Antitryptase antibody, its composition, and its use

Abstract

To provide a composition containing an anti-tryptase antibody and a pharmaceutical composition thereof, and a method for using the composition.SOLUTION: Provided is an isolated antibody that binds to a human tryptase beta 1 or an antigen-binding fragment thereof, where the antibody includes the following six hypervariable regions (HVR): (a) HVR-H1 including an amino acid sequence of GYAIT; (b) HVR-H2 including an amino acid sequence of GISSAATTFYSSWAKS; (c) HVR-H3 including an amino acid sequence of DPRGYGAALDRLDL; (d) HVR-L1 including an amino acid sequence of QSIKSVYNNRLG; (e) HVR-L2 including an amino acid sequence of ETSILTS; and (f) HVR-L3 including an amino acid sequence of AGGFDRSGDTT.SELECTED DRAWING: None

Description

[Technical Field]

[0001] Cross-reference of related applications This application claims the benefit of U.S. Provisional Patent Application No. 62 / 457,722, filed on 10 February 2017, which is incorporated herein by reference in its entirety.

[0002] Sequence List This application includes a sequence listing, which is filed electronically in ASCII format and is incorporated herein by reference in its entirety. The ASCII copy, created on 9 February 2018, is named 50474-112WO2_Sequence_Listing_2.9.18_ST25 and is 108,967 bytes in size.

[0003] The present invention relates to an anti-tryptase antibody, a pharmaceutical composition, and a method of using the same. [Background technology]

[0004] Human tryptase beta is a trypsin-like serine protease abundant in mast cells and, to a lesser extent, in basophils. Human tryptase beta (with three subtypes: tryptase beta 1, tryptase beta 2, and tryptase beta 3), produced by the TPSAB1 and TPSB2 loci, is the dominant active tryptase produced by human mast cells. These two loci produce four tryptase isoforms: TPSAB1 produces tryptase alpha and tryptase beta 1, while TPSB2 produces tryptase beta 2 and tryptase beta 3. Tryptase alpha, as well as other isoforms such as tryptase gamma, tryptase delta, and tryptase epsilon, are mostly inactive.

[0005] Proteolytically treated, active tryptase beta is stored in the secretory granules of mast cells as a tetramer complexed with heparin. Degranulation of mast cells, which can be triggered by IgE-dependent stimulation (e.g., allergens) or non-IgE-dependent stimulation (e.g., substance P or active tryptase), results in the release of tryptase beta along with other granular enzymes and histamine. Previous studies have observed increased mast cell counts in bronchial smooth muscle and epithelium in asthma patients, as well as increased levels of tryptase beta in bronchoalveolar lavage fluid. In addition, tryptases have been suggested to contribute to airway-bronchoconstriction and airway hyperreactivity, and to play a role in idiopathic pulmonary fibrosis and extracellular matrix turnover, demonstrating airway remodeling processes.

[0006] Tryptase has been suggested to be involved in a variety of diseases and disorders, including asthma and other lung disorders, inflammatory disorders, autoimmune disorders, and fibrotic disorders, where there remains a need for improved treatments and therapeutic methods, including therapeutic anti-tryptase antagonists. Attempts have been made to develop small molecule tryptase inhibitors (see, e.g., Cairns, JA, 2005, Pulmonary Pharmacology & Therapeutics 18:55-66), however, to our knowledge, no biological tryptase antagonist therapy, particularly anti-tryptase antagonist antibodies, has been reported. [Overview of the project]

[0007] The present invention relates to an antitryptase antibody, a pharmaceutical composition thereof, and a method of using the same.

[0008] In one embodiment, the present invention is characterized by an isolated antibody or antigen-binding fragment thereof that binds to human tryptase beta 1, wherein the antibody comprises the following six hypervariable regions (HVRs): (a) HVR-H1 comprising the amino acid sequence of DYGMV (SEQ ID NO: 7), (b) HVR-H2 comprising the amino acid sequence of FISSGSSTVYYADTMKG (SEQ ID NO: 2), (c) HVR-H3 comprising the amino acid sequence of RNYDDWYFDV (SEQ ID NO: 8), (d) HVR-L1 comprising the amino acid sequence of SASSSVTYMY (SEQ ID NO: 4), (e) HVR-L2 comprising the amino acid sequence of RTSDLAS (SEQ ID NO: 5), and (f) HVR-L3 comprising the amino acid sequence of QHYHSYPLT (SEQ ID NO: 6). In some embodiments, the antibody is defined by six HVRs comprising the amino acid sequences of SEQ ID NOs: 7, 2, 8, 4, 5, and 6. In some embodiments, the antibody further comprises S43, P46, and W47 (Kabat numbering) in the light chain variable (VL) domain framework region L2 (FR-L2). In some embodiments, the antibody comprises (a) a heavy chain variable (VH) domain containing an amino sequence having at least 90%, at least 95%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO: 9, (b) a light chain variable (VL) domain containing an amino acid sequence having at least 90%, at least 95%, or at least 99% identity with the amino acid sequence of SEQ ID NO: 10, or (c) a VH domain as in (a) and a VL domain as in (b). In some embodiments, the antibody further comprises the following VH domain framework regions (FR): (a) FR-H1 containing the amino acid sequence of EVQLVESGGGLVQPGGSLRLSCAASGFTFS (SEQ ID NO: 11), (b) FR-H2 containing the amino acid sequence of WVRQAPGKGLEWVA (SEQ ID NO: 12), (c) FR-H3 containing the amino acid sequence of RFTISRDNSKNTLYLQMNSLRAEDTAVYYCTR (SEQ ID NO: 13), and (d) FR-H4 containing the amino acid sequence of WGQGTLVTVSS (SEQ ID NO: 14). In some embodiments, the VH domain contains the amino acid sequence of SEQ ID NO: 9.In some embodiments, the antibody further comprises the following VL domains FR: (a) FR-L1 containing the amino acid sequence DIQMTQSPSSLSASVGDRVTITC (SEQ ID NO: 15), (b) FR-L2 containing the amino acid sequence WYQQKPGKSPKPWIY (SEQ ID NO: 16), (c) FR-L3 containing the amino acid sequence GVPSRFSGSGSGTDFTLTISSLQPEDFATYYC (SEQ ID NO: 17), and (d) FR-L4 containing the amino acid sequence FGQGTKVEIK (SEQ ID NO: 18). In some embodiments, the VL domain contains the amino acid sequence of SEQ ID NO: 10. In some embodiments, the antibody comprises a heavy chain containing (a) the amino acid sequence of SEQ ID NO: 76 and a light chain containing (b) the amino acid sequence of SEQ ID NO: 77. In other embodiments, the antibody comprises a heavy chain containing (a) the amino acid sequence of SEQ ID NO: 78 and a light chain containing (b) the amino acid sequence of SEQ ID NO: 79.

[0009] In another embodiment, the present invention is characterized by an isolated antibody bound to human tryptase beta 1, or an antigen-binding fragment thereof, wherein the antibody comprises (a) a VH domain comprising an amino acid sequence having at least 90%, at least 95%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO: 9, and (b) a VL domain comprising an amino acid sequence having at least 90%, at least 95%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO: 10. In some embodiments, the antibody comprises a VH domain comprising the amino acid sequence of SEQ ID NO: 9 and a VL domain comprising the amino acid sequence of SEQ ID NO: 10. In some embodiments, the antibody comprises a heavy chain comprising (a) the amino acid sequence of SEQ ID NO: 76 and (b) a light chain comprising the amino acid sequence of SEQ ID NO: 77. In other embodiments, the antibody comprises a heavy chain comprising (a) the amino acid sequence of SEQ ID NO: 78 and (b) a light chain comprising the amino acid sequence of SEQ ID NO: 79.

[0010] In another embodiment, the present invention is characterized by an isolated antibody comprising (a) a heavy chain containing the amino acid sequence of SEQ ID NO: 76 and (b) a light chain containing the amino acid sequence of SEQ ID NO: 77.

[0011] In another embodiment, the present invention is characterized by an isolated antibody comprising (a) a heavy chain containing the amino acid sequence of SEQ ID NO: 78 and (b) a light chain containing the amino acid sequence of SEQ ID NO: 79.

[0012] In another embodiment, the present invention is characterized by isolated antibodies or antigen-binding fragments thereof that bind to human tryptase beta 1, the antibodies comprising the following six HVRs: (a) HVR-H1 comprising the amino acid sequence of DYGMV (SEQ ID NO: 7), (b) HVR-H2 comprising the amino acid sequence of FISSGSSTVYYADTMKG (SEQ ID NO: 2), (c) HVR-H3 comprising the amino acid sequence of RDNYDWYFDV (SEQ ID NO: 29), (d) HVR-L1 comprising the amino acid sequence of SASSSVTYMY (SEQ ID NO: 4), (e) HVR-L2 comprising the amino acid sequence of RTSDLAS (SEQ ID NO: 5), and (f) HVR-L3 comprising the amino acid sequence of QHYHSYPLT (SEQ ID NO: 6). In some embodiments, the antibody further comprises the following VH domain FR: (a) FR-H1 containing the amino acid sequence of EVKLVESGGGSVQPGGSRKLSCAASGFTFS (SEQ ID NO: 21), (b) FR-H2 containing the amino acid sequence of WVRQAPGKGLEWVA (SEQ ID NO: 22), (c) FR-H3 containing the amino acid sequence of RFTISRDNPKNTLFLQMSSLRSEDTAMYYCAR (SEQ ID NO: 23), and (d) FR-H4 containing the amino acid sequence of WGTGTTVTVSS (SEQ ID NO: 24). In some embodiments, the VH domain contains the amino acid sequence of SEQ ID NO: 19. In some embodiments, the antibody further comprises the following VL domains FR: (a) FR-L1 containing the amino acid sequence QIVLTQSPAIMSASPGEKVTISC (SEQ ID NO: 25), (b) FR-L2 containing the amino acid sequence WYQQKPGSSPKPWIY (SEQ ID NO: 26), (c) FR-L3 containing the amino acid sequence GVPARFSGSGSGTSYSLTISSMEAEDAATYYC (SEQ ID NO: 27), and (d) FR-L4 containing the amino acid sequence FGAGTKLELK (SEQ ID NO: 28). In some embodiments, the VL domain contains the amino acid sequence of SEQ ID NO: 20.

[0013] In another embodiment, the present invention is characterized by an isolated antibody or antigen-binding fragment thereof that binds to human tryptase beta 1, the antibody comprising (a) a VH domain having at least 90%, at least 95%, or at least 99% identity with the amino acid sequence of SEQ ID NO: 19, (b) a VL domain comprising the amino acid sequence of SEQ ID NO: 20, or (c) a VH domain as in (a) and a VL domain as in (b). In some embodiments, the antibody further comprises the following VH domains FR: (a) FR-H1 comprising the amino acid sequence of EVKLVESGGGSVQPGGSRKLSCAASGFTFS (SEQ ID NO: 21), (b) FR-H2 comprising the amino acid sequence of WVRQAPGKGLEWVA (SEQ ID NO: 22), (c) FR-H3 comprising the amino acid sequence of RFTISRDNPKNTLFLQMSSLRSEDTAMYYCAR (SEQ ID NO: 23), and (d) FR-H4 comprising the amino acid sequence of WGTGTTVTVSS (SEQ ID NO: 24). In some embodiments, the VH domain includes the amino acid sequence of SEQ ID NO: 19. In some embodiments, the antibody further comprises the following VL domains FR: (a) FR-L1 including the amino acid sequence of QIVLTQSPAIMSASPGEKVTISC (SEQ ID NO: 25), (b) FR-L2 including the amino acid sequence of WYQQKPGSSPKPWIY (SEQ ID NO: 26), (c) FR-L3 including the amino acid sequence of GVPARFSGSGSGTSYSLTISSMEAEDAATYYC (SEQ ID NO: 27), and (d) FR-L4 including the amino acid sequence of FGAGTKLELK (SEQ ID NO: 28). In some embodiments, the VL domain includes the amino acid sequence of SEQ ID NO: 20.

[0014] In another embodiment, the present invention is characterized by an isolated antibody that binds to human tryptase beta 1, or an antigen-binding fragment thereof, comprising (a) a VH domain comprising an amino acid sequence having at least 99% sequence identity with the amino acid sequence of SEQ ID NO: 19, and (b) a VL domain comprising an amino acid sequence having at least 99% sequence identity with the amino acid sequence of SEQ ID NO: 20.

[0015] In some embodiments of any of the above-described aspects, the antibody binds to an epitope on human tryptase beta 1 containing at least one, at least two, at least three, or all four residues selected from the group consisting of His51, Val80, Lys81, and Asp82 of SEQ ID NO: 71. In some embodiments, the antibody binds to an epitope on human tryptase beta 1 containing at least one, at least two, at least three, or all four residues selected from the group consisting of His51, Val80, Lys81, and Asp82 of SEQ ID NO: 71. In some embodiments, the antibody binds to an epitope on human tryptase beta 1 containing His51 of SEQ ID NO: 71 and at least one, at least two, or all three residues selected from the group consisting of Val80, Lys81, and Asp82. In some embodiments, the epitope on human tryptase beta 1 further comprises one or more amino acid residues selected from the group consisting of Gln67, Leu83, Ala84, Ala85, Arg87, Pro103, Val104, Ser105, Arg106, Glu128, Glu129, and Pro130 of SEQ ID NO: 71. In some embodiments, the epitope on human tryptase beta 1 comprises at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, at least ten, at least eleven, or all twelve amino acid residues selected from the group consisting of Gln67, Leu83, Ala84, Ala85, Arg87, Pro103, Val104, Ser105, Arg106, Glu128, Glu129, and Pro130 of SEQ ID NO: 71. In some embodiments, the epitopes on human tryptase beta 1 include His51, Gln67, Val80, Lys81, Asp82, Leu83, Ala84, Ala85, Arg87, Pro103, Val104, Ser105, Arg106, Glu128, Glu129, and Pro130 of SEQ ID NO: 71. In some embodiments, the epitopes are associated with human tryptase beta 1 monomers or tetramers. In some embodiments, the epitopes are determined by X-ray crystallography models.In some embodiments, the antibody can dissociate both the small interface and the large interface of the tetrameric human tryptase beta 1.

[0016] In another embodiment, the present invention is characterized by an isolated antibody or antigen-binding fragment thereof that binds to human tryptase beta 1, the antibody comprising the following six HVRs: (a) HVR-H1 comprising the amino acid sequence of GYAIT (SEQ ID NO: 30), (b) HVR-H2 comprising the amino acid sequence of GISSAATTFYSSWAKS (SEQ ID NO: 31), (c) HVR-H3 comprising the amino acid sequence of DPRGYGAALDRLDL (SEQ ID NO: 32), (d) HVR-L1 comprising the amino acid sequence of QSIKSVYNNRLG (SEQ ID NO: 33), (e) HVR-L2 comprising the amino acid sequence of ETSILTS (SEQ ID NO: 34), and (f) HVR-L3 comprising the amino acid sequence of AGGFDRSGDTT (SEQ ID NO: 35). In some embodiments, the antibody is defined by six HVRs comprising the amino acid sequences of SEQ ID NOs: 30, 31, 32, 33, 34, and 35. In some embodiments, the antibody further comprises Arg71 and Val78 (Kabat numbering) in the VH domain FR-H3. In some embodiments, the antibody further comprises the following VH domains FR: (a) FR-H1 containing the amino acid sequence of EVQLVESGPGLVKPSETLSLTCTVSRFSLI (SEQ ID NO: 38), (b) FR-H2 containing the amino acid sequence of WIRQPPGKGLEWIG (SEQ ID NO: 42), (c) FR-H3 containing the amino acid sequence of RVTISRDTSKNQVSLKLSSVTAADTAVYYCAR (SEQ ID NO: 43), and (d) FR-H4 containing the amino acid sequence of WGQGTLVTVSS (SEQ ID NO: 41). In some embodiments, the VH domain contains the amino acid sequence of SEQ ID NO: 36. In some embodiments, the antibody comprises the following VL domains FR: (a) FR-L1 containing the amino acid sequence DIQMTQSPSSLSASVGDRVTITC (SEQ ID NO: 64), (b) FR-L2 containing the amino acid sequence WYQQKPGKAPKLLIY (SEQ ID NO: 65), (c) FR-L3 containing the amino acid sequence GVPSRFSGSGSGTDFTLTISSLQPEDFATYYC (SEQ ID NO: 66), and (d) FR-L4 containing the amino acid sequence FGQGTKVEIK (SEQ ID NO: 63). In some embodiments, the VL domain contains the amino acid sequence of SEQ ID NO: 37.In some embodiments, the antibody comprises a heavy chain containing (a) the amino acid sequence of SEQ ID NO: 80 and (b) a light chain containing the amino acid sequence of SEQ ID NO: 81. In other embodiments, the antibody comprises a heavy chain containing (a) the amino acid sequence of SEQ ID NO: 82 and (b) a light chain containing the amino acid sequence of SEQ ID NO: 83.

[0017] In another embodiment, the present invention is characterized by an isolated antibody bound to human tryptase beta 1, or an antigen-binding fragment thereof, the antibody comprising (a) a VH domain comprising an amino acid sequence having at least 90%, at least 95%, or at least 99% sequence identity with any one of the amino acid sequences of SEQ ID NOs: 36, 47, 48, 49, 50, 51, and 52; (b) a VL domain comprising an amino acid sequence having at least 90%, at least 95%, or at least 99% identity with any one of the amino acid sequences of SEQ ID NOs: 37, 53, 58, or 59; or (c) a VH domain as in (a) and a VL domain as in (b). In some embodiments, the antibody further comprises the following VH domains FR: (a) FR-H1 containing the amino acid sequence of EVQLVESGPGLVKPSETLSLTCTVSRFSLI (SEQ ID NO: 38), (b) FR-H2 containing the amino acid sequence of WIRQPPGKGLEWIG (SEQ ID NO: 42), (c) FR-H3 containing the amino acid sequence of RVTISRDTSKNQVSLKLSSVTAADTAVYYCAR (SEQ ID NO: 43), and (d) FR-H4 containing the amino acid sequence of WGQGTLVTVSS (SEQ ID NO: 41). In some embodiments, the VH domain contains the amino acid sequence of SEQ ID NO: 36. In some embodiments, the antibody comprises the following VL domains FR: (a) FR-L1 containing the amino acid sequence DIQMTQSPSSLSASVGDRVTITC (SEQ ID NO: 64), (b) FR-L2 containing the amino acid sequence WYQQKPGKAPKLLIY (SEQ ID NO: 65), (c) FR-L3 containing the amino acid sequence GVPSRFSGSGSGTDFTLTISSLQPEDFATYYC (SEQ ID NO: 66), and (d) FR-L4 containing the amino acid sequence FGQGTKVEIK (SEQ ID NO: 63). In some embodiments, the VL domain contains the amino acid sequence of SEQ ID NO: 37. In some embodiments, the antibody comprises a heavy chain containing (a) the amino acid sequence of SEQ ID NO: 80 and a light chain containing the amino acid sequence of SEQ ID NO: 81. In other embodiments, the antibody comprises a heavy chain containing (a) the amino acid sequence of SEQ ID NO: 82 and a light chain containing the amino acid sequence of SEQ ID NO: 83.

[0018] In another embodiment, the present invention is characterized by an isolated antibody or an antigen-binding fragment thereof that binds to human tryptase, the antibody comprising (a) a VH domain comprising an amino acid sequence having at least 90%, at least 95%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO: 36, and (b) a VL domain comprising an amino acid sequence having at least 90%, at least 95%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO: 37. In some embodiments, the antibody comprises a VH domain comprising the amino acid sequence of SEQ ID NO: 36 and a VL domain comprising the amino acid sequence of SEQ ID NO: 37. In some embodiments, the antibody comprises a heavy chain comprising (a) the amino acid sequence of SEQ ID NO: 80 and (b) a light chain comprising the amino acid sequence of SEQ ID NO: 81. In other embodiments, the antibody comprises a heavy chain comprising (a) the amino acid sequence of SEQ ID NO: 82 and (b) a light chain comprising the amino acid sequence of SEQ ID NO: 83.

[0019] In another embodiment, the present invention is characterized by an isolated antibody comprising (a) a heavy chain containing the amino acid sequence of SEQ ID NO: 80 and (b) a light chain containing the amino acid sequence of SEQ ID NO: 81.

[0020] In another embodiment, the present invention is characterized by an isolated antibody comprising (a) a heavy chain containing the amino acid sequence of SEQ ID NO: 82 and (b) a light chain containing the amino acid sequence of SEQ ID NO: 83.

[0021] In another embodiment, the present invention is characterized by an isolated antibody or an antigen-binding fragment thereof that binds to human tryptase, the antibody comprising (a) a VH domain comprising an amino acid sequence having at least 90%, at least 95%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO: 52, and (b) a VL domain comprising an amino acid sequence having at least 90%, at least 95%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO: 53.

[0022] In some embodiments of any of the above-described aspects, the antibody binds to an epitope on human tryptase beta 1 containing at least one, at least two, or all three residues selected from the group consisting of Gln100, Leu101, and Leu102 of SEQ ID NO: 71. In some embodiments, the epitope on human tryptase beta 1 further contains one or more amino acid residues selected from the group consisting of Trp55, Gln67, Asp82, Leu83, Ala84, Arg87, Pro103, Val104, Ser105, Arg106, Glu126, Leu127, Glu128, and Glu129 of SEQ ID NO: 71. In some embodiments, the epitope on human tryptase beta 1 includes at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, at least ten, at least eleven, at least twelve, at least thirteen, or all fourteen amino acid residues selected from the group consisting of Trp55, Gln67, Asp82, Leu83, Ala84, Arg87, Pro103, Val104, Ser105, Arg106, Glu126, Leu127, Glu128, and Glu129 of SEQ ID NO: 71. In some embodiments, the epitopes include Gln35, Trp55, Gln67, Asp82, Leu83, Ala84, Arg87, Gln100, Leu101, Leu102, Pro103, Val104, Ser105, Arg106, Glu126, Leu127, Glu128, Glu129, and Arg216 of SEQ ID NO: 71. In some embodiments, the epitopes are associated with the human tryptase beta-1 monomer or tetramer. In some embodiments, the epitopes are associated with the human tryptase beta-1 tetramer, and the epitopes on human tryptase beta-1 further include one or both of Gln35 and Arg216 of SEQ ID NO: 71. In some embodiments, the epitopes are determined by an X-ray crystallography model. In some embodiments, the antibody can dissociate the small and / or large interfaces of human tryptase beta 1.

[0023] In some embodiments of any of the aforementioned aspects, the antibody further binds to cynomolgus monkey (cyno) tryptase. In some embodiments, the antibody further binds to human tryptase alpha. In some embodiments, the antibody further binds to human tryptase beta 2 or human tryptase beta 3. In some embodiments, the antibody binds to both human tryptase beta 2 and human tryptase beta 3.

[0024] In some embodiments of any of the aforementioned aspects, the antibody is Kn about 1 nM or less. D It binds to tryptase. In some embodiments, K D This is measured by a surface plasmon resonance (SPR) assay. In some embodiments, the antibody is approximately 120 pM to approximately 0.5 nM K D It then binds to tryptase. In some embodiments, the antibody is diluted to about 120 pM to about 300 pM K D It then binds to tryptase. In some embodiments, the antibody is used at a concentration of about 120 pM to about 200 pM of K. D It binds to tryptase. In some embodiments, the antibody is about 180 pM K D It binds to tryptase. In some embodiments, the antibody is about 400 pM K D It binds to tryptase. In some embodiments, the SPR assay is performed at 25°C. In some embodiments, K Dis measured, for example, using a BIACORE® SPR assay as described in section (A)(vii) of Example 1. In some embodiments, the SPR assay can use a BIAcore® T200 or equivalent instrument. In some embodiments, a BIAcore® Series S CM5 sensor chip (or equivalent sensor chip) is immobilized with a monoclonal mouse anti-human IgG (Fc) antibody, and the anti-tryptase antibody is sequentially captured on the flow cell. Serial 3-fold dilutions of His-tagged human tryptase beta1 monomer (SEQ ID NO: 128) are injected at a flow rate of 30 μl / min. Each sample is analyzed for 3 minutes of association and 10 minutes of dissociation. The assay is performed at 25°C. After each injection, the chip is regenerated using 3 M MgCl2. The binding response is corrected by subtracting the response units (RU) from a flow cell that captures an unrelated IgG of similar density. k on and k off The simultaneous fitting of the 1:1 Languir model of is used for kinetic analysis.

[0025] In some embodiments of any of the aforementioned aspects, the antibody can inhibit the enzymatic activity of human tryptase beta 1. In some embodiments, the antibody inhibits tryptase activity with an IC50 of about 2.5 nM or less, as determined by a human tryptase beta enzyme assay using synthetic peptide S-2288 as a substrate. In some embodiments, the antibody inhibits tryptase activity with an IC50 of about 550 pM to about 2.5 nM. In some embodiments, the antibody inhibits tryptase activity with an IC50 of about 500 pM to about 2 nM. In some embodiments, the antibody inhibits tryptase activity with an IC50 of about 550 nM to about 1.5 nM. In some embodiments, the antibody inhibits tryptase activity with an IC50 of about 500 pM to about 700 pM. In some embodiments, the inhibitory activity of the antibody is determined as described in Example 1(A)(viii)(a). In some embodiments, the final concentration of heparin in a human tryptase beta enzyme assay using synthetic peptide S-2288 is 66 μg / ml. In some embodiments, recombinant human tryptase beta-1 tetramer active enzyme is diluted to 0.75 nM in TNH buffer (200 mM Tris, 150 mM NaCl, 0.1 mg / mL heparin, 0.01% TRITON® X-100, pH 8.0) and combined 1:1 with anti-tryptase antibody (diluted in PBS) in a 384-well plate. The plate is incubated at ambient temperature for 1 hour with gentle agitation. Chromometric substrate S-2288 (Chromogenix, catalog number 82-0852-39), or an equivalent substrate, is diluted to 1200 μM in TNH buffer and added to the plate. In some embodiments, the final well concentrations are 400 μM S-2288, 0.25 nM recombinant human tryptase beta-1 tetramer, 66 μg / mL heparin, and 0.10–222 nM anti-tryptase antibody. The plate is incubated at ambient temperature for 40 minutes with gentle agitation, and then A 405 The IC50 of anti-tryptase antibodies is determined from the 4-parameter fit of their respective curves.

[0026] In some embodiments of any of the aforementioned models, the antibody can inhibit the enzymatic activity of human tryptase beta-1 at pH 6. Specifically, the inhibitory activity of the antibody can be determined at pH 6.

[0027] In some embodiments, the antibody can inhibit tryptase-mediated stimulation of bronchial smooth muscle cell proliferation and / or collagenous system contraction. In some embodiments, the antibody can inhibit mast cell histamine release. In some embodiments, the antibody can inhibit IgE-induced histamine release and / or tryptase-induced histamine release. In some embodiments, the antibody can inhibit cynomolgus monkey tryptase D1 as assessed by an active tryptase ELISA assay. In some embodiments, the antibody can inhibit tryptase activity in cynomolgus monkey bronchoalveolar lavage (BAL) or nasal absorption samples. In some embodiments, the antibody can dissociate tetrameric human tryptase beta 1. In some embodiments, the antibody can dissociate tetrameric human tryptase beta 1 when in monovalent format. In some embodiments, the monovalent format is the Fab format. In some embodiments, the antibody can dissociate tetrameric human tryptase beta 1 in the presence of heparin. In some embodiments, the antibody can dissociate tetrameric tryptasebeta in the presence of 66 μg / ml heparin.

[0028] In another embodiment, the present invention features an antibody that binds to the same epitope as any one of the antibodies described above. In some embodiments, whether antibodies bind to the same epitope or compete for binding to human tryptase beta 1 is determined by an epitope binning assay. In some embodiments, the epitope binning assay is the OCTET® epitope binning assay, as described in Section C of Example 3. In some embodiments, the human tryptase beta 1 monomer protein is biotinylated by reacting it with NHS-PEG4 biotin at a Lys residue. The biotinylated monomer is diluted to 5 μg / ml with a kinetic buffer (ForteBio, Inc.) and immobilized on a streptavidin sensor chip (ForteBio, Inc.). After the immobilization step, the human tryptase beta 1 immobilized sensor is saturated with a first antibody, diluted to 10-20 μg / ml, and then bound with a second antibody diluted to 2.5 μg / ml. In some embodiments, the epitope binning assay is performed at 30°C.

[0029] In another embodiment, the present invention features an antibody that competes with, cross-blocks, or is cross-blocked by, any one of the aforementioned antibodies for binding to human tryptase beta 1.

[0030] In some embodiments of any of the aforementioned aspects, the antibody is monoclonal, human, humanized, or chimeric. In some embodiments, the antibody is humanized.

[0031] In some embodiments of any of the embodiments described above, the antibody is an antibody fragment that binds to tryptase. In some embodiments, the antibody fragment is selected from the group consisting of Fab, Fab'-SH, Fv, scFv, and (Fab')2 fragments.

[0032] In some embodiments of the aforementioned models, the antibody is a full-length antibody. In some embodiments, the antibody is an IgG antibody. In some embodiments, the IgG antibody is an IgG1 antibody. In some embodiments, the IgG antibody is an IgG4 antibody. In some embodiments, the IgG4 antibody contains a mutation in the hinge region. In some embodiments, the mutation is a substitution mutation. In some embodiments, the substitution mutation is at amino acid residue S228 (EU numbering). In some embodiments, the IgG4 antibody contains an S228P mutation (EU numbering).

[0033] In some embodiments of any of the above-described aspects, the antibody is a monospecific antibody.

[0034] In some embodiments of any of the aforementioned embodiments, the antibody is a multispecific antibody. In some embodiments, the antibody is a bispecific antibody. In some embodiments, the antibody comprises a first binding domain that binds to tryptase and a second binding domain that binds to a second biological molecule, the second biological molecule being selected from the group consisting of interleukin-13 (IL-13), interleukin-4 (IL-4), interleukin-5 (IL-5), interleukin-17 (IL-17), IgE, and interleukin-33 (IL-33). In some embodiments, the second biological molecule is IL-13. In some embodiments, the second biological molecule is IL-33. In some embodiments, the second biological molecule is IgE.

[0035] In another embodiment, the present invention features an isolated nucleic acid encoding any of the antibodies described herein, or a set of isolated nucleic acids encoding an antibody together.

[0036] In another embodiment, the present invention features an isolated nucleic acid encoding an antibody comprising a VH domain comprising the amino acid sequence of SEQ ID NO: 9 and / or a VL domain comprising the amino acid sequence of SEQ ID NO: 10, or a set of isolated nucleic acids encoding an antibody together, wherein the nucleic acid comprises a sequence that is at least 85%, at least 90%, at least 95%, or at least 99% identical to the sequence of SEQ ID NO: 104 and / or SEQ ID NO: 105. In some embodiments, the antibody comprises (a) a heavy chain comprising the amino acid sequence of SEQ ID NO: 76 and / or (b) a light chain comprising the amino acid sequence of SEQ ID NO: 77, wherein the nucleic acid or set comprises a sequence that is at least 85%, at least 90%, at least 95%, or at least 99% identical to the sequence of SEQ ID NO: 106 and / or SEQ ID NO: 107. In some embodiments, the antibody comprises a heavy chain containing (a) the amino acid sequence of SEQ ID NO: 78 and / or (b) a light chain containing the amino acid sequence of SEQ ID NO: 79, wherein the nucleic acid or set comprises a sequence that is at least 85%, at least 90%, at least 95%, or at least 99% identical to the sequence of SEQ ID NO: 108 and / or SEQ ID NO: 107. In some embodiments, the nucleic acid or set comprises the sequence of SEQ ID NO: 108 and / or SEQ ID NO: 107.

[0037] In another embodiment, the present invention features an isolated nucleic acid encoding an antibody comprising a VH domain comprising the amino acid sequence of SEQ ID NO: 36 and / or a VL domain comprising the amino acid sequence of SEQ ID NO: 37, or a set of isolated nucleic acids encoding an antibody together, wherein the nucleic acid comprises a sequence that is at least 85%, at least 90%, at least 95%, or at least 99% identical to the sequence of SEQ ID NO: 109 and / or SEQ ID NO: 110. In some embodiments, the antibody comprises (a) a heavy chain comprising the amino acid sequence of SEQ ID NO: 80 and / or (b) a light chain comprising the amino acid sequence of SEQ ID NO: 81, wherein the nucleic acid or set comprises a sequence that is at least 85%, at least 90%, at least 95%, or at least 99% identical to the sequence of SEQ ID NO: 111 and / or SEQ ID NO: 112. In some embodiments, the antibody comprises a heavy chain containing (a) the amino acid sequence of SEQ ID NO: 82 and / or (b) a light chain containing the amino acid sequence of SEQ ID NO: 83, wherein the nucleic acid or set comprises a sequence that is at least 85%, at least 90%, at least 95%, or at least 99% identical to the sequence of SEQ ID NO: 113 and / or SEQ ID NO: 112. In some embodiments, the nucleic acid or set comprises the sequence of SEQ ID NO: 113 and / or SEQ ID NO: 112.

[0038] In another embodiment, the present invention features a vector (e.g., an expression vector) or a set of vectors comprising any of the isolated nucleic acids or sets of isolated nucleic acids described herein. In another embodiment, the present invention features a host cell comprising the aforementioned nucleic acids and / or vectors and / or sets of nucleic acids and / or sets of vectors. In some embodiments, the host cell is a mammalian cell. In some embodiments, the mammalian cell is a Chinese hamster ovary (CHO) cell. In some embodiments, the host cell is a prokaryotic cell. In some embodiments, the prokaryotic cell is E. coli.

[0039] In another embodiment, the present invention features a method for producing any of the antibodies described herein, comprising culturing a host cell containing any of the aforementioned vectors (e.g., expression vectors) or sets of vectors in a culture medium under suitable conditions that enable antibody production. In some embodiments, the method further comprises recovering the antibody from the host cell or culture medium.

[0040] In another embodiment, the present invention features a composition (e.g., a pharmaceutical composition) comprising any one of the aforementioned antibodies. In some embodiments, the composition further comprises a pharmaceutically acceptable carrier, excipient, or diluent.

[0041] In another embodiment, the present invention is characterized by a pharmaceutical composition comprising an isolated monoclonal antibody conjugated to human tryptase beta 1, or an antigen-binding fragment thereof, and a pharmaceutically acceptable carrier, excipient, or diluent, wherein the antibody, as determined by an in vitro tryptase enzyme assay using S-2288 as a substrate, has a K content of about 0.1 nM to about 1 nM. D The antibody binds to monomeric tryptase beta-1 and / or inhibits the enzymatic activity of tryptase at half-inhibitory inhibitory concentrations (IC50) of approximately 0.1 nM to approximately 5 nM.

[0042] In some embodiments of the aforementioned model, the antibody has a K content of about 0.5 nM to about 1 nM. D It binds to tryptase. In some embodiments, the antibody is about 0.1 nM to about 0.5 nM of K D It binds to tryptase. In some embodiments, the antibody is about 0.4 nM K D It binds to tryptase. In some embodiments, the antibody is about 0.2 nM K D It binds to tryptase. In some embodiments, K D This is measured by a surface plasmon resonance (SPR) assay. In some embodiments, the SPR assay is performed at 25°C. In some embodiments, K DThis is measured using the BIACORE® SPR assay, for example, as described in Section (A)(vii) of Example 1. In some embodiments, the SPR assay can be performed using a BIAcore® T200 or equivalent instrument. In some embodiments, a BIAcore® Series S CM5 sensor chip (or equivalent sensor chip) is immobilized with monoclonal mouse anti-human IgG (Fc) antibody, and the anti-tryptase antibody is sequentially captured on a flow cell. A series of 3-fold dilutions of His-tagged human tryptase beta-1 monomer (SEQ ID NO: 128) are injected at a flow rate of 30 μl / min. Each sample is analyzed by 3-minute association and 10-minute dissociation. The assay is performed at 25°C. After each injection, the chip is regenerated using 3M MgCl2. The binding response is corrected by subtracting the response unit (RU) from a flow cell capturing unrelated IgG of similar density. on and k off A 1:1 Languir model with simultaneous fitting is used for dynamic analysis.

[0043] In some embodiments of the above-described model, the antibody can inhibit tryptase activity with an IC50 of about 0.5 nM to about 5 nM. In some embodiments, the antibody can inhibit tryptase activity with an IC50 of about 0.1 nM to about 2 nM. In some embodiments, the antibody can inhibit human tryptase activity with an IC50 of about 4 nM. In some embodiments, the antibody can inhibit tryptase activity with an IC50 of about 0.6 nM. In some embodiments, the antibody can inhibit tryptase activity at pH 6 in an in vitro tryptase enzyme assay using S-2288 as a substrate. In some embodiments, the inhibitory activity of the antibody is determined as described in the examples (e.g., Example 1, Section (A)(viii)(a)). In some embodiments, recombinant human tryptase beta-1 tetramer active enzyme is diluted to 0.75 nM in TNH buffer (200 mM Tris, 150 mM NaCl, 0.1 mg / mL heparin, 0.01% TRITON® X-100, pH 8.0) and combined in a 1:1 ratio with anti-tryptase antibody (diluted in PBS) in a 384-well plate. The plate is incubated at ambient temperature for 1 hour with gentle agitation. Chromometric substrate S-2288 (Chromogenix, catalog number 82-0852-39), or an equivalent substrate, is diluted to 1200 μM in TNH buffer and added to the plate. In some embodiments, the final well concentrations are 400 μM S-2288, 0.25 nM recombinant human tryptase beta-1 tetramer, 66 μg / mL heparin, and 0.10–222 nM anti-tryptase antibody. The plate is incubated at ambient temperature for 40 minutes with gentle agitation, and then A 405 The IC50 of the anti-tryptase antibodies is determined from the 4-parameter fit of their respective curves. In some embodiments, the final concentration of heparin in the human tryptase beta enzyme assay using synthetic peptide S-2288 is 66 μg / ml.

