Anti-il-31 antibody constructs and methods of use

Abstract

Antibody constructs that specifically bind to IL-31 and the use of the antibody constructs in the treatment of allergic, inflammatory and / or autoimmune diseases and disorders. The antibody constructs may be monospecific or multispecific, including antibody constructs comprising at least one antigen-binding domain that binds to IL-31 and at least one antigen-binding domain that binds to another cytokine or cytokine receptor, such as IL-4R.

Description

ANTI-IL-31 ANTIBODY CONSTRUCTS AND METHODS OF USE FIELD

[0001] The present disclosure relates to the field of antibody therapeutics and, in particular, to monospecific and bispecific antibodies that bind to IL-31, including bispecific antibodies that bind to IL-31 and IL-4Ra, and methods of using such antibodies.BACKGROUND

[0002] Interleukin 31 (IL-31) is an inflammatory cytokine preferentially produced by Th2 -cells that helps trigger cell-mediated immunity against pathogens. IL-31 is implicated in various chronic inflammatory diseases, such as atopic dermatitis (AD), allergic asthma, allergic rhinitis, inflammatory bowel diseases, malignancies and osteoporosis (see, for example, Bagci etal., 2018, J Allergy Clin Immunol., 141:858-866.

[0003] Antibodies targeting IL-31 or its receptor for treatment of certain inflammatory diseases have been developed. For example, Nemolizumab is a monoclonal antibody that targets IL-31 receptor A. Nemolizumab is clinically proven to be effective at reducing itch in patients suffering from atopic dermatitis (Ruzicka et al., 2017, N Engl J Med., 376:2092-2093). BMS-981164 is an IL-31 -neutralizing antibody that was developed as a treatment for chronic pruritic skin conditions (Lewis etal., 2017, JEurAcad of Dermatol Venereol., 31: 142-150) andNM26-2198 is abispecific antibody that targets IL-31 and IL-4Ra that is in Phase I trials for the treatment of atopic dermatitis (see International Patent Publication Nos. WO 2022 / 136672 and WO 2022 / 136669).

[0004] This background information is provided for the purpose of making known information believed by the applicant to be of possible relevance to the present disclosure. No admission is necessarily intended, nor should be construed, that any of the preceding information constitutes prior art against the claimed invention(s).SUMMARY

[0005] Described herein are anti -IL-31 antibody constructs and methods of use. One aspect of the present disclosure relates to an antibody construct comprising one or more antigen-binding domains, wherein at least one of the antigen-binding domains is an IL-31 antigen-binding domain 1IPTS / 200238736.2that specifically binds to human IL-31, the IL-31 antigen-binding domain comprising the CDR sequences (HCDR1, HCDR2, HCDR3) of the VH domain as set forth in any one of SEQ ID NOs: 47, 48, 49, 50, 51, 52, 53, 54, 55, 57, 58, 59, 60, 61 or 119, and the CDR sequences (LCDR1, LCDR2, LCDR3) of the VL domain as set forth in any one of SEQ ID NOs: 56, 62, 63, 64, 65 or 120.

[0006] Another aspect of the present disclosure relates to a polynucleotide or set of polynucleotides encoding an anti-IL-31 antibody construct as described herein.

[0007] Another aspect of the present disclosure relates to an expression vector or set of expression vectors comprising a polynucleotide or set of polynucleotides encoding an anti-IL-31 antibody construct as described herein.

[0008] Another aspect of the present disclosure relates to a host cell comprising a polynucleotide or set of polynucleotides encoding an anti-IL-31 antibody construct as described herein or an expression vector or set of expression vectors comprising a polynucleotide or set of polynucleotides encoding an anti-IL-31 antibody construct as described herein.

[0009] Another aspect of the present disclosure relates to a method of preparing an anti-IL-31 antibody construct as described herein comprising transfecting a host cell with a polynucleotide or set of polynucleotides encoding the anti-IL-31 antibody construct or an expression vector or set of expression vectors comprising a polynucleotide or set of polynucleotides encoding an anti-IL-31 antibody construct as described herein, and culturing the host cell under conditions suitable for expression of the antibody construct.

[0010] Another aspect of the present disclosure relates to a multispecific antibody construct comprising an IL-31 antigen-binding domain that specifically binds to human IL-31, and one or more additional antigen-binding domains, wherein the one or more additional antigen-binding domains each specifically bind to an antigen other than IL-31, and wherein the IL-31 antigenbinding domain comprises the CDR sequences (HCDR1, HCDR2, HCDR3) of the VH domain as set forth in any one of SEQ ID NOs: 47, 48, 49, 50, 51, 52, 53, 54, 55, 57, 58, 59, 60, 61 or 119, and the CDR sequences (LCDR1, LCDR2, LCDR3) of the VL domain as set forth in any one of SEQ ID NOs: 56, 62, 63, 64, 65 or 120.2IPTS / 200238736.2

[0011] The multispecific antibody construct comprises at least a second target antigen-binding domain that binds to a second target antigen, wherein the second target antigen is a cytokine or cytokine receptor. In some embodiments, the second target antigen is human IL-4Ra.

[0012] Another aspect of the present disclosure relates to a multispecific antibody construct comprising an IL-31 antigen-binding domain that specifically binds to human IL-31, and one or more additional antigen-binding domains, wherein the one or more additional antigen-binding domains each specifically bind to an antigen other than IL-31, wherein the IL-31 antigen-binding domain comprises a VH domain comprising an HCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 15 or 18, an HCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 16 or 19, and an HCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 38 or 39, and a VL domain comprising an LCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 28 or 40, an LCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 44 or 45, and an LCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 30, and wherein the one or more additional antigen-binding domains comprise a second target antigen-binding domain that specifically binds to IL-4Ra, wherein the second target antigen-binding domain comprises an HCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 68 or 71, an HCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 94 or 218, and an HCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 70 or 73, and a VL domain comprising an LCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 86 or 89, an LCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 87 or 90, and an LCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 88.

[0013] Another aspect of the present disclosure relates to a polynucleotide or set of polynucleotides encoding a multispecific antibody construct as described herein.

[0014] Another aspect of the present disclosure relates to an expression vector or set of expression vectors comprising a polynucleotide or set of polynucleotides encoding a multispecific antibody construct as described herein.3IPTS / 200238736.2

[0015] Another aspect of the present disclosure relates to a host cell comprising a polynucleotide or set of polynucleotides encoding a multispecific antibody construct as described herein or an expression vector or set of expression vectors comprising a polynucleotide or set of polynucleotides encoding a bispecific antibody construct as described herein.

[0016] Another aspect of the present disclosure relates to a method of preparing a multispecific antibody construct as described herein comprising transfecting a host cell with a polynucleotide or set of polynucleotides encoding the multispecific antibody construct or an expression vector or set of expression vectors a polynucleotide or set of polynucleotides encoding the multispecific antibody construct, and culturing the host cell under conditions suitable for expression of the multispecific antibody construct.

[0017] Another aspect of the present disclosure relates to a pharmaceutical composition comprising an anti-IL-31 antibody construct as described herein or a multispecific antibody construct as described herein.

[0018] Another aspect of the present disclosure relates to an anti -IL-31 antibody construct as described herein or a multispecific antibody construct as described herein for use in therapy.

[0019] Another aspect of the present disclosure relates to a use of an anti-IL-31 antibody construct as described herein or a multispecific antibody construct as described herein in therapy.

[0020] Another aspect of the present disclosure relates to a use of an anti-IL-31 antibody construct as described herein or a multispecific antibody construct as described herein in the manufacture of a medicament.

[0021] Another aspect of the present disclosure relates to a method of alleviating pruritis in a subject comprising administering to the subject an effective amount of an anti-IL-31 antibody construct as described herein or a multispecific antibody construct as described herein.

[0022] Another aspect of the present disclosure relates to a method of treating an allergic, inflammatory or autoimmune disease in a subject comprising administering to the subject an effective amount of an anti-IL-31 antibody construct as described herein or a multispecific antibody construct as described herein.4IPTS / 200238736.2

[0023] Another aspect of the present disclosure relates to an anti -IL-31 antibody construct as described herein or a multispecific antibody construct as described herein for use to alleviate pruritis in a subject.

[0024] Another aspect of the present disclosure relates to an anti -IL-31 antibody construct as described herein or a multispecific antibody construct as described herein for use in the treatment of an allergic, inflammatory or autoimmune disease in a subject.

[0025] Another aspect of the present disclosure relates to a use of an anti-IL-31 antibody construct as described herein or a multispecific antibody construct as described herein to alleviate pruritis in a subject.

[0026] Another aspect of the present disclosure relates to a use of an anti-IL-31 antibody construct as described herein or a multispecific antibody construct as described herein in the treatment of an allergic, inflammatory or autoimmune disease in a subject.

[0027] Another aspect of the present disclosure relates to a use of an anti-IL-31 antibody construct as described herein or a multispecific antibody construct as described herein in the manufacture of a medicament to alleviate pruritis.

[0028] Another aspect of the present disclosure relates to a use of an anti-IL-31 antibody construct as described herein or a multispecific antibody construct as described herein in the manufacture of a medicament for the treatment of an allergic, inflammatory or autoimmune disease.BRIEF DESCRIPTION OF THE FIGURES

[0029] Figs. 1A-C show binding of the anti-IL-31 x anti-IL-4Ra bispecific IgG4 antibody variants, v41790 and v41791, on human peripheral blood mononuclear cells (PBMCs) by flow cytometry compared to dupilumab (anti-IL-4Ra), NM26-2198 (anti-IL-13 x anti-IL-4Ra) and palivizumab (v36992; negative control). Fig. 1A CD3+CD4+T cells; Fig. 1B CD3⁻CD19⁺ B cells and Fig. 1C CD3⁻CD14⁺CD15⁺CD16⁻ classical monocytes.

[0030] Figs. 2A-2E show inhibition of IL-4 / IL-13 mediated production of the STAT6 inducible secreted embryonic alkaline phosphatase (SEAP) reporter in HEK-Blue™ IL-4 / IL- 13 cells by anti-IL-31 x anti-IL-4Ra bispecific antibody variants. Figs. 2A and 2C show inhibition of IL-45IPTS / 200238736.2stimulated SEAP production by bispecific antibody variants with an IgGl FcKO backbone; Fig.2B shows inhibition of IL- 13 stimulated SEAP production by bispecific antibody variants with an IgGl FcKO backbone; Figs.2D and 2E show inhibition of IL-4 stimulated SEAP production and IL- 13 stimulated SEAP production, respectively, by three bispecific antibody variants with an IgG4 backbone (v41789, v41790 and v41791) and one bispecific antibody variant with an IgGl backbone (v41544). v22277, v39982, v36992 and v42104 are negative controls.

[0031] Figs. 3A-3B show inhibition of IL-31 mediated proliferation in Ba / F3 cells stably expressing human IL-3 IRA and oncostatin M receptor (OSMR) by anti-IL-31 monospecific and anti-IL-31 x anti-IL-4Ra bispecific antibody variants. Fig. 3A shows inhibition by two monospecific variants (v36542 and v36974) and four bispecific antibody variants with an IgGl FcKO backbone (v38728, v39441, v38681 and v39439); Fig. 3B shows inhibition by three bispecific antibody variants with an IgG4 backbone (v41789, v41790 and v41791) and one bispecific antibody variant with an IgGl backbone (v41544). v39982, v42104 and dupilumab) are negative controls.

[0032] Fig. 4 shows inhibition of CD23 upregulation in monocytes by anti-IL-31 x anti-IL-4Ra bispecific IgGl and IgG4 antibody variants following IL-4 stimulation. V42104 is a negative control.

[0033] Fig.5 shows inhibition of CCL2 gene expression by an anti-IL-31 x anti-IL-4Ra bispecific IgG4 antibody variant, v41791, following combined cytokine stimulation of the IL-4, IL- 13 and IL-31 pathways in HEKa cells.

[0034] Figs.6A-6E show inhibition of IL-4 mediated production of CCL17 (TARC) in peripheral blood mononuclear cells (PBMCs) by anti-IL-31 x anti-IL-4Ra bispecific antibody variants. Figs.6A and 6B show inhibition by bispecific antibody variants with an IgGl FcKO backbone in PBMCs from two different donors; Figs. 6C, 6D and 6E show inhibition by three bispecific antibody variants with an IgG4 backbone (v41789, v41790 and v41791) and one bispecific antibody variant with an IgGl backbone (v41544) in PBMCs from three different donors. v36912 is a monospecific anti-IL4Ra antibody, and v36992, v39982 and v42104 are negative controls.6IPTS / 200238736.2

[0035] Fig. 7 shows total antibody concentrations in serum after 3 mg / kg single intravenous dose of the anti-IL-31 x anti-IL-4Ra bispecific IgG4 antibody variants, v41790 and v41791, in Wistar Han rats.

[0036] Fig.8 shows total antibody concentrations in serum after 3 mg / kg single subcutaneous dose of the anti-IL-31 x anti-IL-4Ra bispecific IgG4 antibody variants, v41790 and v41791, in Wistar Han rats.

[0037] Fig. 9 shows total antibody concentrations in serum after 5 mg / kg single intravenous dose of the anti-IL-31 x anti-IL-4Ra bispecific IgGl antibody variants, v41543 and v41544, in Tg32 mice.

[0038] Figs. 10A-10D show total antibody concentrations in serum after 5 mg / kg single intravenous dose of anti-IL-31 x anti-IL-4Ra bispecific antibody variants in Tg32-SCID mice. Fig.10A v41544 (IgGl backbone); Fig. 10B v41789 (IgG4 backbone); Fig. 10C v41790 (IgG4 backbone) and Fig. 10D v41791 (IgG4 backbone).

[0039] Figs. 11A-11D show the results of treating an acute house dust mite (HDM) mouse model with 25, 10, 3 or 1 mg / kg of the anti-IL-31 x anti -IL-4Ra bispecific IgG4 antibody variant, v41791.Fig. 11A serum IgE levels; Fig. 11B lung hIL-4 levels; Fig. 11C lung eosinophil levels, and Fig. 11D lung alveolar macrophage levels. Data are represented as mean + / - SEM. LLOQ = lower limit of quantification.

[0040] Fig. 12 shows the serum pharmacokinetic (PK) profile from cynomolgus monkeys injected intravenously with 10 mg / kg of the anti -IL-31 x anti-IL-4Ra bispecific IgG4 antibody variant, v41791. LLOQ = lower limit of quantification.

[0041] Figs. 13A-13D show inhibition of IL-4 / IL-13 mediated production ofthe STAT6 inducible secreted embryonic alkaline phosphatase (SEAP) reporter in HEK-Blue™ IL-4 / IL- 13 cells by anti-IL-31 x anti-IL-4Ra bispecific IgG4 antibody variants comprising engineered anti-IL-4Ra paratopes. Figs. 13A and 13B v43190 and v43193 compared to v41790 comprising a parental anti-IL-4Ra paratope; Figs. 13C and 13D v43181, v43182 and v43188 compared to v41791 comprising a parental anti-IL-4Ra paratope.7IPTS / 200238736.2

[0042] Fig. 14 presents the CDR sequences for the chimeric parental anti-IL-31 antibody, v33559, and humanized variants based on the parental v33559 sequences (Table Al). Differences compared to parental (v33559) CDR sequences are marked in bold and underlined.

[0043] Fig. 15 presents the CDR sequences for affinity matured, humanized variants based on the chimeric parental anti-IL-31 antibody, v33559, sequences (Table A2). Differences compared to parental (v33559) CDR sequences are marked in bold and underlined.

[0044] Fig. 16 presents the VH and VL sequences for humanized variants based on the chimeric parental anti-IL-31 antibody, v33559 (Table Bl).

[0045] Fig. 17 presents the VH and VL sequences for affinity matured, humanized variants based on the chimeric parental anti -IL-31 antibody, v33559 (Table B2).

[0046] Fig. 18 presents the CDR sequences for humanized anti-IL-4Ra antigen-binding domains comprised by bispecific anti-IL-31 x anti-IL-4Ra antibody variants (Table Cl).

[0047] Fig. 19 presents the CDR sequences for certain engineered anti-IL-4Ra antigen-binding domains comprised by bispecific anti-IL-31 x anti-IL-4Ra antibody variants (v41791, v43181, v43182, v43184, v43188, v43190 and v43193)(Table C2).

[0048] Fig. 20 presents the VH and VL sequences for humanized anti-IL-4Ra antigen-binding domains comprised by bispecific anti -IL-31 x anti-IL-4Ra antibody variants (Table DI), and for certain engineered anti-IL-4Ra antigen-binding domains comprised by bispecific anti -IL-31 x anti-IL-4Ra antibody variants (v41791, v43181, v43182, v43184, v43188, v43190 and v43193)(Table D2).

[0049] Fig. 21 presents a table (Table E) summarizing the monospecific anti-IL-31 antibody variants described in the Examples.

[0050] Fig. 22 presents a table (Table F) summarizing the bispecific anti-IL-31 x anti-IL-4Ra antibody variants described in the Examples.

[0051] Figs.23A-23C show inhibition of IL-4 / IL-13 mediated production ofthe STAT6 inducible secreted embryonic alkaline phosphatase (SEAP) reporter in HEK-Blue™ IL-4 / IL-13 cells by anti-8IPTS / 200238736.2IL-31 x anti-IL-4Ra bispecific antibody variants with an IgG4 backbone (v41791 and v43184).Fig. 23A shows inhibition of IL-4 stimulated SEAP production; Fig. 23B shows inhibition of IL-13 stimulated SEAP production, and Fig. 23C shows inhibition of IL-31 stimulated SEAP production. v42104 is a negative control.

[0052] Fig.24 shows the serum pharmacokinetic (PK) profile from cynomolgus monkeys injected intravenously with 40 mg / kg of the anti-IL-31 x anti-IL-4Ra bispecific IgG4 antibody variant, v44927. (LLOQ: lower limit of quantification).

[0053] Fig.25 shows the serum IgE levels from cynomolgus monkeys injected intravenously with 40 mg / kg of the anti-IL-31 x anti-IL-4Ra bispecific IgG4 antibody variant, v44927.

[0054] Figs. 26A-26C show the results from an oxazolone-induced atopic dermatitis efficacy study of the anti-IL-31 x anti-IL-4Ra bispecific IgG4 antibody variant, v44927, administered at 25 mg / kg. Fig. 26A shows body weight of the treated mice, Fig. 26B shows ear thickness by caliper measurement, and Fig. 26C shows hIL-4 serum concentrations.DETAILED DESCRIPTION

[0055] The present disclosure relates to antibody constructs that specifically bind to IL-31. The antibody constructs may be monospecific antibody constructs comprising one or more antigenbinding domains that bind to the same epitope on IL-31, or they may be biparatopic antibody constructs comprising two antigen-binding domains each of which binds to a different epitope on IL-31, or they may be bispecific or multispecific antibody constructs that comprise at least one antigen-binding domain that binds to IL-31 and at least one antigen-binding domain that binds to a target antigen other than IL-31. In certain embodiments, the bispecific or multispecific antibody constructs comprise at least one antigen-binding domain that binds to IL-31 and at least one antigen-binding construct that binds to another cytokine or cytokine receptor. Certain embodiments relate to bispecific or multispecific antibody constructs comprising at least one antigen-binding domain that binds to IL-31 and at least one antigen-binding domain that binds to IL-4Ra.

[0056] IL-31 has been determined to be involved in a number of allergic, inflammatory and / or autoimmune diseases and disorders, such as atopic dermatitis, allergic rhinitis, allergic asthma and 9IPTS / 200238736.2inflammatory bowel disease. Certain embodiments thus relate to the use of the anti-IL-31 antibody constructs of the present disclosure in the treatment of such IL-31 related allergic, inflammatory and / or autoimmune diseases and disorders.Definitions

[0057] Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art.

[0058] As used herein, the term “about” refers to an approximately + / -10% variation from a given value. It is to be understood that such a variation is always included in any given value provided herein, whether or not it is specifically referred to.

[0059] The use of the word “a” or “an” when used herein in conjunction with the term “comprising” may mean “one,” but it is also consistent with the meaning of “one or more,” “at least one” and “one or more than one.”

[0060] As used herein, the terms “comprising,” “having,” “including” and “containing,” and grammatical variations thereof, are inclusive or open-ended and do not exclude additional, unrecited elements and / or method steps. The term “consisting essentially of’ when used herein in connection with a composition, use or method, denotes that additional elements and / or method steps may be present, but that these additions do not materially affect the manner in which the recited composition, method or use functions. The term “consisting of’ when used herein in connection with a composition, use or method, excludes the presence of additional elements and / or method steps. A composition, use or method described herein as comprising certain elements and / or steps may also, in certain embodiments consist essentially of those elements and / or steps, and in other embodiments consist of those elements and / or steps, whether or not these embodiments are specifically referred to.

[0061] A “complementarity determining region” or “CDR” is an amino acid sequence that contributes to antigen-binding specificity and affinity. “Framework” regions (FR) can aid in maintaining the proper conformation of the CDRs to promote binding between the antigen-binding region and an antigen. From N-terminus to C-terminus, both the light chain variable region (VL) and the heavy chain variable region (VH) of an antibody typically comprise the domains FR1,10IPTS / 200238736.2CDR1, FR2, CDR2, FR3, CDR3, and FR4. The three heavy chain CDRs are referred to herein as HCDR1, HCDR2, and HCDR3, and the three light chain CDRs are referred to as LCDR1, LCDR2, and LCDR3. CDRs provide the majority of contact residues for the binding of the antibody to the antigen or epitope. Often, the three heavy chain CDRs and the three light chain CDRs are required to bind antigen. However, as is known in the art, in some instances, just a single variable domain can confer binding specificity to the antigen, and in some cases, antigen-binding may also occur through a combination of a minimum of one or more CDRs selected from the VH and / or VL domains, for example HCDR3.

[0062] A number of different definitions of the CDR sequences are in common use, including those described by Kabat et al. (1983, Sequences of Proteins of Immunological Interest, NIH Publication No. 369-847, Bethesda, MD), by Chothia et al. (1987, J Mol Biol, 196:901-917), as well as the IMGT, AbM (University of Bath) and Contact (MacCallum, et al., 1996, J Mol Biol, 262(5):732-745) definitions. By way of example, CDR definitions according to Kabat, Chothia, IMGT, AbM and Contact are provided in Table 1 below. Accordingly, as would be readily apparent to one skilled in the art, the exact numbering and placement of CDRs may differ based on the numbering system employed. However, it is to be understood that the disclosure herein of a VH includes the disclosure of the associated (inherent) heavy chain CDRs (HCDRs) as defined by any of the known numbering systems. Similarly, disclosure herein of a VL includes the disclosure of the associated (inherent) light chain CDRs (LCDRs) as defined by any of the known numbering systems.Table 1: Common CDR Definitions1Definition Heavy Chain Light ChainCDR12CDR2 CDR3 CDR1 CDR2 CDR3 Kabat H31-H35 H50-H65 H95-H102 L24-L34 L50-L56 L89-L97 Chothia H26-H32 H52-H56 H95-H102 L24-L34 L50-L56 L89-L97 IMGT H26-H33 H51-H57 H93-H102 L27-L32 L50-L52 L89-L97 AbM H26-H35 H50-H58 H95-H102 L24-L34 L50-L56 L89-L9711IPTS / 200238736.2Definition Heavy Chain Light ChainCDR12CDR2 CDR3 CDR1 CDR2 CDR3 Contact H30-H35 H47-H58 H93-H101 L30-L36 L46-L55 L89-L961Either the Kabat or Chothia numbering system for antibody sequences may be used for HCDR2, HCDR3 and the light chain CDRs for all definitions except Contact, which uses Chothia numbering.2HCDR1 as shown is defined using Chothia numbering. The position in the Kabat numbering system that demarcates the end of the Chothia and IMGT CDR-H1 loop varies depending on the length of the loop because Kabat numbering places insertions outside of those CDR definitions at positions 35A and 35B. However, the IMGT and Chothia CDR-H1 loop can be unambiguously defined using Chothia numbering.

[0063] The term “identical” in the context of two or more polynucleotide or polypeptide sequences, refers to two or more sequences or subsequences that are the same. Sequences are “substantially identical” if they have a percentage of amino acid residues or nucleotides that are the same (for example, about 80%, about 85%, about 90%, about 95%, or about 98% identity, over a specified region) when compared and aligned for maximum correspondence over a comparison window or over a designated region as measured using one of the commonly used sequence comparison algorithms as known to persons of ordinary skill in the art or by manual alignment and visual inspection. For sequence comparison, typically test sequences are compared to a designated reference sequence. When using a sequence comparison algorithm, test and reference sequences are entered into a computer, subsequence coordinates are designated, if necessary, and sequence algorithm program parameters are designated. Default program parameters can be used, or alternative parameters can be designated. The sequence comparison algorithm then calculates the percent sequence identities for the test sequences relative to the reference sequence, based on the program parameters.

[0064] A “comparison window” refers to a segment of a sequence comprising contiguous amino acid or nucleotide positions, the length of which is typically determined based on the length of the test sequence and may be, for example, from about 10 to 600 contiguous amino acid or nucleotide positions, or from about 10 to about 200, or from about 10 to about 150 contiguous amino acid or nucleotide positions over which the test sequence may be compared to a reference sequence of the same number of contiguous positions after the two sequences are optimally aligned. Methods of alignment of sequences for comparison are known to those of ordinary skill in the art. Optimal12IPTS / 200238736.2alignment of sequences for comparison can be conducted, for example, by the local homology algorithm of Smith & Waterman, 1970, Adv. Appl. Math., 2:482c; by the homology alignment algorithm of Needleman & Wunsch, 1970, J. Mol. Biol., 48:443; by the search for similarity method of Pearson & Lipman, 1988, Proc. Natl. Acad. Sci. USA, 85:2444, or by computerized implementations of these algorithms (for example, GAP, BESTFIT, FASTA or TFASTA in the Wisconsin Genetics Software Package, Genetics Computer Group, Madison, WI), or by manual alignment and visual inspection (see, for example, Ausubel et al., Current Protocols in Molecular Biology, (1995 supplement), Cold Spring Harbor Laboratory Press). Examples of available algorithms suitable for determining percent sequence identity are the BLAST and BLAST 2.0 algorithms, which are described in Altschul et al., 1997, Nuc. Acids Res., 25:3389-3402, and Altschul et al., 1990, J. Mol. Biol., 215:403-410, respectively. Software for performing BLAST analyses is publicly available through the website for the National Center for Biotechnology Information (NCBI).

[0065] The term “subject,” as used herein, refers to an animal, in some embodiments a mammal, which is the object of treatment, observation or experiment. The animal may be a human, a nonhuman primate, a companion animal (for example, dog, cat, or the like), farm animal (for example, cow, sheep, pig, horse, or the like) or a laboratory animal (for example, rat, mouse, guinea pig, non-human primate, or the like). In certain embodiments, the subject is a human.

[0066] It is contemplated that any embodiment described herein in relation to the IL-31 antibody constructs can be implemented with respect to any method, use or composition disclosed herein.

[0067] Particular features, structures and / or characteristics described in connection with an embodiment disclosed herein may be combined with features, structures and / or characteristics described in connection with another embodiment disclosed herein in any suitable manner to provide one or more further embodiments.

[0068] It is also to be understood that the positive recitation of a feature in one embodiment, serves as a basis for excluding the feature in an alternative embodiment. For example, where a list of options is presented for a given embodiment or claim, it is to be understood that one or more option may be deleted from the list and the shortened list may form an alternative embodiment, whether or not such an alternative embodiment is specifically referred to.13IPTS / 200238736.2

[0069] As is known in the art, the amino acid residues for the immunoglobulin heavy and light chains may be numbered according to several conventions. Unless otherwise indicated, AbM numbering is used herein for the VH and VL domains, and EU numbering is used herein for the CL, CHI, CH2 and CH3 domains, and the hinge region.ANTI-IL-31 ANTIBODY CONSTRUCTS

[0070] The present disclosure relates to antibody constructs that specifically bind to human IL-31 (anti-IL-31 antibody constructs). In this context, the term “antibody construct” refers to a polypeptide or a set of polypeptides that comprises one or more antigen-binding domains, where each of the one or more antigen-binding domains specifically binds to an epitope or antigen. Where the antibody construct comprises two or more antigen-binding domains, each of the antigenbinding domains may bind the same epitope or antigen (i.e. the antibody construct is monospecific) or they may bind to different epitopes or antigens (i.e. the antibody construct is biparatopic, bispecific or multispecific). The antibody construct may further comprise a scaffold and the one or more antigen-binding domains can be fused or covalently attached to the scaffold, optionally via a linker, as described herein.

[0071] In accordance with the present disclosure, the anti-IL-31 antibody constructs comprise at least one antigen-binding domain that specifically binds to human IL-31 (an “IL-31 antigenbinding domain”). By “specifically binds” to human IL-31, it is meant that the antigen-binding domain binds to human IL-31 and may bind to IL-31 from one or more other non-human species, but does not exhibit significant binding to any other antigen. Specific binding of an antigen-binding domain to a target antigen or epitope may be measured, for example, through an enzyme-linked immunosorbent assay (ELISA), a surface plasmon resonance (SPR) technique (employing, for example, a Biacore™ instrument) (see, for example, Liljeblad et al., 2000, Glyco J, 17:323-329), flow cytometry or a traditional binding assay (see, for example, Heeley, 2002, Endocr Res, 28:217-229).

[0072] In certain embodiments, the anti-IL-31 antibody constructs of the present disclosure may also be capable of binding to IL-31 from one or more non-human species. In certain embodiments, the anti-IL-31 antibody constructs of the present disclosure are capable of binding to cynomolgus monkey IL-31.14IPTS / 200238736.2

[0073] The protein sequence for human IL-31 protein is known in the art and readily available from publicly accessible databases, such as GenBank or UniProtKB. For example, the NCBI reference sequence for human IL-31 is available under GenBank Accession No. NP_001014358 (also provided in Table 2 as SEQ ID NO: 1). The IL-31 sequence for the macaque (cynomolgus) monkey, Macacci fascicularis, is available under GenBank Accession No. EHH66805 (also provided in Table 2 as SEQ ID NO: 2).Table 2: Human and Macaque IL-31 Protein SequencesOrganism Sequence SEQ ID NOHomo MASHSGPSTSVLFLFCCLGGWLASHTLPVRLLRPSDDVQKIVE 1 sapiens ELQSLSKMLLKDVEEEKGVLVSQNYTLPCLSPDAQPPNNIHSP AIRAYLKTIRQLDNKSVIDEIIEHLDKLIFQDAPETNISVPTDTH ECKRFILTISQQFSECMDLALKSLTSGAQQATTMacaca MASHSGPATSVLFLLCCLGGWLTSHTLPVHFLQPSDIQKIVEE 2 fascicularis LQSLSKMLLKDVKEDKGVLVSQNYTLPCLTPDAQPPNIIHSPA IRAYLKTIRQLDNKSVIDEIIEHLDKLIFQDAPETNISVPTDTHE CKRFILTISQQFSECMDLALKSLTSGAQQATTIL-31 Antigen-Binding Domains

[0074] The anti-IL-31 antibody constructs of the present disclosure comprise at least one antigenbinding domain that specifically binds to human IL-31 (an “IL-31 antigen-binding domain”), which is an immunoglobulin-based binding domain, such as an antigen-binding antibody fragment. Examples of an antigen-binding antibody fragment include, but are not limited to, a Fab fragment (Fab), a Fab’ fragment (Fab’), a single chain Fab (scFab), a single chain Fv (scFv) and a single domain antibody (sdAb).15IPTS / 200238736.2

[0075] A “Fab fragment” contains the constant domain of the light chain (CL) and the first constant domain of the heavy chain (CHI) along with the variable domains of the light and heavy chains (VL and VH, respectively). Fab' fragments differ from Fab fragments by the addition of a few amino acid residues at the C-terminus of the heavy chain CHI domain, including one or more cysteines from the antibody hinge region. A Fab fragment may also be a single-chain Fab molecule, i.e. a Fab molecule in which the Fab light chain and the Fab heavy chain are connected by a peptide linker to form a single peptide chain. For example, the C-terminus of the Fab light chain may be connected to the N-terminus of the Fab heavy chain in the single-chain Fab molecule.

[0076] An “scFv” includes a heavy chain variable domain (VH) and a light chain variable domain (VL) of an antibody in a single polypeptide chain. The scFv may optionally further comprise a polypeptide linker between the VH and VL domains which enables the scFv to form a desired structure for antigen binding. For example, an scFv may include a VL connected from its C-terminus to the N-terminus of a VH by a polypeptide linker. Alternately, an scFv may comprise a VH connected through its C-terminus to the N-terminus of a VL by a polypeptide linker (see review by Pluckthun in The Pharmacology of Monoclonal Antibodies, vol. 113, Rosenburg and Moore eds., Springer-Verlag, New York, pp. 269-315 (1994)).

[0077] An “sdAb” format refers to a single immunoglobulin domain. The sdAb may be, for example, of camelid origin. Camelid antibodies lack light chains and their antigen-binding sites consist of a single domain, termed a “VHH.” An sdAb comprises three CDR / hypervariable loops that form the antigen-binding site: CDR1, CDR2 and CDR3. sdAbs are fairly stable and easy to express, for example, as a fusion with the Fc chain of an antibody (see, for example, Harmsen & De Haard, 2007, Appl. Microbiol Biotechnol., 76:13-22).

[0078] In certain embodiments, the IL-31 antigen-binding domain is a Fab or an scFv. In some embodiments, the IL-31 antigen-binding domain is a Fab.

[0079] In those embodiments in which the anti-IL-31 antibody constructs comprise two or more antigen-binding domains, each additional antigen-binding domain may independently be an immunoglobulin-based domain, such as an antigen-binding antibody fragment, or a non-immunoglobulin-based domain, such as a non-immunoglobulin-based antibody mimetic, or other polypeptide or small molecule capable of specifically binding to its target, for example, a natural 16IPTS / 200238736.2or engineered ligand. Non-immunoglobulin-based antibody mimetic formats include, for example, anticalins, fynomers, affimers, alphabodies, DARPins and avimers. The additional antigen-binding domains may bind to IL-31 or they may bind to a different antigen. In certain embodiments, each additional antigen-binding domain is an immunoglobulin-based binding domain, such as an antigen-binding antibody fragment.

[0080] The present disclosure describes the identification of an antibody that specifically binds IL-31 (40A12; variant v33559; see Example 1 and Table E), as well as representative humanized versions of this antibody (variants v34731, v34734, v34735, v34736, v34737, v34738, v34739, v34740 and v34741) and representative affinity-matured versions of this antibody (for example, variants v36535, v36539, v36540, v36541, v36542, v36545 and v36974) (see Examples and Table E). The CDR sequences of the parental antibody v33559 and representative humanized and / or affinity-matured versions of this antibody are shown in Figs. 14 and 15 (Tables Al and A2). The VH and VL sequences of the parental antibody v33559 are shown in Table 1.1 (see Example 1) and the VH and VL sequences of representative humanized and / or affinity-matured versions of this antibody are shown in Figs. 16 and 17 (Tables Bl and B2).

[0081] In certain embodiments, the anti-IL-31 antibody constructs of the present disclosure comprise at least one IL-31 antigen-binding domain, where the IL-31 antigen-binding domain comprises the CDR sequences (HCDR1, HCDR2, HCDR3) of the VH domain as set forth in any one of SEQ ID NOs: 47, 48, 49, 50, 51, 52, 53, 54, 55, 57, 58, 59, 60, 61 or 119. In some embodiments, the anti-IL-31 antibody constructs of the present disclosure comprise at least one IL-31 antigen-binding domain, where the IL-31 antigen-binding domain comprises the CDR sequences (HCDR1, HCDR2, HCDR3) of the VH domain as set forth in SEQ ID NO: 59 or 61.

[0082] In certain embodiments, the anti-IL-31 antibody constructs of the present disclosure comprise at least one IL-31 antigen-binding domain, where the IL-31 antigen-binding domain comprises the CDR sequences (LCDR1, LCDR2, LCDR3) of the VL domain as set forth in any one of SEQ ID NOs: 56, 62, 63, 64, 65 or 120. In some embodiments, the anti-IL-31 antibody constructs of the present disclosure comprise at least one IL-31 antigen-binding domain, where the IL-31 antigen-binding domain comprises the CDR sequences (LCDR1, LCDR2, LCDR3) of the VL domain as set forth in SEQ ID NO: 65.17IPTS / 200238736.2

[0083] In certain embodiments, the anti-IL-31 antibody constructs of the present disclosure comprise at least one IL-31 antigen-binding domain, where the IL-31 antigen-binding domain comprises the CDR sequences (HCDR1, HCDR2, HCDR3) of the VH domain as set forth in any one of SEQ ID NOs: 47, 48, 49, 50, 51, 52, 53, 54, 55, 57, 58, 59, 60, 61 or 119, and the CDR sequences (LCDR1, LCDR2, LCDR3) of the VL domain as set forth in any one of SEQ ID NOs: 56, 62, 63, 64, 65 or 120. In some embodiments, the anti-IL-31 antibody constructs of the present disclosure comprise at least one IL-31 antigen-binding domain, where the IL-31 antigen-binding domain comprises the CDR sequences (HCDR1, HCDR2, HCDR3) of the VH domain as set forth in SEQ ID NO: 59 or 61, and the CDR sequences (LCDR1, LCDR2, LCDR3) of the VL domain as set forth in SEQ ID NO: 65.

[0084] In certain embodiments, the anti-IL-31 antibody constructs of the present disclosure comprise at least one IL-31 antigen-binding domain, where the IL-31 antigen-binding domain comprises a VH domain comprising heavy chain CDR amino acid sequences (HCDR1, HCDR2 and HCDR3) comprising the sequences as set forth in SEQ ID NOs: 3, 4 and 5, and a VL domain comprising light chain CDR amino acid sequences (LCDR1, LCDR2 and LCDR3) comprising the sequences as set forth in SEQ ID NOs: 6, 7 and 8 (see Table 3). SEQ ID NOs: 3, 4 and 5, and SEQ ID NOs: 6, 7 and 8 are consensus sequences based on the HCDR and LCDR sequences, respectively, of v33559 and representative humanized and / or affinity-matured versions of this antibody when defined by the IMGT system.

[0085] In certain embodiments, the anti-IL-31 antibody constructs of the present disclosure comprise at least one IL-31 antigen-binding domain, where the IL-31 antigen-binding domain comprises a VH domain comprising heavy chain CDR amino acid sequences (HCDR1, HCDR2 and HCDR3) comprising the sequences as set forth in SEQ ID NOs: 9, 10 and 11, and a VL domain comprising light chain CDR amino acid sequences (LCDR1, LCDR2 and LCDR3) comprising the sequences as set forth in SEQ ID NOs: 12, 13 and 14 (see Table 4). SEQ ID NOs: 9, 10 and 11, and SEQ ID NOs: 12, 13 and 14 are consensus sequences based on the HCDR and LCDR sequences, respectively, of v33559 and representative humanized and / or affinity-matured versions of this antibody when defined by the Kabat system.18IPTS / 200238736.2Table 3: Consensus CDR Sequences by IM GTCDR Sequence Variable (X) SEQ ID NOHeavy ChainHCDR1 GIDLSSYF 3HCDR2 ISTGGNT 4HCDR3 ARGWLRDYLDX1X1is R or L 5Light ChainLCDR1 QSVYRENR 6LCDR2 RX2S X2is G orA 7LCDR3 AGGX3SSGSDHA X3is D or A 8Table 4: Consensus CDR Sequences by KabatCDR Sequence Variable (X) SEQ ID NOHeavy ChainHCDR1 SYFMS 9HCDR2 TISTGGNTYYAX4VKG X4is SW orAP 10HCDR3 GWLRDYLDX5X5is R or L 11Light ChainLCDR1 X6QSVYRENRLA X6is RSRor QAS 12LCDR2 RX7SKLEX8X7is G or A 13X8is S orKLCDR3 AGGX9SSGSDHA X9is D or A 14

[0086] In certain embodiments, the anti-IL-31 antibody constructs of the present disclosure comprise at least one IL-31 antigen-binding domain, where the IL-31 antigen-binding domain comprises a VH domain comprising heavy chain CDR amino acid sequences (HCDR1, HCDR2 and HCDR3) comprising the sequences as set forth in SEQ ID NOs: 3, 4 and 5, respectively, and a VL domain comprising light chain CDR amino acid sequences (LCDR1, LCDR2 and LCDR3) comprising the sequences as set forth in SEQ ID NOs: 6, 7 and 8, respectively, and where X1is R, X2is G and X3is D. In certain embodiments, the anti -IL-31 antibody constructs of the present disclosure comprise at least one IL-31 antigen-binding domain, where the IL-31 antigen-binding domain comprises a VH domain comprising heavy chain CDR amino acid sequences (HCDR1,19IPTS / 200238736.2HCDR2 and HCDR3) comprising the sequences as set forth in SEQ ID NOs: 9, 10 and 11, respectively, and a VL domain comprising light chain CDR amino acid sequences (LCDR1, LCDR2 and LCDR3) comprising the sequences as set forth in SEQ ID NOs: 12, 13 and 14, respectively, and where X4is SW, X5is R, X6is RSR, X7is G, X8is S, and X9is D.

[0087] In certain embodiments, the anti-IL-31 antibody constructs of the present disclosure comprise at least one IL-31 antigen-binding domain, where the IL-31 antigen-binding domain comprises a VH domain comprising an HCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 15 or 18, an HCDR2 amino acid sequence comprising the sequence as set forth in any one of SEQ ID NOs: 16, 19 or 37, and an HCDR3 amino acid sequence comprising the sequence as set forth in any one of SEQ ID NOs: 17, 20, 38 or 39. In some embodiments, the anti-IL-31 antibody constructs comprise at least one IL-31 antigen-binding domain, where the IL-31 antigen-binding domain comprises a VH domain comprising an HCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 15 or 18, an HCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 16 or 19, and an HCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 38 or 39.

[0088] In certain embodiments, the anti-IL-31 antibody constructs of the present disclosure comprise at least one IL-31 antigen-binding domain, where the IL-31 antigen-binding domain comprises a VL domain comprising an LCDR1 amino acid sequence comprising the sequence as set forth in any one of SEQ ID NOs: 28, 31, 36 or 40, an LCDR2 amino acid sequence comprising the sequence as set forth in any one of SEQ ID NOs: 29, 32, 41, 44 or 45, and an LCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 30 or 42. In some embodiments, the anti-IL-31 antibody constructs comprise at least one IL-31 antigen-binding domain, where the IL-31 antigen-binding domain comprises a VL domain comprising an LCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 28 or 40, an LCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 44 or 45, and an LCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 30.

[0089] In certain embodiments, the anti-IL-31 antibody constructs of the present disclosure comprise at least one IL-31 antigen-binding domain, where the IL-31 antigen-binding domain comprises a VH domain comprising an HCDR1 amino acid sequence comprising the sequence as20IPTS / 200238736.2set forth in SEQ ID NO: 15 or 18, an HCDR2 amino acid sequence comprising the sequence as set forth in any one of SEQ ID NOs: 16, 19 or 37, and an HCDR3 amino acid sequence comprising the sequence as set forth in anyone of SEQ ID NOs: 17, 20, 38 or 39, and a VL domain comprising an LCDR1 amino acid sequence comprising the sequence as set forth in any one of SEQ ID NOs: 28, 31, 36 or 40, an LCDR2 amino acid sequence comprising the sequence as set forth in any one of SEQ ID NOs: 29, 32, 41, 44 or 45, and an LCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 30 or 42.

