence identity with the amino acid sequence of SEQ ID NO:7. In another aspect, the VL domain has at least 95% sequence identity with the amino acid sequence of SEQ ID NO:8. In one aspect, the dissociation constant (KD) of the antibody binding to C3bBb is reduced by up to 10-fold or increased by up to 10-fold compared to the dissociation constant (KD) of the antibody comprising the VH sequence of SEQ ID NO:7 and the VL sequence of SEQ ID NO:8.

[0156] In another aspect, an anti-C3bBb antibody comprises a heavy chain variable domain (VH) sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity with the amino acid sequence of SEQ ID NO:7. In another aspect, the anti-C3bBb antibody comprises a heavy chain variable domain (VH) sequence having at least 95% sequence identity with the amino acid sequence of SEQ ID NO:8. In some aspects, the VH sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity contains substitutions (e.g., conserved substitutions), insertions, or deletions relative to a reference sequence, but the anti-C3bBb antibody containing this sequence retains its ability to bind to C3bBb. In some aspects, in SEQ ID NO:7, a total of 1 to 10 amino acids have been substituted, inserted, and / or deleted. In some respects, substitution, insertion, or deletion occurs in regions outside the CDR (i.e., in the FR). Optionally, the anti-C3bBb antibody comprises the VH sequence shown in SEQ ID NO:7, including post-translational modifications of that sequence. In one specific aspect, the VH comprises one, two, or three CDRs selected from: (a) CDR-H1 comprising the amino acid sequence of SEQ ID NO:467, (b) CDR-H2 comprising the amino acid sequence of SEQ ID NO:468, and (c) CDR-H3 comprising the amino acid sequence of SEQ ID NO:469. In another aspect, an anti-C3bBb antibody is provided, wherein the antibody comprises a light chain variable domain (VL) sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity with the amino acid sequence of SEQ ID NO:8. In one aspect, the anti-C3bBb antibody comprises a light chain variable domain (VL) sequence having at least 95% sequence identity with the amino acid sequence of SEQ ID NO:8. In some aspects, the VL sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity contains substitutions (e.g., conserved substitutions), insertions, or deletions relative to the reference sequence, but the anti-C3bBb antibody containing this sequence retains its ability to bind to C3bBb. In some aspects, in SEQ ID NO:8, a total of 1 to 10 amino acids have been substituted, inserted, and / or deleted. In some aspects, the substitution, insertion, or deletion occurs in regions outside the CDR (i.e., in the FR).Optionally, the anti-C3bBb antibody comprises the VL sequence shown in SEQ ID NO:8, including post-translational modifications of that sequence. In one specific aspect, the VL comprises one, two, or three CDRs selected from the following: (a) CDR-L1 comprising the amino acid sequence of SEQ ID NO:470, (b) CDR-L2 comprising the amino acid sequence of SEQ ID NO:471, and (c) CDR-L3 comprising the amino acid sequence of SEQ ID NO:472.

[0157] In another aspect, an anti-C3bBb antibody is provided, wherein the antibody comprises a VH sequence as provided in any of the aspects above and a VL sequence as provided in any of the aspects above. In one aspect, the antibody comprises the VH sequence and the VL sequence as in SEQ ID NO:7 and SEQ ID NO:8, respectively, including post-translational modifications of those sequences.

[0158] On the other hand, an anti-C3bBb antibody is provided, wherein the antibody comprises the heavy chain amino acid sequence of SEQ ID NO:441 and the light chain amino acid sequence of SEQ ID NO:442.

[0159] In one aspect, an antibody is provided comprising the VH domain as described above and the heavy chain constant domain amino acid sequence as included in SEQ ID NO:441, and the VL domain comprising the light chain constant domain amino acid sequence as described above and as included in SEQ ID NO:442.

[0160] Antibody P1AG9391

[0161] In another aspect, the present invention provides an antibody comprising: (a) CDR-H1, comprising the amino acid sequence of SEQ ID NO:473; (b) CDR-H2, comprising the amino acid sequence of SEQ ID NO:474; (c) CDR-H3, comprising the amino acid sequence of SEQ ID NO:475; (d) CDR-L1, comprising the amino acid sequence of SEQ ID NO:476; (e) CDR-L2, comprising the amino acid sequence of SEQ ID NO:477; and (f) CDR-L3, comprising the amino acid sequence of SEQ ID NO:478 (corresponding to antibody #5, P1AG9391).

[0162] In another embodiment, the anti-C3bBb antibody comprises one or more CDR sequences of VH of SEQ ID NO:9. In another embodiment, the anti-C3bBb antibody comprises one or more CDR sequences of VL of SEQ ID NO:10. In yet another embodiment, the anti-C3bBb antibody comprises the CDR sequence of VH of SEQ ID NO:9 and the CDR sequence of VL of SEQ ID NO:10.

[0163] On the other hand, the anti-C3bBb antibody comprises the amino acid sequences of CDR-H1, CDR-H2 and CDR-H3 of the VH domain of SEQ ID NO:9 and the amino acid sequences of CDR-L1, CDR-L2 and CDR-L3 of the VL domain of SEQ ID NO:10.

[0164] In one aspect, the anti-C3bBb antibody comprises one or more heavy chain CDR amino acid sequences of the VH domain of SEQ ID NO:9 and a frame amino acid sequence having at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, and 99% sequence identity with the frame amino acid sequence of the VH domain of SEQ ID NO:9. In another aspect, the anti-C3bBb antibody comprises three heavy chain CDR amino acid sequences of the VH domain of SEQ ID NO:9 and a frame amino acid sequence having at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, and 99% sequence identity with the frame amino acid sequence of the VH domain of SEQ ID NO:9. On one hand, the anti-C3bBb antibody comprises the three heavy chain CDR amino acid sequences of the VH domain of SEQ ID NO:9 and a frame with at least 95% sequence identity to the frame amino acid sequence of the VH domain of SEQ ID NO:9. On the other hand, the anti-C3bBb antibody comprises the three heavy chain CDR amino acid sequences of the VH domain of SEQ ID NO:9 and a frame with at least 98% sequence identity to the frame amino acid sequence of the VH domain of SEQ ID NO:9.

[0165] In one aspect, the anti-C3bBb antibody comprises one or more light chain CDR amino acid sequences of the VL domain of SEQ ID NO:10 and a frame having at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, and 99% sequence identity with the frame amino acid sequence of the VL domain of SEQ ID NO:10. In another aspect, the anti-C3bBb antibody comprises three light chain CDR amino acid sequences of the VL domain of SEQ ID NO:10 and a frame having at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, and 99% sequence identity with the frame amino acid sequence of the VL domain of SEQ ID NO:10. On one hand, the anti-C3bBb antibody comprises the three light chain CDR amino acid sequences of the VL domain of SEQ ID NO:10 and a frame having at least 95% sequence identity with the frame amino acid sequence of the VL domain of SEQ ID NO:10. On the other hand, the anti-C3bBb antibody comprises the three light chain CDR amino acid sequences of the VL domain of SEQ ID NO:10 and a frame having at least (particularly at least) 98% sequence identity with the frame amino acid sequence of the VH domain of SEQ ID NO:10.

[0166] In one aspect, the anti-C3bBb antibody comprises: (a) a CDR-H1 comprising the amino acid sequence of SEQ ID NO:473; (b) a CDR-H2 comprising the amino acid sequence of SEQ ID NO:474; (c) a CDR-H3 comprising the amino acid sequence of SEQ ID NO:475; (d) a CDR-L1 comprising the amino acid sequence of SEQ ID NO:476; (e) a CDR-L2 comprising the amino acid sequence of SEQ ID NO:477; and (f) a CDR-L3 comprising the amino acid sequence of SEQ ID NO:478; and a VH domain having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity with the amino acid sequence of SEQ ID NO:9 and a VH domain having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity with the amino acid sequence of SEQ ID NO:10. The amino acid sequence of the VL domain has at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity. In one aspect, the VH domain has at least 95% sequence identity with the amino acid sequence of SEQ ID NO:9. In another aspect, the VL domain has at least 95% sequence identity with the amino acid sequence of SEQ ID NO:10.

[0167] In one aspect, the anti-C3bBb antibody comprises: (a) a CDR-H1 comprising the amino acid sequence of SEQ ID NO:473; (b) a CDR-H2 comprising the amino acid sequence of SEQ ID NO:474; (c) a CDR-H3 comprising the amino acid sequence of SEQ ID NO:475; (d) a CDR-L1 comprising the amino acid sequence of SEQ ID NO:476; (e) a CDR-L2 comprising the amino acid sequence of SEQ ID NO:477; and (f) a CDR-L3 comprising the amino acid sequence of SEQ ID NO:478; and a VH domain having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity with the amino acid sequence of SEQ ID NO:9; and a VH domain having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity with the amino acid sequence of SEQ ID NO:10 ... The antibody has a VL domain with at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity with the amino acid sequence of SEQ ID NO:9; wherein the antibody specifically binds to human C3bBb. In one aspect, the VH domain has at least 95% sequence identity with the amino acid sequence of SEQ ID NO:9. In another aspect, the VL domain has at least 95% sequence identity with the amino acid sequence of SEQ ID NO:10. In one aspect, the dissociation constant (KD) of the antibody binding to C3bBb is reduced by up to 10-fold or increased by up to 10-fold compared to the dissociation constant (KD) of the antibody comprising the VH sequence of SEQ ID NO:9 and the VL sequence of SEQ ID NO:10.

[0168] In another aspect, an anti-C3bBb antibody comprises a heavy chain variable domain (VH) sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity with the amino acid sequence of SEQ ID NO:9. In another aspect, the anti-C3bBb antibody comprises a heavy chain variable domain (VH) sequence having at least 95% sequence identity with the amino acid sequence of SEQ ID NO:10. In some aspects, the VH sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity contains substitutions (e.g., conserved substitutions), insertions, or deletions relative to a reference sequence, but the anti-C3bBb antibody containing this sequence retains its ability to bind to C3bBb. In some aspects, in SEQ ID NO:9, a total of 1 to 10 amino acids have been substituted, inserted, and / or deleted. In some respects, substitution, insertion, or deletion occurs in regions outside the CDR (i.e., in the FR). Optionally, the anti-C3bBb antibody comprises the VH sequence shown in SEQ ID NO:9, including post-translational modifications of that sequence. In one specific aspect, the VH comprises one, two, or three CDRs selected from: (a) CDR-H1 comprising the amino acid sequence of SEQ ID NO:473, (b) CDR-H2 comprising the amino acid sequence of SEQ ID NO:474, and (c) CDR-H3 comprising the amino acid sequence of SEQ ID NO:475. In another aspect, an anti-C3bBb antibody is provided, wherein the antibody comprises a light chain variable domain (VL) sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity with the amino acid sequence of SEQ ID NO:10. In one aspect, the anti-C3bBb antibody comprises a light chain variable domain (VL) sequence having at least 95% sequence identity with the amino acid sequence of SEQ ID NO:10. In some aspects, the VL sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity contains substitutions (e.g., conserved substitutions), insertions, or deletions relative to the reference sequence, but the anti-C3bBb antibody containing this sequence retains its ability to bind to C3bBb. In some aspects, in SEQ ID NO:10, a total of 1 to 10 amino acids have been substituted, inserted, and / or deleted. In some aspects, the substitution, insertion, or deletion occurs in a region outside the CDR (i.e., in the FR).Optionally, the anti-C3bBb antibody comprises the VL sequence shown in SEQ ID NO:10, including post-translational modifications of that sequence. In one specific aspect, the VL comprises one, two, or three CDRs selected from the following: (a) CDR-L1 comprising the amino acid sequence of SEQ ID NO:476, (b) CDR-L2 comprising the amino acid sequence of SEQ ID NO:477, and (c) CDR-L3 comprising the amino acid sequence of SEQ ID NO:478.

[0169] In another aspect, an anti-C3bBb antibody is provided, wherein the antibody comprises a VH sequence as provided in any of the aspects above and a VL sequence as provided in any of the aspects above. In one aspect, the antibody comprises the VH sequence and VL sequence as in SEQ ID NO:9 and SEQ ID NO:10, respectively, including post-translational modifications of those sequences.

[0170] On the other hand, an anti-C3bBb antibody is provided, wherein the antibody comprises the heavy chain amino acid sequence of SEQ ID NO:443 and the light chain amino acid sequence of SEQ ID NO:444.

[0171] In one aspect, an antibody is provided comprising the VH domain as described above and the heavy chain constant domain amino acid sequence as included in SEQ ID NO:443, and the VL domain comprising the light chain constant domain amino acid sequence as described above and as included in SEQ ID NO:444.

[0172] Antibody P1AH1205

[0173] In another aspect, the present invention provides an antibody comprising: (a) CDR-H1, comprising the amino acid sequence of SEQ ID NO:479; (b) CDR-H2, comprising the amino acid sequence of SEQ ID NO:480; (c) CDR-H3, comprising the amino acid sequence of SEQ ID NO:481; (d) CDR-L1, comprising the amino acid sequence of SEQ ID NO:482; (e) CDR-L2, comprising the amino acid sequence of SEQ ID NO:483; and (f) CDR-L3, comprising the amino acid sequence of SEQ ID NO:484 (corresponding to antibody #6, P1AH1205).

[0174] In another embodiment, the anti-C3bBb antibody comprises one or more CDR sequences of VH of SEQ ID NO:11. In another embodiment, the anti-C3bBb antibody comprises one or more CDR sequences of VL of SEQ ID NO:12. In yet another embodiment, the anti-C3bBb antibody comprises the CDR sequence of VH of SEQ ID NO:11 and the CDR sequence of VL of SEQ ID NO:12.

[0175] On the other hand, the anti-C3bBb antibody comprises the amino acid sequences of CDR-H1, CDR-H2 and CDR-H3 of the VH domain of SEQ ID NO:11 and the amino acid sequences of CDR-L1, CDR-L2 and CDR-L3 of the VL domain of SEQ ID NO:12.

[0176] In one aspect, the anti-C3bBb antibody comprises one or more heavy chain CDR amino acid sequences of the VH domain of SEQ ID NO:11 and a frame having at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, and 99% sequence identity with the frame amino acid sequence of the VH domain of SEQ ID NO:11. In another aspect, the anti-C3bBb antibody comprises three heavy chain CDR amino acid sequences of the VH domain of SEQ ID NO:11 and a frame having at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, and 99% sequence identity with the frame amino acid sequence of the VH domain of SEQ ID NO:11. On one hand, the anti-C3bBb antibody comprises the three heavy chain CDR amino acid sequences of the VH domain of SEQ ID NO:11 and a frame with at least 95% sequence identity to the frame amino acid sequence of the VH domain of SEQ ID NO:11. On the other hand, the anti-C3bBb antibody comprises the three heavy chain CDR amino acid sequences of the VH domain of SEQ ID NO:11 and a frame with at least 98% sequence identity to the frame amino acid sequence of the VH domain of SEQ ID NO:11.

[0177] In one aspect, the anti-C3bBb antibody comprises one or more light chain CDR amino acid sequences of the VL domain of SEQ ID NO:12 and a frame having at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, and 99% sequence identity with the frame amino acid sequence of the VL domain of SEQ ID NO:12. In another aspect, the anti-C3bBb antibody comprises three light chain CDR amino acid sequences of the VL domain of SEQ ID NO:12 and a frame having at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, and 99% sequence identity with the frame amino acid sequence of the VL domain of SEQ ID NO:12. On one hand, the anti-C3bBb antibody comprises the three light chain CDR amino acid sequences of the VL domain of SEQ ID NO:12 and a frame having at least 95% sequence identity with the frame amino acid sequence of the VL domain of SEQ ID NO:12. On the other hand, the anti-C3bBb antibody comprises the three light chain CDR amino acid sequences of the VL domain of SEQ ID NO:12 and a frame having at least (particularly at least) 98% sequence identity with the frame amino acid sequence of the VH domain of SEQ ID NO:12.

[0178] In one aspect, the anti-C3bBb antibody comprises: (a) a CDR-H1 comprising the amino acid sequence of SEQ ID NO:479; (b) a CDR-H2 comprising the amino acid sequence of SEQ ID NO:480; (c) a CDR-H3 comprising the amino acid sequence of SEQ ID NO:481; (d) a CDR-L1 comprising the amino acid sequence of SEQ ID NO:482; (e) a CDR-L2 comprising the amino acid sequence of SEQ ID NO:483; and (f) a CDR-L3 comprising the amino acid sequence of SEQ ID NO:484; and a VH domain having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity with the amino acid sequence of SEQ ID NO:11 and a VH domain having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity with the amino acid sequence of SEQ ID NO:12. The amino acid sequence of the VL domain has at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity. In one aspect, the VH domain has at least 95% sequence identity with the amino acid sequence of SEQ ID NO:11. In another aspect, the VL domain has at least 95% sequence identity with the amino acid sequence of SEQ ID NO:12.

[0179] In one aspect, the anti-C3bBb antibody comprises: (a) a CDR-H1 comprising the amino acid sequence of SEQ ID NO:479; (b) a CDR-H2 comprising the amino acid sequence of SEQ ID NO:480; (c) a CDR-H3 comprising the amino acid sequence of SEQ ID NO:481; (d) a CDR-L1 comprising the amino acid sequence of SEQ ID NO:482; (e) a CDR-L2 comprising the amino acid sequence of SEQ ID NO:483; and (f) a CDR-L3 comprising the amino acid sequence of SEQ ID NO:484; and a VH domain having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity with the amino acid sequence of SEQ ID NO:11; and a VH domain having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity with the amino acid sequence of SEQ ID NO:12 ... The antibody has a VL domain with at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity with the amino acid sequence of SEQ ID NO:11; wherein the antibody specifically binds to human C3bBb. In one aspect, the VH domain has at least 95% sequence identity with the amino acid sequence of SEQ ID NO:11. In another aspect, the VL domain has at least 95% sequence identity with the amino acid sequence of SEQ ID NO:12. In one aspect, the dissociation constant (KD) of the antibody binding to C3bBb is reduced by up to 10-fold or increased by up to 10-fold compared to the dissociation constant (KD) of the antibody comprising the VH sequence of SEQ ID NO:11 and the VL sequence of SEQ ID NO:12.

[0180] In another aspect, an anti-C3bBb antibody comprises a heavy chain variable domain (VH) sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity with the amino acid sequence of SEQ ID NO:11. In another aspect, the anti-C3bBb antibody comprises a heavy chain variable domain (VH) sequence having at least 95% sequence identity with the amino acid sequence of SEQ ID NO:12. In some aspects, the VH sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity contains substitutions (e.g., conserved substitutions), insertions, or deletions relative to a reference sequence, but the anti-C3bBb antibody containing this sequence retains its ability to bind to C3bBb. In some aspects, in SEQ ID NO:11, a total of 1 to 10 amino acids have been substituted, inserted, and / or deleted. In some respects, substitution, insertion, or deletion occurs in regions outside the CDR (i.e., in the FR). Optionally, the anti-C3bBb antibody comprises the VH sequence shown in SEQ ID NO:11, including post-translational modifications of that sequence. In one specific aspect, the VH comprises one, two, or three CDRs selected from: (a) CDR-H1 comprising the amino acid sequence of SEQ ID NO:479, (b) CDR-H2 comprising the amino acid sequence of SEQ ID NO:480, and (c) CDR-H3 comprising the amino acid sequence of SEQ ID NO:481. In another aspect, an anti-C3bBb antibody is provided, wherein the antibody comprises a light chain variable domain (VL) sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity with the amino acid sequence of SEQ ID NO:12. In one aspect, the anti-C3bBb antibody comprises a light chain variable domain (VL) sequence having at least 95% sequence identity with the amino acid sequence of SEQ ID NO:12. In some aspects, the VL sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity contains substitutions (e.g., conserved substitutions), insertions, or deletions relative to the reference sequence, but the anti-C3bBb antibody containing this sequence retains its ability to bind to C3bBb. In some aspects, in SEQ ID NO:12, a total of 1 to 10 amino acids have been substituted, inserted, and / or deleted. In some aspects, the substitution, insertion, or deletion occurs in regions outside the CDR (i.e., in the FR).Optionally, the anti-C3bBb antibody comprises the VL sequence shown in SEQ ID NO:12, including post-translational modifications of that sequence. In one specific aspect, the VL comprises one, two, or three CDRs selected from the following: (a) CDR-L1 comprising the amino acid sequence of SEQ ID NO:482, (b) CDR-L2 comprising the amino acid sequence of SEQ ID NO:483, and (c) CDR-L3 comprising the amino acid sequence of SEQ ID NO:484.

[0181] In another aspect, an anti-C3bBb antibody is provided, wherein the antibody comprises a VH sequence as provided in any of the aspects above and a VL sequence as provided in any of the aspects above. In one aspect, the antibody comprises the VH sequence and VL sequence as in SEQ ID NO:11 and SEQ ID NO:12, respectively, including post-translational modifications of those sequences.

[0182] On the other hand, an anti-C3bBb antibody is provided, wherein the antibody comprises the heavy chain amino acid sequence of SEQ ID NO:445 and the light chain amino acid sequence of SEQ ID NO:446.

