Antibodies targeting c5ar

Abstract

To provide novel antibodies or antibody fragments specifically binding to human C5aR.SOLUTION: The present invention relates to antibodies or antibody fragments that have combined beneficial properties and are therefore useful for the treatment of inflammatory or autoimmune diseases or cancer.SELECTED DRAWING: None

Description

[Technical field]

[0001] The present disclosure relates to antibodies that interact with C5aR, and in particular with human C5aR. The disclosure also relates to nucleic acid compositions, vector compositions and host cells capable of expressing said antibodies, pharmaceutical compositions comprising said antibodies, and the use of said antibodies for the treatment of specific diseases and / or for diagnostic purposes. [Background technology]

[0002] C5a anaphylatoxin chemotactic receptor 1 (C5aR), also known as CD88, is a G-protein coupled receptor (GPCR) belonging to the rhodopsin family and is one of two high affinity receptors for the ligand, C5a, which is produced in serum as one of the core effector components of the complement response. Under physiological conditions, C5a acts as a chemoattractant for inflammatory cells, stimulating their respiratory burst and the release of cytokines and chemokines and the function to increase vascular permeability.

[0003] C5aR appears to be widely expressed by various cell types. Maximum C5aR expression levels have been described for neutrophils. Low to moderate expression levels have been shown for macrophages / monocytes, dendritic cells, mast cells, eosinophils, pulmonary vascular smooth muscle cells, astrocytes, microglia, osteoblasts, osteoclasts, epithelial and endothelial cells (Non-Patent Document 1, Non-Patent Document 2). Low expression levels have been observed for human T cells (Non-Patent Document 3).

[0004] The cellular response to C5a is tightly regulated by ligand-induced receptor internalization. It has been described that C5aR is rapidly and dose-dependently internalized upon C5a treatment, with up to 90% of the receptors being recycled back to the cell surface. Thus, high expression levels of C5aR, coupled with its rapid turnover rate, may be limiting in terms of efficiency due to the target-mediated drug elimination (TMDD) effect.

[0005] The interaction of C5aR with C5a has been described in many different disease settings; most of them are involved in inflammation and autoimmune diseases (4, 5). Several early studies have been performed to identify the underlying mechanisms by which C5a stimulates tumor growth (6, 7, 8). Most of the data suggests that C5a increases cancer cell proliferation, intratumoral angiogenesis, and promotes tumor invasiveness and metastasis. More recent data also suggest a role for C5a / C5aR in generating an immunosuppressive environment in the context of solid tumors (9, 10, 11), resulting in the promotion of primary tumor growth by inhibiting antitumor responses (e.g., increased recruitment of C5aR-expressing myeloid cells such as myeloid-derived suppressor cells (MDSCs) or M2 macrophages). Based on these findings, combination strategies with known antitumor agents, such as immune checkpoint protein inhibitors, to promote a subject's immune response by reducing the immunosuppressive microenvironment are the focus of research and clinical development (Non-Patent Document 12).

[0006] To date, only one specific complement therapy has been approved, which targets the upstream molecule of C5a, namely C5. The humanized anti-C5 mAb eculizumab is capable of binding to C5 and preventing its cleavage and formation of C5a and C5b proteins, followed by MAC formation. Various other therapeutic monoclonal C5-specific antibodies, such as ALXN1210 or LFG316, are under clinical evaluation (Non-Patent Document 5). However, since increased risk of infection is a major concern with chronic C5 treatment, it is clearly advantageous to specifically target downstream molecules while preventing the biological activity of other complement components. Thus, the development of many antagonistic C5a-specific monoclonal antibodies is underway.

[0007] Direct targeting of C5aR has many advantages over targeting C5 or C5a, respectively. First, receptor-only inhibition preserves MAC activity, thereby reducing the potential risk of infection. Second, C5aR blockade allows continued C5a interaction with its second receptor, C5L2. Since C5L2 has been reported to have anti-inflammatory effects, maintaining an effective C5L2 signaling pathway may result in improved efficacy or reduced dosing requirements. Third, direct C5aR targeting may offer pharmacodynamic advantages over inhibition of soluble C5a due to its low molecular weight and high turnover rate. Overall, there is strong interest in developing C5aR inhibitors, including aptamers, peptides, and non-peptide small molecules that are being tested in preclinical and clinical trials.

[0008] Neutralizing polyclonal antisera or monoclonal antibodies directed against the N-terminal extracellular domain of human C5aR and capable of disrupting C5aR-C5a interaction have been described in the art (e.g., (Non-Patent Document 13), (Non-Patent Document 14)).

[0009] However, these C5aR-specific antibodies are not suitable for clinical development and therapeutic use in humans, particularly because they are of animal origin (making them immunogenic in human patients), are clonal, and / or lack cross-reactivity to related animal species.

[0010] Therapeutic antibodies targeting C5aR are being considered for clinical development. However, clinical development of the antagonistic C5aR-specific antibody Neutrazumab, a humanized IgG4 mAb, was halted in phase II clinical trials due to problems with immune cell depletion and immunogenicity (Non-Patent Document 15). Because C5aR appears to be constitutively expressed on a variety of cell types, it is important that antagonistic antibodies do not induce any depletion of target cells.

[0011] To overcome the shortcomings of Neutrazumab, a second generation C5aR-specific antibody, namely NNC0215-0384 (Patent Document 1) (NOVO NORDISK); clone 32F3A6GL), was generated. This antibody is a human IgG1 antibody derived from a transgenic mouse and is currently in clinical development in the cancer field as IPH5401 (Non-Patent Document 16). IPH5401 possesses a silenced human IgG1 Fc region to eliminate the antibody's ability to induce effector functions. This antibody is referred to herein as RefMAB#1. [Prior art documents] [Patent documents]

[0012] [Patent Document 1] US Patent Application Publication No. 2013 / 0295116 [Non-patent literature]

[0013] [Non-Patent Document 1] Monk PN et al.,Br J Pharmacol.2007,152:429-448 [Non-Patent Document 2] Wetsel RA, Immunol Lett.1995,44:183-187 [Non-Patent Document 3] Nataf S,J Immunol.1999,162:4018-4023 [Non-Patent Document 4] Morgan BP et al.,Nat Rev Drug Discov.2015,14:857-877 [Non-Patent Document 5] Hawksworth OA et al.,Mol Immunol.2017,89:36-43 [Non-Patent Document 6] Markiewski MM et al.,Nat Immunol.2008,9:1225-1235 [Non-Patent Document 7] Corrales L.et al.,J Immunol.2012,189:4674-4683

Non-Patent Document 8

Non-Patent Document 9

Non-Patent Document 10

Non-Patent Document 11

Non-Patent Document 12

Non-Patent Document 13

Non-Patent Document 14

Non-Patent Document 15

Non-Patent Document 16

Summary of the Invention

Problems to be Solved by the Invention

[0014] The present disclosure provides novel antibodies and antibody fragments. [Means for solving the problem]

[0015] The antibodies and antibody fragments disclosed herein can specifically bind to human C5aR and preferably cross-react with C5aR from cynomolgus monkeys. Thus, in some embodiments, the disclosed antibodies are specific for human C5aR and cynomolgus monkey C5aR. In some other embodiments, the disclosed antibodies or antibody fragments bind to the N-terminal extracellular domains of human and cynomolgus monkey C5aR.

[0016] This is in contrast to the above-referenced prior art antibody IPH5401, which binds to the second extracellular loop of human C5aR and, as a result, lacks binding to cynomolgus monkey C5aR, a commonly used relevant toxicology species.

[0017] Furthermore, the inventors of the present invention have surprisingly found that the presently claimed C5aR-specific antibodies are not only significantly more potent in neutralizing pathophysiological C5a concentrations when compared to IPH5401, but also demonstrate increased efficacy over time in inhibiting C5-mediated activation of neutrophils in vitro.

[0018] Thus, in some embodiments, the disclosed antibodies can efficiently inhibit C5a-induced C5aR activity in vitro, particularly notably at pathophysiological C5a concentrations. The disclosed antibodies or antibody fragments can also inhibit C5a-induced leukocyte activation in vitro, as determined by their ability to inhibit C5a-induced upregulation of CD11b in granulocytes and / or monocytes. In some embodiments, the disclosed antibodies or antibody fragments have an IC of 42 nM in the presence of 150 nM human C5a in vitro. 50Inhibits human C5a-induced CD11b expression in human granulocytes at concentrations. In some other embodiments, the disclosed antibodies may exhibit increased potency in inhibiting C5a-induced upregulation of CD11b in granulocytes and / or monocytes after extended incubation times. The disclosed antibodies may also be effective in inhibiting C5a-induced neutrophil migration.

[0019] In summary, the present disclosure provides novel antibodies that are superior to known C5aR-specific antibodies from the art. In particular, the antibodies of the present disclosure are human antibodies with high affinity binding to human C5aR, which preferably cross-react with cynomolgus monkey C5aR, and have favorable functional and safety properties not previously observed. These properties make the antibodies of the present disclosure highly desirable for therapeutic applications, such as for preventing and / or treating inflammatory and autoimmune diseases and cancer.

[0020] The present disclosure provides isolated antibodies or antibody fragments that specifically bind to human C5aR, having CDR regions according to Table 1 or Table 2 herein. The present disclosure also provides isolated antibodies or antibody fragments specific for human C5aR, having a variable heavy chain region (VH) and a variable light chain region (VL) that comprise amino acid sequences according to Table 1 or Table 2 herein. The present disclosure also provides isolated antibodies or antibody fragments specific for C5aR, having a heavy chain (HC) and a light chain (LC) that comprise amino acid sequences according to Table 1 or Table 2 herein.

[0021] The isolated antibody of the present disclosure induces substantially no effector function in vitro. Such effector function may include ADCP, ADCC, or CDC. Furthermore, the isolated antibody or antibody fragment of the present disclosure comprises one or more amino acid substitutions selected from the group of L234A, L235E, G237A, A330S, and P331S, as numbered according to the EU index. In particular, the isolated antibody or antibody fragment of the present disclosure comprises a variant human IgG1 Fc region, which comprises the following amino acid substitutions: L234A, L235E, G237A, A330S, and P331S, as numbered according to the EU index.

[0022] The present disclosure also provides an isolated antibody or antibody fragment of the present disclosure for use in medicine.

[0023] The present disclosure also provides a method for treating a subject suffering from a disease, such as an inflammatory or autoimmune disease or cancer, by administering to said subject an effective amount of an antibody or antibody fragment of the present disclosure. Preferably, said subject is a human.

[0024] The present disclosure also provides a pharmaceutical composition comprising an isolated antibody or antibody fragment of the present disclosure and a pharma- ceutically acceptable carrier.

[0025] The present disclosure also provides nucleic acid compositions encoding the isolated antibodies or antibody fragments of the present disclosure.The present disclosure also provides vector compositions comprising the nucleic acid compositions encoding the isolated antibodies or antibody fragments of the present disclosure.The present disclosure also provides host cells comprising vector compositions or nucleic acid compositions encoding the isolated antibodies or antibody fragments of the present disclosure.

[0026] The present disclosure also provides a method for treating a subject suffering from a disease, such as an inflammatory disease, an autoimmune disease, or cancer, by administering to the subject an effective amount of an isolated antibody or antibody fragment of the present disclosure, preferably the subject is a human.

[0027] The present disclosure also provides a pharmaceutical composition comprising an isolated antibody or antibody fragment of the present disclosure and a pharma- ceutically acceptable carrier.

[0028] The claimed antibodies or antibody fragments have utility, as do the claimed methods for identifying such antibodies or antibody fragments.

[0029] The use of the claimed antibodies or antibody fragments is to modulate the biological activity of human C5aR. In particular, the claimed antibodies or antibody fragments are for therapeutic applications such as the treatment of inflammatory or autoimmune diseases or cancer. [Brief description of the drawings]

[0030] [Figure 1] Cellular binding of MAB#1, MAB#2, RefMAB#1 and negative isotype control MOR03207 to human and cynomolgus C5aR as determined via FACS. A Dose-response binding to human C5aR overexpressed on Flp-In CHO cells. B Dose-response binding to cynomolgus C5aR overexpressed on Flp-In CHO cells. C Mean dose-response binding to purified human neutrophils from whole blood of three different donors. D Mean dose-response binding to purified cynomolgus neutrophils from whole blood from three different monkeys. [Diagram 2] Cellular binding of MAB#1, MAB#2, RefMAB#1 and the negative isotype control MOR03207 to human, cynomolgus monkey and rodent C5aR, and to the C5aR-related GPCRs human C5L2, human C3aR, human FPR1 and human ChemR23, determined via FACS at an IgG concentration of 600 nM. [Diagram 3] ELISA binding of MAB#1 to two naturally occurring variants of human C5aR and to mouse C5aR expressed on virus-like particles (VLPs). [Figure 4] PathHunter®-β-arrestin assay from DiscoveRx. Neutralization of human C5a-induced β-arrestin recruitment. A Log dose-response curves for increasing concentrations of recombinant human C5a in the absence or presence of MAB#1 (50 nM), MAB#2 (50 nM) and RefMAB#1 (50 nM). B % inhibition was calculated for three increasing concentrations of human C5a (1.2 nM, 11 nM and 100 nM respectively) at a final IgG concentration of 50 nM. [Diagram 5]Inhibition of human C5a-induced CD11b upregulation in human granulocytes at typical and pathophysiological C5a concentrations. A+B Comparison of MAB#1, MAB#2 and RefMAB#1 in a CD11b whole blood assay. Log dose inhibition curves are shown. Human granulocytes were gated as target cells. As a quantitative readout, the IgG concentration required to reach 50% inhibition of CD11b upregulation was calculated (IC50 concentration). The IC50 concentration for each IgG is shown below the x-axis. A Log dose response curves for increasing concentrations of IgG and 15 nM human C5a. B Log dose response curves for increasing concentrations of IgG and 150 nM human C5a. [Figure 6] Inhibition of human C5a-induced CD11b upregulation in human granulocytes and monocytes determined over extended incubation times. Comparison of MAB#1 and RefMAB#1 in the CD11b whole blood assay after incubation of IgG with target cells for either 20 or 300 min. IgG was added in serial dilutions and incubated for either 20 or 300 min, followed by stimulation with 15 nM human C5a. Gating was performed on either granulocytes (FIGS. 6A and B) or monocytes (FIGS. 6C and D) as target cells. CD11b levels were determined. Log dose inhibition curves are shown. Data are expressed as % inhibition of CD11b upregulation with 15 nM human C5a. Results for MAB#1 are shown in FIG. 6A and C, and results for RefMAB#1 are provided in FIG. 6B and D. [Figure 7] Inhibition of human C5a-induced human neutrophil migration. MAB#1 and negative control MOR03207 were each tested at two IgG concentrations (100 nM and 600 nM, respectively) in the presence of 10 nM human C5a. Mean values ​​from three independent assay runs at three different time points (15 min, 25 min, 35 min) are shown. Neutrophils were obtained from three different human donors. % inhibition was calculated based on neutrophil migration in the absence of antibody. [Figure 8]Promega ADCC and ADCP reporter bioassays. Comparison of MAB#1 and a monoclonal anti-C5aR control IgG carrying either a wild-type (non-inhibitory) human IgG1 Fc region or a mutated (inhibitory) Fc region identical to that of MAB#1. In addition, an isotype control antibody MOR03207 with either a mutant or wild-type human IgG1 Fc region was included. The assays were performed according to the supplier's instructions using either modified Jurkat cells expressing FcγRIIa_H to mimic the ADCP pathway or Jurkat cells expressing FcγRIIIa, the V158 high affinity mutant to mimic the ADCC pathway, and C5aR expressing CHO cells. A Results from an ADCP reporter bioassay at an IgG concentration of 10 μg / ml. B Results from an ADCC reporter bioassay at an IgG concentration of 10 μg / ml. Data are provided as the mean fluorescence signal over background. [Figure 9] Group mean pharmacokinetic profile of MAB#1 in Han Wistar rats following a single intravenous dose of 10 mg / kg IgG. Data are presented as mean (IgG concentration over time) ± standard deviation (S) (n=3). [Figure 10] Protein panel profiling (3P) results for MAB#1 and MAB#2. Numbers for each antibody represent the resulting binding signal for each antibody and protein tested, compared to the binding of the isotype negative control antibody MOR03207. Antibodies were tested at concentrations of 10 nM and 100 nM, respectively. [Figure 11] Cytokine release by in vitro matured M1 and M2 macrophages derived from monocytes. After treatment with MAB#1 and overnight incubation with C5a, IL-10 and IL-12 levels were determined by ELISA. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0031] The present disclosure relates to a number of human antibodies that recognize human C5aR.

[0032] definition The term "C5aR" refers to a protein known as the C5a anaphylatoxin chemotactic receptor 1 or CD88.

[0033] Human C5aR (Uniprot:P21730|1-350) (herein referred to as the "D / K mutant") is MDSFNYTTPDYGHYDDKDTLDLNTPVDKTSNTLRVPDILALVIFAVVFLVGVLGNALVVWVTAFEAKRTINAIWFLNLAVADFLSCLALPILFTSIVQHHHWPFGGAACSILPSLILLNMYASILLLATISADRFLLVFKPIWCQNFRGAGLAWIACAVAWGLALLLTIPSFLYRVVREEYFPPKVLCGVDYSHDKRRERAVAIVRLVLGFLWPLLTLTICYTFILLRTWSRRATRSTKTLKVVVAVVASFFIFWLPYQVTGIMMSFLEPSSPTFLLKKLDSLCVSFAYINCCINPIIYVVAGQGFQGRLRKSLPSLLRNVLTEESVVRESKSFTRSTVDTMAQKTQAV (SEQ ID NO: 1) It has the amino acid sequence:

[0034] Two naturally occurring missense mutations in human C5aR have been described (http: / / www.uniprot.org / uniprot / P21730): Reference SNP (refSNP) Cluster Report: rs4467185 (MAF: 0.03) and Cluster Report: rs11880097 (MAF: 0.03). One mutation is located within the N-terminal extracellular domain of human C5aR (position 2 of SEQ ID NO:1) and results in a D to N substitution.

[0035] The human C5aR protein containing both naturally occurring missense mutations in its sequence (also referred to herein as the "N / N mutant") MNSFNYTTPDYGHYDDKDTLDLNTPVDKTSNTLRVPDILALVIFAVVFLVGVLGNALVVWVTAFEAKRTINAIWFLNLAVADFLSCLALPILFTSIVQHHHWPFGGAACSILPSLILLNMYASILLLATISADRFLLVFKPIWCQNFRGAGLAWIACAVAWGLALLLTIPSFLYRVVREEYFPPKVLCGVDYSHDKRRERAVAIVRLVLGFLWPLLTLTICYTFILLRTWSRRATRSTKTLKVVVAVVASFFIFWLPYQVTGIMMSFLEPSPTFLLLNKLDSLCVSFAYINCCINPIIYVVAGQGFQGRLRKSLPSLLRNVLTEESVVRESKSFTRSTVDTMAQKTQAV (SEQ ID NO: 2) It has the amino acid sequence:

[0036] The cynomolgus monkey (Macaca fascicularis) C5aR MDPFSSTTLDYEHYDGKNVLDSDTPVDKTSNTLRVPDILALVVFAVVFLVGVLGNALVVWVTAFEVKRTINAIWFLNLAVADFLSCLALPILFTSIVQHHHWPFGGTACRILPSLILLNMYASILLLATISADRFLLVFNPIWCQNFRGAGLAWIACAVAWGLALLLTIPSFLYRAVRQEEYSPKVLCGVDYNNDTRRERAVAIVRLVLGFLWPLLTLMICYTFLLLRTWSRRATRSTKTLKVVVAVVASFFIFWLPYQVTGTMMSFLRPSSPTYLQLKKLDSLSISFAYINCCINPVIYVVAGQGFQGRLRKSLPSLLRNVLTEESVVRESKSFTRSTVDTMTEKTQAV (SEQ ID NO: 3) It has the amino acid sequence:

[0037] Mouse (Mus musculus) C5aR MDPIDNSSFEINYDHYGTMAPNIPADGIHLPKRQPGDVAALIIYSVVFLVGVPGNALVVWVTAFEARRAVNAIWFLNLAVADLLSCLALPVLFTTVLNHNYWYFDATACIVLPSLILLNMYASILLLATISADRFLLVFKPIWCQKVRGTGLAWMACGVAWVLALLLTIPSFVYREAYKDFYSEHTVCGINYGGGSFPKEKAVAILRLMVGFVLPLLTLNICYTFLLLRTWSRKATRSTKTLKVVMAVVICFFIFWLPYQVTGVMIAWLPPSSPTLKRVEKLNSLCVSLAYINCCVNPIIYVMAGQGFHGRLLRSLPSIIRNALSEDSVGRDSKTFTPSTTDTSTRKSQAV (SEQ ID NO: 4) It has the amino acid sequence:

[0038] Rat (Rattus norvegicus) C5aR MDPISNDSSEITYDYSDGTPNPDMPADGVYIPKMEPGDIAALIIYLAVFLVGVTGNALVVWVTAFEAKRTVNAIWFLNLAVADLLSCLALPILFTSIVKHNHWPFGDQACIVLPSLILLNMYSSILLLATISADRFLLVFKPIWCQKFRRPGLAWMACGVTWVLALLLTIPSFVFRRIHKDPYSDSILCNIDYSKGPFFIEKAIAILRLMVGFVLPLLTLNICYTFLLIRTWSRKATRSTKTLKVVMAVVTCFFVFWLPYQVTGVILAWLPRSSSTFQSVERLNSLCVSLAYINCCVNPIIYVMAGQGFHGRLRRSLPSIIRNVLSEDSLGRDSKSFTRSTMDTSTQKSQAV (SEQ ID NO: 5) It has the amino acid sequence:

[0039] The term "C5a" refers to the protein known as human complement component C5a.

[0040] Human C5a (Uniprot: P01031|678-751) TLQKKIEEIAAKYKHSVVKKCCYDGACVNNDETCEQRAARISLGPRCIKAFTECCVVASQLRANISHKDMQLGR (SEQ ID NO: 6) It has the amino acid sequence:

[0041] The term "C5L2" refers to a protein known as the C5a anaphylatoxin chemotactic receptor 2.

[0042] Human C5L2 is MGNDSVSYEYGDYSDLSDRPVDCLDGACLAIDPLRVAPLPLYAAIFLVGVPGNAMVAWVAGKVARRRVGATWLLHLAVADLLCCLSLPILAVPIARGGHWPYGAVGCRALPSIILLTMYASVLLLAALSADLCFLALGPAWWSTVQRACGVQVACGAAWTLALLLTVPSAIYRRLHQEHFPARLQCVVDYGGSSSTENAVTAIRFLFGFLGPLVAVASCHSALLCWAARRCRPLGTAIVVGFFVCWAPYHLLGLVLTVAAPNSALLARALRAEPLIVGLALAHSCLNPMLFLYFGRAQLRRSLPAACHWALRESQGQDESVDSKKSTSHDLVSEMEV (SEQ ID NO: 7) It has the amino acid sequence:

[0043] The term "C3aR" refers to a protein known as the C3a anaphylatoxin chemotactic receptor.

[0044] Human C3aR is MASFSAETNSTDLLSQPWNEPPVILSMVILSLTFLLGLPGNGLVLWVAGLKMQRTVNTIWFLHLTLADLLCCLSLPFSLAHLALQGQWPYGRFLCKLIPSIIVLNMFASVFLLTAISLDRCL VVFKPIWCQNHRNVGMACSICGCIWVVACVMCIPVFVYREIFTTDNHNRCGYKFGLSSSLDYPDFYGDPLENRSLENIVQPPGEMNDRLDPSSFQTNDHPWTVPTVFQPQTFQRPSADSLPR GSARLTSQNLYSNVFKPADVVSPKIPSGFPIEDHETSPLDNSDAFLSTHLKLFPSASSNSFYESELPQGFQDYYNLGQFTDDDQVPTPLVAITITRLVVGFLLPSVIMIACYSFIVFRMQRGRFAKSQSKTFRVAVVVVAVFLVCWTPYHIFGVLSLLTDPETPLGKTLMSWDHVCIALASANSCFNPFLYALLGKDFRKKARQSIQGILEAAFSEELTRSTHCPSNNVISERNSTTV (SEQ ID NO: 8) It has the amino acid sequence:

[0045] The term "FPR1" refers to a protein known as the fMet-Leu-Phe receptor.

[0046] Human FPR1 is METNSSLPTNISGGTPAVSAGYLFLDIITYLVFAVTFVLGVLGNGLVIWVAGFRMTHTVTTISYLNLAVADFCFTSTLPFFMVRKAMGGHWPFGWFLCKFLFTIVDINLFGSVFLIALIALDRCVCVLHPVWTQNHRTVSLAKKVIIGPWVMALLLTLPVIIRVTTVPGKTGTVACTFNFSPWTNDPKERINVAVAMLTVRGIIRFIIGFSAPMSIVAVSYGLIATKIHKQGLIKSSPPLRVLSFVAAAFFLCWSPYQVVALIATVRIRELLQGMYKEIGIAVDVTSALAFFNSCLNPMLYVFMGQDFRERLIHALPASLERALTEDSTQTSDTATNSTLPSAEVALQAK (SEQ ID NO: 9) It has the amino acid sequence:

[0047] The term "ChemR23" refers to a protein known as chemokine-like receptor 1.

[0048] Human ChemR23 is MRMEDEDYNTSISYGDEYPDYLDSIVVLEDLSPLEARVTRIFLVVVYSIVCFLGILGNGLVIIIATFKMKKTVNMVWFLNLAVADFLFNVFLPIHITYAAMDYHWVFGTAMCKISNFLLIHNMFTSVFLLTIISSDRCISVLLPVWSQNHRSVRLAYMACMVIWVLAFFLSSPSLVFRDTANLHGKISCFNNFSLSTPGSSSWPTHSQMDPVGYSRHMVVTVTRFLCGFLVPVLIITACYLTIVCKLHRNRLAKTKKPFKIIVTIIITFFLCWCPYHTLNLLELHHTAMPGSVFSLGLPLATALAIANSCMNPILYVFMGQDFKKFKVALFSRLVNALSEDTGHSSYPSHRSFTKMSSMNERTSMNERETGML (SEQ ID NO: 10) It has the amino acid sequence:

[0049] The term "antibody" as used herein refers to a protein that comprises at least two heavy (H) chains and two light (L) chains interconnected by disulfide bonds and that interacts with an antigen. Each heavy chain is composed of a heavy chain variable region (abbreviated herein as VH) and a heavy chain constant region. The heavy chain constant region is composed of three domains, CH1, CH2 and CH3. Each light chain is composed of a light chain variable region (abbreviated herein as VL) and a light chain constant region. The light chain constant region is composed of one domain, CL. The VH and VL regions can be further divided into regions called complementarity determining regions (CDRs) that are hypervariable, interspersed with regions that are more conserved, called framework regions (FRs). Each VH and VL is composed of three CDRs and four FRs, arranged from amino terminus to carboxy terminus in the following order: FR1, CDR1, FR2, CDR2, FR3, CDR3 and FR4. The variable regions of the heavy and light chains contain binding domains that interact with antigens. The constant regions of the antibody may mediate the binding of the immunoglobulin to host tissues or factors, including various cells of the immune system (e.g., effector cells) and the first component (C1q) of the classical complement system. The term "antibody" includes, for example, monoclonal, human, humanized, camelized, and chimeric antibodies. The antibody may be of any isotype (e.g., IgG, IgE, IgM, IgD, IgA, and IgY), class (e.g., IgG1, IgG2, IgG3, IgG4, IgA1, and IgA2), or subclass. Both the light and heavy chains are divided into regions of structural and functional homology.

