Anti-IL2 receptor gamma antigen binding proteins

Abstract

The invention provides anti-IL2 receptor gamma antigen binding proteins. The present invention relates to antibodies that bind to the anti-IL2 receptor gamma protein and methods of use thereof. The present invention provides antibodies and antigen-binding fragments (e.g., human antibodies) that specifically bind to human IL2 receptor gamma (IL2R gamma). Also provided are methods for using the antibodies and fragments to treat or prevent diseases mediated by IL2R gamma (e.g., graft versus host disease) and methods of making the antibodies and fragments.

Description

[0001] This application is a divisional application with the same invention title as the parent application. The Chinese application number of the parent application is 202080012091.4, the international application number is PCT / US2020 / 015841, and the filing date is January 30, 2020.

[0002] This application claims the benefit of U.S. Provisional Patent Application No. 62 / 799,851, filed on February 1, 2019, the entire content of which is incorporated herein by reference. TECHNICAL FIELD

[0003] The present invention relates to antibodies that bind to the anti-IL2 receptor γ protein and methods of using the same, such as methods for treating or preventing diseases. BACKGROUND ART

[0004] The common cytokine receptor γ chain (γc) was initially identified as the third chain of the interleukin-2 (IL-2) receptor complex and named IL-2Rγ. The same subunit was identified as part of several other cytokine receptor complexes: IL-4, IL-7, IL-9, IL-15, and IL-21, and can therefore be referred to as γc (common cytokine receptor γ chain). γc is involved in signal transduction and ligand binding of these cytokine receptors.

[0005] Binding of a cytokine to its receptor activates the Janus kinase (JAK) family of protein tyrosine kinases JAK1 and JAK3, and triggers the transphosphorylation of tyrosine by JAK1 and JAK3. JAK1 associates with the unique α or β chain of the receptor, and JAK3 associates with the γc of the receptor. The phosphorylated JAKs in turn can activate signal transducer and activator of transcription (STAT) proteins, which together form the JAK / STAT signaling pathway. Phosphorylation of STAT results in dimerization of STAT, which now adopts high-affinity DNA binding activity and translocates to the nucleus. Here, it acts as a transcription factor to induce transcription of target genes.

[0006] The γc gene (IL2RG) is located on chromosome Xq13. IL-2Rγ is mutated in patients with X-linked severe combined immunodeficiency (X-SCID). Due to the lack of T cells, NK cells, and fully mature B cells, patients with this disease exhibit severe immunodeficiency.

[0007] IL-7, IL-9, and IL-15 are associated with psoriasis and rheumatoid arthritis (Pathak, The expanding role of IL-7 and thymic stromal lymphopoietin as therapeutic target for rheumatoid arthritis. Expert Opin Ther Targets. 18(5):581-94(2014); Hughes-Austin et al., Multiple cytokines and chemokines are associated with rheumatoid arthritis-related autoimmunity in first-degree relatives without rheumatoid arthritis: Studies of the Aetiology of Rheumatoid Arthritis (SERA), Ann Rheum Dis.; 72(6):901-7(2013); Dantas et al., Increased Serum Interleukin-9 Levels in Rheumatoid Arthritis and Systemic Lupus Erythematosus: Pathogenic Role or Just an Epiphenomenon?, Dis Markers. 2015;2015:519638; Yang et al., Therapeutic potential of IL-15 in rheumatoid arthritis, Hum Immunol. 2015 Nov;76(11):812-8; Lesiak et al., Are interleukin-15 and -22 a new pathogenic factor in pustular palmoplantar psoriasis?, Postepy Dermatol Alergol. 33(5):336-339(2016); Raeber et al., The role of cytokines in T-cell memory in health and disease, Immunol Rev. 283(1):176-193(2018)).

[0008] IL-4 and IL-9 blockade has been shown to improve asthma symptoms in mice (Generoso et al., Prospects for Monoclonal Antibody Therapy in Pediatric Asthma, Curr Allergy Asthma Rep. 18(9):45(2018); Tashkin & Wechsler, Role of eosinophils in airway inflammation of chronic obstructive pulmonary disease, Int J Chron Obstruct Pulmo B Dis. 13:335-349(2018); Buzney et al., Asthma and Atopic Dermatitis: A Review of Targeted Inhibition of Interleukin-4 and Interleukin-13 As Therapy for Atopic Disease, J Drugs Dermatol. 15(2):165-71(2016); Lloyd & Harker, Epigenetic Control of Interleukin-9 in Asthma, N Engl J Med. 379(1):87-89(2018); Neurath & Finotto, IL-9 signaling as key driver of chronic inflammation in mucosal immunity, Cytokine Growth Factor Rev. 29:93-9(2016)).

[0009] IL-21 is associated with a variety of inflammatory disorders, including Crohn's disease and rheumatoid arthritis. (Holm et al., Evaluating IL-21 as a Potential Therapeutic Target in Crohn's Disease, Gastroenterol Res Pract. 2018:5962624 (2018); Dinesh & Rasool Multifaceted role of IL-21 in rheumatoid arthritis: Current understanding and future perspectives, J Cell Physiol. 233(5):3918 - 3928 (2018)). Summary of the Invention

[0010] The present invention provides an isolated antigen-binding protein (e.g., an antibody or an antigen-binding fragment thereof, e.g., which is monospecific or multispecific), characterized by one or more of the following: having a K at about 2.75×10 -9 M to about 3.36×10 -7 M for binding to human IL2Rγ at 25°C; having a K at about 6.42×10 D M to about 3.53×10 -9 M for binding to human IL2Rγ at 37°C; or having a K less than about 3.53×10 -7 M for binding; having a K at about 3.18×10 D M to about 2.38×10 -7 M for binding to Macaca fascicularis IL-2Rγ at 25°C; having a K at about 8.29×10 D M to about 3.20×10 -9 M for binding to Macaca fascicularis IL-2Rγ at 37°C; or having a K less than about 3.20×10 -7 M for binding; having a K at about 2.45×10 D M to about 1.20×10 -9 M for binding to human IL2Rγ at 25°C; or having a K less than about 1.20×10 -7 M for binding; having a K at about 1.86×10 D M to about 3.00×10 -7 M for binding; or having a K less than about 1.20×10 D M for binding; having a K at about 2.45×10 -9 M to about 1.20×10 -8 M for binding to human IL2Rγ at 25°C; or having a K less than about 1.20×10 D M for binding; having a K at about 1.86×10 -8 M to about 3.00×10 D M for binding at 37°C; having a K at about 1.86×10 -11 M to about 3.00×10-8 K of M D binds to human IL2Rγ; or binds with a K less than about 3.00×10 -8 K of M D binds; at 25 °C with a K of about 1.84×10 -8 M, 3.76×10 -9 M, 1.08×10 -7 M, 2.17×10 -8 M, 6.02×10 -9 M or 7.93×10 -8 K of M D binds to mouse IL2Rγ; or has no detectable binding; at 37 °C with a K of about 5.59×10 -8 M, 6.11×10 -9 M, 3.87×10 - 7 M, 5.16×10 -8 M, 8.70×10 -9 M or 2.15×10 -7 K of M D binds to mouse IL2Rγ; or has no detectable binding; at 25 °C with a K from about 3.32×10 -9 M to about 1.97×10 -7 K of M D binds to human IL2Rγ domain 1; or has no detectable binding; at 37 °C with a K from about 4.13×10 -9 M to about 2.25×10 -7 K of M D binds to human IL2Rγ domain 1; or has no detectable binding; at 25 °C with a K from about 2.91×10 -7 M to about 5.35×10 -10 K of D binds to human IL2Rγ domain 2; or has no detectable binding; at 37 °C with a K of about 1.14×10 -8 or about 1.27×10 -8 K of DBind to human IL2Rγ domain 2; or have no detectable binding; block STAT phosphorylation in T cells induced by IL-2, IL-4, IL7, IL-15, and / or IL-21; block STAT phosphorylation in mast cells induced by IL-9; reduce the number of human immune cells injected into mice; reduce the levels of human cytokines and / or murine serum cytokines in mice with human immune cells; have no detectable binding to murine or rat IL2Rγ; protect mice from weight loss and / or death due to GvHD in a GvHD mouse model; block the binding of heteromeric receptors containing IL2Rγ complexed with cytokine-specific receptor subunits to IL-2, IL-4, IL-7, IL-9, IL-15, and / or IL-21; and / or reduce the number of CD45+ cells, B cells, T cells, and / or NK cells (but optionally not, for example, neutrophils) in the blood or serum of a subject. Antibodies and antigen-binding fragments that specifically bind to IL2Rγ as described below form part of the present invention, which are variants of any of the antibodies or fragments whose sequences are specifically shown herein and which are characterized by one or more of the above features.

[0011] The present invention also provides the following isolated antigen-binding proteins, e.g., antibodies or antigen-binding fragments thereof, which antigen-binding proteins: (i) specifically bind to the same epitope on IL2Rγ as a reference antibody or antigen-binding fragment thereof; or (ii) compete with a reference antibody or antigen-binding fragment thereof for binding to an IL2Rγ polypeptide, wherein the reference antibody or antigen-binding fragment thereof comprises: (a) an immunoglobulin heavy chain or variable region thereof, which comprises CDR-H1, CDR-H2 and CDR-H3 of an immunoglobulin heavy chain or variable region thereof containing the amino acid sequence shown in SEQ ID NO: 2, 18, 22, 38, 42, 58, 62, 77, 81, 97, 101, 115, 119, 134, 138, 152, 156, 170, 174, 186, 190, 198, 200, 208, 210, 216, 218, 234, 238, 254, 258, 272, 276, 284, 286, 294, 296, 311, 315, 331, 335, 343, 345, 357, 361 and / or 376 or a variant thereof; and / or (b) an immunoglobulin light chain or variable region thereof, which comprises CDR-L1, CDR-L2 and CDR-L3 of an immunoglobulin light chain or variable region thereof containing the amino acid sequence shown in SEQ ID NO: 10, 20, 30, 40, 50, 60, 70, 79, 89, 99, 109, 117, 127, 136, 146, 154, 164, 172, 182, 188, 226, 236, 246, 256, 266, 274, 304, 313, 323, 333, 353, 359, 368 and / or 378 or a variant thereof. In one embodiment of the present invention, the reference antibody or fragment is pre-bound to the IL2Rγ antigen before addition of the antigen-binding protein and evaluated for binding. In one embodiment of the present invention, the antigen-binding protein is pre-bound to the antigen before addition of the reference antibody or fragment and evaluated for binding.

[0012] The present invention also provides the following isolated antigen-binding proteins (e.g., antibodies or antigen-binding fragments thereof), which comprise: (a) an immunoglobulin heavy chain or variable region thereof, which comprises CDR-H1, CDR-H2, and CDR-H3 of an immunoglobulin heavy chain or variable region thereof that contains the amino acid sequence shown in SEQ ID NO: 2, 18, 22, 38, 42, 58, 62, 77, 81, 97, 101, 115, 119, 134, 138, 152, 156, 170, 174, 186, 190, 198, 200, 208, 210, 216, 218, 234, 238, 254, 258, 272, 276, 284, 286, 294, 296, 311, 315, 331, 335, 343, 345, 357, 361, and / or 376 or a variant thereof; and / or (b) an immunoglobulin light chain or variable region thereof, which comprises CDR-L1, CDR-L2, and CDR-L3 of an immunoglobulin light chain or variable region thereof that contains the amino acid sequence shown in SEQ ID NO: 10, 20, 30, 40, 50, 60, 70, 79, 89, 99, 109, 117, 127, 136, 146, 154, 164, 172, 182, 188, 226, 236, 246, 256, 266, 274, 304, 313, 323, 333, 353, 359, 368, and / or 378 or a variant thereof.

[0013] In one embodiment of the present invention, it comprises (a) an immunoglobulin heavy chain or its variable region, which comprises an amino acid sequence having at least 90% amino acid sequence identity with the amino acid sequences shown in SEQ ID NO: 2, 18, 22, 38, 42, 58, 62, 77, 81, 97, 101, 115, 119, 134, 138, 152, 156, 170, 174, 186, 190, 198, 200, 208, 210, 216, 218, 234, 238, 254, 258, 272, 276, 284, 286, 294, 296, 311, 315, 331, 335, 343, 345, 357, 361 and / or 376; and / or (b) an immunoglobulin light chain or its variable region, which comprises an amino acid sequence having at least 90% amino acid sequence identity with the amino acid sequences shown in SEQ ID NO: 10, 20, 30, 40, 50, 60, 70, 79, 89, 99, 109, 117, 127, 136, 146, 154, 164, 172, 182, 188, 226, 236, 246, 256, 266, 274, 304, 313, 323, 333, 353, 359, 368 and / or 378.For example, in one embodiment of the present invention, the antigen-binding protein comprises: (a) an immunoglobulin heavy chain or its variable region, which comprises CDR-H1, CDR-H2 and CDR-H3 of an immunoglobulin heavy chain or its variable region containing the amino acid sequences shown in SEQ ID NO: 2, 18, 22, 38, 42, 58, 62, 77, 81, 97, 101, 115, 119, 134, 138, 152, 156, 170, 174, 186, 190, 198, 200, 208, 210, 216, 218, 234, 238, 254, 258, 272, 276, 284, 286, 294, 296, 311, 315, 331, 335, 343, 345, 357, 361 and / or 376 and amino acid sequences having at least 90% amino acid sequence identity with the amino acid sequences shown in SEQ ID NO: 2, 18, 22, 38, 42, 58, 62, 77, 81, 97, 101, 115, 119, 134, 138, 152, 156, 170, 174, 186, 190, 198, 200, 208, 210, 216, 218, 234, 238, 254, 258, 272, 276, 284, 286, 294, 296, 311, 315, 331, 335, 343, 345, 357, 361 and / or 376; and / or (b) an immunoglobulin light chain or its variable region, which comprises CDR-L1, CDR-L2 and CDR-L3 of an immunoglobulin light chain or its variable region containing the amino acid sequences shown in SEQ ID NO: 10, 20, 30, 40, 50, 60, 70, 79, 89, 99, 109, 117, 127, 136, 146, 154, 164, 172, 182, 188, 226, 236, 246, 256, 266, 274, 304, 313, 323, 333, 353, 359, 368 and / or 378 and amino acid sequences having at least 90% amino acid sequence identity with the amino acid sequences shown in SEQ ID NO: 10, 20, 30, 40, 50, 60, 70, 79, 89, 99, 109, 117, 127, 136, 146, 154, 164, 172, 182, 188, 226, 236, 246, 256, 266, 274, 304, 313, 323, 333, 353, 359, 368 and / or 378.

[0014] In one embodiment of the present invention, the antigen-binding protein comprises:

[0015] (i) Heavy chain CDR set: CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 4, CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 6, and CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 8; and / or CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 24, CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 26, and CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 28; and / or CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 44, CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 46, and CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 48; and / or CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 64, CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 66, and CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 68; and / or CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 83, CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 85, and CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 87; and / or CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 103, CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 105, and CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 107; and / or CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 121, CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 123, and CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 125; and / or CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 140, CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 142, and CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 144; and / or CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 158, CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 160, and CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 162; and / or CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 176, CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 178, and CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 180;and / or CDR-H1 comprising the amino acid sequence set forth in SEQ ID NO: 192, CDR-H2 comprising the amino acid sequence set forth in SEQ ID NO: 194, and CDR-H3 comprising the amino acid sequence set forth in SEQ ID NO: 196; and / or CDR-H1 comprising the amino acid sequence set forth in SEQ ID NO: 202, CDR-H2 comprising the amino acid sequence set forth in SEQ ID NO: 204, and CDR-H3 comprising the amino acid sequence set forth in SEQ ID NO: 206; and / or CDR-H1 comprising the amino acid sequence set forth in SEQ ID NO: 176, CDR-H2 comprising the amino acid sequence set forth in SEQ ID NO: 212, and CDR-H3 comprising the amino acid sequence set forth in SEQ ID NO: 214; and / or CDR-H1 comprising the amino acid sequence set forth in SEQ ID NO: 220, CDR-H2 comprising the amino acid sequence set forth in SEQ ID NO: 222, and CDR-H3 comprising the amino acid sequence set forth in SEQ ID NO: 224; and / or CDR-H1 comprising the amino acid sequence set forth in SEQ ID NO: 240, CDR-H2 comprising the amino acid sequence set forth in SEQ ID NO: 242, and CDR-H3 comprising the amino acid sequence set forth in SEQ ID NO: 244; and / or CDR-H1 comprising the amino acid sequence set forth in SEQ ID NO: 260, CDR-H2 comprising the amino acid sequence set forth in SEQ ID NO: 262, and CDR-H3 comprising the amino acid sequence set forth in SEQ ID NO: 264; and / or CDR-H1 comprising the amino acid sequence set forth in SEQ ID NO: 278, CDR-H2 comprising the amino acid sequence set forth in SEQ ID NO: 280, and CDR-H3 comprising the amino acid sequence set forth in SEQ ID NO: 282; and / or CDR-H1 comprising the amino acid sequence set forth in SEQ ID NO: 288, CDR-H2 comprising the amino acid sequence set forth in SEQ ID NO: 290, and CDR-H3 comprising the amino acid sequence set forth in SEQ ID NO: 292; and / or CDR-H1 comprising the amino acid sequence set forth in SEQ ID NO: 298, CDR-H2 comprising the amino acid sequence set forth in SEQ ID NO: 300, and CDR-H3 comprising the amino acid sequence set forth in SEQ ID NO: 302; and / or CDR-H1 comprising the amino acid sequence set forth in SEQ ID NO: 317, CDR-H2 comprising the amino acid sequence set forth in SEQ ID NO: 319, and CDR-H3 comprising the amino acid sequence set forth in SEQ ID NO: 321;and / or CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 337, CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 339, and CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 341; and / or CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 347, CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 349, and CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 351; and / or CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 363, CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 66, and CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 366; and / or;

[0016] (ii) Light chain CDR sets: CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 12, CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 14, and CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 16; and / or CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 32, CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 34, and CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 36; and / or CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 52, CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 54, and CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 56; and / or CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 72, CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 54, and CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 75; and / or CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 91, CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 93, and CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 95; and / or CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 111, CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 54, and CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 113; and / or CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 129, CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 54, and CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 132; and / or CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 148, CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 54, and CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 150; and / or CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 166, CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 14, and CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 168; and / or CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 72, CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 54, and CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 184;and / or CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 72, CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 54, and CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 184; and / or CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 72, CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 54, and CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 184; and / or CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 72, CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 54, and CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 184; and / or CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 228, CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 230, and CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 232; and / or CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 248, CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 250, and CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 252; and / or CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 268, CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 54, and CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 270; and / or CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 72, CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 54, and CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 184; and / or CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 72, CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 54, and CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 184; and / or CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 306, CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 230, and CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 309; and / or CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 325, CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 327, and CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 329;and / or CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 72, CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 54, and CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 184; and / or CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 72, CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 54, and CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 355; and / or CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 370, CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 372, and CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 374.;

[0017] In one embodiment of the present invention, the antigen-binding protein of the present invention comprises the following heavy-chain CDR set and light-chain CDR set:

[0018] (i) A heavy chain variable region comprising a CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 4, a CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 6, and a CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 8; and a light chain variable region comprising a CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 12, a CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 14, and a CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 16; (ii) A heavy chain variable region comprising a CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 24, a CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 26, and a CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 28; and a light chain variable region comprising a CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 32, a CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 34, and a CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 36; (iii) A heavy chain variable region comprising a CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 44, a CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 46, and a CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 48; and a light chain variable region comprising a CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 52, a CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 54, and a CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 56; (iv) A heavy chain variable region comprising a CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 64, a CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 66, and a CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 68; and a light chain variable region comprising a CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 72, a CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 54, and a CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 75; (v) A heavy chain variable region comprising a CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 83, a CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 85, and a CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 87;and a light chain variable region comprising CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 91, CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 93, and CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 95; (vi) a heavy chain variable region comprising CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 103, CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 105, and CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 107; and a light chain variable region comprising CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 111, CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 54, and CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 113; (vi) a heavy chain variable region comprising CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 121, CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 123, and CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 125; and a light chain variable region comprising CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 129, CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 54, and CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 132; (vii) a heavy chain variable region comprising CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 140, CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 142, and CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 144; and a light chain variable region comprising CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 148, CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 54, and CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 150; (viii) a heavy chain variable region comprising CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 158, CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 160, and CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 162; and a light chain variable region comprising CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 166, CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 14, and CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 168;(ix) A heavy chain variable region comprising a CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 176, a CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 178, and a CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 180; and a light chain variable region comprising a CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 72, a CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 54, and a CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 184; (x) A heavy chain variable region comprising a CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 192, a CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 194, and a CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 196; and a light chain variable region comprising a CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 72, a CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 54, and a CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 184; (xi) A heavy chain variable region comprising a CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 202, a CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 204, and a CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 206; and a light chain variable region comprising a CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 72, a CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 54, and a CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 184; (xii) A heavy chain variable region comprising a CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 176, a CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 212, and a CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 214; and a light chain variable region comprising a CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 72, a CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 54, and a CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 184; (xiii) A heavy chain variable region comprising a CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 220, a CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 222, and a CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 224;and a light chain variable region comprising CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 228, CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 230, and CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 232; (xiv) a heavy chain variable region comprising CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 240, CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 242, and CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 244; and a light chain variable region comprising CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 248, CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 250, and CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 252; (xv) a heavy chain variable region comprising CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 260, CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 262, and CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 264; and a light chain variable region comprising CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 268, CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 54, and CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 270; (xvi) a heavy chain variable region comprising CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 278, CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 280, and CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 282; and a light chain variable region comprising CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 72, CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 54, and CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 184; (xvii) a heavy chain variable region comprising CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 288, CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 290, and CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 292; and a light chain variable region comprising CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 72, CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 54, and CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 184;(xviii) A heavy chain variable region comprising a CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 298, a CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 300, and a CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 302; and a light chain variable region comprising a CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 306, a CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 230, and a CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 309; (xix) A heavy chain variable region comprising a CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 317, a CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 319, and a CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 321; and a light chain variable region comprising a CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 325, a CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 327, and a CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 329; (xx) A heavy chain variable region comprising a CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 337, a CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 339, and a CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 341; and a light chain variable region comprising a CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 72, a CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 54, and a CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 184; (xxi) A heavy chain variable region comprising a CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 347, a CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 349, and a CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 351; and a light chain variable region comprising a CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 72, a CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 54, and a CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 355; (xxii) A heavy chain variable region comprising a CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 363, a CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 66, and a CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 366;and a light chain variable region comprising CDR-L1 having the amino acid sequence shown in SEQ ID NO: 370, CDR-L2 having the amino acid sequence shown in SEQ ID NO: 372, and CDR-L3 having the amino acid sequence shown in SEQ ID NO: 374.;

[0019] A complex comprising an antigen-binding protein of the invention that binds to an IL2Rγ polypeptide or an antigenic fragment thereof is also part of the invention.

[0020] The invention also provides methods for preparing an antigen-binding protein (e.g., an antibody or an antigen-binding fragment thereof) or an immunoglobulin chain thereof (e.g., V H 、V L 、HC or LC), comprising: (a) introducing into a host cell (e.g., a CHO cell) one or more polynucleotides encoding one or more immunoglobulin chains of the antigen-binding protein (or a vector comprising such polynucleotides); (b) culturing the host cell under conditions favorable for the expression of the polynucleotide; and (c) optionally, isolating the antigen-binding protein or immunoglobulin chain from the host cell and / or the medium in which the host cell was cultured. An antigen-binding protein or immunoglobulin chain that is a product of such a method also constitutes part of the invention.

[0021] The present invention also provides polypeptides, which comprise: (a) CDR-H1, CDR-H2 and CDR-H3 of an immunoglobulin heavy chain or its variable region containing the amino acid sequence shown in SEQ ID NO: 2, 18, 22, 38, 42, 58, 62, 77, 81, 97, 101, 115, 119, 134, 138, 152, 156, 170, 174, 186, 190, 198, 200, 208, 210, 216, 218, 234, 238, 254, 258, 272, 276, 284, 286, 294, 296, 311, 315, 331, 335, 343, 345, 357, 361 and / or 376 or its variant; and / or (b) CDR-L1, CDR-L2 and CDR-L3 of an immunoglobulin light chain or its variable region containing the amino acid sequence shown in SEQ ID NO: 10, 20, 30, 40, 50, 60, 70, 79, 89, 99, 109, 117, 127, 136, 146, 154, 164, 172, 182, 188, 226, 236, 246, 256, 266, 274, 304, 313, 323, 333, 353, 359, 368 and / or 378 or its variant; or (c) an amino acid sequence shown in a member selected from SEQ ID NO: 1 to 378 or its variant. The present invention also provides a polynucleotide encoding one or more such polypeptides or a vector (e.g., a plasmid) containing such a polynucleotide.

[0022] The present invention also provides a host cell (e.g., a CHO cell) which comprises an antigen-binding protein (e.g., an antibody or its antigen-binding fragment) or an immunoglobulin chain (e.g., V H , V L , HC or LC) or a polypeptide or a polynucleotide or a vector as shown herein.

[0023] The present invention also provides a composition or a kit, which comprises one or more antigen-binding proteins (e.g., an antibody or an antigen-binding fragment thereof) as shown herein, optionally in combination with additional therapeutic agents (e.g., an anti-inflammatory agent, an anti-TNFα antibody or binding protein, infliximab, adalimumab, etanercept, golimumab, corticoid, prednisolone, methylprednisolone, antithymocyte globulin, alemtuzumab, dacluzimab, extracorporeal photophoresis, mycophenolate mofetil, sirolimus, pentostatin, mesenchymal stem cells, inolimomab, denileukin or basiliximab).

[0024] The present invention also provides a pharmaceutical formulation, which comprises an antigen-binding protein (e.g., an antibody or an antigen-binding fragment thereof) as shown herein and a pharmaceutically acceptable carrier, and optionally, additional therapeutic agents (e.g., an anti-inflammatory agent, an anti-TNFα antibody or binding protein, infliximab, adalimumab, etanercept, golimumab, corticoid, prednisolone, methylprednisolone, antithymocyte globulin, alemtuzumab, dacluzimab, extracorporeal photophoresis, mycophenolate mofetil, tacrolimus, cyclosporine, sirolimus, pentostatin, mesenchymal stem cells, inolimomab, denileukin or basiliximab).

[0025] The present invention also provides a container or an injection device (e.g., a vial, a syringe, a prefilled syringe or an autoinjector), which comprises an antigen-binding protein or a composition (e.g., a pharmaceutical formulation) as shown herein.

[0026] The present invention also provides methods for administering an antigen-binding protein or composition as shown herein to a subject (e.g., a human), which include introducing, e.g., injecting (e.g., subcutaneously, intravenously, or intramuscularly) the antigen-binding protein or composition into the subject. The present invention also provides methods for treating or preventing an IL2Rγ-mediated disease or disorder (e.g., graft-versus-host disease, organ transplant rejection, skin transplant rejection, heart transplant rejection, lung transplant rejection, kidney transplant rejection, liver transplant rejection, birdshot chorioretinopathy, multiple sclerosis, uveitis, autoimmune disease, Type I diabetes, multiple sclerosis, rheumatoid arthritis, systemic lupus erythematosus, and / or myasthenia gravis) in a subject in need thereof, which include administering, e.g., injecting an effective amount of an antigen-binding protein or composition as shown herein.

[0027] The present invention also provides a method for performing the following in a subject: blocking STAT phosphorylation in peripheral blood mononuclear cells (e.g., T cells) induced by cytokines (e.g., IL-2, IL-4, IL-7, IL-15, and / or IL-21); blocking STAT (e.g., STAT3) phosphorylation in mast cells induced by cytokines (e.g., IL-9); reducing the serum levels of interferon-γ, tumor necrosis factor-α, IL-6, IL-8, IL-10, and / or mKC / GRO (e.g., in a subject receiving a graft); blocking JAK-STAT-mediated (e.g., STAT3) intracellular signaling (e.g., in NK cells) induced by cytokines in the IL2Rγ family (e.g., IL-2, IL-4, IL-7, IL-9, IL-15, and / or IL-21); and / or reducing the serum levels of CD45+ immune cells, NK cells, T cells, and / or B cells (e.g., excluding neutrophils); the method comprising administering to the subject an effective amount of an anti-IL2Rγ antigen-binding protein or a composition or formulation thereof as shown herein. In one embodiment of the invention, the subject has an IL2Rγ-mediated disease or disorder, such as graft-versus-host disease, organ transplant rejection, b-islet cell transplant rejection, skin transplant rejection, heart transplant rejection, lung transplant rejection, kidney transplant rejection, liver transplant rejection, birdshot chorioretinopathy, multiple sclerosis, uveitis, autoimmune disease, type I diabetes, multiple sclerosis, rheumatoid arthritis, systemic lupus erythematosus, myasthenia gravis, aplastic anemia, atopic dermatitis, asthma, mast cell activation disorder, mast cell activation syndrome (MCAS), systemic mastocytosis (SM), and / or mast cell leukemia (MCL). BRIEF DESCRIPTION OF THE DRAWINGS

[0028] Panels (A) to (E) of FIG. 1. Blockade of human (panel (A)) IL-2-, (panel (B)) IL-4-, (panel (C)) IL7-, (panel (D)) IL-15-, and (panel (E)) IL-21-induced STAT phosphorylation in human CD4+ T cells by various concentrations of anti-IL-2Rγ antibodies H4H12857P, H4H12874P, H4H12886P, H4H12889P, and H4H12922P2, as well as antibodies REGN1945 and COMP1499.

[0029] Figure 2Blocking of human IL-9-induced STAT3 phosphorylation in in vitro-differentiated human mast cells by anti-IL-2Rγ antibodies H4H12874P, H4H12886P, H4H12889P, H4H12922P2, and antibodies COMP1499 and REGN1945.