[0044] In some embodiments of the aforementioned aspects, the antibody can inhibit tryptase-mediated stimulation of bronchial smooth muscle cell proliferation and / or collagenous system contraction. In some embodiments, the antibody can inhibit mast cell histamine release. In some embodiments, the antibody can inhibit IgE-induced histamine release and / or tryptase-induced histamine release. In some embodiments, the antibody can inhibit tryptase activity in cynomolgus monkey bronchoalveolar lavage (BAL) or nasal absorption samples. In some embodiments, the antibody can dissociate tetrameric human tryptase beta 1. In some embodiments, the antibody can dissociate tetrameric human tryptase beta 1 when in monovalent format. In some embodiments, the monovalent format is Fab format. In some embodiments, the antibody can dissociate tetrameric human tryptase beta 1 in the presence of heparin. In some embodiments, the antibody can dissociate tetrameric tryptase beta in the presence of 66 μg / ml heparin.

[0045] In some embodiments of the above-described model, the antibody comprises the following six hypervariable regions (HVRs): (a) HVR-H1 containing the amino acid sequence of DYGMV (SEQ ID NO: 7), (b) HVR-H2 containing the amino acid sequence of FISSGSSTVYYADTMKG (SEQ ID NO: 2), (c) HVR-H3 containing the amino acid sequence of RNYDDWYFDV (SEQ ID NO: 8), (d) HVR-L1 containing the amino acid sequence of SASSSVTYMY (SEQ ID NO: 4), (e) HVR-L2 containing the amino acid sequence of RTSDLAS (SEQ ID NO: 5), and (f) HVR-L3 containing the amino acid sequence of QHYHSYPLT (SEQ ID NO: 6). In some embodiments, the antibody comprises (a) a heavy chain variable (VH) domain containing an amino acid sequence having at least 90%, at least 95%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO: 9, (b) a light chain variable (VL) domain containing an amino acid sequence having at least 90%, at least 95%, or at least 99% identity with the amino acid sequence of SEQ ID NO: 10, or (c) a VH domain as in (a) and a VL domain as in (b). In some embodiments, the antibody further comprises the following VH domains FR: (a) FR-H1 containing the amino acid sequence of EVQLVESGGGLVQPGGSLRLSCAASGFTFS (SEQ ID NO: 11), (b) FR-H2 containing the amino acid sequence of WVRQAPGKGLEWVA (SEQ ID NO: 12), (c) FR-H3 containing the amino acid sequence of RFTISRDNSKNTLYLQMNSLRAEDTAVYYCTR (SEQ ID NO: 13), and (d) FR-H4 containing the amino acid sequence of WGQGTLVTVSS (SEQ ID NO: 14). In some embodiments, the VH domain of the antibody contains the amino acid sequence of SEQ ID NO: 9. In some embodiments, the antibody further comprises the following VL domains FR: (a) FR-L1 containing the amino acid sequence of DIQMTQSPSSLSASVGDRVTITC (SEQ ID NO: 15), (b) FR-L2 containing the amino acid sequence of WYQQKPGKSPKPWIY (SEQ ID NO: 16), (c) FR-L3 containing the amino acid sequence of GVPSRFSGSGSGTDFTLTISSLQPEDFATYYC (SEQ ID NO: 17), and (d) FR-L4 containing the amino acid sequence of FGQGTKVEIK (SEQ ID NO: 18).In some embodiments, the VL domain of the antibody contains the amino acid sequence of SEQ ID NO: 10. In some embodiments, the antibody contains (a) a heavy chain containing the amino acid sequence of SEQ ID NO: 76 and (b) a light chain containing the amino acid sequence of SEQ ID NO: 77. In some embodiments, the antibody contains (a) a heavy chain containing the amino acid sequence of SEQ ID NO: 78 and (b) a light chain containing the amino acid sequence of SEQ ID NO: 79. In some embodiments, the antibody binds to an epitope on human tryptase beta 1 containing at least one, at least two, at least three, or all four residues selected from the group consisting of His51, Val80, Lys81, and Asp82 of SEQ ID NO: 71. In some embodiments, the antibody binds to an epitope on human tryptase beta 1 containing His51 of SEQ ID NO: 71 and at least one, at least two, or all three residues selected from the group consisting of Val80, Lys81, and Asp82. In some embodiments, the epitope on human tryptase beta 1 further comprises one or more amino acid residues selected from the group consisting of Gln67, Leu83, Ala84, Ala85, Arg87, Pro103, Val104, Ser105, Arg106, Glu128, Glu129, and Pro130 of SEQ ID NO: 71. In some embodiments, the epitope on human tryptase beta 1 comprises at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, at least ten, at least eleven, or all twelve amino acid residues selected from the group consisting of Gln67, Leu83, Ala84, Ala85, Arg87, Pro103, Val104, Ser105, Arg106, Glu128, Glu129, and Pro130 of SEQ ID NO: 71. In some embodiments, the epitopes on human tryptase beta 1 include His51, Gln67, Val80, Lys81, Asp82, Leu83, Ala84, Ala85, Arg87, Pro103, Val104, Ser105, Arg106, Glu128, Glu129, and Pro130 of SEQ ID NO: 71. In some embodiments, the epitopes are associated with human tryptase beta 1 monomers or tetramers.In some embodiments, the epitope is determined by an X-ray crystallography model. In some embodiments, the antibody can dissociate both the small interface and the large interface of the tetrameric human tryptase beta 1.

[0046] In other embodiments of the aforementioned model, the antibody includes the following six HVRs: (a) HVR-H1 containing the amino acid sequence of GYAIT (SEQ ID NO: 30), (b) HVR-H2 containing the amino acid sequence of GISSAATTFYSSWAKS (SEQ ID NO: 31), (c) HVR-H3 containing the amino acid sequence of DPRGYGAALDRLDL (SEQ ID NO: 32), (d) HVR-L1 containing the amino acid sequence of QSIKSVYNNRLG (SEQ ID NO: 33), (e) HVR-L2 containing the amino acid sequence of ETSILTS (SEQ ID NO: 34), and (f) HVR-L3 containing the amino acid sequence of AGGFDRSGDTT (SEQ ID NO: 35). In some embodiments, the antibody comprises (a) a VH domain containing an amino acid sequence having at least 90%, at least 95%, or at least 99% sequence identity with any one of the amino acid sequences of SEQ ID NOs: 36, 47, 48, 49, 50, 51, and 52; (b) a VL domain containing an amino acid sequence having at least 90%, at least 95%, or at least 99% identity with any one of the amino acid sequences of SEQ ID NOs: 37, 53, 58, or 59; or (c) a VH domain as in (a) and a VL domain as in (b). In some embodiments, the antibody further comprises the following VH domains FR: (a) FR-H1 containing the amino acid sequence of EVQLVESGPGLVKPSETLSLTCTVSRFSLI (SEQ ID NO: 38), (b) FR-H2 containing the amino acid sequence of WIRQPPGKGLEWIG (SEQ ID NO: 42), (c) FR-H3 containing the amino acid sequence of RVTISRDTSKNQVSLKLSSVTAADTAVYYCAR (SEQ ID NO: 43), and (d) FR-H4 containing the amino acid sequence of WGQGTLVTVSS (SEQ ID NO: 41). In some embodiments, the VH domain of the antibody contains the amino acid sequence of SEQ ID NO: 36.In some embodiments, the antibody further comprises the following VL domains FR: (a) FR-L1 containing the amino acid sequence DIQMTQSPSSLSASVGDRVTITC (SEQ ID NO: 64), (b) FR-L2 containing the amino acid sequence WYQQKPGKAPKLLIY (SEQ ID NO: 65), (c) FR-L3 containing the amino acid sequence GVPSRFSGSGSGTDFTLTISSLQPEDFATYYC (SEQ ID NO: 66), and (d) FR-L4 containing the amino acid sequence FGQGTKVEIK (SEQ ID NO: 63). In some embodiments, the VL domain of the antibody contains the amino acid sequence of SEQ ID NO: 37. In some embodiments, the antibody comprises a heavy chain containing the amino acid sequence of (a) SEQ ID NO: 80 and a light chain containing the amino acid sequence of (b) SEQ ID NO: 81. In some embodiments, the antibody comprises a heavy chain containing the amino acid sequence of (a) SEQ ID NO: 82 and a light chain containing the amino acid sequence of (b) SEQ ID NO: 83. In some embodiments, the antibody binds to an epitope on human tryptase beta 1 containing at least one, at least two, or all three residues selected from the group consisting of Gln100, Leu101, and Leu102 of SEQ ID NO: 71. In some embodiments, the epitope on human tryptase beta 1 further contains one or more amino acid residues selected from the group consisting of Trp55, Gln67, Asp82, Leu83, Ala84, Arg87, Pro103, Val104, Ser105, Arg106, Glu126, Leu127, Glu128, and Glu129 of SEQ ID NO: 71. In some embodiments, the epitope on human tryptase beta 1 includes at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, at least ten, at least eleven, at least twelve, at least thirteen, or all fourteen amino acid residues selected from the group consisting of Trp55, Gln67, Asp82, Leu83, Ala84, Arg87, Pro103, Val104, Ser105, Arg106, Glu126, Leu127, Glu128, and Glu129 of SEQ ID NO: 71.In some embodiments, the epitopes include Gln35, Trp55, Gln67, Asp82, Leu83, Ala84, Arg87, Gln100, Leu101, Leu102, Pro103, Val104, Ser105, Arg106, Glu126, Leu127, Glu128, Glu129, and Arg216 of SEQ ID NO: 71. In some embodiments, the epitopes are associated with the human tryptase beta-1 monomer or tetramer. In some embodiments, the epitopes are associated with the human tryptase beta-1 tetramer, and the epitopes on human tryptase beta-1 further include one or both of Gln35 and Arg216 of SEQ ID NO: 71. In some embodiments, the epitopes are determined by an X-ray crystallography model. In some embodiments, the antibody can dissociate the small and / or large interfaces of human tryptase beta 1.

[0047] In another embodiment, the present invention features a composition (e.g., a pharmaceutical composition) comprising an isolated monoclonal antibody or its antigen-binding fragment, and a pharmaceutically acceptable carrier, excipient, or diluent, wherein the antibody binds to an epitope on human tryptase beta 1 comprising at least one, at least two, at least three, or at least four residues selected from the group consisting of His51, Val80, Lys81, and Asp82 of SEQ ID NO: 71. In some embodiments, the antibody binds to an epitope on human tryptase beta 1 comprising His51 of SEQ ID NO: 71 and at least one, at least two, or at least three residues selected from the group consisting of Val80, Lys81, and Asp82. In some embodiments, the epitope on human tryptase beta 1 further comprises one or more amino acid residues selected from the group consisting of Gln67, Leu83, Ala84, Ala85, Arg87, Pro103, Val104, Ser105, Arg106, Glu128, Glu129, and Pro130 of SEQ ID NO: 71. In some embodiments, the epitope on human tryptase beta 1 comprises at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, at least ten, at least eleven, or all twelve amino acid residues selected from the group consisting of Gln67, Leu83, Ala84, Ala85, Arg87, Pro103, Val104, Ser105, Arg106, Glu128, Glu129, and Pro130 of SEQ ID NO: 71. In some embodiments, the epitopes on human tryptase beta 1 include His51, Gln67, Val80, Lys81, Asp82, Leu83, Ala84, Ala85, Arg87, Pro103, Val104, Ser105, Arg106, Glu128, Glu129, and Pro130 of SEQ ID NO: 71. In some embodiments, the epitopes are associated with human tryptase beta 1 monomers or tetramers. In some embodiments, the epitopes are determined by X-ray crystallography models.In some embodiments, the antibody can dissociate both the small interface and the large interface of the tetrameric human tryptase beta 1. In some embodiments, the antibody comprises the following six hypervariable regions (HVRs): (a) HVR-H1 containing the amino acid sequence of DYGMV (SEQ ID NO: 7), (b) HVR-H2 containing the amino acid sequence of FISSGSSTVYYADTMKG (SEQ ID NO: 2), (c) HVR-H3 containing the amino acid sequence of RNYDDWYFDV (SEQ ID NO: 8), (d) HVR-L1 containing the amino acid sequence of SASSSVTYMY (SEQ ID NO: 4), (e) HVR-L2 containing the amino acid sequence of RTSDLAS (SEQ ID NO: 5), and (f) HVR-L3 containing the amino acid sequence of QHYHSYPLT (SEQ ID NO: 6). In some embodiments, the antibody comprises (a) a heavy chain variable (VH) domain containing an amino acid sequence having at least 90%, at least 95%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO: 9, (b) a light chain variable (VL) domain containing an amino acid sequence having at least 90%, at least 95%, or at least 99% identity with the amino acid sequence of SEQ ID NO: 10, or (c) a VH domain as in (a) and a VL domain as in (b). In some embodiments, the antibody further comprises the following VH domains FR: (a) FR-H1 containing the amino acid sequence of EVQLVESGGGLVQPGGSLRLSCAASGFTFS (SEQ ID NO: 11), (b) FR-H2 containing the amino acid sequence of WVRQAPGKGLEWVA (SEQ ID NO: 12), (c) FR-H3 containing the amino acid sequence of RFTISRDNSKNTLYLQMNSLRAEDTAVYYCTR (SEQ ID NO: 13), and (d) FR-H4 containing the amino acid sequence of WGQGTLVTVSS (SEQ ID NO: 14). In some embodiments, the VH domain of the antibody contains the amino acid sequence of SEQ ID NO: 9.In some embodiments, the antibody further comprises the following VL domains FR: (a) FR-L1 containing the amino acid sequence DIQMTQSPSSLSASVGDRVTITC (SEQ ID NO: 15), (b) FR-L2 containing the amino acid sequence WYQQKPGKSPKPWIY (SEQ ID NO: 16), (c) FR-L3 containing the amino acid sequence GVPSRFSGSGSGTDFTLTISSLQPEDFATYYC (SEQ ID NO: 17), and (d) FR-L4 containing the amino acid sequence FGQGTKVEIK (SEQ ID NO: 18). In some embodiments, the VL domain of the antibody contains the amino acid sequence of SEQ ID NO: 10. In some embodiments, the antibody comprises a heavy chain containing (a) the amino acid sequence of SEQ ID NO: 76 and a light chain containing the amino acid sequence of SEQ ID NO: 77. In some embodiments, the antibody comprises a heavy chain containing (a) the amino acid sequence of SEQ ID NO: 78 and a light chain containing the amino acid sequence of SEQ ID NO: 79.

[0048] In another embodiment, the present invention features a composition (e.g., a pharmaceutical composition) comprising an isolated monoclonal antibody or an antigen-binding fragment thereof that binds to human tryptase beta 1, and a pharmaceutically acceptable carrier, excipient, or diluent, wherein the antibody binds to an epitope on human tryptase beta 1 comprising at least one, at least two, or all three residues selected from the group consisting of Gln100, Leu101, and Leu102 of SEQ ID NO: 71. In some embodiments, the epitope on human tryptase beta 1 further comprises one or more amino acid residues selected from the group consisting of Trp55, Gln67, Asp82, Leu83, Ala84, Arg87, Pro103, Val104, Ser105, Arg106, Glu126, Leu127, Glu128, and Glu129 of SEQ ID NO: 71. In some embodiments, the epitope on human tryptase beta 1 includes at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, at least ten, at least eleven, at least twelve, at least thirteen, or all fourteen amino acid residues selected from the group consisting of Trp55, Gln67, Asp82, Leu83, Ala84, Arg87, Pro103, Val104, Ser105, Arg106, Glu126, Leu127, Glu128, and Glu129 of SEQ ID NO: 71. In some embodiments, the epitopes include Gln35, Trp55, Gln67, Asp82, Leu83, Ala84, Arg87, Gln100, Leu101, Leu102, Pro103, Val104, Ser105, Arg106, Glu126, Leu127, Glu128, Glu129, and Arg216 of SEQ ID NO: 71. In some embodiments, the epitopes are associated with the human tryptase beta-1 monomer or tetramer. In some embodiments, the epitopes are associated with the human tryptase beta-1 tetramer, and the epitopes on human tryptase beta-1 further include one or both of Gln35 and Arg216 of SEQ ID NO: 71. In some embodiments, the epitopes are determined by an X-ray crystallography model.In some embodiments, the antibody can dissociate the small and / or large interfaces of human tryptase beta 1. In some embodiments, the antibody comprises the following six HVRs: (a) HVR-H1 containing the amino acid sequence of GYAIT (SEQ ID NO: 30), (b) HVR-H2 containing the amino acid sequence of GISSAATTFYSSWAKS (SEQ ID NO: 31), (c) HVR-H3 containing the amino acid sequence of DPRGYGAALDRLDL (SEQ ID NO: 32), (d) HVR-L1 containing the amino acid sequence of QSIKSVYNNRLG (SEQ ID NO: 33), (e) HVR-L2 containing the amino acid sequence of ETSILTS (SEQ ID NO: 34), and (f) HVR-L3 containing the amino acid sequence of AGGFDRSGDTT (SEQ ID NO: 35). In some embodiments, the antibody comprises (a) a VH domain containing an amino acid sequence having at least 90%, at least 95%, or at least 99% sequence identity with any one of the amino acid sequences of SEQ ID NOs: 36, 47, 48, 49, 50, 51, and 52; (b) a VL domain containing an amino acid sequence having at least 90%, at least 95%, or at least 99% identity with any one of the amino acid sequences of SEQ ID NOs: 37, 53, 58, or 59; or (c) a VH domain as in (a) and a VL domain as in (b). In some embodiments, the antibody further comprises the following VH domains FR: (a) FR-H1 containing the amino acid sequence of EVQLVESGPGLVKPSETLSLTCTVSRFSLI (SEQ ID NO: 38), (b) FR-H2 containing the amino acid sequence of WIRQPPGKGLEWIG (SEQ ID NO: 42), (c) FR-H3 containing the amino acid sequence of RVTISRDTSKNQVSLKLSSVTAADTAVYYCAR (SEQ ID NO: 43), and (d) FR-H4 containing the amino acid sequence of WGQGTLVTVSS (SEQ ID NO: 41). In some embodiments, the VH domain of the antibody contains the amino acid sequence of SEQ ID NO: 36.In some embodiments, the antibody further comprises the following VL domains FR: (a) FR-L1 containing the amino acid sequence DIQMTQSPSSLSASVGDRVTITC (SEQ ID NO: 64), (b) FR-L2 containing the amino acid sequence WYQQKPGKAPKLLIY (SEQ ID NO: 65), (c) FR-L3 containing the amino acid sequence GVPSRFSGSGSGTDFTLTISSLQPEDFATYYC (SEQ ID NO: 66), and (d) FR-L4 containing the amino acid sequence FGQGTKVEIK (SEQ ID NO: 63). In some embodiments, the VL domain of the antibody contains the amino acid sequence of SEQ ID NO: 37. In some embodiments, the antibody comprises a heavy chain containing the amino acid sequence of (a) SEQ ID NO: 80 and a light chain containing the amino acid sequence of (b) SEQ ID NO: 81. In some embodiments, the antibody comprises a heavy chain containing the amino acid sequence of (a) SEQ ID NO: 82 and a light chain containing the amino acid sequence of (b) SEQ ID NO: 83.

[0049] In some embodiments, the antibody can further bind to human tryptase alpha, tryptase beta 2, tryptase beta 3, and / or cynomolgus tryptase D1.

[0050] In any of the aforementioned compositions (e.g., pharmaceutical compositions), the antibody may be monoclonal, human, humanized, or chimeric. In some embodiments, the antibody is humanized.

[0051] In any of the aforementioned compositions (e.g., pharmaceutical compositions), the composition may be intended for use in humans.

[0052] Any of the aforementioned compositions (e.g., pharmaceutical compositions) can be freeze-dried. In other embodiments, the aforementioned compositions (e.g., pharmaceutical compositions) may be liquids.

[0053] The aforementioned compositions (e.g., pharmaceutical compositions) and excipients may be antioxidants. In some embodiments, the composition comprises one or more antioxidants selected from the group consisting of N-acetyltryptophan, tryptophan, methionine, cysteine, glutathione, thiosorbitol, ascorbic acid, monothioglycerol, cyclodextrin, trolox (6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid), pyridoxine, mannitol, and metal chelating agents. In some embodiments, the composition comprises N-acetyltryptophan or methionine. In some embodiments, the composition comprises N-acetyltryptophan and methionine.

[0054] In another embodiment, the present invention is characterized by (i) a composition (e.g., a pharmaceutical composition) comprising an isolated antibody bound to human tryptase beta 1, or an antigen-binding fragment thereof, wherein the antibody comprises the following six hypervariable regions (HVRs): (a) HVR-H1 comprising the amino acid sequence of DYGMV (SEQ ID NO: 7), (b) HVR-H2 comprising the amino acid sequence of FISSGSSTVYYADTMKG (SEQ ID NO: 2), (c) HVR-H3 comprising the amino acid sequence of RNYDDWYFDV (SEQ ID NO: 8), (d) HVR-L1 comprising the amino acid sequence of SASSSVTYMY (SEQ ID NO: 4), (e) HVR-L2 comprising the amino acid sequence of RTSDLAS (SEQ ID NO: 5), and (f) HVR-L3 comprising the amino acid sequence of QHYHSYPLT (SEQ ID NO: 6), wherein the oxidation of tryptophan at position 6 of HVR-H3 (SEQ ID NO: 8) is 30% or less. In some embodiments, the oxidation of tryptophan at position 6 of HVR-H3 (SEQ ID NO: 8) is 28% or less, 25% or less, 20% or less, 15% or less, 10% or less, or 6% or less. In some embodiments, the oxidation of tryptophan at position 6 of HVR-H3 (SEQ ID NO: 8) is determined according to the AAPH stress test. In some embodiments, the oxidation of tryptophan at position 6 of HVR-H3 (SEQ ID NO: 8) is determined within one year from the initial production of the composition.

[0055] Any of the aforementioned compositions (e.g., pharmaceutical compositions) may contain N-acetyltryptophan at a concentration of about 0.1 mM to about 5 mM. In some embodiments, the concentration of N-acetyltryptophan is about 0.1 mM to about 1 mM. In some embodiments, the concentration of N-acetyltryptophan is about 0.3 mM. In some embodiments, the composition contains methionine at a concentration of about 1 mM to about 20 mM. In some embodiments, the concentration of methionine is about 1 mM to about 10 mM. In some embodiments, the concentration of methionine is about 5 mM.

[0056] In another embodiment, the present invention relates to (i) an isolated antibody or antigen-binding fragment thereof that binds to human tryptase, wherein the antibody comprises the following six hypervariable regions (HVRs): (a) HVR-H1 comprising the amino acid sequence of DYGMV (SEQ ID NO: 7), (b) HVR-H2 comprising the amino acid sequence of FISSGSSTVYYADTMKG (SEQ ID NO: 2), (c) HVR-H3 comprising the amino acid sequence of RNYDDWYFDV (SEQ ID NO: 8), and (d) SASSSVTYMY (SEQ ID NO: 8). The present invention is characterized by an isolated antibody or its antigen-binding fragment comprising (e) HVR-L1 containing the amino acid sequence of (sequence number 4), (e) HVR-L2 containing the amino acid sequence of RTSDLAS (sequence number 5), and (f) HVR-L3 containing the amino acid sequence of QHYHSYPLT (sequence number 6), and a composition (e.g., a pharmaceutical composition) comprising (ii) N-acetyltryptophan at a concentration of about 0.1 mM to about 1 mM, and (iii) methionine at a concentration of about 1 mM to about 10 mM.

[0057] In any of the aforementioned compositions (e.g., pharmaceutical compositions), the antibody concentration may range from about 1 mg / ml to about 250 mg / ml. In some embodiments, the antibody concentration is about 150 mg / ml.

[0058] Any of the aforementioned compositions (e.g., pharmaceutical compositions) may further comprise one or more additional excipients selected from the group consisting of stabilizers, buffers, surfactants, and tonicity agents. In some embodiments, the composition further comprises a buffer. In some embodiments, the buffer is arginine succinate and / or histidine succinate. In some embodiments, the buffer comprises arginine succinate and histidine succinate. In some embodiments, the concentration of arginine succinate is about 50 mM to about 500 mM. In some embodiments, the concentration of arginine succinate is about 100 mM to about 300 mM. In some embodiments, the concentration of arginine succinate is about 200 mM. In some embodiments, the concentration of histidine succinate is about 1 mM to about 50 mM. In some embodiments, the concentration of histidine succinate is about 15 mM to about 25 mM. In some embodiments, the concentration of histidine succinic acid is about 20 mM. In some embodiments, the composition further comprises a surfactant. In some embodiments, the surfactant is poloxamer 188 or polysorbate 20. In some embodiments, the surfactant is poloxamer 188. In some embodiments, the concentration of poloxamer 188 is about 0.005% to about 0.1%. In some embodiments, the concentration of poloxamer 188 is about 0.005% to about 0.05%. In some embodiments, the concentration of poloxamer 188 is about 0.02%. In some embodiments, the pH of the composition is about 4.5 to about 7.0. In some embodiments, the pH of the composition is about 4.5 to about 6.5. In some embodiments, the pH of the composition is about 5.5. In some embodiments, the composition is contained in a light-shielding container. In some embodiments, the composition is contained in a pre-filled syringe.

[0059] Any of the aforementioned compositions (e.g., pharmaceutical compositions) may further include an IL-13 axially coupled antagonist, an IL-5 axially coupled antagonist, an IL-33 axially coupled antagonist, an M1 prime antagonist, an IgE antagonist, a TRPA1 antagonist, a CRTH2 antagonist, a bronchodilator or asthma symptom controller, an immunomodulator, a corticosteroid, a Th2 pathway inhibitor, a tyrosine kinase inhibitor, or a phosphodiesterase inhibitor. In some embodiments, the IL-13 axially coupled antagonist is an anti-IL-13 antibody. In some embodiments, the anti-IL-13 antibody is levukizumab. In some embodiments, the IL-5 axially coupled antagonist is an IL-5 conjugated antagonist or an IL-5 receptor conjugated antagonist. In some embodiments, the IL-33 axially coupled antagonist is an IL-33 conjugated antagonist or an ST2 conjugated antagonist. In some embodiments, the IL-33 conjugated antagonist is an anti-IL-33 antibody. In some embodiments, the M1 prime antagonist is kilizumab. In some examples, the IgE antagonist is omalizumab (XOLAIR®).

[0060] Any of the aforementioned compositions (e.g., pharmaceutical compositions) may be formulated for administration to humans. In certain embodiments, the pharmaceutical composition comprises an antibody that does not contain a non-human constant region sequence. In certain embodiments, the pharmaceutical composition comprises an antibody that does not contain a non-human framework and a non-human constant region sequence. In certain embodiments, the pharmaceutical composition comprises an antibody that is a human antibody, a humanized antibody, or a chimeric antibody.

[0061] In some embodiments, any one of the aforementioned antibodies can be used as a pharmaceutical product.

[0062] In some embodiments, any one of the aforementioned antibodies may be used to treat a disorder. In some embodiments, the disorder is selected from the group consisting of lung disorders, autoimmune disorders, inflammatory disorders, fibrous disorders, granulocytic (neutrophilic or eosinophilic) disorders, monocytic disorders, lymphocytic disorders, or disorders associated with an increased number or distribution of normal or abnormal tissue resident cells (such as mast cells, macrophages, or lymphocytes) or stromal cells (such as fibroblasts, myofibroblasts, smooth muscle cells, epithelium, or endothelium). In some embodiments, the disorder is a lung disorder. In some embodiments, the lung disorder is selected from the group consisting of asthma, airway hyperreactivity, and chronic obstructive pulmonary disease (COPD). In some embodiments, the lung disorder is asthma. In some embodiments, the asthma is Th2 hyperasthma or Th2 hypoasthma. In some embodiments, the autoimmune disorder is selected from the group consisting of rheumatoid arthritis, psoriasis, eosinophilic esophagitis, inflammatory bowel disease (IBD), and Crohn's disease. In some embodiments, the inflammatory disorder is chronic idiopathic urticaria (CIU, also known as chronic idiopathic urticaria, CSU), anaphylaxis, anaphylactic shock, atopic dermatitis, or allergic rhinitis. In some embodiments, the fibrous disorder is idiopathic pulmonary fibrosis (IPF). In some embodiments, the disorder is related to an increased number or distribution of normal or abnormal tissue resident cells (such as mast cells, macrophages, or lymphocytes) or stromal cells (such as fibroblasts, myofibroblasts, smooth muscle cells, epithelium, or endothelium). In some embodiments, the disorder is mastocytosis. In some embodiments, the antibody is intended for use in combination with further therapeutic agents. In some embodiments, further therapeutic agents include IL-13 axially coupled antagonists, IL-5 axially coupled antagonists, IL-33 axially coupled antagonists, M1 prime antagonists, IgE antagonists, TRPA1 antagonists, CRTH2 antagonists, bronchodilators or asthma symptom controllers, immunomodulators, corticosteroids, Th2 pathway inhibitors, tyrosine kinase inhibitors, or phosphodiesterase inhibitors.In some embodiments, the IL-13 axis-conjugated antagonist is an anti-IL-13 antibody. In some embodiments, the anti-IL-13 antibody is levukizumab. In some embodiments, the IL-5 axis-conjugated antagonist is an IL-5 conjugated antagonist or an IL-5 receptor conjugated antagonist. In some embodiments, the IL-33 axis-conjugated antagonist is an IL-33 conjugated antagonist or an ST2 conjugated antagonist. In some embodiments, the IL-33 conjugated antagonist is an anti-IL-33 antibody. In some embodiments, the M1 prime antagonist is kirizumab. In some embodiments, the antibody is intended for subcutaneous, intravenous, intramuscular, topical, oral, percutaneous, intraperitoneal, intraorbital, implantable, inhaled, intraarachnoid, intravesical, intravesical, or intranasal administration. In some embodiments, the antibody is intended for subcutaneous administration. In some embodiments, the antibody is intended for use in human subjects.

[0063] In some embodiments, any one of the aforementioned compositions (e.g., pharmaceutical compositions) can be used as a pharmaceutical product.

[0064] In some embodiments, any of the aforementioned compositions (e.g., pharmaceutical compositions) may be used to treat a disorder. In some embodiments, the disorder is selected from the group consisting of lung disorders, autoimmune disorders, inflammatory disorders, fibrous disorders, granulocytic (neutrophilic or eosinophilic) disorders, monocytic disorders, lymphocytic disorders, or disorders related to an increased number or distribution of normal or abnormal tissue resident cells (such as mast cells, macrophages, or lymphocytes) or stromal cells (such as fibroblasts, myofibroblasts, smooth muscle cells, epithelium, or endothelium). In some embodiments, the disorder is a lung disorder. In some embodiments, the lung disorder is selected from the group consisting of asthma, airway hyperreactivity, and chronic obstructive pulmonary disease (COPD). In some embodiments, the lung disorder is asthma. In some embodiments, the asthma is Th2-hyperasthma or Th2-hypoasthma. In some embodiments, the autoimmune disorder is selected from the group consisting of rheumatoid arthritis, psoriasis, eosinophilic esophagitis, inflammatory bowel disease (IBD), and Crohn's disease. In some embodiments, the inflammatory disorder is chronic idiopathic urticaria (CIU or CSU), anaphylaxis, anaphylactic shock, atopic dermatitis, or allergic rhinitis. In some embodiments, the fibrous disorder is idiopathic pulmonary fibrosis (IPF). In some embodiments, the disorder is a disorder related to an increased number or distribution of normal or abnormal tissue resident cells (such as mast cells, macrophages, or lymphocytes) or stromal cells (such as fibroblasts, myofibroblasts, smooth muscle cells, epithelium, or endothelium). In some embodiments, the disorder is mastocytosis. In some embodiments, the composition (e.g., a pharmaceutical composition) is intended for use in combination with further therapeutic agents. In some embodiments, further therapeutic agents include IL-13 axially coupled antagonists, IL-5 axially coupled antagonists, IL-33 axially coupled antagonists, M1 prime antagonists, IgE antagonists, TRPA1 antagonists, CRTH2 antagonists, bronchodilators or asthma symptom controllers, immunomodulators, corticosteroids, Th2 pathway inhibitors, tyrosine kinase inhibitors, or phosphodiesterase inhibitors.In some embodiments, the IL-13 axis-binding antagonist is an anti-IL-13 antibody. In some embodiments, the anti-IL-13 antibody is levukizumab. In some embodiments, the IL-5 axis-binding antagonist is an IL-5-binding antagonist or an IL-5 receptor-binding antagonist. In some embodiments, the IL-33 axis-binding antagonist is an IL-33-binding antagonist or an ST2-binding antagonist. In some embodiments, the IL-33-binding antagonist is an anti-IL-33 antibody. In some embodiments, the M1 prime antagonist is kirizumab. In some embodiments, the composition (e.g., pharmaceutical composition) is intended for administration subcutaneously, intravenously, intramuscularly, topically, orally, percutaneously, intraperitoneally, intraorbitally, implantably, by inhalation, intraarachnoidally, intravesically, or nasally. In some embodiments, the composition (e.g., pharmaceutical composition) is administered subcutaneously. In some embodiments, the composition (e.g., a pharmaceutical composition) is intended for use in human subjects.

[0065] In some embodiments, any one of the aforementioned antibodies may be used in the manufacture of a pharmaceutical product for treating a disorder. In some embodiments, the disorder is selected from the group consisting of lung disorders, autoimmune disorders, inflammatory disorders, fibrous disorders, granulocytic (neutrophilic or eosinophilic) disorders, monocytic disorders, lymphocytic disorders, or disorders associated with an increased number or distribution of normal or abnormal tissue resident cells (such as mast cells, macrophages, or lymphocytes) or stromal cells (such as fibroblasts, myofibroblasts, smooth muscle cells, epithelium, or endothelium). In some embodiments, the disorder is a lung disorder. In some embodiments, the lung disorder is selected from the group consisting of asthma, airway hyperreactivity, and chronic obstructive pulmonary disease (COPD). In some embodiments, the lung disorder is asthma. In some embodiments, the asthma is Th2 hyperasthma or Th2 hypoasthma. In some embodiments, the autoimmune disorder is selected from the group consisting of rheumatoid arthritis, psoriasis, eosinophilic esophagitis, inflammatory bowel disease (IBD), and Crohn's disease. In some embodiments, the inflammatory disorder is chronic idiopathic urticaria (CIU or CSU), anaphylaxis, anaphylactic shock, atopic dermatitis, or allergic rhinitis. In some embodiments, the fibrous disorder is idiopathic pulmonary fibrosis (IPF). In some embodiments, the disorder is related to an increased number or distribution of normal or abnormal tissue resident cells (such as mast cells, macrophages, or lymphocytes) or stromal cells (such as fibroblasts, myofibroblasts, smooth muscle cells, epithelium, or endothelium). In some embodiments, the disorder is mastocytosis. In some embodiments, the drug is formulated for use in combination with further therapeutic agents. In some embodiments, further therapeutic agents include IL-13 axially coupled antagonists, IL-5 axially coupled antagonists, IL-33 axially coupled antagonists, M1 prime antagonists, IgE antagonists, TRPA1 antagonists, CRTH2 antagonists, bronchodilators or asthma symptom controllers, immunomodulators, corticosteroids, Th2 pathway inhibitors, tyrosine kinase inhibitors, or phosphodiesterase inhibitors. In some embodiments, the IL-13 axially coupled antagonist is an anti-IL-13 antibody.In some embodiments, the anti-IL-13 antibody is levukizumab. In some embodiments, the IL-5 axis-binding antagonist is an IL-5-binding antagonist or an IL-5 receptor-binding antagonist. In some embodiments, the IL-33 axis-binding antagonist is an IL-33-binding antagonist or an ST2-binding antagonist. In some embodiments, the IL-33-binding antagonist is an anti-IL-33 antibody. In some embodiments, the M1 prime antagonist is kirizumab. In some embodiments, the drug is formulated for subcutaneous, intravenous, intramuscular, topical, oral, transdermal, intraperitoneal, intraorbital, implantable, inhaled, intraarachnoid, intravesical, intravesical, or intranasal administration. In some embodiments, the drug is formulated for subcutaneous administration. In some embodiments, the drug is formulated for use in human subjects.

[0066] In some embodiments, any one of the aforementioned compositions (e.g., pharmaceutical compositions) may be used in the manufacture of a pharmaceutical product for the treatment of a disorder. In some embodiments, the disorder is selected from the group consisting of lung disorders, autoimmune disorders, inflammatory disorders, fibrous disorders, granulocytic (neutrophilic or eosinophilic) disorders, monocytic disorders, lymphocytic disorders, or disorders related to an increased number or distribution of normal or abnormal tissue resident cells (such as mast cells, macrophages, or lymphocytes) or stromal cells (such as fibroblasts, myofibroblasts, smooth muscle cells, epithelium, or endothelium). In some embodiments, the disorder is a lung disorder. In some embodiments, the lung disorder is selected from the group consisting of asthma, airway hyperreactivity, and chronic obstructive pulmonary disease (COPD). In some embodiments, the lung disorder is asthma. In some embodiments, the asthma is Th2-hyperasthma or Th2-hypoasthma. In some embodiments, the autoimmune disorder is selected from the group consisting of rheumatoid arthritis, psoriasis, eosinophilic esophagitis, inflammatory bowel disease (IBD), and Crohn's disease. In some embodiments, the inflammatory disorder is chronic idiopathic urticaria (CIU or CSU), anaphylaxis, anaphylactic shock, atopic dermatitis, or allergic rhinitis. In some embodiments, the fibrous disorder is idiopathic pulmonary fibrosis (IPF). In some embodiments, the disorder is a disorder related to an increased number or distribution of normal or abnormal tissue resident cells (such as mast cells, macrophages, or lymphocytes) or stromal cells (such as fibroblasts, myofibroblasts, smooth muscle cells, epithelium, or endothelium). In some embodiments, the disorder is mastocytosis. In some embodiments, the drug is formulated for use in combination with further therapeutic agents. In some embodiments, further therapeutic agents include IL-13 axially coupled antagonists, IL-5 axially coupled antagonists, IL-33 axially coupled antagonists, M1 prime antagonists, IgE antagonists, TRPA1 antagonists, CRTH2 antagonists, bronchodilators or asthma symptom controllers, immunomodulators, corticosteroids, Th2 pathway inhibitors, tyrosine kinase inhibitors, or phosphodiesterase inhibitors.In some embodiments, the IL-13 axis-binding antagonist is an anti-IL-13 antibody. In some embodiments, the anti-IL-13 antibody is levukizumab. In some embodiments, the IL-5 axis-binding antagonist is an IL-5-binding antagonist or an IL-5 receptor-binding antagonist. In some embodiments, the IL-33 axis-binding antagonist is an IL-33-binding antagonist or an ST2-binding antagonist. In some embodiments, the IL-33-binding antagonist is an anti-IL-33 antibody. In some embodiments, the M1 prime antagonist is kirizumab. In some embodiments, the drug is formulated for subcutaneous, intravenous, intramuscular, topical, oral, transdermal, intraperitoneal, intraorbital, implantable, inhaled, intraarachnoid, intravesical, intravesical, or nasal administration. In some embodiments, the drug is formulated for subcutaneous administration. In some embodiments, the drug is formulated for use in human subjects.