[0050] The phrase "antibody fragment" as used herein refers to one or more portions of an antibody that retain the ability to specifically interact with an antigen (e.g., by binding, steric hindrance, stabilized spatial distribution). Examples of binding fragments include, but are not limited to, Fab fragments, a monovalent fragment consisting of the VL, VH, CL and CH1 domains; F(ab)2 fragments, a bivalent fragment containing two Fab fragments linked by a disulfide bridge at the hinge region; Fd fragments consisting of the VH and CH1 domains; Fv fragments consisting of the VL and VH domains of a single arm of an antibody; dAb fragments consisting of the VH domain (Ward et al., (1989) Nature 341:544-546); and isolated complementarity determining regions (CDRs). Furthermore, although the two domains of the Fv fragment, VL and VH, are encoded by separate genes, they can be linked using recombinant methods by a synthetic linker that allows them to form a single-chain protein in which the VL and VH regions pair to form a monovalent molecule (known as single-chain Fv (scFv); see, e.g., Bird et al., (1988) Science 242:423-426; and Huston et al., (1988) Proc. Natl. Acad. Sci. 85:5879-5883). Such single-chain antibodies are also encompassed within the term "antibody fragments". These antibody fragments are obtained using conventional techniques known to those of skill in the art, and the fragments are screened for utility in the same manner as intact antibodies. Antibody fragments can also be incorporated into single domain antibodies, maxibodies, minibodies, intrabodies, diabodies, triabodies, tetrabodies, v-NARs and bis-scFvs (see, e.g., Hollinger and Hudson, (2005) Nature Biotechnology 23:1126-1136). Antibody fragments can be grafted onto scaffolds based on polypeptides such as fibronectin type III (Fn3) (see U.S. Pat. No. 6,703,199, which describes fibronectin polypeptide monobodies).Antibody fragments can be assembled into single-chain molecules comprising a pair of tandem Fv segments (VH-CH1-VH-CH1) which, together with complementary light chain polypeptides, form a pair of antigen-binding sites (Zapata et al., (1995) Protein Eng. 8:1057-1062; and U.S. Pat. No. 5,641,870).

[0051] "Human antibodies" or "human antibody fragments", as used herein, are antibodies and antibody fragments having variable regions in which both the framework and CDR regions are derived from sequences of human origin. Human antibodies can also be derived from synthetic libraries or from transgenic mice (e.g., Xenomouse), provided that each system produces antibodies with variable regions in which both the framework and CDR regions are derived from sequences of human origin. Furthermore, if the antibody contains a constant region, the constant region is also derived from such sequences. Human origin includes, for example, antibodies containing human germline sequences or mutated versions of human germline sequences or consensus framework sequences derived from human framework sequence analysis, e.g., as described in Knappik et al., (2000) J Mol Biol 296:57-86).

[0052] The structure and location of immunoglobulin variable domains, e.g., CDRs, can be defined using well-known numbering schemes, such as the Kabat numbering scheme, the Chothia numbering scheme, or a combination of Kabat and Chothia (e.g., Sequences of Proteins of Immunological Interest, USDepartment of Health and Human Services (1991), eds. Kabat et al.; Lazikani et al., (1997) J. Mol. Bio. 273:927-948); Kabat et al., (1991) Sequences of Proteins of Immunological Interest, 5th edit., NIH Publication no. 91-3242 USDepartment of Health and Human Services; Chothia et al., (1987) J. Mol. Biol. 196:901-917; Chothia et al., (1989) Nature 342:877-883; and Al-Lazikani et al. See al., (1997) J. Mol. Biol. 273:927-948.

[0053] A "humanized antibody" or "humanized antibody fragment" is defined herein as an antibody molecule having constant antibody regions derived from sequences of human origin, and only the variable antibody regions or parts thereof, or the CDRs, are derived from another species. For example, a humanized antibody can have CDRs grafted, in which the CDRs of the variable domain are of non-human origin, while one or more frameworks of the variable domain are of human origin, and the constant domains (if any) are of human origin.

[0054] The term "chimeric antibody" or "chimeric antibody fragment" is defined herein as an antibody molecule having constant antibody regions derived from or corresponding to sequences found in one species and variable antibody regions derived from another species. Preferably, the constant antibody regions are derived from or correspond to sequences found in humans and the variable antibody regions (e.g. VH, VL, CDR or FR regions) are derived from sequences found in a non-human animal, such as mouse, rat, rabbit or hamster.

[0055] The term "isolated antibody" refers to an antibody or antibody fragment that is substantially free of other antibodies or antibody fragments having different antigen specificities. Furthermore, an isolated antibody or antibody fragment may be substantially free of other cellular material and / or chemicals. Thus, in some embodiments, the antibody provided is an isolated antibody, which is separated from antibodies with different specificities. An isolated antibody may be a monoclonal antibody. An isolated antibody may be a recombinant monoclonal antibody. However, an isolated antibody that specifically binds to a target epitope, isoform, or variant may have cross-reactivity to other related antigens, such as those from other species (e.g., species homologs).

[0056] The term "recombinant antibody" as used herein includes all antibodies prepared, expressed, created, or isolated by means not occurring in nature, such as antibodies isolated from a host cell transformed to express the antibody, antibodies selected and isolated from a recombinant, combinatorial human antibody library, and antibodies prepared, expressed, created, or isolated by any other means including splicing of all or part of a human immunoglobulin gene, sequence to other DNA sequences, or antibodies isolated from an animal (e.g., a mouse) that is transgenic or transchromosomal for human immunoglobulin genes, or hybridomas prepared therefrom. Preferably, such recombinant antibodies have variable regions in which the framework and CDR regions are derived from human germline immunoglobulin sequences. However, in certain embodiments, such recombinant human antibodies may be subjected to in vitro mutagenesis (or in vivo somatic mutagenesis when animals transgenic for human Ig sequences are used) such that the amino acid sequences of the VH and VL regions of the recombinant antibodies, while derived from and related to human germline VH and VL sequences, are sequences that may not naturally occur within the human antibody germline repertoire in vivo. The recombinant antibodies may be monoclonal antibodies. In one embodiment, the antibodies and antibody fragments disclosed herein are isolated from a HuCAL library (Rothe et al., J. Mol. Biol. (2008) 376, 1182-1200).

[0057] As used herein, binding specificity is not absolute, but a relative property, when such an antibody can distinguish such an antigen from one or more reference antigens, so that an antibody "binds specifically to, specifically binds to" is "specific for" or "specifically recognizes" an antigen, such as human C5aR. For example, a standard ELISA assay can be performed. Scoring can be performed by standard color development (e.g. secondary antibody with horseradish peroxidase and tetramethylbenzidine with hydrogen peroxide). The reaction is scored by optical density, e.g. 450 nm, in a given well. A typical background (=negative reaction) can be 0.1 OD; a typical positive reaction can be 1 OD. This means that the difference between positive / negative can be more than 10-fold. Generally, the determination of binding specificity is performed by using a series of about 3-5 unrelated antigens, such as milk powder, BSA, transferrin, etc., rather than a single reference antigen.

[0058] As used herein, the term "affinity" refers to the strength of interaction between a polypeptide and its target at a single site. Within each site, the binding region of the polypeptide interacts with its target at many sites through weak non-covalent forces; the more interactions, the stronger the affinity.

[0059] "K D " as used herein refers to the dissociation constant, which is K on k for off (i.e., k off / k on ), expressed as a molar concentration (M). For example, the K D Values ​​can be determined using methods well established in the art, e.g., the K DMethods for determining C5aR are SET (solution equilibrium titration) or surface plasmon resonance using a biosensor system such as the Biacore® system. In the present disclosure, antibodies specific for C5aR are generally in the range of 5×10 -2 Less than M, 10 -2 Less than M, 5×10 -3 Less than M, 10 -3 Less than M, 5×10 -4 Less than M, 10 -4 Less than M, 5×10 -5 Less than M, 10 -5 Less than M, 5×10 -6 Less than M, 10 -6 Less than M, 5×10 -7 Less than M, 10 -7 Less than M, 5×10 -8 Less than M, 10 -8 Less than M, 5×10 -9 Less than M, 10 -9 Less than M, 5×10 -10 Less than M, 10 -10 Less than M, 5×10 -11 Less than M, 10 -11 Less than M, 5×10 -12 Less than M, 10 -12 Less than M, 5×0 -13 Less than M, 10 -13 Less than M, 5×10 -14 Less than M, 10 -14 Less than M, 5×10 -15 Less than M or 10 -15 Dissociation rate constant (K D )(k off / k on ).

[0060] The term "epitope" includes any proteinaceous region that is specifically recognized by an antibody or antibody fragment thereof or that otherwise interacts with a molecule. Generally, epitopes are chemically active surface groupings of molecules such as amino acids or carbohydrate or sugar side chains and generally may have specific three-dimensional structural characteristics as well as specific charge characteristics. As will be recognized by one of skill in the art, virtually anything to which an antibody specifically binds may be an epitope.

[0061] The "compositions" of the present disclosure may be used for therapeutic or prophylactic applications. Thus, the present disclosure includes pharmaceutical compositions containing an antibody or antibody fragment as disclosed herein and a pharma- ceutically acceptable carrier or excipient therefor. In a related aspect, the present disclosure provides methods for treating cancer. Such methods include administering to a subject in need thereof an effective amount of a pharmaceutical composition containing an antibody or antibody fragment as described herein.

[0062] The present disclosure provides a method of treatment comprising the administration of a therapeutically effective amount of an antibody or antibody fragment as disclosed herein to a subject in need of such treatment. "Therapeutically effective amount" or "effective amount" as used herein refers to the amount of C5aR antibody necessary to induce a desired biological response. According to the present disclosure, a therapeutically effective amount is the amount of C5aR antibody necessary to treat and / or prevent a disease.

[0063] "Administered" or "administration" includes, but is not limited to, delivery of a drug in an injectable form, such as by intravenous, intramuscular, intradermal or subcutaneous routes, or mucosal routes, such as as a nasal spray or aerosol for inhalation, or as an ingestible solution, capsule or tablet. Preferably, administration is by an injectable form.

[0064] As used herein, "treatment," "treat," or "treating," and the like, refer to clinical intervention in an attempt to alter the natural course of a disease in the subject being treated, and may be performed either prophylactically or during the course of clinical disease. 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 consequence of the disease, preventing metastasis, slowing the rate of disease progression, remission or alleviation of the disease state, and ameliorating or improving prognosis. In some embodiments, an antibody or antibody fragment according to the present disclosure is used to delay disease onset or slow disease progression.

[0065] The term "effector function" refers to a biological activity attributable to the Fc region of an antibody, which varies with antibody isotype. Non-limiting examples of antibody effector functions include C1q binding and complement-dependent cytotoxicity (CDC); Fc receptor binding and antibody-dependent cell-mediated cytotoxicity (ADCC) and / or antibody-dependent cellular phagocytosis (ADCP); downregulation of cell surface receptors (e.g., B cell receptors); and B cell activation.

[0066] "Antibody-dependent cell-mediated cytotoxicity" or "ADCC" refers to a form of cytotoxicity in which antibodies bound to Fc receptors (FcR) present on certain cytotoxic cells (e.g., NK cells, neutrophils, and macrophages) enable these cytotoxic effector cells to specifically bind to antigen-bearing target cells and subsequently cytotoxically kill the target cell. NK cells, the primary cells for mediating ADCC, express only FcγRIII, whereas monocytes express FcγRI, FcγRII, and FcγRIII.

[0067] "Complement dependent cytotoxicity" or "CDC" refers to the lysis of a target cell in the presence of complement. Activation of the classical complement pathway is initiated by the binding of the first component of the complement system (C1q) to antibodies (of the appropriate subclass) of the present disclosure that are bound to their cognate antigen.

[0068] "Antibody-dependent cellular phagocytosis" or "ADCP" refers to the mechanism of elimination of antibody-coated target cells by internalization by phagocytic cells, such as macrophages or dendritic cells.

[0069] "Preventing" or "prevention" refers to reducing the risk of acquiring or developing a disease (i.e., preventing at least one clinical symptom of a disease from developing in a subject who may be exposed to a disease-causing agent or who is susceptible to the disease prior to disease onset). "Prevention" also refers to methods aimed at preventing or delaying the onset of a disease or its symptoms.

[0070] "Subject" or "species" or as used in this context refers to any mammal, including rodents, such as mice or rats, and primates, such as cynomolgus monkeys (Macaca fascicularis), rhesus monkeys (Macaca mulatta), or humans (Homo sapiens). Preferably, the subject is a primate, most preferably a human.

[0071] Throughout this specification, unless the context requires otherwise, the words "comprise," "have," and "include," and individual variations thereof such as "comprises," "comprising," "has," "having," "includes," and "including," are understood to indicate the inclusion of a stated element or integer or group of elements or integers but not the exclusion of any other element or integer or group of elements or integers.

[0072] The terms "engineered" or "modified" as used herein include the manipulation of nucleic acids or polypeptides by synthetic means (e.g., by recombinant techniques, by in vitro peptide synthesis, by enzymatic or chemical coupling of peptides, or by some combination of these techniques). Preferably, an antibody or antibody fragment according to the present disclosure is engineered or modified to improve one or more properties, such as antigen binding, stability, half-life, effector functions, immunogenicity, safety, etc.

[0073] "Variant," as used herein, refers to a polypeptide that differs from a reference polypeptide by one or more modifications, such as an amino acid substitution, insertion, or deletion.

[0074] The term "amino acid mutation" as used herein is meant to encompass amino acid substitution, deletion, insertion and modification. Any combination of substitution, deletion, insertion and modification may be made as long as the final construct retains the desired characteristics, such as reduced binding to Fc receptors. Deletion and insertion of amino acid sequences include N- and / or C-terminal deletion and insertion of amino acid residues. A specific amino acid mutation is an amino acid substitution. Amino acid substitution includes replacement with unnatural amino acids or with natural amino acid derivatives of the 20 standard amino acids. Amino acid mutations may be made using genetic or chemical methods well known in the art. Genetic methods may include site-directed mutagenesis, PCR, gene synthesis, and the like. It is contemplated that methods of changing the side chain group of an amino acid residue by methods other than genetic modification, such as chemical modification, may also be useful. Various nomenclature systems may be used herein to indicate the same amino acid mutation. For example, a glycine to alanine substitution at position 327 of the Fc region may be indicated as 237A, G337, G337A, or Gly329Ala.

[0075] "EC 50The term "as used herein" refers to the concentration of antibody or antibody fragment or ligand that induces a response in the assay midway between baseline and maximum. It therefore corresponds to the antibody or ligand concentration at which 50% of the maximum effect is observed.

[0076] "I C 50 The term "antibody concentration" as used herein refers to a concentration of antibody or antibody fragment that inhibits a response in an assay midway between the maximum response and the baseline. This corresponds to the antibody concentration at which the given response is reduced by 50%.

[0077] The term "inhibition" or "inhibit" or "reduction" or "reduce" or "neutralization" or "neutralize" refers to the attenuation or cessation of any phenotypic characteristic (such as binding or biological activity or function) or the reduction or cessation of the incidence, extent, or likelihood of that characteristic. "Inhibition", "reduction" or "neutralization" need not be complete, as long as it is detectable using an appropriate assay. In some embodiments, "reduce" or "inhibit" or "neutralize" refers to the ability to cause a reduction of 20% or more. In another embodiment, "reduce" or "inhibit" or "neutralize" refers to the ability to cause a reduction of 50% or more. In yet another embodiment, "reduce" or "inhibit" or "neutralize" refers to the ability to cause an overall reduction of 75%, 85%, 90%, 95% or more.

[0078] The term "antagonist" antibody, as used herein, refers to an antibody or antibody fragment that interacts with an antigen and partially or completely inhibits or neutralizes the biological activity or function or some other phenotypic characteristic of the target antigen.

[0079] A "wild type" protein is a version or variant of a protein as it is found in nature. The amino acid sequence of a wild type protein, such as the Fc region of a human IgG1 antibody, is the amino acid sequence of the protein as it occurs in nature. Due to allotypic differences, there may be multiple amino acid sequences for the wild type protein. For example, there are several allotypes of the naturally occurring human IGg1 heavy chain constant region (see, e.g., Jeffries et al. (2009) mAbs 1:1).

[0080] "Fc region" is used to define the C-terminal region of an immunoglobulin heavy chain. The Fc region of an immunoglobulin generally comprises two constant domains, a CH2 domain and a CH3 domain. Although the boundaries of the Fc region of an IgG heavy chain may vary slightly, the human IgG heavy chain Fc region is usually defined as extending from Cys226, or from Pro230, to the C-terminus of the heavy chain. However, the C-terminal lysine (Lys447) of the Fc region may or may not be present. Unless otherwise specified herein, the numbering of amino acid residues in the Fc region follows the EU numbering system, also called the EU index, as described in Kabat et al., Sequences of Proteins of Immunological Interest, 5th Ed. Public Health Service, National Institutes of Health, Bethesda, MD, 1991.

[0081] Embodiments: In one embodiment, the present disclosure refers to an isolated antibody or antibody fragment specific for human C5aR, said antibody or antibody fragment comprising: a) an HCDR1 region comprising the amino acid sequence of SEQ ID NO: 27, an HCDR2 region comprising the amino acid sequence of SEQ ID NO: 28, an HCDR3 region comprising the amino acid sequence of SEQ ID NO: 29, an LCDR1 region comprising the amino acid sequence of SEQ ID NO: 32, an LCDR2 region comprising the amino acid sequence of SEQ ID NO: 33, and an LCDR3 region comprising the amino acid sequence of SEQ ID NO: 34, or b) an HCDR1 region comprising the amino acid sequence of SEQ ID NO: 27, an HCDR2 region comprising the amino acid sequence of SEQ ID NO: 39, an HCDR3 region comprising the amino acid sequence of SEQ ID NO: 40, an LCDR1 region comprising the amino acid sequence of SEQ ID NO: 32, an LCDR2 region comprising the amino acid sequence of SEQ ID NO: 33, and an LCDR3 region comprising the amino acid sequence of SEQ ID NO: 34 Includes.

[0082] In one embodiment, the present disclosure refers to an isolated antibody or antibody fragment specific for human C5aR, said antibody or antibody fragment comprising: a) the HCDR1 region of SEQ ID NO: 27, the HCDR2 region of SEQ ID NO: 28, the HCDR3 region of SEQ ID NO: 29, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33 and the LCDR3 region of SEQ ID NO: 34, or b) the HCDR1 region of SEQ ID NO: 27, the HCDR2 region of SEQ ID NO: 39, the HCDR3 region of SEQ ID NO: 40, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33 and the LCDR3 region of SEQ ID NO: 34 Includes.

[0083] In one embodiment, the present disclosure refers to an isolated antibody or antibody fragment specific for human C5aR, said antibody or antibody fragment comprising: a) an HCDR1 region comprising the amino acid sequence of SEQ ID NO: 30, an HCDR2 region comprising the amino acid sequence of SEQ ID NO: 31, an HCDR3 region comprising the amino acid sequence of SEQ ID NO: 29, an LCDR1 region comprising the amino acid sequence of SEQ ID NO: 32, an LCDR2 region comprising the amino acid sequence of SEQ ID NO: 33, and an LCDR3 region comprising the amino acid sequence of SEQ ID NO: 34, or b) an HCDR1 region comprising the amino acid sequence of SEQ ID NO: 30, an HCDR2 region comprising the amino acid sequence of SEQ ID NO: 41, an HCDR3 region comprising the amino acid sequence of SEQ ID NO: 40, an LCDR1 region comprising the amino acid sequence of SEQ ID NO: 32, an LCDR2 region comprising the amino acid sequence of SEQ ID NO: 33, and an LCDR3 region comprising the amino acid sequence of SEQ ID NO: 34 Includes.

[0084] In one embodiment, the present disclosure refers to an isolated antibody or antibody fragment specific for human C5aR, said antibody or antibody fragment comprising: a) the HCDR1 region of SEQ ID NO: 30, the HCDR2 region of SEQ ID NO: 31, the HCDR3 region of SEQ ID NO: 29, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33 and the LCDR3 region of SEQ ID NO: 34, or b) the HCDR1 region of SEQ ID NO: 30, the HCDR2 region of SEQ ID NO: 41, the HCDR3 region of SEQ ID NO: 40, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33 and the LCDR3 region of SEQ ID NO: 34 Includes.

[0085] In one embodiment, the present disclosure refers to an isolated antibody or antibody fragment specific for human C5aR, said antibody or antibody fragment comprising: a) an HCDR1 region comprising the amino acid sequence of SEQ ID NO: 27, an HCDR2 region comprising the amino acid sequence of SEQ ID NO: 28, an HCDR3 region comprising the amino acid sequence of SEQ ID NO: 29, an LCDR1 region comprising the amino acid sequence of SEQ ID NO: 32, an LCDR2 region comprising the amino acid sequence of SEQ ID NO: 33, and an LCDR3 region comprising the amino acid sequence of SEQ ID NO: 34, or b) an HCDR1 region comprising the amino acid sequence of SEQ ID NO: 30, an HCDR2 region comprising the amino acid sequence of SEQ ID NO: 31, an HCDR3 region comprising the amino acid sequence of SEQ ID NO: 29, an LCDR1 region comprising the amino acid sequence of SEQ ID NO: 32, an LCDR2 region comprising the amino acid sequence of SEQ ID NO: 33 and an LCDR3 region comprising the amino acid sequence of SEQ ID NO: 34, or c) an HCDR1 region comprising the amino acid sequence of SEQ ID NO: 27, an HCDR2 region comprising the amino acid sequence of SEQ ID NO: 39, an HCDR3 region comprising the amino acid sequence of SEQ ID NO: 40, an LCDR1 region comprising the amino acid sequence of SEQ ID NO: 32, an LCDR2 region comprising the amino acid sequence of SEQ ID NO: 33, and an LCDR3 region comprising the amino acid sequence of SEQ ID NO: 34, or d) an HCDR1 region comprising the amino acid sequence of SEQ ID NO: 30, an HCDR2 region comprising the amino acid sequence of SEQ ID NO: 41, an HCDR3 region comprising the amino acid sequence of SEQ ID NO: 40, an LCDR1 region comprising the amino acid sequence of SEQ ID NO: 32, an LCDR2 region comprising the amino acid sequence of SEQ ID NO: 33, and an LCDR3 region comprising the amino acid sequence of SEQ ID NO: 34 Includes.

[0086] In one embodiment, the present disclosure refers to an isolated antibody or antibody fragment specific for human C5aR, said antibody or antibody fragment comprising: a) the HCDR1 region of SEQ ID NO: 27, the HCDR2 region of SEQ ID NO: 28, the HCDR3 region of SEQ ID NO: 29, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33 and the LCDR3 region of SEQ ID NO: 34, or b) the HCDR1 region of SEQ ID NO: 30, the HCDR2 region of SEQ ID NO: 31, the HCDR3 region of SEQ ID NO: 29, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33 and the LCDR3 region of SEQ ID NO: 34, or c) the HCDR1 region of SEQ ID NO: 27, the HCDR2 region of SEQ ID NO: 39, the HCDR3 region of SEQ ID NO: 40, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33 and the LCDR3 region of SEQ ID NO: 34, or d) the HCDR1 region of SEQ ID NO: 30, the HCDR2 region of SEQ ID NO: 41, the HCDR3 region of SEQ ID NO: 40, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33, and the LCDR3 region of SEQ ID NO: 34 Includes.

[0087] In one embodiment, the present disclosure refers to an isolated antibody or antibody fragment specific for human C5aR, said antibody or antibody fragment comprising an HCDR1 region of SEQ ID NO: 27, an HCDR2 region of SEQ ID NO: 28, an HCDR3 region of SEQ ID NO: 29, an LCDR1 region of SEQ ID NO: 32, an LCDR2 region of SEQ ID NO: 33 and an LCDR3 region of SEQ ID NO: 34.

[0088] In one embodiment, the present disclosure refers to an isolated antibody or antibody fragment specific for human C5aR, said antibody or antibody fragment comprising an HCDR1 region of SEQ ID NO: 27, an HCDR2 region of SEQ ID NO: 39, an HCDR3 region of SEQ ID NO: 40, an LCDR1 region of SEQ ID NO: 32, an LCDR2 region of SEQ ID NO: 33 and an LCDR3 region of SEQ ID NO: 34.

[0089] In one embodiment, the present disclosure refers to an isolated antibody or antibody fragment specific for human C5aR, said antibody or antibody fragment comprising an HCDR1 region of SEQ ID NO: 30, an HCDR2 region of SEQ ID NO: 31, an HCDR3 region of SEQ ID NO: 29, an LCDR1 region of SEQ ID NO: 32, an LCDR2 region of SEQ ID NO: 33 and an LCDR3 region of SEQ ID NO: 34.

[0090] In one embodiment, the present disclosure refers to an isolated antibody or antibody fragment specific for human C5aR, said antibody or antibody fragment comprising an HCDR1 region of SEQ ID NO: 30, an HCDR2 region of SEQ ID NO: 41, an HCDR3 region of SEQ ID NO: 40, an LCDR1 region of SEQ ID NO: 32, an LCDR2 region of SEQ ID NO: 33 and an LCDR3 region of SEQ ID NO: 34.

[0091] In one embodiment, the disclosure refers to an isolated antibody or antibody fragment specific for human C5aR that comprises the six CDRs as defined by Kabat of any one of the antibodies disclosed in Table 1 or Table 2.

[0092] In one embodiment, the present disclosure refers to an isolated antibody or antibody fragment specific for human C5aR that contains the six Chothia-defined CDRs of any one of the antibodies disclosed in Table 1 or Table 2.

[0093] In one embodiment of the disclosure, the isolated antibody or antibody fragment is a monoclonal antibody or antibody fragment. In one embodiment of the disclosure, the isolated antibody or antibody fragment is a human, humanized or chimeric antibody or antibody fragment. In another embodiment of the disclosure, the isolated antibody or antibody fragment is a recombinant antibody or antibody fragment. In one embodiment of the disclosure, the isolated antibody or antibody fragment is an IgG isotype. In one embodiment of the disclosure, the isolated antibody or antibody fragment is an IgG1 class. In another embodiment of the disclosure, the isolated antibody or antibody fragment induces substantially no effector function in vitro.