[0090] In certain embodiments, the anti-IL-31 antibody constructs of the present disclosure comprise at least one IL-31 antigen-binding domain, where the IL-31 antigen-binding domain comprises a VH domain comprising an HCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 15 or 18, an HCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 16 or 19, and an HCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 17 or 20, and a VL domain comprising either(i) an LCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 28 or 31, an LCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 29 or 32, and an LCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 30, or(ii) an LCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 28 or 36, an LCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 29 or 32, and an LCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 30.

[0091] In certain embodiments, the anti-IL-31 antibody constructs of the present disclosure comprise at least one IL-31 antigen-binding domain, where the IL-31 antigen-binding domain comprises a VH domain comprising either(i) an HCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 15 or 18, an HCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 16 or 37, and an HCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 38 or 39, or21IPTS / 200238736.2(ii) an HCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 15 or 18, an HCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 16 or 19, and an HCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 38 or 39, and

[0092] a VL domain comprising an LCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 28 or 40, an LCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 29 or 41, and an LCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 42.

[0093] In certain embodiments, the anti-IL-31 antibody constructs of the present disclosure comprise at least one IL-31 antigen-binding domain, where the IL-31 antigen-binding domain comprises a VH domain comprising an HCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 15 or 18, an HCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 16 or 19, and an HCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 38 or 39, and a VL domain comprising:(i) an LCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 28 or 40, an LCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 29 or 41, and an LCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 30, or(ii) an LCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 28 or 40, an LCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 29 or 32, and an LCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 30, or(iii) an LCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 28 or 40, an LCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 44 or 45, and an LCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 30.22IPTS / 200238736.2

[0094] In certain embodiments, the anti-IL-31 antibody constructs of the present disclosure comprise at least one IL-31 antigen-binding domain, where the IL-31 antigen-binding domain comprises:(a) a VH domain comprising an HCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 15 or 18, an HCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 16 or 19, and an HCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 17 or 20, and a VL domain comprising an LCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 28 or 31, an LCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 29 or 32, and an LCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 30, or(b) a VH domain comprising an HCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 15 or 18, an HCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 16 or 19, and an HCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 17 or 20, and a VL domain comprising an LCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 28 or 36, an LCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 29 or 32, and an LCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 30, or(c) a VH domain comprising an HCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 15 or 18, an HCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 16 or 37, and an HCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 38 or 39, and a VL domain comprising an LCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 28 or 40, an LCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 29 or 41, and an LCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 42, or(d) a VH domain comprising an HCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 15 or 18, an HCDR2 amino acid sequence comprising the sequence 23IPTS / 200238736.2as set forth in SEQ ID NO: 16 or 19, and an HCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 38 or 39, and a VL domain comprising an LCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 28 or 40, an LCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 29 or 41, and an LCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 42, or(e) a VH domain comprising an HCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 15 or 18, an HCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 16 or 19, and an HCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 38 or 39, and a VL domain comprising an LCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 28 or 40, an LCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 29 or 41, and an LCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 30, or(f) a VH domain comprising an HCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 15 or 18, an HCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 16 or 19, and an HCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 38 or 39, and a VL domain comprising an LCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 28 or 40, an LCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 29 or 32, and an LCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 30, or(g) a VH domain comprising an HCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 15 or 18, an HCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 16 or 19, and an HCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 38 or 39, and a VL domain comprising an LCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 28 or 40, an LCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 44 or24IPTS / 200238736.245, and an LCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 30.

[0095] In some embodiments, the anti-IL-31 antibody constructs of the present disclosure comprise at least one IL-31 antigen-binding domain, where the IL-31 antigen-binding domain comprises a VH domain comprising an HCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 15 or 18, an HCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 16 or 19, and an HCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 38 or 39, and a VL domain comprising an LCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 28 or 40, an LCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 44 or 45, and an LCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 30.

[0096] In certain embodiments, the anti-IL-31 antibody constructs of the present disclosure comprise at least one IL-31 antigen-binding domain, where the IL-31 antigen-binding domain comprises a VH domain having a sequence that is at least about 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the VH sequence as set forth in any one of SEQ ID NOs: 47, 48, 49, 50, 51, 52, 53, 54, 55, 57, 58, 59, 60 or 61. In some embodiments, the anti-IL-31 antibody constructs comprise at least one IL-31 antigen-binding domain, where the IL-31 antigen-binding domain comprises a VH domain having a sequence that is at least about 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the VH sequence as set forth in any one of SEQ ID NOs: 47, 48, 49, 50, 51, 52, 53, 54, 55, 57, 58, 59, 60 or 61. In some embodiments, the anti-IL-31 antibody constructs comprise at least one IL-31 antigen-binding domain, where the IL-31 antigen-binding domain comprises a VH domain having a sequence as set forth in any one of SEQ ID NOs: 47, 48, 49, 50, 51, 52, 53, 54, 55, 57, 58, 59, 60 or 61. In some embodiments, the anti-IL-31 antibody constructs comprise at least one IL-31 antigen-binding domain, where the IL-31 antigen-binding domain comprises a VH domain having a sequence that is at least about 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the VH sequence as set forth in SEQ ID NO: 59 or 61. In some embodiments, the anti-IL-31 antibody constructs comprise at least one IL-31 antigen-binding domain, where the IL-31 antigen-binding domain comprises a VH domain having the sequence as set forth in SEQ ID NO: 59 or 61. In some embodiments, the anti-IL-31 antibody constructs comprise at least one IL-31 antigen-binding domain, where the IL-31 antigen-binding domain25IPTS / 200238736.2comprises a VH domain having a sequence that is at least about 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the VH sequence as set forth in SEQ ID NO: 59. In some embodiments, the anti-IL-31 antibody constructs comprise at least one IL-31 antigen-binding domain, where the IL-31 antigen-binding domain comprises a VH domain having the sequence as set forth in SEQ ID NO: 59.

[0097] In certain embodiments, the anti-IL-31 antibody constructs of the present disclosure comprise at least one IL-31 antigen-binding domain, where the IL-31 antigen-binding domain comprises a VL domain having a sequence that is at least about 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the VL sequence as set forth in any one of SEQ ID NOs: 56, 62, 63, 64 or 65. In some embodiments, the anti -IL-31 antibody constructs comprise at least one IL-31 antigen-binding domain, where the IL-31 antigen-binding domain comprises a VL domain having a sequence that is at least about 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the VL sequence as set forth in any one of SEQ ID NOs: 56, 62, 63, 64 or 65. In some embodiments, the anti -IL-31 antibody constructs comprise at least one IL-31 antigen-binding domain, where the IL-31 antigen-binding domain comprises a VL domain having the sequence as set forth in any one of SEQ ID NOs: 56, 62, 63, 64 or 65. In some embodiments, the anti-IL-31 antibody constructs comprise at least one IL-31 antigen-binding domain, where the IL-31 antigen-binding domain comprises a VL domain having a sequence that is at least about 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the VL sequence as set forth in SEQ ID NO: 65. In some embodiments, the anti-IL-31 antibody constructs comprise at least one IL-31 antigen-binding domain, where the IL-31 antigen-binding domain comprises a VL domain having the sequence as set forth in SEQ ID NO: 65.

[0098] In certain embodiments, the anti-IL-31 antibody constructs of the present disclosure comprise at least one IL-31 antigen-binding domain, where the IL-31 antigen-binding domain comprises a VH domain having a sequence that is at least about 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the VH sequence as set forth in any one of SEQ ID NOs: 47, 48, 49, 50, 51, 52, 53, 54, 55, 57, 58, 59, 60 or 61, and a VL domain having a sequence that is at least about 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the VL sequence as set forth in any one of SEQ ID NOs: 56, 62, 63, 64 or 65. In some embodiments, the anti-IL-31 antibody constructs comprise at least one IL-31 antigen-binding domain, where the IL-31 antigen-binding domain comprises a26IPTS / 200238736.2VH domain having the sequence as set forth in any one of SEQ ID NOs: 47, 48, 49, 50, 51, 52, 53, 54, 55, 57, 58, 59, 60 or 61, and a VL domain having the sequence as set forth in any one of SEQ ID NOs: 56, 62, 63, 64 or 65.

[0099] In certain embodiments, the anti-IL-31 antibody constructs of the present disclosure comprise at least one IL-31 antigen-binding domain, where the IL-31 antigen-binding domain comprises a VH domain having a sequence that is at least about 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the VH sequence as set forth in any one of SEQ ID NOs: 47, 48, 49, 50, 51, 52, 53, 54 or 55, and a VL domain having a sequence that is at least about 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the VL sequence as set forth in SEQ ID NO: 56. In some embodiments, the anti-IL-31 antibody constructs comprise at least one IL-31 antigen-binding domain, where the IL-31 antigen-binding domain comprises a VH domain having the sequence as set forth in any one of SEQ ID NOs: 47, 48, 49, 50, 51, 52, 53, 54 or 55, and a VL domain having the sequence as set forth in SEQ ID NO: 56.

[0100] In certain embodiments, the anti -IL-31 antibody constructs of the present disclosure comprise at least one IL-31 antigen-binding domain, where the IL-31 antigen-binding domain comprises a VH domain having a sequence that is at least about 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the VH sequence as set forth in any one of SEQ ID NOs: 57, 58, 59, 60 or 61, and a VL domain having a sequence that is at least about 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the VL sequence as set forth in any one of SEQ ID NOs: 63, 63, 64 or 65. In some embodiments, the anti -IL-31 antibody constructs comprise at least one IL-31 antigen-binding domain, where the IL-31 antigen-binding domain comprises a VH domain having the sequence as set forth in any one of SEQ ID NOs: 57, 58, 59, 60 or 61, and a VL domain having the sequence as set forth in any one of SEQ ID NOs: 63, 63, 64 or 65.

[0101] In certain embodiments, the anti -IL-31 antibody constructs of the present disclosure comprise at least one IL-31 antigen-binding domain, where the IL-31 antigen-binding domain comprises either(i) a VH domain having the sequence as set forth in SEQ ID NO: 57 or 58, and a VL domain having the sequence as set forth in SEQ ID NO: 62 or 63, or27IPTS / 200238736.2(ii) a VH domain having the sequence as set forth in any one of SEQ ID NOs: 59, 60 or 61, and a VL domain having the sequence as set forth in SEQ ID NO: 64 or 65.

[0102] In certain embodiments, the anti -IL-31 antibody constructs of the present disclosure comprise at least one IL-31 antigen-binding domain, where the IL-31 antigen-binding domain comprises(i) a VH domain having the sequence as set forth in SEQ ID NO: 57 or 58, and a VL domain having the sequence as set forth in SEQ ID NO: 62, or(ii) a VH domain having the sequence as set forth in SEQ ID NO: 58, and a VL domain having the sequence as set forth in any one of SEQ ID NOs: 62 or 63, or(iii) a VH domain having the sequence as set forth in SEQ ID NO: 59, and a VL domain having the sequence as set forth in SEQ ID NO: 64 or 65, or(iv) a VH domain having the sequence as set forth in SEQ ID NO: 59 or 60, and a VL domain having the sequence as set forth in SEQ ID NO: 64, or(v) a VH domain having the sequence as set forth in SEQ ID NO: 59 or 61, and a VL domain having the sequence as set forth in SEQ ID NO: 65.

[0103] In some embodiments, the anti-IL-31 antibody constructs of the present disclosure comprise at least one IL-31 antigen-binding domain, where the IL-31 antigen-binding domain comprises a VH domain having the sequence as set forth in SEQ ID NO: 59 or 61, and a VL domain having the sequence as set forth in SEQ ID NO: 65. In some embodiments, the anti-IL-31 antibody constructs comprise at least one IL-31 antigen-binding domain, where the IL-31 antigen-binding domain comprises a VH domain as set forth in SEQ ID NO: 59, and a VL domain having the sequence as set forth in SEQ ID NO: 65.Formats

[0104] The anti-IL-31 antibody constructs of the present disclosure may have various formats. The minimal component of the anti -IL-31 antibody construct is an antigen-binding domain that binds to human IL-31. The anti-IL-31 antibody constructs may further optionally28IPTS / 200238736.2comprise one or more additional antigen-binding domains and / or a scaffold. In those embodiments in which the anti -IL-31 antibody construct comprises two or more antigen-binding domains, each additional antigen-binding domain may bind to the same epitope within IL-31, may bind to a different epitope within IL-31, or may bind to a different antigen. Thus, the anti-IL-31 antibody construct may be, for example, monospecific, biparatopic, bispecific or multispecific.

[0105] In certain embodiments, the anti-IL-31 antibody construct comprises at least one IL-31 antigen-binding domain and a scaffold, where the IL-31 antigen-binding domain is operably linked to the scaffold. The term “operably linked,” as used herein, means that the components described are in a relationship permitting them to function in their intended manner. Examples of suitable scaffolds are described below.

[0106] In certain embodiments, the anti-IL-31 antibody construct comprises two antigenbinding domains optionally operably linked to a scaffold, where at least one of the antigen-binding domains is an IL-31 antigen-binding domain. In some embodiments, the anti -IL-31 antibody construct may comprise three or four antigen-binding domains and optionally a scaffold, where at least one of the antigen-binding domains is an IL-31 antigen-binding domain. In these formats, when comprising a scaffold, at least a first antigen-binding domain is operably linked to the scaffold and the remaining antigen-binding domain(s) may each independently be operably linked to the scaffold or to the first antigen-binding domain or, when more than two antigen-binding domains are present, to another antigen-binding domain.

[0107] Anti-IL-31 antibody constructs that lack a scaffold may comprise a single IL-31 antigen-binding domain in an appropriate format, such as an sdAb, or they may comprise two or more antigen-binding domains optionally operably linked by one or more linkers, where at least one of the antigen-binding domains is an IL-31 antigen-binding domain. In such anti -IL-31 antibody constructs, the antigen-binding domains may be in the form of scFvs, Fabs, sdAbs, or a combination thereof. For example, using scFvs as the antigen-binding domains, formats such as a tandem scFv ((scFv)2 or taFv) may be constructed, in which the scFvs are connected together by a flexible linker. scFvs may also be used to construct diabody formats, which comprise two scFvs connected by a short linker (usually about 5 amino acids in length). The restricted length of the linker results in dimerization of the scFvs in a head-to-tail manner. In any of the preceding formats,29IPTS / 200238736.2the scFvs may be further stabilized by inclusion of an interdomain disulfide bond. For example, a disulfide bond may be introduced between VL and VH through introduction of an additional cysteine residue in each chain (for example, at position 44 in VH and position 100 in VL) (see, for example, Fitzgerald et al., 1997, Protein Engineering, 10: 1221-1225), or a disulfide bond may be introduced between two VHs to provide a construct having a DART format (see, for example, Johnson etal., 2010, JMol. Biol., 399:436-449).

[0108] Similarly, formats comprising two sdAbs, such as VHs or VHHs, connected together through a suitable linker may be employed in some embodiments. Other examples of anti-IL-31 antibody construct formats that lack a scaffold include those based on Fab fragments, for example, Fab2 and F(ab’)2 formats, in which the Fab fragments are connected through a linker or an IgG hinge region.

[0109] Combinations of antigen-binding domains in different forms may also be employed to generate alternative scaffold-less formats. For example, an scFv or a sdAb may be fused to the C-terminus of either or both of the light and heavy chain of a Fab fragment resulting in a bivalent (Fab-scFv / sdAb) construct.

[0110] In certain embodiments, the anti-IL-31 antibody construct may be in an antibody format that is based on an immunoglobulin (Ig). In certain embodiments, the anti -IL-31 antibody construct may be based on an IgG class immunoglobulin, for example, an IgGl, IgG2, IgG3 or IgG4 immunoglobulin. In some embodiments, the anti-IL-31 antibody construct may be based on an IgGl or IgG4 immunoglobulin. In the context of the present disclosure, when an anti-IL-31 antibody construct is based on a specified immunoglobulin isotype, it is meant that the anti-IL-31 antibody construct comprises all or a portion of the constant region of the specified immunoglobulin isotype. For example, an anti-IL-31 antibody construct based on a given Ig isotype may comprise at least one IL-31 antigen-binding domain operably linked to an Ig scaffold, where the scaffold comprises an Fc region from the given isotype and optionally an Ig hinge region from the same or a different isotype. It is to be understood that the anti-IL-31 antibody constructs may also comprise hybrids of isotypes and / or subclasses in some embodiments. It is also to be understood that the Fc region and / or hinge region may optionally be modified to impart one or more desirable functional properties as is known in the art.30IPTS / 200238736.2

[0111] In some embodiments, the anti-IL-31 antibody constructs may be derived from two or more immunoglobulins that are from different species, for example, the anti-IL-31 antibody construct may be a chimeric antibody or a humanized antibody. The terms “chimeric antibody” and “humanized antibody” both refer generally to antibodies that combine immunoglobulin regions or domains from more than one species.

[0112] A “chimeric antibody” typically comprises at least one variable domain from a nonhuman antibody, such as a rabbit or rodent (for example, murine) antibody, and at least one constant domain from a human antibody. The human constant domain of a chimeric antibody need not be of the same isotype as the non-human constant domain it replaces. Chimeric antibodies are discussed, for example, in Morrison etal., 1984, Proc. Natl. Acad. Sci. USA, 81:6851-55, and U. S. Patent No. 4,816,567.

[0113] A “humanized antibody” is a type of chimeric antibody that contains minimal sequence derived from a non-human antibody. Generally, humanized antibodies are human immunoglobulins (recipient antibody) in which residues from a hypervariable region (CDR) of the recipient are replaced by residues from a hypervariable region (CDR) of a non-human species (donor antibody), such as mouse, rat, rabbit or non-human primate, having the desired specificity and affinity for a target antigen. This technique for creating humanized antibodies is often referred to as “CDR grafting.”

[0114] In some instances, additional modifications may be made to a humanized antibody to further refine antibody performance. For example, framework region (FR) residues of the human immunoglobulin are replaced by corresponding non-human residues, or the humanized antibodies may comprise residues that are not found in either the recipient antibody or the donor antibody. In general, a variable domain in a humanized antibody will comprise all or substantially all of the CDRs from a non-human immunoglobulin and all or substantially all of the FRs from a human immunoglobulin sequence. Humanized antibodies are described in more detail in Jones, et al., 1986, Nature, 321:522-525; Riechmann, etal., 1988, Nature, 332:323-329, and Presta, 1992, Curr. Op. Struct. Biol., 2:593-596, for example.

[0115] A number of approaches are known in the art for selecting the most appropriate human frameworks into which to graft the non-human CDRs. Early approaches used a limited 31IPTS / 200238736.2subset of well-characterised human antibodies, irrespective of the sequence identity to the nonhuman antibody providing the CDRs (the “fixed frameworks” approach). More recent approaches have employed variable regions with high amino acid sequence identity to the variable regions of the non-human antibody providing the CDRs (“homology matching” or “best-fit” approach). An alternative approach is to select fragments of the framework sequences within each light or heavy chain variable region from several different human antibodies. CDR grafting may in some cases result in a partial or complete loss of affinity of the grafted molecule for its target antigen. In such cases, affinity can be restored by back-mutating some of the residues of human origin to the corresponding non-human ones. Methods for preparing humanized antibodies by these approaches are well-known in the art (see, for example, Tsurushita & Vasquez, 2004, Humanization of Monoclonal Antibodies, Molecular Biology of B Cells, 533-545, Elsevier Science (USA); Jones et al., 1986, Nature, 321:522-525; Riechmann et al., 1988, Nature, 332:323-329; Presta et al., 1997, Cancer Res, 57(20):4593-4599).

[0116] Alternatively, or in addition to, these traditional approaches, more recent technologies may be employed to further reduce the immunogenicity of a CDR-grafted humanized antibody. For example, frameworks based on human germline sequences or consensus sequences may be employed as acceptor human frameworks rather than human frameworks with somatic mutation(s). Another technique that aims to reduce the potential immunogenicity of non-human CDRs is to graft only specificity-determining residues (SDRs). In this approach, only the minimum CDR residues required for antigen-binding activity (the “SDRs”) are grafted into a human germline framework. This method improves the “humanness” (i.e. the similarity to human germline sequence) of the humanized antibody and thus may help reduce the risk of immunogenicity of the variable region. These techniques have been described in various publications (see, for example, Almagro & Fransson, 2008, Front Biosci, 13:1619-1633; Tan, et al., 2002, J Immunol, 169:1119-1125; Hwang, etal., 2005, Methods, 36:35-42; Pelat, etal., 2008, J Mol Biol, 384:1400-1407; Tamura, et al., 2000, J Immunol, 164:1432-1441; Gonzales, et al., 2004, Mol Immunol, 1:863-872, and Kashmiri, et al., 2005, Methods, 36:25-34).

[0117] In certain embodiments, the anti -IL-31 antibody construct of the present disclosure comprises humanized antibody sequences, for example, one or more humanized variable domains. In some embodiments, the anti -IL-31 antibody construct is a humanized antibody. Non-limiting32IPTS / 200238736.2examples of humanized antibodies based on the anti-IL-31 antibody 40A12 (variant v33559; see Example 1 and Table E) are described herein (variants v34731, v34734, v34735, v34736, v34737, v34738, v34739, v34740 and v34741; see Examples and Table E).

[0118] Humanized antibodies may also be “affinity matured” in order to improve binding to the target antigen. In vitro affinity maturation usually involves a diversification of the antibody base sequence, followed by stringent selections to isolate higher-affinity binders. Various affinity maturation techniques are known in the art (see, for example, Li, et al., 2023, Int J Biological Macromolecules, 247:125733, Kielczewska, et al., 2022, JBC, 298(2): 101533). In certain embodiments, the anti-IL-31 construct of the present disclosure comprises humanized, affinity matured antibody sequences, for example, one or more humanized, affinity matured variable domains. In some embodiments, the anti -IL-31 antibody construct is a humanized, affinity matured antibody. Non-limiting examples of humanized, affinity matured antibodies based on the anti -IL-31 antibody 40A12 (variant v33559; see Example 1 and Table E) are described herein (variants v36535, v36539, v36540, v36541, v36542, v36545 and v36974; see Examples and Table E). Scaffolds

[0119] In certain embodiments, the anti -IL-31 antibody constructs of the present disclosure comprise one or more IL-31 antigen-binding domains operably linked to a scaffold. The antigenbinding domain(s) may be in one or a combination of the forms described above (for example, scFvs, Fabs and / or sdAbs). Examples of suitable scaffolds are described in more detail below and include, but are not limited to, immunoglobulin Fc regions, albumin, albumin analogues and derivatives, heterodimerizing peptides (such as leucine zippers, heterodimer-forming “zipper” peptides derived from Jun and Fos, IgG CHI and CL domains or bamase-barstar toxins), cytokines, chemokines or growth factors. Other examples include antibodies based on the DOCK-AND-LOCK™ (DNL™) technology developed by IBC Pharmaceuticals, Inc. and Immunomedics, Inc. (see, for example, Chang, et al., 2007, Clin. Cancer Res., 13:5586s-5591s).

[0120] A scaffold may be a peptide, polypeptide, polymer, nanoparticle or other chemical entity. Where the scaffold is a polypeptide, each antigen-binding domain of the anti -IL-31 antibody construct may be linked to either the N- or C-terminus of the polypeptide scaffold. Anti-IL-31 antibody constructs comprising a polypeptide scaffold in which one or more of the antigen-binding 33IPTS / 200238736.2domains are linked to a region other than the N- or C-terminus, for example, via the side chain of an amino acid with or without a linker, are also contemplated in certain embodiments.

[0121] In embodiments where the anti-IL-31 antibody construct comprises a scaffold that is a peptide or polypeptide, the antigen-binding domain(s) may be linked to the scaffold by genetic fusion or chemical conjugation. Typically, when the scaffold is a peptide or polypeptide, the antigen-binding domain(s) are linked to the scaffold by genetic fusion. In some embodiments, where the scaffold is a polymer or nanoparticle, the antigen-binding domain(s) may be linked to the scaffold by chemical conjugation.

[0122] A number of protein domains are known in the art that comprise selective pairs of two different polypeptides and may be used to form a scaffold. An example is leucine zipper domains such as Fos and Jun that selectively pair together (Kostelny, et al., J Immunol, 148: 1547-53 (1992); Wranik, et al., J. Biol. Chem., 287: 43331-43339 (2012)). Other selectively pairing molecular pairs include, for example, the bamase-barstar pair (Deyev, et al., Nat Biotechnol, 21:1486-1492 (2003)), DNA strand pairs (Chaudri, etal., FEBS Letters, 450(l-2):23-26 (1999)) and split fluorescent protein pairs (International Patent Application Publication No. WO 2011 / 135040).

[0123] Other examples of protein scaffolds include immunoglobulin Fc regions, albumin, albumin analogues and derivatives, toxins, cytokines, chemokines and growth factors. The use of protein scaffolds in combination with antigen-binding moieties has been described (see, for example, Muller et al., 2007, J. Biol. Chem., 282:12650-12660; McDonaugh et al., 2012, Mol. Cancer Ther., 11:582-593; Vallera et al., 2005, Clin. Cancer Res., 11:3879-3888; Song et al., 2006, Biotech. Appl. Biochem., 45:147-154, and U. S. Patent Application Publication No.2009 / 0285816).

[0124] For example, fusing antigen-binding moieties such as scFvs, diabodies or single chain diabodies to albumin has been shown to improve the serum half-life of the antigen-binding moieties (Muller et al., ibid.). Antigen-binding moieties may be fused at the N- and / or C-termini of albumin, optionally via a linker. Derivatives of albumin in the form of heteromultimers that comprise two transporter polypeptides obtained by segmentation of an albumin protein such that the transporter polypeptides self-assemble to form quasi-native albumin have been described (see 34IPTS / 200238736.2International Patent Application Publication Nos. WO 2012 / 116453 and WO 2014 / 012082). As a result of the segmentation of albumin, the heteromultimer includes four termini and thus can be fused to up to four different antigen-binding moieties, optionally via linkers.

[0125] In certain embodiments, the anti-IL-31 antibody construct may comprise a protein scaffold. In some embodiments, the anti-IL-31 antibody construct may comprise a protein scaffold that is based on an immunoglobulin Fc region, an albumin or an albumin analogue or derivative. In some embodiments, the anti -IL-31 antibody construct may comprise a protein scaffold that is based on an immunoglobulin Fc region, for example, an IgG Fc region.Fc Regions

[0126] The terms “Fc region,” “Fc” or “Fc domain” as used herein refer to a C-terminal region of an immunoglobulin heavy chain that contains at least a portion of the constant region. The term includes native sequence Fc regions and variant Fc regions. Unless otherwise specified herein, numbering of amino acid residues in the Fc region or constant region is according to 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).

[0127] In certain embodiments, the anti -IL-31 antibody constructs of the present disclosure may comprise a scaffold that is based on an immunoglobulin Fc region. The Fc region may be dimeric and composed of two Fc polypeptides or alternatively, the Fc region may be composed of a single polypeptide. In certain embodiments, the anti -IL-31 antibody constructs comprise an Fc region that is dimeric and composed of two Fc polypeptides.

[0128] An “Fc polypeptide” in the context of a dimeric Fc refers to one of the two polypeptides forming the dimeric Fc domain, i. e. a polypeptide comprising one or more C-terminal constant regions of an immunoglobulin heavy chain that is capable of stable self-association. When referring to the polypeptides forming a dimeric Fc region, the terms “first Fc polypeptide” and “second Fc polypeptide” may be used interchangeably provided that the Fc region comprises one first Fc polypeptide and one second Fc polypeptide.35IPTS / 200238736.2

[0129] An Fc region may comprise a CH3 domain or it may comprise both a CH3 and a CH2 domain. For example, in certain embodiments, an Fc polypeptide of a dimeric IgG Fc region may comprise an IgG CH2 domain sequence and an IgG CH3 domain sequence. In such embodiments, the CH3 domain comprises two CH3 sequences, one from each of the two Fc polypeptides of the dimeric Fc region, and the CH2 domain comprises two CH2 sequences, one from each of the two Fc polypeptides of the dimeric Fc region. In certain embodiments, the Fc region comprises a CH3 domain, a CH2 domain and an IgG hinge region, with each Fc polypeptide comprising a CH3 domain sequence, a CH2 domain sequence and an IgG hinge region sequence.

[0130] In some embodiments, the anti-IL-31 antibody construct may comprise a scaffold that is an IgG Fc region. In some embodiments, the anti -IL-31 antibody construct may comprise a scaffold that is a human IgG Fc region. In some embodiments, the anti-IL-31 antibody construct may comprise a scaffold that is an IgGl or IgG4 Fc region. In some embodiments, the anti-IL-31 antibody construct may comprise a scaffold that is a human IgGl or IgG4 Fc region.

[0131] In certain embodiments, the anti-IL-31 antibody construct may comprise a scaffold based on an IgG Fc region, which is a homodimeric Fc region, comprising a first Fc polypeptide and a second Fc polypeptide, each comprising a CH3 sequence, and optionally a CH2 sequence and in which the amino acid sequences of the first and second Fc polypeptides are the same. In certain embodiments, the homodimeric Fc region further comprises an IgG hinge region.

[0132] In certain embodiments, the anti-IL-31 antibody construct may comprise a scaffold based on an IgG Fc region, which is a heterodimeric Fc region, comprising a first Fc polypeptide and a second Fc polypeptide, each comprising a CH3 sequence, and optionally a CH2 sequence and in which the amino acid sequences of the first and second Fc polypeptides are different. In certain embodiments, the heterodimeric Fc region further comprises an IgG hinge region. Heterodimeric Fc regions may be particularly useful in those embodiments in which the anti-IL-31 antibody construct comprises two or more different antigen-binding domains, for example, when the anti-IL-31 antibody construct is biparatopic, bispecific or multispecific.

[0133] In some embodiments, the anti-IL-31 antibody construct may comprise a scaffold based on an Fc region which comprises two CH3 sequences, at least one of which comprises one or more amino acid modifications. In some embodiments, the anti-IL-31 antibody construct may 36IPTS / 200238736.2comprise a scaffold based on an Fc region which comprises two CH3 sequences and two CH2 sequences, at least one of the CH2 sequences comprising one or more amino acid modifications. In some embodiments, the anti-IL-31 antibody construct may comprise a scaffold based on an Fc region which comprises two CH3 sequences and two CH2 sequences in which at least one of the CH3 sequences comprises one or more mutations and at least one of the CH2 sequences comprises one or more amino acid modifications.

[0134] In some embodiments, the anti-IL-31 antibody construct may comprise a heterodimeric Fc region comprising a modified CH3 domain, where the modified CH3 domain is an asymmetrically modified CH3 domain comprising one or more asymmetric amino acid modifications. As used herein, an “asymmetric amino acid modification” refers to a modification, such as a substitution or an insertion, in which an amino acid at a specific position on a first CH3 or CH2 sequence is different to the amino acid on a second CH3 or CH2 sequence at the same position. These asymmetric amino acid modifications can be a result of modification of only one of the two amino acids at the same respective amino acid position on each sequence, or different modifications of both amino acids at the same respective position on each of the first and second CH3 or CH2 sequences. Each of the first and second CH3 or CH2 sequences of a heterodimeric Fc may comprise one or more than one asymmetric amino acid modification.

[0135] In some embodiments, the anti-IL-31 antibody construct may comprise a heterodimeric Fc comprising a modified CH3 domain, where the modified CH3 domain comprises one or more amino acid modifications that promote formation of the heterodimeric Fc over formation of a homodimeric Fc. In some embodiments, one or more of the amino acid modifications are asymmetric amino acid modifications.

[0136] Amino acid modifications that may be made to the CH3 domain of an Fc in order to promote formation of a heterodimeric Fc are known in the art and include, for example, those described in International Publication No. WO 96 / 027011 (“knobs into holes”), Gunasekaran et al., 2010, J Biol Chem, 285, 19637-46 (“electrostatic steering”), Davis etal., 2010, Prot Eng Des Sci, 23(4): 195-202 (strand exchange engineered domain (SEED) technology) and Labrijn et al., 2013, Proc Natl Acad Sci USA, 110(13):5145-50 (Fab-arm exchange). Other examples include approaches combining positive and negative design strategies to produce stable asymmetrically37IPTS / 200238736.2modified Fc regions as described in International Publication Nos. WO 2012 / 058768 and WO 2013 / 063702. In certain embodiments, the anti -IL-31 antibody construct may comprise a scaffold based on a modified Fc region as described in International Publication No. WO 2012 / 058768 or WO 2013 / 063702.

[0137] Table 5 provides the amino acid sequences of a human IgGl Fc region (SEQ ID NO:66) and a human IgG4 Fc region (SEQ ID NO:67). Table 6 shows CH3 domain amino acid amino acid substitutions that promote formation of a heterodimeric Fc as described in International Patent Publication Nos. WO 2012 / 058768 and WO 2013 / 063702.

[0138] In certain embodiments, the anti-IL-31 antibody construct may comprise a heterodimeric Fc scaffold based on an IgGl or IgG4 Fc region having a modified CH3 domain comprising the amino acid substitutions of any one of Variant 1, Variant 2, Variant 3, Variant 4 or Variant 5, as shown in Table 6.Table 5: Amino Acid Sequence of the Human IgGl and IgG4 Fc Regions (CH2 and CH3 Domains)Fc Sequence SEQ Region ID NOIgGl APELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFN 66 WYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEY KCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLT CLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTV DKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGKIgG4 APEFLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSQEDPEVQFNW 67 YVDGVEVHNAKTKPREEQFNSTYRVVSVLTVLHQDWLNGKEYKC KVSNKGLPSSIEKTISKAKGQPREPQVYTLPPSQEEMTKNQVSLTCL VKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSRLTVDK SRWQEGNVFSCSVMHEALHNHYTQKSLSLSLGKTable 6: CH3 Domain Amino Acid Substitutions Promoting Heterodimer Formation Variant No Chain Mutations1 A L351Y_F405A_Y407VB T366L K392M_T394W38IPTS / 200238736.2Variant No Chain Mutations2 A L351Y_F405A_Y407VB T366L_K392L_T394W3 A T350V_L351Y_F4O5A_Y4O7VB T350V_T366L K392L_T394W4 A T350V_L351Y_F4O5A_Y4O7VB T350V_T366L K392M_T394W5 A T350V_L351Y_S4OOE_F4O5A_Y4O7VB T350V_T366L_N390R_K392M_T394W

[0139] In those embodiments in which the anti-IL-31 antibody construct comprises an IgG4 Fc region as a scaffold, the Fc region may comprise one or more amino acid mutations to decrease or eliminate Fab arm exchange and / or to improve stability. Examples of such amino acid mutations include the amino acid substitutions Y219C, G220C, S228P and R409K (see, for example, Handlogten, et al., 2020, MAbs, 12(1): 1779974; Namisaki, et al., 2020, PLoS ONE, 15(3): e0229027). In certain embodiments, the anti-IL-31 antibody construct may comprise an Fc scaffold based on an IgG4 Fc region comprising the amino acid substitution S228P (EU numbering; S241P Kabat numbering), where the Fc region may be homodimeric or heterodimeric. In certain embodiments, the anti-IL-31 antibody construct may comprise an Fc scaffold based on an IgG4 Fc region comprising the amino acid substitution R409K, where the Fc region may be homodimeric or heterodimeric. In certain embodiments, the anti-IL-31 antibody construct may comprise an Fc scaffold based on an IgG4 Fc region comprising the amino acid substitutions S228P and R409K, where the Fc region may be homodimeric or heterodimeric. In some embodiments, the anti-IL-31 antibody construct may comprise a heterodimeric IgG4 Fc region comprising the amino acid substitution R409K and amino acid substitutions of any one of Variant 1, Variant 2, Variant 3, Variant 4 or Variant 5, as shown in Table 6. In some embodiments, the anti -IL-31 antibody construct may comprise a heterodimeric IgG4 Fc region comprising the amino acid substitutions S228P and R409K and amino acid substitutions of any one of Variant 1, Variant 2, Variant 3, Variant 4 or Variant 5, as shown in Table 6.

[0140] In some embodiments, the anti-IL-31 antibody construct may comprise a scaffold based on an Fc region comprising two CH3 sequences and two CH2 sequences, at least one of the39IPTS / 200238736.2CH2 sequences comprising one or more amino acid modifications that affect the binding of Fc receptors (FcRs) to the Fc, such as receptors of the FcyRI, FcyRII and FcyRIII subclasses. In some embodiments, the anti-IL-31 antibody construct comprises a scaffold based on an IgG Fc having a modified CH2 domain, wherein the modification of the CH2 domain results in altered binding to one or more of the FcyRI, FcyRII and FcyRIII receptors.

[0141] A number of amino acid modifications to the CH2 domain that selectively alter the affinity of the Fc for different Fey receptors are known in the art (see, for example, Lu, et al., 2011, J Immunol Methods, 365(1-2): 132-41; Stavenhagen, et al. 2007, Cancer Res 67(18):8882-90; Nordstrom JL, et al., 2011, Breast Cancer Res, 13(6): R123; Stewart, etal., 2011, Protein Eng Des Sci., 24(9):671-8; Shields, et al., 2001, J Biol Chem, 276(9):6591-604; Lazar, et al., 2006, Proc Natl Acad Sci USA, 103(11):4005-10; Chu, et al., 2008, Mol Immunol, 45(15):3926-33, and International Publication No. WO 2021 / 232162). Amino acid modifications that result in increased binding and amino acid modifications that result in decreased binding can each be useful in certain indications. For example, increasing binding affinity of an Fc for FcyRIIIa (an activating receptor) may result in increased antibody dependent cell-mediated cytotoxicity (ADCC), which in turn results in increased lysis of the target cell. Decreased binding to FcyRIIb (an inhibitory receptor) likewise may be beneficial in some circumstances. In certain indications, a decrease in, or elimination of, ADCC and complement-mediated cytotoxicity (CDC) may be desirable. In such cases, modified CH2 domains comprising amino acid modifications that result in increased binding to FcyRIIb or amino acid modifications that decrease or eliminate binding of the Fc region to all Fey receptors (“knock-out” variants) may be useful.

[0142] In certain embodiments, the anti -IL-31 antibody construct comprises a scaffold based on an IgG Fc having a modified CH2 domain, in which the modified CH2 domain comprises one or more amino acid modifications that result in decreased or eliminated binding of the Fc region to all Fey receptors (i.e. a “knock-out” variant). Various publications describe strategies that have been used to engineer antibodies to produce “knock-out” variants (see, for example, Strohl, 2009, Curr Opin Biotech 20:685-691, and Strohl & Strohl, “Antibody Fc engineering for optimal antibody performance" In Therapeutic Antibody Engineering, Cambridge: Woodhead Publishing, 2012, pp 225-249). These strategies include reduction of effector function through modification of glycosylation, use of IgG2 / IgG4 scaffolds, or the introduction of mutations in the 40IPTS / 200238736.2hinge or CH2 domain of the Fc (see also, U. S. Patent Publication No. 2011 / 0212087, International Publication No. WO 2006 / 105338, U. S. Patent Publication No. 2012 / 0225058, U. S. Patent Publication No. 2012 / 0251531 and Strop et al., 2012, Mol. Biol., 420: 204-219).

[0143] Examples of mutations that may be introduced into the hinge or CH2 domain of an IgGl Fc to produce a “knock-out” variant include the amino acid substitutions E234A / E235A and E234A / E235A / D265S, which are typically introduced into both chains of the CH2 domain. In certain embodiments, the anti-IE-31 antibody construct comprises a scaffold based on an IgGl Fc having a modified CH2 domain that comprises the amino acid substitutions L234A / L235 A / D265 S in both chains of the CH2 domain.

[0144] In certain embodiments, the anti-IL-31 antibody constructs described herein may comprise a scaffold based on an IgG Fc in which native glycosylation has been modified. As is known in the art, glycosylation of an Fc may be modified to increase or decrease effector function. For example, mutation of the conserved asparagine residue at position 297 (N297) to alanine, glutamine, lysine or histidine (i.e. N297A, Q, K or H) results in an aglycoslated Fc that lacks all effector function (Bolt et al., 1993, Eur. J. Immunol., 23:403-411; Tao & Morrison, 1989, J. Immunol., 143:2595-2601).

[0145] Other amino acid mutations in the CH2 domain that may be useful include amino acid mutations that result in increased binding to the neonatal Fc receptor (FcRn). Increased binding to FcRn may improve the in vivo half-life of the antibody construct. Amino acid substitutions in the CH2 domain that enhance binding to FcRn include, for example, the amino acid substitutions M252Y / S254T / T256E (“YTE mutations”) in both chains of the CH2 domain. In certain embodiments, the anti-IL-31 antibody constructs comprise YTE mutations.

[0146] In certain embodiments, the anti-IL-31 antibody constructs comprise an IgGl Fc region and comprise knock-out mutations and YTE mutations. In certain embodiments, the anti-IL-31 antibody constructs comprise an IgG4 Fc region and comprise YTE mutations.

[0147] In certain embodiments, the anti-IL-31 antibody constructs have the format of a full-size antibody (FSA). In some embodiments, the anti-IL-31 antibody constructs have the format of an IgG FSA, for example, an IgGl or IgG4 FSA. In some embodiments, the anti-IL-3141IPTS / 200238736.2antibody construct is a FSA comprising a first heavy chain sequence (Hl), a second heavy chain sequence (H2), a first light chain sequence (LI) and a second light chain sequence (L2), where Hl pairs with LI and H2 pairs with L2. In some embodiments, the anti -IL-31 antibody construct is a monospecific FSA with a homodimeric Fc and comprises Hl, H2, LI and L2 sequence, where Hl and H2 have the same amino acid sequence, and LI and L2 have the same amino acid sequence. In some embodiments, the anti-IL-31 antibody construct is a monospecific FSA with a heterodimeric Fc and comprises Hl, H2, LI and L2 sequences, where Hl and H2 have different amino acid sequences, and LI and L2 have the same amino acid sequence. In some embodiments, the anti-IL-31 antibody construct is a biparatopic, bispecific or multispecific FSA with a heterodimeric Fc and comprises Hl, H2, LI and L2 sequences, where Hl and H2 have different amino acid sequences, and LI and L2 have different amino acid sequences.

[0148] In certain embodiments, the anti-IL-31 antibody construct is a FSA having a set of Hl, H2, LI and L2 sequences comprising the Hl, H2, LI and L2 amino acid sequences as set forth in Tables G and H for any one of variants v33559, v34731, v34734, v34735, v34736, v34737, v34738, v34739, v34740, v34741, v36535, v36539, v36540, v36541, v36542, v36545 or v36974. As is known in the art, expression of antibody heavy chain sequences in certain cell lines or from certain expression vector may result in the inclusion of a C-terminal lysine residue on one or both of the heavy chains. Accordingly, certain embodiments of the present disclosure relate to anti-IL-31 antibody constructs that are FSAs having a set of Hl, H2, LI and L2 sequences comprising the Hl, H2, LI and L2 amino acid sequences as set forth in Tables G and H for any one of variants v33559, v34731, v34734, v34735, v34736, v34737, v34738, v34739, v34740, v34741, v36535, v36539, v36540, v36541, v36542, v36545 or v36974, in which one or both of the Hl and H2 sequences comprise a C-terminal lysine.MULTISPECIFIC ANTIBODY CONSTRUCTS

[0149] Certain embodiments of the present disclosure relate to multispecific antibody constructs that specifically bind to IL-31 and to one or more other target antigens, where the one or more other target antigens are not IL-31. The multispecific antibody constructs of the present disclosure thus comprise an IL-31 antigen-binding domain and one or more additional antigenbinding domains, where the one or more additional antigen-binding domains each bind to an42IPTS / 200238736.2antigen other than IL-31. In certain embodiments, the multispecific antibody constructs are bispecific, trispecific or tetraspecific. In some embodiments, the multispecific antibody constructs are bispecific or trispecific.