[0183] In one aspect, an antibody is provided comprising the VH domain as described above and the heavy chain constant domain amino acid sequence as included in SEQ ID NO:445, and the VL domain comprising the light chain constant domain amino acid sequence as described above and as included in SEQ ID NO:446.

[0184] Antibody P1AH1199

[0185] In another aspect, the present invention provides an antibody comprising: (a) CDR-H1, comprising the amino acid sequence of SEQ ID NO:485; (b) CDR-H2, comprising the amino acid sequence of SEQ ID NO:486; (c) CDR-H3, comprising the amino acid sequence of SEQ ID NO:487; (d) CDR-L1, comprising the amino acid sequence of SEQ ID NO:488; (e) CDR-L2, comprising the amino acid sequence of SEQ ID NO:489; and (f) CDR-L3, comprising the amino acid sequence of SEQ ID NO:490 (corresponding to antibody #7, P1AH1199).

[0186] In another embodiment, the anti-C3bBb antibody comprises one or more CDR sequences of VH of SEQ ID NO:13. In another embodiment, the anti-C3bBb antibody comprises one or more CDR sequences of VL of SEQ ID NO:14. In yet another embodiment, the anti-C3bBb antibody comprises the CDR sequence of VH of SEQ ID NO:13 and the CDR sequence of VL of SEQ ID NO:14.

[0187] On the other hand, the anti-C3bBb antibody comprises the amino acid sequences of CDR-H1, CDR-H2 and CDR-H3 of the VH domain of SEQ ID NO:13 and the amino acid sequences of CDR-L1, CDR-L2 and CDR-L3 of the VL domain of SEQ ID NO:14.

[0188] In one aspect, the anti-C3bBb antibody comprises one or more heavy chain CDR amino acid sequences of the VH domain of SEQ ID NO:13 and a frame amino acid sequence having at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, and 99% sequence identity with the frame amino acid sequence of the VH domain of SEQ ID NO:13. In another aspect, the anti-C3bBb antibody comprises three heavy chain CDR amino acid sequences of the VH domain of SEQ ID NO:13 and a frame amino acid sequence having at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, and 99% sequence identity with the frame amino acid sequence of the VH domain of SEQ ID NO:13. On one hand, the anti-C3bBb antibody comprises the three heavy chain CDR amino acid sequences of the VH domain of SEQ ID NO:13 and a frame with at least 95% sequence identity to the frame amino acid sequence of the VH domain of SEQ ID NO:13. On the other hand, the anti-C3bBb antibody comprises the three heavy chain CDR amino acid sequences of the VH domain of SEQ ID NO:13 and a frame with at least 98% sequence identity to the frame amino acid sequence of the VH domain of SEQ ID NO:13.

[0189] In one aspect, the anti-C3bBb antibody comprises one or more light chain CDR amino acid sequences of the VL domain of SEQ ID NO:14 and a frame having at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, and 99% sequence identity with the frame amino acid sequence of the VL domain of SEQ ID NO:14. In another aspect, the anti-C3bBb antibody comprises three light chain CDR amino acid sequences of the VL domain of SEQ ID NO:14 and a frame having at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, and 99% sequence identity with the frame amino acid sequence of the VL domain of SEQ ID NO:14. On one hand, the anti-C3bBb antibody comprises the three light chain CDR amino acid sequences of the VL domain of SEQ ID NO:14 and a frame having at least 95% sequence identity with the frame amino acid sequence of the VL domain of SEQ ID NO:14. On the other hand, the anti-C3bBb antibody comprises the three light chain CDR amino acid sequences of the VL domain of SEQ ID NO:14 and a frame having at least (particularly at least) 98% sequence identity with the frame amino acid sequence of the VH domain of SEQ ID NO:14.

[0190] In one aspect, the anti-C3bBb antibody comprises: (a) a CDR-H1 comprising the amino acid sequence of SEQ ID NO:485; (b) a CDR-H2 comprising the amino acid sequence of SEQ ID NO:486; (c) a CDR-H3 comprising the amino acid sequence of SEQ ID NO:487; (d) a CDR-L1 comprising the amino acid sequence of SEQ ID NO:488; (e) a CDR-L2 comprising the amino acid sequence of SEQ ID NO:489; and (f) a CDR-L3 comprising the amino acid sequence of SEQ ID NO:490; and a VH domain having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity with the amino acid sequence of SEQ ID NO:13 and a VH domain having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity with the amino acid sequence of SEQ ID NO:14. The amino acid sequence of the VL domain has at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity. In one aspect, the VH domain has at least 95% sequence identity with the amino acid sequence of SEQ ID NO:13. In another aspect, the VL domain has at least 95% sequence identity with the amino acid sequence of SEQ ID NO:14.

[0191] In one aspect, the anti-C3bBb antibody comprises: (a) a CDR-H1 comprising the amino acid sequence of SEQ ID NO:485; (b) a CDR-H2 comprising the amino acid sequence of SEQ ID NO:486; (c) a CDR-H3 comprising the amino acid sequence of SEQ ID NO:487; (d) a CDR-L1 comprising the amino acid sequence of SEQ ID NO:488; (e) a CDR-L2 comprising the amino acid sequence of SEQ ID NO:489; and (f) a CDR-L3 comprising the amino acid sequence of SEQ ID NO:490; and a VH domain having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity with the amino acid sequence of SEQ ID NO:13; and a VH domain having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity with the amino acid sequence of SEQ ID NO:14 ... The antibody has a VL domain with at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity with the amino acid sequence of SEQ ID NO:13; wherein the antibody specifically binds to human C3bBb. In one aspect, the VH domain has at least 95% sequence identity with the amino acid sequence of SEQ ID NO:13. In another aspect, the VL domain has at least 95% sequence identity with the amino acid sequence of SEQ ID NO:14. In one aspect, the dissociation constant (KD) of the antibody binding to C3bBb is reduced by up to 10-fold or increased by up to 10-fold compared to the dissociation constant (KD) of the antibody comprising the VH sequence of SEQ ID NO:13 and the VL sequence of SEQ ID NO:14.

[0192] In another aspect, an anti-C3bBb antibody comprises a heavy chain variable domain (VH) sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity with the amino acid sequence of SEQ ID NO:13. In another aspect, the anti-C3bBb antibody comprises a heavy chain variable domain (VH) sequence having at least 95% sequence identity with the amino acid sequence of SEQ ID NO:14. In some aspects, the VH sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity contains substitutions (e.g., conserved substitutions), insertions, or deletions relative to a reference sequence, but the anti-C3bBb antibody containing this sequence retains its ability to bind to C3bBb. In some aspects, in SEQ ID NO:13, a total of 1 to 10 amino acids have been substituted, inserted, and / or deleted. In some respects, substitution, insertion, or deletion occurs in regions outside the CDR (i.e., in the FR). Optionally, the anti-C3bBb antibody comprises the VH sequence shown in SEQ ID NO:13, including post-translational modifications of that sequence. In one specific aspect, the VH comprises one, two, or three CDRs selected from: (a) CDR-H1 comprising the amino acid sequence of SEQ ID NO:485, (b) CDR-H2 comprising the amino acid sequence of SEQ ID NO:486, and (c) CDR-H3 comprising the amino acid sequence of SEQ ID NO:487. In another aspect, an anti-C3bBb antibody is provided, wherein the antibody comprises a light chain variable domain (VL) sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity with the amino acid sequence of SEQ ID NO:14. In one aspect, the anti-C3bBb antibody comprises a light chain variable domain (VL) sequence having at least 95% sequence identity with the amino acid sequence of SEQ ID NO:14. In some aspects, the VL sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity contains substitutions (e.g., conserved substitutions), insertions, or deletions relative to the reference sequence, but the anti-C3bBb antibody containing this sequence retains its ability to bind to C3bBb. In some aspects, in SEQ ID NO:14, a total of 1 to 10 amino acids have been substituted, inserted, and / or deleted. In some aspects, the substitution, insertion, or deletion occurs in regions outside the CDR (i.e., in the FR).Optionally, the anti-C3bBb antibody comprises the VL sequence shown in SEQ ID NO:14, including post-translational modifications of that sequence. In one specific aspect, the VL comprises one, two, or three CDRs selected from the following: (a) CDR-L1 comprising the amino acid sequence of SEQ ID NO:488, (b) CDR-L2 comprising the amino acid sequence of SEQ ID NO:489, and (c) CDR-L3 comprising the amino acid sequence of SEQ ID NO:490.

[0193] In another aspect, an anti-C3bBb antibody is provided, wherein the antibody comprises a VH sequence as provided in any of the aspects above and a VL sequence as provided in any of the aspects above. In one aspect, the antibody comprises the VH sequence and VL sequence as in SEQ ID NO:13 and SEQ ID NO:14, respectively, including post-translational modifications of those sequences.

[0194] On the other hand, an anti-C3bBb antibody is provided, wherein the antibody comprises the heavy chain amino acid sequence of SEQ ID NO:447 and the light chain amino acid sequence of SEQ ID NO:448.

[0195] In one aspect, an antibody is provided comprising the VH domain as described above and the heavy chain constant domain amino acid sequence as included in SEQ ID NO:447, and the VL domain comprising the light chain constant domain amino acid sequence as described above and as included in SEQ ID NO:448.

[0196] Other aspects and embodiments

[0197] In another aspect of the invention, the anti-human C3bBb antibody according to any of the foregoing aspects is a monoclonal antibody, including chimeric, humanized, or human antibodies. In another aspect, the anti-human C3bBb antibody is an antibody fragment, such as Fv, Fab, Fab', scFv, biantibody, or F(ab')2 fragment.

[0198] On the other hand, the antibody is a full-length antibody, such as a full-length IgG1 antibody as defined herein or other antibody classes or isotypes. In one embodiment, the antibody is a full-length IgG1 antibody.

[0199] In one embodiment, the antibody is a monovalent antibody. In one embodiment, the antibody is a bivalent antibody. In one embodiment, the antibody is a trivalent antibody. In one embodiment, the antibody is a quadrivalent antibody.

[0200] On the other hand, the anti-human C3bBb antibody according to any of the foregoing aspects may incorporate, alone or in combination, any of the features described in sections 1 to 8 below:

[0201] 1. Antibody affinity

[0202] In some respects, the dissociation constant (KD) of the antibodies provided herein is ≤ 1 μM, ≤ 100 nM, ≤ 10 nM, ≤ 1 nM, ≤ 0.1 nM, ≤ 0.01 nM, or ≤ 0.001 nM. In some embodiments, the dissociation constant (KD) of the antibodies provided herein is 10. -6 M or smaller, in one embodiment 10 -7 M or smaller, in one embodiment 10 -8 M or smaller, in one embodiment 10 -8 M to 10 -13 M, in one embodiment, is 10 -9 M to 10 -13 M.

[0203] In one embodiment, as measured by SPR, the antibody binds to human C3bBb with an affinity of ≤ 300 pM at about 37°C. In one embodiment, as measured by SPR, the antibody binds to human C3bBb with an affinity of about 1 to 300 pM at about 37°C. In one embodiment, as measured by SPR, the antibody binds to human C3bBb with an affinity of about 1 to about 200 pM at about 37°C.

[0204] In one embodiment, as measured by SPR, the antibody binds to human C3bBb with an affinity of ≤100 pM at about 25°C. In one embodiment, as measured by SPR, the antibody binds to human C3bBb with an affinity of about 1 to 100 pM at about 25°C. In one embodiment, as measured by SPR, the antibody binds to human C3bBb with an affinity of ≤50 pM at about 25°C.

[0205] In one aspect, KD is measured using BIACORE® surface plasmon resonance measurement. In one embodiment, KD is measured as described in Example 5.

[0206] 2. Antibody fragments

[0207] In some respects, the antibodies presented in this article are antibody fragments.

[0208] On one hand, antibody fragments are Fab, Fab', Fab'-SH, or F(ab')2 fragments, particularly Fab fragments. Papain digestion of an intact antibody produces two identical antigen-binding fragments called "Fab" fragments. Each "Fab" fragment contains a heavy chain variable domain and a light chain variable domain (VH and VL, respectively), as well as a light chain constant domain (CL) and a heavy chain first constant domain (CH1). Therefore, the term "Fab fragment" refers to an antibody fragment comprising a light chain containing the VL and CL domains and a heavy chain containing the VH and CH1 domains. Fab' fragments differ from Fab fragments in that residues are added to the carboxyl terminus of the CH1 domain, including one or more cysteine ​​residues from the antibody hinge region. Fab'-SH is a Fab' fragment in which the cysteine ​​residues of the constant domain have free thiol groups. Pepsin treatment produces the F(ab')2 fragment, which has two antigen-binding sites (two Fab fragments) and a portion of the Fc region. For a discussion of Fab and F(ab')2 fragments containing salvage receptor-binding epitope residues and having an increased in vivo half-life, see U.S. Patent No. 5,869,046.

[0209] In another respect, antibody fragments are bisomatic, trisomatic, or tetrasomatic antibodies. A “bisomatic antibody” is an antibody fragment having two antigen-binding sites, which can be bivalent or bispecific. See, for example, EP 404,097; WO 1993 / 01161; Hudson et al., Nat. Med. 9:129-134 (2003); and Hollinger et al., Proc. Natl. Acad. Sci. USA 90: 6444-6448 (1993). Trisomatic and tetrasomatic antibodies are also described by Hudson et al. in Nat. Med. 9:129-134 (2003).

[0210] On the other hand, the antibody fragment is a single-chain Fab fragment. A “single-chain Fab fragment” or “scFab” is a polypeptide composed of an antibody heavy chain variable domain (VH), an antibody heavy chain constant domain 1 (CH1), an antibody light chain variable domain (VL), an antibody light chain constant domain (CL), and a linker, wherein the antibody domains and linker have one of the following sequences in the N-terminal to C-terminal direction: a) VH-CH1-linker-VL-CL, b) VL-CL-linker-VH-CH1, c) VH-CL-linker-VL-CH1, or d) VL-CH1-linker-VH-CL. Specifically, the linker is a polypeptide of at least 30 amino acids, preferably between 32 and 50 amino acids. The single-chain Fab fragment is stabilized via a native disulfide bond between the CL domain and the CH1 domain. Furthermore, these single-chain Fab fragments can be further stabilized by generating interchain disulfide bonds via the insertion of cysteine ​​residues (e.g., position 44 in the variable heavy chain and position 100 in the variable light chain according to Kabat numbering).

[0211] On the other hand, antibody fragments are single-chain variable fragments (scFv). A "single-chain variable fragment" or "scFv" is a fusion protein of the antibody's heavy chain variable domain (VH) and light chain variable domain (VL), linked by a linker. Specifically, the linker is a short polypeptide of approximately 10 to 25 amino acids, typically rich in glycine for flexibility and serine or threonine for solubility, and can link the N-terminus of the VH to the C-terminus of the VL, or vice versa. Despite the removal of the constant region and the introduction of the linker, the protein retains the specificity of the original antibody. For reviews of scFv fragments, see, for example, Plückthun, in The Pharmacology of Monoclonal Antibodies, Vol. 113, edited by Rosenburg and Moore (Springer-Verlag, New York), pp. 269–315 (1994); see also WO 93 / 16185; and U.S. Patent Nos. 5,571,894 and 5,587,458.

[0212] On the other hand, antibody fragments are single-domain antibodies. A "single-domain antibody" is an antibody fragment containing all or part of the variable heavy chain domain or all or part of the variable light chain domain of an antibody. In some respects, single-domain antibodies are human single-domain antibodies (Domantis, Inc., Waltham, MA; see, for example, U.S. Patent No. 6,248,516 B1).

[0213] Antibody fragments can be prepared using various techniques, including but not limited to the proteolytic digestion of intact antibodies and recombinant production from recombinant host cells (e.g., E. coli), as described herein.

[0214] 3. Chimeric and humanized antibodies

[0215] In some respects, the antibodies described herein are chimeric antibodies. Certain chimeric antibodies are described, for example, in U.S. Patent No. 4,816,567 and Morrison et al., Proc. Natl. Acad. Sci. USA, 81:6851-6855 (1984). In one instance, a chimeric antibody comprises a non-human variable region (e.g., a variable region derived from a mouse, rat, hamster, rabbit, or non-human primate, such as a monkey) and a human constant region. In another instance, a chimeric antibody is a “class-switching” antibody in which the class or subclass has been altered from that of the parent antibody. Chimeric antibodies include their antigen-binding fragment.

[0216] In some respects, chimeric antibodies are humanized antibodies. Typically, nonhuman antibodies are humanized to reduce immunogenicity in humans while retaining the specificity and affinity of the parent nonhuman antibody. Humanized antibodies typically contain one or more variable domains, wherein the CDR (or a portion thereof) is derived from the nonhuman antibody, and the FR (or a portion thereof) is derived from the human antibody sequence. Humanized antibodies may also optionally contain at least a portion of the human constant region. In some respects, some FR residues in the humanized antibody are replaced by corresponding residues from the nonhuman antibody (e.g., the antibody from which the CDR residues are derived), for example, to restore or improve antibody specificity or affinity.

[0217] Humanized antibodies and methods for their preparation are reviewed, for example, in Almagro and Fransson, Front. Biosci. 13:1619-1633 (2008), and further described, for example, in Riechmann et al., Nature 332:323-329 (1988); Queen et al., Proc. Nat'l Acad. Sci. USA 86:10029-10033 (1989); US Patent Nos. 5,821,337, 7,527,791, 6,982,321, and 7,087,409; Kashmiri et al., Methods 36:25-34 (2005) (describing specificity-determining region (SDR) transplantation); Padlan, Mol. Immunol. 28:489-498 (1991) (describing “surface rework”); Dall'Acqua et al., Methods 36:43-60 (2005) (describes “FR reorganization”); and Osbourn et al., Methods 36:61-68 (2005) and Klimka et al., Br. J. Cancer, 83:252-260 (2000) (describes the “guided selection” method for FR reorganization).

[0218] Human frame regions that can be used for humanization include, but are not limited to: frame regions selected using a “best fit” method (see, for example, Sims et al., J. Immunol. 151:2296 (1993)); frame regions derived from the common sequences of human antibodies from specific subgroups of the light or heavy chain variable regions (see, for example, Carter et al., Proc. Natl. Acad. Sci. USA, 89:4285 (1992); and Presta et al., J. Immunol., 151:2623 (1993)); human mature (somatic mutation) frame regions or human germline frame regions (see, for example, Almagro and Fransson, Front. Biosci. 13:1619-1633 (2008)); and frame regions derived from screening FR libraries (see, for example, Baca et al., J. Biol. Chem. 272:10678-10684 (1997) and Rosok et al., J. Biol. Chem. 271:22611-22618 (1996)).

[0219] 4. Human antibodies

[0220] In some respects, the antibodies provided herein are human antibodies. Human antibodies can be generated using a variety of techniques known in the art. Human antibodies are generally described in van Dijk and van de Winkel, Curr Opin Pharmacol. 5:368-74 (2001) and Lonberg, Curr Opin Immunol. 20:450-459 (2008).

[0221] Human antibodies can be prepared by administering an immunogen to a transgenic animal that has been modified to produce intact human antibodies or complete antibodies with human variable regions in response to antigen stimulation. These animals typically contain all or part of a human immunoglobulin locus, which replaces an endogenous immunoglobulin locus, or are present extrachromosomally or randomly integrated into the animal's chromosome. In such transgenic mice, the endogenous immunoglobulin locus is usually inactivated. For a review of methods for obtaining human antibodies from transgenic animals, see Lonberg, Nat. Biotech. 23:1117-1125 (2005). See also, for example, U.S. Patent Nos. 6,075,181 and 6,150,584 describing XENOMOUSE™ technology; U.S. Patent No. 5,770,429 describing HUMAB® technology; U.S. Patent No. 7,041,870 describing KM MOUSE® technology; and U.S. Patent Application Publication No. US 2007 / 0061900 describing VELOCIMOUSE® technology. The human variable region derived from intact antibodies produced by such animals can be further modified, for example, by combining it with different human constant regions.

[0222] Human antibodies can also be prepared using hybridoma-based methods. Human myeloma and mouse-human hybrid myeloma cell lines used to produce human monoclonal antibodies have been described. (See, for example, Kozbor J. Immunol., 133: 3001 (1984); Brodeur et al., Monoclonal Antibody Production Techniques and Applications, pp. 51-63 (Marcel Dekker, Inc., New York, 1987); and Boerner et al., J. Immunol., 147: 86 (1991).) Human antibodies produced via human B-cell hybridoma technology are also described in Li et al., Proc. Natl. Acad. Sci. USA, 103:3557-3562 (2006). Other methods include, for example, those described in U.S. Patent No. 7,189,826 (which describes the production of monoclonal human IgM antibodies from hybridoma cell lines) and Ni, Xiandai Mianyixue, 26(4):265-268 (2006) (which describes human-human hybridoma). Human hybridoma technology (Trioma technology) is also described in Vollmers and Brandlein, Histology and Histopathology, 20(3):927-937 (2005) and Vollmers and Brandlein, Methods and Findings in Experimental and Clinical Pharmacology, 27(3):185-91 (2005).

[0223] Human antibodies can also be generated by isolating variable domain sequences selected from human phage display libraries. These variable domain sequences can then be bound to the desired human constant domain. The technique for selecting human antibodies from antibody libraries is described below.