[0030] Figure 3 Panels (A) to (F). Initial body weight percentage over time of mice with human PBMCs administered with anti-IL2Rγ antibodies (panel (E)) H4H12889P and (panel (F)) H4H12922P2 and antibody (panel (D)) COMP1499. Control experiments in mice (panel (B)) not administered with antibodies, (panel (C)) administered with isotype control antibody, or (panel (A)) without human PBMCs are also shown. The start of antibody injection on day 21 and the end of antibody injection on day 59 are indicated by dashed lines.

[0031] Figure 4 . Survival over time of mice injected with anti-IL2Rγ antibodies H4H12889P and H4H12922P2, antibody COMP1499, antibody REGN1945, and mice not injected with antibodies is shown. The group without huPBMCs is not shown. Differences in animal survival relative to the isotype control antibody group were analyzed by Mantel-Cox log-rank test. A P value < 0.05 was considered statistically significant. **, P value < 0.0021; ****, P value < 0.0001. The start of antibody injection on day 21 and the end of antibody injection on day 59 are indicated by dashed lines.

[0032] Figure 5 Panels (A) to (D). Absolute human cell numbers in blood on day 35 after injection of huPBMCs in mice not administered with antibodies (no IgG), administered with REGN1945, COMP1499, or anti-IL2Rγ antibodies H4H12889P or H4H12922P2 ((panel (A)) human CD45 cells; (panel (B)) human T cells; (panel (C)) human CD4 T cells; and (panel (D)) human CD8 T cells). The "no huPBMC" group is not shown; #, significantly different from the "no huPBMC" group; significantly different from the "huPBMC-no IgG" group; *, significantly different from the "huPBMC-REGN1945" group. Each symbol represents one mouse. For plotting purposes (log scale), zero values were arbitrarily changed to a value of 0.01.

[0033] Figure 6Panels (A) through (D). Blood counts of human (Panel (A)) CD45+ cells, (Panel (B)) T cells, (Panel (C)) CD4+ T cells, and (Panel (D)) CD8+ T cells over time in mice administered with anti-IL2Rγ antibodies H4H12889P or H4H12922P2, or COMP1499 or isotype control antibody. The start of antibody injection on day 21 and the end of antibody injection on day 59 are indicated by the dashed lines.

[0034] Figure 7 Panels (A) through (I). Serum levels of human and murine cytokines ((Panel (A)) human interferon-γ; (Panel (B)) human TNFα; (Panel (C)) human IL-6; (Panel (D)) human IL-8; (Panel (E)) human IL-10; (Panel (F)) murine TNFα; (Panel (G)) murine IL-6; (Panel (H)) murine KC / GRO; and (Panel (I)) murine IL-10) on day 42 after injection of huPBMC in mice administered with no antibody (no IgG), REGN1945, COMP1499, or anti-IL2Rγ antibodies H4H12889P or H4H12922P2, or in mice without human PBMC. #, significantly different from the "no huPBMC" group; significantly different from the "huPBMC-no IgG" group; *, significantly different from the "huPBMC-REGN1945" group. Each symbol represents one mouse.

[0035] Figure 8 Panels (A) through (D). Serum levels of (Panel (A)) human IFN-γ, (Panel (B)) human TNFα, (Panel (C)) murine TNFα, and (Panel (D)) murine IL-6 over time in mice administered with anti-IL2Rγ antibodies H4H12889P or H4H12922P2, or COMP1499 or isotype control antibody.

[0036] Panels (A) through (E) of Figure 9. Levels of total human antibody or CD45+ immune cells (Panel (A)), NK cells (Panel (B)), T cells (Panel (C)), B cells (Panel (D)), or neutrophils (Panel (E)) in the blood of mice treated with various doses of antibody REGN1945 or H4H12889P.

[0037] Figure 10 . Experimental design for in vivo skin graft rejection experiments.

[0038] Figure 11. Time to onset of skin graft rejection in mice not administered an antibody, administered REGN1945, or administered H4H12889P.

[0039] Figure 12 . Time to complete skin graft rejection in mice not administered an antibody, administered REGN1945, or administered H4H12889P.

[0040] Figure 13 . Total donor - specific IgG antibodies in non - transplanted mice or transplanted mice not administered an antibody, administered REGN1945, or administered H4H12889P. Detailed Description

[0041] The present invention provides antibodies and antigen - binding fragments thereof that specifically bind to human and cynomolgus monkey IL2Rγ and exhibit unique biological activities, particularly in blocking cytokine - induced STAT phosphorylation in T cells and blocking graft - versus - host disease in applicable mouse models.

[0042] In accordance with the present invention, conventional molecular biology, microbiology, and recombinant DNA techniques known in the art can be employed. Such techniques are well explained in the literature. See, for example, Sambrook, Fritsch & Maniatis, Molecular Cloning: A Laboratory Manual, 2nd ed. (1989) Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y. (hereinafter referred to as "Sambrook, et al., 1989"); DNA Cloning: A Practical Approach, Volumes I and II (D.N. Glover ed., 1985); Oligonucleotide Synthesis (M.J. Gait ed., 1984); Nucleic Acid Hybridization (B.D. Hames & S.J. Higgins eds. (1985)); Transcription And Translation (B.D. Hames & S.J. Higgins, eds. (1984)); Animal Cell Culture (R.I. Freshney, ed. (1986)); Immobilized Cells And Enzymes (IRL Press, (1986)); B. Perbal, A Practical Guide To Molecular Cloning (1984); F.M. Ausubel, et al. (eds.), Current Protocols in Molecular Biology, John Wiley & Sons, Inc. (1994).

[0043] IL-2Rγ

[0044] Interleukin-2 receptor subunit gamma is also known as CD132, common cytokine receptor gamma c chain, IL-2RG, IL-2Rg, IL2R gamma, IL-2Rγ, IMD4, P64:SCIDX, or SCIDX1. IL2Rγ is a subunit common to several interleukin receptors including IL-2R, IL-4R, IL-7R, IL-9R, IL-15R, and IL21R.

[0045] In one embodiment of the present invention, human IL2Rγ is encoded by the nucleotide sequence shown by Genbank accession number NM_000206. In one embodiment of the present invention, human IL2Rγ comprises the amino acid sequence shown by Genbank accession number NP_000197.

[0046] Antigen-binding protein

[0047] The present invention provides antigen-binding proteins that specifically bind to the IL2Rγ protein or an antigenic fragment thereof (e.g., the extracellular domain of IL2Rγ), such as antibodies (e.g., human antibodies, monoclonal antibodies, and recombinant antibodies) and antigen-binding fragments thereof. Antigen-binding proteins that bind to the same epitope on IL2Rγ as any of the antigen-binding proteins shown herein or that compete with any of the antigen-binding proteins shown herein for binding to IL2Rγ are also part of the present invention.

[0048] The present invention also provides any polypeptide comprising the amino acid sequence shown below or a variant thereof:

[0049] SEQ ID NO: 2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42, 44, 46, 48, 50, 52, 54, 56, 58, 60, 62, 64, 66, 68, 70, 72, 75, 77, 79, 81, 83, 85, 87, 89, 91, 93, 95, 97, 99, 101, 103, 105, 107, 109, 111, 113, 115, 117, 119, 121, 123, 125, 127, 129, 132, 134, 136, 138, 140, 142, 144, 146, 148, 150, 152, 154, 156, 158, 160, 162, 164, 166, 168, 170, 172, 174, 176, 178, 180, 182, 184, 186, 188, 190, 192, 194, 196, 198, 200, 202, 204, 206, 208, 210, 212, 214, 216, 218, 220, 222, 224, 226, 228, 230, 232, 234, 236, 238, 240, 242, 244, 246, 248, 250, 252, 254, 256, 258, 260, 262, 264, 266, 268, 270, 272, 274, 276, 278, 280, 282, 284, 286, 288, 290, 292, 294, 296, 298, 300, 302, 304, 306, 309, 311, 313, 315, 317, 319, 321, 323, 325, 327, 329, 331, 333, 335, 337, 339, 341, 343, 345, 347, 349, 351, 353, 355, 357, 359, 361, 363, 366, 368, 370, 372, 374, 376 and / or 378

[0050] Optionally, the polypeptide is fused to one or more additional polypeptides, such as human Fc (such as human IgG such as IgG1 or IgG4 (such as, comprising the S108P mutation)).

[0051] As used herein, the term "antibody" refers to an immunoglobulin molecule (i.e., "complete antibody molecule") (such as IgG) that comprises four polypeptide chains: two heavy chains (HC) and two light chains (LC) that are interconnected by disulfide bonds - such as

[0052] H4H12857P; H4H12858P; H4H12859P; H4H12863P; H4H12874P; H4H12871P; H4H12884P; H4H12886P; H4H12889P; H4H12890P; H4H12899P; H4H12900P; H4H12908P; H4H12913P2; H4H12922P2; H4H12924P2; H4H12926P2; H4H12927P2; H4H12934P2; H4H13538P; H4H13541P; H4H13544P2; or H4H13545P2.

[0053] In one embodiment of the invention, each antibody heavy chain (HC) comprises a heavy chain variable region ("HCVR" or "V H ") (e.g., SEQ ID NO: 2, 22, 42, 62, 81, 101, 119, 138, 156, 174, 190, 200, 210, 218, 238, 258, 276, 286, 296, 315, 335, 345 or 361 or a variant thereof) and a heavy chain constant region (comprising domains C H 1, C H 2 and C H 3); and each antibody light chain (LC) comprises a light chain variable region ("LCVR or "V L ") (e.g., SEQ ID NO: 10, 30, 50, 70, 89, 109, 127, 146, 164, 182, 226, 246, 266, 304, 323, 353 or 368 or a variant thereof) and a light chain constant region (C L ). The V H and V L regions can be further subdivided into hypervariable regions called complementarity determining regions (CDRs), which are interspersed with more conserved regions called framework regions (FRs). The V H and V L each contain three CDRs and four FRs arranged in the following order from the amino terminus to the carboxyl terminus: FR1, CDR1, FR2, CDR2, FR3, CDR3, FR4. In certain embodiments of the invention, the FRs of the antibody (or its antigen-binding fragment) are identical to the germline sequences or are naturally or artificially modified.

[0054] Generally, the variable domains of both immunoglobulin heavy and light chains contain three hypervariable regions, also called complementarity determining regions (CDRs), located within relatively conserved framework regions (FRs). Generally, from the N-terminus to the C-terminus, the variable domains of both the light and heavy chains contain FR1, CDR1, FR2, CDR2, FR3, CDR3, and FR4. In one embodiment of the invention, amino acids are assigned to each domain according to the definitions in: Sequences of Proteins of Immunological Interest, Kabat, et al.; National Institutes of Health, Bethesda, Md.; 5th Edition; NIH Publ. No. 91-3242 (1991); Kabat (1978) Adv. Prot. Chem. 32:1-75; Kabat, et al., (1977) J. Biol. Chem. 252:6609-6616; Chothia, et al., (1987) J Mol. Biol. 196:901-917 or Chothia, et al., (1989) Nature 342:878-883. Accordingly, the present invention includes antibodies and antigen-binding fragments containing the CDRs of V H and the CDRs of V L , wherein the V H and V L comprise amino acid sequences (or variants thereof) as shown herein, wherein the CDRs are defined according to Kabat and / or Chothia.

[0055] As used herein, terms such as "antigen-binding portion" or "antigen-binding fragment" of an antibody or antigen-binding protein include any naturally occurring, enzymatically obtainable, synthetic, or genetically engineered polypeptide or glycoprotein that specifically binds to an antigen to form a complex. Non-limiting examples of antigen-binding fragments include: (i) Fab fragments; (ii) F(ab’)2 fragments; (iii) Fd fragments (the heavy chain portion of a Fab fragment cleaved with papain); (iv) Fv fragments (V H or V L); and (v) single-chain Fv (scFv) molecules; consisting of amino acid residues that mimic the hypervariable regions of antibodies (e.g., isolated complementarity-determining regions (CDRs), such as CDR3 peptides) or restricted FR3-CDR3-FR4 peptides. Other engineered molecules, such as domain-specific antibodies, single-domain antibodies, domain-deleted antibodies, chimeric antibodies, CDR-grafted antibodies, diabodies, triabodies, tetra-bodies, micro-antibodies, and small modular immunopharmaceuticals (SMIPs), are also included within the expression "antigen-binding fragment" as used herein. In one embodiment of the invention, the antigen-binding fragment comprises three or more of the following CDRs (e.g., CDR-H1, CDR-H2, and CDR-H3; or CDR-L1, CDR-L2, and CDR-L3):

[0056] H4H12857P; H4H12858P; H4H12859P; H4H12863P; H4H12874P; H4H12871P; H4H12884P; H4H12886P; H4H12889P; H4H12890P; H4H12899P; H4H12900P; H4H12908P; H4H12913P2; H4H12922P2; H4H12924P2; H4H12926P2; H4H12927P2; H4H12934P2; H4H13538P; H4H13541P; H4H13544P2; or H4H13545P2.

[0057] In one embodiment of the invention, the antigen-binding protein of the invention (e.g., an antibody or an antigen-binding fragment thereof) comprises: an immunoglobulin heavy chain containing V H (e.g., HC), the V H comprising a combination of heavy chain CDRs (CDR-H1, CDR-H2, and CDR-H3) shown in Table A below; and / or an immunoglobulin light chain containing VL (e.g., LC), the V L comprising a combination of light chain CDRs (CDR-L1, CDR-L2, and CDR-L3) shown in Table B below.

[0058] Table A. Heavy Chain CDRs in the Immunoglobulins of the Invention

[0059] CDR-H combination CDR-H1 CDR-H2 CDR-H3 1 4 6 8 2 24 26 28 3 44 46 48 4 64 66 68 5 83 85 87 6 103 105 107 7 121 123 125 8 140 142 144 9 158 160 162 10 176 178 180 11 192 194 196 12 202 204 206 13 176 212 214 14 220 222 224 15 240 242 244 16 260 262 264 17 278 280 282 18 288 290 292 19 298 300 302 20 317 319 321 21 337 339 341 22 347 349 351 23 363 66 366

[0060] * The numbers correspond to the amino acid sequences shown in the SEQ ID NO.

[0061] Table B. Light Chain CDRs in the Immunoglobulins of the Invention

[0062]

[0063]

[0064] *The numbering corresponds to the amino acid sequence shown in that SEQ ID NO.

[0065] In one embodiment of the invention, the antigen-binding protein of the invention (e.g., an antibody or an antigen-binding fragment thereof) correspondingly contains immunoglobulin heavy and light chains containing V H (e.g., HC) and V L (e.g., LC), which contain the combinations of heavy and light chain CDRs (CDR-H1, CDR-H2, and CDR-H3; and CDR-L1, CDR-L2, and CDR-L3) shown in Table C below.

[0066] Table C. Heavy and Light Chain CDRs in the Immunoglobulins of the Invention

[0067]

[0068]

[0069] *The numbering corresponds to the amino acid sequence shown in that SEQ ID NO.

[0070] The invention includes antigen-binding proteins (e.g., antibodies or antigen-binding fragments thereof) comprising polypeptide pairs containing the following V H and V L amino acid sequences:

[0071] SEQ ID NO: 2 and SEQ ID NO: 10;

[0072] SEQ ID NO: 22 and SEQ ID NO: 30;

[0073] SEQ ID NO: 42 and SEQ ID NO: 50;

[0074] SEQ ID NO: 62 and SEQ ID NO: 70;

[0075] SEQ ID NO: 81 and SEQ ID NO: 89;

[0076] SEQ ID NO: 101 and SEQ ID NO: 109;

[0077] SEQ ID NO: 119 and SEQ ID NO: 127;

[0078] SEQ ID NO: 138 and SEQ ID NO: 146;

[0079] SEQ ID NO: 156 and SEQ ID NO: 164;

[0080] SEQ ID NO: 174 and SEQ ID NO: 182;

[0081] SEQ ID NO: 190 and SEQ ID NO: 182;

[0082] SEQ lD NO: 200 and SEQ lD NO: 182;

[0083] SEQ ID NO: 210 and SEQ ID NO: 182;

[0084] SEQ ID NO: 218 and SEQ ID NO: 226;

[0085] SEQ ID NO: 238 and SEQ lD NO: 246;

[0086] SEQ ID NO: 258 and SEQ ID NO: 266;

[0087] SEQ ID NO: 276 and SEQ ID NO: 182;

[0088] SEQ ID NO: 286 and SEQ lD NO: 182;

[0089] SEQ ID NO: 296 and SEQ ID NO: 304;

[0090] SEQ ID NO: 315 and SEQ ID NO: 323;

[0091] SEQ ID NO: 335 and SEQ lD NO: 182;

[0092] SEQ ID NO: 345 and SEQ ID NO: 353; or

[0093] SEQ ID NO: 361 and SEQ lD NO: 368..

[0094] The present invention includes an antigen-binding protein (e.g., an antibody or an antigen-binding fragment thereof) comprising an amino acid sequence pair encoding HC and LC as follows:

[0095] SEQ ID NO: 18 and SEQ ID NO: 20;

[0096] SEQ ID NO: 38 and SEQ ID NO: 40;

[0097] SEQ ID NO: 58 and SEQ ID NO: 60;

[0098] SEQ ID NO: 77 and SEQ ID NO: 79;

[0099] SEQ ID NO: 97 and SEQ ID NO: 99;

[0100] SEQ ID NO: 115 and SEQ ID NO: 117;

[0101] SEQ ID NO: 134 and SEQ ID NO: 136;

[0102] SEQ ID NO: 152 and SEQ ID NO: 154;

[0103] SEQ ID NO: 170 and SEQ ID NO: 172;

[0104] SEQ ID NO: 186 and SEQ ID NO: 188;

[0105] SEQ ID NO: 198 and SEQ ID NO: 188;

[0106] SEQ ID NO: 208 and SEQ ID NO: 188;

[0107] SEQ ID NO: 216 and SEQ ID NO: 188;

[0108] SEQ ID NO: 234 and SEQ ID NO: 236;

[0109] SEQ ID NO: 254 and SEQ ID NO: 256;

[0110] SEQ ID NO: 272 and SEQ ID NO: 274;

[0111] SEQ ID NO: 284 and SEQ ID NO: 188;

[0112] SEQ ID NO: 294 and SEQ ID NO: 188;

[0113] SEQ ID NO: 311 and SEQ ID NO: 313;

[0114] SEQ ID NO: 331 and SEQ ID NO: 333;

[0115] SEQ ID NO: 343 and SEQ ID NO: 188;

[0116] SEQ ID NO: 357 and SEQ ID NO: 359; or

[0117] SEQ ID NO: 376 and SEQ ID NO: 378..

[0118] Some embodiments of the invention also include antigen-binding proteins containing immunoglobulin V H and V L domains, or HC and LC, such as anti-IL2Rγ antibodies and antigen-binding fragments thereof, wherein the immunoglobulin V H and V L domains, or HC and LC contain variant amino acid sequences having 70% or more (e.g., 80%, 85%, 90%, 95%, 97% or 99%) overall amino acid sequence identity or similarity to the amino acid sequences of the corresponding V H and V L domains, HC or LC specifically shown herein, provided that the CDR-L1, CDR-L2, CDR-L3, CDR-H1, CDR-H2 and CDR-H3 of such immunoglobulins are not variants and contain the amino acid sequences shown herein. Thus, in some such embodiments, the CDRs in the variant antigen-binding proteins are not themselves variants.

[0119] The invention includes monoclonal anti-IL2Rγ antigen-binding proteins, such as antibodies and antigen-binding fragments thereof, and monoclonal compositions comprising a plurality of isolated monoclonal antigen-binding proteins. As used herein, the term "monoclonal antibody" or "mAb" refers to a member of a population of antibodies that is substantially homogeneous, i.e., the population comprises antibody molecules that are identical in amino acid sequence except for possible naturally occurring mutations that may be present in minor amounts. The "plurality" of such monoclonal antibodies and fragments in the composition refers to a concentration of the same (i.e., as discussed above, identical in amino acid sequence except for possible naturally occurring mutations that may be present in minor amounts) antibodies and fragments that is higher than that normally present in nature (e.g., in the blood of a host organism (e.g., a mouse or a human)).

[0120] In one embodiment of the present invention, the anti-IL2Rγ antigen-binding protein (e.g., an antibody or antigen-binding fragment) comprises a heavy chain constant domain, such as an IgA (e.g., IgA1 or IgA2), IgD, IgE, IgG (e.g., IgG1, IgG2, IgG3, and IgG4 (e.g., comprising S228P and / or S108P mutations)), or IgM type heavy chain constant domain. In one embodiment of the present invention, the antigen-binding protein (e.g., an antibody or antigen-binding fragment) comprises a light chain constant domain, such as a K or λ type light chain constant domain. The present invention includes the following antigen-binding proteins that comprise variable domains (e.g., H4H12857P; H4H12858F; H4H12859P; H4H12863P; H4H12874P; H4H12871P; H4H12884P; H4H12886P; H4H12889P; H4H12890P; H4H12899P; H4H12900P; H4H12908P; H4H12913P2; H4H12922P2; H4H12924P2; H4H12926P2; H4H12927P2; H4H12934P2; H4H13538P; H4H13541P; H4H13544P2; or H4H13545P2) linked to a heavy chain and / or light chain constant domain (e.g., as shown above).

[0121] As used herein, the term "human" antigen-binding protein, such as an antibody or antigen-binding fragment, includes antibodies and fragments having variable and constant regions derived from human germline immunoglobulin sequences, whether in human cells or transplanted into non-human cells (e.g., murine cells). See, e.g., US8502018, US6596541, or US5789215. In one embodiment of the invention, the human antibodies and antigen-binding fragments of the invention may contain amino acid residues not encoded by human germline immunoglobulin sequences (e.g., having mutations introduced by in vitro random or site-specific mutagenesis or by in vivo somatic mutation), such as in the CDRs, particularly in CDR3. However, the term "human antibody" as used herein is not intended to include mAbs in which CDR sequences from the germline of another mammalian species (e.g., murine) have been grafted onto human FR sequences. The term includes antibodies recombinantly produced in non-human mammals or cells of non-human mammals. The term is not intended to include antibodies isolated from or produced in a human subject. The invention includes human antigen-binding proteins (e.g., antibodies or antigen-binding fragments thereof, such as H4H12857P; H4H12858P; H4H12859P; H4H12863P; H4H12874P; H4H12871P; H4H12884P; H4H12886P; H4H12889P; H4H12890P; H4H12899P; H4H12900P; H4H12908P; H4H12913P2; H4H12922P2; H4H12924P2; H4H12926P2; H4H12927P2; H4H12934P2; H4H13538P; H4H13541P; H4H13544P2; or H4H13545P2).

[0122] The present invention includes anti-IL2Rγ chimeric antigen-binding proteins, such as antibodies and antigen-binding fragments thereof, and methods of using the same. As used herein, a "chimeric antibody" is an antibody having variable domains from a first antibody and constant domains from a second antibody, wherein the first and second antibodies are from different species. (See, e.g., US 4816567; and Morrison et al., (1984) Proc. Natl. Acad. Sci. USA 81: 6851-6855). The present invention includes chimeric antibodies containing the variable domains shown herein (e.g., the variable domains from H4H12857P; H4H12858P; H4H12859P; H4H12863P; H4H12874P; H4H12871P; H4H12884P; H4H12886P; H4H12889P; H4H12890P; H4H12899P; H4H12900P; H4H12908P; H4H12913P2; H4H12922P2; H4H12924P2; H4H12926P2; H4H12927P2; H4H12934P2; H4H13538P; H4H13541P; H4H13544P2; or H4H13545P2).

[0123] The term "recombinant" antigen-binding protein, such as an antibody or antigen-binding fragment thereof, refers to such a molecule that is produced, expressed, isolated, or obtained by techniques or methods known in the art as recombinant DNA technology, which includes, for example, DNA splicing and transgenic expression. The term includes antibodies expressed in non-human mammals (including transgenic non-human mammals, such as transgenic mice) or host cells (such as Chinese hamster ovary (CHO) cells) or cell expression systems, or antibodies isolated from recombinant combinatorial human antibody libraries. The present invention includes recombinant antigen-binding proteins as shown herein (e.g., H4H12857P; H4H12858P; H4H12859P; H4H12863P; H4H12874P; H4H12871P; H4H12884P; H4H12886P; H4H12889P; H4H12890P; H4H12899P; H4H12900P; H4H12908P; H4H12913P2; H4H12922P2; H4H12924P2; H4H12926P2; H4H12927P2; H4H12934P2; H4H13538P; H4H13541P; H4H13544P2; or H4H13545P2).

[0124] In one embodiment of the invention, an antigen-binding fragment of an antibody comprises at least one variable domain. The variable domain can be of any size or amino acid composition and generally comprises at least one (e.g., 3) CDRs adjacent to or in-frame with one or more framework sequences. In an antigen-binding fragment having a V L domain associated with a V H domain, the V H and V L domains can be positioned relative to each other in any suitable arrangement. For example, the variable region can be dimeric and contain V H -V H , V H -V L or V L -V L dimers. Alternatively, the antigen-binding fragment of an antibody can comprise non-covalently associated monomeric V H and / or V L domains.

[0125] In certain embodiments, the antigen-binding fragment of an antibody can comprise at least one variable domain covalently linked to at least one constant domain. Non-limiting exemplary configurations of variable and constant domains that can be present in the antigen-binding fragments of the antibodies of the invention include: (i) V H -C H 1; (ii) V H -C H 2; (iii) V H -C H 3; (iv) V H -C H 1-C H 2; (v) V H -C H 1-C H 2-C H 3; (vi) V H -C H 2-C H 3; (vii) V H -C L ; (viii) V L -C H 1; (ix) V L -C H 2; (x) V L -CH3; (xi) V L -C H 1-C H 2; (xii) V L -C H 1-C H 2-C H 3; (xiii) VL -C H 2-C H 3; and (xiv) V L -C L 。In any configuration of variable and constant domains including any of the exemplary configurations listed above, the variable and constant domains can be directly connected to each other or can be connected by a full or partial hinge region or linker region. The hinge region can consist of at least 2 (e.g., 5, 10, 15, 20, 40, 60 or more) amino acids, which results in a flexible or semi-flexible connection between adjacent variable and / or constant domains in a single polypeptide molecule. Additionally, antigen-binding fragments of the antibodies of the present invention can comprise any of the variable and constant domain configurations listed above non-covalently associated with each other and / or a homodimer or heterodimer (or other multimer) of one or more monomeric V H or V L domains (e.g., via one or more disulfide bonds). The present invention includes antigen-binding fragments of the antigen-binding proteins shown herein, e.g.,

[0126] H4H12857P; H4H12858P; H4H12859P; H4H12863P; H4H12874P; H4H12871P; H4H12884P; H4H12886P; H4H12889P; H4H12890P; H4H12899P; H4H12900P; H4H12908P; H4H12913P2; H4H12922P2; H4H12924P2; H4H12926P2; H4H12927P2; H4H12934P2; H4H13538P; H4H13541P; H4H13544P2; or H4H13545P2..

[0127] Antigen-binding proteins (e.g., antibodies and antigen-binding fragments) can be monospecific or multispecific (e.g., bispecific). Multispecific antigen-binding proteins are further discussed herein. The present invention includes monospecific as well as multispecific (e.g., bispecific) antigen-binding fragments that comprise one or more variable domains from the antigen-binding proteins specifically shown herein (e.g., H4H12857P; H4H12858P; H4H12859P; H4H12863P; H4H12874P; H4H12871P; H4H12884P; H4H12886P; H4H12889P; H4H12890P; H4H12899P; H4H12900P; H4H12908P; H4H12913P2; H4H12922P2; H4H12924P2; H4H12926P2; H4H12927P2; H4H12934P2; H4H13538P; H4H13541P; H4H13544P2; or H4H13545P2).

[0128] The term "specifically binds" refers to those antigen-binding proteins (e.g., antibodies or antigen-binding fragments thereof) that bind to an antigen (e.g., the IL2Rγ protein) with a binding affinity represented by K D of at least about 10 -7 M (e.g., 10 -8 M, 10 -9 M, 10 -10 M, 10 -11 M or 10 -12 M), as measured by real-time, label-free biolayer interferometry, e.g., at 25 °C or 37 °C, e.g., HTX biosensor, or by surface plasmon resonance, e.g., BIACORE TM , or by solution affinity ELISA. The present invention includes antigen-binding proteins that specifically bind to the IL2Rγ protein. In one embodiment of the present invention, the anti-IL2Rγ antigen-binding protein comprises a K D value for binding to human and / or murine and / or cynomolgus monkey and / or rat IL2Rγ or a domain thereof, as shown in any of Tables 3-1 to 3-12. "Anti-IL2Rγ" refers to an antigen-binding protein (or other molecule) that specifically binds to IL2Rγ, e.g., an antibody or an antigen-binding fragment thereof.

[0129] "Isolated" antigen-binding proteins (e.g., antibodies or antigen-binding fragments thereof), polypeptides, polynucleotides, and vectors are at least partially free of other biomolecules from the cells or cell culture in which they are produced. Such biomolecules include nucleic acids, proteins, other antibodies or antigen-binding fragments, lipids, carbohydrates, or other substances such as cell debris and growth media. Isolated antigen-binding proteins may also be at least partially free of expression system components, such as biomolecules from the host cell or its growth media. Generally, the term "isolated" is not intended to mean: the complete absence of such biomolecules (e.g., a small or insignificant amount of impurities may remain); the absence of water, buffer, or salt; or the components of a pharmaceutical formulation containing the antigen-binding protein (e.g., antibody or antigen-binding fragment).

[0130] The present invention includes antigen-binding proteins, such as antibodies or antigen-binding fragments, that bind the same epitope as the antigen-binding proteins of the present invention (e.g., H4H12857P; H4H12858P; H4H12859P; H4H12863P; H4H12874P; H4H12871P; H4H12884P; H4H12886P; H4H12889P; H4H12890P; H4H12899P; H4H12900P; H4H12908P; H4H12913P2; H4H12922P2; H4H12924P2; H4H12926P2; H4H12927P2; H4H12934P2; H4H13538P; H4H13541P; H4H13544P2; or H4H13545P2).