[0067] In another embodiment, the present invention is characterized by a method for treating a disorder in a subject in need thereof, the method comprising administering to the subject a therapeutically effective amount of any one of the aforementioned antibodies. In some embodiments, the disorder is selected from the group consisting of lung disorder, autoimmune deficiency, inflammatory disorder, fibrous disorder, granulocytic (neutrophilic or eosinophilic) disorder, monocytic disorder, lymphocytic disorder, or disorder related to an increased number or distribution of normal or abnormal tissue resident cells (such as mast cells, macrophages, or lymphocytes) or stromal cells (such as fibroblasts, myofibroblasts, smooth muscle cells, epithelium, or endothelium). In some embodiments, the disorder is lung disorder. In some embodiments, the lung disorder is selected from the group consisting of asthma, airway hyperreactivity, and chronic obstructive pulmonary disease (COPD). In some embodiments, the lung disorder is asthma. In some embodiments, the asthma is Th2-hyperasthma or Th2-hyperasthma. In some embodiments, the autoimmune disorder is selected from the group consisting of rheumatoid arthritis, psoriasis, eosinophilic esophagitis, inflammatory bowel disease (IBD), and Crohn's disease. In some embodiments, the inflammatory disorder is chronic idiopathic urticaria (CIU or CSU), anaphylaxis, anaphylactic shock, atopic dermatitis, or allergic rhinitis. In some embodiments, the fibrous disorder is idiopathic pulmonary fibrosis (IPF). In some embodiments, the disorder is a disorder related to an increased number or distribution of normal or abnormal tissue resident cells (such as mast cells, macrophages, or lymphocytes) or stromal cells (such as fibroblasts, myofibroblasts, smooth muscle cells, epithelium, or endothelium). In some embodiments, the disorder is mastocytosis. In some embodiments, the method further comprises administering a further therapeutic agent to the target. In some embodiments, further therapeutic agents include IL-13 axially coupled antagonists, IL-5 axially coupled antagonists, IL-33 axially coupled antagonists, M1 prime antagonists, IgE antagonists, TRPA1 antagonists, CRTH2 antagonists, bronchodilators or asthma symptom controllers, immunomodulators, corticosteroids, Th2 pathway inhibitors, tyrosine kinase inhibitors, or phosphodiesterase inhibitors.In some embodiments, the IL-13 axis-conjugated antagonist is an anti-IL-13 antibody. In some embodiments, the anti-IL-13 antibody is levukizumab. In some embodiments, the IL-5 axis-conjugated antagonist is an IL-5 conjugated antagonist or an IL-5 receptor conjugated antagonist. In some embodiments, the IL-33 axis-conjugated antagonist is an IL-33 conjugated antagonist or an ST2 conjugated antagonist. In some embodiments, the IL-33 conjugated antagonist is an anti-IL-33 antibody. In some embodiments, the M1 prime antagonist is kirizumab. In some embodiments, the IgE antagonist is omalizumab (Xolair®). In some embodiments, the antibody is administered subcutaneously, intravenously, intramuscularly, topically, orally, percutaneously, intraperitoneally, intraorbitally, implantably, by inhalation, intraarachnoidally, intravesically, or intranasally. In some embodiments, the subject is human.

[0068] In another embodiment, the present invention is characterized by a method for treating a disorder in a subject in need thereof, the method comprising administering to the subject a therapeutically effective amount of any one of the aforementioned compositions (e.g., pharmaceutical compositions). In some embodiments, the disorder is selected from the group consisting of lung disorders, autoimmune disorders, inflammatory disorders, fibrous disorders, granulocytic (neutrophilic or eosinophilic) disorders, monocytic disorders, lymphocytic disorders, or disorders related to an increased number or distribution of normal or abnormal tissue resident cells (such as mast cells, macrophages, or lymphocytes) or stromal cells (such as fibroblasts, myofibroblasts, smooth muscle cells, epithelium, or endothelium). In some embodiments, the disorder is a lung disorder. In some embodiments, the lung disorder is selected from the group consisting of asthma, airway hyperreactivity, and chronic obstructive pulmonary disease (COPD). In some embodiments, the lung disorder is asthma. In some embodiments, the asthma is Th2-hyperasthma or Th2-hypoasthma. In some embodiments, the autoimmune disorder is selected from the group consisting of rheumatoid arthritis, psoriasis, eosinophilic esophagitis, inflammatory bowel disease (IBD), and Crohn's disease. In some embodiments, the inflammatory disorder is chronic idiopathic urticaria (CIU or CSU), anaphylaxis, anaphylactic shock, atopic dermatitis, or allergic rhinitis. In some embodiments, the fibrous disorder is idiopathic pulmonary fibrosis (IPF). In some embodiments, the disorder is a disorder related to an increased number or distribution of normal or abnormal tissue resident cells (such as mast cells, macrophages, or lymphocytes) or stromal cells (such as fibroblasts, myofibroblasts, smooth muscle cells, epithelium, or endothelium). In some embodiments, the disorder is mastocytosis. In some embodiments, the method further comprises administering a further therapeutic agent to the target. In some embodiments, further therapeutic agents include IL-13 axially coupled antagonists, IL-5 axially coupled antagonists, IL-33 axially coupled antagonists, M1 prime antagonists, IgE antagonists, TRPA1 antagonists, CRTH2 antagonists, bronchodilators or asthma symptom controllers, immunomodulators, corticosteroids, Th2 pathway inhibitors, tyrosine kinase inhibitors, or phosphodiesterase inhibitors.In some embodiments, the IL-13 axis-conjugated antagonist is an anti-IL-13 antibody. In some embodiments, the anti-IL-13 antibody is levukizumab. In some embodiments, the IL-5 axis-conjugated antagonist is an IL-5 conjugated antagonist or an IL-5 receptor conjugated antagonist. In some embodiments, the IL-33 axis-conjugated antagonist is an IL-33 conjugated antagonist or an ST2 conjugated antagonist. In some embodiments, the IL-33 conjugated antagonist is an anti-IL-33 antibody. In some embodiments, the M1 prime antagonist is omalizumab (Xolair®). In some embodiments, the composition is administered subcutaneously, intravenously, intramuscularly, topically, orally, percutaneously, intraperitoneally, intraorbitally, implantably, by inhalation, intraarachnoidally, intravesically, or intranasally. In some embodiments, the subject is human. [Brief explanation of the drawing]

[0069] [Figure 1-1] This shows the sequence arrangement of the VH and VL domains of hu31A.v11 and huE104.v2, which represent the complementarity-determining region (CDR) according to Kabat, Chothia, and Contact notation. The hypervariable region (HVR) is underlined. [Figure 1-2] This shows the sequence arrangement of the VH and VL domains of hu31A.v11 and huE104.v2, which represent the complementarity-determining region (CDR) according to Kabat, Chothia, and Contact notation. The hypervariable region (HVR) is underlined. [Figure 2A] This is a series of graphs showing the results of inhibition analysis of hu31A.v11 and huE104.v2 IgG, as determined by human tryptase enzyme assay. Both antibodies completely inhibited tryptase enzyme activity. [Figure 2B]This graph shows the results of human early airway smooth muscle cell (SMC) proliferation (Figure 2B) and contraction (Figure 2C) assays. Addition of tryptase beta stimulated human early airway SMC proliferation, which was inhibited in a dose-dependent manner by the addition of anti-tryptase antibodies hu31A.v11 IgG4 or huE104.v2 IgG4 (Figure 2B). Addition of tryptase also stimulated human early airway SMC contraction, which was also inhibited by the addition of hu31A.v11 IgG4 and huE104.v2 IgG4 (Figure 2C). [Figure 2C] This graph shows the results of human early airway smooth muscle cell (SMC) proliferation (Figure 2B) and contraction (Figure 2C) assays. Addition of tryptase beta stimulated human early airway SMC proliferation, which was inhibited in a dose-dependent manner by the addition of anti-tryptase antibodies hu31A.v11 IgG4 or huE104.v2 IgG4 (Figure 2B). Addition of tryptase also stimulated human early airway SMC contraction, which was also inhibited by the addition of hu31A.v11 IgG4 and huE104.v2 IgG4 (Figure 2C). [Figure 2D] This graph shows the results of a mast cell degranulation assay in vitro, in which mast cells were stimulated by the addition of tryptase beta or anti-4-hydroxy-3-nitrophenylacetyl (NP) IgE and NP. Addition of tryptase resulted in histamine release, which was blocked by the addition of hu31A.v11 (Figure 2D). A catalytically inactive mutant tryptase (S195A) served as a control. Addition of IgE and NP also resulted in histamine release, which was inhibited (30–50%) by the addition of a tryptase small molecule inhibitor (SMI) or hu31A.v11 (Figure 2E). [Figure 2E]This graph shows the results of a mast cell degranulation assay in vitro, in which mast cells were stimulated by the addition of tryptase beta or anti-4-hydroxy-3-nitrophenylacetyl (NP) IgE and NP. Addition of tryptase resulted in histamine release, which was blocked by the addition of hu31A.v11 (Figure 2D). A catalytically inactive mutant tryptase (S195A) served as a control. Addition of IgE and NP also resulted in histamine release, which was inhibited (30–50%) by the addition of a tryptase small molecule inhibitor (SMI) or hu31A.v11 (Figure 2E). [Figure 3A] This graph shows the results of dissociation of human tryptase beta-1 tetramer by hu31A.v11 Fab, analyzed by size exclusion chromatography (SEC). Three tests were analyzed by SEC: Test 1 contained only WT tetramer tryptase, which yielded peak 1 with a retention time Tr=26 min and retention volume Vr=13 ml; Test 2 contained WT tetramer tryptase + Fab hu31A.v11 + heparin, which yielded peak 2 (Tr=27.6 min, Vr=13.8 ml) and peak 4 (Tr=31 min, Vr=15.5 ml); Test 3 contained WT tetramer tryptase + Fab hu31A.v11 without heparin, which yielded peak 3 (Tr=28.1 min, Vr=14 ml) and peak 4 (Tr=31 min, Vr=15.5 ml). [Figure 3B] This graph shows the results of dissociation of human tryptase beta-1 tetramer by huE104.v2 Fab. Three trials were analyzed by SEC: Trial 1 contained His-tagged monomeric tryptase + Fab huE104.v2, which yielded peak 2 (Tr=25.8 min) and peak 6 (31.6 min); Trial 2 contained WT tetramer tryptase + Fab huE104.v2, which yielded peak 3 (Tr=26 min) and peak 7 (Tr=31.8 min); Trial 3 contained WT tetramer tryptase + Fab huE104.v2 + heparin, which yielded peak 1 (Tr=21 min), peak 4 (Tr=27.2 min), and peak 5 (Tr=31.2 min). [Figure 4A] This is a series of graphs showing the results of pharmacokinetic (PK) simulations comparing the PK and neutralizing activity of a dissociated anti-tryptase antibody with that of a tryptase tetramer-specific stabilizing antibody at baseline tryptase levels (4 ng / ml serum, 10 ng / ml lung tissue, Figure 4A) or high tryptase levels (10 ng / ml serum, 40 ng / ml lung tissue, Figure 4B). [Figure 4B] This is a series of graphs showing the results of pharmacokinetic (PK) simulations comparing the PK and neutralizing activity of a dissociated anti-tryptase antibody with that of a tryptase tetramer-specific stabilizing antibody at baseline tryptase levels (4 ng / ml serum, 10 ng / ml lung tissue, Figure 4A) or high tryptase levels (10 ng / ml serum, 40 ng / ml lung tissue, Figure 4B). [Figure 4C] This is a series of graphs showing the results of neutralization activity simulations comparing a dissociated anti-tryptase antibody with a tryptase tetramer-specific stabilizing antibody at baseline or high tryptase levels. [Figure 5A] Images of Coomassie blue-stained sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) gels (top panel) are shown, indicating that human tryptase beta-1 cleaved fibrinogen into peptide fragments at both pH 6 (Figure 5A) and 7.5 (Figure 5B). Under experimental conditions of high heparin concentration, the anti-tryptase antibody hu31A.v11 Fab blocked fibrinogen cleavage at both pH 6 and pH 7.5, while huE102.v2 Fab did not. Lane 1 shows fibrinogen only, with uncleaved alpha, beta, and gamma chains of fibrinogen; Lane 2 shows fibrinogen and tryptase beta; Lane 3 shows fibrinogen, tryptase beta, and hu31A.v11 Fab; Lane 4 shows fibrinogen, tryptase beta, and B12 IgG; Lane 5 shows fibrinogen, tryptase beta 1, and huE104.v2 Fab; Lane 6 shows B12 mIgG1 only. The decrease in alpha chain strength indicates tryptase proteolytic activity, which was analyzed and quantified in the panel below. [Figure 5B]Images of Coomassie blue-stained sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) gels (top panel) are shown, indicating that human tryptase beta-1 cleaved fibrinogen into peptide fragments at both pH 6 (Figure 5A) and 7.5 (Figure 5B). Under experimental conditions of high heparin concentration, the anti-tryptase antibody hu31A.v11 Fab blocked fibrinogen cleavage at both pH 6 and pH 7.5, while huE102.v2 Fab did not. Lane 1 shows fibrinogen only, with uncleaved alpha, beta, and gamma chains of fibrinogen; Lane 2 shows fibrinogen and tryptase beta; Lane 3 shows fibrinogen, tryptase beta, and hu31A.v11 Fab; Lane 4 shows fibrinogen, tryptase beta, and B12 IgG; Lane 5 shows fibrinogen, tryptase beta 1, and huE104.v2 Fab; Lane 6 shows B12 mIgG1 only. The decrease in alpha chain strength indicates tryptase proteolytic activity, which was analyzed and quantified in the panel below. [Figure 6A] This graph shows the results of dissociation of WT or mutant tetramers by hu31A.v11 Fab. Four trials were analyzed by SEC: Trial 1 contained the WT tetramer + huE104.v1 Fab, which yielded a reference peak (Tr=21.6 min); Trial 2 contained the tetramer Y75C variant + hu31A.v11 Fab, which yielded peak 1 (Tr=25.6 min) and peak 4 (Tr=31.6 min); Trial 3 contained the tetramer I99C variant + hu31A.v11 Fab, which yielded peak 2 (Tr=23.9 min) and peak 4 (Tr=31.6 min); Trial 4 contained the WT tetramer + hu31A.v11 Fab, which yielded peak 3 (Tr=28.1 min) and peak 4 (Tr=31.6 min). [Figure 6B] Figure 6A shows the results of Coomassie blue stained SDS-PAGE gel analysis of the size exclusion chromatography peak. hu31A.v11 Fab formed a complex containing tryptase mutants Y75C and I99C, and the tetramer dissociated into a covalently bonded dimer. [Figure 6C]This graph shows the results of dissociation of WT or mutant tetramers by huE104.v2 Fab. Three tests were analyzed by SEC: Test 1 contained the tetramer Y75C variant + huE104.v2 Fab, which yielded peak 1 (Tr=21.6 min) and peak 4 (Tr=31 min); Test 2 contained the tetramer I99C variant + huE104.v2 Fab, which yielded peak 2 and peak 5 (Tr=31 min); Test 3 contained the WT tetramer + huE104.v2 Fab, which yielded peak 3 (Tr=26 min) and peak 6 (Tr=31.8 min). The results indicate that huE104.v2 Fab formed complexes with the tryptase mutants Y75C and I99C, and dissociated only the I99C large interface-locked tetramer into a covalently bonded dimer. Peaks 4, 5, and 6 all contain excessive Fab when determined by SDS-PAGD (data not shown). [Figure 7-1] The amino acid sequence of mature human tryptase beta 1 is shown, following the gene sequence numbering and the chymotrypsinogen numbering ("chymo-numb") system typically used for mammalian serinetrypsins. [Figure 7-2] The amino acid sequence of mature human tryptase beta 1 is shown, following the gene sequence numbering and the chymotrypsinogen numbering ("chymo-numb") system typically used for mammalian serinetrypsins. [Figure 8] This shows the binding epitope of Fab hu31A.v11 on human tryptase beta 1 (all tryptase residues are numbered by chymotrypsinogen). [Figure 9] This is a drawing showing a model of Fab hu31A.v11 on a tryptase tetramer. The tryptase monomer compounded with Fab hu31A.v11 is aligned with protomers A and C in the tryptase tetramer. The heavy and light chains are shown. Collisions between the light chains of Fab hu31A.v11 on adjacent tryptase protomers in this model are marked by dotted ellipses. [Figure 10]The conformational changes in the 60s loop of tryptase detected in the complex structure are shown. Val60c and Val90 (indicated by bars) create hydrophobic pockets for the binding of Tyr173d from the adjacent protomer as part of the protein-protein interaction at the large interface of the tetrameric tryptase. The tryptase protomer in the tetrameric structure is shown. The tryptase bound to Fab hu31A.v11 overlaps one of the protomers of the tetrameric complex. The conformation of Val60c changes when Fab hu31A.v11 is bound, creating steric hindrance which is expected to prevent Tyr173d from binding to its pocket (all tryptase residues are chymotrypsinogen numbered). [Figure 11] This shows the conformational change in the 30s loop of the tryptase located at the small interface detected in the complex structure after binding with hu31A.v11. [Figure 12A] This is a drawing of the crystal structure of the WT tryptase tetramer compounded with four huE104.v1 Fab molecules. The tryptase protomers are indicated by letter labeling or according to the protomer (see Pereira et al. Nature 392:306-311, 1998). [Figure 12B] This graph illustrates the effect of huE104.v1 binding at the small interface of the tryptase tetramer when evaluated by hydrogen-deuterium exchange (HDX). [Figure 13] The graphs show the results of pharmacokinetic (PK) analyses of humanized anti-tryptase antibodies huE104.v2 and hu31A.v11 administered intravenously (IV) at doses of 1 or 10 mg / kg to C57BL / 6 mice, respectively. The graphs show the concentration of the anti-tryptase antibody (μg / mL) as a function of time (days). The results are from three animals. [Figure 14]This graph shows the results of PK analysis of humanized anti-tryptase antibodies huE104.v2 and hu31A.v11 compared to a control anti-gD IgG4 antibody. Each antibody was administered intravenously to cynomolgus monkeys (cyno) at a dose of 30 mg / kg. The graph shows the concentration (μg / mL) of the anti-tryptase antibody as a function of time (days). [Figure 15-1] This is a schematic diagram of an assay for measuring the amount of active tryptase (left panel) and total tryptase (right panel) in a sample. Tryptase monomers and tetramers are shown. In the left panel, a soybean trypsin inhibitor (SBTI) is added. Next, an exemplary biotinylated activity reference probe (ABP) was added to the tryptase-containing sample (e.g., bronchoalveolar lavage fluid (BAL)) to label the active tryptase. Labeling was stopped by adding an exemplary small molecule inhibitor G02849855 (e.g., BMS-262084, Sutton et al., Bioorg. Med. Chem. Lett. 12:3229-33, 2002, Qian et al., J. Org. Chem. 2002, 67:3595-3600). The tryptase tetramer was dissociated by adding hu31A.v11 antibody. Next, labeled tryptase was detected by enzyme-linked immunosorbent assay (ELISA) using horseradish peroxidase complexed with streptavidin. In the whole tryptase assay, hu31A.v11 was added to the sample to dissociate the tryptase tetramer in the sample. The amount of tryptase was then determined using ELISA. [Figure 15-2]This is a schematic diagram of an assay for measuring the amount of active tryptase (left panel) and total tryptase (right panel) in a sample. Tryptase monomers and tetramers are shown. In the left panel, a soybean trypsin inhibitor (SBTI) is added. Next, an exemplary biotinylated activity reference probe (ABP) was added to the tryptase-containing sample (e.g., bronchoalveolar lavage fluid (BAL)) to label the active tryptase. Labeling was stopped by adding an exemplary small molecule inhibitor G02849855 (e.g., BMS-262084, Sutton et al., Bioorg. Med. Chem. Lett. 12:3229-33, 2002, Qian et al., J. Org. Chem. 2002, 67:3595-3600). The tryptase tetramer was dissociated by adding hu31A.v11 antibody. Next, labeled tryptase was detected by enzyme-linked immunosorbent assay (ELISA) using horseradish peroxidase complexed with streptavidin. In the whole tryptase assay, hu31A.v11 was added to the sample to dissociate the tryptase tetramer in the sample. The amount of tryptase was then determined using ELISA. [Figure 16] The results of an active tryptase assay performed on BAL samples obtained from cynomolgus monkeys administered 30 mg / kg of the anti-tryptase antibody hu31A.v11 intravenously (IV) are shown. The panel above shows a schematic diagram of the experimental protocol. [Figure 17] This is a schematic diagram showing the experimental protocol of the cynomolgus monkey roundworm challenge model described in Example 6. [Figure 18A] This graph shows the results of the active tryptase assay (Figure 18A) and the total tryptase assay (Figure 18B) performed on BAL obtained from individual animals in the cynomolgus monkey roundworm challenge experiment described in Example 6. [Figure 18B] This graph shows the results of the active tryptase assay (Figure 18A) and the total tryptase assay (Figure 18B) performed on BAL obtained from individual animals in the cynomolgus monkey roundworm challenge experiment described in Example 6. [Figure 18C]This graph shows the results of a total tryptase assay to determine the amount of total tryptase in the nasal mucosal lining fluid (MLF) obtained by nasal absorption using synthetic absorbable matrix (SAM) from individual animals in the cynomolgus monkey roundworm challenge experiment described in Example 6. [Figure 19] This graph shows that administration of the anti-tryptase antibody hu31A.v11 inhibited IgE-mediated passive systemic anaphylaxis in human-transplanted mice. In IgE challenge, mice treated with the anti-tryptase antibody hu31A.v11 showed improved thermoregulation compared to mice treated with a control anti-gD antibody. ***P<0.0001 (paired t-test). [Modes for carrying out the invention]

[0070] I. Definition As used herein, the term “approximately” refers to the normal range of error for each value, which is readily known to those skilled in the art. References to “approximately” values ​​or parameters herein include (and are described) embodiments relating to the value or parameter itself.

[0071] For the purposes of this specification, “acceptor human framework” is a framework comprising the amino acid sequence of a light chain variable domain (VL) framework or a heavy chain variable domain (VH) framework derived from a human immunoglobulin framework or a human consensus framework, as defined below. An acceptor human framework “derived from” a human immunoglobulin framework or a human consensus framework may contain the same amino acid sequence or may contain amino acid sequence variations. In some embodiments, the number of amino acid variations is 10 or less, 9 or less, 8 or less, 7 or less, 6 or less, 5 or less, 4 or less, 3 or less, or 2 or less. In some embodiments, the VL acceptor human framework is sequence-identical with the VL human immunoglobulin framework sequence or the human consensus framework sequence.

[0072] "Affinity" refers to the sum of the non-covalent interactions between a single binding site of a molecule (e.g., an antibody) and its binding partner (e.g., an antigen). Unless otherwise indicated, as used herein, "binding affinity" refers to the intrinsic binding affinity that reflects the 1:1 interaction between the members of a binding pair (e.g., an antibody and an antigen). The affinity of molecule X for its partner Y is generally expressed by the dissociation constant (K). D ) can be expressed by. Affinity can be measured by common methods known in the art, including the methods described herein. Specific illustrative and exemplary embodiments for measuring binding affinity are described below.

[0073] "K D The term "measured by surface plasmon resonance assay" is used in the context of the claims, K D However, this means that the measurement is performed according to the method described in Example 1(A)(vii), which measures the kinetic parameters of the binding of the anti-tryptase antibody to the human tryptase beta-1 monomer, e.g., the His6-tagged tryptase monomer shown in Sequence ID No. 128, which does not spontaneously form a tryptase tetramer. The assay can be performed using a BIAcore® T200 or equivalent instrument. Briefly, a BIAcore® Series S CM5 sensor chip (or equivalent sensor chip) is immobilized with monoclonal mouse anti-human IgG(Fc) antibody, and the anti-tryptase antibody is sequentially captured on a flow cell. A series of 3-fold dilutions of human tryptase beta-1 monomer are injected at a flow rate of 30 μl / min. Each sample is analyzed by 3-minute association and 10-minute dissociation. The assay is performed at 25°C. After each injection, the chip is regenerated using 3M MgCl2. The binding response is corrected by subtracting the response unit (RU) from a flow cell that captures unrelated IgG of similar density. on and k off A 1:1 Languir model with simultaneous fitting is used for dynamic analysis.

[0074] Affinity-matured antibodies are those having one or more modifications in one or more HVR and / or framework regions, resulting in improved antibody affinity to the antigen compared to parent antibodies without those modifications. Preferred affinity-matured antibodies will have nanomolar or even picomolar affinity to the target antigen. Affinity-matured antibodies are produced by procedures known in the art. For example, Marks et al. Bio / Technology 10:779-783, 1992 describes affinity maturation by VH and VL domain shuffling. Random mutagenesis of HVR and / or framework residues is described by Barbas et al. Proc.Natl.Acad.Sci.USA 91:3809-3813,1994, Schier et al. Gene 169:147-155,1995, Yelton et al. J.Immunol.155:1994-2004,1995, Jackson et al. J.Immunol.154(7):3310-3319,1995, and Hawkins et al. J.Mol.Biol.226:889-896,1992.

[0075] The term "antibody" as used herein is used in its broadest sense and encompasses a variety of antibody structures, including, but not limited to, monoclonal antibodies, polyclonal antibodies, multispecific antibodies (e.g., bispecific antibodies), and antibody fragments, as long as they exhibit the desired antigen-binding activity.

[0076] As used herein, the term “tryptase” refers to any natural tryptase from any vertebrate source, including mammals such as primates (e.g., humans) and rodents (e.g., mice and rats), unless otherwise specified. Tryptases are also known in the art as mast cell tryptases, mast cell protease II, cutaneous tryptases, pulmonary tryptases, pituitary tryptases, mast cell neutral proteinases, and mast cell serine proteinase II. The term “tryptase” encompasses tryptase alpha (encoded in humans by TPSAB1), tryptase beta (encoded in humans by TPSAB1 and TPSB2; see below), tryptase delta (encoded in humans by TPSD1), tryptase gamma (encoded in humans by TPSG1), and tryptase epsilon (encoded in humans by PRSS22). Tryptase alpha, beta, and gamma proteins are soluble, while tryptase epsilon protein is membrane-bound. Tryptase beta and gamma are active serine proteases, but they have different specificities. Tryptase alpha and delta proteins are mostly inactive proteases because they have residues in important positions different from typical active serine proteases. An exemplary full-length protein sequence of tryptase alpha can be found under NCBI GenBank accession number ACZ98910.1 (SEQ ID NO: 118). An exemplary full-length protein sequence of tryptase gamma can be found under Uniprot accession number Q9NRR2 or GenBank accession numbers Q9NRR2.3, AAF03695.1, NP_036599.3, or AAF76457.1. Exemplary tryptase delta full-length protein sequences can be found under Uniprot accession number Q9BZJ3 or GenBank accession number NP_036349.1. Several tryptase genes are clustered on human chromosome 16p13.3. The term "full-length" encompasses untreated tryptase as well as any form of tryptase resulting from intracellular processing.Tryptase beta is the primary tryptase expressed in mast cells, while tryptase alpha is the primary tryptase expressed in basophils. Tryptase alpha and tryptase beta typically contain a leader sequence of approximately 30 amino acids and a catalytic sequence of approximately 245 amino acids (see, for example, Schwartz, Immunol. Allergy Clin. N.Am. 26:451-463, 2006).

[0077] As used herein, the term “tryptase beta” refers to any natural tryptase beta from any vertebrate source, including mammals such as primates (e.g., humans) and rodents (e.g., mice and rats), unless otherwise specified. Tryptase beta is a serine protease that is a major component of mast cell secretory granules. As used herein, the term encompasses tryptase beta 1 (encoded by the TPSAB1 gene, which also encodes tryptase alpha 1), tryptase beta 2 (encoded by the TPSB2 gene), and tryptase beta 3 (also encoded by the TPSB2 gene). An exemplary human tryptase beta 1 sequence is shown in Sequence ID No. 71 (see also GenBank accession number NP_003285.2). An exemplary human tryptase beta 2 sequence is shown in Sequence ID No. 72 (see also GenBank accession number AAD13876.1). An exemplary human tryptase beta 3 sequence is shown in Sequence ID No. 73 (see also GenBank accession number NP_077078.5). The term tryptase beta encompasses "full-length," untreated tryptase beta, as well as tryptase beta resulting from post-translational modifications, including proteolytic treatment. Full-length, protryptase beta is thought to be treated in two proteolytic steps. First, R -3Autocatalytic intermolecular cleavage occurs, particularly at an acidic pH and in the presence of polyanions (e.g., heparin or dextran sulfate). The remaining prodipeptide is then removed (likely by dipeptidyl petidase I). For full-length human tryptase beta-1, in the following SEQ ID NO: 71, the underlined amino acid residues correspond to the natural leader sequence, and the bold and gray-colored amino acid residues correspond to the prodomain, which is cleaved to form the mature protein (see, e.g., Sakai et al. J. Clin. Invest. 97:988-995, 1996). TIFF0007853934000001.tif35170

[0078] Mature, enzyme-active tryptase beta is typically homotetramer or heterotetramer, but active monomers have also been reported (see, e.g., Fukuoka et al. J.Immunol. 176:3165, 2006). The subunits of the tryptase beta tetramer are held together by hydrophobic and polar interactions between subunits and stabilized by polyanions (particularly heparin and dextran sulfate). The term tryptase can refer to either the tryptase tetramer or the tryptase monomer. Exemplary sequences of mature human tryptase beta 1, beta 2, and beta 3 are shown in SEQ ID NOs: 97, 116, and 117, respectively. Each subunit active site faces the central pore of the tetramer, which measures approximately 50 × 30 angstroms (see, e.g., Pereira et al. Nature 392:306-311, 1998). The size of the central pore typically restricts access to the active site by inhibitors. Exemplary substrates of tryptase beta include, but are not limited to, PAR2, C3, fibrinogen, fibronectin, and kininogen.

[0079] The terms “anti-tryptase antibody,” “tryptase-binding antibody,” and “tryptase-specific binding antibody” refer to an antibody that can bind to tryptase with sufficient affinity to be useful as a diagnostic and / or therapeutic agent when the antibody targets tryptase. In one embodiment, the degree of binding of the anti-tryptase antibody to unrelated non-tryptase proteins is less than about 10% of the binding of the antibody to tryptase, as measured, for example, by radioimmunoassay (RIA). In certain embodiments, the tryptase-binding antibody has a viscosity of ≤1 μM, ≤100 nM, ≤10 nM, ≤1 nM, ≤0.1 nM, ≤0.01 nM, or ≤0.001 nM (e.g., 10 -8 M or less, for example, 10 -8 M~10 -13 M, for example, 10 -9 M~10 -13 The dissociation constant (K) of M D ) has. In certain embodiments, the anti-tryptase antibody binds to an epitope of tryptase that is conserved among tryptases from different species.

[0080] An antibody that "binds to the same epitope as the reference antibody" refers to an antibody that, compared to the reference antibody, contacts overlapping sets of amino acid residues of the antigen or blocks the binding of the reference antibody to that antigen by 50% or more, 60% or more, 70% or more, 80% or more, or 90% or more in a competitive assay. In some embodiments, the set of amino acid residues contacted by the antibody may completely or partially overlap with the set of amino acid residues contacted by the reference antibody. In some embodiments, an antibody that binds to the same epitope as the reference antibody blocks the binding of the reference antibody to that antigen by 50% or more, 60% or more, 70% or more, 80% or more, or 90% or more in a competitive assay, and conversely, the reference antibody blocks the binding of the antibody to that antigen by 50% or more, 60% or more, 70% or more, 80% or more, or 90% or more in a competitive assay. Exemplary competitive assays are provided herein.

[0081] In the context of the claims, the term “determined by epitope binning assay” means that the antibody is determined to bind to the same epitope and / or compete for binding with a reference anti-tryptase antibody (e.g., hu31A.v11 or huE104.v2) using an OCTET® epitope binning assay, such as the one described in Section C of Example 3. Briefly, the human tryptase beta-1 monomer protein is biotinylated by reacting it with NHS-PEG4 biotin at a Lys residue. The biotinylated monomer is diluted to 5 μg / ml with a kinetic buffer (ForteBio, Inc.) and immobilized on a streptavidin sensor chip (ForteBio, Inc.). After the immobilization step, the human tryptase beta-1 immobilized sensor is saturated with a first antibody, diluted to 10-20 μg / ml, and then bound with a second antibody diluted to 2.5 μg / ml. The test temperature for such epitope binding assays is 30°C. A binding signal from the second antibody means that the two antibodies can simultaneously bind to the antigen at different, non-overlapping epitopes, but the binding signal does not mean that they share a common epitope. In some cases, a partial signal from the second antibody is observed (i.e., the signal is less than the signal observed when the first antibody is not added, but greater than the background), which means that the epitopes partially overlap.

[0082] An "antibody fragment" includes a portion of an intact antibody, preferably the antigen-binding region or variable region of an intact antibody. Examples of antibody fragments include Fab, Fab', F(ab')2, and Fv fragments; diabodies; linear antibodies (see U.S. Patent No. 5,641,870, Example 2, Zapata et al. Protein Eng. 8(10):1057-1062, 1995); single-chain antibody molecules; and multispecific antibodies formed from antibody fragments.

[0083] Papain digestion of the antibody produces two identical antigen-binding fragments called "Fab" fragments and one residual "Fc" fragment, denoted to reflect its ability to readily crystallize. The Fab fragments consist of the entire light chain along with the variable region domain (VH) of the heavy chain, as well as the first constant region domain (C) of one heavy chain. H 1) consists of two disulfide-bonded Fab fragments, which generally have divalent antigen-binding activity and can still crosslink to the antigen. The Fab' fragment contains one or more cysteine ​​derived from the hinge region of the antibody. H It differs from the Fab fragment in that it has several additional residues at the carboxyl terminus of one domain. Fab'-SH is the heretical notation for Fab' in which the cysteine ​​residue(s) of the constant domain have a free thiol group. The F(ab')2 antibody fragment was originally produced as a pair with the Fab' fragment, which has a hinged cysteine ​​in between. Other chemical conjugations of the antibody fragment are also known.

[0084] The term “Fc region” as used herein is used to define the C-terminal region of an immunoglobulin heavy chain that includes at least a portion of the constant region. This term includes both the native sequence Fc region and variant Fc regions. In one embodiment, the human IgG heavy chain Fc region extends from Cys226 or Pro230 to the carboxyl terminus of the heavy chain. However, the C-terminal lysine (Lys447) of the Fc region may or may not be present. Unless otherwise specified herein, the numbering of amino acid residues within the Fc region or constant region follows the EU numbering system, also known as the EU index, as described in Kabat et al., Sequences of Proteins of Immunological Interest, 5th Ed., Public Health Service, National Institutes of Health, Bethesda, MD, 1991.

[0085] "Fv" consists of a dimer of one heavy chain variable domain and one light chain variable domain, which are closely non-covalently bonded. The folding of these two domains creates six hypervariable loops (three from the H chain and three from the L chain) that provide amino acid residues for antigen binding and confer antigen-binding specificity to the antibody. However, even a single variable domain (or half of Fv containing only three antigen-specific H domains) has the ability to recognize and bind to an antigen, although its affinity may be lower than that of the entire binding site.

[0086] "Single-chain Fv," also abbreviated as "sFv" or "scFv," is an antibody fragment containing VH and VL antibody domains bound to a single polypeptide chain. Preferably, the sFv polypeptide further includes a polypeptide linker between the VH and VL domains, enabling the sFv to form a desired structure for antigen binding. For further information on sFv, see Pluckthun in The Pharmacology of Monoclonal Antibodies, vol. 113, Rosenburg and Moore eds., Springer-Verlag, New York, pp. 269-315, 1994.

[0087] The term "diabody" refers to a small antibody fragment prepared by constructing an sFv fragment (see previous paragraph) using a short linker (approximately 5-10 residues) between the VH and VL domains so that inter-chain V-domain pairing is achieved, rather than intra-chain, resulting in a bivalent fragment, i.e., a fragment having two antigen-binding sites. A bispecific diabody is a heterodimer of two "cross-resolved" sFv fragments in which the VH and VL domains of two antibodies reside on different polypeptide chains. Diabodies are fully described, for example, by EP404,097, WO93 / 11161, and Hollinger et al., Proc. Natl. Acad. Sci. USA 90:6444-6448, 1993.