[0224] 5. Antibodies derived from a library

[0225] In some respects, the antibodies provided herein are derived from libraries. The antibodies of the present invention can be isolated by screening combinatorial libraries for antibodies having one or more desired activities. Methods for screening combinatorial libraries are reviewed, for example, in Lerner et al., Nature Reviews 16:498-508 (2016). For example, various methods are known in the art for generating phage display libraries and screening such libraries to obtain antibodies with desired binding characteristics. Such methods are reviewed in, for example, Frenzel et al., mAbs 8:1177-1194 (2016); Bazan et al., Human Vaccines and Immunotherapeutics 8:1817-1828 (2012); and Zhao et al., Critical Reviews in Biotechnology 36:276-289 (2016), as well as Hoogenboom et al., Methods in Molecular Biology 178:1-37 (edited by O'Brien et al., Human Press, Totowa, NJ, 2001) and Marks and Bradbury, Methods in Molecular Biology 248:161-175 (edited by Lo, Human Press, Totowa, NJ, 2003).

[0226] In some phage display methods, all components of the VH and VL genes are individually cloned by polymerase chain reaction (PCR) and randomly recombined in a phage library. Antigen-binding phages can then be screened from this library, as described by Winter et al., Annual Review of Immunology 12: 433-455 (1994). Phages typically display antibody fragments as single-chain Fv (scFv) fragments or Fab fragments. Libraries from immunized sources provide high-affinity antibodies against immunogens without the need for hybridoma construction. Alternatively, an initial library (e.g., from humans) can be cloned to provide a single source of antibodies against a variety of non-self antigens and self antigens without any immunization, as described by Griffiths et al., EMBO Journal 12: 725-734 (1993). In addition, natural libraries are synthesized by cloning an unrearranged V gene segment from stem cells and using PCR primers containing random sequences to encode the highly variable CDR3 region and perform in vitro rearrangement, as described by Hoogenboom and Winter in Journal of Molecular Biology 227: 381-388 (1992). Patent publications describing human antibody phage libraries include, for example, U.S. Patent Nos. 5,750,373; 7,985,840; 7,785,903 and 8,679,490, and U.S. Patent Publications Nos. 2005 / 0079574, 2007 / 0117126, 2007 / 0237764 and 2007 / 0292936.

[0227] Other examples of methods known in the art for screening combinatorial libraries of antibodies with one or more desired activities include ribosome and mRNA display, as well as methods for displaying and selecting antibodies on bacterial, mammalian, insect, or yeast cells. Methods for yeast surface display are reviewed, for example, in Scholler et al., Methods in Molecular Biology 503:135-56 (2012), Cherf et al., Methods in Molecular Biology 1319:155-175 (2015), and Zhao et al., Methods in Molecular Biology 889:73-84 (2012). Methods for ribosome display are described, for example, in He et al., Nucleic Acids Research 25:5132-5134 (1997), and Hanes et al., PNAS 94:4937-4942 (1997).

[0228] In this paper, antibodies or antibody fragments isolated from human antibody libraries are considered to be human antibodies or human antibody fragments.

[0229] 6. Multispecific antibodies

[0230] In some respects, the antibodies presented herein are multispecific antibodies, particularly bispecific antibodies. A “multispecific antibody” is a monoclonal antibody that has binding specificity to at least two distinct sites (i.e., different epitopes on different antigens or different epitopes on the same antigen). In some respects, multispecific antibodies have three or more binding specificities. In some respects, one of the binding specificities is against human C3bBb, while the others are against any other antigen. In some respects, bispecific antibodies can bind to two (or more) distinct epitopes of human C3bBb. Multispecific (e.g., bispecific) antibodies can also be used to localize cytotoxic agents or cells to cells expressing human C3bBb. Multispecific antibodies can be prepared as full-length antibodies or antibody fragments.

[0231] Techniques for preparing multispecific antibodies include, but are not limited to, recombinant co-expression of heavy-light chain pairs of two immunoglobulins with different specificities (see Milstein and Cuello, Nature 305: 537 (1983)) and engineered “mortar and pestle structures” (see, for example, U.S. Patent 5,731,168 and Atwell et al., J. Mol. Biol. 270:26 (1997)). Multispecific antibodies can also be prepared by: engineering electrostatic manipulation effects for the preparation of antibody Fc-heterodimer molecules (see, for example, WO 2009 / 089004); crosslinking two or more antibodies or fragments (see, for example, U.S. Patent 4,676,980, and Brennan et al., Science, 229: 81 (1985)); using leucine zippers to generate bispecific antibodies (see, for example, Kostelny et al., J. Immunol., 148 (5):1547-1553 (1992) and WO 2011 / 034605); using common light chain techniques to avoid light chain mismatch problems (see, for example, WO 98 / 50431); using “dimeric antibody” techniques for the preparation of bispecific antibody fragments (see, for example, Hollinger et al., Proc. Natl. Acad. Sci. USA, 90:6444-6448 (1993)); and using single-chain Fc-heterodimers. (sFv) dimers (see, for example, Gruber et al., J. Immunol., 152:5368 (1994)); and the preparation of trispecific antibodies as described in Tutt et al., J. Immunol. 147:60 (1991).

[0232] This document also includes engineered antibodies having three or more antigen-binding sites, including, for example, “octopus antibodies” or DVD-Ig (see, for example, WO 2001 / 77342 and WO 2008 / 024715). Other examples of multispecific antibodies having three or more antigen-binding sites can be found in WO 2010 / 115589, WO 2010 / 112193, WO 2010 / 136172, WO 2010 / 145792 and WO 2013 / 026831. Bispecific antibodies or their antigen-binding fragments also include “dual-acting FAbs” or “DAFs”, which contain antigen-binding sites that bind to human C3bBb and two different epitopes of another different antigen or human C3bBb (see, for example, US 2008 / 0069820 and WO 2015 / 095539).

[0233] Multispecific antibodies can also be provided in an asymmetric form, wherein there is domain interchange in one or more binding arms having the same antigen specificity, i.e., by exchanging the VH / VL domain (see, for example, WO 2009 / 080252 and WO2015 / 150447), the CH1 / CL domain (see, for example, WO 2009 / 080253), or the complete Fab arm (see, for example, WO 2009 / 080251, WO 2016 / 016299, and also Schaefer et al., PNAS, 108 (2011) 1187-1191, and Klein et al., MAbs 8 (2016) 1010-20). In one aspect, multispecific antibodies contain cross-Fab fragments. The terms "cross-Fab fragment," "xFab fragment," or "crossover Fab fragment" refer to Fab fragments in which the variable or constant regions of the heavy and light chains are exchanged. Cross-Fab fragments comprise polypeptide chains consisting of a light chain variable region (VL) and a heavy chain constant region 1 (CH1), as well as polypeptide chains consisting of a heavy chain variable region (VH) and a light chain constant region (CL). Asymmetric Fab arms can also be engineered by introducing charged or uncharged amino acid mutations into the domain interfaces to guide correct Fab pairing. See, for example, WO 2016 / 172485.

[0234] Various other molecular forms of multispecific antibodies are known in the art and are included herein (see, for example, Spiess et al., Mol Immunol 67 (2015) 95-106).

[0235] This article also includes a specific type of multispecific antibody, which is a bispecific antibody designed to simultaneously bind to a surface antigen on a target cell (e.g., tumor cells) and an activation-invariant component of the T cell receptor (TCR) complex (such as CD3), for retargeting T cells to kill the target cell. Therefore, in some respects, the antibodies presented herein are multispecific antibodies, particularly bispecific antibodies, wherein one binding specificity is against human C3bBb and the other against CD3.

[0236] Examples of bispecific antibody forms that can be used for this purpose include, but are not limited to, so-called “BiTE” (bispecific T-cell conjugate) molecules, in which two scFv molecules are fused via a flexible linker (see, for example, WO 2004 / 106381, WO 2005 / 061547, WO 2007 / 042261 and WO 2008 / 119567; Nagorsen and Bäuerle, ExpCell Res 317, 1255-1260 (2011)); bispecific antibodies (Holliger et al., Prot Eng 9, 299-305 (1996)) and their derivatives, such as tandem bispecific antibodies (“TandAb”; Kipriyanov et al., J Mol Biol 293, 41-56). (1999)); “DART” (dual affinity retargeting) molecules, which are based on bisomatic antibody forms but characterized by C-terminal disulfide bridges for additional stabilization (Johnson et al., J Mol Biol 399, 436-449 (2010)), and so-called triomab antibodies, which are fully hybridized mouse / rat IgG molecules (reviewed in Seimetz et al., Cancer Treat Rev 36, 458-467 (2010)). The specific T-cell bispecific antibody forms included in this article are described in the following references: WO 2013 / 026833; WO 2013 / 026839; WO 2016 / 020309; Bacac et al., Oncoimmunology 5(8) (2016) e1203498.

[0237] 7. Antibody variants

[0238] In some respects, amino acid sequence variants of the antibodies presented herein are envisioned. For example, it may be desirable to alter the binding affinity and / or other biological properties of the antibody. Amino acid sequence variants of the antibody can be prepared by introducing appropriate modifications into the nucleotide sequence encoding the antibody or by peptide synthesis. Such modifications include, for example, deletion, and / or insertion and / or substitution of residues within the antibody amino acid sequence. Any combination of deletions, insertions, and substitutions can be performed to achieve the final construct, provided that the final construct possesses the desired characteristics, such as antigen binding.

[0239] a) Substitution, insertion, and deletion variants

[0240] In some respects, antibody variants with one or more amino acid substitutions are provided. Sites of interest for substitution mutagenesis include CDR and FR. Conserved substitutions are shown under the heading “Preferred Substitutions” in Table D2. More substantial variations are provided under the heading “Exemplary Substitutions” in Table 1, and are further described below with reference to the amino acid side chain categories. Amino acid substitutions can be introduced into target antibodies, and the products can be screened for desired activities (e.g., retained / improved antigen binding, reduced immunogenicity, or improved ADCC or CDC).

[0241] Table D2: Amino Acid Substitutions

[0242]

[0243] Amino acids can be grouped based on common side-chain characteristics:

[0244] (1) Hydrophobicity: Leucine, Met, Ala, Val, Leu, Ile;

[0245] (2) Neutral hydrophilicity: Cys, Ser, Thr, Asn, Gln;

[0246] (3) Acidic: Asp, Glu;

[0247] (4) Alkaline: His, Lys, Arg;

[0248] (5) Residues affecting chain orientation: Gly, Pro;

[0249] (6) Fang ethnic group: Trp, Tyr, Phe.

[0250] Non-conservative substitution would require swapping members of one of these categories for members of another category.

[0251] One type of substitution variant involves replacing one or more highly variable region residues of a parent antibody (e.g., a humanized antibody or a human antibody). Typically, one or more resulting variants selected for further research will alter (e.g., improve) certain biological properties (e.g., increased affinity, decreased immunogenicity) and / or will substantially retain certain biological properties of the parent antibody, relative to the parent antibody. An exemplary substitution variant is an affinity-matured antibody, which can be conveniently generated, for example, using phage display-based affinity maturation techniques such as those described herein. Briefly, one or more CDR residues are mutated and the variant antibody is displayed on a phage and screened for specific biological activities (e.g., binding affinity).

[0252] For example, alterations (e.g., substitutions) can be made in the CDR to improve antibody affinity. Such alterations can occur in CDR “hotspots,” residues encoded by codons that undergo high-frequency mutations during somatic maturation (see, for example, Chowdhury, Methods Mol. Biol. 207:179-196 (2008)) and / or residues in contact with the antigen, and the binding affinity of the resulting variant VH or VL is detected. Affinity maturation achieved by constructing and reselecting from a secondary library has been described, for example, by Hoogenboom et al. in Methods in Molecular Biology 178:1-37 (edited by O'Brien et al., Human Press, Totowa, NJ, (2001)). In some aspects of affinity maturation, diversity is introduced into the variable gene selected for maturation by any of a variety of methods (e.g., error-prone PCR, strand shuffling, or oligonucleotide directed mutagenesis). A secondary library is then created. This library is subsequently screened to identify any antibody variant with the desired affinity. Another approach to introducing diversity involves CDR targeting, where several CDR residues (e.g., 4 to 6 residues at a time) are randomized. CDR residues involved in antigen binding can be specifically identified, for example, using alanine scanning mutations or modeling. Specifically, CDR-H3 and CDR-L3 are often targeted.

[0253] In some respects, substitution, insertion, or deletion can occur within one or more CDRs, as long as such changes do not substantially reduce the antibody's ability to bind to the antigen. For example, conserved changes that do not substantially reduce binding affinity (e.g., conserved substitutions as provided herein) can be made within the CDR. Such changes can, for example, be external to the antigen-contacting residues in the CDR. In some variant VH and VL sequences provided above, each CDR either remains unchanged or contains no more than one, two, or three amino acid substitutions.

[0254] A method for identifying antibody residues or regions that can be targeted for mutation is called "alanine scan mutation," as described in Cunningham and Wells, (1989) Science, 244:1081-1085. In this method, a residue or a group of target residues (e.g., charged residues such as arg, asp, his, lys, and glu) is identified and replaced with a neutral or negatively charged amino acid (e.g., alanine or polyalanine) to determine if the antibody-antigen interaction is affected. Additional substitutions can be introduced at amino acid positions that exhibit functional sensitivity to the initial substitution. Alternatively or additionally, the contact points between the antibody and antigen can be identified using the crystal structure of the antigen-antibody complex. Such contact residues and adjacent residues can be targeted or eliminated as candidates for substitution. Variants can be screened to determine if they possess the desired properties.

[0255] Amino acid sequence insertions include the fusion of amino and / or carboxyl terms of peptides ranging in length from one residue to one hundred or more residues, as well as intra-sequence insertions of one or more amino acid residues. Examples of terminal insertions include antibodies having an N-terminal methionine residue. Other insertion variants of antibody molecules include the fusion of the N-terminus or C-terminus of the antibody with an enzyme (e.g., for ADEPT (antibody-directed enzyme prodrug therapy)) or peptide that increases the antibody's serum half-life.

[0256] b) Glycosylation variants

[0257] In some respects, the antibodies presented herein can be modified to increase or decrease the degree of antibody glycosylation. The addition or deletion of glycosylation sites to antibodies can be conveniently achieved by altering the amino acid sequence to create or remove one or more glycosylation sites.

[0258] When an antibody contains an Fc region, the oligosaccharide associated with it can be modified. Natural antibodies produced by mammalian cells typically contain branched biantennary oligosaccharides, which are usually linked to Asn297 of the CH2 domain of the Fc region via N-bonding. See, for example, Wright et al., TIBTECH 15:26-32 (1997). Oligosaccharides can include various carbohydrates, such as mannose, N-acetylglucosamine (GlcNAc), galactose, and sialic acid, as well as fucose of GlcNAc attached to the “backbone” of the biantennary oligosaccharide structure. In some aspects, the oligosaccharides in the antibodies of the present invention can be modified to produce antibody variants with certain improved properties.

[0259] On the one hand, antibody variants with unfucosylated oligosaccharides are provided, i.e., oligosaccharide structures lacking (directly or indirectly) fucose linked to the Fc region. Such unfucosylated oligosaccharides (also known as "defucosylated" oligosaccharides) are particularly N-linked oligosaccharides that lack the fucose residues linking the first GlcNAc in the stem of the biantennary oligosaccharide structure. On the other hand, antibody variants with an increased proportion of unfucosylated oligosaccharides in the Fc region compared to natural or parental antibodies are provided. For example, the proportion of unfucosylated oligosaccharides can be at least about 20%, at least about 40%, at least about 60%, at least about 80%, or even about 100% (i.e., no fucose oligosaccharides present). The percentage of defucosylated oligosaccharides, as described, for example, in WO 2006 / 082515, and measured by MALDI-TOF mass spectrometry, is the (average) amount of oligosaccharides lacking fucosylate residues relative to the sum of all oligosaccharides linked to Asn 297 (e.g., complex, hybrid, and high-mannose structures). Asn 297 refers to the asparagine residue (EU number of Fc region residues) located at approximately position 297 in the Fc region; however, due to minor sequence variations in antibodies, Asn 297 can also be located approximately ±3 amino acids upstream or downstream of position 297, i.e., between positions 294 and 300. Such antibodies with an increased proportion of non-fucosylated oligosaccharides in the Fc region may exhibit improved FcγRIIIa receptor binding and / or improved effector function, particularly improved ADCC function. See, for example, US 2003 / 0157108 and US 2004 / 0093621.

[0260] Examples of cell lines capable of producing antibodies with reduced fucosylation include Lec13 CHO cells lacking protein fucosylation (Ripka et al. Arch. Biochem. Biophys. 249:533-545 (1986); US2003 / 0157108; and WO 2004 / 056312, especially in Example 11), and knockout cell lines such as α-1,6-fucosylation gene, FUT8, knockout CHO cells (see, for example, Yamane-Ohnuki et al. Biotech. Bioeng. 87:614-622 (2004); Kanda, Y. et al. Biotechnol. Bioeng., 94(4):680-688 (2006); and WO 2004 / 056312, especially in Example 11), and knockout cell lines such as α-1,6-fucosylation gene, FUT8, knockout CHO cells (see, for example, Yamane-Ohnuki et al. Biotech. Bioeng. 87:614-622 (2004); Kanda, Y. et al. Biotechnol. Bioeng., 94(4):680-688 (2006); and WO 2004 / 056312, especially in Example 11), and knockout cell lines. 2003 / 085107), or cells that have reduced or eliminated activity of GDP-fucose synthesis or transport proteins (see, for example, US2004259150, US2005031613, US2004132140, US2004110282).

[0261] On the other hand, antibody variants provide bipartite oligosaccharides, for example, in which biantennary oligosaccharides linked to the Fc region of the antibody are bipartitely divided by GlcNAc. As mentioned above, such antibody variants can have reduced fucosylation and / or improved ADCC function. Examples of such antibody variants are described, for example, in Umana et al., Nat Biotechnol 17, 176-180 (1999); Ferrara et al., Biotechn Bioeng 93, 851-861 (2006); WO 99 / 54342; WO 2004 / 065540, WO 2003 / 011878.

[0262] Antibody variants having at least one galactose residue in the oligosaccharide linked to the Fc region are also provided. Such antibody variants may have improved CDC function. Such antibody variants are described, for example, in WO 1997 / 30087, WO 1998 / 58964 and WO 1999 / 22764.

[0263] c) Fc region variant

[0264] In some respects, one or more amino acid modifications may be introduced into the Fc region of the antibody provided herein, thereby generating an Fc region variant. The Fc region variant may contain a human Fc region sequence (e.g., a human IgG1, IgG2, IgG3, or IgG4 Fc region) containing an amino acid modification (e.g., substitution) at one or more amino acid positions.

[0265] In some respects, the present invention considers antibody variants possessing some, but not all, of the effector functions, making them ideal candidates for applications where the half-life of the antibody in vivo is important, but certain effector functions (such as complement-dependent cytotoxicity (CDC) and antibody-dependent cell-mediated cytotoxicity (ADCC)) are unnecessary or detrimental. In vitro and / or in vivo cytotoxicity assays can be performed to confirm a reduction / depletion of CDC and / or ADCC activity. For example, Fc receptor (FcR) binding assays can be performed to ensure that the antibody lacks FcγR binding (and therefore may lack ADCC activity), but retains FcRn binding capacity. The primary cells mediating ADCC, NK cells, express only FcγRIII, while monocytes express FcγRI, FcγRII, and FcγRIII. FcR expression on hematopoietic cells is summarized in Table 3 on page 464 of Ravetch and Kinet, Annu. Rev. Immunol. 9:457-492 (1991). Non-limiting examples of in vitro assays for evaluating the ADCC activity of target molecules are described in U.S. Patent No. 5,500,362 (see, for example, Hellstrom, I. et al., Proc. Nat'l. Acad. Sci. USA 83:7059-7063 (1986)) and Hellstrom, I. et al., Proc. Nat'l. Acad. Sci. USA 82:1499-1502 (1985); 5,821,337 (see Bruggemann, M. et al., J. Exp. Med. 166:1351-1361 (1987)). Alternatively, non-radioactive assays may be used (see, for example, the ACTI™ non-radioactive cytotoxicity assay for flow cytometry (Cell Technology, Inc., Mountain View, CA); and the CytoTox 96® non-radioactive cytotoxicity assay (Promega, Madison, Wisconsin)). Useful effector cells for such assays include peripheral blood mononuclear cells (PBMCs) and natural killer (NK) cells. Alternatively or additionally, the ADCC activity of the target molecule can be assessed in vivo, for example, in animal models (such as those disclosed in Clynes et al., Proc. Nat'l Acad. Sci. USA 95:652-656 (1998)). C1q binding assays can also be performed to confirm that the antibody cannot bind C1q and therefore lacks CDC activity. See, for example, C1q and C3c binding ELISAs in WO 2006 / 029879 and WO 2005 / 100402.To assess complement activation, a CDC assay can be performed (see, for example, Gazzano-Santoro et al., J. Immunol. Methods 202:163 (1996); Cragg, MS et al., Blood 101:1045-1052 (2003); and Cragg, MS and MJ Glennie, Blood 103:2738-2743 (2004)). FcRn binding and in vivo clearance / half-life assays can also be performed using methods known in the art (see, for example, Petkova, SB et al., Int'l. Immunol. 18(12):1759-1769 (2006); WO 2013 / 120929 A1).