[0131] An antigen is, for example, a molecule to which an antibody binds, such as a peptide (e.g., IL2Rγ or a fragment thereof (antigenic fragment)). The specific region on an antigen that an antibody recognizes and binds is called an epitope. The antigen-binding proteins of the present invention (e.g., antibodies) that specifically bind to such antigens are part of the present invention.

[0132] The term "epitope" refers to the specific antigen-binding site of an antigen-binding protein, such as an antigenic determinant (e.g., on IL2Rγ) that interacts with the variable region of an antibody molecule, called a paratope. A single antigen can have more than one epitope. Thus, different antibodies can bind to different regions on an antigen and can have different biological effects. The term "epitope" can also refer to the site on an antigen to which B cells and / or T cells respond, and / or to the region of an antigen that is bound by an antibody. Epitopes can be defined as structural or functional. Functional epitopes are generally a subset of structural epitopes and have those residues that directly contribute to the affinity of the interaction. Epitopes can be linear or conformational, i.e., composed of non-linear amino acids. In certain embodiments, an epitope can comprise determinants that are chemical reactive surface groups of a molecule such as an amino acid, sugar side chain, phosphoryl or sulfonyl group, and in certain embodiments can have specific three-dimensional structural features and / or specific charge characteristics. The epitopes bound by the antigen-binding proteins of the present invention can be contained in a fragment of IL2Rγ (e.g., human IL2Rγ), such as its extracellular domain, domain 1 or domain 2. The antigen-binding proteins (e.g., antibodies) of the present invention that bind to such epitopes are part of the present invention.

[0133] Methods for determining the epitopes of antigen-binding proteins, such as antibodies or fragments or polypeptides, include alanine-scanning mutagenesis analysis, peptide blotting analysis (Reineke (2004) Methods Mol. Biol. 248:443-63), peptide cleavage analysis, crystallographic studies, and NMR analysis. In addition, methods such as epitope excision, epitope extraction, and chemical modification of an antigen (Tomer i2000) Prot. Sci. 9:487-496) can also be employed. Another method that can be used to identify the amino acids in a polypeptide that interact with an antigen-binding protein (e.g., an antibody or fragment or polypeptide) is by mass spectrometry detection of hydrogen / deuterium exchange. See, e.g., Ehring (1999) Analytical Biochemistry 267:252-259; Engen and Smith (2001) Anal. Chem. 73:256A-265A.

[0134] The present invention includes antigen-binding proteins that compete with the antigen-binding proteins of the present invention, such as for example, for binding to IL2Rγ, such as the variant IL2Rγ epitopes discussed herein.

[0135] H4H12857P; H4H12858P; H4H12859P; H4H12863P; H4H12874P; H4H12871P; H4H12884P; H4H12886P; H4H12889P; H4H12890P; H4H12899P; H4H12900P; H4H12908P; H4H12913P2; H4H12922P2; H4H12924P2; H4H12926P2; H4H12927P2; H4H12934P2; H4H13538P; H4H13541P; H4H13544P2; or H4H13545P2. As used herein, the term "compete" refers to an antigen-binding protein (e.g., an antibody or an antigen-binding fragment thereof) that binds to an antigen (e.g., IL2Rγ) and inhibits or blocks the binding of another antigen-binding protein (e.g., an antibody or an antigen-binding fragment thereof) to that antigen. Unless otherwise specified, the term also includes competition between two antigen-binding proteins, e.g., antibodies, in both directions, i.e., the first antibody binds to the antigen and blocks the binding of the second antibody, and vice versa. Thus, in one embodiment of the invention, competition occurs in one such direction. In certain embodiments, the first antigen-binding protein (e.g., an antibody) and the second antigen-binding protein (e.g., an antibody) can bind to the same epitope. Alternatively, the first and second antigen-binding proteins (e.g., antibodies) can bind to different but e.g., overlapping or non-overlapping epitopes, where the binding of one inhibits or blocks, e.g., by steric hindrance, the binding of the second antibody. Competition between antigen-binding proteins (e.g., antibodies) can be measured by methods known in the art, e.g., by real-time, label-free biolayer interferometry. In addition, the binding competition between anti-IL2Rγ antigen-binding proteins (e.g., monoclonal antibodies (mAbs)) can be determined using real-time, label-free biolayer interferometry performed on an Octet RED384 biosensor (Pall ForteBio Corp.).

[0136] Generally, an antibody or antigen-binding fragment of the invention that is modified in some way retains the ability to specifically bind to IL2Rγ, e.g., retains at least 10% of its IL2Rγ-binding activity (when compared to the parental antibody), when the activity is expressed on a molar basis. Preferably, an antibody or antigen-binding fragment of the invention retains at least 20%, 50%, 70%, 80%, 90%, 95% or 100% or more of the IL2Rγ-binding affinity of the parental antibody. It is also contemplated that an antibody or antigen-binding fragment of the invention can contain conservative or non-conservative amino acid substitutions that substantially do not alter its biological activity (referred to as "conservative variants" or "functionally conservative variants" of the antibody).

[0137] Polypeptides, such as immunoglobulin chains (e.g., H4H12857P, H4H12858P, H4H12859P, H4H12863P, H4H12874P, H4H12871P, H4H12884P, H4H12886P, H4H12889P, H4H12890P, H4H12899P, H4H12900P, H4H12908P, H4H12913P2, H4H12922P2, H4H12924P2, H4H12926P2, H4H12927P2, H4H12934P2, H4H13538P, H4H13541P, H4H13544P2 or H4H13545P2 V H 、V L, a "variant" of HC or LC or its CDR, which comprises the amino acid sequence specifically shown above, refers to a polypeptide that comprises at least about 70% to 99.9% (e.g., at least 70%, 72%, 74%, 75%, 76%, 79%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 99.5% or 99) of the reference amino acid sequence shown herein (e.g., any one of SEQ ID NO: 2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42, 44, 46, 48, 50, 52, 54, 56, 58, 60, 62, 64, 66, 68, 70, 72, 75, 77, 79, 81, 83, 85, 87, 89, 91, 93, 95, 97, 99, 101, 103, 105, 107, 109, 111, 113, 115, 117, 119, 121, 123, 125, 127, 129, 132, 134, 136, 138, 140, 142, 144, 146, 148, 150, 152, 154, 156, 158, 160, 162, 164, 166, 168, 170, 172, 174, 176, 178, 180, 182, 184, 186, 188, 190, 192, 194, 196, 198, 200, 202, 204, 206, 208, 210, 212, 214, 216, 218, 220, 222, 224, 226, 228, 230, 232, 234, 236, 238, 240, 242, 244, 246, 248, 250, 252, 254, 256, 258, 260, 262, 264, 266, 268, 270, 272, 274, 276, 278, 280, 282, 284, 286, 288, 290, 292, 294, 296, 298, 300, 302, 304, 306, 309, 311, 313, 315, 317, 319, 321, 323, 325, 327, 329, 331, 333, 335, 337, 339, 341, 343, 345, 347, 349, 351, 353, 355, 357, 359, 361, 363, 366, 368, 370, 372, 374, 376 or 378).amino acid sequences that are the same as or similar to the reference polypeptide (e.g., 9%); when the comparison is performed by the BLAST algorithm, the parameters of the algorithm are selected to give the maximum match between the corresponding sequences over the entire length of the corresponding reference sequence (e.g., expectation threshold: 10; word length: 3; maximum match in the query range: 0; BLOSUM 62 matrix; gap penalty: existence 11, extension 1; conditional combination scoring matrix adjustment).

[0138] In addition, variants of the polypeptide may include the following polypeptides, such as immunoglobulin chains (e.g., H4H12857P, H4H12858P, H4H12859P, H4H12863P, H4H12874P, H4H12871P, H4H12884P, H4H12886P, H4H12889P, H4H12890P, H4H12899P, H4H12900P, H4H12908P, H4H12913P2, H4H12922P2, H4H12924P2, H4H12926P2, H4H12927P2, H4H12934P2, H4H13538P, H4H13541P, H4H13544P2, or H4H13545P2 V H 、V L 、HC or LC or their CDRs), which may contain the amino acid sequence of the reference polypeptide specifically shown herein, but with one or more (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10) mutations, such as one or more missense mutations (e.g., conservative substitutions), nonsense mutations, deletions, or insertions. For example, the present invention includes an anti-IL2Rγ antigen-binding protein, which comprises: an immunoglobulin light chain (or V L ) variant containing the amino acid sequence shown in SEQ ID NO: 10 but having one or more such mutations, and / or an immunoglobulin heavy chain (or V H ) variant containing the amino acid sequence shown in SEQ ID NO: 2 but having one or more such mutations. In one embodiment of the present invention, the anti-IL2Rγ antigen-binding protein comprises: an immunoglobulin light chain variant containing CDR-L1, CDR-L2, and CDR-L3, wherein one or more (e.g., 1 or 2 or 3) of such CDRs have one or more such mutations (e.g., conservative substitutions); and / or an immunoglobulin heavy chain variant containing CDR-H1, CDR-H2, and CDR-H3, wherein one or more (e.g., 1 or 2 or 3) of such CDRs have one or more such mutations (e.g., conservative substitutions).

[0139] The following references relate to the BLAST algorithms commonly used in sequence analysis: BLAST ALGORITHMS: Altschul et al. (2005) FEBS J. 272(20): 5101-5109; Altschul, S.F., et al., (1990) J. Mol. Biol. 215: 403-410; Gish, W., et al., (1993) Nature Genet. 3: 266-272; Madden, T.L., et al., (1996) Meth. Enzymol. 266: 131-141; Altschul, S.F., et al., (1997) Nucleic Acids Res. 25: 3389-3402; Zhang, J., et al., (1997) Genome Res. 7: 649-656; Wootton, J.C., et al., (1993) Comput. Chem. 17: 149-163; Hancock, J.M. et al., (1994) Comput. Appl. Biosci. 10: 67-70; ALIGNMENT SCORING SYSTEMS: Dayhoff, M.O., et al., "A model of evolutionary change in proteins." Atlas of Protein Sequence and Structure, (1978) vol. 5, suppl. 3. M.O. Dayhoff (ed.), pp. 345-352, Natl. Biomed. Res. Found., Washington, D.C.; Schwartz, R.M., et al., "Matrices for detecting distant relationships." Atlas of Protein Sequence and Structure, (1978) vol. 5, suppl. 3. "M.O. Dayhoff (ed.), pp. 353-358, Natl. Biomed. Res. Found., Washington, D.C.; Altschul, S.F., (1991) J. Mol. Biol. 219: 555-565; States, D.J., et al., (1991) Methods 3: 66-70; Henikoff, S., et al., (1992) Proc. Natl. Acad. Sci. USA 89: 10915-10919; Altschul, S.F., et al., (1993) J. Mol. Evol. 36: 290-300; ALIGNMENT STATISTICS: Karlin, S., et al., (1990) Proc. Natl. Acad. Sci. USA 87: 2264-2268; Karlin, S., et al., (1993) Proc. Natl. Acad. Sci. USA 90: 5873-5877; Dembo, A., et al., (1994) Ann. Prob. 22: 2022-2039; and Altschul, S.F. "Evaluating the statistical significance of multiple distinct local alignments." Theoretical and Computational Methods in Genome Research (S. Suhai ed.), (1997) pp. 1-14, Plenum, N.Y.

[0140] A "conservative modification variant" or "conservative substitution," e.g., of an immunoglobulin chain as shown herein, refers to a variant in which one or more amino acids in a polypeptide are replaced with other amino acids having similar characteristics (e.g., charge, side chain size, hydrophobicity / hydrophilicity, backbone conformation, and rigidity, etc.). Such changes can often be made without significantly disrupting the biological activity of the antibody or fragment. Those skilled in the art recognize that, generally, a single amino acid substitution in a non-essential region of a polypeptide will substantially not change biological activity (see, e.g., Watson et al. (1987) Molecular Biology of the Gene, The Benjamin / Cummings Pub. Co., p. 224 (4th ed.)). In addition, substitutions of amino acids that are structurally or functionally similar are not likely to significantly disrupt biological activity. The present invention includes anti-IL2Rγ antigen-binding proteins containing such conservatively modified variant immunoglobulin chains.

[0141] Examples of groups of amino acids with side chains having similar chemical properties include: 1) aliphatic side chains: glycine, alanine, valine, leucine, and isoleucine; 2) aliphatic hydroxyl side chains: serine and threonine; 3) amide-containing side chains: asparagine and glutamine; 4) aromatic side chains: phenylalanine, tyrosine, and tryptophan; 5) basic side chains: lysine, arginine, and histidine; 6) acidic side chains: aspartic acid and glutamic acid, and 7) sulfur-containing side chains: cysteine ​​and methionine. Alternatively, a conservative substitution is any change disclosed in Gonnet et al. (1992) Science 256: 1443-45 that has a positive value in the PAM250 log-likelihood matrix.

[0142] The anti-IL2Rγ antigen binding proteins disclosed herein, e.g., anti-IL2Rγ antigen binding proteins comprising variant immunoglobulin chains, may exhibit one or more of the following properties:

[0143] At 25°C, about 2.75×10 -9 M is about 3.36×10 -7 M of K D binds to human IL2Rγ (e.g., a fusion thereof, such as a myc-myc-His6 fusion);

[0144] At 37°C, the -9 M is about 3.53×10 -7 M of K D binds to human IL2Rγ (e.g., a fusion thereof, such as a myc-myc-His6 fusion) (or binds to less than about 3.53×10 -7 M of K D Combined);

[0145] At 25°C, about 3.18×10 -9 M is about 2.38×10 -7 M of K D binds to cynomolgus IL-2Rγ (e.g., a fusion thereof, such as a myc-myc-His6 fusion);

[0146] At 37°C, the K of about 8.29×10-9M to about 3.20×10-7M D Binds to cynomolgus IL-2Rγ (e.g., a fusion thereof, such as a myc-myc-His6 fusion) (or binds to less than about 3.20×10 -7 M of K D Combined);

[0147] At 25°C, about 2.45×10 -9 M is about 1.20×10 -8K of M D binds to human IL2Rγ (e.g., its fusion, such as the fusion with the C-terminal murine IgG2a Fc tag) (or binds with a K less than about 1.20×10 -8 K of M D );

[0148] · at 37 °C with a K of about 1.86×10 -11 M to about 3.00×10 -8 K of M D binds to human IL2Rγ (e.g., its fusion, such as the fusion with the C-terminal murine IgG2a Fc tag) (or binds with a K less than about 3.00×10 -8 K of M D );

[0149] · at 25 °C with a K of about 1.84×10 -8 M, 3.76×10 -9 M, 1.08×10 -7 M, 2.17×10 -8 M, 6.02×10 - 9 M or 7.93×10 -8 K of M D binds (or does not bind) to murine IL2Rγ (e.g., its fusion, such as the myc-myc-His6 fusion);

[0150] · at 37 °C with a K of about 5.59×10 -8 M, 6.11×10 -9 M, 3.87×10 -7 M, 5.16×10 -8 M, 8.70×10 - 9 M or 2.15×10 -7 K of M D binds (or does not bind) to murine IL2Rγ (e.g., its fusion, such as the myc-myc-His6 fusion);

[0151] · at 25 °C with a K of about 3.32×10 -9 M to about 1.97×10 -7 K of M D binds (or does not bind) to human IL2Rγ domain 1 (e.g., its fusion, such as the myc-myc-His6 fusion);

[0152] · at 37 °C with a K of about 4.13×10 -9 M to about 2.25×10 -7 K of M DBinds (or does not bind) to human IL2Rγ domain 1 (e.g., its fusion, such as myc-myc-His6 fusion);

[0153] · At 25 °C at about 2.91×10 -7 M to about 5.35×10 -10 K D Binds (or does not bind) to human IL2Rγ domain 2 (e.g., its fusion, such as myc-myc-His6 fusion);

[0154] · At 37 °C at about 1.14×10 -8 or about 1.27×10 -8 K D Binds (or does not bind) to human IL2Rγ domain 2 (e.g., its fusion, such as myc-myc-His6 fusion);

[0155] · Blocks STAT phosphorylation in T cells (e.g., human CD4 + T cells), such as STAT phosphorylation induced by IL-2 (e.g., at about 10 nM), IL-4 (e.g., at about 50 pM), IL7 (e.g., at about 1 pM), IL-15 (e.g., at about 0.5 nM) and / or IL-21 (e.g., at about 50 pM), such as at about 1 nM to about 0.5 nM IC 50 ;

[0156] · Blocks STAT phosphorylation in mast cells (e.g., differentiated human mast cells), such as STAT phosphorylation induced by IL-9 (e.g., at about 2 nM), such as at about 4×10 -10 M IC 50 ;

[0157] · Reduces the number of human immune cells (e.g., human PBMC (peripheral blood mononuclear cells), such as human CD45+ cells, human T cells, human CD4+ T cells and / or human CD8+ T cells) in mice (e.g., NOD-scid IL2rγ null (NSG) mice) after injection with human peripheral blood mononuclear cells (PBMC);

[0158] · Reduces the levels of serum human cytokines (e.g., human IFN-γ, human TNFα, human IL-6, human IL-8 and / or human IL-10) and / or mouse cytokines (e.g., mouse TNFα, mouse IL-6, mouse KC / GRO and / or mouse IL-10) in mice (e.g., NOD-scidIL2rγnull (NSG) mice) after injection with human peripheral blood mononuclear cells (PBMC);

[0159] ·Competitive bind to human IL-2Rγ with any one or more of the anti-IL2Rγ antibodies shown herein, such as human IL-2Rγ on the cell surface (e.g., labeled with a C-terminal myc-myc-hexahistidine tag);

[0160] ·Bind to the same epitope on IL2Rγ as any one or more of the anti-IL2Rγ antibodies shown herein, such as IL2Rγ on the cell surface (e.g., labeled with a C-terminal myc-myc-hexahistidine tag);

[0161] ·Do not detectably bind to mouse or rat IL2Rγ (e.g., as measured by Biacore at 37°C);

[0162] ·Protect mice from weight loss and / or death due to GvHD in a GvHD mouse model;

[0163] ·Block the binding of a heteromeric receptor comprising IL2Rγ complexed with a cytokine-specific receptor subunit to cytokines such as IL-2, IL-4, IL-7, IL-9, IL-15, and / or IL-21; and / or

[0164] ·Inhibit IL2Rγ intracellular signaling through the JAK-STAT pathway induced by, for example, IL2, IL4, IL7, IL9, IL15, and / or IL21 (e.g., in human B lymphocytes or human natural killer cells), such as as measured by luciferase expression in cells comprising a luciferase gene operably linked to a STAT3 response element.

[0165] Unless otherwise indicated, "H4H12857P", "H4H12858P", "H4H12859P", "H4H12863P", "H4H12874P", "H4H12871P", "H4H12884P", "H4H12886P", "H4H12889P", "H4H12890P", "H4H12899P", "H4H12900P", "H4H12908P", "H4H12913P2", "H4H12922P2", "H4H12924P2", "H4H12926P2", "H4H12927P2", "H4H12934P2", "H4H13538P", "H4H13541P", "H4H13544P2", or "H4H13545P2" refer to an anti-IL2Rγ antigen-binding protein, such as an antibody and its antigen-binding fragments (including multispecific antigen-binding proteins), which respectively comprise: an immunoglobulin heavy chain or its variable region (V containing the amino acid sequences (e.g., SEQ ID NO: 2, 18, 22, 38, 42, 58, 62, 77, 81, 97, 101, 115, 119, 134, 138, 152, 156, 170, 174, 186, 190, 198, 200, 208, 210, 216, 218, 234, 238, 254, 258, 272, 276, 284, 286, 294, 296, 311, 315, 331, 335, 343, 345, 357, 361, or 376) (or variants thereof) specifically shown herein for H4H12857P, H4H12858P, H4H12859P, H4H12863P, H4H12874P, H4H12871P, H4H12884P, H4H12886P, H4H12889P, H4H12890P, H4H12899P, H4H12900P, H4H12908P, H4H12913P2, H4H12922P2, H4H12924P2, H4H12926P2, H4H12927P2, H4H12934P2, H4H13538P, H4H13541P, H4H13544P2, or H4H13545P2 H), and / or comprising an immunoglobulin light chain or variable region thereof containing the amino acid sequences specifically shown herein for H4H12857P, H4H12858P, H4H12859P, H4H12863P, H4H12874P, H4H12871P, H4H12884P, H4H12886P, H4H12889P, H4H12890P, H4H12899P, H4H12900P, H4H12908P, H4H12913P2, H4H12922P2, H4H12924P2, H4H12926P2, H4H12927P2, H4H12934P2, H4H13538P, H4H13541P, H4H13544P2 or H4H13545P2 (e.g., SEQ ID NO: 10, 20, 30, 40, 50, 60, 70, 79, 89, 99, 109, 117, 127, 136, 146, 154, 164, 172, 182, 188, 226, 236, 246, 256, 266, 274, 304, 313, 323, 333, 353, 359, 368 or 378) (or variants thereof) (or the variable region (V L ); and / or comprising: a heavy chain containing its CDRs (CDR-H1 (or variants thereof), CDR-H2 (or variants thereof) and CDR-H3 (or variants thereof)) or V H , and / or a light chain containing its CDRs (CDR-L1 (or variants thereof), CDR-L2 (or variants thereof) and CDR-L3 (or variants thereof)) or V L . In one embodiment of the invention, V H is linked to an IgG constant heavy chain domain, such as a human IgG constant heavy chain domain (e.g., IgG1 or IgG4 (e.g., containing the S228P and / or S108P mutations)); and / or V L is linked to a light chain constant domain, such as a human light chain constant domain (e.g., lambda or kappa constant light chain domain). Polynucleotides encoding one or more of any such immunoglobulin chains (e.g., V H , V L , HC and / or LC) form part of the invention.

[0166] The invention includes "neutralizing" or "antagonist" anti-IL2Rγ antigen-binding proteins (e.g., antibodies or antigen-binding fragments) that include molecules that inhibit IL2Rγ activity to any detectable extent (e.g., inhibit the binding of a heteromeric receptor comprising IL2Rγ complexed with a cytokine-specific receptor subunit to cytokines such as IL-2, IL-4, IL-7, IL-9, IL-15 and / or IL-21).

[0167] The antibodies and antigen-binding fragments of the present invention (e.g., H4H12857P; H4H12858P; H4H12859P; H4H12863P; H4H12874P; H4H12871P; H4H12884P; H4H12886P; H4H12889P; H4H12890P; H4H12899P; H4H12900P; H4H12908P; H4H12913P2; H4H12922P2; H4H12924P2; H4H12926P2; H4H12927P2; H4H12934P2; H4H13538P; H4H13541P; H4H13544P2; or H4H13545P2) comprise immunoglobulin chains containing the amino acid sequences (and variants thereof) specifically shown herein, as well as cellular and in vitro post-translational modifications of the antibody or fragment. For example, the present invention includes antibodies and antigen-binding fragments that specifically bind to IL2Rγ, which comprise the heavy chain and / or light chain amino acid sequences shown herein; and antibodies and fragments in which one or more asparagine, serine, and / or threonine residues are glycosylated, one or more asparagine residues are deamidated, one or more residues (e.g., Met, Trp, and / or His) are oxidized, the N-terminal glutamine is pyroglutamic acid (pyroE), and / or the C-terminal lysine or other amino acids are deleted.

[0168] The present invention provides containers (e.g., plastic or glass vials (e.g., with caps or columns), hollow needles, or syringe barrels) that contain the anti-IL2Rγ antigen-binding proteins of the present invention, e.g.,

[0169] H4H12857P; H4H12858P; H4H12859P; H4H12863P; H4H12874P; H4H12871P; H4H12884P: H4H12886P; H4H12889P; H4H12890P; H4H12899P; H4H12900P; H4H12908P; H4H12913P2; H4H12922P2; H4H12924P2; H4H12926P2; H4H12927P2; H4H12934P2; H4H13538P; H4H13541P; H4H13544P2; or H4H13545P2.

[0170] The present invention also provides injection devices that contain one or more antigen-binding proteins (e.g., antibodies or antigen-binding fragments) that specifically bind to IL2Rγ, e.g.,

[0171] H4H12857P; H4H12858P; H4H12859P; H4H12863P; H4H12874P; H4H12871P; H4H12884P; H4H12886P; H4H12889P; H4H12890P; H4H12899P; H4H12900P; H4H12908P; H4H12913P2; H4H12922P2; H4H12924P2; H4H12926P2; H4H12927P2; H4H12934P2; H4H13538P; H4H13541P; H4H13544P2; or H4H13545P2

[0172] , or a pharmaceutical formulation thereof. The injection device can be packaged in a medicine box. The injection device is a device for introducing a substance into an object's body through a parenteral route such as intraocular, intravitreal, intramuscular, subcutaneous, or intravenous. For example, the injection device can be a syringe or an autoinjector (e.g., pre-filled with a pharmaceutical formulation), which for example includes a barrel or cartridge for containing the fluid to be injected (e.g., containing an antibody or fragment or a pharmaceutical formulation thereof); a needle for piercing the skin, blood vessel, or other tissue for injecting the fluid; and a plunger for pushing the fluid out of the barrel and through the needle hole and into the object's body.

[0173] The present invention also provides a method for administering the anti-IL2Rγ antigen-binding protein of the present invention to an object, such as

[0174] H4H12857P; H4H12858P; H4H12859P; H4H12863P; H4H12874P; H4H12871P; H4H12884P; H4H12886P; H4H12889P; H4H12890P; H4H12899P; H4H12900P; H4H12908P; H4H12913P2; H4H12922P2; H4H12924P2: H4H12926P2: H4H12927P2: H4H12934P2: H4H13538P: H4H13541P:

[0175] such as H4H13544P2; or H4H13545P2, which includes introducing the antigen-binding protein into the body of an object (e.g., a human), such as parenterally. For example, the method includes piercing the object's body with a needle of a syringe and injecting the antigen-binding protein into the object's body, such as injecting it into the object's vein, artery, eye, muscle tissue, or subcutaneous tissue.

[0176] Polynucleotides and preparation methods

[0177] Polynucleotides include DNA and RNA. The present invention includes any polynucleotide of the present invention, for example, encoding

[0178] Immunoglobulin V of H4H12857P; H4H12858P; H4H12859P; H4H12863P; H4H12874P; H4H12871P; H4H12884P; H4H12886P; H4H12889P; H4H12890P; H4H12899P; H4H12900P; H4H12908P; H4H12913P2; H4H12922P2; H4H12924P2; H4H12926P2; H4H12927P2; H4H12934P2; H4H13538P; H4H13541P; H4H13544P2; or H4H13545P2 H 、V L 、CDR-H, CDR-L, HC or LC, optionally operably linked to a promoter or other expression control sequence. For example, the present invention provides any polynucleotide (e.g., DNA) comprising a nucleotide sequence shown in any of the following:

[0179] SEQ ID NO: 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31, 33, 35, 37, 39, 41, 43, 45, 47, 49, 51, 53, 55, 57, 59, 61, 63, 65, 67, 69, 71, 73, 74, 76, 78, 80, 82, 84, 86, 88, 90, 92, 94, 96, 98, 100, 102, 104, 106, 108, 110, 112, 114, 116, 118, 120, 122, 124, 126, 128, 130, 131, 133, 135, 137, 139, 141, 143, 145, 147, 149, 151, 153, 155, 157, 159, 161, 163, 165, 167, 169, 171, 173, 175, 175, 177, 179, 181, 183, 185, 187, 189, 191, 193, 195, 197, 199, 201, 203, 205, 207, 209, 211, 213, 215, 217, 219, 221, 223, 225, 227, 229, 231, 233, 235, 237, 239, 241, 243, 245, 247, 249, 251, 253, 255, 257, 259, 261, 263, 265, 267, 269, 271, 273, 275, 277, 279, 281, 283, 285, 287, 289, 291, 293, 295, 297, 299, 301, 303, 305, 307, 308, 310, 312, 314, 316, 318, 320 ， 322, 324, 326, 328, 330, 332, 334, 336, 338, 340, 342, 344, 346, 348, 350, 352, 354, 356, 358, 360, 362, 364, 365, 367, 369, 371, 373, 375 or 377.

[0180] . In one embodiment of the present invention, the polynucleotide of the present invention is fused with a secretion signal sequence. The polypeptide encoded by such a polynucleotide is also within the scope of the present invention.

[0181] Generally, a "promoter" or "promoter sequence" is a DNA regulatory region capable of binding RNA polymerase in a cell (e.g., directly or through other promoter-binding proteins or substances) and initiating transcription of a coding sequence. The promoter may be operably linked to other expression control sequences including enhancer and repressor sequences and / or to the polynucleotides of the present invention. Promoters useful for controlling gene expression include, but are not limited to, the cytomegalovirus (CMV) promoter (U.S. Pat. Nos. 5,385,839 and 5,168,062), the SV40 early promoter region (Benoist, et al., (1981) Nature 290:304-310), the promoter contained in the Rous sarcoma virus 3' long terminal repeat (Yamamoto, et al., (1980) Cell 22:787-797), the herpes thymidine kinase promoter (Wagner, et al., (1981) Proc. Natl. Acad. Sci. USA 78:1441-1445), the regulatory sequences of the metallothionein gene (Brinster, et al., (1982) Nature 296:39-42); prokaryotic expression vectors such as the β-lactamase promoter (Villa-Komaroff, et al., (1978) Proc. Natl. Acad. Sci. USA 75:3727-3731), or the tac promoter (DeBoer, et al., (1983) Proc. Natl. Acad. Sci. USA 80:21-25); see also "Useful proteins from recombinant bacteria", Scientific American (1980) 242:74-94; and promoter elements from yeast or other fungi such as the Gal4 promoter, the ADC (alcohol dehydrogenase) promoter, the PGK (phosphoglycerate kinase) promoter or the alkaline phosphatase promoter.

[0182] A polynucleotide encoding a polypeptide is "operably linked" to a promoter or other expression control sequence when, in a cell or other expression system, the sequence directs the RNA polymerase-mediated transcription of the coding sequence into RNA, preferably mRNA, which can then be RNA spliced (if it contains introns) and optionally, translated into the protein encoded by the coding sequence.