[0088] A "binding domain" refers to a portion of a compound or molecule that specifically binds to a target epitope, antigen, ligand, or receptor. Binding domains include, but are not limited to, antibodies (e.g., monoclonal, polyclonal, recombinant, humanized, and chimeric antibodies), antibody fragments or portions thereof (e.g., Fab fragments, Fab'2, scFv antibodies, SMIP, domain antibodies, diabodies, minibodies, scFv-Fc, aphibodies, nanobodies, and the VH and / or VL domains of antibodies), receptors, ligands, aptamers, and other molecules having identified binding partners.

[0089] A “blocking” antibody or “antagonist” antibody is an antibody that inhibits or reduces the biological activity of the antigen to which it binds. Certain blocking antibodies or antagonist antibodies substantially or completely inhibit the biological activity of the antigen. In some embodiments, the activity may be tryptase enzyme activity, e.g., protease activity. In other examples, the activity may be tryptase-mediated stimulation of bronchial smooth muscle cell proliferation and / or collagenous system contraction. In other examples, the activity may be mast cell histamine release (e.g., IgE-induced histamine release and / or tryptase-induced histamine release). The antibodies of the present invention can inhibit the biological activity of tryptase by at least about 1%, about 5%, about 10%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, about 96%, about 97%, about 98%, about 99%, or about 100%.

[0090] The phrase "as determined by a human tryptase beta enzyme assay using synthetic peptide S-2288 as substrate" means, in the context of the claims, that the inhibitory activity is measured according to the assay described in Example 1(A)(viii)(a). Briefly, recombinant human tryptase beta-1 tetramer active enzyme is diluted to 0.75 nM in TNH buffer (200 mM Tris, 150 mM NaCl, 0.1 mg / mL heparin, 0.01% TRITON® X-100, pH 8.0) and combined 1:1 with antitryptase antibody (diluted in PBS) in a 384-well plate. The plate is incubated at ambient temperature for 1 hour with gentle agitation. Chromometric substrate S-2288 (Chromogenix, catalog number 82-0852-39), or an equivalent substrate, is diluted to 1200 μM in TNH buffer and added to the plate. The final well concentrations were 400 μM S-2288, 0.25 nM recombinant human tryptase beta-1 tetramer, 66 μg / mL heparin, and 0.10–222 nM anti-tryptase antibody. The plate was incubated at ambient temperature for 40 minutes with gentle agitation, and then A 405 The IC50 of anti-tryptase antibodies is determined from the 4-parameter fit of their respective curves.

[0091] The "class" of an antibody refers to the type of constant domain or constant region its heavy chain possesses. There are five main classes of antibodies: IgA, IgD, IgE, IgG, and IgM, some of which are further classified into subclasses (isotypes), such as IgG1, IgG2, IgG3, IgG4, IgA1, and IgA2. The heavy chain constant domains corresponding to different classes of immune globulins are called α, δ, ε, γ, and μ, respectively.

[0092] The "effector function" of an antibody refers to the biological activity attributed to the antibody's Fc region (either the Fc region of the natural sequence or the Fc region of an amino acid sequence variant), and is diverse depending on the antibody isotype. Examples of antibody effector functions include C1q binding and complement-dependent cytotoxicity, Fc receptor binding, antibody-dependent cell-mediated cytotoxicity (ADCC), phagocytosis, downregulation of cell surface receptors (e.g., B cell receptors), and B cell activation.

[0093] Antibody-dependent cell-mediated cytotoxicity, or ADCC, refers to a form of cytotoxicity in which secreted immunoglobulin (Ig) binds to Fc receptors (FcRs) present on certain cytotoxic cells (e.g., natural killer (NK) cells, neutrophils, and macrophages), enabling these cytotoxic effector cells to specifically bind to antigen-carrying target cells and subsequently kill them cytotoxicly. Antibodies "equip" cytotoxic cells and are absolutely necessary for such killing. Primary cells that mediate ADCC, such as NK cells, express only FcγRIII, while monocytes express FcγRI, FcγRII, and FcγRIII. FcR expression in hematopoietic cells is summarized in Table 3 on page 464 of Ravetch et al. Annu. Rev. Immunol. 9:457-492, 1991. To evaluate the ADCC activity of the target molecule, in vitro ADCC assays such as those described in U.S. Patent No. 5,500,362 or No. 5,821,337 may be performed. Useful effector cells for such assays include peripheral blood mononuclear cells (PBMCs) and natural killer (NK) cells. Alternatively, or in addition, the ADCC activity of the target molecule may be evaluated in vivo in animal models, such as those disclosed in Clynes et al. Proc. Natl. Acad. Sci. USA 95:652-656, 1998.

[0094] "Fc receptor" or "FcR" refers to a receptor that binds to the Fc region of an antibody. Preferred FcRs are human FcRs with natural sequences. Furthermore, preferred FcRs are those that bind to IgG antibodies (gamma receptors) and include the FcγRI, FcγRII, and FcγRIII subclass receptors, including allele variants and, alternatively, splicing forms of these receptors. FcγRII receptors include FcγRIIA ("activating receptor") and FcγRIIB ("inhibiting receptor"), which have similar amino acid sequences, primarily differing in their cytoplasmic domains. The activating receptor FcγRIIA contains an immunoreceptor tyrosine system activating motif (ITAM) in its cytoplasmic domain. The inhibiting receptor FcγRIIB contains an immunoreceptor tyrosine system inhibitory motif (ITIM) in its cytoplasmic domain (see overview M. in Daeron, Annu. Rev. Immunol. 15:203-234, 1997). FcRs are outlined, for example, in Ravetch et al. Annu. Rev. Immunol. 9:457-492, 1991, Capel et al. Immunomethods 4:25-34, 1994, and de Haas et al. J. Lab. Clin. Med. 126:330-41, 1995. Other FcRs, including those to be identified in the future, are encompassed by the term “FcR” as used herein. This term also includes neonatal receptor FcRn, which is involved in the transfer of maternal IgGs to the fetus (see, for example, Guyer et al. J. Immunol. 117:587, 1976, and Kim et al. J. Immunol. 24:249, 1994).

[0095] "Human effector cells" are leukocytes that express one or more FcRs and perform effector functions. Preferably, these cells express at least FcγRIII and perform ADCC effector functions. Examples of human leukocytes that mediate ADCC include peripheral blood mononuclear cells (PBMCs), natural killer (NK) cells, monocytes, cytotoxic T cells, and neutrophils, with PBMCs and NK cells being preferred. Effector cells can be isolated from natural sources, such as blood.

[0096] Complement-dependent cytotoxicity, or CDC, refers to the lysis of target cells in the presence of complement. Activation of the classical complement pathway is initiated by the binding of the first component of the complement system (C1q) to an antibody (of an appropriate subclass), to which its congener antigens bind. To assess complement activation, a CDC assay, such as that described in Gazzano-Santoro et al. J.Immunol. Methods 202:163, 1996, may be performed.

[0097] An "epitope" is the portion of an antigen to which an antibody selectively binds. For polypeptide antigens, a linear epitope is a peptide portion consisting of approximately 4 to 15 amino acid residues (e.g., 4, 5, 6, 7, 8, 9, 10, 11, or 12). A nonlinear conformational epitope may consist of residues of a close-proximity polypeptide sequence in the three-dimensional (3D) structure of the protein. In some embodiments, the epitope contains amino acids within 4 angstroms (Å) of any atom of the antibody. In some embodiments, the epitope contains amino acids of a tryptase protomer within 4 Å of any atom of the partner Fab. In certain embodiments, the epitope contains amino acids within 3.5 Å, 3 Å, 2.5 Å, or 2 Å of any atom of the antibody. The antibody amino acid residues that come into contact with the antigen (i.e., the paratope) can be determined, for example, by determining the crystal structure of the antibody conjugated with the antigen, or by performing hydrogen / deuterium exchange.

[0098] When used in the context of a claim, the phrase "the epitope is determined by an X-ray crystallographic model" means that an atom of a tryptase amino acid residue (e.g., a human tryptase beta-1 residue) is determined to be within 4 Å of any atom of an anti-tryptase antibody (e.g., any anti-tryptase antibody described herein, e.g., hu31A.v11 or huE104.v2) in an X-ray crystallographic model, such as the one described in Example 3. In some embodiments, the X-ray crystallographic model has a resolution of about 3.5 Å or less, about 3 Å or less, about 2.5 Å or less, about 2.15 Å or less, or about 2 Å or less.

[0099] The terms “full-length antibody,” “intact antibody,” and “whole antibody” are used herein synonymously to refer to antibodies that have a structure substantially similar to that of a natural antibody or that have a heavy chain containing an Fc region as defined herein.

[0100] A "human antibody" is one that has the amino acid sequence corresponding to an antibody produced by a human, and / or is produced using any of the techniques for creating human antibodies. This definition of a human antibody specifically excludes humanized antibodies that contain non-human antigen-binding residues.

[0101] The "Human Consensus Framework" is a framework representing the most commonly occurring amino acid residues in the selection of human immunoglobulin VL or VH framework sequences. Generally, the selection of human immunoglobulin VL or VH sequences derives from subgroups of variable domain sequences. Generally, these subgroups of sequences are those found in Kabat et al., Sequences of Proteins of Immunological Interest, Fifth Edition, NIH Publication 91-3242, Bethesda MD, vols. 1-3, 1991. In one embodiment, with respect to VL, the subgroup is subgroup Kappa III or Kappa IV, as found in Kabat et al. (see above). In one embodiment, with respect to VH, the subgroup is subgroup III in Kabat et al. (see above).

[0102] Humanized antibodies of non-human (e.g., rodent) antibodies are chimeric antibodies that contain minimal sequences derived from the non-human antibody. Mostly, a humanized antibody is a human immunoglobulin (recipient antibody) in which residues from the recipient's hypervariable region are replaced by residues from the hypervariable region of a non-human species (donor antibody), such as mouse, rat, rabbit, or non-human primate, possessing the desired antibody specificity, affinity, and capability. In some cases, framework region (FR) residues of the human immunoglobulin are replaced by corresponding non-human residues. Furthermore, humanized antibodies may contain residues not found in the recipient or donor antibody. These modifications are made to further enhance the antibody's capabilities. Generally, a humanized antibody will contain at least one, typically two, variable domains in which all or substantially all of the hypervariable loops correspond to the hypervariable loops of the non-human immunoglobulin, and all or substantially all of the FRs are FRs of the human immunoglobulin sequence. Humanized antibodies may optionally include the constant region (Fc) of immunoglobulins, typically at least a portion of the constant region of human immunoglobulins. For further details, see Jones et al. Nature 321:522-525, 1986, Riechmann et al. Nature 332:323-329, 1988, and Presta, Curr. Op. Struct. Biol. 2:593-596, 1992.

[0103] An "immune complex" is an antibody that complexes with one or more heterologous molecules, including but not limited to cytotoxic agents.

[0104] Where used to describe the various antibodies disclosed herein, the term “isolated” means an antibody that has been identified, separated, and / or recovered from the cells or cell cultures on which it expressed. Contaminations from its natural environment are typically materials that interfere with the diagnostic or therapeutic use of the polypeptide and may include enzymes, hormones, and other proteinaceous or non-proteinaceous solutes. In some embodiments, the antibody is purified to a purity of 95% or greater than 99%, as determined by methods such as electrophoresis (e.g., sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), isoelectric focusing (IEF), capillary electrophoresis) or chromatography (e.g., ion exchange or reverse-phase HPLC). For an overview of methods for determining antibody purity, see, for example, Flatman et al. J. Chromatogr. B 848:79-87, 2007. In preferred embodiments, the antibody is purified (1) using a spinning cup sequencer to a degree sufficient to obtain at least 15 residues of the N-terminal or internal amino acid sequence, or (2) using Coomassie blue or preferably silver staining, by SDS-PAGE under non-reducing or reducing conditions until homogeneous. Isolated antibodies include those of insights within recombinant cells, because at least one component of the polypeptide's natural environment is absent. However, the isolated polypeptide is usually prepared by at least one purification step.

[0105] As used herein, the term “monoclonal antibody” refers to an antibody obtained from a substantially homogeneous population of antibodies, i.e., the individual antibodies constituting that population are identical and / or bind to the same epitope on the antigen, except for variant antibodies that contain, for example, naturally occurring mutations or variant antibodies that may arise during the production of the monoclonal antibody preparation, the presence of such variants generally in small amounts. In contrast to polyclonal antibody preparations, which typically contain different antibodies directed to different determinants (epitopes), each monoclonal antibody in a monoclonal antibody preparation is directed to a single determinant on the antigen. Therefore, the modifier “monoclonal” indicates a characteristic of the antibody that it is obtained from a substantially homogeneous population of antibodies and should not be interpreted as requiring the production of the antibody by any particular method. For example, monoclonal antibodies used in accordance with the present invention may be produced by a variety of techniques, including but not limited to hybridoma methods, recombinant DNA methods, phage display methods, and methods utilizing transgenic animals containing all or part of the human immunoglobulin locus, and such methods and other exemplary methods for producing monoclonal antibodies are described herein. In certain embodiments, the term "monoclonal antibody" encompasses bispecific antibodies.

[0106] The term "bivalent antibody" refers to an antibody that has two binding sites for an antigen. Bivalent antibodies can be in either IgG format or F(ab')2 format, without limitation.

[0107] The term "multispecific antibody" is used most broadly to encompass antibodies that bind to two or more determinants or epitopes on one antigen, or to two or more determinants or epitopes on two or more antigens. Such multispecific antibodies include, but are not limited to, full-length antibodies, antibodies having two or more VL and VH domains, antibody fragments, e.g., Fab, Fv, dsFv, scFv, diabodies, bispecific diabodies, and triabodies, and covalently or non-covalently bound antibody fragments. "Polyepitope specificity" refers to the ability to specifically bind to two or more different epitopes on the same or different target(s). In certain embodiments, a multispecific antibody is a bispecific antibody. "Dual specificity" or "bispecificity" refers to the ability to specifically bind to two different epitopes on the same or different target(s). However, in contrast to bispecific antibodies, bispecific antibodies have two antigen-binding arms with identical amino acid sequences, and each Fab arm can recognize two antigens. Due to bispecificity, the antibody can interact with two different antigens as a single Fab or IgG molecule with high affinity. According to one embodiment, a multispecific antibody binds to each epitope with affinities of 5 μM to 0.001 pM, 3 μM to 0.001 pM, 1 μM to 0.001 pM, 0.5 μM to 0.001 pM, or 0.1 μM to 0.001 pM. "Single specificity" refers to the ability to bind to only one epitope.

[0108] A "naked antibody" refers to an antibody that is not complexed with a heterogeneous portion (e.g., a cytotoxic portion) or a radiolabeled molecule. Naked antibodies may be present in a pharmaceutical composition.

[0109] Regarding the binding of an antibody to a target molecule, the terms “binding,” “binding,” “specific binding,” “specifically binding,” or “specific to” a particular polypeptide or epitope on a particular polypeptide target mean a binding that is somewhat different from a nonspecific interaction. Specific binding can be measured, for example, by comparing the binding of a molecule to the binding of a control molecule. For example, specific binding can be determined by competition with a control molecule similar to the target, e.g., an excess of unlabeled targets. In this case, specific binding is indicated when the binding of a labeled target to a probe is competitively inhibited by an excess of unlabeled targets. The terms “specific binding,” “specifically binding to,” or “specifically binding to” an epitope on a particular polypeptide or epitope on a particular polypeptide target, as used herein, mean, for example, K to a target. D However, 10 -4 M or less, or 10 -5 M or less, or 10 -6 M or less, or 10 -7 M or less, or 10 -8 M or less, or 10 -9 M or less, or 10 -10 M or less, or 10 -11 M or less, or 10 -12 M or less, or 10 -4 M~10 -6 M, or 10 -6 M~10 -10 M or 10 -7 M~10 -9 This can be exhibited by molecules in the M range. As will be understood by those skilled in the art, affinity and K D This is inversely proportional. A high affinity for an antigen corresponds to a low K D It is measured by a value. In one embodiment, the term “specific binding” refers to a binding in which a molecule binds to a specific polypeptide or an epitope on a specific polypeptide without substantially binding to any other polypeptide or polypeptide epitope.

[0110] A "paratope" refers to a portion of an antibody that binds to an epitope of an antigen. Typically, a paratope is a region of approximately 15-22 amino acid residues in the Fv region of an antibody, and the V region of an antibody. H and V L It may contain amino acids from the chain.

[0111] The term "variable" refers to the fact that certain segments of a variable domain differ extensively in sequence across antibodies. The variable, or "V," domain mediates antigen binding and defines the specificity of a particular antibody to that particular antigen. However, variability is not evenly distributed across the entire 110-amino acid-length variable domain. Instead, the V region consists of a relatively invariant range called a framework region (FR) of 15-30 amino acids, separated by shorter, highly variable regions called "hypervariable regions," each 9-12 amino acids long. When used herein, the term "hypervariable region" or "HVR" refers to the amino acid residues of an antibody involved in antigen binding. The hypervariable region generally includes amino acid residues from approximately 24-34 (L1), 50-56 (L2), and 89-97 (L3) in VL, and approximately 26-35 (H1), 49-65 (H2), and 95-102 (H3) in VH (in one embodiment, H1 is approximately 31-35), Kabat et al (see above) and / or the "hypervariable loop" (e.g., amino acid residues from 26-32 (L1), 50-52 (L2), and 91-96 (L3) in VL, and 26-32 (H1), 53-55 (H2), and 96-101 (H3) in VH; Chothia et al. al.J.Mol.Biol.196:901-917,1987. The variable domains of the natural heavy and light chains each contain four FRs that primarily adopt a beta-sheet configuration, connected by three hypervariable regions that form loops connecting beta-sheet structures and, in some cases, forming part of them. The hypervariable region in each chain is held in close proximity to the hypervariable region of the other chain by the FRs, contributing to the formation of the antibody's antigen-binding site (see Kabat et al. (see above)). Therefore, the HVR and FR sequences generally appear in the following sequence in VH (or VL): FR1-H1(L1)-FR2-H2(L2)-FR3-H3(L3)-FR4. The constant domain does not directly participate in the binding of the antibody to the antigen, but exhibits various effector functions, such as the antibody's involvement in antibody-dependent cytotoxicity (ADCC).

[0112] The terms “Kabat-like variable domain residue numbering” or “Kabat-like amino acid position numbering,” and their variations, refer to the numbering system used in Kabat et al. (see above) for the heavy-chain or light-chain variable domains of antibody edits. Using this numbering system, the actual linear amino acid sequence may contain fewer or more amino acids corresponding to the shortening or insertion of FR or HVR in the variable domain. For example, a heavy-chain variable domain may contain a single amino acid insertion fragment after H2 residue 52 (residue 52a by Kabat) and a residue inserted after heavy-chain FR residue 82 (e.g., residues 82a, 82b, and 82c by Kabat). The Kabat numbering of residues can be determined for a given antibody by alignment of the antibody sequence in homology regions with a “standard” Kabat-numbered sequence.

[0113] The Kabat numbering system is generally used to refer to residues within the variable domain (approximately residues 1-107 of the light chain and residues 1-113 of the heavy chain) (e.g., Kabat et al. (see above)). The "EU numbering system" or "EU index" is generally used to refer to residues within the constant region of the immunoglobulin heavy chain (e.g., the EU index reported by Kabat et al. (see above)). "EU index as found in Kabat" refers to the residue numbering of human IgG1 EU antibodies. Unless otherwise indicated herein, references to residue numbers within the variable domain of an antibody mean residue numbering according to the Kabat numbering system. Unless otherwise indicated herein, references to residue numbers within the constant domain of an antibody mean residue numbering according to the EU numbering system (e.g., see U.S. Provisional Patent Application No. 60 / 640,323, drawings relating to EU numbering).

[0114] "Disorder" or "disease" is any condition that would benefit from treatment with an antibody (e.g., any tryptase antibody described herein). This includes chronic and acute disorders or diseases, including pathological conditions that make a mammal susceptible to the disorder in question. In some embodiments, the disorder is a lung disorder, autoimmune disorder, inflammatory disorder, fibrous disorder, granulocytic (neutrophilic or eosinophilic) disorder, monocytic disorder, lymphocytic disorder, or a disorder related to an increased number or distribution of normal or abnormal tissue resident cells (such as mast cells, macrophages, or lymphocytes) or stromal cells (such as fibroblasts, myofibroblasts, smooth muscle cells, epithelium, or endothelium). In some embodiments, the disorder is a lung disorder. In some examples, the disorder may be a tryptase-associated disorder or tryptase-mediated disorder.

[0115] As used herein, the terms “tryptase-related disorder” and “tryptase-mediated disorder” refer to any disorder or condition mediated by or associated with tryptase. In some embodiments, tryptase-related disorder is associated with excessive tryptase levels or activity that may result in atypical symptoms due to local and / or systemic tryptase levels or activity in the body.

[0116] In some embodiments, the lung disorder is asthma. In some embodiments, the asthma is persistent chronic severe asthma with acute events of worsening symptoms (exacerbations or sudden onsets) that may be fatal. In some embodiments, the asthma is atopic (also known as allergic) asthma, non-allergic asthma (e.g., often triggered by respiratory viruses (e.g., influenza, parainfluenza, rhinovirus, human metapneumovirus, and respiratory rash virus), or infections caused by inhaled irritants (air pollutants, fumes, diesel particles, volatile chemicals and gases indoors or outdoors, or even cold, dry air)).

[0117] In some embodiments, asthma is asthma resulting from intermittent or exercise-induced, or acute or chronic direct or indirect exposure to “smoke” (typically cigarettes, cigars, or pipes), inhalation, or “vapor inhalation” (tobacco, cannabis, or other such substances), or asthma induced by recent ingestion of aspirin or related NSAIDs. In some embodiments, asthma is mild or corticosteroid-unsensitized asthma, newly diagnosed and untreated asthma, or asthma that previously did not require chronic use of topical or systemic inhaled steroids to control symptoms (cough, wheezing, shortness of breath / dyspnea, or chest pain). In some embodiments, asthma is chronic, corticosteroid-resistant, corticosteroid-refractory, or asthma that is not controlled by corticosteroids or other chronic asthma controllers.

[0118] In some embodiments, asthma is moderate to severe asthma. In certain embodiments, asthma is Th2 hyperasthma. In some embodiments, asthma is severe asthma. In some embodiments, asthma is atopic asthma, allergic asthma, non-allergic asthma (e.g., caused by infection and / or respiratory rash virus (RSV)), exercise-induced asthma, aspirin-sensitive / exacerbated asthma, mild asthma, moderate to severe asthma, corticosteroid-unsensitized asthma, chronic asthma, corticosteroid-resistant asthma, corticosteroid-refractory asthma, newly diagnosed untreated asthma, smoking-induced asthma, and asthma that is not controlled by corticosteroids. In some embodiments, asthma is type 2 T helper lymphocyte (Th2) or type 2 (Th2) hyperT helper lymphocyte, or type 2 (T2) driven asthma. In some embodiments, asthma is eosinophilic asthma. In some embodiments, the asthma is allergic asthma. In some embodiments, the individual is determined to be eosinophilic inflammation-positive (EIP). See WO2015 / 061441. In some embodiments, the asthma is hyperperiostin asthma (e.g., having a periostin level of at least about 20 ng / mL, 25 ng / mL, or 50 ng / mL serum). In some embodiments, the asthma is hypereosinophilic asthma (e.g., at least about 150, 200, 250, 300, 350, or 400 eosinophil counts / ml blood). In certain embodiments, the asthma is hypothoracic or non-Th2 driven asthma. In some embodiments, the individual is determined to be eosinophilic inflammation-negative (EIN). See WO2015 / 061441. In some embodiments, the asthma is hypothoracic (e.g., having a periostin level of less than about 20 ng / mL serum). In some embodiments, the asthma is hypoeosinophilic asthma (e.g., less than approximately 150 eosinophils / μl blood or less than approximately 100 eosinophils / μl blood).

[0119] As used herein, the term “Th2 hyperasthma” refers to asthma exhibiting high levels of one or more Th2 cell-associated cytokines, such as IL13, IL4, IL9, and IL5, or exhibiting Th2 cytokine-associated inflammation. In certain embodiments, the term Th2 hyperasthma may be used interchangeably with eosinophilic hyperasthma. In certain embodiments, Th2 hyperasthma is Th2-driven asthma. In some embodiments, the asthma patient is determined to be eosinophilic inflammation-positive (EIP). See, for example, International Patent Application Publication WO2015 / 061441, which is incorporated herein in its entirety by reference. In certain embodiments, the individual is determined to have an elevated level of at least one of the eosinophilic characteristic genes compared to a control or reference level. See WO2015 / 061441. In certain embodiments, Th2 hyperasthma is periostin hyperasthma. In some embodiments, the individual has high serum periostin levels. In certain embodiments, the individual is 18 years of age or older. In certain embodiments, an individual is determined to have an elevated level of serum periostin compared to a control or reference level. In certain embodiments, the control or reference level is an intermediate level of periostin in the population. In certain embodiments, an individual is determined to have serum periostin of 20 ng / ml or higher. In certain embodiments, an individual is determined to have serum periostin of 25 ng / ml or higher. In certain embodiments, an individual is determined to have serum periostin of 50 ng / ml or higher. In certain embodiments, the control or reference level of serum periostin is 20 ng / ml, 25 ng / ml, or 50 ng / ml. In certain embodiments, asthma is defined as eosinophilic asthma. In certain embodiments, an individual is determined to have an elevated eosinophil count compared to a control or reference level. In certain embodiments, the control or reference level is an intermediate level in the population. In certain embodiments, an individual is determined to have an eosinophil count of 150 or higher / μl blood. In a particular embodiment, an individual is determined to have a blood eosinophil count of 200 or more per μl.In certain embodiments, the individual is determined to have an eosinophil count of 250 or more per μl of blood. In certain embodiments, the individual is determined to have an eosinophil count of 300 or more per μl of blood. In certain embodiments, the individual is determined to have an eosinophil count of 350 or more per μl of blood. In certain embodiments, the individual is determined to have an eosinophil count of 400 or more per μl of blood. In certain embodiments, the individual is determined to have an eosinophil count of 450 or more per μl of blood. In certain embodiments, the individual is determined to have an eosinophil count of 500 or more per μl of blood. In certain preferred embodiments, the individual is determined to have an eosinophil count of 300 or more per μl of blood. In certain embodiments, eosinophils are peripheral blood eosinophils. In certain embodiments, eosinophils are sputum eosinophils. In certain embodiments, the individual exhibits elevated levels of FeNO (exhaled nitric oxide) and / or elevated levels of IgE. For example, in some cases, individuals exhibit FeNO levels greater than approximately 250 parts per billion (ppb), greater than approximately 275 ppb, greater than approximately 300 ppb, greater than approximately 325 ppb, greater than approximately 325 ppb, or greater than approximately 350 ppb. In some cases, individuals have IgE levels greater than 50 IU / ml.

[0120] As used herein, the terms “Th2 hypoasthma” or “non-Th2 hyperasthma” refer to asthma exhibiting low levels of one or more Th2 cell-associated cytokines, such as IL13, IL4, IL9, and IL5, or exhibiting non-Th2 cytokine-associated inflammation. In certain embodiments, the term Th2 hypoasthma may be used interchangeably with eosinophil hypoasthma. In some embodiments, the asthma patient is determined to be eosinophilic inflammation-negative (EIN). See, for example, WO2015 / 061441. In certain embodiments, Th2 hypoasthma is Th17-driven asthma. In certain embodiments, Th2 hypoasthma is periostin hypoasthma. In certain embodiments, the individual is 18 years of age or older. In certain embodiments, the individual is determined to have reduced levels of serum periostin compared to a control or reference level. In certain embodiments, the control or reference level is an intermediate level of periostin in the population. In certain embodiments, the individual is determined to have serum periostin less than 20 ng / ml. In certain embodiments, asthma is defined as hypoeosinophilic asthma. In certain embodiments, an individual is determined to have a reduced eosinophil count compared to a control or reference level. In certain embodiments, the control or reference level is an intermediate level for the population. In certain embodiments, an individual is determined to have an eosinophil count of less than 150 / μl blood. In certain embodiments, an individual is determined to have an eosinophil count of less than 100 / μl blood. In certain preferred embodiments, an individual is determined to have an eosinophil count of less than 300 / μl blood.

[0121] In some embodiments, autoimmune disorders, inflammatory disorders, fibrotic disorders, granulocytic (neutrophilic or eosinophilic) disorders, monocytic disorders, or lymphocytic disorders include esophagitis (e.g., eosinophilic esophagitis), allergic rhinitis, non-allergic rhinitis, rhinosinusitis with polyps, nasal polyps, bronchitis, chronic pneumonia, allergic bronchopulmonary aspergillosis, airway inflammation, allergic rhinitis, bronchiectasis, and / or chronic bronchitis.

[0122] In some embodiments, autoimmune deficiency, inflammatory disorders, fibrous disorders, granulocytic (neutrophilic or eosinophilic) disorders, monocytic disorders, or lymphocytic disorders are arthritis. In some embodiments, arthritis is rheumatoid arthritis. In some embodiments, arthritis is osteoarthritis, rheumatoid arthritis, juvenile arthritis, early rheumatoid arthritis, polyarticular rheumatoid arthritis, systemic rheumatoid arthritis, enteric arthritis, reactive arthritis, psoriatic arthritis, and / or arthritis as a result of injury.

[0123] In some embodiments, autoimmune deficiency, inflammatory disorders, fibrous disorders, granulocytic (neutrophilic or eosinophilic) disorders, monocytic disorders, or lymphocytic disorders are gastrointestinal conditions. In some embodiments, gastrointestinal conditions include IBD (inflammatory bowel disease), ulcerative colitis (UC), Crohn's disease (CD), colitis (e.g., colitis caused by external stimuli (e.g., caused by or associated with treatment regimens, e.g., chemotherapy, radiotherapy, etc.)), infectious colitis, ischemic colitis, collagenous or lymphocytic colitis, necrotizing enterocolitis, colitis in conditions such as chronic granulomatous disease or celiac disease, food allergies, gastritis, gastroenteritis, infectious gastritis or enteritis (e.g., chronic active gastritis due to Helicobacter pylori infection), esophagitis, and other forms of gastrointestinal inflammation caused by infectious agents, or ulcerative colitis.

[0124] In some embodiments, autoimmune deficiency, inflammatory disorders, fibrotic disorders, granulocytic (neutrophilic or eosinophilic) disorders, monocytic disorders, or lymphocytic disorders constitute a gastroenteritis state. In some embodiments, the gastroenteritis state is IBD (inflammatory bowel disease). In some embodiments, the inflammatory bowel disease is ulcerative colitis (UC) or Crohn's disease (CD). In some embodiments, the gastrointestinal inflammatory state is colitis (e.g., colitis caused by external stimuli (e.g., caused by or associated with treatment regimens, e.g., chemotherapy, radiotherapy, etc.)), infectious colitis, ischemic colitis, collagenous or lymphocytic colitis, necrotizing enterocolitis, colitis in conditions such as chronic granulomatous disease or celiac disease, food allergies, gastritis, gastroenteritis, infectious gastritis or enteritis (e.g., chronic active gastritis due to Helicobacter pylori infection), and other forms of gastrointestinal inflammation caused by an infectious agent, or ulcerative colitis. In some embodiments, the gastrointestinal inflammatory state is ulcerative colitis (UC) or Crohn's disease (CD). In some embodiments, the gastrointestinal inflammatory state is ulcerative colitis (UC). In some embodiments, ulcerative colitis is mild to moderate distal colitis. In some embodiments, ulcerative colitis is mild to moderate widespread colitis. In some embodiments, ulcerative colitis is severe colitis. In some embodiments, the gastrointestinal inflammatory state is Crohn's disease (CD). In some embodiments, Crohn's disease is in the acute phase. In some embodiments, Crohn's disease is in the induced clinical remission phase. In some embodiments, Crohn's disease is in the sustained response / remission phase. In some embodiments, Crohn's disease is a mild to moderate disease. In some embodiments, Crohn's disease is a moderate to severe disease. In some embodiments, Crohn's disease is a severe / fulminant disease. In some embodiments, Crohn's disease is an ileal, ileocolonic, or colonic disease.

[0125] In some embodiments, autoimmune deficiency, inflammatory disorders, fibrous disorders, granulocytic (neutrophilic or eosinophilic) disorders, monocytic disorders, or lymphocytic disorders, or disorders associated with an increased number or distribution of normal or abnormal tissue resident cells (such as mast cells, macrophages, or lymphocytes) or stromal cells (fibroblasts, myofibroblasts, smooth muscle cells, epithelium, or endothelium) are one or more organ-specific symptoms of lupus or systemic lupus erythematosus (SLE), or lupus (e.g., lupus nephritis (LN) affecting the kidneys, or extrarenal lupus (ERL) affecting blood and / or lymphatic organs (lymph nodes, spleen, thymus, and associated lymphatic vessels), and / or joints and / or other organs, but not necessarily the kidneys).

[0126] In some embodiments, autoimmune disorders, inflammatory disorders, or fibrotic disorders include idiopathic (of unknown etiology) conditions such as sepsis and / or trauma, HIV infection, or ANCA-associated vasculitis (AAV), granulomatosis with polyangiitis (previously known as Wegener's granulomatosis), Behçet's disease, cardiovascular disease, eosinophilic bronchitis, Reiter's syndrome, SEA syndrome (seronegative, enthesopathy, arthropathy syndrome), and ankylosis. Spondylitis, dermatomyositis, scleroderma, systemic sclerosis (also known as systemic sclerosis), vasculitis (e.g., temporal arteritis, cranial arteritis, or giant cell arteritis (GCA), also known as Horton's disease), myositis, polymyositis, dermatomyositis, arteritis, polymyalgia rheumatica, sarcoidosis, primary biliary cirrhosis, sclerosing cholangitis, Sjögren's syndrome, psoriasis, psoriasis vulgaris, guttate psoriasis, reverse psoriasis, pustular psoriasis, erythrodermic psoriasis Dermatitis, atopic dermatitis, pemphigus, e.g., pemphigus vulgaris, atherosclerosis, lupus, Still's disease, myasthenia gravis, celiac disease, relapsing-remitting (RRMS), primary progressive (PPMS), or secondary progressive (SPMS) subtypes of multiple sclerosis (MS), Guillain-Barré disease, type 1 diabetes mellitus (T1DM), insulin-dependent diabetes mellitus (IDDM), or juvenile-onset diabetes mellitus, thyroiditis It is associated with adenitis (e.g., Graves' disease), celiac disease, Churg-Strauss syndrome, myalgia syndrome, eosinophilic syndrome, edema reactions including eosinophilic angioedema, helminthic infections, onchocerciasis, eosinophilic esophagitis, eosinophilic enteritis, eosinophilic colitis, obstructive sleep apnea, endocardial fibrosis, Addison's disease, Raynaud's disease or phenomenon, autoimmune hepatitis, graft-versus-host disease (GVHD), or organ transplant rejection.

[0127] In some embodiments, the disorder is an inflammatory disorder of the skin. In some embodiments, the disorder is atopic dermatitis or onchocerciasis. In some embodiments, the disorder is chronic idiopathic urticaria (CIU or CSU).

[0128] In some embodiments, autoimmune deficiency, inflammatory disorders, fibrotic disorders, neutrophil disorders, or eosinophilic disorders are fibrotic disorders. In some embodiments, fibrotic disorders include pulmonary fibrosis, hepatic fibrosis (e.g., fibrosis associated with cirrhosis (e.g., alcohol-induced cirrhosis, viral-induced cirrhosis, post-hepatitis C cirrhosis, and primary biliary cirrhosis), schistosomiasis, cholangitis (e.g., sclerosing cholangitis), and autoimmune-induced hepatitis), renal fibrosis (e.g., tubulointerstitial fibrosis, scleroderma, diabetic nephritis, and glomerulonephritis), cutaneous fibrosis (e.g., scleroderma, hypertrophic and keloid scarring, renal fibrotic skin). This includes fibrosis (and burns), myelofibrosis, neurofibromatosis, fibroma, intestinal fibrosis, and fibrous adhesions resulting from surgical procedures), cardiac fibrosis (e.g., fibrosis associated with myocardial infarction), vascular fibrosis (e.g., fibrosis associated with post-angioplasty arterial restenosis and atherosclerosis), ocular fibrosis (e.g., fibrosis associated with proliferative vitreoretinopathy after cataract surgery, and posterior orbital fibrosis), and myelofibrosis (e.g., idiopathic myelofibrosis and drug-induced myelofibrosis). Fibrosis can be organ-specific or systemic (e.g., systemic sclerosis, and fibrosis associated with GVHD). In some embodiments, the fibrotic disorder is pulmonary fibrosis. In some embodiments, pulmonary fibrosis is fibrous interstitial pneumonia. In some embodiments, pulmonary fibrosis is idiopathic pulmonary fibrosis (IPF), also known as idiopathic fibrotic alveolitis. In some embodiments, IPF is sex, age, and physiological function (GAP) stage I. In some embodiments, IPF is GAP stage II. In some embodiments, IPF is GAP stage III. In some embodiments, pulmonary fibrosis is sporadic IPF. In some embodiments, pulmonary fibrosis is familial pulmonary fibrosis. In some embodiments, pulmonary fibrosis is combined pulmonary fibrosis and emphysema.In some embodiments, pulmonary fibrosis is associated with one or more of the following: typical interstitial pneumonia, idiopathic interstitial pneumonia, desquamative interstitial pneumonia, bronchiolitis interstitial lung disease, acute interstitial pneumonia, nonspecific interstitial pneumonia, sarcoidosis, idiopathic organizing pneumonia, eosinophilic pneumonia, infection, occupational or environmental exposure to substances, cigarette smoking, drug or radiation-induced interstitial lung disease, rheumatic disease-associated interstitial lung disease, lymphoid interstitial pneumonia, upper lobe-predominant pulmonary fibrosis, pulmonary Langerhans cell histiocytosis, systemic scleroderma interstitial lung disease, Hermanskie-Puddlak syndrome, and diseases due to telomere shortening.