[0266] Antibodies with reduced effector function include those with substitutions of one or more of the Fc region residues 238, 265, 269, 270, 297, 327, and 329 (US Patent No. 6,737,056). Such Fc mutants include Fc mutants with substitutions at two or more of the amino acids at positions 265, 269, 270, 297, and 327, including the so-called “DANA” Fc mutant, in which residues 265 and 297 are substituted with alanine (US Patent No. 7,332,581).

[0267] Certain antibody variants with improved or reduced binding to FcR are described. (See, for example, U.S. Patent No. 6,737,056; WO 2004 / 056312, and Shields et al., J. Biol. Chem. 9(2):6591-6604 (2001).)

[0268] In some respects, antibody variants contain an Fc region with one or more amino acid substitutions that improve ADCC, such as substitutions at positions 298, 333, and / or 334 of the Fc region (EU numbers of the residues).

[0269] In some aspects, the antibody variant comprises an Fc region having one or more amino acid substitutions that reduce FcγR binding, such as substitutions at positions 234 and 235 of the Fc region (EU numbers of the residues). In one aspect, the substitutions are L234A and L235A (LALA). In some aspects, the antibody variant further comprises D265A and / or P329G in an Fc region derived from the human IgG1 Fc region. In one aspect, in the Fc region derived from the human IgG1 Fc region, the substitutions are L234A, L235A, and P329G (LALA-PG). (See, for example, WO 2012 / 130831). In another aspect, in the Fc region derived from the human IgG1 Fc region, the substitutions are L234A, L235A, and D265A (LALA-DA).

[0270] In some respects, such as those described in U.S. Patent Nos. 6,194,551, WO 99 / 51642 and Idusogie et al. J. Immunol. 164: 4178-4184 (2000), alterations are made in the Fc region resulting in altered (i.e., improved or reduced) C1q binding and / or complement-dependent cytotoxicity (CDC).

[0271] Antibodies with prolonged half-life and improved neonatal Fc receptor (FcRn) binding (responsible for transferring maternal IgG to the fetus) (Guyer, RL et al., J. Immunol. 117:587 (1976), and Kim, JK et al., J. Immunol. 24:249 (1994)) are described in US2005 / 0014934 (Hinton et al.). These antibodies contain an Fc region with one or more substitutions that improve the binding of the Fc region to FcRn. Such Fc variants include Fc variants with substitutions at one or more of the following Fc region residues: 238, 252, 254, 256, 265, 272, 286, 303, 305, 307, 311, 312, 317, 340, 356, 360, 362, 376, 378, 380, 382, ​​413, 424, or 434, for example, a substitution of Fc region residue 434 (see, for example, U.S. Patent No. 7,371,826; Dall'Acqua, WF et al. J. Biol. Chem. 281 (2006) 23514-23524).

[0272] Studies of the human Fc-human FcRn complex have shown that residues I253, S254, H435, and Y436 are crucial for the interaction (Firan, M. et al., Int. Immunol. 13 (2001) 993; Shields, RL et al., J. Biol. Chem. 276 (2001) 6591-6604). Various mutants of residues 248 to 259, 301 to 317, 376 to 382, ​​and 424 to 437 have been reported and examined in Yeung, YA et al. (J. Immunol. 182 (2009) 7667-7671).

[0273] In some respects, antibody variants contain Fc regions with one or more amino acid substitutions that reduce FcRn binding, for example, substitutions at positions 253, and / or 310, and / or 435 of the Fc region (EU numbers of the residues). In some respects, antibody variants contain Fc regions with amino acid substitutions at positions 253, 310, and 435. In one instance, in the Fc region derived from the human IgG1 Fc region, the substitutions are I253A, H310A, and H435A. See, for example, Grevys, A. et al., J. Immunol. 194 (2015) 5497-5508.

[0274] In some respects, antibody variants comprise Fc regions with one or more amino acid substitutions that reduce FcRn binding, for example, substitutions at positions 310, and / or 433, and / or 436 of the Fc region (EU numbers of the residues). In some respects, antibody variants comprise Fc regions with amino acid substitutions at positions 310, 433, and 436. In one instance, in the Fc region derived from the human IgG1 Fc region, the substitutions are H310A, H433A, and Y436A. (See, for example, WO 2014 / 177460A1).

[0275] In some respects, antibody variants comprise an Fc region having one or more amino acid substitutions that increase FcRn binding, such as substitutions at positions 252, and / or 254, and / or 256 of the Fc region (EU numbers of the residues). In some respects, antibody variants comprise an Fc region having amino acid substitutions at positions 252, 254, and 256. In one respect, in the Fc region derived from the human IgG1 Fc region, the substitutions are M252Y, S254T, and T256E. See also Duncan & Winter, Nature 322:738-40 (1988); U.S. Patent No. 5,648,260; U.S. Patent No. 5,624,821; and WO 94 / 29351 for other examples of Fc region variants.

[0276] The C-terminus of the heavy chain of the antibody reported herein may be a full C-terminus ending with the amino acid residue PGK. The C-terminus of the heavy chain may be a shortened C-terminus in which one or two C-terminal amino acid residues have been removed. In a preferred aspect, the C-terminus of the heavy chain is a shortened C-terminus ending with PG. In one aspect of all aspects reported herein, as specified herein, an antibody comprising a heavy chain including a C-terminal CH3 domain comprises a C-terminal glycine-lysine dipeptide (G446 and K447, EU index numbers of amino acid positions). In one aspect of all aspects reported herein, as specified herein, an antibody comprising a heavy chain including a C-terminal CH3 domain comprises a C-terminal glycine residue (G446, EU index number of amino acid position).

[0277] d) Cysteine-engineered antibody variants

[0278] In some respects, it may be desirable to generate cysteine-engineered antibodies, such as THIOMAB™ antibodies, in which one or more residues of the antibody are substituted with cysteine ​​residues. In certain respects, the substituted residues are present at accessible sites on the antibody. As further described herein, by substituting those residues with cysteine, reactive thiol groups are thereby located at accessible sites on the antibody and can be used to conjugate the antibody to other parts (such as drug portions or linker-drug portions) to produce immunoconjugates. Cysteine-engineered antibodies can be generated, for example, as described in U.S. Patent Nos. 7,521,541, 8,30,930, 7,855,275, 9,000,130, or WO 2016040856.

[0279] e) Antibody derivatives

[0280] In some respects, the antibodies provided herein can be further modified to include additional non-protein moieties known in the art and readily available. Suitable moieties for antibody derivatization include, but are not limited to, water-soluble polymers. Non-limiting examples of water-soluble polymers include, but are not limited to, polyethylene glycol (PEG), copolymers of ethylene glycol / propylene glycol, carboxymethyl cellulose, dextran, polyvinyl alcohol, polyvinylpyrrolidone, poly-1,3-dioxolane, poly-1,3,6-trioxane, ethylene / maleic anhydride copolymers, polyamino acids (homogeneous or random copolymers) and dextran or poly(n-vinylpyrrolidone) polyethylene glycol, propylene glycol homopolymers, polypropylene oxide / ethylene oxide copolymers, polyoxyethyleneized polyols (e.g., glycerol), polyvinyl alcohol, and mixtures thereof. PEG-propionaldehyde may be advantageous in manufacturing due to its stability in water. Polymers can have any molecular weight and can be branched or unbranched. The number of polymers attached to the antibody can vary, and if more than one polymer is attached, they can be the same or different molecules. Typically, the number and / or type of polymers used for derivatization can be determined based on the following considerations, including but not limited to the specific properties or functions of the antibody to be improved, and whether the antibody derivative will be used for a limited therapy.

[0281] 8. Immunoconjugates

[0282] The present invention also provides immunoconjugates comprising the anti-human C3bBb antibody described herein, which is conjugated (chemically bonded) to one or more therapeutic agents, such as cytotoxic agents, chemotherapeutic agents, drugs, growth inhibitors, toxins (e.g., protein toxins, bacterial, fungal, plant or animal-derived enzyme-active toxins or fragments thereof) or radioisotopes.

[0283] On the one hand, immunoconjugates are antibody-drug conjugates (ADCs), in which an antibody is conjugated to one or more therapeutic agents. A linker is typically used to connect the antibody to one or more therapeutic agents. An overview of ADC technology is provided in Pharmacol Review 68:3-19 (2016), which includes examples of therapeutic agents, drugs, and linkers.

[0284] In another aspect, the immunoconjugate comprises an antibody conjugated herein to an enzyme-active toxin or a fragment thereof, including but not limited to diphtheria A chain, a non-conjugated active fragment of diphtheria toxin, exotoxin A chain (from Pseudomonas aeruginosa), ricin A chain, absinthecin A chain, senna root toxin A chain, α-arbusculin, tung oil protein, caryophyllein protein, pokeweed antiviral protein (PAPI, PAPII, and PAP-S), bitter melon inhibitor, curcumin, crotonin, soapwort inhibitor, gelatin, mitochondriin, localized aspergillin, phenolmycin, enoxacin, and trichosporine.

[0285] In another aspect, immunoconjugates comprise antibodies described herein that are conjugated with radioactive atoms to form radioconjugates. A variety of radioisotopes can be used to produce radioconjugates. Examples include radioisotopes of At211, I131, I125, Y90, Re186, Re188, Sm153, Bi212, P32, Pb212, and Lu. When a radioconjugate is used for detection, it may contain radioactive atoms for scintillation studies, such as tc99m or I123, or spin labels for nuclear magnetic resonance (NMR) imaging (also known as magnetic resonance imaging, MRI), such as iodine-123, iodine-131, indium-111, fluorine-19, carbon-13, nitrogen-15, oxygen-17, gadolinium, manganese, or iron.

[0286] A variety of bifunctional protein conjugates, such as N-succinimino-3-(2-pyridyldithio)propionate (SPDP), 4-(N-maleiminomethyl)cyclohexane-1-carboxylic acid succinimide ester (SMCC), iminothiacyclopentane (IT), bifunctional derivatives of imino esters (such as dimethyl adipate HCl), active esters (such as disuccinimide octanoate), aldehydes (such as glutaraldehyde), diazid compounds (such as bis(p-azidobenzoyl)hexamethylenediamine), dinitrogen derivatives (such as bis-(p-diazobenzoyl)-ethylenediamine), diisocyanates (such as toluene 2,6-diisocyanate), and difluorinated compounds (such as 1,5-difluoro-2,4-dinitrobenzene), can be used to prepare conjugates of antibodies and cytotoxic agents. For example, ricin immunotoxin can be prepared as described in Vitetta et al., Science 238:1098 (1987). Carbon-14 labeled 1-isothiocyanobenzyl-3-methyldiethylenetriaminepentaacetic acid (MX-DTPA) is an exemplary chelating agent for conjugating radioactive nucleotides to antibodies. See WO 94 / 11026. The adapter can be a “cleavable adapter” that promotes the release of cytotoxic drugs from cells. For example, acid-labile adapters, peptidase-sensitive adapters, light-labile adapters, dimethyl adapters, or disulfide-containing adapters can be used (Chari et al., Cancer Res. 52:127-131 (1992); U.S. Patent No. 5,208,020).

[0287] The immunoconjugates or ADCs discussed in this article are explicitly considered, but not limited to, such conjugates prepared with cross-linking agents, including but not limited to commercially available (e.g., from Pierce Biotechnology, Inc., Rockford, IL., USA) BMPS, EMCS, GMBS, HBVS, LC-SMCC, MBS, MPBH, SBAP, SIA, SIAB, SMCC, SMPB, SMPH, sulfon-EMCS, sulfon-GMBS, sulfon-KMUS, sulfon-MBS, sulfon-SIAB, sulfon-SMCC, sulfon-SMPB, and SVSB (succinimino-(4-vinyl sulfone)benzoate).

[0288] B. Recombination methods and compositions

[0289] Antibodies can be generated using recombinant methods and compositions, such as those described in US 4,816,567. For these methods, one or more isolated nucleic acids encoding the antibody are provided.

[0290] In the case of natural antibodies or fragments of natural antibodies, two nucleic acids are required: one for the light chain or a fragment thereof, and one for the heavy chain or a fragment thereof. These nucleic acids encode the amino acid sequence of the VL containing the antibody and / or the amino acid sequence of the VH containing the antibody (e.g., the light chain and / or heavy chain of the antibody). These nucleic acids can be expressed on the same expression vector or on different expression vectors.

[0291] In the case of bispecific antibodies with heterodimeric heavy chains, four nucleic acids are required: one for the first light chain, one for the first heavy chain containing the Fc region of the first heteromonomer polypeptide, one for the second light chain, and one for the second heavy chain containing the Fc region of the second heteromonomer polypeptide. These four nucleic acids can be contained in one or more nucleic acid molecules or expression vectors. These nucleic acids encode the amino acid sequence constituting the first VL of the antibody and / or the amino acid sequence constituting the first VH containing the Fc region of the first heteromonomer and / or the amino acid sequence constituting the second VL of the antibody and / or the amino acid sequence constituting the second VH containing the Fc region of the second heteromonomer (e.g., the first light chain and / or the second light chain and / or the first heavy chain and / or the second heavy chain of the antibody). These nucleic acids can be on the same expression vector or on different expression vectors; typically, these nucleic acids are located on two or three expression vectors, meaning that one vector can contain more than one of these nucleic acids. Examples of these bispecific antibodies are cross-Mabs (see, for example, Schaefer, W. et al., PNAS, 108 (2011) 11187-1191). For instance, one of the heterologous single-chain heavy chains contains a so-called "palm-like mutation" (T366W, and optionally one of S354C or Y349C), and the other contains a so-called "mortar-like mutation" (T366S, L368A, and Y407V, and optionally Y349C or S354C) (see, for example, Carter, P. et al., Immunotechnol. 2 (1996) 73), according to EU index number.

[0292] In one aspect, isolated nucleic acids encoding antibodies used in the methods reported herein are provided.

[0293] In one aspect, a method for preparing an anti-human C3bBb antibody is provided, wherein the method includes culturing a host cell comprising a nucleic acid encoding an antibody as provided above under conditions suitable for antibody expression, and optionally recovering the antibody from the host cell (or host cell culture medium).

[0294] For the recombinant production of anti-human C3bBb antibodies, the nucleic acid encoding the antibody (e.g., as described above) is isolated and inserted into one or more vectors for further cloning and / or expression in host cells. Such nucleic acids can be readily isolated and sequenced using routine procedures (e.g., by using oligonucleotide probes capable of specifically binding to genes encoding the heavy and light chains of the antibody), or obtained through recombinant methods or chemical synthesis.

[0295] Suitable host cells for cloning or expressing vectors encoding antibodies include prokaryotic or eukaryotic cells as described herein. Antibodies can be generated in bacteria, for example, particularly when glycosylation and Fc effector function are not required. For information on the expression of antibody fragments and peptides in bacteria, see, for example, US 5,648,237, US 5,789,199, and US 5,840,523. (See also Charlton, KA, in: Methods in Molecular Biology, Vol. 248, Lo, BKC (ed.), Humana Press, Totowa, NJ (2003), pp. 245-254, describing the expression of antibody fragments in *E. coli*.) Antibodies can be separated from the bacterial cell paste in a soluble fraction after expression and can be further purified.

[0296] Besides prokaryotes, eukaryotic microorganisms such as filamentous fungi or yeast are also suitable cloning or expression hosts for antibody-encoding vectors. These eukaryotic microorganisms, including fungal and yeast strains, have "humanized" glycosylation pathways, resulting in antibodies with partial or complete human glycosylation patterns. See Gerngross, TU, Nat. Biotech. 22 (2004) 1409-1414; and Li, H. et al., Nat. Biotech. 24 (2006) 210-215.

[0297] Suitable host cells for expressing (glycosylated) antibodies also originate from multicellular organisms (invertebrates and vertebrates). Examples of invertebrate cells include plant cells and insect cells. Many baculovirus strains have been identified that can be used in conjunction with insect cells, particularly for transfecting cells of the meadow armyworm (Spodoptera frugiperda).

[0298] Plant cell cultures can also be used as hosts. See, for example, US 5,959,177, US 6,040,498, US 6,420,548, US 7,125,978 and US 6,417,429 (which describe PLATNIBODIES™ technology for producing antibodies in transgenic plants).

[0299] Vertebrate cells can also be used as hosts. For example, mammalian cell lines adapted for growth in suspension may be useful. Other examples of useful mammalian host cell lines include the monkey kidney CV1 line (COS-7) transformed with SV40; human embryonic kidney cell lines (such as 293 or 293T cells as described, for example, in Graham, FL et al., J. Gen Virol. 36 (1977) 59-74); hamster kidney cells (BHK); mouse Sertoli cells (such as TM4 cells described, for example, in Mather, JP, Biol. Reprod. 23 (1980) 243-252); monkey kidney cells (CV1); African green monkey kidney cells (VERO-76); human cervical cancer cells (HeLa); canine kidney cells (MDCK); Buffalo rat hepatocytes (BRL 3A); human lung cells (W138); human hepatocytes (Hep G2); mouse mammary tumors (MMT 060562); and TRI cells, such as those described, for example, in Mather, JP et al., Annals NYAcad. (As described in Sci. 383 (1982) 44-68); MRC 5 cells; and FS4 cells. Other useful mammalian host cell lines include Chinese hamster ovary (CHO) cells, including DHFR-CHO cells (Urlaub, G. et al., Proc. Natl. Acad. Sci. USA 77 (1980) 4216-4220); and myeloma cell lines such as Y0, NSO, and Sp2 / 0. For a review of certain mammalian host cell lines suitable for antibody production, see, for example, Yazaki, P. and Wu, AM, Methods in Molecular Biology, Vol. 248, Lo, BKC (ed.), Humana Press, Totowa, NJ (2004), pp. 255-268.

[0300] On the one hand, the host cells are eukaryotic cells, such as Chinese hamster ovary (CHO) cells or lymphocytes (e.g., Y0, NSO, Sp20 cells).

[0301] C. Measurement

[0302] The physical / chemical properties and / or biological activity of the anti-C3bBb antibody provided herein can be identified, screened, or characterized by various assays known in the art.

[0303] 1. Combining measurements with other measurements

[0304] In one aspect, the antibodies of the present invention are tested for their antigen-binding activity by known methods such as ELISA and Western blotting.

[0305] On the other hand, competitive assays can be used to identify antibodies that compete with any of the antibodies described herein for binding to C3bBb. In some aspects, such competing antibodies bind to the same epitope (e.g., a linear or conformational epitope) that the antibody of the present invention binds to. Detailed exemplary methods for mapping the epitope bound by the antibody are provided in Morris (1996) “Epitope Mapping Protocols” in Methods in Molecular Biology, Volume 66 (Humana Press, Totowa, NJ).

[0306] In an exemplary competitive assay, immobilized C3bBb is incubated in a solution containing a first labeled antibody (e.g., the antibody of the present invention) that binds to C3bBb and a second unlabeled antibody whose ability to compete with the first antibody for binding C3bBb is being tested. This second antibody may be present in the hybridoma supernatant. As a control, the immobilized C3bBb is incubated in a solution containing the first labeled antibody but not the second unlabeled antibody. After incubation under conditions allowing the first antibody to bind to C3bBb, excess unbound antibody is removed, and the amount of label associated with the immobilized C3bBb is measured. If the amount of label associated with the immobilized C3bBb is substantially reduced relative to the control sample in the test sample, it indicates that the second antibody competes with the first antibody for binding C3bBb. See Harlow and Lane (1988) Antibodies: A Laboratory Manual ch.14 (Cold Spring Harbor Laboratory, Cold Spring Harbor, NY).

[0307] 2. Activity Assay

[0308] In one aspect, an assay is provided for identifying anti-C3bBb antibodies that possess biological activity. Biological activity may include, for example, inhibiting the formation, stability, activity, and dissociation of the C3 convertase protein complex, and affecting interactions with other proteins (e.g., properdin or complement factor I). Antibodies possessing such biological activity in vivo and / or in vitro are also provided.

[0309] In some respects, such biological activities of the antibodies of the present invention are tested.

[0310] D. Methods and compositions for diagnosis and detection

[0311] In some respects, any of the anti-C3bBb antibodies provided herein can be used to detect the presence of C3bBb in biological samples. As used herein, the term "detection" encompasses both quantitative and qualitative detection.

[0312] In one aspect, an anti-C3bBb antibody is provided for use in a diagnostic or detection method. In another aspect, a method for detecting the presence of C3bBb in a biological sample is provided. In some aspects, the method includes contacting the biological sample with the anti-C3bBb antibody under conditions that allow both the anti-C3bBb antibody and C3bBb to bind, and detecting whether a complex has formed between the anti-C3bBb antibody and C3bBb. This method may be an in vitro or in vivo method. In one aspect, the anti-C3bBb antibody is used to select subjects eligible for treatment with the anti-C3bBb antibody, for example, where C3bBb is a biomarker used for patient selection.