[0150] In certain embodiments, the multispecific antibody constructs comprise a first antigen-binding domain that binds to IL-31 (the IL-31 antigen-binding domain) and at least a second antigen-binding domain that binds to a target antigen other than IL-31 (a second target antigen-binding domain).

[0151] In some embodiments, the multispecific antibody construct is a bispecific antibody construct and comprises at least one IL-31 antigen-binding domain and at least one second target antigen-binding domain. The bispecific antibody constructs may be, for example, bivalent, trivalent or tetravalent. Higher valencies are also contemplated in certain embodiments. In some embodiments, the bispecific antibody constructs are bivalent and comprise one IL-31 antigenbinding domain and one second target antigen-binding domain. In some embodiments, the bispecific antibody constructs are trivalent or tetravalent and comprise one or two IL-31 antigenbinding domains and one or two second target antigen-binding domains.

[0152] In certain embodiments, the multispecific antibody constructs of the present disclosure have higher specificities (for example, trispecific or tetraspecific) and include additional target antigen-binding domains, each binding to a different target antigen. For example, trispecific antibody constructs comprise at least one IL-31 antigen-binding domain, at least one second target antigen-binding domain that specifically binds to a second target antigen and at least one third target antigen-binding domain that specifically binds to a third target antigen, where the second and third target antigens are not IL-31 and are different from each other. A tetraspecific antibody construct comprises at least one IL-31 antigen-binding domain, at least one second target antigenbinding domain that specifically binds to a second target antigen, at least one third target antigenbinding domain that specifically binds to a third target antigen and at least one fourth target antigen-binding domain that specifically binds to a fourth target antigen, where the second, third and fourth target antigens are not IL-31 and are different from each other.43IPTS / 200238736.2

[0153] In accordance with the present disclosure, the at least one IL-31 antigen-binding domain comprised by the multispecific antibody constructs may be the IL-31 antigen-binding domain of any one of the embodiments described above.

[0154] In certain embodiments, the multispecific antibody constructs comprise at least a second target antigen-binding domain that binds to a second target antigen, where the second target antigen is a cytokine or a cytokine receptor. In some embodiments, the second target antigen is a cytokine or cytokine receptor associated with an inflammatory or autoimmune disorder. In some embodiments, the second target antigen is a cytokine or cytokine receptor associated with atopic dermatitis and / or psoriasis, for example, IL-13, IL-31Ra, IL-4 or IL-4Ra. In some embodiments, the second target antigen is IL-4Ra.

[0155] The additional antigen-binding domains comprised by the multispecific antibody constructs may be derived from a known antibody that binds to the target antigen, for example, from a known antibody that binds to IL- 13, IL-31Ra, IL-4 or IL-4Ra, or it may be derived from an antibody against the target antigen generated by standard antibody generation techniques. Examples of known antibodies that bind to IL- 13 include, but are not limited to, lebrikizumab, tralokinumab, dectrekumab, anrukinzumab, cendakimab, romilkimab and GSK679586. An example of a known antibody that binds to IL-31Ra is nemolizumab. Examples of known antibodies that bind to IL-4 include, but are not limited to, pascolizumab, romilkimab and GSK2434735. Examples of known antibodies that bind IL-4Ra include, but are not limited to, dupilumab, stapokibart, rademikibart, AMG317 and NM26-2198. The VH and VL sequences for these known antibodies may be obtained from publicly accessible databases, such as the Therapeutic Antibody Database (Tabs).

[0156] In certain embodiments, the second target antigen-binding domain comprised by the multispecific antibody constructs binds to IL-4Ra (an “IL-4Ra antigen-binding domain”).

[0157] In certain embodiments, the antigen-binding domains (for example, the IL-31 antigen-binding domain and the second, third and fourth target antigen-binding domains) comprised by the multispecific antibody constructs are immunoglobulin-based binding domains, such as antigen-binding antibody fragments. Examples of an antigen-binding antibody fragment include, but are not limited to, a Fab fragment (Fab), a Fab’ fragment (Fab’), a single chain Fab 44IPTS / 200238736.2(scFab), a single chain Fv (scFv) and a single domain antibody (sdAb). In certain embodiments, the antigen-binding domains are each independently a Fab or an scFv.IL-4Ra Antigen-Binding Domains

[0158] In certain embodiments, the multispecific antibody constructs of the present disclosure comprise at least one IL-31 antigen-binding domain as described in any one of the embodiments defined above and at least one IL-4Ra antigen-binding domain. In these embodiments, the at least one IL-4Ra antigen-binding domain comprised by the multispecific antibody construct is capable of specifically binding human IL-4Ra. In certain embodiments, the at least one IL-4Ra antigen-binding domain comprised by the multispecific antibody construct may be capable of binding to human IL-4Ra and to an IL-4Ra from one or more non-human species. In certain embodiments, the at least one IL-4Ra antigen-binding domain comprised by the multispecific antibody construct is capable of binding to human IL-4Ra and cynomolgus monkey IL-4Ra.

[0159] In certain embodiments, the multispecific antibody constructs of the present disclosure are bispecific anti-IL-31 x anti-IL-4Ra antibody constructs comprising at least one IL-31 antigen-binding domain as described in any one of the embodiments defined above and at least one IL-4Ra antigen-binding domain.

[0160] The protein sequences of IL-4Ra from various sources are known in the art and readily available from publicly accessible databases, such as GenBank or UniProtKB. Examples of human IL-4Ra sequences include those provided under NCBI reference numbers P24394, NP_000409.1, NP_001244335.1 and NP_001244336.1. IL-4Ra is a transmembrane protein with a 207 amino acid extracellular domain, a 24 amino acid transmembrane segment, and a 569 amino acid cytoplasmic domain. Anti-IL-4Ra antigen-binding domains typically bind to the extracellular domain of IL-4Ra. The amino acid sequence of the extracellular domain of an exemplary human IL-4Ra protein is provided in Table 7 as SEQ ID NO: 68 (amino acids 24-232 of NCBI Accession No. P24394).45IPTS / 200238736.2Table 7: Amino Acid Sequence of the Extracellular Domain of Human IL-4Ra Organism Sequence SEQ ID NOHomo GNMKVLQEPTCVSDYMSISTCEWKMNGPTNCSTELRLLYQL 68 sapiens VFLLSEAHTCIPENNGGAGCVCHLLMDDVVSADNYTLDLW AGQQLLWKGSFKPSEHVKPRAPGNLTVHTNVSDTLLLTWSN PYPPDNYLYNHLTYAVNIWSENDPADFRIYNVTYLEPSLRIA ASTLKSGISYRARVRAWAQCYNTTWSEWSPSTKWHNSYREP FEQH

[0161] In certain embodiments, the one or more IL-4Ra antigen-binding domains comprised by the multispecific antibody constructs is an antigen-binding antibody fragment, such as a Fab fragment (Fab), a Fab’ fragment (Fab’), a single chain Fab (scFab), a single chain Fv (scFv) and a single domain antibody (sdAb). In certain embodiments, the IL-4Ra antigen-binding domain is a Fab or an scFv. In some embodiments, the IL-4Ra antigen-binding domain is a Fab.

[0162] In certain embodiments, the IL-4Ra antigen-binding domain(s) comprised by the multispecific antibody constructs of the present disclosure may be derived from a known anti-IL-4Ra antibody, such as dupilumab, stapokibart, rademikibart, AMG317 or NM26-2198. In some embodiments, the multispecific antibody constructs may comprise one of the IL-4Ra antigenbinding domains described in the Examples provided herein, for example, the IL-4Ra binding domain of any one of the variants v38597, v38504, v38681, v38727, v41791, v43181, v43182, v43184, v43185, v43186, v43187, v43188, v43189, v43190, v43191, v43192 or v43193. In some embodiments, the multispecific antibody constructs may comprise the IL-4Ra antigen-binding domain from any one of variants v38597, v38504, v38681, v38727, v41791, v43181, v43182, v43184, v43188, v43190 or v43193.

[0163] The CDR sequences of the IL-4Ra antigen-binding domains comprised by variants v38597, v38504, v41791, v43181, v43182, v43184, v43188, v43190 and v43193 described in the Examples herein are provided in Fig. 18 (Table Cl) and Fig. 19 (Table C2) and the VHand VL sequences are provided in Fig. 20 (Tables DI and D2). The CDR sequences and the VH and VL sequences of the IL-4Ra antigen-binding domains comprised by variants v43184, v43185, v43186,46IPTS / 200238736.2v43187, v43189, v43192 and v43193 described in the Examples herein are provided in Tables G and H.

[0164] In certain embodiments, the multispecific antibody constructs of the present disclosure comprise at least one IL-4Ra binding domain, where the IL-4Ra binding domain comprises the heavy chain CDR sequences (HCDR1, HCDR2, HCDR3) and the light chain CDR sequences (LCDR1, LCDR2, LCDR3) of the IL-4Ra binding domain of any one of variants v38597, v38504, v38681, v38727, v41791, v43181, v43182, v43184, v43185, v43186, v43187, v43188, v43189, v43190, v43191, v43192 and v43193. In some embodiments, the multispecific antibody constructs comprise at least one IL-4Ra binding domain, where the IL-4Ra binding domain comprises the heavy chain CDR sequences (HCDR1, HCDR2, HCDR3) and the light chain CDR sequences (LCDR1, LCDR2, LCDR3) of the IL-4Ra binding domain of any one of variants v38597, v38504, v38681, v38727, v41791, v43181, v43182, v43184, v43188, v43190 or v43193.

[0165] In certain embodiments, the multispecific antibody constructs of the present disclosure comprise at least one IL-4Ra antigen-binding domain, where the IL-4Ra antigenbinding domain comprises the CDR sequences of the VH domain as set forth in any one of SEQ ID NOs: 103, 104, 106, 107, 108, 109, 110 or 219. In certain embodiments, the multispecific antibody constructs of the present disclosure comprise at least one IL-4Ra antigen-binding domain, where the IL-4Ra antigen-binding domain comprises the CDR sequences of the VL domain as set forth in any one of SEQ ID NOs: 105, 111, 112, 113 or 114.

[0166] In certain embodiments, the multispecific antibody constructs of the present disclosure comprise at least one IL-4Ra antigen-binding domain, where the IL-4Ra antigenbinding domain comprises the CDR sequences of the VH domain as set forth in any one of SEQ ID NOs: 103, 104, 106, 107, 108, 109, 110 or 219, and the CDR sequences of the VL domain as set forth in any one of SEQ ID NOs: 105, 111, 112, 113 or 114.

[0167] In certain embodiments, the multispecific antibody constructs of the present disclosure comprise at least one IL-4Ra antigen-binding domain, where the IL-4Ra antigenbinding domain comprises a VH domain comprising an HCDR1 amino acid sequence comprising the sequence as set forth in any one of SEQ ID NOs: 68, 71, 81 or 82, an HCDR2 amino acid 47IPTS / 200238736.2sequence comprising the sequence as set forth in any one of SEQ ID NOs: 69, 72, 94, 95, 99, 100, 101, 102 or 218, and an HCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 70 or 73. In some embodiments, the multispecific antibody constructs comprise at least one IL-4Ra antigen-binding domain, where the IL-4Ra antigen-binding domain comprises a VH domain comprising an HCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 68 or 71, an HCDR2 amino acid sequence comprising the sequence as set forth in any one of SEQ ID NOs: 69, 72, 94, 95, 99, 100, 101, 102 or 218, and an HCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 70 or 73.

[0168] In certain embodiments, the multispecific antibody constructs of the present disclosure comprise at least one IL-4Ra antigen-binding domain, where the IL-4Ra antigenbinding domain comprises a VL domain comprising an LCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 86 or 89, an LCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 87 or 90, and an LCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 88.

[0169] In certain embodiments, the multispecific antibody constructs of the present disclosure comprise at least one IL-4Ra antigen-binding domain, where the IL-4Ra antigenbinding domain comprises a VH domain comprising an HCDR1 amino acid sequence comprising the sequence as set forth in any one of SEQ ID NOs: 68, 71, 81 or 82, an HCDR2 amino acid sequence comprising the sequence as set forth in any one of SEQ ID NOs: 69, 72, 94, 95, 99, 100, 101, 102 or 218, and an HCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 70 or 73, and a VL domain comprising an LCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 86 or 89, an LCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 87 or 90, and an LCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 88. In some embodiments, the multispecific antibody constructs of the present disclosure comprise at least one IL-4Ra antigen-binding domain, where the IL-4Ra antigen-binding domain comprises a VH domain comprising an HCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 68 or 71, an HCDR2 amino acid sequence comprising the sequence as set forth in any one of SEQ ID NOs: 69, 72, 94, 95, 99, 100, 101, 102 or 218, and an HCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 70 or 73, and a VL domain comprising an LCDR1 amino acid sequence comprising the48IPTS / 200238736.2sequence as set forth in SEQ ID NO: 86 or 89, an LCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 87 or 90, and an LCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 88.

[0170] In certain embodiments, the multispecific antibody constructs of the present disclosure comprise at least one IL-4Ra antigen-binding domain, where the IL-4Ra antigenbinding domain comprises a VH domain comprising:(i) an HCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 68 or 71, an HCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 69 or 72, and an HCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 70 or 73, or(ii) an HCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 81 or 82, an HCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 69 or 72, and an HCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 70 or 73, or(iii) an HCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 68 or 71, an HCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 94 or 95, and an HCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 70 or 73, or(iv) an HCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 68 or 71, an HCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 94 or 99, and an HCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 70 or 73, or(v) an HCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 68 or 71, an HCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 94 or 100, and an HCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 70 or 73, or49IPTS / 200238736.2(vi) an HCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 68 or 71, an HCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 69 or 101, and an HCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 70 or 73, or(vii) an HCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 68 or 71, an HCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 69 or 102, and an HCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 70 or 73, or(viii) an HCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 68 or 71, an HCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 94 or 218, and an HCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 70 or 73, and

[0171] a VL domain comprising an LCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 86 or 89, an LCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 87 or 90, and an LCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 88.

[0172] In certain embodiments, the multispecific antibody constructs of the present disclosure comprise at least one IL-4Ra antigen-binding domain, where the IL-4Ra antigenbinding domain comprises a VH domain having a sequence that is at least about 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the VH sequence as set forth in any one of SEQ ID NOs: 103, 104, 106, 107, 108, 109, 110 or 219. In some embodiments, the multispecific antibody constructs comprise at least one IL-4Ra antigen-binding domain, where the IL-4Ra antigenbinding domain comprises a VH domain having a sequence that is at least about 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the VH sequence as set forth in SEQ ID NO: 103. In some embodiments, the multispecific antibody constructs comprise at least one IL-4Ra antigen-binding domain, where the IL-4Ra antigen-binding domain comprises a VH domain having a sequence as set forth in any one ofSEQ ID NOs: 103, 104, 106, 107, 108, 109, 110 or 219.50IPTS / 200238736.2

[0173] In certain embodiments, the multispecific antibody constructs of the present disclosure comprise at least one IL-4Ra antigen-binding domain, where the IL-4Ra antigenbinding domain comprises a VL domain having a sequence that is at least about 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the VL sequence as set forth in any one of SEQ ID NOs: 105, 111, 112, 113 or 114. In some embodiments, the multispecific antibody constructs comprise at least one IL-4Ra antigen-binding domain, where the IL-4Ra antigen-binding domain comprises a VL domain having a sequence that is at least about 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the VL sequence as set forth in SEQ ID NO: 105. In some embodiments, the multispecific antibody constructs comprise at least one IL-4Ra antigen-binding domain, where the IL-4Ra antigen-binding domain comprises a VL domain having a sequence as set forth in any one of SEQ ID NOs: 105, 111, 112, 113 or 114.

[0174] In certain embodiments, the multispecific antibody constructs of the present disclosure comprise at least one IL-4Ra antigen-binding domain, where the IL-4Ra antigenbinding domain comprises a VH domain having a sequence that is at least about 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the VH sequence as set forth in any one of SEQ ID NOs: 103, 104, 106, 107, 108, 109, 110 or 219, and a VL domain having a sequence that is at least about 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the VL sequence as set forth in any one of SEQ ID NOs: 105, 111, 112, 113 or 114. In some embodiments, the multispecific antibody constructs comprise at least one IL-4Ra antigen-binding domain, where the IL-4Ra antigenbinding domain comprises a VH domain having a sequence that is at least about 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the VH sequence as set forth in SEQ ID NO: 103, and a VL domain having a sequence that is at least about 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the VL sequence as set forth in SEQ ID NO: 105. In some embodiments, the multispecific antibody constructs comprise at least one IL-4Ra antigen-binding domain, where the IL-4Ra antigen-binding domain comprises a VH domain having a sequence as set forth in any one ofSEQ ID NOs: 103, 104, 106, 107, 108, 109, 110 or 219, and a VL domain having a sequence as set forth in any one of SEQ ID NOs: 105, 111, 112, 113 or 114.51IPTS / 200238736.2Multispecific Formats

[0175] The multispecific antibody constructs described herein comprise at least one IL-31 antigen-binding domain and at least one additional antigen-binding domain, for example, an IL-4Ra antigen-binding domain. In certain embodiments, the multispecific antibody constructs are bivalent (i.e. comprise two antigen-binding domains), trivalent (i.e. comprise three antigenbinding domains) or tetravalent (i.e. comprise four antigen-binding domains). In certain embodiments, the multispecific antibody constructs may further comprise a scaffold, such as a scaffold as described in any one of the embodiments defined above, and the antigen-binding domains are linked directly or indirectly (for example, via a linker or via one of the other antigenbinding domains) to the scaffold.

[0176] In certain embodiments, the multispecific antibody constructs may lack a scaffold and thus comprise two or more antigen-binding domains optionally operably linked by one or more linkers. In such antibody constructs, the antigen-binding domains may be in the form of scFvs, Fabs, sdAbs, or a combination thereof. For example, using scFvs as the antigen-binding domains, formats such as a tandem scFv ((scFv)2 or taFv) may be constructed, in which the scFvs are connected together by a flexible linker. scFvs may also be used to construct diabody formats, which comprise two scFvs connected by a short linker (usually about 5 amino acids in length). The restricted length of the linker results in dimerization of the scFvs in a head-to-tail manner. In any of the preceding formats, the scFvs may be further stabilized by inclusion of an interdomain disulfide bond. For example, a disulfide bond may be introduced between VL and VH through introduction of an additional cysteine residue in each chain (for example, at position 44 in VH and position 100 in VL) (see, for example, Fitzgerald c7 al., 1997, Protein Engineering, 10:1221-1225), or a disulfide bond may be introduced between two VHs to provide a construct having a DART format (see, for example, Johnson et al., 2010, J Mol. Biol., 399:436-449).

[0177] Similarly, formats comprising two sdAbs, such as VHs or VHHs, connected together through a suitable linker may be employed in some embodiments. Other examples of antibody construct formats that lack a scaffold include those based on Fab fragments, for example, Fab2 and F(ab’)2 formats, in which the Fab fragments are connected through a linker or an IgG hinge region.52IPTS / 200238736.2

[0178] Combinations of antigen-binding domains in different forms may also be employed to generate alternative scaffold-less formats. For example, an scFv or a sdAb may be fused to the C-terminus of either or both of the light and heavy chain of a Fab fragment resulting in a bivalent (Fab-scFv / sdAb) construct.

[0179] In certain embodiments, the multispecific antibody constructs may be in an antibody format that is based on an immunoglobulin (Ig), such as an IgG class immunoglobulin, and thus comprise an IgG Fc region as a scaffold. In certain embodiments, the multispecific antibody constructs may be based on an IgGl, IgG2, IgG3 or IgG4 immunoglobulin. In some embodiments, the multispecific antibody constructs may be based on an IgGl or IgG4 immunoglobulin. In the context of the present disclosure, when an antibody construct is based on a specified immunoglobulin isotype, it is meant that antibody construct comprises all or a portion of the constant region of the specified immunoglobulin isotype. For example, a multispecific antibody construct based on a given Ig isotype comprises an Ig scaffold to which at least one antigen-binding domain is operably linked, where the scaffold comprises an Fc region from the given isotype and optionally an Ig hinge region from the same or a different isotype. It is to be understood that the multispecific antibody constructs may also comprise hybrids of isotypes and / or subclasses in some embodiments. It is also to be understood that the Fc region and / or hinge region may optionally be modified to impart one or more desirable functional properties as is known in the art. Examples of such modified Fc and hinge regions are described above.

[0180] In certain embodiments, the multispecific antibody constructs comprise a scaffold based on an IgGl or IgG4 Fc region. Examples of such scaffolds are described in detail above. In some embodiments, the multispecific antibody constructs comprise two antigen-binding domains (i.e. are bivalent) and a scaffold based on an IgGl or IgG4 Fc region. In some embodiments, the multispecific antibody constructs comprise two antigen-binding domains and a scaffold based on an IgG4 Fc region. In some embodiments, the multispecific antibody constructs comprise three antigen-binding domains (i.e. are trivalent) and a scaffold based on an IgGl or IgG4 Fc region. In some embodiments, the multispecific antibody constructs comprise three antigen-binding domains and a scaffold based on an IgG4 Fc region. In some embodiments, the multispecific antibody constructs comprise four antigen-binding domains (i.e. are tetravalent) and a scaffold based on an IgGl or IgG4 Fc region. In some embodiments, the multispecific antibody constructs comprise53IPTS / 200238736.2four antigen-binding domains and a scaffold based on an IgG4 Fc region. In those embodiments in which the multispecific antibody construct comprises an Fc region, the Fc region typically comprises a first Fc polypeptide and a second Fc polypeptide.

[0181] A bivalent multispecific antibody construct comprises one antigen-binding domain that binds to IL-31 and a second antigen-binding domain that binds to a second target antigen (i.e. is bispecific), whereas a trivalent multispecific antibody construct may be bispecific (comprising two antigen-binding domains that bind to one target antigen and a third antigen-binding domain that binds to a different target antigen) or it may be trispecific (comprising three antigen-binding domains that each bind to a different target antigen). Similarly, a tetravalent multispecific antibody construct may be bispecific (for example, comprising two antigen-binding domains that bind to one target antigen and two antigen-binding domains that binds to a different target antigen) or it may be trispecific (comprising two antigen-binding domains that bind to a first target antigen and two other antigen-binding domains that each bind to a different target antigen) or it may be tetraspecific (comprising four antigen-binding domains that each bind to a different target antigen).

[0182] In some embodiments, the multispecific antibody constructs are bivalent and bispecific and comprise a scaffold based on an IgGl or IgG4 Fc region. In some embodiments, the multispecific antibody constructs are trivalent and bispecific and comprise a scaffold based on an IgGl or IgG4 Fc region. In some embodiments, the multispecific antibody constructs are trivalent and trispecific and comprise a scaffold based on an IgGl or IgG4 Fc region. In some embodiments, the multispecific antibody constructs are tetravalent and bispecific and comprise a scaffold based on an IgGl or IgG4 Fc region. In some embodiments, the multispecific antibody constructs are tetravalent and trispecific and comprise a scaffold based on an IgGl or IgG4 Fc region. In some embodiments, the multispecific antibody constructs are tetravalent and tetraspecific and comprise a scaffold based on an IgGl or IgG4 Fc region.

[0183] As would be appreciated by one of skill in the art, the multispecific antibody constructs may be constructed in various formats. For example, each of the antigen-binding domains comprised by the multispecific antibody may be in a different format (for example, Fab, scFv), or they may all be in the same format. Where the multispecific antibody construct comprises an Fc region, each antigen-binding domain may be operably linked to the N-terminus of an Fc54IPTS / 200238736.2polypeptide, to the C-terminus of an Fc polypeptide, or to one of the other antigen-binding domains.

[0184] In certain embodiments, the antigen-binding domains comprised by the multispecific antibody construct are all in Fab format. In some embodiments, at least one of the antigen-binding domains comprised by the multispecific antibody construct is in Fab format and the other antigen-binding domains are in Fab or scFv format. In some embodiments, at least one of the antigen-binding domains comprised by the multispecific antibody construct is in scFv format and the other antigen-binding domains are in Fab or scFv format. In some embodiments, the antigen-binding domains comprised by the multispecific antibody construct are all in scFv format.

[0185] In certain embodiments, the multispecific antibody constructs are bivalent and comprise one IL-31 antigen-binding domain and one second target antigen-binding domain (for example, an IL-4Ra antigen-binding domain) and an IgG Fc region, where (a) both antigenbinding domains are in Fab format, or (b) both antigen-binding domains are in scFv format, or (c) one antigen-binding domain is in Fab format and the other is in scFv format. In some embodiments, in such bivalent antibody constructs, the IgG Fc region is an IgGl or IgG4 Fc region. In some embodiments, in such bivalent antibody constructs, one antigen-binding domain is linked to the N-terminus of one Fc polypeptide and the other antigen-binding domain is linked to the N-terminus of the other Fc polypeptide. In certain embodiments, in such bivalent antibody constructs, each of the antigen-binding domains is linked to the N-terminus of its respective Fc polypeptide via an IgG hinge region.

[0186] In those embodiments in which the multispecific antibody constructs comprise a scaffold based on an IgGl or IgG4 Fc region, the Fc region will typically comprise a heterodimeric Fc comprising amino acid substitutions in the CH3 domain that promote formation of a heterodimeric Fc over a homodimeric Fc, as described above. In certain embodiments in which the multispecific antibody constructs comprise a scaffold based on an IgGl or IgG4 Fc region, the Fc region may comprise a modified CH3 domain comprising the amino acid substitutions of any one of Variant 1, Variant 2, Variant 3, Variant 4 or Variant 5, as shown in Table 6.

[0187] When the multispecific antibody construct comprises two or more antigen-binding domains in Fab format, the CHI and CL domains of each Fab may comprise sets of mutations to 55IPTS / 200238736.2drive the correct pairing between the heavy and light chains of each antigen-binding domain. Examples of sets of mutations that may be used in this context include those described in International Patent Publication Nos. WO 2014 / 082179, WO 2015 / 181805 and WO 2017 / 059551. In some embodiments, the multispecific antibody constructs comprise at least two antigen-binding domains in Fab format and each antigen-binding domain comprises a set of mutations to drive the correct pairing between the heavy and light chains of the antigen-binding domain, where the set of mutations is Set 1 or Set 2 as shown in Table 8.Table 8: Sets of CH1 / CL Mutations to Promote Correct PairingSet of Chain Mutations*Mutations1 CHI HCA A141W / L145E / K147T / Q175ECL LCA Fl 16A / Q124R / L135V / T178RCHI HCB Q175RCL LCB Q124E / L135W / T178E / T180E2 CHI HCA L145RCL LCA Q124E / V133ECHI HCB L145E / K147T / Q175ECL LCB Q124R / T178R* Numbering of amino acid positions according to EU56IPTS / 200238736.2

[0188] In certain embodiments, the multispecific antibody constructs comprise at least an IL-31 antigen-binding domain and an IL-4Ra antigen-binding domain. In some embodiments, the multispecific antibody constructs comprise an IL-31 antigen-binding domain and an IL-4Ra antigen-binding domain, where the HCDR1, HCDR2, HCDR3, LCDR1, LCDR2 and LCDR3 sequences of the IL-31 antigen-binding domain and the HCDR1, HCDR2, HCDR3, LCDR1, LCDR2 and LCDR3 sequences of the IL-4Ra antigen-binding domain have amino acid sequences as set forth for the HCDR1, HCDR2, HCDR3, LCDR1, LCDR2 and LCDR3 of the respective IL-31 antigen-binding domain and IL-4Ra antigen-binding domain of any one of variants v38681, v38727, v41791, v43181, v43182, v43184, v43185, v43186, v43187, v43188, v43189, v43190, v43191, v43192 or v43193 (see Tables G and H). In certain embodiments, the multispecific antibody constructs comprise an IL-31 antigen-binding domain and an IL-4Ra antigen-binding domain, where the VH and VL sequences of the IL-31 antigen-binding domain and the VH and VL sequences of the IL-4Ra antigen-binding domain have amino acid sequences as set forth for the VH and VL of the respective IL-31 antigen-binding domain and IL-4Ra antigen-binding domain of any one of variants v38681, v38727, v41791, v43181, v43182, v43184, v43185, v43186, v43187, v43188, v43189, v43190, v43191, v43192 or v43193 (see Tables G and H).

[0189] In certain embodiments, the multispecific antibody constructs are bispecific and bivalent and comprise one IL-31 antigen-binding domain and one IL-4Ra antigen-binding domain. In some embodiments, the multispecific antibody constructs are bispecific and bivalent and comprise an IL-31 antigen-binding domain and an IL-4Ra antigen-binding domain, where the HCDR1, HCDR2, HCDR3, LCDR1, LCDR2 and LCDR3 sequences of the IL-31 antigen-binding domain and the HCDR1, HCDR2, HCDR3, LCDR1, LCDR2 and LCDR3 sequences of the IL-4Ra antigen-binding domain have amino acid sequences as set forth for the HCDR1, HCDR2, HCDR3, LCDR1, LCDR2 and LCDR3 of the respective IL-31 antigen-binding domain and IL-4Ra antigen-binding domain of any one of variants v38681, v38727, v41791, v43181, v43182, v43184, v43185, v43186, v43187, v43188, v43189, v43190, v43191, v43192 or v43193 (see Tables G and H). In certain embodiments, the multispecific antibody constructs are bispecific and bivalent and comprise an IL-31 antigen-binding domain and an IL-4Ra antigen-binding domain, where the VH and VL sequences of the IL-31 antigen-binding domain and the VH and VL sequences of the IL-4Ra antigen-binding domain have amino acid sequences as set forth for the VH and VL of the respective IL-31 antigen-binding domain and IL-4Ra antigen-binding domain 57IPTS / 200238736.2of any one of variants v38681, v38727, v41791, v43181, v43182, v43184, v43185, v43186, v43187, v43188, v43189, v43190, v43191, v43192 or v43193 (see Tables G and H).

[0190] In certain embodiments, the multispecific antibody constructs have the format of a full-size antibody (FSA), for example, an IgGl or IgG4 FSA. In some embodiments, the multispecific antibody construct is an FSA comprising a first heavy chain sequence (Hl), a second heavy chain sequence (H2), a first light chain sequence (LI) and a second light chain sequence (L2), where Hl pairs with LI and H2 pairs with L2, and where Hl and H2 have different amino acid sequences, and LI and L2 have different amino acid sequences.

[0191] In certain embodiments, the multispecific antibody construct is a FSA having a set of Hl, H2, LI and L2 sequences comprising the Hl, H2, LI and L2 amino acid sequences as set forth in Tables G and H for any one of variants v38681, v38727, v41791, v43181, v43182, v43184, v43185, v43186, v43187, v43188, v43189, v43190, v43191, v43192 or v43193. In some embodiments, the multispecific antibody construct is a FSA having a set of Hl, H2, LI and L2 sequences comprising the Hl, H2, LI and L2 amino acid sequences as set forth in Tables G and H for any one of variants v38681, v38727, v41791, v43181, v43182, v43188, v43190 or v43193.

[0192] As is known in the art, expression of antibody heavy chain sequences in certain cell lines or from certain expression vector may result in the inclusion of a C-terminal lysine residue on one or both of the heavy chains. Accordingly, certain embodiments of the present disclosure relate to multispecific antibody constructs that are FSAs having a set of Hl, H2, LI and L2 sequences comprising the Hl, H2, LI and L2 amino acid sequences as set forth in Tables G and H for any one of variants v38681, v38727, v41791, v43181, v43182, v43184, v43185, v43186, v43187, v43188, v43189, v43190, v43191, v43192 or v43193, in which one or both of the Hl and H2 sequences comprise a C-terminal lysine.METHODS OF PREPARING ANTIBODY CONSTRUCTS

[0193] The anti-IL-31 antibody constructs and multispecific antibody constructs described herein may be produced using standard recombinant methods known in the art (see, for example, U. S. Patent No. 4,816,567 and “Antibodies: A Laboratory Manual, ” 2ndEdition, Ed. Greenfield, Cold Spring Harbor Laboratory Press, New York, 2014).58IPTS / 200238736.2

[0194] Typically, for recombinant production of an antibody construct, a polynucleotide or set of polynucleotides encoding the antibody construct is generated and inserted into one or more vectors for further cloning and / or expression in a host cell. Polynucleotide(s) encoding the antibody construct may be produced by standard methods known in the art (see, for example, Ausubel et al., Current Protocols in Molecular Biology, John Wiley & Sons, New York, 1994 & update, and “ Antibodies: A Laboratory Manual,” 2ndEdition, Ed. Greenfield, Cold Spring Harbor Laboratory Press, New York, 2014). As would be appreciated by one of skill in the art, the number of polynucleotides required for expression of the antibody construct will be dependent on the format of the construct, including whether or not the antibody construct comprises a scaffold. For example, for a monospecific antibody in full-size antibody format with two Fab antigenbinding domains, a set of two polynucleotides will be required: one polynucleotide encoding the heavy chain and one polynucleotide encoding the light chain. Whereas, for a multispecific antibody construct in a full-size antibody format with two Fab antigen-binding domains, each binding a different antigen, two polynucleotides each encoding a different heavy chain and two polynucleotides each encoding a different light chain will be required. When multiple polynucleotides are required, they may be incorporated into one vector or into more than one vector.

[0195] Generally, for expression, the polynucleotide or set of polynucleotides is incorporated into an expression vector or vectors together with one or more regulatory elements, such as transcriptional elements, which are required for efficient transcription of the polynucleotide. Examples of such regulatory elements include, but are not limited to, promoters, enhancers, terminators, and polyadenylation signals. One skilled in the art will appreciate that the choice of regulatory elements is dependent on the host cell selected for expression of the antibody construct and that such regulatory elements may be derived from a variety of sources, including bacterial, fungal, viral, mammalian or insect genes. The expression vector may optionally further contain heterologous nucleic acid sequences that facilitate expression or purification of the expressed protein. Examples include, but are not limited to, signal peptides and affinity tags such as metal-affinity tags, histidine tags, avidin / streptavidin encoding sequences, glutathione-S-transferase (GST) encoding sequences and biotin encoding sequences. The expression vector may be an extrachromosomal vector or an integrating vector.59IPTS / 200238736.2

[0196] Suitable host cells for cloning or expression of the antibody constructs include various prokaryotic or eukaryotic cells as known in the art. Eukaryotic host cells include, for example, mammalian cells, plant cells, insect cells and yeast cells (such as Saccharomyces or Pichia cells). Prokaryotic host cells include, for example, E. coli, A. salmonicida or B. subtilis cells.

[0197] In certain embodiments, the antibody construct may be produced in bacteria, in particular when glycosylation and Fc effector function are not needed, as described for example in U. S. Patent Nos. 5,648,237; 5,789,199, and 5,840,523, and in Charlton, Methods in Molecular Biology, Vol. 248, pp. 245-254, B. K. C. Lo, ed., Humana Press, Totowa, N. J., 2003.

[0198] Eukaryotic microbes such as fdamentous fungi or yeast may be suitable expression host cells in certain embodiments, in particular fungi and yeast strains whose glycosylation pathways have been “humanized” resulting in the production of an antibody construct with a partially or fully human glycosylation pattern (see, for example, Gemgross, 2004, Nat. Biotech. 22:1409-1414, and Li et al., 2006, Nat. Biotech. 24:210-215).

[0199] Suitable host cells for the expression of glycosylated antibody constructs are usually eukaryotic cells. For example, U. S. Patent Nos. 5,959,177, 6,040,498, 6,420,548, 7,125,978 and 6,417,429 describe PLANTIBODIES™ technology for producing antigen-binding constructs in transgenic plants. Mammalian cell lines adapted to grow in suspension may be particularly useful for expression of antibody constructs. Examples include, but are not limited to, monkey kidney CV1 line transformed by SV40 (COS-7), human embryonic kidney (HEK) line 293 or 293 cells (see, for example, Graham et al., \9T1, J. Gen Virol., 36:59), baby hamster kidney cells (BHK), mouse sertoli TM4 cells (see, for example, Mather, 1980, Biol Reprod, 23:243-251), monkey kidney cells (CV1), African green monkey kidney cells (VERO-76), human cervical carcinoma (HeLa) cells, canine kidney cells (MDCK), buffalo rat liver cells (BRL 3A), human lung cells (W138), human liver cells (Hep G2), mouse mammary tumour (MMT 060562), TRI cells (see, for example, Mather et al., 1982, Annals N. Y. Acad Sci, 383:44-68), MRC 5 cells, FS4 cells, Chinese hamster ovary (CHO) cells (including DHFR CHO cells, see Urlaub et al., 1980, Proc Natl Acad Sci USA, 77:4216), and myeloma cell lines (such as Y0, NS0 and Sp2 / 0). Exemplary mammalian host cell lines suitable for production of antibody constructs are reviewed60IPTS / 200238736.2in Yazaki & Wu, Methods in Molecular Biology, Vol. 248, pp. 255-268 (B. K. C. Lo, ed., Humana Press, Totowa, N. J., 2003).

[0200] In certain embodiments, the host cell used to produce the antibody constructs may be a transient or stable higher eukaryotic cell line, such as a mammalian cell line. In some embodiments, the host cell may be a mammalian HEK293T, CHO, HeLa, NS0 or COS cell line, or a cell line derived from any one of these cell lines. In some embodiments, the host cell may be a stable cell line that allows for mature glycosylation of the antibody construct.

[0201] The host cells comprising the expression vector(s) encoding the antibody construct may be cultured using routine methods to produce the antibody construct. Alternatively, in some embodiments, host cells comprising the expression vector(s) encoding the antibody construct may be used therapeutically or prophylactically to deliver the antibody construct to a subject, or polynucleotides or expression vectors may be administered to a cell from a subject ex vivo and the cell then returned to the body of the subject.

[0202] Typically, the antibody constructs are purified after expression. Proteins may be isolated or purified in a variety of ways known to those skilled in the art (see, for example, Protein Purification: Principles and Practice, 3rdEd., Scopes, Springer-Verlag, NY, 1994). Standard purification methods include chromatographic techniques, including ion exchange, hydrophobic interaction, affinity, sizing (or gel filtration), and reverse-phase, carried out at atmospheric pressure or at high pressure using systems such as HPLC or UPLC. Additional purification methods include electrophoretic, immunological, precipitation, dialysis and chromatofocusing techniques. Ultrafiltration and diafiltration techniques, in conjunction with protein concentration, are also useful. As is well known in the art, a variety of natural proteins bind Fc and antibodies, and these proteins may be used for purification of certain antibody constructs. For example, the bacterial proteins A and G bind to the Fc region. Likewise, the bacterial protein L binds to the Fab region of some antibodies. Purification may also be enabled by a particular fusion partner. For example, antibodies may be purified using glutathione resin if a GST fusion is employed, Ni+2affinity chromatography if a His-tag is employed or immobilized anti-flag antibody if a flagtag is used. The degree of purification necessary will vary depending on the use of the antibody constructs. In some instances, no purification may be necessary.61IPTS / 200238736.2

[0203] In certain embodiments, the antibody constructs are substantially pure. The term “substantially pure” (or “substantially purified”) when used in reference to an antibody construct described herein, means that the antibody construct is substantially or essentially free of components that normally accompany or interact with the protein as found in its naturally occurring environment, such as a native cell, or a host cell in the case of recombinantly produced construct. In certain embodiments, an antibody construct that is substantially pure is a protein preparation having less than about 30%, less than about 25%, less than about 20%, less than about 15%, less than about 10%, or less than about 5% (by dry weight) of contaminating protein.

[0204] Certain embodiments of the present disclosure relate to a method of making an anti-IL-31 antibody construct or a multispecific antibody construct as described herein comprising culturing a host cell into which one or more polynucleotides encoding the antibody construct, or one or more expression vectors encoding the antibody construct, have been introduced, under conditions suitable for expression of the antibody construct, and optionally recovering the antibody construct from the host cell or from host cell culture medium.Post-Translational Modifications

[0205] In certain embodiments, the antibody constructs described herein may comprise one or more post-translational modifications. Such post-translational modifications may occur in vivo, or they be conducted in vitro after isolation of the antibody construct from the host cell.

[0206] Post-translational modifications include various modifications as are known in the art (see, for example, Proteins - Structure and Molecular Properties, 2nd Ed., T. E. Creighton, W. H. Freeman and Company, New York, 1993; Post-Translational Covalent Modification of Proteins, B. C. Johnson, Ed., Academic Press, New York, pgs. 1-12, 1983; Seifter et al., 1990, Meth. Enzymol., 182:626-646, and Rattan et al., 1992, Aww. N. Y. Acad. Sci., 663:48-62). In those embodiments in which the antibody construct comprises one or more post-translational modifications, the construct may comprise the same type of modification at one or several sites, or it may comprise different modifications at different sites.

[0207] Examples of post-translational modifications include glycosylation, acetylation, phosphorylation, amidation, derivatization by known protecting / blocking groups, formylation,62IPTS / 200238736.2oxidation, reduction, proteolytic cleavage or specific chemical cleavage by cyanogen bromide, trypsin, chymotrypsin, papain, V8 protease or NaBH4.

[0208] Other examples of post-translational modifications include, for example, addition or removal of N-linked or O-linked carbohydrate chains, chemical modifications of N-linked or O-linked carbohydrate chains, processing of N-terminal or C-terminal ends, attachment of chemical moieties to the amino acid backbone, and addition or deletion of an N-terminal methionine residue resulting from prokaryotic host cell expression.

[0209] Post-translational modifications may also include modification with a detectable label, such as an enzymatic, fluorescent, luminescent, isotopic or affinity label to allow for detection and isolation of the protein. Examples of suitable enzyme labels include, but are not limited to, horseradish peroxidase, alkaline phosphatase, beta-galactosidase and acetylcholinesterase. Examples of suitable prosthetic group complexes include, but are not limited to, streptavidin / biotin and avidin / biotin. Examples of suitable fluorescent materials include, but are not limited to, umbelliferone, fluorescein, fluorescein isothiocyanate, rhodamine, dichlorotriazinylamine fluorescein, dansyl chloride and phycoerythrin. Examples of luminescent materials include luminol, and bioluminescent materials such as luciferase, luciferin and aequorin. Examples of suitable radioactive materials include iodine, carbon, sulfur, tritium, indium, technetium, thallium, gallium, palladium, molybdenum, xenon and fluorine.