[0094] In one embodiment, the present disclosure refers to an isolated antibody or antibody fragment specific for human C5aR, said antibody or antibody fragment comprising: a) comprising an HCDR1 region of SEQ ID NO: 27, an HCDR2 region of SEQ ID NO: 28, an HCDR3 region of SEQ ID NO: 29, an LCDR1 region of SEQ ID NO: 32, an LCDR2 region of SEQ ID NO: 33 and an LCDR3 region of SEQ ID NO: 34, and further comprising a VH of SEQ ID NO: 35 or a VL of SEQ ID NO: 36; or b) comprising an HCDR1 region of SEQ ID NO: 30, an HCDR2 region of SEQ ID NO: 31, an HCDR3 region of SEQ ID NO: 29, an LCDR1 region of SEQ ID NO: 32, an LCDR2 region of SEQ ID NO: 33 and an LCDR3 region of SEQ ID NO: 34, and further comprising a VH of SEQ ID NO: 35 or a VL of SEQ ID NO: 36; or c) comprising an HCDR1 region of SEQ ID NO: 27, an HCDR2 region of SEQ ID NO: 39, an HCDR3 region of SEQ ID NO: 40, an LCDR1 region of SEQ ID NO: 32, an LCDR2 region of SEQ ID NO: 33 and an LCDR3 region of SEQ ID NO: 34, and further comprising a VH of SEQ ID NO: 42 or a VL of SEQ ID NO: 43; or d) comprising an HCDR1 region of SEQ ID NO: 30, an HCDR2 region of SEQ ID NO: 41, an HCDR3 region of SEQ ID NO: 40, an LCDR1 region of SEQ ID NO: 32, an LCDR2 region of SEQ ID NO: 33 and an LCDR3 region of SEQ ID NO: 34, and further comprising a VH of SEQ ID NO: 42 or a VL of SEQ ID NO: 43.

[0095] In one embodiment, the present disclosure refers to an isolated antibody or antibody fragment specific for human C5aR, said antibody or antibody fragment comprising: a) a VH of SEQ ID NO: 35 and a VL of SEQ ID NO: 36, or b) VH of SEQ ID NO: 42 and VL of SEQ ID NO: 43 Includes.

[0096] In one embodiment, the disclosure refers to an isolated antibody or antibody fragment specific for human C5aR, said antibody or antibody fragment comprising a VH of SEQ ID NO:35 and a VL of SEQ ID NO:36.

[0097] In one embodiment, the disclosure refers to an isolated antibody or antibody fragment specific for human C5aR, said antibody or antibody fragment comprising a VH of SEQ ID NO:42 and a VL of SEQ ID NO:43.

[0098] In one embodiment, the disclosure refers to an isolated antibody or antibody fragment specific for human C5aR, said antibody or antibody fragment comprising a HC of SEQ ID NO:37 and a LC of SEQ ID NO:38.

[0099] In one embodiment, the disclosure refers to an isolated antibody or antibody fragment specific for human C5aR, said antibody or antibody fragment comprising a HC of SEQ ID NO:44 and a LC of SEQ ID NO:45.

[0100] In one embodiment, the disclosure refers to an isolated antibody or antibody fragment specific for human C5aR comprising the variable heavy chain (VH) and variable light chain (VL) of any one of the antibodies disclosed in Table 1 or Table 2.

[0101] In one embodiment, the disclosure refers to an isolated antibody or antibody fragment specific for human C5aR comprising the heavy chain (HC) and light chain (LC) of any one of the antibodies disclosed in Table 1 or Table 2.

[0102] In one embodiment, the present disclosure refers to an isolated antibody or antibody fragment specific for human C5aR, said antibody or antibody fragment comprising an HCDR1 region of SEQ ID NO:27, an HCDR2 region of SEQ ID NO:28, an HCDR3 region of SEQ ID NO:29, an LCDR1 region of SEQ ID NO:32, an LCDR2 region of SEQ ID NO:33 and an LCDR3 region of SEQ ID NO:34, and further comprising a VH of SEQ ID NO:35 or a VL of SEQ ID NO:36.

[0103] In one embodiment, the present disclosure refers to an isolated antibody or antibody fragment specific for human C5aR, said antibody or antibody fragment comprising an HCDR1 region of SEQ ID NO:27, an HCDR2 region of SEQ ID NO:39, an HCDR3 region of SEQ ID NO:40, an LCDR1 region of SEQ ID NO:32, an LCDR2 region of SEQ ID NO:33 and an LCDR3 region of SEQ ID NO:34, and further comprising a VH of SEQ ID NO:42 or a VL of SEQ ID NO:43.

[0104] In one embodiment, the present disclosure refers to an isolated antibody or antibody fragment specific for human C5aR, said antibody or antibody fragment comprising an HCDR1 region of SEQ ID NO: 30, an HCDR2 region of SEQ ID NO: 31, an HCDR3 region of SEQ ID NO: 29, an LCDR1 region of SEQ ID NO: 32, an LCDR2 region of SEQ ID NO: 33 and an LCDR3 region of SEQ ID NO: 34, and further comprising a VH of SEQ ID NO: 35 or a VL of SEQ ID NO: 36.

[0105] In one embodiment, the present disclosure refers to an isolated antibody or antibody fragment specific for human C5aR, said antibody or antibody fragment comprising an HCDR1 region of SEQ ID NO: 30, an HCDR2 region of SEQ ID NO: 41, an HCDR3 region of SEQ ID NO: 40, an LCDR1 region of SEQ ID NO: 32, an LCDR2 region of SEQ ID NO: 33 and an LCDR3 region of SEQ ID NO: 34, and further comprising a VH of SEQ ID NO: 42 or a VL of SEQ ID NO: 43.

[0106] In one embodiment of the disclosure, the isolated antibody or antibody fragment is a monoclonal antibody or antibody fragment. In one embodiment of the disclosure, the isolated antibody or antibody fragment is a human, humanized or chimeric antibody or antibody fragment. In another embodiment of the disclosure, the isolated antibody or antibody fragment is a recombinant antibody or antibody fragment.

[0107] In one embodiment, the present disclosure refers to an isolated antibody or antibody fragment specific for human C5aR, said antibody or antibody fragment comprising: a) a VH of SEQ ID NO: 35 and a VL of SEQ ID NO: 36, or b) a VH of SEQ ID NO: 42 and a VL of SEQ ID NO: 43, or VH and VL having at least 80%, at least 85%, at least 90% or at least 95% identity to VH of SEQ ID NO: 35 or SEQ ID NO: 42 and VL of SEQ ID NO: 36 or SEQ ID NO: 43 Includes.

[0108] In one embodiment, the present disclosure refers to an isolated antibody or antibody fragment specific for human C5aR, said antibody or antibody fragment comprising a VH of SEQ ID NO: 35 and a VL of SEQ ID NO: 36, or a VH and a VL having at least 80%, at least 85%, at least 90% or at least 95% identity to the VH of SEQ ID NO: 35 and to the VL of SEQ ID NO: 36.

[0109] In one embodiment, the present disclosure refers to an isolated antibody or antibody fragment specific for human C5aR, said antibody or antibody fragment comprising a VH of SEQ ID NO: 42 and a VL of SEQ ID NO: 43, or a VH and a VL having at least 80%, at least 85%, at least 90% or at least 95% identity to the VH of SEQ ID NO: 42 and to the VL of SEQ ID NO: 43.

[0110] In one embodiment, the present disclosure refers to an isolated antibody or antibody fragment specific for human C5aR, said antibody or antibody fragment comprising: a) HC of SEQ ID NO: 37 and LC of SEQ ID NO: 38, or b) HC of SEQ ID NO: 44 and LC of SEQ ID NO: 45, or HC and LC having at least 80%, at least 85%, at least 90%, or at least 95% identity to the HC of SEQ ID NO:37 or SEQ ID NO:44 and to the LC of SEQ ID NO:38 or SEQ ID NO:45. Includes.

[0111] In one embodiment, the present disclosure refers to an isolated antibody or antibody fragment specific for human C5aR, said antibody or antibody fragment comprising a HC of SEQ ID NO: 37 and a LC of SEQ ID NO: 38, or a HC and LC having at least 80%, at least 85%, at least 90% or at least 95% identity to the HC of SEQ ID NO: 37 and to the LC of SEQ ID NO: 38.

[0112] In one embodiment, the present disclosure refers to an isolated antibody or antibody fragment specific for human C5aR, said antibody or antibody fragment comprising a HC of SEQ ID NO: 44 and a LC of SEQ ID NO: 45, or a HC and LC having at least 80%, at least 85%, at least 90% or at least 95% identity to the HC of SEQ ID NO: 44 and to the LC of SEQ ID NO: 45.

[0113] In one embodiment of the disclosure, the isolated antibody or antibody fragment is a monoclonal antibody or antibody fragment. In one embodiment of the disclosure, the isolated antibody or antibody fragment is a human, humanized or chimeric antibody or antibody fragment. In another embodiment of the disclosure, the isolated antibody or antibody fragment is a recombinant antibody or antibody fragment. In one embodiment, the isolated antibody or antibody fragment is a human or humanized antibody or antibody fragment. In one embodiment, the isolated antibody or antibody fragment is a human antibody or antibody fragment.

[0114] In one embodiment, the isolated antibody or antibody fragment specific for human C5aR of the present disclosure is a recombinant or synthetic antibody or antibody fragment. In a further embodiment, the isolated antibody or antibody fragment specific for C5aR according to the present disclosure is an isolated recombinant monoclonal antibody or antibody fragment. In a further embodiment, the isolated antibody or antibody fragment specific for C5aR according to the present disclosure is an isolated recombinant monoclonal human antibody or antibody fragment.

[0115] In one embodiment, the isolated antibody or antibody fragment specific for C5aR according to the present disclosure is of the IgG isotype. In another embodiment, the isolated antibody or antibody fragment is of the IgG1 class. In another embodiment, the isolated antibody or antibody fragment is of the human IgG1 class.

[0116] nucleic acid In one embodiment, the present disclosure refers to a nucleic acid composition comprising a nucleic acid sequence or a plurality of nucleic acid sequences encoding an isolated antibody or antibody fragment specific for human C5aR, said isolated antibody or antibody fragment comprising: a) the HCDR1 region of SEQ ID NO: 27, the HCDR2 region of SEQ ID NO: 28, the HCDR3 region of SEQ ID NO: 29, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33 and the LCDR3 region of SEQ ID NO: 34, or b) the HCDR1 region of SEQ ID NO: 30, the HCDR2 region of SEQ ID NO: 31, the HCDR3 region of SEQ ID NO: 29, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33 and the LCDR3 region of SEQ ID NO: 34, or c) the HCDR1 region of SEQ ID NO: 27, the HCDR2 region of SEQ ID NO: 39, the HCDR3 region of SEQ ID NO: 40, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33 and the LCDR3 region of SEQ ID NO: 34, or d) the HCDR1 region of SEQ ID NO: 30, the HCDR2 region of SEQ ID NO: 41, the HCDR3 region of SEQ ID NO: 40, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33, and the LCDR3 region of SEQ ID NO: 34 Includes.

[0117] In another embodiment, the present disclosure refers to a nucleic acid composition comprising a nucleic acid sequence or sequences encoding heavy and light chain sequences of an isolated antibody or antibody fragment specific for human C5aR, said nucleic acid sequence or sequences comprising: a) the HCDR1 region of SEQ ID NO: 46, the HCDR2 region of SEQ ID NO: 47, the HCDR3 region of SEQ ID NO: 48, the LCDR1 region of SEQ ID NO: 51, the LCDR2 region of SEQ ID NO: 52 and the LCDR3 region of SEQ ID NO: 53, or b) an HCDR1 region of SEQ ID NO: 49, an HCDR2 region of SEQ ID NO: 50, an HCDR3 region of SEQ ID NO: 48, an LCDR1 region of SEQ ID NO: 51, an LCDR2 region of SEQ ID NO: 52 and an LCDR3 region of SEQ ID NO: 53, or c) the HCDR1 region of SEQ ID NO: 58, the HCDR2 region of SEQ ID NO: 59, the HCDR3 region of SEQ ID NO: 60, the LCDR1 region of SEQ ID NO: 63, the LCDR2 region of SEQ ID NO: 64 and the LCDR3 region of SEQ ID NO: 65, or d) the HCDR1 region of SEQ ID NO: 61, the HCDR2 region of SEQ ID NO: 62, the HCDR3 region of SEQ ID NO: 60, the LCDR1 region of SEQ ID NO: 63, the LCDR2 region of SEQ ID NO: 64 and the LCDR3 region of SEQ ID NO: 65 Includes.

[0118] In another embodiment, the present disclosure refers to a nucleic acid composition comprising a nucleic acid sequence or sequences encoding heavy and light chain sequences of an isolated antibody or antibody fragment specific for human C5aR, said nucleic acid sequence or sequences comprising: a) the HCDR1 region of SEQ ID NO: 70, the HCDR2 region of SEQ ID NO: 71, the HCDR3 region of SEQ ID NO: 72, the LCDR1 region of SEQ ID NO: 75, the LCDR2 region of SEQ ID NO: 76 and the LCDR3 region of SEQ ID NO: 77, or b) the HCDR1 region of SEQ ID NO: 73, the HCDR2 region of SEQ ID NO: 74, the HCDR3 region of SEQ ID NO: 72, the LCDR1 region of SEQ ID NO: 75, the LCDR2 region of SEQ ID NO: 76 and the LCDR3 region of SEQ ID NO: 77, or c) the HCDR1 region of SEQ ID NO: 82, the HCDR2 region of SEQ ID NO: 83, the HCDR3 region of SEQ ID NO: 84, the LCDR1 region of SEQ ID NO: 87, the LCDR2 region of SEQ ID NO: 88 and the LCDR3 region of SEQ ID NO: 89, or d) the HCDR1 region of SEQ ID NO: 85, the HCDR2 region of SEQ ID NO: 86, the HCDR3 region of SEQ ID NO: 84, the LCDR1 region of SEQ ID NO: 87, the LCDR2 region of SEQ ID NO: 88 and the LCDR3 region of SEQ ID NO: 89 Includes.

[0119] In one embodiment, the present disclosure refers to a nucleic acid composition comprising a nucleic acid sequence or a plurality of nucleic acid sequences encoding an isolated antibody or antibody fragment specific for human C5aR, wherein the nucleic acid sequence or a plurality of nucleic acid sequences comprises a VH of SEQ ID NO: 54 and a VL of SEQ ID NO: 55, or a VH and a VL having at least 60%, at least 70%, at least 80%, at least 85%, at least 90% or at least 95% identity to the VH of SEQ ID NO: 54 and / or the VL of SEQ ID NO: 55.

[0120] In one embodiment, the present disclosure refers to a nucleic acid composition comprising a nucleic acid sequence or a plurality of nucleic acid sequences encoding an isolated antibody or antibody fragment specific for human C5aR, wherein the nucleic acid sequence or a plurality of nucleic acid sequences comprises a VH of SEQ ID NO: 66 and a VL of SEQ ID NO: 67, or a VH and a VL having at least 60%, at least 70%, at least 80%, at least 85%, at least 90% or at least 95% identity to the VH of SEQ ID NO: 66 and / or the VL of SEQ ID NO: 67.

[0121] In one embodiment, the present disclosure refers to a nucleic acid composition comprising a nucleic acid sequence or a plurality of nucleic acid sequences encoding an isolated antibody or antibody fragment specific for human C5aR, wherein the nucleic acid sequence or a plurality of nucleic acid sequences comprises a VH of SEQ ID NO: 78 and a VL of SEQ ID NO: 79, or a VH and a VL having at least 60%, at least 70%, at least 80%, at least 85%, at least 90% or at least 95% identity to the VH of SEQ ID NO: 78 and / or the VL of SEQ ID NO: 79.

[0122] In one embodiment, the present disclosure refers to a nucleic acid composition comprising a nucleic acid sequence or a plurality of nucleic acid sequences encoding an isolated antibody or antibody fragment specific for human C5aR, wherein the nucleic acid sequence or a plurality of nucleic acid sequences comprises a VH of SEQ ID NO: 90 and a VL of SEQ ID NO: 91, or a VH and a VL having at least 60%, at least 70%, at least 80%, at least 85%, at least 90% or at least 95% identity to the VH of SEQ ID NO: 90 and / or the VL of SEQ ID NO: 91.

[0123] In one embodiment, the present disclosure refers to a nucleic acid composition comprising a nucleic acid sequence or a plurality of nucleic acid sequences encoding an isolated antibody or antibody fragment specific for human C5aR, wherein the nucleic acid sequence or a plurality of nucleic acid sequences comprises a HC of SEQ ID NO: 56 and a LC of SEQ ID NO: 57, or a HC and a LC having at least 60%, at least 70%, at least 80%, at least 85%, at least 90% or at least 95% identity to the HC of SEQ ID NO: 56 and / or the LC of SEQ ID NO: 57.

[0124] In one embodiment, the present disclosure refers to a nucleic acid composition comprising a nucleic acid sequence or a plurality of nucleic acid sequences encoding an isolated antibody or antibody fragment specific for human C5aR, wherein the nucleic acid sequence or a plurality of nucleic acid sequences comprises a HC of SEQ ID NO: 68 and a LC of SEQ ID NO: 69, or a HC and a LC having at least 60%, at least 70%, at least 80%, at least 85%, at least 90% or at least 95% identity to the HC of SEQ ID NO: 68 and / or the LC of SEQ ID NO: 69.

[0125] In one embodiment, the present disclosure refers to a nucleic acid composition comprising a nucleic acid sequence or a plurality of nucleic acid sequences encoding an isolated antibody or antibody fragment specific for human C5aR, wherein the nucleic acid sequence or a plurality of nucleic acid sequences comprises a HC of SEQ ID NO: 80 and a LC of SEQ ID NO: 81, or a HC and a LC having at least 60%, at least 70%, at least 80%, at least 85%, at least 90% or at least 95% identity to the HC of SEQ ID NO: 80 and / or the LC of SEQ ID NO: 81.

[0126] In one embodiment, the present disclosure refers to a nucleic acid composition comprising a nucleic acid sequence or a plurality of nucleic acid sequences encoding an isolated antibody or antibody fragment specific for human C5aR, wherein the nucleic acid sequence or a plurality of nucleic acid sequences comprises a HC of SEQ ID NO: 92 and a LC of SEQ ID NO: 93, or a HC and a LC having at least 60%, at least 70%, at least 80%, at least 85%, at least 90% or at least 95% identity to the HC of SEQ ID NO: 92 and / or the LC of SEQ ID NO: 93.

[0127] In one embodiment, the present disclosure refers to a nucleic acid composition comprising a nucleic acid sequence or a plurality of nucleic acid sequences encoding an isolated antibody or antibody fragment specific to human C5aR, wherein the nucleic acid sequence or a plurality of nucleic acid sequences comprises a HC of SEQ ID NO:56 and a LC of SEQ ID NO:57.

[0128] In one embodiment, the present disclosure refers to a nucleic acid composition comprising a nucleic acid sequence or a plurality of nucleic acid sequences encoding an isolated antibody or antibody fragment specific to human C5aR, wherein the nucleic acid sequence or a plurality of nucleic acid sequences comprises a HC of SEQ ID NO:68 and a LC of SEQ ID NO:69.

[0129] In one embodiment, the present disclosure refers to a nucleic acid composition comprising a nucleic acid sequence or a plurality of nucleic acid sequences encoding an isolated antibody or antibody fragment specific to human C5aR, wherein the nucleic acid sequence or plurality of nucleic acid sequences comprises a HC of SEQ ID NO: 80 and a LC of SEQ ID NO: 81.

[0130] In one embodiment, the present disclosure refers to a nucleic acid composition comprising a nucleic acid sequence or a plurality of nucleic acid sequences encoding an antibody or antibody fragment specific to human C5aR disclosed herein, wherein the nucleic acid sequence or a plurality of nucleic acid sequences comprises the HC of SEQ ID NO: 92 and the LC of SEQ ID NO: 93.

[0131] In another embodiment, the present disclosure refers to a nucleic acid composition comprising a nucleic acid sequence or a plurality of nucleic acid sequences encoding an isolated antibody or fragment specific to human C5aR, wherein the nucleic acid sequence or plurality of nucleic acid sequences comprises the VH and VL of any one of the antibodies disclosed in Table 1 or Table 2.

[0132] In one embodiment, the disclosure provides an isolated antibody or antibody fragment specific for human C5aR encoded by any one of the nucleic acid sequence or sequences disclosed in Table 1 or Table 2.

[0133] In one embodiment, the disclosure provides a nucleic acid composition comprising a nucleic acid sequence or sequences encoding any one of the isolated antibodies or antibody fragments disclosed in Table 1 or Table 2.

[0134] In one embodiment, the present disclosure provides a nucleic acid composition comprising a nucleic acid sequence or sequences encoding any one of the isolated antibodies or antibody fragments specific for human C5aR according to the present disclosure.

[0135] In another embodiment, the present disclosure refers to a nucleic acid composition comprising a nucleic acid sequence or a plurality of nucleic acid sequences encoding an isolated antibody or antibody fragment specific for human C5aR, said nucleic acid sequence or plurality of nucleic acid sequences comprising the HC and LC of any one of the antibodies or antibody fragments disclosed in Table 1 or Table 2.

[0136] In one embodiment, said nucleic acid composition and / or said nucleic acid sequence and / or nucleic acid sequences are isolated.

[0137] vector In one embodiment, the present disclosure provides a vector composition comprising a vector or a plurality of vectors comprising a nucleic acid composition comprising a nucleic acid sequence or a plurality of nucleic acid sequences encoding an isolated antibody or antibody fragment specific for human C5aR according to the present disclosure.

[0138] In one embodiment, the present disclosure provides a vector composition comprising a vector or a plurality of vectors comprising a nucleic acid composition comprising a nucleic acid sequence or a plurality of nucleic acid sequences encoding any one of the isolated antibodies or antibody fragments specific for human C5aR disclosed in Table 1 or Table 2.

[0139] In one embodiment, the disclosure provides a vector composition comprising a vector or vectors comprising a nucleic acid sequence or sequences disclosed in Table 1 or Table 2.

[0140] In one embodiment, the vector composition and / or the vector and / or vectors are isolated.

[0141] host cell In one embodiment, the present disclosure provides a host cell comprising a vector or a vector composition comprising a plurality of vectors comprising a nucleic acid composition comprising a nucleic acid sequence or a plurality of nucleic acid sequences encoding an isolated antibody or antibody fragment specific to human C5aR according to the present disclosure.

[0142] In one embodiment, the present disclosure refers to a host cell comprising a vector or a vector composition comprising a plurality of vectors comprising a nucleic acid composition comprising a nucleic acid sequence or a plurality of nucleic acid sequences encoding an isolated antibody or antibody fragment specific for C5aR disclosed in Table 1 or Table 2.

[0143] In one embodiment, a host cell according to the present disclosure is capable of expressing an isolated antibody or antibody fragment specific for human C5aR encoded by the vector composition or nucleic acid composition.

[0144] In a further embodiment, the host cell is an isolated host cell. In a further embodiment, the host cell is a mammalian cell. In one embodiment, the mammalian cell is a human cell. In another embodiment, the mammalian cell is a CHO cell. In one embodiment, the cell is a HEK cell. In another embodiment, the cell is a PERC.6 cell. In one embodiment, the cell is an HKB11 cell.

[0145] The skilled artisan will appreciate that the nucleic acid sequence or sequences encoding the heavy and / or light chains of an antibody or antibody fragment of the disclosure may be cloned into different vectors or into the same vector.

[0146] The vector can be introduced into a suitable host cell, such as a prokaryotic (e.g., bacterial) or eukaryotic (e.g., yeast or mammalian) cell, by methods well known in the art (see, e.g., "Current Protocols in Molecular Biology", Ausubel et al. (eds.), Greene Publishing Assoc. and John Wiley Interscience, New York, 1989 and 1992). Many cloning vectors are known to those skilled in the art, and the selection of an appropriate cloning vector is a matter of choice. The gene can be placed under the control of a promoter, a ribosome binding site (in the case of bacterial expression) and optionally an operator (collectively referred to herein as "control" elements), such that a nucleic acid sequence encoding the desired protein is transcribed into RNA in a host cell transformed with a vector containing this expression construct. The coding sequence may or may not contain a signal peptide or leader sequence. Upon expression in a host cell, the antibody or antibody fragment of the disclosure is obtained. These steps can be accomplished in different ways, as would be known by a person skilled in the art. In general, such steps generally include transforming or transfecting a suitable host cell with a nucleic acid composition or vector composition or infectious particle encoding the antibody or antibody fragment. Furthermore, such steps generally include culturing the host cell under conditions suitable for the proliferation (growth) of the host cell and culturing under conditions suitable for the production (expression, synthesis) of the encoded antibody or antibody fragment. Cultivating the host cell under conditions suitable for growth or expression is generally accomplished in the presence of a medium containing components suitable for cell growth or expression induction. In certain embodiments, the method for producing an antibody or antibody fragment of the present disclosure further includes isolating and purifying the produced antibody or antibody fragment from the host cell or medium. If the expression system secretes the protein into the growth medium, the protein can be purified directly from the medium. If the protein is not secreted, it is isolated from a cell lysate or recovered from a cell membrane fraction. The selection of appropriate growth conditions and recovery methods is within the skill of the art.The antibodies or antibody fragments of the present disclosure can then be purified by a number of techniques known to those of skill in the art.

[0147] In one embodiment, the disclosure refers to a method of producing an isolated antibody or antibody fragment specific for human C5aR of any of the antibodies disclosed in Table 1 or Table 2. In one embodiment, a method of producing an isolated antibody or antibody fragment according to the disclosure is provided, comprising culturing a host cell comprising a vector or a vector composition comprising a plurality of vectors, comprising a nucleic acid sequence or a nucleic acid composition comprising a plurality of nucleic acid sequences encoding an antibody or antibody fragment according to the disclosure, under conditions suitable for expression of the antibody or antibody fragment, and isolating the antibody or antibody fragment from the host cell or host cell culture medium. Antibodies or antibody fragments isolated as described herein may be purified by techniques known in the art, such as high performance liquid chromatography (HPLC), ion exchange chromatography, gel electrophoresis, affinity chromatography, size exclusion chromatography, etc. The conditions used to purify a particular antibody or antibody fragment will depend, in part, on factors such as overall charge, hydrophobicity, hydrophilicity, etc., which will be apparent to one of skill in the art. For affinity chromatography purification, an antibody, ligand, receptor, or antigen to which the antibody or antibody fragment binds may be used. For example, for affinity chromatography purification of antibodies or antibody fragments according to the present disclosure, matrices with Protein A or Protein G can be used. Purity of the antibody or antibody fragment can be determined by any of a variety of well-known analytical methods, including gel electrophoresis, high pressure liquid chromatography, and the like.

[0148] specificity In one embodiment, the present disclosure relates to an isolated antibody or antibody fragment specific for human C5aR disclosed in Table 1 or Table 2. In one embodiment, the isolated antibody or antibody fragment according to the present disclosure is specific for human C5aR.

[0149] In one embodiment, the isolated antibody or antibody fragment according to the present disclosure is specific for a human C5aR encoded by the amino acid sequence of SEQ ID NO: 1 and / or SEQ ID NO: 2. In one embodiment, the isolated antibody or antibody fragment according to the present disclosure is specific for a polypeptide comprising the amino acid sequence of SEQ ID NO: 1 and / or SEQ ID NO: 2. In one embodiment, the isolated antibody or antibody fragment according to the present disclosure is specific for a polypeptide consisting of the amino acid sequence of SEQ ID NO: 1 and / or SEQ ID NO: 2.