[0183] The present invention includes polynucleotides comprising pairs of polynucleotides encoding V H and V L :

[0184] SEQ ID NO: 1 and SEQ ID NO: 9;

[0185] SEQ ID NO: 21 and SEQ ID NO: 29;

[0186] SEQ ID NO: 41 and SEQ ID NO: 49;

[0187] SEQ ID NO: 61 and SEQ ID NO: 69;

[0188] SEQ ID NO: 80 and SEQ ID NO: 88;

[0189] SEQ ID NO: 100 and SEQ ID NO: 108;

[0190] SEQ ID NO: 118 and SEQ ID NO: 126;

[0191] SEQ ID NO: 137 and SEQ ID NO: 145;

[0192] SEQ ID NO: 155 and SEQ ID NO: 163;

[0193] SEQ ID NO: 173 and SEQ ID NO: 181;

[0194] SEQ ID NO: 189 and SEQ ID NO: 181;

[0195] SEQ ID NO: 199 and SEQ ID NO: 181;

[0196] SEQ ID NO: 209 and SEQ ID NO: 181;

[0197] SEQ ID NO: 217 and SEQ ID NO: 225;

[0198] SEQ ID NO: 237 and SEQ ID NO: 245;

[0199] SEQ ID NO: 257 and SEQ ID NO: 265;

[0200] SEQ ID NO: 275 and SEQ ID NO: 181;

[0201] SEQ ID NO: 285 and SEQ ID NO: 181;

[0202] SEQ ID NO: 295 and SEQ ID NO: 303;

[0203] SEQ ID NO: 314 and SEQ ID NO: 322;

[0204] SEQ ID NO: 334 and SEQ ID NO: 181;

[0205] SEQ ID NO: 344 and SEQ ID NO: 352; or

[0206] SEQ ID NO: 360 and SEQ ID NO: 367.

[0207] The present invention includes polynucleotides comprising a polynucleotide group encoding the following CDR-H1, CDR-H2, CDR-H3, CDR-L1, CDR-L2 and CDR-L3:

[0208] SEQ ID NO: 3, 5, 7, 11, 13 and 15;

[0209] SEQ ID NO: 23, 25, 27, 31, 33 and 35;

[0210] SEQ ID NO: 43, 45, 47, 51, 53 and 55;

[0211] SEQ ID NO: 63, 65, 67, 71, 73 and 74;

[0212] SEQ ID NO: 82, 84, 86, 90, 92 and 94;

[0213] SEQ ID NO: 102, 104, 106, 110, 73 and 112;

[0214] SEQ ID NO: 120, 122, 124, 128, 130 and 131;

[0215] SEQ ID NO: 139, 141, 143, 147, 73 and 149;

[0216] SEQ ID NO: 157, 159, 161, 165, 13 and 167;

[0217] SEQ ID NO: 175, 177, 179, 71, 73 and 183;

[0218] SEQ ID NO: 191, 193, 195, 71, 73 and 183;

[0219] SEQ ID NO: 201, 203, 205, 71, 73 and 183;

[0220] SEQ ID NO: 175, 211, 213, 71, 73 and 183;

[0221] SEQ ID NO: 219, 221, 223, 227, 229 and 231;

[0222] SEQ ID NO: 239, 241, 243, 247, 249 and 251;

[0223] SEQ ID NO: 259, 261, 263, 267, 73 and 269;

[0224] SEQ ID NO: 277, 279, 281, 71, 73 and 183;

[0225] SEQ ID NO: 287, 289, 291, 71, 73 and 183;

[0226] SEQ ID NO: 297, 299, 301, 305, 307 and 308;

[0227] SEQ ID NO: 316, 318, 320, 324, 326 and 328;

[0228] SEQ ID NO: 336, 338, 340, 71, 73 and 183;

[0229] SEQ ID NO: 346, 348, 350, 71, 73 and 354; or

[0230] SEQ ID NO: 362, 364, 365, 369, 371 and 373.

[0231] The present invention includes polynucleotides comprising polynucleotide pairs encoding HC and LC as follows:

[0232] SEQ ID NO: 17 and SEQ ID NO: 19;

[0233] SEQ ID NO: 37 and SEQ ID NO: 39;

[0234] SEQ ID NO: 57 and SEQ ID NO: 59;

[0235] SEQ ID NO: 76 and SEQ ID NO: 78;

[0236] SEQ ID NO: 96 and SEQ ID NO: 98;

[0237] SEQ ID NO: 114 and SEQ ID NO: 116;

[0238] SEQ ID NO: 133 and SEQ ID NO: 135;

[0239] SEQ ID NO: 151 and SEQ ID NO: 153;

[0240] SEQ ID NO: 169 and SEQ ID NO: 171;

[0241] SEQ ID NO: 185 and SEQ ID NO: 187;

[0242] SEQ ID NO: 197 and SEQ ID NO: 187;

[0243] SEQ ID NO: 207 and SEQ ID NO: 187;

[0244] SEQ ID NO: 215 and SEQ ID NO: 187;

[0245] SEQ ID NO: 233 and SEQ ID NO: 235;

[0246] SEQ ID NO: 253 and SEQ ID NO: 255;

[0247] SEQ ID NO: 271 and SEQ ID NO: 273;

[0248] SEQ ID NO: 283 and SEQ ID NO: 187;

[0249] SEQ ID NO: 293 and SEQ ID NO: 187;

[0250] SEQ ID NO: 310 and SEQ ID NO: 312;

[0251] SEQ ID NO: 330 and SEQ ID NO: 332;

[0252] SEQ ID NO: 342 and SEQ ID NO: 187;

[0253] SEQ ID NO: 356 and SEQ ID NO: 358; or

[0254] SEQ ID NO: 375 and SEQ ID NO: 377.

[0255] The present invention includes polynucleotides encoding immunoglobulin polypeptide chains, which polynucleotides are variants of those whose nucleotide sequences are specifically shown herein. A "variant" of a polynucleotide refers to a polynucleotide comprising a nucleotide sequence that is at least about 70-99.9% (e.g., 70%, 72%, 74%, 75%, 76%, 79%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 99.5%, 99.9%) identical to the reference nucleotide sequence shown herein; when compared by the BLAST algorithm, wherein the parameters of the algorithm are selected to give the maximum match between the sequences over the entire length of each reference sequence (e.g., expectation threshold: 10; word length: 28; maximum number of matches in query range: 0; match / mismatch scores: 1, -2; gap penalties: linear). In one embodiment of the invention, variants of the nucleotide sequences specifically shown herein comprise one or more (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 or 12) point mutations, insertions (e.g., in-frame insertions) or deletions (e.g., in-frame deletions) of one or more nucleotides. In one embodiment of the invention, such mutations can be missense mutations or nonsense mutations. In one embodiment of the invention, such variant polynucleotides encode immunoglobulin polypeptide chains that can be incorporated into an anti-IL2Rγ antigen-binding protein, i.e., such that the protein retains specific binding to IL2Rγ.

[0256] Eukaryotic and prokaryotic host cells, including mammalian cells, can be used as hosts for expressing anti-IL2Rγ antigen-binding proteins (e.g., antibodies or antigen-binding fragments thereof). Such host cells are well known in the art and many are available from the American Type Culture Collection (ATCC). These host cells include, in particular, Chinese hamster ovary (CHO) cells, NSO, SP2 cells, HeLa cells, baby hamster kidney (BHK) cells, monkey kidney cells (COS), human hepatocellular carcinoma cells (e.g., Hep G2), A549 cells, 3T3 cells, HEK-293 cells, and many other cell lines. Mammalian host cells include cells from humans, mice, rats, dogs, monkeys, pigs, goats, cows, horses, and hamsters. Other cell lines that can be used are insect cell lines (e.g., Spodoptera frugiperda or Trichoplusia ni), amphibian cells, bacterial cells, plant cells, and fungal cells.Fungal cells include cells of yeasts and filamentous fungi, including, for example, Pichia, Pichia pastoris, Pichia finlandica, Pichia trehalophila, Pichia koclamae, Pichia membranaefaciens, Pichia minuta (Ogataea minuta, Pichia lindneri), Pichia opuntiae, Pichia thermotolerans, Pichia salictaria, Pichia quercuum, Pichia pijperi, Pichia stiptis, Pichia methanolica, Pichia sp., Saccharomyces cerevisiae, Saccharomyces sp., Hansenula polymorpha, Kluyveromyces sp., Kluyveromyces lactis, Candida albicans, Aspergillus nidulans, Aspergillus niger, Aspergillus oryzae, Trichoderma reesei, Chrysosporium lucknowense, Fusarium sp., Fusarium gramineum, Fusarium venenatum, Physcomitrella patens, and Neurospora crassa. The invention includes isolated host cells (e.g., CHO cells or any of the above types of host cells) that contain an antigen-binding protein, V. H 、V L, HC, LC, or their CDRs (or variants thereof), such as H4H12857P; H4H12858P; H4H12859P; H4H12863P; H4H12874P; H4H12871P; H4H12884P; H4H12886P; H4H12889P; H4H12890P; H4H12899P; H4H12900P; H4H12908P; H4H12913P2; H4H12922P2; H4H12924P2; H4H12926P2; H4H12927P2; H4H12934P2; H4H13538P; H4H13541P; H4H13544P2; or H4H13545P2;

[0257] ; and / or a polynucleotide encoding one or more of its immunoglobulin chains (e.g., as discussed herein).

[0258] The invention also includes cells expressing IL2Rγ or an antigenic fragment or fusion (e.g., His6, Fc, and / or myc) thereof that binds to an antigen-binding protein of the invention (e.g., an antibody or an antigen-binding fragment thereof) such as: H4H12857P; H4H12858P; H4H12859P; H4H12863P; H4H12874P; H4H12871P; H4H12884P; H4H12886P; H4H12889P; H4H12890P; H4H12899P; H4H12900P; H4H12908P; H4H12913P2; H4H12922P2; H4H12924P2; H4H12926P2; H4H12927P2; H4H12934P2; H4H13538P; H4H13541P; H4H13544P2; or H4H13545P2, e.g., wherein the cells are in vivo or in vitro in a subject.

[0259] In addition, the invention also provides a complex comprising an anti-IL2Rγ antigen-binding protein as discussed herein, e.g., an antibody or an antigen-binding fragment thereof, complexed with an IL2Rγ polypeptide or an antigenic fragment or fusion thereof and / or with a secondary antibody or an antigen-binding fragment thereof (e.g., a detectably labeled secondary antibody) that specifically binds to an anti-IL2Rγ antibody or fragment. In one embodiment of the invention, the complex is in vitro (e.g., immobilized on a solid substrate) or in vivo in a subject.

[0260] The recombinant anti-IL2Rγ antigen-binding proteins disclosed herein, e.g., antibodies and antigen-binding fragments, can also be produced in an E. coli / T7 expression system. In this embodiment, encoding the anti-IL2Rγ antibody immunoglobulin molecule of the invention (e.g.

[0261] HC, LC, V of H4H12857P; H4H12858P; H4H12859P; H4H12863P; H4H12874P; H4H12871P; H4H12884P; H4H12886P; H4H12889P; H4H12890P; H4H12899P; H4H12900P; H4H12908P; H4H12913P2; H4H12922P2; H4H12924P2; H4H12926P2; H4H12927P2; H4H12934P2; H4H13538P; H4H13541P; H4H13544P2; or H4H13545P2 H and / or V L Polynucleotides encoding the same (e.g., CDR) can be inserted into pET-based plasmids and expressed in the E. coli / T7 system. For example, the present invention includes a method for expressing an antibody or an antigen-binding fragment thereof or an immunoglobulin chain thereof in a host cell (e.g., a bacterial host cell such as E. coli such as BL21 or BL21DE3), the method comprising expressing T7 RNA polymerase in the cell, which cell further comprises a polynucleotide encoding an immunoglobulin chain operably linked to a T7 promoter (e.g., comprising a nucleotide sequence of any one or more of the following, or a variant thereof:

[0262] SEQ ID NO: 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31, 33, 35, 37, 39, 41, 43, 45, 47, 49, 51, 53, 55, 57, 59, 61, 63, 65, 67, 69, 71, 73, 74, 76, 78, 80, 82, 84, 86, 88, 90, 92, 94, 96, 98, 100, 102, 104, 106, 108, 110, 112, 114, 116, 118, 120, 122, 124, 126, 128, 130, 131, 133, 135, 137, 139, 141, 143, 145, 147, 149, 151, 153, 155, 157, 159, 161, 163, 165, 167, 169, 171, 173, 175, 175, 177, 179, 181, 183, 185, 187, 189, 191, 193, 195, 197, 199, 201, 203, 205, 207, 209, 211, 213, 215, 217, 219, 221, 223, 225, 227, 229, 231, 233, 235, 237, 239, 241, 243, 245, 247, 249, 251, 253, 255, 257, 259, 261, 263, 265, 267, 269, 271, 273, 275, 277, 279, 281, 283, 285, 287, 289, 291, 293, 295, 297, 299, 301, 303, 305, 307, 308, 310, 312, 314, 316, 318, 320, 322, 324, 326, 328, 330, 332, 334, 336, 338, 340, 342, 344, 346, 348, 350, 352, 354, 356, 358, 360, 362, 364, 365, 367, 369, 371, 373, 375 or 377). For example, in one embodiment of the present invention, a bacterial host cell, such as Escherichia coli, contains a polynucleotide encoding a T7 RNA polymerase gene operably linked to a lac promoter, and the expression of the polymerase and the strand is induced by incubating the host cell with IPTG (isopropyl-β-D-thiogalactopyranoside).See US4952496 and US5693489 or Studier & Moffatt, Use of bacteriophage T7 RNA polymerase to direct selective high-level expression of cloned genes, J. Mol. Biol. May 5, 1986; 189(1): 113-30.

[0263] Several methods for producing recombinant antibodies are known in the art. US4816567 discloses an example of a method for the recombinant production of antibodies.

[0264] Transformation can be carried out by any known method for introducing polynucleotides into a host cell. Methods for introducing heterologous polynucleotides into mammalian cells are well known in the art and include dextran-mediated transfection, calcium phosphate precipitation, polybrene-mediated transfection, protoplast fusion, electroporation, encapsulation of polynucleotides (multiple polynucleotides) in liposomes, biolistic injection, and direct microinjection of DNA into the nucleus. In addition, nucleic acid molecules can be introduced into mammalian cells by viral vectors. Methods for transforming cells are well known in the art. See, for example, U.S. Pat. Nos. 4,399,216, 4,912,040, 4,740,461, and 4,959,455. Accordingly, the present invention includes recombinant methods for preparing the anti-IL2Rγ antigen-binding proteins of the present invention, such as antibodies or antigen-binding fragments thereof, or immunoglobulin chains thereof, the method comprising (i) introducing one or more polynucleotides encoding an antigen-binding protein, such as

[0265] the immunoglobulin light chain and / or heavy chain of H4H12857P; H4H12858P; H4H12859P; H4H12863P; H4H12874P; H4H12871P; H4H12884P; H4H12886P; H4H12889P; H4H12890P; H4H12899P; H4H12900P; H4H12908P; H4H12913P2; H4H12922P2; H4H12924P2; H4H12926P2; H4H12927P2; H4H12934P2; H4H13538P; H4H13541P; H4H13544P2; or H4H13545P2 (e.g., comprising the nucleotide sequence in any one or more of the following, or a variant thereof:

[0266] SEQ ID NO: 1, 9, 17, 19, 21, 29, 37, 39, 41, 49, 57, 59, 61, 69, 76, 78, 80, 88, 96, 98, 100, 108, 114, 116, 118, 126, 133, 135, 137, 145, 151, 153, 155, 163, 169, 171, 173, 181, 185, 187, 189, 197, 199, 207, 209, 215, 217, 225, 233, 235, 237, 245, 253, 255, 257, 265, 271, 273, 275, 283, 285, 293, 295, 303, 310, 312, 314, 322, 330, 332, 334, 342, 344, 352, 356, 358, 360, 367, 375 or 377) is introduced into a host cell, e.g., wherein the polynucleotide is in a vector; and / or integrated into the host cell chromosome and / or operably linked to a promoter; (ii) culturing the host cell (e.g., CHO or Pichia or Pichia pastoris) under conditions conducive to the expression of the polynucleotide, and (iii) optionally, isolating the antigen-binding protein (e.g., antibody or antigen-binding fragment) or chain from the host cell and / or the medium in which the host cell was cultured. When preparing an antigen-binding protein comprising more than one immunoglobulin chain (e.g., an antibody or antigen-binding fragment), e.g., preparing an antibody comprising two immunoglobulin heavy chains and two immunoglobulin light chains, co-expression of these chains in a single host cell results in the association of the chains, e.g., intracellularly or on the cell surface or extracellularly (if such chains are secreted), to form the antigen-binding protein (e.g., an antibody or antigen-binding fragment). The methods of the invention include those wherein only an immunoglobulin heavy chain or only an immunoglobulin light chain or both are expressed in the cell (e.g., any of the methods discussed herein include mature fragments and / or their variable domains). For example, such single chains are available, e.g., useful as intermediates in the expression of an antibody or antigen-binding fragment comprising such a chain. For example, the invention also includes anti-IL2Rγ antigen-binding proteins, e.g., antibodies and their antigen-binding fragments, which are products of the production methods described herein and optionally products of the purification methods described herein.

[0267] In one embodiment of the invention, a method for preparing an anti-IL2Rγ antigen-binding protein (e.g., an antibody or antigen-binding fragment thereof) includes purifying the antigen-binding protein, e.g., by column chromatography, precipitation, and / or filtration. As discussed, the products of such methods also form part of the invention.

[0268] Preparation of human antibodies

[0269] The anti-IL2Rγ antibody of the present invention can be a fully human antibody. Methods for generating monoclonal antibodies, including fully human monoclonal antibodies, are known in the art. Any such known method can be used in the context of the present invention to prepare human antibodies that specifically bind to human IL2Rγ.

[0270] Using, for example, VELOCIMMUNE TM technology or any other similar known method for generating fully human monoclonal antibodies, a high-affinity chimeric antibody against IL2Rγ having a human variable region and a murine constant region is first isolated. As described in the experimental section below, the antibody is characterized and selected for desired characteristics, including affinity, ligand blocking activity, selectivity, epitope, etc. If necessary, the murine constant region is replaced with a desired human constant region, such as wild-type or modified IgG1 or IgG4, to generate a fully human anti-IL2Rγ antibody. Although the selected constant region may vary depending on the particular use, the high-affinity antigen binding and target specificity are characterized by the variable region. In some cases, the fully human anti-IL2Rγ antibody is directly isolated from antigen-positive B cells. See, for example, US 6,596,541, Regeneron Pharmaceuticals,

[0271] Anti-IL2Rγ antibodies comprising Fc variants

[0272] According to certain embodiments of the present invention, anti-IL2Rγ antibodies comprising an Fc domain are provided, the Fc domain comprising one or more mutations that enhance or attenuate, for example, enhance or attenuate the binding of the antibody to the FcRn receptor at acidic pH compared to neutral pH. For example, the present invention includes anti-IL2Rγ antibodies comprising mutations in the C H 2 or C H 3 region, wherein one or more mutations increase the affinity of the Fc domain for FcRn in an acidic environment (e.g., in endosomes having a pH range of about 5.5 to about 6.0). When administered to an animal, such mutations can result in an extended serum half-life of the antibody.

[0273] Non-limiting examples of such Fc modifications include, for example, modifications at the following sites:

[0274] · 250 (e.g., E or Q);

[0275] · 250 and 428 (e.g., L or F);

[0276] · 252 (e.g., L / Y / F / W or T);

[0277] · 254 (e.g., S or T); and / or

[0278] · 256 (e.g., S / R / Q / E / D or T);

[0279] and / or modifications at the following sites:

[0280] · 428 and / or 433 (e.g., H / L / R / S / P / Q or K), and / or

[0281] · 434 (e.g., H / F or Y);

[0282] and / or modifications at the following sites:

[0283] · 250 and / or 428;

[0284] and / or modifications at the following sites:

[0285] · 307 or 308 (e.g., 308F, V308F), and / or

[0286] · 434.

[0287] In one embodiment of the present invention, the modifications include:

[0288] · 428L (e.g., M428L) and 434S (e.g., N434S) modifications;

[0289] · 428L, 259I (e.g., V259I), and 308F (e.g., V308F) modifications;

[0290] · 433K (e.g., H433K) and 434 (e.g., 434Y) modifications;

[0291] · 252, 254 and 256 (e.g., 252Y, 254T and 256E) modifications;

[0292] · 250Q and 428L modifications (e.g., T250Q and M428L); and / or

[0293] · 307 and / or 308 modifications (e.g., 308F or 308P).

[0294] For example, the present invention includes an anti-IL2Rγ antibody comprising an Fc domain, the Fc domain comprising one or more pairs or groups of mutations selected from the following:

[0295] · 250Q and 248L (e.g., T250Q and M248L);

[0296] · 252Y, 254T and 256E (e.g., M252γ, S254T and T256E);

[0297] · 428L and 434S (e.g., M428L and N434S); and

[0298] · 433K and 434F (such as H433K and N434F).

[0299] In one embodiment of the invention, the heavy chain constant domain is γ4 comprising the S228P and / or S108P mutations. See Angal et al. A single amino acid substitution abolishes the heterogeneity of chimeric mouse / human (IgG4) antibody, Mol Immunol. 1993 Jan; 30(1): 105-108.

[0300] All possible combinations of the above Fc domain mutations and other mutations within the antibody variable domains disclosed herein are expected to be within the scope of the invention.

[0301] The anti-IL2Rγ antibodies of the invention may comprise a modified Fc domain having reduced effector function. As used herein, "modified Fc domain having reduced effector function" means any Fc portion of an immunoglobulin that has been modified, mutated, truncated, etc. relative to the wild-type, naturally occurring Fc domain, such that a molecule comprising the modified Fc exhibits a reduction in the severity or degree of at least one effector selected from cell killing (e.g., ADCC and / or CDC), complement activation, phagocytosis, and opsonization relative to a comparator molecule comprising the wild-type, naturally occurring form of the Fc portion. In certain embodiments, the "modified Fc domain having reduced effector function" is an Fc domain with reduced or attenuated binding to an Fc receptor (e.g., FcγR).

[0302] In certain embodiments of the invention, the modified Fc domain is a variant IgG1 Fc or variant IgG4 Fc that comprises a substitution in the hinge region. For example, the modified Fc for use in the context of the present invention may comprise a variant IgG1 Fc in which at least one amino acid of the IgG1 Fc hinge region is replaced with the corresponding amino acid from the IgG2 Fc hinge region. Alternatively, the modified Fc for use in the context of the present invention may comprise a variant IgG4 Fc in which at least one amino acid of the IgG4 Fc hinge region is replaced with the corresponding amino acid from the IgG2 Fc hinge region. Non-limiting, exemplary modified Fc regions that may be used in the context of the present invention and any functional equivalent variants of the modified Fc regions described therein are set forth in U.S. Patent Application Publication No. 2014 / 0243504, the disclosure of which is hereby incorporated by reference in its entirety.

[0303] Other modified Fc domains and Fc modifications that can be used in the context of the present invention include any modifications as described in US2014 / 0171623, US8697396, US2014 / 0134162, WO2014 / 043361, the disclosures of which are hereby incorporated by reference in their entirety. Methods for constructing antibodies or other antigen-binding fusion proteins comprising a modified Fc domain as described herein are known in the art.

[0304] Multispecific antigen-binding protein

[0305] The present invention includes anti-IL2Rγ antigen-binding proteins, such as antibodies and antigen-binding fragments thereof, and methods of using and preparing such antigen-binding proteins. The terms "anti-IL2Rγ" or "anti-IL2R gamma" antigen-binding proteins, such as antibodies or antigen-binding fragments, include multispecific (e.g., bispecific or biparatopic) molecules that comprise at least a first antigen-binding domain that specifically binds to IL2Rγ (e.g., an antigen-binding domain from H4H12857P; H4H12858P; H4H12859P; H4H12863P; H4H12874P; H4H12871P; H4H12884P; H4H12886P; H4H12889P; H4H12890P: H4H12899P; H4H12900P; H4H12908P; H4H12913P2; H4H12922P2; H4H12924P2; H4H12926P2; H4H12927P2; H4H12934P2; H4H13538P; H4H13541P; H4H13544P2; or H4H13545P2) and at least a second antigen-binding domain that binds to a different antigen or an epitope on IL2Rγ that is different from the epitope of the first antigen-binding domain. In one embodiment of the present invention, the first and second epitopes overlap. In another embodiment of the present invention, the first and second epitopes do not overlap.

[0306] Multispecific binding refers to binding to two or more different epitopes, which may be on the same antigen or different antigens. Multispecificity includes bispecificity, trispecificity, and tetraspecificity.

[0307] “H4H12857P”; “H4H12858P”; “H4H12859P”; “H4H12863P”; “H4H12874P”; “H4H12871P”; “H4H12884P”; “H4H12886P”: “H4H12889P”; “H4H12890P”: “H4H12899P”; “H4H12900P”; “H4H12908P”; “H4H12913P2”; “H4H12922P2”; “H4H12924P2”; “H4H12926P2”; “H4H12927P2”; “H4H12934P2”; “H4H13538P”: “H4H13541P”; “H4H13544P2”; or “H4H13545P2”

[0308] Comprising multispecific molecules, such as antibodies or antigen-binding fragments, each of which comprises

[0309] The HCDR and LCDR, V of “H4H12857P”: “H4H12858P”; “H4H12859P”; “H4H12863P”; “H4H12874P”; “H4H12871P”; “H4H12884P”; “H4H12886P”; “H4H12889P”; “H4H12890P”; “H4H12899P”; “H4H12900P”; “H4H12908P”; “H4H12913P2”; “H4H12922P2”; “H4H12924P2”; “H4H12926P2”; “H4H12927P2”; “H4H12934P2”; “H4H13538P”; “H4H13541P”; “H4H13544P2”; or “H4H13545P2” H and V L 、or HC and LC, and one or more antigen-binding domains that bind to different epitopes.

[0310] In one embodiment of the invention, the antigen-binding domain that can be included in the multispecific molecule and specifically binds to IL2Rγ comprises: (1)

[0312] (i) Heavy chain variable domain (V H) A sequence comprising CDR-H1, CDR-H2, and CDR-H3 from an immunoglobulin heavy chain, said immunoglobulin heavy chain comprising an amino acid sequence selected from: SEQ ID NO: 2, 22, 42, 62, 81, 101, 119, 138, 156, 174, 190, 200, 210, 218, 238, 258, 276, 286, 296, 315, 335, 345, and 361 (or variants thereof); and

[0313] (ii) A light chain variable domain (V L ) sequence comprising CDR-L1, CDR-L2, and CDR-L3 from an immunoglobulin light chain, said immunoglobulin light chain comprising an amino acid sequence selected from: SEQ ID NO: 10, 30, 50, 70, 89, 109, 127, 146, 164, 182, 226, 246, 266, 304, 323, 353, and 368 (or variants thereof);

[0314] Or, (2)

[0316] (i) A heavy chain variable domain (V H ) comprising an amino acid sequence selected from: SEQ ID NO: 2, 22, 42, 62, 81, 101, 119, 138, 156, 174, 190, 200, 210, 218, 238, 258, 276, 286, 296, 315, 335, 345, and 361 (or variants thereof); and

[0317] (ii) A light chain variable domain (V L ) comprising an amino acid sequence selected from: SEQ ID NO: 10, 30, 50, 70, 89, 109, 127, 146, 164, 182, 226, 246, 266, 304, 323, 353, and 368 (or variants thereof);

[0318] And

[0319] One or more antigen-binding domains that bind to different epitopes.

[0320] In one embodiment of the invention, the bispecific antigen-binding fragment comprises a first scFv having binding specificity for a first epitope (e.g., IL2Rγ) (e.g., comprising

[0321] V of H4H12857P; H4H12858P; H4H12859P; H4H12863P; H4H12874P; H4H12871P; H4H12884P; H4H12886P; H4H12889P; H4H12890P; H4H12899P; H4H12900P; H4H12908P; H4H12913P2; H4H12922P2; H4H12924P2; H4H12926P2; H4H12927P2; H4H12934P2; H4H13538P; H4H13541P; H4H13544P2; or H4H13545P2 H and V L ) and a second scFv that has binding specificity for a second, different epitope. For example, in one embodiment of the invention, the first and second scFvs are linked by a linker, such as a peptide linker (e.g., a GS linker, such as (GGGGS)n (SEQ ID NO: 386), where n is, for example, 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10).

[0322] Other bispecific antigen-binding fragments include the F(ab)2 of a bispecific IgG antibody, which comprises

[0323] the heavy and light chain CDRs of H4H12857P; H4H12858P; H4H12859P; H4H12863P; H4H12874P; H4H12871P; H4H12884P; H4H12886P; H4H12889P; H4H12890P; H4H12899P; H4H12900P; H4H12908P; H4H12913P2; H4H12922P2; H4H12924P2; H4H12926P2; H4H12927P2; H4H12934P2; H4H13538P; H4H13541P; H4H13544P2; or H4H13545P2 and the heavy and light chain CDRs of another antibody that binds to a different epitope.

[0324] Immunoconjugates

[0325] The invention encompasses an anti-IL2Rγ antigen-binding protein, such as an antibody or antigen-binding fragment, conjugated to another moiety, such as a therapeutic moiety ("immunoconjugate"). In one embodiment of the invention, the anti-IL2Rγ antigen-binding protein (such as an antibody or antigen-binding fragment) is conjugated to any of the other therapeutic agents described herein. As used herein, the term "immunoconjugate" refers to an antigen-binding protein, such as an antibody or antigen-binding fragment, that is chemically or biologically linked to another antigen-binding protein, drug, radioactive agent, reporter moiety, enzyme, peptide, protein, or therapeutic agent.