[0210] Additional examples of post-translational modifications include acylation, ADP-ribosylation, amidation, covalent attachment of flavin, covalent attachment of a heme moiety, covalent attachment of a nucleotide or nucleotide derivative, covalent attachment of a lipid or lipid derivative, covalent attachment of phosphotidylinositol, cross-linking, cyclization, disulfide bond formation, demethylation, formation of covalent cross-links, formation of cysteine, formation of pyroglutamate, gamma-carboxylation, GPI anchor formation, hydroxylation, iodination, methylation, myristylation, pegylation, prenylation, racemization, selenoylation, sulfation, transfer-RNA mediated addition of amino acids to proteins such as arginylation, and ubiquitination.63IPTS / 200238736.2POLYNUCLEOTIDES, VECTORS AND HOST CELLS

[0211] Certain embodiments of the present disclosure relate to an isolated polynucleotide or a set of polynucleotides encoding an anti-IL-31 antibody construct or multispecific antibody construct described herein.

[0212] The terms “nucleic acid,” “nucleic acid molecule” and “polynucleotide” are used interchangeably herein and refer to a polymeric form of nucleotides of any length, either deoxyribonucleotides or ribonucleotides, or analogues thereof. Non-limiting examples of polynucleotides include a gene, a gene fragment, messenger RNA (mRNA), cDNA, recombinant polynucleotides, plasmids, vectors, isolated DNA, isolated RNA, nucleic acid probes, and primers.

[0213] A polynucleotide that “encodes” a given polypeptide is a polynucleotide that is transcribed (in the case of DNA) and translated (in the case of mRNA) into a polypeptide in vivo when placed under the control of appropriate regulatory sequences. The boundaries of the coding sequence are determined by a start codon at the 5' (amino) terminus and a translation stop codon at the 3' (carboxy) terminus. A transcription termination sequence may be located 3' to the coding sequence.

[0214] Certain embodiments of the present disclosure relate to vectors (such as expression vectors) comprising one or more polynucleotides encoding an anti-IL-31 antibody construct or a multispecific antibody construct as described herein. The polynucleotide(s) may be comprised by a single vector or by more than one vector. In some embodiments, the polynucleotides are comprised by a multicistronic vector.

[0215] Certain embodiments of the present disclosure relate to host cells comprising polynucleotide(s) encoding an anti-IL-31 antibody construct or a multispecific antibody construct as described herein or one or more vectors comprising the polynucleotide (s). In some embodiments, the host cell is eukaryotic, for example, a Chinese Hamster Ovary (CHO) cell, a human embryonic kidney (HEK) cell or a lymphoid cell (e.g. Y0, NS0, Sp20 cell).64IPTS / 200238736.2PHARMACEUTICAL COMPOSITIONS

[0216] For therapeutic use, the anti-IL-31 antibody construct or multispecific antibody construct may be provided in the form of pharmaceutical compositions comprising the antibody construct and a pharmaceutically acceptable carrier or diluent. The compositions may be prepared by known procedures using well-known and readily available ingredients.

[0217] Pharmaceutical compositions may be formulated for administration to a subject by, for example, parenteral, oral (including, for example, buccal or sublingual), topical, rectal or vaginal routes, or by inhalation or spray. “Parenteral” administration may be subcutaneous injection, or intradermal, intra-articular, intravenous, intramuscular, intravascular, intrastemal or intrathecal injection or infusion. The pharmaceutical composition will typically be formulated in a format suitable for administration to the subject, for example, as a syrup, elixir, tablet, troche, lozenge, hard or soft capsule, pill, suppository, oily or aqueous suspension, dispersible powder or granule, emulsion, injectable or solution. Pharmaceutical compositions may be provided as unit dosage formulations.

[0218] In certain embodiments, pharmaceutical compositions comprising the antibody constructs may be formulated for parenteral administration by infusion or in a unit dosage injectable form, for example as lyophilized formulations or aqueous solutions.

[0219] Pharmaceutically acceptable carriers are generally nontoxic to recipients at the dosages and concentrations employed. Examples of such carriers include, but are not limited to, buffers such as phosphate, citrate, and other organic acids; antioxidants such as ascorbic acid and methionine; preservatives such as octadecyldimethylbenzyl ammonium chloride, hexamethonium chloride, benzalkonium chloride, benzethonium chloride, phenol, butyl alcohol, benzyl alcohol, alkyl parabens (such as methyl or propyl paraben), catechol, resorcinol, cyclohexanol, 3 -pentanol and m-cresol; low molecular weight (less than about 10 residues) polypeptides; proteins such as serum albumin or gelatin; hydrophilic polymers such as polyvinylpyrrolidone; amino acids such as glycine, glutamine, asparagine, histidine, arginine or lysine; monosaccharides, disaccharides, and other carbohydrates such as glucose, mannose or dextrins; chelating agents such as EDTA; sugars such as sucrose, mannitol, trehalose or sorbitol; salt-forming counter-ions such as sodium;65IPTS / 200238736.2metal complexes such as Zn-protein complexes, and non-ionic surfactants such as polyethylene glycol (PEG).

[0220] In certain embodiments, pharmaceutical compositions comprising the antibody constructs may be in the form of a sterile injectable aqueous or oleaginous solution or suspension. Such suspensions may be formulated using suitable dispersing or wetting agents and / or suspending agents that are known in the art. The sterile injectable solution or suspension may comprise the antibody construct in a non-toxic parentally acceptable diluent or solvent. Acceptable diluents and solvents that may be employed include, for example, 1,3-butanediol, water, Ringer’s solution or isotonic sodium chloride solution. In addition, sterile, fixed oils may be employed as a solvent or suspending medium. For this purpose, various bland fixed oils may be employed, including synthetic mono- or diglycerides. In addition, fatty acids such as oleic acid find use in the preparation of injectables. Adjuvants such as local anesthetics, preservatives and / or buffering agents may also be included in the injectable solution or suspension.

[0221] In certain embodiments, pharmaceutical compositions comprising the antibody constructs may be formulated for parenteral administration to a subject, for example a human. Typically, compositions for parenteral administration are solutions in sterile isotonic aqueous buffer. Where necessary, the composition may also include a solubilizing agent and / or a local anaesthetic such as lignocaine to ease pain at the site of the injection. Generally, the ingredients are supplied either separately or mixed together in unit dosage form, for example, as a dry lyophilized powder or water-free concentrate in a hermetically sealed container such as an ampoule or sachet indicating the quantity of active agent. Where the composition is to be administered by infusion, it can be dispensed with an infusion bottle containing sterile pharmaceutical grade water or saline. Where the composition is administered by injection, an ampoule of sterile water for injection or saline can be provided so that the ingredients may be mixed prior to administration.

[0222] Other pharmaceutical compositions and methods of preparing pharmaceutical compositions are known in the art and are described, for example, in “Remington: The Science and Practice of Pharmacy" (formerly “Remingtons Pharmaceutical Sciences”) Gennaro, A., Lippincott, Williams & Wilkins, Philadelphia, PA (2000).66IPTS / 200238736.2METHODS OF USE

[0223] Certain aspects of the present disclosure relate to the therapeutic use of the anti-IL-31 antibody constructs and multispecific antibody constructs described herein.

[0224] Certain embodiments relate to use of the anti -IL-31 antibody constructs and multispecific antibody constructs in the treatment of an allergic, inflammatory or autoimmune disease. Some embodiments relate to methods of treating a subject having an allergic, inflammatory or autoimmune disease comprising administering an effective amount of the anti-IL-31 antibody construct or multispecific antibody construct to the subject.

[0225] Examples of allergic, inflammatory or autoimmune diseases that may be treated with the anti-IL-31 antibody constructs or multispecific antibody constructs in certain embodiments include, but are not limited to, atopic dermatitis, dermatomycosis, eczema, psoriasis, urticaria, lupus, scleroderma, Sjogren’s syndrome, vitiligo, celiac disease, acute allergic contact dermatitis, chronic spontaneous urticaria, bullous pemphigoid, alopecia areata, dermatomyositis, prurigo nodularis, allergic rhinitis, inflammatory bowel disease, allergic asthma and atopic asthma.

[0226] The terms “treat” and “treatment” and grammatical variations thereof as used herein, refer to an intervention performed with the intention of alleviating the symptoms associated with, or altering the pathology of, a disease, disorder or condition. Thus, the terms include in various embodiments one or more of alleviation, moderation, reduction or curing of a disease, disorder or condition.

[0227] Certain embodiments of the present disclosure relate to use of the anti-IL-31 antibody constructs and multispecific antibody constructs to alleviate pruritis. Some embodiments relate to methods of treating a subject suffering from pruritis comprising administering an effective amount of the anti-IL-31 antibody construct or multispecific antibody construct to the subject. Some embodiments relate to methods of alleviating pruritis in a subject comprising administering an effective amount of the anti-IL-31 antibody construct or multispecific antibody construct to the subject. Pruritis (or itching) is a common symptom of a number of diseases and conditions, for example, atopic dermatitis, dermatomycosis, eczema, psoriasis and urticaria. In certain67IPTS / 200238736.2embodiments, the anti-IL-31 antibody construct or multispecific antibody construct may be used to treat pruritis associated with atopic dermatitis, dermatomycosis, eczema, psoriasis or urticaria.

[0228] Certain embodiments of the present disclosure relate to use of the anti-IL-31 antibody constructs and multispecific antibody constructs to treat an allergic, inflammatory or autoimmune disease that causes pruritis. Some embodiments relate to use of the anti-IL-31 antibody constructs and multispecific antibody constructs to treat atopic dermatitis, dermatomycosis, eczema, psoriasis, urticaria, acute allergic contact dermatitis, chronic spontaneous urticaria and prurigo nodularis.

[0229] The dosage of the anti -IL-31 antibody construct or multispecific antibody construct to be administered is not subject to defined limits, but it will be a therapeutically effective amount. A “therapeutically effective amount” refers to that amount of an antibody construct described herein which, when administered to a subject, is sufficient to effect a treatment of the particular indication.PHARMACEUTICAL KITS

[0230] Certain embodiments relate to pharmaceutical kits (or articles of manufacture) comprising an anti -IL-31 antibody construct or a multispecific antibody construct as described herein.

[0231] The kit typically will comprise a container holding the antibody construct and a label and / or package insert on or associated with the container. The label or package insert contains instructions customarily included in commercial packages of therapeutic products, providing information about the indications, usage, dosage, administration, contraindications and / or warnings concerning the use of such therapeutic products. The label or package insert may further include a notice in the form prescribed by a governmental agency regulating the manufacture, use or sale of pharmaceuticals or biological products, which notice reflects approval by the agency of manufacture, use or sale for human or animal administration. In some embodiments, the container may have a sterile access port. For example, the container may be an intravenous solution bag or a vial having a stopper that may be pierced by a hypodermic injection needle.68IPTS / 200238736.2

[0232] In addition to the container holding the antibody construct, the kit may optionally comprise one or more additional containers comprising other components of the kit. For example, a pharmaceutically acceptable buffer (such as bacteriostatic water for injection (BWFI), phosphate-buffered saline, Ringer’s solution or dextrose solution), other buffers or diluents.

[0233] Suitable containers include, for example, bottles, vials, syringes, intravenous solution bags, and the like. The containers may be formed from a variety of materials such as glass or plastic. If appropriate, one or more components of the kit may be lyophilized or provided in a dry form, such as a powder or granules, and the kit can additionally contain a suitable solvent for reconstitution of the lyophilized or dried component(s).

[0234] The kit may further include other materials desirable from a commercial or user standpoint, such as filters, needles, and syringes.EXAMPLES

[0235] The practice of the present disclosure will employ, unless otherwise indicated, conventional methods of protein chemistry, biochemistry, recombinant DNA techniques and pharmacology, within the skill of the art. Such techniques are explained fully in the literature. See, e.g., T. E. Creighton, Proteins: Structures and Molecular Properties (W. H. Freeman and Company, 1993); A. L. Lehninger, Biochemistry (Worth Publishers, Inc., current addition); Sambrook, et al., Molecular Cloning: A Laboratory Manual (2nd Edition, 1989); Methods In Enzymology (S. Colowick and N. Kaplan eds., Academic Press, Inc.); Remington 's Pharmaceutical Sciences, 18th Edition (Easton, Pennsylvania: Mack Publishing Company, 1990).EXAMPLE 1: PREPARATION AND CHARACTERIZATION OF HUMANIZED AND AFFINITY-MATURED ANTI-IL-31 ANTIBODIES1.1 Antibody Generation

[0236] Antibodies that specifically bind IL-31 were generated by immunizing rabbits with human IL-31 as described below.

[0237] Two New Zealand White rabbits were subcutaneously immunized with recombinant human IL-31 antigen (Sino Biological, Beijing, China; Cat. 11557-H08H) over 6369IPTS / 200238736.2days, after which blood was drawn. Anti-human IL-31 antibody titers were determined by flow cytometry using streptavidin beads (Spherotech, Lake Forest, IL) coated with IL-31 antigen (R& D Systems, Minneapolis, MN). Test bleed sera mounted a significant response again human IL-31.

[0238] Immunized rabbits were sacrificed and the spleens harvested. Splenocytes for each rabbit were used for B cell enrichment and sorted on a FACSAria™ Cell Sorter (Becton, Dickinson & Co., Franklin Lakes, NJ) into wells containing lysis buffer with a modified protocol based on the Selected Lymphocyte Antibody Method (SLAM) (Babcook et al., 1996, Proc Natl Acad Set USA, 93(15):7843-7848).

[0239] Total RNA from wells containing a single B cell was used as template with SuperScript™ III (Thermo Fisher Scientific Corp., Waltham, MA) and oligo-dT20 (Integrated DNA Technologies, Inc., Coralville, IA) to transcribe cDNA from mRNA. Initial PCR of heavy and light chain antibody-coding sequences was performed using primers and methods modified from Babcook etal., \996, ProcNatlAcadSci USA, 93(15):7843-7848, von Boehmer etal., 2016, NatProtoc., 11(10): 1908, and Peng etal., 2017, J. Mol. Biol, 429(19):2954-2973, with cDNA as the nucleic acid template. A subsequent PCR reaction was then performed on these unique sequences using V-segment family and J-segment family-specific primers and the resulting amplicons were cloned into pTT5-based expression plasmids (National Research Council of Canada). Unique heavy chain sequences and light chain sequences emerging from a single well sample were co-expressed in Expi293F cells (Thermo Fisher Scientific, Waltham, MA; Cat# A14527).

[0240] Cell supernatants containing secreted antibodies were assessed for blocking human IL-31Ra receptor in a receptor ligand assay. The wells corresponding to human IL-31 antibody containing supernatant were selected for sequencing.

[0241] Heavy and light chain PCR amplicons were sequenced using NGS-based Amplicon-EZ and analyzed for unique antibody-coding sequences. The VH and VL sequences identified for the rabbit anti -human IL-31 antibody 40A12 are shown in Table 1.1.70IPTS / 200238736.2Table 1.1: Rabbit VH and VL Sequences for Anti-Human IL-31 Antibody 40A12 and Chimeric Antibody Construct, v33559Description Sequence SEQ ID NORabbit VH QSLEESGGRLVTPGGSLTLTCTVSGIDLSSYFMSWVRQAPGK 119 GLEYIGTISTGGNTYYASWVKGRFTISKTSTTVDLKITSPTTED TATYFCARGWLRDYLDLWGQGTLVTIS SRabbit VL QVLTQTPSPVSAAVGGTVSISCQASQSVYRENRLAWYQQKV 120 (kappa) GQPPKLLIYRASKLESGVPSRFSGSGSGTEFTLTISGVQCDDAATYYCAGGDSSGSDHAFGGGTEVVVK

[0242] These rabbit VH and VL sequences were used to prepare a rabbit-human chimeric IgGl / kappa antibody construct, v33559, as follows. Coding sequences for antibody variable regions were cloned in frame into a human IgGl expression vector or a human C kappa expression vector (based on the pTT5 vector). The human IgGl constant region starts at alanine Kabat-114, and human C kappa constant region starts at arginine Kabat-108. The activities of the resultant recombinant chimeric antibody construct were confirmed in specificity binding assays.1.2 Humanization

[0243] The rabbit VH and VL sequences from chimeric antibody construct v33559 were aligned against human immunoglobulin germline sequences to select basis germline sequences for humanization. Human germline IGHV3-15*01 with IGHJ4*01 was selected for VH humanization. Human germline IGKVl-5*01 with IGKJ4*01 was selected for VL humanization. The CDR sequences by AbM definition for CDRH1, CDRH2 and IMGT definition for CDRH3 from v33559 were swapped into the selected human germline frameworks to create a basis humanized construct. Several areas of the basis construct were identified for back mutation or deletion to the original rabbit parental sequence to minimize potential disruption to antigen binding.

[0244] Humanization was performed in two rounds with eight candidate humanized VH sequences, and five candidate humanized VL sequences created in Round 1 and twenty-one additional candidate humanized VH sequences created in Round 2. Seven constructs represented by 6 humanized VH and 2 humanized VL sequences from Round 1 were expressed in Expi293F cells and purified by Protein A affinity chromatography using MabSelect™ SuRe™ resin (GE71IPTS / 200238736.2Healthcare) following the protocol described in Section 1.4. Binding to IL-31 was assessed by KinExA™ (Sapidyne Inc., Boise, ID) for a representative variant following the protocol described in Section 1.5 and the purified variants were also screened for activity using a Ba / F3 assay (as described in Example 2). None of the Round 1 humanized variants showed activity comparable to the parental rabbit chimeric construct.

[0245] The twenty-one candidate humanized VH sequences created in Round 2 were combined with one VL sequence from Round 1 (LI) and the resulting 21 human IgGl-based monoclonal antibodies were expressed and purified following the protocol described in Section 1.4. Binding to IL-31 was assessed by KinExA™ following the protocol described in Section 1.5 and the purified variants were screened for activity using a Ba / F3 assay (see Example 2). Nine variants represented by 9 humanized VH and 1 humanized VL sequences (see Tables 1.2 and 1.3, and Fig. 16 (Table Bl)), which demonstrated activity similar to the parental rabbit chimeric construct, were selected for further analysis. The CDR sequences of the parental rabbit chimeric construct and the nine humanized variants are shown in Fig. 14 (Table Al).Table 1.2: Round 2 Humanized Anti-IL-31 Antibody Variant VH and VL Domain Sequences VH / VL Sequence SEQ ID NOHll QSLVESGGGLVKPGGSLRLSCAVSGIDLSSYFMSWVRQAPGKGLEYI 47 GTISTGGNTYYASWVKGRFTISKDSSKNTVYLQMNSLKTEDTAVYYC ARGWLRDYLDLWGQGTLVTVSSH14 QSLVESGGGLVKPGGSLRLSCAVSGIDLSSYFMSWVRQAPGKGLEYI 48 GTISTGGNTYYASWVKGRFTISKTSTTVYLQMNSLKTEDTAVYYCAR GWLRDYLDLWGQGTLVTVS SH15 QSLVESGGGLVKPGGSLRLSCAVSGIDLSSYFMSWVRQAPGKGLEYI 49 GTISTGGNTYYASWVKGRFTISKTSTTVYLQINSLKTEDTAVYYCARG WLRDYLDLWGQGTLVTVSSH16 QSLVESGGGLVKPGGSLRLSCAVSGIDLSSYFMSWVRQAPGKGLEYI 50 GTISTGGNTYYASWVKGRFTISKTSTTVYLQINSPKTEDTAVYYCARG WLRDYLDLWGQGTLVTIS SH17 QSLVESGGGLVKPGGSLRLSCAVSGIDLSSYFMSWVRQAPGKGLEYI 51 GTISTGGNTYYASWVKGRFTISKTSTTVYLQINSPKTEDATYFCARGW LRDYLDLWGQGTLVTIS S72IPTS / 200238736.2VH / VL Sequence SEQ ID NOH18 QSLVESGGGLVKPGGSLRLSCAVSGIDLSSYFMSWVRQAPGKGLEYI 52 GTISTGGNTYYASWVKGRFTISKTSTDVYLQMNSLKTEDTAVYYCAR GWLRDYLDLWGQGTLVTVS SH19 QSLVESGGGLVKPGGSLRLSCAVSGIDLSSYFMSWVRQAPGKGLEYI 53 GTISTGGNTYYASWVKGRFTISKTSTGVYLQMNSLKTEDTAVYYCAR GWLRDYLDLWGQGTLVTVS SH20 QSLVESGGGLVKPGGSLRLSCAVSGIDLSSYFMSWVRQAPGKGLEYI 54 GTISTGGNTYYASWVKGRFGISKTSTTVYLQMNSLKTETAVYYCARG WLRDYLDLWGQGTLVTVSSH21 QSLVESGGGLVKPGGSLRLSCAVSGIDLSSYFMSWVRQAPGKGLEYI 55 GTISTGGNTYYASWVKGRFDISKTSTTVYLQMNSLKTEDTAVYYCAR GWLRDYLDLWGQGTLVTVS S LI DIQMTQSPSTLSASVGDRVTITCRASQSVYRENRLAWYQQKPGKAPK 56 LLIYRASKLESGVPSRFSGSGSGTEFTLTISSLQPDDFATYYCAGGDSS GSDHAFGGGTKVEIKTable 1.3: VH and VL Composition of Selected Round 2 Humanized Anti-IL-31 Antibody VariantsVariant VH VLv34090* chimeric chimericv34731 Hll LIv34734 H14 LIv34735 H15 LIv34736 H16 LIv34737 H17 LIv34738 H18 LIv34739 H19 LIv34740 H20 LIv34741 H21 LI* Identical to chimeric variant v33559 but incudes the CH2 amino acid substitutions: L234A, L235A and D265S

[0246] The nine selected humanized variants (v34731, v34734, v34735, v34736, v34737, v34738, v34739, v34740, v34741) plus the parental rabbit chimeric construct (v34090) were 73IPTS / 200238736.2expressed in 50 mL Expi293F cultures and purified by Protein A following the protocol described in Section 1.4, and assessed for a dose response in the Ba / F3 assay using both human and cynomolgus IL-31 orthologues as described in Example 2. All humanized variants showed activity with similar or higher potency than the parental rabbit chimeric construct and the benchmark antibody (BMS-981164) with equivalent species cross-reactivity (see Example 2). Binding to IL-31 was assessed by KinExA™ for two representative constructs following the protocol described in Section 1.5 and the binding affinity (KD) was determined to be similar to the benchmark antibody (BMS-981164) (see Table 1.4).Table 1.4: Binding Affinity of Humanized Anti-IL-31 Antibody Variants v34731 and v34734 to Human IL-31Variant Affinity (KD) 1 (pM)v34731 1.9v34734 1.3v30675 (BMS-981164) 2.31.3 Affinity Maturation

[0247] Four VH sequences (H8 from round 1, and Hl 1, H14 and H18 from round 2) and one VL sequence (LI from round 1) were affinity matured using the HuTarg™ system (Innovative Targeting Solutions, Vancouver, BC, Canada) resulting in 6 affinity matured monoclonal antibody variants.

[0248] Briefly, genetic recombination was applied to the variable regions of the humanized VH and VL regions, and high affinity mutants were identified using next-generation sequencing (NGS). The CDR loops of the humanized variable domains were interspersed with RAG1 / 2 recombination signal sequences (RSS) and cloned into plasmid E951 (Innovative Targeting Solutions, Vancouver, BC, Canada). E951-based plasmid pools were integrated into HuTarg™ cells, and RAG1 / 2 expression induced for 48 hours. HuTarg™ cells displaying successfully recombined antibodies, shown after staining with PE-conjugated goat anti-human kappa light chain antibody (Bio-Rad Laboratories, Hercules, CA; cat#206009) were then selected and further subjected to multiple rounds of FACS-based sorting on a BD FACSAria™ Flow Cytometer (BD 74IPTS / 200238736.2Biosciences, Franklin Lakes, NJ), with each round using a reduced amount of biotinylated soluble IL-31 antigen (R& D Systems, Minneapolis, MN; Cat. No. 2824-IL / CF). Detection employed streptavidin conjugated to AlexaFluor-647 (Thermo Fisher Scientific Corp., Waltham, MA; cat# S 11223). HuTarg™ cells that exhibited increased binding to IL-31 antigen were sorted directly to RNAzol™ RT (Sigma- Aldrich, St. Louis, MI; cat# R4533) and total RNA from cells lysed in RNAzol™ was isolated as per manufacturer’s instructions.

[0249] RNA was then digested with ezDNase™ (Thermo Fisher Scientific Corp., Waltham, MA; cat# 11766051), and cDNA transcribed using Superscript™ IV (Thermo Fisher Scientific Corp., Waltham, MA; cat# 18090010) and a gene-specific primer. VH and VL domains were targeted for PCR amplification, and molecularly barcoded with NEBNext® Ultra™ DNA Library Prep Kit (New England Biolabs, Ipswich, MA; cat# E7370L). Samples were pooled and run on an Illumina MiSeq™ sequencer with a 500-cycle kit using v2 chemistry (Illumina, San Diego, CA; cat# MS-102-2003). Sequence analysis was performed to identify mutations within VH and VL sequences that exhibited a high likelihood of conferring an increase in affinity. DNA sequences encoding mutated VH and VL domains were synthesized as “MiniGenes” (Integrated Technologies, Inc., Coralville, IA) and cloned into expression vectors to provide expression plasmids coding for complete human IgGl heavy chains and human kappa light chains, respectively. Expression plasmids were matrixed with one another to pair every heavy chain plasmid with every light chain plasmid. This matrix was recombinantly expressed in ExpiCHO™ cells (Thermo Fisher Scientific Corp., Waltham, MA) according to manufacturer’s instructions and the supernatants were assessed by single point Ba / F3 assay with both human and cynomolgus IL-31 as described in Example 2. Seven variants represented by 5 affinity matured VH sequences and 4 affinity matured VL sequences (see Tables 1.5 and 1.6, and Fig. 17 (Table B2)) were expressed in 100 mL ExpiCHO™ cultures as described in Section 1.4 and the purified samples were assessed in a dose response Ba / F3 assay using both human and cynomolgus IL-31 as described in Example 2. The CDR sequences of the seven affinity matured variants are shown in Fig. 15 (Table A2).75IPTS / 200238736.2Table 1.5: Affinity Matured Anti-IL-31 Antibody VH and VL Domain Sequences NVINY ID Sequence SEQ ID NOH8-mut07 EVQLVESGGGLVKPGGSLRLSCAVVGIDLSSYFMSWVRQAPG 57 KGLEYIGTISTGGNTYYAAPVKGRFTISKTSTTVYLQINSPKTE DTAVYYCARGWLRDYLDRWGQGTLVTVSSHll-mut07 EVQLVESGGGLVKPGGSLRLSCAVVGIDLSSYFMSWVRQAPG 58 KGLEYIGTISTGGNTYYASWVKGRFTISKDSSKNTVYLQMNSL KTEDTAVYYCARGWLRDYLDRWGQGTLVTVS SHl l-mut05 EVQLVESGGGLVKPGGSLRLSCAVSGIDLSSYFMSWVRQAPG 59 KGLEYIGTISTGGNTYYASWVKGRFTISKDSSKNTVYLQMNSL KTEDTAVYYCARGWLRDYLDRWGQGTLVTVS SH14-mut05 EVQLVESGGGLVKPGGSLRLSCAVSGIDLSSYFMSWVRQAPG 60 KGLEYIGTISTGGNTYYASWVKGRFTISKTSTTVYLQMNSLKT EDTAVYYCARGWLRDYLDRWGQGTLVTVSSH18-mut05 EVQLVESGGGLVKPGGSLRLSCAVSGIDLSSYFMSWVRQAPG 61 KGLEYIGTISTGGNTYYASWVKGRFTISKTSTDVYLQMNSLKT EDTAVYYCARGWLRDYLDRWGQGTLVTVSSLl-mutl 1 DIQMTQSPSTLSASVGDRVTITCRSRQSVYRENRLAWYQQKPG 62 KAPKLLIYRASKLEKGVASRFSGSGSGTEFTLTISSLQPDDFAT YYCAGGASSGSDHAFGGGTKVEIKLl-mut06 DIQMTQSPSTLSASVGDRVTITCRSRQSVYRENRLAWYQQKPG 63 KAPKLLIYRASKLEKGVASRFSGSGSGTEFTLTISSLQPDDFAT YYCAGGDSSGSDHAFGGGTKVEIKLl-mutOl DIQMTQSPSTLSASVGDRVTITCRSRQSVYRENRLAWYQQKPG 64 KAPKLLIYRASKLESGVPSRFSGSGSGTEFTLTISSLQPDDFATY YCAGGDSSGSDHAFGGGTKVEIKLl-mut05 DIQMTQSPSTLSASVGDRVTITCRSRQSVYRENRLAWYQQKPG 65 KAPKLLIYRGSKLESGVPSRFSGSGSGTEFTLTISSLQPDDFATY YCAGGDSSGSDHAFGGGTKVEIKTable 1.6: VH and VL Composition of Selected Affinity Matured Anti-IL-31 Antibody VariantsVariant VH VLv36535 H8-mut07 Ll-mutl 1v36539 Hll-mut07 Ll-mutl 176IPTS / 200238736.2Variant VHVLv36540 Hll-mut07 Ll-mut06v36541 Hll-mut05 Ll-mutOlv36542 Hll-mut05 Ll-mut05v36545 H14-mut05 Ll-mutOlv36974 H18-mut05 Ll-mut051.4 Production and Characterization of Affinity Matured Anti-IL-31 Antibody Variants

[0250] Each of the 7 affinity matured antibody variants was produced in full-size antibody (FSA) format containing two identical full length heavy chains and two identical kappa light chains.

[0251] The full-length heavy chain contained the human CHl-hinge-CH2-CH3 domain sequence of IGHGl*01 (SEQ ID NO: 115; see Table 1.7) in which the following CH2 amino acid substitutions had been made: L234A, L235A and D265S (amino acid residues in the Fc region are identified according to the EU index). These mutations “knock out” the ability of the Fc region to bind to the FcyRs. IgGl Fc regions comprising these mutations are referred to herein as “IgGl FcKO” and have the sequence set forth in SEQ ID NO: 116 (Table 1.7). The light chain contained human kappa CL sequence of IGKC*01 (SEQ ID NO: 117; see Table 1.7).Table 1.7: Sequences of Wild-Type IgGl Heavy Chain, FcKO Heavy Chain and Kappa Light ChainName Sequence SEQ ID NO CHl-hinge- ASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALT 115 CH2-CH3 SGVHTFPAVLQSSGLYSLSSWTVPSSSLGTQTYICNVNHKPSNTKV (IGHGl*01) DKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEV TCVWDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRW SVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVY TLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPP VLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSL SLSPG CHl-hinge- ASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALT 116 CH2-CH3 SGVHTFPAVLQSSGLYSLSSWTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPEAAGGPSVFLFPPKPKDTLMISRTPE77IPTS / 200238736.2Name Sequence SEQ ID NO(IgGl VTCVWSVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRV FcKO)1VSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQV YTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTT PPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQK SLSLSPG CL (kappa) RTVAAPSVFIFPPSDEQLKSGTASWCLLNNFYPREAKVQWKVDNA 117 (IGKC*01) LQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC1L234A, L235A and D265S mutations marked in bold and underline

[0252] Each of the affinity matured VH domain sequences was appended to a human CH1-hinge-CH2-CH3 domain sequence of IGHG1* and each of the affinity matured VL domain sequences was appended to the human kappa CL sequence of IGKC*01. All sequences were reverse translated to DNA, codon optimized for mammalian expression and gene synthesized.

[0253] Heavy chain vector inserts comprising a signal peptide (artificially designed sequence: MRPTWAWWLFLVLLLALWAPARG [SEQ ID NO: 118] (Barash et al., 2002, Biochem and Biophys Res. Comm., 294:835-842)) and the heavy chain clone terminating at residue G446 (EU numbering) of the CH3 domain were ligated into a pTT5 vector to produce heavy chain expression vectors. Light chain vector inserts comprising the same signal peptide were ligated into a pTT5 vector to produce light chain expression vectors. The resulting heavy and light chain expression vectors were sequenced to confirm correct reading frame and sequence of the coding DNA.

[0254] Gene products were cloned (by GenScriptBioTech Corporation, Piscataway, NJ) into the pTT5 mammalian expression vector and transiently expressed in ExpiCHO™ cells (ThermoFisher Scientific) using ExpiFectamine™ CHO Transfection Kit (ThermoFisher Scientific, Cat No. A29129). Transfected cells were cultured in ExpiCHO™ expression medium (ThermoFisher Scientific, Cat No. A2910001) according to the “ExpiCHO™ Expression System User Guide” (ThermoFisher Scientific, 2018) for the ExpiCHO™ Max Titer protocol. Cells were harvested 7 days post transfection, centrifuged and the supernatant filtered on a 0.22 pm Millipore filter.78IPTS / 200238736.2

[0255] The clarified culture medium was loaded onto MabSelect™ SuRe™ Protein A resin (GE Healthcare) in a gravity column and washed with 10 column volumes of PBS buffer at pH 7.2 - 7.4. The antibody variant was eluted with 10 column volumes of 0.1 M citrate buffer at pH 3.6 and neutralized with 1 M TRIS at pH 9. The amount of antibody variant was then quantified based on A280 nm (NanoDrop™ Spectrophotometer; Thermo Fisher Scientific).

[0256] Purity of samples was assessed by electrophoresis under non-reducing and reducing conditions using the High Throughput Protein Express assay and Caliper LabChip™ GXII or GXII Touch HT (Perkin Elmer, Waltham, MA). Procedures were carried out according to HT Protein Express LabChip™ User Guide version 2 with the following modifications. Antibody samples, at either 2 pl or 5 pl (concentration range 5-2000 ng / pl), were added to separate wells in 96 well plates (BioRad, Hercules, CA; Cat No. HSP9601B) along with 7pl ofHT Protein Express Sample Buffer (Perkin Elmer; Cat No. 760328). Antibody samples were then denatured at 70°C for 15 mins. The LabChip™ instrument was operated using the HT Protein Express Chip (Perkin Elmer, Waltham, MA) and the Ab-200 assay setting.

[0257] Species homogeneity of the affinity matured antibody variants and parental rabbithuman chimeric antibody variant samples was assessed by UPLC-SEC. UPLC-SEC was performed on Agilent Technologies 1290 Infinity LC System with a diode array detector (DAD) using the Agilent Technologies AdvanceBio SEC300A column (7.8 x 150 mm, 2.7 pm) at 25 °C. The mobile phase was 200 mM KPOr + 200 mM KC1, pH 7.4 and the flow rate was 1 mL / min. Total run time for each injection was 7 min with atotal injection of lOug of protein sample. Elution was monitored by UV absorbance in the range 190-400 nm and chromatograms were extracted at 280 nm. Peak integration was performed using OpenLAB CDS ChemStation software (Agilent Technologies, Inc., Santa Clara, CA).

[0258] The antibody variants were further purified by gel filtration chromatography using a Superdex™ 200 Increase column (GE Healthcare) via an AKTA Pure™ chromatography system at a flowrate of 1 mL / min. PBS buffer at pH 7.4 was used at a flow-rate of 1 mL / min. Fractions of eluted antibody construct were collected based on absorbance at A280 nm and the fractions were assessed by non-reducing and reducing High Throughput Protein Express assay using Caliper LabChip™ GXII and UPLC-SEC using Agilent Technologies 1290 Infinity LC System with DAD79IPTS / 200238736.2and the Agilent Technologies AdvanceBio SEC300A column (7.8 x 150 mm, 2.7 um) at 25 °C. Fractions with high purity were pooled and endotoxin levels were determined by the limulus amebocyte lysate (LAL) assay using the Endosafe™ Portable Test System (PTS) (Charles River, Wilmington, MA).

[0259] The yields of all 7 affinity matured antibody variants ranged from 8-43 mg (from 120 mL or 125 mL culture) and all 7 were obtained at >96% purity (see Table 1.8).Table 1.8: Post Purification Yield and Final Purity (%) of Affinity Matured Anti-IL-31 Antibody VariantsVolume Titer % Purity by Variant Yield (mg)(mL) (mg / L) UPLC-SEC v36535 120 233 25 96v36539 120 485 17 98v36540 120 317 8 96v36541 120 921 38 100v36542 120 916 43 100v36545 120 587 35 100v36974 125 927 40.5 1001.5 Characterization of Anti-IL-31 Antibody Variants1.5.1 Binding of Anti-IL-31 Antibody Variants to Human IL-31

[0260] The equilibrium binding affinity of the affinity matured antibody variants to human IL-31 was determined by KinExA™ (Sapidyne Inc., Boise, ID). Briefly, human IL-31 (R& D Systems, Minneapolis, MN; Cat No. 2824-IL / CF) was immobilized on azlactone beads. The antibody variants were used as the constant binding partner (CBP) and antigen (human IL-31) was used as the titrant. The antibody variants were tested at constant concentrations of 10 pM and 100 pM or 5 pM and 50 pM. The titrant, human IL-31, was titrated by 2-fold serial dilutions of 0.5 nM. The antibody / antigen mixture was incubated at 25 °C for 4 days and assessed using goat antihuman IgG (H+L) conjugated with Alexa Fluor™ 647 for detection, with data analyzed using the KinExA™ software.80IPTS / 200238736.21.5.2 Thermal Stability Assessment of Anti-IL-31 Antibody Variants

[0261] Thermal stability of the anti -IL-31 antibody variants was assessed by measuring changes in hydrodynamic radius with increasing temperature using Dynamic Light Scattering (DLS). The aggregation temperature (Tagg) was measured for each antibody variant diluted at 1 mg / mL concentration in either PBS pH 7.4 buffer or 20 mM histidine pH 6.0 buffer using a DLS plate reader (Wyatt Technology Corporation, Goleta, CA). Samples were continuously heated at a ramping rate of ~ 0.1 °C / min. Four DLS acquisitions were made for 5 seconds each to determine the hydrodynamic radius using DYNAMICS™ software (Wyatt Technology). The Taggvalues indicate that all humanized antibody variants assessed have higher thermal stability than the parental chimeric antibody (v34090) and the anti-IL-31 antibody control (BMS-981164) (see Table 1.9). The results also suggest that all tested variants have a higher stability in 20 mM histidine pH 6.0 buffer than in PBS pH 7.4 buffer.Table 1.9: TaggValues for Select Humanized and Affinity Matured Anti-IL-31 Antibody VariantsTagg(°C)Variant PBS pH 7.4 20 mM Histidine pHBuffer 6.0 Bufferv34090 59.49 60.81v34731 72.24 76.12v34734 72.28 75.78v34735 70.98 75.30v34736 69.99 74.16v34737 70.15 73.86v34738 71.84 75.49v34739 72.27 75.57v34740 71.66 75.96v34741 71.01 75.77v36535 64.80 67.40v36539 69.30 74.20v36540 71.30 74.7081IPTS / 200238736.2Tagg (°C)Variant PBS pH 7.4 20 mM Histidine pHBuffer 6.0 Bufferv36541 71.60 75.50v36542 73.00 75.20v36545 71.20 74.60v36974 69.6 65.43*v30675 (BMS-981164) 68.94 71.91* Sample buffer was 20 mM Histidine + 150 mM NaCl.1.5.3 Accelerated Stability Assessment of Anti-IL-31 Antibody Variants

[0262] Accelerated stability studies were performed to assess long term stability of the anti-IL-31 variants. The antibody variants were stored at 40 °C for 1 month at 1 mg / mL concentration in 20 mM histidine pH6.0 buffer. The purity of the samples post 1 month storage was assessed by HPLC-SEC analysis to determine the changes in % monomer, % high molecular weight species (HMWs) and % low molecular weight species (LMWs). Briefly, 10 pL filtered sample was injected on a Waters™ BEH Acquity UPLC Protein BEH SEC 200 A, 4.6 x 300 mm column (Waters Corporation, Milford, MA) attached to a Waters™ Acquity H-Class UPLC system using 100 mM sodium phosphate, 300 mM sodium chloride (NaCl) pH 6.8 mobile phase at a flow rate of 0.3 mL / min with a 15 minute run time and detection of absorbance at 280 nm. The chromatogram obtained was analyzed to determine the impurities (HMW and LMW) in the sample and compared to no treatment condition (see Table 1.10). Any loss in % monomer observed after 1 month at 40 °C was accompanied by an increased LMWs indicating that the loss was caused by fragmentation of the antibody (see Table 1.10).Table 1.10: Accelerated Stability Assessment of Anti-IL-31 Antibody VariantsNo Treatment 1 Month at 40 °C Variant % % % % % %HMWs Monomer LMWs HMWs Monomer LMWs v36535 0.00 81.76 18.24 0.00 79.34 20.66 v36539 0.15 92.8 7.05 0.00 91.05 8.95 v36540 0.10 94.32 5.58 0.28 92.10 7.6282IPTS / 200238736.2No Treatment 1 Month at 40 °C Variant % % % % % %HMWs Monomer LMWs HMWs Monomer LMWs v36541 0.15 99.12 0.73 0.12 98.31 1.56 v36542 0.07 99.36 0.57 0.04 98.37 1.59 v36545 0.18 99.12 0.70 0.11 98.27 1.62 v36974 0.35 99.29 0.36 0.34 98.00 1.65EXAMPLE 2: FUNCTIONAL CHARACTERIZATION OF ANTLIL-31 ANTIBODY VARIANTS - Ba / F3 ASSAY

[0263] Selected Round 2 humanized anti-IL-31 antibody variants and affinity matured anti -IL-31 antibody variants were assessed for inhibition of IL-31 mediated proliferation in Ba / F3 cells stably expressing human IL-3 IRA and oncostatin M receptor (OSMR) as described below. A bivalent anti-IL-31 antibody (based on BMS-981164) was used as a control.

[0264] Briefly, test articles were serially diluted with a top concentration of 5 nM in assay media, RPMI + 10% heat inactivated fetal bovine serum (ThermoFisher Scientific, Waltham, MA) and plated in a 384-well black flat bottom assay plate. 1 pM human IL- 31 (R& D Systems, Minneapolis, MN) was added. Ba / F3 cells (expressing human IL-3 IRA and OSMR) were washed in assay media to remove any mouse IL-3 remaining from the growth media. 500 Ba / F3 cells were seeded per well and incubated for 3 days at 37°C, 5% CO2. Following incubation, cell proliferation was assessed by CellTiter-Glo® Luminescent Cell Viability Assay (Promega Corporation, Madison, WI) and analyzed on a Synergy™ plate reader (BioTek Instruments, Winooski, VT).

[0265] The results are shown in Tables 2.1, 2.2 and 2.3.83IPTS / 200238736.2Table 2.1: Ba / F3 Assay Potency with Human and Cynomolgus IL-31 for Selected Round 2 Humanized Anti-IL-31 Antibody VariantsIC50Variant Human IL-31 Cynomolgus IL-31(pM) (pM)v3409041 33(chimeric)v34731 25 22v34734 15 19v34735 15 19v34736 21 29v34737 23 22v34738 17 21v34739 30 44v34740 20 20v34741 19 28Control22 24BMS-981164Table 2.2: Ba / F3 Assay Potency with Human and Cynomolgus IL-31 for Humanized Affinity Matured Anti-IL-31 Antibody VariantsIC50(Exp. 1) IC50(Exp.2)* Variant Human IL-31 Cynomolgus IL-31 Human IL-31 Cynomolgus IL-31(pM) (pM) (pM) (pM) v36535 2 4 4 41 v36539 3 13 2 35 v36540 3 6 3 29 v36541 7 7 4 28 v36542 8 8 5 16 v36545 5 4 4 15 Control 30 15 32 6584IPTS / 200238736.2IC50(Exp. 1) IC50(Exp. 2)* Variant Human IL-31 Cynomolgus IL-31 Human IL-31 Cynomolgus IL-31(pM) (pM) (pM) (pM) BMS-981164* Different lot of cynomolgus IL-31 was used in Exp. 2.Table 2.3: Ba / F3 Assay Potency with Human and Cynomolgus IL-31 for Humanized Affinity Matured Anti-IL-31 Antibody VariantsIC50Variant Human IL-31 Cynomolgus IL-31(pM) (pM)v36974 3 40Control32 145BMS-981164EXAMPLE 3: PREPARATION AND CHARACTERIZATION OF ANTLIL-31 x ANTLIL-4Ra BISPECIFIC ANTIBODIES (IgGl BACKBONE)

[0266] Bivalent, bispecific anti-IL-31 x anti-IL-4Ra antibody variants were produced in an IgGl background using various anti-IL-31 and anti-IL-4Ra antibody paratopes as described below.