[0313] In some respects, labeled anti-C3bBb antibodies are provided. Labeling includes, but is not limited to, labels or portions for direct detection (such as fluorescence, chromogenicity, electronic density, chemiluminescence, and radioactive labeling) and portions for indirect detection (e.g., via enzymatic reactions or molecular interactions) (such as enzymes or ligands). Exemplary labels include, but are not limited to, radioisotopes. 32 P, 14 C 125 I, 3 H and 131I; fluorophores, such as rare earth chelates or fluorescein and its derivatives, rhodamine and its derivatives, dansyl, umbelliferone; luciferases, such as firefly luciferase and bacterial luciferase (US Patent No. 4,737,456); luciferin; dihydrophthalazinedione; horseradish peroxidase (HRP); alkaline phosphatase; β-galactosidase; glucosylamylase; lysozyme; sugar oxidases, such as glucose oxidase, galactose oxidase, and glucose-6-phosphate dehydrogenase; heterocyclic oxidases, such as urate oxidase and xanthine oxidase; conjugated with enzymes that use hydrogen peroxide to oxidize dye precursors (such as HRP, lactoperoxidase, or microperoxidase); biotin / antibiotin protein; spinning labeling; phage labeling; stable free radicals, etc.

[0314] E. Pharmaceutical Composition

[0315] In another aspect, pharmaceutical compositions comprising any of the antibodies provided herein are provided, for example, in any of the following treatment methods. In one aspect, the pharmaceutical composition comprises any of the antibodies provided herein and a pharmaceutically acceptable carrier. In another aspect, the pharmaceutical composition comprises any of the antibodies provided herein and at least one additional therapeutic agent, such as those described below.

[0316] The pharmaceutical compositions (formulations) of anti-C3bBb antibodies described herein can be prepared by combining the antibody with a pharmaceutically acceptable carrier or excipient known to a person skilled in the art. See, for example, Remington's Pharmaceutical Sciences, 16th edition, Osol, A. (ed., 1980), Shire S., Monoclonal Antibodies: Meeting the Challenges in Manufacturing, Formulation, Delivery and Stability of Final Drug Product, 1st edition, Woodhead Publishing (2015), §4, and Falconer R.J., Biotechnology Advances (2019), 37, 107412. Exemplary pharmaceutical compositions of anti-C3bBb antibodies as described herein are lyophilized, aqueous, frozen, etc.

[0317] Pharmaceutically acceptable carriers are generally non-toxic to recipients at the doses and concentrations used, and include, but are not limited to: buffers such as histidine, phosphates, citrates, acetates, and other organic acids; antioxidants, including ascorbic acid and methionine; preservatives (such as octadecyl dimethyl benzyl ammonium chloride; hexamethyl diammonium chloride; benzalkonium chloride; benzyl chloride; phenol, butanol, or benzyl alcohol; alkyl esters of p-hydroxybenzoate, such as methylparaben or propylparaben; catechol; resorcinol; cyclohexanol; 3-pentanol; m-cresol); and low molecular weight (less than about 10). (1 residue) polypeptides; proteins, such as serum albumin, gelatin, or immunoglobulins; hydrophilic polymers, such as polyvinylpyrrolidone; amino acids, such as glycine, glutamine, asparagine, histidine, arginine, or lysine; monosaccharides, disaccharides, and other carbohydrates, including glucose, mannose, or dextrin; chelating agents, such as EDTA; sugars, such as sucrose, mannitol, trehalose, or sorbitol; salt-forming counterions, such as sodium; metal complexes (e.g., zinc protein complexes); and / or nonionic surfactants, such as polyethylene glycol (PEG).

[0318] The pharmaceutical compositions described herein may also contain more than one active ingredient essential for the specific indication being treated, preferably those active ingredients having complementary activities that do not adversely affect each other. Such active ingredients are suitably present in a combination of amounts effective for the intended purpose.

[0319] Pharmaceutical compositions intended for internal administration are typically sterile. For example, sterility can be readily achieved through filtration using a sterile filter membrane.

[0320] F. Treatment methods and routes of administration

[0321] Any of the anti-C3bBb antibodies provided in this article can be used in treatment methods.

[0322] In one aspect, an anti-C3bBb antibody is provided for use as a drug. In another aspect, an anti-C3bBb antibody is provided for treating eye diseases. In one embodiment, the ocular disease is selected from: AMD (in one embodiment, wet AMD, dry AMD, moderate AMD, advanced AMD, and geographic atrophy (GA)), macular degeneration, macular edema, DME (in one embodiment, localized, non-central DME, and diffuse, central DME), retinopathy, diabetic retinopathy (in one embodiment, proliferative DR (PDR), non-proliferative DR (NPDR), and high-altitude DR), other ischemic retinopathy, ROP, retinal vein occlusion (RVO) (in one embodiment, central (CRVO) and branched (BRVO) forms), CNV (in one embodiment, myopic CNV), corneal neovascularization, diseases associated with corneal neovascularization, retinal neovascularization, diseases associated with retinal / choroidal neovascularization, central serous retinopathy (CSR), pathological myopia, Hippel-Lindau syndrome, ocular histoplasmosis, FEVR, Coronary disease, Norrie's disease, and osteoporosis-pseudoglioma syndrome (OPPG). Related retinal abnormalities, subconjunctival hemorrhage, redness and swelling, ocular neovascularization, neovascular glaucoma, retinitis pigmentosa (RP), hypertensive retinopathy, retinal angioma proliferation, macular telangiectasia, iris neovascularization, intraocular neovascularization, retinal degeneration, cystic macular edema (CME), vasculitis, papillary edema, retinitis (including but not limited to CMV retinitis), ocular melanoma, retinoblastoma, conjunctivitis (in one embodiment infectious conjunctivitis and non-infectious conjunctivitis (in one embodiment allergic conjunctivitis)), Lieber's congenital amaurosis (also known as Reye's amaurosis or LCA), uveitis (including infectious and non-infectious uveitis), choroiditis (in one embodiment multifocal choroiditis), ocular histoplasmosis, blepharitis, dry eye, ocular trauma, dry eye disease, and other ophthalmic diseases (wherein such disease is associated with ocular neovascularization, vascular leakage, and / or retinal edema or retinal atrophy). In one embodiment, the eye disease is selected from: AMD (in one embodiment, wet AMD, dry AMD, moderate AMD, advanced AMD, and geographic atrophy (GA)), macular degeneration, macular edema, DME (in one embodiment, localized, non-central DME, and diffuse, centrally involved DME), retinopathy, and diabetic retinopathy (DR) (in one embodiment, proliferative DR (PDR), non-proliferative DR (NPDR), and high-altitude DR).In one embodiment, the eye disease originates from AMD, and in another embodiment, from wet AMD.

[0323] In some aspects, an anti-C3bBb antibody is provided for use in a treatment method. In some aspects, the invention provides an anti-C3bBb antibody for use in a method of treating an individual suffering from an eye disease, the method comprising administering an effective amount of the anti-C3bBb antibody to the individual. In one such aspect, as described below, the method further comprises administering an effective amount of at least one additional therapeutic agent (e.g., one, two, three, four, five, or six additional therapeutic agents) to the individual.

[0324] In another aspect, the present invention provides the use of anti-C3bBb antibodies in the preparation or manufacture of pharmaceuticals. In one aspect, the pharmaceutical remedy is used to treat an eye disease. In another aspect, the pharmaceutical remedy is used in a method of treating an eye disease, the method comprising administering an effective amount of the pharmaceutical remedy to an individual suffering from an eye disease. In one such aspect, as described below, the method further comprises administering an effective amount of at least one other therapeutic agent to the individual. The "individual" according to any of the foregoing aspects can be a human being.

[0325] In another aspect, the present invention provides a method for treating an eye disease. In one aspect, the method includes administering an effective amount of an anti-C3bBb antibody to an individual suffering from such an eye disease. In one such aspect, as described below, the method further includes administering an effective amount of at least one other therapeutic agent to the individual. The "individual" according to any of the foregoing aspects can be a human being.

[0326] In another aspect, the present invention provides a pharmaceutical composition comprising any of the anti-C3bBb antibodies provided herein, for example, in any of the above-described treatment methods. In one aspect, a pharmaceutical composition comprising any of the anti-C3bBb antibodies provided herein, and a pharmaceutically acceptable carrier. In another aspect, the pharmaceutical composition comprises any of the anti-C3bBb antibodies provided herein and at least one additional therapeutic agent, for example, as described below.

[0327] The antibodies of the present invention can be administered alone or in combination therapy. For example, such combination therapy includes administering the antibody of the present invention and administering at least one additional therapeutic agent (e.g., one, two, three, four, five, or six additional therapeutic agents). In some aspects, such combination therapy includes administering the antibody of the present invention and administering at least one additional therapeutic agent, such as a VEGF antagonist.

[0328] Any suitable AMD therapeutic agent can be administered as an additional treatment in combination with the human C3bBb-binding antibody described herein for the treatment of ocular diseases (e.g., AMD, DME, DR, RVO, or GA). Suitable AMD therapeutic agents include, but are not limited to: VEGF antagonists such as anti-VEGF antibodies (e.g., LUCENTIS® (ranibizumab), RTH-258 (formerly ESBA-1008, an anti-VEGF single-chain antibody fragment; Novartis) or bispecific anti-VEGF antibodies (e.g., anti-VEGF / anti-angiogenic peptide 2 bispecific antibodies, such as faricimab or zifibancimig; Roche)), soluble VEGF receptor fusion proteins (e.g., EYLEA® (apasip)), anti-VEGF DARPin® (e.g., abiciparpegol; Molecular Partners AG / Allergan), or anti-VEGF aptamers (e.g., MACUGEN® (pilgatanib sodium)); platelet-derived growth factor (PDGF). Antagonists, such as anti-PDGF antibodies, anti-PDGFR antibodies (e.g., REGN2176-3), anti-PDGF-BB pegylated aptamers (e.g., FOVISTA®; Ophthotech / Novartis), soluble PDGFR receptor fusion proteins, or dual PDGF / VEGF antagonists (e.g., small molecule inhibitors (e.g., DE-120 (Santen) or X-82 (TyrogeneX)) or bispecific anti-PDGF / anti-VEGF antibodies)); VISUDYNE® (verteporfin) in combination with photodynamic therapy; antioxidants; complement system antagonists, e.g., complement factor C5 antagonists (e.g., small molecule inhibitors (e.g., ARC-1905; Opthotech) or anti-C5 antibodies (e.g., LFG-316; Novartis), properdin antagonists (e.g., anti-properdin antibodies, e.g., CLG-561; Alcon), or complement factor D Antagonists (e.g., anti-complement factor D antibodies, such as lampalizumab; Roche)); C3 blocking peptides (e.g., APL-2, Appellis); visual cycle modulators (e.g., emixustat hydrochloride); squalene (e.g., OHR-102; Ofr Pharmaceutical); vitamin and mineral supplements (e.g., those described in Study 1 (AREDS1; zinc and / or antioxidants) and Study 2 (AREDS2; zinc, antioxidants, lutein, zeaxanthin, and / or omega-3 fatty acids) of age-related eye diseases); cell-based therapies, such as NT-501 (Renexus);PH-05206388 (Pfizer), huCNS-SC cell transplantation (StemCells), CNTO-2476 (umbilical cord stem cell line; Janssen), OpRegen (RPE cell suspension; Cell Cure Neurosciences), or MA09-hRPE cell transplantation (Ocata Therapeutics); tissue factor antagonists (e.g., hI-con1; Iconic Therapeutics); α-adrenergic receptor agonists (e.g., brimonidine tartrate; Allergan); peptide vaccines (e.g., S-646240; Shionogi); amyloid β antagonists (e.g., anti-β amyloid monoclonal antibodies, e.g., GSK-933776); S1P antagonists (e.g., anti-S1P antibodies, e.g., iSONEP™; Lpath Inc); ROBO4 antagonists (e.g., anti-ROBO4). Antibodies, such as DS-7080a; Daiichi Sankyo Co., Ltd.); lentiviral vectors expressing endostatin and angiostatin (e.g., RetinoStat); and any combinations thereof. In some cases, AMD treatments (including any prior AMD treatments) may be co-formulated. For example, the anti-PDGFR antibody REGN2176-3 may be co-formulated with aflibercept (EYLEA®). In some cases, such co-formulations may be administered in combination with antibodies that bind to human C3bBb. In some cases, the ocular condition is AMD (e.g., wet AMD).

[0329] Any suitable DME and / or DR therapeutic agent can be administered in combination with the antibody of the present invention that binds to human C3bBb for the treatment of ocular diseases (e.g., AMD, DME, DR, RVO, or GA). The suitable DME and / or DR therapeutic agent includes, but is not limited to, VEGF antagonists (e.g., LUCENTIS® or EYLEA®), corticosteroids (e.g., corticosteroid implants such as OZURDEX® (dexamethasone intravitreal implant) or ILUVIEN® (fluocinolone acetonide intravitreal implant), or corticosteroids formulated for intravitreal injection (e.g., triamcinolone acetonide) or combinations thereof. In some cases, the ocular disease is DME and / or DR.

[0330] An antibody that binds to human C3bBb, as described herein, can be used in combination with therapies or surgical procedures for treating ocular diseases (e.g., AMD, DME, DR, RVO, or GA), including, for example, laser photocoagulation (e.g., panretinal photocoagulation (PRP)), drusen laser, macular hole surgery, macular displacement surgery, implantable microtelescopes, PHI motor angiography (also known as microlaser therapy and feeder vascular therapy), proton beam therapy, microstimulation therapy, retinal detachment and vitrectomy, scleral buckling, macular surgery, transpupillary thermotherapy, photosystem I therapy, RNA interference (RNAi), extracorporeal membrane abortion (also known as membrane differential filtration and rheological therapy), microchip implantation, stem cell therapy, gene replacement therapy, ribozyme gene therapy (including gene therapy for hypoxia-responsive elements, Oxford Biomedica; Lentipak, Genetix; and PDEF gene therapy, GenVec), photoreceptor / retinal cell transplantation (including transplantable retinal epithelial cells, Diacryn, ... Inc.; retinal cell transplantation, such as Astellas Pharma US, Inc., ReNeuron, CHABiotech; acupuncture and combinations thereof.

[0331] Such combination therapies include combined administration (where two or more therapeutic agents are included in the same or different formulations) and single administration. In the case of single administration, the administration of the antibody of the present invention that binds to human C3bBb can be performed before, simultaneously with, and / or after the administration of additional one or more therapeutic agents. In one embodiment, the administration of the antibody of the present invention that binds to human C3bBb and the administration of additional therapeutic agents each occur within about one, two, three, four, or five months, or within about one, two, or three weeks, or within about one, two, three, four, five, or six days.

[0332] The antibodies (and any other therapeutic agents) of this invention can be administered by any suitable means, including parenteral, intrapulmonary, and intranasal administration, and, if desired, for local treatment or intralesional application. Parenteral infusion includes intramuscular, intravenous, intra-arterial, intraperitoneal, or subcutaneous administration. Administration can be carried out by any suitable route, such as by injection, such as intravenous or subcutaneous injection, depending in part on whether the administration is transient or long-term. Various dosing schedules are considered herein, including but not limited to single or multiple administrations at various time points, bolus administration, and pulsatile infusion.

[0333] The antibodies of this invention will be formulated, administered, and applied in accordance with good medical practice. Factors considered in this context include the specific disease being treated, the specific mammal being treated, the individual patient's clinical condition, the cause of the disease, the site of delivery of the agent, the method of administration, the timing of administration, and other factors known to a medical practitioner. The antibody is not mandatory but may optionally be formulated in conjunction with one or more formulations currently used for the prevention or treatment of the disease in question. The effective amount of these other formulations depends on the amount of antibody present in the pharmaceutical composition, the type of disease or treatment, and other factors discussed above. These are generally used at the same dosage and route of administration as described herein, or at about 1% to 99% of the dosage described herein, or at any dosage and via any route determined empirically / clinically to be appropriate.

[0334] For the prevention or treatment of disease, the appropriate dosage of the antibody of the present invention (when used alone or in combination with one or more other additional therapeutic agents) will depend on the type of disease to be treated, the type of antibody, the severity and course of the disease, whether the molecule is administered for preventive or therapeutic purposes, the patient's medical history and response to the antibody, and the discretion of the attending physician. The antibody is appropriately administered to the patient once or in a series of treatments. For repeated administration over several days or longer, treatment will generally continue until the desired suppression of disease symptoms is achieved, depending on the condition. The progress of this therapy can be easily monitored using conventional techniques and assays.

[0335] 3. Specific embodiments of the present invention

[0336] Specific embodiments of the present invention are listed below.

[0337] 1. An antibody that specifically binds to human C3bBb.

[0338] 2. An antibody that binds to human C3bBb, wherein the antibody:

[0339] - Combined with wild-type C3bBb; and / or

[0340] - It binds to recombinant human C3bBb, which contains the Bb subunit of factor B (FB) according to SEQ ID NO:491;

[0341] - Binds to recombinant human C3bBb, which contains the Bb subunit of factor B (FB) according to SEQ ID NO:492; and / or

[0342] - Binds to recombinant human C3bBb, which contains the Bb subunit of factor B (FB) according to SEQ ID NO:493; and / or

[0343] - Binds to recombinant human C3bBb, which contains the Bb subunit of the recombinant factor B (FB) protein, which contains the D279G mutation and optional mutations K350N and / or M458I.

[0344] - Suppress bypass pathways; and / or

[0345] - An agonist or antagonist of C3bBb activity; and / or

[0346] -Specifically binds to cynomolgus monkey C3bBb and human C3bBb, and / or

[0347] -Specifically binds to African green monkey C3bBb and human C3bBb, and / or

[0348] -Binding with an affinity of ≤ 300 pM at 37°C, as measured by SPR.

[0349] 3. An antibody that specifically binds to recombinant human C3bBb, which comprises the Bb subunit of the FB protein having a D279G mutation and optionally further mutations K350N and / or M458I.

[0350] 4. An antibody that specifically binds to recombinant human C3bBb, the recombinant human C3bBb comprising the Bb subunit of factor B (FB) according to SEQ ID NO:491.

[0351] 5. An antibody that specifically binds to recombinant human C3bBb, the recombinant human C3bBb comprising the Bb subunit of factor B (FB) according to SEQ ID NO:492.

[0352] 6. An antibody that specifically binds to recombinant human C3bBb, the recombinant human C3bBb comprising the Bb subunit of factor B (FB) according to SEQ ID NO:493.

[0353] 7. An antibody that specifically binds to an epitope of human C3bBb, the epitope comprising an amino acid residue of the C3b subunit and an amino acid residue of the Bb subunit of human C3bBb.

[0354] 8. An antibody that binds to human C3bBb, wherein the antibody binds to an epitope containing the following amino acid residues on C3bBb, as detected by cryo-electron microscopy:

[0355] -C3b subunits: Arg444, Lys534, Gly539, Ser540, Val524, Lys544, Gly546, Gln547, Ser548, Arg551, Gln557, Gln558, Thr560, Lys562, Glu564, Glu758, Pro759, Lys761, Asn762, Ile764, Leu768, Asn770, Asp797, and

[0356] -Bb subunits: Tyr465, Lys473, Ile474, Ser475, Ile477, Gly482, His483, Lys513, Val514, Ser515, Lys520, Arg521, Asp522, Glu610, and Lys613.

[0357] 9. An antibody that binds to the same or overlapping epitopes of an antibody (P1AF8499) having a VH domain of SEQ ID NO: 39 and a VL domain of SEQ ID NO: 40.

[0358] 10. An antibody that binds to human C3bBb, wherein the antibody comprises a heavy chain CDR and a light chain CDR selected from antibodies in Table D1.

[0359] 11. An antibody that binds to human C3bBb, wherein the antibody comprises a heavy chain variable domain and a light chain variable domain selected from antibodies in Table D1.

[0360] 12. An antibody according to one of the foregoing embodiments, comprising a set of six CDRs, and in one embodiment a set of VH and VL domains, as an antibody selected from antibodies #1 to #217 in Table D1.

[0361] 13. An antibody according to one of the foregoing embodiments, comprising a set of six CDRs, and in one embodiment a set of VH and VL domains, as an antibody selected from antibodies #1 to #122 in Table D1.

[0362] 14. An antibody according to one of the foregoing embodiments, comprising a set of six CDRs, and in one embodiment a set of VH and VL domains, as antibodies selected from antibodies #1 to #7 and antibodies #22 to #217 in Table D1.

[0363] 15. An antibody according to one of the foregoing embodiments, comprising a set of six CDRs, and in one embodiment a set of VH and VL domains, as an antibody selected from antibodies #1 to #7 in Table D1.

[0364] 16. The antibody according to any one of the foregoing embodiments, comprising...