[0326] Administration and treatment

[0327] The present invention provides a method for treating or preventing an IL2Rγ-mediated disease or disorder in a subject, the method comprising administering to the subject a therapeutically effective dose of an anti-IL2Rγ antigen-binding protein

[0328] H4H12857P; H4H12858P; H4H12859P; H4H12863P; H4H12874P; H4H12871P; H4H12884P; H4H12886P; H4H12889P; H4H12890P; H4H12899P; H4H12900P; H4H12908P; H4H12913P2; H4H12922P2; H4H12924P2; H4H12926P2; H4H12927P2; H4H12934P2; H4H13538P; H4H13541P; H4H13544P2; or (H4H13545P2).

[0329] "IL2Rγ-mediated disease or disorder" refers to any disease disorder whose symptoms are mediated by one or more of the cytokines IL-2, IL-4, IL-7, IL-9, IL-15, and IL-21 and / or the activity of a receptor that binds such cytokines; for example, autoimmunity and / or inflammation mediated by such cytokines and / or receptors. For example, IL2Rγ-mediated diseases or disorders include graft-versus-host disease (GvHD), organ transplant rejection (e.g., skin transplantation (skin graft), b-islet cell graft, heart transplantation, lung transplantation, kidney transplantation, and / or liver transplantation), birdshot chorioretinopathy, multiple sclerosis, uveitis, autoimmune diseases (e.g., type I diabetes, multiple sclerosis, rheumatoid arthritis, systemic lupus erythematosus, and myasthenia gravis), aplastic anemia; atopic dermatitis; asthma; and mast cell activation disorders (e.g., mast cell activation syndrome (MCAS), systemic mastocytosis (SM), or mast cell leukemia (MCL)).

[0330] The present invention also includes an antigen-binding protein (e.g., an antibody or an antigen-binding fragment thereof) that specifically binds to IL2Rγ for administration to a subject, such as a subject suffering from an IL2Rγ-mediated disease or disorder, for example

[0331] A method of H4H12857P; H4H12858P; H4H12859P; H4H12863P; H4H12874P; H4H12871P; H4H12884P; H4H12886P; H4H12889P; H4H12890P; H4H12899P; H4H12900P; H4H12908P; H4H12913P2; H4H12922P2; H4H12924P2; H4H12926P2; H4H12927P2; H4H12934P2; H4H13538P; H4H13541P; H4H13544P2; or H4H13545P2, the method comprising introducing an antigen-binding protein into a subject, such as by injection.

[0332] GvHD is a condition that can occur after allogeneic transplantation. For example, in GvHD, donated bone marrow or peripheral blood stem cells can recognize the recipient's body as foreign, and the donated cells / bone marrow attack the body. GvHD can occur, for example, after hematopoietic cell transplantation (HCT; e.g., in a subject with acute myeloid leukemia (AML) or acute lymphocytic leukemia (ALL) and / or myelodysplastic syndrome or myeloproliferative neoplasm), blood transfusion, thymus transplantation, or in a patient with thymoma. Types of GvHD include steroid-refractory GvHD, acute graft-versus-host disease (aGvHD), and chronic graft-versus-host disease (cGvHD). An allogeneic transplant recipient may experience aGvHD or cGvHD or both forms, or neither. The present invention includes methods for treating or preventing GvHD (any type) in a subject, which comprise administering to the subject a therapeutically effective dose of an anti-IL2Rγ antigen-binding protein.

[0333] Symptoms of aGvHD can include a rash or red areas on the skin (signs of cutaneous aGvHD); yellow discoloration of the skin and / or eyes, and abnormal blood test results (signs of hepatic aGvHD); nausea, vomiting, diarrhea, or abdominal cramps (signs of aGvHD in the digestive tract or "gut"); and / or increased dryness / irritation of the eyes (signs of ocular GvHD).

[0334] The symptoms of cGvHD can include rashes, raised or discolored areas, thickening or tightening of the skin (signs of cutaneous cGvHD); abdominal distension, yellow discoloration of the skin and / or eyes, and abnormal blood test results (signs of hepatic cGvHD); dry eyes or changes in vision (signs of ocular cGvHD); dry mouth, white patches in the mouth, pain, or sensitivity to spicy foods (signs of oral cGvHD - oral); shortness of breath or changes seen on a chest X-ray (signs of dry cough pulmonary cGvHD - pulmonary); difficulty swallowing, pain on swallowing, or weight loss (signs of gastrointestinal or "intestinal" cGvHD); fatigue, muscle weakness, or pain (signs of neuromuscular cGvHD - neural and muscular); and / or increased need to urinate (frequency), burning or bleeding during urination, vaginal dryness / tightening, or penile dysfunction (signs of urogenital, bladder, or sexual organ cGvHD).

[0335] Organ transplant rejection refers to the rejection of a transplanted organ by the recipient's immune system. Hyperacute rejection occurs within minutes after transplantation, acute rejection occurs within one week to 3 months after transplantation, and chronic rejection occurs over many years. Transplanted organs include, for example, solid organs such as skin, pancreas, kidney, liver, heart, and lung. The present invention includes methods for treating or preventing organ transplantation (of any kind) in a subject, the method comprising administering to the subject a therapeutically effective dose of an anti-IL2Rγ antigen-binding protein.

[0336] Birdshot chorioretinopathy is a rare form of posterior uveitis - an inflammation of the uvea, which is the part of the eye that provides most of the blood supply to the retina. Birdshot chorioretinopathy can be caused by autoimmunity. The symptoms of birdshot chorioretinopathy can include night blindness, color vision problems, sensitivity to bright light, seeing flashes, visual distortion, pain in the eyes, and loss of depth perception and / or peripheral vision. The present invention includes methods for treating or preventing birdshot chorioretinopathy or uveitis in a subject, the method comprising administering to the subject a therapeutically effective dose of an anti-IL2Rγ antigen-binding protein, for example, by intravitreal administration, such as intravitreal injection.

[0337] The present invention also provides a method for treating or preventing any autoimmune disease or disorder by inhibiting IL2Rγ. Blocking the signal transduction of one or more cytokines in the γc family may benefit patients suffering from autoimmunity due to the inhibitory effects on the secretion of inflammatory cytokines and the production of autoantibodies. Multiple sclerosis (MS) is a disease of the brain and spinal cord (central nervous system (CNS)) in which the immune system attacks the myelin sheaths of nerve fibers and causes communication problems between the brain and the rest of the body. Eventually, the disease can cause the nerves themselves to deteriorate or become permanently damaged. Rheumatoid arthritis (RA) is an autoimmune disease in which the body's immune system attacks the joints. This produces inflammation, which causes the tissue (synovium) lining the inside of the joints to thicken, resulting in swelling and pain within and around the joints. Psoriasis is an autoimmune disease that primarily affects the skin. Inflammation can also affect the joints, vascular system, and eyes of psoriasis patients. Type 1 diabetes is an autoimmune disease in which the immune system attacks and destroys the insulin-producing β cells in the pancreas. The pancreas then produces little or no insulin. Systemic lupus erythematosus (SLE) is a systemic autoimmune disease that occurs when the body's immune system attacks its own tissues and organs. Inflammation caused by lupus can affect many different body systems - including the joints, skin, kidneys, blood cells, brain, heart, and lungs. Myasthenia gravis is an autoimmune disease in which antibodies block the receptors for acetylcholine at the neuromuscular junction, which prevents muscle contraction. In most individuals with myasthenia gravis, this is caused by antibodies against the acetylcholine receptor itself. However, antibodies against other proteins, such as the MuSK (muscle-specific kinase) protein, can also cause impaired transmission at the neuromuscular junction. The present invention includes a method for treating or preventing an autoimmune disease or disorder (e.g., multiple sclerosis or any other central nervous system inflammation, rheumatoid arthritis, psoriasis, type I diabetes, systemic lupus erythematosus, and / or myasthenia gravis) in a subject, the method comprising administering to the subject a therapeutically effective dose of an anti-IL2Rγ antigen-binding protein.

[0338] The effective dose or therapeutically effective dose of an anti-IL2Rγ antigen-binding protein, such as an antibody or antigen-binding fragment, for treating or preventing an IL2Rγ-mediated disease or disorder refers to the amount of the antigen-binding protein sufficient to alleviate one or more signs and / or symptoms of the disease or disorder in the subject being treated, whether by inducing regression or elimination of these signs and / or symptoms or by inhibiting the progression of these signs and / or symptoms. In one embodiment of the invention, the effective dose or therapeutically effective dose of the anti-IL2Rγ antigen-binding protein is about 0.05 - 50 mg / kg body weight. The amount of the dose can vary depending on the age and size of the subject to be administered, the target disease, condition, route of administration, etc. In certain embodiments, a second or multiple subsequent doses of the antigen-binding protein can be administered after the initial dose, and the amount of the subsequent dose can be approximately equal to or less than or greater than the initial dose, where the subsequent doses are separated by at least 1 day to 3 days, at least one week, at least 2 weeks, at least 3 weeks, at least 4 weeks, at least 5 weeks, at least 6 weeks, at least 7 weeks, at least 8 weeks, at least 9 weeks, at least 10 weeks, at least 12 weeks, or at least 14 weeks.

[0339] As used herein, the term "subject" refers to a mammal (e.g., rat, mouse, cat, dog, cow, sheep, horse, goat, rabbit), preferably a human, who, for example, needs to prevent and / or treat an IL2Rγ-mediated disease. The subject can have an IL2Rγ-mediated disease or be prone to such a disease.

[0340] "Preventing" an IL2Rγ-mediated disease or disorder when it relates to the use of the anti-IL2Rγ antigen-binding protein of the present invention means administering to the subject before the disease or disorder manifests in the subject to stop such manifestation from occurring.

[0341] Combinations and pharmaceutical formulations

[0342] The present invention provides a composition comprising an anti-IL2Rγ antigen-binding protein in combination with one or more components; as well as methods of using the same and methods of preparing such a composition. A pharmaceutical formulation comprising an anti-IL2Rγ antigen-binding protein and a pharmaceutically acceptable carrier or excipient is part of the present invention.

[0343] For the preparation of an anti-IL2Rγ antigen-binding protein, such as an antibody and its antigen-binding fragments (e.g.,

[0344] A pharmaceutical preparation of (H4H12857P; H4H12858P; H4H12859P; H4H12863P; H4H12874P; H4H12871P; H4H12884P; H4H12886P; H4H12889P; H4H12890P; H4H12899P; H4H12900P; H4H12908P; H4H12913P2; H4H12922P2: H4H12924P2: H4H12926P2; H4H12927P2; H4H12934P2; H4H13538P; H4H13541P; H4H13544P2; or H4H13545P2) comprises mixing an antigen-binding protein with a pharmaceutically acceptable carrier or excipient. See, for example, Remington's Pharmaceutical Sciences and U.S. Pharmacopeia: National Formulary, Mack Publishing Company, Easton, Pa. (1984), Hardman, et al. (2001) Goodman and Gilman's The Pharmacological Basis of Therapeutics, McGraw-Hill, New York, N.Y.; Gennaro (2000) Remington: The Science and Practice of Pharmacy, Lippincott, Williams, and Wilkins, New York, N.Y.; Avis, et al. (eds.) (1993) Pharmaceutical Dosage Forms: Parenteral Medications, Marcel Dekker, NY; Lieberman, et al. (eds.) (1990) Pharmaceutical Dosage Forms: Tablets, Marcel Dekker, NY; Lieberman, et al. (eds.) (1990) Pharmaceutical Dosage Forms: Disperse Systems, Marcel Dekker, NY; Weiner and Kotkoskie (2000) Excipient Toxicity and Safety, Marcel Dekker, Inc., New York, N.Y. In one embodiment of the invention, the pharmaceutical preparation is sterile. Such compositions are part of the invention.

[0345] The pharmaceutical preparation of the present invention comprises an anti-IL2Rγ antigen-binding protein and a pharmaceutically acceptable carrier, and the carrier includes, for example, water, buffer, preservative and / or detergent.

[0346] The scope of the present invention includes dried, for example, lyophilized compositions comprising an anti-IL2Rγ antigen-binding protein, such as an antibody or an antigen-binding fragment thereof (such as H4H12857P; H4H12858P; H4H12859P; H4H12863P; H4H12874P; H4H12871P; H4H12884P; H4H12886P; H4H12889P; H4H12890P; H4H12899P; H4H12900P; H4H12908P; H4H12913P2; H4H12922P2; H4H12924P2; H4H12926P2; H4H12927P2; H4H12934P2; H4H13538P; H4H13541P; H4H13544P2; or H4H13545P2), or a pharmaceutical preparation thereof that comprises a pharmaceutically acceptable carrier but is substantially free of water.

[0347] In another embodiment of the present invention, according to Physicians’ Desk Reference 2003 (Thomson Healthcare; 57th edition (Nov. 1, 2002)), another therapeutic agent administered to a subject in combination with the anti-IL2Rγ antigen-binding protein, such as an antibody or an antigen-binding fragment thereof (such as,

[0348] H4H12857P; H4H12858P; H4H12859P; H4H12863P; H4H12874P; H4H12871P; H4H12884P; H4H12886P; H4H12889P; H4H12890P; H4H12899P; H4H12900P; H4H12908P; H4H12913P2; H4H12922P2; H4H12924P2; H4H12926P2; H4H12927P2; H4H12934P2; H4H13538P; H4H13541P; H4H13544P2; or H4H13545P2) is administered to the subject.

[0349] The mode of administration of the anti-IL2Rγ antigen-binding protein or its composition can be changed. The administration routes include parenteral, non-parenteral, oral, rectal, transmucosal, intestinal, parenteral, intramuscular, subcutaneous, intradermal, intramedullary, intrathecal, direct intraventricular, intravenous, intraperitoneal, intranasal, intraocular, inhalation, insufflation, topical, dermal, intraocular, intravitreal, transdermal or intraarterial.

[0350] The present invention provides methods for administering an anti-IL2Rγ antigen-binding protein, such as an antibody or an antigen-binding fragment thereof (e.g., H4H12857P; H4H12858P; H4H12859P; H4H12863P; H4H12874P; H4H12871P; H4H12884P; H4H12886P; H4H12889P; H4H12890P; H4H12899P; H4H12900P; H4H12908P; H4H12913P2; H4H12922P2; H4H12924P2; H4H12926P2; H4H12927P2; H4H12934P2; H4H13538P; H4H13541P; H4H13544P2; or H4H13545P2) to a subject, the method comprising introducing the protein or a pharmaceutical formulation thereof into the subject. For example, in one embodiment of the invention, the method comprises piercing, e.g., with a needle of a syringe, the body of the subject and injecting the antigen-binding protein or a pharmaceutical formulation thereof into the subject, e.g., into the eye, vein, artery, muscle tissue, or subcutaneously of the subject.

[0351] The present invention provides a container (e.g., a plastic bottle or a glass bottle, e.g., having a cap or a chromatography column, a hollow needle, or a syringe barrel) that contains any anti-IL2Rγ antigen-binding protein, such as an antibody or an antigen-binding fragment thereof (e.g., H4H12857P; H4H12858P; H4H12859P; H4H12863P; H4H12874P; H4H12871P; H4H12884P; H4H12886P; H4H12889P; H4H12890P; H4H12899P; H4H12900P; H4H12908P; H4H12913P2; H4H12922P2; H4H12924P2; H4H12926P2; H4H12927P2; H4H12934P2; H4H13538P; H4H13541P; H4H13544P2; or H4H13545P2), or a pharmaceutical formulation thereof that contains a pharmaceutically acceptable carrier.

[0352] The present invention includes a combination comprising an anti-IL2Rγ antigen-binding protein of the present invention, such as an antibody or an antigen-binding fragment thereof (e.g., H4H12857P; H4H12858P; H4H12859P; H4H12863P; H4H12874P; H4H12871P; H4H12884P; H4H12886P; H4H12889P; H4H12890P; H4H12899P; H4H12900P; H4H12908P; H4H12913P2; H4H12922P2; H4H12924P2; H4H12926P2; H4H12927P2; H4H12934P2; H4H13538P; H4H13541P; H4H13544P2; or H4H13545P2), in combination with one or more additional therapeutic agents. The anti-IL2Rγ antigen-binding protein and the additional therapeutic agent(s) can be in a single composition or in separate compositions. For example, in one embodiment of the present invention, the additional therapeutic agent is an immunosuppressive drug. In one embodiment of the present invention, the additional therapeutic agent is an anti-TNFα antibody or binding protein (e.g., infliximab, adalimumab, etanercept, or golimumab), tacrolimus, cyclosporine, corticosteroid, prednisolone, methylprednisolone, antithymocyte globulin, alemtuzumab, daclizumab, extracorporeal photopheresis, mycophenolate mofetil, sirolimus, pentostatin, mesenchymal stem cells, ibritumomab, denileukin, a multispecific (e.g., bispecific) antibody or antigen-binding fragment thereof that binds BCMA (B cell maturation antigen) and CD3, and / or basiliximab.

[0353] A method for performing such treatment or prophylaxis in a subject in need of treatment or prophylaxis of an IL2Rγ-mediated disease by co-administering an anti-IL2Rγ antigen-binding protein (e.g., H4H12857P; H4H12858P; H4H12859P; H4H12863P; H4H12874P; H4H12871P; H4H12884P; H4H12886P; H4H12889P; H4H12890P; H4H12899P; H4H12900P; H4H12908P; H4H12913P2; H4H12922P2; H4H12924P2; H4H12926P2; H4H12927P2; H4H12934P2; H4H13538P; H4H13541P; H4H13544P2; or H4H13545P2) with an additional therapeutic agent is part of the present invention.

[0354] The term "in combination with" means that components: the anti-IL2Rγ antigen-binding protein of the present invention (e.g., an antibody or an antigen-binding fragment thereof) and another agent (e.g., methotrexate) can be formulated into a single composition, e.g., for simultaneous delivery, or separately into two or more compositions (e.g., a kit containing each ingredient). Components administered in combination with each other can be administered to a subject at a different time than when administering the other component; for example, each administration can be given non-simultaneously at intervals (e.g., separately or sequentially) over a given period of time. The separate components administered in combination with each other can also be administered sequentially but substantially simultaneously during the same administration period. In addition, the separate components administered in combination with each other can be administered to a subject by the same or different routes.

[0355] The disclosure herein includes the following embodiments:

[0356] 1. An isolated antigen-binding protein that specifically binds to IL2Rγ or an antigenic fragment thereof, characterized by one or more of the following:

[0357] · having a K that binds to the extracellular domain of human IL2Rγ at about 2.75×10 -9 M to about 3.36×10 -7 M, about 2.45×10 -9 M to about 1.20×10 -8 M, or less than about 1.20×10 -8 M at 25°C; D

[0358] · having a K that binds to the extracellular domain of human IL2Rγ at about 6.42×10 -9 M to about 3.53×10 -7 M, about 1.86×10 -11 M to about 3.00×10 -8 M, less than about 3.00×10 -8 M, or less than about 3.53×10 -7 M at 37°C; D

[0359]

[0360] · having a K that binds to the extracellular domain of cynomolgus monkey IL-2Rγ at about 3.18×10 -9 M to about 2.38×10 -7 M at 25°C; D

[0360] · having a K that binds to the extracellular domain of cynomolgus monkey IL-2Rγ at about 8.29×10 -9 M to about 3.20×10 -7 M at 37°C; or having a K of less than about 3.20×10 D M that binds to the extracellular domain of cynomolgus monkey IL-2Rγ; -7 DD Combined with;

[0361] · At 25 °C, with a K of approximately 1.84×10 -8 M, 3.76×10 -9 M, 1.08×10 -7 M, 2.17×10 -8 M, 6.02×10 - 9 M or 7.93×10 -8 M binds to the extracellular domain of murine IL2Rγ; or there is no detectable binding; D

[0362] · At 37 °C, with a K of approximately 5.59×10 -8 M, 6.11×10 -9 M, 3.87×10 -7 M, 5.16×10 -8 M, 8.70×10 - 9 M or 2.15×10 -7 M binds to the extracellular domain of murine IL2Rγ; or there is no detectable binding; D

[0363] · At 25 °C, with a K ranging from approximately 3.32×10 -9 M to approximately 1.97×10 -7 M binds to human IL2Rγ domain 1; or there is no detectable binding; D

[0364] · At 37 °C, with a K ranging from approximately 4.13×10 -9 M to approximately 2.25×10 -7 M binds to human IL2Rγ domain 1; or there is no detectable binding; D

[0365] · At 25 °C, with a K ranging from approximately 2.91×10 -7 M to approximately 5.35×10 -10 binds to human IL2Rγ domain 2; or there is no detectable binding; D

[0366] · At 37 °C, with a K of approximately 1.14×10 -8 or approximately 1.27×10 -8 binds to human IL2Rγ domain 2; or there is no detectable binding; D

[0367] · There is no detectable binding to murine or rat IL2Rγ;

[0368] ​​​​​​· Block STAT phosphorylation in T cells induced by IL-2, IL-4, IL7, IL-15, and / or IL-21;

[0369] · Block STAT phosphorylation in mast cells induced by IL-9;

[0370] · Reduce the number of such cells and / or human cytokines in the blood or serum of immunodeficient mice injected with human peripheral blood mononuclear cells (PBMCs);

[0371] · Protect mice from weight loss and / or death due to GvHD in a GvHD mouse model;

[0372] · Block the binding of a heteromeric receptor containing IL2Rγ complexed with a cytokine-specific receptor subunit to IL-2, IL-4, IL-7, IL-9, IL-15, and / or IL-21;

[0373] · Inhibit IL2Rγ intracellular signaling through the JAK-STAT pathway induced by IL2, IL4, IL7, IL9, ILl5, and / or IL21;

[0374] · Specifically bind to the same epitope on IL2Rγ as a reference antibody or its antigen-binding fragment, wherein the reference antibody or its antigen-binding fragment is H4H12857P; H4H12858P; H4H12859P; H4H12863P; H4H12874P; H4H12871P; H4H12884P; H4H12886P; H4H12889P; H4H12890P; H4H12899P; H4H12900P; H4H12908P; H4H12913P2; H4H12922P2; H4H12924P2; H4H12926P2; H4H12927P2; H4H12934P2; H4H13538P; H4H13541P; H4H13544P2; or H4H13545P2;

[0375] · Compete with a reference antibody or its antigen-binding fragment for binding to an IL2Rγ polypeptide or its antigenic fragment, wherein the reference antibody or its antigen-binding fragment is

[0376] H4H12857P; H4H12858P; H4H12859P; H4H12863P; H4H12874P: H4H12871P; H4H12884P: H4H12888P; H4H12889P; H4H12890P; H4H12899P; H4.H12900P; H4H12908P; H4H12913P2; H4H12922P2; H4H12924P2; H4H12926P2; H4H12927P2; H4H12934.P2: H4H13538P; H4H1354.1P; H4H13544P2: or H4H1354.5P2; and / or

[0377] · Reduce CD45+ cells, B cells, T cells, and / or NK cells in blood or serum.

[0378] 2. The antigen-binding protein according to embodiment 1, which specifically binds to IL2Rγ or an antigenic fragment thereof, and the antigen-binding protein is an antibody or an antigen-binding fragment thereof.

[0379] 3. The antigen-binding protein according to embodiment 2, which specifically binds to IL2Rγ or an antigenic fragment thereof, and the antigen-binding protein is an antibody.

[0380] 4. An isolated antigen-binding protein that specifically binds to IL2Rγ or an antigenic fragment thereof, comprising:

[0381] (a) An immunoglobulin heavy chain or its variable region, which contains CDR-H1, CDR-H2, and CDR-H3 of an immunoglobulin heavy chain or its variable region containing the amino acid sequence shown in SEQ ID NO: 2, 18, 22, 38, 42, 58, 62, 77, 81, 97, 101, 115, 119, 134, 138, 152, 156, 170, 174, 186, 190, 198, 200, 208, 210, 216, 218, 234, 238, 254, 258, 272, 276, 284, 286, 294, 296, 311, 315, 331, 335, 343, 345, 357, 361, or 376 or a variant thereof; and / or

[0382] (b) An immunoglobulin light chain or variable region thereof, comprising CDR-L1, CDR-L2, and CDR-L3 of an immunoglobulin light chain or variable region thereof containing an amino acid sequence shown in SEQ ID NO: 10, 20, 30, 40, 50, 60, 70, 79, 89, 99, 109, 117, 127, 136, 146, 154, 164, 172, 182, 188, 226, 236, 246, 256, 266, 274, 304, 313, 323, 333, 353, 359, 368, or 378, or a variant thereof.

[0383] 5. The antigen-binding protein according to any one of embodiments 1 to 4, which specifically binds to IL2Rγ or an antigenic fragment thereof, wherein the antigen-binding protein comprises:

[0384] (a) An immunoglobulin heavy chain or variable region thereof, comprising an amino acid sequence having at least 90% amino acid sequence identity with the amino acid sequence shown in SEQ ID NO: 2, 18, 22, 38, 42, 58, 62, 77, 81, 97, 101, 115, 119, 134, 138, 152, 156, 170, 174, 186, 190, 198, 200, 208, 210, 216, 218, 234, 238, 254, 258, 272, 276, 284, 286, 294, 296, 311, 315, 331, 335, 343, 345, 357, 361, or 376; and / or

[0385] (b) An immunoglobulin light chain or variable region thereof, comprising an amino acid sequence having at least 90% amino acid sequence identity with the amino acid sequence shown in SEQ ID NO: 10, 20, 30, 40, 50, 60, 70, 79, 89, 99, 109, 117, 127, 136, 146, 154, 164, 172, 182, 188, 226, 236, 246, 256, 266, 274, 304, 313, 323, 333, 353, 359, 368, or 378.

[0386] 6. The antigen-binding protein according to any one of embodiments 1 to 5, which specifically binds to IL2Rγ or an antigenic fragment thereof, wherein the antigen-binding protein comprises:

[0387] (a) An immunoglobulin heavy chain or variable region thereof, comprising CDR-H1, CDR-H2 and CDR-H3 of an immunoglobulin heavy chain or variable region thereof that contains the amino acid sequence shown in SEQ ID NO: 2, 18, 22, 38, 42, 58, 62, 77, 81, 97, 101, 115, 119, 134, 138, 152, 156, 170, 174, 186, 190, 198, 200, 208, 210, 216, 218, 234, 238, 254, 258, 272, 276, 284, 286, 294, 296, 311, 315, 331, 335, 343, 345, 357, 361 or 376 and an amino acid sequence having at least 90% amino acid sequence identity to the amino acid sequence shown in SEQ ID NO: 2, 18, 22, 38, 42, 58, 62, 77, 81, 97, 101, 115, 119, 134, 138, 152, 156, 170, 174, 186, 190, 198, 200, 208, 210, 216, 218, 234, 238, 254, 258, 272, 276, 284, 286, 294, 296, 311, 315, 331, 335, 343, 345, 357, 361 or 376; and / or

[0388] (b) An immunoglobulin light chain or variable region thereof, comprising CDR-L1, CDR-L2 and CDR-L3 of an immunoglobulin light chain or variable region thereof that contains the amino acid sequence shown in SEQ ID NO: 10, 20, 30, 40, 50, 60, 70, 79, 89, 99, 109, 117, 127, 136, 146, 154, 164, 172, 182, 188, 226, 236, 246, 256, 266, 274, 304, 313, 323, 333, 353, 359, 368 or 378 and an amino acid sequence having at least 90% amino acid sequence identity to the amino acid sequence shown in SEQ ID NO: 10, 20, 30, 40, 50, 60, 70, 79, 89, 99, 109, 117, 127, 136, 146, 154, 164, 172, 182, 188, 226, 236, 246, 256, 266, 274, 304, 313, 323, 333, 353, 359, 368 or 378.