[0150] In one embodiment, an isolated antibody or antibody fragment according to the present disclosure specifically binds to the extracellular domain human C5aR. In one embodiment, an isolated antibody or antibody fragment according to the present disclosure specifically binds to the N-terminal extracellular domain of human C5aR. In one embodiment, an isolated antibody or antibody fragment according to the present disclosure specifically binds to the N-terminal extracellular domain of human C5aR, which N-terminal extracellular domain comprises the amino acid sequence of SEQ ID NO: 13. In one embodiment, an isolated antibody or antibody fragment according to the present disclosure specifically binds to the N-terminus of human C5aR comprising the amino acid sequence of SEQ ID NO: 13. In one embodiment, an isolated antibody or antibody fragment according to the present disclosure specifically binds to a peptide comprising the amino acid sequence of SEQ ID NO: 13. In one embodiment, an isolated antibody or antibody fragment according to the present disclosure specifically binds to a peptide consisting of the amino acid sequence of SEQ ID NO: 13.

[0151] In one embodiment, the isolated antibody or antibody fragment specific for human C5aR comprises: a) the HCDR1 region of SEQ ID NO: 27, the HCDR2 region of SEQ ID NO: 28, the HCDR3 region of SEQ ID NO: 29, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33 and the LCDR3 region of SEQ ID NO: 34, or b) the HCDR1 region of SEQ ID NO: 30, the HCDR2 region of SEQ ID NO: 31, the HCDR3 region of SEQ ID NO: 29, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33 and the LCDR3 region of SEQ ID NO: 34, or c) the HCDR1 region of SEQ ID NO: 27, the HCDR2 region of SEQ ID NO: 39, the HCDR3 region of SEQ ID NO: 40, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33 and the LCDR3 region of SEQ ID NO: 34, or d) the HCDR1 region of SEQ ID NO: 30, the HCDR2 region of SEQ ID NO: 41, the HCDR3 region of SEQ ID NO: 40, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33, and the LCDR3 region of SEQ ID NO: 34 Includes.

[0152] In one embodiment, the isolated antibody or antibody fragment specific for human C5aR comprises: a) a VH of SEQ ID NO: 35 and a VL of SEQ ID NO: 36, or b) a VH of SEQ ID NO: 42 and a VL of SEQ ID NO: 43, or VH and VL having at least at least 80%, at least 85%, at least 90% or at least 95% identity to VH of SEQ ID NO: 35 or 42 and to VL of SEQ ID NO: 36 or 43 Includes.

[0153] In one embodiment, the isolated antibody or antibody fragment specific for human C5aR comprises: a) HC of SEQ ID NO: 37 and LC of SEQ ID NO: 38, or b) HC of SEQ ID NO: 44 and LC of SEQ ID NO: 45, or HC and LC having at least at least 80%, at least 85%, at least 90% or at least 95% identity to the HC of SEQ ID NO: 37 or 44 and to the LC of SEQ ID NO: 38 or 45 Includes.

[0154] In one embodiment, said isolated antibody or antibody fragment specific for human C5aR is a monoclonal antibody or antibody fragment. In one embodiment, said isolated antibody or antibody fragment specific for human C5aR is a human, humanized or chimeric antibody or antibody fragment. In one embodiment, said isolated antibody or antibody fragment specific for human C5aR is a recombinant antibody or antibody fragment. In one embodiment, said isolated antibody or antibody fragment specific for human C5aR is an isolated recombinant human monoclonal antibody or antibody fragment.

[0155] Species cross-reactivity In a further embodiment, the isolated antibody or antibody fragment according to the present disclosure is cross-reactive to cynomolgus monkey (cynomolgus) C5aR. In another embodiment, the isolated antibody or antibody fragment according to the present disclosure is specific for human C5aR and cynomolgus monkey C5aR.

[0156] In one embodiment, the present disclosure refers to an isolated antibody or antibody fragment specific for human C5aR, said antibody or antibody fragment cross-reactively binding to cynomolgus C5aR. In one embodiment, the isolated antibody or antibody fragment according to the present disclosure specifically binds to the extracellular region of human C5aR and cynomolgus C5aR. In one embodiment, the isolated antibody or antibody fragment according to the present disclosure binds to the N-terminal extracellular region of human C5aR and cynomolgus C5aR. In one embodiment, the isolated antibody or antibody fragment according to the present disclosure binds to the N-terminal extracellular region of human C5aR and cynomolgus C5aR, wherein the N-terminal extracellular region of C5aR comprises the amino acid sequence of SEQ ID NO: 13 or SEQ ID NO: 14.

[0157] In yet another embodiment, the isolated antibody or antibody fragment according to the present disclosure does not bind to rodent C5aR, such as mouse or rat C5aR.

[0158] In one embodiment, an antibody or antibody fragment according to the present disclosure binds to a peptide comprising the amino acid sequence of SEQ ID NO:13 and / or SEQ ID NO:14.

[0159] In one embodiment, the antibody or antibody fragment according to the present disclosure binds to a peptide consisting of the amino acid sequence of SEQ ID NO:13 and / or SEQ ID NO:14.

[0160] In one embodiment, the isolated antibody or antibody fragment specific for human C5aR and cynomolgus C5aR is a) the HCDR1 region of SEQ ID NO: 27, the HCDR2 region of SEQ ID NO: 28, the HCDR3 region of SEQ ID NO: 29, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33 and the LCDR3 region of SEQ ID NO: 34, or b) the HCDR1 region of SEQ ID NO: 30, the HCDR2 region of SEQ ID NO: 31, the HCDR3 region of SEQ ID NO: 29, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33 and the LCDR3 region of SEQ ID NO: 34, or c) the HCDR1 region of SEQ ID NO: 27, the HCDR2 region of SEQ ID NO: 39, the HCDR3 region of SEQ ID NO: 40, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33 and the LCDR3 region of SEQ ID NO: 34, or d) the HCDR1 region of SEQ ID NO: 30, the HCDR2 region of SEQ ID NO: 41, the HCDR3 region of SEQ ID NO: 40, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33, and the LCDR3 region of SEQ ID NO: 34 Includes.

[0161] In one embodiment, the isolated antibody or antibody fragment specific for human C5aR comprises: a) a VH of SEQ ID NO: 35 and a VL of SEQ ID NO: 36, or b) a VH of SEQ ID NO: 42 and a VL of SEQ ID NO: 43, or VH and VL having at least at least 80%, at least 85%, at least 90% or at least 95% identity to VH of SEQ ID NO: 35 or 42 and to VL of SEQ ID NO: 36 or 43 Includes.

[0162] In one embodiment, the isolated antibody or antibody fragment specific for human C5aR comprises: a) HC of SEQ ID NO: 37 and LC of SEQ ID NO: 38, or b) HC of SEQ ID NO: 44 and LC of SEQ ID NO: 45, or HC and LC having at least at least 80%, at least 85%, at least 90% or at least 95% identity to the HC of SEQ ID NO: 37 or 44 and to the LC of SEQ ID NO: 38 or 45 Includes.

[0163] In one embodiment, said isolated antibody or antibody fragment specific for human C5aR is a monoclonal antibody or antibody fragment. In one embodiment, said isolated antibody or antibody fragment is a recombinant antibody or antibody fragment. In one embodiment, said isolated antibody or antibody fragment is a human, humanized or chimeric antibody or antibody fragment.

[0164] Monovalent affinity for C5aR peptides In a further embodiment, the disclosure refers to an isolated antibody or antibody fragment specific for human C5aR, said antibody or antibody fragment having a K of 100 nM or less, such as 90 nM or less, 80 nM or less, 70 nM or less, 60 nM or less, 50 nM or less, 40 nM or less, 30 nM or less, 20 nM or less, 10 nM or less, 5 nM or less, or 1 nM. D and has monovalent affinity for the human C5aR peptide comprising SEQ ID NO:13.

[0165] In one embodiment, the monovalent affinity is determined in an IgG format. In certain embodiments, the monovalent affinity is determined by surface plasmon resonance (SPR) as described in Example 4 herein.

[0166] In one embodiment, the isolated antibody or antibody fragment specific for human C5aR comprises: a) the HCDR1 region of SEQ ID NO: 27, the HCDR2 region of SEQ ID NO: 28, the HCDR3 region of SEQ ID NO: 29, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33 and the LCDR3 region of SEQ ID NO: 34, or b) the HCDR1 region of SEQ ID NO: 30, the HCDR2 region of SEQ ID NO: 31, the HCDR3 region of SEQ ID NO: 29, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33 and the LCDR3 region of SEQ ID NO: 34, or c) the HCDR1 region of SEQ ID NO: 27, the HCDR2 region of SEQ ID NO: 39, the HCDR3 region of SEQ ID NO: 40, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33 and the LCDR3 region of SEQ ID NO: 34, or d) the HCDR1 region of SEQ ID NO: 30, the HCDR2 region of SEQ ID NO: 41, the HCDR3 region of SEQ ID NO: 40, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33, and the LCDR3 region of SEQ ID NO: 34 Includes.

[0167] In one embodiment, the isolated antibody or antibody fragment specific for human C5aR comprises: a) a VH of SEQ ID NO: 35 and a VL of SEQ ID NO: 36, or b) a VH of SEQ ID NO: 42 and a VL of SEQ ID NO: 43, or VH and VL having at least at least 80%, at least 85%, at least 90% or at least 95% identity to VH of SEQ ID NO: 35 or 42 and to VL of SEQ ID NO: 36 or 43. Includes.

[0168] In one embodiment, the isolated antibody or antibody fragment specific for human C5aR comprises: a) HC of SEQ ID NO: 37 and LC of SEQ ID NO: 38, or b) HC of SEQ ID NO: 44 and LC of SEQ ID NO: 45, or HC and LC having at least at least 80%, at least 85%, at least 90% or at least 95% identity to the HC of SEQ ID NO: 37 or 44 and to the LC of SEQ ID NO: 38 or 45 Includes.

[0169] In one embodiment, said isolated antibody or antibody fragment specific for human C5aR is a monoclonal antibody or antibody fragment. In one embodiment, said isolated antibody or antibody fragment is a recombinant antibody or antibody fragment. In one embodiment, said isolated antibody or antibody fragment is a human, humanized or chimeric antibody or antibody fragment.

[0170] Apparent affinity for C5aR peptides (bivalent) In one embodiment, the disclosure refers to an isolated antibody or antibody fragment specific for human C5aR, said isolated antibody having a K of 1 nM or less, such as 0.5 nM or less, 0.4 nM or less, 0.3 nM or less, 0.2 nM or less, 0.1 nM or less, 90 pM or less, 80 pM or less, 70 pM or less, 60 pM or less, 50 pM or less, 40 pM or less, 30 pM or less, 20 pM or less, 10 pM or less, 5 pM or less, or 1 pM or less. D The C5aR peptide has an apparent affinity for human C5aR peptides containing SEQ ID NO:13 at .

[0171] In one embodiment, the disclosure refers to an isolated antibody or antibody fragment specific for human C5aR, said isolated antibody having a K of 300 nM or less, such as 250 nM or less, 200 nM or less, 150 nM or less, 100 nM or less, 90 nM or less, 80 nM or less, 70 nM or less, 60 nM or less, 50 nM or less, 40 nM or less, 30 nM or less, 20 nM or less, 10 nM or less, or 1 nM or less. D at 100 nm and has an apparent affinity for the cynomolgus C5aR peptide comprising SEQ ID NO:14.

[0172] In one embodiment, the disclosure refers to an isolated antibody or antibody fragment specific for human C5aR, said isolated antibody having a K of 0.3 nM or less. D for human C5aR peptides comprising SEQ ID NO: 13 at 100 nm and a K D at 100 nm and has an apparent affinity for the cynomolgus C5aR peptide comprising SEQ ID NO:14.

[0173] In one embodiment, the disclosure refers to an isolated antibody or antibody fragment specific for human C5aR, said isolated antibody having a K of 0.05 nM or less. D for human C5aR peptides comprising SEQ ID NO:13 at 1000 Hz and a K of 80 nM or less. D at 100 nm and has an apparent affinity for the cynomolgus C5aR peptide comprising SEQ ID NO:14.

[0174] In certain embodiments, the apparent affinity is determined in an IgG format. In one embodiment, the apparent affinity is determined by biolayer interferometry (BLI) as described in Example 5 herein.

[0175] In one embodiment, the isolated antibody or antibody fragment specific for human C5aR comprises: a) the HCDR1 region of SEQ ID NO: 27, the HCDR2 region of SEQ ID NO: 28, the HCDR3 region of SEQ ID NO: 29, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33 and the LCDR3 region of SEQ ID NO: 34, or b) the HCDR1 region of SEQ ID NO: 30, the HCDR2 region of SEQ ID NO: 31, the HCDR3 region of SEQ ID NO: 29, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33 and the LCDR3 region of SEQ ID NO: 34, or c) the HCDR1 region of SEQ ID NO: 27, the HCDR2 region of SEQ ID NO: 39, the HCDR3 region of SEQ ID NO: 40, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33 and the LCDR3 region of SEQ ID NO: 34, or d) the HCDR1 region of SEQ ID NO: 30, the HCDR2 region of SEQ ID NO: 41, the HCDR3 region of SEQ ID NO: 40, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33, and the LCDR3 region of SEQ ID NO: 34 Includes.

[0176] In one embodiment, the isolated antibody or antibody fragment specific for human C5aR comprises: a) a VH of SEQ ID NO: 35 and a VL of SEQ ID NO: 36, or b) a VH of SEQ ID NO: 42 and a VL of SEQ ID NO: 43, or VH and VL having at least at least 80%, at least 85%, at least 90% or at least 95% identity to VH of SEQ ID NO: 35 or 42 and to VL of SEQ ID NO: 36 or 43 Includes.

[0177] In one embodiment, the isolated antibody or antibody fragment specific for human C5aR comprises: a) HC of SEQ ID NO: 37 and LC of SEQ ID NO: 38, or b) HC of SEQ ID NO: 44 and LC of SEQ ID NO: 45, or HC and LC having at least at least 80%, at least 85%, at least 90% or at least 95% identity to the HC of SEQ ID NO: 37 or 44 and to the LC of SEQ ID NO: 38 or 45 Includes.

[0178] In one embodiment, said isolated antibody or antibody fragment specific for human C5aR is a monoclonal antibody or antibody fragment. In one embodiment, said isolated antibody or antibody fragment is a recombinant antibody or antibody fragment. In one embodiment, said isolated antibody or antibody fragment is a human, humanized or chimeric antibody or antibody fragment.

[0179] Apparent affinity for full-length C5aR (bivalent) In a further embodiment, the disclosure refers to an isolated antibody or antibody fragment specific for human C5aR, said isolated antibody having a K of 10 nM or less, such as 5 nM or less, 4 nM or less, 3 nM or less, 2 nM or less, 1 nM or less, 0.5 nM or less, 0.4 nM or less, 0.3 nM or less, 0.2 nM or less, or 0.1 nM or less. D It has an apparent affinity for human C5aR at

[0180] In embodiments, the disclosure refers to an isolated antibody or antibody fragment specific for human C5aR, said isolated antibody having a K of 10 nM or less, such as 9 nM or less, 8 nM or less, 7 nM or less, 6 nM or less, 5 nM or less, 4 nM or less, 3 nM or less, or 1 nM or less. D It has an apparent affinity for cynomolgus C5aR at

[0181] In one embodiment, the disclosure refers to an isolated antibody or antibody fragment specific for human C5aR, said isolated antibody having a K of 0.5 nM or less. D and K<5 nM for human C5aR at D It has apparent affinity for cynomolgus monkey C5aR at .

[0182] In one embodiment, the human C5aR comprises the amino acid sequence of SEQ ID NO: 1. In one embodiment, the human C5aR comprises the amino acid sequence of SEQ ID NO: 2. In one embodiment, the cynomolgus C5aR comprises the amino acid sequence of SEQ ID NO: 3.

[0183] In certain embodiments, the apparent affinity is determined in an IgG format. In embodiments, the human C5aR or cynomolgus C5aR is expressed on a cell. In embodiments, the human C5aR or cynomolgus C5aR is expressed on a modified CHO cell expressing full-length human. In embodiments, the CHO cell is a Flp-In CHO cell.

[0184] In certain embodiments, the apparent affinity is determined as described in Example 6 herein.

[0185] In one embodiment, the isolated antibody or antibody fragment specific for human C5aR comprises: a) the HCDR1 region of SEQ ID NO: 27, the HCDR2 region of SEQ ID NO: 28, the HCDR3 region of SEQ ID NO: 29, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33 and the LCDR3 region of SEQ ID NO: 34, or b) an HCDR1 region of SEQ ID NO: 30, an HCDR2 region of SEQ ID NO: 31, an HCDR3 region of SEQ ID NO: 29, an LCDR1 region of SEQ ID NO: 32, an LCDR2 region of SEQ ID NO: 33 and an LCDR3 region comprising the amino acid sequence of SEQ ID NO: 34, or c) an HCDR1 region of SEQ ID NO: 27, an HCDR2 region of SEQ ID NO: 39, an HCDR3 region of SEQ ID NO: 40, an LCDR1 region of SEQ ID NO: 32, an LCDR2 region of SEQ ID NO: 33 and an LCDR3 region comprising the amino acid sequence of SEQ ID NO: 34, or d) an HCDR1 region of SEQ ID NO: 30, an HCDR2 region of SEQ ID NO: 41, an HCDR3 region of SEQ ID NO: 40, an LCDR1 region of SEQ ID NO: 32, an LCDR2 region of SEQ ID NO: 33, and an LCDR3 region comprising the amino acid sequence of SEQ ID NO: 34; Includes.

[0186] In one embodiment, the isolated antibody or antibody fragment specific for human C5aR comprises: a) a VH of SEQ ID NO: 35 and a VL of SEQ ID NO: 36, or b) a VH of SEQ ID NO: 42 and a VL of SEQ ID NO: 43, or VH and VL having at least at least 80%, at least 85%, at least 90% or at least 95% identity to VH of SEQ ID NO: 35 or 42 and to VL of SEQ ID NO: 36 or 43 Includes.

[0187] In one embodiment, the isolated antibody or antibody fragment specific for human C5aR comprises: a) HC of SEQ ID NO: 37 and LC of SEQ ID NO: 38, or b) HC of SEQ ID NO: 44 and LC of SEQ ID NO: 45, or HC and LC having at least at least 80%, at least 85%, at least 90% or at least 95% identity to the HC of SEQ ID NO: 37 or 44 and to the LC of SEQ ID NO: 38 or 45 Includes.

[0188] In one embodiment, said isolated antibody or antibody fragment specific for human C5aR is a monoclonal antibody or antibody fragment. In one embodiment, said isolated antibody or antibody fragment is a recombinant antibody or antibody fragment. In one embodiment, said isolated antibody or antibody fragment is a human, humanized or chimeric antibody or antibody fragment.

[0189] Apparent EC for full-length C5aR 50 In a further embodiment, the disclosure refers to an isolated antibody or antibody fragment specific for human C5aR, said isolated antibody having an EC of 20 nM or less, 15 nM or less, 10 nM or less, 5 nM or less, 4 nM or less, 3 nM or less, 2 nM or less, 1 nM or less, 0.5 nM or less, 0.4 nM or less, 0.3 nM or less, 0.2 nM or less, or 0.1 nM or less. 50 It binds to human C5aR at concentrations

[0190] In embodiments, the disclosure refers to an isolated antibody or antibody fragment specific for human C5aR, said isolated antibody having an EC of 20 nM or less, 10 nM or less, 9 nM or less, 8 nM or less, 7 nM or less, 6 nM or less, 5 nM or less, 4 nM or less, 3 nM or less, or 1 nM or less. 50 Binds to cynomolgus monkey C5aR at concentrations

[0191] In one embodiment, the disclosure refers to an isolated antibody or antibody fragment specific for human C5aR, said isolated antibody having a K D E-commerce 50 It binds to human C5aR and cynomolgus monkey C5aR at concentrations

[0192] In one embodiment, the human C5aR comprises the amino acid sequence of SEQ ID NO: 1. In one embodiment, the human C5aR comprises the amino acid sequence of SEQ ID NO: 2. In one embodiment, the cynomolgus C5aR comprises the amino acid sequence of SEQ ID NO: 3.

[0193] In certain embodiments, the EC 50 The concentration is determined in an IgG format. In embodiments, the human C5aR or cynomolgus C5aR is expressed on a cell. In embodiments, the human C5aR or cynomolgus C5aR is expressed on a modified CHO cell expressing full-length human. In embodiments, the CHO cell is a Flp-In CHO cell. In certain embodiments, the human C5aR or cynomolgus C5aR is expressed on a neutrophil. In embodiments, the human C5aR is expressed on a human neutrophil. In embodiments, the cynomolgus C5aR is expressed on a cynomolgus neutrophil. In certain embodiments, the neutrophil is from whole blood. In certain embodiments, the EC 50 Concentrations are determined as described in Example 7 herein.

[0194] In further embodiments, the isolated antibody or antibody fragment specific for human C5aR does not substantially bind to a C5aR-associated antigen selected from the group consisting of human C5L2, human ChemR23, human FPR1 and C3aR. In certain embodiments, the isolated antibody or antibody fragment specific for human C5aR does not substantially bind to a C5aR-associated antigen selected from the group consisting of human C5L2, human ChemR23, human FPR1 and C3aR at an IgG concentration of 300 nM. In one embodiment, the binding is determined as described in Example 7.

[0195] In one embodiment, the isolated antibody or antibody fragment specific for human C5aR comprises: a) the HCDR1 region of SEQ ID NO: 27, the HCDR2 region of SEQ ID NO: 28, the HCDR3 region of SEQ ID NO: 29, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33 and the LCDR3 region of SEQ ID NO: 34, or b) an HCDR1 region of SEQ ID NO: 30, an HCDR2 region of SEQ ID NO: 31, an HCDR3 region of SEQ ID NO: 29, an LCDR1 region of SEQ ID NO: 32, an LCDR2 region of SEQ ID NO: 33 and an LCDR3 region comprising the amino acid sequence of SEQ ID NO: 34, or c) an HCDR1 region of SEQ ID NO: 27, an HCDR2 region of SEQ ID NO: 39, an HCDR3 region of SEQ ID NO: 40, an LCDR1 region of SEQ ID NO: 32, an LCDR2 region of SEQ ID NO: 33 and an LCDR3 region comprising the amino acid sequence of SEQ ID NO: 34, or d) an HCDR1 region of SEQ ID NO: 30, an HCDR2 region of SEQ ID NO: 41, an HCDR3 region of SEQ ID NO: 40, an LCDR1 region of SEQ ID NO: 32, an LCDR2 region of SEQ ID NO: 33, and an LCDR3 region comprising the amino acid sequence of SEQ ID NO: 34; Includes.

[0196] In one embodiment, the isolated antibody or antibody fragment specific for human C5aR comprises: a) a VH of SEQ ID NO: 35 and a VL of SEQ ID NO: 36, or b) a VH of SEQ ID NO: 42 and a VL of SEQ ID NO: 43, or VH and VL having at least at least 80%, at least 85%, at least 90% or at least 95% identity to VH of SEQ ID NO: 35 or 42 and to VL of SEQ ID NO: 36 or 43 Includes.

[0197] In one embodiment, the isolated antibody or antibody fragment specific for human C5aR comprises: a) HC of SEQ ID NO: 37 and LC of SEQ ID NO: 38, or b) HC of SEQ ID NO: 44 and LC of SEQ ID NO: 45, or HC and LC having at least at least 80%, at least 85%, at least 90% or at least 95% identity to VH of SEQ ID NO: 37 or 44 and to VL of SEQ ID NO: 38 or 45 Includes.

[0198] In one embodiment, said isolated antibody or antibody fragment specific for human C5aR is a monoclonal antibody or antibody fragment. In one embodiment, said isolated antibody or antibody fragment is a recombinant antibody or antibody fragment. In one embodiment, said isolated antibody or antibody fragment is a human, humanized or chimeric antibody or antibody fragment.

[0199] In one embodiment, the disclosure refers to an isolated antibody or antibody fragment specific for human C5aR, said antibody or antibody fragment having an EC5aR of 5 nM or less, such as 4 nM or less, 3 nM or less, 2 nM or less, 1 nM or less, or 0.5 nM or less. 50 The antibodies bind to human C5aR comprising SEQ ID NO:1 and / or SEQ ID NO:2 at concentrations

[0200] In certain embodiments, the human C5aR is displayed on a virus-like particle. In certain embodiments, the human C5aR is expressed as a fusion protein. In certain embodiments, the human C5aR is fused to a GAG protein. In embodiments, the fusion protein comprises an amino acid sequence disclosed in Table 8. In certain embodiments, the binding is determined by ELISA as described in Example 8.

[0201] In one embodiment, the isolated antibody or antibody fragment specific for human C5aR comprises: a) the HCDR1 region of SEQ ID NO: 27, the HCDR2 region of SEQ ID NO: 28, the HCDR3 region of SEQ ID NO: 29, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33 and the LCDR3 region of SEQ ID NO: 34, or b) the HCDR1 region of SEQ ID NO: 30, the HCDR2 region of SEQ ID NO: 31, the HCDR3 region of SEQ ID NO: 29, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33 and the LCDR3 region of SEQ ID NO: 34, or c) the HCDR1 region of SEQ ID NO: 27, the HCDR2 region of SEQ ID NO: 39, the HCDR3 region of SEQ ID NO: 40, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33 and the LCDR3 region of SEQ ID NO: 34, or d) the HCDR1 region of SEQ ID NO: 30, the HCDR2 region of SEQ ID NO: 41, the HCDR3 region of SEQ ID NO: 40, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33, and the LCDR3 region of SEQ ID NO: 34 Includes.

[0202] In one embodiment, the isolated antibody or antibody fragment is specific for human C5aR. a) a VH of SEQ ID NO: 35 and a VL of SEQ ID NO: 36, or b) a VH of SEQ ID NO: 42 and a VL of SEQ ID NO: 43, or A VH and VL having at least at least 80%, at least 85%, at least 90% or at least 95% identity to the VH of SEQ ID NO: 35 or 42 and to the VL of SEQ ID NO: 36 or 43.

[0203] In one embodiment, the isolated antibody or antibody fragment specific for human C5aR comprises: a) HC of SEQ ID NO: 37 and LC of SEQ ID NO: 38, or b) HC of SEQ ID NO: 44 and LC of SEQ ID NO: 45, or A HC and LC having at least at least 80%, at least 85%, at least 90% or at least 95% identity to the HC of SEQ ID NO: 37 or 44 and to the LC of SEQ ID NO: 38 or 45.

[0204] In one embodiment, said isolated antibody or antibody fragment specific for human C5aR is a monoclonal antibody or antibody fragment. In one embodiment, said isolated antibody or antibody fragment is a recombinant antibody or antibody fragment. In one embodiment, said isolated antibody or antibody fragment is a human, humanized or chimeric antibody or antibody fragment.

[0205] Functional-C5A-induced activation of C5aR In general, an isolated antibody or antibody fragment specific for human C5aR according to the present disclosure can be used to prevent or inhibit the interaction between human C5aR and human C5a, thereby preventing, inhibiting, neutralizing or reducing signal transduction pathways mediated by C5aR and / or modulating biological pathways and mechanisms involving C5aR.

[0206] In one embodiment, the disclosure relates to an isolated antibody or antibody fragment specific for human C5aR, said isolated antibody or antibody fragment specifically interferes with C5aR-mediated signal transduction.