[0129] In some embodiments, lung damage, autoimmune deficiency, inflammatory disorders, fibrotic disorders, neutrophil disorders, and eosinophilic disorders constitute chronic obstructive pulmonary disease (COPD). In some embodiments, COPD is the international guideline for chronic obstructive pulmonary disease (GOLD) category A. In some embodiments, COPD is GOLD category B. In some embodiments, COPD is GOLD category C. In some embodiments, COPD is GOLD category D. In some embodiments, COPD is chronic bronchitis. In some embodiments, COPD is emphysema. In some embodiments, emphysema is proximal acinar, panlobular, or distal acinar emphysema. In some embodiments, emphysema is tobacco-induced emphysema. In some embodiments, COPD is associated with exposure to particulate matter, chemical fumes, and / or air pollution. In some embodiments, COPD is associated with impaired lung development. In some embodiments, COPD is chronic obstructive asthma. In some embodiments, COPD is associated with alpha-1 antitrypsin deficiency. In some embodiments, COPD is associated with the fission of serine protease inhibitor clade E, member 2 (SERPINE2). In some embodiments, COPD is COPD with persistent systemic inflammation. In some embodiments, COPD is associated with eosinophilic or type 2 T helper (T) H2) The patient has high COPD. In some embodiments, COPD is COPD with persistent bacterial colonization. In some embodiments, COPD is COPD with frequent exacerbations. In some embodiments, autoimmune deficiency, inflammatory disorder, fibrous disorder, neutrophil disorder, or eosinophilic disorder constitutes asthma-COPD overlap syndrome (ACOS). In some embodiments, ACOS is eosinophilic, neutrophilic, mixed pattern, or non-inflammatory (nongranulocytic) ACOS. In some embodiments, autoimmune deficiency, inflammatory disorder, fibrous disorder, neutrophil disorder, or eosinophilic disorder constitutes COPD-obstructive sleep apnea (OSA) overlap syndrome.

[0130] The above list is not exhaustive, and those skilled in the art will understand that diseases or disorders can fall into various categories.

[0131] The term “packaging insert” is used to refer to the instructions typically included in the commercial packaging of a therapeutic product, which contain information about the indications, use, dosage, administration, combination therapies, contraindications, and / or warnings regarding the use of such therapeutic product.

[0132] The terms “pharmaceutical preparation” and “pharmaceutical composition” are used interchangeably herein and refer to preparations in which the biological activity of the active ingredients contained herein is effective and which do not contain additional ingredients that are toxic to an extent unacceptable to the subject to which the preparation is administered. Such preparations are sterile. In preferred embodiments, the pharmaceutical composition or pharmaceutical preparation is administered to a human subject.

[0133] A "sterilized" pharmaceutical preparation is either sterile, free from all viable microorganisms and their spores, or essentially free from them.

[0134] A “stable” pharmaceutical formulation is one in which the internal proteins (e.g., antibodies such as anti-tryptase antibodies) inherently retain their physical and / or chemical stability and / or biological activity during storage. Preferably, the formulation inherently retains its physical and chemical stability, as well as its biological activity, during storage. The storage period is generally selected based on the intended shelf life of the formulation. Various analytical techniques for measuring protein stability are available in the art and are outlined, for example, in Peptide and Protein Drug Delivery, 247-301, Vincent Lee Ed., Marcel Dekker, Inc., New York, NY, Pubs. (1991) and Jones, A. Adv. Drug Delivery Rev. 10:29-90 (1993). Stability can be measured over a selected period of time at a selected exposure level and / or temperature. Stability can be evaluated qualitatively and / or quantitatively by a variety of different methods, including: assessment of aggregate formation (e.g., by measuring turbidity using size exclusion chromatography and / or visual inspection); assessment of ROS formation (e.g., by using a photostress assay or AAPH stress assay); oxidation of specific amino acid residues of the protein (e.g., Trp and / or Met residues of monoclonal antibodies); assessment of charge heterogeneity using cation exchange chromatography, image capillary isoelectric focusing (icIEF), or capillary zone electrophoresis; amino-terminus or carboxy-terminus sequence analysis; mass spectrometry; SDS-PAGE analysis for comparing reduced intact antibodies; peptide mapping (e.g., trypsin or LYS-C) analysis; and evaluation of the biological activity or target binding function of the protein (e.g., antigen binding function of the antibody). Instability may be accompanied by one or more of the following: aggregation, deamide (e.g., Asn deamide), oxidation (e.g., Met oxidation and / or Trp oxidation), isomerization (e.g., Asp isomerization), clipping / hydrolysis / fragmentation (e.g., hinge region fragmentation), succinimide formation, unpaired cysteine(s), N-terminal elongation, C-terminal processing, glycosylation differences, etc.

[0135] An antibody (e.g., an anti-tryptase antibody) "maintains its physical stability" in a pharmaceutical formulation if it shows little or no signs of aggregation, precipitation, fragmentation, and / or denaturation when measured by visual inspection of color and / or clarity, or by UV light scattering or size exclusion chromatography.

[0136] An antibody (e.g., an anti-tryptase antibody) "retains its chemical stability" in a pharmaceutical formulation if its chemical stability at a given time point is such that the antibody is still considered to retain its biological activity as defined below. Chemical stability can be assessed by detecting and quantifying chemically modified forms of the antibody. Chemical modifications may include, for example, protein oxidation, which can be assessed using trypsin peptide mapping, reversed-phase high-performance liquid chromatography (HPLC), and liquid chromatography-mass spectrometry (LC / MS). Other types of chemical modifications include, for example, charge modification of the antibody, which can be assessed by ion-exchange chromatography or icIEF.

[0137] An antibody (e.g., an anti-tryptase antibody) "retains its biological activity" in a pharmaceutical formulation if its biological activity at a given time point is within approximately 20% (e.g., within approximately 10%) of the biological activity exhibited at the time the pharmaceutical formulation is prepared, as determined, for example, by an antigen-binding assay or in vitro inhibition assay of a monoclonal antibody (e.g., an anti-tryptase monoclonal antibody) (within the error of the assay). In some embodiments, the biological activity of the antibody at a given time point is approximately 25%, approximately 30%, approximately 35%, approximately 40%, approximately 45%, or approximately 50% of the biological activity exhibited at the time the pharmaceutical formulation is prepared.

[0138] As used herein, “biological activity” of an antibody (e.g., an anti-tryptase antibody) refers to the antibody’s ability to bind to a target, for example, the ability of a monoclonal antibody to bind to an antigen. This may further include a biological response that can be measured in vitro or in vivo. Such activity may be antagonist activity or agonist activity.

[0139] "Oxidation-sensitive" proteins (e.g., antibodies such as anti-tryptase antibodies) are proteins containing one or more residues that have been found to be easily oxidized, including but not limited to methionine (Met), cysteine ​​(Cys), histidine (His), tryptophan (Trp), and tyrosine (Tyr). For example, the tryptophan amino acid in the Fab portion of a monoclonal antibody or the methionine amino acid in the Fc portion of a monoclonal antibody may be oxidation-sensitive.

[0140] The term "oxidation percentage" refers to the percentage of an antibody in a formulation (e.g., a pharmaceutical composition) that is oxidized by a specific amino acid residue, such as a Trp residue (e.g., Trp100 in HVR-H3 of hu31A.v11) or a Met residue. The oxidation percentage can be determined, for example, by mass spectrometry (MS) of one or more trypsin peptides that contain one or more specific oxidized prawn amino acid residues. In a particular embodiment, the oxidation percentage of Trp100 in HVR-H3 of hu31A.v11 is determined by the mass of oxidized trypsin peptides, where Trp100 is present in excess of the total (oxidized and unoxidized) trypsin peptide mass when determined by MS analysis. The oxidation percentage can be determined, for example, within 9 months, 12 months, 18 months, or 2 years from the initial production of the antibody or its pharmaceutical composition.

[0141] As used herein, the term “determined according to the AAPH stress test” means that the percentage of oxidation at a specific amino acid residue (e.g., Trp100 in HVR-H3 of hu31A.v11) is determined by mass spectrometry of the trypsin peptide after compounding the antibody at 150 mg / ml in 5 mM AAPH for 25 hours at 40°C, as described in Example 5, for example. The stress antibody was digested with trypsin, and the digested peptide was subjected to ultra-high-performance liquid chromatography-high-resolution mass spectrometry (UHPLC-HRMS) to determine the percentage of oxidation.

[0142] As used herein, “buffer” refers to a buffer solution that resists changes in pH due to the action of its acid-base complex components. The buffers of this invention preferably have a pH in the range of about 4.5 to about 8.0 (e.g., about 4.5, about 5, about 5.5, about 6, about 6.5, about 7, about 7.5, or about 8), for example, about pH 5.5. For example, histidine acetate is an example of a buffer that controls pH within this range. Another preferred buffer is arginine succinate and / or histidine succinate.

[0143] A “preservative” is a compound that may be optionally included in a formulation to, for example, essentially reduce internal bacterial activity and thus facilitate the production of a multipurpose formulation. Examples of possible preservatives include, for example, octadecyldimethylbenzylammonium chloride, hexamethonium chloride, benzalkonium chloride (a mixture of alkylbenzyldimethylammonium chlorides in which the alkyl group is a long-chain compound), and benzethonium chloride. Other types of preservatives include aromatic alcohols, such as phenol, butyl, and benzyl alcohol; alkylparabens, such as methyl or propylparaben; catechol, resorcinol, cyclohexanol, 3-pentanol, and m-cresol. In one embodiment, the preservative as used herein is benzyl alcohol.

[0144] As used herein, “surfactant” means a surfactant, preferably a nonionic surfactant. Examples of surfactants as used herein include polysorbate (e.g., polysorbate 20 and polysorbate 80), poloxamer (e.g., poloxamer 188), Triton (registered trademark), sodium dodecyl sulfate (SDS), sodium lauryl sulfate, sodium octyl glucoside, lauryl sulfobetaine, myristyl sulfobetaine, linoleyl sulfobetaine, or stearyl sulfobetaine, lauryl sarcosine, myristyl sarcosine, linoleyl sarcosine, or stearyl sarcosine, linoleyl betaine. Examples include myristyl betaine, or cetyl betaine, lauroamidopropyl betaine, cocamidopropyl betaine, linoleamidopropyl betaine, myristamidopropyl betaine, palmidopropyl betaine, or isostearamidopropyl betaine (e.g., lauroamidopropyl), myristamidopropyl dimethylamine, palmidopropyl dimethylamine, or isostearamidopropyl dimethylamine, sodium methyl cocoyl taurate or disodium methyl oleyl taurate, as well as the MONAQUAT® series (Mona Industries, Inc., Paterson, NJ), polyethyl glycol, polypropyl glycol, and copolymers of ethylene glycol and propylene glycol (e.g., PLURONIC® block copolymer type, e.g., PLURONIC® F-68). In one embodiment, the surfactant as used herein is polysorbate 20. In another embodiment, the surfactant as used herein is poloxamer 188.

[0145] A "pharmaceutically acceptable carrier" refers to a component in a pharmaceutical preparation other than the active ingredient that is non-toxic to the target substance. Pharmaceutically acceptable carriers include, but are not limited to, buffers, excipients, stabilizers, or preservatives.

[0146] As used in this application, the term “prodrug” refers to a precursor or derivative form of a pharmaceutically active substance that is less cytotoxic to tumor cells than the parent drug and can be activated by enzymes or converted into a more active parent form. See, for example, Wilman, “Prodrugs in Cancer Chemotherapy,” Biochemical Society Transactions, 14, pp. 375-382, 615th Meeting Belfast (1986) and Stella et al., “Prodrugs: A Chemical Approach to Targeted Drug Delivery,” Directed Drug Delivery, Borchardt et al. (ed.), pp. 247-267, Humana Press (1985). The prodrugs of the present invention include, but are not limited to, phosphate-containing prodrugs, thiophosphate-containing prodrugs, sulfate-containing prodrugs, peptide-containing prodrugs, D-amino acid-modified prodrugs, glycosylated prodrugs, β-lactam-containing prodrugs, optionally substituted phenoxyacetamide-containing prodrugs or optionally substituted phenylacetamide-containing prodrugs, 5-fluorocytosine, and other 5-fluorouridine prodrugs, and these can be converted into more active cytotoxic free drugs. Examples of cytotoxic agents that can be derivatized into prodrug forms for use in the present invention include, but are not limited to, the chemotherapeutic agents mentioned above.

[0147] The "subjects" are vertebrates, preferably mammals, and more preferably humans. Mammals include, but are not limited to, domesticated animals (such as cattle and sheep), sports animals, companion animals (such as cats, dogs, and horses), primates (e.g., humans and non-human primates such as monkeys (e.g., crab-eating macaques)), and rodents (e.g., mice and rats).

[0148] As used herein, “administer” means a method of giving a certain dosage of a compound (e.g., the anti-tryptase antibody of the present invention or a further therapeutic agent) or composition (e.g., a pharmaceutical composition, e.g., the anti-tryptase antibody of the present invention, and optionally a further therapeutic agent, which may include excipients such as antioxidants (e.g., N-acetyltryptophan and / or methionine)) to a subject. The compositions used in the methods described herein may be administered, for example, intravitreous, intramuscular, intravenous, intradermal, percutaneously, intraarterial, intraperitoneal, intrafocal, intracranial, intraarticular, intraprostatically, intrapleural, intratracheal, intraarachnoid, intranasal, vaginal, rectal, topically, intratumoral, intraperitoneal, subcutaneous, subconjunctival, intravesicular, transmucosal, intrapericardial, intraumbilical, intraocular, intraorbital, transdermally, periorbitally, conjunctiva, subtenon's capsule, anterior chamber, subretinal, retroocular, intracanalicularly, by inhalation, injection, implantation, infusion, continuous infusion, topical perfusion directly into target cells, by catheter, lavage, cream, or lipid composition. The compositions used in the methods described herein may also be administered systemically or topically. The method of administration can vary depending on various factors (e.g., the compound or composition being administered, and the severity of the condition, disease, or disorder being treated).

[0149] The “effective dose” or “therapeutic effective dose” of a drug, such as an anti-tryptase antibody or pharmaceutical preparation (e.g., a pharmaceutical preparation containing an anti-tryptase antibody, which may contain excipients such as antioxidants (N-acetyltryptophan and / or methionine)), refers to the amount effective in achieving the desired therapeutic or preventive outcome over the required period of time at the required dosage. A therapeutic effective dose of an antibody or antibody fragment (e.g., an anti-tryptase antibody), or a composition thereof, can alleviate or treat a disorder or disease, or prevent, reduce, alleviate, or treat symptoms associated with a disorder or disease.

[0150] As used herein, “treatment” (and its grammatical variations such as “to treat” or “to treat”) refers to a clinical intervention aimed at altering the natural course of the individual being treated, which may be performed for preventive purposes or during a clinicopathological process. Desired effects of treatment include, but are not limited to, prevention of disease onset or recurrence, symptom relief, reduction of any direct or indirect pathological consequences of the disease, prevention of metastasis, reduction of the rate of disease progression, recovery or remission of the condition, and remission or improvement of prognosis. In some embodiments, antibodies of the present invention are used to delay the development of the disease or slow the progression of the disease. A patient’s “treatment” of asthma may be considered successful if, for example, after receiving an asthma treatment, the patient shows an observable and / or measurable reduction or absence of one or more of the following: recurrent wheezing, cough, dyspnea, chest tightness, symptoms that occur or worsen at night, symptoms triggered by cold, exercise, or exposure to allergens.

[0151] The term “interleukin-5 (IL-5)” as used herein refers to any natural IL-5 from any vertebrate source, including mammals such as primates (e.g., humans) and rodents (e.g., mice and rats), unless otherwise specified. The term encompasses any form of “full-length” IL-5, including untreated IL-5, mature IL-5, and IL-5 resulting from post-translational modifications. The term also encompasses naturally occurring variants of IL-5, such as splice variants or allele variants. An example amino acid sequence of IL-5 can be found, for example, as UniProtKB acceptance number P05113.

[0152] The term "IL-5 axis-binding antagonist" refers to a molecule that reduces, blocks, inhibits, suppresses, or interferes with the signal transduction resulting from the interaction between IL-5 and one or more of its binding partners, such as the IL-5 receptor alpha (IL5RA). Examples of IL-5 axis-binding antagonists that can be used in the methods of the present invention include, for example, IL-5 binding antagonists (e.g., anti-IL-5 antibodies (e.g., mepolizumab, benralizumab, and reslizumab) and anti-IL-5 receptor binding antagonists (e.g., anti-IL-5R antibodies)).

[0153] As used herein, the term “interleukin-13 (IL-13)” refers to any natural IL-13 from any vertebrate source, including mammals such as primates (e.g., humans) and rodents (e.g., mice and rats), unless otherwise specified. IL-13 is a cytokine secreted by many cell types, including type 2 T helper (Th2) cells. The term encompasses any form of IL-13, including “full-length,” untreated IL-13, mature IL-13, and IL-13 resulting from post-translational modifications. An example amino acid sequence of human IL-13 can be found, for example, under UniProtKB accession number P35225.

[0154] The term "IL-13 axis-binding antagonist" refers to a molecule that reduces, blocks, inhibits, suppresses, or interferes with the signal transduction resulting from the interaction between IL-13 and one or more of its binding partners, such as IL-4 receptor alpha (IL4Rα), IL-13 receptor alpha 1 (IL13RA1), and IL-13 receptor alpha 2 (IL13RA2). Examples of IL-13 axis-binding antagonists include IL-13-binding antagonists (e.g., anti-IL-13 antibodies, e.g., levukizumab, 228B / C-1, 228A-4, 227-26, and 227-43 (see, for example, U.S. Patent Nos. 7,674,459, 8,067,199, 8,088,618, 8,318,160, and 8,734,797) and IL-13 receptor-binding antagonists (e.g., anti-IL4Rα antibody, anti-IL13RA1 antibody, or anti-IL13RA2 antibody).

[0155] As used herein, the term “interleukin-17 (IL-17)” means any natural IL-17 from any vertebrate source, including mammals such as primates (e.g., humans) and rodents (e.g., mice and rats), unless otherwise specified, and includes family members IL-17A, IL-17B, IL-17C, IL-17D, IL-17E, and IL-17F. The term encompasses any form of “full-length” IL-17, untreated IL-17, mature IL-17, and IL-17 resulting from post-translational modifications. An example amino acid sequence of human IL-17A can be found, for example, under UniProtKB acceptance number Q16552. An example amino acid sequence of human IL-17B can be found, for example, under UniProtKB acceptance number Q9UHF5. An example amino acid sequence of human IL-17C can be found, for example, under UniProtKB acceptance number Q9P0M4. The amino acid sequence of an exemplary human IL-17D can be found, for example, under UniProtKB acceptance number Q8TAD2. The amino acid sequence of an exemplary human IL-17E can be found, for example, under UniProtKB acceptance number Q9H293. The amino acid sequence of an exemplary human IL-17F can be found, for example, under UniProtKB acceptance number Q96PD4.

[0156] The term "IL-17 axis-binding antagonist" refers to a molecule that reduces, blocks, inhibits, suppresses, or interferes with the signal transduction resulting from the interaction between IL-17 and one or more of its binding partners, which are interleukin-17 receptor (IL-17R) family member proteins, such as interleukin-17 receptor A (IL17RA), interleukin-17 receptor B (IL17RB), interleukin-17 receptor C (IL17RC), interleukin-17 receptor D (IL17RD), interleukin-17 receptor E (IL17RE), and interleukin-17 receptor E-like (IL17REL). Examples of IL-17 axis-binding antagonists include, for example, IL-17-binding antagonists (e.g., anti-IL-17 antibodies (e.g., secukinumab (AIN417), ixekizumab (LY2439821), bimekizumab, and NI-1401)) and IL-17 receptor-binding antagonists (e.g., anti-IL-17R antibodies (e.g., brodalumab (AMG-827))). See, for example, WO2006 / 013107, WO2007 / 070750, WO2012 / 156219, and U.S. Patent No. 8,715,669.

[0157] As used herein, the term “interleukin-33 (IL-33)” refers to any naturally occurring IL-33 derived from any vertebrate source, including mammals such as primates (e.g., humans and cynomolgus monkeys) and rodents (e.g., mice and rats), unless otherwise specified. IL-33 is also referred in the art to high endothelial venule nuclear factor (NF-HEV, see, e.g., Baekkevold et al. Am.J. Pathol. 163(1):69-79, 2003), DVS27, C9orf26, and interleukin-1 family member 11 (IL-1F11). This term encompasses any form of “full-length” untreated IL-33, mature IL-33, and IL-33 resulting from post-translational modifications. Human full-length, untreated IL-33 contains 270 amino acids (aa). 1-270It is sometimes referred to as such. Examples of human IL-33 processing methods include IL-33 95-270 IL-33 99-270 IL-33 109-270 IL-33 112-270 IL-33 1-178 , and IL-33 179-270 Examples include (Lefrancais et al. Proc. Natl. Acad. Sci. 109(5):1673-1678, 2012 and Martin, Semin. Immunol. 25:449-457, 2013). In some embodiments, the processing method of human IL-33, for example, IL-33 95-270 IL-33 99-270 IL-33 109-270 Other forms, such as those treated with proteases like calpain, proteinase 3, neutrophil elastase, and cathepsin G, may have increased biological activity compared to full-length IL-33. This term also includes naturally occurring variants of IL-33, e.g., splice variants (e.g., the constitutively active splice variant lacking exon 3, spIL-33, Hong et al. J. Biol. Chem. 286(22):20078-20086, 2011) or allele variants. IL-33 may exist intracellularly (e.g., in the nucleus) or as a secreted cytokine. The full-length IL-33 protein contains a helix-turn-helix DNA-binding motif that includes a nuclear localization sequence (amino acids 1-75 of human IL-33), which in turn contains a chromatin-binding motif (amino acids 40-58 of human IL-33). The processed and secreted form of IL-33 lacks these N-terminal motifs. An exemplary amino acid sequence of human IL-33 can be found, for example, as UniProtKB acceptance number O95760.

[0158] The terms “interleukin-1 receptor-like 1 (IL1RL1)” and “ST2,” as used interchangeably herein, refer to any native ST2 from any vertebrate source, including mammals such as primates (e.g., humans) and rodents (e.g., mice and rats), unless otherwise specified. ST2 is also referred to in the art as DER4, T1, and FIT-1. The terms encompass any form of “full-length” ST2, including untreated ST2, mature ST2, and ST2 resulting from post-translational modifications. At least four isoforms of ST2 are known in the art, including soluble (sST2, also known as IL1RL1-a) and transmembrane (ST2L, also known as IL1RL1-b) resulting from differential mRNA expression from dual promoter systems, as well as ST2V and ST2LV resulting from alternative splicing, as described below. The domain structure of ST2L includes three extracellular immunoglobulin-like C2 domains, a transmembrane domain, and a cytoplasmic tol / interleukin-1 receptor (TIR) ​​domain. sST2 lacks the transmembrane and cytoplasmic domains present in ST2L and contains a unique nine-amino acid (aa) C-terminal sequence (see, e.g., Kakkar et al. Nat. Rev. Drug Disc. 7:827-840, 2008). sST2 may function as a decoy receptor inhibiting soluble IL-33. This term also encompasses native variants of ST2, e.g., splice variants (e.g., ST2V lacking a third immunoglobulin motif and possessing a unique hydrophobic tail, and ST2LV lacking the transmembrane domain of ST2L) or allele variants (e.g., variants that protect against or pose an asthmatic risk, as described herein). An exemplary human ST2 amino acid sequence can be found, for example, under UniProtKB accession number Q01638. ST2 is part of the IL-33 receptor along with the co-receptor protein IL-1RAcP.IL-33 binds to ST2 and its co-receptor, interleukin-1 receptor co-protein (IL-1RAcP), forming a 1:1:1 triple signaling complex that promotes downstream signaling (see, for example, Lingel et al. Structure 17(10):1398-1410, 2009, and Liu et al. Proc. Natl. Acad. Sci. 110(37):14918-14924, 2013).

[0159] The "IL-33 axis" refers to nucleic acids (e.g., genes, or mRNA transcribed from genes) or polypeptides involved in IL-33 signal conversion. For example, the IL-33 axis may include the ligand IL-33, the receptor (e.g., ST2 and / or IL-1RAcP), the adapter molecule (e.g., MyD88), or proteins associated with the receptor molecule and / or the adapter molecule (e.g., kinases such as interleukin-1 receptor-related kinase 1 (IRAK1) and interleukin-1 receptor-related kinase 4 (IRAK4), or E3 ubiquitin ligases such as TNF receptor-related factor 6 (TRAF6)).

[0160] An "IL-33 axial-binding antagonist" refers to a molecule that inhibits the interaction between an IL-33 axial-binding partner and one or more of its binding partners. As used herein, an IL-33 axial-binding antagonist includes IL-33 binding antagonists, ST2 binding antagonists, and IL1RAcP binding antagonists.Examples of IL-33 axis-binding antagonists include anti-IL-33 antibodies and their antigen-binding fragments (e.g., anti-IL-33 antibodies such as ANB-020 (Anaptys Bio, Inc.), or EP1725261, US8187596, WO2011 / 031600, WO2014 / 164959, WO2015 / 099175, or WO2015 / 106080, respectively, the whole of which is incorporated herein by reference). Any of the antibodies described in ); polypeptides that bind to IL-33 and / or its receptor (ST2 and / or IL-1RAcP) and block ligand-receptor interactions (e.g., ST2-Fc protein, e.g., as described in WO2014 / 152195 (the whole of which is incorporated herein by reference); immunoadhesins, peptide bodies, and soluble ST2, or derivatives thereof); anti-IL-33 receptor antibodies (e.g., For example, anti-ST2 antibodies, e.g., AMG-282 (Amgen) or STLM15 (Janssen), or any of the anti-ST2 antibodies described in WO2013 / 173761 and WO2013 / 165894 (each of which is incorporated herein by reference in its entirety); or ST2-Fc proteins, e.g., those described in WO2013 / 173761, WO2013 / 165894, or WO2014 / 152195 (each of which is incorporated herein by reference in its entirety); as well as IL-33 receptor antagonists such as small molecule inhibitors, aptamers that bind to IL-33, and nucleic acids that hybridize with the IL-33 axis nucleic acid sequence under stringent conditions (e.g., short interfering RNA (siRNA) or clustered, regularly arranged short palindromic repeat RNA (CRISPR-RNA or crRNA) (Mali Examples include single-stranded guide RNAs (sgRNAs) having crRNA and tracrRNA sequences as described in et al. (Science. 339:823-26, 2013) (the entire work is incorporated herein by reference).

[0161] The terms “anti-IL-33 antibody,” “antibody that binds to IL-33,” and “antibody that specifically binds to IL-33” refer to an antibody that can bind to IL-33 with sufficient affinity to be useful for targeting IL-33 as a diagnostic and / or therapeutic agent. In one embodiment, the degree of binding of an anti-IL-33 antibody to unrelated non-IL-33 proteins is less than about 10% of the binding of this antibody to IL-33, as measured, for example, by radioimmunoassay (RIA). In certain embodiments, the antibody that binds to IL-33 has a viscosity of ≤1 μM, ≤100 nM, ≤10 nM, ≤1 nM, ≤0.1 nM, ≤0.01 nM, or ≤0.001 nM (e.g., 10 -8 M or less, for example, 10 -8 M~10 -13 M, for example, 10 -9 M~10 -13 The dissociation constant (K) of M D ) has. In a particular embodiment, the anti-IL-33 antibody binds to an IL-33 epitope that is conserved among IL-33s from different species.

[0162] The term "ST2-binding antagonist" refers to a molecule that inhibits the interaction of ST2 with IL-33, IL1RAcP, and / or a second ST2 molecule. The ST2-binding antagonist may be a protein such as an "ST2-Fc protein" containing an IL-33 binding domain (e.g., all or part of the ST2 or IL1RAcP protein) and a multimerization domain (e.g., the Fc portion of an immunoglobulin, e.g., isotypes IgG1, IgG2, IgG3, and IgG4, and the Fc domain of IgG selected from any allotype within each isotype group), which is directly or indirectly bound to one another through a linker (e.g., a serine-glycine (SG) linker, a glycine-glycine (GG) linker, or variants thereof (e.g., SGG, GGS, SGS, or GSG linker)), including, but not limited to, the ST2-Fc proteins and variants thereof described in WO2013 / 173761, WO2013 / 165894, and WO2014 / 152195, whose wholes are incorporated herein by reference, respectively. In some embodiments, the ST2-binding antagonist may be an anti-ST2 antibody, such as AMG-282 (Amgen) or STLM15 (Janssen) or any of the anti-ST2 antibodies described in WO2013 / 173761 and WO2013 / 165894.

[0163] "Isolated nucleic acids" refer to nucleic acid molecules separated from components in their natural environment. Isolated nucleic acids typically contain nucleic acid molecules found within cells, but these nucleic acid molecules exist outside of chromosomes or at chromosomal locations different from their natural chromosomal locations.

[0164] The term "regulatory sequence" refers to a DNA sequence required for the expression of a manipulably linked coding sequence in a particular host organism. For example, a regulatory sequence suitable for prokaryotes includes a promoter, optionally an operator sequence, and a ribosome binding site. Eukaryotic cells are known to utilize promoters, polyadenylation signals, and enhancers.

[0165] The terms “host cell,” “host cell line,” and “host cell culture” are used synonymously and refer to cells into which exogenous nucleic acids have been introduced, including the offspring of such cells. Host cells include “transformers” and “transformed cells,” which include primary transformed cells and their offspring, regardless of the number of passages. Offspring may contain mutations, although their nucleic acid content may not be exactly the same as that of the parent cells. Mutant offspring having the same function or biological activity as those screened or selected for the initially transformed cells are included herein.

[0166] Nucleic acids are "operably ligated" when they are placed in a functional relationship with another nucleic acid sequence. For example, DNA for a pre-sequence or secretion leader is operably ligated to DNA for a polypeptide when it is expressed as a pre-protein involved in polypeptide secretion; promoters or enhancers are operably ligated to coding sequences when they affect the transcription of the sequence; or ribosome binding sites are operably ligated to coding sequences when they are positioned to facilitate translation. Generally, "operably ligated" means that the DNA sequences to be ligated are contiguous, and in the case of secretion leaders, contiguous and in the reading phase. Enhancers, however, do not need to be contiguous. Ligation is achieved by ligation at a convenient restriction site. If such a site does not exist, synthetic oligonucleotide adapters or linkers are used according to conventional practice.

[0167] The “amino acid sequence identity percentage (%)” for polypeptide sequences identified herein is defined as the percentage of amino acid residues in a candidate sequence that are identical to amino acid residues in the polypeptide being compared, after the sequences have been aligned and gaps introduced as necessary to obtain the maximum possible sequence identity percentage, with no conservative substitutions considered to be part of the sequence identity. Alignment for the purpose of determining the amino acid sequence identity percentage can be achieved by various means within the scope of the art, for example, using publicly available computer software such as BLAST, BLAST-2, ALIGN, or Megalign (DNASTAR) software. Those skilled in the art can determine appropriate parameters for measuring the alignment, including any algorithm necessary to achieve the maximum alignment over the entire length of the sequences being compared. However, for the purposes herein, the amino acid sequence identity % values ​​are generated using the sequence comparison computer program ALIGN-2. The ALIGN-2 sequence comparison computer program was written by Genentech, Inc., and its source code, along with user documentation, has been filed with the U.S. Copyright Office (Washington DC, 20559) and is registered under U.S. Copyright No. TXU510087. The ALIGN-2 program is publicly available from Genentech, Inc., South San Francisco, California. The ALIGN-2 program should be compiled for use with the UNIX operating system, preferably Digital UNIX V4.0D. All sequence comparison parameters are set by the ALIGN-2 program and do not change.

[0168] In situations where ALIGN-2 is used for amino acid sequence comparison, the amino acid sequence identity percentage of a given amino acid sequence A to, with, or relative to a given amino acid sequence B (or, as can be expressed, a given amino acid sequence A that has, or contains, a specific amino acid sequence identity percentage to, with, or relative to a given amino acid sequence B) is calculated as follows: 100 x fraction X / Y In the formula, X is the number of amino acid residues scored as a perfect match in the alignment of A and B by the sequence alignment program ALIGN-2, and Y is the total number of amino acid residues in B. It is understood that if the length of amino acid sequence A is not equal to the length of amino acid sequence B, the amino acid sequence identity % of A to B is not equal to the amino acid sequence identity % of B to A. Unless otherwise specifically indicated, all amino acid sequence identity % values ​​used herein are obtained using the ALIGN-2 computer program as described in the preceding paragraph.

[0169] Unless otherwise indicated, the amino acid sequences described herein are continuous amino acid sequences.

[0170] When used herein, the term “vector” is intended to refer to a nucleic acid molecule capable of transporting another nucleic acid molecule to which it is bound. One type of vector is a “plasmid,” which refers to a circular double-stranded DNA loop to which further DNA segments can be ligated. Another type of vector is a phage vector. Another type of vector is a viral vector, in which further DNA segments are ligated into the viral genome. Certain vectors are capable of self-replication in the host cell to which they are introduced (e.g., bacterial vectors with bacterial replication and episomal mammalian vectors). Other vectors (e.g., non-episomal mammalian vectors) can be integrated into the host cell's genome upon introduction into the host cell and thus replicate with the host genome. Furthermore, certain vectors are capable of inducing the expression of genes to which they are functionally linked. Such vectors are referred herein as “recombinant expression vectors” (or simply “recombinant vectors” or “expression vectors”). Generally, expression vectors used in recombinant DNA technology are often in the form of plasmids. Hereinafter, “plasmid” and “vector” may be used interchangeably.

[0171] II. Compositions and Methods In one embodiment, the present invention is partially based on a novel antibody that binds to tryptase. In another embodiment, the present invention is partially based on the discovery that certain residues of anti-tryptase antibodies (e.g., HVR residues such as HVR-H3 W100 pointing to W100 in the VH domain of the anti-tryptase antibody hu31A.v11) may be oxidation-sensitive. The present invention provides a pharmaceutical composition comprising an antioxidant (e.g., N-acetyltryptophan and / or methionine) for reducing or preventing the oxidation of the antibodies described herein (e.g., anti-tryptase antibodies). Other suitable antioxidant excipients include, but are not limited to, free tryptophan, cyclodextrin, trolox (6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid), pyridoxine, polyols (e.g., mannitol), and metal chelating agents (e.g., EDTA). See, for example, Ji et al., Biotechnology 98:4485-4500, 2009. The antibodies and pharmaceutical compositions of the present invention are useful for the diagnosis and / or treatment of disorders (e.g., lung disorders, autoimmune disorders, inflammatory disorders, fibrous disorders, granulocytic (neutrophilic or eosinophilic) disorders, monocytic disorders, lymphocytic disorders, or disorders related to an increased number or distribution of normal or abnormal tissue resident cells (such as mast cells, macrophages, or lymphocytes) or stromal cells (such as fibroblasts, myofibroblasts, smooth muscle cells, epithelium, or endothelium), or tryptase-related disorders or tryptase-mediated disorders. In another embodiment, the present invention provides lyophilized pharmaceutical compositions for reducing or eliminating the oxidation of antibodies described herein (e.g., anti-tryptase antibodies).

[0172] A. Exemplary anti-tryptase antibody The present invention provides an isolated antibody that binds to tryptase. In certain embodiments, the anti-tryptase antibody of the present invention has a K content of about 100 nM or less (e.g., 100 nM or less, 10 nM or less, 1 nM or less, 100 pM or less, 10 pM or less, 1 pM or less, or 0.1 pM or less) DIt binds to tryptase. In some embodiments, the antibody has a K content of 10 nM or less (e.g., 10 nM or less, 1 nM or less, 100 pM or less, 10 pM or less, 1 pM or less, or 0.1 pM or less). D It binds to tryptase. In some embodiments, the antibody has a K content of 1 nM or less (e.g., 1 nM or less, 100 pM or less, 10 pM or less, 1 pM or less, or 0.1 pM or less). D It binds to tryptase. In some embodiments, the antibody has a K content of 0.5 nM or less (e.g., 0.5 nM or less, 400 pM or less, 300 pM or less, 200 pM or less, 100 pM or less, 50 pM or less, 25 pM or less, 10 pM or less, 1 pM or less, or 0.1 pM or less). D It binds to tryptase. In some embodiments, the antibody has a K content of about 0.1 nM to about 0.5 nM (e.g., about 0.1 nM, about 0.2 nM, about 0.3 nM, about 0.4 nM, or about 0.5 nM). DIt binds to trypsin. In some embodiments, the antibody has a K of about 1 pM to about 500 pM, about 1 pM to about 400 pM, about 1 pM to about 300 pM, about 1 pM to about 200 pM, about 1 pM to about 100 pM, about 1 pM to about 50 pM, about 25 pM to about 500 pM, about 25 pM to about 400 pM, about 25 pM to about 300 pM, about 25 pM to about 100 pM, about 50 pM to about 500 pM, about 50 pM to about 450 pM, about 50 pM to about 425 pM, about 50 pM to about 400 pM, about 50 pM to about 375 pM, about 50 pM to about 350 pM, about 50 pM to about 325 pM, about 50 pM to about 300 pM, about 50 pM to about 275 pM, about 50 pM to about 250 pM, about 50 pM to about 200 pM, about 50 pM to about 180 pM, about 50 pM to about 175 pM, about 50 pM to about 150 pM, about 50 pM to about 125 pM, about 50 pM to about 75 pM, about 100 pM to about 500 pM, about 100 pM to about 475 pM, about 100 pM to about 450 pM, about 100 pM to about 425 pM, about 100 pM to about 400 pM, about 100 pM to about 375 pM, about 100 pM to about 350 pM, about 100 pM to about 325 pM, about 100 pM to about 300 pM, about 100 pM to about 275 pM, about 100 pM to about 250 pM, about 100 pM to about 225 pM, about 100 pM to about 200 pM, about 100 pM to about 180 pM, about 100 pM to about 175 pM, about 100 pM to about 150 pM, about 100 pM to about 125 pM, about 150 pM to about 500 pM, about 150 pM to about 475 pM, about 150 pM to about 450 pM, about 150 pM to about 425 pM, about 150 pM to about 400 pM, about 150 pM to about 375 pM, about 150 pM to about 350 pM, about 150 pM to about 325 pM, about 150 pM to about 300 pM, about 150 pM to about 375 pM, about 150 pM to about 350 pM, about 150 pM to about 325 pM, about 150 pM to about 300 pM, about 150 pM to about 275 pM, about 150 pM to about 225 pM, about 150 pM to about 200 pM, about 175 pM to about 500 pM, about 175 pM to about 475 pM, about 175 pM to about 450 pM, about 175 pM to about 425 pM, about 175 pM to about 400 pM, about 175 pM to about 375 pM, about 175 pM to about 350 pM, about 175 pM to about 325 pM, about 175 pM to about 300 pM, or about 180 pM to about 400 pM DIt binds to tryptase. In some embodiments, the antibody is about 0.4 nM K D It binds to tryptase. In some embodiments, the antibody is about 0.2 nM K D It binds to tryptase. In some embodiments, the antibody is about 0.18 nM K D It binds to the tryptase. In some embodiments, the tryptase is human tryptase, for example, human tryptase beta (e.g., human tryptase beta 1, human tryptase beta 2, and / or human tryptase beta 3). In some embodiments, K D This is determined by the BIACORE® SPR assay. In certain embodiments, the tryptase is human tryptase alpha. In certain embodiments, the antibody is human or humanized antibody.