[0389] 7. The antigen-binding protein according to any one of embodiments 1 to 6, which specifically binds to IL2Rγ or an antigenic fragment thereof, wherein the antigen-binding protein comprises:

[0390] (i)

[0391] CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 4;

[0392] CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 6; and

[0393] CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 8;

[0394] and / or

[0395] CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 24;

[0396] CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 26; and

[0397] CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 28;

[0398] and / or

[0399] CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 44;

[0400] CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 46; and

[0401] CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 48;

[0402] and / or

[0403] CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 64;

[0404] CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 66; and

[0405] CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 68;

[0406] and / or

[0407] CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 83;

[0408] CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 85; and

[0409] CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 87;

[0410] and / or

[0411] CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 103;

[0412] CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 105; and

[0413] CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 107;

[0414] and / or

[0415] CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 121;

[0416] CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 123; and

[0417] CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 125;

[0418] and / or

[0419] CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 140;

[0420] CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 142; and

[0421] CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 144;

[0422] and / or

[0423] CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 158;

[0424] CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 160; and

[0425] CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 162;

[0426] and / or

[0427] CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 176;

[0428] CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 178; and

[0429] CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 180;

[0430] and / or

[0431] CDR-H1 comprising the amino acid sequence shown in SEQ ID NO: 192;

[0432] CDR-H2 comprising the amino acid sequence shown in SEQ ID NO: 194; and

[0433] CDR-H3 comprising the amino acid sequence shown in SEQ ID NO: 196;

[0434] and / or

[0435] CDR-H1 comprising the amino acid sequence shown in SEQ ID NO: 202;

[0436] CDR-H2 comprising the amino acid sequence shown in SEQ ID NO: 204; and

[0437] CDR-H3 comprising the amino acid sequence shown in SEQ ID NO: 206;

[0438] and / or

[0439] CDR-H1 comprising the amino acid sequence shown in SEQ ID NO: 176;

[0440] CDR-H2 comprising the amino acid sequence shown in SEQ ID NO: 212; and

[0441] CDR-H3 comprising the amino acid sequence shown in SEQ ID NO: 214;

[0442] and / or

[0443] CDR-H1 comprising the amino acid sequence shown in SEQ ID NO: 220;

[0444] CDR-H2 comprising the amino acid sequence shown in SEQ ID NO: 222; and

[0445] CDR-H3 comprising the amino acid sequence shown in SEQ ID NO: 224;

[0446] and / or

[0447] CDR-H1 comprising the amino acid sequence shown in SEQ ID NO: 240;

[0448] CDR-H2 comprising the amino acid sequence shown in SEQ ID NO: 242; and

[0449] CDR-H3 comprising the amino acid sequence shown in SEQ ID NO: 244;

[0450] and / or

[0451] A CDR-H1 comprising the amino acid sequence set forth in SEQ ID NO: 260;

[0452] A CDR-H2 comprising the amino acid sequence set forth in SEQ ID NO: 262; and

[0453] A CDR-H3 comprising the amino acid sequence set forth in SEQ ID NO: 264;

[0454] and / or

[0455] A CDR-H1 comprising the amino acid sequence set forth in SEQ ID NO: 278;

[0456] A CDR-H2 comprising the amino acid sequence set forth in SEQ ID NO: 280; and

[0457] A CDR-H3 comprising the amino acid sequence set forth in SEQ ID NO: 282;

[0458] and / or

[0459] A CDR-H1 comprising the amino acid sequence set forth in SEQ ID NO: 288;

[0460] A CDR-H2 comprising the amino acid sequence set forth in SEQ ID NO: 290; and

[0461] A CDR-H3 comprising the amino acid sequence set forth in SEQ ID NO: 292;

[0462] and / or

[0463] A CDR-H1 comprising the amino acid sequence set forth in SEQ ID NO: 298;

[0464] A CDR-H2 comprising the amino acid sequence set forth in SEQ ID NO: 300; and

[0465] A CDR-H3 comprising the amino acid sequence set forth in SEQ ID NO: 302;

[0466] and / or

[0467] A CDR-H1 comprising the amino acid sequence set forth in SEQ ID NO: 317;

[0468] A CDR-H2 comprising the amino acid sequence set forth in SEQ ID NO: 319; and

[0469] A CDR-H3 comprising the amino acid sequence set forth in SEQ ID NO: 321;

[0470] and / or

[0471] CDR-H1 comprising the amino acid sequence set forth in SEQ ID NO: 337;

[0472] CDR-H2 comprising the amino acid sequence set forth in SEQ ID NO: 339; and

[0473] CDR-H3 comprising the amino acid sequence set forth in SEQ ID NO: 341;

[0474] and / or

[0475] CDR-H1 comprising the amino acid sequence set forth in SEQ ID NO: 347;

[0476] CDR-H2 comprising the amino acid sequence set forth in SEQ ID NO: 349; and

[0477] CDR-H3 comprising the amino acid sequence set forth in SEQ ID NO: 351;

[0478] and / or

[0479] CDR-H1 comprising the amino acid sequence set forth in SEQ ID NO: 363;

[0480] CDR-H2 comprising the amino acid sequence set forth in SEQ ID NO: 66; and

[0481] CDR-H3 comprising the amino acid sequence set forth in SEQ ID NO: 366;

[0482] and / or

[0483] (ii)

[0484] CDR-L1 comprising the amino acid sequence set forth in SEQ ID NO: 12;

[0485] CDR-L2 comprising the amino acid sequence set forth in SEQ ID NO: 14; and

[0486] CDR-L3 comprising the amino acid sequence set forth in SEQ ID NO: 16;

[0487] and / or

[0488] CDR-L1 comprising the amino acid sequence set forth in SEQ ID NO: 32;

[0489] CDR-L2 comprising the amino acid sequence set forth in SEQ ID NO: 34; and

[0490] CDR-L3 comprising the amino acid sequence set forth in SEQ ID NO: 36;

[0491] and / or

[0492] a CDR-L1 comprising the amino acid sequence shown in SEQ ID NO: 52;

[0493] a CDR-L2 comprising the amino acid sequence shown in SEQ ID NO: 54; and

[0494] a CDR-L3 comprising the amino acid sequence shown in SEQ ID NO: 56;

[0495] and / or

[0496] a CDR-L1 comprising the amino acid sequence shown in SEQ ID NO: 72;

[0497] a CDR-L2 comprising the amino acid sequence shown in SEQ ID NO: 54; and

[0498] a CDR-L3 comprising the amino acid sequence shown in SEQ ID NO: 75;

[0499] and / or

[0500] a CDR-L1 comprising the amino acid sequence shown in SEQ ID NO: 91;

[0501] a CDR-L2 comprising the amino acid sequence shown in SEQ ID NO: 93; and

[0502] a CDR-L3 comprising the amino acid sequence shown in SEQ ID NO: 95;

[0503] and / or

[0504] a CDR-L1 comprising the amino acid sequence shown in SEQ ID NO: 111;

[0505] a CDR-L2 comprising the amino acid sequence shown in SEQ ID NO: 54; and

[0506] a CDR-L3 comprising the amino acid sequence shown in SEQ ID NO: 113;

[0507] and / or

[0508] a CDR-L1 comprising the amino acid sequence shown in SEQ ID NO: 129;

[0509] a CDR-L2 comprising the amino acid sequence shown in SEQ ID NO: 54; and

[0510] a CDR-L3 comprising the amino acid sequence shown in SEQ ID NO: 132;

[0511] and / or

[0512] a CDR-L1 comprising the amino acid sequence set forth in SEQ ID NO: 148;

[0513] a CDR-L2 comprising the amino acid sequence set forth in SEQ ID NO: 54; and

[0514] a CDR-L3 comprising the amino acid sequence set forth in SEQ ID NO: 150;

[0515] and / or

[0516] a CDR-L1 comprising the amino acid sequence set forth in SEQ ID NO: 166;

[0517] a CDR-L2 comprising the amino acid sequence set forth in SEQ ID NO: 14; and

[0518] a CDR-L3 comprising the amino acid sequence set forth in SEQ ID NO: 168;

[0519] and / or

[0520] a CDR-L1 comprising the amino acid sequence set forth in SEQ ID NO: 72;

[0521] a CDR-L2 comprising the amino acid sequence set forth in SEQ ID NO: 54; and

[0522] a CDR-L3 comprising the amino acid sequence set forth in SEQ ID NO: 184;

[0523] and / or

[0524] a CDR-L1 comprising the amino acid sequence set forth in SEQ ID NO: 228;

[0525] a CDR-L2 comprising the amino acid sequence set forth in SEQ ID NO: 230; and

[0526] a CDR-L3 comprising the amino acid sequence set forth in SEQ ID NO: 232;

[0527] and / or

[0528] a CDR-L1 comprising the amino acid sequence set forth in SEQ ID NO: 248;

[0529] a CDR-L2 comprising the amino acid sequence set forth in SEQ ID NO: 250; and

[0530] a CDR-L3 comprising the amino acid sequence set forth in SEQ ID NO: 252;

[0531] and / or

[0532] a CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 268;

[0533] a CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 54; and

[0534] a CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 270;

[0535] and / or

[0536] a CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 306;

[0537] a CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 230; and

[0538] a CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 309;

[0539] and / or

[0540] a CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 325;

[0541] a CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 327; and

[0542] a CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 329;

[0543] and / or

[0544] a CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 72;

[0545] a CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 54; and

[0546] a CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 355;

[0547] and / or

[0548] a CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 370;

[0549] a CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 372; and

[0550] a CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 374.

[0551] The antigen-binding protein according to any one of embodiments 1 to 7, which specifically binds to IL2Rγ or an antigenic fragment thereof, and the antigen-binding protein comprises one or more members selected from the following:

[0552] (i) A heavy chain variable region comprising:

[0553] CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 4;

[0554] CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 6; and

[0555] CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 8; and

[0556] A light chain variable region comprising:

[0557] CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 12;

[0558] CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 14; and

[0559] CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 16;

[0560] (ii) A heavy chain variable region comprising:

[0561] CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 24;

[0562] CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 26; and

[0563] CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 28; and

[0564] A light chain variable region comprising:

[0565] CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 32;

[0566] CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 34; and

[0567] CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 36;

[0568] (iii) A heavy chain variable region comprising:

[0569] CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 44;

[0570] CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 46; and

[0571] CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 48; and

[0572] A light chain variable region comprising:

[0573] CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 52;

[0574] CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 54; and

[0575] CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 56;

[0576] (iv) A heavy chain variable region comprising:

[0577] CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 64;

[0578] CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 66; and

[0579] CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 68; and

[0580] A light chain variable region comprising:

[0581] CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 72;

[0582] CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 54; and

[0583] CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 75;

[0584] (v) A heavy chain variable region comprising:

[0585] CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 83;

[0586] CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 85; and

[0587] CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 87; and

[0588] A light chain variable region, comprising:

[0589] CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 91;

[0590] CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 93; and

[0591] CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 95;

[0592] (vi) A heavy chain variable region, comprising:

[0593] CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 103;

[0594] CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 105; and

[0595] CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 107; and

[0596] A light chain variable region, comprising:

[0597] CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 111;

[0598] CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 54; and

[0599] CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 113;

[0600] (vi) A heavy chain variable region, comprising:

[0601] CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 121;

[0602] CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 123; and

[0603] CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 125; and

[0604] A light chain variable region, comprising:

[0605] CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 129;

[0606] CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 54; and

[0607] CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 132;

[0608] (vii) A heavy chain variable region comprising:

[0609] CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 140;

[0610] CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 142; and

[0611] CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 144; and

[0612] A light chain variable region comprising:

[0613] CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 148;

[0614] CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 54; and

[0615] CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 150;

[0616] (viii) A heavy chain variable region comprising:

[0617] CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 158;

[0618] CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 160; and

[0619] CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 162; and

[0620] A light chain variable region comprising:

[0621] CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 166;

[0622] CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 14; and

[0623] CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 168;

[0624] (ix) A heavy chain variable region comprising:

[0625] CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 176;

[0626] The CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 178; and

[0627] The CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 180; and

[0628] A light chain variable region comprising:

[0629] The CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 72;

[0630] The CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 54; and

[0631] The CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 184;

[0632] (x) A heavy chain variable region comprising:

[0633] The CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 192;

[0634] The CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 194; and

[0635] The CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 196; and

[0636] A light chain variable region comprising:

[0637] The CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 72;

[0638] The CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 54; and

[0639] The CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 184;

[0640] (xi) A heavy chain variable region comprising:

[0641] The CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 202;

[0642] The CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 204; and

[0643] The CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 206; and

[0644] A light chain variable region comprising:

[0645] CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 72;

[0646] CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 54; and

[0647] CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 184;

[0648] (xii) A heavy chain variable region comprising:

[0649] CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 176;

[0650] CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 212; and

[0651] CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 214; and

[0652] A light chain variable region comprising:

[0653] CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 72;

[0654] CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 54; and

[0655] CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 184;

[0656] (xiii) A heavy chain variable region comprising:

[0657] CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 220;

[0658] CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 222; and

[0659] CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 224; and

[0660] A light chain variable region comprising:

[0661] CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 228;

[0662] CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 230; and

[0663] CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 232;

[0664] (xiv) A heavy chain variable region comprising:

[0665] CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 240;

[0666] CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 242; and

[0667] CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 244; and

[0668] A light chain variable region comprising:

[0669] CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 248;

[0670] CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 250; and

[0671] CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 252;

[0672] (xv) A heavy chain variable region comprising:

[0673] CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 260;

[0674] CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 262; and

[0675] CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 264; and

[0676] A light chain variable region comprising:

[0677] CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 268;

[0678] CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 54; and

[0679] CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 270;

[0680] (xvi) A heavy chain variable region comprising:

[0681] CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 278;

[0682] CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 280; and

[0683] A CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 282; and

[0684] A light chain variable region comprising:

[0685] A CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 72;

[0686] A CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 54; and

[0687] A CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 184;

[0688] (xvii) A heavy chain variable region comprising:

[0689] A CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 288;

[0690] A CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 290; and

[0691] A CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 292; and

[0692] A light chain variable region comprising:

[0693] A CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 72;

[0694] A CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 54; and

[0695] A CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 184;

[0696] (xviii) A heavy chain variable region comprising:

[0697] A CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 298;

[0698] A CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 300; and

[0699] A CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 302; and

[0700] A light chain variable region comprising:

[0701] A CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 306;

[0702] A CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 230; and

[0703] A CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 309;

[0704] (xix) A heavy chain variable region comprising:

[0705] A CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 317;

[0706] A CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 319; and

[0707] A CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 321; and

[0708] A light chain variable region comprising:

[0709] A CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 325;

[0710] A CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 327; and

[0711] A CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 329;

[0712] (xx) A heavy chain variable region comprising:

[0713] A CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 337;

[0714] A CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 339; and

[0715] A CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 341; and

[0716] A light chain variable region comprising:

[0717] A CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 72;

[0718] A CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 54; and

[0719] A CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 184;

[0720] (xxi) A heavy chain variable region comprising:

[0721] CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 347;

[0722] CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 349; and

[0723] CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 351; and

[0724] A light chain variable region comprising:

[0725] CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 72;

[0726] CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 54; and

[0727] CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 355;

[0728] (xxii) A heavy chain variable region comprising:

[0729] CDR-H1 containing the amino acid sequence shown in SEQ ID NO: 363;

[0730] CDR-H2 containing the amino acid sequence shown in SEQ ID NO: 66; and

[0731] CDR-H3 containing the amino acid sequence shown in SEQ ID NO: 366; and

[0732] A light chain variable region comprising:

[0733] CDR-L1 containing the amino acid sequence shown in SEQ ID NO: 370;

[0734] CDR-L2 containing the amino acid sequence shown in SEQ ID NO: 372; and

[0735] CDR-L3 containing the amino acid sequence shown in SEQ ID NO: 374.

[0736] 9. An antigen-binding protein that specifically binds to IL2Rγ or an antigenic fragment thereof, which is an antibody or an antigen-binding fragment, and which comprises:

[0737] A heavy chain variable region containing the amino acid sequence shown in SEQ ID NO: 2 and a light chain variable region containing the amino acid sequence shown in SEQ ID NO: 10;

[0738] A heavy chain variable region containing the amino acid sequence shown in SEQ ID NO: 22 and a light chain variable region containing the amino acid sequence shown in SEQ ID NO: 30;

[0739] A heavy chain variable region containing the amino acid sequence shown in SEQ ID NO: 42 and a light chain variable region containing the amino acid sequence shown in SEQ ID NO: 50;

[0740] A heavy chain variable region containing the amino acid sequence shown in SEQ ID NO: 62 and a light chain variable region containing the amino acid sequence shown in SEQ ID NO: 70;

[0741] A heavy chain variable region containing the amino acid sequence shown in SEQ ID NO: 81 and a light chain variable region containing the amino acid sequence shown in SEQ ID NO: 89;

[0742] A heavy chain variable region containing the amino acid sequence shown in SEQ ID NO: 101 and a light chain variable region containing the amino acid sequence shown in SEQ ID NO: 109;

[0743] A heavy chain variable region containing the amino acid sequence shown in SEQ ID NO: 119 and a light chain variable region containing the amino acid sequence shown in SEQ ID NO: 127;

[0744] A heavy chain variable region containing the amino acid sequence shown in SEQ ID NO: 138 and a light chain variable region containing the amino acid sequence shown in SEQ ID NO: 146;

[0745] A heavy chain variable region containing the amino acid sequence shown in SEQ ID NO: 156 and a light chain variable region containing the amino acid sequence shown in SEQ ID NO: 164;

[0746] A heavy chain variable region containing the amino acid sequence shown in SEQ ID NO: 174 and a light chain variable region containing the amino acid sequence shown in SEQ ID NO: 182;

[0747] A heavy chain variable region containing the amino acid sequence shown in SEQ ID NO: 190 and a light chain variable region containing the amino acid sequence shown in SEQ ID NO: 182;

[0748] A heavy chain variable region containing the amino acid sequence shown in SEQ ID NO: 200 and a light chain variable region containing the amino acid sequence shown in SEQ ID NO: 182;

[0749] A heavy chain variable region containing the amino acid sequence shown in SEQ ID NO: 210 and a light chain variable region containing the amino acid sequence shown in SEQ ID NO: 182;

[0750] A heavy chain variable region containing the amino acid sequence shown in SEQ ID NO: 218 and a light chain variable region containing the amino acid sequence shown in SEQ ID NO: 226;

[0751] A heavy chain variable region containing the amino acid sequence shown in SEQ ID NO: 238 and a light chain variable region containing the amino acid sequence shown in SEQ ID NO: 246;

[0752] A heavy chain variable region containing the amino acid sequence shown in SEQ ID NO: 258 and a light chain variable region containing the amino acid sequence shown in SEQ ID NO: 266;

[0753] A heavy chain variable region containing the amino acid sequence shown in SEQ ID NO: 276 and a light chain variable region containing the amino acid sequence shown in SEQ ID NO: 182;

[0754] A heavy chain variable region containing the amino acid sequence shown in SEQ ID NO: 286 and a light chain variable region containing the amino acid sequence shown in SEQ ID NO: 182;

[0755] A heavy chain variable region containing the amino acid sequence shown in SEQ ID NO: 296 and a light chain variable region containing the amino acid sequence shown in SEQ ID NO: 304;

[0756] A heavy chain variable region containing the amino acid sequence shown in SEQ ID NO: 315 and a light chain variable region containing the amino acid sequence shown in SEQ ID NO: 323;

[0757] A heavy chain variable region containing the amino acid sequence shown in SEQ ID NO: 335 and a light chain variable region containing the amino acid sequence shown in SEQ ID NO: 182;

[0758] A heavy chain variable region containing the amino acid sequence shown in SEQ ID NO: 345 and a light chain variable region containing the amino acid sequence shown in SEQ ID NO: 353; and / or

[0759] A heavy chain variable region containing the amino acid sequence shown in SEQ ID NO: 361 and a light chain variable region containing the amino acid sequence shown in SEQ ID NO: 368.

[0760] 10. An antigen-binding protein that specifically binds to IL2Ry or an antigenic fragment thereof, which is an antibody or an antigen-binding fragment, and which comprises:

[0761] An immunoglobulin heavy chain containing the following amino acid sequence:

[0762] and

[0763] an immunoglobulin light chain having the following amino acid sequence:

[0764]

[0765] an immunoglobulin heavy chain having the following amino acid sequence:

[0766]

[0767] and

[0768] an immunoglobulin light chain having the following amino acid sequence:

[0769]

[0770] an immunoglobulin heavy chain having the following amino acid sequence:

[0771] and

[0772] an immunoglobulin light chain having the following amino acid sequence:

[0773]

[0774] an immunoglobulin heavy chain having the following amino acid sequence:

[0775] and

[0776] an immunoglobulin light chain having the following amino acid sequence:

[0777]

[0778] an immunoglobulin heavy chain having the following amino acid sequence:

[0779] and

[0780] an immunoglobulin light chain having the following amino acid sequence:

[0781]

[0782] an immunoglobulin heavy chain having the following amino acid sequence:

[0783] and

[0784] an immunoglobulin light chain having the following amino acid sequence:

[0785]

[0786] Immunoglobulin heavy chains containing the following amino acid sequences:

[0787]

[0788] and

[0789] Immunoglobulin light chains containing the following amino acid sequences:

[0790]

[0791] Immunoglobulin heavy chains containing the following amino acid sequences:

[0792] and

[0793] Immunoglobulin light chains containing the following amino acid sequences:

[0794]

[0795] Immunoglobulin heavy chains containing the following amino acid sequences:

[0796] and

[0797] Immunoglobulin light chains containing the following amino acid sequences:

[0798]

[0799] Immunoglobulin heavy chains containing the following amino acid sequences:

[0800]

[0801] and

[0802] Immunoglobulin light chains containing the following amino acid sequences:

[0803]

[0804] Immunoglobulin heavy chains containing the following amino acid sequences:

[0805] and

[0806] Immunoglobulin light chains containing the following amino acid sequences:

[0807] and

[0808] Immunoglobulin heavy chains containing the following amino acid sequences:

[0809] and

[0810] An immunoglobulin light chain having the following amino acid sequence:

[0811]

[0812] An immunoglobulin heavy chain having the following amino acid sequence:

[0813]

[0814] And

[0815] An immunoglobulin light chain having the following amino acid sequence:

[0816]

[0817] An immunoglobulin heavy chain having the following amino acid sequence:

[0818] And

[0819] An immunoglobulin light chain having the following amino acid sequence:

[0820]

[0821] An immunoglobulin heavy chain having the following amino acid sequence:

[0822] And

[0823] An immunoglobulin light chain having the following amino acid sequence:

[0824]

[0825] An immunoglobulin heavy chain having the following amino acid sequence:

[0826] And

[0827] An immunoglobulin light chain having the following amino acid sequence:

[0828]

[0829] An immunoglobulin heavy chain having the following amino acid sequence:

[0830] And

[0831] An immunoglobulin light chain having the following amino acid sequence:

[0832]

[0833] An immunoglobulin heavy chain having the following amino acid sequence:

[0834]

[0835] and

[0836] an immunoglobulin light chain having the following amino acid sequence:

[0837]

[0838] an immunoglobulin heavy chain having the following amino acid sequence:

[0839] and

[0840] an immunoglobulin light chain having the following amino acid sequence:

[0841]

[0842] an immunoglobulin heavy chain having the following amino acid sequence:

[0843] and

[0844] an immunoglobulin light chain having the following amino acid sequence:

[0845]

[0846] an immunoglobulin heavy chain having the following amino acid sequence:

[0847] and

[0848] an immunoglobulin light chain having the following amino acid sequence:

[0849]

[0850] an immunoglobulin heavy chain having the following amino acid sequence:

[0851] and

[0852] an immunoglobulin light chain having the following amino acid sequence:

[0853] and / or

[0854] an immunoglobulin heavy chain having the following amino acid sequence:

[0855] and

[0856] an immunoglobulin light chain having the following amino acid sequence:

[0857]

[0858] 11. The antigen-binding protein according to any one of embodiments 1 to 10, which is multispecific.

[0859] 12. A complex comprising the antigen-binding protein according to any one of embodiments 1 to 11 that binds to an IL2Rγ polypeptide or an antigenic fragment thereof.

[0860] 13. A method for preparing the antigen-binding protein according to any one of embodiments 1 to 11 or an immunoglobulin chain thereof, comprising:

[0861] (a) introducing one or more polynucleotides encoding the immunoglobulin chains of the antigen-binding protein into a host cell;

[0862] (b) culturing the host cell under conditions conducive to the expression of the polynucleotide; and

[0863] (c) optionally, isolating the antigen-binding protein or immunoglobulin chain from the host cell and / or the culture medium in which the host cell was cultured.

[0864] 14. The method according to embodiment 13, wherein the host cell is a Chinese hamster ovary cell.

[0865] 15. An antigen-binding protein or immunoglobulin chain that is a product of the method according to any one of embodiments 13 to 14.

[0866] 16. A polypeptide comprising:

[0867] (a) CDR-H1, CDR-H2, and CDR-H3 of an immunoglobulin heavy chain or its variable region containing the amino acid sequence shown in SEQ ID NO: 2, 18, 22, 38, 42, 58, 62, 77, 81, 97, 101, 115, 119, 134, 138, 152, 156, 170, 174, 186, 190, 198, 200, 208, 210, 216, 218, 234, 238, 254, 258, 272, 276, 284, 286, 294, 296, 311, 315, 331, 335, 343, 345, 357, 361, and / or 376 or a variant thereof; and / or

[0868] (b) CDR-L1, CDR-L2, and CDR-L3 of an immunoglobulin light chain or its variable region containing the amino acid sequence shown in SEQ ID NO: 10, 20, 30, 40, 50, 60, 70, 79, 89, 99, 109, 117, 127, 136, 146, 154, 164, 172, 182, 188, 226, 236, 246, 256, 266, 274, 304, 313, 323, 333, 353, 359, 368, and / or 378, or a variant thereof;

[0869] Alternatively,

[0870] (c) an amino acid sequence shown in a member selected from SEQ ID NO: 1 to 385, or a variant thereof.

[0871] 17. A polynucleotide encoding one or more polypeptides according to any one of embodiments 16.

[0872] 18. A vector comprising the polynucleotide according to embodiment 17.

[0873] 19. A host cell comprising an antigen-binding protein, an immunoglobulin chain, a polypeptide, a polynucleotide, and / or a vector according to any one of embodiments 1 to 11 and 15 to 18.

[0874] 20. A composition or a kit comprising one or more antigen-binding proteins according to any one of embodiments 1 to 11 and 15, optionally in combination with an additional therapeutic agent.

[0875] 21. A pharmaceutical formulation comprising an antigen-binding protein according to any one of embodiments 1 to 11 and 15, a pharmaceutically acceptable carrier or excipient, and optionally, an additional therapeutic agent.

[0876] 22. The composition, kit, or formulation according to any one of embodiments 20 to 21, in combination with an additional therapeutic agent, wherein the additional therapeutic agent is an anti-inflammatory agent.

[0877] 23. The composition, kit, or formulation according to any one of embodiments 20 to 21, in combination with an additional therapeutic agent, wherein the additional therapeutic agent is one or more members selected from the following: an anti-TNFα antibody or binding protein, infliximab, adalimumab, etanercept, golimumab, corticosteroid, prednisolone, methylprednisolone, antithymocyte globulin, alemtuzumab, daclizumab, tacrolimus, cyclosporine, extracorporeal photopheresis, mycophenolate mofetil, sirolimus, pentostatin, mesenchymal stem cells, ibritumomab, denileukin, and basiliximab.

[0878] 24. A container or injection device comprising the antigen-binding protein or composition or formulation according to any one of embodiments 1 to 11, 15, 20, 21, 22 or 23.

[0879] 25. A method for administering to a subject the antigen-binding protein, composition or formulation according to any one of embodiments 1 to 11, 15, 20, 21, 22 or 23, comprising injecting the antigen-binding protein, composition or formulation into the subject.

[0880] 26. A method for treating or preventing an IL2Rγ-mediated disease or disorder in a subject in need thereof, comprising administering an effective amount of the antigen-binding protein or composition or formulation according to any one of embodiments 1 to 11, 15, 20, 21, 22 or 23.

[0881] 27. The method according to embodiment 26, wherein the IL2Rγ-mediated disease or disorder is graft-versus-host disease, organ transplant rejection, b-islet cell transplant rejection, skin transplant rejection, heart transplant rejection, lung transplant rejection, kidney transplant rejection, liver transplant rejection, birdshot chorioretinopathy, multiple sclerosis, uveitis, autoimmune disease, type I diabetes, multiple sclerosis, rheumatoid arthritis, systemic lupus erythematosus, myasthenia gravis, aplastic anemia, atopic dermatitis, asthma, mast cell activation disorder, mast cell activation syndrome (MCAS), systemic mastocytosis (SM) and / or mast cell leukemia (MCL).

[0882] 28. The method according to any one of embodiments 25 to 27, wherein the antigen-binding protein is administered to the subject by subcutaneous, intravenous or intramuscular injection.

[0883] 29. A method for performing in a subject:

[0884] - blocking STAT phosphorylation in peripheral blood mononuclear cells induced by cytokines;

[0885] - blocking STAT phosphorylation in mast cells induced by cytokines;

[0886] - reducing the serum levels of interferon-γ, tumor necrosis factor-α, IL-6, IL-8, IL-10 and / or mKC / GRO;

[0887] - blocking JAK-STAT-mediated intracellular signal transduction induced by cytokines in the ILRγ family; and / or

[0888] - Reduce the serum levels of CD45+ immune cells, NK cells, T cells, and / or B cells; the method comprises administering to the subject an effective amount of the antigen-binding protein or composition or formulation according to any one of embodiments 1 to 11, 15, 20, 21, 22, or 23.

[0889] 30. The method according to embodiment 29, wherein the subject has an IL2Rγ-mediated disease or disorder.

[0890] 31. The method according to embodiment 30, wherein the IL2Rγ-mediated disease or disorder is graft-versus-host disease, organ transplant rejection, b-islet cell transplant rejection, skin transplant rejection, heart transplant rejection, lung transplant rejection, kidney transplant rejection, liver transplant rejection, birdshot chorioretinopathy, multiple sclerosis, uveitis, autoimmune disease, type I diabetes, multiple sclerosis, rheumatoid arthritis, systemic lupus erythematosus, myasthenia gravis, aplastic anemia, atopic dermatitis, asthma, mast cell activation disorder, mast cell activation syndrome (MCAS), systemic mastocytosis (SM), and / or mast cell leukemia (MCL).

[0891] Examples

[0892] The following examples are provided to give a complete disclosure and description to those of ordinary skill in the art of how to make and use the methods and compositions of the invention, and are not intended to limit the scope that the inventors regard as their invention.

[0893] Example 1: Identification and isolation of anti-IL2Rγ antibodies

[0894] Immunize with an IL2Rγ protein immunogen comprising the IL2Rγ extracellular sequence (ecto domain) Mice (i.e., engineered mice containing DNA encoding the variable regions of the human immunoglobulin heavy and K light chains) to obtain anti-IL2Rγ antibodies.

[0895] Specifically, the immunogen human IL2Rg ecto-mmh comprises:

[0896] · Amino acids (1-240): human IL2Rg ecto (L23-A262 of NP_000197.1), and

[0897] · Amino acids (241-268): Myc-Myc-hexahistidine tag (underlined);

[0898] It comprises the amino acid sequence:

[0899]

[0900] *Expression with mROR signal sequence

[0901] The antibody immune response was monitored using an IL2Rγ-specific immunoassay. Fully human anti-IL2Rγ antibodies were isolated and purified.

[0902] Table 1-1. Summary of anti-IL2Rγ VH, VK, and CDR amino acid sequences*

[0903]

[0904] *The numbers refer to SEQ ID NO corresponding to the indicated sequences.

[0905] Table 1-2. Summary of anti-IL2Rγ heavy and light chain amino acid sequences*

[0906]

[0907] *The numbers refer to SEQ ID NO corresponding to the indicated sequences.

[0908] The amino acid sequences of the anti-IL2Rγ antibody immunoglobulin heavy and light chains are shown below (CDRs are underlined; variable regions are in bold).