[0207] In further embodiments of the disclosure, the isolated antibody or antibody fragment specific for human C5aR specifically interferes with the interaction of C5a with C5aR expressed on a cell, hi yet further embodiments, said isolated antibody or antibody fragment is capable of specifically interfering with C5a-induced signal transduction mediated by C5aR.

[0208] Methods for assaying for the functional activity of C5aR-specific antibodies may utilize binding assays, such as enzyme-linked immunosorbent assays (ELISA), radioimmunoassays (RIA), fluorescence-activated cell sorting (FACS) and other methods well known in the art (see Hampton, R. et al. (1990; Serological Methods a Laboratory Manual, APS Press, St Paul, MN) and Maddox, DE et al. (1983; J. Exp. Med. 158:1211-1216)). Alternatively, the assay may test the ability of an isolated antibody or antibody fragment of the present disclosure in eliciting a biological response as a result of binding to C5aR, either in vivo or in vitro. Such assays are described in the Examples disclosed herein. Other suitable assays will be known to those of skill in the art.

[0209] In certain embodiments, the isolated antibody or antibody fragment of the disclosure antagonizes human C5aR activity. In one embodiment, the isolated antibody or antibody fragment of the disclosure neutralizes human C5aR activity. In one embodiment, the isolated antibody or antibody fragment of the disclosure inhibits human C5aR activity. In one embodiment, the isolated antibody or antibody fragment of the disclosure inhibits human C5aR signaling. In one embodiment, the human C5aR activity or human C5aR signaling is induced by C5a. In one embodiment, the human C5aR activity or human C5aR signaling is induced by the interaction of human C5a with human C5aR. In one embodiment, the C5aR activity or C5aR signaling is induced by binding of human C5a to human C5aR. In one embodiment, the C5aR is expressed on a cell. In one embodiment, the human C5aR activity or human C5aR signaling is inhibited in vitro and / or ex vivo and / or in vivo.

[0210] C5A-induced β-arrestin recruitment The ability of isolated antibodies or antibody fragments specific for human C5aR according to the present disclosure to inhibit C5a-induced C5aR activation was tested in a β-arrestin recruitment assay as described in Example 14, revealing that both antibodies are even more potent antagonists of C5aR activity when compared to the prior art antibody RefMAB#1, especially at high pathophysiological C5a concentrations.

[0211] Thus, in one embodiment of the present disclosure, an isolated antibody or antibody fragment specific for human C5aR inhibits the ability of C5a to induce C5aR activity. In one embodiment, said C5a-induced C5aR activity is determined in a β-arrestin recruitment assay as described in Example 14. In one embodiment, said C5a-induced C5aR activity is determined in vitro.

[0212] In one such embodiment, the disclosure provides an isolated antibody or antibody fragment specific for human C5aR, wherein said isolated antibody or antibody fragment inhibits the ability of human C5a to induce human C5aR-mediated β-arrestin recruitment. In one embodiment, said isolated antibody or antibody fragment inhibits human C5a-induced β-arrestin recruitment. In one embodiment, said isolated antibody or antibody fragment inhibits human C5aR-mediated β-arrestin recruitment.

[0213] In a further embodiment of the disclosure, an isolated antibody or antibody fragment specific for human C5aR inhibits human C5a-induced human C5aR interaction with β-arrestin. In one such embodiment, said human C5a-induced human C5aR interaction with β-arrestin and / or its human C5a-induced β-arrestin recruitment is measured using β-galactosidase enzyme fragment complementation. In one embodiment, said human C5a-induced human C5aR interaction with β-arrestin and / or its human C5a-induced β-arrestin recruitment is determined as described in Example 14. In one such embodiment, said human C5a-induced human C5aR interaction with β-arrestin and / or its human C5a-induced β-arrestin recruitment is tested at an IgG concentration of 50 nM.

[0214] The ability of an isolated antibody or antibody fragment according to the present disclosure to inhibit human C5a-induced human C5aR activity, such as inhibiting human C5a-induced human C5aR interaction with β-arrestin and / or human C5a-induced human C5aR-mediated β-mobilization, can be determined by generating dose-response curves against increasing concentrations of human C5a and a fixed concentration of IgG and calculating the individual EC50 concentrations.

[0215] In one embodiment, the disclosure provides an isolated antibody or antibody fragment specific for human C5aR, said isolated antibody or antibody fragment having an EC determined in the absence of said isolated antibody or antibody fragment. 50 EC determined against human C5a in a β-arrestin recruitment assay at an IgG concentration of 50 nM compared to 50 The concentration is increased by at least 5-fold or more, such as at least 6-fold, at least 7-fold, at least 8-fold, at least 9-fold, at least 10-fold, at least 11-fold, at least 12-fold, or at least 13-fold.

[0216] In one embodiment, the disclosure provides an isolated antibody or antibody fragment specific for human C5aR, said isolated antibody or antibody fragment having an EC determined in the absence of said isolated antibody or antibody fragment.50 EC determined against human C5a in a β-arrestin recruitment assay at an IgG concentration of 50 nM when compared to 50 The concentration is increased by about 5-fold, about 6-fold, about 7-fold, about 8-fold, about 9-fold, about 10-fold, about 11-fold, about 12-fold, or about 13-fold.

[0217] In one embodiment, the present disclosure provides an antibody or antibody fragment specific for human C5aR, wherein the human C5a needs to be present at a concentration of at least 5-fold higher, such as at least 6-fold higher, at least 7-fold higher, at least 8-fold higher, at least 9-fold higher, at least 10-fold higher, at least 11-fold higher, at least 12-fold higher or at least 13-fold higher, in order to induce the same human C5aR activity in a β-arrestin recruitment assay at an IgG concentration of 50 nM compared to the concentration of human C5a in the absence of the antibody or antibody fragment.

[0218] In one embodiment, the β-arrestin recruitment assay is performed as described in Example 14. In one embodiment, the β-arrestin recruitment assay is performed in vitro.

[0219] Alternatively, the ability of an isolated antibody or antibody fragment specific for C5aR according to the present disclosure to inhibit C5a-induced C5aR-mediated β-arrestin recruitment can be determined by calculating the % inhibition against different human C5a concentrations.

[0220] In one such embodiment, an isolated antibody or antibody fragment specific for C5aR according to the present disclosure inhibits human C5a-induced human C5aR-mediated β-arrestin recruitment by at least 50%, at least 55%, at least 60%, at least 70%, at least 80% or at least 90% in the presence of 1.2 nM or 11 nM human C5a at an IgG concentration of 50 nM, compared to the level of human C5a-induced human C5aR-mediated β-arrestin recruitment in the presence of 1.2 nM or 11 nM human C5a and in the absence of the antibody or antibody fragment.

[0221] In one such embodiment, an isolated antibody or antibody fragment specific for C5aR according to the present disclosure inhibits human C5a-induced human C5aR-mediated β-arrestin recruitment by at least 25%, such as at least 30%, at least 35%, at least 40%, at least 45% or at least 50%, at an IgG concentration of 50 nM and in the presence of 1.2 nM or 11 nM human C5a, compared to the level of human C5a-induced human C5aR-mediated β-arrestin recruitment in the presence of 100 nM human C5a and in the absence of the antibody or antibody fragment.

[0222] In one embodiment, the β-arrestin recruitment assay is performed as described in Example 14. In one embodiment, the β-arrestin recruitment assay is performed in vitro.

[0223] In one embodiment, the isolated antibody or antibody fragment specific for human C5aR comprises: a) the HCDR1 region of SEQ ID NO: 27, the HCDR2 region of SEQ ID NO: 28, the HCDR3 region of SEQ ID NO: 29, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33 and the LCDR3 region of SEQ ID NO: 34, or b) the HCDR1 region of SEQ ID NO: 30, the HCDR2 region of SEQ ID NO: 31, the HCDR3 region of SEQ ID NO: 29, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33 and the LCDR3 region of SEQ ID NO: 34, or c) the HCDR1 region of SEQ ID NO: 27, the HCDR2 region of SEQ ID NO: 39, the HCDR3 region of SEQ ID NO: 40, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33 and the LCDR3 region of SEQ ID NO: 34, or d) an HCDR1 region of SEQ ID NO: 30, an HCDR2 region of SEQ ID NO: 41, an HCDR3 region of SEQ ID NO: 40, an LCDR1 region of SEQ ID NO: 32, an LCDR2 region of SEQ ID NO: 33, and an LCDR3 region comprising the amino acid sequence of SEQ ID NO: 34; Includes.

[0224] In one embodiment, the isolated antibody or antibody fragment specific for human C5aR comprises: a) a VH of SEQ ID NO: 35 and a VL of SEQ ID NO: 36, or b) a VH of SEQ ID NO: 42 and a VL of SEQ ID NO: 43, or VH and VL having at least at least 80%, at least 85%, at least 90% or at least 95% identity to VH of SEQ ID NO: 35 or 42 and to VL of SEQ ID NO: 36 or 43 Includes.

[0225] In one embodiment, the isolated antibody or antibody fragment specific for human C5aR comprises: a) HC of SEQ ID NO: 37 and LC of SEQ ID NO: 38, or b) HC of SEQ ID NO: 44 and LC of SEQ ID NO: 45, or HC and LC having at least at least 80%, at least 85%, at least 90% or at least 95% identity to the HC of SEQ ID NO: 37 or 44 and to the LC of SEQ ID NO: 38 or 45 Includes.

[0226] In one embodiment, said isolated antibody or antibody fragment specific for human C5aR is a monoclonal antibody or antibody fragment. In one embodiment, said isolated antibody or antibody fragment is a recombinant antibody or antibody fragment. In one embodiment, said isolated antibody or antibody fragment is a human, humanized or chimeric antibody or antibody fragment.

[0227] C5A-induced upregulation of CD11b expression C5a, as a potent activator of human neutrophils and monocytes, induces upregulation of the cell surface antigen CD11b in such cells. Thus, the ability of an isolated antibody or antibody fragment specific for human C5aR according to the present disclosure to inhibit C5a-induced activation of granulocytes and / or monocytes can be assessed by determining the CD11b expression level in such cells.

[0228] The ability of isolated antibodies or antibody fragments according to the present disclosure to inhibit CD11b expression on granulocytes and / or monocytes was determined by generating dose-response curves to increasing concentrations of IgG and a fixed concentration of human C5a and assessing the individual IC 50Alternatively, the ability of an isolated antibody or antibody fragment according to the present disclosure to inhibit CD11b expression on granulocytes and / or monocytes C5a can be determined by calculating the % inhibition of CD11b expression for different IgG concentrations.

[0229] In one such embodiment, an isolated antibody or antibody fragment specific for human C5aR according to the present disclosure has an IC of 30 nM or less, 25 nM or less, 20 nM or less, 15 nM or less, 10 nM or less, or 5 nM or less in the presence of 15 nM human C5a. 50 At concentrations, it inhibits human C5a-induced CD11b expression in human granulocytes and / or human monocytes.

[0230] In another embodiment, an isolated antibody or antibody fragment specific for human C5aR according to the present disclosure inhibits human C5a-induced CD11b expression in human granulocytes and / or human monocytes by at least 70%, at least 75%, 80%, at least 85% or at least 90% in the presence of 15 nM human C5a and an IgG concentration of 600 nM compared to the CD11b expression level in the presence of 15 nM human C5a and in the absence of the antibody or antibody fragment.

[0231] In further embodiments, the isolated antibody or antibody fragment specific for human C5aR according to the present disclosure has an IC of 150 nM or less, 125 nM or less, 100 nM or less, 90 nM or less, 80 nM or less, 70 nM or less, 60 nM or less, 50 nM or less, or 40 nM or less in the presence of 150 nM human C5a. 50 At concentrations, it inhibits human C5a-induced CD11b expression in human granulocytes.

[0232] In one embodiment, an isolated antibody or antibody fragment specific for human C5aR according to the present disclosure has an IC of 42 nM in the presence of 150 nM human C5a. 50 At concentrations, it inhibits human C5a-induced CD11b expression in human granulocytes.

[0233] In a further embodiment, an isolated antibody or antibody fragment specific for human C5aR according to the present disclosure inhibits human C5a-induced CD11b expression in human granulocytes by at least 65%, at least 70%, at least 75%, at least 80% or at least 90% in the presence of 150 nM human C5a and an IgG concentration of 600 nM, compared to the CD11b expression level in the presence of 150 nM human C5a and in the absence of the antibody or antibody fragment.

[0234] In one embodiment, an isolated antibody or antibody fragment specific for human C5aR according to the present disclosure inhibits C5a-induced CD11b expression in human granulocytes by at least 45%, at least 50%, at least 55%, at least 60% or at least 65% in the presence of 150 nM human C5a and an IgG concentration of 100 nM, compared to the CD11b expression level in the presence of 150 nM human C5a and in the absence of the antibody or antibody fragment.

[0235] In one embodiment, said determining of CD11b expression is performed as described in Example 15. In one embodiment, said determining of CD11b expression is performed in vitro and / or ex vivo.

[0236] In one embodiment, the isolated antibody or antibody fragment specific for human C5aR comprises: a) the HCDR1 region of SEQ ID NO: 27, the HCDR2 region of SEQ ID NO: 28, the HCDR3 region of SEQ ID NO: 29, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33 and the LCDR3 region of SEQ ID NO: 34, or b) the HCDR1 region of SEQ ID NO: 30, the HCDR2 region of SEQ ID NO: 31, the HCDR3 region of SEQ ID NO: 29, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33 and the LCDR3 region of SEQ ID NO: 34, or c) the HCDR1 region of SEQ ID NO: 27, the HCDR2 region of SEQ ID NO: 39, the HCDR3 region of SEQ ID NO: 40, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33 and the LCDR3 region of SEQ ID NO: 34, or d) the HCDR1 region of SEQ ID NO: 30, the HCDR2 region of SEQ ID NO: 41, the HCDR3 region of SEQ ID NO: 40, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33, and the LCDR3 region of SEQ ID NO: 34 Includes.

[0237] In one embodiment, the isolated antibody or antibody fragment specific for human C5aR comprises: a) a VH of SEQ ID NO: 35 and a VL of SEQ ID NO: 36, or b) a VH of SEQ ID NO: 42 and a VL of SEQ ID NO: 43, or VH and VL having at least at least 80%, at least 85%, at least 90% or at least 95% identity to VH of SEQ ID NO: 35 or 42 and to VL of SEQ ID NO: 36 or 43 Includes.

[0238] In one embodiment, the isolated antibody or antibody fragment specific for human C5aR comprises: a) HC of SEQ ID NO: 37 and LC of SEQ ID NO: 38, or b) HC of SEQ ID NO: 44 and LC of SEQ ID NO: 45, or HC and LC having at least at least 80%, at least 85%, at least 90% or at least 95% identity to the HC of SEQ ID NO: 37 or 44 and to the LC of SEQ ID NO: 38 or 45 Includes.

[0239] In one embodiment, said isolated antibody or antibody fragment specific for human C5aR is a monoclonal antibody or antibody fragment. In one embodiment, said isolated antibody or antibody fragment is a recombinant antibody or antibody fragment. In one embodiment, said isolated antibody or antibody fragment is a human, humanized or chimeric antibody or antibody fragment.

[0240] Moreover, the inhibitory activity of the isolated antibodies or antibody fragments according to the present disclosure on human C5a-induced CD11b expression over time was further analyzed, meaning that the antibodies were pre-incubated with granulocytes and / or monocytes present in whole blood for various lengths of time (e.g. 300 min vs. 20 min) before the addition of human C5a. Surprisingly, this experiment revealed that the isolated antibodies according to the present disclosure are even more potent antagonists of C5aR activity over longer periods, e.g. long incubation times, especially when compared to the prior art antibody RefMAB#1.

[0241] Thus, the ability of the isolated antibodies or antibody fragments according to the present disclosure to inhibit human C5a-induced CD11b expression in granulocytes and / or monocytes over time was demonstrated by generating dose-response curves against increasing concentrations of IgG and a fixed concentration of human C5a, and by assessing the individual ECs after incubating the isolated antibodies with the granulocytes and / or monocytes for different incubation times. 50 It can be determined by calculating the value.

[0242] As shown in Figures 6A and C, the isolated antibodies or antibody fragments of the present disclosure exhibited a clear shift in the dose-response curves determined after 300 minutes of incubation to lower IgG concentrations when compared to the dose-response curves determined after 20 minutes of incubation. Interestingly, RefMAB#1 demonstrated no increase in potency over time (see Figures 6B and D).

[0243] Thus, in one embodiment, the isolated antibody or antibody fragment according to the present disclosure is more potent in inhibiting C5a-induced CD11b expression in human granulocytes and / or human monocytes after extended incubation times with said cells.

[0244] In one such embodiment, an isolated antibody or antibody fragment according to the present disclosure has an IC 50IC200 at least 2-fold, at least 4-fold, at least 5-fold, or at least 6-fold reduced when determined after a long incubation time of 50, 100, 150, 200, 250, or 300 minutes compared to the 50 At concentrations, it inhibits human C5a-induced CD11b expression in human granulocytes and / or human monocytes.

[0245] In one embodiment, an isolated antibody or antibody fragment according to the present disclosure has an IC 50 The IC was approximately 5-fold lower when determined after a longer incubation time of 300 min compared to the 50 At concentrations, it inhibits human C5a-induced CD11b expression in human granulocytes and / or human monocytes.

[0246] In one embodiment, the isolated antibody or antibody fragment according to the present disclosure is a corresponding IC of RefMAB#1. 50 The IC50 concentration was at least 3-fold, at least 4-fold, at least 5-fold, at least 10-fold, at least 15-fold, or at least 19-fold lower when determined after a long incubation time of 300 minutes compared to the 50 At concentrations, it inhibits human C5a-induced CD11b expression in human granulocytes and / or human monocytes.

[0247] In one embodiment, the isolated antibody or antibody fragment according to the present disclosure has an IC of 3 nM or less, 2.5 nM or less, 2 nM or less, 1.5 nM or less, or 1 nM or less after a prolonged incubation time of 300 minutes with the cells. 50 At concentrations, it inhibits human C5a-induced CD11b expression in human granulocytes and / or human monocytes.

[0248] In one embodiment, the isolated antibody or antibody fragment specific for human C5aR comprises: a) the HCDR1 region of SEQ ID NO: 27, the HCDR2 region of SEQ ID NO: 28, the HCDR3 region of SEQ ID NO: 29, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33 and the LCDR3 region of SEQ ID NO: 34, or b) the HCDR1 region of SEQ ID NO: 30, the HCDR2 region of SEQ ID NO: 31, the HCDR3 region of SEQ ID NO: 29, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33 and the LCDR3 region of SEQ ID NO: 34, or c) the HCDR1 region of SEQ ID NO: 27, the HCDR2 region of SEQ ID NO: 39, the HCDR3 region of SEQ ID NO: 40, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33 and the LCDR3 region of SEQ ID NO: 34, or d) the HCDR1 region of SEQ ID NO: 30, the HCDR2 region of SEQ ID NO: 41, the HCDR3 region of SEQ ID NO: 40, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33, and the LCDR3 region of SEQ ID NO: 34 Includes.

[0249] In one embodiment, the isolated antibody or antibody fragment specific for human C5aR comprises: a) a VH of SEQ ID NO: 35 and a VL of SEQ ID NO: 36, or b) a VH of SEQ ID NO: 42 and a VL of SEQ ID NO: 43, or VH and VL having at least at least 80%, at least 85%, at least 90% or at least 95% identity to VH of SEQ ID NO: 35 or 42 and to VL of SEQ ID NO: 36 or 43 Includes.

[0250] In one embodiment, the isolated antibody or antibody fragment specific for human C5aR comprises: a) HC of SEQ ID NO: 37 and LC of SEQ ID NO: 38, or b) HC of SEQ ID NO: 44 and LC of SEQ ID NO: 45, or HC and LC having at least at least 80%, at least 85%, at least 90% or at least 95% identity to the HC of SEQ ID NO: 37 or 44 and to the LC of SEQ ID NO: 38 or 45 Includes.

[0251] In one embodiment, said isolated antibody or antibody fragment specific for human C5aR is a monoclonal antibody or antibody fragment. In one embodiment, said isolated antibody or antibody fragment is a recombinant antibody or antibody fragment. In one embodiment, said isolated antibody or antibody fragment is a human, humanized or chimeric antibody or antibody fragment.

[0252] C5A-induced migration of neutrophils In further assays, the ability of isolated antibodies or antibody fragments specific for human C5aR according to the present disclosure to inhibit human C5a-induced migration of human neutrophils was evaluated, revealing that MAB#1 efficiently inhibited C5-induced neutrophil migration in vitro.

[0253] Thus, in one embodiment of the present disclosure, said isolated antibody or antibody fragment specific for human C5aR of the present disclosure inhibits human C5a-induced migration of human neutrophils in vitro.

[0254] In further embodiments, the isolated antibody or antibody fragment inhibits human C5a-induced migration of human neutrophils in vitro by at least 25%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 75 or at least 80%, compared to migration levels in the presence of 10 nM human C5a and in the absence of the isolated antibody or antibody fragment.

[0255] In one embodiment, the migration of human neutrophils is determined after 15 minutes, 25 minutes, and / or 35 minutes. In certain embodiments, the isolated antibody or antibody fragment according to the present disclosure is tested at an IgG concentration of 100 nM and / or 600 nM.

[0256] In one embodiment, the isolated antibody or antibody fragment of the disclosure inhibits human C5a-induced migration of human neutrophils by at least 25% at an IgG concentration of 100 nM after 35 minutes compared to the level of migration after 35 minutes in the presence of 10 nM human C5a and in the absence of the antibody or antibody fragment.

[0257] In one embodiment, the isolated antibody or antibody fragment of the disclosure inhibits human C5a-induced migration of human neutrophils by at least 60% at IgG concentrations of 100 nM and / or 600 nM after 25 minutes compared to the level of migration after 25 minutes in the presence of 10 nM human C5a and in the absence of the antibody or antibody fragment.

[0258] In one embodiment, the isolated antibody or antibody fragment of the disclosure inhibits human C5a-induced migration of human neutrophils by at least 40% at an IgG concentration of 600 nM after 35 minutes compared to the level of migration after 35 minutes in the presence of 10 nM human C5a and in the absence of the antibody.

[0259] In one embodiment, the isolated antibody or antibody fragment specific for human C5aR comprises: a) the HCDR1 region of SEQ ID NO: 27, the HCDR2 region of SEQ ID NO: 28, the HCDR3 region of SEQ ID NO: 29, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33 and the LCDR3 region of SEQ ID NO: 34, or b) the HCDR1 region of SEQ ID NO: 30, the HCDR2 region of SEQ ID NO: 31, the HCDR3 region of SEQ ID NO: 29, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33 and the LCDR3 region of SEQ ID NO: 34, or c) the HCDR1 region of SEQ ID NO: 27, the HCDR2 region of SEQ ID NO: 39, the HCDR3 region of SEQ ID NO: 40, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33 and the LCDR3 region of SEQ ID NO: 34, or d) the HCDR1 region of SEQ ID NO: 30, the HCDR2 region of SEQ ID NO: 41, the HCDR3 region of SEQ ID NO: 40, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33, and the LCDR3 region of SEQ ID NO: 34 Includes.

[0260] In one embodiment, the isolated antibody or antibody fragment specific for human C5aR comprises: a) a VH of SEQ ID NO: 35 and a VL of SEQ ID NO: 36, or b) a VH of SEQ ID NO: 42 and a VL of SEQ ID NO: 43, or VH and VL having at least at least 80%, at least 85%, at least 90% or at least 95% identity to VH of SEQ ID NO: 35 or 42 and to VL of SEQ ID NO: 36 or 43 Includes.

[0261] In one embodiment, the isolated antibody or antibody fragment specific for human C5aR comprises: a) HC of SEQ ID NO: 37 and LC of SEQ ID NO: 38, or b) HC of SEQ ID NO: 44 and LC of SEQ ID NO: 45, or HC and LC having at least at least 80%, at least 85%, at least 90% or at least 95% identity to the HC of SEQ ID NO: 37 or 44 and to the LC of SEQ ID NO: 38 or 45 Includes.

[0262] In one embodiment, said isolated antibody or antibody fragment specific for human C5aR is a monoclonal antibody or antibody fragment. In one embodiment, said isolated antibody or antibody fragment is a recombinant antibody or antibody fragment. In one embodiment, said isolated antibody or antibody fragment is a human, humanized or chimeric antibody or antibody fragment.

[0263] Effector Function The Fc region of an immunoglobulin generally confers favorable pharmacokinetic properties to an antibody, such as an extended serum half-life, and the ability to induce effector functions via binding to Fc receptors expressed on cells. However, binding to Fc receptors can also result in undesired activation of certain cell surface receptors, leading to unwanted cytokine release and severe side effects upon systemic administration.

[0264] Thus, for certain therapeutic situations, it may be desirable to reduce or eliminate the normal binding of a wild-type Fc region of an antibody, such as a wild-type IgG Fc region, to one or more or all of the Fc receptors and / or binding to complement components, such as C1q, in order to reduce or eliminate the ability of the antibody to induce effector functions. For example, it may be desirable to reduce or eliminate binding of the Fc region of an antibody to one or more or all of the Fcγ receptors, such as FcγRI, FcγRIIa, FcγRIIb, FcγRIIIa, etc.

[0265] Effector functions may include, but are not limited to, one or more of the following: complement dependent cytotoxicity (CDC), antibody-dependent cell-mediated cytotoxicity (ADCC), antibody-dependent cellular phagocytosis (ADCP), cytokine secretion, immune complex-mediated antigen uptake by antigen presenting cells, binding to NK cells, binding to macrophages, binding to monocytes, binding to polymorphonuclear cells, direct signaling including apoptosis, crosslinking of target-bound antibodies, dendritic cell maturation, or T cell priming.

[0266] Reduced or eliminated binding of the Fc region to Fc receptors and / or to C1q is generally achieved by mutating a wild-type Fc region, such as an IgG1 Fc region, more particularly a human IgG1 Fc region, resulting in a mutant or modified Fc region of said wild-type Fc region, such as a mutant human IgG1 Fc region. Substitutions that reduce binding may be useful. To reduce or eliminate the binding properties of an Fc region to an Fc receptor, non-conservative amino acid substitutions, i.e. replacing one amino acid with another amino acid with different structure and / or chemical properties, are preferred.

[0267] Thus, in one embodiment, an isolated antibody or antibody fragment specific for human C5aR according to the present disclosure comprises a mutant Fc region that has reduced or eliminated binding to an Fc receptor and / or to C1q when compared to a wild-type Fc region. In one such embodiment, the isolated antibody or antibody fragment according to the present disclosure comprises a mutant Fc region that reduces or eliminates the ability of the antibody to induce effector function. In a further embodiment, the isolated antibody or antibody fragment according to the present disclosure does not substantially induce effector function.

[0268] In certain embodiments, the effector function is one or more selected from the group consisting of CDC, ADCC, and ADCP. In one embodiment, the effector function is ADCC. In one embodiment, the effector function is CDC. In one embodiment, the effector function is ADCP. In one embodiment, an isolated antibody or antibody fragment according to the present disclosure does not substantially induce ADCC and / or CDC and / or ADCP. In one embodiment, an isolated antibody or antibody fragment according to the present disclosure does not induce ADCC or ADCP in vitro.