[0173] In other embodiments, in some embodiments, the anti-tryptase antibodies of the present invention (including any of the aforementioned anti-tryptase antibodies) can inhibit tryptase activity. In some embodiments, the anti-tryptase antibodies of the present invention can inhibit the proteolytic activity of tryptase, for example, as determined in an in vitro tryptase enzyme assay. In some embodiments, an artificial substrate, such as the synthetic peptide S-2288, can be used as a substrate in an in vitro tryptase enzyme assay. In some embodiments, the anti-tryptase antibodies of the present invention can inhibit human tryptase activity at half-percentage inhibitory concentrations (IC50) of about 100 nM or less (e.g., ≤100 nM, ≤10 nM, ≤5 nM, ≤2.5 nM, ≤1 nM, ≤100 pM, ≤10 pM, ≤1 pM, or ≤0.1 pM) as determined by an in vitro tryptase enzyme assay using S-2288 as a substrate. In some embodiments, the antibody can inhibit human tryptase activity with an IC50 of approximately 10 nM or less (e.g., ≤10 nM, ≤5 nM, ≤2.5 nM, ≤1 nM, ≤100 pM, ≤10 pM, ≤1 pM, ≤1 pM, or ≤0.1 pM) when determined, for example, by an in vitro tryptase enzyme assay using S-2288 as a substrate. In some embodiments, the antibody can inhibit human tryptase activity with an IC50 of approximately 2.5 nM or less (e.g., ≤2.5 nM, ≤1 nM, ≤100 pM, ≤10 pM, ≤1 pM, or ≤0.1 pM) when determined, for example, by an in vitro tryptase enzyme assay using S-2288 as a substrate. In some embodiments, antibodies can inhibit the activity of human tryptase with an IC50 of about 0.1 nM to about 2 nM (e.g., about 0.1 nM, about 0.2 nM, about 0.3 nM, about 0.4 nM, about 0.5 nM, about 0.6 nM, about 0.7 nM, about 0.8 nM, about 0.9 nM, about 1.0 nM, about 1.1 nM, about 1.2 nM, about 1.3 nM, about 1.4 nM, about 1.5 nM, about 1.6 nM, about 1.7 nM, about 1.8 nM, about 1.9 nM, or about 2.0 nM).In some embodiments, antibodies can inhibit the activity of human tryptase with an IC50 of about 0.5 nM to about 2.5 nM (e.g., about 0.5 nM, about 0.6 nM, about 0.7 nM, about 0.8 nM, about 0.9 nM, about 1.0 nM, about 1.1 nM, about 1.2 nM, about 1.3 nM, about 1.4 nM, about 1.5 nM, about 1.6 nM, about 1.7 nM, about 1.8 nM, about 1.9 nM, about 2.0 nM, about 2.1 nM, about 2.2 nM, about 2.3 nM, about 2.4 nM, or about 2.5 nM).In some embodiments, the antibody concentration is approximately 1 pM to 2.5 nM, 25 pM to 2.5 nM, 50 pM to 2.5 nM, 75 pM to 2.5 nM, 100 pM to 2.5 nM, 125 pM to 2.5 nM, 150 pM to 2.5 nM, 175 pM to 2.5 nM, 200 pM to 2.5 nM, 225 pM to 2.5 nM, 250 pM to 2.5 nM, 300 pM to 2.5 nM, 325 pM to 2.5 nM, 325 pM to 2.5 nM, and 350 pM. ~about 2.5nM, about 375pM to about 2.5nM, about 400pM to about 2.5nM, about 425pM to about 2.5nM, about 450pM to about 2.5nM, about 500pM to about 2.5nM, about 450pM to about 2.5nM, about 500pM to about 2.5nM, about 550pM to about 2.5nM, about 600pM to about 2.5nM, about 650pM to about 2.5nM, about 700pM to about 2.5nM, about 750pM to about 2.5nM, about 800pM to about 2.5nM, about 850pM to about 2.5nM, about 900pM to about 2. 5nM, about 950pM to about 2.5nM, about 1nM to about 2.5nM, about 1.1nM to about 2.5nM, about 1.2nM to about 2.5nM, about 1.3nM to about 2.5nM, about 1.4nM to about 2.5nM, about 1.5nM to about 2.5nM, about 1.6nM ~about 2.5nM, about 1.7nM to about 2.5nM, about 1.8nM to about 2.5nM, about 1.9nM to about 2.5nM, about 2.0nM to about 2.5nM, about 2.1nM to about 2.5nM, about 2.2nM to about 2.5nM, about 2.3nM to about 2.5nM, about Human tryptase activity can be inhibited with an IC50 of 500 pM to approximately 1.9 pM, approximately 750 pM to approximately 1.9 pM, approximately 1 nM to approximately 1.9 pM, approximately 1.25 nM to approximately 1.9 pM, approximately 1.5 nM to approximately 1.9 pM, approximately 1 nM to approximately 1.85 nM, approximately 1.25 nM to approximately 1.85 nM, approximately 1.5 nM to approximately 1.85 nM, approximately 1 nM to approximately 1.8 nM, approximately 1.25 nM to approximately 1.8 nM, approximately 1.5 nM to approximately 1.8 nM, or approximately 1.6 nM to approximately 1.8 nM. In some embodiments, antibodies can inhibit human tryptase activity with an IC50 of approximately 1.8 nM.In other embodiments, antibodies can inhibit the activity of human tryptase with an IC50 of about 0.5 nM to about 1 nM (e.g., about 0.5 nM, about 0.6 nM, about 0.7 nM, about 0.8 nM, about 0.9 nM, or about 1.0 nM).In some embodiments, the antibody is approximately 1 pM to 1 nM, approximately 25 pM to 1 nM, approximately 50 pM to 1 nM, approximately 75 pM to 1 nM, approximately 100 pM to 1 nM, approximately 125 pM to 1 nM, approximately 150 pM to 1 nM, approximately 175 pM to 1 nM, approximately 200 pM to 1 nM, approximately 225 pM to 1 nM, approximately 250 pM to 1 nM, approximately 300 pM to 1 nM, approximately 325 pM to 1 nM, approximately 350 pM to 1 nM, approximately 375 pM to 1 nM, approximately 400 pM to 1 nM, approximately 425 pM to 1 nM, and approximately 450 pM to 1 nM. , about 500pM to about 1nM, about 450pM to about 1nM, about 500pM to about 1nM, about 550pM to about 1nM, about 600pM to about 1nM, about 650pM to about 1nM, about 700pM to about 1nM, about 750pM, about 250pM to about 800pM, about 300pM Approximately 800pM, approximately 325pM to approximately 800pM, approximately 325pM to approximately 800pM, approximately 350pM to approximately 800pM, approximately 375pM to approximately 800pM, approximately 400pM to approximately 800pM, approximately 425pM to approximately 800pM, approximately 450pM to approximately 800pM, approximately 500pM to approximately 800pM, about 450pM to about 800pM, about 500pM to about 800pM, about 550pM to about 800pM, about 600pM to about 800pM, about 650pM to about 800pM, about 700pM to about 800pM, about 750pM to about 800pM, about 1pM to about 600 pM, approx. 25 pM ~ approx. 600 pM, approx. 50 pM ~ approx. 600 pM, approx. 75 pM ~ approx. 600 pM, approx. 100 pM ~ approx. 600 pM, approx. 125 pM ~ approx. 600 pM, approx. Human tryptase activity can be inhibited with an IC50 of 225 pM to approximately 600 pM, approximately 250 pM to approximately 600 pM, approximately 300 pM to approximately 600 pM, approximately 325 pM to approximately 600 pM, approximately 325 pM to approximately 600 pM, approximately 350 pM to approximately 600 pM, approximately 375 pM to approximately 600 pM, approximately 400 pM to approximately 600 pM, approximately 425 pM to approximately 600 pM, approximately 450 pM to approximately 600 pM, approximately 500 pM to approximately 600 pM, approximately 450 pM to approximately 600 pM, approximately 500 pM to approximately 600 pM, or approximately 550 pM to approximately 600 pM. In some embodiments, antibodies can inhibit human tryptase activity with an IC50 of approximately 0.6 nM.In some embodiments, the tryptase is human tryptase, e.g., human tryptase beta (e.g., human tryptase beta 1, human tryptase beta 2, and / or human tryptase beta 3). In some examples, the inhibitory activity of the antibody is determined, as described herein, for example, by examples (e.g., Example 1, specifically section (A)(viii)(a)) or by other approaches known in the art. In certain embodiments, the antibody is human or humanized antibody. In some embodiments, the antibody can inhibit the activity of human tryptase as a monovalent antibody or its antigen-binding antibody fragment (e.g., Fab). In other embodiments, the antibody can inhibit the activity of human tryptase as a bivalent antibody (e.g., IgG antibody (e.g., IgG1 or IgG4 antibody) or F(ab')2).

[0174] In some cases, any of the anti-tryptase antibodies described herein may inhibit tryptase-stimulated contraction of human early airway smooth muscle cells. In other cases, any of the anti-tryptase antibodies described herein may inhibit tryptase-stimulated contraction of human early airway smooth muscle cells. In yet another case, any of the anti-tryptase antibodies described herein may inhibit tryptase or IgE-stimulated mast cell degranulation and / or histamine release. In yet another case, any of the anti-tryptase antibodies described herein may reduce the amount of active tryptase (e.g., in a sample such as bronchoalveolar lavage fluid or nasal absorption sample) upon administration to a subject. For example, any of the anti-tryptase antibodies described herein can reduce the amount of active tryptase by about 1%, about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 75%, about 80%, about 90%, about 95%, about 96%, about 97%, about 98%, about 99%, or more. The reduction may be related to a reference amount of active tryptase, for example, the amount of active tryptase in the sample before administration of the anti-tryptase antibody.

[0175] In some examples, the antibody (e.g., anti-tryptase antibody) is: (a) HVR-H1 containing the amino acid sequence X1X2GMX3 (SEQ ID NO: 1) where X1 is Asp or Ser, X2 is Tyr or Phe, and X3 is Val or His; (b) HVR-H2 containing the amino acid sequence FISSGSSTVYYADTMKG (SEQ ID NO: 2); (c) HVR-H3 containing the amino acid sequence RX1X2X3DWYFDV (SEQ ID NO: 3) where X1 is Asn or Asp, X2 is Tyr or Asn, and X3 is Asp or Tyr; (d) HVR-L1 containing the amino acid sequence SASSSVTYMY (SEQ ID NO: 4); (e) This may include at least one, two, three, four, five, or six hypervariable regions (HVRs) selected from HVR-L2 containing the amino acid sequence of RTSDLAS (SEQ ID NO: 5) and HVR-L3 containing the amino acid sequence of (f)QHYHSYPLT (SEQ ID NO: 6), or a combination of one or more of the above HVRs, as well as one or more variants thereof having at least approximately 80% sequence identity with any of SEQ ID NOs. 1 to 6 (e.g., 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity).

[0176] For example, in some embodiments, the antibody (e.g., anti-tryptase antibody) is (a) HVR-H1 containing the amino acid sequence of DYGMV (SEQ ID NO: 7), (b) HVR-H2 containing the amino acid sequence of FISSGSSTVYYADTMKG (SEQ ID NO: 2), (c) HVR-H3 containing the amino acid sequence of RNYDDWYFDV (SEQ ID NO: 8), (d) HVR-L1 containing the amino acid sequence of SASSSVTYMY (SEQ ID NO: 4), (e) HVR-L2 containing the amino acid sequence of RTSDLAS (SEQ ID NO: 5), and (f) QHYHSYPLT (SEQ ID NO: 6). This may include at least one, two, three, four, five, or six hypervariable regions (HVRs) selected from HVR-L3 containing the amino acid sequence of ), or a combination of one or more of the above HVRs, and one or more variants thereof having at least about 80% sequence identity with any one of SEQ ID NOs. 2 or 4-8 (e.g., 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity).

[0177] In one particular embodiment, in some embodiments, the antibody (e.g., anti-tryptase antibody) may include (a) HVR-H1 containing the amino acid sequence of DYGMV (SEQ ID NO: 7), (b) HVR-H2 containing the amino acid sequence of FISSGSSTVYYADTMKG (SEQ ID NO: 2), (c) HVR-H3 containing the amino acid sequence of RNYDDWYFDV (SEQ ID NO: 8), (d) HVR-L1 containing the amino acid sequence of SASSSVTYMY (SEQ ID NO: 4), (e) HVR-L2 containing the amino acid sequence of RTSDLAS (SEQ ID NO: 5), and (f) HVR-L3 containing the amino acid sequence of QHYHSYPLT (SEQ ID NO: 6). In some embodiments, the antibody (e.g., an anti-tryptase antibody) comprises (a) a heavy chain variable (VH) domain containing an amino acid sequence having at least 90% sequence identity with the amino acid sequence of SEQ ID NO: 9 (e.g., at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%), or (b) a light chain variable (VL) domain containing an amino acid sequence having at least 90% sequence identity with the amino acid sequence of SEQ ID NO: 10 (e.g., at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%), or (c) a VH domain as in (a) and a VL domain as in (b). In some embodiments, the antibody (e.g., an anti-tryptase antibody) comprises one, two, three, or four of the following heavy chain framework regions (FRs): (a) FR-H1 comprising the amino acid sequence of EVQLVESGGGLVQPGGSLRLSCAASGFTFS (SEQ ID NO: 11), (b) FR-H2 comprising the amino acid sequence of WVRQAPGKGLEWVA (SEQ ID NO: 12), (c) FR-H3 comprising the amino acid sequence of RFTISRDNSKNTLYLQMNSLRAEDTAVYYCTR (SEQ ID NO: 13), and (d) FR-H4 comprising the amino acid sequence of WGQGTLVTVSS (SEQ ID NO: 14).In some embodiments, the antibody (e.g., an anti-tryptase antibody) comprises one, two, three, or four of the following light chains FR: (a) FR-L1 containing the amino acid sequence of DIQMTQSPSSLSASVGDRVTITC (SEQ ID NO: 15), (b) FR-L2 containing the amino acid sequence of WYQQKPGKSPKPWIY (SEQ ID NO: 16), (c) FR-L3 containing the amino acid sequence of GVPSRFSGSGSGTDFTLTISSLQPEDFATYYC (SEQ ID NO: 17), and (d) FR-L4 containing the amino acid sequence of FGQGTKVEIK (SEQ ID NO: 18). In some embodiments, the antibody (e.g., an anti-tryptase antibody) comprises a VH domain containing the amino acid sequence of SEQ ID NO: 9, such as antibody hu31A.v11, and a VL domain containing the amino acid sequence of SEQ ID NO: 10.

[0178] In other specific embodiments, in some embodiments, the antibody (e.g., anti-tryptase antibody) may include (a) HVR-H1 containing the amino acid sequence of DYGMV (SEQ ID NO: 7), (b) HVR-H2 containing the amino acid sequence of FISSGSSTVYYADTMKG (SEQ ID NO: 2), (c) HVR-H3 containing the amino acid sequence of RDNYDWYFDV (SEQ ID NO: 29), (d) HVR-L1 containing the amino acid sequence of SASSSVTYMY (SEQ ID NO: 4), (e) HVR-L2 containing the amino acid sequence of RTSDLAS (SEQ ID NO: 5), and (f) HVR-L3 containing the amino acid sequence of QHYHSYPLT (SEQ ID NO: 6). In some embodiments, the antibody (e.g., an antitryptase antibody) comprises (a) a heavy chain variable (VH) domain containing an amino acid sequence having at least 90% sequence identity with the amino acid sequence of SEQ ID NO: 19 (e.g., at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%), or (b) a light chain variable (VL) domain containing an amino acid sequence having at least 90% sequence identity with the amino acid sequence of SEQ ID NO: 20 (e.g., at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%), or (c) a VH domain as in (a) and a VL domain as in (b). In some embodiments, the antibody (e.g., an anti-tryptase antibody) comprises one, two, three, or four of the following heavy chain framework regions (FRs): (a) FR-H1 comprising the amino acid sequence of EVKLVESGGGSVQPGGSRKLSCAASGFTFS (SEQ ID NO: 21), (b) FR-H2 comprising the amino acid sequence of WVRQAPGKGLEWVA (SEQ ID NO: 22), (c) FR-H3 comprising the amino acid sequence of RFTISRDNPKNTLFLQMSSLRSEDTAMYYCAR (SEQ ID NO: 23), and (d) FR-H4 comprising the amino acid sequence of WGTGTTVTVSS (SEQ ID NO: 24).In some embodiments, the antibody (e.g., an anti-tryptase antibody) comprises one, two, three, or four of the following light chains FR: (a) FR-L1 containing the amino acid sequence QIVLTQSPAIMSASPGEKVTISC (SEQ ID NO: 25), (b) FR-L2 containing the amino acid sequence WYQQKPGSSPKPWIY (SEQ ID NO: 26), (c) FR-L3 containing the amino acid sequence GVPARFSGSGSGTSYSLTISSMEAEDAATYYC (SEQ ID NO: 27), and (d) FR-L4 containing the amino acid sequence FGAGTKLELK (SEQ ID NO: 28). In some embodiments, the antibody (e.g., an anti-tryptase antibody) comprises a VH domain containing the amino acid sequence of SEQ ID NO: 19, such as antibody 31a, and a VL domain containing the amino acid sequence of SEQ ID NO: 20.

[0179] In some examples, an antibody (e.g., an antitryptase antibody) comprises (a) a heavy chain variable (VH) domain containing an amino acid sequence having at least 90% sequence identity (e.g., at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%) to any one of the amino acid sequences of SEQ ID NOs. 9, 101, 102, 103, and 104, or an amino acid sequence having that sequence; (b) a light chain variable (VL) domain containing an amino acid sequence having at least 90% sequence identity (e.g., at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%) to any one of the amino acid sequences of SEQ ID NOs. 10, 105, and 106, or an amino acid sequence having that sequence; or (c) a VH domain as in (a) and a VL domain as in (b). For example, in some cases, the antibody contains a VH domain containing the amino acid sequence of SEQ ID NO: 98 and a VL domain containing the amino acid sequence of SEQ ID NO: 102. In other cases, the antibody contains a VH domain containing the amino acid sequence of SEQ ID NO: 98 and a VL domain containing the amino acid sequence of SEQ ID NO: 10. In other cases, the antibody contains a VH domain containing the amino acid sequence of SEQ ID NO: 98 and a VL domain containing the amino acid sequence of SEQ ID NO: 103. In other cases, the antibody contains a VH domain containing the amino acid sequence of SEQ ID NO: 99 and a VL domain containing the amino acid sequence of SEQ ID NO: 102. In other cases, the antibody contains a VH domain containing the amino acid sequence of SEQ ID NO: 99 and a VL domain containing the amino acid sequence of SEQ ID NO: 10. In other cases, the antibody contains a VH domain containing the amino acid sequence of SEQ ID NO: 99 and a VL domain containing the amino acid sequence of SEQ ID NO: 103. In other cases, the antibody contains a VH domain containing the amino acid sequence of SEQ ID NO: 100 and a VL domain containing the amino acid sequence of SEQ ID NO: 102. In other examples, the antibody includes a VH domain containing the amino acid sequence of SEQ ID NO: 100 and a VL domain containing the amino acid sequence of SEQ ID NO: 10. In other examples, the antibody includes a VH domain containing the amino acid sequence of SEQ ID NO: 100 and a VL domain containing the amino acid sequence of SEQ ID NO: 103. In other examples, the antibody includes a VH domain containing the amino acid sequence of SEQ ID NO: 9 and a VL domain containing the amino acid sequence of SEQ ID NO: 102.In other examples, the antibody includes a VH domain containing the amino acid sequence of SEQ ID NO: 9 and a VL domain containing the amino acid sequence of SEQ ID NO: 10. In other examples, the antibody includes a VH domain containing the amino acid sequence of SEQ ID NO: 9 and a VL domain containing the amino acid sequence of SEQ ID NO: 103. In other examples, the antibody includes a VH domain containing the amino acid sequence of SEQ ID NO: 101 and a VL domain containing the amino acid sequence of SEQ ID NO: 102. In other examples, the antibody includes a VH domain containing the amino acid sequence of SEQ ID NO: 101 and a VL domain containing the amino acid sequence of SEQ ID NO: 10. In other examples, the antibody includes a VH domain containing the amino acid sequence of SEQ ID NO: 101 and a VL domain containing the amino acid sequence of SEQ ID NO: 103.

[0180] In some examples, one of the aforementioned antibodies binds to an epitope on human tryptase beta 1 containing at least one, at least two, at least three, or at least four residues selected from the group consisting of His51, Val80, Lys81, and Asp82 of SEQ ID NO: 71. In some embodiments, the antibody binds to an epitope on human tryptase beta 1 containing at least one, at least two, at least three, or at least four residues selected from the group consisting of His51, Val80, Lys81, and Asp82 of SEQ ID NO: 71. In some embodiments, the antibody binds to an epitope on human tryptase beta 1 containing His51 of SEQ ID NO: 71 and at least one, at least two, or at least three residues selected from the group consisting of Val80, Lys81, and Asp82. In some embodiments, the epitope on human tryptase beta 1 further comprises one or more amino acid residues selected from the group consisting of Gln67, Leu83, Ala84, Ala85, Arg87, Pro103, Val104, Ser105, Arg106, Glu128, Glu129, and Pro130 of SEQ ID NO: 71. In some embodiments, the epitope on human tryptase beta 1 comprises at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, at least ten, at least eleven, or all twelve amino acid residues selected from the group consisting of Gln67, Leu83, Ala84, Ala85, Arg87, Pro103, Val104, Ser105, Arg106, Glu128, Glu129, and Pro130 of SEQ ID NO: 71. In some embodiments, the epitopes on human tryptase beta 1 include His51, Gln67, Val80, Lys81, Asp82, Leu83, Ala84, Ala85, Arg87, Pro103, Val104, Ser105, Arg106, Glu128, Glu129, and Pro130 of SEQ ID NO: 71. In some embodiments, the epitopes are associated with human tryptase beta 1 monomers or tetramers. In some embodiments, the epitopes are determined by X-ray crystallography models.In some embodiments, the antibody can dissociate both the small interface and the large interface of the tetrameric human tryptase beta 1.

[0181] In some examples, any of the aforementioned anti-tryptase antibodies contain a paratope that binds to tryptase (e.g., human tryptase beta-1) containing one or more amino acid residues selected from the group consisting of light chain variable region amino acid residues Val30, Thr31, Tyr32, Tyr34, Arg50, Tyr90, His92, Ser93, and Tyr94, and heavy chain variable region amino acid residues Phe50, Ser52, Gly53, Ser54, Ser55, Thr56, Tyr58, Arg95, Tyr97, and Asp98 (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or 19 amino acid residues).

[0182] For example, in some cases, anti-tryptase antibodies contain a paratope that binds to tryptase (e.g., human tryptase beta-1) containing light chain variable region amino acid residues Val30, Thr31, Tyr32, Tyr34, Arg50, Tyr90, His92, Ser93, and Tyr94 or heavy chain variable region amino acid residues Phe50, Ser52, Gly53, Ser54, Ser55, Thr56, Tyr58, Arg95, Tyr97, and Asp98. In some cases, anti-tryptase antibodies contain a paratope that binds to tryptase (e.g., human tryptase beta-1) containing the light chain variable region amino acid residues Val30, Thr31, Tyr32, Tyr34, Arg50, Tyr90, His92, Ser93, and Tyr94, as well as the heavy chain variable region amino acid residues Phe50, Ser52, Gly53, Ser54, Ser55, Thr56, Tyr58, Arg95, Tyr97, and Asp98.

[0183] In some examples, antibodies (e.g., anti-tryptase antibodies) include: (a) HVR-H1 containing the amino acid sequence of GYAIT (SEQ ID NO: 30), (b) HVR-H2 containing the amino acid sequence of GISSAATTFYSSWAKS (SEQ ID NO: 31), (c) HVR-H3 containing the amino acid sequence of DPRGYGAALDRLDL (SEQ ID NO: 32), (d) HVR-L1 containing the amino acid sequence of QSIKSVYNNRLG (SEQ ID NO: 33), (e) HVR-L2 containing the amino acid sequence of ETSILTS (SEQ ID NO: 34), and (f) AGGFDRSGDTT (sequence) This may include at least one, two, three, four, five, or six hypervariable regions (HVRs) selected from HVR-L3 containing the amino acid sequence of number 35), or a combination of one or more of the above HVRs, and one or more variants thereof having at least about 80% sequence identity with any one of sequence numbers 30-35 (e.g., 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity).

[0184] In some examples, an antibody (e.g., an antitryptase antibody) comprises (a) a heavy chain variable (VH) domain containing an amino acid sequence having at least 90% sequence identity (e.g., at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%) to any one of the amino acid sequences of SEQ ID NOs. 36, 47, 48, 49, 50, 51, and 52, or an amino acid sequence having that sequence; (b) a light chain variable (VL) domain containing an amino acid sequence having at least 90% sequence identity (e.g., at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%) to any one of the amino acid sequences of SEQ ID NOs. 37, 53, 58, or 59, or an amino acid sequence having that sequence; or (c) a VH domain as in (a) and a VL domain as in (b).

[0185] In some examples, one of the aforementioned antibodies (e.g., anti-tryptase antibodies) contains an amino acid sequence having at least 90% sequence identity (e.g., at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%) of the following heavy chain framework regions (FRs): (a) FR-H1 containing an amino acid sequence having at least 90% sequence identity (e.g., at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%) of the It may include one, two, three, or four of the following: FR-H3 containing an amino acid sequence having at least 90% sequence identity (e.g., at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%) to the amino acid sequence of RX1TISX2DTSKNQX3SLKLSSVTAADTAVYX4CAR (SEQ ID NO: 40), where X2 is l or Ser, X3 is Arg or Val, X3 is Val or Phe, and X4 is Tyr or Phe; and FR-H4 containing an amino acid sequence having at least 90% sequence identity (e.g., at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%) to the amino acid sequence of (d)WGQGTLVTVSS (SEQ ID NO: 41), where X2 is Arg or Val, X3 is Val or Phe, and X4 is Tyr or Phe.

[0186] For example, in some cases, any of the aforementioned antibodies (e.g., anti-tryptase antibodies) may contain one, two, three, or four of the following heavy chain FRs: (a) FR-H1 containing the amino acid sequence of EVQLVESGPGLVKPSETLSLTCTVSRFSLI (SEQ ID NO: 38), (b) FR-H2 containing the amino acid sequence of WIRQPPGKGLEWIG (SEQ ID NO: 42), (c) FR-H3 containing the amino acid sequence of RVTISRDTSKNQVSLKLSSVTAADTAVYYCAR (SEQ ID NO: 43), and (d) FR-H4 containing the amino acid sequence of WGQGTLVTVSS (SEQ ID NO: 41).

[0187] In other embodiments, in some examples, any of the aforementioned antibodies (e.g., anti-tryptase antibodies) may comprise one, two, three, or four of the following heavy chain FRs: (a) FR-H1 containing the amino acid sequence of EVQLVESGGGLVQPGGSLRLSCAVSRFSLI (SEQ ID NO: 44), (b) FR-H2 containing the amino acid sequence of WVRQAPGKGLEWIG (SEQ ID NO: 45), (c) FR-H3 containing the amino acid sequence of RSTISRDTSKNTVYLQMNSLRAEDTAVYFCAR (SEQ ID NO: 46), and (d) FR-H4 containing the amino acid sequence of WGQGTLVTVSS (SEQ ID NO: 41).

[0188] In other embodiments, in some examples, any of the aforementioned antibodies (e.g., anti-tryptase antibodies) may comprise one, two, three, or four of the following heavy chain FRs: (a) FR-H1 containing the amino acid sequence of EVQLVESGGGLVQPGGSLRLSCAVSRFSLI (SEQ ID NO: 44), (b) FR-H2 containing the amino acid sequence of WVRQAPGKGLEWIG (SEQ ID NO: 45), (c) FR-H3 containing the amino acid sequence of RSTISRDTSKNTVYLQMNSLRAEDTAVYFCAR (SEQ ID NO: 46), and (d) FR-H4 containing the amino acid sequence of WGQGTLVTVSS (SEQ ID NO: 41).

[0189] In some cases, one of the aforementioned antibodies (e.g., anti-tryptase antibodies) has at least 90% sequence identity (e.g., at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity) to the amino acid sequence of DX1QX2TQSPSSLSASVGDRVTITC (SEQ ID NO: 60), where X1 is Ile or Ala and X2 is Met or Leu, or FR-L1 containing an amino acid sequence having that sequence, or (b) at least 90% sequence identity (e.g., at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity) to the amino acid sequence of WYQQKPGKX1PKLLIY (SEQ ID NO: 61), where X1 is Ala or Pro, or the sequence having that sequence (c)FR-L2 containing an amino sequence having (c)X1 is Gly or Glu and X2 is Tyr or Phe, with at least 90% sequence identity (e.g., at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity) of the amino acid sequence VPSRFSGSGSX1TDFTLTISSLQPEDFATYX2C (SEQ ID NO: 62), or FR-L3 containing an amino sequence having the same sequence, and (d)FR-L4 containing an amino sequence having at least 90% sequence identity (e.g., at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity) of the amino acid sequence FGQGTKVEIK (SEQ ID NO: 63), or FR-L4 containing an amino sequence having the same sequence.

[0190] For example, in a particular case, any of the aforementioned antibodies (e.g., anti-tryptase antibodies) may contain one, two, three, or four of the following light chain FRs: (a) FR-L1 containing the amino acid sequence DIQMTQSPSSLSASVGDRVTITC (SEQ ID NO: 64), (b) FR-L2 containing the amino acid sequence WYQQKPGKAPKLLIY (SEQ ID NO: 65), (c) FR-L3 containing the amino acid sequence GVPSRFSGSGSGTDFTLTISSLQPEDFATYYC (SEQ ID NO: 66), and (d) FR-L4 containing the amino acid sequence FGQGTKVEIK (SEQ ID NO: 63).

[0191] In some embodiments, any of the aforementioned antibodies (e.g., anti-tryptase antibodies) may comprise one, two, three, or four of the following light chains FR: (a) FR-L1 comprising the amino acid sequence AAVLTQTPASVSAAVGGTVSISC (SEQ ID NO: 67), (b) FR-L2 comprising the amino acid sequence WYQQKPGQPPKLLIY (SEQ ID NO: 68), (c) FR-L3 comprising the amino acid sequence GVPSRFKGSGSETQFTLTISDVQX1DDAATYFC (SEQ ID NO: 69) where X1 is Cys or Ala, and (d) FR-L4 comprising the amino acid sequence FGQGTKVEIK FGGGTEVVVK (SEQ ID NO: 70).

[0192] In some examples, an antibody (e.g., an antitryptase antibody) comprises (a) a heavy chain variable (VH) domain containing an amino acid sequence having at least 90% sequence identity (e.g., at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%) to any one of the amino acid sequences of SEQ ID NOs. 36, 47, 48, 49, 50, 51, and 52, or an amino acid sequence having that sequence; (b) a light chain variable (VL) domain containing an amino acid sequence having at least 90% sequence identity (e.g., at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%) to any one of the amino acid sequences of SEQ ID NOs. 37, 53, 58, or 59, or an amino acid sequence having that sequence; or (c) a VH domain as in (a) and a VL domain as in (b). For example, in some cases, the antibody contains a VH domain containing the amino acid sequence of SEQ ID NO: 36 and a VL domain containing the amino acid sequence of SEQ ID NO: 37. In some cases, the antibody contains a VH domain containing the amino acid sequence of SEQ ID NO: 47 and a VL domain containing the amino acid sequence of SEQ ID NO: 37. In some cases, the antibody contains a VH domain containing the amino acid sequence of SEQ ID NO: 48 and a VL domain containing the amino acid sequence of SEQ ID NO: 37. In some cases, the antibody contains a VH domain containing the amino acid sequence of SEQ ID NO: 49 and a VL domain containing the amino acid sequence of SEQ ID NO: 37. In some cases, the antibody contains a VH domain containing the amino acid sequence of SEQ ID NO: 50 and a VL domain containing the amino acid sequence of SEQ ID NO: 37. In some cases, the antibody contains a VH domain containing the amino acid sequence of SEQ ID NO: 50 and a VL domain containing the amino acid sequence of SEQ ID NO: 37. In some cases, the antibody contains a VH domain containing the amino acid sequence of SEQ ID NO: 51 and a VL domain containing the amino acid sequence of SEQ ID NO: 37. In some cases, the antibody contains a VH domain containing the amino acid sequence of SEQ ID NO: 52 and a VL domain containing the amino acid sequence of SEQ ID NO: 37. In some examples, the antibody contains a VH domain with the amino acid sequence of SEQ ID NO: 36 and a VL domain with the amino acid sequence of SEQ ID NO: 53.In some examples, the antibody contains a VH domain containing the amino acid sequence of SEQ ID NO: 47 and a VL domain containing the amino acid sequence of SEQ ID NO: 53. In some examples, the antibody contains a VH domain containing the amino acid sequence of SEQ ID NO: 48 and a VL domain containing the amino acid sequence of SEQ ID NO: 53. In some examples, the antibody contains a VH domain containing the amino acid sequence of SEQ ID NO: 49 and a VL domain containing the amino acid sequence of SEQ ID NO: 53. In some examples, the antibody contains a VH domain containing the amino acid sequence of SEQ ID NO: 50 and a VL domain containing the amino acid sequence of SEQ ID NO: 53. In some examples, the antibody contains a VH domain containing the amino acid sequence of SEQ ID NO: 51 and a VL domain containing the amino acid sequence of SEQ ID NO: 53. In some examples, the antibody contains a VH domain containing the amino acid sequence of SEQ ID NO: 52 and a VL domain containing the amino acid sequence of SEQ ID NO: 53. In some examples, the antibody contains a VH domain containing the amino acid sequence of SEQ ID NO: 36 and a VL domain containing the amino acid sequence of SEQ ID NO: 58. In some examples, the antibody contains a VH domain containing the amino acid sequence of SEQ ID NO: 47 and a VL domain containing the amino acid sequence of SEQ ID NO: 58. In some examples, the antibody contains a VH domain containing the amino acid sequence of SEQ ID NO: 48 and a VL domain containing the amino acid sequence of SEQ ID NO: 58. In some examples, the antibody contains a VH domain containing the amino acid sequence of SEQ ID NO: 49 and a VL domain containing the amino acid sequence of SEQ ID NO: 58. In some examples, the antibody contains a VH domain containing the amino acid sequence of SEQ ID NO: 50 and a VL domain containing the amino acid sequence of SEQ ID NO: 58. In some examples, the antibody contains a VH domain containing the amino acid sequence of SEQ ID NO: 51 and a VL domain containing the amino acid sequence of SEQ ID NO: 58. In some examples, the antibody contains a VH domain containing the amino acid sequence of SEQ ID NO: 36 and a VL domain containing the amino acid sequence of SEQ ID NO: 59. In some examples, the antibody contains a VH domain containing the amino acid sequence of SEQ ID NO: 47 and a VL domain containing the amino acid sequence of SEQ ID NO: 59. In some cases, the antibody contains a VH domain containing the amino acid sequence of SEQ ID NO: 48 and a VL domain containing the amino acid sequence of SEQ ID NO: 59.In some examples, the antibody contains a VH domain containing the amino acid sequence of SEQ ID NO: 49 and a VL domain containing the amino acid sequence of SEQ ID NO: 59. In some examples, the antibody contains a VH domain containing the amino acid sequence of SEQ ID NO: 50 and a VL domain containing the amino acid sequence of SEQ ID NO: 59. In some examples, the antibody contains a VH domain containing the amino acid sequence of SEQ ID NO: 51 and a VL domain containing the amino acid sequence of SEQ ID NO: 59.