[0909] H4H12857P

[0910] Heavy chain (SEQ ID NO: 311)

[0911]

[0912] Light chain (SEQ ID NO: 313)

[0913]

[0914] H4H12858P

[0915] Heavy chain (SEQ ID NO: 331)

[0916]

[0917] Light chain (SEQ ID NO: 333)

[0918]

[0919] H4H12859P

[0920] Heavy chain (SEQ ID NO: 18)

[0921]

[0922] Light chain (SEQ ID NO: 20)

[0923]

[0924] H4H12863P

[0925] Heavy chain (SEQ ID NO: 38)

[0926]

[0927] Light chain (SEQ ID NO: 40)

[0928]

[0929] H4H12874P

[0930] Heavy chain (SEQ ID NO: 58)

[0931]

[0932] Light chain (SEQ ID NO: 60)

[0933]

[0934] H4H12871P

[0935] Heavy chain (SEQ ID NO: 376)

[0936]

[0937] Light chain (SEQ ID NO: 378)

[0938]

[0939] H4H12884P

[0940] Heavy chain (SEQ ID NO: 77)

[0941]

[0942] Light chain (SEQ ID NO: 79)

[0943]

[0944] H4H12886P

[0945] Heavy chain (SEQ ID NO: 97)

[0946]

[0947] Light chain (SEQ ID NO: 99)

[0948]

[0949] H4H12889P

[0950] Heavy chain (SEQ ID NO: 357)

[0951]

[0952] Light chain (SEQ ID NO: 359)

[0953]

[0954] H4H12890P

[0955] Heavy chain (SEQ ID NO: 115)

[0956]

[0957] Light chain (SEQ ID NO: 117)

[0958]

[0959] H4H12899P

[0960] Heavy chain (SEQ ID NO: 134)

[0961]

[0962] Light chain (SEQ ID NO: 136)

[0963]

[0964] H4H12900P

[0965] Heavy chain (SEQ ID NO: 152)

[0966]

[0967] Light chain (SEQ ID NO: 154)

[0968]

[0969] H4H12908P

[0970] Heavy chain (SEQ ID NO: 170)

[0971]

[0972] Light chain (SEQ ID NO: 172)

[0973]

[0974] H4H12913P2

[0975] Heavy chain (SEQ ID NO: 186)

[0976]

[0977] Light chain (SEQ ID NO: 188)

[0978]

[0979] H4H12922P2

[0980] Heavy chain (SEQ ID NO: 343)

[0981]

[0982] Light chain (SEQ ID NO: 188)

[0983]

[0984] H4H12924P2

[0985] Heavy chain (SEQ ID NO: 198)

[0986]

[0987] Light chain (SEQ ID NO: 188)

[0988]

[0989] H4H12926P2

[0990] Heavy chain (SEQ ID NO: 208)

[0991]

[0992] Light chain (SEQ ID NO: 188)

[0993]

[0994] H4H12927P2

[0995] Heavy chain (SEQ ID NO: 216)

[0996]

[0997] Light chain (SEQ ID NO: 188)

[0998]

[0999] H4H12934P2

[1000] Heavy chain (SEQ ID NO: 234)

[1001]

[1002] Light chain (SEQ ID NO: 236)

[1003]

[1004] H4H13538P

[1005] Heavy chain (SEQ ID NO: 254)

[1006]

[1007] Light chain (SEQ ID NO: 256)

[1008]

[1009] H4H13541P

[1010] Heavy chain (SEQ ID NO: 272)

[1011]

[1012] Light chain (SEQ ID NO: 274)

[1013]

[1014] H4H13544P2

[1015] Heavy chain (SEQ ID NO: 284)

[1016]

[1017] Light chain (SEQ ID NO: 188)

[1018]

[1019] H4H13545P2

[1020] Heavy chain (SEQ ID NO: 294)

[1021]

[1022] Light chain (SEQ ID NO: 188)

[1023]

[1024] *The antibodies mentioned in these examples are antibodies having immunoglobulin chains containing the amino acid sequences specifically shown in Example 1.

[1025] Example 2 : Surface plasmon resonance binding assay

[1026] Using a real-time surface plasmon resonance-based Biacore 4000 biosensor platform, the dissociation rate constant (k d ) of the IL-2Rγ reagent binding to the purified anti-IL2Rγ monoclonal antibody was determined. All binding studies were performed at 25 °C and 37 °C using two running buffers: (i) 1.9 mM NaH2PO4, 8.1 mM Na2HPO4, 2.7 mM KCl, 137 mM NaCl, 0.03% NaN3, 0.05% v / v surfactant Tween-20, pH 7.4 (PBS-T-pH 7.4) and (ii) 8.8 mM NaH2PO4, 1.2 mM Na2HPO4, 2.7 mM KCl, 137 mM NaCl, 0.03% NaN3, 0.05% v / v surfactant Tween-20, pH 6.0 (PBS-T-pH 6.0). The anti-IL2Rγ monoclonal antibody expressing human IgG4 Fc was captured using an amine-derivatized CM5 Biacore sensor surface conjugated with a monoclonal mouse anti-human Fc antibody (GE, catalog #BR-1008-39). All IL2Rγ reagents were expressed with a C-terminal myc-myc-hexahistidine tag (subsequently denoted with the -MMH suffix). Different concentrations of the extracellular domain of human IL2Rγ expressing a C-terminal myc-myc-hexahistidine tag (hIL-2Rg-MMH; SEQ ID NO: 379) or the extracellular domain of cynomolgus monkey IL2Rγ expressing a C-terminal myc-myc-hexahistidine tag (mfIL-2Rg-MMH; SEQ ID NO: 380) were prepared in PBS-T-pH 7.4 running buffer (100 nM - 11.11 nM, 3-fold serial dilution) and injected at a flow rate of 30 μL / min for 4 minutes. The dissociation of the bound IL-2Rg-MMH was carried out in PBS-T-pH 7.4 or PBS-T-pH 6.0 running buffer for 6 minutes.

[1027] By fitting the real-time binding sensorgram to a 1:1 binding model using Scrubber 2.0c curve fitting software, the dissociation rate constant (k d ) in the two running buffers was determined. The dissociation rate values of the anti-Hemojuvelin mAb binding to hIL-2RG-MMH and mfIL-2RG-MMH at 25 °C and 37 °C in PBS-T-pH 7.4 and PBS-T-pH 6.0 are shown in Tables 2-1 to 2-8.

[1028] Table 2-1. Dissociation rate constants of anti-IL-2Rγ mAb binding to hIL-2Rγ-MMH in PBS-T-pH 7.4 at 25 °C

[1029]

[1030] Table 2-2. Dissociation rate constants of anti-IL-2Rγ mAb binding to hIL-2Rγ-MMH in PBS-T-pH 6.0 at 25 °C

[1031]

[1032] Table 2-3. Dissociation rate constants of anti-IL-2Rγ mAb binding to hIL-2Rγ-MMH in PBS-T-pH 7.4 at 37 °C

[1033]

[1034] Table 2-4. Dissociation rate constants of anti-IL-2Rγ mAb binding to hIL-2Rγ-MMH in PBS-T-pH 6.0 at 37 °C

[1035]

[1036] Table 2-5. Dissociation rate constants of anti-IL-2Rγ mAb binding to mfIL-2Rγ-MMH in PBS-T-pH 7.4 at 25 °C

[1037]

[1038] Table 2-6. Dissociation rate constants of anti-IL-2Rγ mAb binding to mfIL-2Rγ-MMH in PBS-T-pH 6.0 at 25 °C

[1039]

[1040] Table 2-7. Dissociation rate constants of anti-IL-2Rγ mAb binding to mfIL-2Rγ-MMH in PBS-T-pH 7.4 at 37 °C

[1041]

[1042] Table 2-8. Dissociation rate constants of anti-IL-2Rγ mAb binding to mfIL-2Rγ-MMH in PBS-T-pH 6.0 at 37 °C

[1043]

[1044] Example 3 : Binding kinetics

[1045] The equilibrium dissociation constant (K D d) of the binding of IL-2Rγ to the purified anti-IL-2Rγ monoclonal antibody was determined using a Biacore 4000 instrument equipped with a real-time surface plasmon resonance biosensor. All binding studies were performed at 25 °C and 37 °C in a running buffer of 10 mM HEPES, 150 mM NaCl, 3 mM EDTA, and 0.05% v / v surfactant Tween-20, pH 7.4 (HBS-ET). The Biacore sensor surface was first derivatized with amine conjugated to monoclonal mouse anti-human Fc antibody (GE, #BR-1008-39) to capture the anti-IL2Rγ monoclonal antibody.

[1046] Binding studies were performed on the following IL-2Rγ reagents:

[1047] · Human IL2Rγ extracellular domain (hIL-2Rg-MMH; SEQ ID NO: 379) expressing with a C-terminal myc-myc-hexahistidine tag, which contains:

[1048] Amino acids (1 - 240): Human IL2Rg extracellular (L23 - A262 of NP_000197.1)

[1049] Amino acids (241 - 268): Myc-Myc-hexahistidine tag (underlined)

[1050] It contains the amino acid sequence:

[1051]

[1052] * Expressing with an mROR signal sequence

[1053] · Cynomolgus monkey IL2Rγ extracellular domain (mfIL-2Rg-MMH; SEQ ID NO: 380) expressing with a C-terminal myc-myc-hexahistidine tag, which contains:

[1054] Amino acids (1 - 240): Cynomolgus monkey IL2Rg extracellular (L23 - A262 of XP_005593949.1)

[1055] Amino acids (241 - 268): Myc-Myc-hexahistidine tag (underlined)

[1056] It contains the amino acid sequence:

[1057]

[1058] · Express the extracellular domain of human IL2Rγ with a C-terminal murine IgG2a Fc tag (hIL-2Rg-mFc; SEQ ID NO: 381), which contains:

[1059] Amino acids (1-240): extracellular human IL2Rg (L23-A262 of NP_000197.1)

[1060] Amino acids (241-473): murine IgG2a Fc tag (underlined)

[1061] It contains the amino acid sequence:

[1062]

[1063] * Express with the mROR signal sequence

[1064] · Express the D1 domain of the extracellular domain of human IL-2Rγ with a C-terminal myc-myc-hexahistidine tag (hIL-2Rg_D1-MMH; SEQ ID NO: 382), which contains:

[1065] Amino acids (1-131): human IL2Rg domain 1 (L23-I153 of NP_000197.1)

[1066] Amino acids (132-159): Myc-Myc-hexahistidine tag (underlined)

[1067] It contains the amino acid sequence:

[1068]

[1069] * Express with the mROR signal sequence

[1070] · Express the D2 domain of the extracellular domain of human IL2Rγ with a C-terminal myc-myc-hexahistidine tag (hIL-2Rg_D2-MMH; SEQ ID NO: 383), which contains:

[1071] Amino acids (1-88): human IL2Rg domain 2 (P154-S241 of NP_000197.1)

[1072] Amino acids (89-116): Myc-Myc-hexahistidine tag (underlined)

[1073] It contains the amino acid sequence:

[1074]

[1075] * Express with the mROR signal sequence

[1076] · Express the extracellular domain of murine IL2Rγ with a C-terminal myc-myc-hexahistidine tag (mIL-2Rg-MMH; SEQ ID NO: 384), which comprises:

[1077] Amino acids (1-241): extracellular murine IL2Rg (W23-A263 of NP_038591.1)

[1078] Amino acids (242-269): Myc-Myc-hexahistidine tag (underlined)

[1079] It comprises the amino acid sequence:

[1080]

[1081] · Express the extracellular domain of rat IL2Rγ with a C-terminal myc-myc-hexahistidine tag (rIL-2Rg-MMH; SEQ ID NO: 385), which comprises:

[1082] Amino acids (1-240): extracellular rat IL2Rg (W23-A262 of NP_543165.1)

[1083] Amino acids (241-268): Myc-Myc-hexahistidine tag (underlined)

[1084] It comprises the amino acid sequence:

[1085]

[1086] * Express with the mROR signal sequence

[1087] Prepare IL2Rγ reagents at different concentrations in HBS-ET running buffer (100 nM - 6.25 nM; 4-fold serial dilution or 50 nM - 3.125 nM; 4-fold serial dilution for hIL-2Rg-mFc) and inject at a flow rate of 30 μL / min for 4 minutes onto the surface of an anti-human Fc captured anti-IL2Rγ monoclonal antibody. Monitor the dissociation of the monoclonal antibody-bound IL2Rγ reagent in HBS-ET running buffer for 8 to 10 minutes. The kinetic association (k a ) and dissociation (k d ) rate constants are determined by fitting the real-time sensorgrams to a 1:1 binding model using Scrubber2.0c curve fitting software. The binding dissociation equilibrium constant (K D ) and dissociation half-life (t1 / 2) are calculated from the kinetic rate constants as follows:

[1088] and

[1089] The kinetic parameters of the binding of various IL-2Rγ reagents to different IL2Rγ monoclonal antibodies at 25 °C and 37 °C are shown in Tables 3-1 to 3-14.

[1090] Table 3-1. Binding kinetic parameters of hIL-2Rg-MMH binding to IL-2Rγ monoclonal antibody at 25 °C

[1091]

[1092] Table 3-2. Binding kinetic parameters of hIL-2Rg-MMH binding to IL-2Rγ monoclonal antibody at 37 °C

[1093]

[1094] *NB indicates that no binding was observed under the current experimental conditions.

[1095] Table 3-3. Binding kinetic parameters of mfIL-2Rg-MMH binding to IL-2Rγ monoclonal antibody at 25 °C

[1096]

[1097] Table 3-4. Binding kinetic parameters of mfIL-2Rg-MMH binding to IL-2Rγ monoclonal antibody at 37 °C

[1098]

[1099] *NB indicates that no binding was observed under the current experimental conditions.

[1100] Table 3-5. Binding kinetic parameters of hIL-2Rg-mFc binding to IL-2Rγ monoclonal antibody at 25 °C

[1101]

[1102] *NB indicates that no binding was observed under the current experimental conditions.

[1103] # Indicates that no dissociation was observed under the current experimental conditions, and the k d value was artificially fixed at 1.00E-05 s -1 .

[1104] Table 3-6. Binding kinetic parameters of hIL-2Rg-mFc binding to IL-2Rγ monoclonal antibody at 37 °C

[1105]

[1106] *NB indicates that no binding was observed under the current experimental conditions.

[1107] # Indicates that no dissociation was observed under the current experimental conditions, and the k d value was artificially fixed at 1.00E-05 s -1 .

[1108] $ Indicates that the binding data is inconclusive for generating reliable binding kinetic values and K D .

[1109] Table 3-7. Binding kinetic parameters of mIL-2Rg-MMH binding to IL-2Rγ monoclonal antibody at 25 °C

[1110]

[1111] *NB indicates that no binding was observed under the current experimental conditions.

[1112] Table 3-8. Binding kinetic parameters of mIL-2Rg-MMH binding to IL-2Rγ monoclonal antibody at 37 °C

[1113]

[1114] *NB indicates that no binding was observed under the current experimental conditions.

[1115] Table 3-9. Binding kinetic parameters of rat IL-2Rg-MMH binding to IL-2Rγ monoclonal antibody at 25 °C

[1116]

[1117] *NB indicates that no binding was observed under the current experimental conditions.

[1118] Table 3-10. Binding kinetic parameters of rat IL-2Rg-MMH binding to IL-2Rγ monoclonal antibody at 37 °C

[1119]

[1120] *NB indicates that no binding was observed under the current experimental conditions.

[1121] Table 3-11. Binding kinetic parameters of hIL-2Rg_D1-MMH binding to IL-2Rγ monoclonal antibody at 25 °C

[1122]

[1123] *NB indicates that no binding was observed under the current experimental conditions.

[1124] Table 3-12. Binding kinetic parameters of hIL-2Rg_D1-MMH binding to IL-2Rγ monoclonal antibody at 37 °C

[1125]

[1126] *NB indicates that no binding was observed under the current experimental conditions.

[1127] Table 3-13. Binding kinetic parameters of hIL-2Rg_D2-MMH binding to IL-2Rγ monoclonal antibody at 25 °C

[1128]

[1129] *NB indicates that no binding was observed under the current experimental conditions.

[1130] Table 3-14. Binding kinetic parameters of hIL-2Rg_D2-MMH binding to IL-2Rγ monoclonal antibody at 37 °C

[1131]

[1132] *NB indicates that no binding was observed under the current experimental conditions.

[1133] Example 4 : Octet cross-competition between different anti-IL-2Rγ monoclonal antibodies

[1134] Binding competition between a panel of anti-IL2Rγ monoclonal antibodies was determined using real-time, label-free biolayer interferometry assays on an Octet HTX biosensor platform (Pall ForteBio). The entire experiment was conducted at 25 °C in 10 mM HEPES, 150 mM NaCl, 3 mM EDTA, and 0.05% v / v surfactant Tween-20, 1 mg / mL BSA, pH 7.4 (HBS-EBT) buffer using a plate shaker speed of 1000 rpm. To evaluate whether two antibodies compete with each other for binding to their respective epitopes on human IL2Rγ extracellular domain (hIL-2Rγ-MMH; SEQ ID: 379) expressing a C-terminal myc-myc-hexahistidine tag, Octet biosensor tips coated with anti-penta-histidine antibody (Fortebio Inc, #18-5122) were used to capture approximately 0.27 nM hIL-2Rγ-MMH by dipping the biosensor tips into wells containing 10 μg / mL hIL-2Rγ-MMH for 3 minutes. The biosensor tips capturing the antigen were then saturated with the first anti-IL2Rγ monoclonal antibody (subsequently referred to as mAb-1) by dipping into wells containing 50 μg / mL mAb-1 for 300 seconds. Subsequently, the biosensor tips were then dipped into wells containing 50 μg / mL of the second anti-IL2Rγ monoclonal antibody (subsequently referred to as mAb-2) for 240 seconds. Between each step of the experiment, the biosensor tips were washed in HBS-ETB buffer. Real-time binding responses were monitored throughout the experiment, and the binding responses at the end of each step were recorded. The response of mAb-2 binding to hIL-2Rγ-MMH pre-complexed with mAb-1 was compared, and the competitive / non-competitive behavior of different anti-IL2Rγ monoclonal antibodies was determined, as shown in Table 4-1.

[1135] Table 4-1. Cross-competition between anti-IL-2Rγ monoclonal antibodies

[1136]

[1137]

[1138]

[1139]

[1140] Example 5 : Flow cytometry analysis of STAT phosphorylation in human CD4+ T cells (human PBMC)

[1141] To evaluate the in vitro properties of the IL2Rγ antibodies of the present invention, by flow cytometry (BD TMThe Phosflow assay measured their ability to block CD4 T cell activation induced by IL-2, IL-4, IL-7, IL-15, and IL-21. + BD TM Phosflow allows simultaneous analysis of intracellular phosphoproteins (e.g., STAT proteins) and cell surface markers to analyze cell signaling in discrete cell subsets. This technique was used to analyze STAT phosphorylation in human CD4 T cells following cytokine stimulation from the γc family. +

[1142] Human peripheral blood mononuclear cells (PBMCs) were isolated from fresh whole blood (Bioreclammation IVT) by density gradient centrifugation. K2 EDTA whole blood was diluted 1:1 in X-VIVO 15 medium (Lonza), added to a SepMate tube (StemCell) containing Ficoll Paque PLUS (Healthcare), and centrifuged to isolate PBMCs. The upper layer containing PBMCs was transferred to a new tube and washed twice with DPBS (Life Technologies). The PBMCs were then resuspended at a concentration of approximately 5.0 x 10 cells / mL in X-VIVO 15 medium, plated in 96-well plates (50 μL of cells / well; approximately 250,000 cells / well), and incubated at 37 °C for 2 hours before adding cytokines and antibodies. TM 6 TM 15 medium, and incubated at 37 °C for 2 hours before adding cytokines and antibodies.

[1143] Serial dilutions (1:5) of antibodies were prepared in pre-warmed X-VIVO 15 medium and added to the cells (50 μL), with the final antibody concentration starting at 400 nM. Fixed cytokine concentrations were prepared in pre-warmed X-VIVO 15 medium and added to the cells (100 μL): the final concentration of IL-7 (R&D Systems) was 1 pM, the final concentrations of IL-4 (R&D Systems) and IL-21 (eBioscience) were 50 pM, the final concentration of IL-15 (R&D Systems) was 0.5 nM, and the final concentration of IL-2 (R&D Systems) was 10 nM; the final volume per well was 200 μL. TM TM

[1144] For cytokine dose responses, serial dilutions (1:5) of each cytokine were also prepared in pre-warmed X-VIVO TMPrepared in 15 medium, the final cytokine concentrations of IL-4, IL-7 and IL-21 start from 5 nM, or the final cytokine concentrations of IL-2 and IL-15 start from 50 nM. First, add 50 μL of X-VIVO TM 15 medium, then add 100 μL of serial dilutions of cytokines, and the total volume per well is 200 μL. After adding cytokines and antibodies to the cells, incubate them at 37 °C for 15 minutes to allow PBMC activation (STAT phosphorylation). Then stop the stimulation by adding 200 μL of warm Cytofix (BD) to each well and incubate the cells at 37 °C for 10 minutes (fixation step). Then wash the cells twice with staining buffer (BD) and keep them overnight at 4 °C. The next day, centrifuge the cells and permeabilize the cells by slowly adding 100 μL of cold permeabilization buffer III (BD) to the pellet. Incubate the cells at 4 °C for 30 minutes, then wash them twice with staining buffer. To be able to analyze the CD4 + T cell population for measuring STAT phosphorylation, stain the cells with a mixture of human FcR-binding inhibitor (eBioscience; 1 / 10), anti-CD33-PE (BD; 1 / 200), anti-CD4-PacificBlue (BD; 1 / 200), anti-CD3-PECy7 (BD; 1 / 200) and the relevant anti-phospho-STAT-AlexaFluor647 (BD) prepared in staining buffer:

[1145] - Anti-phospho STAT3 (1 / 10): for cells stimulated by IL-21,

[1146] - Anti-phospho STAT5 (1 / 20): for cells stimulated by IL-2, IL-7 and IL-15,

[1147] - Anti-phospho STAT6 (1 / 10): for cells stimulated by IL-4.

[1148] Keep the samples in the dark at room temperature for 1 hour. Then centrifuge the cells and wash them twice with staining buffer. Acquire sample data on an LSR Fortessa X-20 cell analyzer using an HTS attachment (BD). Perform data analysis using FlowJo X software (Tree Star, OR). Define CD4 + T cells as intact cells, singlets, CD33 - , CD3 + , CD4 + ; and analyze STAT phosphorylation (MFI = mean fluorescence intensity) in this cell population.

[1149] Both H4H12889P and H4H12922P2 similarly and effectively blocked STAT phosphorylation induced by all cytokines (IL-2, IL-4, IL-7, IL-15, and IL-21) tested in this assay, while H4H12874P, H4H12886P, H4H12857P, and the comparator antibody COMP1499 (anti-IL2Rγ antibody CP.B8, see US2002 / 0028202) only partially blocked or did not block cytokine-induced STAT phosphorylation.

[1150] Table 5-1. Anti-IL-2Rγ antibodies H4H12889P and H4H12922P2 block human CD4 + STAT phosphorylation induced by human IL-2, IL-4, IL7, IL-15, and IL-21 in T cells

[1151] IC50 [M] IL-2 IL-4 IL-7 IL-15 IL-21 Constant 10 nM 50 pM 1 pM 0.5 nM 50 pM H4H12889P 2.06E-09 1.10E-09 8.92E-10 2.55E-09 2.28E-09 H4H12922P2 1.87E-09 8.54E-10 5.80E-10 2.46E-09 2.21E-09

[1152] *IC50 values measured for the two antibodies are shown at the indicated concentrations for different interleukins.

[1153] See also insets (A)-(E) of Figure 1, where the STAT phosphorylation levels were determined for each tested antibody concentration.

[1154] Example 6 : Flow cytometry analysis of STAT3 phosphorylation in in vitro differentiated human mast cells

[1155] To evaluate the in vitro properties of the anti-IL2Rγ antibodies of the present invention, their ability to block IL-9-induced activation of human mast cells was measured by flow cytometry (BD TM Phosflow assay). We used this technique to observe STAT3 phosphorylation in in vitro differentiated human mast cells after human IL-9 stimulation.

[1156] Briefly, human mast cells were generated in vitro from bone marrow CD133 + progenitor cells cultured for 6 weeks in StemSpan serum-free medium supplemented with human SCF, IL-6, and IL-3.

[1157] Human mast cells were resuspended at a concentration of approximately 4.0x10 6 cells / mL in X-VIVO TM 15 medium, plated in 96-well plates (50 μL cells / well; approximately 200,000 cells / well), and incubated at 37 °C for 2 hours before adding cytokines and antibodies.

[1158] In pre-warmed X-VIVO TMPrepare serial dilutions (1:5) of the antibody in X-VIVO TM 15 medium and add it to the cells (50 μL), starting with a final antibody concentration of 400 nM.

[1159] For cytokine dose response, serial dilutions (1:5) of IL-9 are also prepared in pre-warmed X-VIVO TM 15 medium, starting with a final cytokine concentration of 100 nM. First, add 50 μL of X-VIVO TM 15 medium to the cells, then add 100 μL of the serial dilutions of the cytokine, with a total volume of 200 μL per well.

[1160] After adding the cytokine and antibody to the cells, incubate them at 37 °C for 15 minutes to allow mast cell activation (measured by STAT3 phosphorylation). Then stop the stimulation by adding 200 μL of warm Cytofix (BD) to each well and incubate the cells at 37 °C for 10 minutes (fixation step). Then wash the cells twice with staining buffer (BD) and keep them at 4 °C overnight. The next day, centrifuge the cells and permeabilize the cells by slowly adding 100 μL of cold permeabilization buffer III (BD) to the pellet. Incubate the cells at 4 °C for 30 minutes, then wash them twice with staining buffer. Then, stain the mast cells with a mixture of human FcR-binding inhibitor (eBioscience; 1 / 10), anti-c-Kit-PE (BD; 1 / 100), and anti-phospho-STAT3-AlexaFluor647 (BD; 1 / 10) prepared in staining buffer.

[1161] Keep the samples in the dark at room temperature for 1 hour. Then centrifuge the cells and wash them twice with staining buffer. Acquire sample data on an LSR Fortessa X-20 cell analyzer using an HTS attachment (BD). Perform data analysis using FlowJo X software (Tree Star, OR). Define mast cells as intact cells, singlet, c-Kit + ; and analyze STAT3 phosphorylation (MFI = mean fluorescence intensity) in this cell population.

[1162] Both H4H12889P and H4H12922P2 similarly and effectively blocked IL-9-induced STAT3 phosphorylation.

[1163] Table 6-1. Blockade of IL-9-induced STAT3 phosphorylation in human mast cells differentiated in vitro by anti-IL-2Rγ antibodies H4H12889P and H4H12922P2

[1164] IC50 [M] IL-9 Constant 2 nM H4H12889P 4.41E-10 H4H12922P2 4.16E-10

[1165] *Indicates the IC50 values measured for the two antibodies when the IL-9 concentration was 2 nM.

[1166] See also Figure 2 , in which the level of STAT phosphorylation induced by IL-9 was determined for each concentration of the antibody tested.

[1167] Example 7 : In vivo model for assessing the blocking activity of IL-2Rγ antibodies as a therapeutic treatment: monoclonal antibody testing in a xenogeneic acute graft-versus-host disease model

[1168] To determine the role of our anti-IL2Rγ antibodies H4H12889P and H4H12922P2 and the comparator IL-2Rγ antibody COMP1499 in relevant in vivo models, a xenogeneic acute graft-versus-host disease (GvHD) study was conducted. Briefly, to induce GvHD in mice, human peripheral blood mononuclear cells (huPBMC) were injected into NOD-scid IL2rγ null (NSG) mice (Jackson Laboratory). After transplantation, human immune cells recognize the mouse host as xenogeneic and mount a strong immune response against its tissues.

[1169] In this experiment, 10 million huPBMCs (ReachBio) resuspended in DPBS were retro-orbitally injected into NSG mice (Jackson Lab) (10 million cells / 100 μL; 5 groups, 10 mice per group). Briefly, on the day of injection, human PBMCs were thawed in IMDM medium (Irvine Scientific) supplemented with 10% FBS (Seradigm) and incubated in this supplemented medium at 37 °C for 2 hours. The cells were then washed in DPBS (Life Technologies) and resuspended at 10 million cells / 100 μl for injection. 100 μL of PBS was retro-orbitally injected into the control group (10 mice). Four groups of huPBMC-transplanted NSG mice began subcutaneous injection of 25 mg / kg of H4H12889P, H4H12922P2, COMP1499, or isotype control antibody (REGN1945; human anti-domestic cat (Felis domesticus) Fel d1 antibody (IgG4(S108P) / K)) 3 weeks after huPBMC injection, then twice a week for 6 weeks. The experiment was terminated by sacrificing the remaining mice on day 161 after huPBMC transplantation. The experimental drug administration and treatment regimens for each group of mice are shown in Table 7-1.

[1170] Table 7-1. Experimental drug administration and treatment regimens for each group of mice

[1171] Group NSG mice huPBMC injection Antibody 1 10 None None 2 10 10 million None 3 10 10 million Isotype control antibody (REGN1945) 4 10 10 million IL-2Rγ antibody (COMP1499) 5 10 10 million IL-2Rγ antibody (H4H12889P) 6 10 10 million IL-2Rγ antibody (H4H12922P2)

[1172] Throughout the experiment, the body weight loss and death of the mice were monitored twice a week (to evaluate the effect of the therapeutic antibody on survival). The human cell engraftment in the blood and the serum mouse and human cytokine levels were evaluated at different time points, as shown in Table 7-2.

[1173] Table 7-2. Blood / serum collection dates and readouts

[1174]

[1175] Throughout the experiment, the body weight loss of the mice was monitored twice a week ( Figure 3 panels (A)-(F); % of the initial body weight on the day of huPBMC engraftment) and death ( Figure 4 ; to evaluate the effect of the therapeutic antibody on survival). Animals showing a 20% body weight loss from the initial body weight were euthanized.