[0269] In one embodiment, the variant Fc region of the isolated antibody or antibody fragment according to the disclosure comprises one or more amino acid substitutions that reduce or eliminate binding of the variant Fc region to one or more Fc receptors and / or to C1q when compared to the wild-type Fc region. In one embodiment, the variant Fc region of the isolated antibody or antibody fragment according to the disclosure comprises one or more amino acid substitutions that reduce or eliminate the ability of the antibody to induce effector function when compared to the wild-type Fc region.

[0270] In certain embodiments, the one or more amino acid substitutions may reduce the binding affinity of the mutant Fc region to one or more Fc receptors and / or to C1q by at least 2-fold, at least 5-fold, at least 10-fold, at least 20-fold, or even at least 50-fold when compared to the wild-type Fc region. In alternative embodiments, the one or more amino acid substitutions may reduce the ability of the isolated antibody or antibody fragment according to the present disclosure to induce effector function by at least 2-fold, at least 5-fold, at least 10-fold, at least 20-fold, or even at least 50-fold when compared to the wild-type Fc region.

[0271] In one embodiment, the mutated Fc region of the isolated antibody or antibody fragment according to the present disclosure does not substantially bind to one or more Fc receptors and / or C1q. In one embodiment, the mutated Fc region of the antibody according to the present disclosure substantially eliminates the ability of said antibody to induce effector function. In one embodiment, the antibody or antibody fragment according to the present disclosure does not substantially induce effector function. In one embodiment, said effector function is ADCC and / or ADCP and / or CDC. In one embodiment, the antibody or antibody fragment according to the present disclosure does not substantially induce effector function, meaning that the level of induced effector function is not significantly above background when measured in the absence of said antibody.

[0272] In one embodiment, the Fc receptor is a human Fc receptor. In one embodiment, the Fc receptor is an Fcγ receptor. In one embodiment, the Fc receptor is human FcγRIIIa, FcγRI, FcγRIIa and / or FcγRIIb.

[0273] In one embodiment, the effector function is one or more selected from the group of CDC, ADCC and ADCP. In a particular embodiment, the effector function is ADCC, CDC and ADCP. In a more particular embodiment, the effector function is ADCC and ADCP.

[0274] In one embodiment, the wild-type Fc region is an IgG1 Fc region. In one embodiment, the wild-type Fc region is a human IgG1 Fc region. In one embodiment, the wild-type Fc region is a human IgG1 Fc region. In one embodiment, the wild-type Fc region is This is a human IgG1 Fc region comprising the amino acid sequence of PELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK (sequence number 11).

[0275] In one embodiment, an isolated antibody or antibody fragment according to the disclosure comprises a mutant human IgG1 Fc region comprising one or more amino acid substitutions compared to a wild-type human IgG1 Fc region, which in one embodiment reduce or eliminate binding of the mutant Fc region to an Fc receptor and / or to C1q and / or reduce the ability of the antibody to induce effector function when compared to the wild-type Fc region.

[0276] In one embodiment, the mutated human IgG1 Fc region of an antibody or antibody fragment according to the present disclosure comprises amino acid substitutions at one or more positions selected from the group of 234, 235, 237, 330 and 331, as numbered according to the EU index, compared to a wild-type IgG1 Fc region comprising the amino acid sequence of SEQ ID NO:11.

[0277] In one embodiment, the mutant human IgG1 Fc region comprises an amino acid substitution at one or more positions selected from the group of L234, L235 and G237, as numbered according to the EU index, compared to a wild-type IgG1 Fc region comprising the amino acid sequence of SEQ ID NO: 11.

[0278] In one embodiment, the mutated human IgG1 Fc region comprises amino acid substitutions at one or more positions selected from the group of L234A, L235E G237A, numbered according to the EU index, compared to a wild-type IgG1 Fc region comprising the amino acid sequence of SEQ ID NO:11.

[0279] In one embodiment, the variant human IgG1 Fc region comprises the amino acid substitutions L234A and L235E, numbered according to the EU index, compared to a wild-type IgG1 Fc region comprising the amino acid sequence of SEQ ID NO:11.

[0280] In one embodiment, the mutated human IgG1 Fc region comprises the amino acid substitutions L234A, L235E and G237A, numbered according to the EU index, compared to a wild-type IgG1 Fc region comprising the amino acid sequence of SEQ ID NO:11.

[0281] In one embodiment, the mutated human IgG1 Fc region comprises an amino acid substitution at one or more positions selected from the group of A330 and P331, as numbered according to the EU index, compared to a wild-type IgG1 Fc region comprising the amino acid sequence of SEQ ID NO:11.

[0282] In one embodiment, the mutated human IgG1 Fc region comprises amino acid substitutions at one or more positions selected from the group of A330S and P331S, as numbered according to the EU index, compared to a wild-type IgG1 Fc region comprising the amino acid sequence of SEQ ID NO:11.

[0283] In one embodiment, the mutated human IgG1 Fc region comprises the amino acid substitutions A330S and P331S, numbered according to the EU index, compared to a wild-type IgG1 Fc region comprising the amino acid sequence of SEQ ID NO:11.

[0284] In one embodiment, the variant human IgG1 Fc region comprises the amino acid substitutions L234A, L235E, G237A, A330S and P331S, numbered according to the EU index, compared to a wild-type IgG1 Fc region comprising the amino acid sequence of SEQ ID NO:11.

[0285] In one embodiment, an antibody or antibody fragment according to the present disclosure comprises a mutated human IgG1 Fc region that comprises one or more amino acid substitutions that reduce or eliminate the binding affinity of the mutated Fc region to one or more Fc receptors and / or to C1q and / or reduce the ability of the antibody to induce effector function compared to a wild-type human IgG1 Fc region comprising the sequence of SEQ ID NO: 11, wherein the one or more amino acid substitutions are L234A, L235E, G237A, A330S and P331S, as numbered according to the EU index.

[0286] In one embodiment, the isolated antibody or antibody fragment according to the present disclosure is of human IgG1 class. In one embodiment, the isolated antibody or antibody fragment according to the present disclosure is of mutated human IgG1 class. In one embodiment, the isolated antibody or antibody fragment according to the present disclosure does not substantially induce effector function in vitro. In one embodiment, the mutated human IgG1 Fc region does not substantially bind to one or more Fc receptors and / or C1q. In one such embodiment, the isolated antibody or antibody fragment according to the present disclosure comprises an amino acid substitution selected from the group of L234A, L235E, G237A, A330S and P331S, as numbered according to the EU index, compared to a wild-type IgG1 Fc region comprising the amino acid sequence of SEQ ID NO: 11.

[0287] In one embodiment, the isolated antibody or antibody fragment according to the present disclosure is of the human IgG1 class, comprising the amino acid substitutions L234A, L235E, G237A, A330S and P331S, as numbered according to the EU index.

[0288] In one embodiment, the isolated antibody or antibody fragment according to the disclosure comprises the amino acid substitutions L234A, L235E, G237A, A330S and P331S, numbered according to the EU index.

[0289] In one embodiment, the isolated antibody or antibody fragment according to the disclosure comprises the amino acid substitutions L234A, L235E, G237A, A330S and P331S, numbered according to the EU index, compared to a wild-type IgG1 Fc region comprising the amino acid sequence of SEQ ID NO:11.

[0290] In one such embodiment, the isolated antibody or antibody fragment according to the present disclosure comprises the amino acid substitutions L234A, L235E, G237A, A330S and P331S, as numbered according to the EU index, and does not substantially induce effector function in vitro, hi one embodiment, the effector function is one or more selected from the group of CDC, ADCC and ADCP.

[0291] In one such embodiment, the isolated antibody or antibody fragment according to the disclosure comprises the amino acid substitutions L234A, L235E, G237A, A330S and P331S, as numbered according to the EU index, compared to a wild-type human IgG1 Fc region comprising the amino acid sequence of SEQ ID NO:11, and does not substantially bind to one or more Fc receptors and / or C1q in vitro.

[0292] In one embodiment, the isolated antibody or antibody fragment according to the present disclosure is of the human IgG1 class. In one embodiment, the isolated antibody or antibody fragment according to the present disclosure does not induce substantially effector function in vitro. In one embodiment, the isolated antibody or antibody fragment according to the present disclosure comprises one or more amino acid substitutions selected from the group of L234A, L235E, G237A, A330S and P331S as numbered according to the EU index.

[0293] In one embodiment, the isolated antibody or antibody fragment specific for human C5aR comprises: a) the HCDR1 region of SEQ ID NO: 27, the HCDR2 region of SEQ ID NO: 28, the HCDR3 region of SEQ ID NO: 29, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33 and the LCDR3 region of SEQ ID NO: 34, or b) the HCDR1 region of SEQ ID NO: 30, the HCDR2 region of SEQ ID NO: 31, the HCDR3 region of SEQ ID NO: 29, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33 and the LCDR3 region of SEQ ID NO: 34, or c) the HCDR1 region of SEQ ID NO: 27, the HCDR2 region of SEQ ID NO: 39, the HCDR3 region of SEQ ID NO: 40, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33 and the LCDR3 region of SEQ ID NO: 34, or d) the HCDR1 region of SEQ ID NO: 30, the HCDR2 region of SEQ ID NO: 41, the HCDR3 region of SEQ ID NO: 40, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33, and the LCDR3 region of SEQ ID NO: 34 Includes.

[0294] In one embodiment, the isolated antibody or antibody fragment specific for human C5aR comprises: a) a VH of SEQ ID NO: 35 and a VL of SEQ ID NO: 36, or b) a VH of SEQ ID NO: 42 and a VL of SEQ ID NO: 43, or VH and VL having at least at least 80%, at least 85%, at least 90% or at least 95% identity to VH of SEQ ID NO: 35 or 42 and to VL of SEQ ID NO: 36 or 43 Includes.

[0295] In one embodiment, the isolated antibody or antibody fragment specific for human C5aR comprises: a) HC of SEQ ID NO: 37 and LC of SEQ ID NO: 38, or b) HC of SEQ ID NO: 44 and LC of SEQ ID NO: 45, or HC and LC having at least at least 80%, at least 85%, at least 90% or at least 95% identity to the HC of SEQ ID NO: 37 or 44 and to the LC of SEQ ID NO: 38 or 45 Includes.

[0296] In one embodiment, said isolated antibody or antibody fragment specific for human C5aR is a monoclonal antibody or antibody fragment. In one embodiment, said isolated antibody or antibody fragment is a recombinant antibody or antibody fragment. In one embodiment, said isolated antibody or antibody fragment is a human, humanized or chimeric antibody or antibody fragment.

[0297] Binding of an antibody to an Fc receptor via its Fc region can be easily determined, for example, by ELISA or by surface plasmon resonance (SPR) using standard equipment such as a Biacore® instrument (GE Healthcare), and the Fc receptor can be obtained by recombinant expression. Alternatively, the binding affinity of the Fc region can be evaluated using a cell line known to express a particular Fc receptor, such as NK cells expressing the FcγIIIa receptor. The effector function of an antibody can be measured by methods known in the art. Suitable in vitro assays for assessing the ADCC activity of a molecule of interest are described, for example, in WO2012130831. Useful effector cells for such assays include peripheral blood mononuclear cells (PBMC) and natural killer (NK) cells. Alternatively or additionally, the ADCC activity of a molecule of interest can be evaluated in vivo in an animal model, such as, for example, that disclosed in Clynes et al., Proc Natl Acad Sci USA 95, 652-656 (1998). To assess complement activation, a CDC assay may be performed (see, e.g., Gazzano-Santoro et al., J Immunol Methods 202, 163 (1996); Cragg et al., Blood 101, 1045-1052 (2003); and Cragg and Glennie, Blood 103, 2738-2743 (2004)). A C1q binding assay (such as an ELISA) may be performed to determine whether an antibody is capable of binding to C1q and thus has CDC activity (see, e.g., WO 2006 / 029879). In vitro methods for assessing binding to Fc receptors or for assessing immune effector function are described in Examples 10-13 herein.

[0298] Fusion proteins An isolated antibody or antibody fragment according to the present disclosure may or may not be fused to one or more other amino acid residues, polypeptides or moieties. Such fusion proteins may be prepared as appropriate, including by genetic or chemical approaches. The linked moieties may contain secretion or leader sequences, sequences that facilitate detection, expression, separation or purification, or sequences that improve protein stability, for example during recombinant production. Non-limiting examples of possible moieties include beta-galactosidase, glutathione-S-transferase, luciferase, T7 polymerase fragments, secretion signal peptides, antibodies or antibody fragments, toxins, reporter enzymes, moieties capable of binding to metal ions such as polyhistidine tags, tags suitable for detection and / or purification, homo- or hetero-association domains, moieties that improve protein solubility or moieties that contain enzymatic cleavage sites.

[0299] Thus, an isolated antibody or antibody fragment according to the present disclosure may optionally contain one or more moieties for binding to other targets or target proteins of interest. It should be clear that such additional moieties may or may not provide additional functionality to the antibody and may or may not modify the properties of the isolated antibody or antibody fragment according to the present disclosure. Diagnostic applications

[0300] In one embodiment, the present disclosure provides the use of an isolated antibody or antibody fragment specific for human C5aR according to the present disclosure for disease diagnosis. In one embodiment, the present disclosure provides the use of an antibody or antibody fragment according to the present disclosure for detection of C5aR, in particular human C5aR and / or cynomolgus C5aR. In one embodiment, the present disclosure provides a method for detecting C5aR in a subject or sample, comprising contacting said subject or sample with an isolated antibody or antibody fragment specific for human C5aR of the present disclosure. In one embodiment, the present disclosure provides a method for diagnosing a disease in said subject, comprising contacting said subject or sample with an isolated antibody or antibody fragment according to the present disclosure.

[0301] In one embodiment, the isolated antibody or antibody fragment specific for human C5aR comprises: a) the HCDR1 region of SEQ ID NO: 27, the HCDR2 region of SEQ ID NO: 28, the HCDR3 region of SEQ ID NO: 29, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33 and the LCDR3 region of SEQ ID NO: 34, or b) the HCDR1 region of SEQ ID NO: 30, the HCDR2 region of SEQ ID NO: 31, the HCDR3 region of SEQ ID NO: 29, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33 and the LCDR3 region of SEQ ID NO: 34, or c) the HCDR1 region of SEQ ID NO: 27, the HCDR2 region of SEQ ID NO: 39, the HCDR3 region of SEQ ID NO: 40, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33 and the LCDR3 region of SEQ ID NO: 34, or d) the HCDR1 region of SEQ ID NO: 30, the HCDR2 region of SEQ ID NO: 41, the HCDR3 region of SEQ ID NO: 40, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33, and the LCDR3 region of SEQ ID NO: 34 Includes.

[0302] In one embodiment, the isolated antibody or antibody fragment specific for human C5aR comprises: a) a VH of SEQ ID NO: 35 and a VL of SEQ ID NO: 36, or b) a VH of SEQ ID NO: 42 and a VL of SEQ ID NO: 43, or VH and VL having at least at least 80%, at least 85%, at least 90% or at least 95% identity to VH of SEQ ID NO: 35 or 42 and to VL of SEQ ID NO: 36 or 43 Includes.

[0303] In one embodiment, the isolated antibody or antibody fragment specific for human C5aR comprises: a) HC of SEQ ID NO: 37 and LC of SEQ ID NO: 38, or b) HC of SEQ ID NO: 44 and LC of SEQ ID NO: 45, or HC and LC having at least at least 80%, at least 85%, at least 90% or at least 95% identity to the HC of SEQ ID NO: 37 or 44 and to the LC of SEQ ID NO: 38 or 45 Includes.

[0304] In one embodiment, said isolated antibody or antibody fragment specific for human C5aR is a monoclonal antibody or antibody fragment. In one embodiment, said isolated antibody or antibody fragment is a recombinant antibody or antibody fragment. In one embodiment, said isolated antibody or antibody fragment is a human, humanized or chimeric antibody or antibody fragment.

[0305] Treatment method The isolated antibodies or antibody fragments according to the present disclosure may be used in therapeutic methods. The antibodies or antibody fragments according to the present disclosure may be used for the treatment of diseases such as cancer, autoimmune diseases, or inflammatory diseases.

[0306] In one embodiment, the disclosure provides a method for the treatment of a disease.

[0307] In one embodiment, the disclosure provides an isolated antibody or antibody fragment according to the disclosure for the treatment of a disease. In one embodiment, the disclosure provides an isolated antibody or antibody fragment according to the disclosure for use in the treatment of a disease. In one embodiment, the disclosure provides an isolated antibody or antibody fragment according to the disclosure for use in the treatment of a disease in a subject in need of such treatment.

[0308] In one embodiment, the disclosure provides the use of an isolated antibody or antibody fragment according to the disclosure for the manufacture of a medicament. In one embodiment, the disclosure provides an isolated antibody or antibody fragment according to the disclosure for use as a medicament. In one embodiment, the disclosure provides an isolated antibody or antibody fragment according to the disclosure for use in a medicine. In one embodiment, the disclosure provides an isolated antibody or antibody fragment according to the disclosure for use as a medicament for the treatment of a subject in need of treatment.

[0309] In one embodiment, the disease is associated with the unwanted presence of C5aR, particularly human C5aR. In one embodiment, the disease is associated with the unwanted presence of C5a, particularly human C5a.

[0310] In one embodiment, the disease to be treated is a proliferative disease. In a particular embodiment, the disease is cancer. Non-limiting examples of cancer include bladder cancer, brain cancer, head and neck cancer, pancreatic cancer, lung cancer, breast cancer, ovarian cancer, uterine cancer, cervical cancer, endometrial cancer, esophageal cancer, colon cancer, colorectal cancer, rectal cancer, gastric cancer, prostate cancer, blood cancer, sarcoma, skin cancer, squamous cell carcinoma, bone cancer, melanoma, renal cell carcinoma, and kidney cancer.

[0311] In one embodiment, the disease to be treated is an autoimmune or inflammatory disease. Non-limiting examples of autoimmune or inflammatory diseases include rheumatoid arthritis (RA), psoriasis, psoriatic arthritis, systemic lupus erythematosus (SLE), lupus nephritis, type I diabetes, Graves' disease, inflammatory bowel disease (IBD), Crohn's disease (CD), ulcerative colitis (UC), irritable bowel syndrome, multiple sclerosis (MS), autoimmune myocarditis, Kawasaki disease, coronary artery disease, chronic obstructive pulmonary disease (COPD), interstitial lung disease, autoimmune thyroiditis, scleroderma, systemic sclerosis, osteoarthritis, atopic dermatitis, vitiligo, graft-versus-host disease, Sjögre's disease, and inflammatory bowel disease. These include, but are not limited to, idiopathic pulmonary disease, pulmonary edema, pulmonary fibrosis ...

[0312] In one embodiment, the present disclosure provides an isolated antibody or antibody fragment specific for human C5aR according to the present disclosure for use in a method of treating a subject having a disease comprising administering to the subject a therapeutically effective amount of an antibody or antibody fragment according to the present disclosure.

[0313] In one embodiment, the method further comprises administering to the subject a therapeutically effective amount of at least one additional therapeutic agent. The subject in need of treatment is generally a mammal, more specifically a human. For use in a method of treatment, the isolated antibody or antibody fragment according to the present disclosure is formulated, dosed, and administered in a manner compatible with good medical practice.

[0314] In one embodiment, the isolated antibody or antibody fragment specific for human C5aR comprises: a) the HCDR1 region of SEQ ID NO: 27, the HCDR2 region of SEQ ID NO: 28, the HCDR3 region of SEQ ID NO: 29, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33 and the LCDR3 region of SEQ ID NO: 34, or b) the HCDR1 region of SEQ ID NO: 30, the HCDR2 region of SEQ ID NO: 31, the HCDR3 region of SEQ ID NO: 29, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33 and the LCDR3 region of SEQ ID NO: 34, or c) the HCDR1 region of SEQ ID NO: 27, the HCDR2 region of SEQ ID NO: 39, the HCDR3 region of SEQ ID NO: 40, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33 and the LCDR3 region of SEQ ID NO: 34, or d) the HCDR1 region of SEQ ID NO: 30, the HCDR2 region of SEQ ID NO: 41, the HCDR3 region of SEQ ID NO: 40, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33, and the LCDR3 region of SEQ ID NO: 34 Includes.

[0315] In one embodiment, the isolated antibody or antibody fragment specific for human C5aR comprises: a) a VH of SEQ ID NO: 35 and a VL of SEQ ID NO: 36, or b) a VH of SEQ ID NO: 42 and a VL of SEQ ID NO: 43, or VH and VL having at least at least 80%, at least 85%, at least 90% or at least 95% identity to VH of SEQ ID NO: 35 or 42 and to VL of SEQ ID NO: 36 or 43 Includes.

[0316] In one embodiment, the isolated antibody or antibody fragment specific for human C5aR comprises: a) HC of SEQ ID NO: 37 and LC of SEQ ID NO: 38, or b) HC of SEQ ID NO: 44 and LC of SEQ ID NO: 45, or HC and LC having at least at least 80%, at least 85%, at least 90% or at least 95% identity to the HC of SEQ ID NO: 37 or 44 and to the LC of SEQ ID NO: 38 or 45 Includes.

[0317] In one embodiment, said isolated antibody or antibody fragment specific for human C5aR is a monoclonal antibody or antibody fragment. In one embodiment, said isolated antibody or antibody fragment is a recombinant antibody or antibody fragment. In one embodiment, said isolated antibody or antibody fragment is a human, humanized or chimeric antibody or antibody fragment.

[0318] Pharmaceutical Compositions In one embodiment, the disclosure provides a pharmaceutical composition comprising an isolated antibody or antibody fragment according to the disclosure and a pharma- ceutically acceptable carrier or excipient.

[0319] The pharmaceutical composition may further comprise at least one other pharma- ceutical active compound.The pharmaceutical composition according to the present disclosure may be used in the diagnosis, prevention and / or treatment of diseases associated with an undesirable presence of C5aR, particularly human C5aR.In particular, the present disclosure provides pharmaceutical compositions comprising an antibody or antibody fragment according to the present disclosure suitable for prophylactic, therapeutic and / or diagnostic use in mammals, more particularly in humans.

[0320] In general, an antibody or antibody fragment according to the present disclosure may be formulated as a pharmaceutical composition comprising at least one antibody or antibody fragment according to the present disclosure, at least one pharma- ceutically acceptable carrier or excipient, and, optionally, one or more additional pharma- ceutical active compounds. Such a formulation may be suitable for oral, parenteral, topical, or by inhalation administration. Thus, a pharmaceutical composition comprising at least one antibody or antibody fragment according to the present disclosure may be administered parenterally, such as intravenously or intramuscularly or subcutaneously. Alternatively, an antibody of the present invention may be administered orally or via a parenteral route, such as topically. In a preferred embodiment, a pharmaceutical composition comprising an antibody or antibody fragment according to the present disclosure is administered intravenously or subcutaneously.

[0321] In particular, the antibodies or antibody fragments according to the present disclosure may be used in combination with one or more pharma- ceutically active compounds that are or may be used for the prevention and / or treatment of a disease involving the target antigen of interest, which may or may not result in a synergistic effect. Examples of such compounds, as well as routes, methods and pharmaceutical formulations or compositions for their administration, will be apparent to the clinician.

[0322] In one embodiment, the present disclosure provides a pharmaceutical composition comprising an antibody or antibody fragment according to the present disclosure for use in the prevention and / or treatment of a disease associated with the unwanted presence of C5aR. In one embodiment, the present disclosure provides a pharmaceutical composition comprising an antibody or antibody fragment according to the present disclosure for use as a medicament. In one embodiment, the present disclosure provides a pharmaceutical composition comprising an antibody or antibody fragment according to the present disclosure for use in the prevention and / or treatment of an autoimmune disease and / or an inflammatory disease and / or cancer.

[0323] In one embodiment, the present disclosure provides a method for the treatment of an autoimmune disease and / or an inflammatory disease and / or cancer in a subject in need thereof using a pharmaceutical composition comprising an antibody or antibody fragment according to the present disclosure.

[0324] Further provided is a method of producing an antibody or antibody fragment according to the present disclosure in a form suitable for administration in vivo, the method comprising (a) obtaining an antibody or antibody fragment by a method according to the present disclosure and (b) formulating said antibody or antibody fragment with at least one pharma- ceutically acceptable carrier or excipient, whereby an antibody or antibody fragment preparation is formulated for administration in vivo. Pharmaceutical compositions according to the present disclosure comprise a therapeutically effective amount of one or more antibodies or antibody fragments according to the present disclosure dissolved in a pharma- ceutical acceptable carrier or excipient.

[0325] In one embodiment, the isolated antibody or antibody fragment specific for human C5aR according to the present disclosure comprises: a) the HCDR1 region of SEQ ID NO: 27, the HCDR2 region of SEQ ID NO: 28, the HCDR3 region of SEQ ID NO: 29, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33 and the LCDR3 region of SEQ ID NO: 34, or b) the HCDR1 region of SEQ ID NO: 30, the HCDR2 region of SEQ ID NO: 31, the HCDR3 region of SEQ ID NO: 29, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33 and the LCDR3 region of SEQ ID NO: 34, or c) the HCDR1 region of SEQ ID NO: 27, the HCDR2 region of SEQ ID NO: 39, the HCDR3 region of SEQ ID NO: 40, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33 and the LCDR3 region of SEQ ID NO: 34, or d) the HCDR1 region of SEQ ID NO: 30, the HCDR2 region of SEQ ID NO: 41, the HCDR3 region of SEQ ID NO: 40, the LCDR1 region of SEQ ID NO: 32, the LCDR2 region of SEQ ID NO: 33, and the LCDR3 region of SEQ ID NO: 34 Includes.

[0326] In one embodiment, the isolated antibody or antibody fragment specific for human C5aR comprises: a) a VH of SEQ ID NO: 35 and a VL of SEQ ID NO: 36, or b) a VH of SEQ ID NO: 42 and a VL of SEQ ID NO: 43, or VH and VL having at least at least 80%, at least 85%, at least 90% or at least 95% identity to VH of SEQ ID NO: 35 or 42 and to VL of SEQ ID NO: 36 or 43 Includes.

[0327] In one embodiment, the isolated antibody or antibody fragment specific for human C5aR comprises: a) HC of SEQ ID NO: 37 and LC of SEQ ID NO: 38, or b) HC of SEQ ID NO: 44 and LC of SEQ ID NO: 45, or HC and LC having at least at least 80%, at least 85%, at least 90% or at least 95% identity to VH of SEQ ID NO: 37 or 44 and to VL of SEQ ID NO: 38 or 45 Includes.

[0328] In one embodiment, said isolated antibody or antibody fragment specific for human C5aR is a monoclonal antibody or antibody fragment. In one embodiment, said isolated antibody or antibody fragment is a recombinant antibody or antibody fragment. In one embodiment, said isolated antibody or antibody fragment is a human, humanized or chimeric antibody or antibody fragment.

[0329] Antibody sequence

[0330] [Table 1]

[0331] [Table 2]

[0332] [Table 3]

[0333] [Table 4]

[0334]

Table 5

[0335]

Table 6

[0336]

Table 7

[0337]

Table 8

[0338]

Table 9

[0339]

Table 10

[0340]

Table 11

[0341]

Table 12

[0342]

Table 13

[0343]

Table 14

[0344]

Table 15

[0345] Example 1: Antigen production and quality control The amino acid sequences of C5aR and C5aR-related GPCRs from various species were retrieved from public sources (eg, Uniprot), verified, and generated in-house or by external vendors.