[0365] i) a heavy chain variable domain (VH) comprising: (a) CDR-H1 comprising the amino acid sequence of SEQ ID NO:449, (b) CDR-H2 comprising the amino acid sequence of SEQ ID NO:450, and (c) CDR-H3 comprising the amino acid sequence of SEQ ID NO:451; and a light chain variable domain (VL) comprising: (d) CDR-L1 comprising the amino acid sequence of SEQ ID NO:452, (e) CDR-L2 comprising the amino acid sequence of SEQ ID NO:453, and (f) CDR-L3 comprising the amino acid sequence of SEQ ID NO:454 (corresponding to antibody #1, P1AG9426);

[0366] ii) Heavy chain variable domain (VH) comprising: (a) CDR-H1 comprising the amino acid sequence of SEQ ID NO:455, (b) CDR-H2 comprising the amino acid sequence of SEQ ID NO:456, and (c) CDR-H3 comprising the amino acid sequence of SEQ ID NO:457; and light chain variable domain (VL) comprising: (d) CDR-L1 comprising the amino acid sequence of SEQ ID NO:458, (e) CDR-L2 comprising the amino acid sequence of SEQ ID NO:459, and (f) CDR-L3 comprising the amino acid sequence of SEQ ID NO:460 (corresponding to antibody #2, P1AG9376);

[0367] iii) Heavy chain variable domain (VH) comprising: (a) CDR-H1 comprising the amino acid sequence of SEQ ID NO:461, (b) CDR-H2 comprising the amino acid sequence of SEQ ID NO:462, and (c) CDR-H3 comprising the amino acid sequence of SEQ ID NO:463; and light chain variable domain (VL) comprising: (d) CDR-L1 comprising the amino acid sequence of SEQ ID NO:464, (e) CDR-L2 comprising the amino acid sequence of SEQ ID NO:465, and (f) CDR-L3 comprising the amino acid sequence of SEQ ID NO:466 (corresponding to antibody #3, P1AG9372);

[0368] iv) Heavy chain variable domain (VH) comprising: (a) CDR-H1 comprising the amino acid sequence of SEQ ID NO:467, (b) CDR-H2 comprising the amino acid sequence of SEQ ID NO:468, and (c) CDR-H3 comprising the amino acid sequence of SEQ ID NO:469; and light chain variable domain (VL) comprising: (d) CDR-L1 comprising the amino acid sequence of SEQ ID NO:470, (e) CDR-L2 comprising the amino acid sequence of SEQ ID NO:471, and (f) CDR-L3 comprising the amino acid sequence of SEQ ID NO:472 (corresponding to antibody #4, P1AG9420);

[0369] v) Heavy chain variable domain (VH), comprising: (a) CDR-H1 containing the amino acid sequence of SEQ ID NO:473, (b) CDR-H2 containing the amino acid sequence of SEQ ID NO:474, and (c) CDR-H3 containing the amino acid sequence of SEQ ID NO:475; and light chain variable domain (VL), comprising: (d) CDR-L1 containing the amino acid sequence of SEQ ID NO:476, (e) CDR-L2 containing the amino acid sequence of SEQ ID NO:477, and (f) CDR-L3 containing the amino acid sequence of SEQ ID NO:478 (corresponding to antibody #5, P1AG9391);

[0370] vi) Heavy chain variable domain (VH), comprising: (a) CDR-H1 containing the amino acid sequence of SEQ ID NO:479, (b) CDR-H2 containing the amino acid sequence of SEQ ID NO:480, and (c) CDR-H3 containing the amino acid sequence of SEQ ID NO:481; and light chain variable domain (VL), comprising: (d) CDR-L1 containing the amino acid sequence of SEQ ID NO:482, (e) CDR-L2 containing the amino acid sequence of SEQ ID NO:483, and (f) CDR-L3 containing the amino acid sequence of SEQ ID NO:484 (corresponding to antibody #6, P1AH1205); or

[0371] vii) Heavy chain variable domain (VH), comprising: (a) CDR-H1 containing the amino acid sequence of SEQ ID NO:485, (b) CDR-H2 containing the amino acid sequence of SEQ ID NO:486, and (c) CDR-H3 containing the amino acid sequence of SEQ ID NO:487; and light chain variable domain (VL), comprising: (d) CDR-L1 containing the amino acid sequence of SEQ ID NO:488, (e) CDR-L2 containing the amino acid sequence of SEQ ID NO:489, and (f) CDR-L3 containing the amino acid sequence of SEQ ID NO:490 (corresponding to antibody #7, P1AH1199).

[0372] 17. The antibody according to any one of the foregoing embodiments, comprising a sequence selected from the group consisting of:

[0373] (a) A VH sequence having at least 95% sequence identity with the amino acid sequence of SEQ ID NO:1;

[0374] (b) A VL sequence having at least 95% sequence identity with the amino acid sequence of SEQ ID NO:2; and

[0375] (c) VH sequences as defined in (a) and VL sequences as defined in (b).

[0376] 18. The antibody according to any one of the foregoing embodiments, comprising a sequence selected from the group consisting of:

[0377] (a) A VH sequence having at least 95% sequence identity with the amino acid sequence of SEQ ID NO:3;

[0378] (b) A VL sequence having at least 95% sequence identity with the amino acid sequence of SEQ ID NO:4; and

[0379] (c) VH sequences as defined in (a) and VL sequences as defined in (b).

[0380] 19. The antibody according to any one of the foregoing embodiments, comprising a sequence selected from the group consisting of:

[0381] (a) A VH sequence having at least 95% sequence identity with the amino acid sequence of SEQ ID NO:5;

[0382] (b) A VL sequence having at least 95% sequence identity with the amino acid sequence of SEQ ID NO:6; and

[0383] (c) VH sequences as defined in (a) and VL sequences as defined in (b).

[0384] 20. The antibody according to any one of the foregoing embodiments, comprising a sequence selected from the group consisting of:

[0385] (a) A VH sequence having at least 95% sequence identity with the amino acid sequence of SEQ ID NO:7;

[0386] (b) A VL sequence having at least 95% sequence identity with the amino acid sequence of SEQ ID NO:8; and

[0387] (c) VH sequences as defined in (a) and VL sequences as defined in (b).

[0388] 21. The antibody according to any one of the foregoing embodiments, comprising a sequence selected from the group consisting of:

[0389] (a) A VH sequence having at least 95% sequence identity with the amino acid sequence of SEQ ID NO:9;

[0390] (b) A VL sequence having at least 95% sequence identity with the amino acid sequence of SEQ ID NO:10; and

[0391] (c) VH sequences as defined in (a) and VL sequences as defined in (b).

[0392] 22. The antibody according to any one of the foregoing embodiments, comprising a sequence selected from the group consisting of:

[0393] (a) A VH sequence having at least 95% sequence identity with the amino acid sequence of SEQ ID NO:11;

[0394] (b) A VL sequence having at least 95% sequence identity with the amino acid sequence of SEQ ID NO:12; and

[0395] (c) VH sequences as defined in (a) and VL sequences as defined in (b).

[0396] 23. The antibody according to any one of the foregoing embodiments, comprising a sequence selected from the group consisting of:

[0397] (a) A VH sequence having at least 95% sequence identity with the amino acid sequence of SEQ ID NO:13;

[0398] (b) A VL sequence having at least 95% sequence identity with the amino acid sequence of SEQ ID NO:14; and

[0399] (c) VH sequences as defined in (a) and VL sequences as defined in (b).

[0400] 24. The antibody according to any one of the foregoing embodiments, comprising...

[0401] i) Contains the VH domain of SEQ ID NO:1 and the VL domain of SEQ ID NO:2 (corresponding to antibody #1, P1AG9426).

[0402] ii) Contains the VH domain of SEQ ID NO:3 and the VL domain of SEQ ID NO:4 (corresponding to antibody #2, P1AG9376).

[0403] iii) Contains the VH domain of SEQ ID NO:5 and the VL domain of SEQ ID NO:6 (corresponding to antibody #3, P1AG9372);

[0404] iv) Contains the VH domain of SEQ ID NO:7 and the VL domain of SEQ ID NO:8 (corresponding to antibody #4, P1AG9420);

[0405] v) Contains the VH domain of SEQ ID NO:9 and the VL domain of SEQ ID NO:10 (corresponding to antibody #5, P1AG9391).

[0406] vi) Containing the VH domain of SEQ ID NO:11 and the VL domain of SEQ ID NO:12 (corresponding to antibody #6, P1AH1205); or

[0407] vii) contains the VH domain of SEQ ID NO:13 and the VL domain of SEQ ID NO:14 (corresponding to antibody #7, P1AH1199).

[0408] 25. The antibody according to any one of the foregoing embodiments, comprising the heavy chain of SEQ ID NO:435 and the light chain of SEQ ID NO:436.

[0409] 26. The antibody according to any one of the foregoing embodiments, comprising the heavy chain of SEQ ID NO:437 and the light chain of SEQ ID NO:438.

[0410] 27. The antibody according to any one of the foregoing embodiments, comprising the heavy chain of SEQ ID NO:439 and the light chain of SEQ ID NO:440.

[0411] 28. The antibody according to any one of the foregoing embodiments, comprising the heavy chain of SEQ ID NO:441 and the light chain of SEQ ID NO:442.

[0412] 29. The antibody according to any one of the foregoing embodiments, comprising the heavy chain of SEQ ID NO:443 and the light chain of SEQ ID NO:444.

[0413] 30. The antibody according to any one of the foregoing embodiments, comprising the heavy chain of SEQ ID NO:445 and the light chain of SEQ ID NO:446.

[0414] 31. The antibody according to any one of the foregoing embodiments, comprising the heavy chain of SEQ ID NO:447 and the light chain of SEQ ID NO:447.

[0415] 32. The antibody according to any one of the foregoing embodiments is a monoclonal antibody.

[0416] 33. The antibody according to any one of the foregoing embodiments is a human antibody.

[0417] 34. The antibody according to any one of the foregoing embodiments, wherein the antibody comprises a VH domain having a VH3 framework and a VL domain having a Vκ1 framework.

[0418] 35. The antibody according to any one of the foregoing embodiments, wherein the antibody comprises the human constant region of the IgG isotype.

[0419] 36. The antibody according to any one of the foregoing embodiments is an antibody fragment that binds to human C3bBb.

[0420] 37. The antibody according to any one of the foregoing embodiments is a full-length IgG1 antibody.

[0421] 38. The antibody according to any one of the foregoing embodiments, wherein, as measured by SPR, the antibody binds to human C3bBb with an affinity of ≤300 pM.

[0422] 39. The antibody according to any one of the foregoing embodiments, wherein the antibody is a multispecific antibody.

[0423] 40. An antibody that competitively binds to human C3bBb with an antibody according to any one of the foregoing embodiments.

[0424] 41. An isolated nucleic acid encoding an antibody according to any one of the foregoing embodiments.

[0425] 42. A host cell comprising the nucleic acid as described in Example 42.

[0426] 43. A method for generating an antibody that binds to human C3bBb, comprising culturing host cells as described in Example 42 under conditions suitable for antibody expression.

[0427] 44. The method according to Example 43 further includes recovering antibodies from host cells.

[0428] 45. An antibody produced by the method according to Example 43.

[0429] 46. ​​A pharmaceutical composition comprising an antibody according to any one of Examples 1 to 40 and a pharmaceutically acceptable carrier.

[0430] 47. The pharmaceutical composition according to Example 46 further comprises additional therapeutic agents.

[0431] 48. The antibody according to any one of Examples 1 to 40 or the pharmaceutical composition according to any one of Examples 46 to 47, used as a medicine.

[0432] 49. An antibody according to any one of Examples 1 to 40 or a pharmaceutical composition according to any one of Examples 46 to 47, for the treatment of eye diseases.

[0433] 50. Use of the antibody according to any one of Examples 1 to 40 or the pharmaceutical composition according to any one of Examples 46 to 47 in the manufacture of a medicament for treating an eye disease.

[0434] 51. A method of treating an individual suffering from an eye disease, the method comprising administering to the individual an effective amount of an antibody according to any one of Examples 1 to 40 or a pharmaceutical composition according to any one of Examples 46 to 47.

[0435] Description of amino acid sequence

[0436]

[0437] Example

[0438] The following examples are provided to aid in understanding the invention, the true scope of which is set forth in the appended claims. It should be understood that modifications may be made to the described procedures without departing from the spirit of the invention.

[0439] Example 1:

[0440] Stabilization of human C3bBb complex

[0441] Wild-type C3bBb is a rapidly decaying complex and is unsuitable for antibody discovery purposes. To assess the impact of different mutations within the factor B (FB) subunit on the stability of C3bBb, the following mutations were introduced into the FB protein: D279G, K350N, and M458I. Unmutated FB is indicated in SEQ ID NO: 491, FB with the D279G mutation is indicated in SEQ ID NO: 492, and FB with the D279G, K350N, and M458I mutations is indicated in SEQ ID NO: 493. The mutations are numbered based on the amino acid sequence of human factor B containing the signal peptide (SEQ ID NO: 504). The assembly and decay of the described different C3bBb variants were evaluated on a C3b-coated sensor by biolayer interference (BLI) analysis.

[0442] Stability testing was performed using BLI:

[0443] a) C3bBb assembly in the presence of Mg ions

[0444] To analyze the stability of the formed C3bBb complexes with different Bb subunits, plasma-purified C3b (Complement Technology) was incubated overnight at 4°C in the presence of a 5 mol excess of maleimide-PEG3-biotin (Thermo Fisher Scientific), followed by purification on a Zeba spinning column at 40K (Thermo Fisher Scientific), resulting in biotinylation of C3b at cysteine ​​10¹⁰. This material was then used to load at high density (5 nm BLI signal) onto a streptavidin-functionalized BLI sensor (SA sensor; Fortebio) in an Octet RED 384 device (Fortebio). C3b-loaded sensors were assembled at 22°C in the presence of 54 nM plasma-purified factor D (Complement Technology) in HBS-T(Mg) buffer (150 mM NaCl, 10 mM HEPES, 1 mM MgCl2, 0.005% Tween 20, pH 7.4) with different recombinant factor B variants (i.e., the unmutated D279G variant of SEQ ID NO: 491; the D279G K350N M458I triple variant of SEQ ID NO: 493). After the convertase equilibrated for 12 minutes, the sensor was transferred to 200 µl of HBS-T(Mg) buffer to initiate the dissociation of the assembled C3bBb. Convertase decay was then allowed to occur for three hours.

[0445] After subtracting the signals recorded from the parallel reference sensor (omitting the C3b loading step) and the control sensor (omitting factors B and D), the dissociation phase data were fitted to a double exponential ((A*exp(((-1)*B)*x)))+(C*exp(((-1)*D)*x)) using XLfit (IDBS software), thus allowing for heterogeneity in the assembled invertase population. The half-life (= LN(2) / B) and the relative importance of the major components are reported in Table E1.

[0446] b) C3bBb assembly in the presence of Ni ions

[0447] In another experiment, the Mg structure coordinated in factor B VWA was... 2+ Ions replaced with Ni 2+To further stabilize the C3 convertase, the effect on the stability of the C3bBb complex was investigated using the same setup as previous experiments, except that HBS-T(Mg) buffer (150 mM NaCl, 10 mM HEPES, 1 mM NiCl2, 0.005% Tween 20, pH 7.4) was used instead of HBS-T(Mg).

[0448] The correction for the BLI trace is the same as in the experiment in the presence of magnesium. (Compared to the experiment containing Mg) 2+ Unlike the C3bBb complex, only one substance was detected during dissociation, and analysis was performed using a single exponential decay (A*exp(((-1)*B)*x)). Ni was added during the assembly of C3bBb using different Bb proteins (i.e., wild-type Bb, single-mutant Bb D279G, triple-mutant Bb D279G, K350N, and M458I). 2+ The effect of ions on the formation of the C3bBb complex is shown in Table E1.

[0449] Table E1: t1 / 2 of C3bBb complexes formed using different Bb proteins as measured by BLI

[0450]

[0451] The wild-type C3bBb complex formed in the presence of magnesium ions has a half-life of less than five minutes, and is stabilized by mutations within the Bb subunit of FB and in Ni 2+ The presence of ions leads to the formation of further stabilized C3bBb, resulting in a half-life of up to 11 hours and 15 minutes (equivalent to 675 minutes, which is more than 135 times the C3bBb half-life).

[0452] Assembly and purification of stable C3bBb complexes for antibody discovery

[0453] C3b purified from plasma (Complement Technology) was biotinylated at cysteine ​​1010 by incubation at room temperature for 2 h in the presence of 0.4 mM EZ-Link HPDP-biotin (Thermo Fisher Scientific) followed by purification on a NAP-10 column (Thermo Fisher Scientific). The biotinylated C3b was then mixed at a 1:1 molar ratio with hu factor B (i.e., the unmutated D279G variant of SEQ ID NO: 491; the D279G K350N M458I triple variant of SEQ ID NO: 493), with NiSO4 added to a final concentration of 5 mM. After pre-incubation at room temperature for 10 minutes, factor D (Complement Technology) was added at a molar ratio of 1 (C3b): 1 (factor B): 0.02 (factor D), and incubated for another 10 minutes at room temperature in reaction buffer (20 mM Hepes, 150 mM NaCl, 5 mM NiSO4 pH 7.4). Immediately afterwards, purification was performed at room temperature in storage buffer (20 mM Hepes, 150 mM NaCl, 1 mM NiSO4 pH 7.4) using size exclusion chromatography with Superdex 200 (Cytavia).

[0454] Example 2:

[0455] Antibody screening using a stable C3bBb complex

[0456] The following are the screening methods for antibodies that specifically bind to a stable C3bBb complex:

[0457] Taoxuan

[0458] For initial selection, phage library panning was performed in four rounds, with the first round using 800 nM biotinylated C3bBb triple mutant-HPDP pre-immobilized on Dynabeads M-280 streptavidin (Thermofisher catalog number 11206D) (where C3bBb is formed using FB of SEQ ID NO: 493 in the presence of nickel ions). Rounds 2 through 4 were panned in solution using a 150 nM biotinylated target, followed by capture of the Fab / target complex on the phage on SpeedBead magnetic neutral avidin-coated particles (Cytiva, catalog number 78152104010350, rounds 2 and 4) or Dynabeads M-280 streptavidin (round 3). In summary, in the first round of panning, the phage / target / bead complex was washed 2x with HBS-NT + NiSO4 (supplemented with 0.1% Tween-20 and 1 mM NiSO4 in 0.01 M HEPES pH 7.4; 0.15 M NaCl; Cytiva catalog BR-1006-70) and 1x with HBS-N + NiSO4 (supplemented with 1 mM NiSO4 in 0.01 M HEPES pH 7.4; 0.15 M NaCl; Cytiva catalog BR-1006-70). In panning rounds 2, 3, and 4, the phage / target / bead complex was washed 5x with HBS-NT + NiSO4 and 2x with 1x HBS-N + NiSO4. According to the standard protocol, phage clones with target-specific Fab captured from M-280 beads were eluted using 100mM DTT. These phage clones were then used to infect logarithmic TG1 Escherichia coli (E. coli) cells and rescued using M13K07 helper phages.

[0459] filter

[0460] For the selection of outputs, small-scale preparations of polyclonal plasmids for the corresponding selection rounds were prepared from infected TG1 *E. coli* cells. The plasmids were reformatted to generate soluble Fab in *E. coli* supernatant, which was tagged with a T7 at the C-terminus of the Fab CH1 domain. The ligation polyclonal plasmid encoding the T7-tagged Fab was transformed into TG1 *E. coli* cells (Zymo Research catalog number T3017), and single colonies were picked into microtiter plates. The soluble Fab was expressed in the microtiter plates, and the supernatant was clarified by centrifugation.

[0461] The binding of antibodies to C3bBb was analyzed using different ELISA assays to assess the binding to C3bBb formed from factor B with three mutations: D279G, K350N, and M458I (referred to as "C3bBb triple mutant ELISA"), and to evaluate the binding to different relevant components of the complement system (i.e., C3, C3a, C3b and factors B, Ba, Bb). In short, the following protocol was used:

[0462] C3bBb triple mutant Elisa

[0463] Buffer solution:

[0464] Use 50 mM Hepes pH 7.4 containing 150 mM NaCl, 0.5% BSA, and 0.1% Tween 20 as the ELISA diluent. Use PBS containing 0.1% Tween 20 as the wash buffer.

[0465] method:

[0466] A mixture of 20 µl of 500 ng / ml human C3bBb-human factor B triple mutant (D279G_M350N_M458I)-Cys-HPDP-biotin, 1:3000 mouse anti-human κ HRP (Southern Biotek 9230-05), and 1:2000 mouse IgG anti-T7 tag HRP (Novagen 69048-3) was added to a microcoated streptavidin-coated 384-well microplate (Microcoat 11974998001). 25 µl of sample was added to the plate, and the plate was incubated at room temperature for 1 hour. After washing the culture plate with 90 µl / well of wash buffer using a Biotek EL406 washer, 30 µl of TMB (3,3′,5,5′-3,3,5,5-tetramethylbenzidine, Roche 11835033001) was added. After incubating at room temperature for 5 minutes, the optical density (OD) was measured at 370 nm using an EnVision multimode plate reader (Perkin Elmer).

[0467] C3, C3a, C3b Reverse Screening Elisa

[0468] Buffer solution:

[0469] Use 50 mM Hepes pH 7.4 and 150 mM NaCl as coating buffer. Use 2% BSA from 50 mM Hepes pH 7.4 containing 150 mM NaCl, 0.5% BSA, and 0.1% Tween 20 as blocking buffer. Use 50 mM Hepes pH 7.4 containing 150 mM NaCl, 0.5% BSA, and 0.1% Tween 20 as ELISA diluent. Use PBS containing 0.1% Tween 20 as washing buffer.