[0193] In other specific embodiments, in some embodiments, the anti-tryptase antibody may include (a) HVR-H1 containing the amino acid sequence of GYAIT (SEQ ID NO: 30), (b) HVR-H2 containing the amino acid sequence of GISSAATTFYSSWAKS (SEQ ID NO: 31), (c) HVR-H3 containing the amino acid sequence of DPRGYGAALDRLDL (SEQ ID NO: 32), (d) HVR-L1 containing the amino acid sequence of QSIKSVYNNRLG (SEQ ID NO: 33), (e) HVR-L2 containing the amino acid sequence of ETSILTS (SEQ ID NO: 34), and (f) HVR-L3 containing the amino acid sequence of AGGFDRSGDTT (SEQ ID NO: 35). In some embodiments, the antitryptase antibody comprises (a) a heavy chain variable (VH) domain containing an amino acid sequence having at least 90% sequence identity with the amino acid sequence of SEQ ID NO: 36 (e.g., at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%), or (b) a light chain variable (VL) domain containing an amino acid sequence having at least 90% sequence identity with the amino acid sequence of SEQ ID NO: 37 (e.g., at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%), or (c) a VH domain as in (a) and a VL domain as in (b). In some embodiments, the anti-tryptase antibody comprises one, two, three, or four of the following heavy chain framework regions (FRs): (a) FR-H1 comprising the amino acid sequence of EVQLVESGPGLVKPSETLSLTCTVSRFSLI (SEQ ID NO: 38), (b) FR-H2 comprising the amino acid sequence of WIRQPPGKGLEWIG (SEQ ID NO: 42), (c) FR-H3 comprising the amino acid sequence of RVTISRDTSKNQVSLKLSSVTAADTAVYYCAR (SEQ ID NO: 43), and (d) FR-H4 comprising the amino acid sequence of WGQGTLVTVSS (SEQ ID NO: 41).In some embodiments, the anti-tryptase antibody comprises one, two, three, or four of the following light chains FR: (a) FR-L1 containing the amino acid sequence of DIQMTQSPSSLSASVGDRVTITC (SEQ ID NO: 64), (b) FR-L2 containing the amino acid sequence of WYQQKPGKAPKLLIY (SEQ ID NO: 65), (c) FR-L3 containing the amino acid sequence of GVPSRFSGSGSGTDFTLTISSLQPEDFATYYC (SEQ ID NO: 66), and (d) FR-L4 containing the amino acid sequence of FGQGTKVEIK (SEQ ID NO: 63). In some embodiments, the anti-tryptase antibody comprises a VH domain containing the amino acid sequence of SEQ ID NO: 36, such as anti-tryptase antibody huE104.v2, and a VL domain containing the amino acid sequence of SEQ ID NO: 37.

[0194] In other specific embodiments, in some embodiments, the antibody (e.g., anti-tryptase antibody) may include (a) HVR-H1 containing the amino acid sequence of GYAIT (SEQ ID NO: 30), (b) HVR-H2 containing the amino acid sequence of GISSAATTFYSSWAKS (SEQ ID NO: 31), (c) HVR-H3 containing the amino acid sequence of DPRGYGAALDRLDL (SEQ ID NO: 32), (d) HVR-L1 containing the amino acid sequence of QSIKSVYNNRLG (SEQ ID NO: 33), (e) HVR-L2 containing the amino acid sequence of ETSILTS (SEQ ID NO: 34), and (f) HVR-L3 containing the amino acid sequence of AGGFDRSGDTT (SEQ ID NO: 35). In some embodiments, the antibody (e.g., an antitryptase antibody) comprises (a) a heavy chain variable (VH) domain containing an amino acid sequence having at least 90% sequence identity with the amino acid sequence of SEQ ID NO: 52 (e.g., at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%), or (b) a light chain variable (VL) domain containing an amino acid sequence having at least 90% sequence identity with the amino acid sequence of SEQ ID NO: 53 (e.g., at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%), or (c) a VH domain as in (a) and a VL domain as in (b). In some embodiments, the antibody (e.g., an anti-tryptase antibody) comprises one, two, three, or four of the following heavy chain framework regions (FRs): (a) FR-H1 comprising the amino acid sequence of QXSLEESGGGLFKPTDTLTLTCTVSRFSLI (SEQ ID NO: 54), (b) FR-H2 comprising the amino acid sequence of WVRQSPENGLEWIG (SEQ ID NO: 55), (c) FR-H3 comprising the amino acid sequence of RSTITRNTNENTVTLKMTSLTAADTATYFCAR (SEQ ID NO: 56), and (d) FR-H4 comprising the amino acid sequence of WGQGTLVTVSS (SEQ ID NO: 57).In some embodiments, the antibody (e.g., an anti-tryptase antibody) comprises one, two, three, or four of the following light chains FR: (a) FR-L1 comprising the amino acid sequence AAVLTQTPASVSAAVGGTVSISC (SEQ ID NO: 67), (b) FR-L2 comprising the amino acid sequence WYQQKPGQPPKLLIY (SEQ ID NO: 68), (c) FR-L3 comprising the amino acid sequence GVPSRFKGSGSETQFTLTISDVQX1DDAATYFC (SEQ ID NO: 69) where X1 is Cys or Ala, and (d) FR-L4 comprising the amino acid sequence FGQGTKVEIKFGGGTEVVVK (SEQ ID NO: 70). In some embodiments, the antibody (e.g., an anti-tryptase antibody) comprises a VH domain comprising the amino acid sequence of SEQ ID NO: 52 and a VL domain comprising the amino acid sequence of SEQ ID NO: 53, such as antibody E104. In some embodiments, the antibody (e.g., an anti-tryptase antibody) includes a VH domain containing the amino acid sequence of SEQ ID NO: 52 and a VL domain containing the amino acid sequence of SEQ ID NO: 59.

[0195] In some examples, the present invention provides an antibody comprising (a) a heavy chain comprising an amino acid sequence having at least 90% sequence identity with the amino acid sequence of SEQ ID NO: 76 (e.g., at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%), or having the same sequence, and / or (b) a light chain comprising an amino acid sequence having at least 90% sequence identity with the amino acid sequence of SEQ ID NO: 77 (e.g., at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%), or having the same sequence. In some examples, the antibody comprises a heavy chain comprising the amino acid sequence of SEQ ID NO: 76 and a light chain comprising the amino acid sequence of SEQ ID NO: 77. In some embodiments, the heavy chain further comprises a lysine (K) residue at the C-terminus.

[0196] In some examples, the present invention provides an antibody comprising (a) a heavy chain comprising an amino acid sequence having at least 90% sequence identity with the amino acid sequence of SEQ ID NO: 78 (e.g., at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%), or having the same sequence, and / or (b) a light chain comprising an amino acid sequence having at least 90% sequence identity with the amino acid sequence of SEQ ID NO: 79 (e.g., at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%), or having the same sequence. In some examples, the antibody comprises a heavy chain comprising the amino acid sequence of SEQ ID NO: 78 and a light chain comprising the amino acid sequence of SEQ ID NO: 79. In some embodiments, the heavy chain further comprises a lysine (K) residue at the C-terminus.

[0197] In some examples, the present invention provides an antibody comprising (a) a heavy chain comprising an amino acid sequence having at least 90% sequence identity with the amino acid sequence of SEQ ID NO: 80 (e.g., at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%), or having the same sequence, and / or (b) a light chain comprising an amino acid sequence having at least 90% sequence identity with the amino acid sequence of SEQ ID NO: 81 (e.g., at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%), or having the same sequence. In some examples, the antibody comprises a heavy chain comprising the amino acid sequence of SEQ ID NO: 80 and a light chain comprising the amino acid sequence of SEQ ID NO: 81. In some embodiments, the heavy chain further comprises a lysine (K) residue at the C-terminus.

[0198] In some examples, the present invention provides an antibody comprising (a) a heavy chain comprising an amino acid sequence having at least 90% sequence identity with the amino acid sequence of SEQ ID NO: 82 (e.g., at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%), or having the same sequence, and / or (b) a light chain comprising an amino acid sequence having at least 90% sequence identity with the amino acid sequence of SEQ ID NO: 83 (e.g., at least 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%), or having the same sequence. In some examples, the antibody comprises a heavy chain comprising the amino acid sequence of SEQ ID NO: 82 and a light chain comprising the amino acid sequence of SEQ ID NO: 83. In some embodiments, the heavy chain further comprises a lysine (K) residue at the C-terminus.

[0199] In some examples, one of the aforementioned antibodies binds to an epitope on human tryptase beta 1 containing at least one, at least two, or all three residues selected from the group consisting of Gln100, Leu101, and Leu102 of SEQ ID NO: 71. In some embodiments, the epitope on human tryptase beta 1 further contains one or more amino acid residues selected from the group consisting of Trp55, Gln67, Asp82, Leu83, Ala84, Arg87, Pro103, Val104, Ser105, Arg106, Glu126, Leu127, Glu128, and Glu129 of SEQ ID NO: 71. In some embodiments, the epitope on human tryptase beta 1 includes at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, at least ten, at least eleven, at least twelve, at least thirteen, or all fourteen amino acid residues selected from the group consisting of Trp55, Gln67, Asp82, Leu83, Ala84, Arg87, Pro103, Val104, Ser105, Arg106, Glu126, Leu127, Glu128, and Glu129 of SEQ ID NO: 71. In some embodiments, the epitopes include Gln35, Trp55, Gln67, Asp82, Leu83, Ala84, Arg87, Gln100, Leu101, Leu102, Pro103, Val104, Ser105, Arg106, Glu126, Leu127, Glu128, Glu129, and Arg216 of SEQ ID NO: 71. In some embodiments, the epitopes are associated with the human tryptase beta-1 monomer or tetramer. In some embodiments, the epitopes are associated with the human tryptase beta-1 tetramer, and the epitopes on human tryptase beta-1 further include one or both of Gln35 and Arg216 of SEQ ID NO: 71. In some embodiments, the epitopes are determined by an X-ray crystallography model. In some embodiments, the antibody can dissociate the small and / or large interfaces of human tryptase beta 1.

[0200] In some examples, any one of the aforementioned anti - tryptase antibodies comprises a paratope that binds to tryptase (e.g., human tryptase beta 1) and includes one or more amino acid residues selected from the group consisting of light chain variable region amino acid residues Tyr29, Asn30, Arg32, and Arg94, and heavy chain variable region amino acid residues Gly31, Tyr32, Ser52, Ser53, Ala54, Thr56, Phe58, Pro96, Arg97, Gly98, Tyr99, and Arg100e (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, or 16, 19 amino acid residues).

[0201] For example, in some examples, the anti - tryptase antibody comprises a paratope that binds to tryptase (e.g., human tryptase beta 1) and includes light chain variable region amino acid residues Tyr29, Asn30, Arg32, and Arg94 or heavy chain variable region amino acid residues Gly31, Tyr32, Ser52, Ser53, Ala54, Thr56, Phe58, Pro96, Arg97, Gly98, Tyr99, and Arg100e. In some examples, the anti - tryptase antibody comprises a paratope that binds to tryptase (e.g., human tryptase beta 1) and includes light chain variable region amino acid residues Tyr29, Asn30, Arg32, and Arg94, and heavy chain variable region amino acid residues Gly31, Tyr32, Ser52, Ser53, Ala54, Thr56, Phe58, Pro96, Arg97, Gly98, Tyr99, and Arg100e.

[0202] In some examples, any one of the aforementioned anti - tryptase antibodies binds to human tryptase. In some examples, any one of the aforementioned antibodies binds to cynomolgus (cyno) tryptase. In some examples, the antibody binds to human tryptase alpha or human tryptase beta. In some examples, the antibody binds to human tryptase beta 1, human tryptase beta 2, or human tryptase beta 3.

[0203] In another embodiment, the present invention provides an anti-tryptase antibody that binds to an epitope on a tryptase (e.g., human tryptase beta 1) containing one or more amino acid residues selected from the group consisting of His51, Val80, Lys81, Asp82, Leu83, Ala84, and Ala85, which can refer to the amino acid sequence of SEQ ID NO: 71 (e.g., one, two, three, four, five, six, or seven amino acid residues), or the corresponding amino acid of any tryptase protein. For example, in some embodiments, the antibody binds to an epitope on a tryptase (e.g., human tryptase beta 1) containing at least one, at least two, at least three, or at least four residues selected from the group consisting of His51, Val80, Lys81, and Asp82 of SEQ ID NO: 71, or the corresponding amino acid of any tryptase protein. In some embodiments, the antibody binds to an epitope on a tryptase (e.g., human tryptase beta-1) that contains His51 of SEQ ID NO: 71 and at least one, at least two, or all three residues selected from the group consisting of Val80, Lys81, and Asp82, or the corresponding amino acids of any tryptase protein. In some embodiments, the epitope on the tryptase (e.g., human tryptase beta-1) further comprises one or more amino acid residues selected from the group consisting of Gln67, Leu83, Ala84, Ala85, Arg87, Pro103, Val104, Ser105, Arg106, Glu128, Glu129, and Pro130 of SEQ ID NO: 71, or the corresponding amino acids of any tryptase protein.In some embodiments, the epitope on tryptase (e.g., human tryptase beta 1) comprises at least 2, at least 3, at least 4, at least 5, at least 6, at least 7, at least 8, at least 9, at least 10, at least 11, or all 12 amino acid residues selected from the group consisting of Gln67, Leu83, Ala84, Ala85, Arg87, Pro103, Val104, Ser105, Arg106, Glu128, Glu129, and Pro130 of SEQ ID NO: 71, or the corresponding amino acids of any tryptase protein. In some embodiments, the epitope on tryptase (e.g., human tryptase beta 1) comprises His51, Gln67, Val80, Lys81, Asp82, Leu83, Ala84, Ala85, Arg87, Pro103, Val104, Ser105, Arg106, Glu128, Glu129, and Pro130 of SEQ ID NO: 71, or the corresponding amino acids of any tryptase protein. In some embodiments, the epitope is associated with the tryptase (e.g., human tryptase beta 1) monomer or tetramer. In some embodiments, the epitope is determined by an X-ray crystallography model. In some embodiments, the antibody is capable of dissociating both the small interface of the tetrameric tryptase and the large interface of the tetrameric tryptase tryptase (e.g., human tryptase beta 1).

[0204] In yet another aspect, the present invention provides an anti-tryptase antibody that binds to an epitope of a tryptase (e.g., human tryptase beta-1) containing one or more amino acid residues selected from the group consisting of Gln100, Leu101, Leu102, Pro103, Val104, Ser105, and Arg106 (e.g., one, two, three, four, five, six, or seven amino acid residues), or the corresponding amino acid of any tryptase protein, which can refer to the amino acid sequence of SEQ ID NO: 71. In some embodiments, the epitope on the tryptase (e.g., human tryptase beta-1) further comprises one or more amino acid residues selected from the group consisting of Trp55, Gln67, Asp82, Leu83, Ala84, Arg87, Pro103, Val104, Ser105, Arg106, Glu126, Leu127, Glu128, and Glu129 of SEQ ID NO: 71, or the corresponding amino acids of any tryptase protein. In some embodiments, the epitope on tryptase (e.g., human tryptase beta-1) includes at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, at least ten, at least eleven, at least twelve, at least thirteen, or fourteen amino acid residues selected from the group consisting of Trp55, Gln67, Asp82, Leu83, Ala84, Arg87, Pro103, Val104, Ser105, Arg106, Glu126, Leu127, Glu128, and Glu129 of SEQ ID NO: 71, or the corresponding amino acids of any tryptase protein. In some embodiments, the epitope comprises Gln35, Trp55, Gln67, Asp82, Leu83, Ala84, Arg87, Gln100, Leu101, Leu102, Pro103, Val104, Ser105, Arg106, Glu126, Leu127, Glu128, Glu129, and Arg216 of SEQ ID NO: 71, or the corresponding amino acids of any tryptase protein. In some embodiments, the epitope is associated with a tryptase (e.g., human tryptase beta-1) monomer or tetramer.In some embodiments, the epitope is associated with a tetramer, and the epitope on the tryptase (e.g., human tryptase beta-1) contains one or both of Gln35 and Arg216 of SEQ ID NO: 71, or the corresponding amino acid of any tryptase protein. In some embodiments, the epitope is determined by an X-ray crystallography model. In some embodiments, the antibody can dissociate the small and / or large interfaces of the tryptase (e.g., human tryptase beta-1).

[0205] In some embodiments, any of the aforementioned antibodies bind to an epitope on a tryptase (e.g., human tryptase beta-1) containing one or more amino acid residues selected from the group consisting of Gln67, Asp82, Leu83, Ala84, Arg87, Pro103, Val104, Ser105, Arg106, Glu128, and Glu129 (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, or 11 amino acid residues) that can reference the amino acid sequence of SEQ ID NO: 71, or the corresponding amino acid of any tryptase protein.

[0206] For example, in some cases, one of the aforementioned antibodies binds to an epitope on a tryptase (e.g., human tryptase beta 1) containing the Gln67 amino acid of SEQ ID NO: 71 or the corresponding amino acid of any tryptase protein. In some cases, the antibody binds to an epitope on a tryptase (e.g., human tryptase beta 1) containing the Asp82 amino acid of any tryptase protein. In some cases, the antibody binds to an epitope on a tryptase (e.g., human tryptase beta 1) containing the Leu83 amino acid of any tryptase protein. In some cases, the antibody binds to an epitope on a tryptase (e.g., human tryptase beta 1) containing the Ala84 amino acid of any tryptase protein. In some cases, the antibody binds to an epitope on a tryptase (e.g., human tryptase beta 1) containing the Arg87 amino acid of any tryptase protein. In some cases, the antibody binds to an epitope on a tryptase (e.g., human tryptase beta 1) containing Pro103 of SEQ ID NO: 71 or the corresponding amino acid of any tryptase protein. In some cases, the antibody binds to an epitope on a tryptase (e.g., human tryptase beta 1) containing Val104 of SEQ ID NO: 71 or the corresponding amino acid of any tryptase protein. In some cases, the antibody binds to an epitope on a tryptase (e.g., human tryptase beta 1) containing Ser105 of SEQ ID NO: 71 or the corresponding amino acid of any tryptase protein. In some cases, the antibody binds to an epitope on a tryptase (e.g., human tryptase beta 1) containing Arg106 of SEQ ID NO: 71 or the corresponding amino acid of any tryptase protein. In some cases, the antibody binds to an epitope on a tryptase (e.g., human tryptase beta 1) containing Glu128 of SEQ ID NO: 71 or the corresponding amino acid of any tryptase protein.In some cases, the antibody binds to an epitope on a tryptase (e.g., human tryptase beta-1) that contains the corresponding amino acid of Glu129 in SEQ ID NO: 71 or any other tryptase protein.

[0207] In some embodiments, any of the aforementioned antibodies bind to an epitope on a tryptase (e.g., human tryptase beta-1) containing two or more, three or more, four or more, five or more, six or more, seven or more, eight or more, nine or more, ten or more, or eleven amino acid residues selected from the group consisting of Gln67, Asp82, Leu83, Ala84, Arg87, Pro103, Val104, Ser105, Arg106, Glu128, and Glu129, which may refer to the amino acid sequence of SEQ ID NO: 71, or the corresponding amino acids of any tryptase protein.

[0208] In some cases, one of the aforementioned antibodies binds to an epitope on a tryptase (e.g., human tryptase beta 1) that further contains one or more additional amino acid residues (e.g., one, two, three, four, or five additional amino acid residues) selected from the group consisting of His51, Val80, Lys81, Ala85, and Pro130, which may refer to the amino acid sequence of SEQ ID NO: 71, or the corresponding amino acid of any tryptase protein. For example, in some cases, the antibody binds to an epitope on a tryptase (e.g., human tryptase beta 1) that further contains His51 of SEQ ID NO: 71 or the corresponding amino acid of any tryptase protein. In some cases, the antibody binds to an epitope on a tryptase (e.g., human tryptase beta 1) that further contains Val80 of SEQ ID NO: 71 or the corresponding amino acid of any tryptase protein. In some embodiments, the anti-tryptase antibody binds to an epitope on a tryptase (e.g., human tryptase beta-1) further containing the corresponding amino acid of Lys81 of SEQ ID NO: 71 or any tryptase protein. In some examples, the antibody binds to an epitope on a tryptase (e.g., human tryptase beta-1) further containing the corresponding amino acid of Ala85 of SEQ ID NO: 71 or any tryptase protein. In some examples, the antibody binds to an epitope on a tryptase (e.g., human tryptase beta-1) further containing the corresponding amino acid of Pro130 of SEQ ID NO: 71 or any tryptase protein.

[0209] In some cases, the antibody binds to an epitope on a tryptase (e.g., human tryptase beta-1) that contains two or more, three or more, four or more, or five or more additional amino acid residues selected from the group consisting of His51, Val80, Lys81, Ala85, and Pro130, which may refer to the amino acid sequence of SEQ ID NO: 71, or the corresponding amino acids of any tryptase protein.

[0210] In some cases, anti-tryptase antibodies bind to epitopes on tryptase (e.g., human tryptase beta-1) that contain His51, Gln67, Val80, Lys81, Asp82, Leu83, Ala84, Ala85, Arg87, Pro103, Val104, Ser105, Arg106, Glu128, Glu129, and Pro130 of SEQ ID NO: 71, or the corresponding amino acids of any tryptase protein, which may refer to the amino acid sequence of SEQ ID NO: 71. In some cases, anti-tryptase antibodies bind to epitopes on tryptase (e.g., human tryptase beta-1) consisting of His51, Gln67, Val80, Lys81, Asp82, Leu83, Ala84, Ala85, Arg87, Pro103, Val104, Ser105, Arg106, Glu128, Glu129, and Pro130, as shown in SEQ ID NO: 71.

[0211] In other examples, any of the aforementioned anti-tryptase antibodies bind to an epitope on a tryptase (e.g., human tryptase beta 1) that further contains one or more (e.g., one, two, three, four, five, six, seven, or eight) amino acid residues selected from the group consisting of Gln35, Trp55, Gln100, Leu101, Leu102, Glu126, Leu127, and Arg216, which may refer to the amino acid sequence of SEQ ID NO: 71, or to an epitope on a tryptase (e.g., human tryptase beta 1) that further contains the corresponding amino acid of any tryptase protein. For example, in some examples, the antibody binds to an epitope on a tryptase (e.g., human tryptase beta 1) that further contains Gln35 of SEQ ID NO: 71 or the corresponding amino acid of any tryptase protein. In some examples, the antibody binds to an epitope on a tryptase (e.g., human tryptase beta 1) that further contains Trp55 of SEQ ID NO: 71 or the corresponding amino acid of any tryptase protein. In some cases, the antibody binds to an epitope on a tryptase (e.g., human tryptase beta 1) further containing the corresponding amino acid of Glu100 in SEQ ID NO: 71 or any tryptase protein. In some cases, the antibody binds to an epitope on a tryptase (e.g., human tryptase beta 1) further containing the corresponding amino acid of Leu101 in SEQ ID NO: 71 or any tryptase protein. In some cases, the antibody binds to an epitope on a tryptase (e.g., human tryptase beta 1) further containing the corresponding amino acid of Leu102 in SEQ ID NO: 71 or any tryptase protein. In some cases, the antibody binds to an epitope on a tryptase (e.g., human tryptase beta 1) further containing the corresponding amino acid of Glu126 in SEQ ID NO: 71 or any tryptase protein. In some cases, the antibody binds to an epitope on a tryptase (e.g., human tryptase beta 1) further containing the corresponding amino acid of Leu127 in SEQ ID NO: 71 or any tryptase protein. In some cases, the antibody binds to an epitope on a tryptase (e.g., human tryptase beta-1) that further contains Arg216 of SEQ ID NO: 71 or the corresponding amino acid of any tryptase protein.In some cases, the antibody binds to an epitope on a tryptase (e.g., human tryptase beta-1) that contains two or more, three or more, four or more, five or more, six or more, seven or more, or eight or more additional amino acid residues selected from the group consisting of Gln35, Trp55, Gln100, Leu101, Leu102, Glu126, Leu127, and Arg216, which may refer to the amino acid sequence of SEQ ID NO: 71, or the corresponding amino acids of any tryptase protein.

[0212] In some cases, anti-tryptase antibodies bind to epitopes on tryptase (e.g., human tryptase beta-1) that contain the amino acids Gln35, Trp55, Gln67, Asp82, Leu83, Ala84, Arg87, Gln100, Leu101, Leu102, Pro103, Val104, Ser105, Arg106, Glu126, Leu127, Glu128, Glu129, and Arg216, or the corresponding amino acids of any tryptase protein, which may refer to the amino acid sequence of Sequence ID No. 71. In some cases, anti-tryptase antibodies bind to epitopes on tryptase (e.g., human tryptase beta-1) consisting of Gln35, Trp55, Gln67, Asp82, Leu83, Ala84, Arg87, Gln100, Leu101, Leu102, Pro103, Val104, Ser105, Arg106, Glu126, Leu127, Glu128, Glu129, and Arg216 as of SEQ ID NO: 71.

[0213] In another embodiment, the present invention provides an antibody that competes with any of the aforementioned antibodies for binding to a tryptase (e.g., human tryptase beta-1).

[0214] In another embodiment, the present invention provides an antibody that binds to the same epitope or duplicate epitope as any of the aforementioned antibodies.

[0215] In some embodiments, any one of the aforementioned antibodies may have the ability to cleave tryptase having a tetrameric structure (such as mature tryptase present in or released from mast cell secretory granules) to form small molecular weight species, such as monomers, dimers, and / or trimers.

[0216] In a further aspect, an antibody according to any of the above embodiments may incorporate any one or combination of the features described in Sections 1 to 7 below.

[0217] 1. Antibody affinity In certain embodiments, the antibodies provided herein have a dissociation constant (K -6 ) of ≦1 μM, ≦100 nM, ≦10 nM, ≦1 nM, ≦0.1 nM, ≦0.01 nM, ≦1 pM, or ≦0.1 pM (e.g., 10 -6 M or less, e.g., 10 -9 M to 10 -9 M or less, e.g., 10 -13 M to 10 D ). In some embodiments, the anti-tryptase antibodies of the present invention bind to tryptase (e.g., human tryptase, e.g., human tryptase beta) with a K D of about 100 nM or less (e.g., 100 nM or less, 10 nM or less, 1 nM or less, 100 pM or less, 10 pM or less, 1 pM or less, or 0.1 pM or less). In some embodiments, the antibody binds to tryptase (e.g., human tryptase, e.g., human tryptase beta) with a K D of 10 nM or less (e.g., 10 nM or less, 1 nM or less, 100 pM or less, 10 pM or less, 1 pM or less, or 0.1 pM or less). In some embodiments, the antibody binds to tryptase (e.g., human tryptase, e.g., human tryptase beta) with a K DIt binds to trypsin (e.g., human trypsin, e.g., human trypsin beta). In some embodiments, any of the anti-trypsin antibodies described above or herein has a K of about 0.5 nM or less (e.g., 0.5 nM or less, 400 pM or less, 300 pM or less, 200 pM or less, 100 pM or less, 50 pM or less, 25 pM or less, 10 pM or less, 1 pM or less, or 0.1 pM or less). D It binds to trypsin (e.g., human trypsin, e.g., human trypsin beta). In some embodiments, the antibody has a K of from about 0.1 nM to about 0.5 nM (e.g., about 0.1 nM, about 0.2 nM, about 0.3 nM, about 0.4 nM, or about 0.5 nM). D It binds to trypsin (e.g., human trypsin, e.g., human trypsin beta). In some embodiments, the antibody has a K of about 0.4 nM. D It binds to trypsin (e.g., human trypsin, e.g., human trypsin beta). In some embodiments, the antibody has a K of about 0.18 nM. D It binds to trypsin (e.g., human trypsin, e.g., human trypsin beta).

[0218] In one embodiment, K D is measured by a radioimmunoassay (RIA). In one embodiment, the RIA is performed using the Fab version of the antibody of interest and its antigen. For example, the solution binding affinity of the Fab for the antibody is determined in the presence of a series of titrations of unlabeled antigen with the lowest concentration of Fab( 125I) The antibody is measured by equilibrating it with a labeled antigen and then capturing the bound antibody with a plate coated with anti-Fab antibody (see, for example, Chen et al. J. Mol. Biol. 293:865-881, 1999). To establish conditions for the assay, MICROTITER® multiwell plates (Thermo Scientific) are coated overnight with 5 μg / ml of capture anti-Fab antibody (Cappel Labs) in 50 mM carbonate (pH 9.6), followed by blocking with 2% (w / v) bovine serum albumin in PBS for 2–5 hours at room temperature (approximately 23°C). In a non-adsorbent plate (Nunc No. 269620), 100 pM or 26 pM [ 125 Mix the [I]-antigen with serial dilutions of the target Fab (e.g., consistent with the assessment of anti-VEGF antibody, Fab-12, Presta et al. Cancer Res. 57:4593-4599, 1997). Incubate the target Fab overnight, although this incubation may be extended for a longer period (e.g., about 65 hours) to ensure equilibrium is achieved. Transfer the mixture to a capture plate for incubation at room temperature (e.g., 1 hour). Remove this solution and wash the plate eight times with 0.1% polysorbate 20 (TWEEN-20®) in PBS. Once the plate is dry, add 150 μL / well of scintillant (MICROSCINT-20®, Packard) and count the plate for 10 minutes using a TOPCOUNT® gamma counter (Packard). Select the concentration of each Fab that confers a maximum binding of 20% or less for use in competitive binding assays.

[0219] According to another embodiment, K DThis is measured using the BIACORE® surface plasmon resonance assay. For example, assays using BIACORE®-2000 or BIACORE®-3000 (BIAcore, Inc., Piscataway, NJ) are performed at 25°C using an immobilized antigen CM5 chip at approximately 10 response units (RUs). In one embodiment, a carboxymethylated dextran biosensor chip (CM5, BIACORE, Inc.) is activated with N-ethyl-N'-(3-dimethylaminopropyl)-carbodimide hydrochloride (EDC) and N-hydroxysuccinimide (NHS) according to the supplier's instructions. The antigen is diluted to 5 μg / ml (approximately 0.2 μM) with 10 mM sodium acetate at pH 4.8 and injected at a flow rate of 5 μl / min to achieve approximately 10 response units (RUs) of the coupled protein. Following the injection of the antigen, 1 M ethanolamine is injected to block unreacted groups. For kinetic measurements, serially diluted 2-fold dilutions of Fab (0.78 nM to 500 nM) are injected into phosphate-buffered saline (PBS) containing 0.05% polysorbate 20 (TWEEN® 20) surfactant (PBST) at 25°C at a flow rate of approximately 25 μL / min. The association rate (k on ) and dissociation rate (k off The equilibrium dissociation constant (K) is calculated by simultaneously fitting the association sensorgram and dissociation sensorgram using a simple one-to-one Langmuir couple model (BIACORE® evaluation software version 3.2). D ) is the ratio k off / k on Calculate as follows. For example, see Chen et al. (J Mol. Biol. 293:865-881, 1999). The ON velocity is 10 by the surface plasmon resonance assay described above. 6 M -1 s -1If it exceeds this value, the ON rate can be determined by using a fluorescence quenching technique, which measures the increase or decrease in fluorescence emission intensity (excitation = 295 nm, emission = 340 nm, 16 nm band-passing) of a 20 nM anti-antigen antibody (Fab type) in PBS (pH 7.2) at 25°C in the presence of gradually increasing concentrations of antigen, measured with a spectrometer such as an Aviv Instruments stop-flow spectrophotometer equipped with a stirred cuvette or an 8000 series SLM-AMINCO™ spectrophotometer (ThermoSpectronic).

[0220] In some embodiments, K D This is measured using the BIACORE® SPR assay, for example, as described in Section (A)(vii) of Example 1. In some embodiments, the SPR assay can be performed using a BIAcore® T200 or equivalent instrument. In some embodiments, a BIAcore® Series S CM5 sensor chip (or equivalent sensor chip) is immobilized with monoclonal mouse anti-human IgG (Fc) antibody, and the anti-tryptase antibody is sequentially captured on a flow cell. A series of 3-fold dilutions of His-tagged human tryptase beta-1 monomer (SEQ ID NO: 128) are injected at a flow rate of 30 μl / min. Each sample is analyzed by 3-minute association and 10-minute dissociation. The assay is performed at 25°C. After each injection, the chip is regenerated using 3M MgCl2. The binding response is corrected by subtracting the response unit (RU) from a flow cell capturing unrelated IgG of similar density. on and k off A 1:1 Languir model with simultaneous fitting is used for dynamic analysis.

[0221] 2. Antibody fragment In certain embodiments, the antibodies provided herein are antibody fragments. Antibody fragments include, but are not limited to, Fab, Fab’, Fab’-SH, F(ab’)2, Fv, and scFv fragments, and other fragments described below. For a review of certain antibody fragments, see Hudson et al. Nat. Med. 9:129-134 (2003). For a review of scFv fragments, see, e.g., Pluckthun, in The Pharmacology of Monoclonal Antibodies, vol. 113, Rosenberg and Moore eds., (Springer-Verlag, New York), pp. 269-315 (1994), as well as WO93 / 16185, and U.S. Patent Nos. 5,571,894 and 5,587,458. For a discussion of Fab and F(ab’)2 fragments that include salvage receptor binding epitope residues and have increased in vivo half-lives, see U.S. Patent No. 5,869,046.

[0222] A diabody is an antibody fragment having two antigen-binding sites that can be bivalent or bispecific. See, e.g., EP404,097, WO1993 / 01161, Hudson et al. Nat. Med. 9:129-134, 2003, and Hollinger et al. Proc. Natl. Acad. Sci. USA 99:6444-6448, 1993. Triabodies and tetrabodies are also described in Hudson et al., Nat. Med. 9:129-134, 2003.

[0223] Single domain antibodies are antibody fragments that include all or part of the heavy chain variable domain of an antibody, or all or part of the light chain variable domain. In certain embodiments, the single domain antibodies are human single domain antibodies (see, e.g., U.S. Patent No. 6,248,516 B1).

[0224] Antibody fragments can be prepared by a variety of techniques, including, but not limited to, the proteolysis of intact antibodies and the production of recombinant host cells (e.g., E. coli or phages) as described herein.

[0225] 3. Chimeric antibodies and humanized antibodies In certain embodiments, the antibodies provided herein are chimeric antibodies. Certain chimeric antibodies are described, for example, in U.S. Patent No. 4,816,567 and Morrison et al. Proc. Natl. Acad. Sci. USA, 81:6851-6855, 1984. In one example, a chimeric antibody includes a non-human variable region (e.g., a variable region derived from a mouse, rat, hamster, rabbit, or non-human primate, e.g., a monkey) and a human constant region. In further examples, a chimeric antibody is a “class-switched” antibody in which the class or subclass has changed from the class or subclass of the parent antibody. A chimeric antibody includes its antigen-binding fragment.

[0226] In certain embodiments, the chimeric antibody is a humanized antibody. Typically, a non-human antibody is humanized to reduce its immunogenicity to humans while retaining the specificity and affinity of the non-human parent antibody. Generally, a humanized antibody contains one or more variable domains in which the HVR (or portions thereof) are derived from the non-human antibody and the FR (or portions thereof) are derived from the human antibody sequence. The humanized antibody will optionally also contain at least a portion of the human constant region. In some embodiments, some FR residues in the humanized antibody are replaced with corresponding residues derived from the non-human antibody (e.g., the antibody from which the HVR residues originated) to restore or improve antibody specificity or affinity, for example.

[0227] Humanized antibodies and methods for producing them are outlined, for example, in Almagro et al. Front. Biosci. 13:1619-1633, 2008, and also in, for example, Riechmann et al. Nature 332:323-329, 1988, Queen et al. Proc. Natl. Acad. Sci. USA 86:10029-10033, 1989, U.S. Patents No. 5,821,337, No. 7,527,791, No. 6,982,321, and No. 7,087,409, and Kashmiri et al. Methods. Further details can be found in 36:25-34, 2005 (explaining specificity-determining region (SDR) transplantation), Padlan, Mol.Immunol.28:489-498, 1991 (explaining "resurfacing"), Dall'Acqua et al. Methods 36:43-60, 2005 (explaining "FR shuffling"), and Osbourn et al. Methods 36:61-68, 2005 and Klimka et al. Br.J.Cancer, 83:252-260, 2000 (explaining the "inducible selection" approach to FR shuffling).

[0228] Human framework regions that can be used for humanization include framework regions selected using the "best fit" method (see, e.g., Sims et al. J.Immunol. 151:2296, 1993), framework regions derived from consensus sequences of human antibodies of specific subgroups of light chain or heavy chain variable regions (see, e.g., Carter et al. Proc.Natl.Acad.Sci.USA, 89:4285, 1992, and Presta et al. J.Immunol., 151:2623, 1993); human mature (somatically mutant) framework regions or human germline framework regions (see, e.g., Almagro et al. Front.Biosci. 13:1619-1633, 2008), as well as framework regions derived from screening of FR libraries (see, e.g., Baca et al. J.Biol.Chem. 272:10678-10684, 1997 and Rosok et al. (See al.J.Biol.Chem.271:22611-22618,1996) and other examples, but not limited to these.

[0229] 4. Human antibodies In certain embodiments, the antibodies provided herein are human antibodies. Human antibodies can be produced using various techniques known in the art. Human antibodies are generally described in van Dijk et al. Curr. Opin. Pharmacol. 5:368-74, 2001 and Lonberg, Curr. Opin. Immunol. 20:450-459, 2008.

[0230] Human antibodies can be prepared by administering an immunogen to transgenic animals modified to produce intact human antibodies or intact antibodies with human variable regions in response to antigen administration. Such animals typically contain all or part of human immunoglobulin loci that replace endogenous immunoglobulin loci or are located extrachromosomally or randomly incorporated into the animal's chromosomes. In such transgenic mice, endogenous immunoglobulin loci are generally inactivated. For an overview of methods for obtaining human antibodies from transgenic animals, see Lonberg, Nat. Biotech. 23:1117-1125, 2005. See, for example, U.S. Patent Nos. 6,075,181 and 6,150,584 (describes XENOMOUSE® technology), U.S. Patent No. 5,770,429 (describes HUMAB® technology), U.S. Patent No. 7,041,870 (describes KM MOUSE® technology), and U.S. Patent Publication No. 2007 / 0061900 (describes VELOCIMOUSE® technology). Human variable regions derived from intact antibodies produced by such animals can be further modified, for example, by combining them with different human constant regions.