[0346] Synthetic Peptides A linear peptide covering the N-terminal extracellular region of human C5aR was used as antigen for initial panning and screening. The peptide was chemically synthesized (JPT) with a biotin tag, RP-HPLC purified, and delivered as lyophilized material. The lyophilized peptide was stored at -80°C. Alternatively, the peptide was conjugated to transferrin or bovine serum albumin (BSA).

[0347] [Table 16]

[0348] Linear peptides containing the N-terminal region of human and cynomolgus C5aR were used as antigens for subsequent binding studies. The peptides were chemically synthesized (Genscript) with a biotin tag, RP-HPLC purified, and delivered as lyophilized material. Lyophilized peptides were stored at -80°C. For reconstitution, peptides were dissolved in the desired volume of PBS and stored at -80°C.

[0349] [Table 17]

[0350] Recombinant Proteins C1q protein C1q protein purified from pooled normal human plasma was purchased from Complement Technology, Inc. (Catalog #A099).

[0351] Human C5a protein Recombinant human C5a was either purchased from R&D Systems (CAT#:2037-C5) or produced in-house.

[0352] For in-house production, DNA encoding the amino acids of human C5a (Uniprot:P01031|Lys679-Arg751) together with the N-terminal ompA signal sequence followed by a sequence encoding the maltose-binding protein (MBP), a FXa cleavage site, and a GS linker was cloned in frame into the pET21a expression vector (Novagen).

[0353] Human C5a (hC5a) was expressed in E. coli BL21(DE3) cells (Novagen) as an N-terminally tagged maltose binding protein (MBP) fusion protein. Protein expression was induced by addition of IPTG and cultures were further grown for 20-23 h. Cells were harvested by centrifugation and the pellet resuspended in lysis buffer (PBS buffer + 2 mM MgCl2, 20 U / ml Benzonase (Roche) and 1 tablet / 50 mL cOmplete, EDTA-free protease inhibitor cocktail tablet (Roche)). Cells were disrupted by either chemical lysis or high pressure homogenization. The resulting suspension was centrifuged and the supernatant was sterile filtered for further purification steps.

[0354] hC5a-MBP-fusion protein was purified by dextrin-sepharose affinity chromatography using MBP-Trap columns (GE-Healthcare) and optionally polished by cation exchange chromatography using Hi-Trap SP FF columns (GE LifeSciences). Purified MBP fusion was buffer exchanged into FXa-digestion buffer (20 mM Tris / HCl pH 8.0; 100 mM NaCl; 2 mM CaCl2) using PD10 columns (GE Healthcare). hC5a protein was released from maltose binding protein by addition of factor Xa (1:100 (w / w)) and incubated O / N on a rotary shaker at room temperature. Released hC5a was purified by cation exchange chromatography using Hi-Trap SP FF columns (GE LifeSciences). All affinity chromatography steps were performed using an AKTA Avant 25 preparative chromatography system.

[0355] Buffer exchange into PBS was performed using PD10 columns (GE Healthcare). Samples were sterile filtered and hC5a concentrations were determined by UV-spectrophotometry. Samples were analyzed for purity and integrity in denaturing, reducing or non-reducing SDS-PAGE, SEC-HPLC and mass spectrometry.

[0356] Fc gamma receptors (FcγR) and FcRn receptors DNA encoding the extracellular regions of human FcγRI, human FcγRIIa (131H), human FcγRIIa (131R), human FcγRIIb, human FcγRIIIa (158F), and human FcγRIIIa (158V) were cloned in frame with an N-terminal Vκ leader sequence and a C-terminal 6xHis-tag into the pMAX expression vector, a modified expression vector based on pcDNA3.1 (Thermo Fisher).

[0357] DNA encoding the extracellular domain of human, cynomolgus, mouse or rat FcRn large subunit p51 was cloned in frame with an N-terminal Vkappa leader sequence and a C-terminal AVI-6xHis-tag into the pMAX expression vector, a modified expression vector based on pcDNA3.1 (Thermo Fisher). Additionally, DNA encoding human, cynomolgus, mouse or rat FcRn small subunit p14 (=identical to beta-2 microglobulin) protein was cloned in frame into the second open reading frame with an N-terminal Vk leader sequence. The amino acid sequences of the receptors produced are summarized in Tables 6 and 7.

[0358] The HEK293-6E cell line was developed by the National Research Council of Canada (NRC). Cells were maintained in Freestyle F17 medium (Thermo Scientific) at 37°C and 6% CO2 in a humidified CO2-incubator. HKB11 (parent clone: ​​US Patent No. 6,136,599. J. Biomed. Sci. 2002;9:631-638) is a human hybrid cell line derived from the fusion of HEK293 human embryonic kidney and 2B8 Burkitt's lymphoma cells. HKB11#52 cells were maintained in MAC1.0 medium containing 1% FCS at 37°C and 6% CO2 in a humidified CO2 incubator.

[0359] Transient transfection was performed on HKB11#52 or HEK293-6E cells 1 day after seeding with commercial transfection reagents according to the manufacturer's instructions. Cells were cultured for 3 days and conditioned cell culture supernatants were harvested by centrifugation followed by sterile filtration (0.22 μm). Stably transfected HKB11#52 pools were generated by transfection of cells followed by selection with 800 μg / mL G418 (Thermo Scientific). Expression of antigens from stable pools was performed 4 days after seeding. Conditioned cell culture supernatants were harvested by centrifugation followed by sterile filtration (0.22 μm).

[0360] Individual proteins were purified by IMAC using a Protino Ni-NTA column (Macherey-Nagel). All chromatography steps were performed using an AKTA chromatography system (GE Healthcare). Samples were buffer exchanged into D-PBS using a PD10 column (GE Healthcare). In some cases, a preparative SEC step was performed using a Superdex 200 column (GE Healthcare) with polishing in D-PBS.

[0361] Biotinylation of FcRn heterodimers was performed by in vitro biotinylation using the BirA Kit (Avidity), followed by preparative SEC using a Superdex 200 column (GE Healthcare).

[0362] The quality of the samples was analyzed by denaturing, reducing or non-reducing SDS-PAGE, streptavidin-shift assay, HP-SEC and DLS.

[0363] [Table 18]

[0364] [Table 19]

[0365] Virus-like particles (VLPs) VLPs were produced in-house stably expressing human C5aR (D / K or N / N mutant) and one of the two naturally occurring variants of mouse C5aR as described in WO 2015 / 193143. All cloning experiments were performed using standard techniques. The antigen of interest was cloned in a suitable two-vector system for expression in mammalian cells, where one vector expresses a GAG and the other a GPCR-GAG fusion protein. Expression in these vectors is under the control of the CMV promoter. Host cells were then transfected with the two vectors. The resulting constructs were produced as fusion proteins, with the antigen of interest fused N-terminally to the GAG-protein (HV1B1 (Uni-Prot ID: P03347)). Protein expression and VLP production were performed under standard conditions in suspension culture. The host cells used in this experiment were HKB11 cells (ATCC; CRL-12568) and HEK cells (Life Technologies). Three days after gene transfection, the supernatant containing the VLPs was harvested and purified using standard procedures, including precipitation and ion exchange chromatography. The isolated proteins were subjected to SDS-PAGE chromatography. The supernatants were examined using a commercially available anti-GAG antibody or a commercially available anti-C5aR antibody, revealing that co-expression of GAG and GPCR-GAG fusion proteins resulted in higher expression levels of GAG and GPCR-GAG in the VLPs. This confirmed that the C5aR antigen was efficiently incorporated into the VLPs and that the C5aR antigen was detectable by the antibody. The amino acid sequences of the produced fusion proteins are summarized in Table 8.

[0366] [Table 20]

[0367] [Table 21]

[0368] cell line We generated CHO Flp-In cells stably expressing full-length human C5aR, cynomolgus monkey C5aR, mouse C5aR, rat C5aR and the C5aR-related GPCRs human C5L2, human C3aR, human FPR1 and human ChemR23. For the generation of Flp-In CHO cells, various vector constructs were gene-synthesized in-house and cells were transfected according to the instructor's manual (Thermofischer / Invitrogen). All constructs contained an N-terminal V5 / His tag. To confirm the expression of individual GPCRs on the surface of the cell lines, even in the absence of commercially available specific anti-GPCR tool antibodies, commercially available anti-His (e.g. Dianova CAT#DIA-910) or anti-V5 (e.g. AbD Serotec CAT#MCA2285GA) detection antibodies were used.

[0369] Example 2: Generation of human C5aR-specific antibodies from the HuCAL PLATINUM® library For antibody generation, the HuCAL Platinum® library was used to select antibodies with specificity for human C5aR. The HuCAL PLATINUM® library is a phagemid library based on the HuCAL concept (Knappik et al., (2000) J Mol Biol 296:57-86) and uses CysDisplay® technology to display Fabs on the phage surface (Lohning et al., WO 2001 / 05950).

[0370] To identify human C5aR-specific antibodies, a panning strategy was performed using human and cynomolgus C5aR antigenic material to select for species cross-reactive antibodies. Each panning run consisted of at least three individual selection rounds against different C5aR antigens (either as soluble recombinant antigens or overexpressed on cells).

[0371] Although the overall homology between cynomolgus and human C5aR is fairly high at 90%, the extracellular domains share only 75% protein sequence identity. Indeed, identification of cynomolgus C5aR cross-reactive antibodies has proven difficult, with the ancestor antibodies of MAB#1 and MAB#2 being among the few candidates that demonstrate specific cell binding to human C5aR expressed on cells, cross-reactivity to cynomolgus C5aR, and no binding to other related GPCRs.

[0372] Bead-based solution panning against peptides corresponding to the N-terminus (NT) of human C5aR identified the ancestor antibodies MAB#1 and MAB#2 that were specific for human and cynomolgus C5aR and capable of binding to full-length human C5aR expressed on cells.

[0373] To further improve affinity and specificity for human and cynomolgus C5aR, this clone was subjected to two successive rounds of affinity maturation panning using CHO Flp-In cells engineered to overexpress either human or cynomolgus C5aR. Furthermore, antibody engineering was performed to remove potential post-translational modification sites (PTM motifs) and for germline purposes to further improve specificity.

[0374] This final step in the engineering process identified MAB#1 and MAB#2 as potential therapeutic candidates, since both candidates are derived from the same ancestor and therefore share similar amino acid sequences and in vitro characteristics.

[0375] These two antibodies are further described in the Examples as outlined below.

[0376] Example 3: Production of human C5aR-specific antibodies Both MAB#1 and MAB#2 are of the human IgG1f isotype but have been modified in the Fc region to abolish the ability of the antibodies to mediate immune effector functions. The Fc region contains five amino acid substitutions compared to the wild-type human IgG1 Fc region: L234A, L235E, G237A, A330S, and P331S (h_IgG1f_AEASS), numbered according to the EU index.

[0377] This antibody consists of the heavy chain framework VH3-15 and the antibody light chain framework lambda1.

[0378] Transient Antibody Production - Advanced Micro Scale Production HKB11 Eukaryotic HKB11#52 cells were transiently transfected with mammalian expression vectors encoding the heavy and light chains of MAB#1 or MAB#2 (human IgG1_AEASS), respectively. Seven days after transfection, cell culture supernatants were harvested and subjected to Protein A affinity chromatography (MabSelect SuRe | GE Healthcare) using a liquid handling station. Samples remained in neutralized elution buffer (NaPS: 137 mM NaPhosphate, 81 mM NaCl, pH 7). Samples were sterile filtered (0.2 μm pore size). Protein concentrations were determined by UV-spectrophotometry at 280 nm and IgG purity was analyzed under denaturing, reducing conditions using CE-SDS (LabChip GXII | Perkin Elmer). UHP-SEC was performed to analyze IgG preparations in the native state.

[0379] Transient antibody production - discovery scale production in CHO CHO3-E7 cells were transiently transfected with mammalian expression vectors encoding the heavy and light chains of MAB#1 or MAB#2 (human IgG1_AEASS), respectively. Six days after transfection, cell culture supernatants were harvested and subjected to standard Protein A affinity chromatography (MabSelect SuRe | GE Healthcare). Unless otherwise stated, buffer exchange was performed into 1x Dulbecco's PBS (pH 7.2 | Invitrogen) and samples were sterile filtered (0.2 μm pore size). Protein concentration was determined by UV-spectrophotometry at 280 nm and IgG purity was analyzed under denaturing, reducing and non-reducing conditions using CE-SDS (LabChip GXII | Perkin Elmer). UHP-SEC was performed to analyze IgG preparations in the native state.

[0380] Results Transient production Data on production quality (SEC monomer content) and productivity of MAB#1 and MAB#2 are summarized in Table 9. Overall, acceptable monomer content (>95%) and yields (>55 mg / L in HKB11 cells) were achieved. Volumetric yields from CHO3E-7 transient research-scale production were also in the expected range.

[0381] [Table 22]

[0382] Generation of stable HKB11 pools To generate HKB11#52 pools stably expressing MAB#1 or MAB#2, a two-vector system was used for co-transfection.

[0383] To facilitate pool selection, these vectors contain Zeocin and Neocin resistance cassettes. The two vectors were transiently co-transfected in a 1:1 ratio into actively dividing HKB11#52 cells. One day after transfection, selection was initiated by the addition of 160 μg / mL Zeocin and 800 μg / mL Geneticin to the cell suspension. During selection, cell number and viability initially decreased. Cell harvesting was initiated 20 days after transfection. Upon reaching approximately 80% viability, stable pools were expanded to the desired volume depending on the amount required. Batch-produced cell culture supernatants were harvested on the sixth day after seeding.

[0384] Large-scale purification of MAB#1 and MAB#2 Purification of MAB#1 and MAB#2 from cell culture supernatants of stable pools of HKB11#52 was performed via Protein A affinity chromatography (MabSelect SURE | GE Healthcare) using 100 mM citric acid, 150 mM NaCl pH 3.5 as elution buffer. After 60 min incubation at pH 3.5, samples were neutralized. Buffer exchange to 150 mM histidine, pH 6.0 was performed and samples were sterile filtered (0.2 μm pore size). Protein concentration was determined by UV-spectrophotometry at 280 nm and IgG purity was analyzed under denaturing, reducing and non-reducing conditions using CE-SDS (LabChip GXII | Perkin Elmer). UHP-SEC was performed to analyze IgG preparations in the native state.

[0385] Results Large-scale production As summarized in Table 10, production of MAB#1 and MAB#2 resulted in favorable yields, purity, and integrity.

[0386] [Table 23]

[0387] Control antibodies Various control antibodies were generated and included in the experiments for comparison purposes.

[0388] RefMAB#1: Benchmark antibody Nucleotide sequences encoding the VH and VL regions from the human C5aR specific antibody "IPH5401" were retrieved from US Patent Publication No. 2013 / 0295116 (NOVO NORDISK-clone 32F3A6GL). The nucleotide sequences were gene synthesized as linear DNA fragments with appropriate flanking regions (e.g. appropriate restriction enzyme recognition sites, linker sequences) either in-house or by external vendors. By using standard molecular biology methods, the DNA fragments were cloned into appropriate mammalian IgG expression vectors encoding the heavy and light chains of human IgG1_AEASS as described above. RefMAB#1 was transiently produced as described above. The heavy and light chain amino acid sequences of RefMAB#1 are shown in Table 3.

[0389] Additional control antibodies: An in-house negative isotype control antibody with specificity against hen egg lysozyme (MOR03207) as well as an in-house positive control antibody with specificity against the N-terminus of human C5aR (anti-C5aR antibody) in either human IgG1 or human IgG1_AEASS format were transiently produced as described above.

[0390] Characterization of the binding properties of MAB#1 and MAB#2 Example 4: Monovalent affinity determination for MAB#1 and MAB#2 against C5a RN-terminal peptide using SPR method K via IgG capture setting at 25 °C using a Biacore T200 instrument (Biacore, GE Healthcare). DDeterminations were performed. Approximately 500 RU of IgG diluted in HBS-EP+, pH 7.4 was captured on a CM5 chip (Biacore, GE Healthcare) immobilized with anti-human Fc antibody (GE Healthcare) using standard EDC-NHS amine coupling chemistry. The reference flow cell 1 was simply activated and deactivated. Kinetic measurements were performed using six different human or cynomolgus C5aR_NT-bio peptide concentrations (SEQ ID NO: 13 or SEQ ID NO: 14, respectively) (2n serial dilutions, 2000-62.5 nM) (injection time 300 s; dissociation time 600 s; flow rate 30 μL / min) with HBS-EP+ (GE Healthcare) as running buffer. After each cycle, the sensor chip was regenerated with 3 × 20 s injections of 3 mM MgCl2 to remove bound peptide / antibody complexes. A blank injection of running buffer was used for double referencing. k on and k off To determine the kinetic constants, all sensorgrams were fitted using Biacore T200 Evaluation Software 3.1 (Biacore, GE Healthcare) and these were used to calculate the K D The raw data were fitted to a 1:1 binding model to calculate the parameter R max was set to local and RI was set to 0.

[0391] result The results are summarized in Table 11. Both antibodies showed similar binding to the human C5aR peptide, with D values ​​were in the low double-digit nanomolar range, revealing weaker binding to the cynomolgus C5aR peptide.

[0392] [Table 24]

[0393] Example 5: Apparent affinity (bivalent) determination of MAB#1 and MAB#2 for C5a RN-terminal peptide using Octet method Apparent K via the C5aR_NT-bio peptide capture setup was measured using an Octet HTX instrument (ForteBIO, Pall Life Sciences). D Determinations were performed at 27°C. Approximately 0.03 nm of peptide diluted in PBS, pH 7.4 was loaded onto a streptavidin (SA) sensor (ForteBIO, Pall Life Sciences). Kinetic measurements were performed using seven different IgG concentrations (3-fold serial dilutions, 200-0.27 nM for human C5aR_NT-bio peptide (SEQ ID NO: 13) and 1000-1.4 nM for cynomolgus C5aR_NT-bio peptide (SEQ ID NO: 14) in Octet buffer (PBS, 0.05% (v / v) Tween-20, 0.1% (w / v) BSA) with 480 s association time and 900 s dissociation time. After each dissociation step, the sensor was regenerated to remove bound antibody (3 × 30 s Gly / HCl, pH 1.5). The apparent k on and apparent k off To determine the rate constants, all sensorgrams were fitted using Octet Data Analysis Software 10.0 (ForteBIO) and these were used to calculate the apparent K D was calculated (using a 1:1 binding model).

[0394] result The results are summarized in Table 12. Both antibodies had apparent K D The binding to the cynomolgus monkey C5aR peptide was approximately 1000-fold lower. The weaker binding observed to the cynomolgus monkey C5aR peptide was mainly due to the fast k off In terms of apparent affinity for human C5aR_NT, MAB#2 showed approximately 10-fold reduced binding affinity when compared to MAB#1 (K D :290pM vs. 20pM).

[0395] [Table 25]

[0396] Example 6: Apparent affinity (bivalent) determination for MAB#1 on full-length C5aR expressed on cells by using KinExA Since C5aR belongs to the GPCR family, it is very difficult to generate recombinant full-length antigenic material that can be used for SPR measurements. Therefore, we performed KinExA measurements using Flp-In CHO cells stably expressing human C5aR or cynomolgus C5aR.

[0397] method Apparent K in C5aR-expressing cells at RT using a KinExA3200 instrument (Sapidyne Instruments) D The determination was performed. PBS (Gibco) supplemented with BSA (1 mg / mL) and 0.02% (v / v) NaN3 was used as the assay buffer. MAB#1 (concentrations: 2 nM and 30 pM) was used as the analyte and Flip-In CHO hC5aR_V5 / His cells (final concentrations 3 Mio cells / mL and 1 Mio cells / mL, 2n serial dilutions) or Flip-In CHO cyC5aR_V5 / His cells (final concentrations 25 Mio cells / ml and 6 Mio cells / ml, 2n serial dilutions) as the titrants, equilibrated overnight at RT in an overhead shaker. After equilibration, the samples were centrifuged and the supernatant was used for analysis. Free concentration of MAB#1 was determined using polymethylmethacrylate (PMMA) beads coated with MabSSL (GE Healthcare) and anti-human Fab2 Alexa Fluor 647 (500 ng / mL) was used for detection. Using KinExA software, single K D The apparent K value is calculated by "n-curve analysis" which simultaneously fits all of a certain curve to the D obtained.

[0398] result The results are summarized in Table 13. MAB#1 has an apparent K DBinding to full-length cynomolgus monkey C5aR revealed strong binding with an apparent K of approximately 3 nM. D Similar findings regarding binding to cynomolgus C5aR were previously observable in Octet and Biacore measurements (see Examples 4 and 5).

[0399] [Table 26]

[0400] Example 7: Binding of MAB#1 and MAB#2 to full-length C5aR expressed on Flp-In CHO cells and whole blood-derived neutrophils (FACS analysis) Cells were examined via FACS for binding to CHO Flp-In cell lines stably expressing various full-length C5aR antigens and associated GPCRs, as well as to purified human or cynomolgus neutrophils endogenously expressing human or cynomolgus C5aR. Cynomolgus neutrophils were obtained from whole cynomolgus blood from three different animals (LPT Hamburg).

[0401] method C5aR-CHO Flp-In cell lines were blocked and IgG was added either at serial dilutions or at a single (high) concentration of 300 nM. For detection of IgG binding, R-phycoerythrin (R-PE)-conjugated anti-human IgG (Fc-gamma fragment specific)2 antibody was added and fluorescence was measured using a FACS array or Novocyte instrument.

[0402] Neutrophils were purified from EDTA whole blood samples using the MACSexpress Neutrophil Isolation Cocktail from Miltenyi Biotec. Briefly, the kit was used to isolate cells using magnetic labeling and negative magnetic separation. After removal of red blood cells, the cells were stained by adding serial dilutions of IgG followed by a detection antibody specific for Alexa Fluor 647-conjugated anti-human F(ab)2 fragments. Measurements were performed on a FACS array instrument. FACS data were evaluated using FlowJo and entered into GraphPad Prism (v4.0) and fitted to a sigmoidal dose-response curve using nonlinear regression to calculate EC50.

[0403] result The results are summarized in Table 14 and Figures 1 and 2. Figures 1A and 1C show the binding of MAB#1, MAB#2 and RefMAB#1 to human and cynomolgus C5aR present on engineered CHO cells expressing the respective full-length receptors. Overall, EC 50 Highly comparable binding curves to human C5aR at nearly identical concentrations were determined for MAB#1, MAB#2 and RefMAB#1. Similar results were obtained for MAB#1 and MAB#2 to cynomolgus monkey C5aR. As expected, RefMAB#1 demonstrated no binding to cynomolgus monkey C5aR expressed on CHO cells.

[0404] Binding to cynomolgus and human C5aR was also confirmed using purified neutrophils from human or cynomolgus whole blood (Figures 1B and D). Again, EC 50 Very similar binding curves with almost identical values ​​were observable for MAB#1. Interestingly, MAB#2 revealed an overall lower signal to background levels in cynomolgus monkey neutrophils when compared to MAB#1. This finding was not observable when analyzing binding to cynomolgus monkey C5aR expressed on CHO cells. For RefMAB#1, the lack of binding to cynomolgus monkey neutrophils was confirmed.

[0405] Due to the low sequence homology to rodent C5aR (66% overall identity), no cross-reactivity to rat and mouse C5aR was expected. Indeed, when tested at an IgG concentration of 300 nM, neither MAB#1 nor MAB#2 showed any significant binding to rat or mouse C5aR expressed on CHO-Flp cells (Figure 2). To exclude cross-reactivity to any other members of the C5aR subfamily, binding to full-length human C5L2, ChemR23, FPR1 and C3aR expressed on CHO Flp-In cells via FACS was also determined (compared to non-transfected parental CHO Flp-In cells). As shown in Figure 2, even at an IgG concentration of 300 nM, no significant cellular binding of MAB#1 and MAB#2 to any of the C5aR-related GPCRs or to parental CHO cells was detectable.

[0406] Taken together, both MAB#1 and MAB#2 demonstrated specific binding to human and cynomolgus C5aR.

[0407] [Table 27]

[0408] Example 8: Binding of MAB#1 and MAB#2 to full-length human C5aR displayed on virus-like particles (VLPs) - Binding to naturally occurring variants of human C5aR Two naturally occurring mutants of human C5aR (SEQ ID NO:1; D / K mutant and SEQ ID NO:2; N / N mutant) are reported.

[0409] To compare the binding of MAB#1 and Mab#2 to the two naturally occurring variants, VLPs expressing either of the two C5aR variants were generated as described in Example 1. As a negative control, VLPs expressing mouse C5aR were included, since MAB#1 and Mab#2 do not cross-react with mouse C5aR.

[0410] method For binding assessment, generated VLPs were coated overnight and, after a blocking step, IgG titrations were added the next day. Binding of IgG to the coated antigen was detected using alkaline phosphatase-conjugated anti-human IgG2 antibody and AttoPhos as substrate.

[0411] result The results are summarized in Table 15 and shown in Figure 3. Both MAB#1 and MAB#2 demonstrated comparable titration curves against both natural variants of human C5aR, with EC 50 The concentrations were equal. As expected, no binding to mouse C5aR was detected. Based on these data, high affinity binding of MAB#1 and MAB#2 to human C5aR in vivo can be predicted, independent of the natural variants present on the respective target cells.

[0412] [Table 28]

[0413] Example 9: Protein Panel Profiling (3P) Potential non-specific off-target binding to MAB#1 was determined in a general 3P assay.

[0414] method: Protein panel profiling was performed primarily as described by Frese et al. (mAbs 5:2, 279-287; March / April 2013). Thirty-two different proteins and controls were coated on two 384-well MSD standard plates at a concentration of 1.0 μg / mL overnight at 4° C. The coating solution was discarded and the plates were blocked with 50 μL 3% (w / v) BSA in PBS for 1 h at RT on a microtiter plate shaker (approximately 500 rpm), followed by washing three times with 50 μL wash buffer (PBS with 0.05% (v / v) Tween 20). IgG samples were diluted to 100 nM and 10 nM in assay buffer (PBS with 0.5% (w / v) BSA, 0.05% (v / v) Tween 20). As controls, isotype control antibody MOR03207 (IgG1f_AEASS) and assay buffer were used. Samples and controls were added at 30 μL / well and incubated for 3 h at RT on a microtiter plate shaker. Plates were washed 3 times and 30 μL detection antibody (ECL-labeled anti-human Fab) was added per well and incubated for 1 h on a microtiter plate shaker (approximately 500 rpm). After washing of the MSD plate and addition of 35 μL / well of MSD Read Buffer T with detergent, electro-chemiluminescence signals were detected using a SECTOR Imager S600 instrument (Meso Scale Diagnostics).

[0415] For evaluation, the binding signal of an antibody sample for a given protein was divided by the individual binding signal of the reference antibody MOR03207, resulting in a binding ratio (BR).The cumulative binding ratio (CBR) of all proteins excluding the controls (total of 25) was then calculated: a CBR of 150 or less indicates an IgG with no detectable non-specific binding. A value above 150 indicates an IgG with increased non-specific binding compared to the reference antibody MOR03207.