[0470] method:

[0471] Clear 384-well Nunc MaxiSorp plates (Nunc, 464718) were coated with a mixture of 25 µl of 5 nM human C3 (Complement Technology, A113), 5 nM human C3a (Complement Technology, A118), and 2,5 nM human C3b (Complement Technology, A114) in coating buffer and incubated at room temperature for 1 hour. After a single wash with 90 µl / well wash buffer on an EL406 Biotek washer, 90 µl of blocking buffer was added and the plates were incubated at room temperature for 30 minutes. After washing the plates three times with 90 µl / well wash buffer on an EL406 Biotek washer, 25 µl of sample was added to the plates and the plates were incubated at room temperature for 1 hour. After washing the plate as described above, add 25 µl of the detection mixture (1:3000 mouse anti-human κ HRP (Southern Biotek 9230-05) and 1:2000 mouse IgG anti-T7 tag HRP (Novagen 69048-3). After incubating at room temperature for 1 hour, wash the plate 6 times with 90 µl / well wash buffer using a Biotek EL406 washer, and add 30 µl of TMB (3,3′,5,5′-3,3,5,5-tetramethylbenzidine, Roche 11835033001). After incubating at room temperature for 5 minutes, measure the optical density (OD) at 370 nm using an EnVision multimode plate reader (Perkin Elmer).

[0472] Factors B, Ba, Bb reverse screening ELISA

[0473] Buffer solution:

[0474] Use 50 mM Hepes pH 7.4 and 150 mM NaCl as coating buffer. Use 2% BSA from 50 mM Hepes pH 7.4 containing 150 mM NaCl, 0.5% BSA, and 0.1% Tween 20 as blocking buffer. Use 50 mM Hepes pH 7.4 containing 150 mM NaCl, 0.5% BSA, and 0.1% Tween 20 as ELISA diluent. Use PBS containing 0.1% Tween 20 as washing buffer.

[0475] method:

[0476] Clear 384-well Nunc MaxiSorp plates (Nunc, 464718) were coated with a mixture of 25 µl of 10 nM human factor B (Complement Technology, A135), 5 nM human Ba ​​(Complement Technology, A154), and 10 nM human Bb (Complement Technology, A155) in coating buffer and incubated at room temperature for 1 hour. After a single wash with 90 µl / well wash buffer on an EL406 Biotek washer, 90 µl of blocking buffer was added and the plates were incubated at room temperature for 30 minutes. After washing the plates three times with 90 µl / well wash buffer on an EL406 Biotek washer, 25 µl of sample was added to the plates and the plates were incubated at room temperature for 1 hour. Wash the plate as described above, and add 25 µl of the detection mixture (1:3000 mouse anti-human κ HRP (Southern Biotek 9230-05) and 1:2000 mouse IgG anti-T7 tag HRP (Novagen 69048-3)) to the sample area. After incubating at room temperature for 1 hour, wash the plate 6 times with 90 µl / well wash buffer using a Biotek EL406 washer, and add 30 µl of TMB (3,3′,5,5′-3,3,5,5-tetramethylbenzidine, Roche 11835033001). After incubating at room temperature for 5 minutes, measure the optical density (OD) at 370 nm using an EnVision multimode plate reader (Perkin Elmer).

[0477] Example 3:

[0478] Identification and antigen binding of C3bBb antibodies

[0479] As described in Example 2, various antibody molecules exhibiting activity in binding to C3bBb are identified from panning and screening, and optionally, the derived antibody molecules are further engineered.

[0480] All antibodies were initially expressed as Fab fragments containing the constant light chain domain of SEQ ID NO:502 and the constant heavy chain domain of SEQ ID NO:503. Table E2 includes an overview of the VH and VL amino acid sequences.

[0481] Table E2: Amino acid sequences of the antibodies identified in Example 2.

[0482]

[0483] From the antibodies identified during the screening period, candidate P1AF8499 (#20 in Table E2, containing VH SEQ ID NO: 39 and VL SEQ ID NO: 40) was selected for further optimization based on its selectivity, invertase inhibition and effect on invertase stability characteristics.

[0484] To this end, two phage libraries were created, each containing diversity of parental candidate antibodies in either the heavy chain CDR or the light chain CDR. As described in Example 2, the libraries were panned and screened against the target.

[0485] Candidates with improved properties compared to the parental molecule were selected, and the heavy and light chains of individual molecules were combined into a single molecule in a combinatorial manner. Improved versions of the parental antibody candidates are antibodies #1 to #7 and antibodies #23 to #217, as further indicated in Table E2 below.

[0486] As described in Example 2, the binding of antibodies to human C3bBb was analyzed using different ELISA assays to analyze the binding to human C3bBb formed from factor B with three mutations: D279G, K350N, and M458I (referred to herein as "C3bBb triple mutant ELISA"), and to assess the binding to different relevant components of the complement system (i.e., C3, C3a, C3b and factors B, Ba, Bb). The results of the different ELISA assays of the antibodies as defined in Table E2 are shown in Table E3.

[0487] Table E3: Antigen binding of C3bBb complexes of antibodies identified in Table E2 using C3bBb triple mutant ELISA

[0488]

[0489] Seven candidate antibodies were generated and their potential for therapeutic applications were further characterized, including five antibodies as full-length IgG1 (Mab) and two antibodies as Fab fragments (see Table E4).

[0490] All Mabs and Fabs were produced using the ExpiCHO expression system (Thermo Fisher Scientific) in serum-free medium. For ExpiCHO cell culture, appropriate high-titer protocols were used. The molar plasmid ratio was target-dependent 1:1 or 1:3 light to heavy chain.

[0491] The antibody was purified by size exclusion chromatography using Kappa Select (Cytavia) followed by Superdex200 (Cytavia) in 20 mM histidine, 140 mM NaCl, pH 6.0. Fab purification began with Kappa Select (Cytavia), followed by a CaptureSelect CH1-XL (Thermo Fisher Scientific) step, and finally by Superdex200 (Cytavia) in 20 mM histidine, 140 mM NaCl, pH 6.0.

[0492] Table E4: Amino acid sequences of the clinical candidate antibodies identified in Example 2.

[0493]

[0494] Example 4:

[0495] The trimer complex formed by C3bBb and the Fab fragment of this invention was analyzed by cryo-electron microscopy.

[0496] The trimer complex of the parental antibody P1AF8499 (#20) and the C3bBb complex was analyzed by electron microscopy. In short, the following protocol was followed:

[0497] Cryo-EM mesh preparation. The recombinant complex of human C3bBb triple mutant + Fab P1AF8499, purified in 20 mM HEPES, 150 mM NaCl, 1 mM NiSO4, pH 7.4, was frozen in liquid nitrogen (0.15 mg / ml, 0.7 µM). After thawing, 3 µl was applied to a fresh glow-discharged Quantifoil R1.2 / 1.3 400-mesh copper grid (Plano). The grid was imprinted (Whatman filter paper, Grade 1) at 20 °C for 4 s at 95% room humidity, and the stopper was frozen in liquid ethane using an EM GP2 plunger freezer (Leica).

[0498] Cryo-EM Data Acquisition Two datasets were collected at the University of Basel C-CINA on the FEI Titan Krios (Thermo Fisher Scientific, Waltham, MATLAB, USA), which was running at 300 keV and equipped with a Gatan Quantum-LS imaging energy filter (GIF, 20 eV energy loss window; Gatan Inc, Pleasanton, CA, USA). In total, 7550 videos of 40 frames were acquired in counting mode using SerialEM software [LK1] with a K2 Summit direct electron detector (Gatan Inc) at a physical pixel size of 0.82 Å and a total electron dose of 56 e− / Ų. Initial quality checks of the captured images were performed using the FOCUS program [LK3], which employed the MotionCor2 program [LK2] for frame shift correction and its dose weighting, and the CTFFIND4 program [LK4] for CTF parameter estimation. Images with a resolution lower than 7 Å or an average drift greater than 2 Å per frame were excluded from the analysis.

[0499] Cryo-EM data processing. Image stacks were processed using cryoSPARC v3.3 (Structura Biotechnology). Frame motion was corrected, and contrast transfer function (CTF) parameters were fitted from the video using the 25–4 Å band of the spectrum. Particle picking was performed on images with a CTF fit of 4.5 Å or higher, selected using a spherical speckle picker with maximum and feature diameters of 100 and 200 Å, respectively. Three rounds of 150-class 2D classification with a box size of 384 pixels were performed to sort C3bBb-P1AF8499 protein particles from debris and other false positives. The remaining particles were subjected to de novo reconstruction and homogeneous refinement to obtain an initial 3D reconstruction, which was used to create a low-pass filter template for another round of particle picking. The picking and classification procedure was repeated to obtain a particle selection of 497,884 particles. A reference volume for 3D heterogeneous refinement was obtained by classifying the particles into 4 classes through de novo reconstruction, and the particles were further classified using a batch size of 2,500 and 4 full iterations. Non-uniform refinement was performed using a category containing the best structural features of 172,991 particles. A sharpened image was used for model building and graph preparation. The `blocres` implementation in `cryoSPARC` was used to filter the image based on local resolution.

[0500] Model Building. The structural solution of C3bBb (PDB 2WIN) was structured into four independent models (TED-CUB, B2-MG, C345C, and the splitting factor B). These models were fitted as rigid bodies to the cryo-EM graph. After modeling P1AF8499 and performing manual adjustments primarily related to the P1AF8499 CDR, a single round of real-space refinement was performed using the phenix.real_space_refinement tool with strict secondary structure constraints (TC1) to correct for global structural discrepancies between the initial model and the graph. The model was further manually adjusted in Coot (TC2) through iterative rounds of model building and real-space refinement. The model-to-graph FSC curve was calculated using Phenix.mtriage, which was consistent with our resolution estimate obtained from the half-graph FSC measurements.

[0501] The results are shown in Figure 1. The left panel shows the trimer complex formed between the P1AF8499 Fab fragment and C3bBb. The epitopes bound by P1AF8499 contain amino acids from both components, C3b and Bb (Figure 1, right panel). Therefore, cross-reactivity with a single subunit of C3bBb should be minimized or even excluded. The following epitopes bound by the P1AF8499 Fab fragment were identified by defining the interactions of amino acid residues within 5 Å of P1AF8499:

[0502] Table E5: Epitopes bound to the parental antibody P1AF8499 Fab fragment

[0503]

[0504] Example 5:

[0505] Antigen binding to the human and cynomolgus monkey C3bBb complex was analyzed by SPR.

[0506] The antibodies in Example 3 were evaluated for binding to C3bBb antigens from different species using the following settings via SPR:

[0507] The Biacore T200 system was precharged with 1x HBS-P+ buffer (10 mM HEPES, 150 mM NaCl, and 0.05% v / v surfactant P20) prepared from a 10x stock (Cytiva BR100671). The CAP chip (part of the Biotin CAPture kit - Cytiva 28920233) was connected to the system and equilibrated overnight.

[0508] Human C3bBb binding characteristics

[0509] The following describes the... C3bBbComplex Measurement. The run buffer was replaced with HBS-P+ supplemented with 5 mM NiSO4 to obtain stability of the C3bBb complex, which was then de novo built up on the chip surface in each cycle. Flow cells (FCs) 1 and 2 were used as active FCs. The sample chamber temperature was set to 12 °C, while the analysis temperatures were defined as 25 °C and 37 °C, respectively, depending on the assay procedure. Each analysis cycle began with an injection of undiluted CAP reagent (part of the Biotin CAPture kit - Cytiva 28920233) at 5 µl / min for 300 s through both FCs. Biotinylated-C3b was injected at 9 nM and 10 µl / min into FCs 1 and FC 2 for 30 s (IgG assay) and 60 s (Fab assay), respectively. Injections were performed separately on each FC to obtain comparable levels between the two FCs. D-biotin (Invitrogen B20656) was injected into two FCs at 50 µM and 10 µl / min for 60 s to block the remaining biotin binding sites. The FB triple mutant was injected into FC2 at 100 nM and 5 µl / min for 300 s to construct the intermediate C3bB on FC2. Subsequently, FD was injected into FC2 at 100 nM and 5 µl / min for 300 s to construct the C3bBb complex. After a 60 s stabilization period, single-cycle kinetic sample injection was initiated. The sample was injected into both FCs at 30 µl / min for 60 s with five 3-fold dilutions at progressively increasing concentrations. Due to the highly stable interactions, the final dissociation time was set at 1200 s at 30 µl / min. To regenerate the surface in the next cycle, standard CAP chip regeneration 8 M Gua-NaOH was injected into both FCs for 120 s. Finally, run buffer was injected into both FCs at 30 µl / min for 60 s, with a final stabilization period of 60 s.

[0510] For the analysis, a dual-reference approach was performed, where the reference flow cell (FC1) signal was subtracted and a surrounding reference blank cycle was performed using only buffer injection. BIAevaluation software The curves were fitted using a 1:1 fitting model of Fab and a heteroligand model of IgG, respectively, to separate affinity binding and affinity binding.

[0511] The results of the interaction with human C3bBb are shown in Table E6.

[0512] Table E6: Human C3bBb binding of the clinical candidate antibody in Example 3.

[0513]

[0514] C3bBb combination characteristics of cynomolgus monkeys

[0515] For measurement Crab-eating macaques C3bBb The kinetics were determined using the settings described above. To construct cynomolgus C3bBb, the corresponding cynomolgus fractions were used: biotinylated cynomolgus C3b (SEQ ID NO: 497), cynomolgus FB (triple mutant, SEQ ID NO: 500), and cynomolgus FD (SEQ ID NO: 498). The concentrations and injection times of the three fractions were modified as follows: biotinylated cynomolgus C3b was injected at 60 nM and 10 µl / min for 180 s (IgG assay) and 300 s (Fab assay), respectively; the cynomolgus FB triple mutant was injected at 400 nM and 5 µl / min for 400 s; and cynomolgus FD was injected at 600 nM and 5 µl / min for 400 s. Each concentration of analyte was injected for 75 s, and the dissociation time was shortened to 300 s. All remaining parameters remained the same as those measured for the human C3bBb complex.

[0516] The results are shown in Table E7.

[0517] Table E7: Cynomolgus monkey C3bBb binding of the clinical candidate antibody in Example 3.

[0518]

[0519] Example 6:

[0520] Assessment of cross-reactivity of other components in the bypass pathway by SPR analysis

[0521] To demonstrate the specificity of the binder for the C3bBb complex, the binding of the complex to individual components, as well as precursors and degradation products, was tested in Biacore (also confirmed in ELISA and BLI). The proteins tested were: C3b (component of the complex), C3 (precursor of C3b), C3d, iC3b, C3a (degradation product of C3b), Bb (component of the complex), FB (precursor of Bb), and Ba (degradation product of FB).

[0522] The off-target SPR measurement is set up as described below.

[0523] The Biacore T200 system was equipped with a CM5 chip and pre-charged with 1x HBS-N (10 mM HEPES, 150 mM NaCl) prepared from a 10x stock solution (Cytiva BR100670). Anti-human Fab antibody (Thermo Fisher Scientific 7103082100) was immobilized in 10 mM acetate at pH 5 at 10 µl / min and 30 µg / ml using default settings and a capture antibody injection time of 420 s on all flow cells (FC1-4) of the amine conjugate.

[0524] For the following analysis, the system was pre-charged with HBS-P + 1 mM NiSO4. After at least three initial conditioning cycles of capture and regeneration, the three antibodies were captured for 20 s at 10 µg / ml on FC 2, 3, and 4, respectively, in each cycle. Off-target injection was performed at 200 nM for 60 s, followed by dissociation at a flow rate of 10 µl / min for 60 s. At the end of each cycle, regeneration was performed for 70 s using 10 mM glycine at pH 2.1 at a final stabilization time of 30 µl / min for 120 s. For each antibody, different off-target injection cycles were surrounded by buffer injection cycles for dual reference purposes.

[0525] To analyze potential off-target binding, default binding and stability reporting points were analyzed. The RU values ​​at both reporting points were extracted after the dual reference curves (using the empty flow cell FC1 and the corresponding empty loop minus the active flow cell). The RU values ​​were normalized relative to binding [%] and stability [%] using the following equations:

[0526] Binding [%] or stability [%] = (Dual reference binding or stability [RU] / capture level [RU]) * (capture molecule [MW] / analyte [MW]) * 100

[0527] The results are shown in Table E8, which reports the response levels immediately after binding and after stability is achieved (calculated as theoretical Rmax in %).

[0528] Table E8: Cross-reactivity of indicator antibodies with bypass pathway components in Example 3

[0529]

[0530] Example 7:

[0531] Further features of the antibody of the present invention

[0532] The thermal stability of the antibody in Example 3 was assessed by exposing it to increasing temperatures in a controlled gradient, and the structural unfolding (T3) was determined. m (through inherent fluorescence) and / or aggregation tendency (T) agg (via SLS).

[0533] Apparent hydrophobicity was assessed by determining the retention time on a hydrophobic HPLC column compared to known highly hydrophobic and lowly hydrophobic standard molecules.

[0534] FcRn affinity was assessed as follows: interaction with immobilized FcRn was measured via SPR, as no specific interaction was expected: biotinylated scFcRn was immobilized onto an S-series sensor chip SA (500 to 600 RU). The antibody was injected at 10 µl / min at 5 nM for 60 s, followed by dissociation for 90 s. The assay was run at 25°C in PBS-P pH 6, followed by surface regeneration at 30 µl / min in PBS-P pH 8 for 60 s. Sensing plots with dual reference signals were compared with FcRn binding capacity (Herceptin, wt FC fraction) and incapacitance (Faricimab, Triple A FC fraction). The positive control was serially diluted to 3.125% of 5 nM using a double-dilution series. Results were evaluated using Biacore T200 evaluation software 3.0 with dual references (control flow cell and buffer cycling). Standard curves were created using the binding response (RU) of the positive control at different concentrations. The residual binding percentage (derived from binding RU) of all candidates at 100% concentration (5 nM) was calculated. All candidates showed very low (< 1 or 5%) FcRn surface interactions compared to the positive control. The negative control (Faricimab 3A) showed a signal similar to P1AH1205 and P1AH1199.

[0535] Heparin affinity chromatography was performed on a specific HPLC affinity column. The corresponding retention times were compared with predetermined thresholds based on known molecules with good and bad PK.

[0536] The quality of antibody integrity was measured by monitoring high molecular weight substances with shorter retention times than the main substance. This was accomplished using a size-limited HPLC setup with UV A 280 nm detection. The obtained peaks were integrated using a custom baseline, and the area under the curve (AUC) was calculated. The relative area of ​​the total integral is displayed. The chromatographic conditions for this method were selected based on the molecular size and the manufacturer's recommendations for protein analysis. Molecular fragmentation was assessed using reduced and untreated capillary gel electrophoresis with intrinsic protein fluorescence detection. Low molecular weight substances with shorter migration times than the main substance were monitored, and the AUC was integrated. The relative area of ​​the total integral is displayed. The functional integrity of the molecule was assessed using a specific binding assay for the molecular target (as described in Example 5). Relative activity concentrations were determined in the case of thermally stressed materials. Estimated asparagine and aspartic acid degradation hotspots were experimentally confirmed or excluded by trypsin peptide mapping mass spectrometry. The peak areas of the extracted ion chromatograms (XIC) were integrated for wild-type and modified peptides. When assessing potential modification sites in the predicted complementarity-determining region, the ratio of the modified peptide XIC area to the wild-type XIC area was considered.

[0537] Viscosity was assessed at different concentrations of the target molecule in a salt-free universal buffer system. This was accomplished by co-incubating the target molecule at different concentrations with beads and detecting the results using dynamic light scattering. The maximum feasible concentration was defined by viscosity limits associated with the characteristics of the target product and determined by fitting available data to an appropriate model and applying relevant thresholds.

[0538] Table E9: Further characteristics of indicator antibodies

[0539]

[0540] Example 8:

[0541] Integrity of the antibody under thermal stress in this invention

[0542] The stability of the antibody in Example 3 was assessed under different stress conditions as follows: Molecules were stored in solutions in screw-cap polypropylene vials under simulated physiological conditions (phosphate-buffered saline, pH 7.4) and shelf-indicating conditions (histidine / HCl, sodium chloride buffer, pH 6.0). The molecular solutions were then subjected to relevant temperatures (physiological: 37°C and elevated stress conditions: 40°C). Subsequent analyses focused on changes observed in the stored samples compared to the untreated control.

[0543] Use the methods for integrity determination from Example 7 to analyze changes.

[0544] Table E10: Residual activity of antibodies treated under stress conditions [%]

[0545]

[0546] Example 9:

[0547] The potency of the antibody of this invention (Wieslab assay and hemolysis assay)

[0548] Wieslab Assay: The potency of the antibody in Example 3 was assessed using the Wieslab Complement System Alternate Pathway (AP) Kit. This kit is an enzyme immunoassay for determining the functional complement alternative pathway in human serum.

[0549] It combines the hemolytic assay principle of complement activation with the detection of generated neoantigens using specific antibodies, which are proportional to the functional activity of the complement pathway. In short, 2-fold dilution series of antibody candidates and control compounds Pegcetacoplan (APL-2, C3 inhibitor, Eur J Clin Invest. 2015 Apr;45(4): 423–440. doi:10.1111 / eci.12419) and Ipoxopan (LNP023, FB inhibitor, Proc Natl Acad Sci USA. 2019 Apr 16;116(16):7926-7931. doi: 10.1073 / pnas.1820892116. Epub 2019 Mar 29) were provided in histidine buffer and then pre-incubated on ice with human serum for 15 min. The actual serum concentration at this point was 90%. The mixture was diluted 1:18 with a specific blocking agent (to activate only AP) and incubated for another 15 min at room temperature. The solution was added to 96-well plates coated with an AP-specific activator and incubated at 37°C for 60 min. The wells were washed according to protocol, and C5b-9 was detected. After 30 min, the absorbance was measured at 405 nm, and the amount of complement activation was calculated using Xlfit.