[0267] Bispecific antibody variants that are monovalent for each antigen were prepared in a format in which both the IL-31 and IL-4Ra antigen binding domains are Fab domains and in which the Fc region is a heterodimeric Fc region. These bispecific antibody variants comprise four different chains - 2 heavy chains (heavy chain A and heavy chain B) and 2 light chains (light chain A and light chain B) as described in Table 3.1.

[0268] The heavy chains comprised by the bispecific antibody variants described in Table 3.1 are human IgGl heavy chains that comprise amino acid substitutions in the CHI domain that promote correct heavy and light chain pairing and amino acid substitutions in the CH3 domain that promote formation of a heterodimeric Fc over a homodimer Fc.85IPTS / 200238736.2

[0269] The CHI domain amino acid substitutions promoting correct heavy and light chain pairing are:(i) HetFab 1Chain A: A141W / L145E / K147T / Q175E (HetFabl HCA) and Chain B: Q175R (HetFabl HCB), or(ii) HetFab 2Chain A: L145R (HetFab2 HCA) and Chain B: L145E / K147T / Q175E (HetFab2 HCB), or(iii) HetFab 3Chain A: L145E / K147T / Q175E (HetFab3 HCA) and Chain B: L145R (HetFab3 HCB).

[0270] The CH3 domain amino acid substitutions promoting heterodimeric Fc formation are:Chain A: T350V / L351Y / F405A / Y407V (HetFcA) and Chain B: T350V / T366L / K392L / T394W (HetFcB).

[0271] Fc regions comprising the HetFcA and HetFcB CH3 domains are referred to as “Het Fc.” Numbering of amino acids in the CHI domain and in the Fc region (CH2 and CH3 domains) is according to the EU index.

[0272] The Fc region of the bispecific antibody variants described in Table 3.1 also comprises the following amino acid substitutions in the CH2 domain of both heavy chains which eliminate (“knock out”) FcyR binding: L234A / L235A / D265S. These CH2 amino acid substitutions are referred to herein as “FcKO.”

[0273] In addition, certain of the bispecific antibody variants further comprise amino acid substitutions in the CH2 domain that enhance binding to the neonatal Fc receptor (FcRn). These amino acid substitutions, referred to herein “YTE mutations” or “YTE,” are: M252Y / S254T / T256E in both Fc polypeptide chains.86IPTS / 200238736.2

[0274] The light chains comprised by the bispecific antibody variants described in Table 3.1 are human kappa light chains which comprise amino acid substitutions in the CL domain that promote formation of correct heavy and light chain pairs. The CL domain amino acid substitutions are:(i) HetFab 1Chain A: F116A / Q124R / L135V / T178R (HetFabl LCA) and Chain B: Q124E / L135W / A144I / T178E / T180E (HetFabl LCB), or(ii) HetFab 2Chain A: Q124E / V133E / A144I (HetFab2 LCA) and Chain B: Q124R / T178R (HetFab2 LCB), or(iii) HetFab 3Chain A: Q124R / T178R (HetFab3 LCA) and Chain B: Q124E / V133E / A144I (HetFab3 LCB).

[0275] Numbering of amino acids in the CL domain is according to the EU index.

[0276] The genes encoding the antibody heavy and light chains were constructed via gene synthesis using codons optimized for human / mammalian expression. The anti-IL-31 Fab domain sequences were generated from the VH and VL sequences of one of the humanized, affinity matured anti-IL-31 antibody variants v36542 or v36974 described in Example 1 (see Table 1.6). The anti-IL-4Ra Fab domain sequences were generated from the VH and VL sequences of one of the anti-IL-4Ra monoclonal antibody variants v38597 or v38504. The VH and VL sequences for these anti-IL-4Ra antibody variants are provided in Table 3.2 and Fig. 20 (Table DI). The CDR sequences for these anti-IL-4Ra antibody variants are provided in Fig. 18 (Table Cl).87IPTS / 200238736.2Table 3.1: Description of Anti-IL-31 x Anti-IL-4Ra Bispecific Antibody Variants (IgGl) Heavy Chain Heavy Chain Light Chain Light ChainVariant Fc A B A Bv38681 v36542 VH- v38597 VH - v36542 VL- v38597 VL - HetFc, FcKO CHl-HetFabl CHl-HetFabl CL-HetFabl CL-HetFablHCA-CH3- HCB-CH3- LCA LCBHetFcA HetFcBv38727 v36542 VH - v38504 VH - v36542 VL - v38504 VL - HetFc, FcKO CHl-HetFab2 CHl-HetFab2 CL-HetFab2 CL-HetFab2HCA-CH3- HCB-CH3- LCA LCBHetFcA HetFcBv39439 v38597 VH - v36974 VH - v38597 VL - v36974 VL - HetFc, FcKO CHl-HetFab3 CHl-HetFab3 CL-HetFab3 CL-HetFab3HCA-CH3- HCB-CH3- LCA LCBHetFcA HetFcBv39441 v38504 VH - v36974 VH - v38504 VL - v36974 VL - HetFc, FcKO CHl-HetFab3 CHl-HetFab3 CL-HetFab3 CL-HetFab3HCA-CH3- HCB-CH3- LCA LCBHetFcA HetFcBv41543* v36542 VH - v38597 VH - v36542 VL - v38597 VL - HetFc, FcKO CHl-HetFabl CHl-HetFabl CL-HetFabl CL-HetFablHCA-CH3- HCB-CH3- LCA LCBHetFcA HetFcBv41544 v36542 VH - v38597 VH - v36542 VL - v38597 VL - HetFc, CHl-HetFabl CHl-HetFabl CL-HetFabl CL-HetFabl FcKO, YTE HCA-CH3- HCB-CH3- LCA LCBHetFcA HetFcB* v41543 is identical to v38681 except that v41543 includes a lysine residue at the C-terminus of the heavy chains Table 3.2: VH and VL Sequences of Anti-IL-4Ra Antibody VariantsVariant Domain Sequence SEQ ID NOv38597 VH QVQLVQSGAEVKKPGSSVKVSCKGSGYTFTLKAMH 103 WVRQAPGQGLEWIGIISTYYGNTNYNQKFKGRATMT VDKSTSTAYMELSSLRSEDTAVYYCARGERFYYFDY WGQGTTVTVSS VL DIQLTQSPSSLSASVGDRVTITCRASKSISKYLAWYQQ 105 KPGKAPKLLIYKRSTLQSGVPSRFSGSGSHTDFTLTISS LQPEDFATYYCQQHSEYPFTFGQGTKLEIK88IPTS / 200238736.2Variant Domain Sequence SEQ ID NOv38504 VH QVQLVQSGAEVKKPGSSVKVSCKGSGPRIYAMHWVR 104 QAPGQGLEWIGIISTYYGNTNYNQKFKGRATMTVDKS TSTAYMELS SLRSEDTAVYYCARGERFYYFDYWGQG TTVTVSS VL DIQLTQSPSSLSASVGDRVTITCRASKSISKYLAWYQQ 105 KPGKAPKLLIYKRSTLQSGVPSRFSGSGSHTDFTLTISS LQPEDFATYYCQQHSEYPFTFGQGTKLEIK3.1 Production of Bispecific Anti-IL-31 x Anti-IL-4Ra Antibody Variants

[0277] The bispecific antibody variants were expressed and characterized following generally the protocols described in International Patent Publication No. WO 2015 / 109131.

[0278] The final gene products were sub-cloned into a mammalian expression vector (typically, the mammalian expression vector pTT5 (NRC-BRI, Canada)) and expressed in CHO cells or CHO-K1 cells. Briefly, the cells were transfected in exponential growth phase (1.5 to 2 million cells / mL) with aqueous 1 mg / mL 25 kDa polyethyleneimine (PEI, Polysciences) using a PEI: DNA ratio of 2.5:1. In order to determine the optimal concentration range for forming heterodimers, the DNA was transfected in optimal DNA ratios of the heavy chain A (HC-A), heavy chain B (HC-B), and light chain A (LC-A) and light chain B (LC-B) that allow for heterodimer formation (for example, HC-A: HC-B: LC-A: LC-B ratios of 3:3: 10:4 or 15: 15:53: 17 or 15: 15:35:35 or 22:8:53:17 or 22:8:35:35). Transfected cells were harvested after 5-6 days and the culture medium collected after centrifugation at 4000 rpm and clarified using a 0.45 pm filter. The clarified culture medium was loaded onto a MabSelect™ SuRe™ Protein A column (GE Healthcare) and washed with 10 column volumes of PBS buffer at pH 7.2 - 7.4. The antibody variant was eluted with 10 column volumes of 0.1 M citrate buffer at pH 3.6 with the pooled fractions containing antibody variant neutralized with 1 M TRIS at pH 9 or 11. The amount of antibody variant was then quantified based on A280 nm (NanoDrop™ Spectrophotometer; Thermo Fisher Scientific).

[0279] The antibody variants were further purified by gel filtration chromatography using a Superdex™ 200 HiLoad™ 16 / 600200pg column (GE Healthcare) via an AKTA Express FPLC or AKTA Pure chromatography system at a flowrate of 1 mL / min with PBS buffer at pH 7.4 or 89IPTS / 200238736.2H6-NaCl buffer (20 mM histidine, 150 mM sodium chloride, pH 6.0) used at a flow-rate of 1 mL / min. Some antibody variants were purified by ion exchange chromatography using a Capto™ SP ImpRes column (Cytiva) or POROS™ XS column (Thermo Fisher Scientific) using a salt gradient with HEPES buffer, pH 7.5 (step elution with 200 mM and 400 mM sodium chloride). Fractions of eluted antibody variant were collected based on absorbance at A280 nm and the fractions were assessed by non-reducing and reducing CE-SDS or High Throughput Protein Express assay using Caliper LabChip™ GXII (both from Perkin Elmer, Waltham, MA). Procedures were carried out according to HT Protein Express LabChip™ User Guide version 2 and LabChip™ GXII User Manual, with the following modifications. Antibody variant samples, at either 2 pl or 5 pl (concentration range 5-2000 ng / pl), were added to separate wells in 96 well plates (BioRad, Hercules, CA) along with 7 pl of HT Protein Express Sample Buffer (Perkin Elmer Cat No. 760328). Antibody variant samples were then denatured at 90 °C for 5 mins. The LabChip™ instrument was operated using the HT Protein Express Chip (Perkin Elmer, Waltham, MA) and the Ab-200 assay setting. Fractions corresponding to the purified antibody variants were collected, buffer exchanged into 20 mM histidine, pH 6.0 using a Zeba™ Spin desalting column (Thermo Fisher Scientific), concentrated to ~1 mg / mL or 20 mg / mL, and stored at -80 °C.

[0280] Endotoxin levels were determined by the LAL (limulus amebocyte lysate) assay using the Endosafe™ Portable Test System (PTS, Charles River, Wilmington, MA). Antibody variants were qualified based on A280 nm absorbance (NanoDrop™ spectrophotometer) post protein-A and SEC purification. UPLC-SEC was performed using a Waters™ Acquity BEH200 SEC column (2.5 mL, 4.6x150 mm, stainless steel, 1.7 pm particles) (Waters Corporation, Mississauga, ON) set to 30 °C or 25 °C and mounted on a Waters™ Acquity UPLC H-Class Bio system with a photodiode array (PDA) detector. Run times were 7 min with a total volume per injection of 2.8 pL or 5 pL and a running buffer of Dulbecco’s phosphate-buffered saline (DPBS) or DPBS with 0.02% Tween™ 20 pH 7.4 or 200mM potassium phosphate with 200 mM potassium chloride pH 7.0 at 0.4 mL / min. Elution was monitored by UV absorbance in the range 210-500 nm, and chromatograms were extracted at 280 nm. Peak integration was performed using Empower™ 3 software (Waters Corporation).

[0281] The apparent purity and yield of the final antibody variant was estimated by UPLC- SEC and LC / MS following protocols as described in International Patent Publication No. WO90IPTS / 200238736.22015 / 109131. All antibody variants were expressed and purified to >80% heterodimer purity without contaminating homodimers (see Table 3.3).Table 3.3: Post Purification Yield and Purity for Anti-IL-31 x Anti-IL-4Ra Bispecific Antibody Variants (IgGl)Titer MonomerVariant Volume Yield LCMS(mg / L) Buffer Purity (%)' Purity (%)2(mg / L) (L)v38681 58 20 37 99.9 94.9 20 mM Histidine pH 6.0 V38727 72 25 16 99.4 82.9 20 mM Histidine pH 6.0 V39439 74 20 40 99.6 90.1 20 mM Histidine pH 6.0 V39441 55 25 22 99.9 89.7 20 mM Histidine pH 6.0 V41543 1080 20 548 99.9 99.9 20 mM Histidine pH 6.0 V41544 1150 20 410 99.9 99.9 20 mM Histidine pH 6.01As determined by size-exclusion chromatography HPLC (HPLC-SEC)2As determined by LCMS intensity3.2 Characterization of Bispecific Anti-IL-31 x Anti-IL-4Ra Antibody Variants (IgGl) 3.2.1 Thermal Stability of Bispecific Antibody Variants

[0282] The thermal stability of the bispecific anti-IL-31 x anti-IL-4Ra antibody variants was assessed by differential scanning calorimetry (DSC) and differential scanning fluorimetry (DSF).

[0283] All DSC experiments were carried out using a VP-Capillary DSC instrument (Malvern Instruments Ltd). The proteins were diluted to 0.4 mg / mL or 1 mg / mL with 0.4 mL loaded into the 96 well plates and measured with a scan rate of 1 °C / min from 20 °C to 100 °C. Data was analyzed using the Origin 7.0 software (OriginLab Corporation) with the VP-Capillary DSC option and with the 20 mM histidine, pH 6.0 buffer background subtracted.

[0284] All DSF experiments were carried out using a CFX96 Touch™ Real-Time PCR instrument (BioRad Laboratories, Inc., Hercules, CA). The proteins were diluted to 1 mg / mL and 10 pg was loaded into 35 pL buffer in 96 well plates with 5 pL of 40x Invitrogen™ SYPRO™ Orange Protein Gel Stain (5,000X concentrate in DMSO) (Thermo Fisher Scientific) and measured with a scan rate of 0.5 °C / min from 25°C to 95°C. Data was analyzed using Microsoft Excel with91IPTS / 200238736.2the 20 mM histidine, pH 6.0 buffer background subtracted. Some experiments were carried out on aNanoDSF instrument (NanoTemper Technologies) using a similar protocol.

[0285] The results are shown in Table 3.4, which shows the maximum melting temperatures (Tm) for each of the peaks in the thermograms of the tested bispecific antibody variants. The results demonstrate that the tested antibody variants exhibited thermostability profiles comparable to those of conventional IgGl antibodies.Table 3.4: Thermostability of Anti-IL-31 x Anti-IL-4Ra Bispecific Antibody Variants (IgGl)DSC DSFVariantTml (°C) Tml (°C) Tm2 (°C)v38681 70.5 69.2 - v38727 70.5 69.3 - v41543 ND 68 76v41544 ND 63.3 76Trastuzumab 70.4 68 78(IgGl control)3.2.2 Accelerated Stability of Bispecific Antibody Variants

[0286] The accelerated stability of the bispecific anti-IL-31 x anti-IL-4Ra antibody variants was evaluated by assessing the purity after storage at 40 °C for 4 weeks. The bispecific variants were stored at 1 mg / mL in 20 mM histidine pH 6.0 buffer and the purity was assessed by CE-SDS and UPLC-SEC (as described in Example 1).

[0287] The results are shown in Tables 3.5 and 3.6 and demonstrate that the bispecific antibody variants displayed minimal changes in % monomer purity as measured by UPLC-SEC and CE-SDS, respectively. There was slight decrease in purity observed after 4 weeks at 40 °C with an increase in low molecular weight species (LMWs) but not high molecular weight species (HMWs) as measured by UPLC-SEC (see Table 3.5).92IPTS / 200238736.2Table 3.5: Purity of Anti-IL-31 x Anti-IL-4Ra Bispecific Antibody Variants Assessed by UPLC-SEC after Storage at 40 °C for 4 Weeks% Monomer % HMWs % LMWs VariantDay 0 4 weeks Day 0 4 weeks Day 0 4 weeks v38681 97.6 95.8 0.8 0.4 1.6 3.8v38727 99.0 96.3 0.6 1.1 0.3 2.6Table 3.6: Purity of Anti-IL-31 x Anti-IL-4Ra Bispecific Antibody Variants Assessed by CE-SDS after Storage at 40 °C for 4 Weeks% MonomerVariantDay 0 4 weeksv38681 95.1 93.1v38727 96.7 94.8EXAMPLE 4: PREPARATION AND CHARACTERIZATION OF ANTI-IL-31 x ANTI-IL-4Ra BISPECIFIC ANTIBODIES (IgG4 BACKBONE) #1

[0288] Bivalent, bispecific anti-IL-31 x anti-IL-4Ra antibody variants were produced in an IgG4 background using various anti-IL-31 and anti-IL-4Ra antibody paratopes as described below.

[0289] Bispecific antibody variants that are monovalent for each antigen were prepared in a format in which both the IL-31 and IL-4Ra antigen binding domains are Fab domains and in which the Fc region is a heterodimeric Fc region. These bispecific antibody variants comprise four different chains - 2 heavy chains (heavy chain A and heavy chain B) and 2 light chains (light chain A and light chain B) as described in Table 4.1.

[0290] The heavy chains comprised by the bispecific antibody variants described in Table 4.1 are human IgG4 heavy chains that comprise amino acid substitutions in the CHI domain that promote correct heavy and light chain pairing and amino acid substitutions in the CH3 domain that 93IPTS / 200238736.2promote formation of a heterodimeric Fc over a homodimer Fc. The heavy chains also contained the human IgG4 hinge domain which further comprised amino acid substitution S228P.

[0291] The CHI domain amino acid substitutions promoting correct heavy and light chain pairing are:HetFab 1Chain A: Q175R (HetFab HCA) and Chain B: A141W / L145E / K147T / Q175E (HetFab HCB)

[0292] The CH3 domain amino acid substitutions promoting heterodimeric Fc formation are:Chain A: T350V / L351Y / F405A / Y407V / R409K (HetFcA) and Chain B: T350V / T366L / K392L / T394W / R409K (HetFcB).

[0293] Fc regions comprising the HetFcA and HetFcB CH3 domains are referred to as “Het Fc.” Numbering of amino acids in the CHI domain and in the Fc region (CH2 and CH3 domains) is according to the EU index.

[0294] In addition, certain of the bispecific antibody variants further comprise amino acid substitutions in the CH2 domain that enhance binding to the neonatal Fc receptor (FcRn). These amino acid substitutions, referred to herein “YTE mutations” or “YTE,” are: M252Y / S254T / T256E in both Fc polypeptide chains.

[0295] The light chains comprised by the bispecific antibody variants described in Table 4.1 are human kappa light chains which comprise amino acid substitutions in the CL domain that promote formation of correct heavy and light chain pairs. The CL domain amino acid substitutions are:HetFab 1Chain A: Q124E / L135W / A144I / T178E / T180E (HetFab LCA) and Chain B: Fl 16A / Q124R / L135V / T178R (HetFab LCB)94IPTS / 200238736.2

[0296] Numbering of amino acids in the CL domain is according to the EU index.

[0297] In the bispecific antibody variants, the VH and VL sequences of each of the anti-IL-31 Fab domain and the anti-IL-4Ra Fab domain were coupled to human IgG4 CHI and CL sequences, respectively.

[0298] The anti-IL-31 Fab domain sequences were generated from the VH and VL sequences of the humanized, affinity matured anti-IL-31 antibody variant v36542 described in Example 1 (see Table 1.6). The anti-IL-4Ra Fab domain sequences were generated from the VH and VL sequences of the anti-IL-4Ra monoclonal antibody variant v38597 (see Example 3) or from the VH and VL sequences of the anti-IL-4Ra monoclonal antibody v38597-Y54A. The VH and VL sequences ofv38597-Y54Aare identical to those ofv38597 except that an amino acid substitution, Y54A, was made in CDRH2 of the VH domain to potentially reduce hydrophobicity. The VH and VL sequences of v38597-Y54A are provided in Table 4.2.Table 4.1: Description of Anti-IL-31 x Anti-IL-4Ra Bispecific Antibody Variants (IgG4) Heavy Chain A Heavy Chain B Light Chain A Light Chain BVariant Fc (anti-IL-4Ra) (anti-IL-31) (anti-IL-4Ra) (anti-IL-31)v38597 VH- v36542 VH- V38597 VL- v36542 VL- CHl-HetFab CHl-HetFab HetFc, v41789 CL-HetFab CL-HetFabHCA-CH3- HCB-CH3- IgG4 LCA LCBHetFcA HetFcBv38597 VH- v36542 VH- V38597 VL- v36542 VL- HetFc, CHl-HetFab CHl-HetFabv41790 CL-HetFab CL-HetFab IgG4, HCA-CH3- HCB-CH3- LCA LCB YTEHetFcA HetFcBV38597-Y54A v36542 VH- V38597-VL- v36542 VL- HetFc, VH-CH1- CHl-HetFabv41791 CL-HetFab CL-HetFab IgG4, HetFab HCA- HCB-CH3- LCA LCB YTE CH3-HetFcA HetFcB95IPTS / 200238736.2Table 4.2: VH and VL Sequences of Anti-IL-4Ra Antibody Variant v38597-Y54A Variant Domain Sequence SEQ ID NO VH QVQLVQSGAEVKKPGSSVKVSCKGSGYTFTLKAMH 121* WVRQAPGQGLEWIGIISTYAGNTNYNQKFKGRATMT VDKSTSTAYMELSSLRSEDTAVYYCARGERFYYFDYv38597- WGQGTTVTVSSY54AVL DIQLTQSPSSLSASVGDRVTITCRASKSISKYLAWYQQ 105 KPGKAPKLLIYKRSTLQSGVPSRFSGSGSHTDFTLTISS LQPEDFATYYCQQHSEYPFTFGQGTKLEIK* SEQ ID NO: 121 is identical to SEQ ID NO: 106 and SEQ ID NO: 2244.1 Preparation of Bispecific IL-31 x IL-4Ra Antibody Variants

[0299] The bispecific antibody variants were expressed and characterized following the protocols described in International Patent Publication No. WO 2015 / 109131.

[0300] Briefly, the genes encoding the bispecific antibody heavy and light chains were constructed via gene synthesis using codons optimized for human / mammalian expression. The final gene products were sub-cloned into a mammalian expression vector and expressed in CHO KI cells. The CHO cells were transfected in exponential growth phase (1.5 to 2 million cells / mL). In order to determine the optimal concentration range for forming heterodimers, the DNA was transfected in optimal DNA ratios of the heavy chain A (HC-A), heavy chain B (HC-B), and light chain A (LC-A) and light chain B (LC-B) that allow for heterodimer formation (for example, HC-A: HC-B: LC-A: LC-B ratios of 15:15:53:17 or 15:15:35:35 or 8:22:35:35). Transfected cells were harvested after 5-6 days and the culture medium collected after centrifugation at 4000 rpm and clarification using a 0.45 pm filter. The clarified culture medium was loaded onto a MabSelect™ SuRe™ Protein A column (GE Healthcare) and eluted with 0.1 M citrate buffer or 0.1M acetate buffer at pH 3.0 with the pooled fractions containing antibody variant neutralized with 1 M TRIS at pH 9 or 11. The amount of antibody variant was then quantified based on A280 nm (NanoDrop™ Spectrophotometer; Thermo Fisher Scientific).

[0301] The antibody variants were further purified by cation exchange chromatography using a POROS™ XS column with a salt gradient of 20 mM sodium acetate, IM NaCl buffer, pH 5.5. Fractions of eluted antibody variant were collected based on absorbance at A280 nm and the 96IPTS / 200238736.2fractions were assessed by non-reducing SDS-PAGE. Fractions corresponding to the purified antibody variants were collected, buffer exchanged into 20 mM histidine, pH 6.0, concentrated to ~20 mg / mL, and stored at -80 °C.

[0302] The apparent purity and yield of the final antibody variant was estimated by HPLC-SEC and LC / MS following generally the protocols as described in International Patent Publication No. WO 2015 / 109131. All antibody variants were expressed and purified to >95% heterodimer purity without contaminating homodimers (see Table 4.3).Table 4.3: Post Purification Yield and Purity for Anti-IL-31 x Anti-IL-4Ra Bispecific Antibody Variants (IgG4)Titer Volume Yield Monomer LCMSVariant Buffe (mg / L) (mg / L) Purity (%)' Purity (%)2r(L)V41789 1570 1 500 99.7 95.4 20 mM Histidine pH 6.0 V41790 1290 7 462 99.5 99.4 20 mM Histidine pH 6.0 V41791 1216 7 493 100 99.7 20 mM Histidine pH 6.01As determined by size-exclusion chromatography HPLC (HPLC-SEC)2As determined by LCMS intensity4.2 Characterization of Bispecific Anti-IL-31 x Anti-IL-4Ra Antibody Variants (IgG4) 4.2.1 Thermal Stability of Bispecific Antibody Variants

[0303] The thermal stability of the bispecific anti-IL-31 x anti-IL-4Ra antibody variants was assessed by Differential Scanning Calorimetry (DSC) and Differential Scanning Fluorimetry (DSF).

[0304] All DSC experiments were carried out using a TA Instruments Nano DSC instrument (Waters Corporation). The proteins were diluted to 1 mg / mL with 1 mL loaded into the 96 well plates and measured with a scan rate of 1 °C / min from 20 °C to 90 °C. Data was analyzed using the Nano DSC analysis software with the 20 mM histidine, pH 6.0 buffer background subtracted.

[0305] All DSF experiments were carried out using a CFX96 Touch™ Real-Time PCR instrument (BioRad Laboratories, Inc., Hercules, CA) as described in Example 3 (Section 3.2.1).97IPTS / 200238736.2

[0306] The results are shown in Table 4.4, which shows the maximum melting temperatures (Tm) for each of the peaks in the thermograms of the tested bispecific antibody variants. The results demonstrate that the tested antibody variants exhibited thermostability profiles comparable to those of the corresponding IgGl bispecific constructs in Example 3. Table 4.4: Thermostability of Anti-IL-31 x Anti-IL-4Ra Bispecific Antibody Variants (IgG4)DSC DSFVariantTml (°C) Tm2 (°C) Tm3 (°C) Tml (°C) Tm2 (°C) v41789 65.4 71.7 77.7 64 73v41790 58.7 71.5 77.7 58.8 73v41791 59 71.2 77.8 59 72.74.2.2 Stability of Bispecific Antibody Variants

[0307] The bispecific antibody variants were tested for freeze-thaw stability and long-term storage stability, as well as stability under an accelerated stress test. Purity of the bispecific antibody variants pre- and post-treatment was assessed by UPLC-SEC except where noted.

[0308] For freeze-thaw stability, the bispecific antibody variants were subjected to 5 cycles of freezing to -80 °C followed by thawing to 4 °C using protein concentrations of 20 mg / mL or 150 mg / mL in 20 mM histidine pH 6.0 buffer. No significant change in purity was observed for any of the bispecific antibody variants. See Table 4.5 (20 mg / mL) and Table 4.8 (150 mg / mL).

[0309] For long-term storage stability, the bispecific antibody variants were stored at -80 °C for 10 weeks using a protein concentration of 20 mg / mL in 20 mM histidine pH 6.0 buffer. No significant change in purity was observed for any of the bispecific antibody variants. See Table 4.7.

[0310] For the accelerated stress test for stability, the bispecific antibody variants were incubated at 40 °C for 14 days using a protein concentration of 20 mg / mL or 150 mg / mL in 20 mM histidine pH 6.0 buffer. The bispecific antibody variants showed only a minimal change in purity after the 14 day incubation. See Table 4.6 (20 mg / mL) and Table 4.8 (150 mg / mL).98IPTS / 200238736.2Table 4.5: Purity (%) of Bispecific Antibody Variants After 5 Cycles of Freeze / Thaw (5xF / T)% Monomer % HMWs1% LMWs2VariantDay 0 5x F / T Day 0 5x F / T Day 0 5x F / T v41789 98.54 97.66 1.46 2.34 0 0v41790 98.54 98.43 1.46 1.57 0 0v41791 99.52 99.54 0.48 0.46 0 0Table 4.6: Purity (%) of Bispecific Antibody Variants After 4 Weeks Storage at 40 °C% Monomer % HMWs1% LMWs2VariantDay 0 4 weeks Day 0 4 weeks Day 0 4 weeks v41789 ND3ND3ND3ND3ND3ND3v41790 99.62 98.42 0.38 1.58 0 0 v41791 99.52 98.76 0.44 1.24 0.01 0Table 4.7: Purity (%) of Bispecific Antibody Variants After 2 Months Storage at -80 °C% Monomer % HMWs1% LMWs2VariantDay 0 2 months Day 0 2 months Day 0 2 months v41789 98.54 97.28 1.46 1.1 0 1.64 v41790 98.54 99.19 1.46 0.82 0 0 v41791 99.52 98.92 0.48 0.41 0 0.68Table 4.8: Purity (%)4of Bispecific Antibody Construct After 5 Cycles of Freeze / Thaw and After 4 Weeks Storage at 40 °Cv41789 v41790 v41791No treatment 100 100 1005 cycles freeze / thaw 100 100 1004 weeks storage at 40 °C 100 100 10099IPTS / 200238736.21HMWs = High molecular weight species2LMWs = Low molecular weight species3ND = Not determined4Monomer Purity (%) by HPLC-SEC AnalysisEXAMPLE 5: CELLULAR BINDING OF ANTI-IL-31 x ANTI-IL-4Ra BISPECIFIC ANTIBODIES (IgG4 BACKBONE)

[0311] To test the ability of the bispecific antibody variants to bind human and cynomolgus IL-4Ra, selected antibody variants were assessed for binding to PBMCs by flow cytometry as described below. The anti-IL-4Ra antibody dupilimab was used as a positive control for IL-4Ra binding and a human IgG4 isotype (palivizumab, v36992) was used as a non-specific negative control. A tetravalent, bispecific anti-IL-31 x anti-IL-4Ra antibody based on NM26-2198 (Numab Therapeutics) was also included as a positive control.

[0312] Flow cytometry was performed on human PBMCs (StemCell Technologies, Vancouver Canada) or cynomolgus PBMCs (BioIVT, Westbury, NY). Briefly, PBMCs were thawed and plated in 96 well v-bottom plates at 1×105cells / well. Antibody variants were serially diluted in flow cytometry staining buffer (5% FBS, 2.5mM EDTA, PBS). Cells were stained with Fixable Viability Dye eFluor™ 506 (Thermo Fisher Scientific, Waltham, MA) for 15 minutes in PBS. Cells were washed once with PBS, washed once with flow cytometry staining buffer, then incubated with antibody variants at 4°C for 1 hour in flow cytometry staining buffer to allow binding. Cells were washed and stained for 30 minutes with fluorescently-conjugated antibodies against human cell surface markers (CD3, CD4, CD14, CD15, CD16, CD19) or cynomolgus cell surface markers (CD4, CD8, CD1 lb, CD14, CD16, CD20, TCR-J3) to allow phenotyping of cells by flow cytometry. An anti-human-IgG4 antibody (Southern Biotech, Birmingham, AL) was included in the antibody panel to detect antibody variant binding to PBMCs. Cells were washed with flow cytometry staining buffer, fixed for 10 minutes in BD Cytofix™ Fixation Buffer (BD Biosciences, San Francisco, CA), washed with flow cytometry staining buffer, resuspended in flow cytometry staining buffer, and analyzed by flow cytometry on an LSR Fortessa™ X-20 flow cytometer (BD Biosciences). Geometric mean fluorescent intensity (gMFI) of anti-human-IgG was used to assess binding of constructs on CD4+T cell (Human: CD3+CD19-CD4+; Cynomolgus: CD3+CD20-TCR-β+CD4+CD8-), B cell (Human: CD3" CD19+; Cynomolgus:100IPTS / 200238736.2TCR-P" CD20+) and monocyte (CD14+CD16+ / ) populations as defined by phenotyping antibody panel staining.

[0313] Bispecific antibody variants were shown to bind IL-4Ra across multiple cell types known to express IL-4Ra (T cells, B cells, and monocytes) as shown by the presence of an anti-human-IgG positive population. Furthermore, the gMFI of anti-human-IgG staining increased as bispecific antibody variant concentration increased, while the isotype control gMFI did not, indicating specific binding of the bispecific antibody variants to their target. Results for human PBMC binding are shown in Fig. 1A (CD3+CD4+T cells), Fig. IB (CD3 CD19+B cells) and Fig.1C (CD3 CD14 CD15 CD16" classical monocytes).EXAMPLE 6: FUNCTIONAL CHARACTERIZATION OF BISPECIFIC ANTIBODY VARIANTS (IgGl AND IgG4 BACKBONE) - IL-4 / 13 REPORTER GENE ASSAY

[0314] To determine the impact of the anti-IL-31 x anti-IL-4Ra bispecific antibody variants on IL-4 / IL-13 activation of the STAT6 signalling pathway, selected variants were assessed for inhibition of IL-4 / IL-13 mediated production of the STAT6 inducible secreted embryonic alkaline phosphatase (SEAP) reporter in HEK-Blue™ IL-4 / IL-13 cells as described below. The variants tested were: v38727, v39441, v38681, v39439, v41789, v41790, v41791, v41544 (IL-13 and IL-4 stimulation) and v41543 (IL-4 stimulation only) (See Tables 3.1 and 4.1). A bivalent anti-IL-31 antibody (BMS-981164), a bivalent anti-IL-4Ra antibody (dupilumab), a tetravalent anti-IL-4Ra x anti IL-31 bispecific antibody (NM26-2198), ananti-RSV IgGl antibody (v22277), an anti-RSV IgGl FcKO antibody (v39982), an anti-RSV IgG4 antibody (v36992) and an IgG4 isotype antibody (v42104) were used as controls.

[0315] Briefly, test articles were serially diluted starting at a concentration of 20 nM in DMEM + 10% heat-inactivated fetal bovine serum (ThermoFisher Scientific, Waltham, MA) and plated into a 384-well black flat bottom assay plate. 125 pM IL-4 or 500 - 5000 pM IL- 13 (R& D Systems, Minneapolis, MN) was added, followed by 5000 - 12500 HEK-Blue™ IL-4 / IL-13 cells (InvivoGen, San Diego, CA). After 24 hr at 37°C, 5% CO2, SEAP production was assessed by incubation of supernatant with QUANTI-Blue™ solution (InvivoGen, San Diego, CA) and measuring OD620nmon a Synergy™ plate reader (BioTek Instruments, Inc., Winooski, VT).101IPTS / 200238736.2

[0316] The results are shown in Figs. 2A-E. All bispecific antibody variants blocked IL-4 and IL- 13 mediated production of STAT6 inducible SEAP reporter in HEK-Blue™ IL-4 / IL-13 cells. The level of inhibitory activity was agnostic of Fc format (IgGl vs IgG4) and the presence or absence of the YTE mutations. All bispecific antibody variants showed similar or superior inhibitory activity compared to bivalent anti-IL-4Ra antibody control (dupilumab) and tetravalent anti-IL-4Ra x anti -IL-31 bispecific antibody control (NM26-2198). IgG4 isotype and anti-RSV controls showed no activity as expected.EXAMPLE 7: FUNCTIONAL CHARACTERIZATION OF BISPECIFIC ANTIBODY VARIANTS (IgGl AND IgG4 BACKBONE) - Ba / F3 PROLIFERATION ASSAY

[0317] Selected anti-IL-31 x anti-IL-4Ra bispecific antibody variants (see below) were assessed for inhibition of IL-31 mediated proliferation in Ba / F3 cells stably expressing human IL-3 IRA and oncostatin M receptor (OSMR) following the protocol described in Example 2. A bivalent anti-IL-31 antibody (BMS-981164), a bivalent anti-IL-4Ra antibody (dupilumab), a tetravalent anti-IL-4Ra x anti IL-31 bispecific antibody (NM26-2198), an anti-RSV IgGl FcKO antibody (v39982) and an IgG4 isotype antibody (v42104) were used as controls. The monospecific affinity matured anti-IL-31 antibody variants v36542 and v36974 (see Table 1.6) were also included.

[0318] Bispecific antibody variants tested were: v38727, v39441, v38681, v39439,V41544, v41789, v41790 and v41791 (see Table 3.1 and 4.1).

[0319] The results are shown in Figs. 3A & 3B. All bispecific antibody variants blocked IL-31 mediated proliferation of Ba / F3 cells. The level of inhibition of proliferation was agnostic of Fc format (IgGl vs IgG4) and the presence or absence of the YTE mutations. All bispecific antibody variants showed higher activity compared to the bivalent anti-IL-31 antibody control (BMS-981164) and tetravalent anti-IL-4Ra x anti IL-31 bispecific antibody control (NM26-2198). Dupilumab, IgG4 isotype and anti-RSV controls showed no activity as expected.102IPTS / 200238736.2EXAMPLE 8: FUNCTIONAL CHARACTERIZATION OF BISPECIFIC ANTIBODY VARIANTS (IgGl AND IgG4 BACKBONE) - IL-4 PBMC (CD23) ASSAY

[0320] To test the functional impact of blockade on IL-4Ra signalling by the bispecific antibodies, selected anti-IL-31 x anti-IL-4Ra bispecific antibody variants were assessed for inhibition of CD23 upregulation by PBMCs following IL-4 stimulation. Flow cytometry was used to assess CD23 expression following IL-4 stimulation as described below. Dupilumab was used as a positive control for blocking Il-4Ra signalling, while an anti-hemagglutinin (HA) IgG4 antibody (v42104) was used as a non-specific negative control.

[0321] The antibody variants tested were: v41544, v41789, v41790 and v41791 (see Table 3.1 and Table 4.1).

[0322] Human PBMCs (StemCell, Vancouver Canada) were plated at 2xl05cells / well in 96 well round-bottom plates and incubated with serially diluted concentrations of antibody variants for 30 minutes to allow binding. IL-4 (R& D Systems, Minneapolis, MN) was added to each well to a final concentration of 2ng / mL and cells were incubated at 37°C for 48 hours in 10% FBS RPMI (ThermoFisher Scientific, Waltham, MA). Following incubation, cells were stained with Fixable Viability Dye eFluor™ 506 (Thermo Fisher Scientific, Waltham, MA) for 15 minutes in PBS, washed with PBS, washed with flow cytometry staining buffer (5% FBS, 2.5mM EDTA, PBS), and stained in flow cytometry staining buffer for 30 minutes with fluorescently-conjugated antibodies against human cell surface markers (CD3, CD14, CD16, CD19, CD23, CD27) to allow phenotyping of cells by flow cytometry. Cells were washed twice with flow cytometry staining buffer and fixed for 10 minutes in BD Cytofix™ Fixation Buffer (BD Biosciences, San Francisco, CA). Cells were washed and resuspended in flow cytometry staining buffer then analyzed by flow cytometry on an LSR Fortessa™ X-20 flow cytometer (BD Biosciences). Geometric mean fluorescent intensity (gMFI) of CD23 on various immune cell populations as defined by phenotyping antibodies (B-cells: CD19+CD3" CD27+ / ", T-cells: CD3+CD19", Monocytes: CD14+CD16+ / ) was used to assess inhibition of IL-4Ra signalling.

[0323] The bispecific antibody variants were shown to limit changes to cellular phenotypes resulting from IL-4Ra stimulation as evidenced by decreasing CD23 expression as antibody variant concentration increases. Specifically, naive B-cells and monocytes, which have been 103IPTS / 200238736.2shown to increase CD23 expression following IL-4Ra stimulation, were both shown to have reduced CD23 expression which was dependent on antibody variant concentration. This effect was not seen in cells treated with isotype control showing that the effect was mediated by inhibition of IL-4Ra stimulation by the antibody variants. Furthermore, the antibody variants with an IgG4 backbone (v41789, v41790 and v41791) outperformed the corresponding antibody variant with a shared binding sequence but an IgGl backbone (v41544) suggesting that the IgG4 backbone provides increased efficacy in preventing activation of key cell populations which respond to II-4Ra stimulation. Results for CD23 expression on monocytes are shown in Fig. 4.EXAMPLE 9: FUNCTIONAL CHARACTERIZATION OF BISPECIFIC ANTIBODY VARIANTS (IgG4 BACKBONE) - IL-4 / 13 / 31 HEKALPHA (CCL2) ASSAY

[0324] The anti-IL-31 x anti-IL-4Ra bispecific antibody variant v41791 (see Table 4.1) was assessed for the ability to inhibit CCL2 gene expression in HEKa cells following combined cytokine stimulation of the IL-4, IL- 13 and IL-31 pathways as described below. An anti-IL-4Ra monospecific antibody (dupilumab) and an anti -IL-31 monospecific antibody (BMS-981164) were used as controls.

[0325] Briefly, 9xl04HEKa cells (ATCC, Manassas, VA) were plated in a flat-bottom 24 well plate in assay media (1:1 mix of DMEM and Ham’s F12K (both from Sigma-Aldrich, St. Louis, MO), and 10% FBS) and incubated at 37°C for 24 hours. All cytokines were prepared separately in assay media for a final concentration in cell cultures of 50nM per cytokine. Prior to adding to cell cultures, antibody variants were added into IL-31 cytokine (R& D Systems, Minneapolis, MN) preparations and incubated for 30 minutes at room temperature to allow binding of cytokine by the antibody variant. Media was aspirated from cell cultures and ImL of cytokineantibody variant mixtures were added to applicable wells. Cell cultures were incubated for 30 minutes at room temperature to allow the antibody variant to bind IL-4Ra. After 30 minutes, IL-13 (R& D Systems, Minneapolis, MN) was added to wells for a final concentration of 50nM. Cells were incubated at 37°C for 6 hours, then media was aspirated out of the wells and cells were washed twice with cold PBS. RNA was isolated from the cells via RNeasy™ Mini Kit columns (Qiagen, Hilden, Germany) following manufacturer’s instructions and cDNA was generated from the isolated RNA using a High-Capacity cDNA Reverse Transcription Kit (Applied Biosystems,104IPTS / 200238736.2Waltham, MA) following manufacturer’s instructions. qPCR was then performed using TaqMan™ Gene Expression Assay primers and TaqMan™ Universal PCR Master Mix (Applied Biosystems, Waltham, MA) following manufacturer’s instructions, and a QuantStudio™ 3 Real-Time PCR machine (ThermoFisher Scientific, Waltham, MA) to quantify the amount of CCL2 and ubiquitin C (UBC) mRNA in the original isolated RNA sample. CCL2 mRNA production was used as a proxy for measuring cell stimulation after combined cytokine exposure. Ubiquitin C (UBC) was used as the housekeeping gene. Relative quantification of CCL2 mRNA was determined by calculating the double delta Ct (threshold cycle) for each sample. Briefly, the Ct of CCL2 cDNA was subtracted from the Ct of UBC cDNA from the same sample to give the delta Ct. This value was then subtracted by the delta Ct of unstimulated control samples to provide the delta delta Ct value. Taking the natural log of the negative of this value gives the fold change in CCL2 gene expression in experimental samples as compared to unstimulated controls.