[1176] At different time points after huPBMC injection, blood samples from mice were collected into Microtainer tubes (BD, Cat# 3659740), and human cell engraftment was evaluated by observing the absolute number of human cells in the blood by flow cytometry. Briefly, 50 μL of each blood sample was incubated in ACK lysis buffer (Gibco) for 5 minutes at room temperature to lyse red blood cells. The cells were then washed in DPBS, stained with LIVE / DEAD Fixable Dead Cell Stain (Invitrogen), washed in MACS buffer (Miltenyi Biotec), and labeled with a mixture of antibodies for identification of human CD45 + cells, T cells, CD4 + T cells, and CD8 + T cells (anti-human CD45, anti-human CD3, anti-human CD4, and anti-human CD8 [BD] diluted 1 / 50 in Brilliant Staining Buffer [BD], and human and mouse Fc inhibitor antibodies [eBioscience and BD, respectively]). Finally, the samples were washed in MACS buffer, fixed in BD CytoFix (BD), and then resuspended in MACS buffer containing CountBright beads (Life Technologies) to calculate the absolute number of cells in each sample. Sample data were acquired on an LSR Fortessa X-20 cell analyzer using the HTS accessory (BD). Data analysis was performed using FlowJo X software (Tree Star, OR). Human CD45 + T cells were defined as live cells, singlets, CD45 + , and within this population, CD4 + T cells and CD8 + T cells were further defined as CD3 + , CD4 + and CD3 + , CD8 + .

[1177] Table 7-3. Human immune cells in blood on days 35 and 56 after huPBMC injection (mean ± SD, cells / μL blood)

[1178]

[1179] Note:Shows the statistical significance determined by Kruskal-Wallis one-way ANOVA and Dunn's multiple comparison post hoc test (* = p < 0.05, ** = p < 0.01, *** = p < 0.001, compared to Group 3: huPBMC-isotype control antibody). n = number of mice analyzed.

[1180] As an example, Figure 5 The absolute human cell numbers in blood on day 35 after huPBMC injection are shown in Panels (A) - (D). Blood counts of human CD45+ cells, T cells, CD4+ T cells, and CD8+ T cells over time are shown in Figure 6 Panels (A) - (D).

[1181] Serum was collected from mice at different days after huPBMC injection, and the serum levels of mouse and human cytokines were evaluated. Briefly, whole blood was collected into Microtainer tubes (BD, Cat#365967) and allowed to clot by leaving it undisturbed at room temperature for at least 30 minutes. The clotted blood and cells were pelleted by centrifugation at 15,000 x g for 10 minutes at 4°C. The resulting supernatant (referred to as serum) was transferred to a clean plate, and cytokine concentrations in the serum were measured using two pro-inflammatory (mouse and human) multiplex immunoassay kits (Meso Scale Discovery) according to the manufacturer's instructions. The plates were washed with PBS containing 0.05% (w / v) Tween-20 (Life Technologies). Electrochemiluminescence was read immediately on an MSD Spector instrument. Data analysis was performed using FlowJo X software (Tree Star, OR).

[1182] Table 7-4. Serum human cytokine concentrations on days 42 and 62 after huPBMC injection (mean ± SD, in pg / mL)

[1183]

[1184] Note: Shows the statistical significance determined by Kruskal-Wallis one-way ANOVA and Dunn's multiple comparison post hoc test (* = p < 0.05, ** = p < 0.01, *** = p < 0.001, compared to Group 3: huPBMC-isotype control antibody). n = number of mice analyzed.

[1185] Table 7-5. Serum mouse cytokine concentrations on days 42 and 62 after huPBMC injection (mean ± SD, in pg / mL)

[1186]

[1187] Note: Shows the statistical significance determined by Kruskal-Wallis one-way ANOVA and Dunn's multiple comparison post hoc test (* = p < 0.05, ** = p < 0.01, *** = p < 0.001, compared to Group 3: huPBMC-isotype control antibody). n = number of mice analyzed.

[1188] In addition, as an example, the serum human and murine cytokine levels on Day 42 after huPBMC injection are shown in Figure 7 panels (A) - (I). Figure 8 The serum levels of human IFN-γ, human TNFα, murine TNFα, and murine IL-6 over time are shown in panels (A) - (D).

[1189] This in vivo study demonstrated the efficacy of the anti-IL2Rγ antibodies H4H12889P and H4H12922P2 when administered therapeutically in a graft-versus-host disease model. Both H4H12889P and H4H12922P2 were able to effectively block the development of GvHD in mice, while COMP1499 was not. Mice treated therapeutically with either of these two antibodies were spared from weight loss and death, and this was associated with a dramatic decrease in murine and human serum cytokine levels as well as the number of human T cells in the blood. See Tables 7-3, 7-4, and 7-5.

[1190] Example 8 : Bioluminescence assays performed using NK92 / hIL7R / STAT3-Luc and Ramos.2G6.4C10 / STAT3-Luc cells

[1191] IL2Rγ cytokine family: IL-2, IL-4, IL-7, IL-9, IL-15, and IL-21 signal through the JAK-STAT (Janus kinase-signal transducer and activator of transcription) pathway (Rochman et al., New insights into the regulation of T cells by gamma(c)family cytokines. Nat Rev Immunol. 2009 Jul;9(7):480-90). To assess the inhibition of cytokine signaling by anti-IL2Rγ antibodies, a bioassay was developed using NK-92 cells (human natural killer cell line, ATCC) stably expressing a luciferase reporter (STAT3-Luc; SA Biosciences, #CLS-6028L). NK-92 endogenously expresses IL2Rγ and ligand-selective receptors that mediate IL-2, IL-9, IL-15, and IL-21 signaling. To also assess the regulation of IL-7 signaling, NK-92 cells were transduced with a lentivirus containing human IL-7R, and stably expressing cells were selected and maintained in G418. The resulting cell line is hereinafter referred to as NK-92 / hIL7R / STAT3-Luc. To test the regulation of IL-4-mediated signaling, Ramos.2G6.4C10 (human B lymphocyte cell line, ATCC) cells endogenously expressing IL2Rγ and the IL-4R receptor were transduced with the STAT3-luc reporter, and the resulting cell line is called Ramos.2G6.4C10 / STAT3-Luc.

[1192] The inhibition of the anti-IL2γ antibody of the present invention on human IL-2 (hIL-2), human IL-7 (hIL-7), human IL-9 (hIL-9), human IL-15 (hIL-15) or human IL-21 (hIL-21) signaling was tested by plating 20,000 NK-92 / hIL7R / STAT3-Luc cells per well in a 96-well plate in growth medium (prepared according to the instructions of ATCC, but without IL-2) and incubating overnight at 37 °C in 5% CO2. The next day, the anti-IL2Rγ antibody or isotype control was serially diluted from 500 to 0.008 nM in assay buffer (plus samples containing only buffer without the test molecule), added to the cells and incubated for 30 minutes. After incubation, the ligands were added to the cells at the following final concentrations: 30 pM hIL-2, 50 pM hIL-7, 20 pM hIL-9, 60 pM or 100 pM hIL-15, or 5 pM or 3 pM hIL-21. Serial dilutions of the ligands (from 10 nM to 0.2 pM) (plus samples containing only buffer without the ligand) were added to the cells to determine dose-dependent activation. After incubation for 5 hours at 37 °C in 5% CO2, luciferase activity was measured using OneGlo TM Reagent (Promega, #E6031) and Victor TM X Multilabel Plate Reader (PerkinElmer).

[1193] To test the inhibition of the anti-IL2γ antibody of the present invention on human IL-4 (hIL-4) signaling, Ramos.2G6.4C10 / STAT3-Luc cells were plated at a density of 100,000 cells per well in a 96-well plate in growth medium (prepared according to the instructions of ATCC). The anti-IL2Rγ antibody or isotype control was serially diluted from 500 to 0.008 nM in assay buffer (plus samples containing only buffer without the test molecule), added to the cells and incubated for 20 minutes. After incubation, hIL-4 was added to the cells at a final concentration of 250 pM or 200 pM. Serial dilutions of hIL-4 (from 10 nM to 0.2 pM) (plus samples containing only buffer without the ligand) were added to the cells to determine dose-dependent activation. After overnight incubation at 37 °C in 5% CO2, luciferase activity was measured using OneGlo TM Reagent (Promega, #E6031) and Victor TM X Multilabel Plate Reader (Perkin Elmer).

[1194] The results were analyzed using nonlinear regression (4-parameter logistic) with Prism 5 software (GraphPad) to obtain EC 50and IC 50 Value. The inhibition percentage was calculated from the RLU value using the following equation:

[1195]

[1196] In this equation, "RLU 基线 " is the luminescence value from cells treated with a constant amount of ligand in the absence of antibody, "RLU 抑制 " is the minimum luminescence value from cells treated with a dose response of a specific antibody at a specific ligand concentration, and "RLU 背景 " is the luminescence value from cells not treated with any ligand or antibody.

[1197] Table 8-1. Inhibition of IL2Rγ signaling by 19 anti-IL2γ antibodies in bioassays using NK-92 / hIL7R / STAT3-Luc and Ramos.2G6.4C10 / STAT3-Luc cells

[1198]

[1199] NB: No blockade

[1200] WB: Weak blockade

[1201] Table 8-2. Inhibition of IL2Rγ signaling by four anti-IL27 antibodies in bioassays using NK-92 / hlL7R / STAT3-Luc and Ramos.2G6.4C10 / STAT3-Luc cells

[1202]

[1203] The ability of 23 anti-IL2γ antibodies of the present invention to inhibit the signaling of IL2Rγ family cytokines was tested using a bioassay. As shown in Table 8-1, 19 out of 23 anti-IL2γ antibodies inhibited the activation of IL2Rγ to varying degrees, and as shown in Table 8-2, four out of 23 anti-IL2γ antibodies showed no inhibition of IL2Rγ activation by the ligand.

[1204] Example 9 : Cell binding assays of NK-92, Jurkat, NIH / 3T3, MC / 9, and HEK293 cells by flow cytometry

[1205] To evaluate the binding of anti-IL2Rγ antibodies to human and murine IL-2Rγ expressed on cells, flow cytometry analysis was performed on cell lines endogenously expressing IL-2Rγ: NK-92 (human natural killer cell line), Jurkat (human T lymphocyte cell line), and MC / 9 (murine mast cell line) cells. The NIH / 3T3 (murine fibroblast) and HEK293 (human embryonic kidney) cell lines were included as negative controls.

[1206] For flow cytometry analysis, cells were pre-incubated with mouse IgG at 100 μg / ml for 15 minutes at room temperature (RT) to block antibody binding to Fc receptors. The anti-IL2Rγ antibodies of the present invention and isotype control antibodies were used at 10 μg / ml, with 0.5 to 1 x 10 6 cells / well, in PBS without calcium and magnesium containing 1% FBS (for Jurkat, NIH / 3T3, and HEK293 cells) or in growth medium (prepared according to the ATCC instructions) (for NK-92 and MC / 9) for 30 to 45 minutes at room temperature. The cells were washed and incubated with an anti-human antibody conjugated to allophycocyanin (APC) (Jackson ImmunoResearch, #109-136-170) on ice for 30 minutes. The cells were washed, fixed using BD CytoFix TM (BD biosciences, #554655), and analyzed on a flow cytometer or Accuri flow cytometer (BD). For all cell lines, controls of unstained and secondary antibody alone were also included. The results were analyzed using software to determine the geometric mean fluorescence intensity (MFI) of live cells. The binding ratio was calculated by normalizing the MFI of the test sample with the MFI of the unstained sample.

[1207] As shown in Table 9-1, 19 out of 23 anti-IL2Rγ antibodies of the present invention tested at 10 μg / ml showed binding to Jurkat and NK-92 cells, with binding ratios of 1-19 and 1-94, respectively. The anti-IL2Rγ antibodies showed binding to NIH / 3T3 and MC / 9 cells, with binding ratios of 1-13 and 1, respectively. The human isotype control antibody REGN1945 and the secondary antibody alone control condition showed a binding ratio of 1-13 for all tested cell lines.

[1208] As shown in Table 9-2, among the 23 anti-IL2Rγ antibodies of the present invention tested at 10 μg / ml, 4 showed binding to NK-92 cells with a binding ratio of 1-37, and also bound to HEK293 cells with a binding ratio of 1-3. The human isotype control antibody REGN1945 and the secondary antibody only control condition showed a binding ratio of 1-2 to NK-92 and HEK293 cells.

[1209] Table 9-1. Flow cytometry analysis of the binding of 19 out of 23 anti-IL2Rγ antibodies to NIH / 3T3, MC / 9, Jurkat, and NK-92 cells

[1210]

[1211] Table 9-2. Flow cytometry analysis of the binding of four out of 23 anti-IL2Rγ antibodies to HEK293 and NK-92 cells

[1212]

[1213] Example 10 : In vivo immunosuppression experiment to evaluate the effect of anti-IL2Rγ antibody H4H12889P on immune cell populations in blood

[1214] Experimental procedure. Genetically modified (VG) background mice (C57BL / 6NTac (75%) / 129S6SvEvTac (25%)) from a Regeneron breeding colony, in which the endogenous IL2RG extracellular domain was replaced with the corresponding human sequence, were subcutaneously administered with the isotype control (REGN1945) or H4H12889P at a dose of 10 mg / kg or 25 mg / kg, with a frequency of twice a week for 3 weeks (a total of 6 doses), or not administered.

[1215] Table 10-1. Experimental drug administration and treatment protocols for each group of mice

[1216] Group Recipient strain n mAb treatment A <![CDATA[II2rg hu / hu > 8 No mAb B <![CDATA[II2rg hu / hu > 8 REGN1945 (isotype), 10 mg / kg C <![CDATA[II2rg hu / hu > 8 REGN1945 (isotype), 25 mg / kg D <![CDATA[II2rg hu / hu > 8 H4H12889P (anti-hIL2RG), 10 mg / kg E <![CDATA[II2rg hu / hu > 8 H4H12889P (anti-hIL2RG), 25 mg / kg

[1217] Analysis of immune cell populations in blood over a period of time by flow cytometry. Total immune cells, B cells, T cells, NK cells, and neutrophil counts in peripheral blood at different time points (once a week) were analyzed by flow cytometry to evaluate the effect of H4H12889P on the absolute numbers of these cell types. Briefly, at each time point, blood samples from mice were collected into Microtainer tubes with K2EDTA [BD #365974], and 30 - 75 μL of each blood sample was incubated in red blood cell lysis buffer [Sigma #R7757] for 5 minutes at room temperature to lyse red blood cells. If needed, a second round of lysis was performed. Then the cells were washed in DPBS [Gibco #14190 - 144] and stained with LIVE / DEAD TM Fixable Near - Infrared Dead Cell Stain [Invitrogen #L34962] for 20 minutes, washed again in DPBS, and then blocked with purified anti - mouse CD16 / CD32 (Fc Blocking Reagent) [Tonbo Biosciences, #70 - 0161 - M001] diluted 1:50 in MACS buffer [autoMACS Running Buffer; Miltenyi Biotec, #130 - 091 - 221]. Subsequently, the cells were stained for cell surface markers by adding a mixture of fluorescently labeled antibodies (described in Table 2) diluted in BD horizon brilliant staining buffer [BD #566349] to identify CD45 + cells, T cells, B cells, NK cells, and neutrophils. Finally, the samples were washed in MACS buffer, fixed in BDCytoFix [BD #554655] diluted 1:4 in DPBS, and then washed and resuspended in MACS buffer before acquisition. Sample data was acquired on a FACSymphony A5 analyzer using the HTS attachment [BD]. A fixed volume of each sample was run. Data analysis was performed using FlowJov10 software [Tree Star, OR]. CD45 + immune cells were defined as singlet, live cells, CD45 + ; within this population, T cells were further defined as CD3 + , B cells were further defined as CD3 - CD19 + , NK cells were further defined as CD3 - CD19 - NKp46 + , neutrophils were further defined as F4 / 80 - Ly6G +The absolute number of each cell type run through the analyzer, the volume of sample run, and the volume of blood initially stained are used to calculate the cell / μL blood count for each sample.

[1218] Table 10-2. Antibodies Used in Flow Cytometry Analysis

[1219] Antibody Fluorescent dye Manufacturer Final dilution NKp46 FITC ebioscience 1∶200 Ly6G BB700 BD 1∶100 F4 / 80 PE BD 1∶500 CD3 PE-Cy7 BD 1∶200 CD4 BV786 BD 1∶200 CD8a BUV395 BD 1∶200 CD19 BUV737 BD 1∶200 CD45 Alexa Fluor 700 BioLegend 1∶200

[1220] Analysis of serum therapeutic antibody levels over time by antigen capture ELISA. Serum levels of the IL2Rγ antibody or isotype control antibody were measured weekly using a human total IgG Platinum ELISA kit. Each antibody was serially diluted in a 0.5% solution of BSA in PBS to generate a standard curve of 1.56 to 100 ng / mL for H4H12889P and REGN1945. Absorbance was measured at 450 nm on a SpectraMax M5 microplate reader [Molecular Devices]. Data analysis was performed using Prism 8.1.2 [GraphPad].

[1221] Results Overview and Conclusions. Treatment with H4H12889P (10 mg / kg and 25 mg / kg) resulted in a significant decrease in the total CD45 + immune cells (inset (A) of Figure 9), NK cells (inset (B) of Figure 9), T cells (inset (C) of Figure 9), and B cells (inset (D) of Figure 9) in the blood, while the neutrophil count (inset (E) of Figure 9) was unaffected. After the 3-week dosing period, the serum concentration of H4H12889P decreased over time. This decrease in H4H12889P concentration was associated with a continuous increase in the number of total CD45 + immune cells (inset (A) of Figure 9), NK cells (inset (B) of Figure 9), T cells (inset (C) of Figure 9), and B cells (inset (D) of Figure 9). By the end of the study, all of these populations had recovered to levels similar to those observed prior to treatment and to levels observed in untreated or REGN1945 (isotype control)-treated mice.

[1222] Example 11 : In Vivo Skin Graft Rejection Model for Evaluating the Blocking Activity of the IL2Rγ Antibody H4H12889P

[1223] Experimental Procedures. BALB / cJ mice obtained from The Jackson Laboratory (Bar Harbor, ME) were used as skin graft donors, and MHC-mismatched genetically modified mice from the Regeneron breeding colony that had been engineered to replace the extracellular domain of endogenous IL2RG with the corresponding human sequence (VG) Background mice (C57BL / 6NTac (75%) / 129S6SvEvTac (25%)) were used as recipients. Skin grafts were taken from the tails of donor mice. The skin was dissected with forceps and punched with a biopsy punch with a diameter of 10 mm. VG mice (humanized for IL2Rγ) used as graft recipients were subcutaneously administered an isotype control (REGN1945) or H4H12889P at a dose of 25 mg / kg, twice a week, starting 3 weeks before transplantation and continuing until rejection, or no administration was performed. The shaved recipients at the surgical site were anesthetized with isoflurane via a nasal cone and an analgesic (buprenorphine sustained-release preparation) (ZooPharm) was administered. The shaved dorsal area was treated by applying povidone iodine and alcohol wipes. The graft bed was created transversely in the middle between the dorsal and ventral sides of the mouse by grasping the skin with forceps and then excising the skin using a sterile biopsy skin punch with a diameter of 10 mm. The graft was then placed on the graft bed and covered with an adhesive bandage, which was fixed to the skin with two sterile surgical staples. Aseptic technique was used throughout the procedure. Five days later, the bandage and staples were removed and then monitoring was carried out.

[1224] Table 11-1. Experimental drug administration and treatment protocols for mice in each group

[1225] Group Recipient strain n Donor strain Donor tissue mAb treatment A <![CDATA[II2rg hu / hu > 10 BALB / cJ Tail skin No mAb B <![CDATA[II2rg hu / hu > 10 BALB / cJ Tail skin REGN1945 (isotype) C <![CDATA[II2rg hu / hu > 10 BALB / cJ Tail skin H4H12889P (anti-hIL2Rγ)

[1226] The experimental design is shown in Figure 10 .

[1227] Monitoring of skin graft rejection. Monitoring of skin grafts included the following criteria: (1) Skin grafts that could not be properly vascularized were considered technical failures and excluded from the analysis. These grafts would show crusting and shrinkage hours after the bandage was removed. (2) "Crusting" and shrinkage of the graft at a later stage were used as indicators of graft rejection. The complete rejection time point was recorded as the first day when 100% of the transplanted tissue was necrotic ( Figure 12 ). The onset of rejection was recorded as the first day when signs of rejection (i.e., redness) appeared ( Figure 11 ). Significance was determined by the log-rank test (Mantel-Cox) with Bonferroni correction (adjusted p-value 0.005, K = 9).

[1228] Detection of donor-specific antibodies by flow cytometry. Blood samples were taken at the time point of day 56 after transplantation to evaluate the formation of donor-specific antibodies ( Figure 13 ).

[1229] CT26.WT ( CRL-2638 TM) Cells were cultured in tissue culture flasks until 80% confluent. The cells were washed with 1X DPBS and dissociated with TrypLE Express reagent (Gibco) by incubating at room temperature for 5 minutes and washing the flask with complete RPMI 1640 medium. The cells were then centrifuged (500g, 10 minutes) and resuspended in 1X DPBS with 1:50 diluted 4 μg / ml Fc blocker (Tonbo) at 5 million cells / ml for 15 minutes at room temperature. The suspension was plated at 250,000 cells / well (50 μL) in 384-well V-bottom plates.

[1230] Fifty microliters of serially diluted sample sera from transplanted mice and non-transplanted wild-type VG mice (C57BL / 6NTac (75%) / 129S6SvEvTac (25%)) and wild-type BALB / cJ mice obtained from The Jackson Laboratory were added to their respective wells and incubated at 37 °C for 45 minutes. After washing twice with MACS buffer (500g, 4 minutes), the cells were resuspended in 50 μL of LIVE / DEAD TM Fixable Blue Dead Cell Staining Kit (Invitrogen) at a total volume of 50 μL per well and incubated at room temperature for 15 minutes. After centrifuging at 500g for 4 minutes, the supernatant was discarded, the cells were resuspended in 25 μL of Fc blocker (Tonbo), and incubated at 4 °C for 15 minutes. Then 25 μL of 2X antibody mixture (Table 11-2) was added and incubated at 4 °C for 25 minutes. After centrifugation (500g, 4 minutes), the cells were washed in MACS TM buffer by adding 100 μL of MACS buffer to each well. The cells were fixed by resuspending them in 100 μL of Cytofix TM Fixation Buffer (BD) diluted 1:4 in 1X DPBS and incubated at 4 °C for 15 minutes. Then, after centrifuging and discarding the fixative, the samples were resuspended in MACS buffer. Cells were acquired on a BD Fortessa X-20. The acquired events were analyzed with FlowJo (BD). MFI was derived from cells that were doubly discriminated (FSC-H, FSC-A) and then negative for the Live / Dead dye. The results plotted were the median fluorescence intensity values at a 1 / 512 dilution of the sample sera.

[1231] Table 11-2. Antibodies used in the flow cytometry staining mixture

[1232] Antigen Conjugate Clone Supplier Dilution (1 / ) CD45 BV421 30-F11 BioLegend 200 IgG APC Poly4053 BioLegend 200 B220 BUV395 RA3-6B2 BD 200 IgG1 PE-Cy7 RMG1-1 BioLegend 200 IgM APC-Cy7 RMM-1 BioLegend 200 IgG2a FITC R19-15 BD 200 IgG2c FITC Goat polyclonal IgG Bio-Rad 200

[1233] Results summary and conclusions. In a skin transplantation model (BALB / cJ to VG mice), treatment with H4H12889P (anti-IL2Rγ Ab) delayed the onset of skin graft rejection and improved the overall survival of skin grafts. Treatment with H4H12889P also prevented the production of donor-specific antibodies in this transplantation model.

Claims

1. An isolated antibody or antigen-binding fragment thereof that specifically binds to interleukin-2 receptor gamma (IL2Rγ), wherein the antibody or antigen-binding fragment thereof comprises a heavy chain variable region (HCVR) and a light chain variable region (LCVR), the heavy chain variable region comprises three heavy chain complementarity determining regions CDR-H1, CDR-H2, and CDR-H3, the light chain variable region comprises three light chain complementarity determining regions CDR-L1, CDR-L2, and CDR-L3, wherein CDR-H1 comprises the amino acid sequence of SEQ ID NO:337, CDR-H2 comprises the amino acid sequence of SEQ ID NO:339, CDR-H3 comprises the amino acid sequence of SEQ ID NO:341, CDR-L1 comprises the amino acid sequence of SEQ ID NO:72, CDR-L2 comprises the amino acid sequence of SEQ ID NO:54, and CDR-L3 comprises the amino acid sequence of SEQ ID NO:

184.

2. The antibody or antigen-binding fragment thereof according to claim 1, which is an antibody.

3. The antibody or antigen-binding fragment thereof according to claim 2, wherein the antibody is a human antibody.

4. The antibody or antigen-binding fragment thereof according to claim 1, which is an antigen-binding fragment of an antibody.

5. The antibody or antigen-binding fragment thereof according to claim 1, wherein the HCVR comprises the amino acid sequence of SEQ ID NO:335 and / or the LCVR comprises the amino acid sequence of SEQ ID NO:

182.

6. The antibody or antigen-binding fragment thereof according to claim 1, wherein the HCVR comprises the amino acid sequence of SEQ ID NO:335 and the LCVR comprises the amino acid sequence of SEQ ID NO:

182.

7. The antibody or antigen-binding fragment thereof according to claim 1, which comprises a heavy chain containing the amino acid sequence of SEQ ID NO:343 and / or a light chain containing the amino acid sequence of SEQ ID NO:

188.

8. The antibody or antigen-binding fragment thereof according to claim 1, which comprises a heavy chain and a light chain, the heavy chain comprises the amino acid sequence of SEQ ID NO:343, and the light chain comprises the amino acid sequence of SEQ ID NO:

188.

9. The antibody or antigen-binding fragment thereof according to claim 1, wherein the HCVR comprises an amino acid sequence having at least 90% sequence identity with the amino acid sequence of SEQ ID NO:335; and / or the LCVR comprises an amino acid sequence having at least 90% sequence identity with the amino acid sequence of SEQ ID NO:

182.

10. The antibody or antigen-binding fragment thereof according to claim 1, comprising: a heavy chain comprising an amino acid sequence having at least 90% sequence identity to the amino acid sequence of SEQ ID NO: 343; and / or a light chain comprising an amino acid sequence having at least 90% sequence identity to the amino acid sequence of SEQ ID NO:

188.

11. The antibody or antigen-binding fragment thereof according to claim 1, which is multispecific.

12. A complex comprising the antibody or antigen-binding fragment thereof according to any one of claims 1 to 11, which binds to an IL2Rγ polypeptide or an antigenic fragment thereof.

13. A method for preparing the antibody or antigen-binding fragment thereof according to any one of claims 1 to 11, comprising: (a) Introduce one or more polynucleotides encoding the HCVR and LCVR of the antibody or antigen-binding fragment thereof into a host cell; (b) Culture the host cell under conditions favorable for the expression of the polynucleotide; and (c) Isolate the antibody or antigen-binding fragment thereof from the host cell and / or the culture medium in which the host cell was cultured.

14. The method according to claim 13, wherein the host cell is a Chinese hamster ovary cell.

15. An antibody or antigen-binding fragment thereof, which is a product of the method according to any one of claims 13 to 14.

16. A polynucleotide encoding the antibody or antigen-binding fragment thereof according to any one of claims 1 to 11.

17. A vector comprising the polynucleotide according to claim 16.

18. A host cell comprising the antibody or antigen-binding fragment thereof according to any one of claims 1 to 11, or the polynucleotide according to claim 16, or the vector according to claim 17.

19. A composition or kit comprising the antibody or antigen-binding fragment thereof according to any one of claims 1 to 11 and 15.

20. The composition or kit according to claim 19, further comprising an additional therapeutic agent.

21. A pharmaceutical formulation comprising the antibody or antigen-binding fragment thereof according to any one of claims 1 to 11 and 15 and a pharmaceutically acceptable carrier or excipient.

22. The pharmaceutical formulation according to claim 21, further comprising an additional therapeutic agent.

23. The composition or kit according to claim 20 or the formulation according to claim 22, wherein the additional therapeutic agent is an anti-inflammatory agent.

24. The composition or kit according to claim 20 or the preparation according to claim 22, wherein the additional therapeutic agent is one or more members selected from the following: infliximab, adalimumab, etanercept, golimumab, prednisolone, methylprednisolone, antithymocyte globulin, alemtuzumab, daclizumab, tacrolimus, cyclosporine, mycophenolate mofetil, sirolimus, pentostatin, mesenchymal stem cells, ibritumomab, denileukin and basiliximab.

25. The composition or kit according to claim 20 or the preparation according to claim 22, wherein the additional therapeutic agent is one or more members selected from the following: anti-TNFα antibody or binding protein and adrenocortical hormone.

26. A container or injection device comprising the antibody or antigen-binding fragment thereof according to any one of claims 1 to 11 and 15, or the composition defined by any one of claims 19, 20 and 23-25, or the preparation defined by any one of claims 21 to 25.

27. Use of the antibody or antigen-binding fragment thereof according to any one of claims 1 to 11 and 15 in the manufacture of a medicament for treating or preventing an IL2Rγ-mediated disease or disorder in a subject in need thereof, wherein the IL2Rγ-mediated disease or disorder is graft-versus-host disease or an autoimmune disease.

28. The use according to claim 27, wherein the autoimmune disease is rheumatoid arthritis, systemic lupus erythematosus or type I diabetes.

29. The use according to claim 27, wherein the antibody or antigen-binding fragment thereof is administered by subcutaneous, intravenous or intramuscular injection into the subject.

30. Use of the antibody or antigen-binding fragment thereof according to any one of claims 1 to 11 and 15 in the manufacture of a medicament for treating or preventing an IL2Rγ-mediated disease or disorder in a subject in need thereof, wherein the IL2Rγ-mediated disease or disorder is β-islet cell transplant rejection, skin transplant rejection, heart transplant rejection, lung transplant rejection, kidney transplant rejection or liver transplant rejection.

31. The use according to claim 30, wherein the antibody or antigen-binding fragment thereof is administered by subcutaneous, intravenous or intramuscular injection into the subject.

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