[0231] Human antibodies can also be produced by hybridoma-based methods. Human myeloma and mouse-human heterozygous myeloma cell lines for the production of human monoclonal antibodies are described. (See, for example, Kozbor J. Immunol. 133:3001, 1984, Brodeur et al. Monoclonal Antibody Production Techniques and Applications, pp. 51-63 (Marcel Dekker, Inc., New York, 1987), and Boerner et al. J. Immunol. 147:86, 1991). Human antibodies produced via human B-cell hybridoma technology are also described in Li et al. Proc. Natl. Acad. Sci. USA, 103:3557-3562, 2006. Further methods include, for example, those described in U.S. Patent No. 7,189,826 (regarding the production of monoclonal human IgM antibodies from hybridoma cell lines) and Ni, Xiandai Mianyixue, 26(4):265-268 (2006) (describing human-human hybridomas). Human hybridoma technology (trioma technology) is also described in Vollmers et al. Histology and Histopathology 20(3):927-937, 2005 and Vollmers et al. Methods and Findings in Experimental and Clinical Pharmacology 27(3):185-91, 2005.

[0232] Human antibodies can also be generated by isolating Fv clone variable domain sequences selected from human-derived phage display libraries. Such variable domain sequences can then be combined with desired human constant domains. Techniques for selecting human antibodies from antibody libraries are described below.

[0233] 5. Antibodies derived from libraries The antibodies of the present invention can be isolated by screening a combinatorial library for antibodies having desired activity(s). For example, various methods are known in the art for generating phage display libraries and screening such libraries for antibodies possessing desired binding properties. Such methods are outlined, for example, in Hoogenboom et al. in Methods in Molecular Biology 178:1-37 (O'Brien et al., ed., Human Press, Totowa, NJ, 2001), and also in, for example, McCafferty et al. Nature 348:552-554, 1990, Clackson et al. Nature 352:624-628, 1991, Marks et al. J.Mol.Biol.222:581-597, 1992, Marks et al. in Methods in Molecular Biology 248:161-175 (Lo, ed., Human Press, Totowa, NJ, 2003), Sidhu et al. J.Mol.Biol.338(2):299-310, 2004, Lee et al. Further explanation can be found in al. J. Mol. Biol. 340(5):1073-1093, 2004, Fellouse, Proc. Natl. Acad. Sci. USA 101(34):12467-12472, 2004, and Lee et al. J. Immunol. Methods 284(1-2):119-132, 2004.

[0234] In certain phage presentation methods, the repertoire of VH and VL genes can be separately cloned by polymerase chain reaction (PCR), randomly recombined within a phage library, and subsequently screened for antigen-binding phages as described in Winter et al. Ann. Rev. Immunol., 12:433-455, 1994. The phages typically display antibody fragments as either single-stranded Fv (scFv) fragments or Fab fragments. Libraries derived from immunization sources provide high-affinity antibodies to immunogens without requiring hybridoma construction. Alternatively, as described in Griffiths et al. EMBOJ. 12:725-734, 1993, naive repertoires can be cloned (e.g., from humans) to provide a single source of antibodies against a wide range of non-self and autoantigens without immunization. Finally, as described in Hoogenboom et al. J. Mol. Biol., 227:381-388, 1992, naive libraries can also be synthetically constructed by cloning an unreorganized V gene segment from stem cells, encoding a highly variable HVR3 region using PCR primers containing random sequences, and achieving in vitro rearrangement. Patent publications describing human antibody phage libraries include, for example, U.S. Patent No. 5,750,373, and U.S. Patent Application Publications 2005 / 0079574, 2005 / 0119455, 2005 / 0266000, 2007 / 0117126, 2007 / 0160598, 2007 / 0237764, 2007 / 0292936, and 2009 / 0002360.

[0235] Antibodies or antibody fragments isolated from a human antibody library are considered human antibodies or human antibody fragments in this specification.

[0236] 6. Multispecific antibodies In certain embodiments, the antibodies provided herein are multispecific antibodies, for example, bispecific antibodies. A multispecific antibody is a monoclonal antibody having binding specificity to at least two different sites. In certain embodiments, a bispecific antibody may bind to two different epitopes of tryptase. In certain embodiments, one of the binding specificities is for tryptase and the other is for any other antigen (e.g., a second biological molecule). In some embodiments, a bispecific antibody can bind to two different epitopes of tryptase. In other embodiments, one of the binding specificities is for tryptase (e.g., human tryptase, e.g., human tryptase beta) and the other is for any other antigen (e.g., a second biological molecule, e.g., IL-13, IL-4, IL-5, IL-17, IL-33, IgE, M1 prime, CRTH2, or TRPA). Therefore, bispecific antibodies may have binding specificity to tryptase and IL-13, tryptase and IL-4, tryptase and IL-5, tryptase and IL-17, or tryptase and IL-33. In particular, bispecific antibodies may have binding specificity to tryptase and IL-13 or tryptase and IL-33. Bispecific antibodies can be prepared as full-length antibodies or antibody fragments.

[0237] For example, in some cases, a bispecific antibody includes a first binding domain that binds to tryptase and a second binding domain that binds to IL-13. In some embodiments, the first binding domain that binds to tryptase is, for example, (a) HVR-H1 containing the amino acid sequence X1X2GMX3 (SEQ ID NO: 1) where X1 is Asp or Ser, X2 is Tyr or Phe, and X3 is Val or His; (b) HVR-H2 containing the amino acid sequence FISSGSSTVYYADTMKG (SEQ ID NO: 2); (c) HVR-H3 containing the amino acid sequence RX1X2X3DWYFDV (SEQ ID NO: 3) where X1 is Asn or Asp, X2 is Tyr or Asn, and X3 is Asp or Tyr; and (d) HVR-L1 containing the amino acid sequence SASSSVTYMY (SEQ ID NO: 4). This may include, at least one, two, three, four, five, or six hypervariable regions (HVRs) selected from (e) HVR-L2 containing the amino acid sequence of RTSDLAS (SEQ ID NO: 5) and (f) HVR-L3 containing the amino acid sequence of QHYHSYPLT (SEQ ID NO: 6), or a combination of one or more of the above HVRs, and one or more variants thereof having at least about 80% sequence identity with any one of SEQ ID NOs. 1 to 6 (e.g., 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity).In some examples, the second binding domain that binds to IL-13 is, for example, (a) HVR-H1 containing the amino acid sequence of AYSVN (SEQ ID NO: 84), (b) HVR-H2 containing the amino acid sequence of MIWGDGKIVYNSALKS (SEQ ID NO: 85), (c) HVR-H3 containing the amino acid sequence of DGYYPYAMDN (SEQ ID NO: 86), (d) HVR-L1 containing the amino acid sequence of RASKSVDSYGNSFMH (SEQ ID NO: 87), (e) HVR-L2 containing the amino acid sequence of LASNLES (SEQ ID NO: 88), and (f) QQNNEDP The second binding domain may include at least one, two, three, four, five, or six HVRs selected from HVR-L3 containing the amino acid sequence of RT (SEQ ID NO: 89), or a combination of one or more of the above HVRs, and one or more variants thereof having at least about 80% sequence identity with any one of SEQ ID NOs. 84-89 (e.g., 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity). In some embodiments, the second binding domain includes one, two, three, four, five, or six HVRs of anti-IL-13 lebrikizumab.

[0238] For example, in some cases, the first binding domain that binds to tryptase includes at least one, two, three, four, five, or six hypervariable regions (HVRs) selected from (a) HVR-H1 containing the amino acid sequence of DYGMV (SEQ ID NO: 7), (b) HVR-H2 containing the amino acid sequence of FISSGSSTVYYADTMKG (SEQ ID NO: 2), (c) HVR-H3 containing the amino acid sequence of RNYDDWYFDV (SEQ ID NO: 8), (d) HVR-L1 containing the amino acid sequence of SASSSVTYMY (SEQ ID NO: 4), (e) HVR-L2 containing the amino acid sequence of RTSDLAS (SEQ ID NO: 5), and (f) HVR-L3 containing the amino acid sequence of QHYHSYPLT (SEQ ID NO: 6). In some examples, the second binding domain that binds to IL-13 may include at least one, two, three, four, five, or six HVRs selected from, for example, (a) HVR-H1 containing the amino acid sequence of AYSVN (SEQ ID NO: 84), (b) HVR-H2 containing the amino acid sequence of MIWGDGKIVYNSALKS (SEQ ID NO: 85), (c) HVR-H3 containing the amino acid sequence of DGYYPYAMDN (SEQ ID NO: 86), (d) HVR-L1 containing the amino acid sequence of RASKSVDSYGNSFMH (SEQ ID NO: 87), (e) HVR-L2 containing the amino acid sequence of LASNLES (SEQ ID NO: 88), and (f) HVR-L3 containing the amino acid sequence of QQNNEDPRT (SEQ ID NO: 89). In some embodiments, the second binding domain includes one, two, three, four, five, or six HVRs of anti-IL-13 lebrikizumab. In some embodiments, the first binding domain comprises the VH and / or VL amino acid sequences of hu31a.v11, and the second binding domain comprises the VH and / or VL amino acid sequences of the anti-IL-13 antibody lebrikizumab.

[0239] One of the aforementioned bispecific antitryptase / anti-IL-13 antibodies has (a) at least 80% sequence identity with SEQ ID NO: 9 (e.g., 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity), or a VH domain containing an amino acid sequence having that sequence, or (b) at least 80% sequence identity with SEQ ID NO: 10 It may include a VL domain containing an amino acid sequence having a % sequence identity (e.g., 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%), or a first binding domain that binds to a tryptase, containing a VH domain as in (a) and a VL domain as in (b). One of the aforementioned bispecific antitryptase / anti-IL-13 antibodies is (a) a VH domain containing an amino acid sequence having at least 80% sequence identity with SEQ ID NO: 90 or SEQ ID NO: 114 (e.g., 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%), or (b) SEQ ID NO: 91 or SEQ ID NO: It may include a VL domain containing an amino acid sequence having at least 80% sequence identity with 115 (e.g., 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity), or a second binding domain that binds to IL-13, containing a VH domain as in (c)(a) and a VL domain as in (b). In some examples, the second binding domain contains the VH and / or VL domains of leburikizumab.

[0240] In other embodiments, in some examples, the bispecific antibody comprises a first binding domain that binds to tryptase and a second binding domain that binds to IL-33. The second binding domain that binds to IL-33 may include, for example, one of the anti-IL-33 antibodies described in U.S. Patent Publication No. 2016 / 0168242, which is incorporated herein in whole by reference. In some embodiments, the first binding domain that binds to tryptase is, for example, (a) HVR-H1 comprising the amino acid sequence X1X2GMX3 (SEQ ID NO: 1) where X1 is Asp or Ser, X2 is Tyr or Phe, and X3 is Val or His; (b) HVR-H2 comprising the amino acid sequence FISSGSSTVYYADTMKG (SEQ ID NO: 2); (c) HVR-H3 comprising the amino acid sequence RX1X2X3DWYFDV (SEQ ID NO: 3) where X1 is Asn or Asp, X2 is Tyr or Asn, and X3 is Asp or Tyr; and (d) HVR-L1 comprising the amino acid sequence SASSSVTYMY (SEQ ID NO: 4). This may include, at least one, two, three, four, five, or six hypervariable regions (HVRs) selected from (e) HVR-L2 containing the amino acid sequence of RTSDLAS (SEQ ID NO: 5) and (f) HVR-L3 containing the amino acid sequence of QHYHSYPLT (SEQ ID NO: 6), or a combination of one or more of the above HVRs, and one or more variants thereof having at least about 80% sequence identity with any one of SEQ ID NOs. 1 to 6 (e.g., 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity).In some examples, the second binding domain that binds to IL-33 is, for example, (a) HVR-H1 containing the amino acid sequence of SFSMS (SEQ ID NO: 120), (b) HVR-H2 containing the amino acid sequence of TISGGKTFTDYVDSVKG (SEQ ID NO: 121), (c) HVR-H3 containing the amino acid sequence of ANYGNWFFEV (SEQ ID NO: 122), (d) HVR-L1 containing the amino acid sequence of RASESVKYGLSLLN (SEQ ID NO: 123), (e) HVR-L2 containing the amino acid sequence of AASNRGS (SEQ ID NO: 124), and (f) QQSKE The second binding domain may include at least one, two, three, four, five, or six HVRs selected from HVR-L3 containing the amino acid sequence of VPFT (SEQ ID NO: 125), or a combination of one or more of the above HVRs, and one or more variants thereof having at least about 80% sequence identity with any one of SEQ ID NOs. 120-125 (e.g., 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity). In some embodiments, the second binding domain includes one, two, three, four, five, or six HVRs from the anti-IL-33 antibody 10C12.38.H6.87Y.58I.

[0241] For example, in some cases, the first binding domain that binds to tryptase includes at least one, two, three, four, five, or six hypervariable regions (HVRs) selected from (a) HVR-H1 containing the amino acid sequence of DYGMV (SEQ ID NO: 7), (b) HVR-H2 containing the amino acid sequence of FISSGSSTVYYADTMKG (SEQ ID NO: 2), (c) HVR-H3 containing the amino acid sequence of RNYDDWYFDV (SEQ ID NO: 8), (d) HVR-L1 containing the amino acid sequence of SASSSVTYMY (SEQ ID NO: 4), (e) HVR-L2 containing the amino acid sequence of RTSDLAS (SEQ ID NO: 5), and (f) HVR-L3 containing the amino acid sequence of QHYHSYPLT (SEQ ID NO: 6). In some examples, the second binding domain that binds to IL-33 may include at least one, two, three, four, five, or six HVRs selected from, for example, (a) HVR-H1 containing the amino acid sequence of SFSMS (SEQ ID NO: 120), (b) HVR-H2 containing the amino acid sequence of TISGGKTFTDYVDSVKG (SEQ ID NO: 121), (c) HVR-H3 containing the amino acid sequence of ANYGNWFFEV (SEQ ID NO: 122), (d) HVR-L1 containing the amino acid sequence of RASESVAKYGLSLLN (SEQ ID NO: 123), (e) HVR-L2 containing the amino acid sequence of AASNRGS (SEQ ID NO: 124), and (f) HVR-L3 containing the amino acid sequence of QQSKEVPFT (SEQ ID NO: 125). In some embodiments, the second binding domain contains one, two, three, four, five, or six HVRs from the anti-IL-33 antibody 10C12.38.H6.87Y.58I. In some embodiments, the first binding domain contains the VH and / or VL amino acid sequence of hu31a.v11, and the second binding domain contains the VH and / or VL amino acid sequence of the anti-IL-33 antibody 10C12.38.H6.87Y.58I.

[0242] One of the aforementioned bispecific antitryptase / anti-IL-33 antibodies has (a) at least 80% sequence identity with SEQ ID NO: 9 (e.g., 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity), or a VH domain containing an amino acid sequence having that sequence, or (b) at least 80% sequence identity with SEQ ID NO: 10 It may include a VL domain containing an amino acid sequence having a % sequence identity (e.g., 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%), or a first binding domain that binds to a tryptase, containing a VH domain as in (a) and a VL domain as in (b). One of the aforementioned bispecific antitryptase / anti-IL-33 antibodies is (a) a VH domain containing an amino acid sequence having at least 80% sequence identity with SEQ ID NO: 126 (e.g., 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity), or (b) at least SEQ ID NO: 127 It may include a VL domain containing an amino acid sequence with 80% sequence identity (e.g., 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity), or a second binding domain that binds to IL-33 containing a VH domain as in (c)(a) and a VL domain as in (b). In some examples, the second binding domain contains the VH and / or VL domains of 10C12.38.H6.87Y.58I.

[0243] Techniques for producing multispecific antibodies include, but are not limited to, the recombinant co-expression of two immunoglobulin heavy-light chain pairs with different specificities (see Milstein et al. Nature 305:537, 1983, WO93 / 08829, and Traunecker et al. EMBOJ.10:3655, 1991) and the "knob-in-hole" operation (see, for example, U.S. Patent No. 5,731,168). Multispecific antibodies can also be produced by manipulating the electrostatic steering effect to create antibody Fc-heterodimer molecules (WO2009 / 089004A1), crosslinking two or more antibodies or fragments (see, e.g., U.S. Patent No. 4,676,980 and Brennan et al. Science, 229:81, 1985), producing bispecific antibodies using leucine zippers (see, e.g., Kostelny et al. J.Immunol., 148(5):1547-1553, 1992), producing bispecific antibody fragments using "diabody" technology (see, e.g., Hollinger et al. Proc.Natl.Acad.Sci.USA 90:6444-6448, 1993), and using single-stranded Fv(scFv) dimers (see, e.g., Gruber et al. They can be prepared by (see al. J. Immunol. 152:5368, 1994) and by preparing triplicate antibodies (for example, as described in Tutt et al. J. Immunol. 147:60, 1991).

[0244] Modified antibodies having three or more functional antigen-binding sites, including "octopus antibodies," are also included herein (see, for example, US2006 / 0025576A1).

[0245] The antibodies or fragments described herein also include "Dual Acting FAb" or "DAF" which contain an antigen-binding site that binds to both tryptase and another different antigen (see, for example, US2008 / 0069820).

[0246] Knobs-into-Holes The use of knob-in-hole as a method for producing multispecific antibodies is described, for example, in U.S. Patent No. 5,731,168, WO2009 / 089004, U.S.2009 / 0182127, U.S.2011 / 0287009, Marvin and Zhu, Acta Pharmacol. Sin. (2005) 26(6):649-658, and Kontermann (2005) Acta Pharmacol. Sin. 26:1-9. A brief, non-restrictive discussion is provided below.

[0247] A “protrusion” refers to at least one amino acid side chain that can stabilize a heteromultimer and thereby protrude from the interface of the first polypeptide, for example, to favor heteromultimerization over homomultimerization, and thus be positioned within a complementary cavity at the adjacent interface (i.e., the interface of the second polypeptide). Protrusions may be present within the original interface or may be introduced synthetically (e.g., by altering the nucleic acid encoding the interface). In some embodiments, the nucleic acid encoding the interface of the first polypeptide is altered to encode a protrusion. To achieve this, the nucleic acid encoding at least one “original” amino acid residue at the interface of the first polypeptide is replaced with a nucleic acid encoding at least one “imported” amino acid residue having a larger side chain volume than the original amino acid residue. It will be understood that there may be two or more original residues and corresponding imported residues. Side chain volumes of various amino residues are shown, for example, in Table 1 of US2011 / 0287009 or Table 1 of U.S. Patent No. 7,642,228.

[0248] In some embodiments, the introduced residue for ridge formation is a naturally occurring amino acid residue selected from arginine (R), phenylalanine (F), tyrosine (Y), and tryptophan (W). In some embodiments, the introduced residue is tryptophan or tyrosine. In some embodiments, the prototype residue for ridge formation has a small side-chain volume, such as alanine, asparagine, aspartic acid, glycine, serine, threonine, or valine. See, for example, U.S. Patent No. 7,642,228.

[0249] A “cavity” refers to at least one amino acid side chain that is recessed from the interface of the second polypeptide and thus accommodates a corresponding bulge on the interface of the adjacent first polypeptide. The cavity may exist within the original interface or may be introduced by synthesis (e.g., by altering the nucleic acid encoding the interface). In some embodiments, the nucleic acid encoding the interface of the second polypeptide is modified to encode a cavity. To achieve this, the nucleic acid encoding at least one “original” amino acid residue of the interface of the second polypeptide is replaced with DNA encoding at least one “imported” amino acid residue having a smaller side chain volume than the original amino acid residue. It will be understood that there may be two or more original residues and corresponding imported residues. In some embodiments, the imported residue for cavity formation is a native amino acid residue selected from alanine (A), serine (S), threonine (T), and valine (V). In some embodiments, the imported residue is serine, alanine, or threonine. In some embodiments, the prototype residue for cavity formation has a large side chain volume, such as tyrosine, arginine, phenylalanine, or tryptophan.

[0250] The ridges are "situable" within the cavity, meaning that the spatial position of the ridge and cavity at the interface of the first and second polypeptides, respectively, and the size of the ridge and cavity, are such that the ridge can be positioned within the cavity without significantly disrupting the normal association of the first and second polypeptides at the interface. Since ridges such as Tyr, Phe, and Trp typically do not extend perpendicular to the interface axis and do not have a favorable three-dimensional structure, the alignment of the ridge with the corresponding cavity may, in some cases, depend on modeling of the ridge / cavity pair based on the three-dimensional structure, such as obtained by X-ray crystallography or nuclear magnetic resonance (NMR). This can be achieved using techniques widely accepted in the art.

[0251] In some embodiments, the knob mutation within the IgG1 constant region is T366W. In some embodiments, the hole mutation within the IgG1 constant region includes one or more mutations selected from T366S, L368A, and Y407V. In some embodiments, the hole mutation within the IgG1 constant region includes T366S, L368A, and Y407V.

[0252] In some embodiments, the knob mutation within the IgG4 constant region is T366W. In some embodiments, the hole mutation within the IgG4 constant region includes one or more mutations selected from T366S, L368A, and Y407V. In some embodiments, the hole mutation within the IgG4 constant region includes T366S, L368A, and Y407V.

[0253] 7. Antibody variants In certain embodiments, amino acid sequence variants of antibodies provided herein are intended. For example, it may be desirable to improve the binding affinity and / or other biological properties of the antibody, such as inhibitory activity. Amino acid sequence variants of antibodies can be prepared by introducing appropriate modifications into the nucleotide sequence encoding the antibody or by peptide synthesis. Such modifications include, for example, deletions from and / or insertions into residues and / or substitutions of residues within the amino acid sequence of the antibody. Deletions, insertions, and substitutions can be arbitrarily combined to arrive at a final construct, provided that the final construct has the desired characteristics, such as antigen-binding ability.

[0254] a) Substitution, insertion, and deletion variants In certain embodiments, antibody variants having one or more amino acid substitutions are provided. Target sites for substitutional mutagenesis include HVR and FR. Conservative substitutions are shown in Table 1 under the heading "Preferred Substitutions." More substantial variations are provided in Table 1 under the heading "Exemplary Substitutions" and are further described below with reference to amino acid side chain classes. Amino acid substitutions are introduced into the target antibody, and the product can be screened for desired activity, such as retention / improvement of antigen binding, reduction of immunogenicity, or improvement of ADCC or CDC. Table 1 TIFF0007853934000002.tif171170

[0255] Amino acids can be grouped according to their general side-chain properties: (1) Hydrophobic: norleucine, Met, Ala, Val, Leu, Ile; (2) Neutral hydrophilic: Cys, Ser, Thr, Asn, Gln; (3) Acidic: Asp, Glu; (4) Basicity: His, Lys, Arg; (5) Residues that affect chain orientation: Gly, Pro; (6) Aromatic: Trp, Tyr, Phe.

[0256] Non-conservative substitution involves swapping one member of one of these classes with one of another.

[0257] Certain types of substitutional variants involve substituting one or more hypervariable region residues of a parent antibody (e.g., a humanized antibody or a human antibody). Generally, the resulting variant(s) selected for further testing have a modification (e.g., improvement) of certain biological properties (e.g., increased affinity, decreased immunogenicity) compared to the parent antibody, and / or substantially retain certain biological properties of the parent antibody. Exemplary substitutional variants are affinity-mature antibodies that can be readily generated using phage display-based affinity maturation techniques, such as the techniques described herein. Briefly, one or more HVR residues are mutated, the variant anti...

Claims

1. An isolated antibody that binds to human tryptase beta 1, or an antigen-binding fragment thereof, wherein the antibody has the following six hypervariable regions (HVRs): (a) HVR-H1 containing the amino acid sequence of GYAIT (SEQ ID NO: 30) (b) HVR-H2 containing the amino acid sequence of GISSAATTFYSSWAKS (SEQ ID NO: 31) (c) HVR-H3 containing the amino acid sequence of DPRGYGAALDRLDL (SEQ ID NO: 32) (d) HVR-L1 containing the amino acid sequence of QSIKSVYNNNRLG (SEQ ID NO: 33), (e) HVR-L2 containing the amino acid sequence of ETSILTS (SEQ ID NO: 34), (f) An isolated antibody that binds to human tryptase beta 1, or an antigen-binding fragment thereof, comprising HVR-L3 containing the amino acid sequence AGGFDRSGDTT (SEQ ID NO: 35).

2. The antibody according to claim 1, wherein the antibody comprises (a) a heavy chain variable (VH) domain having at least 90% sequence identity with the amino acid sequence of SEQ ID NO: 36, (b) a light chain variable (VL) domain having at least 90% sequence identity with the amino acid sequence of SEQ ID NO: 37, or (c) the VH domain described in (a) and the VL domain described in (b).

3. The antibody according to claim 1, wherein the VH domain comprises the amino acid sequence of SEQ ID NO:

36.

4. The antibody according to claim 1, wherein the VL domain comprises the amino acid sequence of SEQ ID NO:

37.

5. The antibody according to claim 1, wherein the antibody can inhibit the enzymatic activity of human tryptase beta 1.

6. The aforementioned antibody has a K content of approximately 180 pM. Ｄ The antibody according to claim 5, which binds to the tryptase.

7. The antibody according to claim 5, wherein the antibody inhibits tryptase activity with an IC50 of about 2.5 nM or less when determined by a human tryptase beta enzyme assay using a chromogenic synthetic peptide substrate.

8. (i) The antibody can inhibit the enzymatic activity of human tryptase beta 1 at pH 6; (ii) The antibody can inhibit tryptase-mediated stimulation of bronchial smooth muscle cell proliferation and / or collagenous contraction; (iii) The antibody can dissociate the tetrameric human tryptase beta-1; (iv) The antibody can dissociate tetrameric human tryptase beta-1 when it is in monovalent format; and / or (v) The antibody according to claim 5, wherein the antibody can dissociate tetrameric human tryptase beta 1 in the presence of a low concentration of heparin.

9. The antibody according to claim 1, wherein the antibody can dissociate the small interface of the tetrameric human tryptase beta 1.

10. The antibody according to claim 1, wherein the antibody further binds to cynomolgus tryptase, human tryptase alpha, human tryptase beta 2 and / or human tryptase beta 3.

11. The antibody according to claim 1, wherein the antibody is humanized.

12. The antibody according to claim 1, wherein the antibody is an IgG antibody.

13. The antibody according to claim 12, wherein the IgG antibody is an IgG1 antibody or an IgG4 antibody.

14. The antibody according to claim 13, wherein the IgG4 antibody contains an S228P mutation (according to EU numbering) in the heavy chain constant region.

15. The antibody according to claim 1, wherein the antibody is a monospecific antibody.

16. The antibody according to claim 1, wherein the antibody is a multispecific antibody.

17. The antibody according to claim 16, wherein the multispecific antibody is a bispecific antibody.

18. The antibody according to claim 17, wherein the antibody comprises a first binding domain that binds to human tryptase beta 1 and a second binding domain that binds to a second biological molecule, the second biological molecule being selected from the group consisting of interleukin-13 (IL-13), interleukin-4 (IL-4), interleukin-5 (IL-5), interleukin-17 (IL-17), IgE, and interleukin-33 (IL-33).

19. A pharmaceutical composition comprising the antibody described in claim 1, and a pharmaceutically acceptable carrier, excipient, or diluent.

20. The pharmaceutical composition according to claim 19, wherein the excipient is an antioxidant.

21. The pharmaceutical composition according to claim 20, wherein the excipient contains N-acetyltryptophan at a concentration of about 0.1 mM to about 1 mM and methionine at a concentration of about 1 mM to about 10 mM.

22. The pharmaceutical composition according to claim 20, contained in a light-shielding container or in a pre-filled syringe.

23. A pharmaceutical composition comprising the antibody according to claim 5, and a pharmaceutically acceptable carrier, excipient, or diluent.

24. (a) an isolated antibody comprising a heavy chain containing the amino acid sequence of SEQ ID NO: 80 and a light chain containing the amino acid sequence of SEQ ID NO: 81, or (b) a heavy chain containing the amino acid sequence of SEQ ID NO: 82 and a light chain containing the amino acid sequence of SEQ ID NO:

83.

25. A pharmaceutical composition comprising the antibody according to claim 24, and a pharmaceutically acceptable carrier, excipient, or diluent.

26. An isolated nucleic acid, or set of isolated nucleic acids, encoding an antibody that binds to human tryptase beta 1, or an antigen-binding fragment thereof, wherein the antibody has the following six hypervariable regions (HVRs): (a) HVR-H1 containing the amino acid sequence of GYAIT (SEQ ID NO: 30) (b) HVR-H2 containing the amino acid sequence of GISSAATTFYSSWAKS (SEQ ID NO: 31) (c) HVR-H3 containing the amino acid sequence of DPRGYGAALDRLDL (SEQ ID NO: 32) (d) HVR-L1 containing the amino acid sequence of QSIKSVYNNNRLG (SEQ ID NO: 33), (e) HVR-L2 containing the amino acid sequence of ETSILTS (SEQ ID NO: 34), (f) An isolated nucleic acid, or set of isolated nucleic acids, containing HVR-L3 containing the amino acid sequence AGGFDRSGDTT (SEQ ID NO: 35).

27. The isolated nucleic acid or set of isolated nucleic acids according to claim 26, wherein the antibody comprises (a) a VH domain having at least 90% sequence identity with the amino acid sequence of SEQ ID NO: 36, (b) a VL domain having at least 90% sequence identity with the amino acid sequence of SEQ ID NO: 37, or (c) the VH domain described in (a) and the VL domain described in (b).

28. The isolated nucleic acid or set of isolated nucleic acids according to claim 27, wherein the antibody comprises (a) a VH domain having at least 95% sequence identity with the amino acid sequence of SEQ ID NO: 36, (b) a VL domain having at least 95% sequence identity with the amino acid sequence of SEQ ID NO: 37, or (c) the VH domain described in (a) and the VL domain described in (b).

29. The isolated nucleic acid or set of isolated nucleic acids according to claim 28, wherein the antibody comprises (a) a VH domain having at least 99% sequence identity with the amino acid sequence of SEQ ID NO: 36, (b) a VL domain having at least 99% sequence identity with the amino acid sequence of SEQ ID NO: 37, or (c) the VH domain described in (a) and the VL domain described in (b).

30. The isolated nucleic acid or set of isolated nucleic acids according to claim 26, wherein the antibody comprises a VH domain containing the amino acid sequence of SEQ ID NO:

36.

31. The isolated nucleic acid or set of isolated nucleic acids according to claim 26, wherein the antibody comprises a VL domain containing the amino acid sequence of SEQ ID NO:

37.

32. The isolated nucleic acid or set of isolated nucleic acids according to claim 26, wherein the antibody can dissociate both the small interface and the large interface of the tetrameric human tryptase beta 1.

33. The isolated nucleic acid or set of isolated nucleic acids according to claim 26, wherein the antibody further binds to cynomolgus tryptase, human tryptase alpha, human tryptase beta 2 and / or human tryptase beta 3.

34. The aforementioned antibody has a K content of approximately 1 nM or less. Ｄ The isolated nucleic acid or set of isolated nucleic acids according to claim 26, which binds to the tryptase.

35. The aforementioned antibody has a K content of approximately 180 pM. Ｄ The isolated nucleic acid or set of isolated nucleic acids according to claim 34, which binds to the tryptase.

36. The isolated nucleic acid or set of isolated nucleic acids according to claim 26, wherein the antibody can inhibit the enzymatic activity of human tryptase beta 1.

37. The isolated nucleic acid or set of isolated nucleic acids according to claim 36, wherein the antibody inhibits tryptase activity with an IC50 of about 2.5 nM or less, as determined by a human tryptase beta enzyme assay using a chromogenic synthetic peptide substrate.

38. (i) The antibody can inhibit the enzymatic activity of human tryptase beta 1 at pH 6; (ii) The antibody can inhibit tryptase-mediated stimulation of bronchial smooth muscle cell proliferation and / or collagenous contraction; (iii) The antibody can dissociate the tetrameric human tryptase beta-1; (iv) The antibody can dissociate tetrameric human tryptase beta-1 when it is in monovalent format; and / or (v) The isolated nucleic acid or set of isolated nucleic acids according to claim 26, wherein the antibody can dissociate tetrameric human tryptase beta 1 in the presence of a low concentration of heparin.

39. The isolated nucleic acid or set of isolated nucleic acids according to claim 26, wherein the antibody is a monoclonal antibody or is humanized.

40. The isolated nucleic acid or set of isolated nucleic acids according to claim 26, wherein the antibody is an IgG antibody.

41. The isolated nucleic acid or set of isolated nucleic acids according to claim 40, wherein the IgG antibody is an IgG1 antibody or an IgG4 antibody.

42. The isolated nucleic acid or set of isolated nucleic acids according to claim 41, wherein the IgG4 antibody contains an S228P mutation (according to EU numbering) in the heavy chain constant region.

43. The isolated nucleic acid or set of isolated nucleic acids according to claim 29, wherein the antibody is a monospecific antibody or a multispecific antibody.

44. The isolated nucleic acid or set of isolated nucleic acids according to claim 43, wherein the multispecific antibody is a bispecific antibody.

45. The isolated nucleic acid or set of isolated nucleic acids according to claim 44, wherein the antibody comprises a first binding domain that binds to human tryptase beta 1 and a second binding domain that binds to a second biological molecule, and the second biological molecule is selected from the group consisting of interleukin-13 (IL-13), interleukin-4 (IL-4), interleukin-5 (IL-5), interleukin-17 (IL-17), IgE, and interleukin-33 (IL-33).

46. An isolated nucleic acid, or a set of isolated nucleic acids, encoding an antibody that binds to human tryptase beta 1, or an antigen-binding fragment thereof, wherein the antibody comprises (a) a VH domain comprising the amino acid sequence of SEQ ID NO: 36 and (b) a VL domain comprising the amino acid sequence of SEQ ID NO: 37, and the nucleic acid or set of nucleic acids comprises a sequence having at least 90% sequence identity with the sequence of SEQ ID NO: 109 and / or SEQ ID NO:

110.

47. An isolated nucleic acid, or set of isolated nucleic acids, according to claim 46, comprising a sequence having at least 90% sequence identity with sequence number 109 and / or sequence number 110.

48. An isolated nucleic acid, or set of isolated nucleic acids, according to claim 47, comprising a sequence having at least 95% sequence identity with sequence number 109 and / or sequence number 110.

49. An isolated nucleic acid, or set of isolated nucleic acids, according to claim 48, comprising a sequence having at least 99% sequence identity with sequence number 109 and / or sequence number 110.

50. An isolated nucleic acid, or set of isolated nucleic acids, according to claim 46, comprising the sequence of sequence number 109 and / or sequence number 110.

51. The isolated nucleic acid or set of isolated nucleic acids according to claim 46, wherein the antibody comprises (a) a heavy chain containing the amino acid sequence of SEQ ID NO: 80 and / or (b) a light chain containing the amino acid sequence of SEQ ID NO: 81, and the nucleic acid or set of nucleic acids comprises a sequence having at least 90% sequence identity with the sequence of SEQ ID NO: 111 and / or SEQ ID NO:

112.

52. An isolated nucleic acid, or set of isolated nucleic acids, according to claim 51, comprising a sequence having at least 90% sequence identity with sequence number 111 and / or sequence number 112.

53. An isolated nucleic acid, or set of isolated nucleic acids, according to claim 52, comprising a sequence having at least 95% sequence identity with sequence number 111 and / or sequence number 112.

54. An isolated nucleic acid, or set of isolated nucleic acids, according to claim 53, comprising a sequence having at least 99% sequence identity with sequence number 111 and / or sequence number 112.

55. An isolated nucleic acid, or set of isolated nucleic acids, according to claim 54, comprising the sequence of sequence number 111 and / or sequence number 112.

56. The isolated nucleic acid or set of isolated nucleic acids according to claim 46, wherein the antibody comprises (a) a heavy chain containing the amino acid sequence of SEQ ID NO: 82 and / or (b) a light chain containing the amino acid sequence of SEQ ID NO: 83, and the nucleic acid or set of nucleic acids comprises a sequence having at least 90% sequence identity with the sequence of SEQ ID NO: 113 and / or SEQ ID NO:

112.

57. An isolated nucleic acid, or set of isolated nucleic acids, according to claim 56, comprising a sequence having at least 90% sequence identity with sequence number 113 and / or sequence number 112.

58. An isolated nucleic acid, or set of isolated nucleic acids, according to claim 57, comprising a sequence having at least 95% sequence identity with sequence number 113 and / or sequence number 112.

59. An isolated nucleic acid, or set of isolated nucleic acids, according to claim 58, comprising a sequence having at least 99% sequence identity with sequence number 113 and / or sequence number 112.

60. An isolated nucleic acid, or set of isolated nucleic acids, according to claim 59, comprising the sequence of sequence number 113 and / or sequence number 112.

61. An isolated nucleic acid, or a set of isolated nucleic acids, encoding an antibody that binds to human tryptase beta 1, or an antigen-binding fragment thereof, wherein the antibody comprises a VH domain containing the amino acid sequence of SEQ ID NO: 36 and a VL domain containing the amino acid sequence of SEQ ID NO:

37.

62. (a) an antibody comprising a heavy chain containing the amino acid sequence of SEQ ID NO: 80 and a light chain containing the amino acid sequence of SEQ ID NO: 81, or (b) an antibody comprising a heavy chain containing the amino acid sequence of SEQ ID NO: 82 and a light chain containing the amino acid sequence of SEQ ID NO: 83, an isolated nucleic acid, or a set of isolated nucleic acids.

63. A vector or set of vectors comprising an isolated nucleic acid or a set of isolated nucleic acids as described in claim 26.

64. An isolated host cell comprising the vector or set of vectors according to claim 63.

65. The isolated host cell according to claim 64, wherein the host cell is a mammalian cell or a prokaryotic cell.

66. A method for producing an antibody that binds to human tryptase beta 1, comprising culturing the isolated host cells described in claim 65 in a culture medium under suitable conditions that enable the production of the antibody.

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