[0416] result The results for MAB#1 and MAB#2 are summarized in Figure 10. In summary, there was no significant non-specific binding detectable to any of the proteins tested. Very low (MAB#2) and low (MAB#1) binding to bovine transferrin was observed. Binding to bovine transferrin was confirmed in an Octet-based binding assay, but no binding to rat or cynomolgus transferrin was detected (data not shown). Therefore, the observed binding to bovine transferrin was deemed insignificant.

[0417] Final antibody format and safety C5aR is expressed on various immune cells such as leukocytes, neutrophils and lymphocytes, and human IgG1 Fc-mediated depletion of such cells must be prevented to avoid unwanted side effects. Therefore, the final IgG format of the anti-C5aR antibody must be inhibitory for its ability to induce any effector function during therapeutic intervention.

[0418] Both MAB#1 and MAB#2 contain five amino acid substitutions in the Fc region of human IgG1f, namely L234A, L235E, G237A, A330S and P331S (hIgG1f_AEASS, numbered according to the EU index), to abolish antibody-induced effector functions. The clinical safety of this format for C5aR antibody therapy has been described in the art (Wagner F et al., Annals of the Rheumatic Diseases. 2014;73:499. doi:10.1136 / annrheumdis-2014-eular.2156).

[0419] As outlined below, the lack of effector function induction ability of MAB#1 and MAB#2 was confirmed in various assays, including binding studies to Fcγ receptors or C1q and in vitro ADCC and ADCP assays.

[0420] Example 10: Binding of MAB#1 and MAB#2 to the FcRn receptor using Octet method Apparent K values ​​for immobilized neonatal Fc receptors (FcRn) from different species were measured using an Octet HTX instrument (ForteBIO, Pall Life Sciences). D Determinations were performed at pH 6.0 and 7.2 at 27 °C. Biotinylated human, cynomolgus, mouse and rat FcRn was captured at 0.5 nm on a streptavidin (SA) sensor (ForteBIO, Pall Life Sciences). Kinetic measurements were performed using eight different concentrations of IgG (3n serial dilutions, 1000–0.46 nM) in Octet buffer (PBS, 0.05% / v / v) Tween-20, 0.1% (w / v) BSA, with 240 s association time and 180 s dissociation time. After each cycle, the sensor was regenerated to remove bound ligand / antibody complexes (2 × 30 s in HBS-EP+, pH 8.0). To determine the apparent affinity, all sensorgrams were fitted using Data Analysis Software 10.0 (ForteBIO, Pall Life Sciences) and the data were fitted with a steady-state model.

[0421] result The results are summarized in Table 16. MAB#1 and MAB#2 revealed apparent binding affinities to FcRn from different species in the expected affinity range (comparable to isotype control antibody MOR03207 IgG1f, and for both IgG molecules physiological binding behavior for binding to human FcRn could be confirmed, i.e., no binding at neutral pH (7.2) was detectable). Thus, the five mutations introduced into the Fc region of the antibodies did not adversely affect human FcRn binding.

[0422] [Table 29]

[0423] Example 11: Binding of MAB#1 and MAB#2 to human Fcγ receptors using Octet method Using an Octet (ForteBIO, Pall Life Sciences), K via IgG capture setup at 27 °C D Determinations were performed. 2.0 nM IgG diluted in Octet assay buffer (PBS, 0.05% (v / v) Tween-20, 0.1% (w / v) BSA) was captured on a Protein A sensor (ForteBIO, Pall Life Sciences). Kinetic measurements were performed using seven concentrations (2n serial dilutions) of Fc gamma receptor in assay buffer. After each cycle, the sensor was regenerated (2 × 30 s in 10 mM Gly / HCl, pH 1.5) to remove bound ligand / antibody complexes. All sensorgrams were fitted and k calculated using Data Analysis Software 10.0 (ForteBIO, Pall Life Sciences). on and k off Determine the rate constant, K D was used to calculate the parameter R max We set locally and fitted the 1:1 binding model to the raw data.

[0424] result The results are summarized in Table 17. No or only very weak binding of MAB#1 and MAB#2 could be detected to any of the Fcγ receptors tested, confirming that the mutations introduced into the human IgG1 Fc region were effective in abolishing Fcγ receptor binding.

[0425] [Table 30]

[0426] Example 12: Binding of MAB#1 and MAB#2 to C1q using Octet method Apparent (bivalent) K via IgG capture setup at 27 °C using Octet (Fortebio Pall Life Sciences) D Determinations were performed. 2.0 nM IgG diluted in Octet buffer was captured on anti-hu Fab CH1 kappa / lambda (BAC) immobilized on a streptavidin (SA) sensor (ForteBIO, Pall Life Sciences). Kinetic measurements were performed using eight concentrations of C1q (3n serial dilutions, 500–0.69 nM) in Octet assay buffer (see above) with 240 s association and 240 s dissociation times. After each cycle, the sensor was regenerated (2 × 50 mM NaOH, 1 × 10 mM Gly / HCl, pH 1.5, 30 s each) to remove bound ligand / antibody complexes. To determine the apparent affinity, all sensorgrams were fitted using Data Analysis Software 9 (ForteBIO, Pall Life Sciences). The data were fitted with a steady-state model.

[0427] result The results are summarized in Table 18. As expected, no binding of MAB#1 and MAB#2 to C1q (isolated from pooled human plasma) was observable, confirming that the mutations introduced into the human IgG1 Fc region were effective in abolishing C1q binding.

[0428] [Table 31]

[0429] Example 13: ADCC and ADCP in vitro activity of MAB#1 method ADCC and ADCP activity against MAB#1 was tested using Promega ADCC and ADCP reporter bioassays according to the manufacturer's instructions (Cat#G7017 and Cat#G988A, respectively). The kit uses modified Jurkat cells as effector cells. The cells stably express either the FcyRIIIa receptor for ADCC, the V158 (high affinity) mutant and the FcγRIIa_H receptor for ADCP, and the NFAT response element driving firefly luciferase expression. As target cells, CHO Flp-In cells expressing human C5aR were used. Binding of the effector cells to the target through an antibody bridge (e.g., through MAB#1 or MAB#2) initiates a cascade of events in the NFAT pathway, resulting in the expression of firefly luciferase protein. This enzymatic reaction produces light that is proportional to the luciferase concentration, which directly correlates with ADCC or ADCP activity. ADCC or ADCP activity was analyzed for MAB#1 by plotting the average signal against the background value.

[0430] For MAB#1, wild-type human IgG1f version was not available, so wild-type IgG1f as well as Fc-silent human IgG1f_AEASS version of in-house human anti-C5aR control antibody were included as positive controls. Additionally, Fc-silent (hIgG1_AEASS) and wild-type (hIgG1f) versions of isotype control antibody MOR03207 were included as negative controls.

[0431] result The results are summarized in Table 19 and shown in Figures 8A (ADCP) and B (ADCC) for an IgG concentration of 10 μg / ml. MAB#1, like the Fc-silent version of the anti-C5aR control antibody and the Fc-silent version of MOR03207, did not induce FcγRIIIa or FcγRIIa_H activation of the NFAT pathway in engineered effector cells in the presence of C5aR-overexpressing CHO cells. The wild-type (non-silent) version of the C5aR-specific control antibody clearly induced luciferase production in engineered Jurkat cells in the presence of C5aR-expressing CHO cells.

[0432] Taken together, this experiment clearly confirmed that the mutations introduced into the wild-type human IgG1 Fc region were efficient in preventing ADCC and ADCP activity.

[0433] [Table 32]

[0434] Functional characterization of MAB#1 and MAB#2 The neutralizing activities of MAB#1 and MAB#2 were analyzed in a different in vitro assay that monitors C5a-induced activation of the C5aR.

[0435] Since high levels of C5a have been described under pathophysiological conditions, the ability of C5aR antagonistic antibodies to neutralize the high concentrations of C5a that may be locally present at disease sites is expected to provide beneficial therapeutic effects in vivo. Therefore, in vitro experiments to reflect such in vivo pathological conditions were also set up.

[0436] Example 14: PathHunter® β-Arrestin Assay (DiscoveRx) method: The PathHunter® β-arrestin assay from DiscoveRx was performed according to the manufacturer's instructions. Briefly, human C5a-induced human C5aR activity was measured by detecting the interaction of activated C5aR with β-arrestin using β-galactosidase enzyme fragment complementation.

[0437] Recombinant human C5a was used to induce β-arrestin recruitment and enzyme activity was measured using chemiluminescence detection reagents from DiscoverRx. Chinese Hamster Ovary (CHO) cells expressing a modified version of human C5aR were seeded overnight and serial dilutions of human C5a (R&D Systems) were added and incubated for 1.5 h at 37° C. and 5% CO2 (titration curves were generated in the absence of antagonist). In parallel, titration curves of human C5a were determined in the presence of C5aR-specific antibodies (fixed IgG concentration 50 nM). To do so, cells were added with IgG and then stimulated with human C5a and incubated for 1 h at 37° C. and 5% CO2. Results were expressed as relative luminescence units. Titration curves for human C5a in the presence and absence of antagonistic IgG were generated via GraphPad Prism.

[0438] For the β-arrestin assay, the shift in the dose-response curves to human C5a towards higher doses (i.e., horizontally to the right on the dose axis) was compared for MAB#1, MAB#2 and RefMAB#1 (Figure 4A). Additionally, the % inhibition at three increasing C5a concentrations (1.2 nM, 11 nM and 100 nM) was calculated and compared (Figure 4B).

[0439] result The results from the β-arrestin assay are shown in Figures 4A and 4B and summarized in Tables 20 and 21. In the absence of antibody, the mean EC 50 A dose-response curve was obtained to human C5a at a concentration of 2.9 nM (FIG. 4A).

[0440] Addition of MAB#1 or MAB#2 to human C5a at a final IgG concentration of 50 nM resulted in a significant shift of the dose-response curve by more than 12-fold to higher doses of human C5a. More precisely, in the presence of 50 nM MAB#1 or MAB#2, the mean EC 50 A dose-response curve for human C5a at a concentration of 37 nM was obtained (Figure 4A, Table 20). In other words, the presence of MAB#1 or MAB#2 significantly reduced the ability of human C5 to induce C5aR activation, or alternatively, in the presence of MAB#1 or MAB#2, human C5a had to be added at least 12-fold higher concentrations to induce the same C5aR activity when compared to its activity in the absence of antibody.

[0441] This observation was also reflected when the % inhibition at three increasing C5a concentrations was calculated (Figure 4B). At 1.2 nM human C5a, all three tested antibodies, MAB#1, MAB#2 and RefMAB#1, revealed comparable inhibition of C5a-induced C5aR activity by more than 80% at an IgG concentration of 50 nM. However, when a 10-fold higher concentration of human C5a (11 nM) was used to activate C5aR, RefMAB#1 was only able to neutralize about 50% of C5a-induced C5aR activity. This was quite different from MAB#1 and MAB#2, which were still able to neutralize up to 80% of C5a-induced C5aR activity.

[0442] This effect was even more pronounced when a 100-fold higher concentration of human C5a (100 nM) was used for C5aR activation, where barely detectable neutralizing activity was observed for RefMAB#1, whereas MAB#1 and MAB#2 were still able to neutralize approximately 40% and 30% of human C5a-induced C5aR activity, respectively.

[0443] Thus, both MAB#1 and MAB#2 are effective in neutralizing pathophysiological C5a concentrations in vitro and are significantly more potent compared to RefMAB#1.

[0444] [Table 33]

[0445] [Table 34]

[0446] Example 15: Inhibition of neutrophil and monocyte activation by MAB#1 and MAB#2 - CD11b assay The neutralization potency of MAB#1 and MAB#2 was further determined in a functional CD11b whole blood assay, which corresponds to a more physiological setting. CD11b combines with CD18 to form the integrin Mac-1 complex, which acts as a multi-ligand receptor. CD11b is constitutively expressed on the surface of >50% of peripheral blood leukocytes; upon leukocyte activation, its expression is upregulated through the fusion of CD11b-containing secretory granules to the cell membrane. Thus, CD11b expression is widely used as a marker of leukocyte activation both in vivo and in vitro.

[0447] C5a, as a potent activator of human neutrophils and monocytes, induces the upregulation of the surface antigen CD11b. Therefore, the ability of MAB#1 and MAB#2 to prevent C5a-induced activation of granulocytes and monocytes was examined by evaluating CD11b levels in whole blood-derived granulocytes and monocytes. This experiment was essentially performed as described in US Patent Publication No. 2013 / 0295116 (NOVO NORDISK).

[0448] method In the first assay setup, heparinized whole blood was mixed with IgG (serial dilutions) and incubated for 20 min at 37° C., 5% CO2. Human C5a was added at a standard concentration of 15 nM and incubated for 20 min at 37° C., 5% CO2. Anti-CD11b-PE or isotype control antibody MOR03207 was added and the plate was incubated for 20 min at 37° C., 5% CO2. Finally, red blood cell lysis buffer was added and incubated for 15 min at room temperature in the dark, and the cells were washed and resuspended again in lysis buffer.

[0449] In a second experimental setup, human C5a was added at the more clinically relevant pathophysiological concentration of 150 nM, as described above, without modifying the remaining assay setup.

[0450] To examine the receptor residence time of MAB#1, the assay was further adapted by extending the incubation time of heparinized whole blood with IgG from 20 to 300 min.

[0451] Fluorescence was measured using a FACS array or Novocyte instrument. Samples were gated to exclude dead cells and debris. Monocytes and granulocytes were identified and gated according to their FSC and SSC profiles. The median fluorescence intensity (MFI) of gated granulocytes and / or monocytes in the CD11b-PE channel (Yellow-A) was calculated. Results were expressed as a percentage of inhibition (% inhibition). Maximal CD11b expression (MFI Max ) was the mean MFI of cells incubated with C5a but without IgG. Minimum (background) CD11b expression (MFI Min ) was the average MFI of cells incubated without C5a and without IgG. The formula used to calculate the % inhibition for each sample was as follows: % Inhibition = 100-(((MFI 試料 -MFI Min )) / ((MFI Max -MFI Min ))×100)

[0452] Data were evaluated using FlowJo, entered into GraphPad Prism (v4.0), and fitted to a sigmoidal dose-response curve using nonlinear regression to determine IC 50 was calculated.

[0453] Results for MAB#1 using 15nM C5a ligand Results for MAB#1 from the CD11b whole blood assay are summarized in Table 22. IC in the single digit nM range for both gated cell populations (monocytes and granulocytes). 50Values ​​were determined with a maximal inhibition of approximately 88% for granulocytes and 83% for monocytes.

[0454] [Table 35]

[0455] Results for MAB#1, MAB#2 and RefMAB#1 using 15nM vs. 150nM C5a ligand In addition to the addition of 15 nM human C5a for stimulation of granulocytes (as above), a 10-fold higher concentration (150 nM) of ligand was used to mimic the higher pathophysiological ligand concentration.

[0456] Again, dose titrations of MAB#1, MAB#2 and RefMAB#1 were performed and inhibition curves were generated using GraphPad Prism via nonlinear regression function, and the results were quantitatively expressed as "antagonist concentration required to induce 50% inhibition."

[0457] The results from this CD11b whole blood assay are shown in Figures 5A and B.

[0458] Figure 5A shows the neutralizing activity of MAB#1, MAB#2 and RefMAB#1 at 15 nM C5a, and Figure 5B shows the individual neutralizing activities at 150 nM C5a. As a quantitative readout, the IgG concentration to achieve 50% inhibition of CD11b upregulation was calculated and the results are shown below the x-axis in both figures.

[0459] Overall, similar observations were made for the β-arrestin assay (Example 14) when using increasing C5a concentrations in a CD11b whole blood assay gated on granulocytes as the initial target cells. To block the effect of 15 nM human C5a by 50%, an IgG concentration of 12 nM was required for MAB#1 and RefMAB#1, whereas an IgG concentration of 17 nM was sufficient for MAB#2. However, significantly different amounts of antagonist were required to block 50% of the CD11b upregulation induced by a 10-fold higher C5a concentration (150 nM). For MAB#1, an IgG concentration of 42 nM was now sufficient to inhibit 50% of the C5a-induced activation of C5aR, whereas a 7-fold higher concentration of RefMAB#1 (291 nM) was required to achieve the same blocking effect. For MAB#2, an IgG concentration of 114 nM was sufficient.

[0460] Thus, a ranking of the three IgGs with respect to potency (MAB#1>MAB#2>RefMAB#1) could be made, similar to what was already seen in the b-arrestin assay, with both MAB#1 and MAB#2 appearing to be highly effective in neutralizing pathophysiological C5a concentrations in vitro, being significantly more potent compared to RefMAB#1.

[0461] Results for MAB#1 - Effect on Receptor Residence Time Seow and coworkers (Seow V et al., Sci. Rep. 2016;6:24575) reported that C5a generation can be localized to the cell membrane and be significantly higher for short but repeated periods of time. It was therefore suggested that antagonists of C5aR with longer residence times may be advantageous in systems with rapid transient signaling. It was also concluded that increased receptor residence time measured in vitro may translate into duration of action and degree of efficacy in vivo.

[0462] To compare the receptor residence time of the two antibodies in a similar setting as published by Seow and coworkers, log dose inhibition curves over 20 and 300 min of IgG incubation with granulocytes and monocytes present in whole blood were assessed as described above.

[0463] Results from this experimental setup of the CD11b whole blood assay are shown in Figures 6A-D. Surprisingly, a significant increase in neutralizing activity over extended incubation times was observable with MAB#1. As shown in Figures 6A and C, the IC calculated for MAB#1 for both granulocyte and monocyte populations was 0.01. 50 The concentration decreased to about one-sixth over time (IC 50 (Values ​​are summarized in Table 23.) In the case of RefMAB#1, no shift or decrease in the inhibition curve could be observed after 300 min (Figures 6B and D).

[0464] [Table 36]

[0465] In summary, the combination of both effective neutralization of pathophysiological C5a levels and increased efficacy over time is expected to be beneficial in vivo.

[0466] Example 16: C5a-induced migration of neutrophils C5a-induced chemotaxis of purified neutrophils was analyzed. The experiment was performed essentially as described in US Patent Application Publication No. 2013 / 0295116.

[0467] method Neutrophils were isolated and purified from whole blood of three different human donors using the MACSxpress Whole Blood Neutrophil Isolation Kit, human (Miltenyi Biotec Cat# 130-104-434 and the MACSxpress Erythrocyte Depletion Kit, human (Miltenyi Biotec, CAT# 130-098-196).

[0468] The potency of IgG to inhibit hC5a-dependent neutrophil migration was analyzed by Boyden chamber technique using FluoroBlok® 3.0 μM pore size 96-well plates. The membrane of the Boyden chamber pore was coated with 1 mg / mL human fibrinogen for 2 h at 37° C. After washing, the membrane was blocked with a solution containing 2% bovine serum albumin (BSA) for 1 h at 37° C. Purified neutrophils were then stained with calcein and incubated with 10 mL of 100 mM sodium citrate or 10 mL of 100 mM sodium citrate with or without antagonistic IgG (100 and 600 nM). 5 Stained cells were added to the upper compartment of the Boyden chamber. hC5a (R&D Systems, 10 nM) was applied to the lower compartment of the Boyden chamber. The plate was read at 485 / 538 nm in a plate reader (Tecan M1000Pro) every 5 min at 37° C. for 60 min. Calcein-stained neutrophils passing through the fluoroblock membrane were measured to determine the ability of neutrophils to migrate into the lower chamber. The results are expressed as kinetic migration curves (5-60 min) calculated at selected time points (15, 25 and 35 min) as well as percentage inhibition.

[0469] The formula used to calculate % inhibition was: % Inhibition = 100-(((RFU 試料 -RFU Min )) / ((RFU Max -RFU Min ))×100)

[0470] result The mean percentage inhibition of neutrophil migration at selected time points at the two test IgG concentrations was calculated from three independent experiments using neutrophils from three different human donors.

[0471] As shown in Figure 7, after 15 min of transmigration, almost complete inhibition of C5a-induced neutrophil migration was achieved for MAB#1. After 35 min of transmigration and at the highest IgG concentration tested (600 nM), MAB#1 still demonstrated >50% inhibition. The negative isotype control antibody MOR03207 showed no inhibitory effect on neutrophil migration.

[0472] Example 17: C5a-induced release of cytokines by macrophages Macrophages release pro- or anti-inflammatory cytokines upon activation, depending on their polarization. Therefore, we assessed whether blocking C5a / C5aR interactions with MAB#1 affected C5a-induced release of cytokines by M1 and M2 macrophages. Cytokine ELISA was performed to detect the production of anti-inflammatory IL-10 by M2 macrophages and pro-inflammatory IL-12 by M1 macrophages.

[0473] method Briefly, peripheral blood mononuclear cells (PBMCs) were prepared from whole blood of healthy human donors by density gradient centrifugation using Biocoll separation solution and SepMate tubes (Stemcell). Monocytes were isolated from PBMCs using a CD14+ selection kit (Miltenyi Biotech) and cultured in RPMI supplemented with 10% FCS and 1×GlutaMax in 96-well plates. Overnight plated monocytes were polarized into M1 macrophages for 24 h at 37° C. using LPS (20 ng / ml) and IFNγ (50 ng / ml), pre-incubated with MAB#1 (30 nM) for 30 min, followed by the addition of C5a (15 nM) for an additional 24 h. M1 macrophages stimulated with C5a in the absence of MAB#1 and untreated controls were included. After 24 hours, cell supernatants were analyzed by IL-12 / IL23 DuoSet ELISA (R&D Systems) according to the manufacturer's instructions. For generation of M2 macrophages, monocytes were pre-differentiated into macrophages by culture in RPMI / 10% FCS supplemented with M-CSF for 5 days, and M-CSF, IL-4, IL-13 and IL-6 (40 ng / ml each) were added on day 5. C5a (15 nM) + / - MAB#1 (30 nM) was added daily from day 5 to day 9. On day 9, cell supernatants were analyzed by IL-10 DuoSet ELISA (R&D Systems) according to the manufacturer's instructions.

[0474] result While incubation with C5a led to reduced IL-12 production in M1 macrophages, no reduction in IL-12 levels could be observed after pretreatment with MAB#1 compared to untreated controls, whereas treatment with MAB#1 inhibited C5a-induced IL-10 production in M2 macrophages (see FIG. 11).

[0475] In conclusion, MAB#1 efficiently inhibited C5a-induced production of anti-inflammatory IL-10 by M2 macrophages and restored the production of pro-inflammatory IL-12 by M1 macrophages, thereby revealing the mode of action of MAB#1 in vitro.

[0476] Pharmacokinetics Example 18: Pharmacokinetics The pharmacokinetic profile of MAB#1 was evaluated in male Han-Wistar rats (n=3) after a single intravenous (iv) dose of 10 mg / kg IgG.

[0477] method Plasma samples were collected from each animal via retro-orbital plexus or submandibular vein puncture at the following time points: pre-dose, 0.083, 1, 3, 8, 24, 48, 72, 96, 168, 240, 336 and 504 hours post-dose.

[0478] Free bioactive MAB#1 concentrations in rat plasma were determined using an MSD-based ligand binding assay. Briefly, biotinylated N-terminal human C5aR peptide was coated on the surface of a 96-well streptavidin-MSD plate. Bound analytes were detected using drug-specific anti-idiotypic ECL-labeled antibodies. Based on the free drug concentrations in plasma, noncompartmental data analysis (NCA) was used to evaluate the pharmacokinetic properties of MAB#1.

[0479] result The mean plasma concentrations over time are shown in Figure 9. In all three animals, the mean maximum plasma concentration of MAB#1 after a single iv dose was observed 5 minutes (i.e., 0.083 hours) after dosing (T max ) (i.e., the first sampling time point after dosing). The mean volume of distribution (Vz) of 106 mL / kg was between the plasma volume and the extracellular volume (Davies et al., 1993). The mean terminal elimination half-life after iv dosing was determined to be 9.0 days and the mean total clearance was determined to be 0.341 mL / h / kg.

[0480] Overall, MAB#1 revealed a typical pharmacokinetic profile of a human IgG1 antibody in rat plasma, without cross-reactivity to rodent C5aR: no indication of anti-drug antibody (ADA)-mediated clearance could be detected.

Claims

1. 1. An isolated antibody or antibody fragment specific for human C5aR, comprising: a) the HCDR1 region of SEQ ID NO:27, the HCDR2 region of SEQ ID NO:28, the HCDR3 region of SEQ ID NO:29, the LCDR1 region of SEQ ID NO:32, the LCDR2 region of SEQ ID NO:33 and the LCDR3 region of SEQ ID NO:34, or b) the HCDR1 region of SEQ ID NO:27, the HCDR2 region of SEQ ID NO:39, the HCDR3 region of SEQ ID NO:40, the LCDR1 region of SEQ ID NO:32, the LCDR2 region of SEQ ID NO:33, and the LCDR3 region of SEQ ID NO:

34.

2. An isolated antibody or antibody fragment comprising:

2. a) a VH of SEQ ID NO: 35 and a VL of SEQ ID NO: 36, or b) VH of SEQ ID NO: 42 and VL of SEQ ID NO: 43 3. The isolated antibody or antibody fragment of claim 1 or claim 2, comprising:

3. a) a HC of SEQ ID NO: 37 and a LC of SEQ ID NO: 38, or b) HC of SEQ ID NO: 44 and LC of SEQ ID NO: 45 3. The isolated antibody or antibody fragment of claim 1 or claim 2, comprising:

4. 4. The isolated antibody or antibody fragment of any one of claims 1 to 3, wherein the antibody is of the human IgG1 class.

5. 5. The isolated antibody or antibody fragment of any one of claims 1 to 4, wherein the antibody does not substantially induce effector function in vitro.

6. 6. The isolated antibody or antibody fragment of any one of claims 1 to 5, wherein the antibody comprises one or more amino acid substitutions selected from the group consisting of L234A, L235E, G237A, A330S and P331S as numbered according to the EU index.

7. 7. The isolated antibody or antibody fragment of any one of claims 1 to 6, which is specific for human C5aR and cynomolgus C5aR.

8. In vitro, in the presence of 150 nM human C5a, IC ５０ 8. The isolated antibody or antibody fragment of any one of claims 1 to 7, which inhibits human C5a-induced CD11b expression in human granulocytes at a concentration

9. 9. The isolated antibody or antibody fragment of any of claims 1 to 8, which is a monoclonal antibody or antibody fragment.

10. 10. The isolated antibody or antibody fragment of any of claims 1 to 9, which is a human, humanized or chimeric antibody or antibody fragment.

11. 11. An isolated antibody or antibody fragment according to any one of claims 1 to 10 for use in medicine.

12. A nucleic acid composition comprising a nucleic acid sequence or a plurality of nucleic acid sequences encoding the isolated antibody or antibody fragment of any one of claims 1 to 10.

13. A vector composition comprising a vector or a plurality of vectors comprising the nucleic acid composition of claim 12.

14. A host cell comprising the vector composition of claim 13 or the nucleic acid composition of claim 12.

15. A pharmaceutical composition comprising the isolated antibody or antibody fragment of claims 1-10 and a pharma- ceutically acceptable carrier or excipient.

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