[0326] The results are shown in Fig. 5. Cells incubated with the bispecific antibody variant v41791 show a lower level of CCL2 mRNA production compared to cells incubated with either cytokine alone, cytokine combinations, monospecific antibody controls or combinations of the monospecific antibody controls. This suggests that the bispecific antibody can inhibit CCL2 mRNA production by blocking stimulation through both IL-4Ra signalling and by sequestering free-floating IL-31, preventing cells which express IL-4Ra from receiving IL-31 stimulation. EXAMPLE 10: FUNCTIONAL CHARACTERIZATION OF BISPECIFIC ANTIBODY VARIANTS (IgGl AND IgG4 BACKBONE) -IL-4 INDUCED CCL17 PRODUCTION

[0327] Selected anti-IL-31 x anti-IL-4Ra bispecific antibody variants were assessed for inhibition of IL-4 mediated production of CCL17 (also known as thymus and activation regulated chemokine (TARC)) in peripheral blood mononuclear cells (PBMCs). An anti-IL-4Ra antibody (dupilumab), an anti-IL-31 monospecific antibody (BMS-981164), a tetravalent anti-IL-4Ra x IL-31 bispecific antibody (NM26-2198), an anti-RSV IgGl FcKO antibody (v39982), an anti-RSV IgG4 antibody (v36992) and an IgG4 isotype antibody (v42104) were used as controls. Also included was the anti-IL-4Ra monospecific antibody (v36912) that is the parental antibody for the anti-IL-4Ra paratopes included in the anti-IL-4Ra x anti IL-31 bispecific variants.105IPTS / 200238736.2

[0328] The bispecific antibody variants tested were: v41543, v41789, v41790, v41791 and v41544 (see Table 3.1 and Table 4.1).

[0329] Test articles were serially diluted with a top concentration of 20 nM in RPMI + 10% heat inactivated fetal bovine serum) (ThermoFisher Scientific, Waltham, MA) and plated in a 384-well black flat bottom assay plate (ThermoFisher Scientific, Waltham, MA). 300 pM human IL-4 (R& D Systems, Minneapolis, MN) was added followed by 40 000 freshly thawed PBMCs (StemCell Technologies, Vancouver, Canada). After 24 hours at 37°C, 5% CO2, CCL17 levels in supernatant were quantified with U-PLEX™ human TARC assay (Meso Scale Discovery, Rockville, MD) following the manufacturer’s instructions.

[0330] The results are shown in Figs. 6A-6E. All bispecific antibody variants demonstrated blockade of IL-4 mediated CCL17 production by PBMCs. The level of inhibitory activity was agnostic of Fc format (IgGl vs IgG4) and the presence or absence of the YTE mutations. All bispecific antibody variants showed comparable inhibitory activity to the bivalent anti-IL-4Ra antibody controls (dupilumab and parental antibody, v36912) and to the tetravalent anti-IL-4Ra x IL-31 bispecific antibody control (NM26-2198).EXAMPLE 11: PHARMACOKINETICS OF BISPECIFIC ANTIBODY VARIANTS (IgG4 BACKBONE) IN A RAT MODEL

[0331] The pharmacokinetics (PK) of two anti-IL-31 x anti-IL-4Ra bispecific antibody variants, v41790 and v41791 (see Table 4.1), were assessed in a rat PK model as described below. An anti-IL-4Ra monospecific antibody (dupilumab) and a tetravalent, bispecific anti-IL-31 x anti-IL-4Ra antibody (NM26-2198) were used as controls.

[0332] Wistar Han rats were treated intravenously (IV) or subcutaneously (SC) at 3 mg / kg with v41790, v41791 or control antibody. Serum samples were collected at 15 minutes, 2 hrs, 8hrs, day 1, day 2, day 3, day 5, day 7, day 10, day 14 and day 21 post injection. Levels of antibody in the serum were quantified by a Meso Scale Discovery (MSD)-based PK assay using a biotinylated anti-human IgG-Fc antibody (Thermo Fisher Scientific, Waltham, MA) as the capture antibody and a SULFO-TAG™-conjugated anti-human IgG-Fc antibody (Fortis Life Sciences, Waltham, MA) as the detection antibody. The magnitude of electrochemiluminescence signal correlates with106IPTS / 200238736.2the antibody level and was read by a MESO SECTOR™ S 600MM instrument (Meso Scale Discovery, Rockville, MD) and analyzed by DISCOVERY WORKBENCH 4.0 Analysis Software (Meso Scale Discovery). PK parameters were generated from non-compartmental analysis (NCA) using the Phoenix WinNonlin™ software (Certara, Radnor, PA) (NCA calculation method: Linear log Trapezoidal; Half-life calculation method: Time range day 7-21).

[0333] The results are shown in Figs. 7 and 8, and Tables 11.1 and 11.2. Figs. 7 and 8 show the serum concentrations of the anti-IL-31 x anti-IL-4Ra bispecific antibody variants and controls. Tables 11.1 and 11.2 provide summaries of PK parameters for IV and SC dosing, respectively.107IPTS / 200238736.2Table 11.1: NCA of Total Serum Antibody Concentrations After 3 mg / kg Single IV Dose in Wistar Han Rats Half-life Cm ax Tmax Clast Tlast AUClast AUCINF obs Vz obs Cl obs Variant Rsq(day) (jrg / mL) (day) (jrg / mL) (day) (day* jrg / mL) (day* jrg / mL) (mL / kg) (mL / day / kg) v41790 0.995 10.1 124.3 0.01 4.6 21 279 345 126.0 8.7 v41791 0.944 8.8 142.2 0.01 5.6 21 410 480 79.1 6.3 NM26-2198 0.998 15.7 123.1 0.08 13.0 21 573 867 78.2 3.5Dupilumab 0.989 10.3 142.8 0.01 12.2 21 604 785 56.8 3.8Table 11.2: NCA of Total Serum Antibody Concentrations After 3 mg / kg Single SC Dose in Wistar Han Rats Bioavailability Variant Rsq Half-life Cmax Tmax Clast Tlast AUClast AUCINF obs Vz F obs Cl F obs(day) (fig / mL) (day) (jig / mL) (day) (day* fig / mL) (day* fig / mL) (mL / kg) (mL / day / kg) (%) V41790 0.997 9.6 8.6 5 2.7 21 116 154 271.0 19.5 44.5 V41791 0.998 8.5 9.7 5 2.8 21 130 165 223.0 18.2 34.3 NM26-2198 0.995 6.7 25.2 5 5.3 21 308 359 81.0 8.4 41.4Dupilumab 0.993 10.7 35.8 5 9.8 21 414 566 82.0 5.3 72.1EXAMPLE 12: ASSESSMENT OF HALF-LIFE OF BISPECIFIC ANTIBODY VARIANTS (IgGl AND IgG4 BACKBONE) IN A TG2 MOUSE MODEL

[0334] The pharmacokinetics (PK) of the anti-IL-31 x anti-IL-4Ra bispecific antibody variants v41543, v41544, v41789, v41790 and v41791 (see Tables 3.1 and 4.1), were evaluated in Tg32 or Tg32-SCID mice as described below. An anti-IL-4Ra monospecific antibody (dupilumab), an anti -IL-31 monospecific antibody (BMS-981164) and a tetravalent, bispecific anti-IL-31 x anti-IL-4Ra antibody (NM26-2198) were used as controls.

[0335] Tg32 or Tg32-SCID mice (Jackson Laboratories, Bar Harbor, ME) were injected intravenously with 5.0 mg / kg test antibody. Serum samples were collected at 10 minutes (or 1 hour), 24 hours, day 7, day 11, day 14, day 18 (or day 19) and day 21 post-injection. The levels of antibody in the serum were quantified by a Meso Scale Discovery (MSD)-based PK assay using a biotinylated anti-human IgG-Fc antibody (Thermo Fisher Scientific, Waltham, MA) as the capture antibody and a SULFO-TAG™-conjugated anti-human IgG-Fc antibody (Fortis Life Sciences, Waltham, MA) as the detection antibody. The magnitude of electrochemiluminescence signal correlates with the antibody level and was read by the MESO SECTOR™ S 600MM instrument (Meso Scale Discovery, Rockville, MD) and analyzed by the DISCOVERY WORKBENCH 4.0 Analysis Software (Meso Scale Discovery). PK parameters were generated from noncompartmental analysis (NCA) using the Phoenix WinNonlin™ software (Certara, Radnor, PA) (NCA calculation method: Linear log Trapezoidal; Half-life calculation method: Time range day 7-21).

[0336] The results are shown in Figs. 9 and 10A-D, and Tables 12.1 and 12.2. Figs. 9 and 10A-D show the serum concentrations of the anti-IL-4Ra x anti -IL-31 antibody variants and controls. Tables 12.1 and 12.2 provide summaries of PK parameters.109IPTS / 200238736.2Table 12.1: NCA of Total Serum Antibody Concentrations After 5 mg / kg Single IV Dose in Tg32 Mice Half-life Cm ax Tmax Clast Tlast AUClast AUCINF obs Vz obs Cl obs Variant Rsq(day) (jrg / ml) (day) (jig / ml) (day) (day* jrg / mL) (day* jrg / mL) (mL / kg) (mL / day / kg) v41543 0.997 5.3 129.8 0.04 2.7 21 344 365 104.1 13.7 v41544 0.932 5.2 242.0 0.04 4.8 21 601 637 58.7 7.8Dupilumab 0.920 7.1 183.4 0.04 23.2 21 1412 1651 31.4 3.1Table 12.2: NCA of Total Serum Antibody Concentrations After 5 mg / kg Single IV Dose in Tg32-SCID MiceHalf-life Cm ax Tmax Clast Tlast AUClast AUCINF obs Vz obs Cl obs V ariant Rsq(day) (jig / mL) (day) (jrg / mL) (day) (day* jrg / mL) (day* jrg / mL) (mL / kg) (mL / day / kg) v41544 0.966 6.0 204.7 0.01 6.4 21 644 699 61.5 7.2 v41789 0.999 4.6 351.2 0.01 6.8 21 918 964 34.7 5.2 v41790 0.959 8.4 301.9 0.01 13.2 21 837 998 61.0 5.0 v41791 0.984 5.6 233.7 0.01 13.3 21 1047 1154 35.0 4.3 Dupilumab 0.926 8.5 138.3 1.00 25.7 21 1527 1842 33.3 2.7 BMS- 0.938 6.2 210.5 0.01 10.5 21 1048 1142 39.2 4.4 981164NM26-2198 0.981 7.2 231.0 0.01 21.5 21 1275 1500 34.9 3.3EXAMPLE 13: IN VIVO EFFICACY STUDY OF A BISPECIFIC ANTIBODY VARIANT (IgG4 BACKBONE) IN A CHRONIC HOUSE DUST MITE MOUSE MODEL

[0337] The anti-inflammatory activity of the anti -IL-31 x anti-IL-4Ra antibody variant v41791 (see Table 4.1) was assessed in vivo in an acute house dust mite (HDM) mouse model as described below. The anti-IL-4Ra monospecific antibody dupilumab was used as a positive control, and an anti-hemagglutinin (HA) IgG4 antibody and no HDM treatment were used as negative controls. The experiments were run by GemPharmatech Co., Ltd. (Nanjing, China).

[0338] Briefly, C57B1 / 6 background human IL-4 / IL-4Ra knock in, mouse IL-4 / IL-4Ra knockout mice (Biocytogen, Waltham, MA) were treated with 50 ug of HDM or saline intranasally under isofluorane anesthesia three times weekly for 4 consecutive weeks. Three days prior to initiation of HDM treatment, mice were treated with test antibody or saline solution at doses of 1, 3, 10, or 25 mg / kg by subcutaneous injection twice weekly. Body weights and the general health and welfare of animals were monitored three times a week. 28- or 29-days post HDM treatment initiation, mice were sacrificed. Serum was collected for detection of circulating total IgE levels by ELISA assay. Lung tissue was divided for analysis of lung immune infiltration and cytokine levels. Lung homogenates were analyzed for cytokine levels of human IL-4 by ELISA. For the immunophenotyping, mice were injected with an anti-CD45 antibody conjugated to Brilliant Violet™ 605 dye 5 minutes prior to sacrifice to differentiate the circulating versus tissue resident cells. Immune cells were identified by flow cytometry using the cellular surface markers: CD45, CD1 lb, Ly6G, Ly6C, CD3, CD4, CD8, ST2, CD19, MERTK and Siglec-F.

[0339] The results are shown in Figs. 11A-D. As shown in Fig. 11A, treatment with 25 mg / kg v41791 decreased serum levels of IgE to levels similar to those observed in the no HDM control group. The IgE level reduction was similar to that observed with 25 mg / kg and 10 mg / kg dupilumab treatment.

[0340] To specifically monitor the immune response in the lung, human IL-4 (hIL-4) levels were quantified at endpoint. As shown in Fig. 11B, treatment with 25 mg / kg v41791 decreased levels of hIL-4 to levels similar to those observed in the no HDM control group. This decrease was also observed in mice treated with 25 mg / kg and 10 mg / kg dupilumab.111IPTS / 200238736.2

[0341] In addition to monitoring hIL-4 levels in the lung, immune infiltrate was characterized following treatment. As shown in Fig. 11C, tissue resident eosinophils were decreased in number in mice treated with 25 mg / kg v41791 compared to the negative control antibody group. Furthermore, as shown in Fig. 11D, mice treated with 25 mg / kg v41791 had an increased number of alveolar macrophages in the lung compared to the negative control antibody group. Both the changes in eosinophil number and alveolar macrophage number were also observed in mice treated with 25 mg / kg or 10 mg / kg dupilumab.

[0342] The dose response difference observed between v41791 and dupilumab in the levels of serum IgE, lung hIL-4 and lung infdtrate may be attributed to the bivalency of dupilumab compared to the monovalency of the anti-IL-4Ra arm of the v41791 bispecific antibody. Overall, the data shown in Figs. 11A-D indicate that the anti-IL-31 x anti-IL-4Ra bispecific antibody, v41791, is capable of regulating IL-4 and IL-13 linked immune processes associated with inflammatory disease.EXAMPLE 14: PHARMACOKINETIC STUDY OF A BISPECIFIC ANTIBODY VARIANT (IgG4 BACKBONE) IN CYNOMOLGUS MONKEYS

[0343] Pharmacokinetics (PK) of the anti -IL-31 x anti-IL-4Ra antibody variant v41791 (see Table 4.1) were assessed in vivo in cynomolgus monkeys as described below. The tetravalent, bispecific anti-IL-31 x anti-IL-4Ra antibody NM26-2198 was used as a control antibody.

[0344] Two cynomolgus monkeys were injected intravenously with 10 mg / kg of test antibody or an equal volume of saline. Blood was collected pre-dose, and Omin (after infusion), 3h, 8h, 24h, 48h, 96h, 168h, 240h, 336h, 408h, 504h, 576h, 672h, 744h, 840h, 912h, and 984h post injection and was analyzed for serum PK by an anti-human IgG ELISA assay. Blood samples were collected throughout the study to monitor clinical chemistry, hematology and coagulation parameters.

[0345] Overall, the study showed that treatment with v41791 was well tolerated in monkeys. No adverse responses were observed throughout the study. Analysis for serum PK showed that v41791 had antibody-like PK similar to that observed with NM26-2198 treatment (see Fig. 12).112IPTS / 200238736.2EXAMPLE 15: PREPARATION AND CHARACTERIZATION OF ANTI-IL-31 x ANTI-IL-4Ra BISPECIFIC ANTIBODIES (IgG4 BACKBONE) #2

[0346] Additional anti-IL-31 x anti-IL-4Ra bispecific antibody variants were prepared using engineered versions of the IL-4Ra paratopes described in Example 4. The engineered versions of the IL-4Ra paratopes included additional mutations to modulate the total charge and / or charge asymmetry in the VH and VL domains, with a view to improving the pharmacokinetic properties of the bispecific antibody variants. The engineered IL-4Ra paratopes are described in Table 15.1. The CDR sequences of the engineered IL-4Ra paratopes are shown in Fig. 19 (Table C2) and the VH and VL sequences are shown in Fig. 20 (Table D2).

[0347] Bispecific antibody variants that are monovalent for each antigen were prepared in a format in which both the IL-31 and IL-4Ra antigen binding domains are Fab domains and in which the Fc region is a heterodimeric IgG4 Fc region as described in Example 4. The heavy chains (heavy chain A and heavy chain B) and light chains (light chain A and light chain B) of each of the bispecific antibody variants are described in Table 15.2. All bispecific antibody variants comprised the HetFabl, HetFc and S228P mutations described in Example 4. In addition, certain of the bispecific antibody variants further comprise the YTE mutations (M252Y / S254T / T256E) as noted in Table 15.2.Table 15.1: IL-4Ra Paratope Mutational Designs*Design Name Mutations in VH Domain Mutations in VL Domain — V38597-Y54A Y54A None1 v38597-Mut09 Q1E, K13Q, K23A, Q61D, R18P, K42Q, K45Q, S60D, K62Q, K73T Q79E2 v38597-Mutl8 None R18S, K42Q, S60D3 v38597-Mut29 Q1E, G65E S9E, S12E4 v38597-Mut42 Q61D, K73E K42E5 v38597-Mut57 Q61D, K73E R18T, K42E6 v38597-Mut65 K73E Q3E, R18T, K42E7 v38597-Mut66 K12A, K73E Q55E, I106E8 v38597-Mut88 K64Q, K73E K42E113IPTS / 200238736.2* All amino acid positions are numbered using AbM numbering.Table 15.2: Description of Anti-IL-31 x Anti-IL-4Ra Bispecific Antibody Variants (IgG4)Light Chain Light Chain Heavy Chain A Heavy Chain BVariant Design A B Fc (anti-IL-4Ra) (anti-IL-31)(anti-IL-4Ra) (anti-IL-31) v38597 VH- v36542 VH- V38597 VL- v36542 VL- Het Fc, CHl-HetFabl CHl-HetFablv41790 None CL-HetFabl CL-HetFabl IgG4,HCA-CH3- HCB-CH3- LCA LCB YTEHetFcA HetFcBV38597-Y54A v36542 VH- V38597 VL- v36542 VL- Het Fc, VH-CH1- CHl-HetFablv41791 Y54A CL-HetFabl CL-HetFabl IgG4,HetFabl HCA- HCB-CH3- LCA LCB YTE CH3-HetFcA HetFcBv38597-Mut09- v36542 VH- v38597- v36542 VL- Het Fc, 1 + Y54A VH-CH1- CHl-HetFabl Mut09 VL- v43181 CL-HetFabl IgG4,Y54A HetFabl HCA- HCB-CH3- CL-HetFablLCB YTE CH3-HetFcA HetFcB LCAv38597-Mutl8- v36542 VH- v38597- v36542 VL- Het Fc, 2 + Y54A VH-CH1- CHl-HetFabl Mutl8 VL- v43182 CL-HetFabl IgG4,Y54A HetFabl HCA- HCB-CH3- CL-HetFablLCB YTE CH3-HetFcA HetFcB LCAv38597-Mut29- v36542 VH- v38597- v36542 VL- Het Fc, 3 + Y54A VH-CH1- CHl-HetFabl Mut29 VL- v43183 CL-HetFabl IgG4,Y54A HetFabl HCA- HCB-CH3- CL-HetFablLCB YTE CH3-HetFcA HetFcB LCAv38597-Mut42- v36542 VH- v38597- v36542 VL- Het Fc, 4 + Y54A VH-CH1- CHl-HetFabl Mut42 VL- v43184 CL-HetFabl IgG4,Y54A HetFabl HCA- HCB-CH3- CL-HetFablLCB YTE CH3-HetFcA HetFcB LCAv38597-Mut57- v36542 VH- v38597- v36542 VL- Het Fc, 5 + Y54A VH-CH1- CHl-HetFabl Mut57 VL- v43185 CL-HetFabl IgG4,Y54A HetFabl HCA- HCB-CH3- CL-HetFablLCB YTE CH3-HetFcA HetFcB LCAv38597-Mut65- v36542 VH- v38597- v36542 VL- Het Fc, 6 + Y54A VH-CH1- CHl-HetFabl Mut65 VL- v43186 CL-HetFabl IgG4,Y54A HetFabl HCA- HCB-CH3- CL-HetFablLCB YTE CH3-HetFcA HetFcB LCA114IPTS / 200238736.2Light Chain Light Chain Heavy Chain A Heavy Chain BVariant Design A B Fc (anti-IL-4Ra) (anti-IL-31)(anti-IL-4Ra) (anti-IL-31) v38597-Mut66- v36542 VH- v38597- v36542 VL- Het Fc, 7 + Y54A VH-CH1- CHl-HetFabl Mut66 VL- v43187 CL-HetFabl IgG4,Y54A HetFabl HCA- HCB-CH3- CL-HetFablLCB YTE CH3-HetFcA HetFcB LCAv38597-Mut88- v36542 VH- v38597- v43188 v36542 VL- Het Fc,8 + Y54A VH-CH1- CHl-HetFabl Mut88 VL- CL-HetFabl IgG4, Y54A HetFabl HCA- HCB-CH3- CL-HetFablLCB YTE CH3-HetFcA HetFcB LCAv38597-Mut09 v36542 VH- v38597- v36542 VL- Het Fc, VH-CH1- CHl-HetFabl Mut09 VL- v43189 1 CL-HetFabl IgG4,HetFabl HCA- HCB-CH3- CL-HetFablLCB YTE CH3-HetFcA HetFcB LCAv38597-Mut42 v36542 VH- v38597- v36542 VL- Het Fc, VH-CH1- CHl-HetFabl Mut42 VL- v43190 4 CL-HetFabl IgG4,HetFabl HCA- HCB-CH3- CL-HetFablLCB YTE CH3-HetFcA HetFcB LCAv38597-Mut57 v36542 VH- v38597- v36542 VL- Het Fc, VH-CH1- CHl-HetFabl Mut57 VL- v43191 5 CL-HetFabl IgG4,HetFabl HCA- HCB-CH3- CL-HetFablLCB YTE CH3-HetFcA HetFcB LCAv38597-Mut65 v36542 VH- v38597- v36542 VL- Het Fc, VH-CH1- CHl-HetFabl Mut65 VL- v43192 6 CL-HetFabl IgG4,HetFabl HCA- HCB-CH3- CL-HetFablLCB YTE CH3-HetFcA HetFcB LCAv38597-Mut88- v36542 VH- v38597- v36542 VL- Het Fc, VH-CH1- CHl-HetFabl Mut88 VL- v43193 8 CL-HetFabl IgG4,HetFabl HCA- HCB-CH3- CL-HetFablLCB YTE CH3-HetFcA HetFcB LCA15.1 Preparation of Bispecific Anti-IL-31 x Anti-IL-4Ra Antibody Variants

[0348] The antibody variants shown in Table 15.2 were expressed in ExpiCHO™ cells at a 200 mL culture volume. Generally, the final gene products were sub-cloned into the mammalian expression vector pTT5 (NRC-BRI, Canada) or other mammalian expression vector. Cells were transfected in exponential growth phase (1.5 to 2 million cells / mL) with aqueous 1 mg / mL 25 kDa polyethylenimine (PEI) using a PEEDNA ratio of 2.5: 1. DNA was transfected at an optimal DNA 115IPTS / 200238736.2ratio of the heavy chain A (HC-A), heavy chain B (HC-B), and light chain A (LC-A) and light chain B (LC-B) that allows for heterodimer formation (for example, HC-A: HC-B: LC-A: LC-B ratio of 15:15:35:35). Transfected cells were harvested after 5-6 days and the culture medium collected after centrifugation at 4000 rpm and clarification using a 0.45 pm filter. The clarified culture medium was loaded onto a MabSelect™ SuRe™ (GE Healthcare) protein A column and washed with 10 column volumes of PBS buffer at pH 7.2 - 7.4. The antibody variant was eluted with 10 column volumes of 0.1 M citrate buffer at pH 3.6 and the pooled fractions containing the antibody variant were neutralized with 1 M TRIS at pH 9. The amount of antibody variant was then quantified based on A280 nm (NanoDrop™ Spectrophotometer; Thermo Fisher Scientific).

[0349] The antibody variants were further purified by gel filtration chromatography using a Superdex™ 200 HiLoad™ 16 / 600 200pg column (GE Healthcare) via an AKTA Pure chromatography system at a flowrate of 1 mL / min with 20 mM histidine, 150 mM sodium chloride, pH 6.0 buffer. Fractions of eluted antibody variant were collected based on absorbance at A280 nm and the fractions were assessed by non-reducing and reducing CE-SDS or High Throughput Protein Express assay using Caliper LabChip™ GXII (Perkin Elmer, Waltham, MA) and UPLC-SEC using a Waters Acquity™ BEH200 SEC column (2.5 mL, 4.6x150 mm, stainless steel, 1.7 pm particles) (Waters Corporation, Mississauga, ON). Fractions corresponding to the purified antibody variants were collected, buffer exchanged into 20 mM histidine, pH 6.0 using a Zeba™ Spin desalting column (Thermo Fisher Scientific), concentrated to ~1 mg / mL and stored at -80°C.

[0350] Endotoxin levels were determined by the limulus amebocyte lysate (LAL) assay using the Endosafe™ Portable Test System (PTS) (Charles River Laboratories, Wilmington, MA). Antibody variants were quantified based on A280 nm absorbance ((NanoDrop™ Spectrophotometer) after protein A and SEC purification. UPLC-SEC was performed using a Waters Acquity™ BEH200 SEC column (2.5 mL, 4.6x150 mm, stainless steel, 1.7 pm particles) (Waters Corporation, Mississauga, ON) set to 30°C or 25°C and mounted on a Waters Acquity™ UPLC H-Class Bio system with a photodiode array (PDA) detector. Run times were 7 min with a total volume per injection of 5 pL and a running buffer of 200mM potassium phosphate pH 7.0 at 0.4 mL / min. Elution was monitored by UV absorbance in the range 210-500 nm, and chromatograms were extracted at 280 nm. Peak integration was performed using Agilent OpenLab software (Agilent Technologies, Inc., Santa Clara, CA).116IPTS / 200238736.2

[0351] The apparent purity and yield of the final antibody variant was estimated by UPLC-SEC and LC / MS as described in International Patent Publication No. WO 2015 / 109131. All antibody variants expressed and five of the variants were purified to >80% heterodimer purity without contaminating homodimers as shown in Table 15.3 below.Table 15.3: Post Purification Yield and Purity for Bispecific Anti-IL-31 x Anti-IL-4Ra Antibody Variants (20 mM Histidine pH 6.0 Buffer)Titer Yield MonomerVariant(mg / mL) (mg / L) Purity (%)*v43181 155.7 7.3 91.3v43182 164.1 4.6 91.2v43184 394 0.9 35.5v43185 289.2 0.4 38.5v43186 318.7 0.6 32.2v43187 130.3 0 0v43188 400.7 7.2 92.5v43189 9.79 0 0v43190 400.2 5 83.5v43191 238 1.6 69.4v43192 257.7 0.2 19.4v43193 173.8 8.5 95.3* As determined by HPLC-SEC (size-exclusion chromatography HPLC)15.3 Functional Screening of the Engineered Anti-IL-4Ra Paratope in the Bispecific Anti-IL-31 x Anti-IL-4Ra Antibody Variants

[0352] To determine the impact of the bispecific antibody variants on IL-4 / IL- 13 activation of the STAT6 pathway, selected variants (v43181, v43182, v43188, v43190 and v43193) were assessed for inhibition of IL-4 / IL-13 mediated production of STAT6 inducible secreted embryonic alkaline phosphatase (SEAP) reporter in HEK-Blue™ IL-4 / IL-13 cells as described below. An anti-IL-4Ra antibody (dupilumab), a tetravalent anti-IL-4Ra x IL-31 bispecific antibody (NM26-117IPTS / 200238736.22198), an anti-RSV IgGl FcKO antibody (v39982) and an IgG4 isotype antibody (v42104) were used as controls.

[0353] Briefly, test articles were serially diluted starting at 20 000 pM in DMEM + 10% heat-inactivated fetal bovine serum (ThermoFisher Scientific, Waltham, MA) and plated into a 384-well black flat bottom assay plate. 125 pM IL-4 or 500-5000 pM IL- 13 (R& D Systems, Minneapolis, MN) was added, followed by 5000-12500 HEK-Blue™ IL-4 / IL-13 cells (InvivoGen, San Diego, CA). After 24 hr at 37°C, 5% CO2, SEAP production was assessed by incubating supernatant with QUANTI-Blue™ solution (InvivoGen, San Diego, CA) and measuring OD620nm on Synergy™ plate reader (BioTek Instruments, Winooski, VT).

[0354] The results are shown in Figs. 13A-D. All tested bispecific antibody variants blocked IL-4 and IL- 13 mediated production of STAT6 inducible SEAP reporter in HEK-Blue™ IL-4 / IL-13 cells. Bispecific antibody variants including mutational variations of the IL-4Ra paratope showed a decreased level of inhibition compared to the parental variant (v41791). EXAMPLE 16: PREPARATION AND CHARACTERIZATION OF ANTI-IL-31 x ANTI-IL-4Ra BISPECIFIC ANTIBODIES (IgG4 BACKBONE) #3

[0355] Selected antibody variants from Example 15 were re-expressed in CHO3E7 cells at a 500 mL culture volume. In brief, cells were transfected in exponential growth phase (1.5 to 2 million cells / mL) with aqueous 1 mg / mL polyethyleneimine (PEI) using a PELDNA ratio of 2.5: 1. DNA was transfected at an optimal DNA ratio of the heavy chain A (HC-A), heavy chain B (HC-B), and light chain A (LC-A) and light chain B (LC-B) that allows for heterodimer formation (for example, HC-A: HC-B: LC-A: LC-B ratio of 15:15:53:17 or 15:15:35:35). Transfected cells were harvested after 5-6 days and the antibody variant recovered from the culture medium as described in Example 15 and quantified based on A280 nm.

[0356] The antibody variants were further purified by gel filtration chromatography using one or two Superdex™ 200 HiLoad™ 26 / 600 200pg columns (Cytiva) in tandem via an AKTA Pure chromatography system following the procedure described in Example 15. Endotoxin levels were determined by the limulus amebocyte lysate (LAL) assay using the Endosafe™ Portable Test System (PTS) (Charles River Laboratories, Wilmington, MA). Antibody variants were quantified118IPTS / 200238736.2based on A280 nm absorbance ((NanoDrop™ Spectrophotometer) after protein A and SEC purification. UPLC-SEC was performed using a Waters Acquity™ BEH200 SEC column (2.5 mL, 4.6x150 mm, stainless steel, 1.7 pm particles) (Waters Corporation, Mississauga, ON) set to 30°C or 25 °C and mounted on a Waters Acquity™ UPLC H-Class Bio system with a photodiode array (PDA) detector. Run times were 7 min with a total volume per injection of 5 pL and a running buffer of 200mM potassium phosphate pH 7.0 at 0.4 mL / min. Elution was monitored by UV absorbance in the range 210-500 nm, and chromatograms were extracted at 280 nm.

[0357] The apparent purity and yield of the final antibody variant was estimated by UPLC- SEC and LC / MS as generally described in International Patent Publication No. WO 2015 / 109131. All antibody variants expressed and were purified to >93% heterodimer purity without contaminating homodimers as shown in Table 16.1.Table 16.1: Post Purification Yield and Purity for Bispecific Anti-IL-31 x Anti-IL-4Ra Antibody Variants (20 mM Histidine pH 5.5 Buffer)Monomer Heterodimer Variant Titer (mg / L) Yield (mg / L)Purity (%)' Purity (%)2v43181 84 35.8 99.9 96.7 v43182 371 31 93.8 97.2 v43183 266 48.8 95.9 95.8 v43184 231 484 98.5 96.0 v43185 429 208 98.1 97.0 v43186 194 71.4 97.5 96.9 v43187 163 17.8 94.2 97.4 v43188 254 58.6 97.1 97.01As determined by UPLC-SEC2As determined by LC / MS intensity119IPTS / 200238736.216.1 Thermal Stability of Bispecific Antibody Variants by Differential Scanning Fluorimetry (DSF)

[0358] The thermal stability of the bispecific anti-IL-31 x anti-IL-4Ra antibody variants was assessed by Differential Scanning Fluorimetry (DSF). All DSF experiments were carried out using a CFX96 Touch™ Real-Time PCR instrument (BioRad Laboratories, Inc., Hercules, CA) as described in Example 3 (Section 3.2.1).

[0359] The results are shown in Table 16.2, which shows the maximum melting temperatures (Tm) for each of the peaks in the thermograms of the tested bispecific antibody variants.Table 16.2: Thermal Stability of Bispecific Antibody VariantsVariant Tml (°C) Tm2 (°C)v43181 55 72.5v43182 55.5 72.5v43183 55.5 72.75v43184 55.5 72.5v43185 55.5 72.75v43186 55.5 72.5v43187 55 72.5v43188 55.5 72.516.2 Stability of Bispecific Antibody Variants under an Accelerated Stress

[0360] Stability of the bispecific antibody variants v43184, v43185 and v43187 was tested using an accelerated stress test as described below. Purity of the bispecific antibody variants pre-and post-treatment was assessed by UPLC-SEC.

[0361] Bispecific antibody variants were incubated at 40 °C for 14 and 28 days using a protein concentration of 5 mg / mL in 20 mM histidine, pH 5.5 buffer. The results are shown in Table 16.3. All bispecific antibody variants tested showed only a minimal change in purity after the 28-day incubation.120IPTS / 200238736.2Table 16.3: Purity (%) of Bispecific Antibody Variants After 28 Days at 40 °C% Monomer % HMWs % LMWs VariantDay 0 Day 28 Day 0 Day 28 Day 0 Day 28 v43184 98.93 96.36 1.07 1.08 0.00 2.56 v43185 97.92 96.26 2.08 1.61 0.00 2.13 v43187 94.24 91.85 4.20 2.27 1.55 5.88EXAMPLE 17: FUNCTIONAL CHARACTERIZATION OF BISPECIFIC ANTIBODY VARIANT WITH ENGINEERED 11 -4 Ro PARATOPE (IgG4 BACKBONE)

[0362] The impact of the anti -IL-31 x anti-IL-4Ra bispecific antibody variant v43184 comprising an engineered IL-4Ra paratope (see Table 15.2) on IL-4 / IL-13 activation ofthe STAT6 signalling pathway was assessed by measuring inhibition of IL-4 / IL-13 mediated production of the STAT6 inducible secreted embryonic alkaline phosphatase (SEAP) reporter in HEK-Blue™ IL-4 / IL-13 cells following the protocol described in Example 6. The impact of the anti-IL-31 x anti-IL-4Ra bispecific antibody variants on IL-31 activation of the STAT5 signalling pathway was assessed by measuring inhibition of IL-31 mediated production of the STAT5 inducible SEAP reporter in HEK-Blue™ IL-31 cells as described below. The anti-IL-31 x anti-IL-4Ra bispecific antibody variant v41791 (see Table 4.1 ), a tetravalent, bispecific anti -IL-31 x anti-IL-4Ra antibody based on NM26-2198 (Numab Therapeutics), a bivalent anti-IL-31 antibody (BMS-981164), a bivalent anti-IL-4Ra antibody (dupilumab) and an IgG4 isotype antibody (v42104) were also included in the assay.

[0363] Briefly, test articles were serially diluted starting at a concentration of 20 nM in DMEM + 10% heat-inactivated fetal bovine serum (ThermoFisher Scientific, Waltham, MA) and plated into a 384-well black flat bottom assay plate. 3 pM IL-31 (R& D Systems, Minneapolis, MN) was added, followed by 12500 HEK-Blue™ IL-31 cells (InvivoGen, San Diego, CA). After 24 hr at 37°C, 5% CO2, SEAP production was assessed by incubation of supernatant with QUANTI-Blue™ solution (InvivoGen, San Diego, CA) and measuring OD620nm on a Synergy™ plate reader (BioTek Instruments, Inc., Winooski, VT).

[0364] The results for IL-4 and IL- 13 mediated production of STAT6 inducible SEAP reporter in HEK-Blue™ IL-4 / IL-13 cells are shown in Figs.23A & B. Both bispecific antibody 121IPTS / 200238736.2variants showed similar inhibitory activity to each other and to both the bivalent anti-IL-4Ra antibody control (dupilumab) and the tetravalent, bispecific anti-IL-31 x anti-IL-4Ra antibody NM26-2198. The anti-IL-31 antibody control (BMS-981164) and the IgG4 isotype control showed no activity as expected.

[0365] The results for IL-31 mediated production of STAT6 inducible SEAP reporter in HEK-Blue™ IL-31 cells are shown in Fig.23C. Both bispecific antibody variants showed similar inhibitory activity to each other and similar or better inhibitory activity compared to the bivalent anti-IL-31 antibody control (BMS-981164) and the tetravalent, bispecific anti-IL-31 x anti-IL-4Ra antibody NM26-2198. The anti-IL-4Ra antibody control (dupilumab) and the IgG4 isotype control showed no activity as expected.EXAMPLE 18: PHARMACOKINETIC / PHARMACODYNAMIC STUDY OF A BISPECIFIC ANTIBODY VARIANT (IgG4 BACKBONE) IN CYNOMOLGUS MONKEYS

[0366] Pharmacokinetic (PK) and pharmacodynamic (PD) parameters for the anti -IL-31 x anti-IL-4Ra antibody variant v44927 were assessed in vivo in naive cynomolgus monkeys as described below. v44927 is identical to v43184 (see Table 15.2) except that v44927 includes a lysine residue at the C-terminus of the heavy chains.

[0367] The test group, consisting of two cynomolgus monkeys (one female and one male), was injected intravenously with 40 mg / kg of v44927. A control group of two cynomolgus monkeys was injected intravenously and subcutaneously with equal volumes of saline. All monkeys were injected once a week for four weeks (QWx4). Injections were performed on days 1, 8, 15, and 22.

[0368] Blood was collected at each pre-dose and 5min after each infusion, as well as 3h, 8h, 24h, 48h, 96h, 507h, 512h, 528h, 552h, 600h, and 672h post first injection and analyzed for serum PK by an anti-human IgG ELISA assay. Serum IgE levels were monitored by ELISA assay at each pre-dose and at 672h post first dose (study termination). Blood samples were collected throughout the study to monitor clinical chemistry, hematology and coagulation parameters.

[0369] Treatment with v44927 was well tolerated in monkeys. No adverse responses were observed throughout the study. Analysis for serum PK of the test article-injected monkeys showed 122IPTS / 200238736.2that v44927 had antibody-like PK, with a modest decrease in AUC after the second, third, and fourth administration and differing outcomes per test animal (see Fig. 24).

[0370] To determine the activity of the IL-31 x IL-4Ra bispecific antibody, serum IgE levels were monitored. IL-4 signalling triggers class switch recombination (CSR) in B cells to produce IgE, a critical mediator that induces mast cell degranulation and the release of histamine during allergic reactions. Blocking IL-4 signalling should result in a decrease in serum IgE levels. As the monkeys used in this study are naive (i.e. are not showing allergic inflammation symptoms), the initial IgE serum levels will be low. Thus, changes in serum IgE levels are expected to be modest after treatment with the bispecific antibody blocking IL-4Ra.

[0371] Compared to saline-administered monkeys in the control group, treatment with v44927 once a week for four weeks (QWx4) transiently decreased serum IgE levels between days 8 and 15 (see Fig. 25). A reduction of serum IgE levels has been similarly observed in patients treated with IL-4Ra targeting monoclonal antibody therapies indicating that v44927 can function in a similar manner to clinically approved IL-4Ra targeting monoclonal antibodies.EXAMPLE 19: OXAZOLONE-INDUCED ATOPIC DERMATITIS EFFICACY STUDY OF A BISPECIFIC ANTIBODY VARIANT (IgG4 BACKBONE)

[0372] The anti-IL-31 x anti-IL-4Ra antibody variant v44927 was assessed in vivo in an oxazolone-induced atopic dermatitis mouse model using hIL3 l / hIL3 lRA / hOSMR / hIL4 / hIL4RA C57BL / 6mice. v44927 is identical to v43184 (see Table 15.2) except that v44927 includes a lysine residue at the C-terminus of the heavy chains. Body weight, ear thickness and human IL-4 (hlL-4) serum cytokine levels for the mice were assessed as described below.

[0373] Three groups of hIL3 l / hIL3 lRA / hOSMR / hIL4 / hIL4RA C57BL / 6 mice were used with six mice allocated to each study group. The first group was treated subcutaneously with 25 mg / kg v44927. The second group was treated with 25 mg / kg isotype control antibody. The third group of animals was left untreated. The test article and isotype control administrations were performed twice a week for a total of eight doses, on days -1, 2, 6, 9, 13, 16, 20 and 23. Induction of atopic dermatitis was initiated by application of 0.8% oxazolone solution (in a 4: 1 mixture of acetone and olive oil) onto the right ear of each antibody-treated animal on day 0, followed by123IPTS / 200238736.2application of 0.4% oxazolone solution on days 7, 9, 11, 13, 16, 20 and 23. The third group of untreated animals received a vehicle solution only (4: 1 mixture of acetone and olive oil).

[0374] Body weight and ear thickness (measured by caliper) were recorded on days -2, 0, 7, 9, 11, 14, 16, 18, 21, 23, 25 and 26. Blood was collected on days 9 and 26 to determine hIL-4 serum cytokine concentrations using the Mesoscale Discovery (MSD) method.

[0375] Body weight of the treated mice did not change over the course of the experiment (see Fig. 26A) indicating that the oxazolone-mediated interventions, as well as test article (v44927) and isotype administrations, were well tolerated with no adverse events observed throughout the study.

[0376] Compared to isotype-treated animals, treatment with v44927 significantly reduced ear thickness on days 14, 16, and 18 after initial oxazolone sensitization on day 0 (see Fig. 26B). In addition, compared to isotype-treated mice, treatment with v44927 significantly diminished hIL-4 serum cytokine concentrations on days 9 and 26 after initial oxazolone sensitization on day 0 (see Fig. 26C). The hIL-4 serum cytokine concentrations in animals treated with v44927 were similar to those detected in vehicle-treated control mice.