[0550] Table E11: IC50 of indicator antibodies in Wieslab AP functional assay

[0551]

[0552] Hemolysis Assay: In a separate assay, antibody potency was assessed via a bypass hemolysis assay. Briefly, a 2-fold dilution series of the antibody candidate and control compounds APL-2 (C3-inhibitor) and LNP023 (FB-inhibitor) were provided in histidine buffer. Samples were prepared at 4°C or held on ice to prevent the initiation of complement cascade. Washed rabbit red blood cells (RBCs) were added to the appropriate bypass buffer in a V-shaped 96-well plate along with the antibody dilution series. Normal human serum was added to a final assay concentration of 6.7% to initiate complement activation by incubating the plate at 37°C. The plate was shaken at 800 rpm for 1 min every 10 min to prevent complete RBC sedimentation. After 60 min, the hemolysis reaction was stopped, and ice-cold NaCl solution containing 10 mM EDTA was added. The plate was centrifuged at 1250 x g for 3 min at 4°C to precipitate unlysed RBCs. Carefully transfer the supernatant and measure the absorbance at 405 nm. Calculate the percentage of hemolysis: (OD test sample - OD BG or blank) / (OD completely lysed - OD BG or blank) x 100. Calculate the IC50 value using Xlfit.

[0553] Table E12: IC50 of indicator antibodies in hemolysis assays

[0554]

[0555] In another experiment, antibody potency was assessed via a BLI-based C3b deposition assay.

[0556] BLI-based C3b deposition assay: During C3 cleavage, the activated carbonyl group generated by C3 invertase reacts with a nucleophile near the enzyme to form a covalent adduct. We utilize this property to measure invertase activity by quantitatively depositing C3b on a BLI sensor loaded with C3bBb and exposing it to C3.

[0557] To this end, biotinylated C3b was loaded at a low density onto a streptavidin-functionalized SAX BLI sensor (Sartorius) in an Octet red 384 device to achieve a signal of 1 nm. The invertase was assembled by exposing it to 270 nM plasma purification factor B and 27 nM plasma purification factor D in HBS-T buffer (150 mM NaCl, 10 mM HEPES, 1 mM MgCl2, 0.005% Tween 20, pH 7.4) for 5 min at 30 °C, at which point a dynamic equilibrium was reached between invertase assembly and dissociation. The active sensor was then exposed to different concentrations of C3 (500 nM) and a control sensor for individual testing for 15 min, at which point the invertase was considered completely dissociated. After further exposure to buffer to allow the release of non-covalently related materials for 5 min, the deposited C3b was calculated based on the signal difference between the active and control sensors and used as an indicator for activity and IC50 calculations.

[0558] Table E13: IC50 of indicator antibodies in BLI assay

[0559]

[0560] Example 10:

[0561] The potency of the antibody of this invention compared to existing clinical candidates targeting alternative pathways.

[0562] In the controlled trials, the assays described in previous examples were used to compare the potency of existing-technology clinical candidates targeting bypass pathways. As controls, APL-2 peptide binding to C3 (Eur J Clin Invest. 2015 Apr; 45(4): 423–440. doi:10.1111 / eci.12419) and LNP023 inhibiting FB (Proc Natl AcadSci USA. 2019 Apr 16;116(16):7926-7931. doi: 10.1073 / pnas.1820892116.Epub 2019 Mar 29) were used.

[0563] Table E14: IC50 of the compared compounds

[0564]

[0565] Example 11:

[0566] Cross-reactivity of the antibodies of this invention with different monkey species

[0567] The cross-reactivity of the antibodies of the present invention to C3bBb from different monkey species (namely, the cynomolgus macaque (Macaca Fascicularis / Cynomolgus) and the green monkey (Chlorocebus sabaeus (African green monkey)) was assessed using a potency assay (hemolysis assay), which was based on substances slightly different from those described in the foregoing examples.

[0568] In short, a 2-fold dilution series of the antibody candidate and control compounds APL-2 (C3-inhibitor) and LNP023 (FB-inhibitor) are provided in histidine buffer. Samples are prepared at 4°C or held on ice to prevent the initiation of complement cascades. Washed rabbit red blood cells (RBCs) are added along with the antibody dilution series to a buffer suitable for the bypass pathway in a V-shaped 96-well plate.

[0569] For cynomolgus monkeys, normal human serum was added at a final assay concentration of 5% (serum bank of 2 individuals). Complement activation was initiated by incubating the plate at 37°C. The plate was shaken at 800 rpm for 1 min every 10 min to prevent complete sedimentation of RBCs. After 20 min, the hemolysis reaction was stopped, and ice-cold NaCl solution containing 10 mM EDTA was added. The plate was centrifuged at 1000 xg at 4°C for 3 min to precipitate unlysed RBCs. The supernatant was carefully transferred, and the absorbance was measured at 405 nm. The percentage of hemolysis was calculated as: (OD test sample - OD BG or blank) / (OD completely lysed - OD BG or blank) x 100. The IC50 value was calculated using Xlfit.

[0570] For African green monkeys (AGM), normal human serum was added on an individual basis, with the final assay concentration varying between 4.2% and 6.7%. These concentrations were determined in previous experiments and, along with incubation time and temperature, to obtain appropriate levels of hemolysis. The biggest difference from a “routine” hemolysis assay is that the plate was incubated at room temperature, activation was stopped after 20 min, and the plate was further processed as described above.

[0571] Table E15: Cross-reactivity of indicator compounds in hemolysis assays

[0572]

[0573] Example 12:

[0574] The effect of the antibody of this invention on the dissociation of convertase

[0575] Different anti-C3bBb antibodies (inhibitors) target three-molecule complexes containing C3bBb inhibitors and C3bBb. Effects of decay

[0576] The effect of the C3bBb inhibitor (Inh) on the stability of invertase was investigated by analyzing the decay of the C3bBb:Inh ternary complex assembled on a biolayer interferometry sensor.

[0577] To this end, biotinylated C3b was loaded at a low density onto a streptavidin-functionalized SAX BLI sensor on an Octet red384 device (Sartorius) to achieve a signal of 1.5 nm. The invertase was assembled by exposing the sensor to 270 nM plasma buffer (150 mM NaCl, 10 mM HEPES, 1 mM MgCl2, 0.005% Tween 20, pH 7.4) at 30 °C for 5 min, at which point a dynamic equilibrium was reached between invertase assembly and dissociation. The invertase-loaded sensor was then exposed to 500 nM of assays containing FB and FD for 10 min, allowing the inhibitors to reach binding equilibrium with the invertase.

[0578] The signal decay of the C3bBb:Inh ternary complex was tracked on a control sensor, and the half-life of the ternary complex was calculated by fitting an exponential function using XL fitting (IDBS software). Comparison of the data with parallel assays performed in the absence of inhibitors revealed that some explored C3bBb-specific binders strongly stabilized the invertase, which is a possible role of C3b-Bb interface binding, while other inhibitors did not stabilize the C3 invertase.

[0579] Table E16a:

[0580]

[0581] In another experiment, the effect of the selected inhibitor on the invertase half-life was quantitatively examined under conditions ensuring that the invertase was saturated with the test compound. A similar design to the previous experiment was also implemented, except that C3b loading was limited to a 1 nm BLI signal. Compounds were tested in a 316-fold dilution range, equilibrated with the invertase for 15 min, and exposed to C3 (parallel invertase decay on the control sensor) for 15 min.

[0582] At the highest concentrations tested (31.6 nM, 63.2 nM, 320 nM, and 5 µM of Mab and Fab, respectively), invertase activity was essentially completely inhibited, indicating that invertase occupancy was close to 100% under the experimental conditions. Furthermore, analysis of the BLI signal shift associated with Mab and Fab association confirmed saturation. Therefore, the reported half-life refers to the three-molecule complex, not the mixture of bound / unbound invertases.

[0583] Table E16b:

[0584]

[0585] Example 13:

[0586] The antibody of this invention is a selective inhibitor of the bypass pathway and does not inhibit the classical pathway or the lectin pathway.

[0587] To confirm that the antibody of the present invention is a selective inhibitor of the bypass pathway, a classical pathway hemolysis assay was performed as follows.

[0588] In summary, a 2-fold dilution series of the antibody candidate and control compounds APL-2 (C3-inhibitor) and LNP023 (FB-inhibitor) were provided in histidine buffer. Everything was prepared at room temperature except that the serum was kept on ice to prevent the complement cascade from initiating. Washed Ab-sensitized sheep erythrocytes (EA) were added to a buffer suitable for the classical route in V-shaped 96-well plates along with the antibody dilution series. Normal human serum was added at a final assay concentration of 2% and gently mixed with the other components. Complement activation was initiated by incubating the plate at 37°C with continuous shaking at 600 rpm to prevent total EA sedimentation. After 30 min, hemolysis was stopped by adding an ice-cold NaCl solution containing 10 mM EDTA. The plate was centrifuged at 1000 x g at 4°C for 2 min to precipitate unlysed EA. The supernatant was carefully transferred and the absorbance was measured at 405 nm. Calculate the percentage of hemolysis: (OD test sample - OD BG or EDTA control) / (OD complete lysis - OD BG or EDTA control) x 100. Calculate the IC50 value using Xlfit.

[0589] Table E17: IC50 of indicator molecules assessed by the classical hemolysis assay

[0590]

[0591] As expected, the factor B inhibitor LNP023 does not inhibit the classical pathway. It is specific only for the alternative pathway.

[0592] Example 14:

[0593] Combination with wild-type C3bBb

[0594] The specific binding of the antibody P1AG9426 (#1) of the present invention to the wild-type human C3bBb complex was analyzed by SPR.

[0595] Wild-type human C3bBb SPR determination

[0596] The Biacore T200 system was equipped with a CM5 chip and pre-charged with 1x HBS-N (10 mM HEPES, 150 mM NaCl) prepared from a 10x stock solution (Cytiva BR100670). Anti-human Fab antibody (Thermo Fisher Scientific 7103082100) was immobilized in 10 mM acetate at pH 5 at 10 µl / min and 30 µg / ml using default settings and a capture antibody injection time of 420 s on all flow cells (FC1-4) of the amine conjugate.

[0597] The following method was performed in HBS-P + 1 mM NiSO4. Unless otherwise specified, the sample was diluted in run buffer. The antibody was captured at 10 µl / min (diluted in HBS-N) for 60 s. The pre-constructed C3bBb wt complex was then injected at 10 µl / min for 600 s for dissociation, followed by injection for 120 s. The complex was pre-constructed by incubating with 300 nM human C3b, 310 nM human wt factor B, and 40 nM human factor D at room temperature for 5 min. Surface regeneration was performed by injecting 10 mM glycine at pH 2.1 at 30 µl / min for 60 s. Data were analyzed using signal and buffer cycling on dual reference FC2 to FC1 (injecting run buffer instead of the C3bBb wt complex).

[0598] Example 15:

[0599] C5 invertase inhibition

[0600] Hemolysis assay: The standard hemolysis assay was modified to assess the antibody-specific inhibitory potential at the C5-convertase level in the bypass pathway. Samples were prepared at 4°C or held on ice to prevent the initiation of complement cascade. Washed rabbit red blood cells (RBCs) were added to a low-salt (60 mM NaCl) buffer in a V-shaped 96-well plate, suitable for the bypass pathway but also containing a 200 nM C5 inhibitor (clolimab) to prevent MAC formation. Cells were preheated at 37°C for 10 min, followed by the addition of ice-cold normal human serum at a final assay concentration of 6.7% and incubated for another 10 min with constant oscillation to allow C3-convertase and C5-convertase to form, but without C5 cleavage. To stop further conversion of the convertase, cells were centrifuged by adding a buffer containing 40 mM EDTA, and the pellet was resuspended in two consecutive steps: first with a buffer containing 1.25% final guinea pig serum (as a source of complement protein C5-9 required for MAC formation), EDTA, and antibody (see Table E18 for concentrations), followed by a buffer containing Mg-EGTA. Under these conditions, no new convertase was formed, and the inhibition of hemolysis was a direct result of the inhibition of early-formed C5-convertase. Hemolysis was measured after incubating cells at 37°C with constant oscillation for 1 h.

[0601] Table E18: Inhibition of C5 convertase, indicating antibody levels

[0602]

Claims

1. An antibody that binds to human C3bBb.

2. An antibody that binds to human C3bBb, wherein the antibody: - Combined with wild-type C3bBb; and / or - Combined with recombinant human C3bBb, which comprises the Bb subunit of factor B (FB) according to SEQ ID NO:491; -To bind to recombinant human C3bBb, wherein the recombinant human C3bBb comprises a Bb subunit of factor B (FB) according to SEQ ID NO:492; and / or -To bind to recombinant human C3bBb, wherein the recombinant human C3bBb comprises a Bb subunit of factor B (FB) according to SEQ ID NO:493; and / or - Binds to recombinant human C3bBb, which comprises the Bb subunit of recombinant factor B (FB) protein, wherein the recombinant FB protein comprises a D279G mutation and optionally a K350N mutation and / or an M458I mutation; - Suppress bypass pathways; and / or - An agonist or antagonist of C3bBb activity; and / or -Specifically binds to cynomolgus monkey C3bBb and human C3bBb. -Specifically binds to African green monkey C3bBb and human C3bBb. - As measured by SPR, it binds with an affinity of ≤ 300 pM at 37°C, and / or - Do not stabilize C3 invertase.

3. An antibody that binds to human C3bBb, said antibody binding to an epitope of human C3bBb, said epitope comprising amino acid residues of a C3b subunit and amino acid residues of a Bb subunit of human FB.

4. An antibody that binds to human C3bBb, said antibody binding to an epitope, as detected by cryo-electron microscopy, containing the following amino acid residues on C3bBb: -C3b subunits: Arg444, Lys534, Gly539, Ser540, Val524, Lys544, Gly546, Gln547, Ser548, Arg551, Gln557, Gln558, Thr560, Lys562, Glu564, Glu758, Pro759, Lys761, Asn762, Ile764, Leu768, Asn770, Asp797, and -Bb subunits: Tyr465, Lys473, Ile474, Ser475, Ile477, Gly482, His483, Lys513, Val514, Ser515, Lys520, Arg521, Asp522, Glu610, Lys613.

5. An antibody that binds to the same or overlapping epitopes of an antibody (P1AF8499) having a VH domain of SEQ ID NO: 39 and a VL domain of SEQ ID NO:

40.

6. An antibody that binds to human C3bBb, wherein the antibody comprises a heavy chain CDR and a light chain CDR selected from antibodies in Table D1.

7. The antibody according to any one of the preceding claims, comprising: i) a heavy chain variable domain (VH) comprising: (a) CDR-H1 containing the amino acid sequence of SEQ ID NO:449, (b) CDR-H2 containing the amino acid sequence of SEQ ID NO:450, and (c) CDR-H3 containing the amino acid sequence of SEQ ID NO:451; and a light chain variable domain (VL) comprising: (d) CDR-L1 containing the amino acid sequence of SEQ ID NO:452, (e) CDR-L2 containing the amino acid sequence of SEQ ID NO:453, and (f) CDR-L3 containing the amino acid sequence of SEQ ID NO:454 (corresponding to antibody #1, P1AG9426); ii) Heavy chain variable domain (VH) comprising: (a) CDR-H1 comprising the amino acid sequence of SEQ ID NO:455, (b) CDR-H2 comprising the amino acid sequence of SEQ ID NO:456, and (c) CDR-H3 comprising the amino acid sequence of SEQ ID NO:457; and light chain variable domain (VL) comprising: (d) CDR-L1 comprising the amino acid sequence of SEQ ID NO:458, (e) CDR-L2 comprising the amino acid sequence of SEQ ID NO:459, and (f) CDR-L3 comprising the amino acid sequence of SEQ ID NO:460 (corresponding to antibody #2, P1AG9376); iii) Heavy chain variable domain (VH) comprising: (a) CDR-H1 comprising the amino acid sequence of SEQ ID NO:461, (b) CDR-H2 comprising the amino acid sequence of SEQ ID NO:462, and (c) CDR-H3 comprising the amino acid sequence of SEQ ID NO:463; and light chain variable domain (VL) comprising: (d) CDR-L1 comprising the amino acid sequence of SEQ ID NO:464, (e) CDR-L2 comprising the amino acid sequence of SEQ ID NO:465, and (f) CDR-L3 comprising the amino acid sequence of SEQ ID NO:466 (corresponding to antibody #3, P1AG9372); iv) Heavy chain variable domain (VH) comprising: (a) CDR-H1 comprising the amino acid sequence of SEQ ID NO:467, (b) CDR-H2 comprising the amino acid sequence of SEQ ID NO:468, and (c) CDR-H3 comprising the amino acid sequence of SEQ ID NO:469; and light chain variable domain (VL) comprising: (d) CDR-L1 comprising the amino acid sequence of SEQ ID NO:470, (e) CDR-L2 comprising the amino acid sequence of SEQ ID NO:471, and (f) CDR-L3 comprising the amino acid sequence of SEQ ID NO:472 (corresponding to antibody #4, P1AG9420); v) Heavy chain variable domain (VH), comprising: (a) CDR-H1 containing the amino acid sequence of SEQ ID NO:473, (b) CDR-H2 containing the amino acid sequence of SEQ ID NO:474, and (c) CDR-H3 containing the amino acid sequence of SEQ ID NO:475; and light chain variable domain (VL), comprising: (d) CDR-L1 containing the amino acid sequence of SEQ ID NO:476, (e) CDR-L2 containing the amino acid sequence of SEQ ID NO:477, and (f) CDR-L3 containing the amino acid sequence of SEQ ID NO:478 (corresponding to antibody #5, P1AG9391); vi) Heavy chain variable domain (VH), comprising: (a) CDR-H1 containing the amino acid sequence of SEQ ID NO:479, (b) CDR-H2 containing the amino acid sequence of SEQ ID NO:480, and (c) CDR-H3 containing the amino acid sequence of SEQ ID NO:481; and light chain variable domain (VL), comprising: (d) CDR-L1 containing the amino acid sequence of SEQ ID NO:482, (e) CDR-L2 containing the amino acid sequence of SEQ ID NO:483, and (f) CDR-L3 containing the amino acid sequence of SEQ ID NO:484 (corresponding to antibody #6, P1AH1205); or vii) Heavy chain variable domain (VH), comprising: (a) CDR-H1 containing the amino acid sequence of SEQ ID NO:485, (b) CDR-H2 containing the amino acid sequence of SEQ ID NO:486, and (c) CDR-H3 containing the amino acid sequence of SEQ ID NO:487; and light chain variable domain (VL), comprising: (d) CDR-L1 containing the amino acid sequence of SEQ ID NO:488, (e) CDR-L2 containing the amino acid sequence of SEQ ID NO:489, and (f) CDR-L3 containing the amino acid sequence of SEQ ID NO:490 (corresponding to antibody #7, P1AH1199).

8. The antibody according to any one of the preceding claims, wherein the antibody comprises a VH domain having a VH3 framework and a VL domain having a Vκ1 framework.

9. The antibody according to any one of the preceding claims, comprising a sequence selected from the group consisting of: i) Contains the VH domain of SEQ ID NO:1 and the VL domain of SEQ ID NO:2 (corresponding to antibody #1, P1AG9426). ii) Contains the VH domain of SEQ ID NO:3 and the VL domain of SEQ ID NO:4 (corresponding to antibody #2, P1AG9376). iii) Contains the VH domain of SEQ ID NO:5 and the VL domain of SEQ ID NO:6 (corresponding to antibody #3, P1AG9372); iv) Contains the VH domain of SEQ ID NO:7 and the VL domain of SEQ ID NO:8 (corresponding to antibody #4, P1AG9420); v) Contains the VH domain of SEQ ID NO:9 and the VL domain of SEQ ID NO:10 (corresponding to antibody #5, P1AG9391). vi) Containing the VH domain of SEQ ID NO:11 and the VL domain of SEQ ID NO:12 (corresponding to antibody #6, P1AH1205); or vii) contains the VH domain of SEQ ID NO:13 and the VL domain of SEQ ID NO:14 (corresponding to antibody #7, P1AH1199).

10. An isolated nucleic acid encoding an antibody according to any one of claims 1 to 8.

11. A host cell comprising the nucleic acid according to claim 9.

12. A method for generating an antibody that binds to human C3bBb, the method comprising culturing a host cell according to claim 10 under conditions suitable for the expression of the antibody.

13. A pharmaceutical composition comprising an antibody according to any one of claims 1 to 8 and a pharmaceutically acceptable carrier.

14. The antibody according to any one of claims 1 to 8 or the pharmaceutical composition according to claim 12, used as a drug.

15. Use of the antibody according to any one of claims 1 to 8 or the pharmaceutical composition according to claim 12 in the manufacture of a medicament for treating eye diseases.

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