[0377] The disclosures of all patents, patent applications, publications and database entries referenced in this specification are hereby specifically incorporated by reference in their entirety to the same extent as if each such individual patent, patent application, publication and database entry were specifically and individually indicated to be incorporated by reference

[0378] Modifications of the specific embodiments described herein that would be apparent to those skilled in the art are intended to be included within the scope of the following claims.124IPTS / 200238736.2SEQUENCE TABLES Table G: Clone Numbers for VariantsClone NumbersVariantID Hl H2 LI L2 33559 24891 24891 24892 24892 34090 25272 25272 25262 25262 34731 25900 25900 25259 25259 34734 25903 25903 25259 25259 34735 25904 25904 25259 25259 34736 25905 25905 25259 25259 34737 25906 25906 25259 25259 34738 25907 25907 25259 25259 34739 25908 25908 25259 25259 34740 25909 25909 25259 25259 34741 25910 25910 25259 25259 36535 27394 27394 27392 27392 36539 27396 27396 27392 27392 36540 27396 27396 27391 27391 36541 27395 27395 27389 27389 36542 27395 27395 27390 27390 36545 27397 27397 27389 27389 36974 27813 27813 27390 27390 38681 29426 29418 29425 29417 38727 29420 29414 29424 29412 39439 29448 29882 29424 29412 39441 29440 29882 29424 29412 41543 31575 31576 31317 31580 41544 31577 31578 31317 31580 41789 31743 31745 31317 31580 41790 31744 31746 31317 31580 41791 31747 31746 31317 31580 43181 32864 32191 32190 29417 43182 32861 32191 32139 29417 43183 32860 32191 32134 29417 43184 32783 32191 32147 29417 43185 32783 32191 32151 29417 43186 32862 32191 32153 29417 43187 32863 32191 32154 2941743188 32781 32191 32147 29417125IPTS / 200238736.2Clone NumbersVariantID Hl H2 LI L2 43189 32133 32191 32190 29417 43190 32104 32191 32147 29417 43191 32104 32191 32151 29417 43192 32102 32191 32153 2941743193 32097 32191 32147 29417126IPTS / 200238736.2Table H: Clone SequencesClone Region Sequence SEQ ID ID NO24891 Full QSLEESGGRLVTPGGSLTLTCTVSGIDLSSYFMSWVRQAPGKGLE 122YIGTISTGGNTYYASWVKGRFTISKTSTTVDLKITSPTTEDTATYFC ARGWLRDYLDLWGQGTLVTISSASTKGPSVFPLAPSSKSTSGGTA ALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSV VTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCP APELLGGPSVFLFPPKPKDTLMISRTPEVTCVWDVSHEDPEVKFN WYVDGVEVHNAKTKPREEQYNSTYRWSVLTVLHQDWLNGKE YKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVS LTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSK LTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPG24892 Full QVLTQTPSPVSAAVGGTVSISCQASQSVYRENRLAWYQQKVGQP 123PKLLIYRASKLESGVPSRFSGSGSGTEFTLTISGVQCDDAATYYCA GGDSSGSDHAFGGGTEWVKRTVAAPSVFIFPPSDEQLKSGTASV VCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC25259 Full DIQMTQSPSTLSASVGDRVTITCRASQSVYRENRLAWYQQKPGK 124APKLLIYRASKLESGVPSRFSGSGSGTEFTLTISSLQPDDFATYYCA GGDSSGSDHAFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASW CLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSS TLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC25262 Full QVLTQTPSPVSAAVGGTVSISCQASQSVYRENRLAWYQQKVGQP 125PKLLIYRASKLESGVPSRFSGSGSGTEFTLTISGVQCDDAATYYCA GGDSSGSDHAFGGGTEWVKRTVAAPSVFIFPPSDEQLKSGTASV VCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC25272 Full QSLEESGGRLVTPGGSLTLTCTVSGIDLSSYFMSWVRQAPGKGLE 126YIGTISTGGNTYYASWVKGRFTISKTSTTVDLKITSPTTEDTATYFC ARGWLRDYLDLWGQGTLVTISSASTKGPSVFPLAPSSKSTSGGTA ALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSV VTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCP APEAAGGPSVFLFPPKPKDTLMISRTPEVTCVWSVSHEDPEVKFN WYVDGVEVHNAKTKPREEQYNSTYRWSVLTVLHQDWLNGKE YKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVS LTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSK LTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPG25900 Full QSLVESGGGLVKPGGSLRLSCAVSGIDLSSYFMSWVRQAPGKGLE 127YIGTISTGGNTYYASWVKGRFTISKDSSKNTVYLQMNSLKTEDTA VYYCARGWLRDYLDLWGQGTLVTVSSASTKGPSVFPLAPSSKST SGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGL YSLSSWTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTH TCPPCPAPEAAGGPSVFLFPPKPKDTLMISRTPEVTCVWSVSHED PEVKFNWYVDGVEVHNAKTKPREEQYNSTYRWSVLTVLHQDW LNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELT KNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPG127IPTS / 200238736.2Clone Region Sequence SEQ ID ID NO25903 Full QSLVESGGGLVKPGGSLRLSCAVSGIDLSSYFMSWVRQAPGKGLE 128YIGTISTGGNTYYASWVKGRFTISKTSTTVYLQMNSLKTEDTAVY YCARGWLRDYLDLWGQGTLVTVSSASTKGPSVFPLAPSSKSTSG GTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSL SSWTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCP PCPAPEAAGGPSVFLFPPKPKDTLMISRTPEVTCWVSVSHEDPEV KFNWYVDGVEVHNAKTKPREEQYNSTYRWSVLTVLHQDWLN GKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKN QVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFL YSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPG25904 Full QSLVESGGGLVKPGGSLRLSCAVSGIDLSSYFMSWVRQAPGKGLE 129YIGTISTGGNTYYASWVKGRFTISKTSTTVYLQINSLKTEDTAVYY CARGWLRDYLDLWGQGTLVTVS S ASTKGPS VFPL APS SKSTSGGT AALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSS WTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPC PAPEAAGGPSVFLFPPKPKDTLMISRTPEVTCWVSVSHEDPEVKF NWYVDGVEVHNAKTKPREEQYNSTYRWSVLTVLHQDWLNGK EYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQV SLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYS KLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPG25905 Full QSLVESGGGLVKPGGSLRLSCAVSGIDLSSYFMSWVRQAPGKGLE 130YIGTISTGGNTYYASWVKGRFTISKTSTTVYLQINSPKTEDTAVYY CARGWLRDYLDLWGQGTLVTISSASTKGPSVFPLAPSSKSTSGGT AALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSS WTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPC PAPEAAGGPSVFLFPPKPKDTLMISRTPEVTCWVSVSHEDPEVKF NWYVDGVEVHNAKTKPREEQYNSTYRWSVLTVLHQDWLNGK EYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQV SLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYS KLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPG25906 Full QSLVESGGGLVKPGGSLRLSCAVSGIDLSSYFMSWVRQAPGKGLE 131YIGTISTGGNTYYASWVKGRFTISKTSTTVYLQINSPKTEDTATYF CARGWLRDYLDLWGQGTLVTISSASTKGPSVFPLAPSSKSTSGGT AALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSS WTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPC PAPEAAGGPSVFLFPPKPKDTLMISRTPEVTCWVSVSHEDPEVKF NWYVDGVEVHNAKTKPREEQYNSTYRWSVLTVLHQDWLNGK EYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQV SLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYS KLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPG25907 Full QSLVESGGGLVKPGGSLRLSCAVSGIDLSSYFMSWVRQAPGKGLE 132YIGTISTGGNTYYASWVKGRFTISKTSTDVYLQMNSLKTEDTAVY YCARGWLRDYLDLWGQGTLVTVSSASTKGPSVFPLAPSSKSTSG GTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSL SSWTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCP PCPAPEAAGGPSVFLFPPKPKDTLMISRTPEVTCWVSVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRWSVLTVLHQDWLN128IPTS / 200238736.2Clone Region Sequence SEQ ID ID NO GKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKN QVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFL YSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPG25908 Full QSLVESGGGLVKPGGSLRLSCAVSGIDLSSYFMSWVRQAPGKGLE 133YIGTISTGGNTYYASWVKGRFTISKTSTGVYLQMNSLKTEDTAVY YCARGWLRDYLDLWGQGTLVTVSSASTKGPSVFPLAPSSKSTSG GTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSL SSWTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCP PCPAPEAAGGPSVFLFPPKPKDTLMISRTPEVTCWVSVSHEDPEV KFNWYVDGVEVHNAKTKPREEQYNSTYRWSVLTVLHQDWLN GKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKN QVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFL YSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPG25909 Full QSLVESGGGLVKPGGSLRLSCAVSGIDLSSYFMSWVRQAPGKGLE 134YIGTISTGGNTYYASWVKGRFGISKTSTTVYLQMNSLKTEDTAVY YCARGWLRDYLDLWGQGTLVTVSSASTKGPSVFPLAPSSKSTSG GTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSL SSWTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCP PCPAPEAAGGPSVFLFPPKPKDTLMISRTPEVTCWVSVSHEDPEV KFNWYVDGVEVHNAKTKPREEQYNSTYRWSVLTVLHQDWLN GKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKN QVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFL YSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPG25910 Full QSLVESGGGLVKPGGSLRLSCAVSGIDLSSYFMSWVRQAPGKGLE 135YIGTISTGGNTYYASWVKGRFDISKTSTTVYLQMNSLKTEDTAVY YCARGWLRDYLDLWGQGTLVTVSSASTKGPSVFPLAPSSKSTSG GTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSL SSWTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCP PCPAPEAAGGPSVFLFPPKPKDTLMISRTPEVTCWVSVSHEDPEV KFNWYVDGVEVHNAKTKPREEQYNSTYRWSVLTVLHQDWLN GKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKN QVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFL YSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPG27389 Full DIQMTQSPSTLSASVGDRVTITCRSRQSVYRENRLAWYQQKPGKA 136PKLLIYRASKLESGVPSRFSGSGSGTEFTLTISSLQPDDFATYYCAG GDSSGSDHAFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASVVC LLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSST LTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC27390 Full DIQMTQSPSTLSASVGDRVTITCRSRQSVYRENRLAWYQQKPGKA 137PKLLIYRGSKLESGVPSRFSGSGSGTEFTLTISSLQPDDFATYYCAG GDSSGSDHAFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASVVC LLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSST LTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC27391 Full DIQMTQSPSTLSASVGDRVTITCRSRQSVYRENRLAWYQQKPGKA 138PKLLIYRASKLEKGVASRFSGSGSGTEFTLTISSLQPDDFATYYCAGGDSSGSDHAFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASW129IPTS / 200238736.2Clone Region Sequence SEQ ID ID NO CLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSS TLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC27392 Full DIQMTQSPSTLSASVGDRVTITCRSRQSVYRENRLAWYQQKPGKA 139PKLLIYRASKLEKGVASRFSGSGSGTEFTLTISSLQPDDFATYYCA GGASSGSDHAFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASW CLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSS TLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC27394 Full EVQLVESGGGLVKPGGSLRLSCAVVGIDLSSYFMSWVRQAPGKG 140LEYIGTISTGGNTYYAAPVKGRFTISKTSTTVYLQINSPKTEDTAV YYCARGWLRDYLDRWGQGTLVTVSSASTKGPSVFPLAPSSKSTS GGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLY SLSSWTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHT CPPCPAPEAAGGPSVFLFPPKPKDTLMISRTPEVTCVWSVSHEDP EVKFNWYVDGVEVHNAKTKPREEQYNSTYRWSVLTVLHQDWL NGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTK NQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFF LYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPG27395 Full EVQLVESGGGLVKPGGSLRLSCAVSGIDLSSYFMSWVRQAPGKG 141LEYIGTISTGGNTYYASWVKGRFTISKDSSKNTVYLQMNSLKTED TAVYYCARGWLRDYLDRWGQGTLVTVSSASTKGPSVFPLAPSSK STSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSG LYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKT HTCPPCPAPEAAGGPSVFLFPPKPKDTLMISRTPEVTCVWSVSHE DPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRWSVLTVLHQD WLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDE LTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDG SFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPG27396 Full EVQLVESGGGLVKPGGSLRLSCAVVGIDLSSYFMSWVRQAPGKG 142LEYIGTISTGGNTYYASWVKGRFTISKDSSKNTVYLQMNSLKTED TAVYYCARGWLRDYLDRWGQGTLVTVSSASTKGPSVFPLAPSSK STSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSG LYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKT HTCPPCPAPEAAGGPSVFLFPPKPKDTLMISRTPEVTCVWSVSHE DPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRWSVLTVLHQD WLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDE LTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDG SFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPG27397 Full EVQLVESGGGLVKPGGSLRLSCAVSGIDLSSYFMSWVRQAPGKG 143LEYIGTISTGGNTYYASWVKGRFTISKTSTTVYLQMNSLKTEDTA VYYCARGWLRDYLDRWGQGTLVTVSSASTKGPSVFPLAPSSKST SGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGL YSLSSWTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTH TCPPCPAPEAAGGPSVFLFPPKPKDTLMISRTPEVTCVWSVSHED PEVKFNWYVDGVEVHNAKTKPREEQYNSTYRWSVLTVLHQDW LNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELT KNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPG130IPTS / 200238736.2Clone Region Sequence SEQ ID ID NO27813 Full EVQLVESGGGLVKPGGSLRLSCAVSGIDLSSYFMSWVRQAPGKG 144LEYIGTISTGGNTYYASWVKGRFTISKTSTDVYLQMNSLKTEDTA VYYCARGWLRDYLDRWGQGTLVTVSSASTKGPSVFPLAPSSKST SGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGL YSLSSWTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTH TCPPCPAPEAAGGPSVFLFPPKPKDTLMISRTPEVTCVWSVSHED PEVKFNWYVDGVEVHNAKTKPREEQYNSTYRWSVLTVLHQDW LNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELT KNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSF FLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPG29412 Full DIQMTQSPSTLSASVGDRVTITCRSRQSVYRENRLAWYQQKPGKA 145PKLLIYRGSKLESGVPSRFSGSGSGTEFTLTISSLQPDDFATYYCAG GDSSGSDHAFGGGTKVEIKRTVAAPSVFIFPPSDEELKSGTASVEC LLNNFYPREIKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTL TLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECKabat RSRQSVYRENRLA 40 CDR1Kabat RGSKLES 45 CDR2Kabat AGGDSSGSDHA 30 CDR329412 VL DIQMTQSPSTLSASVGDRVTITCRSRQSVYRENRLAWYQQKPGKA 146PKLLIYRGSKLESGVPSRFSGSGSGTEFTLTISSLQPDDFATYYCAG GDSSGSDHAFGGGTKVEIK29414 Full EVQLVESGGGLVKPGGSLRLSCAVSGIDLSSYFMSWVRQAPGKG 147LEYIGTISTGGNTYYASWVKGRFTISKDSSKNTVYLQMNSLKTED TAVYYCARGWLRDYLDRWGQGTLVTVSSASTKGPSVFPLAPSSK STSGGTAALGCRVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSG LYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKT HTCPPCPAPEAAGGPSVFLFPPKPKDTLMISRTPEVTCVWSVSHE DPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRWSVLTVLHQD WLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYVYPPSRDE LTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDG SFALVSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGKabat SYFMS 18 CDR1Kabat TISTGGNTYYASWVKG 19 CDR2Kabat GWLRDYLDR 39 CDR329414 VH EVQLVESGGGLVKPGGSLRLSCAVSGIDLSSYFMSWVRQAPGKG 148LEYIGTISTGGNTYYASWVKGRFTISKDSSKNTVYLQMNSLKTEDTAVYYCARGWLRDYLDRWGQGTLVTVSS131IPTS / 200238736.2Clone Region Sequence SEQ ID ID NO29417 Full DIQMTQSPSTLSASVGDRVTITCRSRQSVYRENRLAWYQQKPGKA 149PKLLIYRGSKLESGVPSRFSGSGSGTEFTLTISSLQPDDFATYYCAG GDSSGSDHAFGGGTKVEIKRTVAAPSVAIFPPSDERLKSGTASWC VLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSR LTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECKabat RSRQSVYRENRLA 40 CDR1Kabat RGSKLES 35 CDR2Kabat AGGDSSGSDHA 40 CDR329417 VL DIQMTQSPSTLSASVGDRVTITCRSRQSVYRENRLAWYQQKPGKA 150PKLLIYRGSKLESGVPSRFSGSGSGTEFTLTISSLQPDDFATYYCAG GDSSGSDHAFGGGTKVEIK29418 Full EVQLVESGGGLVKPGGSLRLSCAVSGIDLSSYFMSWVRQAPGKG 151LEYIGTISTGGNTYYASWVKGRFTISKDSSKNTVYLQMNSLKTED TAVYYCARGWLRDYLDRWGQGTLVTVSSASTKGPSVFPLAPSSK STSGGTAWLGCEVTDYFPEPVTVSWNSGALTSGVHTFPAVLESSG LYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKT HTCPPCPAPEAAGGPSVFLFPPKPKDTLMISRTPEVTCVWSVSHE DPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRWSVLTVLHQD WLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYVYPPSRDE LTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDG SFALVSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGKabat SYFMS 18 CDR1Kabat TISTGGNTYYASWVKG 19 CDR2Kabat GWLRDYLDR 39 CDR329418 VH EVQLVESGGGLVKPGGSLRLSCAVSGIDLSSYFMSWVRQAPGKG 152LEYIGTISTGGNTYYASWVKGRFTISKDSSKNTVYLQMNSLKTED TAVYYCARGWLRDYLDRWGQGTLVTVSS29420 Full QVQLVQSGAEVKKPGSSVKVSCKGSGPRIYAMHWVRQAPGQGL 153EWIGIISTYYGNTNYNQKFKGRATMTVDKSTSTAYMELSSLRSED TAVYYCARGERFYYFDYWGQGTTVTVSSASTKGPSVFPLAPSSKS TSGGTAALGCEVTDYFPEPVTVSWNSGALTSGVHTFPAVLESSGL YSLSSWTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTH TCPPCPAPEAAGGPSVFLFPPKPKDTLMISRTPEVTCVWSVSHED PEVKFNWYVDGVEVHNAKTKPREEQYNSTYRWSVLTVLHQDW LNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYVLPPSRDELT KNQVSLLCLVKGFYPSDIAVEWESNGQPENNYLTWPPVLDSDGSF FLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGKabat AMH 82 CDR1Kabat IISTYYGNTNYNQKFKG 72CDR2132IPTS / 200238736.2Clone Region Sequence SEQ ID ID NOKabat GERFYYFDY 73 CDR329420 VH QVQLVQSGAEVKKPGSSVKVSCKGSGPRIYAMHWVRQAPGQG 154LEWIGIISTYYGNTNYNQKFKGRATMTVDKSTSTAYMELSSLRSE DTAVYYCARGERFYYFDYWGQGTTVTVSS29424 Full DIQLTQSPSSLSASVGDRVTITCRASKSISKYLAWYQQKPGKAPKL 155LIYKRSTLQSGVPSRFSGSGSHTDFTLTISSLQPEDFATYYCQQHSE YPFTFGQGTKLEIKRTVAAPSVFIFPPSDERLKSGTASWCLLNNF YPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSRLTLSK ADYEKHKVYACEVTHQGLS SPVTKSFNRGECKabat RASKSISKYLA 89 CDR1Kabat KRSTLQS 90 CDR2Kabat QQHSEYPFT 88 CDR329424 VL DIQLTQSPSSLSASVGDRVTITCRASKSISKYLAWYQQKPGKAPKL 156LIYKRSTLQSGVPSRFSGSGSHTDFTLTISSLQPEDFATYYCQQHSE YPFTFGQGTKLEIK29425 Full DIQLTQSPSSLSASVGDRVTITCRASKSISKYLAWYQQKPGKAPKL 157LIYKRSTLQSGVPSRFSGSGSHTDFTLTISSLQPEDFATYYCQQHSE YPFTFGQGTKLEIKRTVAAPSVFIFPPSDEELKSGTASVVCWLNNF YPREIKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSELELSKA DYEKHKVYACEVTHQGLS SPVTKSFNRGECKabat RASKSISKYLA 89 CDR1Kabat KRSTLQS 90 CDR2Kabat QQHSEYPFT 88 CDR329425 VL DIQLTQSPSSLSASVGDRVTITCRASKSISKYLAWYQQKPGKAPKL 158LIYKRSTLQSGVPSRFSGSGSHTDFTLTISSLQPEDFATYYCQQHSE YPFTFGQGTKLEIK29426 Full QVQLVQSGAEVKKPGSSVKVSCKGSGYTFTLKAMHWVRQAPGQ 159GLEWIGIISTYYGNTNYNQKFKGRATMTVDKSTSTAYMELSSLRS EDTAVYYCARGERFYYFDYWGQGTTVTVSSASTKGPSVFPLAPSS KSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLRSS GLYSLSSWTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDK THTCPPCPAPEAAGGPSVFLFPPKPKDTLMISRTPEVTCWVSVSH EDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQ DWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYVLPPSRD ELTKNQVSLLCLVKGFYPSDIAVEWESNGQPENNYLTWPPVLDSD GSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSP GKabat LKAMH 71CDR1133IPTS / 200238736.2Clone Region Sequence SEQ ID ID NOKabat IISTYYGNTNYNQKFKG 72 CDR2Kabat GERFYYFDY 73 CDR329426 VH QVQLVQSGAEVKKPGSSVKVSCKGSGYTFTLKAMHWVRQAPGQ 160GLEWIGIISTYYGNTNYNQKFKGRATMTVDKSTSTAYMELSSLRS EDTAVYYCARGERFYYFDYWGQGTTVTVSS29440 Full QVQLVQSGAEVKKPGSSVKVSCKGSGPRIYAMHWVRQAPGQGL 161EWIGIISTYYGNTNYNQKFKGRATMTVDKSTSTAYMELSSLRSED TAVYYCARGERFYYFDYWGQGTTVTVSSASTKGPSVFPLAPSSKS TSGGTAALGCEVTDYFPEPVTVSWNSGALTSGVHTFPAVLESSGL YSLSSWTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTH TCPPCPAPEAAGGPSVFLFPPKPKDTLMISRTPEVTCVWSVSHED PEVKFNWYVDGVEVHNAKTKPREEQYNSTYRWSVLTVLHQDW LNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYVYPPSRDEL TKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGS FALVSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGKabat AMH 82 CDR1Kabat IISTYYGNTNYNQKFKG 72 CDR2Kabat GERFYYFDY 73 CDR329440 VH QVQLVQSGAEVKKPGSSVKVSCKGSGPRIYAMHWVRQAPGQG 162LEWIGIISTYYGNTNYNQKFKGRATMTVDKSTSTAYMELSSLRSE DTAVYYCARGERFYYFDYWGQGTTVTVSS29448 Full QVQLVQSGAEVKKPGSSVKVSCKGSGYTFTLKAMHWVRQAPGQ 163GLEWIGIISTYYGNTNYNQKFKGRATMTVDKSTSTAYMELSSLRS EDTAVYYCARGERFYYFDYWGQGTTVTVSSASTKGPSVFPLAPSS KSTSGGTAALGCEVTDYFPEPVTVSWNSGALTSGVHTFPAVLESS GLYSLSSWTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDK THTCPPCPAPEAAGGPSVFLFPPKPKDTLMISRTPEVTCWVSVSH EDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQ DWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYVYPPSRD ELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSD GSFALVSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSP GKabat LKAMH 71 CDR1Kabat IISTYYGNTNYNQKFKG 72 CDR2Kabat GERFYYFDY 73 CDR329448 VH QVQLVQSGAEVKKPGSSVKVSCKGSGYTFTLKAMHWVRQAPGQ 164GLEWIGIISTYYGNTNYNQKFKGRATMTVDKSTSTAYMELSSLRSEDTAVYYCARGERFYYFDYWGQGTTVTVSS134IPTS / 200238736.2Clone Region Sequence SEQ ID ID NO29882 Full EVQLVESGGGLVKPGGSLRLSCAVSGIDLSSYFMSWVRQAPGKG 165LEYIGTISTGGNTYYASWVKGRFTISKTSTDVYLQMNSLKTEDTA VYYCARGWLRDYLDRWGQGTLVTVSSASTKGPSVFPLAPSSKST SGGTAALGCRVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGL YSLSSWTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTH TCPPCPAPEAAGGPSVFLFPPKPKDTLMISRTPEVTCVWSVSHED PEVKFNWYVDGVEVHNAKTKPREEQYNSTYRWSVLTVLHQDW LNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYVLPPSRDELT KNQVSLLCLVKGFYPSDIAVEWESNGQPENNYLTWPPVLDSDGSF FLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGKabat SYFMS 18 CDR1Kabat TISTGGNTYYASWVKG 19 CDR2Kabat GWLRDYLDR 39 CDR329882 VH EVQLVESGGGLVKPGGSLRLSCAVSGIDLSSYFMSWVRQAPGKG 166LEYIGTISTGGNTYYASWVKGRFTISKTSTDVYLQMNSLKTEDTA VYYCARGWLRDYLDRWGQGTLVTVSS31317 Full DIQLTQSPSSLSASVGDRVTITCRASKSISKYLAWYQQKPGKAPKL 167LIYKRSTLQSGVPSRFSGSGSHTDFTLTISSLQPEDFATYYCQQHSE YPFTFGQGTKLEIKRTVAAPSVFIFPPSDEELKSGTASVVCWLNNF YPREIKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSELELSKA DYEKHKVYACEVTHQGLS SPVTKSFNRGECKabat RASKSISKYLA 89 CDR1Kabat KRSTLQS 90 CDR2Kabat QQHSEYPFT 88 CDR331317 VL DIQLTQSPSSLSASVGDRVTITCRASKSISKYLAWYQQKPGKAPKL 168LIYKRSTLQSGVPSRFSGSGSHTDFTLTISSLQPEDFATYYCQQHSE YPFTFGQGTKLEIK31575 Full QVQLVQSGAEVKKPGSSVKVSCKGSGYTFTLKAMHWVRQAPGQ 169GLEWIGIISTYYGNTNYNQKFKGRATMTVDKSTSTAYMELSSLRS EDTAVYYCARGERFYYFDYWGQGTTVTVSSASTKGPSVFPLAPSS KSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLRSS GLYSLSSWTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDK THTCPPCPAPEAAGGPSVFLFPPKPKDTLMISRTPEVTCWVSVSH EDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQ DWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYVLPPSRD ELTKNQVSLLCLVKGFYPSDIAVEWESNGQPENNYLTWPPVLDSD GSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSP GKKabat LKAMH 71CDR1135IPTS / 200238736.2Clone Region Sequence SEQ ID ID NOKabat IISTYYGNTNYNQKFKG 72 CDR2Kabat GERFYYFDY 73 CDR331575 VH QVQLVQSGAEVKKPGSSVKVSCKGSGYTFTLKAMHWVRQAPGQ 170GLEWIGIISTYYGNTNYNQKFKGRATMTVDKSTSTAYMELSSLRS EDTAVYYCARGERFYYFDYWGQGTTVTVSS31576 Full EVQLVESGGGLVKPGGSLRLSCAVSGIDLSSYFMSWVRQAPGKG 171LEYIGTISTGGNTYYASWVKGRFTISKDSSKNTVYLQMNSLKTED TAVYYCARGWLRDYLDRWGQGTLVTVSSASTKGPSVFPLAPSSK STSGGTAWLGCEVTDYFPEPVTVSWNSGALTSGVHTFPAVLESSG LYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKT HTCPPCPAPEAAGGPSVFLFPPKPKDTLMISRTPEVTCVWSVSHE DPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRWSVLTVLHQD WLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYVYPPSRDE LTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDG SFALVSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPG KKabat SYFMS 18 CDR1Kabat TISTGGNTYYASWVKG 19 CDR2Kabat GWLRDYLDR 39 CDR331576 VH EVQLVESGGGLVKPGGSLRLSCAVSGIDLSSYFMSWVRQAPGKG 172LEYIGTISTGGNTYYASWVKGRFTISKDSSKNTVYLQMNSLKTED TAVYYCARGWLRDYLDRWGQGTLVTVSS31577 Full QVQLVQSGAEVKKPGSSVKVSCKGSGYTFTLKAMHWVRQAPGQ 173GLEWIGIISTYYGNTNYNQKFKGRATMTVDKSTSTAYMELSSLRS EDTAVYYCARGERFYYFDYWGQGTTVTVSSASTKGPSVFPLAPSS KSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLRSS GLYSLSSWTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDK THTCPPCPAPEAAGGPSVFLFPPKPKDTLYITREPEVTCVWSVSH EDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQ DWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYVLPPSRD ELTKNQVSLLCLVKGFYPSDIAVEWESNGQPENNYLTWPPVLDSD GSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSP GKKabat LKAMH 71 CDR1Kabat IISTYYGNTNYNQKFKG 72 CDR2Kabat GERFYYFDY 73 CDR331577 VH QVQLVQSGAEVKKPGSSVKVSCKGSGYTFTLKAMHWVRQAPGQ 174GLEWIGIISTYYGNTNYNQKFKGRATMTVDKSTSTAYMELSSLRSEDTAVYYCARGERFYYFDYWGQGTTVTVSS136IPTS / 200238736.2Clone Region Sequence SEQ ID ID NO31578 Full EVQLVESGGGLVKPGGSLRLSCAVSGIDLSSYFMSWVRQAPGKG 175LEYIGTISTGGNTYYASWVKGRFTISKDSSKNTVYLQMNSLKTED TAVYYCARGWLRDYLDRWGQGTLVTVSSASTKGPSVFPLAPSSK STSGGTAWLGCEVTDYFPEPVTVSWNSGALTSGVHTFPAVLESSG LYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKT HTCPPCPAPEAAGGPSVFLFPPKPKDTLYITREPEVTCVWSVSHE DPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRWSVLTVLHQD WLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYVYPPSRDE LTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDG SFALVSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPG KKabat SYFMS 18 CDR1Kabat TISTGGNTYYASWVKG 19 CDR2Kabat GWLRDYLDR 39 CDR331578 VH EVQLVESGGGLVKPGGSLRLSCAVSGIDLSSYFMSWVRQAPGKG 176LEYIGTISTGGNTYYASWVKGRFTISKDSSKNTVYLQMNSLKTED TAVYYCARGWLRDYLDRWGQGTLVTVSS31580 Full DIQMTQSPSTLSASVGDRVTITCRSRQSVYRENRLAWYQQKPGKA 177PKLLIYRGSKLESGVPSRFSGSGSGTEFTLTISSLQPDDFATYYCAG GDSSGSDHAFGGGTKVEIKRTVAAPSVAIFPPSDERLKSGTASWC VLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSR LTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGECKabat RSRQSVYRENRLA 40 CDR1Kabat RGSKLES 45 CDR2Kabat AGGDSSGSDHA 30 CDR331580 VL DIQMTQSPSTLSASVGDRVTITCRSRQSVYRENRLAWYQQKPGKA 178PKLLIYRGSKLESGVPSRFSGSGSGTEFTLTISSLQPDDFATYYCAG GDSSGSDHAFGGGTKVEIK31743 Full QVQLVQSGAEVKKPGSSVKVSCKGSGYTFTLKAMHWVRQAPGQ 179GLEWIGIISTYYGNTNYNQKFKGRATMTVDKSTSTAYMELSSLRS EDTAVYYCARGERFYYFDYWGQGTTVTVSSASTKGPSVFPLAPC SRSTSESTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLRS SGLYSLSSWTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKYGP PCPPCPAPEFLGGPSVFLFPPKPKDTLMISRTPEVTCVWDVSQEDP EVQFNWYVDGVEVHNAKTKPREEQFNSTYRWSVLTVLHQDWL NGKEYKCKVSNKGLPSSIEKTISKAKGQPREPQVYVYPPSQEEMT KNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSF ALVSKLTVDKSRWQEGNVFSCSVMHEALHNHYTQKSLSLSLGKKabat LKAMH 71CDR1137IPTS / 200238736.2Clone Region Sequence SEQ ID ID NOKabat IISTYYGNTNYNQKFKG 72 CDR2Kabat GERFYYFDY 73 CDR331743 VH QVQLVQSGAEVKKPGSSVKVSCKGSGYTFTLKAMHWVRQAPGQ 180GLEWIGIISTYYGNTNYNQKFKGRATMTVDKSTSTAYMELSSLRS EDTAVYYCARGERFYYFDYWGQGTTVTVSS31744 Full QVQLVQSGAEVKKPGSSVKVSCKGSGYTFTLKAMHWVRQAPGQ 181GLEWIGIISTYYGNTNYNQKFKGRATMTVDKSTSTAYMELSSLRS EDTAVYYCARGERFYYFDYWGQGTTVTVSSASTKGPSVFPLAPC SRSTSESTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLRS SGLYSLSSWTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKYGP PCPPCPAPEFLGGPSVFLFPPKPKDTLYITREPEVTCVWDVSQEDP EVQFNWYVDGVEVHNAKTKPREEQFNSTYRWSVLTVLHQDWL NGKEYKCKVSNKGLPSSIEKTISKAKGQPREPQVYVYPPSQEEMT KNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSF ALVSKLTVDKSRWQEGNVFSCSVMHEALHNHYTQKSLSLSLGKKabat LKAMH 71 CDR1Kabat IISTYYGNTNYNQKFKG 72 CDR2Kabat GERFYYFDY 73 CDR331744 VH QVQLVQSGAEVKKPGSSVKVSCKGSGYTFTLKAMHWVRQAPGQ 182GLEWIGIISTYYGNTNYNQKFKGRATMTVDKSTSTAYMELSSLRS EDTAVYYCARGERFYYFDYWGQGTTVTVSS31745 Full EVQLVESGGGLVKPGGSLRLSCAVSGIDLSSYFMSWVRQAPGKG 183LEYIGTISTGGNTYYASWVKGRFTISKDSSKNTVYLQMNSLKTED TAVYYCARGWLRDYLDRWGQGTLVTVSSASTKGPSVFPLAPCSR STSESTAWLGCEVTDYFPEPVTVSWNSGALTSGVHTFPAVLESSG LYSLSSVVTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKYGPPC PPCPAPEFLGGPSVFLFPPKPKDTLMISRTPEVTCWVDVSQEDPE VQFNWYVDGVEVHNAKTKPREEQFNSTYRWSVLTVLHQDWLN GKEYKCKVSNKGLPSSIEKTISKAKGQPREPQVYVLPPSQEEMTK NQVSLLCLVKGFYPSDIAVEWESNGQPENNYLTWPPVLDSDGSFF LYSKLTVDKSRWQEGNVFSCSVMHEALHNHYTQKSLSLSLGKKabat SYFMS 18 CDR1Kabat TISTGGNTYYASWVKG 19 CDR2Kabat GWLRDYLDR 39 CDR331745 VH EVQLVESGGGLVKPGGSLRLSCAVSGIDLSSYFMSWVRQAPGKG 184LEYIGTISTGGNTYYASWVKGRFTISKDSSKNTVYLQMNSLKTEDTAVYYCARGWLRDYLDRWGQGTLVTVSS138IPTS / 200238736.2Clone Region Sequence SEQ ID ID NO31746 Full EVQLVESGGGLVKPGGSLRLSCAVSGIDLSSYFMSWVRQAPGKG 185LEYIGTISTGGNTYYASWVKGRFTISKDSSKNTVYLQMNSLKTED TAVYYCARGWLRDYLDRWGQGTLVTVSSASTKGPSVFPLAPCSR STSESTAWLGCEVTDYFPEPVTVSWNSGALTSGVHTFPAVLESSG LYSLSSVVTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKYGPPC PPCPAPEFLGGPSVFLFPPKPKDTLYITREPEVTCWVDVSQEDPEV QFNWYVDGVEVHNAKTKPREEQFNSTYRVVSVLTVLHQDWLNG KEYKCKVSNKGLPSSIEKTISKAKGQPREPQVYVLPPSQEEMTKN QVSLLCLVKGFYPSDIAVEWESNGQPENNYLTWPPVLDSDGSFFL YSKLTVDKSRWQEGNVFSCSVMHEALHNHYTQKSLSLSLGKKabat SYFMS 18 CDR1Kabat TISTGGNTYYASWVKG 19 CDR2Kabat GWLRDYLDR 39 CDR331746 VH EVQLVESGGGLVKPGGSLRLSCAVSGIDLSSYFMSWVRQAPGKG 186LEYIGTISTGGNTYYASWVKGRFTISKDSSKNTVYLQMNSLKTED TAVYYCARGWLRDYLDRWGQGTLVTVSS31747 Full QVQLVQSGAEVKKPGSSVKVSCKGSGYTFTLKAMHWVRQAPGQ 187GLEWIGIISTYAGNTNYNQKFKGRATMTVDKSTSTAYMELSSLRS EDTAVYYCARGERFYYFDYWGQGTTVTVSSASTKGPSVFPLAPC SRSTSESTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLRS SGLYSLSSWTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKYGP PCPPCPAPEFLGGPSVFLFPPKPKDTLYITREPEVTCVWDVSQEDP EVQFNWYVDGVEVHNAKTKPREEQFNSTYRWSVLTVLHQDWL NGKEYKCKVSNKGLPSSIEKTISKAKGQPREPQVYVYPPSQEEMT KNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSF ALVSKLTVDKSRWQEGNVFSCSVMHEALHNHYTQKSLSLSLGKKabat LKAMH 71 CDR1Kabat IISTYAGNTNYNQKFKG 72 CDR2Kabat GERFYYFDY 73 CDR331747 VH QVQLVQSGAEVKKPGSSVKVSCKGSGYTFTLKAMHWVRQAPGQ 188GLEWIGIISTYAGNTNYNQKFKGRATMTVDKSTSTAYMELSSLRSEDTAVYYCARGERFYYFDYWGQGTTVTVSS139IPTS / 200238736.2Clone Region Sequence SEQ ID ID NO32097 Full QVQLVQSGAEVKKPGSSVKVSCKGSGYTFTLKAMHWVRQAPGQ 189GLEWIGIISTYYGNTNYNQKFQGRATMTVDESTSTAYMELSSLRS EDTAVYYCARGERFYYFDYWGQGTTVTVSSASTKGPSVFPLAPC SRSTSESTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLRS SGLYSLSSWTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKYGP PCPPCPAPEFLGGPSVFLFPPKPKDTLYITREPEVTCVWDVSQEDP EVQFNWYVDGVEVHNAKTKPREEQFNSTYRWSVLTVLHQDWL NGKEYKCKVSNKGLPSSIEKTISKAKGQPREPQVYVYPPSQEEMT KNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSF ALVSKLTVDKSRWQEGNVFSCSVMHEALHNHYTQKSLSLSLGKKabat LKAMH 71 CDR1Kabat IISTYYGNTNYNQKFQG 102 CDR2Kabat GERFYYFDY 73 CDR332097 VH QVQLVQSGAEVKKPGSSVKVSCKGSGYTFTLKAMHWVRQAPGQ 190GLEWIGIISTYYGNTNYNQKFQGRATMTVDESTSTAYMELSSLRS EDTAVYYCARGERFYYFDYWGQGTTVTVSS32102 Full QVQLVQSGAEVKKPGSSVKVSCKGSGYTFTLKAMHWVRQAPGQ 191GLEWIGIISTYYGNTNYNQKFKGRATMTVDESTSTAYMELSSLRS EDTAVYYCARGERFYYFDYWGQGTTVTVSSASTKGPSVFPLAPC SRSTSESTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLRS SGLYSLSSWTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKYGP PCPPCPAPEFLGGPSVFLFPPKPKDTLYITREPEVTCVWDVSQEDP EVQFNWYVDGVEVHNAKTKPREEQFNSTYRWSVLTVLHQDWL NGKEYKCKVSNKGLPSSIEKTISKAKGQPREPQVYVYPPSQEEMT KNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSF ALVSKLTVDKSRWQEGNVFSCSVMHEALHNHYTQKSLSLSLGKKabat LKAMH 71 CDR1Kabat IISTYYGNTNYNQKFKG 72 CDR2Kabat GERFYYFDY 73 CDR332102 VH QVQLVQSGAEVKKPGSSVKVSCKGSGYTFTLKAMHWVRQAPGQ 192GLEWIGIISTYYGNTNYNQKFKGRATMTVDESTSTAYMELSSLRSEDTAVYYCARGERFYYFDYWGQGTTVTVSS140IPTS / 200238736.2Clone Region Sequence SEQ ID ID NO32104 Full QVQLVQSGAEVKKPGSSVKVSCKGSGYTFTLKAMHWVRQAPGQ 193GLEWIGIISTYYGNTNYNDKFKGRATMTVDESTSTAYMELSSLRS EDTAVYYCARGERFYYFDYWGQGTTVTVSSASTKGPSVFPLAPC SRSTSESTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLRS SGLYSLSSWTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKYGP PCPPCPAPEFLGGPSVFLFPPKPKDTLYITREPEVTCVWDVSQEDP EVQFNWYVDGVEVHNAKTKPREEQFNSTYRWSVLTVLHQDWL NGKEYKCKVSNKGLPSSIEKTISKAKGQPREPQVYVYPPSQEEMT KNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSF ALVSKLTVDKSRWQEGNVFSCSVMHEALHNHYTQKSLSLSLGKKabat LKAMH 71 CDR1Kabat IISTYYGNTNYNDKFKG 101 CDR2Kabat GERFYYFDY 73 CDR332104 VH QVQLVQSGAEVKKPGSSVKVSCKGSGYTFTLKAMHWVRQAPGQ 194GLEWIGIISTYYGNTNYNDKFKGRATMTVDESTSTAYMELSSLRS EDTAVYYCARGERFYYFDYWGQGTTVTVSS32133 Full EVQLVQSGAEVKQPGSSVKVSCAGSGYTFTLKAMHWVRQAPGQ 195GLEWIGIISTYYGNTNYNDQFKGRATMTVDTSTSTAYMELSSLRS EDTAVYYCARGERFYYFDYWGQGTTVTVSSASTKGPSVFPLAPC SRSTSESTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLRS SGLYSLSSWTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKYGP PCPPCPAPEFLGGPSVFLFPPKPKDTLYITREPEVTCVWDVSQEDP EVQFNWYVDGVEVHNAKTKPREEQFNSTYRWSVLTVLHQDWL NGKEYKCKVSNKGLPSSIEKTISKAKGQPREPQVYVYPPSQEEMT KNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSF ALVSKLTVDKSRWQEGNVFSCSVMHEALHNHYTQKSLSLSLGKKabat LKAMH 71 CDR1Kabat IISTYYGNTNYNDQFKG 196 CDR2Kabat GERFYYFDY 73 CDR332133 VH EVQLVQSGAEVKQPGSSVKVSCAGSGYTFTLKAMHWVRQAPGQ 197GLEWIGIISTYYGNTNYNDQFKGRATMTVDTSTSTAYMELSSLRS EDTAVYYCARGERFYYFDYWGQGTTVTVSS32134 Full DIQLTQSPESLEASVGDRVTITCRASKSISKYLAWYQQKPGKAPKL 198LIYKRSTLQSGVPSRFSGSGSHTDFTLTISSLQPEDFATYYCQQHSE YPFTFGQGTKLEIKRTVAAPSVFIFPPSDEELKSGTASVVCWLNNF YPREIKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSELELSKA DYEKHKVYACEVTHQGLS SPVTKSFNRGECKabat RASKSISKYLA 89 CDR1Kabat KRSTLQS 90CDR2141IPTS / 200238736.2Clone Region Sequence SEQ ID ID NOKabat QQHSEYPFT 88 CDR332134 VL DIQLTQSPESLEASVGDRVTITCRASKSISKYLAWYQQKPGKAPKL 199LIYKRSTLQSGVPSRFSGSGSHTDFTLTISSLQPEDFATYYCQQHSE YPFTFGQGTKLEIK32139 Full DIQLTQSPSSLSASVGDSVTITCRASKSISKYLAWYQQKPGQAPKL 200LIYKRSTLQSGVPDRFSGSGSHTDFTLTISSLQPEDFATYYCQQHSE YPFTFGQGTKLEIKRTVAAPSVFIFPPSDEELKSGTASVVCWLNNF YPREIKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSELELSKA DYEKHKVYACEVTHQGLS SPVTKSFNRGECKabat RASKSISKYLA 89 CDR1Kabat KRSTLQS 90 CDR2Kabat QQHSEYPFT 88 CDR332139 VL DIQLTQSPSSLSASVGDSVTITCRASKSISKYLAWYQQKPGQAPKL 201LIYKRSTLQSGVPDRFSGSGSHTDFTLTISSLQPEDFATYYCQQHSE YPFTFGQGTKLEIK32147 Full DIQLTQSPSSLSASVGDRVTITCRASKSISKYLAWYQQKPGEAPKL 202LIYKRSTLQSGVPSRFSGSGSHTDFTLTISSLQPEDFATYYCQQHSE YPFTFGQGTKLEIKRTVAAPSVFIFPPSDEELKSGTASVVCWLNNF YPREIKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSELELSKA DYEKHKVYACEVTHQGLS SPVTKSFNRGECKabat RASKSISKYLA 89 CDR1Kabat KRSTLQS 90 CDR2Kabat QQHSEYPFT 88 CDR332147 VL DIQLTQSPSSLSASVGDRVTITCRASKSISKYLAWYQQKPGEAPKL 203LIYKRSTLQSGVPSRFSGSGSHTDFTLTISSLQPEDFATYYCQQHSE YPFTFGQGTKLEIK32151 Full DIQLTQSPSSLSASVGDTVTITCRASKSISKYLAWYQQKPGEAPKL 204LIYKRSTLQSGVPSRFSGSGSHTDFTLTISSLQPEDFATYYCQQHSE YPFTFGQGTKLEIKRTVAAPSVFIFPPSDEELKSGTASVVCWLNNF YPREIKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSELELSKA DYEKHKVYACEVTHQGLS SPVTKSFNRGECKabat RASKSISKYLA 89 CDR1Kabat KRSTLQS 90 CDR2Kabat QQHSEYPFT 88 CDR332151 VL DIQLTQSPSSLSASVGDTVTITCRASKSISKYLAWYQQKPGEAPKL 205LIYKRSTLQSGVPSRFSGSGSHTDFTLTISSLQPEDFATYYCQQHSEYPFTFGQGTKLEIK142IPTS / 200238736.2Clone Region Sequence SEQ ID ID NO32153 Full DIELTQSPSSLSASVGDTVTITCRASKSISKYLAWYQQKPGEAPKL 206LIYKRSTLQSGVPSRFSGSGSHTDFTLTISSLQPEDFATYYCQQHSE YPFTFGQGTKLEIKRTVAAPSVFIFPPSDEELKSGTASVVCWLNNF YPREIKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSELELSKA DYEKHKVYACEVTHQGLS SPVTKSFNRGECKabat RASKSISKYLA 89 CDR1Kabat KRSTLQS 90 CDR2Kabat QQHSEYPFT 88 CDR332153 VL DIELTQSPSSLSASVGDTVTITCRASKSISKYLAWYQQKPGEAPKL 207LIYKRSTLQSGVPSRFSGSGSHTDFTLTISSLQPEDFATYYCQQHSE YPFTFGQGTKLEIK32154 Full DIQLTQSPSSLSASVGDRVTITCRASKSISKYLAWYQQKPGKAPKL 208LIYKRSTLESGVPSRFSGSGSHTDFTLTISSLQPEDFATYYCQQHSE YPFTFGQGTKLEEKRTVAAPSVFIFPPSDEELKSGTASWCWLNNF YPREIKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSELELSKA DYEKHKVYACEVTHQGLS SPVTKSFNRGECKabat RASKSISKYLA 89 CDR1Kabat KRSTLES 209 CDR2Kabat QQHSEYPFT 88 CDR332154 VL DIQLTQSPSSLSASVGDRVTITCRASKSISKYLAWYQQKPGKAPKL 210LIYKRSTLESGVPSRFSGSGSHTDFTLTISSLQPEDFATYYCQQHSE YPFTFGQGTKLEEK32190 Full DIQLTQSPSSLSASVGDPVTITCRASKSISKYLAWYQQKPGQAPQL 211LIYKRSTLQSGVPDRFSGSGSHTDFTLTISSLEPEDFATYYCQQHSE YPFTFGQGTKLEIKRTVAAPSVFIFPPSDEELKSGTASVVCWLNNF YPREIKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSELELSKA DYEKHKVYACEVTHQGLS SPVTKSFNRGECKabat RASKSISKYLA 89 CDR1Kabat KRSTLQS 90 CDR2Kabat QQHSEYPFT 88 CDR332190 VL DIQLTQSPSSLSASVGDPVTITCRASKSISKYLAWYQQKPGQAPQL 212LIYKRSTLQSGVPDRFSGSGSHTDFTLTISSLEPEDFATYYCQQHSEYPFTFGQGTKLEIK143IPTS / 200238736.2Clone Region Sequence SEQ ID ID NO32191 Full EVQLVESGGGLVKPGGSLRLSCAVSGIDLSSYFMSWVRQAPGKG 213LEYIGTISTGGNTYYASWVKGRFTISKDSSKNTVYLQMNSLKTED TAVYYCARGWLRDYLDRWGQGTLVTVSSASTKGPSVFPLAPCSR STSESTAWLGCEVTDYFPEPVTVSWNSGALTSGVHTFPAVLESSG LYSLSSVVTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKYGPPC PPCPAPEFLGGPSVFLFPPKPKDTLYITREPEVTCWVDVSQEDPEV QFNWYVDGVEVHNAKTKPREEQFNSTYRVVSVLTVLHQDWLNG KEYKCKVSNKGLPSSIEKTISKAKGQPREPQVYVLPPSQEEMTKN QVSLLCLVKGFYPSDIAVEWESNGQPENNYLTWPPVLDSDGSFFL YSKLTVDKSRWQEGNVFSCSVMHEALHNHYTQKSLSLSLGKKabat SYFMS 18 CDR1Kabat TISTGGNTYYASWVKG 19 CDR2Kabat GWLRDYLDR 39 CDR332191 VH EVQLVESGGGLVKPGGSLRLSCAVSGIDLSSYFMSWVRQAPGKG 214LEYIGTISTGGNTYYASWVKGRFTISKDSSKNTVYLQMNSLKTED TAVYYCARGWLRDYLDRWGQGTLVTVSS32781 Full QVQLVQSGAEVKKPGSSVKVSCKGSGYTFTLKAMHWVRQAPGQ 215GLEWIGIISTYAGNTNYNQKFQGRATMTVDESTSTAYMELSSLRS EDTAVYYCARGERFYYFDYWGQGTTVTVSSASTKGPSVFPLAPC SRSTSESTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLRS SGLYSLSSWTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKYGP PCPPCPAPEFLGGPSVFLFPPKPKDTLYITREPEVTCVWDVSQEDP EVQFNWYVDGVEVHNAKTKPREEQFNSTYRWSVLTVLHQDWL NGKEYKCKVSNKGLPSSIEKTISKAKGQPREPQVYVYPPSQEEMT KNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSF ALVSKLTVDKSRWQEGNVFSCSVMHEALHNHYTQKSLSLSLGKKabat LKAMH 71 CDR1Kabat IISTYAGNTNYNQKFQG 100 CDR2Kabat GERFYYFDY 73 CDR332781 VH QVQLVQSGAEVKKPGSSVKVSCKGSGYTFTLKAMHWVRQAPGQ 216GLEWIGIISTYAGNTNYNQKFQGRATMTVDESTSTAYMELSSLRSEDTAVYYCARGERFYYFDYWGQGTTVTVSS144IPTS / 200238736.2Clone Region Sequence SEQ ID ID NO32783 Full QVQLVQSGAEVKKPGSSVKVSCKGSGYTFTLKAMHWVRQAPGQ 217GLEWIGIISTYAGNTNYNDKFKGRATMTVDESTSTAYMELSSLRS EDTAVYYCARGERFYYFDYWGQGTTVTVSSASTKGPSVFPLAPC SRSTSESTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLRS SGLYSLSSWTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKYGP PCPPCPAPEFLGGPSVFLFPPKPKDTLYITREPEVTCVWDVSQEDP EVQFNWYVDGVEVHNAKTKPREEQFNSTYRWSVLTVLHQDWL NGKEYKCKVSNKGLPSSIEKTISKAKGQPREPQVYVYPPSQEEMT KNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSF ALVSKLTVDKSRWQEGNVFSCSVMHEALHNHYTQKSLSLSLGKKabat LKAMH 71 CDR1Kabat IISTYAGNTNYNDKFKG 218 CDR2Kabat GERFYYFDY 73 CDR332783 VH QVQLVQSGAEVKKPGSSVKVSCKGSGYTFTLKAMHWVRQAPGQ 219GLEWIGIISTYAGNTNYNDKFKGRATMTVDESTSTAYMELSSLRS EDTAVYYCARGERFYYFDYWGQGTTVTVSS32860 Full EVQLVQSGAEVKKPGSSVKVSCKGSGYTFTLKAMHWVRQAPGQ 220GLEWIGIISTYAGNTNYNQKFKERATMTVDKSTSTAYMELSSLRS EDTAVYYCARGERFYYFDYWGQGTTVTVSSASTKGPSVFPLAPC SRSTSESTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLRS SGLYSLSSWTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKYGP PCPPCPAPEFLGGPSVFLFPPKPKDTLYITREPEVTCVWDVSQEDP EVQFNWYVDGVEVHNAKTKPREEQFNSTYRWSVLTVLHQDWL NGKEYKCKVSNKGLPSSIEKTISKAKGQPREPQVYVYPPSQEEMT KNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSF ALVSKLTVDKSRWQEGNVFSCSVMHEALHNHYTQKSLSLSLGKKabat LKAMH 71 CDR1Kabat IISTYAGNTNYNQKFKE 221 CDR2Kabat GERFYYFDY 73 CDR332860 VH EVQLVQSGAEVKKPGSSVKVSCKGSGYTFTLKAMHWVRQAPGQ 222GLEWIGIISTYAGNTNYNQKFKERATMTVDKSTSTAYMELSSLRSEDTAVYYCARGERFYYFDYWGQGTTVTVSS145IPTS / 200238736.2Clone Region Sequence SEQ ID ID NO32861 Full QVQLVQSGAEVKKPGSSVKVSCKGSGYTFTLKAMHWVRQAPGQ 223GLEWIGIISTYAGNTNYNQKFKGRATMTVDKSTSTAYMELSSLRS EDTAVYYCARGERFYYFDYWGQGTTVTVSSASTKGPSVFPLAPC SRSTSESTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLRS SGLYSLSSWTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKYGP PCPPCPAPEFLGGPSVFLFPPKPKDTLYITREPEVTCVWDVSQEDP EVQFNWYVDGVEVHNAKTKPREEQFNSTYRWSVLTVLHQDWL NGKEYKCKVSNKGLPSSIEKTISKAKGQPREPQVYVYPPSQEEMT KNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSF ALVSKLTVDKSRWQEGNVFSCSVMHEALHNHYTQKSLSLSLGKKabat LKAMH 71 CDR1Kabat IISTYAGNTNYNQKFKG 95 CDR2Kabat GERFYYFDY 73 CDR332861 VH QVQLVQSGAEVKKPGSSVKVSCKGSGYTFTLKAMHWVRQAPGQ 224GLEWIGIISTYAGNTNYNQKFKGRATMTVDKSTSTAYMELSSLRS EDTAVYYCARGERFYYFDYWGQGTTVTVSS32862 Full QVQLVQSGAEVKKPGSSVKVSCKGSGYTFTLKAMHWVRQAPGQ 225GLEWIGIISTYAGNTNYNQKFKGRATMTVDESTSTAYMELSSLRS EDTAVYYCARGERFYYFDYWGQGTTVTVSSASTKGPSVFPLAPC SRSTSESTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLRS SGLYSLSSWTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKYGP PCPPCPAPEFLGGPSVFLFPPKPKDTLYITREPEVTCVWDVSQEDP EVQFNWYVDGVEVHNAKTKPREEQFNSTYRWSVLTVLHQDWL NGKEYKCKVSNKGLPSSIEKTISKAKGQPREPQVYVYPPSQEEMT KNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSF ALVSKLTVDKSRWQEGNVFSCSVMHEALHNHYTQKSLSLSLGKKabat LKAMH 71 CDR1Kabat IISTYAGNTNYNQKFKG 95 CDR2Kabat GERFYYFDY 73 CDR332862 VH QVQLVQSGAEVKKPGSSVKVSCKGSGYTFTLKAMHWVRQAPGQ 226GLEWIGIISTYAGNTNYNQKFKGRATMTVDESTSTAYMELSSLRSEDTAVYYCARGERFYYFDYWGQGTTVTVSS146IPTS / 200238736.2Clone Region Sequence SEQ ID ID NO32863 Full QVQLVQSGAEVAKPGSSVKVSCKGSGYTFTLKAMHWVRQAPGQ 227GLEWIGIISTYAGNTNYNQKFKGRATMTVDESTSTAYMELSSLRS EDTAVYYCARGERFYYFDYWGQGTTVTVSSASTKGPSVFPLAPC SRSTSESTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLRS SGLYSLSSWTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKYGP PCPPCPAPEFLGGPSVFLFPPKPKDTLYITREPEVTCVWDVSQEDP EVQFNWYVDGVEVHNAKTKPREEQFNSTYRWSVLTVLHQDWL NGKEYKCKVSNKGLPSSIEKTISKAKGQPREPQVYVYPPSQEEMT KNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSF ALVSKLTVDKSRWQEGNVFSCSVMHEALHNHYTQKSLSLSLGKKabat LKAMH 71 CDR1Kabat IISTYAGNTNYNQKFKG 95 CDR2Kabat GERFYYFDY 73 CDR332863 VH QVQLVQSGAEVAKPGSSVKVSCKGSGYTFTLKAMHWVRQAPGQ 228GLEWIGIISTYAGNTNYNQKFKGRATMTVDESTSTAYMELSSLRS EDTAVYYCARGERFYYFDYWGQGTTVTVSS32864 Full EVQLVQSGAEVKQPGSSVKVSCAGSGYTFTLKAMHWVRQAPGQ 229GLEWIGIISTYAGNTNYNDQFKGRATMTVDTSTSTAYMELSSLRS EDTAVYYCARGERFYYFDYWGQGTTVTVSSASTKGPSVFPLAPC SRSTSESTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLRS SGLYSLSSWTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKYGP PCPPCPAPEFLGGPSVFLFPPKPKDTLYITREPEVTCVWDVSQEDP EVQFNWYVDGVEVHNAKTKPREEQFNSTYRWSVLTVLHQDWL NGKEYKCKVSNKGLPSSIEKTISKAKGQPREPQVYVYPPSQEEMT KNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSF ALVSKLTVDKSRWQEGNVFSCSVMHEALHNHYTQKSLSLSLGKKabat LKAMH 71 CDR1Kabat IISTYAGNTNYNDQFKG 99 CDR2Kabat GERFYYFDY 73 CDR332864 VH EVQLVQSGAEVKQPGSSVKVSCAGSGYTFTLKAMHWVRQAPGQ 230GLEWIGIISTYAGNTNYNDQFKGRATMTVDTSTSTAYMELSSLRSEDTAVYYCARGERFYYFDYWGQGTTVTVSS147IPTS / 200238736.2

Claims

WE CLAIM:

1. An antibody construct comprising one or more antigen-binding domains, wherein at least one of the antigen-binding domains is an IL-31 antigen-binding domain that specifically binds to human IL-31, the IL-31 antigen-binding domain comprising the CDR sequences (HCDR1, HCDR2, HCDR3) of the VH domain as set forth in any one of SEQ ID NOs: 47, 48, 49, 50, 51, 52, 53, 54, 55, 57, 58, 59, 60, 61 or 119, and the CDR sequences (LCDR1, LCDR2, LCDR3) of the VL domain as set forth in any one of SEQ ID NOs: 56, 62, 63, 64, 65 or 120.

2. The antibody construct according to claim 1, wherein the IL-31 antigen-binding domain comprises the CDR sequences (HCDR1, HCDR2, HCDR3) of the VH domain as set forth in SEQ ID NO: 59 or 61, and the CDR sequences (LCDR1, LCDR2, LCDR3) of the VL domain as set forth in any one of SEQ ID NO: 65.

3. The antibody construct according to claim 1, wherein the IL-31 antigen-binding domain comprises a VH domain comprising heavy chain CDR amino acid sequences (HCDR1, HCDR2 and HCDR3) comprising the sequences as set forth in SEQ ID NOs: 3, 4 and 5, and a VL domain comprising light chain CDR amino acid sequences (LCDR1, LCDR2 and LCDR3) comprising the sequences as set forth in SEQ ID NOs: 6, 7 and 8.

4. The antibody construct according to claim 1, wherein the IL-31 antigen-binding domain comprises a VH domain comprising heavy chain CDR amino acid sequences (HCDR1, HCDR2 and HCDR3) comprising the sequences as set forth in SEQ ID NOs: 9, W and 11, and a VL domain comprising light chain CDR amino acid sequences (LCDR1, LCDR2 and LCDR3) comprising the sequences as set forth in SEQ ID NOs: 12, 13 and 14.

5. The antibody construct according to claim 3, wherein X1is R, X2is G and X3is D.

6. The antibody construct according to claim 4, wherein X4is SW, X5is R, X6is RSR, X7is G, X8is S, and X9is D.

7. The antibody construct according to claim 1, wherein the IL-31 antigen-binding domain comprises:148IPTS / 200238736.2(a) a VH domain comprising an HCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 15 or 18, an HCDR2 amino acid sequence comprising the sequence as set forth in any one of SEQ ID NOs: 16, 19 or 37, and an HCDR3 amino acid sequence comprising the sequence as set forth in any one of SEQ ID NOs: 17, 20, 38 or 39, and(b) a VL domain comprising an LCDR1 amino acid sequence comprising the sequence as set forth in any one of SEQ ID NOs: 28, 31, 36 or 40, an LCDR2 amino acid sequence comprising the sequence as set forth in any one of SEQ ID NOs: 29, 32, 41, 44 or 45, and an LCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 30 or 42.

8. The antibody construct according to claim 1, wherein the IL-31 antigen-binding domain comprises:(a) a VH domain comprising an HCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 15 or 18, an HCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 16 or 19, and an HCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 17 or 20, and a VL domain comprising an LCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 28 or 31, an LCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 29 or 32, and an LCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 30, or(b) a VH domain comprising an HCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 15 or 18, an HCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 16 or 19, and an HCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 17 or 20, and a VL domain comprising an LCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 28 or 36, an LCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 29 or 32, and an LCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 30, or(c) a VH domain comprising an HCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 15 or 18, an HCDR2 amino acid sequence comprising the sequence149IPTS / 200238736.2as set forth in SEQ ID NO: 16 or 37, and an HCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 38 or 39, and a VL domain comprising an LCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 28 or 40, an LCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 29 or 41, and an LCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 42, or(d) a VH domain comprising an HCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 15 or 18, an HCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 16 or 19, and an HCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 38 or 39, and a VL domain comprising an LCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 28 or 40, an LCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 29 or 41, and an LCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 42, or(e) a VH domain comprising an HCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 15 or 18, an HCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 16 or 19, and an HCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 38 or 39, and a VL domain comprising an LCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 28 or 40, an LCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 29 or 41, and an LCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 30, or(f) a VH domain comprising an HCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 15 or 18, an HCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 16 or 19, and an HCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 38 or 39, and a VL domain comprising an LCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 28 or 40, an LCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 29 or150IPTS / 200238736.232, and an LCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 30, or(g) a VH domain comprising an HCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 15 or 18, an HCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 16 or 19, and an HCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 38 or 39, and a VL domain comprising an LCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 28 or 40, an LCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 44 or 45, and an LCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 30.

9. The antibody construct according to claim 1, wherein the IL-31 antigen-binding domain comprises a VH domain comprising an HCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 15 or 18, an HCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 16 or 19, and an HCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 38 or 39, and a VL domain comprising an LCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 28 or 40, an LCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 44 or 45, and an LCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 30.

10. The antibody construct according to any one of claims 1 to 9, wherein the IL-31 antigen binding domain comprises a VH domain comprising a sequence that is at least about 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to the VH sequence as set forth in any one of SEQ ID NOs: 47, 48, 49, 50, 51, 52, 53, 54, 55, 57, 58, 59, 60 or 61.

11. The antibody construct according to any one of claims 1 to 10, wherein the IL-31 antigen binding domain comprises a VL domain comprising a sequence that is at least about 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to the VL sequence as set forth in any one of SEQ ID NOs: 56, 62, 63, 64 or 65.

12. The antibody construct according to any one of claims 1, 3, 4 or 9, wherein the IL-31 antigen binding domain comprises a VH domain comprising a sequence that is at least about 90%,151IPTS / 200238736.295%, 96%, 97%, 98%, 99% or 100% identical to the VH sequence as set forth in SEQ ID NO: 59 or 61.

13. The antibody construct according to any one of claims 1, 3, 4, 9 or 12, wherein the IL-31 antigen binding domain comprises a VL domain comprising a sequence that is at least about 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the VL sequence as set forth in SEQ ID NO: 65.

14. The antibody construct according to any one of claims 1, 3, 4 or 9, wherein the IL-31 antigen binding domain comprises a VH domain comprising a sequence as set forth in SEQ ID NO: 59 or 61, and a VL domain comprising a sequence as set forth in SEQ ID NO: 65.

15. The antibody construct according to any one of claims 1 to 14, wherein the IL-31 antigen binding domain also binds to cynomolgus monkey IL-31.

16. The antibody construct according to any one of claims 1 to 15, wherein the IL-31 antigen-binding domain is a Fab or an scFv.

17. The antibody construct according to any one of claim 1 to 16, wherein the antibody construct comprises two, three or four antigen-binding domains.

18. The antibody construct according to any one of claims 1 to 17, wherein the antibody construct comprises two IL-31 antigen-binding domains.

19. The antibody construct according to claim 18, wherein the two IL-31 antigen-binding domains are the same.

20. The antibody construct according to any one of claims 1 to 19, further comprising a scaffold, wherein the IL-31 antigen-binding domain is operably linked to the scaffold.

21. The antibody construct according to claim 20, wherein the scaffold is an IgG Fc region.

22. The antibody construct according to claim 21, wherein the scaffold is a human IgG Fc region.

23. The antibody construct according to claim 21 or claim 22, wherein the IgG Fc region is an IgGl or IgG4 Fc region.152IPTS / 200238736.

224. The antibody construct according to any one of claims 1 to 17, wherein the antibody construct comprises a second target antigen-binding domain that specifically binds to a second target antigen, wherein the second target antigen is not IL-31.

25. The antibody construct according to claim 24, wherein the second target antigen is a cytokine or cytokine receptor.

26. The antibody construct according to claim 25, wherein the second target antigen is human IL-4Ra.

27. The antibody construct according to any one of claims 24 to 26, wherein the second target antigen-binding domain is a Fab or an scFv.

28. The antibody construct according to any one of claims 24 to 27, further comprising a scaffold, wherein the IL-31 antigen-binding domain and the second target antigen-binding domain are both operably linked to the scaffold.

29. The antibody construct according to claim 28, wherein the scaffold is an IgG Fc region.

30. The antibody construct according to claim 29, wherein the scaffold is a human IgG Fc region.

31. The antibody construct according to claim 29 or claim 30, wherein the IgG Fc region is an IgGl or IgG4 Fc region.

32. The antibody construct according to any one of claims 29 to 31, wherein the IgG Fc region is a heterodimeric Fc region comprising a first Fc polypeptide and a second Fc polypeptide.

33. The antibody construct according to claim 32, wherein the heterodimeric Fc region comprises a modified CH3 domain, and wherein the modified CH3 domain comprises one or more amino acid modifications that promote formation of the heterodimeric Fc over formation of a homodimeric Fc.

34. The antibody construct according to claim 33, wherein:153IPTS / 200238736.2(a) the first Fc polypeptide comprises the amino acid substitutions L351Y, F405A and Y407V, and the second Fc polypeptide comprises the amino acid substitutions T366L, K392M and T394W; or(b) the first Fc polypeptide comprises the amino acid substitutions L351Y, F405A and Y407V, and the second Fc polypeptide comprises the amino acid substitutions T366L, K392L and T394W; or(c) the first Fc polypeptide comprises the amino acid substitutions T350V, L351Y, F405A and Y407V, and the second Fc polypeptide comprises the amino acid substitutions T350V, T366L, K392M and T394W; or(d) the first Fc polypeptide comprises the amino acid substitutions T350V, L351Y, F405A and Y407V, and the second Fc polypeptide comprises the amino acid substitutions T350V, T366L, K392L and T394W; or(e) the first Fc polypeptide comprises the amino acid substitutions T350V, L351Y, S400E, F405A and Y407V, and the second Fc polypeptide comprises the amino acid substitutions T350V, T366L, N390R, K392M and T394W,and wherein the numbering of amino acids is EU numbering.

35. The antibody construct according to any one of claims 21 to 23 and 29 to 34, wherein the IgG Fc region comprises the amino acid substitutions M252Y, S254T and T256E, and wherein the numbering of amino acids is EU numbering.

36. The antibody construct according to any one of claims 21 to 23 and 29 to 35, wherein the IgG Fc region is an IgGl Fc region.

37. The antibody construct according to claim 36, wherein the IgGl Fc region comprises the amino acid substitutions L234A, L235A and D265S, and wherein the numbering of amino acids is EU numbering.

38. The antibody construct according to any one of claims 21 to 23 and 29 to 35, wherein the IgG Fc region is an IgG4 Fc region.154IPTS / 200238736.

239. The antibody construct according to claim 38, wherein the IgG4 Fc region comprises the amino acid substitution S228P and / or the amino acid substitution R409K.

40. A polynucleotide or set of polynucleotides encoding the antibody construct according to any one of claims 1 to 39.

41. An expression vector or set of expression vectors comprising the polynucleotide or set of polynucleotides according to claim 40.

42. A host cell comprising the polynucleotide or set of polynucleotides according to claim 40 or the expression vector or set of expression vectors according to claim 41.

43. A method of preparing the antibody construct according to any one of claims 1 to 39 comprising transfecting a host cell with the polynucleotide or set of polynucleotides according to claim 40 or the expression vector or set of expression vectors according to claim 41, and culturing the host cell under conditions suitable for expression of the antibody construct.

44. A multispecific antibody construct comprising an IL-31 antigen-binding domain that specifically binds to human IL-31, and one or more additional antigen-binding domains, wherein the one or more additional antigen-binding domains each specifically bind to an antigen other than IL-31, and wherein the IL-31 antigen-binding domain comprises the CDR sequences (HCDR1, HCDR2, HCDR3) of the VH domain as set forth in any one of SEQ ID NOs: 47, 48, 49, 50, 51, 52, 53, 54, 55, 57, 58, 59, 60, 61 or 119, and the CDR sequences (LCDR1, LCDR2, LCDR3) of the VL domain as set forth in any one of SEQ ID NOs: 56, 62, 63, 64, 65 or 120.

45. The multispecific antibody construct according to claim 44, wherein the IL-31 antigenbinding domain comprises the CDR sequences (HCDR1, HCDR2, HCDR3) of the VH domain as set forth in SEQ ID NO: 59 or 61, and the CDR sequences (LCDR1, LCDR2, LCDR3) of the VL domain as set forth in any one of SEQ ID NO: 65.

46. The multispecific antibody construct according to claim 44 or claim 45, wherein the IL-31 antigen-binding domain comprises a VH domain comprising heavy chain CDR amino acid sequences (HCDR1, HCDR2 and HCDR3) comprising the sequences as set forth in SEQ ID NOs:155IPTS / 200238736.23, 4 and 5, and a VL domain comprising light chain CDR amino acid sequences (LCDR1, LCDR2 and LCDR3) comprising the sequences as set forth in SEQ ID NOs: 6, 7 and 8.

47. The multispecific antibody construct according to any one of claims 44 to 46, wherein the IL-31 antigen-binding domain comprises a VH domain comprising heavy chain CDR amino acid sequences (HCDR1, HCDR2 and HCDR3) comprising the sequences as set forth in SEQ ID NOs: 9, 10 and 11, and a VL domain comprising light chain CDR amino acid sequences (LCDR1, LCDR2 and LCDR3) comprising the sequences as set forth in SEQ ID NOs: 12, 13 and 14.

48. The bispecific antibody construct according to claim 46, wherein X1is R, X2is G and X3is D.

49. The bispecific antibody construct according to claim 47, wherein X4is SW, X5is R, X6is RSR, X7is G, X8is S, and X9is D.

50. The bispecific antibody construct according to claim 44, wherein the IL-31 antigen-binding domain comprises:(a) a VH domain comprising an HCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 15 or 18, an HCDR2 amino acid sequence comprising the sequence as set forth in any one of SEQ ID NOs: 16, 19 or 37, and an HCDR3 amino acid sequence comprising the sequence as set forth in any one of SEQ ID NOs: 17, 20, 38 or 39, and (b) a VL domain comprising an LCDR1 amino acid sequence comprising the sequence as set forth in any one of SEQ ID NOs: 28, 31, 36 or 40, an LCDR2 amino acid sequence comprising the sequence as set forth in any one of SEQ ID NOs: 29, 32, 41, 44 or 45, and an LCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 30 or 42.

51. The multispecific antibody construct according to claim 44, wherein the IL-31 antigenbinding domain comprises:(a) a VH domain comprising an HCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 15 or 18, an HCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 16 or 19, and an HCDR3 amino acid sequence comprising the 156IPTS / 200238736.2sequence as set forth in SEQ ID NO: 17 or 20, and a VL domain comprising an LCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 28 or 31, an LCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 29 or 32, and an LCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 30, or(b) a VH domain comprising an HCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 15 or 18, an HCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 16 or 19, and an HCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 17 or 20, and a VL domain comprising an LCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 28 or 36, an LCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 29 or 32, and an LCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 30, or(c) a VH domain comprising an HCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 15 or 18, an HCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 16 or 37, and an HCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 38 or 39, and a VL domain comprising an LCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 28 or 40, an LCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 29 or 41, and an LCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 42, or(d) a VH domain comprising an HCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 15 or 18, an HCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 16 or 19, and an HCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 38 or 39, and a VL domain comprising an LCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 28 or 40, an LCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 29 or 41, and an LCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 42, or157IPTS / 200238736.2(e) a VH domain comprising an HCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 15 or 18, an HCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 16 or 19, and an HCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 38 or 39, and a VL domain comprising an LCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 28 or 40, an LCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 29 or 41, and an LCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 30, or(f) a VH domain comprising an HCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 15 or 18, an HCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 16 or 19, and an HCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 38 or 39, and a VL domain comprising an LCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 28 or 40, an LCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 29 or 32, and an LCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 30, or(g) a VH domain comprising an HCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 15 or 18, an HCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 16 or 19, and an HCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 38 or 39, and a VL domain comprising an LCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 28 or 40, an LCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 44 or 45, and an LCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 30.

52. The multispecific antibody construct according to claim 44, wherein the IL-31 antigenbinding domain comprises a VH domain comprising an HCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 15 or 18, an HCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 16 or 19, and an HCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 38 or 39, and a VL domain comprising an LCDR1 amino158IPTS / 200238736.2acid sequence comprising the sequence as set forth in SEQ ID NO: 28 or 40, an LCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 44 or 45, and an LCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 30.

53. The multispecific antibody construct according to any one of claims 44 to 52, wherein the IL-31 antigen binding domain comprises a VH domain comprising a sequence that is at least about 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the VH sequence as set forth in any one of SEQ ID NOs: 47, 48, 49, 50, 51, 52, 53, 54, 55, 57, 58, 59, 60 or 61.

54. The multispecific antibody construct according to any one of claims 44 to 53, wherein the IL-31 antigen binding domain comprises a VL domain comprising a sequence that is at least about 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the VL sequence as set forth in any one of SEQ ID NOs: 56, 62, 63, 64 or 65.

55. The multispecific antibody construct according to any one of claims 44 to 52, wherein the IL-31 antigen binding domain comprises a VH domain comprising a sequence that is at least about 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the VH sequence as set forth in SEQ ID NO: 59 or 61.

56. The multispecific antibody construct according to any one of claims 44 to 52 and 55, wherein the IL-31 antigen binding domain comprises a VL domain comprising a sequence that is at least about 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the VL sequence as set forth in SEQ ID NO: 65.

57. The multispecific antibody construct according to any one of claims 44 to 52, wherein the IL-31 antigen binding domain comprises a VH domain comprising a sequence as set forth in SEQ ID NO: 59 or 61, and a VL domain comprising a sequence as set forth in SEQ ID NO: 65.

58. The multispecific antibody construct according to any one of claims 44 to 57, wherein the IL-31 antigen binding domain also binds to cynomolgus monkey IL-31.

59. The multispecific antibody construct according to any one of claims 44 to 58, wherein the IL-31 antigen-binding domain is a Fab or an scFv.159IPTS / 200238736.

260. The multispecific antibody construct according to any one of claims 44 to 59 comprising at least a second target antigen-binding domain that binds to a second target antigen, wherein the second target antigen is a cytokine or cytokine receptor.

61. The multispecific antibody construct according to claim 60, wherein the second target antigen is human IL-4Ra.

62. The multispecific antibody construct according to claim 61, wherein the second target antigen-binding domain comprises the CDR sequences of the VH domain as set forth in any one of SEQ ID NOs: 103, 104, 106, 107, 108, 109, 110 or 219, and the CDR sequences of the VL domain as set forth in any one of SEQ ID NOs: 105, 111, 112, 113 or 114.

63. The multispecific antibody construct according to any one of claims 60 to 62, wherein the second target antigen-binding domain comprises a VH domain comprising an HCDR1 amino acid sequence comprising the sequence as set forth in any one of SEQ ID NOs: 68, 71, 81 or 82, an HCDR2 amino acid sequence comprising the sequence as set forth in any one of SEQ ID NOs: 69, 72, 94, 95, 99, 100, 101, 102 or 218, and an HCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 70 or 73, and an LCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 86 or 89, an LCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 87 or 90, and an LCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 88.

64. The multispecific antibody construct according to claim 63, wherein the VH domain comprises an HCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 68 or 71, an HCDR2 amino acid sequence comprising the sequence as set forth in any one of SEQ ID NOs: 69, 72, 94, 95, 99, 100, 101, 102 or 218, and an HCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 70 or 73.

65. The multispecific antibody construct according to any one of claims 60 to 64, wherein the second target antigen-binding domain comprises a VH domain comprising:(i) an HCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 68 or 71, an HCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID160IPTS / 200238736.2NO: 69 or 72, and an HCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 70 or 73, or(ii) an HCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 81 or 82, an HCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 69 or 72, and an HCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 70 or 73, or(iii) an HCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 68 or 71, an HCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 94 or 95, and an HCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 70 or 73, or(iv) an HCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 68 or 71, an HCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 94 or 99, and an HCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 70 or 73, or(v) an HCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 68 or 71, an HCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 94 or 100, and an HCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 70 or 73, or(vi) an HCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 68 or 71, an HCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 69 or 101, and an HCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 70 or 73, or(vii) an HCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 68 or 71, an HCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 69 or 102, and an HCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 70 or 73, or161IPTS / 200238736.2(viii) an HCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 68 or 71, an HCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 94 or 218, and an HCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 70 or 73, anda VL domain comprising an LCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 86 or 89, an LCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 87 or 90, and an LCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 88.

66. The multispecific antibody construct according to any one of claims 60 to 65, wherein the second target antigen-binding domain comprises a VH domain having a sequence that is at least about 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the VH sequence as set forth in any one ofSEQ IDNOs: 103, 104, 106, 107, 108, 109, 110 or 219, and a VL domain having a sequence that is at least about 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the VL sequence as set forth in any one of SEQ ID NOs: 105, 111, 112, 113 or 114.

67. The multispecific antibody construct according to any one of claims 60 to 65, wherein the second target antigen-binding domain comprises a VH domain having a sequence as set forth in any SEQ ID NO: 219, and a VL domain having a sequence as set forth in SEQ ID NO:

113.

68. The multispecific antibody construct according to any one of claims 60 to 67, wherein the second target antigen-binding domain is a Fab or an scFv.

69. The multispecific antibody construct according to any one of claims 44 to 67, further comprising a scaffold, wherein the IL-31 antigen-binding domain and the one or more additional antigen-binding domains are operably linked to the scaffold.

70. The multispecific antibody construct according to claim 69, wherein the scaffold is an IgG Fc region.

71. The multispecific antibody construct according to claim 69 or claim 70, wherein the scaffold is a human IgG Fc region.162IPTS / 200238736.

272. The multispecific antibody construct according to claim 70 or claim 71, wherein the IgG Fc region is an IgGl or IgG4 Fc region.

73. The multispecific antibody construct according to any one of claims 70 to 72, wherein the IgG Fc region is a heterodimeric Fc region comprising a first Fc polypeptide and a second Fc polypeptide.

74. The multispecific antibody construct according to claim 73, wherein the heterodimeric Fc region comprises a modified CH3 domain, and wherein the modified CH3 domain comprises one or more amino acid modifications that promote formation of the heterodimeric Fc over formation of a homodimeric Fc.

75. The multispecific antibody construct according to claim 74, wherein:(a) the first Fc polypeptide comprises the amino acid substitutions L351Y, F405A and Y407V, and the second Fc polypeptide comprises the amino acid substitutions T366L, K392M and T394W; or(b) the first Fc polypeptide comprises the amino acid substitutions L351Y, F405A and Y407V, and the second Fc polypeptide comprises the amino acid substitutions T366L, K392L and T394W; or(c) the first Fc polypeptide comprises the amino acid substitutions T350V, L351Y, F405A and Y407V, and the second Fc polypeptide comprises the amino acid substitutions T350V, T366L, K392M and T394W; or(d) the first Fc polypeptide comprises the amino acid substitutions T350V, L351Y, F405A and Y407V, and the second Fc polypeptide comprises the amino acid substitutions T350V, T366L, K392L and T394W; or(e) the first Fc polypeptide comprises the amino acid substitutions T350V, L351Y, S400E, F405A and Y407V, and the second Fc polypeptide comprises the amino acid substitutions T350V, T366L, N390R, K392M and T394W,and wherein the numbering of amino acids is EU numbering.163IPTS / 200238736.

276. The multispecific antibody construct according to any one of claims 70 to 75, wherein the IgG Fc region comprises the amino acid substitutions M252Y, S254T and T256E, and wherein the numbering of amino acids is EU numbering.

77. The multispecific antibody construct according to any one of claims 70 to 76, wherein the IgG Fc region is an IgGl Fc region.

78. The multispecific antibody construct according to claim 77, wherein the IgGl Fc region comprises the amino acid substitutions L234A, L235A and D265S, and wherein the numbering of amino acids is EU numbering.

79. The multispecific antibody construct according to any one of claims 70 to 76, wherein the IgG Fc region is an IgG4 Fc region.

80. The multispecific antibody construct according to claim 79, wherein the IgG4 Fc region comprises the amino acid substitution S228P and / or the amino acid substitution R409K.

81. A multispecific antibody construct comprising an IL-31 antigen-binding domain that specifically binds to human IL-31, and one or more additional antigen-binding domains, wherein the one or more additional antigen-binding domains each specifically bind to an antigen other than IL-31,wherein the IL-31 antigen-binding domain comprises a VH domain comprising an HCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 15 or 18, an HCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 16 or 19, and an HCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 38 or 39, and a VL domain comprising an LCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 28 or 40, an LCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 44 or 45, and an LCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 30, andwherein the one or more additional antigen-binding domains comprise at least a second target antigen-binding domain that specifically binds to IL-4Ra, wherein the second target antigen-binding domain comprises an HCDR1 amino acid sequence comprising the164IPTS / 200238736.2sequence as set forth in SEQ ID NO: 68 or 71, an HCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 94 or 218, and an HCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 70 or 73, and a VL domain comprising an LCDR1 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 86 or 89, an LCDR2 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 87 or 90, and an LCDR3 amino acid sequence comprising the sequence as set forth in SEQ ID NO: 88.

82. A polynucleotide or set of polynucleotides encoding the multispecific antibody construct according to any one of claims 44 to 81.

83. An expression vector or set of expression vectors comprising the polynucleotide or set of polynucleotides according to claim 82.

84. A host cell comprising the polynucleotide or set of polynucleotides according to claim 82 or the expression vector or set of expression vectors according to claim 83.

85. A method of preparing the multispecific antibody construct according to any one of claims 44 to 81 comprising transfecting a host cell with the polynucleotide or set of polynucleotides according to claim 82 or the expression vector or set of expression vectors according to claim 83, and culturing the host cell under conditions suitable for expression of the antibody construct.

86. A pharmaceutical composition comprising the antibody construct according to any one of claims 1 to 39 or the multispecific antibody construct according to any one of claims 44 to 81.

87. An antibody construct according to any one of claims 1 to 39 or a multispecific antibody construct according to any one of claims 44 to 81 for use in therapy.

88. Use of an antibody construct according to any one of claims 1 to 39 or a multispecific antibody construct according to any one of claims 44 to 81 in therapy.

89. Use of an antibody construct according to any one of claims 1 to 39 or a multispecific antibody construct according to any one of claims 44 to 81 in the manufacture of a medicament.165IPTS / 200238736.

290. A method of alleviating pruritis in a subject comprising administering to the subject an effective amount of the antibody construct according to any one of claims 1 to 39 or the multispecific antibody construct according to any one of claims 44 to 81.

91. A method of treating an allergic, inflammatory or autoimmune disease in a subject comprising administering to the subject an effective amount of the antibody construct according to any one of claims 1 to 39 or the multispecific antibody construct according to any one of claims 44 to 81.

92. An antibody construct according to any one of claims 1 to 39 or a multispecific antibody construct according to any one of claims 44 to 81 for use to alleviate pruritis in a subject.

93. An antibody construct according to any one of claims 1 to 39 or a multispecific antibody construct according to any one of claims 44 to 81 for use in the treatment of an allergic, inflammatory or autoimmune disease in a subject.

94. Use of an antibody construct according to any one of claims 1 to 39 or a multispecific antibody construct according to any one of claims 44 to 81 to alleviate pruritis in a subject.

95. Use of an antibody construct according to any one of claims 1 to 39 or a multispecific antibody construct according to any one of claims 44 to 81 in the treatment of an allergic, inflammatory or autoimmune disease in a subject.

96. Use of an antibody construct according to any one of claims 1 to 39 or a multispecific antibody construct according to any one of claims 44 to 81 in the manufacture of a medicament to alleviate pruritis.

97. Use of an antibody construct according to any one of claims 1 to 39 or a multispecific antibody construct according to any one of claims 44 to 81 in the manufacture of a medicament for the treatment of an allergic, inflammatory or autoimmune disease.166IPTS / 200238736.2

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